Objective To study the effects of EP4 and EP2 antagonists on the differentiation of Treg/ Th17 cells and disease progression in mice of collagen-induced arthritis (CIA) model.Methods DBA/1 mice wereimmunized subcutaneously twice at the root of the tail with type Ⅱ collagen emulsified in Freund's complete adjuvant.EP2 and EP4 antagonist therapies were intraperitoneally administrated for 14 consecutive days after the second immunization.Clinical signs,histological manifestation,serum interleukin (IL)-17 and quantity of CD4+CD25+Foxp3+ Treg cells were determined.ANOVA and t-test were used for statistical analysis.Results Clinical signs of the disease appeared on day 27 and peaked on day 35 after the first immunization.The quantity of CD4+CD25+Foxp3+ Treg cells in spleens [(1.67±0.15)％] and draining inguinal lymph nodes [(3.30±0.36)％] isolated from CIA mice were significantly lower than those of normal DBA/1 mice [(2.77±0.45)％ and (4.73 ±0.45)％ respectively,P＜0.05].Serum IL-17 level of CIA mice [(27±7) pg/ml] was significantly higher than that of normal DBA/1 mice [(14±4) pg/ml,P＜0.05].Intra-peritoneal injection of EP4 but not EP2 antagonist to CIA mice decreased paw edema and swelling,and alleviated the histological manifestations (1.8±1.0 vs 3.5±0.6,P＜0.05) on day 35 after the first immunization.The percentages of CD4+CD25+Foxp3+ Treg cells in both inguinal lymph nodes [(4.20±0.32)％] and spleens [(2.63±0.40)％] were significantly higher in EP4 antagonist-treated but not EP2 antagonist-treated CIA mice compared with CIA mice group [(3.30±0.36)％ and (1.67±0.15)％ respectively,P＜0.05].The level of serum IL-17 was significantly lower in EP4 antagonist-treated [(15±7) pg/ml] but not EP2 antagonist-treated CIA mice compared with CIA mice group [(27±7) pg/ml,P＜0.05].Conclusion EP4 antagonist therapy alleviates clinical symptoms of CIA,improves the histological manifestations,decreases the serum IL-17 level and increases the percentages of CD4+CD25+Foxp3+ Treg cells in both spleens and draining inguinal lymph nodes,so targeting EP4 receptor may be a new possible therapeutic possibility in the prevention and treatment of rheumatoid arthritis.

Objective Cancer, a disease induced by abnormally regulated cell growth and apoptosis, is imposing a global threat to human health.This study was to explore the effects of Chinese herbal extracts ( CHE) in inducing the apoptosis and inhibiting the proliferation of human lung cancer cells. Methods Human lung cancer A549 cells were divided into a negative control, a high-dose CHE (680 ng/mL), a medium-dose CHE (340 ng/mL), and a low-dose CHE (170 ng/mL) group.The inhibitory effect of CHE on the proliferation of the lung cancer cells was detected by CCK8 and LDH assays, the apoptosis of the cells was assessed by fluorescence microscopy and flow cytometry, and the expressions of hTERT mRNA, cleaved caspase-3 and cleaved PARP were deter-mined by RT-PCR and Western blot. Results CHE inhibited the proliferation of the A549 cells with an IC50 value of 510 ng/mL. Treatment with high-dose CHE for 48 hours significantly suppressed the proliferation of the cells, induced the release of LDH, and promo-ted the apoptosis of the cells by 72.3%.RT-PCR and Western blot showed that 24-hour treatment with medium-dose CHE reduced the expression of hTERT mRNA by 4 times that of the negative control and up-regulated the expressions of cleaved caspase-3 and cleaved
PARP. Conclusion Chinese herbal extracts can induce cell apoptosis by decreasing the expression of hTERT mRNA and increasing those of the cleaved caspase 3 and cleaved PARP proteins.

Objective To study the receptors signaling of prostaglandin E2 on the differentiation of regulatory T (Treg) cells and Th17 cells.Methods The expression of prostaglandin E2 receptors (EP1/EP2/EP3/EP4) on the MACS-purified CD4+CD62L+ T (Th0) cells was analyzed by flow cytometry and reverse transcription polymerase chain reaction (RT-PCR).The quantity of CD25+Foxp3+ cells was examined by flow cytometry,the expression of FoxP3 mRNA and RORγt mRNA were detected using real-time RT-PCR,the level of IL-17 in the culture supernatants was detected by enzyme-linked immunosorbent assay (ELISA).ANOVA,LSD-t,Dunnett T3 were used for statistical analysis.Results EP1,EP2,EP3,EP4 were expressed on Th0 cells at different levels,and EP2 [(89.7±9.1)％] had the strongest expression.PGE2 [(3.0± 2.2) ％],EP2 agonist [(4.5± 1.0) ％] and EP4 agonist [(8.8 ±2.5) ％] decreased the quantity of CD25 +Foxp3 + cells compared with the control group [(28.6±6.8)％] (t=7.156,P=0.021; t=6.958,P=0.032; t=5.359,P=0.044).PGE2(0.210±0.020),EP1 agonist (0.833±0.045),EP2 agonist (0.227±0.025) and EP4 agonist (0.450±0.060) decreased the expression of Foxp3 mRNA compared with the control group (1.000) (t=23.817,t=5.026,t=23.313,t=16.581; all P=0.000).PGE2 [(22±6)pg/ml],EP2 agonist [(24±5)pg/ml]and EP4 agonist [(207±19) pg/ml] decreased the secretion of IL-17 compared with the control group [(678±87) pg/ml] (t=14.925,P=0.004; t=14.873,P=0.004; t=10.480,P=0.008).PGE2 (0.141±0.027),EP1 agonist (0.869±0.033),EP2 agonist (0.176±0.029) and EP4 agonist(0.371±0.042) decreased the expression of RORγt mRNA compared with the control group (1.000) (t=34.046,t=5.184,t=32.673,t=24.962,all P=0.000).Conclusion EP1,EP2,EP3,EP4 receptors are expressed on CD4+CD62L+ T (Th0) cells at different levels.Prostaglandin E2 inhibits the differentiation of Treg cells and Th17 cells via the EP2 and EP4 receptors signaling.

Aurora kinases are a family of serine/threoning kinases whose structures and functions are highly conserved in different model organisms. They play significant roles in many events during cell mitosis, such as centrosome maturation and separation, spindle assembly and maintenance, chromosome segregation, cytokinesis. Overexpression of aurora kinases has been observed in some tumor ceils and aberrations in aurora kinases have been proved to be strongly associated with tumorigenesis. Up to now, some small molecule aurora kinase inhibitors with anti-tumor activity have been developed. Some of those with promising pre-clinical results have reached clinical trial. This review describes the recent progress of aurora kinases and their small molecule inhibitors.

Objective To probe into the pathogenesis of gestational diabetes mellitus (GDM) by studying expression of TLR4 acetylation in peripheral blood mononuclear cells of gravida with GDM as well as its role in TLR4 inflammatory pathway. Methods 30 normal gravidas and 30 gravidas with GDM were enrolled in the study, 15 mL peripheral blood from every participant was collected for extraction of mononuclear cells via density gradient centrifugation and culturing in vitro by LPS. The expression of TLR4 acetylation in the mononuclear cells was detected using immunoprecipitation and western blot and the levels of inflammatory cytokines, TNF-α, IL-1 and IL-10 in the supernatant were detected by ELISA. Results TLR4 acetylation appeared positive in GDM group, and negative in the control group. After the LPS intervention, the degree of TLR4 acetylation was enhanced significantly and the latter was significantly higher than those of the GDM group and the normal + Lps group (P < 0.05). The differences in the levels of inflammatory cytokines, TNF-α, IL-1, IL-10, were statistically significant (P < 0.05). Conclusion TLR4 acetylation exists in mononuclear cells of gravida with GDM, and mediates the release of inflammatory cytokines by influencing the activation of TLR4 inflammatory pathobenesis, which contributes to the promoted antiinflammatory - proinflammatory imbalance in gravidas. In this way, it is involved in the growth of GDM.

Aging ; physiology ; Animals ; Female ; Male ; Mitochondria, Heart ; enzymology ; Mitochondrial Membrane Transport Proteins ; physiology ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism ; Ventricular Function, Left ; physiology

OBJECTIVETo obtain very end full-length cDNA of hepatitis C virus (HCV) 5' untranslated region (5' UTR), and analyse its primary and secondary structure.

METHODSBy reverse transcription-nested polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP), a patient infected with genotype 2a HCV was found. Total RNA isolated from the serum as template, the cDNA of 5' noncoding region was amplified using rapid amplification of cDNA ends methods (RACE), the fragments were recombined by A-T clone strategy, the recombinants were confirmed by RFLP and PCR then sequenced. Secondary structures were analysed by RNA draw.

RESULTSVery end full-length cDNA of 2a genotype HCV 5' UTR was obtained by RACE. In five clones obtained, three contained full-length 5' UTR cDNA, and A21G, G170A, T222C, T247C, C339T substitutions were found compared with HC-J6. he homologies with HCV-1,HC-J6,HC-C2, HC-J8 were 93.6%-94.4%, 92.1%-93.0%, 98.8%-99.7%, 96.2%-96.5%, respectively; however, the substitutions did not alter the secondary structure. Two out of five clones were deleted to have 53 and 144 bases at 5' terminus of HCV 5' UTR, respectively.

CONCLUSIONSRACE is rapid and effective, works well to obtain very end of virus genome. With that, Authors obtained full-length cDNA of genotype 2a of HCV 5' UTR. There are genes deleted at 5' terminus circulated in hepatitis C patients.

5' Untranslated Regions ; genetics ; Base Sequence ; DNA, Complementary ; genetics ; DNA, Viral ; genetics ; Hepacivirus ; classification ; genetics ; isolation & purification ; Hepatitis C ; virology ; Humans ; Molecular Sequence Data ; Nucleic Acid Amplification Techniques ; methods ; Polymorphism, Restriction Fragment Length ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA

OBJECTIVETo observe the action potential duration restitution (APDR) change and potential association with ventricular arrhythmia (VA) in Langendorff-perfused chronic heart failure rabbit hearts.

METHODSMale rabbits were randomly divided into two groups: control (CTL, n=15) group and chronic heart failure (CHF, n=15) group. CHF was induced by injecting isoproterenol (300 µg×kg(-1) ×d(-1)) for 14 days. Four weeks later, cardiac function and structure change of both groups were assessed by echocardiography. In the whole Langendorff-perfused hearts, the monophasic action potential (MAP) and the effective refractory period (ERP) were recorded from left anterior basal ventricle, left anterior free wall, left anterior apex and left posterior basal ventricle, left posterior free wall and left posterior apex, the APD curves were also constructed in both groups; at the six sites of every isolated heart, the programmed electrical stimulation and burst pacing were used to induce action potential duration (APD) alternans and VA, respectively.

RESULTSLeft ventricular ejection was reduced and end-dimension was enlarged in rabbits of CHF group. Compared with the same sites of CTL group, the 90% of MAP duration (MAPD90), the ERP, the max slope (Smax) of APDR curves, the pacing cycle length of inducing the APD alternans and the VAs were significantly increased (all P<0.05) in CHF group; the spatial dispersions of MAPD90, ERP and Smax of APDR curves in CHF group were also greater than in CTL group (all P<0.05).

CONCLUSIONThe ventricular APD alternans might be linked with occurrence of the VA in CHF rabbits. Increase of the Smax from APDR curves and the spatial dispersions of Smax in this CHF model might facilitate the development of ventricular arrhythmia.

Action Potentials ; Animals ; Arrhythmias, Cardiac ; physiopathology ; Electrocardiography ; Heart Failure ; physiopathology ; Heart Ventricles ; physiopathology ; Male ; Rabbits ; Ventricular Fibrillation ; physiopathology

3' Untranslated Regions ; genetics ; Base Sequence ; China ; DNA, Complementary ; chemistry ; genetics ; Genetic Variation ; Hepacivirus ; genetics ; Humans ; Molecular Sequence Data ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Alignment ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid

OBJECTIVETo evaluate the feasibility and safety of total colonic exclusion plus side to side antiperistaltic ileorectal anastomosis in the treatment for elderly patients with slow transit constipation (STC).

METHODSClinical data of 13 patients with severe idiopathic STC undergoing the above novel procedure in Zhongnan Hospital of Wuhan University between May 2009 and September 2012 were retrospectively analyzed. The Wexner constipation score and gastrointestinal quality of life index (GIQLI) before and 6 months after operation were compared.

RESULTSThere were 8 female and 5 male patients, with a mean age of 74 years (range 63-82 years). No procedure-related deaths or any serious complication occurred. The length of follow-up ranged from 6 to 29 months (median,12 months). The duration of surgery was (55±4) min. Blood loss was (30±2) ml. The postoperative hospital stay ranged 10 to 16 days (mean 11.4 days). The first bowel movement occurred in the 4th day (range 2nd-8th day) after operation. There was no intestinal occlusion and anastomotic leakage that required surgery in all the patients. No fecal incontinence or constipation recurrence was found. One patient developed blind loop syndrome 14 months after operation. Postoperative complications included incision fat liquefaction in 2 cases, anorectal bearing-down while bowel movement in 2 cases, and minor defecate difficulty needed glycerin enema in 1 case. Wexner scores was significantly improved from 22.8±3.3 before operation to 5.4±2.1 six months after operation (P<0.05). GQLI was significantly increased from 93.6±20.5 before operation to 120.8±13.0 six months after operation (P<0.05). At 6 months after operation, the outcome was excellent in 11 patients and good in 2 patients.

CONCLUSIONTotal colonic exclusion plus side to side antiperistaltic ileorectal anastomosis is easy, safe and effective in the treatment for selected elderly patients with STC.

Aged ; Aged, 80 and over ; Anastomosis, Surgical ; methods ; Colon ; surgery ; Constipation ; surgery ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Retrospective Studies