The neonatal Fc receptor (FcRn) is important for the metabolic fate of IgG antibodies in vivo. Analysis of the interaction between FcRn and IgG in vitro might provide insight into the structural and functional integrity of therapeutic IgG that may affect pharmacokinetics (PK) in vivo. We developed a standardized pH gradient FcRn affinity liquid chromatography method with conditions closely resembling the physiological mechanism of interaction between IgG and FcRn. This method allows the separation of molecular IgG isoforms, degradation products and engineered molecules based on their affinity to FcRn. Human FcRn was immobilized on the column and a linear pH gradient from pH 5.5 to 8.8 was applied. FcRn chromatography was used in comparison to surface plasmon resonance to characterize different monoclonal IgG preparations, e.g., oxidized or aggregated species. Wild-type and engineered IgGs were compared in vitro by FcRn chromatography and in vivo by PK studies in huFcRn transgenic mice. Analytical FcRn chromatography allows differentiation of IgG samples and variants by peak pattern and retention time profile. The method can distinguish: 1) IgGs with different Fabs, 2) oxidized from native IgG, 3) aggregates from monomer and 4) antibodies with mutations in the Fc part from wild-type IgGs. Changes in the FcRn chromatographic behavior of mutant IgGs relative to the wild-type IgG correlate to changes in the PK profile in the FcRn transgenic mice. These results demonstrate that FcRn affinity chromatography is a useful new method for the assessment of IgG integrity.

Objective. Rheumatoid arthritis therapies that are based on inhibition of a single cytokine, e.g., tumor necrosis factor ␣ (TNF␣) or interleukin-6 (IL-6), produce clinically meaningful responses in only about half of the treated patients. This study was undertaken to investigate whether combined inhibition of TNF␣ and IL-17 has additive or synergistic effects in the suppression of mesenchymal cell activation in vitro and inflammation and tissue destruction in arthritis in vivo. Methods. Cultures of human fibroblast-like synoviocytes (FLS

The cytokines SDF-1alpha and -1beta are two alternatively spliced variants of the CXC (alpha) chemokines that are highly conserved among species. SDF-1alpha was shown to function as a B-cell maturation factor, a ligand for the CXCR4 (LESTR/fusin) chemokine receptor, thereby inhibiting replication of T cell-tropic HIV-1 strains and inducing cell death in human neuronal cell lines. In this report the cloning of the rat SDF-1beta cDNA and a new SDF-1 isoform, SDF-1gamma, are presented. Using Northern blot analysis, the expression pattern of both isoforms was studied in different tissues and it is shown that during postnatal development of the central and peripheral nervous system SDF-1beta- and SDF-1gamma-mRNA expression is inversely regulated. Whilst SDF-1beta-mRNA is the predominant isoform in embryonic and early postnatal nerve tissue, SDF-1gamma-mRNA is expressed at higher levels in adulthood. After peripheral nerve lesion a transient increase in SDF-1beta-mRNA expression is observed. As revealed by in situ hybridization, neurons and Schwann cells are the main cellular sources of both SDF-1beta and SDF-1gamma mRNAs in the nervous system. Computer-assisted analysis revealed that both transcripts encode secreted peptides with putative proteolytic cleavage sites which might generate novel neuropeptides.

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We have isolated a 1.476 bp cDNA (NTII11) representing a transcript that is differntially expressed during sciatic nerve development and regeneration in the rat. Nucleotide sequence comparison indicates partial identity with a recently isolated plasmolipin cDNA. However, our clone extends the published sequence by 234 bp at the 5' end and predicts a protein that contains an additional 25 amino acids at th N-terminus. The open reading frame of th NTII11 transcript encodes a 19.4 kDa protein with four putative transmembrane domains. Northern blot analyses revealed a tissue-specific expression was confirmed by in situ hybridization, and cellular localization of plasmolipin mRNA was demonstrated in Schwann cells of the sciatic nerve and in glial cells of myelinated brain structures. The steady-state levels of plasmolipin mRNA were markedly altered (i) during development of sciatic nerve and brain. (ii) after sciatic nerve injury, and (ii) in cured Schwann cells maintained under different conditions of cell growth and arrest. Our data indicate a function of plasmolipin during myelination in the central as well as in the peripheral nervous system.