We collected the retrospective medical records of 22 vocal fold nodule patients treating with TCM from July 2018 to August 2020. We found that the most used herbs were P. grandiflorus and licorice. Further, we used the two herbs for porcine VFE culture. And the two herbs significantly inhibited the P38, ERK, and JNK pathways in TNF-alpha induced inflammation.
The finished herbal products (FHP) we prescribed are granulated compounds concentrated from Chinese herbal remedies, including single herbs and herbal formulae.(24) They are officially approved in the national healthcare systems not only in Taiwan, but also in Japan, Korea and mainland China. They are widely used in the East Asia. (25)
In our retrospective analysis, patients regularly return to clinic in a two-week to one-month manner. Since our clinic is comparatively remote, most of our patients found that returning to TCM clinic is more convenient than attending voice therapy. From Fig. 2, we can see the most used herbs were Licorice and P. grandifloras. The total amount were more than 2000 grams.
Glycyrrhiza glabra, the licorice species we used here, is brought from Chiayi Chang Gung Memorial Hospital (Taiwan). G. glabra is grown in Eurasia, northern Africa and western Asia.(26) Its rhizomes and roots are the most important medicinal parts, which have been reported to be used alone or in combination with other herbs for treating multiple diseases such as digestive disorders, respiratory tract disorders, epilepsy, fever, sexual disability, and etc. (27) P. grandiflorus contains numerous amino acids, vitamins, and trace elements. Also, it has the ability to treat the following disorders: cough and asthma relief, antitumor activity, anti-inflammatory and antibacterial effects, antioxidation, hypoglycemic effects, liver protection, and immunity enhancement (28). Both Glycyrrhiza glabra and P. grandiflorus can be used as food and flavoring agents.
The combined use of these two herbs can be traced to ancient China. In the Chinese classic Shang Han Lun (English title: Treatise on Cold Damage Diseases, written by ZHANG Zhong-jing, 150–219 A.D.), the two herbs were combined in a 2:1 ratio and were used mainly to treat sore throat and Fei Yon (pulmonary abscess) (29, 30). Pharmacologically, they can affect the metabolic process of each other, improving the bioavailability of their compounds (31, 32).
Clinically, inflammation is commonly observed in patients with chronic voice disorders, such as vocal fold nodules. For example, an immediate increase in inflammatory cytokines, including TNF-α, is observed in response to vocal fold injury. The effect of TNF-α on the vocal fold epithelium upregulates mucin expression (16). TNF-α may activate the NF-κB p65 pathway in PVFE cells, but this inflammatory pathway is inhibited by the corticosteroid dexamethasone(Dex) and high doses of G. glabra and P. grandiflorus. This means that the anti-inflammatory effect of TCM herbs on the vocal folds depends on their accumulation in vocal epithelium cells.
MAPK families are critical in complex cellular programs like proliferation, differentiation, development, transformation, and apoptosis. The MAPK pathways involve a series of protein kinase cascades, which play a crucial role in the regulation of cell proliferation. The three major MAPK families, namely mammalian ERKs, c-JNK, and p38 kinases, are activated by many stimuli. JNK and p38 are activated by cytokines and stressors, and ERK is mainly activated by growth factors (33). Figure 3C shows that G. glabra and P. grandiflorus deregulated the activation of p38, p-ERK, and p-JNK. This result may be explained by the beneficial effects of G. glabra and P. grandiflorus on vocal nodules due to downregulation of the MAPK pathway (34).
In our prescription pattern, a 1:1 ratio of G. glabra to P. grandiflorus may be more beneficial for VFN patients. Our sample size is relatively small. Also, the dosage and duration varied, which may be due to individual difference. For in vitro study, though the above inflammatory pathway down-regulation was seen, whether there exists synergistic effect or other interaction between the herbs are still unknown. Further prospective and functional studies are needed to confirm the results of this study and clarify the mechanism of action.
In conclusion, by reviewing clinical patient data and conducting an in vitro study, we confirmed that TCM herbal treatment is an alternative treatment option for VFNs. Patients’ voices exhibited improvement after daily TCM herb use, and VFNs were reduced. From the above results, we may imply that our most prescribed herbs G. glabra and P. grandiflorus as potential VFN alternative treatment.