Journal of Traditional Chinese Medicine ›› 2023, Vol. 43 ›› Issue (1): 105-112.DOI: 10.19852/j.cnki.jtcm.2023.01.009
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Huoxue Jiedu Huayu recipe (活血解毒化瘀方) alleviates contralateral renal fibrosis in unilateral ureteral obstruction rats by inhibiting the transformation of macrophages to myofibroblast
XIONG Yunzhao1, LIU Lingjin2, LIU Ziqian2, CHEN Gege2, HAO Juan2, GAO Xiaomeng2, QIANG Panpan2, WANG Zheng3(
), XU Qingyou3()
- 1 Graduate School, Hebei University of Chinese Medicine; Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns, Hebei University of Chinese Medicine; Department of Internal Medicine, Hebei University of Chinese Medicine, Shijiazhuang 050091, China
2 Graduate School, Hebei University of Chinese Medicine; Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns, Hebei University of Chinese Medicine, Shijiazhuang 050091, China
3 Graduate School, Hebei University of Chinese Medicine; Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns, Hebei University of Chinese Medicine; Department of Internal Medicine, Hebei University of Chinese Medicine, Shijiazhuang 050091, China
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Received:2022-05-18Accepted:2022-09-14Online:2023-02-15Published:2023-01-10 -
Contact:WANG Zheng,XU Qingyou -
About author:Prof. WANG Zheng, Department of Internal Medicine, Hebei University of Chinese Medicine, Shijiazhuang 050091, China. 1017425887@qq.com. Telephone:+86-15350567700
Prof. XU Qingyou, Department of Internal Medicine, Hebei University of Chinese Medicine, Shijiazhuang 050091, China. qingyouxu@sohu.com;
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Supported by:Effect of Macrophage-to-Myofibroblast Transition in Contralateral Kidney of Unilateral Ureteral Obstruction Rats Through the Aldosterone/ MR/SGK1 Pathway and Inhibition of Chinese Herbs(81873251);Aldosterone-induced Macrophage Polarization-lymphatic Endothelial Cell-like Trans-formation Involved in Renal Fibrosis and the Protective Effect of Yiqi Jiedu Huayu Herbs(82205006);Mechanism of Renal Fibrosis Induced by NF-kB/IL-8 Induced MMT in UUO Rats(BJK2022004)
Cite this article
XIONG Yunzhao, LIU Lingjin, LIU Ziqian, CHEN Gege, HAO Juan, GAO Xiaomeng, QIANG Panpan, WANG Zheng, XU Qingyou. Huoxue Jiedu Huayu recipe (活血解毒化瘀方) alleviates contralateral renal fibrosis in unilateral ureteral obstruction rats by inhibiting the transformation of macrophages to myofibroblast[J]. Journal of Traditional Chinese Medicine, 2023, 43(1): 105-112.
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Figure 1 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal histology and fibrosis of the contralateral kidneys A-D: kidney sections from all groups were stained with HE for morphological changes (× 40). A: sham group; B: UUO group; C: EPL group; D: HJHR group. E-H: kidney sections from all groups were stained with Masson’s trichrome for fibrosis (× 40). E: sham group; F: UUO group; G: EPL group; H: HJHR group. I-L: Kidney sections from all groups were stained with immunohistochemistry staining using antibodies against α-SMA to examine renal infiltration of myofibroblasts in the kidneys (× 40). I: sham group; J: UUO group; K: EPL group; L: HJHR group. M-P: kidney sections from all groups were stained with immunohistochemistry staining using antibodies against vimentin to examine renal infiltration of myofibroblasts in the kidneys (× 40). M: sham group; N: UUO group; O: EPL group; P: HJHR group. UUO: unilateral ureteral obstruction; HE: hematoxylin-eosin; EPL: eplerenone; HJHR: Huoxue Jiedu Huayu recipe; α-SMA: α-smooth muscle actin.
Figure 1 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal histology and fibrosis of the contralateral kidneys A-D: kidney sections from all groups were stained with HE for morphological changes (× 40). A: sham group; B: UUO group; C: EPL group; D: HJHR group. E-H: kidney sections from all groups were stained with Masson’s trichrome for fibrosis (× 40). E: sham group; F: UUO group; G: EPL group; H: HJHR group. I-L: Kidney sections from all groups were stained with immunohistochemistry staining using antibodies against α-SMA to examine renal infiltration of myofibroblasts in the kidneys (× 40). I: sham group; J: UUO group; K: EPL group; L: HJHR group. M-P: kidney sections from all groups were stained with immunohistochemistry staining using antibodies against vimentin to examine renal infiltration of myofibroblasts in the kidneys (× 40). M: sham group; N: UUO group; O: EPL group; P: HJHR group. UUO: unilateral ureteral obstruction; HE: hematoxylin-eosin; EPL: eplerenone; HJHR: Huoxue Jiedu Huayu recipe; α-SMA: α-smooth muscle actin.
Figure 2 M2 macrophages are the major source of MMT in the contralateral kidneys of long-term UUO rats A-D: kidney sections from UUO group were stained with immunofluorescent co-staining of CD68 with the M2 macrophage marker CD206. A: nuclei were stained with DAPI in blue; B: macrophage marker CD68 were stained with FITC in green; C: M2 macrophage marker CD206 were stained with TRITC in red; D: immunofluorescent co-staining of CD68 (green) with the M2 macrophage marker CD206 (red). E-H: kidney sections from UUO group were stained with immunofluorescent co-staining of CD68 with the M1 macrophage marker iNOS. E: nuclei were stained with DAPI in blue; F: macrophage marker CD68 were stained with FITC in green; G: M2 macrophage marker iNOS were stained with TRITC in red; H: Immunofluorescent co-staining of CD68 (green) with the M1 macrophage marker iNOS (red). MMT: macrophage-to-myofibroblast transition; UUO: unilateral ureteral obstruction; DAPI: 4,6-diamino-2-phenyl indole; FITC: fluorescein isothiocyanate; TRITC: tetraethyl rhodamine isothiocyanate; iNOS: inducible nitric oxide synthase. All pictures are selected from the UUO group.
Figure 2 M2 macrophages are the major source of MMT in the contralateral kidneys of long-term UUO rats A-D: kidney sections from UUO group were stained with immunofluorescent co-staining of CD68 with the M2 macrophage marker CD206. A: nuclei were stained with DAPI in blue; B: macrophage marker CD68 were stained with FITC in green; C: M2 macrophage marker CD206 were stained with TRITC in red; D: immunofluorescent co-staining of CD68 (green) with the M2 macrophage marker CD206 (red). E-H: kidney sections from UUO group were stained with immunofluorescent co-staining of CD68 with the M1 macrophage marker iNOS. E: nuclei were stained with DAPI in blue; F: macrophage marker CD68 were stained with FITC in green; G: M2 macrophage marker iNOS were stained with TRITC in red; H: Immunofluorescent co-staining of CD68 (green) with the M1 macrophage marker iNOS (red). MMT: macrophage-to-myofibroblast transition; UUO: unilateral ureteral obstruction; DAPI: 4,6-diamino-2-phenyl indole; FITC: fluorescein isothiocyanate; TRITC: tetraethyl rhodamine isothiocyanate; iNOS: inducible nitric oxide synthase. All pictures are selected from the UUO group.
Figure 3 M2 macrophages are the major source of MMT in the contralateral kidneys of long-term UUO rats A-D: kidney sections from UUO group were stained with immunofluorescent multi-staining of CD68, the M2 macrophage marker CD206 and α-SMA. A: macrophage marker CD68 were stained with Alexa-405 in blue; B: M2 macrophage marker CD206 were stained with FITC in green; C: α-SMA were stained with TRITC in red; D: immunofluorescent multi -staining of CD68 (blue), CD206 (green) with the α-SMA (red). E-H: kidney sections from UUO group were stained with immunofluorescent multi-staining of CD68, the M1 macrophage marker iNOS and α-SMA. E: macrophage marker CD68 were stained with Alexa-405 in blue; F: M1 macrophage marker iNOS were stained with FITC in green; G: α-SMA were stained with TRITC in red; H: Immunofluorescent multi-staining of CD68 (blue), iNOS (green) with the α-SMA (red). MMT: macrophage-to-myofibroblast transition; UUO: unilateral ureteral obstruction; α-SMA: α-smooth muscle actin.; iNOS: inducible nitric oxide synthase; FITC: fluorescein isothiocyanate; TRITC: tetraethyl rhodamine isothiocyanate. All pictures are selected from the UUO group.
Figure 3 M2 macrophages are the major source of MMT in the contralateral kidneys of long-term UUO rats A-D: kidney sections from UUO group were stained with immunofluorescent multi-staining of CD68, the M2 macrophage marker CD206 and α-SMA. A: macrophage marker CD68 were stained with Alexa-405 in blue; B: M2 macrophage marker CD206 were stained with FITC in green; C: α-SMA were stained with TRITC in red; D: immunofluorescent multi -staining of CD68 (blue), CD206 (green) with the α-SMA (red). E-H: kidney sections from UUO group were stained with immunofluorescent multi-staining of CD68, the M1 macrophage marker iNOS and α-SMA. E: macrophage marker CD68 were stained with Alexa-405 in blue; F: M1 macrophage marker iNOS were stained with FITC in green; G: α-SMA were stained with TRITC in red; H: Immunofluorescent multi-staining of CD68 (blue), iNOS (green) with the α-SMA (red). MMT: macrophage-to-myofibroblast transition; UUO: unilateral ureteral obstruction; α-SMA: α-smooth muscle actin.; iNOS: inducible nitric oxide synthase; FITC: fluorescein isothiocyanate; TRITC: tetraethyl rhodamine isothiocyanate. All pictures are selected from the UUO group.
Figure 4 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules A: Western blots using antibodies against SGK-1, NGAL, M-CSF and NF-κB to examine the activation of MR in the kidneys; B: protein ratio of SGK-1/GAPDH; C: protein ratio of NGAL/GAPDH; D: protein ratio of M-CSF/β-actin; E: protein ratio of NF-κB/β-actin. Western blots using antibodies against SGK-1, NGAL, M-CSF and NF-κB to examine the activation of MR in the kidneys 1: Sham group; 2: UUO group; 3: EPL group; 4: HJHR group. SGK-1: serine-glucocorticoid kinase-1; NGAL: neutrophil gelatinase-associated lipocalin; M-CSF: macrophage-colony-stimulating factor; NF-κB: nuclear factor-κB; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; MR: mineralocorticoid receptor; UUO: unilateral ureteral obstruction; EPL: eplerenone; HJHR: Huoxue Jiedu Huayu recipe. Images are representative of n = 12 rats in each group. Staining quantification data are presented as the mean ± stand error of mean, n = 6. aP < 0.05 vs the sham; bP < 0.05 vs the UUO.
Figure 4 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules A: Western blots using antibodies against SGK-1, NGAL, M-CSF and NF-κB to examine the activation of MR in the kidneys; B: protein ratio of SGK-1/GAPDH; C: protein ratio of NGAL/GAPDH; D: protein ratio of M-CSF/β-actin; E: protein ratio of NF-κB/β-actin. Western blots using antibodies against SGK-1, NGAL, M-CSF and NF-κB to examine the activation of MR in the kidneys 1: Sham group; 2: UUO group; 3: EPL group; 4: HJHR group. SGK-1: serine-glucocorticoid kinase-1; NGAL: neutrophil gelatinase-associated lipocalin; M-CSF: macrophage-colony-stimulating factor; NF-κB: nuclear factor-κB; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; MR: mineralocorticoid receptor; UUO: unilateral ureteral obstruction; EPL: eplerenone; HJHR: Huoxue Jiedu Huayu recipe. Images are representative of n = 12 rats in each group. Staining quantification data are presented as the mean ± stand error of mean, n = 6. aP < 0.05 vs the sham; bP < 0.05 vs the UUO.
Table 1 Expression of SGK-1, NF-κB, MCP-1 and TGF-β1 mRNA in kidney tissue of UUO rats
| Group | n | SGK-1 mRNA | NF-κB mRNA | MCP-1 mRNA | TGF-β1 mRNA |
|---|---|---|---|---|---|
| Sham | 6 | 1.000±0.000 | 1.000±0.000 | 1.000±0.000 | 1.000±0.000 |
| UUO | 6 | 2.553±0.346a | 3.426±0.195a | 1.908±0.174a | 7.503±1.962a |
| EPL | 6 | 0.922±0.102b | 1.889±0.367b | 1.377±0.168b | 2.318±0.517b |
| HJHR | 6 | 0.964±0.322b | 1.964±0.475b | 1.445±0.071b | 5.262±0.653b |
Table 1 Expression of SGK-1, NF-κB, MCP-1 and TGF-β1 mRNA in kidney tissue of UUO rats
| Group | n | SGK-1 mRNA | NF-κB mRNA | MCP-1 mRNA | TGF-β1 mRNA |
|---|---|---|---|---|---|
| Sham | 6 | 1.000±0.000 | 1.000±0.000 | 1.000±0.000 | 1.000±0.000 |
| UUO | 6 | 2.553±0.346a | 3.426±0.195a | 1.908±0.174a | 7.503±1.962a |
| EPL | 6 | 0.922±0.102b | 1.889±0.367b | 1.377±0.168b | 2.318±0.517b |
| HJHR | 6 | 0.964±0.322b | 1.964±0.475b | 1.445±0.071b | 5.262±0.653b |
Figure S1 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal histology and fibrosis of the contralateral kidneys
Figure S1 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal histology and fibrosis of the contralateral kidneys
Figure S2 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal fibrosis of the contralateral kidneys
Figure S2 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal fibrosis of the contralateral kidneys
Figure S3 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal fibrosis of the contralateral kidneys
Figure S3 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on renal fibrosis of the contralateral kidneys
Figure S4 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules
Figure S4 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules
Figure S5 Macrophage-to-myofibroblast transition (MMT) in the contralateral kidneys of long-term UUO rats
Figure S5 Macrophage-to-myofibroblast transition (MMT) in the contralateral kidneys of long-term UUO rats
Figure S6 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules
Figure S6 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules
Figure S7 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules
Figure S7 Effects of long-term UUO and treatment with Huoxue Jiedu Huayu recipe on the expression of MR downstream molecules
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