Synergistic suppression of apoptosis in salivary acinar cells by IGF1 and EGF

Abstract

Tissue homeostasis requires balancing cell proliferation and programmed cell death. IGF1 significantly suppressed etoposide-induced apoptosis, measured by caspase 3 activation and quantitation of cellular subG 1 DNA content, in rat parotid salivary acinar cells (C5). Transduction of C5 cells with an adenovirus expressing a constitutively activated mutant of Akt-suppressed etoposide-induced apoptosis, whereas a kinase-inactive mutant of Akt suppressed the protective effect of IGF1. IGF1 also suppressed apoptosis induced by taxol and brefeldin A. EGF was unable to suppress apoptosis induced by etoposide, but was able to synergize with IGF1 to further suppress caspase 3 activation and DNA cleavage after etoposide treatment. The catalytic activity of Akt was significantly higher following stimulation with both growth factors compared to stimulation with IGF1 or EGF alone. These results suggest that a threshold of activated Akt is required for suppression of apoptosis and the cooperative action of growth factors in regulating salivary gland homeostasis.

MDM2 Is Required for Suppression of Apoptosis by Activated Akt1 in Salivary Acinar Cells

“…1B, lanes 1 and 2), 11.5-fold in sublingual (Fig. 1B, lanes 3 and 4), and 4.9-fold in parotid ( ylation of a substrate peptide (50). In myr-Akt1 female mice, Akt kinase activity increased 1.56-fold in the submandibular gland, 1.78-fold in the sublingual gland, and 1.33-fold in the parotid gland relative to endogenous Akt (Fig.…”

“…In some cases, membranes were then stripped, as previously described (35), reblocked in Tris-buffered saline (10 mM Tris [pH 7.4], 150 mM NaCl, and 0.05% Tween 20) with 5% nonfat dry milk (Carnation), and probed with a second antibody. The activation of Akt kinase activity was quantitated with a radioactive Akt kinase assay kit (Upstate Biotechnology, Lake Placid, NY) using 300 g of tissue lysates according to the manufacturer's instructions (50).…”

“…), and VOL. 26,2006 SUPPRESSION OF SALIVARY ACINAR CELL APOPTOSIS BY Akt 8841 primary parotid or submandibular acinar cells were prepared under sterile conditions similar to previously published protocols (50,51,72 (3,51). Activation of caspase 3 was quantitated with the BioMol QuantiZyme Colorimetric assay kit (Plymouth Meeting, PA).…”

“…Recombinant adenoviruses expressing a kinase-inactive mutant of Akt (Ad-KD-Akt1) or LacZ (Ad-LacZ) were used as previously described (50). Briefly, primary cells were transduced with the desired recombinant adenovirus at equal multiplicities of infection for 1 hour in medium lacking fetal calf serum.…”

Satellite Cell Dysfunction and Impaired IGF-1 Signaling Cause CKD-Induced Muscle Atrophy

“…[33][34][35][36][37] Musaro and colleagues 37,38 demonstrated that overexpression of locally acting IGF-1 (mIGF-1) improves muscle function in mouse models of senescence, injury, myodystrophy, or amyotrophic lateral sclerosis. In muscle and satellite cells, we found that CKD impaired IGF-1R-mediated signaling (Supplemental Figure 2, A and B), and when we suppressed the IGF-1 receptor in mice (IGF-1R-KO), satellite cell activation and muscle regeneration were reduced as in CKD mice (Figure 4).…”