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The effects of insulin pre-administration in mice exposed to ethanol: alleviating hepatic oxidative injury through anti-oxidative, anti-apoptotic activities and deteriorating hepatic steatosis through SRBEP-1c activation.

Liu J, Wang X, Peng Z, Zhang T, Wu H, Yu W, Kong D, Liu Y, Bai H, Liu R, Zhang X, Hai C - Int. J. Biol. Sci. (2015)

Bottom Line: Previous studies have showed that insulin, a classic regulator of glucose metabolism, has significant anti-oxidative function and plays an important role in maintaining the redox balance.First, we found insulin pre-administration alleviated acute ethanol exposure-induced liver injury and inflammation reflected by the decrease of serum AST and ALT activities, the improvement of pathological alteration and the inhibition of TNF-α and IL-6 expressions.Our study provided novel insight about the effects and mechanisms of insulin on ethanol-induced liver injury.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Toxicology, the Ministry of Education Key Lab of Hazard Assessment and Control in Special Operational Environment, Shaanxi Provincial Key Lab of Free radical biology and medicine, School of Public Health, The Fourth Military Medical University, Xi'an, 710032, P. R. China;

ABSTRACT
Alcoholic liver disease (ALD) has become an important liver disease hazard to public and personal health. Oxidative stress is believed to be responsible for the pathological changes in ALD. Previous studies have showed that insulin, a classic regulator of glucose metabolism, has significant anti-oxidative function and plays an important role in maintaining the redox balance. For addressing the effects and mechanisms of insulin pre-administration on ethanol-induced liver oxidative injury, we investigated histopathology, inflammatory factors, apoptosis, mitochondrial dysfunction, oxidative stress, antioxidant defense system, ethanol metabolic enzymes and lipid disorder in liver of ethanol-exposed mice pretreatment with insulin or not. There are several novel findings in our study. First, we found insulin pre-administration alleviated acute ethanol exposure-induced liver injury and inflammation reflected by the decrease of serum AST and ALT activities, the improvement of pathological alteration and the inhibition of TNF-α and IL-6 expressions. Second, insulin pre-administration could significantly reduce apoptosis and ameliorate mitochondrial dysfunction in liver of mice exposed to ethanol, supporting by decreasing caspases-3 activities and the ratio of Bax/Bcl-2, increasing mitochondrial viability and mitochondrial oxygen consumption, inhibition of the decline of ATP levels and mitochondrial ROS accumulation. Third, insulin pre-administration prevented ethanol-mediated oxidative stress and enhance antioxidant defense system, which is evaluated by the decline of MDA levels and the rise of GSH/GSSG, the up-regulations of antioxidant enzymes CAT, SOD, GR through Nrf-2 dependent pathway. Forth, the modification of ethanol metabolism pathway such as the inhibition of CYP2E1, the activation of ALDH might be involved in the anti-oxidative and protective effects exerted by insulin pre-administration against acute ethanol exposure in mice. Finally, insulin pre-administration deteriorated hepatic steatosis in mice exposed to ethanol might be through SRBEP-1c activation. In summary, these results indicated that insulin pre-administration effectively alleviated liver oxidative injury through anti-inflammatory, anti-oxidative and anti-apoptotic activities but also deteriorated hepatic steatosis through SRBEP-1c activation in mice exposed to ethanol. Our study provided novel insight about the effects and mechanisms of insulin on ethanol-induced liver injury.

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Insulin pre-administration ameliorated acute ethanol exposure-induced mitochondrial dysfunction in liver of mice. (A) Mitochondrial viability measured by MTT; (B) Hepatic ATP level; (C) Mitochondrial oxygen consumption in a thermostatically controlled oxygen apparatus; (D) Rate of mitochondrial oxygen consumption expressed as the folds of control; (E) Mitochondrial ROS accumulation analyzed by MitoSOX staining using flow cytometer; (F) Values of mitochondrial ROS accumulation calculated by MitoSOX fluorescence intensity expressed as the folds of control; (G) MMP analyzed by rhodamine 123 staining using flow cytometer; (H) Values of MMP calculated by rhodamine 123 fluorescence intensity expressed as the folds of control. Data of A and B were expressed as mean ± SD (n = 8). Data of D, F and H were expressed as the mean ± SD of three independent experiments (n = 3). *P< 0.05, compared to control group; #P < 0.05, compared to EtOH group.
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Figure 3: Insulin pre-administration ameliorated acute ethanol exposure-induced mitochondrial dysfunction in liver of mice. (A) Mitochondrial viability measured by MTT; (B) Hepatic ATP level; (C) Mitochondrial oxygen consumption in a thermostatically controlled oxygen apparatus; (D) Rate of mitochondrial oxygen consumption expressed as the folds of control; (E) Mitochondrial ROS accumulation analyzed by MitoSOX staining using flow cytometer; (F) Values of mitochondrial ROS accumulation calculated by MitoSOX fluorescence intensity expressed as the folds of control; (G) MMP analyzed by rhodamine 123 staining using flow cytometer; (H) Values of MMP calculated by rhodamine 123 fluorescence intensity expressed as the folds of control. Data of A and B were expressed as mean ± SD (n = 8). Data of D, F and H were expressed as the mean ± SD of three independent experiments (n = 3). *P< 0.05, compared to control group; #P < 0.05, compared to EtOH group.

Mentions: To address whether insulin pre-administration could alleviate ethanol exposure-induced mitochondrial dysfunction in liver, we next investigated the mitochondrial function by measuring mitochondrial viability, ATP level, Rate of mitochondrial oxygen consumption, mitochondrial ROS accumulation and MMP in mice receiving different treatments. As shown in Fig.3A, treatment with ethanol resulted in significant decrease in mitochondrial viability, which was markedly prevented by pre-administration with insulin. As noted in Fig.3B, the liver from ethanol-treated mice showed a marked decrease in ATP level compared with control group, but mice pre-administration with insulin exhibited a higher ATP level compared with the ethanol-treated ones, indicating that insulin improved the energy metabolism in mitochondria. Cellular respiration process occurred in the mitochondria needs oxygen supplement. As illustrated in Fig.3C and D, results using Clark electrode showed that there was remarkable inhibition of mitochondrial oxygen consumption rate in mice receiving ethanol, while insulin pre-administration effectively alleviated the inhibition. As shown in Fig.3E and F, we confirmed that there was significant increase in mitochondrial ROS accumulation in ethanol-treated mice, and insulin pre-administration almost completely abolished the increase. As illustrated in Fig.3G and H, ethanol exposure notably decreased MMP compared with the control group, however, a significantly increase of MMP in liver cells of mice pre-administration with insulin was observed compared to that of ethanol treatment mice. Taking together with the data about apoptosis in Fig.2, above results indicated that the inhibition of mitochondrial dysfunction might be responsible for the anti-apoptotic and protective effects exerted by insulin pre-administration against ethanol toxicity.


The effects of insulin pre-administration in mice exposed to ethanol: alleviating hepatic oxidative injury through anti-oxidative, anti-apoptotic activities and deteriorating hepatic steatosis through SRBEP-1c activation.

Liu J, Wang X, Peng Z, Zhang T, Wu H, Yu W, Kong D, Liu Y, Bai H, Liu R, Zhang X, Hai C - Int. J. Biol. Sci. (2015)

Insulin pre-administration ameliorated acute ethanol exposure-induced mitochondrial dysfunction in liver of mice. (A) Mitochondrial viability measured by MTT; (B) Hepatic ATP level; (C) Mitochondrial oxygen consumption in a thermostatically controlled oxygen apparatus; (D) Rate of mitochondrial oxygen consumption expressed as the folds of control; (E) Mitochondrial ROS accumulation analyzed by MitoSOX staining using flow cytometer; (F) Values of mitochondrial ROS accumulation calculated by MitoSOX fluorescence intensity expressed as the folds of control; (G) MMP analyzed by rhodamine 123 staining using flow cytometer; (H) Values of MMP calculated by rhodamine 123 fluorescence intensity expressed as the folds of control. Data of A and B were expressed as mean ± SD (n = 8). Data of D, F and H were expressed as the mean ± SD of three independent experiments (n = 3). *P< 0.05, compared to control group; #P < 0.05, compared to EtOH group.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4400388&req=5

Figure 3: Insulin pre-administration ameliorated acute ethanol exposure-induced mitochondrial dysfunction in liver of mice. (A) Mitochondrial viability measured by MTT; (B) Hepatic ATP level; (C) Mitochondrial oxygen consumption in a thermostatically controlled oxygen apparatus; (D) Rate of mitochondrial oxygen consumption expressed as the folds of control; (E) Mitochondrial ROS accumulation analyzed by MitoSOX staining using flow cytometer; (F) Values of mitochondrial ROS accumulation calculated by MitoSOX fluorescence intensity expressed as the folds of control; (G) MMP analyzed by rhodamine 123 staining using flow cytometer; (H) Values of MMP calculated by rhodamine 123 fluorescence intensity expressed as the folds of control. Data of A and B were expressed as mean ± SD (n = 8). Data of D, F and H were expressed as the mean ± SD of three independent experiments (n = 3). *P< 0.05, compared to control group; #P < 0.05, compared to EtOH group.
Mentions: To address whether insulin pre-administration could alleviate ethanol exposure-induced mitochondrial dysfunction in liver, we next investigated the mitochondrial function by measuring mitochondrial viability, ATP level, Rate of mitochondrial oxygen consumption, mitochondrial ROS accumulation and MMP in mice receiving different treatments. As shown in Fig.3A, treatment with ethanol resulted in significant decrease in mitochondrial viability, which was markedly prevented by pre-administration with insulin. As noted in Fig.3B, the liver from ethanol-treated mice showed a marked decrease in ATP level compared with control group, but mice pre-administration with insulin exhibited a higher ATP level compared with the ethanol-treated ones, indicating that insulin improved the energy metabolism in mitochondria. Cellular respiration process occurred in the mitochondria needs oxygen supplement. As illustrated in Fig.3C and D, results using Clark electrode showed that there was remarkable inhibition of mitochondrial oxygen consumption rate in mice receiving ethanol, while insulin pre-administration effectively alleviated the inhibition. As shown in Fig.3E and F, we confirmed that there was significant increase in mitochondrial ROS accumulation in ethanol-treated mice, and insulin pre-administration almost completely abolished the increase. As illustrated in Fig.3G and H, ethanol exposure notably decreased MMP compared with the control group, however, a significantly increase of MMP in liver cells of mice pre-administration with insulin was observed compared to that of ethanol treatment mice. Taking together with the data about apoptosis in Fig.2, above results indicated that the inhibition of mitochondrial dysfunction might be responsible for the anti-apoptotic and protective effects exerted by insulin pre-administration against ethanol toxicity.

Bottom Line: Previous studies have showed that insulin, a classic regulator of glucose metabolism, has significant anti-oxidative function and plays an important role in maintaining the redox balance.First, we found insulin pre-administration alleviated acute ethanol exposure-induced liver injury and inflammation reflected by the decrease of serum AST and ALT activities, the improvement of pathological alteration and the inhibition of TNF-α and IL-6 expressions.Our study provided novel insight about the effects and mechanisms of insulin on ethanol-induced liver injury.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Toxicology, the Ministry of Education Key Lab of Hazard Assessment and Control in Special Operational Environment, Shaanxi Provincial Key Lab of Free radical biology and medicine, School of Public Health, The Fourth Military Medical University, Xi'an, 710032, P. R. China;

ABSTRACT
Alcoholic liver disease (ALD) has become an important liver disease hazard to public and personal health. Oxidative stress is believed to be responsible for the pathological changes in ALD. Previous studies have showed that insulin, a classic regulator of glucose metabolism, has significant anti-oxidative function and plays an important role in maintaining the redox balance. For addressing the effects and mechanisms of insulin pre-administration on ethanol-induced liver oxidative injury, we investigated histopathology, inflammatory factors, apoptosis, mitochondrial dysfunction, oxidative stress, antioxidant defense system, ethanol metabolic enzymes and lipid disorder in liver of ethanol-exposed mice pretreatment with insulin or not. There are several novel findings in our study. First, we found insulin pre-administration alleviated acute ethanol exposure-induced liver injury and inflammation reflected by the decrease of serum AST and ALT activities, the improvement of pathological alteration and the inhibition of TNF-α and IL-6 expressions. Second, insulin pre-administration could significantly reduce apoptosis and ameliorate mitochondrial dysfunction in liver of mice exposed to ethanol, supporting by decreasing caspases-3 activities and the ratio of Bax/Bcl-2, increasing mitochondrial viability and mitochondrial oxygen consumption, inhibition of the decline of ATP levels and mitochondrial ROS accumulation. Third, insulin pre-administration prevented ethanol-mediated oxidative stress and enhance antioxidant defense system, which is evaluated by the decline of MDA levels and the rise of GSH/GSSG, the up-regulations of antioxidant enzymes CAT, SOD, GR through Nrf-2 dependent pathway. Forth, the modification of ethanol metabolism pathway such as the inhibition of CYP2E1, the activation of ALDH might be involved in the anti-oxidative and protective effects exerted by insulin pre-administration against acute ethanol exposure in mice. Finally, insulin pre-administration deteriorated hepatic steatosis in mice exposed to ethanol might be through SRBEP-1c activation. In summary, these results indicated that insulin pre-administration effectively alleviated liver oxidative injury through anti-inflammatory, anti-oxidative and anti-apoptotic activities but also deteriorated hepatic steatosis through SRBEP-1c activation in mice exposed to ethanol. Our study provided novel insight about the effects and mechanisms of insulin on ethanol-induced liver injury.

Show MeSH
Related in: MedlinePlus