# Adipose Stem Cell Response to Borophosphate Bioactive Glass
## 4 Discussion
This agrees with other studies that have shown the impact of borate released from BBGs on angiogenesis [14; 49; 50; 51; 52]. The phosphate-rich, acidic X0 glass completely inhibited osteogenesis in all three ASC donors. The adipogenic ability was also reduced compared to the controls, though not significantly. This correlates with the observation that pre-treating MSCs with an acidic environment enhances the stemness of MSCs, thus resulting in a lack of differentiation ability [53]. The borate-rich, alkaline X60 glass, on the other hand, reduced adipogenic differentiation. This coincides with previous work showing that boron prevented lipid deposition and suppressed adipogenesis transcriptional programming while maintaining cell viability [54]. Interestingly, despite all donors being similar in age, BMI, race, and sex, there was a donor difference in all differentiation assays except for X0 treatment in osteogenesis. The differences in donors' phenotypic changes from glass treatment warrant further investigation. Most of the literature on bioactive glass, including BPBG, focuses on cell viability, proliferation, or differentiation of ASCs [45; 55; 56; 57]. However, there is very little reported on the effect of BG on the ASC secretome. ASCs have a broad secretory profile of different growth factors, cytokines, and other proteins that impact the body's response to injuries, healing, and tissue regeneration. These secreted proteins all work to keep the body in homeostasis [33; 58]. The present study showed that BPBGs can alter the ASC's secretome. Our study showed that, regardless of the glass composition, the expression of 6-Ckine, EpCAM, FasL, TRAIL R4, and IL-4 was promoted, while they were not secreted without any of the glass treatments. Each of these biomolecules is worth investigating, as BGs may be used as a cell treatment to increase the efficacy of the cells. For example, IL-4 overexpression in BMSCs polarized macrophages from an inflammatory M1 macrophage into a tissue regenerative M2 phenotype [59]. Evaluating these molecules will allow biomaterial designers to cater their materials for different applications. ASCs treated with the pH-neutral X40 glass increased the secretion of anti-proliferation and anti-migration proteins. Because X40 and X0 had similar effects on the pH of the extracellular fluid, the pH cannot be solely responsible for the increase in these proteins. The greater borate or phosphate release from X40 may have impacted the function of ASCs. The concentration of borate ions affected the ability of ASCs to secrete 22 proteins: 2 decreased with increased borate while 20 increased, which included many pro-inflammatory (IL-1, IL-6, IL17, etc.) and angiogenic (VEGF, L-selectin, TIMP-2, etc) proteins. As borate is anti-inflammatory in other applications, it is worth looking into how the controlled release of borate ions in different pH ranges may have different control over cells [60]. These results correlate to previous work from other groups showing that the release of therapeutic ions from bioactive glass influences cell function and phenotype [6; 12; 61]. One limitation of this study is that the pH was measured from the extracellular fluid in its entirety and not at the cell surface. Although the ions released from the glass most likely impacted cell phenotype, it is uncertain if the ions contributed to a localized change in pH near the cell surface. Bioactive glasses have the ability to alter local pH [21; 22; 23; 24]. Cells use pH gradients, local heterogeneity, and dynamic fluctuations to regulate biophysical phenomena, such as metabolism, migration, vesicular traffic, and the spatial organization of tissues during development [62]. The fluctuation of the pH gradient at the cell surface can determine cell fate by activating molecular pathways, for example via the GPCR, a family of proton-sensing receptors on cell membranes [63].