Talaromyces marneffei is the only dimorphic species in its genus and causes a fatal systemic mycosis named talaromycosis. Our previous study indicated that knockdown of AcuD gene (encodes isocitrate lyase of glyoxylate bypass) of T. marneffei by RNA interference approach attenuated the virulence of T. marneffei, while the virulence of the AcuD knockout strains was not studied. In this study, T. marneffei-zebrafish infection model was successfully established through hindbrain microinjection with different amounts of T. marneffei yeast cells. After co-incubated at 28°C, the increasing T. marneffei inoculum doses result in greater larval mortality; and hyphae generation might be one virulence factor involved in T. marneffei-zebrafish infection. Moreover, the results demonstrated that the virulence of the ΔAcuD was significantly attenuated in this Zebrafish infection model.
Gene Knockout Techniques ; Hyphae ; Isocitrate Lyase ; Microinjections ; Mortality ; Rhombencephalon ; RNA Interference ; Talaromyces ; Virulence ; Yeasts ; Zebrafish

Cells, Cultured ; Cyclic Nucleotide Phosphodiesterases, Type 2 ; antagonists & inhibitors ; metabolism ; Fibroblasts ; drug effects ; metabolism ; Glucosides ; pharmacology ; Humans ; Lung ; cytology ; embryology ; Phenols ; pharmacology ; Phosphodiesterase Inhibitors ; pharmacology ; Tumor Suppressor Protein p53 ; metabolism

OBJECTIVE: To clone the promoter sequence of acute monocytic leukemia new antigen gene.MLAA-34 and identify its promoter core region. METHODS: The full-length fragment of MLAA-34 gene promoter region was amplified by PCR, then was ligated into pGL3-Basic vector, and the recombinant plasmid was cloned. Constructed a series of MLAA-34 gene promoter 5' flanking region truncated plasmid. These recombinant plasmids were transfected into U937 and HEK293 cells, and the dual luciferase reporter gene was used to detect the promoter activity of each fragment to determine the minimum active region. Transcription factor binding sites were analyzed by bioinformatics methods. RESULTS: The recombinant plasmid containing MLAA-34 promoter sequence and its truncated plasmid were successfully constructed, and the promoter activity was significantly increased as compared with the empty vector (P＜0.001). The minimal active region of MLAA-34 located between 402 bp and 200 bp. It contained multiple transcription factor binding sites such as E2F1, MZF-1, SP1, USF2 and STAT3. CONCLUSION The promoter of luciferase reporter gene has been successfully constructed with different deletion fragments of MLAA-34, and its core promoter region may contain multiple transcription factor sequence.
Adult ; Antigens, Neoplasm ; genetics ; Apoptosis Regulatory Proteins ; genetics ; Cloning, Molecular ; Genes, Reporter ; HEK293 Cells ; Humans ; Leukemia, Monocytic, Acute ; genetics ; Luciferases ; Promoter Regions, Genetic

A UPLC method has been developed for simultaneous determination of nine furanocoumarins of Angelica dahurics,and was used for quality evaluation of A. dahurica from different habitats. ACQUITY UPLC BEH C18 chromatographic column was employed,the separation was performed with the mobile phase consisting of acetonitrile and water,and the detection wavelength was set at254 nm. This method was used to simultaneously determine the content of xanthotoxol,oxypeucedaninhydrate,byak-angelicin,psoralen,xanthotoxin,bergapten,oxypeucedanin,imperatorin and isoimperatorin in A. dahurica from different habitats. Then,the further quality assessment of the drug was carried out by similarity evaluation,cluster analysis( CA),principal component analysis( PCA),and orthogonal partial least squares discriminant analysis( OPLS-DA). The content order of measured furanocoumarins from high to low was: oxypeucedanin>imperatorin>isoimperatorin>oxypeucedaninhydrate>bergapten>byak-angelicin>xanthotoxin>xanthotoxol>psoralen,with the mean content 2. 844,1. 277,0. 649 2,0. 216 2,0. 129 8,0. 062 68,0. 052 68,0. 019 30,0. 018 19 mg·g-1,respectively. There were difference between the batches of the drug,and the quality was influenced by smouldering sulphur based on the results of chemical pattern recognition and content determination. Finally,six active ingredients were recognized as the quality makers using OPLS-DA method. The validated UPLC fingerprint combined with chemical pattern recognition method can be used in the quality control and evaluation of A. dahurica.
Angelica ; chemistry ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; standards ; Ecosystem ; Furocoumarins ; analysis ; Quality Control

The expert consensus on clinical practice of Chinese patent medicines shall be regarded as the same with clinical practice guidelines,which shall be concise,readable and applicable. Therefore,a formal expert consensus is unable to contain the whole compilation process and the content of its important process documents. It is necessary to write instructions for continuation,expansion and publicity. In 2018 Standardization Department of China Association of Chinese Medicine invited methodologists with the background of evidence-based medicine drafted a series of standards for expert consensus on clinical practice of Chinese patent medicines,including the guidance for compilation the instructions. It will be useful for readers of expert consensus to understand the work behind the compilation,so as to make the compilation process more transparent and standardized. We proposed nine requirements for compilation the instructions,namely content and requirements,work overview,main technical content,compilation process,process and basis for handling major disagreements,expert consensus implementation requirements and measure suggestions,other issues need to be explained,appendix and format for instructions.
China ; Consensus ; Evidence-Based Medicine ; Medicine, Chinese Traditional ; standards ; Nonprescription Drugs ; standards

In 2018,Standardization Department of China Association of Chinese Medicine invited methodologists with the background of evidence-based medicine to discuss and draft a series of standards for expert consensus on clinical practice of Chinese patent medicines. These standards have been made by reference to the published standards for developing expert consensus and clinical practice guidelines. These standards were made based on full consideration of the current evidence status and the history of clinical practice of Chinese patent medicines. These standards were composed of four parts,namely information summary items,normative general items,normative technical items and information supplementary items,including cover,content,preface,introduction,title,scope,basic Information of Chinese patent medicine,suggestions for clinical application,safety,conflict of interest,appendix,and references,so as to provides reference for improving the quality of expert consensus-based compilation and enhancing the applicability of expert consensus.
China ; Consensus ; Evidence-Based Medicine ; Humans ; Medicine, Chinese Traditional ; standards ; Nonprescription Drugs ; standards

OBJECTIVE: The aim of this study was to determine whether low calf circumference (CC) could predict nutritional risk and the cutoff values of CC for predicting nutritional risk in hospitalized patients aged ⪖ 80 years. METHODS: A total of 1,234 consecutive patients aged ⪖ 80 years were enrolled in this study. On admission, demographic data, CC, and laboratory parameters were obtained. Patients with Nutritional Risk Screening 2002 (NRS-2002) total score ⪖ 3 were considered as having nutritional risk. RESULTS: CC values were significantly lower in patients with nutritional risk compared to those in patients without nutritional risk [27.00 (24.50-31.00) vs. 31.00 (29.00-33.50], P < 0.001]. CC was negatively correlated with age and nutritional risk scores. Logistic regression analysis of nutritional risk revealed that body mass index, albumin level, hemoglobin level, cerebral infarction, neoplasms, and CC (OR, 0.897; 95% confidence interval, 0.856-0.941; P < 0.001) were independent impact factors of nutritional risk. Nutritional risk scores increased with a decrease in CC. In men, the best CC cutoff value for predicting nutritional risk according to the NRS-2002 was 29.75 cm. In women, the cutoff value was 28.25 cm. CONCLUSION CC is a simple, noninvasive, and valid anthropometric measure to predict nutritional risk for hospitalized patients aged ⪖ 80 years.
Aged, 80 and over ; Anthropometry ; methods ; China ; Female ; Hospitalization ; statistics & numerical data ; Humans ; Leg ; anatomy & histology ; Male ; Nutritional Status ; Risk Assessment ; methods

OBJECTIVE: By comparing the clinical efficacy of short-segment and long-segment fixation for single-segment thoracic and lumbar spine III stage Kümmell disease to explore a more suitable fixed segment for the disease. METHODS: The clinical data of 46 patients with single-segment thoracic and lumbar spine III stage Kümmell disease treated from July 2013 to December 2016 were retrospectively analyzed. Forty-six patients were divided into short-segment fixation group(one vertebra above and below the diseased vertebra) and long-segment fixation group(two vertebrae on the upper and lower of the diseased vertebra) according to different methods of cement stick fixation. There were 25 patients in the short-segment fixation group, including 9 males and 16 females, with an average age of (75.3±4.5) years old, lumbar spine bone mineral density T-value of (-3.1±0.3) g/cm³, follow-up time of (13.0±2.3) months; there were 21 patients in long-segment fixation group, 6 males and 15 females, with an average age of (74.5±3.9) years old, lumbar spine bone mineral density T-value of (-3.2±0.3) g/cm³, follow-up time of (14.7±3.6) months.The gender, age, follow-up time, operation time, intraoperative blood loss, cement leakage, and the rate of adjacent vertebrae fractures were compared between two groups, as well as pain VAS score, ODI, and kyphosis angle before and after surgery. RESULTS: There were no significant differences in age, gender, bone density, pain VAS score, ODI, and kyphosis between two groups before surgery. The operation time and intraoperative blood loss of short-segment fixation group were less than that of long-segment fixation group. The pain VAS score, ODI and kyphosis of the two groups were significantly improved at 7 days after the operation and at the latest follow-up, there was no significant difference between two groups. There were no significant differences in bone cement leakage(9/25 vs 11/21) and adjacent vertebrae fractures(4/25 vs 3/21). CONCLUSIONS Both long-segment fixation and short-segment fixation can effectively relieve pain, correct kyphosis, improve functional index, and achieve better clinical results, but short-segment fixation has less operation time and less intraoperative blood. So single-segment thoracic and lumbar spine III stage Kümmell disease does not need to extend the fixed segment, short-segment fixation is more in line with clinical needs and worthy of further study.
Aged ; Female ; Fracture Fixation, Internal ; Humans ; Kyphosis ; Lumbar Vertebrae ; Male ; Retrospective Studies ; Spinal Fractures ; Thoracic Vertebrae ; Treatment Outcome

OBJECTIVE: To evaluate the performance of the chemiluminescence immune assay (CLIA) and the electro-chemiluminescence immuneoassay(ECLIA) for Treponemapallidum antibody（anti-TP） screening in blood donors. METHODS: The sero-panel samples from NCCL were tested with ELISA, CLIA and ECLIA assays synchronously to evaluate their performances respectively. RESULTS: The sensitivity and the negative predictive value of the CLIA were 100%, which were the same as one kind of ELISA, and better than the other ELISA; The specificity of the CLIA was 88.46%, the accuracy rate was 97.02%, the positive predictive value was 96.13%, which were higher than both ELISA. Due to the significant interference of sample heat inactivation in ECLIA detection, the result can not demonstrate the true performance of ECLIA in this study. The preliminary result was as follows: the sensitivity was 98.93%, the negative predictive value was 96.75%, and the accuracy rate, specificity and positive predictive value of ECLIA were 97.02%, 91.54% and 97.10% respectively. CONCLUSION Compared with ELISA, the CLIA has higher sensitivity and specificity and can be used for Treponemal antibody screening in blood bank. Unfortunately, the data in this study cannot come to a conclusion for ECLIA and needs more testing.
Blood Donors ; Enzyme-Linked Immunosorbent Assay ; Humans ; Luminescence ; Luminescent Measurements ; Sensitivity and Specificity

OBJECTIVE: To investigate the effect and molecular mechanism of interferon-α (INF-α) on the apoptosis of the mouse podocyte cell line MPC5 induced by hepatitis B virus X (HBx) protein. METHODS: MPC5 cells were transfected with the pEX plasmid carrying the HBx gene. RT-PCR was used to measure the mRNA expression of HBx at different time points. MPC5 cells were divided into 4 groups: control group (MPC5 cells cultured under normal conditions), INF-α group (MPC5 cells cultured with INF-α), HBx group (MPC5 cells induced by HBx), and HBx+INF-α group (MPC5 cells induced by HBx and cultured with INF-α). After 48 hours of intervention under different experimental conditions, flow cytometry was used to measure the apoptosis of MPC5 cells, and quantitative real-time PCR and Western blot were used to measure the mRNA and protein expression of slit diaphragm-related proteins (nephrin, CD2AP, and synaptopodin) and the cytoskeleton-related protein transient receptor potential cation channel 6 (TRPC6). RESULTS: MPC5 cells transfected by pEX-HBx had the highest expression of HBx mRNA at 48 hours after transfection (P<0.05). Compared with the control, INF-α and HBx+INF-α groups, the HBx group had a significant increase in the apoptosis rate of MPC5 cells (P<0.05). Compared with the control and INF-α groups, the HBx group had significant reductions in the mRNA and protein expression of nephrin, synaptopodin, and CD2AP and significant increases in the mRNA and protein expression of TRPC6 (P<0.05). Compared with the HBx group, the HBx+INF-α group had significant increases in the mRNA and protein expression of nephrin, synaptopodin, and CD2AP and significant reductions in the mRNA and protein expression of TRPC6 (P<0.05). CONCLUSIONS INF-α can inhibit the apoptosis of podocytes induced by HBx, possibly through improving the abnormal expression of slit diaphragm-related proteins (CD2AP, nephrin, and synaptopodin) and cytoskeleton-related protein (TRPC6) induced by HBx.
Animals ; Apoptosis ; Hepatitis B virus ; Interferon-alpha ; Mice ; Podocytes ; Trans-Activators