Peripheral Glomerular Basement Membrane Research Articles

Serial renal biopsies in normo- and microalbuminuric patients with type 2 diabetes demonstrate that loss of renal function is associated with a reduction in glomerular filtration surface secondary to mesangial expansion

AimsThe aim of the present study was to explore the relationship between changes in renal structure in patients with type 2 diabetes at an early stage of diabetic nephropathy using serial renal biopsies, and change in renal function. MethodsThe study population comprised 10 patients with type 2 diabetes with normo- or microalbuminuria at baseline. Light and electron microscopy-based morphometric analyses were performed to quantitatively evaluate glomerular and interstitial structural changes. Urinary albumin excretion (UAE) and glomerular filtration rate (GFR) were measured annually. A second renal biopsy was performed after a mean of 6.1 ± 2.4 years of follow-up. ResultsUAE, GFR, blood pressure and severity of diabetic retinopathy (DR) did not change between the baseline and follow-up. The annual decrease in the surface density of the peripheral glomerular basement membrane (GBM) (Sv[PGBM/glom]) was correlated with the rate of loss of GFR during the six-year follow-up period. The annual change in the Sv(PGBM/glom) was negatively correlated with the change in mesangial volume fraction. ConclusionsDecreases in the GFR in patients with type 2 diabetes with normo- or microalbuminuria at baseline were associated with a decreased glomerular filtration surface, as a result of mesangial expansion during a mean six years of observation. These findings confirm ongoing pathological progression of glomerulopathy despite no significant change in albuminuria or retinopathy status.

Role of Kidney Biopsies for Biomarker Discovery in Diabetic Kidney Disease.

Although estimated glomerular filtration rate and albuminuria are well-established biomarkers of diabetic kidney disease (DKD), additional biomarkers are needed, especially for the early stages of the disease when both albuminuria and estimated glomerular filtration rate may still be in the normal range and are less helpful for identifying those at risk of progression. Traditional biomarker studies for early DKD are challenging because of a lack of good early clinical end points, and most rely on changes in existing imprecise biomarkers to assess the value of new biomarkers. There are well-characterized changes in kidney structure, however, that are highly correlated with kidney function, always precede the clinical findings of DKD and, at preclinical stages, predict DKD progression. These structural parameters may thus serve as clinically useful end points for identifying new biomarkers of early DKD. In addition, investigators are analyzing tissue transcriptomic data to identify pathways involved in early DKD which may have associated candidate biomarkers measurable in blood or urine, and differentially expressed microRNAs and epigenetic modifications in kidney tissue are beginning to yield important observations which may be useful in identifying new clinically useful biomarkers. This review examines the emerging literature on the use of kidney tissue in biomarker discovery in DKD.

Urinary IgG4 and Smad1 Are Specific Biomarkers for Renal Structural and Functional Changes in Early Stages of Diabetic Nephropathy.

Diabetic nephropathy (DN) is the major cause of end-stage kidney disease, but early biomarkers of DN risk are limited. Herein we examine urinary IgG4 and Smad1 as additional early DN biomarkers. We recruited 815 patients with type 2 diabetes; 554 patients fulfilled the criteria of an estimated glomerular filtration rate (eGFR) >60 mL/min and no macroalbuminuria at baseline, with follow-up for 5 years. Patients without macroalbuminuria were also recruited for renal biopsies. Urinary IgG4 and Smad1 were determined by enzyme-linked immunoassays using specific antibodies. The specificity, sensitivity, and reproducibility were confirmed for each assay. Increased urinary IgG4 was significantly associated with lower eGFR. The level of urinary IgG4 also significantly correlated with surface density of peripheral glomerular basement membrane (Sv PGBM/Glom), whereas Smad1 was associated with the degree of mesangial expansion-both classic pathological findings in DN. Baseline eGFR did not differ between any groups; however, increases in both urinary IgG4 and Smad1 levels at baseline significantly predicted later development of eGFR decline in patients without macroalbuminuria. These data suggest that urinary IgG4 and Smad1 at relatively early stages of DN reflect underlying DN lesions and are relevant to later clinical outcomes.

Plasma bradykinin and early diabetic nephropathy lesions in type 1 diabetes mellitus.

ObjectiveTo examine the association of bradykinin and related peptides with the development of diabetic nephropathy lesions in 243 participants with type 1 diabetes (T1D) from the Renin-Angiotensin System Study who, at baseline, were normoalbuminuric, normotensive and had normal or increased glomerular filtration rate (GFR).DesignPlasma concentrations of bradykinin and related peptides were measured at baseline by quantitative mass spectrometry. All participants were randomly assigned at baseline to receive placebo, enalapril or losartan during the 5 years between kidney biopsies. Kidney morphometric data were available from kidney biopsies at baseline and after 5 years. Relationships of peptides with changes in morphometric variables were assessed using multiple linear regression after adjustment for age, sex, diabetes duration, HbA1c, mean arterial pressure, treatment assignment and, for longitudinal analyses, baseline structure.ResultsBaseline median albumin excretion rate of study participants was 5.0 μg/min, and mean GFR was 128 mL/min/1.73 m2. After multivariable adjustment, higher plasma concentration of bradykinin (1–8) was associated with greater glomerular volume (partial r = 0.191, P = 0.019) and total filtration surface area (partial r = 0.211, P = 0.010), and higher bradykinin (1–7) and hyp3-bradykinin (1–7) were associated with lower cortical interstitial fractional volume (partial r = -0.189, P = 0.011; partial r = -0.164, P = 0.027 respectively). In longitudinal analyses, higher bradykinin was associated with preservation of surface density of the peripheral glomerular basement membrane (partial r = 0.162, P = 0.013), and for participants randomized to losartan, higher hyp3-bradykinin (1–8) was associated with more limited increase in cortical interstitial fractional volume (partial r = -0.291, P = 0.033).ConclusionsHigher plasma bradykinin and related peptide concentrations measured before clinical onset of diabetic nephropathy in persons with T1D were associated with preservation of glomerular structures, suggesting that elevations of these kinin concentrations may reflect adaptive responses to early renal structural changes in diabetic nephropathy.

Role of Different Molecular Pathways in the Development of Diabetes-Induced Nephropathy

Diabetes mellitus is heterogeneous primary disorder of carbohydrate metabolism with multiple etiologic factors that generally involves absolute or relative insulin deficiency or insulin resistance or both, which results in hyperglycemia. According to WHO projection; it will be the single largest non-communicable disease worldwide by the year 2025 with the largest diabetic population in India. India leads the world with largest diabetic population thus, being termed the ‘‘Diabetes Capital of the World’’. However, the prevalence of diabetes is consistently increasing, but still an effective treatment is lacking for the management of this epidemic. The uncontrolled and chronic diabetes mellitus often leads to cardiomyopathy, macrovascular complications and microvascular complications that include retinopathy, neuropathy and nephropathy. Diabetic Nephropathy (DN) is mainly characterized by decreased Glomerular Filteration Rate (GFR), excessive deposition of extracellular matrix proteins, thickening of the peripheral glomerular basement membrane, glomerular hypertrophy, tubulointerstitial fibrosis, increased excretion of albumin and decreased creatinine clearance. Formation of Advanced Glycation End-Products (AGEs), activation of Protein Kinase C (PKC), c-Jun N-terminal kinase (JNK), Mitogen Activated Protein Kinases (MAPKs), oxidative stress are the possible molecular mechanisms of DN. Despite many molecular mechanisms, the pathophysiology of DN is not clearly understood and its management is unsatisfactory.

Glomerulopathy in spontaneously obese rhesus monkeys with type 2 diabetes: a stereological study.

Animal models could provide insights into the diabetic nephropathy pathogenesis; however, available rodent models do not mirror the heterogeneity of lesions in type 2 diabetic patients, and do not progress to end-stage renal disease. Previous studies showed that spontaneously obese type 2 diabetic rhesus monkeys develop many of the features of human diabetic glomerulopathy, and may progress to end-stage renal disease. Here, in order to further characterize diabetic glomerulopathy in this model, we used electron microscopic stereology. Renal biopsies from 17 diabetic, 17 pre-diabetic/metabolic syndrome and 11 non-diabetic monkeys were studied. Fractional volumes of mesangium [Vv(Mes/glom)], mesangial matrix [Vv(MM/glom)] and mesangial cells [Vv(MC/glom)], glomerular basement membrane width and peripheral glomerular basement membrane surface density per glomerulus [Sv(PGBM/glom)] were estimated. Glomerular filtration and albumin excretion rates were measured in a limited number of animals. Glomerular structural and biochemical/metabolic data were compared among the groups. Compared to non-diabetic monkeys, diabetic rhesus monkeys showed classic diabetic nephropathy changes, including glomerular basement membrane thickening (p = 0.001), increased fractional volumes of mesangium (p = 0.02), and reduced peripheral glomerular basement membrane surface density per glomerulus (p = 0.03) compared to non-diabetic monkeys. Increased fractional volumes of mesangium was primarily due to increased mesangial matrix (p = 0.03). Glomerular structural parameter inter-relationships in diabetic monkeys mirrored those of human diabetic glomerulopathy. Albumin excretion rate was greater (p = 0.03) in diabetic vs. non-diabetic monkeys. There was trend for a positive correlation between albumin excretion rate and fractional volumes of mesangium. This rhesus primate model shares many features of human diabetic glomerulopathy. Mesangial expansion in this model, similar to human diabetic nephropathy and different from available rodent models of the disease, is primarily due to increased mesangial matrix.

Membranoproliferative Glomerulonephritis Type III in a Simultaneous Infection of Leishmania Infantum and Dirofilaria Immitis in a Dog

In this report a 9-year-old female German Shepherd dog with a membranoproliferative glomerulonephritis (MPGN) type III associated with concomitant infection of Dirofilaria immitis and Leishmania infantum is presented. Light microscopic evaluation of kidney revealed a diffuse hypercellularity and thickening of glomerular basement membrane. Heavy and coarse granular complement C(3) deposition and a weaker positive reaction to immunoglobulin G were present along peripheral glomerular basement membrane and in the mesangium in the immunofluorescent study. Transmission electron microscopy revealed deposits in the mesangium, subendothelium, and subepithelium. These lesions are compatible with membranoproliferative glomerulonephritis type III in humans.

Podocyte Lesions in Patients With Obesity-Related Glomerulopathy

Obesity-related glomerulopathy is an important complication of obesity that shares some pathophysiological factors relevant to renal damage with diabetic nephropathy. It was recognized that podocyte lesions lead to proteinuria and glomerulosclerosis in patients with some proteinuric glomerular diseases, especially diabetic nephropathy. However, podocyte changes in patients with obesity-related glomerulopathy and the underlying mechanisms are unclear. Glomerular volume, podocyte number, and foot-process width were evaluated in 46 patients with biopsy-proven obesity-related glomerulopathy. Renal tissue from 10 kidney donors served as controls. Glomerular volume was enlarged markedly in patients with obesity-related glomerulopathy (P < 0.001), with a corresponding decrease in podocyte density (P < 0.001) compared with controls. Foot-process width on the peripheral glomerular basement membrane was increased significantly, whereas mesangial volume fraction did not differ between patients with obesity-related glomerulopathy and controls. Degree of proteinuria was associated strongly with decreased podocyte density (P = 0.001), increased foot-process width (P = 0.001), and decreased podocyte number (P = 0.002). In addition, the endogenous creatinine clearance rate correlated with decreased podocyte number (P = 0.008). Interestingly, podocyte number and density correlated significantly with abnormalities in fasting glucose (P < 0.001) and insulin levels (P < 0.001) and Homeostasis Model Assessment of Insulin Resistance (P < 0.001). Decreased podocyte density and number were observed in patients with obesity-related glomerulopathy, and changes in podocytes correlated with degree of proteinuria and renal function impairment in these patients. In addition to the enlargement in glomerular volume with consequential mechanical stretch, metabolic disorders also may contribute to the development of podocyte lesions in patients with obesity-related glomerulopathy.

Cellular basis of diabetic nephropathy: IV Antioxidant enzyme mRNA expression levels in skin fibroblasts of type 1 diabetic sibling pairs

Blunted cultured skin fibroblast (SF) antioxidant enzyme responses to hyperglycaemia are associated with diabetic nephropathy risk. The present study explores whether this association is, at least in part, genetically determined. We measured glomerular structure and SF mRNA expression for catalase and glutathione peroxidase in 21 sibling pairs concordant for type 1 diabetes. All patients had four or more (mean 21.5) years of diabetes and glomerular filtration rate>40 ml/min/1.73 m2. Thirty-four patients were normoalbuminuric, four were microalbuminuric, three were proteinuric and one was not classifiable. Heritability of patient characteristics was assessed by intra-class correlation and by a genetic variance component model. Mesangial fractional volume, mesangial matrix fractional volume, glomerular basement membrane width and surface density of peripheral glomerular basement membrane per glomerulus were significantly correlated in these sibling pairs. Catalase mRNA expression levels were also related and highly heritable in these sibling pairs. The association between sibship and glutathione peroxidase mRNA expression levels did not reach statistical significance. This study suggests that SF catalase mRNA expression levels, known to be associated with diabetic nephropathy risk, are in part genetically determined.

Effects of duration and age at onset of type 1 diabetes on preclinical manifestations of nephropathy.

Several studies have reported that the effect of type 1 diabetes disease duration on nephropathy may be greater during or after puberty than an equivalent number of years before puberty. The International Diabetic Nephropathy Study examined the effects of disease duration and age at onset on glomerular morphometry obtained from kidney biopsy in 243 young type 1 diabetic subjects with albumin excretion rates <100 micro g/min: 184 with prepubertal onset, 35 with pubertal onset, and 24 with postpuberty onset. Outcomes included the volume fraction of the glomerulus occupied by the mesangium [Vv(Mes/glom)], glomerular basement membrane (GBM) width, and the surface density of peripheral glomerular capillary basement membrane per glomerulus Sv(PGBM/glom). Vv(Mes/glom) progressed slowly in the first 14 or 15 years after disease onset but more rapidly thereafter. GBM width increased, while Sv(PGBM/glom) decreased with increasing disease duration. No statistically significant differences in the effect of duration were observed among the three age-at-onset subgroups, even with multivariate adjustment for sex, center, baseline HbA(1c) concentration, diastolic blood pressure, height, and BMI. Our results may explain the conflicting results of previous studies regarding the effects of pre-versus postpuberty disease duration.

The early natural history of nephropathy in type 1 diabetes: II. Early renal structural changes in type 1 diabetes.

Renal structural abnormalities are known to precede the development of proteinuria, hypertension, and reduced renal function in patients with type 1 diabetes. The determinants of these early structural abnormalities are, however, largely unknown. The International Diabetic Nephropathy Study (IDNS) has recruited 243 children and adults (aged 10-40 years) in Montreal, Minneapolis, and Paris to identify and quantify these determinants. All study subjects were normotensive and had normal-to-high glomerular filtration rates (GFRs) and urinary albumin excretion rates (AERs) <100 microg/min at study entry. Only 8 of 243 had an AER > or =20 microg/min (microalbuminuria). Two renal biopsies are obtained at a 5-year intervals, with baseline and follow-up measures of renal function, blood pressure (BP), HbA(1c), plasma lipids, and AER. Herein, we examine the baseline renal biopsy morphometric findings in these subjects and in 87 kidney donor control subjects and explore the associations between findings and clinical and demographic variables. The principal morphometric abnormalities were increased glomerular basement membrane (GBM) width and fractional volume of mesangium [Vv(Mes/glom)] and mesangial matrix [Vv(MM/glom)]. The frequency of these abnormalities increased with increasing duration of diabetes but was observed as early as 2-8 years after onset. Diastolic BP (DBP), but not HbA(1c), was directly associated with these abnormalities. Elevated GFR was associated with only a small increase in peripheral glomerular capillary basement membrane filtration surface density. Center differences were detected in renal structural, renal functional, and BP parameters, especially between the Paris and North American centers. GBM width, Vv(Mes/glom), and Vv(MM/glom) are significantly increased even within a few years of onset of type 1 diabetes. These changes are detectable in normoalbuminuric patients and are related to duration, BP, and study site. Changes in these and other morphometric measures over 5-year follow-up should help clarify the roles of glycemia and other determinants of the rates of development of diabetic nephropathy lesions, as well as their relationships to early changes in BP, albumin excretion, and renal function.

Cellular basis of diabetic nephropathy: 1. Study design and renal structural-functional relationships in patients with long-standing type 1 diabetes.

This study was designed to elucidate the cellular basis of risk of or protection from nephropathy in patients with type 1 diabetes. Entry criteria included diabetes duration of > or =8 years (mean duration, 22.5 years) and glomerular filtration rate (GFR) >30 ml x min(-1) x 1.73 m(-2). Patients were classified, on the basis of the estimated rate of mesangial expansion, as "fast-track" (upper quintile) or "slow-track" (lower quintile). A total of 88 patients were normoalbuminuric, 17 were microalbuminuric, and 19 were proteinuric. All three groups had increased glomerular basement membrane (GBM) width and mesangial fractional volume [Vv(Mes/glom)], with increasing severity from normoalbuminuria to microalbuminuria to proteinuria but with considerable overlap among groups. Vv(Mes/glom) (r = 0.75, P < 0.001) and GBM width (r = 0.63, P < 0.001) correlated with albumin excretion rate (AER), whereas surface density of peripheral GBM per glomerulus [Sv(PGBM/glom)] (r = 0.50, P < 0.001) and Vv(Mes/glom) (r = -0.48, P < 0.001) correlated with GFR. Vv(Mes/glom) and GBM width together explained 59% of AER variability. GFR was predicted by Sv(PGBM/glom), AER, and sex. Fast-track patients had worse glycemic control, higher AER, lower GFR, more hypertension and retinopathy, and, as expected, worse glomerular lesions than slow-track patients. Thus, there are strong relationships between glomerular structure and renal function across the spectrum of AER, but there is considerable structural overlap among AER categories. Given that normoalbuminuric patients may have advanced glomerulopathy, the selection of slow-track patients based on glomerular structure may better identify protected patients than AER alone.

Glomerular endothelial cells synthesize collagens but little gelatinase A and B.

Mesangial sclerosis is a major feature of progressive renal disease. The mesangium contains mesangial cells and is bounded by the peripheral glomerular basement membrane and endothelial cells. Mesangial cells synthesize and degrade extracellular matrix. Whereas both mesangial and endothelial cells synthesize extracellular matrix components, the degradative pathway, well studied in the former, has not been investigated in endothelial cells. This study examines lines of all three glomerular cell types derived from female B6SJLF1/J mice, as well as mRNA levels for collagens alpha1(I), alpha1(IV), alpha3 (IV), alpha5 (IV), and alpha1 (VI), laminin, tenascin, matrix metalloproteinase-2 (MMP-2), and MMP-9. Type I and IV collagen synthesis was confirmed by enzyme-linked immunosorbent assay. MMP-2 and MMP-9 enzyme activity was measured by zymography. It was found that glomerular endothelial cells are a significant source of collagens, laminin, and tenascin. However, they express only low levels of MMP-2 and no detectable MMP-9. Stimulation with exogenous transforming growth factor-beta1 leads to a significant increase in collagen I, tissue inhibitors of metalloproteinase-1, and MMP-9 in conditioned media. These data suggest that glomerular endothelial cells may play an active role in extracellular matrix remodeling in glomerular disease.

Sequential renal biopsies in insulin-dependent diabetic patients: Structural factors associated with clinical progression

Quantitative structural studies in native kidneys of IDDM patients have almost all been cross sectional, and little is known regarding the dynamics of progression of structural lesions in relation to clinical progression. It has been suggested that interstitial may be more important than glomerular changes in determining functional outcome. This study evaluated renal structure in sequential biopsies from IDDM patients with established renal lesions to determine whether glomerular, arteriolar and interstitial changes progress together and in concordance with measures of renal function. Eleven long-term IDDM patients [age 29 +/- 10 years, duration 17 +/- 7 years (mean +/- SD)] had renal function studies and kidney biopsies performed at two occasions 5.6 +/- 1.6 years apart. HbA1 as well as creatinine clearance (CCr) did not change over this time; albumin excretion rate (AER) increased from 12 (6 to 280) to 19 (5 to 2462) [median (range)] mg/24 hr (P < 0.03). AER increased in the three patients with abnormal albuminuria at first observation, and two normoalbuminuric patients became microalbuminuric. Blood pressure (BP) did not change; however, the number of patients on antihypertensive therapy increased from 1 to 5. All structural parameters were abnormal at first evaluation. Mesangial fractional volume [Vv(mes/glom)] and mean glomerular volume increased and the surface density of the peripheral glomerular basement membrane (GBM) decreased, while GBM width did not change over the five years of the study. Also, arteriolar hyalinosis lesions progressed, while the fractional volume of cortical interstitium [Vv(interstitium/cortex)] and the percent of globally sclerosed glomeruli did not change. The only structural change that correlated with the increasing AER was the change in Vv(mes/glom). Changes in structural parameters, AER or CCr did not significantly correlate with baseline BP or change in BP over the five years. Although based on a small number of patients, this study suggests that at the stage of disease where renal lesions are established and where some IDDM patients are in transition to microalbuminuria or early clinical nephropathy, continuing mesangial expansion is the central variable. Interstitial changes were not occurring over this time. Progressive interstitial expansion at the later stages of diabetic nephropathy may thus be consequent to advanced diabetic glomerular injury.

Structural-functional relationships in Alport syndrome.

Renal morphometric analysis was performed in 15 (13 male) Alport syndrome patients ages 4 to 26 years, along with 10 controls ages 3 to 26 years, to better understand the structural basis of renal dysfunction in Alport syndrome. The glomerular basement membrane (GBM) width class frequencies of controls were normally distributed; those of Alport syndrome patients were slightly skewed, especially toward thicker classes, although there was also an increase in the proportion of thinner classes. Mesangial volume fraction was not different between Alport syndrome patients (0.21 +/- 0.09) and controls (0.19 +/- 0.04). There was an inverse correlation between mesangial volume fraction and creatinine clearance in Alport syndrome patients (r = -0.72, P < 0.01); however, the creatinine clearances in Alport syndrome patients were far less than in insulin-dependent diabetic patients with similar mesangial volume fraction. Similarly, there was no significant difference in the surface density of the peripheral GBM (in square micrometers per cubic micrometer) in Alport syndrome patients (0.12 +/- 0.04) versus controls (0.13 +/- 0.02). The surface density of the peripheral GBM correlated with creatinine clearance in Alport syndrome patients (r = 0.71, P < 0.01). However, there was a greater reduction in creatinine clearance as related to declining the surface density of the peripheral GBM in Alport syndrome than in diabetic patients. The cortical interstitial volume fraction was highly inversely correlated with creatinine clearance in Alport syndrome patients (r = -0.85, P < 0.01). Global glomerular sclerosis was 0% in five and 5 to 61% in nine Alport syndrome patients and correlated inversely with creatinine clearance (r = -0.74, P < 0.01). However, the creatinine clearance was lower in Alport syndrome than in diabetic patients with similar cortical interstitial volume fraction and percent glomerular sclerosis. There was no significant difference in an index of glomerular number between Alport syndrome patients and controls. Thus, changes in mesangial volume fraction, cortical interstitial volume fraction, percent glomerular sclerosis, and surface density of the peripheral GBM in Alport syndrome patients only partially account for the reduction in creatinine clearance. It was speculated that decreased glomerular capillary wall hydraulic conductivity in Alport syndrome could explain many of these observations.

Structural-functional correlations of diabetic nephropathy

Patient 1. A 30-year-old woman had had type-I (insulin-dependent) diabetes mellitus (IDDM) for 27 years. At the time of evaluation, she weighed 50 kg, was 157 cm tall, and had a blood pressure of 114/80 mm Hg. She had background retinopathy and no clinical neuropathy. Her urine was positive for protein by dipstick and the albumin excretion rate was 2500 /tg/min. Her hemoglobin A1 was 8.9%; serum creatinine, 1.2 mg/dl; and creatinine clearance, 79 mI/mm/i .73 m2. As part of the study of renal structure and function in siblings concordant for IDDM, she underwent a percutaneous renal biopsy. Light microscopy revealed that 30% of the glomeruli were globally sclerosed. The others all had moderately severe diffuse mesangial expansion. Many afferent and efferent glomerular arterioles showed varying amounts of hyalinosis, with some vessel walls completely replaced by the waxy, homogeneous, pink hyaline material. Mild diffuse interstitial expansion was evident, and patchy areas of more marked interstitial fibrosis were found, mainly in association with atrophic tubules or sclerotic glomeruli. Electron microscopic morphometric analysis showed the mesangial volume fraction (that is, fraction of glomerular volume occupied by mesangium [VvMesJ) to be 0.40 (upper limit of normal, 0.22). The VvMes cells measured 0.11 and VvMes matrix 0.26; the ratio of mesangial matrix to total mesangium was 0.65 (normal ratio 0.5). The surface density of the peripheral glomerular basement membrane (that is, the filtration surface per unit of tuft area {SvPGBM]) was 0.087 (lower limit of normal, 0.095) and the GBM width was 661 nm (upper limit of normal, 450 nm). Patient 2. The 28-year-old sister of Patient 1 developed IDDM at age 5. She had no retinopathy or neuropathy. She weighed 60 kg, was 164 cm tall, and had a blood pressure of 102/64 mm Hg. The urine was negative for protein by dipstick; urine albumin excretion rate was 75 g/min. Her hemoglobin A1 was 14%; serum creatinine, 0.9 mg/dl; and creatinine clearance, 71 ml/min/l .73 m2. Light microscopic evaluation of her renal biopsy specimen was similar to that of her sister. Morphometric analysis revealed a VvMes of 0.45, a ratio of mesangial matrix to total mesangium of 0.63, an SvPGBM of 0.83, and a GBM width of 712 nm.

Glomerular extracellular matrices in rat diabetic glomerulopathy by scanning electron microscopy.

Characteristic pathological changes in the glomeruli in diabetic nephropathy include expansion of the mesangial matrix and thickening of the glomerular basement membrane (GBM). Using an acellular digestion technique combined with scanning electron microscopy, the three-dimensional ultrastructural changes in glomerular extracellular matrices were studied in rats with diabetic glomerulopathy. Diabetes was induced by the intravenous injection of streptozotocin and morphological analyses were performed 3, 6 and 11 months after the injection. Expansion of mesangial area and GBM thickening became evident with time. After treatment with the series of detergents, all cellular components were completely removed leaving the extracellular matrices intact. In normal controls, the mesangial matrix appeared as fenestrated septa with oval or round stomata between the glomerular capillaries. In diabetic glomerulopathy, expansion of mesangial matrix and narrowing of the mesangial fenestrae were observed. These changes in the mesangial matrices seem to play a vital role in the progression of glomerulosclerosis in rat diabetes. A subendothelial thin layer of the GBM was continuous with the mesangial matrix. One cause of GBM thickening in streptozotocin diabetes may be expansion of the mesangial matrix into the peripheral GBM.

The pathogenesis of Alport syndrome involves type IV collagen molecules containing the α3(IV) chain: Evidence from anti-GBM nephritis after renal transplantation

Mutations in the COL4A5 collagen gene have been implicated as the primary defect in Alport syndrome, a heritable disorder characterized by sensorineural deafness and glomerulonephritis that progresses to end-stage renal failure. In the present study, the molecular nature of the defect in Alport glomerular basement membrane (GBM) was explored using anti-GBM alloantibodies (tissue-bound and circulating) produced in three Alport patients subsequent to renal transplantation. The alloantibodies bound to the alpha 3(IV)NC1 domain of type IV collagen and not to any other basement membrane component. In tissue sections, the alloantibodies bound specifically to peripheral GBM in normal kidney and the affected renal transplant but not to that of Alport kidney. These results establish that: the alpha 3 chain in type IV collagen molecules, the Goodpasture autoantigen, is the target alloantigen in post-transplant anti-GBM nephritis in patients with Alport syndrome, and that a molecular commonality exists in the pathogenesis of anti-GBM nephritis causing loss of renal allografts in patients with Alport syndrome and renal failure in patients with Goodpasture syndrome. These findings implicate: (1) defective assembly of type IV collagen molecules containing the alpha 3(IV) chain in Alport GBM; and (2) the existence of a mechanism linking the assembly of molecules containing the alpha 3(IV) chain with those containing the alpha 5(IV) chain.

Effect of mesangial cell lysis and proliferation on glomerular hemodynamics in the rat

To elucidate an involvement of mesangial cells in the regulation of glomerular hemodynamics, renal micropuncture techniques and glomerular morphometry were employed in Munich-Wistar rats with mesangial cell lytic or proliferative lesions induced by administration of an antibody reactive with Thy-1.1-like antigens on the mesangial cell surface. The antibody-induced mesangial cell lysis at day 1 resulted in a significant decrease in glomerular ultrafiltration coefficient, leading to reduction in single nephron glomerular filtration rate (SNGFR) in spite of a significant increase in both glomerular hydrostatic pressure and single nephron plasma flow (SNPF). During the antibody-induced proliferative lesion at day 6, glomerular ultrafiltration coefficient and SNGFR remained reduced; however, SNPF was now decreased. Morphometric analysis showed the enlargement of capillary luminal volume and the development of new open space in the mesangium accessible for blood flow in the mesangial cell-lytic glomeruli at day 1. An increase in mesangial cell volume was found in the proliferative glomeruli at day 6. The total area of peripheral glomerular basement membrane, presumed as the probable filtration area, was unchanged in these glomeruli. These results indicate that mesangial lesions decrease glomerular ultrafiltration coefficient, and suggest that mesangial cells participate in regulation of glomerular filtration rate.

Three-dimensional architecture of glomerular extracellular matrices in diabetic glomerulosclerosis

The three-dimensional ultrastructural changes of the glomerular extracellular matrices in diabetic glomerulosclerosis were studied in acellular rat and human diabetic glomeruli by scanning electron microscopy. The mesangial matrix appeared as fenestrated septa with oval stomata between the glomerular capillaries in normal control specimens. In diabetic glomerulosclerosis, both in humans and rats, expansion of mesangial matrix and narrowing of the mesangial fenestrae were observed. A thin layer of the mesangial matrix extended into the peripheral glomerular basement membrane (GBM) subendothelially. Thickening of the GBM in diabetic nephropathy might be due to expansion of the mesangial matrix into the peripheral GBM.