To find out relevant detrimental
To find out relevant detrimental signaling molecules involved in Nrf2 deficiency-induced injury, IL-6 was selected as a target because it has been implicated involving in the Nrf2 signaling pathway in other study . Our results suggested that circulating IL-6 was significantly elevated in Nrf2 KO mice during Ang II infusion, indicating the involvement of IL-6 as well as the interaction between Nrf2 and IL-6 during Ang II-induced hypertrophic cardiomyopathy. Accumulating evidence suggests that the IL-6/STAT3 pathway is critical in myocardial function and cardiac protection [, , ]. We observed that there was no difference of p-STAT3 protein abundance between WT and Nrf2 KO heart at baseline. However, after Ang II stimulation, p-STAT3 was significantly induced in Nrf2 KO mice heart. To further investigate the activation of IL-6/STAT3 signaling pathway in Nrf2 KO heart during Ang II-infusion, we performed CD68 staining of heart tissue from the four groups. The result suggested an elevated presence of macrophages in the Nrf2 KO heart compared to WT during Ang II stimulation, which is consistent with the increased p-STAT3 abundance as well as p-STAT3/STAT3 ratio observed in Nrf2 KO-Ang II hearts.
To investigate whether IL-6/STAT3 signaling is downstream of Nrf2, we also used IL-6 KO mice treated with Ang II infusion or not, and our data showed that STAT3 activation was abrogated in the hearts deficient for IL-6 during Ang II infusion. Additionally, the amount of CD68 positive SB 203580 was much less affected by Ang ll in the heart deficient for IL-6 compared to the heart of WT, which supports our conclusion that Nrf2 negatively regulates IL-6/STAT3 signaling pathway during Ang II-induced hypertrophic cardiomyopathy.
In our study, by using Ang II-induced hypertensive models in vivo and in vitro, we show that Nrf2 is a negative regulator of hypertensive cardiomyopathy. The deficiency of Nrf2 promoted an inflammatory response by activating an important signaling pathway, which involves IL-6 and STAT3 and causes cardiac inflammation. Moreover, Nrf2 deficiency increased Ang II-induced oxidative stress. The above two effects together induce cardiac injury, therefore maladaptation and dysfunction. Our study is the first to report that genetic deletion of Nrf2 contributes to cardiac injury via enhancing the effect of IL-6/STAT3 signaling pathway. Therefore, these findings reveal a potential role of Nrf2 in attenuating agonist-induced cardiomyopathy and its link to IL-6/STAT3 signaling during Ang II-induced cardiac injury.
Funding This work was supported by the National Natural Science Foundation of China (Grant No. 81500179, No. 81573484); Fundació La Marató de TV3 (Grant No. 20153810); the Natural Science Foundation of Jiangsu Province (Grant No. BK20150696); The Open Project of State Key Laboratory of Natural Medicines (Grant No. 3144060108); the National Found for Fostering Talents of Basic Science (NFFTBS) (Grant No. J1310032); the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
Conflicts of interest
Introduction To cope with the enormous range of ambient light levels encountered in daily life, the vertebrate retina utilizes two types of photoreceptors: rods that function in dim light and cones that function in bright light and mediate color vision. Although the signaling pathway for phototransduction is common for rods and cones, the signaling proteins are mostly coded by distinct sets of rod and cone specific genes. The human retina has one type of rod and three types of cone. Rods express the visual pigment rhodopsin, whereas the three types of cones express the short (S, blue), medium (M, green) or long (L, red) wavelength cone opsin. The murine retina has one type of rod and only S- (blue) and M- (green) cones; some cones express both S and M opsins. The zebrafish has one type of rod and four types of cone: L-, M-, S- and UV-cone. The fourth type expresses a cone opsin that is maximally sensitive to UV light [, , , , , ]. During development, the rods, cones and five other major classes of retinal cells are produced from a common pool of pluripotent progenitor cells. The progenitors undergo mitosis and the post-mitotic cells then undergo differentiation into specific types of retinal cells. Differentiation is conceptualized as a process of selective expression of sets of specific genes [, , , , , ]. For rods and cones, several transcription factors that control photoreceptor-specific gene expression have been identified. Cone-rod homeobox transcription factor (Crx) is central to photoreceptor determination [, , ] and required for the development of photoreceptor outer segment (OS) and expression of photoreceptor-specific genes [, , , ]. Crx is expressed both in rod and cone photoreceptors. Mutations in Crx cause photoreceptor degeneration in humans (autosomal dominant cone-rod dystrophy (CRD) [13,18], autosomal dominant retinitis pigmentosa (adRP)  and autosomal recessive Leber congenital amaurosis (LCA) ) and in mice . Neural retina leucine zipper protein (Nrl) is a leucine zipper transcription factor that is required for rod development. Nrl is preferentially expressed in rods. It acts synergistically with Crx to regulate rhodopsin transcription. Mutations in human Nrl cause retinal degeneration: autosomal dominant retinitis pigmentosa (adRP) , autosomal recessive retinitis pigmentosa (arRP)  and enhanced S-cone syndrome (ESCS) . More details about ESCS and RP will be presented later. Deletion of Nrl in mice caused the complete loss of rod function and super-normal S-cone function. In the absence of Nrl, the rods acquired some of the characteristics of S-cones. Accordingly, Nrl seems to act as a molecular switch during rod-cell development by directly modulating rod-specific genes while simultaneously inhibiting the S-cone pathway through the activation of Nr2e3 .