Previous reports have demonstrated the ability of antigen-presenting cells (APCs), genetically modified to express Fas ligand (FasL), to inhibit T-cell responses through the induction of apoptosis of antigen-specific T cells. Here we have examined the ability of primary mouse bone marrow-derived dendritic cells (DCs), genetically modified by adenoviral infection to express FasL, to inhibit progression of established collagen-induced arthritis (CIA) in DBA/1 mice. Systemic injection of DC/FasL into mice with established CIA resulted in substantial disease amelioration as determined by analysis of paw swelling, arthritic index, and number of arthritic paws. Moreover, a single injection of DC/FasL resulted in extended suppression of disease. We also demonstrate that treatment of arthritic mice with DC/FasL suppressed interferon-gamma (IFN-gamma) production from spleen-derived lymphocytes and reduced T-cell proliferation following collagen stimulation without affecting the levels of anti-collagen antibody isotypes. These results demonstrate that systemic administration of DC/FasL is able to suppress collagen-reactive T cells, resulting in effective and sustained treatment of established CIA.

Biodegradable and non-toxic multi-block copolymers based on poly(L-lysine) and poly(ethylene glycol) were synthesized. Synthesized copolymers showed almost negligible cytotoxicity above 95% cell viability and transfection efficiency compared to the PLL homopolymer with molecular weight of 25,700. Biodegradation under physiological conditions revealed that the molecular weight of copolymers decreased to 20% of the initial molecular weight within 72 h. Transfection efficiencies of copolymers were not affected by the presence of serum, while that of PLL homopolymer decreased to the level of naked DNA in the presence of serum. Based on the results, the new copolymers are believed to be a potentially efficient carrier for the delivery of bioactive agents.

Intra-articular expression of antagonists of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) in arthritic rabbit knee and mouse ankle joints by direct adenoviral-mediated intraarticular delivery results in amelioration of disease pathology in both the treated and contralateral untreated joints. Previous experiments suggest that direct adenoviral infection of resident antigen-presenting cells (APCs) and subsequent traveling of these cells to other sites of inflammation and lymph nodes might be responsible for this "contralateral effect." To determine whether genetic modification of APCs is required for the contralateral effect, we have used an ex vivo approach utilizing genetically modified fibroblasts to express IL-1 receptor antagonist protein (IL-1Ra) and soluble TNF-alpha receptor (sTNFR) locally in arthritic joints. Retroviral vectors carrying IL-1Ra, sTNFR-Ig, or both genes together were used to stably infect autologous rabbit fibroblasts that were then injected intra-articularly into arthritic rabbit knee joints. The intra-articular delivery of either IL-1Ra- or sTNFR-Ig-expressing fibroblasts was antiinflammatory and chondro-protective in both the injected and noninjected contralateral joints. In addition, we demonstrate that the co-delivery of both antagonists in combination results in a synergistic effect in disease amelioration in both the treated and nontreated joints. These ex vivo results suggest that trafficking of vector-modified inflammatory cells is not the main mechanism responsible for the observed distal spread of the therapeutic effect. Moreover, the results demonstrate that local, ex vivo gene therapy for arthritis could be effective in blocking pathologies within untreated, distant arthritic joints.

Coprinus congregutushas a membrane-associated lactase which is not secreted into the culture media. A mutant monokaryon has been generated by UV irradiation which shows slower radial growth than the wild-type (72.7%) but which secretes lactase. The lactase of the mutant was compared with that of wild-type by native PAGE analysis, and showed identical mobility.