Elsevier

Biomedicine & Pharmacotherapy

Volume 95, November 2017, Pages 520-528
Biomedicine & Pharmacotherapy

Original article
Erhuang Formula ameliorates renal damage in adenine–induced chronic renal failure rats via inhibiting inflammatory and fibrotic responses

https://doi.org/10.1016/j.biopha.2017.08.115Get rights and content

Abstract

Aims

The present study aimed to evaluate the protective effects of Erhuang Formula (EHF) and explore its pharmacological mechanisms on adenine-induced chronic renal failure (CRF).

Materials and methods

The compounds in EHF were analyzed by HPLC/MS. Adenine-induced CRF rats were administrated by EHF. The effects were evaluated by renal function examination and histology staining. Immunostaining of some proteins related cell adhesion was performedin renal tissues, including E-cadherin, β-catenin, fibronectin and laminin. The qRT-PCR was carried out determination of gene expression related inflammation and fibrosis including NF-κB, TNF-α, TGF-β1, α-SMA and osteopontin (OPN).

Results

Ten compounds in EHF were identified including liquiritigenin, farnesene, vaccarin, pachymic acid, cycloastragenol, astilbin, 3,5,6,7,8,3′,4′-heptemthoxyflavone, physcion, emodin and curzerene. Abnormal renal function and histology had significant improvements by EHF treatment. The protein expression of β-catenin, fibronectin and laminin were significantly increased and the protein expression of E-cadherin significantly decreased in CRF groups. However, these protein expressions were restored to normal levels in EHF group. Furthermore, low expression of PPARγ and high expression of NF-κB, TNF-α, TGF-β1, α-SMA and OPN were substantially restored by EHF treatment in a dose-dependent manner.

Conclusions

EHF ameliorated renal damage in adenine-induced CRF rats, and the mechanisms might involve in the inhibition of inflammatory and fibrotic responses and the regulation of PPARγ, NF-κB and TGF-β signaling pathways.

Introduction

Chronic renal failure (CRF) is a common consequence in a variety of chronic kidney diseases (CKD), which has become an important public health problem in China [1]. The progression of CRF is characterized by the development of glomerular and tubular lesions in which multiple factors can be involved [2]. Although much is known about the progression of CRF, there are not any effective treatments to hold back or treat CRF. Therefore, the development of successful therapeutic strategies remains a clinical challenge [3].

As we know, Traditional Chinese medicines (TCMs) had a long history dating back several thousands of years for clinical application and the discovery of new drugs [4], [5], [6]. Many TCMs possessed long time clinical application as well as beneficial and reliable therapeutic efficacy and they are attracting increased global attention for the treatment of CRF [7], [8], [9], [10], [11], [12]. Chinese herbal formula (CHF), a herbal mixture prescription, has been used for the treatment of various of diseases and syndromes [13]. Many previous studies focused on the renal protective effects of CHF [14], [15], [16]. Erhuang Formula (EHF) is a classical formula for treating kidney injury, which is composed of Astragali radix, Rhei radix etrhizoma, Trigonellae semen, Achyranthis bidentatae radix, Vaccariae semen, Smilacis glabrae rhizoma, Curcumae rhizome (Table 1). EHF has an obvious improvement for CRF patients, and it has been reported that EHF exerted significant protective effects for the treatment of renal fibrosis in CRF rats [17]. However, the mechanisms of its pharmacological effects are still unclear.

Accumulating evidence indicates that inflammatory and fibrotic responses play a significant role in CRF progression and development [18]. Activated nuclear factor-kappaB signaling pathway and pro-inflammatory cytokine tumor necrosis factor (TNF-α) and pro-fibrogenic cytokine transforming growth factor beta 1 (TGF-β1) contribute to the damage of renal failure [19], [20]. Inflammatory stimuli have been found to active resident fibroblasts, and inhibition of inflammation attenuated profibrogenic gene expression and tubulointerstitial fibrosis in kidney diseases. Furthermore, an overview of kidney diseases shows that complementary but different mechanisms are responsible for fibrosis [21]. The adenine-induced CRF model was first reported in rats about thirty years ago [22]. This model was widely used for inflammation and metabolism dysregulation–associated action mechanism of CRF [23], [24] and efficacy evaluation and therapeutic mechanism of TCMs such as Poria cocos [25], [26], [27], Rheum officinale [28], [29], [30], [31] and Polyporus umbellatus [32], [33], [34]. This kind of CRF model can provide valuable information of pathological mechanisms for various complications in a persistent uremic state, and long-term feeding adenine to rats could cause metabolic abnormalities similar to the CKD symptoms in humans [35]. Therefore, in the present study, the adenine-induced CRF model was used to evaluate the protective effects of EHF and explore its pharmacological mechanisms.

Section snippets

Materials

Preparation of Erhuang Formula: EHF was prepared and purchased from Shanghai Chinese Medicine Pharmaceutical Technology Ltd. (Shanghai, China), and the preparation method was as follows. The air-dried herbs identified by authority were powdered and subjected to reflux extraction with 10 times water for 1 h. The aqueous extracts were filtered and collected. The extraction was repeated twice with the method introduced above, and then all the extracts were evaporated to dryness under reduced

Identification of ten compounds in EHF

The aqueous extract of mixed ten Chinese medicinal materials was measured by HPLC/ESI–MS in negative and positive-ion mode (Fig. 1A-L). Ten compounds 1  10, with the retention time at 3.59 min, 4.73 min, 4.94 min, 5.01 min, 5.42 min, 5.95 min, 6.92 min, 8.89 min, 10.18 min, and 10.25 min, were identified as liquiritigenin (1), farnesene (2), vaccarin (3), pachymic acid (4), cycloastragenol (5), astilbin (6), 3,5,6,7,8,3′,4′-heptemthoxyflavone (7), physcion (8), emodin (9), and curzerene (10), on the basis

Discussion

The effective therapy of CRF remains a major unmet clinical medical need. Adenine produces metabolic abnormalities resembling chronic renal insufficiency in humans [22], [36]. The present study showed the protective effect of EHF on CRF rats induced by adenine. We examined renal function including Scr and BUN and histology. Significantly increased Scr and BUN concentrations are observed in our study, which are consistent with the previous publications [37], [38], [39]. Furthermore, histological

Conflict of interests

The authors have declared that there is no conflict of interests.

Acknowledgments

This work was supported by National Natural Science Foundation of China (grant number 81503332), Shanghai Municipal Health and Family Planning Commission (grant number ZY3-JSFC-2-2010, 20154Y0063), Outstanding Leaders Training Program of Pudong Health Bureau of Shanghai (grant number PWR12015-05), Excellent Youth Medical Talents Training Program of Pudong Health Bureau of Shanghai (grant number PWRq2015-11), and the Open Research Fund of State Key Laboratory Breeding Base of Systematic

References (65)

  • Y.Y. Zhao

    UPLC-Q-TOF/HSMS/MS(E)-based metabonomics for adenine-induced changes in metabolic profiles of rat faeces and intervention effects of ergosta-4,6,8(14),22-tetraen-3-one

    Chem. Biol. Interact.

    (2013)
  • H. Chen

    Traditional uses, fermentation, phytochemistry and pharmacology of Phellinus linteus: a review

    Fitoterapia

    (2016)
  • Y.N. Song

    Huangqi decoction alleviates dimethylnitrosamine-induced liver fibrosis: an analysis of bile acids metabolic mechanism

    J. Ethnopharmacol.

    (2016)
  • Y. Tu

    Dahuang Fuzi Decoction ameliorates tubular epithelial apoptosis and renal damage via inhibiting TGF-beta1-JNK signaling pathway activation in vivo

    J. Ethnopharmacol.

    (2014)
  • L. Wang

    You-gui Pill ameliorates renal tubulointerstitial fibrosis via inhibition of TGF-beta/Smad signaling pathway

    J. Ethnopharmacol.

    (2015)
  • Y.Y. Zhao

    Renal metabolic profiling of early renal injury and renoprotective effects of Poria cocos epidermis using UPLC Q-TOF/HSMS/MSE

    J. Pharm. Biomed. Anal.

    (2013)
  • Y.Y. Zhao

    Urinary metabonomic study of the surface layer of Poria cocos as an effective treatment for chronic renal injury in rats

    J. Ethnopharmacol.

    (2013)
  • Q. Zhang

    Rhein reverses Klotho repression via promoter demethylation and protects against kidney and bone injuries in mice with chronic kidney disease

    Kidney Int.

    (2017)
  • Y.Y. Zhao

    Effect of ergosta-4,6,8(14),22-tetraen-3-one (ergone) on adenine-induced chronic renal failure rat: a serum metabonomic study based on ultra performance liquid chromatography/high-sensitivity mass spectrometry coupled with MassLynx i-FIT algorithm

    Clin. Chim. Acta

    (2012)
  • J. Wang

    Effect and mechanism of fucoidan derivatives from Laminaria japonica in experimental adenine-induced chronic kidney disease

    J. Ethnopharmacol.

    (2012)
  • Y.Y. Zhao

    Urinary metabonomics study on biochemical changes in an experimental model of chronic renal failure by adenine based on UPLC Q-TOF/MS

    Clin. Chim. Acta

    (2012)
  • A. Chawla

    A PPAR gamma-LXR-ABCA1 pathway in macrophages is involved in cholesterol efflux and atherogenesis

    Mol. Cell

    (2001)
  • C. Guijarro et al.

    Transcription factor-kappa b (NF-kappa B) and renal disease

    Kidney Int.

    (2001)
  • D.Q. Chen

    Gene and protein expressions and metabolomics exhibit activated redox signaling and wnt/β-catenin pathway are associated with metabolite dysfunction in patients with chronic kidney disease

    Redox Biol.

    (2017)
  • H. Chen

    Metabolomics insights into activated redox signaling and lipid metabolism dysfunction in chronic kidney disease progression

    Redox Biol.

    (2016)
  • J.M. Fan

    Transforming growth factor-beta regulates tubular epithelial-myofibroblast transdifferentiation in vitro

    Kidney Int.

    (1999)
  • L. Chen

    Role of RAS/Wnt/beta-catenin axis activation in the pathogenesis of podocyte injury and tubulo-interstitial nephropathy

    Chem. Biol. Interact.

    (2017)
  • V.A. Stevens

    The interdependence of EGF-R and SGK-1 in fibronectin expression in primary kidney cortical fibroblast cells

    Int. J. Biochem. Cell Biol.

    (2007)
  • J.R. Lu

    Protective effects of bu-Shen-Huo-Xue formula against 5/6 nephrectomy-Induced chronic renal failure in rats

    Evid. Based Complement. Alternat. Med.

    (2014)
  • H.X. Pan

    Protective effect of the Erhuang compound remedy on the rats with kidney subtotal excision

    Anhui Med. Pharm. J.

    (2015)
  • Y.Y. Zhao

    Metabolomics analysis reveals the association between lipid abnormalities and oxidative stress, inflammation, fibrosis, and Nrf2 dysfunction in aristolochic acid-induced nephropathy

    Sci. Rep.

    (2015)
  • M.L. Kielar

    Docosahexaenoic acid ameliorates murine ischemic acute renal failure and prevents increases in mRNA abundance for both TNF-alpha and inducible nitric oxide synthase

    J. Am. Soc. Nephrol.

    (2003)
  • Cited by (23)

    • Flavonoids and triterpenes isolated from Eucryphia cordifolia (Cunoniaceae)

      2022, Biochemical Systematics and Ecology
      Citation Excerpt :

      Cunoniaceae) (Barrientos et al., 2020). Astilbine-containing plants and herbs were traditionally used in China and some other Asian countries in the treatment of various disorders such as hyperuricemia (Sun et al., 2015), chronic renal failure (Zhang et al., 2017), immunosuppression (Guo et al., 2007). Catechin (6) was identified in the aerial parts of E. cordifolia (Bridi et al., 2019), and it has also been observed in the fruits of Davidsonia pruriens and Davidsonia jerseyana F. Muell (Cunoniaceae) (Nirmal et al., 2021).

    • Renoprotective effects of enzyme-hydrolyzed polysaccharides from Auricularia polytricha on adenine-induced chronic kidney diseases in mice

      2021, Biomedicine and Pharmacotherapy
      Citation Excerpt :

      Furthermore, more and more studies have indicated that NF-κB-related inflammatory pathways may play a pivotal role in many nephropathies [8,27]. Oxidative stress could trigger inflammatory reactions in CKD animals by activating NF-κB in the damaged kidney, which could be evidenced by increased translocation of p65 [7,13]. Briefly, NF-κB is a transcription factor promoting the expression of many inflammatory genes, such as TNF-α and IL-1β [18].

    • ASTILBIN: A PROMISING UNEXPLORED COMPOUND WITH MULTIDIMENSIONAL MEDICINAL AND HEALTH BENEFITS

      2020, Pharmacological Research
      Citation Excerpt :

      These studies include the effect of astilbin isolated from Smilax china and Rhododendron oldhamii leaves directly in renal injury induced by potassium oxonate [84,87] and fructose (10%) [17]. Also, effects of different extracts from various plants were evaluated like smilax glabra in potassium oxonate [96] and gentamicin [52] induced nephropathy and Erhuang Formula (a traditionally used combination of herbs in China) in renal failure induced by adenine [101]. In these studies in which effects were examined using plant extracts, it was indicated that astilbin is one of the main active component.

    • Evaluation of renoprotective effect of Chinese chive extracts on adenine-induced chronic renal failure

      2018, Food Science and Human Wellness
      Citation Excerpt :

      Thus, it was further confirmed that CCWE was the main renoprotective fraction of Chinese chive. The levels of serum SCr and BUN were considered as the important index to reflect the renal filtration function [18–20]. To further evaluate their renoprotective function, the effects of CCWE and CCEE on the serum SCr and BUN levels of CRF mice were investigated.

    View all citing articles on Scopus
    View full text