Nitrite reduction to nitric oxide by deoxyhemoglobin vasodilates the human circulation.

Nitrite anions comprise the largest vascular storage pool of nitric oxide (NO), provided that physiological mechanisms exist to reduce nitrite to NO. We evaluated the vasodilator properties and mechanisms for bioactivation of nitrite in the human forearm. Nitrite infusions of 36 and 0.36 micromol/min into the forearm brachial artery resulted in supra- and near-physiologic intravascular nitrite concentrations, respectively, and increased forearm blood flow before and during exercise, with or without NO synthase inhibition. Nitrite infusions were associated with rapid formation of erythrocyte iron-nitrosylated hemoglobin and, to a lesser extent, S-nitroso-hemoglobin. NO-modified hemoglobin formation was inversely proportional to oxyhemoglobin saturation. Vasodilation of rat aortic rings and formation of both NO gas and NO-modified hemoglobin resulted from the nitrite reductase activity of deoxyhemoglobin and deoxygenated erythrocytes. This finding links tissue hypoxia, hemoglobin allostery and nitrite bioactivation. These results suggest that nitrite represents a major bioavailable pool of NO, and describe a new physiological function for hemoglobin as a nitrite reductase, potentially contributing to hypoxic vasodilation.

Enhancement of vaccine-mediated antitumor immunity in cancer patients after depletion of regulatory T cells.

In this study, we investigated whether elimination of CD4+/CD25+ Tregs using the recombinant IL-2 diphtheria toxin conjugate DAB(389)IL-2 (also known as denileukin diftitox and ONTAK) is capable of enhancing the immunostimulatory efficacy of tumor RNA-transfected DC vaccines. We show that DAB(389)IL-2 is capable of selectively eliminating CD25-expressing Tregs from the PBMCs of cancer patients without inducing toxicity on other cellular subsets with intermediate or low expression of CD25. DAB(389)IL-2-mediated Treg depletion resulted in enhanced stimulation of proliferative and cytotoxic T cell responses in vitro but only when DAB(389)IL-2 was omitted during T cell priming. DAB(389)IL-2 significantly reduced the number of Tregs present in the peripheral blood of metastatic renal cell carcinoma (RCC) patients and abrogated Treg-mediated immunosuppressive activity in vivo. Moreover, DAB(389)IL-2-mediated elimination of Tregs followed by vaccination with RNA-transfected DCs significantly improved the stimulation of tumor-specific T cell responses in RCC patients when compared with vaccination alone. Our findings may have implications in the design of immune-based strategies that may incorporate the Treg depletion strategy to achieve potent antitumor immunity with therapeutic impact.

Sympathomimetic Effects of Acute E-Cigarette Use: Role of Nicotine and Non-Nicotine Constituents.

BACKGROUND: Chronic electronic (e) cigarette users have increased resting cardiac sympathetic nerve activity and increased susceptibility to oxidative stress. The purpose of the present study is to determine the role of nicotine versus non-nicotine constituents in e-cigarette emissions in causing these pathologies in otherwise healthy humans. METHODS AND RESULTS: Thirty-three healthy volunteers who were not current e-cigarette or tobacco cigarette smokers were studied. On different days, each participant used an e-cigarette with nicotine, an e-cigarette without nicotine, or a sham control. Cardiac sympathetic nerve activity was determined by heart rate variability, and susceptibility to oxidative stress was determined by plasma paraoxonase activity. Following exposure to the e-cigarette with nicotine, but not to the e-cigarette without nicotine or the sham control, there was a significant and marked shift in cardiac sympathovagal balance towards sympathetic predominance. The decrease in high-frequency component and the increases in the low-frequency component and the low-frequency to high-frequency ratio were significantly greater following exposure to the e-cigarette with nicotine compared with exposure to the e-cigarette without nicotine or to sham control. Oxidative stress, as estimated by plasma paraoxonase, did not increase following any of the 3 exposures. CONCLUSIONS: The acute sympathomimetic effect of e-cigarettes is attributable to the inhaled nicotine, not to non-nicotine constituents in e-cigarette aerosol, recapitulating the same heart rate variability pattern associated with increased cardiac risk in multiple populations with and without known cardiac disease. Evidence of oxidative stress, as estimated by plasma paraoxonase activity, was not uncovered following acute e-cigarette exposure.

Radical perineal prostatectomy for treatment of localized prostate cancer in obese and nonobese patients: a matched control study.

OBJECTIVES: To compare the perioperative outcomes of severely obese and nonobese patients undergoing radical perineal prostatectomy (RPP). METHODS: A cohort of 71 severely obese patients, as defined by a body mass index of 35 kg/m2 or more, who underwent RPP between 1992 and 2003 was retrospectively identified. These patients were matched by age, American Society of Anesthesiologists class, and year of surgery to a cohort of 71 nonobese patients (body mass index less than 25 kg/m2). Statistical testing was performed to compare the estimated blood loss, transfusion requirements, and complication rates (primary endpoints), as well as the length of surgery, intraoperative anesthesia requirements, postoperative hematocrit level, length of stay, and surgical margin status (secondary endpoints). RESULTS: The mean body mass index +/- standard deviation of patients in the obese and nonobese group was 38.9 +/- 4.7 and 22.9 +/- 1.6 kg/m2 (P = 0.001), respectively. Patients were similar with regard to baseline characteristics. Obese and nonobese patients did not demonstrate significant differences in mean estimated blood loss (571 +/- 391 and 494 +/- 317 mL, respectively; P = 0.06), transfusion rates (2.8% and 7.0%, respectively; P = 0.45), or positive surgical margin rates (14.1% and 9.9%, respectively; P = 0.22). The overall complication rates were significantly different at 16.9% and 7.0% (P = 0.03). CONCLUSIONS: Severely obese patients undergoing RPP had blood transfusion rates similar to those of the nonobese patients. Obese RPP patients were at increased risk of surgical and anesthesia-related perioperative complications, many of which might be avoidable. Specifically, efforts should be directed toward preventing the development of lower extremity neurapraxia by minimizing the operative time and optimizing patient positioning.

Radical perineal prostatectomy for the treatment of localized prostate cancer in morbidly obese patients.

PURPOSE: We assessed the feasibility of radical perineal prostatectomy (RPP) in morbidly obese patients with clinically organ confined prostate cancer. MATERIALS AND METHODS: Of 1,265 consecutive patients who underwent RPP at our institution from 1992 to 2003 we identified 18 with a body mass index (BMI) of 40 kg/m or greater. Demographic and clinical patient characteristics were obtained from the medical records, which were further reviewed to identify the perioperative incidence of surgical and anesthesia related complications. RESULTS: Median BMI was 41.7 kg/m (range 40.2 to 62.6). Five patients had a BMI of 45.0 kg/m or greater. No intraoperative or anesthesia related complication occurred. Mean operative time +/- SD was 188 +/- 32 minutes and estimated blood loss was 573 +/- 285 ml. None of the 18 patients received blood transfusions. During the immediate postoperative period 4 complications occurred in the form of lower extremity neuropraxia in 2 patients, local skin bleeding in 1 and early sepsis in 1 requiring rehospitalization for intravenous antibiotics. Mean operative time and estimated blood loss were significantly lower when surgery was performed by a highly experienced surgeon compared with experienced surgeons (174 +/- 21 vs 235 +/- 10 minutes and 485 +/- 258 vs 838 +/- 197 ml, p = 0.001 and 0.027, respectively). CONCLUSIONS: RPP in morbidly obese patients is feasible and it is associated with acceptable perioperative morbidity. The perineal approach should be considered in morbidly obese patients seeking surgical treatment for clinically localized prostate cancer.

Cross-sectional survey of long-term quality of life after radical perineal prostatectomy.

OBJECTIVES: To evaluate the late health-related quality of life (HRQOL) after radical perineal prostatectomy (RPP) and identify the predictors of outcome. METHODS: We performed a cross-sectional study of 266 consecutive patients who underwent RPP for clinically localized prostate cancer between July 1998 and December 2000. Of the 236 patients successfully contacted, 187 (79.2%) returned a validated patient self-assessment questionnaire, the Expanded Prostate Cancer Index Composite, a mean of 42.1 months (range 29 to 64) months after surgery. The median HRQOL scores were calculated in four disease-specific domains: urinary, bowel, sexual, and hormonal. Preoperative baseline information from a separate group of 144 consecutive RPP candidates from January 2002 to May 2003 was used for comparison. Univariate and multivariate logistic regression analyses were used to identify predictors of more favorable long-term HRQOL outcomes. RESULTS: No statistically significant differences were found in any of the domain-specific summary scores between the study and reference groups, except in the sexual domain (median score 19.2 versus 56.4; P = 0.001). The number of medical comorbidities was a statistically significant predictor of HRQOL summary scores in all domains (P <0.05). In addition, the urinary summary score was statistically significantly associated with income (P = 0.03), sexual summary with the use of erectile aids (P = 0.003), bowel summary with secondary radiotherapy (P = 0.001) and income (P = 0.002), and hormonal summary with androgen ablation (P = 0.004). CONCLUSIONS: The results of this study have shown that the long-term HRQOL of RPP patients in the urinary, bowel, and hormonal domains is favorable. HRQOL outcomes depend on a spectrum of factors, including the presence of comorbid disease, socioeconomic status, and secondary cancer treatments. Future studies should seek to address the efficacy of preserving the sexual domain HRQOL in patients undergoing bilateral nerve-sparing RPP.

Telomerase mRNA-transfected dendritic cells stimulate antigen-specific CD8+ and CD4+ T cell responses in patients with metastatic prostate cancer.

Telomerase reverse transcriptase (hTERT) represents an attractive target for cancer immunotherapy because hTERT is reactivated in most human tumors. A clinical trial was initiated in which hTERT mRNA-transfected dendritic cells (DC) were administered to 20 patients with metastatic prostate cancer. Nine of these subjects received DC transfected with mRNA encoding a chimeric lysosome-associated membrane protein-1 (LAMP) hTERT protein, allowing for concomitant induction of hTERT-specific CD8+ and CD4+ T cell responses. Treatment was well tolerated. Intense infiltrates of hTERT-specific T cells were noted at intradermal injection sites after repeated vaccination. In 19 of 20 subjects, expansion of hTERT-specific CD8+ T cells was measured in the peripheral blood of study subjects, with 0.9-1.8% of CD8+ T cells exhibiting Ag specificity. Patients immunized with the chimeric LAMP hTERT vaccine developed significantly higher frequencies of hTERT-specific CD4+ T cells than subjects receiving DC transfected with the unmodified hTERT template. Moreover, CTL-mediated killing of hTERT targets was enhanced in the LAMP hTERT group, suggesting that an improved CD4+ response could augment a CTL response. Vaccination was further associated with a reduction of prostate-specific Ag velocity and molecular clearance of circulating micrometastases. Our findings provide a rationale for further development of hTERT-transfected DC vaccines in the treatment of prostate and other cancers.

Prospective and longitudinal patient self-assessment of health-related quality of life following radical perineal prostatectomy.

PURPOSE: We provide a comprehensive, longitudinal assessment of health related quality of life (HRQOL) following radical perineal prostatectomy (RPP). MATERIALS AND METHODS: We report the results of a prospective cohort study of 109 patients with at least 3 months of followup who underwent RPP between January 2001 and July 2003. A validated patient self-assessment questionnaire, the Expanded Prostate Cancer Index Composite, was administered preoperatively, and 1, 3, 6, 9, 12 and 18 months postoperatively. Mean domain specific HRQOL scores were calculated as well as the proportion of patients achieving an individual baseline by each interval. The Cox proportional hazards model was used to identify predictors of a successful return to baseline of disease specific HRQOL scores. RESULTS: HRQOL scores were lowest 1 month postoperatively and they increased with time. By 6 months a majority of patients had recovered baseline summary scores in urinary (65.1%), bowel (93.6%) and hormonal (91.7%) domains at a median of 5.8 (95% CI 3.6 to 6.2), 1.3 (95% CI 1.1 to 1.5) and 1.3 (95% CI 1.2 to 1.8) months, respectively. One in 4 patients recovered the sexual summary score by 18 months. Significant independent predictors for the recovery of domain summary scores were younger age in urinary (p = 0.001), individual surgeon in bowel (p = 0.022), and older age (p = 0.017) and absent medical comorbidities (p = 0.012) in hormonal domains. CONCLUSIONS: A majority of patients undergoing RRP experience an early recovery of individual urinary, bowel and hormonal HRQOL. Future studies should establish the benefit of bilateral nerve sparing RPP on the recovery of sexual domain HRQOL.

S-Nitrosohemoglobin is unstable in the reductive erythrocyte environment and lacks O2/NO-linked allosteric function.

Our previous results run counter to the hypothesis that S-nitrosohemoglobin (SNO-Hb) serves as an in vivo reservoir for NO from which NO release is allosterically linked to oxygen release. We show here that SNO-Hb undergoes reductive decomposition in erythrocytes, whereas it is stable in purified solutions and in erythrocyte lysates treated with an oxidant such as ferricyanide. Using an extensively validated methodology that eliminates background nitrite and stabilizes erythrocyte S-nitrosothiols, we find the levels of SNO-Hb in the basal human circulation, including red cell membrane fractions, were 46 +/- 17 nm in human arterial erythrocytes and 69 +/- 11 nm in venous erythrocytes, incompatible with the postulated reservoir function of SNO-Hb. Moreover, we performed experiments on human red blood cells in which we elevated the levels of SNO-Hb to 10,000 times the normal in vivo levels. The elevated levels of intra-erythrocytic SNO-Hb fell rapidly, independent of oxygen tension and hemoglobin saturation. Most of the NO released during this process was oxidized to nitrate. A fraction (25%) was exported as S-nitrosothiol, but this fraction was not increased at low oxygen tensions that favor the deoxy (T-state) conformation of Hb. Results of these studies show that, within the redox-active erythrocyte environment, the beta-globin cysteine 93 is maintained in a reduced state, necessary for normal oxygen affinity, and incapable of oxygen-linked NO storage and delivery.