Changes in plasma sclerostin level associated with use of a medium cut-off dialyzer in end-stage renal disease.

BACKGROUND: Larger middle molecules are important substances associated with cardiovascular complications in end- stage renal disease. Unfortunately, larger middle molecules are not reliably removed by a high-flux dialyzer. A medium cut-off (MCO) membrane could effectively remove larger middle molecules. This study aimed to identify the long -term effect of the MCO membrane for changes of larger middle molecules. METHODS: Thirty-four patients were prospectively analyzed for 12 months. The enrolled patients were divided into control and MCO groups. We measured the plasma levels of growth differentiation factor 15, sclerostin, and fibroblast growth factor 23 in larger middle molecules and those of biomarkers including small solutes. Single-pool Kt/V (spKt/V) and reduction ratios also were evaluated. RESULTS: Plasma sclerostin did not increase significantly in patients using the MCO dialyzer (135.3 [-637.7 to 908.3], p = 0.715). And there was a significant difference in change of plasma sclerostin level between the two groups (-1,646.9 [-3,015.2 to -278.7], p = 0.033). Furthermore, a negative association between calcium and sclerostin was not observed in the MCO group (r = -0.142, p = 0.587). Solute clearance of larger middle molecules in the MCO group was significantly higher. Moreover, spKt/V values for patients in the MCO group were significantly increased without albumin loss. Values are presented as mean (95% confidence interval [CI]) or adjusted mean (95% CI). CONCLUSION: The MCO dialyzer can increase dialytic adequacy and suppress the increase in plasma sclerostin level without significant albumin loss in patients with end-stage renal disease.

Human CD36 overexpression in renal tubules accelerates the progression of renal diseases in a mouse model of folic acid-induced acute kidney injury.

BACKGROUND: Acute kidney injury (AKI) is a risk factor for progression to chronic kidney disease, with even subclinical AKI episodes progressing to chronic kidney disease. Several risk factors such as preexisting kidney disease, hyperglycemia, and hypertension may aggravate renal disease after AKI. However, mechanisms underlying the progression of AKI are still unclear. This study identified the effect of human cluster of differentiation 36 (CD36) overexpression on the progression of folic acid-induced AKI. METHODS: Pax8-rtTA/tetracycline response element-human CD36 transgenic mice were used to elucidate the effect of human CD36 overexpression in the proximal tubules on folic acid-induced AKI. RESULTS: Results of histological analysis showed severely dilated tubules with casts and albuminuria in folic acid-treated transgenic mice overexpressing human CD36 compared with folic acid-treated wild-type mice. In addition, analysis of mRNA expression showed a significant increase in the collagen 3a1 gene in folic acid-treated transgenic mice overexpressing human CD 36 compared with folic acid-treated wild type mice. CONCLUSION: Human CD36-overexpressing transgenic mice showed severe pathological changes and albuminuria compared with wild-type mice. Moreover, mRNA expression of the collagen 3a1 gene increased in folic acid-treated transgenic mice. These results suggest that human CD36 overexpression is a risk factor of AKI and its progression to chronic kidney disease.

TGF-beta1-induced PINCH-1-ILK-alpha-parvin complex formation regulates mesangial cell proliferation and hypertrophy.

TGF-beta1-induced glomerular mesangial cell (GMC) injury is a prominent characteristic of renal pathology in several kidney diseases, and a ternary protein complex consisting of PINCH-1, integrin-linked kinase (ILK) and alpha-parvin plays a pivotal role in the regulation of cell behavior such as cell proliferation and hypertrophy. We report here that PINCH-1-ILK-alpha-parvin (PIP) complex regulates the TGF-beta1-induced cell proliferation and hypertrophy in cultured rat GMCs. When GMCs were treated with TGF-beta1 for 1, 2 and 3 days, the PIP complex formation was up-regulated after 1 day, but it was down-regulated on day 2. Cell numbers were significantly elevated on day 2, but dramatically decreased on day 3. In contrast, a significant increase in cellular protein contents was observed 3 days after TGF-beta1-treatment. TGF-beta1 induced early increase of caspase-3 activity. In GMCs incubated with TGF-beta1 for 2 days, cytosolic expression of p27(Kip1) was dramatically reduced, but its nuclear expression was remarkably elevated. A significantly decreased expression of phospho-Akt (Ser 473) was observed in the cells treated with TGF-beta1 for 1 day. TGF-beta1 induced early increase of phospho-p27(Kip1) (Thr 157) expression with subsequent decrease, and similar responses to TGF-beta1 were observed in the p38 phosphorylation (Thr 180/Thr 182). Taken together, TGF-beta1 differently regulates the PIP complex formation of GMCs in an incubation period-dependant fashion. The TGF-beta1-induced up- and down-regulation of the PIP complex formation likely contributes to the pleiotropic effects of TGF-beta1 on mesangial cell proliferation and hypertrophy through cellular localization of p27(Kip1) and alteration of Akt and p38 phosphorylation. TGF-beta1-induced alteration of the PIP complex formation may be importantly implicated in the development and progression of glomerular failure shown in several kidney diseases.

Ganglioside GM3 inhibits the high glucose- and TGF-beta1-induced proliferation of rat glomerular mesangial cells.

Abrupt proliferation of glomerular mesangial cells (GMCs) is a common feature in the early stage of diabetic glomerulopathy, and ganglioside GM3 (NeuAcalpha3Galbeta4Glcbeta1Cer) is thought to regulate the proliferation of many cell types. Recently, we have reported ganglioside GM3 as a modulator of glomerular hypertrophy in streptozotocin-induced diabetic rats []. This study examined whether modulation of cellular ganglioside GM3 could regulate the high glucose- and transforming growth factor-beta1 (TGF-beta1)-induced proliferation of GMCs. To pharmacologically modulate the cellular ganglioside GM3, GMCs originated from rat kidneys were cultured with exogenous ganglioside GM3 or d-threo-PDMP, an inhibitor of ganglioside synthesis, in the RPMI 1640 media containing normal (5.6 mM, NG) or high (25 mM, HG) glucose. HG, TGF-beta1 (10 ng/ml) and d-threo-PDMP (20 microM) significantly stimulated the mesangial cell proliferation, whereas these increments were remarkable attenuated by exogenous ganglioside mixture (0.1-0.2 mg/ml) or GM3 (20-100 microM) in a dose-dependent manner. The mesangial cell proliferation caused by HG, TGF-beta1 and d-threo-PDMP was closely correlated with decreases in both cellular sialic acid contents and ganglioside GM3 synthase activity. Based upon the mobility on high-performance thin-layer chromatography (HPTLC), GMCs showed a complex pattern of ganglioside expression that consisted, at least, of five different components of gangliosides, mainly ganglioside GM3. HG, TGF-beta1 and d-threo-PDMP induced a significant reduction of ganglioside expression with apparent changes in the composition of ganglioside GM3, and semi-quantitative analysis by HPTLC showed that ganglioside GM3 expression reduced to about 35-54% of control. These results provide a pathophysiological link between mesangial cell proliferation and ganglioside GM3 expression, indicating that exogenously added ganglioside GM3 inhibits the high-ambient glucose- and TGF-beta1-induced proliferation of cultured GMCs.

Modulation of cell proliferation and hypertrophy by gangliosides in cultured human glomerular mesangial cells.

Glomerular mesangial cells (GMCs) in diverse renal diseases undergo cell proliferation and/or hypertrophy, and gangliosides have been reported to play an important role in modulating cell structure and function. This study compared the effects of transforming growth factor-beta1 (TGF-beta1) and the effects of the application of exogenous gangliosides on GMCs and investigated whether the application of exogenous gangliosides regulated cellular proliferation and hypertrophy. Human GMCs were cultured with exogenous gangliosides and TGF-beta1 in a media containing 10% fetal bovine serum and in a media without the fetal bovine serum. Exogenous gangliosides biphasically changed the proliferation of human GMCs (0.1-1.0 mg/mL). A low concentration (0.1 mg/mL) of gangliosides mainly increased the number of human GMCs, whereas cellular proliferation was significantly reduced by raising the concentration of exogenous gangliosides. TGF-beta1 greatly reduced the number of human GMCs in a concentration-dependent manner (1-10 ng/mL). Serum deprivation accelerated the gangliosides- and TGF-beta1-induced inhibition of mesangial cell proliferation to a greater extent. Gangliosides (1.0 mg/ mL) and TGF-beta1 (10 ng/mL) both caused a significant increase in the incorporation of [3H]leucine per cell in the serum-deprived condition, whereas it was completely reversed in serum-supplemented condition. Similar results to the [3H]leucine incorporation were also observed in the changes in cell size measured by flow cytometric analysis. These results show that exogenous gangliosides modulate cell proliferation and hypertrophy in cultured human GMCs, and these cellular responses were regulated differently based on whether the media contained serum or not. Results from the present study raise new possibilities about the potential involvement of gangliosides in the development of mesangial cell proliferation and hypertrophy.

Decreases of ganglioside GM3 in streptozotocin-induced diabetic glomeruli of rats.

Ganglioside GM(3) (NeuAcalpha3Galbeta4Glcbeta1Cer) is known to regulate the proliferation of many cell types and to maintain the charge-selective filtration barrier of glomeruli. Based on these, this study examined whether altered expression of ganglioside GM(3) was pathologically related with glomerular hypertrophy and proteinuria occurring in diabetic human and rat kidneys. Diabetic rats were produced by intraperitoneal injection of streptozotocin (80 mg/kg, I.P.). At 15 days after the induction of diabetes, glomerular volume and fibrotic matrix were dramatically elevated, whereas glomerular sialic acid contents were significantly reduced compared with control. Based upon mobility on high-performance thin-layer chromatography (HPTLC) and reactivity to anti-GM(3) monoclonal antibody, normal glomeruli showed a complex ganglioside pattern that consisted of six different components of gangliosides, mainly GM(3), and diabetes caused a severe reduction of these gangliosides with apparent changes in the composition of major ganglioside GM(3). Semi-quantitative analysis by HPTLC showed that ganglioside GM(3) was reduced to 57% of control in diabetic glomeruli. A prominent immunofluorescence microscopy showed a dramatic disappearance of GM(3) expression in diabetic glomeruli. These results indicate that diabetic glomeruli can be characterized by decreases of glomerular sialic acid content and ganglioside GM(3) expression, which may cause loss of charge-selective filtration barrier in renal glomeruli. These changes may be account, at least in part, for the development of glomerular hypertrophy and proteinuria seen in the early stage of diabetic glomerulopathy.

Mechanism for the negative regulation of cell proliferation by ganglioside GM3 in high glucose-treated glomerular mesangial cells.

We recently identified ganglioside GM3 as a modulator of glomerular hypertrophy in streptozotocin-induced diabetic rats (Life Sci., 72: 1997-2006, 2003). This study examined whether alteration of ganglioside GM3 expression could modulate the high glucose-induced proliferation of glomerular mesangial cells (GMCs). GMCs isolated from rat kidneys were cultured under normal (5.6 mM) or high (25 mM) glucose condition for 24-72 hrs. Cell proliferation was predominantly stimulated when GMCs were cultured with high glucose as well as 20 microM of d-threo-PDMP, an inhibitor of ganglioside biosynthesis, for 24 hrs, whereas raising ambient glucose significantly reduced the mesangial sialic acid contents. Based upon mobility on high-performance thin-layer chromatography (HPTLC), GMCs showed a complex pattern of ganglioside expression that consisted of three major components of gangliosides, mainly GM3. High glucose induced a significant reduction of ganglioside expression with apparent changes in the composition of major ganglioside GM3, and semi-quantitative analysis by HPTLC showed that ganglioside GM3 was reduced to 62% of GMCs cultured under normal glucose condition. A prominent immunofluorescence microscopy using anti-GM3 monoclonal antibody also showed a dramatic disappearance of immunoreactivity in high glucose-treated GMCs. Moreover, high glucose significantly lowered the Km values of GM3 synthase (16 microM vs. 49 microM), but did not change the Vmax. These results provide the pathophysiological relationship between the high glucose-induced proliferation of GMCs and the decreased expression of ganglioside GM3, indicating a mechanism for the negative regulation of mesangial proliferation by ganglioside GM3. This mechanism may play an important role in the development of diabetic glomerulopathy.

Effects of adjuvant androgen on anemia and nutritional parameters in chronic hemodialysis patients using low-dose recombinant human erythropoietin.

BACKGROUND: Recombiant human erythropoietin (epoetin) has greatly contributed to improvement of the anemia of chronic renal failure patients on hemodialysis. However, the reduced erythropoietic effect to epoetin and its high cost have induced lots of supplementary treatments. Therefore, we performed a prospective study to evaluate the effects of adjuvant low-dose androgen therapy in patients using a lower-dose of epoetin than the commonly recommended dose on anemia and the nutritional parameters. METHODS: 17 patients of hemoglobin (Hgb) less than 9 g/dL even after being treated with 1,000 U epoetin subcutaneously (s.c.) 3 times per week on a stable status for more than 6 months, who were on hemodialysis at our institution were examined. They were injected with the same dose of epoetin s.c. and nandrolone decanoate 100 mg intramuscularly (i.m.) weekly for another 6 months. Blood test was performed every month before therapy for 6 months and after therapy for 6 months and the mean values were reviewed for comparison. RESULTS: Hgb (7.75 +/- 0.9 vs 8.99 +/- 1.39 g/dL, p < 0.01) and hematocrit (Hct) (23.68 +/- 2.85 vs 26.66 +/- 3.91%, p < 0.01) were apparently changed before and after adjuvant therapy. Hgb and Hct, weekly dose of epoetin were not statistically different in 9 male patients before and after adjuvant therapy. The weekly dose of epoetin was not statistically different in 8 female patients, but Hgb and Hct (8.02 +/- 0.6 vs 9.72 +/- 1.31 g/dL, 24.54 +/- 1.7 vs 28.74 +/- 3.06%, p < 0.01) were statistically different before and after adjuvant therapy. In comparison between male and female groups, weekly doses of epoetin and nandrolone decanoate were significantly greater in the female group than the male group (epoetin: 50.66 +/- 6.23 vs 61.18 +/- 8.76 U/kg/week, nandrolone decanoate: 1.69 +/- 0.2 vs 2.04 +/- 0.29 mg/kg/week, p < 0.05). CONCLUSION: Our data show that the adjuvant androgen therapy is effective for the anemia of hemodialysis patients who did not recover from anemia even after being continuously treated with low-dose epoetin.

alpha-Spinasterol isolated from the root of Phytolacca americana and its pharmacological property on diabetic nephropathy.

Based on an inhibitory activity-guided fractionation for the high glucose-induced proliferation of glomerular mesangial cells (GMCs), chloroform extracts of the roots of Phytolacca americana were found to contain alpha-spinasterol (C (29)H (48)O), a delta (7)-sterol. This phytosterol proved to be a potent inhibitor (IC (50) = 3.9 x 10 (-12) g/mL, 9.5 pmol/L) of glomerular mesangial cell proliferation caused by high-ambient glucose (5.6 mM vs. 25 mM), and its inhibitory potency was about 1,000 times higher than that of simvastatin, an HMG-CoA reductase inhibitor used as a positive control. alpha-Spinasterol also significantly reduced the increases of serum triglycerides, renal weight and urinary protein excretion in streptozotocin-induced diabetic mice, and these were comparable to the results observed in insulin-treated diabetic mice. Therefore, the results obtained in this study suggest that alpha-spinasterol has a significant therapeutic potential to modulate the development and/or progression of diabetic nephropathy.