BackgroundPancreatic β-cells release insulin via an electrogenic response triggered by an increase in plasma glucose concentrations. The critical plasma glucose concentration has been determined to be ~3 mM, at which time both insulin and GABA are released from pancreatic β-cells. Taurine, a β-sulfonic acid, may be transported into cells to balance osmotic pressure. The taurine transporter (TauT) has been described in pancreatic tissue, but the function of taurine in insulin release has not been established. Uptake of taurine by pancreatic β-cells may alter membrane potential and have an effect on ion currents. If taurine uptake does alter β-cell current, it might have an effect on exocytosis of cytoplasmic vesicle. We wished to test the effect of taurine on regulating release of insulin from the pancreatic β-cell.MethodsPancreatic β-cell lines Hit-TI5 (Syrian hamster) and Rin-m (rat insulinoma) were used in these studies. Cells were grown to an 80% confluence on uncoated cover glass in RPMI media containing 10% fetal horse serum. The cells were then adapted to a serum-free, glucose free environment for 24 hours. At that time, the cells were treated with either 1 mM glucose, 1 mM taurine, 1 mM glucose + 1 mM taurine, 3 mM glucose, or 3 mM glucose + 1 mM taurine. The cells were examined by confocal microscopy for cytoplasmic levels of insulin.ResultsIn both cell lines, 1 mM glucose had no effect on insulin levels and served as a control. Cells starved of glucose had a significant reduction (p<0.001) in the level of insulin, but this level was significantly higher than all other treatments. As expected, the 3 mM glucose treatment resulted in a statistically lower (p<0.001) insulin level than control cells. Interestingly, 1 mM taurine also resulted in a statistically lower level of insulin (p<0.001) compared to controls when either no glucose or 1 mM glucose was present. Cells treated with 1 mM taurine plus 3 mM glucose showed a level of insulin similar to that of 3 mM glucose alone.ConclusionsTaurine administration can alter the electrogenic response in β-cell lines, leading to a change in calcium homeostasis and a subsequent decrease in intracellular insulin levels. The consequence of these actions could represent a method of increasing plasma insulin levels leading to a decrease in plasma glucose levels.

Inhibition of outer segment (OS) processing by retinal pigment epithelium (RPE) has been linked to photoreceptor injury and retinopathy onset. Sub-retinal clearance by RPE is facilitated by specialized phagocytosis featuring both RPE-specific and traditional FCγR mediated signaling cascades. As a result of this combinatory approach, RPE are capable of internalizing both specific and non-specific external targets alike. The discovery that lack of c-Met signaling results in impairment of phagocytosis in alveolar and hepatocyte macrophages [1] suggests cMet's role as modulator of this activity in post-mitotic cells secreting HGF. Since activated PI3K has been identified as an activator of Rac1during FCγR mediated phagocytosis, we hypothesize that c-Met activation by HGF and subsequent PI3K activation is capable of mediating OS clearance by RPE.To test our hypotheses, cultured ARPE-19 cells were grown to 70% confluence, then serum starved for 24 hr. Post starvation, cells were exposed to various concentrations of HGF for 24hr before fixation with 2.5% paraformaldehyde and .5% glutaraldehyde. Cells were then prepared for immunohistochemistry for receptor expression (non-phosphorylated and phosphorylated forms), focal adhesion kinase (FAK) and binding of fluorescently-labeled E. coli. Intensity values suggest that ARPE-19 respond maximally to concentrations of 25 ng/ml of HGF when compared to controls (Fig 1). While phosphorylated c-Met was not significantly altered (Fig 2) at 24 hr, this may be attributed to the transient expression of phospho-c-Met following activation by HGF. Our findings suggest that RPE respond to increases of exogenous HGF concentrations by up-regulating its receptor and subsequent second messengers systems. In addition, our data show a significant increase of fluorescently labeled E. coli (Fig 3). Taken together, these findings suggest that RTK cross-talk initiated by c-Met activation may be sufficient in mediating general uptake of external debris by RPE. Future studies including RPE challenge with fluorescently labeled OS during peak c-Met phosphorylation evoked by increased HGF exposure will provide evidence for HGF's role as a mediator of specialized phagocytosis of OS.

Retinopathy of prematurity (ROP) has abnormal temporal and spatial regulations of vascular endothelial growth factor (VEGF) and insulin-like growth factor-1 (IGF-1). These are manifested as retinal neovascularization that requires the activation of receptor tyrosine kinases (RTKs) by VEGF and IGF-1 ligands resulted in downstream pathways promoting cell survival, cell proliferation, vascular permeability and cell migration. In addition, hypoxia-inducible factor (HIF) has been shown to be the transcription factor important in inducing VEGF secretion and expression of the VEGF receptor: VEGFR2 in a variety type of cells. The goals of this study are to demonstrate in vitro tube formation assay as a model of ROP using human retinal microvascular endothelial cells; and to determine the role of HIF-1α in angiogenesis affected by VEGF and IGF1 synergy.Immortalized human retinal pigment epithelial cells (ARPE-19) were grown in Lab Tek II chamber slides, once confluent, the cells were treated with VEGF, IGF-1 or both to observe the upregulation of VEGFR2 and HIF-1α using confocal microscopy. In separate experiments, human retinal microvascular endothelial cell (HRMVEC) lines were seeded in growth factorreduced matrigel medium on 96-well plate for 3 days with activated growth medium in the incubator, then serum free media for 24 hours. The cells were then treated with serum free media supplemented with VEGF, IGF-1 or both under normoxic incubator environment and photographs were taken for 48 -72 hours. The VEGFR2 and HIF-1α in ARPE-19 cells were upregulated with the IGF-1 treatment. Tube formation assay showed more prominent with the presence of both growth factors, but not on control or in the presence of a single growth factor. This study demonstrates the crucial roles of IGF-1, and the signaling proteins involved in VEGFdependent angiogenesis in human retinal microvascular endothelial cells.