A Glycyrrhizin Derivative with a More Potent Inhibitory Activity against High-Mobility Group Box 1 Efficiently Discovered by Chemical Synthesis Inspired by the Bioconversion Products of an Endophytic Fungus Isolated from Licorice.

Glycyrrhizin (GL) from licorice alleviates intracerebral hemorrhage (ICH) injuries by interacting with high-mobility group box (HMGB) 1, an inflammatory factor. We found that GL is bioconverted by endophyte coexisting with licorice and succeeded in isolating two derivatives. The aim of this study was to identify the compound with more potent HMGB1 inhibitory activity inspired by these GL derivatives. We took advantage of a ketone introduced by an endophyte at the C-3 position and attempted methyl esterification at the C-30 position because it was suggested that the water or lipid solubility of the molecule plays an important role. Among three derivatives synthesized, the product that is both ketonized and esterified showed more potent HMGB1 inhibitory activity than GL in macrophages and significantly improved adverse events occurred in ICH in vivo. These results suggest that modification of the hydrophilicity of GL, particularly at the C-3 and C-30 positions, enhances the HMGB1 inhibitory activity.

Fungal endophytes Fusarium solani SGGF14 and Alternaria tenuissima SGGF21 enhance the glycyrrhizin production by modulating its key biosynthetic genes in licorice (Glycyrrhiza glabra L.).

AIMS: To identify promising fungal endophytes that are able to produce glycyrrhizin and enhance it in licorice and the mechanisms involved. METHODS AND RESULTS: Fifteen fungal endophytes were isolated from Glycyrrhiza glabra L. rhizomes among which SGGF14 and SGGF21 isolates were found to produce glycyrrhizin by 4.29 and 2.58 µg g-1 dry weight in the first generation of their culture. These isolates were identified as Fusarium solani and Alternaria tenuissima, respectively, based on morphological characteristics and sequence analysis of internal transcribed spacer, TEF1, ATPase, and CAL regions. Subsequently, G. glabra plants were inoculated with these fungal isolates to examine their effect on glycyrrhizin production, plant growth parameters and the expression of key genes involved in glycyrrhizin pathway: SQS1, SQS2, bAS, CAS, LUS, CYP88D6, and CYP72A154. Endophytes were able to enhance glycyrrhizin content by 133%-171% in the plants. Natural control (NC) plants, harboring all natural endophytes, had better growth compared to SGGF14- and SGGF21-inoculated and endophyte-free (EF) plants. Expression of SQS1, SQS2, CYP88D6, and CYP72A154 was upregulated by inoculation with endophytes. LUS and CAS were downregulated after endophyte inoculation. Expression of bAS was higher in SGGF21-inoculated plants when compared with NC, EF, and SGGF14-inoculated plants. CONCLUSIONS: Two selected fungal endophytes of G. glabra can produce glycyrrhizin and enhance glycyrrhizin content in planta by modulating the expression of key genes in glycyrrhizin biosynthetic pathway.

Alleviation of Salt Stress and Changes in Glycyrrhizic Acid Accumulation by Dark Septate Endophytes in Glycyrrhiza glabra Grown under Salt Stress.

This study aimed to investigate the mechanisms by which dark septate endophytes (DSE) regulate salt tolerance and the accumulation of bioactive constituents in licorice. First, the salt stress tolerance and resynthesis with the plant effect of isolated DSE from wild licorice were tested. Second, the performance of licorice inoculated with DSE, which had the best salt-tolerant and growth-promoting effects, was examined under salt stress. All isolated DSE showed salt tolerance and promoted plant growth, withCurvularia lunata D43 being the most effective. Under salt stress, C. lunata D43 could promote growth, increase antioxidant enzyme activities, enhance glycyrrhizic acid accumulation, improve key enzyme activities in the glycyrrhizic acid synthesis pathway, and induce the expression of the key enzyme gene and salt tolerance gene of licorice. The structural equation model demonstrated that DSE alleviate the negative effects of salt stress through direct and indirect pathways. Variations in key enzyme activities, gene expression, and bioactive constituent concentration can be attributed to the effects of DSE. These results contribute to revealing the value of DSE for cultivating medicinal plants in saline soils.

Introducing the glycyrrhizic acid and glabridin rich genotypes from the cultivated Iranian licorice (Glycyrrhiza glabra L.) populations to exploit in production systems.

Currently, the stable, uniform, and highly efficient production of raw materials for pharmaceutical companies has received special attention. To meet these criteria and reduce harvesting pressure on the natural habitats of licorice (Glycyrrhiza glabra L.), cultivation of this valuable plant is inevitable. In the present study, to introduce the glycyrrhizic acid (GA)- and glabridin-rich genotypes from cultivated Iranian licorice, forty genotypes from eight high-potential wild populations were cultivated and evaluated under the same environmental conditions. The GA content varied from 5.00 ± 0.04 mg/g DW (TF2 genotype) to 23.13 ± 0.02 mg/g DW (I5 genotype). The highest and lowest glabridin content were found in the K2 (0.72 ± 0.021 mg/g DW) and M5 (0.02 ± 0.002 mg/g DW) genotypes, respectively. The rutin content in the leaves of the studied genotypes varied from 1.27 ± 0.02 mg/g DW in E4 to 3.24 ± 0.02 mg/g DW in BO5 genotypes. The genotypes from the Ilam population were characterized by higher vegetative growth and yield traits in the aerial parts and roots. The average root dry yield was 2.44 tons per hectare (t/ha) among the studied genotypes and a genotype from Ilam (I5) yielded the maximum value (3.08 ± 0.034 t/ha). The highest coefficient of variation among the genotypes was observed for leaf width (CV = 34.9%). The GA and glabridin-rich genotypes introduced in this study can be used in the future breeding programs to release new bred licorice cultivars.

Network pharmacology and in vitro experimental verification unveil glycyrrhizin from glycyrrhiza glabra alleviates acute pancreatitis via modulation of MAPK and STAT3 signaling pathways.

Acute pancreatitis (AP) is a severe gastrointestinal inflammatory disease with increasing mortality and morbidity. Glycyrrhiza glabra, commonly known as Liquorice, is a widely used plant containing bioactive compounds like Glycyrrhizin, which possesses diverse medicinal properties such as anti-inflammatory, antioxidant, antiviral, and anticancer activities. The objective of this study is to investigate the active components, relevant targets, and underlying mechanisms of the traditional Chinese medicine Glycyrrhiza glabra in the treatment of AP. Utilizing various computational biology methods, we explored the potential targets and molecular mechanisms through Glycyrrhizin supplementation. Computational results indicated that Glycyrrhizin shows promising pharmacological potential, particularly with mitogen-activated protein kinase 3 (MAPK3) protein (degree: 70), forming stable complexes with Glycyrrhizin through ionic and hydrogen bonding interactions, with a binding free energy (ΔGbind) of -33.01 ± 0.08 kcal/mol. Through in vitro experiments, we validated that Glycyrrhizin improves primary pancreatic acinar cell injury by inhibiting the MAPK/STAT3/AKT signaling pathway. Overall, MAPK3 emerges as a reliable target for Glycyrrhizin's therapeutic effects in AP treatment. This study provides novel insights into the active components and potential targets and molecular mechanisms of natural products.

Glycyrrhizic acid alleviated MI/R-induced injuries by inhibiting Hippo/YAP signaling pathways.

BACKGROUND: Previous research has demonstrated that glycyrrhizic acid (GA) exhibits antioxidant, anti-inflammatory, and antiapoptotic characteristics. Using myocardial ischemia/reperfusion injury as a case study, this study aims to clarify the functional significance of GA and to elucidate the mechanisms involved. MATERIALS AND METHODS: In this study, an MI/R injury model was established both in vivo and in vitro to investigate the impact of GA on MI/R injury. The viability of H9c2 cells was evaluated using the Cell Counting Kit-8. Myocardial damage was assessed through the measurement of creatine kinase myocardial band (CK-MB) levels and lactate dehydrogenase (LDH), HE staining, and MASSON staining. Inflammatory cytokine levels (IL-6, IL-1ß, IL-10, and TNF-α) were measured to determine the presence of inflammation. Cellular oxidative stress was evaluated by measuring ROS and MMP levels, while cardiac function was assessed using cardiac color Doppler ultrasound. Immunofluorescence staining to determine the nuclear translocation of YAP, TUNEL to determine apoptosis, and western blotting to determine gene expression. RESULTS: GA treatment effectively alleviated myocardial injury induced by MI/R, as evidenced by reduced levels of inflammatory cytokines (IL-1ß, IL-6, IL-10, and TNF-α) and cardiac biomarkers (CK-MB, LDH) in MI/R rats. Moreover, There was a significant increase in cell viability in vitro after GA treatment and inhibited reactive oxygen species (ROS) during oxidative stress, while also increasing mitochondrial membrane potential (MMP) in vitro. The Western blot findings indicate that GA treatment effectively suppressed apoptosis in both in vivo and in vitro settings. Additionally, GA demonstrated inhibitory effects on the activation of the Hippo/YAP signaling pathway triggered by MI/R and facilitated YAP nuclear translocation both in vitro and in vivo. It has been found, however, in vitro, that silencing the YAP gene negates GA's protective effect against hypoxia/reoxygenation-induced myocardial injury. CONCLUSION: This study suggests that GA regulates YAP nuclear translocation by inhibiting the Hippo/YAP signaling pathway, which protects ists against MI/R injury. This finding may present a novel therapeutic approach for the treatment of MI/R.

Glycyrrhizin regulates antioxidation through Nrf2 signaling pathway in rat temporomandibular joint osteoarthritis.

BACKGROUND: Regulation of redox homeostasis could reduce osteoarthritis severity and limit disease progression, while glycyrrhizin (GL) shows great antioxidant and anti-inflammatory capacity. OBJECTIVE: The aim of this study was to investigate the role of GL on oxidative stress and the potential regulatory mechanism in rat temporomandibular joint (TMJ) chondrocytes under oxidative stress, and investigate the effect of GL in the rat temporomandibular joint osteoarthritis (TMJOA) model. METHODS: Rat TMJ chondrocytes were cultured in oxidative stress with different doses of GL. The effect of glycyrrhizin on the nuclear factor-erythroid 2-related factor 2 (Nrf2) in oxidative stress was evaluated by western blot and immunofluorescence staining. A rat model of TMJOA was treated with GL. Micro-computed tomography, histological and immunohistochemical analysis were used to assess the pathological change of TMJOA. RESULTS: The expression of superoxide dismutase 1 (SOD1), heme oxygenase-1 (HO-1), and peroxiredoxin 6 (PRDX6) were decreased, and intracellular Nrf2 signaling pathway was activated in chondrocytes in oxidative stress. GL upregulates the expression of antioxidants, especially PRDX6, as well as increases Nrf2 expression and nuclear translocation in rat condylar chondrocytes. Administration of GL attenuates condylar bone destruction, cartilage degeneration, and synovitis in rats TMJOA. Meanwhile, GL alleviated oxidative stress and enhanced the antioxidant capacity of TMJOA cartilage. CONCLUSION: This study suggested that GL alleviates rat TMJOA by regulating oxidative stress in condylar cartilage.

Glycyrrhizin Production in Licorice Hairy Roots Based on Metabolic Redirection of Triterpenoid Biosynthetic Pathway by Genome Editing.

Glycyrrhizin, a type of the triterpenoid saponin, is a major active ingredient contained in the roots of the medicinal plant licorice (Glycyrrhiza uralensis, G. glabra and G. inflata), and is used worldwide in diverse applications, such as herbal medicines and sweeteners. The growing demand for licorice threatens wild resources and therefore a sustainable method of supplying glycyrrhizin is required. With the goal of establishing an alternative glycyrrhizin supply method not dependent on wild plants, we attempted to produce glycyrrhizin using hairy root culture. We tried to promote glycyrrhizin production by blocking competing pathways using CRISPR/Cas9-based gene editing. CYP93E3 CYP72A566 double-knockout (KO) and CYP93E3 CYP72A566 CYP716A179 LUS1 quadruple-KO variants were generated, and a substantial amount of glycyrrhizin accumulation was confirmed in both types of hairy root. Furthermore, we evaluated the potential for promoting further glycyrrhizin production by simultaneous CYP93E3 CYP72A566 double-KO and CYP88D6-overexpression. This strategy resulted in a 3-fold increase (∼1.4 mg/g) in glycyrrhizin accumulation in double-KO/CYP88D6-overexpression hairy roots, on average, compared with that of double-KO hairy roots. These findings demonstrate that the combination of blocking competing pathways and overexpression of the biosynthetic gene is important for enhancing glycyrrhizin production in G. uralensis hairy roots. Our findings provide the foundation for sustainable glycyrrhizin production using hairy root culture. Given the widespread use of genome editing technology in hairy roots, this combined with gene knockout and overexpression could be widely applied to the production of valuable substances contained in various plant roots.

Disruption of a licorice cellulose synthase-derived glycosyltransferase gene demonstrates its in planta role in soyasaponin biosynthesis.

KEY MESSAGE: CRISPR-Cas9-mediated disruption of a licorice cellulose synthase-derived glycosyltransferase gene, GuCSyGT, demonstrated the in planta role of GuCSyGT as the enzyme catalyzing 3-O-glucuronosylation of triterpenoid aglycones in soyasaponin biosynthesis. Triterpenoid glycosides (saponins) are a large, structurally diverse group of specialized metabolites in plants, including the sweet saponin glycyrrhizin produced by licorice (Glycyrrhiza uralensis) and soyasaponins that occur widely in legumes, with various bioactivities. The triterpenoid saponin biosynthetic pathway involves the glycosylation of triterpenoid sapogenins (the non-sugar part of triterpenoid saponins) by glycosyltransferases (GTs), leading to diverse saponin structures. Previously, we identified a cellulose synthase-derived GT (CSyGT), as a newly discovered class of triterpenoid GT from G. uralensis. GuCSyGT expressed in yeast, which could transfer the sugar glucuronic acid to the C3 position of glycyrrhetinic acid and soyasapogenol B, which are the sapogenins of glycyrrhizin and soyasaponin I, respectively. This suggested that GuCSyGT is involved in the biosynthesis of glycyrrhizin and soyasaponin I. However, the in planta role of GuCSyGT in saponin biosynthesis remains unclear. In this study, we generated GuCSyGT-disrupted licorice hairy roots using CRISPR-Cas9-mediated genome editing and analyzed the saponin content. This revealed that soyasaponin I was completely absent in GuCSyGT-disrupted lines, demonstrating the in planta role of GuCSyGT in saponin biosynthesis.

Multi-Omics Elucidates Difference in Accumulation of Bioactive Constituents in Licorice (Glycyrrhiza uralensis) under Drought Stress.

Licorice is a frequently applied herb with potential edible and medicinal value based on various flavonoids and triterpenes. However, studies on detailed flavonoid and triterpene metabolism and the molecular basis of their biosynthesis in licorice are very limited, especially under drought conditions. In the present study, we carried out transcriptome, proteome, and metabolome experiments. To ultimately combine three omics for analysis, we performed a bioinformatics comparison, integrating transcriptome data and proteome data through a Cloud platform, along with a simplified biosynthesis of primary flavonoids and triterpenoids in the KEGG pathway based on metabolomic results. The biosynthesis pathways of triterpenes and flavonoids are enriched at both gene and protein levels. Key flavonoid-related genes (PAL, 4CL, CHS, CHI, CYP93C, HIDH, HI4OMT, and CYP81E1_7) and representative proteins (HIDH, CYP81E1_7, CYP93C, and VR) were obtained, which all showed high levels after drought treatment. Notably, one R2R3-MYB transcription factor (Glyur000237s00014382.1), a critical regulator of flavonoid biosynthesis, achieved a significant upregulated expression as well. In the biosynthesis of glycyrrhizin, both gene and protein levels of bAS and CYP88D6 have been found with upregulated expression under drought conditions. Most of the differentially expressed genes (DEGs) and proteins (DEPs) showed similar expression patterns and positively related to metabolic profiles of flavonoid and saponin. We believe that suitable drought stress may contribute to the accumulation of bioactive constituents in licorice, and our research provides an insight into the genetic study and quality breeding in this plant.

Site-specific crosslinking and assembly of tetrameric ß-glucuronidase improve glycyrrhizin hydrolysis.

In this study, eight nonconserved residues with exposed surfaces and flexible conformations of the homotetrameric PGUS (ß-glucuronidase from Aspergillus oryzae Li-3) were identified. Single-point mutation into cysteine enabled the thiol-maleimide reaction and site-specific protein assembly using a two-arm polyethylene glycol (PEG)-maleimide crosslinker (Mal2 ). The Mal2 (1k) (with 1 kDa PEG spacer)-crosslinked PGUS assemblies showed low crosslinking efficiency and unimproved thermostability except for G194C-Mal2 (1k). To improve the crosslinking efficiency, a lengthened crosslinker Mal2 (2k) (with 2 kDa PEG spacer) was used to produce PGUS assembly and a highly improved thermostability was achieved with a half-life of 47.2-169.2 min at 70°C, which is 1.04-3.74 times that of wild type PGUS. It is found that the thermostability of PGUS assembly was closely associated with the formation of inter-tetramer assembly and intratetramer crosslinking, rather than the PEGylation of the enzyme. Therefore, the four-arm PEG-maleimide crosslinker Mal4 (2k) (with 2 kDa PEG spacer) was employed to simultaneously increase the inter-tetramer assembly and intratetramer crosslinking, and the resulting PGUS assemblies showed further improved thermostabilities compared with Mal2 (2k)-crosslinked assemblies. Finally, the application of PGUS assemblies with significantly improved thermostability to the bioconversion of GL proved that the PGUS assembly is a strong catalyst for glycyrrhizin (GL) hydrolysis in industrial applications.

Glycyrrhizic Acid Protects Glomerular Podocytes Induced by High Glucose by Modulating SNARK/AMPK Signaling Pathway.

OBJECTIVE: Diabetic nephropathy is one of the most important microvascular complications of diabetes, which mainly refers to glomerular capillary sclerosis. Podocytes are an important part of glomerular capillaries. Previous clinical and basic studies have shown that fibrosis is the main factor of diabetic nephropathy. This study aimed to assess the protective mechanism of glycyrrhizic acid (GA) on glomerular podocytes induced by high glucose as we hypothesized that GA may have antifibrotic and anti-inflammatory effects on podocytes through regulation of the adenosine 5'-monophosphate-activated protein kinase (AMPK)/sucrose nonfermenting AMPK-related kinase (SNARK) signaling pathway. METHODS: SNARK siRNA was used to transfect podocytes. Real-time quantitative polymerase chain reaction and immunofluorescence staining assays were used for molecular and pathological analysis. The expression levels of key pathway proteins (including TGF-ß1, α-SMA, SITR1, AMPKα, LKB1, PGC-1α, NF-κB, IL-6, and TNF-α) were verified by Western blotting. The expression of inflammatory factors in podocytes was detected by ELISA. RESULTS: We demonstrated that GA decreased the expression of podocyte fibrosis signaling pathway-related factors by upregulating the AMPK pathway and its related factors. However, after transfection of podocytes with SNARK siRNA, there was an increased expression of fibrosis-related factors and inflammation-related factors. CONCLUSION: GA can protect podocytes and alleviate fibrosis and inflammation induced by high glucose, which is related to the AMPK signaling pathway. Meanwhile, knockdown of SNARK protein can inhibit the AMPK signaling pathway, aggravate fibrosis, and increase inflammation.

Exploring the binding effect and mechanism of glycyrrhizin to ovomucin by combining spectroscopic analysis and molecular docking.

Ovomucin (OVM) is an ideal natural macromolecular glycoprotein extracted from eggs with good adhesion. Based on the defect that glycyrrhizin (GL) has good antiviral activity but fast metabolism, this study aimed to explore the binding effect and mechanism of GL to OVM, using multi-spectroscopic techniques, isothermal titration calorimetry (ITC), and molecular docking. The adhesion ability of OVM to the hydrophilic interface and GL was first demonstrated by dual polarization interferometry (DPI) analysis and binding capacity assay, and the OVM-GL complex exhibited a similar affinity for the spike protein of COVID-19. The spectroscopic results show that GL can quench the inherent fluorescence and change the glycosidic bond and secondary structure of OVM. The ITC measurements suggested that the binding was exothermic, the hydrogen bond was the dominant binding force for forming OVM-GL. Finally, molecular docking results indicated that GL has hydrogen bond interaction with several amino acid residues located in α-OVM and ß-OVM while embedding into the hydrophobic pocket of OVM via hydrophobic interactions. In conclusion, OVM can adhere to the hydrophilic interface and bind to GL through hydrogen bonding and hydrophobic interactions to form a stable complex, that is expected to be helpful in virus prophylaxis.

[Adverse effects of licorice consumed as food: An update]. / Toxicités de l'exposition alimentaire à la réglisse : mise au point.

The word "licorice" refers to the plant, its root, and its aromatic extract. From a commercial point of view, Glycyrrhiza glabra is the most important species with a wide range of uses (herbal medicine, tobacco industry, cosmetics, food and pharmaceutical). Glycyrrhizin is one of the main constituents of licorice. Glycyrrhizin is hydrolyzed in the intestinal lumen by bacterial ß-glucuronidases to 3ß-monoglucuronyl-18ß-glycyrrhetinic acid (3MGA) and 18ß-glycyrrhetinic acid (GA), which are metabolized in the liver. Plasma clearance is slow due to enterohepatic cycling. 3MGA and GA can bind to mineralocorticoid receptors with very low affinity, and 3MGA induces apparent mineralocorticoid excess syndrome through dose-dependent inhibition of 11ß-hydroxysteroid dehydrogenase type 2 in renal tissue. The cases of apparent mineralocorticoid excess syndrome reported in the literature are numerous and sometimes severe, even fatal, most often in cases of chronic high dose consumption. Glycyrrhizin poisonings are characterized by hypertension, fluid retention, and hypokalemia with metabolic alkalosis and increased kaliuresis. Toxicity depends on the dose, the type of product consumed, the mode of consumption (acute or chronic) and a very large inter-individual variability. The diagnosis of glycyrrhizin-induced apparent mineralocorticoid excess syndrome is based on the history, clinical examination, and biochemical analysis. Management is primarily based on symptomatic care and stopping licorice consumption.

Glycyrrhizin, an inhibitor of HMGB1 induces autolysosomal degradation function and inhibits Helicobacter pylori infection.

BACKGROUND: Helicobacter pylori is a key agent for causing gastric complications linked with gastric disorders. In response to infection, host cells stimulate autophagy to maintain cellular homeostasis. However, H. pylori have evolved the ability to usurp the host's autophagic machinery. High mobility group box1 (HMGB1), an alarmin molecule is a regulator of autophagy and its expression is augmented during infection and gastric cancer. Therefore, this study aims to explore the role of glycyrrhizin (a known inhibitor of HMGB1) in autophagy during H. pylori infection. MAIN METHODS: Human gastric cancer (AGS) cells were infected with the H. pylori SS1 strain and further treatment was done with glycyrrhizin. Western blot was used to examine the expression of autophagy proteins. Autophagy and lysosomal activity were monitored by fluorescence assays. A knockdown of HMGB1 was performed to verify the effect of glycyrrhizin. H. pylori infection in in vivo mice model was established and the effect of glycyrrhizin treatment was studied. RESULTS: The autophagy-lysosomal pathway was impaired due to an increase in lysosomal membrane permeabilization during H. pylori infection in AGS cells. Subsequently, glycyrrhizin treatment restored the lysosomal membrane integrity. The recovered lysosomal function enhanced autolysosome formation and concomitantly attenuated the intracellular H. pylori growth by eliminating the pathogenic niche. Additionally, glycyrrhizin treatment inhibited inflammation and improved gastric tissue damage in mice. CONCLUSION: This study showed that inhibiting HMGB1 restored lysosomal activity to ameliorate H. pylori infection. It also demonstrated the potential of glycyrrhizin as an antibacterial agent to address the problem of antimicrobial resistance.

Glycyrrhizic acid improves tacrolimus-induced renal injury by regulating autophagy.

Tacrolimus (TAC)-induced renal injury is detrimental to long-term kidney function, but a treatment medication is not available. Glycyrrhizic acid (GA) is an active ingredient in licorice widely used to treat kidney disease. Thus, this study explored the mechanisms of renoprotection by GA on TAC-induced renal injury. C57BL/6 mice were subjected daily to TAC or a combination of TAC and GA for 4 weeks, and then renal function, histopathology, and autophagy were assessed to examine the effect of GA on a renal injury. Next, Human kidney proximal tubular epithelial (HK-2) cells were pretreated with GA for 2 h and then treated with TAC for 24 h. The effect of GA on TAC-induced HK-2 cell injury was assessed by measuring cell viability, apoptosis, autophagy, and lysosomes. Mice exposed to TAC and treated with GA had significantly greater improvements in renal function and tubulointerstitial fibrosis in comparison to mice not treated with GA. In addition, fibrosis-related protein expression, including α-smooth muscle actin and fibronectin, decreased after GA treatment. GA treatment also relieved autophagic clearance in TAC-induced renal injury. Several in vitro studies found that TAC inhibited cell viability, autophagy, lysosomal acidification, and promoted apoptosis. However, these results were less pronounced with GA pretreatment. In addition, bafilomycin A1 (which inhibits lysosomal function) reduced the protective effect of GA, indicating that lysosomal function plays an important role in this effect. Our data suggest that GA improves lysosomal function and regulates autophagy to protect against TAC-induced renal injury.

Incorporation of glycyrrhizic acid and polyene phosphatidylcholine in lipid nanoparticles ameliorates acute liver injury via delivering p65 siRNA.

Liver injury caused by hepatitis is the pathological basis of varied hepatic diseases with high morbidity and mortality. Although siRNA appears promising in therapeutics of hepatitis, efficient and safe delivery remains a challenge. In this study, we developed a new strategy of incorporating glycyrrhizic acid (GA) and polyene phosphatidylcholine (PPC) into lipid nanoparticles (GA/PPC-modified LNPs), which was capable of promoting cellular uptake, enhancing gene-silencing, reducing cytotoxicity and improving siRNA stability. GA/PPC-modified LNP and siRNA lipoplex targeting NF-κB, a key mediator of inflammation, mitigates acute liver injury, as assessed by liver histology, hematological and pro-inflammatory cytokine analysis. Furthermore, GA/PPC-modified LNPs reveal efficiently intracellular delivery of antisense oligonucleotides (ASOs) and mRNA inhibiting viral infection. In conclusion, GA/PPC-modified LNPs could be used as a promising delivery system for nucleic acid-based therapy.

Antioxidant monoammonium glycyrrhizinate alleviates damage from oxidative stress in perinatal cows.

This study was conducted to evaluate the antioxidant capability of dietary supplementation with monoammonium glycyrrhizinate (MAG) in perinatal cows. Glycyrrhizic acid has been shown to have strong antioxidant activity and we hypothesised that the aglycone of glycyrrhizin and MAG, could reduce damage from oxidative stress in perinatal cows by enhancing antioxidant capacity. Blood and milk samples were collected from three groups of healthy perinatal cows that were similar in body weight, parity, milk yield in the last milk cycle, etc., receiving dietary MAG supplementation ([Day 0 = parturition]: 0 g/day, [n = 13)] 3 g/day [n = 13] or 6 g/day [n = 11]) from -28 to 56 day (0 day = parturition). Compared with 0 g/day controls (CON), milk fat was significantly decreased in cows fed with MAG, and 3 g/day had the greatest effect. A diet containing 3 g/day MAG decreased the serum alanine aminotransferase (ALT) level compared with CON at -7 day post-partum. ALT was also lower at 5 day post-partum in cows fed with 3 g/day MAG compared to 6 g/day. The administration of 3 g/day and 6 g/day MAG decreased serum aspartate transaminase (AST) at 3 day post-partum. Supplementation of MAG in cows increased total antioxidant capacity (T-AOC) in serum, and cows given 3 g MAG per day had higher T-AOC than controls on post-partum 7 day. At the end of the experiment, we isolated and cultured primary hepatocytes to determine the effect of MAG on oxidative stress caused by incubation with the sodium oleate (SO). SO increased lipid synthesis, but pre-treatment with MAG prevented the fatty buildup. SO treatment increased AST and ALT levels and malondialdehyde concentration, but decreased T-AOC and superoxide dismutase (SOD). Incubation with MAG increased antioxidant capacity and inhibited oxidant damage in bovine hepatocytes. SO stimulated expression of the antioxidant genes, NAD(P)H quinone dehydrogenase 1 (NQO1) and SOD1, in the nuclear factor erythroid 2-related factor 2 (NRF2) pathway, and catalase 1 (CAT1); this increase was accentuated by MAG pre-treatment. The results suggest that MAG can alleviate the damage caused by oxidative stress in perinatal cows by enhancing antioxidant activity.

LPS-induced macrophage exosomes promote the activation of hepatic stellate cells and the intervention study of total astragalus saponins combined with glycyrrhizic acid.

Huangqi decoction, also known as Huangqi Liuyi decoction, was first recorded in the prescriptions of the Bureau of Taiping People's Welfare Pharmacy. It comprises astragalus and licorice, which is a commonly used prescription in traditional Chinese medicine for the clinical treatment of chronic liver disease, especially liver cirrhosis. Total astragalus saponins (AST) is the main component of astragalus, and glycyrrhizic acid (GA) is the main component of licorice. In this study, normal macrophage exosomes were extracted, and the exosomes incubated with lipopolysaccharides (LPS) and those incubated with LPS + AST + GA were co-cultured with JS1 cells (hepatic stellate cell line). The survival rate and the activation of key signaling pathways of JS1 cells in each group were detected and compared. We found that the co-culture of LPS-induced macrophage exosomes with JS1 cells could significantly increase the expression levels of Collagen-1 (Col-1) and Alpha smooth muscle actin (α-SMA)in JS1 cells. However, a significant reversal effect was observed after pretreatment with AST combined with GA. Further evaluation found that the expression levels of phospho (p)-Smad2 and p-Smad3 in the JS1 cells were significantly increased after macrophages were induced with LPS, whereas pretreatment with AST + GA could significantly decrease the expression levels of p-Smad2 and p-Smad3. Preliminary results of this study indicated that LPS-induced macrophage exosomes can promote the activation of hepatic stellate cells, and the pretreatment of AST combined with GA can exert a significant intervention effect. In this study, the new mechanism of anti-hepatic fibrosis effect of traditional Chinese medicine components of Huangqi Decoction was analyzed from the perspective of exosomes.

Modulation of gut microbiota by glycyrrhizic acid may contribute to its anti-NAFLD effect in rats fed a high-fat diet.

AIMS: Glycyrrhizic acid is a natural anti-non-alcoholic fatty liver disease (NAFLD) compound isolated from licorice, while its action mechanism deserves to be fully elucidated. MATERIALS AND METHODS: Enlightened by the widely discovered associations between the NAFLD and gut microbiota, this study aimed to explore whether glycyrrhizic acid, licorice flavonoids, and licorice extract can regulate the gut microbiota of rats fed a high-fat diet. KEY FINDINGS: It was found that glycyrrhizic acid, licorice flavonoids, and licorice extract could significantly reduce the level of triglycerides in the liver of NAFLD model rats, and the effect of glycyrrhizic acid was stronger than licorice flavonoids and licorice extract. Moreover, they caused significant changes in the structural composition of the gut microbiota. Correlation analysis showed that the regulation of hepatic total cholesterol and triglyceride levels by glycyrrhizic acid treatment was closely related to the decrease of the relative abundances of Lachnospiraceae, Coriobacteriaceae, Blautia, and Collinsella and the increase of the relative abundances of Romboutsia and Turicibacter on the gut microbiota. Meanwhile, the functional predictive analysis of the gut microbiota indicated that the function of carbohydrate transport and metabolism was significantly decreased by drugs treatment, which might contribute to the decrease of fat accumulation in the liver of rats. SIGNIFICANCE: In conclusion, this study revealed the ameliorating effects of glycyrrhizic acid, licorice flavonoids, and licorice extract on NAFLD, and suggested that the effect of glycyrrhizic acid on NAFLD may be related to the improvement of the dysbiosis of gut microbiota.