AIM: Direct exposure of cells to reactive oxygen species can induce apoptosis. In this study we investigate how oxidative stress induces cell death in HepG2 cells and characterize the molecular events involved. METHODS: Oxidative stress was created by exposing HepG2 cells to 2 mmol/L H2O2. Apoptosis was determined by analysis of DNA fragmentation by agarose gel electorphoresis. The mitochondrial membrane potential was analyzed using DePsipher fluorescent staining and the expression of cytochrome c in the cytosolic fraction was measured by Western blotting analysis.The caspase activity was detected using fluorometric assay kit by a fluorescence microplate reader. RESULTS: When HepG2 cells were treated with 2 mmol/L H2O2, the cells displayed DNA fragmentation, a typical feature of apoptosis, after 12 h. The mitochondrial membrane potential appeared different in two group of cells. H2O2 -treated cells appeared green fluorescence as early as 4 h, which represents de - energized mitochondria, the untreated cells appeared red fluorescence,a feature of mitochondria with intact membrane potential. In treated cells, the expression of cytochrome c increased and accumulated in cytosolic fraction with treatment time, caspase - 3 activity increased by 6.7 - fold ( P ＜ 0.01 ) at 8 h and caspase -9 activity increased by 3.6 - fold (P ＜ 0.01 ) at 12 h, respectively, however, the activity of caspase - 8 remained unchanged. CONCLUSION: These findings suggest that oxidative stress can induce apoptotic cell death in HepG2 cells, and the mechanism is related to mitochondrial pathway, which activates caspase -9 and- 3, but not caspase -8.

Aim To investigate the role of endoplasmic reticulum stress in adenosine induced HepG2 cell apoptosis.Methods HepG2 cells were treated with different concentrations of ADO for 36 h,and the effect of ADO on cell proliferation was measured by MTT assay.Cell nuclei DAPI staining was used to detect the nuclei change after being treated with different concentrations ADO for 36 h or 2.0 mmol?L-1 ADO for different time.The effect of ADO on HepG2 cell cycle was analysed by flow cytometry after being treated with 2.0 mmol?L-1 ADO for 12 h or 24 h.Translocation of CHOP and Caspase-3 were measured by immunofluorescence after being treated with 2.0 mmol?L-1 ADO for 36 h.The proteins expressions of CHOP,Caspase-4,Caspase-3 and JNK were assayed by western blot.Results The viability of HepG2 cell decreased in a dose-dependent manner;the relative cell viability of 0.5,1,2,4,6 mmol?L-1 decreased by 13.48%?0.12%,27.92%?0.25%,35.21%?0.42%,51.46%?0.24%,71.42%?0.58%,compared with the control group respectively.The nuclei of HepG2 cells treated with different ADO concentrations or 2.0 mmol?L-1 ADO showed condensation,rounding and shrinkage,then fragmentation,which demonstrated cell apoptosis.After being treated with 2.0 mmol?L-1 ADO for 12 h or 24 h,cell cycle analysis showed sub-G1 phase increased;the apoptotic ratio of control,12 h and 24 h was 1.55%?0.12%,10.96%?0.07% and 21.04%?0.26% respectively.Immunofluorescence assay showed the CHOP and Caspase-3 translocation to the nuclei after being treated with 2.0 mmol?L-1 ADO.The expressions of CHOP,Caspase-3 and Caspase-4 increased after being treated with different concentrations ADO for 36 h,while the expression of JNK did not change.Conclusion Endoplasmic reticulum stress is involved in adenosine-induced HepG2 cell apoptosis.

Objective To investigate the clinical characteristics of patients with polymyositis(PM) and dermatomyositis (DM), and compare the differences of PM/DM to help the understanding of clinical diagnosis and treatment. Methods One hundred and forty-five hospitalized PM/DM patients from Department of Rheumatology of the First Affiliated Hospital of Xiˊan Jiaotong University were collected from May 2008 to December 2014, and the clinical manifestations, muscle enzymes, electromyogram, muscle biopsy, treatment outcome were retrospectively analyzed. Mann-Whitney U test and χ2 test were used for statistical analysis. Results The most common initial symptom of PM was muscle weakness, accounted for 51.2%, while rash was the initial presentation in most DM patients(43.1%). The incidence of interstitial lung disease (ILD) (62.7% vs 39.5%, χ2=11.009, P=0.001), and the elevation of CRP (48.9% vs 26.8%, χ2=10.272, P=0.001) were all higher in DM than PM, while the elevation of level of CK (85.4% vs 61.8%, U=-2.668, P=0.008) and CKMB (82.9%vs 41.2%, U=-3.303, P=0.001) were more common in PM compared with DM. The pathological study showed degeneration of muscle fiber, connective tissue hyperplasia in most PM patients, and perimysium atrophy, vacuoles degeneration, muscle bundles, perivascular inflammatory cell infiltration were observed in most DM patients. During the follow-up, the clinical remission rate was 57.5%, the relapse rate and the mortality rate was 7.5%and 31.1%respectively. The mortality rate was higher in DM than PM (34.6% vs 21.4%, χ2=4.861, P=0.027). Infection and tumors were the major causes of death, and the lung was the most common site of infection. Conclusion Differences in the clinical features, muscle enzymes, CRP level, pathology and the mortality rate between PM and DM are evident, while ILD, infection and the higher mortality rate are more common in DM than in PM.

[ ABSTRACT] AIM:To investigate the effects of tanshinone IIA ( Tan IIA) on proliferation, apoptosis and its mo-lecular mechanism in human hepatoma HepG2 cells under hypoxic condition.METHODS:Hypoxia model was established by treatment with cobalt chloride ( CoCl2 ) .The cells were divided into normoxia control group, hypoxia control group and hypoxia combined at different concentrations of Tan IIA groups.After HepG2 cells were incubated with different concentra-tions of Tan IIA (0.5, 1.0, 2.0, 5.0 and 10.0 mg/L) for 24 h, 48 h and 72 h under hypoxic condition, the cell prolifer-ation was determined by MTT assay.After Tan IIA was added to the media at different concentrations for 24 h and 48 h, the apoptotic cells were observed by Hoechst 33258 staining.The protein levels of hypoxia-inducible factor 1 alpha (HIF-1α) , vascular endothelial growth factor ( VEGF) and wild-type P53 were detected by Western blotting after cultured with different concentrations of Tan IIA for 48 h.RESULTS:Tan IIA inhibited the proliferation of HepG2 cells in a dose-and time-dependent manner.Tan IIA induced the typical morphology of apoptotic cells and increased the apoptotic rate in a dose-and time-dependent manner after treatment with 1.0 mg/L~5.0 mg/L for 24 h and 48 h under hypoxic condition. The protein levels of HIF-1αand VEGF were weakly expressed in HepG2 cells under normoxia but up-regulated after incu-bated under hypoxia for 48 h.The protein expression of HIF-1αand VEGF were decreased with the increase in the concen-tration of Tan IIA under hypoxia.The protein expression of wild-type P53 was increased with the increase in the concentra-tions of Tan IIA under hypoxia.CONCLUSION:Tan IIA significantly inhibits the proliferation and induces the apoptosis of human hepatoma HepG2 cells under hypoxia, which may be related to the down-regulation of HIF-1αand VEGF and up-regulation of wild-type P53.

Aim To investigate the mechanism of demethylation on adenosine (ADO )and homocysteine (HCY)-induced apoptosis in human hepatoma HepG2 cells .Methods HepG 2 cells were treated with differ-ent concentrations of ADO (1.0、2.0、4.0 mol · L-1 ) alone or in combination with HCY for 6h,12h and 24h,5-aza-2-deoxycytidine (5-Aza-CdR)as a positive control.Cell viabilities were assessed by CCK8 assay. Cell apoptosis was observed by AnnexinV-FITC/PI staining.The mitochondrial membrane potentials(ΔΨ) were measured by flow cytometry.The mRNA and pro-tein expressions of caspase-3,caspase-8,caspase-9, MDM-2,p53,Cytochrome C,DNMT1,DNMT3a,DN-MT3 b were detected by RT-qPCR and Western blot re-spectively.Results ADO alone or in combination with HCY significantly reduced the viability of HepG2 cells in a dose and time-dependent manner.The apoptotic rates of HepG2 cells after combination treatment with ADO and HCY at 1 .0,2.0,4.0 mol · L-1 for 24 h were (1 8.63 ± 1.25 )%,(29.42 ±2.37 )% and (42.47 ±3.09 )%,compared with the control group (1.30 ±0.82 )%,P <0.01;and the mitochondrial membrane potentials were decreased from 674.15 ± 82.8%(black control group)to (428.38 ±54.5)%, (297.78 ±30.5)%,(74.45 ±5.73)%,P<0.01, respectively.The expressions of caspase-3,caspase-8, caspase-9,MDM-2,p53,Cytochrome C were up-regula-ted and MDM-2 were down-regulated after combination treatment of ADO and HCY.The mRNA expressions of DNMT1 ,DNMT3 a and DNMT3 b were down-regulated after combination treatment with ADO and HCY or 5-Aza-CdR alone.Conclusion Combination treatment of ADO and HCY can cause cellular methylation chan-ges.The effects of demethylation of ADO and HCY may activate p53 gene and mitochondrial pathway, which at last leads to HepG2 cell apoptosis.

Objective:To establish a prediction model of postoperative 1-year mortality risk in elderly patients undergoing hip fracture surgery and verify its efficacy.Methods:Patients of both sexes, aged ≥65 yr, of American Society of Anesthesiologists physical status Ⅰ-Ⅳ, who underwent an operation for traumatic hip fracture in the Second Affiliated Hospital of Wenzhou Medical University from January 2017 to December 2018, were enrolled and randomly assigned to model group and verification group in a ratio of 3∶1.The demographic characteristics, clinical data and results such as laboratory examinations were collected.In model group, the logistic regression analysis was used to recognize the independent risk factors for 1-year mortality after procedure, and the prediction model was established.In verification group, the prediction efficacy was analyzed using the receiver operating characteristic curve, and the degree of fitting was evaluated by Hosmer-Lemeshow goodness-of-fit test.Results:Multivariate logistic analysis indicated that age ≥84 yr, Charlson comorbidity index ≥2 points, Braden score on admission to hospital ≤16 points, preoperative urea nitrogen ≥8.8 mmol/L and postoperative albumin ≤ 29.6 g/L were the independent risk factors for 1-year mortality after hip fracture surgery in elderly patients ( P<0.05). The prediction model was established based on the risk factors mentioned above.The area under receiver operating characteristic curve was 0.870, and the sensitivity and specificity were 82.8% and 80.0%, respectively.The prediction model showed good fitting ( χ2=4.672, P=0.700). Conclusion:Age ≥84 yr, Charlson comorbidity index ≥2 points, Braden score on admission to hospital≤16 points, preoperative urea nitrogen ≥8.8 mmol/L and postoperative albumin ≤ 29.6 g/L are the independent risk factors for 1-year mortality after hip fracture surgery in elderly patients, and the prediction model established based on the above indicators has good efficacy.

【Objective】 To investigate the clinical characteristics and risk factors of systemic lupus erythematosus （SLE） combined with symptomatic knee osteonecrosis （KON）. 【Methods】 We retrospectively analyzed the clinical data of 26 cases of SLE with KON treated in the Department of Rheumatology and Immunology， The First Affiliated Hospital of Xi’an Jiaotong University， from April 2013 to December 2019. 【Results】 The age of the 26 patients （2 males and 24 females） was （35.3±9.0） years old at the diagnosis of KON， and the course of SLE was （48.7±35.1） months. The time from glucocorticoids initiation to the development of KON was （37.8±42.7） months， the maximum dosage of methylprednisolone was （197.7±290.7）mg and the cumulative dosage was （6.02±6.66）g. Six of the patients had a history of large-dose glucocorticoids impulse therapy. All of them had a history of immunosuppressant treatment. SLEDAI score was （11.23±5.46） at the onset of SLE and （4.46±4.81） at the onset of KON. The most common initial symptoms were edema and arthritis. The most common systemic damages were blood system damage and lupus nephritis. The most common immunological abnormalities were positive antinuclear antibody （25/26）， positive anti-SSA/Ro52kD antibody （16/26）， and positive anti-SmD1 antibody （15/26）. There were 4 patients with positive anticardiolipin antibody （ACA）. Bone metabolism was characterized by vitamin D3 （Vit-D3） deficiency， insufficient N-terminal osteocalcin （N-OST）， and increased β-C-terminal telopeptide of type I collagen （β-CTx）. In 9 out of the 26 patients， SLE was combined with aseptic necrosis of the femoral head. Multifocal bone necrosis （at least 3 lesions） was common （12/26）. Longer disease course and glucocorticoids using time， larger cumulative dose and ACA positive were seen in patients with multifocal bone necrosis compared with those who had one lesion site. 【Conclusion】 KON most possibly occurs 3 to 4 years after the diagnosis of SLE， which is associated with high disease activity， large hormone dose， and long duration in the treatment process. Multifocal bone necrosis is easily seen in patients with severe disease， large initial hormone dose and high cumulative dose， as well as in ACA positive patients.

【Objective】 To evaluate musculoskeletal ultrasound (MSUS) detected subclinical synovitis of rheumatoid arthritis (RA) with different clinical remission criteria so as to explore the clinical characteristics of subclinical synovitis. 【Methods】 Forty-six consecutive patients with RA in clinical remission [disease activity score-28 (DAS28)≤2.6] underwent clinical and MSUS examinations at baseline and 1 year follow-up. Clinical remission was defined according to the DAS28 using the erythrocyte sedimentation rate (DAS28-ESR) and C-reactive protein level (DAS28-CRP), clinical disease activity index (CDAI), simplified clinical disease activity index (SDAI), and American College of Rheumatology/European League Against Rheumatism criteria Boolean (ACR/EULAR criteria). Subclinical synovitis was assessed by MSUS. Differences between the subclinical synovitis and non-subclinical synovitis groups were analyzed. 【Results】 The percentages of patients who achieved DAS28-ESR, DAS28-CRP, CDAI, SDAI, and ACR/EULAR remission at baseline and 1 year were 97.8%, 95.6%, 67.4%, 54.3%, 52.2% and 91.3%, 93.5%, 54.3%, 50.0%, and 45.6%, respectively. Subclinical synovitis was detected in 55.5%, 54.5%, 45.2%, 40.0%, 41.6% and 45.2%, 46.5%, 40.0%, 39.1%, and 38.1% of these patients, respectively. There were 45.6% and 41.3% patients who fulfilled all the criteria, yet 38.1% and 36.8% still had evidence of subclinical synovitis at baseline and 1 year. Compared with the patients without subclinical synovitis, those with subclinical synovitis had a significantly positive rate of anti-CCP antibody and a higher disease activity score at baseline (P<0.05). 【Conclusion】 MSUS detected subclinical synovitis is common. The positive anti-CCP antibody and higher disease activity score at baseline may be related to subclinical synovitis in patients with RA in clinical remission.