CTRP3 disruption promotes a tendon-like matrix signature in tenogenically induced canine adipose-derived stem cells

Abstract

Introduction Tendon injuries heal through disorganized extracellular matrix (ECM) deposition rather than the restoration of native collagen architecture, thus resulting in suboptimal functional recovery. Although adipose-derived stem cells (ADSCs) are attractive for tendon regeneration because of their accessibility and expandability, their tenogenic differentiation efficiency is lower than that of tendon-derived stem cells (TDSCs). C1q/TNF-related protein 3 (CTRP3) is an adipokine implicated in ECM remodeling and fibrosis; however, its role in tendon lineage regulation is unknown. Objective CTRP3 (C1q/TNF-related protein 3) contributes to extracellular-matrix remodeling and, in tendon, is associated with a pathological fibrotic matrix signature; we therefore reasoned that reducing CTRP3 would favor a more mature tendon-like matrix. Using a loss-of-function approach based on clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9), we investigated whether CTRP3 disruption alters tenogenic marker expression, at both the transcript and protein levels, and the tendon-associated ECM marker profile of canine ADSCs relative to wild-type ADSCs and native TDSCs. Methods CRISPR/Cas9-mediated CTRP3–disrupted ADSCs were generated and validated using sequencing, quantitative polymerase chain reaction, and Western blotting. Wild-type ADSCs, CTRP3-disrupted ADSCs, and TDSCs were subjected to tenogenic induction for 0–3 weeks. Tenogenic transcriptional and ECM markers were analyzed at mRNA and protein levels. Results CTRP3-disrupted ADSCs showed increased collagen type I and tenomodulin (TNMD) expression during tenogenic induction compared with wild-type ADSCs. COL III expression showed a transient increase followed by a decline, with lower expression in CTRP3-disrupted cADSCs than in WT cADSCs at week 2. CTRP3 disruption increased early Mkx and Scx expression at the earliest time points but did not consistently alter the early transcriptional program, which rose comparably across all groups. CTRP3-disrupted cADSCs showed partial, marker- and time-point-dependent overlap with native cTDSCs, mainly for selected late ECM markers rather than across the overall transcript and protein profiles. Conclusion CTRP3 disruption shifted the tendon-associated ECM marker profile during the tenogenic differentiation of ADSCs, with reduced COL III accumulation, rather than uniformly enhancing tenogenic differentiation. Although early transcriptional changes were observed, the more consistent differences were found in late tendon-associated ECM markers. The modulation of CTRP3 may represent a candidate strategy worth further investigation for stem cell–based tendon regeneration in veterinary medicine.

The authors' abstract, as published at the source. Regenerative Therapy, 2026 · DOI ↗