Cardiovascular and renal outcomes following percutaneous coronary intervention in a population with renal disease: a case-control study.

BACKGROUND: Patients with renal disease are less likely to undergo percutaneous coronary intervention (PCI) due to concerns about poor outcomes. AIM: We describe outcomes following PCI in individuals with chronic kidney disease (CKD), as compared with matched controls with comparable CKD who did not undergo PCI. We also identified factors predictive of poor outcomes following PCI amongst patients with CKD. DESIGN: Retrospective observational case-control study. METHODS: Cases were individuals with CKD (stages 1-5) undergoing PCI between 2008 and 2014. Controls were age, gender and creatinine-matched individuals not requiring PCI. We compared mortality between groups using Kaplan-Meier curves and Cox regression modelling. We assessed changes in serum creatinine using Wilcoxon Rank testing. We explored the relationship between biochemical and haematological measures (baseline creatinine, calcium, phosphate, calcium-phosphate product, parathyroid hormone, white cell count, haemoglobin, platelet count, c-reactive protein and total cholesterol) and post-PCI mortality, using logistic regression. RESULTS: We identified 144 cases and 144 controls. Mortality was significantly lower amongst cases compared with controls [hazard ratio 0.46 (95% confidence intervals 0.31, 0.69)]. PCI did not result in a significant change in renal function (P=0.52). Amongst cases, serum creatinine and calcium-phosphate product were predictors of mortality following PCI. CONCLUSION: Cases undergoing PCI had lower mortality, and PCI was not associated with accelerated CKD progression. On this data, PCI should not be deferred as a treatment option in patients with CKD. Serum creatinine and calcium-phosphate product predict mortality following PCI in this cohort, and may be useful in risk-stratifying patients with CKD being considered for PCI.

The influence of socioeconomic status on presentation and outcome of acute kidney injury.

AIM: Although socioeconomic background is known to impact on the incidence and progression of chronic kidney disease, its influence of on the presentation and outcome for acute kidney injury is not known and is the subject of this study. DESIGN: The Welsh National electronic AKI reporting system was used to identify all cases of AKI in patients >18 years of age between March 2015 and November 2017. METHODS: Socioeconomic classification of patients was derived from the Welsh Index Multiple Deprivation score (WIMD). Patients were grouped according to the WIMD score by their postcode, and the ranked data were categorized into percentiles and correlated with incidence and measures of AKI severity and outcome. RESULTS: Date was collected on a total of 57 654 patients. Increased deprivation was associated with higher AKI incidence rates, more episodes of AKI per patient and more severe AKI at presentation. In contrast 90-day mortality was highest in the most affluent areas. Mortality in affluent areas was driven by increased patient age. Corrected for age 90-day mortality was higher in areas of increased deprivation. CONCLUSION: This study highlights that AKI incidence presentation and outcomes are adversely affected by social deprivation. Further studies are required to understand the extent to which these differences reflect patient related factors or regional differences in provision and access to care.

Adding a new dimension to the weekend effect: an analysis of a national data set of electronic AKI alerts.

BACKGROUND: Increased mortality related to differences in delivery of weekend clinical care is the subject of much debate. AIM: We compared mortality following detection of acute kidney injury (AKI) on week and weekend days across community and hospital settings. DESIGN: A prospective national cohort study, with AKI identified using the Welsh National electronic AKI reporting system. METHODS: Data were collected on outcome for all cases of adult AKI in Wales between 1 November 2013 and 31 January 2017. RESULTS: There were a total of 107 298 episodes. Weekday detection of AKI was associated with 28.8% (26 439); 90-day mortality compared to 90-day mortality of 31.9% (4551) for AKI detected on weekdays (RR: 1.11, 95% CI: 1.08-1.14, P < 0.001, HR: 1.16 95% CI: 1.12-1.20, P < 0.001). There was no 'weekend effect' for mortality associated with hospital-acquired AKI. Weekday detection of community-acquired AKI (CA-AKI) was associated with a 22.6% (10 356) mortality compared with weekend detection of CA-AKI, which was associated with a 28.6% (1619) mortality (RR: 1.26, 95% CI: 1.21-1.32, P < 0.001, HR: 1.34, 95%CI: 1.28-1.42, P < 0.001). The excess mortality in weekend CA-AKI was driven by CA-AKI detected at the weekend that was not admitted to hospital compared with CA-AKI detected on weekdays which was admitted to hospital (34.5% vs. 19.1%, RR: 1.8, 95% CI: 1.69-1.91, P < 0.001, HR: 2.03, 95% CI: 1.88-2.19, P < 0.001). CONCLUSION: 'Weekend effect' in AKI relates to access to in-patient care for patients presenting predominantly to hospital emergency departments with AKI at the weekend.

Community acquired acute kidney injury: findings from a large population cohort.

BACKGROUND: The extent of patient contact with medical services prior to development of community acquired-acute kidney injury (CA-AKI)is unknown. AIM: We examined the relationship between incident CA-AKI alerts, previous contact with hospital or primary care and clinical outcomes. DESIGN: A prospective national cohort study of all electronic AKIalerts representing adult CA-AKI. METHODS: Data were collected for all cases of adult (≥18 years of age) CA-AKI in Wales between 1 November 2013 and 31 January 2017. RESULTS: There were a total of 50 560 incident CA-AKI alerts. In 46.8% there was a measurement of renal function in the 30 days prior to the AKI alert. In this group, in 63.8% this was in a hospital setting, of which 37.6% were as an inpatient and 37.5% in Accident and Emergency. Progression of AKI to a higher AKI stage (13.1 vs. 9.8%, P < 0.001) (or for AKI 3 an increase of > 50% from the creatinine value generating the alert), the proportion of patients admitted to Intensive Care (5.5 vs. 4.9%, P = 0.001) and 90-day mortality (27.2 vs. 18.5%, P < 0.001) was significantly higher for patients with a recent test. 90-day mortality was highest for patients with a recent test taken in an inpatient setting prior to CA-AKI (30.9%). CONCLUSION: Almost half of all patients presenting with CA-AKI are already known to medical services, the majority of which have had recent measurement of renal function in a hospital setting, suggesting that AKI for at least some of these may potentially be predictable and/or avoidable.

Acute kidney injury electronic alerts in primary care - findings from a large population cohort.

BACKGROUND: Electronic reporting of AKI has been used to aid early AKI recognition although its relevance to CA-AKI and primary care has not been described. AIMS: We described the characteristics and clinical outcomes of patients with CA-AKI, and AKI identified in primary care (PC-AKI) through AKI e-Alerts. DESIGN: A prospective national cohort study was undertaken to collect data on all e-alerts representing adult CA-AKI. METHOD: The study utilized the biochemistry based AKI electronic (e)-alert system that is established across the Welsh National Health Service. RESULTS: 28.8% of the 22 723 CA-AKI e-alerts were classified as PC-AKI. Ninety-day mortality was 24.0% and lower for PC-AKI vs. non-primary care (non-PC) CA-AKI. Hospitalization was 22.3% for PC-AKI and associated with greater disease severity, higher mortality, but better renal outcomes (non-recovery: 18.1% vs. 21.6%; progression of pre-existing CKD: 40.5% vs. 58.3%). 49.1% of PC-AKI had a repeat test within 7 days, 42.5% between 7 and 90 days, and 8.4% was not repeated within 90 days. There was significantly more non-recovery (24.0% vs. 17.9%) and progression of pre-existing CKD (63.3% vs. 47.0%) in patients with late repeated measurement of renal function compared to those with early repeated measurement of renal function. CONCLUSION: The data demonstrate the clinical utility of AKI e-alerts in primary care. We recommend that a clinical review, or referral together with a repeat measurement of renal function within 7 days should be considered an appropriate response to AKI e-alerts in primary care.

The long-term impact of eGFR reporting on referral patterns.

BACKGROUND: The prognostic significance of CKD has driven the widespread introduction of automated estimated glomerular filtration rate (eGFR) reporting, and the incorporation of CKD in the revised Quality Outcomes Framework (QOF) of the General Medical Services (GMS) contract in the U.K. AIMS: To assess the long-term impact of the introduction of these two initiatives, on patient referral numbers to a nephrology service. METHODS: Data was collected on the numbers and basic characteristics of all new patients referred from April 2005 to March 2011, to one NHS Health Board. RESULTS: Introduction of eGFR reporting and CKD QOF domains was associated with a significant increase in the number of referrals, which was sustained. The initiatives also led to a sustained increase in the mean age of the patients at referral, predominantly due to an increase in the age of female patients referred. There was also an increase in the proportion of female patients referred. In the immediate aftermath of the introduction of change there was a transient decrease in the average eGFR at referral, a decrease in age of patients referred with an eGFR <15ml/min and an increase in the eGFR of patients >70yrs of age. CONCLUSIONS: The data demonstrates significant and sustained increase in numbers of referrals. In the short term this was associated with a reduction in referral of elderly patients with stage 5 CKD and an increase in elderly patients with mild renal impairment. In the longer term we saw an increase in referral of an older female population.

Severe hyperkalaemia: demographics and outcome.

BACKGROUND: Few studies have evaluated the prevalence of severe hyperkalaemia in unselected patient populations. We identified all episodes of severe hyperkalaemia occurring in 1 year, and described patient demographics, clinical response and outcome. We also assessed junior doctor knowledge of its causes and significance. MATERIALS AND METHODS: A retrospective interrogation of the database of the regional biochemical laboratory identified all episodes of severe hyperkalaemia (K≥ 6.5 mmol/L) occurring in 2011. The understanding of trainee doctors of the importance, causes and treatment of severe hyperkalaemia was assessed by structured questionnaire. RESULTS: Severe hyperkalaemia was recorded in 433 samples (365 patients) giving a prevalence of 0.11%. Thirty-six per cent of episodes occurred in patients under the care of a nephrologist, who were significantly younger than those not under the care of a nephrologist. In the nephrology cohort, 86% occurred in patients with chronic kidney disease (CKD), the majority of which had CKD Stage 5. In the non-nephrology cohort, only 65% occurred in the context of CKD, which was equally distributed between Stages 3 and 5 CKD. In both patient groups, roughly 50% of episodes occurred in association with acute kidney injury (AKI). Acute mortality (death within 48 h of documented severe hyperkalaemia) was higher in the non-nephrology compared with the nephrology cohort. Time to repeat serum potassium was influenced by the clinical setting with shorter time to repeat for acute care compared with ward settings. Assessment of trainee doctor's knowledge suggested significant deficiencies in relation to severe hyperkalaemia. CONCLUSIONS: The prevalence of severe hyperkalaemia was low and occurred predominantly in the context of CKD and/or AKI. The majority of episodes occurred in patients not under the care of a nephrologist. Variability in time to repeat serum potassium levels suggested deficiencies in care, and assessment of trainee doctor's knowledge suggests the need for further educational initiatives to highlight its importance.

Applying estimated glomerular filtration rate to an ageing population: are we in danger of becoming ageist?

BACKGROUND: Management of CKD is a major public health concern. The introduction of automated eGFR reporting has seen an increase in labelling of elderly patients with CKD. The prognostic significance of the CKD label in this population remains controversial. AIM: To investigate rates of specialist intervention in the over 75's to determine whether these patients may be more appropriately managed in primary care, relieving the burden of excessive outpatient visits in this population. METHODS: Retrospective review of patient notes and laboratory reports over 25 consecutive renal outpatient clinics within a single NHS trust. RESULTS: 546 patients were studied. The mean age of patient was 68.7 years (SD+/-14.9). The over 75's had more advanced renal disease compared to under 75's (mean eGFR 28.2 vs. 41.3 ml/min/1.73 m(2)), but there was no significant difference in eGFR stability between the older and younger cohort or in the overall rate of intervention (32.5% vs. 30.7% p=0.86). The over 75's had a lower mean haemoglobin (11.8 vs. 12.6, p=<0.001) necessitating greater EPO requirements (25.2% vs. 10.5%, p=<0.001). The greatest intervention was seen in the more advanced CKD patients (29% of CKD 3 vs. 55% of CKD 5), and in those with diagnoses requiring immunosuppression. CONCLUSION: Intervention to management may be predictable on the basis of specific diagnoses and advancing CKD stage but not by patient age. We can provide no evidence to suggest that elderly CKD patients are managed any differently to younger patients and in fact have a higher need for attention to and treatment of renal anaemia, validating their attendance in nephrology clinic.

Impact of chronic kidney disease management in primary care.

BACKGROUND: The introduction of eGFR reporting and publication of national CKD guidelines has led to major challenges in primary and secondary care, leading to an increase in the number of referrals to nephrology clinics. We have shown that introduction of a renal patient care pathway reduces nephrology referrals and enables managed discharges of CKD patients to primary care. The aim of this article is to examine the outcome of patients discharged to primary care to find out if there is an associated risk with increased discharge supported by the patient pathway. METHODS: The study was carried out within a single NHS Trust covering a population of 560,000. All patients discharged from the trust's renal outpatient clinic between June 2007 and July 2008 were identified. Patient notes and the local laboratory database systems were used to determine the source and timing of tests. RESULTS: A total of 31 new referrals and 57 regular follow-ups were discharged during this period. The median age of discharge was 67.5 years. Most subjects (60%) had CKD stage 3 at the time of discharge. A total of 23% of discharges were categorized as CKD stages 1, 2 or normal and 17% of patients had CKD stage 4. Overall, 93% had stable eGFRs prior to discharge, 77.5% of patients had blood pressure within threshold (140/90 according to UK CKD guidelines) and 97.7% of patients had haemoglobins >10 g/dl. Post-discharge 83% of patients had eGFRs recorded by their general practitioner and 92.6% of these were measured within appropriate time frames as per CKD guidelines. The majority of patients (82%) had either improved or stable eGFR post-discharge and only three patients had a significant decline in their eGFR. CONCLUSION: These data indicate that selected CKD patients can be appropriately discharged from secondary care and adequately monitored in primary care. Furthermore, we have shown that this was a safe practice for patients.

Renal quality outcomes framework and eGFR: impact on secondary care.

BACKGROUND: The prognostic significance of impaired renal function has driven the need for its early recognition and the widespread introduction of the estimated glomerular filtration rate (eGFR) reporting, and the incorporation of Chronic Kidney Disease (CKD) in the revised Quality Outcomes Framework (QOF) of the General Medical Services (GMS) contract in the UK. AIM: To characterize the effect of these changes on referral numbers and appropriateness to a nephrology service, and the impact of a newly introduced Map of Medicine(R)-based patient care pathway coupled to the systematic screening of all new referrals. METHODS: The study was carried out within a single NHS Trust covering five primary health care Local Health Boards and a population of 560,000. RESULTS: Introduction of eGFR reporting and CKD QOF domains was associated with a rapid 61% increase in new patient referral, and an increase in the mean age of the patients at referral from 63.0 +/- 18.1 to 69.1 +/- 18.5. The referrals did not correlate with the QOF reported prevalence of CKD. Systematic screening of new referrals demonstrated 36% to be either inappropriate or inadequate in terms of clinical information supplied. Introduction of the renal patient care pathway was associated with a fall in both the number of inadequate and total new referrals received. Overall 62% of all primary care practices registered with the Map of Medicine(R) and these sent a higher proportion of appropriate referrals and were less likely to generate referrals with inadequate information. The initiative also enabled managed discharges from secondary to primary care settings, freeing up outpatient capacity. CONCLUSION: The study describes the impact of the introduction eGFR reporting and revision of the GMS contract with Renal QOF, on patient referrals to a nephrology service. In addition, we provide evidence that a new management pathway has helped to regulate and proactively manage the increased demand within the current resources.

Y-box protein-1 controls transforming growth factor-beta1 translation in proximal tubular cells.

Transforming growth factor-beta1 (TGF-beta1) mRNA has low basal translational efficiency in proximal tubule cells; however, its translation is stimulated by profibrotic cytokines. We studied the role of the multifunctional Y-box protein-1 (YB-1) in regulating proximal tubule cell TGF-beta1 translation. Using RNA-electrophoretic mobility shift assays and ultraviolet crosslinking, we found two protein complexes of 50 and 100 kDa, which bound to the TGF-beta1 mRNA 5'-untranslated region. Supershift studies using antibodies to YB-1 showed that both sites contained YB-1 as did studies with recombinant YB-1, which demonstrated that it was sufficient to form both complexes. RNA competition experiments confirmed YB-1 binding to the two predicted binding sites; one with high affinity and the other with lower affinity. Strong basal YB-1 association with TGF-beta1 mRNA was found in proximal tubule cells, which decreased when platelet-derived growth factor was used to activate TGF-beta1 translation. In contrast, knockdown of proximal tubule cell YB-1 expression abrogated TGF-beta1 synthesis. Our results suggest that TGF-beta1 translation in proximal tubule cells requires YB-1 binding to a high-affinity site in the 5'-untranslated region of its mRNA; however, binding to a low-affinity site inhibits basal translation.

Characterization of hyaluronan cable structure and function in renal proximal tubular epithelial cells.

Alteration in the glycosaminoglycan hyaluronan (HA) has been demonstrated in numerous renal diseases. We have demonstrated that renal proximal tubular epithelial cells (PTCs) surround themselves in vitro with HA in an organized pericellular matrix or 'coat', which is associated with cell migration, and also form pericellular HA cable-like structures which modulate PTC-mononuclear leukocytes interactions. The aim of this study was to characterize potential regulatory mechanism in the assembly of PTC-HA into pericellular cables. HA cables are generated by PTCs in the absence of serum. Immunohistochemical analysis demonstrates the incorporation of components of the inter-alpha-inhibitor (IalphaI) family of proteins and versican into HA cables. Addition of an antibody to IalphaI/PalphaI (pre-alpha-inhibitor) inhibits cable formation. In contrast, inhibition of tumor necrosis factor-alpha-stimulated gene 6 (TSG-6) has no effect on cable formation, suggesting that their generation is independent of the known heavy-chain transfer activity of TSG-6. Overexpression of HAS3 is associated with induction of HA cable formation, and also increased incorporation of HA into pericellular coats. Functionally, this resulted in enhanced HA-dependent monocyte binding and cell migration, respectively. Cell surface expression of CD44 and trypsin-released cell-associated HA were increased in HAS3-overexpressing cells. In addition, hyaluronidase (hyal1 and hyal2) and bikunin mRNA expression were increased, whereas PalphaI HC3 mRNA expression was unchanged in the transfected cells. The data demonstrate the importance of IalphaI/PalphaI in cable formation and suggest that expression of HAS3 may be critical for HA cable assembly.

Butyrate modulates TGF-beta1 generation and function: potential renal benefit for Acacia(sen) SUPERGUM (gum arabic)?

Anecdotal evidence suggests that high fibre supplementation of dietary intake may have health benefits in renal disease related to alterations in circulating levels of short-chain fatty acids. The aim of the study was to examine the hypothesis that dietary manipulation may increase serum butyrate and thus have potential beneficial effects in renal disease. We examined the effect of dietary supplementation with a gum arabic sample of standardized molecular characteristics, Acacia(sen) SUPERGUM EM2 (SUPERGUM), on systemic levels of butyrate in normal human subjects. In an in vitro study, we also examined the potential role of butyrate in modifying the generation of the profibrotic cytokine transforming growth factor-beta (TGF-beta1) by renal epithelial cells. Following 8 weeks of dietary supplementation with 25 g/day of SUPERGUM, there was a two-fold increase in serum butyrate (n=7, P=0.03). In vitro work demonstrated that exposure of renal epithelial cells to elevated concentrations of butyrate suppressed both basal and stimulated TGF-beta1 synthesis. The action of butyrate was mediated by suppression of the extracellular signal-regulated kinase/mitogen-activated protein kinase signalling pathway. In addition, butyrate exposures reduced the response of renal epithelial cells to TGF-beta1 as assessed by luciferase activity of a TGF-beta-responsive reporter construct. Attenuation of TGF-beta1 signalling was associated with reduced phosphorylation of Smad 3 and decreased trafficking of TGF-beta1 receptors into signalling, non-lipid raft-associated membrane fractions. In conclusion, the data demonstrate that dietary supplementation with SUPERGU increased serum butyrate, which at least in vitro has beneficial effects on renal pro-fibrotic cytokine generation.

Mechanisms of irbesartan in prevention of renal lesion in streptozotocin-induced diabetic rats.

AIM: To investigate the mechanisms of angiotensin II receptor antagonist irbesartan (Irb) in prevention of renal lesion in streptozotocin (STZ)-induced diabetic rats. METHODS: Sprague-Dawley (SD) rats were randomly divided into three groups: normal control (group N), diabetic nephropathy (group DN), and diabetic nephropathy treated with Irb (group DNI). Diabetes was induced by injection of STZ ip after rats had received uninephrectomy. Blood glucose (BG), body weight (BW), urinary albumin excretion (Ualb), and 24-h proteinuria (24hUpro) were observed in the rats at week 4, 8, and 12, respectively. Creatinine clearance (Ccr), the kidney weight (KW), profile of kidney hypertrophy (KW/BW), renal tissue protein contents (RTP), glomerular area (AG), glomerular volume (VG), and width of glomerular basement membrane (GBM) were determined after the rats were sacrificed at week 12. Renal expression of connective tissue growth factor (CTGF) and transforming growth factor-beta1 (TGF-beta1) were determined by immunohistochemistry. RESULTS: There was no significant difference in BG between group DN and DNI (P >0.05). Irb prevented the increasing of Ualb excretion, 24 hUpro, and Ccr in diabetic rats ( P < 0.01). Furthermore, Irb markedly inhibited the increasing of KW, KW/BW, RTP, AG, and VG shown in diabetic rats (P <0.05, P <0.01, respectively). Irb prevented the thickening of GBM and immunostaining of CTGF (P <0.01). The extent of CTGF expression was positively correlated with the glomerular immunostaining for TGF-beta1 and size of VG (P <0.01). CONCLUSION: Irb exerts an early renal protective role to diabetic nephropathy, possibly through inhibition of renal hypertrophy and expression of CTGF.

Diabetic nephropathy: the central role of renal proximal tubular cells in tubulointerstitial injury.

Diabetic nephropathy is now the commonest cause of end stage renal disease and accounts for 30-40% of all patients requiring renal replacement therapy. Furthermore, the incidence of diabetic nephropathy continues to increase, in part due to the improved survival of type 2 diabetic patients as the cardiovascular mortality in this group declines (Ritz and Stefanski, 1996). Clinically incipient nephropathy is first manifest by the onset of persistent microalbuminuria, after which, overt diabetic nephropathy is heralded by the appearance of persistent proteinuria. Subsequently, there is a progressive decline in glomerular filtration rate (GFR) resulting, within 5 years, in end stage renal disease in 50% of patients (Hasslacher et al., 1989). The pathology of the renal lesions are similar in type I and II diabetes (Taft et al., 1994), although it has been suggested that there is more heterogeneity in type II diabetes (Chihara et al., 1986). Studies analysing structural-functional relationships have demonstrated that the development of proteinuria correlates with the degree of mesangial expansion (Mauer et al., 1984; White and Bilous, 2000). Although diabetic nephropathy was traditionally considered a primarily glomerular disease, it is now widely accepted that the rate of deterioration of function correlates best with the degree of renal tubulointerstitial fibrosis (Mauer et al., 1984, Bohle et al., 1991). This suggests that although in the majority of patients the primary event is a condition manifest by glomerular changes resulting in proteinuria, the long-term outcome is determined by events in the renal interstitium. With the increasing awareness of the importance of these pathological interstitial changes, interest has focused on the role of cells, such as the epithelial cells of the proximal tubule (PTC) or the interstitial myofibroblast, in the initiation of fibrosis. The aim of the present review is to analyse the available data supporting the role for the PTC in orchestrating renal interstitial fibrosis in diabetic nephropathy as a result of glucose-dependent alterations in PTC function. The potential for subsequent effects on PTC-fibroblast cross-talk will also be considered.

Translational regulation of renal proximal tubular epithelial cell transforming growth factor-beta1 generation by insulin.

We have previously demonstrated that the proximal tubular cell may contribute to the pathogenesis of renal interstitial fibrosis in diabetes. Transforming growth factor (TGF)-beta1 is one of a group of pro-fibrotic cytokines and growth factors, which have been associated with the development of interstitial fibrosis. The aim of the current study was to examine the effect of insulin on the generation of TGF-beta1 by proximal tubular cells. HK-2 cells were grown to confluence in the absence of insulin, and serum deprived for 48 hours before all experimental manipulations. Addition of insulin (5 microg/ml) to the culture medium led to a time-dependent increase in TGF-beta1 concentration in the cell culture supernatant, and increased incorporation of radiolabeled amino acids into TGF-beta1 suggestive of de novo TGF-beta1 protein synthesis. Addition of insulin did not alter TGF-beta1 mRNA expression as assessed by reverse transcriptase-polymerase chain reaction or Northern analysis. Insulin-induced increase in TGF-beta1 concentration was not abrogated by actinomycin D, however, stimulation by insulin, in the presence of cycloheximide led to a dose-dependent decrease in TGF-beta1 production. Addition of insulin had no effect on TGF-beta1 mRNA stability as assessed by actinomycin D chase, but led to increased binding of a cytoplasmic protein to a putative stem loop structure in the 5'-UTR of TGF-beta1 mRNA, previously implicated in the posttranscriptional control of TGF-beta1 synthesis. To address the functional significance of insulin-induced alteration in TGF-beta1 synthesis, we examined its effect on matrix turnover. Insulin stimulated type IV collagen gene expression and an increase in the concentrations of the type IV collagen laid down in the extracellular matrix. This increase in type IV collagen was abrogated when cells were stimulated by insulin in the presence of an anti-TGF-beta1-blocking antibody. In conclusion the data demonstrate that insulin may directly alter the production of TGF-beta1 by renal proximal tubular cells by a posttranscriptional mechanism, and that this may have implications for the increase in extracellular matrix that accompanies diabetic nephropathy.

Regulation of renal proximal tubular epithelial cell fibroblast growth factor-2 generation by heparin.

Progression of renal disease is closely correlated to the degree of renal interstitial fibrosis, and evidence is increasing that epithelial cells of the renal proximal tubule (PTCs) may contribute to its pathogenesis. Such cytokines as basic fibroblast growth factor (FGF-2) have been implicated in progressive renal injury, and we previously showed that PTCs are a source of this cytokine. FGF-2 is characterized by its high affinity for heparin, and numerous studies have suggested that heparin may modify the progression of renal disease. The current study examined how heparin influenced FGF-2 generation and bioactivity in the human renal epithelial PTC line, HK-2. Incubation of HK-2 cells with heparin led to a dose- and time-dependent increase in FGF-2 concentration in the culture supernatant that was not accompanied by alterations in FGF-2 messenger RNA expression, assessed by reverse-transcriptase polymerase chain reaction and Northern analysis. The heparin-induced increase in FGF-2 concentration was accompanied by a decrease in the amount of FGF-2 bound to the extracellular matrix, although this accounted for only a small proportion of the total FGF-2 generated. Induction of FGF-2 by 2-O-desulfated heparin, together with a reduction in total cell-associated FGF-2 and anti-FGF-2 antibody binding to fixed permeabilized cells after the addition of heparin, suggested that the FGF-2 released was mainly derived from a preformed intracellular source. That FGF-2 was predominantly derived from an intracellular pool was also confirmed by pulse chase experiments. The addition of heparin resulted in the generation of bioinactive FGF-2, judged by in vitro fibroblast proliferation. Conversely, heparitinase treatment of supernatant samples from heparin-treated cells and the addition of 2-O-desulfated heparin resulted in the generation of active FGF-2, suggesting that the generation of bioinactive FGF-2 was related to binding of FGF-2 by extracellular heparin after its release from cells. These data show that heparin depletes both the cell and surrounding matrix of FGF-2 and suggest that FGF-2 released from cells was mainly derived from a preformed intracellular source. Furthermore, FGF-2 released from epithelial PTCs after the application of heparin was bioinactive. This likely resulted from released FGF-2 binding to an excess of extracellular heparin. Results presented here therefore suggest a mechanism by which heparin, through its effect on depletion of matrix and cells of FGF-2 and its generation in an inactive form, may influence progressive renal interstitial fibrosis.

Regulation of renal proximal tubular epithelial cell hyaluronan generation: implications for diabetic nephropathy.

BACKGROUND: Proximal tubular cells (PTCs) contribute to pathological changes in the renal interstitium by the generation of cytokines and alterations in the composition of the extracellular matrix. Hyaluronan (HA) is a ubiquitous connective tissue polysaccharide that regulates cell function and tissue remodeling. In the current study, we investigated the regulation of HA generation by PTCs. METHODS: Primary cultures of human PTCs were grown to confluence and stimulated under serum-free conditions with either interleukin-1 (IL-1) or 25 mmol/L D-glucose. Alterations in HA generation were detected by enzyme-linked immunosorbent assay, and alterations in HA synthase gene expression were examined by reverse transcription-polymerase chain reaction. Subsequently, the mechanisms of IL-1 beta and glucose-induced alterations in HA were examined utilizing HK-2 cells. RESULTS: Stimulation of human PTCs (HPTCs) with either IL-1 beta or 25 mmol/L D-glucose led to a significant increase in the HA concentration in the culture supernatant. In contrast, stimulation of HPTCs with transforming growth factor-beta1, basic fibroblast growth factor, or platelet-derived growth factor-AB did not stimulate HA production. The addition of IL-1 beta or 25 mmol/L D-glucose also increased HA generation in HK-2 cells and was associated with the induction of HAS2 mRNA. HAS3 mRNA was constitutively expressed and was not influenced by the addition of either stimulus. HAS1 mRNA expression was not detected in either unstimulated or stimulated cells. Inhibition of gene transcription or protein synthesis abolished HA production in response to either IL-1 beta or glucose. Inhibition of nuclear factor-kappa B (NF-kappa B) activation either by sulindac or by the proteosome inhibitor (PSI) abrogated both IL-1 beta and glucose-mediated alteration in HA synthesis. CONCLUSION: This study demonstrates, to our knowledge for the first time, that increased HA synthesis in response to either IL-1 beta or elevated 25 mmol/L D-glucose is associated with NF-kappa B-activated transcription of HAS2.

Association of prolonged hyperglycemia with glomerular hypertrophy and renal basement membrane thickening in the Goto Kakizaki model of non-insulin-dependent diabetes mellitus.

The aim of the current study was to characterize the effects of prolonged hyperglycemia on renal structure and function using a model of non-insulin-dependent diabetes mellitus: the Goto Kakizaki (GK) rat, which does not have confounding variables, such as hyperlipidemia, obesity, or elevated blood pressure. The data show that hyperglycemia in this model was not associated with the development of significant proteinuria, but it was associated with the development of definitive age-dependent renal structural changes. These changes consisted of thickening of glomerular basement membrane at 35 weeks and tubular basement membrane. This thickening was accompanied by marked glomerular hypertrophy resulting from a parallel increase in total capillary luminal volume and mesangial volume, but fractional capillary and mesangial volumes remained unchanged. There was evidence of podocyte injury, as assessed by de novo expression of desmin. In contrast, there was no evidence of mesangial cell activation, as assessed by their de novo expression of alpha-SMA. Interstitial monocyte/macrophage influx increased significantly in GK rats at 12 weeks compared with Wistar controls. Glomerular macrophage infiltration was elevated significantly in 35-week GK rats. The structural changes described in the GK rat are similar to those described in prolonged non-insulin-dependent diabetes mellitus patients who have not developed overt renal disease. This model allows us to investigate further the mechanisms involved in the pathogenesis of the consequences of prolonged hyperglycemia.