Objective:To investigate the effect of cPLA 2(cytosolic PLA 2, 85-kD PLA 2) on the signaling pathways of IL-1?、IL-6 release in LPS induced Hela cells.Methods:①LPS stimulated Hela cells were exposed to the complex of cPLA 2 initiation site-directed antisense oligonucleotide and liposome in the indicated doses and time, and then,analyzed the changes of cPLA 2 both in mRNA and in protein by RT-PCR and Western blot.②At the same time, the supernatants were collected to detect the level of IL-1? and IL-6 under radioimmunoassay(RIA).Results:①The transfection led to a dose-dependent decrease in cPLA 2 protein, while the change of mRNA is not notable.②The levels of IL-1?、IL-6 in the transfected cells were reduced following the repression of cPLA 2 compared with that of LPS added only.Conclusion:cPLA 2 probably plays an important role in the signaling mechanisms on IL-1?、IL-6 release in LPS-induced Hela cell. [

OBJECTIVE:To establish an LC-MS method for determination of picamilon in human plasma. METHODS:Picamilon and repaglinide(as internal standard)were separated on Agilent C18 column,using methanol-water(85 ∶ 15) as the mobile phase at a flow rate of 0.2 mL?min-1. Column temperature was set at 20 ℃. SIM was used for determination of picamilon in human plasma.Ion mass spectral(m/z) of 207.0→121.1 was selected for picamilon and 451.3→379.2 for repaglinide.RESULTS:The linear range was 50～10 000 ?g?L-1(r=0.999 1); the method recovery was within 99.14%～104.27%. The RSD of inter-day and intra-day validation were 2.12%～5.94% and 7.30%～8.44%,respectively. CONCLUSION:The method is sensitive and accurate for the determination of picamilon in human plasma and its pharmacokinetic study.

AIM: To establish a HPLC quality control for Chizhihuang Capsules(Radix Paeoniae Rubra, Fructus Gardeniae, Radix et Rhizoma Rhei, etc.). METHODS: Shim pack C 18 column was used, and jasminoidin and paeoniflorin were determined at the same time and detection wavelength at 230nm. RESULTS: The standard curve of jasminoidin was linear in the range of 0.30～1.50?g. The average recovery and RSD were 99.58 and 1.57% ( n =5), respectively. The standard curve of paeoniflorin was linear in the range of 0.26～ 1.30?g . The average recovery and RSD were 99.11 and 1.29% ( n =5), respectively. CONCLUSIONS: This method is a convenient, reliable and accurate for the quality control of Chizhihuang Capsules.

Objective To evaluate the prevalence of subclinical atherosclerosis in rheumatoid arthritis (RA) and the related risk factors.Methods Fifty RA patients without overt atherosclerotic disease and 121 control subjects matched for age and sex were recruited.Duplex carotid sonography was used to measure intima-media thickness (IMT) and plaque formation assessment.Differences between RA and the control group were compared, and the risk factors were explored.RA patients then were divided into two subgroups according to IMT and the comparison between the two subgroups were completed.T test, Mann-Whitney U test, chi-square test, Pearson's correlation and Logistic regression analysis were used for statistical analysis.Results Although RA patients had lower level of serum lipids and body mass index than the control group, the mean IMT value was significantly higher in the RA group than that in the control group [(0.78±0.18) mm vs (0.62±0.14) mm, t=5.853, P=0.000], and plaque formation was more prevalent [56.0％(28/50) vs 36.4％ (44/121),x2=5.596, P=0.018].The difference was especially significant in the younger groups (＜50 years old group and 50-60 years old group).Logistic regression showed that the presence of RA [OR=7.34, 95％CI (2.53, 21.25)], male [OR=2.0, 95％CI (91.25, 3.17)] and old age [OR=1.1, 95％CI (1.04, 21.15)] were the independent risk factors for abnormal IMT (thickened or the presence of carotid plaques).The RA patients were divided into two subgroups according to IMT.Compared with patients with normal IMT, patients with abnormal IMT were older and more common in postmenopausal, and had longer RA duration and higher cholesterol level.In treatment, less patients with abnormal IMT had been taking methotrexate (MTX) for more than 3 months compared with patients with normal IMT.Among these parameters, old age [OR=1.13, 95％CI (1.03, 1.23)] was shown to be the independent risk factor for abnormal IMT in RA patients, and more than 3 months of MTX treatment [OR=0.12, 95％CI (0.02, 0.71)] was the protective factor.Conclusion Atherosclerosis occurs frequently and prematurely in patients with RA and the presence of RA is an independent risk factor for atherosclerosis.Early primary prevention for atherosclerosis should be recommended.MTX probably has a positive effect on preventing atherosclerosis for RA patients, which needs to be confirmed by further study.

Objective:To verify the fracture risk assessment tool ( FRAX) to estimate the probability of osteoporotic fracture in patients with rheumatoid arthritis ( RA ) with or without bone mineral density (BMD), and identify associated risk factors of osteoporosis .Methods: In the study, 200 patients with rheumatoid arthritis aged more than 40 years in Peking University First Hospital from Dec .2009 to Dec. 2012 were recruited.Clinical information was obtained from a questionnaire of their case history and medical records.FRAX tool was administered.Their lumber spine and left femoral BMD were determined by dual energy X ray absorptiometry.The gender, age, disease duration, menopause status, body mass index ( BMI) and accumulative dose of glucocorticoid were obtained in retrospect .Correlation analysis was conducted between the BMD and clinical information .Results:The study population ( female, 77.5%) had a mean age of 59.4 years, in which 10 (13%) patients showed a normal BMD, 67 (87%) were osteopenia or osteoporosis , while 32 patients (16%) had fragile fracture.Compared with the patients with normal BMD, the subjects with low BMD had significantly older age , longer period for corticoids usage , higher day dose and accumulated dose of corticoids .The 10-year fracture risk of sustai-ning major osteoporotic fractures and hip fracture was higher .No significant difference was observed be-tween the 10-year fracture risks calculated with BMD and without BMD .The values of the different area under the receiver operating characteristic ( ROC) curve ( AUC) for major and hip fractures calculated in three ways:without BMD, with the femoral neck BMD, and with T-score.The best result was for FRAX tool for hip fracture with the T-score ( AUC 0 .899 ) .A stepwise multivariate linear regression model was constructed to explore the relationship between the different clinical factors studied and a low BMD . Three statistically significant variables for lumber BMD were pain on visual assessment scale ( VAS ) (P=0.02), fracture history (P=0.003) and a higher steroid accumulated dose (P=0.008).Three statistically significant variables for left hip BMD were age (P<0.001), fracture history (P=0.05) and lower BMI ( P=0.03) .Conclusion:Low BMD is a common complication in RA patients .Risk factors for major fracture and hip fracture are increased .There is a positive correlation between FRAX calculated with and without BMD or T score .FRAX with the femoral neck T score or BMD presents a discriminatory capacity better than FRAX without BMD , according to the AUC ROC .

Objective To investigate the outcomes of patients with rheumatoid arthritis (RA) treated by different combination of synthetic disease modifying antirheumatic drugs (DMARDs) under the guidance of treat-to-target strategy.Methods Forty-two RA patients with high disease activity were enrolled into this randomized,open-label and prospective study.It was comprised of a maximal 36-week induction phase and then followed by a maintenance phase up to 84 weeks.Combination of synthetic DMARDs was initiated in the induction phase,with or without low dose glucocorticoids (GCs) during the first 12 weeks.Patients who achieved low disease activity (LDA) were randomized into two maintenance groups.An increase of DAS28 by 0.6 was defined as relapse.The patients achieved LDA in the induction phase,relapsed during maintenance phase and possible relevant risk factors were analyzed.Results Twenty-seven (64％) patients achieved LDA during the induction phase.More non-smoking patients achieved LDA than those smoked [85％ (11/13) vs 55％(16/29),P＜0.05].During the maintenance phase,14 (61％) out of 27 patients relapsed.Patients taking GCs during the first 12 weeks had a significantly higher relapse rate compared to those without GC (83％ vs 36％,P=0.021).Patients who entered maintenance phase at week 12 had a significantly higher tendency to relapse compared to those who entered the maintenance phase at week 24 [75％(9/12) vs 33％(3/9),P=0.026].Conclusion Smoking seems to be a risk factor for RA patients who fail to reach LDA.Low dose GCs as a bridge therapy may require a longer duration.High relapse rates in both the maintenance groups indicat that a longer tight induction phase may be appropriate before downstairs therapy.

AIM: To detect the effect of sepsis on hepatic,renal functions and corresponding enzymes in intestine of mice,and to explore the role of leptin in acute inflammation.METHODS: A mice model of sepsis was made by cecum ligation and perforation,and 96-well spectrophotometry was used to detecte the levels of alanine aminotransferase(ALT),uric acid(UA) and activities of myeloperoxidase(MPO),glutathin-S-transferase(GST),xanthine oxidase(XOD),superoxide dismutase(SOD) in serum and intestinal homogenized fluids,respectively.Hematoxylin-eosin staining was used simultaneously to check the histopathologic changes of intestine.RESULTS: Compared with sham group(330.12 ?mol/L?94.15 ?mol/L),serum UA level(521.92 ?mol/L?91.86 ?mol/L) at 6 h after sepsis was significantly higher.12 h after sepsis,both serum ALT(83.55 U/L?40.44 U/L) and UA(474.03 ?mol/L?75.22 ?mol/L) were significantly higher than those in sham group(66.23 U/L?16.80 U/L and 320.95 ?mol/L?99.14 ?mol/L,respectively).12 h after leptin injection(0.1 mg/kg,ip) or indomethacin injection(2 mg/kg,ip),the serum ALT and UA levels significantly decreased(vs sepsis group,P

OBJECTIVE: To explore the change of orexin-A expression in hepatic reperfusion and their association with liver injury, and to find out the role of orexin-A in traumatic stress responses. METHODS: A 70% hepatic reperfusion model of rats was established, setting groups of sham-operation and injury ones with different reperfusion time. A self-produced radioimmunoassay and relevant kits were used to detect the protein level of orexin-A in the plasma and the hypothalamus, serum glucose, total anti-oxidation capacity and alanine transaminase, HE staining and immunohistochemistry were used to investigate the pathological variation and protein expression of orexin-A in the liver, while RT-PCR was applied to observe mRNA expression of orexin-A in the hypothalamus and the liver. RESULTS: Both the shape of standard curve and metrical results of the self-produced orexin-A radioimmunoassay were good. Protein levels of orexin-A in the plasma and the hypothalamus in each reperfusion group showed no significant change. Serum glucose and total anti-oxidation capacity increased significantly at the later phase of injury. There was significant and positive linear correlation between the plasma orexin-A and serum glucose and total anti-oxidation capacity; serum alanine transaminase in each reperfusion group was significantly higher, and liver damage was significantly alleviated at the later phase of the injury. Different extents of variation were observed in protein expression of orexin-A in the liver and its mRNA expression in the hypothalamus and the liver. CONCLUSION Orexin-A undergoes significant changes during hepatic reperfusion, indicating that orexin-A participates in the modulation of hepatic reperfusion-induced liver injury and internal disorders.
Animals ; Hypothalamus ; metabolism ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Liver ; blood supply ; metabolism ; Male ; Neuropeptides ; genetics ; metabolism ; Orexins ; RNA, Messenger ; genetics ; metabolism ; Radioimmunoassay ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; metabolism

ObjectiveTo evaluate the clinical and radiographic efficacy and safety of the combination of recombinant human tumor necrosis factor-αt receptor Ⅱ IgG Fc fusion protein (rhTNFR:Fc) and methotrexate (MTX) in patients with rheumatoid arthritis (RA). MethodsThirty patients with highly active RA were treated with rhTNFR:Fc (25 mg subcutaneously twice weekly) and oral MTX (up to 15 mg weekly). Clinical efficacy was assessed using ACR response criteria and the disease activity score in 28 joints (DAS28).Radiographs of the hands and wrists were assessed with the modified Sharp score. Chi-square test, Fisher is exact test and paired t-test were performed. ResultsAt week 52, ACR20, ACR50 and ACR70 responses were achieved by 90％, 87％ and 67％ respectively. At week 52, mean DAS28 was 3.4±1.1 compared to 6.4±0.6 at base-line(P＜0.01), with 23％ patients achieving clinical remission and 17％ patients in low disease activity. Similarly, the HAQ was improved significantly, declining from 1.18±0.56 at base-line to 0.25t±0.34 at week 52 (P＜0.01). No radiographic progression was found in 22 cases. Adverse events were mild in general.ConclusionTreatment with rhTNFR:Fc plus MTX has shown good efficacy throughout 52 study period in reducing disease activity, improving function, and retarding radiographic progression. Combination therapy for 52 weeks can achieve disease remission and no radiographic progression, which are the two goals of therapy for RA.

Objective:To investigate the protective effect of naringenin on acute lung injury related with sepsis; To discuss its possible mechanism.Methods:Totally 30 male SD rats were randomly divided into sham-operation group, model group, naringin low-, medium- and high-dosage groups, with 6 rats in each group. The sepsis-related acute lung injury model was established by cecal ligation and puncture in all groups except the sham-operation group. After modeling, naringin low-, medium- and high-dosage groups were given naringin 20 mg/kg, 40 mg/kg and 80 mg/kg, respectively for gavage, while the sham-operation group and the model group were given the same volume of distilled water by gavage, once a day, for 2 days. Pathological changes in lung tissue were observed using HE staining. The levels of 1L-1, IL-6 and IL-18 in bronchoalveolar lavage fluid (BALF) were measured by ELISA; the expression of TNF-α in lung tissue was detected by immunofluorescence histopathology; the expressions of TGF-β1, TGF-βR1 and Smad2 were detected by Western Blot. An agonist group and a naringin plus agonist group were set up, with 6 mice in each group, and the expressions of TGF-β1 and Smad2 protein in the lung tissue of each group were detected by immunohistochemical staining to verify the effect of naringin on the expressions of TGF-β1 and Smad2 protein.Results:Compared with the model group, the pathological injury of lung tissue in naringin groups were obviously alleviated, and the levels of IL-1β, IL-6 and IL-18 in BALF decreased ( P<0.01), the protein expressions of TNF-α, TGF-β1, TGF-βR1 and Smad2 in lung tissue decreased ( P<0.01 or P<0.05). Further verification found that the expressions of TGF-β1 and Smad2 in the agonist group increased ( P<0.01), while the expressions of TGF-β1 and Smad2 in the naringin agonist group decreased ( P<0.01). Conclusion:Naringin can reduce the inflammatory response in the lung of the rats to protect against sepsis-related acute lung injury, and its protective effect could be related to the inhibition of the TGF-β1/Smad2 signaling pathway.