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Apoptosis of bone marrow mesenchymal stem cells caused by homocysteine via activating JNK signal.

Cai B, Li X, Wang Y, Liu Y, Yang F, Chen H, Yin K, Tan X, Zhu J, Pan Z, Wang B, Lu Y - PLoS ONE (2013)

Bottom Line: Apoptosis of BMSCs in response to various pathological stimuli leads to the attenuation of healing ability of BMSCs.Furthermore, apoptotic appearance and mitochondrial membrane potential depolarization in homocysteine-treated BMSCs was significantly reversed by JNK inhibitor but not p38 MAPK and ERK inhibitors.Homocysteine treatment caused a significant reduction of BMSCs-secreted VEGF and IGF-1 in the culture medium.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Harbin Medical University, Harbin, Heilongjiang Province, China.

ABSTRACT
Bone marrow mesenchymal stem cells (BMSCs) are capable of homing to and repair damaged myocardial tissues. Apoptosis of BMSCs in response to various pathological stimuli leads to the attenuation of healing ability of BMSCs. Plenty of evidence has shown that elevated homocysteine level is a novel independent risk factor of cardiovascular diseases. The present study was aimed to investigate whether homocysteine may induce apoptosis of BMSCs and its underlying mechanisms. Here we uncovered that homocysteine significantly inhibited the cellular viability of BMSCs. Furthermore, TUNEL, AO/EB, Hoechst 333342 and Live/Death staining demonstrated the apoptotic morphological appearance of BMSCs after homocysteine treatment. A distinct increase of ROS level was also observed in homocysteine-treated BMSCs. The blockage of ROS by DMTU and NAC prevented the apoptosis of BMSCs induced by homocysteine, indicating ROS was involved in the apoptosis of BMSCs. Moreover, homocysteine also caused the depolarization of mitochondrial membrane potential of BMSCs. Furthermore, apoptotic appearance and mitochondrial membrane potential depolarization in homocysteine-treated BMSCs was significantly reversed by JNK inhibitor but not p38 MAPK and ERK inhibitors. Western blot also confirmed that p-JNK was significantly activated after exposing BMSCs to homocysteine. Homocysteine treatment caused a significant reduction of BMSCs-secreted VEGF and IGF-1 in the culture medium. Collectively, elevated homocysteine induced the apoptosis of BMSCs via ROS-induced the activation of JNK signal, which provides more insight into the molecular mechanisms of hyperhomocysteinemia-related cardiovascular diseases.

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Effects of homocysteine on MAPKs and apoptotic proteins expression in BMSCs.(a) The total and phosphorylated JNK, p38 and ERK1/2 protein was detected by western blotting in BMSCs after treatment with homocysteine at the time point of 0, 2, 4, 8, 12 and 24 h. Homocysteine effectively activated phosphorylated JNK expression after treatment with homocysteine. But homocysteine did not increase the expression of total JNK protein in BMSCs. (b) Influences of homocysteine on the expression of Bcl-2, caspase-3, cleaved caspase-3, and p-p53 proteins in BMSCs. n = 3 independent experiments.
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pone-0063561-g006: Effects of homocysteine on MAPKs and apoptotic proteins expression in BMSCs.(a) The total and phosphorylated JNK, p38 and ERK1/2 protein was detected by western blotting in BMSCs after treatment with homocysteine at the time point of 0, 2, 4, 8, 12 and 24 h. Homocysteine effectively activated phosphorylated JNK expression after treatment with homocysteine. But homocysteine did not increase the expression of total JNK protein in BMSCs. (b) Influences of homocysteine on the expression of Bcl-2, caspase-3, cleaved caspase-3, and p-p53 proteins in BMSCs. n = 3 independent experiments.

Mentions: To confirm that JNK pathway contributed to homocysteine-induced BMSCs apoptosis, western blot was utilized to detect the expression of JNK, p38 and ERK1/2, as well as p-p53, caspase-3, cleaved caspase-3, Bcl-2 proteins in BMSCs with or without homocysteine 300 µM treatment. Figure 6a showed that homocysteine 300 µM can increase phosphorylated JNK expression (Figure 6a). Moreover, homocysteine treatment did not significantly alter phosphorylated p38 and ERK1/2 protein expression in BMSCs. In order to confirm that homocysteine induced BMSCs apoptosis, we also detected the expression of p-p53, caspase-3, cleaved caspase-3 and Bcl-2 proteins after homocysteine treatment. As shown in Figure 6b, homocysteine did not impact the expression of p-p53, but increased cleaved caspase-3 expression. Bcl-2 was markedly decreased by homocysteine treatment in BMSCs.


Apoptosis of bone marrow mesenchymal stem cells caused by homocysteine via activating JNK signal.

Cai B, Li X, Wang Y, Liu Y, Yang F, Chen H, Yin K, Tan X, Zhu J, Pan Z, Wang B, Lu Y - PLoS ONE (2013)

Effects of homocysteine on MAPKs and apoptotic proteins expression in BMSCs.(a) The total and phosphorylated JNK, p38 and ERK1/2 protein was detected by western blotting in BMSCs after treatment with homocysteine at the time point of 0, 2, 4, 8, 12 and 24 h. Homocysteine effectively activated phosphorylated JNK expression after treatment with homocysteine. But homocysteine did not increase the expression of total JNK protein in BMSCs. (b) Influences of homocysteine on the expression of Bcl-2, caspase-3, cleaved caspase-3, and p-p53 proteins in BMSCs. n = 3 independent experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3646804&req=5

pone-0063561-g006: Effects of homocysteine on MAPKs and apoptotic proteins expression in BMSCs.(a) The total and phosphorylated JNK, p38 and ERK1/2 protein was detected by western blotting in BMSCs after treatment with homocysteine at the time point of 0, 2, 4, 8, 12 and 24 h. Homocysteine effectively activated phosphorylated JNK expression after treatment with homocysteine. But homocysteine did not increase the expression of total JNK protein in BMSCs. (b) Influences of homocysteine on the expression of Bcl-2, caspase-3, cleaved caspase-3, and p-p53 proteins in BMSCs. n = 3 independent experiments.
Mentions: To confirm that JNK pathway contributed to homocysteine-induced BMSCs apoptosis, western blot was utilized to detect the expression of JNK, p38 and ERK1/2, as well as p-p53, caspase-3, cleaved caspase-3, Bcl-2 proteins in BMSCs with or without homocysteine 300 µM treatment. Figure 6a showed that homocysteine 300 µM can increase phosphorylated JNK expression (Figure 6a). Moreover, homocysteine treatment did not significantly alter phosphorylated p38 and ERK1/2 protein expression in BMSCs. In order to confirm that homocysteine induced BMSCs apoptosis, we also detected the expression of p-p53, caspase-3, cleaved caspase-3 and Bcl-2 proteins after homocysteine treatment. As shown in Figure 6b, homocysteine did not impact the expression of p-p53, but increased cleaved caspase-3 expression. Bcl-2 was markedly decreased by homocysteine treatment in BMSCs.

Bottom Line: Apoptosis of BMSCs in response to various pathological stimuli leads to the attenuation of healing ability of BMSCs.Furthermore, apoptotic appearance and mitochondrial membrane potential depolarization in homocysteine-treated BMSCs was significantly reversed by JNK inhibitor but not p38 MAPK and ERK inhibitors.Homocysteine treatment caused a significant reduction of BMSCs-secreted VEGF and IGF-1 in the culture medium.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Harbin Medical University, Harbin, Heilongjiang Province, China.

ABSTRACT
Bone marrow mesenchymal stem cells (BMSCs) are capable of homing to and repair damaged myocardial tissues. Apoptosis of BMSCs in response to various pathological stimuli leads to the attenuation of healing ability of BMSCs. Plenty of evidence has shown that elevated homocysteine level is a novel independent risk factor of cardiovascular diseases. The present study was aimed to investigate whether homocysteine may induce apoptosis of BMSCs and its underlying mechanisms. Here we uncovered that homocysteine significantly inhibited the cellular viability of BMSCs. Furthermore, TUNEL, AO/EB, Hoechst 333342 and Live/Death staining demonstrated the apoptotic morphological appearance of BMSCs after homocysteine treatment. A distinct increase of ROS level was also observed in homocysteine-treated BMSCs. The blockage of ROS by DMTU and NAC prevented the apoptosis of BMSCs induced by homocysteine, indicating ROS was involved in the apoptosis of BMSCs. Moreover, homocysteine also caused the depolarization of mitochondrial membrane potential of BMSCs. Furthermore, apoptotic appearance and mitochondrial membrane potential depolarization in homocysteine-treated BMSCs was significantly reversed by JNK inhibitor but not p38 MAPK and ERK inhibitors. Western blot also confirmed that p-JNK was significantly activated after exposing BMSCs to homocysteine. Homocysteine treatment caused a significant reduction of BMSCs-secreted VEGF and IGF-1 in the culture medium. Collectively, elevated homocysteine induced the apoptosis of BMSCs via ROS-induced the activation of JNK signal, which provides more insight into the molecular mechanisms of hyperhomocysteinemia-related cardiovascular diseases.

Show MeSH
Related in: MedlinePlus