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Rupalatha Maddala, Vasanth Rao, ; Expression and Distribution of VLK, a secretory tyrosine kinase involved in phosphorylation of extracellular matrix proteins in the Aqueous humor outflow pathway and optic nerve head . Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3264.
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Extracellular matrix (ECM) proteins are recognized to influence resistance to aqueous humor (AH) outflow and intraocular pressure (IOP). To better understand the role of ECM protein biological responses in the AH outflow pathway, here we determined the distribution and expression pattern of a secretory protein kinase-VLK (Vertebrate Lonesome Kinase) regulating tyrosine phosphorylation of ECM and other secretory proteins in the trabecular pathway, cultured trabecular meshwork (TM) cells, AH and optic nerve head.
The expression and distribution pattern of VLK in human and rodent AH outflow pathway, optic nerve head, AH, TM cells and TM cell conditioned media and in the ECM-enriched fraction of TM cells was determined by RT-PCR, immunofluorescence and immunoblot analyses. Basal status of tyrosine phosphorylation of TM cell secretory proteins and ECM proteins was evaluated by immunoblot and immunofluorescence analyses using an anti-phospho tyrosine specific antibody.
The ~32 kDa isoform of VLK is abundant in the AH of human, conditioned media of human TM cells and TM cell lysates, and in the ECM-enriched fraction of human TM cells based on immunoblot and RT-PCR analyses. Immunofluorescence analysis confirmed distribution of VLK in the human and rodent Trabecular AH outflow pathway and optic nerve head, and in cultured human TM cells. Both, secretory proteins and the ECM-enriched fraction of human TM cells were confirmed to exhibit detectable basal tyrosine phosphorylation by immunoblotting analysis.
This initial and first study demonstrating the distribution and expression of VLK kinase in the trabecular outflow pathway, optic nerve head, AH and TM cells along with evidence for ECM proteins of TM cells undergoing tyrosine phosphorylation, suggests that the effects of ECM proteins on homeostasis of AH outflow, IOP and optic nerve head biomechanics may be mediated via modulation of ECM tyrosine phosphorylation status.
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