DNA Copy Number Research Articles

The development of a custom RNA-sequencing panel for the identification of predictive and diagnostic biomarkers in glioma

PurposeVarious molecular profiles are needed to classify malignant brain tumors, including gliomas, based on the latest classification criteria of the World Health Organization, and their poor prognosis necessitates new therapeutic targets. The Todai OncoPanel 2 RNA Panel (TOP2-RNA) is a custom-target RNA-sequencing (RNA-seq) using the junction capture method to maximize the sensitivity of detecting 455 fusion gene transcripts and analyze the expression profiles of 1,390 genes. This study aimed to classify gliomas and identify their molecular targets using TOP2-RNA.MethodsA total of 124 frozen samples of malignant gliomas were subjected to TOP2-RNA for classification based on their molecular profiles and the identification of molecular targets.ResultsAmong 55 glioblastoma cases, gene fusions were detected in 11 cases (20%), including novel MET fusions. Seven tyrosine kinase genes were found to be overexpressed in 15 cases (27.3%). In contrast to isocitrate dehydrogenase (IDH) wild-type glioblastoma, IDH-mutant tumors, including astrocytomas and oligodendrogliomas, barely harbor fusion genes or gene overexpression. Of the 34 overexpressed tyrosine kinase genes, MDM2 and CDK4 in glioblastoma, 22 copy number amplifications (64.7%) were observed. When comparing astrocytomas and oligodendrogliomas in gene set enrichment analysis, the gene sets related to 1p36 and 19q were highly enriched in astrocytomas, suggesting that regional genomic DNA copy number alterations can be evaluated by gene expression analysis.ConclusionsTOP2-RNA is a highly sensitive assay for detecting fusion genes, exon skipping, and aberrant gene expression. Alterations in targetable driver genes were identified in more than 50% of glioblastoma. Molecular profiling by TOP2-RNA provides ample predictive, prognostic, and diagnostic biomarkers that may not be identified by conventional assays and, therefore, is expected to increase treatment options for individual patients with glioma.

Circulating levels of brain-derived neurotrophic factor (BDNF) in patients with bipolar disorders may be influenced by mitochondrial dysfunction, inflammation, and history of childhood trauma

BackgroundBipolar disorders are underpinned by deregulated immune processes including chronic inflammation. The crosstalk between immune factors and homeostatic molecules remains scarcely studied. The brain-derived neurotrophic factor is a neurotrophic molecule essential for neuronal and synaptic homeostasis. We explored correlations between BDNF levels and factors known to contribute to sustained inflammation, namely mitochondrial dysfunction (mtDNAcn), inflammation, and history of childhood maltreatment (CM). Methods175 patients with BD assessed either during acute mood episode or when euthymic and 85 healthy controls were included. CM history was assessed by the Childhood Trauma Questionnaire. Circulating levels of inflammatory molecules, BDNF and mitochondrial DNA copy number were determined. Correlations between the studied parameters were analyzed. ResultsLow levels of BDNF were observed during acute mood episodes (mania/mixed/depression). We observed that IL-12/23p40 levels and history of CM were negatively correlated with circulating BDNF levels in acutely ill patients. In euthymic patients we found that while TNFα levels correlate negatively with BDNF levels, that of IL16, VEGF and mtDNAcn were positively correlated. Linear regression showed that: (i) in the whole BD sample, low mtDNAcn levels were associated with low BDNF levels (ii) in acutely ill BD patients, high CTQ-AE scores, and high IL-12/23p40 levels were associated with low BDNF levels. (ⅲ) in euthymic BD patients, low mtDNAcn levels were associated with low BDNF levels. LimitationsWe acknowledge the relatively low sample size. ConclusionsWe identified some of the immune factors possibly influencing the levels of BDNF. Future studies involving larger cohorts are warranted.

Traumatic Brain Injury Alters the Trajectory of Age-Related Mitochondrial Change.

Some epidemiologic studies associate traumatic brain injury (TBI) with Alzheimer's disease (AD). To test whether a TBI-induced acceleration of age-related mitochondrial change could potentially mediate the reported TBI-AD association. We administered unilateral controlled cortical impact (CCI) or sham injuries to 5-month-old C57BL/6J and tau transgenic rTg4510 mice. In the non-transgenics, we assessed behavior (1-5 days, 1 month, and 15 months), lesion size (1 and 15 months), respiratory chain enzymes (1 and 15 months), and mitochondrial DNA copy number (mtDNAcn) (1 and 15 months) after CCI/sham. In the transgenics we quantified post-injury mtDNAcn and tangle burden. In the non-transgenics CCI caused acute behavioral deficits that improved or resolved by 1-month post-injury. Protein-normalized complex I and cytochrome oxidase activities were not significantly altered at 1 or 15 months, although complex I activity in the CCI ipsilesional cortex declined during that period. Hippocampal mtDNAcn was not altered by injury at 1 month, increased with age, and rose to the greatest extent in the CCI contralesional hippocampus. In the injured then aged transgenics, the ipsilesional hippocampus contained less mtDNA and fewer tangles than the contralesional hippocampus; mtDNAcn and tangle counts did not correlate. As mice age their brains increase mtDNAcn as part of a compensatory response that preserves mitochondrial function, and TBI enhances this response. TBI may, therefore, increase the amount of compensation required to preserve late-life mitochondrial function. If TBI does modify AD risk, altering the trajectory or biology of aging-related mitochondrial changes could mediate the effect.

Health risks of rare earth elements exposure: Impact on mitochondrial DNA copy number and micronucleus frequency

Toxic effects in terms of mitochondria and hereditary substances have been characterized in vitro for individual rare earth elements, while, the joint effects of mixed elements exposure in the population remain ambiguous. Based on the Occupational Chromate Exposure Dynamic Cohort of China, this study investigated the relationship between 15 blood rare earth elements (cerium, dysprosium, erbium, europium, gadolinium, holmium, lanthanum, lutetium, neodymium, praseodymium, samarium, terbium, thulium, yttrium, and ytterbium) and mitochondrial DNA copy number (MtDNACN) as well as peripheral blood lymphocyte micronucleus frequency (MNF). The elastic net was used to select elements highly correlated with effect indicators, whose dose-response relationships were further illustrated by restricted cubic splines. Bayesian kernel regression was employed to explore the combined effects of elements and the contributions of single element. The results showed that most rare earth elements were positively correlated with effect indicators, with yttrium showing the strongest association (β (95% CI): 0.139 (0.1089 – 0.189) for MtDNACN, 0.937 (0.345 – 1.684) for MNF). In the mixed exposure model, with the exposure level fixed at the 50th percentile as the reference, the effect estimates on MtDNACN and MNF increased by 0.228 and 0.598 units, respectively, at the 75th percentile. The single effect analysis implied that yttrium, lanthanum and terbium contributed the most to the elevation of MtDNACN, while yttrium posed the highest risk for genetic damage, accordingly, we provided recommendations to prioritize these elements of concern. In addition, we observed a chief mediating effect of MtDNACN on the elevation of MNF caused by lanthanum, whereas further mechanistic exploration is required to confirm this finding.

TMEM65 promotes gastric tumorigenesis by targeting YWHAZ to activate PI3K-Akt-mTOR pathway and is a therapeutic target.

Copy number alterations are crucial for the development of gastric cancer (GC). Here, we identified Transmembrane Protein 65 (TMEM65) amplification by genomic hybridization microarray to profile copy-number variations in GC. TMEM65 mRNA level was significantly up-regulated in GC compared to adjacent normal tissues, and was positively associated with TMEM65 amplification. High TMEM65 expression or DNA copy number predicts poor prognosis (P < 0.05) in GC. Furtherly, GC patients with TMEM65 amplification (n = 129) or overexpression (n = 78) significantly associated with shortened survival. Ectopic expression of TMEM65 significantly promoted cell proliferation, cell cycle progression and cell migration/invasion ability, but inhibited apoptosis (all P < 0.05). Conversely, silencing of TMEM65 in GC cells showed opposite abilities on cell function in vitro and suppressed tumor growth and lung metastasis in vivo (all P < 0.01). Moreover, TMEM65 depletion by VNP-encapsulated TMEM65-siRNA significantly suppressed tumor growth in subcutaneous xenograft model. Mechanistically, TMEM65 exerted oncogenic effects through activating PI3K-Akt-mTOR signaling pathway, as evidenced of increased expression of key regulators (p-Akt, p-GSK-3β, p-mTOR) by Western blot. YWHAZ (Tyrosine 3-Monooxygenase/Tryptophan 5-Monooxygenase) was identified as a direct downstream effector of TMEM65. Direct binding of TMEM65 with YWHAZ in the cytoplasm inhibited ubiquitin-mediated degradation of YWHAZ. Moreover, oncogenic effect of TMEM65 was partly dependent on YWHAZ. In conclusion, TMEM65 promotes gastric tumorigenesis by activating PI3K-Akt-mTOR signaling via cooperating with YWHAZ. TMEM65 overexpression may serve as an independent new biomarker and is a therapeutic target in GC.

Influence of Periodontitis on Th17/Treg Percentages in Patients with Rheumatoid Arthritis

Th17/Treg cells and P. gingivalis have been implicated in rheumatoid arthritis (RA) and periodontitis (PE). The aim was to determine and compare Th17/Treg cells and the influence of P. gingivalis on this cell population in healthy subjects (HS) and patients with PE, RA, and PE/RA. The number of copies of P. gingivalis was performed by real-time PCR and flow cytometry was used to analyze Th17/Treg cells. In this study, PE in patients with RA increased the number of DNA copies of P. gingivalis, increasing the percentages of Th17 cells, as well as the number and regulation of Treg cells. Periodontitis influences the immune system, increasing the percentage of pro-inflammatory cells in patients with rheumatoid arthritis, which may could influence the inflammatory condition of this population.

Umbilical cord blood thyroid hormones are inversely related to telomere length and mitochondrial DNA copy number

Hypothyroidism has been linked to reduced mortality rate and increased lifespan and health span. Telomere shortening, enhanced oxidative stress, and reduced cellular mitochondrial content are important hallmarks of aging shown to be related to age-associated diseases. It was proposed that the status of these markers in early life can be predictive of lifespan and the predisposition to certain age-associated disease in adulthood. Animal studies indicated that prenatal injection of thyroid hormones affects postnatal telomere length. Here, we sought to determine whether thyroid hormones TSH and fT4 are related to the telomere length, mitochondrial DNA copy number (mtDNAcn), and oxidative stress resistance marker GPX in the cord blood of newborns. In this study, we analyzed 70 mothers (18–42 years) and neonate dyads born in 2022 at the Nik Nafs maternity Hospital in Rafsanjan. The relative telomere length (RTL) and mtDNAcn were measured in the genomic DNA of cord blood leukocytes using real-time PCR. GPX enzyme activity was measured in the serum using colorimetric assays. In this study the correlation between these markers and the cord blood TSH and fT4 hormones were assessed using regression models. We found a reverse relationship between TSH levels and RTL in the cord blood of neonates. Additionally, our results displayed increased TSH levels associated with enhanced GPX activity. Regarding the mitochondrial DNA copy number, we found an indirect relationship between fT4 level and mtDNAcn only in male newborns. Future analyses of various oxidative stress markers, mitochondrial biogenesis status, telomerase activity, and the level of DNA damage are warranted to demonstrate the underlying mechanism of our observations.

Morphologic and Molecular Heterogeneity of High-grade Serous Carcinoma Precursor Lesions.

Serous tubal intraepithelial carcinoma (STIC) is the fallopian tube precursor lesion for most cases of pelvic high-grade serous carcinoma (HGSC). To date, the morphologic, molecular, and clinical heterogeneity of STIC and a less atypical putative precursor lesion, termed serous tubal intraepithelial lesion, has not been well characterized. Better understanding of precursor heterogeneity could impact the clinical management of women with incidental STICs (without concurrent carcinoma) identified in cases of prophylactic or opportunistic salpingectomy. This study analyzed morphologic and molecular features of 171 STICs and 21 serous tubal intraepithelial lesions. We assessed their histologic features, Ki-67 and p53 staining patterns, and genome-wide DNA copy number alterations. We classified all precursor lesions into 2 morphologic subtypes, one with a flat surface (Flat) and the other characterized by budding, loosely adherent, or detached (BLAD) morphology. On the basis of pathology review by a panel of 8 gynecologic pathologists, we found 87 BLAD, 96 Flat, and 9 indeterminate lesions. As compared with Flat lesions, BLAD lesions were more frequently diagnostic of STIC ( P <0.0001) and were found concurrently with HGSC ( P <0.0001). BLAD morphology was also characterized by higher Ki-67 proliferation index ( P <0.0001), presence of epithelial stratification ( P <0.0001), and increased lymphocyte density ( P <0.0001). BLAD lesions also exhibited more frequent DNA copy number gain/amplification at the CCNE1 or CMYC loci canonical to HGSCs ( P <0.0001). Both BLAD morphology and STIC diagnoses are independent risk factors for an elevated Ki-67 proliferation index. No correlation was observed between BLAD and Flat lesions with respect to patient age, presence of germline BRCA1/2 mutation, or p53 staining pattern. These findings suggest that tubal precursor lesions are morphologically and molecularly heterogeneous, laying the foundation for further studies on the pathogenesis of HGSC initiation and identifying histologic features predictive of poor patient outcomes.

Mitochondrial genetic variation and risk of chronic kidney disease and acute kidney injury in UK Biobank participants

Experimental models suggest an important role for mitochondrial dysfunction in the pathogenesis of chronic kidney disease (CKD) and acute kidney injury (AKI), but little is known regarding the impact of common mitochondrial genetic variation on kidney health. We sought to evaluate associations of inherited mitochondrial DNA (mtDNA) variation with risk of CKD and AKI in a large population-based cohort. We categorized UK Biobank participants who self-identified as white into eight distinct mtDNA haplotypes, which were previously identified based on their associations with phenotypes associated with mitochondrial DNA copy number, a measure of mitochondrial function. We used linear and logistic regression models to evaluate associations of these mtDNA haplotypes with estimated glomerular filtration rate by serum creatinine and cystatin C (eGFRCr-CysC, N = 362,802), prevalent (N = 416 cases) and incident (N = 405 cases) end-stage kidney disease (ESKD), AKI defined by diagnostic codes (N = 14,170 cases), and urine albumin/creatinine ratio (ACR, N = 114,662). The mean age was 57 ± 8 years and the mean eGFR was 90 ± 14 ml/min/1.73 m2. MtDNA haplotype was significantly associated with eGFR (p = 2.8E−12), but not with prevalent ESKD (p = 5.9E−2), incident ESKD (p = 0.93), AKI (p = 0.26), or urine ACR (p = 0.54). The association of mtDNA haplotype with eGFR remained significant after adjustment for diabetes mellitus and hypertension (p = 1.2E−10). When compared to the reference haplotype, mtDNA haplotypes I (β = 0.402, standard error (SE) = 0.111; p = 2.7E−4), IV (β = 0.430, SE = 0.073; p = 4.2E−9), and V (β = 0.233, SE = 0.050; p = 2.7E−6) were each associated with higher eGFR. Among self-identified white UK Biobank participants, mtDNA haplotype was associated with eGFR, but not with ESKD, AKI or albuminuria.

Abstract A036: The genetic landscape of head and neck cancer using brush biopsy

Abstract Oral potentially malignant disorders (OPMDs) are characterized by the presence of dysplasia and DNA copy number aberrations (CNAs) and subsequently progress to oral squamous cell carcinoma (OSCC). OSCC has a 5-year survival rate of only about 50% and varies greatly depending on site and stage. The combination of next-generation sequencing and advanced computational data analysis approaches has revolutionized our understanding of the genomic underpinnings of cancer development and progression. This can reveal individual targetable alterations, mutational load, complex mutation signatures, and tumor-specific antigens, which might inform the utilization of targeted therapies and early diagnosis. Oral brush biopsy represents an attractive non-invasive advantage in the characterization of premalignant and malignant oral lesions. Still, a sufficient number of abnormal cells is mandatory to diagnose. The current study tested the possibility of recreating the genomic landscape of oral cancer using brush biopsy. Materials and methods: As a pilot study to address this question, we analyzed matched tissue and brush biopsy of a patient with synchronous OPMD and OSCC. Brush and tissue biopsy was taken from normal epithelium, OPMD, and OSCC. We scrutinized the evolution and heterogeneity of these three brush/tissue biopsy pairs. The techniques used include whole exome sequencing (WES), mutational signature analysis to evaluate smoking and other aberrant signatures, copy number analysis (using ASCAT), and subclonal architecture reconstruction (using DPclust). Results: To analyze intra-tumoral heterogeneity (ITH), ASCAT and DPclust were used to call copy number aberrations (CNA) and cluster somatic SNVs based on cancer cell fraction (CCF), respectively. We observed a large number of shared SNVs between the OPMD and OSCC lesions suggesting a common evolutionary origin. After adjusting the ploidy and purity based on DPClust results, ASCAT profiles revealed differences in aberrant tumor cell fraction, gains and loss of heterozygosity, and copy number neutral events between the samples. It showed almost similar profiles between the respective lesions' brush and tissue samples, suggesting that brush biopsy can recapitulate the genomic landscape. The OPMD and OSCC lesions showed loss of heterozygosity in chromosome 9 and chromosome 17, which play a potential role in tumorigenesis with loss of function of P53. We inferred the subclonal architecture of each sample, illuminating the clonal relationships between samples and distinct lineages of precursor clones developing independently. Conclusion: Collectively, these results emphasize the possibility of using brush biopsy to reconstruct the genomic characterization of OPMD and OSCC show prognostic potential for diagnosis and intervention strategies, and accelerate progress in precision cancer care. Citation Format: Evit John, Tom Lesluyes, Xiao Zhao, Peter Van Loo. The genetic landscape of head and neck cancer using brush biopsy [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Cancer Evolution and Data Science: The Next Frontier; 2023 Dec 3-6; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_2):Abstract nr A036.

An integrative pan-cancer bioinformatics analysis of MSRB1 and its association with tumor immune microenvironment, prognosis, and immunotherapy

Methionine sulfoxide reductase B1 (MSRB1) is involved in the development and immune regulation of multiple tumors. However, the role of MSRB1 in the tumor microenvironment and its potential as a therapeutic target remain largely unknown. In this study, MSRB1 expression patterns were evaluated using pan-cancer RNA sequencing data from multiple cell lines, tissues, and single cells. The pan-cancer prognostic role of MSRB1 was assessed and the association between MSRB1 expression and certain cancer characteristics was analyzed. We showed that MSRB1 expression levels were increased in several types of cancer (P < 0.05) and in certain cell types (macrophages, dendritic cells, and malignant tumor cells). The upregulation of MSRB1 expression was due to DNA copy number amplification. Furthermore, MSRB1 was significantly associated with the activation of immune pathways (P < 0.05, NES > 0), immune cell infiltration, and expression of immune checkpoint molecules. In addition, high expression of MSRB1 was found in a series of in vivo and in vitro immunotherapy response models (P < 0.05), and showed resistance to most targeted drugs. Our results indicated that MSRB1 may regulate the tumor immune microenvironment through an immunoresponse and potentially influence cancer development. This could make it a promising predictive biomarker and therapeutic target for precise tumor immunotherapy.

Blood-Based mtDNA Quantification Indicates Population-Specific Differences Associated with Alzheimer's Disease-Related Risk.

Age is known to be the biggest risk factor for Alzheimer's disease (AD), and Mexican Americans (MAs), who are one of the fastest-aging populations in the United States, are at a uniquely elevated risk. Mitochondrial stress and dysfunction are key players in the progression of AD and are also known to be impacted by lifestyle and environmental exposures/stressors. This study aimed to identify population-specific differences in indicators of mitochondrial stress and dysfunction associated with AD risk that are detectable in the blood. Examining blood from both non-Hispanic white (NHW) and MA participants (N = 527, MA n = 284, NHW n = 243), mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) copy numbers were assessed through quantitative PCR. Data was stratified by population and sample type, and multiple linear regression analyses were performed to identify factors that may influence this phenotype of mitochondrial dysfunction. In the MA cohort, there was a significant relationship between cellular mtDNA:nDNA ratio and body mass index, CDR sum of boxes score, the APOEɛ2/ɛ3 genotype, and education. Further, there was a significant relationship between cell-free mtDNA copy number and both education and CDR sum score. In the NHW cohort, there was a significant relationship between cellular mtDNA:nDNA ratio and both age and CDR sum score. Age was associated with cell-free mtDNA in the NHW cohort. This evidence supports the existence of population-based differences in the factors that are predictive of this blood-based phenotype of mitochondrial dysfunction, which may be indicative of cognitive decline and AD risk.

Toxic metals and essential trace elements in placenta and their relation to placental function

IntroductionPlacental function is essential for fetal development, but it may be susceptible to malnutrition and environmental stressors. ObjectiveTo assess the impact of toxic and essential trace elements in placenta on placental function. MethodsToxic metals (cadmium, lead, mercury, cobalt) and essential elements (copper, manganese, zinc, selenium) were measured in placenta of 406 pregnant women in northern Sweden using ICP-MS. Placental weight and birth weight were obtained from hospital records and fetoplacental weight ratio was used to estimate placental efficiency. Placental relative telomere length (TL) and mitochondrial DNA copy number (mtDNAcn) were determined by quantitative PCR (n = 285). Single exposure-outcome associations were evaluated using linear or spline regression, and joint associations and interactions with Bayesian kernel machine regression (BKMR), all adjusted for sex, maternal smoking, and age or BMI. ResultsMedian cadmium, mercury, lead, cobalt, copper, manganese, zinc, and selenium concentrations in placenta were 3.2, 1.8, 4.3, 2.3, 1058, 66, 10626, and 166 μg/kg, respectively. In the adjusted regression, selenium (>147 μg/kg) was inversely associated with placental weight (B: −158; 95 % CI: −246, −71, per doubling), as was lead at low selenium (B: −23.6; 95 % CI: −43.2, −4.0, per doubling). Manganese was positively associated with placental weight (B: 41; 95 % CI: 5.9, 77, per doubling) and inversely associated with placental efficiency (B: −0.01; 95 % CI: −0.019, −0.004, per doubling). Cobalt was inversely associated with mtDNAcn (B: −11; 95 % CI: −20, −0.018, per doubling), whereas all essential elements were positively associated with mtDNAcn, individually and joint. ConclusionAmong the toxic metals, lead appeared to negatively impact placental weight and cobalt decreased placental mtDNAcn. Joint essential element concentrations increased placental mtDNAcn. Manganese also appeared to increase placental weight, but not birth weight. The inverse association of selenium with placental weight may reflect increased transport of selenium to the fetus in late gestation.

Biological aging markers in blood and brain tissue indicate age acceleration in alcohol use disorder.

Alcohol use disorder (AUD) is associated with increased mortality and morbidity risk. A reason for this could be accelerated biological aging, which is strongly influenced by disease processes such as inflammation. As recent studies of AUD show changes in DNA methylation and gene expression in neuroinflammation-related pathways in the brain, biological aging represents a potentially important construct for understanding the adverse effects of substance use disorders. Epigenetic clocks have shown accelerated aging in blood samples from individuals with AUD. However, no systematic evaluation of biological age measures in AUD across different tissues and brain regions has been undertaken. As markers of biological aging (BioAge markers), we assessed Levine's and Horvath's epigenetic clocks, DNA methylation telomere length (DNAmTL), telomere length (TL), and mitochondrial DNA copy number (mtDNAcn) in postmortem brain samples from Brodmann Area 9 (BA9), caudate nucleus, and ventral striatum (N = 63-94), and in whole blood samples (N = 179) of individuals with and without AUD. To evaluate the association between AUD status and BioAge markers, we performed linear regression analyses while adjusting for covariates. The majority of BioAge markers were significantly associated with chronological age in all samples. Levine's epigenetic clock and DNAmTL were indicative of accelerated biological aging in AUD in BA9 and whole blood samples, while Horvath's showed the opposite effect in BA9. No significant association of AUD with TL and mtDNAcn was detected. Measured TL and DNAmTL showed only small correlations in blood and none in brain. The present study is the first to simultaneously investigate epigenetic clocks, telomere length, and mtDNAcn in postmortem brain and whole blood samples in individuals with AUD. We found evidence for accelerated biological aging in AUD in blood and brain, as measured by Levine's epigenetic clock, and DNAmTL. Additional studies of different tissues from the same individuals are needed to draw valid conclusions about the congruence of biological aging in blood and brain.

Chromosomal copy number amplification-driven Linc01711 contributes to gastric cancer progression through histone modification-mediated reprogramming of cholesterol metabolism.

Chromosome gains or localized amplifications are frequently observed in human gastric cancer (GC) and are major causes of aberrant oncogene activation. However, the significance of long non-coding RNAs (LncRNAs) in the above process is largely unknown. The copy number aberrations (CNAs) data of GC samples were downloaded and analyzed from the TCGA database. qRT-PCR and fluorescence in situ hybridization were used to evaluate the expression of Linc01711 in GC. The effects of Linc01711 on GC progression were investigated through in vitro and in vivo assays. The mechanism of Linc01711 action was explored through transcriptome sequencing, chromatin immunoprecipitation sequencing, RNA immunoprecipitation, RNA pull-down and chromatin isolation by RNA purification (ChIRP) assays. We report for the first time a novel DNA copy number amplification-driven LncRNA on chromosome 20q13, designated Linc01711 in human GC, which is highly associated with malignant features. Functionally, Linc01711 significantly accelerates the proliferation and metastasis of GC. Mechanistically, Linc01711 acts as a modular scaffold to promote the binding of histone acetyltransferase HBO1 and histone demethylase KDM9. By coordinating the localization of the HBO1/KDM9 complex, Linc01711 specifies the histone modification pattern on the target genes, such as LPCAT1, and consequently facilitates the cholesterol synthesis, thereby contributing to tumor progression. Our findings suggest that copy number amplification-driven Linc01711 may serve as a promising prognostic predictor for GC patients and targeting Linc01711-related cholesterol metabolism pathway may be meaningful in anticancer strategies.

DOP58 Human Leukocyte Antigen drives methylation signatures in primary sclerosing cholangitis but not in IgG4-related sclerosing cholangitis

Abstract Background Primary sclerosing cholangitis (PSC) and IgG4-related sclerosing cholangitis (IgG4-SC) are fibro-inflammatory hepatobiliary conditions, with genetic, environmental, and immunologic risk factors, in which epigenetic alterations may provide insights into pathophysiology and novel biomarkers. Methods Whole blood DNA methylation profiling and genotyping was performed in 70 PSC (57 with PSC and ulcerative colitis (PSC-UC)), 56 IgG4-SC, 66 UC, and 88 healthy controls (HC), with MethylationEPIC and Global Screening arrays (Illumina). UC patients were selected to match the clinical characteristics of UC in the PSC-UC cohort. Methylation data was processed with Minfi and limma, genotyping analysed with Plink, and meQTLs with linear models. Results There were 31, 13 and 8 Holm-significant methylation differences between PSC and controls, IgG4-SC and controls, and UC and controls respectively (8 probes shared). Inflammatory genes (including CEP97, IFNAR1, TXK, and SMAD2) and methylation loci associated with mitochondrial DNA copy number (C5orf36, PYY, and MTRNR2L1) were predominant. Sub-analyses of PSC-UC and PSC without IBD detected 25 and 4 differences with HC respectively (p&amp;lt;0.05). DNA methylation age acceleration was observed in IgG4-SC, but not on PSC, PSC-IBD or UC. MeQTL analyses to identify genetic determinants of methylation revealed a strong HLA signal in PSC but not IgG4-SC, with 19/40 significant findings in 5 clusters (HLA-DRB1, HLA-DPB1, BTNL2, CSNK2B, and SFTA2). There was no overlap with the 36 meQTLs found in IgG4-SC. Using multivariate adaptive shrinkage for meQTLs in PSC, IgG4-SC, and UC yielded 5505 meQTLs with local false discovery rate &amp;lt;0.05 in at least one condition, with 1230 unique to PSC, and 1043 to IgG4-SC. Conclusion We identify characteristic epigenetic alterations in PSC and IgG4-related disease, providing insights into disease pathogenesis; and highlight the potential interaction with germ-line variation.

Nuclear and mitochondrial genetic variants associated with mitochondrial DNA copy number

Mitochondrial DNA copy number (mtDNA-CN) is a biomarker for mitochondrial dysfunction associated with several diseases. Previous genome-wide association studies (GWAS) have been performed to unravel underlying mechanisms of mtDNA-CN regulation. However, the identified gene regions explain only a small fraction of mtDNA-CN variability. Most of this data has been estimated from microarrays based on various pipelines. In the present study we aimed to (1) identify genetic loci for qPCR-measured mtDNA-CN from three studies (16,130 participants) using GWAS, (2) identify potential systematic differences between our qPCR derived mtDNA-CN measurements compared to the published microarray intensity-based estimates, and (3) disentangle the nuclear from mitochondrial regulation of the mtDNA-CN phenotype. We identified two genome-wide significant autosomal loci associated with qPCR-measured mtDNA-CN: at HBS1L (rs4895440, p = 3.39 × 10–13) and GSDMA (rs56030650, p = 4.85 × 10–08) genes. Moreover, 113/115 of the previously published SNPs identified by microarray-based analyses were significantly equivalent with our findings. In our study, the mitochondrial genome itself contributed only marginally to mtDNA-CN regulation as we only detected a single rare mitochondrial variant associated with mtDNA-CN. Furthermore, we incorporated mitochondrial haplogroups into our analyses to explore their potential impact on mtDNA-CN. However, our findings indicate that they do not exert any significant influence on our results.

Rare and common genetic determinants of mitochondrial function determine severity but not risk of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease involving selective vulnerability of energy-intensive motor neurons (MNs). It has been unclear whether mitochondrial function is an upstream driver or a downstream modifier of neurotoxicity. We separated upstream genetic determinants of mitochondrial function, including genetic variation within the mitochondrial genome or autosomes; from downstream changeable factors including mitochondrial DNA copy number (mtCN). Across three cohorts including 6,437 ALS patients, we discovered that a set of mitochondrial haplotypes, chosen because they are linked to measurements of mitochondrial function, are a determinant of ALS survival following disease onset, but do not modify ALS risk. One particular haplotype appeared to be neuroprotective and was significantly over-represented in two cohorts of long-surviving ALS patients. Causal inference for mitochondrial function was achievable using mitochondrial haplotypes, but not autosomal SNPs in traditional Mendelian randomization (MR). Furthermore, rare loss-of-function genetic variants within, and reduced MN expression of, ACADM and DNA2 lead to ∼50 % shorter ALS survival; both proteins are implicated in mitochondrial function. Both mtCN and cellular vulnerability are linked to DNA2 function in ALS patient-derived neurons. Finally, MtCN responds dynamically to the onset of ALS independently of mitochondrial haplotype, and is correlated with disease severity. We conclude that, based on the genetic measures we have employed, mitochondrial function is a therapeutic target for amelioration of disease severity but not prevention of ALS.

Telomere Length, Mitochondrial DNA, and Micronucleus Yield in Response to Oxidative Stress in Peripheral Blood Mononuclear Cells.

Telomere shortening, chromosomal damage, and mitochondrial dysfunction are major initiators of cell aging and biomarkers of many diseases. However, the underlying correlations between nuclear and mitochondrial DNA alterations remain unclear. We investigated the relationship between telomere length (TL) and micronucleus (MN) and their association with mitochondrial DNA copy number (mtDNAcn) in peripheral blood mononuclear cells (PBMCs) in response to 100 μM and 200 μM of hydrogen peroxide (H2O2) at 44, 72, and 96 h. Significant TL shortening was observed after both doses of H2O2 and at all times (all p < 0.05). A concomitant increase in MN was found at 72 h (p < 0.01) and persisted at 96 h (p < 0.01). An increase in mtDNAcn (p = 0.04) at 200 µM of H2O2 was also found. In PBMCs treated with 200 µM H2O2, a significant inverse correlation was found between TL and MN (r = -0.76, p = 0.03), and mtDNA content was directly correlated with TL (r = 0.6, p = 0.04) and inversely related to MN (r = -0.78, p = 0.02). Telomere shortening is the main triggering mechanism of chromosomal damage in stimulated T lymphocytes under oxidative stress. The significant correlations between nuclear DNA damage and mtDNAcn support the notion of a telomere-mitochondria axis that might influence age-associated pathologies and be a target for the development of relevant anti-aging drugs.

Research on quantitative detection technology of raccoon-derived ingredient adulteration in sausage products.

This project presents a quantitative detection method to identify raccoon-derived ingredient adulteration in sausage products. The specific copy gene of the raccoon was selected as the target gene. According to the specificity of its primer and probe, the quantitative detection method of raccoon microdrops by droplet digital PCR was established. In addition, the accuracy of the proposed method was verified by artificially mixed samples, and the applicability of this method was tested based on the commercially available products. The experimental results indicate that the raccoon mass (M) and raccoon-extracted DNA concentration have a good linear relationship when the sample content is 5-100 mg, and there is also a significant linear relationship between DNA content and DNA copy number (C) with R 2 = .9982. Therefore, using DNA concentration as the median signal, the conversion equation between raw raccoon mass (M) and DNA copy number (C) could be obtained as follows: M = (C + 177.403)/16.954. The detection of artificially mixed samples and commercial samples shows that the method is accurate and suitable for quantitative adulteration detection of various sausage products in the market.