Blood mtDNA Research Articles

Serum FGF-21, GDF-15, and blood mtDNA copy number are not biomarkers of Parkinson disease.

Strong evidence of mitochondrial dysfunction exists for both familial and sporadic Parkinson disease (PD). A simple test, reliably identifying mitochondrial dysfunction, could be important for future stratified medicine trials in PD. We previously undertook a comparison of serum biomarkers in classic mitochondrial diseases and established that serum growth differentiation factor 15 (GDF-15) outperforms fibroblast growth factor 21 (FGF-21) when distinguishing patients with mitochondrial diseases from healthy controls. This study aimed to systematically assess serum FGF-21 and GDF-15, together with mitochondrial DNA (mtDNA) copy number levels in peripheral blood cells from patients with PD and healthy controls, to determine whether these measures could act as a biomarker of PD. One hundred twenty-one patients with PD and 103 age-matched healthy controls were recruited from a single center. Serum FGF-21 and GDF-15, along with blood mtDNA copy number, were quantified using established assays. There were no meaningful differences identified for any of the measures when comparing patients with PD with healthy controls. This highlights a lack of diagnostic sensitivity that is incompatible with these measures being used as biomarkers for PD. In this study, serum FGF-21, serum GDF-15, and blood mtDNA levels were similar in patients with PD and healthy controls and therefore unlikely to be satisfactory indicators of mitochondrial dysfunction in patients with PD. This study provides Class III evidence that serum FGF-21, serum GDF-15, and blood mtDNA copy number levels do not distinguish patients with PD from healthy controls. There was no diagnostic uncertainty between patients with PD and healthy controls.

Mitochondrial DNA somatic mutation burden and heteroplasmy are associated with chronological age, smoking, and HIV infection.

The gradual accumulation of mitochondrial DNA (mtDNA) mutations is implicated in aging and may contribute to the accelerated aging phenotype seen with tobacco smoking and HIV infection. mtDNA mutations are thought to arise from oxidative damage; however, recent reports implicate polymerase γ errors during mtDNA replication. Investigations of somatic mtDNA mutations have been hampered by technical challenges in measuring low‐frequency mutations. We use primer ID‐based next‐generation sequencing to quantify both somatic and heteroplasmic blood mtDNA point mutations within the D‐loop, in 164 women and girls aged 2–72 years, of whom 35% were smokers and 56% were HIV‐positive. Somatic mutations and the occurrence of heteroplasmic mutations increased with age. While transitions are theorized to result from polymerase γ errors, transversions are believed to arise from DNA oxidative damage. In our study, both transition and transversion mutations were associated with age. However, transition somatic mutations were more prevalent than transversions, and no heteroplasmic transversions were observed. We also measured elevated somatic mutations, but not heteroplasmy, in association with high peak HIV viremia. Conversely, heteroplasmy was higher among smokers, but somatic mutations were not, suggesting that smoking promotes the expansion of preexisting mutations rather than de novo mutations. Taken together, our results are consistent with blood mtDNA mutations increasing with age, inferring a greater contribution of polymerase γ errors in mtDNA mutagenesis. We further suggest that smoking and HIV infection both contribute to the accumulation of mtDNA mutations, though in different ways.

Quantification of peripheral whole blood, cell-free plasma and exosome encapsulated mitochondrial DNA copy numbers in patients with atrial fibrillation

Mitochondrial DNA (mtDNA) copy number changes have been associated with various diseases. Several studies showed that mtDNA content in peripheral blood was associated with oxidative stress and cardiovascular disease. Atrial fibrillation (AF) is one of the severe cardiovascular diseases. We aimed to determine the mtDNA copy numbers in peripheral blood, in cell-free plasma and in exosomes of AF patients and healthy controls.Peripheral blood was drawn from 60 AF patients and 72 healthy controls. DNA was isolated from EDTA blood and plasma. Exosomes were isolated from cell-free plasma and then exosome encapsulated DNA (exoDNA) was extracted. Quantitative-real-time PCR was performed with Human Mitochondrial DNA (mtDNA) Monitoring Primer Set. Statistical analysis of the data was performed.We found statistically significant difference in mtDNA copy numbers in DNA isolated from peripheral whole blood, cell-free plasma and exosome samples of controls’ (44.4 ± 18.0, 27.2 ± 30.1, 11.5 ± 8.7), and patients’ group (43.4 ± 13.6, 26.2 ± 26.4, 14.5 ± 12.3). However there was no significant difference in mtDNA copy number between the two study groups either in peripheral blood, in cell-free plasma and in exosomes, and even in different sexes and ages.We found the highest copy number of mtDNA in peripheral blood, followed by plasma and exosomes. We did not find differences between patients and controls, neither age nor gender had effect on the mtDNA copy number. According to our results the mtDNA copy numbers did not differ in AF patients.

The Cord Blood Insulin and Mitochondrial DNA Content Related Methylome.

Mitochondrial dysfunction seems to play a key role in the etiology of insulin resistance. At birth, a link has already been established between mitochondrial DNA (mtDNA) content and insulin levels in cord blood. In this study, we explore shared epigenetic mechanisms of the association between mtDNA content and insulin levels, supporting the developmental origins of this link. First, the association between cord blood insulin and mtDNA content in 882 newborns of the ENVIRONAGE birth cohort was assessed. Cord blood mtDNA content was established via qPCR, while cord blood levels of insulin were determined using electrochemiluminescence immunoassays. Then the cord blood DNA methylome and transcriptome were determined in 179 newborns, using the human 450K methylation Illumina and Agilent Whole Human Genome 8 × 60 K microarrays, respectively. Subsequently, we performed an epigenome-wide association study (EWAS) adjusted for different maternal and neonatal variables. Afterward, we focused on the 20 strongest associations based on p-values to assign transcriptomic correlates and allocate corresponding pathways employing the R packages ReactomePA and RDAVIDWebService. On the regional level, we examined differential methylation using the DMRcate and Bumphunter packages in R. Cord blood mtDNA content and insulin were significantly correlated (r = 0.074, p = 0.028), still showing a trend after additional adjustment for maternal and neonatal variables (p = 0.062). We found an overlap of 33 pathways which were in common between the association with cord blood mtDNA content and insulin levels, including pathways of neurodevelopment, histone modification, cytochromes P450 (CYP)-metabolism, and biological aging. We further identified a DMR annotated to Repulsive Guidance Molecule BMP Co-Receptor A (RGMA) linked to cord blood insulin as well as mtDNA content. Metabolic variation in early life represented by neonatal insulin levels and mtDNA content might reflect or accommodate alterations in neurodevelopment, histone modification, CYP-metabolism, and aging, indicating etiological origins in epigenetic programming. Variation in metabolic hormones at birth, reflected by molecular changes, might via these alterations predispose children to metabolic diseases later in life. The results of this study may provide important markers for following targeted studies.

Peripheral Blood Mitochondrial DNA Copy Number Obtained From Genome-Wide Genotype Data Is Associated With Neurocognitive Impairment in Persons With Chronic HIV Infection.

Mitochondrial DNA (mtDNA) copy number varies by cell type and energy demands. Blood mtDNA copy number has been associated with neurocognitive function in persons without HIV. Low mtDNA copy number may indicate disordered mtDNA replication; high copy number may reflect a response to mitochondrial dysfunction. We hypothesized that blood mtDNA copy number estimated from genome-wide genotyping data is related to neurocognitive impairment (NCI) in persons with HIV. In the CNS HIV Antiretroviral Therapy Effects Research (CHARTER) study, peripheral blood mtDNA copy number was obtained from genome-wide genotyping data as a ratio of mtDNA single-nucleotide polymorphism probe intensities relative to nuclear DNA single-nucleotide polymorphisms. In a multivariable regression model, associations between mtDNA copy number and demographics, blood cell counts, and HIV disease and treatment characteristics were tested. Associations of mtDNA copy number with the global deficit score (GDS), GDS-defined NCI (GDS ≥ 0.5), and HIV-associated neurocognitive disorder (HAND) diagnosis were tested by logistic regression, adjusting for potential confounders. Among 1010 CHARTER participants, lower mtDNA copy number was associated with longer antiretroviral therapy duration (P < 0.001), but not with d-drug exposure (P = 0.85). mtDNA copy number was also associated with GDS (P = 0.007), GDS-defined NCI (P < 0.001), and HAND (P = 0.002). In all analyses, higher mtDNA copy number was associated with poorer cognitive performance. Higher mtDNA copy number estimated from peripheral blood genotyping was associated with worse neurocognitive performance in adults with HIV. These results suggest a connection between peripheral blood mtDNA and NCI, and may represent increased mtDNA replication in response to mitochondrial dysfunction.

Prenatal manganese and cord blood mitochondrial DNA copy number: Effect modification by maternal anemic status

IntroductionManganese (Mn) is an essential nutrient but also a toxicant at high exposures, when it can induce oxidative stress (OS). Mn uptake is inversely correlated with iron status, therefore anemic individuals may be more susceptible to Mn overload induced-OS, which can manifest as changes in mitochondrial DNA copy number (mtDNA CN). Our objectives were to: 1) determine stage-specific associations of prenatal Mn exposure with cord blood MtDNA CN; and 2) investigate effect modification by maternal anemia, ferritin, and mean corpuscular volume (MCV). Materials and methodsWe measured whole blood Mn, hemoglobin, serum ferritin, and MCV in the 2nd and 3rd trimester, in maternal blood at birth, and in cord blood from a prospective birth cohort in Mexico City, Mexico (n = 485). We then extracted DNA from cord blood leukocytes to determine mtDNA CN. We used robust regression to measure associations between Mn and mtDNA CN at each trimester and at birth. Anemia (hemoglobin ≤11 g/dL), iron deficiency (ferritin ≤15 ng/mL) and MCV (stratified at median), were examined as effect modifiers. ResultsMn levels increased throughout pregnancy, and Mn was inversely correlated with ferritin. We observed a positive association between Mn in the 3rd trimester and Mn in cord blood and mtDNA CN (β = 0.04–0.05; 95% CI = 0.01, 0.08). Anemia significantly modified the association between mtDNA CN and Mn in the 2nd trimester. We found a positive association between 2nd trimester Mn and mtDNA CN in mothers with normal hemoglobin, and a negative association in those with low hemoglobin. (βhigh = 0.06; 95% CI = 0.01, 0.11; p = 0.01 and βlow = −0.06; 95% CI = 0.03, −0.13; p = 0.06). No associations were detected between anemia, iron deficiency and MCV and mtDNA CN. ConclusionsMaternal blood Mn in the 3rd trimester and in cord blood was positively associated with mtDNA CN, suggesting that higher late pregnancy prenatal Mn exposures can impact newborn mitochondria by promoting OS. Furthermore, 2nd trimester Mn was positively associated with mtDNA in non-anemic mother-child pairs but inversely associated in anemic individuals, indicating potential interactions between Mn and chronic anemia.

Altered cord blood mitochondrial DNA content and pregnancy lead exposure in the PROGRESS cohort

IntroductionLead (Pb) crosses the placenta and can cause oxidative stress, reduced fetal growth and neurological problems. The principal source of oxidative stress in human cells is mitochondria. Therefore, disruption of normal mitochondrial function during pregnancy may represent a primary mechanism behind the adverse effects of lead. We sought to assess the association of Pb exposure during pregnancy with mitochondrial DNA (mtDNA) content, a sensitive marker of mitochondrial function, in cord blood. Materials and methodsThis study comprised mother-infant pairs from the Programming Research in Obesity, Growth, Environment and Social Stressors (PROGRESS) study, a prospective birth-cohort that enrolled 1050 pregnant women from Mexico City who were receiving prenatal care between December 2007 and July 2011. Quantitative PCR was used to calculate relative MtDNA content (mitochondrial-to-nuclear DNA ratio (mtDNA/nDNA)) in cord blood. Lead concentrations in both maternal blood (2nd and 3rd trimester and at delivery day) and in cord blood were measured by ICP-MS. Multivariable regression models adjusting for multiple confounders were fitted with 410 mother-infant pairs for whom complete data for mtDNA content, lead levels, and covariates were available. ResultsMaternal blood Pb measured in the second (mean 3.79 μg/dL, SD 2.63; β = 0.059, 95% CI 0.008, 0.111) and third trimester (mean 3.90 μg/dL; SD 2.84; β = 0.054, 95% CI 0.002, 0.107) during pregnancy and PB in cord blood (mean 3.50 μg/dL, SD 2.59; β = 0.050, 95% CI 0.004; 0.096) were associated with increased cord blood mtDNA content (mean 1.46, SD 0.44). In two-way interaction analyses, cord blood Pb marginally interacted with gestational age leading to an increase in mtDNA content for pre-term births (Benjamini-Hochberg False Discovery Rate correction; BH-FDR = 0.08). ConclusionThis study shows that lead exposure in pregnancy alters mtDNA content in cord blood; therefore, alteration of mtDNA content might be a mechanism underlying the toxicity of lead.

A descriptive pilot study of mitochondrial mutations & clinical phenotype in fibromyalgia syndrome.

Background & objectives:Fibromyalgia syndrome (FMS) is one of the most common chronic pain conditions of unknown aetiology. Mitochondrial dysfunction has been reported in FMS with some studies reporting the presence of mitochondrial mutation namely A3243G, which also causes mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes. This pilot study was conducted to assess this mutation and also detect large deletions in mitochondrial DNA (mtDNA) in patients with FMS.Methods:Thirty female patients with FMS participated and 30 matched controls were included. Genomic DNA was subjected to polymerase chain reaction (PCR) amplification using specific primers followed by restriction digestion with ApaI enzyme to detect the specific A3243G mtDNA mutation. Long-range PCR was done in two sets to detect the large deletions in the mtDNA. Biochemical parameters including thyroid-stimulating hormone and vitamin D levels were also looked at.Results:None of the patients were found to carry the common mutation or large deletions. Low vitamin D level was a common finding. Hypothyroidism was found in a few patients.Interpretation & conclusions:Although the common mutation or large mtDNA deletions were not detected in blood mtDNA in the FMS patients, mutations in the muscle and sequence variation in mtDNA remained a possibility. Future studies in both blood and muscle tissue including mtDNA sequencing are warranted in such patients to determine if a subset of FMS patients have mitochondrial myopathy.

Peripheral blood mitochondrial DNA copy number is a potential biomarker for progress of diabetic nephropathy in type 2 diabetes patients

Mitochondrial DNA (mtDNA) copy number is a surrogate measure of mitochondrial function, and altered mtDNA copy number has been associated with diabetic nephropathy (DN), a leading cause end stage renal disease in diabetic patients. We investigated whether mtDNA copy number in peripheral blood could be utilized as a biomarker for DN progress in type 2 diabetes (T2D) patients. A total of 50 non-diabetic and 100 diabetic subjects were recruited. The diabetic subjects were sub-divided based on urinary albumin-creatinine ratio (ACR) into T2D patients with normoalbuminuria (ACR < 30 mg/g, n = 50), DN patients with microalbuminuria (ACR 30–300 mg/g, n = 29), and DN patients with macroalbuminuria (ACR > 300 mg/g, n = 21). The mtDNA copy number was measured as relative quantity of mtDNA compared to nuclear DNA using real-time PCR. Overall, the mtDNA copy number was lower in T2D and DN patients than in controls (P < 0.001). Sub-group analyses by ACR showed that lower mtDNA copy number was significantly associated with high levels of albuminuria even after adjusting for conventional risk factors (odds ratio [OR] 0.47, confidence interval [CI] 0.299–0.74 for microalbuminuria and OR 0.21, 95% CI 0.1–0.436 for macroalbuminuria). Moreover, mtDNA copy number was correlated negatively with albuminuria (r = −0.546) and positively with glomerular filtration rate (r = 0.572). The areas under the receiver operating characteristic curves (AUCs) of mtDNA copy number in discriminating DN patients from T2D patients and healthy controls were 0.80 (P < 0.001) and 0.885 (P < 0.001), respectively. Our results showed that peripheral blood mtDNA copy number was associated with the development of DN and could serve as a biomarker during DN progression.

Mitochondrial DNA copy number is associated with psychosis severity and anti-psychotic treatment

Mitochondrial pathology has been implicated in the pathogenesis of psychotic disorders. A few studies have proposed reduced leukocyte mitochondrial DNA (mtDNA) copy number in schizophrenia and bipolar disorder type I, compared to healthy controls. However, it is unknown if mtDNA copy number alteration is driven by psychosis, comorbidity or treatment. Whole blood mtDNA copy number was determined in 594 psychosis patients and corrected for platelet to leukocyte count ratio (mtDNAcnres). The dependence of mtDNAcnres on clinical profile, metabolic comorbidity and antipsychotic drug exposure was assessed. mtDNAcnres was reduced with age (β = −0.210, p < 0.001), use of clozapine (β = −0.110,p = 0.012) and risperidone (β = −0.109,p = 0.014), dependent on prescribed dosage (p = 0.006 and p = 0.026, respectively), and the proportion of life on treatment (p = 0.006). Clozapine (p = 0.0005) and risperidone (p = 0.0126) had a reducing effect on the mtDNA copy number also in stem cell-derived human neurons in vitro at therapeutic plasma levels. For patients not on these drugs, psychosis severity had an effect (β = −0.129, p = 0.017), similar to age (β = −0.159, p = 0.003) and LDL (β = −0.119, p = 0.029) on whole blood mtDNAcnres. Further research is required to determine if mtDNAcnres reflects any psychosis-intrinsic mitochondrial changes.

Cord blood leptin and insulin levels in association with mitochondrial DNA content

BackgroundThe developmental origins of health and disease theory states that a disturbance in the early life environment can contribute to disease risk in later life. Leptin and insulin are anorectic hormones involved in energy homeostasis and are crucial for foetal growth. Disturbances in the levels of these hormones contribute to obesity and diabetes. In adults, altered mitochondrial function is an important hallmark of metabolic disorders, including obesity and diabetes. However, the mitochondrial effects of early life metabolic variation are unexplored. We investigated whether there is an association between metabolic hormones and mitochondrial DNA (mtDNA) content in early life.MethodsThe study included 236 newborns from the FLEHS III birth cohort, Flanders (Belgium). Relative mtDNA content of cord blood leukocytes was determined using quantitative PCR. Cord blood levels of leptin and insulin were determined using immunoassays. We studied the association between these metabolic hormones and mtDNA content using multiple linear regression models, while accounting for covariates and potential confounders.ResultsLeptin and insulin levels were positively associated with cord blood mtDNA content. mtDNA content was respectively 4.49% (95% CI 1.15–7.93; p = 0.008) and 1.60% (95% CI 0.31–2.91; p = 0.02) higher for a interquartile range increase of respectively cord blood leptin and insulin levels. In a sensitivity analysis, we observed that insulin and leptin were independently associated to mtDNA content and that insulin was stronger associated to mtDNA content in boys than in girls.ConclusionNeonatal metabolic hormones were associated with cord blood mtDNA content, which suggests that in early life the variation of mtDNA content might accommodate or reflect changes in the metabolic status.

Associations of blood mitochondrial DNA copy number with social-demographics and cancer risk: results from the Mano-A-Mano Mexican American Cohort.

The relationship between blood mitochondrial DNA (mtDNA) copy number and subsequent cancer risk has been investigated previously. However, such association has never been examined in Mexican Americans. In the current study, we examined association between social-demographic factors and blood mtDNA copy number, as well as longitudinal relationship between cancer and mtDNA copy number, among 10,802 Mexican Americans in the Mano-A-Mano Mexican American Cohort. Overall, mtDNA copy number was statistically significantly higher among participants who developed cancer during the study period than among cancer-free controls (0.17 vs 0.13, P = 0.007). Among cancer-free control participants, mtDNA copy number significantly differed by social-demographic characteristics. However, there was a large degree of heterogeneity in these effects across the mtDNA copy number distribution. In the longitudinal analysis, we observed that higher mtDNA copy number was positively associated with increased risk of all cancer types (adjusted hazard ratio [HR], 1.13; 95% confidence interval [CI], 1.09–1.17). Participants with mtDNA copy number in the fourth (highest) quartile had a higher risk of all cancer (adjusted HR, 2.12; 95% CI, 1.65–2.73) than did participants in the first (lowest) quartile. In summary, our results in Mexican Americans support an association between increased mtDNA copy number and cancer risk. Our results also suggest that mtDNA copy number may be influenced by social and demographic factors.

Elevated Cell-Free Mitochondrial DNA in Filtered Plasma Is Associated With HIV Infection and Inflammation

Increased cell-free DNA levels are associated with poor health outcomes, and cell-free mitochondrial DNA (cf-mtDNA) has proinflammatory properties. Given that HIV infection is associated with chronic inflammation, we investigated the relationship between cf-mtDNA and proinflammatory cytokine interleukin-6 (IL-6) in the context of HIV infection. We also optimized separation of cell-free plasma from blood. In this retrospective cross-sectional study, we collected blood, demographic information, and clinical data from 99 HIV-infected and 103 HIV-uninfected adults and children enrolled in the Children and Women: AntiRetrovirals and Markers of Aging pan-Canadian (CARMA) cohort. Plasma was separated from blood by 14,000g centrifugation followed by 0.45-μm filtration to remove cells and platelets. Cf-mtDNA and cell-free nuclear DNA were quantified simultaneously via monochrome, multiplex, quantitative polymerase chain reaction. IL-6 was measured using enzyme-linked immunosorbent assay. Higher speed centrifugation and filtration was necessary to isolate truly cell-free plasma. Higher cf-mtDNA levels were univariately associated with HIV infection, elevated IL-6 levels, younger age, higher white blood cell count, and higher cell-free nuclear DNA levels but not blood mtDNA content or HIV viral load. In a multivariable model, HIV infection (P < 0.001), elevated IL-6 (P = 0.021), younger age (P < 0.001), and higher blood nDNA levels (P = 0.007) were independently associated with higher cf-mtDNA. People living with HIV have higher levels of circulating cf-mtDNA than their uninfected peers. Increased levels of inflammatory marker IL-6 are associated with elevated cf-mtDNA, independent of the effect of HIV infection. Higher cf-mtDNA levels and white blood cell count in younger people may reflect higher cell turnover in that population.

Peripheral blood mitochondrial DNA copy number, a potential marker of non‐alcoholic fatty liver disease?

IntroductionNonalcoholic fatty liver disease (NAFLD) includes a spectrum of histological findings ranging from simple steatosis (SS) to non‐alcoholic steatohepatitis (NASH). Although often considered the hepatic manifestation of metabolic syndrome (MS), longitudinal studies suggest that NAFLD precedes MS. To date, liver biopsy is the most accurate method to differentiate between SS and NASH. However, recent studies indicate an association between mitochondrial DNA (mtDNA) copy number and NAFLD severity in liver biopsies. If a similar association exists in peripheral blood, mtDNA copy number could represent a non‐invasive alternative for differentiating NAFLD disease stages.Given the above, we analyzed whether mtDNA copy number differs in individuals discordant for NAFLD and MS. We also probed for correlations between liver and peripheral blood mtDNA copy number, in addition to associations between NAFLD and mtDNA copy number.ObjectiveTo analyze whether mtDNA copy number is a useful blood‐based marker of NAFLD severity.Materials and methodsCohort = 73 peripheral blood samples and liver biopsies obtained from patients that were operated for gallstones; individuals were separated into 4 groups: controls (n = 27), NAFLD (n = 14), MS (n = 9) and MS + NAFLD (n = 23). NAFLD severity was determined according to the parameters described by Kleiner; mtDNA copy number was determined by semiquantitative PCR.ResultsWe found no significant differences in mtDNA copy number across the four groups analyzed (i.e. control, NAFLD, MS, MS+NAFLD). However, we did find an association between mtDNA copy number and NAFLD severity. In particular, we found that a significant increase in mtDNA copy number in SS relative to control samples, while mtDNA copy number was significantly reduced in NASH relative to SS.ConclusionAkin to the previously published data of mtDNA copy number in liver biopsies of individuals with NAFLD, we uncovered a dynamic relationship between mtDNA copy number and NAFLD severity in peripheral blood samples. This suggests that mtDNA copy number may be a promising blood‐based biomarker for differentiating between SS and NASH. To address this issue, we are currently probing for correlations between liver and peripheral blood mtDNA copy number.Support or Funding InformationMexican National Council for Science and Technology (CONACyT).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Association of mitochondrial DNA content in peripheral blood with cancer-related fatigue and chemotherapy-related cognitive impairment in early-stage breast cancer patients: a prospective cohort study.

Cancer-related fatigue (CRF) and chemotherapy-related cognitive impairment (CRCI) are reported to be associated with mitochondrial dysfunction. Hence, mitochondrial DNA (mtDNA) content, a biomarker of mitochondrial dysfunction, is hypothesized to correlate with the onset of CRF and CRCI. This study aims to evaluate the association between peripheral blood mtDNA content reduction and severity of CRF and CRCI in patients receiving chemotherapy. This was a prospective cohort study. Early-stage breast cancer patients receiving anthracycline- or taxane-based chemotherapy were recruited. CRF was assessed using MFSI-SF, and CRCI was assessed using FACT-Cog and CANTAB at two timepoints: baseline (T1; prior to treatment) and 6weeks after initiation of treatment (T2). mtDNA content was measured at both timepoints using real-time quantitative polymerase chain reaction. Multiple logistic regression was utilized to evaluate the association between mtDNA reduction and worsening of CRF and CRCI, adjusting for age, anxiety, insomnia, plasma cytokines concentrations, and other clinically important covariates. A total of 108 patients (age 52.0±9.2years; 82.4% Chinese; 64.8% receiving anthracycline-based chemotherapy) were recruited. Proportions of patients with worsening of CRF increased from the lower to the upper quartiles of mtDNA reduction (22.2, 33.3, 55.6, and 63.0% in quartiles 1, 2, 3, and 4, respectively,p=0.001 for trend). Reduction of mtDNA content was significantly greater among those with worsening of CRF and CRCI compared to those without CRF [mean reduction (±SD): 36.5 (46.1) vs. 9.4 (34.5), p<0.001]. After adjusting for covariates, every 1-unit reduction of the mtDNA content was associated with a 4% increased risk for worsening of CRF (95% CI, 1-6%; p=0.009). This is the first study to show that the reduction of mtDNA content in peripheral blood is associated with the onset of CRF in patients receiving chemotherapy. Further validation studies are required to confirm the findings.

Lower mitochondrial DNA and altered mitochondrial fuel metabolism in HIV-exposed uninfected infants in Cameroon.

Evaluate blood mitochondrial DNA (mtDNA) content in HIV/antiretroviral-exposed uninfected (HEU) vs. HIV-unexposed uninfected (HUU) infants and investigate differences in mitochondrial-related metabolites by exposure group. We enrolled a prospective cohort of HIV-infected and HIV-uninfected pregnant woman/infant pairs in Cameroon. Dried blood spot mtDNA : nuclear DNA ratio was measured by monochrome multiplex quantitative polymerase chain reaction in HEU infants exposed to in-utero antiretrovirals and postnatal zidovudine (HEU-Z) or nevirapine (HEU-N), and in HUU infants at 6 weeks of life. Acylcarnitines and branch-chain amino acids (BCAAs) were measured via tandem mass spectrometry and consolidated into seven uncorrelated components using principal component analysis. Linear regression models were fit to assess the association between in-utero/postnatal HIV/antiretroviral exposure and infant mtDNA, adjusting for confounders and principal component analysis-derived acylcarnitine/BCAA component scores. Of 364 singleton infants, 38 were HEU-Z, 117 HEU-N, and 209 HUU. Mean mtDNA content was lowest in HEU-Z infants (140 vs. 160 in HEU-N vs. 174 in HUU, P = 0.004). After adjusting for confounders, HEU-Z infants remained at increased risk for lower mtDNA content compared with HUU infants (β: -4.46, P = 0.045), whereas HEU-N infants did not, compared with HUU infants (β: -1.68, P = 0.269. Furthermore, long-chain acylcarnitines were associated with lower (β: -2.35, P = 0.002) and short-chain and BCAA-related acylcarnitines were associated with higher (β: 2.96, P = 0.001) mtDNA content. Compared with HUU infants, HEU infants receiving postnatal zidovudine appear to be at increased risk for decreased blood mtDNA content which may be associated with altered mitochondrial fuel utilization in HEU-Z infants.

Independent impacts of aging on mitochondrial DNA quantity and quality in humans

BackgroundThe accumulation of mitochondrial DNA (mtDNA) mutations, and the reduction of mtDNA copy number, both disrupt mitochondrial energetics, and may contribute to aging and age-associated phenotypes. However, there are few genetic and epidemiological studies on the spectra of blood mtDNA heteroplasmies, and the distribution of mtDNA copy numbers in different age groups and their impact on age-related phenotypes. In this work, we used whole-genome sequencing data of isolated peripheral blood mononuclear cells (PBMCs) from the UK10K project to investigate in parallel mtDNA heteroplasmy and copy number in 1511 women, between 17 and 85 years old, recruited in the TwinsUK cohorts.ResultsWe report a high prevalence of pathogenic mtDNA heteroplasmies in this population. We also find an increase in mtDNA heteroplasmies with age (β = 0.011, P = 5.77e-6), and showed that, on average, individuals aged 70-years or older had 58.5% more mtDNA heteroplasmies than those under 40-years old. Conversely, mtDNA copy number decreased by an average of 0.4 copies per year (β = −0.395, P = 0.0097). Multiple regression analyses also showed that age had independent effects on mtDNA copy number decrease and heteroplasmy accumulation. Finally, mtDNA copy number was positively associated with serum bicarbonate level (P = 4.46e-5), and inversely correlated with white blood cell count (P = 0.0006). Moreover, the aggregated heteroplasmy load was associated with blood apolipoprotein B level (P = 1.33e-5), linking the accumulation of mtDNA mutations to age-related physiological markers.ConclusionsOur population-based study indicates that both mtDNA quality and quantity are influenced by age. An open question for the future is whether interventions that would contribute to maintain optimal mtDNA copy number and prevent the expansion of heteroplasmy could promote healthy aging.

Associations of Annual Ambient Fine Particulate Matter Mass and Components with Mitochondrial DNA Abundance.

Fine particulate matter (PM2.5) represents a mixture of components with potentially different toxicities. However, little is known about the relative effects of PM2.5 mass and PM2.5 components on mitochondrial DNA (mtDNA) abundance, which may lie on the pathway of PM2.5-associated disease. We studied 646 elderly male participants in the Normative Aging Study from Greater Boston to investigate associations of long-term exposure to PM2.5 mass and PM2.5 components with mtDNA abundance. We estimated concentrations of pollutants for the 365-day preceding examination at each participant's address using spatial- and temporal-resolved chemical transport models. We measured blood mtDNA abundance using RT-PCR. We applied a shrinkage and selection method (adaptive LASSO) to identify components most predictive of mtDNA abundance, and fit multipollutant linear mixed-effects models with subject-specific intercept to estimate the relative effects of individual PM component. MtDNA abundance was negatively associated with PM2.5 mass in the previous year and-after adjusting for PM2.5 mass-several PM2.5 components, including organic carbon, sulfate (marginally), and nitrate. In multipollutant models including as independent variables PM2.5 mass and PM2.5 components selected by LASSO, nitrate was associated with mtDNA abundance. An SD increase in annual PM2.5-associated nitrate was associated with a 0.12 SD (95% confidence intervals [CI] = -0.18, -0.07) decrease in mtDNA abundance. Analyses restricted to PM2.5 annual concentration below the current 1-year U.S. Environmental Protection Agency standard produced similar results. Long-term exposures to PM2.5-associated nitrate were related to decreased mtDNA abundance independent of PM2.5 mass. Mass alone may not fully capture the potential of PM2.5 to oxidize the mitochondrial genome.See video abstract at, http://links.lww.com/EDE/B274.

Correlation Between Mitochondrial DNA Content Measured in Myocardium and Peripheral Blood of Patients with Non-Ischemic Heart Failure.

Heart failure (HF) is associated with disturbances in mitochondrial energy production. This mitochondrial dysfunction is reflected by depletion of mitochondrial DNA (mtDNA) in different tissues. Our aims were to determine if there was a correlation between mtDNA content measured in myocardial tissue and the easily accessible peripheral blood cells of patients with non-ischemic HF; and to determine if there was a correlation between myocardial mtDNA and left ventricular (LV) ejection fraction. We prospectively collected paired myocardial tissue and peripheral blood samples from 13 consecutive end-stage non-ischemic HF patients undergoing cardiac transplantation. mtDNA content was assessed with real-time quantitative PCR by calculating the relative ratio of two specific mitochondrial sequences and one nuclear control gene sequence. HF patients with lower myocardial mtDNA content had a significantly lower LV ejection fraction (r = 0.65, p = 0.016). Peripheral blood mtDNA content correlated positively with right ventricular myocardial mtDNA content (r = 0.63, p = 0.021). We also observed that averaged myocardial DNA content tended to correlate with peripheral blood mtDNA content (r = 0.53, p = 0.061). In non-ischemic HF patients, myocardial mtDNA content is positively correlated with peripheral blood mtDNA content and LV function as assessed by echocardiography.

Quantification of mitochondrial DNA copy number in suspected cancer patients by a well optimized ddPCR method

Changes in mitochondrial DNA (mtDNA) content is a useful clinical biomarker for various diseases, however results are controversial as several analytical factors can affect measurement of mtDNA. MtDNA is often quantified by taking ratio between a target mitochondrial gene and a reference nuclear gene (mtDNA/nDNA) using quantitative real time PCR often on two separate experiments. It measures relative levels by using external calibrator which may not be comparable across laboratories. We have developed and optimized a droplet digital PCR (ddPCR) based method for quantification of absolute copy number of both mtDNA and nDNA gene in whole blood. Finally, the role of mtDNA in suspected cancer patients referred to a cancer diagnostic center was investigated.Analytical factors which can result in false quantification of mtDNA have been optimized and both target and reference have been quantified simultaneously with intra- and inter-assay coefficient variances as 3.1% and 4.2% respectively. Quantification of mtDNA show that compared to controls, solid tumors (but not hematologic malignancies) and other diseases had significantly lower copy number of mtDNA. Higher mtDNA (highest quartile) was associated with a significantly lower risk of both solid tumors and other diseases, independent of age and sex. Receiver operating curve demonstrated that mtDNA levels could differentiate controls from patients with solid tumors and other diseases.Quantification of mtDNA by a well optimized ddPCR method showed that its depletion may be a hallmark of general illness and can be used to stratify healthy individuals from patients diagnosed with cancer and other chronic diseases.