Leukemia Genotype Analysis of Children with Acute Lymphoblastic Leukemia in Yunnan Area / 中国实验血液学杂志

OBJECTIVE@#To analyze 43 leukemia genes in children with acute lymphoblastic leukemia (ALL) in Yunnan province, and provide the basis for the diagnosis and treatment of children with ALL in this area.@*METHODS@#The clinical data of 428 children with newly diagnosed ALL in Yunnan area from January 2015 to December 2020 were retrospectively analyzed. Multiple nested PCR technology was used to detect 43 common leukemia genes.@*RESULTS@#Among the 428 children with ALL, 159 were positive for leukemia genes, with a positive rate of 37.15% (159/428), and a total of 15 leukemia genes were detected. Among the 159 leukemia gene-positive children, ETV6-RUNX1+ accounted for 25.79% (41/159), followed by E2A-PBX1+ and BCR-ABL+, accounting for 24.53% (39/159) and 23.27% (37/159) respectively. MLL+ accounted for 6.29% (10/159), WT1+ accounted for 4.40% (7/159), IKZF1 gene deletion and CRLF2+ accounted for 3.77% (6/159) respectively. The positive rate of MLL (46.15%) was the highest in ＜1-year old group, the positive rate of ETV6-RUNX1 (10.56%) was the highest in 1-10-year old group, and BCR-ABL+ rate (23.65%) was the highest in >10-year old group. The distribution of leukemia genes in different age groups was statistically significant (P <0.05).@*CONCLUSION@#The most common fusion gene of children with ALL in Yunnan is ETV6-RUNX1, followed by E2A-PBX1 and BCR-ABL.

Intravenous Tenecteplase for Acute Ischemic Stroke Within 4.5–24 Hours of Onset (ROSE-TNK): A Phase 2, Randomized, Multicenter Study / 대한뇌졸중학회지

Background@#and Purpose Intravenous tenecteplase (TNK) efficacy has not been well demonstrated in acute ischemic stroke (AIS) beyond 4.5 hours after onset. This study aimed to determine the effect of intravenous TNK for AIS within 4.5 to 24 hours of onset. @*Methods@#In this pilot trial, eligible AIS patients with diffusion-weighted imaging (DWI)-fluid attenuated inversion recovery (FLAIR) mismatch were randomly allocated to intravenous TNK (0.25 mg/kg) or standard care within 4.5–24 hours of onset. The primary endpoint was excellent functional outcome at 90 days (modified Rankin Scale [mRS] score of 0–1). The primary safety endpoint was symptomatic intracranial hemorrhage (sICH). @*Results@#Of the randomly assigned 80 patients, the primary endpoint occurred in 52.5% (21/40) of TNK group and 50.0% (20/40) of control group, with no significant difference (unadjusted odds ratio, 1.11; 95% confidence interval 0.46–2.66; P=0.82). More early neurological improvement occurred in TNK group than in control group (11 vs. 3, P=0.03), but no significant differences were found in other secondary endpoints, such as mRS 0–2 at 90 days, shift analysis of mRS at 90 days, and change in National Institutes of Health Stroke Scale score at 24 hours and 7 days. There were no cases of sICH in this trial; however, asymptomatic intracranial hemorrhage occurred in 3 of the 40 patients (7.5%) in the TNK group. @*Conclusion@#This phase 2, randomized, multicenter study suggests that intravenous TNK within 4.5–24 hours of onset may be safe and feasible in AIS patients with a DWI-FLAIR mismatch.

Effect of Modified Xiaoyaosan on Dopamine Transporter in Rats with Hyperprolactinemia / 中国实验方剂学杂志

ObjectiveTo investigate the effect of modified Xiaoyaosan on central dopamine transporter （DAT）/protein kinase C （PKC）-dependent signaling pathway in hyperprolactinemia （HPRL） rats. MethodHPRL rat model was established by chronic combined stress combined with intraperitoneal injection of metoclopramide. Ninety-six rats were randomly divided into six groups， namely， the blank group， model group， western medicine （bromocriptine， 0.001 g·kg-1·d-1） group， and high-， medium-， and low-dose （60， 30， 15 g·kg-1·d-1） modified Xiaoyaosan groups. After 14 days of administration， the serum prolactin （PRL） content was detected by enzyme-linked immunosorbent assay， the expression of tyrosine hydroxylase （TH） in rat hypothalamus by immunohistochemistry， and the protein expression of DAT and PKC in hypothalamus by Western blot. ResultCompared with the blank group， the model group exhibited significantly increased PRL and DAT （P<0.01）， but decreased TH and PKC （P<0.01）. Compared with the model group， bromocriptine and modified Xiaoyaosan at the medium dose significantly lowered the content of PRL （P<0.01）. The modified Xiaoyaosan at the medium and high doses elevated the expression of TH （P<0.05， P<0.01）. The expression levels of PKC in the medium- and low-dose modified Xiaoyaosan groups and the western medicine group were significantly increased （P<0.01）， while the DAT expression levels in the high-， medium-， and low-dose modified Xiaoyaosan groups and the western medicine group were decreased （P<0.01）. ConclusionThe modified Xiaoyaosan is able to up-regulate the expression of TH and down-regulate the level of DAT through PKC-dependent signaling pathway， thereby regulating the PRL.

Ampelopsin inhibited apoptosis and migration of SMMC-7721 cells through MAPKs signaling pathway / 药学学报

This research is aimed to investigate the effect of ampelopsin on apoptosis and migration of human hepatoma SMMC-7721 cells and explore the molecular mechanism. SMMC-7721 cells were pretreated with different doses of ampelopsin and cells proliferation was detected by CCK8 kit. Cell morphology was observed under an inverted microscope. Nuclear morphology was detected by DAPI staining. Apoptotic rate was detected by Annexin V-FITC/PI flow cytometry. Migration and invasion were detected by Transwell and scratch healing test. Western blotting was used to detect cleavage of poly ADP-ribose polymerase (PARP), expression of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), E-cadherin, and N-cadherin, and phosphorylation of ERK, P38 and JNK in MAPKs pathway. Our results showed that ampelopsin significantly inhibited proliferation and induced apoptosis of SMMC-7721 cells, with half inhibition dose (IC50) for 24 h was 38.98 μg·mL-1. With 50 μg·mL-1 ampelopsin treatment, typical apoptotic morphological changes occurred, such as cell detachment, shrinkage and nuclear condensation. Apoptotic rate increased from 15% to 55.1%, with PARP cleavage significantly increased. In addition, treatment of ampelopsin reduced scratch healing of cells and transmembrane cells number. The expression levels of MMP-2 and MMP-9 were decreased. Further analysis of EMT-related proteins showed that after ampelopsin treatment, E-cadherin was up-regulated and N-cadherin was down-regulated. During ampelopsin treatment, ERK reached its peak of activation after 1 h, while the maximum activation time of JNK was 12 h. Meanwhile, P38 was activated within 4 h, with the highest point at 2 h. But after 4 h, ampelopsin inhibited phosphorylation of P38. These results indicated that ampelopsin induced apoptosis and reduced migration through activating MAPKs pathway and reversing EMT process in SMMC-7721 cells. This work provides a mechanistic basis for utilizing ampelopsin for anti-hepatocarcinoma treatment.

Effect of silencing Fisl gene on METH-induced proliferative capacity, mitochondrial membrane potential and ultrastructure of SH-SY5Y cells cultured in vitro / 中国药理学通报

Aim To evaluate the role of fisson I (Fisl) in methamphetamine (METH)-induced injur)' of human neuroblastoma (SH-SY5Y) cells cultured in vitro. Methods SH-SY5Y cells cultured in vitro were divided into different groups by the group design method∗. unsilent groups, silent negative groups and silent groups. Different concentrations of METH induced SH-SY5 Y cells in each group for 24 hours. The expression level of Fisl was detected by Western blot. The effect of METH on the proliferative capacity of SH-SY5Y cells was analyzed by CCK-8 cytotoxicity proliferation assay. The MMP level of METH on SH-SY5Y cells was detected by mitochondrial membrane potential detection kit (JC-1). The effect of METH on the mitochondrial ultrastructure of SH-SY5Y cells was observed by transmission electron microscopy. Results In unsilent group, silent negative group and silent group, the expression level of Fisl increased (P < 0. 05) and the proliferative capacity decreased (P < 0. 05) , and the MMP levels decreased (P <0. 05) with the increase of the concentration of SH-SY5Y cells induced by METH. Compared with the same concentration in unsi-lent group and silent negative group, in silent group, the expression level of Fisl in SH-SY5Y cells de-creased (P < 0. 05) , the proliferative capacity increased (P<0. 05) , and the MMP level increased (P < 0. 05). Compared with control group, 2. 0 mmol • L"1 METH induced unsilent groups, silent negative groups and silent groups, and transmission electron microscopy showed the increase in the mitochondrial small globular structure (P < 0. 01). Conclusion Fisl may play a key role in METH-induced injury of SH-SY5Y cells cultured in vitro.

Study on Constructing Evaluation Indicator System for Clinic Scientific Research on Data Quality Based on Delphi Method / 中国中医药信息杂志

Objective To construct the evaluation indicator system for clinical scientific research on data quality; To provide a basis for assessing data quality of clinic scientific research.Methods Evaluation indicator system for clinic scientific research on data quality was established based on literature review and expert suggestion. A two- rounds Delphi process was conducted and the specialist participating level, concentration degree, coordination degree and authority coefficient were calculated to assess the indicator system.Results Two rounds of Delphi consultations was conducted by 30 experts that from different majors at the department of internal medicine or surgery, and both the effective recovery rates were 100％; the average authority coefficients of expert were 0.79 and 0.81, respectively. In the second round of survey, the final concentration degree ranged from 46.67％ to 54.69％ at full mark rate, and 4.23 to 4.36 at the mean values in 5 primary indicators. The CV for average 5 primary indicators ranged from 0.11 to 0.18. After 2 rounds of expert consultation, the final indicator system was consisted of 5 primary indicators (CRF construction, random control process, informed consent, data capture and data validation), 18 secondary indicators and 63 third-class indicators.Conclusion After two rounds of consultation, the opinions of all experts have been tending to be uniform, with high degree of coordination, and the indicator system can be used in assessment of data quality of clinic scientific research.

Preparetion and analysis on the immunogenicity of Echinococcus granulosus-specific diagnostic antigen Eg-07279 / 中国人兽共患病学报

We screened echinococcosis-specific diagnostic antigen and verified its immunogenicity.The sequencing data of Echinococcus granulosus released by the National Human Genome Research Center was analyzed.The bioinformaties method was used to screen out Eg-07279 antigen gene Eg-07279,which was not expressed in Echinococcus granulosus and highly expressed in the original metacercariae.After expression,the recombinant protein rEg-07279 was purified by affinity chromatography.The specific IgG level was detected in the recombinant protein immunized mice and the immunogenicity was verified by Western blot.The selected antigen molecule Eg-07279 was cloned,expressed and purified to obtain the recombinant protein.The results of ELISA showed that the specific IgG level of Eg-07279 (2.559±0.125) was significantly higher than that of the blank group (0.319 0±0.01),the difference was statistically significant (P＜0.01);the Western blot results showed that the recombinant plasmids could be recognized by the secondary infection of the protoscolex and the serum of the recombinant protein immunized group,while the serum of the blank control group was not recognized by the serum.In summary,the Eg07279 antigen of E.granulosus was obtained and it was proved that the recombinant protein has good immunogenicity and is a good diagnostic antigen candidate molecule.

Curcumin inhibits apoptosis of HK-2 cell line induced by cisplatin / 基础医学与临床

Objective To investigate whether curcumin could inhibit cisplatin-induced apoptosis of HK-2 cells. Methods Using MTT methods to observe inhibitory effects of curcumin and cisplatin on the proliferation of HK-2 cell. Detecting caspases expression after curcumin and cisplatin treatment by Western blot. Results Cisplatin induced cell apoptosis, and the effect was dose-dependent(P＜0.05).Under the synergistic effect of curcumin and cisplatin on HK-2 cell, the number of apoptosis decreased(P＜0.05). In the same time the expression of Bcl-2 protein increased (P < 0.05) and the expression of Bcl-2 protein decreased (P < 0.05). Conclusions Cisplatin enhances the apoptosis of HK-2 cell in a dose-dependent manner, and is inhibited by curcumin. Curcumin can antagonize cisplatin-induced apoptosis in HK-2 by regulating the expressions of Bax and Bcl-2 protein.

Chrysin inhibits lipopolysaccharide-induced inflammatory responses of macrophages via JAK-STATs signaling pathway / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the mechanism of chrysin in regulating lipopolysaccharide (LPS)-induced inflammation in RAW264.7 cells.</p><p><b>METHODS</b>RAW264.7 cells were treated with different concentrations (0, 5, 10, 20, 40, 60, 80, 100, 150, and 200 µg/mL) of chrysin for 24 h, and the cell viability was measured using CCK-8. RAW264.7 cells were pre-treated with 10, 30, or 60 µg/mL chrysin for 2 h before stimulation with LPS for different times. The levels of TNF-α, IL-6 and MCP-1 were detected by ELISA, and Western blotting was used to detect the phosphorylation of JAK- 1, JAK-2, STAT-1 and STAT-3. The level of reactive oxygen species in RAW264.7 cells was detected by CM-H2DCFDA fluorescence probe. The effect of ROS on LPS-induced JAK-STATs signal and the inflammatory response of RAW264.7 cells was detected by ROS scavenger NAC. The transcription factors STAT-1 and STAT-3 nuclear translocation were observed by laser confocal microscopy.</p><p><b>RESULTS</b>Chrysin below 60 µg/mL did not significantly affect the viability of RAW264.7 cells. At 10, 30, and 60 µg/mL, chrysin dose-dependently inhibited the expression of iNOS induced by LPS. Chrysin treatment also inhibited LPS-induced phosphorylation of JAK-STATs, nuclear translocation of STAT1 and STAT3, release of TNF-α, IL-6 and MCP-1, and the production of ROS in RAW264.7 cells; ROS acted as an upstream signal to mediate the activation of JAK-STATs signaling pathway.</p><p><b>CONCLUSION</b>Chrysin blocks the activity of JAK-STATs mediated by ROS to inhibit LPS-induced inflammatory response in RAW264.7 cells.</p>

Neuroprotective effect of Nogo-66 receptor silencing in preterm rats with brain injury caused by intrauterine infection / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the effect of Nogo-66 receptor (NgR) silencing with specific small interfering RNA (siRNA) on brain injury repair in preterm rats with brain injury caused by intrauterine infection and related mechanism of action.</p><p><b>METHODS</b>The pregnant Sprague-Dawley rats (with a gestational age of 15 days) were selected, and premature delivery was induced by RU486 or lipopolysaccharide (LPS). The preterm rats delivered by those treated with RU486 were selected as the control group. The preterm rats with brain injury caused by intrauterine infection induced by LPS were divided into model, empty vector, and NgR-siRNA groups, with 36 rats in each group. The rats in the control and model groups were given routine feeding only, and those in the empty vector and NgR-siRNA groups were given an injection of lentiviral empty vector or NgR-siRNA lentivirus via the lateral ventricle on postnatal day 1 (P1) and then fed routinely. On P3, P7, and P14, 8 rats in each group were randomly selected and sacrificed to harvest the brain tissue. RT-PCR was used to measure the mRNA expression of NgR. Western blot was used to to measure the protein expression of active RhoA. The immunofluorescence histochemistry was used to determine the degree of activation of microglial cells and the morphology of oligodendrocyte precursor cells (OPCs). Hematoxylin and eosin staining was used to observe the pathological changes in brain tissue. The behavioral score was evaluated on P30.</p><p><b>RESULTS</b>On P3, the NgR-siRNA group had significantly lower mRNA expression of NgR and protein expression of active RhoA in brain tissue than the model and empty vector groups (P<0.05). In each group, the mRNA expression of NgR was positively correlated with the protein expression of active RhoA (P<0.05). The results of immunofluorescence histochemistry showed that on P3, the NgR-siRNA group had a significantly reduced fluorescence intensity of the microglial cells labeled with CD11b compared with the model and empty vector groups (P<0.05). The OPCs labeled with O4 antibody in the four groups were mainly presented with tripolar cell morphology. The results of pathological examination showed a normal structure of white matter with clear staining in the periventriclar area in the control group, a loose structure of white matter with disorganized fibers and softening lesions in the model and empty vector groups, and a loose structure of white matter with slightly disorganized fibers, slight gliocyte proliferation, and no significant necrotic lesions in the NgR-siRNA group. As for the behavioral score, compared with the model and empty vector groups, the NgR-siRNA group had a higher score in the suspension test, a longer total activity distance, and greater mean velocity and number of squares crossed, as well as a shorter time of slope test and a shorter time and distance of activity in the central area (P<0.05), while there were no significant differences in these parameters between the NgR-siRNA and control groups (P>0.05).</p><p><b>CONCLUSIONS</b>NgR silencing with specific siRNA can effectively silence the expression of NgR in pertem rats with brain injury caused by interauterine infection and has a significant neuroprotective effect in brain injury repair.</p>

Salidroside protects PC12 cells from HO-induced apoptosis via suppressing NOX2-ROS-MAPKs signaling pathway / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the molecular mechanism by which salidroside protects PC12 cells from HO-induced apoptosis.</p><p><b>METHODS</b>PC12 cells cultured in DMEM supplemented with 10% horse serum and 5% fetal bovine serum were pretreated with different doses of salidroside for 2 h and then stimulated with HOfor different lengths of time. The expression levels of PARP and caspase 3 and the phosphorylation of p38, ERK and JNK were determined with Western blotting. The cell nuclear morphology was observed after DAPI staining. The production of ROS was detected using a ROS detection kit, and the levels of gp91and p47in the membrane and cytoplasm were detected by membrane-cytoplasm separation experiment; the binding between gp91and p47was assayed by coimmunoprecipitation experiment.</p><p><b>RESULTS</b>Salidroside dose-dependently suppressed cell apoptosis, lowered phosphorylation levels of p38, ERK and JNK, inhibited the production of ROS, reduced the binding between gp91and p47, and inhibited the activity of NOX2 in PC12 cells exposed to HO.</p><p><b>CONCLUSION</b>Salidroside protects PC12 cells from HO-induced apoptosis at least partly by suppressing NOX2-ROS-MAPKs signaling pathway.</p>

Effect of sodium aescinate in inducing human breast cancer MCF-7 cells apoptosis by inhibiting AKT, ERK and upstream signal SRC activity / 中国中药杂志

To study the effect of sodium aescinate in inducing human breast cancer MCF-7 cells apoptosis and its possible mechanism. MTT assay was used to detect the inhibitory effect of sodium aescinate on the proliferation of MCF-7 cells. The morphological changes were observed under inverted microscope. DAPI nuclear staining was used to detect the changes in cell nucleus. Annexin V-FITC/PI flow cytometry was adopted to test the apoptosis rate. Changes in apoptosis-related proteins (PARP, cleaved caspase-8 and pro-caspase-3), cell survival-associated signal molecules (AKT and ERK) and their common upstream kinase SRC was detected by Western blotting. The result showed that after different concentrations of sodium aescinate were used to treat breast cancer MCF-7 cells, they inhibited the proliferation of MCF-7 cells in a dose-dependent manner, induced cell apoptosis (typical morphological changes in nucleus, significant increase in cell apoptosis rate). The expressions of cleaved PARP and caspase-8 increased, while the expression of pro-caspase-3 decreased, which further verified sodium aescinate's effect in inducing cell apoptosis. Sodium aescinate significantly inhibited the phosphorylation of cell survival-related signal molecules (AKT, ERK) and down-regulate the activation of their common up-stream kinase SRC. The findings indicated that sodium aescinate can block signals transiting to downstream molecules AKT, ERK, inhibit the proliferation of breast cancer cell MCF-7 cell apoptosis and induced cell apoptosis by suppressing the activation of SRC.

Expression of enterovirus 71 capsid protein VP1 in Escherichia coli and its clinical application

The VPl gene of enterovirus 71 (EV71) was synthesized, construct a recombinant plasmid pET15b/VP1 and expressed in E. coli BL21. The recombinant VP1 protein could specifically react with EV71-infected patient sera without the cross-reaction with serum antibodies of coxsackievirus A16 (CA16), A4, A5, B3 and B5 as well as echovirus 6. In acute and convalescent phases, IgM and IgG antibodies of 182 serum samples were detected by ELISA with recombinant VP1 protein as a coated antigen. The results showed that the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of IgM antibodies in serum samples for the diagnosis of EV71 infection were 90.1, 98.4, 98.8 and 88.7%, respectively; similarly, those of IgG antibodies in serum samples were 82.4, 89.1, 91.5 and 78.1%, respectively. Five of 80 samples (6.25%) from CA16infected patients were detected positive by ELISA with recombinant VP1 protein in which indicated the cross reactions and 0 of 5 samples from patients infected with other enteroviruses including CA4, CA5, CB3, CB5 and echovirus 6. Therefore, the recombinant VP1 protein of EV7l may provide a theoretical reference for establishing an effective antibody screening of IgM for EV71-infected patients with clinically suspected hand, foot, and mouth disease (HFMD).

Differential gene expressions of the MAPK signaling pathway in enterovirus 71-infected rhabdomyosarcoma cells

BACKGROUND: Mitogen-activated protein kinase (MAPK) signaling pathway plays an important role in response to viral infection. The aim of this study was to explore the function and mechanism of MAPK signaling pathway in enterovirus 71 (EV71) infection of human rhabdomyosarcoma (RD) cells. METHODS: Apoptosis of RD cells was observed using annexin V-FITC/PI binding assay under a fluorescence microscope. Cellular RNA was extracted and transcribed to cDNA. The expressions of 56 genes of MAPK signaling pathway in EV71-infected RD cells at 8 h and 20 h after infection were analyzed by PCR array. The levels of IL-2, IL-4, IL-10, and TNF-α in the supernatant of RD cells infected with EV71 at different time points were measured by ELISA. RESULTS: The viability of RD cells decreased obviously within 48 h after EV71 infection. Compared with the control group, EV71 infection resulted in the significantly enhanced releases of IL-2, IL-4, IL-10 and TNF-α from infected RD cells (p < 0.05). At 8 h after infection, the expressions of c-Jun, c-Fos, IFN-i, MEKK1, MLK3 and NIK genes in EV71-infected RD cells were up-regulated by 2.08-6.12-fold, whereas other 19 genes (e.g. AKT1, AKT2, E2F1, IKK and NF-κB1) exhibited down-regulation. However, at 20 h after infection, those MAPK signaling molecules including MEKK1, ASK1, MLK2, MLK3, NIK, MEK1, MEK2, MEK4, MEK7, ERK1, JNK1 and JNK2 were up-regulated. In addition, the expressions of AKT2, ELK1, c-Jun, c-Fos, NF-κB p65, PI3K and STAT1 were also increased. CONCLUSION: EV71 infection induces the differential gene expressions of MAPK signaling pathway such as ERK, JNK and PI3K/AKT in RD cells, which may be associated with the secretions of inflammatory cytokines and host cell apoptosis.

CHRNA5 polymorphism and susceptibility to lung cancer in a Chinese population

Polymorphisms in the nicotinic acetylcholine receptor subunit CHRNA5 gene have been associated with lung cancer positive susceptibility in European and American populations. In the present hospital-based, case-control study, we determined whether polymorphism in rs503464 of CHRNA5 is associated with lung cancer risk in Chinese individuals. A single nucleotide polymorphism in CHRNA5 rs503464, c.-166T>A (hereafter T>A), was identified using TaqMan-MGB probes with sequencing via PCR in 600 lung cancer cases and 600 healthy individuals. Genotype frequencies for rs503464 (T>A) were in Hardy-Weinberg equilibrium for the control population. However, genotype frequencies were significantly different between cases and controls (P < 0.05), while allele frequencies were not significantly different between groups. Compared to homozygous genotypes (TT or AA), the risk of lung cancer in those with the heterozygous genotype (TA) was significantly lower (OR = 0.611, 95%CI = 0.486-0.768, P = 0.001). Using genotype AA as a reference, the risk of lung cancer for those with genotype TA was increased 1.5 times (OR = 1.496, 95%CI = 1.120-1.997, P = 0.006). However, no difference in risk was observed between T allele carriers and A allele carriers (OR = 0.914, 95%CI = 0.779-1.073, P = 0.270). Stratification analysis showed that the protective effect of TA was more pronounced in those younger than 60 years, nonsmokers, or those without a family history of cancer, as well as in patients with adenocarcinoma or squamous cell carcinoma in clinical stages III or IV (P < 0.05). Therefore, the heterozygous genotype c.-166T>A at rs503464 of CHRNA5 may be associated with reduced risk of lung cancer, thus representing a susceptibility allele in Chinese individuals.

Carbapenem and cefoxitin resistance of Klebsiella pneumoniae strains associated with porin OmpK36 loss and DHA-1 β-lactamase production

Clinical isolates of carbapenem-resistant Klebsiella pneumoniae (K. pneumoniae) strains are being increased worldwide. Five pan-resistant K. pneumoniae strains have been isolated from respiratory and ICU wards in a Chinese hospital, and reveal strong resistance to all β-lactams, fluoroquinolones and aminoglycosides. Totally 27 β-lactamase genes and 2 membrane pore protein (porin) genes in 5 K. pneumoniae strains were screened by polymerase chain reaction (PCR). The results indicated that all of 5 K. pneumoniae strains carried blaTEM-1 and blaDHA-1 genes, as well as base deletion and mutation of OmpK35 or OmpK36 genes. Compared with carbapenem-sensitive isolates by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the resistant isolates markedly lacked the protein band of 34-40 kDa, which might be the outer membrane proteins of OmpK36 according to the electrophoresis mobility. In addition, the conjugation test was confirmed that blaDHA-1 mediated by plasmids could be transferred between resistant and sensitive strains. When reserpine (30 µg/mL) and carbonyl cyanide m-chlorophenylhydrazone (CCCP) (50 µg/mL) were added in imipenem and meropenem, the MICs had no change against K. pneumoniae strains. These results suggest that both DHA-1 β-lactamase and loss or deficiency of porin OmpK36 may be the main reason for the cefoxitin and carbapenem resistance in K. pneumoniae strains in our hospital.

Association of TNF-α gene polymorphisms with Graves disease susceptibility and early course thyroid stimulating hormone receptor antibody level in Chinese Han population in Anhui region / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To assess the association of tumour necrosis factor-α (TNF-α) gene polymorphisms at positions -863C/A, -857C/T, -238G/A and Graves disease (GD) susceptibility in Chinese Han population in Anhui region.</p><p><b>METHODS</b>The polymorphisms of TNF-α gene were determined by polymerase chain reaction with specific primers in 254 patients affected with GD and 212 healthy controls. Allelic and genotypic frequencies in GD group and normal controls as well as in different genders were compared. The allelic and genotypic frequencies for different thyroid stimulating hormone receptor antibody (TRAb) levels (TRAb > 12 U/L; ≤12 U/L) were also compared among patients with earlier onset GD.</p><p><b>RESULTS</b>(1) The A allele at -863C/A locus in GD group (16.73%) was significantly greater than that of the control group (11.79%) (P< 0.05, OR = 1.503); the frequency of AA+CA genotype of -863C/A locus in GD group (32.68%) was significantly greater than that of control group (23.58%) (P< 0.05, OR = 1.573). There was no significant difference (P> 0.05) in the allelic and genotypic frequencies of -857C/T, -238G/A loci between the two groups. (2) There was no significant difference (P> 0.05) in the allelic and genotypic frequencies of -863C/A, -857C/T, -238G/A loci between patients of different genders. (3) There was no significant difference (P>0.05) in such frequencies between patients with earlier onset GD and different TRAb levels (TRAb > 12 U/L; ≤12 U/L).</p><p><b>CONCLUSION</b>(1) The -863 A allele of TNF-α gene may contribute to the development of GD in Chinese Han population in Anhui, whilst -857C/T, -238G/A alleles may not. (2) There is no association between TNF-α gene -863C/A, -857C/T, -238G/A polymorphisms and development of GD in different genders. (3) There was no association between above polymorphisms and TRAb levels in patients with earlier onset GD.</p>

Change of cytokine expressions on bone marrow mesenchymal stem cells in patients with bone marrow failure syndromes and its significance / 中国实验血液学杂志

This study was aimed to investigate the expressions of multiple cytokines on bone marrow mesenchymal stem cells (MSC) in patients with aplastic anemia (AA) and myelodysplastic syndrome (MDS), and its significance. The semi-quantitative reverse transcriptase-PCR (RT-PCR) was used to detect the expressions of IL-1β, SCF, G-CSF at mRNA level in bone marrow MSC of patients with AA and MDS. The real time quantitative polymerase chain reaction (RQ-PCR) technique was used to detect the mRNA expression of TPO in bone marrow MSC of AA and MDS patients. The results indicated that the expression of SCF in AA group was much lower than that in the normal control group (p < 0.05), and the expression of TPO in AA group was higher than that in the normal control group (p < 0.05), while the expression of IL-1β of AA had no significant difference when compared with the normal control group (p > 0.05). Compared with normal control group, the expressions of SCF of MDS patients was lower (p < 0.05), but the expressions of IL-1β and TPO did not show significant difference (p > 0.05). The expressions of IL-1β, SCF and TPO were no significant difference between AA and MDS groups (p > 0.05). Neither the AA patients, MDS patients nor the normal control group had the expression of G-CSF. It is concluded that the expression of SCF and TPO in bone marrow MSC of AA patients are obviously abnormal, the expression of SCF is also abnormal in bone marrow MSC of MDS patients.

In vitro investigation on specific anti-leukemia cell effect of CTL induced by sensitized dendritic cells from umbilical cord blood / 中国实验血液学杂志

This study was aimed to investigate the specific anti-leukemia cell effect of cytotoxic T lymphocytes (CTLs) induced by HL-60 or K562 cell-sensitized dendritic cells (DCs) from umbilical cord blood. 12 units of human umbilical cord blood (UCB) were collected and the mononuclear cells (MNCs) were isolated from UCB, then cultured with granulocyte monocyte colony- stimulating factor (GM-CSF), interleukin 3 (IL-3), recombinant human stem cell factor (SCF) and EPO for 3 - 4 weeks. Flow cytometry was used to determine the number of DCs and cell surface antigens before and after culture with monoclonal antibodies including CD83, CD1a, CD11c and CDw123. HL-60 and K562 were frozen-thawed, and released their tumor antigen peptides (TAP). The CTLs were produced by sensitizing T lymphocytes with DC-loaded HL-60 and K562 cell antigens. The test of (3)H-TdR incorporation was used to detect the immunostimulation activity of DCs. MTT assay was applied to evaluate specific cytotoxicity of CTL on leukaemia cells. The results indicated that the MNCs of UCBs cultured with GM-CSF, IL-3, EPO and SCF were shown to differentiate into CD1a(+) CD11c(+) CD83(+) CDw123(+) DCs. Numbers of DCs from UCBs remarkably increased in 2 - 4 weeks and then decreased. After culture with cytokines DCs increased (10.6 - 28.2) x 10(5)/ml in actual numbers. The CTL induced by DC pulsed with HL-60, K562 frozen-thawed lysates were effective to kill HL-60 and K562. Cytotoxicity of CTL to HL60 and K562 were (42.04 +/- 8.46)% and (31.25 +/- 11.07)% respectively. It is concluded that the MNCs of UCBs cultured with cytokines of GM-CSF, SCF, EPO and IL-3 can differentiate into CD1a(+), CD83(+), CD11c(+) and CDw123(+) DCs. The CTL induced by DCs pulsed with HL-60, K562 frozen-thawed lysates can effectively kill HL-60 and K562. These DCs as antigen presenting cells play an important role in cancer immunotherapy.

Expression of mucin glycoproteins and cytokeratins in intrahepatic cholangiocarcinoma / 中华病理学杂志

<p><b>OBJECTIVE</b>To compare the immunoprofiles of intrahepatic cholangiocarcinoma and metastatic colorectal adenocarcinoma for mucin glycoproteins (including MUC1, MUC2, MUC5AC and MUC6) and cytokeratins (including CK7, CK19 and CK20), and to assess their diagnostic value.</p><p><b>METHODS</b>One hundred cases of intrahepatic cholangiocarcinoma and 21 cases of metastatic colorectal adenocarcinoma were enrolled into the study. Immunohistochemical study for MUC1, MUC2, MUC5AC, MUC6, CK7, CK19 and CK20 was carried out in all cases by EnVision method.</p><p><b>RESULTS</b>In intrahepatic cholangiocarcinoma, the expression rates of MUC1, MUC2, MUC5AC and MUC6 were 61.0%, 2.0%, 22.0% and 8.0% respectively, as compared to 57.1%, 47.6%, 19.0% and 23.8% respectively in metastatic colorectal adenocarcinoma. On the other hand, the expression rates of CK7, CK19 and CK20 in intrahepatic cholangiocarcinoma were 73.0%, 53.0% and 15.0% respectively, in contrast to 14.3%, 90.5% and 85.7% respectively in metastatic colorectal adenocarcinoma. The difference in expressions of MUC2, MUC6, CK7 and CK20 carried statistical significance.</p><p><b>CONCLUSIONS</b>The immunoprofile for mucin glycoproteins and cytokeratins provides important clues in distinguishing between intrahepatic cholangiocarcinoma and metastatic colorectal adenocarcinoma to liver. The immunophenotype of MUC2-/MUC6-/CK7+/CK20- indicates the diagnosis of intrahepatic cholangiocarcinoma, while MUC2+/MUC6+/CK7-/CK20+ suggests the possibility of metastatic colorectal adenocarcinoma.</p>