Testosterone Propionate Promotes Proliferation and Viability of Bone Marrow Mesenchymal Stem Cells while Preserving Their Characteristics and Inducing Their Anti-Cancer Efficacy

Background: Various studies have reported the effects of testosterone on different cell types, yet bone marrow-derived mesenchymal stem cells' cellular responses to testosterone remain unknown. Aims: To investigate the effects of testosterone propionate, an oil-soluble short-acting form of testosterone, on human bone marrow-derived mesenchymal stem cells' proliferation and viability after 24 hours of incubation. We also investigated the impact of testosterone propionate on bone marrow-derived mesenchymal stem cell's polarization and cytotoxicity on K562 leukemia cell line. Study Design: In vitro study. Methods: We expanded commercially available bone marrow derived mesenchymal stem cells in vitro and treated them with testosterone propionate at concentrations ranging from 10-6-10-10 M for 24 hours. Ideal concentration was determined by evaluating cellular viability and proliferation with Annexin V/Propidium Iodide assay and carboxyfluorescein succinimidyl ester staining. The characteristic features of bone marrow-derived mesenchymal stem cells were evaluated by immunophenotyping and investigating their differentiation capacities. Bone marrow-derived mesenchymal stem cells' cytotoxic properties upon testosterone propionate treatment were determined by co-culturing the cells with K562 cells and with confocal imaging investigating polarization. Results: Testosterone propionate promoted proliferation and maintained the viability of bone marrow-derived mesenchymal stem at 10-8 M concentration. Further evaluations were conducted with the determined dose. The results showed that, apart from promoting mesenchymal stem cells' polarization and increasing their cytotoxicity on K562 cells, testosterone propionate did not alter differentiation capacities of bone marrow-derived mesenchymal stem cells and certain cell surface markers, but led to a significant increase in HLA-DR expression. Conclusion: The findings reveal that testosterone propionate promotes the proliferation and survival of bone marrow-derived mesenchymal stem cells in a dose-dependent manner without hampering their differentiation capacities, induces their polarization to the pro-inflammatory phenotype, and increases their cytotoxicity on the K562 cell line.

A preliminary study of tracking B-cell kinetics in patients with lung transplantation by monitoring kappa-deleting recombination excision circles.

Background: This study aims to evaluate humoral immune system response by measuring copy numbers of kappa-deleting recombination excision circles (KREC) gene segment from B lymphocytes in patients with lung transplantation. Methods: Between September 2015 and November 2016, a total of 11 patients (8 males, 3 females; mean age: 45.4±12.0 years; range, 23 to 59 years) who underwent lung transplantation with different primary indications were included. The copy numbers of KREC gene segment were quantified using real-time polymerase chain reaction method in peripheral blood samples collected pre- and post-transplantation. The samples of the patients were compared with the KREC l evels i n deoxyribonucleic acid extracted from blood samples of healthy children. Results: There was no significant change in KREC levels between pre- and post-operation (p=0.594 and p=0.657), although the median values indicated that the highest increase in the KREC levels (7x105- 12x105; 85-170) was on Day 7 of transplantation. There was a positive correlation between the KREC levels (mL in blood) and lymphocytes at 24 h after transplantation (p=0.043) and between KREC copies per 106 of blood and age on Day 7. Conclusion: Our preliminary results suggest that KREC l evels a s an indicator of B lymphocyte production are elevated after lung transplantation. A prognostic algorithm by tracking B cell kinetics after post-transplantation for long-term follow-up can be developed following the confirmation of these preliminary results with more patient samples.

5-Alfa reductase type 2 (SRD5A2) gene rs523349 polymorphism is not associated with non-obstructive azoospermia in Turkish patients.

Spermatogenesis is an androgen-dependent event, and testosterone is the major androgen source. The enzyme 5-alpha reductase converts testosterone to dihydrotestosterone (DHT) in testicular and peripheral tissues. Polymorphisms in genes encoding 5-alpha reductase may be associated with impaired male fertility. The present study aimed to investigate the relationship between 5-alpha reductase type 2 (SRD5A2) gene rs523349 polymorphism and non-obstructive azoospermia (NOA) in Turkish patients. The study included 75 NOA patients and 43 fertile men from Turkey. No significant relationship was found between SRD5A2 gene rs523349 polymorphism and male infertility (P = 0.071). There was a statistically significant difference in total testosterone level and total testis volume between NOA patients and the control groups, however there was no significant difference between serum follicle-stimulating hormone and luteinizing hormone levels. Our results showed that SRD5A2 gene rs523349 polymorphism was not associated with NOA in Turkish patients.

House Dust Mites Confer a Distinct Immunological Feature among Dermatitis.

Atopic dermatitis (AD) is a heterogeneous disease with regard to clinical phenotype and natural history. We investigated T cell subtypes and cytokine responses in peripheral blood and skin lesions of AD patients with various sensitivities. Immunological studies were performed in 27 subjects: 9 house dust mite (HDM)-sensitized; 6 subjects with sensitizations other than HDM; 7 non-allergic AD patients and 5 healthy controls. Among those, skin biopsy samples of 13 subjects were evaluated for immunohistochemical analyses, as well. The mean age was 8.93±5.17 years. HDM-allergic AD emerged as a distinct immunologic phenotype, with higher production of interleukin (IL)-4, -5, -2 both at rest and when stimulated by Der p1 or SEB along with higher Th17. As for TH17 cell percentage, it was increased in all AD groups compared to healthy controls, while HDM-allergic group was distinguished with a significantly lower production of IL-17. Patients with sensitizations other than HDM were mostly similar to non-allergic AD, with increased Th17 and CD4+CD69+interferon-gamma (IFN-γ)+ T cells percentage. The biopsy of lesional skin showed that HDM-allergic AD had lower IFN-γ and IFN-γ co-expressing CD8+ T cells compared to patients with other sensitizations (p=0.03 and p=0.04, respectively). Among the HDM allergic patients, pairwise comparison of lesional versus non-lesional skin revealed higher CD4+ T cells numbers, expression of forkhead box P3 (Foxp3) and T-cell-specific transcription factor (T-bet) (p=0.018, p=0.018, p=0.018, respectively). HDM-allergic AD is a distinct subtype with a predominant skewing in Th2 and higher Th17 cell percentage along with a blunted Th1 response in the skin, all of which may have therapeutic implications.

Pleiotropic and Renoprotective Effects of Erythropoietin Beta on Experimental Diabetic Nephropathy Model.

BACKGROUND: This study aimed at investigating the possible protective effect of erythropoietin beta on experimental diabetic nephropathy (DN) model in rats. METHODS: Sprague Dawley rats (n = 32) were allocated into 4 equal groups of 8 each, the control (Group C), diabetes (Group D), erythropoietin beta (Group E), and erythropoietin beta treated DN (Group E + D) groups. Streptozocin (65 mg/kg) was used to induce diabetes in 10-week old rats. Erythropoietin beta was given intraperitoneally at a dose of 500 IU/kg/3 days of a week for 12 weeks. Renal function parameters, intrarenal levels and activities of oxidative stress biomarkers, serum inflammatory parameters and kidney histology were determined. RESULTS: Group E + D had lower mean albumin-to-creatinine ratio (p < 0.001) as well as higher creatinine clearance (p = 0.035) than the diabetic rats (Group D). Intrarenal malondialdehyde levels were significantly lower (p = 0.004); glutathione (GSH) levels (p = 0.003), GSH peroxidase (p = 0.004) and superoxide dismutase (p < 0.005) activities of renal tissue were significantly higher in Group E + D than in Group D. The mean serum levels of interleukin-4 (p < 0.005), interleukin 1 beta (p = 0.012), interferon gamma (p = 0.018) and tumor necrosis factor alpha (p < 0.005) were significantly lower; serum levels of monocyte chemoattractant protein 1 (p = 0.018) was significantly higher in Group E + D when compared to Group D. The mean scores of tubulointerstitial inflammation (p = 0.004), tubular injury (p = 0.013) and interstitial fibrosis (p = 0.003) were also lower in Group E + D when compared to Group D. CONCLUSION: Our data seem to suggest a potential role of erythropoietin beta for reducing the progression of DN in an experimental rat model. This protective effect is, in part, attributable to the suppression of the inflammatory response and oxidative damage.

Effects of paricalcitol and aliskiren combination therapy on experimental diabetic nephropathy model in rats.

BACKGROUND/AIMS: The aim of the present study was to investigate the effect of combination of aliskiren with paricalcitol on experimental diabetic nephropathy (DN) model in rats. METHODS: Forty male Sprague Dawley rats were divided into 5 groups of 8 rats each, namely the control (Group C), diabetes (Group D), aliskiren (Group A), paricalcitol (Group P), and aliskiren plus paricalcitol (Group A+P) groups. Aliskiren was given by oral-gavage at a dose of 50 mg/kg/day once daily for 12 weeks. Paricalcitol was given by intraperitoneally at a dose of 0,4 µg/kg/three day of week for 12 weeks. Renal function parameters, oxidative stress biomarkers, mRNA expression of renin-angiotensin system parameters and kidney histology were determined. RESULTS: Group A+P had lower mean albümin-to-creatinine ratio (ACR) (p=0.004) as well as higher creatinine clearance (CCr) (p<0.005) than the diabetic rats (Group D). Combination therapy significantly increased CCr (Group A+P vs. Group A, p<0.005; Group A+P vs. Group P, p=0.022) and reduced ACR (Group A+P vs. Group A, p=0.018; Group A+P vs. Group P, p<0.005) when compared to monotherapy. Serum malondialdehyde levels were significantly lower (p=0.004); glutathion levels (p=0.003), glutathion peroxidase (p=0.004) and superoxide dismutase (p<0.005) activities were significantly higher in group A+P than in group D. The mean scores of mRNA expression of renin (p<0.005), angiotensin II (p=0.012) and angiotensin type 1 receptor (p=0.018) in group A+P were significantly lower. Although combination therapy showed no additional effect on oxidative system, renin-angiotensin system and renal histology, aliskiren plus paricalcitol significantly decreased interstitial fibrosis volume when compared to monotherapy (Group A+P vs. Group A, p<0.005; Group A+P vs. Group P, p=0.002). CONCLUSION: Our data seem to suggest a potential role of aliskiren plus paricalcitol acting synergystically for reducing the progression of diabetic nephropathy in an experimental rat model.

Aberrant expression of c-met and HGF/c-met pathway provides survival advantage in B-chronic lymphocytic leukemia.

BACKGROUND: B-chronic lymphocytic leukemia (B-CLL) is characterized by accumulation of CD5(+) B lymphocytes. Decreased VLA-4 (Cd49d/CD29) and CD11a expression and defective adhesion in B-CLL have been previously shown, although there was no substantial data about its importance in immunobiology of B-CLL. The hepatocyte growth factor (HGF) receptor, c-met, plays a role in adhesion by acting on VLA-4. c-met and VLA-4 share crucial signaling molecules in cell survival. In this study, relationship between expressions of c-met and CD49d, CD11a, and additional common signaling molecules in B-CLL was investigated. METHODS: White blood cells from 24 patients with CLL were studied by flow cytometry and/or western blotting prior to and after culturing with recombinant HGF. HGF level from sera was measured with a bead-based flow cytometric assay. RESULTS: c-metα and c-metß were expressed on B-CLL cells, while no expression was observed on normal donor CD19+ cells. This increase was inversely correlated with decreased expression of adhesion molecules. Serum level of HGF in B-CLL was found to be increased. In vitro experiments showed that HGF supported survival in B-CLL cells supporting the possible function of HGF/c-met pathway in B-CLL. Furthermore, expressions of critical signaling molecules shared by both VLA-4 and HGF/c-met systems including Bcl-XL, Akt, PI3K, and phospho-bad(136) following HGF stimulations of B-CLL cells have been found to be increased. CONCLUSION: Increased expression of c-met and HGF may bypass the importance of expression of critical adhesion molecules and support survival of B-CLL cells. c-met, being one of the surface tyrosine kinases, may serve as a target for future therapies in B-CLL meriting more attention.