Project description:Inflammatory signals must be regulated and kept in check in order to prevent tissue damage. This is especially true in the cornea, where damage can induce loss of transparency, essential for vision. Toll-like receptors (TLRs) are present at the ocular surface and, in addition to being protective against infection, have also been implicated in the pathogenesis of dry eye syndrome, an inflammatory condition that affects millions of individuals in the United States each year.  Therefore, an important area of research is the development of new anti-inflammatory therapeutics that limit aberrant ocular surface inflammation. Vitamin D has been studied for its role in suppressing inflammation in other tissues. In previous studies, we have demonstrated that vitamin D is able to decrease proinflammatory mediators induced by TLRs in human corneal epithelial cells (HCEC). Therefore, the goal of the current study was to examine this mechanism further through an evaluation of vitamin Dâ??s influence on gene expression in two different HCEC cell lines. Microarray comparing SV40-HCEC, human corneal epithelial cells, treated for 6 hours with Vitamin D (1,25D3) and control (0.1% ethanol) Two-condition arrays: SV40-HCEC cells were treated with either 100nM 1,25D3 or 0.1% Ethanol for 6h. Two biological replicates, one per array, with a dye swap (technical replicate).
Project description:Inflammatory signals must be regulated and kept in check in order to prevent tissue damage. This is especially true in the cornea, where damage can induce loss of transparency, essential for vision. Toll-like receptors (TLRs) are present at the ocular surface and, in addition to being protective against infection, have also been implicated in the pathogenesis of dry eye syndrome, an inflammatory condition that affects millions of individuals in the United States each year. Therefore, an important area of research is the development of new anti-inflammatory therapeutics that limit aberrant ocular surface inflammation. Vitamin D has been studied for its role in suppressing inflammation in other tissues. In previous studies, we have demonstrated that vitamin D is able to decrease proinflammatory mediators induced by TLRs in human corneal epithelial cells (HCEC). Therefore, the goal of the current study was to examine this mechanism further through an evaluation of vitamin Dâs influence on gene expression in two different HCEC cell lines. Microarray comparing hTCEpi, human corneal epithelial cells, treated for 6 hours with Vitamin D (1,25D3) and control (0.1% ethanol) Two-condition arrays: hTCEpi cells were treated with either 100nM 1,25D3 or 0.1% Ethanol for 6h. Two biological replicates, one per array, with a dye swap (technical replicate).
Project description:Inflammatory signals must be regulated and kept in check in order to prevent tissue damage. This is especially true in the cornea, where damage can induce loss of transparency, essential for vision. Toll-like receptors (TLRs) are present at the ocular surface and, in addition to being protective against infection, have also been implicated in the pathogenesis of dry eye syndrome, an inflammatory condition that affects millions of individuals in the United States each year. Therefore, an important area of research is the development of new anti-inflammatory therapeutics that limit aberrant ocular surface inflammation. Vitamin D has been studied for its role in suppressing inflammation in other tissues. In previous studies, we have demonstrated that vitamin D is able to decrease proinflammatory mediators induced by TLRs in human corneal epithelial cells (HCEC). Therefore, the goal of the current study was to examine this mechanism further through an evaluation of vitamin D’s influence on gene expression in two different HCEC cell lines. Microarray comparing SV40-HCEC, human corneal epithelial cells, treated for 6 hours with Vitamin D (1,25D3) and control (0.1% ethanol)
Project description:Inflammatory signals must be regulated and kept in check in order to prevent tissue damage. This is especially true in the cornea, where damage can induce loss of transparency, essential for vision. Toll-like receptors (TLRs) are present at the ocular surface and, in addition to being protective against infection, have also been implicated in the pathogenesis of dry eye syndrome, an inflammatory condition that affects millions of individuals in the United States each year. Therefore, an important area of research is the development of new anti-inflammatory therapeutics that limit aberrant ocular surface inflammation. Vitamin D has been studied for its role in suppressing inflammation in other tissues. In previous studies, we have demonstrated that vitamin D is able to decrease proinflammatory mediators induced by TLRs in human corneal epithelial cells (HCEC). Therefore, the goal of the current study was to examine this mechanism further through an evaluation of vitamin D’s influence on gene expression in two different HCEC cell lines. Microarray comparing hTCEpi, human corneal epithelial cells, treated for 6 hours with Vitamin D (1,25D3) and control (0.1% ethanol)
Project description:MicroRNAs are powerful gene expression regulators, but their corneal repertoire and potential changes in corneal diseases remain unknown. Our purpose was to identify miRNAs altered in the human diabetic cornea by microarray analysis, and to examine their effects on wound healing in cultured telomerase-immortalized human corneal epithelial cells (HCEC) in vitro. Using microarrays, 29 miRNAs were identified as differentially expressed in diabetic samples. Two miRNA candidates showing the highest fold increased in expression in the diabetic cornea were confirmed by Q-PCR and further characterized. HCEC transfection with h-miR-146a or h-miR-424 significantly retarded wound closure, but their respective antagomirs significantly enhanced wound healing vs. controls. Cells treated with h-miR-146a or h-miR-424 had decreased p-p38 and p-EGFR staining, but these increased over control levels close to the wound edge upon antagomir treatment. In conclusion, several miRNAs with increased expression in human diabetic central corneas were found. Two such miRNAs inhibited cultured corneal epithelial cell wound healing. Dysregulation of miRNA expression in human diabetic cornea may be an important mediator of abnormal wound healing.
Project description:Human corneal endothelial cells (HCEC) form a monolayer by adhering tightly through their intercellular adhesion molecules. Located at the posterior corneal surface, they maintain corneal translucency by dehydrating the corneal stroma, mainly through the Na+- and K+-dependent ATPase (Na+/K+-ATPase). Because HCEC proliferative activity is low in vivo,we tried to activate proliferation of HCEC by inhibiting cyclin-dependent kinase inhibitors.We have here demonstrated microarray data of transduced human corneal endothelial cell lines. Affymetrix human U133 plus 2.0 array was used to transcriptionally profile to compare cultured human corneal endothelial cells and transduced human corneal endothelial cells.
Project description:Human corneal endothelial cells (HCEC) form a monolayer by adhering tightly through their intercellular adhesion molecules. Located at the posterior corneal surface, they maintain corneal translucency by dehydrating the corneal stroma, mainly through the Na+- and K+-dependent ATPase (Na+/K+-ATPase). Because HCEC proliferative activity is low in vivo,we tried to activate proliferation of HCEC by inhibiting cyclin-dependent kinase inhibitors.We have here demonstrated microarray data of transduced human corneal endothelial cell lines.
Project description:MicroRNAs are powerful gene expression regulators, but their corneal repertoire and potential changes in corneal diseases remain unknown. Our purpose was to identify miRNAs altered in the human diabetic cornea by microarray analysis, and to examine their effects on wound healing in cultured telomerase-immortalized human corneal epithelial cells (HCEC) in vitro. Using microarrays, 29 miRNAs were identified as differentially expressed in diabetic samples. Two miRNA candidates showing the highest fold increased in expression in the diabetic cornea were confirmed by Q-PCR and further characterized. HCEC transfection with h-miR-146a or h-miR-424 significantly retarded wound closure, but their respective antagomirs significantly enhanced wound healing vs. controls. Cells treated with h-miR-146a or h-miR-424 had decreased p-p38 and p-EGFR staining, but these increased over control levels close to the wound edge upon antagomir treatment. In conclusion, several miRNAs with increased expression in human diabetic central corneas were found. Two such miRNAs inhibited cultured corneal epithelial cell wound healing. Dysregulation of miRNA expression in human diabetic cornea may be an important mediator of abnormal wound healing. Total RNA was extracted from age-matched human autopsy normal (n=6) and diabetic (n=6) central corneas, Flash Tag end-labeled, and hybridized to Affymetrix® GeneChip® miRNA Arrays. Select miRNAs associated with diabetic cornea were validated by quantitative RT-PCR (Q-PCR) and by in situ hybridization (ISH) in independent samples.