Prolonged Exposure to Hypoxic Milieu Improves the Osteogenic Potential of Adipose Derived Stem Cells

Abstract

Mesenchymal stem cells (MSC) have been widely used in orthopedics for several applications. Conventionally, MSC are maintained under 21% O2 which does not reflect the real O2 tension in vivo. Recently, it was reported that different O2 conditions can give different cellular responses. Here, we investigated whether prolonged exposure to hypoxia affects the osteogenic differentiation of adipose-derived stem cells (ASC). ASC from six individuals were cultured under "low" (2-3%) or "air" (21%) oxygen tensions, either without or with osteogenic stimuli. The effect of the O2 tension was evaluated on cell proliferation, surface antigens, stemness and bone-related genes expression, alkaline phosphatase activity (ALP), mineralization activity, and release of osteogenic growth factors. Without differentiating stimuli, hypoxia favored ASC proliferation, reduced the number of CD184+ and CD34+ cells, and preserved the expression of NANOG and SOX2. The combination of hypoxia and osteogenic medium induced a high proliferation rate, a rapid and more pronounced mineralization activity, a higher expression of genes related to the MSC differentiation, a higher release of mitogenic growth factors (bFGF, PDGF-BB), and the decrease in TGF-β secretion, an inhibitor of the early stage of the osteoblast differentiation. We demonstrated that hypoxia acts dually, favoring ASC proliferation and the maintenance of the stemness in the absence of osteogenic stimuli, but inducing the differentiation in a bone-like microenvironment. In conclusion, prolonged cell culture in hypoxic microenvironment represents a proper method to modulate the stem cell function that may be used in several applications, for example, studies on bone pathophysiology or bone-tissue engineering.

The involvement of histone methylation in osteoblastic differentiation of human periosteum‐derived cells cultured in vitro under hypoxic conditions

“…Hsu et al reported that hypoxic conditions, compared with normoxic conditions, suppress the osteoblastic differentiation of human MSCs isolated from bone marrow by inhibiting metabolic switch and mitochondrial function. In contrast, in a study using human MSCs isolated from adipose tissue, Fotia et al suggested that hypoxic conditions provide a culture environment that enhances the osteoblastic potential of adipose‐derived MSCs. In the present study, the osteoblastic phenotypes of hPDCs identified through ALP activity, alizarin red‐S positive mineralization, and mRNA expression levels of osteoblast‐related genes were significantly decreased in cells cultured in hypoxia compared with normoxia.…”

“…Although oxygen tension plays an important role in the viability and maintenance of MSCs in an undifferentiated state, there is controversy regarding the effect of hypoxia on the osteoblastic differentiation of MSCs . Several studies have shown a reduced differentiation potential of MSCs cultured in hypoxia, while others reported that hypoxic conditions enhance the osteoblastic differentiation of MSCs .…”

“…The effect of hypoxia on osteoblastic differentiation of MSCs remains controversial. While several studies have shown that MSCs in hypoxia are more prone to osteogenic differentiation relative to those in normoxia, others have found the opposite correlation …”

“…Likewise, oxygen tension influences the biological characteristics of stem cells, such as self-renewal and the generation of While several studies have shown that MSCs in hypoxia are more prone to osteogenic differentiation relative to those in normoxia, others have found the opposite correlation. [4][5][6][7][8][9] Differentiation of stem cells into specialized cells requires the activation of genes involved in the expression of a specific cell phenotype. Regarding osteogenic differentiation of MSCs, epigenetic regulation may play a role in the increased expression of osteoblastspecific genes.…”

Isolation and Culture of Human Stem Cells from Apical Papilla under Low Oxygen Concentration Highlight Original Properties

“…In addition, BM-MSCs cultured under low oxygen tension before their implantation, exhibit improved repair capabilities for bone defects, tendon tear, stroke and hindlimb ischemia [21,22]. Adipose tissue derived stem cells (ADSCs) proliferate well and maintain stemness with improved differentiation potential when grown under low O 2 concentration [23,24].…”