To investigate the role of connective tissue growth factor (CTGF) in transdifferentiation of human renal tubular epithelial cell (HKC), in vitro cultured HKC cells were divided into 3 groups: negtive control, low dose CTGF-treated group (rh CTGF, 2.5 ng/ml) and high dose CTGF-treated (rhCTGF, 5.0 ng/ml). Then the expression of alpha-smooth muscle actin (alpha-SMA) were assessed by indirect immuno-fluorescence, and the percentage of alpha-SMA positive cells were assessed by flow cytometry. RT-PCR were also performed to examine the mRNA level of alpha-SMA. Upon the stimulation of different concentrations of rhCTGF, the expression of alpha-SMA were markedly stronger than that in negative controls. The percentages of alpha-SMA positive cells were significantly higher in the stimulated groups than that of negative controls (38.9%, 65.5% vs 2.4%, P<0.01). alpha-SMA mRNA levels were also up-regulated by the stimulation of rhCTGF (P<0.01). These results suggest that CTGF can promote the transdifferentiation of human renal tubular epithelial cells towards myofibroblast (Myo-F).
Actins/metabolism ; Cell Differentiation/*drug effects ; Cells, Cultured ; Epithelial Cells/*cytology ; Immediate-Early Proteins/*pharmacology ; Insulin-Like Growth Factor Binding Proteins/pharmacology ; Intercellular Signaling Peptides and Proteins/*pharmacology ; Kidney Tubules/*cytology

Objective To explore whether Cl2MDP-loaded gelatin particles can induce the depletion of macrophage or can depress the immunity of macrophage in SD rat models of immune thrombocytopenic purpura(ITP) to treat the ITP rats.Methods SD rats were intravenously injected with 150 ?L of anti-platelet serum once per 24 hr to induce the ITP model.The platelet count was persistently less than 50?109L-1 in these models.After intravenous injection of a group dose of Cl2 MDP-gelatin particles,the platelet counts of these models were examined at the time of 4 hr,24 hr,48 hr,72 hr and 96 hr,respectively.The pathological change of macrophages in spleen was observed by electron microscopy.Results Cl2MDP-loaded gelatin particles can significantly increase the platelet counts of ITP models from less than 50?109L-1 to mean 180?109L-1 in 24 hr after injection.The pathological changes of macrophages were seen under electron microscope,such as degeneration, necrosis and so on.Furthermore,rats pre-treated with Cl2 MDP-loaded gelatin particles can evade the decrease of platelet counts when they were injected anti-platelet serum.Conclusion Cl2 MDP-loaded gelatin particles can promptly,effectively restoreplatelet counts of ITP models to physiological level with a dose dependent manner.So,the targeting therapy of drug-loaded gelatin particles offers a new approach to treat ITP.

Objective:To investigate the value of 64-slice spiral computed tomography (64SCT) in preoperative evaluation of spinal vascular intervention.Methods: Seventeen patients with spinal vascular malformations(SVM) scheduled for spinal vascular interventional therapy in our hospital between January 2012-December 2014 were collected, all received 64-detector spiral CT angiography (64-MDCTA) before surgery, image postprocessing technologies such as volume rendering (VR), maximum intensity projection(MIP) and multiplanar reconstruction(MPR) were applied to conduct three-dimensional reconstruction, the traveling case of spinal cord feeding artery was analyzed and were compared with DSA test results.Results: All of the 17 patients clearly showed the abnormality of spinal cord blood vessels and lesions scope, the SVM classification and lesions scope were exactly the same as that of DSA and intraoperative performance, the coincidence rate was 100%;1 case of patient was not shown fistula, the rest of the patients clearly shown the draining veins around the spinal cord, open position and flow tendency of radiculomedullary artery, the coincidence rates were 100.00%, 94.12%%, 100.00%.Conclusion: 64-MDCTA is important for SVM patients to preoperative evaluation of spinal vascular intervention, for it can fully and accurately reflect the open position and flow tendency of radiculomedullary artery, spatial relationship with the surrounding vessels, spinal cord and vertebral situation, should be widely applied.

Objective To observe the changes of the blood culture results before and after implementing the quality control on blood culture .Methods The blood culture results in 1 year before implementing the quality control and in 1 year after implementing the quality control were performed the contrastive analysis .Results After implementing the quality control on blood culture ,the positive rate of blood culture was elevated by 2 .4% and the pollution rate was decreased by 8 .6% after the quality control .Conclu‐sion Implementing the total quality control on blood culture can improve the positive rate of blood culture and reduces the pollu‐tion rate of blood culture .The determination of contamination bacteria must combine the laboratory detection data with the clinical situation for ensuring to provide accurate and effective antimicrobial agents for clinic .

Objective To investigate the role of connective tissue growth factor (CTGF) in transdifferentiation of human renal tubular epithelial cells (HKC).Methods Cultured HKC were divided into 3 groups: (1) negative control; (2) low dose CTGF treated (rhCTGF 2. 5 ng/ml); (3) high dose CTGF treated (rhCTGF,5. 0 ng/ml). Expression of ?-smooth muscle actin (?-SMA) and fibronectin(FN) mRNA were measured by RT-PCR. Indirect immunofluorescence and flow cytometry methods were used to assess the level of intracellular ?-SMA protein. Concentration of FN secreted into the media was determined by ELBA. Results Upon the stimulations of different concentrations of rhCTGF,expression of ?-SMA and FN mRNA increased markedly(P

Objective To explore the silence of B-cell specific Moloney murine leukaemia virus insertion site 1(Bmi-1)by RNA interference on the proliferation of human leukemia cell line K562 and its mechanisms.Methods Small interfering RNA(siRNA)targeting Bmi-1 gene was designed and double-stranded siRNA was chemically synthesized.After double-stranded siRNA was transfected into K562 cells with Lipofectamine 2000,the proliferation of K562 cells was detected by MTT colorimetry,cell cycle was determined by flow cytometry,and the expression of Bmi-1 and P16 were analyzed by Western blotting.Results The siRNA targeting human Bmi-1 gene effectively prolonged the double time of K562 cells,increased the percentage of cells at G1 phase,and the expression of Bmi-1 was significantly down-regulated but the expression of P16 was up-regulated.Conclusion The siRNA targeting human Bmi-1 gene inhibits the proliferation of K562 cells,and up-regulates the expression of P16 in the cells.

OBJECTIVETo discuss the applicatioen of flap for the urethroplasty in the distal penile segment in the staged correction of severe hypospadias.

METHODSFrom Oct. 2004 to Aug. 2014, 25 cases with severe hypospadias were treated by staged urethroplasty for urethra reconstruction. The urethral meatus were located at peroscrotum in 4 patinets, at scrotum in 8 patients and at perineum in 13 cases. At the first stage, the urethral plate was divided and chordee was corrected. Then tubularized transverse island flap was used to prefabricate partial distal urethra. The defective urethra was repaired by using the Thiersch-Duplay principle at the second stage.

RESULTSAll patients completed both stages of the operation. The follow-up duration was 6-72 months (average, 24 months). In the first-stage, the modified tabularized transverse preputial island flap was performed on 10 patients, whereas the modified preputial double-faced island flap was performed on the other 15 patients. All of the prefabricated partial distal urethras had no evidence of stenosis or scarring. The result of the second-stage procedure was a complete penis with integrated urethral. All patients were satisfied with cosmetic and functional results. Neither stricture nor diverticula was observed. A good urinary stream during the urination was achieved in 19(19/ 25, 76%) patients. Five cases (5/25, 25%) developed urethrocutaneous fistula and one cases developed meatal stenosis (1/25, 4%) after the second stage repair.

CONCLUSIONSStaged urethroplasty using flap is a good choice for severe hypospadias. The successful rate is relatively high with good cosmetic and functional results.

Cicatrix ; Follow-Up Studies ; Humans ; Hypospadias ; surgery ; Male ; Penis ; surgery ; Perineum ; Reconstructive Surgical Procedures ; methods ; Scrotum ; Surgical Flaps ; Time Factors ; Urethra ; abnormalities ; surgery ; Wound Healing

Objective To explore the role of NG2 proteoglycan in the pathogenesis of glomerulosclerosis. Methods Eukaryotic expression vectors carrying the small hairpin RNA (shRNA) for NG2 mRNA , named as Psilencer-NG2, was constructed. Then, rat mesangial cells (RMC) were transfeeted with Psilencer-NG2, Psilencer-NC (negative control), pcDNA/NG2 (NG2 over-expressive vector) and empty vector pcDNA 1 respectively. The expression of endogenous NG2 in RMCs was examined by real-time PCR and Western blotting. Cell proliferation was analyzed by MTT assay and flow cytometry. The expression of laminin was detected by real-time PCR. Results Transfection of pcDNA/NG2 into HBZY-1 cells resulted in over-expression of NG2 mRNA and protein (P<0.05, P<0.05). Transfection of Psilencer-NG2 led to reduced expression of NG2 mRNA and protein (P<0.01, P<0.01). The expression of laminin β1 significantly increased due to overexpression of NG2 and decreased by treating with NG2 siRNA. According to MTT assay, overexpreasion of NG2 significantly stimulated the proliferation of mesangial cells while NG2 silencing inhibited it. NG2 increased the cell number in S phase and decreased the cell number in G0/G1 phase, while silencing NG2 induced the decrease of cell number in S phase and the increase of cell number in G0/G1 phase. Conclusion NG2 actively participates in the pathogenesis of glomerulosclerosis by stimulating proliferation of RMCs and increasing the deposition of ECM.

objective To assess the effect of aldosterone on the production of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9)and collagen Ⅳ in culture supematants of podocytes and the possible molecular mechanisms involved in the influence of aldosterone on the synthesis and degradation of extracellular matrix produced by podocytes. Methods Podecytes were treated with aldosterone at the concentration of 10-11, 10-9, 10-7 mol/L respectively. Cultured podocytes were examined at 24, 48 and 72 hours respectively. Spironolactone, a receptor antagonist of aldosterone, was added to observe the blocking effect on aldosterone. An inhibitor of TGF-β1 receptor was used to determine whether the effect of aldosterone on podocytes were mediated through TGF-β1 system. The enzymatic activities of MMP-2 and MMP-9 were assayed by gehtin zymography. Collagen Ⅳ 0.5 chain and TGF-β1 proteins released into culture supematants were assessed by Western blot and ELISA analysis. The adhesion rate of podocytes was monitored by flow cytometry. Results Aldosterone increased the activities of MMP-2 and MMP-9 in a dose- and time-dependent manner (P<0.05). Aldosterone decreased the level of collagen Ⅳ or5 chain protein in culture supernatants (P<0.05). Meanwhile, the expression of TGF-β1 was also increased (P<0.05). Spironolactone completely abolished the above-mentioned changes(P< 0.05). Blockage of TGF-β1 signaling with SB431542 prevented the aldosterone-induced upregulation of MMP-2 and MMP-9 as well as the downregulation of the collagen Ⅳ α5 chain protein and the adhesion rate of podocytes (P<0.05). Conclusions Aldosterone increases the activities of MMP-2 and MMP-9 but decreases the expression of collagen Ⅳ α5 chain and the adhension rate of podocytes possibly via TGF-β1 signaling pathway. Such alterations may contribute to glomerular podocyte injury associated with the GBM abnormality caused by the imbalance between matrix synthesis and degradation.

Objective To study the effects of CD2-associated protein (CD2AP) on podocyte adhesion and extension ability and to explore its possible mechanism. Methods Conditionally immortalized murine podocyte cell line was cultured in RPMI 1640 medium at 33℃permissive conditions. The podocytes were transfected with CD2AP small interfering RNA (siRNA) and serambing sequences labeled with fluorescein were taken as control. The transfected podocytes were trypsinized and seed into collagen IV coated plates. The relative cell adhesion and cell area were examined 90 min later. Apoptotic rates of CD2AP siRNA transfected podoeytes and different PAN concentrations incubated podoeytes were detected by flow cytometer. The distribution of F-actin was observed under laser scanning confoeal microscope. Nephrin protein expression and its phosphorylation level were examined by immunofluorescence and Western blot. Results The relative ceil adhesion of CD2AP siRNA transfected podocytes was apparently lower than that of control group[(41.72±6.07)% vs (64.46±8.53)%, P<0.05]. The cell area analysis had the similar result. The apoptotic rate of CD2AP siRNA transfected podocytes was significantly higher than that of the controls [(5.73±0.61)% vs (3.26±0.45)%, P<0.05]. 100 mg/L PAN could markedly induce podocytes to apoptosis and impair cell adhesion ability (P<0.05). Nevertheless, no significant difference was found in cell body spreading (P>0.05). The distribution of F-actin in CD2AP depletion podocytes was apparently altered. The expression of nephrin protein and its phosphorylation level was conspicuously descended to some degree (P<0.05). Conclusions CD2AP depletion facilitates podocyte apoptosis and impairs cell adhesion function. Cytoskeleton confusion and nephrin signaling weakness caused by CD2AP depletion may he partly responsible for the decline of cell adhesion and spreading.