ASCs and BMSCs display similar morphology and growth characteristics when expanded ex vivo. Immunophenotypically, they are both known to be positive for the classical mesenchymal stromal cell markers CD105, CD44 CD90, CD166, and negative for CD14, CD19, and CD45 . There is a continued effort to understand the differences between these MSCs, particularly as they are both being advanced for cell immunotherapeutic purposes. ASCs and BMSCs have comparable suppressive effects on the growth of PHA-stimulated T cells, suggesting that they have equal influence on the adaptive immune response . We explored their comparability in preventing innate immune responses observed in endotoxic inflammation and demonstrate a significant improvement of disease indices by BMSC secreted factors. We discovered that BMSCs secrete significantly greater amounts of two soluble cytokine receptors, sTNFR1 and sVEGFR1, compared to ASCs. Although these soluble cytokine receptors can be considered potency markers unique to BMSCs, other factors are likely to also be responsible for BMSCs’ therapeutic effects in systemic inflammatory responses.
sTNFR1 has been shown to block the effects of TNF-α, an inflammatory cytokine that is released in response to inflammation. With increased distress, TNF-α reaches dangerously high levels and can eventually lead to death . TNF-α converting enzyme (TACE) has been implicated as the enzyme that sheds sTNFR1 from TNFR1. Upon sTNFR1’s release, it binds to circulating TNF-α and prevents its inflammatory effects . A study by Yagi et al. demonstrated the value of circulating sTNFR1 released by intramuscular BMSC transplants in the attenuation of septic shock in rodents. When BMSC transplants were co-administered with a neutralizing sTNFR1 antibody, the cell therapy failed to prevent the infiltration of inflammatory cells in the lungs, liver, and kidney . The downstream mechanism by which sTNFR1 is protective remains to be determined. There is evidence that sTNFR1 may not just simply block TNF-α, but exert its effects by inducing apoptosis in monocytes via transmembrane TNF-α . Waetzig et al. found that sTNFR1 boosts TGFβ1, which in turn has the ability to inhibit T lymphocyte proliferation . There is, however, a delicate balance of the sTNFR1/TNF-α axis where it has been observed that low sTNFR1 levels stabilize TNF-α . Further studies will be needed to unravel the mechanism that BMSCs employ to make sTNFR1 and how this soluble receptor modulates the immune system.
VEGF, an angiogenic factor that supports microcapillary growth from existing vasculature, typically benefits endothelial tissues . Recent studies have shown that elevated levels of VEGF in response to inflammation can cause more hindrance than initially thought. A study by Tsao et al. demonstrated that injecting mice with VEGF after induction of endotoxemia via LPS resulted in 100% mortality . VEGF induces the expression of cell adhesion molecules, which cause leukocytes to bind effectively to the endothelium. This can lead to even greater local cytokine production. VEGF can be silenced by sVEGFR1, the truncated form of membrane-bound VEGF receptor 1 . Heightened sVEGFR1 levels can mitigate this devastating cascade at the blood vessel scale. Mice treated with sVEGFR1 as late as 20 hours after LPS administration have demonstrated a survival rate of 100% . Several theories have been proposed to explain the dichotomous nature of an anti-angiogenic factor, sVEGFR1, as a beneficial influence in inflammatory disease. Some suggest that high levels of sVEGFR1 in sepsis may promote hypocoagulability by recruiting endogenous anti-coagulating molecules, which counteract the tissue damage by allowing microcirculation to remain open and permit the body to recover more quickly from endotoxemia . sVEGFR1 has also been shown to selectively activate endothelial nitric oxide synthase (eNOS), which contributes to arteriogenesis, angiogenesis, and mural cell recruitment . Apart from sepsis, the presence of sVEGFR1 is also thought to regulate angiogenesis by VEGF in the setting of cancerous tumor growth . The release of sVEGR1 by BMSCs may have implications in the therapeutic use of this cell population as well as the endogenous regulation of VEGF signaling in stromalized microenvironments.
Our findings indicate, however, that modulating sVEGFR1 activity alone with a neutralizing antibody does not significantly affect BMSC-CM’s benefit to sepsis survival. For the studies utilizing neutralizing antibodies, the addition of IgG to the BMSC-CM would ideally be used to control for the presence of antibody in the neutralizing antibody-treated CM. Its absence, however, is likely to be inconsequential because the bovine serum albumin IgG in the CM acts as a sufficient control for the neutralizing antibody’s influence on the LPS-challenged mice.
In conclusion, we have uncovered a potency advantage of BMSCs compared to ASCs in a model of sepsis and a unique expression pattern of soluble receptors that are implicated in the resolution of inflammation. This study can guide the positioning and monitoring of these cell populations for therapeutic use in immune-mediated disease and may have ramifications to endogenous stromal cell biology.