LncRNA AP000695.2 promotes glycolysis of lung adenocarcinoma via the miR-335-3p/TEAD1 axis.

AP000695.2 is a novel long non-coding RNA (lncRNA). Its aberrant high expression is remarkably associated with poor prognosis of patients with lung adenocarcinoma (LUAD). However, its role and underlying mechanism in LUAD remains unclear. Previous bioinformatics analysis indicated that AP000695.2 may be closely related to the glycolysis of LUAD. This study aims to verify and explore the mechanism of AP000695.2 in glycolysis of LUAD. Overexpression plasmid and siRNA are used to construct cell models of upregulation and downregulation of AP000695.2, respectively. AP000695.2 is highly expressed in lung cancer cell lines as revealed by qPCR. Western blot analysis, FDG uptake, lactate production assay and ECAR determination results show that high expression of AP000695.2 facilitates glycolysis of LUAD cells. CCK-8, EdU staining, Transwell and wound healing assays show that high expression of AP000695.2 promotes cell growth and migration of LUAD. The relationship between AP000695.2 and miR-335-3p is confirmed by bioinformatics analysis and dual-luciferase reporter assays. Through the dual-luciferase reporter assay, TEA domain transcription factor 1 (TEAD1) is identified as a target gene of miR-335-3p. Rescue experiments are applied to verify the relationship among AP000695.2, miR-335-3p and TEAD1. Our study indicates that AP000695.2 is involved in the mechanism of LUAD through functioning as a ceRNA to competitively sponge miR-335-3p, thereby regulating the expression of TEAD1. In the in vivo models, AP000695.2 depletion restrains tumor growth and glycolysis. AP000695.2 promotes the glycolysis of LUAD by regulating the miR-335-3p/TEAD1 axis, and it may serve as a potential target of anti-tumor energy metabolism therapy.

An integrated multi-omics approach revealed the regulation of melatonin on age-dependent mitochondrial function impair and lipid dyshomeostasis in mice hippocampus.

Melatonin can improve mitochondrial dysfunction associated with the aging process by removing active oxygen, as well as inhibiting lipid peroxidation to maintain biofilm fluidity and resist free radical attack. However, there is poor understanding of the effect of melatonin on age-dependent mitochondrial function and lipid profile changes in brain. In this study, we investigated the energy metabolism of the whole body and brain of mice at 9 months, 13 months, and 25 months of continuous gastric administration of 3 mg/kg/d melatonin once per day morning for two months. In addition, we performed transcriptomic, proteomic and lipidomic analysis in the hippocampus of mice at different ages. Proteomics showed that melatonin regulated mitochondrial electron transport and leucine degradation in mouse hippocampus. Lipomics suggested that the long-chain unsaturated glycerol phospholipids in mouse hippocampus increased in an age-dependent manner, while ceramide and glycerol phospholipids decreased significantly in hippocampus of mouse chronically exposed to melatonin. The combined analysis of proteome and liposome demonstrated that Mpst, Ccsap, Hdhd5, Rpl5 and Flna were the key proteins of the network which involved in the regulation of numerous lipids. Furthermore, ultrastructure observation results illustrated that melatonin could improve the damaged mitochondrial and morphologies of 25-month-old mice hippocampus. In conclusion, we describe a mechanism that age-dependent up-regulation of long-chain unsaturated lipids is a driving risk factor for mitochondrial damage and this effect could be reversed by chronic supplement of low-dose melatonin.

Nicotine Activating α4ß2 Nicotinic Acetylcholine Receptors to Suppress Neuroinflammation via JAK2-STAT3 Signaling Pathway in Ischemic Rats and Inflammatory Cells.

Nicotine plays a role in inhibiting inflammatory factors, which contributes to improving cognitive impairment by activating α4ß2 nAChRs in ischemic rats, but the underlying mechanism has not been fully elucidated. Janus tyrosine kinase 2-signal transducer and activator of transcription 3 (JAK2-STAT3) signaling pathway is involved in cognitive improvement, and there seems to be a relationship between nAChRs and JAK2-STAT3 as well. The aim of this study is to explore the role of JAK2-STAT3 signaling pathway in nicotine-mediated anti-inflammatory effect. Nicotine, DHßE (the strongest competitive antagonist of α4ß2 nAChRs), and AG490 (a specific JAK2-STAT3 blocker) were used to intervene and treat ischemic rats and HEK-293 T-hα4ß2 cells. The Morris water maze (MWM) test and 2-[18F]-A-85380 PET imaging were performed to detect the cognitive function and α4ß2 nAChRs density in ischemic rats. The results demonstrated that nicotine intervention increased the density of α4ß2 nAChRs and improved cognitive impairment, but this effect was blocked by AG490, and the receptors were still upregulated. Essentially, when the JAK2-STAT3 signaling pathway was blocked, nicotine could only upregulate the expression of α4ß2 nAChRs, but not improve the cognitive function. PCR and Western blot analysis further confirmed these results. The cell experiments also showed that nicotine could reduce inflammatory factors stimulated by LPS and upregulate the expression of pJAK2 and pSTAT3 in HEK-293 T-hα4ß2 cells, while AG490 and DHßE reversed the effect of nicotine. To sum up, our work indicated that JAK2-STAT3 signaling pathway played an important role in nicotine-induced cognitive improvement by upregulating α4ß2 nAChRs in ischemic rats.

Targeted inhibition of glutamine metabolism enhances the antitumor effect of selumetinib in KRAS-mutant NSCLC.

Regulated by the tumor microenvironment, the metabolic network of the tumor is reprogrammed, driven by oncogenes and tumor suppressor genes. The metabolic phenotype of tumors of different driven-genes and different tissue types is extremely heterogeneous. KRAS-mutant non-small cell lung cancer (NSCLC) has glutamine dependence. In this study, we demonstrated that glutamine utilization of KRAS-mutant NSCLC was higher than that of KRAS wild-type. CB839, an efficient glutaminase inhibitor, synergized with the MEK inhibitor selumetinib to enhance antitumor activity in KRAS-mutant NSCLC cells and xenografts, and the therapeutic response could be well identified by 18F-FDG PET imaging. Combination therapy induced redox stress, manifesting as a decrease in mitochondrial membrane potential and an increase in ROS levels, and energetic stress manifesting as suppression of glycolysis and glutamine degradation. The phosphorylation of AKT was also suppressed. These effects combined to induce autophagy and thereby caused cancer cell death. Our results suggest that dual inhibition of the MEK-ERK pathway and glutamine metabolism activated by KRAS mutation may be an effective treatment strategy for KRAS-driven NSCLC.

PD-L1 in Lung Adenocarcinoma: Insights into the Role of 18F-FDG PET/CT.

PURPOSE: This study aimed to evaluate the role of 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography (PET)/computed tomography (CT) in expression of tumor programmed death ligand-1 (PD-L1) expression and prognostic significance of 18F-FDG PET/CT at different PD-L1 status in patients with lung adenocarcinoma. PATIENTS AND METHODS: Seventy-three patients with primary lung adenocarcinoma who received 18F-FDG PET/CT before treatment were retrospectively included in this study. Expression of tumor PD-L1, programmed death-1 (PD-1) and glucose metabolic parameters were evaluated. RESULTS: Tumor PD-L1 expression was positively correlated with maximum standardized uptake value (SUVmax), total lesion glycolysis (TLG), hexokinase II (HK-II) and glucose transporter 1 (GLUT-1) (P<0.0001 for all). SUVmax was a unique independent predictor of tumor PD-L1 expression, with an optimal cut-off value of 9.5. For all the patients, tumor stage (P<0.001) and SUVmax (P=0.009) were independent prognostic indicators of disease-free survival (DFS)/progression-free survival (PFS) while carcino-embryonic antigen (CEA) (P=0.003), Ki67 (P=0.042), PD-L1 (P=0.048) and TLG (P=0.004) were independent prognostic indicators of overall survival (OS). Tumor stage (P=0.004) and SUVmax (P=0.022) were independent prognostic indicators of DFS/PFS while TLG (P=0.012) and CEA (P=0.045) were independent prognostic indicators of OS in the PD-L1-positive group. In the PD-L1-negative group, tumor stage (P=0.002) and CEA (P=0.006) were unique independent prognostic indicators of DFS/PFS and OS, respectively. CONCLUSION: 18F-FDG PET/CT may potentially predict tumor PD-L1 expression and play a role in predicting prognosis of PD-L1/PD-1 immunotherapy in lung adenocarcinoma.

Nicotine Induced Neurocognitive Protection and Anti-inflammation Effect by Activating α 4ß 2 Nicotinic Acetylcholine Receptors in Ischemic Rats.

INTRODUCTION: The main objective of this study was to explore the mechanism of nicotine improving cognitive impairments in ischemic rats. METHODS: Twenty adult male Sprague-Dawley (SD) rats underwent ischemic model surgery by injecting endothelin-1 into the left thalamus, which were classified into four different groups with different intervention: nicotine (1.5 mg/kg/d), dihydro-ß-erythroidine (DHßE; 3 mg/kg/d), nicotine (1.5 mg/kg/d) + DHßE (3 mg/kg/d), or saline, after ischemic model surgery. Another five male SD rats also underwent same surgery, while not injecting endothelin-1 but saline, as the control group. Morris water maze (MWM) test was adopted to assess the cognition. All the rats underwent the MWM test, micro positron emission tomography imaging with 2-[18F]-A-85380, and messenger RNA (mRNA) test of α 4 nicotinic acetylcholine receptor (nAChR), ß 2 nAChR, tumor necrosis factor-alpha (TNF-α), IL-1ß, and IL-6. RESULTS: The MWM test showed the rats given nicotine showing better memory than ischemic rats (p < .05), whereas the rats given DHßE or both nicotine and DHßE did not show any statistical difference from the ischemic rats (p > .05). Micro positron emission tomography imaging showed higher uptake of tracer in the left thalamus and whole brain in rats given nicotine than in ischemic rats, but the rats given DHßE or both nicotine and DHßE did not. By real-time PCR test, the mRNA of α 4 nAChR and ß 2 nAChR in rats given nicotine increased significantly compared with ischemic rats and decreased TNF-α, IL-1ß, and IL-6 mRNA (all ps < .05). CONCLUSIONS: By activating α 4ß 2 nAChRs, nicotine plays a role in inhibiting the inflammatory factors, which contributes to improving cognitive impairment in ischemic rats. IMPLICATIONS: It is well acknowledged that vascular cognitive impairment (VCI) is the second most common cause of dementia after Alzheimer's disease. Cholinergic agents have potential for the symptomatic treatment of the cognitive symptoms of dementia, but the exact mechanism still remains unclear. There are potential complex associations and interactions between VCI and inflammation. This study showed that nicotine had anti-inflammatory potency, which is most likely because of the activation of the nAChRs. By activating α4ß2 nAChRs, nicotine played a role in inhibiting the inflammatory factors, which contribute to improving cognitive impairment in ischemic rats.

Effect of IDH3a on glucose uptake in lung adenocarcinoma: A pilot study based on [18 F]FDG.

Subunit of isocitrate dehydrogenase 3 (IDH3a) as upstream of the hypoxia-inducible factor was reported highly expressed in malignant tumors, playing an important role in glucose metabolism reprogramming. As one of rate-limiting enzyme in the Krebs cycle, whether high expression of IDH3a affects glucose uptake in tumors has not been elucidated. This study was aimed to investigate the relationship between IDH3a expression and tumor glucose uptake. Sixty-five patients who underwent 2-[18 F]-2-deoxy-D-glucose ([18 F]-FDG) positron emission tomography/computed tomography (PET/CT) imaging before surgery and pathologically diagnosed as lung adenocarcinoma were included. All patients were divided into high (n = 31) and low (n = 34) groups according IDH3a expression by immunohistochemistry. Comparatively higher [18 F]-FDG uptake was found in high IDH3a expression group. Glucose transporter 1 (GLUT1) level was demonstrated to correlate with IDH3a expression, but not for hexokinase 2 (HK2). Furthermore, A549 and H1299 cells experiment showed, the expression of p-AKT and GLUT1 were significantly downregulated after IDH3a interference. The cellular uptake of [18 F]-FDG and lactate production were significantly reduced in treatment group. In summary, high expression of IDH3a in lung adenocarcinoma patients is associated with higher glucose uptake. IDH3a targets AKT-GLUT1 pathway to affect glucose uptake and metabolites in lung adenocarcinoma.

ATAD2 expression increases [18F]Fluorodeoxyglucose uptake value in lung adenocarcinoma via AKT-GLUT1/HK2 pathway.

[18F]Fluorodeoxyglucose (FDG) PET/CT imaging has been widely used in the diagnosis of malignant tumors. ATPase family AAA domain-containing protein 2 (ATAD2) plays important roles in tumor growth, invasion and metastasis. However, the relationship between [18F]FDG accumulation and ATAD2 expression remains largely unknown. This study aimed to investigate the correlation between ATAD2 expression and [18F]FDG uptake in lung adenocarcinoma (LUAD), and elucidate its underlying molecular mechanisms. The results showed that ATAD2 expression was positively correlated with maximum standardized uptake value (SUVmax), total lesion glycolysis (TLG), glucose transporter type 1 (GLUT1) expression and hexokinase2 (HK2) expression in LUAD tissues. In addition, ATAD2 knockdown significantly inhibited the proliferation, tumorigenicity, migration, [18F]FDG uptake and lactate production of LUAD cells, while, ATAD2 overexpression exhibited the opposite effects. Furthermore, ATAD2 modulated the glycometabolism of LUAD via AKT-GLUT1/HK2 pathway, as assessed using LY294002 (an inhibitor of PI3K/AKT pathway). In summary, to explore the correlation between ATAD2 expression and glycometabolism is expected to bring good news for anti-energy metabolism therapy of cancers. [BMB Reports 2019; 52(7): 457-462].

Long noncoding RNA H19 participates in metformin-mediated inhibition of gastric cancer cell invasion.

Recent research suggests that the first-line oral antidiabetes drug metformin may prevent gastric cancer progression and improve prognosis. Many studies have also shown that long noncoding RNAs (lncRNAs) play important roles in many biological processes. Therefore, we aimed to explore whether lncRNAs participate in the mechanisms by which metformin affects gastric cancer cells. In the current study, we found that metformin significantly inhibited the cellular functions of gastric cancer cells through Cell Counting Kit-8 and invasion assays. We found that lncRNA H19 was greatly downregulated in gastric cancer cells treated with metformin using lncRNA microassays. Based on bioinformatics analyses of the Oncomine and The Cancer Genome Atlas databases, H19 is shown to be overexpressed in gastric cancer tissues, with increased expression of H19 relating to advanced pathological tumor stage and pathological tumor node metastasis stage, indicating that H19 may be associated with the invasive ability of gastric cancer. We knocked down H19 in AGS and SGC7901 cell lines and found that knocked-down H19 could decrease gastric cancer cell invasion and that metformin could not further decrease invasion after the knock down. Moreover, H19 depletion increased AMPK activation and decreased MMP9 expression, and metformin could not further activate AMPK or decrease MMP9 in H19 knocked-down gastric cancer cells. In summary, metformin has a profound antitumor effect on gastric cancer cells, and H19 is a key component in the process of metformin suppressing gastric cancer cell invasion.

Immobility responses between mouse strains correlate with distinct hippocampal serotonin transporter protein expression and function.

Mouse strain differences in immobility and in sensitivity to antidepressants have been observed in the forced swimming test (FST) and the tail suspension test (TST). However, the neurotransmitter systems and neural substrates that contribute to these differences remain unknown. To investigate the role of the hippocampal serotonin transporter (5-HTT), we measured baseline immobility and the immobility responses to fluoxetine (FLX) in the FST and the TST in male CD-1, C57BL/6, DBA and BALB/c mice. We observed strain differences in baseline immobility time, with CD-1 mice showing the longest and DBA mice showing the shortest. In contrast, DBA and BALB/c mice showed the highest sensitivity to FLX, whereas CD-1 and C57BL/6 mice showed the lowest sensitivity. Also we found strain differences in both the total 5-HTT protein level and the membrane-bound 5-HTT level (estimated by V max) as follows: DBA>BALB/c>CD-1=C57BL/6. The uptake efficiency of the membrane-bound 5-HTT (estimated by 1/K m) was highest in DBA and BALB/c mice and lowest in CD-1 and C57BL/6 mice. A correlation analysis of subregions within the hippocampus revealed that immobility time was negatively correlated with V max and positively correlated with K m in the hippocampus. Therefore a higher uptake capacity of the membrane-bound 5-HTT in the hippocampus was associated with lower baseline immobility and greater sensitivity to FLX. These results suggest that alterations in hippocampal 5-HTT activity may contribute to mouse strain differences in the FST and the TST.

[Relationship between DEK oncogene expression and oral squamous cell carcinoma].

PURPOSE: To investigate the expression of gene DEK in oral squamous cell carcinoma, and explore the role of DEK in the development and metastasis of oral squamous cell carcinoma. METHODS: Thirty-three specimens of oral squamous cell carcinoma (OSCC), 28 specimens of cervical lymph nodes metastasis, and 33 specimens of normal oral mucosal tissues adjacent to the cancerous lesions were examined. Polymerase chain reaction with reverse transcription (RT-PCR) and immunohistochemistry were used to detect mRNA and protein expression of DEK, respectively.The results were analysed with SPSS17.0 software package. RUSULTS: RT-PCR and immunohistochemistry showed that the expression of DEK mRNA and DEK protein in metastatic lymph nodes (82.1%, 82.1%) and cancerous tissues (81.2%, 76.8%) of OSCC were significantly higher than in the adjacent tissues (27.3%, 18.2%). The difference between cancerous tissues and adjacent tissues was significant (P<0.05). CONCLUSION: DEK may play an important role in the occurrence and metastasis of OSCC.

Subregion-specific decreases in hippocampal serotonin transporter protein expression and function associated with endophenotypes of depression.

Stress influences the development of depression, and depression is associated with structural and functional changes in the hippocampus. The current study sought to determine whether chronic corticosteroid (CORT) treatment influences serotonin transporter (5-HTT) protein expression and function in the CA1, CA3, and dentate gyrus (DG) subregions of the hippocampus. Male CD-1 mice were subcutaneously injected with CORT at a dose of 20 mg/kg once daily for 3 weeks. Behavioral state was assessed using sucrose preference, physical state of the coat, forced swimming test, and tail suspension test. We then determine 5-HTT protein expression and synaptosomal 5-HT uptake in the CA1, CA3 and DG subregions. CORT treatment induced anhedonia and behavioral despair, two core endophenotypes of clinical depression; 5-HTT protein expression levels and synaptosomal 5-HT uptake were both decreased in a subregion-specific manner, with the greatest decrease observed in the DG, a moderate decrease in the CA3, and the CA1 showed no apparent change. In addition, a reduction in tissue mass was detected in the DG following the CORT treatment. These data indicate that subregion-specific decreases in hippocampal 5-HTT protein expression and function are associated with endophenotypes of depression.

Anti-edema effect of epigallocatechin gallate on spinal cord injury in rats.

Recent studies indicated that epigallocatechin gallate (EGCG) had neuroprotective effects on spinal cord injury (SCI).The current study was performed to determine the anti-edema effect of EGCG after SCI in rats. EGCG (100 mg/kg, i.p.) was administered to rats immediately following SCI. It was found that EGCG (100 mg/kg) could significantly reduce spinal cord water content. In addition, EGCG (100mg/kg) significantly reduced the expression of aquaporin-4(AQP4) and glial fibrillary acidic protein (GFAP) level at 24, 48 and 72h after injury, but it did not have this effect at 12 h after injury. The changes of AQP4 and GFAP protein induced by EGCG (100 mg/kg) treatment were accompanied by a reduction of spinal cord edema. Our results indicated that EGCG (100 mg/kg) could reduce spinal cord edema after SCI, which could be correlated with the down-regulation the expression of AQP4 and GFAP protein level after SCI.

Mn (III) tetrakis (4-benzoic acid) porphyrin protects against neuronal and glial oxidative stress and death after spinal cord injury.

This study explores the ability of a catalytic antioxidant, Mn (III) tetrakis (4-benzoic acid) porphyrin (MnTBAP), to protect against neuronal and glial oxidative stress and death after spinal cord injury (SCI). Nine different doses of MnTBAP were administered into the intrathecal space of the rat spinal cord immediately following moderate SCI to establish dose - response curves for prevention of lipid peroxidation and neuron death. An optimal dose was determined by comparing the effectiveness of MnTBAP protection among doses. The optimal dose was then administered and the cords were removed 24 h post-administration and processed for staining. The cells in the cord sections at different distances from the epicenter were counted to obtain the spatial profiles of MnTBAP protection. Comparison of the counts between MnTBAP- and vehicle-treated groups in the sections double immuno-fluorescence-stained with oxidative and cellular markers demonstrated that MnTBAP significantly reduced numbers of nitrotyrosine- and DNP-positive (stained with an antibody against 2,4-dinitrophenyl hydrazine (DNPH)-labeled protein carbonyls) neurons, astrocytes, and oligodendrocytes. Comparison of the counts between the two treatments in the sections immuno-stained with cellular markers revealed that MnTBAP significantly increased numbers of neurons, motoneurons, astrocytes, and oligodendrocytes. MnTBAP more effectively reduced neuronal than glial cell death. Post-injury treatment with the optimal dose of MnTBAP at 6, 12, 24, 48, and 72 h post-SCI demonstrated that the effective time window for reducing protein nitration and neuron death was at least 12 h. Our results demonstrated that MnTBAP combats oxidative stress, thereby attenuating all types of cell death after SCI.