Single-cell transcriptomic analysis deciphers heterogenous cancer stem-like cells in colorectal cancer and their organ-specific metastasis.

OBJECTIVE: Metastasis is the major cause of cancer death. However, what types of heterogenous cancer cells in primary tumour and how they metastasise to the target organs remain largely undiscovered. DESIGN: We performed single-cell RNA sequencing and spatial transcriptomic analysis in primary colorectal cancer (CRC) and metastases in the liver (lCRC) or ovary (oCRC). We also conducted immunofluorescence staining and functional experiments to examine the mechanism. RESULTS: Integrative analyses of epithelial cells reveal a stem-like cell cluster with high protein tyrosine phosphatase receptor type O (PTPRO) and achaete scute-like 2 (ASCL2) expression as the metastatic culprit. This cell cluster comprising distinct subpopulations shows distinct liver or ovary metastatic preference. Population 1 (P1) cells with high delta-like ligand 4 (DLL4) and MAF bZIP transcription factor A (MAFA) expression are enriched in primary CRC and oCRC, thus may be associated with ovarian metastasis. P3 cells having a similar expression pattern as cholangiocytes are found mainly in primary CRC and lCRC, presuming to be likely the culprits that specifically metastasise to the liver. Stem-like cells interacted with cancer-associated fibroblasts and endothelial cells via the DLL4-NOTCH signalling pathway to metastasise from primary CRC to the ovary. In the oCRC microenvironment, myofibroblasts provide cancer cells with glutamine and perform a metabolic reprogramming, which may be essential for cancer cells to localise and develop in the ovary. CONCLUSION: We uncover a mechanism for organ-specific CRC metastasis.

Novel Platinum-Nickel Composite Trap for Simultaneous and Direct Determination of Mercury and Cadmium in Soil and Its Mechanism Study.

In this work, following a metal-ceramic heater (MCH) as an electrothermal vaporizer (ETV), a novel composite Pt/Ni trap based on platinizing the foamed nickel was first fabricated to trap Hg and Cd simultaneously. So, a solid sampling Hg-Cd analyzer was developed to simultaneously detect trace Hg and Cd in soil samples, mainly consisting of an MCH, a composite Pt/Ni trap, and an atomic fluorescence spectrometer (AFS). This small-size MCH-ETV system only consumes 100 W for the complete vaporization of Hg and Cd in soil matrices. The Pt/Ni trap fulfills the complete trapping of Hg and Cd following the solid sampling MCH-ETV system and then fast releases them by heating. It was proved that trapped and released Hg and Cd by the Pt/Ni trap are atomic species using X-ray photoelectron spectroscopy (XPS) and other approaches; specially, the effective cotrapping of Hg and Cd might be due to forming alloys of Hg + Pt and Cd + Ni on the Pt/Ni trap. Under the optimized conditions, the method detection limits (LODs) of Hg and Cd reached 0.4 µg/kg and 0.04 µg/kg for a 20 mg sample size, the relative standard deviations (RSDs) were within 12% and 8% for soil samples, respectively, and the recoveries ranged from 96% to 105%, indicating favorable analytical sensitivity, precision, and accuracy. The whole analysis time can be controlled within 5 min without the soil digestion process. The proposed Hg-Cd analyzer is thus suitable for rapid detection of Hg and Cd in soil samples with advantages such as simplicity, green, and safety. Further, the proposed solid sampling ETV-composite trap method has a promising application potential in the field and rapid detection for multielements.

ATXN2-mediated translation of TNFR1 promotes esophageal squamous cell carcinoma via m6A-dependent manner.

N6-methyladenosine (m6A) is the most prevalent RNA modification, and the effect of its dysregulation on esophageal squamous cell carcinoma (ESCC) development remains unclear. Here, by performing transcriptome-wide m6A sequencing in 16 ESCC tissue samples, we identified the key roles of m6A in TNFRSF1A (also known as TNFR1)-mediated MAPK and NF-κB activation in ESCC. Mechanistically, a functional protein involved in m6A methylation, ATXN2, is identified that augments the translation of TNFRSF1A by binding to m6A-modified TNFRSF1A mRNA. Upregulation of the TNFRSF1A protein level, a vital upstream switch for TNFRSF1A-mediated signaling events, activates the NF-κB and MAPK pathways and thus promotes ESCC development. Furthermore, TNFRSF1A m6A modifications and protein levels are upregulated in ESCC, and high levels of TNFRSF1A m6A and protein are correlated with poor ESCC patient survival. These results collectively indicate that the m6A-TNFRSF1A axis is critical for ESCC development and thus may serve as a potential druggable target.

A Brief Review on the Potential of Psychedelics for Treating Alzheimer's Disease and Related Depression.

Alzheimer's disease (AD), the most common form of senile dementia, is poised to place an even greater societal and healthcare burden as the population ages. With few treatment options for the symptomatic relief of the disease and its unknown etiopathology, more research into AD is urgently needed. Psychedelic drugs target AD-related psychological pathology and symptoms such as depression. Using microdosing, psychedelic drugs may prove to help combat this devastating disease by eliciting psychiatric benefits via acting through various mechanisms of action such as serotonin and dopamine pathways. Herein, we review the studied benefits of a few psychedelic compounds that may show promise in treating AD and attenuating its related depressive symptoms. We used the listed keywords to search through PubMed for relevant preclinical, clinical research, and review articles. The putative mechanism of action (MOA) for psychedelics is that they act mainly as serotonin receptor agonists and induce potential beneficial effects for treating AD and related depression.

SMALL PLANT AND ORGAN 1 (SPO1) Encoding a Cellulose Synthase-like Protein D4 (OsCSLD4) Is an Important Regulator for Plant Architecture and Organ Size in Rice.

Plant architecture and organ size are considered as important traits in crop breeding and germplasm improvement. Although several factors affecting plant architecture and organ size have been identified in rice, the genetic and regulatory mechanisms remain to be elucidated. Here, we identified and characterized the small plant and organ 1 (spo1) mutant in rice (Oryza sativa), which exhibits narrow and rolled leaf, reductions in plant height, root length, and grain width, and other morphological defects. Map-based cloning revealed that SPO1 is allelic with OsCSLD4, a gene encoding the cellulose synthase-like protein D4, and is highly expressed in the roots at the seedling and tillering stages. Microscopic observation revealed the spo1 mutant had reduced number and width in leaf veins, smaller size of leaf bulliform cells, reduced cell length and cell area in the culm, and decreased width of epidermal cells in the outer glume of the grain. These results indicate the role of SPO1 in modulating cell division and cell expansion, which modulates plant architecture and organ size. It is showed that the contents of endogenous hormones including auxin, abscisic acid, gibberellin, and zeatin tested in the spo1 mutant were significantly altered, compared to the wild type. Furthermore, the transcriptome analysis revealed that the differentially expressed genes (DEGs) are significantly enriched in the pathways associated with plant hormone signal transduction, cell cycle progression, and cell wall formation. These results indicated that the loss of SPO1/OsCSLD4 function disrupted cell wall cellulose synthase and hormones homeostasis and signaling, thus leading to smaller plant and organ size in spo1. Taken together, we suggest the functional role of SPO1/OsCSLD4 in the control of rice plant and organ size by modulating cell division and expansion, likely through the effects of multiple hormonal pathways on cell wall formation.

LncRNA MIR155HG functions as a ceRNA of miR-223-3p to promote cell pyroptosis in human degenerative NP cells.

Nucleus pulposus (NP) cell pyroptosis plays a critical role in the pathogenesis of intervertebral disk degeneration (IDD). MIR155 host gene (MIR155HG) is a long non-coding RNA with pro-inflammatory activity. However, very little is known about its role in NP cell pyroptosis. This study aimed to observe the impact of MIR155HG on cell pyroptosis and to explore the underlying mechanism in human degenerative NP cells. Our results demonstrated that MIR155HG expression was significantly increased in human degenerative NP tissue samples and showed a positive correlation with Pfirrmann score. Overexpression of MIR155HG through a lentiviral vector decreased miR-223-3p levels, up-regulated NLRP3 expression and induced cell pyroptosis in human degenerative NP cells. A ceRNA action mode was identified among MIR155HG, miR-223-3p, and NLRP3. The stimulatory effect of MIR155HG on human degenerative NP cell pyroptosis was significantly reversed by pretreatment with miR-223-3p mimic or NLRP3 siRNA. In summary, these data suggest that MIR155HG sponges miR-223-3p to promote NLRP3 expression, leading to induction of cell pyroptosis in human degenerative NP cells. Targeting MIR155HG could be a novel and promising strategy to slow down the progression of IDD.

Experimental Research on Vibration-Damping Effect of Combined Shear Hinge Prefabricated Steel Spring Floating Slab Track.

OBJECTIVE: The cast-in-place steel spring floating slab track (SSFST) is difficult to maintain and repair, while the mechanical strength of the end of the traditional prefabricated SSFST is poor. In order to overcome the above shortcomings, a shear-hinge-combined prefabricated SSFST was developed, and an indoor test was carried out to analyze its vibration-damping effect. METHODS: A combined shear hinge SSFST connection model with two length sizes was established. The dynamic response amplitude and frequency response characteristics of the foundation (ground) under different isolator installations and fatigue loads were studied, and the vibration-damping performance of two sizes of combined shear hinge SSFST was evaluated. RESULTS: The vibration-damping effect of the steel spring vibration isolator mainly acts in the middle and low-frequency bands of 16-400 Hz, and the vibration near 10 Hz will be aggravated after the vibration isolator is installed. The vibration index and variation law of the two sizes of SSFST are similar, and the vibration response of 4.8 m SSFST is slightly less than 3.6 m SSFST. There is almost no change in each index when the load is 5 million times, and there is a certain range of change when the load is 10 million times, but the overall change is small. CONCLUSIONS: The combined shear hinge prefabricated SSFST can have an excellent isolation effect on vibration and can still maintain good vibration-damping ability within 10 million fatigue loads (about 5 years); 4.8 m SSFST should be laid in straight sections with higher train speeds, while 3.6 m SSFST should be applied in curved sections to ensure smooth lines.

Novel Dielectric Barrier Discharge Trap for Arsenic Introduced by Electrothermal Vaporization: Possible Mechanism and Its Application.

In this work, a novel integrated dielectric barrier discharge (IDBD) reactor coupled to an electrothermal vaporizer (ETV) was established for arsenic determination. It is for the first time gas-phase enrichment (GPE) was fulfilled based on the hyphenation of ETV and DBD. The mechanisms of evolution of arsenic atomic and molecular species during vaporization, transportation, trapping, and release processes were investigated via X-ray photoelectron spectroscopy (XPS) and other approaches. Tentative mechanisms were deduced as follows: the newly designed DBD atomizer (DBDA) tube upstream to the air inlet fulfills the atomization of arsenic nanoparticles in vaporized aerosol, leading to free arsenic atoms that are indispensable for forming arsenic oxides; the DBD trap (DBDT) tube traps arsenic oxides under an O2-domininating atmosphere and then releases arsenic atoms under H2-dominating atmospheres. In essence, this process is a physical-chemical process rather than an electrostatic particle deposition. Such a trap and release sequence separates matrix interference and enhances analytical sensitivity. Under the optimized conditions, the method detection limit (LOD) was 0.04 mg/kg and the relative standard deviations (RSDs) were within 6% for As standard solution and real seafood samples, indicating adequate analytical sensitivity and precision. The mean spiked recoveries for laver, kelp, and Undaria pinnatifida samples were 95-110%, and the results of the certified reference materials (CRMs) were consistent with certified values. This ETV-DBD preconcentration scheme is easy and green and has low cost for As analysis in seafood samples. DBD was proved a novel ETV transportation enhancement and preconcentration technique for arsenic, revealing its potential in rapid arsenic analysis based on direct solid sampling ETV instrumentation.

Roles of ß-Endorphin in Stress, Behavior, Neuroinflammation, and Brain Energy Metabolism.

ß-Endorphins are peptides that exert a wide variety of effects throughout the body. Produced through the cleavage pro-opiomelanocortin (POMC), ß-endorphins are the primarily agonist of mu opioid receptors, which can be found throughout the body, brain, and cells of the immune system that regulate a diverse set of systems. As an agonist of the body's opioid receptors, ß-endorphins are most noted for their potent analgesic effects, but they also have their involvement in reward-centric and homeostasis-restoring behaviors, among other effects. These effects have implicated the peptide in psychiatric and neurodegenerative disorders, making it a research target of interest. This review briefly summarizes the basics of endorphin function, goes over the behaviors and regulatory pathways it governs, and examines the variability of ß-endorphin levels observed between normal and disease/disorder affected individuals.

Network Medicine Approach for Analysis of Alzheimer's Disease Gene Expression Data.

Alzheimer's disease (AD) is the most widespread diagnosed cause of dementia in the elderly. It is a progressive neurodegenerative disease that causes memory loss as well as other detrimental symptoms that are ultimately fatal. Due to the urgent nature of this disease, and the current lack of success in treatment and prevention, it is vital that different methods and approaches are applied to its study in order to better understand its underlying mechanisms. To this end, we have conducted network-based gene co-expression analysis on data from the Alzheimer's Disease Neuroimaging Initiative (ADNI) database. By processing and filtering gene expression data taken from the blood samples of subjects with varying disease states and constructing networks based on that data to evaluate gene relationships, we have been able to learn about gene expression correlated with the disease, and we have identified several areas of potential research interest.

Simultaneous Monitoring of Multi-Enzyme Activity and Concentration in Tumor Using a Triply Labeled Fluorescent In Vivo Imaging Probe.

The use of fluorescent imaging probes that monitor the activity of proteases that experience an increase in expression and activity in tumors is well established. These probes can be conjugated to nanoparticles of iron oxide, creating a multimodal probe serving as both a magnetic resonance imaging (MRI) agent and an indicator of local protease activity. Previous works describe probes for cathepsin D (CatD) and metalloproteinase-2 (MMP2) protease activity grafted to cross-linked iron oxide nanoparticles (CLIO). Herein, we have synthesized a triply labeled fluorescent iron oxide nanoparticle molecular imaging (MI) probe, including an AF750 substrate concentration reporter along with probes for cathepsin B (CatB) sand MMP2 protease activity. The reporter provides a baseline signal from which to compare the activity of the two proteases. The activity of the MI probe was verified through incubation with the proteases and tested in vitro using the human HT29 tumor cell line and in vivo using female nude mice injected with HT29 cells. We found the MI probe had the appropriate specificity to the activity of their respective proteases, and the reporter dye did not activate when incubated in the presence of only MMP2 and CatB. Probe fluorescent activity was confirmed in vitro, and reporter signal activation was also noted. The fluorescent activity was also visible in vivo, with injected HT29 cells exhibiting fluorescence, distinguishing them from the rest of the animal. The reporter signal was also observable in vivo, which allowed the signal intensities of the protease probes to be corrected; this is a unique feature of this MI probe design.

A Pilot Study of Multi-Input Recurrent Neural Networks for Drug-Kinase Binding Prediction.

The use of virtual drug screening can be beneficial to research teams, enabling them to narrow down potentially useful compounds for further study. A variety of virtual screening methods have been developed, typically with machine learning classifiers at the center of their design. In the present study, we created a virtual screener for protein kinase inhibitors. Experimental compound-target interaction data were obtained from the IDG-DREAM Drug-Kinase Binding Prediction Challenge. These data were converted and fed as inputs into two multi-input recurrent neural networks (RNNs). The first network utilized data encoded in one-hot representation, while the other incorporated embedding layers. The models were developed in Python, and were designed to output the IC50 of the target compounds. The performance of the models was assessed primarily through analysis of the Q2 values produced from runs of differing sample and epoch size; recorded loss values were also reported and graphed. The performance of the models was limited, though multiple changes are proposed for potential improvement of a multi-input recurrent neural network-based screening tool.

A Novel Dual Fluorochrome Near-Infrared Imaging Probe for Potential Alzheimer's Enzyme Biomarkers-BACE1 and Cathepsin D.

A molecular imaging probe to fluorescently image the ß-site of the amyloid precursor protein (APP) cleaving enzyme 1 (BACE1) and cathepsin D (CatD) enzymes associated with Alzheimer's disease (AD) was designed and synthesized. This imaging probe was built upon iron oxide nanoparticles (cross-linked dextran iron oxide nanoparticles, or CLIO). Peptide substrates containing a terminal near-infrared fluorochrome (fluorophore emitting at 775 nm for CatD or fluorophore emitting at 669 nm for BACE1) were conjugated to the CLIO nanoparticles. The CatD substrate contained a phenylalanine-phenylalanine cleavage site more specific to CatD than BACE1. The BACE1 substrate contained the sequence surrounding the leucine-asparagine cleavage site of the BACE1 found in the Swedish mutation of APP, which is more specific to BACE1 than CatD. These fluorescently-labeled peptide substrates were then conjugated to the nanoparticle. The nanoparticle probes were purified by gel filtration, and their fluorescence intensities were determined using a fluorescence plate reader. The CatD peptide substrate demonstrated a 15.5-fold increase in fluorescence when incubated with purified CatD enzyme, and the BACE1 substrate exhibited a 31.5-fold increase in fluorescence when incubated with purified BACE1 enzyme. Probe specificity was also demonstrated in the human H4 neuroglioma cells and the H4 cells stably transfected with BACE1 in which the probe monitored enzymatic cleavage. In the H4 and H4-BACE1 cells, BACE1 and active CatD activity increased, an occurrence that was reflected in enzyme expression levels as determined by immunoblotting. These results demonstrate the applicability of this probe for detecting potential Alzheimer's enzyme biomarkers.

PIWI-interacting RNA-36712 restrains breast cancer progression and chemoresistance by interaction with SEPW1 pseudogene SEPW1P RNA.

BACKGROUND: Breast cancer is one of the most common malignancies and the major cause of cancer-related death in women. Although the importance of PIWI-interacting RNAs (piRNAs) in cancer has been increasingly recognized, few studies have been explored the functional mechanism of piRNAs in breast cancer development and progression. METHODS: We examined the top 20 highly expressed piRNAs based on the analysis of TCGA breast cancer data in two patient cohorts to test the roles of piRNAs in breast cancer. The effects of piRNA-36,712 on the malignant phenotypes and chemosensitivity of breast cancer cells were detected in vitro and in vivo. MS2-RIP and reporter gene assays were conducted to identify the interaction and regulation among piRNA-36,712, miRNAs and SEPW1P. Kaplan-Meier estimate with log-rank test was used to compare patient survival by different piRNA-36,712 expression levels. RESULTS: We found piRNA-36,712 level was significantly lower in breast cancer than in normal breast tissues and low level was correlated with poor clinical outcome in patients. Functional studies demonstrated that piRNA-36,712 interacts with RNAs produced by SEPW1P, a retroprocessed pseudogene of SEPW1, and subsequently inhibits SEPW1 expression through competition of SEPW1 mRNA with SEPW1P RNA for microRNA-7 and microRNA-324. We also found that higher SEPW1 expression due to downregulation of piRNA-36,712 in breast cancer may suppress P53, leading to the upregulated Slug but decreased P21 and E-cadherin levels, thus promoting cancer cell proliferation, invasion and migration. Furthermore, we found that piRNA-36,712 had synergistic anticancer effects with the paclitaxel and doxorubicin, two chemotherapeutic agents for breast cancer. CONCLUSIONS: These findings suggest that piRNA-36,712 is a novel tumor suppressor and may serve as a potential predictor for the prognosis of breast cancer patients.

Expression of matrix Metalloproteinases-2 and aquaporin-1 in corneoscleral junction after angle-closure in rabbits.

BACKGROUND: To investigate the expression of Matrix Metalloproteinases 2 and aquaporin-1 in corneoscleral junction and explore the mechanism of trabecular damageafter angle-closure. METHODS: Thirty New Zealand white rabbits were randomly assigned into 2 groups, theexperimental group (Group 1) including twenty five rabbits and the control group (Group 2) including 5 rabbits. The rabbits in the experimental group were used to establish angle-closure models, and the rabbits in the control group were not subjected to any operation. All the rabbits were followed by slit lamp microscopy, Tonopen tonometer, and anterior segment optical coherent tomography (AS-OCT). The expressions of metalloproteinase MMP-2, aquaporin-1, and tissue inhibitors of metalloproteinase-2 in corneoscleral junctionwere evaluatedin both groups byimmunofluorescence, quantitative reverse-transcription polymerase chain reaction (qRT-PCR), and enzyme-linked immunosorbent assay (ELISA). RESULTS: Slit-lamp examination showed that angle-closure model was successfully established in twenty rabbits. The extent of angle-closure was about 2 to 4 clock hours in all the rabbit models, but the intraocular pressure (IOP) of the rabbits distributed from 8.57 to 15.25 mmHg and no significant high IOP was found in the follow-up period. The AQP-1-positive cells mainly located in Schlemm's canal, the inner surface of trabecular meshwork (TM), and the surface of iris, which began to decline on 1 month after angle-closure. MMP2 staining was diffuse in trabecular meshwork and iris. Immunofluorescence signal of MMP2 was strong within 1 month after angle-closure, and subsequently became weak. qRT-PCR and ELISA showed that the expression of MMP-2 and TIMP-2 increased within 1 month after angle-closure and then declined gradually. The AQP-1 levels showed slightly declined on 1 month after angle-closure. CONCLUSIONS: Altered levels of MMPs, TIMPs, and AQP-1 were found in the area of angle-closure, which may be involved in the damage of TM and Schlemm's canal after angle-closure.

Machine Learning Models Combined with Virtual Screening and Molecular Docking to Predict Human Topoisomerase I Inhibitors.

In this work, random forest (RF), support vector machine, k-nearest neighbor and C4.5 decision tree, were used to establish classification models for predicting whether an unknown molecule is an inhibitor of human topoisomerase I (Top1) protein. All these models have achieved satisfactory results, with total prediction accuracies from 89.70% to 97.12%. Through comparative analysis, it can be found that the RF model has the best forecasting effect. The parameters were further optimized to generate the best-performing RF model. At the same time, features selection was implemented to choose properties most relevant to the inhibition of Top1 from 189 molecular descriptors through a special RF procedure. Subsequently, a ligand-based virtual screening was performed from the Maybridge database by the optimal RF model and 596 hits were picked out. Then, 67 molecules with relative probability scores over 0.7 were selected based on the screening results. Next, the 67 molecules above were docked to Top1 using AutoDock Vina. Finally, six top-ranked molecules with binding energies less than -10.0 kcal/mol were screened out and a common backbone, which is entirely different from that of existing Top1 inhibitors reported in the literature, was found.

Manganese causes neurotoxic iron accumulation via translational repression of amyloid precursor protein and H-Ferritin.

For more than 150 years, it is known that occupational overexposure of manganese (Mn) causes movement disorders resembling Parkinson's disease (PD) and PD-like syndromes. However, the mechanisms of Mn toxicity are still poorly understood. Here, we demonstrate that Mn dose- and time-dependently blocks the protein translation of amyloid precursor protein (APP) and heavy-chain Ferritin (H-Ferritin), both iron homeostatic proteins with neuroprotective features. APP and H-Ferritin are post-transcriptionally regulated by iron responsive proteins, which bind to homologous iron responsive elements (IREs) located in the 5'-untranslated regions (5'-UTRs) within their mRNA transcripts. Using reporter assays, we demonstrate that Mn exposure repressed the 5'-UTR-activity of APP and H-Ferritin, presumably via increased iron responsive proteins-iron responsive elements binding, ultimately blocking their protein translation. Using two specific Fe2+ -specific probes (RhoNox-1 and IP-1) and ion chromatography inductively coupled plasma mass spectrometry (IC-ICP-MS), we show that loss of the protective axis of APP and H-Ferritin resulted in unchecked accumulation of redox-active ferrous iron (Fe2+ ) fueling neurotoxic oxidative stress. Enforced APP expression partially attenuated Mn-induced generation of cellular and lipid reactive oxygen species and neurotoxicity. Lastly, we could validate the Mn-mediated suppression of APP and H-Ferritin in two rodent in vivo models (C57BL6/N mice and RjHan:SD rats) mimicking acute and chronic Mn exposure. Together, these results suggest that Mn-induced neurotoxicity is partly attributable to the translational inhibition of APP and H-Ferritin resulting in impaired iron metabolism and exacerbated neurotoxic oxidative stress. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.

BRCA1-Associated Protein Increases Invasiveness of Esophageal Squamous Cell Carcinoma.

BACKGROUND & AIMS: We performed a screen for genes whose expression correlates with invasiveness of esophageal squamous cell carcinoma (ESCC) cells. We studied the effects of overexpression and knockdown of these genes in cell lines and expression levels in patient samples. METHODS: We selected genes for analysis from 11 loci associated with risk of ESCC. We analyzed the effects of knocking down expression of 47 of these genes using RNA interference on-chip analysis in ESCC cells and HeLa cells. Cells with gene overexpression and knockdown were analyzed in migration and invasion assays or injected into nude mice and metastasis of xenograft tumors was quantified. We collected ESCC and non-tumor esophageal tissues from 94 individuals who underwent surgery in China from 2010 and 2014; clinical information was collected and survival time was measured from the date of diagnosis to the date of last follow-up or death. Levels of messenger RNAs (mRNAs) were quantified by RNA sequencing, and levels of proteins were determined from immunoblot analyses. Patient survival was compared with mRNA levels using Kaplan-Meier methods and hazard ratios were calculated by Cox models. RESULTS: We identified 8 genes whose disruption increased migration and 10 genes whose disruption reduced migration. Knockdown of BRCA1-associated protein gene (BRAP) significantly reduced migration of KYSE30, KYSE150, and HeLa cells. In patient tumors, 90% of ESCCs examined had higher levels of BRAP protein than paired non-tumor tissues, and 63.8% had gains in BRAP DNA copy number. Levels of BRAP mRNA in ESCC tissues correlated with patient survival time, and high expression increased risk of death 2.4-fold compared with low expression. ESCCs that had metastasized to lymph node had significantly higher levels of BRAP mRNA than tumors without metastases. Knockdown of BRAP in ESCC and HeLa cell lines significantly reduced migration and invasiveness; these cell lines formed less metastases in mice than control cells. Nuclear translocation of the nuclear factor-κB (NF-κB) P65 subunit and phosphorylation of inhibitor of NF-κB kinase subunit ß (IKBKB or IKKß) increased in cells that overexpressed BRAP and decreased in cells with BRAP knockdown. In immunoprecipitation assays, BRAP interacted directly with IKKß. Expression of matrix metalloproteinase 9 and vascular epithelial growth factor C, which are regulated by NF-κB, was significantly reduced in cells with knockdown of BRAP and significantly increased in cells that overexpressed BRAP. CONCLUSIONS: Expression of BRAP is increased in ESCC samples compared with non-tumor esophageal tissues; increased expression correlates with reduced patient survival time and promotes metastasis of xenograft tumors in mice. BRAP overexpression leads to increased activity of NF-κB and expression of matrix metalloproteinase 9 and vascular epithelial growth factor C.

Solute Carrier Family 39 Member 6 Gene Promotes Aggressiveness of Esophageal Carcinoma Cells by Increasing Intracellular Levels of Zinc, Activating Phosphatidylinositol 3-Kinase Signaling, and Up-regulating Genes That Regulate Metastasis.

BACKGROUND & AIMS: A common variant in the solute carrier family 39 member 6 gene (SLC39A6) has been associated with survival times of patients with esophageal squamous cell carcinoma (ESCC). We investigated the function of SLC39A6 and ways in which this variant affects tumor progression by studying ESCC samples and cell lines. METHODS: SLC39A6 was expressed or knocked down by expression of short hairpin RNAs in ESCC cells (KYSE30 and KYSE450) and HeLa cells using lentiviral vectors; we analyzed effects on proliferation, colony formation, migration, and invasion in vitro. Cells were grown as xenograft tumors in nude mice and tumor volume and metastases were quantified; tumors were collected and analyzed histologically. Cells were also analyzed for levels of intracellular zinc and messenger RNA (mRNA) expression patterns. We obtained ESCC and adjacent normal esophageal tissues from 94 patients who underwent esophagectomy in China from 2010 through 2014. Survival times of patients were measured from the date of diagnosis to the date of last follow-up or death. We sequenced mRNAs and compared levels between tumor and non-tumor tissues using the Wilcox rank-sum test. Total proteins in cell lines or tissue samples were measured by immunoblotting. We searched publicly available databases for variants of SLC39A6 in human tumor and non-tumor tissues. RESULTS: Knockdown of SLC39A6 reduced proliferation of ESCC cells in culture and metastasis of xenograft tumors in mice. Cells that overexpressed SLC39A6 had significant increases in intracellular levels of zinc and were more invasive in assays, activating phosphatidylinositol 3-kinase signaling to AKT serine/threonine kinase 1 and mitogen-activated protein kinase 1. Cells that overexpressed SLC39A6 had increased expression of mRNAs and proteins associated with metastasis, such as matrix metalloproteinase (MMP) 1, MMP3, MYC, and snail family transcriptional repressor 2 (SNAI2 or SLUG). Levels of MMP1, MMP3, MYC, and SLUG mRNAs correlated with levels of SLC39A6 mRNA in ESCC samples from patients. ESCC tissues had increased levels of SLC39A6 mRNA compared with non-tumor tissues; the increase correlated with tumor metastasis to lymph node and reduced patient survival time. CONCLUSIONS: In an analysis of ESCC samples and cell lines, we associated increased expression of SLC39A6 with tumor invasiveness, intracellular level of zinc, and patient survival time. ESCC cell lines that overexpress SLC39A6 up-regulate expression MMP1, MMP3, MYC, and SLUG and form metastatic xenograft tumors in mice. Up-regulation of SLC39A6 might be used to determine prognoses of patients with ESCC or as a therapeutic target.

Network Medicine for Alzheimer's Disease and Traditional Chinese Medicine.

Alzheimer's Disease (AD) is a neurodegenerative condition that currently has no known cure. The principles of the expanding field of network medicine (NM) have recently been applied to AD research. The main principle of NM proposes that diseases are much more complicated than one mutation in one gene, and incorporate different genes, connections between genes, and pathways that may include multiple diseases to create full scale disease networks. AD research findings as a result of the application of NM principles have suggested that functional network connectivity, myelination, myeloid cells, and genes and pathways may play an integral role in AD progression, and may be integral to the search for a cure. Different aspects of the AD pathology could be potential targets for drug therapy to slow down or stop the disease from advancing, but more research is needed to reach definitive conclusions. Additionally, the holistic approaches of network pharmacology in traditional Chinese medicine (TCM) research may be viable options for the AD treatment, and may lead to an effective cure for AD in the future.