Icariin Enhances Transdifferentiation Of BMSCs Into Leydig-like Cells In Vivo

Research background and objectiveThe function of Leydig cells, which are the predominant source of the male sex steroid hormone testosterone, decreases in the aging males, diabetic patients and in male rectal cancer patients receving EBRT. What’s more, androgen deficiency cursed152per100000children with Klinefelter syndrome. Extral hormone intake has become conventional treatment for these patients and is in favor of the maintenance of male secondary sexual characteristics. Whereas, extral testosterone administration is hard for olders with amnesia and children considering the levels of testosterone are dynamic during puberty. What’s more, long time of androgen supplement therapy is not easy to achieve a reasonable dose, there may be side effects and potential risks. Biotherapy, especially autologous stem cells transplantation, has been taking consideration. We hope that the feedback and the negative feedback mechanism supply the required hormone of the body’s physiological activities in order toavoid excessive and inadequate of supplement under the hypothalamus-pituitary-gonadal axis adjustment of the patient s after the stem cell transplantation, this method has the potential to solve the number of the leydig cells decreases and the function of the leydig cells declines.Bone marrow stem cells are adult stem cells which have the ability to differentiate into multiple mesodermal cells. At the same time because it can be used as autologous transplantation of stem cell, there is no ethics problem but also can effectively avoid immune rejection. Leydig cells are originated from the mesoderm. Recent study recovered BMSCs, through stable transfection with steroidogenic factor-1, could transdifferentiated into testicular Leydig-like cells, originating from the intermediate mesoderm. Steroidogenic factor-1has been demonstrated to be a primary transcriptional regulator of steroidogenic-related genes. Icariin, a major flavonoid isolated from Herba epimedii, has been shown to have therapeutic potential in the management of hypoadrogenism. The modern medical studies have shown that icariin has a variety of pharmacological effects, such as improving immunity, regulating endocrine, improving cardiovascular function, especially increasing testosterone levels in vivo. The present study was to evaluate impact of icariin on the transdifferentiation of BMSCs into Leydig-like cells in the testis of rats.Materials and methods4-wk-old SD rats of SPF grade were randomly divided into5groups with10in each:a sham control(group A), a irradiated model(group B), one BMSCs transplanted group(group C), a low dose icariin group after stem cell transplantation(group D) and a high dose icariin group after stem cell transplantation (group E). Group A is normal feeding, Testis of recipient animals of group B, C, D and E were irradiated by2fraction of60Co gamma ray (6Gy+8Gy) with24hours apart. At passage9BMSCs (1*10^6) were injected into irradiated testis at pi (post irradiation)1weeks. Group D and E with transplanted BMSCs were administrated with icariin2mg/100g,5mg/100g respectively for8weeks. Animals were sacrificed, body weight of rats was measured, blood and testis were collected after the last administration of icariin. Parts of testis were fixed in formalin for HE staining and immunofluorescence analysis. Parts were stored at-80degrees for RT-PCR. Serum testosterone and LH was detected using commercial ELISA kits. Transcription level of SF-1and P450scc in the testis was eveluated through RT-PCR. Immunofluorescence and confocal analysis was performed to detect cells with co-localized expression of GFP and Leydig cell marker P450scc.Results:Irradiation reduce weight of testis significantly (p<0.05). Compared with that of sham control, serum testosterone in irradiation group animals reduced, though this difference was not significant (p>0.05). Serum LH was increased by irradiation. BMSCs transplantation, expecially with high dose icariin, appeared to attenuated these damages.Immonofluorescence and confocal analysis revealed GFP-positive donor derived cells were located at the in the interstitium, all of which expressed P450scc in the BMSCs transplanted group as well as two icariin groups.RT-PCR confirmed the increased expression of P450scc in these groups compared with irradiated model groups. At the same time, SF-1mRNA in testis with high dose icariin elevated significantly.ConclusionChinese herbal monomer icariin, a major flavonoid isolated from Herba epimedii, has the ability to enhance transdifferentiation of bone marrow stem cells into testicul-ar Leydig-like cells, this efficiency was found to be related to the enhanced exciteme-nt of SF-1expression.