Dual Strategy with Adipose-Derived Stem Cells and l-arginine Recovered Cavernosal Functions in a Rat Model of Radical Prostatectomy.

As standard therapy for prostate cancer, radical prostatectomy causes cavernous nerve (CN) injury and increases fibrosis and hypoxia-induced penile structural alterations. This study aimed to determine the potential beneficial effects of adipose-derived stem cells (ADSCs) and l-arginine alone or in combination on the penile erection in a rat model of erectile dysfunction caused by bilateral cavernous nerve transection (CNT). Male rats (n = 35) were randomized into five groups: Sham-operated; CNT (4-weeks); CNT plus ADSCs (1 × 106 cells by intracavernosal injection); CNT plus l-arginine (4 weeks, 10 mg/kg/day, oral); and ADSCs combined with l-arginine in CNT. In vivo erectile responses and in vitro relaxant responses were measured. Western blot and immunohistochemistry analyses were used to determine the expression and localization of endothelial nitric oxide synthase, neuronal nitric oxide synthase, transforming growth factor-beta 1, hypoxia-inducible factor-1 alpha (HIF-1α), and apoptosis markers (Bax and Bcl-2). The ratio of smooth muscle to collagen and nerve regeneration were calculated using Masson's trichrome and nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase staining. The combined treatment restored diminished erectile responses, endothelium-dependent acetylcholine, and electrical field stimulation-induced relaxation of the corpus cavernosum in rats with CNT, whereas either monotherapy produced only partial improvements. All treatment regimens restored increases in the protein expression of HIF-1 and Bax in rats with CNT. The decrease in smooth muscle mass and NADPH-diaphorase-positive nerve fibers was partially ameliorated by monotherapy, whereas combined therapy led to recovery. These findings indicate that combined treatment with ADSCs and l-arginine may restore erectile function in rats with CNT by inhibiting hypoxia-induced neurotoxicity and preserving endothelium function and smooth muscle content.

A Review of the Potential of Nuclear Factor [Erythroid-Derived 2]-like 2 Activation in Autoimmune Diseases.

An autoimmune disease is the consequence of the immune system attacking healthy cells, tissues, and organs by mistake instead of protecting them. Inflammation and oxidative stress (OS) are well-recognized processes occurring in association with acute or chronic impairment of cell homeostasis. The transcription factor Nrf2 (nuclear factor [erythroid-derived 2]-like 2) is of major importance as the defense instrument against OS and alters anti-inflammatory activities related to different pathological states. Researchers have described Nrf2 as a significant regulator of innate immunity. Growing indications suggest that the Nrf2 signaling pathway is deregulated in numerous diseases, including autoimmune disorders. The advantageous outcome of the pharmacological activation of Nrf2 is an essential part of Nrf2-based chemoprevention and intervention in other chronic illnesses, such as neurodegeneration, cardiovascular disease, autoimmune diseases, and chronic kidney and liver disease. Nevertheless, a growing number of investigations have indicated that Nrf2 is already elevated in specific cancer and disease steps, suggesting that the pharmacological agents developed to mitigate the potentially destructive or transformative results associated with the protracted activation of Nrf2 should also be evaluated. The activators of Nrf2 have revealed an improvement in the progress of OS-associated diseases, resulting in immunoregulatory and anti-inflammatory activities; by contrast, the depletion of Nrf2 worsens disease progression. These data strengthen the growing attention to the biological properties of Nrf2 and its possible healing power on diseases. The evidence supporting a correlation between Nrf2 signaling and the most common autoimmune diseases is reviewed here. We focus on the aspects related to the possible effect of Nrf2 activation in ameliorating pathologic conditions based on the role of this regulator of antioxidant genes in the control of inflammation and OS, which are processes related to the progression of autoimmune diseases. Finally, the possibility of Nrf2 activation as a new drug development strategy to target pathogenesis is proposed.

Combined levothyroxine and testosterone treatment for restoring erectile dysfunction in propylthiouracil-induced hypothyroid rats.

BACKGROUND: Sexual dysfunction may indicate severe endocrine diseases. Recent research has suggested a link between hypothyroidism, low testosterone (T) levels, and erectile dysfunction (ED); however, the exact cause is unknown. AIM: We sought to investigate possible beneficial effects of levothyroxine and T alone or in combination on ED in propylthiouracil (PTU)-induced hypothyroid rats. METHODS: Adult Wistar rats (n = 35) were divided into 5 groups: control, PTU-induced hypothyroidism, PTU + levothyroxine, PTU + Sustanon (a mixture of 4 types of T: propionate, phenylpropionate, isocaproate, and decanoate) and PTU + levothyroxine + Sustanon. PTU was given in drinking water for 6 weeks. Four weeks after PTU administration, levothyroxine (20 µg microgram kg/day, oral) and Sustanon (10 mg/kg/week, intramuscular) were given for 2 weeks. Serum levels of total T, triiodothyronine (T3), and thyroxine (T4) were determined. In vivo erectile response and in vitro relaxant responses were measured. Localization of neuronal nitric oxide synthase (nNOS), endothelial NOS (eNOS), and phosphodiesterase type 5 (PDE5) were determined using immunohistochemical analysis. The relative area of smooth muscle to collagen was measured using Masson trichrome staining. OUTCOMES: Outcome variables included in vivo erectile function, in vitro relaxant and contractile responses of corpus cavernosum (CC) strips; protein localization of eNOS, nNOS, and PDE5; and smooth muscle content in penile tissue. RESULTS: The rat model of hypothyroidism showed a significant decline in serum levels of total T, T3, and T4. Levothyroxine increased T3 and T4 levels, whereas Sustanon normalized only total T levels. Combined treatment enhanced all hormone levels. Rats with hypothyroidism displayed the lowest erectile response (P < 0.001 vs controls). Combined treatment returned reduced responses, while partial amelioration was observed after levothyroxine and Sustanon treatment alone. Acetylcholine (P < 0.01 vs controls), electrical field stimulation (P < 0.001 vs controls), and sildenafil-induced relaxant responses (P < 0.05 vs controls) were decreased in the CC strips from hypothyroid rats. The combined treatment increased the reduction in relaxation responses. Levothyroxine and Sustanon restored decreases in eNOS and nNOS expression in the hypothyroid group. There was no significant difference in PDE5 expression among groups. Monotreatment partially enhanced reduced smooth muscle mass, while combined therapy completely recovered. CLINICAL IMPLICATIONS: The combination of thyroid hormones and T is likely to be a therapeutic approach for treatment of hypothyroidism-induced ED in men. STRENGTHS AND LIMITATIONS: Beneficial effects of levothyroxine and Sustanon treatment were shown in vitro and in vivo in PTU-induced hypothyroid rats. The main limitation of the study was the lack of measurement of androgen-sensitive organ weights and luteinizing hormone, follicle-stimulating hormone, and prolactin levels. CONCLUSION: These findings demonstrate that neurogenic and endothelium-dependent relaxation responses are reduced by hypothyroidism, which is detrimental to T levels and erectile responses. Levothyroxine and Sustanon combination medication was able to counteract this effect.

Evaluation of non-radioactive endpoints of ex vivo local lymph node assay-BrdU to investigate select contact sensitizers.

The present study sought to verify the utility of the non-radioactive endpoints LLNA BrdU (5-bromo-2'-deoxyuridine) ex vivo incorporation and cytokine release using auricular lymph node cells isolated from BALB/c mice topically treated with a strong (formaldehyde or p-phenylene-diamine [PPD]), moderate sensitizer (cinnamal), or weak sensitizer (eugenol). Stimulation index (SI) and EC3 values were calculated for each agent. Based on the results of ex vivo LLNA-BrdU assays, EC3 values were calculated to be 0.29, 0.09, 1.91, and 16.60% for formaldehyde, PPD, cinnamal, and eugenol, respectively. These results were in good agreement with data from previous standard radioactive LLNA. Cytokine analyses indicated T(H)1 and T(H)2 cytokine involvement in the regulation of murine contact allergy and these could be utilized as endpoints in assessments of contact allergy in mice. In conclusion, the current study provided evidence that the non-radioactive endpoint LLNA BrdU ex vivo incorporation could be of use as a viable alternative approach to assess the skin sensitization potential of test compound with respect to improving animal welfare. This is of particular importance in the case of any laboratory where it might be difficult to handle and/or readily employ radioisotopes. Further studies will be required to confirm--across test agents--the reproducibility as well as the limits of utility of this new ex vivo BrdU method.

The relationship of PON1 QR 192 and LM 55 polymorphisms with serum paraoxonase activities of Turkish diabetic patients.

Paraoxonase (PON1) is a serum esterase responsible for the protection against xenobiotics toxicity such as paraoxon. Alterations in PON1 concentrations have been reported in a variety of diseases including diabetes mellitus (DM). It has been shown that the serum PON1 concentration and activity are decreased in patients with both type 1 and type 2 DM. This study aimed to investigate the lipid profiles and the relationship between PON1 activity and PON1, QR192 and LM55 polymorphisms in Turkish type 2 diabetic patients and non-diabetic control subjects. According to our results, RR variant had significantly higher PON activity than QQ and QR variants (p < 0.01) and LL variant had significantly higher PON activity than MM variant in both control and patient groups (p < 0.05). In conclusion, we found that PON1 192RR and 55LL genotypes are associated with higher PON activity than QQ and MM genotypes. This may be more protective to lipid peroxidation.

Evaluation of auricular lymph node cell lymphocyte proliferation and cytokine production as non-radioactive endpoints during murine contact allergy.

The murine local lymph node assay (LLNA) has been developed as a test method to assess allergic contact dermatitis. In spite of the validity of the LLNA, attention was drawn to the two disadvantages: use of radioactivity for in vivo measurement of lymph node cell proliferation ([(3)H]-thymidine labeling) and the possibility of false positive results caused by non-specific cell activation as a result of inflammatory processes in the skin (irritation). We aimed to investigate the following non-radioactive endpoints of LLNA: 5-bromo-2'-deoxyuridine (BrdU) incorporation ex vivo and in vivo, in vivo and ex vivo cytokine production with or without phytohemagglutinin (PHA) stimulation. Here, 8-12-week-old female BALB/c mice were treated topically with the strong sensitizer 2,4-dinitrochlorobenzene (DNCB) in acetone:olive oil (AOO, 4:1 [v/v]) at levels of 0.025, 0.05, 0.01, or 0.25% (w/v). Ear thickness was also measured to determine the differentiation index (DI) indicating the proportion of non-specific activation due to irritating properties of test compound. At the concentration of 0.05%, stimulation index (SI) value was found to be 3 for DNCB based on in vivo and ex vivo BrdU incorporation. The results of the in vivo and ex vivo non-radioactive LLNA assays were compatible both with each other and with previous radioactive LLNA data. Our results indicate that non-radioactive endpoints may be used as an alternative to the [(3)H]-thymidine LLNA. The levels of T(H)1 cytokines (IL-2 and IFNγ) and T(H)2 cytokines (IL-4 and IL-5) in lymph node cell cultures were significantly (P < 0.01) increased when DNCB was applied at the concentrations of 0.05 and 0.1%, respectively. As the DI was > 1, the applied concentrations of DNCB caused only allergic effect but not any irritant effect. This study reports that the use of these non-radioactive endpoints can assess allergic contact dermatitis caused by chemicals.

Correlation of Ochratoxin A exposure to urinary levels of 8-hydroxydeoxyguanosine and malondialdehyde in a Turkish population.

Ochratoxin A is one of the most abundant food- contaminating mycotoxins in the world that is immunosuppressive, genotoxic, teratogenic and carcinogenic. Malondialdehyde is a naturally occurring product of lipid peroxidation that is mutagenic and carcinogenic. 8-Hydroxydeoxyguanosine is produced during the interaction of reactive oxygen species and DNA. In this study, Ochratoxin A, malondialdehyde and 8-Hydroxydeoxyguanosine levels of individuals in the study group were measured and results were correlated with each other. Additionally, the correlation of biomarker levels to smoking habit, alcohol and coffee consumption, age and gender of individuals was investigated. As a result of these assessments, a significant correlation was observed between Ochratoxin A exposures and malondialdehyde and 8-Hydroxydeoxyguanosine levels.

Are PON1 Q/R 192 and M/L 55 polymorphisms risk factors for diabetes complications in Turkish population?

OBJECTIVES: We investigated whether the human serum paraoxonase (PON1) Q/R 192 and M/L 55 polymorphisms are associated with the complications of the type 2 diabetes (T2D). DESIGN AND METHODS: Study group was consisted of 50 healthy subjects and 100 type 2 diabetes mellitus (DM) patients. Following measuring of serum PON1 activity, isolation of DNA and genotyping analyses were performed. RESULTS: PON1 activity of the patients with complications was significantly reduced by 23.5% compared to the group of diabetic patients and by 26.3% than the controls. According to multivariate analysis, we observed a three times significant effect of Q/R 192 polymorphism on the susceptibility to the occurrence of complications. CONCLUSIONS: Protective effects of paraoxonase against peroxidation of LDL particles are important in T2D complications. Although both of the two polymorphisms are associated, 192 polymorphism seems to be stronger predictor of the risk of diabetic complications.

Possible effect of gene polymorphisms on the release of TNFα and IL1 cytokines in coal workers' pneumoconiosis.

It has been shown that coal dust exposure stimulates inflammatory response leading to increased release of cytokines from monocytes such as TNF-alpha and IL1. These released cytokines play the key role in the pathogenesis of pneumoconiosis including coal workers' pneumoconiosis. In this study, we investigated TNFA, IL1A, IL1B and IL1RA genes variations on basal, lipopolysaccharide and coal dust-induced cytokine release from blood monocytes of homozygous allele and minor variant allele carriers in Turkish coal workers and CWP patients. According to the genotyping results, TNFA -238 gene polymorphism was found as a risk factor in CWP development (OR=3.79) and to in vitro results; release of both TNF-alpha and IL1 cytokines from the monocytes in CWP patients was significantly increased compared to the healthy workers. Also, LPS and coal dust stimulated release of TNF-alpha, which was significantly higher in allele 2 carriers compared to subjects carrying allele 1 in both the groups. These data suggest that the coal dust-induced release of TNF-alpha from monocytes may be a useful biomarker of CWP.

Association of cytokine gene polymorphisms in CWP and its severity in Turkish coal workers.

BACKGROUND: Cytokines appear to play a key role in some inflammatory reactions affecting the interactions among pro- and anti-inflammatory mechanisms that result in several diseases such as coal workers' pneumoconiosis (CWP). In this study, to determine the cytokine gene profiles of Turkish coal miners, we performed genotyping analysis to investigate the polymorphisms of CWP-related pro-inflammatory (TNFA, IL1A, IL1B, and IL6) and anti-inflammatory cytokines (IL-1RN and TGFB1). An additional goal was to observe whether these cytokine gene polymorphisms influence the development risk and severity of. METHODS: Genotyping was carried out by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. RESULTS: TNFA (-238) gene polymorphism principally affected CWP development and severity (OR = 3.47: 95% CI, 1.12-10.77 and OR = 4.30: 95% CI, 1.25-14.74, respectively) and also risk of CWP (OR = 3.79: 95% CI, 1.37-10.46). The TNFA (-308) variant was associated with a risk for the CWP severity (OR = 2.84: 95% CI, 1.08-7.39). A protective effect of IL6 was found on the development (OR = 0.48: 95% CI, 0.21-0.93) and severity of CWP (OR = 0.37: 95% CI, 0.15-0.91). CONCLUSIONS: We suggest that TNFA (-238) variant may be a risk factor in both development and the severity of CWP, while TNFA (-308) variant seems to be important only in disease severity. On the other hand, IL6 variant may have a protective effect on the development and disease severity.

Effects of occupational polycyclic aromatic hydrocarbon exposure on T-lymphocyte functions and natural killer cell activity in asphalt and coke oven workers.

Polycyclic aromatic hydrocarbons (PAHs) are environmental carcinogens exhibiting potent immunosuppressive properties. In order to determine PAH-induced immunotoxicity in humans, we investigated possible immunomodulating effects on T-lymphocyte proliferative responses and natural killer (NK) cell activities, at two different exposure levels, in asphalt and coke oven workers. We evaluated the efficiency of urinary 1-hy droxypyrene as a measure of exposure to PAHs. We found a statistically significant inhibition in T-lymphocyte proliferative responses of asphalt and coke oven workers compared to the controls. On the other hand, interestingly, we found significantly higher NK cell activities at three effector:target (E:T) ratios in the asphalt group compared to coke oven and control groups. We conclude that PAHs may cause suppression of T-lymphocyte proliferation at both exposure levels and augment NK cell activity only at low levels of exposure. Our results are in line with others reported in the literature indicating that chronic exposure to PAHs at different levels may alter some immune responses in different ways.

The oxidative DNA base damage in testes of rats after intraperitoneal cadmium injection.

Cadmium is known to be a carcinogenic metal that especially its compounds have sufficient evidence in both humans and experimental animals beneath its environmental effects. Testis tissue is highly sensitive to the effects of cadmium. It is proposed that cadmium also increases oxygen derived free radicals and lipid peroxidation. As indicators of oxidative DNA damage, 6 oxidative DNA bases were determined by using Gas Chromatography/Mass Spectrometry-Selected Ion Monitoring technique. 45 Sprague-Dawley rats (225-300 g) were used as experimental animals and were divided into 3 groups of 15 rats. A single 2 mg NaCl/kg body wt, 0,5 and 1,25 mg CdCl2/kg body wt were injected intraperitoneally to control, low and high dose groups, respectively. 5-OH Cytosine, 8-OH Adenine and Fapy Guanine lesions were elevated significantly in high dose group in the first day. A clear dose-response relationship was seen between dose groups and 8-OH Adenine levels related with time in all periods. There was a significant dose-response relationship in 2-OH Adenine, Fapy Guanine and 8-OH Guanine, especially in the second week suggesting the inhibition of XPA protein by cadmium after first week. In contrast, the observation of a significant decrease of 5-OH Cytosine levels after first week showed that cadmium could not affect the enzymes repairing the cytosine base lesions.