OBJECTIVES: To propose a hypothesis that aloe-emodin may inhibit scar tissue fibrosis through thrombospondin-1(THBS1)-PI3K-Akt pathway. METHODS: By cultivating fibroblasts derived from scar tissue after cleft palate surgery in humans, aloe emodin of different concentrations (10, 20, 30, 40 and 50 μmol/L) was added to the cells which activity was detected. At the same time, transcriptome sequencing was performed on scar tissue and cells, and bioinformatics methods were used to explore potential targets and signaling pathways of scar tissue fibrosis. RESULTS: Aloe-emodin had a concentration dependent inhibitory effect on fibroblast proliferation,with the 40 μmol/L concentration group showing the most significant effect. The results of tissue and cell sequencing indicated that differentially expressed genes were significantly enriched in extracellular matrix-receptor interaction pathway, and shared a common differential gene which was THBS1. The ORA analysis results indicated that differentially expressed genes, including THBS1, were significantly enriched in the PI3K-Akt signaling pathway. CONCLUSIONS Aloe emodin may inhibit the PI3K-Akt pathway by downregulating THBS1, thereby reducing the proliferation activity of fibroblasts derived from postoperative palatal scar tissue.
Thrombospondin 1/genetics* ; Humans ; Signal Transduction/drug effects* ; Fibroblasts/cytology* ; Proto-Oncogene Proteins c-akt/metabolism* ; Fibrosis ; Phosphatidylinositol 3-Kinases/metabolism* ; Cicatrix/metabolism* ; Cell Proliferation/drug effects* ; Anthraquinones/pharmacology* ; Cells, Cultured

Objective To investigate the effects and molecular mechanism of Homer protein homolog 1a (Homer 1a) overexpression on nerve injury in mice with traumatic brain injury (TBI). Methods Sixty male C57BL/6 mice were randomly divided into five groups: sham group, TBI group, empty lentivirus (Lv-NC) group, Homer 1a overexpression lentivirus (Lv-Homer 1a) group and Lv-Homer 1a + 740 Y-P group, with 12 mice in each group. The lentivirus was orthotopic injected into the cerebral cortex of mice 5 d before modeling, while 740 Y-P was injected intraperitoneally 1 d before modeling. The TBI model was established using the free-fall impact method, and the modified neurological severity scores (mNSS) of the mice was assessed 72 h post-surgery. The water content of brain tissue was quantified, and the histopathological damage and neuronal loss in brain tissue were assessed using HE staining and Nissl staining respectively. The formation of autophagosomes in brain tissue was observed by transmission electron microscopy. The protein expression levels of Homer 1a, microtubule-associated protein 1 light chain 3B (LC3B), Beclin 1, phosphatidylinositol 3-kinase (PI3K), phosphorylation PI3K(p-PI3K), protein kinase B (AKT), p-AKT, mammalian target of rapamycin (mTOR), and p-mTOR in brain tissue were detected by Western blot analysis. Results Compared to the sham group, the mice in the TBI group exhibited a significant increase in mNSS and cerebral water content. Moreover, severe brain tissue pathological damage was observed, accompanied by a substantial loss of neurons and an increase in autophagosome formation. The protein expressions of Homer 1a and Beclin 1, as well as the protein ratio of LC3B-II/LC3B-I, in brain tissues were significantly elevated, while the protein ratios of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR were significantly reduced. Compared to the TBI group, the Lv-Homer 1a group exhibited reduced mNSS and brain water content. Additionally, there was an improvement in pathological brain tissue damage and neuron loss. Furthermore, there was an increase in autophagosome formation and expression of autophagy-related proteins, while the protein ratios of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR were decreased. Compared to the Lv-Homer 1a group, the nerve injury in the Lv-Homer 1a+740 Y-P group was exacerbated, accompanied by a reduction in autophagosome formation and expression of autophagy-related proteins, while the PI3K/AKT/mTOR signaling pathway was activated. Conclusion Overexpression of Homer 1a effectively mitigates neurological damage in TBI mice, potentially through modulation of autophagy mediated by the PI3K/AKT/mTOR signaling pathway.
Animals ; TOR Serine-Threonine Kinases/genetics* ; Autophagy ; Brain Injuries, Traumatic/pathology* ; Male ; Proto-Oncogene Proteins c-akt/genetics* ; Phosphatidylinositol 3-Kinases/genetics* ; Homer Scaffolding Proteins/metabolism* ; Mice, Inbred C57BL ; Signal Transduction ; Mice

Objective To investigate the effect of basic helix-loop-helix family member E40 (BHLHE40) on the invasion and migration of osteosarcoma (OS) cells, and to explore the role of the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) signaling pathway in the biological behavior of OS mediated by BHLHE40, providing a scientific basis for targeted therapy of OS. Methods On the basis of clinical OS samples and OS cell lines, the expression differences of BHLHE40 between OS and adjacent tissues, as well as those between OS cells and normal osteoblast cell lines, were analyzed. BHLHE40 knockdown OS cells were obtained through shRNA transfection. The effects of BHLHE40 on OS cell proliferation, migration, and invasion were examined using CCK-8, EdU staining, wound healing, and Transwell assays. The involvement of the PI3K/AKT signaling pathway was assessed by Western blotting. Further validation was conducted in vivo experiments. Results The expression of BHLHE40 was significantly higher in OS tissues compared to adjacent tissues. In OS cell lines, BHLHE40 protein expression levels were increased compared to normal osteoblasts, and the cell line with the highest BHLHE40 expression was selected for subsequent knockdown experiments. Compared with the knockdown control group, the BHLHE40 knockdown group exhibited reduced cell viability, EdU-positive cell count, colony number, cell migration, and invasion abilities, along with downregulation of phosphorylated PI3K(p-PI3K)/PI3K and p-AKT/AKT protein expression. The aforementioned functions of BHLHE40 were also reproduced in in vivo experiments. Conclusion BHLHE40 is highly expressed in OS tissues, and its knockdown can significantly inhibit OS cell proliferation, migration, and invasion, while reducing PI3K/AKT signaling pathway activity. This suggests that BHLHE40 could serve as a novel therapeutic target for OS.
Osteosarcoma/metabolism* ; Humans ; Proto-Oncogene Proteins c-akt/genetics* ; Cell Proliferation/genetics* ; Cell Movement/genetics* ; Signal Transduction/genetics* ; Phosphatidylinositol 3-Kinases/genetics* ; Cell Line, Tumor ; Animals ; Neoplasm Invasiveness ; Basic Helix-Loop-Helix Transcription Factors/metabolism* ; Bone Neoplasms/metabolism* ; Mice ; Gene Knockdown Techniques ; Male ; Female ; Mice, Nude

Objective To investigate whether miR-15b-5p can alleviate airway inflammation in asthma by negatively regulating ubiquitin specific peptidase 9X (USP9X) to down-regulate the expression of phosphatidylinositol 4, 5-diphosphate 3-kinase catalytic subunit α/AKT serine/threonine kinase 1 (PIK3CA/AKT1) pathway. Methods USP9X was predicted to be a direct target of miR-15b-5p by using an online database (miRWalk), and the luciferase reporter gene assay was performed to verify it. Co-immunoprecipitation (CO-IP) was used to verify the direct binding between USP9X and PIK3CA and the role of USP9X and its small molecule inhibitor WP1130 in the deubiquitination of PIK3CA. C57 mice were randomly divided into Control group, OVA group, OVA combined with NC group and miR-15b-5p agomir group, with 10 mice in each group. BEAS-2B cells were induced with interleukin 13 (IL-13) and treated with miR-15b-5p mimic. HE, Masson, PAS, immunohistochemistry, immunofluorescence staining, flow cytometry, Western blot and quantitative real-time PCR(qRT-PCR) were performed. Results It was found that the administration of miR-15b-5p agomir and mimic could reduce peribronchial inflammatory cells and improve airway inflammation, and miR-15b-5p could target negative regulation of USP9X. USP9X could directly bind to PIK3CA and regulate PIK3CA level in a proteasome-dependent manner, and USP9X could deubiquitinate K29-linked PIK3CA protein. Down-regulation of USP9X could increase PIK3CA ubiquitination level. WP1130, a small molecule inhibitor of USP9X, has the same effect as knockdown of USP9X, both of which could increase the ubiquitination level of PIK3CA and reduce the protein level of PIK3CA. Conclusion The miR-15b-5p/USP9X/PIK3CA/AKT1 signaling pathway may provide potential therapeutic targets for asthma.
Animals ; MicroRNAs/metabolism* ; Asthma/pathology* ; Class I Phosphatidylinositol 3-Kinases/genetics* ; Ubiquitin Thiolesterase/metabolism* ; Proto-Oncogene Proteins c-akt/genetics* ; Mice ; Signal Transduction ; Mice, Inbred C57BL ; Humans ; Inflammation/genetics* ; Cell Line ; Female ; Male

OBJECTIVE: To investigate the effect of moxibustion on central insulin resistance related proteins of the rats suffering from diabetic cognitive decline, and analyze the underlying mechanism of moxibustion for cognition improvement. METHODS: Using the intraperitoneal injection of STZ combined with a high-fat diet, the rat model of diabetic cognitive decline were prepared. Twenty successfully-modeled rats were assigned randomly into a model group and a moxibustion group, 10 rats in each one. Besides, a blank group was set up with 10 rats collected. In the moxibustion group, suspending moxibustion was applied to "Baihui" (GV20), "Shenting" (GV24) and "Dazhui" (GV14) at the same time, 20 min in each intervention, once a day, and 6 interventions were delivered weekly and the duration of treatment was consecutive 4 weeks. The random blood glucose was measured using glucometer, and the learning-memory ability was detected by water maze test. HE staining was used to observe the morphology of neurons in the hippocampal tissue, real-time PCR assay was to detect mRNA expression of insulin receptor substrate 1 (IRS1), phosphatidylinositol 3-kinase (PI3K) and protein kinase B (AKT) in the hippocampal tissue. The Western blot method was employed to detect the protein expression of IRS1, PI3K, AKT, phosphorylated IRS1 (p-IRS1), phosphorylated PI3K (p-PI3K) and phosphorylated AKT (p-AKT) in the hippocampal tissue, and the ratio of p-IRS1/IRS1, p-PI3K/PI3K and p-AKT/AKT was calculated separately. The immunofluorescence intensity of p-IRS1, p-PI3K, and p-AKT was measured using immunofluorescence. RESULTS: Compared with the blank group, the rats of the model group exhibited higher random blood glucose (P<0.001), longer escape latency (P<0.001), severe pathological damage in the hippocampus, lower mRNA expression of IRS1, PI3K, and AKT (P<0.001), reduced ratio of p-IRS1/IRS1, p-PI3K/PI3K and p-AKT/AKT (P<0.001), and declined immunofluorescence intensity of p-IRS1, p-PI3K, and p-AKT in the hippocampal tissue (P<0.001). In comparison with the model group, for the rats of the moxibustion group, the random blood glucose decreased (P<0.05), the escape latency was shortened (P<0.01), the hippocampal pathological damage was attenuated, the mRNA expression of IRS1, PI3K and AKT increased (P<0.01), the ratio of p-IRS1/IRS1, p-PI3K/PI3K and p-AKT/AKT was elevated (P<0.01, P<0.05), and the immunofluorescence intensity of p-IRS1, p-PI3K, and p-AKT in the hippocampal tissue was strengthened (P<0.01, P<0.05). CONCLUSION In diabetic rats experiencing cognitive decline, moxibustion can enhance the learning-memory ability, which may be attributed to modulating the protein expression of IRS1, PI3K, and AKT, and their phosphorylation, activating insulin signal transduction, and reducing central insulin resistance.
Animals ; Moxibustion ; Insulin Resistance ; Rats ; Male ; Insulin Receptor Substrate Proteins/genetics* ; Rats, Sprague-Dawley ; Humans ; Proto-Oncogene Proteins c-akt/genetics* ; Cognitive Dysfunction/genetics* ; Diabetes Mellitus, Experimental/therapy* ; Hippocampus/metabolism* ; Acupuncture Points ; Phosphatidylinositol 3-Kinases/genetics*

OBJECTIVE: To observe the effect of acupuncture on chondrocyte autophagy in rats of knee osteoarthritis (KOA) and explore its underlying mechanisms. METHODS: Forty male SPF-grade SD rats were randomized into a blank group, a model group, a suspension group, an acupuncture group, and a combined therapy group, 8 rats in each one. Except the blank group, KOA model was prepared by the injection with papain. The suspension exercise therapy (10 min each time, three times daily), acupuncture (at "Yanglingquan" [GB34], "Zusanli" [ST36], and "Dubi" [ST35] on the right side, 30 min each intervention, once daily) and the combined therapy (the suspension exercise therapy combined with acupuncture) were delivered in the suspension group, the acupuncture group and the combined therapy group, respectively. The intervention of each group was performed continuously for 6 days, and 4 consecutive weeks, at the interval of 1 day. Before and after intervention, Lequesne MG score was assessed in the rats. After intervention, HE staining was adopted to observe the cartilaginous tissue morphology of the right knee joints, and Mankin score was evaluated; the serum levels of interleukin (IL)-1β, IL-6, and tumor neurosis factor-α (TNF-α) were measured using ELISA; the real-time PCR was provided to determine the mRNA expression of collagen protein type Ⅱ(COL2), collagen protein type Ⅹ (COL10), phosphatidylinositol 3-kinase (PI3K), protein kinase B (Akt), mammalian target of rapamycin (mTOR), and autophagy-regulated protein (Beclin-1) in the cartilaginous tissue of the right knee joint; Western blot was employed to detect the protein expression of PI3K, phosphorylated PI3K (p-PI3K), Akt, phosphorylated Akt (p-Akt), mTOR, phosphorylated mTOR (p-mTOR) and Beclin-1 in the cartilaginous tissue of the right knee joint. RESULTS: Compared with the blank group, the rats in the model group showed the higher Lequesne MG score (P<0.01), thinner cartilage of the right knee, reduced chondrocytes and disordered arrangement, and higher Mankin score (P<0.01). Besides, in the model group, the serum levels of IL-1β, IL-6 and TNF-α were elevated (P<0.01), the mRNA expression of COL2 and Beclin-1 and the protein expression of Beclin-1 decreased (P<0.01), the mRNA expression of COL10, PI3K, Akt and mTOR, and the protein expression of p-PI3K, p-Akt and p-mTOR increased (P<0.01) in the cartilaginous tissue of the right knee joint. Compared with the model group, in the suspension group, the acupuncture group and the combined therapy group, the Lequesne MG scores were reduced (P<0.01), the cartilage of the right knee was thickened, the arrangement of chondrocytes was improved, and the Mankin scores were lower (P<0.01). Besides, in these intervention groups, the serum levels of IL-1β, IL-6 and TNF-α were reduced (P<0.01), the mRNA expression of COL2 and Beclin-1 and the protein expression of Beclin-1 increased (P<0.05, P<0.01), the mRNA expression of COL10, PI3K, Akt and mTOR, and the protein expression of p-PI3K, p-Akt and p-mTOR decreased (P<0.05, P<0.01) in the cartilaginous tissue of the right knee joint. When compared with the suspension group and the acupuncture group, in the combined therapy group, the Lequesne MG score was reduced (P<0.01), and the Mankin score was reduced (P<0.05, P<0.01). Besides, the serum levels of IL-1β, IL-6 and TNF-α were reduced (P<0.05), the mRNA expression of COL2 and Beclin-1 and the protein expression of Beclin-1 increased (P<0.05), the mRNA expression of COL10, PI3K, Akt and mTOR, and the protein expression of p-PI3K, p-Akt and p-mTOR decreased (P<0.05, P<0.01) in the cartilaginous tissue of the right knee joint. CONCLUSION Acupuncture can promote cartilage regeneration of knee joint and autophagy in KOA rats, alleviate inflammation, so as to retard cartilage degeneration, which may be possibly associated with the PI3K/Akt/mTOR signaling pathway.
Animals ; Male ; Acupuncture Therapy ; Proto-Oncogene Proteins c-akt/genetics* ; TOR Serine-Threonine Kinases/genetics* ; Osteoarthritis, Knee/physiopathology* ; Phosphatidylinositol 3-Kinases/genetics* ; Rats ; Signal Transduction ; Rats, Sprague-Dawley ; Chondrocytes/metabolism* ; Humans ; Autophagy ; Acupuncture Points

OBJECTIVE: To explore the effect and mechanism of acupuncture at "Weizhong" (BL40) on microcirculation of paravertebral skeletal muscle in rats with chronic blunt injury of lumbar muscle based on the insulin-like growth factor-1 (IGF-1)/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. METHODS: Forty-eight SPF-grade SD rats were randomized into a blank group (8 rats) and a modeling group (40 rats). Chronic blunt injury model was established by weight impact method in the modeling group. Forty rats were successfully modeled, and were randomly divided into a model group, an acupuncture at Weizhong group (Weizhong group), an acupuncture at non-acupoint group (non-acupoint group), an inhibitor group, and an inhibitor+acupuncture at Weizhong group (inhibitor+Weizhong group), 8 rats in each group. In the Weizhong group and the inhibitor+Weizhong group, acupuncture was applied at bilateral "Weizhong" (BL40). In the non-acupoint group, acupuncture was applied at non-acupoints, i.e. points 0.5 cm inward from bilateral "Weizhong" (BL40). The acupuncture intervention was delivered 20 min each time, once a day for continuous 2 weeks. In the inhibitor group and the inhibitor+Weizhong group, intraperitoneal injection of IGF-1 receptor (IGF-1R) inhibitor was given once a day, at a dosage of 2 mg/100 g, for continuous 2 weeks. Before modeling and on the 1st, 7th and 14th days of intervention, the body mass was measured. Before and after modeling, and after intervention, the limb grip strength and paw withdrawal threshold (PWT) were measured. After intervention, the morphology of psoas muscle was observed by HE staining; the ultrastructure of psoas muscle capillaries was observed by electron microscopy; the levels of serum vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS) were detected by ELISA; and the protein and mRNA expression of IGF-1, IGF-1R, PI3K, AKT of psoas muscle was detected by Western blot and real-time PCR. RESULTS: Compared with the blank group, in the model group, the body mass on the 7th and 14th days of intervention, the limb grip strength, and the PWT of left and right hind feet were decreased (P<0.001, P<0.01); the skeletal muscle cells showed enlarged intercellular space, loosely arranged and irregularly shaped, the capillaries in the psoas muscle tissues were edematous, and the lumen of the blood vessels was obviously atrophied; the levels of serum VEGF and eNOS were decreased (P<0.001); in psoas muscle, the protein expression of IGF-1 and IGF-1R, as well as the p-PI3K/PI3K, p-AKT/AKT values were decreased (P<0.001), the mRNA expression of IGF-1, IGF-1R, PI3K and AKT was decreased (P<0.001, P<0.05). Compared with the model group, in the Weizhong group, the body weight was increased on the 7th and 14th days of intervention (P<0.001), the limb grip strength and the PWT of the left and right hind feet were increased (P<0.001, P<0.01); the arrangement of the skeletal muscle cells was relatively tight and the intercellular space was reduced, the blood vessels tended to be regular and the structure of the basement membrane was continuous, while the lumens of blood vessels were collapsed locally; the levels of serum VEGF and eNOS were increased (P<0.001); in psoas muscle, the protein expression of IGF-1 and IGF-1R, as well as the p-PI3K/PI3K, p-AKT/AKT values were increased (P<0.001), the mRNA expression of IGF-1, IGF-1R, PI3K and AKT was increased (P<0.001, P<0.01). Compared with the model group, in the inhibitor group, the body mass was decreased on the 7th and 14th days of intervention (P<0.05, P<0.01); the limb grip strength and the PWT of the left hind foot were decreased (P<0.01, P<0.001); the intercellular space of skeletal muscle cells was larger, the nuclei of the cells and erythrocytes were scattered in the intercellular space, the damage of the capillaries in the muscular tissues was serious, the collagen fibers were sparsely distributed and disorganized; the levels of serum VEGF and eNOS were decreased (P<0.001, P<0.01); in psoas muscle, the protein expression of IGF-1 and IGF-1R, as well as the p-PI3K/PI3K and p-AKT/AKT values were decreased (P<0.01, P<0.05, P<0.001), the mRNA expression of IGF-1, IGF-1R, PI3K, and AKT was decreased (P<0.01, P<0.001, P<0.05). Compared with the Weizhong group, in the non-acupoint group and the inhibitor+Weizhong group, the body mass was decreased on the 7th and 14th days of intervention (P<0.001, P<0.01), the limb grip strength was decreased (P<0.001); the morphology of muscle cell was relatively poor, with generally irregular, there was mild collapse and atrophy in the vascular lumen, and mild edema in the endothelial cells; the levels of serum VEGF and eNOS were decreased (P<0.001); in psoas muscle, the protein expression of IGF-1 and IGF-1R, as well as the p-PI3K/PI3K and p-AKT/AKT values were decreased (P<0.01, P<0.001), the mRNA expression of IGF-1, IGF-1R, PI3K, and AKT was decreased (P<0.001, P<0.01, P<0.05). Compared with the Weizhong group, the PWT of the left hind foot was decreased in the non-acupoint group (P<0.001), and PWT of the left and right hind feet was decreased in the inhibitor+Weizhong group (P<0.001). CONCLUSION Acupuncture at "Weizhong" (BL40) promotes lumbar muscle repair in chronic low back pain, its mechanism may be related to the activation of the IGF-1/PI3K/AKT pathway, thereby improving the microcirculation.
Animals ; Insulin-Like Growth Factor I/genetics* ; Acupuncture Therapy ; Rats, Sprague-Dawley ; Rats ; Proto-Oncogene Proteins c-akt/genetics* ; Male ; Humans ; Muscle, Skeletal/metabolism* ; Signal Transduction ; Phosphatidylinositol 3-Kinases/genetics* ; Wounds, Nonpenetrating/metabolism* ; Acupuncture Points

BACKGROUND: G protein-coupled receptor kinase 2 (GRK2) could participate in the regulation of diverse cells via interacting with non-G-protein-coupled receptors. In the present work, we explored how paroxetine, a GRK2 inhibitor, modulates the differentiation and activation of immune cells in rheumatoid arthritis (RA). METHODS: The blood samples of healthy individuals and RA patients were collected between July 2021 and March 2022 from the First Affiliated Hospital of Anhui Medical University. C57BL/6 mice were used to induce the collagen-induced arthritis (CIA) model. Flow cytometry analysis was used to characterize the differentiation and function of dendritic cells (DCs)/T cells. Co-immunoprecipitation was used to explore the specific molecular mechanism. RESULTS: In patients with RA, high expression of GRK2 in peripheral blood lymphocytes, accompanied by the increases of phosphatidylinositol 3 kinase (PI3K), protein kinase B (AKT), and mammalian target of rapamycin (mTOR). In animal model, a decrease in regulatory T cells (T regs ), an increase in the cluster of differentiation 8 positive (CD8 + ) T cells, and maturation of DCs were observed. Paroxetine, when used in vitro and in CIA mice, restrained the maturation of DCs and the differentiation of CD8 + T cells, and induced the proportion of T regs . Paroxetine inhibited the secretion of pro-inflammatory cytokines, the expression of C-C motif chemokine receptor 7 in DCs and T cells. Simultaneously, paroxetine upregulated the expression of programmed death ligand 1, and anti-inflammatory cytokines. Additionally, paroxetine inhibited the PI3K-AKT-mTOR metabolic pathway in both DCs and T cells. This was associated with a reduction in mitochondrial membrane potential and changes in the utilization of glucose and lipids, particularly in DCs. Paroxetine reversed PI3K-AKT pathway activation induced by 740 Y-P (a PI3K agonist) through inhibiting the interaction between GRK2 and PI3K in DCs and T cells. CONCLUSION Paroxetine exerts an immunosuppressive effect by targeting GRK2, which subsequently inhibits the metabolism-related PI3K-AKT-mTOR pathway of DCs and T cells in RA.
G-Protein-Coupled Receptor Kinase 2/metabolism* ; Arthritis, Rheumatoid/immunology* ; Animals ; Dendritic Cells/metabolism* ; Paroxetine/therapeutic use* ; Proto-Oncogene Proteins c-akt/metabolism* ; Mice ; Humans ; Mice, Inbred C57BL ; Signal Transduction/drug effects* ; Male ; Phosphatidylinositol 3-Kinases/metabolism* ; Lymphocyte Activation/drug effects* ; Female ; T-Lymphocytes/metabolism* ; Middle Aged

BACKGROUND: Arsenic trioxide (ATO) is indicated as a broad-spectrum medicine for a variety of diseases, including cancer and cardiac disease. While the role of ATO in hepatic ischemia/reperfusion injury (HIRI) has not been reported. Thus, the purpose of this study was to identify the effects of ATO on HIRI. METHODS: In the present study, we established a 70% hepatic warm I/R injury and partial hepatectomy (30% resection) animal models in vivo and hepatocytes anoxia/reoxygenation (A/R) models in vitro with ATO pretreatment and further assessed liver function by histopathologic changes, enzyme-linked immunosorbent assay, cell counting kit-8, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. Small interfering RNA (siRNA) for extracellular signal-regulated kinase (ERK) 1/2 was transfected to evaluate the role of ERK1/2 pathway during HIRI, followed by ATO pretreatment. The dynamic process of autophagic flux and numbers of autophagosomes were detected by green fluorescent protein-monomeric red fluorescent protein-LC3 (GFP-mRFP-LC3) staining and transmission electron microscopy. RESULTS: A low dose of ATO (0.75 μmol/L in vitro and 1 mg/kg in vivo ) significantly reduced tissue necrosis, inflammatory infiltration, and hepatocyte apoptosis during the process of hepatic I/R. Meanwhile, ATO obviously promoted the ability of cell proliferation and liver regeneration. Mechanistically, in vitro  studies have shown that nontoxic concentrations of ATO can activate both ERK and phosphoinositide 3-kinase-serine/threonine kinase (PI3K-AKT) pathways and further induce autophagy. The hepatoprotective mechanism of ATO, at least in part, relies on the effects of ATO on the activation of autophagy, which is ERK-dependent. CONCLUSION Low, non-toxic doses of ATO can activate ERK/PI3K-AKT pathways and induce ERK-dependent autophagy in hepatocytes, protecting liver against I/R injury and accelerating hepatocyte regeneration after partial hepatectomy.
Animals ; Arsenic Trioxide ; Autophagy/physiology* ; Reperfusion Injury/prevention & control* ; Mice ; Male ; Proto-Oncogene Proteins c-akt/physiology* ; Arsenicals/therapeutic use* ; Oxides/therapeutic use* ; Liver/metabolism* ; Extracellular Signal-Regulated MAP Kinases/metabolism* ; Mice, Inbred C57BL

BACKGROUND: Pancreatic cancer is a lethal malignancy prone to gemcitabine resistance. The single-strand selective monofunctional uracil DNA glycosylase (SMUG1), which is responsible for initiating base excision repair, has been reported to predict the outcomes of different cancer types. However, the function of SMUG1 in pancreatic cancer is still unclear. METHODS: Gene and protein expression of SMUG1 as well as survival outcomes were assessed by bioinformatic analysis and verified in a cohort from Fudan University Shanghai Cancer Center. Subsequently, the effect of SMUG1 on proliferation, cell cycle, and migration abilities of SMUG1 cells were detected in vitro . DNA damage repair, apoptosis, and gemcitabine resistance were also tested. RNA sequencing was performed to determine the differentially expressed genes and signaling pathways, followed by quantitative real-time polymerase chain reaction and Western blotting verification. The cancer-promoting effect of forkhead box Q1 (FOXQ1) and SMUG1 on the ubiquitylation of myelocytomatosis oncogene (c-Myc) was also evaluated. Finally, a xenograft model was established to verify the results. RESULTS: SMUG1 was highly expressed in pancreatic tumor tissues and cells, which also predicted a poor prognosis. Downregulation of SMUG1 inhibited the proliferation, G1 to S transition, migration, and DNA damage repair ability against gemcitabine in pancreatic cancer cells. SMUG1 exerted its function by binding with FOXQ1 to activate the Protein Kinase B (AKT)/p21 and p27 pathway. Moreover, SMUG1 also stabilized the c-Myc protein via AKT signaling in pancreatic cancer cells. CONCLUSIONS SMUG1 promotes proliferation, migration, gemcitabine resistance, and c-Myc protein stability in pancreatic cancer via protein kinase B signaling through binding with FOXQ1. Furthermore, SMUG1 may be a new potential prognostic and gemcitabine resistance predictor in pancreatic ductal adenocarcinoma.
Humans ; Pancreatic Neoplasms/pathology* ; Forkhead Transcription Factors/genetics* ; Signal Transduction/genetics* ; Animals ; Cell Line, Tumor ; Proto-Oncogene Proteins c-akt/metabolism* ; Cell Proliferation/physiology* ; Mice ; Uracil-DNA Glycosidase/genetics* ; Female ; Male ; Gemcitabine ; Mice, Nude ; Apoptosis/physiology* ; Deoxycytidine/analogs & derivatives* ; Cell Movement/genetics*