Influence of indigenous microbiota on activities of alkaline phosphatase, phosphodiesterase I, and thymidine kinase in mouse enterocytes

Abstract

An indigenous microflora introduced into the gastrointestinal tracts of animals in a population of germfree mice affected in different ways three enzymes in small bowel enterocytes. Cells were obtained by techniques designed for sequentially removing enterocytes from the tip of the villus to the crypts of Lieberkuhn. The specific activity of alkaline phosphatase, a component of the enterocyte microvillous membrane, did not differ in cells isolated from germfree mice and from those associated with a microflora, while that of phosphodiesterase I, also a part of the microvillous membrane, was approximately 1.5-fold greater in the suspensions from all levels of the villi in germfree mice than in those from the associated animals. By contrast, the specific activity of thymidine kinase, a cytosol enzyme, in suspensions in which the cells were isolated from the lower portion of the villi and crypts was about one-half as great in cells from germfree mice as in those from the same regions of animals with a microbiota. These results support the hypothesis that activities of certain enzymes involved in metabolism, uptake, and incorporation by enterocytes of components of dietary nucleic acids are influenced by a microflora.

Intestinal alkaline phosphatase: multiple biological roles in maintenance of intestinal homeostasis and modulation by diet

“…Conventional chicks, as zebrafish, tend to display a higher intestinal AP activity than germ‐free chicks 61 . However, AP activity was not found to vary significantly in pigs, 17 while it decreased along with bacterial colonization in mice and rats 62–69 . Therefore, AP activity in the intestine may be modulated by the microbiota differently depending on the animal species.…”

Better living through microbial action: the benefits of the mammalian gastrointestinal microbiota on the host

“…Although intrinsic factors like glucocorticoid hormones drive a developmental scheme, the gut microbiota influences intestinal epithelial ontogony. Activities of brush‐border enzymes, such as disaccharide hydrolases, peptidases and alkaline phophatase, which are markers of enterocyte differentiation, are modified by intestinal colonization (Whitt and Savage, 1988; Kozakova et al ., 2006; Siggers et al ., 2008). Postnatal gene expression of β1,4‐galactosyltransferase (βGT), a marker of glycosyltransferase development, increases rapidly in CONV mice, but stays at low levels in GF mice.…”

Stability of enterocytes and certain enzymatic activities in suspensions of cells from the villous tip to the crypt of Lieberkühn of the mouse small intestine

“…Therefore, our emphasis is primarily on villous cells. To study such mechanisms, we compare germfree mice with animals associated with a flora (21,30,31). Equivalent percentages of cell mass, protein content, enterocytes, and enzymatic activities can be removed from the two types of animals (data not shown) (31).…”

“…Thymidine kinase activity was reproducibly assayed when the products on the filters were washed in three solutions with shaking. We have successfully used this procedure to study how lactobacilli and a complex indigenous microbiota affect alkaline phosphatase, phosphodiesterase, and thymidine kinase activities in enterocytes of mice (30,31).…”

“…In this laboratory, we have been assaying the activities of alkaline phosphatase, phosphodiesterase, and thymidine kinase in cell suspensions containing enterocytes from the upper small intestine of germfree mice and ex-germfree mice associated with an indigenous gastrointestinal microflora (21,30,31). To perform such assays, we had to isolate epithelial cells in a sequential fashion with a minimum number of interfering factors.…”