The impact of alterations in the gut microbiome on the immunopathogenesis of diabetic keratopathy

Abstract

Diabetes Mellitus is known to be associated with various ocular disorders, including delayed corneal wound healing. In our study, we aim to investigate the effect of sustained hyperglycemia on corneal epithelial wound healing, the ocular surface and systemic immune response, and microbiome indices in diabetic mice compared to controls after alkaline chemical injury of the eye, and to see if probiotics pre-treatment will ameliorate the attenuated wound healing response due to sustained hyperglycaemia.

Using Akita mice as mice model of type I diabetes, we report that diabetic mice had significantly delayed corneal wound healing compared to controls. This was associated with a reduction in tear levels of VEGF-A, Ang2 and IGF-1 on days 0, 3 and 7 after injury. Furthermore, there was a lack of significant upregulation of peripheral blood and ocular surface CD3+CD4 cell count in response to corneal injury in diabetic mice compared to controls. In response to injury, Akita mice also had a significant reduction in intestinal microbiome diversity indices compared to controls. Individually, at day 7 after injury, Akita mice had significantly lower abundance of Firmicutes bacterium M10-2 compared wild type mice. Furthermore, we found that probiotics pre-treatment promoted corneal epithelial wound healing and had beneficial effect in restoring the altered intestinal microbiome composition as well as attenuated adaptive immune response of the diabetic mice.

Our findings revealed that, in diabetic mice, impaired corneal wound healing was associated with a significant inability to mount a proper systemic and local immune response to ocular chemical injury. Baseline and post-injury differences in intestinal microbial diversity and abundance patterns between diabetic mice and controls may account for this impaired response. Importantly, probiotics treatment has been demonstrated to be a novel and promising therapeutic agent in treating diabetic keratopathy.

However, even without diabetes, corneal chemical burn, in particular alkaline corneal injury, may still result in serious and even permanent defect that significantly impairs visual acuity. Thus, it is meaningful to seek for preventative method that may minimize the hazard on the cornea induced by alkaline burn injury. The second part of our study aims to evaluate the effectiveness of pre-treatment of Lycium barbarum polysaccharide (LBP) in promoting corneal epithelial wound healing after alkaline burn.

After pretreatment of the mouse corneas with either LBP or PBS as vehicle continuously for 7 days, following by induction of the corneal alkaline burn, we have found that, compared to the injury group, mice with LBP pre-treatment revealed a significant decrease in fluorescein-stained area upon injury with increased epithelial layer thickness. The corneal opacity was significantly reduced in the group with LBP pre-treatment followed by injury. The expression of matrix metalloproteinase 12, interleukin- 1 beta, platelet derived growth factor-BB, and aquaporin 5 on the cornea were decreased with LBP pre-treatment.

Our results showed that LBP promoted corneal epithelial growth and minimized disruption of the collagen architecture in vivo. We suggest that LBP, as a natural Traditional Chinese Medicine, may potentially be another novel topical pre-treatment option for patients highly susceptible to chemical injury.