TissueGene-C induces long-term analgesic effects through regulation of pain mediators and neuronal sensitization in a rat monoiodoacetate-induced model of osteoarthritis pain.

OBJECTIVE: TissueGene-C (TG-C), a combination of human allogeneic chondrocytes and irradiated GP2-293 cells engineered to overexpress transforming growth factor-ß1 (TGF-ß1), has been developed as a novel cell-based gene therapy and a candidate for disease modifying osteoarthritis drug (DMOAD). We aim to investigate analgesic mechanism of TG-C in a pre-clinical animal model with monoiodoacetate (MIA)-induced pain. DESIGN: We used a rat MIA model of osteoarthritis (OA) pain. We examined that TG-C can regulate pain by inhibiting the upregulation of various pain mediators in both knee joint tissue and dorsal root ganglia (DRG) (n = 112) and alleviating pain behavior (n = 41) and neuronal hyperexcitability in DRG (n = 60), afferent nerve fiber (n = 24), and spinal cord (n = 35). RESULTS: TG-C significantly alleviated pain-related behavior by restoring altered dynamic weight bearing and reduced mechanical threshold of the affected hindlimb. TG-C significantly suppressed the expression of nerve growth factor (NGF) and calcitonin gene-related peptide (CGRP) in inflamed joint tissue. TG-C significantly suppressed the upregulation of tropomyosin receptor kinase A (TrkA) and nerve injury/regeneration protein (GAP43) and activation of Iba1-positive microglial cells in DRG. TG-C significantly recovered neuronal hyperexcitability by restoring RMP and firing threshold and frequency of DRG neurons, attenuating firing rates of mechanosensitive C- or Aδ-nerve fiber innervating knee joint, and lowering increased miniature and evoked excitatory postsynaptic currents (mEPSCs and eEPSCs) in the spinal cord. CONCLUSION: Our results demonstrated that TG-C exerted potent analgesic effects in a rat MIA model of OA pain by inhibiting the upregulation of pain mediators and modulating neuronal sensitization.

The current state of the osteoarthritis drug development pipeline: a comprehensive narrative review of the present challenges and future opportunities.

In this narrative review article, we critically assess the current state of the osteoarthritis (OA) drug development pipeline. We discuss the current state-of-the-art in relation to the development and evaluation of candidate disease-modifying OA drugs (DMOADs) and the limitations associated with the tools and methodologies that are used to assess outcomes in OA clinical trials. We focus on the definition of DMOADs, highlight the need for an updated definition in the form of a consensus statement from all the major stakeholders, including academia, industry, regulatory agencies, and patient organizations, and provide a summary of the results of recent clinical trials of novel DMOAD candidates. We propose that DMOADs should be more appropriately targeted and investigated according to the emerging clinical phenotypes and molecular endotypes of OA. Based on the findings from recent clinical trials, we propose key topics and directions for the development of future DMOADs.

TissueGene-C promotes an anti-inflammatory micro-environment in a rat monoiodoacetate model of osteoarthritis via polarization of M2 macrophages leading to pain relief and structural improvement.

Osteoarthritis (OA) is the most common form of arthritis, characterized by cartilage destruction, pain and inflammation in the joints. Existing medications can provide relief from the symptoms, but their effects on the progression of the disease are limited. TissueGene-C (TG-C) is a novel cell and gene therapy for the treatment of OA, comprising a mixture of human allogeneic chondrocytes and irradiated cells engineered to overexpress transforming growth factor-ß1 (TGF-ß1). This study aims to investigate the efficacy and mechanism of action of TG-C in a rat model of OA. Using the monosodium-iodoacetate (MIA) model of OA, we examined whether TG-C could improve OA symptoms and cartilage structure in rats. Our results showed that TG-C provided pain relief and cartilage structural improvement in the MIA OA model over 56 days. In parallel with these long-term effects, cytokine profiles obtained on day 4 revealed increased expression of interleukin-10 (IL-10), an anti-inflammatory cytokine, in the synovial lavage fluid. Moreover, the increased levels of TGF-ß1 and IL-10 caused by TG-C induced the expression of arginase 1, a marker of M2 macrophages, and decreased the expression of CD86, a marker of M1 macrophages. These results suggest that TG-C exerts a beneficial effect on OA by inducing a M2 macrophage-dominant micro-environment. Cell therapy using TG-C may be a promising strategy for targeting the underlying pathogenic mechanisms of OA, reducing pain, improving function, and creating a pro-anabolic micro-environment. This environment supports cartilage structure regeneration and is worthy of further evaluation in future clinical trials.

TRPM2 contributes to LPC-induced intracellular Ca2+ influx and microglial activation.

Microglia are the resident immune cells which become activated in some pathological conditions in central nervous system (CNS). Lysophosphatidylcholine (LPC), an endogenous inflammatory phospholipid, is implicated in immunomodulatory function of glial cells in the CNS. Although several studies uncovered that LPC induces intracellular Ca2+ influx and morphologic change in microglia, there is still no direct evidence showing change of phosphorylation of mitogen-activated protein kinase (MAPK) p38 (p-p38), a widely used microglia activation marker, by LPC. Furthermore, the cellular mechanism of LPC-induced microglia activation remains unknown. In this study, we found that LPC induced intracellular Ca2+ increase in primary cultured microglia, which was blocked in the presence of Gd3+, non-selective transient receptor potential (TRP) channel blocker. RT-PCR and whole cell patch clamp recordings revealed molecular and functional expression of TRP melastatin 2 (TRPM2) in microglia. Using western blotting, we also observed that LPC increased phosphorylation of p38 MAPK, and the increase of p-p38 expression is also reversed in TRPM2-knockout (KO) microglia. Moreover, LPC induced membrane trafficking of TRPM2 and intrathecal injection of LPC increased Iba-1 immunoreactivity in the spinal cord, which were significantly reduced in KO mice. In addition, LPC-induced intracellular Ca2+ increase and inward currents were abolished in TRPM2-KO microglia. Taken together, our results suggest that LPC induces intracellular Ca2+ influx and increases phosphorylation of p38 MAPK via TRPM2, which in turn activates microglia.

High-resolution transcriptome analysis reveals neuropathic pain gene-expression signatures in spinal microglia after nerve injury.

Microglial cells, the resident immune cells of the spinal cord, become activated in response to peripheral nerve injury. Microglia activation contributes to the development of neuropathic pain. Here we employed microarray analysis of individually collected pools of 10 spinal microglia cells to identify changes of levels and cell-to-cell expression variance of microglial genes during their activation after peripheral nerve injury. The analysis of microglia on postoperative day 1 (POD1) identified miR-29c as a critical factor for microglial activation and the development of neuropathic pain. Early POD1 microglia exhibited a very distinct expression profile compared to late POD7 microglia, possibly leading to the transition from initiation to maintenance of neuropathic pain. We found sample variance patterns that were consistent with the hypothesis that microglia were highly heterogeneous at the level of individual cells, and variation analysis identified 56 microglial genes potentially linked to the maintenance of neuropathic pain which included Gria1. This study provides insights into spinal microglial biology and reveals novel microglial targets for the treatment of neuropathic pain.