Adult ; Cranial Fossa, Middle ; anatomy & histology ; surgery ; Ear, Inner ; anatomy & histology ; surgery ; Facial Nerve ; anatomy & histology ; surgery ; Humans ; Male

Integrative Medicine ; education ; Internal Medicine ; education

Altitude ; Animals ; Arvicolinae ; Gene Expression ; Hypoxia ; Rats ; Tumor Suppressor Protein p53 ; genetics

OBJECTIVETo explore the effect of total flavonoids of Herba Epimedium (FHE) on BMP-2/RunX2/OSX signaling pathway in promoting osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs).

METHODSPassage 3 BMSCs were randomly divided into the control group, the experimental group, and the inhibitor group. BMSCs in the control group were cultured in 0.2% dimethyl sulfoxide + Osteogenuxic Supplement (OS) fluid + DMEM/F12 culture media. BMSCs in the experimental group were intervened by 20 microg/mL FHE. BMSCs in the inhibitor group were intervened by 20 microg/mL FHE and 1 microg/mL NOGGIN recombinant protein. At day 9 alkaline phosphatase (ALP) activity was measured. Calcium nodules were stained by alizarin red staining and the density was observed. The transcription expression of osteogenic differentiation-related proteins (type I collagen, osteocalcin, and osteopontin) and related factors of BMP-2/RunX2/OSX signaling pathway was assayed by RT-PCR.

RESULTSCompared with the control group, ALP activities were enhanced and the density of calcium nodules significantly increased; type I collagen, osteocalcin, and osteopontin expression levels were increased in the experimental group. The expression of osteogenesis-related transcription factor was also increased in the experimental group. Noggin recombinant protein inhibited FHE promoting BMSCs osteogenesis in the inhibitor group. Compared with the experimental group, ALP activity decreased (P < 0.05), the density of calcium nodules was lowered, expression levels of type I collagen, osteocalcin, osteopontin significantly decreased (P < 0.05) in the inhibitor group.

CONCLUSION20 microg/mL FHE promoted osteogenic differentiation process of BMSCs by BMP-2/RunX2/OSX signaling pathway.

Bone Morphogenetic Protein 2 ; metabolism ; Cell Differentiation ; drug effects ; Cells, Cultured ; Collagen Type I ; metabolism ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Epimedium ; chemistry ; Flavonoids ; pharmacology ; Humans ; Mesenchymal Stromal Cells ; cytology ; drug effects ; Osteocalcin ; metabolism ; Osteogenesis ; drug effects ; Osteopontin ; metabolism ; Signal Transduction ; Sp7 Transcription Factor ; Transcription Factors ; metabolism

Child ; Humans ; Leukemia, Lymphocytic, Chronic, B-Cell

Objective To induce mouse embryonic stem(ES)cells to differentiate into insulin-secreting cells by means of a 5-step model system.Methods E14.1 mouse ES cells were cultured in the presence of leukemia inhibitory factor(LIF)for 2 days(step 1),then the cells were cultured in hanging drops to form embryonic bodies(EBs)and the resulting EBs were cultured in suspension for 6 days in the presence of basic fibroblast growth factor bFGF(step 2).Subsequently the EBs were cultured in the medium containing glucagon- like peptide 1(GLP-1),hepatocyte growth factor(HGF),nerve growth factor(NGF)and nicotinamide for 10 days(step 3).After that,the EBs were dissociated into single cells,and the cells were cultured in monolayer in the presence of GLP-1,betacellulin,activin A,bFGF and nicotinamide for 10 days(step 4).Finally,the cells were cultured in low-glucose medium containing nicotinamide for 4 days(step 5).Insulin and some other islet- related genes expressions were investigated using RT-PCR and insulin expression was also investigated by DTZ- staining and immunohistochemistry.The percentage of insulin-secreting cells was evaluated by flowcytometry and insulin concentrations were measured by RIA.Results mRNA expression of insulin became visible at step 3 and more evident at step 5.Additionally,at step 5,mRNAs of glucagon,somatostatin,pancreatic polypeptide(PP), pancreatic duodenal homeobox 1(PDX-1),beta-cell E box transactivator 2(Beta2)and neurogenin 3(Ngn3) were detected.DTZ-staining positive cells and insulin immunohistochemical staining positive cells were observed. The percentage of insulin-positive cells was(24.0?2.5)%(n=6).In the presence of 5.6 mmol/L and 25 mmol/L glucose,insulin concentrations were(0.05?0.01)?g/L and(0.13?0.02)?g/L respectively(n= 6).Conclusion E14.1 mouse ES cells can be induced to differentiate into insulin-secreting cells by the 5-step model system.Insulin-secreting cells can release insulin into culture medium when treated with glucose,and insulin concentrations increase with rising concentration of glucose.

To examine the effects of heterologous expression of ZrGPD1 (encoding glycerol 3-phosphate dehydrogenase ) cloned from osmotolerant yeast Zygosacharomyces rouxii on glycerol production in wild Pichia farinosa,the URA3 gene was amplified from P. farinosa as the homology integrative region. A recombinant plasmid (pUR-ZG) was constructed then transformed into P. farinosa by electroporation. The transformant pfa-gu was obtained by the selectable marker Zeocin TM . Primary results showed that the biomass of pfa-gu was higher than the wild type in the flask and after 72h fermentation the concentration of glycerol of pfa-gu was 37g/L enhanced 30% in comparison with the wild type. It is concluded that heterologous expression of ZrGPD1 is useful for increasing glycerol production and the ability of osmoregulation in P. farinosa.

Objective To determine whether Fudenine is a novel membrane protein. Methods Green fluorescence protein(GFP)was used to localize Fudenine in vivo. GFP, as a control, was targeted to cytoplasm.Epithelial cell, CBRH7919, and non-epithelial cell, L-6TG, were cultured and transiently transfected by using the lipofectamine reagent. After 48 h, intact cells were examined with fluorescence microscope for Fudenine. Results Reporter plasmid pEGFP-N1, as a control , was expressed and localized to cytoplasm. But Fudenine, driven by the cytomegalovirus promoterenhancer contained in the pEGFP-N1 vector, was overexpressed and targeted to cellular membrane. Conclusions Fudenine is a novel membrane protein. It may play the similar role with its homologues AC133 antigen and prominin in human and mouse.respectively. It might be involved in signaling transduction and regulate blood glucose metabolism in vivo.

OBJECTIVETo investigate the changing laws of serum high mobility group box 1 protein (HMGB1) in septic rats and intervention effect of Xuebijing on it.

METHODSLipopolysaccharide (LPS) (5 mg/kg BW) was intravenously injected into the tail vein of healthy male Wistar rats to prepare the sepsis rat model. In Experiment 1: 50 Wistar rats were randomly divided into three groups, i.e., the normal group (A, n=10); the LPS model group (B, n=10), the LPS +Xuebijing treatment group (C, n=30). Rats in the C group were further divided into three subgroups, i.e., 2 h before LPS injection (group C1), 2 h after LPS injection (group C2), and 8 h after LPS injection (group C3), 10 in each group. Blood samples were collected from the caudal vein to detect serum HMGB1 levels by Western blot at 4, 12, 24, 48, and 72 h after LPS injection. Experiment 2: 30 Wistar rats were equally divided into the LPS model group (D) and the LPS + Xuebijing treatment group (E), 15 in each group. They were treated as rats in the B group and the C1 group respectively. Five rats were sacrificed at 12, 24, and 48 h after LPS injection in the two groups. Blood as well as the tissue samples were harvested to measure such indices as ALT, AST, Cr, and BUN, as well as pathological changes of liver, lung, and kidney.

RESULTS(1) Compared with the A group, serum HMGB1 levels were higher at various time points in the B group (P < 0.05). Compared with the B group, serum HMGB1 levels at 12,24,48, and 72 h decreased in the C1, C2, and C3 groups. Besides, the decrease was more obvious at 24 h and 48 h.The decrement in the C3 group was less than that in the C1 and C2 groups (P < 0.05). (2) In the D group, ALT, AST, Cr, and BUN were significantly higher than those in the A group and reached the peak at 24 h (P < 0.05). Compared with the E group, AST, Cr, and BUN at 24 and 48 h, and ALT at each time point decreased significantly in the E group (P < 0.05). (3)The results of pathological section of liver, lung, and kidney showed local congestion and hemorrhage, cell edema/necrosis/degeneration, infiltration of inflammatory cells, damage of characteristic structures and so on; particularly serious lesion occurred at 24 and 48 h in the D group. The microscopic lesion was obviously alleviated in the E group than in the D group at corresponding time points.

CONCLUSIONSThe serum HMGB1 levels increased in septic rats, with late occurrence of peak value and longer duration of the high value. HMGB1 played an important role in excessive inflammatory response and multiple organ dysfunction. Xuebijing could reduce the serum levels of HMGB1, improve biochemical parameters, and attenuate severe inflammatory response of liver, lung, and kidney tissues in septic rats. Besides, the earlier use, the better effect obtained.

Animals ; Disease Models, Animal ; Drugs, Chinese Herbal ; therapeutic use ; HMGB1 Protein ; blood ; Male ; Rats ; Rats, Wistar ; Sepsis ; blood ; drug therapy

AIM AND METHODSTo study the roles of carbon monoxide on hypoxic pulmonary vasoconstriction (HPV) by investigating the effects of exogenous carbon monoxide and heme oxygenase inhibitor ZnPPIX on hypoxic vasoconstriction reaction of isolated rat pulmonary arterial rings (PAR).

RESULTSHypoxia caused constriction in PAR preconstricted by PE. Both ZnPPIX and carbon monoxide inhibited hypoxic pulmonary constriction significantly by increasing the cGMP level after hypoxia.

CONCLUSIONZnPPIX and exogenous carbon monoxide can inhibit HPV. The reduction of cGMP induced by the decreased of CO may be one of reasons of HPV.

Animals ; Carbon Monoxide ; physiology ; Hypoxia ; In Vitro Techniques ; Male ; Pulmonary Artery ; physiology ; Rats ; Rats, Wistar ; Vasoconstriction ; physiology