Abstract: Hepatitis B virus is a hepatotropic DNA virus that can cause acute or chronic hepatitis B. Hepatitis B has now become a global public health concern, with a high risk of progressing to liver cirrhosis, liver failure, and even hepatocellular carcinoma, thereby posing a serious threat to human health. Traditional Chinese medicines and their bioactive constituents exhibit significant anti–hepatitis B viral activity; this review summarizes the anti–hepatitis B viral activities of these medicinal agents and their underlying mechanisms, with the aim of providing a reference for future research in this field.

The hepatitis B virus is a hepatotropic DNA virus that can cause acute or chronic hepatitis B. Globally, approximately 3.5% of the population is currently infected with chronic hepatitis B. If left untreated, chronic hepatitis B can readily progress to liver cirrhosis, liver failure, and even hepatocellular carcinoma, posing a serious threat to human health. ^[1] Currently, the antiviral drugs commonly used in clinical practice for hepatitis B fall into two main categories: nucleos(t)ide analogs and interferons. The former includes drugs such as entecavir and lamivudine; however, long-term use of these agents readily leads to drug resistance, and abrupt discontinuation can result in a severe rebound phenomenon. In contrast, the latter has a narrow range of indications, significant adverse effects, requires parenteral administration, and is relatively expensive. ^[2] Therefore, there is an urgent need to develop new anti-hepatitis B virus drugs for the current treatment of hepatitis B.

Traditional Chinese medicines and their bioactive constituents exert significant anti-hepatitis B virus activity due to their unique advantages, holding great promise for clinical application. In this review, the author has examined relevant literature on the anti-hepatitis B virus effects of traditional Chinese medicines over the past five years and summarized the latest advances in research on their antiviral activity and mechanisms of action, with the aim of providing a reference for future studies on traditional Chinese medicines as anti-hepatitis B virus agents.

1 Antiviral Effects of Active Ingredients from Single Traditional Chinese Medicinal Herbs Against Hepatitis B Virus

1.1 Artemisia ragweed

Artemisia capillaris is derived from the Asteraceae plant Artemisia scoparia. Artemisia scoparia Waldst . and Kit. or Artemisia capillaris Artemisia capillaris Thunb., with the effects of clearing and draining damp-heat, promoting bile flow, and resolving jaundice. ^[3] Its anti-hepatitis B virus active constituents mainly include glucosides, sesquiterpenes, chlorogenic acid, p-hydroxyacetophenone, and enynes. The Geng research group investigated the anti-hepatitis B virus activity of various Artemisia capillaris extracts in HepG2.2.15 hepatocellular carcinoma cells and found that three glucosides exhibited potent inhibitory effects on hepatitis B virus DNA replication as well as on the secretion of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg), with half-maximal inhibitory concentrations (IC ₅₀ ) are all less than 3.00 μmol/L; eucalyptane sesquiterpenes exhibit strong inhibitory activity against hepatitis B virus DNA replication as well as the secretion of HBsAg and HBeAg, with IC ₅₀ The respective values are 12.01, 15.02, and 9.00 μmol/L; chlorogenic acid and its analogs, p-hydroxyacetophenone, enynes, and other compounds exhibit strong inhibitory activity against hepatitis B virus DNA replication, with IC ₅₀ All were less than 15.00 μmol/L, and exhibited mild inhibitory activity against the secretion of HBsAg and HBeAg. ^[4-8] . The aforementioned Artemisia capillaris extract exhibits significant in vitro anti-hepatitis B virus activity, laying the foundation for further mechanistic studies and the development of anti-hepatitis B virus drugs.

1.2 Polygonum cuspidatum

Polygonum cuspidatum is harvested from the Polygonaceae plant Polygonum cuspidatum. Polygonum cuspidatum Sieb. et Zucc., with effects such as promoting diuresis to resolve jaundice, clearing heat and detoxifying, dispersing blood stasis to relieve pain, and more. ^[3] , its bioactive components with anti-hepatitis B virus activity mainly include quercetin and resveratrol, among others. Cheng et al. ^[9] Upon treatment of HepG2.2.15 cells and Huh7 cells transfected with pCH-9/3091 (which harbors a 1.1-fold genome of the hepatitis B virus) with quercetin, the secretion of HBsAg and HBeAg as well as the replication of hepatitis B virus DNA were markedly inhibited; moreover, this inhibitory activity was further enhanced when quercetin was used in combination with lamivudine, adefovir, entecavir, and other antiviral agents. The underlying mechanism may involve quercetin-mediated downregulation of the expression of heat shock proteins (HSPs) such as Hsp40, Hsp70, and Hsp90-β, thereby suppressing hepatitis B virus transcription. Parvez et al. ^[10] After continuous treatment of HepG2.2.15 cells with quercetin for 5 days, the secretion of HBsAg and HBeAg in the culture supernatant was markedly inhibited; however, prolonged treatment resulted in excessive cell proliferation and cell death. Molecular docking results indicated that quercetin exhibits strong binding affinity to hepatitis B virus polymerase (Pol). Park et al. ^[11] Treatment of Huh7-HBx cells, which stably express the hepatitis B virus X protein (HBx), with resveratrol disrupts serine–threonine protein kinase (Akt) signaling, reduces cyclin D1 expression, and thereby inhibits cell proliferation. In vivo studies using a xenograft mouse model bearing Huh7-HBx cells further confirmed resveratrol’s antitumor activity, suggesting that resveratrol may have the potential to reduce the risk of hepatocellular carcinoma in patients with chronic hepatitis B virus infection. However, Shi et al. ^[12] According to reports, resveratrol can activate the Sirt1–PGC-1α–PPARα signaling axis, thereby enhancing the activity of the hepatitis B virus core promoter, which in turn promotes viral transcription and replication and leads to a worsening of hepatitis B symptoms. The dual nature of resveratrol’s effects on the hepatitis B virus as reported in the literature means that its potential as an antiviral compound against hepatitis B still requires further investigation.

1.3 Salvia miltiorrhiza

Danshen is derived from the Lamiaceae plant Danshen. Salvia miltiorrhiza The dried roots and rhizomes of Bge. exhibit excellent hepatoprotective effects. ^[3] Danshensu, rosmarinic acid, shikonin, and other compounds are the main bioactive constituents of Salvia miltiorrhiza with anti-hepatitis B virus activity. Duan Shupeng et al. ^[13] Studies have shown that treatment of HepG2.2.15 cells with danshensu reduces hepatitis B virus reverse transcriptase activity, thereby directly inhibiting HBV DNA replication and consequently suppressing the expression of HBsAg and HBeAg. Tsukamoto et al. ^[14] The report demonstrates that rosmarinic acid exerts a significant inhibitory effect on the secretion of hepatitis B virus DNA in the culture supernatants of Hep38.7-Tet cells and primary human hepatocytes infected with hepatitis B virus, while exerting a mild inhibitory effect on the secretion of HBsAg. The underlying mechanism may involve interference with the binding of hepatitis B virus pregenomic RNA (pgRNA) to the viral polymerase. In addition, the authors’ study found that the polyphenolic compound shikonin from Salvia miltiorrhiza, when applied to HepG2.2.15 cells, markedly suppresses hepatitis B virus DNA replication. The mechanism is likely mediated by shikonin upregulating the expression of lysosome-associated membrane proteins 1 and 2 (LAMP1/2) and cathepsin D (Cath-D), thereby enhancing lysosomal acidification; furthermore, shikonin inhibits the PI3K/Akt/mTOR signaling pathway, promoting autophagosome formation and ultimately inducing complete autophagy in hepatocytes, which in turn disrupts hepatitis B virus replication.

1.4  Sophora flavescens

Sophora flavescens is a leguminous plant. Sophora flavescens Dried roots of Ait. ^[3] Its anti-hepatitis B virus active components mainly include matrine, oxymatrine, sophocarpine and other matrine-type alkaloids, as well as sophora polysaccharides. Feng Jing et al. ^[15] Studies have shown that matrine exhibits potent inhibitory effects on hepatitis B virus DNA replication in HepG2.2.15 cells, with the mechanism likely involving suppression of polymerase activity; furthermore, its combination with lamivudine can enhance the latter’s antiviral efficacy against hepatitis B virus. Zhang Xuan et al. ^[16] and Sang et al. ^[17] It was found that oxymatrine significantly inhibits the secretion of HBsAg and HBeAg, as well as the replication of hepatitis B virus DNA, in the culture supernatant of HepG2.2.15 cells and in the serum of mice receiving hydrodynamic injection of pAAV-HBV 1.2. Chen et al. ^[18] The report indicates that treatment of HepG2.2.15 cells with sophoridine for 72 hours significantly inhibits the secretion of HBsAg and HBeAg as well as the replication of hepatitis B virus DNA, with greater inhibitory activity than matrine and oxymatrine. Yang et al. ^[19] Studies have shown that matrine polysaccharides significantly inhibit the secretion of HBsAg and HBeAg by HepG2.2.15 cells, with the underlying mechanism likely involving the regulation of inflammatory cytokine secretion, including interferon-γ (IFN-γ), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α).

1.5 Other single-herb active extracts

In addition to the single-herb Chinese medicinal extracts mentioned above, a large number of other Chinese medicinal extracts have been found in recent years to exhibit anti-hepatitis B virus activity, thereby providing an extensive library of potential bioactive compounds for the development of anti-hepatitis B virus drugs. Parvez et al. ^[20] Research has demonstrated that aloe-emodin, an extract from aloe, inhibits the secretion of HBsAg and HBeAg as well as the replication of hepatitis B virus in HepG2.2.15 cells, with the underlying mechanism likely involving suppression of polymerase activity. Furthermore, the findings indicate that aloe-emodin exhibits antiviral activity comparable to that of lamivudine, possibly due to its ability to activate cytochrome P450 enzymes. In our own study, we found that protocatechuic acid suppresses the transcription and replication of hepatitis B virus by downregulating hepatocyte nuclear factors HNF-4α and HNF-1α via activation of the ERK1/2 signaling pathway, thereby modulating the activity of the viral X promoter, core promoter, and pre-S1 promoter. ^[21] Furthermore, the combined use of protocatechuic acid and lamivudine significantly enhances their inhibitory effects on the secretion of hepatitis B virus DNA, HBsAg, HBeAg, and other markers, prolongs the duration of these effects, and exerts a favorable hepatoprotective effect. ^[22] 。

The active constituents and extracts of individual Chinese medicinal herbs, including Yinchen, Polygonum cuspidatum, Salvia miltiorrhiza, and Sophora flavescens, have been reported to exhibit anti-hepatitis B virus activity; see Table 1.

2 Antiviral Effects of Traditional Chinese Medicine Compound Formulas Against Hepatitis B Virus

2.1 Compound Phyllanthus urinaria

Compound Herba Phyllanthi is a traditional Chinese medicine formula composed of Herba Phyllanthi, Astragalus, Panax notoginseng, Glycyrrhiza, and other herbs. Li et al. ^[23] The report indicates that treatment with the compound formulation of Phyllanthus urinaria significantly inhibits the proliferation, migration, and colony-forming ability of HepG2-HBx cells—cells stably expressing HBx—as well as the growth of tumors in mice xenografted with these cells. The underlying mechanism is likely related to the inactivation of the HBx–SHH signaling axis, suggesting that this compound formulation holds therapeutic potential for attenuating the progression of hepatitis B to hepatocellular carcinoma. Tong et al. ^[24] A total of 102 hepatitis B patients with hepatitis B–related cirrhosis accompanied by regenerative or dysplastic nodules were selected as the study subjects, and it was found that the compound formulation of Folium Phyllanthi could to a certain extent inhibit hepatitis B virus DNA replication, thereby preventing or delaying the progression of hepatitis B–related liver disease toward hepatocellular carcinoma. Tan Benren et al. ^[25] The study found that Fufang Yexiazhu Dropping Pills exerted a certain degree of inhibitory effect on HBsAg, HBeAg, and hepatitis B virus DNA replication in the culture supernatants of HepG2.2.15 cells as well as in serum samples from HBsAg-positive patients.

2.2  Xiao Chaihu Tang

Xiao Chaihu Tang originates from the “Shanghan Zabu Lun” and is composed of seven herbal ingredients: Bupleurum, Scutellaria, Ginseng, Pinellia, Licorice, Ginger, and Jujube. ^[26] . Li Jiexia ^[27] The report indicates that Xiaochaihu Tang combined with adefovir demonstrates superior therapeutic efficacy in patients with hepatitis B, outperforming adefovir alone. Chen Shaofang et al. ^[28] After administering modified Xiaochaihu Tang for 7 days to clinical patients with chronic hepatitis B characterized by liver depression and spleen deficiency, the drug-containing serum was used to treat HepG2.2.15 cells, resulting in a significant inhibition of HBsAg secretion in the supernatant. In addition, Wanshichuan ^[29] The study found that Jiawei Xiaochaihu Tang significantly inhibits the secretion of HBsAg and HBeAg by HepG2.2.15 cells, with the underlying mechanism involving suppression of the JAK2-STAT3 signaling pathway.

2.3 Kidney-tonifying compound

The Kidney-Nourishing Compound is a time-tested traditional Chinese medicine formula composed of six herbal ingredients: Astragalus, Ligustrum fruit, Epimedium, Cat’s Claw Herb, Picrorhiza, and Citrus Peel. Ji et al. ^[30] According to the report, among patients with partial virological response to entecavir, HBeAg-positive hepatitis B patients who received combined treatment with a kidney-tonifying herbal formula and entecavir demonstrated significantly better outcomes than those treated with entecavir alone. This may be associated with a reduction in the proportion of regulatory T cells (Tregs) and a downregulation of programmed death-ligand 1 (PD-L1) expression on dendritic cells. Li et al. ^[31] Studies have found that kidney-tonifying compound formulas can reduce serum levels of alanine aminotransferase (ALT) and hepatitis B virus DNA in HBeAg-positive hepatitis B patients, demonstrating particularly good therapeutic efficacy in patients with mild elevations in ALT. The underlying mechanism is believed to involve the reduction of CD4 ^plus CD25 ^plus T-cell ratio and upregulation of CD4 ^plus related to the expression level of IFN-γ in T cells. Meanwhile, Li et al. ^[32] It was also found that, in hepatitis B patients who exhibit poor efficacy with entecavir monotherapy, the combined use of a kidney-tonifying compound formula and entecavir can improve outcomes such as the clearance rates of HBsAg and HBeAg, suggesting that the kidney-tonifying compound formula may serve as an effective adjunct to entecavir.

2.4 Fuzheng Huayu Capsules

Fuzheng Huayu Capsules are composed of six traditional Chinese medicinal ingredients: Salvia miltiorrhiza, peach kernel, fermented Cordyceps sinensis powder, pine pollen, Gynostemma pentaphyllum, and Schisandra chinensis. Wang et al. ^[33] The report indicates that Fuzheng Huayu capsules, when used as an adjunctive therapy in conjunction with standard antiviral treatment for hepatitis B, can more effectively modulate levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), type III procollagen (PCIII), hyaluronic acid (HA), and laminin (LN) in patients with hepatitis B virus–related liver cirrhosis, while also increasing the rate of hepatitis B virus DNA conversion, with no serious adverse reactions. Wu et al. ^[34] Collect peripheral blood CD4 cells from patients with chronic hepatitis B complicated by liver cirrhosis. ^plus T cell CD8 ^plus T cells and natural killer cells were co-cultured in vitro with the human hepatic stellate cell line LX-2, and chronic liver injury was induced using concanavalin A. The results showed that Fuzheng Huayu Capsules significantly increased the proportion of natural killer cells, induced apoptosis and necrosis in LX-2 cells, and downregulated the expression of α-smooth muscle actin and type I procollagen. These findings suggest that Fuzheng Huayu Capsules indirectly inhibit the proliferation and activation of hepatic stellate cells by modulating the proportions and functions of T lymphocyte subsets, while promoting their apoptosis and necrosis, thereby helping to block the progression of fibrosis.

2.5 Other

In addition to the aforementioned traditional Chinese medicine formulas, in recent years, as research on herbal medicines for hepatitis B has deepened, a large number of other TCM formulas have also been reported to exhibit anti-hepatitis B activity. Kim et al. ^[35] Research has found that the traditional Chinese medicine compound KCT-01, composed of Artemisia capillaris, Sanguisorba officinalis, and Curcuma longa, can significantly inhibit the secretion of HBsAg, HBeAg, and viral particles, as well as the synthesis of pgRNA, in HepG2.2.15 cells, without affecting cell viability. At the same time, it suppresses the production of viral particles in the serum of hepatitis B virus–replicating mouse models and the synthesis of inflammatory cytokines, while also reducing the levels of covalently closed circular DNA (cccDNA) in mouse hepatocytes. Furthermore, the combined use of KCT-01 with entecavir enhances the latter’s in vitro and in vivo anti–hepatitis B virus activity. Zhang et al. ^[36] The report indicates that a kidney-tonifying prescription can enhance HBeAg seroconversion, achieve negative conversion of hepatitis B virus DNA, and normalize ALT levels in chronic hepatitis B patients with high baseline ALT levels and kidney-yang deficiency, without causing serious adverse reactions. Zhao et al. ^[37] It was found that the compound extract of Lecanthes and Shilijia can inhibit duck hepatitis B virus infection in a duck hepatitis B virus–infected model, reducing levels of DHBsAg, DHBeAg, and viral replication, while also lowering ALT and AST levels and ameliorating hepatic histopathological changes. Furthermore, this compound extract can suppress the secretion of HBsAg and HBeAg by HepG2.2.15 cells.

Numerous traditional Chinese medicine compound formulations have been reported to exhibit anti-hepatitis B virus activity, as summarized in Table 2.

3 Mechanisms of Traditional Chinese Medicine in Anti-Hepatitis B Virus Activity

Currently, the mechanisms by which traditional Chinese medicines exert anti-hepatitis B virus activity mainly encompass two aspects: direct targeting of the virus and indirect targeting of the host, as illustrated in Figure 1. Reported mechanisms of action of TCM against HBV can be broadly categorized into two major types: direct (virus-targeting) and indirect (host-targeting). The former primarily includes mechanisms such as inhibiting HBV entry (e.g., EGCG, silybin, proanthocyanidins), targeting HBV cccDNA (e.g., punicalagin, curcumin), and suppressing HBV transcription (e.g., saikosaponin C, baicalin, luteolin). The latter mainly involves modulating virus-induced cellular autophagy (e.g., EGCG, humic acid), regulating host immune function (e.g., Bushen Qudu Decoction, oxymatrine), inducing host cell apoptosis (e.g., matrine), and mitigating oxidative stress (e.g., 2,7-dichlorofluorescein, luteolin-7- O —including glucosides—and by modulating cellular signaling pathways such as JAK-STAT and PI3K-Akt-mTOR (e.g., betaine, Isatis indigotica polysaccharides, dihydroartemisinin, etc.).

3.1 Direct route

3.1.1 Inhibition of Hepatitis B Virus Entry Hepatitis B virus adsorption and entry into hepatocytes represent the initial step in the viral life cycle. For hepatitis B virus infection to proceed, the virus must first bind to and be internalized by host hepatocytes via the sodium–taurocholate cotransporting polypeptide (Na ^plus -involvement of sodium-taurocholate cotransporting polypeptide (NTCP) and clathrin ^[38-39] Therefore, NTCP and clathrin are often used as targets in the study of hepatitis B virus entry inhibitors. Huang et al. ^[40] Using HBV-infected Huh-E/2 cells and Huh7 cells transfected with an HA-NTCP expression plasmid, it was found that epigallocatechin gallate (EGCG) induces the translocation of NTCP from the plasma membrane to the cytoplasm, thereby promoting lysosomal degradation of NTCP and inhibiting HBV entry into hepatocytes. This effect may be related to EGCG’s modulation of clathrin-mediated endocytosis of NTCP. Umetsu et al. ^[41] Research has found that silybin, a constituent of milk thistle extract, reduces transferrin uptake in HepG2-NTCP-C4 cells and inhibits clathrin-mediated endocytosis, thereby suppressing hepatitis B virus entry into cells. Tsukuda et al. ^[42] The report demonstrates that proanthocyanidins and their analogs inhibit the binding of the pre-S1 domain of the hepatitis B virus large surface protein to NTCP, thereby blocking viral entry into hepatocytes, reducing viral infectivity, and exhibiting inhibitory activity against all genotypes of hepatitis B virus.

3.1.2 Targeting hepatitis B virus cccDNA. The persistent and stable presence of hepatitis B virus cccDNA is one of the main reasons why chronic hepatitis B remains difficult to cure. Therefore, cccDNA represents an important therapeutic target for the development of anti-hepatitis B viral agents. ^[43-45] . Liu et al. ^[46] A cell-based assay for cccDNA accumulation and stability was established, revealing that three hydrolyzable tannins, including punicalagin, exert anti-hepatitis B virus activity by inhibiting cccDNA formation and promoting its degradation, thereby reducing cccDNA levels. Wei et al. ^[47] Research has shown that curcumin, extracted from plants of the Zingiberaceae and Araceae families such as turmeric, can significantly reduce the acetylation levels of histones H3 and H4 bound to cccDNA, thereby disrupting cccDNA homeostasis. This suggests that curcumin holds promise as a potential anti-hepatitis B virus compound targeting cccDNA.

3.1.3 Inhibition of hepatitis B virus transcription: High antigen load is a major contributor to liver injury in patients with chronic hepatitis B. By suppressing the transcription of cccDNA into various viral RNAs, thereby reducing the expression of hepatitis B surface antigen (HBsAg), core antigen (HBcAg), HBx protein, and other viral proteins, it is possible to effectively alleviate hepatocellular damage in these patients. ^[48] . Pan et al. ^[49] The study found that saikosaponin C downregulates the levels of HNF-1α and HNF-4α by stimulating IL-6 expression, thereby inhibiting the synthesis of hepatitis B virus pgRNA, and that it is effective against both wild-type and drug-resistant hepatitis B viruses. Huang et al. ^[50] The report also indicates that baicalin exerts its inhibitory effects on hepatitis B virus transcription and replication by downregulating the expression of HNF-1α and HNF-4α. Bai et al. ^[51] Studies have shown that luteolin can activate the ERK signaling pathway, thereby inhibiting the expression of HNF-4α and its binding to the hepatitis B virus promoter, ultimately disrupting hepatitis B virus transcription and replication.

3.2 Indirect pathway

3.2.1 Regulation of Hepatitis B Virus–Induced Autophagy Cellular autophagy exerts a dual role in hepatitis B virus infection: in the early stages of chronic HBV infection, the virus can induce incomplete autophagy in host cells to activate its own replication; however, once chronic HBV infection progresses to hepatocellular carcinoma, the activation of autophagy can mitigate further tumor progression by inducing cell death and enhancing anti-tumor immune responses. ^[52-53] . Zhong et al. ^[54] According to the report, epigallocatechin gallate (EGCG), a polyphenolic compound extracted from green tea, can inhibit hepatitis B virus–induced incomplete autophagy in host cells by enhancing lysosomal acidification, thereby reducing viral replication within host hepatocytes. Pant et al. ^[55] The study found that humic acid, the main component of mineral bitumen, can effectively inhibit HBx expression in HepG2.2.15 cells, thereby reducing the formation of autophagosomes induced by hepatitis B virus and lowering the level of viral replication.

3.2.2 Modulation of Host Immune Function: Persistent hepatitis B virus infection leads to a reduction in the number and impaired function of innate immune cells throughout the body, resulting in systemic innate immune tolerance. Meanwhile, prolonged exposure to the antigenically loaded hepatic microenvironment causes progressive dysfunction of hepatitis B virus–specific T cells, rendering them incapable of effectively clearing the virus from the host. ^[56-58] . Liu et al. ^[59] According to the report, the Kidney-Nourishing and Toxin-Removing Decoction modulates immune function by either enhancing the generation of Th1 cells or by boosting Th1 cell function while simultaneously suppressing Th17 cell function, thereby exerting an anti-hepatitis B virus effect. Yao et al. ^[60] Studies have shown that oxymatrine can upregulate the expression of Toll-like receptor 9 (TLR9) signaling molecules, activate TLR9 signaling pathway function, and induce the production of antiviral cytokines such as IFN-α and IFN-γ.

3.2.3 Regulation of Host Cell Apoptosis Although some studies have reported that hepatitis B virus can induce hepatocyte apoptosis, most researchers believe that the virus attenuates the host’s clearance response by inhibiting apoptosis. ^[61-63] . Liu Xin et al. ^[64] The report indicates that matrine can induce apoptosis in HepG2.2.15 cells, increase the proportion of cells in the S phase, and thereby exert an anti-hepatitis B virus effect. Su et al. ^[65] The study found that water-soluble polysaccharides extracted from Sipunculus nudus can induce dose-dependent apoptosis in HepG2.2.15 cells by upregulating the expression of apoptosis-related proteins such as TNF-α and cysteine-aspartate protease-3 (Caspase-3), thereby inhibiting hepatitis B virus DNA replication in these cells.

3.2.4 Regulating hepatitis B virus–induced oxidative stress: Hepatitis B virus variants carrying PreS/S gene mutations exhibit excessive synthesis of surface proteins in host hepatocytes, leading to their accumulation in the endoplasmic reticulum and triggering endoplasmic reticulum stress. This, in turn, results in oxidative DNA damage and genomic instability in hepatocytes, ultimately precipitating hepatocellular injury. ^[66-67] . Arbab et al. ^[68] Report: Medicinal Plants Guiera senegalensis J.F. Gmel extract can ameliorate 2,7-dichlorofluorescein-induced oxidative stress in hepatocytes, thereby protecting these cells and suggesting its potential as an anti-hepatitis B virus–active compound. Cui et al. ^[69] Research has found that lettuce extract luteolin-7- O -Glucosides may inhibit the secretion of HBsAg and HBeAg, as well as hepatitis B virus transcription and replication, by suppressing hepatitis B virus–induced reactive oxygen species production and thereby improving mitochondrial function.

3.2.5 Regulation of cellular signaling pathways: Common signaling pathways such as JAK-STAT, NF-κB, PI3K-Akt-mTOR, and MAPK are closely associated with hepatitis B virus replication, infection, and the progression of virus-induced hepatocellular carcinoma. ^[70-73] The anti-hepatitis B virus activity of active extracts from traditional Chinese medicine often involves the regulation of multiple signaling pathways. For example, betaine and Isatis indigotica polysaccharides enhance JAK-STAT signal transduction, thereby improving the interferon response in HepG2.2.15 cells and demonstrating potent anti-hepatitis B virus activity. ^[74-75] . Hesari et al. ^[76] Studies have shown that curcumin exhibits anti-hepatitis B virus activity, with its mechanism of action associated with the downregulation of the NF-κB and Wnt/β-catenin signaling pathways. Zou et al. ^[77] The study found that dihydroartemisinin exerts its inhibitory effects on the proliferation and migration of HepG2.2.15 cells by suppressing the expression of tripeptidyl peptidase I (TPP I), enhancing the DNA damage response pathway to induce cellular senescence in HepG2.2.15 cells, and downregulating autophagy in these cells mediated by the Akt-mTOR signaling pathway, thereby reducing the tumorigenicity of these cells in mice. Fang et al. ^[78] The report indicates that lentinan can promote the secretion of cytokines such as IL-1β, IL-12, and TNF-α in monocytes/macrophages, thereby alleviating HBsAg-induced impairment of host immune function; the underlying mechanism may involve lentinan-mediated phosphorylation of IκB-α, which activates the NF-κB signaling pathway, as well as activation of the MAPK signaling pathway. Wang et al. ^[79] The report indicates that EGCG downregulates HNF-4α expression by activating and enhancing ERK signaling, thereby reducing C/preS promoter activity and subsequently impairing hepatitis B virus transcription and replication.

4 Summary and Outlook

In recent years, researchers both domestically and internationally have conducted extensive basic research and clinical trials to elucidate the anti–hepatitis B virus efficacy of traditional Chinese medicine extracts and compound formulations. Active constituents (or extracts) with anti–hepatitis B virus activity from single herbs such as Artemisia capillaris, Polygonum cuspidatum, Salvia miltiorrhiza, Sophora flavescens, and Aloe vera have been successively reported in the literature; among compound formulations, those based on Phyllanthus urinaria, Xiaochaihu Tang, kidney-tonifying formulas, and Fuzheng Huayu capsules have been particularly well documented. The reported active extracts and compound formulations generally reduce levels of HBsAg and HBeAg in the supernatant of HepG2.2.15 cell cultures, in duck hepatitis B virus–infected duck models, or in the serum of patients with chronic hepatitis B; some traditional Chinese medicines even exhibit activity against nucleoside analog–resistant strains of hepatitis B virus. In addition, the mechanisms underlying the anti–hepatitis B virus effects of certain traditional Chinese medicines or their extracts have been progressively elucidated, encompassing both direct pathways—such as inhibition of viral entry, transcription, and targeting of covalently closed circular DNA—and indirect pathways—such as modulation of virus-induced autophagy, host immune function, host cell apoptosis, virus-induced oxidative stress, and cellular signaling pathways.

Traditional Chinese medicine holds great promise for the prevention and treatment of hepatitis B virus infection. However, the current research and development of anti-hepatitis B viral TCM still faces the following challenges: (1) TCM products currently lack standardized quality criteria. ^[80] Moreover, the varying disease courses of hepatitis B lead to substantial differences in the therapeutic efficacy of traditional Chinese medicines; (2) in clinical studies of traditional Chinese medicines for anti-hepatitis B virus therapy, few randomized, double-blind, controlled trials have been conducted; most studies have only evaluated the short-term efficacy of these herbs, and many lack comprehensive reports on adverse drug reactions. ^[81] (3) The multi-component, multi-target, and multi-pathway characteristics of traditional Chinese medicines pose additional challenges to research on the therapeutic targets and mechanisms of action of TCM agents against hepatitis B virus. ^[82-83] Therefore, in the future, unified and comprehensive quality standards for traditional Chinese medicines should be established, while modern molecular biology techniques should be employed to quantitatively measure HBsAg levels in patients’ serum, thereby ensuring the consistent therapeutic efficacy of these medicines across hepatitis B patients at different stages of the natural disease course and with different genotypes, and ultimately enabling individualized antiviral treatment. It is also necessary to conduct multicenter, large-scale, prospective, randomized, double-blind controlled trials, coupled with long-term follow-up studies in chronic hepatitis B patients, to obtain more reliable data on drug efficacy and adverse reactions. Furthermore, based on information regarding the active constituents of traditional Chinese medicines, their molecular targets, and the life cycle of the hepatitis B virus, network pharmacology approaches should be utilized to efficiently predict the potential targets of anti-hepatitis B viral agents derived from traditional Chinese medicine. In addition, building upon the inheritance of traditional Chinese medical theory, deep interdisciplinary integration among traditional Chinese medicine, medicine, pharmacology, pharmacokinetics, pathology, biochemistry, structural biology, bioinformatics, genomics, and other fields should be pursued, with close collaboration and coordinated development between basic research and clinical applications, thus laying a solid foundation for the discovery of novel, low-toxicity, highly effective anti-hepatitis B viral drugs from traditional Chinese medicine.