Anti-inflammatory and Antioxidant Effects of Japanese Herbal Medicine Kyoseihatekigan on Vocal Fold Wound Healing

Published:November 23, 2021DOI:



      The Japanese herbal medicine kyoseihatekigan (KHG) has been used to alleviate the symptoms of croaky voice and globus hystericus, and each of its components has anti-inflammatory and antioxidant effects. However, the mechanisms underlying these beneficial actions of KHG on the vocal folds remain largely unknown. We examined the effects of KHG on rat vocal fold wound healing and assessed its anti-inflammatory and antioxidant properties.

      Study Design

      Animal model.


      The vocal folds of Sprague-Dawley rats were unilaterally injured under endoscopy. Rats were divided into three groups based on KHG dosing from pre injury day 4 to post injury day 3: 0 mg/kg/day (sham group), 500 mg/kg/day (1% KHG group) and 1000 mg/kg/day (2% KHG group). Histologic changes were examined to assess the degree of inflammation and oxidative stress at day 3, and fibrosis at day 56. In addition, gene expression related to pro-inflammatory cytokines and transforming growth factor-beta1 (TGF-β1) signaling was examined by quantitative real-time polymerase chain reaction (qPCR).


      Histologic analysis showed that the 1% and 2% KHG treatments significantly decreased cell infiltration and the 4-hydroxy-2-nonenalx-immunopositive area, and increased hyaluronic acid at day 3. Both KHG treatments significantly decreased fibrosis at day 56. qPCR revealed that mRNA of interleukin-1β and cyclooxygenase-2 were significantly suppressed at day 1 and TGF-β1 mRNA was significantly downregulated at day 5 in both KHG groups.


      The current findings suggest that KHG has anti-inflammatory and antioxidant effects in the early phase of vocal fold wound healing, which can lead to better wound healing with less scar formation.

      Key Words

      To read this article in full you will need to make a payment
      Subscribe to Journal of Voice
      Already a print subscriber? Claim online access
      Already an online subscriber? Sign in
      Institutional Access: Sign in to ScienceDirect


        • Kumai Y
        Pathophysiology of fibrosis in the vocal fold: current research, future treatment strategies, and obstacles to restoring vocal fold pliability.
        Int J Mol Sci. 2019; 20: 2551
        • Ohno S
        • Hirano S
        • Kanemaru S
        • et al.
        Transforming growth factor beta3 for the prevention of vocal fold scarring.
        Laryngoscope. 2012; 122: 583-589
        • Coutinho AE
        • Chapman KE
        The anti-inflammatory and immunosuppressive effects of glucocorticoids, recent developments and mechanistic insights.
        Mol Cell Endocrinol. 2011; 335: 2-13
        • Samson LD
        A target to suppress inflammation.
        Science. 2018; 362: 748-749
        • Sheppard D
        Transforming growth factor beta: a central modulator of pulmonary and airway inflammation and fibrosis.
        Proc Am Thorac Soc. 2006; 3: 413-417
        • Cui Y
        • Robertson J
        • Maharaj S
        • et al.
        Oxidative stress contributes to the induction and persistence of TGF-beta1 induced pulmonary fibrosis.
        Int J Biochem Cell Biol. 2011; 43: 1122-1133
        • Wynn TA
        • Ramalingam TR
        Mechanisms of fibrosis: therapeutic translation for fibrotic disease.
        Nat Med. 2012; 18: 1028-1040
        • Ingle JW
        • Helou LB
        • Li NY
        • et al.
        Role of steroids in acute phonotrauma: a basic science investigation.
        Laryngoscope. 2014; 124: 921-927
        • Mizuta M
        • Hirano S
        • Hiwatashi N
        • et al.
        Effect of astaxanthin on vocal fold wound healing.
        Laryngoscope. 2014; 124: E1-E7
        • Li X
        • Zhu L
        • Wang B
        • et al.
        Drugs and targets in fibrosis.
        Front Pharmacol. 2017; 8: 855
        • Liu C
        • Yuan X
        • Tao L
        • et al.
        Xia-yu-xue decoction (XYXD) reduces carbon tetrachloride (CCl4)-induced liver fibrosis through inhibition hepatic stellate cell activation by targeting NF-kappaB and TGF-beta1 signaling pathways.
        BMC Complement Altern Med. 2015; 15: 201
        • Veilleux MP
        • Moriyama S
        • Yoshioka M
        • et al.
        A review of evidence for a therapeutic application of traditional Japanese Kampo medicine for oral diseases/disorders.
        Med (Basel). 2018; 5: 35
        • Yu F
        • Takahashi T
        • Moriya J
        • et al.
        Traditional Chinese medicine and Kampo: a review from the distant past for the future.
        J Int Med Res. 2006; 34: 231-239
        • Kimura I
        • Kagawa S
        • Nishimura K
        Self-administered rhubarb-free kyoseihatekigan has novel anti-asthma effects: two case reports.
        Traditional Kampo Med. 2017; 4: 58-61
        • Takeno Y
        Clinical effect of STR-1 on throat discomfort and hoarseness.
        J New Rem Clin. 1992; 41: 2311-2316
        • Branski RC
        • Verdolini K
        • Rosen CA
        • et al.
        Acute vocal fold wound healing in a rabbit model.
        Ann Otol Rhinol Laryngol. 2016; 114: 19-24
        • Mizuta M
        • Hirano S
        • Ohno S
        • et al.
        Expression of reactive oxygen species during wound healing of vocal folds in a rat model.
        Ann Otol Rhinol Laryngol. 2012; 121: 804-810
        • Tateya T
        • Tateya I
        • Sohn JH
        • et al.
        Histological study of acute vocal fold injury in a rat model.
        Ann Otol Rhinol Laryngol. 2006; 115: 285-292
        • Kawai S
        • Takagi Y
        • Kaneko S
        • et al.
        Effect of three types of mixed anesthetic agents alternate to ketamine in mice.
        Exp Anim. 2011; 60: 481-487
        • Chang Z
        • Ling C
        • Yamashita M
        • et al.
        Microarray-driven validation of reference genes for quantitative real-time polymerase chain reaction in a rat vocal fold model of mucosal injury.
        Anal Biochem. 2010; 406: 214-221
        • Dharmananda S
        Treatment of throat and voice disorders with Chinese medicine.
        ITM, 2002
        • Jung HW
        • Mahesh R
        • Lee JG
        • et al.
        Pinoresinol from the fruits of Forsythia koreana inhibits inflammatory responses in LPS-activated microglia.
        Neurosci Lett. 2010; 480: 215-220
        • Kim JY
        • Hwang YP
        • Kim DH
        • et al.
        Inhibitory effect of the saponins derived from roots of Platycodon grandiflorum on carrageenan-induced inflammation.
        Biosci Biotechnol Biochem. 2006; 70: 858-864
        • Gong G
        • Yuan LB
        • Hu L
        • et al.
        Glycyrrhizin attenuates rat ischemic spinal cord injury by suppressing inflammatory cytokines and HMGB1.
        Acta Pharmacol Sin. 2012; 33: 11-18
        • Choi YA
        • Choi JK
        • Jang YH
        • et al.
        Antiinflammatory effect of Amomum xanthioides in a mouse atopic dermatitis model.
        Mol Med Rep. 2017; 16: 8964-8972
        • Sazwi NN
        • Nalina T
        • Abdul Rahim ZH
        Antioxidant and cytoprotective activities of Piper betle, Areca catechu, Uncaria gambir and betel quid with and without calcium hydroxide.
        BMC Complement Altern Med. 2013; 13: 351
        • Bag A
        • Kumar Bhattacharyya S
        • Kumar Pal N
        • et al.
        Anti-inflammatory, anti-lipid peroxidative, antioxidant and membrane stabilizing activities of hydroalcoholic extract of Terminalia chebula fruits.
        Pharm Biol. 2013; 51: 1515-1520
        • Trevisan SCC
        • Menezes APP
        • Barbalho SM
        • et al.
        Properties of mentha piperita: a brief review.
        World J Pharm Med Res. 2017; 3: 309-313
        • Asl MN
        • Hosseinzadeh H
        Review of pharmacological effects of Glycyrrhiza sp. and its bioactive compounds.
        Phytother Res. 2008; 22: 709-724
        • Kai H
        • Mori T
        • Tokuda K
        • et al.
        Pressure overload-induced transient oxidative stress mediates perivascular inflammation and cardiac fibrosis through angiotensin II.
        Hypertens Res. 2006; 29: 711-718
        • Kolb M
        • Margetts PJ
        • Anthony DC
        • et al.
        Transient expression of IL-1beta induces acute lung injury and chronic repair leading to pulmonary fibrosis.
        J Clin Invest. 2001; 107: 1529-1536
        • Hansen JK
        • Thibeault SL
        • Walsh JF
        • et al.
        In vivo engineering of the vocal fold extracellular matrix with injectable hyaluronic acid hydrogels: early effects on tissue repair and biomechanics in a rabbit model.
        Ann Otol Rhinol Laryngol. 2005; 114: 662-670
        • Sato H
        • Takahashi T
        • Ide H
        • et al.
        Antioxidant activity of synovial fluid, hyaluronic acid, and two subcomponents of hyaluronic acid. Synovial fluid scavenging effect is enhanced in rheumatoid arthritis patients.
        Arthritis Rheumatism. 1988; 31: 63-71
        • Ohno T
        • Hirano S
        • Rousseau B
        Gene expression of transforming growth factor-beta1 and hepatocyte growth factor during wound healing of injured rat vocal fold.
        Laryngoscope. 2009; 119: 806-810
        • Paul BC
        • Rafii BY
        • Gandonu S
        • et al.
        Smad3: an emerging target for vocal fold fibrosis.
        Laryngoscope. 2014; 124: 2327-2331
        • Yao QY
        • Xu BL
        • Wang JY
        • et al.
        Inhibition by curcumin of multiple sites of the transforming growth factor-beta1 signalling pathway ameliorates the progression of liver fibrosis induced by carbon tetrachloride in rats.
        BMC Complement Altern Med. 2012; 12: 156
        • Hirano S
        • Bless DM
        • Rousseau B
        • et al.
        Fibronectin and adhesion molecules on canine scarred vocal folds.
        Laryngoscope. 2003; 113: 966-972
        • Hiwatashi N
        • Hirano S
        • Mizuta M
        • et al.
        The efficacy of a novel collagen-gelatin scaffold with basic fibroblast growth factor for the treatment of vocal fold scar.
        J Tissue Eng Regen Med. 2017; 11: 1598-1609
        • Tateya T
        • Tateya I
        • Sohn JH
        • et al.
        Histologic characterization of rat vocal fold scarring.
        Ann Otol Rhinol Laryngol. 2005; 114: 183-191
        • Lim X
        • Tateya I
        • Tateya T
        • et al.
        Immediate inflammatory response and scar formation in wounded vocal folds.
        Ann Otol Rhinol Laryngol. 2006; 115: 921-929
        • Morisaki T
        • Kishimoto Y
        • Tateya I
        • et al.
        Adipose-derived mesenchymal stromal cells prevented rat vocal fold scarring.
        Laryngoscope. 2018; 128: E33-E40