Model Mice Research Articles

Cryoablation combined with dual immune checkpoint blockade enhances antitumor efficacy in hepatocellular carcinoma model mice

Background Cryoablation (Cryo) is a minimally invasive treatment for tumors. Cryo can activate the body’s immune response, although it is typically weak. The immune response induced by Cryo in hepatocellular carcinoma (HCC) is poorly understood. PD-1 and CTLA-4 monoclonal antibodies are immune checkpoint inhibitors used in immunotherapy for tumors. The combined use of these antibodies with Cryo may enhance the immune effect. Methods A Balb/c mouse model of HCC was established and treated with Cryo, immune checkpoint blockade (ICB), or Cryo + ICB (combination therapy). The growth trend of right untreated tumors and survival time of mice were determined. The expression of apoptosis-related proteins was detected by Western blot (WB) assay. The percentages of immune cells and immunosuppressive cells were analyzed by flow cytometry. The numbers of infiltrating T lymphocytes were checked by immunohistochemistry, and the levels of T-cell-associated cytokines were detected by Quantitative real-time Polymerase Chain Reaction (qRT-PCR) assays and Enzyme-Linked Immunosorbent Assays (ELISA) assays. Results Cryo + ICB inhibited the growth of right untreated tumors, promoted tumor cell apoptosis, and prolonged the survival time of mice. Local T-cell infiltration in right tumor tissues increased after the combination therapy, while the number of immunosuppressive cells was significantly reduced. In addition, the combination therapy may induce the production of multiple Th1-type cytokines but reduce the production of Th2-type cytokines. Conclusions Cryo can activate CD8+ and CD4+ T-cell immune responses. Cryo + ICB can relieve the immunosuppressive tumor microenvironment and shift the Th1/Th2 balance toward Th1 dominance, further enhancing the Cryo-induced T-cell immune response and resulting in a stronger antitumor immune response.

Temperature-sensitive nanogels combined with polyphosphate and cisplatin for the enhancement of tumor artery embolization by coagulation activation

Transcatheter arterial chemoembolization (TACE) is the first-line therapy for hepatocellular carcinoma (HCC). However, the exacerbated hypoxia microenvironment induces tumor relapse and metastasis post-TACE. Here, temperature-sensitive block polymer complexed with polyphosphate–cisplatin (Pt–P@PND) was prepared for the enhancement of tumor artery embolization by coagulation activation. After supra-selective infusion into the tumor vessels, Pt–P@PND nanogels performed efficient embolization of tumor arteries by sol–gel transition at body temperature. Meanwhile, coagulation cascade was evoked to form blood clots in the peripheral arteries inaccessible to the nanogels by released PolyP. The blood clots-filled hydrogel networks composed of gel and clots showed a denser structure and higher modulus, thereby achieving long-term embolization of all levels of tumor arteries. Pt–P@PND nanogels efficiently inhibited tumor growth and reduced the expression of HIF-1α, VEGF, CD31, and MMP-9 on VX2 tumor-bearing rabbit model. The released Nitro-Pt stimulated the immunogenic cell death of tumor cells, thus enhancing the antitumor immune response to suppress tumor relapse and metastasis post-TACE. It is hoped that Pt–P@PND nanogels can be developed as a promising embolic agent with procoagulant activity for enhancing the antitumor immune response through a combination of embolism, coagulation, and chemotherapy. Statement of significanceClinical embolic agents, such as Lipiodol and polyvinyl alcohol (PVA) microspheres, are limited by their rapid elimination or larger size, thus lead to incomplete embolization of trans-catheter arterial chemoembolization (TACE). Herein, temperature-sensitive Pt-P@PND nanogels were developed to achieve long-term embolization of all levels of tumor arteries by gel/clot generation. The released Nitro-Pt induced immunogenic cell death in tumor cells, which improved the antitumor immune microenvironment by the maturation of DCs and lymphocytic infiltration. Pt-P@PND nanogels successfully inhibited tumor growth and activated an antitumor immune response to curb the recurrence and metastasis of residual tumor cells both in VX2 tumor-bearing rabbit model and 4T1 tumor-bearing mouse model. These findings suggested that Pt-P@PND could be developed as an ideal embolic agent for clinical TACE treatment.

Effect of Diets with the Addition of Edible Insects on the Development of Atherosclerotic Lesions in ApoE/LDLR-/- Mice.

Foods enriched with insects can potentially prevent several health disorders, including cardiovascular diseases, by reducing inflammation and improving antioxidant status. In this study, Tenebrio molitor and Gryllus assimilis were selected to determine the effect on the development of atherosclerosis in ApoE/LDLR-/- mice. Animals were fed AIN-93G-based diets (control) with 10% Tenebrio molitor (TM) and 10% Gryllus assimilis (GA) for 8 weeks. The nutritional value as well as antioxidant activity of selected insects were determined. The lipid profile, liver enzyme activity, and the fatty acid composition of liver and adipose tissue of model mice were evaluated. Quantitative analysis of atherosclerotic lesions in the entire aorta was performed using the en face method, and for aortic roots, the cross-section method was used. The antioxidant status of the GA cricket was significantly higher compared to the TM larvae. The results showed that the area of atherosclerosis (en face method) was not significantly different between groups. Dietary GA reduced plaque formation in the aortic root; additionally, significant differences were observed in sections at 200 and 300 µm compared to other groups. Furthermore, liver enzyme ALT activity was lower in insect-fed groups compared to the control group. The finding suggests that a diet containing edible insect GA potentially prevents atherosclerotic plaque development in the aortic root, due to its high antioxidant activity.

A classic prescription alleviates inflammation in CUMS model mice via modulating MYDGF/MAP4K4/NF-κB signaling pathway, verified through UPLC-HRMS and proteomics analysis

BackgroundXiaoyaosan (XYS), a renowned classical traditional Chinese medicinal formula utilized in addressing major depressive disorder (MDD), has garnered significant acclaim for its remarkable efficacy in clinical application. The onset of major depressive disorder (MDD) often correlates with chronic unpredictable mild stress (CUMS), a pivotal instigating factor in its development.Aim of the study: This study aims to clarify the potential anti-inflammatory mechanisms of XYS in treating CUMS model mice. Materials and methodsUtilizing cutting-edge ultra high-performance liquid chromatography - high-resolution mass spectrometry (UPLC-HRMS), the active constituents of XYS were discerned, while employing proteomics analysis to delve into the potential mechanisms of its efficacy. Molecular docking studies, alongside subsequent in vivo experiments utilizing CUMS model mice, were conducted to corroborate the findings derived from the proteomics analysis. ResultsIn vivo experiments demonstrated that XYS not only markedly ameliorated behavioral markers but also attenuated serum inflammatory markers and suppressed IL-6 and TNF-α expression within the brains of CUMS model mice. Proteomics analysis suggested that the pivotal anti-inflammatory mechanism of XYS against CUMS-induced damage might involve modulation of the MAPK signaling pathway. Utilizing UPLC-HRMS, the active constituents of XYS were successfully identified, while molecular docking investigations explored interactions between XYS and MYDGF, PKC, MAP4K4, P-p65, p65, P-IKBα, and IKBα. The findings revealed XYS's regulatory influence on the MYDGF/MAP4K4/NF-κB signaling cascade. ConclusionsThis study is the first to our knowledge to demonstrate that XYS can alleviate inflammation in CUMS model mice by modulating the MYDGF/MAP4K4/NF-κB signaling pathway.

M6A mRNA methylation-mediated MAPK signaling modulates the nasal mucosa inflammatory response in allergic rhinitis.

Allergic rhinitis (AR) is a complex disease in which gene-environment interactions contribute to its pathogenesis. Epigenetic modifications, such as N6-methyladenosine (m6A) modification of mRNA, play important roles in regulating gene expression in multiple physiological and pathological processes. However, the function of m6A modification in AR and the inflammatory response is poorly understood. We used the ovalbumin (OVA) and aluminum hydroxide to induce an AR mouse model. Nasal symptoms, histopathology, and serum cytokines were examined. We performed combined m6A and RNA sequencing to analyze changes in m6A modification profiles. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and methylated RNA immunoprecipitation sequencing qPCR (MeRIP-qPCR) were used to verify differential methylation of mRNAs and the m6A methylation level. Knockdown or inhibition of Alkbh5 in nasal mucosa of mice was mediated by lentiviral infection or IOX1 treatment. We showed that m6A was enriched in a group of genes involved in MAPK signaling pathway. Moreover, we identified a MAPK pathway involving Map3k8, Erk2, and Nfκb1 that may play a role in the disrupted inflammatory response associated with nasal inflammation. The m6A eraser, Alkbh5, was highly expressed in the nasal mucosa of AR model mice. Furthermore, knockdown of Alkbh5 expression by lentiviral infection resulted in high MAPK pathway activity and a significant nasal mucosa inflammatory response. Our findings indicate that ALKBH5-mediated m6A dysregulation likely contributes to a nasal inflammatory response via the MAPK pathway. Together, our data show that m6A dysregulation mediated by ALKBH5, is likely to contribute to inflammation of the nasal mucosa via the MAPK signaling pathway, suggesting that ALKBH5 is a potential biomarker for AR treatment.

Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-ĸB pathway

Radiation-induced thrombocytopenia (RIT) faces a perplexing challenge in the clinical treatment of cancer patients, and current therapeutic approaches are inadequate in the clinical settings. In this researsh, oxymatrine, a new molecule capable of healing RIT was screened out, and the underlying regulatory mechanism associated with magakaryocyte (MK) differentiation and thrombopoiesis was demonstrated. The capacity of oxymatrine to induce MK differentiation was verified in K-562 and Meg-01 cells in vitro. The ability to induce thrombopoiesis was subsequently demonstrated in Tg (cd41:eGFP) zebrafish and RIT model mice. In addition, we carried out network pharmacological prediction, drug affinity responsive target stability assay (DARTS) and cellular thermal shift assay (CETSA) analyses to explore the potential targets of oxymatrine. Moreover, the pathway underlying the effects of oxymatrine was determined by Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, Western blot, and immunofluorescence. Oxymatrine markedly promoted MK differentiation and maturation in vitro. Moreover, oxymatrine induced thrombopoiesis in Tg (cd41:eGFP) zebrafish and accelerated thrombopoiesis and platelet function recovery in RIT model mice. Mechanistically, oxymatrine directly binds to toll-like receptor 2 (TLR2) and further regulates the downstream pathway stimulator of interferon genes (STING)/nuclear factor-kappaB (NF-κB), which can be blocked by C29 and C-176, which are specific inhibitors of TLR2 and STING, respectively. Taken together, we demonstrated that oxymatrine , a novel TLR2 agonist, plays a critical role in accelerating MK differentiation and thrombopoiesis via the STING/NF-κB axis, suggesting that oxymatrine is a promising candidate for RIT therapy.

Mairin from Huangqi Decoction Mitigates Liver Cirrhosis through Suppression of Pro-inflammatory Signaling Pathways: A Network Pharmacology and Experimental Study

Background:: Liver cirrhosis is a consequence of various chronic liver conditions and may lead to liver failure and cancer. Huangqi Decoction (HQD) is a Traditional Chinese Medicine (TCM) effective for treating liver conditions, including cirrhosis. Therefore, both the active ingredients and the pharmacological actions of HQD deserve further exploration. The active components and pharmacological actions of HQD in preventing and treating liver cirrhosis were investigated using network pharmacology. The actions of the principal active ingredient, Mairin, were investigated empirically. Methods:: Using network pharmacology, the critical components of HQD were identified from multiple databases, and UPLC screening and targets were investigated using Swiss Target Prediction. Targets associated with liver cirrhosis were identified using the GeneCards database. GO and KEGG enrichment analysis of targets that overlapped between HQD and cirrhosis were analyzed in DAVID, and a “component-target-pathway” network for HQD was created in Cytoscape 3.7.2. The biological functions of the key active component, Mairin, were investigated using in silico docking, cell experiments, and evaluation in a carbon-tetrachloride (CCl4)-induced mouse model of liver cirrhosis. CCK-8 and F-actin assays were used to measure cell viability and hepatic stellate cell (HSC) activation, respectively; fibrosis was measured by histological and immunohistochemical evaluations, and the levels of the cirrhosis-related protein α-SMA and predicted essential target proteins in the PI3KAKT, NFκB-IκBα, and NLRP3-IL18 pathways were determined by western blotting. Results:: Fourteen active HQD components, 72 targets, and 10 pathways common to HQD and cirrhosis were identified. Network analysis indicated the association of Mairin with most targets and with inflammation through the PI3K/Akt, NF-κB, and NLRP3 pathways. Dose-dependent reductions in the activation and proliferation of LX-2 cells after Mairin treatment were observed. Mairin reversed the histopathological changes in the livers of cirrhosis model mice. Mairin also significantly reduced the α-SMA, NF-κB, IκBα, NLRP3, and IL-18 protein levels while increasing those of p- PI3K and p-Akt, suggesting that Mairin mitigates liver cirrhosis through modulation of the PI3KAKT, NFκB-IκBα, and NLRP3-IL18 pathways. Conclusions:: Using a comprehensive investigative process involving network pharmacology, bioinformatics, and experimental verification, it was found that Mairin, an active component of HQD, may be useful for developing specific treatments for preventing and treating liver cirrhosis.

FGFR3 drives Aβ-induced tau uptake

The amyloid cascade hypothesis suggests that amyloid beta (Aβ) contributes to initiating subsequent tau pathology in Alzheimer’s disease (AD). However, the underlying mechanisms through which Aβ contributes to tau uptake and propagation remain poorly understood. Here, we show that preexisting amyloid pathology accelerates the uptake of extracellular tau into neurons. Using quantitative proteomic analysis of endocytic vesicles, we reveal that Aβ induces the internalization of fibroblast growth factor receptor 3 (FGFR3). Extracellular tau binds to the extracellular domain of FGFR3 and is internalized by the FGFR3 ligand, fibroblast growth factor 2 (FGF2). Aβ accelerates FGF2 secretion from neurons, thereby inducing the internalization of tau-attached FGFR3. Knockdown of FGFR3 in the hippocampus reduces tau aggregation by decreasing tau uptake and improving memory function in AD model mice. These data suggest FGFR3 in neurons as a novel tau receptor and a key mediator of Aβ-induced tau uptake in AD.

Auditory brainstem response audiometry in tauopathy mouse model of human Alzheimer's disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder in which changes in hearing sensitivity precede cognitive decline. Despite a well-known link between dementia and hearing loss, few AD model mouse lines have hearing characterized. We screened for hearing loss using auditory brainstem responses (ABR) in young (3-4 months) and aging (9-10 months) mice with a P301S tauopathy (PS19 mice). Compared to wild types, aging PS19 mice did not show accelerated hearing loss but did show latency differences in centrally generated ABR waveform components. These results suggest that tauopathy causes mild central auditory dysfunction in the absence of overt hearing loss.

LncRNA MAAMT facilitates macrophage recruitment and proinflammatory activation and exacerbates autoimmune myocarditis through the SRSF1/NF-κB axis

Long non-coding RNAs (lncRNAs) have been implicated in dilated cardiomyopathy (DCM). However, the biological functions and regulatory mechanisms of lncRNAs in DCM remain elusive. Using a mouse model of experimental autoimmune myocarditis (EAM) to mimic DCM, we successfully constructed a dynamic lncRNA expression library for EAM by lncRNA microarray and found that the expression of a macrophage-enriched lncRNA, MAAMT, was significantly increased in the myocardial tissue of mice at the acute stage of EAM. Functionally, MAAMT knockdown alleviated the recruitment and proinflammatory activation of macrophages in the heart, spleen, and peripheral blood of mice at the acute stage of EAM, reduced myocardial inflammation and injury, and eventually reversed ventricular remodelling and improved cardiac function in mice at the chronic stage of EAM. Mechanistically, we identified serine/arginine-rich splicing factor 1 (SRSF1) as an MAAMT-interacting protein in macrophages using RNA pull-down assays coupled with mass spectrometry. MAAMT knockdown attenuated the ubiquitination-mediated degradation of SRSF1, increased the protein expression of SRSF1, and restrained the activation of the NF-κB pathway in macrophages, thereby inhibiting the proinflammatory activation of macrophages. Collectively, our results demonstrate that MAAMT is a key proinflammatory regulator of myocarditis that promotes macrophage activation through the SRSF1-NF-κB axis, providing a new insight into early effective treatment strategies for DCM.

Shiikuwasha leaf and peel extracts inhibit allergic reactions by suppressing degranulation in RBL-2H3 rat basophilic leukemia cells and immunoglobulin production in mouse spleen lymphocytes.

This study aims to investigate the antiallergic effects of Shiikuwasha (Citrus depressa Hayata) leaf and peel extracts by examining the regulation of degranulation and inflammatory cytokine production from rat basophilic leukemia (RBL-2H3) cells and antigen-specific antibody production in sensitized mouse spleen lymphocytes. In vivo antiallergic activity was evaluated using the passive cutaneous anaphylaxis (PCA) reaction model. Extracts of Shiikuwasha leaves and peel were prepared using 80% methanol and dissolved in dimethylsulfoxide. The dinitrophenyl-human serum albumin-induced β-hexosaminidase levels in immunoglobulin (Ig) E-sensitized RBL-2H3 cells were assessed using enzymatic assays. Cytokine production was measured by enzyme-linked immunosorbent assay. Antibody production capacity was evaluated using lymphocytes isolated from spleens of type I allergy model mice. Lymphocytes were cultured for 72 h with Shiikuwasha extracts, and ovalbumin-specific IgE, IgG1, and IgG2a levels were measured. Shiikuwasha leaf and peel extract significantly reduced β-hexosaminidase release and suppressed interleukin-4 and tumor necrosis factor-α production from RBL-2H3 cells. Ovalbumin-specific IgE and IgG1 production decreased in Shiikuwasha extract-treated lymphocytes. These extracts also significantly suppressed the PCA reaction. Shiikuwasha leaf and peel extract reduce degranulation in RBL-2H3 cells and antibody production in spleen-derived lymphocytes and therefore exhibit antiallergic effects.

Production of Spinocerebellar Ataxia Type 3 Model Mice by Intravenous Injection of AAV-PHP.B Vectors.

We aimed to produce a mouse model of spinocerebellar ataxia type 3 (SCA3) using the mouse blood-brain barrier (BBB)-penetrating adeno-associated virus (AAV)-PHP.B. Four-to-five-week-old C57BL/6 mice received injections of high-dose (2.0 × 1011 vg/mouse) or low-dose (5.0 × 1010 vg/mouse) AAV-PHP.B encoding a SCA3 causative gene containing abnormally long 89 CAG repeats [ATXN3(Q89)] under the control of the ubiquitous chicken β-actin hybrid (CBh) promoter. Control mice received high doses of AAV-PHP.B encoding ATXN3 with non-pathogenic 15 CAG repeats [ATXN3(Q15)] or phosphate-buffered saline (PBS) alone. More than half of the mice injected with high doses of AAV-PHP.B encoding ATXN3(Q89) died within 4 weeks after the injection. No mice in other groups died during the 12-week observation period. Mice injected with low doses of AAV-PHP.B encoding ATXN3(Q89) exhibited progressive motor uncoordination starting 4 weeks and a shorter stride in footprint analysis performed at 12 weeks post-AAV injection. Immunohistochemistry showed thinning of the molecular layer and the formation of nuclear inclusions in Purkinje cells from mice injected with low doses of AAV-PHP.B encoding ATXN3(Q89). Moreover, ATXN3(Q89) expression significantly reduced the number of large projection neurons in the cerebellar nuclei to one third of that observed in mice expressing ATXN3(Q15). This AAV-based approach is superior to conventional methods in that the required number of model mice can be created simply by injecting AAV, and the expression levels of the responsible gene can be adjusted by changing the amount of AAV injected. Moreover, this method may be applied to produce SCA3 models in non-human primates.

Laurocapram, a transdermal enhancer, boosts cephalosporin’s antibacterial activity against Methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA), a notorious bacterium with high drug resistance and easy recurrence after surgery, has posed significant clinical treatment challenges. In the current scarcity of new antibiotics, the identification of adjuvants to existing antibiotics is a promising approach to combat infections caused by multidrug-resistant Gram-positive bacteria. The in vitro synergy test, which included a MIC assay, time-kill curve, antimicrobial susceptibility testing, and live/dead bacteria staining assay, revealed that laurocapram, a widely used chemical transdermal enhancer, could potentiate the antibacterial activity of cephalosporins against MRSA. In vitro, laurocapram combined with cefixime showed an excellent synergistic activity against MRSA (FICI = 0.28 ± 0.00). In addition, the combination of laurocapram and cefixime may inhibited the formation of MRSA biofilm and caused cell membrane damage. Following that, we discovered that combining laurocapram with cefixime could alleviate the symptoms of mice in the MRSA skin infection model and the MRSA pneumonia model. In conclusion, laurocapram is a promising and low-cost antibacterial adjuvant, providing a new strategy for further exploring the use of lower doses of cephalosporins to combat MRSA infection.

Vagus nerve modulates acute-on-chronic liver failure progression via CXCL9

Abstract Background: Hepatic inflammatory cell accumulation and the subsequent systematic inflammation drive acute-on-chronic liver failure (ACLF) development. Previous studies showed that the vagus nerve exerts anti-inflammatory activity in many inflammatory diseases. Here, we aimed to identify the key molecule mediating the inflammatory process in ACLF and reveal the neuroimmune communication arising from the vagus nerve and immunological disorders of ACLF. Methods: Proteomic analysis was performed and validated in ACLF model mice or patients, and intervention animal experiments were conducted using neutralizing antibodies. PNU-282987 (acetylcholine receptor agonist) and vagotomy were applied for perturbing vagus nerve activity. Single-cell RNA sequencing (scRNA-seq), flow cytometry, immunohistochemical and immunofluorescence staining, and CRISPR/Cas9 technology were used for in vivo or in vitro mechanistic studies. Results: The unbiased proteomics identified C-X-C motif chemokine ligand 9 (CXCL9) as the greatest differential protein in the livers of mice with ACLF and its relation to the systematic inflammation and mortality were confirmed in patients with ACLF. Interventions on CXCL9 and its receptor C-X-C chemokine receptor 3 (CXCR3) improved liver injury and decreased mortality of ACLF mice, which were related to the suppressing of hepatic immune cells’ accumulation and activation. Vagus nerve stimulation attenuated while vagotomy aggravated the expression of CXCL9 and the severity of ACLF. Blocking CXCL9 and CXCR3 ameliorated liver inflammation and increased ACLF-associated mortality in ACLF mice with vagotomy. scRNA-seq revealed that hepatic macrophages served as the major source of CXCL9 in ACLF and were validated by immunofluorescence staining and flow cytometry analysis. Notably, the expression of CXCL9 in macrophages was modulated by vagus nerve-mediated cholinergic signaling. Conclusions: Our novel findings highlighted that the neuroimmune communication of the vagus nerve–macrophage–CXCL9 axis contributed to ACLF development. These results provided evidence for neuromodulation as a promising approach for preventing and treating ACLF.

Distribution of the active components from Xianglian Pill in tissues of healthy and antibiotic-associated diarrhea model mice and the mechanism study

Antibiotic-associated diarrhea (AAD) is a common side effect of antibiotic therapy, characterized by intestinal inflammation which reduces the quality of life of patients. Xianglian Pill (XLP) has long been used to treat abdominal pain, diarrhea, bacillary dysentery and enteritis. Studies found that XLP has curative effect on AAD; however, the chemical constituents and mechanism of XLP have not been fully elucidated because of the lack of in vitro and in vivo studies. In this study, ultra-high performance liquid chromatography mass spectrometry method (UPLC-Q-Exactive-Orbitrap-HRMS) was used to examine the components of the XLP. Then, the binding between active compounds and the key targets was studied using network pharmacology and molecular docking. A comparative tissue distribution study was established for the simultaneous determination of the 10 active components in healthy and AAD mouse models. Forty-six components were characterized from XLP. According to the network pharmacology degree value, a prediction was made that encompassed 42 components and 14 core targets, which were intricately involved in crucial biological pathways, such as the AGE-RAGE signaling, cellular senescence, and MAPK signaling. Tissue distribution analysis showed that the 10 components were widely distributed in the heart, liver, spleen, lungs, kidneys, small intestine, and large intestine of mice, with varying concentrations in healthy and AAD mice. Molecular docking analysis also indicated that the active compounds in the tissue distribution could bind tightly to key targets of network pharmacological studies. This study provides a reference for further investigations of the relationships between the chemical components and pharmacological activities of XLP.

Bifidobacterium longum-Derived Extracellular Vesicles Prevent Hepatocellular Carcinoma by Modulating the TGF-β1/Smad Signaling in Mice.

The involvement of gut microbiota in carcinogenesis has gradually been highlighted in past decades. Bacteria could play its role by the secretion of extracellular vesicles (EVs); however, interrelationship between bacterial EVs and hepatocellular carcinoma (HCC) development has not been investigated much. Diethylnitrosamine (DEN) was utilized to produce HCC model in mice, of which fecal was collected for detecting Bifidobacterium longum (B.longum) with real-time polymerase chain reaction (PCR). EV isolated from B.longum (B.longum-EV) with ultracentrifugation were stained with PKH26 to investigate the cellular uptake of murine hepatocytes (AML12). After treatment with B.longum-EV, TGF-β1-induced AML12 cells were subjected to morphological observation, fibrosis- and apoptosis-related marker detection with western blot, apoptotic ratio and reactive oxygen species (ROS) level analysis with flow cytometry, and oxidative stress biomarker assessment with enzyme-linked immunosorbent assay (ELISA); meanwhile, animal studies including liver function, tumor formation rate, and histological analysis, were also performed to investigate the role of B.longum-EV in the fibrosis, apoptosis, oxidative stress, and carcinogenesis of the liver in vivo. The levels of B.longum were significantly reduced in HCC model mice. B.longum-EV could enter AML12 cells and effectively attenuate TGF-β1-induced fibrosis, apoptosis, and oxidative stress in AML12 cells. In vivo studies showed that B.longum-EV administration alleviated DEN-induced liver fibrosis, apoptosis, and oxidative stress at the early stage. Moreover, B.longum-EV administration also effectively reduced the tumor formation rate and liver function injury in DEN-induced mice and down-regulated TGF-β1 expression and Smad3 phosphorylation of mouse liver. B.longum-EVs protect hepatocytes against fibrosis, apoptosis, and oxidative damage, which exert a potential of preventing HCC development.

Ecklonia cava Polyphenols Have a Preventive Effect on Parkinson's Disease through the Activation of the Nrf2-ARE Pathway.

Parkinson's disease (PD) is a degenerative neurological disorder defined by the deterioration and loss of dopamine-producing neurons in the substantia nigra, leading to a range of motor impairments and non-motor symptoms. The underlying mechanism of this neurodegeneration remains unclear. This research examined the neuroprotective properties of Ecklonia cava polyphenols (ECPs) in mitigating neuronal damage induced by rotenone via the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway. Using human neuroblastoma SH-SY5Y cells and PD model mice, we found that ECP, rich in the antioxidant polyphenol phlorotannin, boosted the gene expression and functionality of the antioxidant enzyme NAD(P)H quinone oxidoreductase-1. ECP also promoted Nrf2 nuclear translocation and increased p62 expression, suggesting that p62 helps sustain Nrf2 activation via a positive feedback loop. The neuroprotective effect of ECP was significantly reduced by Compound C (CC), an AMP-activated protein kinase (AMPK) inhibitor, which also suppressed Nrf2 nuclear translocation. In PD model mice, ECPs improved motor functions impaired by rotenone, as assessed by the pole test and wire-hanging test, and restored intestinal motor function and colon tissue morphology. Additionally, ECPs increased tyrosine hydroxylase expression in the substantia nigra, indicating a protective effect on dopaminergic neurons. These findings suggest that ECP has a preventative effect on PD.

CircTOP1 targeted regulation of PTBP1 expression promotes the progression of coronary artery calcification

Coronary artery calcification (CAC) is a hallmark event in the pathogenesis of cardiovascular disease, involving the phenotypic transformation of vascular smooth muscle cells (VSMC) towards an osteogenic state. Despite this understanding, the molecular mechanisms governing the VSMC osteogenic switch remain incompletely elucidated. Here, we sought to examine the potential role of circular RNA (circRNA) in the context of CAC. Through transcriptome analysis of circRNA-seq, we identified circTOP1 as a potential candidate circRNA in individuals with CAC. Furthermore, we observed that overexpression of circTOP1 exacerbated vascular calcification in a CAC model. Subsequent pull-down assays revealed an interaction between circTOP1 and PTBP1, a putative target gene of circTOP1 in the context of CAC. In both in vivo and in vitro experiments, we observed heightened expression of circTOP1 and PTBP1 in the CAC model, and noted that reducing circTOP1 expression effectively reduced calcium salt deposits and mineralized nodules in model mice. Additionally, in vitro experiments demonstrated that overexpression of PTBP1 reversed the weakening of signaling caused by silencing circTOP1, thereby exacerbating the osteogenic transition and calcification of VSMC. Collectively, our findings suggested that circTOP1 promotes CAC by modulating PTBP1 expression to mediate VSMC transdifferentiation.

Chronic Administration of Cannabinoid Agonists ACEA, AM1241, and CP55,940 Induce Sex-Specific Differences in Tolerance and Sex Hormone Changes in a Chemotherapy-Induced Peripheral Neuropathy.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of chemotherapy treatment, routinely manifesting as increased pain sensitivity (allodynia) in distal extremities. Despite its prevalence, effective treatment options are limited. Cannabinoids are increasingly being evaluated for their ability to treat chronic pain conditions, including CIPN. While previous studies have revealed sex differences in cannabinoid-mediated antinociception in acute and chronic pain models, there is a paucity of studies addressing potential sex differences in the response of CIPN to cannabinoid treatment. Therefore, we evaluated the long-term antiallodynic efficacy of cannabinoid receptor type 1 (CB1)-selective, cannabinoid receptor type 2 (CB2)-selective, and CB1/CB2 mixed agonists in the cisplatin CIPN model, using both male and female mice. CB1 selective agonism was observed to have sex differences in the development of tolerance to antiallodynic effects, with females developing tolerance more rapidly than males, while the antiallodynic effects of selective CB2 agonism lacked tolerance development. Compound-specific changes to the female estrous cycle and female plasma estradiol levels were noted, with CB1 selective agonism decreasing plasma estradiol while CB2 selective agonism increased plasma estradiol. Chronic administration of a mixed CB1/CB2 agonist resulted in increased mRNA expression of proinflammatory cytokines and endocannabinoid regulatory enzymes in female spinal cord tissue. Ovarian tissue was noted to have proinflammatory cytokine mRNA expression following administration of a CB2 acting compound while selective CB1 agonism resulted in decreased proinflammatory cytokines and endocannabinoid regulatory enzymes in testes. These results support the need for further investigation into the role of sex and sex hormones signaling in pain and cannabinoid-mediated antinociceptive effects. SIGNIFICANCE STATEMENT: CIPN is a common side effect of chemotherapy. We have found that both CB1 and CB2 receptor agonism produce antinociceptive effects in a cisplatin CIPN model. We observed that tolerance to CB1-mediated antinociception developed faster in females and did not develop for CB2-mediated antinociception. Additionally, we found contrasting roles for CB1/CB2 receptors in the regulation of plasma estradiol in females, with CB1 agonism attenuating estradiol and CB2 agonism enhancing estradiol. These findings support the exploration of cannabinoid agonists for CIPN.

Suppression of eEF2 phosphorylation alleviates synaptic failure and cognitive deficits in mouse models of Down syndrome.

Cognitive impairment is a core feature of Down syndrome (DS), and the underlying neurobiological mechanisms remain unclear. Translation dysregulation is linked to multiple neurological disorders characterized by cognitive impairments. Phosphorylation of the translational factor eukaryotic elongation factor 2 (eEF2) by its kinase eEF2K results in inhibition of general protein synthesis. We used genetic and pharmacological methods to suppress eEF2K in two lines of DS mouse models. We further applied multiple approaches to evaluate the effects of eEF2K inhibition on DS pathophysiology. We found that eEF2K signaling was overactive in the brain of patients with DS and DS mouse models. Inhibition of eEF2 phosphorylation through suppression of eEF2K in DS model mice improved multiple aspects of DS-associated pathophysiology including de novo protein synthesis deficiency, synaptic morphological defects, long-term synaptic plasticity failure, and cognitive impairments. Our data suggested that eEF2K signaling dysregulation mediates DS-associated synaptic and cognitive impairments. Phosphorylation of the translational factor eukaryotic elongation factor 2 (eEF2) is increased in the Down syndrome (DS) brain. Suppression of the eEF2 kinase (eEF2K) alleviates cognitive deficits in DS models. Suppression of eEF2K improves synaptic dysregulation in DS models. Cognitive and synaptic impairments in DS models are rescued by eEF2K inhibitors.