Activation of the α₂β₁ integrin‐mediated malignant phenotype on type I collagen in pancreatic cancer cells by shifts in the concentrations of extracellular Mg²⁺ and Ca²⁺

Abstract

The authors have previously demonstrated that a 2 b 1 integrin-mediated pancreatic cancer cell adhesion to Type I collagen is Mg 21 -dependent, inhibited by Ca 21 , and that this integrin, purified from cell lysates using Type I-collagen-sepharose in Mg 21 , can be eluted with Ca 21 . In the present study, the authors examined the divalent cation-dependency of a 2 b 1 integrin-mediated pancreatic cancer cell adhesion, migration and proliferation on Type I collagen, an extracellular matrix protein shown to be highly up-regulated, and to promote the malignant phenotype in vitro and in vivo. Integrins are a family of heterodimeric transmembrane receptor proteins that mediate the binding of cells to the extracellular matrix (ECM) as well as, in some cases, other cells. [1][2][3][4][5][6] A common characteristic of all integrins is their absolute requirement for divalent cations, such as Mg 21 and Ca 21 , to function. 3,4,[6][7][8][9][10][11] These cations presumably exert their effects by binding to the 3-5 putative cation-binding domains located on all integrin a subunits, 12 and possibly by interacting directly with b subunits. 13 We have recently demonstrated that a 2 b 1 integrin-mediated pancreatic cancer cell adhesion to Type I collagen, an ECM protein shown to be highly upregulated in pancreatic cancer [14][15][16][17] and to promote the malignant phenotype in vitro and in vivo, 18-25 is promoted in 1.5 mM Mg 21 and inhibited in 1.5 mM Ca 21 , 21 and that the a 2 b 1 integrin from pancreatic cancer cell lysates bound specifically to Type I collagen by affinity chromatography in 3 mM Mg 21 , can be eluted with 3 mM Ca. 2126 These data are consistent with our previous observations regarding the a 2 b 1 integrin, Type I collagen, and the various cell types involved in cutaneous wound repair, 27-29 of which strong parallels to the pancreatic cancer paradigm have been described. 30,31 During normal cutaneous wound healing in vivo, shifts in the concentrations of extracellular Mg 21 and Ca 21 have been shown to occur early in the process, activating the a 2 b 1 integrin-mediated migration of various wound healing cell types on Type I collagen, including epithelial keratinocytes. 29 This shift in extracellular divalent cation concentration is characterized by increased Mg 21 and decreased Ca 21 . Interestingly, similar shifts probably occur in pancreatic cancer as well as pancreatic juice, produced in the range of 1,500-2,000 mL/day, contains Mg 21 in the range of 137-244 lM and Ca 21 in the range of 0. 111-0.197 lM. 32 This represents a more than 1,200-fold increase of extracellular Mg 21 relative to Ca 21 . As pancreatic ductal epithelial basement membranes have been shown to be discontinuous or absent in pancreatic cancer, 15,16,33 pancreatic juice could be expected to leak into the pancreatic cancer tumor microenvironment. Additionally, and similar to cutaneous wound healing, 34 solid tumors are distinguished by increased Mg 21 load, even at the expense of healthy adjacent tissue. 35,36 Collectively, these data...

Saccharomyces boulardii Improves Intestinal Cell Restitution through Activation of the α2β1 Integrin Collagen Receptor

“…In accordance with this, some data have highlighted dynamic changes in integrin affinities for ligands. These changes are thought to occur in response to extracellular cues, and can modify cell migration [35]. In line with these data, we demonstrated that factors secreted by Sb can activate α2β1integrin.…”

Divalent Cations Modulate α2β1 Integrin-Mediated Malignancy in a Novel 3-Dimensional In Vitro Model of Pancreatic Cancer

“…We have also shown in 2D cell adhesion assays on type I collagen, that α 2 β 1 integrin-mediated pancreatic cancer cell attachment is Mg 2+ -dependent and inhibited by Ca 2+ 22,27,28. In the present study, we examined the divalent cation-dependency of pancreatic cancer cell adhesion on 3D type I collagen in titration experiments.…”

“…Additionally, we demonstrated further that the α 2 β 1 integrin specifically mediates pancreatic cancer cell proliferation on type I collagen in 2D in the presence of altered divalent cations 27,28. We therefore examined the specificity of divalent cation-dependent, α 2 β 1 integrin-mediated interactions of pancreatic cancer cells with 3D type I collagen/GAG scaffolds using inhibition of proliferation assays and anti-integrin function-blocking monoclonal antibodies.…”

“…Pancreatic cancer cell proliferation was determined after 72 hours as described above in the presence or absence of the indicated function-blocking anti-integrin monoclonal antibodies (P1E6, anti-α 2 ; P1D6, anti-α 5 ; P5D2, anti-β 1 )47-49 at 25 μg/ml (Millipore Corp., Temecula, CA). Antibody concentrations used are based on previous inhibition of adhesion, migration, and proliferation assays with these pancreatic cancer cell lines as well as fibroblasts, keratinocytes and endothelial cells in both 2D and 3D in vitro assays 21,26,27,29,46,50. Antibodies were added 24 hours after initial cell adhesion as previously described 27…”

“…And while Mg 2+ supports and Ca 2+ inhibits pancreatic cancer cell migration and proliferation on type I collagen, when Mg 2+ is present at concentrations that promote maximal cell adhesion and Ca 2+ is present at concentrations less than Mg 2+ , both cell migration and proliferation can be significantly increased by as much as two-fold 27,28. These data are consistent with our previous observations regarding α 2 β 1 integrin-mediated cellular interactions with type I collagen, including the various cell types involved in cutaneous wound repair29-31, of which strong parallels to the pancreatic cancer paradigm have been described 32,33…”