There are plenty of haematopoietic stem cells in umbilical cord blood, which are regarded as important resources for transplantation therapy. There are some arguments on whether or not mesenchymal stem cells(MSCs) exist in umbilical cord blood. Some researchers have such opinions that there exist a number of MSCs in umbilical cord blood as similar with one from bone marrow in many aspects. Others believe that the content of MSCs in umbilical cord blood is too low to expand in vitro. We summarized the data from last five years researches. On umbilical cord blood MSCs during last to help further research and application of this resource of MSCs.

AIM: To explore the possibility of differentiating functional insulin-producing cells from human BM-derived stem cells. METHODS: Mesenchymal stem cells were isolated from human bone marrow. Then these cells were induced with epidermal growth factor, β-mercaptoethanol and high concentration of glucose. The gene expression related to islet β cells was detected by RT-PCR. Insulin in the treated cells was examined by immunocytochemistry. In addition, the levels of insulin secretion and glucose-stimulated insulin release were examined by microparticle enzyme immunoassay. Finally, the induced cells were implanted into the right renal subcapsular space of diabetic mice. Blood glucose levels were monitored 16 d after implantation. The right kidneys of the treated mice were harvested for immunohistochemistry. RESULTS: The key genes related to pancreatic β cells had been confirmed to express by PCR and insulin was detected by immunocytochemistry in differentiated human BM-derived stem cells induced by high glucose, which responded to glucose challenge. Furthermore, implantation of the cells in renal subcapsular space was able to lower the glucose levels in hyperglycemic mice. After 16 days, the implanted cells were determined still to be insulin positive cells by immunohistochemistry. CONCLUSION: These results indicate human BM-derived stem cells are capable of differentiating into functional insulin-producing cells and may represent a pool of cells for the treatment of diabetes.

Objective To discuss the possibility of differentiation of mesenchymal stem cells(MSCs)from umbilical cord blood(UCB)into hepatocyte-like cells in the in vitro culture.Methods MSCs were isolated from UCB,cultured and passaged.The surface markers were examined by flow cytometry.When cells were cultured to the third passage,they were inoculated into a 6-well plate.A two-stage induction method was used:MSCs in the first phase were cultured in DMEM/F12 medium supplemented with dexamethasone(final concentration of 0.5 μmol/L,the same below),hepatocyte growth factor(HGF,10 ng/ml),epidermal growth factor(10 ng/m1)and 1×insulin-transferrin-Se(ITS)for two weeks,then in DMEM/F12 supplemented with 0.5 μmol/L dexamethasone,10 ng/ml HGF,1×ITS,10 ng/ml Oncostatin M for another two weeks.Morphological changes were observed under a microscope.The gene expression correlated with hepatocytes was detected by using RT-PCR.Immunofluorescence staining was used to identify the expression of specific protein related to hepatocytes(AFP,Albumin,CK-18).Ultrastructure was detected under an electron microscope.Results In the cultured MSCs from UCB,CD34/CD45/CD14,CD54,CD49f and HLA-DR were not detected,there was low expression of CD106 and strong expression of CD29,CD44 and CD13.The gene expression of AFP,albumin,CK-18 and TAT was discovered and three kinds of protein AFP,albumin and CK-18 were positively showed in cytoplasm after 4 weeks'induction.The hepatin granules and fatty drops in cytoplasm of cells induced for 4 weeks were found under an electron microscope.Conclusion The MSCs fromUCB can differentiate into hepatocyte-like ceils in the in vitro culture under some conditions.

The experiments of rabbit mydriasis & isolated cat ciliare muscle paralysis of atropine Methobromide (AMB) have demonstrated that : AMB has much larger mydriasis effect. It effects faster than Atropine Sulfate, Homatropine & Tropinexamide in acetylcholin -induced ciliare muscle contraction. The combined-force of AMB on ciliare muscle is less than Atropine & Homatropine, but a little larger than Tropinexamide. There fore AMB is a rapid & short-time mydriasis agent & ciliare muscle paralysis agent.

Objective: To investigate the effect of the dosageof chemotherapeutic agent on tumor chemotherapy linked with biotherapy and provide experimentalevidence for the dose choice of chemical drugs in the combination of chemotherapy and biotherapy.Methods: The high- and low-dosage of MMC was determined by injection of mice with different dosages of MMC. Mice inoculated with H22 hepatic carcinoma cells were treated with different dosages of MMC followed by three different kinds of biotherapy: transfection with plasmid pCH510 in vivo, immunization with Hsp70-tumor antigen peptide complexes and the combination of these two elements. Results: By toxicity test of MMC to mice, it was determined that 100 ?g of MMC was high dosage and 50 ?g was low dosage. The curative effect was significantly improved if chemotherapy was followed by different elements of biotherapy. Better efficacy was obtained when biotherapy elements were used to follow the chemotherapy with high dosage of MMC. In the case of low dosage of MMC, no difference could be observed in curative effect of three different kinds of biotherapy. When high dosage of MMC was used, the curative effect of three different kinds of biotherapy was signiferently different. The best efficacy was obtained if chemotherapy was followed by the combination of two biotherapy elements, transfection with plasmid pCH510 in vivo and immunization with Hsp70-tumor antigen peptide complexes. Conclusions: Using different chemical dosages, the curative effect of chemotherapy linked with biotherapy is different. In the case of high dose, the chemotherapy linking with biotherapy can reach more better efficacy.

Objective: To invistigate the relationship between time and efficacy of the linkage of tumor biotherapy after chemotherapy by studying the influence of chemotherapeutic drugs on immunocytes.Methods: The cytotoxicity of chemotherapeutic drugs to tumor cells, mouse peritoneal macrophages and spleen lymphocytes was observed by cell culture technique. The influence of chemotherapeutic drugs to the metabolism and activation of macrophages and lymphocytes at different time after peritoneal injection of drugs was observed. The mice were inoculated with tumor cells two days after the injection of drugs, and the growth of tumor was measured 14 days after inoculation by anatomizing mice. Results: Chemotherapeutic drugs had cytotoxicity in vitro to different cells, suppressed the function of immunocytes, and decreased the number of immunocytes in vivo. After injection of drugs, the number of immunocytes was the lowest on the third day and recovered to the nomal level on the 10th day. If drugs was injected two days before the inoculation of tumor cells, the growth of tumor became faster than control group. Conclusions: Chemotherapy not only decreases the number of immunocytes but also suppresses the function of immunocytes, and it can promote the growth of tumor after its cytotoxicity disappeared. So it is not good that biotherapy, which depends on immunocyte to kill tumor cells, is used immediately after chemotherapy and it is also not good for using biotherapy with a long interval after chemotherapy . It is good time to use biotherapy when the number of immunocytes is lowest or the recovery just starts.

Objective To optimized the culture conditions of the human umbilical cord blood mesenchymal stem cells.Methods The umbilical cord blood was taken in Guangzhou Hua Qiao Hospital between Oct.2004 to Feb.2005.The isolation method,planting density,the time of the first medium changing,the influence of culture medium on the growth of MSC were analyzed. Mesenchymal stem cells were identified using the surface marker by flow cytometry.Osteoblast was identified by Von Kossa staining and alkaline phosphatase staining,adipocyte was identified by Oil Red O staining. Results When the other conditions were the same,the hespan isolation was better than Ficoll isolation;5?10~6/cm~2 was the best planting density;the best first medium changing was on the seventh day at primary culture. Under optimized conditions,the MSC expressed adhesion molecules CD_ 13 ,CD_ 29 and CD_ 44 ,but not antigens of hematopoietic CD_ 34 ,CD_ 45 ,CD_ 14 and not antigens of endothelia CD_ 106 . Exposure of these cells to osteogenic inductive agents resulted in an increase in expression of alkaline phosphatase and the appearance of hydroxyapatite nodules. Incubation with adipogenic inductive agents resulted in morphological change and staining with Oil Red O. Conclusion Mesenchymal stem cells exist in Cord blood,but slower to establish in culture.Cord blood may prove to be a new source of cells for cellular therapeutics.

BACKGROUND: At present,there are many reports regarding the differentiation of bone marrow-derived mesenchymal stem cells or pancreatic gland stem cells into pancreatic islet β-like cells.But little is about umbilical cord blood-derived mesenchymal stem cells(UCBMSCs)differentiation into pancreatic islet β-like cells in vitro.OBJECTIVE: To investigate whether UCBMSCs can differentiate into pancreatic islet β-like cells in vitro and the optimal inducing condition.METHODS: UCB samples were obtained sterilely from healthy parturients.Nucleated cells were isolated by sedimentation with hydroxyethyl starch and MSCs were obtained by adherent method.Then purified UCBMSCs were induced with epidermal growth factor,β-mercaptoethanol,high glucose,Activin A and hepatocyte growth factor(HGF).Following cell morphology observation,induced cells were identified by insulin immunofluorescence.In addition,insulin secretion and glucose-stimulated insulin release were examined by chemiluminescence immunoassay.RESULTS AND CONCLUSION: After induction,many cells exhibited a round appearance and produced islet-like cell clusters.Immunofluorescence assay showed insulin positive in the treated cells.In addition,chemiluminescence immunoassay demonstrated low expression of insulin and secretion of insulin upon glucose challenge.UCBMSCs can differentiate into pancreatic islet β-like cells in vitro.

BACKGROUND: Reproductive activity of hepatocytes is limited. There are numerous studies concerning stem cells differentiation into hepatocytes, including embryonic stem cells, bone marrow cells, pancreas stem cells, neural stem cells, various sources of mesenchymal stem cells.OBJECTIVE: To explore the possibility of induction of mesenchymal stem cells (MSCs) from umbilical cord blood (UCB) into hepatocyte-like cells in vitro culture.DESIGN, TIME AND SETTING: A cytological in vitro study was performed at the Institute of Hematology, Medical College of Jinan University from October 2005 to April 2006.MATERIALS: Fetus cord blood was obtained from spontaneous delivery and caesarean delivered healthy pregnant women at the Department of Obstetrics and Gynecology, First Affiliated Hospital of Jinan University. The parturients signed the informed consents.METHODS: UCB-MSCs were incubated in vitro, and digested in trypsin-EDTA. The third passage of cells at 5 × 10~4 cells/cm~2 wereinoculated. Original medium was removed 48 hours later. Cells were washed in phosphate-buffered saline. In the first phase, cells were incubated in F12 medium supplemented with dexamethasone, hepatocyte growth factor, epidermal growth factor and 1 ×its for 2 weeks. In the second phase, cells were incubated in F12 medium containing dexamethasone, hepatocyte growth factor,oncostatin M and 1 × ITS for two weeks.MAIN OUTCOME MEASURES: The following parameters were measured: expression of surface marker of UCB-MSCs using flow cytometry, expression of related gene and protein of hepatocytes following induction respectively using RT-PCR and immunofluorescence staining.RESULTS: No CD34/CD45/CD14 of hematopoietic markers were detected, either no the CD54, CD49f, HLA-DR were found. The low expression of CD106 and high expression of CD29, CD44, CD13 were found. The gene expression of a-fetoprotein, albumin,CK-18 and TAT were discovered, and three kind of protein a-fetoprotein, albumin, CK-18 were positively observed in cytoplasm after 4 weeks of induction using immunofluorescence staining.CONCLUSION: UCB-MSCs are able to differentiate into hepatocyte-like cells in vitro culture following combination with many growth differentiation factors, such as dexamethasone, hepatocyte growth factor, epidermal growth factor, tumorigenesis M and ITS.

Objective:To sutdy the effect of 2 chloroadenosine(2 ClA),which is specifically cytotoxic to macrophages,on MTT assay in activation test and cytotoxicity test of lymphocyte.Methods:Using cell culture technique,mouse splenic lymphocytes and peritoneal macrophages were cultured.Lymphocyte activation and specific cytotoxicity to tumor cells and toxicity of 2 ClA to macrophages were measured by MTT assay in the presence or absence of 2 ClA.Results:2 ClA had a strong cytotoxic effect on macrophages.When the activation test and cytotoxicity test of lymphocyte were measured by MTT assay,the optical density values of 2 ClA group was lower than that of control group,and statistic analysis showed P