Efficacy and Safety of Wuling Powder in the Treatment of Patients with Diabetic Nephropathy: A Systematic Review and Meta-Analysis

Background Wuling powder is a classical formula of traditional Chinese medicine (TCM), which is extensively applied to treat diabetic nephropathy (DN). However, there are no related reports on systematically evaluating the efficacy of Wuling powder in the treatment of DN. Targeted at this, this study was developed. Materials and Methods This study systematically searched related articles from nine databases, including PubMed, Cochrane Library, Embase, Web of Science, China Knowledge Infrastructure (CNKI), China Biomedical CD-ROM (Sino Med), Wanfang database, Vipers database (VIP), and the China Clinical Trials Registry website. The randomized controlled trials (RCTs) involving Wuling Power to treat DN were included, which were published from the established data of the above databases to March 2022. In addition, the language of the studies was not restricted. Studies were meta-analyzed using the RevMan 5.4 software given in the Cochrane Collaboration Network. The treatment efficacy was measured using the weighted mean differences (WMD) and 95% confidence intervals (CI). Results 24 studies were included for the final analysis. 24 h urine volume (WMD = 357.95; 95% CI [322.83, 393.06], p < 0.00001), 24 h urine protein quantification(24 h UPQ) (WMD = −1.30; 95% CI [−1.82, −0.78], p < 0.00001), serum creatinine (Scr) (WMD = −10.17; 95% CI [−11.13, −9.21], p < 0.00001), blood urea nitrogen (BUN) (WMD = −1.62; 95% CI [−2.30, −0.93], p < 0.00001), urinary albumin excretion rate (UAER) (WMD = −24.73; 95% CI [−35.46, −13.99], p < 0.00001), fasting blood glucose (FBG) (WMD = −0.63.95% CI [−0.97, −0.30], p = 0.002), glycated hemoglobin (WMD = −0.11; 95% CI [−0.30, 0.08], p=0.26), total cholesterol (TC) (WMD = −0.63; 95% CI [−1.23, −0.04], p=0.04), triglycerides (TG) (WMD = −0.46. 95% CI [−0.70, −0.23], p=0.0001), high-density lipoprotein cholesterol (HDL-C) (WMD = −0.32; 95% CI [0.03, 0.62], p=0.03), low-density lipoprotein cholesterol (LDL-C) (WMD = −0.57; 95% CI [−0.77, −0.37], p < 0.00001), and total effective rate (TER) (response ratio (RR) = 1.40; 95% CI [1.32, 1.48]; p < 0.00001) were concluded. The Wuling powder in the treatment of DN was statistically significant in all the above outcome indicators, and the efficacy of the treatment group was better than that of the control group. Conclusion The results of this study provided evidence for the clinical application of Wuling powder to treat the DN, but it had to be further validated in higher-quality clinical studies.


Introduction
Diabetic nephropathy (DN) is the most common and serious microvascular complication of diabetes mellitus (DM). It is clinically characterized by persistent albuminuria and/or progressive decline in glomerular filtration rate and microangiopathy, and it may result in end-stage renal disease in severe cases. erefore, it becomes one of the leading causes of death in patients with DM [1]. e number of patients with DM in China ranks first all over the world, and its incidence is increasing year by year [2,3]. In recent years, studies have shown that the incidence of nephropathy

Design.
e protocol for this systematic study was registered on the Open Science Framework (INPLASY) platform (https://inplasy.com/) (registration number: INPLASY202240071). It was implemented and executed according to the preferred reporting guidelines for systematic review and meta-analysis protocols [12]. e final report would be in line with the PRISMA recommendations for systematic view reporting of the medical interventions in meta-analysis [13].

Database and Literature Search.
is study systematically searched nine databases from their establishment data to March 2022, including PubMed, Cochrane Library, Embase, Web of Science, China Knowledge Infrastructure (CNKI), China Biomedical CD-ROM (Sino Med), Wanfang database, Vipers database (VIP), and China Clinical Trials Registry website. In addition, the patent databases were searched to exclude the clinical trials that were not published due to patent applications for Wuling powder (approval number: Z11020702).

Types of Research.
e selected RCTs were on the Wuling powder plus or minus formula for the treatment of DN.

Subjects of the Studies.
In the included studies, the subjects were patients who met the internationally recognized diagnostic criteria for DN at the time of the study, who had been definitively diagnosed with DN by a clinician, and who had excluded primary nephropathy and other causes of renal disease.

Interventions.
All patients in the treatment group were treated with Wuling powder plus or minus formula, while those in the control group received other hypoglycaemic drugs; or patients in both treatment and control groups received the same conventional diabetes medication and based on which patients in the treatment group took Wuling powder plus or minus formula. Studies with multiple interventions or where Wuling powder was not the primary intervention were excluded. ere were no requirements for the course of the disease, course of treatment, and dose of medication.

Exclusion Criteria.
e studies satisfying the below conditions had to be excluded: (1) duplicate publications; (2) reviews, summaries of expert experience, evaluative articles, or theoretical elaborations; (3) not RTCs, or animal studies; (4) nonclinical studies such as pharmacology; and (5) with incomplete documentation of data.

Literature Screening.
e RCTs were screened independently by two researchers to exclude those who failed to meet the inclusion criteria. After the elimination of the duplicates, the abstracts of the searched RCTs were read for initial screening to exclude the RCTs not meeting the inclusion criteria and all the RCTs were downloaded. Next, the full texts of these RCTs were read. Finally, the RCTs on Wuling powder for DN treatment that met the inclusion criteria were selected. Any different opinions between the two researchers were referred to a third party for adjudication.  e authors, year of publication, mean age, number of trials and controls, course of the disease, number of cases by gender, interventions, duration of treatment, and outcome indicators of the included RCTs were selected. en, the authors were contacted if the required data were incomplete. Two researchers crosschecked the information entered in the RCTs, and any disagreements were referred to a third party for adjudication.

Assessment on Risk of Bias.
e risk of bias of the included RCTs was assessed by using the Risk of Bias Assessment Tool recommended by the Cochrane Collaboration, in terms of seven aspects: (1) random sequence generation method; (2) allocation protocol concealment; (3) blindness to subjects and intervention providers; (4) blindness to outcome assessors; (5) completeness of outcome data; (6) selective reporting of study results; and (7) other biases. For each of the included RCTs, "high risk," "low risk," and "unclear risk" were assessed for each aspect [14]. In case of any disagreement on assessment results of the risk of bias, it should be discussed with a third researcher.

Data Analysis.
e RevMan 5.4 software provided by the Cochrane Collaboration Network was adopted for metaanalysis. Dichotomous variables were expressed with the response ratio (RR), and the continuous variables were expressed using mean differences (MD). e χ 2 test was performed for heterogeneity. If p > 0.1 and I 2 < 50%, the heterogeneity among all RCTs was low and the fixed effect model (FEM) was used for meta-analysis; while if p < 0.1 and I 2 ≥ 50%, the heterogeneity was statistically significant and the random effect model (REM) was used for the meta-analysis. p < 0.05 indicated a statistically significant difference.

Heterogeneity Assessment and Sensitivity Analysis.
If I 2 ≥ 50%, there was a statistical heterogeneity and the REM was adopted to analyse the data; while the FEM was adopted if the test for heterogeneity was not significant (I 2 < 50%). As there was a variation in heterogeneity, the sensitivity analysis or subgroup analysis was essential to explore the potential causes of heterogeneity and to exclude the RCTs with a high risk of bias, so as to ensure the robustness of results.

Subgroup
Analysis. Due to differences in heterogeneity, subgroup analysis was required to analyze the possible reasons for heterogeneity. e main subgroup analysis items included different ages, control treatments, duration of treatment, region, and safety.

Assessment of Publication Bias.
Publication bias was detected using a funnel plot. A significant asymmetry in the funnel plot meant a publication bias. Figure 2 showed the flowchart to screen the RCTs. 222 relevant publications were searched from various databases, and 100 duplicates were excluded after the initial screening. Next, 88 not satisfying the inclusion criteria were excluded. en, after the full texts were read carefully, the publications which were not RCTs, not related to the treatment of DN, and lacked details of the results were excluded. Finally, 24 RCTs  were included in this meta-analysis.

3.2.
Characteristics of the Included RCTs. 2,018 patients were included in the 24 RCTs, including 1,030 in the treatment group and 988 in the control group. e mean age of the patients ranged from 50.57 ± 15.17 years old to 69.58 ± 1.65 years old. e intervention for patients in all treatment groups was Wuling powder combined with conventional Western medicine. All 24 RCTs were conducted in China and published in 2003∼2021. e shortest and longest durations of treatment were 3 weeks and 12 weeks, respectively. Table 2 showed in detail the basic characteristics of the 24 RCTs.

Risk of Bias in the Included RCTs.
is meta-analysis investigated the risk of bias for all RCTs included. All research projects were randomized into a five-linger group and a control group. A comparison of the 24 RCTs revealed inconsistent randomization of treatment; 9 RCTs [21-23, 28, 30, 31, 34, 36, 38] were determined as low risk of bias because they generated random sequences by random number tables and random coin flips, while the remaining RCTs described only "random allocation" and therefore were determined to be an unclear risk of bias. In addition, 3 RCTs [31,34,36] reported allocation concealment, so they were classified as low risk of bias. In terms of performance bias, only 1 RCT [31] reported the double-blind trials, so the remaining 23 RCTs were classified as having a high risk of bias. In terms of reporting bias, only 1 RCT [34] had shedding of participant data and was, therefore, determined to have a high risk of bias. All RCTs were balanced at baseline examination and showed no other bias. e full and detailed analysis results on the risk of bias are shown in Figures 3 and 4. [16,17,24,25,32,37] involving 334 DN patients provided data on 24 h urine volume before and after the intervention. A heterogeneity test (P � 0.03 and I 2 � 59%) and a sensitivity analysis revealed that the statistical heterogeneity became lower after the study of Wang et al. [32] was removed (χ 2 � 5.16, p � 0.27, I 2 � 23%; Figure 5), so the FEM was used. e results showed that the 24 h urine volume was significantly higher for patients in the treatment group treated with Wuling powder combined with conventional western medicine (WMD � 357.95; 95% CI [322.83, 393.06], p < 0.00001; Figure 5). e subgroup analysis suggested that, in the different age subgroups (p � 0.01), the group <60 years (χ 2 � 3.52, p � 0.32, I 2 � 15%) was homogeneous with the group ≥60 years (χ 2 � 0.56, p � 0.45, I 2 � 0%), and the difference was statistically significant. e difference between different regional subgroups (p � 0.14) was not significant (Table 3, Supplementary Material Figures 1 and 2). [15-17, 19, 20, 23, 24, 26, 29, 31, 35-37] (including 1,054 patients) reported the data of 24 UPQ. Significant heterogeneity was found in these 13 RCTs (χ 2 � 760.27, p < 0.00001, I 2 � 98%; Figure 6), so the REM was used. e 24 h UPQ for patients treated with Wuling powder combined with conventional western medicine was higher than that in the control group (WMD � -1.30; 95% CI [−1.82, −0.78], p < 0.00001; Figure 6). Because of the large heterogeneity, the subgroup analysis was conducted based on the age, control treatment, duration of treatment, and region; and significant differences were found in intervention effects of different ages (p � 0.002), control treatments (p < 0.0001), and regions (p < 0.0001). It was reflected in the glipizide group (control treatment) (χ 2 � 0.1, p � 0.95, I 2 � 0%) and in Guangdong province (region) (χ 2 � 0.36, p � 0.55, I 2 � 0%), while significant heterogeneity was still observed in the rest (

Scr.
14 RCTs [15, 17-19, 21, 24, 27-29, 31, 35-38] (including 1,210 participants) reported the Scr data. Significant heterogeneity was found (χ 2 � 3628.59, p < 0.00001, I 2 � 100%; Figure 7), so the meta-analysis was conducted by using a REM. e analysis showed a statistically significant difference in Scr between the treatment and control groups , indicating that the Scr was significantly better in the treatment group. In addition, the subgroup analysis was conducted in different durations of treatment and regions, and different durations of treatment showed no statistically significant difference (p � 0.11) and different regions showed statistically obvious difference (p � 0.007), but significant heterogeneity was still observed (  [15,19,21,24,27,28,31,35,36,38] (including 965 patients) reported the BUN data. e tests showed significant heterogeneity (χ 2 � 137.59, p < 0.00001, I 2 � 93%; Figure 8), so the meta-analysis was conducted by using a REM. e analysis showed a statistically significant difference in BUN between the treatment and control groups (WMD � −1.62; 95% CI [−2.30, −0.93], p < 0.00001; Figure 8), indicating that the BUN was significantly better in the treatment group. Subgroup analysis in different durations of treatment and regions showed no significant difference in intervention effects between the two groups (p � 0.26 and 0.41 respectively), but significant heterogeneity was still observed (  [18,20,21,27,28,38] (including 482 patients) reported on UAER. Tests showed significant heterogeneity (χ 2 � 128.55, p < 0.00001, I 2 � 96%; Figure 9), so a REM was used. A statistically significant difference was       Figure 9), indicating that the UAER was significantly better in the treatment group. A subgroup analysis suggested the intervention effect was not significantly different in age (p � 0.11) and duration of treatment (p � 0.93) but showed an obvious difference in regions (p � 0.02), with no heterogeneity in the Guangdong region (χ 2 � 0.24, p � 0.62, I 2 � 0%) and the rest still observed significant heterogeneity (  Figure 10), so a REM was adopted for metaanalysis. e analysis results revealed a statistically significant difference in FBG between the treatment and control groups (WMD � −0.63; 95% CI [−0.97, −0.30], p � 0.002; Figure 10), suggesting that Wuling powder combined with conventional western medicine may significantly reduce the FBG in DN patients. Based on different ages, control treatments, durations of treatment, regions, and safety, the subgroup analysis was conducted, and the results showed no significant differences in intervention effects (p � 0.49, 0.45, 0.49, 0.67, and 0.78, respectively); no heterogeneity was found in the ≥60 years group (χ 2 � 0.26, p � 0.61, I 2 � 0%) in the age subgroup and in Guangdong Province (χ 2 � 0.26, p � 0.61, I 2 � 0%) in the region, while significant heterogeneity was still observed in the rest (  Figure 12). e REM analysis results showed the TC was statistically significant (WMD � −0.63; 95% CI [−1.23, −0.04], p � 0.04; Figure 12). In the subgroup analysis, the differences in age, control treatment, duration of treatment, and region (p � 0.06, 0.22, 0.05, and 0.99, respectively) were not statistically significant (  Figure 13). e REM analysis results showed (WMD � −0.46; 95% CI [−0.70, −0.23], p � 0.0001; Figure 13) the differences in TG between the two groups were statistically significant. In the subgroup analysis, the differences were not statistically significant in age, control treatment, and duration of treatment (p � 0.08, 0.11, and 0.7, respectively), were statistically significant in regions (p < 0.00001), and not heterogeneous in Guangdong Province (p � 0.69, I2 � 0%) (  Figure 14), indicating high heterogeneity, and using a random effects model, the results showed (WMD � −0.32; 95% CI [0.03,0.62], p � 0.03; see Figure 14), the difference between the two groups was statistically significant. In the subgroup analysis, the differences were statistically significant across sessions and regions (all p � 0.0002) ( (4) 4 RCTs [18,21,28,29] (314 patients) recorded LDL values, and the heterogeneity among them was proved to be high (χ 2 � 10.28, p � 0.02, I 2 � 71%; Figure 15), which became lower after the study by Chen et al. [21] was removed (χ 2 � 3.34, p � 0.19, I 2 � 40%; Figure 15), so the  In summary, Wuling powder showed better performance compared with the conventional methods in the treatment of DN in terms of TC, TG, HDL-C, and LDL-C.

Adverse Effects.
Adverse reactions were reported in 2 out of 24 RTCs. e adverse reactions reported in the study by Shen and Shu [23] were nausea, vomiting, abdominal distension, diarrhea, skin rash, and mild hypoglycemia; while those reported by Jing et al. [28] included nausea, vomiting, back pain, skin pruritus, swelling of lower limbs, thirst, and excessive drinking. No adverse reactions were reported in the remaining RCTs.

Certainty of Evidence.
e GRADEpro was employed to assess the certainty of the evidence in this study. Table 4 showed that the results of 24 h urine output, LDL, and TER were moderate-quality evidence, while other outcomes were low-quality evidences. e high heterogeneity of some outcomes, the low methodological quality, and the high risk of bias were reasons for the poor quality of the evidence. erefore, Wuling powder should be considered cautiously in the clinical use for DN treatment.

Discussion
4.1. Results. As one of the common vascular disease complications in DM patients, DN seriously affects the prognosis of patients and should be treated as early as possible. In recent years, TCM has become an essential adjunctive drug treatment for most Chinese patients with DN due to its stable efficacy and low side effects. Many studies confirm that the combination of Wuling powder with conventional symptomatic supportive treatment for DN is effective in      is meta-analysis study provided a sound theoretical basis for the application of Wuling powder in the treatment of DN. erefore, the results of this study may provide an important reference for the adjuvant treatment of DN with TCM.
Although our results were statistically significant, some of the results were subject to greater heterogeneity. e outcome markers were divided into subgroups based on characteristics of the patients such as different age, control treatment, duration of treatment, and region for comparison to seek reasons for heterogeneity. In the subgroup analysis, 24 h urine volume reduced heterogeneity after different age subgroup analysis, 24 h urine protein quantification reduced heterogeneity after control treatment and regional subgroup analysis, urine albumin excretion rate reduced heterogeneity after regional subgroup analysis, FBG reduced heterogeneity after age and regional subgroup analysis, and glycated hemoglobin and TG both reduced heterogeneity after regional subgroup analysis. It is worth noting that most of the outcome indicators showed lower heterogeneity in the Guangdong region, which may be related to the origin of the herbs. In contrast, significant heterogeneity was observed in Scr and BUN. e subgroup analysis was performed and there were no significant differences in intervention effects between groups, and the size of such heterogeneity was not reduced following the use of a REM.
It is believed that these heterogeneities arise from the following points. Firstly, the reasons for the large heterogeneity are most likely related to the variety, origin,  Evidence-Based Complementary and Alternative Medicine harvesting season, storage and processing, dosage form, and route of administration of the herbal medicines. Such contents are not described in detail in the literature, So they could not be analyzed further in this study. Secondly, only 24 relevant studies were included in this study, and most of them only mentioned the word simple randomisation without considering the specific implementation methods, which affected the scientific validity of the study results. In addition, three studies [31,34,36] reported allocation concealment and only one study [31] reported the use of a double-blind trial. e rest studies did not report the allocation concealment and were unblinded, which was susceptible to a variety of artifacts and may lead to heterogeneity with different study participants and various interventions. e randomization grouping resulted in selective bias, which could reduce the overall quality of the    Total (95% CI) Heterogeneity: Tau 2 = 0.03; Chi 2 = 11.16, df = 5 (P = 0.05); I 2 = 55% Test for overall effect: Z = 1. 13   meta-analysis. Furthermore, some of the included studies did not mention the methods of testing for outcome indicators, and there were uncontrollable factors such as different experimental instruments, which affected the objectivity of the results. In addition, one study [34] had a shedding of participant data, which may affect the final analysis of the results. Meanwhile, some of the outcome indicators were combined despite high heterogeneity, which affected the reliability of the study. Taken together, these may have contributed to the high heterogeneity of some of the outcome indicators.
In addition, many other factors were evident in this study in terms of their impacts on the results. Firstly, the studies included in this study were limited to English and Chinese, and the final analysis was conducted on all Chinese literature, which would result in a potential publication bias.            Evidence-Based Complementary and Alternative Medicine kind of Western medical treatment was adopted. In addition, whether the dose was controlled strictly would also be another factor resulting in publication bias. Clinical trials are concerned with the follow-up of patients' long-term outcomes. Most of the included studies were limited to a shortterm treatment after the drug intervention, which also impacts the bias in outcome efficacy. In the subgroup analysis, the cut-off points for age and duration of disease were mainly based on relevant studies, however, more biological support is needed. Finally, the funnel plot showed the publication bias in this study, which may be due to the ease of publishing positive results and the difficulty of publishing negative results, severely limiting the validity and objectivity of the efficacy of Wuling powder to treat DN. In addition, the results of the GRADE analysis showed that the reliability of the outcome indicators was mostly low to moderate. erefore, Wuling powder could be cautiously recommended as an adjunctive treatment for DN.

Strengths and Limitations.
e strengths of this study could be summarized as follows. Firstly, this was probably the first meta-analysis to assess the efficacy and safety of Wuling powder in the treatment of DN. Secondly, all the literature included were RCTs, ensuring the credibility of the results of this study. irdly, the results of this study provided a new therapeutic option for the treatment of DN. e results of this study suggested that the combination of the Wuling powder with conventional treatment for DN had positive clinical implications, which was superior to the Western medicine treatment alone. It implied that Wuling powder may enhance the effectiveness of conventional treatment and improve the overall clinical outcome, reflecting the uniqueness and superiority of TCM. Due to the holistic treatment theory, the use of TCM in the adjunctive treatment of disease is increasingly reported and researched. It is found that TCM can play a better therapeutic advantage in the treatment of both DN and its complications, and exert a positive effect on the safety, suffering reduction, and life improvement of patients. Systematically assessing the efficacy of TCM in DN and providing corresponding evidencebased medical evidence are of high significance to promote the TCM culture worldwide and search for new breakthroughs in the treatment of DN patients.
However, there are some limitations to this study. Firstly, it was limited by the quality of the literature. Most of the studies included in the study did not report allocation concealment and the use of blinding, leading to the measurement and implementation of various biases. Secondly, the included studies were RCTs with small samples and were of low quality. irdly, the lack of DN staging in most of the   Evidence-Based Complementary and Alternative Medicine studies in this study affected the effectiveness of Wuling powder in patients with different degrees of DN. In addition, the lack of a placebo prevented us from analysing the difference in efficacy between using and not using Wuling powder. Finally, the patients in this study were all selected from the Chinese region and may not be globally representative, with some degree of clinical bias applied.

Conclusion
In conclusion, Wuling powder combined with conventional drugs showed outstanding efficacy and positive effect in the treatment of DN. However, there were still some limitations in this systematic evaluation, so applying Wuling powder in clinical treatment should be considered cautiously. erefore, some clinical studies with larger samples, higher study quality, and more rigorous study design should be taken in the future to validate the accurate and objective assessment of DN, and then obtain more valuable meta-analysis results, providing more reliable and effective new ideas for the treatment of DN.

TCM:
Traditional Chinese medicine DN: Diabetic nephropathy CNKI: China knowledge infrastructure Sino Med: China biomedical CD-ROM VIP: Vipers database RCTs: Randomized controlled trials WMD: Weighted mean differences CI: Confidence intervals 24 h UPQ: 24 h urine protein quantification Scr: Serum creatinine BUN: Blood urea nitrogen UAER: Urinary albumin excretion rate FBG: Fasting blood glucose TC: Total cholesterol TG: Triglycerides HDL-C: High-density lipoprotein cholesterol LDL-C: Low-density lipoprotein cholesterol TER: Total effective rate RR: Response ratio DM: Diabetes mellitus MD: Mean differences FEM: Fixed effect model REM: Random effect model.

Data Availability
All data relevant to the study are included in the article.

Conflicts of Interest
e authors declare that there are no conflicts of interest regarding the publication of the paper.

Authors' Contributions
YYY, LT, and CL conceived the study. YYY, SWJ, HKK, XYY, TSF, and YHP conducted the literature search and data extraction. YYY and SWJ analysed the data, performed the statistical analysis, and wrote the manuscript. LT and CL participated in the correction of the manuscript and supervised every step of the study. All authors read and approved the final manuscript and decided to publish it. LT and CL equally contributed to this work and should be considered cocorresponding authors.

Supplementary Materials
Supplementary Table 1