Synergistic Effect of Hyperthermia and Neferine on Reverse Multidrug Resistance in Adriamycin-resistant SGC7901/ADM Gastric Cancer Cells

Multidrug resistance（MDR） plays a major obstacle to successful gastric cancer chemotherapy.The purpose of this study was to investigate the MDR reversal effect and mechanisms of hyperthermia in combination with neferine（Nef） in adriamycin（ADM） resistant human SGC7901/ADM gastric cancer cells.The MDR cells were heated at 42℃ and 45℃ for 30 min alone or combined with 10 μg/mL Nef.The cytotoxic effect of ADM was evaluated by MTT assay.Cellular plasma membrane lipid fluidity was detected by fluorescence polarization technique.Intracellular accumulation of ADM was monitored with high performance liquid chromatography.Mdr-1 mRNA,P-glycoprotein（P-gp）,γH2AX expression and γH2AX foci formation were determined by real-time PCR,Western blot and immunocytochemical staining respectively.It was found that different heating methods induced different cytotoxic effects.Water submerged hyperthermia had the strongest cytotoxicity of ADM and Nef combined with hyperthermia had a synergistic cytotoxicity of ADM in the MDR cells.The water submerged hyperthermia increased the cell membrane fluidity.Both water submerged hyperthermia and Nef increased the intracellular accumulation of ADM.The water submerged hyperthermia and Nef down-regulated the expression of mdr-1 mRNA and P-gp.The water submerged hyperthermia could damage DNA and increase the γH2AX expression of SGC7901/ADM cells.The higher temperature was,the worse effect was.Our results show that combined treatment of hyperthermia with Nef can synergistically reverse MDR in human SGC7901/ADM gastric cancer cells.

Reversing multidrug resistance by RNA interference through the suppression of MDR1 gene in human hepatoma cells

AIM： To reverse the multidrug resistance （MDR） by RNA interference （RNAi）-mediated MDRI suppression in heparoma cells.METHODS： For reversing MDR by RNAi technology, two different short hairpin RNAs （shRNAs） were designed and constructed into pGenSil-1 plasmid, respectively. They were then transfected into a highly adriarnycin-resistant HepG2 hepatorna cell line （HepG2/ADM）. The RNAi effect on MDR was evaluated by real-time PCR, cell cytotoxicity assay and rhodarnine 123 （Rh123） efflux assy. RESULTS： The stably-transfected clones showed various degrees of reversal of MDR phenotype. Surprisingly, the MDR phenotype was completely reversed in two transfected clones. CONCLUSION： MDR can be reversed by the shRNAmediated MDRI suppression in HepG2/ADM cells, which provides a valuable clue to make multidrug-resistant hepatoma cells sensitive to anti-cancer drugs.

Association of single nucleotide polymorphisms of brain-derived neurotrophic factor gene and multidrug resistance 1 gene to refractory epilepsy in Chinese Han children

BACKGROUND： There are two hypotheses for the underlying cause of refractory epilepsy： ＂target＂ and ＂transport＂. Studies have shown that brain-derived neurotrophic factor （BDNF） is over-expressed in refractory epilepsy. Multidrug resistance 1 （MDR1） gene encodes for P-glycoprotein, the primary ATP-binding cassette transporter in the human body. Some single nucleotide polymorphisms of the MDR1 gene have been associated with refractory epilepsy. OBJECTIVE： To investigate the association between BDNF gene C270T polymorphism and MDR1 T-129C polymorphism with refractory epilepsy in Chinese Han children through the use of polymerase chain reaction （PCR）-restriction fragment length polymorphism analysis. DESIGN, TIME AND SETTING： A case-control, genetic association study was performed at the Central Laboratory, Third Xiangya Hospital of Central South University from June 2005 to November 2007. PARTICIPANTS： A total of 84 cases of unrelated children with epilepsy, including 41 cases of refractory epilepsy and 43 cases of drug-responsive epilepsy, were enrolled. An additional 30 healthy, Chinese Han children, whose ages and gender matched the refractory epilepsy patients, were selected as normal controls. METHODS： Venous blood was collected and genomic DNA was extracted from the blood specimens. C270T polymorphism in BDNF gene and T-129C polymorphism in MDR1 gene were genotyped using PCR-restriction fragment length polymorphism analysis. Association analysis using the Ftest and Chi-square test was statistically performed between C270T polymorphism in BDNF gene and T-129C polymorphism in MDR1 gene and refractory epilepsy. MAIN OUTCOME MEASURES： The distribution of genotypes and allele frequencies of C270T polymorphism in BDNF gene and T-129C polymorphism in MDR1 gene. RESULTS： The distribution of CC, CT, and TT genotypes, as well as C and T allele frequencies, in the BDNF gene was not significantly different between the refractory epilepsy group, drug-responsive epilepsy group, or the normal control group （P 〉 0.05）. The distribution of TT genotype and T allele frequencies of the MDR1 gene was significantly different in the refractory epilepsy group compared with the drug-responsive epilepsy and normal control groups （P 〈 0.05）. Comparison of haplotype combinations demonstrated that there were no significant differences in combinations of TT＋CC, -FI-＋CT, TC＋CC, and TC＋CT among the three groups （P 〉 0.05）. CONCLUSION： C270T polymorphism of the BDNF gene was not associated with refractory epilepsy in Chinese Han children, but T-129C polymorphism in the MDR1 gene was associated with refractory epilepsy in Chinese Han children. The TT genotype and T allele frequencies could serve as susceptibility loci for refractory epilepsy. Interactions between C270T in BDNF gene and T-129C in MDR1 gene were not observed in refractory epilepsy in Chinese Han children.

Differential expression of breast cancer-resistance protein,lung resistance protein,and multidrug resistance protein 1 in retinas of streptozotocin-induced diabetic mice

AIM：To investigate the altering expression profiles of efflux transporters such as breast cancer-resistance protein（BCRP）,lung resistance protein（LRP）,and multidrug resistance protein 1（MDR1） at the inner blood-retinal barrier（BRB） during the development of early diabetic retinopathy（DR） and/or aging in mice.METHODS：Relative m RNA and protein expression profiles of these three efflux transporters in the retina during the development of early DR and/or aging in mice were examined.The differing expression profiles of Zonula occludens 1（ ZO-1） and vascular endothelial growth factor-A（ VEGFA） in the retina as well as the perfusion characterization of fluorescein isothiocyanate（FITC）-dextran and Evans blue were examined to evaluate the integrity of the inner BRB.RESULTS：There were significant alterations in these three efflux transporters＇ expression profiles in the m RNA and protein levels of the retina during the development of diabetes mellitus and/or aging.The development of early DR was confirmed by the expression profiles of ZO-1 and VEGFA in the retina as well as the compromised integrity of the inner BRB.CONCLUSION：The expression profiles of some efflux transporters such as BCRP,LRP,and MDR1 in mice retina during diabetic and/or aging conditions are tested,and the attenuated expression of BCRP,LRP,and MDR1 along with the breakdown of the inner BRB is found,which may be linked to the pathogenesis of early DR.

Roles of sulfonylurea receptor 1 and multidrug resistance protein 1 in modulating insulin secretion in human insulinoma

BACKGROUND:Sulfonylurea receptor 1(SUR1)and multidrug resistance protein 1(MRP1)are two prominent members of multidrug resistance proteins associated with insulin secretion. The aims of this study were to investigate their expression in insulinomas and their sole and synergistic effects in modulating abnormal insulin secretion. METHODS:Fasting glucose,insulin and C-peptide were measured in 11 insulinoma patients and 11 healthy controls. Prolonged oral glucose tolerance tests were performed in 6 insulinoma patients.Insulin content,SUR1 and MRP1 were detected in 11 insulinoma patients by immunohistochemistry. SUR1 and MRP1 were also detected in 6 insulinoma patients by immunofluorescence. RESULTS:Insulinoma patients presented the typical demons-trations of Whipple’s triad.Fasting glucose of each insulinoma patient was lower than 2.8 mmol/L,and simultaneous insulin and C-peptide were increased in insulinoma patients. Prolonged oral glucose tolerance tests showed that insulin secretion in insulinoma patients were also stimulated by high glucose.Immunohistochemistry and immunofluorescence staining showed that SUR1 increased,but MRP1 decreased in insulinoma compared with the adjacent islets. CONCLUSIONS:The hypersecretion of insulin in insulinomas might be,at least partially,due to the enrichment of SUR1. In contrast,MRP1,which is down-regulated in insulinomas, might reflect a negative feedback in insulin secretion.

Expression of multidrug resistance 1 gene and C3435T genetic polymorphism in peripheral blood of patients with intractable epilepsy

BACKGROUND： Increased expression of multidrug resistance 1 （MDR1） mRNA in peripheral blood of patients with intractable epilepsy is not due to epilepsy drugs, but epilepsy behavior. Monitoring MDR1 expression in peripheral blood is a target for MDR1 gene evaluation. OBJECTIVE： To investigate the influence of antiepileptic drugs and seizures on MDR expression in intractable epilepsy, and to analyze the genetic polymorphisms of C3435T in the MDRl gene. DESIGN, TIME AND SETTING： Factorial designs and comparative observations at the experimental center of the Affiliated Hospital of Qingdao Medical College, Qingdao University between October 2003 and October 2004. PARTICIPANTS： A total of 120 subjects were recruited from the epilepsy clinical department of the Affiliated Hospital of Qingdao Medical College. Four groups （n = 30） were classified according to statistical factorial design： intractable epilepsy, treatment response, no treatment, and normal control groups. METHODS： One-step semi-quantitative reverse-transcription polymerase chain reaction technology was used to test expressions of the MDR1 gene in 120 subjects. C3435T polymorphisms in intractable epilepsy group and normal control groups were analyzed by polymerase chain reaction-restriction fragment length polymorphism. MAIN OUTCOME MEASURES： Expression of MDR1 mRNA in the four groups, and C3435T genetic polymorphisms in intractable epilepsy and normal control groups. RESULTS： MDRl gene expression was increased in the intractable epilepsy group, due to the factor seizures, but not the antiepileptic drugs. However, the interaction between the two factors was not statistically significant. Of the 30 subjects in the intractable epilepsy group, the following genotypes were exhibited： 3 （10%） C/C genotype, 9 （30%） C/T genotype, and 18 （60%） T/T genotype at the site of C3435T, while 4 （13%）, 10 （33%）, and 16 （53%） subjects were determined to express these genotypes in the normal control group, respectively. C and T allele frequency were 25% and 75% in the intractable epilepsy group, and 30% and 70% in the normal control group, respectively. However, there was no statistical difference between the groups. CONCLUSION： Results demonstrated that seizures, not antiepileptic drugs, induced MDR1 gene expression in intractable epilepsy. Genetic polymorphisms of C3435T in the MDR1 gene did not contribute to the development of multidrug resistance in patients with intractable epilepsy.

Detection and clinical significance of multidrug resistance-1 mRNA in bone marrow cells in children with acute lymphoblastic leukemia by real-time fluorescence quantitative RT-PCR

Objective： Multidrug resistance（MDR） is one of the most important reasons for treatment failure and recurrence of acute leukemia. Its manifestations are different in children with acute lymphoblastic leukemia（ALL） which may be due to different detection methods. This study was to detect the expression of MDR1 mRNA in bone marrow cells of children with ALL by real-time fluorescence- quantitative reverse transcription polymerase-chain reaction（FQ-RT-PCR）, and combine minimal residual desease（MRD） detection by flow cytometry（FCM） and to study their relationship with treatment response and prognosis of ALL. Methods：The MDR1 mRNA levels in bone marrow cells from 67 children with ALL[28 had newly diagnosed disease, 27 had achieved complete remission（CR）, 12 recurrent] and 22 children without leukemia were detected by FQ-RT-PCR. MRD was detected by FCM. The patients were observed for 9-101 months, with a median of 64 months. Results：Standard curves of human MDR1 and GAPDH genes were constructed successfully. MDR1 mRNA was detected in all children with a positive rate of 100%. The mRNA level of MDR1 was similar among the newly diagnosed ALL group, CR group, and control group（P 〉 0.05）, but significantly higher in the recurrence group than that in newly diagnosed disease group and control group（0.50 ± 0.55 vs. 0.09 ± 0.26 and 0.12 ± 0.23, P〈 0.05）. 54 ALL patients were followed up, and it was found that MDR1 mRNA level was significantly higher in ALL patients within 3 years duration than that of ALL patients with 3-6 years and over 6 years duration（0.63 ± 0.56 vs. 0.11 ± 0.12 and 0.04 ± 0.06, P〈 0.01）. For the 28 children with newly diagnosed disease, the MDR1 mRNA level was similar between WBC 〉 50 ~ 109 group and WBC〈50 × 10^9 group（P〉 0.05）. In the 33 CR patients, the MDR1 mRNA level was significantly higher in MRD〉10a group than that in MRD〈10a group（0.39 ± 0.47 vs. 0.03 ± 0.03, P 〈 0.05）. Conclusion：The sensitivity and specificity of FQ-RT-PCR in detecting MDR1 mRNA in bone marrowy cells of children with ALL patients are high. MDR1 mRNA is expressed in children with and without leukemia. MDR1 mRNA is highly expressed in the CR ALL patients with high MRD, recurrence and short duration（within 3 years）. Monitoring MRD and the MDR1 mRNA level might be helpful for individual treatment.

Reversal Effect of BM-cyclin 1 on Multidrug Resistance by Down-regulating MRP2 in BALB/C Nude Mice Bearing C-A120 Cells

Our previous study demonstrated that BM-cyclin 1, a traditional anti-mycoplasma drug, could effectively reverse the multidrug resistance （MDR） of C-A120 cells. The present study aims to explore the reversal effect of BM-cyclin 1 on MDR and its mechanisms in BALB/C nude mice bearing C-A120 cells. Irnmunoblotting analysis and reverse transcription-polymerase chain reaction （RT-PCR） were used to study the change in multidrug resistance-associated protein 2 （MRP2） induced by BM-cyclin 1. We found that the expression levels of MRP2 protein and mRNA in C-A120 cells treated with BM-cyclin 1 were reduced significantly. Chemical colorimetry revealed no significant change in the level of glutathione （GSH）. In the xenografl model, the inhibitory rate of C-A120 cells growth in BM-cyclin 1 plus adriamycin （ADM） group was 52%, which was significantly higher than in control group （P〈0.01）. The immunoblotting and RT-PCR results conclusively demonstrated that BM-cycin 1 could significantly reduce the expression of MRP2 in transplanted tumor. In conclusion, BM-cyclin 1 could effectively reverse the MDR of C-A 120 cells in vivo by suppressing the expression of MRP2.

Inhibition of apoptosis-regulatory protein Siva-1 reverses multidrug resistance in gastric cancer by targeting PCBP1

Introduction:Siva-1,as a pro-apoptotic protein,has been shown to induce extensive apoptosis in a number of different cell lines.In our previous study,we showed that overexpressed Siva-1 decreased the apoptosis of gastric cancer cells.So,we believe that it can also work as an anti-apoptotic protein.The present study aimed to determine the specific role of Siva-1 in anticancer drug resistance in gastric cancer in vivo and in vitro and preliminarily reveal the mechanism.Materials and Methods:A vincristine-resistant MKN-28/VCR gastric cancer cell line with stably downregulated Siva-1 was established.The effect of Siva-1 downregulation on chemotherapeutic drug resistance was assessed by measuring the IC50 and pump rate of doxorubicin.Proliferation,apoptosis of cells,and cell cycle were detected via colony formation assay andflow cytometry,respectively.Additionally,migration and invasion of cells was detected via wound healing and transwell assays.Moreover,we determined in vivo effects of LV-Siva-1-RNAi on tumor size,and apoptotic cells in tumor tissues were detected using TUNEL and hematoxylin and eosin staining.Results:Siva-1 downregulation reduced the pump rate of doxorubicin and enhanced the response to drug treatment.Siva-1 negatively regulated proliferation and promoted apoptosis of cells by potentiality G2-M phase arresting.Inhibition of Siva-1 expression in MKN-28/VCR cells significantly weakened wound healing ability and decreased invasion ability.Poly(C)-binding protein 1(PCBP1)was identified as a Siva-1-interacting protein in yeast two-hybrid screening.Semiquantitative RT-PCR and western blotting revealed that Siva-1 downregulation could inhibit expression of PCBP1,Akt,and NF-κB and eventually decrease the expression of MDR1 and MRP1.Conclusion:The current study demonstrated that the downregulation of Siva-1,which functions as a regulator of MDR1 and MRP1 gene expression in gastric cancer cells by inhibiting PCBP1/Akt/NF-κB signaling pathway expression,enhanced the sensitivity of gastric cancer cells to certain chemotherapies.

Role of Hypoxia-inducible Factor-1α in Formation of Muttidrug Resistance Induced by Microenvironment in Hepatocellular Carcinoma

Objective： To explore the role of hypoxia-inducible factor-1α （HIF-1α） in formation of multidrug resistance （MDR） induced by microenvironment and to find a new and effective molecular target on preventing and reversing chemoresistance in hepatocellular carcinoma （HCC）. Methods： In HepG2 cells exposed to hypoxia, low glucose or transfected by plasmid pcDNA3/HBX, the expression of HIF-1α mRNA and protein was respectively detected using real-time fluorescent quantitative PCR and Westernblot technique and its expression localization was investigated by immunocytochemical technique. Plasmid pcDNA3/HIF-1α was transfected into HepG2 cells and then the expression of multidrug resistance related genes mdrl, multidrug resistance-associated protein 1 （MRP1） and lung resistance protein （LRP） in transfected cells was determined by the same methods. Results： In HepG2 cells respectively exposed to hypoxia, low glucose or transfected by plasmid pcDNA3/HBX, HIF-1α was overexpressed at mRNA and protein levels to varying degrees and translocated into nucleus. The gene expression levels of mdrl, MRP1 and LRP in HepG2 cells transfected by plasmid pcDNA3/HIF-1α were respectively increased by 2.4±0.2, 2.2±0.3 and 2.3±0.4 folds as compared with those in non-transfected HepG2 cells （all P〈0.01） and similar changes were observed in protein level. Conclusion： Microenvironmental factors around HCC could modulate the transcription of the MDR related genes by nuclear transcript factor HIF-1α, thereby conferred MDR of HCC. Up-regulation of HIF-1α expression could hold a central position in the formation of MDR of HCC induced by microenvironment. HIF-1α probably becomes a new and effective molecular target on preventing and reversing MDR in HCC.

白介素37下调多药耐药基因-1逆转肺腺癌紫杉醇耐药的研究

目的 本研究旨在探究白介素37(IL-37)在抑制肺腺癌细胞多药耐药性方面的潜在作用,及其对于耐紫杉醇的A549/TAX细胞的影响。方法 通过细胞培养、处理程序、实时荧光定量PCR、Western blot分析和统计学分析等实验方法,系统研究了IL-37对耐紫杉醇A549/TAX细胞的影响。结果 紫杉醇明显抑制了A549和A549/TAX细胞的增殖,其中A549/TAX的耐药指数RI为16.88。100ng/mL的rhIL-37显著抑制了A549/TAX细胞的增殖。在紫杉醇和rhIL-37联合处理组,细胞增殖的抑制率显著高于仅用紫杉醇处理组(P<0.05)。此外,rhIL-37在24小时后显著抑制了A549/TAX细胞的迁移和侵袭。非细胞毒性浓度的rhIL-37也能显著抑制A549/TAX细胞的集落形成。经rhIL-37作用48小时后,A549/TAX细胞中MDR1的表达水平比对照组下降了约66%(P<0.05)。结论 IL-37与紫杉醇联合处理可有效抑制A549/TAX细胞的增殖、迁移和侵袭,同时通过降低MDR1基因的表达水平可能逆转细胞的耐药性,为IL-37在肺腺癌治疗中的潜在应用提供了实验依据。

鸡源mcr-1阳性多重耐药沙门菌的鉴定和特征分析

试验旨在对1株鸡源mcr-1阳性多重耐药沙门菌株进行鉴定以及特征分析,从河南鸡场采集病死鸡样品总计187份,通过SS选择培养基进行沙门菌的分离,利用聚合酶链式反应(PCR)方法扩增16S rRNA和invA对菌株进行鉴定,基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)复核鉴定结果,并采用PCR测序的方法检测黏菌素耐药基因mcr的携带情况。采用微量稀释法测定沙门菌对常见16种抗菌药的敏感性,通过全基因组测序分析沙门菌的特征。结果显示,共分离鉴定出32株鸡源沙门菌,分离率为17.11%,黏菌素耐药率为18.75%;在1株沙门菌中检测到了黏菌素耐药基因mcr-1,检出率为3.13%。mcr-1阳性鸡源沙门菌的黏菌素最小抑菌浓度为16μg/mL,多序列位点分型结果为ST9。药敏结果显示,该菌株对氨苄西林、头孢噻呋、庆大霉素、大观霉素、恩诺沙星、环丙沙星、氟苯尼考、氯霉素、磺胺异恶唑、甲氧嘧啶、四环素耐药;携带blaOXA-1、blaTEM-1B、oqxAB、tet(A)、floR、catB3、sul1、sul2、sul3、ant(3'')、dfrA等多种耐药基因。接合试验证实mcr-1基因可发生接合转移。该研究结果为临床治疗鸡源沙门菌感染中合理使用抗生素以及有效预防、控制耐黏菌素鸡源沙门菌的传播提供了科学依据。

Plasmid mediated antibiotic resistance of Vibrio cholerae Ol biotype E1 Tor serotype Ogawa associated with an outbreak in Kolkata,India

Objective:To determine the antibiotic resistance of Vibrio choleras(V.cholerae) O1 biotype El Tor serotype Ogawa isolates involved in an outbreak of watery diarrhea in Kolkata,and to explore the role of plasmid in mediating antibiotic resistance.Methods:Antibiotic susceptibility and minimum inhibitory concentration(MIC) values of antibiotics for the isolated V.cholerae 01 Ogawa(n=12) were determined by disk diffusion and agar dilution methods,respectively,using ampicillin(Am),chloramphenicol(C),trimethoprim(Tm),tetracycline(T).erythromycine(Er), nalidixic acid(Nx).ciprofloxacin(Cp),amikacin(Ak) and cefotaxime(Cf).Plasmid curing of multidrug resistant(MDR) V.cholerae 01 Ogawa strains was done following ethidium bromide treatment.Following electrophoresis,the plasmid DNAs,extracted from the isolated MDR V. cholerae O1 Ogawa strains and their cured derivatives,were visualized and documented in ’gel doc’ system.Results:The outbreak causing V.cholerae O1 Ogawa isolates were MDR as determined by disk diffusion susceptibility test,and MIC determination.The isolates showed three different drug resistance patterns:AmTmTErNx(for 6 isolates).TmTErCp(for 5 isolates), and AmTniNx(for one isolate),and showed uniform sensitivity to C,Ak and Cf.The loss of plasmids with the concomitant loss of resistance to Am,Tm,T and Er of the isolates occurred following ethidium bromide treatment.Conclusions:The current findings suggest that the V. cholerae O1 Ogawa associated with the cholera outbreak were MDR,and resistance to Am,Tm,T and Er among the isolates were plasmid mediated.

Reversal of Adriamycin Resistance of Hepatocellular Carcinoma by Targeting with Recombined Adenovirus Caning Antisense mdr1 RNA

OBJECTIVE Chemotherapy is an important therapy for hepatocellular carcinoma （HCC）. However, it is not effective in many cases due to recurrence and metastasis even if the initial treatment produces a response. Multidrug resistance （MDR） is considered to be one of the considerable causes. The aim of this study was to reverse MDR of HepG2/ADM cells by blocking mdr1 with an adenovirus vector carrying antisense mdr1 in a tumor transplantated in athymic mice. METHODS PCMV IE was removed from the pshuttle vector. A 0.3 kb AFP promoter was inserted into the pshuttle vector and pCMV changed into pAFP. The pAFP and asmdr1 PCR products were doubly digested with Kpnl and Apal, the digested products were ligated by T4 ligase, the asmdr1 gene was inserted into pAFP and a newly plasmid pAFP-asmdr1 was constructed. Following digestion with PI-SceI/I-Ceu I, pAFP-asmdr1 was ligated with Adeno-X genome DNA and amplified in E.coli XL1-Blue. The HEK293 cells were transfected and virus collected. The HepG2 MDR cells （HepG2/ADM） were induced by graded resistance to ADM and were inoculated into athymic mice. After adeno-asmdr1 was injected, the expression of mdr1-mRNA and the volume of the transplantated tumor and its cells were observed. RESULTS Following injection with Adeno-asmdr1, the tumor volume in the ADM＋Adeno-asmdr1 group did not increase. However the tumor volume in the PBS plus ADM group did significantly increase （P〈0.05）. In the tumor xenograft cells, mdr1 mRNA in the xenografts was assessed by RT-PCR and was found to be reduced at 1 week and 4 weeks in the ADM＋asmdr1 group, but it was stable in the ADM group. It was only 20% in the ADM＋asmdr1 group compared to the ADM group at the 4th week （P〈0.05）. Evidence of apoptosis was observed in the tumor xenograft cells treated with Adeno-asmdr1, but there was rare or no apoptosis in the group treated with ADM and PBS. CONCLUSION Adenovirus carrying antisense mdr1 RNA can partially reverse the MDR of HepG2/ADM cells and inhibit tumor growth by down-regulating mdr1 mRNA resulting in tumor cell apoptosis.

Characterization of Wnt1-inducible Signaling Pathway Protein-1 in Obese Children and Adolescents

Wnt1-inducible signaling pathway protein-1(WISP1),a member of the CCN family,is increasingly being recognized as a potential target for obesity and type 2 diabetes mellitus.Recent studies have shown that WISP1 can regulate low-grade inflammation in obese mice,and circulating WISP1 levels are associated with obesity and type 2 diabetes mellitus in adults.Herein,we measured serum WISP1 levels in obese youth and explored its relationships with pro-inflammatory cytokine interleukin 18(IL-18)and other metabolic indexes.Totally,44 normal-weight and 44 obese children and adolescents were enrolled.Physical and laboratory data were recorded,and then serum levels of WISP1 and IL-18 were determined by enzyme-linked immunosorbent assays.Results showed that serum levels of WISP1 were significantly higher in obese children and adolescents than in normal-weight healthy controls (1735.444-15.29 vs. 1364.084-18.69 pg/mL).WISP1 levels were significantly positively correlated with body mass index (BMI)and BMI z-score (r=0.392,P=0.008;r=0.474,P=0.001,respectively) in obese group;circulating IL-18 was increased in obese individuals (1229.064-29.42 vs. 295.874-13.30 pg/mL).Circulating WISP1 levels were significantly correlated with IL-18 (r=0.542,P<0.001),adiponectin (r=0.585,P<0.001)and leptin (r=0.592,P<0.001).The multivariate stepwise regression analysis showed that higher IL-18 levels represented the main determinant of increased WISP1 levels after adjusting for BMI,waist circumference, fasting insulin,homeostatic model assessment of insulin resistance (HOMA-IR)and HbAlc in obese individuals (β=0.542,P=0.000).WISP1 can be involved in glucose/lipid metabolism in obese youth,which may be modulated by IL-18.Increased WISP1 levels may be a risk factor of obesity and insulin resistance,and WISP1 has a potential therapeutic effect on insulin resistance in obese children and adolescents.

Pathophysiology of insulin resistance and steatosis in patients with chronic viral hepatitis

Chronic hepatitis due to any cause leads to cirrhosis and end-stage liver disease.A growing body of literature has also shown that fatty liver due to overweight or obesity is a leading cause of cirrhosis.Due to the obesity epidemic,fatty liver is now a significant problem in clinical practice.Steatosis has an impact on the acceleration of liver damage in patients with chronic hepatitis due to other causes.An association between hepatitis C virus (HCV) infection,steatosis and the onset of insulin resistance has been reported.Insulin resistance is one of the leading factors for severe fibrosis in chronic HCV infections.Moreover,hyperinsulinemia has a deleterious effect on the management of chronic HCV.Response to therapy is increased by decreasing insulin resistance by weight loss or the use of thiazolidenediones or metformin.The underlying mechanisms of this complex interaction are not fully understood.A direct cytopathic effect of HCV has been suggested.The genomic structure of HCV (suggesting that some viral sequences are involved in the intracellular accumulation of triglycerides),lipid metabolism,the molecular links between the HCV core protein and lipid droplets (the core protein of HCV and its transcriptional regulatory function which induce a triglyceride accumulation in hepatocytes) and increased neolipogenesis and inhibited fatty acid degradation in mitochondria have been investigated.

MDR1基因多态性与耐药结核的相关性研究

目的对多药耐药基因1(MDR1)基因单核苷酸多态性与耐药结核的关系进行研究。方法采用病例对照研究,收集兰州市肺科医院2020年10月至2021年1月结核患者共97例,测序MDR1基因位点c.3435C>T(rs1045642)、c.1236C>T(rs1128503)和c.2677G>T/A(rs2032582)的单核苷酸多态性,统计后进行分析,比较耐药组与药物敏感组单核苷酸多态性的分布差异。结果在汉族人群耐药组与药物敏感组、利福平耐药组与利福平敏感组、乙胺丁醇耐药组和乙胺丁醇敏感组间分别比较MDR1基因rs10456423、rs1128503、rs2032582位点的基因型频率和等位基因频率,差异均无统计学意义(P>0.05)。结论MDR1基因单核苷酸多态性可能与汉族人群患耐药结核、乙胺丁醇耐药结核、利福平耐药结核的易感基因无关。

基因捕获元件int1和ISCR1在临床菌株中的分布及与细菌耐药性的关系研究

目的研究临床多种细菌中的Ⅰ型整合酶编码基因int1和ISCR1的分布情况,探究其与细菌多药耐药表型的关系。方法收集2011-2012年河北省某医院临床菌株,应用VITEK2COMPACT全自动微生物生化鉴定系统和16SrDNA序列分析进行种属鉴定,利用PCR方法进行int1基因和ISCR1筛查;两种可移动元件的携带情况和菌株的多药耐药表型之间的关系采用卡方检验进行统计计算。结果共收集临床菌株372株,包括325株革兰氏阴性菌和47株革兰氏阳性菌;int1基因和ISCR1在22种(属)175株和18种(属)90株革兰氏阴性细菌中检出,同时在17种(属)71株革兰氏阴性细菌中检出,革兰氏阳性细菌int1基因和ISCR1检测均为阴性;通过对数据分层对比,统计分析发现,在int1基因不存在时,ISCR1的存在与否与菌株是否为多药耐药的关系明显,差异有统计学意义(P<0.01),其存在亦能介导更广泛种类药物耐受,差异亦有统计学意义(P<0.01)。在int1基因存在时,ISCR1的存在能介导更广泛种类药物耐受,差异有统计学意义(P=0.02);在ISCR1不存在时,int1基因的存在与否与菌株是否多药耐药的关系明显,差异有统计学意义(P<0.01),其存在亦能介导更广泛种类药物耐受,差异亦具有统计学意义(P<0.01)。而在ISCR1存在的情况下,上述两种关系不明显。结论基因捕获元件int1基因及ISCR1在革兰氏阴性临床菌株中分布广泛,并与细菌的多重耐药、泛耐药明显相关,特别是ISCR1;应加强可移动元件int1基因或ISCR1流行状况的监测,为其在耐药基因捕获中的作用机制研究和控制多药耐药细菌在临床散播提供可靠的基础数据。

非M3型急性白血病患者中MUC1基因和MDR1基因表达及其与临床疗效的关系

背景与目的:MUC1基因在胃癌、卵巢癌、多发性骨髓瘤、恶性淋巴瘤等肿瘤中有表达,在急性白血病患者中有较高的表达。但MUC1基因和多药耐药基因(MDR1)相互关系以及两者的表达与急性白血病治疗效果的关系尚有待探讨。本研究拟探讨MUC1基因与MDR1基因表达及其与非M3型急性白血病患者治疗效果的关系。方法:应用逆转录鄄聚合酶链反应(RT鄄PCR)法检测34例初治非M3型急性白血病患者MUC1和MDR1的表达,并观察两种基因表达及其与临床疗效的关系。结果:34例初治非M3型急性白血病患者中MUC1基因阳性率为50%,MDR1基因阳性率为29.4%。MUC1基因阳性患者的MDR1阳性率为52.9%,明显高于MUC1阴性者的5.9%(P=0.003)。MUC1基因阴性者完全缓解(CR)率达94.1%,阳性患者CR率52.9%,两组有显著性差异(P<0.05);MDR1基因阴性者CR率为91.7%,明显高于阳性患者的50.0%(P<0.05)。MUC1基因和MDR1基因均阳性者CR率为55.6%,MUC1基因和MDR1基因均阴性者16例,全部获得CR。结论:非M3型急性白血病MUC1基因阳性者MDR1基因表达率较高,MUC1基因及MDR1基因均为阴性者治疗缓解率高。提示联合检测MUC1基因和MDR1基因对判断初治非M3型急性白血病的疗效有良好的预测作用,可作为临床判断疗效的一项有意义的指标。

靶向MRP1基因的shRNA稳定逆转肺癌的多药耐药性

目的:利用短发夹RNA(shRNA)表达载体逆转肺癌细胞株(A549/DDP)的多药耐药性。方法:构建2个多药耐药相关蛋白1(MRP1)基因特异的shRNA表达载体pSilencer 2.1-U6 neo-MRP1,稳定电转染A549/DDP细胞,实时荧光定量PCR分析MRP1 mRNA的表达,免疫荧光检测细胞MRP1蛋白的表达,流式细胞术检测细胞内罗丹明123(Rho123)的潴留情况。MTT法检测细胞活力。结果:成功构建了shRNA表达载体pSilencer 2.1-U6 neo-MRP1,稳定转染sh-MRP1-2.1-1和sh-MRP1-2.1-2后,A549/DDP细胞MRP1 mRNA和蛋白表达均显著降低,细胞内Rho123相对荧光强度由16.93%±0.58%分别升高至89.02%±0.59%和82.56%±1.37%;A549/DDP亲本细胞顺铂的IC50分别由(101.45±0.64)μmol/L降至(38.06±0.05)μmol/L和(53.72±0.36)μmol/L,5-氟尿嘧啶的IC50分别由(263.20±2.00)μmol/L降至(98.82±1.16)μmol/L和(141.81±0.49)μmol/L。结论:shRNA干扰表达载体pSilencer 2.1-U6 neo-RMP1能够稳定、持久地抑制MRP1基因,有效地逆转了A549/DDP细胞的多药耐药性。