Cervical smear sampling quality

926

SUMMARY OF CLINICAL AND LABORATORY FINDINGS

recovering from malnutrition. After a few days

on a maintenance energy diet without added zinc, the children were fed recovery diets in which the energy and protein contents varied. 10 children were given supplementary zinc (2 mg/kg per day) that was added to the recovery milk-based diet throughout catch-up growth. Plasma IGF-1 concentration was measured at weekly intervals. Both energy and protein intakes were significantly associated with plasma IGF-1(p=0003 and p=0001, respectively). However, multiple regression analysis showed that zinc was significantly associated with plasma IGF-1after controlling for energy and protein intakes (p < 0’0001). Zinc was also significantly related to the rate of weight gain (p < 0001). However, after controlling for energy and protein intake, this association was not significant. Zinc may augment IGF production through linkage with peptide-hormone cell-surface receptors. Although your editorial notes that binding of human growth hormone to the growth hormone receptor was not found to be zinc dependent by Genentech researchers, some evidence exists to show that zinc exerts a dose-dependent stimulation of human-growth-hormonespecific binding to isolated rat adipocytes.5 Hence the zinc/growthhormone-receptor complex may initiate a series of cytosolic events that ultimately lead to IGF-1 mRNA synthesis and expression. Since zinc also modulates insulin sensitivity, and insulin is involved in IGF-1 synthesis and release, both growth hormone and insulin receptors may be involved in the zinc link.

Tropical Metabolism Research Unit, University of the West Indies, Kingston 7, Jamaica

HAZEL M. PAYNE-ROBINSON MICHAEL H. N. GOLDEN BARBARA E. GOLDEN DONALD T. SIMEON

1. Golden BE, Golden MHN. Plasma zinc and the clinical features of malnutrition. Am J Clin Nutr 1979; 32: 2940. 2. Robinson HM, Picou D. A comparison of fasting plasma insulin and growth hormone concentrations in marasmic, kwashiorkor, marasmic-kwashiorkor and underweight children. Pediatr Res 1977; 11: 637. 3. Editorial. Laron dwarfism. Lancet 1991; 337: 146. 4. Maes M, Underwood LE, Ketelslegers JM. Plasma somatomedin-C in fasted and refed rats: close relationship between changes in liver somatogenic but not lactogenic binding sites. J Endocrinol 1983; 97: 243. 5. Herington AC. Effect of zinc on the binding and action of growth hormone in isolated rat adipocytes. Biochem Int 1985; 11: 853. 6. Arquilla ER, Packer S, Tarmas W, Miyamoto S. The effect of zinc on insulin metabolism. Endocrinology 1978; 103: 1440.

Amphotericin

B for second-line treatment of

Indian kala-azar SIR,-Amphotericin B has sometimes been used in leishmaniasis ’l-3 but at the usual dosage for systemic mycoses it tends to be toxic. We postulated that it should be easier to get rid of a parasite than a saprophyte infecting an immunocompromised host; if this agent were effective at lower doses it might be an acceptable alternative to pentamidine as a second-line drug in visceral leishmaniasis. We studied 15 patients with Indian kala-azar (10 men, 5 women; mean age 26 years, mean weight 47 kg). They had all been treated with sodium stibogluconate but had failed to respond or had relapsed. 3 patients had been given pentamidine and had relapsed. A bone marrow smear was stained with Leishman and only those patients in whom the smear yielded the amastigote form of leishmania were studied. Patients with cardiac, renal, pulmonary, or hepatic complications were excluded. We used amphotericin B solution (10 ml sterile water added to a vial containing 50 mg) and a test dose of 1 mg; the dose was stepped up in daily increments of 5 mg to 05 mg/kg on alternate days for 28 days. Every time 540 ml 5% dextrose solution was infused over 4--6 h. Injections of hydrocortisone and pheniramine were kept to hand. If there was an adverse reaction lower doses were continued until the patient started tolerating the drug. Patients were examined daily and laboratory investigations were done weekly. Parasitological cure was defined by absence of Leishman-Donovan bodies at the end of treatment. All 15 patients responded clinically; their hepatosplenomegaly regressed and leucocyte and platelet counts and haemoglobin levels rose. All became parasitologically negative after 1 month of therapy.

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Before vs after treatment ’p < 0 01, tp < 0 001

after 5-14 months the spleens showed further without regression any further treatment (table). Only 1 patient relapsed (after 4 months) and had to be treated with pentamidine.1 patient became hyperpyrexial and needed a hydrocortisone injection; another patient had herpes zoster and treatment had to be interrupted for 10 days. Mild adverse effects (fever, chills, diarrhoea, trace albuminuria) were quite common after the first four or five injections, after which they disappeared. Unresponsiveness to kala-azar is a serious problem necessitating the use of toxic drugs. While newer drugs such as gold4 are in the offmg we felt amphotericin B could do with re-evaluation. This agent proved very effective and reasonably safe. The paediatric scalp vein needles and the high dilutions we used may have prevented many adverse effects, including thrombophlebitis. At

follow-up

Darbhanga Medical College, Darbhanga, Bihar-846003, India

MOHAN MISHRA M. P. SINGH D. CHOUDHURY V. P. SINGH A. B. KHAN

JD. Chemotherapy for leishmaniasis: biochemical mechanisms, clinical efficacy, and future strategies. Rev Infect Dis 1988; 10: 560-86. 2. Prat A. Treatment of kala-azar with amphotericin B. Trans R Soc Med Hyg 1963; 57: 1. Berman

266-268. 3. WHO Expert Committee. The leishmaniases. WHO Tech Rep Ser 1984; 701: 104 4. Singh MP, Mishra M, Khan AB, Ramadas SL, Panjiyar S. Gold treatment for kala-azar. Br Med J 1989; 299: 1318.

Cervical

smear

sampling quality

SIR,-Dr Mitchell and Dr Medley report (Feb 2, p 265) a longitudinal study, showing no higher rate of pathological cells in cervical smears after two years in women whose first smear lacked endocervical cells, than in those from women whose first smear contained those cells. They conclude that the absence of endocervical cells should not be a reason for early rescreening. Without advocating an immediate or early repeat test, we hope that Mitchell and Medley’s conclusions will not cause a reduction in the efforts of sample takers to provide good quality smears. Mitchell and Medley cite studies showing a relation between the presence of endocervical cells and the rate of reported abnormal findings, as well as those identifying the lack of endocervical cells as a major predictive factor for false-negative smears. We are preparing a report, based on 17 000 smears, of the relation between five possible quality criteria and the rate of abnormal findings. The presence of endocervical cells has proved an important predictive factor with an odds ratio (OR) of 1 ’5 adjusted for the other criteria, age, and the identity of the cytologist. The large number of studies on the impact of endocervical cells may wrongly suggest that sampling quality and the presence of endocervical cells are the same. Other factors are also important, however. A restrospective study on paired spatula-’Cytobrush’ smears showed a three-fold increase in false-negative diagnoses when endocervical (or metaplastic) cells were absent, and a five-fold increase when squamous cells were lacking (Acta Cytol, in press). Preliminary analysis of a continuing study on the effect of ectopia on the presence of squamous cells suggests that an ectopia with a diameter of 1 cm or more increases (relative risk 2-5) the possibility of an endocervical sample without squamous cells. Our data also suggest a very important relation between late or insufficient fixation and the rate of pathological smears (OR 7 14, adjusted for the other criteria, age, and identity of the cytologist).

927

Smear takers should concentrate on providing high quality cervical smears, containing endocervical as well as squamous cells, with immediate fixation. The cytologist should report on these quality criteria. The provision of the best devices (spatula with cytobrush or ’Cervex’), training in techniques, and feed-back on adequacy of smears should lead to a high level of sampling quality. We think this is more important than whether the lack of endocervical cells should be reason for an early repeat cervical smear. We also suggest that publications about cervical intraepithelial neoplasia, especially those on cervical screening, should provide information on sampling quality. Whenever possible, an inter-observer assessment should also be included. Department of General Practice, State University Maastricht, 6200 MD Maastrich, Netherlands

F. BUNTINX H. F. J. M. CREBOLDER J. A. KNOTTNERUS

Department of Gynaecology, University Hospital, Maastricht

G. G. M. ESSED

The

Division of Immunological Medicine, MRC Clinical Research Centre, Harrow HA1 3UJ, UK

powerful placebo

concerned about Professor Ernst and colleagues’ report (March 9, p 611). The ethics of prescribing a placebo preparation outside a clinical trial, whether ointment or tablet, have to be questioned. Many chronic, subjectively reported symptoms, such as they describe, respond well to medical consultation alone. 85% of patients referred to a specialist breast clinic with breast pain (mastalgia) do not need active treatment and, of those treated, a clinically useful placebo response occurs in around 19%.1 Active drug treatments used in mastalgia such as danazol, bromocriptine, and evening primrose oil all produce a significantly greater irnprovement than placebo, whereas progestogens do not, and therefore should not be prescribed in mastalgia. I would suggest that if, as Ernst et al state, in patients with varicose veins placebo ointments do produce as good a response as pharmacologically active ones, it is because ointments are not the treatment of choice for symptoms of this condition.

SIR,-I

am

Department of Surgery, University Hospital of South Manchester, West Didsbury, Manchester M20 8LR, UK 1

the analogous sites on the other subtypes, despite sequence similarities at residues other than position 59. There may be other reasons for the rarity of AS and related disorders in certain racial groups. HLA-B27-positive first-degree relatives of B27 AS patients have a 5-16-fold higher incidence of the disease than unrelated B27 subjects,’,’ pointing to another strongly predisposing gene or group of genes. The location and nature of this second AS locus is unknown, but its frequency in certain populations might well explain the rarity of these disorders. A further complication arises when geographically diverse populations are examined because exposure to postulated environmental factors2 may also vary. The notion of a reactive sulphydryl group at the B27 epitope is not necessarily incompatible with the "arthritogenic peptide" model discussed by Hill et al. Reaction with the sulphydryl side chain of Cys67 might be critical in B27 forming an unusually stable or stoichiometrically rigid interaction with a cysteine-containing bacterial peptide,7 or in controlling access to portions of the antigen presentation groove on B27.2

C. A. GATELEY

Inflammation Research Group, London Hospital Medical College

I. L. MACLEAN

J. R. ARCHER M. A. WHELAN

1. El-Zaatari FAK, Sams KC, Taurog JD. In vitro mutagenesis of HLA-B27. Amino add substitutions at position 67 disrupt anti-B27 monoclonal antibody binding in direct relation to the size of the substituted side chain. J Immunol 1990; 144: 1512-17. 2. Benjamin R, Parham P. Guilt by association: HLA-B27 and ankylosing spondylitis. Immunol Today 1990; 11: 137-42. 3. Archer JR, Whelan MA, Badakere SS, MacLean IL, Archer IVJ, Winrow VR. Effect of a free sulphydryl group on expression of HLA-B27. Scand J Rheumatol 1990; 87: 44-50. 4. MacLean IL, Winrow VR, Perrett D, Archer JR. Status of an unpaired thiol group on the HLA-B27 epitope. Clin Exp Immunol 1989; 77: 417-21. 5. Calin A, Marder A, Becks E, Bums T. Genetic differences between B27 positive patients with ankylosing spondylitis and B27 positive healthy controls. Arthritis Rheum 1986; 26: 1460-64 6. van der Linden SM, Valkenburg HA, de Jongh BM, Cats A. The risk of developing ankylosing spondylitis in HLA-B27 positive individuals: a comparison of relatives of spondylitis patients with the general population. Arthritis Rheum 1984; 27: 241-49. 7. Stieglitz H, Fosmire S, Lipsky P Identification of a 2-Md plasmid from Shigella flexnen associated with reactive arthritis. Arthritis Rheum 1989; 32: 937-46.

Gateley CA, Mansel RE. Management ofcyclical breast pain. Br J Hosp Med 1990; 43:

Mitochondrial leucine tRNA mutation in neurological diseases

330-32

HLA-B27 subtypes SjR,—Dr Hill and colleagues (March 16, p 640) used the polymerase chain reaction and dot-blot hybridisation to show that 11 of 18 healthy HLA-B27-positive Gambians had the B*2703 subtype. They postulate that the rarity of ankylosing spondylitis (AS) and other B27-related disorders in African blacks is due to B*2703 conferring a much lower risk than the other subtypes. B*2703 differs from the AS-associated B*2705 subtype by a single Tyr->His substitution at position 59 on the al domain of the molecule. The B27 epitope features an unpaired cysteine residue at aminoacid 67.1 A unique combination of neighbouring aminoacids includes asparagine at position 97 and lysine at position 70,2 whose positively charged side chain may favour the more reactive-Ssulphydryl side chain variant at the adjacent Cys67.3 Sulphydryl reactivity of a proportion of B27 molecules’ suggests that the fine conformation of the epitope will vary according to the oxidative state of this cysteine, the "altered self’ model. Hill et al cite the postulated absence of disease predisposition with B*2703, which shares Cys67, Lys70, and Asn97 with the other B27 subtypes, as evidence against this model. We have two major reservations about this argument. As confmned by their observation that cytotoxic T lymphocytes responding to B*2702 and B*2705 fail to recognise B*2703, the conformation of this region will be different in B*2703. The accessibility and reactive state of Cys67 may be very different from

SIR,-Mutations in mitochondrial DNA (mtDNA) have been several neurological diseases, including chronic progressive external ophthalmoplegia (CPEO), Kearns-Sayre syndrome,! myoclonus epilepsy and ragged-red fibres (MERRF),22 and mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS).3,4 Two independent Japanese groups have reported a point mutation in mitochondrial leucine tRNA (UUR) -namely an A-to-G transition at position 3234-in 26 out of 313 and in all of 44 patients with MELAS. This mutation was also detected in one patient with CPEO (1/29)3 who had a positive family history of mitochondrial disease. To verify the significance of this mutation in other populations, we studied mtDNA from American probands with mitochondrial encephalomyopathies. An mtDNA region bracketing the tRNA leucine gene was synthesised by polymerase chain reaction amplification, from positions 1591 (12S rRNA) to 3650 (ND-1), and was digested with the restriction enzyme ApaI. The 3243 mutation creates an Apal recognition site (GAGCCC to GGGCCC) and the wild-type 2059 base PCR product is cleaved into 1652 base and 407 base products. We detected the 3243 mutation in all of 6 probands with MELAS, in 1 patient with Kearns-Sayre syndrome, and in another with CPEO. Neither the patient with Kearns-Sayre syndrome nor the CPEO patient had a positive family history of mitochondrial disease. None of these 8 patients had a detectable mtDNA deletion by Southern blot analysis or by widely interspaced primer PCR.S In each patient and tissue examined, the 3243 mutation coexisted with found in