Characterization of a base-resolution common deletion region within CDKN2A gene in cancers and its biological and clinical significances

Background: Frequency of somatic copy number deletion of CDKN2A gene is upto 60% in human esophageal squamous cell cancer. However, it is unknown whether CDKN2A deletion could be a biomarker for esophageal squamous cell dysplasia (ESCdys) due to absence of a feasible detection method. Methods: Information on base-resolution common deletion region (CDR) for CDKN2A were extracted from published articles and confirmed with whole genome sequencing (WGS). A quantitative PCR targeted to the CDR (P16-Light) was established and used to detect CDKN2A copy number in ESCdys biopsies from patients (n=205) enrolled in a multicentre follow-up study. Results: A 5.1-kb CDR from the CDKN2A/P16INK4A promoter to intron-2 was firstly characterized in 90% (83/92) of cancer cell lines and confirmed with WGS. The CDR covers CDKN2A exon-2 which is the essential coding exon for both P16INK4a and P14ARF. And CDKN2A exon-2 deletion markedly promoted the proliferation and invasion and inhibited the apoptosis of HEK293T cells. In the follow-up study, both somatic CDKN2A deletion and amplification are prevalent in mild/moderate (m/M) ESCdys. CDKN2A deletion was less common among 70 patients whose ESCdys regressed than among 135 patients whose ESCdys progressed or remained stable, and CDKN2A amplification was more common in the patients who regressed than in the patients whose m/M ESCdys persisted or progressed over a median of 37 months of follow-up (p<0.0001). Conclusion: There is A 5.1-kb CDR within CDKN2A gene in many cancers. CDR deletion could inactivate both P16INK4a and P14ARF and associate with prognosis of ESCdys.

Altered Glycolytic Capacity, Not Mitochondrial Common Deletion, May Be Responsible For Age-Related Increases in Rat Intestinal Permeability

Some evidence points to a link between aging-related increased intestinal permeability and mitochondrial dysfunction in in-vivo models. Several studies have also demonstrated age-related accumulation of the of specific deletion 4834-bp of “common” mitochondrial DNA (mtDNA) in various rat tissues and suggest that this deletion may disrupt mitochondrial metabolism. The present study aimed to investigate possible associations among the mtDNA common deletion, mitochondrial function, intestinal permeability, and aging in rats. Our data showed were no significant differences in the abundance of mtDNA4834 deletions in intestinal tissue between young adult ((4-month-old) and aged (24-month-old) rats). In addition, Spearman's correlation coefficients of measured parameters in serum samples for intestinal permeability did not correlate with deletion frequency and measured levels of mitochondrial energy function parameters. However, the tissue lactate/pyruvate ratio (L/P) was three times lower in old rats than in young rats. Additionally, there were significant negative correlations between intestinal permeability parameters and L/P ratios. Considering our findings showing that the intestinal tissues of aged rats are not prone to accumulate mtDNA common deletion, we suggest that this mutation does not explain the age-related increase in intestinal permeability. Some data support the thought that altered glycolytic capacity could be a possible mechanism linked to increased intestinal permeability with age.

Activation of NF-κB signaling via cytosolic mitochondrial RNA sensing in kerotocytes with mitochondrial DNA common deletion

Scar formation as a result of corneal wound healing is a leading cause of blindness. It is a challenge to understand why scar formation is more likely to occur in the central part of the cornea as compared to the peripheral part. The purpose of this study was to unravel the underlying mechanisms. We applied RNA-seq to uncover the differences of expression profile in keratocytes in the central/peripheral part of the cornea. The relative quantity of mitochondrial RNA was measured by multiplex qPCR. The characterization of mitochondrial RNA in the cytoplasm was confirmed by immunofluoresence microscope and biochemical approach. Gene expression was analyzed by western blot and RT qPCR. We demonstrate that the occurrence of mitochondrial DNA common deletion is greater in keratocytes from the central cornea as compared to those of the peripheral part. The keratocytes with CD have elevated oxidative stress levels, which leads to the leakage of mitochondrial double-stranded RNA into the cytoplasm. The cytoplasmic mitochondrial double-stranded RNA is sensed by MDA5, which induces NF-κB activation. The NF-κB activation thereafter induces fibrosis-like extracellular matrix expressions and IL-8 mRNA transcription. These results provide a novel explanation of the different clinical outcome in different regions of the cornea during wound healing.

Mitochondrial Common Deletion Level in Blood: New Insight Into the Effects of Age and Body Mass Index

Background: Age-related decrease in mitochondrial activity has been reported in several tissues. Reactive Oxygen Species (ROS) produced from defected mitochondria lead to aging and accumulate through time. However, studies about the mitochondrial DNA mutation level in blood are contradictory. Other lifestyle factors may modify the effects of age in post-mitotic tissues such as blood. The BMI represents the sum of the various lifestyle factors. Objective: We proposed that age, obesity and mtDNA deletion are three ROS producing factors, which may interact with each other and induce senescence. Methods: In a cross-sectional study, 172 male and female volunteers without known mitochondrial diseases were selected and the presence of common mitochondrial 4977bp deletion (ΔmtDNA4977) evaluated using Nested-PCR. Results: Our results showed that a high percentage of samples (54.06%) harbor common deletion in blood. Furthermore, both BMI and the ΔmtDNA4977 levels significantly decrease with age. The chronological age, BMI and ΔmtDNA4977 reciprocally affect each other. Conclusion: Our data suggest that age affects purifying selection and BMI, which may influence the relative level of the mtDNA common deletion in blood.