The Efficacy and Mechanism of Chinese Herbal Medicines in Lowering Serum Uric Acid Levels: A Systematic Review

Background. Chinese herbal medicines are widely used to lower serum uric acid levels. However, no systemic review summarizes and evaluates their efficacies and the underlying mechanisms of action. Objectives. To evaluate the clinical and experimental evidences for the effectiveness and the potential mechanism of Chinese herbal medicines in lowering serum uric acid levels. Methods. Four electronic databases PubMed, Wed of Science, the Cochrane Library and Embase were used to search for Chinese herbal medicines for their effects in lowering serum uric acid levels, dated from 1 January 2009 to 19 August 2020. For clinical trials, randomized controlled trials (RCTs) were included; and for experimental studies, original articles were included. The methodological quality of RCTs was assessed according to the Cochrane criteria. For clinical trials, a meta-analysis of continuous variables was used to obtain pooled effects. For experimental studies, lists were used to summarize and integrate the mechanisms involved. Results. A total of 10 clinical trials and 184 experimental studies were included. Current data showed that Chinese herbal medicines have promising clinical efficacies in patients with elevated serum uric acid levels (SMD: −1.65, 95% CI: −3.09 to −0.22; p = 0.024). There was no significant difference in serum uric acid levels between Chinese herbal medicine treatments and Western medicine treatments (SMD: −0.13, 95% CI: −0.99 to 0.74; p = 0.772). Experimental studies revealed that the mechanistic signaling pathways involved in the serum uric acid lowering effects include uric acid synthesis, uric acid transport, inflammation, renal fibrosis and oxidative stress. Conclusions. The clinical studies indicate that Chinese herbal medicines lower serum uric acid levels. Further studies with sophisticated research design can further demonstrate the efficacy and safety of these Chinese herbal medicines in lowering serum uric acid levels and reveal a comprehensive picture of the underlying mechanisms of action.


INTRODUCTION
Hyperuricemia refers to an abnormally high concentration of serum uric acid (sUA), typically defined as >7 mg/dL in men and >6 mg/dL in women. The data from the National Health and Nutrition Examination Survey (NHANES) [2007][2008][2009][2010][2011][2012][2013][2014][2015][2016] showed that the prevalence rates of hyperuricemia were 20.2% among men and 20.0% among women between 2015 and 2016 in the United States (Chen-Xu et al., 2019). A cross-sectional survey in China showed that the prevalence of hyperuricemia was 8.4% among Chinese adults from 2009 to 2010 (Liu et al., 2014a). Hyperuricemia and gout remain as a considerable burden, which not only adversely affect patients' health and quality of life (Burke et al., 2015;Gamala and Jacobs, 2019), but also cast an economic burden in the society (Rai et al., 2015). SUA is associated with cardiovascular diseases, such as hypertension (Kuwabara et al., 2018) and atrial fibrillation (Tang et al., 2014). Elevated sUA can lead to decreased renal function, which in turn reduces the excretion of UA in urine, resulting in an increased risk of hyperuricemia or gout (Sato et al., 2019). Elevated sUA may also contribute to the pathogenesis of metabolic syndrome (Battelli et al., 2018;Battelli et al., 2019), non-alcoholic fatty liver disease , diabetes (Kim et al., 2015). Uratelowering therapy is a therapeutic strategy for controlling gout, chronic kidney disease, metabolic syndrome and many other diseases. Current interventions for elevated sUA include xanthine oxidase inhibitors, uricosuric agents and anti-inflammatory drugs. Although both febuxostat and allopurinol are effective in reducing sUA, allopurinol may produce a mild skin rash and severe cutaneous reactions (Strilchuk et al., 2019), while febuxostat has a higher risk of all-cause and cardiovascular mortality (White et al., 2018). Benzbromarone is a uricosuric drug which is widely used, but studies have reported possible complications such as hepatotoxicity (Strilchuk et al., 2019). Although these drugs are clinically used, their efficacies are unsatisfactory, and are usually coupled with adverse side effects in the long-term use (Dalbeth et al., 2016).
Hyperuricemia belongs to the arthromyodynia disease category in traditional Chinese medicine. Chinese herbal medicine has been used to treat hyperuricemia for a long time and has significant clinical efficacy . Chinese herbal medicines with high efficacy and low incidence of adverse reactions have drawn increasing attention from scholars. Studies have compared Chinese herbal medicine with Western medicine for their efficacies in lowering sUA levels. However, these studies differ in their treatment protocols and evaluation methodologies, which greatly limit their clinical applicability. In addition, the mechanism of Chinese herbal medicine in lowering sUA levels is still being explored. Some herbs such as Phellodendri Chinrnsis Cortex, Atractylodes Lancea (Thunb.)Dc. (Chen et al., 2015b), Smilacis Glabrae Rhixoma , reduce UA intake and/or increase UA excretion by regulating various physiological and cellular pathways. Some herbs like bergenin (Chen et al., 2020a), alpinia oxyphylla seed extract (Lee et al., 2019b) and rhizoma smilacis glabrae extracts , promote renal and gut uric acid excretion in hyperuricemia models and also decrease the serum levels of inflammatory cytokines. Most Chinese herbal medicines act on multiple targets to achieve their sUA lowering effects. Therefore, the aim of this systematic review is to evaluate the evidence for the efficacy of Chinese herbal medicines in lowering sUA levels in patients with hyperuricemia, compared to no intervention, placebo or urate-lowering agents, and to comprehensively summarize the mechanisms underlying the sUA lowering effects reported from experimental studies.

Data Source and Search Strategy
Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) (Moher et al., 2009) was used to construct the report of the current study and the completed checklist is provided in Supplementary material, Supplementary Table S1. The electronic databases PubMed, Wed of Science, the Cochrane Library and Embase were systematically searched by three researchers (Liqian Chen, Zhengmao Luo and Ming Wang). For any discrepancies between researchers, consensus was reached through discussion. We reviewed literatures published from 1 January 2009 to 19 August 2020 on elevated sUA that had been treated with Chinese herbal medicines. The following combination of terms were used as search keywords: (Traditional Chinese Medicine OR Herbal Medicine OR Chinese Herbal Drugs OR Chinese Plant Extracts OR (Plants, Medicinal) OR Phytochemicals OR herb* OR natural product) combined with (gout OR pain paralysis OR hyperuricemia OR uric acid). The full search strategy in PubMed was provided in Supplementary material, Supplementary Table S2. The search did not exclude articles based on language. To look for additional relevant studies, references of all potentially relevant articles were also retrieved, and authors of studies that met the inclusion criteria but lacked data would be contacted. Abstracts, meeting proceedings, and personal communications were not used for the purpose of this study.

Study Selection
The articles in this review included clinical trials and in vivo and in vitro studies. When screening clinical trials, the followings are the inclusion criteria: A). Types of trials: All randomized controlled trials (RCTs) investigating the use of Chinese herbal medicine in lowering sUA will be eligible for inclusion. B). Types of participants: In accordance with the NHANES-III laboratory definition, hyperuricemia is typically defined as >7 mg/dL in men and >6 mg/dL in women (Choi et al., 2020). All adult patients (18 years and older, no upper age limit) with a diagnosis of hyperuricemia will be considered for this review. C). Types of interventions: According the General guidelines for methodologies on research and evaluation of traditional medicine from WHO, herbal medicines are defined as materials or products derived from plants that have medical or other beneficial effects on human health, including herbs, herbal materials, herbal products, finished herbal products that contain parts of plants or other plant materials or compositions as active ingredients, as well as materials of inorganic or animal sources. The interventions included in this study include Chinese herbal medicine in various prescriptions, such as herbal formulas, herbal extracts, active ingredients of herbs. Clinical trials comparing Chinese herbal medicine with no intervention, placebo or urate-lowering agents were included in our study. Chinese herbal medicine plus placebo or combined with urate-lowering agents compared to the same medications was also included. We do not limit the formulations or administration of herbal preparations for clinical use. D). Types of outcome measures: Outcome measures should include at least one essential outcome, such as change in sUA levels after treatment and the overall efficacy.
When screening in vivo or vitro experimental studies using the herbs, the following conditions should be met before inclusion: A) Original article investigating the use of Chinese herbal medicine in lowering sUA will be eligible for inclusion. B) Studies using experimental models of mice, rats, rabbits or cell cultures will be considered. All models used should present corresponding pathological symptoms. C) The model should only be treated with Chinese herbal medicine before or after intervention. If medication other than herbs were being used, both the treatment group and the control group must be administered. D) Outcome measures should include change in sUA after treatment.

Data extraction
The data were extracted by three researchers (Jingru Cheng, Fei Li and Hanqi Lu) independently to obtain the following information: A) the design of study; B) characteristics of trial participants (including sample size, age, period); C) type of intervention (including dose regimen, duration); D) type of outcome measure (including the level of sUA). The reported mean (standard deviation [SD]) or risk estimates and 95% confidence intervals (CIs) of sUA were also extracted.

Quality Assessment
The quality assessment of the studies was performed independently by three researchers (Qiuxing He, Yanting You and Xinghong Zhou), and all discrepancies between researchers were resolved through discussion. The methodological quality of RCTs was assessed according to the Cochrane Handbook for Systematic reviews of Interventions. The scores for each bias domain and the final score of risk of systematic bias were graded as low, high or unclear risk. The overall level of evidence was considered "strong" if there were consistent findings among multiple high quality RCTs, and "moderate" if findings were consistent among multiple lowquality RCTs and/or one high-quality RCT. Level of evidence was "conflicting" if findings were inconsistent across the studies, and "no evidence from trials," if there were no RCTs.

Statistical Analysis
Statistical analysis was performed with the software STATA version 16.0. The d index and the standard deviation (SD d ) values of sUA for each RCT were calculated before using STATA. The d index and SD d values of sUA were continuous, standard mean difference (SMD) with 95% confidence interval (95% CI) was calculated. The overall effect was calculated by a Z-test, and p < 0.05 (2-tailed) was deemed statistically significant. Potential heterogeneity was assessed by I 2 statistics. A fixed effects model was chosen if I 2 ≤ 50%, otherwise, a random effects model was applied. Subgroup analysis was performed to illuminate the heterogeneity according to the study characteristics, such as interventions other than Chinese herbal medicine. A 'leaveone-out' sensitivity analysis was carried out to test the reliability of the results. Potential publication bias was evaluated by the Egger's test and Begg's test. When the geometric mean and CIs were reported, tools provided in the Cochrane Handbook were used to convert the geometric mean and CIs to arithmetic mean and SD of the raw data.

Literature Flow
The initial electronic search of the literature yielded 4981 potentially relevant citations. After duplicate removal and title/ abstract screening, 374 full-text articles were retrieved for detailed assessment. Of these studies, 180 articles did not meet the inclusion criteria. Finally, 194 articles were included in the review with 10 clinical trials, 169 in vivo experiments, 0 in vitro experiments, and 15 that were a combination of both in vitro and in vivo experiments ( Figure 1).

Study Characteristics
The specified characteristics of the included clinical trials as well as their study populations were summarized in Table 1. All clinical trials included were RCTs which mainly explored the effects of Chinese herbal medicine on patients with elevated sUA levels such as hyperuricemia and gout. In order to objectively observe the therapeutic effects of herbs on lowering sUA levels, the changes of sUA levels before and after treatment in 10 clinical trials were listed in Supplementary material, Supplementary Table  S3. Of the 10 included trials, nine were treated with herbal formula, and the remaining one was treated with a combination of herbal extracts and active ingredients. In the subgroup analyses of intervention, two studies compared the therapeutic effects of Chinese herbal medicine and placebo (Rozza et al., 2016;Xie et al., 2017), three studies compared Chinese herbal medicine and Western medicine (Zhang et al., 2009;Zhou et al., 2013;Wang et al., 2014), two studies compared Chinese herbal medicine, Western medicine and placebo or no intervention Wang et al., 2019b), one compared two kinds of Chinese herbal medicine and Western medicine (Yu et al., 2018), and two compared Chinese herbal medicine combined Western medicine and Western medicine (Chen et al., 2009;Xiang et al., 2009).

Meta-Analysis
In order to investigate the efficacy of Chinese herbal medicine in lowering sUA, we performed a subgroup analysis based on interventions. A random effects model was used for the analysis because I 2 96.4%.

Chinese Herbal Medicine vs Placebo or No Intervention
Four RCTs were analyzed. The subgroup meta-analysis showed that all of them were statistically significant differences between Chinese herbal medicine and placebo or no intervention. The combined SMD was −1.65 with a 95% CI of −3.09 to −0.22 (p 0.024). Therefore, there was a significant difference between the Chinese herbal medicine and placebo or no intervention in the reduction of sUA ( Figure 2).

Chinese Herbal Medicine vs Western Medicine
Seven RCTs were analyzed. On subgroup meta-analysis, the combined SMD was −0.13 with a 95% CI of −0.99 to 0.74 (p 0.772), indicating no significant difference between the Chinese herbal medicine and Western medicine in the reduction of sUA ( Figure 2).

Chinese Herbal Medicine Plus Western Medicine vs Western Medicine
Two RCTs were analyzed. Subgroup meta-analysis ( Figure 2) showed no statistical significance between the Chinese herbal medicine plus Western medicine and Western medicine in the reduction of sUA (SMD −2.27, 95% CI: −5.84 to 1.31, p 0.214).

Meta-regression Analyses
The meta-regression showed that samples size, duration of treatment, type of diseases and intervention other than Chinese herbal medicine did not influence these results (all p values >0.05) (Supplementary Figure S1).

Quality Assessment
RCT was assessed according to the Cochrane Handbook for Systematic reviews of Interventions (Table 2). Generally, the methodological quality was assessed to be moderate. Most of the studies (8/10, 80%) have details on the random grouping of patients, but only half of the studies (5/10, 50%) fully reported the scheme of concealment allocation. Only four trials (4/10, 40%) had their subjects and investigators blinded during the study, and four trials (4/10, 40%) had all the subjects, investigators and outcome evaluators blinded.

Sensitivity Analysis
Sensitivity analysis was conducted to confirm the efficacy Chinese herbal medicine in lowering sUA. The pooled SMDs were  repeated by sequentially removing one of the included studies with a random-effects model (Supplementary Figure S2). None of the studies changed the overall effect.

Publication Bias
There was no evidence of publication bias according to the Begg's test (p 0.428) and Egger's test (p 0.344) for the meta-analysis of Chinese herbal medicine on lowering sUA (Supplementary Figure S3).

Results of experimental studies
We categorized 184 in vivo and in vitro experiments into three groups: 56 active ingredients (Table 3), 78 natural products (Table 4), and 52 herbal formulas ( Table 5). Among them, one article described two active ingredients , and one article (Su et al., 2014) studied one active ingredient and one natural product concurrently. To assure the quality of the studies included, detailed information on herbs (including source, concentration, quality assessment, chemical analysis, and compound purity) was summarized in Supplementary material, Supplementary Tables S4-S6.

Active Ingredients That Lower Serum Urate in vivo and in vitro Studies
Active ingredient is a single ingredient and studies have shown that it plays an important therapeutic role in   reducing sUA. Among the active ingredients with potential of lowering sUA ( Table 3), 25 of which act on uric acid synthesis, 28 target uric acid transporter, 19 resolve inflammation, nine possess kidney protective function, and four regulate the oxidative stress.

Natural Products That Lower Serum Urate in vivo and in vitro Studies
Natural products include herbs and relatively complex extracts derived from herbs. Among the natural products with potential of lowering sUA ( Table 4), 40 of which act on uric acid synthesis, 29 target uric acid transporter, 17 resolve inflammation, eight possess kidney protective functions, and three regulate the oxidative stress.

Herbal Formulas That Lower Serum Urate in vivo and in vitro Studies
Herbal formula is a combination of a variety of herbs. The composition of the herbal formula included in the current study were listed in Supplementary material, Supplementary Table S7.

DISCUSSION
The efficacies of Chinese herbal medicines lowering Serum uric Acid levels This systematic review compares the efficacies of the Chinese herbal medicines and the Western medicine in lowering sUA levels by analysing10 RCTs with a total of 1,060 patients. The meta-analysis has three important findings. First, there is a significant difference between the Chinese herbal medicine, placebo or no intervention in the reduction of sUA levels. Second, the efficacies of Chinese herbal medicines in lowering sUA levels are comparable to that of Western medicine. Third, the efficacies of Chinese herbal medicines plus Western medicine in lowering sUA levels are comparable to that of Western medicine. The heterogeneity of the meta-analysis was high. To investigate the source of heterogeneity in our analysis, we conducted a sensitivity analysis, removing one study at one time from the primary analysis did not change the main finding. In addition, our findings were confirmed by the lack of publication bias and effect modifiers according to the Begg's test, Egger's test and meta-regression analysis.
The results indicate that the efficacies of Chinese herbal medicines in lowering sUA levels are comparable to that of Western medicine, which is consistent with the analysis result of Lin et al. . Compared with the study of Lin et al., our study is more comprehensive, because we also compared the sUA lowering efficacy between Chinese herbal medicine and placebo or no intervention, and between Chinese herbal medicine plus western medicine and western medicine. In addition, we have summarized the underlying mechanisms of herbs in lowering sUA. It should be noted that among the 10 RCTs included, a total of six RCTs used one or several components of Simiao Pills, which is a famous formula in traditional Chinese medicine, including Cortex Phellodendri Chinensis, Atractylodes Lancea (Thunb.) DC., Coix lacrymajobi L.var.ma-yuen (Roman.) Stapf (Yi Yi), Cyathula officinalis Kuan (Xiang et al., 2009;Zhang et al., 2009;Zhang et al., 2011;Zhou et al., 2013;Xie et al., 2017;Yu et al., 2018). The specific doses of herbs in each study have been listed in the  Supplementary  Tables  S4-S6. According to the Pharmacopoeia of the People's Republic of China revised by the Food and Drug Administration in 2015, except for the use of Cortex Phellodendri Chinensis, Atractylodes Lancea (Thunb.) DC., Cyathula officinalis Kuan in one RCT (Xiang et al., 2009) exceeded the recommended dosage, the dosages of these four herbs in the remaining five RCTs were within the recommended dosage range. Except for the RCT in which the dosage exceeded the recommended dosage (Xiang et al., 2009), the dosage of these four herbs in the remaining five RCTs were all at the high level or even reached the critical value within the recommended range. Despite the high dosage, the three RCTs describing the adverse reactions in the six RCTs showed that the incidence of adverse reactions was either lower than that of the placebo group or was not statistically significant compared with the control group.
In addition to clinical studies, experimental studies have also suggested the urate lowering effects of Simiao Pills, which act on xanthine oxidase (XO), uric acid transport-related proteins urate anion transporter 1 (URAT1) and glucose transporter 9 (GLUT9), and also modulate the inflammation, oxidative stress and other processes Hua et al., 2012;Shi et al., 2013;Pan et al., 2014;Ma et al., 2015;Lin et al., 2020). The finding suggests that Chinese herbal medicines are mostly multi-targeted or have interplay with other signaling pathways to lower sUA levels. In contrast, conventional Western medicines used to lower sUA levels include allopurinol, probenecid and benzbromarone, which generally act on specific targets. Allopurinol competitively inhibits xanthine oxidase (Strilchuk et al., 2019). Probenecid and benzbromarone are typical uratepromoting drugs target at URAT1 (Azevedo et al., 2019). These may partly explain why the combination of Chinese and Western medicine can improve the index of renal function while lowering sUA levels in patients with hyperuricemia (Chen et al., 2009;Xiang et al., 2009).
Nevertheless, several limitations of the meta-analysis are worth considering. Due to limited reports, participants with elevated sUA levels in this meta-analysis included patients with hyperuricemia and gout, and most participants were from China. These facts indicate that the current metaanalysis could have potential bias. Further trials need to be carried out in a larger comprehensive population to demonstrate the efficacy of Chinese herbal medicines in lowering sUA. In addition, long-term tracking of sUA is necessary to determine whether Chinese herbal medicine can effectively control sUA. However, long-term follow-ups were not available in the current included studies. The incidence of adverse reactions is an important indicator to compare the efficacy and MSU SD rats TNF-α, IL-1β, NF-κB Ma et al. (2009) Abbreviations can be referred to Table 3. ICAM1, intercellular adhesion molecule one; IFN, interferon; AMPK, adenosine 5′-monophosphate-activated protein kinase; FOXO3α, forkhead box O3 alpha; SDF-1, stromal cell-derived factor 1; CXCR4, SDF-1 receptor; MKK, mitogen-activated kinase; TRAIL, the tumor necrosis factor-related apoptosis-inducing ligand.
Frontiers in Pharmacology | www.frontiersin.org January 2021 | Volume 11 | Article 578318 safety of Chinese herbal medicine and Western medicine in lowering sUA levels. However, of the six included studies comparing Chinese herbal medicine and Western medicine, only 2 has reported the adverse reaction events in details. Thus, we cannot evaluate the safety of Chinese herbal medicine vs. western medicine in lowering sUA levels. Moreover, the standardization of the methodologies and the small number of the included trials may lead to an overestimation of the overall efficacy of Chinese herbal medicine. Therefore, studies with high quality are needed to confirm the efficacy and safety of Chinese herbal medicine in lowering sUA levels.

The mechanisms of Chinese herbal medicines in lowering Serum uric Acid
In current review, 186 in vivo and in vitro experiments with 56 active ingredients (Table 3), 78 natural products (Table 4), and 52 herbal formulas ( Table 5) are included to explore the common mechanism of Chinese herbal medicine in lowering sUA levels.
According to the summary of the targets of Chinese herbal medicine in lowering sUA, it is clearly revealed that most Chinese herbal medicine lowers sUA by acting on multiple targets or multiple pathways. The therapeutic mechanism of Chinese herbal medicine included in this study mainly involved the UA synthesis, UA transport, inflammation, renal fibrosis and oxidative stress. Uric acid, produced primarily in the liver, is the final product of diet and endogenous purine metabolism. Problems with key enzymes involved in UA production can cause abnormal UA levels, including phosphoribosyl pyrophosphate (PRPP) synthetase, purine nucleoside phosphorylase, xanthine oxidase, hypoxanthine-guanine phosphoribosyl transferase (HGPRT) (Cammalleri and Malaguarnera, 2007;Maiuolo et al., 2016). In the circulation, UA exists mainly in the form of urate anion under physiologic pH. The saturation level of monosodium urate in human plasma is limited. Hence, UA must be excreted continuously to prevent its accumulation and reduce the toxicity. The primary scavenger of urate is the kidney, which expels about 75% of urate every day (Chaudhary et al., 2013). In recent years, studies have revealed the complex interaction of transporters involved in urate metabolism (Wright et al., 2010). Several transporters involved in urate metabolism have been identified, including URAT1, GLUT9, organic anion transporters (OAT1/3), and adenosine triphosphate (ATP)binding cassette transporter 2 (ABCG2) (Enomoto et al., 2002;Lipkowitz, 2012;Mandal and Mount, 2015). Previous studies have found that inflammation, oxidative stress, mitochondrial Abbreviations can be referred to Tables 3, 4. FABP1, fatty acid-binding protein; HPRT1, hypoxanthine-guanine phosphoribosyl transferase; APOB, apolipoprotein B; FOS, one subunit of activator protien-1; FN1, fibronectin1; MIP-1α, MIP-1 β, serum proinflammatory cytokines; TXNIP, thioredoxin interacting proteins; PTEN, phosphate and tension homology deleted on chromsome ten; NALP1/6, NACHT, LRR and PYD domains-containing protein one and six; ADAMTs, a family of metalloproteinases with thrombospondin motifs; TIMP, tissue inhibitor of metalloproteinase; CD2AP, CD2-associated protein.

Herbs Lower Serum Uric Acid by Targeting Uric Acid Synthesis
The substrate for UA synthesis is ribose-5-phosphate which can be converted to PRPP via PRPP synthase and then to inosine monophosphate. This intermediate compound produces adenosine monophosphate and guanosine monophosphate, which subsequently release adenosine and guanosine molecules, respectively. Adenosine deaminase converts adenosine to inosine, while guanosine to free guanine. Inosine is degraded to hypoxanthine via purine nucleoside phosphorylase. Xanthine oxidase, one of the key enzymes involved in UA synthesis, converts hypoxanthine to xanthine which then converts to UA. Guanine is directly converted to xanthine, which subsequently to UA by XO. On the other hand, hypoxanthine and guanine enter a salvage pathway through the activity of HGPRT, which converts these purine into their respective nucleotides (Lipkowitz, 2012;Lima et al., 2015;Maiuolo et al., 2016). Thus, PRPP synthase, XO and HGPRT are all key enzymes that can cause abnormal sUA levels. PRPP gene mutations have been implicated in a number of human diseases. Overexpression of PRPP results in the enhanced activity of phosphoribosyl pyrophosphate synthetase-I, which can lead to excessive production of purine. Patients with active phosphoribosyl pyrophosphate synthetase-I may result in UA over production (Mittal et al., 2015). Lesch-Nyhan disease is caused by a wide variety of mutations in the HGPRT gene and is one of the models of gout caused by the increased production of UA (Fu et al., 2014;Harris, 2018). Lack of HGPRT can lead to Kelley-Seegmiller syndrome, which is characterized by hyperuricemia, hyperuricosuria, gouty arthritis and urolithiasis (Chavarriaga et al., 2019). Among these key enzymes, XO is the most commonly studied enzyme. 3,5,2′,4′-tetrahydroxychalcone significantly inhibits the activities of XO in liver, and the decreased content of PRPP in liver will suppresse the UA production (Niu et al., 2018). In our review, nearly half of the studies (83/186, 45%) confirm that herbs regulate XO synthesis and activity. For example, pallidifloside D, a saponin glycoside constituent from the total saponins of Smilax riparia, enhances the hypouricemic effect of allopurinol by regulating XO activity. The combination of allopurinol and Pallidifloside D significantly up-regulates HGPRT expression and down-regulates the expressions of PRPP in PC12 cells .

Herbs Lower Serum Uric Acid by Targeting Uric Acid Transporter
About two-thirds of UA is excreted by the kidneys, and the remaining is excreted via the gastrointestinal tract. Renal excretion of UA consists of four steps: 1) glomerular filtration, 2) presecretory reabsorption, 3) secretion and 4) post-secretory reabsorption (Maiuolo et al., 2016). Urate is filtered through the glomerulus, so the reabsorption and secretion of UA after glomerulus play an important role in regulating the excretion of UA. Most of the urate (99%) filtered through the glomerulus is reabsorbed in the early S1 segment of the proximal tubule (presecretory reabsorption). Uric acid is then secreted in the S2 segment of the proximal tubule to return approximately 50% of the filtered urate into the tubule lumen. Post-secretory reabsorption occurs primarily in the distal S3 segment of the proximal tubule, followed by about 10% of the secreted urate in the urine (Diamond and Paolino, 1973;Mandal and Mount, 2015). Urate transporters such as URAT1, OAT1, OAT3, GLUT9, ABCG2 play important roles in regulating sUA, and their dysfunction may cause abnormal urate transport. URAT1, the main urate-anion exchanger in the luminal membrane of the proximal tubules, can be inhibited by the uricosuric agents, likes probenecid, benzbromarone and losartan (Enomoto et al., 2002). URAT1 mutations have been found in patients with familial hypouricemia and UA levels below 1 mg/dL, and the UA transport is inactivated when this mutant transporter is expressed in xenopus oocytes (Dinour et al., 2011). Clinical studies have shown that inhibition of URAT1 effectively reduces sUA levels and resolves the gout symptoms (Lee et al., 2019a). URAT1 is a branch of the organic anion transporter (OAT). OAT1 and OAT3 exchange urate with bivalent anions, suggesting that they are suitable for basolateral entry of urate during urate secretion (Mandal and Mount, 2015). Among the 14 members of the GLUT family transport glucose or other monosaccharides, GLUT9 does transport essentially urate (Caulfield et al., 2008). GLUT9 mediates urate transport, which is independent of sodium, chlorine and anions, but voltage-dependent (Anzai et al., 2008). Compared with the mutation in URAT1, the complete loss of GLUT9 results in the net secretion of urate (Preitner et al., 2009). ABCG2 is a multidrug resistance transporter that is also implicated as an important urate transporter. Its gene variation has become the main cause of elevated sUA levels (Dehghan et al., 2008). ABCG2-mediated loss or reduction of renal urate secretion will lead to increased renal urate reabsorption (Woodward et al., 2009). In our review, more than one-third of the studies (72/186, 39%) show that Chinese herbal medicines target uric acid transporters. Alpinia oxyphylla seed extract enhances UA excretion in the kidney by reducing URAT1 and up-regulating OTA1 (Lee et al., 2019b). Wuling San down-regulates mRNA and protein levels of URAT1 and GLUT9, as well as up-regulates OAT1 in the kidney of hyperuricemic mice. Moreover, Wuling San also up-regulates organic cation/carnitine transporters which are associated with impaired renal function, leading to kidney protection (Ding et al., 2013). Gypenosides, natural saponins extracted from Gynostemma pentaphyllum, significantly lower sUA levels by reducing XO and increasing in urate excretion through regulating URAT1, GLUT9, and OAT1 (Pang et al., 2017).

Herbs Lowering Serum Uric Acid Resolve Inflammation
Uric acid belongs to the damage-associated molecular patterns, altered metabolites of necrotic or stressed cells that the innate immune system sees as an alarm signal (Patel, 2018). Elevated UA levels alters the physiology, boosting the expressions of inflammatory proteins by triggering complex proinflammatory cascades that damage cells and tissues (Chen and Lan, 2017). Clinical trials have shown that serum levels of IL-6 and TNF-α are significantly higher in hyperuricemia patients than in healthy people, and that of IL-6 and TNF-α are significantly increased as sUA levels increase (Zhou et al., 2018b). SUA above 9 mg/dl is associated with a gouty arthritis incidence of 4.9%. The accumulation of monosodium urate (MSU) crystals induce a mass of inflammatory cells (such as neutrophils and monocytes) to infiltrate into the site of MSU crystal deposits in patients, resulting in an acute inflammatory response, manifested as acute gout flares (Dalbeth et al., 2016). IL-1 released from these immune cells further triggers the release of various pro-inflammatory cytokines and chemokines, such as IL-8, IL-6 and TNF-α, which can further enhance neutrophil recruitment (El Ridi and Tallima, 2017). Inflammatory cytokines, especially IL-1β, are the key mediators of gouty inflammation. A phase III, international safety study of patients with acute gout arthritis treated with rilonacept (an IL-1 blocker) for 16 weeks shows that rilonacept significantly reduces the risk of gout attacks by 70.3% (Sundy et al., 2014). In experimental studies included in this review, 37 articles (11 active ingredients, 13 natural products, and 13 herbal formulas) describe the effects of these herbs on IL-1. It is worth mentioning that these herbs not only act on IL-1, but also as XO, UA transporter, other inflammatory factors such as IL-6, IL-8, TNF-α, NF-κB, NLRP3, caspase 1, etc.
There are 16 herbs included in this review that interfere with the NLRP3 inflammasome. Consistent with reported studies, elevated UA can be effectively reduced by regulating NLRP3 inflammasome -IL-1 pathway (Dhanasekar and Rasool, 2016;Szekanecz et al., 2019). Acute gout is an inflammatory response to MSU crystals. Innate immune pathways are essential in the pathogenesis of gout, particularly the activation of NLRP3 inflammasome, which leads to the release of IL-1β and other pro-inflammatory cytokines (So and Martinon, 2017). MSU crystals must first be coated with serum proteins and then interact with articular cell's surface membrane directly or via receptors, to stimulate a cytosolic molecular platform involved in innate immunity and promote the assembly and activation of the NLRP3 inflammasome (El Ridi and Tallima, 2017). NLRP3 inflammasomes are formed by the recruitment of the apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), and subsequent recruitment of caspase-1. Caspase-1 activates pro-inflammatory cytokines IL-1β and IL-18 by cleaving their respective precursor proteins, pro-IL-1β and pro-IL-18 (So and Martinon, 2017). Neutrophils are recruited and activated in response to the spillover of IL-1, producing ROS, proteolytic enzymes, pro-inflammatory chemokines, cytokines and so on, which recruit and activate macrophages (El Ridi and Tallima, 2017). Thus, the release of IL-1β mediated by inflammasome and the rapid recruitment of neutrophils lead to acute inflammatory episodes in gout patients. It is noteworthy that hyperuricemia may stimulate inflammatory leukocytes through epigenetic modification such as histone methylation, even without MSU crystals, thereby increasing production of IL-1β, IL-6, and TNF-α (Wasilewska et al., 2012;Zhou et al., 2018b).
The activation and maturation of IL-1β in response to endogenous and exogenous stimuli are also involved in the NF-κB pathway (Chen and Lan, 2017). In our review, there are 13 articles focused on NF-κB signaling pathway, showing the inhibitory effect of herbs on NF-κB activation under elevated sUA levels. It has been proved that NF-κB can be activated by UA (Spiga et al., 2017). Its activation transcribes a large number of pro-inflammatory genes. In a resting state, NF-κB combines with IκBα (inhibitor of NF-κB kinase subunit α) form a dimer in the cytoplasm, after IκBα kinase is activated, it phosphorylates IκBα. Subsequently, NF-κB is transferred from the cytoplasm to the nucleus, leading to the transcription and expression of genes related to inflammation. Studies have shown that inhibition of NF-κB restrains inflammation and improves hyperuricemia or gout conditions Wang et al., 2019c). Interestingly, studies focused on NF-κB activity is also related to NLRP3. NF-κB is essential for the initiation, assembly and activation of NLRP3 inflammasome, which is also a key step in the release of inflammatory cytokines in MSU crystal-induced inflammation (So and Martinon, 2017). P38 mitogen-activated protein kinase (MAPK) is also involved in the inflammatory cascade of NF-κB. Uric acid activates NF-κB through MAPK signal pathway, thus leading to the release of inflammatory factors such as TNF-α (Tang et al., 2017).

Herbs Lowering Serum Uric Acid Protect Against Renal Fibrosis
Evidence suggests that UA levels can be used to predict the prognosis of chronic kidney disease and end-stage kidney disease. Elevated sUA level is an independent risk factor for kidney disease and lead to renal fibrosis . Mice with systemic GLUT9 knockout showed moderate hyperuricemia, excessive hyperaciduria and obstructive nephropathy, along with progressive inflammatory fibrosis (Preitner et al., 2009). Once UA exceeds the maximum amount the kidneys can excrete, it gets deposit in the kidney and firstly causes direct pathological damage to the kidney. Secondly, the deposition of UA in the kidney results in the accumulation of neutrophils and monocytes as well as the release of inflammatory factors, which lead to glomerulosclerosis and interstitial fibrosis and aggravate renal injury (Ye et al., 2018). Prevention and treatment of renal fibrosis is the best treatment for kidney diseases caused by elevated sUA. However, at present, modern medical treatments are not effective in reducing renal fibrosis and preventing the progression of diseases.
A total of 24 studies included in this review are focused on hyperuricemic nephropathy, suggesting that herbal medicines improve renal injury induced by elevated sUA levels by regulating renal fibrosis-related signal pathways. Fibrosis is usually associated with strong inflammatory reactions and immunocyte infiltration. Therefore, inhibition of inflammatory cytokines might be a potential method to prevent fibrosis. Vaticaffinol, one of the herbs included in this review, markedly down-regulates NLRP3, ASC, caspase-1, IL-1β, IL-18, IL-6 and TNF-α in hyperuricemic mice, thus significantly decreases sUA levels and improves kidney function (Chen et al., 2017b). Transforming growth factor-β1/Mothers against decapentaplegic homolog 3 (TGF-β1/ Smad3) signaling is the most potent fibrogenic factor in the regulation of renal interstitial fibrosis process (Loeffler and Wolf, 2015). Pterostilbene, also mentioned in this review, suppresses the activation of TGF-β1/Smad3 and protooncogene tyrosine-protein kinase Src/Signal transducer and activator of transcription 3 (Src/STAT3) signaling pathway as to decrease sUA level and alleviate renal fibrosis in hyperuricemic mice (Pan et al., 2019). These findings highlight the fact that herbal medicines may be the potential antifibrotic therapeutics for hyperuricemic nephropathy treatment.

Herbs Lowering Serum Uric Acid Modulate Oxidative Stress
In addition to the inflammatory process, oxidative stress is one of the early events related to the elevated sUA. Uric acid entering cells can rapidly induce oxidative stress (Ko et al., 2019;Yin et al., 2019). This state of oxidative stress is governed by the balance between ROS production and their elimination by antioxidants. Since the cell membrane is impermeable to urate anion, cellular concentration of urate depends on the specific transporter of urate and xanthine oxidoreductase (XOR). In mammals, this enzyme exists in two forms, xanthine dehydrogenase (XDH) and XO. XDH transfers electron to NAD + and generates NADH, and XO transfers electron to O 2 and generates oxidative stress (Isaka et al., 2016). In addition, XOR may lead to the production of the superoxide anion and nitric oxide, especially under low pH or hypoxia conditions (Godber et al., 2000;Battelli et al., 2019). It can be seen that the generation of UA mediated by XO is closely related to the production of ROS. On the other hand, antioxidant enzymes that scavenge ROS are ubiquitous, including superoxide dismutase, glutathione peroxidase and catalase, the changes of these enzymatic activities may lead to oxidative stress. However, in inflammatory diseases including hyperuricemia, the production of the superoxide anion is often faster than its removal by superoxide dismutase (Zamudio-Cuevas et al., 2015).
There is an increasing interest in using herbs to resolve the inflammatory conditions, including hyperuricemia, because they play an anti-inflammatory role by inhibiting the production of ROS, such as procyanidins, Shizhifang, Zisheng Shenqi decoction, quercetin, Modified Simiao decoction and other herbs included in this review. Antioxidants like quercetin inhibits inflammation in rat models of chronic MSU-induced arthritis by decreasing inflammatory mediators such as IL-1β . Other herbs mentioned above also show significant antioxidant effects. For example, Shizhifang effectively suppresses the NLRP3-ASC-caspase-1 axis through accommodating the ROS pathway, thereby alleviates potassium oxonate -induced hyperuricemia . In summary, the herbs that possess anti-oxidant effects may be developed as anti-inflammatory agents to treat inflammatory diseases such as gout and hyperuricemia.

Limitations and Perspectives From the Experimental Studies
As shown in Tables 3-5, there are fewer studies on the active ingredients of herbs than on the herbal extracts and herbal formulas. Aside from herbal formula, there can be hundreds of active ingredients in a single-flavored herb, making it difficult to identify the ones that actually mediate the therapeutic effects. This partly explains why multiple signaling pathways are involved in herbal treatments. Extraction of components that mediate the therapeutic effects is difficult. In general, safe and effective single-flavored herb can be screened out from literatures or experimental studies, and its effective active ingredients can be separated by pharmacological methods. Then, the mechanism of action of the drug can be explored based on the identified active ingredients from the herbs. In recent years, there are new approaches to identify the active ingredients of herbs and predict their targets, such as Systems Pharmacology. Researchers used Systems Pharmacology as a basis to screen out active ingredients from herbal formulas or herbs, and predict the targets of active ingredients, and finally verify them through in vivo and in vitro experimental studies (Zhou et al., 2016;Hong et al., 2017;Zhao et al., 2019). It really opens up new ideas for the studies of herbs and shortens the time to find out the mechanisms of action underlying the therapeutic effects of the herbs. However, with the increasing number of herbal studies using Systems Pharmacology, we can find that it also has disadvantages, such as limited Systems Pharmacology databases, data not updated, limited prediction results and so on. In view of this, it is still necessary to explore how to efficiently screen out the active ingredients of the herbs and identify their molecular targets.

CONCLUSION
In conclusion, the results of meta-analysis indicate that Chinese herbal medicines have potent therapeutic effects in lowering sUA levels. The signaling pathways involved in the sUA lowering effects include UA synthesis, UA transport, inflammation, renal fibrosis and oxidative stress. Further studies with sophisticated research design can further demonstrate the efficacy and safety of these Chinese herbal medicines in lowering sUA levels. Identification of the active ingredients and delineation of the underlying mechanisms of action can facilitate the clinical translation and application of these ingredients.

AUTHOR CONTRIBUTIONS
All the authors contributed sufficiently for their participation in the study, as follows: XZ and LZ conceived, designed, and supervised the study; LC, ZL, MW, JC, FL, and HL conducted the literature search, data extraction, data analysis and interpretation; QH, YY, and XZ appraised the articles; LC, ZL, and MW wrote the paper, HK modified the paper. All authors have read and approved the final manuscript.