Posters

Acrolein and Hypoxia Induced VEGF Secretion by 661W Cone Photoreceptors

Discipline Track

Biomedical Science

Abstract

Diabetic retinopathy (DR) is a leading cause of blindness in the United States with vascular endothelial growth factor (VEGF) being one of the main hallmarks that exacerbates this condition. In this study, we will determine the effects of acrolein and hypoxia on VEGF secretion under different glucose concentrations in mouse cone photoreceptors (661W) to study the molecular mechanisms of DR. 661W cells were cultured in P100 dishes and then seeded in 6 wells plates at 300k cells/per well. Upon confluency of 80%, cells were subjected to the appropriate treatments consisting of complete culture media with normal (NG; 5.5mM) or high glucose (HG; 30 mM). Hypoxia was induced chemically by using cobalt (ll) chloride hexahydrate solution of 300ug/ml. Acrolein concentrations consisted of low (25uM and 50uM) and high (100uM and 200uM). Elisa was used to measure the VEGF protein in conditioned media. Hypoxia had a significant impact by increasing the amount of VEGF secretion (NG=234 pg./ml, NG + hypoxia=658, HG=246, HG+ hypoxia =630; p=0.002 and 0.0275 respectively) and decrease number of viable cells (NG=2,200,000, NG + hypoxia=762,333, HG=2,230,000, HG+ hypoxia =1,040,000; p=0.003 and 0.037 respectively). Additionally, acrolein played a significant role in decreasing cell viability (p=0.035 in HG with 25uM acrolein=1,470,000, 50uM =760,200, 100uM =95,833, 200uM =76,233 and p=0.0064 in NG with 25uM acrolein=1,500,000, 50uM =1,290,000, 100uM =228,500, 200uM =58,600) and decreased VEGF secretion (p=0.05 in HG with 25uM acrolein=204 pg./ml, 50uM =112, 100uM =27.4, 200uM =18 and p=<0.0001 in NG with 25uM acrolein =272, 50uM =283, 100uM =56.6, 200uM =19.4) in 661W photoreceptor cells. Hypoxia exerted significant effects to reduce 661W cell viability and increase VEGF secretion. However acrolein induced a reduction in cell viability, along with a decrease VEGF secretion. Since hypoxia (low oxygen) and acrolein (a highly reactive aldehyde) are known to affect significantly oxidative pathways, it is possible that their effects may be mediated by the TGFb pathway. Further experiments will be conducted to determine the underlying molecular mechanism.

Presentation Type

Poster

Academic Level

Medical Student

Mentor/PI Department

Molecular Science

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Acrolein and Hypoxia Induced VEGF Secretion by 661W Cone Photoreceptors

Diabetic retinopathy (DR) is a leading cause of blindness in the United States with vascular endothelial growth factor (VEGF) being one of the main hallmarks that exacerbates this condition. In this study, we will determine the effects of acrolein and hypoxia on VEGF secretion under different glucose concentrations in mouse cone photoreceptors (661W) to study the molecular mechanisms of DR. 661W cells were cultured in P100 dishes and then seeded in 6 wells plates at 300k cells/per well. Upon confluency of 80%, cells were subjected to the appropriate treatments consisting of complete culture media with normal (NG; 5.5mM) or high glucose (HG; 30 mM). Hypoxia was induced chemically by using cobalt (ll) chloride hexahydrate solution of 300ug/ml. Acrolein concentrations consisted of low (25uM and 50uM) and high (100uM and 200uM). Elisa was used to measure the VEGF protein in conditioned media. Hypoxia had a significant impact by increasing the amount of VEGF secretion (NG=234 pg./ml, NG + hypoxia=658, HG=246, HG+ hypoxia =630; p=0.002 and 0.0275 respectively) and decrease number of viable cells (NG=2,200,000, NG + hypoxia=762,333, HG=2,230,000, HG+ hypoxia =1,040,000; p=0.003 and 0.037 respectively). Additionally, acrolein played a significant role in decreasing cell viability (p=0.035 in HG with 25uM acrolein=1,470,000, 50uM =760,200, 100uM =95,833, 200uM =76,233 and p=0.0064 in NG with 25uM acrolein=1,500,000, 50uM =1,290,000, 100uM =228,500, 200uM =58,600) and decreased VEGF secretion (p=0.05 in HG with 25uM acrolein=204 pg./ml, 50uM =112, 100uM =27.4, 200uM =18 and p=<0.0001 in NG with 25uM acrolein =272, 50uM =283, 100uM =56.6, 200uM =19.4) in 661W photoreceptor cells. Hypoxia exerted significant effects to reduce 661W cell viability and increase VEGF secretion. However acrolein induced a reduction in cell viability, along with a decrease VEGF secretion. Since hypoxia (low oxygen) and acrolein (a highly reactive aldehyde) are known to affect significantly oxidative pathways, it is possible that their effects may be mediated by the TGFb pathway. Further experiments will be conducted to determine the underlying molecular mechanism.