Laura Clarke^1,2, Simon L. Broadley^1,2, Thao Nguyen^3,4, Samantha Johnston^3,4, Natalie Eaton^3,4, Donald Staines^3,4, Sonya Marshall-Gradisnik^3,4*

Show more
¹ School of Medicine, Griffith University, Gold Coast, Australia.
² Department of Neurology, Gold Coast University Hospital, Gold Coast, Australia.
³ The National Centre for Neuroimmunology and Emerging Diseases, Menzies Health Institute, Griffith University, Gold Coast, Australia.
⁴ School of Medical Science, Griffith University, Brisbane, Australia.
Show less

Abstract: Background: Natural killer (NK) cell phenotypes have reported to be implicated in the pathomechanism of Multiple Sclerosis (MS). Several investigators have observed reduced peripheral numbers, reduced cytotoxic activity, and altered CD56^Dim and CD56^Bright NK cell phenotypes. This current project, for the first time, investigates the NK cell cytotoxicity, calcium mobilisation and transient receptor potential melastatin 3 (TRPM3) surface expression. Methods: NK cell cytotoxic activity and calcium signaling were examined in CD56^Dim and CD56^Bright NK cells before and after stimulation using Ionomycin, Pregnenolone sulphate, 2-Aminoethoxydiphenyl borate and Thapsigargin. Purified NK cells were labelled with antibodies to determine TRPM3, CD69 and CD107a surface expression using flow cytometry. Results: Twenty-two MS patients and 22 healthy controls were recruited for this project. Twelve of the 22 previously received Alemtuzumab (Lemtrada®) and the remaining ten reported nil medication. We report TRPM3 was significantly increased in untreated MS patients compared with healthy controls and treated MS patients (p-value 0.034). There was a significant decrease in CD69 surface expression on CD56^Dim NK cell phenotype for untreated MS patients (p-value 0.031) and treated MS patients (p-value 0.036). We report altered calcium mobilisation in CD56^Bright NK cells and to a lesser extent CD56^Dim NK cells between healthy controls, treated and untreated MS patients. Conclusion: This investigation suggests variations in TRPM3 expression and calcium mobilisation of NK cells may be implicated in the pathogenesis of MS. Further investigation is required to determine the mechanism by which alemtuzumab alters calcium signaling in NK cells.

Keywords: Natural Killer Cells, Multiple Sclerosis, Calcium Signalling, Transient Receptor Potential Melastatin 3 Ion Channels

Cite this paper: Clarke, L. , Broadley, S. , Nguyen, T. , Johnston, S. , Eaton, N. , Staines, D. and Marshall-Gradisnik, S. (2018) Transient Receptor Potential Melastatin 3 and Intracellular Calcium in Natural Killer Cells in Multiple Sclerosis. International Journal of Clinical Medicine, 9, 541-565. doi: 10.4236/ijcm.2018.97047.

References

[1]   Gandhi, R., Laroni, A. and Weiner, H.L. (2010) Role of the Innate Immune System in the Pathogenesis of Multiple Sclerosis. Journal of Neuroimmunology, 221, 7-14.
https://doi.org/10.1016/j.jneuroim.2009.10.015

[2]   Hemmer, B., Kerschensteiner, M. and Korn, T. (2015) Role of the Innate and Adaptive Immune Responses in the Course of Multiple Sclerosis. The Lancet Neurology, 14, 406-419.
https://doi.org/10.1016/S1474-4422(14)70305-9

[3]   Goldenberg, M.M. (2012) Multiple Sclerosis Review. Pharmacy and Therapeutics, 37, 175-184.

[4]   Kingwell, E., et al. (2013) Incidence and Prevalence of Multiple Sclerosis in Europe: A Systematic Review. BMC Neurology, 13, 128.

[5]   Segal, B.M. (2007) The Role of Natural Killer Cells in Curbing Neuroinflammation. Journal of Neuroimmunology, 191, 2-7.
https://doi.org/10.1016/j.jneuroim.2007.09.006

[6]   Benczur, M., et al. (1980) Dysfunction of Natural Killer Cells in Multiple Sclerosis: A Possible Pathogenetic Factor. Clinical and Experimental Immunology, 39, 657-662.

[7]   Merrill, J., Jondal, M., Seeley, J., Ullberg, M. and Sidén, A. (1982) Decreased NK Killing in Patients with Multiple Sclerosis: An Analysis on the Level of the Single Effector Cell in Peripheral Blood and Cerebrospinal Fluid in Relation to the Activity of the Disease. Clinical and Experimental Immunology, 47, 419-430.

[8]   Duffy, S.S., Lees, J.G. and Moalem-Taylor, G. (2014) The Contribution of Immune and Glial Cell Types in Experimental Autoimmune Encephalomyelitis and Multiple Sclerosis. Multiple Sclerosis International, 2014, Article ID: 285245.

[9]   Caligiuri, M.A. (2008) Human Natural Killer Cells. Blood, 112, 461-469.
https://doi.org/10.1182/blood-2007-09-077438

[10]   Vranes, Z., Poljakovic, Z. and Marusic, M. (1989) Natural Killer Cell Number and Activity in Multiple Sclerosis. Journal of the Neurological Sciences, 94, 115-123.
https://doi.org/10.1016/0022-510X(89)90222-0

[11]   Uchida, A., Maida, E.M., Lenzhofer, R. and Micksche1, M. (1982) Natural Killer Cell Activity in Patients with Multiple Sclerosis: Interferon and Plasmapheresis. Immunobiology, 160, 392-402.
https://doi.org/10.1016/S0171-2985(82)80003-X

[12]   Hirsch, R.L. and Johnson, K.P. (1985) Natural Killer Cell Activity in Multiple Sclerosis Patients Treated with Recombinant Interferon-α2. Clinical Immunology and Immunopathology, 37, 236-244.
https://doi.org/10.1016/0090-1229(85)90155-2

[13]   Braakman, E., van Tunen, A., Meager, A. and Lucas, C.J. (1986) Natural Cytotoxic Activity in Multiple Sclerosis Patients: Defects in IL-2/Interferon Gamma-Regulatory Circuit. Clinical & Experimental Immunology, 66, 285-294.

[14]   Bielekova, B., et al. (2006) Regulatory CD56bright Natural Killer Cells Mediate Immunomodulatory Effects of IL-2Rα-Targeted Therapy (Daclizumab) in Multiple Sclerosis. Proceedings of the National Academy of Sciences of the United States of America, 103, 5941-5946.
https://doi.org/10.1073/pnas.0601335103

[15]   Kastrukoff, L.F., Lau, A., Wee, R., Zecchini, D., White, R. and Paty, D.W. (2003) Clinical Relapses of Multiple Sclerosis Are Associated with “Nove” Valleys in Natural Killer Cell Functional Activity. Journal of Neuroimmunology, 145, 103-114.
https://doi.org/10.1016/j.jneuroim.2003.10.001

[16]   Feske, S., Wulff, H. and Skolnik, E.Y. (2015) Ion Channels in Innate and Adaptive Immunity. Annual Review of Immunology, 33, 291-353.
https://doi.org/10.1146/annurev-immunol-032414-112212

[17]   Schwarz, E.C., Qu, B. and Hoth, M. (2013) Calcium, Cancer and Killing: The Role of Calcium in Killing Cancer Cells by Cytotoxic T Lymphocytes and Natural Killer Cells. Biochimica et BiophysicaActa (BBA)-Molecular Cell Research, 1833, 1603-1611.
https://doi.org/10.1016/j.bbamcr.2012.11.016

[18]   Nilius, B. and Owsianik, G. (2011) The Transient Receptor Potential Family of Ion Channels. Genome Biology, 12, 218.
https://doi.org/10.1186/gb-2011-12-3-218

[19]   Nilius, B. and Flockerzi, V. (2014) Mammalian Transient Receptor Potential (TRP) Cation Channels. Vol. 2, Springer, Berlin.
https://doi.org/10.1007/978-3-642-54215-2

[20]   Fleig, A. and Penner, R. (2004) The TRPM Ion Channel Subfamily: Molecular, Biophysical and Functional Features. Trends in Pharmacological Sciences, 25, 633-639.
https://doi.org/10.1016/j.tips.2004.10.004

[21]   Moran, M.M., McAlexander, M.A., Bíró, T. and Szallasi, A. (2011) Transient Receptor Potential Channels as Therapeutic Targets. Nature Reviews Drug Discovery, 10, 601-620.
https://doi.org/10.1038/nrd3456

[22]   Clapham, D.E., Runnels, L.W. and Strübing, C. (2001) The TRP Ion Channel Family. Nature Reviews Neuroscience, 2, 387-396.
https://doi.org/10.1038/35077544

[23]   Schattling, B., et al. (2012) TRPM4 Cation Channel Mediates Axonal and Neuronal Degeneration in Experimental Autoimmune Encephalomyelitis and Multiple Sclerosis. Nature Medicine, 18, 1805-1811.
https://doi.org/10.1038/nm.3015

[24]   Paltser, G., et al. (2013) TRPV1 Gates Tissue Access and Sustains Pathogenicity in Autoimmune Encephalitis. Molecular Medicine, 19, 149-159.
https://doi.org/10.2119/molmed.2012.00329

[25]   Grimm, C., Kraft, R., Sauerbruch, S., Schultzand, G. and Harteneck, C. (2003) Molecular and Functional Characterization of the Melastatin-Related Cation Channel TRPM3. Journal of Biological Chemistry, 278, 21493-21501.
https://doi.org/10.1074/jbc.M300945200

[26]   Papanikolaou, M., Lewis, A. and Butt, A. (2017) Store-Operated Calcium Entry Is Essential for Glial Calcium Signalling in CNS White Matter. Brain Structure and Function, 222, 2993-3005.
https://doi.org/10.1007/s00429-017-1380-8

[27]   Vriens, J., et al. (2011) TRPM3 Is a Nociceptor Channel Involved in the Detection of Noxious Heat. Neuron, 70, 482-494.
https://doi.org/10.1016/j.neuron.2011.02.051

[28]   Oberwinkler, J. and Philipp, S. (2007) TRPM3. In: Flockerzi, V. and Nilius, B., Eds., Transient Receptor Potential (TRP) Channels, Springer, Berlin, 253-267.
https://doi.org/10.1007/978-3-540-34891-7_15

[29]   Nguyen, T., Staines, D., Nilius, B., Smith, P. and Marshall-Gradisnik, S. (2016) 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. Biological Research, 49, 27.
https://doi.org/10.1186/s40659-016-0087-2

[30]   Zierler, S., Hampe, S. and Nadolni, W. (2017) TRPM Channels as Potential Therapeutic Targets against Pro-Inflammatory Diseases. Cell Calcium, 67, 105-115.
https://doi.org/10.1016/j.ceca.2017.05.002

[31]   Launay, P., et al. (2004) TRPM4 Regulates Calcium Oscillations after T Cell Activation. Science, 306, 1374-1377.
https://doi.org/10.1126/science.1098845

[32]   Nguyen, T., Johnston, S., Clarke, L., Smith, P. Staines, D. and Marshall‐Gradisnik, S. (2017) Impaired Calcium Mobilization in Natural Killer Cells from Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients Is Associated with Transient Receptor Potential Melastatin 3 Ion Channels. Clinical & Experimental Immunology, 187, 284-293.
https://doi.org/10.1111/cei.12882

[33]   Willis, M.D. and Robertson, N.P. (2016) Alemtuzumab for Multiple Sclerosis. Current Neurology and Neuroscience Reports, 16, 84.
https://doi.org/10.1007/s11910-016-0685-y

[34]   Jiang, L., et al. (2009) Variable CD52 Expression in Mature T Cell and NK Cell Malignancies: Implications for Alemtuzumab Therapy. British Journal of Haematology, 145, 173-179.
https://doi.org/10.1111/j.1365-2141.2009.07606.x

[35]   Moreau, T., et al. (1994) Preliminary Evidence from Magnetic Resonance Imaging for Reduction in Disease Activity after Lymphocyte Depletion in Multiple Sclerosis. The Lancet, 344, 298-301.
https://doi.org/10.1016/S0140-6736(94)91339-0

[36]   Coles, A.J., et al. (1999) Monoclonal Antibody Treatment Exposes Three Mechanisms Underlying the Clinical Course of Multiple Sclerosis. Annals of Neurology, 46, 296-304.
https://doi.org/10.1002/1531-8249(199909)46:3<296::AID-ANA4>3.0.CO;2-#

[37]   Hu, Y., Turner, M.J., Shields J., et al. (2009) Investigation of the Mechanism of Action of Alemtuzumab in a Human CD52 Transgenic Mouse Model. Immunology, 128, 260-270.
https://doi.org/10.1111/j.1365-2567.2009.03115.x

[38]   Polman, C.H., et al. (2011) Diagnostic Criteria for Multiple Sclerosis: 2010 Revisions to the McDonald Criteria. Annals of Neurology, 69, 292-302.
https://doi.org/10.1002/ana.22366

[39]   Wagner, T.F., et al. (2008) Transient Receptor Potential M3 Channels Are Ionotropic Steroid Receptors in Pancreatic β Cells. Nature Cell Biology, 10, 1421-1430.
https://doi.org/10.1038/ncb1801

[40]   Lytton, J., Westlin, M. and Hanley, M.R. (1991) Thapsigargin Inhibits the Sarcoplasmic or Endoplasmic Reticulum Ca-ATPase Family of Calcium Pumps. Journal of Biological Chemistry, 266, 17067-17071.

[41]   Huth, T., Brenu E.W., Ramos, S., et al. (2016) Pilot Study of Natural Killer Cells in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis and Multiple Sclerosis. Scandinavian Journal of Immunology, 83, 44-51.
https://doi.org/10.1111/sji.12388

[42]   Aubry, J.P., et al. (1999) Annexin V Used for Measuring Apoptosis in the Early Events of Cellular Cytotoxicity. Cytometry, 37, 197-204.
https://doi.org/10.1002/(SICI)1097-0320(19991101)37:3<197::AID-CYTO6>3.0.CO;2-L

[43]   Rice, G.P.A., Casali, P., Merigan, T.C. and Oldstone, M.B.A. (1983) Natural Killer Cell Activity in Patients with Multiple Sclerosis Given α Interferon. Annals of Neurology, 14, 333-338.
https://doi.org/10.1002/ana.410140312

[44]   Santoli, D., et al. (1981) Cytotoxic Activity and Interferon Production by Lymphocytes from Patients with Multiple Sclerosis. Journal of Immunology, 126, 1274-1278.

[45]   Moretta, A., et al. (1991) CD69-Mediated Pathway of Lymphocyte Activation: Anti-CD69 Monoclonal Antibodies Trigger the Cytolytic Activity of Different Lymphoid Effector Cells with the Exception of Cytolytic T Lymphocytes Expressing T Cell Receptor Alpha/Beta. Journal of Experimental Medicine, 174, 1393-1398.
https://doi.org/10.1084/jem.174.6.1393

[46]   Gross, C.C., et al. (2016) Alemtuzumab Treatment Alters Circulating Innate Immune Cells in Multiple Sclerosis. Neurology-Neuroimmunology & Neuroinflammation, 3, e289.
https://doi.org/10.1212/NXI.0000000000000289

[47]   Laroni, A., et al. (2016) Dysregulation of Regulatory CD56bright NK Cells/T Cells Interactions in Multiple Sclerosis. Journal of Autoimmunity, 72, 8-18.
https://doi.org/10.1016/j.jaut.2016.04.003

[48]   Alter, G., Malenfant, J.M. and Altfeld, M. (2004) CD107a as a Functional Marker for the Identification of Natural Killer Cell Activity. Journal of Immunological Methods, 294, 15-22.
https://doi.org/10.1016/j.jim.2004.08.008

[49]   Saraste, M., Irjala, H. and Airas, L. (2007) Expansion of CD56Bright Natural Killer Cells in the Peripheral Blood of Multiple Sclerosis Patients Treated with Interferon-β. Neurological Sciences, 28, 121-126.
https://doi.org/10.1007/s10072-007-0803-3

[50]   Lee, N., et al. (2003) Expression and Characterization of Human Transient Receptor Potential Melastatin 3 (hTRPM3). Journal of Biological Chemistry, 278, 20890-20897.
https://doi.org/10.1074/jbc.M211232200

[51]   Harteneck, C. (2013) Pregnenolone Sulfate: From Steroid Metabolite to TRP Channel Ligand. Molecules, 18, 12012-12028.
https://doi.org/10.3390/molecules181012012