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1.
Nat Metab ; 2021 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-34663975

RESUMO

5-diphosphoinositol pentakisphosphate (5-IP7) is a signalling metabolite linked to various cellular processes. How extracellular stimuli elicit 5-IP7 signalling remains unclear. Here we show that 5-IP7 in ß cells mediates parasympathetic stimulation of synaptotagmin-7 (Syt7)-dependent insulin release. Mechanistically, vagal stimulation and activation of muscarinic acetylcholine receptors triggers Gαq-PLC-PKC-PKD-dependent signalling and activates IP6K1, the 5-IP7 synthase. Whereas both 5-IP7 and its precursor IP6 compete with PIP2 for binding to Syt7, Ca2+ selectively binds 5-IP7 with high affinity, freeing Syt7 to enable fusion of insulin-containing vesicles with the cell membrane. ß-cell-specific IP6K1 deletion diminishes insulin secretion and glucose clearance elicited by muscarinic stimulation, whereas mice carrying a phosphorylation-mimicking, hyperactive IP6K1 mutant display augmented insulin release, congenital hyperinsulinaemia and obesity. These phenotypes are absent in mice lacking Syt7. Our study proposes a new conceptual framework for inositol pyrophosphate physiology in which 5-IP7 acts as a GPCR second messenger at the interface between peripheral nervous system and metabolic organs, transmitting Gq-coupled GPCR stimulation to unclamp Syt7-dependent, and perhaps other, exocytotic events.

3.
J Med Chem ; 64(18): 13475-13486, 2021 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-34506131

RESUMO

Beclin 1 is an essential autophagy gene and a haploinsufficient tumor suppressor. Beclin 1 is the scaffolding member of the Class III phosphatidylinositol-3-kinase complex (PI3KC3) and recruits two positive regulators Atg14L and UVRAG through its coiled-coil domain to upregulate PI3KC3 activity. Our previous work has shown that hydrocarbon-stapled peptides targeted to the Beclin 1 coiled-coil domain reduced Beclin 1 homodimerization and promoted the Beclin 1-Atg14L/UVRAG interaction. These peptides also induced autophagy and enhanced the endolysosomal degradation of cell surface receptors like EGFR. Here, we present the optimization of these Beclin 1-targeting peptides by staple scanning and sequence permutation. Placing the hydrocarbon staple closer to the Beclin 1-peptide interface enhanced their binding affinity by ∼10- to 30-fold. Optimized peptides showed potent antiproliferative efficacy in cancer cells that overexpressed EGFR and HER2 by inducing necrotic cell death but not apoptosis. Our Beclin 1-targeting stapled peptides may serve as effective therapeutic candidates for EGFR- or HER2-driven cancer.

4.
J Phys Condens Matter ; 33(48)2021 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-34492654

RESUMO

A special kind of pentamode metamaterial, which is composed of double-cone elements and forms simple-cubic lattice, is investigated in this paper. Based on finite-element method, the phonon band structure and the transmission spectrum of the pentamode metamaterial are calculated, and then the pentamodal behavior of this structure is theoretically verified from two aspects, i.e. the physical properties and the mathematical definition. Results show that in the phonon band structure, there is a wider single-mode band gap, corresponding to the remarkable loss in the transmission spectrum. The ratio of bulk modulusBto shear modulusGis more than 300, which is obviously larger than that of traditional materials. Except for the isotropic bulk modulusB, the other mechanical moduli are all anisotropic. Five of six eigenvalues of elastic coefficient matrix are nearly zero for the microstructure, and only one is non-zero. These results demonstrate that this metamaterial microstructure performs excellent pentamodal characteristics, and also conforms to the mathematical definition of 'pentamode'.

5.
Int J Mol Sci ; 22(15)2021 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-34360755

RESUMO

Increasing attention is being focused on the use of polypeptide-based N-methyl-d-aspartate (NMDA) receptor antagonists for the treatment of nervous system disorders. In our study on Achyranthes bidentata Blume, we identified an NMDA receptor subtype 2B (NR2B) antagonist that exerts distinct neuroprotective actions. This antagonist is a 33 amino acid peptide, named bidentatide, which contains three disulfide bridges that form a cysteine knot motif. We determined the neuroactive potential of bidentatide by evaluating its in vitro effects against NMDA-mediated excitotoxicity. The results showed that pretreating primary cultured hippocampal neurons with bidentatide prevented NMDA-induced cell death and apoptosis via multiple mechanisms that involved intracellular Ca2+ inhibition, NMDA current inhibition, and apoptosis-related protein expression regulation. These mechanisms were all dependent on bidentatide-induced inhibitory regulation of NR2B-containing NMDA receptors; thus, bidentatide may contribute to the development of neuroprotective agents that would likely possess the high selectivity and safety profiles inherent in peptide drugs.


Assuntos
Achyranthes/química , Hipocampo/metabolismo , Neurônios/metabolismo , Fármacos Neuroprotetores , Peptídeos , Proteínas de Plantas , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/isolamento & purificação , Fármacos Neuroprotetores/farmacologia , Peptídeos/química , Peptídeos/isolamento & purificação , Peptídeos/farmacologia , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/metabolismo
6.
Anal Chem ; 93(32): 11072-11080, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34342978

RESUMO

Pyrylium salts are considered efficient chemical tags for amino groups. However, the apparent steric selectivity of pyrylium salts limits their application in the field of chemical labeling, especially during the labeling of sterically hindered compounds like amino acids, peptides, and proteins. Herein, we have investigated the effects of the α-substitution of pyrylium salts on their reactivity. We have also investigated the mechanism of nucleophilic reactions with pyrylium salts and further proposed that the reactivity of pyrylium salts mainly depends on the position and type of their substituents. A series of pyrylium salts were synthesized, and a highly active α-monosubstituted pyrylium salt, 2,4,5-triphenylpyrylium, was developed for efficient chemical labeling. All of the 15 amino acids studied were efficiently labeled under optimized reaction conditions. The 2,4,5-triphenylpyrylium salt was highly efficient in comparison to the previously reported 2,4,6-triphenylpyrylium salt developed for lysine-specific modifications. Furthermore, we successfully used 2,4,5-triphenylpyrylium salt for the hydrophobic labeling of peptides and protein hydrolysates. The most striking observation was that the ionization efficiency of short-chain multilabeled peptides in mixed samples, after derivatization, increased by up to 60 times. The increase in ionization efficiency gradually decreased with increasing peptide chain length. During the "soft" collision-induced dissociation (CID) process, the peptide was tagged at the N-terminus with 2,4,5-triphenylpyrylium, producing abundant a-type ions and b-type ions (Δ = 28), which eases the peptide resequencing process and assists in cracking the peptide codes. Moreover, 2,4,5-triphenylpyrylium has been utilized for the proteomic analysis of HeLa cell digests. In addition, 215 additional proteins were identified in the labeled products and the coverage of most proteins was improved.


Assuntos
Peptídeos , Proteômica , Células HeLa , Humanos , Indicadores e Reagentes , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem
7.
Anal Chim Acta ; 1175: 338759, 2021 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-34330437

RESUMO

We report here an easy-fabricated and disposable polymer-film microfluidic impedance cytometer (PMIC) integrated with inertial focusing and parallel facing electrodes for cell sensing. The cells are first focused in an asymmetric serpentine channel, and then their impedance signals are measured when passing through the electrode region. The proposed PMIC device is the first impedance cytometer that is fabricated into a flexible sheet (with a thickness of 0.45 mm) by using the materials of commonly-available ITO-coated polymer films and double-sided adhesive tapes, the whole fabrication process is shortened from traditional 3-4 days to less than 5 min by using UV laser cutting. To verify the feasibility of our device for cell sensing, we explore the focusing behaviors of three differently sized particles and two types of tumor cells, and analyze their impedance signals. The results show that our device is capable of obtaining impedance information on numbers, diameters, and longitudinal positions of cells. We envision that our PMIC device is promising in label-free cell sensing owning to the advantages of low cost, small footprint, and simple fabrication.


Assuntos
Microfluídica , Polímeros , Impedância Elétrica , Eletrodos , Dispositivos Lab-On-A-Chip
8.
Proteomics Clin Appl ; : e2000058, 2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34329527

RESUMO

PURPOSE: There are great demands for identifying biomarkers of major depressive disorder (MDD), a common mental illness with a prevalence of approximately 6%. Finding potential biomarkers to aid MDD diagnosis is in high demand. EXPERIMENTAL DESIGN: In this study, a combination of pretreatment methods named salt-out assisted liquid-liquid extraction (SALLE) and nontargeted peptidomics based on nano-LC-Orbitrap/MS was primarily employed to discover the candidate peptide markers from the plasma of 238 subjects. RESULTS: Many peptides were enriched and identified from the plasma, 42 of which showed significant differences between MDD patients and controls by univariate statistical analysis. A binary logistic regression (BLR) model combined four peptide markers (P1, P9, P17, P29) was established, yielding an overall prediction accuracy of 91.7% and 82.2% in the discovery and validation sets, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: In conclusion, the excellent performance of the BLR model in both discovery and validation sets demonstrates the robustness of the four peptide markers panel. It is very valuable for quantification of the absolute content of four peptides and further verification.

9.
Talanta ; 233: 122571, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34215067

RESUMO

Single-cell analysis has gained considerable attention for disease diagnosis, drug screening, and differentiation monitoring. Compared to the well-established flow cytometry, which uses fluorescent-labeled antibodies, microfluidic impedance cytometry (MIC) offers a simple, label-free, and noninvasive method for counting, classifying, and monitoring cells. Superior features including a small footprint, low reagent consumption, and ease of use have also been reported. The MIC device detects changes in the impedance signal caused by cells passing through the sensing/electric field zone, which can extract information regarding the size, shape, and dielectric properties of these cells. According to recent studies, electrode configuration has a remarkable effect on detection accuracy, sensitivity, and throughput. With the improvement in microfabrication technology, various electrode configurations have been reported for improving detection accuracy and throughput. However, the various electrode configurations of MIC devices have not been reviewed. In this review, the theoretical background of the impedance technique for single-cell analysis is introduced. Then, two-dimensional, three-dimensional, and liquid electrode configurations are discussed separately; their sensing mechanisms, fabrication processes, advantages, disadvantages, and applications are also described in detail. Finally, the current limitations and future perspectives of these electrode configurations are summarized. The main aim of this review is to offer a guide for researchers on the ongoing advancement in electrode configuration designs.


Assuntos
Técnicas Analíticas Microfluídicas , Microfluídica , Impedância Elétrica , Eletrodos , Citometria de Fluxo , Dispositivos Lab-On-A-Chip
10.
Nat Commun ; 12(1): 2461, 2021 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-33911083

RESUMO

COP1 and COP9 signalosome (CSN) are the substrate receptor and deneddylase of CRL4 E3 ligase, respectively. How they functionally interact remains unclear. Here, we uncover COP1-CSN antagonism during glucose-induced insulin secretion. Heterozygous Csn2WT/K70E mice with partially disrupted binding of IP6, a CSN cofactor, display congenital hyperinsulinism and insulin resistance. This is due to increased Cul4 neddylation, CRL4COP1 E3 assembly, and ubiquitylation of ETV5, an obesity-associated transcriptional suppressor of insulin secretion. Hyperglycemia reciprocally regulates CRL4-CSN versus CRL4COP1 assembly to promote ETV5 degradation. Excessive ETV5 degradation is a hallmark of Csn2WT/K70E, high-fat diet-treated, and ob/ob mice. The CRL neddylation inhibitor Pevonedistat/MLN4924 stabilizes ETV5 and remediates the hyperinsulinemia and obesity/diabetes phenotypes of these mice. These observations were extended to human islets and EndoC-ßH1 cells. Thus, a CRL4COP1-ETV5 proteolytic checkpoint licensing GSIS is safeguarded by IP6-assisted CSN-COP1 competition. Deregulation of the IP6-CSN-CRL4COP1-ETV5 axis underlies hyperinsulinemia and can be intervened to reduce obesity and diabetic risk.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Secreção de Insulina/fisiologia , Células Secretoras de Insulina/metabolismo , Proteínas Nucleares/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Complexos Ubiquitina-Proteína Ligase/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Complexo do Signalossomo COP9/metabolismo , Linhagem Celular , Ciclopentanos/farmacologia , Diabetes Mellitus/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Glucose/metabolismo , Células HEK293 , Humanos , Hiperinsulinismo/tratamento farmacológico , Secreção de Insulina/genética , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/tratamento farmacológico , Pirimidinas/farmacologia
11.
J Biomol Struct Dyn ; : 1-12, 2021 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-33826484

RESUMO

Interaction between the SARS-COV-2 (2019 novel coronavirus) spike protein and ACE2 receptors expressed on cellular surfaces initialises viral attachment and consequent infection. Blocking this interaction shows promise for blocking or ameliorating the virus' pathological effects on the body. By contrast to work focusing on the coronavirus, which has significant potential diversity through possible accumulation of mutations during transmission, targeting the conserved ACE2 protein expressed on human cells offers an attractive alternative route to developing pharmacological prophylactics against viral invasion. In this study, we screened a virtual database of natural peptides in silico, with ACE2 as the target, and performed structural analyses of the interface region in the SARS-COV-2 RBD/ACE2 complex. These analyses have identified 15 potentially effective compounds. Analyses of ACE2/polypeptide interactions suggest that these peptides can block viral invasion of cells by stably binding in the ACE2 active site pocket. Molecular simulation results for Complestatin and Valinomycin indicate that they may share this mechanism. The discovery of this probable binding mechanism provides a frame of reference for further optimization, and design of high affinity ACE2 inhibitors that could serve as leads for production of drugs with preventive and therapeutic effects against SARS-COV-2.Communicated by Ramaswamy H. Sarma.

12.
J Phys Condens Matter ; 33(18)2021 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-33721850

RESUMO

An effective method for realizing ultra-low-frequency single-mode band gap in pentamode metamaterials is proposed based on constituent materials. Results show that the decreasing ratioE/ρ(stiffness/mass density) of constituent material can significantly lower the frequency range of single-mode band gap. By merely replacing the constituent material from Al to rubber, the center frequencyfcof single-mode band gap can be reduced nearly 600 times (from 3621 Hz to 6.5 Hz), while the normalized bandwidth Δf/fcand the ratio of bulk modulusBto shear modulusGof pentamode structure keep substantially stable. The nonlinear fitting demonstrates that the relation betweenfcandE/ρsatisfies the logarithmic function. The two-component pentamode structure is designed to further explore the ultra-low-frequency single-mode band gap. The effects of thick-end diameterDof double-cone, diameterD0and material type of additional sphere, on single-mode band gap of two-component system are analyzed. This work is attractive for several ∼Hz acoustic/elastic wave regulations using pentamode metamaterials.

13.
J Neuroinflammation ; 18(1): 49, 2021 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-33602234

RESUMO

BACKGROUND: The integrin VLA-4 (α4ß1) plays an important role in leukocyte trafficking. This study investigated the efficacy of a novel topical α4ß1 integrin inhibitor (GW559090, GW) in a mouse model for non-infectious posterior uveitis (experimental autoimmune uveitis; EAU) and its effect on intraocular leukocyte subsets. METHODS: Mice (female; B10.RIII or C57Bl/6; aged 6-8 weeks) were immunized with specific interphotoreceptor retinoid-binding protein (IRBP) peptides to induce EAU. Topically administered GW (3, 10, and 30 mg/ml) were given twice daily either therapeutically once disease was evident, or prophylactically, and compared with vehicle-treated (Veh) and 0.1% dexamethasone-treated (Dex) controls. Mice were sacrificed at peak disease. The retinal T cell subsets were investigated by immunohistochemistry and immunofluorescence staining. The immune cells within the retina, blood, and draining lymph nodes (dLNs) were phenotyped by flow cytometry. The effect of GW559090 on non-adherent, adherent, and migrated CD4+ T cell subsets across a central nervous system (CNS) endothelium was further assayed in vitro and quantitated by flow cytometry. RESULTS: There was a significant reduction in clinical and histological scores in GW10- and Dex-treated groups as compared to controls either administered therapeutically or prophylactically. There were fewer CD45+ leukocytes infiltrating the retinae and vitreous fluids in the treated GW10 group (P < 0.05). Immunofluorescence staining and flow cytometry data identified decreased levels of retinal Th17 cells (P ≤ 0.001) in the GW10-treated eyes, leaving systemic T cell subsets unaffected. In addition, fewer Ly6C+ inflammatory monocyte/macrophages (P = 0.002) and dendritic cells (P = 0.017) crossed the BRB following GW10 treatment. In vitro migration assays confirmed that Th17 cells were selectively suppressed by GW559090 in adhering to endothelial monolayers. CONCLUSIONS: This α4ß1 integrin inhibitor may exert a modulatory effect in EAU progression by selectively blocking Th17 cell migration across the blood-retinal barrier without affecting systemic CD4+ T cell subsets. Local α4ß1 integrin-directed inhibition could be clinically relevant in treating a Th17-dominant form of uveitis.

14.
J Ocul Pharmacol Ther ; 37(4): 241-247, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33524301

RESUMO

Purpose: This study presents clinical features and prognosis after long-term (12-18 months) antitubercular therapy (ATT) in patients with ocular tuberculosis (OTB) in East China, an endemic area of tuberculosis. Methods: This retrospective study reviewed data from OTB patients treated at the Eye and ENT Hospital of Fudan University from 2008 to 2018. All the patients completed a minimum follow-up of 6 months after the cessation of ATT. Results: Sixty-six patients with OTB were studied. The ocular manifestations included retinal vasculitis (51.6%), choroiditis (24.2%), panuveitis (23.2%), intermediate uveitis (7.4%), scleritis (5.3%), anterior uveitis (2.1%), and optic neuropathy (1%). Except for two patients (ATT for 6 months), all other patients (64/66, 96.97%) received ATT for at least 12 months (6 patients for 12 months, 30 patients for 15 months, and 28 patients for 18 months). Treatment in conjunction with oral corticosteroids was used in 48 patients (72.7%). The average initial best-corrected visual acuity (BCVA) was 0.8 ± 0.64 (LogMAR), which improved to 0.31 ± 0.35 (LogMAR) at the last follow-up (P < 0.05). The final BCVA was significantly associated with the initial BCVA and the duration of clinical symptoms. A complete remission of uveitis was achieved in 97% of the patients. Conclusions: This study observed a favorable prognosis with long-term ATT regimens. Patients with better baseline visual acuity and a shorter duration of clinical symptoms before diagnosis had a better prognosis.

15.
Talanta ; 222: 121580, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33167267

RESUMO

Feature detection is a crucial pre-processing step for high-resolution liquid chromatography-mass spectrometry (LC-MS) data analysis. Typical practices based on thresholds or rigid mathematical assumptions can cause ineffective performance in detecting low abundance and non-ideal distributed compounds. We herein introduce a novel feature detection method based on deep learning named SeA-M2Net that considers feature detection as an image-based object detection task. By fully employing raw data directly, and integrating all related factors (e.g., LC elution, charge state, and isotope distribution) with two-dimensional pseudo color images to calculate the probability of the presence of the compound, low abundance compounds can be well preserved and observed. More importantly, SeA-M2Net, with deep multilevel and multiscale structures focuses on compound pattern detection in a learned method instead of assuming a mathematical parametric model. All parameters in SeA-M2Net are learned from data in the training procedure, thus allowing for maximum flexibility of pattern distribution deformation. The algorithm is tested on several LC-MS datasets of multiple biological samples obtained from different instruments with varied experimental settings. We demonstrate the superiority of the new approach in handling complex compound patterns (e.g., low abundance, overlapping regions, LC shifts, and missing values). Our experiments indicate that SeA-M2Net outperforms widely used detection methods in terms of detection accuracy.

16.
Cancers (Basel) ; 12(12)2020 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-33291756

RESUMO

Diagnosis of ovarian cancer is difficult due to the lack of clinical symptoms and effective screening algorithms. In this study, we aim to develop models for ovarian cancer diagnosis by detecting metabolites in urine and plasma samples. Ultra-high-performance liquid chromatography and quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) in positive ion mode was used for metabolome quantification in 235 urine samples and 331 plasma samples. Then, Urine and plasma metabolomic profiles were analyzed by univariate and multivariate statistics. Four groups of samples: normal control, benign, borderline and malignant ovarian tumors were enrolled in this study. A total of 1330 features and 1302 features were detected from urine and plasma samples respectively. Based on two urine putative metabolites, five plasma putative metabolites and five urine putative metabolites, three models for distinguishing normal-ovarian tumors, benign-malignant (borderline + malignant) and borderline-malignant ovarian tumors were developed respectively. The AUC (Area Under Curve) values were 0.987, 0876 and 0.943 in discovery set and 0.984, 0.896 and 0.836 in validation set for three models. Specially, the diagnostic model based on 5 plasma putative metabolites had better early-stage diagnosis performance than CA125 alone. The AUC values of the model were 0.847 and 0.988 in discovery and validation set respectively. Our results showed that normal and ovarian tumors have unique metabolic signature in urine and plasma samples, which shed light on the ovarian cancer diagnosis and classification.

17.
J Ginseng Res ; 44(5): 680-689, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32913397

RESUMO

Background: Peptides have diverse and important physiological roles in plants and are ideal markers for species identification. It is unclear whether there are specific peptides in Panax quinquefolius L. (PQ). The aims of this study were to identify Quinetides, a series of diverse posttranslational modified native peptides of the ribonuclease-like storage protein (ginseng major protein), from PQ to explore novel peptide markers and develop a new method to distinguish PQ from Panax ginseng. Methods: We used different fragmentation modes in the LTQ Orbitrap analysis to identify the enriched Quinetide targets of PQ, and we discovered Quinetide markers of PQ and P. ginseng using ultrahigh-performance liquid chromatography-quadrupole time-of-flight mass spectrometry analysis. These "peptide markers" were validated by simultaneously monitoring Rf and F11 as standard ginsenosides. Results: We discovered 100 Quinetides of PQ with various post-translational modifications (PTMs), including a series of glycopeptides, all of which originated from the protein ginseng major protein. We effectively distinguished PQ from P. ginseng using new "peptide markers." Four unique peptides (Quinetides TP6 and TP7 as markers of PQ and Quinetides TP8 and TP9 as markers of P. ginseng) and their associated glycosylation products were discovered in PQ and P. ginseng. Conclusion: We provide specific information on PQ peptides and propose the clinical application of peptide markers to distinguish PQ from P. ginseng.

18.
Front Pharmacol ; 11: 949, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32848721

RESUMO

Osteoarthritis (OA), as one of the top 10 causes of physical disability, is characterized by inflammation of the synovial membrane and progressive destruction of the articular cartilage. Cinnamic aldehyde (CA), an α,ß-unsaturated aldehyde extracted from the traditional Chinese herbal medicine cinnamon (Cinnamomum verum J.Presl), has been reported to have anti-inflammatory, antioxidant, and anticancer properties. However, the anti-inflammatory effect of CA on OA remains unclear. The purpose of the present study was to investigate the effects of CA on inflammation, and cartilage degeneration in OA. A CCK-8 assay was performed to assess the potential toxicity of CA on cultured human OA chondrocytes. Following treatment with lipopolysaccharide (LPS) and CA, the expression of proinflammatory cytokines, including interleukin (IL)-1ß, IL-6, and tumor necrosis factor-alfa (TNF-α), was evaluated using quantitative real-time polymerase chain reaction (RT-qPCR) analysis, enzyme-linked immunosorbent assay, and Western blotting (WB). The production of matrix metalloproteinase-13 (MMP-13) and a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS-5) was also examined using RT-qPCR and WB. Furthermore, to investigate the potential anti-inflammatory mechanism of CA, biomarkers of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway (p65, IKB-α) were detected using WB. The results demonstrated that CA significantly inhibited the expressions of IL-1ß, IL-6, TNF-α, MMP-13, and ADAMTS-5 in LPS-induced OA chondrocytes. CA dramatically suppressed LPS-stimulated NF-κB activation. Collectively, these results suggest that CA treatment may effectively prevent OA.

19.
Zhongguo Gu Shang ; 33(7): 643-8, 2020 Jul 25.
Artigo em Chinês | MEDLINE | ID: mdl-32700488

RESUMO

OBJECTIVE: To observe effects of Tongluo Zhitong (, TLZT) gel preparation on p53, miR-502-5p, NF-κBp65 in synovial tissue of knee osteoarthritis (KOA), and to explore mechanism of TLZT gel preparation in treating KOA. METHODS: Thirthy-six Wistar rats aged 8 weeks and weighed 200 to 220 g (meaned 208 g) were randomly divided into normal group, model group and traditional Chinese medicine (TCM) group, 12 rats in each group. KOA model was established by modified Hulth method. After 4 weeks of modeling, TCM group treated with TLZT gel preparation for external use, 3 times daily for 2 weeks;normal group and model group were fed normally without intervention. After treatment, morphological changes of specimens in each group were observed, changes of miR-502-5p in synovial tissue were detected by qPCR, and contents of p53, NF-κBp65, IL-1ß, TNF-α, MMP-13 in synovial tissue were detected by qPCR and Western Blot respectively. RESULTS: (1)Morphological observation of specimens showed that the articular cartilage in model group was hyaline and uneven, the synovial membranes were hypertrophic and proliferative with a large number of inflammatory cells infiltrating, the joint fluid was thicker in texture;the articular cartilage in TCM group was more transparent and smooth, synovial hyperplasia was mild with a small amount of inflammatory cell infiltration, the texture of articular fluid was clear and sparse. (2) Compared with normal group, content of miR-502-5p of synovial tissue in model and TCM group were increased, mRNA and expression of p53 decreased, expression of NF-κBp65, IL-1ß, TNF-α, MMP-13 increased. (3)Compared with model group, content of miR-502-5p in synovial tissue of TCM group decreased (P<0.05), mRNA and protein expression of p53 increased (P<0.05), mRNA and protein expression of NF-κBp65, IL-1ß, TNF-α, MMP-13 decreased (P<0.05). CONCLUSION: Expression of p53, miR-502 -5p, NF -κBp65 in synovial tissue is closely related to synovial hyperplasia and inflammatory reaction, TLZT gel preparation may reduce proliferation and inflammatory reaction of KOA synovium by regulating the expression of p53, miR- 502-5p, NF-κBp65 in synovial tissues.


Assuntos
MicroRNAs , Osteoartrite do Joelho , Animais , Medicamentos de Ervas Chinesas , Ratos , Ratos Wistar , Membrana Sinovial , Proteína Supressora de Tumor p53
20.
Eur J Immunol ; 50(12): 1941-1951, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32652562

RESUMO

Immunopathogenic roles for both Th1 (CD4+ IFN-γ+ ) and Th17 (CD4+ IL-17A+ ) cells have been demonstrated in experimental autoimmune uveitis (EAU). However, the role for Th17/Th1 (CD4+ T cells co-expressing IFN-γ and IL-17A) cells in EAU is not yet understood. Using interphotoreceptor retinoid-binding protein peptide-induced EAU in mice, we found increased levels of Th17/Th1 cells in EAU retinae (mean 9.6 ± 4.2%) and draining LNs (mean 8.4 ± 3.9%; p = 0.01) relative to controls. Topical dexamethasone treatment effectively reduced EAU severity and decreased retinal Th1 cells (p = 0.01), but had no impact on retinal Th17/Th1 or Th17 cells compared to saline controls. Using in vitro migration assays with mouse CNS endothelium, we demonstrated that Th17/Th1 cells were significantly increased within the migrated population relative to controls (mean 15.6 ± 9.5% vs. 1.9 ± 1.5%; p = 0.01). Chemokine receptor profiles of Th17/Th1 cells (CXCR3 and CCR6) did not change throughout the transendothelial migration process and were unaffected by dexamethasone treatment. These findings support a role for Th17/Th1 cells in EAU and their resistance to steroid inhibition suggests the importance of targeting both Th17 and Th17/Th1 cells for improving therapy.


Assuntos
Doenças Autoimunes/imunologia , Movimento Celular/imunologia , Interferon gama/imunologia , Interleucina-17/imunologia , Células Th1/imunologia , Células Th17/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL
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