Effect of electroacupuncture stimulation of "Sibai"(ST2) and "Quanliao"(SI18) on excitatory neurons of sensorimotor cortex in mice. / 电针"四白"和"颧髎"æ¿€æ´»å°é¼ èº¯ä½“æ„Ÿè§‰è¿åŠ¨çš®å±‚çš„å ´å¥‹æ€§ç¥žç»å ƒ.

OBJECTIVES: To observe the activation state and neuronal types of somatosensory cortex and the primary motor cortex induced by electroacupuncture (EA) stimulation of "Sibai" (ST2) and "Quanliao" (SI18) acupoints in mice. METHODS: Male C57BL/6J mice were randomly divided into blank control and EA groups, with 6 mice in each group. Rats of the EA group received EA stimulation (2 Hz, 0.6 mA) at ST2 and SI18 for 30 minutes. Samples were collected after EA intervention, and immunofluorescence staining was performed to quantify the expression of the c-Fos gene (proportion of c-Fos positive cells) in the somatosensory cortex and primary motor cortex. The co-labelled cells of calcium/calmodulin-dependent protein kinase Ⅱ (CaMKⅡ) and gamma-aminobutyric acid (GABA) in the somatosensory cortex and primary motor cortex were observed and counted by using microscope after immunofluorescence staining. Another 10 mice were used to detect the calcium activity of excitatory neurons in the somatosensory cortex and primary motor cortex by fiber photometry. RESULTS: In comparison with the blank control group, the number of c-Fos positive cells, and the proportion of c-Fos and CaMKⅡ co-labelled cells in both the somatosensory cortex and primary motor cortex were significantly increased after EA stimulation (P<0.05). No significant changes were found in the proportion of c-Fos and GABA co-labeled cells in both the somatosensory cortex and primary motor cortex after EA. Results of fiber optic calcium imaging technology showed that the spontaneous calcium activity of excitatory neurons in both somatosensory cortex and primary motor cortex were obviously increased during EA compared with that before EA (P<0.01), and strikingly reduced after cessation of EA compared with that during EA (P<0.05). CONCLUSIONS: Under physiological conditions, EA of ST2 and SI18 can effectively activate excitatory neurons in the somatosensory cortex and primary motor cortex.

A more accurate indicator to evaluate oxidative stress in rat plasma with osteoporosis.

Background: oxidative stress is linked to various human diseases which developed into the idea of "disrupted redox signaling". Osteoporosis (OP) is a chronic skeletal disorder characterized by low bone mineral density and deterioration of bone microarchitecture among which estrogen deficiency is the main cause. Lack of estrogen leads to the imbalance between oxidation and anti-oxidation in patients, and oxidative stress is an important link in the pathogenesis of OP. The ratio of the reduced to the oxidized thiols can characterize the redox status. However, few methods have been reported for the simultaneous determination of reduced forms and their oxidized forms of thiols in plasma. Methods: we developed a hollow fiber centrifugal ultrafiltration (HFCF-UF) method for sample preparation and validated a high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method to determine two reduced forms of thiols-homocysteine (Hcy), cysteine (Cys) levels and their respective oxidized compounds, homocystine (HHcy) and cystine (Cyss) in rat plasma simultaneously for the first time. Thirty-six female rats were randomly divided into three groups: normal control (NC), oxidative stress (ovariectomy, OVX) and ovariectomy with hydrogen-rich saline administration (OVX + HRS). Results: the validation parameters for the methodological results were within the acceptance criteria. There were both significant differences of Hcy/HHcy (Hcy reduced/oxidized) and Cys/Cyss (Cys reduced/oxidized) in rat plasma between three groups with both p < 0.05 and meanwhile, the p values of malondialdehyde, superoxide dismutase and glutathione peroxidase were all less than 0.01. The value of both Hcy/HHcy and Cys/Cyss were significantly decreased with the change of Micro-CT scan result of femoral neck in OVX group (both the trabecular thickness and trabecular number significantly decreased with a significant increase of trabecular separation) which demonstrate OP occurs. The change of Hcy/HHcy is more obvious and prominent than Cys/Cyss. Conclusions: the Hcy/HHcy and Cys/Cyss could be suitable biomarkers for oxidative stress and especially Hcy/HHcy is more sensitive. The developed method is simple and accurate. It can be easily applied in clinical research to further evaluate the oxidative stress indicator for disease risk factors.

Identification of Gαi3 as a novel molecular therapeutic target of cervical cancer.

Here we studied expression and potential functions of Gαi3 in cervical cancer. The bioinformatics analysis together with the results from local patients' tissues revealed that Gαi3 expression was remarkably elevated in human cervical cancer tissues and different cervical cancer cells, and was associated with poor overall survival and poor disease-specific survival of patients. Gαi3 depletion resulted in profound anti-cervical cancer activity. In primary or immortalized cervical cancer cells, Gαi3 shRNA or CRISPR/Cas9-caused Gαi3 knockout/KO largely hindered cell proliferation and migration, and provoked apoptosis. On the contrast, ectopic Gαi3 overexpression further enhanced cervical cancer proliferation and migration. Akt-mTOR activation in primary cervical cancer cells was significantly reduced after Gαi3 silencing or KO, but was augmented following Gαi3 overexpression. Further studies revealed that the transcription factor GATA4 binding to Gαi3 promoter region was significantly enhanced in cervical cancer tissues and cells. Gαi3 expression was decreased by GATA4 shRNA, but upregulated following GATA4 overexpression. In vivo, the growth of cervical cancer xenografts was robustly suppressed after Gαi3 silencing or KO. Gαi3 depletion and Akt-mTOR inactivation were detected in Gαi3-silenced/-KO cervical cancer xenograft tissues. Together, upregulated Gαi3 is a valuable oncotarget of cervical cancer.

The sodium/myo-inositol co-transporter SLC5A3 promotes non-small cell lung cancer cell growth.

Identification of novel molecular signaling targets for non-small cell lung cancer (NSCLC) is important. The present study examined expression, functions and possible underlying mechanisms of the sodium/myo-inositol co-transporter SLC5A3 in NSCLC. The Cancer Genome Atlas (TCGA) database and local NSCLC tissue results demonstrated that SLC5A3 expression in NSCLC tissues (including patient-derived primary NSCLC cells) was significantly higher than that in normal lung tissues and lung epithelial cells. In primary NSCLC cells and immortalized lines, SLC5A3 depletion, using small hairpin RNA (shRNA) and CRSIRP/Cas9 methods, robustly impeded cell proliferation and migration, simultaneously provoking cell cycle arrest and apoptosis. Conversely, ectopic overexpression of SLC5A3 further enhanced proliferation and migration in primary NSCLC cells. The intracellular myo-inositol contents and Akt-mTOR activation were largely inhibited by SLC5A3 silencing or knockout (KO), but were augmented following SLC5A3 overexpression in primary NSCLC cells. Significantly, SLC5A3 KO-induced anti-NSCLC cell activity was largely ameliorated by exogenously adding myo-inositol or by a constitutively-active Akt construct. By employing the patient-derived xenograft (PDX) model, we found that the growth of subcutaneous NSCLC xenografts in nude mice was largely inhibited by intratumoral injection SLC5A3 shRNA adeno-associated virus (AAV). SLC5A3 silencing, myo-inositol depletion, Akt-mTOR inactivation and apoptosis induction were detected in SLC5A3 shRNA virus-injected NSCLC xenograft tissues. Together, elevated SLC5A3 promotes NSCLC cell growth possibly by maintaining myo-inositol contents and promoting Akt-mTOR activation.

The requirement of phosphoenolpyruvate carboxykinase 1 for angiogenesis in vitro and in vivo.

We here examined the potential biological function of phosphoenolpyruvate carboxykinase 1 (PCK1) in angiogenesis. shRNA- or CRISPR-Cas9-induced PCK1 depletion potently inhibited endothelial cell proliferation, migration, sprouting, and tube formation, whereas ectopic PCK1 overexpression exerted opposite activity. In HUVECs, Gαi3 expression and Akt activation were decreased following PCK1 depletion, but were augmented by ectopic PCK1 overexpression. In vivo, retinal expression of PCK1 gradually increased from postnatal day 1 (P1) to P5. The intravitreous injection of endothelial-specific PCK1 shRNA adenovirus at P1 potently inhibited the radial extension of vascular plexus at P5. Conditional endothelial knockdown of PCK1 in adult mouse retina increased vascular leakage and the number of acellular capillaries while decreasing the number of RGCs in murine retinas. In diabetic retinopathy patients, PCK1 mRNA and protein levels were up-regulated in retinal tissues. Together, PCK1 is essential for angiogenesis possibly by mediating Gαi3 expression and Akt activation.

The sphingosine kinase inhibitor SKI-V suppresses cervical cancer cell growth.

Overexpression and/or overactivation of sphingosine kinase 1/2 (SphK1/2) is important for tumorigenesis and progression of cervical cancer. The current study examined the potential activity and signaling mechanisms of SKI-V, a non-lipid small molecule SphK inhibitor, against cervical cancer cells. In different primary and immortalized cervical cancer cells, SKI-V exerted significant anti-cancer activity by inhibiting cell viability, colony formation, proliferation, cell cycle progression and cell migration. Significant apoptosis activation was detected in SKI-V-treated cervical cancer cells. Significantly, SKI-V also provoked programmed necrosis cascade in cervical cancer cells, as it induced mitochondrial p53-cyclophilin-D-adenine nucleotide translocator-1 (ANT1) complexation, mitochondrial membrane potential collapse, reactive oxygen species production and the release of lactate dehydrogenase into the medium. Further, SKI-V blocked SphK activation and induced ceramide accumulation in primary cervical cancer cells, without affecting SphK1/2 expression. SKI-V-induced cytotoxicity in cervical cancer cells was largely inhibited by sphingosine-1-phosphate or the SphK1 activator K6PC-5, but was sensitized by adding the short-chain ceramide C6. Moreover, SKI-V inhibited Akt-mTOR (mammalian target of rapamycin) activation in primary cervical cancer cells, and its cytotoxicity was mitigated by a constitutively-active Akt. In vivo, daily intraperitoneal injection of SKI-V significantly inhibited subcutaneous primary cervical cancer xenograft growth in nude mice. Together, the SphK inhibitor SKI-V suppresses cervical cancer growth in vitro and in vivo.

Pseudomonas aeruginosa heteroresistance to levofloxacin caused by upregulated expression of essential genes for DNA replication and repair.

Pseudomonas aeruginosa (P. aeruginosa), a common cause of severe chronic infections, has developed heteroresistance to several antibiotics, thus hindering successful treatment. In this study, we aimed to investigate the characteristics and mechanisms underlying levofloxacin (LVX) heteroresistance in P. aeruginosa PAS71 and PAS81 clinical isolates using a combination of physiological and biochemical methods, bacterial genomics, transcriptomics, and qRT-PCR. The six P. aeruginosa strains, namely PAS71, PAS72, PAS81, PAS82, ATCC27853, and PAO1, were studied. The Kirby-Bauer (K-B), minimum inhibitory concentration (MIC) test, and population analysis profile (PAP) experimental results showed that PAS71, PAS81, ATCC27853, and PAO1 were heteroresistant to LVX, with MIC of 0.25, 1, 0.5, and 2 µg/ml, respectively; PAS72 and PAS82 were susceptible to LVX with a MIC of 0.25 and 0.5 µg/ml, respectively. The resistance of PAS71 and PAS81 heteroresistant subpopulations was unstable and had a growth fitness cost. Genomic and transcriptomic results proved that the unstable heteroresistance of PAS71 and PAS81 was caused by elevated expression of essential genes involved in DNA replication and repair, and homologous recombination, rather than their genomic single-nucleotide polymorphism (SNP) and insertion-deletion (InDel) mutations. Additionally, PAS71 and PAS81 enhanced virulence and physiological metabolism, including bacterial secretion systems and biosynthesis of siderophore group nonribosomal peptides, in response to LVX stress. Our results suggest that the upregulation of key genes involved in DNA replication and repair, and homologous recombination causes unstable heteroresistance in P. aeruginosa against LVX. This finding provides novel insights into the occurrence and molecular regulation pathway of P. aeruginosa heteroresistant strains.

A simple and accurate HFCF-UF method for the analysis of homocysteine, cysteine, cysteinyl-glycine, and glutathione in human blood.

The presence of reduced aminothiols, including homocysteine (Hcy), cysteine (Cys), cysteinyl-glycine (CG), and glutathione (GSH), is significantly increased in the pathological state. However, there have been no reports on the relationship between reduced aminothiols (Hcy, Cys, CG, and GSH) and different genders, ages, and drug combinations in human blood. The accurate quantification of these reduced thiols in biological fluids is important for monitoring some special pathological conditions of humans. However, the published methods typically not only require cumbersome and technically challenging processing procedures to ensure reliable measurements, but are also laborious and time-consuming, which may disturb the initial physiological balance and lead to inaccurate results. We developed a hollow fiber centrifugal ultrafiltration (HFCF-UF) method for sample preparation coupled with a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method and used it to determine four reduced aminothiols (Hcy, Cys, CG, and GSH) in human blood for the first time. A total of 96 clinical patients were enrolled in our study. The influence of different genders, ages, and drug combinations on the levels of four reduced thiols in human blood was also discussed by SPSS 24.0. The sample preparation was simplified to a single 5 min centrifugation step in a sealed system that did not disturb the physiological environment. The validation parameters for the methodological results were excellent. The procedure was successfully applied to monitoring the concentrations of four reduced aminothiols (Hcy, Cys, CG, and GSH) in 96 clinical blood samples. There were no significant differences in Hcy, Cys, CG, or GSH for the different genders, ages, or combinations with methotrexate or vancomycin (P > 0.05). However, there was a significant increase in Hcy concentration in patients treated with valproic acid who were diagnosed with epilepsy (p=0.0007). It is advisable to measure reduced Hcy level in patients taking valproic acid. The developed HFCF-UF method was simple and accurate. It can be easily applied in clinical research to evaluate oxidative stress in further study.

Neuronal-driven glioma growth requires Gαi1 and Gαi3.

Neuroligin-3 (NLGN3) is necessary and sufficient to promote glioma cell growth. The recruitment of Gαi1/3 to the ligand-activated receptor tyrosine kinases (RTKs) is essential for mediating oncogenic signaling. Methods: Various genetic strategies were utilized to examine the requirement of Gαi1/3 in NLGN3-driven glioma cell growth. Results: NLGN3-induced Akt-mTORC1 and Erk activation was inhibited by decreasing Gαi1/3 expression. In contrast ectopic Gαi1/3 overexpression enhanced NLGN3-induced signaling. In glioma cells, NLGN3-induced cell growth, proliferation and migration were attenuated by Gαi1/3 depletion with shRNA, but facilitated with Gαi1/3 overexpression. Significantly, Gαi1/3 silencing inhibited orthotopic growth of patient-derived glioma xenografts in mouse brain, whereas forced Gαi1/3-overexpression in primary glioma xenografts significantly enhanced growth. The growth of brain-metastatic human lung cancer cells in mouse brain was largely inhibited with Gαi1/3 silencing. It was however expedited with ectopic Gαi1/3 overexpression. In human glioma Gαi3 upregulation was detected, correlating with poor prognosis. Conclusion: Gαi1/3 mediation of NLGN3-induced signaling is essential for neuronal-driven glioma growth.

Relationship Between the Free and Total Methotrexate Plasma Concentration in Children and Application to Predict the Toxicity of HD-MTX.

High-dose methotrexate (HD-MTX) can be highly effective as well as extremely toxic. Many drug molecules can bind to plasma proteins to different extents in vivo, whereas only the free drug can reach the site of action to exert a pharmacological effect and cause toxicity. However, free MTX concentrations in plasma have not been reported. Traditional analyses of free drugs are both cumbersome and inaccurate. We collected 92 plasma samples from 52 children diagnosed with ALL or NHL or other lymphomas that were treated with HD-MTX. The hollow fiber centrifugal ultrafiltration (HFCF-UF) was used to prepare plasma samples for analysis of the free MTX concentration. Protein precipitation was employed to measure the total MTX concentration. The HFCF-UF is a simple method involving a step of ordinary centrifugation; the validation parameters for the methodological results were satisfactory and fell within the acceptance criteria. A linearity coefficient r 2 of 0.910 was obtained for the correlation between the free and total MTX plasma concentrations in 92 plasma samples. However, the free and total MTX concentrations was only weakly correlated in 16 clinical plasma specimens with total MTX concentrations >2 µmol L-1 (r 2 = 0.760). Both the free and total MTX concentrations at 42 h were negatively correlated with the creatinine clearance (CCr) level (P = 0.023, r = -0.236 for total MTX and P = 0.020, r = -0.241for free MTX, respectively). The free MTX concentration could not be accurately estimated from the total MTX concentration for patients with high MTX levels which are conditions under which toxic reactions are more likely to occur. High plasma MTX levels could become a predictor of the occurrence of MTX nephrotoxicity to draw people's attention. The proposed HFCF-UF method is a simple and accurate way to evaluate efficacy and toxicity in clinical therapeutic drug monitoring.

LncRNA PINK1-AS promotes Gαi1-driven gastric cancer tumorigenesis by sponging microRNA-200a.

Gastric cancer (GC) is one of the leading causes of human mortality around the world. We have previously shown that Gαi1 (the inhibitory subunit 1 of the heterotrimeric guanine nucleotide-binding protein) recruitment to ligand-activated receptor tyrosine kinases (RTKs) is essential for signaling. Testing its role in GC cancer-promoting functions, we found that Gαi1 is upregulated in human GC, correlating with poor overall survival. In established and primary human GC cells, Gαi1 shRNA (small hairpin RNA) or knockout produced significant anti-GC cell activity, proliferation and migration was inhibited, and apoptosis was activated. Conversely, ectopic Gαi1 overexpression promoted proliferation and migration of GC cells in vitro. By examining the tumor-suppressive miRNA microRNA-200a (miR-200a), we found that miR-200a directly silenced Gαi1 to induce anti-GC cell activity. The expression of miR-200a was downregulated in human GC, correlating with upregulation of a novel miR-200a-targeting long non-coding RNA (LncRNA), PINK1 (PTEN Induced Kinase 1)-AS. RNA immunoprecipitation, RNA-pull down, and RNA fluorescence in situ hybridization assays confirmed that PINK1-AS directly binds to miR-200a. Silencing PINK1-AS in GC cells led to miR-200a accumulation, Gαi1 downregulation, and inhibition of GC cell progression in vitro, whereas PINK1-AS upregulation produced the converse results. Significantly, anti-GC cell activity induced by PINK1-AS shRNA was ameliorated by the expression of miR-200a antisense or the 3'-UTR (untranslated region)-depleted Gαi1. In vivo, the growth of subcutaneous MGC-803 xenografts in nude mice was inhibited by PINK1-AS shRNA, but accelerated by PINK1-AS overexpression. Patient-derived GC xenograft growth in nude mice was largely inhibited after intratumoral injection of PINK1-AS shRNA lentivirus. In conclusion, PINK1-AS promotes Gαi1-driven GC progression by sponging miR-200a.

MAFG-driven osteosarcoma cell progression is inhibited by a novel miRNA miR-4660.

Osteosarcoma (OS) is the most common primary bone malignancy in the adolescent population. MAFG (v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog G) forms a heterodimer with Nrf2 (NF-E2-related factor 2), binding to antioxidant response element (ARE), which is required for Nrf2 signaling activation. We found that MAFG mRNA and protein expression is significantly elevated in human OS tissues as well as in established and primary human OS cells. In human OS cells, MAGF silencing or knockout (KO) largely inhibited OS cell growth, proliferation, and migration, simultaneously inducing oxidative injury and apoptosis activation. Conversely, ectopic overexpression of MAFG augmented OS cell progression in vitro. MicroRNA-4660 (miR-4660) directly binds the 3' untranslated region (UTR) of MAFG mRNA in the cytoplasm of OS cells. MAFG 3' UTR luciferase activity and expression as well as OS cell growth were largely inhibited with forced miR-4660 overexpression but augmented with miR-4660 inhibition. In vivo, MAGF short hairpin RNA (shRNA) or forced overexpression of miR-4660 inhibited subcutaneous OS xenograft growth in severe combined immunodeficient mice. Furthermore, MAFG silencing or miR-4660 overexpression inhibited OS xenograft in situ growth in proximal tibia of the nude mice. In summary, MAFG overexpression-driven OS cell progression is inhibited by miR-4660. The miR-4660-MAFG axis could be novel therapeutic target for human OS.

Requirement of Gαi1 and Gαi3 in interleukin-4-induced signaling, macrophage M2 polarization and allergic asthma response.

IL-4 induces Akt activation in macrophages, required for full M2 (alternative) polarization. We examined the roles of Gαi1 and Gαi3 in M2 polarization using multiple genetic methods. Methods and Results: In MEFs and primary murine BMDMs, Gαi1/3 shRNA, knockout or dominant negative mutations attenuated IL-4-induced IL4Rα endocytosis, Gab1 recruitment as well as Akt activation, leaving STAT6 signaling unaffected. Following IL-4 stimulation, Gαi1/3 proteins associated with the intracellular domain of IL-4Rα and the APPL1 adaptor, to mediate IL-4Rα endosomal traffic and Gab1-Akt activation in BMDMs. In contrast, gene silencing of Gαi1/3 with shRNA or knockout resulted in BMDMs that were refractory to IL-4-induced M2 polarization. Conversely, Gαi1/3-overexpressed BMDMs displayed preferred M2 response with IL-4 stimulation. In primary human macrophages IL-4-induced Akt activation and Th2 genes expression were inhibited with Gαi1/3 silencing, but augmented with Gαi1/3 overexpression. In Gαi1/3 double knockout (DKO) mice, M2 polarization, by injection of IL-4 complex or chitin, was potently inhibited. Moreover, in a murine model of asthma, ovalbumin-induced airway inflammation and hyperresponsiveness were largely impaired in Gαi1/3 DKO mice. Conclusion: These findings highlight novel and essential roles for Gαi1/3 in regulating IL-4-induced signaling, macrophage M2 polarization and allergic asthma response.

The histone acetyltransferase HBO1 functions as a novel oncogenic gene in osteosarcoma.

HBO1 (KAT7 or MYST2) is a histone acetyltransferase that acetylates H3 and H4 histones. Methods: HBO1 expression was tested in human OS tissues and cells. Genetic strategies, including shRNA, CRISPR/Cas9 and overexpression constructs, were applied to exogenously alter HBO1 expression in OS cells. The HBO1 inhibitor WM-3835 was utilized to block HBO1 activation. Results:HBO1 mRNA and protein expression is significantly elevated in OS tissues and cells. In established (MG63/U2OS lines) and primary human OS cells, shRNA-mediated HBO1 silencing and CRISPR/Cas9-induced HBO1 knockout were able to potently inhibit cell viability, growth, proliferation, as well as cell migration and invasion. Significant increase of apoptosis was detected in HBO1-silenced/knockout OS cells. Conversely, ectopic HBO1 overexpression promoted OS cell proliferation and migration. We identified ZNF384 (zinc finger protein 384) as a potential transcription factor of HBO1. Increased binding between ZNF384 and HBO1 promoter was detected in OS cell and tissues, whereas ZNF384 silencing via shRNA downregulated HBO1 and produced significant anti-OS cell activity. In vivo, intratumoral injection of HBO1 shRNA lentivirus silenced HBO1 and inhibited OS xenograft growth in mice. Furthermore, growth of HBO1-knockout OS xenografts was significantly slower than the control xenografts. WM-3835, a novel and high-specific small molecule HBO1 inhibitor, was able to potently suppressed OS cell proliferation and migration, and led to apoptosis activation. Furthermore, intraperitoneal injection of a single dose of WM-3835 potently inhibited OS xenograft growth in SCID mice. Conclusion: HBO1 overexpression promotes OS cell growth in vitro and in vivo.

Adherence to the 2018 World Cancer Research Fund/American Institute for Cancer Research cancer prevention recommendations and pancreatic cancer incidence and mortality: A prospective cohort study.

BACKGROUND: Whether adherence to the World Cancer Research Fund/American Institute for Cancer Research (WCRF/AICR) cancer prevention recommendations is associated with a reduced risk of pancreatic cancer remains controversial. Additionally, no study has investigated this association in the US population. Hence, we investigated the association of adherence to the 2018 WCRF/AICR cancer prevention recommendations with pancreatic cancer incidence and mortality in a US population. METHODS: A population-based cohort of 95 962 participants was identified. A score incorporating eight WCRF/AICR components was constructed to reflect adherence to the WCRF/AICR guidelines, with higher scores representing greater adherence to the guidelines. Cox and competing risk regression were used to calculate risk estimates for pancreatic cancer incidence and mortality, respectively. Restricted cubic spline functions were used to test nonlinearity. RESULTS: In the fully adjusted model, higher overall WCRF/AICR scores were shown to be associated with lower risks of developing pancreatic cancer (hazard ratiotertile 3 vs 1 :0.67; 95% confidence interval: 0.49, 0.90; Ptrend  = .0099) and mortality due to this cancer (subdistribution hazard ratiotertile 3 vs 1 0.65; 95% confidence interval: 0.47, 0.89; Ptrend  = .0108) in a linear dose-response manner (all Pnonlinearity  > .05). The component "be physically active" was shown to be a key contributor to the observed associations. No association of the diet-specific WCRF/AICR score with pancreatic incidence and mortality was found. CONCLUSIONS: Adherence to the 2018 WCRF/AICR guidelines, especially "be physically active," confers reduced risks of pancreatic cancer incidence and mortality in the US population; however, adherence to dietary components alone does not confer such beneficial effects.

Comparison of clinicopathological traits and prognostic factors of hepatocellular carcinoma with and without cirrhotic background.

The difference of the patients bearing hepatocellular carcinoma (HCC) with and without cirrhosis at clinical level has not been completely determined. This study compared their differences in clinicopathological traits and prognostic factors for relapse-free survival (RFS) and overall survival (OS). Animal model was established to validate the result of clinical observation. As a result, 82 patients bearing HCC with no cirrhosis (HCC-NC) and 146 patients bearing HCC with cirrhosis (HCC-C) were included. HCC-NC exhibited shorter prothrombin time and higher plasma albumin than HCC-C. In HCC-NC, satellite nodule was an independent risk factor for OS, and high γ-glutamyl transpeptidase was an independent risk factor for RFS. In HCC-C, female sex was an independent risk factor for OS. Stratified analysis showed the OS and RFS of HCC-NC were better than HCC-C in conditions like without cancer embolus (in the portal vein or bile duct), without lymphadenopathy in hepatic portal, without satellite nodule and with small or high-differentiated tumor. Animal model analysis showed HCC-NC had a higher liver/body weight ratio, less tumor count and smaller max tumor volume than HCC-C. In conclusion, clinicopathological traits and risk factors influencing postoperative OS and RFS differed between patients with HCC-C and HCC-NC.

MicroRNA-6862 inhibition elevates sphingosine kinase 1 and protects neuronal cells from MPP+-induced apoptosis.

MPP+ (1-methyl-4-phenylpyridinium)-induced dopaminergic neuronal cell apoptosis is associated with sphingosine kinase 1 (SphK1) inhibition. We here tested the potential effect of microRNA-6862 (miR-6862), a novel SphK1-targeting miRNA, on MPP+-induced cytotoxicity in neuronal cells. MiR-6862 locates in the cytoplasm of SH-SY5Y neuronal cells. It directly binds to SphK1 mRNA. In SH-SY5Y cells and HCN-2 cells, ectopic overexpression of miR-6862 decreased SphK13'-untranslated region luciferase reporter activity and downregulated its expression. miR-6862 inhibition exerted opposite activity and elevated SphK1 expression. In neuronal cells, MPP+-induced cell death was significantly inhibited through miR-6862 inhibition. Conversely, ectopic overexpression of miR-6862 or CRISPR/Cas9-induced SphK1 knockout augmented MPP+-induced apoptosis in the neuronal cells. Importantly, antagomiR-6862 failed to inhibit MPP+-induced apoptosis in SphK1-knockout SH-SY5Y cells. These results suggest that inhibition of miR-6862 induces SphK1 elevation and protects neuronal cells from MPP+-induced cell death.

SC79, a novel Akt activator, protects dopaminergic neuronal cells from MPP+ and rotenone.

In pathogenesis of Parkinson's disease (PD), mitochondrial dysfunction causes substantial reactive oxygen species (ROS) production and oxidative stress, leading to dopaminergic (DA) neuronal cell death. Mitochondrial toxins, including MPP+ (1-methyl-4-phenylpyridinium ion) and rotenone, induce oxidative injury in cultured DA neuronal cells. The current study tested the potential effect of SC79, a first-in-class small-molecule Akt activator, against the process. In SH-SY5Y cells and primary murine DA neurons, SC79 significantly attenuated MPP+- and rotenone-induced viability reduction, cell death, and apoptosis. SC79 activated Akt signaling in DA neuronal cells. Akt inhibition (by LY294002 and MK-2206) or CRISPR-Cas9-mediated Akt1 knockout completely abolished SC79-induced DA neuroprotection against MPP+. Further studies demonstrated that SC79 attenuated MPP+- and rotenone-induced ROS production, mitochondrial depolarization, and lipid peroxidation in SH-SY5Y cells and primary DA neurons. Moreover, upregulation of Nrf2-dependent genes (HO1 and NQO1) and Nrf2 protein stabilization were detected in SC79-treated SH-SY5Y cells and primary DA neurons. Together we show that SC79 protects DA neuronal cells from mitochondrial toxins possibly via activation of Akt-Nrf2 signaling.