Tag: monocytes

Single-cell transcriptomics of the immune system in ME/CFS at baseline and following symptom provocation

Summary:

ME/CFS is a serious and poorly understood disease. To understand immune dysregulation in ME/CFS, we used single-cell RNA-seq (scRNA-seq) to examine immune cells in cohorts of patients and controls. Post-exertional malaise (PEM), an exacerbation of symptoms following strenuous exercise, is a characteristic symptom of ME/CFS. Thus, to detect changes coincident with PEM, we also performed scRNA-seq on the same cohorts following exercise. At baseline, ME/CFS patients displayed dysregulation of classical monocytes suggestive of inappropriate differentiation and migration to tissue. We were able to identify both diseased and more normal monocytes within patients, and the fraction of diseased cells correlated with metrics of disease severity. Comparing the transcriptome at baseline and post-exercise challenge, we discovered patterns indicative of improper platelet activation in patients, with minimal changes elsewhere in the immune system. Taken together, these data identify immunological defects present at baseline in patients and an additional layer of dysregulation following exercise.

Highlights ME/CFS is a debilitating disease with unknown causes. Here, we provide, for the first time, an extensive single cell resolution dataset detailing the gene expression programs of circulating immune cells of ME/CFS cases at baseline and after symptom provocation. We were able to detect robust dysregulation in certain immune cells from patients, with dysregulation of classical monocytes manifesting the strongest signal. Indeed, the fraction of aberrant monocytes in ME/CFS patients correlated with the degree of disease severity. Surprisingly, platelet transcriptomes were also altered in ME/CFS, and they were the only component of the immune system that showed large-scale changes following symptom provocation.

Source: Faraz Ahmed, Luyen Tien Vu, Hongya Zhu, David Shing Huk Iu, Elizabeth A. Fogarty, Yeonui Kwak, Weizhong Chen, Carl J. Franconi, Paul R. Munn, Susan M. Levine, Jared Stevens, Xiangling Mao, Dikoma C. Shungu, Geoffrey E. Moore, Betsy A. Keller, Maureen R. Hanson, Jennifer K. Grenier, Andrew Grimson. Single-cell transcriptomics of the immune system in ME/CFS at baseline and following symptom provocation. bioRxiv 2022.10.13.512091; doi: https://doi.org/10.1101/2022.10.13.512091 https://www.cell.com/cell-reports-medicine/fulltext/S2666-3791(23)00602-X (Full text)

Deep phenotyping of myalgic encephalomyelitis/chronic fatigue syndrome in Japanese population

Abstract:

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex and debilitating disease with no molecular diagnostics and no treatment options. To identify potential markers of this illness, we profiled 48 patients and 52 controls for standard laboratory tests, plasma metabolomics, blood immuno-phenotyping and transcriptomics, and fecal microbiome analysis. Here, we identified a set of 26 potential molecular markers that distinguished ME/CFS patients from healthy controls. Monocyte number, microbiome abundance, and lipoprotein profiles appeared to be the most informative markers.

When we correlated these molecular changes to sleep and cognitive measurements of fatigue, we found that lipoprotein and microbiome profiles most closely correlated with sleep disruption while a different set of markers correlated with a cognitive parameter. Sleep, lipoprotein, and microbiome changes occur early during the course of illness suggesting that these markers can be examined in a larger cohort for potential biomarker application. Our study points to a cluster of sleep-related molecular changes as a prominent feature of ME/CFS in our Japanese cohort.

Source: Kitami T, Fukuda S, Kato T, Yamaguti K, Nakatomi Y, Yamano E, Kataoka Y, Mizuno K, Tsuboi Y, Kogo Y, Suzuki H, Itoh M, Morioka MS, Kawaji H, Koseki H, Kikuchi J, Hayashizaki Y, Ohno H, Kuratsune H, Watanabe Y. Deep phenotyping of myalgic encephalomyelitis/chronic fatigue syndrome in Japanese population. Sci Rep. 2020 Nov 16;10(1):19933. doi: 10.1038/s41598-020-77105-y. PMID: 33199820; PMCID: PMC7669873.  https://www.nature.com/articles/s41598-020-77105-y (Full text)

Inclusion of Family Members Without ME/CFS in Research Studies Promotes Discovery of Biomarkers Specific for ME/CFS

Abstract:

Background: The search for a biomarker specific for ME/CFS (myalgic encephalomyelitis/chronic fatigue syndrome) has been long, arduous and, to date, unsuccessful. Researchers need to consider their expenditures on each new candidate biomarker. In a previous study of antibody-dependent cell-mediated cytotoxicity (ADCC) by natural killer lymphocytes, we found lower ADCC for ME/CFS patients vs. unrelated donors but ruled against low ADCC as a biomarker because of similar ADCC for patients vs. their family members without ME/CFS.

Objective: We applied inclusion of family members without ME/CFS, from families with multiple CFS patients, as a second non-ME/CFS control group in order to re-examine inflammation in ME/CFS.

Method: Total and CD16A-positive ‘non-classical’ anti-inflammatory monocytes were monitored.

Results: Non-classical monocytes were elevated for patients vs. unrelated healthy donors but these differences were insignificant between patients vs. unaffected family members.

Conclusions: Inclusion of family members ruled against biomarker considerations for the monocytes characterized. These pilot findings for the non-classical monocytes are novel in the field of ME/CFS. We recommend that occupational therapists advocate and explain to family members without ME/CFS the need for the family members’ participation as a second set of controls in pilot studies to rapidly eliminate false biomarkers, optimize patient participation, and save researchers’ labor.

Source: Tokunaga K, Sung AP, Tang JJ, et al. Inclusion of family members without ME/CFS in research studies promotes discovery of biomarkers specific for ME/CFS [published online ahead of print, 2020 Jun 16]. Work. 2020;10.3233/WOR-203177. doi:10.3233/WOR-203177 https://pubmed.ncbi.nlm.nih.gov/32568152/

MicroRNAs hsa-miR-99b, hsa-miR-330, hsa-miR-126 and hsa-miR-30c: Potential Diagnostic Biomarkers in Natural Killer (NK) Cells of Patients with Chronic Fatigue Syndrome (CFS)/ Myalgic Encephalomyelitis (ME

Abstract:

BACKGROUND: Chronic Fatigue Syndrome (CFS/ME) is a complex multisystem disease of unknown aetiology which causes debilitating symptoms in up to 1% of the global population. Although a large cohort of genes have been shown to exhibit altered expression in CFS/ME patients, it is currently unknown whether microRNA (miRNA) molecules which regulate gene translation contribute to disease pathogenesis. We hypothesized that changes in microRNA expression in patient leukocytes contribute to CFS/ME pathology, and may therefore represent useful diagnostic biomarkers that can be detected in the peripheral blood of CFS/ME patients.

METHODS: miRNA expression in peripheral blood mononuclear cells (PBMC) from CFS/ME patients and healthy controls was analysed using the Ambion Bioarray V1. miRNA demonstrating differential expression were validated by qRT-PCR and then replicated in fractionated blood leukocyte subsets from an independent patient cohort. The CFS/ME associated miRNA identified by these experiments were then transfected into primary NK cells and gene expression analyses conducted to identify their gene targets.

RESULTS: Microarray analysis identified differential expression of 34 miRNA, all of which were up-regulated. Four of the 34 miRNA had confirmed expression changes by qRT-PCR. Fractionating PBMC samples by cell type from an independent patient cohort identified changes in miRNA expression in NK-cells, B-cells and monocytes with the most significant abnormalities occurring in NK cells. Transfecting primary NK cells with hsa-miR-99b or hsa-miR-330-3p, resulted in gene expression changes consistent with NK cell activation but diminished cytotoxicity, suggesting that defective NK cell function contributes to CFS/ME pathology.

CONCLUSION: This study demonstrates altered microRNA expression in the peripheral blood mononuclear cells of CFS/ME patients, which are potential diagnostic biomarkers. The greatest degree of miRNA deregulation was identified in NK cells with targets consistent with cellular activation and altered effector function.

 

Source: Petty RD, McCarthy NE, Le Dieu R, Kerr JR. MicroRNAs hsa-miR-99b, hsa-miR-330, hsa-miR-126 and hsa-miR-30c: Potential Diagnostic Biomarkers in Natural Killer (NK) Cells of Patients with Chronic Fatigue Syndrome (CFS)/ Myalgic Encephalomyelitis (ME). PLoS One. 2016 Mar 11;11(3):e0150904. doi: 10.1371/journal.pone.0150904. ECollection 2016. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4788442/ (Full article)

 

Increased production of interleukin-6 by adherent and non-adherent mononuclear cells during ‘natural fatigue’ but not following ‘experimental fatigue’ in patients with chronic fatigue syndrome

Abstract:

In an investigator-blinded study, adherent (monocytes) and non-adherent cells (lymphocytes) from patients with chronic fatigue syndrome (CFS) were examined on two separate occasions (when feeling ‘fatigued’ and when feeling ‘rested’) for in vitro spontaneous, phytohemagglutinin- (PHA, for lymphocytes), and lipopolysaccharide- (LPS, for monocytes) induced production of IL-6 by ELISA assay.

A group of CFS patients and controls were also subjected to exercise-induced fatigue (‘experimental fatigue’) and IL-6 production was compared, in a double-blinded manner, prior to and following induction of fatigue.

A significant increase in spontaneous, PHA- and LPS-induced IL-6 secretion by both lymphocytes and monocytes was observed in CFS patients during ‘natural fatigue’ as compared to during state. However, no such changes in IL-6 production were observed during ‘experimental fatigue’.

These data suggest a role of IL-6 in natural symptomatology and perhaps in the pathogenesis of CFS. In addition, the data demonstrate that laboratory-induced fatigue (experimental fatigue) may not be a good model to study immunological changes in CFS; immunological parameters should be studied in a longitudinal manner during the natural course of the disease.

 

Source: Gupta S, Aggarwal S, Starr A. Increased production of interleukin-6 by adherent and non-adherent mononuclear cells during ‘natural fatigue’ but not following ‘experimental fatigue’ in patients with chronic fatigue syndrome. Int J Mol Med. 1999 Feb;3(2):209-13. http://www.ncbi.nlm.nih.gov/pubmed/9917531

 

Cytokine production by adherent and non-adherent mononuclear cells in chronic fatigue syndrome

Abstract:

It has been suggested that cytokines play a role in certain clinical manifestations of chronic fatigue syndrome (CFS). In this study adherent (monocytes) and non-adherent (lymphocytes) mononuclear cells were stimulated in the presence or absence of phytohemagglutinin (PHA) or lipopolysaccharide (LPS), respectively, and supernatants were assayed for IL-6, TNF-alpha, and IL-10 by ELISA. IL-6 was also measured at the mRNA level by polymerase chain reaction.

The levels of spontaneously (unstimulated) produced TNF-alpha by non-adherent lymphocytes and spontaneously produced IL-6 by both adherent monocytes and non-adherent lymphocytes were significantly increased as compared to simultaneously studied matched controls. The abnormality of IL-6 was also observed at mRNA level.

In contrast, spontaneously produced IL-10 by both adherent and non-adherent cells and by PHA-activated non-adherent cells were decreased. This preliminary study suggests that an aberrant production of cytokines in CFS may play a role in the pathogenesis and in some of the clinical manifestations of CFS.

 

Source: Gupta S, Aggarwal S, See D, Starr A. Cytokine production by adherent and non-adherent mononuclear cells in chronic fatigue syndrome. J Psychiatr Res. 1997 Jan-Feb;31(1):149-56. http://www.ncbi.nlm.nih.gov/pubmed/9201656

 

A comprehensive immunological analysis in chronic fatigue syndrome

Abstract:

A detailed analysis of cell-mediated and antibody-mediated immunity was performed in 20 CDC-defined patients with chronic fatigue syndrome (CFS) and 20 age- and sex-matched healthy controls.

CD3+, CD4+, CD8+, and CD20+ lymphocytes were comparable in two groups. Natural killer cells as defined by CD16, CD56 and CD57 antigens were significantly reduced in CFS. A significant increase in the proportions of CD4+ ICAM 1+ T cells was observed in CFS. Monocytes from CFS displayed increased density (as determined by mean fluorescence channel numbers) of intercellular adhesion molecule 1 (ICAM-1) and lymphocyte function associated antigen 1 (LFA-1), but showed decreased enhancing response to recombinant interferon-gamma in vitro.

The lymphocyte DNA synthesis in response to phytohaemoglobulin (PHA), Concanavalin A (Con A) and pokeweed mitogen (PWM) was normal but the response to soluble antigens was significantly reduced. Serum IgM, IgG, IgA, and IgG subclasses were normal. In vivo specific antibody response to pneumococcus vaccine was depressed in CFS.

Forty percent of patients showed titres of anti-human herpes virus 6 (anti-HHV-6) antibody higher than that in the controls (greater than or equal to 1/80). These data suggest immunological dysfunction in patients with chronic fatigue syndrome. The significance of these observations is discussed.

 

Source: Gupta S, Vayuvegula B. A comprehensive immunological analysis in chronic fatigue syndrome. Scand J Immunol. 1991 Mar;33(3):319-27. http://www.ncbi.nlm.nih.gov/pubmed/1849315

 

Naloxone-reversible monocyte dysfunction in patients with chronic fatigue syndrome

Abstract:

We studied monocyte function in 35 consecutive patients with chronic fatigue syndrome (CFS) and 25 healthy controls. Eighty-five per cent of the patients showed monocyte dysfunction characterized by marked reduction in the number of monocytes displaying immunoreactive cytoskeletal vimentin filaments, a low phagocytosis index, and a reduced expression of HLA-DR antigens. These values increased dramatically after incubation of the patients’ monocytes with the opioid antagonist naloxone.

Other immunological abnormalities also noted in the patients were low lymphocyte blastogenesis and diminished numbers of monocytes displaying receptors for Fc of IgG (FcR) and C3b (CR1). These findings suggest that an increased opioid activity acting through a classical receptor mechanism is active on monocytes from a high proportion of patients with CFS and that this represents a novel example of immunomodulation by opioid peptides in human disease.

We suggest that endogenous opioids are involved in the pathogenesis of the chronic fatigue syndrome.

 

Source: Prieto J, Subirá ML, Castilla A, Serrano M. Naloxone-reversible monocyte dysfunction in patients with chronic fatigue syndrome. Scand J Immunol. 1989 Jul;30(1):13-20. http://www.ncbi.nlm.nih.gov/pubmed/2526966