Objective:To investigate the expression of serum caspase-cleaved cytokeratin 18(CCCK-18) in patients with cerebral ischemic stroke and its diagnostic value.Methods:One hundred and six patients with cerebral ischemic stroke who were diagnosed and treated in Affiliated Dongfeng Hospital from October 2018 to October 2019 were selected as the study group. Ninety patients who underwent digital subtraction angiography (DSA) during the same period and showed no other abnormalities inside or outside the skull were selected as control group. The baseline data of gender, age, drinking history, smoking history, hypertension history, diabetes history, coronary heart disease, and other subjects in the two groups had no significant differences ( P>0.05). Cubital venous blood of 5 ml from two groups of subjects were collected, and the level of serum CCCK-18 was detected by enzyme-linked immunosorbent assay. The levels of total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C) were detected by enzymatic method. The receiver operating characteristic curve (ROC curve) was used to analyze the diagnostic efficacy of serum CCCK-18 in patients with ischemic stroke, and the relationship between serum CCCK-18 and TC, TG, LDL-C, HDL-C were analyzed by Pearson test. Results:The levels of serum CCCK-18, TC, TG, and LDL-C in the study group were significantly higher than those in the control group: (158.10 ± 50.89) U/L vs. (85.57 ± 35.25) U/L, (4.26 ± 0.92) mmol/L vs. (3.92 ± 0.80) mmol/L, (2.34 ± 0.53) mmol/L vs. (1.83 ± 0.47) mmol/L, (3.12 ± 0.73) mmol/L vs. (2.61 ± 0.67) mmol/L, and HDL-C level was lower than that in the control group: (1.20 ± 0.24) mmol/L vs. (1.32 ± 0.28) mmol/L, and there were significant differences ( P<0.05). Logistic regression analysis showed that CCCK-18, TC, TG, LDL-C, and HDL-C were independent risk factors for patients with ischemic stroke ( P<0.05). The area under the curve(AUC) of serum CCCK-18 to distinguish ischemic stroke from the control group was 0.878, with a sensitivity of 84.91% and a specificity of 78.89%. The AUC of serum CCCK-18 to identify patients with mild ischemic stroke was 0.763, with a sensitivity of 70.37% and a specificity of 78.89%. Correlation analysis showed that serum CCCK-18 was positively correlated with TC, TG, and LDL-C in patients with ischemic stroke ( r = 0.711, 0.722, 0.705), and negatively correlated with HDL-C ( r = - 0.714), and there were significant differences ( P<0.01). Conclusions:Serum CCCK-18 levels are significantly increased in patients with cerebral ischemic stroke, which can be used as a biomarker for diagnosis and judgment of disease severity.

Aim To investigate the interactions be-tween the neuroinflammation caused by lipopolysaccha-ride(LPS) and brain CYP2E1.Methods The human cholinergic neuroblastoma cell line IMR-32 was treated with LPS ( 0.1 mg · L-1 , 1.0 mg · L-1 ) , and the LDH and SOD activities were determined after 24 h in-cubation .In order to determine the roles of MAPK sig-naling pathway in the regulation of CYP 2E1 by LPS, the IMR-32 cells were treated with p38 pathway inhibi-tor SB203580 or ERK pathway inhibitor U 0126 for 45 min before the incubation with LPS .The human do-paminergic neuroblastoma cell line SH-SY5Y with CYP2 E1 over-expression was established . The LDH and SOD activities were determined in SH-SY5 Y cells over-expressed CYP2 E1 and control cells treated with LPS(0.1 mg· L-1 , 1.0 mg· L-1 ) for 24 h.Results
The levels of LDH in IMR-32 cells treated with high-dose LPS were increased by 1.38-fold ( P <0.01 ) compared with the control group , and the levels of SOD reduced by 15.0%( P <0.01 ) .Compared with the control, CYP2E1 mRNA and protein levels in IMR-32 cells treated with high-dose LPS were increased by 1.25-fold(P<0.01) and 1.19-fold(P<0.05).The up-regulation of CYP2E1 by LPS could be attenuated by SB203580 and U0126 pretreatment.Compared with the control cells, the CYP2E1 over-expression in-creased LDH levels by 1.28-fold ( P<0.01 ) and de-creased SOD levels by 3.53-fold ( P<0.01 ) after the low-dose of LPS treatment .The CYP2E1 over-expres-sion increased LDH levels by 1.54-fold ( P <0.01 ) and decreased SOD levels by 2.17-fold( P<0.01) af-ter the high-dose of LPS treatment , compared with the control cells.Conclusions LPS can induce CYP2E1 mRNA and protein levels , and the p38 and ERK sig-naling pathway may be involved in the regulation .The elevated CYP2 E1 levels aggravate the damage to neuro-nal cells caused by LPS .Brain CYP2E1 may be an im-portant contributing factor to the pathological process of neuroinflammatory injury .

10.3969/j.issn.2095-4344.2013.23.026

Objective In order to investigate the roles of hIGF\|1 in treatment of diabetes mellitus and diabetic syndromes,the gene of human insulin like growth factor type Ⅰ(IGF\|1) was cloned and constructed into eukaryotic expression vector,then the expression and activity were determined. Methods Total cellular RNA of human fetal liver was abstracted and the RT\|PCR amplification of the cDNA fragment was performed.The fragment was cloned into pUCM\|T vector and sequenced.The eukaryiotic expression vector was recombined and transfected into fibroblast cell line,COS\|7.The expression of hIGF\|1 was examined by in situ hybridization and immunohistochemistry.The effect of hIGF\|1 on cultured islet cells was observed by glucose\|stimulated insulin release assay. Results The cDNA fragment of 710bp with additional Kozak sequence was amplified by RT\|PCR.Eukaryiotic expression vector pCI\|neo/hIGF\|1 was constructed and IGF\|1 gene expressed in COS\|7.The biological activity of hIGF\|1 was proved by increasing inslin secretion from islet cells.Conclusion\ The newly constructed vector,pCI\|neo/hIGF\|1 could be transfected into COS7 cells and its expressed product showed to have the biology activity of hIGF\|1.\;[

Objective To compare the infectivity between laboratory adapted human inununodefi- ciency virus(HIV-1) and primary HIV-1 isolates for different mucosal epithelial cell lines. Methods Mu-cosal epithelial cells Caco-2, T-84, HeLa and lymphocyte MT-4 were infected with laboratory adapted HIV-1 SF33 and 2 primary HIV-1 isolates (02010561, 02010141). Culture supernatant and cells were collected respectively on 3-4 days interval after virus inoculation. The former was tested for HIV-1 antigen P24 level and viral load, and the latter was tested for total viral DNA and integrated viral DNA. Results All 3 virus strains could infect MT-4 cells and integrate into their genome. Only HIV-1 SF33 could infect Caco-2 cells but could not integrate into their genomic DNA. Both HIV-1 SF33 and 02010561 infected HeLa cells but only integration of HIV-1 SF33 was detected. All the 3 HIV-1 strains infected T-84 cells but only the integra-tion of HIV-1 SF33 and 02010141 was observed. Conclusion Although laboratory adapted and primary HIV-1 strains are able to infect human mucosal epithelial cell lines, transient or productive infection estab-lished in different mucosal epithelial cells is dependent on the character of cells and virus strains.

Purpose To detect the expressions of stress induced phosphoprotein 1 (STIP1) and estrogen receptor-α(ER-α) in papil-lary thyroid carcinoma and to analyse the relationship between STIP1 and ER-α. Methods 54 cases of paraffin-embedded tissues of papillary thyroid carcinoma, 18 cases of papillary thyroid carcinoma with lymph node metastasis, 15 cases of Hashimoto’ s thyroiditis, 10 cases of adjacent normal thyroid tissue were collected. The expressions of STIP1 and ER-αwere detected by immunohistochemistry, and the relationship between the expressions and clinicopathological features of papillary thyroid carcinoma was analyzed. Results The expression of STIP1 and ER-α in papillary thyroid cancer group ( 55. 6% and 44. 4%) were higher than that of normal thyroid group (10% and 0) and Hashimoto’s thyroiditis group (8. 3% and 0, all P<0. 05). STIP1 expressions was related to lymph node metastasis ( P<0. 05 ) , while ER-α expression was related to gender, TG-Ab and the merger of nodular goiter, but not related to lymph node metastasis (P>0. 05). The expressions of STIP1 and ER-α in papillary thyroid carcinoma were not related to patients’ age , tumor location, number of tumors, tumer size, invasion of capsule, the concomitant Hashimoto’ s thyroiditis and TPO-Ab ( all P>0. 05). And the expressions of STIP1 was not related to gender, TG-Ab and the merger of nodular goiter (all P>0. 05). A positive correlation was found between the expressions of STIP1 and ER-αin thyroid papillary carcinoma (P<0. 05). Conclusion STIP1 and ER-α in papillary thyroid carcinoma may be related with lymph node metastasis.

OBJECTIVETo investigate the effect of oligochitosan in promoting intestinal absorption of protoberberine alkaloids in extracts from Corydalis saxicola total alkaloids.

METHODThe in vitro single-pass intestinal perfusion model in rats was established to study the changes in absorption kinetic parameters of dehydrocavidine, berberine hydrochloride and palmatine chloride in C. saxicola total alkaloids after the addition of different concentrations oligochitosan and evaluate the effect of oligochitosan in promoting intestinal absorption of the drugs.

RESULTThe concentration of oligochitosan had different effects on the absorption rate constant (Ka) and apparent permeability coefficient (Peff) of the three active component in rat intestines. Ka and Peff in 0.5% oligochitosan group significantly increased, indicating a stronger effect in promoting the absorption.

CONCLUSIONOligochitosan has a certain effect in promoting the intestinal absorptions of protoberberine alkaloids in C. saxicola total alkaloids.

Animals ; Berberine Alkaloids ; administration & dosage ; pharmacokinetics ; Chitin ; administration & dosage ; analogs & derivatives ; Corydalis ; chemistry ; Drugs, Chinese Herbal ; administration & dosage ; pharmacokinetics ; Intestinal Absorption ; drug effects ; Intestines ; drug effects ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley

[ ABSTRACT] AIM:To investigate the effect of oxidized low-density lipoprotein ( ox-LDL) on autophagy in mac-rophages and the underlying molecular mechanisms.METHODS:RAW264.7 macrophages were pretreated with 2 mg/L anti-CD36 monoclonal antibody (anti-CD36 mAb), 5 μmol/L diphenyleneiodonium (DPI), 3 mmol/L 3-methyladenine (3-MA) or 1μmol/L rapamycin for 1 h and then treated with ox-LDL (100 mg/L) for 12 h.The viability of the cells was measured by MTT assay.The activities of lactic dehydrogenase ( LDH) in the medium and nicotinamide adenine dinucleoti-de phosphate ( NADPH) oxidase, superoxide dismutase ( SOD) in the cells as well as the levels of intracellular reactive ox-ygen species ( ROS) and malondialdehyde ( MDA) were determined to characterize the membrane integrity and the oxida-tive stress, respectively.The protein levels of beclin-1 and microtubule-associated protein 1 light chain 3-II ( LC3-II) , 2 important molecular markers of autophagy, were examined by Western blotting.RESULTS:ox-LDL induced autophagy in
RAW264.7 macrophages as assessed by upregulation of beclin-1 and LC3-II.Similar to 3-MA, an autophagy inhibitor, an-ti-CD36 mAb significantly inhibited the ox-LDL-induced upregulation of beclin-1 and LC3-II.Anti-CD36 mAb suppressed the ox-LDL-induced oxidative stress as revealed by decreased NADPH oxidase activation, ROS and MDA generation as well as increased SOD activity.Similar results were observed in the cells pretreated with DPI, a NADPH oxidase inhibitor.Mo-reover, DPI significantly inhibited the ox-LDL-induced upregulation of beclin-1 and LC3-II.Inaddition, the decrease in the cell viability and increase in LDH release induced by ox-LDL were promoted by 3-MA and blocked by rapamycin ( an auto-phagy inducer).CONCLUSION: ox-LDL induces autophagy in RAW264.7 macrophages, which may be involved in CD36-mediated ox-LDL uptake and subsequent activation of oxidative stress, and moderate activation of autophagy may pro-tect macrophages from ox-LDL-induced injury.

Objective To identify Isatidis Radix in different areas by SDS-PAGE;To provide the basis for the identification and quality evaluation of Isatidis Radix. Methods By using SDS-PAGE technology, the protein profiles of Isatidis Radix in Heilongjiang and Hebei regions were established. Results Isatidis Radix protein in Heilongjiang contained about 13 protein subunits, and the major subunits were 10.5 kD, 23.0 kD, 24.9 kD, and 44.1 kD. Isatidis Radix protein in Hebei contained about 12 subunits, and the major subunits were 55.6 kD, 45.9 kD, 34.3 kD, 18.9 kD, 12.4 kD, and 10.5 kD. Conclusion SDS-PAGE technology can be used as one of the references to identify Isatidis Radix in different areas.

SHIV-CN97001 played an important role in assessing the immune effect and strategy of the AIDS vaccine which included genes of the predominant prevalent HIV-1 strain in China. In this study, SHIV-CN97001 was in vivo passaged serially to construct pathogenic SHIV-CN97001/rhesus macaques model. To identify variation in the gp120 region of SHIV-CN97001 during passage, the fragments of gp120 gene were amplified by RT-PCR from the plasma of SHIV-CN97001 infected animals at the peak viral load time point and the gene distances (divergence, diversity) were calculated using DISTANCE. The analysis revealed that the genetic distances of SHIV-CN97001 in the third passage animals were the highest during in vivo passage. It had a relationship between viral divergence from the founder strain and viral replication ability. The nucleic acid sequence of the V3 region was highly conservative. All of the SHIV-CN97001 strains had V3 loop central motif (GPGQ) and were predicted to be using CCR5 co-receptor on the basis of the critical amino acids within V3 loop. These results show that there was no significant increase in the genetic distance during serial passage, and SHIV-CN97001 gp120 gene evolved toward ancestral states upon transmission to a new host. This could partly explain why there was no pathogenic viral strain obtained during in vivo passage.