Pancreatic cancer is a disease with poor prognosis mainly due to low resection rates and late diagnosis. To increase resectability and improve survival rates, a better understanding of pancreatic cancer pathogenesis and more effective screening techniques are required. New methods, such as genetic and molecular alterations, may suggest novel approaches for pancreatic cancer diagnosis and treatment. We immunohistochemically investigated 44 formalin-fixed, paraffin-embedded specimens of pancreatic ductal adenocarcinoma using monoclonal anti-p16 antibodies and monoclonal anti-p53 antibodies. The expressions of p16 and p53 proteins were compared using the Chi-square test with SPSS. Disease-free survival was analyzed using the Kaplan-Meier method, verified by the Log- Rank test. Loss of p16 expression was noted in 20 (45.5%) cases and aberrant p53 protein expression was detected in 14 (31.8%) cases. Loss of p16 expression was associated with a higher incidence of lymph node metastasis (p=0.040) and a more advanced stage (p=0.015), although there was no significant correlation between p16 expression and survival. Aberrant p53 protein expression correlated with histologic grade (p= 0.038). Disease-free survival rate was significantly lower in the aberrant p53 protein positive group compared to the negative group (p=0.029). From our results, we suggest that p53 is not a prognostic factor; however, p16 and p53 genes do play important roles in the progression of pancreatic ductal adenocarcinoma.
Adult ; Aged ; Female ; Genes, p16 ; Genes, p53 ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Neoplasm Staging ; Pancreatic Neoplasms/*chemistry/genetics/mortality/pathology ; Protein p16/*analysis ; Protein p53/*analysis ; Sex Characteristics

OBJECTIVETo contrast the feature of different periods of p53, p16, and cyclin D1 expression in cells of hepatocarcinoma.

METHODSSixty hepatocarcinoma specimens were collected and divided into three periods in one yearly cycle with each period of 20 cases. Hepatocellular carcinoma was documented in all cases by pathomorphological classification. Edmondson-Steinet grading belonged to II~III stages. The expression of gene proteins of p53, p16, and cyclin D1 was determined by immunohistochemical assay (S-P method). The expressive intensity was analysed by the rank test.

RESULTSp16 expression showed significant difference of period (H=10.334, P<0. 05). Arpil-July was the high expressive period.

CONCLUSIONSp16 may take an important role in the chronobiologic mechanism of gene control of hepatocyte canceration and hepatocarcinoma cell growth.

Carcinoma, Hepatocellular ; chemistry ; Cyclin D1 ; analysis ; Cyclin-Dependent Kinase Inhibitor p16 ; analysis ; Female ; Humans ; Immunohistochemistry ; Liver Neoplasms ; chemistry ; Male ; Middle Aged ; Tumor Suppressor Protein p53 ; analysis

OBJECTIVETo investigate the relationship between alterations of p16(INK4a) and p14(ARF) genes and gastric carcinogenesis.

METHODSThe tumors and neighboring gastric tissues from 48 patients with gastric cancer were studied. The homozygous deletion, mutation, methylation of the CpG islands, and mRNA expression of p16(INK4a) and p14(ARF) genes were assessed by PCR, PCR-SSCP, PCR based methylation assay, and RT-PCR.

RESULTS(1) The homozygous deletion rate of p16(INK4a) and p14(ARF) was 35.4% (17/48), and no homozygous deletion was examined in any gastric tissue neighboring the tumor. (2) There was no point mutation of p16(INK4a) and p14(ARF) in 31 gastric cancers without homozygous deletion or in the matched gastric tissues adjacent to the tumor. (3) Methylation of the CpG islands of p16(INK4a) and p14(ARF) was detected in 47.9% (23/48) of gastric cancers, while methylation was observed only in 2 of 48 gastric tissues neighboring the cancer with a significant difference (P < 0.01). (4) The loss rate of p16(INK4a) mRNA was 47.9% (23/48) in gastric cancer, and the patients of the combined methylation of exons 1alpha and 2 had a higher loss rate (100%, 6/6) of p16(INK4a) mRNA than those of the methylation of the other exons (11.8%, 2/17, P < 0.01); the loss rate of p14(ARF) mRNA was 45.8% (22/48) in gastric cancer, and patients with the combined methylation of exons 1beta and 2 had a higher loss rate (100%, 3/3) of p14(ARF) mRNA than those of the methylation of the other exons (15%, 3/20, P < 0.05). (5) The combined loss of p16(INK4a) and p14(ARF) mRNAs was examined in 1 (5.6%) of 18 patients of well and moderately-differentiated carcinomas, and 11 (36.7%) of 30 patients of poorly and not-differentiated carcinomas with a significant difference (P < 0.05).

CONCLUSIONp16(INK4a) and p14(ARF) genes are frequently inactivated by homozygous deletion and methylation of the 5'CpG islands in gastric cancer, which may play an important role in the carcinogenesis of gastric cancer.

Adolescent ; Adult ; Aged ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; Gene Deletion ; Genes, p16 ; Humans ; Middle Aged ; RNA, Messenger ; analysis ; Stomach Neoplasms ; genetics ; Tumor Suppressor Protein p14ARF ; genetics

OBJECTIVETo investigate the relationship between alterations of p16INK4a and p14ARF genes and gastric carcinogenesis.

METHODSTumors and gastric tissues neighboring carcinoma from 48 patients with gastric cancer were studied. Homozygous deletion, mutation, methylation of the CpG islands, mRNA expression of p16INK4a and p14ARF genes were assessed by polymerase chain reaction (PCR), PCR-single strand conformation polymorphism (SSCP), PCR based methylation assay, and reverse transcription (RT)-PCR.

RESULTS(1) The overall homozygous deletion rate of p16INK4a and p14ARF was 35.4% (17/48), and no homozygous deletion was examined in all the gastric tissues neighboring tumor. (2) There was no point mutation of p16INK4a and p14ARF in 31 gastric cancers without homozygous deletion and in the matched gastric tissues adjacent to tumor. (3) Methylation of the CpG islands of p16INK4a and p14ARF was detected in 47.9% (23/48) of gastric cancers, while methylation was observed only in 2 of 48 gastric tissues neighboring cancers with a significant difference (P <0.01). (4) The rate of p16INK4a mRNA loss was 47.9% (23/48) in gastric cancer, and the cases of combined methylation of exons 1alpha and 2 had a higher loss rate (100%, 6/6) of p16INK4a mRNA than those of methylation form the other exons (11.8%, 2/17) (P <0.01). The loss rate of p14ARF mRNA was 45.8% (22/48) in gastric cancer, and patients with combined methylation of exons 1beta and 2 had a higher loss rate (100%, 3/3) of p14ARF mRNA than those of the methylation from the other exons (15%, 3/20) (P < 0.05). (5) The combined loss of p16INK4a and p14ARF mRNAs was examined in 1 (5.6%) of 18 cases of well and moderately-differentiated carcinomas, and 11 (36.7%) of 30 cases of poorly and not-differentiated carcinomas with significant difference (P <0.05).

CONCLUSIONp16INK4a and p14ARF genes were frequently inactivated by homozygous deletion and methylation of the 5' CpG islands in gastric cancer, which might have played an important role in the development of gastric cancer.

Adenocarcinoma ; genetics ; Adolescent ; Adult ; Aged ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; Female ; Gene Deletion ; Genes, p16 ; Humans ; Male ; Middle Aged ; RNA, Messenger ; analysis ; Stomach Neoplasms ; genetics ; Tumor Suppressor Protein p14ARF ; genetics

OBJECTIVETo investigate the mechanism of pilose antler polypeptides (PAP) resisting replicative senescence of rat chondrocyte serially subcultivated in vitro by means of PAP interfering and controlled experiment.

METHODSThe successive tert-generation (2nd passage, 3rd passage, 4th passage) chondrocytes and the 4th passage cells intervented by PAP were studied for senenscence mechanism. In this course, immunocytochemistry was applied for pl6, pRb, E2F, CyclinD, CDK4 and TRAP-ELISA (telomerase repeat amplification protocol assay-enzyme linked immunosorbent assay) was applied for telomerase activation to observe targets' changing regarding to senescence and the function of PAP.

RESULTSAlong with cell's replicative senescence, pl6, pRb and Cyclin D express significantly rised (P < 0.01), while E2F, CDK4 and telomerase express significantly lowerd (P < 0.01). Meanwhile, in PAP interfered group compared with which in 4th passage group, pl6, pRb and Cyclin D express significantly lowerd (P < 0.01l), while E2F, CDK4 and telomerase express significantly rised (P < 0.01).

CONCLUSIONPAP has function that it reversingly affect the express of factors which controlling cell life cycle and cell growth to postpone chondrocyte senenscence.

Animals ; Antlers ; chemistry ; Cellular Senescence ; drug effects ; Chondrocytes ; cytology ; drug effects ; Cyclin D ; Cyclin-Dependent Kinase 4 ; analysis ; Cyclin-Dependent Kinase Inhibitor p16 ; analysis ; Cyclins ; analysis ; E2F Transcription Factors ; analysis ; Peptides ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Retinoblastoma Protein ; analysis

BACKGROUND: This study investigates the prevalence of human papillomavirus (HPV) 52 and 58 genotypes among women residing in Busan, and the expression of p16 and p53 proteins in cervical neoplasia with HPV 52 and 58 infections. METHODS: A total of three hundred fifteen cases were analyzed using the HPV DNA chip test for HPV genotypes, and of these, we retrospectively examined p16 and p53 expression in 62 cases of cervical tissues infected with HPV 52 and 58 using immunohistochemistry. RESULTS: HPV 52 and 58 genotypes were identified in 62 (54.9%) out of 113 high-risk, HPV-infected cases. Of the cases examined, there were 19 single HPV 52 infections (16.8%), 23 single HPV 58 infections (20.4%), 4 multiple HPV 52 infections (3.5%), and 16 multiple HPV-58 infections (14.2%). Immunoreactivity of p16 and p53 was observed in 41 (66.1%) and 23 (37.1%) of the 62 cases of cervical neoplasia infected with HPV 52 and 58 genotypes, respectively. CONCLUSIONS: This study demonstrates a high prevalence of HPV 52 and 58 genotypes, in addition to HPV 16, among high-risk strains of cervical neoplasia in Korea. These findings suggest that development of more vaccines would be beneficial for the prevention of the various HPV genotypes.
Busan ; Cyclin-Dependent Kinase Inhibitor p16 ; Female ; Genotype* ; Human papillomavirus 16 ; Humans* ; Immunohistochemistry ; Korea ; Oligonucleotide Array Sequence Analysis ; Prevalence ; Retrospective Studies ; Tumor Suppressor Protein p53 ; Vaccines

PURPOSE: The methylation status of the CpG promoter regions of the p16INK4A and p14ARF genes, mutations of 4 exons of the CDKN2A gene, and the expression of the corresponding proteins were examined. Prognostic implications were assessed in osteosarcoma. MATERIALS AND METHODS: Methylation-specific PCR, sequence analysis, and immunohistochemical staining were performed upon 32 frozen osteosarcoma tissues. RESULTS: Methylation of p16INK4A was found in 16%, and methylation of p14ARF in 47%. Metastasis and poor survival was statistically related to the methylation of p14ARF. The methylation of p14ARF correlated with the repression of the corresponding protein, and repression of p14ARF with the repression of p21 and of wild type of p53. No sequence alterations were found in the four exons of the CDKN2A gene. Methylation of p14 showed highest hazard ratio by multivariate survival analysis. CONCLUSION: Our data suggest that methylation of the CDKN2A gene seems to be the main mechanism of protein repression. For p14ARF, the methylation of its promoter region was related to the repression of p21 and wild type p53, distant metastasis and a poor prognosis. Further study regarding cell cycle regulatory factors should shed light on oncogenesis and the possibility of a new treatment strategy for osteosarcoma.
Carcinogenesis ; Cell Cycle ; Exons ; Genes, p16 ; Methylation* ; Neoplasm Metastasis ; Osteosarcoma* ; Phosphotransferases* ; Polymerase Chain Reaction ; Prognosis* ; Promoter Regions, Genetic ; Repression, Psychology ; Sequence Analysis ; Tumor Suppressor Protein p14ARF

The ability of viral oncoproteins to subvert cell cycle checkpoints may constitute a mechanism by which viral oncoproteins induce genetic instability. HPV 16 E6 and E7 disrupt cell cycle checkpoints, particularly affecting nearly all cyclin-dependent kinase inhibitors linked to the G1- and G2- checkpoints, in each case by means of a different mechanism. HPV 16 E7 shows homology with the pRb binding sites of cyclin D1, which consequently releases E2F. In addition, E7 directly binds to p21, and releases PCNA and other S-phase promoting genes. In turn, released E2F activates cyclin E, and cyclin E accelerates p27 proteolysis as a function of the antagonistic reaction of its own inhibitor. The induction of p16 expression is assumed to be indirectly associated with E7, which is upregulated only after prolonged inactivation of Rb. HPV 16 E6 decreased the fidelity of multiple checkpoints controlling both entry into and exit from mitosis, with the mechanism of p53 inactivation. In addition, HPV 16 E6 increased the sensitivity to chemically induced S-phase premature mitosis and decreased mitotic spindle assembly checkpoint function. Alongside the impressive advances made in the understanding of the molecular mechanisms, which HPV disrupts, the validity of these conclusions should be evaluated in the diagnostic and prognostic fields.
Cervix Neoplasms/*pathology/virology ; Cyclin-Dependent Kinases/*antagonists & inhibitors ; Cyclins/analysis ; Female ; *G1 Phase ; *G2 Phase ; Human ; Microfilament Proteins/analysis ; Papillomavirus Infections/*pathology ; *Papillomavirus, Human ; Proliferating Cell Nuclear Antigen/analysis ; Protein p16/analysis ; Tumor Virus Infections/*pathology

OBJECTIVETo investigate the role of HPV16E6 and E7 during the transformation of oral epithelial cells.

METHODSAn human immortalized oral epithelial cell line (HIOEC) was established by transfecting HPV16E6, E7 open reading frames using recombinant retroviral system plxsn to human normal oral epithelial cells. Expression of HPV16E6, E7, Rb, P16 and Cycin D1 were analyzed by Western blot in HIOEC and human normal oral epithelial cells. Formation of complex of HPV16E7 and Rb were analyzed by Immunoprecipitation-western blot. Human normal oral epithelial cells and the oral epithelial cells transfected with plxsn were used as control groups.

RESULTSHIOEC expressed HPV16 E6 and E7; HIOEC expressed both hyperphosphorylated and underphosphorylated Rb while oral epithelial cells in two control groups only expressed hyperphosphorylated Rb. HPV16 E7 formed complex with underphosphorylated Rb; the level of P16 and Cyclin D1 had no remarkable change.

CONCLUSIONSHPV16E7 plays an important role in the immortalization of oral epithelial cells induced by HPV16.

Blotting, Western ; Cell Line ; Cell Transformation, Neoplastic ; Cyclin D1 ; analysis ; Cyclin-Dependent Kinase Inhibitor p16 ; analysis ; Humans ; Mouth Mucosa ; metabolism ; pathology ; virology ; Oncogene Proteins, Viral ; physiology ; Papillomavirus E7 Proteins ; Phosphorylation ; Repressor Proteins ; Retinoblastoma Protein ; analysis

Deranged expression of cell cycle modulators has been reported to contribute to the development and progression of hepatocellular carcinoma (HCC). However, their expression patterns remain poorly understood in hepatitis B virus (HBV)-related HCC, which constitutes about 65-70% of HCC in Korea. The aims of this study were to evaluate the expressions of G1-S modulators in HBV-related HCCs and dysplastic nodules (DNs), and to correlate with the histopathologic features of HCCs. Immunohistochemical expressions of cyclin D1, cyclin E, p53, p27, p21, p16, Rb, and PCNA proteins were investigated in 80 HCCs and 22 DNs. Cyclin D1 overexpression showed positive relationships with advanced tumor stage, poor differentiation, larger tumor size, microvascular invasion, intrahepatic meta-stasis, no tumor capsule formation, infiltrative growth, aberrant p53 expression, and high PCNA labeling index (LI) of HCC (p<0.05). Aberrant p53 expression showed positive relationship with poor differentiation of HCC (p<0.01). Expression of cyclin D1 or p53 was not observed in DNs. The p27 LI and p16 LI were lower in HCCs with intrahepatic metastasis (p<0.05). Cyclin D1 overexpression and aberrant p53 expression could be associated with the progression of HBV-related HCC, and might have a less crucial role in the DN-HCC sequence. In addition, elevated expression of p27 and p16 proteins might have inhibitory action to the intrahepatic metastasis of HBV-related HCC.
Adult ; Aged ; Carcinoma, Hepatocellular/chemistry/etiology/*pathology ; Cyclin D1/*analysis ; Female ; G1 Phase ; Hepatitis B/*complications ; Human ; Immunohistochemistry ; Liver Neoplasms/chemistry/etiology/*pathology ; Male ; Microfilament Proteins/analysis ; Middle Age ; Precancerous Conditions/*virology ; Proliferating Cell Nuclear Antigen/analysis ; Protein p16/analysis ; Protein p53/*analysis ; Retinoblastoma Protein/analysis ; S Phase