Tag: NK cells

Further clues in the fight against chronic fatigue syndrome

New findings regarding the pathology of Chronic Fatigue Syndrome (CFS) are bringing Griffith University researchers closer to identifying the cause of this disabling illness.

This is the news from a team at the National Centre for Neuroimmunology and Emerging Diseases at the Menzies Health Institute Queensland.

Professors Marshall-Gradisnik and Don Staines and their research team have identified significant impairments in cellular function of people with CFS.

CFS — sometimes known as ME (myalgic encephalomyelitis) — is a complex illness characterized by impaired memory and concentration, metabolic, cardiac, gut and immune dysfunction and debilitating muscle pain and fatigue on exertion (also known as neuroimmune exhaustion).

It is estimated that the prevalence rate of CFS/ME worldwide is between 1 and 2 per cent.

“While the patho-mechanism of CFS/ME is unknown, these recent findings by NCNED researchers provide further evidence for the pathology of this illness,” says Professor Sonya Marshall-Gradisnik, who speaks as we approach International CFS Awareness Day on Thursday May 12.

Published in the Journal of Translational Medicine, the results report significant differences in intracellular signalling of cells with CFS patients.

“In this group, we see that dysfunctional signalling may contribute to impaired cell activity. These findings are consistent with our previous findings and align with the presentation of symptoms in patients,” says Professor Staines.

The current research findings build upon recent discoveries including novel identification of key genetic changes in cells of the immune system.

The NCNED — internationally recognised for research into CFS/ME — will present a seminar on current research findings on this disease on International CFS/ME Awareness Day, Thursday May 12 at Griffith University, Gold Coast Campus, commencing 1pm, location G17, Lecture theatre 3.

Griffith University will also be illuminating the Griffith Health Centre in blue to further help raise awareness for CFS/ME.

Journal Reference: Teilah Kathryn Huth, Donald Staines, Sonya Marshall-Gradisnik. ERK1/2, MEK1/2 and p38 downstream signalling molecules impaired in CD56dimCD16 and CD56brightCD16dim/− natural killer cells in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis patients. Journal of Translational Medicine, 2016; 14 (1) DOI: 10.1186/s12967-016-0859-z

 

Source: Griffith University. “Further clues in the fight against chronic fatigue syndrome.” ScienceDaily. ScienceDaily, 10 May 2016. https://www.sciencedaily.com/releases/2016/05/160510093906.htm

 

Dysregulation of Protein Kinase Gene Expression in NK Cells from Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients

Abstract:

BACKGROUND: The etiology and pathomechanism of chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) are unknown. However, natural killer (NK) cell dysfunction, in particular reduced NK cytotoxic activity, is a consistent finding in CFS/ME patients. Previous research has reported significant changes in intracellular mitogen-activated protein kinase pathways from isolated NK cells. The purpose of this present investigation was to examine whether protein kinase genes have a role in abnormal NK cell intracellular signaling in CFS/ME.

METHOD: Messenger RNA (mRNA) expression of 528 protein kinase genes in isolated NK cells was analyzed (nCounter GX Human Kinase Kit v2 (XT); NanoString Technologies) from moderate (n = 11; age, 54.9 ± 10.3 years) and severe (n = 12; age, 47.5 ± 8.0 years) CFS/ME patients (classified by the 2011 International Consensus Criteria) and nonfatigued controls (n = 11; age, 50.0 ± 12.3 years).

RESULTS: The expression of 92 protein kinase genes was significantly different in the severe CFS/ME group compared with nonfatigued controls. Among these, 37 genes were significantly upregulated and 55 genes were significantly downregulated in severe CFS/ME patients compared with nonfatigued controls.

CONCLUSIONS: In severe CFS/ME patients, dysfunction in protein kinase genes may contribute to impairments in NK cell intracellular signaling and effector function. Similar changes in protein kinase genes may be present in other cells, potentially contributing to the pathomechanism of this illness.

 

Source: Chacko A, Staines DR, Johnston SC, Marshall-Gradisnik SM. Dysregulation of Protein Kinase Gene Expression in NK Cells from Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients. Gene Regul Syst Bio. 2016 Aug 28;10:85-93. doi: 10.4137/GRSB.S40036. ECollection 2016. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5003121/ (Full article)

 

Killer Cell Immunoglobulin-like Receptor Genotype and Haplotype Investigation of Natural Killer Cells from an Australian Population of Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients

Abstract:

Killer cell immunoglobulin-like receptor (KIR) genes encode for activating and inhibitory surface receptors, which are correlated with the regulation of Natural Killer (NK) cell cytotoxic activity. Reduced NK cell cytotoxic activity has been consistently reported in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) patients, and KIR haplotypes and allelic polymorphism remain to be investigated.

The aim of this article was to conduct a pilot study to examine KIR genotypes, haplotypes, and allelic polymorphism in CFS/ME patients and nonfatigued controls (NFCs). Comparison of KIR and allelic polymorphism frequencies revealed no significant differences between 20 CFS/ME patients and 20 NFCs.

A lower frequency of the telomeric A/B motif (P < 0.05) was observed in CFS/ME patients compared with NFCs. This pilot study is the first to report the differences in the frequency of KIR on the telomeric A/B motif in CFS/ME patients. Further studies with a larger CFS/ME cohort are required to validate these results.

 

Source: Huth TK, Brenu EW, Staines DR, Marshall-Gradisnik SM. Killer Cell Immunoglobulin-like Receptor Genotype and Haplotype Investigation of Natural Killer Cells from an Australian Population of Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients. Gene Regul Syst Bio. 2016 Jun 19;10:43-9. doi: 10.4137/GRSB.S39861. ECollection 2016. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4913894/ (Full article)

 

Novel identification and characterisation of Transient receptor potential melastatin 3 ion channels on Natural Killer cells and B lymphocytes: effects on cell signalling in Chronic fatigue syndrome/Myalgic encephalomyelitis patients

Abstract:

BACKGROUND: Transient receptor potential melastatin 3 (TRPM3) cation channels are ubiquitously expressed by multiple cells and have an important regulatory role in calcium-dependent cell signalling to help maintain cellular homeostasis. TRPM3 protein expression has yet to be determined on Natural Killer (NK) cells and B lymphocytes. Multiple single nucleotide polymorphisms have been reported in TRPM3 genes from isolated peripheral blood mononuclear cells, NK and B cells in Chronic fatigue syndrome/Myalgic encephalomyelitis (CFS/ME) patients and have been proposed to correlate with illness presentation. The object of the study was to assess TRPM3 surface expression on NK and B lymphocytes from healthy controls, followed by a comparative investigation examining TRPM3 surface expression, and cytoplasmic and mitochondrial calcium influx in CD19(+) B cells, CD56(bright) and CD56(dim) cell populations from CFS/ME patients.

RESULTS: TRPM3 cell surface expression was identified for NK and B lymphocytes in healthy controls (CD56(bright) TRPM3 35.72 % ± 7.37; CD56(dim) 5.74 % ± 2.00; B lymphocytes 2.05 % ± 0.19, respectively). There was a significant reduction of TRPM3 surface expression on CD19(+) B cells (1.56 ± 0.191) and CD56(bright) NK cells (17.37 % ± 5.34) in CFS/ME compared with healthy controls. Anti-CD21 and anti-IgM conjugated biotin was cross-linked with streptavidin,and subsequently treatment with thapsigargin. This showed a significant reduction in cytoplasmic calcium ion concentration in CD19(+) B lymphocytes. CD56(bright) NK cells also had a significant decrease in cytoplasmic calcium in the presence of 2-APB and thapsigargin in CFS/ME patients.

CONCLUSIONS: The results from this preliminary investigation identify, for the first time, TRPM3 surface expression on both NK and B lymphocytes in healthy controls. We also report for the first time, significant reduction in TRPM3 cell surface expression in NK and B lymphocytes, as well as decreased intracellular calcium within specific conditions in CFS/ME patients. This warrants further examination of these pathways to elucidate whether TRPM3 and impaired calcium mobilisation has a role in CFS/ME.

 

Source: Nguyen T, Staines D, Nilius B, Smith P, Marshall-Gradisnik S. Novel identification and characterisation of Transient receptor potential melastatin 3 ion channels on Natural Killer cells and B lymphocytes: effects on cell signalling in Chronic fatigue syndrome/Myalgic encephalomyelitis patients. Biol Res. 2016 May 31;49(1):27. doi: 10.1186/s40659-016-0087-2. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4888729/ (Full article)

 

Natural killer cells and single nucleotide polymorphisms of specific ion channels and receptor genes in myalgic encephalomyelitis/chronic fatigue syndrome

Abstract:

AIM: The aim of this paper was to determine natural killer (NK) cytotoxic activity and if single nucleotide polymorphisms (SNPs) and genotypes in transient receptor potential (TRP) ion channels and acetylcholine receptors (AChRs) were present in isolated NK cells from previously identified myalgic encephalomyelitis (ME)/chronic fatigue syndrome (CFS) patients.

SUBJECTS AND METHODS: A total of 39 ME/CFS patients (51.69±2 years old) and 30 unfatigued controls (47.60±2.39 years old) were included in this study. Patients were defined according to the 1994 Centers for Disease Control and Prevention criteria. Flow cytometry protocols were used to examine NK cytotoxic activity. A total of 678 SNPs from isolated NK cells were examined for 21 mammalian TRP ion channel genes and for nine mammalian AChR genes via the Agena Bioscience iPlex Gold assay. SNP association and genotype was determined using analysis of variance and Plink software.

RESULTS: ME/CFS patients had a significant reduction in NK percentage lysis of target cells (17%±4.68%) compared with the unfatigued control group (31%±6.78%). Of the 678 SNPs examined, eleven SNPs for TRP ion channel genes (TRPC4, TRPC2, TRPM3, and TRPM8) were identified in the ME/CFS group. Five of these SNPs were associated with TRPM3, while the remainder were associated with TRPM8, TRPC2, and TRPC4 (P<0.05). Fourteen SNPs were associated with nicotinic and muscarinic AChR genes: six with CHRNA3, while the remainder were associated with CHRNA2, CHRNB4, CHRNA5, and CHRNE (P<0.05). There were sixteen genotypes identified from SNPs in TRP ion channels and AChRs for TRPM3 (n=5), TRPM8 (n=2), TRPC4 (n=3), TRPC2 (n=1), CHRNE (n=1), CHRNA2 (n=2), CHRNA3 (n=1), and CHRNB4 (n=1) (P<0.05).

CONCLUSION: We identified a number of SNPs and genotypes for TRP ion channels and AChRs from isolated NK cells in patients with ME/CFS, suggesting these SNPs and genotypes may be involved in changes in NK cell function and the development of ME/CFS pathology. These anomalies suggest a role for dysregulation of Ca(2+) in AChR and TRP ion channel signaling in the pathomechanism of ME/CFS.

 

Source: Marshall-Gradisnik S, Huth T, Chacko A, Johnston S, Smith P, Staines D. Natural killer cells and single nucleotide polymorphisms of specific ion channels and receptor genes in myalgic encephalomyelitis/chronic fatigue syndrome. Appl Clin Genet. 2016 Mar 31;9:39-47. doi: 10.2147/TACG.S99405. ECollection 2016. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4821384/ (Full article)

 

ERK1/2, MEK1/2 and p38 downstream signalling molecules impaired in CD56 dim CD16+ and CD56 bright CD16 dim/- natural killer cells in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis patients

Abstract:

BACKGROUND: Natural Killer (NK) cell effector functions are dependent on phosphorylation of the mitogen-activated protein kinases (MAPK) pathway to produce an effective immune response for the clearance of target cells infected with viruses, bacteria or malignantly transformed cells. Intracellular signals activating NK cell cytokine production and cytotoxic activity are propagated through protein phosphorylation of MAPKs including MEK1/2, ERK1/2, p38 and JNK. Reduced NK cell cytotoxic activity is consistently reported in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) patients and intracellular signalling by MAPK in NK cells remains to be investigated. Therefore, the purpose of this paper was to investigate MAPK downstream signalling molecules in NK cell phenotypes from CFS/ME patients.

METHODS: Flow cytometric protocols were used to measure phosphorylation of the MAPK pathway in CD56(bright)CD16(dim/-) and CD56(dim)CD16(+) NK cells following stimulation with K562 tumour cells or phorbol-12-myristate-13-acetate plus ionomycin. NK cell cytotoxic activity, degranulation, lytic proteins and cytokine production were also measured as markers for CD56(bright)CD16(dim/-) and CD56(dim)CD16(+) NK cell function using flow cytometric protocols.

RESULTS: CFS/ME patients (n = 14) had a significant decrease in ERK1/2 in CD56(dim)CD16(+) NK cells compared to the non-fatigued controls (n = 11) after incubation with K562 cells. CD56(bright)CD16(dim/-) NK cells from CFS/ME patients had a significant increase in MEK1/2 and p38 following incubation with K562 cells.

CONCLUSIONS: This is the first study to report significant differences in MAPK intracellular signalling molecules in CD56(dim)CD16(+) and CD56(bright)CD16(dim/-) NK cells from CFS/ME patients. The current results highlight the importance of intracellular signalling through the MAPK pathway for synergistic effector function of CD56(dim)CD16(+) and CD56(bright)CD16(dim/-) NK cells to ensure efficient clearance of target cells. In CFS/ME patients, dysfunctional MAPK signalling may contribute to reduced NK cell cytotoxic activity.

 

Source: Huth TK, Staines D, Marshall-Gradisnik S. ERK1/2, MEK1/2 and p38 downstream signalling molecules impaired in CD56 dim CD16+ and CD56 bright CD16 dim/- natural killer cells in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis patients. J Transl Med. 2016 Apr 21;14:97. doi: 10.1186/s12967-016-0859-z. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4839077/ (Full article)

 

Regulatory T, natural killer T and γδ T cells in multiple sclerosis and chronic fatigue syndrome/myalgic encephalomyelitis: a comparison

Abstract:

BACKGROUND: Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME), and Multiple Sclerosis (MS) may share some similarities in relation to reduced NK cell activity. It is likely that other cells such as regulatory T (Tregs), invariant Natural Killer T (iNKT) and gamma delta T (γδ T) cells may also be dysregulated in CFS/ME and MS.

OBJECTIVE: To evaluate and compare specific immune regulatory cells of patients with CFS/ME, patients with MS and healthy controls.

METHOD: Sixty three volunteers were included in this study: 24 were CFS/ME patients, 11 were MS patients and 27 were healthy controls. Blood samples were obtained from all participants for flow cytometry analysis of iNKT cells, Tregs and γδ T cell phenotypes.

RESULTS: We observed a significant increase in Tregs in the CFS/ME group (p≤0.05) compared to the healthy control group. Total γδ and γδ2 T cells were significantly reduced in MS patients in comparison with the healthy control group. Conversely, CD4+iNKT percentage of iNKT, was significantly increased in the CFS/ME group compared with healthy controls and the double-negative iNKT percentage of iNKT significantly decreased compared with the healthy control group.

CONCLUSIONS: This study has not identified any immunological disturbances that are common in both MS and CFS/ME patients. However, the differential expression of cell types between the conditions investigated suggests different pathways of disease. These differences need to be explored in further studies.

 

Source: Ramos S, Brenu E, Broadley S, Kwiatek R, Ng J, Nguyen T, Freeman S, Staines D, Marshall-Gradisnik S. Regulatory T, natural killer T and γδ T cells in multiple sclerosis and chronic fatigue syndrome/myalgic encephalomyelitis: a comparison. Asian Pac J Allergy Immunol. 2016 Dec;34(4):300-305. doi: 10.12932/AP0733. http://apjai-journal.org/wp-content/uploads/2016/12/8RegulatoryTnaturalkillerAPJAIVol34No4December2016P300.pdf (Full text as PDF)

 

Pilot Study of Natural Killer Cells in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis and Multiple Sclerosis

Abstract:

Patients with chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) and multiple sclerosis (MS) suffer from debilitating fatigue which is not alleviated by rest. In addition to the fatigue-related symptoms suffered by patients with CFS/ME and MS, dysfunction of the immune system and, in particular, reduced natural killer (NK) cell cytotoxic activity has also been reported in CFS/ME and MS. The purpose of this pilot study was to compare NK cellular mechanisms in patients with CFS/ME and MS to investigate potential dysfunctions in the NK cell activity pathway. Flow cytometry protocols assessed CD56(dim) CD16(+) and CD56(bright) CD16(+/-) NK cell expression of adhesion molecules, NK activating and inhibiting receptors, NK cell maturation and lytic proteins.

All participants in this study were female and included 14 patients with CFS/ME, nine patients with MS and 19 non-fatigued controls. The patient groups and the non-fatigued controls were not taking any immunosuppressive or immune-enhancing medications. In the MS cohort, KIR2DL5 was significantly increased on CD56(bright) CD16(+/-) NK cells and expression of CD94 was significantly increased on CD56(dim) CD16(+) NK cells in comparison with the controls. Co-expression of CD57 and perforin was significantly increased on CD56(dim) CD16(+) NK cells from patients with CFS/ME compared to the MS and non-fatigued control participants.

The results from this pilot study suggest that NK cells from patients with CFS/ME and MS may have undergone increased differentiation in response to external stimuli which may affect different mechanisms in the NK cell cytotoxic activity pathway.

© 2015 The Authors. Scandinavian Journal of Immunology published by John Wiley & Sons Ltd on behalf of The Foundation for the Scandinavian Journal of Immunology.

 

Source: Huth TK, Brenu EW, Ramos S, Nguyen T, Broadley S, Staines D, Marshall-Gradisnik S. Pilot Study of Natural Killer Cells in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis and Multiple Sclerosis. Scand J Immunol. 2016 Jan;83(1):44-51. doi: 10.1111/sji.12388. http://onlinelibrary.wiley.com/wol1/doi/10.1111/sji.12388/full (Full article)

 

Natural killer cells in patients with severe chronic fatigue syndrome

Abstract:

Maintenance of health and physiological homeostasis is a synergistic process involving tight regulation of proteins, transcription factors and other molecular processes. The immune system consists of innate and adaptive immune cells that are required to sustain immunity. The presence of pathogens and tumour cells activates innate immune cells, in particular Natural Killer (NK) cells.

Stochastic expression of NK receptors activates either inhibitory or activating signals and results in cytokine production and activation of pathways that result in apoptosis of target cells. Thus, NK cells are a necessary component of the immunological process and aberrations in their functional processes, including equivocal levels of NK cells and cytotoxic activity pre-empts recurrent viral infections, autoimmune diseases and altered inflammatory responses. NK cells are implicated in a number of diseases including chronic fatigue syndrome (CFS). The purpose of this review is to highlight the different profiles of NK cells reported in CFS patients and to determine the extent of NK immune dysfunction in subtypes of CFS patients based on severity in symptoms.

 

Source: Brenu EW, Hardcastle SL, Atkinson GM, van Driel ML, Kreijkamp-Kaspers S, Ashton KJ, Staines DR, Marshall-Gradisnik SM. Natural killer cells in patients with severe chronic fatigue syndrome. Auto Immun Highlights. 2013 Apr 16;4(3):69-80. doi: 10.1007/s13317-013-0051-x. ECollection 2013. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4389023/ (Full article)

 

Immunophenotyping in post-giardiasis functional gastrointestinal disease and chronic fatigue syndrome

Abstract:

BACKGROUND: A Giardia outbreak was associated with development of post-infectious functional gastrointestinal disorders (PI-FGID) and chronic fatigue syndrome (PI-CFS). Markers of immune dysfunction have given conflicting results in CFS and FGID patient populations. The aim of this study was to evaluate a wide selection of markers of immune dysfunction in these two co-occurring post-infectious syndromes.

METHODS: 48 patients, reporting chronic fatigue in a questionnaire study, were clinically evaluated five years after the outbreak and grouped according to Fukuda criteria for CFS (n=19) and idiopathic chronic fatigue (n=5) and Rome II criteria for FGIDs (n=54). 22 Giardia exposed non-fatigued individuals and 10 healthy unexposed individuals were recruited as controls. Peripheral blood lymphocyte subsets were analyzed by flow cytometry.

RESULTS: In peripheral blood we found significantly higher CD8 T-cell levels in PI-FGID, and significantly lower NK-cell levels in PI-CFS patients. Severity of abdominal and fatigue symptoms correlated negatively with NK-cell levels. A tendency towards lower T-cell CD26 expression in FGID was seen.

CONCLUSION: Patients with PI-CFS and/or PI-FGID 5 years after Giardia lamblia infection showed alterations in NK-cell and CD8-cell populations suggesting a possible immunological abnormality in these conditions. We found no significant changes in other markers examined in this well-defined group of PI-CFS and PI-FGID elicited by a gastrointestinal infection. Controlling for co-morbid conditions is important in evaluation of CFS-biomarkers.

 

Source: Hanevik K, Kristoffersen EK, Sørnes S, Mørch K, Næss H, Rivenes AC, Bødtker JE, Hausken T, Langeland N. Immunophenotyping in post-giardiasis functional gastrointestinal disease and chronic fatigue syndrome. BMC Infect Dis. 2012 Oct 14;12:258. doi: 10.1186/1471-2334-12-258. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3553045/ (Full article)