Investigation of murine host sex as a biological variable in epithelial barrier function and muscle contractility in human intestinal organoids

Abstract

Intestinal failure (IF) occurs when intestinal surface area or function is not sufficient to support digestion and nutrient absorption. Human intestinal organoid (HIO)-derived tissue-engineered intestine is a potential cure for IF. Research to date has demonstrated successful HIO transplantation (tHIO) into mice with significant in vivo maturation. An area lacking in the literature is exploration of murine host sex as a biological variable (SABV) in tHIO function. In this study, we investigate murine host SABV in tHIO epithelial barrier function and muscle contractility. HIOs were generated in vitro and transplanted into nonobese diabetic, severe combined immunodeficiency gamma chain deficient male and female mice. tHIOs were harvested after 8-12 weeks in vivo. Reverse transcriptase polymerase chain reaction and immunohistochemistry were conducted to compare tight junctions and contractility-related markers in tHIOs. An Ussing chamber and contractility apparatus were used to evaluate tHIO epithelial barrier and muscle contractile function, respectively. The expression and morphology of tight junction and contractility-related markers from tHIOs in male and female murine hosts is not significantly different. Epithelial barrier function as measured by transepithelial resistance, short circuit current, and fluorescein isothiocyanate-dextran permeability is no different in tHIOs from male and female hosts, although these results may be limited by HIO epithelial immaturity and a short flux time. Muscle contractility as measured by total contractile activity, amplitude, frequency, and tension is not significantly different in tHIOs from male and female hosts. The data suggest that murine host sex may not be a significant biological variable influencing tHIO function, specifically epithelial barrier maintenance and muscle contractility, though limitations exist in our model.

Engineered Synthetic Matrices for Human Intestinal Organoid Culture and Therapeutic Delivery

“…This effect may be due to cell line variability as it is known that PDOs retain the donor's identity. [ 42–44 ] When compared to the mouse intestinal organoids studies discussed in this section, the patient‐derived organoids exhibited a cyst‐like morphology instead of crypt‐like containing structures as observed with the murine organoids. Differences in cell source species may contribute to this observation.…”

“…[41] PDOs have been shown to maintain tissue donor identity, including regional intestinal tissue characteristics, [42] donor's age, [43] and donor's sex. [44] Additionally, the donor's gastrointestinal disease state is also maintained, thus different disease organoids models can be developed. [1,16,45] Depending on the state of the initial tissue source (e.g., fresh biopsy, established PDO lines), either freshly isolated intestinal crypt fragments or single cells are embedded with a 3D basement membrane (e.g., Matrigel).…”

“…[ 41 ] PDOs have been shown to maintain tissue donor identity, including regional intestinal tissue characteristics, [ 42 ] donor's age, [ 43 ] and donor's sex. [ 44 ] Additionally, the donor's gastrointestinal disease state is also maintained, thus different disease organoids models can be developed. [ 1,16,45 ]…”