Background: Cyclooxygenase (COX) is the rate-limiting enzyme that catalyzes the formation of prostaglandins. The inducible isoform of COX (COX-2) is highly expressed in aggressive metastatic breast cancers and may play a critical role in cancer progression (i.e. growth and metastasis). However, the exact mechanism(s) for COX-2-enhanced metastasis has yet to be clearly defined. It is well established that one of the direct results of COX-2 action is increased prostaglandin production, especially prostaglandin E 2 (PGE 2 ). Here, we correlate the inhibition of COX-2 activity with decreased breast cancer cell proliferation, migration, invasion and matrix metalloproteinase (MMP) expression.

An investigation was conducted to assess the effects of various beta-galactoside specific lectins on the growth of vascular cells in vitro. The plant lectins from peanut (Arachis hypogaea), mushroom (Agaricus bisporus), and coral tree (Erythrina corallodendron) were used in these studies with the ultimate purpose of comparing those findings with data derived with the lectin isolated from rat lung. Peanut lectin was added to confluent and subconfluent cultures of smooth muscle cells (SMC), pulmonary arterial (PEC), and aortic endothelial cells (BAEC) at concentrations of 2, 3.5, and 7.0 micrograms/ml. There was a dose-dependent increase in cell proliferation for both confluent and subconfluent SMC, with maximal stimulation noted between 3.5 and 7 micrograms/ml of peanut lectin. A dose-dependent stimulation of PEC proliferation was also found with maximal stimulation between 3.5 and 7.0 micrograms/ml. Peanut lectin did not stimulate BAEC to multiply. The stimulation of PEC and SMC by peanut lectin could be prevented by the addition of 50 mM lactose. Peanut and mushroom lectin stimulated the proliferation of sparse cultures of SMC in a dose-dependent fashion in both standard (10% fetal bovine serum, or FBS) or low (0.5% FBS) serum to about the same degree. Coral tree lectin did not have a significant stimulation of proliferation under either serum conditions. The incorporation of [3H]thymidine into the DNA of PEC was increased 30 and 150% by peanut lectin and lung galaptin, respectively, under standard serum conditions. However, under low serum conditions, both lectins increased incorporation by about the same extent (93 and 78% for peanut lectin and galaptin, respectively). Both lectins produced a 30% increase in DNA synthesis by SMC under standard serum conditions, and about a 200% increase under low serum conditions. These studies indicate that beta-galactoside specific lectins such as lung galaptin have mitogenic activity toward vascular cells.

Galaptins are small, soluble, lectins with a specificity for beta-galactose residues. Many galaptins are inactivated by atmospheric oxygen and are protected by disulphide-reducing reagents. We find that each subunit of rat lung galaptin contains one residue of tryptophan and six of cysteine. Oxygen inactivates rat lung galaptin by oxidation of the cysteine residues. During oxidation, the normal dimeric structure is maintained and all disulphide bonds are formed within individual subunits. Exogenous thiols protect against inactivation, but oxidized thiols accelerate inactivation. Human lung fibroblast galaptin is almost completely inactivated within 1 h in tissue culture medium at 37 degrees C. Alkylation of native rat lung galaptin with iodoacetate or ethyleneimine causes substantial loss of activity. The dimeric galaptin structure is maintained. In contrast, alkylation with iodoacetamide yields carboxamidomethyl-galaptin, which is fully active and stable to atmospheric oxygen in the absence of disulphide-reducing reagents. This derivative is very useful for studies of galaptin properties and function.

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BackgroundNeovascularization (angiogenesis) is a multistep process, controlled by opposing regulatory factors, which plays a crucial role in several ocular diseases. It often results in vitreous hemorrhage, retinal detachment, neovascularization glaucoma and subsequent vision loss. Hypoxia is considered to be one of the key factors to trigger angiogenesis by inducing angiogenic factors (like VEGF) and their receptors mediated by hypoxia inducible factor-1 (HIF-1α) a critical transcriptional factor. Another factor, nuclear factor kappa B (NFκB) also regulates many of the genes required for neovascularization, and can also be activated by hypoxia. The aim of this study was to elucidate the mechanism of interaction between HRPC and HUVEC that modulates a neovascularization response.MethodsHuman retinal progenitor cells (HRPC) and human umbilical vein endothelial cells (HUVEC) were cultured/co-cultured under normoxia (control) (20% O2) or hypoxia (1% O2) condition for 24 hr. Controls were monolayer cultures of each cell type maintained alone. We examined the secretion of VEGF by ELISA and influence of conditioned media on blood vessel growth (capillary-like structures) via an angiogenesis assay. Total RNA and protein were extracted from the HRPC and HUVEC (cultured and co-cultured) and analyzed for the expression of VEGF, VEGFR-2, NFκB and HIF-1α by RT-PCR and Western blotting. The cellular localization of NFκB and HIF-1α were studied by immunofluorescence and Western blotting.ResultsWe found that hypoxia increased exogenous VEGF expression 4-fold in HRPC with a further 2-fold increase when cultured with HUVEC. Additionally, we found that hypoxia induced the expression of the VEGF receptor (VEGFR-2) for HRPC co-cultured with HUVEC. Hypoxia treatment significantly enhanced (8- to 10-fold higher than normoxia controls) VEGF secretion into media whether cells were cultured alone or in a co-culture. Also, hypoxia was found to result in a 3- and 2-fold increase in NFκB and HIF-1α mRNA expression by HRPC and a 4- and 6-fold increase in NFκB and HIF-1α protein by co-cultures, whether non-contacting or contacting.Treatment of HRPC cells with hypoxic HUVEC-CM activated and promoted the translocation of NFκB and HIF-1α to the nuclear compartment. This finding was subsequently confirmed by finding that hypoxic HUVEC-CM resulted in higher expression of NFκB and HIF-1α in the nuclear fraction of HRPC and corresponding decrease in cytoplasmic NFκB and HIF-1α. Lastly, hypoxic conditioned media induced a greater formation of capillary-like structures (angiogenic response) compared to control conditioned media. This effect was attenuated by exogenous anti-human VEGF antibody, suggesting that VEGF was the primary factor in the hypoxic conditioned media responsible for the angiogenic response.ConclusionsThese findings suggest that intercellular communications between HRPC and HUVEC lead to the modulation of expression of transcription factors associated with the production of pro-angiogenic factors under hypoxic conditions, which are n...