Introducing a frameshift mutation to the POL sequence of HIV-1 provirus and evaluation of the immunogenic characteristics of the mutated virions (RINNL4-3).

Inactivation of the reverse transcriptase (RT) and integrase (IN) enzymes can abolish the replication of the human immunodeficiency virus (HIV) and, thus, its infectivity. Here, inactivated HIV particles convenient for designing virus-like particle (VLP) based vaccines have been produced. Inactivated HIV-provirus was created by introducing a frame shift mutation. HIV provirus DNA was cut in the pol region by Age I restriction enzyme, followed by filling of sticky ends using the Klenow fragment before ligation. The resulting plasmid was named as pRINNL4-3. HEK-293T cells were used as producer, after being transfected with the modified plasmid. Viral particle production and biological activity were assayed by virus capsid protein (p24) quantification and syncytium formation in MT2 cells, respectively. The immunogenicity of the RINNL4-3 virions was investigated in a mouse model. The mutation was expected to inactivate the virus RT and IN enzymes. The results showed that the VLPs were assembled, as measured by the p24 load of the culture supernatant, and contained functional envelope proteins (Env) as monitored by the syncytium formation. However, these VLPs had no ability to infect target MT2 cells, as well as their VSVG (vesicular stomatitis virus-glycoprotein) pseudotyped counterparts infected HEK-293T cells. A high level of antibody response was observed in immunized mice. Since RINNL4-3 virions are replication incompetent, they are convenient for production and use in biomedical studies. Also, RINNL4-3 is a candidate for a vaccine development due to it contains envelope and structural virus proteins which are crucial for triggering neutralizing antibodies and the cellular immune response.

Inhibition of HIV and HSV infection by vaginal lactobacilli in vitro and in vivo.

BACKGROUND AND THE PURPOSE OF THE STUDY: The cervico-vaginal mucosa which is populated with microflora (mostly includes lactobacilli) is the portal of entry for sexually transmitted pathogens. METHODS: The in vitro anti-viral effect of vaginal and non-vaginal lactobacillus was evaluated using single cycle HIV-1 replication and HSV-2 plaque reduction assays. The XTT proliferation assay was used to monitor the cellular toxicity. The in vivo anti-HSV-1 activity was evaluated in BALB/c mouse model by monitoring skin lesion and immune response development. RESULTS AND MAJOR CONCLUSION: DMEM culture supernatant of L. Gasseri and L. fermentum (PH 7.3) did not show toxic effect but inhibited 50% of HIV replication at 12 and 31% concentrations, respectively. Co-culture of L. gasseri (1000 CFU/ target cell) showed mild cytotoxicity but inhibited 68% of HIV replication. The supernatant of L. crispatus inhibited 50% of HSV replication at 4% and also co-culture of L. gasseri, L. rhamnosus and L. crispatus revokes almost all of the HSV multiplication. Culture supernatants of L. gasseri and L. crispatus had significant virucidal effect against the HIV and HSV and inhibited HSV infection in a stage before viral entry to the target cells. Alive L. gasseri cells showed high potential for inhibiting HSV-1 infection in vivo condition. Current data indicates that lactobacilli supernatant encompasses components with neutralizing activity against HIV and HSV and it would be a determinant factor for viral diseases transmission and promising lead for anti-viral probiotic design.

Erectile dysfunction severity as a risk predictor for coronary artery disease.

INTRODUCTION: Erectile dysfunction (ED) is now beginning to be considered as an early manifestation of a subclinical systemic vascular disorder and may be an index of subclinical coronary artery disease (CAD). AIM: To further evaluate whether ED is a predicting factor for CAD while adjusting for other common risk factors. METHODS: One hundred eighty-three patients with newly diagnosed and documented CAD and 134 participants without CAD were enrolled in this case-control study at our referral center. Univariate and multivariate logistic regression analysis were performed to assess the effect of classic risk factors and ED severity on CAD; calculating odds ratio (OR) and 95% confidence interval (CI). Adjustments were made for potential confounding factors including age, hypertension, diabetes, dyslipidemia, obesity, and smoking. MAIN OUTCOME MEASURES: The prevalence of ED and the distribution of CAD risk factors (age, smoking, lipid profile, hypertension, obesity, and diabetes mellitus) were evaluated. The 5-item International Index of Erectile Function was used to evaluate the presence and the severity of ED. RESULTS: The prevalence of ED in CAD-positive and CAD-negative groups was 88.5% and 64.2%, respectively (P < 0.05). A statistically significant difference was found for all risk factors (except total cholesterol and low-density lipoprotein levels), and also ED prevalence between studied groups. Adjusted OR for age, diabetes, hypertension, hypercholesterolemia, and smoking demonstrated a significant confounding effect. Our results also revealed a significant association between severe ED and CAD (OR: 2.22, 95% CI: 1.11-6.03; P < 0.05). CONCLUSION: This study suggests that ED could be considered as a surrogate marker which can predict the occurrence of CAD, and severe ED could be regarded as an independent risk predictor in addition to the established ones.

Suprapubic prostatectomy with a novel catheter.

PURPOSE: We evaluated the postoperative morbidity and technical complications of a new handmade catheter used for suprapubic prostatectomy. MATERIALS AND METHODS: A total of 146 patients with lower urinary tract symptoms who had indications for open prostatectomy were enrolled in the study from January 2003 to August 2004. Of the patients 96 were treated with a novel technique without a urethral catheter but with a special handmade cystostomy catheter, while the other 50 underwent the traditional procedure using cystostomy and a urethral catheter. The same surgical team operated on the 2 groups. Clot retention episodes, hemoglobin decreases, irritative symptoms, voiding status after cystostomy removal and incontinence were evaluated postoperatively. Three months later we followed the patients with symptom score, maximum flow rate and cystoscopic examination. RESULTS: Preoperatively mean International Prostate Symptom Score was 31.6 in each group. Postoperatively none of the patients with the novel catheter complained of significant irritative urinary symptoms, clot retention and true or stress urinary incontinence. Of the cohorts 94% were satisfied with voiding but 86% of controls were satisfied. There was no report of urinary tract infection or epididymo-orchitis in the cohorts, while we found epididymo-orchitis in 4 controls (8%). Three months after the operation the mean International Prostate Symptom Score was 4.4 (range 1 to 7) and the mean maximum flow rate was 22.6 ml per second (range 14 to 25) in patients with the novel catheter, and 4.2 (range 1 to 7) and 22.5 ml per second (range 15 to 25), respectively, in those with the traditional catheter. At followup there was no bladder neck contracture but 4 patients (4.1%) showed some degree of membranous urethral stricture. We also noted 7 controls (14%) with urethral stricture. CONCLUSIONS: Transurethral prostate resection has been introduced as the surgical treatment of choice in patients with benign prostatic hyperplasia. However, open adenomectomy still has a place. Urethral catheter-free suprapubic prostatectomy can be safely applied with a low postoperative risk of infection, incontinence and stricture formation.

Posttransplant infectious complications: a prospective study on 142 kidney allograft recipients.

INTRODUCTION: We evaluated the posttransplant complications resulting from infections and their association with graft function, immunosuppressive drugs, and mortality. MATERIALS AND METHODS: A total of 142 kidney allograft recipients were followed for 1 year after transplantation. The patients' status was assessed during regular visits, and data including clinical characteristics, infections, serum creatinine level, acute rejection episodes, immunosuppressive regimen, graft function, and mortality were recorded and analyzed. RESULTS: Infections occurred in 77 patients (54%). The lower urinary (42%) and respiratory (6.3%) tracts were the most common sites of infection. The most frequent causative organisms were Klebsiella in 34 (24%) and cytomegalovirus in 25 patients (18%). Wound infection occurred in 7 patients (5%). The mortality rate was 7.7% and infection-related death was seen in 5 patients (3.5%) who developed sepsis. Graft loss was seen in 16 patients (11%), of whom 2 developed cytomegalovirus infection, 2 experienced urinary tract infection, and 5 developed sepsis and died. Mycobacterial and hepatitis C infections were noticeably rare (0.7% and 2.8%, respectively). CONCLUSION: This study showed that infections are important causes of morbidity and mortality during the posttransplant period. We recommend that serologic tests be performed before and after transplantation to recognize and meticulously follow those who are at risk. In our study, high-risk patients were those with elevated serum creatinine levels who received high doses of immunosuppressive drugs. As the urinary tract is the most common site of infection, early removal of urethral catheter is recommended to reduce the risk of infection.

Preoperative corporal biopsy as a predictor of postoperative results in venoocclusive erectile dysfunction.

INTRODUCTION: Our aim was to investigate the association of corporeal cavernosal pathology with venoocclusive erectile dysfunction (ED) and whether preoperative corporeal biopsy can help predict postoperative results. MATERIALS AND METHODS: A total of 36 patients with venoocclusive ED underwent corporeal cavernosal biopsy and venous ligation. Preoperative assessment included complete physical examination, international index of erectile dysfunction (IIEF) scoring, nocturnal penile tumescence, penile Doppler ultrasonography, cavernosography, and, if needed, cavernosometry. Three months after surgery, all patient parameters were reevaluated and compared with the preoperative results. Biopsy results of 43 patients with penile fracture were used for controls. RESULTS: The mean follow-up was 49.0 +/- 24.1 months. The mean age of the patients with ED was 32.1 +/- 8.6 years. Venous leakage was clearly revealed by cavernosography in all patients, preoperatively. The IIEF score and peak systolic velocity of the cavernosal artery in the patients did not differ postoperatively. The mean end diastolic velocity (EDV), however, decreased from 11.0 cm/s to 5.1 cm/s (P = .023). Only 2 patients had satisfactory penile rigidity after venous ligation, 2 were helped by intracorporeal injection to achieve full rigidity, and 1 used oral sildenafil to achieve full rigidity. Pathologically, 23 patients had a slight decrease of cavernosal smooth muscle cells, while in 9 patients, the cavernosal smooth muscles were markedly decreased and replaced by collagen fibers. Four patients had normal histologies, and all responded either partially or completely to surgical therapy. In the control group, 41 of 43 patients had a normal histologies, and 2 had a slight decrease of smooth muscle cell mass. CONCLUSION: Decreased cavernosal smooth muscle mass may impair erectile function. Its association with venoocclusive ED may be a poor prognostic factor of the outcome of surgical therapy. For the preoperative evaluation of patients, we propose cavernosal biopsy.