Modified protocol for improvement of differentiation potential of menstrual blood-derived stem cells into adipogenic lineage.

OBJECTIVES: To characterize potency of menstrual blood-derived stem cells (MenSCs) for future cell therapies, we examined differentiation potential of MenSCs into adipocytes. MATERIALS AND METHODS: Differentiation potential of MenSCs in comparison to bone marrow stem cells (BMSCs) was assessed in conventional culture medium. Differentiation potential of MenSCs into adipocytes was improved using different combinations of growth factors and hormones. RESULTS: First, we demonstrated that MenSCs preserve their appearance and karyotypic stability during passages. Although these cells express mesenchymal stem cells markers, they cannot simply be classified as mesenchymal stem cells due to expression of embryonic stem cells marker, OCT-4. Oil red O staining showed that differentiated MenSCs in conventional medium with/without retinoic acid (protocols 1 and 2) did not attain adipocyte characteristics, whereas differentiated BMSCs in conventional medium accumulated oil vacuoles typically. Nevertheless, real-time RT-PCR results showed that LPL gene expression was up-regulated in both protocols 1 and 2, whereas LEPR was up-regulated only in protocol 2 (fortified with retinoic acid). Surprisingly, protocol 3 (including rosiglitazone) had odd influence on mRNA expression of all genes (LEPR, LPL and PPAR-γ). Oil red O staining confirmed fat-producing ability of MenSCs under protocol 3. CONCLUSIONS: Presented data suggest an efficient differentiation protocol for in vitro production of MenSC-derived adipocytes. These cells are suggested to be an apt alternative to BMSCs for future stem cell therapy of soft tissue injuries.

Regulation of luteinizing hormone receptor in hippocampal neurons following different long-lasting treatments of castrated adult rats.

The aim of this study was to investigate the effects of different Luteinizing hormone (LH) and steroid hormones levels on LH receptor (LHR) expression in the hippocampal cells. Rats (24 males and 24 females) were assigned to four groups: one control and three experimental [gonadectomy (GDX), gonadectomy + gonadotropin releasing hormone analogue (GDX+GnRHa) and GDX+GnRHa+estradiol (E2) or testosterone (T)] independently for each gender. All experimental rats were gonadectomized; then GnRHa was administrated to GDX+GnRHa group, and GnRHa plus steroid hormone to GDX+GnRHa+E2 or T group in both genders for four-month. LHR mRNA expression and its protein level in hippocampal cells were measured using QRT-PCR and Western blotting. Quantification of mRNA revealed a decrease in LHR transcripts level in GDX+GnRHa group of females. A significant change was observed between GDX groups and GDX+GnRHa+E2 or T versus GDX+GnRHa group in females. High levels of LH decreased significantly the immature isoform of LHR in GDX group compared to control group in both genders, but low LH concentrations in GDX+GnRHa group induced immature LHR isoform production only in females. Therefore increased LH concentration induces production of incomplete LHR transcripts in hippocampal cells and decreases immature LHR at the protein level. This implies that LH decreases the efficiency of translation through either producing non-functional LHR molecules or preventing their translation.

Comparison of intracytoplasmic sperm injection outcomes between spermatozoa retrieved from testicular biopsy and from ejaculation in cryptozoospermic men.

The infrequent presence of spermatozoa in cryptozoospermic men ejaculate is a limiting factor in the treatment of them. Sometimes, this consideration impels us to apply meticulous microscopic search in ejaculate or testicular sperm extraction (TESE) method. The aim of this study was to assess putative effectiveness of sperm origin, ejaculated or testicular, in cryptozoospermia treatment. In this context, were evaluated intracytoplasmic sperm injection (ICSI) outcomes in two parameters including fertilisation rate (2PN) and embryo quality, independently. We compared the outcome in two groups: patients who underwent ejaculate/ICSI and ones who underwent TESE/ICSI process. Nineteen ICSI cycles performed with testicular spermatozoa and the rest of cycles (n = 208) carried out with ejaculated spermatozoa. Result analysis showed similar fertilisation rate between testicular and ejaculated spermatozoa (respectively, 60% versus 68%, P ≥ 0.05). Also, on the other hand, embryo quality did not show significant differences between two groups, except grade A with low significance. With regard to almost equal performance of both methods in results and being invasive of TESE as surgical sperm retrieval method, the use of ejaculated sperm more than testicular sperm should be recommended in patients with cryptozoospermia whenever possible.

Relationship between sperm chromatin status and ICSI outcome in men with obstructive azoospermia and unexplained infertile normozoospermia.

This study was done to evaluate the effect of sperm source on chromatin integrity and ICSI outcomes. One hundred and thirteen samples containing epididymal aspirates of 57 obstructive azoospermic men and 56-ejaculated semen of normozoospermic men were included in this study. Sperm chromatin status was evaluated by Chromomycin A3 (CMA3), Aniline Blue (AB) and Toluidine Blue (TB). Fertilization rate and embryo quality were recorded. In epididymal group the percentage of sperms stained with AB, CMA3 and TB were significantly higher compared to ejaculate group while fertilization rate (60.6% vs. 74.04%) was significantly lower. However, embryo quality was not significantly different between two groups. In addition, abnormal sperm chromatin condensation and DNA fragmentation were not correlated with fertilization rate and embryo quality. Our results highlight the role of epididymis in sperm maturation and confirm that ICSI using ejaculated sperm is the gold standard for treatment of infertile men.