Granzyme K mediates IL-23-dependent inflammation and keratinocyte proliferation in psoriasis

Psoriasis is an inflammatory disease with systemic manifestations that most commonly presents as itchy, erythematous, scaly plaques on extensor surfaces. Activation of the IL-23/IL-17 pro-inflammatory signaling pathway is a hallmark of psoriasis and its inhibition is key to clinical management. Granzyme K (GzmK) is an immune cell-secreted serine protease elevated in inflammatory and proliferative skin conditions. In the present study, human psoriasis lesions exhibited elevated GzmK levels compared to non-lesional psoriasis and healthy control skin. In an established murine model of imiquimod (IMQ)-induced psoriasis, genetic loss of GzmK significantly reduced disease severity, as determined by delayed plaque formation, decreased erythema and desquamation, reduced epidermal thickness, and inflammatory infiltrate. Molecular characterization in vitro revealed that GzmK contributed to macrophage secretion of IL-23 as well as PAR-1-dependent keratinocyte proliferation. These findings demonstrate that GzmK enhances IL-23-driven inflammation as well as keratinocyte proliferation to exacerbate psoriasis severity

Fibroblast cell-based therapy prevents induction of alopecia areata in an experimental model

YesAlopecia areata (AA) is an autoimmune hair loss disease with infiltration of proinflammatory cells into hair follicles. Current therapeutic regimens are unsatisfactory mainly because of the potential for side effects and/or limited efficacy. Here we report that cultured, transduced fibroblasts, which express the immunomodulatory molecule indoleamine 2,3-dioxygenase (IDO), can be applied to prevent hair loss in an experimental AA model. A single intraperitoneal (IP) injection of IDO-expressing primary dermal fibroblasts was given to C3H/HeJ mice at the time of AA induction. While 60–70% of mice that received either control fibroblasts or vehicle injections developed extensive AA, none of the IDO-expressing fibroblast-treated mice showed new hair loss up to 20 weeks post injection. IDO cell therapy significantly reduced infiltration of CD4+ and CD8+ T cells into hair follicles and resulted in decreased expression of TNF-α, IFN-γ and IL-17 in the skin. Skin draining lymph nodes of IDO fibroblast-treated mice were significantly smaller, with more CD4+ CD25+ FoxP3+ regulatory T cells and fewer Th17 cells than those of control fibroblast and vehicle-injected mice. These findings indicate that IP injected IDO-expressing dermal fibroblasts can control inflammation and thereby prevent AA hair loss.Canadian Institutes of Health Researches (Funding Reference Number: 134214 and 136945)

Novel, Blended Polymeric Microspheres for the Controlled Release of Methotrexate: Characterization and In Vivo Antifibrotic Studies

Low dose methotrexate (MTX) is known to effectively decrease type I collagen production in dermal fibroblasts, while increasing the matrix metalloproteinase-1 (MMP-1) production in vitro. For in vivo use as an antifibrotic agent on wounds, a linear and extended controlled release formulation of MTX is required. The objective of this study was to optimize the fabrication of MTX-loaded polymeric microspheres with such properties, and to test the efficacy for the prevention of fibrosis in vivo. Poly lactic-co-glycolic acid (PLGA), Poly (L-lactic acid) (PLLA) and the diblock copolymer, methoxypolyethylene glycol-block-poly (D, L-lactide) (MePEG-b-PDLLA), were used to fabricate microspheres, which were then characterized in terms of size, drug encapsulation efficiency, and in vitro release profiles. The optimized formulation (PLGA with diblock copolymer) showed high drug encapsulation efficiency (>80%), low burst release (~10%) and a gradual release of MTX. The amphipathic diblock copolymer is known to render the microsphere surface more biocompatible. In vivo, these microspheres were effective in reducing fibrotic tissue which was confirmed by quantitative measurement of type I collagen and α-smooth muscle actin expression, demonstrating that MTX can be efficiently encapsulated in PLGA microspheres to provide a delayed, gradual release in wound beds to reduce fibrosis in vivo

Efficacy of kynurenic acid and methotrexate in prevention of fibrosis

In postnatal life, tissue injuries with sufficient intensity, such as surgical procedures, elicit a repair process rather than regeneration. Hypertrophic scaring, capsular contracture around implants and peritendinous adhesions are common fibroproliferative reactions observed following surgery. These complications not only adversely affect the patients’ quality of life but also impose a significant economic burden on health care system. Despite recent advances in knowledge, refinements of techniques, and various therapeutic methods, the management of post- surgical fibrosis remains a challenge. Recently, Kynurenic acid (KynA), an end product of tryptophan metabolism, has been found to have an anti-fibrotic effect both in vitro and in vivo through suppression of type 1 collagen and enhancement of the matrix metalloproteinase 1 (MMP-1) expression in dermal fibroblasts. Another potential anti-fibrotic molecule is methotrexate (MTX). Systemic low dose MTX, which is a popular first line drug in the management of early and established rheumatoid arthritis (RA), has also been reported to be effective in prevention and treatment of keloid in few case reports. Since fibroproliferative disorders usually appear after the wound is healed, the intact skin can be an obstacle to effective delivery of topical KynA or MTX, and serious side effects limit the systemic use of MTX. In this dissertation, it is hypothesized that localized controlled release of anti-fibrosis agents from biodegradable, biocompatible polymer microspheres, embedded within the wound bed before skin closure, will reduce fibrotic scar formation following surgical procedures. To test this hypothesis, three specific objectives were employed: (1) Developing and validating a suitable animal model for in vivo studies, (2) Fabricating controlled release microspheres loaded with KynA and evaluating the efficacy of fabricated microspheres in vivo and (3) Determining the anti-fibrotic effect of MTX in vitro and evaluating the efficacy of fabricated MTX microspheres in vivo. The findings of these specific objectives demonstrated that the implantation of the controlled release polymer microspheres containing MTX and/or KynA efficiently reduces the fibrotic tissue formation in the PVA sponge-implant model of fibrosis, supporting our hypothesis that controlled local delivery of small anti-fibrosis agents to the wound bed is a feasible method for prevention of post surgical fibrosis.Medicine, Faculty ofExperimental Medicine, Division ofMedicine, Department ofGraduat

Localized Controlled Release of Kynurenic Acid Encapsulated in Synthetic Polymer Reduces Implant—Induced Dermal Fibrosis

Excessive fibrosis following surgical procedures is a challenging condition with serious consequences and no effective preventive or therapeutic option. Our group has previously shown the anti-fibrotic effect of kynurenic acid (KynA) in vitro and as topical cream formulations or nanofiber dressings in open wounds. Here, we hypothesized that the implantation of a controlled release drug delivery system loaded with KynA in a wound bed can prevent fibrosis in a closed wound. Poly (lactic-co-glycolic acid) (PLGA), and a diblock copolymer, methoxy polyethylene glycol-block-poly (D, L-lactide) (MePEG-b-PDLLA), were used for the fabrication of microspheres which were evaluated for their characteristics, encapsulation efficiency, in vitro release profile, and in vivo efficacy for reduction of fibrosis. The optimized formulation exhibited high encapsulation efficiency (>80%), low initial burst release (~10%), and a delayed, gradual release of KynA. In vivo evaluation of the fabricated microspheres in the PVA model of wound healing revealed that KynA microspheres effectively reduced collagen deposition inside and around PVA sponges and α-smooth muscle actin expression after 66 days. Our results showed that KynA can be efficiently encapsulated in PLGA microspheres and its controlled release in vivo reduces fibrotic tissue formation, suggesting a novel therapeutic option for the prevention or treatment of post-surgical fibrosis.Pharmaceutical Sciences, Faculty ofNon UBCReviewedFacultyResearche