2024 AIChE Annual Meeting

Alginate and Poly-L-Ornithine Degeling Techniques for Cell RNA Extraction

Osteoporosis affects approximately 8 million women in the U.S. and is the leading cause of bone fracture with costs expected to reach $95 billion by 2040. Prior to 2002, one widely used treatment for osteoporosis in women who lacked ovarian function consisted of pharmacological hormone treatments (pHTs) that replace the hormones naturally secreted by healthy ovaries. These were found to be effective for reducing osteoporotic fractures but increased women’s risks of developing cardiovascular issues and some cancers. Thus the use of pHTs has decreased in use since 2002. New pharmacological treatments are not more effective than hormone based treatments and have the potential for severe, potentially life threatening side effects as well as minor side effects that lead to poor patient compliance.

One reason for the safety risks inherent to pHTs is their inability to regulate the hypothalamus-pituitary ovary (HPO) axis. A cell-based hormone treatment (cHT) that can secrete the hormones produced by healthy ovaries while regulating the HPO axis is a potential approach to safe and effective use of hormone therapies to mitigate hormone-related osteoporosis. This potential treatment involves creating a two-compartment alginate system consisting of an inner core of granulosa cells and an outer core of theca cells taken from a donor. These constructs are coated with perm-selective poly-L-ornithine as a way to mitigate immune attack on the encapsulated cells.

To better understand the molecular mechanisms of action for our cHT approach, we are interested in removing the cells from the construct for RNA extraction. While there are methods of digesting alginate such as alginase and EDTA/ citrate, the presence of PLO makes removal of cells from the constructs challenging. The ability to isolate cells and RNA from the cHT constructs would allow us to determine the mechanisms by which the cells work.