Mesenchymal stromal cells (MSCs) are self-renewing, culture-expandable adult stem cells that have been isolated from a variety of tissues, and possess multipotent differentiation capacity, immunomodulatory properties, and are relatively non-immunogenic. Due to this unique set of characteristics, these cells have attracted great interest in the field of regenerative medicine and have been shown to possess pronounced therapeutic potential in many different pathologies. MSCs' mode of action involves a strong paracrine component resulting from the high levels of bioactive molecules they secrete in response to the local microenvironment. For this reason, MSCs' secretome is currently being explored in several clinical contexts, either using MSC-conditioned media (CM) or purified MSC-derived extracellular vesicles (EVs) to modulate tissue response to a wide array of injuries. Rather than being a constant mixture of molecular factors, MSCs' secretome is known to be dependent on the diverse stimuli present in the microenvironment that MSCs encounter. As such, the composition of the MSCs' secretome can be modulated by preconditioning the MSCs during in vitro culture. This manuscript reviews the existent literature on how preconditioning of MSCs affects the therapeutic potential of their secretome, focusing on MSCs' immunomodulatory and regenerative features, thereby providing new insights for the therapeutic use of MSCs' secretome.

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Low back pain is a highly prevalent clinical problem and intervertebral disc (IVD) degeneration is now accepted as the major pathophysiological mechanism responsible for this condition. Accumulating evidence suggests that inflammation plays a crucial role in the progression of human IVD degeneration, with macrophages being pointed as the key immune cell players in this process since their infiltration in degenerated IVD samples has been extensively demonstrated. Since they are highly plastic, macrophages can play different roles depending on the microenvironmental cues. The study of inflammation associated with IVD degeneration has been somehow neglected and one of the reasons is related with lack of adequate models. To overcome this, we established and characterized a new model of IVD organ culture under pro-inflammatory conditions to further dissect the role of macrophages in IVD associated immune response. For that, human monocyte-derived macrophages were co-cultured either with bovine caudal IVD punches in the presence of the pro-inflammatory cytokine IL-1β, or IVD-conditioned medium (CM), to investigate how IVD-produced factors influence macrophage phenotype. After 72 h, metabolic activity, gene expression and cytokine profile of macrophages and IVD cells were measured. Our results show that macrophages and IVDs remain metabolically active in the presence of IL-1β, significantly upregulate CCR7 gene expression and increase production of IL-6 on macrophages. When treating macrophages with IL-1β-IVD-CM, CCR7 upregulation follows the same trend, while for IL-6 an opposite effect was observed. On the other hand, macrophages interfere with IVD ECM remodeling, decreasing MMP3 expression and downregulating aggrecan and collagen II gene expression in the presence of IL-1β. Overall, the co-culture model established in this study can be considered a suitable approach to address the cellular and molecular pathways that regulate macrophage-IVD crosstalk, suggesting that degenerated IVD tissue tends to polarize human macrophages toward a more pro-inflammatory profile, which seems to aggravate IVD degeneration. This model could be used to improve the knowledge of the mechanisms that link IVD degeneration and the immune response.