Caloric restriction alleviates aging-related fibrosis of kidney through downregulation of miR-21 in extracellular vesicles.

Glomerulosclerosis and renal interstitial fibrosis occur with the aging kidney. In this study, we examined the expression of miR-21, peroxisome proliferator-activated receptor(PPARα), hypoxia-inducible factor(HIF-1α) in the kidney of 3-month-old rats fed ad libitum (YAL), 24-month-old rats fed ad libitum (OAL) and 24-month-old rats subjected to a 70% calorie-restricted diet for 8 months (OCR). We found long-term caloric restriction (CR) ameliorated aging and aging-related fibrosis. CR ameliorated the increment of miR-21 and HIF-1α, as well as the decrement of PPARα in old ad libitum group. Human proximal tubular cells (HPTCs) presented phenotypes of senescence and epithelial to mesenchymal transition (EMT) under high-glucose conditions, in which senescence occurred earlier than EMT. Senescent cells secreted extracellular vesicles (EVs) which contained miR-21 into the recipient cells. Inhibiting miR-21 of donor cells prevented the occurrence of EMT in recipient cells. In addition, miR-21 induced EMT through targeting PPARα protein and consequently enhancing HIF-1α expression, although other pathways cannot be ruled out. These findings demonstrated that miR-21-containing EVs derived from the senescent cells could facilitate EMT of HPTCs via PPARα-HIF-1α signaling pathway. Long-term caloric restriction and caloric restriction mimetics alleviated aging-related-fibrosis of kidney through downregulation of miR-21.

Alleviation of senescence and epithelial-mesenchymal transition in aging kidney by short-term caloric restriction and caloric restriction mimetics via modulation of AMPK/mTOR signaling.

Renal fibrosis contributes to declining renal function in the elderly. What is unclear however, is whether epithelial-mesenchymal transition (EMT) contributes to this age-related renal fibrosis. Here, we analyzed indicators of EMT during kidney aging and investigated the protective effects and mechanisms of short-term regimens of caloric restriction (CR) or caloric restriction mimetics (CRMs), including resveratrol and metformin. High glucose was used to induce premature senescence and EMT in human primary proximal tubular cells (PTCs) in vitro. To test the role of AMPK-mTOR signaling, siRNA was used to deplete AMPK. Cellular senescence and AMPK-mTOR signaling markers associated with EMT were detected. CR or CRMs treatment alleviated age-related EMT in aging kidneys, which was accompanied by activation of AMPK-mTOR signaling. High glucose induced premature senescence and EMT in PTCs in vitro, which was accompanied by down-regulation of AMPK/mTOR signaling. CRMs alleviated high glucose-induced senescence and EMT via stimulation of AMPK/mTOR signaling. Activation of AMPK/mTOR signaling protected PTCs from high glucose-induced EMT and cellular senescence. Short-term regimens of CR and CRMs alleviated age-related EMT via AMPK-mTOR signaling, suggesting a potential approach to reducing renal fibrosis during aging.

Pathoanatomy and incidence of the posterolateral fractures in bicondylar tibial plateau fractures: a clinical computed tomography-based measurement and the associated biomechanical model simulation.

OBJECTIVES: The aim of our study is to evaluate the incidence and pathoanatomy of posterolateral fragments and analyze the associated fracture mechanism in bicondylar tibial plateau fractures. METHODS: From 1.1.2008 to 3.15.2012, all patients suffering bicondylar tibial plateau fractures were identified, scanned and analyzed at the Shanghai Clinical Trauma Center. Furthermore cadaver knees were selected into three groups of 30/60/90 knee flexion to simulate the posterolateral tibial plateau fracture by an impact device. RESULTS: One hundred and sixty-four (44.32 %) bicondylar tibial plateau fractures finally satisfied our requirements. Fifty-three and ninety-four cases were measured eventually in the groups of posterolateral split and depression. The posterolateral articular fragment proportion was 15.43 %. The posterolateral articular fragment angle showed an average of 12.94°. The posterolateral fragment cortical height was on average 2.96 cm. The posterolateral sagittal fragment angle averaged at 72.06°. Ninety-four cases were measured in the posterolateral depression group. The average posterolateral articular depression proportion was 16.74 %. The average posterolateral articular depression height was 2.47 cm. In the biomechanical modeling of such kinds of fracture patterns, posterolateral split fractures in 30° and 60° flexion are significantly more than those in 90° flexion. Posterolateral splits combined with anterolateral depression fractures in 30° flexion are significantly more than those in 90° flexion. CONCLUSION: The incidence of posterolateral fractures is 44.32 % in bicondylar tibial plateau fractures. The morphology of posterolateral area can be referenced for the surgeon in the future clinical work. The information is also helpful for the design of locking plate and fracture modeling in biomechanical test. In addition, that posterolateral split and posterolateral depression might be caused by different injury mechanisms. Different angles of knee flexion under the axial impact loading are possibly the interpretations for these two fracture patterns.

AMPK-mediated downregulation of connexin43 and premature senescence of mesangial cells under high-glucose conditions.

Diabetic nephropathy is associated with premature senescence. Our previous study showed that glomerular mesangial cells (GMCs) appeared to take on senescent phenotypes under high-glucose conditions in conjunction with the downregulation of connexin43 (Cx43). In this study, we investigated whether AMPK-mediated Cx43 expression and premature senescence in diabetic nephropathy are associated with mTOR activation. From in vivo and in vitro studies, we found decreased expression of Cx43 and p-AMPK but increased expression of p21 both in the glomeruli of diabetic nephropathy and in primary GMCs cultured in high glucose. Activating AMPK or inhibiting mTOR prevented the downregulation of Cx43 and reversed GMC senescence. Dominant-negative AMPK expression both reduced Cx43 expression and induced GMC senescence. Furthermore, AMPK regulated Cx43 expression and GMC senescence mainly through the inhibition of mTOR, although other pathways cannot be ruled out. This study demonstrated that AMPK signaling pathways play an important role in the regulation of the Cx43 expression that accompanies GMC senescence under high-glucose conditions.

Expression and displaying of ß-glucosidase from Streptomyces coelicolor A3 in Escherichia coli.

Two genes encoding ß-glucosidase from Streptomyces coelicolor A3(2) were cloned and expressed in Escherichia coli BL21 (DE3). Two recombinant enzymes (SC1059 and SC7558) were purified and characterized. The molecular mass of the purified SC1059 and SC7558 as determined by SDS-PAGE agrees with the calculated values (51.0 and 52.2 kDa, respectively). Optimal temperature and pH for the two enzymes were both at 35 °C and 6.0. SC7558 exhibited to be much more active than SC1059 under optimal conditions, and it was recombined with ice nucleation protein which could anchor on the surface of the cell. The optimal temperature and pH of the recombinant cells were 55 °C and 8.0, respectively. The resultant cells were to be used as material for immobilized ß-glucosidase, which is convenient to catalyze substrates in various complicated conditions.