To investigate the mechanisms by which Jian-Pi-Yi-Shen Recipe (JPYSR) affects chronic kidney disease (CKD) progression, focusing on stress granules (SGs), lysosomal integrity and autophagy regulation. A CKD model was induced in mice using a 0.2% adenine diet and treated with JPYSR (15.60 g/kg/day) via gavage. Renal function was assessed using serum creatinine (Scr) and blood urea nitrogen (BUN) levels. Pathological changes were evaluated using PAS and Masson's trichrome staining. Protein and gene expressions were analyzed using Western blot, qPCR, immunohistochemistry and immunofluorescence. In vitro studies were conducted on human renal tubular epithelial cells (HK2). Statistical analyses were performed using GraphPad Prism (version: 9) software. CKD progression was associated with lysosomal impairment and reduced autophagy. JPYSR treatment significantly improved renal function, reduced pathological changes, decreased renal fibrosis, promoted SG formation, alleviated lysosomal damage and maintained baseline autophagy. Inflammation was also diminished, as confirmed by in vitro experiments. JPYSR may slow CKD progression and reduce renal fibrosis by modulating SG formation, lysosomal function and autophagy levels.

The diagnosis and treatment of chronic glomerulonephritis (CGN) during childhood pose distinct challenges. Acteoside (ACT) is the primary active ingredient extracted from the leaves of Rehmannia glutinosa by our research group, accounting for 2.15 %, which possess multiple biological activities, especially for nephropathy treatment. However, its mechanism intervention on CGN in children remains obscure. In this study, we established a model of purinomycin aminonucleoside (PAN)-induced CGN in childhood rats to assess the potential therapeutic effect and underlying mechanisms of ACT. Leveraging network pharmacology and multi-omics technology, we delved into the effects and mechanisms of ACT intervention on CGN. And these findings were further corroborated through qRT-PCR, western blot and targeted metabolomics. Our results demonstrated that ACT had significantly efficient in the treatment of CGN in childhood rats by improving the key indicators and pathological changes. Further, ACT could significantly regulate differences in endogenous small molecules and genes based on non-target metabolomics and transcriptomics. Meanwhile, target capture analysis found the crucial targets of ACT treatment in CGN. Integrated analysis of multi-omics study indicated that PI3K/Akt signaling pathway and its downstream amino acid metabolism were significantly enriched, hinting at the essential regulatory pathway for ACT in treating of CGN. Finally, through qRT-PCR, western blot and targeted metabolomics, it was verified that ACT could ameliorate CGN through Cxcr4-PI3K-Akt-eNOS signaling pathway, thereby regulating amino acid metabolism. The collective results were consistent with those of multi-omics analysis. Our study illuminated that ACT had notable curative effect on CGN rats, and preliminarily elucidated its mechanism of action. Our research will provide solid basis for the treatment of chronic glomerulonephritis in children with ACT and developing it into innovative traditional Chinese medicine.

This study aims to comprehensively analyze the material basis of toad visceral oil(hereafter referred to as toad oil), and explore the pharmacological effect of toad oil on atopic dermatitis(AD). Ultra-high performance liquid chromatography-linear ion trap/orbitrap high-resolution mass spectrometry(UHPLC-LTQ-Orbitrap-MS) and gas chromatography-mass spectrometry(GC-MS) were employed to comprehensively identify the chemical components in toad oil. The animal model of AD was prepared by the hapten stimulation method. The modeled animals were respectively administrated with positive drug(0.1% hydrocortisone butyrate cream) and low-and high-doses(1%, 10%) of toad oil by gavage. The effect of toad oil on AD was evaluated with the AD score, ear swelling rate, spleen index, and pathological section results as indicators. A total of 99 components were identified by UHPLC-LTQ-Orbitrap-MS, including 14 bufadienolides, 7 fatty acids, 6 alkaloids, 10 ketones, 18 amides, and other compounds. After methylation of toad oil samples, a total of 20 compounds were identified by GC-MS. Compared with the model group, the low-and high-dose toad oil groups showed declined AD score, ear swelling rate, and spleen index, alleviated skin lesions, and reduced infiltrating mast cells. This study comprehensively analyzes the chemical composition and clarifies the material basis of toad oil. Meanwhile, this study proves that toad oil has a good therapeutic effect on AD and is a reserve resource of traditional Chinese medicine for external use in the treatment of AD.

Bioenergetic therapy based on tumor glucose metabolism is emerging as a promising therapeutic modality. To overcome the poor bioavailability and toxicity of arenobufagin (ArBu), a MOF-derived intelligent nanosystem, ZIAMH, was designed to facilitate energy deprivation by simultaneous interventions of glycolysis, OXPHOS and TCA cycle. Herein, zeolitic imidazolate framework-8 was loaded with ArBu and indocyanine green, encapsulated within metal-phenolic networks for chemodynamic therapy and hyaluronic acid modification for tumor targeting. ZIAMH nanoparticles can release ArBu in the tumor microenvironment for chemtherapy, and ICG enables photothermal therapy under near-infrared laser irradiation. In vitro and in vivo mechanism studies revealed that the ZIAMH nanoplatform downregulated glucose metabolism related genes, resulting in the reduction of energy substances and metabolites in tumors. Additionally, it significantly promoted cell apoptosis by upregulating pro-apoptotic proteins such as Bax, Bax/Bcl-2, cytochrome C. Animal studies have shown that the tumor inhibition efficiency of ZIAMH nanomedicines was three fold higher than that of free drugs. Therefore, this study provides a new strategy for glucose metabolism-mediated bioenergetic therapy and PTT/CDT/CT combined therapy for tumors.

Microbiota-gut-brain axis communication represents another crucial pathway in the pathogenesis of Alzheimer's disease (AD), whereby gut microbiota significantly impacts AD pathology by modulating immune, metabolic, digestive, and neurological functions. Although research on treating AD through gut microbiota interventions is advancing, substantial breakthroughs remain limited. Given AD's complex pathological mechanisms, Traditional Chinese Medicine (TCM) presents a clear advantage with its multi-target effects. During the processes of TCM intake, absorption, and therapeutic action, the gut microbiota serves both as a mediator and as a therapeutic target. However, the mechanisms by which TCM interacts with gut microbiota to exert beneficial effects on AD remain largely unclarified. Here, we review the mechanisms through which TCM may intervene in AD from the perspective of gut microbiota, examining the potential mechanisms and clinical application prospects of Chinese herbal medicine in regulating the gut microbiome. This provides a novel theoretical foundation and methodological support for further research into herbal therapies for AD.

BACKGROUND:Although Yinhua Miyanling tablet has demonstrated promising efficacy in managing uncomplicated urinary tract infections (UTIs), robust clinical evidence from large-scale studies remains scarce. This systematic review aims to comprehensively evaluate the therapeutic effectiveness and safety profile of Yinhua Miyanling tablet in the treatment of uncomplicated UTIs, providing an evidence-based assessment for clinical practice. METHODS:A comprehensive computerized search was conducted across multiple databases, including China National Knowledge Infrastructure, China Science and Technology Journal Database, Wanfang Data, Chinese Biomedical Literature Database, Big Medical Knowledge Base, Medline, PubMed, Embase, Cochrane Library, and Web of Science. The search period spanned from the inception of each database to October 2023, aiming to identify randomized controlled trials (RCTs) investigating the efficacy of Yinhua Miyanling tablets in treating uncomplicated UTIs. The selected studies were subjected to meta-analysis using RevMan 5.3 software (The Cochrane Collaboration, London, United Kingdom). The methodological quality of the included literature was assessed using the Cochrane Risk of Bias Tool (version 5.1.0; The Cochrane Collaboration, London, United Kingdom) and the modified Jadad scale. Additionally, the GRADE system was employed to evaluate the overall quality of evidence. RESULTS:A total of 14 RCTs, involving 1773 patients, were included in the meta-analysis. The results demonstrated that Yinhua Miyanling tablets, whether used alone or in combination with Western medicine, significantly improved clinical efficacy and bacterial clearance rates while reducing recurrence rates (all P < .05). Notably, there was no statistically significant difference in adverse reactions between the treatment and control groups, suggesting a favorable safety profile. CONCLUSION:The Yinhua Miyanling tablet demonstrates therapeutic potential in managing uncomplicated UTIs. However, the current evidence is limited by the relatively low methodological quality of included studies. Further validation through large-scale, multicenter RCTs with rigorous designs is warranted to strengthen these findings. According to the Grading of Recommendations Assessment, Development and Evaluation evidence assessment, the available data provide moderate confidence in the efficacy of Yinhua Miyanling tablet for treating uncomplicated UTIs.

BackgroundJianwei Xiaoshi oral liquid (JWXS), a classical traditional prescription comprising various edible medicinal plants, has demonstrated significant efficacy in treating paediatric indigestion. It originates from Jianpi Pill, which is developed in the Ming Dynasty and nourishes the spleen and regulates gastrointestinal function. However, the specific molecular mechanisms involved remain unclear.MethodsTo elucidate the material base of JWXS and its underlying mechanism in treating dyspepsia, the UHPLC-Q-Orbitrap HRMS method and network pharmacology were utilized. This was followed by pharmacological experiments, transcriptomics analyses and gut microbiota studies to further investigate the effects of JWXS on dyspepsia.ResultsA total of 105 compounds, mainly flavonoids, alkaloids, organic acids and cyclic peptides, were identified. According to the five principles of generic drug properties, 43 candidate compounds were screened out. Their efficacy was verified through gastric emptying and intestinal propulsion experiments. Transcriptomic analysis revealed that JWXS primarily alleviated dyspepsia symptoms by regulating the secretion of 8 key proteins in the pancreatic secretion pathway. The differences in the gut microbiota, as identified through 16S rRNA and ITS2 sequencing, were subsequently more pronounced than those observed in the bacterial microbiota of the model group. In total, 15 differential bacteria and 16 differential fungi were identified. Targeted metabolomics analysis of SCFAs revealed a significant decrease in valeric acid (VA), acetic acid (AA), and isovaleric acid (IVA) levels in the model group, which were restored to the corresponding levels after the administration of JWXS. Correlation analysis revealed that VA, AA, and IVA were positively correlated with Lactobacillus and Bacteroides, and negatively correlated with Aspergillus and Candida. This further suggested that JWXS might alleviate symptoms of indigestion by regulating the composition of the microbiota, increasing the variety and quantity of beneficial bacteria, reducing fungal contamination, and further increasing the levels of SCFAs in the body.ConclusionJWXS improved functional dyspepsia in immature rats via a mechanism involving the regulation of the secretion of 8 key proteins in the pancreatic secretion pathway and the amelioration of flora disorders.

Background Huashi Baidu granule (HSBD), an approved herbal formula for treating COVID-19, demonstrates safety and efficacy. Despite its market approval, the detailed methodology and identification of its active components remain unexplored, leaving its bioactive constituents and action mechanisms unclear. Methods This study investigated the potential mechanisms of HSBD’s active ingredient in treating COVID-19. Our approach integrated various techniques, including the UHPLC-QqQ-MS/MS method, analysis of the GEO database, network pharmacology, surface plasmon resonance, molecular docking and molecular dynamics simulations, to formulate a comprehensive research strategy. Results The UHPLC-QqQ-MS/MS method employed for HSBD analysis proved stable, reliable, and reproducible. We identified 25 principal components in HSBD, with 7 compounds detected in plasma, namely pogostone, p-hydroxybenzoic acid, Paeoniflorin, Rhein, Emodin, ephedrine hydrochloride, and pseudoephedrine hydrochloride. Protein-Protein Interaction (PPI) network analysis identified MMP9 as a pivotal target. Surface plasmon resonance analysis revealed that Paeoniflorin and Rhein exert their antiviral effects by interacting with RBD and ACE2. In contrast, Emodin’s antiviral mechanism predominantly involves binding to MMP9. Molecular docking results indicated strong binding affinities of Rhein and Paeoniflorin to the hACE2 protein, and high binding affinities of Emodin to the MMP9 protein, all of which were corroborated by molecular dynamics simulations. Conclusion We investigated the methodology and identified the active components of HSBD, focusing on those absorbed into the plasma, to elucidate the effective material basis of HSBD in the treatment of COVID-19, our research offered insightful exploration into its mechanisms of action against COVID-19.

This study explored the biosynthesis of bufadienolides(BDs) in Bufo bufo gargarizans to solve the dilemma of the decreasing resources of B. bufo gargarizans and provide a theoretical basis for the sustainable utilization of the resources. Ultra-high performance liquid chromatography-Orbitrap-mass spectrometry(UHPLC-Orbitrap-MS) was employed to detect the synthesis sites of BDs in B. bufo gargarizans, and the results were verified by desorption electrospray ionization-mass spectrometry imaging(DESI-MSI) and homogenate incubation experiments. BDs in B. bufo gargarizans had the highest content in the liver and the highest concentration in the gallbladder, in addition to the parotid gland and skin, which suggested that the liver could synthesize BDs. The results of DESI-MSI also showed that BDs were mainly enriched in the liver rather than the immature parotid gland. The incubation experiment of liver homogenates demonstrated the liver of B. bufo gargarizans had the ability to synthesize BDs. This study showed that the liver was a major organ for the synthesis of BDs in B. bufo gargarizans during metamorphosis, development, and growth, which provided strong theoretical support for the biosynthesis of BDs and the sustainable utilization of B. bufo gargarizans resources.

Background: Diabetic nephropathy (DN) was one of the most popular and most significant microvascular complications of diabetes mellitus. Qingxin Lianzi Yin Decoction (QXLZY) was a traditional Chinese classical formula, suitable for chronic urinary system diseases. QXLZY had good clinical efficacy in early DN, but the underlying molecular mechanism remained unrevealed. Purpose: This study aimed to establish the content determination method of QXLZY index components and explore the mechanism of QXLZY on DN by network pharmacology and metabolomics studies. Methods: Firstly, the content determination methods of QXLZY were established with calycosin-7-O-beta-d-gluco- side, acteoside, baicalin and glycyrrhizic acid as index components. Secondly, pharmacological experiments of QXLZY were evaluated using db/db mice. UHPLC-LTQ-Orbitrap MS was used to carry out untargeted urine metabolomics, serum metabolomics, and kidney metabolomics studies. Thirdly, employing network pharmacology, key components and targets were analyzed. Finally, targeted metabolomics studies were performed on the endogenous constituents in biological samples for validation based on untargeted metabolomics results. Results: A method for the simultaneous determination of multiple index components in QXLZY was established, which passed the comprehensive methodological verification. It was simple, feasible, and scientific. The QXLZY treatment alleviated kidney injury of db/db mice, included the degree of histopathological damage and the level of urinary microalbumin/creatinine ratio. Untargeted metabolomics studies had identified metabolic dysfunction in pathways associated with amino acid metabolism in db/db mice. Treatment with QXLZY could reverse metabolite abnormalities and influence the pathways related to energy metabolism and amino acid metabolism. It had been found that pathways with a high degree were involved in signal transduction, prominently on amino acids metabolism and lipid metabolism, analyzed by network pharmacology. Disorders of amino acid metabolism did occur in db/db mice. QXLZY could revert the levels of metabolites, such as quinolinic acid, arginine, and asparagine. Conclusion: This study was the first time to demonstrate that QXLZY alleviated diabetes-induced pathological changes in the kidneys of db/db mice by correcting disturbances in amino acid metabolism. This work could provide a new experimental basis and theoretical guidance for the rational application of QXLZY on DN, exploring the new pharmacological effect of traditional Chinese medicine, and promoting in-depth research and development.

Background Vascular calcification (VC) significantly raises cardiovascular mortality in chronic kidney disease (CKD) patients. VC is characterized by the phenotypic transformation of vascular smooth muscle cells (VSMCs) to osteoblast-like cells, mediated by exosomes derived from calcified VSMCs and the exosomal microRNAs (miRNA) which may trigger some signals to recipient VSMCs. Bushen Huoxue (BSHX) formula has demonstrated its clinical efficacy in CKD and its protective role in CKD-VC rats has also been observed. However, little is known about its underlying mechanism.Methods To establish a VC model, aortic VSMCs from rats were induced to osteogenic differentiation by high-level phosphate (HP) in vitro. The expression of exosome and calcification makers were analyzed by western blot, including CD9, CD63, alpha-SMA, BMP-2, and Runx2, respectively. Differential expression of exosomal miRNAs in normal and HP-induced VSMCs were identified by using whole miRNA microarray technology. GO and KEGG analyses were performed to determine the significant enrichment of functions and signaling pathways in the target genes. In vivo, the CKD-VC rat model was established by administering adenine gavage combined with a high phosphorus diet. The rats were divided into normal control, model, low-dose BSHX, medium-dose BSHX, high-dose BSHX groups, and sevelamer groups. The blood biochemical parameters were measured. Renal histopathology and aortic calcification were observed. Western blot detected the levels of the calcification markers. Quantitative real-time PCR (qPCR) assay detected exosomal microRNA-32 (miR-32) mRNA expression in the aorta, the most differentially expressed exosomal miRNA previously identified. Phosphatase and tensin homolog located on chromosome ten (PTEN)/phosphatidylinositol-3 kinase (PI3K)/protein kinase B (AKT) signaling pathway components were also tested by western blot.Results Exosomal miRNA-32 and PI3K/AKT signaling pathways were highly differentially expressed between normal and HP-induced VSMCs. In vivo, BSHX improved blood biochemical parameters, renal histopathology, and aortic calcification in CKD-VC rats. BSHX increased the expression level of alpha-SMA and decreased the level of BMP-2 and Runx2. BSHX also lowered the expression level of exosomal miR-32 mRNA, enhanced PTEN expression, therefore, reduced p-PI3K and p-AKT levels in the aorta.Conclusion BSHX alleviated VC in CKD rats by downregulating exosomal miR-32 expression in the aorta, thereby promoting PTEN expression and inhibiting the PI3K/AKT signaling pathway.

Abstract Idiopathic pulmonary fibrosis (IPF) represents a chronic interstitial lung disease with an unclear underlying mechanism and currently lacks a definitive treatment. Cordyceps sinensis (CS), renowned for its pharmacological properties in traditional Chinese medicine and extensive use in lung disease treatment, holds promise as a therapeutic agent for IPF. However, the specific role of CS in treating IPF remains unclear. In this study, we aimed to assess the efficacy of CS in treating IPF and unravel potential underlying mechanisms. Our results demonstrate that CS treatment effectively mitigated pulmonary inflammation and collagen deposition in bleomycin‐induced IPF mice. Proteomics analysis revealed that the regulation of mitochondrial oxidative phosphorylation may serve as a potential protective mechanism of CS against IPF in mice. Further investigation unveiled that CS could suppress the excessive production of mitochondrial reactive oxygen species in lung tissues induced by bleomycin through moderating the expression and activity of mitochondrial complexes, thus safeguarding the integrity and function of mitochondria. Overall, our findings not only underscore the effectiveness of CS in preventing bleomycin‐induced IPF but also highlight mitochondrial‐mediated oxidative stress as a promising therapeutic target for treating IPF.

Background: The pathogenesis of metabolic syndrome was strongly associated with compromised metabolism homeostasis and gut microbiota imbalance. NAFLD is a progressive metabolic liver disease for which effective interventions are lacking. Bile acids exhibited appreciable metabolic regulatory effects and selective antimicrobial activity. Aim of the study: This study was designed to investigate the effect of BBBP, which mainly contained bile acids, on NAFLD from the perspectives of gut microbiota and metabolomics. Materials and methods: The present study was initiated on the anti-NAFLD effect of BBBP in HFD-fed mice. The efficacy of BBBP was evaluated by mice phenotypes, liver histopathological analysis and serum lipid and glucose levels. The activation of bile acid receptors such as Nr1h4, Nr1i2 and S1pr2 were detected by qRT-PCR analysis. Subsequently, untargeted metabolomics coupled with microbiomics was used to explore the mechanism of BBBP against NAFLD. Human L02 hepatocytes induced by OA and PA were used to investigate the effect of GABA on reducing lipid accumulation in vitro. Results: BBBP significantly and dose-dependently alleviated the obese phenotype, lipid accumulation and liver injury in mice subjected to 18 weeks HFD diet. Untargeted metabolomics and microbiomics analysis revealed that BBBP could alleviate the disturbance of lipid and amino acid metabolism and the imbalance of gut microbiota. Furthermore, BBBP oral gavage activated liver bile acid receptors, as indicated by elevated mRNA levels of Nr1h4, Nr1i2 and S1pr2. Surprisingly, we determined that BBBP, which mainly contained bile acids that possessed antimicrobial activity, could promote the growth of Lactobacillus. Correlation analysis showed a remarkable correlation between Lactobacillus and endogenous metabolites such as valine, serine, glutamine, et al. Among them, GABA which could be produced by Lactobacillus significantly reduced the lipid accumulation in L02 cells. Conclusions: The role of BBBP in regulating lipid metabolism might be achieved by activating bile acid receptors, or partially by promoting the levels of Lactobacillus and its metabolites such as GABA. Our study provided evidence that BBBP could be a novel therapeutic candidate for the treatment of NAFLD.

Sanhan Huashi formula (SHHS), a traditional Chinese medicine (TCM), has shown significant therapeutic effects on coronavirus disease 2019 (COVID-19) in clinical settings. However, its specific mechanism and components still require further clarification. In vitro experiments with Vero-E6 cells infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) demonstrated that SHHS effectively inhibited viral invasion and proliferation. Complementary in vivo experiments using K18-human angiotensin converting enzyme 2 (hACE2) mice exposed to virus-like particles (VLPs) further confirmed that SHHS impeded SARS-CoV-2 entry. Although SHHS did not demonstrate direct antiviral effects in K18-hACE2 mice challenged with SARS-CoV-2, it significantly alleviated pathological damage and decreased the expression of chemokines such as C–C motif ligand (CCL)-2, CCL-3, C–X–C motif ligand (CXCL)-1, CXCL-6, CXCL-9, CXCL-10, and CXCL-11 in the lungs, suggesting that SHHS exerts immunomodulatory and anti-inflammatory effects via the CCL-2–CXCL axis. Additional research using a lipopolysaccharide (LPS)-induced acute lung injury (ALI) and RAW264.7 cell model validated the ability of SHHS to reduce the levels of inflammatory biomarkers, including interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). Using advanced analytical techniques such as ultrahigh-performance liquid chromatography coupled with linear trap quadrupole Orbitrap mass spectrometry (UHPLC-LTQ-Orbitrap-MS) and surface plasmon resonance (SPR), nodakenin was identified as a potent antiviral component of SHHS that targets the 3C-like protease (3CLpro), a finding supported by the hydrogen–deuterium exchange mass spectrometry (HDX-MS) and molecular docking analyses. Furthermore, nodakenin demonstrated a significant antiviral effect, reducing the viral load by more than 66%. This investigation reveals that SHHS can combat COVID-19 by inhibiting viral invasion and promoting anti-inflammatory effects.

Bailing capsule (BLC), a drug that is clinically administered to modulate the autoimmune system, exhibits promising therapeutic potential in the treatment of thyroiditis. This study elucidates the chemical profile of BLC and its potential therapeutic mechanism in thyroiditis, leveraging network pharmacology and molecular docking techniques. Utilizing ultra‐high‐performance liquid chromatography coupled with linear trap‐Orbitrap mass spectrometry (UHPLC‐LTQ‐Orbitrap MS), 58 compounds were identified, the majority of which were nucleosides and amino acids. Utilizing the ultra‐high‐performance liquid chromatography coupled with triple quadrupole tandem mass spectrometry (UHPLC QqQ MS/MS) strategy, 16 representative active components from six batches of BLCs were simultaneously determined. Network pharmacology analysis further revealed that the active components included 5′‐adenylate, guanosine, adenosine, cordycepin, inosine, 5′‐guanylic acid, and l‐lysine. Targets with higher connectivity included AKT1, MAPK3, RAC1, and PIK3CA. The signaling pathways primarily focused on thyroid hormone regulation and the Ras, PI3K/AKT, and MAPK pathways, all of which were intricately linked to inflammatory immunity and hormonal regulation. Molecular docking analysis corroborated the findings from network pharmacology, revealing that adenosine, guanosine, and cordycepin exhibited strong affinity toward AKT1, MAPK3, PIK3CA, and RAC1. Overall, this study successfully elucidated the material basis and preliminary mechanism underlying BLC's intervention in thyroiditis, thus laying a solid basis for further exploration of its in‐depth mechanisms.

ETHNOPHARMACOLOGICAL RELEVANCE:Diabetic nephropathy (DN) was a major cause of end-stage renal failure and a common microvascular complication in patients with diabetes mellitus (DM). Acteoside (ACT) was the main ingredient extracted from the leaves of Rehmannia glutinosa, which had the functions of entering the lung, moisturizing the skin and relieving itching, nourishing yin and tonifying the kidney, cooling blood, and stopping bleeding. ACT had attracted worldwide interest because of its therapeutic effects on DM and its complications.AIM OF THE STUDY:To clarify the metabolic profiles and targets of ACT in db/db mice based on metabolomics and network pharmacology studies.MATERIALS AND METHODS:Db/db mice were used to observe the biochemical indices and histopathological changes in the kidney to evaluate the pharmacological effects of ACT on DN. Untargeted metabolomics studies were performed to investigate by UHPLC-LTQ-Orbitrap MS on urine, serum, and kidney samples. The key targets and pathways were analyzed by network pharmacology. For the pathways enriched by untargeted metabolomics, targeted metabolomics by UHPLC-QQQ-MS/MS was performed in kidney samples for validation. Sensitive biomarkers in kidney samples were evaluated. The effect of ACT on the improvement of DN from the perspective of metabolism of small molecules in vivo was described.RESULTS:ACT could delay the progression of DN and improve the degree of histopathological damage to the kidney. The pathways were focused on amino acid metabolism by untargeted metabolomics. Through network pharmacology analysis, the effect pathways were related to signal transduction, carbohydrate, lipid, amino acid metabolism and mainly affected the endocrine and immune systems. Amino acid metabolism was disturbed in the kidney of db/db mice, which could be callback by ACT, such as tryptophan, glutamine, cysteine, leucine, threonine, proline, phenylalanine, histidine, serine, arginine, asparagine by targeted metabolomics.CONCLUSIONS:In conclusion, this study provided strong support for ACT on DN treatment in clinics. Meanwhile, the Rehmannia glutinosa was used fully to raise the income level of farmers economically, while achieving the social benefit of empowering rural revitalization.

Objective: This study aims to elucidate and quantify the composition of Jiangtang Qingre formula (JQF), delineate the absorbed components in the bloodstream, predict the major biologically active components, and identify potential targets for the treatment of diabetes mellitus (DM). Materials and Methods: The chemical composition and metabolites of JQF were elucidated using ultra-high-performance liquid chromatography (UHPLC)-linear ion trap quadrupole-orbitrap high-resolution mass spectrometry (MS). The various components of JQF were concurrently determined using UHPLC-triple–quadrupole MS. Network pharmacological analysis was employed to explore the bioactive components and potential therapeutic targets in DM. Results: A total of 63 compounds were identified and provisionally characterized, with flavones, organic acids, and alkaloids emerging as the major chemical constituents. A robust analytical method that enables the simultaneous quantification of 24 representative components was successfully developed. The contents of 11 batches of samples were assessed. Ten prototype components were identified in rat plasma. The pathways associated with the efficacy of JQF in DM treatment were linked to signal transduction, endocrine and immune systems, lipid metabolism, and amino acid metabolism. Conclusion: This study systematically and comprehensively characterized the major chemical components and patterns in JQF, laying the groundwork for understanding its pharmacodynamic mechanisms and clinical applications.