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Oxysterol as a Deterrent of Adipogenesis: in vitro and in vivo Studies
Reena Bakshi, MD, Akishige Hokugo, DDS, PhD, Luis Segovia, MD, Situo Zhou, MD, Kameron Rezzadeh, MD, Patricia Zuk, PhD, Reza Jarrahy, MD.
UCLA, Los Angeles, CA, USA.

Purpose
Therapeutic options for obesity and related diseases are limited and often carry significant consequences. New therapies targeting key stages in adipocyte commitment and maturation have shown clinical promise. One potential strategy is to use oxysterols—naturally occurring bioactive molecules—to influence adipose tissue metabolism. To determine if this feature might function as an anti-adipogenic agent, a series of in vitro studies was designed to determine if one isoform of oxysterol, Oxy133, might influence the differentiation process of already committed premature adipose cells. We also examined the in vivo effects of Oxy133 on adipogenesis with adipose-derived stem cells grafting in a mouse model.
Methods
Mouse preadipocyte, 3T3-L1 cell lines was cultured with basal growth medium (GM). After cells reached confluency, they were re-seeded on multi-well plates. Cells were treated with conventional adipogenic medium (AM) containing DMEM, 10% FBS, penicillin-streptomycin with isobutyl methylxanthine, dexamethasone, and insulin with or without Oxy133. A negative control using GM alone was also included. Measuring differentiation specific gene expressions (PPARγ, CEBP, GPD, and Adiposin) tested for active adipogenesis, and an Oil Red O stain established mature adipocyte functionality and morphology. For the in vivo study, human adipose-derived stem cells (hASC) were cultured and maintained until 90% confluence. Cells were then cultured in adipogenic medium (control) or treated with adipogenic medium with Oxy133. We subsequently collected and injected the cells into the subcutaneous tissue on the scalp of nude mice. After 12 weeks, the specimens were harvested for histologic analysis.
Results
Expression of adipogenic differentiation specific genes were reduced in cells exposed to the Oxy133, in contrast to positive control groups. Functionality and morphology of mature adipocytes attenuated in the Oxy133-treated cells. Mice injected with hASCs treated with Oxy133 demonstrated significantly decreased adipose tissue as compared to control animals after 12 weeks.
Conclusions
Our studies demonstrated the ability of Oxy133 to impede differentiation of preadipocytes into mature functional adipocytes in vitro. Oxy133 also impeded adipogenesis of hASCS in vivo. Oxy133 represents a potentially viable therapy for anti-fat agent in obesity.


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