Ursodeoxycholic acid and 18ß-glycyrrhetinic acid alleviate ethinylestradiol-induced cholestasis via downregulating RORγt and CXCR3 signaling pathway in iNKT cells.

Estrogen-induced intrahepatic cholestasis (IHC) is a mild but potentially serious risk and urges for new therapeutic targets and effective treatment. Our previous study demonstrated that RORγt and CXCR3 signaling pathway of invariant natural killer T (iNKT) 17 cells play pathogenic roles in 17α-ethinylestradiol (EE)-induced IHC. Ursodeoxycholic acid (UDCA) and 18ß-glycyrrhetinic acid (GA) present a protective effect on IHC partially due to their immunomodulatory properties. Hence in present study, we aim to investigate the effectiveness of UDCA and 18ß-GA in vitro and verify the accessibility of the above targets. Biochemical index measurement indicated that UDCA and 18ß-GA presented efficacy to alleviate EE-induced cholestatic cytotoxicity. Both UDCA and 18ß-GA exhibited suppression on the CXCL9/10-CXCR3 axis, and significantly restrained the expression of RORγt in vitro. In conclusion, our observations provide new therapeutic targets of UDCA and 18ß-GA, and 18ß-GA as an alternative treatment for EE-induced cholestasis.

Glycyrrhetinic acid attenuates endoplasmic reticulum stress-induced hepatocyte apoptosis via CHOP/DR5/Caspase 8 pathway in cholestasis.

Bile acid (BA)-induced apoptosis is a common pathologic feature of cholestatic liver injury. Glycyrrhetinic acid (GA) is the hepatoprotective constituent of licorice. In the present study, the anti-apoptotic potential of GA was investigated in wild type and macrophage-depleted C57BL/6 mice challenged with alpha-naphthyl isothiocyanate (ANIT), and hepatocytes stimulated with Taurocholic acid (TCA) or Tumor necrosis factor-alpha (TNF-α). Apoptosis was determined by TUNEL positive cells and expression of executioner caspases. Firstly, we found that GA markedly alleviated liver injury, accompanied with reduced positive TUNEL-staining cells, and expression of caspases 3, 8 and 9 in mice modeled with ANIT. Secondly, GA mitigated apoptosis in macrophage-depleted mice with exacerbated liver injury and augmented cell apoptosis. In vitro study, pre-treatment with GA reduced the expression of activated caspases 3 and 8 in hepatocytes stimulated with TCA, but not TNF-α. The ability of GA to ameliorate apoptosis was abolished in the presence of Tauroursodeoxycholic Acid (TUDCA), a chemical chaperon against Endoplasmic reticulum stress (ER stress). Furthermore, GA attenuated the over-expression of Glucose regulated protein 78 (GRP78), and blocked all three branches of Unfolded protein reaction (UPR) in cholestatic livers of mice induced by ANIT. GA also downregulated C/EBP homologous protein (CHOP) expression, accompanied with reduced expression of Death receptor 5 (DR5) and activation of caspase 8 in both ANIT-modeled mice and TCA-stimulated hepatocytes. The results indicate that GA inhibits ER stress-induced hepatocyte apoptosis in cholestasis, which correlates with blocking CHOP/DR5/Caspase 8 pathway.

Exploring the molecular mechanism of glycyrrhetinic acid in the treatment of gastric cancer based on network pharmacology and experimental validation.

There is a wide range of pharmacological effects for glycyrrhetinic acid (GRA). Previous studies have shown that GRA could inhibit the proliferation of tumor cells, showing a promising value in the treatment of gastric cancer (GC). Nonetheless, the precise mechanism of the effect of GRA on GC remains unclear. We explored cellular and molecular mechanisms of GRA based on network pharmacology and in vitro experimental validation. In this study, we predicted 156 potential therapeutic targets for GC with GRA from public databases. We then screened the hub targets using protein-protein interaction network (PPI) and conducted clinical correlation analysis. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment showed that GRA made anti-GC effects through multiple targets and pathways, particularly the MAPK signaling pathway. Next, molecular docking results revealed a potential interaction between GRA and MAPK3. In addition, qRT-PCR experiments revealed that 18ß-GRA was able to suppress mRNA expression of KRAS, ERK1 and ERK2 in AGS cells. Western blotting results also revealed that 18ß-GRA was able to suppress the expression of KRAS and p-ERK1/2 proteins in AGS cells. Additionally, immunofluorescence assays revealed that 18ß-GRA inhibited p-ERK1/2 nuclear translocation in AGS cells. These results systematically reveal that 18ß-GRA may have anti-tumor effects on GC by modulating the MAPK signaling pathway.

18ß-glycyrrhetinic Acid Modulated Autophagy is Cytotoxic to Breast Cancer Cells.

The development of endocrine therapy resistance in the luminal A subtype of breast cancer is related to the appearance of protective autophagy. The bioactive component from the root of licorice, 18ß-glycyrrhetinic acid (18ß-GA), has many antitumor properties. Whether 18ß-GA can modulate autophagy to inhibit proliferation of the luminal A subtype is still unclear. The proportion of apoptosis caused by 18ß-GA in MCF-7 and T-47D cells was determined using flow cytometry. The autophagy marker, LC3-II conversion, was investigated using Western blotting, and a PremoTM Tandem Autophagy Sensor Kit. We found that the concentration (150-µM) of 18ß-GA caused caspase-dependent apoptosis and LC3-II accumulation or blocked autophagic flux. Moreover, 18ß-GA-mediated apoptosis was improved using rapamycin but reversed by 3-methyladenine (3-MA) addition. The phosphorylation level of Jun-amino-terminal kinase (JNK) was increased significantly in the 18ß-GA treatment and combined incubation using rapamycin. A JNK inhibitor (SP600125) significantly inhibited 18ß-GA-mediated apoptosis, LC3-II accumulation and rescued the numbers of MCF-7 and T-47D colony formation. Especially, 18ß-GA can inhibit xenograft tumor growth in BALB/c nude mice. These data indicate the combination of 18ß-GA with rapamycin or 3-MA can sensitize or decrease MCF-7 and T-47D cells to 18ß-GA-induced apoptosis, respectively. 18ß-GA modulated autophagy is cytotoxic to luminal A subtype breast cancer cells through apoptosis promotion and JNK activation.

Construction of redox-sensitive liposomes modified by glycyrrhetinic acid and evaluation of anti-hepatocellular carcinoma activity.

The aim of this study was to construct a bifunctional liposome with hepatic-targeting capacity by modifying with a targeting ligand and an intracellular tumor reduction response functional group to deliver drugs precisely to focal liver tissues and release them in large quantities in hepatocellular carcinoma cells. This could improve drug efficacy and reduce toxic side effects at the same time. First, the bifunctional ligand for liposome was successfully obtained by chemically synthesizing it from the hepatic-targeting glycyrrhetinic acid (GA) molecule, cystamine, and the membrane component cholesterol. Then the ligand was used to modify the liposomes. The particle size, PDI and zeta potential of the liposomes were determined with a nanoparticle sizer, and the morphology was observed by transmission electron microscopy. The encapsulation efficiency and drug release behavior were also determined. Further, the stability in vitro of the liposomes and the changes in the simulated reducing environment were determined. Finally, the antitumor activity in vitro and cellular uptake efficiency of the drug-loaded liposomes were investigated by performing cellular assays. The results showed that the prepared liposomes had a uniform particle size of 143.6 ± 2.86 nm with good stability and an encapsulation rate of 84.3 ± 2.1 %. Moreover, the particle size of the liposomes significantly increased and the structure was destroyed in a DTT reducing environment. Cellular experiments showed that the modified liposoes had better cytotoxic effects on hepatocarcinoma cells than both normal liposomes and free drugs. This study has great potential for tumor therapy and provides novel ideas for the clinical use of oncology drugs in dosage forms.

Synthesis of glycyrrhetinic acid-modified liposomes to deliver Murrayafoline A for treatment of hepatocellular carcinoma.

Hepatocellular carcinoma is a common type of cancer associated with a high mortality rate. Among several bioactive compounds, Murrayafoline A (MuA) has been proved as a bio substance that exhibits great potentials in treating liver cancer. In order to overcome the high cytotoxicity and low solubility of MuA, a delivery system based on nanocarriers is necessary to deliver MuA towards the desired target. In the present study, 18ß-glycyrrhetinic acid (GA), which is known as a ligand for liver targeting, was used to construct the cholesterol-poly (ethylene glycol)-glycyrrhetinic acid (GA-PEG-Chol) conjugate and liposome for MuA administration. The compound was then examined for therapeutic efficacy and safety in HUVEC and HepG2 cells in 2D and 3D cell cultures. Results have shown that MuA-loaded liposomes had IC50 value of 2 µM in HepG2 and had the cytosolic absorption of 8.83 ± 0.97 ng/105 cells, while The IC50 value of MuA-loaded liposomes in HUVEC cell lines was 15 µM and the the cytosolic absorption was recorded as 3.62 ± 0.61 cells. The drug test on the 3D cancer sphere platform of the HepG2 cancer sphere showed that MuA-loaded GA liposomes had the highest efficacy at a concentration of 100 µg/mL. In short, these results suggest that MuA-loaded GA liposomes have the potential for maintenance drug delivery and liver targeting.

Glycyrrhetinic acid regulates impaired macrophage autophagic flux in the treatment of non-alcoholic fatty liver disease.

Macrophages are involved in hepatocyte steatosis and necroinflammation and play an important role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Impaired autophagy function (decreased autophagy or blocked autophagic flow) leads to cell damage and death and promotes NAFLD progression. The experimental and clinical research of glycyrrhetinic acid (GA) in the treatment of NAFLD has gradually attracted attention with clear pharmacological activities such as immune regulation, antiviral, antitumor, antioxidant, liver protection, and anti-inflammatory. However, the effects of GA on the STAT3-HIF-1α pathway and autophagy in macrophages are still unclear, and its mechanism of action in the treatment of NAFLD remains to be further elucidated. We constructed a NAFLD mouse model through a high-fat and high-sugar diet to investigate the therapeutic effects of GA. The results showed that GA reduced weight, improved the pathological changes and hepatic lipid deposition of liver, and abnormally elevated the levels of serum biochemical (AST, ALT, TG, T-CHO, LDL-C, and HDL-C) and inflammatory indexes (IL-1ß, IL-4, IL-6, MCP-1, and TNF-α) in NAFLD mice. Further examination revealed that GA ameliorates excessive hepatic macrophage infiltration and hepatocyte apoptosis. The results of the cell experiments further elaborated that GA modulated the PA-induced macrophage STAT3-HIF-1α pathway and ameliorated impaired autophagic flux (blockade of autophagosome-lysosome fusion) and overactivation of inflammation. Excessive hepatocyte apoptosis caused by the uncontrolled release of inflammatory cytokines was also suppressed by GA. Conclusion: This study demonstrated that GA could regulate the STAT3-HIF-1α pathway of macrophages, ameliorate the impaired autophagy flux, and reduce the excessive production of inflammatory cytokines to improve the excessive apoptosis of liver cells, thus playing a therapeutic role on NAFLD.

Glycyrrhetinic acid: A potential drug for the treatment of COVID-19 cytokine storm.

BACKGROUND: The cytokine storm (CS) triggered by coronavirus disease 2019 (COVID-19) has caused serious harm to health of humanity and huge economic burden to the world, and there is a lack of effective methods to treat this complication. PURPOSE: In this research, we used network pharmacology and molecular docking to reveal the interaction mechanism in the glycyrrhetinic acid (GA) for the treatment of CS, and validated the effect of GA intervention CS by experiments. STUDY DESIGN: First, we screened corresponding target of GA and CS from online databases, and obtained the action target genes through the Venn diagram. Then, protein-protein interaction (PPI) network, Gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment of the action target genes were acquired by R language to predict its mechanism. Next, molecular docking was performed on core targets. Finally, experiments in which GA intervened in lipopolysaccharide (LPS)-induced CS were implemented. RESULTS: 84 action target genes were obtained from online database. The PPI network of target genes showed that TNF, IL6, MAPK3, PTGS2, ESR1 and PPARG were considered as the core genes. The results of GO and KEGG showed that action target genes were closely related to inflammatory and immune related signaling pathways, such as TNF signaling pathway, IL-17 signaling pathway, Human cytomegalovirus infection, PPAR signaling pathway and so on. Molecule docking results prompted that GA had fine affinity with IL6 and TNF proteins. Finally, in vivo and in vitro experimental results showed that GA could significantly inhibit LPS-induced CS. CONCLUSION: GA has a potential inhibitory effect on CS, which is worthy of further exploration.

Can Antiviral Activity of Licorice Help Fight COVID-19 Infection?

The phytotherapeutic properties of Glycyrrhiza glabra (licorice) extract are mainly attributed to glycyrrhizin (GR) and glycyrrhetinic acid (GA). Among their possible pharmacological actions, the ability to act against viruses belonging to different families, including SARS coronavirus, is particularly important. With the COVID-19 emergency and the urgent need for compounds to counteract the pandemic, the antiviral properties of GR and GA, as pure substances or as components of licorice extract, attracted attention in the last year and supported the launch of two clinical trials. In silico docking studies reported that GR and GA may directly interact with the key players in viral internalization and replication such as angiotensin-converting enzyme 2 (ACE2), spike protein, the host transmembrane serine protease 2, and 3-chymotrypsin-like cysteine protease. In vitro data indicated that GR can interfere with virus entry by directly interacting with ACE2 and spike, with a nonspecific effect on cell and viral membranes. Additional anti-inflammatory and antioxidant effects of GR cannot be excluded. These multiple activities of GR and licorice extract are critically re-assessed in this review, and their possible role against the spread of the SARS-CoV-2 and the features of COVID-19 disease is discussed.

Glycyrrhetinic acid remodels the tumor microenvironment and synergizes with doxorubicin for breast cancer treatment in a murine model.

We aimed to combine glycyrrhetinic acid with doxorubicin to prepare, characterize and evaluate a drug delivery nano-system with REDOX sensitivity for the treatment of breast cancer. M-DOX-GA NPs prepared by nano sedimentation were spherical, with a particle size of 181 nm. And the maximum encapsulation efficiency and drug loading in M-DOX-GA NPs were 89.28% and 18.22%, respectively. Cytotoxicity and cellular uptake experiments of nanoparticles to KC cells, Cal-27 cells and 4T1 cells were studied by the CCK-8 method. The result indicated that M-DOX-GA NPs could accurately release the drug into the tumor cells, thus achieving the targeted release of the drug. Comparing the survival rate of the above three cells, it was found that M-DOX-GA NPs had a good tumor selectivity and had a more significant therapeutic effect on breast cancer. A 4T1-bearing mouse model was established, and the tumor inhibition rate was 77.37% after injection of nanoparticle solution for 14 d. Normal tissue H&E stained sections and TUNEL assay were verified M-DOX-GA NPs have excellent tumor suppressive effect, and can efficiently reduce the toxic side effects on normal organisms, and effectively avoided 4T1 cells metastasis. Immunofluorescence detection and Western-blot analysis figured a decline in both CUGBP1 and α-SMA, which verifying the TME remodeling induced by glycyrrhetinic acid. Collectively, the combination of doxorubicin and glycyrrhetinic acid is an effective and safe strategy for remodeling fibrotic TME by improving the therapeutic outcome for breast cancer.

Administration of glycyrrhetinic acid reinforces therapeutic effects of mesenchymal stem cell-derived exosome against acute liver ischemia-reperfusion injury.

Recent studies have shown that mesenchymal stem cell-derived exosome could attenuate ischaemia-reperfusion (I/R) injury by suppressing inflammatory response in the liver. Glycyrrhetinic acid was also shown to be capable of repressing the TLR4 signalling pathway. However, it remains to be explored as whether the combined administration of mesenchyma stem cell (MSC)-derived exosome and glycyrrhetinic acid (GA) could increase their therapeutic effects on I/R injury. Western blot was performed to evaluate the expression of proteins associated with inflammatory response in THP-1 cells and I/R rat models treated under different conditions. Flow cytometry was carried out to analyse the proportions of different subtypes of peripheral blood cells in I/R rats. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured to assess the liver injury in I/R rats. Combined treatment with MSC-derived exosome and GA effectively maintained the expression of key proteins involved in inflammatory response in LPS stimulated THP-1 cells and THP-1 cells treated under hypoxia conditions. In the established of I/R rat models, GA administration reinforced the therapeutic efficiency of MSC-derived exosomes by maintaining the proportion of different subgroups of peripheral blood cells, decreasing the concentration of ALT and AST, and restoring the expression of dysregulated proteins associated with inflammation. Our results demonstrated that treatment with exosomes derived from mesenchymal stem cells (MSCs) attenuated liver I/R injury, while the pre-treatment with GA may further promote the therapeutic effect of mesenchymal stem cell-derived exosome against acute liver ischaemia-reperfusion injury.

Glycyrrhetinic acid alleviates acute lung injury by PI3K/AKT suppressing macrophagic Nlrp3 inflammasome activation.

Glycyrrhetinic acid (GA), a triterpene saponins, has been widely proven to have multiple medicinal properties. Our study aimed to figure out the protective effect of GA on acute lung injury (ALI) and the underlying mechanism. The LPS-induced ALI model mice were intratracheally administrated with 10 mg/kg LPS. Pretreatment with GA (10, 20, 40 mg/kg, i.g.) ameliorated acute lung injury pathological damage, macrophage infiltration and lung edema. In the lung tissue, immunofluorescence (IF) and Immunohistochemistry (IHC) were performed to detect macrophage Nod-like receptor 3 (Nlrp3) inflammasome activation and interleukin-1ß (IL-1ß) protein expression. In macrophages, the co-localization of Nlrp3 with caspase-1 and Nlrp3 with ASC were assessed by IF. The translational and transcriptional level of Nlrp3, cle-caspase-1 and apoptosis-associated speck-like protein containing CARD (ASC), were examined by Western blot and Real time PCR (RT-PCR). The protein expression of Cle-caspase-1 was remarkably suppressed via sh-Nlrp3 transfection compared with LPS groups. GA notably attenuated ALI by inhibiting Nlrp3 formation and activation. Furthermore, GA downregulated the production of reactive oxygen species (ROS) and the phosphorylation level of PI3K and AKT in macrophages. These findings indicate that GA ameliorated ALI in mice by suppressing the activation of Nlrp3 inflammasome which may be mediated by ROS-PI3K/AKT pathway. GA may serve as a promising agent for the attenuation of ALI-related inflammation and pathology.

Synthesis and antitumor effects of novel 18ß-glycyrrhetinic acid derivatives featuring an exocyclic α,ß-unsaturated carbonyl moiety in ring A.

A series of novel 18ß-glycyrrhetinic acid (GA) derivatives featuring an exocyclic α,ß-unsaturated carbonyl moiety in ring A were synthesized and evaluated for their antitumor activities. Compounds 5c and 5l showed stronger cytotoxicity than other compounds and reported GA analogue CDODA-Me (methyl 2-cyano-3,11-dioxo-18ß-olean-1,12-dien-30-oate). 5c and 5l induced apoptosis in cancer cells accompanying with c-Flip reduction and Noxa induction, associated with decreased HDAC3 expression and increased acetylation of H3. 5l displayed better stability properties than 5c and CDODA-Me in microsomes and plasma, 5l also showed favorable pharmacokinetic profiles and inhibited tumor growth in mice. Compound 5l represents a new type of GA derivatives with improved antitumor activity.

Clinical and noninvasive instrumental evaluation of the efficacy of a nonsteroidal anti-inflammatory 8-beta glycyrrhetinic acid cream for the treatment of erythema in rosacea.

Rosacea is a very common chronic facial dermatosis characterized by a multiphase evolution. Inflammation is an important reaction in rosacea not only due to inflammatory reactions to cutaneous microorganisms, such as Demodex follicolorum, but also to ultraviolet damage that generates reactive oxygen species. This study evaluated the efficacy and tolerability of a nonsteroidal anti-inflammatory 18-beta glycyrrhetinic acid cream for the treatment of mild rosacea by means of noninvasive methods. A total of 24 subjects suffering from erythemato-telangiectatic or mild papulo-pustular rosacea were recruited in the trial. Twelve patients applied an anti-inflammatory cream with 18-beta glycyrrhetinic acid twice daily for 20 days and 12 patients, recruited as control, applied the same formulation without 18-beta glycyrrhetinic acid. After 10 days of treatment, a significant reduction of erythema was recorded in the patient sample who applied the 18-beta glycyrrhetinic acid cream, the mean change from baseline showed an increase in hydration level of the skin surface but it was not statistically significant. The use of 18-beta glycyrrhetinic acid cream can be helpful in managing symptoms and condition of rosacea skin, especially in the management of erythema.

Gan Shen Fu Fang ameliorates liver fibrosis in vitro and in vivo by inhibiting the inflammatory response and extracellular signal-regulated kinase phosphorylation.

BACKGROUND: Liver fibrosis is a common health problem worldwide and there is still a lack of effective medicines. The Chinese herbal medicine, Gan Shen Fu Fang (GSFF) is composed of salvianolic acid B and diammonium glycyrrhizinate. In this study, we observed the effects of GSFF on liver fibrosis in vivo and in vitro in an attempt to provide some hope for the treatment. AIM: To observe the effects of GSFF on liver fibrosis in vivo and in vitro and investigate the mechanism from the perspective of the inflammatory response and extracellular signal-regulated kinase (ERK) phosphorylation. METHODS: Common bile duct-ligated rats were used for in vivo experiments. Hepatic stellate cells-T6 (HSC-T6) cells were used for in vitro experiments. Hematoxylin and eosin staining and Masson staining, biochemical assays, hydroxyproline (Hyp) assays, enzyme-linked immunoasorbent assay and western blotting were performed to evaluate the degree of liver fibrosis, liver function, the inflammatory response and ERK phosphorylation. The CCK8 assay, immunofluorescence and western blotting were applied to test the effect of GSFF on HSC-T6 cell activation and determine whether GSFF had an effect on ERK phosphorylation in HSC-T6 cells. RESULTS: GSFF improved liver function and inhibited liver fibrosis in common bile duct-ligated rats after 3 wk of treatment, as demonstrated by histological changes, hydroxyproline assays and collagen I concentrations. GSFF alleviated inflammatory cell infiltration and reduced the synthesis of pro-inflammatory cytokines [tumor necrosis factor-α (TNF-α) and interlukin-1ß] and NF-κB. In addition, GSFF decreased ERK phosphorylation. In vitro, GSFF inhibited the viability of HSC-T6 cells with and without transforming growth factor ß1 (TGF-ß1) stimulation and decreased the synthesis of collagen I. GSFF had the greatest effect at a concentration of 0.5 µmol/L. GSFF inhibited the expression of α-smooth muscle actin (α-SMA), a marker of HSC activation, in HSC-T6 cells. Consistent with the in vivo results, GSFF also inhibited the phosphorylation of ERK and downregulated the expression of NF-κB. CONCLUSION: GSFF inhibited liver fibrosis progression in vivo and HSC-T6 cell activation in vitro. These effects may be related to an alleviated inflammatory response and downregulated ERK phosphorylation.

18ß-Glycyrrhetinic acid induces human HaCaT keratinocytes apoptosis through ROS-mediated PI3K-Akt signaling pathway and ameliorates IMQ-induced psoriasis-like skin lesions in mice.

BACKGROUND: Psoriasis is a chronic inflammatory skin disease affecting 2-3% of the population worldwide. Hyperproliferative keratinocytes were thought to be an amplifier of inflammatory response, thereby sustaining persistence of psoriasis lesions. Agents with the ability to inhibit keratinocyte proliferation or induce apoptosis are potentially useful for psoriasis treatment. 18ß-Glycyrrhetinic acid (GA), an active metabolite of glycyrrhizin, exhibits diverse pharmacological activities, including anti-inflammatory, anti-bacteria and anti-proliferation. The current study aims to evaluate the effects of GA on the proliferation and apoptosis of human HaCaT keratinocytes in vitro and investigate the effects of GA on the skin lesions of imiquimod (IMQ)-induced psoriasis-like mouse model in vivo. METHODS: Cell viability was assayed by CCK-8. Flow cytometry was performed to measure apoptosis and reactive oxygen species (ROS), with Annexin V-FITC/PI detection kit and DCFH-DA probe respectively. Caspase 9/3 activities were measured using caspase activity assay kits. The protein levels of Akt and p-Akt were determined using Western blotting. IMQ was applied to induce psoriasis-like skin lesions in mice. The histological change in mouse skin lesions was detected using hematoxylin and eosin (H&E) staining. The severity of skin lesions was scored based on Psoriasis Area Severity Index (PASI). RT-PCR was employed to examine the relative expression of TNF-α, IL-22 and IL-17A in mouse skin lesions. RESULTS: GA decreased HaCaT keratinocytes viability and induced cell apoptosis in a dose-dependent manner. In the presence of GA, intracellular ROS levels were significantly elevated. NAC, a ROS inhibitor, attenuated GA-mediated HaCaT keratinocytes growth inhibition and apoptosis. In addition, GA treatment remarkably decreased p-Akt protein level, which could be restored partially when cells were co-treated with GA and NAC. LY294002 (a PI3K inhibitor) treatment significantly enhanced GA-mediated cytotoxicity. Moreover, GA ameliorated IMQ-induced psoriasis-like skin lesions in mice. CONCLUSIONS: GA inhibits proliferation and induces apoptosis in HaCaT keratinocytes through ROS-mediated inhibition of PI3K-Akt signaling pathway, and ameliorates IMQ-induced psoriasis-like skin lesions in mice.

HMGB1 in nasal inflammatory diseases: a reappraisal 30 years after its discovery.

INTRODUCTION: High mobility group protein box 1 (HMGB1) is a protein belonging to the alarmin family. HMGB1 has a relevant role in starting the inflammatory cascade by means of receptors, such as RAGE and TLR. HMGB1 supports transcription of many genes in interactions with many transcription factors, including NF-kB. The axis HMGB1-RAGE-NF-kB has, therefore, a pivotal role in the inflammatory cascade. HMGB1 controls the production of several pro-inflammatory cytokines and the proliferation and activation of many inflammatory cells. AREAS COVERED: The present report concerns the role of HMGB1 in nasal inflammatory disorders, including allergic and non-allergic rhinitis, and chronic rhinosinusitis with nasal polyps. HMGB1 modulation has been the aim of several studies. The literature search included recent papers that covered this topic. EXPERT OPINION: As HMGB1 has a pivotal role in inflammatory events, its modulation could be attractive for designing new therapeutic strategies. In this regard, glycyrrhetic acid (GA), the active component of Glycyrrhiza glabra, can efficiently block HMGB1. Promising reports seem to suggest that GA could exert favorable anti-inflammatory activity in patients with nasal inflammatory disorders.

A Previously Unknown Drug-Drug Interaction Is Suspected in Delayed Elimination of Plasma Methotrexate in High-Dose Methotrexate Therapy.

Background: High-dose methotrexate (HD-MTX) therapy is widely implemented for leukemia, osteosarcoma, and lymphoma. Although various measures have been taken to avoid toxicity from high serum MTX concentrations, there are many cases of delayed elimination of MTX. Objective: We suspected that delayed elimination of serum MTX was caused by unknown interactions between MTX and concomitant drugs. Methods: Concerning concomitant drugs in the case of delayed elimination of MTX, we performed screening tests in 35 patients who had undergone HD-MTX therapy. We then investigated the risk factors for delayed MTX elimination in 94 patients with leukemia, lymphoma, or osteosarcoma retrospectively. Results: The percentages of concomitant use of Stronger Neo-Minophagen C (SNMC), a glycyrrhizin preparation, and vincristine were higher in the delayed group. The percentage of delayed MTX elimination in patients receiving HD-MTX therapy was 41%. Multiple logistic regression analysis revealed that the concomitant use of SNMC solely was a significant risk factor for delayed MTX (odds ratio = 12.20; 95% CI = 1.06-139.84). Conclusion and Relevance: Concomitant use of SNMC was shown to be related to delayed elimination of serum MTX, and our results suggested a previously unknown drug-drug interaction between MTX and SNMC.

Glycyrrhetinic acid pretreatment attenuates liver ischemia/reperfusion injury via inhibiting TLR4 signaling cascade in mice.

Glycyrrhetinic acid (GA), the main bioactive substances of glycyrrhiza uralensis Fisch, has been reported to exhibit hepatoprotective and anti-inflammatory properties. However, the effects and underlying mechanisms of GA in liver ischemia/reperfusion (I/R) injury remain elusive. In this study, mice were pretreated with GA (100 mg/kg) three times a day by gavage prior to I/R injury, and then hepatic histopathological damages, biochemical parameters and inflammatory molecules were evaluated. We found that mice performed with liver I/R showed a significantly increase in plasma aminotransferase (ALT), aspartate aminotransferase (AST), liver cell apoptosis and infiltration of neutrophils compared with the control group. GA pretreatment notably improved liver function, histopathology of liver tissues, and lowered liver cell apoptosis and infiltration of neutrophils. Besides, further analysis indicated that GA pretreatment reduced I/R-induced expression of extracellular HMGB1, inhibited activation of TLR4 and following phosphorylation of IRAK1, ERK, P38 and NF-κB, and attenuated TNF-α and IL-1ß production. These data suggested that GA protected against liver I/R injury through a HMGB1-TLR4 signaling pathway and it might be a promising drug for future clinical use in liver transplantation.

Protective effects of 18ß-glycyrrhetinic acid on pulmonary arterial hypertension via regulation of Rho A/Rho kinsase pathway.

PURPOSE: Excessive proliferation, migration and anti-apoptosis of pulmonary artery smooth muscle cells (PASMCs) are the basis for the development of pulmonary vascular remodeling, and it is the driving force for pulmonary arterial hypertension (PAH). 18ß-glycyrrhetinic acid (18ß-GA) is the main active substance extracted from Chinese herbal medicine licorice, with outstanding anti-inflammatory, anti-oxidation and anti-proliferative effects. Our team found in previous studies that 18ß-GA has protective effects on monocrotaline-induced PAH in rats. However, the anti-angiogenic effect of 18ß-GA on PAH remains unclear. Therefore, in order to further investigate whether the beneficial effects of 18ß-GA on PAH are related to its antiproliferative effect, we conducted experiments in vivo and in vitro. METHODS AND RESULTS: In vivo, 18ß-GA relieved mean pulmonary arterial pressure, right ventricular systolic pressure, and right ventricular hypertrophy index, improving pulmonary remodeling. In vitro, 18ß-GA significantly inhibited PDGF-BB-induced proliferation and DNA synthesis of HPASMCs, blocking the progression of G0/G1 to S phase of the cell cycle. Furthermore, after treatment with 18ß-GA, the expression of Rho A, ROCK1, ROCK2 was decreased and ROCK activity was inhibited in HPASMC. In addition, 18ß-GA also attenuated PDGF-induced changes in p27kip1, Bax and Bcl-2. CONCLUSIONS: In summary, these results indicate that 18ß-GA regulates the activity of RhoA-ROCK signaling pathway, inhibits the proliferation of HPASMCs, and has potential value in the treatment of PAH.