Janus Kinase 2/Signal Transducer and Activator of Transcription 3/Cyclooxygenase 2 Signaling Pathway Mediates the Effect of Central Angiotensin II on the Elevation of Rostral Ventrolateral Medulla Prostaglandin E 2 ‐Induced Oxidative Stress in Hypertension

Figures

• 

  Figure 1. Effect of PGE2 in the RVLM on sympathetic activity. The PGE2 concentration in the RVLM in the WKY rats and SHRs (A). Representative original recordings of ABP, MAP, HR, and RSNA response (B) and percentage changes in MAP (C) and RSNA (D) to microinjection of PGE2 into the RVLM of WKY rats. MAP (E) and plasma NE level (F) in SHRs with intracisternal infusion of aCSF or angiotensin II. Values are mean±SD, n=5 per group. ABP indicates arterial blood pressure; aCSF, artificial cerebrospinal fluid; HR, heart rate; Inte‐RSNA, integrated RSNA; MAP, mean arterial pressure; NE, norepinephrine; PGE2, prostaglandin E2; RSNA, renal sympathetic nerve activation; RVLM, rostral ventrolateral medulla; SHRs, spontaneously hypertensive rats; and WKY, Wistar‐Kyoto; WYK, Wistar‐Kyoto Rats.

• 

  Figure 2. Effect of PGE2 in the RVLM on oxidative stress. ROS level in microinjection of PGE2 into the rostral ventrolateral medulla of WKY rats (A). Representative original recordings of ABP, MAP, HR, and RSNA response (B) and percentage changes in MAP (C) and RSNA (D) to microinjection of Tempol and PGE2 into the rostral ventrolateral medulla of WKY rats. ROS level (E) in SHRs with intracisternal infusion of aCSF or L‐798106. Values are mean±SD, n=5 per group. ABP indicates arterial blood pressure; aCSF, artificial cerebrospinal fluid; HR, heart rate; Inte‐RSNA, integrated RSNA; MAP, mean arterial pressure; PGE2, prostaglandin E2; ROS, reactive oxygen species; RSNA, renal sympathetic nerve activation; RVLM; rostral ventrolateral medulla; SHRs, spontaneously hypertensive rats; and WKY, Wistar‐Kyoto.

• 

  Figure 3. Effect of central Ang II on PGE2 and oxidative stress in the RVLM. PGE2 concentration in the RVLM of WKY rats with intracisternal infusion Ang II (A) and in SHRs with intracisternal infusion of L‐798106 (B). MAP (C), plasma NE level (D), and ROS level in the RVLM (E) in WKY rats with intracisternal infusion of both Ang II and L‐798106. ROS level in cultured PC12 cells with treatment of Ang II and L‐798106 (F). Representative Western blot bands (G) and densitometric analysis of NOX2 (H), NOX4 (I), SOD1 (J), and SOD2 (K) in cultured PC12 cells with treatment of Ang II and L‐798106. Each bar represents an experimental group, with the annotation below the x axis indicating the drug treatment protocol. Values are mean±SD, n=5 per group in (A) through (E), and n=3 per group in (F) through (K). aCSF indicates artificial cerebrospinal fluid; Ang II, angiotensin II; GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; MAP, mean arterial pressure; NE, norepinephrine; NOX2, NADPH oxidase 2; NOX4, NADPH oxidase 4; PGE2, prostaglandin E2; ROS, reactive oxygen species; RVLM; rostral ventrolateral medulla; SHRs, spontaneously hypertensive rats; SOD1, superoxide dismutase 1; SOD2, superoxide…

• 

  Figure 4. Effect of central Ang II on COX2 expression in the RVLM. Representative Western blot bands (top) and densitometric analysis (bottom) of COX2 (A), COX1 (B), and PGES (C) in the RVLM of WKY rats with intracisternal infusion Ang II. Representative fluorescence images of rat brain sections showing cell localization of COX1 (blue), COX2 (green), and NeuN (orange fluorescence) represent presynaptic neurons in the RVLM (D) (white arrow shows the colocalization of COX2 and NeuN). Values are mean±SD, n=5 per group. aCSF indicates artificial cerebrospinal fluid; Ang II, angiotensin II; COX1, cyclooxygenase 1; COX2, cyclooxygenase 2; GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; NeuN, neuron‐specific nuclear protein; PGES, prostaglandin E synthases; RVLM; rostral ventrolateral medulla; and WKY, Wistar‐Kyoto.

• 

  Figure 5. Effect of COX2 on the PGE2 and blood pressure regulation. PGE2 concentration (A), MAP (B), plasma NE level (C) in WKY‐GFP and WKY‐COX2 group. PGE2 concentration (D), MAP (E), and plasma NE level (F) in the SHR‐GFP group and SHR‐shRNA group. PGE2 concentration (G), MAP (H), and plasma NE level (I) in WT plus aCSF, WT plus Ang II, COX‐2 KO plus aCSF, and COX2 KO plus Ang II group. Values are mean±SD, n=5 per group. aCSF indicates artificial cerebrospinal fluid; Ang II, angiotensin II; COX2, cyclooxygenase 2; GFP, green fluorescent protein; KO, knock‐out; MAP, mean arterial pressure; NE, norepinephrine; PGE2, prostaglandin E2; SHR, spontaneously hypertensive rats; shRNA, short hairpin RNA; WKY, Wistar‐Kyoto; and WT, wild‐type.

• 

  Figure 6. Effect of COX2 on PGE2 and oxidative stress. ROS level in the RVLM in WKY‐GFP and WKY‐COX2 group (A). Representative Western blot bands (top) and densitometric analysis (bottom) of NOX2 (B), NOX4 (C), SOD1 (D), and SOD2 (E) in the RVLM of WKY‐GFP and WKY‐COX2 group. ROS level in the RVLM in WT plus Ang II and COX2‐KO plus Ang II group (F). Values are mean±SD, n=5 per group. Ang II indicates angiotensin II; COX2, cyclooxygenase 2; GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; GFP, green fluorescent protein; KO, knock‐out; NOX2, NADPH oxidase 2; NOX4, NADPH oxidase 4; PGE2, prostaglandin E2; ROS, reactive oxygen species; RVLM; rostral ventrolateral medulla; SOD1, superoxide dismutase 1; SOD2, superoxide dismutase 2; WKY, Wistar‐Kyoto; and WT, wild‐type.

• 

  Figure 7. Effect of JAK2/STAT3 signaling pathway in Ang II‐induced COX2 expression enhancement. Relative COX2 mRNA level in PC12 cells treated by Ang II and signal pathway inhibitors (PD169316, WP1066, A‐443654, Rhosin, and BMS‐345541) (A). Representative Western blot bands (B) and densitometric analysis of p‐JAK2/JAK2 (Phosphorylated Janus kinase 2) level (C), p‐STAT3/STAT3, (Phosphorylated Signal transducer and activator of transcription 3) level (D), and COX2 (E) in the PC12 cells treated by Ang II and WP1066. Relative enrichment fold of COX2 DNA sequence by p‐STAT3 and IgG (negative control) antibody in the PC12 cells with angiotensin or PBS treatment (F). Each bar represents an experimental group, with the annotation below the x axis indicating the drug treatment protocol. Values are mean±SD, n=3 per group. Ang II indicates angiotensin II; COX2, cyclooxygenase 2; GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; IgG, immunoglobulin G; JAK2, Janus kinase 2; and STAT3, signal transducer and activator of transcription 3.