Page 32 - The autoimmune hypothesis of narcolepsy and its unexplored clinical features M.S. Schinkelshoek

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Chapter 2
2019b, Lind et al., 2019). These reports started a discussion about whether different immunological mechanisms may be involved in post-H1N1 NT1. Since these studies are small and only address HLA-DQB1-associations in a cohort of post-H1N1 NT1 patients and a healthy control population, our aim is to compare HLA-DQB1-associations in NT1 patients with disease onset before and after the 2009 H1N1 pandemic in a large Dutch cohort to assess whether differences can be seen which would test the hypothesis that sporadic and post-H1N1 NT1 should be regarded as separate entities based on different immunological mechanisms leading to the disease.
Materials and methods
Participants
We included all NT1 patients of European ancestry who, between 2005 and 2019, were HLA-genotyped for clinical care in the laboratory of the Leiden University Medical Center. Patients came from the sleep clinic of the Department of Neurology, Leiden University Medical Center, the Sleep-Wake Centre of Stichting Epilepsie Instellingen Nederland (SEIN), Heemstede and Zwolle, and the Sleep Centre of Kempenhaeghe, Heeze. All patients were diagnosed with either narcolepsy with cataplexy or NT1 according to the International Classification of Sleep Disorders (ICSD-2 or ICSD-3; (ICSD, 2005, ICSD, 2014)). Age at symptom onset was assessed in all patients to separate those with symptom onset in childhood (<16 years at symptom onset) or adulthood, and we also noted whether the symptoms of NT1 started before or after the 2009 H1N1 pandemic to be able to compare HLA-DQB1-associations before and after the H1N1 pandemic. We also included a cohort of HLA-DQB1*06:02 positive non- related healthy individuals from a panel of randomly selected Dutch individuals (van Rooijen et al., 2012) to be able to assess HLA associations in NT1.
HLA typing
HLA genotyping for HLA-DR and HLA-DQ was performed with a reversed approach of the PCR-sequence-specific oligonucleotide probe technique previously described (Verduyn et al., 1993). If a rare DRB1-DQB1 was found, typing results were confirmed by PCR-SBT, using the SBT Excellerator HLA- DRB and -DQB kit (Genome Diagnostics, Utrecht, The Netherlands).