Human Serum is a Suitable Supplement for the Osteogenic Differentiation of Human Adipose-Derived Stem Cells Seeded on Poly-3-Hydroxibutyrate-Co-3-Hydroxyvalerate Scaffolds

Abstract

Human adipose-derived stem cells (hASCs) are currently a point of focus for bone tissue engineering applications. However, the ex vivo expansion of stem cells before clinical application remains a challenge. Fetal bovine serum (FBS) is largely used as a medium supplement and exposes the recipient to infections and immunological reactions. In this study, we evaluated the osteogenic differentiation process of hASCs in poly-3-hydroxybutyrate-co-3-hydroxyvalerate (PHB-HV) scaffolds with the osteogenic medium supplemented with pooled allogeneic human serum (aHS). The hASCs grown in the presence of FBS or aHS did not show remarkable differences in morphology or immunophenotype. The PHB-HV scaffolds, which were developed by the freeze-drying technique, showed an adequate porous structure and mechanical performance as observed by micro-computed tomography, scanning electron microscopy (SEM), and compression test. The three-dimensional structure was suitable for allowing cell colonization, which was revealed by SEM micrographs. Moreover, these scaffolds were not toxic to cells as shown by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The differentiation capacity of hASCs seeded on scaffolds was confirmed by the reduction of the proliferation, the alkaline phosphatase (AP) activity, expression of osteogenic gene markers (AP, collagen type I, Runx2, and osteocalcin), and the expression of bone markers, such as osteopontin, osteocalcin, and collagen type I. The osteogenic capacity of hASCs seeded on PHB-HV scaffolds indicates that this scaffold is adequate for cell growth and differentiation and that aHS is a promising supplement for the in vitro expansion of hASCs. In conclusion, this strategy seems to be useful and safe for application in bone tissue engineering.

Production of Human Endothelial Cells Free from Soluble Xenogeneic Antigens for Bioartificial Small Diameter Vascular Graft Endothelization

“…The aHS was obtained from the whole blood of healthy donors from all blood group types, as previously described by our group [8]. Briefly, whole blood was collected and allowed to clot spontaneously.…”

“…This study was approved by the Ethics Committee of the Federal University of Minas Gerais (number ETIC 21176413.9.0000.5149). The isolation and culture of hASCs were performed as described, with minor modifications [8, 11]. Briefly, the excess of blood in the raw lipoaspirates was removed with phosphate-buffered saline (PBS) washes and enzymatic digestion of adipose tissue was performed with the incubation of the tissue in a solution of 0.075% type I collagenase I (Life Technologies) in PBS at 37°C for 1 h. Then, the stromal vascular fraction was isolated by centrifugation at 252 g for 10 min, and the pellet was suspended in BM and plated into cell culture flasks (Sarstedt).…”

“…Phenotypic characterization of isolated hASCs was performed as recommended by the Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy [12] and performed as already described elsewhere [8]. Briefly, fourth passage undifferentiated hASCs were detached from cell culture surface and counted.…”

“…Recently, our group has shown that human allogeneic serum (aHS) is a suitable cell culture supplement for hASCs [8]. In addition to successfully maintaining hASCs in vitro , it maintains hASCs multipotency and induces higher proliferation rates without promoting cell senescence or loss of differentiation capacity [8–10].…”

“…In addition to successfully maintaining hASCs in vitro , it maintains hASCs multipotency and induces higher proliferation rates without promoting cell senescence or loss of differentiation capacity [8–10]. Therefore, the use of aHS may be beneficial for tissue engineering approaches.…”