Introduction: Chromogranin A is a neuroendocrine secretory product and its loss is a feature of malignant NEN de-differentiation. We hypothesized that chromogranin A fragments were differentially expressed during NEN metastasis and played a role in the regulation of NEN proliferation. Methods: Chromogranin A mRNA (PCR) and protein (ELISA/western blot) were studied in 10 normal human mucosa, 5 enterochromaffin cell preparations, 26 small intestinal NEN primaries and 9 liver metastases. Cell viability (WST-1 assay), proliferation (bromodeoxyuridine ELISA) and expression of AKT/AKT-P (CASE ELISA/western blot) in response to chromogranin A silencing, inhibition of prohormone convertase and mTOR inhibition (RAD001/AKT antisense) as well as different chromogranin A fragments were examined in 4 SI-NEN cell lines. Results: Chromogranin A mRNA and protein levels were increased (37-340 fold, p < 0.0001) in small intestinal NENs compared to normal enterochromaffin cells. Western blot identified chromogranin A-associated processing bands including vasostatin in small intestinal NENs as well as up-regulated expression of prohormone convertase in metastases. Proliferation in small intestinal NEN cell lines was decreased by silencing chromogranin A as well as by inhibition of prohormone convertase (p < 0.05). This inhibition also decreased secretion of chromogranin A (p < 0.05) and 5-HT (p < 0.05) as well as expression of vasostatin. Metastatic small intestinal NEN cell lines were stimulated (50-80%, p < 0.05) and AKT phosphorylated (Ser473: p < 0.05) by vasostatin I, which was completely reversed by RAD001 (p < 0.01) and AKT antisense (p < 0.05) while chromostatin inhibited proliferation (∼50%, p < 0.05). Conclusion: Chromogranin A was differentially regulated in primary and metastatic small intestinal NENs and cell lines. Chromogranin A fragments regulated metastatic small intestinal NEN proliferation via the AKT pathway indicating that CgA plays a far more complex role in the biology of these tumors than previously considered. © 2013 Giovinazzo et al.
Giovinazzo, F., Schimmack, S., Svejda, B., Alaimo, D., Pfragner, R., Modlin, I., & Kidd, M. (2013). Chromogranin a and its fragments as regulators of small intestinal neuroendocrine neoplasm proliferation. PLoS ONE, 8(11). https://doi.org/10.1371/journal.pone.0081111