Oxidative stress induces release of mitochondrial DNA into the extracellular space in human placental villous trophoblast BeWo cells.

Circulating cell-free mitochondrial DNA (ccf-mtDNA) is an indicator of cell death, inflammation, and oxidative stress. ccf-mtDNA differs in pregnancies with placental dysfunction from healthy pregnancies and the direction of this difference depends on gestational age and method of mtDNA quantification. Reactive oxygen species (ROS) trigger release of mtDNA from non-placental cells; yet it is unknown whether trophoblast cells release mtDNA in response to oxidative stress, a common feature of pregnancies with placental pathology. We hypothesized that oxidative stress would induce cell death and release of mtDNA from trophoblast cells. BeWo cells were treated with antimycin A (10-320 mM) or rotenone (0.2-50 mM) to induce oxidative stress. A multiplex real-time quantitative PCR (qPCR) assay was used to quantify mtDNA and nuclear DNA in membrane bound, non-membrane bound, and vesicular-bound forms in cell culture supernatants and cell lysates. Treatment with antimycin A increased ROS (p<0.0001), induced cell necrosis (p=0.0004) but not apoptosis (p=0.6471) and was positively associated with release of membrane-bound and non-membrane bound mtDNA (p<0.0001). Antimycin A increased mtDNA content in exosome-like extracellular vesicles (vesicular-bound form; p=0.0019) and reduced autophagy marker expression (LC3A/B, p=0.0002; p62, p<0.001). Rotenone treatment did not influence mtDNA release or cell death (p>0.05). Oxidative stress induces release of mtDNA into the extracellular space and causes non-apoptotic cell death and a reduction in autophagy markers in BeWo cells, an established in vitro model of human trophoblast cells. Intersection between autophagy and necrosis may mediate the release of mtDNA from the placenta in pregnancies exposed to oxidative stress.

Oxidative stress induces release of mitochondrial DNA into the extracellular space in human placental villous trophoblast BeWo cells.

Circulating cell-free mitochondrial DNA (ccf-mtDNA) is an indicator of cell death, inflammation, and oxidative stress. ccf-mtDNA differs in pregnancies with placental dysfunction from healthy pregnancies and the direction of this difference depends on gestational age and method of mtDNA quantification. Reactive oxygen species (ROS) trigger release of mtDNA from non-placental cells; yet it is unknown whether trophoblast cells release mtDNA in response to oxidative stress, a common feature of pregnancies with placental pathology. We hypothesized that oxidative stress would induce cell death and release of mtDNA from trophoblast cells. BeWo cells were treated with antimycin A (10-320 µM) or rotenone (0.2-50 µM) to induce oxidative stress. A multiplex real-time quantitative PCR (qPCR) assay was used to quantify mtDNA and nuclear DNA in membrane bound, non-membrane bound, and vesicular-bound forms in cell culture supernatants and cell lysates. Treatment with antimycin A increased ROS (p<0.0001), induced cell necrosis (p=0.0004) but not apoptosis (p=0.6471) and was positively associated with release of membrane-bound and non-membrane bound mtDNA (p<0.0001). Antimycin A increased mtDNA content in exosome-like extracellular vesicles (vesicular-bound form; p=0.0019) and reduced autophagy marker expression (LC3A/B, p=0.0002; p62, p<0.001). Rotenone treatment did not influence mtDNA release or cell death (p>0.05). Oxidative stress induces release of mtDNA into the extracellular space and causes non-apoptotic cell death and a reduction in autophagy markers in BeWo cells, an established in vitro model of human trophoblast cells. Intersection between autophagy and necrosis may mediate the release of mtDNA from the placenta in pregnancies exposed to oxidative stress. NEW & NOTEWORTHY: This is the first study to test whether trophoblast cells release mitochondrial DNA in response to oxidative stress and to identify mechanisms of release and biological forms of mtDNA from this cellular type. This research identifies potential cellular mechanisms that can be used in future investigations to establish the source and biomarker potential of circulating mitochondrial DNA in preclinical experimental models and humans.

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

BACKGROUND: 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. OBJECTIVE: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Mitochondrial SOS: how mtDNA may act as a stress signal in Alzheimer's disease.

BACKGROUND: Alterations in mitochondrial DNA (mtDNA) levels have been observed in Alzheimer's disease and are an area of research that shows promise as a useful biomarker. It is well known that not only are the mitochondria a key player in producing energy for the cell, but they also are known to interact in other important intracellular processes as well as extracellular signaling and communication. BODY: This mini review explores how cells use mtDNA as a stress signal, particularly in Alzheimer's disease. We investigate the measurement of these mtDNA alterations, the mechanisms of mtDNA release, and the immunological effects from the release of these stress signals. CONCLUSION: Literature indicates a correlation between the release of mtDNA in Alzheimer's disease and increased immune responses, showing promise as a potential biomarker. However, several questions remain unanswered and there is great potential for future studies in this area.

Integrative blood-based characterization of oxidative mitochondrial DNA damage variants implicates Mexican American's metabolic risk for developing Alzheimer's disease.

Alzheimer's Disease (AD) continues to be a leading cause of death in the US. As the US aging population (ages 65 +) expands, the impact will disproportionately affect vulnerable populations, e.g., Hispanic/Latino population, due to their AD-related health disparities. Age-related regression in mitochondrial activity and ethnic-specific differences in metabolic burden could potentially explain in part the racial/ethnic distinctions in etiology that exist for AD. Oxidation of guanine (G) to 8-oxo-guanine (8oxoG) is a prevalent lesion and an indicator of oxidative stress and mitochondrial dysfunction. Damaged mtDNA (8oxoG) can serve as an important marker of age-related systemic metabolic dysfunction and upon release into peripheral circulation may exacerbate pathophysiology contributing to AD development and/or progression. Analyzing blood samples from Mexican American (MA) and non-Hispanic White (NHW) participants enrolled in the Texas Alzheimer's Research & Care Consortium, we used blood-based measurements of 8oxoG from both buffy coat PBMCs and plasma to determine associations with population, sex, type-2 diabetes, and AD risk. Our results show that 8oxoG levels in both buffy coat and plasma were significantly associated with population, sex, years of education, and reveal a potential association with AD. Furthermore, MAs are significantly burdened by mtDNA oxidative damage in both blood fractions, which may contribute to their metabolic vulnerability to developing AD.

Maternal and fetal mitochondrial gene dysregulation in hypertensive disorders of pregnancy.

Mitochondrial dysfunction has been implicated in pregnancy-induced hypertension (PIH). The role of mitochondrial gene dysregulation in PIH, and consequences for maternal-fetal interactions, remain elusive. Here, we investigated mitochondrial gene expression and dysregulation in maternal and placental tissues from pregnancies with and without PIH; further, we measured circulating mitochondrial DNA (mtDNA) mutational load, an index of mtDNA integrity. Differential gene expression analysis followed by Time Course Gene Set Analysis (TcGSA) was conducted on publicly available high throughput sequencing transcriptomic data sets. Mutational load analysis was carried out on peripheral mononuclear blood cells from healthy pregnant individuals and individuals with preeclampsia. Thirty mitochondrial differentially expressed genes (mtDEGs) were detected in the maternal cell-free circulating transcriptome, whereas nine were detected in placental transcriptome from pregnancies with PIH. In PIH pregnancies, maternal mitochondrial dysregulation was associated with pathways involved in inflammation, cell death/survival, and placental development, whereas fetal mitochondrial dysregulation was associated with increased production of extracellular vesicles (EVs) at term. Mothers with preeclampsia did not exhibit a significantly different degree of mtDNA mutational load. Our findings support the involvement of maternal mitochondrial dysregulation in the pathophysiology of PIH and suggest that mitochondria may mediate maternal-fetal interactions during healthy pregnancy.NEW & NOTEWORTHY This study identifies aberrant maternal and fetal expression of mitochondrial genes in pregnancies with gestational hypertension and preeclampsia. Mitochondrial gene dysregulation may be a common etiological factor contributing to the development of de novo hypertension in pregnancy-associated hypertensive disorders.

Integrative Blood-Based Characterization of Oxidative Mitochondrial DNA Damage Variants Implicates Mexican Americans' Metabolic Risk for Developing Alzheimer's Disease.

Alzheimer's Disease (AD) continues to be a leading cause of death in the US. As the US aging population (ages 65+) expands, the impact will disproportionately affect vulnerable populations, e.g., Hispanic/Latinx population, due to their AD-related health disparities. Age-related regression in mitochondrial activity and ethnic-specific differences in metabolic burden could potentially explain in part the racial/ethnic distinctions in etiology that exist for AD. Oxidation of guanine (G) to 8-oxo-guanine (8oxoG) is a prevalent lesion and an indicator of oxidative stress and mitochondrial dysfunction. Damaged mtDNA (8oxoG) can serve as an important marker of age-related systemic metabolic dysfunction and upon release into peripheral circulation may exacerbate pathophysiology contributing to AD development and/or progression. Analyzing blood samples from Mexican American (MA) and non-Hispanic White (NHW) participants enrolled in the Texas Alzheimer's Research & Care Consortium, we used blood-based measurements of 8oxoG from both buffy coat PBMCs and plasma to determine associations with population, sex, type-2 diabetes, and AD risk. Our results show that 8oxoG levels in both buffy coat and plasma were significantly associated with population, sex, years of education, and reveal a potential association with AD. Furthermore, MAs are significantly burdened by mtDNA oxidative damage in both blood fractions, which may contribute to their metabolic vulnerability to developing AD.

Mitochondrial DNA oxidative mutations are elevated in Mexican American women potentially implicating Alzheimer's disease.

Mexican Americans (MAs) are the fastest-growing Hispanic population segment in the US; as this population increases in age, so will the societal burden of age-related diseases such as Alzheimer's disease (AD). Mitochondrial DNA (mtDNA) damage may be implicated in MA AD risk since metabolic comorbidities are more prevalent in this group. Oxidative damage to guanosine (8oxoG) is one of the most prevalent DNA lesions and a putative indicator of mitochondrial dysfunction. Testing blood samples from participants of the Texas Alzheimer's Research and Care Consortium, we found mtDNA 8oxoG mutational load to be significantly higher in MAs compared to non-Hispanic whites and that MA females are differentially affected. Furthermore, we identified specific mtDNA haplotypes that confer increased risk for oxidative damage and suggestive evidence that cognitive function may be related to 8oxoG burden. Our understanding of these phenomena will elucidate population- and sex-specific mechanisms of AD pathogenesis, informing the development of more precise interventions and therapeutic approaches for MAs with AD in the future.

Sex differences in neurobehavioral consequences of methamphetamine exposure in adult mice.

RATIONALE: Recreational and medical use of stimulants is increasing, and their use may increase susceptibility to aging and promote neurobehavioral impairments. The long-term consequences of these psychostimulants and how they interact with age have not been fully studied. OBJECTIVES: Our study investigated whether chronic exposure to the prototypical psychostimulant, methamphetamine (METH), at doses designed to emulate human therapeutic dosing, would confer a pro-oxidizing redox shift promoting long-lasting neurobehavioral impairments. METHODS: Groups of 4-month-old male and female C57BL/6 J mice were administered non-contingent intraperitoneal injections of either saline or METH (1.4 mg/kg) twice a day for 4 weeks. Mice were randomly assigned to one experimental group: (i) short-term cognitive assessments (at 5 months), (ii) long-term cognitive assessments (at 9.5 months), and (ii) longitudinal motor assessments (at 5, 7, and 9 months). Brain regions were assessed for oxidative stress and markers of neurotoxicity after behavior testing. RESULTS: Chronic METH exposure induced short-term effects on associative memory, gait speed, dopamine (DA) signaling, astrogliosis in females, and spatial learning and memory, balance, DA signaling, and excitotoxicity in males. There were no long-term effects of chronic METH on cognition; however, it decreased markers of excitotoxicity in the striatum and exacerbated age-associated motor impairments in males. CONCLUSION: In conclusion, cognitive and motor functions were differentially and sex-dependently affected by METH exposure, and oxidative stress did not seem to play a role in the observed behavioral outcomes. Future studies are necessary to continue exploring the long-term neurobehavioral consequences of drug use in both sexes and the relationship between aging and drugs.

Reduced Maternal Circulating Cell-Free Mitochondrial DNA Is Associated With the Development of Preeclampsia.

Background Circulating cell-free mitochondrial DNA (ccf-mtDNA) is a damage-associated molecular pattern that reflects cell stress responses and tissue damage, but little is known about ccf-mtDNA in preeclampsia. The main objectives of this study were to determine (1) absolute concentrations of ccf-mtDNA in plasma and mitochondrial DNA content in peripheral blood mononuclear cells and (2) forms of ccf-mtDNA transport in blood from women with preeclampsia and healthy controls. In addition, we sought to establish the association between aberrance in circulating DNA-related metrics, including ccf-mtDNA and DNA clearance mechanisms, and the clinical diagnosis of preeclampsia using bootstrapped penalized logistic regression. Methods and Results Absolute concentrations of ccf-mtDNA were reduced in plasma from women with preeclampsia compared with healthy controls (P≤0.02), while mtDNA copy number in peripheral blood mononuclear cells did not differ between groups (P>0.05). While the pattern of reduced ccf-mtDNA in patients with preeclampsia remained, DNA isolation from plasma using membrane lysis buffer resulted in 1000-fold higher ccf-mtDNA concentrations in the preeclampsia group (P=0.0014) and 430-fold higher ccf-mtDNA concentrations in the control group (P<0.0001). Plasma from women with preeclampsia did not induce greater Toll-like receptor-9-induced nuclear factor kappa-light-chain enhancer of activated B cells-dependent responses in human embryonic kidney 293 cells overexpressing the human TLR-9 gene (P>0.05). Penalized regression analysis showed that women with preeclampsia were more likely to have lower concentrations of ccf-mtDNA as well as higher concentrations of nuclear DNA and DNase I compared with their matched controls. Conclusions Women with preeclampsia have aberrant circulating DNA dynamics, including reduced ccf-mtDNA concentrations and DNA clearance mechanisms, compared with gestational age-matched healthy pregnant women.

Circulating Cell-Free Mitochondrial DNA in Pregnancy.

Circulating cell-free mitochondrial DNA (ccf-mtDNA) released upon cell injury or death stimulates diverse pattern recognition receptors to activate innate immune responses and initiate systemic inflammation. In this review, we discuss the temporal changes of ccf-mtDNA during pregnancy and its potential contribution to adverse pregnancy outcomes in pregnancy complications.

Accurate profiling of forensic autosomal STRs using the Oxford Nanopore Technologies MinION device.

The high variability characteristic of short tandem repeat (STR) markers is harnessed for human identification in forensic genetic analyses. Despite the power and reliability of current typing techniques, sequence-level information both within and around STRs are masked in the length-based profiles generated. Forensic STR typing using next generation sequencing (NGS) has therefore gained attention as an alternative to traditional capillary electrophoresis (CE) approaches. In this proof-of-principle study, we evaluate the forensic applicability of the newest and smallest NGS platform available - the Oxford Nanopore Technologies (ONT) MinION device. Although nanopore sequencing on the handheld MinION offers numerous advantages, including low startup cost and on-site sample processing, the relatively high error rate and lack of forensic-specific analysis software has prevented accurate profiling across STR panels in previous studies. Here we present STRspy, a streamlined method capable of producing length- and sequence-based STR allele designations from noisy, error-prone third generation sequencing reads. To assess the capabilities of STRspy, seven reference samples (female: n = 2; male: n = 5) were amplified at 15 and 30 PCR cycles using the Promega PowerSeq 46GY System and sequenced on the ONT MinION device in triplicate. Basecalled reads were then processed with STRspy using a custom database containing alleles reported in the STRSeq BioProject NIST 1036 dataset. Resultant STR allele designations and flanking region single nucleotide polymorphism (SNP) calls were compared to the manufacturer-validated genotypes for each sample. STRspy generated robust and reliable genotypes across all autosomal STR loci amplified with 30 PCR cycles, achieving 100% concordance based on both length and sequence. Furthermore, we were able to identify flanking region SNPs in the 15-cycle dataset with > 90% accuracy. These results demonstrate that when analyzed with STRspy ONT reads can reveal additional variation in and around STR loci depending on read coverage. As the first and only third generation sequencing platform-specific method to successfully profile the entire panel of autosomal STRs amplified by a commercially available multiplex, STRspy significantly increases the feasibility of nanopore sequencing in forensic applications.

Multiomics Investigation of Hypertension and White Matter Hyperintensity as a Source of Vascular Dementia or a Comorbidity to Alzheimer's Disease.

BACKGROUND: Age-related comorbidity is common and significantly increases the burden for the healthcare of the elderly. Alzheimer's disease (AD) and hypertension are the two most prevalent age-related conditions and are highly comorbid. While hypertension is a risk factor for vascular dementia (VD), hypertension with AD (ADHyp+) is often characterized as probable vascular dementia. In the absence of imaging and other diagnostic tests, differentiating the two pathological states is difficult. OBJECTIVE: Our goals are to (1) identify differences in CSF-based vascular dementia profiles, if any, between individuals who have AD only (ADHyp-), and individuals with ADHyp+ using CSF levels of amyloid ß, tau and p-tau, and (2) compare genome-wide DNA profiles of ADHyp- and ADHyp+ with an unaffected control population. METHOD: Genotype and clinical data were used to compare healthy controls to AD Hyp- vs. AD Hyp+. We compared the CSF biomarkers followed by evaluating genome wide profiles in three groups, and mapped SNPs to genes based on position and lowest p-value. The significant genes were examined for co-expression and known disease networks. RESULTS: We found no differences between Aß, tau and p-tau levels between ADHyp- and ADHyp+. We found TOMM40 to be associated with ADHyp- as expected but not with ADHyp+. Interestingly, SLC9A3R2 polymorphism was associated with ADHyp+, and significant gene expression changes were observed for neighboring genes. CONCLUSION: Through this exploratory study using a novel cohort stratification design, we highlight the genetic differences in clinically similar phenotypes, indicating the utility of genetic profiling in aiding differential diagnosis of ADHyp+ and VD.

Genetically-regulated transcriptomics & copy number variation of proctitis points to altered mitochondrial and DNA repair mechanisms in individuals of European ancestry.

BACKGROUND: Proctitis is an inflammation of the rectum and may be induced by radiation treatment for cancer. The genetic heritability of developing radiotoxicity and prior role of genetic variants as being associated with side-effects of radiotherapy necessitates further investigation for underlying molecular mechanisms. In this study, we investigated gene expression regulated by genetic variants, and copy number variation in prostate cancer survivors with radiotoxicity. METHODS: We investigated proctitis as a radiotoxic endpoint in prostate cancer patients who received radiotherapy (n = 222). We analyzed the copy number variation and genetically regulated gene expression profiles of whole-blood and prostate tissue associated with proctitis. The SNP and copy number data were genotyped on Affymetrix® Genome-wide Human SNP Array 6.0. Following QC measures, the genotypes were used to obtain gene expression by leveraging GTEx, a reference dataset for gene expression association based on genotype and RNA-seq information for prostate (n = 132) and whole-blood tissue (n = 369). RESULTS: In prostate tissue, 62 genes were significantly associated with proctitis, and 98 genes in whole-blood tissue. Six genes - CABLES2, ATP6AP1L, IFIT5, ATRIP, TELO2, and PARD6G were common to both tissues. The copy number analysis identified seven regions associated with proctitis, one of which (ALG1L2) was also associated with proctitis based on transcriptomic profiles in the whole-blood tissue. The genes identified via transcriptomics and copy number variation association were further investigated for enriched pathways and gene ontology. Some of the enriched processes were DNA repair, mitochondrial apoptosis regulation, cell-to-cell signaling interaction processes for renal and urological system, and organismal injury. CONCLUSIONS: We report gene expression changes based on genetic polymorphisms. Integrating gene-network information identified these genes to relate to canonical DNA repair genes and processes. This investigation highlights genes involved in DNA repair processes and mitochondrial malfunction possibly via inflammation. Therefore, it is suggested that larger studies will provide more power to infer the extent of underlying genetic contribution for an individual's susceptibility to developing radiotoxicity.

Mitochondrial tRNA methylation in Alzheimer's disease and progressive supranuclear palsy.

BACKGROUND: Methylation of mitochondrial tRNAs (mt-tRNA) at the 9th position ("p9 site") is known to impact translational efficiency and downstream mitochondrial function; however, direct assessment of mt-RNA methylation is challenging. Recent RNA sequence-based methods have been developed to reliably identify post-transcriptional methylation. Though p9 methylation has been studied in healthy human populations and in the context of cancer, it has not yet been analyzed in neurodegenerative disease, where mitochondrial dysfunction is a prominent and early hallmark of disease progression. METHODS: Mitochondrial p9 methylation was inferred from multi-allelic calls in RNA-seq data. Gene-based association studies were performed in FUMA. Correlations between nuclear gene expression and p9 methylation were tested using Spearman's rho. Fisher's Exact test was used in PANTHER and IPA to test for overrepresentation and enrichment of biological processes and pathways in the top nuclear genes correlated with p9 methylation. RESULTS: Variable methylation was observed at 11 p9 sites in post-mortem cerebellar tissue of elderly subjects who were either healthy or diagnosed with Alzheimer's disease (AD), progressive supranuclear palsy (PSP) or pathological aging (PA). Similarities in degree of methylation were observed between AD and PSP. Certain nuclear encoded genes were identified as significantly associated with p9 methylation. Expression of 5300 nuclear encoded genes was significantly correlated with p9 methylation, with AD and PSP subjects exhibiting similar expression profiles. Overrepresentation and enrichment testing using the top transcripts revealed enrichment for a number of molecular processes, terms and pathways including many of which that were mitochondrial-related. CONCLUSION: With mitochondrial dysfunction being an established hallmark of neurodegenerative disease pathophysiology, this work sheds light on the potential molecular underpinnings of this dysfunction. Here we show overlap in cerebellar pathophysiology between common tauopathies such as Alzheimer's disease and progressive supranuclear palsy. Whether p9 hypermethylation is a cause or consequence of pathology remains an area of focus.

Differentiation of Hispanic biogeographic ancestry with 80 ancestry informative markers.

Ancestry informative single nucleotide polymorphisms (SNPs) can identify biogeographic ancestry (BGA); however, population substructure and relatively recent admixture can make differentiation difficult in heterogeneous Hispanic populations. Utilizing unrelated individuals from the Genomic Origins and Admixture in Latinos dataset (GOAL, n = 160), we designed an 80 SNP panel (Setser80) that accurately depicts BGA through STRUCTURE and PCA. We compared our Setser80 to the Seldin and Kidd panels via resampling simulations, which models data based on allele frequencies. We incorporated Admixed American 1000 Genomes populations (1000 G, n = 347), into a combined populations dataset to determine robustness. Using multinomial logistic regression (MLR), we compared the 3 panels on the combined dataset and found overall MLR classification accuracies: 93.2% Setser80, 87.9% Seldin panel, 71.4% Kidd panel. Naïve Bayesian classification had similar results on the combined dataset: 91.5% Setser80, 84.7% Seldin panel, 71.1% Kidd panel. Although Peru and Mexico were absent from panel design, we achieved high classification accuracy on the combined populations for Peru (MLR = 100%, naïve Bayes = 98%), and Mexico (MLR = 90%, naïve Bayes = 83.4%) as evidence of the portability of the Setser80. Our results indicate the Setser80 SNP panel can reliably classify BGA for individuals of presumed Hispanic origin.

Cell-free mitochondrial DNA increases in maternal circulation during healthy pregnancy: a prospective, longitudinal study.

Mitochondrial DNA (mtDNA) exposed to the extracellular space due to cell death has immunostimulatory properties. Case-control studies reported a positive association between odds of developing preeclampsia and circulating mtDNA. These findings are based on relative quantification protocols that do not allow determination of absolute concentrations of mtDNA and are highly sensitive to nuclear DNA contamination. Furthermore, circulating mtDNA concentrations in response to normal pregnancy, which is an inflammatory state characterized by continuous placental cell apoptosis, have not been established. The main objective of this study was to determine longitudinal changes in circulating mtDNA from preconception to first trimester, third trimester, and postpartum in healthy pregnant women. Absolute real-time PCR quantification of mtDNA and nuclear DNA (nDNA) was performed on whole genomic extracts from serum using TaqMan probes and chemistry. Serum cell-free mtDNA and nDNA concentrations were greater in late pregnancy as compared with early pregnancy and postpartum. Pregnant women carrying neonates at the upper quartile of birth length distribution had higher concentrations of mtDNA in late pregnancy compared with pregnancies carrying neonates at the lower quartile. The correlation between circulating mtDNA and nDNA concentrations varied by sex (i.e., pregnancies carrying female vs. male fetuses). This study is the first to establish temporal patterns of circulating cell-free mtDNA concentrations in normal human pregnancy using absolute DNA quantification techniques. Concentrations of circulating mtDNA in normal pregnancy may be used as reference values for the development of clinical prognostic or diagnostic tests in pregnant women with, or at risk of developing, gestational complications.

Two-stage Bayesian GWAS of 9576 individuals identifies SNP regions that are targeted by miRNAs inversely expressed in Alzheimer's and cancer.

INTRODUCTION: We compared genetic variants between Alzheimer's disease (AD) and two age-related cancers-breast and prostate -to identify single-nucleotide polymorphisms (SNPs) that are associated with inverse comorbidity of AD and cancer. METHODS: Bayesian multinomial regression was used to compare sex-stratified cases (AD and cancer) against controls in a two-stage study. A ±500 KB region around each replicated hit was imputed and analyzed after merging individuals from the two stages. The microRNAs (miRNAs) that target the genes involving these SNPs were analyzed for miRNA family enrichment. RESULTS: We identified 137 variants with inverse odds ratios for AD and cancer located on chromosomes 19, 4, and 5. The mapped miRNAs within the network were enriched for miR-17 and miR-515 families. DISCUSSION: The identified SNPs were rs4298154 (intergenic), within TOMM40/APOE/APOC1, MARK4, CLPTM1, and near the VDAC1/FSTL4 locus. The miRNAs identified in our network have been previously reported to have inverse expression in AD and cancer.

Genome-Wide Methylation of Mild Cognitive Impairment in Mexican Americans Highlights Genes Involved in Synaptic Transport, Alzheimer's Disease-Precursor Phenotypes, and Metabolic Morbidities.

The Mexican American population is among the fastest growing aging population and has a younger onset of cognitive decline. This group is also heavily burdened with metabolic conditions such as hypertension, diabetes, and obesity. Unfortunately, limited research has been conducted in this group. Understanding methylation alterations, which are influenced by both genetic and lifestyle factors, is key to identifying and addressing the root cause for mild cognitive impairment, a clinical precursor for dementia. We conducted an epigenome-wide association study on a community-based Mexican American population using the Illumina EPIC array. Following rigorous quality control measures, we identified 10 CpG sites to be differentially methylated between normal controls and individuals with mild cognitive impairment annotated to PKIB, KLHL29, SEPT9, OR2C3, CPLX3, BCL2L2-PABPN1, and CCNY. We found four regions to be differentially methylated in TMEM232, SLC17A8, ALOX12, and SEPT8. Functional gene-set analysis identified four gene-sets, RIN3, SPEG, CTSG, and UBE2L3, as significant. The gene ontology and pathway analyses point to neuronal cell death, metabolic dysfunction, and inflammatory processes. We found 1,450 processes to be enriched using empirical Bayes gene-set enrichment. In conclusion, the functional overlap of differentially methylated genes associated with cognitive impairment in Mexican Americans implies cross-talk between metabolically-instigated systemic inflammation and disruption of synaptic vesicular transport.