Cell-based therapies offer a promising strategy to treat tendon injuries and diseases. Both mesenchymal stromal cells (MSCs) and pluripotent stem cells (PSCs) are good candidates for such applications due to their self-renewing and differentiation capacity. However, the translation of cell-based therapies from bench to bedside can be hindered by the use of animal-derived components in ancillary materials and by the lack of standardised media and protocols for in vitro tenogenic differentiation. To address this, we have optimized animal component-free (ACF) workflows for differentiating human MSCs and PSCs to tenocyte-like cells (TLCs) respectively. MSCs isolated from bone marrow (n = 3) or adipose tissue (n = 3) were expanded using MesenCult™-ACF Plus Culture Kit for at least 2 passages, and differentiated to TLCs in 21 days using a step-wise approach. Briefly, confluent cultures were treated with an ACF tenogenic induction medium for 3 days, followed by treatment with an ACF maturation medium for 18 days. Monolayer cultures were maintained at high density without passaging for the entire duration of the protocol, and the medium was changed every 2 – 3 days. In a similar fashion, embryonic (n = 3) or induced PSCs (n = 3) were first differentiated to acquire a mesenchymal progenitor cell (MPC) phenotype in 21 days using STEMdiff™ Mesenchymal Progenitor Kit, followed by the aforementioned tenogenic protocol for an additional 21 days. In all cases, the optimized workflows using ACF formulations consistently activated a tenogenic transcriptional program, leading to the generation of elongated, spindle-shaped tenomodulin-positive (TNMD+) cells and deposition of an extracellular matrix predominantly composed of collagen type I. In summary, here we describe novel workflows that can robustly generate TLCs from MSCs and hPSC-derived MPCs for potential translational applications.

Mesenchymal stromal cells (MSCs) have been one of the most widely studied cell types in preclinical and clinical trials, due to their self-renewing, multipotent capacity, immunomodulatory properties and relative ease of isolation from multiple tissues. Despite limitations and safety concerns, fetal bovine serum (FBS) is still predominantly used for MSC expansion in clinical protocols. In addition, the undefined nature of serum composition and lot-to-lot variability have been linked to reduced reproducibility and efficiency of MSC bioprocessing. Moreover, use of animal serum in human cell culture increases the risk of contamination with adventitious pathogenic microorganisms, such as viruses, prions and bacteria. Hence, a defined serum-free formulation can provide increased safety, better control over physiological responsiveness, consistent performance and reproducible results. Here we present preliminary data on a prototype serum-free medium optimized for in vitro tenogenic differentiation of human bone marrow-derived MSCs. This serum-free formulation is capable of generating tenocyte-like cells in vitro expressing tenogenic markers such as Scx, Tnmd, TnC, Collagen I and Collagen III, whilst repressing expression of specific markers of other mesenchymal lineages.

We aimed to determine the impact of anaesthetic techniques on post-operative analgesic utilisation in people undergoing total knee arthroplasty. We conducted a prospective study at a district general hospital to assess post-operative analgesia use via patient controlled analgesia (PCA) in patients undergoing a elective total knee arthroplasty. Subjects were divided into three groups; group 1 (general anaesthetic and local nerve block, n = 39), group 2 (spinal only, n= 39) and group 3 (general anaesthetic only, n = 38). The primary outcome measure was post-operative morphine consumption through a PCA within the first 24 hours. All subjects were followed up post-operatively by a dedicated pain control nurse. Data was analysed using SPSS version 17 for Windows (SPSS Inc, Chicago, IL, USA).

Results are reported as mean (SD) unless stated otherwise. No significant difference in the age or sex distribution between groups was demonstrable. There was no statistical difference in the average usage of morphine when comparing Group 1 compared to group 2, (77.57 (49.56) vs 65.80 (44.71), p=0.27), group 1 compared to group 3, (77.57 (49.56) vs 77.80 (45.52) p=0.98) and finally group 2 compared to group 3, (65.80 (44.71) vs 77.80 (45.52), p=0.25).

In this cohort of patients undergoing total knee arthroplasty the anaesthetic technique used does not appear to have an influence on the post-operative pain management.