A role for schwann cells in the neuroregenerative effects of a non-immunosuppressive fk506 derivative, jnj460.

UNLABELLED: FK506 and its non-immunosuppressive derivatives represent a class of pharmacological agents referred to as immunophilin ligands that have been reported to promote neuroregeneration and survival in several experimental models; however their cellular and molecular mechanisms of action have not been well established. Here we characterize a new immunophilin ligand that interacts with both FK506 binding protein 12 (FKBP12) and FKBP52, and demonstrate that JNJ460 induces neurite outgrowth from freshly explanted dorsal root ganglia (DRG) in a Schwann cell-dependent manner. Purified cultures of neurons fail to respond to these drugs, but cultures containing Schwann cells and neurons respond with neurite outgrowth, as do neurons grown in conditioned medium from JNJ460-treated Schwann cells. Using microarray analysis and a transcription reporter assay, we show that JNJ460 induces a series of transcriptional changes that occur in a temporal cascade. Among the Schwann cell-expressed genes upregulated following JNJ460 treatment is the POU transcription factor SCIP, which has been shown to regulate Schwann cell gene transcription and differentiation. JNJ460 potentiated transforming growth factor beta (TGF-beta)-induced transcriptional activation and SCIP induction in Schwann cells, by altering the interaction between FKBP12 and the TGF-beta type I receptor, TbetaR1. Finally, to test whether JNJ460 enhances neurite regeneration in vivo, we treated animals with JNJ460 for 30 days following mechanical transection of the sciatic nerve and demonstrated myelin and axonal hypertrophy at the ultrastructural level. Collectively, these data suggest that Schwann cells play an important role in the biological effects of immunophilin ligands by affecting neuron-glial signaling during regeneration. SUMMARY: The cellular and molecular mechanisms responsible for the regenerative effects of immunophilin ligands are not well understood. Here we show that the neuritogenic effects of JNJ460 in a DRG model depend on interactions between neurons and Schwann cells. Treatment of purified Schwann cells with JNJ460 alters Schwann cell gene expression, and promotes the generation of factors that act on neurons. These data indicate that Schwann cells play an important role in the actions of immunophilin ligands.

Polymorphisms in IgG Fc receptor IIB regulatory regions associated with autoimmune susceptibility.

Autoimmune diseases involve multiple genes. While functions of these genes are largely unknown, some may be related to an intrinsic hyperresponsiveness of B cells. B-cell responses are controlled by signaling thresholds through the B-cell antigen receptor (BCR) complex. The B1 isoform of type II IgG Fc receptors (FcgammaRIIB1) is exclusively expressed on B cells and serves as a negative regulator for inhibiting BCR-elicited activation. Thus, its allelic variants associated with functional deficits could be examined for possible associations with susceptibility to autoimmune diseases. We found that there are three types of polymorphisms in the reported FcgammaRIIB transcription regulatory regions in mouse strains. Compared to normal healthy mouse strains (group III), autoimmune disease-prone strains (group I) share three deletion sites: two in the promoter region and one in the third intron. Strains (group II) that per se are not autoimmune-prone, but have potentials to accelerate autoimmune diseases share two deletion sites in the third intron: one identical to that in group I and the other unique to group II. These polymorphisms correlated well with extents of down-regulation of FcgammaRIIB1 expression in germinal-center B cells upon stimulation with antigens and up-regulation of IgG antibody responses. Our data imply that these FcgammaRIIB polymorphisms are selected evolutionarily for natural defense against pathogens, and that such polymorphisms may, in turn, form the basis of one aspect of autoimmune susceptibility.

Genetically determined aberrant down-regulation of FcgammaRIIB1 in germinal center B cells associated with hyper-IgG and IgG autoantibodies in murine systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a multigenic disease associated with IgG hypergammaglobulinemia, IgG anti-nuclear antibodies and immune complex (IC)-type glomerulonephritis. In both human and murine SLE, one susceptibility allele has been mapped to the interval linked to the IgG Fc receptor II (FcgammaRII) gene on chromosome 1. In spontaneous SLE models of NZB and (NZB x NZW) F(1) mice, expression of FcgammaRIIB1, which acts as a negative regulator for B cells, was abnormally down-regulated in follicular germinal center B cells from aged mice, compared to findings in non-SLE NZW, while levels in non-germinal center B cells were practically identical. Such strain differences were also evident in young mice upon in vivo stimulation with foreign antigens. In the FcgammaRIIB promoter region, the NZB allele has two deletion sites, including transcription factor-binding sites. Analyses using (NZB x NZW) F(1) x NZW backcross mice showed that this NZB allele was significantly linked to hyper-IgG, irrespective of the MHC haplotype, while high levels of IgG antibodies specific for DNA were regulated by a combinatorial effect of the F(1)-unique MHC haplotype and the NZB FcgammaRIIB allele. Therefore, the FcgammaRIIB promoter polymorphism may possibly predispose to SLE through germinal center B cells abnormally down-regulating FcgammaRIIB1 expression upon autoantigen stimulations and thus escaping negative signals for IgG production.

Genetic polymorphism of murine tissue plasminogen activator associated with antiphospholipid syndrome.

In a subset of patients with systemic lupus erythematosus (SLE), antiphospholipid syndrome characterized by thrombocytopenia, thrombosis, recurrent abortion and antiphospholipid antibodies develops. Male (NZW x BXSB) F1 mice are widely used as a model for SLE-associated antiphospholipid syndrome. Our earlier genetic studies showed that one susceptibility allele for thrombocytopenia and associated IgG platelet-binding autoantibodies in male (NZW x BXSB) F1 mice was linked to the BXSB-type polymorphic microsatellite D8Mit96, located in proximity to the gene Plat for tissue-type plasminogen activator (t-PA). In the present studies, sequence analyses for structural and promoter regions of Plat revealed a single nucleotide polymorphism encoding a catalytic domain of t-PA, with an amino acid substitution of anionic Glu366 in NZW for a cationic Lys in BXSB. Progeny studies using NZW x (NZW x BXSB) F1 male backcross mice showed that the BXSB Plat allele was significantly associated with high levels of both platelet-binding antibodies and thrombocytopenia. Furthermore, these two traits appeared to be regulated by a complementary effect of two BXSB alleles; one is linked to Plat and the other to the H-2 complex and the gene for plasminogen. Thus, the BXSB-type Plat may be one susceptibility allele for the multigenic antiphospholipid syndrome seen in (NZW x BXSB) F1 mice. Potential mechanisms are discussed.