Tongxinluo exerts protective effects via anti-apoptotic and pro-autophagic mechanisms by activating AMPK pathway in infarcted rat hearts.

What is the central question of this study? In a rat model of acute myocardial infarction (AMI), we investigated the effect of Tongxinluo (TXL) treatment. Does TXL activate autophagy and attenuate apoptosis of cardiomyocytes through the AMPK pathway to facilitate survival of cardiomyocytes and improve cardiac function? What is the main finding and its importance? Major findings are as follows: (i) TXL treatment preserved cardiac function and reduced ventricular remodelling, infarct size and inflammation in rat hearts after AMI; (ii) TXL treatment dramatically increased autophagy and inhibited apoptosis in myocardium; and (iii) the AMPK signalling pathway played a crucial role in mediating the beneficial effects of TXL. Tongxinluo (TXL) has been demonstrated to have a protective role during ischaemia-reperfusion after acute myocardial infarction, but the long-term effects and underlying mechanisms are still unknown. The aim of this study was to investigate whether TXL could have an effect on apoptosis or autophagy of cardiomyocytes through the AMP-activated protein kinase (AMPK) pathway. Male Sprague-Dawley rats (n = 75) were randomly divided to sham, control, TXL (4 mg kg-1  day-1 orally), compound C (i.p. injection of 10 mg kg-1  day-1 ) and TXL + compound C groups. The extent of fibrosis, infarct size and angiogenesis were determined by pathological and histological studies. Four weeks after acute myocardial infarction, TXL treatment significantly increased ejection fraction, promoted angiogenesis in the peri-infarct region and substantially decreased fibrosis and the size of the infarcted area (P < 0.05). Treatment with TXL also increased AMPK/mTOR phosphorylation, upregulated expression of the autophagic protein LC3 and downregulated expression of the apoptotic protein Bax in the infarcted myocardium (P < 0.05). Addition of the AMPK inhibitor, compound C, counteracted these beneficial effects significantly (P < 0.05). The cardioprotective benefits of TXL against myocardial infarction are related to the inhibition of apoptosis and promotion of autophagy in rat hearts after acute myocardial infarction. This effect may occur through the AMPK signalling pathway.

Tongxinluo modulates cytokine secretion by cardiac microvascular endothelial cells in ischemia/reperfusion injury.

Cardiac microvascular endothelial cells (CMECs) extensively secrete cytokines during myocardial ischemia/reperfusion injury (MIRI). Tongxinluo (TXL) has been demonstrated to preserve the function of the endothelium and myocardium against MIRI. This study was designed to identify alterations in the paracrine function of CMECs under hypoxia/reoxygenation (H/R) conditions and assess its modulation by TXL. CMECs were exposed to different concentrations of TXL for 30 min and then subjected to hypoxia and reoxygenation for 12 and 2 h, respectively. Apoptosis was measured to determine the optimal TXL concentration. Protein antibody arrays were used to assess changes in cytokines secreted into conditioned medium by CMECs. A Gene Ontology (GO) analysis was applied to interpret the functional implications of changes in cytokines. TXL inhibited CMEC apoptosis in a concentration-dependent manner after H/R, reaching peak efficacy at a concentration of 800 μg/ml. H/R significantly altered 33 cytokines, and TXL (800 μg/ml) changed the levels of 121 different cytokines compared with the H/R group. Among these cytokines, 10 that were increased by H/R were decreased by TXL, five that were decreased by H/R were increased by TXL, and eight that were attenuated by H/R were further decreased by TXL. Insulin-like growth factor binding protein-1 was up-regulated by H/R and was further increased by TXL. Significantly altered factors were found to be involved in cell proliferation, growth and differentiation, as well as chemotaxis and transport. TXL inhibited the apoptosis of CMECs and modulated their paracrine function in MIRI.

The clinical, angiographic and prognosis characteristics of elderly patients with acute ST-segment elevation myocardial infarction--the first elderly STEMI population study in northwest of China.

Association of green tea consumption with risk of coronary heart disease in Chinese population.

Intravenous administration of atorvastatin-pretreated mesenchymal stem cells improves cardiac performance after acute myocardial infarction: role of CXCR4.

BACKGROUND:The interaction between stromal cell-derived factor 1 (SDF-1) and its receptor CXC chemokine receptor 4 (CXCR4) plays an important role in mesenchymal stem cells (MSCs) migration and engraftment. Statins can increase the survival of MSCs. However, whether statins could enhance MSCs migration and engraftment is still unknown. Therefore, we designed the study to investigate whether atorvastatin (ATV) could enhance CXCR4 expression of MSCs and promote them homing toward the injured myocardium.METHODS AND RESULTS:Expression of CXCR4 was evaluated by flow cytometry and real time PCR. A transwell system was used to assess MSCs migration ability. Recruitment of systematically delivered MSCs to the infarcted heart was evaluated in Sprague-Dawley rats with acute myocardial infarction (AMI). ATV pretreatment enhanced the expression of CXCR4 and stimulated MSCs migration in vitro. However, the effect was largely abolished by CXCR4 neutralizing antibody. In AMI models, we found much more ATV-pretreated MSCs homing toward the infarcted myocardium than non-treated cells and this was accompanied by improved cardiac performance.CONCLUSIONS:ATV increases the migration ability of MSCs and improves cardiac performance due to up-regulated expression of CXCR4. These results suggest that ATV pretreatment of donor MSCs is an effective way to promote cell therapeutic potential for AMI.

Atorvastatin induces autophagy of mesenchymal stem cells under hypoxia and serum deprivation conditions by activating the mitogen-activated protein kinase/extracellular signal-regulated kinase pathway.

BACKGROUND:The survival ratio of implanted mesenchymal stem cells (MSCs) in the infarcted myocardium is low. Autophagy is a complex "self-eating" process and can be utilized for cell survival. We have found that atorvastatin (ATV) can effectively activate autophagy to enhance MSCs survival during hypoxia and serum deprivation (H/SD). The mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK/ERK) pathway is a non-canonical autophagy pathway. We hypothesized that the MEK/ERK pathway mediated ATV-induced autophagy of MSCs under H/SD.METHODS:MSCs were pretreated with ATV (0.01-10 µmol/L) under H/SD for three hours. For inhibitor studies, the cells were pre-incubated with the MEK1/2 inhibitor U0126. Cell autophagy was assessed by acidic vesicular organelles (AVO)-positive cells using flow cytometry, autophagy related protein using Western blotting and autophagosome using transmission electron microscopy.RESULTS:Autophagy was elevated in the H/SD group compared with the normal group. ATV further enhanced the autophagic activity as well as the phosphorylation of ERK1/2 evidenced by more AVO-positive cells ((8.63 ± 0.63)% vs. (5.77 ± 0.44)%, P < 0.05), higher LC3-II/LC3-I ratio (4.36 ± 0.31 vs. 2.52 ± 0.18, P < 0.05) and more autophagosomes. And treatment with U0126 downregulated the phosphorylation of ERK1/2 and attenuated ATV-induced autophagy.CONCLUSION:The MEK/ERK pathway participates in ATV-induced autophagy in MSCs under H/SD, and modulation of the pathway could be a novel strategy to improve MSCs survival.

The efficacy and safety of transradial percutaneous coronary intervention vs transfemoral percutaneous coronary intervention for ST-segment elevation myocardial infarction patients: a meta-analysis.

Atorvastatin treatment improves the effects of mesenchymal stem cell transplantation on acute myocardial infarction: the role of the RhoA/ROCK/ERK pathway.

BACKGROUND:Statins protect mesenchymal stem cells (MSCs) against the harsh microenvironment and improve the efficacy of MSC transplantation after acute myocardial infarction (AMI); however, the mechanism remains uncertain. Furthermore, the transdifferentiation potential of MSCs in the post-infarct heart remains highly controversial. The RhoA/Rho-associated coiled-coil-forming kinase (ROCK) pathway participates in many aspects of the damaged heart after AMI and related to the "pleiotropic" effects of statins. This study aimed to explore whether atorvastatin (ATV) facilitates the survival and therapeutic efficacy of MSCs via the inhibition of RhoA/ROCK pathway and subsequently its downstream molecular extracellular regulated protein kinase (ERK1/2), and to investigate the transdifferentiation potential of MSCs in vivo.METHODS AND RESULTS:Female rats received myocardial injections of male rat MSCs 30 min after AMI. Four weeks after AMI, ATV combined with MSC treatment resulted in improved cardiac function and reduced infarct area. ATV facilitated the MSC survival, as revealed by the increased expression of Y chromosomal genes and the increased number of Y chromosome-positive cells; however, no transdifferentiation markers were observed. ATV inhibited the production of inflammatory cytokines both in vitro and vivo, accompanied by suppression of ROCK and ERK activities. Geranylgeranyl pyrophosphate (GGPP) abrogated the effects of ATV in the H9c2 cells under hypoxia/serum deprivation (H/SD), while the ROCK inhibitor fasudil mimicked the benefits of ATV after AMI.CONCLUSIONS:ATV improves the post-infarct microenvironment via RhoA/ROCK/ERK inhibition and thus facilitates the survival and efficacy of implanted MSCs. Transdifferentiation may be not responsible for the cardiac benefits that follow MSC transplantation.

Mipomersen is a promising therapy in the management of hypercholesterolemia: a meta-analysis of randomized controlled trials.

INTRODUCTION:By inhibiting apolipoprotein B (ApoB) synthesis, mipomersen can significantly reduce ApoB-containing lipoproteins in hypercholesterolemic patients.OBJECTIVE:This study sought to ascertain both the extent to which mipomersen can decrease ApoB-containing lipoproteins and the safety of mipomersen therapy.METHODS:Studies were identified through PubMed, CENTRAL, Embase, Clinical Trials, reviews, and reference lists of relevant papers. The efficacy endpoints were the changes in low-density lipoprotein cholesterol (LDL-C), non-high-density lipoprotein cholesterol (non-HDL-C), ApoB, and lipoprotein (a) [Lp(a)]. The safety endpoints were the incidence of injection-site reactions, flu-like symptoms, and elevated transaminases.RESULTS:Six randomized controlled trials with 444 patients were included in the analysis. Compared with the placebo group, patients who received mipomersen therapy had a significant reduction in LDL-C (33.13%), as well as a reduction in non-HDL-C (31.70%), ApoB (33.27%), and LP(a) (26.34%). Mipomersen therapy was also associated with an obvious increase in injection-site reactions with an odds ratio (OR) of 14.15, flu-like symptoms with an OR of 2.07, and alanine aminotransferase levels ≥ 3 × the upper limit of normal with an OR of 11.21.CONCLUSIONS:Mipomersen therapy is effective for lowering ApoB-containing lipoproteins in patients with severe hypercholesterolemia. Future studies exploring how to minimize side effects of mipomersen therapy are needed.

Tongxinluo decreases apoptosis of mesenchymal stem cells concentration-dependently under hypoxia and serum deprivation conditions through the AMPK/eNOS pathway.

Tongxinluo (TXL), a traditional Chinese medicine, is widely used to treat cardiovascular diseases in China. Our previous study has demonstrated the pro-survival role of TXL on mesenchymal stem cells (MSCs) in vivo. But whether TXL could decrease apoptosis of MSCs in vitro, and the underlying mechanism are still unknown. Moreover, AMPK/eNOS pathway is crucial in regulating cell apoptosis. Therefore, we designed the study to investigate whether TXL could decrease MSCs apoptosis under hypoxia and serum deprivation (H/SD) conditions and to determine the role of AMPK/eNOS pathway. To test the hypothesis, MSCs were treated with TXL (50-400 μg/mL) under H/SD for 6 hours. For inhibitor studies, the cells were preincubated with AMPK inhibitor compound C. Results indicated that TXL decreased MSCs apoptosis concentration-dependently evidenced by reduced Annexin V+/PI- cells and increased red/green ratio of JC-1. Further, TXL enhanced the phosphorylation of AMPK and eNOS. Whereas, treatment with compound C decreased the phosphorylation of AMPK and eNOS and was accompanied by attenuated anti-apoptotic effect of TXL. In conclusion, TXL protected MSCs against H/SD-induced injury at least in part through the AMPK/eNOS pathway, which provides a novel explanation for the multi-effect of TXL on cardiovascular system.

Tongxinluo exerts protective effects via anti-apoptotic and pro-autophagic mechanisms by activating AMPK pathway in infarcted rat hearts.

What is the central question of this study? In a rat model of acute myocardial infarction (AMI), we investigated the effect of Tongxinluo (TXL) treatment. Does TXL activate autophagy and attenuate apoptosis of cardiomyocytes through the AMPK pathway to facilitate survival of cardiomyocytes and improve cardiac function? What is the main finding and its importance? Major findings are as follows: (i) TXL treatment preserved cardiac function and reduced ventricular remodelling, infarct size and inflammation in rat hearts after AMI; (ii) TXL treatment dramatically increased autophagy and inhibited apoptosis in myocardium; and (iii) the AMPK signalling pathway played a crucial role in mediating the beneficial effects of TXL. Tongxinluo (TXL) has been demonstrated to have a protective role during ischaemia-reperfusion after acute myocardial infarction, but the long-term effects and underlying mechanisms are still unknown. The aim of this study was to investigate whether TXL could have an effect on apoptosis or autophagy of cardiomyocytes through the AMP-activated protein kinase (AMPK) pathway. Male Sprague-Dawley rats (n = 75) were randomly divided to sham, control, TXL (4 mg kg-1  day-1 orally), compound C (i.p. injection of 10 mg kg-1  day-1 ) and TXL + compound C groups. The extent of fibrosis, infarct size and angiogenesis were determined by pathological and histological studies. Four weeks after acute myocardial infarction, TXL treatment significantly increased ejection fraction, promoted angiogenesis in the peri-infarct region and substantially decreased fibrosis and the size of the infarcted area (P < 0.05). Treatment with TXL also increased AMPK/mTOR phosphorylation, upregulated expression of the autophagic protein LC3 and downregulated expression of the apoptotic protein Bax in the infarcted myocardium (P < 0.05). Addition of the AMPK inhibitor, compound C, counteracted these beneficial effects significantly (P < 0.05). The cardioprotective benefits of TXL against myocardial infarction are related to the inhibition of apoptosis and promotion of autophagy in rat hearts after acute myocardial infarction. This effect may occur through the AMPK signalling pathway.

Tongxinluo modulates cytokine secretion by cardiac microvascular endothelial cells in ischemia/reperfusion injury.

Cardiac microvascular endothelial cells (CMECs) extensively secrete cytokines during myocardial ischemia/reperfusion injury (MIRI). Tongxinluo (TXL) has been demonstrated to preserve the function of the endothelium and myocardium against MIRI. This study was designed to identify alterations in the paracrine function of CMECs under hypoxia/reoxygenation (H/R) conditions and assess its modulation by TXL. CMECs were exposed to different concentrations of TXL for 30 min and then subjected to hypoxia and reoxygenation for 12 and 2 h, respectively. Apoptosis was measured to determine the optimal TXL concentration. Protein antibody arrays were used to assess changes in cytokines secreted into conditioned medium by CMECs. A Gene Ontology (GO) analysis was applied to interpret the functional implications of changes in cytokines. TXL inhibited CMEC apoptosis in a concentration-dependent manner after H/R, reaching peak efficacy at a concentration of 800 μg/ml. H/R significantly altered 33 cytokines, and TXL (800 μg/ml) changed the levels of 121 different cytokines compared with the H/R group. Among these cytokines, 10 that were increased by H/R were decreased by TXL, five that were decreased by H/R were increased by TXL, and eight that were attenuated by H/R were further decreased by TXL. Insulin-like growth factor binding protein-1 was up-regulated by H/R and was further increased by TXL. Significantly altered factors were found to be involved in cell proliferation, growth and differentiation, as well as chemotaxis and transport. TXL inhibited the apoptosis of CMECs and modulated their paracrine function in MIRI.

The clinical, angiographic and prognosis characteristics of elderly patients with acute ST-segment elevation myocardial infarction--the first elderly STEMI population study in northwest of China.

Association of green tea consumption with risk of coronary heart disease in Chinese population.

Intravenous administration of atorvastatin-pretreated mesenchymal stem cells improves cardiac performance after acute myocardial infarction: role of CXCR4.

BACKGROUND:The interaction between stromal cell-derived factor 1 (SDF-1) and its receptor CXC chemokine receptor 4 (CXCR4) plays an important role in mesenchymal stem cells (MSCs) migration and engraftment. Statins can increase the survival of MSCs. However, whether statins could enhance MSCs migration and engraftment is still unknown. Therefore, we designed the study to investigate whether atorvastatin (ATV) could enhance CXCR4 expression of MSCs and promote them homing toward the injured myocardium.METHODS AND RESULTS:Expression of CXCR4 was evaluated by flow cytometry and real time PCR. A transwell system was used to assess MSCs migration ability. Recruitment of systematically delivered MSCs to the infarcted heart was evaluated in Sprague-Dawley rats with acute myocardial infarction (AMI). ATV pretreatment enhanced the expression of CXCR4 and stimulated MSCs migration in vitro. However, the effect was largely abolished by CXCR4 neutralizing antibody. In AMI models, we found much more ATV-pretreated MSCs homing toward the infarcted myocardium than non-treated cells and this was accompanied by improved cardiac performance.CONCLUSIONS:ATV increases the migration ability of MSCs and improves cardiac performance due to up-regulated expression of CXCR4. These results suggest that ATV pretreatment of donor MSCs is an effective way to promote cell therapeutic potential for AMI.

Atorvastatin induces autophagy of mesenchymal stem cells under hypoxia and serum deprivation conditions by activating the mitogen-activated protein kinase/extracellular signal-regulated kinase pathway.

BACKGROUND:The survival ratio of implanted mesenchymal stem cells (MSCs) in the infarcted myocardium is low. Autophagy is a complex "self-eating" process and can be utilized for cell survival. We have found that atorvastatin (ATV) can effectively activate autophagy to enhance MSCs survival during hypoxia and serum deprivation (H/SD). The mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK/ERK) pathway is a non-canonical autophagy pathway. We hypothesized that the MEK/ERK pathway mediated ATV-induced autophagy of MSCs under H/SD.METHODS:MSCs were pretreated with ATV (0.01-10 µmol/L) under H/SD for three hours. For inhibitor studies, the cells were pre-incubated with the MEK1/2 inhibitor U0126. Cell autophagy was assessed by acidic vesicular organelles (AVO)-positive cells using flow cytometry, autophagy related protein using Western blotting and autophagosome using transmission electron microscopy.RESULTS:Autophagy was elevated in the H/SD group compared with the normal group. ATV further enhanced the autophagic activity as well as the phosphorylation of ERK1/2 evidenced by more AVO-positive cells ((8.63 ± 0.63)% vs. (5.77 ± 0.44)%, P < 0.05), higher LC3-II/LC3-I ratio (4.36 ± 0.31 vs. 2.52 ± 0.18, P < 0.05) and more autophagosomes. And treatment with U0126 downregulated the phosphorylation of ERK1/2 and attenuated ATV-induced autophagy.CONCLUSION:The MEK/ERK pathway participates in ATV-induced autophagy in MSCs under H/SD, and modulation of the pathway could be a novel strategy to improve MSCs survival.

The efficacy and safety of transradial percutaneous coronary intervention vs transfemoral percutaneous coronary intervention for ST-segment elevation myocardial infarction patients: a meta-analysis.

Atorvastatin treatment improves the effects of mesenchymal stem cell transplantation on acute myocardial infarction: the role of the RhoA/ROCK/ERK pathway.

BACKGROUND:Statins protect mesenchymal stem cells (MSCs) against the harsh microenvironment and improve the efficacy of MSC transplantation after acute myocardial infarction (AMI); however, the mechanism remains uncertain. Furthermore, the transdifferentiation potential of MSCs in the post-infarct heart remains highly controversial. The RhoA/Rho-associated coiled-coil-forming kinase (ROCK) pathway participates in many aspects of the damaged heart after AMI and related to the "pleiotropic" effects of statins. This study aimed to explore whether atorvastatin (ATV) facilitates the survival and therapeutic efficacy of MSCs via the inhibition of RhoA/ROCK pathway and subsequently its downstream molecular extracellular regulated protein kinase (ERK1/2), and to investigate the transdifferentiation potential of MSCs in vivo.METHODS AND RESULTS:Female rats received myocardial injections of male rat MSCs 30 min after AMI. Four weeks after AMI, ATV combined with MSC treatment resulted in improved cardiac function and reduced infarct area. ATV facilitated the MSC survival, as revealed by the increased expression of Y chromosomal genes and the increased number of Y chromosome-positive cells; however, no transdifferentiation markers were observed. ATV inhibited the production of inflammatory cytokines both in vitro and vivo, accompanied by suppression of ROCK and ERK activities. Geranylgeranyl pyrophosphate (GGPP) abrogated the effects of ATV in the H9c2 cells under hypoxia/serum deprivation (H/SD), while the ROCK inhibitor fasudil mimicked the benefits of ATV after AMI.CONCLUSIONS:ATV improves the post-infarct microenvironment via RhoA/ROCK/ERK inhibition and thus facilitates the survival and efficacy of implanted MSCs. Transdifferentiation may be not responsible for the cardiac benefits that follow MSC transplantation.

Mipomersen is a promising therapy in the management of hypercholesterolemia: a meta-analysis of randomized controlled trials.

INTRODUCTION:By inhibiting apolipoprotein B (ApoB) synthesis, mipomersen can significantly reduce ApoB-containing lipoproteins in hypercholesterolemic patients.OBJECTIVE:This study sought to ascertain both the extent to which mipomersen can decrease ApoB-containing lipoproteins and the safety of mipomersen therapy.METHODS:Studies were identified through PubMed, CENTRAL, Embase, Clinical Trials, reviews, and reference lists of relevant papers. The efficacy endpoints were the changes in low-density lipoprotein cholesterol (LDL-C), non-high-density lipoprotein cholesterol (non-HDL-C), ApoB, and lipoprotein (a) [Lp(a)]. The safety endpoints were the incidence of injection-site reactions, flu-like symptoms, and elevated transaminases.RESULTS:Six randomized controlled trials with 444 patients were included in the analysis. Compared with the placebo group, patients who received mipomersen therapy had a significant reduction in LDL-C (33.13%), as well as a reduction in non-HDL-C (31.70%), ApoB (33.27%), and LP(a) (26.34%). Mipomersen therapy was also associated with an obvious increase in injection-site reactions with an odds ratio (OR) of 14.15, flu-like symptoms with an OR of 2.07, and alanine aminotransferase levels ≥ 3 × the upper limit of normal with an OR of 11.21.CONCLUSIONS:Mipomersen therapy is effective for lowering ApoB-containing lipoproteins in patients with severe hypercholesterolemia. Future studies exploring how to minimize side effects of mipomersen therapy are needed.

Tongxinluo decreases apoptosis of mesenchymal stem cells concentration-dependently under hypoxia and serum deprivation conditions through the AMPK/eNOS pathway.

Tongxinluo (TXL), a traditional Chinese medicine, is widely used to treat cardiovascular diseases in China. Our previous study has demonstrated the pro-survival role of TXL on mesenchymal stem cells (MSCs) in vivo. But whether TXL could decrease apoptosis of MSCs in vitro, and the underlying mechanism are still unknown. Moreover, AMPK/eNOS pathway is crucial in regulating cell apoptosis. Therefore, we designed the study to investigate whether TXL could decrease MSCs apoptosis under hypoxia and serum deprivation (H/SD) conditions and to determine the role of AMPK/eNOS pathway. To test the hypothesis, MSCs were treated with TXL (50-400 μg/mL) under H/SD for 6 hours. For inhibitor studies, the cells were preincubated with AMPK inhibitor compound C. Results indicated that TXL decreased MSCs apoptosis concentration-dependently evidenced by reduced Annexin V+/PI- cells and increased red/green ratio of JC-1. Further, TXL enhanced the phosphorylation of AMPK and eNOS. Whereas, treatment with compound C decreased the phosphorylation of AMPK and eNOS and was accompanied by attenuated anti-apoptotic effect of TXL. In conclusion, TXL protected MSCs against H/SD-induced injury at least in part through the AMPK/eNOS pathway, which provides a novel explanation for the multi-effect of TXL on cardiovascular system.