We constructed different N-terminal truncated variants based on Bacillus acidopullulyticus pullulanase 3D structure (PDB code 2WAN), and studied the effects of truncated mutation on soluble expression, enzymatic properties, and application in saccharification. Upon expression, the variants of X45 domain deletion existed as inclusion bodies, whereas deletion of CBM41 domain had an effective effect on soluble expression level. The variants that lack of CBM41 (M1), lack of X25 (M3), and lack both of CBM41 and X25 (M5) had the same optimal pH (5.0) and optimal temperature (60 degrees C) with the wild-type pullulanase (WT). The K(m) of M1 and M5 were 1.42 mg/mL and 1.85 mg/mL, respectively, 2.4- and 3.1-fold higher than that of the WT. k(cat)/K(m) value of M5 was 40% lower than that of the WT. Substrate specificity results show that the enzymes exhibited greater activity with the low-molecular-weight dextrin than with high-molecular-weight soluble starch. When pullulanases were added to the saccharification reaction system, the dextrose equivalent of the WT, M1, M3, and M5 were 93.6%, 94.7%, 94.5%, and93.1%, respectively. These results indicate that the deletion of CBM41 domain and/or X25 domain did not affect the practical application in starch saccharification process. Furthermore, low-molecular-weight variants facilitate the heterologous expression. Truncated variants may be more suitable for industrial production than the WT.
Bacillus ; enzymology ; Glycoside Hydrolases ; metabolism ; Molecular Weight ; Protein Conformation ; Sequence Deletion ; Substrate Specificity ; Temperature

Objective To evaluate the effect of pitavastatin on blood glucose in patients with hypercholesterolemia,and to investigate the efficacy of pitavastatin in diabetic patients combined with hypercholesterolemia.Method This study was a 12-week,multi-center,open-label,without parallel-group comparison,phase Ⅳ clinical trail.Results Contrasting to baseline,the prevalences at week 4 and 12 post-treatment of abnormal fasting plasma glucose (FPG) and glycosylated hemoglobin Alc (HbA1c)( FPG:14.2％ vs 14.1％ and 11.0％ ; HbA1c:14.3％ vs 15.1％ and 16.1％ ) in the safety set subjects without diabetes mellitus (DM),as well as in those with DM but not taking glucose-lowering drugs (FPG:7/7 vs 4/7 and 5/7; HbAlc:5/5 vs 4/4 and 5/5) had no significant changes (all P vaules ＞0.05).Contrasting to baseline,the levels of TC [ (6.51±0.94) mmol/L vs (5.12 ±0.93) mmol/L and (4.54 ±1.00) mmol/L],LDL-C [(4.11 ±0.79)mmol/L vs (3.02 ±0.81) mmol/L and (2.51 ±0.70)mmol/L] and TG [2.10(1.53,2.54) mmol/L vs 1.62(1.26,2.00) mmol/L and 1.35(1.10,1.86)mmol/L]at week 4 and 12 post-treatment in the per protocol set 55 subjects with DM were significantly reduced (all P values ＜ 0.05 ) ; 33.3％ of subjects at high risk and 10.0％ of subjects at very high risk had achieved a TC target value; 55.6％ of subjects at high risk and 40.0％ of subjects at very high risk had achieved a LDL-C target value.Conclusion Pitavastatin has a safe effect on blood glucose and it could be used to treat diabetic patients combined with hypercholesterolemia in China.

OBJECTIVETo investigate the expression of CEA mRNA and CK(19) mRNA in peripheral blood cells of patients with lung cancer and evaluate its clinical significance.

METHODSPeripheral blood nucleated cells of 50 patients with lung cancer were studied by RT-PCR to detect the expression of CEA mRNA and CK(19) mRNA.

RESULTSThe combined positive rate of CEA mRNA and CK(19) mRNA in patients with lung cancer (82.0%) was significantly higher than that in patients with benign lung diseases (28.0%) and healthy volunteers (8.1%) (P < 0.001). The expression rate had no relation to the clinical staging or histological type. Compared with single detection, combined detection increased the detection rate but did not decrease the specificity.

CONCLUSIONCombined detection of CEA mRNA and CK(19) mRNA expression in peripheral blood nucleated cells increase the sensitivity of detecting hematogenous dissemination of cancer cells. Long-term survival analysis and more specimens would be helpful for evaluating its clinical significance.

Adenocarcinoma ; blood ; pathology ; Adult ; Aged ; Carcinoembryonic Antigen ; biosynthesis ; blood ; genetics ; Carcinoma, Small Cell ; blood ; pathology ; Carcinoma, Squamous Cell ; blood ; pathology ; Female ; Humans ; Keratins ; biosynthesis ; blood ; genetics ; Lung Neoplasms ; blood ; pathology ; Male ; Middle Aged ; Neoplasm Staging ; RNA, Messenger ; blood ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

Objective To determine kinetics of TNF-α and miR-146a (microRNA-146a)expressions in lipopolysaccharide (LPS)-induced NR8383 alveolar macrophages (AM) at different intervals and their relationships in order to explore regulatory effect and mechanism of miR-146a on alveolar macrophages inflammatory responses.Methods NR8383 alveolar macrophages were seeded in a 6-well plate,and stimulated with 1 μg/ml of LPS for 0 h,3 h,6 h and 12 h separately after 90 min.Cells were harvested and supernatant were collected 0 h,3 h,6 h and 12 h after incubation.The expressions of miR146a and TNF-α mRNA in cells were detected by real-time qPCR and the levels of TNF-α protein in the supematant of cells were assayed by enzyme-linked immunosorbent assay ( ELISA ).Pearson correlation analysis was used to analyze the correlation between miR-146a and TNF-α mRNA.Results ①The level of TNF-α protein in the supernatant of cell was significantly increased 3 h after LPS challenge ( 359.80 ±57.54) pg/ml (P ＜0.01 ),and peaked 12 h later (729.22 ±50.40) pg/ml (P＜0.01 ) ; ②the expression of TNF-α mRNA peaked 3 h after LPS challenge (67.48 ±24.52) fold,P ＜0.01 ),and then decreased gradually; ③the expression of miR-146a mRNA increased continuously until 6 h or 12 h after LPS challenge 6 h:(5.33 ±0.81) fold,12 h:(8.21 ±1.19) fold,(P＜0.01),and it showed an upward tendency;④ the expression of miR-146a mRNA was negatively correlated with TNF-α mRNA ( r =-0.895,P ＜0.01).Conclusions The miR-146a mRNA showed a negative correlation with TNF-α mRNA present in lipopolysaccharide-stimulated alveolar macrophages,suggesting miR-146a mRNA involved in regulating the inflammatory response of alveolar macrophages.

OBJECTIVETo assess the association between brain-derived neurotrophic factor (BDNF) gene Val66Met polymorphism and clinical characteristics of first episode schizophrenia in a Chinese Han population.

METHODSGenotyping of BDNF Val66Met polymorphism was carried out for 135 schizophrenic patients and 483 healthy controls with TaqMan probe technology. The patients' psychotic symptoms were assessed using the positive and negative syndrome scale (PANSS).

RESULTSA significant difference was found in genotype distribution and allelic frequency of the Val66Met polymorphism between the two groups (P< 0.01). In patients, Met homozygotes had a significantly higher score in anxiety/depression factor, cognitive factor and total score of PANSS than Val carriers.

CONCLUSIONBDNF gene Val66Met polymorphism is associated with the pathogenesis of schizophrenia. The Met/Met genotype of BDNF Val66Met variant may be a risk factor for symptoms in first episode schizophrenia patients.

Adolescent ; Adult ; Brain-Derived Neurotrophic Factor ; genetics ; China ; Female ; Gene Frequency ; Genetic Predisposition to Disease ; Genotype ; Humans ; Male ; Middle Aged ; Polymorphism, Single Nucleotide ; Schizophrenia ; genetics ; Young Adult

OBJECTIVETo observe the prevalence, awareness, treatment and control rate of hypertension and to evaluate the 10-year absolute risk of coronary heart disease(CHD) and ischemic cardiovascular disease(ICVD) in Chinese cardiovascular physicians.

METHODSA total of 4032 cardiovascular physicians (28 to 79 years old) from 386 hospitals in 31 provinces, autonomous regions and municipalities were randomly selected and received an epidemiologic survey of prevalence, awareness, and control of hypertension and evaluations of CHD and ICVD risk.

RESULTSThe prevalence of hypertension in Chinese cardiovascular physicians was 13.1%. The awareness rate of hypertension in Chinese cardiovascular physicians was 81.7%. Hypertension treatment rate was 69.6% and blood pressure control rate was 44.6%. The prevalence of hypertension was higher in male physicians than in female physicians before the age of 55 years old. Ten-year absolute risk of CHD and ICVD was 0.08 and 0.03 in hypertensive physicians compared to 0.03 and 0.01 in non-hypertensive physicians.

CONCLUSIONSThe results show suboptimal awareness, treatment and control rate in Chinese cardiovascular physicians for their own hypertension status.Physicians suffering from hypertension face higher risk for cardiovascular disease. It is therefore necessary to improve the self-monitoring of blood pressure in Chinese cardiovascular physicians.

Adult ; Aged ; Asian Continental Ancestry Group ; China ; epidemiology ; Coronary Disease ; epidemiology ; Female ; Health Knowledge, Attitudes, Practice ; Humans ; Hypertension ; epidemiology ; Male ; Middle Aged ; Myocardial Ischemia ; epidemiology ; Physicians ; Prevalence ; Risk Factors

OBJECTIVETo evaluate the outcome of precision-fit surface hemiarthroplasty in the treatment of femoral head osteonecrosis.

METHODSThe clinical data of 41 patients (48 hip joints) with femoral head osteonecrosis were reviewed. Of them, 30 were male and 11 were female, average age was 37 years old (ranging from 29 - 49). Thirty-five patients were at Ficat stage III and 13 at Ficat stage IV, their acetabula were relatively normal. The 41 patients (48 hip joints) underwent precision-fit surface hemiarthroplasties.

RESULTSThe mean duration of follow-up was 5.2 years. The average UCLA hip score at follow-up was improved significantly from 3.1 to 9.1 points for pain, 4.4 to 9.2 points for walking, and 5.5 to 7.1 points for activity (P = 0.001). The satisfaction rate was 88.6% for 35 at Ficat stage III, 69.2% for 13 at stage IV (P = 0.25). Eight hips failed; the UCLA hip score was not improved significantly; the postoperative X-ray examination showed that 7 femoral prostheses were implanted in a varus orientation (the angle between the femoral prosthesis stem and the anatomic axis of the femoral shaft was lower than angle of 130 degrees). Five-year survival rate was 83.0%.

CONCLUSIONSPrecision-fit surface hemiarthroplasty of the hip has the satisfactory result in treatment of the femoral head osteonecrosis on the basis of observing strictly operative indications and improving operative technique.

Adult ; Arthroplasty, Replacement, Hip ; methods ; Female ; Femur Head Necrosis ; surgery ; Hip Prosthesis ; Humans ; Male ; Middle Aged ; Retrospective Studies ; Treatment Outcome

OBJECTIVETo explore the relationship and interaction of elevated fasting glucose and hypertension on cardiocerebral vascular disease.

METHODS10 054 males and females were recruited for our cross-sectional study during May 2007 to August 2007. Unconditional logistic regression was used to analysis the relationship between fasting glucose and hypertension on cardiocerebral vascular disease. A product of fasting glucose and hypertension was added to the logistic regression model to evaluate the multiplicative interaction and relative excess risk of interaction (RERI), attributable proportion (AP) of interaction and synergy index (S) was applied to evaluate the additive interaction of the two factors. Bootstrap was used to calculate 95% confidence intervals (CI) of RERI, AP and S.

RESULTSAfter adjusting age, gender, smoking, drinking, body mass index (BMI) and region, the product of fasting glucose and hypertension was not statistically significant, which means there was no multiplicative interaction between the two. But the additive indexes RERI, AP and S with 95%CI of diabetes and hypertension were 0.64 (0.03, 1.25), 0.27 (0.01, 0.47) and 1.83 (1.02, 5.13) respectively, which means significant additive interaction was shown between the two on cardiovascular disease but not no stroke. And there were no additive interaction between impaired fasting glucose on cardiovascular disease or stroke.

CONCLUSIONSHypertension was independently related to cardiovascular disease and stroke in Beijing citizens, and diabetes were independently related to stroke. There was additive interaction between diabetes and hypertension on cardiovascular disease.

Adult ; Aged ; Blood Glucose ; metabolism ; Blood Pressure ; Cardiovascular Diseases ; epidemiology ; Cerebrovascular Disorders ; epidemiology ; China ; epidemiology ; Cross-Sectional Studies ; Female ; Humans ; Hypertension ; blood ; complications ; Male ; Middle Aged ; Risk Factors

OBJECTIVETo identify histopathologic changes of major organs and to correlate clinical symptoms in patients infected by avian influenza H5N1.

METHODSAutopsy study was performed in two patients died of avian influenza HSN1 infection, following conventional protocols and strict safety procedures. Tissue samples from all major organs of two cases and lung samples of one case were collected and fixed in 4% formaldehyde. Histopathologic changes were evaluated by light microscope.

RESULTSDiffuse alveolar damage (DAD) of the lung was seen in both cases. Lesions at various stages of development were seen involving different areas of the lung. At the early stages, the lungs exhibited exudative changes, including capillary congestion, necrosis of alveolar epithelial cells, and intra-alveolar edema. Hyaline membranes were prominent and diffusely distributed along alveoli. In the middle-late stages of the disease, the lungs exhibited proliferative and fibrotic changes, including proliferation of pneumocytes and bronchial epithelium, fibrosis of the interstitium and alveolar spaces. Lung biopsy tissue of one case showed DAD and interstitial fibrosis in a background of bronchiectasis. Lymph nodes and spleens showed quantity reduction of lymphocytes and active hemophagocytosis. Other changes in major organs included interstitial carditis in one case and acute renal tubular necrosis in one case. In one case, the brain showed edema with cytoplasmic eosinophilia, loss of structure, axon welling and focal necrosis around ventricle. Multiple foci of trophoblastic necrosis with dystrophic calcification were observed in placenta of one pregnant patient. Acute necrotizing deciduitis was found focally. Sections of fetal lung showed edema and scattered interstitial neutrophils were consistent with acute interstitial pneumonitis.

CONCLUSIONSThe respiratory tract is the major target of avian influenza A H5N1 virus infection. The changes of DAD in the lungs resulted in hypoxia, leading to multiple organ failure and death.

Adult ; Alveolar Epithelial Cells ; pathology ; Animals ; Birds ; Fatal Outcome ; Female ; Humans ; Influenza A Virus, H5N1 Subtype ; pathogenicity ; Influenza in Birds ; pathology ; physiopathology ; virology ; Influenza, Human ; pathology ; Male ; Pregnancy ; Pulmonary Fibrosis ; etiology ; pathology

Objective To explore the effect of hydrogen sulfide (H2S) on the expression of survivin in PC12 cells and the neuroprotective function of H2S on PC12 cells.Methods Different concentrations of sodium hydrosulfide (NaHS) were used to treat the PC12 cells at different times.Dose-effect (50-800 μmol/L) and time-effect (0-180 min) on the expression of survivin were evaluated by Western blotting.Cell viability was tested by using cell counter kit-8.Results NariS treatment at the concentrations from 50 to 200 μmol/L for 30 min could up-regulate the expression of survivin in a dose dependent manner,however,when the concentration of NariS was above that,the expression of survivin decreased gradually;when the concentration of NariS reached 800 μmoi/L,the expression level of survivin was lower than the normal level.Treatment with 400 μmol/L NariS within the range of 0-60 min could promote the expression of survivin in a time dependent manner,but with the extension of time,the expression of survivin was declined.On the other hand,400 μmol/L NaHS preconditioning could enhance the expression of survivin promoted by CoCl2 and reduce the injuries of PC12 cells induced by CoCl2 to increase the cell viability.Conclusion H2S increases the expression ofsurvivin in a dose and time dependent manners at certain degree,which may be related to the protection of PC12 cells against chemical hypoxic damage.