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C McCaig

Publications and source records attributed to C McCaig.

11 recordsLinked to original sources
PubMed2006

Insulin-like growth factor binding protein 3 has opposing actions on malignant and nonmalignant breast epithelial cells that are each reversible and dependent upon cholesterol-stabilized integrin receptor complexes.

IGF-binding protein (IGFBP)-3 is generally considered to have actions that counterbalance those of IGFs and is therefore being developed as a cancer treatment. In breast tumors, however, high levels are associated with aggressive tumors and poor prognosis. Consistent with this we have demonstrated that although IGFBP-3 and a non-IGF-binding fragment (serine phosphorylation domain peptide) reduced attachment and enhanced apoptosis of Hs578T breast cancer cells cultured on collagen or laminin, it promoted their attachment and survival on fibronectin, which is abundant in the matrix of aggressive tumors. We have now examined the factors that determine whether IGFBP-3 has positive or negative actions on breast epithelial cells. IGFBP-3 also promoted survival of Hs578T cells in the presence of an antibody to the beta1-integrin subunit or when cholesterol-stabilized complexes were disrupted. These actions were blocked by IGF-I or a MAPK inhibitor. Serine phosphorylation domain peptide had similar actions on MCF-7 cells that were again reversed on fibronectin or with disruption of cholesterol-stabilized complexes and blocked by the beta1-integrin antibody. In contrast, IGFBP-3 promoted growth and survival for nonmalignant MCF-10A cells, but these effects were again reversed on fibronectin and blocked by the beta1 antibody or a MAPK inhibitor or by disruption of cholesterol-stabilized complexes. On Hs578T cells, IGFBP-3 bound to caveolin-1 and beta1-integrins, enhancing their aggregation, the recruitment of focal adhesion kinase, and the activation of MAPK. In summary, with three breast epithelial cell lines, IGFBP-3 had positive or negative effects on growth and survival dependent upon the status of cholesterol-stabilized integrin receptor complexes.

Apoptosis↗
PubMed2002

A non-IGF binding mutant of IGFBP-3 modulates cell function in breast epithelial cells.

We demonstrated previously that IGFBP-3 alone had no effect on cell death, but dramatically modulated apoptosis in Hs578T IGF non-responsive cells. We investigated whether a non-IGF binding mutant of IGFBP-3 retained its intrinsic actions in this cell line, prior to investigating its actions in IGF-responsive cells (MCF-7 and MCF-10A). In the Hs578T cells, the ceramide analogue, C2-induced apoptosis, non-glycosylated, glycosylated or mutant IGFBP-3 alone had no effect but on co-incubation with C2, all forms of IGFBP-3 markedly accentuated triggered apoptosis. In MCF-7 cells, IGFBP-3 was unable to modulate C2-induced death. In the MCF-10A cells, IGFBP-3 acted as a potent survival factor. IGFBP-3 also affected cell growth in the MCF-10A cells (inhibiting at low doses but increasing growth at higher concentrations). These actions of IGFBP-3 in the MCF-10A cells were independent of IGF-1. IGFBP-3 has differential IGF-independent effects on cell death and growth in normal breast and breast cancer cells.

Apoptosis↗
PubMed2002

Effects of a non-IGF binding mutant of IGFBP-5 on cell death in human breast cancer cells.

We have demonstrated previously that IGFBP-5 alone had no effect on cell death but modulated ceramide-induced apoptosis in Hs578T IGF non-responsive cells. To investigate if IGFBP-5 maintains its intrinsic ability to modulate apoptosis in IGF-responsive cells, we used a non-IGF binding mutant of IGFBP-5. In Hs578T cells, non-glycosylated, glycosylated or mutant IGFBP-5 alone each had no effect on cell death, whereas all forms inhibited ceramide-induced apoptosis. In IGF-responsive MCF-7 cells, each wild type form reduced ceramide-induced cell death but mutant IGFBP-5 was without effect. In the presence of mutant IGFBP-5, however, IGF-I no longer conferred survival and in the presence of wild type IGFBP-5, long R3 IGF-I was also unable to confer survival. In summary, all forms of IGFBP-5 modulated ceramide-induced apoptosis in Hs578T cells. In MCF-7 cells, IGF-I-induced survival could be facilitated by IGFBP-5, but also blocked by IGFBP-5 if association with IGFBP-5 was prevented.

Apoptosis↗
PubMed2002

Differential interactions between IGFBP-3 and transforming growth factor-beta (TGF-beta) in normal vs cancerous breast epithelial cells.

In addition to modulating insulin-like growth factors action, it is now clear that insulin-like growth factor-binding protein-3 also has intrinsic effects on cell growth and survival. We have compared the effects of insulin-like growth factor-binding protein-3 and transforming growth factor-beta on cell proliferation and death of Hs578T cells and the normal breast epithelial cell line, MCF-10A. The growth of MCF-10A cells was inhibited at low concentrations of insulin-like growth factor-binding protein-3 but stimulated at high concentrations. These differential effects were unaffected in the presence of an insulin-like growth factor-I receptor antagonist. A synthetic peptide corresponding to the serine phosphorylation domain of insulin-like growth factor-binding protein-3 (that does not bind to insulin-like growth factors) also mimicked these differential actions. The growth of both cell lines was significantly inhibited by transforming growth factor-beta, this was associated with a 14-fold increase of insulin-like growth factor-binding protein-3 secreted by the Hs578T cells but a five-fold decrease of insulin-like growth factor-binding protein-3 secreted by MCF-10A cells. Replacement doses of exogenous insulin-like growth factor-binding protein-3 overcame the transforming growth factor-beta-induced growth inhibition in the MCF-10A cells. Cell death induced by ceramide was significantly reduced by insulin-like growth factor-binding protein-3 in the MCF-10A cells and depleting insulin-like growth factor-binding protein-3 with transforming growth factor-beta in these cells consequently increased their susceptibility to ceramide. In contrast, insulin-like growth factor-binding protein-3 enhanced apoptosis induced by ceramide in the Hs578T cells but transforming growth factor-beta treated Hs578T cells were resistant to apoptosis. The addition of anti-sense mRNA to insulin-like growth factor-binding protein-3 significantly abrogated this effect of transforming growth factor-beta. These data indicate that insulin-like growth factor-binding protein-3 has intrinsic activity capable of inhibiting or enhancing the growth and survival of breast epithelial cells depending on the cell line and exposure to other cytokines.

Apoptosis↗
PubMed2000

Differential insulin-like growth factor (IGF)-independent interactions of IGF binding protein-3 and IGF binding protein-5 on apoptosis in human breast cancer cells. Involvement of the mitochondria.

We have demonstrated previously in Hs578T cells that insulin-like growth factor binding protein (IGFBP)-3 can significantly accentuate ceramide (C2)-induced apoptosis, but has no effect on cell death induced by integrin detachment [using an arginine-glycine-aspartic acid (RGD)-containing peptide]. In contrast we found that IGFBP-5 could inhibit apoptosis induced by either C2 or integrin detachment. It is now clear that the mitochondria not only provide the energy required for cell viability, but can also play an important role during the commitment phase to apoptosis. We used a mitochondrial respiratory chain inhibitor, antimycin A, at both apoptotic and nonapoptotic doses to further investigate the IGF-independent actions of IGFBP-3 and IGFBP-5 on C2 and RGD-induced apoptosis in the Hs578T cells. Hs578T cells had one of three treatments. 1: They were incubated with increasing doses of antimycin A for 24 h. 2: They were coincubated with an apoptotic dose of either C2 or RGD together with a nonapoptotic dose of antimycin A for 24 h. 3: They were incubated with a binding protein (100 ng/ml) for 24 h followed by coincubation of the binding protein with an apoptotic dose of antimycin A for a further 24 h. Cell viability was assessed by trypan blue dye exclusion and MTT assay, and apoptosis was confirmed and measured by morphologic assessment and flow cytometry. We found that antimycin A initiated apoptosis at 10 micromol/L and above. We also demonstrated that a nonapoptotic dose of antimycin A (0.1 micromol/L) significantly inhibited C2-induced apoptosis, whereas it significantly accentuated RGD-induced cell death. In addition, we found that cell death induced by antimycin A can be accentuated by IGFBP-3 but is not affected by IGFBP-5. These data indicate that IGFBP-3 can directly enhance apoptosis triggered via the mitochondria; either directly by a mitochondrial inhibitor or by C2 (which we demonstrate to act via effects on the mitochondria in this model). IGFBP-5, however, appears to confer survival effects via a distinct pathway not involving the mitochondria.

Antimycin A↗
PubMed1997

Contact guidance of CNS neurites on grooved quartz: influence of groove dimensions, neuronal age and cell type.

We used an in vitro system that eliminates competing guidance cues found in embryos to determine whether substratum topography alone provides important neurite guidance information. Dissociated embryonic Xenopus spinal cord neurons and rat hippocampal neurons were grown on quartz etched with a series of parallel grooves. Xenopus neurites grew parallel to grooves as shallow as 14 nm and as narrow as 1 microm. Hippocampal neurites grew parallel to deep, wide grooves but perpendicular to shallow, narrow ones. Grooved substrata determined the sites at which neurites emerged from somas: Xenopus neurites sprouted from regions parallel to grooves but presumptive axons on rat hippocampal neurons emerged perpendicular to grooves and presumptive dendrites emerged parallel to them. Neurites grew faster in the favored direction of orientation and turned through large angles to align on grooves. The frequency of perpendicular alignment of hippocampal neurites depended on the age of the embryos from which neurons were isolated, suggesting that contact guidance is regulated in development. Collectively, the data indicate that substratum topography is a potent morphogenetic factor for developing CNS neurons and suggest that in addition to a role in pathfinding the geometry of the embryo assists in establishing neuronal polarity. In the companion paper (A. M. Rajnicek and C. D. McCaig (1997) J. Cell Sci. 110, 2915-2924) we explore the cellular mechanism for contact guidance of growth cones.

Animals↗
PubMed1997

Guidance of CNS growth cones by substratum grooves and ridges: effects of inhibitors of the cytoskeleton, calcium channels and signal transduction pathways.

We exploited our observation that embryonic Xenopus spinal neurites align parallel to grooves in a quartz surface and that embryonic rat hippocampal neurites align perpendicular to shallow, narrow grooves (see companion paper: A. M. Rajnicek, S. Britland and C. D. McCaig, 1997) (J. Cell Sci. 110, 2905-2913) to investigate the mechanism of growth cone contact guidance. Substratum topography affected the pattern of growth cone filopodia and microtubules but parallel orientation of Xenopus neurites and perpendicular orientation of hippocampal neurites were unperturbed by cytochalasin B, which virtually eliminated filopodia. Hippocampal growth cone orientation and turning in response to grooves was unaffected by disruption of microtubules using taxol or nocodazole. Gross cytoskeletal reorganization on grooved substrata was therefore not required for growth cone steering. Inhibitors were used to identify the signal transduction pathway for perpendicular alignment of hippocampal neurites. Alignment persisted in the presence of gadolinium chloride, a blocker of stretch-activated calcium channels, the G protein inhibitor pertussis toxin, the protein tyrosine kinase inhibitor genistein, the protein kinase A and G inhibitor HA1004, the protein kinase A inhibitor KT5720and the protein kinase G inhibitor KT5823. Low concentrations of the protein kinase C inhibitors stauro-sporine, bisindolylmaleimide or H-7 did not affect perpendicular orientation but higher concentrations inhibited it. The calcium channel blockers flunarizine, nifedipine and diltiazem also inhibited perpendicular orientation. Influx of calcium and protein kinase C activity therefore appear to be involved in perpendicular contact guidance.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1996

Embryonic Xenopus neurites integrate and respond to simultaneous electrical and adhesive guidance cues.

Nerve cells detect and respond to multiple extrinsic guidance cues during development and regeneration using a motile growth cone. Navigational decisions may be required of the growth cone when different guidance cues are encountered simultaneously. We have tested the relative potencies of two opposing cues by presenting Xenopus spinal cord nerve cells growing on a micropatterned laminin culture substratum with an orthogonal DC electric field. Substrata composed of repeating 25-micron laminin tracks and spaces failed to influence the position of neuritogenesis from nerve cell soma. Once established, however, growth cone movement was constrained by laminin tracks such that neurites of 65% of cells were aligned after 5 h in vitro. Two hours after the application of a 100-140 mV/mm DC field the majority of cells remained aligned with the laminin tracks. Around 70% of Xenopus neurites normally orient cathodally on homogenous laminin substrata; therefore the galvanotropic response was impeded by prior exposure to a patterned laminin substrate. However, a proportion of aligned neurites did orient cathodally and evidence of a response to both directional cues was even found within the same cell. Video-enhanced contrast, differential interference contrast (VEC-DIC) microscopy was used to examine the detailed behavior of growth cones on micropatterned laminin substrata. The present study has demonstrated that growth cones can detect and integrate at least two morphogenetic guidance cues simultaneously. The strength of the galvanotropic response in Xenopus growth cones, however, was often insufficient to override established adhesive guidance in this model system.

Animals↗