Epigallocatechin gallate potentially abrogates fluoride induced lung oxidative stress, inflammation via Nrf2/Keap1 signaling pathway in rats: An in-vivo and in-silico study
Authors: Senthilraja Poomalai
Thangapandiyan Shanmugam
Miltonprabu Selvaraj
Journal: International Immunopharmacology (2016)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: EGCG; Lung; NaF; Nrf2; ROS
Affiliations: Department of Zoology, Annamalai University, Annamalainagar 608002, Tamilnadu, India. Electronic address: ., Department of Zoology, Annamalai University, Annamalainagar 608002, Tamilnadu, India., Divisions of Bioinformatics, Annamalai University, Annamalainagar 608002, Tamilnadu, India.

Abstract:  Background: Since this Nrf2-dependent cellular defense response is able to protect multi-organs, including cancer, neurodegenerative diseases, cardiovascular diseases, inflammation and chronic lung injury. The antioxidant and anti-inflammatory potential of Epigallocatechin gallate (EGCG) and Nrf2/Keap1 signaling mechanisms in pulmonary toxicity have not been clarified. In the present study, we demonstrated that protective efficacy of EGCG against fluoride (Fl) induced oxidative stress mediated lung injury in rats. Methods: The animals were divided in to four groups. Group 1: Control rats received normal saline; Group 2 rats received EGCG (40mg/kg/bw) alone for four weeks; Group 3 rats received Fl (25mg/kg/bw) alone for four weeks, Group 4 rats received EGCG (90min before administration) along with Fl for four weeks. Results: Oral administration of Fl (25mg/kg/bw) significantly (p&lt;0.05) increased the ROS, inflammatory cytokines, lung edema, melonaldehyde (MDA) and myeloperoxidase (MPO) in rats. In addition, upon administration of Fl significantly (p&lt;0.05) decreased the antioxidant status, Nrf2, and HO-1 with increased Keap1 protein. Histological and immunohistochemical (iNOS) study also revealed the Fl induced significant (p&lt;0.05) changes in the lung tissue of rats. Pre-administration of EGCG significantly (p&lt;0.05) improved the antioxidant status, and inhibited the oxidative stress, inflammatory cytokines, and Keap1 protein via the activation of Nrf2 translocation in to the nucleus. Moreover, the molecular docking studies also support the antioxidant potential of EGCG and Nrf2 activation. Conclusion: Taken together, our data indicate that EGCG potentially abrogates Fl induced oxidative lung injury by activation of the Nrf2/Keap1 pathway in rats. 

Retracted: No

PubPeer comments:
 Concern about Figure 7. * Panels A (control rats) and B (EGCG alone treated rats) appear to show a large area of overlap. Shown with red boxes. * Panels C (FI treated rats) and D (pretreatment with EGCG, then FI treated) appear to contain repetitive elements, both within and between the panels. Shown with colored rounded boxes. Could the authors please explain what happened here?  , 
 Fig 5.  . (A) Effect of EGCG on Nrf2, HO1, and Keap1 protein expressions in the lung tissue of control and Fl treated rats by western blot. Lane1.Control, Lane2.Control u00b1 EGCG (40 mg kg BW), Lane3.Fl-control (25 mg kg BW), Lane4.EGCG (40 mg kg BW) u00b1 Fl (25 mg kg BW)  Lane 4 of Nrf2 and Lane 1 of HO1 have more in common than I would expect.  , 
 Dear professor, I have addressed the issue which u have mentioned the immunohistochemistry . I have submitted the new original picture after done the experint. Now again you are giving comment for western blotting. Please understand one thing that in science, if the band intensity coming for same to another may happened, we could not stop it also. No one give the reason why the same band intensity coming for both nrf2 and ho1, what we got the intensity after done the experiment we simply projected out and published. By seeing one by one it is same, same means how could we do the experiment once again. Please under stand this same band intensity not identical and not manipulated also. Its true and i had original data. I have already submitted corrigendum to this issue regarding immunohistochemistry to the editor. If again you are making this issue repeated not fair. Thank u. , 
 #3 To help you understand: the comments made in #1 and #2 are by two different peers, and it is likely that both were unaware that you were addressing the concerns raised in #1. They are not affiliated with the journal or this post publication peer review site, but share a goal of making sure the scientific record is correct. The issues raised in #2 could also be addressed if you are able to provide the scans of the original uncropped blots. I share the concern of the close similarities of the two marked bands posted by u201cHoyau201d in #2. ***  , 
 #3 * Dear professor, I have addressed the issue which u have mentioned the immunohistochemistry . I have submitted the new original picture after done the experint. * Thank you Dr Shanmugam. It sounds like you have contacted the journal and asked them to correct this paper, with a new version of Figure 7, is that correct? Readers might be interested in how Figures 7C and 7D came to exist, and specifically who created them. All the little sections repeated between them make it implausible that they are actual microphotographs of real slices of tissue. , 
 Descriptions of Figure 7 have not done it justice. This is a work of art, where only one panel was needed. There were cuts, asymmetric stretches and some creative erasures. Just follow the  birdie .  , 
 #6  This is a work of art To my mind, too derivative of Dubuffet s [ Hourloupe period] . The  Birdies are of course a *hommage* to Max Ernst s Loplop. Some of the repeated motifs in 7C show formal similarities to Philip Guston s unshaven eye heads. , 
 Perplexingly, a correction to Figure 7 was published: March 2020 The authors regret to inform the readers that this paper included unintended mistakes. In Fig. 7 all the panels (Au2013D) have been replaced with the correct images. In Fig. 7A show no expression of iNOS in the lung tissue which was similar to that of the EGCG alone treated rats (Fig. 7B). A significant increase of iNOS expression was observed in the lung tissue of Fl treated rats (Fig. 7C) when compared to the control rats. Pre-administration of EGCG shows significant reduction in the over expression of iNOS in the lung tissue (Fig. 7D). These changes do not affect the overall results and conclusions published in this paper. The correct Fig. 7Au2013D is presented below:  , 
 Dietary polyphenols are not well absorbed from the gut - see this excerpt from Lambert et. al  In summary, we reported here for the first time the pharmacokinetic parameters of EGCG in the mouse. These results point to the potential role of glucuronidation in affecting EGCG bioavailability and demonstrate important species differences among humans, rats and mice. Further, we confirmed the results of previous studies suggesting that the intestine represents a major barrier to the bioavailability of EGCG and that fecal excretion is the major route of elimination (23). These results provide a basis for future studies of the mechanisms that affect EGCG bioavailability. Further studies of the kinetics of low dose, chronic EGCG, in conjunction with appropriate in vivo markers of a particular mechanism(s), are required to delineate clearly the importance of various in vitro mechanisms in whole animals and in humans. One may also look to Cai, Y.; Anavy, N.D.; Chow, H.H. Contribution of presystemic hepatic extraction to the low oral bioavailability of green tea catechins in rats.Drug Metab. Dispos.2002,30, 1246u20131249 ,