Xian-Fang Meng

High Serum Uric Acid and Increased Risk of Type 2 Diabetes: A Systemic Review and Meta-Analysis of Prospective Cohort Studies

Objective Current evidence suggests high serum uric acid may increase the risk of type 2 diabetes, but the association is still uncertain. The aim of the study was to evaluate the association between serum uric acid and future risk of type 2 diabetes by conducting a meta-analysis of prospective cohort studies. Design and Methods We conducted a systematic literature search of the PubMed database through April 2012. Prospective cohort studies were included in meta-analysis that reported the multivariate adjusted relative risks (RRs) and the corresponding 95% confidence intervals (CIs) for the association between serum uric acid and risk of type 2 diabetes. We used both fix-effects and random-effects models to calculate the overall effect estimate. The heterogeneity across studies was tested by both Q statistic and I2 statistic. Begg’s funnel plot and Egger’s regression test were used to assess the potential publication bias. Results We retrieved 7 eligible articles derived from 8 prospective cohort studies, involving a total of 32016 participants and 2930 incident type 2 diabetes. The combined RR of developing type 2 diabetes for the highest category of serum uric acid level compared with the lowest was 1.56(95% CI, 1.39–1.76). Dose-response analysis showed the risk of type 2 diabetes was increased by 6% per 1 mg/dl increment in serum uric acid level (RR 1.06, 95% CI: 1.04–1.07). The result from each subgroup showed a significant association between serum uric acid and risk of type 2 diabetes. In sensitive analysis, the combined RR was consistent every time omitting any one study. Little evidence of heterogeneity and publication bias was observed. Conclusions Our meta-analysis of prospective cohort studies provided strong evidence that high level of serum uric acid is independent of other established risk factors, especially metabolic syndrome components, for developing type 2 diabetes in middle-aged and older people.

Connective tissue growth factor regulates the key events in tubular epithelial to myofibroblast transition in vitro.

Connective tissue growth factor (CTGF) has been reported to play an important role in mediating the profibrotic effects of transforming growth factor‐β (TGF‐β) in various renal diseases. To elucidate the role of CTGF in renal tubular epithelial‐myofibroblast transdifferentiation, we examined the expression of α‐smooth muscle actin (α‐SMA), vimentin, tenascin‐C, and collagen IV expression upon the stimulation of CTGF in cultured human proximal tubular epithelial cell line (HKC), and further investigated the effects of endogenous CTGF blockade on the transdifferentiation process induced by TGF‐β. It is revealed that upon the stimulation of recombinant human CTGF (rhCTGF, 2.5 or 5.0 μg/L), the expression of α‐SMA and tenascin‐C mRNA increased significantly (p < 0.01), while collagen IV gene expression decreased significantly (p < 0.01), all in a dose‐dependent manner. The percentage of α‐SMA‐positive cells was significantly larger in the rhCTGF‐stimulated groups than that in negative control (38.9%, 65.5% vs. 2.4%, respectively, p < 0.01) as confirmed by flow cytometry. Both cytoplasmic and secretory tenascin‐C expression was upregulated by the stimulation of rhCTGF (p < 0.01). Under this condition, collagen IV secreted into the culture media was lowered markedly (p < 0.01). On RT‐PCR analysis, TGF‐β1 upregulated CTGF gene expression, preceding that of α‐SMA. The α‐SMA mRNA expression induced by TGF‐β1 was significantly inhibited by CTGF antisense oligodeoxynucleotide (ODN) transfection (p < 0.01). With prolonged incubation time, CTGF antisense ODN also inhibited intracellular α‐SMA protein synthesis, as demonstrated by indirect immuno‐fluorescence. So it is concluded that CTGF could promote the transdifferentiation of human renal tubular epithelial cells towards myofibroblasts in vitro, both directly and as a downstream mediator of TGF‐β, and CTGF blockade would be a possible therapeutic target against tubulointerstitial fibrosis.

Role of NADPH Oxidase-Mediated Reactive Oxygen Species in Podocyte Injury

Proteinuria is an independent risk factor for end-stage renal disease (ESRD) (Shankland, 2006). Recent studies highlighted the mechanisms of podocyte injury and implications for potential treatment strategies in proteinuric kidney diseases (Zhang et al., 2012). Reactive oxygen species (ROS) are cellular signals which are closely associated with the development and progression of glomerular sclerosis. NADPH oxidase is a district enzymatic source of cellular ROS production and prominently expressed in podocytes (Zhang et al., 2010). In the last decade, it has become evident that NADPH oxidase-derived ROS overproduction is a key trigger of podocyte injury, such as renin-angiotensin-aldosterone system activation (Whaley-Connell et al., 2006), epithelial-to-mesenchymal transition (Zhang et al., 2011), and inflammatory priming (Abais et al., 2013). This review focuses on the mechanism of NADPH oxidase-mediated ROS in podocyte injury under different pathophysiological conditions. In addition, we also reviewed the therapeutic perspectives of NADPH oxidase in kidney diseases related to podocyte injury.

Thioredoxin-interacting protein mediates NALP3 inflammasome activation in podocytes during diabetic nephropathy

Numerous studies have shown that the NALP3 inflammasome plays an important role in various immune and inflammatory diseases. However, whether the NALP3 inflammasome is involved in the pathogenesis of diabetic nephropathy (DN) is unclear. In our study, we confirmed that high glucose (HG) concentrations induced NALP3 inflammasome activation both in vivo and in vitro. Blocking NALP3 inflammasome activation by NALP3/ASC shRNA and caspase-1 inhibition prevented IL-1β production and eventually attenuated podocyte and glomerular injury under HG conditions. We also found that thioredoxin (TRX)-interacting protein (TXNIP), which is a pro-oxidative stress and pro-inflammatory factor, activated NALP3 inflammasome by interacting with NALP3 in HG-exposed podocytes. Knocking down TXNIP impeded NALP3 inflammasome activation and alleviated podocyte injury caused by HG. In summary, the NALP3 inflammasome mediates podocyte and glomerular injury in DN, moreover, TXNIP participates in the formation and activation of the NALP3 inflammasome in podocytes during DN, which represents a novel mechanism of podocyte and glomerular injury under diabetic conditions.

Inhibition of ethanol-induced toxicity by tanshinone IIA in PC12 cells

AbstractAim:To observe the effects of tanshinone IIA (Tan IIA) on the neurotoxicity induced by ethanol in PC12 cells and to explore its protective role.Methods:PC12 cell survival was measured by MTT assay. The formation of reactive oxygen species (ROS) and lactate dehydrogenase (LDH) release were detected by 2′,7′-dichlorofluorescin (DCF) fluorescence and calorimetric method, respectively. The percentage of cell apoptosis was monitored by flow cytometry. The expression of p53 was detected by immuno-fluorescence and flow cytometry.Results:Ethanol significantly impaired the survival of PC 12 cells as demonstrated by MTT assay. Ethanol also induced significant ROS formation and increased LDH release. Pre-incubation with Tan IIA in the culture medium significantly reversed these changes. Ethanol caused cell apoptosis and the upregulation of p53 protein. The anti-apoptosis effects of Tan IIA on ethanol-induced toxicity were accompanied by the downregulation of pro-apoptotic p53 protein expression.Conclusion:Tan IIA can protect neurons from apoptosis and might serve as a potential therapeutic drug for neurological disorders induced by ethanol.

Regulation of CD2-associated protein influences podocyte endoplasmic reticulum stress-mediated apoptosis induced by albumin overload.

Proteinuria is an exacerbating factor of chronic kidney diseases, leading to glomerulosclerosis. However, the molecular mechanisms mediating protein overload-induced podocyte injury are poorly understood. Recent studies have shown that apoptosis mediated by endoplasmic reticulum (ER) stress participated in the progression of a variety of kidney diseases. In the present study, we investigated the role of CD2-associated protein (CD2AP) in protein overload-induced ER stress and subsequent podocyte apoptosis. Conditionally immortalized mouse podocytes were cultured in vitro and treated with different concentrations of bovine serum albumin (BSA). In addition, CD2AP eukaryotic expression vector or siRNA was transfected into podocytes before exposed to BSA. Albumin endocytosis and podocyte apoptosis were visualized by confocal microscopy. The subcellular organelles were observed by transmission electron microscopy. The expressions of GRP78, caspase-12 and CD2AP were detected by RT-PCR or Western blot analysis. It was found that albumin was endocytosed by podocytes in a time-dependent manner. Accumulation of albumin in podocytes induced ER stress and apoptosis in a concentration-dependent manner as indicated by upregulation of GRP78 and caspase-12. Meanwhile, the subcellular organelles were disrupted and the expression of CD2AP was downregulated by high concentration of albumin. Transfection of CD2AP eukaryotic expression vector into podocytes increased CD2AP expression, depressed GRP78 and caspase-12 expressions, and inhibited podocyte apoptosis. In contrast, transfection of CD2AP siRNA deteriorated the above changes induced by BSA. It is concluded protein overload induces podocyte apoptosis via ER stress and CD2AP may play a crucial role in albumin overload-induced ER stress and apoptosis in podocytes.

NADPH Oxidase-Induced NALP3 Inflammasome Activation Is Driven by Thioredoxin-Interacting Protein Which Contributes to Podocyte Injury in Hyperglycemia

Diabetic nephropathy (DN) is one of the major causes of end-stage renal disease, and previously we demonstrated that NALP3 inflammasome was involved in the pathogenesis of DN. Here we investigated the mechanisms of NALP3 inflammasome activation in podocyte injury during DN. We found that, besides the activation of NALP3 inflammasome and upregulated thioredoxin-interacting protein (TXNIP), the glomerular expression of gp91phox, a subunit of NADPH oxidase, was enhanced in DN mice simultaneously. Inhibiting NADPH oxidase abrogated NALP3 inflammasome activation, and IL-1β production and eventually protected podocytes from high glucose- (HG-) induced injury. TXNIP, an inhibitor of thioredoxin, acts as a suppressor for antioxidant defense system. Our observation indicated that in HG-exposed podocytes genetic deletion of TXNIP by shRNA reversed gp91phox overexpression and alleviated the injury of podocyte. Collectively, our findings proposed that HG-induced NADPH oxidase activation was driven by TXNIP which subsequently triggered NALP3 inflammasome activation in podocytes and ultimately led to podocyte injury, and blocking TXNIP/NADPH oxidase signaling may be a promising treatment for DN.

Electroacupuncture Increases CB2 Receptor Expression on Keratinocytes and Infiltrating Inflammatory Cells in Inflamed Skin Tissues of Rats

UNLABELLED Endogenous cannabinoids and peripheral cannabinoid CB2 receptors (CB2Rs) are involved in the antinociceptive effect of electroacupuncture (EA) on inflammatory pain. However, it remains unclear about how EA affects the expression and distribution patterns of peripheral CB2Rs in inflamed skin tissues. To study this, inflammatory pain was induced by local injection of complete Freunds adjuvant into the hindpaw of rats. The mRNA and protein levels of CB2Rs were quantified by using RTPCR and Western blotting, respectively. The distribution of CB2Rs on keratinocytes and immune cells recruited to the inflamed skin tissues was determined by using double-immunofluorescence labeling. Induction of tissue inflammation significantly increased the mRNA and protein levels of CB2Rs in the skin tissue. Also, both 2 Hz and 100 Hz EA, applied to GB30 and GB34, significantly increased the mRNA and protein levels of CB2Rs in inflamed tissues compared to the sham EA group. CB2Rimmunoreactivities were mainly distributed in keratinocytes, macrophages, and T-lymphocytes in the epidermis and dermis of the inflamed skin tissue. Inflammation caused a significant increase in the number of CB2R-immunoreactive keratinocytes, macrophages, and T-lymphocytes. Furthermore, compared to the sham EA group, EA at 2 or 100 Hz significantly increased the number of keratinocytes, macrophages, and T-lymphocytes with CB2R-immunoreactivity in the inflamed skin tissue. Therefore, our findings suggest that EA is associated with upregulation of local CB2Rs in the inflamed skin tissue. EA primarily potentiates the expression of CB2Rs on keratinocytes and infiltrating inflammatory cells at the site of inflammation. PERSPECTIVE This study shows that electroacupuncture increases the CB2 receptor expression on keratinocytes and infiltrating inflammatory cells in inflammatory skin tissues. This finding provides new evidence showing the potential role of CB2 receptors in the analgesic effect of acupuncture on inflammatory pain.

Role of CD2-associated protein in albumin overload-induced apoptosis in podocytes.

Proteinuria is a well‐established exacerbating factor of chronic kidney diseases. However, the harmful effects of protein overload on podocytes and the underlying mechanisms are still poorly understood. In the present study, we examined the effects of high concentrations of albumin on podocytes and investigated the role of CD2AP (CD2‐associated protein) in albumin overload‐induced podocyte apoptosis. Conditionally immortalized mouse podocytes were cultured in vitro and treated with different concentrations of BSA. In addition, CD2AP eukaryotic expression vector or siRNA (small interfering RNA) was transfected into podocytes before they were exposed to BSA. Podocyte apoptosis, expressions of active caspase‐3 (p17) and CD2AP, and the distribution of F‐actin cytoskeleton were detected by flow cytometry, Western‐blot analysis and fluorescent staining respectively. It was found that exposure of podocytes to BSA induced podocyte apoptosis in a concentration‐dependent manner that was accompanied by up‐regulation of active caspase‐3, the disruption of F‐actin cytoskeleton, and decreased expression of CD2AP. Transfection of CD2AP eukaryotic expression vector into podocytes increased CD2AP expression, partially restored F‐actin distribution, blocked active caspase‐3 expression and inhibited podocyte apoptosis. In contrast, transfection of CD2AP siRNA deteriorated the above changes induced by BSA. It is concluded that protein overload induces podocyte apoptosis via the down‐regulation of CD2AP and subsequent disruption of cytoskeleton of podocytes, and CD2AP may play an important role in protein overload‐induced podocyte injury.

Ethanol directly induced HMGB1 release through NOX2/NLRP1 inflammasome in neuronal cells

It has been reported that ethanol contributes to neuronal damage. However, the mechanisms mediating the actions of ethanol on neurons remain elusive. The present study was designed to test whether ethanol directly induced HMGB1 release and to explore the cellular and molecular mechanism mediating its action. It was found that ethanol increased significant HMGB1 release from SH-SY5Y cells and from cultured primary cortical neurons as detected by ELISA assay. Meanwhile, ethanol induced the expression of NOX2 subunits such as gp91 and p47(phox) and increased the activation of NLRP1 inflammasome. However, when cells were pretreated with NADPH oxidase inhibitor, apocynin, HMGB1 release was significantly decreased. Moreover, apocynin also prevented the activation of NLRP1 inflammasome as detected the levels of cleaved caspase-1. In addition, z-YVAD-fmk, an inhibitor of caspase-1, decreased the ethanol-induced HMGB1 release. It is concluded that ethanol-induced HMGB1 release is associated with NOX2/NLRP1 inflammasome signaling, which represents a novel mechanism of ethanol-associated neuron injury.