Proceedings for Annual Meeting of The Japanese Pharmacological Society
Background: Atrial fibrillation (AF) is the most common arrhythmia. AF is highly correlated with ... more Background: Atrial fibrillation (AF) is the most common arrhythmia. AF is highly correlated with multiple risk factors including heart failure, age, obesity, and type 2 diabetes. Among risk factors, the incidence in obesity is increasing worldwide. Recently, it was reported that SGLT2 inhibitors reduced the incidence of atrial fibrillation. However, it is unclear how the treatment with SGLT2 inhibitors has effects on vulnerability to AF. In this study, we examined the effects on the inducibility and duration of AF by treatment with SGLT2 inhibitors in diet-induced obese mice. Methods: Mice were fed a normal chow diet (NCD) or high-fat diet (HFD). Following diet-loading, we randomly divided the animals into groups: NCD+vehicle, HFD+vehicle, and HFD+ SGLT2 treatments. Induction of AF was performed by transesophageal atrial burst pacing. Furthermore, we evaluated cardiac function, blood pressure, atrial fibrosis, and glucose tolerance at the end of the treatments. Results: The results showed that HFD-fed mice increased the inducibility of AF compared to NCD mice. In addition, treatment with the SGLT2 inhibitor in HFD-fed mice dose-dependently reduced the inducibility and duration of AF. There were no significant differences in cardiac function, blood pressure, and fibrosis among all groups. Impairment of glucose tolerance in HFD-induced obesity was improved by treatment with the SGLT2 inhibitor. Conclusion: Treatment with the SGLT2 inhibitor reduced the inducibility of AF and shortened the duration of AF without affecting atrial structural remodeling, suggesting that the SGLT2 inhibitor effectively prevents AF in obesity.
Proceedings for Annual Meeting of The Japanese Pharmacological Society
Background: Doxorubicin (Dox), an anticancer drug, is known to induce cardiac toxicity by causing... more Background: Doxorubicin (Dox), an anticancer drug, is known to induce cardiac toxicity by causing mitochondrial dysfunction. Although CaMKII and its phosphorylation targets, Drp1 to control mitochondrial fission and MCU to control mitochondrial Ca 2 + uptake, regulate mitochondrial homeostasis, the involvement of these molecules in the Dox-induced mitochondrial dysfunction remains unclear. Method: To study the effects of Dox on mitochondrial homeostasis, we evaluated mitochondrial membrane potential (MMP), mitophagy, and mitochondrial Ca 2 + content ([Ca 2 +]m) in H9C2 cells with the following fluorescent dyes, JC-1, Mtphagy, and Rhod2-AM, respectively. To examine the activating effect of Dox on CaMKII, we evaluated the phosphorylation levels of CaMKII by western blotting. To test the involvement of CaMKII, Drp1, and MCU in the Dox-induced mitochondrial dysfunction, the specific inhibitors, KN-93, Mdivi-1, and Ru360, respectively, were used. Result: Dox treatment dose-dependently reduced MMP and increased the number of cells with mitophagy and [Ca 2 +] m (p<0.05 in all). Dox treatment significantly increased the phosphorylation levels of CaMKII (p<0.05). The inhibition of CaMKII suppressed the effects of Dox on the MMP and the mitophagy (p<0.05), but not on [Ca2+]m. Contrarily, the inhibition of Drp1 and MCU failed to suppress the decrease in MMP by Dox. Similarly, the inhibition of Drp1 did not reverse the increase in mitophagy by Dox, nor did the inhibition of MCU suppress the elevation of [Ca 2 +]m by Dox. Conclusion: These results suggested that activated CaMKII, but not Drp1 and MCU, is involved in the impairment of MMP leading to Dox-induced mitochondrial dysfunction and that the excessive fission by Drp1 and the increased uptake [Ca 2 +]m by MCU are not the mechanism for the Dox-induced MMP reduction.
Proceedings for Annual Meeting of The Japanese Pharmacological Society
Objective Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstei... more Objective Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstein-Barr virus (EBV) persists in human B cells and occasionally reactivates. During EBV reactivation, the host B cells differentiate to be plasma cells and produce IgM-dominant antibodies. We have previously observed that TRAb-IgM disruptes thyroid follicular epithelial cells and does not transduce thyroid hormone-producing signals. However, it is still unclear how it works on receptor-binding of TSH. We aimed to investigate this. Methods TRAb-IgM were separated from sera or EBV-reactivated culture media of peripheral blood mononuclear cells from Graves' disease patients. TSH binding-inhibitory activity of TRAb-IgM was assessed by a commercial radio-receptor assay kit. Results & Discussion All TRAb-IgM samples showed gamma-ray counts that were almost twice that of the 0 standard. This meant that two molecules of 125 I-TSH bound to one TRAb-IgM binding complex because TRAb-IgM kept a TSH receptor for the separation procedure. This result indicated that TRAb-IgM bound to almost all TSH receptors coated in the test tubes, and did not inhibit TSH binding to TSH receptors. Although thyroid-stimulating TRAb is IgG type, TRAb-IgM may has particular role on Graves' disease. Conclusions TRAb-IgM did not inhibit TSH binding to the TSH receptor. TRAb-IgM does not function as an antagonist.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2022
Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstein-Barr vir... more Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstein-Barr virus (EBV) persists in human B cells, and occasionally reactivates. We have reported that both Graves' disease patients and healthy controls have EBV-infected B cells that have TRAb as their surface globulin (TRAb(+) EBV(+) cells). The peripheral blood mononuclear cells containing TRAb(+) EBV(+) cells produced TRAbs along with EBV reactivation. We proposed the EBV reactivation-induced Ig production as alternative system of Ig production. The antibodies produced by EBV reactivation-induced system are IgM dominant, and skewed to be autoreactive. However, the class of thyroid stimulating TRAb is known to be IgG. We studied about the role of the IgM antibody. We purified TRAb-IgM from culture medium of TRAb(+) EBV(+) cells. Then, we cultured porcine thyroid cells with the TRAb-IgM and complements, and then, measured cAMP and LDH levels to estimate thyroid stimulating effect and cell injury, respectively. We observed the increase of the levels of LDH, but could not detect cAMP. We considered that TRAb-IgM did not have thyroid stimulating effect, but it could injure the thyroid cells and release thyroid antigens including TSH receptor antigen. The relevance of EBV reactivation to Graves' disease may have a possibility for the new therapy.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2021
Objective: Skeletal muscle fiber conversion bears a part in multiple myopathies, and one of those... more Objective: Skeletal muscle fiber conversion bears a part in multiple myopathies, and one of those is Sarcopenia which is known to correlate with aging, nutritional deficiency, and so on. In this study, we focused on muscle specific enriched microRNAs (myomirs) to understand the mechanisms of muscle fiber conversion. Method & Results: In-silico study, we selected the microRNA-133a-3p (miR-133a), a myomir relevant to the skeletal myogenesis. Using the human iPS cell-derived skeletal myogenesis system, we identified that expression level of miR-133a was elevated with a peak at day 5 after the induction of myogenesis. Overexpression of miR-133a mimic or inhibitor at the beginning of myogenesis increased mRNA expression of Myh7, a specific marker of slow oxidative myotube, or Myh1, a marker of fast-type myotube, respectively. Furthermore, we found that, under the serumdeprived culture condition, overexpression of miR-133a mimic preserved the cell feature of oxidative fibers, although miR-133a inhibition deformed the tubular formation of oxidative myotubes. Conclusion: MicroRNA-133a functioned to buttress the gene expression of oxidative fiber-specified myosin heavy chain, and to preserve myotubes from nutritional stress, suggesting the possibility that miR-133a would be a therapeutic target for sarcopenia.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2020
It has been known that the increase of visceral fat is closely involved in abnormal glucose toler... more It has been known that the increase of visceral fat is closely involved in abnormal glucose tolerance, hypertension, and hyperlipidemia, leading to the other diseases, such as atherosclerosis and type 2 diabetes. Recent reports have shown that serum copper concentration was increased in obese and/or diabetic patients. Here, we investigated the effects of copper chelator, cuprizone on high fat diet (HFD)-induced obesity in mice. We administered cuprizone (0.2%w) mixed in food pellets. Mice were divided in 4 groups, fed with (1) normal chow (NCD), (2) NCD with cuprizone (NCD+C), (3) HFD or (4) HFD with cuprizone (HFD+C) for 4 weeks, and then metabolic parameters were obtained. Serum copper levels were decreased in both cuprizone groups. Cuprizone significantly decreased the body weight in HFD+C without changes of food intake. Specifically in HFD+C, cuprizone decreased 60% of epididymal and inguinal fat weights, but did not change the weight of skeletal muscle, both soleus and gastrocnemius. Furthermore, we found that cuprizone ameliorated HFD-induced insulin resistance (ipGTT and ITT) with the modifications of gene expression pattern in liver, based on the comparison between NCD, HFD and HFD+C. These results suggest that cuprizone would be a candidate of leading compounds for anti-obesity agent.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2020
Background: Obesity is a risk factor for atrial fibrillation (AF). However, the mechanisms underl... more Background: Obesity is a risk factor for atrial fibrillation (AF). However, the mechanisms underlying AF in obesity remain unclear. In this study, we established the diet-induced obese-rat model with high AF inducibility and evaluated the relationship between AF inducibility and cardiac function. Methods: Male Sprague-Dawley rats were fed with normal chow diet + normal drinking water (NCD) or high-fat diet + 30% fructose in drinking water (HFFr) for 12 weeks. These rats were subjected to hemodynamic measurements, echocardiography to assess the evaluation of the cardiac structure and function., and transesophageal burst atrial pacing for the induction of AF. Results: HFFr-fed rats were divided into 2 groups: obese and non-obese HFFr-fed rats. Compared with NCD-fed rats, the inducibility of AF significantly increased in obese HFFr-fed rats, but not in non-obese HFFr-fed rats. On echocardiography, LVEF (an indicator of LV systolic function), and E/A ratio (a marker of LV diastolic function) didn't change among these rats. For hemodynamic measurements, LVSP, dP/dtmax, and heart rate increased in obese HFFr-fed rats. Conclusion: We established the obese rat model with high AF inducibility while maintaining normal cardiac function. This model would be useful to elucidate the mechanisms of obesity-related AF.
Proceedings for Annual Meeting of The Japanese Pharmacological Society
Background: Atrial fibrillation (AF) is the most common arrhythmia. AF is highly correlated with ... more Background: Atrial fibrillation (AF) is the most common arrhythmia. AF is highly correlated with multiple risk factors including heart failure, age, obesity, and type 2 diabetes. Among risk factors, the incidence in obesity is increasing worldwide. Recently, it was reported that SGLT2 inhibitors reduced the incidence of atrial fibrillation. However, it is unclear how the treatment with SGLT2 inhibitors has effects on vulnerability to AF. In this study, we examined the effects on the inducibility and duration of AF by treatment with SGLT2 inhibitors in diet-induced obese mice. Methods: Mice were fed a normal chow diet (NCD) or high-fat diet (HFD). Following diet-loading, we randomly divided the animals into groups: NCD+vehicle, HFD+vehicle, and HFD+ SGLT2 treatments. Induction of AF was performed by transesophageal atrial burst pacing. Furthermore, we evaluated cardiac function, blood pressure, atrial fibrosis, and glucose tolerance at the end of the treatments. Results: The results showed that HFD-fed mice increased the inducibility of AF compared to NCD mice. In addition, treatment with the SGLT2 inhibitor in HFD-fed mice dose-dependently reduced the inducibility and duration of AF. There were no significant differences in cardiac function, blood pressure, and fibrosis among all groups. Impairment of glucose tolerance in HFD-induced obesity was improved by treatment with the SGLT2 inhibitor. Conclusion: Treatment with the SGLT2 inhibitor reduced the inducibility of AF and shortened the duration of AF without affecting atrial structural remodeling, suggesting that the SGLT2 inhibitor effectively prevents AF in obesity.
Proceedings for Annual Meeting of The Japanese Pharmacological Society
Background: Doxorubicin (Dox), an anticancer drug, is known to induce cardiac toxicity by causing... more Background: Doxorubicin (Dox), an anticancer drug, is known to induce cardiac toxicity by causing mitochondrial dysfunction. Although CaMKII and its phosphorylation targets, Drp1 to control mitochondrial fission and MCU to control mitochondrial Ca 2 + uptake, regulate mitochondrial homeostasis, the involvement of these molecules in the Dox-induced mitochondrial dysfunction remains unclear. Method: To study the effects of Dox on mitochondrial homeostasis, we evaluated mitochondrial membrane potential (MMP), mitophagy, and mitochondrial Ca 2 + content ([Ca 2 +]m) in H9C2 cells with the following fluorescent dyes, JC-1, Mtphagy, and Rhod2-AM, respectively. To examine the activating effect of Dox on CaMKII, we evaluated the phosphorylation levels of CaMKII by western blotting. To test the involvement of CaMKII, Drp1, and MCU in the Dox-induced mitochondrial dysfunction, the specific inhibitors, KN-93, Mdivi-1, and Ru360, respectively, were used. Result: Dox treatment dose-dependently reduced MMP and increased the number of cells with mitophagy and [Ca 2 +] m (p<0.05 in all). Dox treatment significantly increased the phosphorylation levels of CaMKII (p<0.05). The inhibition of CaMKII suppressed the effects of Dox on the MMP and the mitophagy (p<0.05), but not on [Ca2+]m. Contrarily, the inhibition of Drp1 and MCU failed to suppress the decrease in MMP by Dox. Similarly, the inhibition of Drp1 did not reverse the increase in mitophagy by Dox, nor did the inhibition of MCU suppress the elevation of [Ca 2 +]m by Dox. Conclusion: These results suggested that activated CaMKII, but not Drp1 and MCU, is involved in the impairment of MMP leading to Dox-induced mitochondrial dysfunction and that the excessive fission by Drp1 and the increased uptake [Ca 2 +]m by MCU are not the mechanism for the Dox-induced MMP reduction.
Proceedings for Annual Meeting of The Japanese Pharmacological Society
Objective Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstei... more Objective Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstein-Barr virus (EBV) persists in human B cells and occasionally reactivates. During EBV reactivation, the host B cells differentiate to be plasma cells and produce IgM-dominant antibodies. We have previously observed that TRAb-IgM disruptes thyroid follicular epithelial cells and does not transduce thyroid hormone-producing signals. However, it is still unclear how it works on receptor-binding of TSH. We aimed to investigate this. Methods TRAb-IgM were separated from sera or EBV-reactivated culture media of peripheral blood mononuclear cells from Graves' disease patients. TSH binding-inhibitory activity of TRAb-IgM was assessed by a commercial radio-receptor assay kit. Results & Discussion All TRAb-IgM samples showed gamma-ray counts that were almost twice that of the 0 standard. This meant that two molecules of 125 I-TSH bound to one TRAb-IgM binding complex because TRAb-IgM kept a TSH receptor for the separation procedure. This result indicated that TRAb-IgM bound to almost all TSH receptors coated in the test tubes, and did not inhibit TSH binding to TSH receptors. Although thyroid-stimulating TRAb is IgG type, TRAb-IgM may has particular role on Graves' disease. Conclusions TRAb-IgM did not inhibit TSH binding to the TSH receptor. TRAb-IgM does not function as an antagonist.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2022
Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstein-Barr vir... more Thyrotropin receptor antibody (TRAb) is a causative antibody of Graves' disease. Epstein-Barr virus (EBV) persists in human B cells, and occasionally reactivates. We have reported that both Graves' disease patients and healthy controls have EBV-infected B cells that have TRAb as their surface globulin (TRAb(+) EBV(+) cells). The peripheral blood mononuclear cells containing TRAb(+) EBV(+) cells produced TRAbs along with EBV reactivation. We proposed the EBV reactivation-induced Ig production as alternative system of Ig production. The antibodies produced by EBV reactivation-induced system are IgM dominant, and skewed to be autoreactive. However, the class of thyroid stimulating TRAb is known to be IgG. We studied about the role of the IgM antibody. We purified TRAb-IgM from culture medium of TRAb(+) EBV(+) cells. Then, we cultured porcine thyroid cells with the TRAb-IgM and complements, and then, measured cAMP and LDH levels to estimate thyroid stimulating effect and cell injury, respectively. We observed the increase of the levels of LDH, but could not detect cAMP. We considered that TRAb-IgM did not have thyroid stimulating effect, but it could injure the thyroid cells and release thyroid antigens including TSH receptor antigen. The relevance of EBV reactivation to Graves' disease may have a possibility for the new therapy.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2021
Objective: Skeletal muscle fiber conversion bears a part in multiple myopathies, and one of those... more Objective: Skeletal muscle fiber conversion bears a part in multiple myopathies, and one of those is Sarcopenia which is known to correlate with aging, nutritional deficiency, and so on. In this study, we focused on muscle specific enriched microRNAs (myomirs) to understand the mechanisms of muscle fiber conversion. Method & Results: In-silico study, we selected the microRNA-133a-3p (miR-133a), a myomir relevant to the skeletal myogenesis. Using the human iPS cell-derived skeletal myogenesis system, we identified that expression level of miR-133a was elevated with a peak at day 5 after the induction of myogenesis. Overexpression of miR-133a mimic or inhibitor at the beginning of myogenesis increased mRNA expression of Myh7, a specific marker of slow oxidative myotube, or Myh1, a marker of fast-type myotube, respectively. Furthermore, we found that, under the serumdeprived culture condition, overexpression of miR-133a mimic preserved the cell feature of oxidative fibers, although miR-133a inhibition deformed the tubular formation of oxidative myotubes. Conclusion: MicroRNA-133a functioned to buttress the gene expression of oxidative fiber-specified myosin heavy chain, and to preserve myotubes from nutritional stress, suggesting the possibility that miR-133a would be a therapeutic target for sarcopenia.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2020
It has been known that the increase of visceral fat is closely involved in abnormal glucose toler... more It has been known that the increase of visceral fat is closely involved in abnormal glucose tolerance, hypertension, and hyperlipidemia, leading to the other diseases, such as atherosclerosis and type 2 diabetes. Recent reports have shown that serum copper concentration was increased in obese and/or diabetic patients. Here, we investigated the effects of copper chelator, cuprizone on high fat diet (HFD)-induced obesity in mice. We administered cuprizone (0.2%w) mixed in food pellets. Mice were divided in 4 groups, fed with (1) normal chow (NCD), (2) NCD with cuprizone (NCD+C), (3) HFD or (4) HFD with cuprizone (HFD+C) for 4 weeks, and then metabolic parameters were obtained. Serum copper levels were decreased in both cuprizone groups. Cuprizone significantly decreased the body weight in HFD+C without changes of food intake. Specifically in HFD+C, cuprizone decreased 60% of epididymal and inguinal fat weights, but did not change the weight of skeletal muscle, both soleus and gastrocnemius. Furthermore, we found that cuprizone ameliorated HFD-induced insulin resistance (ipGTT and ITT) with the modifications of gene expression pattern in liver, based on the comparison between NCD, HFD and HFD+C. These results suggest that cuprizone would be a candidate of leading compounds for anti-obesity agent.
Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2020
Background: Obesity is a risk factor for atrial fibrillation (AF). However, the mechanisms underl... more Background: Obesity is a risk factor for atrial fibrillation (AF). However, the mechanisms underlying AF in obesity remain unclear. In this study, we established the diet-induced obese-rat model with high AF inducibility and evaluated the relationship between AF inducibility and cardiac function. Methods: Male Sprague-Dawley rats were fed with normal chow diet + normal drinking water (NCD) or high-fat diet + 30% fructose in drinking water (HFFr) for 12 weeks. These rats were subjected to hemodynamic measurements, echocardiography to assess the evaluation of the cardiac structure and function., and transesophageal burst atrial pacing for the induction of AF. Results: HFFr-fed rats were divided into 2 groups: obese and non-obese HFFr-fed rats. Compared with NCD-fed rats, the inducibility of AF significantly increased in obese HFFr-fed rats, but not in non-obese HFFr-fed rats. On echocardiography, LVEF (an indicator of LV systolic function), and E/A ratio (a marker of LV diastolic function) didn't change among these rats. For hemodynamic measurements, LVSP, dP/dtmax, and heart rate increased in obese HFFr-fed rats. Conclusion: We established the obese rat model with high AF inducibility while maintaining normal cardiac function. This model would be useful to elucidate the mechanisms of obesity-related AF.
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Papers by Agung Priyono