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The aim of the study was to optimize the in vitro induction and expression of the human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and also study the processes of its denaturation, renaturation, and purification. The pGEX-6P-1/TRAIL114-281 plasmid was induced by isopropyl-β-D-1-thiogalactopyranoside (IPTG) in Escherichia coli BL21 (DE3), and the expressed target protein was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The protein expressed in the form of inclusion body was first denaturalized and then renaturalized by dilution and dialysis technique. GST-rTRAIL114-281 fusion protein was purified by Glutathione-Superflow Resin affinity chromatography and confirmed by Western blot. The molecular weight of GST-rTRAIL expressed in E. coli BL21 (DE3) was approximately 40 kDa. GST-rTRAIL was mainly expressed in the form of inclusion bodies. An optimum expression was induced by IPTG at a concentration of 0.2 mM for 8 h at 37 °C. Glutathione-Superflow Resin affinity chromatography yielded the purified GST-rTRAIL protein which was confirmed by Western blot using anti-GST mouse monoclonal antibody. The optimum prokaryotic cell expression of the human GST-rTRAIL was obtained by 0.2 mM IPTG induction for 8 h at 37 °C. The denatured inclusion body protein can be refolded by dilution and dialysis and purified by Glutathione-Superflow Resin affinity chromatography.

The most common malignancy of urinary tract is bladder cancer. It is one of the most widespread cancers of the world and ranks nine among frequent malignancies existing in world. The only solution to above burning problem is timely diagnosis at earlier stage, and the cancer research is being forwarded in this direction. There are various prominent gene modifications responsible for growth of bladder cancer. The present review is focused on recent advances in the field of cancer makers involving, genetic, urinary, pathological, etc., approaches to contain the deadly process of carcinogenesis. The present review provides an insight on the emerging biomarkers that could be developed to boost current bladder cancer detection strategies. This shall help timely diagnosis of this deadly disease at early stage, thereby, helping bladder cancer patients to fight against this iniquity of life.

Wusen Erlian Granules are a traditional Chinese medicine and we sought to determine the antiviral activity of Wusen Erlian Granules against Coxsackievirus B3 infection. First, cytotoxicity of Wusen Erlian Granules was determined in cultured cardiomyocytes isolated from day-old Wister rat pups. Later, cardiomyocytes were infected with Coxsackievirus B3 and the protective effect of Wusen Erlian Granules against cell injury was compared with that of ribavirin. Cell injury indicators including myoglobin, MB isozyme of Creatine Kinase, and cardiac Troponin were assessed by enzyme-linked immunosorbent assay (ELISA) and antiviral effect was assessed by MTT assay. We found that the 50 %-Toxic (TC50) and 50 %-Effective (EC50) concentrations of Wusen Erlian Granules were 394.05 and 30.26 μg/ml, respectively. Following infection of cardiomyocytes with Coxsackievirus B3, cell injury index of Wusen Erlian Granules, as determined by ELISA, was 125 μg/ml which yielded significant protection from virus-induced cell damage. The antiviral activity of Wusen Erlian Granules i.e., therapeutic index in MTT assay was higher (13.02) than that of ribavirin (6.93). It was, therefore, concluded that the Wusen Erlian Granules exerted better antiviral effect than ribavirin using Coxsackievirus B3 in vitro infection model in terms of rat cardiomyocytes protection from virus-induced cell injury.

Chronic allograft nephropathy (CAN) is the most common cause of chronic graft dysfunction leading to graft failure, our study investigates the expression and significance of p-Akt in the pathogenesis of CAN in rats. Kidneys of Fisher (F344) rats were orthotopically transplanted into Lewis (LEW) rats. The animals were evaluated at 4, 8, 12, 16, and 24 weeks post-transplantation for renal function and histopathology. Phosphorate Akt (p-Akt) protein expression was determined by Western blot and immunohistological assays. Our data show that 24-h urinary protein excretion in CAN rats increased significantly at week 16 as compared with F344/LEW controls. Allografts got severe interstitial infiltration of mononuclear cells at week 4 and week 8, but it was degraded as the time went on after week 16. Allografts markedly presented with severe interstitial fibrosis (IF) and tubular atrophy at 16 and 24 weeks. p-Akt expression was upregulated in rat kidneys with CAN, and the increase became more significant over time after transplantation. p-Akt expression correlated significantly with 24-h urinary protein excretion, serum creatinine levels, tubulointerstitial mononuclear cells infiltration, smooth muscle cells (SMCs) migration in vascular wall, and IF. It was concluded that p-Akt overexpression might be the key event that involved mononuclear cells infiltration and vascular SMCs migration at early stage, and IF and allograft nephroangiosclerosis at the late stage of CAN pathogenesis in rats.

The objective of this study was to compare surgical treatments for non-invasive bladder tumor. Hundred and forty patients with non-invasive bladder tumor were studied. Seventy-three patients were treated by transurethral resection of bladder tumor (TURBT) and Repeated-Transurethral Resection of Bladder Tumor (R-TURBT), while 67 by partial cystectomy. Operation time, blood loss, postoperative complications, and postoperative recurrence rate were better in the TURBT+R-TURBT group compared with the partial cystectomy group. Further, TURBT+R-TURBT offers advantages, such as simple surgical manipulation, less trauma, faster recovery, repeatedly performable procedure, and safety. In conclusion, this is an optimal therapy for treatment of non-invasive bladder tumor.

We wished to establish a mouse model of hypospadias using injections of estradiol benzoate for investigating the molecular mechanisms of hypospadias. Fifty timed pregnant mice were randomly divided into five study groups: A, B, C, D, and E. These groups were injected subcutaneously with estradiol benzoate mixed with sesame oil at, respectively, the doses of 0, 0.1, 0.5, 2.5, or 12.5 mg kg(-1) days(-1) from gestation day (GD) 12 to GD 16. The pups' mortality was recorded on the day of delivery. Urethras and positions of testes were examined on postnatal day 28. The numbers of live pups were significantly lower in the study groups D and E compared to study group A (p 0.01). Hypospadias was seen in groups C (3.3 %; 1/30), D (18.2 %; 4/22), and E (21.4 %; 3/14), while cryptorchidism was observed in groups C (10 %; 3/30), D (31.8 %; 7/22), and E (57.1 %; 8/14) on postnatal day 28. The experimental model of hypospadias induced by estradiol benzoate in the group D (2.5 mg kg(-1) days(-1)) was more reliable considering high mortality of the study group E. The dose of estradiol benzoate used in the group D is suitable for establishing mouse model of hypospadias.

To investigate the mechanism of apoptosis induced by BDI-1 monoclonal antibody (MAb) coupled Staphylococcal superantigen-A (SEA) in human bladder cancer cell line BIU-87. Human PBMC (effector cells) mediated cytotoxic killing of BIU-87 cells (target cells) was studied by culturing the BIU-87 cells in the presence of effector cells plus medium after their activation by treatment with SEA-targeted by MAb, SEA alone or vehicle (control). Proliferation and apoptosis of BIU-87 cells was measured after the treatments. Expression of Bax and Bcl-2 and cytokine concentration in co-culture supernatants were detected by Western blot and ELISA, respectively. Proliferation of MAb-targeted SEA BIU-87 cells decreased significantly (P < 0.05) as compared to control and SEA groups. Flow cytometry revealed apoptosis in SEA alone and more prominently in targeted-SEA treated in BIU-87 cells, which is significantly more than in controI cells (P < 0.05). In addition, Western blot analysis indicated that the ratio of Bax/Bcl-2 significantly increased by targeted SEA treatment, even at low concentration, as compared to cells treated with SEA alone or control cells (P < 0.05). However, there were no significant differences in IL-2, TNF-α and IFN-γ levels in the culture medium between SEA and targeted SEA groups, even though they are several folds higher than in control cells. SEA targeting by MAb significantly increases apoptosis in BIU-87 cells, possibly through the up-regulation of proapoptotic protein Bax and down regulation of antiapoptotic protein, Bcl-2.

The objective is to evaluate clinical indications and safety of tubeless percutaneous nephrolithotomy (PCNL) for the treatment of stones in upper urinary tract. A total of 231 subjects accepted to undergo tubeless PCNL, both micro-channel and standard operation were performed. After PCNL, renal gastrostomy tube was not placed also catheters and double-J stents were subsequently removed 3–5 and 14–28 days, respectively. Primary PCNL resulted in stone clearance of 98.27 %. There were five cases of postoperative hemorrhage with bleeding of about 500 ml, which was stopped by keeping patients in bed and medication. Three cases had pleural injury and thoracentesis plus drainage was performed in one case. Other two cases did not receive any special treatment due to a small amount of pleural effusion. No urinary leakage, perineal hematoma, infection, and other complications were observed in both of these cases. One and three months check-ups after discharge showed no recurrence of stones or perineal urinoma, and hydronephrosis were relieved in varying degrees. Thus, the application of tubeless PCNL is a safe and effective procedure for upper urinary tract calculi and the presence of stones was confirmed during operation showing good location with double-J stent and no obvious bleeding or damage.

We wished to study the protective effects of Wusen Erlian granules, a therapy from traditional Chinese medicine, in experimental viral myocarditis (VMC). Sixty mice were divided into six groups: control group, infection group, ribavirin group, and three Wusen Erlian groups, treated with low, intermediate, or high doses (4, 12, or 20 mg/kg) of Wusen Erlian. Control animals were intraperitoneally injected with culture medium, while animals in other groups received intraperinoneal injections of CoxB3 virus. The Wusen Erlian granules were intragastrically administered on days 0, 1, 2, 3, 4, and 5 after virus inoculation. The experiment was terminated 2 h after the final drug administration. The mice were weighed, and specimens were collected for detection of myocardial enzymes, measurement of organ index, and natural killer (NK) cell activity. The levels of creatine kinase isoenzyme, troponin, and myoglobin were significantly increased in infected animals (all p

To develop a transurethral endoscopy technique of the transurethral seminal vesiculoscopy to examine and treat seminal vesicle disease. A total of 61 patients with seminal vesicle disease were diagnosed and treated with the transurethral seminal vesiculoscopy through the distal seminal tracts and vesicles. 58 cases were successfully treated using transurethral seminal vesiculoscopy via the seminal vesicles. The operation took 25 ~ 85 min, with an average of (35.6) mins. In this group, seven cases were diagnosed as ejaculatory orifice cyst, 14 cases had blood clots in the seminal vesicles, and nine patients had stones in the seminal vesicles. All patients were treated properly. Follow-up occurred at 3 months, with two cases showing post-operative discomfort in perineal region. One patient had recurrence with seminal vesiculitis, which improved with treatment. Four infertile patients had a significant increase in sperm count and ejaculation volume and two of these patients were able to naturally inseminate within seven to 18 months post-surgery. This approach enables a new endoscopic technique with the transurethral seminal vesiculoscopy to diagnose and treat seminal vesicle disease through the normal anatomic pathway which can be easily performed with few post-operative complications.

To construct an adenovirus carrying SEA gene and regulated by telomerase reverse transcriptase (hTERT) and hypoxia-inducible factor (HIF) promoters and investigate its anti-tumor function in vitro, as well as its role in lymphocyte production. hTERT and HIF genes were cloned into adenovirus E1A and E1B shuttle plasmids. The control vector for SEA gene expression is under the regulation of CMV and SV40 promoters. Double regulation was obtained through homologous recombination. The positive clones of replicable adenovirus H2-SEA-Ad were selected by plaque assay. The adenovirus was purified, titrated, and DNA was verified by PCR. The obtained virus was used to infect EJ bladder tumor cells and the SEA Mrna, and protein expression was measured by RT-PCR, western blot, and immunofluorescence microscopy, respectively. Co-culture of lymphocytes and tumor cells was observed dynamically under microscope. The construction of shuttle plasmid p315-CSS-SEA was confirmed by PCR and DNA sequencing. Insertion of superantigen SEA gene in adenovirus (H2-SEA-Ad.SEA) was obtained by homologous recombination. In lymphocytes and tumor cell co-culture, the number of viable tumor cells in test groups was significantly lower than that in control group after 12, 24, and 48 h of treatment. Production of interleukin-2, interleukin-4, and tumor necrosis factor were higher in test groups than in control group. Expression of SEA gene in bladder tumor cells by adenoviral vector caused reduced tumor cell proliferation, as well as stimulation of inflammatory cytokine productions in co-cultures with lymphocytes.

The purpose of this study was to investigate the expression and significance of integrin-linked kinase (ILK) in the pathogenesis of chronic allograft nephropathy (CAN) in rats. For this, kidneys of Fisher (F344) rats were orthotopically transplanted into Lewis (LEW) rats. The animals were evaluated at 4, 8, 12, 16, and 24 weeks post-transplantation for renal function and histopathology. ILK protein expression was determined by Western-blot and immunohistological assays, and mRNA by RT-PCR. Our data show that 24-h urinary protein excretion in CAN rats increased significantly at week 16 as compared with F344/LEW controls. Allografts showed markedly increased mononuclear cells infiltration and presented with severe interstitial fibrosis and tubular atrophy at 16 and 24 weeks. ILK expression (protein/mRNA) was upregulated in rat kidneys with CAN, and the increase became more significant over time after transplantation. ILK expression correlated significantly with 24-h urinary protein excretion, serum creatinine levels, tubulointerstitial mononuclear cells infiltration, smooth muscle cells (SMCs) migration in vascular wall, and interstitial fibrosis. Therefore, it was concluded that ILK overexpression was the key event that involved mononuclear cells infiltration and vascular SMCs migration at early stage, and interstitial fibrosis and allograft nephroangiosclerosis at later stage of CAN pathogenesis in rats.