Effects of growth regulation on conditionally-transformed alginate-entrapped insulin secreting cell lines in vitro

The ability to control cell growth is an issue of critical importance for the use of transformed ββ-cell lines within a bioartificial pancreas. Such control can be achieved either by entrapping the cells in a biomaterial that can inhibit cell proliferation or by genetically modifying the cells to regulate growth. Integrating tetracycline-off or -on operon systems into murine insulinoma cell lines (βTCβTC-tet and R7T1, respectively) allows cell growth regulation upon exposure to tetracycline (TC) or its derivative doxycycline (Dox), respectively. However, the effects of this regulatory approach on the long-term phenotypic metabolic and secretory stability of alginate-entrapped cells have yet to be thoroughly investigated. In this study, cultures of βTCβTC-tet and R7T1 cells entrapped in alginate beads were allowed to grow freely, or were growth-regulated, either at the onset, or after 20 days of growth. The data show that growth regulation of alginate-entrapped cells is achievable with chronic administration of the regulatory compound in a concentration-dependent manner. However, as these cultures age, the amount of insulin released does not always reflect the metabolic and histological characteristics of the cultures. This change, coupled with a loss of glucose stimulated insulin secretion in the Dox treated R7T1 cell line, indicate a phenotypic shift of cells with an activated tet-operon. These observations have implications on the selection and long-term function of three-dimensional bioartificial pancreatic constructs that include conditionally transformed ββ-cell lines.