Differential sensitivity to apoptosis between the human small and large intestinal mucosae: Linkage with segment‐specific regulation of Bcl‐2 homologs and involvement of signaling pathways*

Abstract

The small and large intestines differ in their expression profiles of Bcl-2 homologs. Intestinal segment-specific Bcl-2 homolog expression profiles are acquired as early as by mid-gestation (18-20 weeks) in man. In the present study, we examined the question whether such distinctions underlie segment-specific control mechanisms of intestinal cell survival. Using mid-gestation human jejunum and colon organotypic cultures, we analyzed the impact of growth factors (namely insulin; 10 microg/ml) and pharmacological compounds that inhibit signal transduction molecules/pathways (namely tyrosine kinases, Fak, P13-K/Akt, and MEK/Erk) on cell survival and Bcl-2 homolog expression (anti-apoptotic: Bcl-2, Bcl-X(L), Mcl-1; pro-apoptotic: Bax, Bak, Bad). The relative activation levels of p125Fak, p42Erk-2, and p57Akt were analyzed as well. Herein, we report that (1) the inhibition of signal transduction molecules/pathways revealed striking differences in their impact on cell survival in the jejunum and colon (e.g., the inhibition of p125Fak induced apoptosis with a significantly greater extent in the jejunum [approximately 43%] than in the colon [approximately 24%]); (2) sharp distinctions between the two segments were noted in the modulatory effects of the various treatments on Bcl-2 homolog steady-state levels (e.g., inhibition of tyrosine kinase activities in the jejunum down-regulated all anti-apoptotics analyzed while increasing Bax, whereas the same treatment in the colon down-regulated Bcl-X(L) only and increased all pro-apoptotics); and (3) in addition to their differential impact on cell survival and Bcl-2 homolog expression, the MEK/Erk and P13-K/Akt pathways were found to be distinctively regulated in the jejunum and colon mucosae (e.g., insulin in the jejunum increased p42Erk-2 activation without affecting that of p57Akt, whereas the same treatment in the colon decreased p42Erk-2 activation while increasing that of p57Akt). Altogether, these data show that intestinal cell survival is characterized by segment-specific susceptibilities to apoptosis, which are in turn linked with segmental distinctions in the involvement of signaling pathways and the regulation of Bcl-2 homolog steady-state levels. Therefore, these indicate that cell survival is subject to segment-specific control mechanisms along the proximal-distal axis of the intestine.

Human Intestinal Epithelial Cell Survival and Anoikis

“…For treatments, cells were maintained 24 -48 h in either of the following conditions: 1) with 1 M cytochalasin (CD), which has been shown to inhibit Fak at this concentration range without affecting actin filament organization (12,30,31,34,43,44); 2) with 50 g/ml of the monoclonal antibody P4C10 (Invitrogen and Chemicon), which inhibits the binding activities of the ␤ 1 integrin subunit (10,12,30,45,46); 3) with 100 g/ml nonimmune mouse IgGs, as control for experiments using the P4C10 antibody; or 4) with 30 M Ly294002 (Calbiochem), a specific inhibitor of PI3K (13)(14)(15). The working concentrations of the reagents used were established previously (10 -12, 30, 31, 34, 46, 47).…”

“…Total proteins (50 g/well) were resolved by SDS-PAGE on 4 -15% gels (Bio-Rad), electrotransferred onto nitrocellulose membranes (Hybond-ECL; Amersham Biosciences), and probed for Fak, Akt isoforms, and SI, as described previously (12,30,31,34). Broad range molecular mass markers (Bio-Rad) were used as standards.…”

“…For instance, undifferentiated/crypt cells exhibit a susceptibility to apoptosis and/or anoikis that is distinct from the differentiated/villus cells as follows: 1) their expression and regulation of cell death regulators/effectors (e.g. Bcl-2 homologs and caspases) (27, 29 -34); 2) a differential involvement of the PI3K/ Akt and MEK/Erk pathways in their survival (30,31,34); 3) distinct roles enacted by integrins and Fak in the suppression of anoikis (12,30,31,34); and 4) a differentiation state-selective involvement of p38 stress-activated MAPK isoforms in the induction of apoptosis/anoikis (12,34). This in turn fits well with the fact that intestinal epithelial cells express differential integrin profiles and interact with specific basement membrane components along the crypt-villus axis, depending on their state of differentiation (35).…”

Tissue-Specific Attenuation of Endogenous DNA I-Compounds in Rats by Carcinogen Azoxymethane: Possible Role of Dietary Fish Oil in Colon Cancer Prevention

“…In addition, a critical step in the prevention of gastrointestinal cancer by DNA alkylation or oxidation is the removal of DNA adducts, either through repair (66) or targeted apoptosis (53,67). The colon exhibits lower rates of apoptosis and a reduction in sensitivity to apoptosis induction by a variety of stimuli compared with the small intestine (22).…”

“…There are a variety of possible differences in epithelial cells from the small intestine and large intestine that may increase the likelihood of colon carcinogenesis. These include differences in substrate metabolism (20), enhanced DNA damage in colon stem cell populations (21), reduced apoptotic removal of damaged colon cells (21,22), and effects of luminal contents on these cells (23,24).…”