Objective To investigate the fertilization ability of spermatozoa derived from neonatal mouse testes grafted into immunodeficient mice by intracytoplasmic sperm injection(ICSI).Methods Neonatal Kun-ming mouse testes containing primitive spermatogenic cells as the only germ cells were grafted under the back skin of castrated BALB/c-nu/nu mice.Grafts were taken out after 8 weeks.The initiation and restoration of spermatogenesis in grafts was documented by histological analyses.The ultrastructure of spermatogenic cells in grafts was observed using transmission electron microscopy.The activity of acrosomal enzyme of spermatozoa was analyzed with gelatin substrate test.The fertilization ability of spermatozoa was determined by using intracytoplasmic sperm injection.Results Histological analysis showed complete restoration of spermatogenesis in the grafts after 8 weeks.1?10~(3) spermatozoa with normal morphological appearance in 1mg suspended graft tissue were counted.Spermatogonia and spermatocytes with normal ultrastructure were observed.Gelatin substrate test positive spermatozoa were found implicating the activity of acrosomal enzyme of the spermatozoa.Cleavages were observed both in the ICSI group and the ICSI plus electric stimulation group.Conclusion Spermatozoa with normal morphology and fertilization ability can be developed by allografting neonatal mouse testes under the back skin of immunodeficient mice.

Objective Stromal and glandular cells of human endometrium were successfully isolated and cultured in this experiment. Methods These two types of cells were observed by light microscopy and verified by immunocytochemical staining. Results The result showed that some of stromal cells became adherent after.0 5h of culture.Two shapes of stromal cells were found.One was spindle and the another was polygonal.Immunocytochemical staining method showed that both shapes of stromal cells were positive for vimentin,with 95% of positive rate,and negative for cytokeratin 1,indicating that these cells were endometrial stromal cells.Most of glandular cells became adherent after 24h of culture and formed tightly packed whorls after 4d.Individual cells were polygonal and had a large and round nucleus.Immunocytochemical staining method showed that glandular epithelial cells were positive for cytokeratin 1,with 90% of positive rate,suggesting their nature of epithelial cells. Conclusion Stromal and glandular cells of human endometrium were successfully isolated by using two series of filter and cultured in this experiment. [

Objective To investigate the developmental feasibility of early human fetal testes (<3 months) using xenografting technique and to acquire an accessible donor derivation that is essential for studying human germ cell development. Methods Nine testes from 10-13 weeks aborted fetus were grafted under the back skin of 6 castrated nude mice. Grafts were collected at different time point according to the growth of the donor tissues and the health condition of the recipients. Morphological and histological analyses were performed for the observation of the development of grafted immature testicular tissues. Results The mass of grafts was increased from about 5-7mg to 84.1mg (the biggest). Six of 9 testes were to be in developing. Histological observations showed a significant expansion of seminiferous tubules from (44.26±3.14)μm to (77.69±7.47)μm. Cells dispersedly distributed in seminiferous cords at the time of grafting migrated towards the basal part of seminiferous epithelium. Some germ cells with spermatogonium-like characteristics located on the basement membrane. Sertoli cells were in stages from immature into matured with abundant cytoplasm which were orderly arranged around spermatogonia forming a niche-like structure. Conclusion Testes from early aborted human fetus grafted under the back skin of castrated nude mice showed further development and therefore could be used as an easier accessible donor tissues for the investigation of human spermatogenetic mechanism.

Objective:To explore the effect of the microenvironment induced by damaged mouse hepatic cells on the conversion of human umbilical cord blood-derived cells into hepatocyte-like cells. Methods: A hepatic injury-like microenvironment was mimicked using carbon tetrachloride damaged mouse hepatic cells, where mononuclear cells (MNC) from human umbilical cord blood were cultured in a compartment separated by trans-well membrane. Histochemical staining, reversed transcription-polymerase chain reaction (RT-PCR) and gene sequencing were performed for the information on the conversion of human umbilical cord blood MNC. Results: A number of PAS positive stained cells in MNC co-cultured with damaged mouse hepatic cells were observed after 72 h. Cells expressing mature hepatocyte markers, human albumin (hALB) and human GATA-4 (hGATA-4) mRNA were detected by RT-PCR, which was further confirmed with sequencing. Relevant control groups, MNC co-cultured with normal mouse hepatic cells and MNC cultured alone remained negative. Conclusion: The culture system using damaged mouse hepatic cells as stimulator could be a potential in vitro system for the conversion of human umbilical cord blood-derived cells into hepatocyte-like cells.

OBJECTIVETo investigate the development of xenografted primitive human germ cells by using fetal testicular tissues as donor tissues and an immunodeficient mouse as the recipient.

METHODSTesticular tissue fragments of a 26-week fetus were grafted under the back skin of a castrated immunodeficient mouse. Grafts were taken out after 135 days and processed for morphological and histological analyses.

RESULTSThe mass of grafts grew from about 1 mm in diameter and 5 mg in wet weight to about 3 mm and more than 20 mg 135 days after grafting. Histological observations showed a significant expansion of seminiferous tubules after grafting (80 +/- 25 microm in diameter) in comparison with seminiferous cords at the time of grafting (60 +/- 15 microm in diameter). The seminiferous cords developed into seminiferous tubules with the epithelial border and lumen. After 135 days of grafting, most of the dispersedly distributed primitive Sertoli cells and germ cells migrated to the basal part of seminiferous epithelium, located on the basement membrane and few of germ cells differentiated into spermatogonia.

CONCLUSIONHuman fetal testicular tissues could survive and continuously develop after being xenograft into castrated immunodeficient mice.

Animals ; Fetal Tissue Transplantation ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Spermatids ; growth & development ; Testis ; cytology ; transplantation ; Transplantation, Heterologous

OBJECTIVE To investigate the effects of Forsythia suspensa ethanol extract on the proliferation， migration and invasion of lung cancer cells NCI-H226. METHODS As research objects， lung cancer cells NCI-H226 were divided into control group， F. suspensa ethanol extract low-， medium- and high-concentration groups （5， 10， 20 mg/mL）， activator group [10 mg/mL F. suspensa ethanol extract+0.5 μmol/L nuclear factor kappa B （NF-κB） signaling pathway activator PMA]， inhibitor group （10 mg/mL F. suspensa ethanol extract+10 μmol/L NF-κB signaling pathway inhibitor BAY 11-7082） and positive control group （20 μg/mL cisplatin）. Except for the control group of cells without intervention， all other groups of cells were cultured with corresponding drugs for 24 hours； the proliferation， migration and invasion of cells were all detected， and the proliferation rate， migration rate， and the number of invading cells were also calculated； protein expressions of NF-κB p65， NF-κB inhibitory protein α （IκBα）， phosphorylated NF-κB p65 （p-NF-κB p65） and phosphorylated IκBα （p-IκBα） were determined. RESULTS Compared with control group， the proliferation rate， migration rate， and the number of invading cells as well as the protein expressions of p- IκBα and p-NF-κB p65 were decreased significantly in F. suspensa ethanol extract groups and positive control group （P＜0.05）. Compared with F. suspensa ethanol extract medium-concentration group， the proliferation rate， migration rate， and the number of invading cells as well as above protein expressions were all decreased significantly in inhibitor group （P＜0.05）， while those of activator group were increased significantly （P＜0.05）. CONCLUSIONS F. suspensa ethanol extract can inhibit the proliferation， migration and invasion of lung cancer cells NCI-H226， and the mechanism of which may be related to the inhibition of NF-κB signaling pathway.