Protective effects and mechanisms of omega-3 polyunsaturated fatty acid on intestinal injury and macrophage polarization in peritoneal dialysis rats.

AIM: This study was conducted to investigate the chronic injury of peritoneal glucose injection on the peritoneum and intestine and the protective effects of omega-3 polyunsaturated fatty acid (ω-3PUFA) during peritoneal dialysis (PD). METHODS: Peritoneal dialysis animal models were established by intraperitoneal injection of 4.25% glucose for 28 days. Protein expression in ileum and peritoneum was measured by immunofloresence and immunohistochemistry. Protein expression in macrophages was measured by Western blot. Fibrosis was analyzed by Masson staining. RESULTS: Peritoneal dialysis significantly increased the structural injury and decreased junction-related protein ZO-1 and occludin expression in ileum, the expression of proteins relating to the activation of M2 (Erg2, IRF4), but not M1 (CD38, IRF5) macrophages. PD significantly increased the expression of TGF-ß1, VEGF and ALK5 protein in peritoneal tissues. PD significantly increased fibrosis (Masson staining) and the expression of fibroblast marker α-SMA in peritoneal tissues. Injection of macrophage clean reagent and ω-3PUFA significantly inhibited M2 activation, and decreased Masson staining, α-SMA, TGF-ß1, VEGF and ALK5 protein expression in peritoneal tissues in PD treated rats. ω-3PUFA injection significantly decreased PD-induced injury in ileum and normalized the expression of ZO-1 and occludin in the ileum of PD rats. CONCLUSION: Omega-3 fatty acids can provide a protective role on PD-induced peritoneal fibrosis and injury of the intestine.

Effects of Omega-3 Fatty Acids on Markers of Inflammation in Patients With Chronic Kidney Disease: A Controversial Issue.

Chronic kidney disease (CKD) is a global problem which contributes to a significant morbidity and mortality in China. Concomitant inflammatory state further boosts the mortality due to cardiovascular events in patients with CKD undergoing dialysis. There is a general notion that Omega-3 fatty acids including docosahexaenoic acids (DHA) and eicosapentaenoic (EPA) have certain health benefits perhaps via the regulation of inflammation. However, the anti-inflammatory effect of omega-3 fatty acids in patients with CKD is controversial. We analyzed the data of oral supplementation of omega-3 fatty acids in CKD patients by searching literature on database from inception to August 2016. The analysis included randomized controlled trials (RCTs) derived from multiple databases, and the effect of omega-3 fatty acids supplementation versus the control cohorts were compared. All of the data analysis was calculated by RevMan 5.2. A total of 12 RCTs involving 487 patients were included in the meta-analysis. Among them 254 patients received omega-3 fatty acids and 233 patients served as controls who received placebo. The meta-analysis revealed no statistical significance in serum levels of C-reactive protein (CRP) (SMD, -0.20; 95% CI, -0.44 to 0.05; P = 0.11), IL-6 (SMD, 0.00; 95% CI, -0.33 to 0.33; P = 0.99) and TNF-α (SMD, 0.14; 95% CI, -0.17 to 0.44; P = 0.38) between the omega-3 fatty acids supplementation group and control. This suggested that there is insufficient evidence to conclude the benefit of omega-3 fatty acids oral supplementation in reducing serum levels of CRP, IL-6 and TNF-α in patients with CKD.

Renoprotective approaches and strategies in acute kidney injury.

Acute kidney injury (AKI) is a major renal disease associated with high mortality rate and increasing prevalence. Decades of research have suggested numerous chemical and biological agents with beneficial effects in AKI. In addition, cell therapy and molecular targeting have been explored for reducing kidney tissue damage and promoting kidney repair or recovery from AKI. Mechanistically, these approaches may mitigate oxidative stress, inflammation, cell death, and mitochondrial and other organellar damage, or activate cytoprotective mechanisms such as autophagy and pro-survival factors. However, none of these findings has been successfully translated into clinical treatment of AKI. In this review, we analyze these findings and propose experimental strategies for the identification of renoprotective agents or methods with clinical potential. Moreover, we propose the consideration of combination therapy by targeting multiple targets in AKI.

Efficacy of triptolide on the apoptosis of tonsillar mononuclear cells from patients with IgA nephropathy.

AIMS: This study aimed to evaluate the extent of apoptosis of tonsillar mononuclear cells (TMCs) derived from patients with IgA nephropathy (IgAN) and the effects of triptolide (TP) on the apoptosis of these TMCs. METHODS: TMCs were isolated from tonsillar tissues of patients with IgAN or chronic tonsillitis (control group). Rates of TMCs apoptosis were measured by annexin V-fluorescein isocyanate (FITC)/propidium iodide (PI)-labeled flow cytometry (FCM). Expression levels of Bcl-2 family proteins were quantified by immunohistochemistry of fixed tonsillar sections and Western blot analyzes of TMCs lysates. TMCs from IgAN patients were treated 10, 20, or 30 ng/mL TP for 24 h and then evaluated for viability by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, for the percentage of apoptotic cells by FCM, and for the relative expression levels of Bcl-2 family proteins by Western blot analysis. RESULTS: Compared to TMCs from the control group, TMCs from the IgAN group demonstrated lower rates of apoptosis, higher expression levels of the anti-apoptosis proteins Bcl-2 and Bcl-xL, and lower expression levels of the pro-apoptosis protein Bax. Treatment of IgAN patient-derived TMCs with 10, 20, or 30 ng/mL TP for 24 h suppressed the viability and promoted the apoptosis of TMCs in a dose-dependent manner. Western blot analysis revealed a TP dose-dependent decrease in Bcl-2 and Bcl-xL expression levels, in parallel with increased Bax protein levels. CONCLUSION: TMCs from IgAN patients may be in a state of inhibited apoptosis mediated by Bcl-2 family proteins, which may be reversed by TP treatment.

[Role of protein phosphatase 2A in renal interstitial fibrosis].

BACKGROUND: To explore the role of protein phosphatase 2A (PP2A) in renal interstitial fibrosis by using rat model of unilateral ureteral obstructive (UUO) or cell model of human kidney proximal tubular epithelial (HK)-2 cells treated with transforming growth factor-ß1 (TGF-ß1).
 METHODS: 1) A total of 15 Sprague-Dawley rats were randomly divided into a sham group, a UUO group and an okadaic acid (OA) treated group (OA group) (n=5 in each group). The OA [30 µg/(kg·d)], diluted with 1.8% alcohol, was given to the rats in the OA group through gastric tube after at 72 h after the surgery, while the equal volume of 1.8% alcohol was given to the rats in the sham group and the UUO group. After sacrificing rats, the blood and kidney were collected to detect the renal function and the expression of PP2Ac, fibronectin (FN), collagen-I (Col-I), E-cadherin (E-cad) and α-smooth muscle actin (α-SMA) by immunohistochemistry, Western blot and RT-PCR, respectively; 2) The likely concentration of OA was determined by Trypan blue dye exclusive assay and methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. The HK-2 cells were incubated with serum-free Dulbecco's modified eagle medium (DMEM) for 24 h; then they were divided into a control group, a TGF-ß1 group (treated with 5 ng/mL TGF-ß1 for 24 h) and a TGF-ß1+OA group (treated with 5 ng/mL TGF-ß1 and 40 nmol/L OA for 24 h). The HK-2 cells were collected and the expression of PP2Ac, FN, Col-I, E-cad and α-SMA were detected by Western blot.
 RESULTS: 1) Compared with the sham group, the BUN and Scr in the UUO group increased (both P<0.05); compared with the UUO group, the BUN and Scr in the OA group decreased (both P<0.05); the expression of PP2Ac, FN, Col-I and α-SMA was up-regulated while the expression of E-cad was down-regulated in the UUO group compared with those in the sham group (all P<0.05). The expression of PP2Ac, FN, Col-I and α-SMA was down-regulated while the expressions of E-cad was up-regulated in the OA group compared with those in the UUO group (all P<0.05); 2) The likely concentration of OA was 40 nmol/L. Western blot showed that the expression of PP2Ac, FN, Col-I and α-SMA was up-regulated while the expressions of E-cad was down-regulated in the TGF-ß1 group compared with those in the control group (all P<0.05); the expression of PP2Ac, FN, Col-I and α-SMA were down-regulated while the expression of E-cad was up-regulated in the TGF-ß1+OA group compared with those in the TGF-ß1 group (all P<0.05).
 CONCLUSIONS: PP2A might be able to promote the renal interstitial fibrosis.
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Protective effects of curcumin on acute gentamicin-induced nephrotoxicity in rats.

BACKGROUND: Gentamicin-induced nephrotoxicity is one of the most common causes of acute kidney injury (AKI). The phenotypic alterations that contribute to acute kidney injury include inflammatory response and oxidative stress. Curcumin has a wide range biological functions, especially as an antioxidant. This study was designed to evaluate the renoprotective effects of curcumin treatment in gentamicin-induced AKI. METHODS: Gentamicin-induced AKI was established in female Sprague-Dawley rats. Rats were treated with curcumin (100 mg/kg body mass) by intragastric administration, once daily, followed with an intraperitoneal injection of gentamicin sulfate solution at a dose of 80 mg/kg body mass for 8 consecutive days. At days 3 and 8, the rats were sacrificed, and the kidneys and blood samples were collected for further analysis. RESULTS: The animals treated with gentamicin showed marked deterioration of renal function, together with higher levels of neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule 1 (KIM-1) in the plasma as compared with the controls. Animals that underwent intermittent treatment with curcumin exhibited significant improvements in renal functional parameters. We also observed that treatment with curcumin significantly attenuated renal tubular damage, apoptosis, and oxidative stress. Curcumin treatment exerted anti-apoptosis and anti-oxidative effects by up-regulating Nrf2/HO-1 and Sirt1 expression. CONCLUSIONS: Our data clearly demonstrate that curcumin protects kidney from gentamicin-induced AKI via the amelioration of oxidative stress and apoptosis of renal tubular cells, thus providing hope for the amelioration of gentamicin-induced nephrotoxicity.

Use of Ophiocordyceps sinensis (syn. Cordyceps sinensis) combined with angiotensin-converting enzyme inhibitors (ACEI)/angiotensin receptor blockers (ARB) versus ACEI/ARB alone in the treatment of diabetic kidney disease: a meta-analysis.

Ophiocordyceps sinensis (O. sinensis; syn. Cordyceps sinensis) has been used in clinical therapy for diabetic kidney disease (DKD) for more than 15 years. O. sinensis is a household name in china and it is available even in supermarket. However, the precise role of O. sinensis has not been fully elucidated with meta-analysis. The aim of this study was to review existing evidence on the effectiveness of O. sinensis for the treatment of DKD. We identified 60 trials involving 4288 participants. Overall, O. sinensis combined with ACEI/ARB had a better effect when compared to ACEI/ARB alone on 24 h UP (MD = -0.23 g/d, 95% CI: - 0.28 to -0.19, p < 0.00001), UAER (MD = -19.71 µg/min, 95% CI: -22.76 to -16.66, p < 0.00001), MAU (MD = -45.09 mg/d, 95% CI: -55.68 to -34.50, p < 0.00001), BUN (MD = -0.70 mmol/L, 95% CI: -1.02 to -0.39, p < 0.0001), SCr (MD = -8.37 µmol/L, 95% CI: -12.41 to -4.32, p < 0.0001), CRP (MD = -1.32 mg/L; 95% CI: -1.78 to -0.86; p < 0.00001), TG (MD = -0.51 mmol/L; 95% CI: -0.69 to -0.34, p < 0.00001), TC (MD = -0.64 mmol/L; 95% CI: -0.91 to -0.37, p < 0.00001), and SBP (MD = -2.01 mmHg; 95% CI: -3.45 to -0.58, p = 0.006). However, no effects were found for DBP, FBG, and HbA1C. This meta-analysis suggested that use of O. sinensis combined with ACEI/ARB may have a more beneficial effect on the proteinuria, inflammatory, dyslipidemia status as compared to ACEI/ARB alone in DKD III-IV stage patients, while there is no evidence that O. sinensis could improve the hyperglycemia status. However, with regard to low-quality and significant heterogeneity of included trials, to further verify the current results from this meta-analysis, long-term and well-designed RCTs with high-quality study are warranted to ascertain the long-term efficacy of O. sinensis.

[Effects of Shenxiong pill on infarct brain volumes and NF-kappaB expression in SD rats with middle cerebral artery occlusion (MCAO)].

OBJECTIVE: To investigate the effects of Shenxiong Pill on the infarct volume and expression of NF-kappaB in brains of rats with middle cerebral artery occlusion. METHODS: 169 SD rats were randomly divided into five groups: normal group, sham group, model group, cyclophosphamide group and Shenxiong Pill group. MCAO rat models were established by string ligation (for model, cyclophosphamide-treated and Shenxiong-treated groups). Rats in the Shenxiong Pill group was further randomly divided into sub-groups, receiving a range of high dose treatment (5 to 20 times of clinical dosage). Brains of the rats were examined 48 h or 72 h after interventions in a random order. Image processing software was used in the calculation of volume of cerebral infarction. Conventional HE staining was used for observation of brain tissue. Immunohistochemical method was used to determine NF-kappaB expression. RESULTS: Compared with the model group, rats treated with Shenxiong Pill and cyclophosphamide had lower infarct brain volumes (P < 0.05). NF-kappaB positive inflammatory cells were not found in the normal and sham groups. But the MCAO model rats had increased numbers of NF-kappaB positive inflammatory cells and higher integral optical density of NF-kappaB over time. Compared with the model group, lower numbers and expression of NF-kappaB positive inflammatory cells were found in those treated with Shenxiong Pill (P<0. 05). Higher dosage of Shenxiong was associated with lower numbers and expression of NF-gB inflammatory cells (P<0. 05). CONCLUSION: Shenxiong Pill can reduce pathological damage to brains as a result of cerebral ischemia, possibly through inhibiting the expression and activation of NF-kappaB.

Effect of L-carnitine therapy on patients in maintenance hemodialysis: a systematic review and meta-analysis.

BACKGROUND: L-Carnitine has been used as adjuvant therapy in hemodialysis (HD) patients for many years. However, there is controversy whether L-carnitine supplementation is beneficial. Therefore we performed a meta-analysis of randomized controlled trials (RCTs) to assess the effect of L-carnitine on HD patients. METHODS: RCTs of L-carnitine versus placebo for HD patients were searched from Medline, EMBASE and the Cochrane Central Register of Controlled Trials. We screened relevant studies according to predefined inclusion and exclusion criteria, and performed meta-analyses using Revman 5.1 software. RESULTS: Meta-analysis showed L-carnitine could not increase the total score of 36-item Short-Form Health Survey Questionnaire (SF-36) (SMD 0.76, 95 % CI -0.13 to 1.65, P = 0.09), and L-carnitine therapy did not improve serum C-reactive protein (SMD -0.37, 95 % CI -0.88 to 0.14, P = 0.16), oxidized low-density lipoprotein (SMD 0.04, 95 % CI -0.43 to 0.50, P = 0.87), albumin (SMD 0.25, 95 % CI -0.31 to 0.81, P = 0.38;), hemoglobin (SMD 0.23, 95 % CI -0.23 to 0.68, P = 0.33), cholesterol (SMD -0.24, 95 % CI -0.71 to 0.24, P = 0.33), triglycerides (SMD 0.02, 95 % CI -0.4 to 0.44, P = 0.91) or parathyroid hormone (SMD 0.21, 95 % CI -0.35 to 0.76, P = 0.46) levels. CONCLUSIONS: There is no evidence that L-carnitine can improve the inflammation, oxidative stress, nutrition, anemia, dyslipidemia, hyperparathyroidism status or quality of life in HD patients. However, given methodological limitations and lack of hard endpoints, high-quality, long-term randomized trials are required to fully elucidate the clinical value of L-carnitine administration in hemodialysis patients.

Is lead chelation therapy effective for chronic kidney disease? A meta-analysis.

The heavy metal lead (Pb) is a major environmental and occupational hazard. Epidemiological studies have demonstrated a strong association between lead exposure and the presence of chronic kidney injury. Some studies have suggested that chelation therapy with calcium disodium ethylenediaminetetraacetic acid (calcium disodium EDTA) might help decrease the progression of chronic kidney disease among patients with measurable body lead burdens. However, calcium disodium EDTA chelation in lead exposure is controversial due to the potential for adverse effects such as acute tubular necrosis. Therefore, we investigated the available randomized controlled trials assessing the renoprotective effects of calcium disodium EDTA chelation therapy. Our meta-analysis shows that calcium disodium EDTA chelation therapy can effectively delay the progression of chronic kidney disease in patients with measurable body lead burdens reflected by increasing the levels of estimated glomerular filtration rate (eGFR) and creatinine clearance rate (Ccr). There appears to be no conclusive evidence that calcium disodium EDTA can decrease proteinuria.

Low-dose paclitaxel ameliorates fibrosis in the remnant kidney model by down-regulating miR-192.

Transforming growth factor (TGF)-ß has been shown to play a central role in the development of tubulointerstitial fibrosis, which can be corrected via treatment with paclitaxel. The biology of microRNA (miR) can be modulated by paclitaxel. We hypothesized that paclitaxel may attenuate renal fibrosis in a rat model of remnant kidney disease by inhibiting TGF-ß induced-miRs. Rats in groups of 12 were subjected to 5/6 nephrectomy and received low-dose intraperitoneal injection of paclitaxel. Renal functions were assessed at 8 weeks. The TGF-ß signalling cascade and ECM proteins were evaluated by real-time polymerase chain reaction (TRT-PCR) and immunofluorescence microscopy. Animals with remnant kidneys developed hypertension, which was not relieved with paclitaxel treatment. However, paclitaxel treatment resulted in dampening the proteinuric response, reduction in serum BUN, creatinine levels and urine protein : creatinine ratio and normalization of creatinine clearance. These effects were accompanied by the inhibition of Smad2/3 activation, attenuation of renal fibrosis and normalization of integrin-linked kinase (ILK), COL(I)A1, COL(IV)A2 and α-SMA expression. Also, paclitaxel down-regulated the expression of miR-192, miR-217 and miR -377, while miR-15 was up-regulated in the remnant kidney. In vitro, in tubular epithelial cells (NRK-52E), paclitaxel also inhibited TGF-ß1-induced Smad2/3 activation and normalized ILK, COL(I)A1, COL(IV)A2 and α-SMA expression. Furthermore, ChIP analyses indicated that Taxol suppressed Smad3-mediated miR-192 transcriptional activity. Over-expression of miR-192 in NRK-52E mimicked the changes seen in the remnant kidney, while inclusion of miR-192 inhibitor in the culture medium blocked TGF-ß1-induced COL(I)A1 and COL(IV)A2 expression, while ILK and α-SMA were unaffected. These data suggest that low-dose paclitaxel ameliorates renal fibrosis via modulating miR-192 pathobiology and TGF-ß/Smad signalling.

Norcantharidin inhibits the expression of extracellular matrix and TGF-ß1 in HK-2 cells induced by high glucose independent of calcineurin signal pathway.

Norcantharidin (NCTD) was shown in our previous studies to attenuate renal tubulointerstitial fibrosis in rat models with diabetic nephropathy (DN). The aim of this study was to determine the effects of NCTD on the expression of extracellular matrix (ECM) and TGF-ß1 in HK-2 cells stimulated by high glucose and on calcineurin (CaN)/NFAT pathway. Whether or not the antifibrotic effect of NCTD on renal interstitium was dependent on its inhibition of CaN pathway was also investigated. Experimental concentrations of NCTD were verified by cytotoxic test and MTT assay. HK-2 cells were transfected with CaN small interference RNA (siRNA). The mRNA and protein expressions of FN, ColIV, TGF-ß1, and CaN in HK-2 cells were detected by real-time PCR and western blot. The CaN/NFAT pathway was examined by indirect immunofluorescence and western blot. Our study revealed that NCTD concentrations over 5 mg/l had overt cytotoxicity on HK-2 cells. Meanwhile, both 2.5 and 5 mg/l NCTD inhibited HK-2 cell proliferation (P < 0.05). NCTD inhibited the upregulation of FN, ColIV, and TGF-ß1 of HK-2 cells stimulated by high glucose (P < 0.05), and also significantly downregulated the expression of CaN mRNA and protein in HK-2 cells (P < 0.05). In addition, not only was the nuclear translocation of NFATc inhibited, but its protein level in the nucleus was also reduced. Following CaN siRNA transfection, CaN mRNA and protein expression were significantly decreased. In contrast, the protein levels of FN, ColIV, and TGF-ß1 increased in HK-2 cells stimulated by high glucose (P < 0.05). However, NCTD treatment downregulated their expression. These results indicated that NCTD could decrease the expression of ECM and TGF-ß1 in HK-2 cells stimulated by high glucose, downregulate CaN expression, and block the CaN/NFAT signaling pathway. However, the effect of NCTD on inhibition of the expression of ECM and TGF-ß1 was not associated with its inhibition of the CaN/NFAT pathway.

Norcantharidin attenuates tubulointerstitial fibrosis in rat models with diabetic nephropathy.

OBJECTIVE: To investigate the effects of norcantharidin (NCTD) on tubulointerstitial fibrosis of diabetic nephropathy (DN) in streptozotocin-induced rat model. METHODS: Sprague-Dawley rats were randomly divided into control group, model group, low-dose NCTD (0.05 mg/kg/day) group, and high-dose NCTD (0.1 mg/kg/day) group. The model group was induced by injection intraperitoneally with 30 mg/kg streptozotocin in 0.1 mol/L sodium citrate solution (pH 4.5), after high-calorie foods were given for 2 months. NCTD was administered daily after the DN rat model was built. Rats were sacrificed at the end of the third and the eighth week; renal fibrosis and the expression of FN, collagen IV, TGF-ß1, and calcineurin (CaN) were detected by Masson and immunohistochemistry staining, respectively. RESULTS: Tubulointerstitial fibrosis was observed in DN rats, this kind of pathological changes was ameliorated in NCTD treatment group (p < 0.05). The expressions of FN, collagen IV, and TGF-ß1 protein increased in the tubulointerstitial field of DN rats compared with the rats in control group. NCTD treatment could dose-dependently decrease their expression and reverse the fibrotic degree (p < 0.05). Meanwhile, the expression of CaN was detected in tubular fields of normal kidney and increased in the tubulointerstitial field in DN rats. However, NCTD downregulated its expression in a dose-dependent manner (p < 0.05). CONCLUSIONS: NCTD could downregulate FN, collagen IV, and TGF-ß1 expression in tubulointerstitial fields and attenuate tubulointerstitial fibrosis in the early stage of DN rats. NCTD also alleviated the expression of CaN in tubules in DN. The relationship between the role of NCTD's anti-tubulointerstitial fibrosis and its inhibition to CaN expression remains to be further elucidated.

[Influence of huangqi on TGF-beta 1 inducing extracellular matrix secretion of human peritoneal mesothelial cell].

OBJECTIVE: To study the effect of huangqi on peritoneal sclerosis and its possible mechanism. METHODS: Isolated human peritoneal mesothelial cells (HPMC) was cultured. Cultured HPMC were treated with F12 without serum (control group), F12 with TGF-beta 1 (TGF-beta 1 group), F12 with TGF-beta 1 and huangqi 100 micrograms/ml (huangqi 1 group), F12 with TGF-beta 1 and huangqi 1 mg/ml (huangqi 2 group). Fibronectin(FN) and plasminogen activator inhibitor-1 (PAI-1) were detected by ELISA method. The expression of FN mRNA, PAI-1 mRNA and bFGF mRNA was detected by RT-PCR method. RESULTS: 1. HPMC expressed FN, PAI-1, and bFGF. 2. Fn and PAI-1 significantly increased compared with control with 5 ng/ml TGF-beta 1 stimulated at 24 hours and 48 hours(P < 0.05); 3. The different concentration of huangqi decreased FN and PAI-1 expression compared with TGF-beta 1 group, especially 1 mg/ml huangqi at 24 hours(P < 0.05). 4. FN, PAI-1 and bFGF mRNA expression were higher in 12 hours after stimulation with TGF-beta 1 compared with control. FN, PAI-1 and bFGF mRNA expression were lower in 12 hours after stimulation with different concentration of huangqi (100 micrograms/ml and 1 mg/ml) than those in the TGF-beta 1 group. CONCLUSION: TGF-beta 1 promotes extracellular matrix synthesis and secretion of HPMC. Huangqi partly counteracts the synthesis of human peritoneal mesothelial cell's extracellular matrix by the stimulation of TGF-beta 1.