Tumour necrosis factor alpha, interleukin 1 beta and interferon gamma have detrimental effects on equine tenocytes that cannot be rescued by IL-1RA or mesenchymal stromal cell–derived factors

Abstract

Tendon injuries occur commonly in both human and equine athletes, and poor tendon regeneration leads to functionally deficient scar tissue and an increased frequency of re-injury. Despite evidence suggesting inadequate resolution of inflammation leads to fibrotic healing, our understanding of the inflammatory pathways implicated in tendinopathy remains poorly understood, meaning successful targeted treatments are lacking. Here, we demonstrate IL-1β, TNFα and IFN-γ work synergistically to induce greater detrimental consequences for equine tenocytes than when used individually. This includes altering tendon associated and matrix metalloproteinase gene expression and impairing the cells’ ability to contract a 3-D collagen gel, a culture technique which more closely resembles the in vivo environment. Moreover, these adverse effects cannot be rescued by direct suppression of IL-1β using IL-1RA or factors produced by BM-MSCs. Furthermore, we provide evidence that NF-κB, but not JNK, P38 MAPK or STAT 1, is translocated to the nucleus and able to bind to DNA in tenocytes following TNFα and IL-1β stimulation, suggesting this signalling cascade may be responsible for the adverse downstream consequences of these inflammatory cytokines. We suggest a superior approach for treatment of tendinopathy may therefore be to target specific signalling pathways such as NF-κB.

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“…1f). For all experiments either vehicle only conditions were used as a negative control, or adult tenocytes were used as a positive control since they are known to be adversely affected by inflammatory cytokine stimulation [13].…”

“…Inflammation in tendon injuries is characterised by the endogenous expression of inflammatory mediators including pro-inflammatory cytokines. Amongst these, TNFα (tumour necrosis factor alpha) and IL-1β (interleukin 1 beta) are evidenced to be the most potent inducers of a catabolic response, with recent evidence suggesting these inflammatory cytokines in combination with IFN-γ (interferon gamma) have detrimental consequences for tendon cell function in vitro [13]. Inflammatory cytokines stimulate the activation of specific signalling pathways inside the cell, including nuclear factor kappa B (NF-κB), mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK), and signal transducer and activator of transcription (STATs) [14][15][16].…”

“…Inflammatory cytokines stimulate the activation of specific signalling pathways inside the cell, including nuclear factor kappa B (NF-κB), mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK), and signal transducer and activator of transcription (STATs) [14][15][16]. Our previous work has demonstrated that in adult tenocytes the adverse effects of TNFα and IL-1β are likely caused by their activation of NF-κB [13,17]. This triggers a series of inflammatory processes that eventually lead to the transcription of different regulatory genes, linking extracellular signals to the mechanisms that govern essential cellular functions.…”

“…This triggers a series of inflammatory processes that eventually lead to the transcription of different regulatory genes, linking extracellular signals to the mechanisms that govern essential cellular functions. However, although an inflammatory response is essential to initiate successful wound healing, dysregulation of these signalling pathways may contribute to the degradation of the tendon matrix and ultimately lead to chronic tendinopathy and tissue fibrosis [13,[18][19][20]. Therefore, understanding how these mechanisms regulate inflammation is critical to identifying therapeutic approaches to promote tendon healing.…”

“…The response of adult tenocytes to pro-inflammatory cytokines has been investigated extensively [13,17,[34][35][36], whereas few studies have been performed in equine ESCderived tenocytes [34]. Here, we determine the responses of equine ESC-tenocytes to IFN-γ, TNFα, and IL-1β.…”