BACKGROUND Approximately 40%of colorectal cancer(CRC)cases are linked to Kirsten rat sarcoma viral oncogene homolog(KRAS)mutations.KRAS mutations are associated with poor CRC prognosis,especially KRAS codon 12 mutatio...BACKGROUND Approximately 40%of colorectal cancer(CRC)cases are linked to Kirsten rat sarcoma viral oncogene homolog(KRAS)mutations.KRAS mutations are associated with poor CRC prognosis,especially KRAS codon 12 mutation,which is associated with metastasis and poorer survival.However,the clinicopathological characteristics and prognosis of KRAS codon 13 mutation in CRC remain unclear.AIM To evaluate the clinicopathological characteristics and prognostic value of codonspecific KRAS mutations,especially in codon 13.METHODS This retrospective,single-center,observational cohort study included patients who underwent surgery for stage I-III CRC between January 2009 and December 2019.Patients with KRAS mutation status confirmed by molecular pathology reports were included.The relationships between clinicopathological characteristics and individual codon-specific KRAS mutations were analyzed.Survival data were analyzed to identify codon-specific KRAS mutations as recurrence-related factors using the Cox proportional hazards regression model.RESULTS Among the 2203 patients,the incidence of KRAS codons 12,13,and 61 mutations was 27.7%,9.1%,and 1.3%,respectively.Both KARS codons 12 and 13 mutations showed a tendency to be associated with clinical characteristics,but only codon 12 was associated with pathological features,such as stage of primary tumor(T stage),lymph node involvement(N stage),vascular invasion,perineural invasion,tumor size,and microsatellite instability.KRAS codon 13 mutation showed no associations(77.2%vs 85.3%,P=0.159),whereas codon 12 was associated with a lower 5-year recurrence-free survival rate(78.9%vs 75.5%,P=0.025).In multivariable analysis,along with T and N stages and vascular and perineural invasion,only codon 12(hazard ratio:1.399;95%confidence interval:1.034-1.894;P=0.030)among KRAS mutations was an independent risk factor for recurrence.CONCLUSION This study provides evidence that KRAS codon 13 mutation is less likely to serve as a prognostic biomarker than codon 12 mutation for CRC in a large-scale cohort.展开更多
Ras homolog enriched in brain(Rheb) is a small GTPase that activates mammalian target of rapamycin complex 1(mTORC1).Previous studies have shown that constitutively active Rheb can enhance the regeneration of sensory ...Ras homolog enriched in brain(Rheb) is a small GTPase that activates mammalian target of rapamycin complex 1(mTORC1).Previous studies have shown that constitutively active Rheb can enhance the regeneration of sensory axons after spinal cord injury by activating downstream effectors of mTOR.S6K1 and4E-BP1 are important downstream effectors of mTORC1.In this study,we investigated the role of Rheb/mTOR and its downstream effectors S6K1 and 4E-BP1in the protection of retinal ganglion cells.We transfected an optic nerve crush mouse model with adeno-associated viral 2-mediated constitutively active Rheb and observed the effects on retinal ganglion cell survival and axon regeneration.We found that overexpression of constitutively active Rheb promoted survival of retinal ganglion cells in the acute(14 days) and chronic(21 and 42 days) stages of injury.We also found that either co-expression of the dominant-negative S6K1mutant or the constitutively active 4E-BP1 mutant together with constitutively active Rheb markedly inhibited axon regeneration of retinal ganglion cells.This suggests that mTORC1-mediated S6K1 activation and 4E-BP1 inhibition were necessary components for constitutively active Rheb-induced axon regeneration.However,only S6K1 activation,but not 4E-BP1 knockdown,induced axon regeneration when applied alone.Furthermore,S6K1 activation promoted the survival of retinal ganglion cells at 14 days post-injury,whereas 4E-BP1 knockdown unexpectedly slightly decreased the survival of retinal ganglion cells at 14 days postinjury.Ove rexpression of constitutively active 4E-BP1 increased the survival of retinal ganglion cells at 14 days post-injury.Likewise,co-expressing constitutively active Rheb and constitutively active 4E-BP1 markedly increased the survival of retinal ganglion cells compared with overexpression of constitutively active Rheb alone at 14 days post-injury.These findings indicate that functional 4E-BP1 and S6K1 are neuroprotective and that 4E-BP1 may exert protective effects through a pathway at least partially independent of Rhe b/mTOR.Together,our results show that constitutively active Rheb promotes the survival of retinal ganglion cells and axon regeneration through modulating S6K1 and 4E-BP1 activity.Phosphorylated S6K1 and 4E-BP1 promote axon regeneration but play an antagonistic role in the survival of retinal ganglion cells.展开更多
AIM To demonstrate the relationship betweenH-ras oncogene and hepatocellular carcinoma(HCC) metastasis.METHODS Activated H-ras oncogene wastransfected into SMMC 7721, a cell line derivedfrom human HCC, by calcium phos...AIM To demonstrate the relationship betweenH-ras oncogene and hepatocellular carcinoma(HCC) metastasis.METHODS Activated H-ras oncogene wastransfected into SMMC 7721, a cell line derivedfrom human HCC, by calcium phosphatetransfection method. Some metastasis-relatedparameters were detected in vitro, includingadhesion assay, migration assay, expression ofcollagenase ⅣV (c ⅣV ase) and epidermal growthfactor receptor (EGFR).RESULTS The abilities of H-ras-transfected cellclones in adhesion to laminin (LN) or fibronectin(FN), migration, c Ⅳ ase secretion increasedmarkedly, and the expression of EGFR elevatedmoderately. More importantly, these alterationswere consistent positively with the expressionof p21, the protein product of H-ras oncogene.CONCLUSION H-ras oncogene could inducethe metastatic phenotype of HCC cell in vitro toraise its metastatic potential.展开更多
INTRODUCTIONThe point mutation rate of k-ras gene at codon 12 inpancreatic adenocarcinoma is reported to be as highas 90%,and with no mutations in normalpancreas tissues or other pancreatic disorders.Wehave detected t...INTRODUCTIONThe point mutation rate of k-ras gene at codon 12 inpancreatic adenocarcinoma is reported to be as highas 90%,and with no mutations in normalpancreas tissues or other pancreatic disorders.Wehave detected the presence of k-ras gene展开更多
AIM: To study the effect of some genes especially those involved in cell cycle regulation on hepatocellular carcinoma. METHODS: Paraffin-embedded tissue samples of 25 patients (18 males and 7 females) with hepatocellu...AIM: To study the effect of some genes especially those involved in cell cycle regulation on hepatocellular carcinoma. METHODS: Paraffin-embedded tissue samples of 25 patients (18 males and 7 females) with hepatocellular carcinoma were collected from 22 pathology centers in Tehran during 2000-2001, and stained using immunohistochemistry method (avidin-biotin-peroxidase) for detection of p53, cyclinD1, RB1, c-fos and N-ras proteins. RESULTS: Six (24%), 5 (20%), 12 (48%) and 2 samples (8%) were positive for p53, cyclinD1, C-fos and N-ras expression, respectively. Twenty-two (88%) samples had alterations in the G1 cell-cycle checkpoint protein expression (RB1 or cyclinD1). P53 positive samples showed a higher (9 times) risk of being positive for RB1 protein than p53 negative samples. Loss of expression of RB1 in association with p53 over-expression was observed in 4 (66.7%) of 6 samples. Loss of expression of RB1 was seen in all cyclinD1 positive, 20 (90.9%) N-ras negative, and 11 (50%) C-fos positive samples, respectively. CyclinD1 positive samples showed a higher (2.85 and 4.75 times) risk of being positive for c-fos and N-ras expression than cyclinD1 negative samples. CONCLUSION: The expression of p53, RB1 and c-fos genes appears to have a key role in the pathogenesis of hepatocellular carcinoma in Iran. Simultaneous overexpression of these genes is significantly associated with their loss of expression during development of hepatocellular carcinoma.展开更多
Abnormality of ras gene family was studied in a total of 206 cases of gastric cancer and precancerous lesions by PCR-RFLP, PCR-SSCP and DNA sequencing. The results showed that mutation rate of H-ras 12 codon in metapl...Abnormality of ras gene family was studied in a total of 206 cases of gastric cancer and precancerous lesions by PCR-RFLP, PCR-SSCP and DNA sequencing. The results showed that mutation rate of H-ras 12 codon in metaplasia,atypical hyperplasia, early-stage cancer and advanced cancer was 16. 7%, 31. 2 %, 50. 0%, and 32. 2%, respectively. In the groups of superficial gastritis and normal controls, no mutation were detected in codon 12 of ras. Mutations of Hras 61 codon and N-ras 12 codon in various groups were the same as those in normal control. K-ras 12 codon mutation was detected in only 2 cases of gastric cancer by using PCR-SSCP, but it was not detected by DNA sequencing, which may be polymorphism. All H-ras 12 codon mutations were G→T mutation. There were significant difference between the groups of metaplasia, dysplasia, gastric carcinoma and normal control group (P<0.05, P<0.01, P<0.01,respectively). It was concluded that H-ras 12 codon mutation was an early event and may play an important role in gastric carcinogenesis. Although K-ras, N-ras mutation rates are high in colon cancer and leukemia, it seems to bear no relationship with gastric cancer.展开更多
AIM To study the genetic alteration in ACF andto define the possibility that ACF may be a veryearly morphological lesion with molecularchanges, and to explore the relationshipbetween ACF and colorectal adenoma evencar...AIM To study the genetic alteration in ACF andto define the possibility that ACF may be a veryearly morphological lesion with molecularchanges, and to explore the relationshipbetween ACF and colorectal adenoma evencarcinoma.METHODS DNA from 35 CRC, 15 adenomas, 34ACF and 10 normal mucus was isolated by meansof microdissection. Direct gene sequencing of K-ras gene including codon 12, 13 and 61 as well asthe mutation cluster region (MCR) of APC genewas performed.RESULTS K-ras gene mutation frequency inACF, adenoma and carcinoma was 17.6% (6/34), 13.3% (2/ 15), and 14.3% (5/ 35)respectively, showing no difference ( P > 0.05)in K-fas gene mutation among three pathologicprocedures. The K-ras gene mutation inadenoma, carcinoma and 4 ACF restricted incodon 12 (GGT→GAT), but the other 2 mutationsfrom ACF located in codon 13 (GGC→GAC). K-res gene mutation was found more frequently inolder patients and patients with polypoidcancer. No mutation in codon 61 was found in thethree tissue types. Mutation rate of APO gene inadenoma and carcinoma was 22.9% (8/35) and26.7% (4/ 15), which was higher than ACF(2.9%) (P < 0.05). APC gene mutation incarcinoma was not correlated with age ofpatients, location, size and differentiation oftumor.CONCLUSION ACF might be a very earlymorphological lesion in the tumorogenesis ofcolorectal tumor. The morphological feature andgene mutation status was different in ACF andadenoma. ACF is possibly putative'microadenoma' that might be the precursor ofadenoma. In addition, the development of asubgroup of colorectal carcinomas mightundergo a way of 'normal epithelium→ ACF→carcinomas'.展开更多
AIM: To identify whether the polymorphisms of the Nacetyltransferase (NAT) genes are susceptible to primary liver cancer (PLC) in Luoyang, a PLC low-incidence area of China.METHODS: The NAT1 and NAT2 genotypes of 96 P...AIM: To identify whether the polymorphisms of the Nacetyltransferase (NAT) genes are susceptible to primary liver cancer (PLC) in Luoyang, a PLC low-incidence area of China.METHODS: The NAT1 and NAT2 genotypes of 96 PLC cases and 173 controls were determined by PCR-RFLP.Both interaction between NAT1 or NAT2 and environmental risk factors were analyzed based on case control study.RESULTS: Compared to the control group, the frequencies of alleles NAT1*3, NAT1*4, NAT1*10, NAT1*14B and alleles NAT2*4, NAT2*6, NAT2*7 in PLC group showed no statistically significant difference (x2 = 2.61 and 4.16,respectively, both P>0.05). The frequencies of NAT1 genotypes NAT1*3/*3, NAT1*3/*4, NAT1*3/*10,NAT1*3/*14B, NAT1*4/*4, NAT1*4/*10, NAT1*4/*14B,NAT1*10/*10, NAT1*10/*14B, and NAT2 genotypes NAT2*4/*4, NAT2*4/*6, NAT2*4/*7, NAT2*6/*6,NAT2*6/*7 and NAT2*7/*7 also had no statistically significant difference between the two groups (x2 = 11.86 and 2.94respectively both, P>0.05). Neither the frequencies of rapid and slow NAT1 acetylators nor the frequencies of rapid and slow NAT2 acetylators were significantly different between the two groups (x2 = 0.598 and 0.44,respectively, both P>0.05). The interaction betweenNAT1*10 and occupational exposures was found significant with an odds ratio of 3.40 (x2 = 8.42, P = 0.004,OR 95%CI:1.03-11.22). But no interaction was found between NAT2 and any environmental risk factors.CONCLUSION: The polymorphisms of NAT1 and NAT2are not susceptible to PLC in Luoyang. Allele NAT1*10interacts with occupational exposures.展开更多
AIM: To investigate the frequency and distribution of N-acetyltransferase 2 (NAT2) and uridine 5′-diphosphate (UDP)-glucuronosyltransferase 1A7 (UGT1A7) genes in patients with ulcerative colitis (UC) and Crohn's ...AIM: To investigate the frequency and distribution of N-acetyltransferase 2 (NAT2) and uridine 5′-diphosphate (UDP)-glucuronosyltransferase 1A7 (UGT1A7) genes in patients with ulcerative colitis (UC) and Crohn's disease (CD).METHODS: Frequencies and distributions of NAT2 and UGT1A7SNPs as well as their haplotypes were investigated in 95 patients with UC, 60 patients with CD, and 200gender-matched, unrelated, healthy, control volunteers by PCR-restriction fragment length polymorphism (RFLP),PCR-denaturing high-performance liquid chromatography (DHPLC), and direct DNA sequencing.RESULTS: Multiple logistic regression analysis revealed that the frequency of haplotype, NAT2*7B, significantly increased in CD patients, compared to that in controls (P= 0.0130, OR = 2.802, 95%CI = 1.243-6.316). However,there was no association between NAT2 haplotypes and UC, or between any UGT1A7haplotypes and inflammatory bowel disease (IBD).CONCLUSION: It is likely that the NAT2 gene is one of the determinants for CD in Japanese. Alternatively, a new CD determinant may exist in the 8p22 region, whereNAT2is located.展开更多
Summary: The specific anti-tumor immune response induced by mouse bone marrow dendritic cells (DCs) lransfected with recombinant adenovirus carrying mutant k-ras genes was investighted. DCs were generated from mous...Summary: The specific anti-tumor immune response induced by mouse bone marrow dendritic cells (DCs) lransfected with recombinant adenovirus carrying mutant k-ras genes was investighted. DCs were generated from mouse bone marrow in the presence of rmGM-CSF (3.3 ng/mL) and rmIL-4 (1.3 ng/mL) and detected by FACS, and then transfecled with the recombinant adenovirus encoding mutant k ras gene. The efficacy of transfection and T cell stimulating activity of DCs were detected. CTL activity of the mice vaccinated with DCs was observed. The resuhs showed thai DCs had dendritic veiled morphology. BmDCs highly expressed B7-1(80%), B7-2(77%), MHC Ⅱ (70%), CDllc (65%), CD40 (70%) and CD54 (96%) with FACS, and no significant difference in the expression was observed before and after the transfection (P〈0.05). The DCs transfeeled by mutant k-ras gene could significantly stimulate lymphoeytes proliferation as compared with those transfeeted by Ad e or non-modified DCs (P〈0.05). DC vaccine transfected by mutant k-ras gene could induce CTL activity against Lewis lung cancer, but not against B16. The specific eytotoxicity against Lewis lung cancer in Ad-k-ras/12-transdueed DC group was signifieantly higher than those in the control, vector and non transfeeted DCs groups (P〈0.05). It was concluded that special antitumor response could be induced by DCs transfected with recombinant adenovirus carrying mutant k-ras genes.展开更多
Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well ...Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well studied in bacteria. N6-adenine-specific DNA methyltransferase(N6AMT) is the main enzyme catalyzing the methylation of the adenine base and knowledge of this enzyme was mainly derived from work in prokaryotic models. However, large-scale gene discovery at the genome level in many model organisms indicated that the N6AMT gene also exists in eukaryotes, such as humans, mice, fruit flies and plants. Here, we cloned a N6AMT gene from Nilaparvata lugens(Nlu-N6AMT) and amplified its fulllength transcript. Then, we carried out a systematic investigation of N6AMT in 33 publically available insect genomes, indicating that all studied insects had N6AMT. Genomic structure analysis showed that insect N6AMT has short introns compared with the mammalian homologs. Domain and phylogenetic analysis indicated that insect N6AMT had a conserved N6-adenine Mlase domain that is specific to catalyze the adenine methylation. Nlu-N6AMT was highly expressed in the adult female. We knocked down Nlu-N6AMT by feeding ds RNA from the second instar nymph to adult female, inducing retard development of adult female. In all, we provide the first genome-wide analysis of N6AMT in insects and presented the experimental evidence that N6AMT might have important functions in reproductive development and ovary maturation.展开更多
To evaluate the feasibility and clinical significance of the PCR SSP technique in detecting K ras gene mutation at codon 12 in pancreatic adenocarcinoma tissues. 80 specimens of surgical resection or biopsy samples ...To evaluate the feasibility and clinical significance of the PCR SSP technique in detecting K ras gene mutation at codon 12 in pancreatic adenocarcinoma tissues. 80 specimens of surgical resection or biopsy samples were tested at our hospital from January 1994 to September 1995. Three different special sequence primers (SSP) synthesized according to mutation styles of CGT, GTT, GAT were respectively prepared. Three amplification reactions were performed for each sample. The amplification products were analyzed by conventional polyacrylamide gel electrophoresis, stained with ethidium bromide and observed under UV transillumination. Results: All of the 34 pancreatic adenocarcinoma samples had positive PCR results with the mutation rate 100%. 7 cases were CGT mutation, 18 GGT and 17 GAT mutation, in which 2 types of mutation existed in 8 cases. No mutation appeared in 13 normal pancreatic tissues, 6 insulinomas, 6 chronic pancreatitis, 5 benign pancreatic cysts, 7 bile duct carcinoma, 5 ampulla carcinoma and 4 carcinomas of duodenal papilla. Conclusion: Pancreatic adenocarcinoma is one of the commonly encounted tumors and is still very difficult to diagnose at the early stage and to distinguish from other lesions preoperatively. Our study indicates that PCR SSP is an ideal assay in comparison with other methods to detect K ras gene mutation. It is simple, rapid, specific, sensitive and easily generalized for clinical application on preoperative diagnosis.展开更多
Objective:To study the role of the mutations of p53, APC and K-ras genes in 47 cases of 3 types of intestinal metaplasia (IM) of gastric mucosa. Methods:In 47 cases of IM, exons 5- 8 of p53 and exons 15 of APC were ex...Objective:To study the role of the mutations of p53, APC and K-ras genes in 47 cases of 3 types of intestinal metaplasia (IM) of gastric mucosa. Methods:In 47 cases of IM, exons 5- 8 of p53 and exons 15 of APC were examined with PCR-SSCP and codon 12 of K-ras with PCR-RFLP to detect the existence of any mutations of these structures. Results:Muta- tions of p53, APC and K-ras were found in 29.8% (14/47),6.4% (3/47) and 6.4% (3/47) respectively in our series of patients who consisted of 33 with types I and II and 14 with type III of IM. The mutation rate of p53 was far higher in patients with type III IM (57.1%,8/14) than in those with types I and II IM(18.2%,6/33)(P <0.05). Though the mutation rate of APC and K-ras was also higher in the patients with type III IM than in those with types I and II IM, it was of no statistical significance (P >0.05). In one case of type III IM, mutation of both p53 and K-ras was found. Conclusion: The molecular changes of 3 types of IM are different. The mutation of p53 may be closely related to carcinogenesis in cases of type III IM and it serve as a sign for the early diagnosis of gastric carcinoma.展开更多
文摘BACKGROUND Approximately 40%of colorectal cancer(CRC)cases are linked to Kirsten rat sarcoma viral oncogene homolog(KRAS)mutations.KRAS mutations are associated with poor CRC prognosis,especially KRAS codon 12 mutation,which is associated with metastasis and poorer survival.However,the clinicopathological characteristics and prognosis of KRAS codon 13 mutation in CRC remain unclear.AIM To evaluate the clinicopathological characteristics and prognostic value of codonspecific KRAS mutations,especially in codon 13.METHODS This retrospective,single-center,observational cohort study included patients who underwent surgery for stage I-III CRC between January 2009 and December 2019.Patients with KRAS mutation status confirmed by molecular pathology reports were included.The relationships between clinicopathological characteristics and individual codon-specific KRAS mutations were analyzed.Survival data were analyzed to identify codon-specific KRAS mutations as recurrence-related factors using the Cox proportional hazards regression model.RESULTS Among the 2203 patients,the incidence of KRAS codons 12,13,and 61 mutations was 27.7%,9.1%,and 1.3%,respectively.Both KARS codons 12 and 13 mutations showed a tendency to be associated with clinical characteristics,but only codon 12 was associated with pathological features,such as stage of primary tumor(T stage),lymph node involvement(N stage),vascular invasion,perineural invasion,tumor size,and microsatellite instability.KRAS codon 13 mutation showed no associations(77.2%vs 85.3%,P=0.159),whereas codon 12 was associated with a lower 5-year recurrence-free survival rate(78.9%vs 75.5%,P=0.025).In multivariable analysis,along with T and N stages and vascular and perineural invasion,only codon 12(hazard ratio:1.399;95%confidence interval:1.034-1.894;P=0.030)among KRAS mutations was an independent risk factor for recurrence.CONCLUSION This study provides evidence that KRAS codon 13 mutation is less likely to serve as a prognostic biomarker than codon 12 mutation for CRC in a large-scale cohort.
基金National Natural Science Foundation of China,Nos.82070967,81770930the Natural Science Foundation of Hunan Province,No.2020jj4788 (all to BJ)。
文摘Ras homolog enriched in brain(Rheb) is a small GTPase that activates mammalian target of rapamycin complex 1(mTORC1).Previous studies have shown that constitutively active Rheb can enhance the regeneration of sensory axons after spinal cord injury by activating downstream effectors of mTOR.S6K1 and4E-BP1 are important downstream effectors of mTORC1.In this study,we investigated the role of Rheb/mTOR and its downstream effectors S6K1 and 4E-BP1in the protection of retinal ganglion cells.We transfected an optic nerve crush mouse model with adeno-associated viral 2-mediated constitutively active Rheb and observed the effects on retinal ganglion cell survival and axon regeneration.We found that overexpression of constitutively active Rheb promoted survival of retinal ganglion cells in the acute(14 days) and chronic(21 and 42 days) stages of injury.We also found that either co-expression of the dominant-negative S6K1mutant or the constitutively active 4E-BP1 mutant together with constitutively active Rheb markedly inhibited axon regeneration of retinal ganglion cells.This suggests that mTORC1-mediated S6K1 activation and 4E-BP1 inhibition were necessary components for constitutively active Rheb-induced axon regeneration.However,only S6K1 activation,but not 4E-BP1 knockdown,induced axon regeneration when applied alone.Furthermore,S6K1 activation promoted the survival of retinal ganglion cells at 14 days post-injury,whereas 4E-BP1 knockdown unexpectedly slightly decreased the survival of retinal ganglion cells at 14 days postinjury.Ove rexpression of constitutively active 4E-BP1 increased the survival of retinal ganglion cells at 14 days post-injury.Likewise,co-expressing constitutively active Rheb and constitutively active 4E-BP1 markedly increased the survival of retinal ganglion cells compared with overexpression of constitutively active Rheb alone at 14 days post-injury.These findings indicate that functional 4E-BP1 and S6K1 are neuroprotective and that 4E-BP1 may exert protective effects through a pathway at least partially independent of Rhe b/mTOR.Together,our results show that constitutively active Rheb promotes the survival of retinal ganglion cells and axon regeneration through modulating S6K1 and 4E-BP1 activity.Phosphorylated S6K1 and 4E-BP1 promote axon regeneration but play an antagonistic role in the survival of retinal ganglion cells.
文摘AIM To demonstrate the relationship betweenH-ras oncogene and hepatocellular carcinoma(HCC) metastasis.METHODS Activated H-ras oncogene wastransfected into SMMC 7721, a cell line derivedfrom human HCC, by calcium phosphatetransfection method. Some metastasis-relatedparameters were detected in vitro, includingadhesion assay, migration assay, expression ofcollagenase ⅣV (c ⅣV ase) and epidermal growthfactor receptor (EGFR).RESULTS The abilities of H-ras-transfected cellclones in adhesion to laminin (LN) or fibronectin(FN), migration, c Ⅳ ase secretion increasedmarkedly, and the expression of EGFR elevatedmoderately. More importantly, these alterationswere consistent positively with the expressionof p21, the protein product of H-ras oncogene.CONCLUSION H-ras oncogene could inducethe metastatic phenotype of HCC cell in vitro toraise its metastatic potential.
基金Natural Science Foundation of Jiangsu Scientific Committee
文摘INTRODUCTIONThe point mutation rate of k-ras gene at codon 12 inpancreatic adenocarcinoma is reported to be as highas 90%,and with no mutations in normalpancreas tissues or other pancreatic disorders.Wehave detected the presence of k-ras gene
文摘AIM: To study the effect of some genes especially those involved in cell cycle regulation on hepatocellular carcinoma. METHODS: Paraffin-embedded tissue samples of 25 patients (18 males and 7 females) with hepatocellular carcinoma were collected from 22 pathology centers in Tehran during 2000-2001, and stained using immunohistochemistry method (avidin-biotin-peroxidase) for detection of p53, cyclinD1, RB1, c-fos and N-ras proteins. RESULTS: Six (24%), 5 (20%), 12 (48%) and 2 samples (8%) were positive for p53, cyclinD1, C-fos and N-ras expression, respectively. Twenty-two (88%) samples had alterations in the G1 cell-cycle checkpoint protein expression (RB1 or cyclinD1). P53 positive samples showed a higher (9 times) risk of being positive for RB1 protein than p53 negative samples. Loss of expression of RB1 in association with p53 over-expression was observed in 4 (66.7%) of 6 samples. Loss of expression of RB1 was seen in all cyclinD1 positive, 20 (90.9%) N-ras negative, and 11 (50%) C-fos positive samples, respectively. CyclinD1 positive samples showed a higher (2.85 and 4.75 times) risk of being positive for c-fos and N-ras expression than cyclinD1 negative samples. CONCLUSION: The expression of p53, RB1 and c-fos genes appears to have a key role in the pathogenesis of hepatocellular carcinoma in Iran. Simultaneous overexpression of these genes is significantly associated with their loss of expression during development of hepatocellular carcinoma.
文摘Abnormality of ras gene family was studied in a total of 206 cases of gastric cancer and precancerous lesions by PCR-RFLP, PCR-SSCP and DNA sequencing. The results showed that mutation rate of H-ras 12 codon in metaplasia,atypical hyperplasia, early-stage cancer and advanced cancer was 16. 7%, 31. 2 %, 50. 0%, and 32. 2%, respectively. In the groups of superficial gastritis and normal controls, no mutation were detected in codon 12 of ras. Mutations of Hras 61 codon and N-ras 12 codon in various groups were the same as those in normal control. K-ras 12 codon mutation was detected in only 2 cases of gastric cancer by using PCR-SSCP, but it was not detected by DNA sequencing, which may be polymorphism. All H-ras 12 codon mutations were G→T mutation. There were significant difference between the groups of metaplasia, dysplasia, gastric carcinoma and normal control group (P<0.05, P<0.01, P<0.01,respectively). It was concluded that H-ras 12 codon mutation was an early event and may play an important role in gastric carcinogenesis. Although K-ras, N-ras mutation rates are high in colon cancer and leukemia, it seems to bear no relationship with gastric cancer.
基金This subject is supported by the Fund for Returned Scientists and Scholars,[1999]363.Chinese Ministry of Education.
文摘AIM To study the genetic alteration in ACF andto define the possibility that ACF may be a veryearly morphological lesion with molecularchanges, and to explore the relationshipbetween ACF and colorectal adenoma evencarcinoma.METHODS DNA from 35 CRC, 15 adenomas, 34ACF and 10 normal mucus was isolated by meansof microdissection. Direct gene sequencing of K-ras gene including codon 12, 13 and 61 as well asthe mutation cluster region (MCR) of APC genewas performed.RESULTS K-ras gene mutation frequency inACF, adenoma and carcinoma was 17.6% (6/34), 13.3% (2/ 15), and 14.3% (5/ 35)respectively, showing no difference ( P > 0.05)in K-fas gene mutation among three pathologicprocedures. The K-ras gene mutation inadenoma, carcinoma and 4 ACF restricted incodon 12 (GGT→GAT), but the other 2 mutationsfrom ACF located in codon 13 (GGC→GAC). K-res gene mutation was found more frequently inolder patients and patients with polypoidcancer. No mutation in codon 61 was found in thethree tissue types. Mutation rate of APO gene inadenoma and carcinoma was 22.9% (8/35) and26.7% (4/ 15), which was higher than ACF(2.9%) (P < 0.05). APC gene mutation incarcinoma was not correlated with age ofpatients, location, size and differentiation oftumor.CONCLUSION ACF might be a very earlymorphological lesion in the tumorogenesis ofcolorectal tumor. The morphological feature andgene mutation status was different in ACF andadenoma. ACF is possibly putative'microadenoma' that might be the precursor ofadenoma. In addition, the development of asubgroup of colorectal carcinomas mightundergo a way of 'normal epithelium→ ACF→carcinomas'.
基金Supported by the National Natural Science Foundation of China,No. 39870654
文摘AIM: To identify whether the polymorphisms of the Nacetyltransferase (NAT) genes are susceptible to primary liver cancer (PLC) in Luoyang, a PLC low-incidence area of China.METHODS: The NAT1 and NAT2 genotypes of 96 PLC cases and 173 controls were determined by PCR-RFLP.Both interaction between NAT1 or NAT2 and environmental risk factors were analyzed based on case control study.RESULTS: Compared to the control group, the frequencies of alleles NAT1*3, NAT1*4, NAT1*10, NAT1*14B and alleles NAT2*4, NAT2*6, NAT2*7 in PLC group showed no statistically significant difference (x2 = 2.61 and 4.16,respectively, both P>0.05). The frequencies of NAT1 genotypes NAT1*3/*3, NAT1*3/*4, NAT1*3/*10,NAT1*3/*14B, NAT1*4/*4, NAT1*4/*10, NAT1*4/*14B,NAT1*10/*10, NAT1*10/*14B, and NAT2 genotypes NAT2*4/*4, NAT2*4/*6, NAT2*4/*7, NAT2*6/*6,NAT2*6/*7 and NAT2*7/*7 also had no statistically significant difference between the two groups (x2 = 11.86 and 2.94respectively both, P>0.05). Neither the frequencies of rapid and slow NAT1 acetylators nor the frequencies of rapid and slow NAT2 acetylators were significantly different between the two groups (x2 = 0.598 and 0.44,respectively, both P>0.05). The interaction betweenNAT1*10 and occupational exposures was found significant with an odds ratio of 3.40 (x2 = 8.42, P = 0.004,OR 95%CI:1.03-11.22). But no interaction was found between NAT2 and any environmental risk factors.CONCLUSION: The polymorphisms of NAT1 and NAT2are not susceptible to PLC in Luoyang. Allele NAT1*10interacts with occupational exposures.
文摘AIM: To investigate the frequency and distribution of N-acetyltransferase 2 (NAT2) and uridine 5′-diphosphate (UDP)-glucuronosyltransferase 1A7 (UGT1A7) genes in patients with ulcerative colitis (UC) and Crohn's disease (CD).METHODS: Frequencies and distributions of NAT2 and UGT1A7SNPs as well as their haplotypes were investigated in 95 patients with UC, 60 patients with CD, and 200gender-matched, unrelated, healthy, control volunteers by PCR-restriction fragment length polymorphism (RFLP),PCR-denaturing high-performance liquid chromatography (DHPLC), and direct DNA sequencing.RESULTS: Multiple logistic regression analysis revealed that the frequency of haplotype, NAT2*7B, significantly increased in CD patients, compared to that in controls (P= 0.0130, OR = 2.802, 95%CI = 1.243-6.316). However,there was no association between NAT2 haplotypes and UC, or between any UGT1A7haplotypes and inflammatory bowel disease (IBD).CONCLUSION: It is likely that the NAT2 gene is one of the determinants for CD in Japanese. Alternatively, a new CD determinant may exist in the 8p22 region, whereNAT2is located.
文摘Summary: The specific anti-tumor immune response induced by mouse bone marrow dendritic cells (DCs) lransfected with recombinant adenovirus carrying mutant k-ras genes was investighted. DCs were generated from mouse bone marrow in the presence of rmGM-CSF (3.3 ng/mL) and rmIL-4 (1.3 ng/mL) and detected by FACS, and then transfecled with the recombinant adenovirus encoding mutant k ras gene. The efficacy of transfection and T cell stimulating activity of DCs were detected. CTL activity of the mice vaccinated with DCs was observed. The resuhs showed thai DCs had dendritic veiled morphology. BmDCs highly expressed B7-1(80%), B7-2(77%), MHC Ⅱ (70%), CDllc (65%), CD40 (70%) and CD54 (96%) with FACS, and no significant difference in the expression was observed before and after the transfection (P〈0.05). The DCs transfeeled by mutant k-ras gene could significantly stimulate lymphoeytes proliferation as compared with those transfeeted by Ad e or non-modified DCs (P〈0.05). DC vaccine transfected by mutant k-ras gene could induce CTL activity against Lewis lung cancer, but not against B16. The specific eytotoxicity against Lewis lung cancer in Ad-k-ras/12-transdueed DC group was signifieantly higher than those in the control, vector and non transfeeted DCs groups (P〈0.05). It was concluded that special antitumor response could be induced by DCs transfected with recombinant adenovirus carrying mutant k-ras genes.
基金supported by the National Basic Research Program of China (2012CB114102)
文摘Methylation of the N6 position of adenine, termed N6-methyladenine, protects DNA from restriction endonucleases via the host-specific restriction-modification system. N6-methyladenine was discovered and has been well studied in bacteria. N6-adenine-specific DNA methyltransferase(N6AMT) is the main enzyme catalyzing the methylation of the adenine base and knowledge of this enzyme was mainly derived from work in prokaryotic models. However, large-scale gene discovery at the genome level in many model organisms indicated that the N6AMT gene also exists in eukaryotes, such as humans, mice, fruit flies and plants. Here, we cloned a N6AMT gene from Nilaparvata lugens(Nlu-N6AMT) and amplified its fulllength transcript. Then, we carried out a systematic investigation of N6AMT in 33 publically available insect genomes, indicating that all studied insects had N6AMT. Genomic structure analysis showed that insect N6AMT has short introns compared with the mammalian homologs. Domain and phylogenetic analysis indicated that insect N6AMT had a conserved N6-adenine Mlase domain that is specific to catalyze the adenine methylation. Nlu-N6AMT was highly expressed in the adult female. We knocked down Nlu-N6AMT by feeding ds RNA from the second instar nymph to adult female, inducing retard development of adult female. In all, we provide the first genome-wide analysis of N6AMT in insects and presented the experimental evidence that N6AMT might have important functions in reproductive development and ovary maturation.
文摘To evaluate the feasibility and clinical significance of the PCR SSP technique in detecting K ras gene mutation at codon 12 in pancreatic adenocarcinoma tissues. 80 specimens of surgical resection or biopsy samples were tested at our hospital from January 1994 to September 1995. Three different special sequence primers (SSP) synthesized according to mutation styles of CGT, GTT, GAT were respectively prepared. Three amplification reactions were performed for each sample. The amplification products were analyzed by conventional polyacrylamide gel electrophoresis, stained with ethidium bromide and observed under UV transillumination. Results: All of the 34 pancreatic adenocarcinoma samples had positive PCR results with the mutation rate 100%. 7 cases were CGT mutation, 18 GGT and 17 GAT mutation, in which 2 types of mutation existed in 8 cases. No mutation appeared in 13 normal pancreatic tissues, 6 insulinomas, 6 chronic pancreatitis, 5 benign pancreatic cysts, 7 bile duct carcinoma, 5 ampulla carcinoma and 4 carcinomas of duodenal papilla. Conclusion: Pancreatic adenocarcinoma is one of the commonly encounted tumors and is still very difficult to diagnose at the early stage and to distinguish from other lesions preoperatively. Our study indicates that PCR SSP is an ideal assay in comparison with other methods to detect K ras gene mutation. It is simple, rapid, specific, sensitive and easily generalized for clinical application on preoperative diagnosis.
基金National Natural Science foundation of China, No.39470332
文摘Objective:To study the role of the mutations of p53, APC and K-ras genes in 47 cases of 3 types of intestinal metaplasia (IM) of gastric mucosa. Methods:In 47 cases of IM, exons 5- 8 of p53 and exons 15 of APC were examined with PCR-SSCP and codon 12 of K-ras with PCR-RFLP to detect the existence of any mutations of these structures. Results:Muta- tions of p53, APC and K-ras were found in 29.8% (14/47),6.4% (3/47) and 6.4% (3/47) respectively in our series of patients who consisted of 33 with types I and II and 14 with type III of IM. The mutation rate of p53 was far higher in patients with type III IM (57.1%,8/14) than in those with types I and II IM(18.2%,6/33)(P <0.05). Though the mutation rate of APC and K-ras was also higher in the patients with type III IM than in those with types I and II IM, it was of no statistical significance (P >0.05). In one case of type III IM, mutation of both p53 and K-ras was found. Conclusion: The molecular changes of 3 types of IM are different. The mutation of p53 may be closely related to carcinogenesis in cases of type III IM and it serve as a sign for the early diagnosis of gastric carcinoma.