Celecoxib Inhibits Hepatocellular Carcinoma Cell Growth and Migration by Targeting PNO1.

BACKGROUND Celecoxib has shown anti-tumor activities against several types of cancer. Although the majority of research focuses on its mechanism via cyclooxygenase-2 (COX-2) enzyme inhibition, we identified a distinct mechanism behind celecoxib anti-cancer abilities. MATERIAL AND METHODS We treated hepatocellular carcinoma (HCC) Huh-7 cells and tumor xenograft mice models with celecoxib to test its effects on the tumor. Using gene chip method to identify the differential expressed genes after celecoxib treatment and using pathway enrichment analysis to predict the potential pathways for further study. We transfected cells with lentiviral shRNA to detect the effect of RNA binding gene partner of NOB1 (PNO1) on tumor growth in vitro and in vivo. Further we performed western blot to detect the effect of PNO1 on the protein kinase B (AKT) pathway. RESULTS Celecoxib inhibited HCC cell growth in vitro and in vivo, and gene chip and pathway enrichment analysis revealed that PNO1 may be the potential target of celecoxib in HCC cells. Celecoxib significantly reduced levels of PNO1 in tumor tissue. Knockdown of PNO1 remarkably suppressed tumor growth and metastasis in vitro and in vivo. Disruption of PNO1 expression significantly reduced protein kinase B (AKT)/rapamycin (mTOR) signaling, indicating that this pathway may be involved in PNO1-mediated tumorigenic activity. CONCLUSIONS Celecoxib may exert its anti-tumor activity by inhibiting PNO1, and that AKT/mTOR signaling helps mediate the oncogenic effects of PNO1. This work offers the first evidence for a role of PNO1 as an HCC oncogene, which may open new avenues for prevention and treatment of HCC.

Down-regulation of TRAF4 targeting RSK4 inhibits proliferation, invasion and metastasis in breast cancer xenografts.

Ribosomal S6 protein kinase 4 (RSK4) was known as a novel tumor suppressor gene, and the tumor necrosis factor receptor-associated factor 4 (TRAF4) was linked to carcinogenesis. The purpose of this study is to further investigate the effect of the TRAF4 gene on cell proliferation, invasion and metastasis in vivo and explore whether there is an interaction between TRAF4 and RSK4 in breast cancer. MDA-MB-231 cells were transfected with lentivirus TRAF4-shRNA to specifically block the expression of TRAF4, or transfected with lentivirus negative-shRNA as a negative control. Four-six weeks female BALB/c nude mice were randomly assigned to three groups (n = 14): TRAF4-shRNA, negative and control, and then inoculated subcutaneously with the corresponding cells. In-vivo metastasis model was constructed by injecting above cells into tail vein. Tumor proliferation was assessed in terms of the tumor growth curve, tumor size and weight. Invasion and metastasis were evaluated by the histopathologic examination in lung or/and liver tissues. Measurement of TRAF4 and RSK4 expression and their correlation factors (AKT, NF-κB, TGF-ß1, TNF-α, MMP2 and MMP9) were performed by immunohistochemistry, western blot or fluorescence quantitative RT-PCR. We found that the size and weight of tumors in TRAF4-shRNA group was significantly smaller than the negative and blank group, and the number of the lung and liver metastases lesions was also fewer (P < 0.05). And TRAF4 and its correlation factors (P-AKT, P-NF-κB, TGF-ß1, TNF-α, MMP2 and MMP9) in the TRAF4-shRNA group were significantly decreased compared with the negative and blank group. However, the expression of RSK4 mRNA and protein in TRAF4-shRNA group were significantly increased. Collectively, TRAF4 knockdown significantly inhibited proliferation, invasion and metastasis in the xenograft nude mouse model, possibly involving in the interaction with RSK4 through down-regulation of AKT signaling pathway and then inactivating NF-κB pathway.

Triisobutyl phosphate biodegradation by enriched activated sludge consortia: Degradation mechanism and bioaugmentation potential.

This study investigated the ability of activated sludge (AS) to biodegrade triisobutyl phosphate (TiBP) after acclimation in an AS bioreactor by adding 50 mg/L TiBP. The bioreactor significantly increased the biotransformation rate of TiBP (2.15-12.7 d-1) over two months of acclimation. Seven transformation products (TPs) of TiBP were identified by high-resolution mass spectrometry, and hydrolysis, hydroxylation and dehydrogenation were the major biodegradation pathways of TiBP. TiBP degradation solutions at 0, 3, 7, and 10 h showed significantly toxic effects on zebrafish embryos, while the toxicity of TiBP degradation solutions at 24 h significantly decreased. Pseudomonas was inferred to be a specific bacterial population in the TiBP metabolic microbial consortium (TMMC) that degrades TiBP (p < 0.001). When TMMC (0.5, 1, and 2 gss/L) was introduced into AS, the TiBP biotransformation rates (1.97, 2.05, and 2.26 d-1 at 1.0 mg/L TiBP, and 0.09, 0.11, and 0.83 d-1 at 30.0 mg/L TiBP) were significantly enhanced compared to the control (0.31 and 0.07 d-1) without TMMC inoculation. In general, this study provides new insights into the key species populations that accelerate TiBP degradation and promote the development of TiBP reduction biotechnology in WWTPs.

Aerobic degradation of parent triisobutyl phosphate and its metabolite diisobutyl phosphate in activated sludge: Degradation pathways and degrading bacteria.

Although organophosphate esters (OPEs) degradation has been widely studied, the degradation of their metabolites is always ignored. Triisobutyl phosphate (TiBP), a typical alkyl-OPEs, is of emerging concern because of its potential ecotoxicity in the environment. This study provides comprehensive understanding about the degradation of TiBP and one of its metabolites, diisobutyl phosphate (DiBP) using activated sludge (AS). The results showed that TiBP and DiBP were degraded mainly through hydrolysis, dehydrogenation, and hydroxylation. The degradation kinetics indicated that DiBP had similar transformation rates to its parent TiBP in AS, highlighting the importance of metabolite DiBP study. Dehydrogenase, hydroxylase, phosphotriesterase, phosphodiesterase, and phosphomonoesterase played an important role in contributing to TiBP and its metabolites degradation via enzyme activity analysis. Besides, the expression of genes encoding these enzymes in bacteria and the relative abundance change of bacterial populations indicated that Sphingomonas and Pseudomonas may be the degrading bacteria of TiBP and Pseudomonas may be the main degrading bacteria of DiBP. This study provides new perspectives for metabolite DiBP and its parent TiBP degradation. It highlights that the formation and degradation of metabolites must be considered into the future researches.

Exosomal miR-552-5p promotes tumorigenesis and disease progression via the PTEN/TOB1 axis in gastric cancer.

Purpose Gastric cancer (GC) is associated with rapid disease progression and poor patient prognosis, highlighting the pressing need for new biomarkers to facilitate disease management. Exosomes are released by all cells and are ubiquitous in body fluids, thus giving them great potential as diagnostic biomarkers and therapeutic targets. MicroRNAs (miRNAs) can be transported by exosomes, and are a common target for regulation in cancer. Methods Our screen of miRNAs in the Gene Expression Omnibus and The Cancer Genome Atlas databases identified miR-552-5p as the most overexpressed miRNA in GC, and we investigated its function and mechanism of action. Results We detected high expression of miR-552-5p in GC tissues, plasma samples and cell lines. We found that miR-552-5p binds directly to the 3'-untranslated region of PTEN, and the resulting downregulation of PTEN in turn downregulates the tumor suppressor TOB1. Furthermore, experiments in cell culture and mice showed that miR-552-5p in exosomes is internalized by recipient cells, where it enhances proliferation, migration and the epithelial-mesenchymal transition, while suppressing the caspase-3 apoptotic pathway. These effects were reversed by inhibiting miR-552-5p. Conclusion GC-derived exosomal miR-552-5p facilitates tumorigenesis by interfering with the PTEN/TOB1 axis, providing new potential therapeutic targets.

Silencing ROCK1 ameliorates ventilator-induced lung injury in mice by inhibiting macrophages' NLRP3 signaling.

Rho kinase, including two subtypes, ROCK1 and ROCK2, controls a variety of biological processes helping coordinate the tissues response to stress and injury. Some authors believe that alveolar macrophages (AMs) play a key role in the early phase of ventilator-induced lung injury (VILI), which is closely related to the activation of NLRP3 inflammasome and NF-κB signaling. However, there is currently little known about the relationship between ROCK signaling and NLRP3 inflammasome. Accordingly, we focused on exploring the effect of ROCK for NLRP3 inflammasome, the results showed that VILI in C57BL/6 mice significantly increased NF-κB, NLRP3, ASC, caspase1 expression, and the secretion of cytokines, which was reversed by applying the ROCK Inhibitor-Y27632. Moreover, the use of AMs and mechanical stretching suggested that ROCK regulated transcriptional level of NF-κB and NLRP3 inflammasome in AMs. Specifically, we silenced the ROCK1 and ROCK2 respectively, and found that the inflammation of MH-S cells after LPS and ATP priming could be regulated by ROCK1 and ROCK2, while the NLRP3 was only dependent upon ROCK1. Meantime, the related genes of NLRP3 signal are also regulated by ROCK1. Collectively, our data suggest that silencing ROCK1 ameliorates VILI in mice in part by inhibiting AMs' NLRP3 signaling pathway.

Ear ischemia induced by endovascular therapy for arteriovenous fistula of the sigmoid sinus: A case report.

BACKGROUND: Arteriovenous fistula of the sigmoid sinus is an abnormal connection of arteries with the sigmoid sinus. Endovascular treatments of such lesions are considered safe and with low rates of complications. CASE SUMMARY: A 62-year-old female patient underwent endovascular treatment of an arteriovenous fistula of the right sigmoid sinus on February 7, 2017, but her tinnitus was not cured. She was admitted to the Beijing Tiantan Hospital, Capital Medical University, on March 20, 2017, and her pre-operative diagnosis, by digital subtraction cerebral angiography, was arteriovenous fistula of the sigmoid sinus. She underwent endovascular embolization of the distal occipital artery and posterior auricular artery using Onyx-18. The arteriovenous fistula of the sigmoid sinus was cured, and her tinnitus disappeared, but ischemia of the upper 2/3 of the right auricle occurred without hearing loss. The patient received treatment to improve microcirculation, in addition to fluid supplementation, analgesia, and hyperbaric oxygen, and the swelling due to ischemia in the right auricle did not progress further. The patient reported no tinnitus , and the right auricle had returned to normal 3 years later. CONCLUSION: Ischemic complications of vital organs should be considered when performing embolization procedures for arteriovenous fistulas of cerebral sinuses. Compensation of the organs should be evaluated before the operation, and the related treatment regimens should be planned.

Morphine may act via DDX49 to inhibit hepatocellular carcinoma cell growth.

Pain in hepatocellular carcinoma (HCC) is a frequent cause of low quality of life, and morphine is routinely used as a first-line opiate analgesic in HCC. Morphine may exert not only analgesic effects but also anti-cancer effects via unknown mechanisms. Here we show that morphine can inhibit HCC cell proliferation. We further show that DEAD-box helicase 49 (DDX49) is up-regulated in HCC tumors, and that knocking down the DDX49 gene decreases tumor formation in vivo and in vitro, as well as reduces tumor metastasis in vivo. Morphine decreases DDX49 expression in HCC cells. Our results suggest that DDX49 contributes to HCC, and that morphine may exert anti-cancer effects by down-regulating it.

Ly-6Chigh inflammatory-monocyte recruitment is regulated by p38 MAPK/MCP-1 activation and promotes ventilator-induced lung injury.

Lymphocyte antigen 6Chigh (Ly-6Chigh) inflammatory monocytes, as novel mononuclear cells in the innate immune system, participate in infectious diseases. In this study, we investigated the potential role of these monocytes in ventilator-induced lung injury (VILI) and the possible mechanism involved in their migration to lung tissue. Our results showed that mechanical ventilation with high tidal volume (HTV) increased the accumulation of Ly-6Chigh inflammatory monocytes in lung tissues and that blocking C­C chemokine receptor 2 (CCR2) could significantly reduce Ly-6Chigh inflammatory-monocyte migration and attenuate the degree of inflammation of lung tissues. In addition, inhibition of p38 mitogen-activated protein kinase (p38 MAPK) activity could decrease the secretion of monocyte chemoattractant protein 1 (MCP-1), which in turn decreased the migration of Ly-6Chigh inflammatory monocytes into lung tissue. We also demonstrated that high ventilation caused Ly-6Chigh inflammatory monocytes in the bone marrow to migrate into and aggregate in the lungs, creating inflammation, and that the mechanism was quite different from that of infectious diseases. Ly-6Chigh inflammatory monocytes might play a pro-inflammatory role in VILI, and blocking their infiltration into lung tissue might become a new target for the treatment of this injury.

Mitophagy-Mediated mtDNA Release Aggravates Stretching-Induced Inflammation and Lung Epithelial Cell Injury via the TLR9/MyD88/NF-κB Pathway.

BACKGROUND: In animal models of ventilation-induced lung injury, mitophagy triggers mitochondria damage and the release of mitochondrial (mt) DNA, which activates inflammation. However, the mechanism of this process is unclear. METHODS: A model of cyclic stretching (CS)-induced lung epithelial cell injury was established. The genetic intervention of phosphatase and tensin homolog-induced kinase 1 (PINK1) expression via lentivirus transfection was used to identify the relationship between PINK1-mediated mitophagy and mtDNA release in stretching-induced inflammatory response and injury. Pharmacological inhabitation of Toll-like receptor 9 (TLR9) and myeloid differentiation factor 88 (MyD88) expression was performed via their related inhibitors, while pre-treatment of exogenous mtDNA was used to verify the role of mtDNA in stretching-induced inflammatory response and injury. RESULTS: Using a cell culture model of CS, we found that knocking down PINK1 in lung epithelial cells reduced mitophagy activation and mtDNA release, leading to milder inflammatory response and injury; conversely, up-regulating PINK1 exacerbated stretching-induced inflammation and injury, and similar effects were observed by upregulating TLR9 to induce expression of MyD88 and nuclear factor-κB (NF-κB)/p65. Down-regulating MyD88 protected lung epithelial cells from stretching injury and decreased NF-κB/p65 expression. CONCLUSION: These findings suggest that PINK1-dependent mitophagy and associated TLR9 activation is indeed a major factor in stretch-induced cell injury via a mechanism in which released mtDNA activates TLR9 and thereby the MyD88/NF-κB pathway. Inhibiting this process may be a therapeutic approach to prevent inflammation and cell injury in patients on mechanical ventilation.

[Effect of high tidal volume mechanical ventilation on pulmonary autophagy and mitochondrial damage in rats].

OBJECTIVE: To investigate the relationship between different tidal volume (VT) mechanical ventilation (MV) and autophagy and mitochondrial damage in rats. METHODS: A total of 120 clean-grade male Sprague-Dawley (SD) rats were divided into five groups (n = 24) by random number table method, and then given 0 (spontaneous breathing), 10, 20, 30, 40 mL/kg VT for MV. The rats in each group were subdivided into four subgroups of 1, 2, 3, and 4 hours according to ventilation time, with 6 rats in each subgroup. The lung tissue and bronchoalveolar lavage fluid (BALF) were harvested, and alveolar macrophages (AMs) and type II alveolar epithelial cells (AEC II) were cultured in vitro. The mRNA and protein expressions of autophagy-associated protein microtubule-associated protein 1 light chain 3B-II (LC3B-II) and autophagy-related genes Beclin1 and p62 were determined by reverse transcription-polymerase chain reaction (RT-PCR) or Western Blot. Lung autophagosome formation was observed under transmission electron microscope. The levels of adenosine triphosphate (ATP), reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in lung tissue were determined for assessing mitochondrial damage. RESULTS: There were no significant differences in the mRNA and protein expressions of LC3B-II, p62 and Beclin1 at 1 hour after ventilation among the groups. With the prolonged ventilation time, the mRNA and protein expressions of LC3B-II, p62 and Beclin1 in MV groups were increased gradually, peaked at 2-3 hours, and they were increased significantly in 30 mL/kg VT group as compared with those in spontaneous respiration group with statistical significances [ventilation for 2 hours: LC3B-II mRNA (2-ΔΔCt) was 2.44±0.24 vs. 1.12±0.04, LC3B-II/LC3B-I was 1.42±0.16 vs. 0.57±0.03, p62 mRNA (2-ΔΔCt) was 2.96±0.14 vs. 1.14±0.02, Beclin1 mRNA (2-ΔΔCt) was 2.80±0.13 vs. 1.14±0.02; ventilation for 3 hours: p62/ß-actin was 1.14±0.15 vs. 0.55±0.04, Beclin1/ß-actin was 1.27±0.06 vs. 0.87±0.04, all P < 0.05]. Autophagosomes and autolysosomes were found in AEC II after ventilation for 2 hours at 30 mL/kg VT by transmission electron microscopy, but not in AEC I. Compared with spontaneous breathing group, ATP synthesis in AMs was significantly decreased at 2 hours of ventilation in 30 mL/kg VT group (A value: 0.82±0.05 vs. 1.00±0.00, P < 0.05), ROS accumulate in AMs and AEC II were significantly increased [ROS in AMs: (33.83±4.00)% vs. (6.90±0.62)%, ROS in AEC II: (80.68±0.90)% vs. (2.16±0.19)%, both P < 0.05]. With the increase in VT and the prolongation of ventilation time, ATP and ROS levels in AMs and AEC II were gradually decreased, the ATP (A value) in AMs at 4 hours of ventilation in 40 mL/kg VT group was 0.41±0.05, the ROS in AMs was (12.95±0.88)%, and the ROS in AEC II was (40.43±2.29)%. With the increase in VT and the prolongation of ventilation time, MMP levels were gradually increased, the MMP (green/red fluorescence intensity ratio) in AMs at 2 hours of ventilation in 30 mL/kg VT group was 1.11±0.17, the MMP in AEC II was 0.96±0.04, and the MMP (green/red fluorescence intensity ratio) at 4 hours of ventilation in 40 mL/kg VT group was 0.51±0.07 and 0.49±0.06, respectively. CONCLUSIONS: The MV with high VT could induce autophagy activation and mitochondrial damage in lung tissue of rats, and the longer the ventilation time, the more obvious autophagy in the lung.

RhoA inhibitor suppresses the production of microvesicles and rescues high ventilation induced lung injury.

Microvesicles (MVs) have been extensively identified in various biological fluids including bronchoalveolar lavage fluid (BALF), peripheral blood and ascitic fluids. Our previous study showed that MVs are responsible for acute lung injury, but the exact mechanism underlying MVs formation remains poorly understood. In the present study, we investigate the potential role of RhoA/Rock signaling in MVs generation and the biological activity of MVs in ventilator-induced lung injury (VILI). Our results revealed that high tide ventilation induced super MVs releasing into the lung and subsequently caused lung inflammation. Strikingly, intratracheal instillation of MVs that isolated from highly ventilated mice triggered significant lung inflammation in naive mice. The MVs production is strongly correlated with lung inflammation and the upregulation of RhoA, Rock and phospho-Limk (phosphorylation of Limk is the activated form). RhoA inhibitor decreased the expression of Rock and the phosphorylation of Limk, decreased MVs production and alleviated lung inflammation. Rock inhibitor also decreased the phosphorylation of Limk, decreased MVs production and alleviated lung inflammation. Our data demonstrated that the production of MVs requires RhoA/Rock signaling, and VILI might be potentially prevented by targeting RhoA/Rock signaling pathway.

Non-intubated anesthesia in patients undergoing video-assisted thoracoscopic surgery: A systematic review and meta-analysis.

INTRODUCTION: Non-intubated anesthesia (NIA) has been proposed for video-assisted thoracoscopic surgery (VATS), although how the benefit-to-risk of NIA compares to that of intubated general anesthesia (IGA) for certain types of patients remains unclear. Therefore, the aim of the present meta-analysis was to understand whether NIA or IGA may be more beneficial for patients undergoing VATS. METHODS: A systematic search of Cochrane Library, Pubmed and Embase databases from 1968 to April 2019 was performed using predefined criteria. Studies comparing the effects of NIA or IGA for adult VATS patients were considered. The primary outcome measure was hospital stay. Pooled data were meta-analyzed using a random-effects model to determine the standard mean difference (SMD) with 95% confidence intervals (CI). RESULTS AND DISCUSSION: Twenty-eight studies with 2929 patients were included. The median age of participants was 56.8 years (range 21.9-76.4) and 1802 (61.5%) were male. Compared to IGA, NIA was associated with shorter hospital stay (SMD -0.57 days, 95%CI -0.78 to -0.36), lower estimated cost for hospitalization (SMD -2.83 US, 95% CI -4.33 to -1.34), shorter chest tube duration (SMD -0.32 days, 95% CI -0.47 to -0.17), and shorter postoperative fasting time (SMD, -2.76 days; 95% CI -2.98 to -2.54). NIA patients showed higher levels of total lymphocytes and natural killer cells and higher T helper/T suppressor cell ratio, but lower levels of interleukin (IL)-6, IL-8 and C-reactive protein (CRP). Moreover, NIA patients showed lower levels of fibrinogen, cortisol, procalcitonin and epinephrine. CONCLUSIONS: NIA enhances the recovery from VATS through attenuation of stress and inflammatory responses and stimulation of cellular immune function.

Microvesicles packaging IL-1ß and TNF-α enhance lung inflammatory response to mechanical ventilation in part by induction of cofilin signaling.

Microvesicles shed from pulmonary cells are capable of transferring inflammatory cargo to recipient cells nearby or in distant to enhance inflammation. Some authors believe that cofilin controls actin dynamics and regulates vesicle mobilization. We therefore investigated the potential role and mechanism of microvesicles in ventilator-induced lung injury (VILI). Fifty male C57BL/6 mice were orotracheally intubated and either allowed to breathe spontaneously or they were mechanically ventilated with different tidal volumes (Vt) and ventilation times. Lung tissue injury was assessed in terms of lung histopathologic examination, wet/dry weight ratios, and levels of total proteins and of cytokines. Microvesicle characteristics, sizes, contents and levels as well as cofilin were also measured. We found that lung inflammation increased significantly after ventilation with high Vt for 4 h; these conditions led to secretion of larger and more microvesicles into the alveoli than animals with/without ventilation at low Vt. Intratracheal instillation of microvesicles obtained from animals ventilated with low or high Vt triggered significant lung inflammation in naive mice, and these high-Vt microvesicles not only carried more IL-1ß and TNF-α but also induced more severe lung inflammation compared to low-Vt microvesicles; And high-Vt microvesicles at 2 h carried more molecular cargo than that at 1 h or 4 h, which may involve the shift and amplification of inflammation. Furthermore, blocking the phosphorylation of cofilin can not only inhibit microvesicle formation in the lung, but also reduce lung injury. Collectively, our data suggest that microvesicles packaging IL-1ß and TNF-α enhance lung inflammation in VILI.