RESUMEN

As part of our ongoing research on new anti-diabetic compounds from ethnopharmacologically consumed plants, two previously undescribed lupane-type triterpenoids (1 and 2) with dicarboxylic groups, an undescribed nor-taraxastane-type triterpenoid (3), and 14 known compounds (4-17) were isolated from the leaves of Cleistocalyx operculatus. Extensive spectroscopic analysis (IR, HRESIMS, 1D, and 2D NMR) was used for structure elucidation, while the known compounds were compared to reference data reported in the scientific literature. All the isolates (1-17) were evaluated for their inhibitory effects on the protein tyrosine phosphatase 1B (PTP1B) enzyme. Compounds 6, 9, and 17 showed strong PTP1B inhibitory activities. The mechanism of PTP1B inhibition was studied through enzyme kinetic experiments. A non-competitive mechanism of inhibition was determined using Lineweaver-Burk plots for compounds 6, 9, and 17. Additionally, Dixon plots were employed to determine the inhibition constant. Further insights were gained through a structure-activity relationship study and molecular docking analysis of isolated compounds with the PTP1B crystal structure. Moreover, all isolates (1-17) were tested for their stimulatory effects on the uptake of 2-deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl) amino]-D-glucose (2-NBDG) in differentiated 3T3-L1 adipocyte cells. Compounds 6, 13, and 17 exhibited strong glucose absorption stimulation activity in a dose-dependent manner.

Asunto(s)

Células 3T3-L1 , Glucosa , Simulación del Acoplamiento Molecular , Hojas de la Planta , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Hojas de la Planta/química , Ratones , Animales , Glucosa/metabolismo , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/aislamiento & purificación , Extractos Vegetales/farmacología , Extractos Vegetales/química , Syzygium/química , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Relación Estructura-Actividad , Simulación por Computador

RESUMEN

Fluxes in human copper levels recently garnered attention for roles in cellular signaling, including affecting levels of the signaling molecule cyclic adenosine monophosphate. We herein apply an unbiased temporal evaluation of the signaling and whole genome transcriptional activities modulated by copper level fluctuations to identify potential copper sensor proteins responsible for driving these activities. We find that fluctuations in physiologically relevant copper levels modulate EGFR signal transduction and activation of the transcription factor CREB. Both intracellular and extracellular assays support Cu1+ inhibition of the EGFR phosphatase PTPN2 (and potentially PTPN1)-via ligation to the PTPN2 active site cysteine side chain-as the underlying mechanism. We additionally show i) copper supplementation drives weak transcriptional repression of the copper importer CTR1 and ii) CREB activity is inversely correlated with CTR1 expression. In summary, our study reveals PTPN2 as a physiological copper sensor and defines a regulatory mechanism linking feedback control of copper stimulated EGFR/CREB signaling and CTR1 expression.

Asunto(s)

Transportador de Cobre 1 , Cobre , Proteína de Unión a Elemento de Respuesta al AMP Cíclico , Receptores ErbB , Proteína Tirosina Fosfatasa no Receptora Tipo 2 , Transducción de Señal , Receptores ErbB/metabolismo , Receptores ErbB/genética , Cobre/metabolismo , Humanos , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Transportador de Cobre 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 2/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 2/genética , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Transcripción Genética/efectos de los fármacos

RESUMEN

Protein tyrosine phosphatase non-receptor type 2 (PTPN2) has recently been recognized as a promising target for cancer immunotherapy. Despite extensive structural and functional studies of other protein tyrosine phosphatases, there is limited structural understanding of PTPN2. Currently, there are only five published PTPN2 structures and none are truly unbound due to the presence of a mutation, an inhibitor or a loop (related to crystal packing) in the active site. In this report, a novel crystal packing is revealed that resulted in a true apo PTPN2 crystal structure with an unbound active site, allowing the active site to be observed in a native apo state for the first time. Key residues related to accommodation in the active site became identifiable upon comparison with previously published PTPN2 structures. Structures of PTPN2 in complex with an established PTPN1 active-site inhibitor and an allosteric inhibitor were achieved through soaking experiments using these apo PTPN2 crystals. The increased structural understanding of apo PTPN2 and the ability to soak in inhibitors will aid the development of future PTPN2 inhibitors.

Asunto(s)

Dominio Catalítico , Proteína Tirosina Fosfatasa no Receptora Tipo 2 , Proteína Tirosina Fosfatasa no Receptora Tipo 2/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 2/química , Proteína Tirosina Fosfatasa no Receptora Tipo 2/genética , Humanos , Cristalografía por Rayos X , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Sitios de Unión , Modelos Moleculares , Unión Proteica , Proteína Tirosina Fosfatasa no Receptora Tipo 1/química , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Cristalización , Apoenzimas/química , Apoenzimas/metabolismo , Apoenzimas/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo

RESUMEN

BACKGROUND: Formononetin (FNT) is an isoflavone known for its anti-inflammatory properties and has been shown to reduce insulin resistance in Type 2 Diabetes Mellitus (T2DM). However, its effects and the underlying mechanisms in diabetic liver injury remain largely unexplored. METHODS: We established a T2DM-induced liver injury mouse model by feeding high-fat diet, followed by injecting streptozotocin. The mice were then treated with FNT and the liver function in these mice was assessed. Macrophage markers in FNT-treated T2DM mice or human THP-1 cells were evaluated using flow cytometry, RT-qPCR, and Western blotting. The expression of PTP1B and STAT6 in mouse liver tissues and THP-1 cells was analyzed. Molecular docking predicted the interaction between PTP1B and STAT6, which was validated via co-immunoprecipitation (Co-IP) and phos-tag analysis. Microscale thermophoresis (MST) assessed the binding affinity of FNT to PTP1B. RESULTS: FNT treatment significantly ameliorated blood glucose levels, hepatocyte apoptosis, inflammatory response, and liver dysfunction in T2DM mice. Moreover, FNT facilitated M2 macrophage polarization in both T2DM mice and high glucose (HG)-induced THP-1-derived macrophages. The PTP1B/STAT6 axis, deregulated in T2DM mice, was normalized by FNT treatment, which counteracted the T2DM-induced upregulation of PTP1B and downregulation of phosphorylated STAT6. Molecular docking and subsequent analyses revealed that PTP1B binds to and dephosphorylates STAT6 at the S325A site. In contrast, FNT strongly binds to PTP1B and influences its expression at the K116A site, promoting M2 polarization of THP-1 cells via downregulation of PTP1B. CONCLUSION: Formononetin mitigates diabetic hepatic injury by fostering M2 macrophage polarization via the PTP1B/STAT6 axis.

Asunto(s)

Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Isoflavonas , Macrófagos , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Animales , Humanos , Masculino , Ratones , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Dieta Alta en Grasa , Isoflavonas/farmacología , Isoflavonas/uso terapéutico , Hígado/efectos de los fármacos , Hígado/patología , Hígado/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Simulación del Acoplamiento Molecular , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Transducción de Señal/efectos de los fármacos , Factor de Transcripción STAT6/metabolismo , Estreptozocina , Células THP-1

RESUMEN

Uvaol (UV), a pentacyclic triterpene found in olives and virgin olive oil, is known for its anti-inflammatory and antioxidant effects in various disease models. While olive oil is reported to reduce obesity and insulin resistance, the specific impact of UV on liver lipid metabolism and its molecular mechanisms are not fully understood. In this study, hepatic lipid accumulation was measured using oil red O staining, and protein expression levels in liver cells were assessed via Western blot analysis. Apoptosis was evaluated through cell viability and caspase 3 activity assays. UV treatment reduced lipid accumulation, fatty acid uptake, apoptosis, and ER stress in palmitate-treated liver cells. Additionally, UV enhanced fatty acid oxidation. Mechanistically, increased SIRT6 expression and autophagy were observed in UV-treated cells. SIRT6-targeted siRNA or 3-methyladenine blocked the effects of UV in hyperlipidemic cells. In conclusion, UV improves SIRT6/autophagy signaling, reducing lipid deposition and apoptosis in liver cells under high lipid conditions. This in vitro study provides strong evidence for potential therapeutic strategies for hepatic steatosis.

Asunto(s)

Apoptosis , Estrés del Retículo Endoplásmico , Hepatocitos , Hiperlipidemias , Metabolismo de los Lípidos , Transducción de Señal , Sirtuinas , Hepatocitos/metabolismo , Hepatocitos/efectos de los fármacos , Hepatocitos/efectos de la radiación , Metabolismo de los Lípidos/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Hiperlipidemias/metabolismo , Hiperlipidemias/tratamiento farmacológico , Sirtuinas/metabolismo , Sirtuinas/genética , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Autofagia/efectos de los fármacos , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Humanos , Animales , Triterpenos Pentacíclicos/farmacología

RESUMEN

Four new compounds, racemic chalcone-monoterpene hybrids (1-3) and a chalcone (9), along with nine known compounds (4-8, 10-13), have been isolated from the buds of Cleistocalyx operculatus. The chemical structures of the isolated compounds were identified through NMR data analysis and confirmed by computational methods, including electronic circular dichroism (ECD) calculations, and further synthetic approaches. Compounds 1-5 were synthesized via a Diels-Alder reaction, a process informed by biomimetic condensation studies that combined chalcones and monoterpenes. These synthetic approaches also yielded various unnatural chalcone-monoterpene derivatives (14-23). The inhibitory effects on protein tyrosine phosphatase 1B (PTP1B) of both naturally isolated and synthetically obtained compounds were evaluated. Compounds 4, 9, 13, and 16b exhibited potent PTP1B inhibitory activity, with IC50 values ranging from 0.9 ± 0.2 to 3.9 ± 0.7 µM. The enantiomers (+)-4 and (-)-16b showed enhanced activity compared to their respective enantiomers. Kinetic studies indicate that all active compounds inhibit PTP1B through mixed mechanisms, and molecular docking simulations agree with the experimental assays on PTP1B. Our results suggest that chalcone-meroterpene adducts from the buds of C. operculatus exhibit potential as antidiabetic agents, partly due to their PTP1B enzyme inhibition.

Asunto(s)

Monoterpenos , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Estructura Molecular , Monoterpenos/farmacología , Monoterpenos/química , Monoterpenos/aislamiento & purificación , Chalconas/farmacología , Chalconas/química , Chalconas/aislamiento & purificación , Chalcona/farmacología , Chalcona/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Humanos , Syzygium/química

RESUMEN

Although tumor cell-derived microparticles (MPs) vaccines have reportedly induced antitumor immune reactions for various cancers, the mechanism by which MPs derived from Hepa1-6 cells are taken up by dendritic cells (DCs) and provide the MPs antigens message to CD8+ T cells to exert their anti-hepatocellular carcinoma (HCC) effects remain unclear. Furthermore, the role of MPs in combination with the small-molecule drug MSI-1436, an inhibitor of protein tyrosine phosphatase 1B (PTP1B), in HCC has not yet been reported. In this study, protein mass spectrometry combined with cytology revealed that MPs are mainly taken up by DCs via the clathrin-mediated endocytosis and phagocytosis pathway and localized mainly in lysosomes. High concentration of tumor necrosis factor-α and interferon-γ was detected in CD8+ T cells stimulated with MPs-loaded DCs. Moreover, MPs combined with MSI-1436 further suppressed the proliferation of HCC cells in C57BL/6 tumor-bearing mice, which was closely correlated with CD4+/CD8+ T cells counts in peripheral blood, spleen, and the tumor microenvironment. Mechanistically, the combination of MPs and MSI-1436 exerts a more powerful anti-HCC effect, which may be related to the further inhibition of the expression of PTP1B. Overall, MPs combined with MSI-1436 exerted stronger antitumor effects than MPs or MSI-1436 alone. Therefore, the combination of MPs and MSI-1436 may be a promising means of treating HCC.

Asunto(s)

Linfocitos T CD8-positivos , Vacunas contra el Cáncer , Carcinoma Hepatocelular , Micropartículas Derivadas de Células , Células Dendríticas , Neoplasias Hepáticas , Ratones Endogámicos C57BL , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Animales , Carcinoma Hepatocelular/inmunología , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/terapia , Vacunas contra el Cáncer/inmunología , Neoplasias Hepáticas/inmunología , Neoplasias Hepáticas/patología , Células Dendríticas/inmunología , Células Dendríticas/efectos de los fármacos , Micropartículas Derivadas de Células/inmunología , Ratones , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/efectos de los fármacos , Línea Celular Tumoral , Humanos , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/inmunología , Proliferación Celular/efectos de los fármacos

RESUMEN

Pueraria lobata (Willd.) Ohwi, known as kudzu and used as a "longevity powder" in China, is an edible plant which is rich in flavonoids and believed to be useful for regulating blood sugar and treating diabetes, although the modes of action are unknown. Here, a total of 53 flavonoids including 6 novel compounds were isolated from kudzu using multidimensional preparative liquid chromatography. The flavonoid components were found to lower blood sugar levels, promote urine sugar levels in mice, and reduce the urine volume. Molecular docking and in vitro assays suggested that the antidiabetic effect of kudzu was attributed to at least three targets: sodium-dependent glucose transporter 2 (SGLT2), protein tyrosine phosphatase-1B (PTP1B), and alpha-glucosidase (AG). This study suggests a possible mechanism for the antidiabetic effect that may involve the synergistic action of multiple active compounds from kudzu.

Asunto(s)

Flavonoides , Hipoglucemiantes , Extractos Vegetales , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Pueraria , Pueraria/química , Hipoglucemiantes/química , Hipoglucemiantes/farmacología , Flavonoides/química , Animales , Ratones , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Humanos , Extractos Vegetales/química , Extractos Vegetales/farmacología , Simulación del Acoplamiento Molecular , Masculino , alfa-Glucosidasas/metabolismo , alfa-Glucosidasas/química , Glucemia/metabolismo , Plantas Comestibles/química

RESUMEN

Overexpression of protein tyrosine phosphatase 1B (PTP1B) disrupts signaling pathways and results in numerous human diseases. In particular, its involvement has been well documented in the pathogenesis of metabolic disorders (diabetes mellitus type I and type II, fatty liver disease, and obesity); neurodegenerative diseases (Alzheimer's disease, Parkinson's disease); major depressive disorder; calcific aortic valve disease; as well as several cancer types. Given this multitude of therapeutic applications, shortly after identification of PTP1B and its role, the pursuit to introduce safe and selective enzyme inhibitors began. Regrettably, efforts undertaken so far have proved unsuccessful, since all proposed PTP1B inhibitors failed, or are yet to complete, clinical trials. Intending to aid introduction of the new generation of PTP1B inhibitors, this work collects and organizes the current state of the art. In particular, this review intends to elucidate intricate relations between numerous diseases associated with the overexpression of PTP1B, as we believe that it is of the utmost significance to establish and follow a brand-new holistic approach in the treatment of interconnected conditions. With this in mind, this comprehensive review aims to validate the PTP1B enzyme as a promising molecular target, and to reinforce future research in this direction.

Asunto(s)

Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Humanos , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Neoplasias/metabolismo , Neoplasias/enzimología , Neoplasias/genética , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/enzimología , Inhibidores Enzimáticos/uso terapéutico , Inhibidores Enzimáticos/farmacología , Enfermedades Metabólicas/metabolismo , Enfermedades Metabólicas/enzimología , Animales , Transducción de Señal

RESUMEN

Protein tyrosine phosphatase 1B (PTP1B) is a non-receptor tyrosine phosphatase best known for its role in regulating insulin and leptin signalling. Recently, knowledge on the role of PTP1B as a major regulator of multiple signalling pathways involved in cell growth, proliferation, viability and metabolism has expanded, and PTP1B is recognised as a therapeutic target in several human disorders, including diabetes, obesity, cardiovascular diseases and hematopoietic malignancies. The function of PTP1B in the immune system was largely overlooked until it was discovered that PTP1B negatively regulates the Janus kinase-a signal transducer and activator of the transcription (JAK/STAT) signalling pathway, which plays a significant role in modulating immune responses. PTP1B is now known to determine the magnitude of many signalling pathways that drive immune cell activation and function. As such, PTP1B inhibitors are being developed and tested in the context of inflammation and autoimmune diseases. Here, we provide an up-to-date summary of the molecular role of PTP1B in regulating immune cell function and how targeting its expression and/or activity has the potential to change the outcomes of immune-mediated and inflammatory disorders.

Asunto(s)

Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Transducción de Señal , Humanos , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Animales , Inflamación/metabolismo , Inflamación/inmunología , Diabetes Mellitus/inmunología , Diabetes Mellitus/metabolismo

RESUMEN

During the search for protein tyrosine phosphatase 1B (PTP1B) inhibitory compounds from the natural resources, two new serratane triterpenes, 3-O-dihydro-p-coumaroyltohogenol (1) and 21-O-acetyltohogenol (2), along with four known serratane triterpenes (3-6), were isolated from the whole plant of Huperzia serrata. The chemical structures of compounds 1 and 2 were determined by NMR study, HRMS analysis, and chemical modification. All isolates were evaluated for their PTP1B inhibitory activities. Among the isolates, compounds 1, 3, 5 and 6 exhibit moderate inhibitory activities against PTP1B. Kinetic studies demonstrated that they are competitive inhibitors. Molecular docking studies support these experimental results by showing that compounds 1, 3, 5 and 6 interact with the active site of PTP1B, clarifying the structure-activity relationship. This study suggests that serratane triterpenes from H. serrata have potential as starting skeletons for anti-diabetes or anti-obesity agents.

Asunto(s)

Inhibidores Enzimáticos , Simulación del Acoplamiento Molecular , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Triterpenos , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Triterpenos/química , Triterpenos/farmacología , Triterpenos/aislamiento & purificación , Relación Estructura-Actividad , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/aislamiento & purificación , Humanos , Estructura Molecular , Relación Dosis-Respuesta a Droga

RESUMEN

Due to their low binding affinities, detecting small-molecule fragments bound to protein structures from crystallographic datasets has been a challenge. Here, we report a trove of 65 new fragment hits for PTP1B, an "undruggable" therapeutic target enzyme for diabetes and cancer. These structures were obtained from computational analysis of data from a large crystallographic screen, demonstrating the power of this approach to elucidate many (∼50% more) "hidden" ligand-bound states of proteins. Our new structures include a fragment hit found in a novel binding site in PTP1B with a unique location relative to the active site, one that links adjacent allosteric sites, and, perhaps most strikingly, a fragment that induces long-range allosteric protein conformational responses. Altogether, our research highlights the utility of computational analysis of crystallographic data, makes publicly available dozens of new ligand-bound structures of a high-value drug target, and identifies novel aspects of ligandability and allostery in PTP1B.

Asunto(s)

Sitio Alostérico , Unión Proteica , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Proteína Tirosina Fosfatasa no Receptora Tipo 1/química , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Cristalografía por Rayos X , Humanos , Ligandos , Dominio Catalítico , Modelos Moleculares , Regulación Alostérica , Sitios de Unión , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Conformación Proteica

RESUMEN

Astragalus kurdicus Boiss. roots are used in folk medicine for antidiabetic purposes. Different Astragalus plant metabolites have a notable potential for antidiabetic activity through varying mechanisms. Herein, this study is designed to assess the antidiabetic activity of Astragalus kurdicus total (AKM: methanol extract, yield: 14.53 %) and sub-extracts (AKB: n-butanol, AKC: chloroform, AKW: water, AKH: hexane extracts), utilizing a range of diabetes-related in vitro methodologies, and to investigate the chemical composition of the plant. The highest astragaloside and saponin content was seen in AKB extract. Among the measured saponins, the abundance of Astragaloside IV (27.41 µg/mg in AKM) was the highest in high-performance thin-layer chromatography (HPTLC) analysis. Furthermore, flavonoid-rich AKC was found to be mostly responsible for the high antioxidant activity. According to the results of the activity tests, AKW was the most active extract in protein tyrosine phosphatase 1 B (PTP1B), dipeptidyl peptidase IV (DPP4), and α-amylase inhibition tests (percent inhibitions are: 87.17 %, 82.4 %, and 91.49 % respectively, at 1 mg/mL). AKM and AKW demonstrated the highest efficacy in stimulating the growth of prebiotic microorganisms and preventing the formation of advanced glycation end products (AGEs). Thus, for the first time, the antidiabetic activity of A. kurdicus was evaluated from various perspectives.

Asunto(s)

Planta del Astrágalo , Hipoglucemiantes , Extractos Vegetales , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Hipoglucemiantes/aislamiento & purificación , Planta del Astrágalo/química , Extractos Vegetales/química , Extractos Vegetales/farmacología , Extractos Vegetales/aislamiento & purificación , Saponinas/farmacología , Saponinas/química , Saponinas/aislamiento & purificación , Antioxidantes/farmacología , Antioxidantes/química , Antioxidantes/aislamiento & purificación , Humanos , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , alfa-Amilasas/antagonistas & inhibidores , alfa-Amilasas/metabolismo , Triterpenos/química , Triterpenos/farmacología , Triterpenos/aislamiento & purificación , Dipeptidil Peptidasa 4/metabolismo , Dipeptidil Peptidasa 4/química , Fitoquímicos/farmacología , Fitoquímicos/química , Fitoquímicos/aislamiento & purificación

RESUMEN

Directed cell migration is driven by the front-back polarization of intracellular signalling1-3. Receptor tyrosine kinases and other inputs activate local signals that trigger membrane protrusions at the front2,4-6. Equally important is a long-range inhibitory mechanism that suppresses signalling at the back to prevent the formation of multiple fronts7-9. However, the identity of this mechanism is unknown. Here we report that endoplasmic reticulum-plasma membrane (ER-PM) contact sites are polarized in single and collectively migrating cells. The increased density of these ER-PM contacts at the back provides the ER-resident PTP1B phosphatase more access to PM substrates, which confines receptor signalling to the front and directs cell migration. Polarization of the ER-PM contacts is due to microtubule-regulated polarization of the ER, with more RTN4-rich curved ER at the front and more CLIMP63-rich flattened ER at the back. The resulting ER curvature gradient leads to small and unstable ER-PM contacts only at the front. These contacts flow backwards and grow to large and stable contacts at the back to form the front-back ER-PM contact gradient. Together, our study suggests that the structural polarity mediated by ER-PM contact gradients polarizes cell signalling, directs cell migration and prolongs cell migration.

Asunto(s)

Membrana Celular , Movimiento Celular , Retículo Endoplásmico , Humanos , Línea Celular , Membrana Celular/metabolismo , Polaridad Celular , Retículo Endoplásmico/metabolismo , Microtúbulos/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Transducción de Señal

RESUMEN

BACKGROUND: Spilanthes filicaulis (Schumach. & Thonn.) C. D Adam is a shrubby plant of the Asteraceae family that has medicinal benefits for the pharmaceutical and cosmetic industries. PURPOSE: The purpose of this study was to assess the effectiveness of Spilanthes filicaulis leaf extract in a streptozotocin (STZ)-induced rat model and the associated signaling pathways. METHODS: A sample of 25 male Wistar rats was randomly assigned to groups I, II, III, IV, and V. Each group included five animals, i.e., control rats, diabetic control rats, diabetic rats treated with metformin, and diabetic rats treated with 150 mg/kg/bw and 300 mg/kg/bw of the methanolic extract of S. filicaulis leaves (MESFL). Treatment was administered for 15 successive days via oral gavage. After 15 days, the rats were evaluated for fasting blood glucose (FBG), glycated hemoglobin (HbA1c), alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), triglyceride (TG), total cholesterol (TC), low-density lipoprotein (LDL), high-density lipoprotein (HDL), reduced glutathione (GSH), glutathione-S-transferase (GST), superoxide dismutase (SOD), catalase (CAT), lipid peroxidation (MDA), hexokinase, and glucose-6-phosphatase activities. Gene expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2), peroxisome proliferator-activated receptor gamma (PPAR-γ), kelch-like ECH-associated protein 1 (Keap1), protein tyrosine phosphatase 1B (PTP1B) and the antiapoptotic protein caspase-3 were examined. RESULTS: MESFL was administered to diabetic rats, and changes in body weight, fasting blood glucose (FBG) and HbA1c were restored. Furthermore, in diabetic rats, S. filicaulis significantly reduced the levels of triglycerides (TGs), total cholesterol (TC), low-density lipoprotein (LDL), and very low-density lipoprotein (VLDL) and significantly increased HDL. S. filicaulis improved ALT, AST, and ALP enzyme activity in diabetic rats. MDA levels decreased considerably with increasing activity of antioxidant enzymes, such as GST, SOD, CAT and GSH, in diabetic liver rats treated with S. filicaulis. Diabetic rats treated with MESFL and metformin exhibited upregulated mRNA expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and peroxisome proliferator-activated receptor gamma (PPAR-γ). Kelch-like ECH-associated protein 1 (Keap1) and protein tyrosine phosphatase 1B (PTP1B) mRNA expression in the liver was downregulated in diabetic rats treated with MESFL and metformin. In addition, MESFL downregulated the mRNA expression of caspase-3 in diabetic rats. CONCLUSION: It can be concluded from the data presented in this study that MESFL exerts a protective effect on diabetic rats due to its antidiabetic, antioxidant, antihyperlipidemic and antiapoptotic effects and may be considered a treatment for T2DM.

Asunto(s)

Diabetes Mellitus Experimental , Proteína 1 Asociada A ECH Tipo Kelch , Hígado , Factor 2 Relacionado con NF-E2 , Estrés Oxidativo , PPAR gamma , Extractos Vegetales , Hojas de la Planta , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Ratas Wistar , Transducción de Señal , Animales , Factor 2 Relacionado con NF-E2/metabolismo , Masculino , Extractos Vegetales/farmacología , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Estrés Oxidativo/efectos de los fármacos , Hojas de la Planta/química , Transducción de Señal/efectos de los fármacos , Ratas , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , PPAR gamma/metabolismo , PPAR gamma/genética , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Asteraceae/química , Estreptozocina , Hipoglucemiantes/farmacología

RESUMEN

Type 2 diabetes mellitus (T2DM) is a chronic metabolic disease mainly caused by insulin resistance, which can lead to a series of complications such as cardiovascular disease, retinopathy, and its typical clinical symptom is hyperglycaemia. Glucosidase inhibitors, including Acarbose, Miglitol, are commonly used in the clinical treatment of hypoglycaemia. In addition, Protein tyrosine phosphatase 1B (PTP1B) is also an important promising target for the treatment of T2DM. Gynostemma pentaphyllum is a well-known oriental traditional medicinal herbal plant, and has many beneficial effects on glucose and lipid metabolism. In the present study, three new and nine known dammarane triterpenoids isolated from G. pentaphyllum, and their structures were elucidated by spectroscopic methods including HR-ESI-MS,1H and 13C NMR and X-ray crystallography. All these compounds were evaluated for inhibitory activity against α-glucosidase, α-amylase and PTP1B. The results suggested that compounds 7∼10 were potential antidiabetic agents with significantly inhibition activity against PTP1B in a dose-dependent manner.

Asunto(s)

Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos , Gynostemma , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Gynostemma/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/aislamiento & purificación , Estructura Molecular , Relación Estructura-Actividad , alfa-Glucosidasas/metabolismo , Humanos , alfa-Amilasas/antagonistas & inhibidores , alfa-Amilasas/metabolismo , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Hipoglucemiantes/aislamiento & purificación , Modelos Moleculares , Cristalografía por Rayos X , Triterpenos/farmacología , Triterpenos/química , Triterpenos/aislamiento & purificación , Inhibidores de Glicósido Hidrolasas/farmacología , Inhibidores de Glicósido Hidrolasas/química , Inhibidores de Glicósido Hidrolasas/aislamiento & purificación

RESUMEN

The misregulation of protein phosphatases is a key factor in the development of many human diseases, notably cancers. Here, based on a 100 MHz quartz crystal microbalance (QCM) biosensing platform, the dephosphorylation process of phosphopeptide (P-peptide) caused by protein tyrosine phosphatase 1B (PTP1B) was monitored in real time for the first time and PTP1B activity was assayed rapidly and sensitively. The QCM chip, coated with a gold (Au) film, was used to immobilized thiol-labeled single-stranded 5'-phosphate-DNAs (P-DNA) through Au-S bond. The P-peptide, specific to PTP1B, was then connected to the P-DNA via chelation between Zr4+ and phosphate groups. When PTP1B was injected into the QCM flow cell where the P-peptide/Zr4+/MCH/P-DNA/Au chip was placed, the P-peptide was dephosphorylated and released from the Au chip surface, resulting in an increase in the frequency of the QCM Au chip. This allowed the real-time monitoring of the P-peptide dephosphorylation process and sensitive detection of PTP1B activity within 6 min with a linear detection range of 0.01-100 pM and a detection limit of 0.008 pM. In addition, the maximum inhibitory ratios of inhibitors were evaluated using this proposed 100 MHz QCM biosensor. The developed 100 MHz QCM biosensing platform shows immense potential for early diagnosis of diseases related to protein phosphatases and the development of drugs targeting protein phosphatases.

Asunto(s)

Técnicas Biosensibles , Fosfopéptidos , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Tecnicas de Microbalanza del Cristal de Cuarzo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/análisis , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Tecnicas de Microbalanza del Cristal de Cuarzo/métodos , Fosfopéptidos/análisis , Técnicas Biosensibles/métodos , Fosforilación , Humanos , Circonio/química , Factores de Tiempo , Oro/química , Pruebas de Enzimas/métodos

RESUMEN

The abnormal expression of protein tyrosine phosphatase 1B (PTP1B) is highly related to several serious human diseases. Therefore, an accurate PTP1B activity assay is beneficial to the diagnosis and treatment of these diseases. In this study, a dual-mode biosensing platform that enabled the sensitive and accurate assay of PTP1B activity was constructed based on the high-frequency (100 MHz) quartz crystal microbalance (QCM) and dual-signaling electrochemical (EC) ratiometric strategy. Covalent-organic framework@gold nanoparticles@ferrocene@single-strand DNA (COF@Au@Fc-S0) was introduced onto the QCM Au chip via the chelation between Zr4+ and phosphate groups (phosphate group of the phosphopeptide (P-peptide) on the QCM Au chip and the phosphate group of thiol-labeled single-stranded DNA (S0) on COF@Au@Fc-S0) and used as a signal reporter. When PTP1B was present, the dephosphorylation of the P-peptide led to the release of COF@Au@Fc-S0 from the QCM Au chip, resulting in an increase in the frequency of the QCM. Meanwhile, the released COF@Au@Fc-S0 hybridized with thiol/methylene blue (MB)-labeled hairpin DNA (S1-MB) on the Au NPs-modified indium-tin oxide (ITO) electrode. This caused MB to be far away from the electrode surface and Fc to be close to the electrode, leading to a decrease in the oxidation peak current of MB and an increase in the oxidation peak current of Fc. Thus, PTP1B-induced dephosphorylation of the P-peptide was monitored in real time by QCM, and PTP1B activity was detected sensitively and reliably using this innovative QCM-EC dual-mode sensing platform with an ultralow detection limit. This platform is anticipated to serve as a robust tool for the analysis of protein phosphatase activity and the discovery of drugs targeting protein phosphatase.

Asunto(s)

Técnicas Electroquímicas , Compuestos Ferrosos , Oro , Estructuras Metalorgánicas , Metalocenos , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Tecnicas de Microbalanza del Cristal de Cuarzo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/análisis , Oro/química , Humanos , Estructuras Metalorgánicas/química , Compuestos Ferrosos/química , Metalocenos/química , ADN de Cadena Simple/química , ADN de Cadena Simple/metabolismo , Nanopartículas del Metal/química , Técnicas Biosensibles/métodos , Circonio/química , Pruebas de Enzimas/métodos

RESUMEN

Pueraria lobata (P. lobata), a traditional anti-diabetic medicine mainly composed of flavonoids and isoflavones, has a long history in diabetes treatment in China. However, the anti-diabetic active component is still unclear. Recently, protein tyrosine phosphatase 1B (PTP1B) has been a hot therapeutic target by negatively regulating insulin signaling pathways. In this study, the spectrum-effect relationship analysis method was first used to identify the active components of P. lobata that inhibit PTP1B. The fingerprints of 12 batches of samples were established using high-performance liquid chromatography (HPLC), and sixty common peaks were identified. Meanwhile, twelve components were identified by a comparison with the standards. The inhibition of PTP1B activity was studied in vitro by using the p-nitrophenol method, and the partial least squares discriminant analysis, grey relational analysis, bivariate correlation analysis, and cluster analysis were used to analyze the bioactive compounds in P. lobata. Peaks 6, 9 (glycitin), 11 (genistin), 12 (4'-methoxypuerarin), 25, 34, 35, 36, 53, and 59 were considered as potentially active substances that inhibit PTP1B. The in vitro PTP1B inhibitory activity was confirmed by glycitin, genistin, and 4'-methoxypuerarin. The IC50s of the three compounds were 10.56 ± 0.42 µg/mL, 16.46 ± 0.29 µg/mL, and 9.336 ± 0.56 µg/mL, respectively, indicating the obvious PTP1B inhibitory activity. In brief, we established an effective method to identify PTP1B enzyme inhibitors in P. lobata, which is helpful in clarifying the material basis of P. lobata on diabetes. Additionally, it is evident that the spectrum-effect relationship method serves as an efficient approach for identifying active compounds, and this study can also serve as a reference for screening bioactive constituents in traditional Chinese medicine.

Asunto(s)

Inhibidores Enzimáticos , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Pueraria , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Pueraria/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Cromatografía Líquida de Alta Presión , Isoflavonas/farmacología , Isoflavonas/química , Extractos Vegetales/química , Extractos Vegetales/farmacología , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Humanos

RESUMEN

Tyrosine protein phosphatase non-receptor type 1 (PTP1B; also known as protein tyrosine phosphatase 1B) is a member of the protein tyrosine phosphatase (PTP) family and is a soluble enzyme that plays an essential role in different physiological processes, including the regulation of metabolism, specifically in insulin and leptin sensitivity. PTP1B is crucial in the pathogenesis of type 2 diabetes mellitus and obesity. These biological functions have made PTP1B validated as an antidiabetic and anti-obesity, and potentially anticancer, molecular target. Four main approaches aim to inhibit PTP1B: orthosteric, allosteric, bidentate inhibition, and PTPN1 gene silencing. Developing a potent and selective PTP1B inhibitor is still challenging due to the enzyme's ubiquitous expression, subcellular location, and structural properties. This article reviews the main advances in the study of PTP1B since it was first isolated in 1988, as well as recent contextual information related to the PTP family to which this protein belongs. Furthermore, we offer an overview of the role of PTP1B in diabetes and obesity, and the challenges to developing selective, effective, potent, bioavailable, and cell-permeable compounds that can inhibit the enzyme.

Asunto(s)

Diabetes Mellitus Tipo 2 , Inhibidores Enzimáticos , Hipoglucemiantes , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/química , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Humanos , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Hipoglucemiantes/uso terapéutico , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/uso terapéutico , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/enzimología , Diabetes Mellitus Tipo 2/genética , Animales , Obesidad/tratamiento farmacológico , Obesidad/metabolismo , Obesidad/enzimología , Obesidad/genética