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Brian Leonard, Theint Aung, Melinda Quan, Allison Shannon, VijayKrishna Raghunathan, Sara M Thomasy, Christopher J Murphy; Dose dependent induction and kinetics of human beta-defensin 3 peptide expression after treatment with herbal compounds. Invest. Ophthalmol. Vis. Sci. 2021;62(8):916.
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© ARVO (1962-2015); The Authors (2016-present)
Antimicrobial peptides (AMPs), including defensins, are key effector molecules of innate immunity expressed and secreted by corneal epithelial cells. Previous studies from our lab demonstrate a marked upregulation of human beta-defensin 3 mRNA (DEFB103) expression in corneal epithelial cells treated with herbal compounds andrographolide, oridonin, isoliquiritigenin. The current study focused on quantification of human beta-defensin 3 peptide (hBD3) after induction with the herbal compounds and determination of the induction kinetics.
hTCEpi cells were treated with increasing concentrations of andrographolide (10-100 μM), oridonin (1-15 μM), isoliquiritigenin (5-50 μM) or negative control (vehicle: DMSO or EtOH) for 48 hours. After treatment, the supernatant was collected and 100 μl was used to quantify hBD3 peptide concentration and performed in duplicate for each treatment. To determine kinetics of DEFB103 mRNA upregulation, hTCEpi cells were treated with andrographolide (75 μM), oridonin (10 μM), or isoliquiritigenin (25 μM) for 2, 6, 12, and 24 hours prior to mRNA isolation and comparted with vehicle control. Statistical significance was determined based on an ANOVA and appropriate post-hoc test.
The level of hBD3 peptide was below the threshold of detection when hTCEpi cells were treated with vehicle, 10, 25 and 50 μM of andrographolide, yet resulted in detectable levels of hBD3 peptide of 260 pg/ml and 488 pg/ml with 75 and 100 mM, respectively. Additionally, hBD3 peptide levels were below the level of detection in supernatants from hTCEpi cells treated with oridonin and isoliquiritigenin at all doses. Both isoliquiritigenin and oridonin had an initial upregulation of DEFB103 mRNA expression within 6 hours, peaking by 12 hours and returning to baseline levels by 24 hours post-treatment. Andrographolide had an initial upregulation of DEFB103 mRNA expression by 6 hours that continued to increase at 24 hours post-treatment.
Modulation of AMP expression with herbal compounds (andrographolide, oridonin, isoliquiritigenin) represent a novel approach to treating patients with resistant bacterial infections of the ocular surface. The kinetics of DEFB103 induction with each compound provides insight as to the frequency of application that may be required to sustain antimicrobial activity.
This is a 2021 ARVO Annual Meeting abstract.
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