CALCIUM-INDUCED DIFFERENTIATION IN NORMAL HUMAN COLONOID CULTURES: Cell-cell / cell-matrix adhesion, barrier formation and tissue integrity

Abstract

Colonoid cultures were established from histologically-normal human colon tissue and maintained in a low-calcium (0.25 mM) medium or in medium supplemented with an amount of calcium (1.5 - 3.0 mM) that was shown in a previous study to induce differentiation in colonoids derived from large adenomas. Calcium alone was compared to Aquamin, a multi-mineral natural product that contains magnesium and detectable levels of 72 additional trace elements in addition to calcium. Unlike the previously-studied tumor-derived colonoids (which remained un-differentiated in the absence of calcium-supplementation), normal tissue colonoids underwent differentiation as indicated by gross and microscopic appearance, a low proliferative index and high-level expression of cytokeratin 20 (CK20) in the absence of intervention. Only modest additional changes were seen in these parameters with either calcium alone or Aquamin (providing up to 3.0 mM calcium). In spite of this, proteomic analysis and immunohistochemistry revealed that both interventions induced strong up-regulation of proteins that promote cell-cell and cell-matrix adhesive functions, barrier formation and tissue integrity. Transmission electron microscopy revealed an increase in desmosomes in response to intervention. These findings demonstrate that histologically normal human colonoids can undergo differentiation in the presence of a low ambient calcium concentration. However, higher calcium levels can induce elaboration of proteins that promote cell-cell and cell-matrix adhesion. These changes could lead to improved barrier function and improved colon tissue health.