title: Alginate hydrogels for three-dimensional organ culture of ovaries and oviducts.
abstract: Ovarian cancer is the fifth leading cause of cancer deaths in women and has a 63% mortality rate in the United States(1). The cell type of origin for ovarian cancers is still in question and might be either the ovarian surface epithelium (OSE) or the distal epithelium of the fallopian tube fimbriae(2,3). Culturing the normal cells as a primary culture in vitro will enable scientists to model specific changes that might lead to ovarian cancer in the distinct epithelium, thereby definitively determining the cell type of origin. This will allow development of more accurate biomarkers, animal models with tissue-specific gene changes, and better prevention strategies targeted to this disease. Maintaining normal cells in alginate hydrogels promotes short term in vitro culture of cells in their three-dimensional context and permits introduction of plasmid DNA, siRNA, and small molecules. By culturing organs in pieces that are derived from strategic cuts using a scalpel, several cultures from a single organ can be generated, increasing the number of experiments from a single animal. These cuts model aspects of ovulation leading to proliferation of the OSE, which is associated with ovarian cancer formation. Cell types such as the OSE that do not grow well on plastic surfaces can be cultured using this method and facilitate investigation into normal cellular processes or the earliest events in cancer formation(4). Alginate hydrogels can be used to support the growth of many types of tissues(5). Alginate is a linear polysaccharide composed of repeating units of β-D-mannuronic acid and α-L-guluronic acid that can be crosslinked with calcium ions, resulting in a gentle gelling action that does not damage tissues(6,7). Like other three-dimensional cell culture matrices such as Matrigel, alginate provides mechanical support for tissues; however, proteins are not reactive with the alginate matrix, and therefore alginate functions as a synthetic extracellular matrix that does not initiate cell signaling(5). The alginate hydrogel floats in standard cell culture medium and supports the architecture of the tissue growth in vitro. A method is presented for the preparation, separation, and embedding of ovarian and oviductal organ pieces into alginate hydrogels, which can be maintained in culture for up to two weeks. The enzymatic release of cells for analysis of proteins and RNA samples from the organ culture is also described. Finally, the growth of primary cell types is possible without genetic immortalization from mice and permits investigators to use knockout and transgenic mice.
authors.affiliation: Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, USA.
authors.forename: Shelby M
authors.initials: SM
authors.lastname: King
authors.forename: Suzanne
authors.initials: S
authors.lastname: Quartuccio
authors.forename: Tyvette S
authors.initials: TS
authors.lastname: Hilliard
authors.forename: Kari
authors.initials: K
authors.lastname: Inoue
authors.forename: Joanna E
authors.initials: JE
authors.lastname: Burdette
mesh_terms: D000464:Alginates / Q000737:chemistry*; D000818:Animals; D005260:Female; D020100:Hydrogels / Q000737:chemistry*; D051379:Mice; D009924:Organ Culture Techniques / Q000379:methods*; D010053:Ovary / Q000033:anatomy & histology* / Q000737:chemistry / Q000648:ultrastructure; D010057:Oviducts / Q000033:anatomy & histology* / Q000737:chemistry / Q000648:ultrastructure
chemical_list: D000464:Alginates; D020100:Hydrogels