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The DNA Repair Enzyme XPD Is Partially Regulated by PI3K/AKT Signaling in the Context of Bupivacaine-Mediated Neuronal DNA Damage

ABSTRACT: Bupivacaine, a local anesthetic widely used for regional anesthesia and pain management, has been reported to induce neuronal injury, especially DNA damage. Neurons employ different pathways to repair DNA damage. However, the mechanism underlying bupivacaine-mediated DNA damage repair is unclear. A rat neuronal injury model was established by intrathecal injection of (3%) bupivacaine. An in vitro neuronal injury model was generated by exposing SH-SY5Y cells to bupivacaine (1.5 mmol/L). Then, a cDNA plate array was used to identify the DNA repair genes after bupivacaine exposure. The results showed that xeroderma pigmentosum complementary group D (XPD) of the nuclear excision repair (NER) pathway was closely associated with the repair of DNA damage induced by bupivacaine. Subsequently, Western blot assay and immunohistochemistry indicated that the expression of the repair enzyme XPD was upregulated after DNA damage. Downregulation of XPD expression by a lentivirus aggravated the DNA damage induced by bupivacaine. In addition, phosphatidyl-3-kinase (PI3K)/AKT signaling in neurons was inhibited after exposure to bupivacaine. After PI3K/AKT signaling was inhibited, bupivacaine-mediated DNA damage was further aggravated, and the expression of XPD was further upregulated. However, knockdown of XPD aggravated bupivacaine-mediated neuronal injury but did not affect PI3K/AKT signaling. In conclusion, the repair enzyme XPD, which was partially regulated by PI3K/AKT signaling, responded to bupivacaine-mediated neuronal DNA damage. These results can be used as a reference for the treatment of bupivacaine-induced neurotoxicity.

SUBMITTER: Zhao W

PROVIDER: S-EPMC8516563 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Esophageal squamous cell carcinoma (ESCC) is a lethal disease due to its high aggressiveness. The aim of the present study was to investigate the role of xeroderma pigmentosum complementation group D (XPD) in the growth and invasion of ESCC and to elucidate the potential underlying molecular mechanisms. Western blot analysis and RT?qPCR were used to detect the expression level of XPD in ESCC tissue samples and adjacent normal esophageal tissue samples. The pEGFP?N2/XPD plasmid was transfected into human ESCC cell lines (EC9706 and EC109). The proliferation, apoptosis, migration and invasion of EC9706 or EC109 cells were assessed following transfection with the XPD overexpression plasmid. The chemosensitivity of EC9706 or EC109 cells to cisplatin or fluorouracil was evaluated by CCK?8 assay. The expression levels of phosphoinositide 3?kinase (PI3K)/AKT, nuclear factor (NF)??B, Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) and mitogen?activated protein kinase (MAPK) signaling pathway?related genes were detected by RT?qPCR and western blot analysis. The results demonstrated that the expression level of XPD was markedly lower in ESCC tissue samples than in adjacent normal esophageal tissue samples. The pEGFP?N2/XPD plasmid was successfully transfected into EC9706 or EC109 cells, inducing XPD overexpression. A High XPD expression markedly suppressed cell proliferation, migration and invasion, and increased the apoptotic rate of EC9706 and EC109 cells. Furthermore, the overexpression of XPD significantly increased the chemosensitivity of EC9706 and EC109 cells to cisplatin or fluorouracil. Following XPD overexpression, the expression levels of PI3K, p?AKT, c?Myc, Cyclin D1, Bcl?2, vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP)?9 were markedly downregulated, while the expression level of p21 was markedly upregulated. On the whole, the findings of the present study demonstrate that XPD inhibits the growth and invasion of EC9706 and EC109 cells, whilst also enhancing the chemosensitivity of EC9706 and EC109 cells to cisplatin or fluorouracil by regulating the PI3K/AKT signaling pathway. XPD may thus be an underlying target for ESCC treatment and drug resistance.

Project description:BackgroundEffective removal of pathogenic bacteria is key to improving the prognosis of sepsis. Polymorphonuclear neutrophils (PMNs) are the most important components of innate cellular immunity and play vital roles in clearing pathogenic bacteria. However, the metabolic characteristics and immunomodulatory pathways of PMNs during sepsis have not been investigated. In the present study, we explored the immune metabolism characteristics of PMNs and the mechanism by which neutrophilic glycolysis is regulated during sepsis.MethodsMetabolomics analysis was performed on PMNs isolated from 14 septic patients, 26 patients with acute appendicitis, and 19 healthy volunteers. Transcriptome analysis was performed on the PMNs isolated from the healthy volunteers and the patients with sepsis to assess glycolysis and investigate its mechanism. Lipopolysaccharide (LPS) was used to stimulate the neutrophils isolated from the healthy volunteers at different time intervals to build an LPS-tolerant model. Chemotaxis, phagocytosis, lactate production, oxygen consumption rate (OCR), and extracellular acidification rate (ECAR) were evaluated.ResultsTranscriptomics showed significant changes in glycolysis and the mTOR/HIF-1α signaling pathway during sepsis. Metabolomics revealed that the Warburg effect was significantly altered in the patients with sepsis. We discovered that glycolysis regulated PMNs' chemotaxis and phagocytosis functions during sepsis. Lactate dehydrogenase A (LDHA) downregulation was a key factor in the inhibition of glycolysis in PMNs. This study confirmed that the PI3K/Akt-HIF-1α pathway was involved in the LDHA expression level and also influenced PMNs' chemotaxis and phagocytosis functions.ConclusionsThe inhibition of glycolysis contributed to neutrophil immunosuppression during sepsis and might be controlled by PI3K/Akt-HIF-1α pathway-mediated LDHA downregulation. Our study provides a scientific theoretical basis for the management and treatment of patients with sepsis and promotes to identify therapeutic target for the improvement of immune function in sepsis.

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The DNA Repair Enzyme XPD Is Partially Regulated by PI3K/AKT Signaling in the Context of Bupivacaine-Mediated Neuronal DNA Damage

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