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The failure of bone regeneration has been considered as a serious problem that troubling patients for decades, most of which was resulted by the poor angiogenesis and chronic inflammation after surgery. Among multiple materials applied in the repair of bone defect, silicate bioceramics attracted researchers because of its excellent bioactivity. The purpose of this study was to detect the effect of specific bioactive glass ceramic (AP40, based on crystalline phases of apatite and wollastonite) on angiogenesis and the subsequent bone growth through the modulation of macrophages. Two groups were included in this study: control group (macrophages without any stimulation, denominated as Control) and AP40 group (macrophages incubated on AP40). This study investigated the effect of AP40 on macrophages polarization (RAW264.7) and angiogenesis in vitro and in vivo. Additionally, the changes of angiogenic ability regulated by macrophages were explored. AP40 showed excellent angiogenesis potential and the expression of CD31 was promoted through the modulation of macrophages toward M2 subtype. Additionally, the macrophages incubated on AP40 synthesized more PDGF-BB comparing to macrophages without any stimulation, which contributed to the improved angiogenetic ability of human umbilical vein endothelial cells (HUVECs). Results of in vivo studies indicated increased bone ingrowth along the implants, which indicated the potential of bioceramics for bone defect repair clinically.
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Regeneração Óssea , Cerâmica , Células Endoteliais da Veia Umbilical Humana , Macrófagos , Neovascularização Fisiológica , Impressão Tridimensional , Silicatos , Cerâmica/farmacologia , Cerâmica/química , Animais , Camundongos , Silicatos/química , Silicatos/farmacologia , Regeneração Óssea/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Macrófagos/metabolismo , Humanos , Células RAW 264.7 , Células Endoteliais da Veia Umbilical Humana/metabolismo , Porosidade , Coelhos , AngiogêneseRESUMO
The regeneration of demineralized enamel holds great significance in the treatment of dental caries. Amelogenin (Ame), an essential protein for mediating natural enamel growth, is no longer secreted after enamel has fully matured in childhood. Although biomimetic mineralization based on peptides or proteins has made significant progress, easily accessible, low-cost, biocompatible and highly effective Ame mimics are still lacking. Herein, we construct a series of amphiphilic branched polypeptides (CAMPs) by facile coupling of the Ame's C-terminal segment and poly(γ-benzyl-L-glutamate), which serves to simulate the Ame's hydrophobic N-terminal segment. Among them, CAMP15 is the best biomimetic mineralization template with great self-assembly performance to guide the oriented crystallization of hydroxyapatite and is capable of inhibiting the adhesion of Streptococcus mutans and Staphylococcus aureus on the enamel surfaces. This work highlights the potential application of amphiphilic branched polypeptide as Ame mimics in repairing defected enamel, providing a promising strategy for prevention and treatment of dental caries.
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Amelogenina , Materiais Biomiméticos , Esmalte Dentário , Peptídeos , Streptococcus mutans , Amelogenina/química , Amelogenina/farmacologia , Peptídeos/química , Peptídeos/farmacologia , Peptídeos/síntese química , Esmalte Dentário/química , Esmalte Dentário/efeitos dos fármacos , Streptococcus mutans/efeitos dos fármacos , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologia , Materiais Biomiméticos/síntese química , Staphylococcus aureus/efeitos dos fármacos , Animais , Propriedades de Superfície , Humanos , Aderência Bacteriana/efeitos dos fármacos , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/síntese químicaRESUMO
MALT1 (mucosa-associated lymphoid tissue lymphoma translocation protein 1) is a human paracaspase protein with proteolytic activity via its caspase-like domain. The pharmacological inhibition of MALT1 by MI-2, a specific chemical inhibitor, diminishes the response of endothelial cells to inflammatory stimuli. However, it is largely unknown how MALT1 regulates the functions of vascular smooth muscle cells (SMCs). This study aims to investigate the impact of MALT1 inhibition by MI-2 on the functions of vascular SMCs, both in vitro and in vivo. MI-2 treatment led to concentration- and time-dependent cell death of cultured aortic SMCs, which was rescued by the iron chelator deferoxamine (DFO) or ferrostatin-1 (Fer-1), a specific inhibitor of ferroptosis, but not by inhibitors of apoptosis (Z-VAD-fmk), pyroptosis (Z-YVAD-fmk), or necrosis (Necrostatin-1, Nec-1). MI-2 treatment downregulated the expression of glutathione peroxidase 4 (GPX4) and ferritin heavy polypeptide 1 (FTH1), which was prevented by pre-treatment with DFO or Fer-1. MI-2 treatment also activated autophagy, which was inhibited by Atg7 deficiency or bafilomycin A1 preventing MI-2-induced ferroptosis. MI-2 treatment reduced the cleavage of cylindromatosis (CYLD), a specific substrate of MALT1. Notably, MI-2 treatment led to a rapid loss of contractility in mouse aortas, which was prevented by co-incubation with Fer-1. Moreover, local application of MI-2 significantly reduced carotid neointima lesions and atherosclerosis in C57BL/6J mice and apolipoprotein-E knockout (ApoE-/-) mice, respectively, which were both ameliorated by co-treatment with Fer-1. In conclusion, the present study demonstrated that MALT1 inhibition induces ferroptosis of vascular SMCs, likely contributing to its amelioration of proliferative vascular diseases.
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Prenylated proteins contain C15 or C20 isoprenoids linked to cysteine residues positioned near their C-termini. Here we describe the preparation of isoprenoid diphosphate analogues incorporating diazirine groups that can be used to probe interactions between prenylated proteins and other proteins that interact with them. Studies using synthetic peptides and whole proteins demonstrate that these diazirine analogues are efficient substrates for prenyltransferases. Photo-cross-linking experiments using peptides incorporating the diazirine-functionalized isoprenoids selectively cross-link to several different proteins. These new isoprenoid analogues should be broadly useful in the studies of protein prenylation.
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Diazometano , Difosfatos , Peptídeos , Cisteína , TerpenosRESUMO
Neointima lesion and atherosclerosis are proliferative vascular diseases associated with deregulated proliferation of vascular smooth muscle cells (SMCs). CFI-400945 is a novel, highly effective anticancer drug that inhibits polo-like kinase 4 (PLK4) and targets mitosis. In this study, we aim to investigate how CFI-400945 affects the development of proliferative vascular diseases. In C57BL/6 mice, neointima formation was generated by complete carotid ligation. In apolipoprotein E knockout (ApoE-/-) mice fed a high-fat diet, atherosclerosis was induced by partial carotid ligation. CFI-400945 was directly applied to carotid arteries via a perivascular collar. Our results showed that CFI-400945 drastically inhibited neointima formation but significantly accelerated atherosclerosis. In vitro studies showed that CFI-400945 treatment induced SMC polyploidization and arrested cells in the G2/M phase. CFI-400945 treatment upregulated p53 and p27 expression but decreased p21 and cyclin B1 expression. CFI-400945 also induced SMC apoptosis, which was inhibited by hydroxyurea, a DNA synthesis inhibitor that inhibits polyploidization. Furthermore, CFI-400945 caused supernumerary centrosomes, leading to mitotic failure, resulting in polyploidization. In conclusion, CFI-400945 prevents carotid arterial neointima formation in C57BL/6 mice but accelerates atherosclerosis in ApoE-/- mice, likely through mitotic arrest and subsequent induction of polyploidization and apoptosis.
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OBJECTIVE: To investigate the clinical outcomes of a novel soft tissue repair patch (porcine small intestinal submucosa patch, SIS patch) in the treatment of full-thickness hand skin defects. METHODS: From January 2017 to July 2019, 80 patients with hand soft tissue defects, who met the inclusion criteria, were retrospectively reviewed and divided into two groups. After debridement, patients in group A were treated with the novel SIS patch to cover the wound, and patients in group B were treated with autologous skin graft. The dimensions of skin defect area and healing outcome were evaluated and recorded. Scar assessment was carried out using Scar Cosmesis Assessment and Rating Scale (SCAR scale) at the last follow-up postoperation, and the recovery of wound sensation was assessed at the same time using British Medical Research Council (BMRC) grading of sensorimotor recovery. All the data were collected and statistically analyzed. RESULTS: A total of 80 patients were enrolled in the study with 40 patients in each group. Four patients in group A and 5 patients in group B were excluded due to wound infection and lost to follow-up. There were 36 patients in group A and 35 patients in group B finally got follow-up postoperation with mean interval of 12.75 ± 5.61 months in group A and 14.11 ± 5.42 months in group B. The dimensions of skin defect area in group A ranged from 7.5 to 87.5 cm2 (mean 25.97 ± 18.66 cm2) and in group B ranged from 7.5 to 86.25 cm2 (mean 33.61 ± 19.27 cm2) which have no significant difference (P > 0.05). SCAR scale results of group A and group B were 10.98 ± 0.33 and 9.49 ± 0.35, respectively, and the difference was statistically significant (P < 0.05). BMRC grading results showed 6 cases of S4, 11 cases of S3+, 5 cases of S3, 6 cases of S2, 6 cases of S1 and 2 cases of S0 in group A, and 8 cases of S4, 10 cases of S3+, 7 cases of S3, 4 cases of S2, 5 cases of S1, and 1 case of S0 in group B, which had no significant difference between them (P > 0.05). CONCLUSIONS: The novel SIS patch is an applicable biological material in the treatment of hand skin defect, which could achieve a better cosmetic appearance of the newborn skin tissue.
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Procedimentos de Cirurgia Plástica , Lesões dos Tecidos Moles , Animais , Suínos , Estudos Retrospectivos , Pele/lesões , Transplante de Pele/métodos , Cicatrização , Cicatriz , Resultado do Tratamento , Lesões dos Tecidos Moles/cirurgiaRESUMO
The O-linked ß-N-acetylglucosamine (O-GlcNAc) transferase (OGT) mediates intracellular O-GlcNAcylation modification. O-GlcNAcylation occurs on Ser/Thr residues and is important for numerous physiological processes. OGT is essential for dividing mammalian cells and is involved in many human diseases; however, many of its fundamental substrates during cell division remain unknown. Here, we focus on the effect of OGT on polo-like kinase 1 (PLK1), a mitotic master kinase that governs DNA replication, mitotic entry, chromosome segregation, and mitotic exit. We show that PLK1 interacts with OGT and is O-GlcNAcylated. By utilizing stepped collisional energy/higher-energy collisional dissociation mass spectrometry, we found a peptide fragment of PLK1 that is modified by O-GlcNAc. Further mutation analysis of PLK1 shows that the T291A mutant decreases O-GlcNAcylation. Interestingly, T291N is a uterine carcinoma mutant in The Cancer Genome Atlas. Our biochemical assays demonstrate that T291A and T291N both increase PLK1 stability. Using stable H2B-GFP cells, we found that PLK1-T291A and PLK1-T291N mutants display chromosome segregation defects and result in misaligned and lagging chromosomes. In mouse xenograft models, we demonstrate that the O-GlcNAc-deficient PLK1-T291A and PLK1-T291N mutants enhance uterine carcinoma in animals. Hence, we propose that OGT partially exerts its mitotic function through O-GlcNAcylation of PLK1, which might be one mechanism by which elevated levels of O-GlcNAc promote tumorigenesis.
Assuntos
Divisão Celular , Proteínas Serina-Treonina Quinases , Neoplasias Uterinas , Animais , Feminino , Humanos , Camundongos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Neoplasias Uterinas/enzimologia , Neoplasias Uterinas/genética , Acilação , Divisão Celular/fisiologia , Mutação , Quinase 1 Polo-LikeRESUMO
BACKGROUND: PKA (protein kinase A)-mediated phosphorylation of cardiac RyR2 (ryanodine receptor 2) has been extensively studied for decades, but the physiological significance of PKA phosphorylation of RyR2 remains poorly understood. Recent determination of high-resolution 3-dimensional structure of RyR2 in complex with CaM (calmodulin) reveals that the major PKA phosphorylation site in RyR2, serine-2030 (S2030), is located within a structural pathway of CaM-dependent inactivation of RyR2. This novel structural insight points to a possible role of PKA phosphorylation of RyR2 in CaM-dependent inactivation of RyR2, which underlies the termination of Ca2+ release and induction of cardiac Ca2+ alternans. METHODS: We performed single-cell endoplasmic reticulum Ca2+ imaging to assess the impact of S2030 mutations on Ca2+ release termination in human embryonic kidney 293 cells. Here we determined the role of the PKA site RyR2-S2030 in a physiological setting, we generated a novel mouse model harboring the S2030L mutation and carried out confocal Ca2+ imaging. RESULTS: We found that mutations, S2030D, S2030G, S2030L, S2030V, and S2030W reduced the endoplasmic reticulum luminal Ca2+ level at which Ca2+ release terminates (the termination threshold), whereas S2030P and S2030R increased the termination threshold. S2030A and S2030T had no significant impact on release termination. Furthermore, CaM-wild-type increased, whereas Ca2+ binding deficient CaM mutant (CaM-M [a loss-of-function CaM mutation with all 4 EF-hand motifs mutated]), PKA, and Ca2+/CaMKII (CaM-dependent protein kinase II) reduced the termination threshold. The S2030L mutation abolished the actions of CaM-wild-type, CaM-M, and PKA, but not CaMKII, in Ca2+ release termination. Moreover, we showed that isoproterenol and CaM-M suppressed pacing-induced Ca2+ alternans and accelerated Ca2+ transient recovery in intact working hearts, whereas CaM-wild-type exerted an opposite effect. The impact of isoproterenol was partially and fully reversed by the PKA inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinoline-sulfonamide and the CaMKII inhibitor N-[2-[N-(4-chlorocinnamyl)-N-methylaminomethyl]phenyl]-N-(2-hydroxyethyl)-4-methoxybenzenesulfonamide individually and together, respectively. S2030L abolished the impact of CaM-wild-type, CaM-M, and N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinoline-sulfonamide-sensitive component, but not the N-[2-[N-(4-chlorocinnamyl)-N-methylaminomethyl]phenyl]-N-(2-hydroxyethyl)-4-methoxybenzenesulfonamide-sensitive component, of isoproterenol.
Assuntos
Canal de Liberação de Cálcio do Receptor de Rianodina , Serina , Camundongos , Animais , Humanos , Isoproterenol/farmacologia , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Serina/metabolismo , Serina/farmacologia , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Calmodulina/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Isoquinolinas/farmacologia , Sulfonamidas/farmacologia , Cálcio/metabolismo , Miócitos Cardíacos/metabolismo , Retículo Sarcoplasmático/metabolismoRESUMO
Mitigation of biofouling and the host's foreign body response (FBR) is a critical challenge with biomedical implants. The surface coating with various anti-fouling materials provides a solution to overcome it, but limited options in clinic and their potential immunogenicity drive the development of more alternative coating materials. Herein, inspired by liquid-liquid phase separation of intrinsically disordered proteins (IDPs) to form separated condensates in physiological conditions, we develop a new type of low-fouling biomaterial based on flexible IDP of FUS protein containing rich hydrophilic residues. A chemical structure-defined FUS IDP sequence tagged with a tetra-cysteine motif (IDPFUS) was engineered and applied for covalent immobilization on various surfaces to form a uniform layer of protein tangles, which boosted strong hydration on surfaces, as revealed by molecular dynamics simulation. The IDPFUS-coated surfaces displayed excellent performance in resisting adsorption of various proteins and adhesion of different cells, platelets, and bacteria. Moreover, the IDPFUS-coated implants largely mitigated the host's FBR compared with bare implants and particularly outperformed PEG-coated implants in reducing collagen encapsulation. Thus, this novel low-fouling and anti-FBR strategy provides a potential surface coating material for biomedical implants, which will also shed light on exploring similar applications of other IDP proteins.
Assuntos
Incrustação Biológica , Corpos Estranhos , Proteínas Intrinsicamente Desordenadas , Humanos , Incrustação Biológica/prevenção & controle , Interações Hidrofóbicas e Hidrofílicas , Propriedades de SuperfícieRESUMO
Polypeptides, a kind of very promising biomaterial, have shown a wide range of applications due to their excellent biocompatibility, easy accessibility, and structural variability. To synthesize polypeptides with desired functions, post-polymerization modification (PPM) plays an important role in introducing novel chemical structure on their side-chains. The key to PPM strategy is to develop highly selective and efficient reactions that can couple the additional functional moieties with pre-installed side-chain functionalities on polypeptides. In this minireview, classic PPM approaches and their recent progresses are summarized and categorized into five sections, including various reactions on unsaturated alkyl, oxygen-, nitrogen-, sulfur-containing and other special functional groups. In addition, we also highlight the applications of the resultant structure-diversified polypeptides in the field of biomaterial.
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Materiais Biocompatíveis , Peptídeos , Materiais Biocompatíveis/química , Peptídeos/química , PolimerizaçãoRESUMO
Objective: To clarify the pathogenesis of human atheroma, the origin of deposited lipids, the developmental mechanism of liponecrotic tissue, and the significance of the oxidation of phospholipids were investigated using mass spectrometry-aided imaging and immunohistochemistry.Atherosclerotic lesions in human coronary arteries were divided into 3 groups: pathologic intimal thickening with lipid pool, atheroma with lipid core, and atheroma with necrotic core. The lipid pool and lipid core were characterized by the deposition of extracellular lipids. The necrotic core comprised extracellular lipids and liponecrotic tissue. The proportion of cholesteryl linoleate in cholesteryl linoleate+cholesteryl oleate fraction in the extracellular lipid and liponecrotic regions differed significantly from that of the macrophage foam cell-dominant region, and the plasma-derived components (apolipoprotein B and fibrinogen) were localized in the regions. The liponecrotic region was devoid of elastic and collagen fibers and accompanied by macrophage infiltration in the surrounding tissue. Non-oxidized phospholipid (Non-OxPL), OxPL, and Mox macrophages were detected in the three lesions. In the atheroma with lipid core and atheroma with necrotic core, non-OxPL tended to localize in the superficial layer, whereas OxPL was distributed evenly. Mox macrophages were colocalized with OxPL epitopes.In human atherosclerosis, plasma-derived lipids accumulate to form the lipid pool of pathologic intimal thickening, lipid core of atheroma with lipid core, and necrotic core of atheroma with necrotic core. The liponecrotic tissue in the necrotic core appears to be developed by the loss of elastic and collagen fibers. Non-OxPL in the accumulated lipids is oxidized to form OxPL, which may contribute to the lesion development through Mox macrophages.
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Colesterol/análise , Doença da Artéria Coronariana/metabolismo , Doença da Artéria Coronariana/patologia , Vasos Coronários/química , Vasos Coronários/patologia , Imagem Molecular , Fosfolipídeos/análise , Placa Aterosclerótica , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Autofagia , Biópsia , Estudos de Casos e Controles , Colesterol/sangue , Doença da Artéria Coronariana/sangue , Feminino , Células Espumosas/química , Células Espumosas/patologia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Necrose , Neointima , Oxirredução , Fosfolipídeos/sangue , Valor Preditivo dos TestesRESUMO
Utilizing the interactions between tryptophan, methyl viologen and cucurbit[8]uril, we found that the distance between the targeted peptides/protein and the reactive peptide was shortened, which facilitated the Michael addition reaction between cysteine and dehydroalanine. The highest acceleration was observed on cysteines with suitable pKa and spatial location to tryptophan, suggesting that our system can be used for regioselective cysteine modification.
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Hidrocarbonetos Aromáticos com Pontes/química , Cisteína/química , Imidazóis/química , Estrutura Molecular , EstereoisomerismoRESUMO
[Figure: see text].
Assuntos
Aorta/efeitos dos fármacos , Doenças da Aorta/prevenção & controle , Aterosclerose/prevenção & controle , Colesterol/metabolismo , Proteínas de Choque Térmico/administração & dosagem , Chaperonas Moleculares/administração & dosagem , Pró-Proteína Convertase 9/metabolismo , Receptores de LDL/metabolismo , Vacinas Sintéticas/administração & dosagem , Idoso , Idoso de 80 Anos ou mais , Animais , Anticorpos/sangue , Aorta/enzimologia , Aorta/imunologia , Aorta/patologia , Doenças da Aorta/enzimologia , Doenças da Aorta/imunologia , Doenças da Aorta/patologia , Aterosclerose/enzimologia , Aterosclerose/imunologia , Aterosclerose/patologia , Estudos de Casos e Controles , Modelos Animais de Doenças , Feminino , Proteínas de Choque Térmico/imunologia , Proteínas de Choque Térmico/metabolismo , Células Hep G2 , Humanos , Imunogenicidade da Vacina , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Pessoa de Meia-Idade , Chaperonas Moleculares/imunologia , Chaperonas Moleculares/metabolismo , Placa Aterosclerótica , Receptores de LDL/genética , Vacinação , Vacinas Sintéticas/imunologiaRESUMO
Here, a new amorphous material composed of carbon-coated zinc cobaltate doped with manganese and nickel ZNMC@C (ZnNi0.5Mn0.5CoO4@C) with a spinel structure is proposed as a cathode material for use in aqueous zinc-ion batteries. This cathode material exhibits a high charge/discharge capacity with an initial capacity of about 160 mA h g-1 and its capacity retention rate remains at 60% after 500 cycles at 0.2 A g-1, which is higher than that of some reported spinel cathode materials. This superior electrochemical performance can be ascribed to the synergistic effect of the co-doping of manganese and nickel, which produces reversible multivalence redox transition activity (Co4+/Co3+, Ni4+/Ni3+/Ni2+, and Mn4+/Mn3+) that facilitates the insertion and migration of zinc ions and the existence of an outer amorphous carbon coating that effectively inhibits the dissolution of the cathode structure and stabilizes the cathode structure. In addition, the cycling mechanism of ZNMC@C was analyzed in detail through electrochemical measurements of the different cycling stages, including the kinetic behavior based on cyclic voltammetry and electrochemical impedance spectroscopic analysis and the reaction mechanism from X-ray photoelectron spectroscopy, ex situ X-ray diffractometry and ex situ scanning electron microscopy analysis. These research results suggest that the ZNMC@C composite material could be a competitive cathode material for Abs (aqueous rechargeable batteries).
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A novel STING agonist, CDGSF, ipsilaterally modified with phosphorothioate and fluorine, was synthesized. The phosphorothioate in CDGSF might be a site for covalent conjugation. Injection of CDGSF generated an immunogenic ("hot") tumor microenvironment to suppress melanoma, more efficiently than dithio CDG. In particular, immunization with SARS-CoV-2 spike protein using CDGSF as an adjuvant elicited an exceptionally high antibody titer and a robust T cell response, overcoming the drawbacks of aluminum hydroxide. These results highlighted the therapeutic potential of CDGSF for cancer immunotherapy and the adjuvant potential of the STING agonist in the SARS-CoV-2 vaccine for the first time.
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Adjuvantes Imunológicos/administração & dosagem , Vacinas contra COVID-19/administração & dosagem , COVID-19/prevenção & controle , Melanoma Experimental/tratamento farmacológico , Proteínas de Membrana/agonistas , Nucleotídeos Cíclicos/administração & dosagem , Neoplasias Cutâneas/tratamento farmacológico , Adjuvantes Imunológicos/síntese química , Hidróxido de Alumínio/administração & dosagem , Hidróxido de Alumínio/química , Animais , Anticorpos Antivirais/biossíntese , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/virologia , COVID-19/imunologia , COVID-19/virologia , Vacinas contra COVID-19/química , ELISPOT , Humanos , Imunoterapia/métodos , Interferon gama/biossíntese , Melanoma Experimental/imunologia , Melanoma Experimental/mortalidade , Melanoma Experimental/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Nucleotídeos Cíclicos/síntese química , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Glicoproteína da Espícula de Coronavírus/administração & dosagem , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Análise de Sobrevida , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/virologia , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Vacinação/métodosRESUMO
BACKGROUND: The unique hypervariable C-terminal region (HVR) of K-Ras4B, one of the most frequently mutated proteins in many powerful cancers, contains a C-terminal farnesylated and methylated Cys and a poly-lysine motif, which decides the association of K-Ras4B to the inner leaflet of plasma membrane for activating the downstream signaling activity. In our previous work, we inserted an additional Cys in K-Ras4B HVR peptide synthesis for NCL in the semi-synthesis of K-Ras4b protein, but it is not suitable for application in protein dimerization research. The recently developed selenocysteine (Sec, U) mediated native chemical ligation reaction followed by selective deselenization, which can help to broaden the scope of protein synthesis, requires the generation of the peptide fragment with an N-terminal Sec. OBJECTIVE: To synthesize K-Ras4B HVR peptide containing both N-terminal Sec and C-terminal farnesylated and methylated Cys to achieve traceless protein semi-synthesis. METHODS AND RESULTS: We have developed a facile synthesis approach for producing Boc-Sec)2-OH using economic Se powder, which can facilitate scaling up preparation of peptides containing Sec at the N-terminus. Furthermore, we synthesized K-Ras4B HVR peptide containing selenocystine by utilization of Boc-Sec)2-OH. Finally, we took K-Ras4B HVR peptide as an example to test the compatibility of farnesylation reaction at Cys with the N-terminal Sec)2, and the farnesyl group was successfully added to the thiol group of Cys.
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Cistina/análogos & derivados , Compostos Organosselênicos , Peptídeos , Prenilação de Proteína , Técnicas de Química Sintética , Cisteína/química , Cistina/síntese química , Cistina/química , Humanos , Modelos Moleculares , Compostos Organosselênicos/síntese química , Compostos Organosselênicos/química , Peptídeos/síntese química , Peptídeos/química , Proteínas Proto-Oncogênicas p21(ras)/químicaRESUMO
AIMS: The estrogen-inducible protein Heat Shock Protein 27 (HSP27) as well as anti-HSP27 antibodies are elevated in healthy subjects compared to cardiovascular disease patients. Vaccination of ApoE-/- mice with recombinant HSP25 (rHSP25, the murine ortholog), boosts anti- HSP25 levels and attenuates atherogenesis. As estrogens promote HSP27 synthesis, cellular release and blood levels, we hypothesize that menopause will result in loss of HSP27 atheroprotection. Hence, the rationale for this study is to compare the efficacy of rHSP25 vaccination vs. estradiol (E2) therapy for the prevention of post-menopausal atherogenesis. METHODS AND RESULTS: ApoE-/- mice subjected to ovariectomy (OVX) showed a 65 % increase atherosclerotic burden compared to sham mice after 5 weeks of a high fat diet. Relative to vaccination with rC1, a truncated HSP27 control peptide, atherogenesis was reduced by 5-weekly rHSP25 vaccinations (-43 %), a subcutaneous E2 slow release pellet (-52 %) or a combination thereof (-82 %). Plasma cholesterol levels declined in parallel with the reductions in atherogenesis, but relative to rC1/OVX mice plasma PCSK9 levels were 52 % higher in E2/OVX and 41 % lower in rHSP25/OVX mice (p < 0.0001 for both). Hepatic LDLR mRNA levels did not change with E2 treatment but increased markedly with rHSP25 vaccination. Conversely, hepatic PCSK9 mRNA increased 148 % with E2 treatment vs. rC1/OVX but did not change with rHSP25 vaccination. In human HepG2 hepatocytes E2 increased PCSK9 promoter activity 303 %, while the combination of [rHSP27 + PAb] decreased PCSK9 promoter activity by 64 %. CONCLUSION: The reduction in post-OVX atherogenesis and cholesterol levels with rHSP25 vaccination is associated with increased LDLR but not PCSK9 expression. Surprisingly, E2 therapy attenuates atherogenesis and cholesterol levels post-OVX without altering LDLR but increases PCSK9 expression and promoter activity. This is the first documentation of increased PCSK9 expression with E2 therapy and raises questions about balancing physiological estrogenic / PCSK9 homeostasis and targeting PCSK9 in women - are there effects beyond cholesterol?
Assuntos
Aterosclerose/prevenção & controle , Colesterol/sangue , Estradiol/administração & dosagem , Terapia de Reposição de Estrogênios , Proteínas de Choque Térmico HSP27/metabolismo , Proteínas de Choque Térmico/administração & dosagem , Fígado/efeitos dos fármacos , Chaperonas Moleculares/administração & dosagem , Pró-Proteína Convertase 9/metabolismo , Receptores de LDL/metabolismo , Vacinas/administração & dosagem , Animais , Aterosclerose/sangue , Aterosclerose/enzimologia , Aterosclerose/imunologia , Biomarcadores/sangue , Modelos Animais de Doenças , Regulação para Baixo , Implantes de Medicamento , Feminino , Proteínas de Choque Térmico/imunologia , Células Hep G2 , Humanos , Fígado/enzimologia , Menopausa , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Chaperonas Moleculares/imunologia , Ovariectomia , Pró-Proteína Convertase 9/genética , Receptores de LDL/genética , VacinaçãoRESUMO
We developed a new responsive peptide hydrogel FmocFFpSC(oNB)-PEG, which could achieve gel formation induced by calcium ions and sequential dissolution stimulated by light. It provides a potential delivery system for the efficient encapsulation of drugs and their controlled release in a spatial and temporal way.
Assuntos
Hidrogéis , PeptídeosRESUMO
BACKGROUND: Beta-tricalcium phosphate (ß-TCP) has been employed successfully as a synthetic graft material in maxillary sinus floor augmentation (MSFA) for placing dental implants. However, the lack of osteogenic and osteoinductive properties of this substitute invariably results in bone regeneration of low quality and quantity. The purpose of this study was to determine whether loading dentin matrix protein-1 (DMP1) gene-modified bone marrow mesenchymal stem cells (BMSCs) onto ß-TCP promoted bone regeneration and osteointegration of dental implants in MSFA of dogs. METHODS: BMSCs were transduced with a lentiviral vector overexpressing the DMP1 gene (Lenti-DMP1) and with a lentiviral vector overexpressing enhanced green fluorescent protein (Lenti-EGFP) in vitro and were loaded into ß-TCP scaffolds for autologous sinus grafting. Beagles received bilateral MSFA with four biomaterials (â Lenti-DMP1-transduced BMSCs/ß-TCP, â¡ Lenti-EGFP-transduced BMSCs/ß-TCP, ⢠BMSCs/ß-TCP, ⣠ß-TCP) and simultaneous implant placement at each sinus. Twelve weeks post operation, the maxillae were explanted, and every sinus was evaluated by radiographic observation, micro-CT and histological analysis. The osteogenic outcomes of bone regeneration and osseointegration were compared between the four groups. RESULTS: The sinuses grafted with Lenti-DMP1-transduced BMSCs/ß-TCP constructs presented a significantly higher increase in compact radiopaque area, higher local bone mineral densities, greater bone-implant contact and greater bone density when compared to other three groups. CONCLUSION: These results demonstrated that combinations of ß-TCP and DMP1 gene-modified BMSCs could be used to construct tissue-engineered bone to enhance mineralization of the regenerated bone and osseointegration of dental implants in MSFA.
Assuntos
Implantes Dentários , Levantamento do Assoalho do Seio Maxilar , Animais , Células da Medula Óssea , Dentina , Cães , Maxila/diagnóstico por imagem , Maxila/cirurgia , Seio Maxilar , OsseointegraçãoRESUMO
We developed a host-guest ATP responsive strategy for efficient intracellular delivery of phosphopeptides, employing a pegylated arginine clustered calix[5]arene nanocarrier system that has great capability of recognizing the phosphate moieties on peptides and penetrating the cell membrane.