AIM: To analyze the correlation between the protein expression of p16 and Rb genes in gastric carcinoma (GC),to investigate the role of p16 gene in invasion and lymph node metastasis of GC, and to examine the deletion...AIM: To analyze the correlation between the protein expression of p16 and Rb genes in gastric carcinoma (GC),to investigate the role of p16 gene in invasion and lymph node metastasis of GC, and to examine the deletion and mutation in exon 2 of p16 gene in GC.METHODS: The protein expression of p16 and Rb genes was examined by streptavidin-peroxidase conjugated method (S-P) in normal gastric mucosa, dysplastic gastric mucosa and GC. The deletion and mutation of p16 gene were examined by polymerase chain reaction (PCR) and polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) respectively in normal gastric mucosa and GC.RESULTS: The positive rates of P16 and Rb protein expression respectively were 96% (77/80) and 99%(79/80) in normal gastric mucosa, 92% (45/50) and 80%(40/50) in dysplastic gastric mucosa, 48% (58/122) and 60% (73/122) in GC. The positive rates of P16 and Rb protein expression in GC were significantly lower than that in normal gastric mucosa and dysplastic gastric mucosa (P＜0.05). The positive rate of P16 protein expression in mucoid carcinoma (10%, 1/10) was significantly lower than that in poorly differentiated carcinoma (51%, 21/41),undifferentiated carcinoma (58%, 15/26) and signet ring cell carcinoma (62%, 10/16) (P＜0.05). The positive rates of P16 protein in 30 cases of paired primary and lymph node metastatic GC were 47% (14/30) and 17% (5/30)respectively, being significantly lower in the later than in the former (P＜0.05). There was no mutation in exon 2 of p16 gene in the 25 freshly resected primary GCs. But five cases in the 25 freshly resected primary GCs displayed deletion in exon 2 of p16 gene. The positive rate of both P16 and Rb proteins was 16% (14/90), and the negative rate of both P16 and Rb proteins was 8% (7/90) in 90GCs. The rate of positive P16 protein with negative Rb protein was 33% (30/90). The rate of negative P16 protein with positive Rb protein was 43% (39/90). There was reverse correlation between P16 and Rb expression in 90GCs (P＜0.05).CONCLUSION; The loss protein expression of p16 and Rb genes is related to GC. The loss expression of P16protein is related to the histopathologic subtypes and lymph node metastasis of GC. Expression of P16 and Rb proteins in GC is reversely correlated. The deletion but not mutation in exon 2 of p16 gene may be involved in GC.展开更多
Objective To study the alterations of FHIT gene and P16 gene in malignant transformed human bronchial epithelial cells induced by crystalline nickel sulfide using an immortal human bronchial epithelial cell line, and ...Objective To study the alterations of FHIT gene and P16 gene in malignant transformed human bronchial epithelial cells induced by crystalline nickel sulfide using an immortal human bronchial epithelial cell line, and to explore the molecular mechanism of nickel carcinogenesis. Methods 16HBE cells were treated 6 times with different concentrations of NiS in vitro, and the degree of malignant transformation was determined by assaying the anchorage-independent growth and tumorigenicity. Malignant transformed cells and tumorigenic cells were examined for alterations of FHIT gene and P16 gene using RT-PCR, DNA sequencing, silver staining PCR-SSCP and Western blotting. Results NiS-treated cells exhibited overlapping growth. Compared with that of negative control cells, soft agar colony formation efficiency of NiS-treated cells showed significant increases (P<0.01) and dose-dependent effects. NiS-treated cells could form tumors in nude mice, and a squamous cell carcinoma was confirmed by histopathological examination. No mutation of exon 2 and exons 2-3, no abnormal expression in p16 gene and mutation of FHIT exons 5-8 and exons 1-4 or exons 5-9 were observed in transformed cells and tumorigenic cells. However, aberrant transcripts or loss of expression of the FHIT gene and Fhit protein was observed in transformed cells and tumorigenic cells. One of the aberrant transcripts in the FHIT gene was confirmed to have a deletion of exon 6, exon 7, exon 8, and an insertion of a 36 bp sequence replacing exon 6-8. Conclusions The FHIT gene rather than the P16 gene, plays a definite role in nickel carcinogenesis. Alterations of the FHIT gene induced by crystalline NiS may be a molecular event associated with carcinogen, chromosome fragile site instability and cell malignant transformation. FHIT may be an important target gene activated by nickel and other exotic carcinogens.展开更多
Objective:To investigate the relationship between the genetic polymorphism of CYP1A1 and the genetic susceptibility to lung cancer as well as to study the effects of the methylation in p16 gene on the risk of lung can...Objective:To investigate the relationship between the genetic polymorphism of CYP1A1 and the genetic susceptibility to lung cancer as well as to study the effects of the methylation in p16 gene on the risk of lung cancer in a Chinese population.Methods:A case control study was conducted among 47 cases of lung cancer and 94 controls.The genetic polymorphism of CYP1A1 was tested with method of PCR-RFLP,and a methylation-specific PCR(MSP)was performed to detect p16 methylation.Results:It showed that there was no significant difference in frequencies of the genotypes of CYP1A1 between the two groups(P>0.05).Synergistic effects were not found between smoking and CYP1A1.Methylated p16 gene was found in 44.7%(21/47)of lung cancer tissues and in 17.0%(8/47)of normal lung tissues with significant difference(P< 0.05).Conclusion:The genetic polymorphism of CYP1A1 does not increase the risk of lung cancer in a Chinese population. The methylation in p16 gene may be the most common mechanism to inactivate p16 gene in lung cancer,and is not significantly associated with genotype of CYP1A1.展开更多
Objective: To analyze the aberrant methylation of p16 gene and DAPK gene in sera from primary liver cancer patients ad to evaluate the clinical significance. Methods: A methylation-specific PCR was performed for the d...Objective: To analyze the aberrant methylation of p16 gene and DAPK gene in sera from primary liver cancer patients ad to evaluate the clinical significance. Methods: A methylation-specific PCR was performed for the detection of promoter hypermethylation of p16 gene and DAPK gene in blood DNA from 64 cases of HCC patients, and to analyze the relation of the aberrant methylation of p16 gene and KAPK gene and the clinical pathological data. Results: 76.6%(49/64) of the sera from 64 cases of HCC patients showed hypermethylation for p16 promoter and 40.6% (26/64) for KAPK promoter, whereas no methylated p16 gene promoter and DAPK gene promoter were found in sera from benign liver diseases patients and normal control. Methylated p16 gene and KAPK gene promoters in sera did not strongly correlated with HBsAg, stage, metastasis and differentiation in HCC; but strongly correlated with AFP. Conclusion: Detection of the aberrant methylation of p16 gene and KAPK gene in blood DNA from HCC patients might offer an effective means for the earlier auxiliary diagnosis of the malignancy.展开更多
AIM To explore the association of methylation of the CpG island in the promotor of the p16 tumor suppressor gene with the clinicopathological characteristics of the colorectal cancers.``METHODS Methylation-specific PC...AIM To explore the association of methylation of the CpG island in the promotor of the p16 tumor suppressor gene with the clinicopathological characteristics of the colorectal cancers.``METHODS Methylation-specific PCR (MSP) was used to detect p16 methylation of 62 sporadic colorectal cancer specimens.``RESULTS pi6 methylation was detected in 42% of the tumors. Dukes staging was associated with p16methylation status, p16 methylation occurred more frequently in Dukes C and D patients (75.9%) than in Dukes A and B patients (12.1%).``CONCLUSION p16 rnethylation plays a role in the carcinogenesis of a subset of colorectal cancer, and it might be linked to poor prognosis.展开更多
Objective: To detect promoter hypermethylation of p16 gene in matched pre- and post-operative plasma of patients with gastric adenocarcinoma for evaluating the effectiveness of therapeutic intervention. Methods: Tissu...Objective: To detect promoter hypermethylation of p16 gene in matched pre- and post-operative plasma of patients with gastric adenocarcinoma for evaluating the effectiveness of therapeutic intervention. Methods: Tissue samples, pre- and post-operative plasma of 84 patients were collected. Plasma of 15 healthy people was collected as control. After sodium-bisulfite treatment, extracted DNA was amplified for p16 promoter by methylation-specific polymerase chain reaction (MSP). The PCR products were detected by both gel-ethidium bromide electrophoresis and high performance liquid chromatogram (HPLC). Results: Among 84 patients, p16 hypermethylation was detected in 26 (31.0%) cancer tissues and 2 (0.02%) tumor-adjacent tissues and 12 (14.3%) pre-operative plasma, while negative in plasma of healthy people. For positive plasma cases, the paired tumor tissues were confirmed to be methylated. Within available 30 pairs of matched pre- and post-operative plasma, 6 pre-operative plasma was positive, and only 1 of 6 plasma remained hypermethylated after surgery. The results detected by HPLC exactly matched those by gel-electrophoresis. Conclusion: The alteration of status of p16 hypermethylation in post-operative plasma is considered the consequences of surgical intervention. Although p16 hypermethylation has no role in pre-operative staging of gastric cancer, detecting hypermethylated p16 in plasma could be utilized in monitoring patients after surgery.展开更多
Objective: To examine the occurrence of p16 gene deletion and to analyze p16 expression on paraffinembedded human pituitary adenoma specimens. Efforts were made to optimize the technical conditions for in situ PCR. Me...Objective: To examine the occurrence of p16 gene deletion and to analyze p16 expression on paraffinembedded human pituitary adenoma specimens. Efforts were made to optimize the technical conditions for in situ PCR. Methods: In situ PCR techniques and inimunohistochemistry were used. Results: Immunohistochemically, p16-positive tumor cells were observed in all cases with various proportions. The majority of the stromal cells and part of tumor cells was devoid of p16 immunostaining, but signal of in situ PCR for p16 gene, exon 2, was displayed in these cells. Conclusion: The results implied that pl6 gene might not be deleted in these pituitary adenomas. It also indicated that in situ PCR, both direct and indirect methods, proved feasible and informative to the aim of DNA detection. It is critical to overcome non-specific amplification in direct in situ PCR by means of higher annealing temperature, fewer cycle, lower magnesium concentration and stringent washing. A target DNA-deleted sample as the negative control is extremely necessary. For the indirect method, the way to improve the sensitivity is to loosen the conditions for amplification and washing, so that more amplification products are subject to hybridization, and signal detection is facilitated.展开更多
To stddy the chang of suppressing cancer gene P16 in acute leukemia, the P16 antigen expression of leukemia cell surfaces in 61 cases were investigated with ABC assay and gene structural defects in 51 cases of acute l...To stddy the chang of suppressing cancer gene P16 in acute leukemia, the P16 antigen expression of leukemia cell surfaces in 61 cases were investigated with ABC assay and gene structural defects in 51 cases of acute leukemia were examined with multiple comparative PCR method. It was found that antigen expression of P16 in leukemia was obviously lower than that innormal subjects ( P <0 001). At the same time, antigen expression in All was lower than that AML ( P <0 05). No significant difference was found betwee the complete reission (CR) and non remission (NR) subjects from AML and ALL groups ( P >0 05). THe exon 2 of P16 gene showed homozygous deletion only inn4 cases out of 30 cases in ALL. No stuctural defect was revealed in 21 cases of AML. It was suggested that expression defect of P16 gene was a main cause in development and progression of acute leukemia, and structural defect of exon 2 was not a primary molecular event.展开更多
In this study, we infected human glioma U251 cells with a replication-defective recombinant adeno-virus carrying the p16 gene. This adenovirus constructed was able to transfect exogenous p16 into the human glioma cell...In this study, we infected human glioma U251 cells with a replication-defective recombinant adeno-virus carrying the p16 gene. This adenovirus constructed was able to transfect exogenous p16 into the human glioma cells efficiently, and direct a high level of p16 protein expression. Tumor-inhibition experiments demonstrated that treatment with the adenovirus-p16 significantly inhibited the growth of glioma cells in vitro as well as the in vivo development of tumors in nude mice bearing a brain glioma. The combination of adenovirus-p16 gene treatment and X-ray irradiation resulted in a greater inhibition of tumor growth. Adenovirus-mediated p16 gene therapy conferred a significant antitumor effect against human glioma cells both in vitro and in vivo, and that there was a synergistic effect when X-ray irradiation was also used.展开更多
AIM: To further understand the molecular basis for gastriccardia carcinogenesis and to provide etiological clues.METHODS: Endoscopic mucosa biopsy and histopathologicalexaminations were made on 37 subjects from a high...AIM: To further understand the molecular basis for gastriccardia carcinogenesis and to provide etiological clues.METHODS: Endoscopic mucosa biopsy and histopathologicalexaminations were made on 37 subjects from a high incidencearea for both esophageal and gastric cardia carcinomas innorthem China. All the biopsy samples were fixed in 850 mi. -1 Lalcohol and embedded in paraffin. Each block contained onepiece of tissue and was serially section at 5 μm.Immunohistochemistry (ABC) was carried out on these gastriccardia samples to determine the alterations of p16 and Rb.RESULTS: Based on the histopathlogical examinationtherewere 11 cases of chronic superficial gastritis, 12 cases ofchronic atrophic gastritis and 14 cases of dysplasia. Theimmunostaining demonstrated different levels of unclearimmunostaining of p16 and Rb in normal gastric cardiatissue and the tissues with different severity of lesions. Withthe lesions progressing, the positive immunostaining ratesfor pi6 protein had a decreasing tendency. In contrast, thepositive immunostaining rate for Rb protein had anincreasing tendency. There was a significant negativerelationship between the two parameters. Changes of p16wasCSG 11(100 % ), CAG 7(58 % ), DYS 4(29 % ) andchanges of Rb was CSG 2(18 %), CAG 8(67 %) and DYS 12(86 %), (p＜0.05).CONCLUSION: The alterations of p16 and Rb protein may playa role in the early stages of gastric cardia carcinogenesis.展开更多
The effects of exogenous p16ink4a gene on biological behaviors of human lung cancer cell line with homozygous deletion of p16ink4a gene were investigated. Exogenous p16ink4a gene was transfected by lipofectin into hum...The effects of exogenous p16ink4a gene on biological behaviors of human lung cancer cell line with homozygous deletion of p16ink4a gene were investigated. Exogenous p16ink4a gene was transfected by lipofectin into human lung cell line A549, in which p16ink4a gene was homozygously deleted. The expression of p16ink4a mRNA and protein was detected by RT-PCR and immunocyto-chemistry, respectively. The changes in the behaviors of the transfected cell lines in vitro and in vivo were observed. In the transfected cell line A549, the exogenous p16ink4a gene could be stably ex-pressed. The growth of A549 cells transfected with p16ink4a gene was obviously slowed down. Flow cytometry revealed that transfection of the exogenous p16ink4a gene resulted in A549 cell lines arrest in G1 phase of cell cycle. The tumorigenicity of these transfected cells in nude mice could be inhib-ited, and the tumor growth of nude mice was significantly suppressed. It was concluded that exoge-nous p16ink4a gene may be stably expressed in human lung cancer cell line A549. The expression of the introduced p16ink4a could block lung cancer cells to entry into S phase of cell cycle and inhibit tumor malignant growth both in vitro and in vivo.展开更多
Objective: To determine the effects on the cell growth, tumorigenicity and chemosensitivity of p16/CDK4I in human glioma. Methods: p16 gene was transfected into U251 cells by lipofectin. Expression of exogenous p16 ge...Objective: To determine the effects on the cell growth, tumorigenicity and chemosensitivity of p16/CDK4I in human glioma. Methods: p16 gene was transfected into U251 cells by lipofectin. Expression of exogenous p16 gene was confirmed by immunohistochemistry and Northern blot. The effects of exogenous p16 gene on the growth and chemosensitivity to teniposide were examined. Results: Expression of exogenous p16 gene inhibited the growth dramatically in vitro. G1 arrest of tumor cells was observed. However, wt p16-positive U251 was less sensitive than control cell lines and the number of apoptotic cells after chemotherapy was reduced. Conclusion: The expression of exogenous p16 gene could inhibit the growth of glioma. On the other hand, the chemosensitivity to teniposide of p16-positive U251 was decreased.展开更多
基金Supported by the Grant From the Education Committee of Hunan Province, No. 97B095, No. 01B016 the grant from the Health Bureau of Hunan Province, No. 9301, the Key Programs during the 8th 5-Year Plan Period, the Bureau of Health, Hunan Province, China
文摘AIM: To analyze the correlation between the protein expression of p16 and Rb genes in gastric carcinoma (GC),to investigate the role of p16 gene in invasion and lymph node metastasis of GC, and to examine the deletion and mutation in exon 2 of p16 gene in GC.METHODS: The protein expression of p16 and Rb genes was examined by streptavidin-peroxidase conjugated method (S-P) in normal gastric mucosa, dysplastic gastric mucosa and GC. The deletion and mutation of p16 gene were examined by polymerase chain reaction (PCR) and polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) respectively in normal gastric mucosa and GC.RESULTS: The positive rates of P16 and Rb protein expression respectively were 96% (77/80) and 99%(79/80) in normal gastric mucosa, 92% (45/50) and 80%(40/50) in dysplastic gastric mucosa, 48% (58/122) and 60% (73/122) in GC. The positive rates of P16 and Rb protein expression in GC were significantly lower than that in normal gastric mucosa and dysplastic gastric mucosa (P＜0.05). The positive rate of P16 protein expression in mucoid carcinoma (10%, 1/10) was significantly lower than that in poorly differentiated carcinoma (51%, 21/41),undifferentiated carcinoma (58%, 15/26) and signet ring cell carcinoma (62%, 10/16) (P＜0.05). The positive rates of P16 protein in 30 cases of paired primary and lymph node metastatic GC were 47% (14/30) and 17% (5/30)respectively, being significantly lower in the later than in the former (P＜0.05). There was no mutation in exon 2 of p16 gene in the 25 freshly resected primary GCs. But five cases in the 25 freshly resected primary GCs displayed deletion in exon 2 of p16 gene. The positive rate of both P16 and Rb proteins was 16% (14/90), and the negative rate of both P16 and Rb proteins was 8% (7/90) in 90GCs. The rate of positive P16 protein with negative Rb protein was 33% (30/90). The rate of negative P16 protein with positive Rb protein was 43% (39/90). There was reverse correlation between P16 and Rb expression in 90GCs (P＜0.05).CONCLUSION; The loss protein expression of p16 and Rb genes is related to GC. The loss expression of P16protein is related to the histopathologic subtypes and lymph node metastasis of GC. Expression of P16 and Rb proteins in GC is reversely correlated. The deletion but not mutation in exon 2 of p16 gene may be involved in GC.
基金This work was supported by a grant (No. 39170651 and 30200235) from National Natural Science Foundation of China.
文摘Objective To study the alterations of FHIT gene and P16 gene in malignant transformed human bronchial epithelial cells induced by crystalline nickel sulfide using an immortal human bronchial epithelial cell line, and to explore the molecular mechanism of nickel carcinogenesis. Methods 16HBE cells were treated 6 times with different concentrations of NiS in vitro, and the degree of malignant transformation was determined by assaying the anchorage-independent growth and tumorigenicity. Malignant transformed cells and tumorigenic cells were examined for alterations of FHIT gene and P16 gene using RT-PCR, DNA sequencing, silver staining PCR-SSCP and Western blotting. Results NiS-treated cells exhibited overlapping growth. Compared with that of negative control cells, soft agar colony formation efficiency of NiS-treated cells showed significant increases (P<0.01) and dose-dependent effects. NiS-treated cells could form tumors in nude mice, and a squamous cell carcinoma was confirmed by histopathological examination. No mutation of exon 2 and exons 2-3, no abnormal expression in p16 gene and mutation of FHIT exons 5-8 and exons 1-4 or exons 5-9 were observed in transformed cells and tumorigenic cells. However, aberrant transcripts or loss of expression of the FHIT gene and Fhit protein was observed in transformed cells and tumorigenic cells. One of the aberrant transcripts in the FHIT gene was confirmed to have a deletion of exon 6, exon 7, exon 8, and an insertion of a 36 bp sequence replacing exon 6-8. Conclusions The FHIT gene rather than the P16 gene, plays a definite role in nickel carcinogenesis. Alterations of the FHIT gene induced by crystalline NiS may be a molecular event associated with carcinogen, chromosome fragile site instability and cell malignant transformation. FHIT may be an important target gene activated by nickel and other exotic carcinogens.
基金a grant from the National Natural Sciences Foundationof China(No.30471427).
文摘Objective:To investigate the relationship between the genetic polymorphism of CYP1A1 and the genetic susceptibility to lung cancer as well as to study the effects of the methylation in p16 gene on the risk of lung cancer in a Chinese population.Methods:A case control study was conducted among 47 cases of lung cancer and 94 controls.The genetic polymorphism of CYP1A1 was tested with method of PCR-RFLP,and a methylation-specific PCR(MSP)was performed to detect p16 methylation.Results:It showed that there was no significant difference in frequencies of the genotypes of CYP1A1 between the two groups(P>0.05).Synergistic effects were not found between smoking and CYP1A1.Methylated p16 gene was found in 44.7%(21/47)of lung cancer tissues and in 17.0%(8/47)of normal lung tissues with significant difference(P< 0.05).Conclusion:The genetic polymorphism of CYP1A1 does not increase the risk of lung cancer in a Chinese population. The methylation in p16 gene may be the most common mechanism to inactivate p16 gene in lung cancer,and is not significantly associated with genotype of CYP1A1.
文摘Objective: To analyze the aberrant methylation of p16 gene and DAPK gene in sera from primary liver cancer patients ad to evaluate the clinical significance. Methods: A methylation-specific PCR was performed for the detection of promoter hypermethylation of p16 gene and DAPK gene in blood DNA from 64 cases of HCC patients, and to analyze the relation of the aberrant methylation of p16 gene and KAPK gene and the clinical pathological data. Results: 76.6%(49/64) of the sera from 64 cases of HCC patients showed hypermethylation for p16 promoter and 40.6% (26/64) for KAPK promoter, whereas no methylated p16 gene promoter and DAPK gene promoter were found in sera from benign liver diseases patients and normal control. Methylated p16 gene and KAPK gene promoters in sera did not strongly correlated with HBsAg, stage, metastasis and differentiation in HCC; but strongly correlated with AFP. Conclusion: Detection of the aberrant methylation of p16 gene and KAPK gene in blood DNA from HCC patients might offer an effective means for the earlier auxiliary diagnosis of the malignancy.
基金Supported by the grants from Mjnistry of Public Health of China,No.98-1-303The Educational Committee of Shanghai,No.2000B02.
文摘AIM To explore the association of methylation of the CpG island in the promotor of the p16 tumor suppressor gene with the clinicopathological characteristics of the colorectal cancers.``METHODS Methylation-specific PCR (MSP) was used to detect p16 methylation of 62 sporadic colorectal cancer specimens.``RESULTS pi6 methylation was detected in 42% of the tumors. Dukes staging was associated with p16methylation status, p16 methylation occurred more frequently in Dukes C and D patients (75.9%) than in Dukes A and B patients (12.1%).``CONCLUSION p16 rnethylation plays a role in the carcinogenesis of a subset of colorectal cancer, and it might be linked to poor prognosis.
基金This work was supported in part byScience Research Foundation of Peking University Schoolof Oncology(00-01and04-01)
文摘Objective: To detect promoter hypermethylation of p16 gene in matched pre- and post-operative plasma of patients with gastric adenocarcinoma for evaluating the effectiveness of therapeutic intervention. Methods: Tissue samples, pre- and post-operative plasma of 84 patients were collected. Plasma of 15 healthy people was collected as control. After sodium-bisulfite treatment, extracted DNA was amplified for p16 promoter by methylation-specific polymerase chain reaction (MSP). The PCR products were detected by both gel-ethidium bromide electrophoresis and high performance liquid chromatogram (HPLC). Results: Among 84 patients, p16 hypermethylation was detected in 26 (31.0%) cancer tissues and 2 (0.02%) tumor-adjacent tissues and 12 (14.3%) pre-operative plasma, while negative in plasma of healthy people. For positive plasma cases, the paired tumor tissues were confirmed to be methylated. Within available 30 pairs of matched pre- and post-operative plasma, 6 pre-operative plasma was positive, and only 1 of 6 plasma remained hypermethylated after surgery. The results detected by HPLC exactly matched those by gel-electrophoresis. Conclusion: The alteration of status of p16 hypermethylation in post-operative plasma is considered the consequences of surgical intervention. Although p16 hypermethylation has no role in pre-operative staging of gastric cancer, detecting hypermethylated p16 in plasma could be utilized in monitoring patients after surgery.
基金a grant from Ministry of Public Health of China!(98-l-303)
文摘Objective: To examine the occurrence of p16 gene deletion and to analyze p16 expression on paraffinembedded human pituitary adenoma specimens. Efforts were made to optimize the technical conditions for in situ PCR. Methods: In situ PCR techniques and inimunohistochemistry were used. Results: Immunohistochemically, p16-positive tumor cells were observed in all cases with various proportions. The majority of the stromal cells and part of tumor cells was devoid of p16 immunostaining, but signal of in situ PCR for p16 gene, exon 2, was displayed in these cells. Conclusion: The results implied that pl6 gene might not be deleted in these pituitary adenomas. It also indicated that in situ PCR, both direct and indirect methods, proved feasible and informative to the aim of DNA detection. It is critical to overcome non-specific amplification in direct in situ PCR by means of higher annealing temperature, fewer cycle, lower magnesium concentration and stringent washing. A target DNA-deleted sample as the negative control is extremely necessary. For the indirect method, the way to improve the sensitivity is to loosen the conditions for amplification and washing, so that more amplification products are subject to hybridization, and signal detection is facilitated.
文摘To stddy the chang of suppressing cancer gene P16 in acute leukemia, the P16 antigen expression of leukemia cell surfaces in 61 cases were investigated with ABC assay and gene structural defects in 51 cases of acute leukemia were examined with multiple comparative PCR method. It was found that antigen expression of P16 in leukemia was obviously lower than that innormal subjects ( P <0 001). At the same time, antigen expression in All was lower than that AML ( P <0 05). No significant difference was found betwee the complete reission (CR) and non remission (NR) subjects from AML and ALL groups ( P >0 05). THe exon 2 of P16 gene showed homozygous deletion only inn4 cases out of 30 cases in ALL. No stuctural defect was revealed in 21 cases of AML. It was suggested that expression defect of P16 gene was a main cause in development and progression of acute leukemia, and structural defect of exon 2 was not a primary molecular event.
基金Science and Technology Fund Program of Shaanxi Province, No. 2002K10-G3Xi'an Jiaotong University Innovation Fund, No. 0203207
文摘In this study, we infected human glioma U251 cells with a replication-defective recombinant adeno-virus carrying the p16 gene. This adenovirus constructed was able to transfect exogenous p16 into the human glioma cells efficiently, and direct a high level of p16 protein expression. Tumor-inhibition experiments demonstrated that treatment with the adenovirus-p16 significantly inhibited the growth of glioma cells in vitro as well as the in vivo development of tumors in nude mice bearing a brain glioma. The combination of adenovirus-p16 gene treatment and X-ray irradiation resulted in a greater inhibition of tumor growth. Adenovirus-mediated p16 gene therapy conferred a significant antitumor effect against human glioma cells both in vitro and in vivo, and that there was a synergistic effect when X-ray irradiation was also used.
基金the National Natural Science Foundation of China,No,39770296
文摘AIM: To further understand the molecular basis for gastriccardia carcinogenesis and to provide etiological clues.METHODS: Endoscopic mucosa biopsy and histopathologicalexaminations were made on 37 subjects from a high incidencearea for both esophageal and gastric cardia carcinomas innorthem China. All the biopsy samples were fixed in 850 mi. -1 Lalcohol and embedded in paraffin. Each block contained onepiece of tissue and was serially section at 5 μm.Immunohistochemistry (ABC) was carried out on these gastriccardia samples to determine the alterations of p16 and Rb.RESULTS: Based on the histopathlogical examinationtherewere 11 cases of chronic superficial gastritis, 12 cases ofchronic atrophic gastritis and 14 cases of dysplasia. Theimmunostaining demonstrated different levels of unclearimmunostaining of p16 and Rb in normal gastric cardiatissue and the tissues with different severity of lesions. Withthe lesions progressing, the positive immunostaining ratesfor pi6 protein had a decreasing tendency. In contrast, thepositive immunostaining rate for Rb protein had anincreasing tendency. There was a significant negativerelationship between the two parameters. Changes of p16wasCSG 11(100 % ), CAG 7(58 % ), DYS 4(29 % ) andchanges of Rb was CSG 2(18 %), CAG 8(67 %) and DYS 12(86 %), (p＜0.05).CONCLUSION: The alterations of p16 and Rb protein may playa role in the early stages of gastric cardia carcinogenesis.
基金This project was supported by a grant from the Science Research Foundation of Hubei Province, China (No. 98J102).
文摘The effects of exogenous p16ink4a gene on biological behaviors of human lung cancer cell line with homozygous deletion of p16ink4a gene were investigated. Exogenous p16ink4a gene was transfected by lipofectin into human lung cell line A549, in which p16ink4a gene was homozygously deleted. The expression of p16ink4a mRNA and protein was detected by RT-PCR and immunocyto-chemistry, respectively. The changes in the behaviors of the transfected cell lines in vitro and in vivo were observed. In the transfected cell line A549, the exogenous p16ink4a gene could be stably ex-pressed. The growth of A549 cells transfected with p16ink4a gene was obviously slowed down. Flow cytometry revealed that transfection of the exogenous p16ink4a gene resulted in A549 cell lines arrest in G1 phase of cell cycle. The tumorigenicity of these transfected cells in nude mice could be inhib-ited, and the tumor growth of nude mice was significantly suppressed. It was concluded that exoge-nous p16ink4a gene may be stably expressed in human lung cancer cell line A549. The expression of the introduced p16ink4a could block lung cancer cells to entry into S phase of cell cycle and inhibit tumor malignant growth both in vitro and in vivo.
基金the National Natural Science Foundation of China! (No. 39700143).
文摘Objective: To determine the effects on the cell growth, tumorigenicity and chemosensitivity of p16/CDK4I in human glioma. Methods: p16 gene was transfected into U251 cells by lipofectin. Expression of exogenous p16 gene was confirmed by immunohistochemistry and Northern blot. The effects of exogenous p16 gene on the growth and chemosensitivity to teniposide were examined. Results: Expression of exogenous p16 gene inhibited the growth dramatically in vitro. G1 arrest of tumor cells was observed. However, wt p16-positive U251 was less sensitive than control cell lines and the number of apoptotic cells after chemotherapy was reduced. Conclusion: The expression of exogenous p16 gene could inhibit the growth of glioma. On the other hand, the chemosensitivity to teniposide of p16-positive U251 was decreased.
