Objective To study the effects of high matrix metalloproteinase 9 (MMP 9 ) on the biological behaviors of fibroblasts,in order to clarify the possible mechanisms in the wound healing of diabetic foot.Methods Establish the cell model of skin fibroblast with high expression of MMP 9 by high glucose (22.0 mmol/L) and high homocysteine (100 μmol/L) co-culture.Control group was incubated with normal glucose (5.5 mmol/L).Realtime PCR,ELISA and gelatin zymography were used to detect the MMP 9 mRNA,protein expression and activity of MMP 9.Flow cytometry,CCK-8,ELISA assay,scratch test and transwell were used to detect cell proliferation,viability,collagen (hydroxyproline) secretion,horizontal migration and vertical migration of cells.Results Data were expressed as (x- ± s).Differences were considered significant if their P value was ＜ 0.05.Results The expression of MMP 9 mRNA,protein levels and the activity of MMP 9 in high MMP 9 group were 6.05 folds,4.12 folds and 1.58 folds higher than those in control group (P ＜ 0.01,respectively).The proportion of S phase cells,proliferation index,cell viability,collagen (hydroxyproline) secretion,6 h horizontal migration rate and the number of vertical migration cells in high MMP9 group decreased by 29.8 ％,18.1％,23.3 ％,68.7 ％,45.0 ％ and 21.4 ％ than those in control group (P ＜ 0.01,respectively ). Conclusions Fibroblast with high expression of MMP 9 have decreased proliferation,activity,collagen secretion and reduced migration,which suggests that MMP 9 may inhibit the biological behaviors of fibroblasts.

Objective To evaluate the effect of angiotensin Ⅱ ( Ang Ⅱ ),angiotensin- (1-7) [ Ang- ( 1-7 ) ],and co-action of Ang Ⅱ and Ang-( 1-7 ) on β cell insulin signaling pathway.Methods Mouse pancreatic β cell line NIT-1 was incubated with( 1 )0,10-7,10-6,10-s,10-4 mol/L concentrations of Ang Ⅱ for 24 h ; ( 2 )0,10-7,10-6,10 -5,10-4 mol/L concentrations of Ang- ( 1-7 ) for 24 h; ( 3 ) co-administration of Ang Ⅱ and Ang- ( 1-7 ) was divided into control,10-5mol/L Ang Ⅱ,10-6mol/L Ang-( 1-7 ),10-5mol/L Ang Ⅱ + 10-6mol/L Ang-( 1-7 ) group.Tyrosine phosphorylation of insulin receptor β subunit(IR-β-Tyr) and serine phophorylation of protein kinase B(Akt-Ser) were detected by Western blot.ResultsInsulin-stimulated IR-β-Tyr and Akt-Ser phosphorylation was significantly decreased in Ang Ⅱ 10-5 and 10-4 mol/L group; no significant changes in insulin-stimulated IR-β-Tyr and Akt-Ser phosphorylation were detected between Ang-( 1-7 ) treatment groups and control; Ang-( 1-7 ) blocked the inhibitory effect of Ang Ⅱ on Akt-Ser phosphorylation,yet exerted no effect on Ang Ⅱ-induced IR-β-Tyr phosphorylation inhibition.Conclusion Ang Ⅱ significantly inhibits insulin signaling pathway in β cell; Ang-( 1-7 ) reverts the inhibitory effect of Ang Ⅱ on insulin-stimulated Akt-Ser phosphorylation in β cell.

Overexpression of survivin may partly protect the NIT-1 cells(mouse insulin-secreting cells) from cytokine-induced apoptosis.In addition, NIT-1 cells transfected with survivin had an slightly improved response of insulin secretion to glucose stimulation.

AIM: To compare the effects of three different cell culture protocols: embryonic body(EB) formation,EB formation-monolayer and monolayer on differentiation of mouse embryonic stem(ES) cells into insulin-secreting cells.METHODS: E14.1 mouse ES cells were treated with GLP-1,betacellulin,activin A,bFGF and nicotinamide by using EB formation,EB formation-monolayer and monolayer culture protocol respectively for 30 days,then insulin expression was examined by RT-PCR,DTZ-staining and immunohistochemistry.The percentage of insulin-secreting cells was evaluated by flow cytometry.RESULTS: DTZ-staining positive cells and insulin immunohistochemical staining positive cells were observed in the differentiated cells for all the three groups.mRNAs of insulin and some other islet-related genes were detected,insulin expression was the strongest in EB formation-monolayer,and the weakest was in monolayer.The percentage of insulin-positive cells of the differentiated cells in the EB formation-monolayer group was higher than that in the EB formation group(P

AIM: To study the effects of insulin on the proliferation and function of osteoblasts and the relationship between insulin post-receptor change in osteoblasts and osteoblastic cell growth.METHODS: The effects of different levels of insulin on osteoblasts were assessed by MTT colorimetry.Osteocalcin in medium was measured by RIM.IGF-1 mRNA expression levels were determined by RT-PCR.The concentrations of free IGF-1 protein in serum-free medium were measured by ELISA.In addition,the protein level and phosphorylated protein of P~(44/42)MAPK were determined by Western blotting analysis.RESULTS: Insulin enhanced the proliferation of osteoblasts,depending on its dose and exposure time.Insulin at concentration of 10~(-7) mol/L showed the strongest effect,and the action attained the plateau phase beyond 96 h.The best concentration that stimulated synthesis of osteocalcin by insulin was 10~(-7) mol/L.When the insulin concentration beyond 10~(-7) mol/L,the osteocalcin concentration was decreased.Exposure time had no effect on insulin-stimulated synthesis of osteocalcin of osteoblastic cells.When the concentration of insulin reaches 10~(-6) mol/L,the IGF-1 mRNA expression stimulated by insulin was also decreased.The concentrations of free IGF-1 protein in insulin-stimulated groups were all higher than that in control group(P0.05).Insulin acute stimulation rapidly induced the activity of tyrosine phosphorylation of P~(44/42)MAPK.The degree of tyrosine phosphorylation of P~(44/42)MAPK was increased step by step along with the increasing doses of insulin from 0 to 10~(-7) mol/L(P

Objective To explore the effect of glucose fluctuation on resistin. Methods The phorbol-12-myristate-13-acetate(PMA)-activated and differentiated U937 cells were exposed to experimental condition for 3 days, three groups of cells were formed, each one receiving the following fresh medium every 6 hours, respectively: (1) continuous 11.1 mmol/L glucose concentration medium (Con group), (2)continuous 22.2 mmol/L glucose concentration medium (CHG group), (3) alternating 11.1 mmol/L glucose concentration and 22. 2 mmol/L glucose concentration medium every 6 hours (IHG group). The supernatants of cell mediam at the last 6 hours were collected to test resistin concentration. Besides, 92 subjects were selected and classified into three groups according to the results of oral glucose tolerance test:normal glucose tolerance group ( NGT group, n =30), impaired glucose tolerance patients (IGT group, n =31) and newly diagnosed type 2 diabetes patients (T2DM group, n =31). Blood glucose and serum resistin levels were measured at 0 h and 1 h during oral glucose tolerance test ( OGTT) to compare the glucose fluctuation (△Glu1-0) and the change of serum resistin level (△lnRes1-0) among the three groups. Results Resistin concentration in the Con , CHG and IHG group was (73.62 ± 5.07)ng/L, (97.78 ±7.00)ng/L and(212.49 ± 28. 81 )ng/L respectively and in IHG group it was higher as compared with the other two groups (P<0.05). △Glu1-0 in NGT, IGT and T2DM group was(2.31 ±2.30)mmol/L,(5.70 ±2.08)mmol/L and (8.41 ±2.63)mmol/L respectively; △Glu1-0 increased gradually in all the three groups (P<0.05). Serum resistin level from 0 h to 1 h in the NGT group was 6.41 (1.52-15.76) μg/L to 6. 96( 1.52-22. 70) μg/L, in the IGT group 5.47( 1.49-24. 09)μg/L to 9. 12( 1.27-21.94)μg/L and in the T2DM group 5.77( 1.11-30.10) μg/L to 9. 27(1.02-48.15)μg/L In the IGT and T2DM group serum resistin level increased from 0 h to 1 h (P<0.05), but no difference was observed in the NGT group (P>0. 05).△lnRes1-0 in these 3 groups was (0.05 ± 0.05) μg/L, (0.25 ± 0.04) μg/L and (0.37 ± 0.03 )μg/L respectively and the change in the T2DM group was significant as compared with that in the NGT group,△lnRes1-0 was positively correlated with △Glu1-0 (r = 0.23, P = 0.02). Conclusion Glucose fluctuation induced monocyte/macrophage to secrete resistin, greater the glucose fluctuation, greater the change of amplitude of serum resistin.

Multiple endocrine neoplasia type 2A (MEN2A) is a hereditary syndrome. Here, two different RET proto-oncogen mutation were identified from family members of two MEN2A pedigrees by genetic screening. One RET mutations were found at codons 1893 and 1895 in exon 11 (1893-1895delCGA) from pedigree 1, which is a novel mutation, the other occurs at codon 634 (Cys634Arg) in exon 11 from pedigree 2. However, the clinical characteristics were similar in the patients of the two pedigrees. All the patients were in middle-age at onset. Most of them were firstly diagnosed with bilateral adrenal pheochromocytoma with different degrees of thyroid abnormalities (elevated serum calcitonin with or without thyroid mass, or had been diagnosed with medullary thyroid carcinoma). Some family members were with elevated serum parathyroid hormone but with no other evidences for hyperparathyroidism.