Yuanyuan Qian

Blockage of ROS and NF-κB-mediated inflammation by a new chalcone L6H9 protects cardiomyocytes from hyperglycemia-induced injuries

Increased oxidative stress and cardiac inflammation have been implicated in the pathogenesis of diabetic cardiomyopathy (DCM). We previously found that a novel chalcone derivative, L6H9, was able to reduce LPS-induced inflammatory response in macrophages. This study was designed to investigate its protective effects on DCM and the underlying mechanisms. H9C2 cells were cultured with DMEM containing 33 mmol/L of glucose in the presence or absence of L6H9. Pretreatment with L6H9 significantly reduced high glucose-induced inflammatory cytokine expression, ROS level increase, mitochondrial dysfunction, cell apoptosis, fibrosis, and hypertrophy in H9c2 cells, which may be mediated by NF-κB inhibition and Nrf2 activation. In mice with STZ-induced diabetes, oral administration of L6H9 at 20 mg/kg/day for 8 weeks significantly decreased the cardiac cytokine and ROS level, accompanied by decreasing cardiac apoptosis and hypertrophy, and, finally, improved histological abnormalities and fibrosis, without affecting the hyperglycemia. L6H9 also attenuated the diabetes-induced NF-κB activation and Nrf2 decrease in diabetic hearts. These results strongly suggest that L6H9 may have great therapeutic potential in the treatment of DCM via blockage of inflammation and oxidative stress. This study also provides a deeper understanding of the regulatory role of Nrf2 and NF-κB in DCM, indicating that they may be important therapeutic targets for diabetic complications.

EGFR inhibition protects cardiac damage and remodeling through attenuating oxidative stress in STZ-induced diabetic mouse model.

Diabetes mellitus is strongly associated with cardiomyopathy. The underlying mechanisms for the development of diabetic cardiomyopathy are complex and not completely understood. Recent studies showed that epidermal growth factor receptors (EGFRs) are involved in diabetes-induced cardiac injury. However, the role of EGFR in the diabetic heart has yet to be confirmed. The aim of the present study is to further determine the role of EGRF in the pathogenesis of diabetic heart injury. The type 1 diabetic mice induced by streptozotocin were treated with EGFR inhibitors (AG1478 and 451) for 8 weeks, respectively. It was observed that diabetes induced phospohorylation of EGFR and AKT, increased cardiac ROS levels, and ultimately led to cardiac remodeling including cardiac hypertrophy, disorganization, apoptosis, and fibrosis, while all these molecular and pathological alterations were attenuated by the treatment with EGFR inhibitors. In vitro, either pharmacological inhibition of EGFR/AKT or sh-RNA silencing of EGFR significantly inhibited high concentration glucose (HG)-induced ROS generation and subsequently cell apoptosis in both cardiac H9C2 cells and primary rat cardiomyocytes, respectively. The ROS reduction by EGFR inhibitor was associated with the decreased NADPH oxidase activity and expression in H9c2 cells. HG-induced cardiomyocyte injuries were also reduced by NAC, an inhibitor of ROS. This study provides evidence that EGFR has a key role in the pathogenesis of STZ-induced diabetic cardiac damage and remodeling via ROS generation, and suggests that EGFR may be a potential target in treating diabetic cardiomyopathy.

Saturated palmitic acid induces myocardial inflammatory injuries through direct binding to TLR4 accessory protein MD2

Obesity increases the risk for a number of diseases including cardiovascular diseases and type 2 diabetes. Excess saturated fatty acids (SFAs) in obesity play a significant role in cardiovascular diseases by activating innate immunity responses. However, the mechanisms by which SFAs activate the innate immune system are not fully known. Here we report that palmitic acid (PA), the most abundant circulating SFA, induces myocardial inflammatory injury through the Toll-like receptor 4 (TLR4) accessory protein MD2 in mouse and cell culture experimental models. Md2 knockout mice are protected against PA- and high-fat diet-induced myocardial injury. Studies of cell surface binding, cell-free protein–protein interactions and molecular docking simulations indicate that PA directly binds to MD2, supporting a mechanism by which PA activates TLR4 and downstream inflammatory responses. We conclude that PA is a crucial contributor to obesity-associated myocardial injury, which is likely regulated via its direct binding to MD2.

MD2 mediates angiotensin II-induced cardiac inflammation and remodeling via directly binding to Ang II and activating TLR4/NF-κB signaling pathway

Angiotensin II (Ang II) induces cardiac inflammation and remodeling. Emerging evidence indicates that Ang II may utilize the Toll-like receptor 4 (TLR4) signaling pathway in mediating pro-inflammatory and pro-fibrotic activities. However, the precise mechanism is poorly understood. Myeloid differentiation 2 (MD2), a molecule that physically binds to TLR4, confers lipopolysaccharide responsiveness and may also be involved in mediating the actions of Ang II. We hypothesize that MD2 plays an essential role in cardiac inflammation and remodeling induced by local Ang II, and inhibition of MD2 can attenuate Ang II-induced cardiac dysfunction. Using a specific small molecule MD2 blocker L6H21 and the MD2 knockout mice, we show that MD2 deficiency significantly reduces cardiac inflammation and subsequent fibrosis, hypertrophy, and dysfunction in mice challenged with subcutaneous injection of Ang II. In rat cardiomyocyte-like H9c2 cells as well as rat primary cardiomyocytes, inhibition of MD2 by L6H21 or siRNA knockdown suppressed the Ang II-induced TLR4 signaling pathway activation including MyD88 recruitment, and reduced cardiomyocyte hypertrophy and matrix protein expression. These pro-inflammatory activities of Ang II were independent of the AT1 receptor. Finally, we demonstrated the direct interaction between Ang II and MD2 protein via hydrogen bonds on Arg-90, Glu-92, and Asp-100. Ang II produces an inflammatory response and cardiac remodeling by directly binding to MD2, activating MD2/TLR4 complex, and recruiting MyD88. MD2 may be a new therapeutic target for Ang II-mediated cardiac inflammation and remodeling.

Novel EGFR inhibitors attenuate cardiac hypertrophy induced by angiotensin II.

Cardiac hypertrophy is an important risk factor for heart failure. Epidermal growth factor receptor (EGFR) has been found to play a role in the pathogenesis of various cardiovascular diseases. The aim of this current study was to examine the role of EGFR in angiotensin II (Ang II)‐induced cardiac hypertrophy and identify the underlying molecular mechanisms. In this study, we observed that both Ang II and EGF could increase the phospohorylation of EGFR and protein kinase B (AKT)/extracellular signal‐regulated kinase (ERK), and then induce cell hypertrophy in H9c2 cells. Both pharmacological inhibitors and genetic silencing significantly reduced Ang II‐induced EGFR signalling pathway activation, hypertrophic marker overexpression, and cell hypertrophy. In addition, our results showed that Ang II‐induced EGFR activation is mediated by c‐Src phosphorylation. In vivo, Ang II treatment significantly led to cardiac remodelling including cardiac hypertrophy, disorganization and fibrosis, accompanied by the activation of EGFR signalling pathway in the heart tissues, while all these molecular and pathological alterations were attenuated by the oral administration with EGFR inhibitors. In conclusion, the c‐Src‐dependent EGFR activation may play an important role in Ang II‐induced cardiac hypertrophy, and inhibition of EGFR by specific molecules may be an effective strategy for the treatment of Ang II‐associated cardiac diseases.

Novel Epidermal Growth Factor Receptor Inhibitor Attenuates Angiotensin II–Induced Kidney Fibrosis

Chronic activation of renin-angiotensin system (RAS) greatly contributes to renal fibrosis and accelerates the progression of chronic kidney disease; however, the underlying molecular mechanism is poorly understood. Angiotensin II (Ang II), the central component of RAS, is a key regulator of renal fibrogenic destruction. Here we show that epidermal growth factor receptor (EGFR) plays an important role in Ang II–induced renal fibrosis. Inhibition of EGFR activation by novel small molecules or by short hairpin RNA knockdown in Ang II–treated SV40 mesangial cells in vitro suppresses protein kinase B and extracellular signal-related kinase signaling pathways and transforming growth factor-β/Sma- and Mad-related protein activation, and abolishes the accumulation of fibrotic markers such as connective tissue growth factor, collagen IV. The transactivation of EGFR by Ang II in SV40 cells depends on the phosphorylation of proto-oncogene tyrosine-protein kinase Src (c-Src) kinase. Further validation in vivo demonstrates that EGFR small molecule inhibitor successfully attenuates renal fibrosis and kidney dysfunction in a mouse model induced by Ang II infusion. These findings indicate a crucial role of EGFR in Ang II–dependent renal deterioration, and reveal EGFR inhibition as a new therapeutic strategy for preventing progression of chronic renal diseases.

A newly designed curcumin analog Y20 mitigates cardiac injury via anti-inflammatory and anti-oxidant actions in obese rats.

Obesity is strongly associated with the cause of structural and functional changes of the heart in both human and animal models. Oxidative stress and inflammation play a critical role in the development of obesity-induced cardiac disorders. Curcumin is a natural product from Curcuma Longa with multiple bioactivities. In our previous study, in order to reach better anti-inflammatory and anti-oxidant dual activities, we designed a new mono-carbonyl curcumin analog, Y20, via the structural modification with both trifluoromethyl and bromine. This study was designed to investigate the protective effects of Y20 on obesity-induced cardiac injury and its underlying mechanisms. In high fat diet–fed rats, oral administration of Y20 at 20 mg/kg or curcumin at 50 mg/kg significantly decreased the cardiac inflammation and oxidative stress and eventually improved the cardiac remodeling by mitigating cardiac disorganization, hypertrophy, fibrosis and apoptosis. Y20 at 20 mg/kg showed comparable and even stronger bioactivities than curcumin at 50 mg/kg. The beneficial actions of Y20 are closely associated with its ability to increase Nrf2 expression and inhibit NF-κB activation. Taken together, these results suggest that Y20 may have a great therapeutic potential in the treatment of obesity-induced cardiac injury using Nrf2 and NF-κB as the therapeutic targets for treating obesity-related disorders.

Inhibition of inflammation and oxidative stress by an imidazopyridine derivative X22 prevents heart injury from obesity

Inflammation and oxidative stress plays an important role in the development of obesity‐related complications and cardiovascular disease. Benzimidazole and imidazopyridine compounds are a class of compounds with a variety of activities, including anti‐inflammatory, antioxidant and anti‐cancer. X22 is an imidazopyridine derivative we synthesized and evaluated previously for anti‐inflammatory activity in lipopolysaccharide‐stimulated macrophages. However, its ability to alleviate obesity‐induced heart injury via its anti‐inflammatory actions was unclear. This study was designed to evaluate the cardioprotective effects of X22 using cell culture studies and a high‐fat diet rat model. We observed that palmitic acid treatment in cardiac‐derived H9c2 cells induced a significant increase in reactive oxygen species, inflammation, apoptosis, fibrosis and hypertrophy. All of these changes were inhibited by treatment with X22. Furthermore, oral administration of X22 suppressed high‐fat diet‐induced oxidative stress, inflammation, apoptosis, hypertrophy and fibrosis in rat heart tissues and decreased serum lipid concentration. We also found that the anti‐inflammatory and anti‐oxidative actions of X22 were associated with Nrf2 activation and nuclear factor‐kappaB (NF‐κB) inhibition, respectively, both in vitro and in vivo. The results of this study indicate that X22 may be a promising cardioprotective agent and that Nrf2 and NF‐κB may be important therapeutic targets for obesity‐related complications.

New EGFR inhibitor, 453, prevents renal fibrosis in angiotensin II-stimulated mice

Chronic activation of renin-angiotensin system (RAS) greatly contributes to renal fibrosis through the over expression of angiotensin (Ang) II, ultimately leading to chronic kidney disease (CKD). As the main peptide in the RAS, Ang II is a key regulator of nephrotic inflammation, fibrogenic destruction and hypertensive nephropathy. Controlled by growth factors such as TGF-β, Ang II is thought to be affected by other such growth factors including epidermal growth factor (EGF) due to its ability to stimulate growth, regulate angiogenesis, and desensitize cells from apoptotic stimuli. Here we show that epidermal growth factor receptor (EGFR) plays a key role in Ang II induced renal fibrosis and its inhibition for the use as an effective treatment of CKD. 453, an AG1478 analog, was used to block the EGF-EGFR interaction in vivo in 4-week old mice treated with Ang II and 453. Along with the inhibition of EGFR and its downstream signaling pathways (AKT and ERK), 453 also prevented the activation of fibrotic (collagen, CFGF, TGF-β), inflammatory (COX2, IL-6, IL-1β, TNF-α), apoptosis and oxidative stress pathways. These findings suggest the use of 453 as a novel EGFR-inhibitor for therapeutic use in CKD kidney dysfunction.

A novel imidazopyridine derivative, X22, prevents the retinal ischemia-reperfusion injury via inhibition of MAPKs.

Inflammation is a pathological hallmark of ischemia reperfusion (I/R) injury. The present study was conducted to explore the ability of a new anti-inflammatory compound, X22, to attenuate retinal I/R injury via cytokine-inhibitory mechanism. For the in vitro experiment, ARPE-19 cells were pretreated with X22 (5 or 10 μM) or saline for 2 h, followed by stimulation with tert-butyl hydroperoxide (TBHP, 1000 μM) for an indicated amount of time. The expression of inflammatory mediators, cell viability, and cell apoptosis were evaluated. For the in vivo experiment, the rats were randomized to receive treatment with saline or X22 (0.1 μM/kg, 3 μL) before the induction of I/R injury. Histological evaluation, apoptosis of retinal cells, macrophage infiltration, and retina functional changes were further determined. Our data showed that pretreatment with X22 significantly inhibited TBHP-induced inflammatory cytokine expression in ARPE-19 cells. The anti-inflammatory activity of X22 may be associated with its inhibition on MAPKs, rather than NF-κB. Subsequently, our data proved that TBHP induced apoptosis in ARPE-19 cells, while pretreatment of X22 significantly suppressed TBHP-caused ARPE-19 apoptosis. Finally, the in vivo data revealed that X22 administration maintained better inner retinal layer structures, reduced apoptosis of retinal ganglion cell, and improved retinal function in retinal I/R rat models, which were accompanied with a remarkable decrease in retinal macrophage infiltration. These results suggest that the novel compound X22 is a potential agent for the treatment of retinal I/R-related diseases via the MAPKs-targeting anti-inflammatory mechanism and deserves the further development.