Astragaloside IV inhibits cell viability and glycolysis of hepatocellular carcinoma by regulating KAT2A-mediated succinylation of PGAM1.

BACKGROUND: Astragaloside IV (AS-IV) is one of the basic components of Astragali radix, that has been shown to have preventive effects against various diseases, including cancers. This study aimed to explore the role of AS-IV in hepatocellular carcinoma (HCC) and its underlying mechanism. METHODS: The cell viability, glucose consumption, lactate production, and extracellular acidification rate (ECAR) in SNU-182 and Huh7 cell lines were detected by specific commercial kits. Western blot was performed to analyze the succinylation level in SNU-182 and Huh7 cell lines. The interaction between lysine acetyltransferase (KAT) 2 A and phosphoglycerate mutase 1 (PGAM1) was evaluated by co-immunoprecipitation and immunofluorescence assays. The role of KAT2A in vivo was explored using a xenografted tumor model. RESULTS: The results indicated that AS-IV treatment downregulated the protein levels of succinylation and KAT2A in SNU-182 and Huh7 cell lines. The cell viability, glucose consumption, lactate production, ECAR, and succinylation levels were decreased in AS-IV-treated SNU-182 and Huh7 cell lines, and the results were reversed after KAT2A overexpression. KAT2A interacted with PGAM1 to promote the succinylation of PGAM1 at K161 site. KAT2A overexpression promoted the viability and glycolysis of SNU-182 and Huh7 cell lines, which were partly blocked following PGAM1 inhibition. In tumor-bearing mice, AS-IV suppressed tumor growth though inhibiting KAT2A-mediated succinylation of PGAM1. CONCLUSION: AS-IV inhibited cell viability and glycolysis in HCC by regulating KAT2A-mediated succinylation of PGAM1, suggesting that AS-IV might be a potential and suitable therapeutic agent for treating HCC.

An In Vitro Study on the Cytotoxic, Antioxidant, and Antimicrobial Properties of Yamogenin-A Plant Steroidal Saponin and Evaluation of Its Mechanism of Action in Gastric Cancer Cells.

Yamogenin is a steroidal saponin occurring in plant species such as Asparagus officinalis, Dioscorea collettii, Trigonella foenum-graecum, and Agave sp. In this study, we evaluated in vitro cytotoxic, antioxidant, and antimicrobial properties of yamogenin. The cytotoxic activity was estimated on human colon cancer HCT116, gastric cancer AGS, squamous carcinoma UM-SCC-6 cells, and human normal fibroblasts with MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. The amount of apoptotic and dead AGS cells after treatment with yamogenin was estimated with flow cytometry. Also, in yamogenin-treated AGS cells we investigated the reactive oxygen species (ROS) production, mitochondrial membrane depolarization, activity level of caspase-8 and -9, and gene expression at mRNA level with flow cytometry, luminometry, and RT-PCR, respectively. The antioxidant properties of yamogenin were assessed with DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) assays. The antimicrobial potential of the compound was estimated on Staphylococcus aureus, Bacillus cereus, Klebsiella pneumoniae, Escherichia coli, Salmonella enterica, Helicobacter pylori, Campylobacter coli, Campylobacter jejuni, Listeria monocytogenes, Lactobacillus paracasei, and Lactobacillus acidophilus bacteria strains. Yamogenin showed the strongest cytotoxic effect on AGS cells (IC50 18.50 ± 1.24 µg/mL) among the tested cell lines. This effect was significantly stronger in combinations of yamogenin with oxaliplatin or capecitabine than for the single compounds. Furthermore, yamogenin induced ROS production, depolarized mitochondrial membrane, and increased the activity level of caspase-8 and -9 in AGS cells. RT-PCR analysis revealed that this sapogenin strongly up-regulated TNFRSF25 expression at the mRNA level. These results indicate that yamogenin induced cell death via the extrinsic and intrinsic way of apoptosis. Antioxidant study showed that yamogenin had moderate in vitro potential (IC50 704.7 ± 5.9 µg/mL in DPPH and 631.09 ± 3.51 µg/mL in ABTS assay) as well as the inhibition of protein denaturation properties (with IC50 1421.92 ± 6.06 µg/mL). Antimicrobial test revealed a weak effect of yamogenin on bacteria strains, the strongest one being against S. aureus (with MIC value of 350 µg/mL). In conclusion, yamogenin may be a potential candidate for the treatment and prevention of gastric cancers.

Efficacy of substances containing 3 types of active ingredients-saponins, flavones, and alkaloids in regulation of cytokines in autoimmune diseases a systematic review and Meta-analysis based on animal studies.

OBJECTIVE: To investigate the efficacy of substances containing 3 types of active ingredients-saponins, flavones, and alkaloids on experimental animals with autoimmune diseases (AIDs). METHODS: The protocol for this systematic review and Meta-analysis was prospectively registered with PROSPERO (CRD42023395741). Searches were conducted in the China National Knowledge Infrastructure, Wanfang, Chinese Science and Technology Journals, China Biomedical, PubMed, Cochrane Library, and Embase databases to screen for animal studies investigating the therapeutic effects of saponins, flavones, or alkaloids on autoimmune diseases; consequently, corresponding data extraction tables were prepared. Systematic Review Centre for Laboratory Animal Experimentation was used to assess the risk of methodological bias in the included literature. RevMan 5.4 was used for the Meta-analysis on the 8 serum cytokines. RESULTS: A total of 31 studies were included, all of which were randomized controlled studies. Meta-analysis indicated that substances rich in saponins, flavones, and alkaloids reduced serum levels of interleukin (IL)-1ß [standardized mean difference (SMD) = -1.94, 95% confidence interval (CI) (-2.99, -0.90), P = 0.0003], IL-6 [SMD = -1.65, 95% CI (-2.33, -0.97,) P < 0.000 01], IL-17 [SMD = -2.41, 95% CI (-3.61, -1.20), P < 0.0001], tumor necrosis factor (TNF)-α [SMD = -1.84, 95% CI (-2.61, -1.06), P < 0.0001], and interferon (IFN)-γ [SMD = -1.54, 95% CI (-2.43, -0.65), P = 0.0007], but increased serum levels of IL-4 [SMD = 1.30, 95% CI (0.15, 2.44), P = 0.03) and IL-10 [SMD = 2.05, 95% CI (1.39, 2.70), P < 0.000 01) in animal models. However, no significant regulatory effect of these three active components was observed on serum levels of IL-2 [SMD = -0.63, 95% CI (-1.82, 0.57), P = 0.30]. CONCLUTIONS: Substances containing saponins, flavones, and alkaloids regulated the changes of immune-related cytokines, it may be a novel dietary substance to relieve and control autoimmune diseases in the future.

Platycodon grandiflorum saponins: Ionic liquid-ultrasound-assisted extraction, antioxidant, whitening, and antiaging activity.

This study explored the use of ionic liquid-ultrasound (ILU)-assisted extraction to enhance the extraction rate of Platycodon grandiflorum saponins (PGSs), and the content, extraction mechanism, antioxidant activity, whitening, and antiaging activity of PGSs prepared using ILU, ultrasound-water, thermal reflux-ethanol, and cellulase hydrolysis were compared. The ILU method particularly disrupted the cell wall, improved PGS extraction efficiency, and yielded a high total saponin content of 1.45 ± 0.02 mg/g. Five monomeric saponins were identified, with platycodin D being the most abundant at 1.357 mg/g. PGSs displayed excellent in vitro antioxidant activity and exhibited inhibitory effects on tyrosinase, elastase, and hyaluronidase. The results suggest that PGSs may have broad antioxidant, skin-whitening, and antiaging potential to a large extent. Overall, this study provided valuable insights into the extraction, identification, and bioactivities of PGSs, which could serve as a reference for future development and application of these compounds in the functional foods industry.

Polyphyllin VII Promotes Apoptosis in Breast Cancer by Inhibiting MAPK/ERK Signaling Pathway through Downregulation of SOS1.

Polyphyllin VII is a biologically active herbal monomer extracted from the traditional Chinese herbal medicine Chonglou. Many studies have demonstrated the anticancer activity of polyphyllin VII against various types of cancers, such as colon, liver, and lung cancer, but its effect on breast cancer has not been elucidated. In this study, we demonstrate that polyphyllin VII inhibited proliferation, increased production of intracellular reactive oxygen species, and decreased mitochondrial membrane potential in breast cancer cells. Notably, polyphyllin VII also induced apoptosis via the mitochondrial pathway. Transcriptome sequencing was used to analyze the targets of PPVII in regulating breast cancer cells. Mechanistic studies showed that polyphyllin VII downregulated Son of Sevenless1 (SOS1) and inhibited the MAPK/ERK pathway. Furthermore, PPVII exerted strong antitumor effects in vivo in nude mice injected with breast cancer cells. Our results suggest that PPVII may promote apoptosis through regulating the SOS1/MAPK/ERK pathway, making it a possible candidate target for the treatment of breast cancer.

Panax notoginseng Saponins Activate Nuclear Factor Erythroid 2-Related Factor 2 to Inhibit Ferroptosis and Attenuate Inflammatory Injury in Cerebral Ischemia-Reperfusion.

Panax notoginseng saponins (PNS), the primary medicinal ingredient of Panax notoginseng, mitigates cerebral ischemia-reperfusion injury (CIRI) by inhibiting inflammation, regulating oxidative stress, promoting angiogenesis, and improving microcirculation. Moreover, PNS activates nuclear factor erythroid 2-related factor 2 (Nrf2), which is known to inhibit ferroptosis and reduce inflammation in the rat brain. However, the molecular regulatory roles of PNS in CIRI-induced ferroptosis remain unclear. In this study, we aimed to investigate the effects of PNS on ferroptosis and inflammation in CIRI. We induced ferroptosis in SH-SY5Y cells via erastin stimulation and oxygen glucose deprivation/re-oxygenation (OGD/R) in vitro. Furthermore, we determined the effect of PNS treatment in a rat model of middle cerebral artery occlusion/reperfusion and assessed the underlying mechanism. We also analyzed the changes in the expression of ferroptosis-related proteins and inflammatory factors in the established rat model. OGD/R led to an increase in the levels of ferroptosis markers in SH-SY5Y cells, which were reduced by PNS treatment. In the rat model, combined treatment with an Nrf2 agonist, Nrf2 inhibitor, and PNS-Nrf2 inhibitor confirmed that PNS promotes Nrf2 nuclear localization and reduces ferroptosis and inflammatory responses, thereby mitigating brain injury. Mechanistically, PNS treatment facilitated Nrf2 activation, thereby regulating the expression of iron overload and lipid peroxidation-related proteins and the activities of anti-oxidant enzymes. This cascade inhibited ferroptosis and mitigated CIRI. Altogether, these results suggest that the ferroptosis-mediated activation of Nrf2 by PNS reduces inflammation and is a promising therapeutic approach for CIRI.

The Regulatory Mechanism of Smilax China L. Saponins against Nonalcoholic Fatty Liver Is Revealed by Metabolomics and Transcriptomics.

This study was to investigate the effects of Smilax China L. saponins (SCS) on non-alcoholic fatty liver disease (NAFLD). Rats were fed a high-fat diet (HFD) for 8 weeks to induce NAFLD, followed by SCS treatment for 8 weeks. The effect of SCS on liver injury was observed by H&E staining and the regulative mechanism of SCS on lipid formation was exposed by detecting Oil red O, insulin resistance (IR), and fatty acids synthesis (FAS). Furthermore, transcriptomics and metabolomics were performed to analyze the potential targets. The experimental results indicated that SCS exerted a positive curative effect in alleviating HFD-induced overweight, hepatic injury, steatosis, and lipid formation and accumulation in rats, and the preliminary mechanism studies showed that SCS could alleviate IR, inhibit FAS expression, and reduce Acetyl-CoA levels. Besides, the integrative analysis of transcriptomics and metabolomics exposed the targets of SCS to regulate lipid production likely being the sphingolipid metabolism and glycerophospholipid metabolism pathways. This study demonstrates that SCS significantly ameliorates lipid metabolic disturbance in rats with NAFLD by relieving insulin resistance, inhibiting the FAS enzymes, and regulating the sphingolipid and glycerophospholipid metabolism pathways.

Green Preparation and Antibacterial Activity Evaluation of AgNPs-Blumea balsamifera Oil Nanoemulsion.

Bacterial infection is a thorny problem, and it is of great significance to developing green and efficient biological antibacterial agents that can replace antibiotics. This study aimed to rapidly prepare a new type of green antibacterial nanoemulsion containing silver nanoparticles in one step by using Blumea balsamifera oil (BBO) as an oil phase and tea saponin (TS) as a natural emulsifier and reducing agent. The optimum preparation conditions of the AgNPs@BBO-TS NE were determined, as well as its physicochemical properties and antibacterial activity in vitro being investigated. The results showed that the average particle size of the AgNPs@BBO-TS NE was 249.47 ± 6.23 nm, the PDI was 0.239 ± 0.003, and the zeta potential was -35.82 ± 4.26 mV. The produced AgNPs@BBO-TS NE showed good stability after centrifugation and 30-day storage. Moreover, the AgNPs@BBO-TS NE had an excellent antimicrobial effect on Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. These results demonstrated that the AgNPs@BBO-TS NE produced in this study can be used as an efficient and green antibacterial agent in the biomedical field.

Phytochemical study of Magonia pubescens A. St.-Hil. and cytotoxicity of branches aqueous extract on breast cancer and leukemia cells.

Magonia pubescens is a natural species from the Brazilian cerrado biome. Its fruits and seeds are used in the treatment of seborrheic dermatitis, a common inflammatory skin disease. In this work, the known compounds lapachol, stigmasterol, maniladiol and scopoletin were isolated from hexane and dichloromethane extracts of M. pubescens branches. The aqueous extract of this material was fractioned through a liquid-liquid partition and the obtained fractions were analyzed by UHPLC-MS/MS. The results obtained were compared with data from three databases, leading to the putative identification of 51 compounds from different classes, including flavonoids, saponins and triterpenes. The cytotoxicity of aqueous fractions was assayed against breast cancer (MDA-MB-231) and leukemia (THP-1 and K562) cells. The best activity was observed for fraction AE3 against MDA-MB-231 cells (IC50 30.72 µg.mL-1).

Effects of Saponins on Lipid Metabolism: The Gut-Liver Axis Plays a Key Role.

Unhealthy lifestyles (high-fat diet, smoking, alcohol consumption, too little exercise, etc.) in the current society are prone to cause lipid metabolism disorders affecting the health of the organism and inducing the occurrence of diseases. Saponins, as biologically active substances present in plants, have lipid-lowering, inflammation-reducing, and anti-atherosclerotic effects. Saponins are thought to be involved in the regulation of lipid metabolism in the body; it suppresses the appetite and, thus, reduces energy intake by modulating pro-opiomelanocortin/Cocaine amphetamine regulated transcript (POMC/CART) neurons and neuropeptide Y/agouti-related peptide (NPY/AGRP) neurons in the hypothalamus, the appetite control center. Saponins directly activate the AMP-activated protein kinase (AMPK) signaling pathway and related transcriptional regulators such as peroxisome-proliferator-activated-receptors (PPAR), CCAAT/enhancer-binding proteins (C/EBP), and sterol-regulatory element binding proteins (SREBP) increase fatty acid oxidation and inhibit lipid synthesis. It also modulates gut-liver interactions to improve lipid metabolism by regulating gut microbes and their metabolites and derivatives-short-chain fatty acids (SCFAs), bile acids (BAs), trimethylamine (TMA), lipopolysaccharide (LPS), et al. This paper reviews the positive effects of different saponins on lipid metabolism disorders, suggesting that the gut-liver axis plays a crucial role in improving lipid metabolism processes and may be used as a therapeutic target to provide new strategies for treating lipid metabolism disorders.

Dammarane-type triterpenoids from Gynostemma longipes and their protective activities on hypoxia-induced injury in PC12 cells.

Sixteen new dammarane-type triterpenoid saponins (1-16) featuring diverse structural variations in the side chain at C-17, along with twenty-one known analogues (17-37), have been isolated from the rhizomes of Gynostemma longipes C. Y. Wu, a plant renowned for its medicinal and edible properties. The structural elucidation of these compounds was accomplished through comprehensive analyses of 1D and 2D NMR and HRMS spectroscopic data, supplemented by comparison with previously reported data. Subsequent assays on the isolates for their protective effects against hypoxia-induced damage in pheochromocytoma cells (PC12 cells) revealed that nine saponins exhibited significant anti-hypoxic activities. Further investigation into the anti-hypoxia mechanisms of the representative saponins demonstrated that compounds 22 and 36 markedly reduced the levels of hypoxia-induced apoptosis. Additionally, these compounds were found to decrease the release of lactate dehydrogenase (LDH) and malondialdehyde (MDA), while increasing the activity of superoxide dismutase (SOD), thereby indicating that the saponins could mitigate hypoxia-induced injuries by ameliorating apoptosis and oxidative stress. These findings offer substantial evidence for the future utilization and development of G. longipes, identifying dammarane-type triterpenoid saponins as its active anti-hypoxic constituents.

Anti-MRSA mechanism of spirostane saponin in Rohdea pachynema F.T.Wang & tang.

ETHNOPHARMACOLOGY RELEVANCE: Rohdea pachynema F.T.Wang & Tang (R. pachynema), is a traditional folk medicine used for the treatment of stomach pain, stomach ulcers, bruises, and skin infections in China. Some of the diseases may relate to microbial infections in traditional applications. However few reports on its antimicrobial properties and bioactive components. AIM OF THE STUDY: To identify its bioactive constituents against methicillin-resistant Staphylococcus aureus (MRSA) in vitro and in vivo, and its mechanism. MATERIALS AND METHODS: The anti-MRSA ingredient 6α-O-[ß-D-xylopyranosyl-(1 â†’ 3)-ß-D-quinovopyranosyl]-(25S)-5α-spirostan-3ß-ol (XQS) was obtained from R. pachynema by phytochemical isolation. Subsequently, XQS underwent screening using the broth microdilution method and growth inhibition curves to assess its antibacterial activity. The mechanism of XQS was evaluated by multigeneration induction, biofilm resistance assay, scanning electron microscopy, transmission electron microscopy, and metabolomics. Additionally, a mouse skin infection model was established in vivo. RESULTS: 26 compounds were identified from the R. pachynema, in which anti-MRSA spirostane saponin (XQS) was reported for the first time with a minimum inhibitory concentration (MIC) of 8 µg/mL. XQS might bind to peptidoglycan (PGN) of the cell wall, phosphatidylglycerol (PG), and phosphatidylethanolamine (PE) of the cell membrane, then destroying the cell wall and the cell membrane, resulting in reduced membrane fluidity and membrane depolarization. Furthermore, XQS affected MRSA lipid metabolism, amino acid metabolism, and ABC transporters by metabolomics analysis, which targeted cell walls and membranes causing less susceptibility to drug resistance. Furthermore, XQS (8 mg/kg) recovered skin wounds in mice infected by MRSA effectively, superior to vancomycin (8 mg/kg). CONCLUSIONS: XQS showed anti-MRSA bioactivity in vitro and in vivo, and its mechanism association with cell walls and membranes was reported for the first, which supported the traditional uses of R. pachynema and explained its sensitivity to MRSA.

[Anti-tumor mechanism of total saponins of Paridis Rhizoma on inducing ferroptosis of breast cancer MCF-7 cells].

This study aims to investigate the mechanism of total saponins of Paridis Rhizoma in inducing the ferroptosis of MCF-7 cells and provide a theoretical basis for the clinical treatment of breast cancer with total saponins of Paridis Rhizoma. The methyl thiazolyl tetrazolium(MTT) assay was employed to examine the effects of different concentrations of total saponins of Paridis Rhizoma on the proliferation of MCF-7 cells. A phase contrast inverted microscope was used to observe the morphological changes of MCF-7 cells. The colony formation assay was employed to test the colony formation of MCF-7 cells. The lactate dehydrogenase(LDH) release test was conducted to determine the cell membrane integrity of MCF-7 cells. The cell scratch assay was employed to examine the migration of MCF-7 cells. After that, the level of reactive oxygen species(ROS) in MCF-7 cells was observed by an inverted fluorescence microscope, and the content of Fe~(2+) in MCF-7 cells was detected by the corresponding kit. Transmission electron microscopy was employed to observe the mitochondrial ultrastructure of MCF-7 cells. Western blot was employed to determine the expression of ferroptosis-related proteins, such as p53, solute carrier family 7 member 11(SLC7A11), glutathione peroxidase 4(GPX4), acyl-CoA synthetase long-chain family member 4(ACSL4), and transferrin receptor protein 1(TFR1) in MCF-7 cells. The results showed that 1.5, 3, 4.5, 6, 7.5, and 9 µg·mL~(-1) total saponins of Paridis Rhizoma significantly inhibited the proliferation of MCF-7 cells, with the IC_(50) of 4.12 µg·mL~(-1). Total saponins of Paridis Rhizoma significantly damaged the morphology of MCF-7 cells, leading to the formation of vacuoles and the gradual shrinkage and detachment of cells. Meanwhile, total saponins of Paridis Rhizoma inhibited the colony formation of MCF-7 cells, destroyed the cell membrane(leading to the release of LDH), and shortened the migration distance of MCF-7 cells. Total saponins of Paridis Rhizoma treatment significantly increased the content of ROS, induced oxidative damage, and led to the accumulation of Fe~(2+) in MCF-7 cells. Furthermore, total saponins of Paridis Rhizoma changed the mitochondrial structure, increased the mitochondrial membrane density, led to the decrease or even disappear of ridges, promoted the expression of p53 protein, down-regulated the expression of SLC7A11 and GPX4, and up-regulated the expression of ACSL4 and TFR1. In summary, total saponins of Paridis Rhizoma can significantly inhibit the proliferation and migration of MCF-7 cells and destroy the cell structure by inducing ferroptosis.

[Mechanism of total saponins of Panax japonicus against liver injury induced by acetaminophen in mice based on ERK/NF-κB/COX-2 signaling pathway].

This study investigated the effects and mechanisms of total saponins of Panax japonicus(TSPJ) against liver injury induced by acetaminophen(APAP). Male Kunming mice were randomly divided into a blank control group, TSPJ group(200 mg·kg~(-1), ig), model group, APAP+ TSPJ low-dose group(50 mg·kg~(-1), ig), APAP+ TSPJ medium-dose group(100 mg·kg~(-1), ig), APAP+ TSPJ high-dose group(200 mg·kg~(-1), ig), and APAP+ N-acetyl-L-cysteine group(200 mg·kg~(-1), ip). The administration group received the corresponding medications via ig or ip once a day for 14 consecutive days. After the last administration for one hour, except for the blank control group and TSPJ group, all groups of mice were given 500 mg·kg~(-1) APAP by gavage. After 24 hours, mouse serum and liver tissue were collected for serum alanine aminotransferase(ALT), aspartate aminotransferase(AST), reactive oxygen species(ROS), tumor necrosis factor alpha(TNF-α), interleukin-1 beta(IL-1ß), cyclooxygenase-2(COX-2), IL-6, IL-4, IL-10, as well as lactate dehydrogenase(LDH), glutathione(GSH), superoxide dismutase(SOD), catalase(CAT), total antioxidant capacity(T-AOC), malondialdehyde(MDA), and myeloperoxidase(MPO) liver tissue. Hematoxylin-eosin staining was used to observe the morphological changes of liver tissue. The mRNA expression levels of lymphocyte antigen 6G(Ly6G), galectin 3(Mac-2), TNF-α, IL-1ß, COX-2, IL-6, IL-4, and IL-10 in liver tissue were determined by quantitative real-time polymerase chain reaction(PCR). Western blot was utilized to detect the protein expression levels of Ly6G, Mac-2, extracellular regulated protein kinases(ERK), phosphorylated extracellular regulated protein kinases(p-ERK), COX-2, inhibitor of nuclear factor κB protein α(IκBα), phosphorylated inhibitor of nuclear factor κB protein α(p-IκBα), and nuclear factor-κB subunit p65(NF-κB p65) in cytosol and nucleus in liver tissue. The results manifested that TSPJ dramatically reduced liver coefficient, serum ALT, AST, ROS, TNF-α, IL-1ß, IL-6, and COX-2 levels, LDH, MPO, and MDA contents in liver tissue, and mRNA expressions of TNF-α, IL-1ß, and IL-6 in APAP-induced liver injury mice. It prominently elevated serum IL-4 and IL-10 levels, GSH, CAT, SOD, and T-AOC contents, and mRNA expressions of IL-4 and IL-10 in liver tissue, improved the degree of liver pathological damage, and suppressed neutrophil infiltration and macrophage recruitment in liver tissue. In addition, TSPJ lessened the mRNA and protein expressions of neutrophil marker Ly6G, macrophage marker Mac-2, and COX-2 in liver tissue, protein expressions of p-ERK, p-IκBα, and NF-κB p65 in nuclear, and p-ERK/ERK and p-IκBα/p-IκBα ratios and hoisted protein expression of NF-κB p65 in cytosol. These results suggest that TSPJ has a significant protective effect on APAP-induced liver injury in mice, and it can alleviate APAP-induced oxidative damage and inflammatory response. Its mechanism may be related to suppressing ERK/NF-κB/COX-2 signaling pathway activation, thus inhibiting inflammatory cell infiltration, cytokine production, and liver cell damage.

Astragaloside IV alleviates renal fibrosis by inhibiting renal tubular epithelial cell pyroptosis induced by urotensin II through regulating the cAMP/PKA signaling pathway.

OBJECTIVE: To explore the molecular mechanism of Astragaloside IV (AS-IV) in alleviating renal fibrosis by inhibiting Urotensin II-induced pyroptosis and epithelial-mesenchymal transition of renal tubular epithelial cells. METHODS: Forty SD rats were randomly divided into control group without operation: gavage with 5ml/kg/d water for injection and UUO model group: gavage with 5ml/kg/d water for injection; UUO+ AS-IV group (gavage with AS-IV 20mg/kg/d; and UUO+ losartan potassium group (gavage with losartan potassium 10.3mg/kg/d, with 10 rats in each group. After 2 weeks, Kidney pathology, serum Urotensin II, and cAMP concentration were detected, and the expressions of NLRP3, GSDMD-N, Caspase-1, and IL-1ß were detected by immunohistochemistry. Rat renal tubular epithelial cells were cultured in vitro, and different concentrations of Urotensin II were used to intervene for 24h and 48h. Cell proliferation activity was detected using the CCK8 assay. Suitable concentrations of Urotensin II and intervention time were selected, and Urotensin II receptor antagonist (SB-611812), inhibitor of PKA(H-89), and AS-IV (15ug/ml) were simultaneously administered. After 24 hours, cells and cell supernatants from each group were collected. The cAMP concentration was detected using the ELISA kit, and the expression of PKA, α-SMA, FN, IL-1ß, NLRP3, GSDMD-N, and Caspase-1 was detected using cell immunofluorescence, Western blotting, and RT-PCR. RESULTS: Renal tissue of UUO rats showed renal interstitial infiltration, tubule dilation and atrophy, renal interstitial collagen fiber hyperplasia, and serum Urotensin II and cAMP concentrations were significantly higher than those in the sham operation group (p <0.05). AS-IV and losartan potassium intervention could alleviate renal pathological changes, and decrease serum Urotensin II, cAMP concentration levels, and the expressions of NLRP3, GSDMD-N, Caspase-1, and IL-1ß in renal tissues (p <0.05). Urotensin II at a concentration of 10-8 mol/L could lead to the decrease of cell proliferation, (p<0.05). Compared with the normal group, the cAMP level and the PKA expression were significantly increased (p<0.05). After intervention with AS-IV and Urotensin II receptor antagonist, the cAMP level and the expression of PKA were remarkably decreased (p<0.05). Compared with the normal group, the expression of IL-1ß, NLRP3, GSDMD-N, and Caspase-1 in the Urotensin II group was increased (p<0.05), which decreased in the AS-IV and H-89 groups. CONCLUSION: AS-IV can alleviate renal fibrosis by inhibiting Urotensin II-induced pyroptosis of renal tubular epithelial cells by regulating the cAMP/PKA signaling pathway.

Astragaloside IV protects against autoimmune myasthenia gravis in rats via regulation of mitophagy and apoptosis.

Astragaloside IV (AS­IV) has various pharmacological effects, including antioxidant and immunoregulatory properties, which can improve myasthenia gravis (MG) symptoms. However, the potential mechanism underlying the effects of AS­IV on MG remains to be elucidated. The present study aimed to investigate whether AS­IV has a therapeutic effect on MG and its potential mechanism of action. By subcutaneously immunizing rats with R97­116 peptide, an experimental autoimmune (EA) MG rat model was established. AS­IV (40 or 80 mg/kg/day) treatment was then applied for 28 days after modeling. The results demonstrated that AS­IV significantly ameliorated the weight loss, Lennon score and pathological changes in the gastrocnemius muscle of EAMG rats compared with the model group. Additionally, the levels of acetylcholine receptor antibody (AChR­Ab) were significantly decreased, whereas mitochondrial function [ATPase and cytochrome c (Cyt­C) oxidase activities] and ultrastructure were improved in the AS­IV treated rats. Moreover, the mRNA and protein expression levels of phosphatase and tensin homolog­induced putative kinase 1, Parkin, LC3II and Bcl­2, key signaling molecules for mitophagy and apoptosis, were upregulated, whereas the mRNA and protein expression levels of p62, Cyt­C, Bax, caspase 3 and caspase 9 were downregulated following AS­IV intervention. In conclusion, AS­IV may protect against EAMG in a rat model by modulating mitophagy and apoptosis. These findings indicated the potential mechanism underlying the effects of AS­IV on MG and provided novel insights into treatment strategies for MG.

Astragaloside IV Mediates the PI3K/Akt/mTOR Pathway to Alleviate Injury and Modulate the Composition of Intestinal Flora in ApoE-/- Atherosclerosis Model Rats.

BACKGROUND: Atherosclerosis (AS) is a chronic inflammatory vascular disease with a complex pathogenesis. Astragaloside IV (AST IV), the primary active component of Astragalus, possesses anti-inflammatory, antioxidant, and immunomodulatory properties. This research aims to investigate the outcome of AST IV on AS and its potential molecular mechanism. METHODS: A high-fat diet (21% fat, 50% carbohydrate, 20% protein, 0.15% cholesterol, and 34% sucrose) was utilized to feed Apolipoprotein E deficient (ApoE-/-) SD rats for 8 weeks, followed by continuous intragastric administration of AST IV for 8 weeks. Biochemical detection was conducted for serum lipid levels and changes in vasoactive substances. After Masson staining, aortic root oil red O staining, and Hematoxylin Eosin (HE) staining, the efficacy of AST IV was verified using quantitative reverse transcription polymerase chain reaction (qRT-PCR). The mRNA expression levels of inflammatory factors and endothelial dysfunction-related biomarkers in rat aortic root tissues were appraised. The changes in the composition of intestinal flora in rats after AST IV treatment were appraised using Image J (Multi-point Tool). Western blot was used to evaluate phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway-related protein levels in rat aortic root tissues. RESULTS: AST IV administration alleviated the pathological symptoms of AS rats. AST IV administration reduced serum total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), endothelin-1 (ET-1) and angiotensin (Ang)-II (Ang-II) levels, and augmented serum high-density lipoprotein cholesterol (HDL-C) and nitric oxide (NO) levels. At the same time, AST IV administration inhibited the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1ß, vascular cell adhesion molecule-1 (VCAM-1), matrix metalloproteinase-2 (MMP-2), macrophage inflammatory protein-1 (MCP-1), and intercellular adhesion molecule-1 (ICAM-1) in the aortic root tissue of AS rats. In addition, the intestinal flora changed significantly after AST IV administration. The number of Bifidobacterium, Lactobacillus, and Bacteroides augmented significantly, and Enterobacter, Enterococcus, Fusobacterium, and Clostridium significantly decreased. Mechanistically, AST IV administration inhibited the phosphorylation of PI3K, Akt, and mTOR in AS rats. When combined with Dactolisib (BEZ235) (a PI3K/Akt/mTOR pathway inhibitor), AST IV could further inhibit phosphorylation and reduce inflammation. CONCLUSION: AST IV has a potential anti-AS effect, which can improve the pathological changes of the aorta in ApoE-/- rats fed with a high-fat diet, reduce the level of inflammatory factors, and modulate the composition of intestinal flora via the PI3K/Akt/mTOR pathway.

Saponin and Ribosome-Inactivating Protein Synergistically Trigger Lysosome-Dependent Apoptosis by Inhibiting Lysophagy: Potential to Become a New Antitumor Strategy.

The susceptibility of lysosomal membranes in tumor cells to cationic amphiphilic drugs (CADs) enables CADs to induce lysosomal membrane permeabilization (LMP) and trigger lysosome-dependent cell death (LDCD), suggesting a potential antitumor therapeutic approach. However, the existence of intrinsic lysosomal damage response mechanisms limits the display of the pharmacological activity of CADs. In this study, we report that low concentrations of QS-21, a saponin with cationic amphiphilicity extracted from Quillaja Saponaria tree, can induce LMP but has nontoxicity to tumor cells. QS-21 and MAP30, a type I ribosome-inactivating protein, synergistically induce apoptosis in tumor cells at low concentrations of both. Mechanistically, QS-21-induced LMP helps MAP30 escape from endosomes or lysosomes and subsequently enter the endoplasmic reticulum, where MAP30 downregulates the expression of autophagy-associated LC3 proteins, thereby inhibiting lysophagy. The inhibition of lysophagy results in the impaired clearance of damaged lysosomes, leading to the leakage of massive lysosomal contents such as cathepsins into the cytoplasm, ultimately triggering LDCD. In summary, our study showed that coadministration of QS-21 and MAP30 amplified the lysosomal disruption and can be a new synergistic LDCD-based antitumor therapy.

Paris saponin VII reverses resistance to PARP inhibitors by regulating ovarian cancer tumor angiogenesis and glycolysis through the RORα/ECM1/VEGFR2 signaling axis.

The emergence of Poly (ADP-ribose) polymerase inhibitors (PARPi) has marked the beginning of a precise targeted therapy era for ovarian cancer. However, an increasing number of patients are experiencing primary or acquired resistance to PARPi, severely limiting its clinical application. Deciphering the underlying mechanisms of PARPi resistance and discovering new therapeutic targets is an urgent and critical issue to address. In this study, we observed a close correlation between glycolysis, tumor angiogenesis, and PARPi resistance in ovarian cancer. Furthermore, we discovered that the natural compound Paris saponin VII (PS VII) partially reversed PARPi resistance in ovarian cancer and demonstrated synergistic therapeutic effects when combined with PARPi. Additionally, we found that PS VII potentially hindered glycolysis and angiogenesis in PARPi-resistant ovarian cancer cells by binding and stabilizing the expression of RORα, thus further inhibiting ECM1 and interfering with the VEGFR2/FAK/AKT/GSK3ß signaling pathway. Our research provides new targeted treatment for clinical ovarian cancer therapy and brings new hope to patients with PARPi-resistant ovarian cancer, effectively expanding the application of PARPi in clinical treatment.

Formosanin C inhibits pulmonary metastasis by targeting stearyl CoA desaturase-1.

BACKGROUND: Cisplatin (DDP) as the first-line drug has been used in cancer therapy. However, side effects and drug resistance are the challenges of DDP. Disordered lipid metabolism is related to DDP resistance. STUDY DESIGN: In this study, formosanin C (FC) as the main compound of Rhizoma Paridis saponins (RPS) inhibits pulmonary metastasis by targeting stearyl CoA desaturase-1. METHODS AND RESULTS: RPS prolonged the survival period of mice, reduced pulmonary metastases and alleviated colon toxicity caused by DDP. FC as the main compound of RPS enhanced the anti-tumor and anti-metastatic effects of DDP. FC decreased the mRNA level of SCD1 and the content of lipid droplets (LDs) in lung cancer cells. Molecular dynamics and isothermal titration calorimetry verified the binding stability and spontaneously between FC and SCD1. SiSCD1 reduced the content of LDs in cell lines and increased mitochondria (mtROS), which was consistent with the results of FC treatment. The combination group decreased DNA repair associated protein as well as DDP resistance markers such as ERCC1 and 53bp1, and increased DNA damage marker like γH2AX, which indirectly confirmed the occurrence of mtROS. In addition, FC combination with DDP also affected epithelial-mesenchymal transition-related protein like VIM and CDH1 in vivo experiments, and thereby inhibited pulmonary metastasis. CONCLUSION: Our research indicated that the FC as the main compound of RPS targeted the CY2 domain of SCD1, inhibited lipid metabolism in mice, and thereby suppressed cancer metastases. This provided support for use of FC to treat cancer based on lipid metabolism pathway.