Application of enhanced stromal vascular fraction and fat grafting mixed with PRP in post-traumatic lower extremity ulcers

BackgroundThe authors presented their experience in regenerative surgery of post-traumatic lower extremity ulcers, evaluating the effects related to the use of Enhanced Stromal Vascular Fraction (e-SVF) and Fat Grafting with Platelet rich Plasma (PRP). The authors compared the results of two control groups.MethodThe analysis involved 20 patients aged between 23 to 62 years affected by post-traumatic lower extremity ulcers. 10 patients managed with e-SVF and 10 patients managed with Fat grafting + PRP in the Plastic and Reconstructive Surgery Department at “Tor Vergata” University Rome. Patients in the first control group (n = 10), were treated only with curettage and application of hyaluronic acid in the bed of ulcers. Patients in the second control group (n = 10), were treated only with PRP.ResultsThe authors showed that wounds treated with e-SVF healed better than those treated with hyaluronic acid. In fact, after 9.7 weeks, patients treated with e-SVF underwent 97.9% ± 1.5% reepithelialisation compared to 87.8% ± 4.4% of the first control group (only hyaluronic acid; p < 0.05). Patients treated with PRP and fat grafting also showed an improvement in reepithelialisation; in fact after 9.7 weeks, they underwent a 97.8% ± 1.5% reepithelialisation compared to 89.1% ± 3.8% of the second control group (only PRP; p < 0.05). As reported e-SVF and PRP mixed with fat grafting were the two treatments evidencing improvement in the healing of patients post-traumatic extremity ulcers.ConclusionsThe results obtained proved the efficacy of these treatments, and the satisfaction of the patients confirmed the quality of the results.     

Structural alterations in rat myocardium induced by chronic l-arginine and l-NAME supplementation

Structural changes affecting cardiomyocyte function may contribute to the pathophysiological remodeling underlying cardiac function impairment. Recent reports have shown that endogenous nitric oxide (NO) plays an important role in this process. In order to examine the role of NO in cardiomyocyte remodeling, male rats were acclimated to room temperature (22 ± 1 °C) or cold (4 ± 1 °C) and treated with 2.25% l-arginine·HCl or 0.01% l-NAME (N^ω-nitro-l-arginine methyl ester)·HCl for 45 days. Untreated groups served as controls. Right heart ventricles were routinely prepared for light microscopic examination. Stereological estimations of volume densities of cardiomyocytes, surrounding blood vessels and connective tissue, as well as the morphometric measurements of cardiomyocyte diameters were performed. Tissue sections were also analyzed for structural alterations. We observed that both l-arginine and l-NAME supplementation induced cardiomyocyte hypertrophy, regardless of ambient temperature. However, cardiomyocyte hypertrophy was associated with fibrosis and extra collagen deposition only in the l-NAME treated group. Taken together, our results suggest that NO has a modulatory role in right heart ventricle remodeling by coordinating hypertrophy of cardiomyocytes and fibrous tissue preventing cardiac fibrosis.     

Meal timing and composition influence ghrelin levels, appetite scores and weight loss maintenance in overweight and obese adults

BackgroundAlthough dietary restriction often results in initial weight loss, the majority of obese dieters fail to maintain their reduced weight. Diet-induced weight loss results in compensatory increase of hunger, craving and decreased ghrelin suppression that encourage weight regain. A high protein and carbohydrate breakfast may overcome these compensatory changes and prevent obesity relapse.MethodsIn this study 193 obese (BMI 32.2 ± 1.0 kg/m²), sedentary non diabetic adult men and women (47 ± 7 years) were randomized to a low carbohydrate breakfast (LCb) or an isocaloric diet with high carbohydrate and protein breakfast (HCPb). Anthropometric measures were assessed every 4 weeks. Fasting glucose, insulin, ghrelin, lipids, craving scores and breakfast meal challenge assessing hunger, satiety, insulin and ghrelin responses, were performed at baseline, after a Diet Intervention Period (Week 16) and after a Follow-up Period (Week 32).ResultsAt Week 16, groups exhibited similar weight loss: 15.1 ± 1.9 kg in LCb group vs. 13.5 ± 2.3 kg in HCPb group, p = 0.11. From Week 16 to Week 32, LCb group regained 11.6 ± 2.6 kg, while the HCPb group lost additional 6.9 ± 1.7 kg. Ghrelin levels were reduced after breakfast by 45.2% and 29.5% following the HCPb and LCb, respectively. Satiety was significantly improved and hunger and craving scores significantly reduced in the HCPb group vs. the LCb group.ConclusionA high carbohydrate and protein breakfast may prevent weight regain by reducing diet-induced compensatory changes in hunger, cravings and ghrelin suppression. To achieve long-term weight loss, meal timing and macronutrient composition must counteract these compensatory mechanisms which encourage weight regain after weight loss.     

A targeted high-efficiency angiogenesis strategy as therapy for myocardial infarction

AimsThe aim of this study was to prove that an intramyocardial injection of a mixture of low-dose human growth factor (HGF) plasmid and microbubbles (MB) in combination with insonation was an effective therapy for myocardial infarction.Main methodsTwenty dogs with myocardial infarction were divided into 4 groups: (1) HGF, MB and ultrasound (HGF-US/MB), (2) HGF and US (HGF-US), (3) HGF alone and (4) surgery alone (control). In the HGF-US/MB group, HGF plasmid DNA (500 μg) mixed with 0.5 ml of MB solution was injected 5 min after coronary occlusion followed by insonation. With the exception of the control group, the other dogs were divided into two groups, one treated with the HGF gene and insonation and the other with the HGF gene only.Key findingsCompared to the HGF group, infarct size decreased from 32% ± 7% (control) to 23% ± 5% in the HGF-US/MB group 28 d later (P < 0.05). Capillary density increased from 21.7 ± 4.2/mm² (control) to 114.3 ± 28.9/mm² in the HGF-US/MB group (P < 0.01). Compared to the HGF group, there was a 14% decrease in the ratio of left ventricle weight/body weight and a 25% decrease in hydroxyproline content. We also observed a 29% and 20% decrease in collagen volume fraction of type I and type III collagen, respectively in the HGF-US/MB group.SignificanceIntramyocardial injection of HGF and MB in combination with insonation enhances neovascularization and reduces ventricular remodeling and infarct size.     

Development and validation of a procedure to isolate viable bone marrow cells from the vertebrae of cadaveric organ donors for composite organ grafting

Background aimsDonor-derived vertebral bone marrow (BM) has been proposed to promote chimerism in solid organ transplantation with cadaveric organs. Reports of successful weaning from immunosuppression in patients receiving directed donor transplants in combination with donor BM or blood cells and novel peri-transplant immunosuppression has renewed interest in implementing similar protocols with cadaveric organs.MethodsWe performed six pre-clinical full-scale separations to adapt vertebral BM preparations to a good manufacturing practice (GMP) environment. Vertebral bodies L4–T8 were transported to a class 10 000 clean room, cleaned of soft tissue, divided and crushed in a prototype bone grinder. Bone fragments were irrigated with medium containing saline, albumin, DNAse and gentamicin, and strained through stainless steel sieves. Additional cells were eluted after two rounds of agitation using a prototype BM tumbler.ResultsThe majority of recovered cells (70.9 ± 14.1%, mean ± SD) were eluted directly from the crushed bone, whereas 22.3% and 5.9% were eluted after the first and second rounds of tumbling, respectively. Cells were pooled and filtered (500, 200 μm) using a BM collection kit. Larger lumbar vertebrae yielded about 1.6 times the cells of thoracic vertebrae. The average product yielded 5.2 ± 1.2 × 10¹⁰ total cells, 6.2 ± 2.2 × 10⁸ of which were CD45⁺ CD34⁺. Viability was 96.6 ± 1.9% and 99.1 ± 0.8%, respectively. Multicolor flow cytometry revealed distinct populations of CD34⁺ CD90⁺ CD117^dim hematopoietic stem cells (15.5 ± 7.5% of the CD34 ⁺ cells) and CD45^? CD73⁺ CD105⁺ mesenchymal stromal cells (0.04 ± 0.04% of the total cells).ConclusionsThis procedure can be used to prepare clinical-grade cells suitable for use in human allotransplantation in a GMP environment.     

Is preconditioning by oxytocin administration mediated by iNOS and/or mitochondrial KATP channel activation in the in vivo anesthetized rabbit heart?

AimsOxytocin (OXT) pretreatment protects the heart during ischemia–reperfusion injury by activating ATP-dependent potassium (K_ATP) channels. The aim of the current study was to elucidate the roles of nitric oxide synthaseNOS and myocardial biochemistry in the cardioprotective effects of OXT and ischemic preconditioning (IPC).Main methodsMale New Zealand White anesthetized rabbits (13 groups) were subjected to 30 min of occlusion of the left coronary artery and 120 min of reperfusion with or without IPC.Key findingsIPC (1 cycle), OXT (0.03 μg/kg, i.p.) or IPC + OXT yield significant infarct size reductions (21.8 ± 1.5%, 20.5 ± 1.2% and 19.4 ± 1.4%, respectively, versus 38.9 ± 3.5% in the S-CONT group; P < 0.01) and antiarrhythmic effects, including VF (0%, 0% and 0%, versus 50% in S-CONT group; P < 0.05) sustained VT (13%, 13% and 13%, versus 100% in S-CONT group; P < 0.005) and other arrhythmias (25%, 13% and 25%, versus 100% in S-CONT group; P < 0.005, P < 0.01 and P < 0.005, respectively). Atosiban (ATO, a selective OXT receptor antagonist), 5-HD and l-NAME (a nonspecific NOS inhibitor) abolished the beneficial effects of IPC and OXT, suggesting that the benefits are achieved via selective activation of OXT receptors, mitochondrial K_ATP channels and NO. An iNOS inhibitor (1400 W) blocked the beneficial effects of IPC but not OXT. The IPC, OXT, IPC + OXT and 1400 W + OXT interventions significantly preserved ATP levels in the heart.SignificanceThis study demonstrates similarities between acute OXT pretreatment and IPC in terms of infarct size reduction, antiarrhythmic activity, and metabolic status.     

Plasma renin and aldosterone levels in obese and non-obese women with polycystic ovary syndrome

ObjetiveTo assess plasma renin and aldosterone levels in obese and non-obese women with polycystic ovary syndrome (PCOS).MethodsObese women (body mass index [BMI] > 30 kg/m2; group A, n = 34) and non-obese women (BMI < 25 kg/m2; group B, n = 13) with PCOS were selected. The control group (group C, n =47) consisted of age-matched women with regular menses and normal ultrasonographic ovaries. Luteinizing hormone, follicle-stimulating hormone, androstenedione, testosterone, sex hormone-binding globulin, serum glucose, insulin, renin, plasma renin activity, and aldosterone levels were measured.ResultsObese and non-obese women with PCOS had higher luteinizing hormone, follicle-stimulating hormone, androstenedione, testosterone, and insulin levels as compared to women in the control group (p < 0.05). Women with PCOS had significantly higher renin levels (group A: 50.2 ± 4.9 picoU/mL, group B: 39.9 ± 2.7 picoU/mL, and group C: 24.6 ± 2.6 picoU/mL), plasma renin activity (group A: 3.7 ± 0.3 ng/mL/h, group B: 3.6 ± 0.3 ng/mL/h, and group C: 2.2 ± 0.4 ng/mL/h), and aldosterone levels (group A: 31.2 ± 3.3 ng/dL, group B: 29.3 ± 2.9 ng/dL, and group C: 22.2 ± 3.9 ng/dL) as compared with controls.ConclusionSignificant differences exist in plasma renin and aldosterone levels between obese and non-obese women as compared with polycystic ovary syndrome and normal controls.     

Involvement of NO-guanylyl cyclase pathway for the depression of spinal monosynaptic reflex by Mesobuthus tamulus venom in neonatal rat in vitro

AimsThe present study was undertaken to evaluate the role of nitric oxide (NO) in Mesobuthus tamulus (MBT) venom-induced depression of spinal reflexes.Main methodsExperiments were performed on isolated hemisected spinal cords from 4 to 6 day old rats. Stimulation of a dorsal root with supramaximal strength evoked monosynaptic (MSR) and polysynaptic reflex (PSR) potentials in the corresponding segmental ventral root.Key findingsSuperfusion of MBT venom (0.3 μg/ml) depressed the spinal reflexes in a time-dependent manner and the maximum depression was seen at 10 min (MSR by 63%; PSR by 79%). The time to produce 50% depression (T-50) of MSR and PSR was 7.7 ± 1.3 and 5.7 ± 0.5 min, respectively. Pretreatment with bicuculline (1 μM; GABA_A receptor antagonist) or strychnine (1 μM; glycine_A receptor antagonist) did not block the venom-induced depression of spinal reflexes. However, Nω-nitro-L-arginine methyl ester (L-NAME, 100 or 300 μM; NO synthase inhibitor) or hemoglobin (Hb, 100 μM; NO scavenger) antagonized the venom-induced depression of MSR. Further, soluble guanylyl cylase inhibitors (1 H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one, ODQ; 1 μM or methylene blue, 100 μM) also antagonized the venom-induced depression of MSR but not PSR. Nitrite concentration (indicator of NO activity) of the cords exposed to venom (0.3 μg/ml) was not different from the control group.SignificanceThe results indicate that venom-induced depression of MSR is mediated via NO-guanylyl cyclase pathway without involving GABAergic or glycinergic system.     

Production of interspecies handmade cloned embryos by nuclear transfer of cattle, goat and rat fibroblasts to buffalo (Bubalus bubalis) oocytes

The possibility of producing interspecies handmade cloned (iHMC) embryos by nuclear transfer from donor cells of cattle, goat and rat using buffalo oocytes as recipient cytoplasts was explored. Zona-free buffalo oocytes were enucleated by protrusion cone-guided bisection with a microblade. After electrofusion with somatic cells, reconstructed oocytes were activated by calcimycin A23187, treated with 6-dimethylaminopurine and were cultured in K-RVCL-50^® medium for 8 days. Although the cleavage rate was not significantly different when buffalo, cattle, goat or rat cells were used as donor nuclei (74.6 ± 3.8, 82.8 ± 5.3, 86.0 ± 4.9 and 82.3 ± 3.6%, respectively), the blastocyst rate was significantly higher (P < 0.01) for buffalo (51.4 ± 2.6) than for cattle (3.5 ± 1.0) or the goat (2.2 ± 0.9), whereas none of the embryos crossed the 32-cell stage when rat cells were used. However, the total cell number was similar for buffalo–buffalo (175.0 ± 5.07) and cattle–buffalo embryos (178.0 ± 11.84). Following transfer of 3 buffalo–buffalo embryos each to 6 recipients, 3 were found to be pregnant, though the pregnancies were not carried to full term. These results suggest that interspecies blastocyst stage embryos can be produced by iHMC using buffalo cytoplasts and differentiated somatic cells from cattle and goat and that the source of donor nucleus affects the developmental competence of interspecies embryos.     

Involvement of src-kinase activation in ischemic preconditioning induced protection of mouse brain

AimsTo investigate the role of src-kinase in ischemic preconditioning induced reversal of ischemia and reperfusion induced cerebral injury in mice.Main methodsBilateral carotid artery occlusion of 17 min followed by reperfusion for 24 h was employed to produce ischemia and reperfusion induced cerebral injury in mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining using both by volume and by weight methods differently. Memory was evaluated using elevated plus maze test. Rota rod test was employed to assess motor incoordination.Key findingsBilateral carotid artery occlusion followed by reperfusion produced cerebral infarction and impaired memory and motor co-ordination. Three preceding episodes of bilateral carotid artery occlusion for 1 min and reperfusion of 1 min (ischemic preconditioning) prevented markedly ischemia–reperfusion-induced cerebral injury measured in terms of infarct size (38.5 ± 1.3% and 38.5 ± 2.9% mean infarct of control animals was reduced to 24.3 ± 1.2% and 23.5 ± 1.8% of the preconditioning groups respectively), loss of memory (72.2 ± 3.6 mean transfer latency time of control animals was reduced to 25.6 ± 5.2 of the preconditioning group respectively) and motor coordination (78.3 ± 17.6 s mean falling down latency time of control animals was increased to a mean value of 180.9 ± 6.5 s of the preconditioning groups respectively). SU6656 (2 mg/kg, ip) and PP1 (0.1 mg/kg, ip), highly selective src-kinase inhibitors, attenuated this neuroprotective effect of ischemic preconditioning.SignificanceTherefore, neuroprotective effect of ischemic preconditioning may be due to src-kinase linked mechanism.     

Cryopreservation of umbilical cord mesenchymal cells in xenofree conditions

Background aimsMesenchymal stromal cells (MSC) are being used to treat and prevent a variety of clinical conditions. To be readily available, MSC must be cryopreserved until infusion. However, the optimal cryopreservation methods, cryoprotector solutions and MSC sensitivity to dimethyl sulfoxide (DMSO) exposure are unknown. This study investigated these issues.MethodsMSC samples were obtained from human umbilical cord (n = 15), expanded with Minimal Essential Medium-alpha (α-MEM) 10% human serum (HS), resuspended in 25 mL solution (HS, 10% DMSO, 20% hydroxyethyl starch) and cryopreserved using the BioArchive^® system. After a mean of 18 ± 7 days, cell suspensions were thawed and diluted until a DMSO concentration of 2.5% was reached. Samples were tested for cell quantification and viability, immunophenotype and functional assays.ResultsPost-thaw cell recovery: 114 ± 2.90% (mean ± SEM). Recovery of viable cells: 93.46 ± 4.41%, 90.17 ± 4.55% and 81.03 ± 4.30% at 30 min, 120 min and 24 h post-thaw, respectively. Cell viability: 89.26 ± 1.56%, 72.71 ± 2.12%, 70.20 ± 2.39% and 63.02 ± 2.33% (P < 0.0001) pre-cryopreservation and 30 min, 120 min and 24 h post-thaw, respectively. All post-thaw samples had cells that adhered to culture bottles. Post-thaw cell expansion was 4.18 ± 0.17 ×, with a doubling time of 38 ± 1.69 h, and their capacity to inhibit peripheral blood mononuclear cells (PBMC) proliferation was similar to that observed before cryopreservation. Differentiation capacity, cell-surface marker profile and cytogenetics were not changed by the cryopreservation procedure.ConclusionsA method for cryopreservation of MSC in bags, in xenofree conditions, is described that facilitates their clinical use. The MSC functional and cytogenetic status and morphologic characteristics were not changed by cryopreservation. It was also demonstrated that MSC are relatively resistant to exposure to DMSO, but we recommend cell infusion as soon as possible.     

Right ventricular pacing and sensing function in high posterior septal and apical lead placement in cardiac resynchronization therapy

BackgroundThe conventional right ventricular (RV) lead position in cardiac resynchronization therapy pacemakers (CRT-P) is the RV apex (RV-A). Little is known about electrophysiological stability and associated complications of pacing leads in RV high posterior septal (RV-HS) position in CRT-P.MethodsTwo hundred and thirty-five consecutive CRT-P patients were included from 1999-2010. Pacing thresholds at 0.5 ms and 2.5 V, sensing electrograms and lead impedances were measured at implant and repeated 1,3,6,12,18 and 24 months after CRT-P. Electrophysiological measurements of leads located in RV-A and RV-HS were analyzed retrospectively. Bipolar RV leads were used, including high impedance leads, passive fixation and active fixation.ResultsRV pacing leads were implanted in RV-A (n = 79) and RV-HS (n = 156). Average RV pacing thresholds from CRT implant procedure to 24-month follow-up at 0.5 ms were 0.77 ± 0.69 V in RV-A and 0.71 ± 0.35 V in RV-HS (P = 0.31), and at 2.5 V were 0.06 ± 0.08 ms in RV-A and 0.07 ± 0.05 ms in RV-HS (P = 0.12). Average RV electrogram amplitudes from baseline to 24 months after CRT were 15.3 ± 6.9 mV in RV-A and 12.1 ± 6.0 mV in RV-HS (P = 0.55). Average RV impedances during follow-up were 850 ± 286Ω in RV-A and 618 ± 147Ω in RV-HS (P = 0.57). Similar RV lead revisions between RV-A and RV-HS were observed after 2-year follow-up (P = 0.55).ConclusionsThe RV-HS lead position demonstrated stable and acceptable long-term pacing and sensing function, with rates of complications comparable to conventional RV-A lead position in CRT. The RV-HS lead position is feasible in CRT-P.     

Role of proinflammatory markers and NT-proBNP in patients with an implantable cardioverter-defibrillator and an electrical storm

BackgroundSeveral studies have attempted to identify risk factors for the development of an electrical storm (ES), which is defined as ⩾3 separate ventricular tachyarrhythmic (VT/VF) events, but in the majority of studies no triggers have been found. However, little is known about the role of inflammation and NT-proBNP in patients with ES. The aim of this study was therefore to assess the relationship of Interleukin-6 (IL-6), high-sensitivity C-reactive protein (hs-CRP) and NT-proBNP serum concentrations in ICD-patients with or without single spontaneous ventricular tachyarrhythmic events (VT/VF) and in ES.MethodsMarkers were determined in 51 patients without ICD-intervention, in 15 ICD-patients with single VT/VF-episodes during 9-months follow-up and in 20 ICD-patients with ES (blood sampling performed within 60 min after fulfilling ES criteria). VT/VF-episodes were analysed by stored ICD-electrograms.ResultsAll patients had idiopathic dilated cardiomyopathy (n = 23) or coronary artery disease (n = 63). Patients with ES revealed significantly higher mean serum concentrations of all markers (IL-6 15.19 ± 10.34 pg/mL, hs-CRP 20.12 ± 14.4 mg/L, NT-proBNP 4799 ± 4596 pg/mL) compared to baseline values of patients with single VT/VF-events during follow-up (IL-6 8.37 ± 5.8 pg/mL (p = 0.03), hs-CRP 4.7 ± 5.3 mg/dL (p < 0.001), NT-proBNP 1913 ± 2665 pg/mL (p = 0.04)) and compared to baseline values of ICD-patients without device intervention (IL-6 4.62 ± 3.66 pg/mL (p < 0.001), hs-CRP 4.1 ± 3.4 mg/L (p < 0.001), NT-proBNP 1461 ± 2281 pg/mL (p < 0.001)). In 9/20 patients presenting with ES (45%) baseline values were available. All markers were significantly higher during ES compared to event-free determination (IL-6 14.54 ± 10.43 vs. 7.03 ± 2.83 pg/mL (p = 0.04), hs-CRP 19.07 ± 16.07 vs. 6.5 ± 3.9 mg/L (p = 0.02), NT-proBNP 4218 ± 2561 vs. 2099 ± 1279 pg/mL (p = 0.03)).ConclusionsElectrical storm is associated with significantly elevated IL-6, hs-CRP and NT-proBNP serum concentrations in ICD-patients with structural heart disease. Thus, ES may be triggered by proinflammatory activity. Combined intraindividual elevation of determined markers might help to identify patients at risk of impending electrical storm.     

Application of carbon fiber composite minielectrodes for measurement of kinetic constants of nitric oxide decay in solution

Carbon fiber microelectrodes and carbon fiber composite minielectrodes (CFM/CFCM) have been generally used for measurements of nitric oxide (NO) concentration in chemical and biological systems. The response time of a CFM/CFCM is usually from milliseconds to seconds depending on the electrode size, the thickness of coating layers on the electrode, and NO diffusion coefficients of the coating layers. As a result, the time course of recoded current changes (I–t curves) by the CFM/CFCM may be different from the actual time course of NO concentration changes (c–t curves) if the half-life of NO decay is close to or shorter than the response time of the electrode used. This adds complexity to the process for determining rate constants of NO decay kinetics from the recorded current curves (I–t curves). By computer simulations based on a mathematical model, an approximation method was developed for determining rate constants of NO decay from the recorded current curves. This method was first tested and valuated using a commercial CFCM in several simple reaction systems with known rate constants. The response time of the CFCM was measured as 4.7 ± 0.7 s (n = 5). The determined rate constants of NO volatilization and NO autoxidation in our measurement system at 37 °C are (1.9 ± 0.1) × 10^?3 s^?1 (n = 4) and (2.0 ± 0.3) × 10³ M^?1 s^?1 (n = 7), which are close to the reported rate constants. The method was then applied to determine the rate of NO decay in blood samples from control and smoking exposed mice. It was observed that the NO decay rate in the smoking group is >20% higher than that in control group, and the increased NO decay rate in the smoking group was reversed by 10 μM diphenyleneiodonium chloride (DPI), an inhibitor of flavin enzymes such as leukocyte NADPH oxidase.     

Lipid profile modifications in post-menopausal women treated with testosterone gel

ObjectiveTo assess lipid profile changes in post-menopausal women treated with testosterone gel.MethodsThirty-six oophorectomized women on estradiol treatment who received transdermal testosterone gel (5 mg daily) were enrolled into our study. Cholesterol, triglycerides, high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol (LDL-C), very low density-lipoprotein cholesterol (VLDL-C), and lipoprotein (a) were tested before and after 6 months of treatment.ResultsSelected participants had a mean age of 50.9 ± 4.6 years and a body mass index of 30.1 ± 3.8 kg/m². Significantly decreased cholesterol levels were found after 6 months of treatment (204.5 ± 35.1 mg/dL before treatment as compared to 183.1 ± 21.9 mg/dL after treatment; p < 0.05). A significant reduction was also seen in LDL-C levels after 6 months of treatment with testosterone gel as compared to baseline (130.9 ± 29.7 mg/dL versus 118.5 ± 21.3 mg/dL; p < 0.05). No differences were found in triglyceride, HDL-C, VLDL-C, and lipoprotein (a) levels (p = ns).ConclusionEl gel de testosterona, asociado a tratamiento estrogénico en mujeres ooforectomizadas, produce disminución de las concentraciones de colesterol y LDL-C posterior a 6 meses de tratamiento, sin afectar las concentraciones de triglicéridos, HDL-C, VLDL-C y lipoproteína (a)Testosterone gel, associated to estrogen treatment in oophorectomized women, decreased cholesterol and LDL-C levels after 6 months of treatment, without affecting serum triglyceride, HDL-C, VLDL-C, and lipoprotein (a) levels.     

Role of high-fat diet in regulation of gene expression of drug metabolizing enzymes and transporters

AimsLate phase ischemic preconditioning (LPC) protects the heart against ischemia–reperfusion (I/R) injury. However, its effect on myocardial tissue oxygenation and related mechanism(s) is unknown. The aim of the current study is to determine whether LPC attenuates post-ischemic myocardial tissue hyperoxygenation through preserving mitochondrial oxygen metabolism.Main methodsC57BL/6 mice were subjected to 30 min coronary ligation followed by 60 min or 24 h reperfusion with or without LPC (3 cycles of 5 min I/5 min R): Sham, LPC, I/R, and LPC + I/R group. Myocardial tissue Po₂ and redox status were measured with electron paramagnetic resonance (EPR) spectroscopy.Key findingsUpon reperfusion, tissue Po₂ rose significantly above the pre-ischemic level in the I/R mice (23.1 ± 2.2 vs. 12.6 ± 1.3 mm Hg, p < 0.01). This hyperoxygenation was attenuated by LPC in the LPC + I/R mice (11.9 ± 2.0 mm Hg, p < 0.01). Activities of NADH dehydrogenase (NADH-DH), succinate-cytochrome c reductase (SCR) and cytochrome c oxidase (CcO) were preserved or increased in the LPC group, significantly reduced in the I/R group, and conserved in the LPC + I/R group. Manganese superoxide dismutase (Mn-SOD) protein expression was increased by LPC in the LPC and LPC + I/R mice compared to that in the Sham control (1.24 ± 0.01 and 1.23 ± 0.01, p < 0.05). Tissue redox status was shifted to the oxidizing state with I/R (0.0268 ± 0.0016/min) and was corrected by LPC in the LPC + I/R mice (0.0379 ± 0.0023/min). Finally, LPC reduced the infarct size in the LPC + I/R mice (10.5 ± 0.4% vs. 33.3 ± 0.6%, p < 0.05).SignificanceThus, LPC preserved mitochondrial oxygen metabolism, attenuated post-ischemic myocardial tissue hyperoxygenation, and reduced I/R injury.     

Evaluation of vitamin E in the treatment of erectile dysfunction in aged rats

AimsThe present study aims to elucidate the role of oxidative stress in erectile dysfunction associated with aging and to investigate the effect of treatment with vitamin E in this respect.Main methodsRats were divided into four groups: young (3-month-old), aged rats (18-month-old), aged rats given 80 IU of vitamin E/rat/day for 21-days, aged rats given 5 mg/kg of sildenafil/day for 21-days. Intracavernosal pressure/mean arterial pressure (ICP/MAP), nitric oxide production, TBARS, GSH levels and SOD activity in corpus cavernosum were measured.Key findingsSignificant decrease in ICP/MAP was observed in aged rats at both low and high frequency of stimulation. Significant increase in ICP/MAP was observed in aged rats treated with vitamin E over the range of 0.8 to 5 Hz but young control values were not restored. Percentage potentiation of ICP/MAP than aged group at 0.8 Hz was 326 ± 41.3% and 897 ± 72.2% for vitamin E and sildenafil respectively. Decreased levels of NO₂/NO₃ and SOD activity in the penile tissue observed with aging were elevated back to control by either vitamin E or sildenafil. Penile concentration of TBARS was 20.86 ± 0.83 for aged rats vs. 11.39 ± 0.79 nmol/g tissue for young controls. Both vitamin E and sildenafil reduced penile TBARS in aged rats.SignificanceThis study proves that antioxidant therapy with vitamin E ameliorates the age-associated erectile dysfunction. Sildenafil may exert some antioxidant properties which add to the advantages of its long-term use. The effect of combinations of low-dose sildenafil and vitamin E on age-associated erectile dysfunction merits to be studied.     

Modulation of antioxidant enzyme activities and metabolites ratios by nitric oxide in short-term salt stressed soybean root nodules

Several abiotic factors cause molecular damage to plants either directly or through the accumulation of reactive oxygen species such as hydrogen peroxide (H₂O₂). We investigated if application of nitric oxide (NO) donor 2,2′-(hydroxynitrosohydrazono) bis-ethanimine (DETA/NO) could reduce the toxic effect resulting from short-term salt stress. Salt treatment (150 mM NaCl) alone and in combination with 10 μM DETA/NO or 10 μM DETA were given to matured soybean root nodules for 24 h. Salt stress resulted in high H₂O₂ level and lipid peroxidation while application of DETA/NO effectively reduced H₂O₂ level and prevented lipid peroxidation in the soybean root nodules. NO treatment increased the activities of ascorbate peroxidase and dehydroascorbate reductase under salt stress. Whereas short-term salt stress reduced AsA/DHAsA and GSH/GSSG ratios, application of the NO donor resulted in an increase of the reduced form of the antioxidant metabolites thus increasing the AsA/DHAsA and GSH/GSSG ratios. Our data suggests a protective role of NO against salt stress.     

Prevotella bivia as a source of lipopolysaccharide in the vagina

ObjectivesTo compare vaginal lipopolysaccharides (LPS) concentrations between patients with and without bacterial vaginosis (BV), to evaluate the correlation between Prevotella bivia colonization density and LPS concentration, and to determine the impact of LPS on loss of dopamine neurons (DA).MethodsVaginal washes obtained from patients with (n = 43) and without (n = 59) BV were tested for quantity of P. bivia cells using quantitative PCR and for concentrations of LPS using the Limulus Amebocyte Lysate gel clot method. Prevotella bivia, Gardnerella vaginalis and Escherichia coli sonicated cell extracts were also tested for LPS production. DA neuron cells obtained from embryonic day (E) 14.5 pregnant rats were exposed to fluid from eight vaginal washes; tyrosine hydrolase immunoreactive staining was applied for visualization and cell counts.ResultsThe median LPS concentrations were dramatically higher among patients who had symptoms of BV compared to those who did not have symptoms (3235.0 vs 46.4 EU/ml, respectively, P < 0.001); patients who had BV also had much higher colonization densities of P. bivia (0.06 ± 0.36 vs 5.4 ± 2.2 log₁₀ CFU/ml, respectively, P < 0.001).Prevotella bivia cell lysates resulted in a higher LPS concentration (10,713.0 ± 306.6 EU/ml) than either E. coli (4679.0 ± 585.3 EU/ml) or G. vaginalis (0.07 ± 0.01 EU/ml of LPS).The loss of DA neuron was 20–27% in cultures treated with vaginal washes from BV-negative patients and 58–97% in cultures treated with vaginal washes from patients with BV.ConclusionP. bivia produces high LPS concentration, which may create a toxic vaginal environment that damages DA neurons.     

Aqueous humor nitric oxide in patients with central retinal vein occlusion

PurposeIt has been suggested that nitric oxide (NO) has a role in ischemic retinopathies. Since retinal ischemia may develop in retinal vein occlusion, we investigated the presence of nitric oxide in the pathogenesis of central retinal vein occlusion (CRVO).MethodsEighteen consecutive patients with CRVO were included in this study. Aqueous humor specimens were obtained within 21 days of diagnosis. Samples of aqueous humor were also collected from 20 control patients undergoing cataract surgery. For each sample after reduction of nitrate to nitrite with vanadium chloride (VCl₃), we used spectrophotometric method for simultaneous detection of nitrate and nitrite (NO_x).ResultsMean level of aqueous humor NO_x in CRVO and control group was 94.1 ± 23.2 μmol/l and 55.6 ± 11.0 μmol/l, respectively. The difference between two groups was statistically significant (p < 0.0001).ConclusionsOur results may support involvement of nitric oxide in the pathogenesis of CRVO.