Tag: CD4

Investigating the Genetic and Immunological Aetiology of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome

Abstract:

This thesis describes two investigations into the disease Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS), specifically its genetic aetiology and immune system alterations.

The first study investigated the genetic basis of ME/CFS using Genome-wide Association Studies (GWAS) by attempting to replicate and extend results previously found using UK Biobank cohort data. GWAS attempt to identify associations between DNA variants and phenotypes. T his GWAS was novel, conducted on new phenotypes constructed by combining those in the most up-to-date UK Biobank data release. A new, previously unseen, genome-wide significant association was found on chromosome 6 for males with ME/CFS within the gene PDE10A. Further results were not genome-wide significant, but many were suggestive and hence independent replication may justify further research.

A previous analysis on the UK Biobank cohort had identified an indicative association in females between variants around the SLC25A15 gene at genome-wide significance. I adopted a hypothesis that the dietary protein intake of people with the CFS risk variants would be lower than those with the alternative alleles, due to potentially reduced production of mitochondrial ornithine transporter 1 (ORNT1). However, this association with dietary protein intake was not supported by UK Biobank data.

Additionally, I investigated associations between the human leukocyte antigen (HLA) alleles and the ME/CFS phenotype using UK Biobank data. Associations between alleles within the HLA-C and -DQB1 genes had previously been found in a cohort of Norwegian people with ME/CFS, and my goal was to seek replication of these results in a larger dataset. None of the associations found in the UK Biobank proved to be genome-wide significant.

In my second study I investigated the use of T-cell clonal diversity as a potential biomarker for ME/CFS. This project used cells from CureME Biobank samples in collaboration with Systems Biology Laboratory (SBL). I developed a data analysis pipeline to analyse T-cell receptor (TCR) genomic DNA data based on the best practices currently used in the fields of immunology and mathematical biology. This approach used a mathematical notion of entropy as a measure for the diversity of TCR repertoires, in this way combining all of the most commonly used metrics in mathematical biology. When combined, these measures form a profile for each repertoire, a set of which can be sorted using a machine learning algorithm to partition the repertoires into subgroups.

My hypothesis was that the T-cell clonal expansion of people with ME/CFS would be greater than for healthy controls, and comparable to disease (multiple sclerosis) controls. Although this method was able to effectively classify TCR chains using simulated data, results from experimentally-derived data did not support the hypothesis, with the most effective classifications for both CD4+ and CD8+ cells failing to pass corrections for multiple hypothesis significance testing.

Lay summary

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a disease that affects millions of people around the world. Very little is understood about the cause or progression of the disease, and there is no known cure. At present, there is also no reliable clinical test to determine whether a person has ME/CFS.

This thesis explores the potential for a genetic or immunological basis for ME/CFS, with the goal to eventually find a biomarker that could be used in diagnosis.

The first part of this thesis investigates whether genetic variants are more (or less) common among those with ME/CFS than in the general population. In particular, the region of the genome that encodes immune system proteins was of interest, as previous studies have shown associations between this region and the disease.

Using strict statistical thresholds, none of the previously found associations were replicated. However, one new association was found, with the gene PDE10A, which is implicated in central nervous system diseases, such as Parkinsons and Huntingtons disease. This association has never been seen before, and would require replication in a new cohort before its role in ME/CFS could be confirmed. However, it represents a promising avenue for new research.

The second part of this thesis investigates T-cells. These are highly specialised immune cells in the blood, each of which targets an antigen (foreign substance) such as from a virus. When a T-cell recognises this antigen, it clones itself repeatedly. This clonal expansion is measurable, and can serve as evidence of immune system activation.

My hypothesis was that this immune signature could be used to distinguish people with ME/CFS from healthy controls and others diagnosed with another disease.

I used a mathematical measure of diversity and a machine learning method to sort their immune profiles into groups. However, the pattern of immune activation was not sufficiently clear to provide consistent classification. Hence, the role of the immune system in ME/CFS is still unclear, and the utility of this method as a diagnostic biomarker is not proved.

Source: Joshua James Dibble. Investigating the Genetic and Immunological Aetiology of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome. PhD Thesis [University of Edinburgh]  https://era.ed.ac.uk/bitstream/handle/1842/39763/DibbleJJ_2022.pdf?sequence=1&isAllowed=y (Full text)

A scoping review of regulatory T cell dynamics in convalescent COVID-19 patients – indications for their potential involvement in the development of Long COVID?

Abstract:

Background: Recovery from coronavirus disease 2019 (COVID-19) can be impaired by the persistence of symptoms or new-onset health complications, commonly referred to as Long COVID. In a subset of patients, Long COVID is associated with immune system perturbations of unknown etiology, which could be related to compromised immunoregulatory mechanisms.

Objective: The objective of this scoping review was to summarize the existing literature regarding the frequency and functionality of Tregs in convalescent COVID-19 patients and to explore indications for their potential involvement in the development of Long COVID.

Design: A systematic search of studies investigating Tregs during COVID-19 convalescence was conducted on MEDLINE (via Pubmed) and Web of Science.

Results: The literature search yielded 17 relevant studies, of which three included a distinct cohort of patients with Long COVID. The reviewed studies suggest that the Treg population of COVID-19 patients can reconstitute quantitatively and functionally during recovery. However, the comparison between recovered and seronegative controls revealed that an infection-induced dysregulation of the Treg compartment can be sustained for at least several months. The small number of studies investigating Tregs in Long COVID allowed no firm conclusions to be drawn about their involvement in the syndrome’s etiology. Yet, even almost one year post-infection Long COVID patients exhibit significantly altered proportions of Tregs within the CD4+ T cell population.

Conclusions: Persistent alterations in cell frequency in Long COVID patients indicate that Treg dysregulation might be linked to immune system-associated sequelae. Future studies should aim to address the association of Treg adaptations with different symptom clusters and blood parameters beyond the sole quantification of cell frequencies while adhering to consensualized phenotyping strategies.

Source: Haunhorst S, Bloch W, Javelle F, Krüger K, Baumgart S, Drube S, Lemhöfer C, Reuken P, Stallmach A, Müller M, Zielinski CE, Pletz MW, Gabriel HHW, Puta C. A scoping review of regulatory T cell dynamics in convalescent COVID-19 patients – indications for their potential involvement in the development of Long COVID? Front Immunol. 2022 Dec 13;13:1070994. doi: 10.3389/fimmu.2022.1070994. PMID: 36582234; PMCID: PMC9792979. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9792979/ (Full text)

Study finds differences in energy use by immune cells in ME/CFS

Press Release: New findings published in the Journal of Clinical Investigation suggest that specific immune T cells from people with myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) show disruptions in the way they produce energy. The research was supported by the National Institutes of Health.

“This research gives us additional evidence for the role of the immune system in ME/CFS and may provide important clues to help us understand the mechanisms underlying this devastating disease,” said Vicky Whittemore, Ph.D., program director at NIH’s National Institute of Neurological Disorders and Stroke (NINDS), which partially funded the study.

ME/CFS is a severe, chronic, and debilitating disease that can cause a range of symptoms including pain, severe exhaustion, cognitive impairment, and post-exertional malaise, the worsening of symptoms after physical or mental activity. Estimates suggest that between 836,000 and 2.5 million people in the United States may be affected by ME/CFS. It is unknown what causes the disease and there are no treatments.

Research by Alexandra Mandarano and collaborators in the laboratory of Maureen Hanson, Ph.D., professor of molecular biology and genetics at Cornell University in Ithaca, New York, examined biochemical reactions involved in energy production, or metabolism, in two specific types of immune cells obtained from 45 healthy controls and 53 people with ME/CFS. Investigators focused on CD4 T cells, which alert other immune cells about invading pathogens, and CD8 T cells, which attack infected cells. Dr. Hanson’s team used state-of-the-art methods to look at energy production by the mitochondria within T cells, when the cells were in a resting state and after they had been activated. Mitochondria are biological powerhouses and create most of the energy that drives cells.

Dr. Hanson and her colleagues did not see significant differences in mitochondrial respiration, the cell’s primary energy-producing method, between healthy and ME/CFS cells at rest or after activation. However, results suggest that glycolysis, a less efficient method of energy production, may be disrupted in ME/CFS. Compared to healthy cells, CD4 and CD8 cells from people with ME/CFS had decreased levels of glycolysis at rest. In addition, ME/CFS CD8 cells had lower levels of glycolysis after activation.

“Our work demonstrates the importance of looking at particular types of immune cells that have different jobs to do, rather than looking at them all mixed together, which can hide problems specific to particular cells,” said Dr. Hanson. “Additional studies focusing on specific cell types will be important to unravel what’s gone wrong with immune defenses in ME/CFS.”

Dr. Hanson’s group also looked at mitochondrial size and membrane potential, which can indicate the health of T cell mitochondria. CD4 cells from healthy controls and people with ME/CFS showed no significant differences in mitochondrial size nor function. CD8 cells from people with ME/CFS showed decreased membrane potential compared to healthy cells during both resting and activated states.

Dr. Hanson’s team examined associations between cytokines, chemical messengers that send instructions from one cell to another, and T cell metabolism. The findings revealed different, and often opposite, patterns between healthy and ME/CFS cells, suggesting changes in the immune system. In addition, the presence of cytokines that cause inflammation unexpectedly correlated with decreased metabolism in T cells.

This study was supported in part by the NIH’s ME/CFS Collaborative Research Network, a consortium supported by multiple institutes and centers at NIH, consisting of three collaborative research centers and a data management coordinating center. The research network was established in 2017 to help advance research on ME/CFS.

“In addition to providing valuable insights into the immunology of ME/CFS, we hope that the results coming out of the collaborative research network will inspire more researchers, particularly those in the early stages of their careers, to work on this disease,” said Joseph Breen, Ph.D., section chief, Immunoregulation Section, Basic Immunology Branch, National Institute of Allergy and Infectious Diseases (NIAID), which partially funded the study.

Future research studies will examine metabolism in other subsets of immune cells. In addition, researchers will investigate ways in which changes in metabolism affect the activity of T cells.

This study was supported by NINDS grant U54NS105541, NIAID grant R21AI117595, Simmaron Research, and an anonymous private donor.

NINDS (https://www.ninds.nih.gov/) is the nation’s leading funder of research on the brain and nervous system. The mission of NINDS is to seek fundamental knowledge about the brain and nervous system and to use that knowledge to reduce the burden of neurological disease.

About the National Institutes of Health (NIH): NIH, the nation’s medical research agency, includes 27 Institutes and Centers and is a component of the U.S. Department of Health and Human Services. NIH is the primary federal agency conducting and supporting basic, clinical, and translational medical research, and is investigating the causes, treatments, and cures for both common and rare diseases. For more information about NIH and its programs, visit www.nih.gov.

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Reference: Mandarano et al. “Myalgic encephalomyelitis/chronic fatigue syndrome patients exhibit altered T cell metabolism and cytokine associations,” Journal of Clinical Investigation. December 12, 2019

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome as a hyper-regulated immune system driven by an interplay between regulatory T cells and chronic human herpesvirus infections

Abstract:

Autoimmunity and chronic viral infections are recurrent clinical observations in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS), a complex disease with an unknown cause. Given these observations, the regulatory CD4+ T cells (Tregs) show promise to be good candidates for the underlying pathology due to their known capacity to suppress the immune responses not only to body components but also against infections. Here we discussed the overlooked role of these cells in the chronicity of Human Herpes Virus 6 (HHV6), Herpes Simplex 1 (HSV1) and Epstein-Barr virus (EBV), as often reported as triggers of ME/CFS.

Using simulations of the Cross-regulation model for the dynamics of Tregs, we illustrated that mild infections might lead to a chronically activated immune responses under control of Tregs if the responding clone has a high autoimmune potential. Such infections promote persistent inflammation and possibly fatigue. We then hypothesized that ME/CFS is a condition characterized by a predominance of this type of infections under control of Tregs. In contrast, healthy individuals are hypothesized to trigger immune responses of a virus-specific clone with a low autoimmune potential.

According to this hypothesis, simple model simulations of the CD4+ T-cell repertoire could reproduce the increased density and percentages of Tregs observed in patients suffering from the disease when compared to healthy controls. A deeper analysis of Tregs in the pathogenesis of ME/CFS will help to assess the validity of this hypothesis.

Source: Nuno Sepúlveda, Jorge Carneiro, Eliana M. Lacerda and Luis C. Nacul. Myalgic Encephalomyelitis/Chronic Fatigue Syndrome as a hyper-regulated immune system driven by an interplay between regulatory T cells and chronic human herpesvirus infections. Front. Immunol. | doi: 10.3389/fimmu.2019.02684. https://www.frontiersin.org/articles/10.3389/fimmu.2019.02684/abstract

CD4 T lymphocytes from patients with chronic fatigue syndrome have decreased interferon-gamma production and increased sensitivity to dexamethasone

Abstract:

A disturbed hypothalamus-pituitary-adrenal gland axis and alterations at the immune system level have been observed in patients with chronic fatigue syndrome (CFS). Glucocorticoids are known to modulate T cell responses; therefore, purified CD4 T cells from CFS patients were studied to determine whether they have an altered sensitivity to dexamethasone (DEX).

CD4 T cells from CFS patients produced less interferon-gamma than did cells from controls; by contrast, interleukin-4 production and cell proliferation were comparable. With CD4 T cells from CFS patients (compared with cells from controls), a 10- to 20-fold lower DEX concentration was needed to achieve 50% inhibition of interleukin-4 production and proliferation, indicating an increased sensitivity to DEX in CFS patients.

Surprisingly, interferon-gamma production in patients and controls was equally sensitive to DEX. A differential sensitivity of cytokines or CD4 T cell subsets to glucocorticoids might explain an altered immunologic function in CFS patients.

 

Source: Visser J, Blauw B, Hinloopen B, Brommer E, de Kloet ER, Kluft C, Nagelkerken L. CD4 T lymphocytes from patients with chronic fatigue syndrome have decreased interferon-gamma production and increased sensitivity to dexamethasone. J Infect Dis. 1998 Feb;177(2):451-4. http://jid.oxfordjournals.org/content/177/2/451.long

 

Clinical improvement in chronic fatigue syndrome is not associated with lymphocyte subsets of function or activation

Abstract:

The relationship between markers of immune function and chronic fatigue syndrome (CFS) is controversial. To examine the relationship directly, 43 subjects with CFS entering a randomized controlled trial of a nonpharmacological treatment for CFS gave samples for immunological analysis before and after treatment. Percentage levels of total CD3+ T cells, CD4 T cells, CD8 T cells, and activated subsets did not differ between CFS subjects and controls. Naive (CD45RA+ RO-) and memory (CD45RA- RO+) T cells did not differ between subjects and controls.

Natural killer cells (CD16+/CD56+/CD3-) were significantly increased in CFS patients compared to controls, as was the percentage of CD11b+ CD8 cells.

There were no correlations between any immune variable and measures of clinical status, with the exception of a weak correlation between total CD4 T cells and fatigue. There was a positive correlation between memory CD4 and CD8 T cells and depression scores and a negative correlation between naive CD4 T cells and depression.

No immune measures changed during the course of the study, and there was no link between clinical improvement as a result of the treatment program and immune status. Immune measures did not predict response or lack of response to treatment.

In conclusion, we have been unable to replicate previous findings of immune activation in CFS and unable to find any important associations between clinical status, treatment response, and immunological status.

 

Source: Peakman M, Deale A, Field R, Mahalingam M, Wessely S. Clinical improvement in chronic fatigue syndrome is not associated with lymphocyte subsets of function or activation. Clin Immunol Immunopathol. 1997 Jan;82(1):83-91. http://www.ncbi.nlm.nih.gov/pubmed/9000046

 

Immunological abnormalities in patients with chronic fatigue syndrome

Abstract:

Between January 1991 and January 1993, 265 patients who fulfilled the CDC criteria of the working case definition of Chronic Fatigue Syndrome (CFS) have been observed at our Institution and submitted for clinical and laboratory evaluation.

One hundred and sixty-three patients were females and 102 males, the median age was 35 years (range 4-55 years); all patients reported profound and prolonged fatigue, lasting for a median of 3 years (range 6 months-10 years), preceded or accompanied at appearance by fever in 185 cases, and neuropsychologic problems including inability to concentrate, difficulty in thinking, confusion, irritability, forgetfulness, and depression. The fatigue was so severe that it required 102 patients to stop their working activities for a period of time ranging from 3 months to 2 years (range 7 months).

In 40 consecutive patients a comprehensive immunologic testing by single and two-colour flow cytometry was performed and results compared with a group of 35 healthy, age- and sex-matched controls. Whilst no significant differences were found in the absolute numbers of circulating total T cells (CD3+) and of total helper/inducer (CD4+) or suppressor/cytotoxic (CD8+) T cells, an evident reduction in CD3-/CD16+ and CD57+/CD56+ NK lymphocytes along with an expansion of the CD8+/CD56+ and CD16-/CD56+ NK subsets, were found in the CFS group.

In addition, CD56+ NK cells from CFS subjects were found to express an increased amount of cell adhesion molecules (CD11b, CD11c, CD54) and activation antigens (CD38). Both the percentage and absolute numbers of CD4+ T cells bearing the CD45RA antigen appeared significantly reduced in CFS patients, and CD4+ T lymphocytes from CFS subjects displayed an increased expression of the intercellular adhesion molecule-1 (ICAM-1/CD54).

Finally, the total numbers of circulating (CD19+) B lymphocytes, were significantly higher in CFS cases than in controls, and in 11 out of 30 CFS patients the increase in circulating B cells was sustained by the expansion of the CD5+/CD19+ subset of B lymphocytes.

We conclude that CFS is a syndrome not previously described in Italy, with already known clinical characteristics and appears to be associated with several immunologic abnormalities, including those reported previously in cohort of patients from different countries. We also show for the first time that CD56- NK cell subsets from CFS patients display an abnormally increased expression of cell adhesion molecules and activation markers.

 

Source: Tirelli U, Marotta G, Improta S, Pinto A. Immunological abnormalities in patients with chronic fatigue syndrome. Scand J Immunol. 1994 Dec;40(6):601-8. http://www.ncbi.nlm.nih.gov/pubmed/7997849

 

Biological and molecular characteristics of human herpesvirus 7: in vitro growth optimization and development of a syncytia inhibition test

Abstract:

Two isolates of human herpesvirus 7 (HHV-7) were recovered from phytohemagglutinin-activated peripheral blood mononuclear cells of a patient with chronic fatigue syndrome and of a healthy blood donor. A genetic polymorphism between the two isolates was detected by Southern blot analysis using a novel HHV-7 genomic clone (pVL8) as a probe. We developed optimized conditions for the in vitro propagation of HHV-7 by using enriched populations of activated CD4+ T lymphocytes derived from normal peripheral blood, resulting in the production of high-titered extracellular virus (> 10(6) cell culture infectious doses/ml). Bona fide syncytia formation was documented both in normal CD4+ T lymphocytes and in the Sup-T1 CD4+ T-cell line following infection with high-titered HHV-7. To identify neutralizing antibodies to HHV-7, a syncytia-inhibition test was developed. Variable titers of syncytia-neutralizing antibodies were detected in all the human sera tested, thus confirming the high prevalence of HHV-7 in the human population.

 

Source: Secchiero P, Berneman ZN, Gallo RC, Lusso P. Biological and molecular characteristics of human herpesvirus 7: in vitro growth optimization and development of a syncytia inhibition test. Virology. 1994 Jul;202(1):506-12. http://www.ncbi.nlm.nih.gov/pubmed/8009865

 

Infection of natural killer cells by human herpesvirus 6

Abstract:

Natural killer (NK) cells are a functionally defined subset of non-T, non-B lymphocytes of bone marrow origin, which induce lysis of selected target cells, including neoplastic and virus-infected cells. The NK cell function provides an important mechanism of primary defence against viruses in vivo, as demonstrated by the occurrence of multiple herpesvirus infections in patients congenitally lacking NK cells.

Here we show that functionally competent CD3- NK clones can be productively infected by human herpesvirus 6 (HHV-6), a T-lymphotropic DNA virus that may play a role in the acquired immunodeficiency syndrome (AIDS) and in the chronic fatigue syndrome, two disorders associated with a defective NK cell activity.

The infection is cytopathic and induces de novo expression of CD4, an antigen not expressed within the NK lineage, thereby predisposing NK cells to infection by human immunodeficiency virus type 1 (HIV-1).

These results provide evidence that a herpesvirus can directly target and kill NK cells, a potential strategy to suppress the natural anti-viral immunity of the host.

 

Source: Lusso P, Malnati MS, Garzino-Demo A, Crowley RW, Long EO, Gallo RC. Infection of natural killer cells by human herpesvirus 6. Nature. 1993 Apr 1;362(6419):458-62. http://www.ncbi.nlm.nih.gov/pubmed/7681936

 

Lymphocyte phenotype and function in the chronic fatigue syndrome

Abstract:

Lymphocytes of 18 patients meeting the Centers for Disease Control (CDC) case definition for the chronic fatigue syndrome (CFS), 10 similar, chronically fatigued patients not fully conforming to the CDC case definition, and 17 matched, healthy individuals were studied to determine the presence of abnormalities of peripheral cell phenotype and function.

Extensive phenotypic analyses of B- and T-cell subsets, natural killer (NK) cells, and macrophages were performed using single-, dual-, and three-color flow cytometry. Compared to controls, in CFS patients the percentage of CD4 T cells and CD4,CD45RA, or naive T cells, was reduced. The CD4,CD45RO, or memory T-cell, subset was numerically normal but expressed increased levels of adhesion markers (CD29, CD54, and CD58). CFS patient lymphocytes showed reduced proliferative responses to phytohemagglutinin, concanavalin A, and staphylococcal enterotoxin B. Lymphocytes from fatigue patients not meeting the CDC definition showed similar abnormalities.

These data indicate that peripheral T cells manifest an increased state of differentiation in CFS and related conditions. This may arise as a consequence of an underlying neuropsychiatric and/or neuroendocrine disorder or because of exposure to antigens or superantigens of an infectious agent.

 

Source: Straus SE, Fritz S, Dale JK, Gould B, Strober W. Lymphocyte phenotype and function in the chronic fatigue syndrome. J Clin Immunol. 1993 Jan;13(1):30-40. http://www.ncbi.nlm.nih.gov/pubmed/8095270