Glibenclamide alleviates ß adrenergic receptor activation-induced cardiac inflammation.

ß-Adrenergic receptor (ß-AR) overactivation is a major pathological factor associated with cardiac diseases and mediates cardiac inflammatory injury. Glibenclamide has shown anti-inflammatory effects in previous research. However, it is unclear whether and how glibenclamide can alleviate cardiac inflammatory injury induced by ß-AR overactivation. In the present study, male C57BL/6J mice were treated with or without the ß-AR agonist isoprenaline (ISO) with or without glibenclamide pretreatment. The results indicated that glibenclamide alleviated ISO-induced macrophage infiltration in the heart, as determined by Mac-3 staining. Consistent with this finding, glibenclamide also inhibited ISO-induced chemokines and proinflammatory cytokines expression in the heart. Moreover, glibenclamide inhibited ISO-induced cardiac fibrosis and dysfunction in mice. To reveal the protective mechanism of glibenclamide, the NLRP3 inflammasome was further analysed. ISO activated the NLRP3 inflammasome in both cardiomyocytes and mouse hearts, but this effect was alleviated by glibenclamide pretreatment. Furthermore, in cardiomyocytes, ISO increased the efflux of potassium and the generation of ROS, which are recognized as activators of the NLRP3 inflammasome. The ISO-induced increases in these processes were inhibited by glibenclamide pretreatment. Moreover, glibenclamide inhibited the cAMP/PKA signalling pathway, which is downstream of ß-AR, by increasing phosphodiesterase activity in mouse hearts and cardiomyocytes. In conclusion, glibenclamide alleviates ß-AR overactivation-induced cardiac inflammation by inhibiting the NLRP3 inflammasome. The underlying mechanism involves glibenclamide-mediated suppression of potassium efflux and ROS generation by inhibiting the cAMP/PKA pathway.

Membrane nanotubes facilitate the propagation of inflammatory injury in the heart upon overactivation of the ß-adrenergic receptor.

Acute sympathetic stress quickly induces cardiac inflammation and injury, suggesting that pathogenic signals rapidly spread among cardiac cells and that cell-to-cell communication may play an important role in the subsequent cardiac injury. However, the underlying mechanism of this response is unknown. Our previous study demonstrated that acute ß-adrenergic receptor (ß-AR) signaling activates inflammasomes in the heart, which triggers the inflammatory cascade. In the present study, ß-AR overactivation induced inflammasome activation in both the cardiomyocytes and cardiac fibroblasts (CFs) of mice hearts following a subcutaneous injection of isoproterenol (ISO, 5 mg/kg body weight), a selective agonist of ß-AR. In isolated cardiac cells, ISO treatment only activated the inflammasomes in the cardiomyocytes but not the CFs. These results demonstrated that inflammasome activation was propagated from cardiomyocytes to CFs in the mice hearts. Further investigation revealed that the inflammasomes were activated in the cocultured CFs that connected with cardiomyocytes via membrane nanotubes (MNTs), a novel membrane structure that mediates distant intercellular connections and communication. Disruption of the MNTs with the microfilament polymerization inhibitor cytochalasin D (Cyto D) attenuated the inflammasome activation in the cocultured CFs. In addition, the MNT-mediated inflammasome activation in the CFs was blocked by deficiency of the inflammasome component NOD-like receptor protein 3 (NLRP3) in the cardiomyocytes, but not NLRP3 deficiency in the CFs. Moreover, ISO induced pyroptosis in the CFs cocultured with cardiomyocytes, and this process was inhibited by disruption of the MNTs with Cyto D or by the NLRP3 inhibitor MCC950 and the caspase-1 inhibitor Z-YVAD-FMK (FMK). Our study revealed that MNTs facilitate the rapid propagation of inflammasome activation among cardiac cells to promote pyroptosis in the early phase of ß-adrenergic insult. Therefore, preventing inflammasome transfer is a potential therapeutic strategy to alleviate acute ß-AR overactivation-induced cardiac injury.

α1-AR overactivation induces cardiac inflammation through NLRP3 inflammasome activation.

Acute sympathetic stress causes excessive secretion of catecholamines and induces cardiac injuries, which are mainly mediated by ß-adrenergic receptors (ß-ARs). However, α1-adrenergic receptors (α1-ARs) are also expressed in the heart and are activated upon acute sympathetic stress. In the present study, we investigated whether α1-AR activation induced cardiac inflammation and the underlying mechanisms. Male C57BL/6 mice were injected with a single dose of α1-AR agonist phenylephrine (PE, 5 or 10 mg/kg, s.c.) with or without pretreatment with α-AR antagonist prazosin (5 mg/kg, s.c.). PE injection caused cardiac dysfunction and cardiac inflammation, evidenced by the increased expression of inflammatory cytokine IL-6 and chemokines MCP-1 and MCP-5, as well as macrophage infiltration in myocardium. These effects were blocked by prazosin pretreatment. Furthermore, PE injection significantly increased the expression of NOD-like receptor protein 3 (NLRP3) and the cleavage of caspase-1 (p20) and interleukin-18 in the heart; similar results were observed in both Langendorff-perfused hearts and cultured cardiomyocytes following the treatment with PE (10 µM). Moreover, PE-induced NLRP3 inflammasome activation and cardiac inflammation was blocked in Nlrp3-/- mice compared with wild-type mice. In conclusion, α1-AR overactivation induces cardiac inflammation by activating NLRP3 inflammasomes.

[Effects of Ramipril on the expression of connexin 43 in cerebral arteries of spontaneously hypertensive rats].

The present study was designed to examine whether Ramipril (an inhibitor of angiotensin-converting enzyme) affected spontaneous hypertension-induced injury of cerebral artery by regulating connexin 43 (Cx43) expression. Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) were randomly divided into WKY, WKY + Ramipril, SHR, and SHR + Ramipril groups (n = 8). The arterial pressure was monitored by the tail-cuff method, and vascular function in basilar arteries was examined by pressure myography. Hematoxylin-eosin (HE) staining was used to show vascular remodeling. The expression and distribution of Cx43 was determined by using immunofluorescence and immunohistochemistry analysis. The protein and mRNA levels of Cx43 were examined by Western blot and real-time PCR analysis, respectively. The results showed that chronic Ramipril treatment significantly attenuated blood pressure elevation (P < 0.01, n = 8) and blood vessel wall thickness in SHR (P < 0.01, n = 8). The cerebral artery contraction rate in the SHR group was higher than that in the WKY group (P < 0.05, n = 8). The cerebral artery contraction rate in the SHR + Ramipril group was lower than that in the SHR group (P < 0.05, n = 8). Pretreatment with 2-APB (Cx43 non-specific blocker) or Gap26 (Cx43 specific blocker) significantly decreased the vasoconstriction rate, while pretreatment with AAP10 (Cx43 non-specific agonist) significantly increased the vasoconstriction in the SHR + Ramipril group (P < 0.05, n = 8). In addition, the expression of Cx43 mRNA and protein in cerebral arteries of SHR group was higher than that of WKY group (P < 0.05, n = 8). The mRNA and protein expression of Cx43 in cerebral arteries of SHR + Ramipril group was significantly lower than that of SHR group (P < 0.05, n = 8). These results suggest that Ramipril can down-regulate the expression of Cx43 mRNA and protein in cerebral arterial cells of SHR, lower blood pressure, promote vasodilation, and improve arterial damage and vascular dysfunction caused by hypertension.

Longitudinal Study of Scar Hyperplasia Formation Following Cleft Lip Wound Healing.

The purpose of this study was to observe the hyperplasia trend of scar after the cleft lip surgery in a rabbit animal model, and determine the time-point of the highest hypertrophic degree of scar after cleft lip repair. Forty New Zealand white rabbits from the same offspring were used to establish a cleft lip wound healing model using Millard surgery procedure. The scar volumes were measured and granulation tissues were observed visually in the 2, 3, 4, and 5 weeks after operation. The scar tissues were harvested at the indicated time-points. Immunohistochemical (IHC) and Western Blot analyses were performed to detect the expression level of α-smooth muscle actin (α-SMA) in the scar tissue. The scars shrunk and the volumes reduced at 3 to 4 weeks after surgery; however, at 5 weeks postsurgery, the volumes increased. IHC and Western blot analyses indicated the expression of α-SMA was significantly enhanced 3 to 4 weeks, but reduced in the 5 weeks after surgery. Overall, the degree of scar hyperplasia after cleft lip surgery in rabbits was normally distributed and the scarring was most severe in the 3 to 4 weeks after cleft lip surgery. The study confirms a novel animal model for the assessment of therapies for the treatment of scar hyperplasia of human cleft lip in future.