Purinergic Astrocyte Signaling Driven by TNF-α After Cannabidiol Administration Restores Normal Synaptic Remodeling Following Traumatic Brain Injury.

Traumatic brain injury (TBI) is a prevalent form of cranial trauma that results in neural conduction disruptions and damage to synaptic structures and functions. Cannabidiol (CBD), a primary derivative from plant-based cannabinoids, exhibits a range of beneficial effects, including analgesic, sedative, anti-inflammatory, anticonvulsant, anti-anxiety, anti-apoptotic, and neuroprotective properties. Nevertheless, the effects of synaptic reconstruction and the mechanisms underlying these effects remain poorly understood. TBI is characterized by increased levels of tumor necrosis factor-alpha (TNF-α), a cytokine integral for the modulation of glutamate release by astrocytes. In the present study, the potential of CBD in regulating aberrant glutamate signal transmission in astrocytes following brain injury, as well as the underlying mechanisms involved, were investigated using immunofluorescence double staining, enzyme-linked immunosorbent assay (ELISA), western blot analysis, hematoxylin and eosin (H&E) staining, Nissl staining, transmission electron microscopy, and RT-qPCR. In this study, we examined the impact of CBD on neuronal synapses, focusing on the TNF-α-driven purinergic signaling pathway. Specifically, our research revealed that CBD pretreatment effectively reduced the secretion of TNF-α induced by astrocyte activation following TBI. This reduction inhibited the interaction between TNF-α and P2Y1 receptors, leading to a decrease in the release of neurotransmitters, including Ca2+ and glutamate, thereby initiating synaptic remodeling. Our study showed that CBD exhibits significant therapeutic potential for TBI-related synaptic dysfunction, offering valuable insights for future research and more effective TBI treatments. Further exploration of the potential applications of CBD in neuroprotection is required to develop innovative clinical strategies.

DLGAP1-AS2 promotes human colorectal cancer progression through trans-activation of Myc.

Substantial evidence suggests that non-coding RNA plays a vital role in human cancer, especially long non-coding RNA (lncRNA) with a length greater than 200nt. Herein, we found a lncRNA facilitating human colorectal cancer (CRC) progression. DLGAP1-AS2 was significantly increased in CRC tissues and cell lines. Knockdown of DLGAP1-AS2 inhibited CRC cell proliferation, migration, invasion in vitro, and tumor growth in vivo. The subcellular localization of DLGAP1-AS2 was translocated from the cytoplasm of normal cells to the nucleus of CRC cells due to reduced levels of N6-methyladenosine (m6A) modification. Further, through the screening of a series of signal pathways, we found that Myc pathway was involved in the effect of DLGAP1-AS2. Silencing of DLGAP1-AS2 markedly reduced Myc mRNA and protein levels. Blockade of Myc effectively abolished the enhanced aggressive behaviors of CRC cells caused by DLGAP1-AS2 overexpression. Mechanistically, DLGAP1-AS2 directly bound CTCF, a well-known transcriptional repressor of Myc, resulting in reduced binding of CTCF on Myc promoter and activating Myc transcription. The second hairpin structure of DLGAP1-AS2 was critical for the interaction between DLGAP1-AS2 and CTCF in the nucleus. Taken together, our study reveals the oncogenic regulatory axis of DLGAP1-AS2/CTCF/Myc in CRC, implying a promising targeted therapy for clinical application.

Genomic Characterization of Two Escherichia fergusonii Isolates Harboring mcr-1 Gene From Farm Environment.

The prevalence and transmission of mobile colistin resistance (mcr) genes have led to a severe threat to humans and animals. Escherichia fergusonii is an emerging pathogen which is closely related to a variety of diseases. However, the report of mcr genes harboring E. fergusonii is still rare. One study in Brazil reported the E. fergusonii isolates with IncHI2-type plasmids harboring mcr-1. A Chinese study reported two strains carrying mcr-1 gene with the same plasmid type IncI2. Here, we identified two strains of E. fergusonii carrying mcr-1 gene from farm environments with IncX4-type and IncI2-type plasmids, respectively. To our best knowledge, this is the first report about mcr-1 gene located on IncX4-type plasmid in E. fergusonii. We investigate the resistance mechanism of colistin-resistant Escherichia fergusonii strains 6S41-1 and 5ZF15-2-1 and elucidate the genetic context of plasmids carrying mcr-1 genes. In addition, we also investigated chromosomal mutations mediated colistin resistance in these two strains. Species identification was performed using MALDI-TOF MS and 16S rRNA gene sequencing. The detection of mcr-1 gene was determined by PCR and Sanger sequencing. S1-pulsed-field gel electrophoresis (PFGE), Southern blotting, antimicrobial susceptibility testing, conjugation experiments, complete genome sequencing, and core genome analysis were conducted to investigate the characteristics of isolates harboring mcr-1. The mcr-1 genes on two strains were both plasmids encoded and the typical IS26-parA-mcr-1-pap2 cassette was identified in p6S41-1 while a nikA-nikB-mcr-1 locus sites on the conjugative plasmid p5ZF15-2-1. In addition, Core genome analysis reveals that E. fergusonii 6S41-1 and 5ZF15-2-1 have close genetic relationships. The mcr-1 gene is located on conjugative IncI2-type plasmid p5ZF15-2-1, which provides support for its further transmission. In addition, there's the possibility of mcr-1 spreading to humans through farm environments and thereby threatening public health. Therefore, continuous monitoring and investigations of mcr-1 among Enterobacteriaceae in farm environments are necessary to control the spread.

Emergence and Genetic Characterization of Plasmid-Encoded VIM-2-Producing Pseudomonas stutzeri with Novel Integron In1998 Isolated from Cerebrospinal Fluid.

PURPOSE: To investigate the genomic and plasmid characteristics of a newly discovered Pseudomonas stutzeri strain with a bla VIM-2-carrying plasmid and novel integron In1998 isolated from a cerebrospinal fluid specimen in a teaching hospital. METHODS: Species identification was performed by MALDI-TOF MS, and bla VIM-2 was identified by PCR and Sanger sequencing. Whole-genome sequencing analysis was conducted using the Illumina NovaSeq 6000 and Oxford Nanopore platforms. Integron detection was performed using INTEGRALL. The phylogenetic tree was constructed by using kSNP3.0. Plasmid characteristics were assessed by S1-pulsed-field gel electrophoresis (S1-PFGE), Southern blotting, conjugation experiments, and whole-genome sequencing analysis. Comparative genomics analysis of the plasmid and genetic context of bla VIM-2 were conducted by using BLAST Ring Image Generator (BRIG) and Easyfig 2.3, respectively. RESULTS: ZDHY95, an MDR strain of P. stutzeri harboring bla VIM-2, was identified. It was sensitive only to amikacin and was resistant to carbapenems, ß-lactams, aztreonam, fluoroquinolones, and aminoglycosides. Joint S1-PFGE, Southern blot, conjugation assay, and whole-genome sequencing experiments confirmed that the bla VIM-2 gene was located within class I integron In1722 of the plasmid and that the surrounding genetic environment was 5'CS-aacA4'-30-bla VIM-2-aacA4'-3'CS. The novel class I integron In1998 was detected on the chromosome of P. stutzeri ZDHY95, and the gene cassette array was 5'CS-aacA3-aadA13-cmlA8-bla OXA-246-arr3-dfrA27-3'CS. Phylogenetic analysis showed that antimicrobial resistance gene-carrying P. stutzeri isolates were divided into two clusters, mainly containing isolates from the USA and Pakistan. CONCLUSION: A novel bla VIM-2-carrying conjugative plasmid, pZDHY95-VIM-2, was reported for the first time in P. stutzeri, elucidating the genetic environment and transfer mechanism. The gene structure of the novel class I integron In1998 was also clarified. We explored the phylogenetic relationship of P. stutzeri with drug resistance genes and suggested that Pseudomonas with metallo-ß-lactamases (MBLs) in the hospital environment may cause infection in patients with long-term intubation or after interventional surgery.

Effects of cannabinoid (CBD) on blood brain barrier permeability after brain injury in rats.

Cannabidiol is a natural herbal medicine known to protect the brain from traumatic brain injury (TBI). Here, a TBI rat model was established, with cannabidiol administered intraperitoneally at doses of 5, 10, or 20 mg/kg, 30 min before surgery and 6 h after surgery until sacrifice. Brain water content, body weight, and modified neurological severity scores were determined, and enzyme-linked immunosorbent assay, immunofluorescence staining, hematoxylin and eosin staining, Nissl staining, Evans-blue dye extravasation, and western blotting were performed. Results showed that cannabidiol decreased the number of aquaporin-4-positive and glial fibrillary acidic protein-positive cells. Cannabidiol also significantly reduced the protein levels of proinflammatory cytokines (TNF-α and IL-1ß) and significantly increased the expression of tight junction proteins (claudin-5 and occludin). Moreover, cannabidiol administration significantly mitigated water content in the brain after TBI and blood-brain barrier disruption and ameliorated the neurological deficit score after TBI. Cannabidiol administration improved the integrity and permeability of the blood-brain barrier and reduced edema in the brain after TBI.

Oral Microbiome in Patients with Oesophageal Squamous Cell Carcinoma.

To investigate the oral microflora of patients with oesophageal squamous cell carcinoma (ESCC), saliva samples were collected from 20 patients with ESCC and 21 healthy controls. The V3-V4 region of 16S rDNA was amplified and sequenced by the Illumina MiSeq high-throughput sequencing platform. The final sequences were used for OTU analysis. Alpha and beta diversity analysis showed that the bacterial diversity and richness of the ESCC group were lower than those of the control group, while the variability of the ESCC group was higher than that of the control group. According to the Metastats difference analysis and LEfSe analysis, the high risk of ESCC may be related to Actinomyces and Atopobium, while the healthy control group is closely related to Fusobacterium and Porphyromonas (the analysis was performed at the genus level). The establishment of the relationship between oral microbiota and risk of ESCC may lead to significant advances in understanding the aetiology of cancer and may open a new research paradigm for cancer prevention.

miR­378a­3p inhibits cellular proliferation and migration in glioblastoma multiforme by targeting tetraspanin 17.

Glioblastoma multiforme (GBM) is the most common and aggressive brain tumor and patients with this disease tend to have poor clinical outcome. MicroRNAs (miRs) are important regulators of a number of key pathways implicated in tumor pathogenesis. Recently, the expression of miR­378 was shown to be dysregulated in several different types of cancer, including gastric cancer, colorectal cancer and oral carcinoma. Additional studies have demonstrated that miR­378 may serve as a potential therapeutic target against human breast cancer. However, the underlying mechanisms and potential targets of miR­378a­3p involved in GBM remain unknown. The aim of the present of was to determine the effects of miR­378a­3p and its potential targets. Tetraspanin 17 (TSPAN17) is involved in the neoplastic events in GBM and is a member of the tetraspanin family of proteins. The tetraspanins are involved in the regulation of cell growth, migration and invasion of several different types of cancer cell lines, and may potentially act as an oncogene associated with GBM pathology. The results of the present study showed that high miR­378a­3p and low TSPAN17 expression levels were associated with improved survival in patients with GBM. Additionally, high levels of TSPAN17 were linked to the poor prognosis of patients with GBM aged 50­60, larger tumor sizes (≥5 cm) and an advanced World Health Organization stage. TSPAN17 was identified and confirmed as a direct target of miR­378a­3p using a luciferase reporter assay in human glioma cell lines. Overexpression of miR­378a­3p in either of U87MG or MT­330 cells decreased the expression of TSPAN17, promoted apoptosis and decreased proliferation, migration and invasion. Overexpression of TSPAN17 attenuated the aforementioned effects induced by miR­378a­3p overexpression. The present study indicated that miR­378a­3p suppresses the progression of GBM by reducing TSPAN17 expression, and may thus serve as a potential therapeutic target for treating patients with GBM.

Detection and Genomic Characterization of a Morganella morganii Isolate From China That Produces NDM-5.

The increasing prevalence and transmission of the carbapenem resistance gene bla NDM-5 has led to a severe threat to public health. So far, bla NDM-5 has been widely detected in various species of Enterobacterales and different hosts across various cities. However, there is no report on the bla NDM-5- harboring Morganella morganii. In January 2016, the first NDM-5-producing Morganella morganii L241 was found in a stool sample of a patient diagnosed as recurrence of liver cancer in China. Identification of the species was performed using 16S rRNA gene sequencing. Carbapenemase genes were identified through both PCR and sequencing. To investigate the characteristics and complete genome sequence of the bla NDM-5-harboring clinical isolate, antimicrobial susceptibility testing, S1 nuclease pulsed field gel electrophoresis, Southern blotting, transconjugation experiment, complete genome sequencing, and comparative genomic analysis were performed. M. morganii L241 was found to be resistant to broad-spectrum cephalosporins and carbapenems. The complete genome of L241 is made up from both a 3,850,444 bp circular chromosome and a 46,161 bp self-transmissible IncX3 plasmid encoding bla NDM-5, which shared a conserved genetic context of bla NDM-5 (ΔIS3000-ΔISAba125-IS5-bla NDM-5-ble-trpF-dsbC-IS26). BLASTn analysis showed that IncX3 plasmids harboring bla NDM genes have been found in 15 species among Enterobacterales from 13 different countries around the world thus far. In addition, comparative genomic analysis showed that M. morganii L241 exhibits a close relationship to M. morganii subsp. morganii KT with 107 SNPs. Our research demonstrated that IncX3 is a key element in the worldwide dissemination of bla NDM-5 among various species. Further research will be necessary to control and prevent the spread of such plasmids.

Sex hormones play roles in determining musk composition during the early stages of musk secretion by musk deer (Moschus berezovskii).

Musk is a secreted external hormone or information compound that is stored in musk scent glands of the males of species within the family Moschidae, such as Moschus berezovskii. The secretion of musk changes periodically during the courtship and reproduction periods, with the early stage of secretion occurring from May to July, and the maturation stage occurring from August to April of the following year. In this study, we analyzed the dynamic changes in musk components from June to April of the following year. The result showed that musk morphological character, water content, total ion chromatographic pattern, and composition undergo seasonal change. Luminescence immunoassay and radioimmunoassay analyses were performed to determine corresponding fecal hormone levels. The results showed that testosterone, estrogen, and cortisol levels in feces change on a seasonal basis, and are significantly higher in June than in other months (p < 0.01). Correlation analysis showed that the contents of four examined musk components (muscone, cyclopentadecanone, cholesterol, and cholestenol) from June to August were significantly highly negatively correlated with fecal testosterone and estradiol levels (p < 0.01). In contrast, the correlation coefficients were low or not significant from August to April of the following year. These results indicate that testosterone and estradiol may play a major role in determining musk composition during the early stage of musk secretion but not during the course of musk maturation, which suggests that musk secretion may be promoted by increases in sex hormones in June.

Lactobacilli inhibit cervical cancer cell migration in vitro and reduce tumor burden in vivo through upregulation of E-cadherin.

The present study was designed to investigate the antitumor effects of Lactobacillus and the potential mechanisms. Cell Counting Kit-8 (CCK-8) assays were carried out to determine suitable doses for investigating the inhibitory effect of lactobacilli on cell migration ability of HeLa and U14 cells in vitro. In addition, western blot assays were performed to investigate the possible mechanisms corresponding to its antitumor effects. Furthermore, a xenograft mouse model was established for investigating the E-cadherin expression in tumor tissues after treatment with lactobacilli. Our results showed that live lactobacilli [multiplicity of infection (MOI) of 1,000:1] significantly possessed inhibitory effects on cell migration ability of cervical cancer cells. Lactobacilli (MOI: 1,000:1) significantly upregulated E-cadherin expressions in HeLa and U14 cells (p<0.05). On the contrary, our results showed that inactivated lactobacilli could not affect the E-cadherin expression levels in HeLa and U14 cells. Similar to the western blot assay, immunohistochemistry results also indicated that lactobacilli treatment significantly upregulated E-cadherin in tumor tissues (p<0.05). In conclusion, our results above suggest that lactobacilli have the potential for inhibiting the migratory ability of cervical cancer cell lines, and the possible pharmacological mechanism may be closely related to the upregulation of E-cadherin.

Inhibitory effects of oleoylethanolamide (OEA) on H2O2-induced human umbilical vein endothelial cell (HUVEC) injury and apolipoprotein E knockout (ApoE-/-) atherosclerotic mice.

Atherosclerosis (AS) is initiated by vascular endothelial cell injury, which is induced by lipid and protein oxidation. Oleoylethanolamide (OEA), a dietary fat-derived lipid, has shown atheroprotective effect. In vitro studies demonstrated that OEA showed cytoprotective effects on H2O2-induced primary cultured human umbilical vein endothelial cell (HUVEC) injury model. Further investigation of the cytoprotective effects of OEA demonstrated that OEA exerted its function by scavenging for reactive oxygen species, as well as increasing anti-oxidative enzymes, reducing lipid peroxidation, terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL)-positive cells and apoptosis-related proteins expression. The in vivo study using an ApoE-/- mouse model fed with high-fat diet for 8 weeks showed that OEA (10 mg/kg/day, i.g.) administration reduced blood lipid levels, prevented endothelial cell damage and inhibited early AS plaque formation. In conclusion, our results suggested that OEA exerted a pharmacological effect on ameliorating atherosclerotic plaque formation through the inhibition of oxidative stress-induced endothelial cell injury and therefore OEA can be a potential candidate drug for anti-atherosclerosis.

Trichostatin A, a histone deacetylase inhibitor, suppresses proliferation and promotes apoptosis of esophageal squamous cell lines.

Histone deacetylase (HDAC)­mediated epigenetic modification plays crucial roles in numerous biological processes, including cell cycle regulation, cell proliferation and apoptosis. HDAC inhibitors demonstrate antitumor effects in various cancers, including glioblastoma and breast cancer. HDAC inhibitors are therefore promising antitumor drugs for these tumors. The tumorigenesis and development of esophageal squamous cell carcinoma (ESCC) involve genetic and epigenetic mechanisms. However, the effects of the HDAC inhibitor on ESCC are not fully investigated. In the present study, ESCC cells were treated with trichostatin A (TSA) and its antitumor effects and related mechanisms were investigated. The results indicated that TSA suppressed the proliferation of ESCCs and caused G1 phase arrest by inducing the expression of p21 and p27. TSA also induced cell apoptosis by enhancing the expression of pro­apoptotic protein Bax and decreasing the expression of anti­apoptotic protein Bcl­2. Furthermore, TSA inhibited the expression of phosphatidylinositol­3­kinase (PI3K) and reduced the phosphorylation of Akt and extracellular signal­regulated kinase (ERK)1/2 in EC9706 and EC1 cell lines. High levels of acetylated histone H4 were detected in TSA­treated ESCC cell lines. Overall, these results indicate that TSA suppresses ESCC cell growth by inhibiting the activation of the PI3K/Akt and ERK1/2 pathways. TSA also promotes cell apoptosis through epigenetic regulation of the expression of apoptosis­related protein.

Both enalapril and losartan attenuate sarcolemmal Na+-K+-ATPase remodeling in failing rat heart due to myocardial infarction.

To investigate the mechanisms underlying the depressed sarcolemmal (SL) Na(+)-K(+)-ATPase activity in congestive heart failure (CHF), different isoforms and gene expression of Na(+)-K(+)-ATPase were examined in the failing left ventricle (LV) at 8 weeks after myocardial infarction (MI). In view of the increased activity of renin-angiotensin system (RAS) in CHF, these parameters were also studied after 5 weeks of treatment with enalapril (10 mg x kg-1 x day-1), an angiotensin-converting enzyme inhibitor, and losartan (20 mg.kg-1.day-1), an angiotensin II type 1 receptor antagonist, starting at 3 weeks after the coronary ligation in rats. The infarcted animals showed LV dysfunction and depressed SL Na(+)-K(+)-ATPase activity. Protein content and mRNA levels for Na(+)-K(+)-ATPase alpha2 isoform were decreased whereas those for Na(+)-K(+)-ATPase alpha3 isoform were increased in the failing LV. On the other hand, no significant changes were observed in protein content or mRNA levels for Na(+)-K(+)-ATPase alpha1 and beta1 isoforms. The treatment of infarcted animals with enalapril or losartan improved LV function and attenuated the depression in Na(+)-K(+)-ATPase alpha2 isoform as well as the increase in alpha3 isoform, at both the protein and mRNA levels; however, combination therapy with enalapril and losartan did not produce any additive effects. These results provide further evidence that CHF due to MI is associated with remodeling of SL membrane and suggest that the blockade of RAS plays an important role in preventing these alterations in the failing heart.

Modification of myosin protein and gene expression in failing hearts due to myocardial infarction by enalapril or losartan.

The effects of enalapril, an angiotensin converting enzyme (ACE) inhibitor, and losartan, an angiotensin II receptor type I antagonist, were investigated on alterations in myofibrillar ATPase activity as well as myosin heavy chain (MHC) content and gene expression in failing hearts following myocardial infarction (MI). Three weeks after ligation of the left coronary artery, rats were treated with or without enalapril (10 mg/kg/day), and/or losartan (20 mg/kg/day) for 5 weeks. The infarcted animals exhibited an increase in left ventricle (LV) end diastolic pressure and depressed rates of LV pressure development as well as pressure decay. LV myofibrillar Ca2+ -stimulated ATPase activity was decreased in the infarcted hearts compared with controls, MHC alpha-isoform content was significantly decreased whereas that of MHC beta-isoform was markedly increased. The level of MHC alpha-isoform mRNA was decreased whereas that of MHC beta-isoform was increased in the viable infarcted LV. Treatment of animal with enalapril, losartan, or combination of enalapril and losartan partially prevented the MI induced changes in LV function, myofibrillar Ca2+ -stimulated ATPase activity, MHC protein expression and MHC gene expression. The results suggest that the beneficial effects of the renin-angiotensin system blockade in heart failure are associated with partial prevention of myofibrillar remodeling.

Partial prevention of changes in SR gene expression in congestive heart failure due to myocardial infarction by enalapril or losartan.

Although activation of the renin-angiotensin system (RAS) is known to produce ventricular remodeling and congestive heart failure (CHF), its role in inducing changes in the sarcoplasmic reticulum (SR) protein and gene expression in CHF is not fully understood. In this study, CHF was induced in rats by ligation of the left coronary artery for 3 weeks and then the animals were treated orally with or without an angiotensin converting enzyme inhibitor, enalapril (10 mg/kg/day) or an angiotensin II receptor antagonist, losartan (20 mg/kg/day) for 4 weeks. Sham-operated animals were used as control. The animals were hemodynamically assessed and protein content as well as gene expression of SR Ca(2+)-release channel (ryanodine receptor, RYR), Ca(2+)-pump ATPase (SERCA2), phospholamban (PLB) and calsequestrin (CQS) were determined in the left ventricle (LV). The infarcted animals showed cardiac hypertrophy, lung congestion, depression in LV +dP/dt and -dP/dt, as well as increase in LV end diastolic pressure. Both protein content and mRNA levels for RYR, SERCA2 and PLB were decreased without any changes in CQS in the failing heart. These alterations in LV function as well as SR protein and gene expression in CHF were partially prevented by treatment with enalapril or losartan. The results suggest that partial improvement in LV function by enalapril and losartan treatments may be due to partial prevention of changes in SR protein and gene expression in CHF and that these effects may be due to blockade of the RAS.