Journal of Traditional Chinese Medicine ›› 2026, Vol. 46 ›› Issue (4): 907-915.DOI: 10.19852/j.cnki.jtcm.2026.04.011
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Antihypertensive effect of acupuncture involve reshaping gut microbiota and alleviating gut inflammation in spontaneously hypertensive rats
ZHENG Jie1, DING Zhimin2, CHEN Tingyu1, LÜ Zhuan3, ZHANG Lihong1, LIU Yongtao3, ZHANG Yasu1, JIN Xiaoqin1, WANG Xinyu1, LIU Bin1, FENG Xiaodong1(
)
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School of Rehabilitation Medicine ,Henan University of Chinese Medicine Zhengzhou 450046, China
2College of Rehabilitation Medicine ,Fujian University of Traditional Chinese Medicine Fuzhou 350122, China
3Department of Rehabilitation ,the First Affiliated Hospital of Henan University of Chinese Medicine Zhengzhou 450000, China
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Received:2025-08-09Accepted:2026-04-13Online:2026-08-15Published:2026-08-08 -
Contact:Prof. FENG Xiaodong, School of Rehabilitation Medicine, Henan University of Chinese Medicine, Zhengzhou 450046, China. fxd0502@163.com,Telephone: +86-15516164325; +86-15303828605
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About author:ZHENG Jie and DING Zhimin are co-first authors and contributed equally to this work -
Supported by:Provincial Natural Science Foundation of Henan: Mechanism Underlying Antihypertensive Effect of Acupuncture at Renying (ST9) and Zusanli (ST36) in Spontaneously Hypertensive Rats Based on Gut-immune Axis(202300410260);Key Research Project Program of Higher Education Institutions in Henan Province: Investigation of the Neuroprotective Mechanism of Acupuncture at Renying (ST9) and Zusanli (ST36) in Hypertension via the Gut-Brain Axis(25B360011);Special Project for Scientific Research on Traditional Chinese Medicine of the "Double First-class" Founding Disciplines of Henan Province(HSRP-DFCTCM-T-9)
Cite this article
ZHENG Jie, DING Zhimin, CHEN Tingyu, LÜ Zhuan, ZHANG Lihong, LIU Yongtao, ZHANG Yasu, JIN Xiaoqin, WANG Xinyu, LIU Bin, FENG Xiaodong. Antihypertensive effect of acupuncture involve reshaping gut microbiota and alleviating gut inflammation in spontaneously hypertensive rats[J]. Journal of Traditional Chinese Medicine, 2026, 46(4): 907-915.
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Figure 1 Effect of acupuncture on HMGB1, IL-10, and C1q proteins at Zusanli (ST36) A: concentration of HMGB1 proteins; B: concentration of IL-10 proteins; C: concentration of C1q proteins; A1, B1, C1: D1 after acupuncture; A2, B2, C2: D3 after acupuncture; A3, B3, C3: D5 after acupuncture; A4, B4, C4: D7 after acupuncture; D-F: the temporal changes of HMGB1, IL-10 and C1q proteins after acupuncture (D3 vs D1, D5 vs D3, D7 vs D5). D: HMGB1 proteins; E: IL-10 proteins; F: C1q proteins. WT group: wild-type mice without acupuncture intervention, ACU group: wild-type mice receiving bilateral Zusanli (ST36) acupuncture stimulation. Acupuncture was performed at Zusanli (ST36) using sterilized stainless-steel filiform needles in 8-min daily sessions for 1, 3, 5, or 7 d. HMGB1: high mobility group box 1; IL-10: interleukin-10; C1q: complement component 1q; D: day. Data are presented as mean ± standard deviation (n = 5-7 per group). The t-test was used for data analysis between two groups. aP < 0.05, bP < 0.01, cP < 0.001 versus the WT group.
Figure 1 Effect of acupuncture on HMGB1, IL-10, and C1q proteins at Zusanli (ST36) A: concentration of HMGB1 proteins; B: concentration of IL-10 proteins; C: concentration of C1q proteins; A1, B1, C1: D1 after acupuncture; A2, B2, C2: D3 after acupuncture; A3, B3, C3: D5 after acupuncture; A4, B4, C4: D7 after acupuncture; D-F: the temporal changes of HMGB1, IL-10 and C1q proteins after acupuncture (D3 vs D1, D5 vs D3, D7 vs D5). D: HMGB1 proteins; E: IL-10 proteins; F: C1q proteins. WT group: wild-type mice without acupuncture intervention, ACU group: wild-type mice receiving bilateral Zusanli (ST36) acupuncture stimulation. Acupuncture was performed at Zusanli (ST36) using sterilized stainless-steel filiform needles in 8-min daily sessions for 1, 3, 5, or 7 d. HMGB1: high mobility group box 1; IL-10: interleukin-10; C1q: complement component 1q; D: day. Data are presented as mean ± standard deviation (n = 5-7 per group). The t-test was used for data analysis between two groups. aP < 0.05, bP < 0.01, cP < 0.001 versus the WT group.
Figure 2 Acupuncture-induced C1q involved in regulating the inflammatory responses at Zusanli (ST36) A: representative hematoxylin-eosin staining images of Zusanli (ST36) tissues (Nuclei: blue, cytoplasm: red, Scale bar = 50 µm, n = 3, magnification × 40); A1-A3: the epidermis, dermis layers of the WT, ACU, and C1q-/-+ACU groups; A1:WT groups; A2: ACU groups; A3: C1q-/-+ACU groups; A4-A6: the muscle layers of the WT, ACU, and C1q-/-+ACU groups; A4: WT groups; A5: ACU groups; A6: C1q-/-+ACU groups; B: single-label immunofluorescence of p-p65 (p-p65: green, 1:100, DAPI: blue, Scale bar = 10 µm, n = 3, magnification × 40). B1-B3: representative merged images showing p-p65 with DAPI; B1: WT groups; B2: ACU groups; B3: C1q-/-+ACU groups; C: quantification of p-p65-positive cells by immunofluorescence (n = 3); D: protein concentration of HMGB1 at Zusanli (ST36) measured by ELISA (n = 5-6); E: protein concentration of IL-10 at Zusanli (ST36) measured by ELISA (n = 5-6). WT group: wild-type mice without acupuncture intervention, ACU group: wild-type mice receiving bilateral Zusanli (ST36) acupuncture stimulation, C1q-/-+ACU group: C1q knockout mice receiving acupuncture. Acupuncture was performed at Zusanli (ST36) using sterilized stainless-steel filiform needles in 8-min daily sessions for 7 d. HMGB1: high mobility group box 1; IL-10: interleukin-10; ELISA: enzyme-linked immunosorbent assay; p-p65: phosphorylated p65. Data are presented as mean ± standard deviation. One-way analysis of variance was used for multiple groups comparisons. If the data followed a normal distribution, the least significant difference test was used; otherwise, Dunnett’s T3 test was used. aP < 0.05, cP < 0.01 versus the WT group; bP < 0.05 versus the ACU group.
Figure 2 Acupuncture-induced C1q involved in regulating the inflammatory responses at Zusanli (ST36) A: representative hematoxylin-eosin staining images of Zusanli (ST36) tissues (Nuclei: blue, cytoplasm: red, Scale bar = 50 µm, n = 3, magnification × 40); A1-A3: the epidermis, dermis layers of the WT, ACU, and C1q-/-+ACU groups; A1:WT groups; A2: ACU groups; A3: C1q-/-+ACU groups; A4-A6: the muscle layers of the WT, ACU, and C1q-/-+ACU groups; A4: WT groups; A5: ACU groups; A6: C1q-/-+ACU groups; B: single-label immunofluorescence of p-p65 (p-p65: green, 1:100, DAPI: blue, Scale bar = 10 µm, n = 3, magnification × 40). B1-B3: representative merged images showing p-p65 with DAPI; B1: WT groups; B2: ACU groups; B3: C1q-/-+ACU groups; C: quantification of p-p65-positive cells by immunofluorescence (n = 3); D: protein concentration of HMGB1 at Zusanli (ST36) measured by ELISA (n = 5-6); E: protein concentration of IL-10 at Zusanli (ST36) measured by ELISA (n = 5-6). WT group: wild-type mice without acupuncture intervention, ACU group: wild-type mice receiving bilateral Zusanli (ST36) acupuncture stimulation, C1q-/-+ACU group: C1q knockout mice receiving acupuncture. Acupuncture was performed at Zusanli (ST36) using sterilized stainless-steel filiform needles in 8-min daily sessions for 7 d. HMGB1: high mobility group box 1; IL-10: interleukin-10; ELISA: enzyme-linked immunosorbent assay; p-p65: phosphorylated p65. Data are presented as mean ± standard deviation. One-way analysis of variance was used for multiple groups comparisons. If the data followed a normal distribution, the least significant difference test was used; otherwise, Dunnett’s T3 test was used. aP < 0.05, cP < 0.01 versus the WT group; bP < 0.05 versus the ACU group.
Figure 3 Mechanism of C1q regulation of the inflammatory response at Zusanli (ST36) A: representative Western blot bands showing protein expression of RAGE, LAIR-1, and SHP-1 at Zusanli (ST36) (n = 5-6); B: quantification of LAIR-1 normalized to GAPDH; C: quantification of RAGE normalized to GAPDH; D: quantification of SHP-1normalized to GAPDH; E: double-label immunofluorescence of M1 macrophages (M0: F4/80, red, 1:50; M1: CD86, green, 1:100; Merged: F4/80, CD86, and DAPI; Scale bar = 100 µm; magnification ×40, n = 3); E1: F4/80-label of WT groups; E2: F4/80-label of ACU groups; E3: F4/80-label of C1q-/-+ACU groups, E4: CD86-label of WT groups; E5: CD86-label of ACU groups; E6: CD86-label of C1q-/-+ACU groups; E7: merged of WT groups; E8: merged of ACU groups; E9: merged of C1q-/-+ACU groups, F: double-label immunofluorescence of M2 macrophages (M0: F4/80, red, 1:50; M2: CD206, green, 1:50, Merged: F4/80, CD206, and DAPI; Scale bar = 100 µm; magnification ×40, n = 3); F1: F4/80-label of WT groups; F2: F4/80-label of ACU groups; F3: F4/80-label of C1q-/-+ACU groups, F4: CD206-label of WT groups; F5: CD206-label of ACU groups; F6: CD206-label of C1q-/-+ACU groups; F7: merged of WT groups; F8: merged of ACU groups; F9: merged of C1q-/-+ACU groups; G : quantitative immunofluorescence analysis of M1 macrophages at Zusanli (ST36) (n = 3); H: quantitative immunofluorescence analysis of M2 macrophages at Zusanli (ST36) (n = 3). WT group: wild-type mice without acupuncture intervention, ACU group: wild-type mice receiving bilateral Zusanli (ST36) acupuncture stimulation, C1q-/-+ACU group: C1q knockout mice receiving acupuncture. Acupuncture was performed at Zusanli (ST36) using sterilized stainless-steel filiform needles in 8-min daily sessions for 7 d. RAGE: the receptor for advanced glycation end-products; LAIR-1: leukocyte-associated immunoglobulin-like receptor 1; SHP-1: src homology 2 domain-containing protein tyrosine phosphatase 1; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; M0: Naïve Macrophage; M1: classical activated macrophages; M2: alternatively activated macrophages. Data are presented as mean ± standard deviation. One-way analysis of variance was used for multiple groups comparisons. If the data followed a normal distribution, the least significant difference test was used; otherwise, Dunnett’s T3 test was used. cP < 0.01, aP < 0.001 versus the WT group; bP < 0.05, dP < 0.001 versus the ACU group.
Figure 3 Mechanism of C1q regulation of the inflammatory response at Zusanli (ST36) A: representative Western blot bands showing protein expression of RAGE, LAIR-1, and SHP-1 at Zusanli (ST36) (n = 5-6); B: quantification of LAIR-1 normalized to GAPDH; C: quantification of RAGE normalized to GAPDH; D: quantification of SHP-1normalized to GAPDH; E: double-label immunofluorescence of M1 macrophages (M0: F4/80, red, 1:50; M1: CD86, green, 1:100; Merged: F4/80, CD86, and DAPI; Scale bar = 100 µm; magnification ×40, n = 3); E1: F4/80-label of WT groups; E2: F4/80-label of ACU groups; E3: F4/80-label of C1q-/-+ACU groups, E4: CD86-label of WT groups; E5: CD86-label of ACU groups; E6: CD86-label of C1q-/-+ACU groups; E7: merged of WT groups; E8: merged of ACU groups; E9: merged of C1q-/-+ACU groups, F: double-label immunofluorescence of M2 macrophages (M0: F4/80, red, 1:50; M2: CD206, green, 1:50, Merged: F4/80, CD206, and DAPI; Scale bar = 100 µm; magnification ×40, n = 3); F1: F4/80-label of WT groups; F2: F4/80-label of ACU groups; F3: F4/80-label of C1q-/-+ACU groups, F4: CD206-label of WT groups; F5: CD206-label of ACU groups; F6: CD206-label of C1q-/-+ACU groups; F7: merged of WT groups; F8: merged of ACU groups; F9: merged of C1q-/-+ACU groups; G : quantitative immunofluorescence analysis of M1 macrophages at Zusanli (ST36) (n = 3); H: quantitative immunofluorescence analysis of M2 macrophages at Zusanli (ST36) (n = 3). WT group: wild-type mice without acupuncture intervention, ACU group: wild-type mice receiving bilateral Zusanli (ST36) acupuncture stimulation, C1q-/-+ACU group: C1q knockout mice receiving acupuncture. Acupuncture was performed at Zusanli (ST36) using sterilized stainless-steel filiform needles in 8-min daily sessions for 7 d. RAGE: the receptor for advanced glycation end-products; LAIR-1: leukocyte-associated immunoglobulin-like receptor 1; SHP-1: src homology 2 domain-containing protein tyrosine phosphatase 1; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; M0: Naïve Macrophage; M1: classical activated macrophages; M2: alternatively activated macrophages. Data are presented as mean ± standard deviation. One-way analysis of variance was used for multiple groups comparisons. If the data followed a normal distribution, the least significant difference test was used; otherwise, Dunnett’s T3 test was used. cP < 0.01, aP < 0.001 versus the WT group; bP < 0.05, dP < 0.001 versus the ACU group.
Figure 4 Gut microbiota correlations with blood pressure and inflammatory cytokines A: Spearman’s correlation between gut genera and blood pressure, including SBP and DBP; B: Spearman’s correlation between gut genera and serum inflammatory cytokines, including IL-6, IL-8, IL-1β and TNF-α. WKY and SHR groups: restrained and fixed without any intervention; SHR+A group: treated with acupuncture at bilateral Renying (ST9) and Zusanli (ST36); SHR+SA group: treated with sham acupuncture at non-meridian non-acupuncture points adjacent to bilateral Renying (ST9) and Zusanli (ST36). SBP: systolic blood pressure; DBP: diastolic blood pressure; WKY: Wistar-Kyoto rats; SHRs: spontaneously hypertensive rats; SEM: standard error of the mean. Data are presented as mean ± SEM and Spearman’s correlation analysis was adopted (n = 6 per group). Comparisons between indicators on the horizontal and vertical axes, *P < 0.05; **P < 0.01; ***P < 0.001.
Figure 4 Gut microbiota correlations with blood pressure and inflammatory cytokines A: Spearman’s correlation between gut genera and blood pressure, including SBP and DBP; B: Spearman’s correlation between gut genera and serum inflammatory cytokines, including IL-6, IL-8, IL-1β and TNF-α. WKY and SHR groups: restrained and fixed without any intervention; SHR+A group: treated with acupuncture at bilateral Renying (ST9) and Zusanli (ST36); SHR+SA group: treated with sham acupuncture at non-meridian non-acupuncture points adjacent to bilateral Renying (ST9) and Zusanli (ST36). SBP: systolic blood pressure; DBP: diastolic blood pressure; WKY: Wistar-Kyoto rats; SHRs: spontaneously hypertensive rats; SEM: standard error of the mean. Data are presented as mean ± SEM and Spearman’s correlation analysis was adopted (n = 6 per group). Comparisons between indicators on the horizontal and vertical axes, *P < 0.05; **P < 0.01; ***P < 0.001.
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