ABSTRACT
PANoptosis is manifested with simultaneous activation of biomarkers for both pyroptotic, apoptotic and necroptotic signaling via the molecular platform PANoptosome and it is involved in pathologies of various inflammatory diseases including hemophagocytic lymphohistiocytosis (HLH). Scutellarin is a flavonoid isolated from herbal Erigeron breviscapus (Vant.) Hand.-Mazz. and has been shown to possess multiple pharmacological effects, but it is unknown whether scutellarin has any effects on PANoptosis and related inflammatory diseases. In this study, we found that scutellarin inhibited cell death in bone marrow-derived macrophages (BMDMs) and J774A.1 cells treated with TGF-ß-activated kinase 1 (TAK1) inhibitor 5Z-7-oxozeaenol (OXO) plus lipopolysaccharide (LPS), which has been commonly used to induce PANoptosis. Western blotting showed that scutellarin dose-dependently inhibited the activation biomarkers for pyroptotic (Caspase-1p10 and GSDMD-NT), apoptotic (cleaved Casp3/8/9 and GSDME-NT), and necroptotic (phosphorylated MLKL) signaling. The inhibitory effect of scutellarin was unaffected by NLRP3 or Caspase-1 deletion. Interestingly, scutellarin blocked the assembly of PANoptosome that encompasses ASC, RIPK3, Caspase-8 and ZBP1, suggesting its action on upstream signaling. Consistent with this, scutellarin inhibited mitochondrial damage and mitochondrial reactive oxygen species (mtROS) generation in cells treated with OXO+LPS. Further, mito-TEMPO that can scavenge mtROS significantly inhibited OXO+LPS-induced PANoptotic cell death. In line with the in vitro results, scutellarin markedly alleviated systemic inflammation, multiple organ injury, and activation of PANoptotic biomarkers in mice with HLH. Collectively, our data suggest that scutellarin can inhibit PANoptosis by suppressing mitochondrial damage and mtROS generation and thereby mitigating multiple organ injury in mice with inflammatory disorders.
ABSTRACT
OBJECTIVE: PANoptosis, a new form of regulated cell death, concomitantly manifests hallmarks for pyroptosis, apoptosis, and necroptosis. It has been usually observed in macrophages, a class of widely distributed innate immune cells in various tissues, upon pathogenic infections. The second-generation curaxin, CBL0137, can trigger necroptosis and apoptosis in cancer-associated fibroblasts. This study aimed to explore whether CBL0137 induces PANoptosis in macrophages in vitro and in mouse tissues in vivo. METHODS: Bone marrow-derived macrophages and J774A.1 cells were treated with CBL0137 or its combination with LPS for indicated time periods. Cell death was assayed by propidium iodide staining and immunoblotting. Immunofluorescence microscopy was used to detect cellular protein distribution. Mice were administered with CBL0137 plus LPS and their serum and tissues were collected for biochemical and histopathological analyses, respectively. RESULTS: The results showed that CBL0137 alone or in combination with LPS induced time- and dose-dependent cell death in macrophages, which was inhibited by a combination of multiple forms of cell death inhibitors but not each alone. This cell death was independent of NLRP3 expression. CBL0137 or CBL0137 + LPS-induced cell death was characterized by simultaneously increased hallmarks for pyroptosis, apoptosis and necroptosis, indicating that this is PANoptosis. Induction of PANoptosis was associated with Z-DNA formation in the nucleus and likely assembly of PANoptosome. ZBP1 was critical in mediating CBL0137 + LPS-induced cell death likely by sensing Z-DNA. Moreover, intraperitoneal administration of CBL0137 plus LPS induced systemic inflammatory responses and caused multi-organ (including the liver, kidney and lung) injury in mice due to induction of PANoptosis in these organs. CONCLUSIONS: CBL0137 alone or plus inflammatory stimulation induces PANoptosis both in vitro and in vivo, which is associated with systemic inflammatory responses in mice.
Subject(s)
Carbazoles , DNA, Z-Form , Neoplasms , Mice , Animals , Lipopolysaccharides/pharmacology , Apoptosis , PyroptosisABSTRACT
Aim: To observe the effect of uc.48 + small interference RNA (siRNA) on liver glycogen abnormality in type 2 diabetic rats and its possible mechanism. Methods: The diabetes model was established by feeding high glucose and high fat diet combined with streptozotocin(STZ). After the success of the model, the long noncoding RNA uc. 48 + siRNA was injected into the rat body via tail vein. The changes of blood glucose and the content of liver glycogen were detected dynamically, and the liver glycogen was detected one week after injection. Glucokinase (GK) mRNA and protein expression in liver tissues of each group were detected by qPCR and Western blot. Results: It was observed that postprandial blood glucose and fasting blood glucose decreased in diabetic model rats after treated with uc. 48 + siRNA compared with those in model rats. The level of liver glycogen in diabetic model rats was significantly lower than that in control group. The synthesis of liver glycogen in diabetic model rats with uc. 48 + siRNA treatment increased compared with that in diabetic model group. The expressions of GK mRNA and protein in the diabetic model group were significantly lower than those in control group. The expression of GK mRNA and protein markedly increased after uc. 48 + siRNA treatment. Conclusions: uc. 48 + siRNA reduces blood glucose and increases glycogen synthesis in type 2 diabetic rats, and its mechanism may involve in increasing GK expression and Aktl phosphorylation.
ABSTRACT
A rapid and sensitive liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI/ MS) method for quantification of pentoxyverine citrate in human plasma has been developed and applied for the bioequivalence and pharmacokinetics study. After extracted from plasma samples with ethyl acetate, analysis was performed in selected ion monitoring (SIM) mode with a positive electrospray ionization (ESI) interface with a mobile phase consisted of methanol and water (0.4% glacial acetic acid and 4 mmol x L(-1) ammonium acetate, 43 : 57, v/v). The linear concentration range of the calibration curves was 1.0-160.0 ng x mL(-1) for pentoxyverine citrate, inter- and intra-precision (RSD) was less than 12.5%, accuracy (RE) was in +/- 13.5% and absolute recovery was more than 80%. The method was proved simple, rapid, sensitive, specific and suitable for pharmacokinetic and bioequivalence study of Yufenweilin capsule containing pentoxyverine citrate.
