Maternal undernutrition upregulates apoptosis in offspring nephrogenesis.

Maternal undernutrition (MUN) results in growth-restricted newborns with reduced nephron numbers that is associated with increased risk of hypertension and renal disease. The total adult complement of nephrons is set during nephrogenesis suggesting that MUN affects the staged development of nephrons in as yet unknown manner. A possible cause may be the increased renal apoptosis; therefore, we investigated whether apoptotic signaling and cell death were increased in MUN rat kidneys. Pregnant rat dams were fed an ad libitum diet [control] or were 50% food restricted (MUN) starting at embryonic day (E) 10. Male offspring kidneys (n = 5 each, MUN and control) were analyzed for mRNA using quantitative PCR (E20) and for protein expression using Western blotting and immunohistochemistry (E20 and postnatal day 1, P1). Apoptosis was measured by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Upregulation of pro-apoptotic protein expression was detected at E20 (Fas receptor, caspase 9) and at P1 (caspase 3, Bax). The anti-apoptotic factor Bcl2 was significantly decreased in P1 kidneys. Kidney TUNEL showed apoptotic nuclei significantly increased in the P1 nephrogenic zone (MUN 3.3 + 0.3 v. C 1.6 + 0.5, P = 0.002). The majority of apoptotic nuclei co-localized to mesenchyme and pretubular aggregates in the nephrogenic zone. Differential regulation of apoptosis in mesenchyme and pretubular aggregates following parturition suggests a mechanism for nephropenia in gestational programming of the kidney.

GDNF and MAPK-ERK pathway signaling is reduced during nephrogenesis following maternal under-nutrition.

Maternal under-nutrition (MUN) during gestation results in growth-restricted newborns with reduced glomerular number and subsequent hypertension. We investigated dysregulation of glial derived neurotrophic factor (GDNF) and MAPK-ERK (mitogen-activated protein kinase-extracellular signal-regulated protein kinase) signal pathway gene expression following MUN. MUN rats were 50% food restricted from embryonic day 10 till postnatal day 1. Kidneys were harvested at embryonic day (E)20, and postnatal days (P)1 and 21. Kidney protein expression was determined by Western blot. At E20, protein expression of growth factor receptor alpha 1 (GFRα1) and phosphorylated ERK1/2 and mitogen-activated protein kinase kinase (MEK)1/2 were reduced significantly, and immunohistochemistry confirmed reduction of phosphorylated ERK (pERK) with maintenance of pERK localization. Total MEK and ERK were unchanged. At P1, only GFRα1 and pERK1/2 were reduced significantly while at P21, expression of all growth factors except total MEK was unchanged. Total MEK was increased. Glomerular number was decreased by 19% in P21 kidneys and blood pressure was increased in 12-week-old rats. In conclusion, GDNF and MAPK-ERK signaling are dysregulated during active nephrogenesis in fetal and early newborn offspring kidneys in the MUN model. This may be a key mechanism in reduced offspring nephrogenesis and programmed hypertension.

Glucose and diabetes: effects on podocyte and glomerular p38MAPK, heat shock protein 25, and actin cytoskeleton.

Phosphorylated p38 (pp38) mitogen-activated protein kinase (MAPK) regulates heat shock protein 25 (HSP25), stabilizing fibrillar actin (FA) and preventing cleavage to G-actin (GA). Cultured podocytes (Pods) were exposed to glucose (5.5-50 mM)+/-p38MAPK inhibitor SB202190 (SB) or control SB202474 to assess the effects on FA/GA and Pod structure. The relationship of p38MAPK with in vivo Pod structure and albuminuria (Ualb) was assessed in rats with streptozotocin (SZ)-induced diabetes (DM) for 1 week, 1 month, and 4 months. High glucose induced concentration-dependent increases in pp38MAPK and phosphorylated HSP25 (pHSP25) maintained actin cytoskeleton. Inhibition by SB diminished pp38MAPK and pHSP25, decreased FA/GA, and altered FA and GA immunohistochemical appearance. In SZ-DM, glomerular pp38MAPK and biphosphorylated HSP25 were increased after 1 week, declining at 1 month, and at or below C values at 4 months. Glomerular FA/GA in DM was normal at 1 week, declining at 1 month, and low at 4 months. Ualb/creatinine was similar in DM vs C at 1 week, and increased at 1 and 4 months. Morphometry demonstrated progressively diminishing slit pore density in DM over time, denoting evolving effacement. There were strong correlations between slit membrane density and both glomerular biphosphorylated HSP25 and ln Ualb/cr ratio. The data suggest that increased pp38MAPK and pHSP25 comprise an acute adaptation to glycemic stress. Later depletion of DM may contribute to Pod structural alterations and Ualb.

Recurrent and de novo diabetic nephropathy in renal allografts.

BACKGROUND: Histologic findings of diabetic nephropathy (DN) are observed in allografts of patients with pretransplant (PreTx) diabetes mellitus (DM) and in patients who develop DM posttransplant (PostTx). Patients with allograft biopsies (Bx) were retrospectively studied to determine the incidence of recurrent and de novo DN and to ascertain what, if any, risk factors predispose to histologic DN in either patient population. METHODS: From the renal transplant services at four hospitals from 1992 to 2000, the authors identified all patients with PreTxDM and PostTxDM (n=81). Those with renal biopsies performed >/=18 months PostTx were classified according to the presence or absence of histologic DN (Bx-positive, n=23; Bx-negative, n=35). Patients were then subdivided into four categories-recurrent DN (n=16), de novo DN (n=7), no recurrent DN (n=27), and no de novo DN (n=8)-for analyses. RESULTS: Among these 58 patients, 74.1% had PreTx and 25.9% had PostTx diabetes. Of those with histologic DN, 69.6% were recurrent DN and 30.4% were de novo DN, making de novo DN at least as likely to develop as recurrent DN. After the onset of diabetes in the de novo population, the time to development of histologic DN was similar in the recurrent and the de novo patients (6.68+/-3.86 years vs. 5.90+/-3.13 years, P=0.66) and more rapid than previously reported. Apart from a more frequent family history of hypertension in patients with allograft DN compared with those without allograft DN, known risk factors for the development of native DN did not significantly differ among patients in the four cohorts. Proposed risk factors related to transplantation did not correlate with the development of recurrent or de novo DN. CONCLUSION: Among patients with histologic DN, de novo DN occurred at least as frequently as recurrent DN, and the time to onset of histologically apparent DN was more rapid than previously reported. Neither the usual clinical predictors of DN nor clinical variables related to transplantation clearly distinguished the group with DN from the group without it, potentially implicating novel mechanisms in its pathogenesis.

Proteinuria in patients with arterial/arteriolar nephrosclerosis.

UNLABELLED: Controversy exists regarding the level of proteinuria in patients with nephrosclerosis. MATERIAL AND METHODS: We retrospectively examined the clinical parameters of 67 patients with the histologic diagnosis of nephrosclerosis defined as arteriolar hyalinization and/or arterial intimal fibrosis in the absence of other findings in an adequate renal biopsy. Biopsies were performed for clinical indications and were submitted to Cedars-Sinai Pathology Department from January 1994 to March 1999. RESULTS: The mean age and blood pressure (+/- SD) was 60 +/- 17 years, and 139 +/- 19/80 +/- 12 mmHg. Mean serum creatinine was 2.3 +/- 1.3 mg/dl and 24-hour urinary protein excretion was 0.94 +/- 0.73 g, 66% had < or = 1 g/day, 25% had > 1 but < or = 2 g/day, 6% had > 2 g but < 3 g/day and 1 patient had nephrotic range proteinuria. Twelve patients had no history of hypertension. They had a mean blood pressure of 121 +/- 8/76 +/- 8. Their mean serum creatinine was 1.5 +/- 0.6 mg/dl and their mean 24-hour urinary protein excretion was 0.78 +/- 0.43 g. CONCLUSIONS: Our data do not confirm that of Innes et al. [1993] who reported proteinuria > 1.5 g/day in 40% and nephrotic syndrome in 22% of patients with nephrosclerosis; systemic hypertension may not be a prerequisite for the development of nephrosclerosis.

Glomerular mRNAs in human type 1 diabetes: biochemical evidence for microalbuminuria as a manifestation of diabetic nephropathy.

BACKGROUND: In patients with type 1 diabetes, some consider microalbuminuria to be a predictor of diabetic nephropathy while others believe it is an early feature of diabetic nephropathy. METHODS: Levels of mRNAs that are of pathogenetic relevance in diabetic nephropathy were compared in glomeruli isolated from microalbuminuric and overtly proteinuric subjects and in control normoalbuminuric diabetic subjects and living renal transplant donors. RESULTS: In subjects with microalbuminuria and overt proteinuria, glomerular mRNAs were virtually identical and approximately twofold higher for connective tissue growth factor (CTGF; P < 0.01) and collagen alpha2(IV) (P < 0.03) compared to living renal donors and normoalbuminuric patients. Glomerular glyceraldehyde-3-phosphate dehydrogenase (GAPDH) levels were not significantly different among the groups (P = 0.4). Weak but statistically significant correlations were noted between CTGF mRNA and albuminuria (assessed by rank), fractional mesangial surface area, and a composite renal biopsy index. Glomerular CTGF mRNA correlated inversely with creatinine clearance. Glomerular collagen alpha2(IV) mRNA levels correlated with albuminuria (by rank) and less strongly with fractional mesangial area. CONCLUSION: To our knowledge, these data provide the first biochemical evidence demonstrating that the glomeruli of microalbuminuric patients and those with overt proteinuria do not differ significantly. The data support the concept that microalbuminuria is not "predictive" of diabetic nephropathy, but rather is an earlier point in the spectrum of diabetic nephropathy.

Cyclosporine microemulsion- and mycophenolate mofetil-related lymphoid aggregates are not associated with acute rejection.

BACKGROUND: Microemulsion cyclosporine, mycophenolate mofetil, and prednisone have become a common immunosuppressive protocol in renal transplantation. We identified lymphocytic infiltrates in transplant fine-needle aspirates and core biopsies from patients on this regimen without acute rejection clinically or by standardized morphological criteria and investigated this inflammatory response. METHODS: Twenty-eight aspirates from 21 patients were included and assessed in the standard fashion. Nine core biopsies showing interstitial lymphocytic infiltration were evaluated with antibodies against CD3, CD4, CD8, CD20, CD30, CD56, KP1, and epithelial membrane antigen (EMA). Aspirates and biopsies were assessed for tubular cell major histocompatibility complex (MHC) class II antigen and for gamma-interferon (gamma-IFN), interleukin-4 (IL-4), and IL-10 mRNAs by reverse transcription-polymerase chain reaction. RESULTS: Fifteen aspirates showed immune activation solely due to mature lymphocytes and monocytes; 13 had no immune activation. All aspirates were negative for MHC class II antigens. Of 6 activated aspirates assessed for gamma-IFN mRNA, 5 were negative. All 21 patients had similar clinical characteristics and recovered renal function without rejection treatment. The core biopsies had lymphocytes in 5-30% of the interstitium. The cells were 70-85% CD3+, with 50-85% CD4+, 3-10% KP1+, and rare cells CD56+. No T-cell activation was present (EMA- and CD30-). Seven biopsies were assessed and were negative for gamma-IFN mRNA; only one biopsy had weakly positive MHC class II staining. Two activated aspirates were negative for IL-4 and IL-10 mRNA, while three biopsies each contained IL-4 and IL-10 mRNAs. CONCLUSIONS: Inactive interstitial lymphoid infiltrates are frequent in patients on this drug regimen and should not be interpreted as acute rejection, particularly in aspirate samples. These lymphocytes may play a role in long-term allograft acceptance.

Association of parvovirus B19 infection with idiopathic collapsing glomerulopathy.

BACKGROUND: Collapsing glomerulopathy (CG), a disorder with severe glomerular and tubular involvement, occurs either as an idiopathic lesion or in some patients with human immunodeficiency virus (HIV) infection known as HIV-associated nephropathy (HIVAN). We previously reported a renal transplant recipient with de novo CG and red cell aplasia in association with persistent parvovirus B19 (PVB19) infection. This prompted us to look for an association between PVB19 infection and CG. METHODS: DNA from archived biopsies of patients with CG was analyzed for PVB19 by polymerase chain reaction (PCR). Results were compared with HIVAN, idiopathic focal segmental glomerulosclerosis (FSGS), and controls. In situ hybridization (ISH) was done to localize PVB19 in renal biopsies. Peripheral blood specimens of patients with CG, HIV infection, healthy controls, and randomly selected hospitalized patients (sick controls) were also analyzed for PVB19. RESULTS: PVB19 DNA was detected in renal biopsies of 18 out of 23 (78.3%) patients with CG, 3 out of 19 (15.8%) with HIVAN, 6 out of 27 (22.2%) with FSGS, and 7 out of 27 (25.9%) controls (P < 0.01, CG vs. HIVAN, FSGS, and controls). PVB19 was detected in peripheral blood of 7 out of 8 (87.5%) CG patients, 3 out of 22 (13.6%) with HIV infection, 4 out of 133 (3%) healthy controls, and 2 out of 50 (4%) sick controls (P < 0.001, CG vs. HIV infected, healthy, and sick controls). PVB19 was identified in glomerular parietal and visceral epithelial and tubular cells by ISH. CONCLUSIONS: The significantly higher prevalence of PVB19 DNA in renal biopsies and peripheral blood of CG patients suggests a specific association between PVB19 infection and CG. In susceptible individuals, renal epithelial cell infection with PVB19 may induce CG.

Th1-type responses mediate spontaneous ileitis in a novel murine model of Crohn's disease.

We describe here the immunologic characterization of a new mouse strain, SAMP1/Yit, which spontaneously develops a chronic intestinal inflammation localized to the terminal ileum. The resulting ileitis bears a remarkable resemblance to human Crohn's disease. This strain of mice develops discontinuous, transmural inflammatory lesions in the terminal ileum with 100% penetrance by 30 weeks of age. The intestinal inflammation is characterized by massive infiltration of activated CD4+ and CD8alpha(+)TCRalphabeta(+) T cells into the lamina propria and is accompanied by a dramatic decrease in the intraepithelial lymphocyte CD8alpha(+)TCRgammadelta(+)/CD8alpha(+)TCRalphabeta(+) ratio. The results of adoptive transfer experiments strongly suggest that CD4+ T cells that produce a Th1-like profile of cytokines, e.g., IFN-gamma and TNF, mediate the intestinal inflammation found in SAMP1/Yit mice. In addition, pretreatment of adoptive transfer recipients with a neutralizing anti-TNF antibody prevents the development of intestinal inflammation, suggesting that TNF plays an important role in the pathogenesis of intestinal inflammation in this model. To our knowledge, these data provide the first direct evidence that Th1-producing T cells mediate intestinal inflammation in a spontaneous animal model of human Crohn's disease.

12-lipoxygenase is increased in glucose-stimulated mesangial cells and in experimental diabetic nephropathy.

BACKGROUND: Arachidonic acid-derived 12-lipoxygenase (12-LO) products have potent growth and chemotactic properties. The present studies examined whether 12-LO and fibronectin are induced in cultured rat mesangial cells (MCs) exposed to high glucose and whether they are expressed in experimental diabetic nephropathy. METHODS: To determine the effect of high glucose on MC 12-LO mRNA and protein expression, rat MCs were incubated with RPMI medium containing 100 (NG) or 450 mg/dL glucose (HG). For animal studies, rats were injected with diluent (control) or streptozotocin. The latter were left untreated (DM) or treated with insulin (DM + I). At sacrifice after four months, GAPDH, 12-LO, and fibronectin mRNA were measured by competitive reverse transcription-polymerase chain reaction (RT-PCR) in microdissected glomeruli (G). Renal sections were semiquantitatively scored (0 to 4+) for diabetic changes and for 12-LO and fibronectin by immunohistochemistry. RESULTS: 12-LO mRNA expression in MC exposed to HG (12.71 +/- 1.17 attm/microL) and DM G (1.78 +/- 0.65 x 10-3 attm/glomerulus) was significantly higher than those of MCs in NG media (6.71 +/- 0.78 attm/microL) and control G (0.34 +/- 0.12 x 10-3 attm/glomerulus, P < 0.005), respectively. Western blot revealed a 1.7- and a 2.8-fold increase in MC and G 12-LO protein expression, respectively (P < 0.05). The immunohistochemistry score for G 12-LO and diabetic nephropathy score was significantly greater in DM and DM + I than controls. MC and G GAPDH mRNA remained unchanged. CONCLUSIONS: In MCs exposed to HG and in diabetic rat glomeruli, increments in 12-LO mRNA and protein are associated with changes modeling diabetic nephropathy. These findings suggest a role for the 12-LO pathway in the pathogenesis of diabetic nephropathy.

Chlorambucil-induced acute hepatic failure in a patient with membranous nephropathy.

We present the case of a 23-year-old woman with membranous nephropathy who developed acute hepatic failure after being administered chlorambucil for a month.

Glomerular type IV collagen in patients with diabetic nephropathy with and without additional glomerular disease.

BACKGROUND: Type IV collagen is a constituent of mesangial matrix and is increased in amount in many forms of glomerular injury. METHODS: We performed renal biopsies in patients who (1) were donating a kidney to a relative (LRD, N = 6), (2) had diabetic glomerulopathy with or without nephrosclerosis (DM, N = 6), or (3) had diabetic glomerulopathy with a superimposed glomerular lesion (DM+, N = 5). Glomerular collagen alpha2(IV) and control glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNAs were measured, and the former correlated with clinical and morphological data to assess its usefulness in reflecting glomerular injury. RESULTS: Collagen alpha2(IV) mRNA levels were lowest in LRD (2.9 +/- 0.6 attomol/glomerulus), higher in DM (5.9 +/- 1.6, P = 0.05), and highest in DM+ (12.7 +/- 2.8 attm/glomerulus, P < 0.05 vs. LRD and vs. DM). Control GAPDH mRNA levels were not significantly different (P > 0.05). Levels of proteinuria, serum creatinine, and glomerular size did not correlate with collagen alpha2(IV) mRNA levels. The fractional mesangial area and the fractional mesangial area occupied by type IV collagen were higher in both diabetic groups than in LRD (P < 10-6), but the intensity of type IV collagen staining in the diabetic patients was significantly less than that seen in the LRD (P < 0.01). In DM+ patients, extramesangial type IV collagen was present. Fractional mesangial area and glomerular collagen alpha2(IV) mRNA levels correlated (r = 0.45, P < 0.05). CONCLUSION: These data are consistent with a view of diabetic nephropathy as a lesion of increased alpha2 type IV collagen transcription, increased total amount of collagen present, but decreased mesangial density relative to other matrix molecules. These data further demonstrate that glomerular injury superimposed on diabetic nephropathy contributes to additional structural damage by inducing increased synthesis of type IV collagen at extramesangial sites.

Acetylsalicylic acid reduces vegetation bacterial density, hematogenous bacterial dissemination, and frequency of embolic events in experimental Staphylococcus aureus endocarditis through antiplatelet and antibacterial effects.

BACKGROUND: Platelets are integral to cardiac vegetations that evolve in infectious endocarditis. It has been postulated that the antiplatelet aggregation effect of aspirin (ASA) might diminish vegetation evolution and embolic rates. METHODS AND RESULTS: Rabbits with Staphylococcus aureus endocarditis were given either no ASA (controls) or ASA at 4, 8, or 12 mg. kg-1. d-1 IV for 3 days beginning 1 day after infection. Vegetation weights and serial echocardiographic vegetation size, vegetation and kidney bacterial densities, and extent of renal embolization were evaluated. In addition, the effect of ASA on early S aureus adherence to sterile vegetations was assessed. In vitro, bacterial adherence to platelets, fibrin matrices, or fibrin-platelet matrices was quantified with either platelets exposed to ASA or S aureus preexposed to salicylic acid (SAL). ASA at 8 mg. kg-1. d-1 (but not at 4 or 12 mg. kg-1. d-1) was associated with substantial decreases in vegetation weight (P<0.05), echocardiographic vegetation growth (P<0.001), vegetation (P<0.05) and renal bacterial densities and renal embolic lesions (P<0.05) versus controls. Diminished aggregation resulted when platelets were preexposed to ASA or when S aureus was preexposed to SAL (P<0.05). S aureus adherence to sterile vegetations (P<0.05) or to platelets in suspension (P<0.05), fibrin matrices (P<0.05), or fibrin-platelet matrices (P<0.05) was significantly reduced when bacteria were preexposed to SAL. CONCLUSIONS: ASA reduces several principal indicators of severity and metastatic events in experimental S aureus endocarditis. These benefits involve ASA effects on both the platelet and the microbe.

Selective activation of sar promoters with the use of green fluorescent protein transcriptional fusions as the detection system in the rabbit endocarditis model.

The global regulatory locus sar is composed of three overlapping transcripts initiated from a triple-promoter system (designated P1, P3, and P2). To explore if the individual sar promoters are differentially expressed in vitro and in vivo, we constructed a shuttle plasmid (pALC1434) containing a promoterless gfpUV gene (a gfp derivative [Clontech]) preceded by a polylinker region. Recombinant shuttle vectors containing individual sar promoters upstream of the gfpUV reporter gene were then introduced into Staphylococcus aureus RN6390. Northern and immunoblot analysis revealed that P1 is stronger than the P2 and P3 promoters in vitro. Additionally, the levels of the gfpUV transcript driven by individual sar promoters also correlated with the growth cycle dependency of these promoters in liquid cultures, thus suggesting the utility of pALC1434 as a vehicle for reporter fusion. Using the rabbit endocarditis model, we examined the expression of these three GFPUV fusions in vivo by fluorescence microscopy of infected cardiac vegetations 24 h after initial intravenous challenge. Similar to the in vitro findings, P1 was activated both in the center and on the surface of the vegetations. In contrast, the P3 promoter was silent both in vivo and in vitro as determined by fluorescence microscopy. Remarkably, P2 was silent in vitro but became highly activated in vivo. In particular, the sar P2 promoter was activated on the surface of the vegetation but not in the center of the lesion. These data imply that in vivo promoter activation of sar differed from that observed in vitro. Moreover, the individual sar promoters may be differentially expressed in different areas within the same anatomic niche, presumably reflecting the microbial physiological response to distinct host microenvironments. As the sar locus controls the synthesis of both extracellular and cell wall virulence determinants, these promoter-gfpUV constructs should be useful to characterize many aspects of S. aureus gene regulation in vivo.

Plasmapheresis in the treatment of steroid-resistant focal segmental glomerulosclerosis in native kidneys.

A circulating glomerular capillary albumin permeability factor (P(alb)) has been implicated in the pathogenesis of focal segmental glomerulosclerosis (FSGS), which recurs in transplanted kidneys. Plasmapheresis for recurrent FSGS may reduce proteinuria and stabilize renal function if instituted early. We performed six plasmapheresis treatments over 2 weeks in eight patients with a history of steroid-resistant idiopathic FSGS in native kidneys for an average of 12 +/- 2.3 months to determine whether treatment would decrease proteinuria or stabilize renal function. P(alb) was measured before and after plasmapheresis, and patients were followed-up for a mean of 29 +/- 4 months after the development of clinical symptoms. Proteinuria decreased in two of eight treated patients, although only transiently in one of the two. P(alb) improved in one of the two responding patients. Both patients with an improvement in proteinuria had stable renal function at last follow-up. In six of eight patients, there was no improvement in proteinuria despite an improvement in P(alb) (P < 0.0001) after plasmapheresis. At last follow-up, renal function was stable in two of the six nonresponding patients, and four of the six had significant progression of renal disease or were receiving dialysis treatments. These studies suggest that plasmapheresis may diminish proteinuria and stabilize renal function in a small minority of patients with steroid-resistant idiopathic FSGS. However, the lack of a relationship between the removal of the circulating permeability factor and the development of remission in these patients suggests that local factors associated with advanced renal injury or systemic factors unrelated to glomerular permeability play a significant role in determining proteinuria at this late stage of the disease.

Differential response of glomerular epithelial and mesangial cells after subtotal nephrectomy.

Recent studies in both human and experimental chronic renal disease suggest that there is a linkage between glomerular hypertrophy and glomerulosclerosis. To further define these relationships, we studied the changes in glomerular hypertrophy, procollagen alpha 1(IV) mRNA levels and glomerulosclerosis in rats undergoing 1 2/3 nephrectomy (Nx) or sham nephrectomy (SNx). Glomerular hypertrophy, measured biochemically by RNA/DNA and protein/DNA ratios, was significantly increased in Nx compared to SNx two days after subtotal renal ablation (RNA/DNA: Nx = 133 +/- 8%, SNx = 100 +/- 3% of the mean control value, P < 0.01; protein/DNA: Nx = 164 +/- 22%, SNx = 100 +/- 10%, P < 0.05) and remained elevated after 7 and 15 days (RNA/DNA: seven days Nx = 155 +/- 3%, SNx = 100 +/- 13%, P < 0.01; 15 days Nx = 303 +/- 21%, SNx = 100 +/- 24%, P < 0.001; protein/DNA: seven days Nx = 228 +/- 57%, SNx = 100 +/- 18%, P < 0.05; 15 days Nx = 341 +/- 23%, SNx = 100 +/- 18%, P < 0.01). Light microscopic measures of glomerular tuft volume (GTV) were too insensitive to detect glomerular enlargement until 15 days postoperatively, but GTV measured ultrastructurally demonstrated a 20% increment in Nx compared to SNx as early as two days postoperatively (P < 0.01). The latter increment in GTV was due exclusively to glomerular visceral epithelial cell (GVEC) expansion. Glomerular procollagen alpha 1(IV) mRNA levels were significantly elevated only 15 days after nephrectomy (Nx = 265 +/- 58% of the mean control value, SNx = 100 +/- 12%, P < 0.05; corrected for beta-actin mRNA levels). As this time, exuberant mesangial expansion measured ultrastructurally contributed to a 1.6 +/- 0.1-fold increase in GTV (P < 10(-5)), and to a relative decrement in the GVEC contribution to glomerular cells plus matrix (P < 0.01). Segmental sclerosis was observed only 15 days postoperatively in Nx (Nx = 1.3 +/- 0.4% of glomeruli evaluated, SNx = 0.0%, P < 0.05), and there was a strong correlation between the prevalence of segmental sclerosis and the procollagen alpha 1(IV) mRNA levels in Nx at 15 days (r = 0.93, P < 0.01). There was no significant correlation between the RNA/DNA and protein/DNA ratios and procollagen alpha 1(IV) mRNA levels. Thus, glomerular regions responded differentially to subtotal nephrectomy. Early epithelial cell expansion was followed by later mesangial expansion. Glomerular procollagen alpha 1(IV) mRNA levels were elevated only during the second (mesangial) phase of glomerular hypertrophy, when it correlated with glomerulosclerosis, but not during the initial (epithelial) phase, a pattern consistent with a mesangial origin of the procollagen alpha 1(IV) mRNA.

Corticomedullary procollagen alpha1(IV) mRNA levels and localization after subtotal nephrectomy.

The relationships between tubular hypertrophy/hyperplasia, procollagen alpha1(IV) mRNA levels, and the development of tubular basement membrane thickening were studied in male Sprague-Dawley rats subjected to subtotal renal ablation and sacrificed 2 or 15 days later. Tubular hypertrophy and hyperplasia were demonstrable at 2 days, however no increment in procollagen alpha1 (IV) mRNA levels was discerned at that time, demonstrating a dissociation between mRNA levels for classical type IV collagen and tubular enlargement. At 15 days, tubular procollagen alpha1(IV) mRNA levels did increase approximately 2-fold (p < 0.002), localizing predominantly in proximal tubules in the deep cortex and outer medullary stripe. At this time point, there was still no significant correlation to tubular enlargement, but there was a significant correlation to tubular basement membrane thickening (r = 0.89, p < 0.01). These studies demonstrated that an increase in mRNA for classical type IV collagen is not required for the development of hypertrophy, and that the increment is a better marker for matrix expansion than it is for hypertrophy.

TGF-beta in renal allograft rejection.

The role of TGF-beta in pathological processes in the transplanted kidney is beginning to be investigated both in animal models and in humans. In both settings in acute cell-mediated rejection, TGF-beta, receptor, and message have all been documented to be elevated in the tubulointerstitium, likely a reflection of TGF-beta's role in recruiting leukocytes to areas of injury and downregulation of the inflammatory response. In chronic rejection, expression of TGF-beta, message, and induced proteins is elevated, especially in cortex. TGF-beta mRNA, unlike other inflammatory cytokine mRNAs, correlated very well with interstitial fibrosis, a hallmark of chronic rejection. Thus, a relationship between renal scarring and TGF-beta has been documented by most studies of transplant kidneys. Additionally, this growth factor also appears to have a role in the renal fibrosis associated with cyclosporine administration and perhaps in augmenting this drug's immunosuppressive effects.