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1.
Biochim Biophys Acta Mol Basis Dis ; 1866(1): 165554, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31513833

RESUMO

Activation of interferon (IFN)-I signaling in B cells contributes to the pathogenesis of systemic lupus erythematosus (SLE). Recent studies have shown that myeloid-derived suppressor cells (MDSCs) significantly expand in SLE patients and lupus-prone MRL/lpr mice and contribute to the pathogenesis of SLE. However, the role of SLE-derived MDSCs in regulating IFN-I signaling activation of B cells remains unknown. Here, we demonstrate that expansions of MDSCs, including granulocyte (G)-MDSCs and monocytic (M)-MDSCs, during the progression of SLE were correlated with the IFN-I signature of B cells. Interestingly, G-MDSCs from MRL/lpr mice, but not M-MDSCs, could significantly promote IFN-I signaling activation of B cells and contribute to the pathogenesis of SLE. Mechanistically, we identified that the long non-coding RNA NEAT1 was over-expressed in G-MDSCs from MRL/lpr mice and could induce the promotion of G-MDSCs on IFN-I signaling activation of B cells through B cell-activating factor (BAFF) secretion. Importantly, NEAT1 deficiency significantly attenuated the lupus symptoms in pristane-induced lupus mice. In addition, there was a positive correlation between NEAT1 and BAFF with the IFN signature in SLE patients. In conclusion, G-MDSCs may contribute to the IFN signature in SLE B cells through the NEAT1-BAFF axis, highlighting G-MDSCs as a potential therapeutic target to treat SLE.

2.
Microbes Infect ; 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31678658

RESUMO

CD4+/CD8+ T cells play a major role in conferring immune protection against tuberculosis (TB), but it remains unknown how the immune responses of CD4+/CD8+ T cells exactly correlate with the clinical variables and disease statuses during anti-TB chemotherapy. To address this, several major immune parameters of CD4+/CD8+ T cells in peripheral blood derived from pulmonary TB patients and healthy volunteers were evaluated. We observed that active TB infection induced lower CD3+ T cell and CD4+ T cell levels but higher CD8+T cell levels, while anti-TB chemotherapy reversed these effects. Also, anti-TB treatment induced enhanced production of IL-2 and IFN-γ but reduced expression of IL-10 and IL-6. Moreover, the dynamic changes of CD3, CD4, and CD8 levels did not show a significant association with sputum smear positivity. However, the frequencies of IL-2+CD4+ or IL-10 + CD4+ T effector subpopulation or IL-1ß production in peripheral blood showed significant difference between patients positive for sputum smear and patients negative for sputum smear after anti-TB treatment. These findings implicated that recovery of Th1/CD8+T cell effector levels might be critical immunological events in pulmonary TB patients after treatment and further suggested the importance of these immunological parameters as potential biomarkers for prediction of TB progress and prognosis.

3.
Front Immunol ; 10: 1824, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31428103

RESUMO

Macrophages play a critical role in the pathogenesis of endotoxin shock by producing excessive amounts of pro-inflammatory cytokines. A pan-caspase inhibitor, zVAD, can be used to induce necroptosis under certain stimuli. The role of zVAD in both regulating the survival and activation of macrophages, and the pathogenesis of endotoxin shock remains not entirely clear. Here, we found that treatment of mice with zVAD could significantly reduce mortality and alleviate disease after lipopolysaccharide (LPS) challenge. Notably, in LPS-challenged mice, treatment with zVAD could also reduce the percentage of peritoneal macrophages by promoting necroptosis and inhibiting pro-inflammatory responses in macrophages. In vitro studies showed that pretreatment with zVAD promoted LPS-induced nitric oxide-mediated necroptosis of bone marrow-derived macrophages (BMDMs), leading to reduced pro-inflammatory cytokine secretion. Interestingly, zVAD treatment promoted the accumulation of myeloid-derived suppressor cells (MDSCs) in a mouse model of endotoxin shock, and this process inhibited LPS-induced pro-inflammatory responses in macrophages. Based on these findings, we conclude that treatment with zVAD alleviates LPS-induced endotoxic shock by inducing macrophage necroptosis and promoting MDSC-mediated inhibition of macrophage activation. Thus, this study provides insights into the effects of zVAD treatment in inflammatory diseases, especially endotoxic shock.

4.
Pharm Biol ; 57(1): 412-423, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31230505

RESUMO

Context: Shikonins, a series of natural occurring naphthoquinones extracted from Arnebia euchroma (Royle) Jonst. (Boraginaceae), have antitumor activities and low toxicity. Objective: To illuminate potential activity and mechanism of shikonins against colorectal cancer (CRC). Materials and methods: Five shikonins were isolated from A. euchroma, and elucidated by extensive spectroscopic analysis. Anti-proliferative activities of shikonins (0-100 µg/mL) on human colorectal cells were evaluated by MTT and CCK-8 for 24 or 48 h. Cell apoptosis and cycle distribution were examined by FCM analysis. The expression of PI3K/Akt/mTOR pathway mRNAs and proteins was analysed by RT-PCR and Western blot, respectively. Cell viability, cell apoptosis, cell cycle and protein expression were measured, when co-treated with PI3K/Akt/mTOR pathway inhibitors. The in vivo activity of deoxyshikonin was evaluated using xenograft tumour model. Results: Deoxyshikonin and another four shikonins were isolated and identified. Deoxyshikonin exhibited anti-proliferative activity with IC50 of 10.97 µM against HT29 cells. Moreover, the percentage of early apoptotic cells and G0/G1 cells increased from 1 to 29% and 44 to 67% with 0-50 µg/mL deoxyshikonin, respectively. Deoxyshikonin also down-regulated the expression of PI3K, p-PI3K, Akt, p-Akt308 and mTOR proteins in HT29 and DLD-1 cells. Moreover, LY294002, NVP-BEZ235 and MK-2206 can make deoxyshikonin more cell proliferation inhibited, cell cycle arrested at G0/G1 and apoptosis promoted. In vivo study, the weight of tumour tissues at deoxyshikonin groups was significantly reduced compared with the control group, and PI3K, p-PI3K, Akt, p-Akt308 and mTOR expression was decreased. Discussion and conclusions: We can conclude that deoxyshikonin isolated from Arnebia euchroma inhibited CRC through the PI3K/Akt/mTOR pathway.

5.
Immunology ; 157(3): 257-267, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31120548

RESUMO

Asthma is a chronic inflammatory disease that involves a variety of cytokines and cells. Interleukin-16 (IL-16) is highly expressed during allergic airway inflammation and is involved in its development. However, its specific mechanism of action remains unclear. In the present study, we used an animal model of ovalbumin (OVA)-induced allergic asthma with mice harboring an IL-16 gene deletion to investigate the role of this cytokine in asthma, in addition to its underlying mechanism. Increased IL-16 expression was observed during OVA-induced asthma in C57BL/6J mice. However, when OVA was used to induce asthma in IL-16-/- mice, a diminished inflammatory reaction, decreased bronchoalveolar lavage fluid (BALF) eosinophil numbers, and the suppression of OVA-specific IgE levels in the serum and BALF were observed. The results also demonstrated decreased levels of T helper type 2 (Th2) and Th17 cytokines upon OVA-induced asthma in IL-16-/- mice. Hence, we confirmed that IL-16 enhances the lung allergic inflammatory response and suggest a mechanism possibly associated with the up-regulation of IgE and the promotion of Th2 and Th17 cytokine production. This work explored the mechanism underlying the regulation of IL-16 in asthma and provides a new target for the clinical treatment of asthma.

6.
Zhongguo Zhong Yao Za Zhi ; 44(7): 1450-1456, 2019 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-31090304

RESUMO

The aim of this paper was to investigate the effect of Shaoyao Tang on ulcerative colitis(UC) in rats via regulation of TLR4/NF-κB signal pathway. A total of 56 Wistar rats were randomly divided into 6 groups: normal control group(double distilled water), model group(double distilled water), mesalazine group(10 mL·kg~(-1)), high dose, middle dose and low dose Shaoyao Tang groups(2.4, 1.2, and 0.6 g·mL~(-1)). After UC rat models were established by 2, 4-dinitrochlorobenzene(DNCB)/ethanol enema, the rats received double distilled water or corresponding drugs twice a day for 7 days. After the treatment cycle, the general performance and disease activity index(DAI) of rats were observed on the next day. Then the rats were sacrificed. The length of colon was measured. Macroscopic and histological score of colon were evaluated. Histopathological changes of colon were observed by HE staining. Ultraviolet spectrophotometry detection was used to detect the content of myeloperoxidase(MPO) in blood and colon tissues. The levels of P-selectin, macrophage migration inhibitory factor(MIF) and thromboxane B_2(TXB_2) in blood and colon tissues were determined by ELISA. Immunohistochemistry and Western blot analysis were performed to detect the protein expressions of TLR4 and NF-κB in colon tissues. The results showed that as compared with the model group, Shaoyao Tang of different doses improved the general performance of UC rats. Moreover, high-dose Shaoyao Tang group showed the most obvious effect in scoring of disease activity index(P<0.001); both medium and high doses of Shaoyao Tang significantly inhibited the colon shortening and pathological injury, with significantly decreased expression levels of MPO, P-selectin, MIF and TXB_(2 )in serum and colon tissues of UC rats(P<0.001). Immunohistochemistry and Western blot assay showed that the levels of TLR4 and NF-κB protein expression in the colon tissues of Shaoyao Tang high-dose group were remarkably lower than that in the model group(P<0.001). This study shows that Shaoyao Tang has protective and repairing effects on UC, and its possible mechanism is achieved probably by regulating the TLR4/NF-κB pathway and inhibiting the expressions of MPO, P-selectin, MIF and TXB_2.


Assuntos
Colite Ulcerativa/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , NF-kappa B/metabolismo , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Animais , Colo , Distribuição Aleatória , Ratos , Ratos Wistar
7.
Front Immunol ; 10: 215, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30809230

RESUMO

Dysregulation of macrophage has been demonstrated to contribute to aberrant immune responses and inflammatory diseases. CD11b, expressed on macrophages, plays a critical role in regulating pathogen recognition, phagocytosis, and cell survival. In the present study, we explored the effect of leukadherin-1 (LA1), an agonist of CD11b, on regulating LPS-induced pro-inflammatory response in macrophages and endotoxic shock. Intriguingly, we found that LA1 could significantly reduce mortalities of mice and alleviated pathological injury of liver and lung in endotoxic shock. In vivo studies showed that LA1-induced activation of CD11b significantly inhibited the LPS-induced pro-inflammatory response in macrophages of mice. Moreover, LA1-induced activation of CD11b significantly inhibited LPS/IFN-γ-induced pro-inflammatory response in macrophages by inhibiting MAPKs and NF-κB signaling pathways in vitro. Furthermore, the mice injected with LA1-treated BMDMs showed fewer pathological lesions than those injected with vehicle-treated BMDMs in endotoxic shock. In addition, we found that activation of TLR4 by LPS could endocytose CD11b and activation of CD11b by LA1 could endocytose TLR4 in vitro and in vivo, subsequently blocking the binding of LPS with TLR4. Based on these findings, we concluded that LA1-induced activation of CD11b negatively regulates LPS-induced pro-inflammatory response in macrophages and subsequently protects mice from endotoxin shock by partially blocking LPS-TLR4 interaction. Our study provides a new insight into the role of CD11b in the pathogenesis of inflammatory diseases.

8.
Biochim Biophys Acta Mol Basis Dis ; 1865(3): 535-546, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30557700

RESUMO

Myeloid-derived suppressor cells (MDSCs) play an immunosuppressive role in the pathogenesis of inflammatory diseases. CD180, a TLR-like protein, can regulate the proliferation and activation of immune cells. However, the roles of CD180 in regulating the accumulation and function of MDSCs have not been investigated. Here, we found that, compared with non-treated controls, the expression of CD180 was significantly elevated in MDSCs, especially granulocytic MDSCs (G-MDSCs), from mice challenged with lipopolysaccharide (LPS). Ligation of CD180 by the anti-CD180 antibody not only blocked the expansion of MDSCs by preventing the phosphorylation of signal transducer and activator of transcription 3 (STAT3), but also reduced the immunosuppressive activity of MDSCs on M1 macrophage polarization through inhibition of Arg-1 expression in vitro. In vivo studies showed that injection of anti-CD180 antibody significantly aggravated pathological lesions in mice challenged with LPS. Furthermore, injection of anti-CD180 antibody inhibited the accumulation of G-MDSCs in mice challenged with LPS and reduced the immunosuppressive activity of G-MDSCs on M1 macrophage polarization. Based on these findings, we conclude that ligation of CD180 contributes to the pathogenesis of endotoxic shock by inhibiting the accumulation and immunosuppressive activity of G-MDSCs, thus providing insight into the function of CD180 in inflammatory diseases.


Assuntos
Anticorpos Monoclonais/farmacologia , Antígenos CD/imunologia , Células Supressoras Mieloides/imunologia , Fator de Transcrição STAT3/fisiologia , Choque Séptico/induzido quimicamente , Choque Séptico/imunologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Feminino , Inflamação/induzido quimicamente , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Células Supressoras Mieloides/citologia , Células Supressoras Mieloides/efeitos dos fármacos , Ligação Proteica , Choque Séptico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia
9.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 34(11): 961-968, 2018 Nov.
Artigo em Chinês | MEDLINE | ID: mdl-30591103

RESUMO

Objective To study the effect of CD11b agonist leukadherin-1 (LA1) on the aggregation and immunosuppressive function of myeloid-derived suppressor cells (MDSCs) and its therapeutic effect on the condition of endotoxic shock mice. Methods The percentages of MDSCs , granulocytic myeloid-derived suppressor cells(G-MDSCs)and monocytic myeloid-derived suppressor cells(M-MDSCs)in spleen were detected by flow cytometry, after C57BL/6 female mice were injected of LA1 to activate through abdominal cavity for 12 hours and 48 hours. MDSCs were induced from the femur and tibia of C57BL/6 female mice in vitro. The expression levels of immunosuppressive related factors, such as interleukin 10 (IL-10), NADPH oxidase 1 (NOX1) and inducible nitric oxide synthase (iNOS) , were detected by real time quantitative PCR. C57BL/6 female mice were randomly divided into PBS group, LA1 group, PBS combined LPS group and LA1 combined LPS group. Flow cytometry was utilized to detect the ratio changes of MDSCs, G-MDSCs and M-MDSCs as well as the expression of CD86 and CD40 in macrophage, hematoxylin-eosin staining of lung and liver was utilized to detect the pathological injury, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling(TUNEL)was used to detect the apoptosis of pneumonocyte and hepatocyte and mortality analysis was reflected the severity of the disease. Based on the above indicators, we analyzed the effects of LA1 on the aggregation of MDSCs and the condition of mice in endotoxic shock. Results The ratio of MDSCs was increased by LA1 treatment for 12 and 48 hours. Further analysis of the proportions of G-MDSCs showed that LA1 treatment for 12 hours increased the proportions of G-MDSCs compared with the control group. In vitro, mRNA levels of IL-10, NOX1 and iNOS were increased after LA1 treatment in MDSCs. In vivo experiments, compared with the PBS combined LPS group, the proportions of MDSCs and G-MDSCs in LA1 combined LPS group were increased, the injuries of liver and lung were alleviated, the mortalities were reduced, and the activations of macrophage were decreased. Conclusion The activation of CD11b by LA1 alleviates endotoxin shock by promoting the aggregation of MDSCs and the expression of immunosuppressive related factors.


Assuntos
Benzoatos/farmacologia , Antígeno CD11b/agonistas , Células Supressoras Mieloides/citologia , Choque Séptico/tratamento farmacológico , Tioidantoínas/farmacologia , Animais , Feminino , Interleucina-10/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NADPH Oxidase 1/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Distribuição Aleatória , Baço/citologia
10.
Fitoterapia ; 131: 236-244, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30414877

RESUMO

Six previously undescribed naturally occurring meroterpenoids (2, 5-9) together with seven known meroterpenoids (1, 3, 4, 10-13) were isolated from the root plant of Arnebia euchroma. Their structures and absolute configurations were determined by extensive 1D (1H NMR, 13C NMR) and 2D NMR (1H1H COSY, DEPT, HMQC, HMBC, NOESY) spectroscopic methods, spectroscopy high resolution mass spectrometry, as well as DFT and MM2 force-field calculations. Meroterpenoids 1-13 were evaluated for their cytotoxicities against human liver cancer cell lines SMMC-7721, HepG2, QGY-7703 and HepG2/ADM. Meroterpenoid 5 exhibited the most potent activity with IC50 values of 6.40 ±â€¯0.51, 3.86 ±â€¯0.28, 3.43 ±â€¯0.27 and 11.31 ±â€¯0.67 µM, respectively. Meroterpenoid 4 exhibited significant growth inhibitory effects against HepG2/ADM with IC50 at 18.77 ±â€¯1.23 µM, and meroterpenoid 8 with IC50 at 5.41 ±â€¯0.51 and 6.18 ±â€¯0.47 µM against HepG2 and QGY-7703, respectively. These were more potent than the positive drug, Cisplatin.

11.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 34(8): 695-701, 2018 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-30384867

RESUMO

Objective To investigate the role of interleukin-16 (IL-16) in the development of inflammatory bowel disease (IBD) and clarify its regulatory mechanism involved in the pathogenesis of IBD. Methods Seven-week-old wild-type C57BL/6 (WT) and IL-16 knockout (IL-16-/-) female mice were divided into WT control group, WT dextran sulfate sodium (DSS) treatment group, IL-16-/- control group and IL-16-/- DSS treatment group. The DSS model groups were given the water with 25 g/L DSS for 7 days to establish the IBD models, while the control groups were given the normal water. During the modeling period, the body mass of mice was recorded to calculate the body mass curve. After 7 days, the whole colon of the mice was dissected and the level of IL-16 mRNA in the colon tissue was detected by real-time PCR. The level of IL-16 protein in the colon tissue was detected by ELISA. The expression and localization of IL-16 in the colon tissue were observed by immunofluorescence technique. HE staining was used to detect colonic pathological injury in mice. TUNEL assay was used to detect cell apoptosis of the colon tissue. Flow cytometry was used to detect the number and polarization of macrophages in peritoneal cells (F4/80, CD86). Immunohistochemical staining was used to detect the distribution of macrophages in the colon tissues. Real-time PCR was used to detect IL-6 and IL-12 mRNA levels in the colon tissue, and IL-6 and IL-12 protein levels were detected by ELISA. Results DSS induced high expression of IL-16 in the colon tissue. Compared with WT DSS treatment group, IL-16-/- DSS treatment group showed less changes in body mass, less colon tissue damage, and markedly lower percents of apoptotic cells in the peritoneal or colonic tissues of IL-16-/- mice. What's more, the number of macrophages, the polarization level of M1 macrophages, and the levels of the iconic inflammatory factors IL-6 and IL-12 significantly decreased in IL-16-/- DSS treatment group compared with WT DSS treatment group. Conclusion IL-16 can aggravate DSS-induced IBD by promoting the polarization of M1 macrophages.

12.
Zhongguo Zhong Yao Za Zhi ; 43(11): 2358-2364, 2018 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-29945391

RESUMO

To investigate the inhibitory effect of isobutyrylshikonin on the growth of human colon carcinoma cells in vitro and its effect on the PI3K/Akt/m-TOR pathway. MTT assay was used to detect the inhibitory effect of different concentrations (0, 6.25, 12.5, 25, 50, 100 mg·L⁻¹) of isobutyrylshikonin on the proliferation of human colon carcinoma cell HT29 at 24, 48 h. CCK-8 method was used to detect the inhibitory effect of isobutyrylshikonin on HT29, HCT116, DLD-1 and Caco-2 at 48 h. AnnexinV/propidium iodide staining was applied in detecting the apoptoticrate of HT29 cells treated with different concentrations of isobutyrylshikonin at 24 h and 48 h. Cycletest plus DNA was employed to analyze HT29 apoptosis and cell cycle after 48 h treatment with isobutyrylshikonin at different concentrations. Western blot and RT-PCR assay were used to examine the protein and mRNA expressions of PI3K, p-PI3K, Akt, p-Akt and m-TOR. The results showed that isobutyrylshikonin inhibited the proliferation of different human colon carcinoma cells, and the inhibition rate was in a dose-dependent manner. Isobutyrylshikonin induced apoptosis mainly in the early stage and blocked cells in the G0/G1 or G2/M phase. Isobutyrylshikonin reduced the protein expressions of PI3K, p-PI3K, Akt, p-Akt, m-TOR and the mRNA expressions of PI3K, Akt, m-TOR in a dose-dependent manner. Isobutyrylshikonin can significantly inhibit the proliferation, induce the early apoptosis and change the cycle distribution in colon carcinoma cells.This biological effect may be correlated with the inhibition of PI3K/AKT/m-TOR pathway.

13.
Dig Dis Sci ; 62(3): 639-651, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28035546

RESUMO

PURPOSE: The CD8+CD28+/CD8+CD28- T lymphocyte balance is vital for human ulcerative colitis (UC) but has not been defined in experimental colitis. This investigation will try to identify the changes that occur in the CD8+CD28+/CD8+CD28- T lymphocyte balance during the progression of trinitrobenzenesulfonic acid (TNBS)-induced colitis in rats. METHODS: The frequencies of blood CD8+CD28+ and CD8+CD28- T lymphocytes were detected in the rats belonging to the normal, model, and treated groups on five days using flow cytometry. The treated rats were administered with mesalazine and were euthanized after a 14-day treatment, as were the normal and model rats. The sensitivity and specificity of the CD8+CD28+/CD8+CD28- T lymphocyte balance in diagnosing early colitis were analyzed by receiver operating characteristics (ROC) curves. The frequencies of CD8+CD28+ and CD8+CD28- T lymphocytes in the colon tissue were tested via immunofluorescence. ELISA was used to measure the levels of the cytokines. Immunohistochemistry and Western blotting were used to detect the colonic expression of JAK3, STAT6, NFATc2, and GATA3. RESULTS: We found that the ratio of CD8+CD28+/CD8+CD28- T lymphocytes decreased, as did the level of interleukin-7, but not IL-12p40, IL-13, or IL-15, in the blood; however, the ratio increased along with JAK3, STAT6, NFATc2, and GATA3 in the colon of the rats with colitis. The changes were effectively reversed through the administration of mesalazine for 13 days. Surprisingly, the balance in the blood could sensitively distinguish rats with early colitis from normal rats. CONCLUSION: These data show that increase in CD8+CD28+ T cells in blood and decrease in CD8+CD28- T cells in colon are associated with experimental colitis.


Assuntos
Antígenos CD , Colite , Mesalamina/administração & dosagem , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Antígenos CD/análise , Antígenos CD/metabolismo , Colite/diagnóstico , Colite/etiologia , Colite/imunologia , Modelos Animais de Doenças , Progressão da Doença , Diagnóstico Precoce , Interleucinas/análise , Interleucinas/metabolismo , Prognóstico , Curva ROC , Ratos , Sensibilidade e Especificidade , Ácido Trinitrobenzenossulfônico/farmacologia
14.
J Phys Chem B ; 121(3): 508-517, 2017 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-28030949

RESUMO

pH is highly regulated in mammalian central nervous systems. Neuronal calcium sensor-1 (NCS-1) can interact with numerous target proteins. Compared to that in the NCS-1 protein of Caenorhabditis elegans, evolution has avoided the placement of histidine residues at positions 102 and 83 in the NCS-1 protein of humans and Xenopus laevis, possibly to decrease the conformational sensitivity to pH gradients in synaptic processes. We used all-atom molecular dynamics simulations to investigate the effects of amino acid substitutions between species on human NCS-1 by substituting Arg102 and Ser83 for histidine at neutral (R102H and S83H) and acidic pHs (R102Hp and S83Hp). Our cumulative 5 µs simulations revealed that the R102H mutation slightly increases the structural flexibility of loop L2 and the R102Hp mutation decreases protein stability. Community network analysis illustrates that the R102H and S83H mutations weaken the interdomain and strengthen the intradomain communications. Secondary structure contents in the S83H and S83Hp mutants are similar to those in the wild type, whereas the global structural stabilities and salt-bridge probabilities decrease. This study highlights the conformational dynamics effects of the R102H and S83H mutations on the local structural flexibility and global stability of NCS-1, whereas protonated histidine decreases the stability of NCS-1. Thus, histidines at positions 102 and 83 may not be compatible with the function of NCS-1 whether in the neutral or protonated state.


Assuntos
Histidina/química , Histidina/metabolismo , Simulação de Dinâmica Molecular , Proteínas Sensoras de Cálcio Neuronal/química , Proteínas Sensoras de Cálcio Neuronal/metabolismo , Neuropeptídeos/química , Neuropeptídeos/metabolismo , Substituição de Aminoácidos , Histidina/genética , Humanos , Concentração de Íons de Hidrogênio , Mutação , Proteínas Sensoras de Cálcio Neuronal/genética , Neuropeptídeos/genética
15.
Cancer Biol Ther ; 17(11): 1149-1157, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27686621

RESUMO

This study aimed to understand the exact function and potential mechanism of miR-4500 in colorectal cancer (CRC). In this study, the expression of miR-4500 was decreased in both CRC cells and tissues, and downregulated miR-4500 indicated advanced tumor stage and poor survival. By bisulfite sequencing analysis, we found that the CpG island in the promoter region of miR-4500 was hypermethylated in CRC cells and tissues compared with normal control cells and non-tumor tissues, respectively. Functionally, gain- and loss-of-function analyses indicated the tumor suppressor role of miR-4500: it suppressed cell proliferation, cell cycle progression, migration, and invasion. Predictive algorithms and experimental analyses identified HMGA2 as a direct target of miR-4500. Reintroducing HMGA2 impaired the inhibitory effects of miR-4500 on cell growth and motility. Clinically, higher HMGA2 protein expression in CRC tissues was associated with advanced tumor stage and poor survival. An inverse correlation was found between miR-4500 levels and HMGA2 protein expression. Taken together, this study provides the first evidence that miR-4500 functions as a novel tumor suppressor in the miR-4500/HMGA2 axis in colorectal carcinogenesis, and restoring miR-4500 expression might represent a promising therapeutic strategy for CRC.


Assuntos
Neoplasias Colorretais/genética , Proteína HMGA2/genética , MicroRNAs/genética , Animais , Proliferação de Células/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Metilação de DNA , Regulação para Baixo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Células HCT116 , Proteína HMGA2/metabolismo , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , MicroRNAs/biossíntese , Prognóstico
16.
ACS Appl Mater Interfaces ; 8(35): 23133-42, 2016 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-27538462

RESUMO

The construction of a p-n heterojunction is an efficient strategy to resolve the limited light absorption and serious charge-carrier recombination in semiconductors and enhance the photocatalytic activity. However, the promotion effect is greatly limited by poor interfacial charge transfer efficiency as well as reduced redox ability of charge carriers. In this work, we demonstrate that the embedding of metal Pd into the interface between n-type C3N4 and p-type Cu2O can further enhance the interfacial charge transfer and increase the redox ability of charge carriers through the design of the C3N4-Pd-Cu2O stack nanostructure. The embedded Pd nanocubes in the stack structure not only trap the charge carriers from the semiconductors in promoting the electron-hole separation but also act as a Z-scheme "bridge" in keeping the strong reduction/oxidation ability of the electrons/holes for surface reactions. Furthermore, Pd nanocubes also increase the bonding strength between the two semiconductors. Enabled by this unique design, the hydrogen evolution achieved is dramatically higher than that of its counterpart C3N4-Cu2O structure without Pd embedding. The apparent quantum efficiency (AQE) is 0.9% at 420 nm for the designed C3N4-Pd-Cu2O. This work highlights the rational interfacial design of heterojunctions for enhanced photocatalytic performance.

17.
J Phys Chem B ; 120(14): 3551-9, 2016 Apr 14.
Artigo em Inglês | MEDLINE | ID: mdl-27007011

RESUMO

Neuronal calcium sensor-1 (NCS-1) protein has orthologues from Saccharomyces cerevisiae to human with highly conserved amino acid sequences. NCS-1 is an important factor controlling the animal's response to temperature change. This leads us to investigate the temperature effects on the conformational dynamics of human NCS-1 at 310 and 316 K by all-atom molecular dynamics (MD) simulations and dynamic community network analysis. Four independent 500 ns MD simulations show that secondary structure content at 316 K is similar to that at 310 K, whereas the global protein structure is expanded. Loop 3 (L3) adopts an extended state occuping the hydrophobic crevice, and the number of suboptimal communication paths between residue D176 and V190 is reduced at 316 K. The dynamic community network analysis suggests that the interdomain correlation is weakened, and the intradomain coupling is strengthened at 316 K. The elevated temperature reduces the number of the salt bridges, especially in C-domain. This study suggests that the elevated temperature affects the conformational dynamics of human NCS-1 protein. Comparison of the structural dynamics of R102Q mutant and Δ176-190 truncated NCS-1 suggests that the structural and dynamical response of NCS-1 protein to elevated temperature may be one of its intrinsic functional properties.


Assuntos
Simulação de Dinâmica Molecular , Proteínas Sensoras de Cálcio Neuronal/química , Neuropeptídeos/química , Temperatura Ambiente , Dissulfetos/química , Humanos , Proteínas Sensoras de Cálcio Neuronal/metabolismo , Neuropeptídeos/metabolismo , Domínios Proteicos , Estrutura Secundária de Proteína
18.
Mol Med Rep ; 13(4): 3525-32, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26935771

RESUMO

The aim of the present study was to investigate the mechanism underlying the antitumor effects of ent-11α-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F) in colorectal cancer (CRC). 5F was isolated and used to treat C26 murine colon carcinoma cells, a xenograft tumor mouse model (induced by C26 cells) and a CRC mouse model [induced by 1,2-dimethylhydrazine (DMH)/dextran sodium sulfate (DSS)]. C26 cell growth was inhibited by 5F in a dose- and time-dependent manner in vitro. In addition, 5F induced cell apoptosis and cell cycle arrest in the G2 phase, increased the activity of caspase-3 and caspase-9, but did not affect the activity of cascase­8, suggesting that 5F induced apoptosis via activation of the mitochondrial signaling pathway rather than the death­receptor signaling pathway. Furthermore, treatment of C26 cells with 5F resulted in upregulation of cyclin­dependent kinase inhibitor 1A (p21, Cip1), Bcl­2­associated X protein, nuclear factor of κ light polypeptide gene enhancer in B­cells inhibitor, α and downregulation of B­cell lymphoma 2, nuclear factor κ­light­chain enhancer of activated B cells and survivin. In vivo animal models demonstrated that 5F treatment protected mice from carcinogenesis induced by DMH/DSS and markedly decreased the xenograft tumor weight with minimal side effects. Therefore, 5F may have potential as an anti-CRC therapeutic agent for use in the clinical setting.


Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Diterpenos de Caurano/farmacologia , Animais , Antineoplásicos Fitogênicos/farmacologia , Antineoplásicos Fitogênicos/uso terapêutico , Caspase 3/metabolismo , Linhagem Celular Tumoral , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Diterpenos de Caurano/uso terapêutico , Feminino , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos ICR , Inibidor de NF-kappaB alfa/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transplante Heterólogo , Regulação para Cima/efeitos dos fármacos , Proteína X Associada a bcl-2/metabolismo
19.
ACS Chem Neurosci ; 7(3): 286-96, 2016 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-26815332

RESUMO

The aggregation of the copper-zinc superoxide dismutase (SOD1) protein is linked to familial amyotrophic lateral sclerosis, a progressive neurodegenerative disease. A recent experimental study has shown that the (147)GVIGIAQ(153) SOD1 C-terminal segment not only forms amyloid fibrils in isolation but also accelerates the aggregation of full-length SOD1, while substitution of isoleucine at site 149 by proline blocks its fibril formation. Amyloid formation is a nucleation-polymerization process. In this study, we investigated the oligomerization and the nucleus structure of this heptapeptide. By performing extensive replica-exchange molecular dynamics (REMD) simulations and conventional MD simulations, we found that the GVIGIAQ hexamers can adopt highly ordered bilayer ß-sheets and ß-barrels. In contrast, substitution of I149 by proline significantly reduces the ß-sheet probability and results in the disappearance of bilayer ß-sheet structures and the increase of disordered hexamers. We identified mixed parallel-antiparallel bilayer ß-sheets in both REMD and conventional MD simulations and provided the conformational transition from the experimentally observed parallel bilayer sheets to the mixed parallel-antiparallel bilayer ß-sheets. Our simulations suggest that the critical nucleus consists of six peptide chains and two additional peptide chains strongly stabilize this critical nucleus. The stabilized octamer is able to recruit additional random peptides into the ß-sheet. Therefore, our simulations provide insights into the critical nucleus formation and the smallest stable nucleus of the (147)GVIGIAQ(153) peptide.


Assuntos
Peptídeos beta-Amiloides/química , Modelos Moleculares , Superóxido Dismutase-1/química , Humanos , Peptídeos/química , Conformação Proteica em Folha beta
20.
J Phys Chem B ; 119(44): 14236-44, 2015 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-26447771

RESUMO

Neuronal calcium sensor-1 (NCS-1) protein has been implicated in multiple neuronal functions by binding partners mostly through a largely exposed hydrophobic crevice (HC). In the absence of a ligand, the C-terminal tail (loop L3, residues D176 to V190) binds directly to the HC pocket as a ligand mimetic, occupying the HC and regulating its conformational stability. A recent experimental study reported that L3 deletion resulted in global structure destabilization. However, the influence of C-terminal tail on the conformations of NCS-1 protein is unclear at the atomic level. In this study, we investigated the structural properties and the conformational dynamics of wild type NCS-1 and L3 truncation variant by extensive all-atom molecular dynamics (MD) simulations. Our cumulative 2 µs MD simulations demonstrated that L3 deletion increased the structural flexibility of the C-domain and the distant N-domain. The community network analysis illustrated that C-terminal tail truncation weakened the interdomain correlation. Moreover, our data showed that the variant significantly disrupted the salt bridges network and expanded simultaneously the global structure and HC. These conformational changes caused by C-terminal tail truncation may affect the regulation of target interactions. Our study provides atomic details of the conformational dynamics effects of the C-terminal tail on human wild type NCS-1.


Assuntos
Simulação de Dinâmica Molecular , Proteínas Sensoras de Cálcio Neuronal/química , Neuropeptídeos/química , Humanos , Conformação Proteica
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