Blockade of KLF5/LDH-A feedback loop contributes to Curcumol inhibition of sinusoidal endothelial cell glycolysis and mitigation of liver fibrosis.

BACKGROUND: LSECs (Liver sinusoidal endothelial cells) are the portal of liver, their pathological angiogenesis plays a constructive role in etiopathogenesis of liver fibrosis by affecting liver tissue repair and inflammatory drive. Although intervention in angiogenesis can effectively inhibit abnormal activation of LSEC, no effective drugs have been found to treat liver fibrosis. PURPOSE: We investigated the effect of the natural compound Curcumol on LSEC angiogenesis and elucidated the novel underlying mechanism, expecting to provide a scientific basis for exploring potential therapeutic drugs for liver fibrosis. METHODS: Various cellular and molecular assays, as well as genetic assays, were used to detect pathological angiogenesis and changes in glycolysis levels in cultured rat LSECs and mouse liver fibrosis models. RESULTS: Transcription factor KLF5 is able to influence the angiogenic properties of LSEC by regulating the glycolytic process, and affect the expression of LDH-A by transcriptionally binding to its promoter. In our study, we were surprised to find that LDH-A (the final step of glycolysis) has a strong regulatory effect on the glycolytic process of LSEC. Through in-depth study, we found that LDH-A could affect the transcriptional activity of KLF5, thus forming a positive feedback loop. Curcumol could break this positive feedback loop and inhibit the glycolysis-dependent angiogenic nature of LSEC, thus alleviating liver fibrosis. Curcumol reduced extracellular matrix (ECM) deposition, attenuated pathological angiogenesis in LSEC, and decreased the level of CCl4-induced liver fibrosis in mice. CONCLUSION: Our results demonstrated the great utilization potentiality of KLF5 in liver fibrosis, and the innovative discovery that LDH-A regulates the glycolytic process and forms a malignant feedback loop by exerting non-enzymatic effects. It also reveals the prospect of Curcumol-regulated KLF5/LDH-A feedback loop in the treatment of liver fibrosis, providing a new option for the future medicine of liver fibrosis.

Qingchang Wenzhong Decoction reduce ulcerative colitis in mice by inhibiting Th17 lymphocyte differentiation.

BACKGROUND: Qingchang Wenzhong Decoction (QCWZD), a chinese herbal prescription, is widely used for ulcerative colitis (UC). Nevertheless, the active ingredients and mechanism of QCWZD in UC have not yet been explained clearly. PURPOSE: This research focuses on the identification of the effective ingredients of QCWZD and the prediction and verification of their potential targets. METHODS: The UC mice were established by adding 3.0% dextran sulfate sodium (DSS) to sterile water for one week. Concurrently, mice in the treatment group were gavage QCWZD or mesalazine. LC-MS analyzed the main components absorbed after QCWZD treatment, and network pharmacology predicted their possible targets. ELISA, qPCR, immunohistochemistry and immunofluorescence experiments were used to evaluate the colonic inflammation level and the intestinal barrier completeness. The percentage of Th17 and Treg lymphocytes was detected by flow cytometry. RESULTS: After QCWZD treatment, twenty-seven compounds were identified from the serum. In addition, QCWZD treatment significantly reduced the increased myeloperoxidase (MPO) and inflammatory cell infiltration caused by DSS in the colonic. In addition, QCWZD can reduce the secretion of inflammatory factors in serum and promote the expression of mRNAs and proteins of occludin and ZO-1. Network pharmacology analysis indicated that inhibiting IL-6-STAT3 pathway may be necessary for QCWZD to treat UC. Flow cytometry analysis showed that QCWZD can restore the normal proportion of Th17 lymphocytes in UC mice. Mechanistically, QCWZD inhibited the phosphorylation of JAK2-STAT3 pathway, reducing the transcriptional activation of RORγT and IL-17A. CONCLUSIONS: Overall, for the first time, our work revealed the components of QCWZD absorbed into blood, indicated that the effective ingredients of QCWZD may inhibit IL-6-STAT3 pathway and inhibit the differentiation of Th17 lymphocytes to reduce colon inflammation.

Modification of lysine deacetylation regulates curcumol-induced necroptosis through autophagy in hepatic stellate cells.

The excessive deposition of extracellular matrix (ECM) is the main characteristic of liver fibrosis, and hepatic stellate cells (HSCs) are the main source of ECM. The removal of activated HSCs has a reversal effect on liver fibrosis. Western blot and MTT analysis indicated that curcumol could relieve hepatic fibrosis by promoting HSCs receptor-interacting protein kinase 1/3 (RIP1/RIP3)-dependent necroptosis. Importantly, autophagy flow was monitored by constructing the mRFP-GFP-LC3 plasmid, and it was found that curcumol cleared activated HSCs in a necroptosis manner that was dependent on autophagy. Our study suggested that the activation of necrosome formed by RIP1 and RIP3 depended on Atg5, and that autophagosomes were also necessary for curcumol-induced necroptosis. Furthermore, microscale thermophoresis and co-immunoprecipitation assay results proved that curcumol could target Sirt1 to regulate autophagy by reducing the acetylation level of Atg5. The HSCs-specific silencing of Sirt1 exacerbated CCl4 -induced liver fibrosis in mice. The deacetylation of Atg5 not only accelerated the accumulation of autophagosomes but also enhanced the interaction between Atg5 and RIP1/RIP3 to induce necroptosis. Overall, our study indicated that curcumol could activate Sirt1 to promote Atg5 deacetylation and enhanced its protein-protein interaction function, thereby inducing autophagy and promoting the necroptosis of HSCs to reduce liver fibrosis.

Investigation of the key targets and pharmacological mechanisms of rhamnocitrin against oxaliplatin-induced neuropathic pain based on network pharmacology approach and experimental validation.

Rhamnocitrin (RH) is a bioactive flavonoid of Astragali Radix, which exerts a wide variety of pharmacological effects. However, there are no reports focusing on the therapeutical effects and mechanisms of RH against neuropathic pain (NP). In this study, systematic pharmacology and in vivo experimental approaches were employed to identify the potential targets of RH for treating oxaliplatin-induced NP. Our findings indicated that the therapeutical effect of RH might be closely associated with key genes, including MAPK3, MAPK1, SRC, PTGS2, EGFR, MMP9, and MMP2, as well as potential signaling pathways such as PI3K-Akt signaling pathway, EGFR tyrosine kinase inhibitor resistance, and Rap1 signaling pathway. The in vivo experimental findings demonstrated that RH could suppress oxidative stress, inflammatory response, and down-regulate MMP2 and MMP9 expressions to exert its therapeutic effects against NP. This study used network pharmacology and experimental validation to elucidate the potential targets and underlying mechanisms by which RH improves oxaliplatin-induced NP and offer new insight on drug development for NP.

First report of potato rot nematode, Ditylenchus destructor Thorne, 1945 infecting Codonopsis pilosula in Gansu province, China.

In November 2019, stem nematode was found on Codonopsis pilosula in Tanchang county, Gansu province, China. The population of stem nematode was identified on the basis of both molecular and morphological methods. The morphological and morphometric characteristics of this nematode population matched with Ditylenchus destructor Thorne, 1945. The sequences of rDNA-ITS and D2/D3 region of 28S-rRNA similarity with the D. destructor. The pathogenicity results revealed the symptom of dry rot on C. pilosula was caused by this nematode. To our knowledge, this is the first report that D. destructor on C. pilosula in China.

Identification of the absorbed components and metabolites of Xiao-Ai-Jie-Du decoction and their distribution in rats using ultra high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry.

Xiao-Ai-Jie-Du decoction (XAJDD), a traditional Chinese medicine formula, has long been used for the treatment of hepatocarcinoma, gastric cancer and colorectal cancer. It is composed of six herbal medicines, including Scutellariae Barbatae Herba, Pseudostellariae Radix, Ophiopogonis Radix, Cremastrae Pseudobulbus, Curcumae Rhizoma and Akebiae Fructus. Despite the in-depth study on its pharmacological effects on cancer prevention and treatment, the comprehensive analysis of the chemical components and the absorbed bioactive constituents are not well studied. Thus, an ultra-high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) method was established to detect and identify the chemical constituents in XAJDD. The absorbed components and metabolites after oral administration of XAJDD in rats were also studied. In total, 102 components were identified or tentatively characterized in XAJDD, including 30 flavonoids, 19 triterpenoids, 12 organic acids, 9 steroidal saponins, 9 cyclic peptides, 7 phenanthrenes, 5 amino acids, 3 alkaloids and 8 other compounds. After analysing the metabolites in rat plasma and urine after oral administration of XAJDD, a total of 70 compounds were identified, including 15 primary components and 55 metabolites, and metabolic pathways, including hydrogenation, hydroxylation, methylation, sulfonation, and glucuronidation were evaluated. Among these, methylation and glucuronidation were the main metabolic pathways. In conclusion, the developed UHPLC-Q-TOF-MS method with high sensitivity and resolution is suitable for identifying and characterizing the chemical constituents of XAJDD in vitro and characterizing the primary components and their metabolites in vivo; moreover, the results will provide essential data for further studying the relationship between the chemical components and pharmacological activity of XAJDD.

Wogonin reverses multi-drug resistance of human myelogenous leukemia K562/A02 cells via downregulation of MRP1 expression by inhibiting Nrf2/ARE signaling pathway.

Constitutive NF-E2-related factor 2 (Nrf2) activation has been recently reported to play a pivotal role in enhancing cell survival and resistance to anticancer drugs in many tumors. Previously, much effort has been devoted to the investigation of blocking Nrf2 function in cultured cells and cancer tissues, but few researches have been undertaken to evaluate the precise mechanism of flavonoids-induced sensitivity by inhibiting Nrf2. In this study, we investigated the reversal effect of Wogonin, a flavonoid isolated from the root of Scutellaria baicalensis Georgi, in resistant human myelogenous leukemia. Data indicated that Wogonin had strong reversal potency by inhibiting functional activity and expression of MRP1 at both protein and mRNA in adriamycin (ADR)-induced resistant human myelogenous leukemia K562/A02 cells. Consequently, the inhibition of MRP1 by Wogonin was dependent on Nrf2 through the decreased binding ability of Nrf2 to antioxidant response element (ARE). Further research revealed Wogonin modulated Nrf2 through the reduction of Nrf2mRNA at transcriptional processes rather than RNA degradation, which is regulated by the PI3K/Akt pathway. Moreover, DNA-PKcs was found to be involved in the Wogonin-induced downregulation of Nrf2 mRNA at transcriptional levels. In summary, these results clearly demonstrated the effectiveness of using Wogonin via inhibiting Nrf2 to combat chemoresistance and suggested that Wogonin can be developed into an efficient natural sensitizer for resistant human myelogenous leukemia.