RESUMO
The misfolding and un-natural fibrillation of proteins/peptides are associated with many conformation diseases, such as human islet amyloid polypeptide (hIAPP) in type 2 diabetes (T2D). Inspired by molecular chaperones maintaining protein homeostasis in vivo, many polymer-based artificial chaperones were introduced to regulate protein/peptide folding and fibrillation. However, the pure polymer chaperones prefer to agglomerate into large-size micelles in the physiological environment and thus lose their chaperone functions, which greatly restricts the application of polymer-based chaperones. Here, we designed and prepared a core-shell artificial chaperone based on a dozen poly-(N-isopropylacrylamide-co-N-acryloyl-O-methylated-l-arginine) (PNAMR) anchored on a gold-nanocluster (AuNC) core. The introduction of the AuNC core significantly reduced the size and enhanced the efficacy and stability of polymer-based artificial chaperones. The PNAMR@AuNCs, with a diameter of 2.5 ± 0.5 nm, demonstrated exceptional ability in maintaining the natively unfolded conformation of protein away from the misfolding and the following fibrillation by directly binding to the natively unfolded monomolecular hIAPP and hence in preventing their conversion into toxic oligomers. More excitingly, the PNAMR@AuNCs were able to restore the natural unfolded conformation of hIAPP via dissolving the ß-sheet-rich hIAPP fibrils. Considering the uniform molecular mechanism of protein misfolding and fibrillation in conformation disorders, this finding provides a generic therapeutic strategy for neurodegenerative diseases and other conformation diseases by using PNAMR@AuNC artificial chaperones to restore and maintain the native conformation of amyloid proteins.
Assuntos
Diabetes Mellitus Tipo 2 , Humanos , Diabetes Mellitus Tipo 2/metabolismo , Polipeptídeo Amiloide das Ilhotas Pancreáticas/química , Polímeros/farmacologia , Chaperonas Moleculares , Conformação Proteica , Amiloide/químicaRESUMO
The misfolding, aggregation and fibrillation of human islet amyloid polypeptide (hIAPP) has been acknowledged as a hallmark event in type-II diabetes. Hence, inhibiting the misfolding, aggregation and fibrillation of hIAPP have been accepted as a vital factor to treat the disease. Here cichoric acid was extracted from witloof to explore its inhibition effects on misfolding, aggregation and fibrillation of hIAPP. Thioflavin-T (ThT) fluorescence assay, dynamic light scattering (DLS) and atomic force microscopy (AFM) images showed that cichoric acid inhibited the aggregation and fibrillation of hIAPP in a dosage-dependent manner. Circular dichroism (CD) spectra showed that cichoric acid inhibited the misfolding of hIAPP from unfolded to ß-sheet. Molecular docking and further experiments revealed interactions between hIAPP and cichoric acid. Cichoric acid could bind to K1 and R11 of hIAPP via electrostatic interaction. In addition, cichoric acid could form π-π stacking with hIAPP residues F15 and F23. These interactions inhibited the misfolding, aggregation and fibrillation of hIAPP. These results, together with cichoric acid's good cytocompatibility and significant protective effects in hIAPP lesioned cell models, not only showed that cichoric acid could be used to fight against amyloidosis, but also brought a new perspective for Chinese herbal medicine as natural compound's medical potential.
Assuntos
Amiloide/química , Ácidos Cafeicos/química , Cichorium intybus/química , Polipeptídeo Amiloide das Ilhotas Pancreáticas/química , Succinatos/química , Amiloide/antagonistas & inibidores , Humanos , Polipeptídeo Amiloide das Ilhotas Pancreáticas/antagonistas & inibidores , Polipeptídeo Amiloide das Ilhotas Pancreáticas/metabolismo , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Agregação Patológica de Proteínas , Dobramento de Proteína , Análise EspectralRESUMO
Protein/Peptide amyloidosis is the main cause of several diseases, such as neurodegenerative diseases. It has been widely acknowledged that the unnatural fibrillation of protein/peptides in vivo is significantly affected by the physical and chemical properties of multiscale biological membranes. For example, previous studies have proved that molecule chirality could greatly influence the misfolding, fibrillation and assembly of ß-Amyloid peptides at the flat liquid-solid surface. However, how the nanoscale chirality influences this process remains unclear. Here we used gold nanoparticles (AuNPs, d = 4 ± 1 nm)-modified with N-isobutyl-L(D)-cysteine (L(D)-NIBC) enantiomers-as a model to illustrate the chiral effect on the amylin fibrillation at nano-bio interface. We reported that both two chiral AuNPs could inhibit amylin fibrillation in a dosage-dependent manner but the inhibitory effect of L-NIBC-AuNPs was more effective than that of D-NIBC-AuNPs. In-situ real time circular dichroism (CD) spectra showed that L-NIBC-AuNPs could inhibit the conformation transition process of amylin from random coils to α-helix, while D-NIBC-AuNPs could only delay but not prevent the formation of α-helix; however, they could inhibit the further conformation transition process of amylin from α-helix to ß-sheet. These results not only provide interesting insight for reconsidering the mechanism of peptides amyloidosis at the chiral interfaces provided by biological nanostructures in vivo but also would help us design therapeutic inhibitors for anti-amyloidosis targeting diverse neurodegenerative diseases.
RESUMO
ß-HgS quantum dots (QDs) have drawn enormous attention due to the size-tunable bandgap and the lowest quantum state in conduction band which have been applied to semiconductor transistor and photodetector. Though ß-HgS is the essential component of Tibetan medicine, the potential toxicity of ß-HgS limits its applications, especially in bio-application. Herein, chiral biomolecule enantiomers N-isobutyryl-L(D)-cysteine (L(D)-NIBC) and L(D)-cysteine (L(D)-Cys) were introduced into HgCl2 and Na2S aqueous solution to synthesize chiral ß-HgS QDs in one-pot, which significantly improved their water-solubility and cytocompatibility. Notably, all chiral ß-HgS QDs showed none cytotoxicity even at high concentration (20â¯mg·L-1), and the cytocompatibility of D-ß-HgS QDs was better than corresponding L-ß-HgS QDs at the concentration of 20â¯mg·L-1. This cytotoxicity discrimination was associated with the chirality inversion of chiral ß-HgS QDs compared with the corresponding chiral ligands. In-situ real-time circular dichroism (CD) monitoring indicated that the chirality of ß-HgS QDs originated from the asymmetrical arrangement of chiral ligands on the achiral core surface. Their chiroptical activity, near-infrared optical absorption (800â¯nm), fluorescence emission (900-1000â¯nm), high-performance photothermal conversion and good cytocompatibility, implied chiral ß-HgS QDs could be used as a candidate material for photothermal therapy or a near-infrared fluorescent probe in organism, which brings a novel insight for bio-application of ß-HgS QDs.
Assuntos
Compostos de Mercúrio/síntese química , Pontos Quânticos/química , Sulfetos/química , Água/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Compostos de Mercúrio/química , Compostos de Mercúrio/farmacologia , Fenômenos Ópticos , Tamanho da Partícula , Relação Estrutura-Atividade , Sulfetos/farmacologia , Propriedades de SuperfícieRESUMO
Cyclin dependent kinase 4 (Cdk4) is a cell cycle regulator involved in early G1 cell cycle progression and has been indirectly implicated in angiogenesis in the Min mouse system, a mouse that harbors a mutation in the Apc gene. Apc(+/Min) mice when crossed with Ink4a/arf-/- mice, exhibited increased angiogenesis of colorectal tumors suggesting that dysregulation of Cdk4 (due to loss of Ink4a-mediated suppression) may contribute to enhanced angiogenesis. To demonstrate a direct role for Cdk4 in angiogenesis, we crossed mice that have an activated Cdk4, Cdk4(R24C/R24C) mice, with Apc(+/Min) mice and examined levels of angiogenesis in intestinal tumors formed. Our results show an increase in the percentage of highly vascularized tumors in Cdk4(R24C/R24C):Apc(Min/+) and Cdk4(+/R24C):Apc(Min/+) mice compared to Cdk4(+/+):Apc(Min/+) mice. In addition immunohistochemical analysis showed an increase in CD-31 staining localized to endothelial cells of Cdk4(R24C/R24C):Apc(Min/+) mouse tumors, supporting the hypothesis of increased vasculature in these tumors. Further analysis showed an increase in the expression of the E2F1 target proteins Vegf-b and Cyclin A in Cdk4(R24C/R24C):Apc(+/Min) intestinal tumors. Together these data suggest that the dysregulated Cdk4 gene plays an important role in angiogenesis during intestinal tumor formation and may in part act via increasing E2F1 target proteins. This is the first report to show that Cdk4 has a direct role in angiogenesis in vivo and may be an important drug target to reduce or prevent angiogenesis during intestinal tumor formation.
Assuntos
Quinase 4 Dependente de Ciclina/metabolismo , Genes APC , Neoplasias Intestinais/irrigação sanguínea , Neoplasias Intestinais/metabolismo , Neovascularização Patológica , Animais , Ciclo Celular , Ciclina A/biossíntese , Quinase 4 Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/deficiência , Inibidor p16 de Quinase Dependente de Ciclina/genética , Fator de Transcrição E2F1/biossíntese , Células Endoteliais/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Intestinais/genética , Intestinos/irrigação sanguínea , Camundongos , Mutação , Molécula-1 de Adesão Celular Endotelial a Plaquetas , Fator B de Crescimento do Endotélio Vascular/biossíntese , Fator B de Crescimento do Endotélio Vascular/genéticaRESUMO
Chronic graft-vs-host (cGVH) disease is induced in nonautoimmune mice by the transfer of alloreactive T cells that recognize foreign MHC class II. It closely resembles systemic lupus erythematosus, with antinuclear Abs and immune-mediated nephritis. Recent work has implicated TLRs, particularly TLR9, in the recognition of certain autoantigens in vitro and in vivo. To explore further the role of TLR9 in systemic autoimmunity, we induced cGVH disease in C57BL/6 (B6) mice lacking TLR9, including B6 mice expressing the anti-DNA-encoding IgH transgenes 3H9 or 56R (B6.3H9.TLR9(-/-), B6.56R.TLR9(-/-)). We found that cGVH disease caused breakdown of B cell tolerance to chromatin and DNA in TLR9(-/-) recipients of alloreactive cells, yet that nephritis was less severe and that some autoantibody titers were lower compared with B6-cGVH controls. Spleen lymphocyte analysis showed that cGVH disease strikingly depleted marginal zone B cells in B6 mice, but did not influence T cell subsets in either B6 or B6-TLR9(-/-) hosts. B6.56R.TLR9(-/-) mice had less spontaneous production of autoantibodies than B6.56R mice, but there were no significant differences between B6.56R and B6.56R.TLR9(-/-) postinduction of cGVH disease. Taken together, these results suggested that TLR9 may worsen some aspects of systemic autoimmunity while alleviating others.