Clonally expanded plasma cells in the cerebrospinal fluid of patients with central nervous system autoimmune demyelination produce "oligoclonal bands".

Clonally expanded plasma cells (cePC) and oligoclonal IgG (oligoclonal bands, OCB) in the cerebrospinal fluid (CSF) suggest an involvement of B cell mechanisms in autoimmune CNS demyelination. Due to their CSF-restricted occurrence, OCB are commonly believed to be the products of B cells inside the borders of the blood brain barrier. A comparison of CSF cell Ig transcriptomes and CSF-Ig proteomes recently demonstrated, that in multiple sclerosis patients CSF cells are the origin of CSF immunoglobulins. We expand these findings by applying anti-idiotypic antibodies to detect specific heavy chain CDR3 idiotopes of cePC-produced antibodies amongst OCB in the CSF of a patient each with MS and acute disseminated encephalomyelitis.

p300-mediated acetylation of the Rothmund-Thomson-syndrome gene product RECQL4 regulates its subcellular localization.

RECQL4 belongs to the conserved RecQ family of DNA helicases, members of which play important roles in the maintenance of genome stability in all organisms that have been examined. Although genetic alterations in the RECQL4 gene are reported to be associated with three autosomal recessive disorders (Rothmund-Thomson, RAPADILINO and Baller-Gerold syndromes), the molecular role of RECQL4 still remains poorly understood. Here, we show that RECQL4 specifically interacts with the histone acetyltransferase p300 (also known as p300 HAT), both in vivo and in vitro, and that p300 acetylates one or more of the lysine residues at positions 376, 380, 382, 385 and 386 of RECQL4. Furthermore, we report that these five lysine residues lie within a short motif of 30 amino acids that is essential for the nuclear localization of RECQL4. Remarkably, the acetylation of RECQL4 by p300 in vivo leads to a significant shift of a proportion of RECQL4 protein from the nucleus to the cytoplasm. This accumulation of the acetylated RECQL4 is a result of its inability to be imported into the nucleus. Our results provide the first evidence of a post-translational modification of the RECQL4 protein, and suggest that acetylation of RECQL4 by p300 regulates the trafficking of RECQL4 between the nucleus and the cytoplasm.

Clonally expanded plasma cells in the cerebrospinal fluid of MS patients produce myelin-specific antibodies.

Clonally expanded plasma cells (cePC) and their presumed products, oligoclonal immunoglobulin G bands (OCB), are characteristic findings in the cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS). While cePC and OCB strongly suggest an involvement of B cell-dependent immune mechanisms in the pathogenesis of MS, their actual pathological relevance and target antigens remain unknown. To further understand the potential role played by cePC, we generated a panel of monoclonal antibodies (MS-mAb) from CSF-derived cePC from four patients with early or definite MS. Single-cell RT-PCR of correctly paired heavy and light chain immunoglobulin genes from individual cePC ensured the subsequent resurrection of their original antigen specificity. Immunofluorescence stainings of MS lesion tissue with MS-mAb revealed myelin reactivity in the cePC repertoire of all four patients and intracellular filament reactivity in one patient. While myelin staining by MS-mAb was only rarely detectable in non-MS CNS white matter tissue, it was greatly enhanced at the edge of demyelinating lesions in MS brain tissue. Our findings provide conclusive evidence for the presence of an antigen-driven B cell response in the CSF of MS patients directed against epitopes present in areas of myelin degradation.

Pathogen specificity and autoimmunity are distinct features of antigen-driven immune responses in neuroborreliosis.

Neuroborreliosis (NB) is a chronic infectious disease of the central nervous system (CNS) caused by a tick-borne spirochete, Borrelia burgdorferi. In addition to direct effects of the causative infectious agent, additional immunity-mediated mechanisms are thought to play a role in the CNS pathology of NB. In order to further understand the involvement of humoral immune mechanisms in NB, we dissected the intrathecal antibody responses down to the single-plasma-cell level. Starting with single-cell reverse transcription-PCR of fluorescence-activated cell sorter-sorted cerebrospinal fluid plasma cells from an NB patient, we identified expanded clones and resurrected the antigen specificity of their secreted antibodies through recombinant expression of the correctly paired immunoglobulin heavy- and light-chain genes as monoclonal antibodies (MAbs). As expected, we found specificity for the causative infectious agent, B. burgdorferi, among the clonally expanded plasma cell (cePC)-derived MAbs. However, from an independent cePC of the same patient, we could derive MAbs specific for human CNS myelin, without detectable cross-reactivity with B. burgdorferi antigens. While reactivity against B. burgdorferi is a known feature of humoral immune responses in NB, we show (i) that immune responses specific for self antigens may be a distinct feature of CNS infections independent of pathogen reactivity and (ii) that humoral autoimmunity in NB (since found in cePC) is the result of a truly antigen-driven immune response. Our findings indicate that in NB mechanisms may be at play that induce distinct immune responses specific for pathogen and self antigens independent from "molecular mimicry."

ADAMTS1 proteinase is up-regulated in wounded skin and regulates migration of fibroblasts and endothelial cells.

The metalloproteinase ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin motifs) is induced under inflammatory conditions, and it is also a potent inhibitor of angiogenesis. Due to these properties, we speculated about the role of ADAMTS1 in cutaneous wound repair. Here we have shown up-regulation of ADAMTS1 expression in wounds of normal and particularly of healing-impaired genetically diabetic mice. Immunofluorescence staining identified macrophages as the source of ADAMTS1 in early wounds, whereas keratinocytes and fibroblasts produce this protein at later stages of wound healing. The distribution of ADAMTS1 in the normal and wounded epidermis, its regulation in cultured keratinocytes, as well as the skin phenotype of ADAMTS1 knock-out mice suggests a role of this metalloproteinase in keratinocyte differentiation. Furthermore, we provide evidence for a novel dual function of ADAMTS1 in fibroblast migration; although low concentrations of this protein stimulate fibroblast migration via its proteolytic activity, high concentrations inhibit this process because of binding to fibroblast growth factor-2 and subsequent inhibition of its promotogenic activity. Similar effects were also observed with endothelial cells. Taken together, our results suggest a role of ADAMTS1 in keratinocyte differentiation and migration of fibroblasts and endothelial cells in healing skin wounds.