Global trends and burden of stroke attributable to particulate matter pollution from 1990 to 2019.

OBJECTIVE: To investigate the association between particulate matter and the incidence, disability, and mortality of stroke, we reported the burden of stroke attributable to particulate matter (PM2.5) pollution, including ambient particulate matter pollution (APMP) and household air pollution from solid fuels (HAP), from 1990 to 2019. METHODS: We retrieved the detailed data on the burden of stroke attributable to PM2.5 from the Global Burden of Disease (GBD) 2019. The number of disability-adjusted life-years (DALYs) and deaths, age-standardized death rates (ASMR), and age-standardized disability-adjusted life-years rates (ASDR) attributable to PM2.5 were estimated by age, sex, geographical location, socio-demographic index (SDI), and stroke subtypes (ischemic stroke, intracerebral hemorrhage, and subarachnoid hemorrhage). The estimated annual percentage change (EAPC) was calculated to assess the trends in ASDR and ASMR during the period 1990-2019. RESULTS: Regarding stroke subtypes, the proportion of ischemic stroke burden is increasing, while intracerebral hemorrhage carries the heaviest burden. Both APMP and HAP contributed the most to stroke-related deaths and DALYs of stroke among the elderly populations and males. The highest ASDR and ASMR of stroke attributable to APMP were in the middle SDI regions, especially in East Asia. For HAP, the highest ASDR and ASMR were in the low SDI regions, mainly in Oceania. From 1990-2019, in terms of the EAPC results, APMP caused an increased burden of stroke, whereas the impact of HAP significantly fell. The most pronounced increase in ASDR and ASMR for strokes attributed to APMP were in the low-middle SDI and low SDI regions, particularly among the 25-35 age group. CONCLUSIONS: Stroke attributed to PM2.5 is a global health problem, and the patterns and trends were heterogeneous across APMP and HAP. Targeted interventions should be formulated for APMP and HAP.

The toxic influence of dibromoacetic acid on the hippocampus and pre-frontal cortex of rat: involvement of neuroinflammation response and oxidative stress.

Dibromoacetic acid (DBA) exsits in drinking water as a by-product of disinfection as a result of chlorination or ozonation processes. Hippocampus and pre-frontal cortex are the key structures in memory formation and weanling babies are more sensitive to environmental toxicant than adults, so this study was conducted to evaluate the potential neurotoxicity effects of DBA exposure when administered intragastrically for 4 weeks to weanling Sprague-Dawley rats, at concentration of 0, 20, 50, 125 mg/kg via the neurobehavioral and neurochemical effects. Results indicated that animals weight gain and food consumption were not significantly affected by DBA. However, morris water maze test showed varying degrees of changes between control and high-dose group. Additionally, the level of malondialdehyde (MDA) and generation of reactive oxygen species (ROS) in the hippocampus and pre-frontal cortex of rats increased significantly. The activities of total superoxide dismutase (SOD) and the glutathione (GSH) content in the hippocampus and pre-frontal cortex of rats decreased significantly after treatment with DBA. Treatment with DBA increased the protein and mRNA expression of Iba-1, NF-κB, TNF-α, IL-6, IL-1ß and HO-1 in the hippocampus and pre-frontal cortex of rats. These data suggested that DBA had a toxic influence on the hippocampus and pre-frontal cortex of rats, and that the mechanism of toxicity might be associated with the neuroinflammation response and oxidative stress.

The effects of icariin on the expression of HIF-1α, HSP-60 and HSP-70 in PC12 cells suffered from oxygen-glucose deprivation-induced injury.

CONTEXT: The effects of icariin, a chief constituent of flavonoids from Epimedium brevicornum Maxim (Berberidaceae), on the levels of HIF-1α, HSP-60 and HSP-70 remain unknown. OBJECTIVE: To explore the effects of icariin on the levels of HSP-60, HIF-1α and HSP-70 neuron-specific enolase (NSE) and cell viability. MATERIALS AND METHODS: PC12 cells were treated with icariin (10-7, 10-6 or 10-5 mol/L) for 3 h (1 h before oxygen-glucose deprivation (OGD) plus 2 h OGD). HSP-60, HIF-1α, HSP-70 and NSE were measured using enzyme-linked immunosorbent assay (ELISA). Cell viability was determined by metabolic 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS: After 2 h OGD, levels of HIF-1α, HSP-60, HSP-70 and NSE were increased significantly (HIF-1α: 33.3 ± 1.9 ng/L, HSP-60: 199 ± 16 ng/L, HSP-70: 195 ± 17 ng/L, NSE: 1487 ± 125 ng/L), and cell viability was significantly decreased (0.26 ± 0.03), while icariin (10-7, 10-6, or 10-5 mol/L) significantly reduced the contents of HIF-1α, HSP-60, HSP-70 and NSE (HIF-1α: 14.1 ± 1.4, 22.6 ± 1.8, 15.7 ± 2.1, HSP-60: 100 ± 12, 89 ± 6, 113 ± 11, HSP-70: 139 ± 9, 118 ± 7, 95 ± 9 and NSE: 1121 ± 80, 1019 ± 52, 731 ± 88), and improved cell viability (0.36 ± 0.03, 0.38 ± 0.04, 0.37 ± 0.03) in OGD-treated PC12 cells. DISCUSSION AND CONCLUSION: These results indicate that the protective mechanisms of icariin against OGD-induced injury may be related to down-regulating the expression of HIF-1α, HSP-60 and HSP-70.

Dibromoacetic Acid Induces Thymocyte Apoptosis by Blocking Cell Cycle Progression, Increasing Intracellular Calcium, and the Fas/FasL Pathway in Vitro.

Dibromoacetic acid (DBAA), a haloacetic acid found in drinking water as a disinfection by-product, can cause many adverse effects, including immunotoxicity. In a previous study, we confirmed that DBAA can induce obvious immunotoxicity in mice but that the underlying mechanisms are not clearly understood. In our current study, we confirmed that DBAA induced cytotoxicity and apoptosis in thymocytes isolated from mice by a range of DBAA concentrations (0, 5, 10, 20, or 40 µM). The data showed that DBAA exposure led to a significant decrease in proliferative responses to T-cell mitogens and obvious inhibition in the production of cytokines interleukin-2 and interleukin-4. We found obvious morphological changes of apoptosis in thymocytes and observed the percentage of apoptotic thymocytes to increase significantly as the DBAA concentration increased. Further investigation showed that DBAA can cause G0/G1 arrest in cell cycle analysis, increase intracellular calcium ([Ca(2+)]i) levels, increase the expression of Fas/FasL proteins, and decrease the expression of Bcl-2 protein. It is concluded that in vitro exposure to DBAA can lead to marked cytotoxicity and apoptosis among thymocytes, and the mechanism involved is strongly related to blocking cell cycle progression, increasing intracellular calcium, and increasing Fas/FasL expressions.

Nano-Regulator Inhibits Tumor Immune Escape via the "Two-Way Regulation" Epigenetic Therapy Strategy.

Tumor immune escape caused by low levels of tumor immunogenicity and immune checkpoint-dependent suppression limits the immunotherapeutic effect. Herein, a "two-way regulation" epigenetic therapeutic strategy is proposed using a novel nano-regulator that inhibits tumor immune escape by upregulating expression of tumor-associated antigens (TAAs) to improve immunogenicity and downregulating programmed cell death 1 ligand 1 (PD-L1) expression to block programmed death-1 (PD-1)/PD-L1. To engineer the nano-regulator, the DNA methyltransferase (DNMT) inhibitor zebularine (Zeb) and the bromodomain-containing protein 4 (BRD4) inhibitor JQ1 are co-loaded into the cationic liposomes with condensing the toll-like receptor 9 (TLR9) agonist cytosine-phosphate-guanine (CpG) via electrostatic interactions to obtain G-J/ZL. Then, asparagine-glycine-arginine (NGR) modified material carboxymethyl-chitosan (CMCS) is coated on the surface of G-J/ZL to construct CG-J/ZL. CG-J/ZL is shown to target tumor tissue and disassemble under the acidic tumor microenvironment (TME). Zeb upregulated TAAs expression to improve the immunogenicity; JQ1 inhibited PD-L1 expression to block immune checkpoint; CpG promote dendritic cell (DC) maturation and reactivated the ability of tumour-associated macrophages (TAM) to kill tumor cells. Taken together, these results demonstrate that the nano-regulator CG-J/ZL can upregulate TAAs expression to enhance T-cell infiltration and downregulate PD-L1 expression to improve the recognition of tumor cells by T-cells, representing a promising strategy to improve antitumor immune response.

Drosophila ryanodine receptors mediate general anesthesia by halothane.

BACKGROUND: Although in vitro studies have identified numerous possible targets, the molecules that mediate the in vivo effects of volatile anesthetics remain largely unknown. The mammalian ryanodine receptor (Ryr) is a known halothane target, and the authors hypothesized that it has a central role in anesthesia. METHODS: Gene function of the Drosophila Ryr (dRyr) was manipulated in the whole body or in specific tissues using a collection of mutants and transgenes, and responses to halothane were measured with a reactive climbing assay. Cellular responses to halothane were studied using Ca imaging and patch clamp electrophysiology. RESULTS: Halothane potency strongly correlates with dRyr gene copy number, and missense mutations in regions known to be functionally important in the mammalian Ryrs gene cause dominant hypersensitivity. Tissue-specific manipulation of dRyr shows that expression in neurons and glia, but not muscle, mediates halothane sensitivity. In cultured cells, halothane-induced Ca efflux is strictly dRyr-dependent, suggesting a close interaction between halothane and dRyr. Ca imaging and electrophysiology of Drosophila central neurons reveal halothane-induced Ca flux that is altered in dRyr mutants and correlates with strong hyperpolarization. CONCLUSIONS: In Drosophila, neurally expressed dRyr mediates a substantial proportion of the anesthetic effects of halothane in vivo, is potently activated by halothane in vitro, and activates an inhibitory conductance. The authors' results provide support for Ryr as an important mediator of immobilization by volatile anesthetics.

Ultrasound Images under an Optimized Image Processing Algorithm in Guiding the Neurological Safety of Resection of Lumbar Disc Nucleus Pulposus in Spinal Surgery.

This study was aimed at investigating the effect of an optimized image processing algorithm in ultrasound images and the influence of resection of lumbar disc nucleus pulposus in spinal surgery under the guidance of ultrasound images on the neurological safety of patients. A total of 60 patients with lumbar disc herniation were selected and divided randomly into the control group and observation group. Patients from the control group were treated with resection of lumbar disc nucleus pulposus by an X-ray-guided foraminal microscope, and patients from the observation group underwent the ultrasound image-guided surgeries with an optimized image processing algorithm. Then, the treatment of patients from the two groups was compared. The results showed that the radiotherapy time in the control group was 120 ± 6.3 min and the radiotherapy dose was 129 ± 10.3 min/sec, while the radiotherapy time in the observation group was 4.5 ± 1.2 min and the radiotherapy dose was 22 ± 7.7 min/sec. The time and dose of radiotherapy in the observation group were significantly lower than those in the control group (P < 0.05). In the control group, the numbers of significant effective cases, effective cases, and ineffective cases were 8, 16, and 6, respectively, while those in the observation group were 12, 18, and 0, respectively. The comparison between the groups showed that the number of effective cases and the number of effective cases in the observation group were significantly higher than those in the control group, and the number of ineffective cases was significantly lower than that in the control group (P < 0.05). In conclusion, ultrasound-guided percutaneous foraminal lumbar discectomy could improve patients' clinical symptoms, promote clinical efficacy, and reduce postoperative pain symptoms, thereby accelerating the postoperative rehabilitation of patients. Moreover, it was extremely safe for the nerves.

Tiling of r7 axons in the Drosophila visual system is mediated both by transduction of an activin signal to the nucleus and by mutual repulsion.

The organization of neuronal wiring into layers and columns is a common feature of both vertebrate and invertebrate brains. In the Drosophila visual system, each R7 photoreceptor axon projects within a single column to a specific layer of the optic lobe. We refer to the restriction of terminals to single columns as tiling. In a genetic screen based on an R7-dependent behavior, we identified the Activin receptor Baboon and the nuclear import adaptor Importin-alpha3 as being required to prevent R7 axon terminals from overlapping with the terminals of R7s in neighboring columns. This tiling function requires the Baboon ligand, dActivin, the transcription factor, dSmad2, and retrograde transport from the growth cone to the R7 nucleus. We propose that dActivin is an autocrine signal that restricts R7 growth cone motility, and we demonstrate that it acts in parallel with a paracrine signal that mediates repulsion between R7 terminals.

Atrazine-induced apoptosis of splenocytes in BALB/C mice.

BACKGROUND: Atrazine (2-chloro-4-ethytlamino-6-isopropylamine-1,3,5-triazine; ATR), is the most commonly applied broad-spectrum herbicide in the world. Unintentional overspray of ATR poses an immune function health hazard. The biomolecular mechanisms responsible for ATR-induced immunotoxicity, however, are little understood. This study presents on our investigation into the apoptosis of splenocytes in mice exposed to ATR as we explore possible immunotoxic mechanisms. METHODS: Oral doses of ATR were administered to BALB/C mice for 21 days. The histopathology, lymphocyte apoptosis and the expression of apoptosis-related proteins from the Fas/Fas ligand (FasL) apoptotic pathway were examined from spleen samples. RESULTS: Mice administered ATR exhibited a significant decrease in spleen and thymus weight. Electron microscope histology of ultrathin sections of spleen revealed degenerative micromorphology indicative of apoptosis of splenocytes. Flow cytometry revealed that the percentage of apoptotic lymphocytes increased in a dose-dependent manner after ATR treatment. Western blots identified increased expression of Fas, FasL and active caspase-3 proteins in the treatment groups. CONCLUSIONS: ATR is capable of inducing splenocytic apoptosis mediated by the Fas/FasL pathway in mice, which could be the potential mechanism underlying the immunotoxicity of ATR.

Identification of the VIL2 enhancer in human embryonic kidney cells.

We previously demonstrated that the VIL2 -87/+134 region exhibited promoter activity in some human cells, and a region further upstream of this promoter might contain an enhancer. However, the properties and location of this VIL2 enhancer remain unclear. In this study, we cloned the VIL2 -1541/-706 segment and investigated its transcriptional regulatory properties via luciferase assays in transiently transfected HEK-293 cells (human embryonic kidney cells). The VIL2 -1541/-706 was found to exhibit promoter activity. Furthermore, when this segment was located upstream of the VIL2 or SV40 (simian virus 40) promoters in the forward orientation, the expression levels of luciferase were dramatically enhanced. However, this transcriptional enhancement disappeared when this segment was located upstream of the promoter in the reverse orientation or downstream of the reporter gene in the forward or reverse orientation. In deletion experiments, we found several potential regulatory regions within the VIL2 -1541/-706. When these regions were separately located upstream of the VIL2 or SV40 promoters, only the -1297/-1186 considerably enhanced the activity of these promoters. Although the other regulatory regions exhibited significant transcriptional regulation in deletion experiments, they weakly enhanced VIL2 promoter activity and/or did not regulate SV40 promoter activity. These results suggest that the DNA sequence upstream of the VIL2 promoter functions as an enhancer in a position- and orientation-dependent manner, and the VIL2 -1297/-1186, which acts as a key enhancer, probably regulates VIL2 transcription in combination with other potential regulatory regions located upstream of the VIL2 promoter.

Specificity protein 1 regulates fascin expression in esophageal squamous cell carcinoma as the result of the epidermal growth factor/extracellular signal-regulated kinase signaling pathway activation.

The overexpression of fascin in human carcinomas is associated with aggressive clinical phenotypes and poor prognosis. However, the molecular mechanism underlying the increased expression of fascin in cancer cells is largely unknown. Here, we identified a Sp1 binding element located at -70 to -60 nts of the FSCN1 promoter and validated that Sp1 specifically bound to this element in esophageal carcinoma cells. Fascin expression was enhanced by Sp1 overexpression and blocked by Sp1 RNAi knockdown. Specific inhibition of ERK1/2 decreased phosphorylation levels of Sp1, and thus suppressed the transcription of the FSCN1, resulting in the down-regulation of fascin. Stimulation with EGF could enhance fascin expression via activating the ERK1/2 pathway and increasing phosphorylation levels of Sp1. These data suggest that FSCN1 transcription may be subjected to the regulation of the EGF/EGFR signaling pathway and can be used as a viable biomarker to predict the efficacy of EGFR inhibitors in cancer therapies.

Potential transcriptional regulatory regions exist upstream of the human ezrin gene promoter in esophageal carcinoma cells.

We previously demonstrated that the region -87/+134 of the human ezrin gene (VIL2) exhibited promoter activity in human esophageal carcinoma EC109 cells, and a further upstream region -1324/-890 positively regulated transcription. In this study, to identify the transcriptional regulatory regions upstream of the VIL2 promoter, we cloned VIL2 -1541/-706 segment containing the -1324/-890, and investigated its transcriptional regulatory properties via luciferase assays in transiently transfected cells. In EC109 cells, it was found that VIL2 -1541/-706 possessed promoter and enhancer activities. We also localized transcriptional regulatory regions by fusing 5'- or 3'-deletion segments of VIL2 -1541/-706 to a luciferase reporter. We found that there were three positive and one negative transcriptional regulatory regions within VIL2 -1541/-706 in EC109 cells. When these regions were separately located upstream of the luciferase gene without promoter, or located upstream of the VIL2 promoter or SV40 promoter directing the luciferase gene, only VIL2 -1297/-1186 exhibited considerable promoter and enhancer activities, which were lower than those of -1541/-706. In addition, transient expression of Sp1 increased ezrin expression and the transcriptional activation of VIL2 -1297/-1186. Other three regions, although exhibiting significantly positive or negative transcriptional regulation in deletion experiments, showed a weaker or absent regulation. These data suggested that more than one region upstream of the VIL2 promoter participated in VIL2 transcription, and the VIL2 -1297/-1186, probably as a key transcriptional regulatory region, regulated VIL2 transcription in company with other potential regulatory regions.

The tumor-suppressive effects of alpha-ketoglutarate-dependent dioxygenase FTO via N6-methyladenosine RNA methylation on bladder cancer patients.

N6-methyladenosine (m6A) methylation participates in the progression of bladder cancer (BCa). Nevertheless, the regulatory mechanism of alpha-ketoglutarate-dependent dioxygenase FTO influencing the BCa progression has still remained elusive. In this study, to investigate the tumor-suppressive effects of FTO via m6A RNA methylation on BCa patients, a total of 15 cancer tissues and adjacent normal tissues (ANTs) were collected from BCa patients who received tumor resection in our hospital from September 2015 to December 2019. We found that the FTO expression was significantly reduced in cancer tissues compared with that in ANTs, which indicated a lower malignant potential and a higher overall survival rate. It was revealed that overexpression of FTO in two human urinary BCa cell lines (HT-1197 and HT-1376) significantly decreased the cell proliferation and invasion abilities compared with the negative controls, whereas the cell apoptosis was markedly enhanced. In addition, we noted that the changes in m6A methylation level mainly appeared at 5' untranslated region (5' UTR) of MALAT1 and NOTCH1 transcripts, and at 3' UTR of CSNK2A2 and ITGA6 transcripts, responding to the overexpression of FTO. Mechanistically, we found that the splicing factor, proline- and glutamine-rich (SFPQ) could influence the FTO-mediated m6A RNA demethylation, eventually affecting the gene expression. This study provided a new insight into the relationship between the FTO expression and the m6A RNA methylation, assisting scholars to better understand the pathogenesis of BCa.

Novel XTENylated AWRK6 analog with hypoglycemic activity, and anti-HSV-2 potential in combination with double shRNA.

AIMS: To design and evaluate a novel AWRK6 peptide-based long-acting GLP-1 receptor agonist (GLP-1RA) conjugated a recombinant polyethylene glycol mimetic (XTEN protein) with significant therapeutic potential on type 2 diabetes mellitus (T2DM) alone as well as Herpes simplex virus type 2 (HSV-2) infection in combination with double shRNA. MAIN METHODS: First, four AWRK6 analogs (termed XA-1 to XA-4) were designed and produced by solid phase synthesis strategy. Further surface plasmon resonance (SPR) measurement and in vitro cAMP accumulation assay were performed to detect the GLP-1R binding affinities and GLP-1R activation, respectively. The in vivo efficacy evaluation including pharmacokinetic test, oral glucose tolerance test (OGTT), hypoglycemic duration test and chronic pharmacodynamics study in rodent animals were all carefully performed. KEY FINDINGS: Four XA peptides were synthesized with purity >99%. High binding affinity as well as activation potency of XA-4 for GLP-1R were demonstrated by SPR and cell-based luciferase reporter assay, respectively. Additionally, XA-4 exhibited the long-lasting antidiabetic effects in the multiple OGTTs, hypoglycemic duration test and chronic study in mice. Furthermore, combined treatment of XA-4 and double shRNA (D-shRNA) achieved potent antiviral effects in HSV-2 infected HEK293 cells. SIGNIFICANCE: XA-4 exhibited promising pharmaceutical potential to be a therapeutic drug for treating T2D, and also held potential to against the HSV-2 infection, which is really an accidental discovery. The strategy of recombinant XTENylation can also be applied to other peptides or small molecules for the development of long-acting therapeutic drugs.

Visible-Light-Responsive Nanofibrous α-Fe2O3 Integrated FeOx Cluster-Templated Siliceous Microsheets for Rapid Catalytic Phenol Removal and Enhanced Antibacterial Activity.

Visible-light-driven environmental contaminants control using 2D photocatalytic nanomaterials with an unconfined reaction-diffusion path is advantageous for public health. Here, cost-effective siliceous composite microsheets (FeSiO-MS) combined with two distinct refined α-Fe2O3 nanospecies as photofunctional catalysts were constructed via a one-pot synthesis approach. Through precise control of Fe2+ precursor addition, specially configured α-Fe2O3 nanofibers combined with FeOx cluster-functionalized siliceous microsheets of ∼15 nm gradually evolved from the iron oxide-bearing molecular sieve, endowing a superior light-response characteristic of the formed nanocomposite. The catalytic experiment along with the ESR study demonstrated that the produced FeSiO-MS showed reinforced photo-Fenton reactivity, which was effective for rapid phenol degradation under visible light radiation. Moreover, the phenol removal process was found to be regulated by the specially configured types and concentrations of iron oxides. Notably, the obtained composites exhibited a considerable visible-light-induced bactericidal effect against E. coli. The constructed FeSiO-MS nanocomposites as integrated and eco-friendly photocatalysts exhibit enormous potentials for environmental and hygienic application.

[Isolation PQQ biosynthesis gene cluster from Gluconobacter oxydans based on sorbose-dehydrogenase activity].

OBJECTIVE: To isolate PQQ biosynthesis gene cluster from Gluconobacter oxydans H24 based on sorbose-dehydrogenase activity. METHODS: A library of Gluconobacter oxydans H24 genomic DNA was constructed with host strains Escherichia coli JM109s, which was integrated of sdh gene at the ptsG site on the chromosome of JM109. By detecting sorbose-dehydrogenase activity, clone of PQQ biosynthesis was isolated and subcloned. RESULTS: A positive clone was isolated from Gluconobacter oxydans H24 genomic DNA library. Within the 5,400-base-pair DNA fragment five reading frames are presented, corresponding to five of the pqq genes (pqqABCDE). The nucleotide and amino acid sequence showed highly homology to pqq genes of other bacteria. CONCLUSION: The pqqABCDE gene cluster was successfully isolated from Gluconobacter oxydans H24 by sorbose dehydrogenase activity.

Designing orodispersible films containing everolimus for enhanced compliance and bioavailability.

INTRODUCTION: Everolimus (EVR) has been approved for the treatment of various advanced cancers and its indications are increasingly expanding. Therefore, it is crucial for patients who have difficulty in swallowing, such as pediatric and elderly patients, to obtain a convenient formulation. The oral absorption of EVR is limited due to its low solubility in water, intestinal metabolism by CYP3A4 enzyme, P-gp-mediated efflux, and metabolism in the liver. The aim of this study was to develop a novel sublingual orodispersible film loading everolimus for improving patient compliance and enhancing oral bioavailability of EVR. RESEARCH DESIGN AND METHODS: Sublingual orodispersible films loading EVR were prepared by the solvent casting method and evaluated by in vitro and in vivo studies. RESULTS: The properties of films were determined by scanning electron microscopy, differential scanning calorimetry, X-ray diffraction, and Fourier-transform infrared spectrometry. The addition of acacia gum appeared to be crucial for protecting the drug from oxidation. Pharmacokinetic studies showed that loading into the sublingual orodispersible films significantly increased the oral bioavailability of EVR. CONCLUSION: The EVR-loaded sublingual orodispersible films are a promising, economical, and convenient approach for delivering EVR efficiently in a solid dosage form.

Reduction of liver tumor necrosis factor-alpha expression by targeting delivery of antisense oligonucleotides into Kupffer cells protects rats from fulminant hepatitis.

BACKGROUND: Fulminant liver failure can cause extreme mortality due to the lack of effective and targeting therapeutics for the disease. Novel therapeutics using antisense technology require an efficient and safe delivery system with Kupffer cell targeting ability. METHODS: We explored the capacity of galactosylated low molecular weight chitosan (GLC) to efficiently mediate the antisense oligonucleotide (ASO) TJU-2755 into Kupffer cells, enhance the effect of the oligonucleotides on the suppression of tumor necrosis factor (TNF)-alpha and prolong the active time of the antisense drug in vivo. The protective and therapeutic effect of ASO/GLC in the animal model of D-galactosamine/lipopolysaccharide-induced fulminant hepatitis was tested. RESULTS: ASOs delivered by GLC were concentrated in Kupffer cells and more potent in reducing the expression of TNF-alpha mRNA, as well as reducing serum TNF-alpha levels. Furthermore, the ASO/GLC complex successfully rescued animals from fulminant hepatitis and mortality. Compared to naked ASO, the complex notably reduced the dose administrated in animals and prolonged its effectiveness. A single dose of 5 mg ASO per kg body weight achieved a satisfactory effect after 5 days, and 20 mg ASO per kg body weight preserved 70% of the effect after more than 2 weeks. Its efficacy was affirmed through both pretreatment and therapeutic use after liver damage had begun. CONCLUSIONS: Inhibiting TNF-alpha expression in the liver by this strategy represents a novel therapeutic approach that may be valuable for the treatment of some inflammation-related liver diseases.

From form to function: the ways to know a neuron.

The shape of a neuron, its morphological signature, dictates the neuron's function by establishing its synaptic partnerships. Here, we review various anatomical methods used to reveal neuron shape and the contributions these have made to our current understanding of neural function in the Drosophila brain, especially the optic lobe. These methods, including Golgi impregnation, genetic reporters, and electron microscopy (EM), necessarily incorporate biases of various sorts that are easy to overlook, but that filter the morphological signatures we see. Nonetheless, the application of these methods to the optic lobe has led to reassuringly congruent findings on the number and shapes of neurons and their connection patterns, indicating that morphological classes are actually genetic classes. Genetic methods using, especially, GAL4 drivers and associated reporters have largely superceded classical Golgi methods for cellular analyses and, moreover, allow the manipulation of neuronal activity, thus enabling us to establish a bridge between morphological studies and functional ones. While serial-EM reconstruction remains the only reliable, albeit labor-intensive, method to determine actual synaptic connections, genetic approaches in combination with EM or high-resolution light microscopic techniques are promising methods for the rapid determination of synaptic circuit function.

NGALR is overexpressed and regulated by hypomethylation in esophageal squamous cell carcinoma.

PURPOSE: Neutrophil gelatinase-associated lipocalin receptor (NGALR) mRNA level is reduced in isolated chronic myelogenous leukemia blasts but up-regulated in esophageal squamous cell carcinoma (ESCC). The mechanism of NGALR regulation is unknown. Here, we show the expression pattern of NGALR and examine the aberrant methylation of its gene in ESCC and esophageal carcinoma cell lines. EXPERIMENTAL DESIGN: The expression pattern of NGALR was analyzed by immunohistochemistry in 59 ESCCs and compared with noncancerous tissues. The DNA methylation status was investigated by methylation-specific PCR and by bisulfite genomic sequencing in esophageal carcinoma cell lines and surgically resected samples. Methylated cell lines were treated with a methylation inhibitor to restore NGALR expression. RESULTS: The expression of NGALR in ESCC was significantly higher in tumor cell membrane and cytoplasm than in normal esophageal epithelium (P < 0.01). Methylated alleles were detected in three NGALR-nonexpressing cell lines but were not detected in three NGALR-expressing cell lines. Treatment of methylated cell lines with 5-aza-2'-deoxycytidine, a methylation inhibitor, restored NGALR expression. In surgically resected samples, 31 of 77 (40.3%) primary esophageal carcinomas and 46 of 77 (59.7%) paired normal tissues contained methylated NGALR alleles (P < 0.05). CONCLUSIONS: Our results suggest that NGALR hypomethylation contributes to its expression in esophageal carcinomas and that this overexpression may play a role in the pathogenesis of esophageal carcinomas.