Objective To construct a eukaryotic expression plasmid carrying the PKCI-1/HINT1 gene,to investigate its expression in A375 melanoma cells after transfection,and to evaluate its effects on apoptosis and autophagy of A375 cells.Methods The PKCI-1/HINT1 gene sequence was amplified by reverse transcription PCR (RT-PCR) with total RNA extracted from A375 cells as the template,then inserted into the eukaryotic expression plasmid PCDNA3.1 (+) to construct a recombinant plasmid,PCDNA3.1 (+)-PKCI-1/HINT1.Some A375 cells were classified into two groups to be transiently transfected with the recombinant plasmid (PCDNA3.1 (+)-PKCI-1/HINT1 group) or the empty plasmid PCDNA3.1 (+) (control group).After additional 48-hour culture,RT-PCR and Western blot analysis were performed to quantify the mRNA and protein expressions of PKCI-1/HINT1 respectively,Hoechst 33342 staining was conducted to detect apoptosis of A375 cells,Western blot analysis to detect the expressions of intracellular caspase-3 and autophagy-associated protein beclin1,and cell autophagy was observed by using the green fluorescent protein (GFP)-microtubule-associated protein 1 light chain 3 (LC3) labelling method combined with confocal laser scanning microscopy.Methyl thiazolyl tetrazolium (MTT) assay was performed to evaluate the proliferative activity of A375 cells at 24,48,72 and 96 hours after transfection.Results Enzyme digestion and sequencing analysis confirmed that the eukaryotic expression plasmid PCDNA3.1 (+)-PKCI-1/HINT1 was successfully constructed and effectively expressed in the transfected A375 cells.MTT assay showed that PKCI-1/HINT1 could obviously inhibit the proliferation of A375 cells,and the number of live cells was decreased by 17.0％,25.6％ and 29.4％ in the PCDNA3.1 (+)-PKCI-1/HINT1 group at 48,72 and 96 hours,respectively,compared with the control group (all P ＜ 0.05).Hoechest 33258 staining revealed that PKCI-1/HINT1 could promote the formation of apoptotic bodies in A375 cells.Confocal laser scanning microscopy demonstrated that the overexpression of PKCI-1/HINT1 increased GFP-LC3 puncta formation in A375 cells.In addition,Western blot analysis indicated that PKCI-1/HINT1 up-regulated the protein expressions of caspase-3 and beelin1 in A375 cells.Conclusions The eukaryotic expression plasmid PCDNA3.1 (+)-PKCI-1/HINT1 was successfully constructed,and PKCI-1/HINT1 could be effectively expressed in A375 cells.High-level expression of PKCI-1/HINT1 could suppress cellular proliferation,promote apoptosis,and induce autophagy,of A375 cells.

Objective To introduce a self- assembled adjustable traction frame for the patients with severe trauma and lower limb fracture in ICU, compared with Brawns traction frame. Methods 96 patients with severe trauma and lower limb fracture diagnosed by clinical signs and X-ray who had to be transferred to ICU were randomly divided into the study group (46 cases) and the control group (50 cases) by sealing letters method.The study group were tracted by the self-assembled adjustable traction frame, and the control group were tracted by the Brawns traction frame. The fracture alignment, degree of pain, incidence of pressure ulcers, nursing limit time of changing the sheets of the two groups were observed. Results The results declared that 37 cases with 2 pain score points, 9 cases with 0-1 points in the study group and 19 cases with 2 pain score points, 3 cases with 0-1 points, 28 cases with Grade 3 in the control group (χ2=38.683, P<0.01). In the control group, pressure ulcer were occurred, 5 of them at Level one, 1 of them at Level two, 1 of them at Level three and the incidence rate was 14.00% (7/50). In the study group, pressure ulcer were occurred, 1 of them at Level two and the incidence rate was 2.17%(1/46). The pain level and incidence of pressure sore in the study group were decreased and the difference between the two groups was significant (χ2=8.197, P < 0.05). Conclusions The self-assembly adjustable traction rack can not only ensure the same effect as the Brawns traction frame but also reduce the deficiency caused by Brown traction frame. It is safer and more comfortable for the patients and can reduce nursing complications and improve the quality and efficiency of the nursing work.

Objective: To evaluate the quality consistency of four domestic nifedipine sustained release tablets by dissolution test and virtual bioequivalence study by GastroPlus software.Methods: The dissolution curves of the four preparations were determined with the methods described in Japanese orange book and Chinese Pharmacopeia.The f2 factor of dissolution curves was calculated to compare the similarity.The in vitro dissolution data of the original preparation were combined with GastroPlus software to obtain the simulated in vivo absorption curves which were correlated with the actual concentration-time curves.The suitable dissolution medium was selected to evaluate the quality of domestic nifedipine sustained release tablets according to the better in vivo-in vitro correlation (IVIVC).The simulated in vivo absorption parameters obtained from the dissolution data combined with GastroPlus software were used to conduct the virtual bioequivalence study of domestic nifedipine sustained release tablets compared with the original products.Results: The f2 similar factors of the four domestic nifedipine sustained release tablets compared with the original preparation were all less than 50.Compared with that from the method in Japanese orange book, the correlation between the dissolution profiles in vitro and in vivo of original nifedipine sustained release tablets obtained from the method in Chinese Pharmacopoeia was better.The deviation between the simulated Cmax and AUC0-∞ values of the four test tablets and the measured values of the original preparation was within the range of ±20%.Conclusion: The dissolution curves of the four domestic nifedipine sustained release tablets are not similar to that of the original preparation, however, the four preparations are all bioequivalent to the original preparation according to the simulated absorption parameters based on the dissolution method in Chinese Pharmacopeia and GastroPlus software.

Objective To investigate the effects of continuous positive airway pressure (CPAP) on the levels of hemorheology and C-reactive protein in patients with obstructive sleep hypopnea apnea syndrome (OSAHS). Methods 58 moderate to severe OSAHS subjects were selected as treatment group and 32 healthyadults were selected as control group. Hemorheology and C-reactive protein in all subjects were examined and compared. The levels of hemorheology and C-reactive protein were also examined and compared before and after 3 months CPAP treatment in treatment group. Results In treatment group, the levels of the whole blood viscosity and hs-CRP were significantly higher than those in control group (P < 0.01). After treatment of CPAP for three months, the levels of the whole blood viscosity and hs-CRP obviously decreased (P < 0.01). Conclusion Levels of hemorheology and hs-CRP are elevated in patients with OSAHS and CPAP therapy could effectively decrease serum levels of hemorheology and hs-CRP in patients with OSAHS.

Objective To explore the therapeutic effects of statin in patients with chronic obstructive pulmonary disease (COPD) in stable period. Methods 83 patients with COPD in stable period were recruited and randomly divided into two groups, treatment group (n = 41) and control group (n = 42).The control group received conventional routine therapy, and treatment group was given rosuvastatin calcium 20 mg once per day on the basis of routine treatment. The levels of sera C-reactive protein (CRP), pulmonary function (FEV1、FEVl%pred), and COPD assessment test (CAT) were observed at the time points of 0 week and 12th week, 24th week and 48th week were compared between two groups. Results At 12th week after the treatment, sera CRP levels, FEV1, FEV1%pred and CAT scores in either control group or treatment group showed some extent of improvements as compared to those at the time point of 0 week, and there was statistical difference of sera CRP levels and CAT scores between two groups (P < 0.05). There was no statistical difference of FEV1, FEV1%pred between two groups (P > 0.05) .24th week and 48th week after the treatment; sera CRP levels, FEV, FEV1%pred and CAT scores in two groups all significantly improved and were statistical different between two groups (P < 0.05). Conclusion Statin can alleviate inflammatory reaction, improve pulmonary function and life quality of patients with COPD.

Objectives To investigate the overall prevalence of cardiovascular disease (CVD) in subjects hospitalized for chronic obstructive pulmonary disease (COPD),and explore the prevalence of the major CVD complications and trends in patients with COPD over a 10-year period.Methods Medical records in the PLA General Hospital,Beijing Union Medical College Hospital,and Beijing Hospital from 2000/01/01 to 2010/03/03 were retrospectively reviewed.A total of 4960 patients with COPD were reviewed in the study (3570 males,mean age,72.2 ± 10.5 years; 1390 females,mean age,72.0 ± 10.4 years).Results The prevalence of CVD in COPD patients was 51.7％.The three most prevalent CVDs were ischemic heart disease (28.9％),heart failure (19.6％),and arrhythmia (12.6％).During the 10-year study period,the prevalence of various CVDs in COPD patients showed a gradual increasing trend with increasing age.There was higher morbidity due to ischemic heart disease (P ＜ 0.01) in male COPD patients than in the female counterparts.However,heart failure (P ＜ 0.01)and hypertension (P ＜ 0.01) occurred less frequently in male COPD patients than in female COPD patients.Furthermore,the prevalence of ischemic heart disease decreased year by year.In addition to heart failure,various types of CVD complications in COPD patients tended to occur in younger subjects.The prevalence of all major types of CVD in women tended to increase year by year.Conclusions The prevalence of CVD in patients hospitalized for COPD in Beijing was high.Age,sex and CVD trends,as well as life style changes,should be considered when prevention and control strategies are formulated.

Objective To improve the understanding of cutaneous Mycobacterium haemophilum infection.Methods Clinical and laboratory data were collected from two patients with cutaneous Mycobacterium haemophilum infection.The clinical and histopathologic manifestations, etiology and treatment of cutaneous Mycobacterium haemophilum infection were analyzed.Results The two patients were both immunocompromised.Lesions, which occurred after trauma in both the patients, began as well-marginated subcutaneous nodules, and then gradually progressed into ulceration with crusting and abscess formation.Histopathological examination revealed epithelial cell granuloma or histiocytic infiltrate in the dermis, as well as focal necrosis and neutrophil abscesses.Acid-fast staining showed abundant bacilli in tissues.These bacilli only grew in iron-containing medium, and were finally identified as Mycobacterium haemophilum by PCR and sequencing.According to the result of antimicrobial sensitivity testing, both the patients were treated with rifampicin, clarithromycin and moxifloxacin.Three months later, the condition was improved in both of them.Conclusions Mycobacterium haemophilum infection occurs most frequently in immunocompromised populations after trauma, manifests as subcutaneous nodules and abscesses with histopathological changes consistent with infectious granuloma.Molecular biological approaches are reliable for the identification of Mycobacterium haemophilum.

Objective To investigate the protective effects of memantine on isofluane￣induced decrease of proliferation in neural stem cells ( NSCs) and the potential mechanisms in vitro. Methods Neural stem cells were isolated from rat hippocampi (postnatal day 1) and grew in culture. Cultured NSCs were randomly divided into the control ( Group Control), Vehicle (Group Vehicle), Isoflurane ( Group Iso), Isoflurane +Memantine ( Group Iso +Mem) and Isoflurane + Memantine +LY294002 groups (Group Iso+Mem+LY).Proliferation was assessed by cell counting and BrdU incorporation.Western blot was conducted to detect protein expression of phospho￣Akt. Results Compared with the control group,BrdU incorporation and phospho￣Akt expressions in neural stem cells significantly declined after 2.4% isoflurane exposure for 6 h (P<0.01).However, isofluane￣induced decrease of BrdU incorporation and phospho￣Akt expressions was attenuated by the treatments of memantine (P<0.01)). It was showed that Akt inhibitors LY294002 reversed the protective effects on neural stem cells proliferation by memantine(P<0.01). Conclusion The results suggest that memantine treatment might attenuate isofluane￣induced decrease of proliferation in neural stem cells via Akt signaling pathway.

Objective To preliminarily understand the genotype characteristics of Toxoplasma gondii in blood of HIV?posi?tive persons in Lincang City,Yunnan Province. Method Two segments of SAG2 gene of T. gondii from blood samples of HIV?positive persons in Lincang City were extracted and amplified by using the nested PCR method and the genotype was identified and compared with the standard strain(Type I)of Toxoplasma gondii. Results Thirty?five SAG2 genes(241 bp)and 35 SAG2 genes(221 bp)of T. gondii were amplified from 170 blood samples of the HIV?positive people,and 4 of each case were selected and digested with enzyme,then 2 aim gene fragments of each case were chosen and compared with the standard strain (Type I)of T. gondii. The digestion of SAG2 gene(241 bp)showed the genotype of the blood samples was Type I or Type II, and the digestion of SAG2 gene(221 bp)confirmed that the genotype was Type I. Conclusion It is preliminarily confirmed that the genotype of T. gondii in blood of HIV?positive persons in Lincang City,Yunnan Province is Type I.

ObjectiveTo investigate the protein wasting and energy reserve in chronic renal failure (CRF) patients undergoing maintaining hemodialysis (HD) at different ages.MethodsA total of 129 CRF patients (62 men and 67 women) aged (56.33 ± 14.14) years on HD were enrolled in this study.They were divided into four age groups:below 40,40-69,60-69,and over 70 years.Nutritional status was assessed by body mass index (BMI),normalized protein catabolic rate (nPCR),and biochemical parameters including pre-albumin,cholesterol,and creatinine.Body composition was tested with multi-frequency bioelectric impedance analysis.Meanwhile,83 healthy subjects,matched for age and sex,were enrolled as controls.ResultsBMI showed no significant difference between healthy controls and HD patients.Among HD patients,38％ had an BMI higher than the normal high limit and 6.2％ had a BMI less than 18.5kg/m2 ; patients with a low body weight accounted for 0,6.2％,4.8％,and 11.1％ in the ＜40 years group,40-59 years group,60-69 years group,and above 70 years groups,respectively.nPCR in the above 70 years group was ( 1.46 ±0.28) g/( kg·d),which was significantly lower than that in the 40-59 years group [ (1.54 ±0.28) g/( kg·d) ; P =0.004) ; the serum creatinine in the the above 70 years group was (834.08 ± 184.96) μmol/L,which was significantly lower than that in 40-59 years group [ (976.24 ± 186.86) μmol/l ] ( P=0.037) ; the prealbumin in the the above 70 years group was (272.65 ±79.78) mg/L,which was significantly lower than that in 40-59 year group [ (332.07 ± 73.03 ) mg/L] ( P =0.026).Body cell mass in HD patients was (22.81 ±8.12) kg,which was significantly lower than that in the control group [ (29.95 ±6.73) kg] (P＜0.001) ; furthermore,the lean tissue mass (LTM) [ (40.16±11.90) kg vs.(47.22 ±9.84) kg] and fat [ (17.45± 8.83)vs.(13.66±7.28) kg] were also significantly lower (both P=0.001 ).The lean tissue indicators in the below 40 years group were also significantly lower than those in the healthy controls ( P =0.012).For the fat tissue ingredients,the below 40 years group ( P =0.013 ) and over 70 years group (P =0.039) showed significant differences with the controls,while the 60-69 years group (P =0.191 ) showed no such difference.ConclusionsNutritional status is different among HD patients at different ages.HD patients below 40 years and abover 70 years are more susceptible to malnutrition.Although BMI shows no difference with the normal subjects,protein wasting and increased fat tissue storage do exist in HD patients.The nutrition changes are slightest in 40-59 years old and much severer in over 70 years old patients.