BACKGROUND:Chondromodulin-Ⅰ is expressed mainly in the cartilage, but it is little expressed in mesenchymal stem cells. Combined with the previous study of our group, we concluded that chondromodulin-Ⅰmaybe play an important role in inducing mesenchymal stem cells into chondrocytes accurately.
OBJECTIVE:To construct an expression plasmid stably carrying chondromodulin-Ⅰ to up-regulate the expression of chondromodulin-Ⅰ in bone marrow mesenchymal stem cells.
METHODS:Specific primers were designed in rat cartilage for chondromodulin-Ⅰ gene, then the pcDNA3.1 (+) plasmid expression vector was digested by enzyme and directional connected gene to construct pcDNA3.1(+)/ChM-Ⅰ expression vector. Bone marrow mesenchymal stem cells were obtained from rats using the method of density gradient centrifugation combined with adherent culture. Recombinant plasmid pcDNA3.1(+)/ChM-Ⅰ was transfected into rat bone marrow mesenchymal stem cells with liposome method, and G418 selection was used for stable screen of transfected cells. Reverse transcription-PCR and western blot were used to detect chondromodulin-Ⅰ expression in celllines.
RESULTS AND CONCLUSION:The positive clones were digested by enzyme and were identified and sequenced. The results showed that the reality length and sequence of chondromodulin-Ⅰ gene were consistent with the theoretical values, and reading frame was correct. Reverse transcription-PCR and western blot results showed that the expressions of chondromodulin-ⅠmRNA and protein were markedly up-regulated in bone marrow mesenchymal stem cells. Recombinant plasmid pcDNA3.1(+)/ChM-I was successful y constructed, and transfected into rat bone marrow mesenchymal stem cells. After G418 selection, expression of chondromodulin-Ⅰ was up-regulated stably in rat bone marrow mesenchymal stem cells.

AlM: To investigate the relationship between the anterior ischemic optic neuropathy ( AlON ) and the carotid artery change using doppler ultrasound.METHODS:Fifty-four cases of AlON patients and 54 cases of healthy control were observed, atherosclerotic spots were detected by the application of color ultrasound.RESULTS:ln AlON group of 54 patients, 38 cases appeared carotid atherosclerosis, accounting for 70%. The number of cases with hard plaque, soft plaque and mixed plaques were 18, 13, and 7 respectively, accounting for 33%, 24% and 13%. ln the control group, 20 cases were detected atherosclerotic change, accounting for 37%. And the number of cases with hard plaque, soft plaque and mixed plaques were 12, 5 and 3 respectively, accounting for 22%, 9%, 6%. Significant stenosis and velocity change were showed in neither AlON group nor control group. Compared with the control group, AlON group had more cases of atherosclerotic plaque, the difference was statistically significant (χ2=12. 836, P=0. 005)CONCLUSlON: The incidence of AlON is correlated with carotid atherosclerosis, and carotid ultrasonography is significantly valuable for AlON etiology and diagnosis.

Objective:To prepare a kind of G418 resistance fibroblast feeder layers.Methods: In order to prepare a stock primary embryonic fibroblast (EMFI) cells, the pregnant Smad3ex8+/- mice containing heterozygous neo gene were killed 14 days after coitus to get the embryo. EMFI cells were tested by PCR to identify their gene types. Only one EMFI cell which contained heterozygous neo gene was chosen and treated with mitomycin C.The mitomycin C-treated EMFI feeders were stored at -70℃. Results: The EMFI feeder layers could support the growth of embryonic stem cell line TC-1 efficiently and could live in G418 resistance medium for 1-2 weeks.

ObjectiveTo evaluate effect intervened by the linden relaxation technique (LRT) in the group of long-term naval shipmen's anxiety on surface ship.MethodsSelected naval surface ship unit 1402 shipmen serve as study subjects.140 anxiety shipmen (70 ≥ SAS score ≥ 50) screen out with Zung'self-rating anxiety scale (SAS).The person's group with anxiety mood,but did not meet the diagnostic criteria for anxiety disorders of DSM-Ⅳ.Anxiety shipmen were random divided into the linden relaxation technique group ( n =70) and No-intervention control group ( n =70).In the LRT intervention group:LRT exercises time course was 1 time/ day,and 30 min/time to persist 10 days.The control group without relaxation training.Before and after interventions,anxiety level separately were evaluated with SAS.ResultsBefore intervention,there were not statistically significant between intervention group and control group in SAS anxiety scores,respectively ( ( 54.00 ± 4.43 ) vs (53.91 ± 4.88 )).After intervention,there were statistically significant between intervention group and control group anxiety scores,respectively was ( (40.57 ± 8.98 ) vs ( 53.13 ± 5.51 ) ).The difference before and after intervention,anxiety scoring in intervention group (13.43 ± 7.82 ) was lower than that of the control group (0.79 ± 3.41 ).After following 2 months,SAS scores in the intervention group was more lower (36.76 ± 8.57 ) than the right finish intervention,there was significant different; but there was not significant different belong to the control group (52.34 ± 5.50).ConclusionLinden relaxation technique improves anxiety mood in the naval servicemen.

In order to investigate the antioxidant properties of the polysaccharides from the brown alga Sargassum fusiforme, the crude polysaccharides from S. fusiforme (SFPS) were extracted in hot water, and the lipid peroxidation inhibition assay exhibited that SFPS possessed a potential antioxidant activity. Hence, two purely polymeric fractions, SFPS-1 and SFPS-2 were isolated by the column of DEAE (2-diethylaminoethanol)-Sepharose Fast Flow, with their molecular weights of 51.4 and 30.3 kDa determined by high performance gel permeation chromatography (HPGPC). They were preliminarily characterized using chemical analysis in combination of infrared (IR) and nuclear magnetic resonance (NMR) spectroscopies and found to contain large amounts of uronic acids and beta-glycosidical linkages. The antioxidant activities of these two SFPS fractions were evaluated using superoxide and hydroxyl radical-scavenging assays. The results show that the antioxidant ability of SFPS-2 was higher than that of SFPS-1, probably correlating with the molecular weight and uronic acid content.
Antioxidants ; chemistry ; Hydrogen-Ion Concentration ; Molecular Weight ; Pilot Projects ; Polysaccharides ; chemistry ; Sargassum ; metabolism

OBJECTIVETo investigate the influence and significance of peroxisome proliferator- activated receptor-gamma (PPAR-gamma) agonist rosiglitazone on the expression of I kappa B kinase-beta(IKK-beta) mRNA and protein induced by LPS in Kupffer cells (KCs) cultured in vitro and to investigate the activity of nuclear factor-kappa B (NF-kappa B) together with the expression of cyclooxygenase-2 (COX-2) in livers of rats with non-alcoholic steatohepatitis (NASH).

METHODS(1) KCs from healthy Wistar rats were isolated and purified with IV collagenase digestion and gradient centrifugalization, and then were incubated in the presence or absence of LPS (1 microg/ml) together with two different concentrations of rosiglitazone (10 nmol/L and 50 nmol/L). (2) Thirty-eight healthy Wistar rats were randomly divided into a normal blank control group (10 rats) fed with a normal diet and a NASH model group (28 rats) fed with a fat-rich diet (10% lard + 2% cholesterol + 5% corn oil). After the NASH model was established successfully and confirmed by pathological examination of the livers of 4 rats, 24 rats that continued with the high fat-rich diet, were divided into three groups (8 rats in each group): a control group fed normal saline (NS), a lower dose rosiglitazone group (1 mg.kg(-1).d(-1)) and a higher dose rosiglitazone group (4 mg.kg(-1).d(-1)) for 12 weeks. The mRNA expression of IKK-beta in KCs and COX-2 in livers were measured using reverse transcription-polymerase chain reaction (RT-PCR). The IKK-beta protein in KCs and the NF-kappa B activity of hepatic tissues were determined respectively by Western blot and electrophoretic mobility shift assay (EMSA). The concentration of tumor necrosis factor alpha (TNF alpha) in the supernatant of KCs cultures and serum of the rats was quantified by enzyme linked immunosorbent assay (ELISA).

RESULTSLPS significantly increased the expression of IKK-beta mRNA and protein in the KCs and the concentration of TNF alpha in the supernatant of the KCs cultures. The expressions of COX-2 mRNA and protein were more obvious in rats with NASH than those in the normal control group, and the binding activity of NF-kB correlated positively with the expression of COX-2 in the livers and the level of serum TNF alpha of model rats as well. Rosiglitazone blocked the expression of IKK-beta mRNA and protein induced by LPS in KCs, and also inhibited NF-kappa B activation and reduced COX-2 expression in the rats.

CONCLUSIONSPPAR-gamma specific agonist rosiglitazone can play an anti-inflammatory role by IKK-beta/I kappa B/NF-kappa B/TNF alpha signal ways, and minimize inflammatory reaction at cellular and molecular levels. This may help to provide a new idea for treating NASH effectively.

Animals ; Cyclooxygenase 2 ; metabolism ; Fatty Liver ; drug therapy ; metabolism ; pathology ; Hypoglycemic Agents ; therapeutic use ; I-kappa B Kinase ; metabolism ; Male ; NF-kappa B ; metabolism ; PPAR gamma ; agonists ; Rats ; Rats, Wistar ; Signal Transduction ; Thiazolidinediones ; therapeutic use ; Tumor Necrosis Factor-alpha ; metabolism

Finding the novel drug from the effective components of traditional Chinese herbal medicine is a hotspot of the modern pharmacological research.Hyperoside (HYP) belongs to flavonoid glycosides,and it has various properties,such as anti-inflammation,anti-spasm,anti-diuretic,antitussive,lowering blood pressure,and lowering cholesterol effects as well as protective effects for the cardiac and cerebral blood vessels.The purpose of this study was to investigate the effects of HYP on inflammatory and apoptotic responses in vascular endothelial cells stimulated by lipopolysaccharide (LPS) and further to identify the possible mechanisms underlying these effects.In our study,human umbilical vein endothelial cells (HUVECs) were stimulated with 1 μg/mL LPS in the presence or absence of HYP (10,20 and 50 μrnol/L).Our results indicated that HYP alone exerted no cytotoxicity on HUVECs,while it had an up-regulatory effect on the viability of HUVECs induced by LPS in a dose-dependent manner;increased mRNA expression of IL-1β,IL-6,TNFα and iNOS induced by LPS was attenuated after treatment with HYP both in a dose-and time-dependent manner;LPS-induced HUVECs apoptosis and cleaved-caspase 8,9,3 were all significantly reduced by HYP.Furthermore,the possible pathway involved in apoptosis and inflammation by HYP was detected,and the results showed that when treated with HYP,LPS-induced mitochondrial membrane instability was significantly inhibited through up-regulation of Bcl-2 and down-regulation of Bax.Furthermore,the expression of TLR4 and the phosphorylation of Iκ Bα and p65 in LPS-treated cells were blocked by HYP.Our results suggested that HYP treatment prevented HUVECs from LPS-induced inflammation and apoptosis responses,which might be mediated by inhibiting TLR4/NFκB pathway.

The transgenic mice that express Cre recombinase in a tissue specific manner is a powerful tool in generating the conditional gene knockout mice. The rat insulin promoter was cloned target the expression of Cre in pancreatic tissue. The Cre gene was modified by adding the nuclear localization signal and the sequence for initiation by eukaryotic ribosomes at 5' terminal of the Cre gene. Cre gene was linked to the intron of human growth factor gene. This construct was introduced into the mouse eggs using microinjection. Seven mice were identified as founders carrying the Cre gene by PCR. The results of RT-PCR showed that the transgenic mouse from one founder could transcribe the foreign gene in pancreas. The Southern blot analysis indicated that the Cre recombinase expressed in pancreas of the transgenic mouse was functional.
Animals ; Blotting, Southern ; Female ; Insulin ; genetics ; Integrases ; genetics ; Mice ; Mice, Transgenic ; Pancreas ; metabolism ; Promoter Regions, Genetic ; RNA, Messenger ; analysis ; Rats ; Reverse Transcriptase Polymerase Chain Reaction ; Viral Proteins ; genetics

Child, Preschool ; China ; Female ; Humans ; Infant ; Male ; Nutritional Status ; Rural Population

OBJECTIVETo study the relationship between Helicobacter pylori (Hp) infection and histopathological features of nodular gastritis (NG) in children.

METHODSA total of 213 children who had undergone gastroscopy due to upper gastrointestinal symptoms were enrolled and were divided into NG and non-NG groups according to endoscopic appearance. The histopathological features of gastric mucosa were evaluated using the updated Sydney System. The rates of Hp infection, moderate to severe inflammation and lymphoid follicles formation of gastric mucosa were compared between the two groups.

RESULTSThirty-eight (17.8%) of the subjects were diagnosed with NG. The NG group had significantly increased rates of Hp infection (86.8% vs 14.3%; P<0.01), moderate to severe inflammation (81.6% vs 15.4%; P<0.01) and lymphoid follicles formation of gastric mucosa (52.6% vs 10.3%; P<0.01) compared with the non-NG group. NG had a high specificity (96.8%) and a positive predictive value (86.8%) for the diagnosis of Hp infection. NG was observed in 33 (56.9%) of 58 Hp-positive children and in 5 (3.2%) of 155 Hp-negative children (P<0.01). Hp-positive children had higher rates of moderate to severe inflammation (86.2% vs 5.2%, P<0.01) and lymphoid follicles formation of gastric mucosa (84.2% vs 14.9% P<0.01) compared with Hp-negative children. There were significant differences in Hp colonization, degree of inflammation and inflammation activity in gastric tissues between the NG and non-NG groups (P<0.01).

CONCLUSIONSNG is a special sign of Hp infection in children, which mostly shows moderate to severe inflammation of gastric mucosa, and can be used as an endoscopic indicator of Hp infection. Hp eradication therapy should be considered in the treatment of NG.

Adolescent ; Child ; Child, Preschool ; Female ; Gastric Mucosa ; pathology ; Gastritis ; pathology ; Helicobacter Infections ; pathology ; Helicobacter pylori ; Humans ; Male