CD8+ cells and small viral reservoirs facilitate post-ART control of SIV replication in M3+ Mauritian cynomolgus macaques initiated on ART two weeks post-infection.

Sustainable HIV remission after antiretroviral therapy (ART) withdrawal, or post-treatment control (PTC), remains a top priority for HIV treatment. We observed surprising PTC in an MHC-haplomatched cohort of MHC-M3+ SIVmac239+ Mauritian cynomolgus macaques (MCMs) initiated on ART at two weeks post-infection (wpi). None of the MCMs possessed MHC haplotypes previously associated with SIV control. For six months after ART withdrawal, we observed undetectable or transient viremia in seven of the eight MCMs, despite detecting replication competent SIV using quantitative viral outgrowth assays. In vivo depletion of CD8α+ cells induced rebound in all animals, indicating the observed PTC was mediated, at least in part, by CD8α+ cells. With intact proviral DNA assays, we found that MCMs had significantly smaller viral reservoirs two wpi than a cohort of identically infected rhesus macaques, a population that rarely develops PTC. We found a similarly small viral reservoir among six additional SIV+ MCMs in which ART was initiated at eight wpi, some of whom exhibited viral rebound. These results suggest that an unusually small viral reservoir is a hallmark among SIV+ MCMs. By evaluating immunological differences between MCMs that did and did not rebound, we identified that PTC was associated with a reduced frequency of CD4+ and CD8+ lymphocyte subsets expressing exhaustion markers. Together, these results suggest a combination of small reservoirs and immune-mediated virus suppression contribute to PTC in MCMs. Further, defining the immunologic mechanisms that engender PTC in this model may identify therapeutic targets for inducing durable HIV remission in humans.

Human bronchopulmonary disposition and plasma pharmacokinetics of oral bemnifosbuvir (AT-527), an experimental guanosine nucleotide prodrug for COVID-19.

BACKGROUND: Bemnifosbuvir (AT-527) is a novel oral guanosine nucleotide antiviral drug for the treatment of persons with COVID-19. Direct assessment of drug disposition in the lungs, via bronchoalveolar lavage, is necessary to ensure antiviral drug levels at the primary site of SARS-CoV-2 infection are achieved. OBJECTIVES: This Phase 1 study in healthy subjects aimed to assess the bronchopulmonary pharmacokinetics, safety and tolerability of repeated doses of bemnifosbuvir. METHODS: A total of 24 subjects were assigned to receive bemnifosbuvir twice daily at doses of 275, 550 or 825 mg for up to 3.5 days. RESULTS: AT-511, the free base of bemnifosbuvir, was largely eliminated from the plasma within 6 h post dose in all dosing groups. Antiviral drug levels of bemnifosbuvir were consistently achieved in the lungs with bemnifosbuvir 550 mg twice daily. The mean level of the guanosine nucleoside metabolite AT-273, the surrogate of the active triphosphate metabolite of the drug, measured in the epithelial lining fluid of the lungs was 0.62 µM at 4-5 h post dose. This exceeded the target in vitro 90% effective concentration (EC90) of 0.5 µM for antiviral drug exposure against SARS-CoV-2 replication in human airway epithelial cells. Bemnifosbuvir was well tolerated across all doses tested, and most treatment-emergent adverse events reported were mild in severity and resolved. CONCLUSIONS: The favourable pharmacokinetics and safety profile of bemnifosbuvir demonstrates its potential as an oral antiviral treatment for COVID-19, with 550 mg bemnifosbuvir twice daily currently under further clinical evaluation in persons with COVID-19.

Validity of Using the Load-Velocity Relationship to Estimate 1 Repetition Maximum in the Back Squat Exercise: A Systematic Review and Meta-Analysis.

ABSTRACT: LeMense, AT, Malone, GT, Kinderman, MA, Fedewa, MV, and Winchester, LJ. Validity of using the load-velocity relationship to estimate 1 repetition maximum in the back squat exercise: a systematic review and meta-analysis. J Strength Cond Res 38(3): 612-619, 2024-The one repetition maximum (1RM) test is commonly used to assess muscular strength. However, 1RM testing can be time consuming, physically taxing, and may be difficult to perform in athletics team settings with practice and competition schedules. Alternatively, 1RM can be estimated from bar or movement velocity at submaximal loads using the minimum velocity threshold (MVT) method based on the load-velocity relationship. Despite its potential utility, this method's validity has yielded inconsistent results. The purpose of this systematic review and meta-analysis was to assess the validity of estimated 1RM from bar velocity in the back squat exercise. A systematic search of 3 electronic databases was conducted using combinations of the following keywords: "velocity-based training," "load-velocity profiling," "mean velocity," "mean propulsive velocity," "peak velocity," "maximal strength," "1RM," "estimation," "prediction," "back squat," and "regression." The search identified 372 unique articles, with 4 studies included in the final analysis. Significance was defined as a p level less than 0.05. A total of 27 effects from 71 subjects between the ages of 17-25 years were analyzed; 85.2% of effects were obtained from male subjects. Measured 1RMs ranged from 86.5 to 153.1 kg, whereas estimated 1RMs ranged from 88.6 to 171.6 kg. Using a 3-level random effects model, 1RM back squat was overestimated when derived from bar velocity using the MVT method (effect sizes [ES] = 0.5304, 95% CI: 0.1878-0.8730, p = 0.0038). The MVT method is not a viable option for estimating 1RM in the free weight back squat. Strength and conditioning professionals should exercise caution when estimating 1RM from the load-velocity relationship.

Disparities in physical activity between sexual minority and heterosexual women: A systematic review and meta-analysis.

Research examining potential differences in physical activity (PA) between sexual minority women (SMW) and heterosexual women have yielded inconsistent results. OBJECTIVE: Therefore, the purpose of this systematic review and meta-analysis is to examine potential differences in PA between SMW and heterosexual women and to identify potential moderators that may partially explain observed differences in PA. METHODS: All studies were peer reviewed, published in English, and included a continuous measure of PA for SMW and heterosexual women. A standardized mean difference effect size (ES) was used to compare groups, with random effects models used to estimate a mean ES and 95% CI using a 3-level meta-analysis model to adjust for the correlation between effects nested within studies. RESULTS: The cumulative results of 24 effects gathered from 7 studies indicated there was no difference in PA between SMW (n = 1619) and heterosexual women (n = 103,295) (ES = -0.038, 95%CI -0.179 to 0.102, p = 0.576). Despite no mean differences, moderate-high heterogeneity was observed, indicating that the results were not consistent across effects (I2 = 64.8%, Q23 = 36.7, p = 0.035). The difference in PA was associated with age (ß = -0.018, 95%CI -0.034 to -0.003, p = 0.022) and BMI (ß = -0.145, 95%CI -0.228 to -0.061, p = 0.002), with a quadratic relationship observed for both variables. CONCLUSIONS: Although the results of the current analysis did not indicate significant differences in PA behaviors between SMW and heterosexual women, age and BMI modify the association and are curvilinear in nature; such that smaller differences in PA were observed between SMW and heterosexual women when samples were middle-aged and overweight.

Pharmacokinetics of levonorgestrel and etonogestrel contraceptive implants over 48†weeks with rilpivirine- or darunavir-based antiretroviral therapy.

BACKGROUND: Pharmacokinetic data are lacking for progestin-releasing subdermal contraceptive implants when used with either rilpivirine- or darunavir/ritonavir-based ART. OBJECTIVES: To characterize the pharmacokinetics of etonogestrel or levonorgestrel implants when administered with these ART regimens over 48 weeks. PATIENTS AND METHODS: Two separate, parallel, three-group, non-randomized, pharmacokinetic studies evaluated either etonogestrel or levonorgestrel in women receiving rilpivirine- or darunavir-based ART compared with women without HIV (control group). Participants on ART were switched to rilpivirine-based ART with a run-in period of 6 weeks or darunavir-based ART with a run-in of 2 weeks prior to implant insertion. Plasma was collected on Day 0, and 1, 4, 12, 24, 36 and 48 weeks post-insertion. Plasma progestin concentrations were compared between ART and control groups by geometric mean ratio (GMR) and 90% CI. RESULTS: At the primary endpoint of Week 24, progestin concentrations were similar between the rilpivirine and control groups [etonogestrel: 1.18 (0.99-1.37); levonorgestrel: 1.16 (0.97-1.33)]. At Week 24, progestin exposure was higher in the darunavir groups compared with the control group [etonogestrel: 2.56 (1.69-3.28); levonorgestrel: 1.89 (1.38-2.29)]. Results remained consistent through to Week 48. No differences in etonogestrel-related adverse events were observed, but both ART groups experienced more menstrual abnormalities versus the control group with levonorgestrel. CONCLUSIONS: Etonogestrel and levonorgestrel concentrations were not altered by rilpivirine-based ART. Although progestin concentrations were higher in the ART groups containing ritonavir-boosted darunavir, no implant-related serious adverse events were observed. Both progestin-releasing implants are an appropriate contraceptive option with either rilpivirine- or darunavir/ritonavir-based ART.

Comparing Physical Fitness in Career vs. Volunteer Firefighters.

ABSTRACT: Morris, CE, Arnett, SW, and Winchester, LJ. Comparing physical fitness in career vs. volunteer firefighters. J Strength Cond Res 36(5): 1304-1309, 2022-The purpose of this study was to assess the potential similarities and differences in health and physical fitness profiles between career firefighters (CFF) and volunteer firefighters (VFF). The research protocol consisted of a health and physical fitness assessment, testing the 5 components of health-related fitness using previously published and accepted protocols. The subject population consisted of a total of 138 firefighters, including 119 CFF and 19 VFF. Statistical significance was defined as a p level less than 0.05. An independent t test showed evidence of CFF having a significantly higher value/score for the following variables: height (p = 0.034), V̇o2max (p = 0.006), push-ups completed (p = 0.024), and plank time (p < 0.001). Volunteer firefighters had a significantly higher value for the following variables: fat mass (p = 0.002), body fat percentage (p < 0.001), and absolute grip strength (p = 0.029). There were no significant differences between groups for the following variables: age (p = 0.299), body mass (p = 0.166), fat-free mass (p = 0.281), body mass index (p = 0.057), flexibility (p = 0.106), or relative grip strength (p = 0.887). With regard to physical fitness testing, the VFF had a significantly worse fitness profile across a number of variables than the CFF. Practical applications: Despite the financial and commitment status of volunteer firefighting departments, their members perform an equally dangerous and important job as do firefighters of professional/career firefighting departments, and more attention should be directed toward developing the fitness and performance of these firefighters.

Intramuscular and subcutaneous administration of antiretroviral drugs, compared with oral, enhances delivery to lymphoid tissues in BALB/c mice.

BACKGROUND: Multiple tissue reservoirs are established soon after HIV infection, and some tissues may also be pharmacological sanctuaries. Parenteral administration of antiretroviral (ARV) drugs for treatment and prevention of HIV infection is an active area of drug development. The influence of route of administration on ARV tissue pharmacokinetics is not known. OBJECTIVES: To investigate ARV pharmacokinetics in lymphatic and select non-lymphatic tissues (e.g. brain and testes) after intramuscular and subcutaneous administration compared with oral in BALB/c mice. METHODS: Tissue concentrations of cobicistat, efavirenz, elvitegravir, maraviroc, rilpivirine, tenofovir alafenamide and tenofovir disoproxil fumarate were determined. The tissue penetration ratio (TPR) was the primary measure for comparison; a change in TPR arises from factors affecting tissue distribution controlling for changes in systemic bioavailability. RESULTS: Intramuscular and subcutaneous delivery increased TPRs in the lymph node and spleen for 27 of 28 (96%) drug administration events. Decreased TPRs, however, were found in some tissues such as the brain and testes. CONCLUSIONS: These results demonstrate a change in route of drug administration from oral to intramuscular or subcutaneous can change tissue uptake. This has implications for HIV pharmacotherapy. For example, HIV persists in lymphoid tissues despite long-term oral ARV therapy, and low ARV concentrations have been found in lymphoid tissues. The improved ARV lymphatic tissue bioavailability with intramuscular and subcutaneous administration allows future studies to investigate these routes of drug administration as a therapeutic manoeuvre to limit viral persistence and eliminate viral sanctuaries in the lymphatic tissues, which is a prerequisite for eradication of HIV.

Validity of Smartphone Heart Rate Variability Pre- and Post-Resistance Exercise.

The aim was to examine the validity of heart rate variability (HRV) measurements from photoplethysmography (PPG) via a smartphone application pre- and post-resistance exercise (RE) and to examine the intraday and interday reliability of the smartphone PPG method. Thirty-one adults underwent two simultaneous ultrashort-term electrocardiograph (ECG) and PPG measurements followed by 1-repetition maximum testing for back squats, bench presses, and bent-over rows. The participants then performed RE, where simultaneous ultrashort-term ECG and PPG measurements were taken: two pre- and one post-exercise. The natural logarithm of the root mean square of successive normal-to-normal (R-R) differences (LnRMSSD) values were compared with paired-sample t-tests, Pearson product correlations, Cohen's d effect sizes (ESs), and Bland-Altman analysis. Intra-class correlations (ICC) were determined between PPG LnRMSSDs. Significant, small-moderate differences were found for all measurements between ECG and PPG: BasePre1 (ES = 0.42), BasePre2 (0.30), REPre1 (0.26), REPre2 (0.36), and REPost (1.14). The correlations ranged from moderate to very large: BasePre1 (r = 0.59), BasePre2 (r = 0.63), REPre1 (r = 0.63), REPre2 (r = 0.76), and REPost (r = 0.41)-all p < 0.05. The agreement for all the measurements was "moderate" (0.10-0.16). The PPG LnRMSSD exhibited "nearly-perfect" intraday reliability (ICC = 0.91) and "very large" interday reliability (0.88). The smartphone PPG was comparable to the ECG for measuring HRV at rest, but with larger error after resistance exercise.

Blood Flow Restriction at High Resistance Loads Increases the Rate of Muscular Fatigue, but Does Not Increase Plasma Markers of Myotrauma or Inflammation.

Winchester, LJ, Morris, CE, Badinger, J, Wiczynski, TL, and VanWye, WR. Blood flow restriction at high resistance loads increases the rate of muscular fatigue, but does not increase plasma markers of myotrauma or inflammation. J Strength Cond Res 34(9): 2419-2426, 2020-High-load resistance training and blood flow restriction (BFR) training at low loads both promote protein synthesis and growth through different cell signaling mechanisms. Therefore, co-activation of these pathways could result in a synergistic effect for additional growth enhancement. The purpose of this study was to evaluate how BFR effects performance and physiological responses after an acute bout of high-load barbell squat training. Twelve resistance-trained, college-aged men and women performed 5 sets of barbell squats at 75% of 1 repetition maximum until failure under traditional (TRAD; control) or intermittent BFR conditions. Perceived limb pain and number of repetitions performed were recorded after each set. Blood samples were collected at baseline and 1-hour postexercise after each trial for analysis of myoglobin and interleukin-6 (IL-6). An alpha level of p < 0.05 was used to determine significance. Blood flow restriction trial performance significantly declined at set 3 and was lower than performance during control, whereas control performance did not decrease until set 5. Perceived limb pain was statistically increased with BFR use for the whole trial and was significantly higher with BFR during set 3 than observed during TRAD. Plasma myoglobin and IL-6 were significantly increased after both trials when compared with baseline, but were not significantly different between trials. Intermittent BFR use during high-load barbell squats increases the rate of muscular fatigue and perceived limb pain, but does not increase muscular damage or inflammatory response. Data obtained from this study can be used by fitness professionals as a means of potentially enhancing the rate of muscular hypertrophy.

Changes in Heart Rate Variability and Fatigue Measures Following Moderate Load Resistance Exercise.

The purpose of this study was to determine the relationship between changes in heart rate variability (HRV), neuromuscular performance, and fatigue biomarkers in response to a resistance exercise bout. The root mean square of successive RR interval differences (RMSSD), neuromuscular performance - isometric handgrip (IHG), countermovement jump (CMJ), mean propulsive velocity (MPV) - metabolic stress (lactate [Lac]) and inflammation (interleukin-6 [IL-6]) were measured in 30 subjects who performed 6×10 back squat (BS), 3×10 bench press (BP), and 3×10 bent-over rows (BR) at 70% of 1-repetition maximum (1RM). The RMSSD, neuromuscular performance, and biomarkers were measured 10 min pre-exercise and 30 min post-exercise (Post30); HRV and Lac were also measured immediately post-exercise (Post0). Pre- versus post-exercise differences were evaluated using paired-samples t-tests. Pearson's correlations were used to determine the association between changes. With the exception of IL-6 (P=0.296) and MPVBP (P=0.678), LnRMSSD, neuromuscular performance, and metabolic stress were different post- compared to pre-exercise. We observed moderate associations between ΔLnRMSSD Post0 and ΔLac Post0 (r = -0.44) and ΔLac Post30 (r = -0.55), respectively. Practitioners should use multiple training load indicators to gain an accurate depiction of recovery.

Assessing the lymphoid tissue bioavailability of antiretrovirals in human primary lymphoid endothelial cells and in mice.

BACKGROUND: The secondary lymphoid tissues (LTs), lymph nodes (LNs) and gut-associated lymphoid tissue (GALT) are considered reservoirs for HIV. Antiretrovirals (ARVs) have lower penetration into LT. In vitro models predictive of ARV LT penetration have not been established. OBJECTIVES: To develop an in vitro model of LT bioavailability using human lymphoid endothelial cells (HLECs) and investigate its predictability with in vivo pharmacokinetic (PK) studies in mice. METHODS: ARV bioavailability in HLECs was evaluated at the maximum plasma concentration (Cmax) observed in HIV-infected patients. ARVs were: abacavir, atazanavir, darunavir, dolutegravir, efavirenz, elvitegravir, emtricitabine, maraviroc, raltegravir, rilpivirine, ritonavir, tenofovir disoproxil fumarate and the PK booster cobicistat. The LT PK of representative drugs showing high (efavirenz), intermediate (dolutegravir) and low (emtricitabine) HLEC bioavailability was investigated in BALB/c mice given 50/10/30 mg/kg efavirenz/dolutegravir/emtricitabine orally, daily for 3 days. The concordance of in vitro and in vivo ARV bioavailability was examined. RESULTS: ARVs showed high (>67th percentile; rilpivirine, efavirenz, elvitegravir and cobicistat), intermediate (67th-33rd percentile; ritonavir, tenofovir disoproxil fumarate, dolutegravir and maraviroc) and low (<33rd percentile; atazanavir, darunavir, raltegravir, emtricitabine and abacavir) HLEC bioavailability. The hierarchy of efavirenz, dolutegravir and emtricitabine bioavailability in LN, gut and brain tissues of mice was: efavirenz>dolutegravir>emtricitabine. CONCLUSIONS: ARVs displayed distinct HLEC penetration patterns. PK studies of representative ARVs in LT of mice were concordant with HLEC bioavailability. These findings support further development of this approach and its translational predictability in humans.

Impact of efavirenz pharmacokinetics and pharmacogenomics on neuropsychological performance in older HIV-infected patients.

BACKGROUND: Pharmacokinetics (PK) and pharmacodynamics of efavirenz and its 8-hydroxy metabolite (8-OH-efavirenz) have not been robustly evaluated in older HIV-infected persons. OBJECTIVES: We investigated relationships between neuropsychological (NP) performance and efavirenz and 8-OH-efavirenz PK in HIV-infected individuals >50 years of age. METHODS: A cross-sectional study of HIV-infected adults on an efavirenz-containing regimen. The 12 and 18 h post-dose plasma efavirenz and 8-OH-efavirenz were quantified. CYP2B6 polymorphisms were investigated. Participants underwent neuropsychological tests; surveys were used for depression, sleep quality and anxiety. We investigated potential correlations of efavirenz and 8-OH-efavirenz plasma concentrations with NP performance, sleep, depression, anxiety and CYP2B6 polymorphisms. RESULTS: Thirty participants (24 men and 6 women) with mean age 57 years (range 50-68). Plasma efavirenz concentrations did not correlate with NP performance; however, higher plasma 8-OH-efavirenz correlated with better learning (P = 0.002), language (P = 0.002) and total NP z-scores (P = 0.003). No correlation was seen for efavirenz or 8-OH-efavirenz with sleep, anxiety or depression. Median 12 and 18 h efavirenz plasma concentrations were 1967 ng/mL (IQR 1476-2394) and 1676 ng/mL (IQR 1120-2062), respectively. Median 12 and 18 h 8-OH-efavirenz plasma concentrations were 378 ng/mL (IQR 223-589) and 384 ng/mL (IQR 216-621), respectively. CYP2B6 G516T was associated with significantly higher plasma efavirenz at 12 and 18 h (P = 0.02) but not worse NP function. CONCLUSIONS: Better neurocognitive functioning was associated with higher 8-OH-efavirenz but not efavirenz plasma concentrations. No correlation was observed with sleep or depression. These findings point to a need for greater understanding of the metabolic profile of efavirenz and 8-OH-efavirenz in plasma and the CNS and relationships with antiviral effect and neurotoxicity.

Hyperhomocysteinemia associated skeletal muscle weakness involves mitochondrial dysfunction and epigenetic modifications.

HHcy has been implicated in elderly frailty, but the underlying mechanisms are poorly understood. Using C57 and CBS+/- mice and C2C12 cell line, we investigated mechanisms behind HHcy induced skeletal muscle weakness and fatigability. Possible alterations in metabolic capacity (levels of LDH, CS, MM-CK and COX-IV), in structural proteins (levels of dystrophin) and in mitochondrial function (ATP production) were examined. An exercise regimen was employed to reverse HHcy induced changes. CBS+/- mice exhibited more fatigability, and generated less contraction force. No significant changes in muscle morphology were observed. However, there is a corresponding reduction in large muscle fiber number in CBS+/- mice. Excess fatigability was not due to changes in key enzymes involved in metabolism, but was due to reduced ATP levels. A marginal reduction in dystrophin levels along with a decrease in mitochondrial transcription factor A (mtTFA) were observed. There was also an increase in the mir-31, and mir-494 quantities that were implicated in dystrophin and mtTFA regulation respectively. The molecular changes elevated during HHcy, with the exception of dystrophin levels, were reversed after exercise. In addition, the amount of NRF-1, one of the transcriptional regulators of mtTFA, was significantly decreased. Furthermore, there was enhancement in mir-494 levels and a concomitant decline in mtTFA protein quantity in homocysteine treated cells. These changes in C2C12 cells were also accompanied by an increase in DNMT3a and DNMT3b proteins and global DNA methylation levels. Together, these results suggest that HHcy plays a causal role in enhanced fatigability through mitochondrial dysfunction which involves epigenetic changes.

Alterations in P-Glycoprotein Expression and Function Between Macrophage Subsets.

PURPOSE: Macrophages are an important cellular reservoir in HIV, and exist in two phenotypically dissimilar subsets, the pro-inflammatory M1 phenotype, and the anti-inflammatory M2 phenotype. The role of these two subsets is uncertain. We hypothesized that differences in drug efflux transporters exist between the subsets, which would result in altered intracellular drug concentrations between these cells. METHODS: U937 monocytic cells were polarized to the M1 or M2 phenotype via treatment with interferon-gamma and LPS, or interleukins 4, 13, and LPS, respectively. PGP function was assessed with Hoechst 33342, and expression via western blotting. Intracellular lopinavir was assessed via LC-MS/MS. Data was confirmed with primary monocyte derived macrophages. RESULTS: We observed significant differences in intracellular concentrations of lopinavir, a PGP substrate, with higher concentrations in M1 cells. PGP function and expression was higher in the M2 macrophages. These results were confirmed with primary monocyte derived macrophages. CONCLUSIONS: This data shows that there are previously unreported differences in P-glycoprotein expression between macrophage subsets, and suggests that there may be differences for other transporters. These differences can play a role in intracellular drug concentrations in these cells, and may allow for low-level HIV replication.

Homocysteine elicits an M1 phenotype in murine macrophages through an EMMPRIN-mediated pathway.

INTRODUCTION: Hyperhomocysteinemia (HHcy) is associated with inflammatory diseases and is known to increase the production of reactive oxygen species (ROS), matrix metalloproteinase (MMP)-9, and inducible nitric oxide synthase, and to decrease endothelial nitric oxide production. However, the impact of HHcy on macrophage phenotype differentiation is not well-established. It has been documented that macrophages have 2 distinct phenotypes: the "classically activated/destructive" (M1), and the "alternatively activated/constructive" (M2) subtypes. We hypothesize that HHcy increases M1 macrophage differentiation through extracellular matrix metalloproteinase inducer (EMMPRIN), a known inducer of matrix metalloproteinases. METHODS: murine J774A.1 and Raw 264.7 macrophages were treated with 100 and 500 µmol/L Hcy, respectively, for 24 h. Samples were analyzed using Western blotting and immunocytochemistry. RESULTS: Homocysteine treatment increased cluster of differentiation 40 (CD40; M1 marker) in J774A.1 and Raw 264.7 macrophages. MMP-9 was induced in both cell lines. EMMPRIN protein expression was also increased in both cell lines. Blocking EMMPRIN function by pre-treating cells with anti-EMMPRIN antibody, with or without Hcy, resulted in significantly lower expression of CD40 in both cell lines by comparison with the controls. A DCFDA assay demonstrated increased ROS production in both cell lines with Hcy treatment when compared with the controls. CONCLUSION: Our results suggest that HHcy results in an increase of the M1 macrophage phenotype. This effect seems to be at least partially mediated by EMMPRIN induction.

Hyperhomocysteinemia: a missing link to dysfunctional HDL via paraoxanase-1.

Paraoxanase-1 (PON1) is an HDL-associated enzyme that contributes to the antioxidant and antiatherosclerotic properties of HDL. Lack of PON1 results in dysfunctional HDL. HHcy is a risk factor for cardiovascular disorders, and instigates vascular dysfunction and ECM remodeling. Although studies have reported HHcy during atherosclerosis, the exact mechanism is unclear. Here, we hypothesize that dysfunctional HDL due to lack of PON1 contributes to endothelial impairment and atherogenesis through HHcy-induced ECM re-modeling. To verify this hypothesis, we used C57BL6/J and PON1 knockout mice (KO) and fed them an atherogenic diet. The expression of Akt, ADMA, and DDAH, as well as endothelial gap junction proteins such as Cx-37 and Cx-40 and eNOS was measured for vascular dysfunction and inflammation. We observed that cardiac function was decreased and plasma Hcy levels were increased in PON1 KO mice fed the atherogenic diet compared with the controls. Expression of Akt, eNOS, DDAH, Cx-37, and Cx-40 was decreased, and the expression of MMP-9 and ADMA was increased in PON1 KO mice fed an atherogenic diet compared with the controls. Our results suggest that HHcy plays an intricate role in dysfunctional HDL, owing to the lack of PON1. This contributes to vascular endothelial impairment and atherosclerosis through MMP-9-induced vascular remodeling.

Quantification of cell-associated atazanavir, darunavir, lopinavir, ritonavir, and efavirenz concentrations in human mononuclear cell extracts.

An ultrasensitive assay utilizing high-pressure liquid chromatography and mass spectrometry detection was developed and validated for the quantification of the antiretrovirals atazanavir (ATV), darunavir (DRV), lopinavir (LPV), ritonavir (RTV), and efavirenz (EFV) in human mononuclear cell (MNC) extracts. The assay utilizes 20 µl of cellular extract that contains as few as 50,000 MNCs. The analytical range of the assay is 0.0200 to 10.0 fmol/µl for ATV, 0.0500 to 25.0 fmol/µl for DRV, LPV, and RTV, and 0.200 to 100 fmol/µl for EFV. The assay has proven to be a clinically useful tool for investigating antiretroviral drug concentrations in virologic sanctuaries where harvested cell numbers are extremely low. The assay provides a tool for investigators to explore the clinical pharmacology of strategies for prevention, treatment, and cure in pathophysiologically relevant sites.

Exercise mitigates the adverse effects of hyperhomocysteinemia on macrophages, MMP-9, skeletal muscle, and white adipocytes.

Regular exercise is a great medicine with its benefits encompassing everything from prevention of cardiovascular risk to alleviation of different muscular myopathies. Interestingly, elevated levels of homocysteine (Hcy), also known as hyperhomocysteinemia (HHcy), antagonizes beta-2 adrenergic receptors (ß2AR), gamma amino butyric acid (GABA), and peroxisome proliferator-activated receptor-gamma (PPARγ) receptors. HHcy also stimulates an elevation of the M1/M2 macrophage ratio, resulting in a more inflammatory profile. In this review we discuss several potential targets altered by HHcy that result in myopathy and excessive fat accumulation. Several of these HHcy mediated changes can be countered by exercise and culminate into mitigation of HHcy induced myopathy and metabolic syndrome. We suggest that exercise directly impacts levels of Hcy, matrix metalloproteinase 9 (MMP-9), macrophages, and G-protein coupled receptors (GPCRs, especially Gs). While HHcy promotes the M1 macrophage phenotype, it appears that exercise may diminish the M1/M2 ratio, resulting in a less inflammatory phenotype. HHcy through its influence on GPCRs, specifically ß2AR, PPARγ and GABA receptors, promotes accumulation of white fat, whereas exercise enhances the browning of white fat and counters HHcy-mediated effects on GPCRs. Alleviation of HHcy-associated pathologies with exercise also includes reversal of excessive MMP-9 activation. Moreover, exercise, by reducing plasma Hcy levels, may prevent skeletal muscle myopathy, improve exercise capacity and rescue the obese phenotype. The purpose of this review is to summarize the pathological conditions surrounding HHcy and to clarify the importance of regular exercise as a method of disease prevention.

Lower limb blood flow occlusion increases systemic pressor response without increasing brachial arterial blood flow redistribution in women.

This study was conducted to investigate the systemic hemodynamic and vascular changes in women during and after two commonly used clinical blood flow restriction (BFR) pressures at rest. There are minimal data regarding the independent effects of BFR on hemodynamic and systemic vascular changes due to pressor response, particularly among women. Therefore, this study investigated BFR-induced alterations in pressor response and systemic flow redistribution at rest during two commonly used pressures (50% and 80% limb occlusion pressure [LOP]). Fifteen women (22.1 ± 4.2 years) completed two randomised sessions involving 8-min of bilateral, lower limb restriction at 50% or 80% LOP followed by 8-min of recovery post-deflation. Changes in vascular (arterial diameter [DIA], time-averaged mean velocity [TAMV], volume flow [VF], and area) and hemodynamic (heart rate [HR] and blood pressure) measures over time (pre-, during, post-occlusion) and by session (50% vs. 80% LOP) were tested using repeated measures analysis of variance. Repeated measures correlations (rrm) quantified common intraindividual associations between BFR-induced hemodynamic and vascular responses. HR increased from baseline during 50% LOP and remained elevated during recovery (p < 0.05). HR increased from baseline during 80% LOP, while tibial VF and TAMV decreased (p < 0.03 for all). HR and TAMV values returned to baseline during recovery, while brachial artery VF decreased (p < 0.05). Changes in HR, brachial VF, and brachial TAMV were similar between 50% and 80% LOP (rrm = 0.32-0.70, p < 0.05 for all). At 80% LOP, changes in HR were positively correlated with brachial VF (rrm = 0.38) and TAMV (rrm = 0.43) and negatively correlated with tibial VF (rrm = -0.36) and TAMV (rrm = -0.30) (p < 0.05 for all). Results suggest that BFR at 80% LOP elicits an acute systemic pressor reflex without concomitant increases in brachial arterial flow, while 50% LOP elicits a subdued response.

Body composition estimates using a 2D image analysis system across different environmental conditions: An agreement study.

PURPOSE: This study examined the agreement between %Fat measurements using a smartphone-based application (IMAGE) across different environmental conditions. METHODS: A single reference image was obtained using an 8 MP smartphone camera under Ambient Light in front of a white background. Additional photos were obtained using a 0.7 MP, 5 MP, and 12 MP smartphone cameras; low-, moderate-, and bright-lighting conditions; and various color backgrounds including black, green, orange, and gray. RESULTS: %Fat measured using the 0.7 MP camera (27.8 ± 6.2 %Fat) was higher than the reference (26.8 ± 6.1 %Fat) (p < 0.001). The black (32.0 ± 12.0 %Fat), green (27.5 ± 6.3 %Fat), and gray (27.8 ± 6.3 %Fat) backgrounds yielded higher %Fat than the white (p = 0.03, 0.01, and 0.001). All camera, lighting, and background conditions were strongly correlated with the reference (all intraclass correlation coefficient [ICC] >0.98, all standard error of the estimate [SEE] <1.5 %Fat, all p < 0.001), except the black background which yielded poorer agreement with the white background (ICC = 0.69, SEE = 4.5%, p < 0.001). CONCLUSION: %Fat from IMAGE were strongly correlated across various environmental conditions.