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Mechanisms of the lysophosphatidic acid-induced increase in [Ca(2+)](i) in skeletal muscle cells.

Xu YJ, Tappia PS, Goyal RK, Dhalla NS - J. Cell. Mol. Med. (2008)

Bottom Line: The present study was therefore undertaken to examine the effect of LPA on the [Ca(2+)](i) in C2C12 cells.The LPA effect was also attenuated by ethylene glycolbis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA), an extracellular Ca(2+) chelator, Ni(2+) and KB-R7943, inhibitors of the Na(+)-Ca(2+) exchanger; the receptor operated Ca(2+) channel (ROC) blockers, 2-aminoethoxydiphenyl borate and SK&F 96365.However, the L-type Ca(2+) channel blockers, verapamil and diltiazem; the store operated Ca(2+) channel blockers, La(3+) and Gd(3+); a sarcoplasmic reticulum calcium pump inhibitor, thapsigargin; an inositol trisphosphate receptor antagonist, xestospongin and a phospholipase C inhibitor, U73122, did not prevent the increase [Ca(2+)](i) due to LPA.

View Article: PubMed Central - PubMed

Affiliation: Institute of Cardiovascular Sciences, St. Boniface Hospital Research Centre, Winnipeg, Manitoba, Canada.

ABSTRACT
Although lysophosphatidic acid (LPA) is known to increase intracellularfree calcium concentration ([Ca(2+)](i)) in different cell types, the effect of LPA on the skeletal muscle cells is not known. The present study was therefore undertaken to examine the effect of LPA on the [Ca(2+)](i) in C2C12 cells. LPA induced a concentration and time dependent increase in [Ca(2+)](i), which was inhibited by VPC12249, VPC 32183 and dioctanoyl glycerol pyrophosphate, LPA1/3 receptor antagonists. Pertussis toxin, a G(i) protein inhibitor, also inhibited the LPA-induced increase in [Ca(2+)](i). Inhibition of tyrosine kinase activities with tyrphostin A9 and genistein also prevented the increase in [Ca(2+)](i) due to LPA. Likewise, wortmannin and LY 294002, phosphatidylinositol 3-kinase (PI3-K) inhibitors, inhibited [Ca(2+)](i) response to LPA. The LPA effect was also attenuated by ethylene glycolbis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA), an extracellular Ca(2+) chelator, Ni(2+) and KB-R7943, inhibitors of the Na(+)-Ca(2+) exchanger; the receptor operated Ca(2+) channel (ROC) blockers, 2-aminoethoxydiphenyl borate and SK&F 96365. However, the L-type Ca(2+) channel blockers, verapamil and diltiazem; the store operated Ca(2+) channel blockers, La(3+) and Gd(3+); a sarcoplasmic reticulum calcium pump inhibitor, thapsigargin; an inositol trisphosphate receptor antagonist, xestospongin and a phospholipase C inhibitor, U73122, did not prevent the increase [Ca(2+)](i) due to LPA. Our data suggest that the LPA-induced increase in [Ca(2+)](i) might occur through G(i)-protein coupled LPA(1/3) receptors that may be linked to tyrosine kinase and PI3-K, and may also involve the Na(+)-Ca(2+) exchanger as well as the ROC. In addition, LPA stimulated C2C12 cell proliferation via PI3-K. Thus, LPA may be an important phospholipid in the regulation of [Ca(2+)](i) and growth of skeletal muscle cells.

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Related in: MedlinePlus

Effect of tyrosine kinase inhibitors (A) and phosphatidylinositol kinase 3 inhibitors (B) on LPA-induced changes in intracellular calcium concentration in C2C12 cells. C2C12 cells were pre-treated with 5 μM of the tyrosine kinase inhibitors, tyrphostin A9 (A9) and genistein (GEN), and 100 nM or 10 μM of the phosphatidylinositol 3-kinase inhibitors, wortmannin (WOR) and LY 294002 (LY), respectively, for 10 min prior to the addition of LPA (10 μM). The [Ca2+]i was measured as described in the Materials and methods. Values are means ± S.E.M. of six different experiments. * P<0.05 versusvehicle control value. LPA: sophosphatidic acid, [Ca2+]i: intracellular calcium concentration.
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fig03: Effect of tyrosine kinase inhibitors (A) and phosphatidylinositol kinase 3 inhibitors (B) on LPA-induced changes in intracellular calcium concentration in C2C12 cells. C2C12 cells were pre-treated with 5 μM of the tyrosine kinase inhibitors, tyrphostin A9 (A9) and genistein (GEN), and 100 nM or 10 μM of the phosphatidylinositol 3-kinase inhibitors, wortmannin (WOR) and LY 294002 (LY), respectively, for 10 min prior to the addition of LPA (10 μM). The [Ca2+]i was measured as described in the Materials and methods. Values are means ± S.E.M. of six different experiments. * P<0.05 versusvehicle control value. LPA: sophosphatidic acid, [Ca2+]i: intracellular calcium concentration.

Mentions: To determine the role of tyrosine kinase on LPAinduced increase in [Ca2+]i in C2C12 cells, genistein (5 μM, Sigma-Aldrich, MO, USA) and tyrphostin A9 (A9, 5 μM, Sigma-Aldrich, MO, USA), tyrosine kinase inhibitors [33], were added 10 min prior to the addition of LPA. A significant attenuation of the LPAinduced Ca2+ response was seen by both inhibitors (Fig. 3A). The role of PI3-K in the LPA-induced increase in [Ca2+]i was examined by pre-treating C2C12 cells with the PI3-K inhibitors [17], wortmannin (100 nM) and LY 294002 (10 μM) for 10 min prior to addition of LPA. It can be seen in Figure 3B that both wortmannin and LY 294002 significantly attenuated the Ca2+ response to LPA by 57 % and 63 %, respectively. It should be noted that the pre-treatment the cells with genistein significantly reduced the basal [Ca2+]i (vehicle control: 95 ± 3; genistein: 63 ± 3, P < 0.05). Furthermore, pre-treatment with other agents had no significant effect on the basal [Ca2+]i (vehicle control: 90 ± 10 nM; A9: 85 ± 11 nM; wortmannin: 80 ± 6 nM; LY 294002 85 ± 7 nM).


Mechanisms of the lysophosphatidic acid-induced increase in [Ca(2+)](i) in skeletal muscle cells.

Xu YJ, Tappia PS, Goyal RK, Dhalla NS - J. Cell. Mol. Med. (2008)

Effect of tyrosine kinase inhibitors (A) and phosphatidylinositol kinase 3 inhibitors (B) on LPA-induced changes in intracellular calcium concentration in C2C12 cells. C2C12 cells were pre-treated with 5 μM of the tyrosine kinase inhibitors, tyrphostin A9 (A9) and genistein (GEN), and 100 nM or 10 μM of the phosphatidylinositol 3-kinase inhibitors, wortmannin (WOR) and LY 294002 (LY), respectively, for 10 min prior to the addition of LPA (10 μM). The [Ca2+]i was measured as described in the Materials and methods. Values are means ± S.E.M. of six different experiments. * P<0.05 versusvehicle control value. LPA: sophosphatidic acid, [Ca2+]i: intracellular calcium concentration.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4401138&req=5

fig03: Effect of tyrosine kinase inhibitors (A) and phosphatidylinositol kinase 3 inhibitors (B) on LPA-induced changes in intracellular calcium concentration in C2C12 cells. C2C12 cells were pre-treated with 5 μM of the tyrosine kinase inhibitors, tyrphostin A9 (A9) and genistein (GEN), and 100 nM or 10 μM of the phosphatidylinositol 3-kinase inhibitors, wortmannin (WOR) and LY 294002 (LY), respectively, for 10 min prior to the addition of LPA (10 μM). The [Ca2+]i was measured as described in the Materials and methods. Values are means ± S.E.M. of six different experiments. * P<0.05 versusvehicle control value. LPA: sophosphatidic acid, [Ca2+]i: intracellular calcium concentration.
Mentions: To determine the role of tyrosine kinase on LPAinduced increase in [Ca2+]i in C2C12 cells, genistein (5 μM, Sigma-Aldrich, MO, USA) and tyrphostin A9 (A9, 5 μM, Sigma-Aldrich, MO, USA), tyrosine kinase inhibitors [33], were added 10 min prior to the addition of LPA. A significant attenuation of the LPAinduced Ca2+ response was seen by both inhibitors (Fig. 3A). The role of PI3-K in the LPA-induced increase in [Ca2+]i was examined by pre-treating C2C12 cells with the PI3-K inhibitors [17], wortmannin (100 nM) and LY 294002 (10 μM) for 10 min prior to addition of LPA. It can be seen in Figure 3B that both wortmannin and LY 294002 significantly attenuated the Ca2+ response to LPA by 57 % and 63 %, respectively. It should be noted that the pre-treatment the cells with genistein significantly reduced the basal [Ca2+]i (vehicle control: 95 ± 3; genistein: 63 ± 3, P < 0.05). Furthermore, pre-treatment with other agents had no significant effect on the basal [Ca2+]i (vehicle control: 90 ± 10 nM; A9: 85 ± 11 nM; wortmannin: 80 ± 6 nM; LY 294002 85 ± 7 nM).

Bottom Line: The present study was therefore undertaken to examine the effect of LPA on the [Ca(2+)](i) in C2C12 cells.The LPA effect was also attenuated by ethylene glycolbis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA), an extracellular Ca(2+) chelator, Ni(2+) and KB-R7943, inhibitors of the Na(+)-Ca(2+) exchanger; the receptor operated Ca(2+) channel (ROC) blockers, 2-aminoethoxydiphenyl borate and SK&F 96365.However, the L-type Ca(2+) channel blockers, verapamil and diltiazem; the store operated Ca(2+) channel blockers, La(3+) and Gd(3+); a sarcoplasmic reticulum calcium pump inhibitor, thapsigargin; an inositol trisphosphate receptor antagonist, xestospongin and a phospholipase C inhibitor, U73122, did not prevent the increase [Ca(2+)](i) due to LPA.

View Article: PubMed Central - PubMed

Affiliation: Institute of Cardiovascular Sciences, St. Boniface Hospital Research Centre, Winnipeg, Manitoba, Canada.

ABSTRACT
Although lysophosphatidic acid (LPA) is known to increase intracellularfree calcium concentration ([Ca(2+)](i)) in different cell types, the effect of LPA on the skeletal muscle cells is not known. The present study was therefore undertaken to examine the effect of LPA on the [Ca(2+)](i) in C2C12 cells. LPA induced a concentration and time dependent increase in [Ca(2+)](i), which was inhibited by VPC12249, VPC 32183 and dioctanoyl glycerol pyrophosphate, LPA1/3 receptor antagonists. Pertussis toxin, a G(i) protein inhibitor, also inhibited the LPA-induced increase in [Ca(2+)](i). Inhibition of tyrosine kinase activities with tyrphostin A9 and genistein also prevented the increase in [Ca(2+)](i) due to LPA. Likewise, wortmannin and LY 294002, phosphatidylinositol 3-kinase (PI3-K) inhibitors, inhibited [Ca(2+)](i) response to LPA. The LPA effect was also attenuated by ethylene glycolbis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA), an extracellular Ca(2+) chelator, Ni(2+) and KB-R7943, inhibitors of the Na(+)-Ca(2+) exchanger; the receptor operated Ca(2+) channel (ROC) blockers, 2-aminoethoxydiphenyl borate and SK&F 96365. However, the L-type Ca(2+) channel blockers, verapamil and diltiazem; the store operated Ca(2+) channel blockers, La(3+) and Gd(3+); a sarcoplasmic reticulum calcium pump inhibitor, thapsigargin; an inositol trisphosphate receptor antagonist, xestospongin and a phospholipase C inhibitor, U73122, did not prevent the increase [Ca(2+)](i) due to LPA. Our data suggest that the LPA-induced increase in [Ca(2+)](i) might occur through G(i)-protein coupled LPA(1/3) receptors that may be linked to tyrosine kinase and PI3-K, and may also involve the Na(+)-Ca(2+) exchanger as well as the ROC. In addition, LPA stimulated C2C12 cell proliferation via PI3-K. Thus, LPA may be an important phospholipid in the regulation of [Ca(2+)](i) and growth of skeletal muscle cells.

Show MeSH
Related in: MedlinePlus