gms | German Medical Science

Objectives: Semaphorin 3A (Sema3A) is a neurotransmitter, produced in various tissues, originally identified as an axon guidance molecule. Sema3A is involved in several physiological and pathophysiological processes, like angiogenesis and immune responses, via high affinity binding to a receptor complex consisting of neuropilin-1 (NRP1) and plexin A1. It has been shown that chondrocytes from osteoarthritic (OA) patients have an increased NRP1 gene and Sema3A protein level compared to non-OA chondrocytes. Thus, this work aims to provide new insights into the role of Sema3A in OA pathology and as a potential new treatment option, by inhibiting its receptor subunit NRP1 in OA-chondrocytes.

Methods: Human chondrocytes from OA donors were transfected with NRP1-specific siRNA and proliferation was analysed by BrdU assay, apoptosis via Caspase 3/7 activity assay, adhesion capacity by crystal violet staining, senescence via SA-ß-galactosidase assay and gene and protein expression with qPCR and western blotting. Additionally, NRP1-knockdown- and control cells were stimulated with Sema3A in different concentrations (1-100 ng) and MMP13 gene expression and downstream signalling pathways were analysed.

Results and conclusion: After transient knockdown of NRP1, OA-chondrocytes revealed a decrease in proliferation and an increase in adhesion ability and senescence, but no alteration in apoptosis, even though gene expression of Bcl-2, an anti-apoptotic factor, was increased in NRP1-knockdown cells. Stimulation with Sema3A induced the phosphorylation of ERK1/2 in NRP-1 knockdown and control cells, without affecting the AKT signalling pathway.

The gene expression of VEGFA, and the gene and protein expression of MMP13, two classical hypertrophy marker including the initial stage of OA, was significantly reduced after NRP1 knockdown. When NRP1-knockout- and control cells were stimulated with Sema3A, only control cells increased MMP13 gene expression significantly after Sema3A treatment, whereas NRP-1 knockdown cells did not. This observation points to a direct effect of Sema3A on MMP13 expression via binding to the NRP1 receptor. We conclude that Sema3A-NRP1-binding promotes catabolic processes in OA chondrocytes by stimulating the expression of MMP13 and VEGFA and thereby favour processes such as degradation of the extracellular matrix, probably mediated via the ERK1/2 pathway.

In summary, it is conceivable that blockage of NRP1-signalling during OA pathogenesis could diminish some catabolic mechanisms that promote the degradation of the cartilage matrix and prevent/delay the progression of the disease. However, it is also possible that NRP1 inhibition provokes a reduction in cell proliferation and an increase in senescence, thereby counteracting the attempt of articular chondrocytes to repair damaged matrix regions, a process known as chondrocyte cloning. Therefore, in-depth studies are needed to clarify the potential of NRP1 inhibition to induce rather beneficial or detrimental processes in chondrocytes.