- Email: [email protected]
Procedures related to connective tissue disease
Best Practice & Research Clinical Rheumatology Vol. 19, No. 3, pp. 417–436, 2005 doi:10.1016/j.berh.2005.01.008 available online at http://www.sciencedirect.com
5 Procedures related to connective tissue disease Aike A. Kruize*
MD, PhD
Rheumatologist
Johannes W.J. Bijlsma
MD, PhD
Professor and Head Department of Rheumatology and Clinical Immunology, F02.127, University Medical Center, Box 85500, 3508 GA Utrecht, The Netherlands
In patients with rheumatic and, especially, connective tissue diseases many organ systems apart from the joints may be involved, such as the skin, muscles, salivary glands, nerves, kidneys and blood vessels. Biopsies of these tissues may help in establishing a diagnosis, in assessing the extent and severity of the disease and in monitoring the success of therapy. In this chapter, indications for biopsy, technical procedures, potential results and possible complications are described. The emphasis is on when a biopsy is indicated, how the patient is prepared, how the biopsy is performed, what results can be expected from the biopsy and what are the possible sequelae. For full details of the histology of e.g. a renal biopsy in a lupus patient, the reader is referred to the relevant textbooks. Key words: skin biopsy; subcutaneous fat biopsy; salivary gland biopsy; muscle biopsy; peripheral nerve biopsy; renal biopsy; temporal artery biopsy.
BIOPSIES OF THE SKIN Indications Abnormalities of the skin may provide clues for the diagnosis of a generalised rheumatic disease. To diagnose suspected autoimmune dermatose, skin biopsy material for histological examination, including immunofluorescence, may be of great help. Examples are blistering disorders (perilesional skin) or lupus erythematosus (lesional skin). In addition, skin biopsy is used to diagnose a dermal or epidermal neoplasm or an * Corresponding author. Tel.: C31 3025 07357; Fax: C31 3025 23741. E-mail address: [email protected] (A.A. Kruize). 1521-6942/$ - see front matter Q 2005 Elsevier Ltd. All rights reserved.
418 A. A. Kruize and J. W. J. Bijlsma
infection, by culturing a fresh, unfixed skin biopsy specimen for various organisms including mycobacteria, deep fungi or by examining it for protozoa or filarial worms.1 Technical procedures Skin biopsy is essentially a minimally invasive procedure. A preferably untreated skin lesion, in an early stage and characteristic for the skin disorder as a whole, should be chosen for biopsy. If lesions are present at different stages of evolution, it may be helpful to take biopsies of different lesions. For blistering, ulcerating or necrotic disorders, the biopsy should include both normal and affected skin so that the pathologist can identify the plane of separation for a blister or the early changes adjacent to ulceration and necrosis. For cases of suspected vasculitis the location of the affected vessels must be considered: palpable purpura occur in superficial venules in the dermis while vascular changes in disorders producing necrosis or livedo are in larger vessels deep in the subcutaneous fat. When direct immuno-fluorescence is needed, the biopsy should also include normal-looking skin. In choosing the site of biopsy, one should try to avoid areas that are prone to bad healing and areas that are cosmetically important. Moreover, changes that are secondary to circumstances such as venous stasis of the lower legs in the elderly may interfere with the histopathological abnormalities associated with the lesion and therefore be a reason for avoiding such sites. The skin is anaesthetised by infiltrating 1–2% lidocaine with or without epinephrine around the site of biopsy. Because epinephrine, more than lidocaine, may cause distortion artifacts which interfere with the histological interpretation of the biopsy specimen, some caution with the dosage is advised. It is probable that some of the discussion on the negative effects of adrenaline has been exaggerated. When taking a biopsy of the extremities, however, incidental extreme vasospasm might cause necrosis of the tissue. Slow delivery may decrease the pain associated with the installation of the anaesthesia. Superficial injection, creating an oedematous wheal has immediate efficacy but is more painful. Several techniques for skin biopsy are available, depending on the purpose of the subsequent investigation. For diagnosing a dermatose possibly associated with a generalised rheumatic disease, punch biopsy, incision biopsy and excision biopsy are the most commonly used techniques.2 A punch biopsy samples both the epidermis and dermis; in most cases a 3–4 mm punch is adequate. A disposable punch is generally used, having a cutting edge attached to a handle. A small cylinder of tissue is removed by means of a twisting movement of the punch. The specimen is extracted using forceps but scissors may be needed to detach the base of the tissue sample. The wound can be left to heal by secondary intention after achieving haemostasis; the scar may take several weeks to heal. The wound can be closed primarily and the resulting scar is generally a white or hypo-pigmented linear mark. Although incision (‘wedge’) biopsy should ideally reveal material from a sample of approximately 5 mm in length, cosmetic reasons, however, may necessitate some modesty in taking material. When taking a biopsy of a larger lesion, obtaining adjacent normal skin is important. For cosmetic reasons, the main line of incision should try to follow the natural crease lines of the skin. Lesions O4 mm may have to be stitched. Small lesions may be removed totally and, if malignancy is suspected, it is important to perform an excision biopsy with clear margins. Mechanical damage of the tissue should be avoided. When different fixatives are needed for different studies, it is preferable to take two specimens rather than dividing
Procedures related to connective tissue disease 419
one specimen into smaller parts. For routine microscopy of material embedded in paraffin, 10% formalin is most widely used. For specific techniques, including immunofluorescence, more specific transport media and fixatives are available. Potential results The most commonly encountered skin lesions in patients with generalised rheumatic disease are exanthema, nodules, vesicles and bullae, ulcers, purpura and papulosqamous rashes. As in rheumatic diseases, the diagnosis of skin lesions is commonly based upon pattern recognition. Not all skin lesions warrant skin biopsy. If in doubt, histological examination may confirm a probable diagnosis, because the histological features of several skin lesions are diagnostic, such as palisading granulomas in rheumatoid nodules. Biopsy of a suspicious lesion can confirm the diagnosis of lupus in a patient with other suggestive clinical manifestations.3 Direct immunofluorescence is especially diagnostic in lupus erythematosus and blistering abnormalities. In over 70% of patients with lupus erythematosus the lupus band test, i.e. granular deposition of immunoglobulins at the dermo-epidermal junction, is present, particularly in sun exposed skin, while it is absent in normal skin. Scleroderma involves the skin and subcutaneous structures in a characteristic manner, although similar findings can occur as a localised process (e.g. morphea) or as deeper pathology, such as in the fasciitis syndromes. In scleroderma there is a risk of impaired healing of the biopsy wound, therefore care should be taken in selecting the biopsy location. Vasculitis seen on skin biopsy often prompts a search for systemic involvement and seldom stands alone as a diagnostic finding. Immunoglobulin A deposition indicates Henoch-Scho¨nlein purpura or one of its variants. Not all palpable purpurae are due to vasculitis; biopsy can show other processes, such as microthrombi or cholesterol emboli.2 When panniculitis is suspected, the biopsy should be taken deeply enough to contain subcutaneous fat. When subcutaneous nodules are found in polyarthritis they are often characteristic for rheumatoid granuloma; however, nodular vasculitis and nodules of other material, such as monosodium urate, calcium pyrophosphate, cholesterol or amyloid may occur. Complications Complications during invasive procedures All invasive procedures may lead to the following complications: † Post-biopsy bleeding, especially when aspirin, non-steroidal anti-inflammatory drugs (NSAIDs) or anticoagulants are used. † To completely prevent this bleeding medication should be stopped well before hand. † Infections: all invasive examinations can become a ‘point of entry’ for infections. The presence of cardiac valvular disease raises this risk for infection, especially subacute bacterial endocarditis; prophylactic treatment with antibiotics might be indicated.
420 A. A. Kruize and J. W. J. Bijlsma
If preoperative antibiotics are administered, the dosage needs to be adequate for the control of skin flora, especially Staphylococcus aureus; they should be started less than 2 hours pre-operatively and continued for 48–72 hours. Skin biopsy of the upper trunk has an increased risk for keloid formation in patients who tend to make keloid. Skin biopsy of the lower leg of an elderly patient may result in poor healing and be prone to infection. If an infection develops after 3–5 days, oral antibiotics may be required. The slightest suspicion of malignant melanoma should result in refraining from skin biopsy in favour of a wide excision biopsy because of the (theoretical) risk of disseminating tumor cells.
BIOPSY OF SUBCUTANEOUS FAT Indications Systemic amyloidoses are characterised by the extracellular deposition of fibrils, derived from circulating soluble precursors. In the past, amyloidosis A (AA, reactive or secondary) was quite often found in patients with longstanding rheumatoid arthritis (RA) and it was found sporadically in patients with other chronic inflammatory arthropathies.4 Perhaps due to the improvement in the treatment of most rheumatic diseases over the last decade, AA is now less often present in these patients. Patients with other forms of amyloidosis, such as AL (amyloidosis immunoglobulin light chain, or primary) and Ab2-microglobulin, dialysis associated, amyloidosis may have rheumatic symptoms such as arthritis (with a predilection for the shoulder). To diagnose amyloidosis, the definitive test is biopsy of easily accessible tissues that are expected to contain amyloid, or biopsy of clinically affected organs. Rectal biopsy and subcutaneous fat aspiration are the procedures of choice. Technical procedures Fat samples are taken from the abdominal wall between the xiphoid process and the pubic bone, using an adapter, disposable syringe (10 ml) and a disposable needle (Gauge 0.8 or 0.9). The needle is injected quickly into the subcutaneous fat, a vacuum drawn into the syringe and the needle is then moved in different directions back and forth very quickly. The vacuum is released before the needle is withdrawn from the fat. Fat tissue remaining in the needle is blown with air onto glass slides, where it is allowed to air overnight before being stained with Congo red.5 Potential results In AA patients, subcutaneous fat aspiration has been reported to have a very high sensitivity of 0.97 while in AL patients this was reported to be 0.62. However, these data come from tertiary referral centres and a significantly lower yield for this procedure has been reported in other settings. Because each form of amyloid is defined by the chemical nature of the fibril protein, pathological specimens from any site can be stained with antibodies that are specific for the precursors to establish the character of the responsible disease.
Procedures related to connective tissue disease 421
Complications In theory bleeding and infection may occur, as listed previously. However, no relevant complications have been reported.
SUBLABIAL SALIVARY GLAND BIOPSY Indications In rheumatology, confirmation of a presumed diagnosis of Sjo¨gren’s syndrome is the main indication for sublabial salivary gland biopsy. To assess the oral component of Sjo¨gren’s syndrome various techniques are used, including sialochemistry, nuclear imaging of the salivary glands and radiographical contrast sialography. Focal sialadenitis in sublabial salivary gland specimens reflects salivary gland involvement in Sjo¨gren’s syndrome as a whole, allowing for a relatively simple diagnostic procedure compared to major salivary gland biopsy. Nowadays, sublabial salivary gland biopsy is widely considered and used as the single most specific test for confirming a diagnosis of (primary) Sjo¨gren’s syndrome. Histological changes in salivary gland tissue, however, are not 100% specific. Consequently, focal sialadenitis is not pathognomic for Sjo¨gren’s syndrome: focal sialadenitis is also found in sublabial minor salivary gland specimens overlain by inflamed mucosa due to lichen planus, cheek biting or a mucocele. Consequently, sublabial salivary gland biopsy that is intended for the confirmation of a diagnosis of Sjo¨gren’s syndrome should be taken through healthy, normal appearing mucosa of the lower lip. However, in contrast to acinar atrophy, dilatation of ducts and hyperplasia of sublabial salivary gland tissue— which all increase with age and are related to inflammation—focal sialadenitis is an objective diagnostic parameter for a diagnosis of the salivary component of Sjo¨gren’s syndrome. Other diagnostic findings in sublabial salivary gland specimens include non-caseating granuloma indicating sarcoidosis, amyloidosis, iron depositions indicating haemochromatosis and abnormal appearing lymphocytes compatible with leukaemia. Technical procedures Labial salivary gland biopsy is a simple outpatient procedure. The patient is asked to lie down on his back with the upper half of the body inclined upright at 308. A brief summary explaining the procedure is given. The mucosa of the lower lip is locally infiltrated with anesthetic, e.g. 2% lidocaine in addition to vasoconstrictor epinephrine 1:80 000. A 1 cm superficial linear incision of the epithelium of the mucosa is made between the midline and commissure, parallel to the vermillion, revealing the minor salivary glands (Figure 1). The glands are bluntly dissected from the fascia and excised separately, avoiding damaging the sensory nerves, which can be clearly distinguished. An assistant should be dabbing regularly to enable an adequate view. Because histopathological findings may differ between the various glands, at least five glands have to be taken for examination (Figure 2). The incision is closed with dissolvable sutures (Figure 3). In order to avoid burning due to loss of sensation, the patient is advised not to drink hot beverages for the next few hours.
422 A. A. Kruize and J. W. J. Bijlsma
Figure 1. Lipbiopsy 1: paramedian incision of the lower lip.
The glands are fixed in a formol (-sublimate) solution, before embedding them in paraffin for histopathological examination. Inflammation, expressed as focal lymphocytic sialoadenits, is assessed semi-quantitatively. Salivary gland lobes with dilatation of ducts, parenchymal atrophy or extravascular polymorphonuclear leukocytes are excluded from the assessment. Potential results A focus of focal lymphocytic adenitis is defined as an aggregate of at least 50 lymphocytes and histocytes. Usually these foci are found at the edge of intralobular
Figure 2. Lipbiopsy 2: sublabial salivary gland biopsy specimen.
Procedures related to connective tissue disease 423
Figure 3. Lipbiopsy 3: stitched incision of the lower lip.
ducts, related to small veins, adjacent to the acini of the glands, leading to destruction of the acini while relatively sparing the ducts. More than one focus per 4 mm2 sample of sublabial salivary gland tissue—corresponding to a focal sialadenitis of grade IV6—is a consistent finding and, subsequently, a widely accepted criterion for the histopathological confirmation of Sjo¨gren’s syndrome. It is one of the main objective criteria items of the revised classification criteria proposed by the American–European Consensus group.7 The presence of duct abnormalities and the formation of epimyoepithelial cell islands in major salivary gland tissue are late features in Sjo¨gren’s syndrome, but are generally not seen in minor gland tissue. In addition, immunohistological assessment of labial salivary gland specimens is reported to be more specific for the disease than the lymphocytic focus score criterion. A bivariate classification criterion, based on the percentages of IgA- and IgG-containing plasma cells resulted in a reduction in the number of false positive diagnoses. Using the more simply manageable univariate criterion based on the percentage of IgA-containing plasma cells (!70% IgA-containing plasma cells for confirmation of Sjo¨gren’s syndrome), results in 99 and 96% specificity and sensitivity, respectively.8 Also, immunohistological detection of monotypic plasma cells in labial salivary glands—defined by a kappa:lambda ratio of R3—has been indicated as a prognosticator for systemic lymphoproliferative disease. Complications Complications of sublabial salivary gland biopsy are, apart from the complications listed previously , seldom reported, but they include arterial bleeding due to inadequate, c.q. too deep incision, vasovagal collapse and burning of the mouth while drinking hot beverages due to a temporary loss of sensation because of the anaesthesia. Persistent hypoesthaesia of the lower lip region is reported when a nerve, in spite of the procedure of which all details can be followed by sight, is damaged.
424 A. A. Kruize and J. W. J. Bijlsma
MUSCLE BIOPSY Indications Muscle biopsy is the most crucial test for establishing the diagnosis of inflammatory myopathies.9 The inflammatory myopathies encompass a heterogeneous group of acquired muscle diseases characterised clinically by subacute-onset, proximal and symmetrical muscle weakness and histologically by inflammatory infiltrates within the skeletal muscles. These myopathies comprise three major and discrete subsets: polymyositis (PM), dermatomyositis (DM) and inclusion body myositis (IBM). Each subset retains its characteristic clinical, immunopathological and morphological features regardless of whether it occurs in isolation or in connection with other systemic diseases. Although the diagnosis of these disorders is based on the combination of clinical examination, serum muscle enzyme levels and sometimes the presence of autoantibodies, muscle biopsy offers the most definitive diagnostic information in the majority of cases.10 Although recently discussion was started as to whether or not PM is an over diagnosed entity,11 the value of muscle biopsy is still undisputed. Although there is a long list of differential diagnoses for muscle weakness, such as denervating conditions, neuromuscular junction disorders, muscular dystrophies, myotonic diseases, congenital myopathies, glycogen storage disease, lipid storage myopathies and others, clinical characteristics most often make these diagnoses less likely. Nevertheless, where there is doubt, muscle biopsy will help to differentiate between these different conditions. There is seldom an indication for a repeated muscle biopsy. When the first biopsy is negative, but there is a very strong clinical suspicion, such as for IBM, a second biopsy might be indicated, in which case it is best guided by skeletal muscle computed tomography (CT) or magnetic resonance imaging (MRI) scans. In cases where muscle weakness persists or is aggravated during glucocorticoid treatment a muscle biopsy might help to differentiate a possible glucocorticoid myopathy from undertreatment or relapse of the PM. A quite different indication for performing a muscle biopsy is established when vasculitis, especially polyarteritis nodosa (PAN), is suspected in a patient. In this situation a deep and rather large muscle biopsy is taken from a distal muscle group. Technical procedure Most researchers prefer an open muscle biopsy over a needle biopsy, because the former offers a larger sample that can be properly orientated and is better suited for immuno-cytochemical and ultra-structural studies. In some centres, however, percutaneous needle biopsy is used. A moderately weak muscle is the most suitable for performing a diagnostic biopsy because it offers the best chance to obtain maximum information. If the biopsy is done to seek vasculitis, a distal muscle group should be selected. Open biopsy is performed after infiltrating a local anaesthetic into the skin and subcutaneous tissue in the region of interest, avoiding muscle. Pain fibres are located in the perimysial area and fascia, but not the muscle itself. Hence, pain from the procedure can be minimised. In addition, sedation using a short-acting benzodiazepine or narcotic may further reduce the patient’s discomfort. Open biopsy permits the direct inspection
Procedures related to connective tissue disease 425
of the muscle, enabling an easily processed specimen to be obtained, which is more likely to contain larger blood vessels and no fat.12 A longitudinal incision of 5 cm through subcutaneous tissue and fat to reach the muscle should permit the excision of a sample measuring approximately 1!1!1 cm. The two ends of the sample should be lightly affixed to an object that holds the muscle at resting length. The sample should be kept moist before being placed in fixative to avoid contraction artifact; it should be deep frozen within 2 hours. A second, smaller specimen can be taken for electron microscopy. Percutaneous sampling of muscle is performed using specially designed instruments, such as a trocar with a cutting window that admits tissue by suction, or a springactivated biopsy instrument. Skin overlying an area that has been anaesthetised (as is done for open biopsy), is incised with a scalpel to enable penetration of the deeper tissues by the biopsy instrument. The cutting window of the biopsy instrument is opened to admit tissue, which is cut free and contained by the circling blade. Both percutaneous techniques can be performed at the bedside and are relatively atraumatic. The small size of the samples obtained requires special care in processing. An advantage of percutaneous biopsy is that muscle can be sampled from a wide area by orientating the biopsy instrument in several different directions through the same skin puncture, thus encompassing a much larger region than is available in an open biopsy.13 For transportation, muscle tissue may be kept on saline moistened gauze for several hours; the specimen should be kept cool. A portion of the biopsy specimen should be snap frozen for cryostat sections and embedded in plastic for ultra thin sections and ultra structural studies. Frozen muscle may be safely shipped overnight on dry ice. When muscle is frozen it can be stored at K80 8C. Frozen muscle for histochemistry provides excellent muscle fibre morphology and most diagnostic information using light microscopy. Muscle embedded in paraffin is useful for surveys for inflammation and morphology of inflammatory cells but gives poor muscle fibre morphology. It is always important to check the techniques to be used with the laboratory that is going to analyse the muscle biopsy. It is also advised to use a specific laboratory with expertise in enzyme histochemistry and myopathology. Some people advise the use of CT or MRI scans to select an involved region of a muscle. However, the additional value is not proven and most physicians do not use it for their primary biopsy; only when the biopsy results are negative while the clinical suspicion is very high, is a second biopsy guided by CT or MRI advisable. Potential results In cases of inflammatory myositis, histochemistry, immuno-histology and, sporadically, electron microscopy may help to establish a diagnosis. For details see relevant textbooks cited in Dalakas 2002.10 In idiopathic PM mononuclear cell infiltrates, predominantly consisting of T cells, are found in the endomysial spaces as well as invasion of histologically healthy looking muscle fibres by mononuclear cytotoxic T cells. Less pronounced are necrosis and regeneration of muscle fibres, while macrophages and B cells are sparse. It has been shown that in the clinically evident PM patient, without skin abnormalities, mononuclear cell infiltrates are often localised perimysially and perivascularly.11 In cases of sporadic IBM there are endomysial mononuclear cell infiltrates with invasion of normal-looking muscle fibres and so-called rimmed vacuoles in at least 0.3% of the muscle fibres. In DM the inflammation is predominantly perivascular, in the interfascicular septae rather than within the fascicles;
426 A. A. Kruize and J. W. J. Bijlsma
intramuscular blood vessels show endothelial hyperplasia and the muscle fibres undergo necrosis and phagocytosis. Ischaemia results in atrophy of fibres in the periphery of the fascicles (perifascicular atrophy). Additional biopsy stains are useful when other diagnoses are suspected, e.g. ATPase staining to define myosin loss, elucidating either type-1 or type-2 fibre atrophy, while different enzyme stains are used to elucidate mitochondrial pathology. Enzyme deficiencies and storage diseases can become evident by staining with periodic-acidSchiff (PAS: glycogen, carbohydrate), Alcian blood (mucopolysaccharide), Sudan black, Oil red (lipids) or Congo red (amyloid). Complications There are no serious complications for either open or needle biopsy, as aforesaid. Of course bleeding is a well-known possibility and medication that influences haemostases should be stopped before the procedure. When the biopsy is performed under properly aseptic conditions the possibility for an infection is very low. Sometimes the scar does not heal properly or a dermal nerve branch has been damaged. After biopsy, patients are advised to rest for a few hours, especially after an open biopsy. When an open biopsy of the leg has been performed, the patient is advised not to overuse that leg for a few days.
SURAL NERVE BIOPSY Indications In patients with rheumatic conditions the most frequent reason to perform a nerve biopsy is suspicion of a vasculitis of the nervus suralis. This vasculitis could be part of a systemic vasculitis or accompany a connective tissue disease such as RA. In general, nerve biopsy may be performed to help distinguish between demyelination (destruction of parts of the myelin sheath covering the nerve) and axonal degeneration (destruction of the axon portion of nerve cell), to identify inflammatory neuropathies, or to confirm a specific diagnosis. Peripheral neuropathy is very common; the incidence varies with the specific type of neuropathy. Most often it is a manifestation of conditions that can damage peripheral nerves, such as diabetes mellitus, renal insufficiency or alcoholism. Symptoms differ depending on whether the damage is to the sensory fibres, the motor fibres or both. Technical procedure There is no specific preparation for the patient. A local anaesthetic is used to numb the area. A small incision is made and a section is removed of the whole nerve (not limited to only a few fascicles, since in vasculitis not all fascicles may be involved). Because of the use of the local anaesthetic, discomfort during the procedure is usually minimal. The anaesthetic may burn or sting when first injected. After the procedure the area may feel tender or sore for a few days. The nerve tissue is frozen, paraffin embedded and sectioned in plastic. Morphological changes are studied using a light microscope, but on specific indications an electron microscopical investigation is performed.
Procedures related to connective tissue disease 427
Potential results With regard to specific rheumatic conditions, interest has focused on signs of inflammation, especially cellular infiltrates. Invasion of neutrophils into the vessel walls are suggestive for nercotising vasculitis. Eosinophils can also be seen in vasculitis, especially Churg-Strauss syndrome, but also in fasciitis and eosinophilia myalgia syndrome. Invasion of (epithelioid) histiocytes is seen in sarcoidosis, leprosy and also fasciitis. Plasma cell infiltrations in plasma cell dyscrasia can be seen in vasculitis, in Lyme’s disease and in leprosy. Perineural cell infiltrates are found in perineuritis and leprosy. In addition, information is gathered from nerve biopsies regarding demyelination, differential fascicular loss, neurofilament accumulation, mitochondrial abnormalities, axon loss, regeneration of axons and many other signs. In evaluating some of these differential diagnoses electro-physiological studies are also performed. In a case of suspected vasculitis it is important to select an appropriate nerve for biopsy, to also look at the size of the vessels involved and to look for the integrity of the lamina elastica of the vessel. Immune-mediated neuropathies also include Guillain-Barre´ syndrome (demyelinating disease) and chronic idiopathic axonal polyneuropathy associated with gammopathy. When the diagnosis of mononeuritis multiplex is made, a full examination of the patient is performed. In a case of a wellknown connective tissue disease, such as RA, the association is clear and in general more intensive therapy is indicated. However, if the mononeuritis multiplex is the first sign of a connective tissue disease then an intensive evaluation of the patient is warranted. Complications Removal of a part of the sural nerve leads to nerve damage, with hypaesthesia of the lateral site of the foot. Around 10% of the patients involved report persistent pain in this area. It is uncommon that this nerve damage leads to functional impairment. Of course the procedure will give some discomfort to the patient and there is a small risk of infection and bleeding, see table with listed complications given previously. In some cases an allergic reaction to the local anaesthetic has been reported.
BIOPSIES OF THE KIDNEY Indications In rheumatology, the main indication for biopsy of the kidney will be renal disease in association with an already defined generalised autoimmune disease, especially an earlier, established, diagnosis of systemic lupus erythematosus (SLE). In SLE, during the course of the disease, repeated kidney biopsy may be required. Renal findings in SLE do not correspond with the various subtypes of glomerulonephritis, therefore, renal histology is essential for determining the subtype of SLE nephritis and, consequently for establishing the ‘renal’ prognosis and required treatment. In patients with SLE regular evaluation of urine to identify abnormalities, such as red cells, proteinuria or casts is essential, since an ‘active urine’ may precede other signs of kidney involvement by many months. Also, unexplained persistent
428 A. A. Kruize and J. W. J. Bijlsma
proteinuria and/or haematuria, acute or chronic renal failure, persistent nephritic syndrome and acute nephritic syndrome in any patient with a rheumatic disease including RA, ankylosing spondylitis and reactive arthritis also form an indication for kidney biopsy. A clinical presentation of rapidly progressive glomerulonephritis (RPGN), which needs treatment without delay, requires urgent kidney biopsy. This may occur within the context of an already established rheumatic disease such as SLE, antineutrophil cytoplasmatic antibody (ANCA)-related vasculitis, including Wegener’s granulomatosis, and relapsing polychondritis. However, RPGN may also be part of the initiative phase of a generalised autoimmune or other disease, such as bacterial endocarditis and Goodpasture’s syndrome, requiring completely different treatments. Technical procedures The standard procedure for obtaining renal tissue is a percutaneous needle biopsy (Figures 4–10). Ultrasound is used to position the needle correctly. Radioscopic and CT positioning guidance are also used. In cases of extreme adiposity or repeated failure of percutaneous needle biopsy, alternative procedures such as surgical or laparoscopic biopsy may be required. Before percutaneous needle biopsy the patient is examined for bleeding diathesis (bleeding time, APTT (activated partial thromboplastin time) and PTT (prothrombin time)). NSAIDs are stopped 1 week before the bleeding time is checked. In cases of thrombopathy, prophylactic use of vasopressin may be indicated. Using ultrasound, the presence and position of two kidneys and the absence of congenital abnormalities, such as a horseshoe kidney, is checked. In cases where vasculitis is suspected, especially polyarteritis nodosa, angiographical examination of the kidney before the biopsy is performed is mandatory. If asked for, the patient receives 10 mg of oral diazepam, 60 minutes prior to the procedure, for sedation and muscle relaxation. The patient is asked to lie facedown and, subsequently, the skin is anaesthetised. For a standard
Figure 4. Kidney biopsy 1: material used to perform kidney biopsy.
Procedures related to connective tissue disease 429
Figure 5. Kidney biopsy 2: local anesthetization.
procedure usually two needle-cores of renal tissue are taken, generally using a true cut needle in a kidney-gun. After the procedure, the patient is confined to bed for 24 hours, the first 12 entailing strict bed rest. Blood pressure and heart rate are measured every 30 minutes for the first 4 hours or longer if indicated. Eating and drinking are permitted, provided the patient lies face up. The needle-cores of kidney tissue are fixed in a formaldehyde solution for light microscopical investigations. Subsequently, depending on which of the different procedures is to follow, routine staining includes haematoxylin and eosin, PAS and Masson trichrome, supplemented amongst others with silver stain to investigate possible thickening of the basement membrane, and Congo red stain to evaluate the presence of amyloid deposition. For immunofluorescence, the kidney tissue has to be frozen immediately by placing the cores in a glass cylinder immersed in liquid nitrogen. For transport to the laboratory, a container with dry ice—solid carbon dioxide—may be used. For electron microscopical investigation, the tissue cores are fixed in glutaraldehyde.
430 A. A. Kruize and J. W. J. Bijlsma
Figure 6. Kidney biopsy 3: incision skin.
Potential results In general, the interpretation of renal histology requires a pathologist with extensive experience. The different domains of the renal parenchyma have to be evaluated, namely the vascular, the tubulo-interstitial and the glomerular domains. Within the vascular domain, vasculitis of the arteries and arterioles has to be distinguished from vasculo-occlusive disease and the changes due to hypertension or
Figure 7. Kidney biopsy 4: ultrasound guided kidney biopsy.
Procedures related to connective tissue disease 431
Figure 8. Kidney biopsy 5: ultrasound imaging, arrow indicates biopsy needle.
diabetes mellitus as well as arteriolar changes due to medication. In generalised rheumatic disease, different types of vascular renal disease may be present. For instance in SLE, vasculitis has to be differentiated from thrombotic microangiopathy with antiphospholipid syndrome. Vasculitis of the small vessels is seen in microscopic angiitis, Wegener’s granulomatosis and essential mixed cryoglobulinaemia. Vasculitis with the formation of aneurysms of medium-sized arteries is characteristic of classic polyarteritis nodosa. In RA, the disease itself or medication, including NSAIDs, methotrexate and cyclosporine, may precipitate vasculitis. In systemic sclerosis and mixed connective tissue disease (MCTD), concentric thickening of arterial walls may occur and result in narrowing of the lumen.
Figure 9. Kidney biopsy 6: kidney biopsy specimen.
432 A. A. Kruize and J. W. J. Bijlsma
Figure 10. Kidney biopsy 7: kidney biopsy specimen.
Tubulo-interstitial disease may vary from acute inflammation to chronic fibrosis. Medication, including NSAIDs, cyclosporine and gold salts, may induce tubular interstitial nephritis. Lymphocytic tubulo-interstitial nephritis associated with hypokalaemic renal tubular acidosis has been observed in primary Sjo¨gren’s syndrome, while tubular interstitial nephritis has been observed in SLE, relapsing polychondritis and essential mixed cryoglobulinaemia. Lymhocytic granulomatous infiltrates in the tubulointerstitium can be seen in Wegener’s granulomatosis and eosinophil granulomatous infiltrates are seen in Churg-Strauss syndrome. Peritubular amyloidal deposits may be a late complication of longstanding rheumatic disease including RA and ankylosing spondylitis. Glomerular disease may be associated with a broad variety of well-classified multi-system autoimmune diseases. The glomerular changes may be divided into different categories. Electron microscopy is needed to detect minimal change lesions, which are sometimes seen in NSAID-induced nephropathy. In systemic rheumatic disease, glomerular sclerosis may be the eventual result of prior glomerular disease. Membranous changes, consisting of thickening of the glomerular basement membrane (GBM), may be due to changes on the epithelial side of the GBM in the form of granular deposits of immunoglobulins (‘spikes’), as in idiopathic membranous glomerulopathy. Thickening of the GBM may also be due to changes from the endothelial side in the form of interposition of mesangial cell pseudopodia (‘double contour’) as in mesangiocapillary glomerulonephritis. Proliferative changes in the form of hypercellularity (‘crescent formation’) reflect destructive forms of glomerulonephritis, including RPGN. Mesangial glomerulonephritis has been associated with relapsing polychondritis. Focal proliferative glomerulonephritis may be associated with RA and, if mesangial IgA deposits are present, with reactive arthritis and ankylosing spondylitis. Diffuse proliferative glomerulonephritis with mesangial IgA deposits is seen in HenochScho¨nlein purpura. Membranous and mesangial glomerulonephritis may occur in mixed connective tissue disease.
Procedures related to connective tissue disease 433
All of these types may be seen in lupus nephritis, see below.3,14
World Health Organisation classes of lupus nephritis † Normal, or minimal glomerular abnormality No glomerular abnormalities on light microscopy † Mesangial nephropathy Mesangial expansion, including hypercellularity, increased matrix and immune complex deposits † Focal proliferative glomerulonephritis Predominantly segmental hypercellularityGnecrosis compromising the circulatory space of capillary loops in %50% of glomeruli, mesangial and subendothelial immune complex deposits † Diffuse proliferative glomerulonephritis Characteristically global but irregular hypercellularityGnecrosisGcellular crescents affecting O50% of glomeruli; variable sclerosis, atrophy, fibrosis; mesangial and subendothelial (G subepithelial) immune complex deposits † Membranous nephropathy Generalised thickening of capillary loops; mesangial and subepithelial immune complex deposits † Sclerosing nephropathy Predominantly hyalinised glomeruli, tubular atrophy, interstitial fibrosis; no (or very rare) immune complex deposits Complications The main complication is post-biopsy bleeding. This occurs in 3–5% of the patients, mostly as a subcapsular haematoma but also as heavy haematuria, sometimes causing circulatory shock, ultimately necessitating nephrectomy in 0.2% of the patients. Bleeding may also result in pain in the side and paralytic ileus; coagulation within the urinary tract may cause obstruction and consequently colic. See also table with listed complications given previously.
BIOPSY OF THE TEMPORAL ARTERY Indications Giant cell arteritis is a systemic vasculitis that affects large and medium-sized arteries. Although it may involve any artery, it has a propensity to affect the branches of the external carotid artery, particularly the posterior ciliary arteries that supply the optic nerve and the superficial temporal artery, hence its alternative name of ‘temporal arteritis’. There is an overlap between temporal arteritis, giant cell arteritis and polymyalgia rheumatica, all of them probably being manifestations of one disease spectrum. The indication for temporal biopsy is clinical suspicion of giant cell arteritis in a patient with specific complaints of polymyalgia rheumatica: bilateral shoulder pain
434 A. A. Kruize and J. W. J. Bijlsma
and/or stiffness, age O50, increased ESR (erythrocyte sedimentation rate), depression and/or weight loss. However, non-specific symptoms are common and suspicion should be raised in elderly patients with systemic upset such as weight loss or fatigue. Suspicion is high when clinical symptoms related to the affected arteries are present: headache, tenderness of the scalp, particularly around the temporal and occipital arteries, visual disturbances or jaw claudication. There is, however, no consensus about in whom a biopsy should be performed. Technical procedures The skin is anaesthetised by infiltrating 1–2% lidocaine around the site of biopsy, the often palpable arteria temporalis. It has been suggested, but not proven, that imaging, such as ultrasound or MRI, may increase the yield of a biopsy when the artery is not palpable. For cosmetic reasons, the main line of incision should follow the natural crease lines of the skin. Although incision biopsy should yield a reasonable biopsy of at least 5 cm, for cosmetic reasons the incision should not be overly large. Formalin 10% is most widely used for routine microscopy of material embedded in paraffin. For specific techniques, including immunofluorescence, more specific transport media and fixatives are available. The tissue is embedded in plastic and transverse cuts are made every 1–2 mm to ensure that ‘skip’ lesions are found. Apart from haematoxin-eosin staining an elastica stain (Von Gieson) is mandatory. Potential results The histological appearance of giant cell arteritis is a panarteritis with giant cell granuloma formation, often in close proximity to a disrupted internal elastica lamina (Figures 11 and 12).
Figure 11. Temporal arteritis 1: (A) multinucleated giant cell, (B) narrowed lumen due to hyperplasia of the intima, (C) media, (D) inflammatory cell infiltration.
Procedures related to connective tissue disease 435
Figure 12. Temporal arteritis 2: (A) disruption of the internal elastic lamina, (B) hyperplasia of the intima, (C) narrowed lumen containing erythrocytes, (D) vascular branch.
Large and medium-sized arteries are affected, the involvement is patchy and skip lesions are often found. There is preferential involvement of muscular arteries with well developed internal and external lamina elastica. These gross features are not characteristic. The vessels are enlarged and nodular and have little or no lumen due to intimal proliferation. Thrombosis often develops at sites of active inflammation and later these areas may recanalise.15 The adventitia is usually invaded by mononuclear inflammatory cells; in the media giant cells may be present, which vary from small cells with two to three nuclei up to masses of 100 mm containing many nuclei. A temporal progression can be observed from an active infiltrative phase to a scarred artery with little cellular infiltrate. At first the most severe changes are centred on the internal lamina elastica, which becomes swollen and fragmented. Fragments of elastic tissue have been demonstrated within giant cells. The histological changes of healed arteritis include medial chronic inflammation with ingrowth of new blood vessels, focal medial scarring and a bizarre pattern of intimal fibrosis. The finding of healed vasculitis in a biopsy by no means excludes the possibility of an active lesion elsewhere. Complications Bleeding and infection (see also the table with listed complications given previously) are seldom seen. In some elderly patients, in particular, wound healing is not optimal. Hypesthaesia of the skin may persist for a prolonged period of time. ACKNOWLEDGEMENTS The authors wish to thank their colleagues CAFM Bruynzeel Koomen, RHWM Derksen, CJM Frijns, RJ Hene´, JE Hoogendijk and JWG Jacobs, for their suggestions on an earlier version of this chapter.
436 A. A. Kruize and J. W. J. Bijlsma
REFERENCES 1. Cerio R. Histopathology of the skin: general principles. In Champion JL, Burns DA & Breathnach SM (eds.) Rook/Wilkinson/Ebling Textbook of Dermatology, 6th edn. Oxford: Blackwell, 1998, pp. 159–203. 2. Freedberg IM, Eisen AZ, Wolff K et al. Fitzpatrick’s Dermatology in General Medicine. 6th edn. New York: McGraw-Hill; 2003. 3. Lahita RG. Systemic Lupus Erythematosus. 3rd edn. London: Academic Press; 1999. 4. Hochberg MC, Silman AJ, Smolen JS et al. Rheumatology. 3rd edn. St. Louis: Mosby; 2003. 5. Klemi PJ, Sorsa S & Happonen RP. Fine-needle aspiration biopsy from subcutaneous fat. Scand J Rheumatol 1987; 16: 429–431. 6. Chisholm DM & Mason DK. Labial salivary gland in Sjo¨gren’s syndrome. J Clin Pathol 1968; 21: 656–660. 7. Vitali C, Bombardieri S, Johnson R et al. Classification criteria for Sjo¨gren’s syndrome: a revised version of the European criteria proposed by the American-European Consensus Group. Ann Rheum Dis 2002; 61: 554–558. 8. De Wilde PCM, Kater L, Baak JPA et al. A new and highly sensitive immunohistologic diagnostic criterion for Sjo¨gren’s syndrome. Arthritis Rheum 1989; 32: 1214–1220. 9. Dalakas MC & Hohlfeld R. Polymyositis and dermatomyositis. Lancet 2003; 362: 971–982. 10. Dalakas MC. Muscle biopsy findings in inflammatory myopathies. Rheum Dis Clin North America 2002; 28: 779–798. 11. Van der Meulen MF, Ronner IM, Hoogendijk JE et al. Polymyositis. An over diagnosed entity. Neurology 2003; 61: 316–321. 12. O’Rourke KS & Ike RW. Muscle biopsy. Curr Opin Rheumatol 1995; 7: 426–428. 13. Ike RW. Minimal invasive procedures. In Hochberg MC, Silman AJ, Smolen JS, Weinblatt ME & Weisman MH (eds.) Rheumatology. Edinburgh: Mosby, 2003, pp. 245–247. 14. Ballow JE, Boumpas DT & Austin HA. Systemic lupus erythematosus and the kidney. In Lahita RG (ed.) Systemic Lupus Erythematosus. Amsterdam: Elsevier, 2004, pp. 877–912. 15. Hazleman BL. Polymyalgia rheumatica and giant cell arteritis. In Hochberg MC, Silman AJ, Smolen JS, Weinblatt ME & Weisman MH (eds.) Rheumatology. Edinburgh: Mosby, 2003, pp. 1623–1633.
5 Procedures related to connective tissue disease Aike A. Kruize*
MD, PhD
Rheumatologist
Johannes W.J. Bijlsma
MD, PhD
Professor and Head Department of Rheumatology and Clinical Immunology, F02.127, University Medical Center, Box 85500, 3508 GA Utrecht, The Netherlands
In patients with rheumatic and, especially, connective tissue diseases many organ systems apart from the joints may be involved, such as the skin, muscles, salivary glands, nerves, kidneys and blood vessels. Biopsies of these tissues may help in establishing a diagnosis, in assessing the extent and severity of the disease and in monitoring the success of therapy. In this chapter, indications for biopsy, technical procedures, potential results and possible complications are described. The emphasis is on when a biopsy is indicated, how the patient is prepared, how the biopsy is performed, what results can be expected from the biopsy and what are the possible sequelae. For full details of the histology of e.g. a renal biopsy in a lupus patient, the reader is referred to the relevant textbooks. Key words: skin biopsy; subcutaneous fat biopsy; salivary gland biopsy; muscle biopsy; peripheral nerve biopsy; renal biopsy; temporal artery biopsy.
BIOPSIES OF THE SKIN Indications Abnormalities of the skin may provide clues for the diagnosis of a generalised rheumatic disease. To diagnose suspected autoimmune dermatose, skin biopsy material for histological examination, including immunofluorescence, may be of great help. Examples are blistering disorders (perilesional skin) or lupus erythematosus (lesional skin). In addition, skin biopsy is used to diagnose a dermal or epidermal neoplasm or an * Corresponding author. Tel.: C31 3025 07357; Fax: C31 3025 23741. E-mail address: [email protected] (A.A. Kruize). 1521-6942/$ - see front matter Q 2005 Elsevier Ltd. All rights reserved.
418 A. A. Kruize and J. W. J. Bijlsma
infection, by culturing a fresh, unfixed skin biopsy specimen for various organisms including mycobacteria, deep fungi or by examining it for protozoa or filarial worms.1 Technical procedures Skin biopsy is essentially a minimally invasive procedure. A preferably untreated skin lesion, in an early stage and characteristic for the skin disorder as a whole, should be chosen for biopsy. If lesions are present at different stages of evolution, it may be helpful to take biopsies of different lesions. For blistering, ulcerating or necrotic disorders, the biopsy should include both normal and affected skin so that the pathologist can identify the plane of separation for a blister or the early changes adjacent to ulceration and necrosis. For cases of suspected vasculitis the location of the affected vessels must be considered: palpable purpura occur in superficial venules in the dermis while vascular changes in disorders producing necrosis or livedo are in larger vessels deep in the subcutaneous fat. When direct immuno-fluorescence is needed, the biopsy should also include normal-looking skin. In choosing the site of biopsy, one should try to avoid areas that are prone to bad healing and areas that are cosmetically important. Moreover, changes that are secondary to circumstances such as venous stasis of the lower legs in the elderly may interfere with the histopathological abnormalities associated with the lesion and therefore be a reason for avoiding such sites. The skin is anaesthetised by infiltrating 1–2% lidocaine with or without epinephrine around the site of biopsy. Because epinephrine, more than lidocaine, may cause distortion artifacts which interfere with the histological interpretation of the biopsy specimen, some caution with the dosage is advised. It is probable that some of the discussion on the negative effects of adrenaline has been exaggerated. When taking a biopsy of the extremities, however, incidental extreme vasospasm might cause necrosis of the tissue. Slow delivery may decrease the pain associated with the installation of the anaesthesia. Superficial injection, creating an oedematous wheal has immediate efficacy but is more painful. Several techniques for skin biopsy are available, depending on the purpose of the subsequent investigation. For diagnosing a dermatose possibly associated with a generalised rheumatic disease, punch biopsy, incision biopsy and excision biopsy are the most commonly used techniques.2 A punch biopsy samples both the epidermis and dermis; in most cases a 3–4 mm punch is adequate. A disposable punch is generally used, having a cutting edge attached to a handle. A small cylinder of tissue is removed by means of a twisting movement of the punch. The specimen is extracted using forceps but scissors may be needed to detach the base of the tissue sample. The wound can be left to heal by secondary intention after achieving haemostasis; the scar may take several weeks to heal. The wound can be closed primarily and the resulting scar is generally a white or hypo-pigmented linear mark. Although incision (‘wedge’) biopsy should ideally reveal material from a sample of approximately 5 mm in length, cosmetic reasons, however, may necessitate some modesty in taking material. When taking a biopsy of a larger lesion, obtaining adjacent normal skin is important. For cosmetic reasons, the main line of incision should try to follow the natural crease lines of the skin. Lesions O4 mm may have to be stitched. Small lesions may be removed totally and, if malignancy is suspected, it is important to perform an excision biopsy with clear margins. Mechanical damage of the tissue should be avoided. When different fixatives are needed for different studies, it is preferable to take two specimens rather than dividing
Procedures related to connective tissue disease 419
one specimen into smaller parts. For routine microscopy of material embedded in paraffin, 10% formalin is most widely used. For specific techniques, including immunofluorescence, more specific transport media and fixatives are available. Potential results The most commonly encountered skin lesions in patients with generalised rheumatic disease are exanthema, nodules, vesicles and bullae, ulcers, purpura and papulosqamous rashes. As in rheumatic diseases, the diagnosis of skin lesions is commonly based upon pattern recognition. Not all skin lesions warrant skin biopsy. If in doubt, histological examination may confirm a probable diagnosis, because the histological features of several skin lesions are diagnostic, such as palisading granulomas in rheumatoid nodules. Biopsy of a suspicious lesion can confirm the diagnosis of lupus in a patient with other suggestive clinical manifestations.3 Direct immunofluorescence is especially diagnostic in lupus erythematosus and blistering abnormalities. In over 70% of patients with lupus erythematosus the lupus band test, i.e. granular deposition of immunoglobulins at the dermo-epidermal junction, is present, particularly in sun exposed skin, while it is absent in normal skin. Scleroderma involves the skin and subcutaneous structures in a characteristic manner, although similar findings can occur as a localised process (e.g. morphea) or as deeper pathology, such as in the fasciitis syndromes. In scleroderma there is a risk of impaired healing of the biopsy wound, therefore care should be taken in selecting the biopsy location. Vasculitis seen on skin biopsy often prompts a search for systemic involvement and seldom stands alone as a diagnostic finding. Immunoglobulin A deposition indicates Henoch-Scho¨nlein purpura or one of its variants. Not all palpable purpurae are due to vasculitis; biopsy can show other processes, such as microthrombi or cholesterol emboli.2 When panniculitis is suspected, the biopsy should be taken deeply enough to contain subcutaneous fat. When subcutaneous nodules are found in polyarthritis they are often characteristic for rheumatoid granuloma; however, nodular vasculitis and nodules of other material, such as monosodium urate, calcium pyrophosphate, cholesterol or amyloid may occur. Complications Complications during invasive procedures All invasive procedures may lead to the following complications: † Post-biopsy bleeding, especially when aspirin, non-steroidal anti-inflammatory drugs (NSAIDs) or anticoagulants are used. † To completely prevent this bleeding medication should be stopped well before hand. † Infections: all invasive examinations can become a ‘point of entry’ for infections. The presence of cardiac valvular disease raises this risk for infection, especially subacute bacterial endocarditis; prophylactic treatment with antibiotics might be indicated.
420 A. A. Kruize and J. W. J. Bijlsma
If preoperative antibiotics are administered, the dosage needs to be adequate for the control of skin flora, especially Staphylococcus aureus; they should be started less than 2 hours pre-operatively and continued for 48–72 hours. Skin biopsy of the upper trunk has an increased risk for keloid formation in patients who tend to make keloid. Skin biopsy of the lower leg of an elderly patient may result in poor healing and be prone to infection. If an infection develops after 3–5 days, oral antibiotics may be required. The slightest suspicion of malignant melanoma should result in refraining from skin biopsy in favour of a wide excision biopsy because of the (theoretical) risk of disseminating tumor cells.
BIOPSY OF SUBCUTANEOUS FAT Indications Systemic amyloidoses are characterised by the extracellular deposition of fibrils, derived from circulating soluble precursors. In the past, amyloidosis A (AA, reactive or secondary) was quite often found in patients with longstanding rheumatoid arthritis (RA) and it was found sporadically in patients with other chronic inflammatory arthropathies.4 Perhaps due to the improvement in the treatment of most rheumatic diseases over the last decade, AA is now less often present in these patients. Patients with other forms of amyloidosis, such as AL (amyloidosis immunoglobulin light chain, or primary) and Ab2-microglobulin, dialysis associated, amyloidosis may have rheumatic symptoms such as arthritis (with a predilection for the shoulder). To diagnose amyloidosis, the definitive test is biopsy of easily accessible tissues that are expected to contain amyloid, or biopsy of clinically affected organs. Rectal biopsy and subcutaneous fat aspiration are the procedures of choice. Technical procedures Fat samples are taken from the abdominal wall between the xiphoid process and the pubic bone, using an adapter, disposable syringe (10 ml) and a disposable needle (Gauge 0.8 or 0.9). The needle is injected quickly into the subcutaneous fat, a vacuum drawn into the syringe and the needle is then moved in different directions back and forth very quickly. The vacuum is released before the needle is withdrawn from the fat. Fat tissue remaining in the needle is blown with air onto glass slides, where it is allowed to air overnight before being stained with Congo red.5 Potential results In AA patients, subcutaneous fat aspiration has been reported to have a very high sensitivity of 0.97 while in AL patients this was reported to be 0.62. However, these data come from tertiary referral centres and a significantly lower yield for this procedure has been reported in other settings. Because each form of amyloid is defined by the chemical nature of the fibril protein, pathological specimens from any site can be stained with antibodies that are specific for the precursors to establish the character of the responsible disease.
Procedures related to connective tissue disease 421
Complications In theory bleeding and infection may occur, as listed previously. However, no relevant complications have been reported.
SUBLABIAL SALIVARY GLAND BIOPSY Indications In rheumatology, confirmation of a presumed diagnosis of Sjo¨gren’s syndrome is the main indication for sublabial salivary gland biopsy. To assess the oral component of Sjo¨gren’s syndrome various techniques are used, including sialochemistry, nuclear imaging of the salivary glands and radiographical contrast sialography. Focal sialadenitis in sublabial salivary gland specimens reflects salivary gland involvement in Sjo¨gren’s syndrome as a whole, allowing for a relatively simple diagnostic procedure compared to major salivary gland biopsy. Nowadays, sublabial salivary gland biopsy is widely considered and used as the single most specific test for confirming a diagnosis of (primary) Sjo¨gren’s syndrome. Histological changes in salivary gland tissue, however, are not 100% specific. Consequently, focal sialadenitis is not pathognomic for Sjo¨gren’s syndrome: focal sialadenitis is also found in sublabial minor salivary gland specimens overlain by inflamed mucosa due to lichen planus, cheek biting or a mucocele. Consequently, sublabial salivary gland biopsy that is intended for the confirmation of a diagnosis of Sjo¨gren’s syndrome should be taken through healthy, normal appearing mucosa of the lower lip. However, in contrast to acinar atrophy, dilatation of ducts and hyperplasia of sublabial salivary gland tissue— which all increase with age and are related to inflammation—focal sialadenitis is an objective diagnostic parameter for a diagnosis of the salivary component of Sjo¨gren’s syndrome. Other diagnostic findings in sublabial salivary gland specimens include non-caseating granuloma indicating sarcoidosis, amyloidosis, iron depositions indicating haemochromatosis and abnormal appearing lymphocytes compatible with leukaemia. Technical procedures Labial salivary gland biopsy is a simple outpatient procedure. The patient is asked to lie down on his back with the upper half of the body inclined upright at 308. A brief summary explaining the procedure is given. The mucosa of the lower lip is locally infiltrated with anesthetic, e.g. 2% lidocaine in addition to vasoconstrictor epinephrine 1:80 000. A 1 cm superficial linear incision of the epithelium of the mucosa is made between the midline and commissure, parallel to the vermillion, revealing the minor salivary glands (Figure 1). The glands are bluntly dissected from the fascia and excised separately, avoiding damaging the sensory nerves, which can be clearly distinguished. An assistant should be dabbing regularly to enable an adequate view. Because histopathological findings may differ between the various glands, at least five glands have to be taken for examination (Figure 2). The incision is closed with dissolvable sutures (Figure 3). In order to avoid burning due to loss of sensation, the patient is advised not to drink hot beverages for the next few hours.
422 A. A. Kruize and J. W. J. Bijlsma
Figure 1. Lipbiopsy 1: paramedian incision of the lower lip.
The glands are fixed in a formol (-sublimate) solution, before embedding them in paraffin for histopathological examination. Inflammation, expressed as focal lymphocytic sialoadenits, is assessed semi-quantitatively. Salivary gland lobes with dilatation of ducts, parenchymal atrophy or extravascular polymorphonuclear leukocytes are excluded from the assessment. Potential results A focus of focal lymphocytic adenitis is defined as an aggregate of at least 50 lymphocytes and histocytes. Usually these foci are found at the edge of intralobular
Figure 2. Lipbiopsy 2: sublabial salivary gland biopsy specimen.
Procedures related to connective tissue disease 423
Figure 3. Lipbiopsy 3: stitched incision of the lower lip.
ducts, related to small veins, adjacent to the acini of the glands, leading to destruction of the acini while relatively sparing the ducts. More than one focus per 4 mm2 sample of sublabial salivary gland tissue—corresponding to a focal sialadenitis of grade IV6—is a consistent finding and, subsequently, a widely accepted criterion for the histopathological confirmation of Sjo¨gren’s syndrome. It is one of the main objective criteria items of the revised classification criteria proposed by the American–European Consensus group.7 The presence of duct abnormalities and the formation of epimyoepithelial cell islands in major salivary gland tissue are late features in Sjo¨gren’s syndrome, but are generally not seen in minor gland tissue. In addition, immunohistological assessment of labial salivary gland specimens is reported to be more specific for the disease than the lymphocytic focus score criterion. A bivariate classification criterion, based on the percentages of IgA- and IgG-containing plasma cells resulted in a reduction in the number of false positive diagnoses. Using the more simply manageable univariate criterion based on the percentage of IgA-containing plasma cells (!70% IgA-containing plasma cells for confirmation of Sjo¨gren’s syndrome), results in 99 and 96% specificity and sensitivity, respectively.8 Also, immunohistological detection of monotypic plasma cells in labial salivary glands—defined by a kappa:lambda ratio of R3—has been indicated as a prognosticator for systemic lymphoproliferative disease. Complications Complications of sublabial salivary gland biopsy are, apart from the complications listed previously , seldom reported, but they include arterial bleeding due to inadequate, c.q. too deep incision, vasovagal collapse and burning of the mouth while drinking hot beverages due to a temporary loss of sensation because of the anaesthesia. Persistent hypoesthaesia of the lower lip region is reported when a nerve, in spite of the procedure of which all details can be followed by sight, is damaged.
424 A. A. Kruize and J. W. J. Bijlsma
MUSCLE BIOPSY Indications Muscle biopsy is the most crucial test for establishing the diagnosis of inflammatory myopathies.9 The inflammatory myopathies encompass a heterogeneous group of acquired muscle diseases characterised clinically by subacute-onset, proximal and symmetrical muscle weakness and histologically by inflammatory infiltrates within the skeletal muscles. These myopathies comprise three major and discrete subsets: polymyositis (PM), dermatomyositis (DM) and inclusion body myositis (IBM). Each subset retains its characteristic clinical, immunopathological and morphological features regardless of whether it occurs in isolation or in connection with other systemic diseases. Although the diagnosis of these disorders is based on the combination of clinical examination, serum muscle enzyme levels and sometimes the presence of autoantibodies, muscle biopsy offers the most definitive diagnostic information in the majority of cases.10 Although recently discussion was started as to whether or not PM is an over diagnosed entity,11 the value of muscle biopsy is still undisputed. Although there is a long list of differential diagnoses for muscle weakness, such as denervating conditions, neuromuscular junction disorders, muscular dystrophies, myotonic diseases, congenital myopathies, glycogen storage disease, lipid storage myopathies and others, clinical characteristics most often make these diagnoses less likely. Nevertheless, where there is doubt, muscle biopsy will help to differentiate between these different conditions. There is seldom an indication for a repeated muscle biopsy. When the first biopsy is negative, but there is a very strong clinical suspicion, such as for IBM, a second biopsy might be indicated, in which case it is best guided by skeletal muscle computed tomography (CT) or magnetic resonance imaging (MRI) scans. In cases where muscle weakness persists or is aggravated during glucocorticoid treatment a muscle biopsy might help to differentiate a possible glucocorticoid myopathy from undertreatment or relapse of the PM. A quite different indication for performing a muscle biopsy is established when vasculitis, especially polyarteritis nodosa (PAN), is suspected in a patient. In this situation a deep and rather large muscle biopsy is taken from a distal muscle group. Technical procedure Most researchers prefer an open muscle biopsy over a needle biopsy, because the former offers a larger sample that can be properly orientated and is better suited for immuno-cytochemical and ultra-structural studies. In some centres, however, percutaneous needle biopsy is used. A moderately weak muscle is the most suitable for performing a diagnostic biopsy because it offers the best chance to obtain maximum information. If the biopsy is done to seek vasculitis, a distal muscle group should be selected. Open biopsy is performed after infiltrating a local anaesthetic into the skin and subcutaneous tissue in the region of interest, avoiding muscle. Pain fibres are located in the perimysial area and fascia, but not the muscle itself. Hence, pain from the procedure can be minimised. In addition, sedation using a short-acting benzodiazepine or narcotic may further reduce the patient’s discomfort. Open biopsy permits the direct inspection
Procedures related to connective tissue disease 425
of the muscle, enabling an easily processed specimen to be obtained, which is more likely to contain larger blood vessels and no fat.12 A longitudinal incision of 5 cm through subcutaneous tissue and fat to reach the muscle should permit the excision of a sample measuring approximately 1!1!1 cm. The two ends of the sample should be lightly affixed to an object that holds the muscle at resting length. The sample should be kept moist before being placed in fixative to avoid contraction artifact; it should be deep frozen within 2 hours. A second, smaller specimen can be taken for electron microscopy. Percutaneous sampling of muscle is performed using specially designed instruments, such as a trocar with a cutting window that admits tissue by suction, or a springactivated biopsy instrument. Skin overlying an area that has been anaesthetised (as is done for open biopsy), is incised with a scalpel to enable penetration of the deeper tissues by the biopsy instrument. The cutting window of the biopsy instrument is opened to admit tissue, which is cut free and contained by the circling blade. Both percutaneous techniques can be performed at the bedside and are relatively atraumatic. The small size of the samples obtained requires special care in processing. An advantage of percutaneous biopsy is that muscle can be sampled from a wide area by orientating the biopsy instrument in several different directions through the same skin puncture, thus encompassing a much larger region than is available in an open biopsy.13 For transportation, muscle tissue may be kept on saline moistened gauze for several hours; the specimen should be kept cool. A portion of the biopsy specimen should be snap frozen for cryostat sections and embedded in plastic for ultra thin sections and ultra structural studies. Frozen muscle may be safely shipped overnight on dry ice. When muscle is frozen it can be stored at K80 8C. Frozen muscle for histochemistry provides excellent muscle fibre morphology and most diagnostic information using light microscopy. Muscle embedded in paraffin is useful for surveys for inflammation and morphology of inflammatory cells but gives poor muscle fibre morphology. It is always important to check the techniques to be used with the laboratory that is going to analyse the muscle biopsy. It is also advised to use a specific laboratory with expertise in enzyme histochemistry and myopathology. Some people advise the use of CT or MRI scans to select an involved region of a muscle. However, the additional value is not proven and most physicians do not use it for their primary biopsy; only when the biopsy results are negative while the clinical suspicion is very high, is a second biopsy guided by CT or MRI advisable. Potential results In cases of inflammatory myositis, histochemistry, immuno-histology and, sporadically, electron microscopy may help to establish a diagnosis. For details see relevant textbooks cited in Dalakas 2002.10 In idiopathic PM mononuclear cell infiltrates, predominantly consisting of T cells, are found in the endomysial spaces as well as invasion of histologically healthy looking muscle fibres by mononuclear cytotoxic T cells. Less pronounced are necrosis and regeneration of muscle fibres, while macrophages and B cells are sparse. It has been shown that in the clinically evident PM patient, without skin abnormalities, mononuclear cell infiltrates are often localised perimysially and perivascularly.11 In cases of sporadic IBM there are endomysial mononuclear cell infiltrates with invasion of normal-looking muscle fibres and so-called rimmed vacuoles in at least 0.3% of the muscle fibres. In DM the inflammation is predominantly perivascular, in the interfascicular septae rather than within the fascicles;
426 A. A. Kruize and J. W. J. Bijlsma
intramuscular blood vessels show endothelial hyperplasia and the muscle fibres undergo necrosis and phagocytosis. Ischaemia results in atrophy of fibres in the periphery of the fascicles (perifascicular atrophy). Additional biopsy stains are useful when other diagnoses are suspected, e.g. ATPase staining to define myosin loss, elucidating either type-1 or type-2 fibre atrophy, while different enzyme stains are used to elucidate mitochondrial pathology. Enzyme deficiencies and storage diseases can become evident by staining with periodic-acidSchiff (PAS: glycogen, carbohydrate), Alcian blood (mucopolysaccharide), Sudan black, Oil red (lipids) or Congo red (amyloid). Complications There are no serious complications for either open or needle biopsy, as aforesaid. Of course bleeding is a well-known possibility and medication that influences haemostases should be stopped before the procedure. When the biopsy is performed under properly aseptic conditions the possibility for an infection is very low. Sometimes the scar does not heal properly or a dermal nerve branch has been damaged. After biopsy, patients are advised to rest for a few hours, especially after an open biopsy. When an open biopsy of the leg has been performed, the patient is advised not to overuse that leg for a few days.
SURAL NERVE BIOPSY Indications In patients with rheumatic conditions the most frequent reason to perform a nerve biopsy is suspicion of a vasculitis of the nervus suralis. This vasculitis could be part of a systemic vasculitis or accompany a connective tissue disease such as RA. In general, nerve biopsy may be performed to help distinguish between demyelination (destruction of parts of the myelin sheath covering the nerve) and axonal degeneration (destruction of the axon portion of nerve cell), to identify inflammatory neuropathies, or to confirm a specific diagnosis. Peripheral neuropathy is very common; the incidence varies with the specific type of neuropathy. Most often it is a manifestation of conditions that can damage peripheral nerves, such as diabetes mellitus, renal insufficiency or alcoholism. Symptoms differ depending on whether the damage is to the sensory fibres, the motor fibres or both. Technical procedure There is no specific preparation for the patient. A local anaesthetic is used to numb the area. A small incision is made and a section is removed of the whole nerve (not limited to only a few fascicles, since in vasculitis not all fascicles may be involved). Because of the use of the local anaesthetic, discomfort during the procedure is usually minimal. The anaesthetic may burn or sting when first injected. After the procedure the area may feel tender or sore for a few days. The nerve tissue is frozen, paraffin embedded and sectioned in plastic. Morphological changes are studied using a light microscope, but on specific indications an electron microscopical investigation is performed.
Procedures related to connective tissue disease 427
Potential results With regard to specific rheumatic conditions, interest has focused on signs of inflammation, especially cellular infiltrates. Invasion of neutrophils into the vessel walls are suggestive for nercotising vasculitis. Eosinophils can also be seen in vasculitis, especially Churg-Strauss syndrome, but also in fasciitis and eosinophilia myalgia syndrome. Invasion of (epithelioid) histiocytes is seen in sarcoidosis, leprosy and also fasciitis. Plasma cell infiltrations in plasma cell dyscrasia can be seen in vasculitis, in Lyme’s disease and in leprosy. Perineural cell infiltrates are found in perineuritis and leprosy. In addition, information is gathered from nerve biopsies regarding demyelination, differential fascicular loss, neurofilament accumulation, mitochondrial abnormalities, axon loss, regeneration of axons and many other signs. In evaluating some of these differential diagnoses electro-physiological studies are also performed. In a case of suspected vasculitis it is important to select an appropriate nerve for biopsy, to also look at the size of the vessels involved and to look for the integrity of the lamina elastica of the vessel. Immune-mediated neuropathies also include Guillain-Barre´ syndrome (demyelinating disease) and chronic idiopathic axonal polyneuropathy associated with gammopathy. When the diagnosis of mononeuritis multiplex is made, a full examination of the patient is performed. In a case of a wellknown connective tissue disease, such as RA, the association is clear and in general more intensive therapy is indicated. However, if the mononeuritis multiplex is the first sign of a connective tissue disease then an intensive evaluation of the patient is warranted. Complications Removal of a part of the sural nerve leads to nerve damage, with hypaesthesia of the lateral site of the foot. Around 10% of the patients involved report persistent pain in this area. It is uncommon that this nerve damage leads to functional impairment. Of course the procedure will give some discomfort to the patient and there is a small risk of infection and bleeding, see table with listed complications given previously. In some cases an allergic reaction to the local anaesthetic has been reported.
BIOPSIES OF THE KIDNEY Indications In rheumatology, the main indication for biopsy of the kidney will be renal disease in association with an already defined generalised autoimmune disease, especially an earlier, established, diagnosis of systemic lupus erythematosus (SLE). In SLE, during the course of the disease, repeated kidney biopsy may be required. Renal findings in SLE do not correspond with the various subtypes of glomerulonephritis, therefore, renal histology is essential for determining the subtype of SLE nephritis and, consequently for establishing the ‘renal’ prognosis and required treatment. In patients with SLE regular evaluation of urine to identify abnormalities, such as red cells, proteinuria or casts is essential, since an ‘active urine’ may precede other signs of kidney involvement by many months. Also, unexplained persistent
428 A. A. Kruize and J. W. J. Bijlsma
proteinuria and/or haematuria, acute or chronic renal failure, persistent nephritic syndrome and acute nephritic syndrome in any patient with a rheumatic disease including RA, ankylosing spondylitis and reactive arthritis also form an indication for kidney biopsy. A clinical presentation of rapidly progressive glomerulonephritis (RPGN), which needs treatment without delay, requires urgent kidney biopsy. This may occur within the context of an already established rheumatic disease such as SLE, antineutrophil cytoplasmatic antibody (ANCA)-related vasculitis, including Wegener’s granulomatosis, and relapsing polychondritis. However, RPGN may also be part of the initiative phase of a generalised autoimmune or other disease, such as bacterial endocarditis and Goodpasture’s syndrome, requiring completely different treatments. Technical procedures The standard procedure for obtaining renal tissue is a percutaneous needle biopsy (Figures 4–10). Ultrasound is used to position the needle correctly. Radioscopic and CT positioning guidance are also used. In cases of extreme adiposity or repeated failure of percutaneous needle biopsy, alternative procedures such as surgical or laparoscopic biopsy may be required. Before percutaneous needle biopsy the patient is examined for bleeding diathesis (bleeding time, APTT (activated partial thromboplastin time) and PTT (prothrombin time)). NSAIDs are stopped 1 week before the bleeding time is checked. In cases of thrombopathy, prophylactic use of vasopressin may be indicated. Using ultrasound, the presence and position of two kidneys and the absence of congenital abnormalities, such as a horseshoe kidney, is checked. In cases where vasculitis is suspected, especially polyarteritis nodosa, angiographical examination of the kidney before the biopsy is performed is mandatory. If asked for, the patient receives 10 mg of oral diazepam, 60 minutes prior to the procedure, for sedation and muscle relaxation. The patient is asked to lie facedown and, subsequently, the skin is anaesthetised. For a standard
Figure 4. Kidney biopsy 1: material used to perform kidney biopsy.
Procedures related to connective tissue disease 429
Figure 5. Kidney biopsy 2: local anesthetization.
procedure usually two needle-cores of renal tissue are taken, generally using a true cut needle in a kidney-gun. After the procedure, the patient is confined to bed for 24 hours, the first 12 entailing strict bed rest. Blood pressure and heart rate are measured every 30 minutes for the first 4 hours or longer if indicated. Eating and drinking are permitted, provided the patient lies face up. The needle-cores of kidney tissue are fixed in a formaldehyde solution for light microscopical investigations. Subsequently, depending on which of the different procedures is to follow, routine staining includes haematoxylin and eosin, PAS and Masson trichrome, supplemented amongst others with silver stain to investigate possible thickening of the basement membrane, and Congo red stain to evaluate the presence of amyloid deposition. For immunofluorescence, the kidney tissue has to be frozen immediately by placing the cores in a glass cylinder immersed in liquid nitrogen. For transport to the laboratory, a container with dry ice—solid carbon dioxide—may be used. For electron microscopical investigation, the tissue cores are fixed in glutaraldehyde.
430 A. A. Kruize and J. W. J. Bijlsma
Figure 6. Kidney biopsy 3: incision skin.
Potential results In general, the interpretation of renal histology requires a pathologist with extensive experience. The different domains of the renal parenchyma have to be evaluated, namely the vascular, the tubulo-interstitial and the glomerular domains. Within the vascular domain, vasculitis of the arteries and arterioles has to be distinguished from vasculo-occlusive disease and the changes due to hypertension or
Figure 7. Kidney biopsy 4: ultrasound guided kidney biopsy.
Procedures related to connective tissue disease 431
Figure 8. Kidney biopsy 5: ultrasound imaging, arrow indicates biopsy needle.
diabetes mellitus as well as arteriolar changes due to medication. In generalised rheumatic disease, different types of vascular renal disease may be present. For instance in SLE, vasculitis has to be differentiated from thrombotic microangiopathy with antiphospholipid syndrome. Vasculitis of the small vessels is seen in microscopic angiitis, Wegener’s granulomatosis and essential mixed cryoglobulinaemia. Vasculitis with the formation of aneurysms of medium-sized arteries is characteristic of classic polyarteritis nodosa. In RA, the disease itself or medication, including NSAIDs, methotrexate and cyclosporine, may precipitate vasculitis. In systemic sclerosis and mixed connective tissue disease (MCTD), concentric thickening of arterial walls may occur and result in narrowing of the lumen.
Figure 9. Kidney biopsy 6: kidney biopsy specimen.
432 A. A. Kruize and J. W. J. Bijlsma
Figure 10. Kidney biopsy 7: kidney biopsy specimen.
Tubulo-interstitial disease may vary from acute inflammation to chronic fibrosis. Medication, including NSAIDs, cyclosporine and gold salts, may induce tubular interstitial nephritis. Lymphocytic tubulo-interstitial nephritis associated with hypokalaemic renal tubular acidosis has been observed in primary Sjo¨gren’s syndrome, while tubular interstitial nephritis has been observed in SLE, relapsing polychondritis and essential mixed cryoglobulinaemia. Lymhocytic granulomatous infiltrates in the tubulointerstitium can be seen in Wegener’s granulomatosis and eosinophil granulomatous infiltrates are seen in Churg-Strauss syndrome. Peritubular amyloidal deposits may be a late complication of longstanding rheumatic disease including RA and ankylosing spondylitis. Glomerular disease may be associated with a broad variety of well-classified multi-system autoimmune diseases. The glomerular changes may be divided into different categories. Electron microscopy is needed to detect minimal change lesions, which are sometimes seen in NSAID-induced nephropathy. In systemic rheumatic disease, glomerular sclerosis may be the eventual result of prior glomerular disease. Membranous changes, consisting of thickening of the glomerular basement membrane (GBM), may be due to changes on the epithelial side of the GBM in the form of granular deposits of immunoglobulins (‘spikes’), as in idiopathic membranous glomerulopathy. Thickening of the GBM may also be due to changes from the endothelial side in the form of interposition of mesangial cell pseudopodia (‘double contour’) as in mesangiocapillary glomerulonephritis. Proliferative changes in the form of hypercellularity (‘crescent formation’) reflect destructive forms of glomerulonephritis, including RPGN. Mesangial glomerulonephritis has been associated with relapsing polychondritis. Focal proliferative glomerulonephritis may be associated with RA and, if mesangial IgA deposits are present, with reactive arthritis and ankylosing spondylitis. Diffuse proliferative glomerulonephritis with mesangial IgA deposits is seen in HenochScho¨nlein purpura. Membranous and mesangial glomerulonephritis may occur in mixed connective tissue disease.
Procedures related to connective tissue disease 433
All of these types may be seen in lupus nephritis, see below.3,14
World Health Organisation classes of lupus nephritis † Normal, or minimal glomerular abnormality No glomerular abnormalities on light microscopy † Mesangial nephropathy Mesangial expansion, including hypercellularity, increased matrix and immune complex deposits † Focal proliferative glomerulonephritis Predominantly segmental hypercellularityGnecrosis compromising the circulatory space of capillary loops in %50% of glomeruli, mesangial and subendothelial immune complex deposits † Diffuse proliferative glomerulonephritis Characteristically global but irregular hypercellularityGnecrosisGcellular crescents affecting O50% of glomeruli; variable sclerosis, atrophy, fibrosis; mesangial and subendothelial (G subepithelial) immune complex deposits † Membranous nephropathy Generalised thickening of capillary loops; mesangial and subepithelial immune complex deposits † Sclerosing nephropathy Predominantly hyalinised glomeruli, tubular atrophy, interstitial fibrosis; no (or very rare) immune complex deposits Complications The main complication is post-biopsy bleeding. This occurs in 3–5% of the patients, mostly as a subcapsular haematoma but also as heavy haematuria, sometimes causing circulatory shock, ultimately necessitating nephrectomy in 0.2% of the patients. Bleeding may also result in pain in the side and paralytic ileus; coagulation within the urinary tract may cause obstruction and consequently colic. See also table with listed complications given previously.
BIOPSY OF THE TEMPORAL ARTERY Indications Giant cell arteritis is a systemic vasculitis that affects large and medium-sized arteries. Although it may involve any artery, it has a propensity to affect the branches of the external carotid artery, particularly the posterior ciliary arteries that supply the optic nerve and the superficial temporal artery, hence its alternative name of ‘temporal arteritis’. There is an overlap between temporal arteritis, giant cell arteritis and polymyalgia rheumatica, all of them probably being manifestations of one disease spectrum. The indication for temporal biopsy is clinical suspicion of giant cell arteritis in a patient with specific complaints of polymyalgia rheumatica: bilateral shoulder pain
434 A. A. Kruize and J. W. J. Bijlsma
and/or stiffness, age O50, increased ESR (erythrocyte sedimentation rate), depression and/or weight loss. However, non-specific symptoms are common and suspicion should be raised in elderly patients with systemic upset such as weight loss or fatigue. Suspicion is high when clinical symptoms related to the affected arteries are present: headache, tenderness of the scalp, particularly around the temporal and occipital arteries, visual disturbances or jaw claudication. There is, however, no consensus about in whom a biopsy should be performed. Technical procedures The skin is anaesthetised by infiltrating 1–2% lidocaine around the site of biopsy, the often palpable arteria temporalis. It has been suggested, but not proven, that imaging, such as ultrasound or MRI, may increase the yield of a biopsy when the artery is not palpable. For cosmetic reasons, the main line of incision should follow the natural crease lines of the skin. Although incision biopsy should yield a reasonable biopsy of at least 5 cm, for cosmetic reasons the incision should not be overly large. Formalin 10% is most widely used for routine microscopy of material embedded in paraffin. For specific techniques, including immunofluorescence, more specific transport media and fixatives are available. The tissue is embedded in plastic and transverse cuts are made every 1–2 mm to ensure that ‘skip’ lesions are found. Apart from haematoxin-eosin staining an elastica stain (Von Gieson) is mandatory. Potential results The histological appearance of giant cell arteritis is a panarteritis with giant cell granuloma formation, often in close proximity to a disrupted internal elastica lamina (Figures 11 and 12).
Figure 11. Temporal arteritis 1: (A) multinucleated giant cell, (B) narrowed lumen due to hyperplasia of the intima, (C) media, (D) inflammatory cell infiltration.
Procedures related to connective tissue disease 435
Figure 12. Temporal arteritis 2: (A) disruption of the internal elastic lamina, (B) hyperplasia of the intima, (C) narrowed lumen containing erythrocytes, (D) vascular branch.
Large and medium-sized arteries are affected, the involvement is patchy and skip lesions are often found. There is preferential involvement of muscular arteries with well developed internal and external lamina elastica. These gross features are not characteristic. The vessels are enlarged and nodular and have little or no lumen due to intimal proliferation. Thrombosis often develops at sites of active inflammation and later these areas may recanalise.15 The adventitia is usually invaded by mononuclear inflammatory cells; in the media giant cells may be present, which vary from small cells with two to three nuclei up to masses of 100 mm containing many nuclei. A temporal progression can be observed from an active infiltrative phase to a scarred artery with little cellular infiltrate. At first the most severe changes are centred on the internal lamina elastica, which becomes swollen and fragmented. Fragments of elastic tissue have been demonstrated within giant cells. The histological changes of healed arteritis include medial chronic inflammation with ingrowth of new blood vessels, focal medial scarring and a bizarre pattern of intimal fibrosis. The finding of healed vasculitis in a biopsy by no means excludes the possibility of an active lesion elsewhere. Complications Bleeding and infection (see also the table with listed complications given previously) are seldom seen. In some elderly patients, in particular, wound healing is not optimal. Hypesthaesia of the skin may persist for a prolonged period of time. ACKNOWLEDGEMENTS The authors wish to thank their colleagues CAFM Bruynzeel Koomen, RHWM Derksen, CJM Frijns, RJ Hene´, JE Hoogendijk and JWG Jacobs, for their suggestions on an earlier version of this chapter.
436 A. A. Kruize and J. W. J. Bijlsma
REFERENCES 1. Cerio R. Histopathology of the skin: general principles. In Champion JL, Burns DA & Breathnach SM (eds.) Rook/Wilkinson/Ebling Textbook of Dermatology, 6th edn. Oxford: Blackwell, 1998, pp. 159–203. 2. Freedberg IM, Eisen AZ, Wolff K et al. Fitzpatrick’s Dermatology in General Medicine. 6th edn. New York: McGraw-Hill; 2003. 3. Lahita RG. Systemic Lupus Erythematosus. 3rd edn. London: Academic Press; 1999. 4. Hochberg MC, Silman AJ, Smolen JS et al. Rheumatology. 3rd edn. St. Louis: Mosby; 2003. 5. Klemi PJ, Sorsa S & Happonen RP. Fine-needle aspiration biopsy from subcutaneous fat. Scand J Rheumatol 1987; 16: 429–431. 6. Chisholm DM & Mason DK. Labial salivary gland in Sjo¨gren’s syndrome. J Clin Pathol 1968; 21: 656–660. 7. Vitali C, Bombardieri S, Johnson R et al. Classification criteria for Sjo¨gren’s syndrome: a revised version of the European criteria proposed by the American-European Consensus Group. Ann Rheum Dis 2002; 61: 554–558. 8. De Wilde PCM, Kater L, Baak JPA et al. A new and highly sensitive immunohistologic diagnostic criterion for Sjo¨gren’s syndrome. Arthritis Rheum 1989; 32: 1214–1220. 9. Dalakas MC & Hohlfeld R. Polymyositis and dermatomyositis. Lancet 2003; 362: 971–982. 10. Dalakas MC. Muscle biopsy findings in inflammatory myopathies. Rheum Dis Clin North America 2002; 28: 779–798. 11. Van der Meulen MF, Ronner IM, Hoogendijk JE et al. Polymyositis. An over diagnosed entity. Neurology 2003; 61: 316–321. 12. O’Rourke KS & Ike RW. Muscle biopsy. Curr Opin Rheumatol 1995; 7: 426–428. 13. Ike RW. Minimal invasive procedures. In Hochberg MC, Silman AJ, Smolen JS, Weinblatt ME & Weisman MH (eds.) Rheumatology. Edinburgh: Mosby, 2003, pp. 245–247. 14. Ballow JE, Boumpas DT & Austin HA. Systemic lupus erythematosus and the kidney. In Lahita RG (ed.) Systemic Lupus Erythematosus. Amsterdam: Elsevier, 2004, pp. 877–912. 15. Hazleman BL. Polymyalgia rheumatica and giant cell arteritis. In Hochberg MC, Silman AJ, Smolen JS, Weinblatt ME & Weisman MH (eds.) Rheumatology. Edinburgh: Mosby, 2003, pp. 1623–1633.