Tissue Antigens ISSN 0001-2815
LETTER TO THE EDITOR
Are KIR and HLA class I genes associated with schizophrenia? 1 , D. Frydecka2 , I. Nowak1 , E. Majorczyk1 , D. Senitzer3 , P. Piotrowski2 , A. Beszlej2 , ´ A. Wisniewski 2 1 ´ A. Kiejna & P. Kusnierczyk
1 Laboratory of Immunogenetics and Tissue Immunology, Department of Clinical Immunology, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wrocław, Poland 2 Department of Psychiatry, Wrocław Medical University, Wrocław, Poland 3 City of Hope Comprehensive Cancer Center, Hematology and Hematopoietic Cell Transplantation, Duarte, CA USA
Schizophrenia is a chronic and very serious psychiatric disorder, the etiology and pathophysiology of which has not been firmly established. It is multifactorial, and genetic, immunologic, and environmental factors play a role (1–3). It has been shown that infections and autoimmune diseases increase the risk of schizophrenia (4, 5). Natural killer (NK) cells are elements of innate immunity, important for the defense against viral infections and oncogenesis by lysis of infected and transformed cells, respectively, and they also contribute to autoimmune diseases (6). The activity of NK cells in schizophrenia patients is increased, which may suggest a contribution of these cells to the disease (7, 8). Activation of NK cells depends on a balance of activating and inhibitory signals from cell surface receptors. Among these receptors, the most genetically polymorphic are killer cell immunoglobulin-like receptors (KIRs). These receptors are encoded by 15 genes on chromosome 19, but individuals differ in number and kind (activating vs inhibitory) of KIR genes, which influences their susceptibility to viral infections, neoplasms, and autoimmune diseases (9). Ligands of KIRs, when known, are specific epitopes on human leukocyte antigen (HLA) class I molecules (6): all HLA-C allotypes fall into C1 or C2 group (asparagine and lysine, respectively, at position 80) recognized by KIR2DL2/KIR2DL3 and KIR2DL1/KIR2DS1, respectively. HLA-Bw4 is an epitope present on roughly half of HLA-B allotypes, as well as on some HLA-A molecules, and is recognized by KIR3DL1. Some other KIRs bind also some HLA class I molecules (10). We hypothesized that some KIR and HLA class I genes might be associated with schizophrenia. Here, we report the results of our comparison of KIR genes and HLA class I-encoded KIR ligand frequencies in patients with schizophrenia and healthy control individuals in the Polish Caucasian population. Our case–control study enrolled 200 unrelated schizophrenic patients (113 women and 87 men). The mean age at blood sampling was 37.8 years [standard deviation (SD): 12.25, range: 21–75]. All patients were diagnosed at the Wroclaw Medical University Hospital according to ICD-10 (11) and DSM-IV (12) criteria based on interview data and hospital case notes. In addition to unstructured interviews and review of medical records, the patients were evaluated for lifetime psychotic symptomatology using the Operational Criteria for © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Tissue Antigens, 2014, 84, 503–504
Psychotic Illness (OPCRIT) checklist, which provides a polydiagnostic categorical and dimensional approach to diagnosis of schizophrenia (13). Patients having a history of traumatic brain injury, neurologic disorders, and substance abuse were excluded from the study by detailed medical examination. The control group consisted of 561 (326 women and 235 men) healthy subjects. They were volunteers declaring good health condition and lack of psychiatric abnormalities. The mean age at blood sampling was 39.3 years (SD: 14.67, range: 19–83). The study was approved by the Bioethics Committee of the Wroclaw Medical University. Signed informed consent was obtained from all the subjects. DNA isolation was done using Invisorb Spin Blood Midi Kit (Invitek, Berlin, Germany) following the manufacturer’s instructions, or according to Gustincich et al. (14). KIR genotyping was performed by single (15) or multiplex (16) polymerase chain reaction using sequence-specific primers. KIR ligands were established using primers for HLA-C C1 and C2 (17) as well as a commercial kit (Olerup SSP® KIR HLA Ligand) for HLA-CAsparagine80 – (C1), HLA-CLysine80 – (C2), HLA-ABw4+ , HLA-B Bw4Isoleucine80 , and HLA-BThreonine80 (Olerup SSP AB, Saltsjobaden, Sweden). Statistical analysis was performed as follows: global test for difference between two sets of k dependent proportions i.e. ‖x −x ‖ x1 = (p11 , … , p1k )T and x2 = (p21 , … , p2k )T was T = SE 1 2 2 , ‖x1 −x2 ‖2 where distribution of T statistic was estimated numerically. √ Proportions were transformated to p′ = sin−1 ( p). Odds ratio and its 95% confidence interval was used as a measure of effect size. Comparison of all studied KIR gene frequencies in 200 patients with schizophrenia and 561 healthy control individuals did not reveal any significant difference (pglobal = 0.15). Similarly, no differences were found in distribution of KIR gene ligands (pglobal = 0.73 and 0.67 for the HLA-C1, -C2 and for the HLA-ABw4+ , HLA-B Bw4Ile , and HLA-B Bw4Thr , respectively). These results seem contradictory to those of the recent GWAS genome-wide association study (18), where a strong association of HLA-C*01:02 allele with schizophrenia was observed. The reason for this discrepancy may lay in the fact that (i) we typed only for C1 and C2 epitopes of HLA-C but not for particular HLA-C alleles, whereas the HLA-C*01:02 allele is rare in Caucasians including Poles (19) and constitutes 503
Letter to the Editor
only a small fraction of all C1 alleles; (ii) we examined much lower numbers of patients and controls. Moreover, also analysis of different KIR-ligand combinations revealed no significant differences between patients and controls. In conclusion, we have not detected any associations of KIR and KIR ligand genes, or combinations thereof, with susceptibility to schizophrenia. Nevertheless, to our knowledge, this is the first report on KIR and ligand genetic contribution to schizophrenia. Acknowledgments
This work was supported by the Polish National Science Centre grant no. N402 455238. We would like to express our sincere thanks to the patients and to the healthy volunteers for donating blood for our study and to MSc. Maciej Sobczy´nski for his help in statistical analysis. Conflict of interest
The authors have declared no conflicting interests. Correspondence ´ Dr Andrzej Wisniewski Tel: +48 71 3371172 232 Fax: +48-71-337 2171 e-mail: [email protected] and ´ Prof. Dr Piotr Kusnierczyk Laboratory of Immunogenetics and Tissue Immunology, Department of Clinical Immunology Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences Wrocław Poland Tel: +48 71 3709976 Fax: +48-71-337 2171 e-mail: [email protected] doi: 10.1111/tan.12423
References 1. Van Os J, Rutten BPF, Poulton R. Gene-environment interactions in schizophrenia: review of epidemiological findings and future directions. Schizophr Bull 2008: 34: 1066–82. 2. Takahashi S. Heterogeneity of schizophrenia: genetic and symptomatic factors. Am J Med Genet B Neuropsychiatr Genet 2013: 162B: 648–52. 3. Debnath M, Cannon DM, Venkatasubramanian G. Variation in the major histocompatibility complex [MHC] gene family in schizophrenia: associations and functional implications. Prog Neuropsychopharmacol Biol Psychiatry 2013: 42: 49–62.
504
´ A. Wisniewski et al.
4. Benros ME, Nielsen PR, Nordentoft M, Eaton WW, Dalton SO, Mortensen PB. Autoimmune diseases and severe infections as risk factors for schizophrenia: a 30-year population-based register study. Am J Psychiatry 2011: 168: 1303–10. 5. Chen SJ, Chao YL, Chen CY et al. Prevalence of autoimmune diseases in in-patients with schizophrenia: nationwide population-based study. Br J Psychiatry 2012: 200: 374–80. 6. Parham P. MHC class I molecules and KIRs in human history, health and survival. Nat Rev Immunol 2005: 5: 201–14. 7. Yovel G, Sirota P, Mazeh D, Shakhar G, Rosenne E, Ben-Eliyahu S. Higher natural killer cell activity in schizophrenic patients: the impact of serum factors, medication, and smoking. Brain Behav Immun 2000: 14: 153–69. 8. Vodjgani M, Matloubi H, Nasehi AA et al. Increased natural killer cells activity in schizophrenic patients. IJI 2005: 2: 111–6. 9. Rajalingam R. Human diversity of killer cell immunoglobulin-like receptors and disease. Korean J Hematol 2011: 46: 216–28. 10. Parham P, Norman PJ, Abi-Rached L, Guethlein LA. Human-specific evolution of killer cell immunoglobulin-like receptor recognition of major histocompatibility complex class I molecules. Philos Trans R Soc Lond B Biol Sci 2012: 367: 800–11. 11. World Health Organization. The ICD-10 Classification of Mental and Behavioural Disorders: Clinical Descriptions and Diagnostic Guidelines. Geneva: WHO, 1992. 12. American Psychiatric Association. Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV). Washington DC: American Psychiatric Association, 1994, 143–7. 13. McGuffin P, Farmer A, Harvey I. A polydiagnostic application of operational criteria in studies of psychotic illness. Development and reliability of the OPCRIT system. Arch Gen Psychiatry 1991: 48: 764–70. 14. Gustincich S, Manfioletti G, Del Sal G, Schneider C, Carninci P. A fast method for high-quality genomic DNA extraction from whole human blood. Biotechniques 1991: 11: 298–302. 15. Vilches C, Castaño J, Gómez-Lozano N, Estefanía E. Facilitation of KIR genotyping by a PCR-SSP method that amplifies short DNA fragments. Tissue Antigens 2007: 70: 415–22. 16. Sun JY, Gaidulis L, Miller MM et al. Development of a multiplex PCR-SSP method for killer-cell immunoglobulin-like receptor genotyping. Tissue Antigens 2004: 64: 462–8. 17. Frohn C, Schlenke P, Ebel B, Dannenberg C, Bein G, Kirchner H. DNA typing for natural killer cell inhibiting HLA-Cw groups NK1 and NK2 by PCR-SSP. J Immunol Methods 1998: 218: 155–60. 18. Irish Schizophrenia Genomics Consortium and the Wellcome Trust Case Control Consortium 2. Genome-wide association study implicates HLA-C*01:02 as a risk factor at the major histocompatibility complex locus in schizophrenia. Biol Psychiatry 2012: 15: 620–8. 19. Schmidt AH, Solloch UV, Pingel J et al. High-resolution human leukocyte antigen allele and haplotype frequencies of the Polish population based on 20,653 stem cell donors. Hum Immunol 2011: 72: 558–65.
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Tissue Antigens, 2014, 84, 503–504
Copyright of Tissue Antigens is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
LETTER TO THE EDITOR
Are KIR and HLA class I genes associated with schizophrenia? 1 , D. Frydecka2 , I. Nowak1 , E. Majorczyk1 , D. Senitzer3 , P. Piotrowski2 , A. Beszlej2 , ´ A. Wisniewski 2 1 ´ A. Kiejna & P. Kusnierczyk
1 Laboratory of Immunogenetics and Tissue Immunology, Department of Clinical Immunology, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wrocław, Poland 2 Department of Psychiatry, Wrocław Medical University, Wrocław, Poland 3 City of Hope Comprehensive Cancer Center, Hematology and Hematopoietic Cell Transplantation, Duarte, CA USA
Schizophrenia is a chronic and very serious psychiatric disorder, the etiology and pathophysiology of which has not been firmly established. It is multifactorial, and genetic, immunologic, and environmental factors play a role (1–3). It has been shown that infections and autoimmune diseases increase the risk of schizophrenia (4, 5). Natural killer (NK) cells are elements of innate immunity, important for the defense against viral infections and oncogenesis by lysis of infected and transformed cells, respectively, and they also contribute to autoimmune diseases (6). The activity of NK cells in schizophrenia patients is increased, which may suggest a contribution of these cells to the disease (7, 8). Activation of NK cells depends on a balance of activating and inhibitory signals from cell surface receptors. Among these receptors, the most genetically polymorphic are killer cell immunoglobulin-like receptors (KIRs). These receptors are encoded by 15 genes on chromosome 19, but individuals differ in number and kind (activating vs inhibitory) of KIR genes, which influences their susceptibility to viral infections, neoplasms, and autoimmune diseases (9). Ligands of KIRs, when known, are specific epitopes on human leukocyte antigen (HLA) class I molecules (6): all HLA-C allotypes fall into C1 or C2 group (asparagine and lysine, respectively, at position 80) recognized by KIR2DL2/KIR2DL3 and KIR2DL1/KIR2DS1, respectively. HLA-Bw4 is an epitope present on roughly half of HLA-B allotypes, as well as on some HLA-A molecules, and is recognized by KIR3DL1. Some other KIRs bind also some HLA class I molecules (10). We hypothesized that some KIR and HLA class I genes might be associated with schizophrenia. Here, we report the results of our comparison of KIR genes and HLA class I-encoded KIR ligand frequencies in patients with schizophrenia and healthy control individuals in the Polish Caucasian population. Our case–control study enrolled 200 unrelated schizophrenic patients (113 women and 87 men). The mean age at blood sampling was 37.8 years [standard deviation (SD): 12.25, range: 21–75]. All patients were diagnosed at the Wroclaw Medical University Hospital according to ICD-10 (11) and DSM-IV (12) criteria based on interview data and hospital case notes. In addition to unstructured interviews and review of medical records, the patients were evaluated for lifetime psychotic symptomatology using the Operational Criteria for © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Tissue Antigens, 2014, 84, 503–504
Psychotic Illness (OPCRIT) checklist, which provides a polydiagnostic categorical and dimensional approach to diagnosis of schizophrenia (13). Patients having a history of traumatic brain injury, neurologic disorders, and substance abuse were excluded from the study by detailed medical examination. The control group consisted of 561 (326 women and 235 men) healthy subjects. They were volunteers declaring good health condition and lack of psychiatric abnormalities. The mean age at blood sampling was 39.3 years (SD: 14.67, range: 19–83). The study was approved by the Bioethics Committee of the Wroclaw Medical University. Signed informed consent was obtained from all the subjects. DNA isolation was done using Invisorb Spin Blood Midi Kit (Invitek, Berlin, Germany) following the manufacturer’s instructions, or according to Gustincich et al. (14). KIR genotyping was performed by single (15) or multiplex (16) polymerase chain reaction using sequence-specific primers. KIR ligands were established using primers for HLA-C C1 and C2 (17) as well as a commercial kit (Olerup SSP® KIR HLA Ligand) for HLA-CAsparagine80 – (C1), HLA-CLysine80 – (C2), HLA-ABw4+ , HLA-B Bw4Isoleucine80 , and HLA-BThreonine80 (Olerup SSP AB, Saltsjobaden, Sweden). Statistical analysis was performed as follows: global test for difference between two sets of k dependent proportions i.e. ‖x −x ‖ x1 = (p11 , … , p1k )T and x2 = (p21 , … , p2k )T was T = SE 1 2 2 , ‖x1 −x2 ‖2 where distribution of T statistic was estimated numerically. √ Proportions were transformated to p′ = sin−1 ( p). Odds ratio and its 95% confidence interval was used as a measure of effect size. Comparison of all studied KIR gene frequencies in 200 patients with schizophrenia and 561 healthy control individuals did not reveal any significant difference (pglobal = 0.15). Similarly, no differences were found in distribution of KIR gene ligands (pglobal = 0.73 and 0.67 for the HLA-C1, -C2 and for the HLA-ABw4+ , HLA-B Bw4Ile , and HLA-B Bw4Thr , respectively). These results seem contradictory to those of the recent GWAS genome-wide association study (18), where a strong association of HLA-C*01:02 allele with schizophrenia was observed. The reason for this discrepancy may lay in the fact that (i) we typed only for C1 and C2 epitopes of HLA-C but not for particular HLA-C alleles, whereas the HLA-C*01:02 allele is rare in Caucasians including Poles (19) and constitutes 503
Letter to the Editor
only a small fraction of all C1 alleles; (ii) we examined much lower numbers of patients and controls. Moreover, also analysis of different KIR-ligand combinations revealed no significant differences between patients and controls. In conclusion, we have not detected any associations of KIR and KIR ligand genes, or combinations thereof, with susceptibility to schizophrenia. Nevertheless, to our knowledge, this is the first report on KIR and ligand genetic contribution to schizophrenia. Acknowledgments
This work was supported by the Polish National Science Centre grant no. N402 455238. We would like to express our sincere thanks to the patients and to the healthy volunteers for donating blood for our study and to MSc. Maciej Sobczy´nski for his help in statistical analysis. Conflict of interest
The authors have declared no conflicting interests. Correspondence ´ Dr Andrzej Wisniewski Tel: +48 71 3371172 232 Fax: +48-71-337 2171 e-mail: [email protected] and ´ Prof. Dr Piotr Kusnierczyk Laboratory of Immunogenetics and Tissue Immunology, Department of Clinical Immunology Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences Wrocław Poland Tel: +48 71 3709976 Fax: +48-71-337 2171 e-mail: [email protected] doi: 10.1111/tan.12423
References 1. Van Os J, Rutten BPF, Poulton R. Gene-environment interactions in schizophrenia: review of epidemiological findings and future directions. Schizophr Bull 2008: 34: 1066–82. 2. Takahashi S. Heterogeneity of schizophrenia: genetic and symptomatic factors. Am J Med Genet B Neuropsychiatr Genet 2013: 162B: 648–52. 3. Debnath M, Cannon DM, Venkatasubramanian G. Variation in the major histocompatibility complex [MHC] gene family in schizophrenia: associations and functional implications. Prog Neuropsychopharmacol Biol Psychiatry 2013: 42: 49–62.
504
´ A. Wisniewski et al.
4. Benros ME, Nielsen PR, Nordentoft M, Eaton WW, Dalton SO, Mortensen PB. Autoimmune diseases and severe infections as risk factors for schizophrenia: a 30-year population-based register study. Am J Psychiatry 2011: 168: 1303–10. 5. Chen SJ, Chao YL, Chen CY et al. Prevalence of autoimmune diseases in in-patients with schizophrenia: nationwide population-based study. Br J Psychiatry 2012: 200: 374–80. 6. Parham P. MHC class I molecules and KIRs in human history, health and survival. Nat Rev Immunol 2005: 5: 201–14. 7. Yovel G, Sirota P, Mazeh D, Shakhar G, Rosenne E, Ben-Eliyahu S. Higher natural killer cell activity in schizophrenic patients: the impact of serum factors, medication, and smoking. Brain Behav Immun 2000: 14: 153–69. 8. Vodjgani M, Matloubi H, Nasehi AA et al. Increased natural killer cells activity in schizophrenic patients. IJI 2005: 2: 111–6. 9. Rajalingam R. Human diversity of killer cell immunoglobulin-like receptors and disease. Korean J Hematol 2011: 46: 216–28. 10. Parham P, Norman PJ, Abi-Rached L, Guethlein LA. Human-specific evolution of killer cell immunoglobulin-like receptor recognition of major histocompatibility complex class I molecules. Philos Trans R Soc Lond B Biol Sci 2012: 367: 800–11. 11. World Health Organization. The ICD-10 Classification of Mental and Behavioural Disorders: Clinical Descriptions and Diagnostic Guidelines. Geneva: WHO, 1992. 12. American Psychiatric Association. Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV). Washington DC: American Psychiatric Association, 1994, 143–7. 13. McGuffin P, Farmer A, Harvey I. A polydiagnostic application of operational criteria in studies of psychotic illness. Development and reliability of the OPCRIT system. Arch Gen Psychiatry 1991: 48: 764–70. 14. Gustincich S, Manfioletti G, Del Sal G, Schneider C, Carninci P. A fast method for high-quality genomic DNA extraction from whole human blood. Biotechniques 1991: 11: 298–302. 15. Vilches C, Castaño J, Gómez-Lozano N, Estefanía E. Facilitation of KIR genotyping by a PCR-SSP method that amplifies short DNA fragments. Tissue Antigens 2007: 70: 415–22. 16. Sun JY, Gaidulis L, Miller MM et al. Development of a multiplex PCR-SSP method for killer-cell immunoglobulin-like receptor genotyping. Tissue Antigens 2004: 64: 462–8. 17. Frohn C, Schlenke P, Ebel B, Dannenberg C, Bein G, Kirchner H. DNA typing for natural killer cell inhibiting HLA-Cw groups NK1 and NK2 by PCR-SSP. J Immunol Methods 1998: 218: 155–60. 18. Irish Schizophrenia Genomics Consortium and the Wellcome Trust Case Control Consortium 2. Genome-wide association study implicates HLA-C*01:02 as a risk factor at the major histocompatibility complex locus in schizophrenia. Biol Psychiatry 2012: 15: 620–8. 19. Schmidt AH, Solloch UV, Pingel J et al. High-resolution human leukocyte antigen allele and haplotype frequencies of the Polish population based on 20,653 stem cell donors. Hum Immunol 2011: 72: 558–65.
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Tissue Antigens, 2014, 84, 503–504
Copyright of Tissue Antigens is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
