Objective To observe the changes of extracellular matrix(ECM) production and hypertrophy of glomerular mesangial cells(GMCs) induced by high glucose(HG),and the inhibitory role of myriocin(ISP-1).Methods GMC cultured with normal glucose(5.6 mmol/L D-Glucose,NG),HG(25.2 mmol/L D-Glucose) and HG plus ISP-1(100 mg/L) for different durations(0,24,48,(72 h)).The sizes of GMC were indicated by forward scatter intensity,measured by flow cytometery,and the levels of fibronection(FN),collagen Ⅳ(Col Ⅳ),laminin(LN),precollagen Ⅲ(Pcol Ⅲ) and hyaluronic acid(HA) in the supernatant of cultured GMC were detec-(ted) by ELISA.Results Compared with NG,HG could induce GMC hypertrophy(P

Adult ; Antidotes ; administration & dosage ; therapeutic use ; Female ; Humans ; Insecticides ; poisoning ; Parathion ; poisoning ; Poisoning ; drug therapy ; Pralidoxime Compounds ; administration & dosage ; therapeutic use ; Treatment Outcome

Objective To compare the sub-cellular proteome of isoniazid ( INH)-resistant Mycobacterium tuberculosis (MTB) with that of sensitive strains for identifying of unique proteins of these strains and discussing their preliminary application in clinical diagnosis. MethodsProteins of cell wall and membrane of 5 INH-resistant strains and 5 INH-sensitive strains were extracted by density gradient centrifugation.The extracts were subsequently analyzed using weak cation exchange (WCX) liquid chromatography ( LC )followed reverse phase (RP) liquid chromatography to compare the sub-cellular protein patterns. A total of 1280 fractions were collected and identified by matrix assisted laser desorption ionization-time of flight-tandem mass spectrometry (MALDI-TOF MS/MS). The Database for Annotation, Visualization and Integrated Discovery (DAVID) was used for cell component and biological process analysis. Normalized Spectral Abundance Factors (NASF) was used for semi-quantity of protein expression. 5 proteins significantly up-regulated in INH-resistant strains and 2 proteins significantly up-regulated in INH-sensitive strains were selected for ELISA analysis with autologous sera respectively. Results A total of 347 proteins were identified. Cell component analysis showed that 58％ proteins were cells well or membrane proteins. Biological process analysis showed that 31％ proteins involved in carboxylic/monocarboxylic acid biosynthetic and metabolic process, 26％ and 15％ proteins involved in organic acid or fatty acid biosynthetic and metabolic process,while 28％ proteins involved in lipid biosynthetic , metabolic, transport and localization process. O-succinylbenzoate synthase, monooxygenase, hypothetical protein Rv2255c, nicotinate-nucleotide--dimethylbenzimidazole phosphoribosyltransferase and membrane phosphatidate cytidylyltransferase cdsA were up-regulated in INH-resistant strains and fractions contained these proteins could elicit specific antibody response with autologous sera. The A450 was higher than that with INH-sensitive sera. The differences between the INH-resistant sera and the INH-sensitive sera were significant ( t = 0.028, 0.044, 0.066, 0.064, 0.083, all P＜0.01 ). Chain A of Rv2002 Gene Product and Chain A of Crystal Structure Of Rv2632c were up-regulated in INH-sensitive strains and fractions contained these proteins could elicit specific antibody response with autologous sera. The A450 was higher than that with INH-resistant sera. The differences between the INH-sensitive sera and the INH-resistant sera were significant (t=0.053, 0.073, both P＜0.05). ConclusionThe combination of density gradient centrifugation and 2D-LC MS/MS technology is useful in enrichment and identification of differential expressed proteins between INH-resistant and INH-sensitive strains at sub-cellular level. It is useful in finding antigens associated with INH-resistant MTB infection, which may prove useful for further study in the mechanism of INH resistant, as well as interaction between MTB and host.

Objective:To study the combined use of endoscopic balloon dilation with endoscopic biliary brushings in diagnosis of bile duct strictures.Methods:A prospective single center study was conducted at the Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology. All patients with suspected malignant bile duct strictures shown on CT or MRI imaging from January 2018 to January 2020 were reviewed. All patients gave informed consent to the endascopic retrograde cholangiopancreatography procedures. Their clinical and follow-up data were analyzed. All patients underwent endoscopic balloon dilation of bile duct strictures. Before and after balloon dilation, biliary brush cytology was performed, and the results were used to classify the patients into the control group and the experimental group. Pathological examination of the brush cytology samples was carried out by a single chief pathologist. Presence of cancer cells or significant heterogeneous cells indicated a positive brush cytology test. Negative patients who still highly consider cholangiocarcinoma and agree to surgery and whose gross specimen is confirmed to be malignant after surgery should be considered as false negative by brush examination; it is difficult to judge that patients with cholangiocarcinoma have progress after 2 months of follow-up should be considered as false negative by brush examination. Any progression of disease indicated that the brush test was wrong and the test was again classified as false negative. Only when there was no progression of strictures was the possibility of a benign biliary stricture being considered. The advantage test (McNemar test) was used to analyze the difference between the two diagnostic methods.Results:Of 39 patients who were included in this study, there were 26 males and 13 females, with an age of (68.0 ± 5.2) years. Cholangiocarcinoma was diagnosed by histopathology, surgery or at 2 months follow-up in 35 patients. In the control group, 17 patients had a positive brush test (sensitivity rate was 48.6%, 17/35). In the experimental group, 26 patients had a positive brush test (sensitivity rate was 74.2%, 26/35). In addition, 2 patients in the control group had a positive brush test, while in the experimental group, a negative brush test. A total of 28 patients were positive in the two groups. The sensitivity rate of the brush test was 80.0% (28/35). There were significant differences between the two groups ( P<0.05). Conclusion:Endoscopic balloon dilation combined with endoscopic biliary brushings improved the sensitivity of pathological diagnosis of cholangiocarcinoma, and endoscopic biliary brushings before and after balloon dilation improved the sensitivity of diagnosis.

OBJECTIVETo study the effects of Jisuikang (Chinese characters) on Nogo-NgR gene expression, and to explore the protective effects and mechanism of Jisuikang (Chinese characters) on spinal cord injury in rats.

METHODSOne hundred eighty female rats were randomly assigned to 6 groups(30 rats per group). Sham group: T10 lamina was resected only and spinal cord was untreated. Model group: spine cord injury (SCI) was created with a modified impinger of Allen's by impacting on the T10 spinal cord. Prednisolone group: Prednisolone (0.06 g/kg) was given by intragastric administration at a time interval of 24 hours after operation. The Jisuikang (Chinese characters) high, moderate and low dose groups: Jisuikang (Chinese characters) was supplied with different dose (50 g/kg, 25 g/kg, 12.5 g/kg) by intragastric administration in rats after operation,for the first time at 30 min after surgery. Animals were killed 3, 7, 14 days after surgery. The expression levels of Nogo-A and NgR were observed by Western Blot and Real-time PCR.

RESULTSThe expression of Nogo-A and NgR was at the basic level at all time points in sham group. Compared with model group, the protein expression levels of Nogo-A and NgR in sham, prednisolone, Jisuikang (Chinese characters) moderate dose groups were statistically significant at all time points (P < 0.05). No difference was found in Jisuikang (Chinese characters) high and low dose groups (P > 0.05). Three days after surgery, the mRNA levels of Nogo-A and NgR in treatment group were significantly lower than that in model group (P < 0.01); 7 days after surgery,Nogo-A and NgR mRNA expression were dramatically upregulated and peaked; 14 days after operation, the expression was decreased, but still significantly higher than that in other treatment groups (P < 0.01). Prednisolone and Jisuikang (Chinese characters) moderate dose groups showed the most significant effects among all groups,but there was no statistically significant difference between two groups (P > 0.05).

CONCLUSIONThe decoction Jisuikang (Chinese characters) can promote the nerve cell regeneration by regulating Nogo-A and NgR gene expression, activating Nogo- NgR signaling pathways after acute spinal cord injury.

Animals ; Female ; GPI-Linked Proteins ; analysis ; genetics ; physiology ; Medicine, Chinese Traditional ; Myelin Proteins ; analysis ; genetics ; physiology ; Nerve Regeneration ; drug effects ; Nogo Proteins ; Nogo Receptor 1 ; Rats ; Rats, Sprague-Dawley ; Receptors, Cell Surface ; analysis ; genetics ; physiology ; Signal Transduction ; drug effects ; Spinal Cord Injuries ; drug therapy ; metabolism

OBJECTIVEMyriocin (ISP-1) is a new type of immune inhibitor extracted from cordyceps sinensis. This study was to observe the effects of ISP-1 on the expression of cell cycle regulatory protein D1 (cyclinD1) in high glucose-induced hypertrophy rat glomerular mesangial cells (GMCs).

METHODSRat GMCs were cultured in vitro and divided into three groups: high glucose (450 mg/dL D-glucose), normal glucose (100 mg/dL D-glucose, control) and ISP-1 (450 mg/dL D-glucose plus 100 μg/mL ISP-1). The protein expression of cyclinD1 was detected by flow cytometry.

RESULTSThe expression of cyclinD1 in GMCs in the high glucose group increased significantly in a time-dependent manner compared with that in the control group. ISP-1 treatment significantly inhibited the up-regulated expression of cyclinD1 induced by high concentration glucose, and the expression of cyclinD1 was restored to the level of the control group 48 and 72 hrs after ISP-1 treatment.

CONCLUSIONSHigh concentration of glucose can up-regulate the expression of cyclinD1 in GMCs. ISP-1 may inhibit the up-regulated expression of cyclinD1, which might contribute to the protective effect of ISP-1 against GMC hypertrophy induced by high glucose.

Animals ; Cyclin D1 ; analysis ; Fatty Acids, Monounsaturated ; pharmacology ; G1 Phase ; Glucose ; toxicity ; Hypertrophy ; Mesangial Cells ; chemistry ; pathology ; Rats ; Rats, Sprague-Dawley

H2S has a role of protecting neurons from ischemia-reperfusion injury and significantly reducing the cerebral infarction area, but high concentration of H2S can induce neurotoxicity. Memantine, a N-methyl-D-aspartic acid(NMDA)receptor antagonist, could reduce the neurotoxicity of H2S at high concentration. Nine novel structures(compounds I1-I9)were designed by coupling(4-hydroxy phenyl)-3H-1, 2-dithiole-3-thione(ADT-OH)with memantine through alkanes as linkers and synthesized by four-step reactions from ADT-OH. Their neuroprotection against damage induced by glutamate on HT-22 cells was evaluated by MTT method. The results indicated that these compounds markedly increased the survival rates of damaged HT-22 cells at the concentration of 1 μmol/L(P< 0. 01), which suggested that these compounds could preferably protect neurons against damage induced by glutamate.

OBJECTIVETo select sub-clinical patients with symptoms of knee osteoarthritis (KOA) without X-ray changes by measuring the serum level of cartilage oligomeric matrix protein (COMP) with ELISA, so as to diagnose and treat patients with knee osteoarthritis at early stage.

METHODSThe 115 patients with KOA or with symptomatic primary KOA were enrolled from August 2007 to September 2009, which was OA group; and 35 healthy people in the control group. In OA group, there were 55 males and 60 females,ranging in age from 39 to 76 years, with an average of (55 +/- 13.32) years; the body mass index (BMI) ranged from 15.1 to 29.8; the disease course ranged from 6 to 60 months. In the control group, there were 16 males and 19 females, ranging in age from 36 to 77 years, with an average of (53 +/- 12.53) years; the BMI ranged from 14.8 to 29.2. Patients with symptomatic primary knee OA of Kellgren-Lawrence (K-L) grade I-IV were evaluated. Serum level of COMP and its correlation with OA grade were analyzed by ELISA method. The patients were treated with Celecoxib capsules. The patients in OA group were followed up, and the duration ranged from 24 to 38 months (averaged, 33.4 months), and the serum level of COMP were analyzed before and after treatment.

RESULTSThe serum level of COMP in the control group varied with age (t= 2.50, P=0.02). The serum level of COMP did not correlate with gender (control group: t=0.98, P=0.34; OA group: t=0.18, P= 0.86), BMI (control group: t=0.56, P=0.92; OA group: t=0.17, P=0.85) and smoking (control group: t=1.89, P=0.08; OA group: t=0.70, P=0.49). The serum level of COMP was higher in the patients with higher K-L grades than in the patients with lower K-L grades (F=15.56, P=0.001) . The sub-clinical KOA patients without X-ray changes can be detected significant higher COMP levels than sub-clinical patients with other diseases (t=2.55, P=0.03). Therefore, according to this method, subclinical OA patients can be detected from people with other sub-clinical diseases successfully.

CONCLUSIONThe serum level of COMP can be used as a potential prognostic marker to diagnose KOA.

Adult ; Aged ; Biomarkers ; blood ; Body Mass Index ; Cartilage Oligomeric Matrix Protein ; Enzyme-Linked Immunosorbent Assay ; Extracellular Matrix Proteins ; blood ; Female ; Glycoproteins ; blood ; Humans ; Male ; Matrilin Proteins ; Middle Aged ; Osteoarthritis, Knee ; blood ; diagnosis

OBJECTIVETo investigate the changes in the firing activity of noradrenergic neurons in the locus coeruleus (LC) in a rat model of Parkinson disease (PD).

METHODS2 and 4 weeks after unilateral lesion of the nigrostriatal pathway in the rat by local injection of 6-hydroxydopamine (6-OHDA) into the right substantia nigra pars compacta (SNc), the firing activity of noradrenergic neurons in LC was recorded by extracellular single unit recording.

RESULTSThe firing rate of LC noradrenergic neurons increased significantly 2 and 4 weeks after 6-OHDA lesions compared to normal rats, respectively (P < 0.05). The percentage of irregularly firing neurons was obviously higher than that of normal rats during the fourth week after SNc lesion (P < 0.05).

CONCLUSIONLC noradrenergic neurons are overactive and more irregular in 6-OHDA-lesioned rats. These changes suggest an implication of the LC in the pathophysiological mechanism of PD.

Action Potentials ; physiology ; Animals ; Disease Models, Animal ; Locus Coeruleus ; pathology ; Male ; Neurons ; physiology ; Norepinephrine ; metabolism ; Oxidopamine ; Parkinsonian Disorders ; chemically induced ; pathology ; Rats ; Rats, Sprague-Dawley ; Time Factors

OBJECTIVETo investigate the mutant alleles of thiopurine S-methyltransferase (TPMT) among Jing Chinese.

METHODSPolymerse chain reaction-single strand conformation polymorphism (PCR-SSCP) techniques were developed for assaying exons 5, 7 and 10 of the TPMT gene respectively and were used to detect mutant TPMT alleles among Jing Chinese.

RESULTSTwo cases of TPMT*3C (A719G) heterozygotes were identified in 103 Jing Chinese; other deleterious alleles such as TPMT*2 (G238C), TPMT*3A (G460A/A719G) and TPMT*3B (G460A) were not found; 27 cases of silent mutant allele TPMT*1S (T474C) were also identified (5 homozygotes and 22 heterozygotes).

CONCLUSIONThe PCR-SSCP assay established and adopted in this study was sensitive and reliable, which could be used to detect mutant TPMT alleles. Allele frequency of TPMT*3C is low among Jing Chinese (1.0%), and TPMT*3C appears to be the most prevalent deleterious allele in this population.

Adolescent ; Alleles ; Asian Continental Ancestry Group ; genetics ; China ; Exons ; genetics ; Female ; Gene Frequency ; Genotype ; Humans ; Male ; Methyltransferases ; genetics ; Mutation ; Polymerase Chain Reaction ; Polymorphism, Single-Stranded Conformational