This study was aimed at creating an engineered strain of Bacillus subtilis for efficient expression of biologically active type 2 toxin B(TcdB2)derived from a highly virulent strain of Clostridium difficile.The TcdB2 gene was cloned from ST1/RT027 strain genome DNA,incorporated into the PHT01 vector,and then transformed into B.subtilis strain WB800N for prokaryotic expression.Cell toxicity assays revealed that the recombinant TcdB2 exhibited cytotoxic effects in various cells.The engineered B.subtilis strain effectively expressed biologically active TcdB2,thus providing a basis for further exploration of the pathogenic mechanisms of highly virulent strains of C.difficile and establishing a foundation for potential vaccine can-didate targets.

Background/Aims: Despite the high efficacy of direct-acting antivirals (DAAs), approximately 1–3% of hepatitis C virus (HCV) patients fail to achieve a sustained virological response. We conducted a nationwide study to investigate risk factors associated with DAA treatment failure. Machine-learning algorithms have been applied to discriminate subjects who may fail to respond to DAA therapy. Methods: We analyzed the Taiwan HCV Registry Program database to explore predictors of DAA failure in HCV patients. Fifty-five host and virological features were assessed using multivariate logistic regression, decision tree, random forest, eXtreme Gradient Boosting (XGBoost), and artificial neural network. The primary outcome was undetectable HCV RNA at 12 weeks after the end of treatment. Results: The training (n=23,955) and validation (n=10,346) datasets had similar baseline demographics, with an overall DAA failure rate of 1.6% (n=538). Multivariate logistic regression analysis revealed that liver cirrhosis, hepatocellular carcinoma, poor DAA adherence, and higher hemoglobin A1c were significantly associated with virological failure. XGBoost outperformed the other algorithms and logistic regression models, with an area under the receiver operating characteristic curve of 1.000 in the training dataset and 0.803 in the validation dataset. The top five predictors of treatment failure were HCV RNA, body mass index, α-fetoprotein, platelets, and FIB-4 index. The accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of the XGBoost model (cutoff value=0.5) were 99.5%, 69.7%, 99.9%, 97.4%, and 99.5%, respectively, for the entire dataset. Conclusions Machine learning algorithms effectively provide risk stratification for DAA failure and additional information on the factors associated with DAA failure.

Abstract: Objective To analyze the occupational hazards of enterprises in Pingshan district of Shenzhen in 2017. Methods Occupational hazards were analyzed in 200 enterprises in Pingshan district of Shenzhen City selected using stratified Results random sampling method. A total of 24 industries were involved in the 200 enterprises. The declaration rate of ， occupational hazards was 91.5% and the exposure rate of occupational hazards among workers was 49.2%. The regular monitoring rate of occupational hazard factors in workplaces of the enterprises was 79.5%. There were 129 kinds of occupational ， ， hazard factors of which 19 factors exceeded the national occupational exposure limit accounting for 14.7%. The over standard ， ， ， ， ， ， ， ， rates of noise silica dust cotton dust methanol toluene and other dust were 28.7% 13.6% 11.8% 5.86% 0.5% and ， ， 0.4% respectively. There were 13 kinds of occupational hazard factors in the workplace of metal products industry all of which （ ） exceeded the occupational exposure limit. The exposure rate 56.7% of occupational hazard factors in workers was the highest. Conclusion ， ， The main occupational hazard factors were noise dust and chemical factor and the major occupational hazard industry was metal manufacturing in Pingshan district of Shenzhen City.

Major depressive disorder (MDD) is a common mood disorder that affects almost 20% of the global population. In addition, much evidence has implicated altered function of the gamma-aminobutyric acid (GABAergic) system in the pathophysiology of depression. Recent research has indicated that GABA receptors (GABARs) are an emerging therapeutic target in the treatment of stress-related disorders such as MDD. However, which cell types with GABARs are involved in this process is unknown. As hippocampal dysfunction is implicated in MDD, we knocked down GABARs in the hippocampus and found that knocking down these receptors in astrocytes, but not in GABAergic or pyramidal neurons, caused a decrease in immobility in the forced swimming test (FST) without affecting other anxiety- and depression-related behaviors. We also generated astrocyte-specific GABAR-knockout mice and found decreased immobility in the FST in these mice. Furthermore, the conditional knockout of GABARs in astrocytes selectively increased the levels of brain-derived neurotrophic factor protein in hippocampal astrocytes, which controlled the decrease in immobility in the FST. Taken together, our findings contribute to the current understanding of which cell types expressing GABARs modulate antidepressant activity in the FST, and they may provide new insights into the pathological mechanisms and potential targets for the treatment of depression.

Astrocytes are the most abundant cell type in the central nervous system (CNS). They provide trophic support for neurons, modulate synaptic transmission and plasticity, and contribute to neuronal dysfunction. Many transgenic mouse lines have been generated to obtain astrocyte-specific expression of inducible Cre recombinase for functional studies; however, the expression patterns of inducible Cre recombinase in these lines have not been systematically characterized. We generated a new astrocyte-specific Aldh1l1-CreER knock-in mouse line and compared the expression pattern of Cre recombinase between this and five widely-used transgenic lines (hGfap-CreER from The Jackson Laboratory and The Mutant Mouse Resource and Research Center, Glast-CreER, Cx30-CreER, and Fgfr3-iCreER) by crossing with Ai14 mice, which express tdTomato fluorescence following Cre-mediated recombination. In adult Aldh1l1-CreER:Ai14 transgenic mice, tdTomato was detected throughout the CNS, and five novel morphologically-defined types of astrocyte were described. Among the six evaluated lines, the specificity of Cre-mediated recombination was highest when driven by Aldh1l1 and lowest when driven by hGfap; in the latter mice, co-staining between tdTomato and NeuN was observed in the hippocampus and cortex. Notably, evident leakage was noted in Fgfr3-iCreER mice, and the expression level of tdTomato was low in the thalamus when Cre recombinase expression was driven by Glast and in the capsular part of the central amygdaloid nucleus when driven by Cx30. Furthermore, tdTomato was clearly expressed in peripheral organs in four of the lines. Our results emphasize that the astrocyte-specific CreER transgenic lines used in functional studies should be carefully selected.

Objective To observe the clinical efficacy and safety of sorafenib tablets combined with transcatheter arterial chemoembolization (TACE) in the treatment of unresectable liver cancer. Methods A total of 164 patients with unresectable liver cancer were randomly divided into control and treatment groups with 82 cases per group. Control group was treated with TACE alone, once every 4 weeks. Treatment group was given sorafenib tablets 400 mg per time from 5 d after TACE treatment, bid, orally, on the basis of control group. Two groups were treated for 12 weeks. The clinical efficacy, serum tumor markers, serum vascular endothelial growth factor (VEGF) , levels of basic fibroblast growth factor (bFGF) , and adverse drug reactions were compared between two groups.Results After treatment, the objective remission rates of treatment and control groups were 52. 44％ (43 cases/79 cases) and 28. 05％ (23 cases/79 cases) , the disease control rates were 87. 80％ (72 cases/79 cases) and 68. 29％ (56 cases/79 cases) , the progression free survival time were (15. 32 ± 2. 04) and (10. 83 ± 1. 43) months, the overall survival time were (15. 32 ± 2. 04) and (10. 83 ± 1. 43) months, the differences were statistically significant (all P < 0. 05) . After treatment, the alpha fetoprotein of treatment and control groups were (71. 38 ± 10. 04) and (152. 36 ± 20. 37) ng·m L-1, the carcinoembryonic antigen were (2. 02 ± 0. 27) and (2. 94 ± 0. 34) μg·L-1, the VEGF were (317. 87 ± 32. 76) and (442. 45 ± 35. 09) pg·m L-1, the differences were statistically significant (all P < 0. 05) . The adverse reactions of treatment group and the control group were nausea and vomiting (71. 95％ vs63. 41％) , diarrhea (35. 37％ vs 42. 68％) , myelosuppression (43. 90％ vs 40. 24％) and fever (84. 15％ vs90. 24％) , oral mucositis (32. 93％ vs 6. 10％) , hand-foot skin reaction (69. 51％ vs 2. 44％) , the differences were statistically significant (all P < 0. 05) . Conclusion Sorafenib tablets combined with TACE have a definitive clinical efficacy in the treatment of unresectable liver cancer, which can effectively inhibit the release of tumor markers, decrease the levels of serum VEGF and other cytokines. Although the incidence of adverse drug reactions is high, they can be controlled.

OBJECTIVETo assess the quality of whole spine images obtained by DR and magnetic resonance imaging (MRI) and analyze the whole spinal imaging sagittal parameters for standing DR and supine MRI.

METHODSSixty-one patients aged 49.9∓17.6 years with degenerative spinal disease underwent both standing DR and supine MRI of the whole spine from November, 2010 to March, 2016. The image quality was retrospectively reviewed, and the cervical lordosis (CL), thoracic kyphosis (TK), lumbar lordosis (LL), sacral slope (SS), and sagittal vertical axis (SVA) were measured on the whole spinal lateral DR and middle sagittal MR images.

RESULTSBoth the DR and MR whole spine images had a high quality (100%). The CL, TK, LL, SS, and SVA measured were 28.37mnplus;10.91 °, 29.98mnplus;8.96 °, 45.61mnplus;12.46 °, 34.38mnplus;9.05 °, and 17.20mnplus;26.39 mm on DR images and were 24.34mnplus;9.01 °, 21.22mnplus;8.13 °, 41.45mnplus;12.17 °, 37.45mnplus;8.19 °, and 36.51mnplus;12.44mm on MR images, respectively, showing significant differences in the measurements between the two modalities (P=0.000, 0.000, 0.000, 0.001, and 0.007, respectively). The correlation coefficient between DR and MR images for CL, TK, LL, SS, and SVA were 0.69, 0.68, 0.72, 0.51, and 0.27 (P=0.000, 0.000, 0.000, 0.000, and 0.034, respectively).

CONCLUSIONBoth standing DR and supine MR whole spine imaging can provide high-quality images. The CL, TK, LL, SS, and SVA measured on supine MR whole spine images are correlated with those on standing DR images but differ obviously. Supine MR imaging can not substitute standing DR examinations, and comprehensive assessment of degenerative spinal disease needs the combination of the two imaging techniques.

BACKGROUNDTalaromyces (Penicillium) marneffei (TM) is an emerging dimorphic human pathogenic fungus that is endemic to Southeast Asia. TM mostly occurs as an opportunistic infection in patients with human immunodeficiency virus (HIV). The objective of this study was to compare the clinical and laboratory parameters of patients with TM infections who were HIV-positive and HIV-negative and to assess therapies and outcomes.

METHODSThis was a retrospective analysis of 26 patients diagnosed with disseminated TM infection from September 2005 to April 2014 at Fujian Provincial Hospital, China.

RESULTSPatients with TM infection tend to present with fever, weight loss, and anemia. The time from symptom onset to confirmed diagnosis was greater for HIV-negative patients (n = 7; median: 60 days, range: 14-365 days) than for HIV-positive patients (n = 19; median: 30 days, range: 3-90 days, Mann-Whitney U = 31.50, P= 0.041). HIV-negative patients were more likely to have dyspnea (57.1% vs. 5.3%, χ2 = 8.86, P= 0.010), low neutrophil count (Mann-Whitney U = 27.00, P= 0.029), high CD4 count (Mann-Whitney U = 0.00, P= 0.009), and high lymphocyte count (Mann-Whitney U = 21.00, P= 0.009). There were no significant differences in other demographic, clinical, or biochemical characteristics. Among all the patients, 12 HIV-positive patient and 1 HIV-negative patient received amphotericin and fluconazole treatment, 9 of whom improved, 1 died, 2 had kidney damage, 1 had hypokalemia due to exceeded doses.

CONCLUSIONSHIV-negative patients with TM infections tend to have a longer diagnostic interval, a higher percentage of dyspnea, higher levels of CD4 and lymphocytes, and lower neutrophil counts than TM infection in HIV-positive patients. Treatment programs with amphotericin and fluconazole are mostly effective.

Adult ; Aged ; CD4 Lymphocyte Count ; Female ; HIV Infections ; complications ; Humans ; Male ; Middle Aged ; Mycoses ; diagnosis ; drug therapy ; immunology ; Retrospective Studies ; Talaromyces ; drug effects

OBJECTIVETo evaluate the effect of post-treatment PSA kinetics on the prognosis of prostate cancer (PCa).

METHODSWe retrospectively reviewed the clinical data of 114 cases of locally advanced PCa treated by maximal androgen blockade (MAB) combined with brachytherapy, and analyzed the association of the changes in PSA kinetics with the prognosis of the patients.

RESULTSThe median survival time of the patients was 81 (15 - 144) months, with 1-, 3- and 5-year survival rates of 91. 23%, 78.07% and 68.42% , respectively. Univariate analysis indicated that the baseline PSA level, PSA nadir, the time of PSA decreasing to nadir, PSA doubling time, and the extent of PSA declining were all predictive factors for the survival time of the PCa patients. Multivariate analysis demonstrated that PSA nadir, the time of PSA decreasing to nadir, and the extent of PSA declining were three independent prognostic factors, which prolonged the long-term survival of the patients by 1.7, 3.2 and 6.8 times, respectively.

CONCLUSIONFor locally advanced PCa treated by MAB combined with brachytherapy, PSA nadir <1 micro g/L, the time to nadir <3 months, and the extent of PSA declining >96% are independent prognostic factors.

Aged ; Aged, 80 and over ; Androgens ; administration & dosage ; therapeutic use ; Brachytherapy ; Humans ; Male ; Middle Aged ; Prognosis ; Prostate-Specific Antigen ; metabolism ; Prostatic Neoplasms ; metabolism ; therapy ; Retrospective Studies

OBJECTIVETo establish a rabbit model of intervertebral disc degeneration by puncturing the anulus fibrosus through an approach between the longissimus dorsi muscle and obliquus externus abdominis.

METHODSThe L(4/5) and L(5/6) intervetebral discs of 6 New Zealand white rabbits were punctured by an 18-gauge pin in the anterolateral annular fibrosus through an approach between the longissimus dorsi muscle and the obliquus externus abdominis with the right transverse processes of L(5) and L(6) resected; the L(2/3) discs were used as the control without exposure or needle stab, and the L(3/4) discs were subjected to sham operation with the discs exposed but not punctured after resecting the right transverse process of L(4). X-ray and magnetic resonance imaging (MRI) were performed preoperatively and at the 4th week after puncture. At 4 weeks after the operation, histological and immunohistochemical analyses of the discs were carried out.

RESULTSX-ray of the punctured discs at 4 weeks after the operation presented a significant decrease of disc height, osteophytosis formation, and end-plate stiffness; an obvious decrease of signal intensity on T(2)-weighted images was found in the puncture group but not in the control or sham-operated groups. Gross morphological inspection showed atrophy of the nucleus pulposus, which became loose, soft, and fragile with a light yellow color. Histological and immunohistochemical analyses showed a significant decrease of notochordal cells and type II collagen in the nucleus pulposus in the puncture group as compared to the control and sham-operated groups.

CONCLUSIONPuncture through the approach between the longissimus dorsi muscle and the obliquus externus abdominis allows the establishment of a reliable animal model for studying intervertebral disc degeneration.

Animals ; Disease Models, Animal ; Female ; Intervertebral Disc Degeneration ; Lumbar Vertebrae ; physiopathology ; Male ; Rabbits