Diagnostic utility of a-methylacyl COA racemase in prostate cancer of the Iranian population.

BACKGROUND: Considering the great variations in the reported prevalence of prostate cancer across the world possibly due to different genetic and environmental backgrounds, we aimed to determine the expression pattern and the diagnostic utility of α-methylacyl coenzyme A racemase (AMACR) among Iranian patients with prostate adenocarcinoma. MATERIALS AND METHODS: In this cross-sectional study, formalin-fixed paraffin-embedded tissues of 58 patients with a definitive pathologic diagnosis of prostatic adenocarcinoma were evaluated. The expression of AMACR, intensity, and extensity of its staining was determined in selected samples by immunohistochemical technique. RESULTS: AMACR expression was significantly higher in neoplastic compared to normal tissue (P < 0.05). The expression of AMACR was significantly associated with the age of the patients (P = 0.04). The intensity of the staining was associated with the grade of the prostate adenocarcinoma (P = 0.04). There was no significant relationship between AMACR expression and perineural invasion. The sensitivity, specificity, positive predictive value, and negative predictive value of AMACR were 90%, 96%, 96%, and 90%, respectively. CONCLUSION: Findings from our study indicate that AMACR could be used as a diagnostic tool for the diagnosis of prostate adenocarcinoma. However, due to false-positive staining in the mimicker of prostatic adenocarcinoma, it is recommended to use it in combination with basal cell markers.

An Easy and Fast Method for Production of Chinese Hamster Ovary Cell Line Expressing and Secreting Human Recombinant Activin A.

OBJECTIVE: Growth factors are key elements of embryonic stem cell (ESC) research. Cell line development in eukaryotes is a time-consuming procedure which usually takes 12-18 months. Here, we report an easy and fast method with which production of Chinese hamster ovary (CHO) cells that express and secrete recombinant Activin A, as a major growth factor in endo/mesoderm differentiation of embryonic stem cells is achieved within 3-4 weeks. MATERIALS AND METHODS: In this experimental study, we cloned human Activin A into the pDONR/Zeo gateway entry vector using the BP reaction. Activin A was subcloned next into the pLIX_403 and pLenti6.3/TO/V5-DEST destination vectors by the LR reaction. The result was the production of constructs with which 293T cells were finally transfected for virus production. CHO cells were transduced using viral particles to produce a cell line that secretes the His6- Activin A fusion protein. RESULTS: We developed a quick protocol which saves up to 3-4 weeks of time for producing recombinant proteins in CHO cells. The recombinant cell line produced 90 mg/L of functional Activin A measured in human ESC line Royan H5 (RH5), during in vitro differentiation into meso-endoderm and definitive endoderm. CONCLUSION: Our results showed no significant differences in functionality between commercial Activin A and the one produced using our novel protocol. This approach can be easily used for producing recombinant proteins in CHO.

Artificial Ureter in Patients with Extensive Ureteral Damage.

PURPOSE: loss of significant lengths of ureter when substitution with bowel or bladder fails is a disaster in urology. This study is conducted to evaluate the results of subcutaneous nephron-vesical bypass (SNVB) in ureteral damage of different etiologies. MATERIALS AND METHODS: Seventeen SNVB were employed in patients with ureteral injuries. We employed a device consisted of an internal silicone tube covered by a coiled PTFE tube to replace the ureter. This is called artificial ureter (AU). Proximal end of the AU was introduced in the kidney percutaneously, the tube was passed through a subcutaneous tunnel, while the distal end was inserted in the bladder through a small suprapubic incision. RESULTS: Follow-up ranged from six months to ten years. We removed the prosthetic ureter in one patient due to gross hematuria two months after insertion. One of the patients was reoperated two days after the procedure because of urinary leakage. In all other patients, the procedure was safe and effective. CONCLUSION: Subcutaneous nephron-vesical bypass is a safe and appealing alternative to a nephrostomy tube. This is a permanent device with no need for exchange. The technique can be applied in ureteral injuries due to various causes.

Cloning, expression, and functional characterization of in-house prepared human leukemia inhibitory factor.

OBJECTIVE: Leukemia inhibitory factor (LIF) plays important roles in cellular proliferation, growth promotion and differentiation of various types of target cells. In addition, LIF influences bone metabolism, cachexia, neural development, embryogenesis and inflammation. Human LIF (hLIF) is an essential growth factor for the maintenance of mouse embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) in a pluripotent, undifferentiated state. MATERIALS AND METHODS: In this experimental study, we cloned hLIF into the pENTR-D/ TOPO entry vector by a TOPO reaction. Next, hLIF was subcloned into the pDEST17 destination vector by the LR reaction, which resulted in the production of a construct that was transferred into E. coli strain Rosetta-gami™ 2(DE3) pLacI competent cells to produce the His6-hLIF fusion protein. RESULTS: This straightforward method produced a biologically active recombinant hLIF protein in E. coli that has long-term storage ability. This procedure has provided rapid, cost effective purification of a soluble hLIF protein that is biologically active and functional as measured in mouse ESCs and iPSCs in vitro. CONCLUSION: Our results showed no significant differences in function between laboratory produced and commercialized hLIF.

Concomitant surgical correction of severe stress urinary incontinence and anterior vaginal wall prolapse by anterior vaginal wall wrap: 18 months outcomes.

BACKGROUND: The aim of this study is to evaluate the outcome of an innovative, minimally invasive sling technique with autologous tissue in women with concomitant incontinence and anterior vaginal wall prolapse (AVWP). MATERIALS AND METHODS: Fifty-six women with stress urinary incontinence (SUI) or mixed urinary incontinence and AVWP were randomly assigned into two groups: In Group A (26 patients), anterior colporrhaphy (Kelly placation) and sling placement using a strip of anterior vaginal wall were performed, and in Group B (30 patients), transvaginal mesh correction of AVWP and tension-free vaginal tape (TVT) insertion (retropubic - craniocaudal route) using polypropylene mesh were carried out. The patients were followed-up for over 18 months and were assessed objectively using a 48 h frequency-volume chart, a 48 h pad test and a standardized stress test. Related surgical complications and outcomes were recorded and compared. RESULTS: Surgical cure rates for Group A and Group B at the first (3 days) and last (18 months) post-operative visits were 62% and 84%; and 54%, and 72%, respectively (P = 0.09 and 0.31). Complications occurred in 9 patients (44%) of Group B, but only 3 patients (12%) in Group A. CONCLUSION: Vaginal sling surgery using an anterior vaginal wall strip can improve SUI and in comparison with propylene mesh is associated with lower complication rates. Although, the surgical success rate of this technique is lower than T-Sling, larger studies with selected patients will help assess the suitable patients for this pelvic reconstructive surgery.

E-cadherin expression as a prognostic factor in transitional cell carcinoma of the bladder after transurethral resection.

PURPOSE: To analyze the role of negative versus positive immunoexpression of E-cadherin in recurrence rate of low-grade bladder tumors. MATERIALS AND METHODS: A total of 180 patients with unifocal, superficial, low-grade, papillary transitional cell carcinoma of the bladder were included in this study. The E-cadherin expression was evaluated using E-cadherin antibody. The patients were followed up for 36 months. Thereafter, recurrence rate of the tumor was compared between E-cadherin positive and negative groups. RESULTS: Of 180 low-grade carcinomas, E-cadherin immunoexpression was negative in 101 (56%) and positive in 79 (44%) patients. The recurrence rate in negative and positive groups was 65.6% and 37.9%, respectively. Negative in comparison with positive E-cadherin expression was associated with more disease recurrence (P = .045). CONCLUSION: There is an association between decreased E-Cadherin immunoexpression and tumor recurrence in low-grade and non-muscle invasive transitional cell carcinoma of the bladder.

One-suture, 1-knot technique in renal vascular transplant.

OBJECTIVES: We describe the results of our 1-suture, 1-knot technique for vascular anastomosis in renal transplant. This technique can be used for both of the arterial and venous anastomoses. MATERIALS AND METHODS: Between May 2006 and June 2008, a total of 386 renal transplants were done in our center, using a 1-suture, 1-knot technique. Intraoperative data including the warm and cold ischemic time, arterial and venous anastomotic time, and any early and late postoperative complications in the follow-up were recorded. RESULTS: Mean age of recipients was 37 years. Mean kidney warm and cold ischemia time was 4.8 and 26.2 minutes. Mean arterial and venous anastomotic time was 5.1 and 7.2 minutes. No vascular complications were seen in the early postoperative period. Delayed graft function was diagnosed in 36 patients, but a renal scan showed good perfusion of the allografts of these cases. In the mean follow-up of 18.5 months, we did not encounter any case of renal artery thrombosis or any suspected arterial stenosis. CONCLUSIONS: The 1-suture, 1-knot technique is a safe, rapid, and easy method for arterial and venous anastomosis of the renal allograft with low complication rates. It is especially valuable in obese patients and recipients with deep iliac fossa.

Transplantation of kidneys with duplicated ureters.

OBJECTIVE: Complete duplication of the ureters is one of the most common anomalies of the upper urinary tract. This study evaluates the urological complications after kidney transplantation from donors with complete duplicated ureters. MATERIAL AND METHODS: Between 1990 and 2008, 12 kidney transplantations from donors with complete ureteral duplication were performed in Shahid Labbafinejad Medial Center. The donors included 10 living and 2 deceased donors. The distal ends of the ureters were anastomosed to each other and then the modified Lich technique was used for ureteroneocystostomy. Two ureteral stents were placed in the ureters. RESULTS: No ureteral obstruction, urinary fistula or any other significant ureteral complications were observed in the recipients in the mean follow-up period of 6 years. CONCLUSION: This procedure is feasible and reliable for kidney transplantation of donors with duplicated ureters.

Generation of a novel high-affinity monoclonal antibody with conformational recognition epitope on human IgM.

As IgM is the first isotype of antibody which appears in blood after initial exposure to a foreign antigen in the pattern of primary response, detection, and quantification of this molecule in blood seems invaluable. To approach these goals, generation, and characterization of a highly specific mAb (monoclonal antibody) against human IgM were investigated. Human IgM immunoglobulins were used to immunize Balb/c mice. Spleen cells taken from the immunized animals were fused with SP2/O myeloma cells using PEG (polyethylene glycol, MW 1450) as fusogen. The hybridomas were cultured in HAT containing medium and supernatants from the growing hybrids were screened by enzyme-linked immunosorbent assay (ELISA) using plates coated with pure human IgM and the positive wells were then cloned at limiting dilutions. The best clone designated as MAN-1, was injected intraperitoneally to some Pristane-injected mice. Anti-IgM mAb was purified from the animals' ascitic fluid by protein-G sepharose followed by DEAE-cellulose ion exchange chromatography. MAN-1 interacted with human IgM with a very high specificity and affinity. The purity of the sample was tested by SDS-PAGE and the affinity constant was measured (K(a) = 3.5 x 10(9)M(-1). Immunoblotting and competitive ELISA were done and the results showed that the harvested antibody recognizes a conformational epitope on the mu chain of human IgM and there was no cross-reactivity with other subclasses of immunoglobulins. Furthermore, isotyping test was done and the results showed the subclass of the obtained mAb which was IgG(1)kappa.