Kowarik Markus C, Dzieciatkowska Monika, Wemlinger Scott, Ritchie Alanna M, Hemmer Bernhard, Owens Gregory P, Bennett Jeffrey L
Department of Neurology, 12700 E. 19th Ave, Box B-182, Aurora, CO, 80045, USA.
Department of Biochemistry, University of Colorado Denver, Denver, CO, USA.
J Neuroinflammation. 2015 Jan 28;12:19. doi: 10.1186/s12974-015-0240-9.
Neuromyelitis optica (NMO) is a severe demyelinating disorder of the central nervous system (CNS) associated with the presence of an autoimmune antibody response (AQP4-IgG) against the water channel aquaporin-4 (AQP4). It remains unclear whether pathologic AQP4-IgG in the CNS is produced entirely by peripheral plasma cells or is generated in part by infiltrating B cells. To determine the overlap of AQP4-IgG idiotypes between the CNS and periphery, we compared the immunoglobulin G (IgG) transcriptome of cerebrospinal fluid (CSF) plasmablasts with the CSF and serum IgG proteomes in 7 AQP4-seropositive NMO patients following exacerbation.
CSF variable region Ig heavy- (VH) and light-chain (VL) transcriptome libraries were generated for each patient from CSF plasmablasts by single cell sorting, reverse transcriptase polymerase chain reaction (RT-PCR), and DNA sequencing. Recombinant antibodies were generated from clonally expanded, paired VH and VL sequences and tested for AQP4-reactivity by cell-binding assay. CSF and serum IgG fractions were searched for sequences that matched their respective CSF IgG transcriptome. Matching peptides within the same patient's CSF and serum IgG proteomes were also identified.
In each NMO patient, we recovered CSF IgG VH and VL sequences that matched germline-mutated IgG protein sequences from the patient's CSF and serum IgG proteomes. Although a modest variation was observed between patients, the overlap between the transcriptome and proteome sequences was found primarily, but not exclusively, within the CSF. More than 50% of the CSF IgG transcriptome sequences were exclusively found in the CSF IgG proteome, whereas 28% were found in both the CSF and blood IgG proteome, and 18% were found exclusively in the blood proteome. A comparable distribution was noted when only AQP4-specific IgG clones were considered. Similarly, on average, only 50% of the CSF IgG proteome matched corresponding peptide sequences in the serum.
During NMO exacerbations, a substantial fraction of the intrathecal Ig proteome is generated by an intrathecal B cell population composed of both novel and peripherally-derived clones. Intrathecal CSF B cell clones may contribute to NMO disease exacerbation and lesion formation and may be an important target for preventative therapies.
视神经脊髓炎（NMO）是一种严重的中枢神经系统（CNS）脱髓鞘疾病，与针对水通道蛋白4（AQP4）的自身免疫抗体反应（AQP4-IgG）有关。目前尚不清楚中枢神经系统中病理性AQP4-IgG是完全由外周浆细胞产生，还是部分由浸润的B细胞产生。为了确定中枢神经系统和外周之间AQP4-IgG独特型的重叠情况，我们比较了7例AQP4血清阳性NMO患者病情加重后脑脊液（CSF）浆母细胞的免疫球蛋白G（IgG）转录组与脑脊液和血清IgG蛋白质组。
通过单细胞分选、逆转录聚合酶链反应（RT-PCR）和DNA测序，从每个患者的脑脊液浆母细胞中生成脑脊液可变区Ig重链（VH）和轻链（VL）转录组文库。从克隆扩增的配对VH和VL序列中产生重组抗体，并通过细胞结合试验检测其对AQP4的反应性。在脑脊液和血清IgG组分中搜索与其各自脑脊液IgG转录组匹配的序列。还鉴定了同一患者脑脊液和血清IgG蛋白质组中的匹配肽段。
在每例NMO患者中，我们从患者的脑脊液和血清IgG蛋白质组中获得了与种系突变IgG蛋白序列匹配的脑脊液IgG VH和VL序列。虽然患者之间观察到适度的差异，但转录组和蛋白质组序列之间的重叠主要（但非唯一）存在于脑脊液中。超过50%的脑脊液IgG转录组序列仅在脑脊液IgG蛋白质组中发现，28%在脑脊液和血液IgG蛋白质组中均有发现，18%仅在血液蛋白质组中发现。仅考虑AQP4特异性IgG克隆时，也观察到类似的分布。同样，平均而言，脑脊液IgG蛋白质组中只有50%与血清中的相应肽段序列匹配。
在NMO病情加重期间，鞘内Ig蛋白质组的很大一部分是由由新克隆和外周来源克隆组成的鞘内B细胞群体产生的。鞘内脑脊液B细胞克隆可能导致NMO疾病加重和病变形成，可能是预防性治疗的重要靶点。
