Fusion protein of retinol-binding protein and albumin domain III reduces liver fibrosis.

Activated hepatic stellate cells (HSCs) play a key role in liver fibrosis, and inactivating HSCs has been considered a promising therapeutic approach. We previously showed that albumin and its derivative designed for stellate cell-targeting, retinol-binding protein-albumin domain III fusion protein (referred to as R-III), inactivate cultured HSCs. Here, we investigated the mechanism of action of albumin/R-III in HSCs and examined the anti-fibrotic potential of R-III in vivo. R-III treatment and albumin expression downregulated retinoic acid (RA) signaling which was involved in HSC activation. RA receptor agonist and retinaldehyde dehydrogenase overexpression abolished the anti-fibrotic effect of R-III and albumin, respectively. R-III uptake into cultured HSCs was significantly decreased by siRNA-STRA6, and injected R-III was localized predominantly in HSCs in liver. Importantly, R-III administration reduced CCl4- and bile duct ligation-induced liver fibrosis. R-III also exhibited a preventive effect against CCl4-inducd liver fibrosis. These findings suggest that the anti-fibrotic effect of albumin/R-III is, at least in part, mediated by downregulation of RA signaling and that R-III is a good candidate as a novel anti-fibrotic drug.

PPARgamma agonist and angiotensin II receptor antagonist ameliorate renal tubulointerstitial fibrosis.

The peroxisome proliferator activated receptor (PPAR)gamma agonist is used as antidiabetic agent with antihyperglycemic and antihyperinsulinemic actions. Beyond these actions, antifibrotic effects have been reported. We examined antifibrotic effects of PPARgamma agonist and interaction with angiotensin receptor antagonist in the unilateral ureteral obstruction (UUO) model. After UUO, mice were divided to four groups: no treatment (CONT), pioglitazone treatment, L158809 treatment, and L158809+ pioglitazone treatment. On day 14, CONT mice showed severe fibrosis and all treated mice showed decreased fibrosis. The immunohistochmistry of PAI-1, F4/80 and p-Smad2 demonstrated that their expressions were increased in CONT group and decreased in the all treated groups compared to CONT. PAI-1 and p-Smad2 determined from Western blotting, among treated groups, was decreased compared to CONT group. The expression of TGF-beta1 from real time RT PCR showed markedly increased in the CONT group and decreased in all treated groups compared to CONT. These data suggest the pioglitazone inhibited tubulointerstitial fibrosis, however, the synergism between pioglitazone and L158809 is not clear. Considering decreased expression of PAI-1 and TGF-beta/Smad2 in the treated groups, PAI-1 and TGF-beta are likely linked to the decreased renal tubulointerstitial fibrosis. According to these results, the PPARgamma agonist might be used in the treatment of renal fibrotic disease.

Spironolactone prevents diabetic nephropathy through an anti-inflammatory mechanism in type 2 diabetic rats.

Aldosterone induces myocardial fibrosis and vascular inflammation via proinflammatory and profibrotic cytokines. The effect of spironolactone on renal inflammation and renal function was investigated in type 2 diabetic rats. For define the molecular mechanism of spironolactone, the effect of spironolactone on the synthesis of monocyte chemotactic peptide-1 (MCP-1) and its upstream transcription factor, NF-kappaB, was evaluated in cultured mesangial cells and proximal tubular cells. There were no changes in blood glucose concentration or BP after spironolactone treatment. Spironolactone treatment significantly reduced urinary albumin excretion and ameliorated glomerulosclerosis. Urinary levels of MCP-1 were significantly increased concurrently with renal expression of MCP-1, macrophage migration inhibitory factor, and macrophage infiltration. Spironolactone treatment significantly inhibited urinary excretion of MCP-1 as well as renal MCP-1 and migration inhibitory factor expression and macrophage infiltration. In addition, aldosterone induced upregulation of MCP-1 expression and NF-kappaB transcriptional activity in cultured cells, and spironolactone reduced both NF-kappaB activation and MCP-1 synthesis. Furthermore, NF-kappaB inhibition abolished aldosterone-induced MCP-1 production. Overall, these findings suggest that aldosterone-induced NF-kappaB activation leads to activation of proinflammatory cytokines, ultimately leading to renal injury in this model. These data suggest that mineralocorticoid blockade may be a potential therapeutic target in diabetic nephropathy.

Effect of IgA aggregates on transforming growth factor-beta1 production in human mesangial cells and the intraglomerular expression of transforming growth factor-beta1 in patients with IgA nephropathy.

BACKGROUND: Transforming growth factor-beta (TGF-beta) stimulates renal fibrosis in various renal diseases including IgA nephropathy. METHODS: We examined whether immunoglobulin A (IgA) stimulated TGF-beta1 synthesis in human mesangial cells (MCs), and whether this effect was mediated through the protein kinase C (PKC) pathway. We measured the intraglomerular TGF-beta1 mRNA expression by using competitive RT-PCR, and this was compared with various parameters in IgA nephropathy patients. RESULTS: The IgA aggregate increased the TGF-beta1 mRNA expression in MCs, while this expression was not affected by the culture media or IgG aggregate. Phorbol 12-myristate 13-acetate and calphostin C did not influence the TGF-beta1 mRNA expression that was increased by the IgA aggregate. Intraglomerular TGF-beta1 mRNA expression was significantly correlated with creatinine clearance (r = -0.764, p = 0.027), daily proteinuria (r = 0.781, p = 0.022), serum creatinine (r = 0.884, p = 0.004), and tubulointerstitial changes (r = 0.809, p = 0.015). Glomerular TGF-beta1 mRNA expression was associated with an increased tendency for glomerulosclerosis (r = 0.646, p = 0.084). After 4 years, patients with a high expression of intraglomerular TGF-beta1 mRNA showed a tendency for an decrease of their renal function. CONCLUSION: The IgA aggregate increased TGF-beta1 mRNA expression in MCs, and this was independent of the PKC pathway. The evaluation of intraglomerular TGF-beta1 mRNA expression could be useful in predicting the progression of IgA nephropathy.

Effect of retinoic acid in experimental diabetic nephropathy.

Although the pathogenetic mechanism of diabetic nephropathy has not been elucidated, an inflammatory mechanism has been suggested to contribute to its progression. Monocyte chemoattractant peptide (MCP)-1 attracts macrophages and T cells, and ultimately injures renal tissue. In early diabetic nephropathy, urinary excretion of MCP-1 was elevated, and increased as renal damage became more severe. Podocytes are expected to have an inflammatory role in diabetic nephropathy, as the surface expression of chemokine receptors such as CCR and CXCR on these cells has been recently reported. Although retinoid (retinal), a known anti-inflammatory agent, has been reported to be beneficial in some experimental models of renal disease, it has not been determined to prevent disease progression in diabetic nephropathy. We investigated the effects of all-trans retinoic acid on the production of MCP-1 under high glucose conditions in cultured mouse podocytes. We also evaluated whether all-trans retinoic acid inhibits inflammatory changes and improves renal function during the early stages of diabetic nephropathy in streptozotocin-induced diabetic rats. In cultured podocytes, high glucose stimuli rapidly upregulated the MCP-1 mRNA transcript and protein release. Treatment with retinoic acid tended to suppress the MCP-1 gene transcript, and significantly inhibited MCP-1 protein synthesis induced by high glucose stimulation. Urinary protein excretion and the urinary albumin : creatinine ratio (ACR) were significantly higher in diabetic rats 4 weeks after the induction of diabetes mellitus compared with control rats, and retinoic acid treatment markedly decreased both proteinuria and urinary ACR (proteinuria: 1.25+/-0.69 vs 0.78+/-0.72 mg/mgCr, P=0.056; urinary ACR: 0.47+/-0.25 vs 0.21+/-0.06 mg/mgCr, P=0.088). Urinary excretion of MCP-1 was rapidly increased 2 days after induction of diabetes mellitus in diabetic rats, and further increased until rats were 4 weeks of age, compared with control rats. Retinoic acid treatment resulted in 30% reduction of the urinary level of MCP-1 compared with vehicle-treated diabetic rats (119.3+/-74.2 vs 78.1+/-62.7 pg/mgCr, P=0.078). Immunohistochemistry revealed a significant increase in staining for MCP-1 and anti-monocyte/macrophage (ED-1) protein in the diabetic kidney, and retinoic acid treatment significantly suppressed intrarenal MCP-1 and ED-1 protein synthesis. In conclusion, podocytes are involved in the inflammatory reaction under diabetic circumstances, and these reactions were suppressed by retinoic acid. Retinoic acid also suppressed inflammatory changes in the diabetic rat kidney, and decreased proteinuria in diabetic rats. These results suggest that retinoic acid may have renoprotective effects in the early stages of diabetic nephropathy through an anti-inflammatory activity.

Insomnia in diabetic hemodialysis patients. Prevalence and risk factors by a multicenter study.

BACKGROUND: Insomnia is one of the most common problems in dialysis patients, and likely to contribute impairment in quality of life, which has a positive correlation with patients' survival. In diabetic patients, morbidity and mortality are substantially higher than in the nondiabetic counterparts, and also the incidence of sleep disturbances. However, there is no means to predict sleep disturbance in the dialysis patients especially in diabetics. To define the prevalence and risk factors for insomnia in diabetic patients on hemodialysis, we undertook a cross-sectional multicenter study. METHODS: Eighty-two diabetic patients (50 men/32 women, aged 58.7 +/- 9.23 years) on maintenance hemodialysis for more than 6 months from 12 different hospitals were enrolled. The demographic data, subjective symptoms, depression scale, and insomnia were assessed by questionnaires, and lean body mass, BMI, Kt/V, subjective global assessment, nursing assessment score (NAS), and biochemical parameters were examined. RESULTS: The number of patients with and without insomnia were 56 and 26, respectively, which amounted to 68.2% for insomnia. NAS (28.1 +/- 3.81 vs. 30.8 +/- 2.88, p = 0.002), serum albumin concentration (3.82 +/- 0.44 vs. 4.09 +/- 0.36 g/dl, p = 0.008), and depression scale (25.2 +/- 12.1 vs. 18.9 +/- 10.3, p = 0.025) were significantly different between them. Patients with insomnia were older (60.5 +/- 9.0 vs. 56. 1 +/- 9.60 years, p = 0.053) and felt pain (38.5 vs. 15.3%, p = 0.06) more frequently than those without insomnia. The scale of depression was correlated with NAS (r = -0.455, p < 0.001) and the serum albumin concentration was correlated with NAS (r = 0.337, p = 0.002). NAS, age, and serum albumin concentration were the major risk factors for insomnia in logistic regression analysis. CONCLUSION: The prevalence of insomnia in diabetic hemodialysis patients was 68.2%. Age, nutritional status, and depression were the major risk factors for sleep disturbance in diabetic patients.