Longitudinal Assessment of Bone Mineral Density in Women Living With and Without HIV Across Reproductive Phases.

BACKGROUND: Women living with HIV commonly experience low areal bone mineral density (BMD), but whether this is affected by low ovarian hormonal states (prolonged amenorrhea or menopause) is unknown. We compared rates of BMD loss between women living with HIV and HIV-negative control women and investigated its association with low ovarian hormonal states. SETTING: Women living with HIV were enrolled from Vancouver Canada and controls from 9 Canadian sites. METHODS: This longitudinal analysis included age-matched women living with HIV in the Children and Women: AntiRetrovirals and Markers of Aging cohort and controls in the population-based Canadian Multicentre Osteoporosis Study. Rate of change/year in BMD at the total hip and lumbar spine (L1-L4) between 3 and 5 years was compared between groups, adjusting for sociodemographic and clinical variables. RESULTS: Ninety-two women living with HIV (median [interquartile range] age: 49.5 [41.6-54.1] years and body mass index: 24.1 [20.7-30.8] kg/m 2 ) and 278 controls (age: 49.0 [43.0-55.0] years and body mass index: 25.8 [22.9-30.6] kg/m 2 ) were included. Total hip BMD loss was associated with HIV (ß: -0.003 [95% CI: -0.006 to -0.0001] g/cm 2 /yr), menopause (ß: -0.007 [-0.01 to -0.005] g/cm 2 /yr), and smoking (ß: -0.003 [-0.006 to -0.0002] g/cm 2 /yr); BMD gain was linked with higher body mass index (ß: 0.0002 [0.0007-0.0004] g/cm 2 /yr). Menopause was associated with losing L1-L4 BMD (ß: -0.01 [-0.01 to -0.006] g/cm 2 /yr). Amenorrhea was not associated with BMD loss. CONCLUSIONS: HIV and menopause negatively influenced total hip BMD. These data suggest women living with HIV require hip BMD monitoring as they age.

Robust tobacco smoking self-report in two cohorts: pregnant women or men and women living with or without HIV.

Understanding the true burden of tobacco smoking on adverse pregnancy outcomes is critical in generating appropriate interventions to improve outcomes. Self-reporting of human behaviour that is associated with stigma is associated with underreporting in general and may bias the impact of smoking in studies; however, self-reporting is frequently the most practical method of gleaning this information. The objective of this study was to evaluate concordance between self-reported smoking and concentrations of plasma cotinine, a biomarker of smoking, among participants enrolled in two related HIV cohorts. A total of 100 pregnant women (76 living with HIV [LWH] and 24 negative controls) in their third trimester, and 100 men and non-pregnant women (43 LWH and 57 negative controls) were included. Among all participants, 43 pregnant women (49% LWH and 25% negative controls) and 50 men and non-pregnant women (58% LWH and 44% negative controls) were self-reported smokers. The odds of discordance between self-reported smoking and cotinine levels were not significantly different between self-reported smokers and non-smokers, nor between pregnant women and others, but were significantly increased, regardless of self-reported status, among people LWH compared to negative controls. The overall concordance between plasma cotinine and self-reported data among all participants was 94% with a sensitivity and specificity of 90% and 96%, respectively. Taken together, these data demonstrate that participant surveying in a non-judgemental context can lead to accurate and robust self-report smoking data among both persons LWH and not, including in the context of pregnancy.

Vitamin D intakes among women living with and without HIV in Canada.

BACKGROUND: Patterns of vitamin D intake are relatively unexplored among women living with HIV, despite its importance for women's health. We compared vitamin D dietary and supplement intakes in women with HIV and population-based national controls and investigated barriers to intake. METHODS: In this case-control study, women with HIV in the Children and Women: AntiRetrovirals and Markers of Aging (CARMA) cohort were matched with Canadian Multicentre Osteoporosis Study (CaMos) controls. Participants were queried for vitamin D in dairy consumption, supplementation/dosage, and sociodemographic variables. We assessed barriers to supplementation and factors associated with dietary intake by regression modelling. RESULTS: Ninety-five women living with HIV were age-matched to 284 controls. Women with HIV had lower income and bone mineral density and were more likely to smoke, take multiple medications and be non-white. Vitamin D dietary intake was lower in women living with HIV versus controls [0.76 vs. 1.79 µg/day; adjusted odds ratio (aOR) for greater than or equal to median intake 0.29 (0.12-0.61), p = 0.002], but any supplementation was higher [62.2% vs. 44.7%; aOR = 3.44 (95% CI: 1.16-11.00), p = 0.03]. Total vitamin D intake was similar between groups. Smoking was associated with no supplementation; non-white ethnicity and low income were related to lower dietary intake. CONCLUSIONS: Women living with HIV showed lower dietary vitamin D intake but higher supplementation rates, suggesting that care providers are promoting supplementation. Women living with HIV who smoke, have low incomes and are non-white may particularly benefit from targeted efforts to improve vitamin D intake.

Factors associated with contraceptive use among women living with HIV in Canada: a controlled, cross-sectional study.

BACKGROUND: Multiple contraindications to combined hormonal contraceptives (CHC) use exist. The impact of these factors on contraceptive choice, particularly among women living with HIV (WLWH), is not well understood. We measured and compared the prevalence of contraceptive use and contraindications among WLWH and women not living with HIV (controls). METHODS: We examined cross-sectional survey and medical chart data from 83 WLWH and 62 controls, aged 16-49 and sexually active, from 2013-2017. We compared the age-adjusted prevalence and types of contraceptives used in the last month and the proportion of women with CHC contraindications, including drug interactions, medical comorbidities, and smoking at ≥ 35 years old. All WLWH received care at an interdisciplinary, women-centred HIV clinic. RESULTS: Compared to controls, WLWH were older (median [IQR)] 39 [34-43] vs 31 [23-41] years; p = 0.003), had less post-secondary education (37% vs 73%; p < 0.001), and more often had household income < $15,000/year (49% vs 30%; p = 0.006). WLWH trended to higher contraceptive prevalence than controls (80% vs 63%; p = 0.06 adjusted for age). Overall hormonal contraceptive use was similar. However, despite controlling for age, WLWH used CHC less (4% vs 18%; p = 0.006) than controls, and had more frequently undergone tubal ligation (12% vs 2%; p = 0.03). WLWH also experienced more CHC contraindications (54% vs 13%; p = 0.0001), including smoking at ≥ 35 years old (30% vs 6%; p = 0.0003) or a CHC-related drug interaction (all antiretroviral related) (25% vs 0%; p = 0.0001). CONCLUSIONS: WLWH attending our interdisciplinary clinic used hormonal contraception at similar rates as controls, though with different types. Differences may reflect different distributions of CHC contraindications. CHC contraindications present barriers to accessing the full range of contraceptive choices for WLWH. Guidelines and education for care providers and WLWH regarding contraceptive choices and drug interactions are needed, especially when care is provided without the benefit of an interdisciplinary women-centered healthcare team.

BACKGROUND: There are many reasons why individuals cannot use combined hormonal contraceptives (CHC). The impact of these reasons on contraceptive choice for women living with HIV (WLWH) are poorly understood. We measured and compared the prevalence of contraceptive choice and factors that may preclude their use in WLWH. METHODS: We examined survey and medical chart data from 83 WLWH and 62 controls (women not living with HIV), aged 16­49 and sexually active, from 2013 to 2017. We compared the prevalence and types of contraceptives used in the last month and the proportion of women with factors that would not allow the use of CHC, including drug interactions, medical conditions, and smoking at ≥ 35 years old. All WLWH received care at a women-centred HIV clinic. RESULTS: Compared to controls, WLWH were older, had less post-secondary education, and more often had household income < $15,000/year. WLWH were more likely to use contraception than controls. Overall hormonal contraceptive use was similar. However, even when accounting for age, WLWH used CHC less than controls, and had more frequently undergone tubal ligation. WLWH also had more reasons that would preclude the use of CHC contraindications including smoking at ≥ 35 years old or a CHC-related drug interaction. CONCLUSIONS: WLWH attending our interdisciplinary clinic used combined hormonal contraception at similar rates as controls, though with different types. Differences may reflect the fact that WLWH more often have factors that do not allow the safe use of CHC. Guidelines and education for care providers and WLWH regarding contraceptive choices and drug interactions are needed.

Daily Oral Supplementation with 60 mg of Elemental Iron for 12 Weeks Alters Blood Mitochondrial DNA Content, but Not Leukocyte Telomere Length in Cambodian Women.

There is limited evidence regarding the potential risk of untargeted iron supplementation, especially among individuals who are iron-replete or have genetic hemoglobinopathies. Excess iron exposure can increase the production of reactive oxygen species, which can lead to cellular damage. We evaluated the effect of daily oral supplementation on relative leukocyte telomere length (rLTL) and blood mitochondrial DNA (mtDNA) content in non-pregnant Cambodian women (18-45 years) who received 60 mg of elemental iron as ferrous sulfate (n = 190) or a placebo (n = 186) for 12 weeks. Buffy coat rLTL and mtDNA content were quantified by monochrome multiplex quantitative polymerase chain reaction. Generalized linear mixed-effects models were used to predict the absolute and percent change in rLTL and mtDNA content after 12 weeks. Iron supplementation was not associated with an absolute or percent change in rLTL after 12 weeks compared with placebo (ß-coefficient: -0.04 [95% CI: -0.16, 0.08]; p = 0.50 and ß-coefficient: -0.96 [95% CI: -2.69, 0.77]; p = 0.28, respectively). However, iron supplementation was associated with a smaller absolute and percent increase in mtDNA content after 12 weeks compared with placebo (ß-coefficient: -11 [95% CI: -20, -2]; p = 0.02 and ß-coefficient: -11 [95% CI: -20, -1]; p= 0.02, respectively). Thus, daily oral iron supplementation for 12 weeks was associated with altered mitochondrial homeostasis in our study sample. More research is needed to understand the risk of iron exposure and the biological consequences of altered mitochondrial homeostasis in order to inform the safety of the current global supplementation policy.

Preterm Birth and Antiretroviral Exposure in Infants HIV-exposed Uninfected.

BACKGROUND: Infants HIV-exposed and uninfected (IHEU) who are born to women living with HIV are at an increased risk of preterm birth (PTB). Antenatal exposure to certain maternal antiretroviral therapy (ART) regimens has been associated with PTB, although existing studies in this domain are limited and report discordant findings. We determined odds of PTB among IHEU by antenatal ART regimens and timing of exposure, adjusting for maternal risk factors. METHODS: We retrospectively studied IHEU born in British Columbia (BC), Canada between 1990 and 2012 utilizing provincial health administrative databases. We included data from a control group of infants HIV-unexposed and uninfected (IHUU) matched ~3:1 for each IHEU on age, sex and geocode. RESULTS: A total of 411 IHEU and 1224 IHUU were included in univariable analysis. PTB was more frequent among IHEU (20%) compared with IHUU (7%). IHEU were more often antenatally exposed to alcohol, tobacco, as well as prescription, nonprescription, and illicit drugs (IHEU: 36%, 8% and 35%; vs. IHUU: 3%, 1% and 9%, respectively). After adjusting for maternal substance use and smoking exposure, IHEU remained at increased odds of PTB [adjusted odds ratio (aOR) (95% CI): 2.66; (1.73, 4.08)] compared with matched IHUU controls. ART-exposed IHEU (excluding those with NRTIs only ART) had lower adjusted odds of PTB compared with IHEU with no maternal ART exposure, regardless of regimen [aOR range: 0.16-0.29 (0.02-0.95)]. Odds of PTB between IHEU exposed to ART from conception compared with IHEU exposed to ART postconception did not differ [aOR: 0.91 (0.47, 1.76)]; however, both groups experienced lower odds of PTB compared with IHEU with no maternal ART [preconception: aOR: 0.28 (0.08, 0.89); postconception: aOR 0.30 (0.11, 0.83)]. CONCLUSIONS: BC IHEU were over twice as likely to be born preterm compared with demographically matched controls. Maternal substance use in pregnancy modulated this risk; however, we found no adverse associations of PTB with exposure to antenatal ART.

Inverse relationship between leukocyte telomere length attrition and blood mitochondrial DNA content loss over time.

Leukocyte telomere length (LTL) and whole blood mitochondrial DNA (WB mtDNA) content are aging markers impacted by chronic diseases such as human immunodeficiency virus (HIV) infection. We characterized the relationship between these two markers in 312 women ≥12 years of age living with HIV and 300 HIV-negative controls. We found no relationship between the two markers cross-sectionally. In multivariable models, age, ethnicity, HIV, and tobacco smoking were independently associated with shorter LTL, and the former three with lower WB mtDNA. Longitudinally, among a subgroup of 228 HIV participants and 68 HIV-negative controls with ≥2 biospecimens ≥1 year apart, an inverted pattern was observed between the rates of change in LTL and WB mtDNA content per year, whereby faster decline of one was associated with the preservation of the other. Furthermore, if HIV viral control was not maintained between visits, increased rates of both LTL attrition and WB mtDNA loss were observed. We describe a novel relationship between two established aging markers, whereby rates of change in LTL and WB mtDNA are inversely related. Our findings highlight the importance of maintaining HIV viral control, the complementary longitudinal relationship between the two markers, and the need to consider both in aging studies.

Prolonged Amenorrhea and Low Hip Bone Mineral Density in Women Living With HIV-A Controlled Cross-sectional Study.

BACKGROUND: Women living with HIV (WLWH) have higher rates of prolonged secondary amenorrhea (no flow for ≥1 year) than HIV-negative women. Both having amenorrhea and being HIV positive are associated with lower areal bone mineral density (BMD). However, their combined BMD effects remain unclear. Therefore, we investigated prolonged amenorrhea and BMD in WLWH and controls. METHODS: This cross-sectional study enrolled WLWH and HIV-negative control women aged 19-68 years of similar backgrounds. We assessed BMD (Hologic; as age- and ethnicity-matched Z-scores) in the Children and women: AntiRetrovirals and Markers of Aging cohort. Participants were stratified by amenorrhea history defined as past/present lack of menses for ≥1 year at age 45 and younger and not because of surgery, breastfeeding, pregnancy, or hormonal contraception. Hip and spine Z-scores by amenorrhea/no amenorrhea used linear models with multivariable analysis for relationships within WLWH. RESULTS: WLWH (N = 129) were similar to controls (N = 129) in age, body mass index, ethnicity, and substance use. Among WLWH, 21% experienced prolonged amenorrhea vs. 9% in controls. WLWH had significantly lower total hip (mean ± SD: -0.4 ± 0.9 vs. 0.3 ± 1.1; P < 0.001) and spine (-0.5 ± 1.3 vs. 0.2 ± 1.3; P = 0.001) Z-scores than controls. Amenorrhea was independently associated with hip (P = 0.01) but not spine (P = 0.94) BMD by multivariable linear regression. WLWH with amenorrhea had lower hip Z-scores (-0.8 ± 0.9) than those without (-0.3 ± 0.8; P = 0.01). They also had higher rates of substance use, smoking, opioid therapy, hepatitis C coinfection, and lower CD4 nadir. CONCLUSIONS: WLWH had higher rates of prolonged amenorrhea and lower BMD than controls. WLWH with amenorrhea experienced lower hip BMD Z-scores than those without. Prolonged amenorrhea is an added osteoporosis risk in WLWH.

Comorbidity and polypharmacy among women living with HIV in British Columbia.

OBJECTIVE: To characterize comorbid disease and medication burden among women living with HIV (WLWH) in British Columbia (BC), Canada. DESIGN: We examined baseline data from 267 WLWH and 276 HIV-negative women, aged at least 19 years, enrolled in the Children and Women: Antiretrovirals and Markers of Aging (CARMA) cohort. METHODS: Self-reported demographic, medical condition, medication, vitamin, and substance exposure data were collected at baseline CARMA study visits. We considered conditions with appropriate concomitant medications to be 'treated'. Wilcoxon rank-sum and Fisher's exact tests compared continuous and categorical variables between WLWH and HIV-negative women. Number of diagnoses, prescribed medications (excluding HIV/antiretrovirals), vitamins, and prevalence of depression/anxiety/panic disorder were compared using negative binomial and logistic regressions for continuous and binary variables, respectively. RESULTS: WLWH were younger [median, interquartile range (IQR) 39.9, 33.6-46.9 vs. 43.6, 31.8-54.6 years; P = 0.01], attained lower education (40.5 vs. 69.6% college/university; P < 0.001), and more often currently smoked tobacco (47.9 vs. 31.9%; P < 0.001) or had income less than $15 000/year (49.0 vs. 43.1%; P < 0.001). Although younger, and despite omitting HIV infection, WLWH had a greater number of diagnoses (incidence rate ratio, 95% confidence interval 1.58, 1.38-1.81; P < 0.001), and more depression/anxiety/panic disorder vs. controls (odds ratio, 95% CI 1.86, 1.22-2.83; P = 0.004). Our model predicts that with mean BMI (26.3), WLWH and HIV-negative peers would have two comorbid diagnoses by age 30 and 60, respectively. CONCLUSIONS: WLWH living in BC have more comorbid illness earlier in life than their HIV-negative peers, and have very high rates of depression/anxiety/panic disorder. Addressing mental health and comorbid conditions is essential to improving health outcomes among WLWH.

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.

Dynamics of leukocyte telomere length in pregnant women living with HIV, and HIV-negative pregnant women: A longitudinal observational study.

BACKGROUND: HIV-mediated inflammation and immune activation can accelerate telomere attrition. In addition, antiretrovirals can inhibit telomerase, possibly shortening telomeres. We examined the longitudinal dynamics of leukocyte telomere length (LTL) during pregnancy in a unique cohort of women living with HIV (WLWH) treated with combination antiretroviral therapy (cART), and HIV-negative control women. METHODS: Blood was collected at three visits during pregnancy, at 13-23, >23-30, and >30-40 weeks of gestation, and for WLWH only, at 6 weeks post-partum. LTL was measured by qPCR and both cross-sectional and longitudinal (MANOVA) models were used to examine possible predictors of LTL among participants who attended all three visits during pregnancy. RESULTS: Among WLWH (n = 64) and HIV-negative women (n = 41), within participant LTL were correlated throughout pregnancy (p<0.001). LTL was shorter among WLWH at first visit, but this difference waned by the second visit. WLWH who discontinued cART post-partum experienced a decrease in LTL. Longitudinally, LTL was similar in both groups and increased as gestation progressed, a change that was more pronounced among women under 35 years. Among WLWH, both smoking throughout pregnancy (p = 0.04) and receiving a ritonavir-boosted protease inhibitor-based regimen (p = 0.03) were independently associated with shorter LTL. CONCLUSIONS: LTL increases as pregnancy progresses; the reasons for this are unknown but may relate to changes in blood volume, hormones, and/or cell subset distribution. While our observations need confirmation in an independent cohort, our data suggest that although some cART regimens may influence LTL, being on cART appears overall protective and that stopping cART post-partum may negatively impact LTL. The effect of smoking on LTL is clearly negative, stressing the importance of smoking cessation.

A mitochondrial DNA D loop insertion detected almost exclusively in non-replicating tissues with maternal inheritance across three generations.

Muscle biopsy identified a possibly pathogenic, mitochondrial DNA D-loop insertion, in each of 5 family members from two generations, that was otherwise undetectable in most other tissues. The tissue specific regulation of heteroplasmy is reflected in an age related increase in muscle heteroplasmy level, across the pedigree. This latter finding is in keeping with previous reports (e.g. T408A, C16327) but differs in having a very high muscle heteroplasmy level, and appears maternally transmitted.

Neurodevelopmental outcomes and in-utero antiretroviral exposure in HIV-exposed uninfected children.

OBJECTIVES: To assess and compare neurodevelopmental disorders in HIV-exposed uninfected (HEU) and HIV-unexposed uninfected (HUU) children in British Columbia, Canada. To determine associations between these outcomes and in-utero exposure to antiretroviral drugs. DESIGN: Retrospective controlled cohort study. METHODS: Data were collected on 446 HEU children and 1323 HUU children (matched ∼1 : 3 for age, sex, and geocode) born between 1990 and 2012. Multivariable logistic regressions determined odds ratios of neurodevelopmental disorder diagnoses. RESULTS: HEUs had three times higher odds of being born preterm (P < 0.0001), and a more than two-fold increase in odds for autism, disturbance of emotions, hyperkinetic syndrome, and developmental delay compared with matched HUUs (P < 0.02) in unadjusted analysis. This association was reduced [adjusted neurodevelopmental disorder odds ratio (AOR) = 1.67; 95% confidence interval: 1.12-2.48; P = 0.011] after adjusting for maternal substance use and/or smoking (children born after April 2000). Regardless of antiretroviral exposure type (i.e. none, treatment with one or multiple drug classes), HEUs had higher odds of any neurodevelopmental disorders compared with matched HUUs; however, there was no evidence suggesting any specific classes of antiretroviral drugs or exposure durations increased their likelihood of neurodevelopmental disorders. CONCLUSION: The results suggest no adverse associations between antiretroviral drugs and neurodevelopmental disorders within antiretroviral-exposed HEU children in our cohort. Prevalence of neurodevelopmental disorders is higher in HEUs; however, maternal substance use plays a role, as could other environmental factors not captured. These findings highlight a need for holistic support for pregnant women as well as careful developmental monitoring of HEUs past infancy, and access to early interventions, particularly among those born preterm and those exposed to addictive substances.

A Monochrome Multiplex Real-Time Quantitative PCR Assay for the Measurement of Mitochondrial DNA Content.

Mitochondrial DNA copies per cell (mtDNA content) can fluctuate with cellular aging, oxidative stress, and mitochondrial dysfunction, and has been investigated in cancer, diabetes, HIV, and metabolic disease. mtDNA content testing in both clinical and basic settings is expected to increase as research uncovers its biological relevance. Herein, we present a novel mtDNA content assay developed on monochrome multiplex real-time quantitative PCR (MMqPCR) principles. This assay offers a greater than twofold improvement on time effectiveness and cost-effectiveness over conventional (monoplex) qPCR, as well as improved reproducibility given the reduced effects of human pipetting errors. The new MMqPCR method was compared with the gold standard monoplex qPCR assay on DNA from a variety of sources, including human whole blood, skeletal muscle, and commercial cell lines. The MMqPCR assay is reproducible (n = 98, r = 0.99, P < 0.0001) and highly correlated to the monoplex qPCR assay (n = 160, r > 0.98, P < 0.0001). Intra-assay and interassay variabilities, as established independently by multiple operators, range between 4.3% and 7.9% and between 2.9% and 9.2%, respectively. This robust assay can quantify >82 pg of template DNA per reaction, with a minimum mtDNA/nuclear DNA ratio of 20, and is especially suitable for studies that require high throughput.

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

BACKGROUND: 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. SETTING: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Leukocyte Telomere Length at Birth and During the Early Life of Children Exposed to but Uninfected With HIV After In Utero Exposure to Antiretrovirals.

Background: Maternal combination antiretroviral therapy (cART) during pregnancy could impact the health of human immunodeficiency virus (HIV)-exposed, HIV-uninfected (HEU) children, because some antiretrovirals cross the placenta and can inhibit telomerase. Our objective was to compare leukocyte telomere length (LTL) in HEU children and HIV-unexposed, HIV-uninfected (HUU) children at birth and in early life and to investigate any relationship with cART exposure. Methods: HEU and HUU children's blood LTL was compared cross-sectionally at birth, and during the first three years of life. Longitudinal HEU LTL dynamics was evaluated over that same period. Results: At birth, the LTL in HEU children (n = 114) was not shorter than that in HUU children (n = 86), but female infants had longer LTL than male infants. Maternal cART (duration or type) showed no association with shorter infant LTL. Among 214 HEU children age- and sex-matched at a 1:1 ratio to HUU children, LTL declined similarly in both groups. In a longitudinal analysis, LTL attrition in HEU children was rapid from birth to 1 year of age and gradual thereafter. Zidovudine prophylaxis did not significantly alter LTL. Conclusions: Our results indicate that from birth to 3 years of age, the LTL in HEU children is not negatively affected by exposure to maternal HIV infection and cART, at least not to the regimens used within this Canadian cohort, a reassuring finding.

Competing Factors Link to Bone Health in Polycystic Ovary Syndrome: Chronic Low-Grade Inflammation Takes a Toll.

Chronic inflammation predisposes to poor bone health. Women with polycystic ovary syndrome (PCOS) experience androgen excess, ovulatory disturbances, insulin resistance, abdominal adiposity and chronic inflammation. Our objective was to investigate the relationships among bone health parameters, chronic subclinical inflammation and anthropometric measures in premenopausal women with and without PCOS. In 61 premenopausal women, 22 women with PCOS and 39 controls, we assessed bone parameters (total hip bone mineral density [BMD] by dual-energy X-ray absorptiometry and radius strength-strain index [SSI] by peripheral quantitative computed tomography), inflammation (C-reactive protein/albumin), oxidative stress (leukocyte telomere length, urinary 8-hydroxydeoxyguanosine); hemoglobin A1c; anthropometric measures (body mass index, waist-to-height ratio, cross-sectional muscle area). A diagnosis of PCOS negatively predicted (beta = -0.251, p = 0.022) hip BMD in a regression model including weight. In women with PCOS, inflammation, which was predicted by increased waist-to-height ratio and current use of oral contraceptives, attenuated the positive influences of increased weight and muscle mass on bone strength and was inversely associated with radial SSI (R2 = 0.25, p = 0.018). In conclusion, chronic subclinical inflammation may negatively impact bone physiology in women with PCOS. Strategies focused on reducing abdominal adiposity and avoiding medications that increase inflammation may counter this effect.

Childhood Personality, Betrayal Trauma, and Leukocyte Telomere Length in Adulthood: A Lifespan Perspective on Conscientiousness and Betrayal Traumas as Predictors of a Biomarker of Cellular Aging.

Conscientiousness is associated with longevity. As such, identifying the biological pathways linking personality to mortality is important. This study employs longitudinal data spanning >40 years to test prospective associations with Leukocyte Telomere Length (LTL), a potential marker of cellular aging. Because telomeres shorten over time, and are sensitive to oxidative stress, shorter LTL may reflect cumulative damage associated with negative health behaviors and past stressful events. We investigated childhood conscientiousness as a protective factor, expecting an association with longer LTL in adulthood, possibly reflecting slower LTL shortening. Potential lifespan pathways involving childhood trauma, smoking behaviors, and Body Mass Index (BMI) were explored. Childhood conscientiousness showed a small raw association with LTL (r = .08, p = .04), although this effect did not persist when controlling for age and sex. Despite this lack of a direct effect on LTL, we detected an indirect effect operating jointly through BMI and smoking. Higher rates of childhood betrayal trauma were associated with shorter LTL. Contrary to our hypothesis that conscientiousness would buffer this effect, we found evidence for an interaction with childhood betrayal traumas where the association between childhood betrayal traumas and LTL was larger for those higher on conscientiousness in childhood.

Optimization of a Relative Telomere Length Assay by Monochromatic Multiplex Real-Time Quantitative PCR on the LightCycler 480: Sources of Variability and Quality Control Considerations.

Telomere length (TL) measurement is central to many biomedical research, population, and epidemiology studies, with promising potential as a clinical tool. Various assays are used to determine TL, depending on the type and size of the sample. We describe the detailed optimization of a monochromatic multiplex real-time quantitative PCR (MMqPCR) assay for relative TL using the LightCycler 480. MMqPCR was initially developed using a different instrument with many separate reagents. Differences in instrument performance, reagents, and workflow required substantial optimization for the assay to be compatible with the LightCycler 480. We optimized the chemistry of the assay using a purchased one-component reaction mix and herein describe sources of variability and quality control relevant to the MMqPCR TL assay on any instrument. Finally, the assay was validated against other TL assays, such as terminal restriction fragment, Southern blot, and flow fluorescent in situ hybridization. The correlations obtained between data from MMqPCR and these assays (R(2) = 0.88 and 0.81) were comparable to those seen with the monoplex version (R(2) = 0.85 and 0.82) when the same samples were assayed. The intrarun and interrun CV ranged from 4.2% to 6.2% and 3.2% to 4.9%, respectively. We describe a protocol for measuring TL on the LightCycler platform that provides a robust high-throughput method applicable to clinical diagnostics or large-scale studies of archived specimens.