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IL-8/CXCL8 Upregulates 12-Lipoxygenase Expression in Vascular Smooth Muscle Cells from Spontaneously Hypertensive Rats.

Kim JH, Kang YJ, Kim HS - Immune Netw (2009)

Bottom Line: In the present study, we investigated the direct effect of IL-8/CXCL8 on expression of 12-lipoxygenase (LO), a hypertensive modulator, in SHR VSMC.Treatment with IL-8/CXCL8 greatly increased 12-LO mRNA expression and protein production compared to treatment with angiotensin II.These results suggest that the potential role of IL-8/CXCL8 in hypertensive processes is likely mediated through the 12-LO pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, College of Medicine, Yeungnam University, Daegu, Korea.

ABSTRACT

Background: We previously demonstrated remarkable differences in the expression of IL-8/CXCL8 in aortic tissues and vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) compared to VSMC from normotensive Wistar-Kyoto rats (WKY). In the present study, we investigated the direct effect of IL-8/CXCL8 on expression of 12-lipoxygenase (LO), a hypertensive modulator, in SHR VSMC.

Methods: Cultured aortic VSMC from SHR and WKY were used. Expression of 12-LO mRNA was determined by real-time polymerase chain reaction. Phosphorlyation of ERK1/2 and production of 12-LO and angiotensin II subtype 1 (AT(1)) receptor were assessed by Western blots. IL-8/CXCL8-stimulated DNA synthesis was determined by measuring incorporation of [(3)H]-thymidine. And effect of IL-8/CXCL8 on vascular tone was determined by phenylephrine-induced contraction of thoracic aortic rings.

Results: Treatment with IL-8/CXCL8 greatly increased 12-LO mRNA expression and protein production compared to treatment with angiotensin II. IL-8/CXCL8 also increased the expression of the AT(1) receptor. The increase in 12-LO induced by IL-8/CXCL8 was inhibited by treatment with an AT(1) receptor antagonist. The induction of 12-LO mRNA production and the proliferation of SHR VSMC by IL-8/CXCL8 was mediated by the ERK pathway. The proliferation of SHR VSMC and the vascular contraction in the thoracic aortic ring, both of which were induced by IL-8/CXCL8, were inhibited by baicalein, a 12-LO inhibitor.

Conclusion: These results suggest that the potential role of IL-8/CXCL8 in hypertensive processes is likely mediated through the 12-LO pathway.

No MeSH data available.


Related in: MedlinePlus

Expression of IL-8/CXCL8-induced 12-LO is mediated through the ERK pathway, and proliferation of SHR VSMC by IL-8/CXCL8 is inhibited by baicalein and PD98059. (A, B) VSMC were untreated (NT) or pretreated with PD98059 (ERK inhibitor, 10 µM) for 30 min. Cells were left untreated or treated with IL-8/CXCL8 (100 ng/ml) for 2 h, and the total RNA and cell lysates were isolated. The total RNA was analyzed by real-time PCR (A), and cell lysates were separated on 10% SDS-polyacrylamide gels and then immunoblotted with the phospho-ERK1/2 antibody (B). Bars represent means±SD from three independent experiments. *p<0.05 vs. VSMC treated with IL-8/CXCL8. Data shown are representative of three independent experiments. (C, D) SHR VSMC were treated with 12-HETE (500 nmol/L), with IL-8/CXCL8 (100 ng/ml), with baicalein (12-LO inhibitor, 10 µmol/L, B), or with PD98059 (10 µmol/L, C) for 48 h in medium containing [3H]-thymidine (1 µCi/ml). [3H]-thymidine incorporation is shown on the Y-axis. Bars represent means±SD from three independent experiments run in triplicate. *p<0.05 vs. untreated VSMC. **p<0.01 vs. VSMC treated with IL-8/CXCL8 alone. a: p<0.05 vs. VSMC treated with IL-8/CXCL8 alone.
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Figure 3: Expression of IL-8/CXCL8-induced 12-LO is mediated through the ERK pathway, and proliferation of SHR VSMC by IL-8/CXCL8 is inhibited by baicalein and PD98059. (A, B) VSMC were untreated (NT) or pretreated with PD98059 (ERK inhibitor, 10 µM) for 30 min. Cells were left untreated or treated with IL-8/CXCL8 (100 ng/ml) for 2 h, and the total RNA and cell lysates were isolated. The total RNA was analyzed by real-time PCR (A), and cell lysates were separated on 10% SDS-polyacrylamide gels and then immunoblotted with the phospho-ERK1/2 antibody (B). Bars represent means±SD from three independent experiments. *p<0.05 vs. VSMC treated with IL-8/CXCL8. Data shown are representative of three independent experiments. (C, D) SHR VSMC were treated with 12-HETE (500 nmol/L), with IL-8/CXCL8 (100 ng/ml), with baicalein (12-LO inhibitor, 10 µmol/L, B), or with PD98059 (10 µmol/L, C) for 48 h in medium containing [3H]-thymidine (1 µCi/ml). [3H]-thymidine incorporation is shown on the Y-axis. Bars represent means±SD from three independent experiments run in triplicate. *p<0.05 vs. untreated VSMC. **p<0.01 vs. VSMC treated with IL-8/CXCL8 alone. a: p<0.05 vs. VSMC treated with IL-8/CXCL8 alone.

Mentions: We investigated whether the MAPK signaling pathway is involved in IL-8/CXCL8-induced 12-LO expression. The IL-8/CXCL8-induced expression of 12-LO mRNA was decreased by PD98059 (Fig. 3A), and a high level of phosphorylation of ERK in SHR VSMC treated with IL-8/CXCL8 was detected (Fig. 3B)


IL-8/CXCL8 Upregulates 12-Lipoxygenase Expression in Vascular Smooth Muscle Cells from Spontaneously Hypertensive Rats.

Kim JH, Kang YJ, Kim HS - Immune Netw (2009)

Expression of IL-8/CXCL8-induced 12-LO is mediated through the ERK pathway, and proliferation of SHR VSMC by IL-8/CXCL8 is inhibited by baicalein and PD98059. (A, B) VSMC were untreated (NT) or pretreated with PD98059 (ERK inhibitor, 10 µM) for 30 min. Cells were left untreated or treated with IL-8/CXCL8 (100 ng/ml) for 2 h, and the total RNA and cell lysates were isolated. The total RNA was analyzed by real-time PCR (A), and cell lysates were separated on 10% SDS-polyacrylamide gels and then immunoblotted with the phospho-ERK1/2 antibody (B). Bars represent means±SD from three independent experiments. *p<0.05 vs. VSMC treated with IL-8/CXCL8. Data shown are representative of three independent experiments. (C, D) SHR VSMC were treated with 12-HETE (500 nmol/L), with IL-8/CXCL8 (100 ng/ml), with baicalein (12-LO inhibitor, 10 µmol/L, B), or with PD98059 (10 µmol/L, C) for 48 h in medium containing [3H]-thymidine (1 µCi/ml). [3H]-thymidine incorporation is shown on the Y-axis. Bars represent means±SD from three independent experiments run in triplicate. *p<0.05 vs. untreated VSMC. **p<0.01 vs. VSMC treated with IL-8/CXCL8 alone. a: p<0.05 vs. VSMC treated with IL-8/CXCL8 alone.
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Figure 3: Expression of IL-8/CXCL8-induced 12-LO is mediated through the ERK pathway, and proliferation of SHR VSMC by IL-8/CXCL8 is inhibited by baicalein and PD98059. (A, B) VSMC were untreated (NT) or pretreated with PD98059 (ERK inhibitor, 10 µM) for 30 min. Cells were left untreated or treated with IL-8/CXCL8 (100 ng/ml) for 2 h, and the total RNA and cell lysates were isolated. The total RNA was analyzed by real-time PCR (A), and cell lysates were separated on 10% SDS-polyacrylamide gels and then immunoblotted with the phospho-ERK1/2 antibody (B). Bars represent means±SD from three independent experiments. *p<0.05 vs. VSMC treated with IL-8/CXCL8. Data shown are representative of three independent experiments. (C, D) SHR VSMC were treated with 12-HETE (500 nmol/L), with IL-8/CXCL8 (100 ng/ml), with baicalein (12-LO inhibitor, 10 µmol/L, B), or with PD98059 (10 µmol/L, C) for 48 h in medium containing [3H]-thymidine (1 µCi/ml). [3H]-thymidine incorporation is shown on the Y-axis. Bars represent means±SD from three independent experiments run in triplicate. *p<0.05 vs. untreated VSMC. **p<0.01 vs. VSMC treated with IL-8/CXCL8 alone. a: p<0.05 vs. VSMC treated with IL-8/CXCL8 alone.
Mentions: We investigated whether the MAPK signaling pathway is involved in IL-8/CXCL8-induced 12-LO expression. The IL-8/CXCL8-induced expression of 12-LO mRNA was decreased by PD98059 (Fig. 3A), and a high level of phosphorylation of ERK in SHR VSMC treated with IL-8/CXCL8 was detected (Fig. 3B)

Bottom Line: In the present study, we investigated the direct effect of IL-8/CXCL8 on expression of 12-lipoxygenase (LO), a hypertensive modulator, in SHR VSMC.Treatment with IL-8/CXCL8 greatly increased 12-LO mRNA expression and protein production compared to treatment with angiotensin II.These results suggest that the potential role of IL-8/CXCL8 in hypertensive processes is likely mediated through the 12-LO pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, College of Medicine, Yeungnam University, Daegu, Korea.

ABSTRACT

Background: We previously demonstrated remarkable differences in the expression of IL-8/CXCL8 in aortic tissues and vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) compared to VSMC from normotensive Wistar-Kyoto rats (WKY). In the present study, we investigated the direct effect of IL-8/CXCL8 on expression of 12-lipoxygenase (LO), a hypertensive modulator, in SHR VSMC.

Methods: Cultured aortic VSMC from SHR and WKY were used. Expression of 12-LO mRNA was determined by real-time polymerase chain reaction. Phosphorlyation of ERK1/2 and production of 12-LO and angiotensin II subtype 1 (AT(1)) receptor were assessed by Western blots. IL-8/CXCL8-stimulated DNA synthesis was determined by measuring incorporation of [(3)H]-thymidine. And effect of IL-8/CXCL8 on vascular tone was determined by phenylephrine-induced contraction of thoracic aortic rings.

Results: Treatment with IL-8/CXCL8 greatly increased 12-LO mRNA expression and protein production compared to treatment with angiotensin II. IL-8/CXCL8 also increased the expression of the AT(1) receptor. The increase in 12-LO induced by IL-8/CXCL8 was inhibited by treatment with an AT(1) receptor antagonist. The induction of 12-LO mRNA production and the proliferation of SHR VSMC by IL-8/CXCL8 was mediated by the ERK pathway. The proliferation of SHR VSMC and the vascular contraction in the thoracic aortic ring, both of which were induced by IL-8/CXCL8, were inhibited by baicalein, a 12-LO inhibitor.

Conclusion: These results suggest that the potential role of IL-8/CXCL8 in hypertensive processes is likely mediated through the 12-LO pathway.

No MeSH data available.


Related in: MedlinePlus