Calcium-induced differentiation in normal human colonoid cultures: Cell-cell / cell-matrix adhesion, barrier formation and tissue integrity

• Published in: PloS one 2019-04, Vol.14 (4), p.e0215122-e0215122
• Main Authors: Attili, Durga, McClintock, Shannon D, Rizvi, Areeba H, Pandya, Shailja, Rehman, Humza, Nadeem, Daniyal M, Richter, Aliah, Thomas, Dafydd, Dame, Michael K, Turgeon, Danielle Kim, Varani, James, Aslam, Muhammad N
• Format: Article
• Language: English
• Subjects: Adenoma - metabolism; Adenoma - pathology; Adhesion; Adhesive strength; Backup software; Biology and Life Sciences; Calcium; Calcium - pharmacology; Calcium compounds; Calcium content; Cell adhesion; Cell Adhesion - drug effects; Cell Communication - drug effects; Cell Culture Techniques; Cell differentiation; Cell Differentiation - drug effects; Cell Proliferation - drug effects; Cell-Matrix Junctions - physiology; Cells, Cultured; Colon; Colon - cytology; Colon - drug effects; Colon - metabolism; Colon cancer; Colorectal cancer; Cytokeratin; Desmosomes; Dietary supplements; Differentiation; Electron microscopy; Histology; Human tissues; Humans; Immunohistochemistry; Integrity; Internal medicine; Intervention; Keratin; Kinases; Magnesium; Medical schools; Medicine and Health Sciences; Microscopy; Minerals - pharmacology; Natural products; Organoids - cytology; Organoids - metabolism; Pathology; Proteins; Proteome - analysis; Risk factors; Scanning electron microscopy; Studies; Supplements; Tissues; Trace elements; Transmission electron microscopy; Tumors; Vitamin deficiency
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0215122

The goal of the study was to assess calcium alone and Aquamin, a multi-mineral natural product that contains magnesium and detectable levels of 72 trace elements in addition to calcium, for capacity to affect growth and differentiation in colonoid cultures derived from histologically-normal human colon tissue. Colonoid cultures were maintained in a low-calcium (0.25 mM) medium or in medium supplemented with an amount of calcium (1.5-3.0 mM), either from calcium alone or Aquamin for a period of two weeks. This was shown in a previous study to induce differentiation in colonoids derived from large adenomas. Changes in growth, morphological features and protein expression profile were assessed at the end of the incubation period using a combination of phase-contrast and scanning electron microscopy, histology and immunohistology, proteomic assessment and transmission electron microscopy. 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 in the absence of intervention (i.e., in control condition). 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 colonoids derived from histologically normal human tissue can undergo differentiation in the presence of a low ambient calcium concentration. However, higher calcium levels 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.