Structural changes in the diabetic kidney

3 Structural Changes in the Diabetic Kidney RUTH 0STERBY

The diabetic kidney presents unique features regarding functional and structural aspects over the whole course of diabetes. The tragic result of the se changes is the development of renal failure in a large proportion of long-term diabetics (Andersen et ai, 1983). The late functional deterioration is rooted in structural changes within the glomerular capillary tuft named diabetic glomerulopathy, which is well known and easily recognizable histologically (Thomsen, 1965). It is the end-result of a very long-standing development. Changes begin at the onset of the metabolic disorder and from the very beginning there is an interplay between structure and function. The structure-function relationship in the diabetic kidney at different stages of development works in two ways. The functional consequences of advanced glomerulopathy are obvious and represent one example of structural changes leading to functional abnormalities. The opposite may also be true: functional haemodynamic changes, including increased capillary pressure may cause structural damage both early and late in the course of the disease (Hostetter et ai, 1982). At different stages in the development these relationships as well as their significance are still a matter of debate. The kidney in long-term diabetics may present a multitude of changes some of which are secondary to the glomerulopathy. It is the united effects of the different abnormalities which determine the total function of the kidney and thi s should nor be ' overlooked when in our studies we concentrate on one particular aspect. The glomerulopathy, however, has the dominant role in determining the overall prospects for the patient. In thc present text main emphasis is placed on the very characteristic glomerular changes, and other structural abnormalities in the kidney will only be dealt with very briefly. GLOMERULAR STRUCTURES ON LIGHT AND ELECTRON MICROSCOPY The tools required for the study of glomerular structure are microscopes and stereological methods. Clinics in Endocrinology and Metabolism-Vol. 15, No.4, November 1986

733

734

R.0STERDY

Stereological methods enable us to extract quantitative information from the microscopical pictures (Gundersen, 1980). This text will not deal with the methodology. If correct and unbiased sampling of the two-dimensional pictures is applied, it is possible by simple procedures to obtain data on the three-dimensional tissue under study. This has been very important for the clarification of a large number of detailed questions concerning glomerular structure in diabetes. Observing the glomerular structures with the aid of microscopes we can identify and measure different structural elements, depending on the actual magnification. As an introduction to the results of quantitative structural studies in diabetic glomeruli, a brief review is given of observations made at increasing levels of magnification, at the same time describing changes to be found in diabetes. . Light microscopical inspection is the method used to make the pathoanatomical diagnosis 'diabetic glomerulopathy'. In its moderate or advanced stages this lesion is easily identifiable (Figure 1). The glomerular

•

Figure I. Light micrograph illustrating the diffuse diabetic glomerulopathy in a patient with overt nephropathy (Group VI, Table 1).

STRUcrURAL CHANGES IN THE DIABETIC KIDNEY

735

Figure 2. Light micrograph of a normal glomerulus. Kidney biopsy from one of the cases in Group I (Table 1).

tuft appears more solid with less open space than in normal cases (Figure 2). Thus at light microscopy there is an overall impression of the relative amount of the solid structures. The size of the glomerular profiles can be determined and thereby estimates of glomerular volume are obtainable (0sterby and Gundersen, 1975). Low magnification electron microscopy (EM) (Figure 3) is convenient for comparison with the well-known light microscopical changes. At this level the mesangial regions can be identified: these are the solid spaces in between the capillaries, the capillary walls separating the blood space from the urinary space, thereby constituting the filtration barrier. In the following the term filtration surface (FS) will be used for this peripheral capillary wall. The three different cell types, endothelial, epithelial and mesangial cells , can be identified at this magnification. Figure 4 shows at a higher magnification part of a mesangial region with mesangial cells and mesangial basement membrane-like material (BMLM). For quantitation, these irregular humps of BMLM have to be distinguished strictly from the peripheral basement membrane (PBM). The thickness of the PBM can be estimated at this magnification (Jensen et al, 1979; Hirose et ai, 1982) and the relative volume of the BMLM can be expressed as per cent of mesangial volume: BMLM volume fraction (Vv(BMLMlmes)). Abutting the BM on the external, urinary side are the foot processes of the epithelial cells. Their true width can be determined with a stereological method (Gundersen et aI, 1980).

736

R.0STERDY

Figure 3. Photomontage of a glomerular profile illustrating glomerulopathy as it appears at low magnification electron microscopy. Two profiles of mesangial regions have been delineated. The arrow points to a peripheral capillary wall. For actual measurements of mesangial volume fraction and surface densities about 2400 x magnification is convenient.

Proceeding to higher magnification we may study the fine structural details of cells and BM (Figures 5-7). In glomeruli severely affected by diabetic glomerulopathy several peculiarities in fine structure may be observed as illustrated in the figures. Detailed studies of such changes in fine structure have not as yet been carried out, and any relationship to functional changes of the basement membrane material is not known. Applying very special techniques at this high magnification can give further information on the fine structures which are not observable in ordinary preparations. For instance cytochemical detection of the negatively charged sites which are localized preferentially in the external layer of the BM are under study in diabetic kidney biopsies (Vernier et ai, 1986). In brief, the structural characteristics of the diabetic glomerulopathy are: increase in PBM thickness and expansion of mesangial regions with their BMLM, i.e. increase in the proportion of the tuft that is occupied by mesangial regions. These changes result in the typical appearance at light

STRucrURAL CHANGES IN THE DIABETIC KIDNEY

737

Figure 4. The measurements of PBM thickness and volume fraction of mesangial BI\I-like material arc performed at about 7 000 x magnification. The mesangial region is delineated from the peripheral BM by the straight line. (From a case in Group VI, Table 1.)

Figure 5. At higher magnifications the fine details in BM structure can be studied. In this small segment of a markedly thickened BM from a long-term diabetic, the architecture of the BM material appears normal. C=eapillary lumen. U=urinary space. FP=profile of an epithelial foot process.

738

R.0STERBY

Figure 6. Various structural alterations are somet imes observed in the PBM in diabetic glomerul op athy . In this segment membranous profiles are seen (arrow) as well as an area with more loose structure (*). Their nature and possible significance is unknown .

microscopy (Figure 1) termed the diffuse diabetic glomerulopathy. Sometimes the mesangial BMLM accumulation takes a particular shape giving rise to the so-called Kirnrnelstiel-Wilson nodules. They are seen as circular profiles within the tuft, and their appearance is usually homogeneous and acellular. The nodular type, which is in practical terms specific for diabetes, occurs in only some patients with glomerulopathy, and in these cases only in some of the glomerular profiles (Thomsen, 1965). Apart from the classical diabetic glomerulopathy, structural changes of a completely different nature are also demonstrable in diabetic patients. They consist of glomerular hypertrophy, with enlargement of the capillary tuft and all its constituents. The data that will be presented refer only to patients with insulin dependent (Type I) diabetes, since no exact data are available on glomerulopathy and its development in non-insulin dependent (Type II) patients in whom the time of onset of diabetes is not known . It is thus impossible to draw reasonable conclusions regarding the development of glomerulopathy. It can, however, be stated with certainty that the very characteristic changes of diabetic glomerulopathy develop also in noninsulin dependent diabetics (Thomsen, 1965).

STRUcrURAL CHANGES IN THE DIABETIC KIDNEY

739

m

Figure 7. Section of a mesangial region from a case with advanced glomerulopathy. The Blvl-like material which demonstrates various abnormal structures occupies a great proportion of the region with only sparse islands of mesangial cell cytoplasm (M). C=capillary lumen.

The description of the development of structural changes will follow the chronology in individual diabetic patients so far as it has been elucidated. We can describe certain phases which are to a large extent determined by functional parameters. Much information is still needed from longitudinal studies in individual patients in particular to gain a greater understanding of the pattern of development. In particular we still lack data on those diabetics who do not develop renal functional impairment. GLOMERULAR HYPERTROPHY IN EARLY DIABETES The discovery of glomerular hyperfunction in early diabetes (Mogensen et aI, 1979) led to the demonstration of early diabetic renal and glomerular hypertrophy. The marked elevation in glomerular filtration rate (GFR) present at the clinical onset of diabetes and usually remaining elevated for many years was unexplained. The search for possible structural changes led to a number of positive findings which all corresponded in a relevant way to the functional state. It was found by X-ray analysis, and later ultrasound (Mogensen et aI, 1979; Christiansen et aI, 1981), that the whole kidney is

740

R .0STERBY

larger than in non-diabetics. Light microscopic study of kidney biopsies demonstrated an increase in glomerular size with a glomerular volume 70% larger than in non-diabetics (0sterby and Gundersen , 1975). Finally the total area of the filtration surface was increased by 80% (Kroustrup et al, 1977). So concomitant with the hyperfunction there is a generalized renal hypertrophy, and it is reasonable to assume that the enlargement of the structures is the basis of the hyperfunction. This has gained further support from the demonstration of a close correlation between the level of GFR and the size of the filtration surface (Hirose et al, 1980). However, the cause-effect direction in this relationship cannot be stated with certainty. Both changes are present at the diagnosis so that there are two pos sibilities: either primary altered haemodynamics lead to expansion of the glomerular tuft, or a primary growth of the renal corpuscle with enlargement of the FS gives rise to the increased filtration rate . It is interesting that in thi s acute diabetic state of short duration, BM material has already accumulated in the glomeruli corresponding to the enlargement. Animal experiments have confirmed that such fast changes in the BM volume occur at the onset of the diabetic state (0sterby and Gundersen, 1980). That it is truly an increase in mass, and not for instance merely an osmotic expansion, is seen from the demonstration of increased amounts of RNA, DNA and protein in these kidneys (Seyer-Hansen, 1976). The course of these changes during many years of diabetes remains unclear. The kidney remains large (Ellis et aI, 1985) and the GFR is also elevated for many years, so that the enlargement of the glomeruli and the FS may also remain. It is also uncertain what happens to the glomerular structures during periods of very intensive treatment leading to a decrease in GFR while the kidney size does not change (Christiansen et al, 1981). Animal experiments have shown that the BM accumulation taking place at the induction of diabetes is not reversible within a four week period of metabolic control (Gotzsche et al, 1981). One fundamental unanswered question is whether the functional and/or structural lesions very early in diabetes contribute to the ensuing development of diabetic glomerulopathy (Hostetter et al, 1982; Parving et al , 1982). There are animal experiments which show that hypertrophy is not necessary for the development of glomerulopathy (Brekke et al, 1985); furthermore, the hypertrophy and hyperfunction seen after uninephrectomy are not sufficient to lead to BM thickening (Steffes et al, 1982). Thus the question remains as to whether the altered haemodynamics, in particular -increased capillary pressure, contribute to the development of diabetic glomerulopathy. GLOMERULAR ULTRASTRUCTURE IN THE COURSE OF DIABETES

The clinically silent period The onset of the development of BM thickening most probably starts at the onset of diabetes. Within the initial phase kidney biopsy material has been

STRUcrURAL CHANGES IN THE DIABETIC KIDNEY

741

studied from insulin dependent diabetics identified only by the duration of disease which varied from nought to six years. These patients have normal renal function. At the onset of diabetes the rBM thickness, the volume fraction of mesangial regions and the volume fraction of BMLM are normal (0sterby, 1975). It is an important statement that the structural changes of diabetic glomerulopathy develop only after the onset of the metabolic disorder. Sequential biopsies in some of these cases at the diagnosis and again after two to three years' duration showed that BM thickening is demonstrable after this short period of time, amounting to approximately 15% (Table 1). Diabetics with a duration of about five years demonstrate a thickening of about 25-30% when compared with normal. The volume fraction of mesangial regions did not differ from normal in these patients whereas the content of BMLM relative to the mesangial regions showed an increase in the sequential biopsies. The observation of BM thickening in all of the five patients who were re-biopsied, and in all seven patients biopsied after five years of diabetes suggests that it occurs in almost all diabetics, that it begins at the onset and progresses over many years at a very slow rate. There is unfortunately no information regarding changes after the first five years. Subsequent changes arc only known from biopsy material obtained from selected diabetics defined by their functional status, and represent a different population from that available for study of the initial lesions.

Stages with signs of renal functional impairment preceding overt nephropathy The appearance of permanently increased urinary albumin excretion (UAE) is the first warning that renal function may in the future deteriorate (see Chapter 4). UAE increases gradually over some years into overt proteinuria (Christensen and Mogensen, 1985) and at some stage GFR will start to decline. Structural data are available from patients at different stages in this development. The main findings are seen in Table 1 in which comparisons can be made with controls and early diabetics. Group IV was defined as patients showing UAE of 20-600 ug/min, a higher level than presently used to define incipient nephropathy (Mogensen et ai, 1986) and GFR above 100 mVmin/1.73 m 2 (0sterby et ai, 1984). Some of these patients had an elevated GFR despite a markedly increased UAE. Patients in Groups V and VI with overt nephropathy had a raised UAE but decreased GFR. Group VI is considered separately since all these patients had been in intensive antihypertensive treatment for a number of years (Nyberg et ai, 1984) during which period they showed a stable low level of GFR. The progression of the structural abnormalities characteristic of diabetic glomerulopathy is clearly seen, increasing from the five year group with completely normal function, through incipient to overt nephropathy. Within the. entire group of diabetics all of the structural parameters

742

R.0STERBY

correlated with the duration of diabetes irrespective of the fact that the long-term groups were selected for biopsies on the basis of their functional status. This very selective population sampling may easily blur the relationship. Mauer et al (1984), who studied the same structural parameters, did not find a positive correlation to duration in a series of patients most of whom were candidates for pancreas transplantation. The dependence on duration of diabetes is one of the fundamental characteristics of the various expressions of diabetic microangiopathy (Lundbaek, 1953). However, in order to obtain reliable information on the relationship cross-sectional patient groups should be studied, and such series have not been available for detailed structural investigations. Glomerular occlusion Glomerular occlusion is a phenomenon which is even apparent in diabetics with incipient nephropathy, and becomes increasingly common as functional impairment progresses (Table 1). The estimates of the percentage of glomeruli occluded obtained from biopsy material are not very precise in individual patients. However, this phenomenon is very important in relation to kidney function, since the occluded glomeruli have ceased to function. In relating structure to renal function it is therefore necessary to take this estimate, however vague, into account. Compensatory hypertrophy The structural changes described in the previous section refer only to open functioning glomeruli. Only in these can the individual structural elements be identified. The changes in these glomeruli are not only due to advanced glomerulopathy but also result from the closure of other nephrons. Compensatory hypertrophy develops in all probability when a certain proportion of glomeruli have been occluded (Gundersen and 0sterby, 1977). The estimates of glomerular size shown in Table 1 demonstrate this additional hypertrophy in the advanced stages. By this mechanism a certain gain in capillary surface is achieved: in Group VI it was found that total capillary surface per open glomerulus is retained at the normal level despite the marked increase in mesangial volume fraction (0sterby et aI, 1987). The fact that the glomerular tuft is markedly enlarged also means that the structural changes are much more pronounced than it appears from the relative figures. Thus the total volume of mesangial regions in open glomeruli in Group VI is increased by about 300% and total BM volume (PBM + BMLM) by 600% compared with normal. Distribution pattern of diabetic glomerulopathy A study of the distribution of the diabetic glomerulopathy within the cortex was performed in order to look for possible pathogenetic mechanisms involved in its development (Herlyck et ai, 1986). The material was from autopsy kidney specimens from long-term insulin dependent diabetics.

~

;l:l

c Q c;l:l :> L'

o

:;

Table I. Structural data on diabet ics at different stages of the disease. No.

PBM thickne ss (nm )

mes/tuft (%)

BMLMlm es (%)

Glom erular occlusion ('Yo)

'Glomerular volume (f.lm 3 x 10-")

Non-di abetics

5

0

7

0

8

45 30-5 0 53 49-63 58 45-67 61 55-6 8

0.83 0.50-1.13 1.41 0.79- 2.05

IDD s after 5 years

33 29-4 0 34 29-42 34 30-4 0 41 34-52 51 40-67 57 40-75

49 41-52

IDD s at onset

308 280-350 309 270-3 35 414 370-450 527 380-6 36 668 531- 759 647 495-948

Group

II III IV

IDDs with raised urinary album in excretion but normal GFR V IDDs with raised urinary album in excretion and decreased GFR VI Nephr opathy patients on antihypertensive treatment

9 12 12

59 51- 69

z o rn Vl

Z :l

tn

o

:>t: :Q

0

()

17" 0-27 16

3-4 9

39 24-6 7

2. 15" 1.83-2.64 2.17 1.17- 2.79

co z

rn -<:

2.66 1.74-3.94

IDD = Insulin dependent diabetics (Type I) "n = 3

-...J

.j:>. \;J

744

R.0STERI3Y

It was found that the severity of involvement of the open glomeruli as well as the glomerular occlusion occurs to the same extent in superficial and deep zones of the cortex. This indicates that none of the functional and structural differences that exist between these zones plays any major pathogenetic role. The glomerular occlusion, on the other hand, showed clustering and a tendency to occur in stripes perpendicular to the kidney surface. One explanation could be that the occlusion of one tuft has consequences on the immediate surroundings leading to glomerular occlusion in the observed pattern.

RELATIONSHIP BETWEEN STRUCTURE AND FUNCTION Glomerular structure and GFR The very close correlation in early diabetes between enlargement of the filtration surface (FS) and GFR has been described above. Also in the more advanced stages, when a decline in GFR sets in, a close correlation obtains between the structural and functional parameters. Many glomeruli arc then totally occluded while others are in the process of occlusion. In all of the remaining open glomeruli the changes of diabetic glornerulopathy are present to a varying degree. The relevant structural parameter which correlates with the GFR is the total area of FS in the kidney. When biopsy material is examined the FS per nephron has to be considered. This is obtained by determining the percentage of nephrons that are occluded, as well as the remaining surface in the open ones. Since there is not a constant relationship between these two parameters (percentage of glomeruli occluded and the severity of glomerulopathy in the open tufts) both must be taken into account to give complete information relevant to kidney function. In the biopsy material presented in Table 1 (Groups IV, V and VI) it was found that the level of GFR correlates closely to the estimates of FS per nephron (r = 0.75, 2P < 10- 4 ) : the loss of filtration surface due to the progressing glomerulopathy provides a straightforward explanation for the deterioration in renal function. The compensatory hypertrophy of the least affected glomeruli (Gundersen and 0sterby, 1977) leads to preservation of FS for a time, but finally the advancing glomerulopathy leads to end-stage renal failure. An example of the relationship between GFR and other structural parameters is shown in Figure 8 in which the patients of Group VI are indicated separately. For the whole group the decrease in GFR correlates with increasing BM thickness (2P = 0.01). The patients in Group VI with advanced nephropathy who have been on antihypertensive treatment for a period of years seem to follow their own course. There is no explanation for this observation. It has been shown that intensive antihypertensive treatment leads to a slowing in the decline of GFR (Mogensen, 1982; Parving et al, 1983; Nyberg et ai, 1985). At present nothing is known of the mode of action of this treatment or whether it affects the glomerular structures.

745

STRUcrURAL CHANGES IN THE DIABETIC KIDNEY

GFR rnl/min/1.73m 2 0

150

0

0 0 0

0

0 0

0 0

100

0 0

0 0

0 0

00 0 0

50

•

•• •

, •

• •

0

•

• 0

•

o- t - - - - - - - - , - - - - - - - - - - - - - - , o

500

1000 nm

GLOMERULAR BASEMENT MEMBRANE THICKNESS Figure 8. Relationship between BM thickness and GFR. All diabetic patients had elevated urinary albumin excretion. Cases indicated with • had been on an antihypertensive treatment for a period of years.

Glomerular structures and urinary albumin excretion Contrary to the above structure-function relationships, the structural basis of increased albumin excretion is obscure. Increased leakiness of the glomerular filter in diabetics occurs in two different forms. The albuminuria during periods of poor control in the early stages may have mechanisms that are different from those leading to the microalbuminuria of incipient nephropathy. Reversibility differentiates the two. In the first instance, it is assumed that reversible structural changes in the glomerular capillaries cause the increased passage of albumin. One possible mechanism may result from metabolically determined changes in the negative charges of the PBM. Most of these are associated with the heparan sulphate moieties of the BM (Vernier et al, 1983) which seem to have a faster turnover than the other BM constituents. However, experimental proof for this mechanism is still lacking. Even the non-reversible microalbuminuria may be linked to defects in the negative charges (Viberti et al, 1983a; Deckert et al, 1984). Cytochemical studies of this component of the BM are in progress, and preliminary results in biopsies from diabetics (Vernier et al, 1986) indicate a relationship between increasing UAE and progressive loss of charges. Further studies will appear in the near future. The search for a relationship between UAE and structural parameters has led to some positive correlations: Mauer et al (1984) demonstrated that the level of UAE correlated with the volume fraction of mesangial regions but not with BM thickness. The series presented in this chapter (Table 1)

746

R .0STERBY

demonstrates a significant correlation between UAE and the structural parameters listed in the table. Again, Group VI (patients treated for hypertension) when considered separately failed to show such correlations 0sterby et al, 1987). Since a priori it is known that UAE will gradually increase as the disease progresses (Viberti 1983a; Christensen and Mogensen , 1985) and that the structural lesions also progress, this correlation does not imply a cause-effect relationship. In the series of patients with increased UAE, glomerular occlusion was demonstrated although in some cases this only occurred in a small percentage. Thus it appears that glomerular closure develops concomitantly with the increase in UAE. It is possible that the occlusion process is involved in the mechanism (Mogensen and 0sterby, 1986). Albumin escape could perhaps originate in tufts that are occluding and therefore are exposed to very particular haemodynamic forces, or from those tufts in the neighbourhood that have to handle the local flow changes due to occlusion. Epithelial foot processes (FP) may be involved in the development of albuminuria. Widening of FP has been documented in diabetic glomerulopathy. Mauer (personal communication, 1986) found a correlation with UAE, whereas we found that the degree of widening is independent of the level of UAE (0sterby et al, 1986). This point presently remains open. Changes in the fine structural architecture of the BM material might be related to increased leakiness. In severely affected glomeruli a loosening of the structure is occasionally observed (Figure 6). Such localized areas, constituting about 10% of the capillary walls, could well be the site of albumin leakage (Myers, 1981). It is clear from this presentation that the mechanism of the albuminuria in diabetic nephropathy remains hypothetical. There is still much to learn about albuminuria in diabetics. It is most important to clarify the structural details at the time of onset of microalbuminuria: studies of diabetics without microalbuminuria, and of comparable diabetics with incipient nephropathy need to be undertaken in order to answer the question regarding the structural basis of the onset of microalbuminuria . Mode of structure-function interaction Late onset permanent microalbuminuria in diabetics is undoubtedly closely related to the glomerular structural damage regardless of the mechanism. It has been suggested, however, that the primary change is in leakiness which could in turn affect the structures: abnormal amounts of proteins passing the capillary tuft to the urinary space have been assumed to trigger the glomerular cells to produce increased amounts of BM (Williamson and Kilo, 1977). However, it is clear that incipient nephropathy appears only when rather advanced lesions are present in the glomerular capillaries. It is therefore most reasonable to consider structural changes as the cause of microalbuminuria. The development of BM abnormalities and mesangial expansion over a long period before this state is reached has led some to deny any relationship between the structural changes and the glomerular function in

STRUcrURAL CHANGES IN THE DIABETIC KIDNEY

747

diabetes (Deckert et aI, 1984). It is, however, to be expected that only when the pathological changes have reached a certain severity will they lead to clinically detectable signs. It has been observed in several studies that the appearance of microalbuminuria is highly predictive of later development of overt nephropathy (Chapter 4). Whether the increased permeability at this stage is contributory to the further development of the established glomerulopathy or whether it simply reflects structural damage so advanced that it cannot be arrested, is unknown. The interrelationship between haemodynamic parameters and the glomerular structures in the early hypertrophy/hyperfunction state is of great interest, but it is still indeterminable which is the primary event. Glomerular hypertension occurs both early on in hyperfiltering diabetics as well as late on in remnant nephrons when some are occluded, but it is uncertain whether this accelerates the renal damage. On the other hand, it is well established that once the diabetic glomerulopathy has led to a significant decrease in total filtration surface there is a resulting decrease in GFR. The interaction between blood pressure and renal structures is also. unclear. Since hypertension usually appears only after the onset of microalbuminuria it is most likely due to the renal damage. Once blood pressure is increased it accelerates the course of diabetic nephropathy. The mechanisms at the structural level are not understood. OTHER PATHOLOGICAL LESIONS IN THE DIABETIC KIDNEY It is the development of diabetic glomerulopathy which determines the risk of developing nephropathy and end-stage renal failure. However, in the diabetic kidney a spectrum of other changes may also be present. Thus, other renal diseases occur in diabetics with the same probability as in the general population. They include other glomerulopathies which on rare occasions give rise to diagnostic problems. There arc also other kidney diseases to which diabetics are predisposed. Among them are primarily infections. Pyelonephritis with significant bacteriuria, ascending interstitial inflammation, and even papillary necrosis occur quite often in diabetics (Ditscherlein, 1969). Involvement of the juxta-glomerular arterioles is an almost constant finding in diabetic kidneys with glomerulopathy and is to be considered as part of the microangiopathy. The picture is very characteristic, and the severity of arteriolar changes correlates with the severity of the glomerulopathy (Thomsen, 1965). Undoubtedly an interaction between arteriolar and glomerular changes takes place in both directions. Severe arteriolar hyalinosis could well playa role in glomerular occlusion. The renal arteries frequently demonstrate the morphological changes of arteriosclerosis in long-term diabetics (Ditscherlein, 1969). This 'nephrosclerosis' may be important in non-insulin dependent diabetics. The influence of nephrosclerosis on renal function is not known. The

748

R.0STERBY

observation that glomerular occlusion tends to occur in stripes perpendicular to the kidney surface (Horlyck et ai, 1986) may point to a possible role of the interlobular arteries in the occlusion process. The nephrosclerotic lesions may be considered as part of the generalized long-term diabetic macroangiopathy. Perhaps increased blood flow over several years, or the slightly elevated blood pressure which occurs early on, may accelerate the development of the arterial lesions. The kidneys in long-term diabetics with nephropathy remain large (Ellis et ai, 1985) in contrast to the contracted kidneys seen in non-diabetics with advanced nephrosclerosis. Finally, advanced glomerulopathy naturally leads to secondary changes such as interstitial fibrosis, perhaps with cellular infiltrates and degenerative changes in the distal nephron. It is not usually possible to identify to what extent the changes are merely secondary to the glomerulopathy, or whether there is an independent expansion of the interstitium, with tubular degeneration and BM thickening (Mauer et ai, 1984). In the long-term diabetic kidney several of these changes will be present at the same time and it is the unity of these heterogeneous changes that determine the functional consequences. PATHOGENESIS AND PREVENTION The fundamental facts regarding the development of nephropathy in long-term diabetics are reasonably well established. Detailed knowledge of mechanisms at cellular level or the influence of specific abnormalities such as hyperfunction or moderate hypertension on development of nephropathy is not yet available. Underlying all these changes is of course the diabetic metabolic disorder without which none of them would occur. An effective treatment should therefore be able to prevent all of the functional and structural changes. One vital question which needs to be answered is at what point must intensive treatment be instituted in order to prevent the disease. Once microalbuminuria appears it is already too late (Viberti et ai, 1983b; Feldt-Fasmussen, 1986) which means that other than' functional parameters are needed to examine the effect of intervention. Whether the structural parameters described in this chapter, particularly the BM thickness, would be of value during the silent stage to predict the future course needs further investigation. Longitudinal studies in this clinically silent phase are most urgently needed as Mauer et al (1984) have advocated. The rate of progression varies considerably in individual patients. The appearance of clinical signs occurs at any time from about six to some 30 years' duration. Better understanding of the conditions which are responsible for this diversity in progression rate would enhance our knowledge of the pathogenesis of diabetic nephropathy. The reasons why some diabetics do not develop nephropathy are also obscure: do they just progress very slowly, or are they completely protected by unknown factors? Future studies along these lines should improve our understanding of diabetic nephropathy.

STRUcrURAL CHANGES IN TilE DIABETIC KIDNEY

749

SUl\1l\1ARY

Diabetic glomerulopathy is characterized by a very slow development of basement membrane (BM) accumulation, manifested as thickening of the peripheral BM and increased volume of the mesangial BM-like material (BMLM) with mesangial expansion. The initiation of the process is probably at the onset of diabetes since the BM thickening is detectable after a few years. The BM accumulations at the two sites (PBM and BMLM) in the glomerular tuft are considered as two different expressions of a fundamental BM abnormality. The two locations present different conditions for quantitation, may have a different biochemical make-up, and immediate functional implications of the abnormalities may differ as well. In the long run, however, the two in concert lead to the ultimate solidification of the glomerular tuft with loss of capillary surface. The end-stage is glomerular closure, with elimination of glomerular function. A very close correlation has been found between the total remnant surface area of the glomerular capillaries and the level of GFR. Along with the classical changes of the diabetic glomerulopathy, changes in glomerular size are detectable. In early diabetes during the stages of glomerular hyperfunction, hypertrophy develops acutely at the onset of diabetes, leading to an increase in capillary surface corresponding to the increase in filtration rate. In the advanced stages when glomerular closure involves a proportion of the nephrons compensatory hypertrophy develops, thereby probably helping to preserve capillary surface for a period of time. The exact mechanisms that may influence these developments are not known, but underlying them all are the metabolic abnormalities of diabetes. Acknowledgements My sincere gratitude to all those who have taken part in obtaining data presented in this chapter. The biopsy series were made available by excellent collaboration with G. Gregersen. C.E. Mogensen. Aarhus. G. Nyberg. Gothenburg. and H.-H. Parving, Copenhagen. My sincere thanks go to all of the skilful technicians and all co-workers who have taken part. I am most grateful for very valuable assistance in preparing the present manuscript by Anne Holrnehave, Anette Larsen. Albert Meyer and Lone Nielsen.

REFERENCES Andersen AR. Christiansen JS. Andersen JK. Kreiner S & Deckert T (1983) Diabetic nephropathy in type I (insulin-dependent) diabetes. An epidemiological study. Diabetologia 25: 496-501. Brekke lB. Gundersen lUG & 0sterby R (1985) Thickening of glomerular basement membrane in rats with severe diabetes without kidney hypertrophy. Diabetic Nephropathy 4: 19-22. Christensen CK & Mogensen CE (191'5)The course of incipient diabetic nephropathy: studies of albumin excretion and blood pressure. Diabetic Medicine 2: 97-102. Christiansen JS. Gamrnelgaard J. Frandsen M & Parving H-H (1981) Increased kidney size. glomerular filtration rate and renal plasma flow in short-term insulin-dependent diabetics. Diabetologia 20: 451-456.

750

R.0STERBY

Deckert T, Feldt-Rasmussen B, Mathiesen E & Baker L (1984) Pathogenesis of incipient nephropathy: a hypothesis. Diabetic Nephropathy 3: 83-88. Ditscherlein G (1969) Nierenviinderungen bei Diabetikern, Jena: Gustav Fischer Verlag. 255 pp. Ellis EN, Steffes MW, Goetz Fe, Sutherland DER & Mauer SM (1985) Relationship of renal size to nephropathy in Type I (insulin-dependent) diabetes. Diabetologia 28: 12-15. Feldt-Rasmussen, D., Mathieson ER & Deckert T (1986) No effect on kidney function after 18 months of strict metabolic control in insulin-dependent diabetic patients with incipient nephropathy. Diabetic Nephropathy 5: 42. Gotzsche, 0, Gundersen HJG & 0sterby R (1981) Irreversibility of glomerular basement membrane accumulation despite reversibility of renal hypertrophy with islet transplantation in early experimental diabetes. Diabetes 30: 481-485. Gundersen HJG (1980) Stereology. Or how figures for spatial shape and content are obtained by observation of structures in section. Microscopica Acta 83: 409-426. Gundersen HJG & 0sterby R (1977) Glomerular size and structure in diabetes mellitus. II. Late abnormalities. Diabetologia 13: 43-48. Gundersen HJG, Seefeldt T & 0sterby R (1980) Glomerular epithelial foot processes in normal man and rats: distribution of true width and its intra- and interindividual variation. Cell and Tissue Research 205: 147-155. Hirose K, Tsuchida H, 0sterby R & Gundersen HJG (1980) A strong correlation between glomerular filtration rate and filtration surface in diabetic kidney hyperfunction. Laboratory Investigation 43: 434-437. Hirose K, 0sterby R, Nozawa & Gundersen HJG (1982) Development of glomerular lesions in experimental long-term diabetes in the rat. Kidney International 21: 689-695. Horlyck A, Gundersen HJG & 0sterby R (1986) The cortical distribution pattern of diabetic glomerulopathy. Diabetologia 29: 146-150. Hostetter TH, Rennke HG & Brenner DM (1982) The case for intrarenal hypertension in the initiation and progression of diabetic and other glomerulopathies. American Journal of Medicine 72: 375-380. Jensen EB, Gundersen HJG & 0sterby R (1979) Determination of membrane thickness distribution from orthogonal intercepts. Journal of Microscopy 115: 19-33. Kroustrup JP, Gundersen HJG & 0sterby R (1977) Glomerular size and structure in diabetes mellitus. III. Early enlargement of the capillary surface. Diabetologia 13: 207-210. Lundbrek K (1953) Long-term Diabetes. Copenhagen: Munksgaard, Mauer SM, Steffes MW, Ellis EN et al (1984) Structural-functional relationship in diabetic nephropathy. Journal of Clinical Investigation 74: 1l43-1155. Mauer SM, Steffes MW, Ellis EN & Brown DM (1984) Can the insulin-dependent diabetic patient be managed without kidney biopsy? In Robinson RR (ed.) Nephrology, vol. I, Proceedings of the IXth International Congress of Nephrology, pp 1103-1108. New York: Springer-Verlag. Myers BD (1981) Glomerular barrier function in advanced diabetic nephropathy. Acta Endocrinologica Supplementum 242: 36-38. Mogensen CE (1982) Long-term antihypertensive treatment inhibiting progression of diabetic nephropathy. British Medical Journal 285: 685-688. Mogensen CE & 0sterby R (1986) Structural and functional alterations in the diabetic kidney. In Frontiers in Diabetes. Proceedings from the 2nd International Diabetes Conference. Basel: S Karger. Mogensen CE, 0sterby R & Gundersen lUG (1979) Early functional and morphologic vascular renal consequences of the diabetic state. Diabetologia 17: 71-76. Mogensen CE, Chachati A, Christensen CK et al (1986) Microalbuminuria. An early marker of renal involvement in diabetes. Uremia Investigation (in press). Nyberg G, Larsson 0, Westberg NG et al (1984) A platelet aggregation inhibitorticlopidine-in diabetic nephropathy. Clinical Nephrology 21: 184-187. Nyberg G, Blohme G, Bj6rck S & Norden G (1985) Progression of diabetic nephropathy in relation to plasma glucose control and blood pressure control. Diabetic Nephropathy 4: 53-54.

STRUcrURAL CHANGES IN THE DIABETIC KIDNEY

751

0sterby R (1975) Early phases in the development of diabetic glomerulopathy. A quantitative electron microscopic study. Acta Medica Scandinavica Supplementum 57-1: 1-82. 0sterby R & Gundersen lUG (1975) Glomerular size and structure in diabetes mellitus. I. Early abnormalities. Diabetologia II: 225-229. 0sterby R & Gundersen lUG (1980) Fast accumulation of basement membrane material and the rate of morphological changes in acute experimental diabetic glomerular hypertrophy. Diabetologia 18: 493-500. 0sterby R. Andersen AR. Gundersen HJG et al (1984) Quantitative studies of glomerular ultrastructure in juvenile diabetics with incipient nephropathy. Diabetic Nephropathy 3: 95-100. 0sterby R. Andersen AR. Gregersen G et al (1986) Quantitative structural data on the development of diabetic glomerulopathy. Diabetic Nephropathy 5: 10-11. 0sterby R, Gundersen lUG, Nyberg G & Aurell M (1987) Advanced diabetic glomerulopathy--quantitative structural characterization of the non-occluded glomeruli. Submitted for publication. Parving HH. Vibcrti GC, Keen H et al (1982) Hemodynamic factors in the genesis of diabetic microangiopathy. Metabolism 32: 9-13--9-19. Parving HH, Andersen AR, Smidt UM & Svendsen PA (1983) Early aggressive antihypertensive treatment reduces rate of decline in kidney function in diabetic nephropathy. Lancet i: 1175-1179. Seyer-Hausen K (1976) Renal hypertrophy in streptozotocin-diabetic rats. Clinical Science and Molecular Medicine 51: 551-555. Steffes MW. Vernier RL, Brown DM, Basgen JM & Mauer SM (1982) Diabetic' glomerulopathy in the uninephrcctomized rats resists amelioration following islet transplantation. Diabetologia 23: 347-353. Thomsen AC (1965) The Kidney in Diabetes Mellitus, Copenhagen: Munksgaard. 337 pp. Vernier RL, Klein DJ, Sisson SP et al (1983) Heparan sulfate-rich anionic sites in the human glomerular basement membrane. New England Journal of Medicine 309: 1001-1009. Vernier RL, Sisson-Ross S & Mauer SM (1986) Cyto-chemical studies of the anionic charges in the kidney in Type I diabetes mellitus. Diabetic Nephropathy 5: 15-18. Viberti GC, Mackintosh D & Keen H (1983a) Determinants of the penetration of proteins through the glomerular barrier in insulin-dependent diabetes mellitus. Diabetes 32(supplement 2): 92-95. Viberti GC, Bilous RW, Mackintosh D et al (1983b) Long term correction of hyperglycaemia and progression of renal failure in insulin dependent diabetes. British Medical Journal 286: 598-602. Williamson JR & Kilo C (1977) Current status of capillary basement-membrane disease in diabetes mellitus. Diabetes 26: 65-73.