Intervertebral disc (IVD) degeneration presents a harsh microenvironment characterised by low glucose, low oxygen and matrix acidity posing a significant challenge for cell-based therapies. The objective of this work was to assess the effect of primed bone marrow derived stem cells (BMSC) and articular chondrocytes (AC) in different pH (7.1, 6.8 and 6.5) conditions and assess metabolic activity in terms of oxygen (O₂) and glucose consumption as well as lactate production. Secondly, we investigated pH effects on cell viability and matrix accumulation capacity. Primary cells were encapsulated in alginate beads and cultured in disc-like conditions (5% O₂, 5mM glucose, pH 7.1, 6.8 and 6.5). For growth factor priming, cells were cultured with 10ng/ml TGF-β3 at a pH of 7.4 for 14 days prior to being subjected to acidic pH conditions. AC exhibited superior cell viability and sGAG deposition compared to BMSC at all pH levels which was further enhanced after priming. Priming also reduced O₂ consumption of AC for all pH conditions while lactate production profiles of both cell types were altered with decreasing extracellular pH. This work demonstrates the importance of cell type selection to sustain disc-like microenvironmental conditions. Results show that BMSCs that have not been primed may need additional factors to sustain the harsh acidic microenvironment. In contrast, AC were capable of sustaining the low pH conditions better than BMSC and accumulated more similar disc-like matrix in all conditions. Overall this study highlights that AC may be advantageous for disc regeneration and warrant further investigation for disc repair.

The scarcity of mesenchymal stem cells (MSCs) in iliac crest bone marrow aspirate (ICBMA), and the expense and time in culturing cells, has led to the search for alternative harvest sites. The reamer-irrigation-aspirator (RIA) provides continuous irrigation and suction during reaming of long bones. The aspirated contents pass via a filter, trapping bony fragments, before moving into a ‘waste’ bag from which MSCs have been previously isolated. We examined the liquid and solid phases, performed a novel digestion of the solid phase, and made a comparative assessment in terms of number, phenotype and differentiation capacity with matched ICBMA.

The solid fraction from the filtrate was digested for 60 minutes at 37°C with collagenase. Enumeration was performed via the colony-forming unit fibroblast (CFU-F) assay. Passage (P2) cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages, and their phenotypes assessed using flow cytometry (CD33, CD34, CD45, CD73, CD90, and CD105).

MSCs from the RIA phases were able to differentiate at least as well as those from ICBMA, and all fractions had phenotypes consistent with other established sources. The median number of colonies for the three groups was: ICBMA = 8.5 (2 to 86), RIA-liquid = 19.5 (4 to 90), RIA-solid = 109 (67 to 200) per 200 μl. The mean total yield of cells for the three groups was: ICBMA = 920 (0 to 4275), RIA-liquid = 114 983 (16 500 to 477 750), RIA-solid = 12 785 (7210 to 28 475).

The RIA filtrate contains large numbers of MSCs that could potentially be extracted without enzymatic digestion and used for bone repair without prior cell expansion.