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 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 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.

ObjectiveTo explore the relationship of body fat distribution with serum lipid and its potentially predictive value for dyslipidemia.MethodsA total of 784 Beijing rural residents were enrolled in this study using a cluster sampling method.The body height,weight,waist circumference (WC),hip circumference,body composition,high-density lipoprotein cholesterol ( HDL-C ),low-density lipoprotein cholesterol ( LDL-C),total cholesterol ( TC),and triglycerides (TG) were measured.The body mass index (BMI) and waist to hip ratio (WHR) were calculated.ResultsThe age-adjusted partial correlation analysis showed that WC had the best correlation with HDL-C ( r =- 0.310) and LDL-C ( r =0.204 ),while WHR with TC ( r =0.151 ) and TG ( r =0.271 ).Subgroup analysis with different BMI,WC,WHR,and trunk fat mass (TFM) showed that WC,WHR,and TFM sensitively reflected the changes of body lipids,whereas BMI,WC,WHR,and TFM sensitively reflected the low HDL,high TG,and risk of dyslipidemia.Receiver operating characteristic curve analysis showed that the predictive curves of WC,WHR,BMI,and TFM were above the reference line,and the areas under the receiver operating characteristic curves of WHR (0.684,0.630),WC (0.667,0.616),and TFM (0.661,0.604) showed high tendencies than BMI (0.629,0.597) for both male and female subjects,although no statistically significant differences were found ( all P ＞ 0.05 ).ConclusionsCompared with BMI,the body fat distribution indicators including WHR,WC,and TFM have higher predictive values in evaluating the risk of dyslipidemia.When the maximum Youden index for predicting the risk of dyslipidemia is applied,the ideal cutoff points was 24 kg/m2 for BMI,0.91 for WHR,85cm for WC,7.5kg for TFM in males,and 25 kg/m2,0.91,87cm,and 9.5 kg,respectively,in females.

Objective To investigate the expressions of Na+/H+ exchanger regulatory factor 1 (NHERF1) and β-catenin in extramammary Paget's disease tissue as well as their significance.Methods Immunohistochemistry was performed to detect the protein expressions of NHERF1 and β-catenin in paraffin-embeded tissue samples from 18 patients with in situ and 22 patients with invasive extramammary Paget's disease.Results There was a high expression of NHERF1 protein in 18 (81.82％) invasive and 7 (38.89％) in situ extramammary Paget's disease samples (x2 =7.78,P ＜ 0.01).Statistical differences were observed in the membrane expression rate and cytoplasmic or nuclear expression rate of β-catenin between invasive and in situ extramammary Paget's disease tissue samples (0 (0/22) vs.33.33％ (6/18),x2 =8.63,P ＜ 0.01; 81.82％ (18/22) vs.44.44％ (8/18),x2 =6.08,P ＜ 0.05).In extramammary Paget's disease in situ tissue samples,the expression of NHERF1 was negatively correlated with the cytomembrane expression of β-catenin (ρ =-0.488,P ＜ 0.01),but positively correlated with the cytoplasmic or nuclear expression of β-catenin (ρ =0.623,P ＜ 0.01),and there was a negative correlation between the cytomembrane and cytoplasmic or nuclear expression of β-catenin (ρ =-0.572,P ＜ 0.01).Conclusions There is an abnormal expression of NHERF1 and β-catenin in extramammary Paget's disease tissue,which may be associated with the initiation,progression,and invasion of primary extramammary Paget's disease.

Objective To detect the expression of tissue inhibitor of metalloproteinase-4 (TIMP-4) in cutaneous malignant melanoma (CMM) tissue and to assess its relationship with melanoma proliferation,invasion and metastasis.Methods Western blot was conducted to measure the protein expression of TIMP-4 in five fresh lesional and paratumoral tissue specimens of CMM and three fresh tissue specimens of nevi.Immunohistochemistry was carried out to quantify the expression of TIMP-4,Ki-67,matrix metalloproteinase-2 (MMP-2),vascular endothelial growth factor (VEGF) and CD63 in paraffin-embedded tissue samples from 43 cases of CMM and 51 cases of nevi.The degree of malignancy of melanoma was evaluated in these lesions.Results Western blot analysis showed that the expression of TIMP-4 was significantly higher in 4 of 5 CMM tissue specimens than in corresponding paratumoral tissue specimens and nevus tissue specimens.Immunohistochemistry revealed that the expression rate of TIMP-4 was 86.04％ (37/43) in melanoma tissue,compared to 19.6％ (10/51) in nevus tissue (x2 =31.55,P ＜ 0.05).The expression of TIMP-4 increased sequentially from in situ melanoma to invasive and metastatic melanoma (rs =0.309,P ＜ 0.05).As far as CMM was concerned,the TIMP-4 expression was uncorrelated with any of the known prognostic variables including clinical stage,Clark level,Breslow depth,presence of ulcer,and Ki-67 expression (all P ＞ 0.05),but positively correlated with the expressions of VEGF (rs =0.345,P ＜ 0.05) and CD63 (rs =0.555,P ＜ 0.01).The median expression level of TIMP-4 was significantly higher in MMP-2-positive than in MMP-2-negative melanoma tissue samples (3 vs.0,P ＜ 0.01).Conclusions TIMP-4 protein is highly expressed in CMM tissue,which may be closely associated with the initiation and progression of CMM,especially with the metastasis of and angiogenesis in CMM.

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.

Objective To observe the changes of cell cycle on cancer cells after dihydroartemisinin and X-ray irradiation. Methods Human HeLa cells of cervical cancer with p53 mutation was used and human SiHa cells of cervical cancer with wild p53 was used as control. Flow cytometry was used to detect the effect of dihydroartemisinin (20 and 100 μmol/L) and irradiation (6 Gy)on cell cycle. Western blot was used to measure the levels of cell cycle protein. Results G2 arrest was observed in irradiated HeLa cells, which the proportion of cells in G2 phase was increased from 14.45% to 73. 58% after 6 Gy X-ray irradiation, but it was abrogated by dihydroartemisinin from 73. 58% to 48.31%in HeLa cells, and it had no change on the SiHa cells. The elevated Weel protein and the lowered Cyclin B1 protein were observed with the G2 arrest severity. The expression of radiation-induced Weel protein was suppressed and the Cyclin B1 protein was increased after dihydroartemisinin treatment, which was in accordance with the abrogation of radiation-induced G2 delay. Conclusions The main effect of irradiation on cell cycle of p53 mutated HeLa cells is G2 arrest. Dihydroartemisinin could abrogate it, which is associated with the changes of Weel protein and Cyclin B1 protein. In Siha cells, the main effect of irradiation on cell cycle is G1 arrest, and dihydroartemisinin has no effect on it.

Objective To analyze serum amyloid protein A (SAA) subtype and amino acid mutation sequence of the renal biopsy specimens from patients with renal amyloidosis secondary to ankylosing spondylitis (AS) by laser microdissection combined with mass spectometry.Methods Kidney biopsy formalin-preserved paraffin-embedded (FFPE) specimen slices were stained by Congo red,the positive areas of Congo red staining were selected by microdissection,after trypsin hydrolysis and filtration,peptide samples were subjected to liquid chromatography tandem mass spectrometry.Analysis softwares were used to evaluate the results,and the patient's amino acid sequence of SAA protein was compared to mutant amino acid sequence reported by literature or deduced from mutant SAA gene to determine whether there was a variation.Results SAA1 and SAA2 proteins with high abundance were identified by mass spectrometry,serum amyloid P and apolipoprotein E were also detected.No variation of SAA1 and SAA2 protein was detected.Conclusions The SAA1 and SAA2 proteins in AA amyloidosis secondary to ASwere identified for the first time,which enriched the pathogenesis of amyloidosis secondary to AS and provided a new method for the accurate classification of AA amyloidosis.

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.