Objective To observe the regulatory mechanism of substance P on the tension of human umbilical vein and the calcium ion of endothelial cells. Methods The tension was recorded by conventional physiological recording methods,and confocal laser scanning microscopy and patch\|clamp technique were used to observe the concentration of intracytosolic free calcium ion and the opening probability of membrane calcium ion channel of cultured human umbilical vein endothelial cells. Results Substance P induced the endothelial\|dependent relaxing of human umbilical vein and the increasing of the concentration of intracytosolic free calcium ion and of the opening probability of calcium ion channel in the cultured human umbilical vein endothelial cells.Conclusion\ Substance P could be function as the activator of the releasing of the intracellular stored calcium ion,and the inflowing of calcium ion from the outside cell bodies to relax human umbilical vein.

Objective To investigate the expression characteristics of the phosphatase and tensin homolog deleted on the chromosome 10 (PTEN) in the central nervous system(CNS) of adult rats. Methods Immunohistochemical staining was used to demonstrate the PTEN-like positive immunohistochemical product. Results Expression of PTEN was extensive in the CNS. The distribution of positive cells was strong, moderate or weak and its subcellular localization included the cytoplasm, nuclei and processes. Conclusion The present results identify the expression of PTEN protein in the CNS of adult rats.

AIM: To investigate the effect of sodium nitrite (SN) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) and its underlying mechanism in mice. METHODS: All male Institute of Cancer Research (ICR) mice were randomly divided into five groups: Control group;LPS group;and SN 4.8 nmol/L, SN 48 nmol/L, SN 480 nmol/L (ip) groups. Lung wet weight/dry weight (W/D) ratio and permeability were detected. Neutrophil infiltration in bronchoalveolar lavage fluid (BALF) was measured by cel1 counting and morphological changes in lung tissues were assayed by hematoxylin-eosin staining. The 1evels of interleukin-10 (IL-10) and tumor necrosis factor-α (TNF-α) in lung were detected. Nitric oxide (NO) level and nitric oxide synthase (NOS) activity in lung were measured according to the specification. RESULTS: Compared to lung in LPS-induced ALI mice, at doses of 4.8 nmol/L and 48 nmol/L, not 480 nmol/L, SN markedly decreased the lung W/D ratio, total leukocyte number and neutrophil percentage in the BALF, lung permeability, and TNF-α/IL-10 ratio, in lung. SN at dose of 480 nmol/L markedly increased the lung NO level compared to control group. In addition, SN decreased the total NOS and inducible NOS (iNOS) activities compared to LPS-induced ALI mice. CONCLUSION: These results indicate that the protective effect of SN against LPS-induced ALI in mice is associated with the low dose SN-induced NO, as well as the subsequent decrease in iNOS activity and TNF-α/IL-10 ratio.

BACKGROUND: Neural axon regeneration is one of the difficulties that must be overcome in treatment of injury of central nerve system. Significant therapeutic effects have been obtained in transplantation of neural stem cells (NSCs), embryonic stem cells (ESCs) and Schwann cells. But the bottleneck situation of insufficiency of cell provider has limited the development on it.OBJECTIVE: To observe directional-differentiation of retinoic-acid induced ESCs so as to find optimal condition for neuronal differentiation.DESIGN: Non-randomized controlled experiment was designed.SETTING: Teaching-Research Room of Histology and Embryology, Department of Basic Medicines. Third Military Medical University of Chinese PLAMATERIALS: The experiment was performed in Staff Room of Histology and Embryology, Third Military Medical University of Chinese PLA from January to May 2000. Eighteen Kunming mice in disoestrus were employed, of which. 12 mice were female and 6 mice male. They were placed in same cage at ratio of 2:1 for mating. The date of pregnancy was recorded. MESPU35 ESC line was prepared.METHODS: Removed head. internal organs and four limbs, feeder-layer Feeder-layer adherent culture was used to proliferate MESPU35 ESCs.Classic 4-/4+ method [The embryoid body (EB) grew naturally for 4 days,without retinoic acid added. In the coming 4 days, retinoic acid was added to induce neural EB of high proportion] was applied to induce the directional differentiation of the nerve. EB was cultured with serum of different concentrations. Phase contrast microscope was used to observe nerve-like EB in serum of different concentrations and to count numbers. ②Immunocytochemical technique was used to observe cellular morphological charac ters at various differentiating phase spots (5th. 9th, 14th days) and with retinoic acid at various concentrations. Flow cytometer (FCM) was used to count the proportion of differentiated neurons.MAIN OUTCOME MEASURES: ①Estimated measurement of the length of process and cell body during formation of neural EB after retinoic-acid induced differentiation of MESPU35 ESCs. ② Observation of cell morphology with immunocytochemical staining and proportion of differentiated cells assayed with FCM.RESULTS: ①It was discovered with phase contrast microscope that serum of different concentrations affect neural directional differentiation after EB formation to certain extent. Excessively high and low concentrations of serum reduced the proportion of neural differentiation of EB. The differentiating proportion is high in serum with 5% concentration. ② It is observed with immunocytochemical technique that the proportions of NF200 positive cell and glial fibrillary acidicprotein (GFAP) positive cell in differentiation of MESPU35 ESCs induced by retinoic acid were increased with phase spots in differentiation and increased concentration of retinoic acid. NF200 positive cell is transformed as multipolar neurons from absence of process in morphology. The processes of GFAP positive cell became longer and linked among each other as reticular pattern finally. ③ It was assayed with FCM that the proportion changes of GFAP positive cell and NF200 positive cell manufactured in differentiation were similar to immunocytochemical one.CONCLUSION: Retinoic acid in combination with proper concentration of serum and differentiating phase spots can induce neural-differentiation of MESPU35 ESC at high proportion and its differentiating regulation is in the patterns of concentration dependence and time dependence.

Objective To observe DAT1 expression in neural stem cells (NSCs) and the effects of DAT1 on the differentiation of NSCs. Methods Eukaryotic expression vectors pEGFP-C1-DAT1 and pcDNA4/hisA-DAT1 were constructed and transfected into NSCs of rat cerebellum by lipofectamine2000. The transfected NSCs were observed by immunohistochemistry under fluorescent microscope. Results The overexpression of DAT1 could increase the number of Mash-1 staining cells and promote the NSCs to differentiate into neuron progenitors, and the high levels of DAT1 in NSCs facilitated the differentiation of neurons. The localizations of DAT1 protein in Mash-1 staining cells and NF 200 staining cells changed. This shift may result from the two distinct inducing factors, FBS and nature differentiation, or distinct stages in differentiation of NSCs. Conclusion DAT1 functions in differentiation of NSCs as a multiprotein combined with distinct transcription factors by virtue of different inducer or varied stage of differentiation.

Objective To isolate and identify pancreatic stem cells of Kunming mouse and to observe the differentiation potency of those cells. Methods Pancreas of postnatal Kunming mice were digested by enzymes to isolate pancreatic stem cells. The potency of the cell differentiation was then identified with special marker of cytokeratin 19(CK 19), insulin and glucagons by immunocytochemical staining. Results Few epithelioid cells could be found to survive at the beginning of isolation, but when medium was replaced by that with growth factor, the cells proliferated quickly in fusiform shape and formed colony gradually. The cells were CK 19 immunoreactive positive after transfer of culture. After differentiation induced by high glucose, the cells formed the pancreatic islet like structures. Immunocytochemical staining showed that part of the cells of pancreatic islet like structures were insulin immunoreactive positive and few of the cells were glucagon immunoreactive positive. Conclusion Pancreatic stem cells of Kumming mouse can proliferate when cultured in vitro and have the potency of differentiation into ? and? cells of pancreatic islet.

Objective To observe the effects of fetal bovine serum(FBS) on differentiation of human neural stem cells (NSCs). Methods The effects of FBS with different concentrations on differentiation of human fetal NSCs were observed by cell culture, immunocytochemistry and flow cytometry. Results Human fetal NSCs could be induced to differentiate mainly three types of nerve system cells(neuron, astrocyte and oligodentrocyte). There were 80%～90% astrocytes of differentiated cells from human fetal NSCs with the concentration of 15% FBS induced. Conclusion Concentration dependent FBS in culture medium may have effect on the ratio of neurons to glial cells differentiated from human NSCs in vitro .

Objective　To investigate the mechanism affecting on permeability of vascular endothelial cell by nitric oxide (NO). Methods　Series concentration of sin-1（a donor of NO） were added to ECV 304, a cell line of human umbilical vein endothelium. Cell growth and expression of f-actin, a cytoskeleton protein were observed. Results　Cell growth was inhibited with a dose from 6.25 to 100 μmol/L and was caused to death at the concentration of 50 to 100 μmol/L by sin-1. The expression of f-actin was suppressed obviously after cultured with 100 μmol/L sin-1 for 4 hours. Conclusion　It suggests that anomaly increased NO can increase permeability of blood vessels by suppressing the expression of f-actin, inhibiting cell growth or even resulting in cell death.

Objective To observe the effect of Ginkgolide B of various consistency on the differentiation of neuron stem cells (NSCs).MethodsNSCs were cultured in differentiation medium containing Ginkgolide B of various consistency for 3 and 7 days, the neurites length and cell body area were measured by inverted phase-contrast micrograph, then neurofilament-200 (NF-200), glial fibrillary acidic protein (GFAP), adenomatus polyposis coli (CC-1) expression were detected and counted by fluorescence microscope. The suppressor of cytokine signaling-2 (SOCS2), inhibitor of DNA binding-2 (Id2) were alsoimmunostained. The percentage of positive cells were counted respectively.ResultsThe neurites length and cell body area in Ginkgolide B groups were obviously larger than that in the control group. The percentage of NF, GFAP positive cells in Ginkgolide B groups increased with dosage increasing of Ginkgolide B. Compared with the normal control group, the percentage of SOCS2 positive cells increased significantly ( P<0.01) and the percentage of Id2 positive cells decreased significantly ( P<0.01) in Ginkgolide B groups.ConclusionGinkgolide B can promote NSCs to differentiate into neuron and astrocyte, the percentage of astrocyte is increased with a dosage-dependent relationship with Ginkgolide B.

Cocaine is one of the main addiction drugs in the world. Scientists are always trying to discover why the addiction happens and find the methods to cure the cocaine addiction. The classics mechanism is that the cocaine binds with the dopamine transporter (DAT), then the retake of dopamine was blocked, and this resulted in the sustained excitement of the dopaminergic neuron. Now, it is found that the cocaine influence some systems relate to the gene expression of the "reward" circuit. This influence finally leads to the change of neuron dendritic plastisity in that area. All the changes are sustaining and these may be the foundation of some behavior effects about the cocaine addiction. At present, the main therapy orientation is decrease the contact between the cocaine and the neuron by idiosyncratic antibody, vaccine or enzyme. Here, related mechanism and therapies were mainly reviewed.