Background:Previous research demonstrated that a homozygous mutation of g.136372044G>A(S12N)in caspase recruitment domain family member 9(CARD9)is critical for producing Aspergillus fumigatus-induced(Af-induced)T h...Background:Previous research demonstrated that a homozygous mutation of g.136372044G>A(S12N)in caspase recruitment domain family member 9(CARD9)is critical for producing Aspergillus fumigatus-induced(Af-induced)T helper 2(T_(H)2)-mediated responses in allergic bronchopulmonary aspergillosis(ABPA).However,it remains unclear whether the CARD9^(S12N)mutation,especially the heterozygous occurrence,predisposes the host to ABPA.Methods:A total of 61 ABPA patients and 264 controls(including 156 healthy controls and 108 asthma patients)were recruited for sequencing the CARD9 locus to clarify whether patients with this heterozygous single-nucleotide polymorphisms are predisposed to the development of ABPA.A series of in vivo and in vitro experiments,such as quantitative real-time polymerase chain reaction,flow cytometry,and RNA isolation and quantification,were used to illuminate the involved mechanism of the disease.Results:The presence of the p.S12N mutation was associated with a significant risk of ABPA in ABPA patients when compared with healthy controls and asthma patients,regardless of Aspergillus sensitivity.Relative to healthy controls without relevant allergies,the mutation of p.S12N was associated with a significant risk of ABPA(OR:2.69 and 4.17 for GA and AA genotypes,P=0.003 and 0.029,respectively).Compared with patients with asthma,ABPA patients had a significantly higher heterozygous mutation(GA genotype),indicating that p.S12N might be a significant ABPA-susceptibility locus(aspergillus sensitized asthma:OR:3.02,P=0.009;aspergillus unsensitized asthma:OR:2.94,P=0.005).The mutant allele was preferentially expressed in ABPA patients with heterozygous CARD9^(S12N),which contributes to its functional alterations to facilitate Af-induced T_(H)2-mediated ABPA development.In terms of mechanism,Card9 wild-type(Card9^(WT))expression levels decreased significantly due to Af-induced decay of its messenger RNA compared to the heterozygous Card9 S12N.In addition,ABPA patients with heterozygous CARD9^(S12N)had increased 展开更多
AIM:To investigate the polymorphic simple sequencerepeat in intron 1 of the epidermal growth factor receptorgene(EGFR)(CA-SSRⅠ),which is known to affect theefficiency of gene transcription as a putative target of the...AIM:To investigate the polymorphic simple sequencerepeat in intron 1 of the epidermal growth factor receptorgene(EGFR)(CA-SSRⅠ),which is known to affect theefficiency of gene transcription as a putative target of themismatch repair(MMR)machinery in colorectal tumors.METHODS:The CA-SSRⅠgenotype was analyzed ina total of 86 primary colorectal tumors,selected upontheir microsatellite instability(MSI)status[42 with highfrequency MSI(MSI-H)and 44 microsatellite stable(MSS)]and their respective normal tissue.The effect of the CA-SSRⅠgenotype on the expression of the EGFR gene wasevaluated in 18 specimens using quantitative real-timereverse transcription PCR and immunohistochemistry.RESULTS:Mutations in CA-SSRⅠwere detected in 86%(36 of 42)of MSI-H colorectal tumors and 0%(0 of 44)ofMSS tumors,indicating the EGFR gene as a novel putativespecific target of the defective MMR system(P<0.001).Impaired expression of EGFR was detected in most ofthe colorectal tumors analyzed[6/12(50%)at the mRNAlevel and 15/18(83%)at the peptide level].However,noassociation was apparent between EGFR expression andCA-SSRⅠstatus in tumors or normal tissues.CONCLUSION:Our results suggest that CA-SSRⅠsequence does not contribute to the regulation of EGFRtranscription in colon,and should thus not be consideredas a promising predictive marker for response to EGFRinhibitors in patients with colorectal cancer.展开更多
Objective: To study the features of microsatellite alterations on chromosome 8 and their asso- ciation with clinicopathological characteristics of hepatocellular carcinoma (HCC). Methods: Ten highly- ...Objective: To study the features of microsatellite alterations on chromosome 8 and their asso- ciation with clinicopathological characteristics of hepatocellular carcinoma (HCC). Methods: Ten highly- polymorphic microsatellite markers on chromosome 8 were selected to be detected for loss of heterozygosity (LOH), microsatellite instability (MSI) and allelic imbalance (AI) in 56 HCC using MegaBACE 500 auto- matic DNA analysis system. Results: LOH was found in 37 of 56 HCC (66.1%) on at least 1 locus, and the top three loci were D8S261(53.5%), D8S1721(52.5%) and D8S1771(52.5%). LOH frequency on D8S277 was signi?cantly higher in cases positive for serum HBsAg than in those negative for HBsAg (P <0.01). Similarly, LOH on D8S261, D8S298 and D8S1733 occurred more frequently in patients with negative HB- sAg than those with positive HBsAg (P <0.01). LOH on D8S298 and D8S1771 was more frequent in those tumors larger than 3 cm in size (P <0.05 or P <0.01). LOH frequencies of D8S1721 were signi?cantly higher in the patients with absent or not intact tumor capsule than in those with intact tumor capsule (P <0.05). LOH on D8S298 and D8S1771 was more frequently detected in tumors with intrahepatic metastasis than in those without intrahepatic metastasis (P <0.01). MSI was found in 12.5% (7/56) cases, and AI was found in 19.6% (11/56), no correlation was found between MSI and AI and clinicopathological character- istics of HCC. Conclusion: Frequent microsatellite alterations on chromosome 8 existed in HCC. LOH, which represents tumor suppressor gene pathway, plays a more important role in hepatocarcinogenesis; MSI representing mismatch repair gene pathway ranks next. LOH at a particula locus is associated with the clinicopathological parameters of human HCC.展开更多
AIM： To analyze the DNA copy number of target genes NF2, TIMP3, ST13, TOB2, BIK, and TP and the reference microsatellite markers D22S283, D22S423, and D22S274 mapped on 22q12-qter in liver fluke related cholangiocarc...AIM： To analyze the DNA copy number of target genes NF2, TIMP3, ST13, TOB2, BIK, and TP and the reference microsatellite markers D22S283, D22S423, and D22S274 mapped on 22q12-qter in liver fluke related cholangiocarcinoma （CCA） and define its correlation with clinical parameters. METHODS： Quantitative real time PCR （qPCR） was used for determining allelic imbalances in 65 liver fluke related CCA tissues. Statistical correlations between allelic imbalances and clinicopathological parameters, i.e. age, sex, tumor stage, histological type, blood vessel invasion, nerve invasion and lymphatic invasion were evaluated by means of the X^2 test. Cox regression analysis was used for determining patient＇s survival. RESULTS： Amplifications of the TP （22q13.33）, TOB2 （22q13.2-13.31）, D22S283 （22q12.3）, TIMP3 （22q12.3） and NF2 （22q12.2） were found in 35 （53.8%）, 28 （43.1%）, 27 （41.5%）, 24 （36.9%）, and 24 （36.9%）, respectively. Losses at the D22S423 （22q13.1-13.2）and BIK （22q13.31） were detected in 26 （40%） and 23 （35.4%）, respectively. Significant correlations were observed between lymphatic invasion and allelic losses of BIK （P = 0.025） and D22S283 （P = 0.041）. Univariate and multivariate Cox regression analysis revealed D22S283 amplification as an independent predictor of good prognosis （P = 0.006, death hazard ratio = 0.411, 95% CI = 0.217-0.779） and blood vessel invasion as an independent poor prognostic factor （P = 0.042, death hazard ratio = 1.911, 95% CI = 1.022-3.571） in CCA patients. CONCLUSION： This study provides evidence for the involvement of gene amplification and deletion on chromosome 22q in liver fluke related CCA, This is the first report of D22S283 amplification as an independent indicator of favorable prognosis in liver fluke related CCA.展开更多
AIM:To characterize and evaluate DNA alterations among intrahepatic cholangiocarcinoma (ICC) patients. METHODS:DNA from tumor and corresponding normal tissues of 52 patients was amplified with 33 arbitrary primers. Th...AIM:To characterize and evaluate DNA alterations among intrahepatic cholangiocarcinoma (ICC) patients. METHODS:DNA from tumor and corresponding normal tissues of 52 patients was amplified with 33 arbitrary primers. The DNA fragment that alters most frequently in ICC was cloned,sequenced,and identified by comparison with known nucleotide sequences in the genome database (www.ncbi.nlm.nih.gov). The DNA copy numbers of the allelic alterations in cholangiocarcinoma were determined by quantitative real-time PCR and interpreted as allelic loss or DNA amplification by comparison with the reference gene. Associations between allelic imbalance and clinicopathological parameters of ICC patients were evaluated by χ2-test. The Kaplan-Meier method was used to analyze survival rates. RESULTS:From 33 primers,an altered DNA fragment (518 bp) amplified from BC17 random primer was found frequently in the tumors analyzed and mapped to chromosome 17p13.2. Sixteen of 52 (31%) cases showed DNA amplification,while 7 (13%) showed allelic loss. Interestingly,DNA amplification on chromosome 17p13.2 was associated with a good prognosis,median survival time (wk) of amp vs no amp was 44.14 vs 24.14,P=0.002; whereas allelic loss of this DNA sequence corresponded with a poor prognosis,median survival time (wk) of loss vs no loss was 18.00 vs 28.71,P=0.019). Moreover,Kaplan-Meier curves comparing the DNA alterations with survival depicted highly significant separation that the median survival time equal to DNAamplification,allelic loss,and normal was 44.14 wk,18.00 wk,and 24.29 wk,respectively (P=0.005). CONCLUSION:Alterations in the DNA sequence on chromosome 17p13.2 may be involved in cholangio-carcinogenesis,and could be used as a prognostic marker in the treatment of ICC patients.展开更多
Objective： To investigate the expression of integrin α4 in osteosaxcoma and significance. Methods- Forty-six patients with osteosarcoma （Enneking Ⅰ-Ⅲ） were analyzed for the expression of α4 integrin subunit usi...Objective： To investigate the expression of integrin α4 in osteosaxcoma and significance. Methods- Forty-six patients with osteosarcoma （Enneking Ⅰ-Ⅲ） were analyzed for the expression of α4 integrin subunit using immunohistochemical method. Results： Twenty-nine （63.04%） of 46 samples demonstrated positive （＋-＋＋） integrin α4 expression. Loss expression of integrin α4 was observed in the patients with advanced Enneking stage （P=0.0040） and with metastatic disease at presentation （P=0.0158）. Integrin α4 expression correlated with cell differentiation, the level of malignancy and the invasive behavior of osteosaxcoma. Conclusion： The loss expression of integrin α4 subunit might be a predictor indicating the invasive potential of osteosarcoma and play a role in metastasis of osteosaxcoma patients.展开更多
基金supported by grants from the National Natural Science Foundation of China(Nos.81925001,81970036,and 31970889)the Innovation Program of Shanghai Municipal Education Commission(Nos.202101070007-E00097 and 201901070007E00022)+2 种基金the Program of Shanghai Municipal Science and Technology Commission(No.21DZ2201800)the Shanghai Municipal Health Commission(Nos.201740019 and ZY2018-2020 FWTX3022)Innovative Research Ream of High-Level Local Universities in Shanghai.
文摘Background:Previous research demonstrated that a homozygous mutation of g.136372044G>A(S12N)in caspase recruitment domain family member 9(CARD9)is critical for producing Aspergillus fumigatus-induced(Af-induced)T helper 2(T_(H)2)-mediated responses in allergic bronchopulmonary aspergillosis(ABPA).However,it remains unclear whether the CARD9^(S12N)mutation,especially the heterozygous occurrence,predisposes the host to ABPA.Methods:A total of 61 ABPA patients and 264 controls(including 156 healthy controls and 108 asthma patients)were recruited for sequencing the CARD9 locus to clarify whether patients with this heterozygous single-nucleotide polymorphisms are predisposed to the development of ABPA.A series of in vivo and in vitro experiments,such as quantitative real-time polymerase chain reaction,flow cytometry,and RNA isolation and quantification,were used to illuminate the involved mechanism of the disease.Results:The presence of the p.S12N mutation was associated with a significant risk of ABPA in ABPA patients when compared with healthy controls and asthma patients,regardless of Aspergillus sensitivity.Relative to healthy controls without relevant allergies,the mutation of p.S12N was associated with a significant risk of ABPA(OR:2.69 and 4.17 for GA and AA genotypes,P=0.003 and 0.029,respectively).Compared with patients with asthma,ABPA patients had a significantly higher heterozygous mutation(GA genotype),indicating that p.S12N might be a significant ABPA-susceptibility locus(aspergillus sensitized asthma:OR:3.02,P=0.009;aspergillus unsensitized asthma:OR:2.94,P=0.005).The mutant allele was preferentially expressed in ABPA patients with heterozygous CARD9^(S12N),which contributes to its functional alterations to facilitate Af-induced T_(H)2-mediated ABPA development.In terms of mechanism,Card9 wild-type(Card9^(WT))expression levels decreased significantly due to Af-induced decay of its messenger RNA compared to the heterozygous Card9 S12N.In addition,ABPA patients with heterozygous CARD9^(S12N)had increased
文摘AIM:To investigate the polymorphic simple sequencerepeat in intron 1 of the epidermal growth factor receptorgene(EGFR)(CA-SSRⅠ),which is known to affect theefficiency of gene transcription as a putative target of themismatch repair(MMR)machinery in colorectal tumors.METHODS:The CA-SSRⅠgenotype was analyzed ina total of 86 primary colorectal tumors,selected upontheir microsatellite instability(MSI)status[42 with highfrequency MSI(MSI-H)and 44 microsatellite stable(MSS)]and their respective normal tissue.The effect of the CA-SSRⅠgenotype on the expression of the EGFR gene wasevaluated in 18 specimens using quantitative real-timereverse transcription PCR and immunohistochemistry.RESULTS:Mutations in CA-SSRⅠwere detected in 86%(36 of 42)of MSI-H colorectal tumors and 0%(0 of 44)ofMSS tumors,indicating the EGFR gene as a novel putativespecific target of the defective MMR system(P<0.001).Impaired expression of EGFR was detected in most ofthe colorectal tumors analyzed[6/12(50%)at the mRNAlevel and 15/18(83%)at the peptide level].However,noassociation was apparent between EGFR expression andCA-SSRⅠstatus in tumors or normal tissues.CONCLUSION:Our results suggest that CA-SSRⅠsequence does not contribute to the regulation of EGFRtranscription in colon,and should thus not be consideredas a promising predictive marker for response to EGFRinhibitors in patients with colorectal cancer.
基金This project was supported by "the Hundred Leading Scientists Program of the Public Health Sector of Shanghai " (No. 98BR007), and "the National Science Foundation of China" (No. 30370645).
文摘Objective: To study the features of microsatellite alterations on chromosome 8 and their asso- ciation with clinicopathological characteristics of hepatocellular carcinoma (HCC). Methods: Ten highly- polymorphic microsatellite markers on chromosome 8 were selected to be detected for loss of heterozygosity (LOH), microsatellite instability (MSI) and allelic imbalance (AI) in 56 HCC using MegaBACE 500 auto- matic DNA analysis system. Results: LOH was found in 37 of 56 HCC (66.1%) on at least 1 locus, and the top three loci were D8S261(53.5%), D8S1721(52.5%) and D8S1771(52.5%). LOH frequency on D8S277 was signi?cantly higher in cases positive for serum HBsAg than in those negative for HBsAg (P <0.01). Similarly, LOH on D8S261, D8S298 and D8S1733 occurred more frequently in patients with negative HB- sAg than those with positive HBsAg (P <0.01). LOH on D8S298 and D8S1771 was more frequent in those tumors larger than 3 cm in size (P <0.05 or P <0.01). LOH frequencies of D8S1721 were signi?cantly higher in the patients with absent or not intact tumor capsule than in those with intact tumor capsule (P <0.05). LOH on D8S298 and D8S1771 was more frequently detected in tumors with intrahepatic metastasis than in those without intrahepatic metastasis (P <0.01). MSI was found in 12.5% (7/56) cases, and AI was found in 19.6% (11/56), no correlation was found between MSI and AI and clinicopathological character- istics of HCC. Conclusion: Frequent microsatellite alterations on chromosome 8 existed in HCC. LOH, which represents tumor suppressor gene pathway, plays a more important role in hepatocarcinogenesis; MSI representing mismatch repair gene pathway ranks next. LOH at a particula locus is associated with the clinicopathological parameters of human HCC.
基金Supported by The Thailand Research Fund through The Royal Golden Jubilee PhD Program, Grant No. PHD/0037/2544 for Thanasai J and Limpaiboon T
文摘AIM： To analyze the DNA copy number of target genes NF2, TIMP3, ST13, TOB2, BIK, and TP and the reference microsatellite markers D22S283, D22S423, and D22S274 mapped on 22q12-qter in liver fluke related cholangiocarcinoma （CCA） and define its correlation with clinical parameters. METHODS： Quantitative real time PCR （qPCR） was used for determining allelic imbalances in 65 liver fluke related CCA tissues. Statistical correlations between allelic imbalances and clinicopathological parameters, i.e. age, sex, tumor stage, histological type, blood vessel invasion, nerve invasion and lymphatic invasion were evaluated by means of the X^2 test. Cox regression analysis was used for determining patient＇s survival. RESULTS： Amplifications of the TP （22q13.33）, TOB2 （22q13.2-13.31）, D22S283 （22q12.3）, TIMP3 （22q12.3） and NF2 （22q12.2） were found in 35 （53.8%）, 28 （43.1%）, 27 （41.5%）, 24 （36.9%）, and 24 （36.9%）, respectively. Losses at the D22S423 （22q13.1-13.2）and BIK （22q13.31） were detected in 26 （40%） and 23 （35.4%）, respectively. Significant correlations were observed between lymphatic invasion and allelic losses of BIK （P = 0.025） and D22S283 （P = 0.041）. Univariate and multivariate Cox regression analysis revealed D22S283 amplification as an independent predictor of good prognosis （P = 0.006, death hazard ratio = 0.411, 95% CI = 0.217-0.779） and blood vessel invasion as an independent poor prognostic factor （P = 0.042, death hazard ratio = 1.911, 95% CI = 1.022-3.571） in CCA patients. CONCLUSION： This study provides evidence for the involvement of gene amplification and deletion on chromosome 22q in liver fluke related CCA, This is the first report of D22S283 amplification as an independent indicator of favorable prognosis in liver fluke related CCA.
基金Rajabhat University Suan Dusit FundFaculty of Tropical Medicine,Mahidol UniversityDAAD scholarship through Southeast Asian Ministers of Education Organization,Regional Tropical Medicine and Public Health Network
文摘AIM:To characterize and evaluate DNA alterations among intrahepatic cholangiocarcinoma (ICC) patients. METHODS:DNA from tumor and corresponding normal tissues of 52 patients was amplified with 33 arbitrary primers. The DNA fragment that alters most frequently in ICC was cloned,sequenced,and identified by comparison with known nucleotide sequences in the genome database (www.ncbi.nlm.nih.gov). The DNA copy numbers of the allelic alterations in cholangiocarcinoma were determined by quantitative real-time PCR and interpreted as allelic loss or DNA amplification by comparison with the reference gene. Associations between allelic imbalance and clinicopathological parameters of ICC patients were evaluated by χ2-test. The Kaplan-Meier method was used to analyze survival rates. RESULTS:From 33 primers,an altered DNA fragment (518 bp) amplified from BC17 random primer was found frequently in the tumors analyzed and mapped to chromosome 17p13.2. Sixteen of 52 (31%) cases showed DNA amplification,while 7 (13%) showed allelic loss. Interestingly,DNA amplification on chromosome 17p13.2 was associated with a good prognosis,median survival time (wk) of amp vs no amp was 44.14 vs 24.14,P=0.002; whereas allelic loss of this DNA sequence corresponded with a poor prognosis,median survival time (wk) of loss vs no loss was 18.00 vs 28.71,P=0.019). Moreover,Kaplan-Meier curves comparing the DNA alterations with survival depicted highly significant separation that the median survival time equal to DNAamplification,allelic loss,and normal was 44.14 wk,18.00 wk,and 24.29 wk,respectively (P=0.005). CONCLUSION:Alterations in the DNA sequence on chromosome 17p13.2 may be involved in cholangio-carcinogenesis,and could be used as a prognostic marker in the treatment of ICC patients.
文摘Objective： To investigate the expression of integrin α4 in osteosaxcoma and significance. Methods- Forty-six patients with osteosarcoma （Enneking Ⅰ-Ⅲ） were analyzed for the expression of α4 integrin subunit using immunohistochemical method. Results： Twenty-nine （63.04%） of 46 samples demonstrated positive （＋-＋＋） integrin α4 expression. Loss expression of integrin α4 was observed in the patients with advanced Enneking stage （P=0.0040） and with metastatic disease at presentation （P=0.0158）. Integrin α4 expression correlated with cell differentiation, the level of malignancy and the invasive behavior of osteosaxcoma. Conclusion： The loss expression of integrin α4 subunit might be a predictor indicating the invasive potential of osteosarcoma and play a role in metastasis of osteosaxcoma patients.
