Cdkn2a Tracer Mice: A Novel Model to Study Cellular Senescence in Post-traumatic Osteoarthritis / 中国生物化学与分子生物学报

Osteoarthritis (OA) is one of the most common geriatric motor system diseases in the clinical practice. Aging, whose hallmark is cellular senescence, is an important factor leading to the occurrence and development of OA, but its exact role in the pathological development of OA is not completely clear. Studies have proved that targeting senescence can effectively treat aging-related diseases. In this study, the heterozygous mouse model of Cdkn2a-e (Luc-2A-tdTomato-2a-CreerT2-Wpre-PA)1 was established by the CRISPR/ Cas9 technique. The expression of Cdkn2a (p16, p16INK4a), a classical marker of senescence, can be traced in vivo in mice. We then utilized anterior cruciate ligament transection (ACLT) to induce OA in Cdkn2a mice, and we hope to verify the relationship between aging and the occurrence and development of OA and visualize aging changes during OA development through the mice model. In this study, 10-12 weeks old Cdkn2a mice were randomly divided into no surgery control group, sham operation group and ACLT group. OA model was constructed in mice by ACLT operation. After surgery for 4 weeks, animals were collected for fluorescence imaging detection in vivo, which showed that local fluorescence expression of Cdkn2a increased in knee joints of mice in the ACLT group four weeks after surgery (P < 0. 05) . The Safranin O-Fast Green Staining of mouse knee tissue sections showed degeneration of the knee cartilage in the ACLT group at 4 weeks after surgery (P < 0. 05) . Immunohistochemical staining of Cdkn2a was performed on the knee tissue of mice. Compared with the other two groups, Cdkn2a staining on the cartilage surface of the knee tissue of mice in the ACLT group was deeper. The results showed that the OA model induced by surgery showed local aging, which further verified the relationship between aging and OA. At the same time, the Cdkn2a tracer mouse model can reflect the aging progress of mice in vivo, with combination of imaging examinations, so that the occurrence and progress of the relationship between aging and OA can be observed in real time. This heterozygous mouse model of Cdkn2a-e(Luc-2A-tdTomato-2a-CreerT2-Wpre-PA) 1 is not only useful for mechanism research of aging and OA diseases, but also beneficial for finding more potential therapy targets to OA and aging.

Correlation between methylation level of and genes and aging in healthy individuals / 南方医科大学学报

OBJECTIVE@#To analyze the relationship between and gene methylation with aging in the general population.@*METHODS@#We collected peripheral blood samples from 284 male and 246 female healthy subjects for detection of methylation levels of and genes using quantitative methylation-specific PCR (qMSP). The relationship between the methylation levels of and genes and aging was analyzed using Spearman or Pearson correlation test.@*RESULTS@#We found a significant positive correlation between the methylation levels of the two genes in these subjects ( < 0.05). In the overall population as well in the female subjects, methylation was found to be inversely correlated with age ( < 0.05). The methylation levels of and genes were inversely correlated with TG, ApoE, Lp(a) and AST in the overall population ( < 0.05). In both the female and male subjects, the methylation levels of the two genes were inversely correlated with Lp(a) ( < 0.05). In the male subjects, methylation was inversely correlated with AST ( < 0.05), while methylation was inversely correlated with HDL and ApoE ( < 0.05). In the female subjects, methylation was positively correlated with LDL and inversely correlated with ApoE and AST ( < 0.05).@*CONCLUSIONS@#The methylation levels of and are closely related to age and the levels of multiple proteins in healthy subjects.

Prognostic value of CDKN2A mRNA level in glioblastoma / 肿瘤研究与临床

Objective To investigate the prognostic value of CDKN2A mRNA in glioblastoma (GBM).Methods CDKN2A gene mRNA data were obtained from three different GBM database online (TCGA,REMBARNDT and GSE16011).The correlations between overall survival (OS) and CDKN2A expression were analyzed by Kaplan-Meier method and multivariate Cox regression analysis.Results In the TCGA database (n =358),patients with high CDKN2A mRNA level got longer OS than those with low expression level [median OS:18.0 months (95 ％ CI 15.0-21.0 months) vs 13.9 months (95 ％ CI 12.4-15.4 months),P =0.001].In another two validation datasets,patients with high CDKN2A mRNA level had longer OS than those with low expression level [median OS in REMBRANT:16.6 months (95 ％ CI 13.3-19.8 months) vs 11.8 months (95 ％ CI 7.3-16.4 months),P =0.019; in GSE16011:11.9 months (95 ％ CI 8.3-15.6 months) vs 8.4 months (95 ％ CI 6.2-10.5 months),P =0.005].CDKN2A mRNA level was an independent prognostic factor for GBM.The combination of CDKN2A mRNA expression with MGMT promoter methylation status or G-CIMP status/IDH1 mutations provided an optimized prognostic factor in GBM patients.Conclusion The CDKN2A mRNA has prognostic value in GBM patients,which provided an optimized stratification strategy based on multiple biomarkers.

Genética molecular del melanoma familiar: revisión del tema y aplicación clínica / Genetics of familial cutaneous melanoma

El melanoma maligno cutáneo (MMC) representa el 4 por ciento del total de los tumores malignos de piel, dando cuenta del 80 por ciento de las muertes producidas por cáncer cutáneo. La sobrevida a cinco años de individuos portadores de enfermedad metastásica se estima en 14 por ciento. Las formas de presentación incluyen una variante esporádica y otra familiar o hereditaria. En ambas el papel de diferentes mutaciones genéticas que otorgan susceptibilidad al desarrollo de MMC es indiscutido, así como la interacción entre las características genéticas del individuo y eventos ambientales. En el MMC familiar se han establecido dos genes de alta susceptibilidad con diferente penetrancia y frecuencia: CDK4 y CDKN2A. CDKN2A (Cyclin-dependent kinase inhibitor 2A) es el más importante gen de susceptibilidad a MMC, cuyas mutaciones se han identificado en aproximadamente un 40 por ciento de familias que presentan tres o más casos de MMC. Las características clínicas asociadas a la mutación de CDKN2A son número elevado de individuos afectados por MMC dentro de una familia, MMC primario múltiple y presencia conjunta de MMC y cáncer de páncreas dentro de una familia. En Chile de desconoce la frecuencia y tipos de mutaciones que afectan a CDKN2A en familias con predisposición a MMC familiar.

Cutaneous malignant melanoma (CMM) represents 4 percent of all malignant skin tumors and accounts for 80 percent of deaths related to cutaneous cancer. 5-year survival rates in individuals with metastatic disease is around 14 percent. An hereditary or familial variant of CMM has been identified. It is related to different mutations of so-called susceptibility genes as well as to interactions between genetic characteristics and environmental factors. Familial CMM is related to two genes of elevated susceptibility, penetrance, and frequency: CDK4 and CDKN2A (Cyclin-dependent kinase inhibitor 2A). CDKN2A is the most important susceptibility gene and its mutations have been identified in approximately 40 percent of the families bearing three or more members with CNM. Clinical features associated to CDKN2A mutations are elevated number of family members with CMN, multiple primary CMM, and the presence of both CMN and pancreatic cancer in the same family. In Chile, the incidence and mutation variants of CDKN2A in families with CMM is unknown.

Genes de predisposición al melanoma. Melanoma familiar / Melanoma predisposition genes: familial melanoma

El melanoma es un tumor que ha crecido en frecuencia y para el cual todavía no existe un tratamiento efectivo. En la génesis del melanoma participan factores externos, como la radiación ultravioleta (RUV), y factores genéticos. Si bien el estudio de los genes involucrados en el melanoma aun está en su inicio, se conocen en la actualidad múltiples mutaciones genéticas transmisibles que confieren a su portador mayor riesgo de desarrollar melanoma, tales como las del gen CDK-N2A, del receptor de la melanocortina 1 (MCR1) y las de los genes de la vía de las MAP kinasas, entre otras. Muchos de estos genes determinan la predisposición al melanoma familiar o al melanoma múltiple y aumentan la susceptibilidad a otros tumores no cutáneos. El conocimiento cada vez más exacto y complejo de la acción de cada gen en la génesis y en la progresión del melanoma permitirá en un futuro próximo la creación de tratamientos más eficaces y más específicos para cada paciente.

Cutaneous melanoma is an increasingly frequent tumor that, as yet,does not have a satisfactory treatment. Environmental risk factors such as ultraviolet radiation (UVR) and genetic factors as well, participate in its genesis. Although studies of the genes involved in melanoma are scarce, it is well known that multiple transmissible genetic mutations, such as CDKN2A gene, the melanocortin-receptor 1(MCR1) gene and the MAPK signaling pathway genes, among others,confer to their carrier greater risk of developing melanoma. Many of these genes determine the predisposition to familial or multiple melanoma and increase the susceptibility to noncutaneous tumors. The precise and more complex knowledge of the role of each gene in the genesis and progression of melanoma, will allow the development of more successful and specific treatments for each patient in thenear future.

Methylation status of ANAPC1, CDKN2A and TP53 promoter genes in individuals with gastric cancer

Gastric cancer is the forth most frequent malignancy and the second most common cause of cancer death worldwide. DNA methylation is the most studied epigenetic alteration, occurring through a methyl radical addition to the cytosine base adjacent to guanine. Many tumor genes are inactivated by DNA methylation in gastric cancer. We evaluated the DNA methylation status of ANAPC1, CDKN2A and TP53 by methylation-specific PCR in 20 diffuse- and 26 intestinal-type gastric cancer samples and 20 normal gastric mucosa in individuals from Northern Brazil. All gastric cancer samples were advanced stage adenocarcinomas. Gastric samples were surgically obtained at the João de Barros Barreto University Hospital, State of Pará, and were stored at -80°C before DNA extraction. Patients had never been submitted to chemotherapy or radiotherapy, nor did they have any other diagnosed cancer. None of the gastric cancer samples presented methylated DNA sequences for ANAPC1 and TP53. CDKN2A methylation was not detected in any normal gastric mucosa; however, the CDKN2A promoter was methylated in 30.4 percent of gastric cancer samples, with 35 percent methylation in diffuse-type and 26.9 percent in intestinal-type cancers. CDKN2A methylation was associated with the carcinogenesis process for ~30 percent diffuse-type and intestinal-type compared to non-neoplastic samples. Thus, ANAPC1 and TP53 methylation was probably not implicated in gastric carcinogenesis in our samples. CDKN2A can be implicated in the carcinogenesis process of only a subset of gastric neoplasias.

Significance of the Methylation of Cyclin-Dependent Kinase Inhibitor 2A gene in the Prognosis of Osteosarcoma / 대한정형외과학회잡지

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.

Dectection of p16~(CDKN2A) exon 1 methylation in experimental tongue carcinogenesis in rats / 实用口腔医学杂志

Objective: To detect methylation status of p16 CDKN2A exon 1 during experimental carcinogenesis in rats. Methods:Thirty male clean SD rats were fed with 0.02 g/L of 4-nitroquinoline-oxide (4NQO) in drinking water.13, 16 and 24 weeks after experiment the normal, moderate-severe dysplasia and invasive squamous cell carcinoma tissues were removed from their tongues respectively; then the methylation status of p16 CDKN2A exon 1 were detected by methylation-specific PCR(MSP). Results:A 123 bp-unmethylated product was amplified in all samples but the methylated product was not detected in any of the samples. Conclusion: The p16 CDKN2A exon 1 appeares to be unmethylated during carcinogenesis of tongue cancer in experimental rats.