The bone morphogenetic protein signaling pathway is upregulated in a mouse model of total parenteral nutrition

Abstract

Total parenteral nutrition (TPN) results in intestinal mucosal atrophic changes due to an absence of enteral nutrition; however, the mechanisms responsible for this are not fully understood. It has been shown that bone morphogenetic protein (BMP) activation inhibits intestinal epithelial cell (EC) proliferation. Therefore, we hypothesized that the BMP pathway could be upregulated by TPN. To address this, we randomly assigned mice to receive TPN or to be enterally fed (control) for 7 d. Mucosal EC isolates were harvested from the entire length of small intestine for RNA and protein measurements. Full-thickness, mid-small bowel was processed for histological examination. TPN increased the abundance of BMP2, BMP4, and BMP type II receptor at the RNA and protein levels. Phosphorylation of Smad1, Smad5, and Smad8 also was greater in the TPN group than in the control, which helped to confirm activation of this pathway. Interestingly, the TPN and control groups did not differ in the mRNA expression of the extracellular soluble bmp antagonists, noggin, gremlin, chordin, or follistatin. Compared to the control group, the expression of c-Myc (cellular myelocytomatosis) mRNA was lower, whereas the level of p21(WAF1/CIP1) was greater, in the TPN group. Because the BMP family may function through suppression of Wnt-beta-catenin signaling, this pathway was also examined. mRNA expression of Wnt 3, Wnt5a, and the Wnt receptor Lrp5 were lower in the TPN group compared to controls. The results suggest that the BMP signaling pathway may be involved in the development of intestinal mucosal atrophy due to TPN administration.

FIGURE 1

Expression of intestinal mucosal BMP2 (A,C,D) and BMP4 (B,E,F) detected by Western blotting and immunohistochemical staining in control and TPN mice after 3 d (Expt. 1). Values are means ± SD, n = 6–8. **Different from TPN, P < 0.001. Immunohistochemical staining was moderately weak in controls and greater in the TPN group. Staining was predominately in the lower one-third of villi and the crypt region. A color version of this figure is available (Supplemental Fig. 1).

FIGURE 2

Expression of intestinal mucosal Bmpr1a (A,C,D) and p-Smad 1/5/8 (B,E,F) as detected by Western blotting and immunohistochemical staining (Expt. 1). Values are means ± SD, n = 6–8. *Different from TPN, P < 0.05. Immunohistochemical staining of Bmpr1a and p-Smad 1/5/8 increased in the TPN group. Immunoreactivity to p-Smad 1/5/8 proteins showed a predominant expression in the lower one-third of villi and in the crypt region. A color version of this figure is available (Supplemental Fig. 2).

FIGURE 3

Protein expression of the total PTEN and p-PTEN (A) and p- Erk1/2 (B) from intestinal mucosal specimens as measured by immunoblot analysis (Expt. 1). Values are means ± SD, n = 6–8. *Different from TPN, P < 0.05.

FIGURE 4

Representative histologic images of EC proliferation as detected by BrdU staining in mouse jejunum. Note a marked decrease in BrdU staining at 7 d of TPN. See text for quantitative results. A color version of this figure is available (Supplemental Fig. 3).