Quercetin inhibits mitophagy-mediated apoptosis and inflammatory response by targeting the PPARγ/PGC-1α/NF-κB axis to improve acute liver failure.

Reactive oxygen species (ROS) from mitochondrial dysfunction are critical in triggering apoptosis and inflammation in acute liver failure (ALF). Quercetin (QUE), an antioxidant, is renowned for its therapeutic effects onliverdiseases. There are no studies on whether QUE regulates mitophagy level in hepatocytes to inhibit ALF. This study investigates QUE's protective effects on ALF and elucidates the mechanisms involved. The ALF and hepatocyte inflammatory injury model was established using LPS and D-Galn. To predict potential targets and mechanisms of QUE in ALF treatment, transcriptomics, network pharmacology, molecular docking techniques, and ChIP were employed. The expression level related to mitophagy, apoptosis, and signaling pathways were detected by CCK8, IHC, IF staining, TUNEL, RT-qPCR, TEM, Western blotting, ELISA, and flow cytometry. Network pharmacology and transcriptomics revealed common targets between QUE and ALF. Enrichment analysis showed that the anti-ALF targets of QUE were significantly associated with mitochondria and NF-κB-related pathways. Subsequent experiments showed that QUE pretreatment significantly alleviated the loss of hepatocyte viability, enhanced mitochondrial membrane potential, activated mitophagy, and promoted the clearance of damaged mitochondria, thereby reducing ROS accumulation, significantly reducing cell apoptosis and inflammatory responses, reducing ALT and AST levels, and improving liver tissue pathology. Mechanistically, molecular docking, DARTS, and CETSA analyses confirmed that QUE directly binds to the PPARγ molecule, which reduced binding to IκB and significantly inhibit the NF-κB pathway to exert its protective effects. In short, our results provide the first evidence that QUE improves acute liver failure by promoting mitophagy through regulating the PPARγ/PGC-1α/NF-κB axis and inhibiting apoptosis and inflammatory responses mediated by mitochondrial dysfunction, which provides evidence for the potential of QUE in the treatment of ALF.

Wu H ，Wu L ，Luo L ，Wu YT ，Zhang QX ，Li HY ，Zhang BF ... -  《-》

Kaixinsan regulates neuronal mitochondrial homeostasis to improve the cognitive function of Alzheimer's disease by activating CaMKKβ-AMPK-PGC-1α signaling axis.

Alzheimer's disease (AD) is a neurodegenerative disease primarily characterized by cognitive impairments. With the intensification of population aging, AD has become a major health concern affecting the elderly. Kaixinsan, a classical traditional Chinese formula, consists of Ginseng Panax et Rhizoma, Polygalae Radix, Poria and Acori Tatarinowii Rhizoma, and is commonly used in clinical for treating memory decline. However, its mechanism remains unclear, which hinders its popularization and application. Morris water maze (MWM) was performed to evaluate the effect of Kaixinsan on improving learning and memory ability in SAMP8 (senescence-accelerated mouse prone 8, an AD model mice) mice. Nissl staining, TdT-mediated dUTP Nick End Labeling (TUNEL) and western blotting (Bax and Bcl-2) were used to confirm the effect of Kaixinsan on the neuronal structure and apoptosis of SAMP8 mice. Ultra performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC-Q-TOF/MS) was performed to identify the distribution components in brain tissue after administration of Kaixinsan extraction. Based on the identified brain distribution components, the mechanism of Kaixinsan improving the cognitive function was predicted by network pharmacology. Then, using HSP60 as a mitochondrial marker and RBFOX3 as a neuronal marker, immunofluorescence co-localization was used to confirm the effect of Kaixinsan on neuronal mitochondria quantity in SAMP8 mice. Western blotting was employed to access the expression of predicted proteins (AMPK, CaMKKβ, PGC-1α and HSP90) implicated in mitochondrial homeostasis. To further confirm the mechanism of Kaixinsan, SH-SY5Y cell injury model induced by amyloid β - protein fragment 25-35 (Aβ25-35) was replicated and the effect of Kaixinsan - containing serum on apoptosis in injured SH-SY5Y cells was investigated by flow cytometer. The expression level of apoptosis-associated proteins (Bax and Bcl-2) and mitochondrial homeostasis related proteins (AMPK, CaMKKβ, PGC-1α and HSP90) in the presence or absence of CaMKKβ inhibitor (STO-609) were compared. The results indicate that Kaixinsan can improve the cognitive function of SAMP8 mice, alleviate the hippocampal tissue lesions and inhibit neuron apoptosis. Seventeen brain distribution components of Kaixinsan were identified. Based on the brain distribution components of Kaixinsan, the results of network pharmacology suggest that Kaixinsan may regulate mitochondrial homeostasis through the CaMKKβ-AMPK-PGC-1α signaling axis. In vivo experiments indicated that Kaixinsan could reverse neuronal mitochondrial loss in SAMP8 mice by upregulating CaMKKβ, AMPK, HSP90 and PGC-1α to promote mitochondrial biogenesis and increase the number of neuronal mitochondria. Additionally, the in vitro experiments demonstrated that Kaixinsan can inhibit apoptosis of Aβ25-35 injured SH-SY5Y cells and upregulate mitochondrial homeostasis-related proteins CaMKKβ, AMPK and PGC-1α. However, in addition to CaMKKβ inhibitors, the neuroprotective effect disappeared. The results indicate that Kaixinsan can improve the cognitive function of SAMP8 mice by regulating CaMKKβ-AMPK-PGC-1α signaling axis to maintain mitochondrial homeostasis and inhibit neuronal apoptosis.

Ren J ，Xiang B ，Song L ，René DJ ，Luo Y ，Wen G ，Gu H ，Yang Z ，Zhang Y ... -  《-》

Preventive effects of matrine on LPS-induced inflammation in RAW 264.7 cells and intestinal damage in mice through the TLR4/NF-κB/MAPK pathway.

Matrine is a tetracyclic quinolizidine alkaloid with diverse bioactive properties, including anti-inflammatory and neuroprotective properties. However, the underlying anti-inflammatory mechanisms remain unclear. This study aimed to explore how matrine reduces Lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 cells and to assess its protective effects against LPS-induced intestinal damage. The effect of matrine on cell viability was assessed using the cell counting kit-8 (CCK-8) assay. Additionally, its impact on inflammatory cytokines and macrophage polarization was assessed using enzyme-linked immunosorbent assay (ELISA), flow cytometry, and quantitative real-time polymerase chain reaction (qRT-PCR) analyses. The effects on intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), nitric oxide (NO) production, and oxidative stress were evaluated using 2',7'-dichlorodihydrofluorescein diacetate staining and JC-1 and Griess assays. Immunofluorescence staining was used to observe the translocation of the NF-κB p65 subunit. Western blotting (WB) and qRT-PCR were employed to analyze the expression levels of proteins related to the toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB)/mitogen-activated protein kinase (MAPK) pathway. An LPS-induced mouse model was established to study the intestinal inflammation and barrier injury. Mouse feces characteristics, colon length, and disease activity index (DAI) were recorded. Hematoxylin-eosin (H&E) and alcian blue/periodic acid schiff (AB/PAS) staining were used to observe morphological changes and barrier damage in the duodenum, jejunum, ileum, and colon and to measure villus length, crypt depth, goblet cell count, and positive areas in the duodenum, jejunum, and ileum. The content of short-chain fatty acids (SCFAs) in the colon was determined using gas chromatography (GC). Matrine inhibited LPS-induced inflammatory cytokine levels, suppressed macrophage M1 polarization, and promoted M2 macrophage polarization. Matrine reduced LPS-induced increases in ROS and NO levels and regulates oxidative stress. Additionally, matrine inhibited the nuclear translocation of the NF-κB p65 subunit and exerted anti-inflammatory effects by suppressing the activation of the TLR4/NF-κB/MAPK pathway. In vivo experiments indicated that matrine significantly alleviated LPS-induced diarrhea, increased DAI, and shortened the colon. Matrine reduced the production of the pro-inflammatory cytokine interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α and the pro-inflammatory mediator NO in mouse intestinal tissues while promoting the content of the anti-inflammatory cytokine IL-10. Furthermore, it improved intestinal tissue structure and alleviated LPS-induced intestinal barrier damage. Finally, matrine increased the SCFA levels in the intestine. Matrine exerted its anti-inflammatory effects and protects against intestinal injury through the TLR4/NF-κB/MAPK signaling pathway.

Mao N ，Yu Y ，Lu X ，Yang Y ，Liu Z ，Wang D ... -  《-》

Based on network pharmacology, molecular docking and experimental verification to reveal the mechanism of Andrographis paniculata against solar dermatitis.

Solar dermatitis (SD) is an acute, damaging inflammation of the skin caused by UV exposure, especially UVB. Therefore, the discovery of novel anti-SD therapeutic agents is crucial. Andrographis paniculata (AP) is a medicinal plant with a wide range of pharmacological effects. Increased evidence shows that AP has potential therapeutic effects on SD. However, the therapeutic mechanisms of AP against SD have not yet been completely elucidated, which is an unexplored field. This study employed network pharmacology, molecular docking and experimental verification to ascertain the active constituents, possible targets, and biological pathways associated with AP in the treatment of SD. AP-related active ingredients and their potential targets were screened from TCMSP and Swiss Target Prediction database, respectively. Potential therapeutic targets of SD were collected using the GeneCards, DrugBank and OMIM databases. Then, we established protein-protein interaction (PPI), compound-target-disease (D-C-T-D) through Cytoscape to identify the major components, core targets of AP against SD. Next, the GO and KEGG pathway was identified by the David database of AP in the treatment of SD. Molecular docking techniques were used to estimate the binding force between the components and the hub genes. In this paper, we used UVB-irradiated HaCaT keratinocytes as an in vitro model and established the dorsal skin of UVB-irradiated ICR mice as an in vivo model to explore the mechanism for further verification. There were 24 active components and 63 related target genes in AP against SD. PPI analysis showed that AKT-1, TNF-α, IL6, MMP9, EGFR, and PTGS2 shared the highest centrality among all target genes. KEGG pathway analysis revealed that the PI3K-Akt signaling pathway may be central in the anti-SD system. The molecular docking results showed that the main active components of AP have strong binding affinity with hub genes. In vitro results showed that WG had a protective effect on UVB-intervened HaCat cells. Western blot analysis showed that WG intervention achieved anti-inflammation by reducing the phosphorylated expression of AKT, PI3K proteins in the PI3K-AKT signaling pathway and downregulating the expression of TNF-α, IL-6, EGFR. Furthermore, Histological analysis confirmed that administration of WG to ICR mice significantly ameliorated UVB-induced skin roughness, epidermal thickening, disturbed collagen fiber alignment and wrinkles. Meanwhile, immunohistochemistry showed that administration of WG to ICR mice significantly reduced UVB-induced expression of MMP9, MPO, F4/80 in the skin. These results provide new insights into the contribution of WG to the development of clinical treatment modalities for UVB-induced SD. The crucial element in the fight against SD is WG, with the primary route being PI3K/Akt. The main components and hub genes had robust binding abilities. In vitro and vivo experiments showed that WG could inhibit the expression level of the hub genes by inhibiting the PI3K/Akt pathway. In summary, the information presented in this study indicates that WG might be utilised as a treatment for UVB-induced SD.

Deng Q ，Chen W ，Deng B ，Chen W ，Chen L ，Fan G ，Wu J ，Gao Y ，Chen X ... -  《-》

Dioscin improves fatty liver hemorrhagic syndrome by promoting ERα-AMPK mediated mitophagy in laying hens.

Mitochondria play a crucial role in upholding metabolic homeostasis. Mitochondrial damage closely associated with the pathogenesis of fatty liver hemorrhagic syndrome (FLHS), while mitophagy being among the most effective methods for eliminating the damaged mitochondria. Dioscin, a natural extract, can activate autophagy; however, its effects on FLHS regarding mitophagy regulation remain unelucidated. We explored the impact of dioscin on FLHS induced by a high-energy and low-protein (HELP) diet in laying hens, mainly focused the protective effects of dioscin on mitochondrial injury. To investigate the impact of dioscin on fatty liver syndrome in laying hens, we first induced the condition by feeding them a high-energy and low-protein diet. Then, we assessed lipid metabolism-related markers using oil red staining and a commercial detection kit. In addition, the role of dioscin on fatty liver syndrome in laying hens was confirmed by assessing the activation of hepatocyte fat deposition and hepatocyte apoptosis; and the mechanism of dioscin in FLHS was investigated through LMH cell experiment in vitro. Furthermore, CETSA and molecular docking were conducted for additional confirmation. The results showed that dioscin alleviated mitochondrial damage, relieved the excessive deposition of hepatic lipid droplets and oxidative stress induced by HELP diet in laying hens. Furthermore, dioscin regulated the mitophagy by activating the estrogen receptor α (ERα)/adenosine 5'-monophosphate-activated protein kinase (AMPK) signaling pathway, thus mitigating mitochondria injury and apoptosis in hepatocytes. In addition, we found that dioscin promoted the translocation of nuclear transcription factor into nucleus by activating ERα-AMPK signaling, facilitating autophagic flux in the liver of laying hens and LMH cells. Furthermore, cells pretreated with the lysosomal acidification inhibitor bafilomycin A1 blocked the inhibitory effect of dioscin on the apoptosis induced by palmitic acid (PA)-stimulation in LMH cells, suggesting that dioscin reduces PA-induced apoptosis by activating mitophagy. Moreover, dioscin-induced lysosomal acidification and mitochondrial biogenesis were reversed in PA-induced LMH cells pretreated with ERα-specific inhibitor methylpiperidino pyrazole. This study firstly demonstrated that dioscin alleviates fatty liver syndrome induced by HELP diet in laying hens. The findings from this study illustrated that dioscin plays a significant role in reducing mitochondrial damage and apoptosis, and these beneficial effects mainly achieve through promotion of ERα-AMPK signaling, which mediates autophagy within the liver of laying hens fed a HELP-diets. These findings provide a theoretical basis for considering dioscin as a possible treatment option for mitigating FLHS in egg-laying hens.

Xing Y ，Huang B ，Cui Z ，Zhang Q ，Ma H ... -  《-》