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1.
Gastroenterology ; 164(4): 630-641.e34, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36623778

RESUMO

BACKGROUND & AIMS: The etiology of abdominal pain in postinfectious, diarrhea-predominant irritable bowel syndrome (PI-IBS-D) is unknown, and few treatment options exist. Catechol-O-methyltransferase (COMT), an enzyme that inactivates and degrades biologically active catecholamines, plays an important role in numerous physiologic processes, including modulation of pain perception. Our objective was to determine the mechanism(s) of how decreased colonic COMT in PI-IBS-D patients contributes to the chronic abdominal pain phenotype after enteric infections. METHODS: Colon neurons, epithelial cells, and macrophages were procured with laser capture microdissection from PI-IBS-D patients to evaluate cell-specific colonic COMT, microRNA-155 (miR-155), and tumor necrosis factor (TNF) α expression levels compared to recovered patients (infection cleared: did not develop PI-IBS-D) and control individuals. COMT-/-, colon-specific COMT-/-, and miR-155-/- mice and human colonoids were used to model phenotypic expression of COMT in PI-IBS-D patients and to investigate signaling pathways linking abdominal pain. Citrobacter rodentium and trinitrobenzene sulfonic acid animal models were used to model postinflammatory changes seen in PI-IBS-D patients. RESULTS: Colonic COMT levels were significantly decreased and correlated with increased visual analog scale abdominal pain ratings in PI-IBS-D patients compared to recovered patients and control individuals. Colonic miR-155 and TNF-α were increased in PI-IBS-D patients with diminished colonic COMT. COMT-/- mice had significantly increased expression of miR-155 and TNF-α in both colon tissues and dorsal root ganglia. Introduction of cV1q antibody (anti-TNF-α) into mice reversed visceral hypersensitivity after C rodentium and trinitrobenzene sulfonic acid. CONCLUSIONS: Decreased colonic COMT in PI-IBS-D patients drives abdominal pain phenotypes via the COMT/miR-155/TNF-α axis. These important findings will allow new treatment paradigms and more targeted and personalized medicine approaches for gastrointestinal disorders after enteric infections.


Assuntos
Síndrome do Intestino Irritável , MicroRNAs , Humanos , Camundongos , Animais , Síndrome do Intestino Irritável/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Catecol O-Metiltransferase/genética , Catecol O-Metiltransferase/metabolismo , Nociceptividade , Inibidores do Fator de Necrose Tumoral , Colo/metabolismo , Dor Abdominal/genética , Dor Abdominal/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Trinitrobenzenos/metabolismo , Ácidos Sulfônicos/metabolismo
2.
Environ Sci Technol ; 58(16): 7020-7031, 2024 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-38608167

RESUMO

Adjusting the electronic state of noble metal catalysts on a nanoscale is crucial for optimizing the performance of nanocatalysts in many important environmental catalytic reactions, particularly in volatile organic compound (VOC) combustion. This study reports a novel strategy for optimizing Pt catalysts by modifying their electronic structure to enhance the electron density of Pt. The research illustrates the optimal 0.2Pt-0.3W/Fe2O3 heterostructure with atomic-thick WO3 layers as a bulking block to electronically modify supported Pt nanoparticles. Methods such as electron microscopy, X-ray photoelectron spectroscopy, and in situ Fourier transform infrared spectroscopy confirm Pt's electron-enriched state resulting from electron transfer from atomic-thick WO3. Testing for benzene oxidation revealed enhanced low-temperature activity with moderate tungsten incorporation. Kinetic and mechanistic analyses provide insights into how the enriched electron density benefits the activation of oxygen and the adsorption of benzene on Pt sites, thereby facilitating the oxidation reaction. This pioneering work on modifying the electronic structure of supported Pt nanocatalysts establishes an innovative catalyst design approach. The electronic structure-performance-dependent relationships presented in this study assist in the rational design of efficient VOC abatement catalysts, contributing to clean energy and environmental solutions.

3.
Biomed Chromatogr ; 38(5): e5835, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38304995

RESUMO

The saponin-enriched extract from Celosiae Semen is a promising resource owing to its lipid-lowering activity. However, triterpenoid saponins are difficult to extract owing to their high molecular weight and strong water solubility. The aim of this paper was to explore an eco-friendly and effective technology of extraction and enrichment of total triterpenoid saponins to obtain high lipid-lowering fractions. Initially, Box-Behnken design experiments were employed to optimize the heat reflux extraction process on the basic of mono-factor experiments. Afterwards, the crude extract was further purified using D-101 resin, and the purification parameters were investigated based on adsorption/desorption experiments and biological activity assay. Under optimal conditions, the purity of the finally obtained total triterpenoid saponins was increased by 7.28-fold. The lipid-lowering activities of the six main triterpenoid saponins were evaluated in HepG2 cells induced by palmitic acid. The results of Oil Red O staining showed that the compounds all exhibited potential lipid-lowering activity. The structure-activity relationship analysis suggested that the oligosaccharide chain at C-28 played an essential role in their lipid-lowering activity and the substituent group at C-23 site also showed important effects. The optimal extraction and purification methods may facilitate the utilization of Celosiae Semen for the industrial production as a functional food and drug.


Assuntos
Amaranthaceae , Metabolismo dos Lipídeos , Saponinas/química , Saponinas/farmacologia , Triterpenos/química , Amaranthaceae/química , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Humanos , Células Hep G2 , Etanol/química
4.
Langmuir ; 39(43): 15343-15354, 2023 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-37857276

RESUMO

Volatile organic compounds such as benzene are hazardous air pollutants that require effective elimination. Noble metal-based catalysts exhibit high benzene combustion activity, but their prohibitive cost necessitates strategies to enhance utilization efficiency. This study investigates a Pt-Cu alloy catalyst for improved benzene combustion by preferentially exposing Pt active sites through Cu alloying. Aberration-corrected scanning transmission electron microscopy and X-ray spectroscopy characterize the nanoscale distribution and enrichment of Pt on the alloy surface. Kinetic measurements demonstrate substantially enhanced activity compared with Pt catalysts, attributed to increased Pt metallic site exposure rather than alteration of the reaction mechanism. In situ Fourier transform infrared (FTIR) spectroscopy reveals a higher abundance of terrace-like Pt sites in the alloy, beneficial for benzene adsorption. Partial pressure dependence analyses indicate competitive adsorption of benzene and O2, following Langmuir-Hinshelwood kinetics. These findings provide conceptual insights into tuning surface composition in bimetallic catalysts to optimize noble metal efficiency, with broad applicability for sustainable catalytic process advancement.

5.
Molecules ; 27(17)2022 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-36080196

RESUMO

Transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel, predominantly expressed in a subset of peripheral sensory neurons for pain signaling. Topical application of agonist capsaicin for desensitizing TRPV1 currents has been approved for relief of chronic pain. However, the potent TRPV1 capsaicin is not ingestible and even topical capsaicin causes common side effects such as skin irritation, swelling, erythema and pruritus, suggesting that a mild TRPV1 agonist might be helpful for reducing side effects while reliving pain. In this study, we reported on a partial and selective TRPV1 agonist 4-(5-chloropyridin-2-yl)-N-(1H-indazol-6-yl)piperazine-1-carboxamide named CPIPC that was modified based on targeting the residue Arg557, important for conversion between the channel antagonism and agonism. Whole-cell patch clamp recordings indicated a concentration-dependent activation of TRPV1 currents by CPIPC with an EC50 of 1.56 ± 0.13 µM. The maximum efficacy of CPIPC (30 µM) was about 60% of saturated capsaicin (10 µM). Repetitive additions of CPIPC caused TRPV1 current desensitization in both TRPV1-expressing HEK293 cells and dorsal root ganglion (DRG) sensory neurons. Oral administration of CPIPC dose-dependently alleviated inflammatory pain in mice. Further site-directed mutagenesis combined with molecular docking revealed that residue Arg557 is critical for TRPV1 activation by CPIPC. Taken together, we identified a novel partial and selective TRPV1 agonist CPIPC that exhibits antinociceptive activity in mice.


Assuntos
Capsaicina , Canais de Cátion TRPV , Animais , Capsaicina/farmacologia , Capsaicina/uso terapêutico , Gânglios Espinais , Células HEK293 , Humanos , Camundongos , Simulação de Acoplamento Molecular , Dor/tratamento farmacológico , Células Receptoras Sensoriais , Canais de Cátion TRPV/agonistas
6.
Hematol Oncol ; 39(4): 490-497, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-33908077

RESUMO

This study aimed to identify the prognostic factors in patients with Waldeyer's ring diffuse large B-cell lymphoma (WR-DLBCL), comparing the efficacy of radiotherapy (RT) for the WR-DLBCL patients in the pre-rituximab and rituximab eras. We conducted a retrospective analysis of 134 patients diagnosed with WR-DLBCL. Univariate and multivariate analyses were performed to identify the prognostic factors for WR-DLBCL. Then, we divided these patients into the rituximab plus chemotherapy group (R-chemotherapy) (n = 88) and chemotherapy group (n = 46), and the Kaplan-Meier and Cox regression model analyses were applied to investigate the treatment value of RT in both the groups. Multivariate analysis revealed international prognostic index (IPI) ≥ 3 and chemotherapy without rituximab as significant risk factors for the progression-free survival (PFS, IPI ≥ 3: p = 0.001; chemotherapy without rituximab: p = 0.002) and overall survival (OS, IPI ≥ 3, p < 0.001; chemotherapy without rituximab, p = 0.024). Rituximab combined with chemotherapy significantly improved PFS (p = 0.002) and OS (p = 0.006) in these patients. RT did not significantly contribute to the survival in the overall cohort analysis, whereas in the subgroup analysis, RT significantly improved the PFS (p = 0.025) and OS (p = 0.029) for the patients in the chemotherapy group, but not in the R-chemotherapy group. In conclusion, the WR-DLBCL patients could benefit from RT in the pre-rituximab era, whereas the addition of rituximab to chemotherapy significantly improved the survival of WR-DLBCL patients, and the clinical benefit of RT was reduced.


Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/radioterapia , Rituximab/uso terapêutico , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos Imunológicos/farmacologia , Humanos , Linfoma Difuso de Grandes Células B/patologia , Pessoa de Meia-Idade , Fatores de Risco , Rituximab/farmacologia , Adulto Jovem
7.
FASEB J ; 34(9): 12338-12353, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32729134

RESUMO

Thermosensitive transient receptor potential vanilloid 2 (thermoTRPV2) is a nonselective Ca2+ -permeable cation channel broadly expressed, and is implicated in the pathology of diseases such as diabetes and pancreatitis. However, the physiological and pharmacological functions of TRPV2 channels have not been extensively investigated because of the absence of specific modulators. In this study, we report a pair of natural coumarin derivative enantiomers (-)-murraxocin (B304-1) and (+)-murraxocin (B304-2) from Murraya exotica for their selective inhibition of TRPV2 channels expressed in HEK293 cells and native TRPV2 currents in differentiated brown adipocytes. Whole-cell patch clamp recordings confirmed the enantiomers B304-1 and B304-2 could selectively inhibit the agonist mediated activation of TRPV2 current with IC50 values of 22.2 ± 7.8 µM and 3.7 ± 0.7 µM, respectively. Molecular docking and site-directed mutagenesis revealed a key residue I600 of TRPV2 critical for the binding of the enantiomers. Furthermore, B304-1 and B304-2 significantly reversed TRPV2 agonist-induced inhibition of mouse brown adipocyte differentiation. Taken together, our identification of two natural coumarin enantiomers provides valuable tools and chemical leads for further elucidation of TRPV2 channel function, and pharmacological modulation of thermoTRPV2 in brown adipocytes may represent a new therapeutic strategy for treatment of energy imbalance or metabolic disorders.


Assuntos
Cumarínicos/farmacologia , Murraya/química , Canais de Cátion TRPV/antagonistas & inibidores , Adipócitos Marrons/citologia , Adipócitos Marrons/efeitos dos fármacos , Animais , Diferenciação Celular/efeitos dos fármacos , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Simulação de Acoplamento Molecular , Mutagênese Sítio-Dirigida , Raízes de Plantas/química , Estereoisomerismo , Canais de Cátion TRPV/química , Canais de Cátion TRPV/fisiologia
8.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 52(5): 813-818, 2021 Sep.
Artigo em Zh | MEDLINE | ID: mdl-34622598

RESUMO

OBJECTIVE: To segment images through an unsupervised method as an alternative to manual labeling. METHODS: A total of 100 whole slide image (WSI) data of HE stained and Pap stained slides were selected as the research and test objects, including 70 breast slides, 20 lung slides and 10 thyroid slides. In order to ensure the diversity of data, the breast slides included those of normal tissue, inflammation and tumor, the lung slides were mainly neoplasms in the lower lobe, including those of inflammation and tumor, and the thyroid slides were of cells, all benign, obtained through fine needle aspiration. The maximum total magnification (original magnification) of each image was 400 times, and the file format was NDPI. Each WSI was manually labeled, and the labeled area of each WSI was more than 10 fields of vision. The labeled information was to be used for validity verification. An unsupervised image segmentation technique based on superpixel and fully convolution neural network algorithms was constructed and used to segment any region of interest (ROI) of unlabeled WSI. In comparison with the region adjacency graph merging method, the segmentation effect of the two methods was assessed with the under segmentation error, the boundary recall and the mean Intersection-over-Union, and the efficiency of the two methods was also compared. In the comparison of execution efficiency, the test process included the preprocessing time of superpixel, and excluded the time of loading the deep learning engine. RESULTS: Unsupervised automatic segmentation was implemented for any ROI region of WSI according to the texture and color. The results of the breast slides, lung slides and thyroid slides showed slight differences, and multiple tests yielded stable results. However, the performance of this method in differentiating inflammation and tumor was average. The under-segmentation error, the boundary recall and the mean Intersection-over-Union were 19.10%, 82.06% and 45.06%, respectively. The under segmentation error, the boundary recall and the mean Intersection-over-Union for the region adjacency graph merging method were 21.52%, 78.39% and 44.81%, respectively. The average time consumption of the whole process was 0.27 s in GPU mode and 1.30 s in CPU mode. The average time consumption of the region adjacency graph merging method was 10.5 s in CPU mode because the method of region adjacency graph merging was not realized in the GPU mode. CONCLUSION: This method produced ideal pixel level labeling results through simple human-computer interaction, which could effectively reduce the cost of digital pathology slide data labeling. Compared with the region adjacency graph merging method, this method had better performance in processing image texture and had faster processing speed.


Assuntos
Algoritmos , Aprendizado de Máquina não Supervisionado , Mama , Humanos , Redes Neurais de Computação
9.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 52(4): 693-697, 2021 Jul.
Artigo em Zh | MEDLINE | ID: mdl-34323051

RESUMO

OBJECTIVE: To study the different methods of artificial intelligence (AI)-assisted Ki-67 scoring of clinical invasive ductal carcinoma (IDC) of the breast and to compare the results. METHODS: A total of 100 diagnosed IDC cases were collected, including slides of HE staining and immunohistochemical Ki-67 staining and diagnosis results. The slides were scanned and turned into whole slide image (WSI), which were then scored with AI. There were two AI scoring methods. One was fully automatic counting by AI, which used the scoring system of Ki-67 automatic diagnosis to do counting with the whole image of WSI. The second method was semi-automatic AI counting, which required manual selection of areas for counting, and then relied on an intelligent microscope to conduct automatic counting. The diagnostic results of pathologists were taken as the results of pure manual counting. Then the Ki-67 scores obtained by manual counting, semi-automatic AI counting and automatic AI counting were pairwise compared. The Ki-67 scores obtained from the manual counting (pathological diagnosis results), semi-automatic AI and automatic AI counts were pair-wise compared and classified according to three levels of difference: difference ≤10%, difference of >10%-<30% and difference ≥30%. Intra-class correlation coefficient ( ICC) was used to evaluate the correlation. RESULTS: The automatic AI counting of Ki-67 takes 5-8 minutes per case, the semi-automatic AI counting takes 2-3 minutes per case, and the manual counting takes 1-3 minutes per case. When results of the two AI counting methods were compared, the difference in Ki-67 scores was all within 10% (100% of the total), and the ICC index being 0.992. The difference between manual counting and semi-automatic AI was less than 10% in 60 cases (60% of the total), between 10% and 30% in 37 cases (37% of the total), and more than 30% in only 3 cases (3% of the total), ICC index being 0.724. When comparing automatic AI with manual counting, 78 cases (78% of the total) had a difference of ≤10%, 17 cases (17% of the total) had a difference of between 10% and 30%, and 5 cases (5%) had a difference of ≥30%, the ICC index being 0.720. The ICC values showed that there was little difference between the results of the two AI counting methods, indicating good repeatability, but the repeatability between AI counting and manual counting was not particularly ideal. CONCLUSION: AI automatic counting has the advantage of requiring less manpower, for the pathologist is involved only for the verification of the diagnosis results at the end. However, the semi-automatic method is better suited to the diagnostic habits of pathologists and has a shorter turn-over time compared with that of the fully automatic AI counting method. Furthermore, in spite of its higher repeatability, AI counting, cannot serve as a full substitute for pathologists, but should instead be viewed as a powerful auxiliary tool.


Assuntos
Neoplasias da Mama , Inteligência Artificial , Neoplasias da Mama/diagnóstico , Feminino , Humanos , Antígeno Ki-67 , Microscopia , Reprodutibilidade dos Testes
10.
Anal Chem ; 92(2): 1934-1939, 2020 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-31855414

RESUMO

Fluorescent probes have been used as effective methods for profiling proteins in biological systems because of their high selectivity, sensitivity, and temporal-spatial resolution. A specific fluorescent probe for understanding the function of the transient receptor potential ankyrin 1 (TRPA1) channel that is closely related with various diseases like persistent pain, respiratory, and chronic itch syndromes, however, is still lacking. Here, we report a "turn-on" fluorescent probe (A1CA) for visualizing TRPA1 channels in the plasma membrane of live cells based on a photochromic ligand derived from 4-(phenylazo)benzenamine. Evaluating the specificity and sensitivity of A1CA by electrophysiology and confocal imaging showed that the A1CA probe displays higher affinity and selectivity to TRPA1 channel versus all other ion channels including TRPV1, TRPV3, Nav1.4, Nav1.5, and hERG. Based on the supporting evidence, A1CA has great potential as a molecular imaging probe for high-throughput screening of novel TRPA1 agonists.


Assuntos
Compostos Azo/química , Membrana Celular/química , Cumarínicos/química , Corantes Fluorescentes/química , Canal de Cátion TRPA1/análise , Animais , Compostos Azo/síntese química , Células CHO , Cumarínicos/síntese química , Cricetulus , Eletrofisiologia/métodos , Corantes Fluorescentes/síntese química , Ligantes , Microscopia Confocal/métodos , Canal de Cátion TRPA1/agonistas , Canal de Cátion TRPA1/antagonistas & inibidores
11.
Gut ; 68(6): 996-1002, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30108163

RESUMO

BACKGROUND: More effective treatments are needed for patients with postinfectious, diarrhoea-predominant, irritable bowel syndrome (IBS-D). Accordingly, we conducted a randomised, double-blind, placebo-controlled, 8-week-long trial to assess the efficacy and safety of oral glutamine therapy in patients who developed IBS-D with increased intestinal permeability following an enteric infection. METHODS: Eligible adults were randomised to glutamine (5 g/t.i.d.) or placebo for 8 weeks. The primary end point was a reduction of ≥50 points on the Irritable Bowel Syndrome Severity Scoring System (IBS-SS). Secondary endpoints included: raw IBS-SS scores, changes in daily bowel movement frequency, stool form (Bristol Stool Scale) and intestinal permeability. RESULTS: Fifty-four glutamine and 52 placebo subjects completed the 8-week study. The primary endpoint occurred in 43 (79.6%) in the glutamine group and 3 (5.8%) in the placebo group (a 14-fold difference). Glutamine also reduced all secondary endpoint means: IBS-SS score at 8 weeks (301 vs 181, p<0.0001), daily bowel movement frequency (5.4 vs 2.9±1.0, p<0.0001), Bristol Stool Scale (6.5 vs 3.9, p<0.0001) and intestinal permeability (0.11 vs 0.05; p<0.0001). 'Intestinal hyperpermeability' (elevated urinary lactulose/mannitol ratios) was normalised in the glutamine but not the control group. Adverse events and rates of study-drug discontinuation were low and similar in the two groups. No serious adverse events were observed. CONCLUSIONS: In patients with IBS-D with intestinal hyperpermeability following an enteric infection, oral dietary glutamine supplements dramatically and safely reduced all major IBS-related endpoints. Large randomised clinical trials (RCTs) should now be done to validate these findings, assess quality of life benefits and explore pharmacological mechanisms. TRIAL REGISTRATION NUMBER: NCT01414244; Results.


Assuntos
Suplementos Nutricionais , Enterite/microbiologia , Glutamina/uso terapêutico , Síndrome do Intestino Irritável/tratamento farmacológico , Administração Oral , Adulto , Relação Dose-Resposta a Droga , Método Duplo-Cego , Esquema de Medicação , Enterite/complicações , Feminino , Humanos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/microbiologia , Síndrome do Intestino Irritável/diagnóstico , Síndrome do Intestino Irritável/microbiologia , Masculino , Valores de Referência , Medição de Risco , Índice de Gravidade de Doença , Resultado do Tratamento
12.
Biochem Biophys Res Commun ; 520(1): 60-66, 2019 11 26.
Artigo em Inglês | MEDLINE | ID: mdl-31570165

RESUMO

Human TRDMT1 is a transfer RNA (tRNA) methyltransferase for cytosine-5 methylation and has been suggested to be involved in the regulation of numerous developmental processes. However, little is known about the molecular mechanisms or their biological significance. In this study, we investigated the effects of CRISPR-based TRDMT1 knockdown on phenotypes, mRNA m5C modifications and gene expression changes in HEK293 cells. We found that knockdown of TRDMT1 significantly inhibited cell proliferation and migration but had no effect on clonogenic potential. The inhibitory effects could be attenuated by re-expression of TRDMT1 in HEK293 cells. RNA sequencing (RNA-Seq) and RNA bisulfite sequencing (RNA-BisSeq) were performed in TRDMT1 knockdown and wild-type HEK293 cells. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses indicated that the differentially expressed genes were associated with the cell cycle, RNA transport, and RNA degradation and were enriched in cancer and Notch signaling pathways. We also found that TRDMT1 knockdown could change mRNA methylation levels. For the first time, these findings clarify the role of TRDMT1 in regulating mRNA methylation and inhibiting the proliferation and migration of HEK293 cells. These results provide new insights into a new function of TRDMT1 and elucidate the molecular mechanisms of aberrant RNA m5C during tumorigenesis.


Assuntos
5-Metilcitosina/metabolismo , Movimento Celular , Proliferação de Células , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , RNA Mensageiro/metabolismo , Sistemas CRISPR-Cas , Carcinogênese , Biologia Computacional , Metilação de DNA , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Fenótipo , RNA-Seq , Transdução de Sinais
13.
J Pharmacol Exp Ther ; 368(1): 21-31, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30377214

RESUMO

The temperature-sensitive and calcium-permeable transient receptor potential vanilloid 3 (TRPV3) channel abundantly expressed in keratinocytes plays important functions in skin physiology. Dysfunctional gain-of-function TRPV3 gene mutations cause genetic Olmsted syndrome characterized by periorificial keratoderma, palmoplantar keratoderma, inflammation, and severe itching, which suggests that pharmacological inhibition of overactive TRPV3 function may be beneficial in treating pruritus or skin disorders. To test this hypothesis, we identified natural compound forsythoside B as a TRPV3 inhibitor through screening of human embryonic kidney 293 (HEK293) cells expressing human TRPV3 channels in a calcium fluorescent assay. Whole-cell patch-clamp recordings of HEK293 cells expressing TRPV3 confirmed that forsythoside B selectively inhibited the channel current activated by agonist 2-aminoethoxydiphenyl borate (50 µM) in a dose-dependent fashion, with an IC50 value of 6.7 ± 0.7 µM. In vivo evaluation of scratching behavior demonstrated that pharmacological inhibition of TRPV3 by forsythoside B significantly attenuated acute itch induced by either the TRPV3 agonist carvacrol or the pruritogen histamine, as well as chronic itch induced by acetone-ether-water in a mouse model of dry skin. Furthermore, forsythoside B was able to prevent the death of HEK293 cells or native human immortalized nontumorigenic keratinocyte cells from human keratinocytes expressing a gain-of-function TRPV3 G573S mutant or in the presence of the TRPV3 agonist carvacrol. Taken together, our findings demonstrate the crucial role of TRPV3 in pruritus and keratinocyte toxicity; thus, specific inhibition of overactive TRPV3 by natural forsythoside B may possess therapeutic potential for treatment of chronic pruritus, skin allergy, or inflammation-related skin diseases.


Assuntos
Ácidos Cafeicos/uso terapêutico , Sinalização do Cálcio/efeitos dos fármacos , Glucosídeos/uso terapêutico , Queratinócitos/metabolismo , Prurido/tratamento farmacológico , Canais de Cátion TRPV/metabolismo , Animais , Ácidos Cafeicos/farmacologia , Sinalização do Cálcio/fisiologia , Linhagem Celular Transformada , Cimenos , Relação Dose-Resposta a Droga , Glucosídeos/farmacologia , Células HEK293 , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Monoterpenos/farmacologia , Prurido/metabolismo , Prurido/patologia , Canais de Cátion TRPV/agonistas , Temperatura
14.
J Clin Gastroenterol ; 53(7): e298-e302, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30260809

RESUMO

BACKGROUND: Well over 700,000 United States military personnel participated in the Persian Gulf War in which they developed chronic health disorders of undetermined etiology. Up to 25% of Veterans had persistent and chronic gastrointestinal (GI) symptoms, which they suspected were related to their military service in the Gulf. AIM: The overall aim of the current study was to evaluate intestinal permeability in previously deployed Gulf War Veterans who developed chronic GI symptoms during their tour in the Persian Gulf. METHODS: To accomplish this, we evaluated intestinal permeability (IP) using the urinary lactulose/mannitol test. Measurements of intestinal permeability were then correlated with mean ratings of daily abdominal pain, frequency of bowel movements, and consistency of bowel movements on the Bristol Stool Scale in all Veterans. RESULTS: A total of 73 veterans had documented chronic GI symptoms (diarrhea, abdominal pain) and were included in the study. A total of 29/73 (39%) of veterans has increased IP and had a higher average daily stool frequency (P<0.05); increased liquid stools as indicated by a higher Bristol Stool Scale (P<0.01); and a higher mean M-VAS abdominal pain rating (P<0.01). Pearson correlation coefficients revealed that there was a positive correlation between increased IP and stool frequency, Bristol Stool Scale, and M-VAS abdominal pain rating. CONCLUSIONS: Our study demonstrates that deployed Gulf War Veterans with persistent GI symptoms commonly have increased intestinal permeability that potentiates the severity of abdominal pain, diarrhea, and stool consistency. These new findings in our study are important as they may lead to novel diagnostic biomarkers for returning Gulf War Veterans who suffer from chronic functional gastrointestinal disorders. These advances are also important for an increasing number of veterans who are now serving in the Persian Gulf and are at a high risk of developing these chronic pain disorders.


Assuntos
Dor Abdominal/etiologia , Diarreia/epidemiologia , Gastroenteropatias/fisiopatologia , Intestinos/fisiopatologia , Dor Abdominal/epidemiologia , Adulto , Doença Crônica , Feminino , Gastroenteropatias/diagnóstico , Guerra do Golfo , Humanos , Lactulose/urina , Masculino , Manitol/urina , Pessoa de Meia-Idade , Permeabilidade , Estados Unidos , Veteranos
15.
Acta Pharmacol Sin ; 40(6): 737-745, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30333556

RESUMO

The α7 nicotinic acetylcholine receptor (α7 nAChR) is a ligand-gated Ca2+-permeable homopentameric ion channel implicated in cognition and neuropsychiatric disorders. Pharmacological enhancement of α7 nAChR function has been suggested for improvement of cognitive deficits. In the present study, we characterized a thiazolyl heterocyclic derivative, 6-(2-chloro-6-methylphenyl)-2-((3-fluoro-4-methylphenyl)amino)thiazolo[4,5-d]pyrimidin-7(6H)-one (JWX-A0108), as a novel type I α7 nAChR positive allosteric modulator (PAM), and evaluated its ability to reverse auditory gating and spatial working memory deficits in mice. In Xenopus oocytes expressing human nAChR channels, application of JWX-A0108 selectively enhanced α7 nAChR-mediated inward current in the presence of the agonist ACh (EC50 value = 4.35 ± 0.12 µM). In hippocampal slices, co-application of ACh and JWX-A0108 (10 µM for each) markedly increased both the frequency and amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs) recorded in pyramidal neurons, but JWX-A0108 did not affect GABA-induced current in oocytes expressing human GABAA receptor α1ß3γ2 and α5ß3γ2 subtypes. In mice with MK-801-induced deficits in auditory gating, administration of JWX-A0108 (1, 3, and 10 mg/kg, i.p.) dose-dependently attenuates MK-801-induced auditory gating deficits in five prepulse intensities (72, 76, 80, 84, and 88 dB). Furthermore, administration of JWX-A0108 (0.03, 0.1, or 0.3 mg/kg, i.p.) significantly reversed MK-801-induced impaired spatial working memory in mice. Our results demonstrate that JWX-A0108 is a novel type I PAM of α7 nAChR, which may be beneficial for improvement of cognitive deficits commonly found in neuropsychiatric disorders such as schizophrenia and Alzheimer's disease.


Assuntos
Nootrópicos/uso terapêutico , Inibição Pré-Pulso/efeitos dos fármacos , Filtro Sensorial/efeitos dos fármacos , Tiazóis/uso terapêutico , Receptor Nicotínico de Acetilcolina alfa7/agonistas , Animais , Maleato de Dizocilpina , Hipocampo/efeitos dos fármacos , Hipocampo/fisiopatologia , Humanos , Interneurônios/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Transtornos da Memória/tratamento farmacológico , Camundongos Endogâmicos C57BL , Nootrópicos/farmacocinética , Nootrópicos/farmacologia , Ratos Sprague-Dawley , Esquizofrenia/induzido quimicamente , Esquizofrenia/tratamento farmacológico , Transmissão Sináptica/efeitos dos fármacos , Tiazóis/farmacocinética , Tiazóis/farmacologia , Xenopus
17.
Gut ; 65(5): 797-805, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-25681400

RESUMO

OBJECTIVE: Many patients with irritable bowel syndrome IBS not only have abdominal pain but also may suffer from visceral hypersensitivity and heighted visceral nociception. Moreover, IBS has few effective therapeutic agents and mechanisms of disease are unclear. Our goals were to (i) identify microRNA (miRNA) expression, signalling and targets in human colon (controls; patients with IBS); (ii) verify in vitro, IBS-associated changes in miRNAs, especially miR-199, which is complementary to the transient receptor potential vanilloid type 1 (TRPV1) gene; and (iii) determine whether modulating the expression of miRNAs in vivo, especially miR-199, reverses associated changes and pathological hallmarks of visceral hypersensitivity via TRPV1 signalling. DESIGN: We evaluated 45 patients with diarrhoea-predominant IBS (IBS-D) and 40 controls with (1) visceral pain severity score and (2) colonoscopy with biopsies. miRNA expression was evaluated in human colon following miRNA array analysis. Luciferase assays were done to confirm relationships between miR-199 and TRPV1 expression. A rat model of visceral hypersensitivity was used to study miR-199 and its target gene (TRPV1) expression in dorsal root ganglion (DRG) and colon in vivo. RESULTS: Gut miR-199a/b expression in IBS-D was significantly decreased, which correlated directly with both increased visceral pain scores and TRPV1 expression. In vivo upregulation of miR-199a by intraperitoneal injection of lenti-miR-199a precursors decreased visceral hypersensitivity via diminished TRPV1 signalling. CONCLUSIONS: Decreased colonic miR-199a/b correlates with visceral pain in patients with IBS-D. Similarly, reduced miR-199a expression in rat DRG and colon tissue is associated with heightened visceral hypersensitivity. In vivo upregulation of miR-199a decreases visceral pain via inhibition of TRPV1 signalling. Thus, miR-199 precursors may be promising therapeutic candidates for the treatment in patients with visceral pain.


Assuntos
Síndrome do Intestino Irritável/genética , MicroRNAs/genética , Canais de Cátion TRPV/fisiologia , Regulação para Cima , Dor Visceral/genética , Animais , Colo , Regulação da Expressão Gênica , Humanos , Síndrome do Intestino Irritável/complicações , Masculino , Medição da Dor , Biossíntese de Proteínas , Ratos , Ratos Endogâmicos F344 , Dor Visceral/etiologia
19.
Gastroenterology ; 148(1): 158-169.e8, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25277410

RESUMO

BACKGROUND & AIMS: Some patients with irritable bowel syndrome with diarrhea (IBS-D) have intestinal hyperpermeability, which contributes to their diarrhea and abdominal pain. MicroRNA 29 (MIR29) regulates intestinal permeability in patients with IBS-D. We investigated and searched for targets of MIR29 and investigated the effects of disrupting Mir29 in mice. METHODS: We investigated expression MIR29A and B in intestinal biopsies collected during endoscopy from patients with IBS (n = 183) and without IBS (controls) (n = 36). Levels were correlated with disease phenotype. We also generated and studied Mir29(-/-) mice, in which expression of Mir29a and b, but not c, is lost. Colitis was induced by administration of 2,4,6-trinitrobenzenesulfonic acid; intestinal tissues were collected and permeability was assessed. Microarray analysis was performed using tissues from Mir29(-/-) mice. Changes in levels of target genes were measured in human colonic epithelial cells and small intestinal epithelial cells after knockdown of MIR29 with anti-MIRs. RESULTS: Intestinal tissues from patients with IBS-D (but not IBS with constipation or controls) had increased levels of MIR29A and B, but reduced levels of Claudin-1 (CLDN1) and nuclear factor-κB-repressing factor (NKRF). Induction of colitis and water avoidance stress increased levels of Mir29a and Mir29b and intestinal permeability in wild-type mice; these increased intestinal permeability in colons of far fewer Mir29(-/-) mice. In microarray and knockdown experiments, MIR29A and B were found to reduce levels of NKRF and CLDN1 messenger RNA, and alter levels of other messenger RNAs that regulate intestinal permeability. CONCLUSIONS: Based on experiments in knockout mice and analyses of intestinal tissue samples from patients with IBS-D, MIR29 targets and reduces expression of CLDN1 and NKRF to increase intestinal permeability. Strategies to block MIR29 might be developed to restore intestinal permeability in patients with IBS-D.


Assuntos
Claudina-1/metabolismo , Colite/metabolismo , Colo/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , MicroRNAs/metabolismo , Proteínas Repressoras/metabolismo , Animais , Estudos de Casos e Controles , Linhagem Celular , Claudina-1/genética , Colite/induzido quimicamente , Colite/genética , Colite/patologia , Colo/patologia , Modelos Animais de Doenças , Regulação para Baixo , Técnicas de Silenciamento de Genes , Predisposição Genética para Doença , Humanos , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/genética , Permeabilidade , Fenótipo , RNA Mensageiro/metabolismo , Proteínas Repressoras/genética , Transdução de Sinais , Ácido Trinitrobenzenossulfônico
20.
Phytomedicine ; 130: 155734, 2024 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-38761775

RESUMO

BACKGROUND: Toxic components frequently exhibit unique characteristics and activities, offering ample opportunities for the advancement of anti-cancer medications. As the main hepatotoxic component of Dioscorea bulbifera L. (DB), Diosbulbin B (DIOB) has been widely studied for its anti-tumor activity at nontoxic doses. However, the effectiveness and mechanism of DIOB against non-small cell lung cancer (NSCLC) remains unclear. PURPOSE: To evaluate the anti-NSCLC activity of DIOB and to elucidate the specific mechanism of action. METHOD: The effect of DIOB on NSCLCL in vitro was evaluated through CCK8, colony formation, and flow cytometry. The in vivo efficacy and safety of DIOB in treating NSCLC were assessed using various techniques, including HE staining, tunel staining, immunohistochemistry, and biochemical index detection. To understand the underlying mechanism, cell transfection, western blotting, molecular docking, cellular thermal shift assay (CESTA), and surface plasmon resonance (SPR) were employed for investigation. RESULTS: DIOB effectively hindered the progression of NSCLC both in vitro and in vivo settings at a no-observed-adverse-effect concentration (NOAEC) and a safe dosage. Specifically, DIOB induced significant G0/G1 phase arrest and apoptosis in A549, PC-9, and H1299 cells, while also notably inhibiting the growth of subcutaneous tumors in nude mice. Mechanistically, DIOB could directly interact with oncogene Yin Yang 1 (YY1) and inhibit its expression. The reduction in YY1 resulted in the triggering of the tumor suppressor P53, which induced cell cycle arrest and apoptosis in NSCLC cells by inhibiting the expression of Cyclin A2, B2, CDK1, CDK2, CDK4, BCL-2, and inducing the expression of BAX. In NSCLC cells, the induction of G0/G1 phase arrest and apoptosis by DIOB was effectively reversed when YY1 was overexpressed or P53 was knocked down. Importantly, we observed that DIOB exerted the same effect by directly influencing the expression of YY1-regulated c-Myc and BIM, particularly in the absence of P53. CONCLUSION: For the inaugural investigation, this research unveiled the anti-NSCLC impact of DIOB, alongside its fundamental mechanism. DIOB has demonstrated potential as a treatment agent for NSCLC due to its impressive efficacy in countering NSCLC.


Assuntos
Apoptose , Carcinoma Pulmonar de Células não Pequenas , Pontos de Checagem do Ciclo Celular , Compostos Heterocíclicos de 4 ou mais Anéis , Neoplasias Pulmonares , Camundongos Nus , Fator de Transcrição YY1 , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Humanos , Apoptose/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Animais , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Antineoplásicos Fitogênicos/farmacologia , Dioscorea/química , Camundongos , Camundongos Endogâmicos BALB C , Masculino , Simulação de Acoplamento Molecular , Células A549 , Proteína Supressora de Tumor p53/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
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