NRG1-ErbB4 signaling in the medial amygdala controls mating motivation in adult male mice.

Motivation-driven mating is a basic affair for the maintenance of species. However, the underlying molecular mechanisms that control mating motivation are not fully understood. Here, we report that NRG1-ErbB4 signaling in the medial amygdala (MeA) is pivotal in regulating mating motivation. NRG1 expression in the MeA negatively correlates with the mating motivation levels in adult male mice. Local injection and knockdown of MeA NRG1 reduce and promote mating motivation, respectively. Consistently, knockdown of MeA ErbB4, a major receptor for NRG1, and genetic inactivation of its kinase both promote mating motivation. ErbB4 deletion decreases neuronal excitability, whereas chemogenetic manipulations of ErbB4-positive neuronal activities bidirectionally modulate mating motivation. We also identify that the effects of NRG1-ErbB4 signaling on neuronal excitability and mating motivation rely on hyperpolarization-activated cyclic nucleotide-gated channel 3. This study reveals a critical molecular mechanism for regulating mating motivation in adult male mice.

EGFR promotes the apoptosis of CD4+ T lymphocytes through TBK1/Glut1 induced Warburg effect in sepsis.

INTRODUCTION: Sepsis-induced apoptosis leads to lymphopenia including the decrease of CD4+ T cells thus favoring immunosuppression. OBJECTIVES: Although epidermal growth factor receptor (EGFR) inhibitors significantly improve the survival rate of septic mice, the effect of EGFR on the function and metabolism of CD4+ T cells in sepsis remained unknown. METHODS: CD4+ T cells from septic mice and patients were assessed for apoptosis, activation, Warburg metabolism and glucose transporter 1 (Glut1) expression with or without the interference of EGFR activation. RESULTS: EGFR facilitates CD4+ T cell activation and apoptosis through Glut1, which is a key enzyme that controls glycolysis in T cells. EGFR, TANK binding kinase 1 (TBK1) and Glut1 form a complex to facilitate Glut1 transportation from cytoplasm to cell surface. Both the levels of membrane expression of EGFR and Glut1 and the activation levels of CD4+ T cells were significantly higher in patients with sepsis as compared with healthy subjects. CONCLUSION: Our data demonstrated that through its downstream TBK1/Exo84/RalA protein system, EGFR regulates Glut1 transporting to the cell surface, which is a key step for inducing the Warburg effect and the subsequent cellular activation and apoptosis of CD4+ T lymphocytes and may eventually affect the immune functional status, causing immune cell exhaustion in sepsis.

EGFR tyrosine kinase activity and Rab GTPases coordinate EGFR trafficking to regulate macrophage activation in sepsis.

EGFR phosphorylation is required for TLR4-mediated macrophage activation during sepsis. However, whether and how intracellular EGFR is transported during endotoxemia have largely been unknown. Here, we show that LPS promotes high levels cell surface expression of EGFR in macrophages through two different transport mechanisms. On one hand, Rab10 is required for EEA1-mediated the membrane translocation of EGFR from the Golgi. On the other hand, EGFR phosphorylation prevents its endocytosis in a kinase activity-dependent manner. Erlotinib, an EGFR tyrosine kinase inhibitor, significantly reduced membrane EGFR expression in LPS-activated macrophage. Mechanistically, upon LPS induced TLR4/EGFR phosphorylation, MAPK14 phosphorylated Rab7a at S72 impaired membrane receptor late endocytosis, which maintains EGFR membrane localization though blocking its lysosomal degradation. Meanwhile, Rab5a is also involved in the early endocytosis of EGFR. Subsequently, inhibition of EGFR phosphorylation switches M1 phenotype to M2 phenotype and alleviates sepsis-induced acute lung injury. Mechanistic study demonstrated that Erlotinib suppressed glycolysis-dependent M1 polarization via PKM2/HIF-1ɑ pathway and promoted M2 polarization through up-regulating PPARγ induced glutamine metabolism. Collectively, our data elucidated a more in-depth mechanism of macrophages activation, and provided stronger evidence supporting EGFR as a potential therapeutic target for the treatment of sepsis.

Effects of Continuous Fascia Iliaca Compartment Block on Early Quality of Recovery After Total Hip Arthroplasty in Elderly Patients: A Randomized Controlled Trial.

Purpose: In recent years, patient-centered postoperative quality of recovery has gained attention. This study aimed to assess the influence of ultrasound-guided continuous fascia iliaca compartment block (CFICB) on early quality of recovery in elderly patients after total hip arthroplasty (THA) using the QoR-15 score. Patients and Methods: In this single-center, randomized, prospective study, 60 patients scheduled for unilateral THA were randomized to the CFICB or patient-controlled intravenous analgesia (PCIA) group. The primary outcome was the QoR-15 score. The secondary outcomes were pain score, number of patients requiring rescue analgesics, time of first postoperative ambulation, incidence of postoperative complications, Bromage score, and length of hospital stay. Results: The QoR-15 score was significantly higher in the CFICB group than in the PCIA group at 24 h (P < 0.001) after surgery. However, the QoR-15 score was not significantly different at 48 h (P = 0.074) between the two groups. Pain scores at rest and during movement were lower in the CFICB group than in the PCIA group at 12, 24, and 48 h postoperatively (P < 0.05). There was no difference in the number of patients requiring rescue analgesics, time of first postoperative ambulation, incidence of postoperative complications apart from dizziness, or length of hospital stay between the two groups. In addition, Bromage score of 1 point was reported by four patients in the CFICB group at 24 h (P = 0.048) after THA. Conclusion: In elderly patients following THA, CFICB improved the quality of recovery at 24 h and reduced pain scores compared with PCIA. The time of first postoperative ambulation and length of hospital stay were not significantly affected.

Macrophage-derived exosomal aminopeptidase N aggravates sepsis-induced acute lung injury by regulating necroptosis of lung epithelial cell.

Sepsis-induced acute lung injury (ALI) is a serious sepsis complication and the prevailing cause of death. Circulating plasma exosomes might exert a key role in regulating intercellular communication between immunological and structural cells, as well as contributing to sepsis-related organ damage. However, the molecular mechanisms by which exosome-mediated intercellular signaling exacerbate ALI in septic infection remains undefined. Therefore, we investigated the effect of macrophage-derived exosomal APN/CD13 on the induction of epithelial cell necrosis. Exosomal APN/CD13 levels in the plasma of septic mice and patients with septic ALI were found to be higher. Furthermore, increased plasma exosomal APN/CD13 levels were associated with the severity of ALI and fatality in sepsis patients. We found remarkably high expression of APN/CD13 in exosomes secreted by LPS-stimulated macrophages. Moreover, c-Myc directly induced APN/CD13 expression and was packed into exosomes. Finally, exosomal APN/CD13 from macrophages regulated necroptosis of lung epithelial cells by binding to the cell surface receptor TLR4 to induce ROS generation, mitochondrial dysfunction and NF-κB activation. These results demonstrate that macrophage-secreted exosomal APN/CD13 can trigger epithelial cell necroptosis in an APN/CD13-dependent manner, which provides insight into the mechanism of epithelial cell functional disorder in sepsis-induced ALI.

Cytosolic p53 Inhibits Parkin-Mediated Mitophagy and Promotes Acute Liver Injury Induced by Heat Stroke.

Heat stroke (HS) is a severe condition characterized by increased morbidity and high mortality. Acute liver injury (ALI) is a well-documented complication of HS. The tumor suppressor p53 plays an important role in regulation of mitochondrial integrity and mitophagy in several forms of ALI. However, the role of p53-regulated mitophagy in HS-ALI remains unclear. In our study, we discovered the dynamic changes of mitophagy in hepatocytes and demonstrated the protective effects of mitophagy activation on HS-ALI. Pretreatment with 3-MA or Mdivi-1 significantly exacerbated ALI by inhibiting mitophagy in HS-ALI mice. Consistent with the animal HS-ALI model results, silencing Parkin aggravated mitochondrial damage and apoptosis by inhibiting mitophagy in HS-treated normal human liver cell line (LO2 cells). Moreover, we described an increase in the translocation of p53 from the nucleus to the cytoplasm, and cytosolic p53 binds to Parkin in LO2 cells following HS. p53 overexpression using a specific adenovirus or Tenovin-6 exacerbated HS-ALI through Parkin-dependent mitophagy both in vivo and in vitro, whereas inhibition of p53 using siRNA or PFT-α effectively reversed this process. Our results demonstrate that cytosolic p53 binds to Parkin and inhibits mitophagy by preventing Parkin's translocation from the cytosol to the mitochondria, which decreases mitophagy activation and leads to hepatocyte apoptosis in HS-ALI. Overall, pharmacologic induction of mitophagy by inhibiting p53 may be a promising therapeutic approach for HS-ALI treatment.

Validation of novel hub genes and molecular mechanisms in acute lung injury using an integrative bioinformatics approach.

Acute lung injury (ALI) is an inflammatory pulmonary disease that can easily develop into serious acute respiratory distress syndrome, which has high morbidity and mortality. However, the molecular mechanism of ALI remains unclear, and few molecular biomarkers for diagnosis and treatment have been identified. In this study, we aimed to identify novel molecular biomarkers using a bioinformatics approach. Gene expression data were obtained from the Gene Expression Omnibus database, co-expressed differentially expressed genes (CoDEGs) were identified using R software, and further functional enrichment analyses were conducted using the online tool Database for Annotation, Visualization, and Integrated Discovery. A protein-protein interaction network was established using the STRING database and Cytoscape software. Lipopolysaccharide (LPS)-induced ALI mouse model was constructed and verified. The hub genes were screened and validated in vivo. The transcription factors (TFs) and miRNAs associated with the hub genes were predicted using the NetworkAnalyst database. In total, 71 CoDEGs were screened and found to be mainly involved in the cytokine-cytokine receptor interactions, and the tumor necrosis factor and malaria signaling pathways. Animal experiments showed that the lung injury score, bronchoalveolar lavage fluid protein concentration, and wet-to-dry weight ratio were higher in the LPS group than those in the control group. Real-time polymerase chain reaction analysis indicated that most of the hub genes such as colony-stimulating factor 2 (Csf2) were overexpressed in the LPS group. A total of 20 TFs including nuclear respiratory factor 1 (NRF1) and two miRNAs were predicted to be regulators of the hub genes. In summary, Csf2 may serve as a novel diagnostic and therapeutic target for ALI. NRF1 and mmu-mir-122-5p may be key regulators in the development of ALI.

microRNA-219-5p targets NEK6 to inhibit hepatocellular carcinoma progression.

MicroRNA-219-5p (miR-219-5p) is a key post-transcriptional regulator of gene expression that is known to regulate cancer progression, but its role in the context of hepatocellular carcinoma (HCC) remains to be fully elucidated. Herein, it was found that this miRNA functions as a tumor suppressor. Specifically, significant decreases in miR-219-5p expression were detected in HCC cells and patient serum samples relative to that found in the serum of 15 healthy people, and it was concluded that miR-219-5p overexpression was sufficient to impair HCC cell proliferation in vitro and vivo and migration in vitro. At the mechanistic level, it was found that miR-219-5p was able to suppress the expression of NEK6 (never in mitosis gene a-related kinase 6), thereby resulting in dysregulated ß-catenin/c-Myc-regulated gene expression. When NEK6 was overexpressed in HCC cells, this was sufficient to reverse the inhibitory impact of miR-219-5p on HCC cell proliferation both in vitro and vivo and metastasis in vitro. Bioinformatics analyses were also conducted, and both miR-219-5p and Nek6 were linked to disease progression in HCC patients with advanced disease. More importantly, the serum specimen data showed that reduced perioperative plasma miR-219-5p correlated significantly with increased risk of early recurrence after curative hepatectomy, whereas it was opposed to NEK6. Together, these findings highlight miR-219-5p as a potentially valuable diagnostic biomarker that can potentially be leveraged to improve clinical outcomes in HCC patients.

Bcl-2 Proteins Regulate Mitophagy in Lipopolysaccharide-Induced Acute Lung Injury via PINK1/Parkin Signaling Pathway.

Mitophagy is involved in sepsis-induced acute lung injury (ALI). Bcl-2 family proteins play an important role in mitochondrial homeostasis. However, whether targeting Bcl-2 proteins (Bcl-2 and Bad) could influence mitophagy in ALI remains unclear. In this study, lipopolysaccharide (LPS) was used to induce injury in A549 cells and ALI in mice. LPS treatment resulted in elevated cell apoptosis, enhanced mitophagy, decreased Bcl-2 expression, increased Bad expression, and activation of PINK1/Parkin signaling in cells and lung tissues. Both Bcl-2 overexpression and Bad knockdown attenuated LPS-induced injury, inhibited cell apoptosis and mitophagy, and improved survival. Atg5 knockout (KO) inhibited LPS-induced cell apoptosis. Furthermore, Bcl-2 proteins regulated mitophagy by modulating the recruitment of Parkin from the cytoplasm to mitochondria via direct protein-protein interactions. These results were further confirmed in Park2 KO cells and Park2-/- mice. This is the first study to demonstrate that Bcl-2 proteins regulated mitophagy in LPS-induced ALI via modulating the PINK1/Parkin signaling pathway, promoting new insights into the mechanisms and investigation of therapeutic strategies for a septic patient with ALI.

Propofol inhibits biological functions of leukaemia stem and differentiated cells through suppressing Wnt/ß-catenin and Akt/mTOR.

The biological roles of intravenous anaesthetic propofol in cancer have been shown by various studies using cancer cell lines that represent differentiated cancer cells. However, the activities of propofol in cancer stem cells have not been elucidated. In this work, we examined the effects and mechanisms of propofol on acute myeloid leukaemia (AML) differentiated and CD34+ CD38- stem cells. We found that propofol inhibited growth, differentiation and self-renewal capabilities of AML stem cells regardless of cellular origin and genetic profiling. In addition, propofol inhibited the growth of AML differentiated cells. Propofol significantly induced apoptosis of AML differentiated but not CD34+ CD38- stem cells. We further found that propofol significantly augmented the efficacy of AML standard therapeutic drugs. Consistent with the previous findings, we showed that propofol suppressed the Akt/mTOR pathway in AML cells. We also found that propofol inhibited pathways important for stem cell maintenance and self-renewal, such as Wnt/ß-catenin. Overexpression of constitutively active Akt partially reversed the inhibitory effects of propofol in AML differentiated cells. Stabilization of ß-catenin using genetic and pharmacological approaches also partially rescued the inhibitory effects of propofol in AML differentiated and stem cells. Our work shows that propofol targets leukaemia cells at all stages of development, in a cell type-specific manner. Inhibition of both Akt/mTOR and Wnt/ß-catenin is required for the action of propofol in AML. Our findings also highlight the activities of propofol on cancer stem cells.

Efficacy and Tolerability of Sufentanil, Dexmedetomidine, or Ketamine Added to Propofol-based Sedation for Gastrointestinal Endoscopy in Elderly Patients: A Prospective, Randomized, Controlled Trial.

PURPOSE: To investigate the optimal agent combined with propofol for sedation in elderly patients undergoing gastrointestinal endoscopy. METHODS: A total of 120 elderly patients scheduled for gastrointestinal endoscopy under propofol-based sedation were randomly allocated to receive propofol + saline (control group), propofol + sufentanil 0.1 µg/kg, propofol + dexmedetomidine 0.4 µg/kg, or propofol + ketamine 0.4 mg/kg. Mean arterial pressure, heart rate, pulse oximetry, pressure of end-tidal carbon dioxide, respiratory rate, and Ramsay sedation scale score were recorded. Induction time, procedure time, recovery time, propofol dose, and adverse events were also recorded. FINDINGS: During the sedation procedure, the AUC of HR was lowest in the propofol + dexmedetomidine group (all, P < 0.05), and the AUC of pulse oximetry was significantly higher in the propofol + dexmedetomidine and propofol + ketamine groups compared to the other 2 groups (both, P < 0.05). The propofol + dexmedetomidine group had the highest prevalences of hypotension and bradycardia, and the control group experienced the largest number of hypoxia episodes (all, P < 0.05). The control group consumed the highest dose of propofol, while the propofol + ketamine group needed the lowest dose (all, P < 0.05). IMPLICATIONS: The combination of propofol + ketamine 0.4 mg/kg maintained hemodynamic and respiratory stability, as evidenced by less hypotension, bradycardia, and hypoxia events, in elderly patients undergoing gastrointestinal endoscopy. China clinical trial registration (chictr.org.cn) ID: ChiCTR-INR-17013710.

Propofol inhibits parthanatos via ROS-ER-calcium-mitochondria signal pathway in vivo and vitro.

Parthanatos is a new form of programmed cell death. It has been recognized to be critical in cerebral ischemia-reperfusion injury, and reactive oxygen species (ROS) can induce parthanatos. Recent studies found that propofol, a widely used intravenous anesthetic agent, has an inhibitory effect on ROS and has neuroprotective in many neurological diseases. However, the functional roles and mechanisms of propofol in parthanatos remain unclear. Here, we discovered that the ROS-ER-calcium-mitochondria signal pathway mediated parthanatos and the significance of propofol in parthanatos. Next, we found that ROS overproduction would cause endoplasmic reticulum (ER) calcium release, leading to mitochondria depolarization with the loss of mitochondrial membrane potential. Mitochondria depolarization caused mitochondria to release more ROS, which, in turn, contributed to parthanatos. Also, we found that propofol inhibited parthanatos through impeding ROS overproduction, calcium release from ER, and mitochondrial depolarization in parthanatos. Importantly, our results indicated that propofol protected cerebral ischemia-reperfusion via parthanatos suppression, amelioration of mitochondria, and ER swelling. Our findings provide new insights into the mechanisms of how ER and mitochondria contribute to parthanatos. Furthermore, our studies elucidated that propofol has a vital role in parthanatos prevention in vivo and in vitro, and propofol can be a promising therapeutic approach for nerve injury patients.

Propofol inhibits lipopolysaccharide-induced tumor necrosis factor-alpha expression and myocardial depression through decreasing the generation of superoxide anion in cardiomyocytes.

TNF-α has been shown to be a major factor responsible for myocardial depression in sepsis. The aim of this study was to investigate the effect of an anesthetic, propofol, on TNF-α expression in cardiomyocytes treated with LPS both in vivo and in vitro. In cultured cardiomyocytes, compared with control group, propofol significantly reduced protein expression of gp91phox and phosphorylation of extracellular regulated protein kinases 1/2 (ERK1/2) and p38 MAPK, which associates with reduced TNF-α production. In in vivo mice studies, propofol significantly improved myocardial depression and increased survival rate of mice after LPS treatment or during endotoxemia, which associates with reduced myocardial TNF-α production, gp91phox, ERK1/2, and p38 MAPK. It is concluded that propofol abrogates LPS-induced TNF-α production and alleviates cardiac depression through gp91phox/ERK1/2 or p38 MAPK signal pathway. These findings have great clinical importance in the application of propofol for patients enduring sepsis.

[Effects of hypertonic sodium chloride hydroxyethyl starch solution on cerebral vasospasm following subarachnoid hemorrhage and its mechanism].

OBJECTIVE: To investigate the protective effect and potential mechanisms of hypertonic sodium chloride hydroxyethyl starch solution (HSH) against the cerebral vasospasm (CVS) following subarachnoid hemorrhage (SAH). METHODS: Twenty-four male Sprague-Dawley (SD) rats were randomly assigned to four groups according to the random number table, with 6 rats in each group. The SAH-CVS model was reproduced by injection of the blood twice through the cisterna magna. Rats in both model and HSH treatment groups received 8 mL/kg normal saline (NS) or HSH treatment everyday via caudal vein. Rats in sham group were injected with 1.5 mL/kg NS into cisterna magna followed by 8 mL/kg NS treatment. Rats in normal group received no treatment. Rats were sacrificed to harvest basilar artery after 7 days. The thickness of vessel wall and lumen area were measured using hematoxylin-eosin (HE) staining. The rate of apoptosis of vascular smooth muscle cell (VSMC) was assessed using flow cytometry. Caspase-3 activity was measured by a fluorometric assay. The expressions of Bax and Bcl-2 were determined by Western Blot. Intracellular reactive oxygen species (ROS) was detected by H2DCFDA. RESULTS: Compared with normal group, increased thickness of vessel wall (27.72 ± 1.94 µm vs. 18.30 ± 1.10 µm, P<0.05), decreased lumen area (26 115 ± 1 991 µm² vs. 55 080 ± 2 091 µm², P<0.05), and elevation of rate of apoptosis of VSMCs [(35.05 ± 5.54) % vs. (5.93 ± 1.53) %, P<0.05] were found in model group. Compared with model group, decreased thickness of vessel wall (22.55 ± 1.50 µm vs. 27.72 ± 1.94 µm, P<0.05), increase of lumen area (48 115 ± 2 460 µm² vs. 26 115 ± 1 991 µm², P<0.05), and depressed rate of apoptosis of VSMCs [(16.54 ± 5.94) % vs. (35.05 ± 5.54) %, P<0.05] were found in HSH treatment group. Caspase-3 activity, intracellular ROS level, Bax and Bcl-2 expressions in model group were (188.40 ± 19.35)%, (163.50 ± 17.02)%, (208.71 ± 26.04)% and (44.52 ± 9.61) % of those of normal group, and the differences of these parameters between model and normal groups were statistically significant (all P<0.05). Caspase-3 activity, intracellular ROS level, Bax and Bcl-2 expressions in HSH treatment group were (135.05 ± 19.52)%, (119.44 ± 11.50)%, (139.20 ± 18.04)% and (85.35 ± 13.12)% of those of normal group, respectively, and the differences of these parameters between HSH treatment and model groups were statistically significant (all P<0.05). The differences of all measurements between sham and normal groups were not statistically significant. CONCLUSIONS: The current results demonstrate that HSH attenuates the SAH-induced CVS, alleviates thickness of vessel wall, and increases lumen area via inhibition of VSMCs apoptosis.

Proteomic profiling of the phosphoproteins in the rat thalamus, hippocampus and frontal lobe after propofol anesthesia.

BACKGROUND: Propofol is a safe and effective intravenous anesthetic that is widely used for the induction and maintenance of anesthesia during surgery. However, the mechanism by which propofol exerts its anesthetic effect remains unknown. The rapid onset of phosphorylation modifications coincides with that of propofol anesthesia. METHODS: Propofol-anesthetized rat models were built and phosphorylated proteins in the thalamus, hippocampus and frontal lobe were enriched the to analyze the changes in these phosphoproteins after propofol anesthesia. RESULTS: Sixteen of these phosphoprotein spots were successfully identified using MALDI-TOF MS and a subsequent comparative sequence search in the Mascot database. Of these proteins, keratin 18 and the tubulin 2c chain are cytoskeletal proteins; keratin 18 and gelsolin are relevant to alcohol drowsiness. Based on Western blot analysis, we also confirmed that the phosphorylation of these proteins is directly induced by propofol, indicating that propofol anesthesia may be relevant to cytoskeletal proteins and alcohol drowsiness. CONCLUSIONS: These identified propofol-induced phosphorylations of proteins provide meaningful contributions for further studying the anesthetic mechanism of propofol.

Propofol inhibits the activation of p38 through up-regulating the expression of annexin A1 to exert its anti-inflammation effect.

Inflammatory response is a kind of nonspecific immune response, with the central link of vascular response, which is mainly manifested by changes in neutrophils and vascular endothelial cells. In recent years, the in vivo and in vitro role of intravenous anesthetic propofol in inhibiting inflammatory response has been attracting more and more attention, but the anti-inflammatory mechanisms of propofol for mononuclear cells still remain undefined. In this study, proteomics analysis was applied to investigate protein expression profile changes in serum mononuclear cells following intervention of rats with endotoxemia using propofol. After two-dimensional electrophoresis and mass spectrometric identification, it has been found that the protein Annexin A1 was up-regulated in the propofol intervention group. Annexin A1 is a glucocorticoid-dependent anti-inflammatory protein. After detection using ELISA and Western blot assays, it has also been found that propofol can not only promote the expression of Annexin A1, but also inhibit the phosphorylation level of p38 and release of inflammatory factors (IL-1ß, IL-6 and TNF-α) in rats with endotoxemia. In order to further determine the role of up-regulated expression of Annexin A1 in anti-inflammation of propofol, this gene was silenced in vitro in human THP-1 cells, to detect the phosphorylation status of p38 and release of inflammatory factors. The results show that Annexin A1 can negatively regulate phosphorylation of p38 and release of IL-1ß, IL-6 and TNF-α in THP-1 cells following propofol intervention and lipopolysaccharide (LPS) stimulation. Our results clearly indicate that propofol can up-regulate Annexin A1 to inhibit the phosphorylation level of p38 and release of IL-1ß, IL-6 and TNF-α, so as to inhibit inflammatory response. Therefore, it can be speculated that Annexin A1 might be the key signaling protein in the in vivo and in vitro anti-inflammatory mechanisms of propofol.