Objective To promote the differentiation of Flk1+CD31-CD34-cells isolated from adult adipose tissues into pancreatic islet endocrine cells in vitro.Methods Flk1+CD31-CD34-cells were first cultured and plated in medium supplemented with B27,epidermal growth factor(EGF),and basic fibroblast growth factor(bFGF).Next,the culture medium was changed.The glucose concentration in the serum-free medium was increased.At the same time,betacellulin and nicotinamide were added.Reverse transcription polymerase chain reaction(RT-PCR) was used to detect the expression of nestin,ngn3,insulin promoter factor-1(IPF-1),insulin,and glucagon before and after differentiation induction;immunofluorescent staining for nestin,insulin and glucagon and radioimmunoassay(RIA) for insulin.Results Initially,a nestin positive precursor cell population was found,then small round cells increased in number after 6 days.Later on,they were differentiated into islet-like clusters.The induced cells resulted in the formation of clusters which exhibited higher insulin secretion and other pancreatic endocrine hormones.RT-PCR detected an enhanced expression of pancreatic genes in the differentiated cells.Immunofluorescence revealed a high percentage of insulin-expressing cells in the clusters.Furthermore,the intra-cellular insulin content was detected by RIA after the induction culture.Conclusion These cells represent a previously unidentified adult intrinsic pancreatic precursor population and are a promising candidate for cell-based therapeutic strategies.

Animals ; Dentistry ; Swine ; Swine, Miniature

Objective To evaluate ketamine-induced cerebral protection in mice with traumatic brain injury (TBI) by magnetic resonance imaging (MRI).Methods Thirty-two pathogen-free healthy male C57BL/6 mice,aged 8 weeks,weighing 26-30 g,were divided into 4 groups using a random number table:control group (group C,n=7),ketanine group (group K,n=7),TBI group (n=9) and TBI plus ketamine group (group TBI+K,n =9).TBI was produced with a pneumatically driven controlled cortical impact device.Ketamine 150 mg/kg was intraperitoneally injected at l h after operation in TBI+K and K groups,while the equal volume of normal saline was given instead in TBI and C groups.Open field test was conducted at 24 h,72 h and 7 days after operation.The animals in TBI and TBI+K groups were scanned by T1-weighted MRI at 6,24 and 72 h after operation,the animals in C and K groups were scanned by MRI at 24 h after operation,and the development of cerebral edema was observed.Results MRI scan showed no cerebral edema in C and K groups,and different degrees of cerebral edema were found in TBI and TBI+K groups.Compared with group C,the locomotor distance was significantly shortened at 24 and 72 h after operation in group TBI (P＜0.05).Compared with group TBI,the size of cerebral edema was significantly decreased,and the locomotor distance was prolonged at 24 and 72 h after operation in group TBI+K (P＜0.05 or 0.01).Conclusion MRI method further clarifies that ketamine can produce cerebral protection to some extent in mice with TBI.

The influences of the calmodulin antagonist chlorpromazine (CPZ), and calcium chanmel blocker nimodipine (NLMO) and their combination on cadmium (Cd) poisoning of mice were studied.A series of biochemical parameters including urinary enzyme activities, blood and urine Cd levels, metallothionein (MT) contents in liver and kidney, hepatic ultrastructure and Ca2+ -Mg2- AT Pase activitv in erythrocyte membrane were determined. Animal models for Cd poisoning were established by peritoneal injection of 1/5 LD50 CdCl2. The experimental groups were protected by administration of CPZ, NIMO and CPZ and NIMO in combination I h before the injection of CdCl2. Five days later, samples were collected for analysis. The data showed that CPZ could protect kidney tissue against Cd-induced damage, as the urinary y-glutamyl-traspeptidase (γ-GT) and N-acetyl-β-D-glucosaminidase (NAG) activities were reduced significantly. There was neither evidence of the protective effect of NIMO on kidney tissue nor an indication of a synergistic effecf of CPZ and NIMO.Both CPZ and NIMO showed a considerable protective effect against the decrease in Ca2+ -Mg2+ AT-Pase activity, and a synergistic action was observed. Cd content in blood was reduced significantly by CPZ or the combination of CPZ and NIMO, but elevated by NIMO. Both CPZ and NIMO considerably increased MT contents in livers and kidneys and ameliorated damaged to the hepatic ultrastructures caused by Cd. The results indicated that these inhibitors could protect mice against the toxic effects of Cd in liver and kidney tissues, while CPZ was more efficient than NIMO. The combination of CPZ and NIMO excrted a synergistic action. The protective action of these two drugs might be relevent to the function of MT.

ObjectiveTocomparethedifferencesoftwostocksofguineapigs,thealbinoguineapigsandpigment guinea pigs , in establishing dyslipidemic model , to evaluate their lipid-lowering action , and to compare their properties for development of hyperlipidemia .Methods Two stocks of the 5-week-old guinea pigs were randomly divided into two groups, normal group (NC) and model group (Model).For the NC group, 12 guinea pigs were fed with normal chew .For the model group , after fed with high-fat diet for four weeks , 24 male guinea pigs were randomly grouped and treated with vehicle (VC group) and pitavastatin (Pit group) calcium, respectively, by gavage as well as received high-fat diet.Before and after modeling and pitavastatin treatment , blood samples were collected and subjected to analysis of plasma TC , TG, HDL-C and LDL-C, respectively .Results In the normal group , the blood lipid levels of albino guinea pigs were more stable than that of the pigmented pigs with the increase of age .After fed with high-fat diet , the plasma lipid levels of TC , TG and LDL-C were significantly increased in the two strains of guinea pigs , while HDL-C showed a decrease to varying degrees .Interestingly , the lipid level in the albino guinea pigs was significantly higher than that of pigment guinea pigs . And also, after drug administration for four weeks , pitavastatin treatment significantly decreased the elevated lipid level of TC, TG and LDL-C in the albino guinea pigs compared with that in the pigment guinea pigs .Conclusions The albino guinea pigs and pigment guinea pigs demonstrate certain differences in establishing dyslipidemic model and evaluating lipid -lowering pharmacodynamics .However , compared with the pigment guinea pigs , the albino guinea pigs have obvious superiority because of easy establishment of hyperlipidemia model and are more sensitive to lipid -lowering drugs .

Background Researches demonstrated that dendritic cells(DCs) are uniformly immature in the central cornea but mature in the peripheral region of cornea.So an important question is which factor impact the maturation of DCs,especially in terms of corneal transplant rejection and the known roles of DCs in the development and persistence of some corneal diseases.Objective This study aimed to examine whether corneal stroma cells (CSCs) inhibit DCs maturation through secreting transforming growth factor beta 2 (TGF-β2) and prostaglandin E2 (PGE2).Methods DCs,T cells and CSCs were isolated and cultured from clean BALB/c and C57BL/6 mice.The level of PGE2 and TGF-β2in CSCs culture supernatant and the fresh RPMI 1640 medium were then analyzed by enzyme linked immunosorbent assay (ELISA).During the DCs maturation stage,the neutralizing TGF-β2 antibody and the EP2 receptor antagonist AH6809 were added in the CSCs culture supernatant respectively.According to the different treatment,cultured cells were assigned to different groups as follows:control group,CSCs culture supernatant group,AH6809 group,TGF-β2 antibody group,AH6809 +TGF-β2 antibody group.Subsequently,the cellular surface markers for DCs,including CD11c,CD80,CD86,and MHC- Ⅱ,were analyzed by flow cytometry.The capability of stimulating the proliferation of T lymphocytes was evaluated by allogeneic mixed lymphocyte reactions,and the function of endocytosis was assessed by fluorescein isothiocyanate-dextran(FITC) uptake.Results The data of ELISA showed a higher concentration of TGF-β2 and PGE2 in murine CSCs culture supernatant than in the fresh RPMI 1640 medium.Compared with the CSCs culture supernatant group,the expression of CD80,CD86,and MHC- Ⅱ was up-regulated ( P ＜ 0.05 ),the expression of dextran was down-regulated ( P ＜ 0.05 ),and the stimulate index was increased( P＜ 0.05 ) in the TGF-β2 antibody group; the expression of CD86,and MHC-Ⅱ was up-regulated (P＜0.05),the expression of dextran was down-regulated ( F =13.740,P =0.006 ),and the stimulate index was increased(P＜0.05) in the AH6809 group;the expression of MHC-Ⅱ was up-regulated and the stimulate index was increased with statistical difference in interaction(P＜0.05 ) in the AH6809+TGF-β2 antibody group.Compared with the control group,the expression of CD80 and CD86,and the stimulate index was still lower(P＜0.05 ).Conclusions TGF-β2 and PGE2 contribute to the inhibitory effects on DCs maturation mediated by murine CSCs in vitro and further have additive effect on the immunosuppression of DCs.

Objective We tried to observe the effects of Chinese herb fumigation and washing combined with electromyographic biofeedback on stroke patients with muscle cramp on limbs.Methods 84 patients who had muscle cramp on limbs were selected and divided into two groups randomly,namely the intervention group and the control group and both groups had 42 stroke patients.The intervention group was treated with routine treatment,Chinese herbal fumigation and washing and electromyographic biofeedback treatment.The control group received only routine treatment and electromyographic biofeedback treatment.We had compared and analyzed the Ashworth Spasticity Scale and the Daily Life Abihty Score (Barthel index) of the two groups at the time of the pre therapy,two weeks after the treatment and at the end of the treatment(after four weeks),respectively.Results There were no significant differences between the two groups in Barthel index,however,there were significant differences in Ashworth Spasticity Scale scores between the two groups four weeks after treatment,t=-3.84,P ＜ 0.05.Conclusions The effect of Chinese herb fumigation and washing combining with the electromyo-graphic biofeedback treatment is better than usage of the electromyographic biofeedback treatment alone on stroke patients with muscle cramp on limbs.

Objective To explore the feasibility of establishment of NOD/SCID-mouse model with multiple myeloma by using plasma cells from myeloma patients.Methods The femurs and tibias were removed from the New Zealand white rabbits;the muscles,periosteum and cartilage tissues were cleared.Then each bone was cut into two pieces gently along its middle.The NOD/SCID mice weighing 25 - 30 g (4 - 6 weeks)were anesthetized by intraperitoneal injection;rabbit bone was inserted into the right side of the mouse back and engraftment of the bones was allowed to take place after 4 weeks.The 5000 000 purified plasma cells which expressed CD38 +/CD45 - were immunofluorescence labeled and then injected slowly into the implanted rabbit bone through the distal end.The mice were observed weekly;the plasma cells growth in mice was screened by the living-imaging system and the tumor from the mice was determined by biopsy.Results The implanted rabbit bone survived after 4 weeks.The tumor in mice was observed 2 weeks after the purified myeloma cells were injected into the rabbit bone,and it reached 100 mm3 after 8 weeks.Results of the living-imaging system showed that the myeloma cells had uptake in the rabbit bone after 2 weeks of injection and this phenomenon was more pronounced after 8 weeks of injection (2.4×10 4 vs .1.5× 10 5 ,P < 0.05 ).The tumor infiltrated with numerous plasma cells and osteoclasts increased upon the biopsy. Conclusion Rabbit bone marrow implanted into NOD/SCID mice can effectively support local injection of plasma cells of multiple myeloma patients,and the NOD/SCID-mouse model of myeloma has been established.This model can be used to study in vivo experiments related to myeloma and clinical therapeutic approaches for this disease.

OBJECTIVETo diagnose and detect the molecular defect in a suspected patient with Prader-Willi syndrome.

METHODSGenetic diagnosis and molecular genetic analysis were performed by using chromosome karyotype analysis, methylation-specific PCR (MS-PCR), and linkage analysis using short tandem repeat (STR).

RESULTSThe karyotype of the patient was 45, XX, der(5), t(5;15)(q35;q13), -15, and the parents were 46, XY and 46, XX, respectively, implying that the unbalanced translocation t(5;15) in the patient was de novo. Furthermore, MS-PCR and STR linkage analysis confirmed that the patient's 15q11-13 deletion was resulted from unbalanced translocation on paternal chromosome 15.

CONCLUSIONGenetic analysis should be applied in suspected patients with Prader-Willi syndrome to confirm the diagnosis. Cytogenetic and molecular techniques would be helpful in clinical diagnosis, genetic counseling and prenatal diagnosis.

Chromosomes, Human, Pair 15 ; genetics ; Chromosomes, Human, Pair 5 ; genetics ; DNA Methylation ; Female ; Genetic Linkage ; Humans ; Infant ; Karyotyping ; Male ; Microsatellite Repeats ; genetics ; Polymerase Chain Reaction ; Prader-Willi Syndrome ; diagnosis ; genetics ; pathology ; physiopathology ; Translocation, Genetic ; genetics

OBJECTIVETo investigate the effects of serum containing Gumibao (Chinese character: see text) on the proliferation and differentiation of osteoblast induced by dexamethasone.

METHODSOsteoblasts were extracted from skulls in newly born (within 24 hours) SD rats, and digested with collagenase. The first passage of cells were used for experiments. Cells were cultured in the medium containing different concentrations of dexamethasone (0, 10(-8), 10(-7), 10(-6), 10(-5) ,10(-4) mol/L). Alkaline phosphatase staining were carried out after 1 week and numbers of mineralized nodes with alizarin red staining were observed after 3 weeks. Accordingly, following the treatment of 10(-5) mol/L dexamethasone for 1 week, cells were cultured in the medium with serum containing Gumibao (Chinese character: see text). One week after Cumibao (Chinese character: see text) treatment, cells were stained with Alkaline phosphatase and collagen I and PCNA were examined by Western-blot. However, the observation of numbers of mineralized nodes with alizarin red stain required one more week.

RESULTSHigh concentration of dexamethasone could inhibit the expression of PCNA, collagen I, alkaline phosphatase and reduce the number of mineralized nodes of osteoblast, while serum containing Gumibao (Chinese character: see text) could reverse the inhibition.

CONCLUSIONHigh concentration of dexamethasone could inhibit the proliferation and differentiation of osteoblastic cells, while serum containing Gumibao (Chinese character: see text) could reverse the inhibition.

Animals ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen Type I ; analysis ; Dexamethasone ; pharmacology ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; pharmacology ; Female ; Osteoblasts ; cytology ; drug effects ; physiology ; Proliferating Cell Nuclear Antigen ; analysis ; Rats ; Rats, Sprague-Dawley