Extracellular flux analyses reveal differences in mitochondrial PBMC metabolism between high-fit and low-fit females

Analyzing metabolism of peripheral blood mononuclear cells (PBMCs) can possibly serve as a cellular metabolic read-out for lifestyle factors and lifestyle interventions. However, the impact of PBMC composition on PBMC metabolism is not yet clear, neither is the differential impact of a longer-term lifestyle factor versus a short-term lifestyle intervention. We investigated the effect of aerobic fitness level and a recent exercise bout on PBMC metabolism in females. PBMCs from 31 young female adults divided into a high-fit (V̇O2peak ≥ 47 mL/kg/min, N = 15) and low-fit (V̇O2peak ≤ 37 mL/kg/min, N = 16) group were isolated at baseline and overnight after a single bout of exercise (60 minutes, 70% V̇O2peak). Oxygen consumption rate (OCR) and glycolytic rate (GR) were measured using extracellular flux (XF) assays and PBMC subsets were characterized using fluorescence-activated cell sorting (FACS). Basal OCR, FCCP-induced OCR, spare respiratory capacity, ATP-linked OCR, and proton leak were significantly higher in high-fit compared to low-fit females (all P < 0.01), while no significant differences in glycolytic rate (GR) were found (all P > 0.05). A recent exercise bout did not significantly affect GR or OCR parameters (all P > 0.05). The overall PBMC composition was similar between high-fit and low-fit females. Mitochondrial PBMC function was significantly higher in PBMCs from high-fit compared to low-fit females, which was unrelated to PBMC composition and not impacted by a recent bout of exercise. Our study reveals a link between PBMC metabolism and levels of aerobic fitness, increasing the relevance of PBMC metabolism as a marker to study the impact of lifestyle factors on human health.

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Novel standardized method for extracellular flux analysis of oxidative and glycolytic metabolism in peripheral blood mononuclear cells

Scientific Reports ◽

10.1038/s41598-021-81217-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Joëlle J. E. Janssen ◽

Bart Lagerwaard ◽

Annelies Bunschoten ◽

Huub F. J. Savelkoul ◽

R. J. Joost van Neerven ◽

...

Keyword(s):

Oxygen Consumption ◽

Oxygen Consumption Rate ◽

Consumption Rate ◽

Mononuclear Cells ◽

Flux Analysis ◽

Standard Score ◽

Peripheral Blood Mononuclear ◽

Extracellular Flux ◽

Blood Mononuclear Cells ◽

Glycolytic Rate

AbstractAnalyzing metabolism of peripheral blood mononuclear cells (PBMCs) provides key opportunities to study the pathophysiology of several diseases, such as type 2 diabetes, obesity and cancer. Extracellular flux (XF) assays provide dynamic metabolic analysis of living cells that can capture ex vivo cellular metabolic responses to biological stressors. To obtain reliable data from PBMCs from individuals, novel methods are needed that allow for standardization and take into account the non-adherent and highly dynamic nature of PBMCs. We developed a novel method for extracellular flux analysis of PBMCs, where we combined brightfield imaging with metabolic flux analysis and data integration in R. Multiple buffy coat donors were used to demonstrate assay linearity with low levels of variation. Our method allowed for accurate and precise estimation of XF assay parameters by reducing the standard score and standard score interquartile range of PBMC basal oxygen consumption rate and glycolytic rate. We applied our method to freshly isolated PBMCs from sixteen healthy subjects and demonstrated that our method reduced the coefficient of variation in group mean basal oxygen consumption rate and basal glycolytic rate, thereby decreasing the variation between PBMC donors. Our novel brightfield image procedure is a robust, sensitive and practical normalization method to reliably measure, compare and extrapolate XF assay data using PBMCs, thereby increasing the relevance for PBMCs as marker tissue in future clinical and biological studies, and enabling the use of primary blood cells instead of immortalized cell lines for immunometabolic experiments.

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The impact of 17β-estradiol and progesterone therapy on peripheral blood mononuclear cells of asthmatic patients

Molecular Biology Reports ◽

10.1007/s11033-020-06046-6 ◽

2020 ◽

Author(s):

Leila Nejatbakhsh Samimi ◽

Morteza Fallahpour ◽

Majid Khoshmirsafa ◽

Seyed Ali Javad Moosavi ◽

Paria Bayati ◽

...

Keyword(s):

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Peripheral Blood Mononuclear ◽

Asthmatic Patients ◽

Blood Mononuclear Cells ◽

17Β Estradiol ◽

The Impact

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Regulation of CTLA-4 and PD-L1 Expression in Relapsing-Remitting Multiple Sclerosis Patients after Treatment with Fingolimod, IFNβ-1α, Glatiramer Acetate, and Dimethyl Fumarate Drugs

Journal of Personalized Medicine ◽

10.3390/jpm11080721 ◽

2021 ◽

Vol 11 (8) ◽

pp. 721

Author(s):

Afshin Derakhshani ◽

Zahra Asadzadeh ◽

Hossein Safarpour ◽

Patrizia Leone ◽

Mahdi Abdoli Shadbad ◽

...

Keyword(s):

Multiple Sclerosis ◽

Mononuclear Cells ◽

Demyelinating Disease ◽

Immune Checkpoints ◽

Peripheral Blood Mononuclear ◽

Relapsing Remitting ◽

Remitting Multiple Sclerosis ◽

Multiple Sclerosis Patients ◽

Relapsing Remitting Multiple Sclerosis ◽

The Impact

Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS) that is characterized by inflammation which typically results in significant impairment in most patients. Immune checkpoints act as co-stimulatory and co-inhibitory molecules and play a fundamental role in keeping the equilibrium of the immune system. Cytotoxic T-lymphocyte antigen-4 (CTLA-4) and Programmed death-ligand 1 (PD-L1), as inhibitory immune checkpoints, participate in terminating the development of numerous autoimmune diseases, including MS. We assessed the CTLA-4 and PD-L1 gene expression in the different cell types of peripheral blood mononuclear cells of MS patients using single-cell RNA-seq data. Additionally, this study outlines how CTLA-4 and PD-L1 expression was altered in the PBMC samples of relapsing-remitting multiple sclerosis (RRMS) patients compared to the healthy group. Finally, it investigates the impact of various MS-related treatments in the CTLA-4 and PD-L1 expression to restrain autoreactive T cells and stop the development of MS autoimmunity.

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Understanding the influence of socioeconomic status on the association between combinations of lifestyle factors and adverse health outcomes: a systematic review protocol

BMJ Open ◽

10.1136/bmjopen-2020-042212 ◽

2021 ◽

Vol 11 (5) ◽

pp. e042212

Author(s):

Hamish Foster ◽

Peter Polz ◽

Frances Mair ◽

Jason Gill ◽

Catherine A O'Donnell

Keyword(s):

Systematic Review ◽

Socioeconomic Status ◽

Health Outcomes ◽

Meta Analysis ◽

Lifestyle Factors ◽

Lifestyle Factor ◽

Unhealthy Lifestyle ◽

Adverse Health ◽

Adverse Health Outcomes ◽

The Impact

IntroductionCombinations of unhealthy lifestyle factors are strongly associated with mortality, cardiovascular disease (CVD) and cancer. It is unclear how socioeconomic status (SES) affects those associations. Lower SES groups may be disproportionately vulnerable to the effects of unhealthy lifestyle factors compared with higher SES groups via interactions with other factors associated with low SES (eg, stress) or via accelerated biological ageing. This systematic review aims to synthesise studies that examine how SES moderates the association between lifestyle factor combinations and adverse health outcomes. Greater understanding of how lifestyle risk varies across socioeconomic spectra could reduce adverse health by (1) identifying novel high-risk groups or targets for future interventions and (2) informing research, policy and interventions that aim to support healthy lifestyles in socioeconomically deprived communities.Methods and analysisThree databases will be searched (PubMed, EMBASE, CINAHL) from inception to March 2020. Reference lists, citations and grey literature will also be searched. Inclusion criteria are: (1) prospective cohort studies; (2) investigations of two key exposures: (a) lifestyle factor combinations of at least three lifestyle factors (eg, smoking, physical activity and diet) and (b) SES (eg, income, education or poverty index); (3) an assessment of the impact of SES on the association between combinations of unhealthy lifestyle factors and health outcomes; (4) at least one outcome from—mortality (all cause, CVD and cancer), CVD or cancer incidence. Two independent reviewers will screen titles, abstracts and full texts of included studies. Data extraction will focus on cohort characteristics, exposures, direction and magnitude of SES effects, methods and quality (via Newcastle-Ottawa Scale). If appropriate, a meta-analysis, pooling the effects of SES, will be performed. Alternatively, a synthesis without meta-analysis will be conducted.Ethics and disseminationEthical approval is not required. Results will be disseminated via peer-reviewed publication, professional networks, social media and conference presentations.PROSPERO registration numberCRD42020172588.

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009 Self-Reported Sleep Efficiency and Duration are Associated with Systemic Bioenergetic Function in Community-Dwelling Adults

SLEEP ◽

10.1093/sleep/zsab072.008 ◽

2021 ◽

Vol 44 (Supplement_2) ◽

pp. A4-A4

Author(s):

H Matthew Lehrer ◽

Lauren Chu ◽

Martica Hall ◽

Kyle Murdock

Keyword(s):

Sleep Duration ◽

Mononuclear Cells ◽

Basic Research ◽

Basal Respiration ◽

Community Dwelling ◽

Sleep Efficiency ◽

Flux Analysis ◽

Spare Capacity ◽

Peripheral Blood Mononuclear ◽

Extracellular Flux

Abstract Introduction Sleep is important for aging, health, and disease, but its cellular role in these outcomes is poorly understood. Basic research suggests that disturbed and insufficient sleep impair mitochondrial bioenergetics, which is involved in numerous aging-related chronic conditions. However, the relationship between sleep and bioenergetics has not been examined in humans. We examined associations of self-reported sleep with systemic bioenergetic function in peripheral blood mononuclear cells (PBMCs) of community-dwelling adults. Methods N = 43 adults (79% female) ages 48–70 (M = 61.63, SD = 5.99) completed the Pittsburgh Sleep Quality Index (PSQI) from which key components of sleep (satisfaction, alertness, timing, efficiency, and duration) were calculated. Participants provided blood samples from which PBMCs were isolated and measured for bioenergetics using extracellular flux analysis. Associations of sleep components with bioenergetic parameters, including the Bioenergetic Health Index (BHI), were examined. Results In bivariate analyses, lower sleep efficiency was associated with lower maximal respiration, spare capacity, and BHI (ps < 0.05). Longer sleep duration was associated with lower BHI (p < 0.01) and later sleep timing was associated with higher basal respiration, ATP-linked respiration, maximal respiration, spare capacity, and non-mitochondrial respiration (ps < 0.05). After adjustment for age, sex, and body mass index, lower sleep efficiency (β = 0.52, p < 0.01) and longer sleep duration (β = -0.43, p < 0.01) were associated with lower BHI. Conclusion Self-reported indices of sleep efficiency and duration are related to systemic bioenergetic function in humans, suggesting a possible cellular pathway linking sleep to health. Support (if any) T32HL082610

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Carfilzomib can induce tumor cell death through selective inhibition of the chymotrypsin-like activity of the proteasome

Blood ◽

10.1182/blood-2009-05-223677 ◽

2009 ◽

Vol 114 (16) ◽

pp. 3439-3447 ◽

Cited By ~ 229

Author(s):

Francesco Parlati ◽

Susan J. Lee ◽

Monette Aujay ◽

Erika Suzuki ◽

Konstantin Levitsky ◽

...

Keyword(s):

Tumor Cells ◽

Mononuclear Cells ◽

Proteasome Inhibitors ◽

Selective Inhibition ◽

Tumor Cell Death ◽

Clinical Safety ◽

Peripheral Blood Mononuclear ◽

Non Hodgkin Lymphoma ◽

Proteasome Subunits ◽

The Impact

Abstract Carfilzomib is a proteasome inhibitor in clinical development that primarily targets the chymotrypsin-like (CT-L) subunits in both the constitutive proteasome (c20S) and the immunoproteasome (i20S). To investigate the impact of inhibiting the CT-L activity with carfilzomib, we set out to quantitate the levels of CT-L subunits β5 from the c20S and LMP7 from the i20S in normal and malignant hematopoietic cells. We found that the i20S is a major form of the proteasome expressed in cells of hematopoietic origin, including multiple myeloma (MM) CD138+ tumor cells. Although specific inhibition of either LMP7 or β5 alone was insufficient to produce an antitumor response, inhibition of all proteasome subunits was cytotoxic to both hematologic tumor cells and peripheral blood mononuclear cells. However, selective inhibition of both β5 and LMP7 was sufficient to induce an antitumor effect in MM, non-Hodgkin lymphoma, and leukemia cells while minimizing the toxicity toward nontransformed cells. In MM tumor cells, CT-L inhibition alone was sufficient to induce proapoptotic sequelae, including proteasome substrate accumulation, Noxa and caspase 3/7 induction, and phospho-eIF2α suppression. These data support a hypothesis that hematologic tumor cells are uniquely sensitive to CT-L inhibition and provide a mechanistic understanding of the clinical safety profile and antitumor activity of proteasome inhibitors.

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Integrated microRNA and mRNA Gene Expression in Peripheral Blood Mononuclear Cells in Response to Acute Psychosocial Stress: A Repeated-Measures Within-Subject Pilot Study

10.21203/rs.3.rs-322294/v1 ◽

2021 ◽

Author(s):

Magdalena Maria Jurkiewicz ◽

Anett Müller-Alcazar ◽

Dirk Moser ◽

Indralatha Jayatilaka ◽

Anatoly Mikhailik ◽

...

Keyword(s):

Gene Expression ◽

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Psychosocial Stress ◽

Mononuclear Cells ◽

Peripheral Blood Mononuclear ◽

Time Points ◽

Blood Mononuclear Cells ◽

The Impact ◽

Acute Psychosocial Stress

Abstract Objective: The impact of psychosocial stress on a variety of negative health outcomes is well documented, with current research efforts directed at possible mechanisms. Here, we focused on a potential mechanism involving differential expression of mRNA and microRNA in response to acute psychosocial stress. We utilized a validated behavioral paradigm, the Trier Social Stress Test (TSST), to induce acute psychosocial stress in a cohort of volunteers. Stress reactivity was assessed repeatedly during the TSST using saliva samples that were analyzed for levels of cortisol. Peripheral blood mononuclear cells were extracted from blood drawn at baseline and at two time points following the stress paradigm. Total RNA was extracted, and mRNA and microRNA microarrays were utilized to assess within-subject changes in gene expression between baseline and the two post-stressor time points. Results: For microarray gene expression analysis, we focused on 12 participants who showed a robust cortisol response to the task, as an indicator of robust HPA-axis activation. We discovered a set of mRNAs and miRNAs that exhibited dynamic expression change in response to the TSST in peripheral blood mononuclear cells, further characterizing the link between psychosocial stress and cellular response mechanisms.

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Gene-regulatory network study of rheumatoid arthritis in single-cell chromatin landscapes of peripheral blood mononuclear cells

10.21203/rs.3.rs-899470/v1 ◽

2021 ◽

Author(s):

Cantong Zhang ◽

Xiaoping Hong ◽

Haiyan Yu ◽

Hongwei Wu ◽

Huixuan Xu ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Single Cell ◽

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Peripheral Blood Mononuclear ◽

Blood Mononuclear Cells ◽

High Resolution Map ◽

The Impact ◽

Accessible Chromatin

Abstract Rheumatoid arthritis is a chronic autoinflammatory disease with an elusive etiology. Assays for transposase-accessible chromatin with single-cell sequencing (scATAC-seq) contribute to the progress in epigenetic studies. However, the impact of epigenetic technology on autoimmune diseases has not been objectively analyzed. Therefore, scATAC-seq was performed to generate a high-resolution map of accessible loci in peripheral blood mononuclear cells (PBMCs) of RA patients at the single-cell level. The purpose of our project was to discover the transcription factors (TFs) that were involved in the pathogenesis of RA at single-cell resolution. In our research, we obtained 22 accessible chromatin patterns. Then, 10 key TFs were involved in the RA pathogenesis by regulating the activity of MAP kinase. Consequently, two genes (PTPRC, SPAG9) regulated by 10 key TFs were found that may be associated with RA disease pathogenesis and these TFs were obviously enriched in RA patients (p<0.05, FC>1.2). With further qPCR validation on PTPRC and SPAG9 in monocytes, we found differential expression of these two genes, which were regulated by eight TFs (ZNF384, HNF1B, DMRTA2, MEF2A, NFE2L1, CREB3L4 (var. 2), FOSL2::JUNB (var. 2), MEF2B). What is more, the eight TFs showed highly accessible binding sites in RA patients. These findings demonstrate the value of using scATAC-seq to reveal transcriptional regulatory variation in RA-derived PBMCs, providing insights on therapy from an epigenetic perspective.

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Fecal microbiota transplantation increases colonic IL-25 and dampens tissue inflammation in patients with recurrent Clostridioides difficile

10.1101/2021.07.16.21260643 ◽

2021 ◽

Author(s):

Ning-Jiun Jan ◽

Noah Oakland ◽

Pankaj Kumar ◽

Girija Ramakrishnan ◽

Brian W. Behm ◽

...

Keyword(s):

Mononuclear Cells ◽

Fecal Microbiota Transplantation ◽

Alpha Diversity ◽

The United States ◽

Fecal Microbiota ◽

Peripheral Blood Mononuclear ◽

Clostridioides Difficile ◽

Rdna Sequencing ◽

Clostridioides Difficile Infection ◽

The Impact

Background: Clostridioides difficile infection (CDI) is the most common hospital-acquired infection in the United States. Antibiotic-induced dysbiosis is the primary cause of susceptibility and fecal microbiota transplantation (FMT) has emerged as an effective therapy for recurrence. We previously demonstrated in the mouse model of CDI that antibiotic-induced dysbiosis reduced colonic expression of IL-25, and that FMT protected in part by restoring gut commensal bacteria-mediated IL-25 signaling. Here we conducted a prospective clinical trial to test the impact of FMT on immunity, specifically testing in humans if FMT induced IL-25 expression in the colon. Methods: Subjects received colonic biopsies and blood sampling at the time of FMT and 60-days later. Colon biopsies were assayed for IL-25 by immunoassay, for mRNA by RNAseq, and for bacterial content by 16 S rDNA sequencing. High dimensional flow cytometry was also conducted on peripheral blood mononuclear cells pre- and post-FMT. Results: All 10 subjects who received FMT had no CDI recurrences over a 2 year follow-up post FMT. FMT increased alpha diversity of the colonic microbiota and was associated with several immunologic changes. The cytokine IL-25 was increased in colonic tissue. In addition, increased expression of homeostatic genes and repression of inflammatory genes was observed in colonic mRNA transcripts. Finally, circulating Th17 cells were decreased post-FMT. Conclusion: The increase in the cytokine IL-25 accompanied by decreased inflammation is consistent with FMT acting in part to protect from recurrent CDI via restoration of commensal activation of type 2 immunity.

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