Connexin 40-formed GJIC increases the phototoxicity of photodynamic therapy through ROS- and calcium-mediated pathways / 中国药理学与毒理学杂志

OBJECTIVE To explore the effect of connexin (Cx) 40-formed gap junctional intercellular communication (GJIC) on Photofrin- photodynamic therapy (PDT) phototoxicity in Cx40- transfected HeLa cells and its potential mechanisms. METHODS HeLa cell line stably transfected to express Cx40 was seeded at high and low cell density, respectively, to assess in vitro photosensitivity using CCK8 assay. Western blot assay was performed to detect the expression of Cx40. The intracellular ROS and Ca2 +concentrations were determined using flow cytometer. 4-HNE and ceramide were measured using ELISA assay. RESULTS Cx40-composed GJ formation at high density enhances the phototoxicity of Photofrin-PDT. When the Cx40 is not expressed or Cx40 channels are blocked, the phototoxicity in high-density cultures substantially reduces, indicating that the enhanced PDT phototoxicity at high density is mediated by Cx40-composed GJIC. The GJIC-mediated increase in PDT phototoxicity was associated with ROS and calcium-mediated stress signaling pathways. CONCLUSION The work uniquely presents the ability of Cx40-composed GJIC to enhance the sensitivity of malignant cells to PDT, and indicates that mainte?nance or increase of Cx40-formed GJIC may be a profitable strategy towards the enhancement of PDT therapeutic efficiency.

Type 2 Diabetes Induces Prolonged P-wave Duration without Left Atrial Enlargement

Prolonged P-wave duration has been observed in diabetes. However, the underlying mechanisms remain unclear. The aim of this study was to elucidate the possible mechanisms. A rat model of type 2 diabetes mellitus (T2DM) was used. P-wave durations were obtained using surface electrocardiography and sizes of the left atrium were determined using echocardiography. Cardiac inward rectifier K+ currents (I(k1)), Na+ currents (I(Na)), and action potentials were recorded from isolated left atrial myocytes using patch clamp techniques. Left atrial tissue specimens were analyzed for total connexin-40 (Cx40) and connexin-43 (Cx43) expression levels on western-blots. Specimens were also analyzed for Cx40 and Cx43 distribution and interstitial fibrosis by immunofluorescent and Masson trichrome staining, respectively. The mean P-wave duration was longer in T2DM rats than in controls; however, the mean left atrial sizes of each group of rats were similar. The densities of I(k1) and I(Na) were unchanged in T2DM rats compared to controls. The action potential duration was longer in T2DM rats, but there was no significant difference in resting membrane potential or action potential amplitude compared to controls. The expression level of Cx40 protein was significantly lower, but Cx43 was unaltered in T2DM rats. However, immunofluorescent labeling of Cx43 showed a significantly enhanced lateralization. Staining showed interstitial fibrosis was greater in T2DM atrial tissue. Prolonged P-wave duration is not dependent on the left atrial size in rats with T2DM. Dysregulation of Cx40 and Cx43 protein expression, as well as fibrosis, might partly account for the prolongation of P-wave duration in T2DM.

Antioxidant and anti-atrial fibrillation effects of Guanfu base A / 中国药科大学学报

@#This study aimed at evaluating the antioxidant effects of Guanfu base A(GFA)on acetylcholine(Ach)/CaCl2(CaCl2 10 mg/mL, Ach 66 μg/mL)-induced atrial fibrillation(AF)in rats. SD rats were rando-mized into normal group, model group, GFA treatment groups(6 mg/kg, 12 mg/kg), Amiodarone(Ami)treatment group(50 mg/kg)and Lovastatin(Lov)treatment group(10 mg/kg). The AF durations were measured by electrocardiogram(ECG). The effective refractory periods(AERP)were measured in the left atrial appendage. Oxidative stress-related gene and protein expression was evaluated by RT-PCR and Western blot. The activity of antioxidant enzymes was measured by enzymatic assay. Results indicated that, in comparison with that in the vehicle-treated AF rats, treatment with GFA(6 mg/kg, 12 mg/kg, po), significantly shortened the AF duration and prolonged the AERP in rats. In addition, treatment with GFA reduced the levels of plasma and myocardium malondialdehyde, increased the activity of plasma superoxide dismutase in a dose-dependent manner. Moreover, treatment with GFA mitigated AF-up-regulated p22phox, p47phox, gp91phox, and p67phox NADPH oxidase expression, and AF-increased ratios of membrane to cytosolic Rac-1 in the atrium. It also significantly prevented AF-down-regulated atrial connexin40 expression in rats. Data suggested that GFA(6 mg/kg, 12 mg/kg)has potent anti-oxidant activity and inhibits oxidative-stress-related AF in rats.

Inhibiting activation of ERK1/2 pathway improves atrial fibrosis and connexin40 remodeling in rats / 第二军医大学学报

Objective To investigate the effects of extracellular signal-regulated kinase l/2(ERKl/2) pathway inhibitor U0126 on isopreterenol(ISO)-induced atrial fibrosis and connexin 40 (Cx40) remodeling in rats. Methods Thirty-two male SD rats were evenly randomized into control group, DMSO group, ISO (5 mg/[kg • d])+DMSO group (fibrosis group), and ISO (5 mg/[kg • d]) + U0126 (0. 5 mg/[kg • d]) + DMSO group (U0126-treated group). The corresponding reagents were given to each group once a day and the rats were killed and the myocardial tissues were collected after 7 d. The Ang IJ contents in the myocardial tissues were measured by radioimmunoassay; H-E staining and Masson staining were applied to measure the degree of atrial fibrosis; p-MEKl/2, p-ERKl/2, and Cx40 were detected by immunohistochemistry method. Results (1) The contents of Ang IJ were similar between control group ([242. 133 ± 4. 870] ng/L) and DMSO ([239. 412 ± 1. 795] ng/L) group (P>0. 05). Compared with the above two groups, Ang IJ contents in fibrosis group ([500. 250 ± 8. 869] ng/L)and U0126-treated group([498. 695 ± 9. 340]ng/L) were significantly increased (P alKO. 01). (2) Control group and DMSO group had no atrial fibrosis; the degree of atrial fibrosis in U0126-treated group was significantly lower than that in the fibrosis group (P0. 05), and those in the fibrosis group were significantly increased compared with control group and DMSO group(P0. 05), and was significantly decreased compared with the fibrosis group(P0. 05), and Cx40 was distributed in myocardial cell intercalated disc in a linear manner. The content of Cx40 was significantly reduced (P0. 05)and most of the Cx40 was linearly distributed in myocardial cell intercalated disc. Meanwhile, the reduce degree of Cx40 content in U0126-treated group was significantly decreased than that in the fibrosis group(P<0. 01), and some Cx40 was linearly distributed in myocardial cell intercalated disc. Conclusion Long-term Ang TJ elevation in myocardium may be involved in atrial fibrosis and Cx40 remodeling, and U0126 can efficiently improve atrial fibrosis and Cx40 remodeling by inhibiting the activation of ERK1/2 pathway.