Effect of Zirconium oxide nanoparticle on serum level of testosterone and spermatogenesis in the rat: An experimental study.

BACKGROUND: Zirconium nanoparticles are used as health agents, pharmaceutical carriers, and in dental and orthopedic implants. OBJECTIVE: This studyaimed to investigate the effects of Zirconium oxide nanoparticles on the process of spermatogenesis in rat. MATERIALS AND METHODS: In this experimental study, 32 male Wistar rats (150-200 gr), with range of age 2.5 to 3 months were used and divided into four groups of eight per each. The control group received 0.5 ml of distilled water and the three experimental groups received 50, 200, and 400 ppm doses of Zirconium oxide nanoparticles solution over a 30-day period, respectively. At the end of the experiment, tissue sections were taken from the testis and stained with hematoxylin-eosin. Serum concentration of testosterone was measured by enzyme-linked immunosorbent assay. RESULTS: In the experimental group receiving 400 ppm Zirconium oxide nanoparticles, the number of Spermatogonia cells (p ≤ 0.01), Spermatocytes (p ≤ 0.01), Spermatids (p ≤ 0.001), and sertoli and Leydig cells (p ≤ 0.05) showed a significant decrease compared to the control group. Serum testosterone concentration did not change significantly in all experimental groups receiving Zirconium oxide nanoparticles compared to the control group. Experimental group received 400 ppm Zirconium oxide nanoparticles shrinkage of seminal tubules and reduced lumen space compared to control group. CONCLUSION: Zirconium oxide nanoparticles are likely to damage the testes by increasing Reactive oxygen species production and free radicals.

ROCK Y-27632 Inhibitor, Ascorbic Acid, and Trehalose Increase Survival of Human Wharton Jelly Mesenchymal Stem Cells After Cryopreservation.

OBJECTIVES: Wharton jelly mesenchymal stem cells are good candidates for application in different aspects of regenerative medicine, and their long-time banking is important. In this study, the effects of trehalose, ascorbic acid, and Y-27632 on proliferation and survival rate of these cells after cryopreservation were investigated. MATERIALS AND METHODS: Mesenchymal stem cells were isolated from human umbilical cord Wharton jelly and frozen using a slow-rate cooling process. Different concentrations of trehalose (35, 75, and 125 mM), ascorbic acid (0.06, 0.125, 0.25, and 0.5 mM), and Y-27632 (10 µM) were used to treat culture medium and/or to supplement freezing medium. Assessment of cell viability after thawing was performed using Trypan blue staining, and MTT assay was performed to measure the cell proliferation rate. RESULTS: We observed significantly increased postthaw viability, increased cell proliferation, and decreased doubling time of cells when 75 mM trehalose, 0.25 and 0.5 mM ascorbic acid, and 10 mM Y-27632 were used. In addition, increased viability, proliferation, and attachment were observed after 24 hours of pretreatment with these cryoprotective agents and when they were added to conventional freezing medium. CONCLUSIONS: The use of different cryoprotective agents in culture and freezing media could be useful for long-term storage of Wharton jelly mesenchymal stem cells.

In vitro differentiation of human umbilical cord Wharton's jelly mesenchymal stromal cells to insulin producing clusters.

AIM: To investigate the differentiation of human Wharton's jelly derived mesenchymal stromal cells (WJ-MSCs) to insulin producing clusters (IPC) this study was conducted. METHODS: The umbilical cords samples were collected from full term caesarian section mothers and the WJ-MSCS were cultured from tissue explants in High glucose-Dulbecco's Modified Eagle Medium (H-DMEM); H-DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics. The expression of CD90, CD44, CD105, CD34 and CD133 as well as osteogenic and adipogenic differentiation of cells in appropriate medium were also evaluated. The cells were differentiated toward IPC with changing the culture medium and adding the small molecules such as nicotinic acid, epidermal growth factor, and exendin-4 during 3 wk period. The gene expression of PDX1, NGN3, Glut2, insulin was monitored by reveres transcription polymerase chain reaction method. The differentiated clusters were stained with Dithizone (DTZ) which confirms the presence of insulin granules. The insulin challenge test (low and high glucose concentration in Krebs-Ringer HEPES buffer) was also used to evaluate the functional properties of differentiated clusters. RESULTS: WJ-MSCS were positive for mesenchymal surface markers (CD90, CD44, CD105), and negative for CD34 and CD133. The accumulation of lipid vacuoles and deposition of calcium mineral in cells were considered as adipogenic and osteogenic potential of WJ-MSCS. The cells also expressed the transcriptional factors such as Nanog and OCT4. During this three step differentiation, the WJ-MSCS morphology was gradually changed from spindle shaped cells in to epithelioid cells and eventually to three dimensional clusters. The clusters expressed PDX1, NGN3, Glut2, and insulin. The cells became bright red color when stained with DTZ and the insulin secretion was also confirmed. In glucose challenge test a significant increase in insulin secretion from 0.91 ± 0.04 µIu/mL (2.8 mmol/L glucose) to to 8.34 ± 0.45 µIu/mL (16.7 mmol/L glucose) was recorded (P < 0.05). The insulin secretion of undifferentiated WJ-MSCS was not changed in this challenge test. CONCLUSION: WJ-MSCs have the ability to differentiate in to islet-like cells in vitro. However, this process needs further optimization in order to generate efficient and functional IPCs.

Immunomodulatory effects of mice mesenchymal stem cells on maturation and activation of dendritic cells.

BACKGROUND: Mesenchymal stem cells (MSCs) possess a wide range of immunomodulatory functions mostly in immune cells including dendritic cells (DCs). DCs are the key cells in the immune response and play an important role in initiating cell-mediated immunity. OBJECTIVE: To evaluate the immunomodulatory effects of MSCs supernatant on maturation and function of DCs. METHODS: Bone marrow derived mice MSCs were isolated and cultured. Twenty-four, forty-eight and seventy-two hours after passage 6, supernatants were collected and MSCs were assessed by flow cytometric analysis for the expression of CD34, CD44, CD45 and SCA-1. Splenic DCs were isolated using MACS and then co-cultured with MSCs supernatant. Expression of CD86, CD40 and MHC-II on DCs were also evaluated by flow cytometry. H3-thymidine incorporation by proliferating T cells was determined in two separate MLR assay settings. In one setting, DCs were co-cultured with T cells in the presence of MSCs supernatant, and in the other setting DCs were treated with MSCs supernatant and then were co-cultured with T cells. Production of IL-12, IL-6 and IL-10 cytokines was measured in the supernatant of DCs treated with MSCs supernatant. We also measured IFN-γ and IL-4 levels in MLR supernatant. RESULTS: The results showed that 72h MSCs supernatant could decrease the expression of MHC-II and CD86. The T cell proliferation was inhibited in the presence of MSCs supernatant and MSCs supernatant treated DCs as demonstrated by MLR assay. A significant increase in IL-4 level and a non significant decrease in IFN-γ level in MLR supernatant was observed. However, IL-6, IL-10 and IL-12 production did not change significantly. CONCLUSION: MSCs supernatant has a time dependent effect on the maturation of DCs. Also, it could alter cytokine production from responding T cells toward Th2. Generally, the findings of this study supported the immunomodulatory effect of MSCs supernatant on DCs maturation and function.