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1.
Braz. j. biol ; 84: e250556, 2024. ilus
Article in English | LILACS, VETINDEX | ID: biblio-1360208

ABSTRACT

Exosomes are 30-120nm bio particles transferred from donor to recipient cells leading to modification in their regulatory mechanisms depending upon the coded message in the form of loaded biomolecule. Cancer cells derived exosomes the true representatives of the parent cells have been found to modify the tumor surrounding/distinct regions and participate in metastasis, angiogenesis and immune suppression. Tis study was aimed to study the effects of tumor mice derived exosomes on the normal mice spleen isolated T cells by using co-culture experiments and flow cytometer analysis. We mainly focused on some of the T cells population and cytokines including IFN-γ, FOXP3+ regulatory T (Treg) cells and KI67 (proliferation marker). Overall results indicated random changes in different set of experiments, where the cancer derived exosomes reduced the IFN-γ expression in both CD4 and CD8 T cells, similarly the Treg cells were also found decreased in the presence of cancer exosomes. No significant changes were observed on the Ki67 marker expression. Such studies are helpful in understanding the role of cancer exosomes in immune cells suppression in tumor microenvironment. Cancer exosomes will need to be validated in vivo and in vitro on a molecular scale in detail for clinical applications.


Os exossomos são biopartículas de 30-120 nm transferidas de células doadoras para células receptoras, levando à modificação em seus mecanismos reguladores, dependendo da mensagem codificada na forma de biomolécula carregada. Verificou-se que exossomos derivados de células cancerosas ­ os verdadeiros representantes das células-mãe ­ modificam as regiões circundantes / distintas do tumor e participam da metástase, angiogênese e imunossupressão. Este estudo teve como objetivo estudar os efeitos de exossomos derivados de camundongos com tumor nas células T isoladas de baço de camundongos normais, usando experimentos de cocultura e análise de citômetro de fluxo. Concentrou-se, principalmente, em algumas populações de células T e citocinas, incluindo IFN-γ, células T reguladoras FOXP3 + (Treg) e KI67 (marcador de proliferação). Os resultados gerais indicaram mudanças aleatórias em diferentes conjuntos de experimentos, em que os exossomos derivados de câncer reduziram a expressão de IFN-γ em células T CD4 e CD8, da mesma forma que as células Treg também foram encontradas diminuídas na presença de exossomos de câncer. Nenhuma mudança significativa foi observada na expressão do marcador Ki67. Esses dados são úteis para a compreensão do papel dos exossomos do câncer na supressão de células do sistema imunológico no microambiente tumoral. Exossomos de câncer precisarão ser validados in vivo e in vitro em escala molecular com detalhes para aplicações clínicas.


Subject(s)
Animals , Mice , Exosomes , Tumor Microenvironment , Immune System , Neoplasm Metastasis , Neoplasms
2.
Rev. Ciênc. Méd. Biol. (Impr.) ; 21(1): 40-45, maio 05,2022. fig
Article in Portuguese | LILACS | ID: biblio-1370563

ABSTRACT

Introduction: dengue is a most common mosquito-borne viral disease in the Americas and tropical countries. Objective: in this work, mice were hyperimmunized with DENV 4 antigen to produce monoclonal antibodies (mAbs). Methodology: DENV 4 (GenBank KC806069) was inoculated in C6/36 cell monolayers cultivated in Leibovitz's 15 medium supplemented with 5% fetal bovine serum and incubated at 28 oC. The virus stock was submitted to concentration and ultracentrifugation and stored at -80 oC until use (VC DENV 4). Balb/c mice were injected intraperitoneally with 50µg of DENV-4 and successive intraperitoneal injections of 25 µg of VCDENV 4 with Freund's incomplete adjuvant were performed. The spleen cells were fused to SP2/0 myeloma cells with PEG 1540 and distributed in 96-well microplates with Iscove's modified medium with Hipoxantina­Aminopterina­Timidina. Hybridoma screening by indirect ELISA showed positive results for six mAbs, and their characterization was performed by Western blotting and Indirect Immunofluorescence (IFI) techniques. Results: the six mAbs showed strong recognition of prM (24/29 kDa), and minor reaction to E protein (66 kDa), E/E protein dimer (105 kDa), and NS1 (49 kDa) protein in two mAbs. The use of mAbs anti-prM as a diagnostic tool using IFI has been demonstrated to detect DENV-4 antigen in infected cells or tissues. Conclusion: DENV 4 generate mAbs with strong reactivity to prM with potential use to confirm the presence of DENV 4 antigen in tissues or infected cells.


Introdução: a dengue é uma doença viral transmitida por mosquitos comumente das Américas e países tropicais. Objetivo: neste trabalho, camundongos foram hiperimunizados com antígeno DENV 4 para produzir anticorpos monoclonais (mAbs). Metodologia: DENV 4 (GenBank KC806069) foi inoculado em monocamadas de células C6 / 36 cultivadas em meio Leibovitz 15 suplementado com 5% de soro fetal bovino e incubadas a 28oC. O estoque viral foi submetido à concentração, ultracentrifugação e armazenado a -80 oC (VC DENV 4). Camundongos Balb / c foram injetados intraperitonealmente com 50 µg de VC DENV-4 e injeções intraperitoneais sucessivas de 25 µg de antigeno com adjuvante incompleto de Freund. As células do baço foram misturadas a células SP2/0 com PEG 1540 e distribuídas em microplacas de 96 poços com meio Iscove Modificado em presença de Hipoxantina ­ Aminopterina ­ Timidina. A triagem de hibridomas por ELISA indireto apresentou resultados positivos para seis mAbs, e sua caracterização foi realizada por técnicas de Western blotting e Imunofluorescência Indireta (IFI). Resultados: os seis mAbs mostraram forte reconhecimento de prM (24/29 kDa) e reação menor à proteína E (66 kDa), dímero de proteína E / E (105 kDa) e proteína NS1 (49 kDa) em dois mAbs. O uso de mAbs anti-prM como uma ferramenta de diagnóstico utilizando IFI demonstrou eficacia em detectar o antígeno DENV-4 em células ou tecidos infectados. Conclusão: o mAbs produzidos para DENV 4 demonstraram uma forte reatividade contra prM, e poderiam ser uma ferramenta de uso potencial no diagnóstico de DENV 4 .


Subject(s)
Animals , Rats , Dengue , Mice , Antibodies, Monoclonal
3.
Arq. ciências saúde UNIPAR ; 26(1): 75-87, Jan-Abr. 2022.
Article in Portuguese | LILACS | ID: biblio-1362684

ABSTRACT

O cigarro eletrônico surgiu como uma tentativa para minimizar a dependência ao uso de tabaco, entretanto, engloba controvérsias e dúvidas acerca das reais implicações para o organismo humano. Diante disso, o presente estudo tem como objetivo realizar uma revisão da literatura a fim de relacionar o uso de cigarro eletrônico com suas consequências para os humanos. Os estudos analisados relatam experimentos in vitro e in vivo em camundongos, demonstrando menor concentração de poluentes e nocividades no cigarro eletrônico comparado ao convencional, porém, seu potencial efeito maléfico está relacionado à composição do e-líquido, à maneira do uso e à variedade de aromas presentes nos produtos. Além disso, foram verificadas lesões celulares, hiperreatividade das vias aéreas, liberação de citocinas ­ IL-8, IL-10 e TNF, redução da ação antimicrobiana de queratinócitos e potencial apoptose nas células alveolares. Foi observado também um aumento em até cinco vezes da concentração de carboxihemoglobina em comparação ao cigarro comum e um aumento na auto renovação de células de adenocarcinoma pulmonar de células não pequenas, devido à expressão de SOX2. Observa-se também que em casos de DPOC, o cigarro eletrônico não apresenta agravamentos na fisiologia respiratória, contrapondo outras ocorrências como asma, pneumonia, câncer de pulmão e doenças infecciosas que podem ser ocasionadas ou exacerbadas pelo seu uso. Contudo, pelo curto prazo de observação de seus efeitos, não é possível determinar com precisão a segurança dos cigarros eletrônicos, dessa forma, faz-se necessário que mais pesquisas longitudinais sejam desenvolvidas, auxiliando, assim, na construção de evidências sobre a segurança dos cigarros eletrônicos e na regulamentação futura do produto.


Electronic cigarettes emerged as an attempt to minimize tobacco dependence. However, its use is surrounded by controversies and doubts about the real implications for the human organism. Therefore, this study aims at performing a review of the most recent literature to corelate the use of e-cigarettes with their consequences for the human body. The analyzed studies relate in vitro and in vivo experiments on mice, demonstrating lower concentration of pollutants and harmfulness in the electronic cigarette than in conventional cigarettes. However, its potential harmful effect is related to the composition of the e-liquid, in its use and in the variety of aromas in the products. In addition, cellular lesions, airway hyperreactivity, release of IL-8, IL-10 and TNF cytokines could be observed, as well as reduced keratinocyte antimicrobial action and potential apoptosis in alveolar cells. An increase of up to five-fold the concentration of carboxyhemoglobin in comparison to ordinary cigarettes and an increase in self-renewal of non-small pulmonary adenocarcinoma cells due to the expression of SOX2 have also been related. It could also be observed that in COPD cases, e-cigarettes do not present worsening in respiratory physiology, which contrasts with other occurrences such as asthma, pneumonia, lung cancer, and infectious diseases that can be caused or exacerbated by its use. However, due to the short term of observation of the effects, the safety of e-cigarettes could not be accurately determined, thus, the need for further longitudinal research is necessary, which could be used to help build evidence about the safety of e-cigarettes and also to create future regulation of the product.


Subject(s)
Animals , Mice , Rats , Electronic Nicotine Delivery Systems/instrumentation , Lung Diseases , Pneumonia/complications , Asthma/complications , Tobacco Use Disorder/complications , Smoking , Disease , Lung Injury , Tobacco Use , Vaping , Smokers , E-Cigarette Vapor/adverse effects , Lung Neoplasms
4.
Neuroscience Bulletin ; (6): 113-134, 2022.
Article in English | WPRIM | ID: wpr-922681

ABSTRACT

Mutations of the X-linked methyl-CpG-binding protein 2 (MECP2) gene in humans are responsible for most cases of Rett syndrome (RTT), an X-linked progressive neurological disorder. While genome-wide screens in clinical trials have revealed several putative RTT-associated mutations in MECP2, their causal relevance regarding the functional regulation of MeCP2 at the etiologic sites at the protein level requires more evidence. In this study, we demonstrated that MeCP2 was dynamically modified by O-linked-β-N-acetylglucosamine (O-GlcNAc) at threonine 203 (T203), an etiologic site in RTT patients. Disruption of the O-GlcNAcylation of MeCP2 specifically at T203 impaired dendrite development and spine maturation in cultured hippocampal neurons, and disrupted neuronal migration, dendritic spine morphogenesis, and caused dysfunction of synaptic transmission in the developing and juvenile mouse cerebral cortex. Mechanistically, genetic disruption of O-GlcNAcylation at T203 on MeCP2 decreased the neuronal activity-induced induction of Bdnf transcription. Our study highlights the critical role of MeCP2 T203 O-GlcNAcylation in neural development and synaptic transmission potentially via brain-derived neurotrophic factor.


Subject(s)
Animals , Humans , Methyl-CpG-Binding Protein 2/metabolism , Mice , Neurodevelopmental Disorders/genetics , Rett Syndrome/genetics , Synaptic Transmission , Threonine
5.
Neuroscience Bulletin ; (6): 181-199, 2022.
Article in English | WPRIM | ID: wpr-922672

ABSTRACT

The glymphatic system plays a pivotal role in maintaining cerebral homeostasis. Chronic cerebral hypoperfusion, arising from small vessel disease or carotid stenosis, results in cerebrometabolic disturbances ultimately manifesting in white matter injury and cognitive dysfunction. However, whether the glymphatic system serves as a potential therapeutic target for white matter injury and cognitive decline during hypoperfusion remains unknown. Here, we established a mouse model of chronic cerebral hypoperfusion via bilateral common carotid artery stenosis. We found that the hypoperfusion model was associated with significant white matter injury and initial cognitive impairment in conjunction with impaired glymphatic system function. The glymphatic dysfunction was associated with altered cerebral perfusion and loss of aquaporin 4 polarization. Treatment of digoxin rescued changes in glymphatic transport, white matter structure, and cognitive function. Suppression of glymphatic functions by treatment with the AQP4 inhibitor TGN-020 abolished this protective effect of digoxin from hypoperfusion injury. Our research yields new insight into the relationship between hemodynamics, glymphatic transport, white matter injury, and cognitive changes after chronic cerebral hypoperfusion.


Subject(s)
Animals , Brain Ischemia , Carotid Stenosis/drug therapy , Cognitive Dysfunction/etiology , Digoxin , Disease Models, Animal , Mice , Mice, Inbred C57BL , White Matter
6.
Neuroscience Bulletin ; (6): 1-15, 2022.
Article in English | WPRIM | ID: wpr-922671

ABSTRACT

Parkinson's disease (PD) is the second most common and fastest-growing neurodegenerative disorder. In recent years, it has been recognized that neurotransmitters other than dopamine and neuronal systems outside the basal ganglia are also related to PD pathogenesis. However, little is known about whether and how the caudal zona incerta (ZIc) regulates parkinsonian motor symptoms. Here, we showed that specific glutamatergic but not GABAergic ZIc


Subject(s)
Animals , Mice , Neurons , Parkinson Disease , Parkinsonian Disorders , Substantia Nigra , Zona Incerta
7.
Neuroscience Bulletin ; (6): 29-46, 2022.
Article in English | WPRIM | ID: wpr-922666

ABSTRACT

A large number of putative risk genes for autism spectrum disorder (ASD) have been reported. The functions of most of these susceptibility genes in developing brains remain unknown, and causal relationships between their variation and autism traits have not been established. The aim of this study was to predict putative risk genes at the whole-genome level based on the analysis of gene co-expression with a group of high-confidence ASD risk genes (hcASDs). The results showed that three gene features - gene size, mRNA abundance, and guanine-cytosine content - affect the genome-wide co-expression profiles of hcASDs. To circumvent the interference of these features in gene co-expression analysis, we developed a method to determine whether a gene is significantly co-expressed with hcASDs by statistically comparing the co-expression profile of this gene with hcASDs to that of this gene with permuted gene sets of feature-matched genes. This method is referred to as "matched-gene co-expression analysis" (MGCA). With MGCA, we demonstrated the convergence in developmental expression profiles of hcASDs and improved the efficacy of risk gene prediction. The results of analysis of two recently-reported ASD candidate genes, CDH11 and CDH9, suggested the involvement of CDH11, but not CDH9, in ASD. Consistent with this prediction, behavioral studies showed that Cdh11-null mice, but not Cdh9-null mice, have multiple autism-like behavioral alterations. This study highlights the power of MGCA in revealing ASD-associated genes and the potential role of CDH11 in ASD.


Subject(s)
Animals , Autism Spectrum Disorder/genetics , Brain , Cadherins/genetics , Gene Expression , Mice , Mice, Knockout
8.
Neuroscience Bulletin ; (6): 16-28, 2022.
Article in English | WPRIM | ID: wpr-922663

ABSTRACT

Chronic stress leads to many psychiatric disorders, including social and anxiety disorders that are associated with over-activation of neurons in the basolateral amygdala (BLA). However, not all individuals develop psychiatric diseases, many showing considerable resilience against stress exposure. Whether BLA neuronal activity is involved in regulating an individual's vulnerability to stress remains elusive. In this study, using a mouse model of chronic social defeat stress (CSDS), we divided the mice into susceptible and resilient subgroups based on their social interaction behavior. Using in vivo fiber photometry and in vitro patch-clamp recording, we showed that CSDS persistently (after 20 days of recovery from stress) increased BLA neuronal activity in all the mice regardless of their susceptible or resilient nature, although impaired social interaction behavior was only observed in susceptible mice. Increased anxiety-like behavior, on the other hand, was evident in both groups. Notably, the CSDS-induced increase of BLA neuronal activity correlated well with the heightened anxiety-like but not the social avoidance behavior in mice. These findings provide new insight to our understanding of the role of neuronal activity in the amygdala in mediating stress-related psychiatric disorders.


Subject(s)
Amygdala , Animals , Anxiety/etiology , Anxiety Disorders , Avoidance Learning , Mice , Mice, Inbred C57BL , Social Behavior , Stress, Psychological/complications
9.
Article in English | WPRIM | ID: wpr-922575

ABSTRACT

OBJECTIVE@#To investigate the protective effect of Chinese herbal formula Huangqin Decoction (HQD) on ulcerative colitis mouse model induced by dextran sulphate sodium (DSS) and human intestinal epithelial cell injury induced by tumour necrosis factor-α (TNF-α).@*METHODS@#In vivo, 30 male C57BL/6 mice were divided into 5 groups using a random number table (n=6 per group), including control, DSS, 5-aminosalicylic acid (5-ASA), HQD low- (HQD-L) and high-dose (HQD-H) groups. The colitis mouse model was established by 3% (w/v) DSS water for 5 days. Meanwhile, mice in the HQD-L, HQD-H and 5-ASA groups were administrated with 100, 200 mg/kg HQD or 100 mg/kg 5-ASA, respectively, once daily by gavage. After 9 days of administration, the body weight, disease activity index (DAI) score and colon length of mice were measured, the pathological changes of colons were analyzed by hematoxylin-eosin staining (HE) staining, and the levels of serum interleukin (IL)-6, IL-1β and TNF-α were measured by enzyme linked immunosorbent assay. In vitro, the human colon epithelial normal cells (FHC cells) were exposed to HQD (0.6 mg/mL) for 12 h and then treated with TNF-α (10 ng/mL) for 24 h. The tight junction (TJ) protein expression levels of Claudin-4 and Occludin, and the protein phosphorylation levels of p65 and inhibitor of nuclear factor kappaB (NF-κB)-α (IκBα) were measured by Western blot.@*RESULTS@#In vivo, compared with the DSS group, HQD-H treatment attenuated the weight loss and reduced DAI score of mice on the 8th day (P<0.05). Moreover, HQD-H treatment ameliorated the colon shortening in the DSS-induced colitis mice (P<0.05). HE staining showed HQD attenuated the pathological changes of colitis mice, and the histological scores of HQD-H and 5-ASA groups were significantly decreased compared with the DSS group (P<0.05). Meanwhile, HQD-H and 5-ASA significantly decreased the serum IL-1β, IL-6 and TNF-α levels of mice (P<0.05). In vitro experiments showed that HQD up-regulated Occludin and Claudin-4 protein expressions and inhibited p-p65 and p-IκBα levels in FHC cells compared with the TNF-α group (P<0.05).@*CONCLUSION@#HQD significantly relieved the symptoms in DSS-induced colitis mice by inhibiting pro-inflammatory cytokines expression and maintained the homeostasis of TJ protein in FHC cells by suppressing TNF-α-induced NF-κB activation.


Subject(s)
Animals , Colitis, Ulcerative/drug therapy , Dextran Sulfate , Disease Models, Animal , Male , Mice , Mice, Inbred C57BL , NF-kappa B , Scutellaria baicalensis , Tumor Necrosis Factor-alpha
10.
Article in English | WPRIM | ID: wpr-922534

ABSTRACT

OBJECTIVE@#The chemo-preventative and therapeutic properties of selenium nanoparticles (SeNPs) have been documented over recent decades and suggest the potential uses of SeNPs in medicine. Biogenic SeNPs have higher biocompatibility and stability than chemically synthesized nanoparticles, which enhances their medical applications, especially in the field of cancer therapy. This study evaluated the potential of green-synthetized SeNPs by using berberine (Ber) as an antitumor agent and elucidated the mechanism by which these molecules combat Ehrlich solid tumors (ESTs).@*METHODS@#SeNPs containing Ber (SeNPs-Ber) were synthesized using Ber and Na@*RESULTS@#Treatment with SeNPs-Ber significantly improved the survival rate and decreased the body weight and tumor size, compared to the EST group. SeNPs-Ber reduced oxidative stress in tumor tissue, as indicated by a decrease in the lipid peroxidation and nitric oxide levels and an increase in the glutathione levels. Moreover, SeNPs-Ber activated an apoptotic cascade in the tumor cells by downregulating the B-cell lymphoma 2 (Bcl-2) expression rate and upregulating the Bcl-2-associated X protein and caspase-3 expression rates. SeNPs-Ber also considerably improved the histopathological alterations in the developed tumor tissue, compared to the EST group.@*CONCLUSION@#Our study provides a new insight into the potential role of green-synthesized SeNPs by using Ber as a promising anticancer agent, these molecules could be used alone or as supplementary medication during chemotherapy.


Subject(s)
Animals , Antineoplastic Agents , Antioxidants , Berberine , Male , Mice , Nanoparticles , Selenium
11.
Article in English | WPRIM | ID: wpr-929267

ABSTRACT

Ma-Mu-Ran Antidiarrheal Capsules (MMRAC) is traditional Chinese medicine that has been used to treat diarrhea caused by acute enteritis (AE) and bacillary dysentery in Xinjiang (China) for many years. However, the potential therapeutic mechanism of MMRAC for AE and its regulatory mechanism on host metabolism is unclear. This study used fecal metabolomics profiling with GC/MS and 16S rRNA gene sequencing analysis to explore the potential regulatory mechanisms of MMRAC on a dextran sulfate sodium salt (DSS)-induced mouse model of AE. Fecal metabolomics-based analyses were performed to detect the differentially expressed metabolites and metabolic pathways. The 16S rRNA gene sequencing analysis was used to assess the altered gut microbes at the genus level and for functional prediction. Moreover, Pearson correlation analysis was used to integrate differentially expressed metabolites and altered bacterial genera. The results revealed that six intestinal bacteria and seven metabolites mediated metabolic disorders (i.e., metabolism of amino acid, carbohydrate, cofactors and vitamins, and lipid) in AE mice. Besides, ten altered microbes mediated the differential expression of eight metabolites and regulated these metabolisms after MMRAC administration. Overall, these findings demonstrate that AE is associated with metabolic disorders and microbial dysbiosis. Further, we present that MMRAC exerts protective effects against AE by improving host metabolism through the intestinal flora.


Subject(s)
Animals , Antidiarrheals/pharmacology , Capsules , Enteritis/genetics , Feces/microbiology , Genes, rRNA , Metabolomics , Mice , RNA, Ribosomal, 16S/genetics
12.
Article in English | WPRIM | ID: wpr-929264

ABSTRACT

Abelmoschus manihot (L.) Medik. (A. manihot) is a traditional Chinese herbal medicine with a variety of pharmacological properties. It was first recorded in Jiayou Materia Medica dating back to the Song dynasty to eliminate urinary tract irritation by clearing away heat and diuretic effect. However, its pharmacological action on urinary tract infections has not been investigated. The present study aims to evaluate the anti-inflammatory activity of A. manihot on a mouse model of lipopolysaccharide (LPS)-induced cystitis. The results showed that A. manihot decreased white blood cell (WBC) count in urine sediments of the cystitis mice, alleviated bladder congestion, edema, as well as histopathological damage, reduced the expression levels of tumor necrosis factor-α, interleukin-6, and interleukin-1β simultaneously. Moreover, A. manihot administration significantly downregulated the expression levels of TLR4, MYD88, IκBα, p-IκBα, NF-κB p65, and p-NF-κB p65 in LPS-induced cystitis mice. These findings demonstrated the protective effect of A. manihot against LPS-induced cystitis, which is attributed to its anti-inflammatory profile by suppressing TLR4/MYD88/NF-κB pathways. Our results suggest that A. manihot could be a potential candidate for cystitis treatment.


Subject(s)
Abelmoschus/metabolism , Animals , Anti-Inflammatory Agents/therapeutic use , Cystitis , Female , Humans , Lipopolysaccharides/pharmacology , Male , Mice , Myeloid Differentiation Factor 88/metabolism , NF-KappaB Inhibitor alpha/metabolism , NF-kappa B/metabolism , Signal Transduction , Toll-Like Receptor 4/metabolism
13.
Article in English | WPRIM | ID: wpr-929263

ABSTRACT

A series of 26 novel derivatives have been synthesized through structural modification of gentiopicroside, a lead COX-2 inhibitor. And their in vivo and in vitro anti-inflammatory activities have been investigated. The in vitro anti-inflammatory activities were evaluated against NO, PGE2, and IL-6 production in the mouse macrophage cell line RAW264.7 stimulated by LPS. Results showed that most compounds had good inhibitory activity. The in vivo inhibitory activities were further tested against xylene-induced mouse ear swelling. Results demonstrated that several compounds were more active than the parent compound gentiopicroside. The inhibition rate of the most active compound P23 (57.26%) was higher than positive control drug celecoxib (46.05%) at dose 0.28 mmol·kg-1. Molecular docking suggested that these compounds might bind to COX-2 and iNOS. Some of them, e.g P7, P14, P16, P21, P23, and P24, had high docking scores in accordance with their potency of the anti-inflammatory activitiy, that downregulation of the inflammatory factors, NO, PGE2, and IL-6, was possibly associated with the suppression of iNOS and COX-2. Therefore, these gentiopicroside derivatives may represent a novel class of COX-2 and iNOS inhibitors.


Subject(s)
Animals , Anti-Inflammatory Agents/pharmacology , Cyclooxygenase 2/chemistry , Dinoprostone , Interleukin-6/metabolism , Iridoid Glucosides , Mice , Molecular Docking Simulation , Pyridinolcarbamate
14.
Article in English | WPRIM | ID: wpr-929262

ABSTRACT

Chemical fractionation of the n-BuOH partition, which was generated from the EtOH extract of the flower buds of Tussilago farfara, afforded a series of polar constituents including four new sesquiterpenoids (1-4), one new sesquiterpenoid glucoside (5) and one known analogue (6) of the eudesmane type, as well as five known quinic acid derivatives (7-11). Structures of the new compounds were unambiguously characterized by detailed spectroscopic analyses, with their absolute configurations being established by X-ray crystallography, electronic circular dichroism (ECD) calculation and induced ECD experiments. The inhibitory effect of all the isolates against LPS-induced NO production in murine RAW264.7 macrophages was evaluated, with isochlorogenic acid A (7) showing significant inhibitory activity.


Subject(s)
Animals , Flowers/chemistry , Glucosides/pharmacology , Mice , Sesquiterpenes/pharmacology , Sesquiterpenes, Eudesmane/pharmacology , Tussilago/chemistry
15.
Article in English | WPRIM | ID: wpr-929260

ABSTRACT

Wuzi-Yanzong-Wan (WZYZW) is a classic prescription for male infertility. Our previous investigation has demonstrated that it can inhibit sperm apoptosis via affecting mitochondria, but the underlying mechanisms are unclear. The purpose of the present study was to explore the actions of WZYZW on mitochondrial permeability transition pore (mPTP) in mouse spermatocyte cell line (GC-2 cells) opened by atractyloside (ATR). At first, WZYZW-medicated serum was prepared from rats following oral administration of WZYZW for 7 days. GC-2 cells were divided into control group, model group, positive group, as well as 5%, 10%, 15% WZYZW-medicated serum group. Cyclosporine A (CsA) was used as a positive control. 50 μmol·L-1 ATR was added after drugs incubation. Cell viability was assessed using CCK-8. Apoptosis was detected using flow cytometry and TUNEL method. The opening of mPTP and mitochondrial membrane potential (MMP) were detected by Calcein AM and JC-1 fluorescent probe respectively. The mRNA and protein levels of voltage-dependent anion channel 1 (VDAC1), cyclophilin D (CypD), adenine nucleotide translocator (ANT), cytochrome C (Cyt C), caspase 3, 9 were detected by RT-PCR (real time quantity PCR) and Western blotting respectively. The results demonstrated that mPTP of GC-2 cells was opened after 24 hours of ATR treatment, resulting in decreased MMP and increased apoptosis. Pre-protection with WZYZ-medicated serum and CsA inhibited the opening of mPTP of GC-2 cells induced by ATR associated with increased MMP and decreased apoptosis. Moreover, the results of RT-qPCR and WB suggested that WZYZW-medicated serum could significantly reduce the mRNA and protein levels of VDAC1 and CypD, Caspase-3, 9 and CytC, as well as a increased ratio of Bcl/Bax. However, ANT was not significantly affected. Therefore, these findings indicated that WZYZW inhibited mitochondrial mediated apoptosis by attenuating the opening of mPTP in GC-2 cells. WZYZW-medicated serum inhibited the expressions of VDAC1 and CypD and increased the expression of Bcl-2, which affected the opening of mPTP and exerted protective and anti-apoptotic effects on GC-2 cell induced by ATR.


Subject(s)
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine , Animals , Atractyloside/pharmacology , Cyclophilin D , Male , Matrix Metalloproteinases , Mice , Mitochondrial Membrane Transport Proteins/metabolism , Mitochondrial Permeability Transition Pore , RNA, Messenger , Rats
16.
Article in English | WPRIM | ID: wpr-929259

ABSTRACT

Hepatic sinusoidal obstruction syndrome (HSOS) via exposure to pyrrolizidine alkaloids (PAs) is with high mortality and there is no effective treatment in clinics. Bear bile powder (BBP) is a famous traditional animal drug for curing a variety of hepatobiliary diseases such as cholestasis, inflammation, and fibrosis. Here, we aim to evaluate the protective effect of BBP against HSOS induced by senecionine, a highly hepatotoxic PA compound. Our results showed that BBP treatment protected mice from senecionine-induced HSOS dose-dependently, which was evident by improved liver histology including reduced infiltration of inflammatory cells and collagen positive cells, alleviated intrahepatic hemorrhage and hepatic sinusoidal endothelial cells, as well as decreased conventional serum liver function indicators. In addition, BBP treatment lowered matrix metalloproteinase 9 and pyrrole-protein adducts, two well-known markers positively associated with the severity of PA-induced HSOS. Further investigation showed that BBP treatment prevents the development of liver fibrosis by decreasing transforming growth factor beta and downstream fibrotic molecules. BBP treatment also alleviated senecionine-induced liver inflammation and lowered the pro-inflammatory cytokines, in which tauroursodeoxycholic acid played an important role. What's more, BBP treatment also decreased the accumulation of hydrophobic bile acids, such as cholic acid, taurocholic acid, glycocholic acid, as well. We concluded that BBP attenuates senecionine-induced HSOS in mice by repairing the bile acids homeostasis, preventing liver fibrosis, and alleviating liver inflammation. Our present study helps to pave the way to therapeutic approaches of the treatment of PA-induced liver injury in clinics.


Subject(s)
Animals , Bile , Bile Acids and Salts , Endothelial Cells/metabolism , Hepatic Veno-Occlusive Disease/pathology , Inflammation/pathology , Liver Cirrhosis/drug therapy , Mice , Powders , Pyrrolizidine Alkaloids/adverse effects , Ursidae
17.
Article in English | WPRIM | ID: wpr-929258

ABSTRACT

Colorectal cancer (CRC) is the third most lethal cancer and leading cause of cancer mortality worldwide. A key driver of CRC development is colon inflammatory responses especially in patients with inflammatory bowl disease (IBD). It has been proved that Panax notoginseng saponins (PNS) have anti-inflammatory, anti-oxidant and anti-tumor effects. The chemopreventive and immunomodulatory functions of PNS on colitis-associated colorectal cancer (CAC) have not been evaluated.This present study was designed to study the potential protective effects of PNS on AOM/DSS-induced CAC mice to explore the possible mechanism of PNS against CAC. Our study showed that PNS significantly alleviated colitis severity and prevented the occurrence of CAC. Functional assays revealed that PNS relieved immunosuppression of Treg cells in the CAC microenvironment by inhibiting the expression of IDO1 mediated directly by signal transducer and activator of transcription 1 (STAT1) rather than phosphorylated STAT1. Ultimately, Rh1, one of the PNS metabolites, exhibited the best inhibitory effect on IDO1 enzyme activity. Our study showed that PNS exerted significant chemopreventive function and immunomodulatory properties on CAC. It could reduce macrophages accumulation and Treg cells differentiation to reshape the immune microenvironment of CAC. These findings provided a promising approach for CAC intervention.


Subject(s)
Animals , Colitis/drug therapy , Colitis-Associated Neoplasms/drug therapy , Humans , Macrophages , Mice , Panax notoginseng , Saponins/therapeutic use , Tumor Microenvironment
18.
Article in English | WPRIM | ID: wpr-929248

ABSTRACT

Simiao Wan (SMW) is a traditional Chinese formula, including Atractylodis Rhizoma, Achyranthis Bidentatae Radix, Phellodendri Chinensis Cortex and Coicis Semen at the ratio of 1:1:2:2. It can be used to the treatment of diabetes. However, its bioactive compounds and underlying mechanism are unclear. This study aimed to screen the antilipolytic fraction from SMW and investigate its therapeutic mechanisms on hepatic insulin resistance. Different fractions of SMW were prepared by membrane separation combined with macroporous resin and their antilipolytic activities were screened in fasted mice. The effects of 60% ethanol elution (ESMW) on lipolysis were investigated in 3T3-L1 adipocytes stimulated by palmitic acid (PA) and high fat diet (HFD)-fed mice. In our study, ESMW is the bioactive fraction responsible for the antilipolytic activity of SMW and 13 compounds were characterized from ESMW by UHPLC-QTOF-MS/MS. ESMW suppressed protein kinase A (PKA)-hormone-sensitive lipase (HSL) related lipolysis and increased AMP-activated protein kinase (AMPK) phosphorylation in PA challenged 3T3-L1 adipocytes. AMPKα knockdown abolished the inhibitory effects of ESMW on IL-6 and HSL pSer-660, revealing that the antilipolytic and anti-inflammatory activities of ESMW are AMPK dependent. Furthermore, ESMW ameliorated insulin resistance and suppressed lipolysis in HFD-fed mice. It inhibited diacylglycerol accumulation in the liver and inhibited hepatic gluconeogenesis. Conditional medium collected from ESMW-treated 3T3-L1 cells ameliorated insulin action on hepatic gluconeogenesis in liver cells, demonstrating the antilipolytic activity contributed to ESMW beneficial effects on hepatic glucose production. In conclusion, ESMW, as the antilipolytic fraction of SMW, inhibited PKA-HSL related lipolysis by activating AMPK, thus inhibiting diacylglycerol (DAG) accumulation in the liver and thereby improving insulin resistance and hepatic gluconeogenesis.


Subject(s)
AMP-Activated Protein Kinases/metabolism , Animals , Insulin/metabolism , Lipolysis/physiology , Liver/metabolism , Mice , Tandem Mass Spectrometry
19.
Article in English | WPRIM | ID: wpr-929246

ABSTRACT

Three new ursane-type triterpenoids, 3-oxours-12-en-20, 28-olide (1), 3β-hydroxyurs-12-en-20, 28-olide (2) and 3β-hydroxyurs-11, 13(18)-dien-20, 28-olide (3), were isolated from a potent anti-inflammatory and antibacterial fraction of the ethanolic extract of Rosmarinus officinalis. Their structures were elucidated by a combination of extensive 1D- and 2D-NMR experiments, MS data and comparisons with literature reports. Compounds 1-3 exhibited significantly inhibitory effects on nitric oxide production in lipopolysaccharide-activated mouse RAW264.7 macrophages, but no antibacterial activity was found at a concentration of 128 μg·mL-1.


Subject(s)
Animals , Drugs, Chinese Herbal/chemistry , Mice , Molecular Structure , Rosmarinus , Triterpenes/chemistry
20.
Article in English | WPRIM | ID: wpr-929241

ABSTRACT

Parkinson's disease (PD) is a multifactorial disorder of the nervous system where a progressive loss of dopaminergic neurons exist. However, the pathogenesis of PD remains undefined, which becomes the main limitation for the development of clinical PD treatment. Demethylenetetrahydroberberine (DMTHB) is a novel derivative of natural product berberine. This study was aimed to explore the neuroprotective effects and pharmacological mechanism of DMTHB on Parkinson's disease using C57BL/6 mice. A PD model of mice was induced by administration of MPTP (20 mg·kg-1) and probenecid (200 mg·kg-1) twice per week for five weeks. The mice were administered with DMTHB daily by gavage at the dose of 5 and 50 mg·kg-1 for one- week prophylactic treatment and five-week theraputic treatment. The therapeutic effects of DMTHB were evaluated by behavior tests (the open field, rotarod and pole tests), immunohistochemical staining of tyrosine hydroxylase (TH), Nissl staining and biochemical assays. The molecular mechanisms of DMTHB on the key biomarkers of PD pathological states were analyzed by Western blot (WB) and qRT-PCR. DMTHB treatment alleviated the behavioral disorder induced by MPTP-probenecid. Nissl staining and TH staining showed that the damage of dopaminergic neurons in the substantia nigra was remarkably suppressed by DMTHB treatment. Western blot results showed that the ratio of Bcl-2/Bax and TH increased, but the level of α-synuclein (α-syn) was remarkably reduced, which indicated that the apoptosis of dopaminergic neurons in mice was significantly reduced. The protein phosphorylation of p-PI3K, p-AKT and p-mTOR also increased about 2-fold, compared with the model group. Furthermore, qRT-PCR results demonstrated that the mRNA levels of pro-inflammatory cytokines, IL-1β and TNF-α, were reduced, but the level of anti-inflammatory cytokine IL-10 increased after DMTHB treatment. Finally, the cellular assay displayed that DMTHB was also a strong antioxidant to protect neuron cell line PC12 by scavenging ROS. In this study, we demonstrated DMTHB alleviates the behavioral disorder and protects dopaminergic neurons through multiple-target effects includubg anti-apoptotic, anti-inflammatory and antioxidant effects.


Subject(s)
Animals , Dopaminergic Neurons/pathology , Mice , Mice, Inbred C57BL , Parkinson Disease/pathology , Parkinsonian Disorders/chemically induced , Substantia Nigra
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