PDLIM2 integrates cytoskeletal signaling with gene expression to enable reversible differentiation of epithelial cancer cells. PDLIM2 associates with the COP9 signalosome and controls its nuclear translocation and the stability of key transcription factors necessary for either a mesenchymal or an epithelial phenotype.

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Matrix ligation of integrins ␣v␤3/␣v␤5 is critical for endothelial survival and angiogenesis. We have previously shown that ceramide, a proapoptotic lipid second messenger, increases during endothelial anoikis (detachment-induced apoptosis). We now show that RGDfV, an integrin ␣v␤3/␣v␤5 cyclic function-blocking peptide, increased ceramide and decreased sphingomyelin in human brain microvascular endothelial cells (HBMECs) plated on vitronectin, suggesting that sphingomyelin hydrolysis contributes to RGDfVinduced ceramide increase. Desipramine and imipramine, inhibitors of acid sphingomyelinase (ASMase), suppressed RGDfV-induced ceramide increase. Importantly, desipramine, imipramine, and a third ASMase inhibitor, SR33557, but not inhibitors of neutral sphingomyelinase, suppressed RGDfV-induced apoptosis, suggesting that ASMase was required for integrin-mediated apoptosis. Myriocin, an inhibitor of de novo ceramide synthesis, had no effect on RGDfV-induced HBMEC apoptosis. Interestingly, ASMase inhibitors also suppressed the RGDfV-induced loss of spreading on vitronectin. RGDfV IntroductionIntegrins are heterodimeric cell-surface receptors composed of ␣ and ␤ subunits. Integrins regulate functions such as cell movement, gene expression, cell cycle regulation, and cell survival, using complex signaling cascades with both inside-out as well as outside-in signaling. [1][2][3][4] Integrins ␣v␤3 and ␣v␤5 are preferentially expressed on angiogenic endothelial cells, and their inhibition induces apoptosis. [5][6][7][8][9] The signal mediated by ␣v␤3 and ␣v␤5 requires their binding to matrix proteins such as vitronectin, fibronectin, osteopontin, and tenascin. This binding is via arginineglycine-aspartic acid (RGD) sequences and can be specifically abrogated by function-blocking cyclic RGDfV peptides containing this sequence. 10 In vivo, inhibition of integrins ␣v␤3 and/or ␣v␤5 results in suppression of new blood vessel formation, disruption of existing angiogenic vasculature, inhibition of tumor growth, and tumor regression, [5][6][7][8][11][12][13] providing rationale for inhibition of integrins ␣v␤3 and ␣v␤5 in antiangiogenic therapy. Indeed, one of the cyclic RGDfV peptides (cilengitide, EMD 121974), monoclonal antibodies, and other inhibitors of integrins ␣v␤3 and/or ␣v␤5 are currently in clinical trials that attempt to harness their antiangiogenic potential. [14][15][16][17] Integrin ␣v␤3/␣v␤5 signaling regulates migration, proliferation, and survival of endothelial cells, thereby affecting angiogenesis. Signaling from integrin ␣v␤3 leads to inhibition of p53 transcriptional activity, decreased expression of p21 WAF1/CIP1 , and suppression of the bax cell death pathway in endothelial cells. 12 However, as demonstrated in wild-type and p53-null mice, inhibition of ␣v-integrin ligation in developing retinas induces p21 WAF1 independently of p53, underscoring the complexity and diversity of this pathway. 18 On osteopontin, the ␣v␤3-dependent signals for endothelial cell survival are mediated via nuclear factor B (NF-B...

Aims/hypothesis. Premature death of retinal pericytes is a pathophysiological hallmark of diabetic retinopathy. Among the mechanisms proposed for pericyte death is exposure to AGE, which accumulate during diabetes. The current study used an in vitro model, whereby retinal pericytes were exposed to AGE-modified substrate and the mechanisms underlying pericyte death explored. Methods. Pericytes were isolated from bovine retinal capillaries and propagated on AGE-modified basement membrane (BM) extract or non-modified native BM. The extent of AGE modification was analysed. Proliferative responses of retinal pericytes propagated on AGE-modified BM were investigated using a 5-bromo-2-deoxy-uridine-based assay. The effect of extrinsically added platelet-derived growth factor (PDGF) isoforms on these proliferative responses was also analysed alongside mRNA expression of the PDGF receptors. Apoptotic death of retinal pericytes grown on AGE-modified BM was investigated using terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling labelling, mitochondrial membrane depolarisation and by morphological assessment. We also measured both the ability of PDGF to reverse Akt dephosphorylation that was mediated by AGE-modified BM, and increased pericyte apoptosis. Results. Retinal pericytes exposed to AGE-modified BM showed reduced proliferative responses in comparison to controls (p<0.05-0.01), although this effect was reversed at low-AGE modifications. PDGF mRNA levels were differentially altered by exposure to low and high AGE levels, and AGE-modified BM caused significantly increased apoptosis in retinal pericytes. Pre-treatment of AGE-modified BM with PDGF-AA and -BB reversed the apoptosis (p<0.05-0.001) and restored Akt phosphorylation in retinal pericytes. Conclusions/interpretation. Evidence suggests that substrate-derived AGE such as those that occur during diabetes could have a major influence on retinal pericyte survival. During diabetic retinopathy, AGE modification of vascular BM may reduce bioavailability of pro-survival factors for retinal pericytes.