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While immune checkpoint blockade (ICB) has shown promise for clinical cancer therapy, its efficacy has only been observed in a limited subset of patients and the underlying mechanisms regulating innate and acquired resistance to ICB of tumor cells remain poorly understood. Here, we identified ependymin-related protein 1 (EPDR1) as an important tumor-intrinsic regulator of PD-L1 expression and tumor immune evasion. Aberrant expression of EPDR1 in hepatocellular carcinoma is associated with immunosuppression. Mechanistically, EPDR1 binds to E3 ligase TRIM21 and disrupts its interaction with IkappaB kinase-b, suppressing its ubiquitylation and autophagosomal degradation and enhancing NF-κB-mediated transcriptional activation of PD-L1. Further, we validated through a mouse liver cancer model that EPDR1 mediates exhaustion of CD8+ T cells and promotes tumor progression. In addition, we observed a positive correlation between EPDR1 and PD-L1 expression in both human and mouse liver cancer samples. Collectively, our study reveals a previously unappreciated role of EPDR1 in orchestrating tumor immune evasion and cancer progression.
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Mulberry leaves have been used in traditional Chinese medicine due to their antioxidant, antidiabetic, and antihyperlipidemic properties. A previous study showed that ultraviolet-B radiation followed by dark incubation could improve the contents of active ingredients in mulberry leaves, such as moracin N and chalcomoracin. The endoplasmic reticulum (ER) serves as a protein quality control center and the location for protein synthesis, which is involved in the response to the environmental stress in plants. To investigate the mechanisms in response to ultraviolet-B radiation followed by dark incubation (UV + D), ER proteomics was performed on mulberry leaves. The ER protein markers, glucose-regulated protein (GRP78), and calnexin (CNX), were significantly higher in the ER fraction than in the total protein fraction, indicating that the ER was purified. Compared to the control, the abundance of protein disulfide isomerase, UDP-glucose glycoprotein glucosyltransferase, CNX, and calreticulin proteins decreased, while of the abundance of heat shock-related proteins increased under stress. P450 enzyme system-related proteins and ribosomal proteins showed significant increases. These results suggest that under UV + D stress, mulberry leaves activated the cell redox and ER quality control systems, enhancing protein synthesis and weakening N-glycan biosynthesis in the ER to resist the damage.
Assuntos
Morus , Proteômica , Calnexina/metabolismo , Retículo Endoplasmático/metabolismo , Morus/metabolismo , Folhas de Planta/metabolismo , Proteômica/métodosRESUMO
Volatile organic compounds (VOCs) in urine are potential biomarkers of breast cancer. Previously, our group has investigated breast cancer through analysis of VOCs in mouse urine and identified a panel of VOCs with the ability to monitor tumor progression. However, an unanswered question is whether VOCs can be exploited similarly to monitor the efficacy of antitumor treatments over time. Herein, subsets of tumor-bearing mice were treated with pitavastatin at high (8 mg/kg) and low (4 mg/kg) concentrations, and urine was analyzed through solid-phase microextraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS). Previous investigations using X-ray and micro-CT analysis indicated pitavastatin administered at 8 mg/kg had a protective effect against mammary tumors, whereas 4 mg/kg treatments did not inhibit tumor-induced damage. VOCs from mice treated with pitavastatin were compared to the previously analyzed healthy controls and tumor-bearing mice using chemometric analyses, which revealed that mice treated with pitavastatin at high concentrations were significantly different than tumor-bearing untreated mice in the direction of healthy controls. Mice treated with low concentrations demonstrated significant differences relative to healthy controls and were reflective of tumor-bearing untreated mice. These results show that urinary VOCs can accurately and noninvasively predict the efficacy of pitavastatin treatments over time.
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Neoplasias Mamárias Animais , Compostos Orgânicos Voláteis , Animais , Quimiometria , Cromatografia Gasosa-Espectrometria de Massas/métodos , Camundongos , Quinolinas , Microextração em Fase Sólida/métodos , Compostos Orgânicos Voláteis/análiseRESUMO
In the pandemic of COVID-19, there are exposed individuals who are infected but lack distinct clinical symptoms. In addition, the diffusion of related information drives aware individuals to spontaneously seek resources for protection. The special spreading characteristic and coevolution of different processes may induce unexpected spreading phenomena. Thus we construct a three-layered network framework to explore how information-driven resource allocation affects SEIS (susceptible-exposed-infected-susceptible) epidemic spreading. The analyses utilizing microscopic Markov chain approach reveal that the epidemic threshold depends on the topology structure of epidemic network and the processes of information diffusion and resource allocation. Conducting extensive Monte Carlo simulations, we find some crucial phenomena in the coevolution of information diffusion, resource allocation and epidemic spreading. Firstly, when E-state (exposed state, without symptoms) individuals are infectious, long incubation period results in more E-state individuals than I-state (infected state, with obvious symptoms) individuals. Besides, when E-state individuals have strong or weak infectious capacity, increasing incubation period has an opposite effect on epidemic propagation. Secondly, the short incubation period induces the first-order phase transition. But enhancing the efficacy of resources would convert the phase transition to a second-order type. Finally, comparing the coevolution in networks with different topologies, we find setting the epidemic layer as scale-free network can inhibit the spreading of the epidemic.
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While urine has been considered as a useful bio-fluid for health monitoring, its dynamic changes to physical activity are not well understood. We examined urine's possible antitumor capability in response to medium-level, loading-driven physical activity. Urine was collected from mice subjected to 5-minute skeletal loading and human individuals before and after 30-minute step aerobics. Six cancer cell lines (breast, prostate, and pancreas) and a mouse model of the mammary tumor were employed to evaluate the effect of urine. Compared to urine collected prior to loading, urine collected post-activity decreased the cellular viability, proliferation, migration, and invasion of tumor cells, as well as tumor weight in the mammary fat pad. Detection of urinary volatile organic compounds and ELISA assays showed that the loading-conditioned urine reduced cholesterol and elevated dopamine and melatonin. Immunohistochemical fluorescent images presented upregulation of the rate-limiting enzymes for the production of dopamine and melatonin in the brain. Molecular analysis revealed that the antitumor effect was linked to the reduction in molecular vinculin-linked molecular force as well as the downregulation of the Lrp5-CSF1-CD105 regulatory axis. Notably, the survival rate for the high expression levels of Lrp5, CSF1, and CD105 in tumor tissues was significantly lowered in the Cancer Genome Atlas database. Collectively, this study revealed that 5- or 10-minute loading-driven physical activity was sufficient to induce the striking antitumor effect by activating the neuronal signaling and repressing cholesterol synthesis. The result supported the dual role of loading-conditioned urine as a potential tumor suppressor and a source of diagnostic biomarkers.
Assuntos
Urina/fisiologia , Adolescente , Adulto , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Dopamina/urina , Exercício Físico/fisiologia , Feminino , Humanos , Masculino , Neoplasias Mamárias Animais/urina , Melatonina/urina , Camundongos , Camundongos Endogâmicos C57BL , Células PC-3 , Transdução de Sinais/fisiologia , Adulto JovemRESUMO
Mechanical stimulations can prevent bone loss, but their effects on the tumor-invaded bone or solid tumors are elusive. Here, we evaluated the effect of knee loading, dynamic loads applied to the knee, on metastasized bone and mammary tumors. In a mouse model, tumor cells were inoculated to the mammary fat pad or the proximal tibia. Daily knee loading was then applied and metabolic changes were monitored mainly through urine. Urine samples were also collected from human subjects before and after step aerobics. The result showed that knee loading inhibited tumor progression in the loaded tibia. Notably, it also reduced remotely the growth of mammary tumors. In the urine, an altered level of cholesterol was observed with an increase in calcitriol, which is synthesized from a cholesterol derivative. In urinary proteins, knee loading in mice and step aerobics in humans markedly reduced WNT1-inducible signaling pathway protein 1, WISP1, which leads to poor survival among patients with breast cancer. In the ex vivo breast cancer tissue assay, WISP1 promoted the growth of cancer fragments and upregulated tumor-promoting genes, such as Runx2, MMP9, and Snail. Collectively, the present preclinical and human study demonstrated that mechanical stimulations, such as knee loading and step aerobics, altered urinary metabolism and downregulated WISP1. The study supports the benefit of mechanical stimulations for locally and remotely suppressing tumor progression. It also indicated the role of WISP1 downregulation as a potential mechanism of loading-driven tumor suppression.
Assuntos
Neoplasias Ósseas/terapia , Neoplasias da Mama/terapia , Proteínas de Sinalização Intercelular CCN/metabolismo , Terapia por Exercício , Neoplasias Mamárias Experimentais/terapia , Condicionamento Físico Animal , Proteínas Proto-Oncogênicas/metabolismo , Animais , Neoplasias Ósseas/secundário , Neoplasias da Mama/patologia , Proteínas de Sinalização Intercelular CCN/urina , Linhagem Celular Tumoral , Colesterol/urina , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas/urinaRESUMO
PURPOSE: To describe breakthrough vitreous hemorrhage secondary to polypoidal choroidal vasculopathy (PCV). METHODS: Patients with the diagnosis of PCV from January 2005 to March 2020 at Peking Union Medical College Hospital were retrospectively reviewed, cases with breakthrough vitreous hemorrhage were analyzed. Subgroup analysis was conducted regarding pachychoroid PCV and nonpachychoroid PCV. RESULTS: Among 722 PCV patients (834 eyes), 103 eyes with breakthrough vitreous hemorrhage (12.4%) were included. Pars plana vitrectomy and proper further interventions could significantly improve the best-corrected visual acuity from logMAR 2.15 ± 0.48 (Snellen 20/2825) to 1.65 ± 0.67 (20/893). Hemorrhagic retinal detachment, baseline central macular thickness, and best-corrected visual acuity were factors associated with final best-corrected visual acuity (P < 0.05). In the pachychoroid PCV group, patients were younger, all had hemorrhagic pigment epithelial detachment, with a higher prevalence of choroidal vascular hyperpermeability and hemorrhagic retinal detachment, thicker subfoveal choroidal thickness, and thinner central macular thickness; besides, the initial pars plana vitrectomy were more complicated, more additional surgeries had to be performed. More eyes in the nonpachychoroid PCV group had received anti-vascular endothelial growth factor or photodynamic therapy, mostly fibrovascular pigment epithelial detachment, the best-corrected visual acuity and the status of the fellow eye were significantly worse. For the final ocular status, more eyes in nonpachychoroid PCV group were taking anti-vascular endothelial growth factor monotherapy, whereas more eyes in pachychoroid PCV group were stable. The choroidal parameters of these two groups were all significantly different. CONCLUSION: Breakthrough vitreous hemorrhage is a troublesome complication of PCV. Pars plana vitrectomy and additional interventions are required for better prognosis. Vitreous hemorrhage secondary to pachychoroid PCV or nonpachychoroid PCV have different characteristics and prognosis.
Assuntos
Doenças da Coroide/complicações , Corioide/irrigação sanguínea , Pólipos/complicações , Acuidade Visual , Hemorragia Vítrea/epidemiologia , Idoso , China/epidemiologia , Corioide/diagnóstico por imagem , Doenças da Coroide/diagnóstico , Feminino , Angiofluoresceinografia/métodos , Seguimentos , Fundo de Olho , Humanos , Incidência , Masculino , Pólipos/diagnóstico , Prognóstico , Estudos Retrospectivos , Tomografia de Coerência Óptica/métodos , Hemorragia Vítrea/classificação , Hemorragia Vítrea/etiologiaRESUMO
Urinary volatile terpene (VT) levels are significantly altered with induced models of breast cancer in mice. The question arises whether VTs can detect the efficacy of antitumor treatments. BALB/c mice were injected with 4T1.2 murine tumor cells in the mammary pad or iliac artery to model localized breast cancer and induced bone metastasis. The effect of two dopaminergic antitumor agents was tested by conventional histology and altered VT levels. The headspace of urine specimens was analyzed by gas chromatography-mass spectrometry. In the localized model, the statistical significance (p < 0.05) was identified for 26% of VTs, and in the metastasis model, 19% of VTs. The authors discovered separate VT panels classifying localized/control [area under the curve (AUC) = 1.0] and metastasis/control (AUC = 0.98). Treatment samples were tested using these panels, which showed that mice treated with either agent were statistically significantly different from cancer samples, which is consistent with conventional analysis.
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Neoplasias , Compostos Orgânicos Voláteis , Animais , Cromatografia Gasosa-Espectrometria de Massas , Camundongos , Camundongos Endogâmicos BALB C , Microextração em Fase Sólida , Terpenos , Compostos Orgânicos Voláteis/análiseRESUMO
Bone is a frequent site of metastasis from breast cancer, and a desirable drug could suppress tumor growth as well as metastasis-linked bone loss. Currently, no drug is able to cure breast cancer-associated bone metastasis. In this study, we focused on statins that are known to inhibit cholesterol production and act as antitumor agents. After an initial potency screening of 7 U.S. Food and Drug Administration-approved statins, we examined pitavastatin as a drug candidate for inhibiting tumor and tumor-induced bone loss. In vitro analysis revealed that pitavastatin acted as an inhibitor of tumor progression by altering stress to the endoplasmic reticulum, down-regulating peroxisome proliferator-activated receptor γ, and reducing Snail and matrix metalloproteinase 9. In bone homeostasis, it blocked osteoclast development by suppressing transcription factors c-Fos and JunB, but stimulated osteoblast mineralization by regulating bone morphogenetic protein 2 and p53. In a mouse model, pitavastatin presented a dual role in tumor inhibition in the mammary fat pad, as well as in bone protection in the osteolytic tibia. In mass spectrometry-based analysis, volatile organic compounds (VOCs) that were linked to lipid metabolism and cholesterol synthesis were elevated in mice from the tumor-grown placebo group. Notably, pitavastatin-treated mice reduced specific VOCs that are linked to lipid metabolites in the mevalonate pathway. Collectively, the results lay a foundation for further investigation of pitavastatin's therapeutic efficacy in tumor-induced bone loss, as well as VOC-based diagnosis of tumor progression and treatment efficacy.-Wang, L., Wang, Y., Chen, A., Teli, M., Kondo, R., Jalali, A., Fan, Y., Liu, S., Zhao, X., Siegel, A., Minami, K., Agarwal, M., Li, B.-Y., Yokota, H. Pitavastatin slows tumor progression and alters urine-derived volatile organic compounds through the mevalonate pathway.
Assuntos
Ácido Mevalônico/metabolismo , Quinolinas/farmacologia , Compostos Orgânicos Voláteis/metabolismo , Animais , Neoplasias Ósseas/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Feminino , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/metabolismo , Metabolismo dos Lipídeos/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Osteoblastos/metabolismo , Células RAW 264.7RESUMO
Interleukin-27 (IL-27) has shown promise in halting tumor growth and mediating tumor regression in several models, including prostate cancer. We describe our findings on the effects of IL-27 on the gene expression changes of TC2R prostate adenocarcinoma cells. We utilized RNAseq to assess profile differences between empty vector control, vector delivering IL-27 modified at its C-terminus with a non-specific peptide, and IL-27 modified at the C-terminus with a peptide targeting the IL-6-Rα. The targeted IL-27 had higher bioactivity and activity in vivo in a recent study by our group, but the mechanisms underlying this effect had not been characterized in detail at the gene expression level on tumor cells. In the present work, we sought to examine potential mechanisms for targeted IL-27 enhanced activity directly on tumor cells. The targeted IL-27 appeared to modulate several changes that would be consistent with an anti-tumor effect, including upregulation in the Interferon (IFN) and Interferon regulatory factor (IRF), oxidative phosphorylation, Janus kinase/Signal transducers and activators of transcription (JAK/STAT), and eukaryotic initiation factor 2 (EIF2) signaling. Of these signaling changes predicted by ingenuity pathway analyses (IPA), the novel form also with the highest significance (-log(Benjamini-Hochberg (B-H)) p-value) was the EIF2 signaling upregulation. We validated this predicted change by assaying for eukaryotic initiation factor 2 alpha (eIF2α), or phosphorylated eIF2α (p-eIF2α), and caspase-3 levels. We detected an increase in the phosphorylated form of eIF2α and in the cleaved caspase-3 fraction, indicating that the EIF2 signaling pathway was upregulated in these prostate tumor cells following targeted IL-27 gene delivery. This approach of targeting cytokines to enhance their activity against cancer cells is a novel approach to help augment IL-27's bioactivity and efficacy against prostate tumors and could be extended to other conditions where it could help interfere with the EIF2α pathway and promote caspase-3 activation.
Assuntos
Adenocarcinoma/metabolismo , Marcação de Genes/métodos , Terapia Genética/métodos , Interleucina-27/genética , Neoplasias da Próstata/metabolismo , Receptores de Interleucina-6/genética , Transdução de Sinais , Adenocarcinoma/genética , Animais , Linhagem Celular Tumoral , Fator de Iniciação 2 em Eucariotos/metabolismo , Interferons/metabolismo , Interleucina-27/química , Interleucina-27/metabolismo , Janus Quinases/metabolismo , Masculino , Camundongos , Neoplasias da Próstata/genética , Domínios Proteicos , Receptores de Interleucina-6/metabolismo , Fatores de Transcrição STAT/metabolismoRESUMO
Ultraviolet (UV)-B radiation acts as an elicitor to enhance the production of secondary metabolites in medicinal plants. To investigate the mechanisms, which lead to secondary metabolites in Catharanthus roseus under UVB radiation, a phosphoproteomic technique was used. ATP content increased in the leaves of C. roseus under UVB radiation. Phosphoproteins related to calcium such as calmodulin, calcium-dependent kinase, and heat shock proteins increased. Phosphoproteins related to protein synthesis/modification/degradation and signaling intensively changed. Metabolomic analysis indicated that the metabolites classified with pentoses, aromatic amino acids, and phenylpropanoids accumulated under UVB radiation. Phosphoproteomic and immunoblot analyses indicated that proteins related to glycolysis and the reactive-oxygen species scavenging system were changed under UVB radiation. These results suggest that UVB radiation activates the calcium-related pathway and reactive-oxygen species scavenging system in C. roseus. These changes lead to the upregulation of proteins, which are responsible for the redox reactions in secondary metabolism and are important for the accumulation of secondary metabolites in C. roseus under UVB radiation.
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Catharanthus/metabolismo , Fosfoproteínas/genética , Proteínas de Plantas/metabolismo , Metabolismo Secundário/efeitos da radiação , Cálcio/metabolismo , Calmodulina/genética , Calmodulina/metabolismo , Catharanthus/genética , Catharanthus/efeitos da radiação , Fosfoproteínas/efeitos da radiação , Folhas de Planta/metabolismo , Folhas de Planta/efeitos da radiação , Proteínas de Plantas/efeitos da radiação , Raízes de Plantas/metabolismo , Raízes de Plantas/efeitos da radiação , Plantas Medicinais/efeitos da radiação , Metabolismo Secundário/genética , Transdução de Sinais/efeitos da radiação , Raios UltravioletaRESUMO
Bone is a mechano-sensitive tissue that alters its structure and properties in response to mechanical loading. We have previously shown that application of lateral dynamic loads to a synovial joint, such as the knee and elbow, suppresses degradation of cartilage and prevents bone loss in arthritis and postmenopausal mouse models, respectively. While loading effects on pathophysiology have been reported, mechanical effects on the loaded joint are not fully understood. Because the direction of joint loading is non-axial, not commonly observed in daily activities, strain distributions in the laterally loaded joint are of great interest. Using elbow loading, we herein characterized mechanical responses in the loaded ulna focusing on the distribution of compressive strain. In response to 1-N peak-to-peak loads, which elevate bone mineral density and bone volume in the proximal ulna in vivo, we conducted finite-element analysis and evaluated strain magnitude in three loading conditions. The results revealed that strain of ~ 1000 µstrain (equivalent to 0.1% compression) or above was observed in the limited region near the loading site, indicating that the minimum effective strain for bone formation is smaller with elbow loading than axial loading. Calcein staining indicated that elbow loading increased bone formation in the regions predicted to undergo higher strain.
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Análise de Elementos Finitos , Membro Anterior/fisiologia , Ulna/fisiologia , Animais , Densidade Óssea , Força Compressiva , Modelos Animais de Doenças , Feminino , Camundongos Endogâmicos BALB C , Tamanho do Órgão , Osteogênese/fisiologia , Estresse Mecânico , Ulna/diagnóstico por imagem , Suporte de Carga/fisiologiaRESUMO
Morus alba is an important medicinal plant that is used to treat human diseases. The leaf, branch, and root of Morus can be applied as antidiabetic, antioxidant, and anti-inflammatory medicines, respectively. To explore the molecular mechanisms underlying the various pharmacological functions within different parts of Morus, organ-specific proteomics were performed. Protein profiles of the Morus leaf, branch, and root were determined using a gel-free/label-free proteomic technique. In the Morus leaf, branch, and root, a total of 492, 414, and 355 proteins were identified, respectively, including 84 common proteins. In leaf, the main function was related to protein degradation, photosynthesis, and redox ascorbate/glutathione metabolism. In branch, the main function was related to protein synthesis/degradation, stress, and redox ascorbate/glutathione metabolism. In root, the main function was related to protein synthesis/degradation, stress, and cell wall. Additionally, organ-specific metabolites and antioxidant activities were analyzed. These results revealed that flavonoids were highly accumulated in Morus root compared with the branch and leaf. Accordingly, two root-specific proteins named chalcone flavanone isomerase and flavonoid 3,5-hydroxylase were accumulated in the flavonoid pathway. Consistent with this finding, the content of the total flavonoids was higher in root compared to those detected in branch and leaf. These results suggest that the flavonoids in Morus root might be responsible for its biological activity and the root is the main part for flavonoid biosynthesis in Morus.
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Morus/metabolismo , Especificidade de Órgãos , Proteômica/métodos , Coloração e Rotulagem , Antioxidantes/metabolismo , Ciclo do Ácido Cítrico , Flavonoides/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Glicólise , Metaboloma , Morus/genética , Especificidade de Órgãos/genética , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Raízes de Plantas/genética , Metabolismo SecundárioRESUMO
Previous studies reported that miR-146a was involved in small intestine ischemia-reperfusion (I/R) injury, but the mechanism is largely vague. Here, we aimed to identify the change of miR-146a in patients with mesenteric ischemia and explore the potential regulatory mechanism of miR-146a in intestine epithelial cells survival under ischemia and I/R injury. The plasma of 20 patients with mesenteric ischemia and 25 controls was collected to examine the miR-146a expression by qPCR. Rat intestinal epithelial cells (IEC-6) and 24 male Sprague-Dawley rats were included to build ischemia and I/R model in vitro and in vivo. The qPCR results showed that miR-146a decreased both in the plasma of patients with mesenteric ischemia and in IEC-6 cells and rat small intestine tissues in ischemia and I/R model compared to controls. Both the in vitro and in vivo results showed that I/R resulted in more severe apoptotic injury than ischemia. Cleaved-caspase 3, TLR4, TRAF6, and nuclear NF-κB p65 were up-regulated accompanying reduced XIAP and SOCS3 expression in intestinal ischemia and I/R injury. After up-regulation of miR-146a in IEC-6 cells, increased cell survival and decreased cell apoptosis were observed, concomitant with decreased cleaved-caspase 3 and down-regulated TLR4/TRAF6/NF-κB pathway. What is more, this protective effect was blocked by TRAF6 overexpression and increased nuclear NF-κB p65 nuclear. Taken together, this study revealed that miR-146a expression was decreased in small intestine ischemia and I/R injury. And miR-146a improves intestine epithelial cells survival under ischemia and I/R injury through inhibition TLR4, TRAF6, and p-IκBα, subsequently leading to decreased NF-κB p65 nuclear translocation.
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Intestino Delgado/metabolismo , Isquemia Mesentérica/metabolismo , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Traumatismo por Reperfusão/metabolismo , Fator 6 Associado a Receptor de TNF/metabolismo , Receptor 4 Toll-Like/metabolismo , Adulto , Animais , Apoptose , Estudos de Casos e Controles , Caspase 3/metabolismo , Hipóxia Celular , Linhagem Celular , Modelos Animais de Doenças , Feminino , Humanos , Intestino Delgado/irrigação sanguínea , Intestino Delgado/patologia , Peptídeos e Proteínas de Sinalização Intracelular , Masculino , Isquemia Mesentérica/genética , Isquemia Mesentérica/patologia , MicroRNAs/genética , Pessoa de Meia-Idade , Inibidor de NF-kappaB alfa/genética , Inibidor de NF-kappaB alfa/metabolismo , NF-kappa B/genética , Fosforilação , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/patologia , Traumatismo por Reperfusão/prevenção & controle , Transdução de Sinais , Fator 6 Associado a Receptor de TNF/genética , Fatores de Tempo , Fator de Transcrição RelA/metabolismo , TransfecçãoRESUMO
Pharmaceutically active compounds from medical plants are attractive as a major source for new drug development. Prenylated stilbenoids with increased lipophilicity are valuable secondary metabolites which possess a wide range of biological activities. So far, many prenylated stilbenoids have been isolated from Morus alba but the enzyme responsible for the crucial prenyl modification remains unknown. In the present study, a stilbenoid-specific prenyltransferase (PT), termed Morus alba oxyresveratrol geranyltransferase (MaOGT), was identified and functionally characterized in vitro. MaOGT recognized oxyresveratrol and geranyl diphosphate (GPP) as natural substrates, and catalyzed oxyresveratrol prenylation. Our results indicated that MaOGT shared common features with other aromatic PTs, e.g. multiple transmembrane regions, conserved functional domains and targeting to plant plastids. This distinct PT represents the first stilbenoid-specific PT accepting GPP as a natural prenyl donor, and could help identify additional functionally varied PTs in moraceous plants. Furthermore, MaOGT might be applied for high-efficiency and large-scale prenylation of oxyresveratrol to produce bioactive compounds for potential therapeutic applications.
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Dimetilaliltranstransferase/metabolismo , Difosfatos/metabolismo , Diterpenos/metabolismo , Morus/enzimologia , Estilbenos/metabolismo , Catálise , Dimetilaliltranstransferase/genética , Morus/genética , Morus/metabolismo , Organismos Geneticamente Modificados , Filogenia , Extratos Vegetais/metabolismo , Folhas de Planta/enzimologia , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas , Prenilação , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Alinhamento de Sequência , Especificidade por Substrato , NicotianaRESUMO
BACKGROUND: The inflammatory immune response plays an important role in mesenteric ischemia and ischemia-reperfusion injury. Toll-like receptor 4 (TLR4) is a critical receptor in transduction of the inflammatory response and plays an important role in intestinal homeostasis. Tumor necrosis factor receptor-associated factor 6 (TRAF6), known as a key adaptor protein downstream of TLR4, is involved in the inflammatory response by activating multiple apoptotic signaling pathways. However, mechanisms of the suppressor of cytokine signaling-1 (SOCS-1) in regulating cell inflammation and apoptosis are still obscure. OBJECTIVES: To investigate the TLR4-TRAF6 signaling pathway in intestinal ischemia and reperfusion injury, as well as SOCS-1 expression after ischemic preconditioning in the rat intestine. METHODS: The small bowel ischemia, ischemia-reperfusion, and preconditioning models were induced using ligation of the superior mesenteric artery in male Sprague-Dawley rats; then, the mRNA and protein levels of TLR4, TRAF6, and SOCS-1 were analyzed using real-time PCR, Western blot, and immunohistochemistry, respectively. RESULTS: The expression of TLR4 and TRAF6 was gradually increased with increasing intestinal ischemia duration, but increased substantially after ischemia-reperfusion injury. After ischemic preconditioning, TLR4 and TRAF6 expressions decreased; however, expression of SOCS-1 and the TLR4-TRAF6 pathway inhibitor was increased. CONCLUSION: These data show that ischemic preconditioning may induce the activation of SOCS-1 to inhibit the TLR4-TRAF6 signaling pathway, thereby playing a protective role in ischemia-reperfusion injury.
Assuntos
Intestino Delgado/imunologia , Precondicionamento Isquêmico , Isquemia Mesentérica/imunologia , Traumatismo por Reperfusão/imunologia , Proteína 1 Supressora da Sinalização de Citocina/imunologia , Fator 6 Associado a Receptor de TNF/imunologia , Receptor 4 Toll-Like/imunologia , Animais , Apoptose/imunologia , Western Blotting , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Intestino Delgado/patologia , Ligadura , Masculino , Artéria Mesentérica Superior/cirurgia , Isquemia Mesentérica/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Proteína Serina-Treonina Quinases de Interação com Receptores , Traumatismo por Reperfusão/patologia , Transdução de Sinais , Proteína 1 Supressora da Sinalização de Citocina/genética , Fator 6 Associado a Receptor de TNF/genética , Receptor 4 Toll-Like/genéticaRESUMO
Salubrinal is an agent that reduces the stress to the endoplasmic reticulum by inhibiting de-phosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α). We and others have previously shown that the elevated phosphorylation of eIF2α stimulates bone formation and attenuates bone resorption. In this study, we applied salubrinal to a mouse model of osteogenesis imperfecta (Oim), and examined whether it would improve Oim's mechanical property. We conducted in vitro experiments using RAW264.7 pre-osteoclasts and bone marrow derived cells (BMDCs), and performed in vivo administration of salubrinal to Oim (+/-) mice. The animal study included two control groups (wildtype and Oim placebo). The result revealed that salubrinal decreased expression of nuclear factor of activated T cells cytoplasmic 1 (NFATc1) and suppressed osteoclast maturation, and it stimulated mineralization of mesenchymal stem cells from BMDCs. Furthermore, daily injection of salubrinal at 2 mg/kg for 2 months made stiffness (N/mm) and elastic module (GPa) of the femur undistinguishable to those of the wildtype control. Collectively, this study supported salubrinal's beneficial role to Oim's femora. Unlike bisphosphonates, salubrinal stimulates bone formation. For juvenile OI patients who may favor strengthening bone without inactivating bone remodeling, salubrinal may present a novel therapeutic option.
Assuntos
Remodelação Óssea/efeitos dos fármacos , Cinamatos/farmacologia , Cinamatos/uso terapêutico , Fêmur/efeitos dos fármacos , Osteogênese Imperfeita/tratamento farmacológico , Tioureia/análogos & derivados , Animais , Fenômenos Biomecânicos/efeitos dos fármacos , Fenômenos Biomecânicos/fisiologia , Remodelação Óssea/fisiologia , Linhagem Celular , Proteínas de Ligação a DNA/antagonistas & inibidores , Feminino , Fêmur/patologia , Fêmur/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Osteogênese Imperfeita/patologia , Osteogênese Imperfeita/fisiopatologia , Tioureia/farmacologia , Tioureia/uso terapêutico , Fatores de Transcrição/antagonistas & inibidoresRESUMO
Integrated stress responses (ISR) may lead to cell death and tissue degeneration via eukaryotic translation initiation factor 2 α (eIF2α)-mediated signaling. Alleviating ISR by modulating eIF2α phosphorylation can reduce the symptoms associated with various diseases. Guanabenz is known to elevate the phosphorylation level of eIF2α and reduce pro-inflammatory responses. However, the mechanism of its action is not well understood. In this study, we investigated the signaling pathway through which guanabenz induces anti-inflammatory effects in immune cells, in particular macrophages. Genome-wide mRNA profiling followed by principal component analysis predicted that colony stimulating factor 2 (Csf2, or GM-CSF as granulocyte macrophage colony stimulating factor) is involved in the responses to guanabenz. A partial silencing of Csf2 or eIF2α by RNA interference revealed that Interleukin-6 (IL6), Csf2, and Cyclooxygenase-2 (Cox2) are downregulated by guanabenz-driven phosphorylation of eIF2α. Although expression of IL1ß and Tumor Necrosis Factor-α (TNFα) was suppressed by guanabenz, their downregulation was not directly mediated by eIF2α signaling. Collectively, the result herein indicates that anti-inflammatory effects by guanabenz are mediated by not only eIF2α-dependent but also eIF2α-independent signaling.
Assuntos
Anti-Inflamatórios/farmacologia , Regulação para Baixo , Fator de Iniciação 2 em Eucariotos/metabolismo , Guanabenzo/farmacologia , Transdução de Sinais , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Células Cultivadas , Ciclo-Oxigenase 2/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Inflamação/metabolismo , Interleucina-6/metabolismo , Células Jurkat , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Transcriptoma , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND/AIMS: Promyelocytic leukemia (PML) protein is a tumor suppressor that fuses with retinoic acid receptor-α (PML-RARα) to contribute to the initiation of acute promyelocytic leukemia (APL). Arsenic trioxide (ATO) upregulates expression of TGF-ß1, promoting collagen synthesis in osteoblasts, and ATO binds directly to PML to induce oligomerization, sumoylation, and ubiquitination. However, how ATO upregulates TGF-ß1 expression is uncertain. Thus, we suggested that PML sumoylation is responsible for regulation of TGF-ß1 protein expression. METHODS: Kunming mice were treated with ATO, and osteoblasts were counted under scanning electron microscopy. Masson's staining was used to quantify collagen content. hFOB1.19 cells were transfected with siRNA against UBC9 or RNF4, and then treated with ATO or FBS. TGF-ß1, PML expression, and sumoylation were quantified with Western blot, and collagen quantified via immunocytochemistry. RESULTS: ATO enhanced osteoblast accumulation, collagen synthesis, and PML-NB formation in vivo. Knocking down UBC9 in hFOB1.19 cells inhibited ATO- and FBS-induced PML sumoylation, TGF-ß1 expression, and collagen synthesis. Conversely, knocking down RNF4 enhanced ATO- and FBS-induced PML sumoylation, TGF-ß1 expression, and collagen synthesis. CONCLUSION: These data suggest that PML sumoylation is required for ATO-induced collagen synthesis in osteoblasts.
Assuntos
Colágeno/metabolismo , Óxidos/toxicidade , Sumoilação/efeitos dos fármacos , Proteínas Supressoras de Tumor/metabolismo , Animais , Trióxido de Arsênio , Arsenicais , Linhagem Celular , Fêmur/patologia , Humanos , Masculino , Camundongos , Microscopia Eletrônica , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Proteína da Leucemia Promielocítica , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Proteína SUMO-1/metabolismo , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Enzimas de Conjugação de Ubiquitina/antagonistas & inibidores , Enzimas de Conjugação de Ubiquitina/genética , Enzimas de Conjugação de Ubiquitina/metabolismoRESUMO
Morus plants played a pivotal role in ancient Chinese sericulture and silk production, which served as critical components of economy and culture. Besides, many parts of mulberry trees, including roots, leaves, stems, and fruits, hold various medicinal value, and have been utilized in traditional medicine for thousands of years. The chemical composition of mulberry has been reported in many literatures, while the characteristic compounds have not been systematically summarized. In this review, we focused on the polyphenolic compounds in mulberry, including flavonoids, 2-arylbenzofurans, and Diels-Alder (D-A) adducts, and summarized their structural features, structure-activity relationships, and potential biosynthetic pathways. The results revealed a characteristic class of 2'-hydroxylated flavonoids and stilbenes which played an important role in the biosynthesis of downstream 2-arylbenzofurans and D-A adducts in mulberry but had been overlooked by most studies. The prenylated modifications of different compounds were also discussed and their function as precursors of D-A adducts was emphasized. We also describe the effects of different modifications on biological activities. Besides, the chemical composition of Morus was most similar to that of Artocarpus in the Moraceae family in that they had almost identical characteristic compounds. Finally, a putative total biosynthetic pathway of D-A adducts in mulberry was proposed based on structure derivation and combination of verified reactions. This review contributes to the understanding of the biological activity and biosynthesis of the characteristic components of Morus plants.