Quercetin Attenuates Lactate Production and Extracellular Matrix Secretion in Keratoconus

Abstract

Keratoconus(KC) is an ecstatic corneal disease leading to corneal-thinning and the formation of a cone-like cornea. Elevated lactate levels, increased oxidative stress, and myofibroblast formation have all been previously reported. In the current study, we assess the role of Quercetin on collagen secretion and myofibroblast formation in KC in vitro. Human corneal fibroblasts(HCFs) and human keratoconus cells(HKCs) were treated with a stable Vitamin C derivative and cultured for 4 weeks, stimulating formation of a self-assembled extracellular matrix. All samples were analyzed using Western blots and targeted tandem mass spectrometry. Our data showed that Quercetin significantly down regulates myofibroblast differentiation and fibrotic markers, such as α-smooth muscle actin (α-SMA) and Collagen III (Col III), in both HCFs and HKCs. Collagen III secretion was reduced 80% in both HCFs and HKCs following Quercetin treatment. Furthermore, Quercetin reduced lactate production by HKCs to normal HCF levels. Quercetin down regulated TGF-βR2 and TGF-β2 expression in HKCs suggesting a significant link to the TGF-β pathway. These results assert that Quercetin is a key regulator of fibrotic markers and ECM assembly by modulating cellular metabolism and TGF-β signaling. Our study suggests that Quercetin is a potential therapeutic for treatment of corneal dystrophies, such as KC.

Caveolins and caveolae in ocular physiology and pathophysiology

“…Quercetin also suppresses oxidant-induced expression of Cav-1 in RPE cells and protects them from oxidative stress (Kook et al, 2008). Intriguingly, quercetin has also recently been shown to reduce fibrotic changes in human corneal fibroblasts, in vitro (McKay et al, 2015). Whether the anti-inflammatory, anti-oxidant, and anti-fibrotic properties of polyphenolic compounds are mediated through suppression of caveolin expression remains to be determined.…”

Quercetin modulates keratoconus metabolism in vitro

“…In the retina, quercetin was shown to protect retinal pigment epithelium (RPE) from hydrogen peroxide‐induced cell death and reverse oxidative damage in the RPE . In the cornea, we have recently shown that quercetin inhibits the fibrotic phenotype exhibited by HKCs in vitro with a decrease in α‐SMA and Collagen III expression . Herein, we explored the role of quercetin in modulating cellular metabolism in HKCs as a possible mechanism of action..…”

“…Second, in HCFs, the TCA cycle showed downregulation of citrate while HKCs showed upregulation of this metabolite as well as isocitrate and malate, indicating a faster and rather overloading TCA cycling. These results become even more significant when we consider that quercetin significantly downregulates fibrotic markers in these same cell types and the same 3D culture model, as shown by our group recently . This suggests that the metabolic mechanism for corneal fibrosis differs between specific conditions and diseases.…”

“…Studies have shown that flavonoids, including quercetin, play a protective role against UV‐light damage in plants, whereby their biosynthesis is altered dependent on cellular stress by exposure to UV‐light . We have recently reported quercetin as a key regulator of corneal fibrosis and ECM assembly in KC . As a natural progression to our recent findings, this study establishes the metabolic activity of quercetin in both HCFs and HKCs.…”

Acute hypoxia influences collagen and matrix metalloproteinase expression by human keratoconus cells in vitro

“…Cell lysis was performed at the end of week 4 (post-1 week in hypoxia) using 1X RIPA buffer containing protease inhibitors, as previously described [25]. Briefly, constructs were isolated and washed 1X with PBS.…”