Opposing Roles of Wnt Inhibitors IGFBP-4 and Dkk1 in Cardiac Ischemia by Differential Targeting of LRP5/6 and ß-catenin.

BACKGROUND: Myocardial infarction is one of the leading causes of morbidity and mortality worldwide, triggering irreversible myocardial cell damage and heart failure. The role of low-density lipoprotein receptor-related proteins 5 and 6 (LRP5/6) as coreceptors of the Wnt/ß-catenin pathway in the adult heart remain unknown. Insulin-like growth factor binding protein 4 and dickkopf-related protein 1 (Dkk1) are 2 secreted LRP5/6 binding proteins that play a crucial role in heart development through preventing Wnt/ß-catenin pathway activation. However, their roles in the adult heart remain unexplored. METHODS: To understand the role of LRP5/6 and ß-catenin in the adult heart, we constructed conditional cardiomyocyte-specific LRP5/6 and ß-catenin knockout mice and induced surgical myocardial infarction. We also directly injected recombinant proteins of insulin-like growth factor binding protein 4 and Dkk1 into the heart immediately following myocardial infarction to further examine the mechanisms through which these proteins regulate LRP5/6 and ß-catenin. RESULTS: Deletion of LRP5/6 promoted cardiac ischemic insults. Conversely, deficiency of ß-catenin, a downstream target of LRP5/6, was beneficial in ischemic injury. It is interesting to note that although both insulin-like growth factor binding protein 4 and Dkk1 are secreted Wnt/ß-catenin pathway inhibitors, insulin-like growth factor binding protein 4 protected the ischemic heart by inhibiting ß-catenin, whereas Dkk1 enhanced the injury response mainly through inducing LRP5/6 endocytosis and degradation. CONCLUSIONS: Our findings reveal previously unidentified dual roles of LRP5/6 involved in the cardiomyocyte response to ischemic injury. These findings suggest new therapeutic strategies in ischemic heart disease by fine-tuning LRP5/6 and ß-catenin signaling within the Wnt/ß-catenin pathway.

Increased apoptosis and decreased proliferation of colorectal cancer cells using insulin-like growth factor binding protein-4 gene delivered locally by gene transfer.

OBJECTIVE: Insulin-like growth factor (IGF)-I induces proliferation of transformed cells. Its binding proteins (IGFBP) are involved in local regulation of IGF. This study assessed the effects of overexpression of IGFBP-4 on the development of cancer in vivo. METHOD: Nude mice were subcutaneously inoculated with HT-29 colorectal cancer cells (3 x 10(6)). When the tumour became visible (1 week after inoculation), animals received either 150 microg of mammalian expression vector containing IGFBP-4 cDNA or vector alone (n = 6 each) by peritumoural injection. Tumour size was measured during the growth. After 3 weeks of IGFBP-4 induction, animals were killed and tumour tissue samples were collected for examining the level of IGFBP-4 expression. Tumour mitotic activities were determined by counting numbers of mitotic cells on the tissue section. Apoptosis was investigated by terminal deoxynucleotidyl transferase-mediated dUDP nick end labelling assay. RESULTS: Following IGFBP-4 treatment, tumour showed large necrotic areas, significantly increased numbers of apoptotic cells (36.67 +/- 7.36 vs 7.07 +/- 1.91, P < 0.01 vs control), decreased cells undergoing mitosis (2.31 +/- 0.32 vs 3.61 +/- 0.27, P < 0.01 vs control) and higher expression of IGFBP-4 (P < 0.05 vs control). CONCLUSION: IGFBP-4 gene transfer increased apoptosis and decreased mitosis, but tumour volume was not significantly altered possibly due to cellular debris filling the centre of tumours.

Overexpression of an inhibitory insulin-like growth factor binding protein (IGFBP), IGFBP-4, delays onset of prostate tumor formation.

Insulin-like growth factor (IGF) binding proteins (IGFBPs) have been shown to either inhibit or enhance the action of IGF, or act in an IGF-independent manner in the prostate. We have overexpressed the IGF-inhibitory IGFBP-4 in the malignant M12 prostate epithelial cell line to determine the effects on tumor formation and apoptosis. Overexpression was determined by Northern, Western immunoblot and Western radioligand blot analysis. IGF-induced proliferation was reduced in the IGFBP-4 transfected cells compared with control cells (P < or = 0.01). Colony formation in soft agar was significantly inhibited up to 14 days after plating in the IGFBP-4 transfected cells when compared with the M12 controls (P < or = 0.01): however, in the presence of des(1-3)IGF-I, there was no significant difference between the control and IGFBP-4 transfectants in colony formation in soft agar. Apoptosis in an IGFBP-4 transfected cell line was significantly increased in response to induction by 6-hydroxyurea compared with the control line. When injected s.c. into male athymic/nude mice, a marked delay was noted in tumor formation in animals receiving IGFBP-4 transfected cells (P < or = 0.01). Interestingly, IGFBP-2 protein levels were reduced in the conditioned media of all IGFBP-4 transfected cell cultures. These data indicate that an inhibitory IGFBP may significantly delay the growth of malignant prostate epithelial cells and enhance the sensitivity of these cells to apoptosis.