RESUMEN
Microglial cells are crucial in maintaining central nervous system (CNS) homeostasis and mediating CNS disease pathogenesis. Increasing evidence supports that alterations in the mechanical properties of CNS microenvironments influence glial cell phenotypes, but the mechanisms regulating microglial cell function remain elusive. Here, we examined the mechanosensitive Piezo1 channel in microglial cells, particularly, how Piezo1 channel activation regulates pro-inflammatory activation and production of pro-inflammatory cytokines, using BV2 and primary microglial cells. Piezo1 expression in microglial cells was detected both at mRNA and protein levels. Application of Piezo1 channel activator Yoda1 induced Ca2+ flux to increase intracellular Ca2+ concentration that was reduced by treatment with ruthenium red, a Piezo1 inhibitor, or Piezo1-specific siRNA, supporting that Piezo1 functions as a cell surface Ca2+ -permeable channel. Priming with lipopolysaccharide (LPS) induced microglial cell activation and production of TNF-α and IL-6, which were inhibited by treatment with Yoda1. Furthermore, LPS priming induced the activation of ERK, p38 MAPKs, and NF-κB. LPS-induced activation of NF-κB, but not ERK and p38, was inhibited by treatment with Yoda1. Yoda1-induced inhibition was blunted by siRNA-mediated depletion of Piezo1 expression and, furthermore, treatment with BAPTA-AM to prevent intracellular Ca2+ increase. Collectively, our results support that Piezo1 channel activation downregulates the pro-inflammatory function of microglial cells, especially production of TNF-α and IL-6, by initiating intracellular Ca2+ signaling to inhibit the NF-κB inflammatory signaling pathway. These findings reveal Piezo1 channel activation as a previously unrecognized mechanism regulating microglial cell function, raising an interesting perspective on targeting this molecular mechanism to alleviate neuroinflammation and associated CNS pathologies.
Asunto(s)
Lipopolisacáridos , FN-kappa B , FN-kappa B/metabolismo , Lipopolisacáridos/toxicidad , Factor de Necrosis Tumoral alfa/metabolismo , Interleucina-6/metabolismo , Microglía/metabolismo , Transducción de Señal , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismoRESUMEN
Large-conductance Ca2+-activated K+ (BKCa) channel and L-type voltage-dependent Ca2+ channel (L-VDCC) play important roles in regulating uterine contractility. The uterus stretch, occurring during pregnancy, is a critical factor to trigger uterine contraction. However, how mechanical stimuli impact the two channels remains unknown. Here we investigated the effects of exposure to mechanical stretches with varying magnitudes and durations on expressions of the two channels in rat uterine smooth muscle cells. Our results show that stretch down-regulates the BKCa channel expression but upregulates the L-VDCC expression. These findings are helpful to better understand the roles of L-VDCC and BKCa channel in stretch-triggered uterine contraction.
Asunto(s)
Canales de Calcio Tipo L , Canales de Potasio de Gran Conductancia Activados por el Calcio , Miocitos del Músculo Liso , Contracción Uterina , Útero , Animales , Canales de Calcio Tipo L/genética , Canales de Calcio Tipo L/metabolismo , Femenino , Canales de Potasio de Gran Conductancia Activados por el Calcio/genética , Canales de Potasio de Gran Conductancia Activados por el Calcio/metabolismo , Miocitos del Músculo Liso/fisiología , Embarazo , Ratas , Útero/fisiologíaRESUMEN
Esophageal squamous cell carcinoma (ESCC) is a common subtype of esophageal cancer with high incidence. Surgery remains the main strategy for treatment of ESCC at early stage. However, the treatment outcome is unsatisfactory. Therefore, finding new therapeutics is of great importance. In the present study, we measured the level of NEDD4L, an ubiquitin protein ligase, in clinical samples and investigated the effects of NEDD4L on cell viability, cell cycle progression, and glutamine metabolism in TE14 cells determined by CCK-8 assay, flow cytometry and biochemical analysis, respectively. The results show that NEDD4L is significantly decreased in ESCC specimens, and its decreased expression is associated with a poor clinical outcome. Overexpression of NEDD4L significantly inhibits cell viability, cell cycle progression, and glutamine metabolism in TE14 cells. Mechanistic study indicates that NEDD4L regulates tumor progression through ubiquitination of c-Myc and modulation of glutamine metabolism. NEDD4L inhibits cell viability, cell cycle progression, and glutamine metabolism in ESCC by ubiquitination of c-Myc to decrease the expressions of GLS1 and SLC1A5. Our findings highlight the importance of NEDD4L/c-Myc signaling in ESCC.
Asunto(s)
Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Genes myc , Proteínas Proto-Oncogénicas c-myc , Humanos , Sistema de Transporte de Aminoácidos ASC/metabolismo , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas de Esófago/genética , Carcinoma de Células Escamosas de Esófago/metabolismo , Glutamina/metabolismo , Antígenos de Histocompatibilidad Menor/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Transducción de Señal , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación , Genes myc/genéticaRESUMEN
Vascular stiffening, an early and common characteristic of cardiovascular diseases (CVDs), stimulates vascular smooth muscle cell (VSMC) proliferation which reciprocally accelerates the progression of CVDs. However, the mechanisms by which extracellular matrix stiffness accompanying vascular stiffening regulates VSMC proliferation remain largely unknown. In the present study, we examined the role of the intermediate-conductance Ca2+ -activated K+ (IKCa ) channel in the matrix stiffness regulation of VSMC proliferation by growing A7r5 cells on soft and stiff polydimethylsiloxane substrates with stiffness close to these of arteries under physiological and pathological conditions, respectively. Stiff substrates stimulated cell proliferation and upregulated the expression of the IKCa channel. Stiff substrate-induced cell proliferation was suppressed by pharmacological inhibition using TRAM34, an IKCa channel blocker, or genetic depletion of the IKCa channel. In addition, stiff substrate-induced cell proliferation was also suppressed by reducing extracellular Ca2+ concentration using EGTA or intracellular Ca2+ concentration using BAPTA-AM. Moreover, stiff substrate induced activation of extracellular signal-regulated kinases (ERKs), which was inhibited by treatment with TRAM34 or BAPTA-AM. Stiff substrate-induced cell proliferation was suppressed by treatment with PD98059, an ERK inhibitor. Taken together, these results show that substrates with pathologically relevant stiffness upregulate the IKCa channel expression to enhance intracellular Ca2+ signaling and subsequent activation of the ERK signal pathway to drive cell proliferation. These findings provide a novel mechanism by which vascular stiffening regulates VSMC function.
Asunto(s)
Señalización del Calcio , Proliferación Celular , Dimetilpolisiloxanos/química , Canales de Potasio de Conductancia Intermedia Activados por el Calcio/metabolismo , Mecanotransducción Celular , Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/metabolismo , Animales , Técnicas de Cultivo de Célula , Línea Celular , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Canales de Potasio de Conductancia Intermedia Activados por el Calcio/genética , RatasRESUMEN
Mechanical stimulation is an important factor regulating mesenchymal stem cell (MSC) functions such as proliferation. The Ca2+ -activated K+ channel, KCa 3.1, is critically engaged in MSC proliferation but its role in mechanical regulation of MSC proliferation remains unknown. Here, we examined the KCa 3.1 channel expression and its role in rat bone marrow-derived MSC (BMSC) proliferation in response to mechanical stretch. Application of mechanical stretch stimulated BMSC proliferation via promoting cell cycle progression. Such mechanical stimulation up-regulated the KCa 3.1 channel expression and pharmacological or genetic inhibition of the KCa 3.1 channel strongly suppressed stretch-induced increase in cell proliferation and cell cycle progression. These results support that the KCa 3.1 channel plays an important role in transducing mechanical forces to MSC proliferation. Our finding provides new mechanistic insights into how mechanical stimuli regulate MSC proliferation and also a viable bioengineering approach to improve MSC proliferation.
Asunto(s)
Canales de Potasio de Conductancia Intermedia Activados por el Calcio/metabolismo , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Estrés Mecánico , Animales , Proliferación Celular , Masculino , Ratas Sprague-DawleyRESUMEN
Protein tyrosine phosphatase 1B (PTP1B) is a widely confirmed target of the type 2 diabetes mellitus (T2DM) treatment. Herein, we reported a highly specific PTP1B inhibitor 2,2',3,3'-tetrabromo-4,4',5,5'-tetrahydroxydiphenylmethane (compound 1), which showed promising hypoglycemic activity in diabetic BKS db mice. With the IC50 value of 2.4 µM, compound 1 could directly bind to the catalytic pocket of PTP1B through a series of hydrogen bonds. Surface plasmon resonance analysis revealed that the target affinity [KD (equilibrium dissociation constant) value] of compound 1 binding to PTP1B was 2.90 µM. Moreover, compound 1 could activate the insulin signaling pathway in C2C12 skeletal muscle cells. We further evaluated the long-term effects of compound 1 in diabetic BKS db mice. Notably, oral administration of compound 1 significantly reduced the blood glucose levels of diabetic mice with increasing insulin sensitivity. In addition, the dyslipidemia of diabetic mice was also significantly improved by compound 1 gavage. The histological experiments showed that compound 1 treatment significantly ameliorated the disordered hepatic and pancreatic architecture and increased the glycogen content in the liver tissues as well as improved the insulin secretion function of pancreas. Taken together, our results manifested that the natural product compound 1 was a highly specific PTP1B inhibitor, which could activate insulin signaling pathway and ameliorate hyperglycemia and dyslipidemia in diabetic BKS db mice.
Asunto(s)
Compuestos de Bencidrilo , Diabetes Mellitus Tipo 2 , Hipoglucemiantes , Extractos Vegetales , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Animales , Masculino , Ratones , Administración Oral , Compuestos de Bencidrilo/administración & dosificación , Compuestos de Bencidrilo/química , Compuestos de Bencidrilo/farmacología , Compuestos de Bencidrilo/uso terapéutico , Dominio Catalítico , Línea Celular , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Glucógeno/metabolismo , Enlace de Hidrógeno , Hipoglucemiantes/administración & dosificación , Hipoglucemiantes/química , Hipoglucemiantes/farmacología , Hipoglucemiantes/uso terapéutico , Concentración 50 Inhibidora , Insulina/metabolismo , Resistencia a la Insulina , Hígado/efectos de los fármacos , Hígado/metabolismo , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Simulación del Acoplamiento Molecular , Mioblastos/efectos de los fármacos , Mioblastos/metabolismo , Extractos Vegetales/administración & dosificación , Extractos Vegetales/química , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/química , Proteína Tirosina Fosfatasa no Receptora Tipo 1/aislamiento & purificación , Rhodophyta/química , Transducción de Señal/efectos de los fármacosRESUMEN
RNA-binding proteins (RBPs) lie at the center of posttranscriptional regulation and the dysregulation of RBPs contributes to diabetes. Therefore, the modulation of RBPs is anticipated to become a potential therapeutic approach to diabetes. CYC27 is a synthetic derivative of marine bromophenol BDB, which is isolated from red alga Rhodomela confervoides. In this study, we found that CYC27 significantly lowered the blood glucose levels of diabetic BKS db mice. Moreover, CYC27 effectively ameliorated dyslipidemia in BKS db mice by reducing their total serum cholesterol (TC) and triglyceride (TG) levels. Furthermore, CYC27 was an insulin-sensitizing agent with increased insulin-stimulated phosphorylation of insulin receptors and relevant downstream factors. Finally, to systemically study the mechanisms of CYC27, label-free quantitative phosphoproteomic analysis was performed to investigate global changes in phosphorylation. Enriched GO annotation showed that most regulated phosphoproteins were related to RNA splicing and RNA processing. Enriched KEGG analysis showed that a spliceosome-associated pathway was the predominant pathway after CYC27 treatment. Protein-protein interaction (PPI) analysis showed that CYC27 modulated the process of mRNA splicing via phosphorylation of the relevant RBPs, including upregulated Cstf3 and Srrt. Our results suggested that CYC27 treatment exerted promising anti-diabetic effects by sensitizing the insulin signaling pathways and modulating RNA splicing-associated RBPs.
Asunto(s)
Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hipoglucemiantes/farmacología , Bifenilos Polibrominados/farmacología , Proteínas de Unión al ARN/metabolismo , Rhodophyta/química , Animales , Glucemia/análisis , Glucemia/efectos de los fármacos , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/genética , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Humanos , Hipoglucemiantes/síntesis química , Concentración 50 Inhibidora , Insulina/metabolismo , Masculino , Ratones , Fosforilación/efectos de los fármacos , Bifenilos Polibrominados/síntesis química , Mapas de Interacción de Proteínas/efectos de los fármacos , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Empalme del ARN/efectos de los fármacos , Transducción de Señal/efectos de los fármacosRESUMEN
BACKGROUND: Hemolytic streptococcus gangrene is a life threatening invasive bacterial infection. Hemolytic streptococcus gangrene in the danger triangle of the face is too lethal to operate. A case of the confirmed hemolytic streptococcus gangrene in the danger triangle of the face caused by Group A beta-hemolytic streptococcus (GAS) in 20-months old boy is presented to draw attention of clinicians to this uncommon but frequently fatal infection. CASE PRESENTATION: Previously healthy 20 months old boy suddenly developed paranasal gangrene on the left side of the danger triangle of the face, followed by rapidly progressive thrombocytopenia and hepatitis. The clinical features, liver function, and hematological and serological parameters resembled to a description of streptococcal toxic shock syndrome (STSS). Aggressive antibiotics, substitutional and supportive therapy were conducted without surgical debridement of facial tissues. Prompt diagnosis and aggressive timely treatment completely cured the disease in 28 days. CONCLUSIONS: The present case report demonstrates prompt diagnosis and timely treatment as a strategy to cure the fatal hemolytic streptococcus gangrene located in too risky body part to operate.
Asunto(s)
Cara/patología , Gangrena/complicaciones , Gangrena/microbiología , Hepatitis/complicaciones , Infecciones Estreptocócicas/complicaciones , Streptococcus pyogenes , Trombocitopenia/complicaciones , Antibacterianos/uso terapéutico , Farmacorresistencia Bacteriana , Diagnóstico Precoz , Humanos , Lactante , Masculino , Meropenem/uso terapéutico , Penicilina G/uso terapéutico , Infecciones Estreptocócicas/diagnóstico , Infecciones Estreptocócicas/tratamiento farmacológico , Vancomicina/uso terapéuticoRESUMEN
Both fibronectin (FN) and filamentous actin (F-actin) fibers play a critical role for endothelial cells (ECs) in responding to shear stress and modulating cell alignment and functions. FN is dynamically coupled to the F-actin cytoskeleton via focal adhesions. However, it is unclear how ECs cooperatively remodel their subcellular FN matrix and intracellular F-actin cytoskeleton in response to shear stress. Current studies are hampered by the lack of a reliable and sensitive quantification method of FN orientation. In this study, we developed a MATLAB-based feature enhancement method to quantify FN and F-actin orientation. The role of F-actin in FN remodeling was also studied by treating ECs with cytochalasin D. We have demonstrated that FN and F-actin codistributed and coaligned parallel to the flow direction, and that F-actin alignment played an essential role in regulating FN alignment in response to shear stress. Our findings offer insight into how ECs cooperatively remodel their subcellular ECM and intracellular F-actin cytoskeleton in response to mechanical stimuli, and are valuable for vascular tissue engineering.
Asunto(s)
Actinas/metabolismo , Fibronectinas/metabolismo , Estrés Mecánico , Estrés Fisiológico/fisiología , Citoesqueleto de Actina/fisiología , Animales , Células Cultivadas , Citocalasina D/farmacología , Células Endoteliales , Ratas , Ratas Sprague-DawleyRESUMEN
One of the major challenges in tissue engineering of small-diameter vascular grafts is to inhibit intimal hyperplasia and keep long-term patency after implantation. Rapid endothelialization of the grafts could be an effective approach. In this study, QK, a peptide mimicking vascular endothelial growth factor, was selected as the bioactive substrate and loaded in electrospun membranes for enhancement of vascular endothelial cell growth. In detail, QK peptide was firstly introduced with poly(ethylene glycol) diacrylate into a thiolated chitosan solution that could transfer into hydrogel. Then, suspensions or emulsions of poly(ethylene glycol)-b-poly(L-lactide-co-ε-caprolactone) (PELCL) containing QK peptide (with or without chitosan hydrogel) were electrospun into fibrous membranes. For comparison, the electrospun PELCL membrane without QK was also fabricated. Results of release behaviors showed that the electrospun membranes, especially that contained chitosan hydrogel prepared by suspension electrospinning, could successfully encapsulate QK peptide and maintain its secondary structure after released. In vitro cell culture studies exhibited that the release of QK peptide could accelerate the proliferation of vascular endothelial cells in the 9 days. It was suggested that the electrospun PELCL membranes loaded with QK peptide might have potential applications in vascular tissue engineering.
Asunto(s)
Células Endoteliales/efectos de los fármacos , Membranas Artificiales , Péptidos/farmacología , Poliésteres/química , Polietilenglicoles/química , Polímeros/química , Prótesis Vascular , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Liberación de Fármacos , Humanos , Ensayo de Materiales , Microscopía Electrónica de Rastreo , Péptidos/químicaRESUMEN
Previous studies have indicated that hormonal control is essential for plant root growth. The root of the carrot is an edible vegetable with a high nutritional value. However, molecular mechanisms underlying hormone-mediated root growth of carrot have not been illustrated. Therefore, the present study collected carrot root samples from four developmental stages, and performed transcriptome sequencing to understand the molecular functions of plant hormones in carrot root growth. A total of 160,227 transcripts were generated from our transcriptome, which were assembled into 32,716 unigenes with an average length of 1,453 bp. A total of 4,818 unigenes were found to be differentially expressed between the four developmental stages. In total, 87 hormone-related differentially expressed genes were identified, and the roles of the hormones are extensively discussed. Our results suggest that plant hormones may regulate carrot root growth in a phase-dependent manner, and these findings will provide valuable resources for future research on carrot root development.
Asunto(s)
Daucus carota/genética , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Reguladores del Crecimiento de las Plantas/biosíntesis , Raíces de Plantas/genética , Transcriptoma/genética , Análisis por Conglomerados , Daucus carota/crecimiento & desarrollo , Daucus carota/metabolismo , Ontología de Genes , Secuenciación de Nucleótidos de Alto Rendimiento , Anotación de Secuencia Molecular , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/metabolismo , Factores de TiempoRESUMEN
The large conductance Ca(2+)-activated K(+) (BK(Ca)) channel in vascular smooth muscle cell (VSMC) is an important potassium channel that can regulate vascular tone. Recent work has demonstrated that abnormalities in BK(Ca) channel function are associated with changes in cell proliferation and the onset of vascular disease. However, until today there are rare reports to show whether this channel is involved in VSMC proliferation in response to fluid shear stress (SS). Here we investigated a possible role of BK(Ca) channel in VSMC proliferation under laminar SS. Rat aortic VSMCs were plated in parallel-plate flow chambers and exposed to laminar SS with varied durations and magnitudes. VSMC proliferation was assessed by measuring proliferating cell nuclear antigen (PCNA) expression and DNA synthesis. BK(Ca) protein and gene expression was determined by flow cytometery and RT-PCR. The involvement of BK(Ca) in SS-induced inhibition of proliferation was examined by BK(Ca) inhibition using a BK(Ca) specific blocker, iberiotoxin (IBTX), and by BK(Ca) transfection in BK(Ca) non-expressing CHO cells. The changes in [Ca(2+)](i) were determined using a calcium-sensitive dye, fluo 3-AM. Membrane potential changes were detected with a potential-sensitive dye, DiBAC(4)(3). We found that laminar SS inhibited VSMC proliferation and stimulated BK(Ca) channel expression. Furthermore, laminar SS induced an increase in [Ca(2+)](i) and membrane hyperpolarization. Besides in VSMC, the inhibitory effect of BK(Ca) channel activity on cell proliferation in response to SS was also confirmed in BK(Ca)-transfected CHO cells showing a decline in proliferation. Blocking BK(Ca) channel reversed its inhibitory effect, providing additional support for the involvement of BK(Ca) in SS-induced proliferation reduction. Our results suggest, for the first time, that BK(Ca) channel mediates laminar SS-induced inhibition of VSMC proliferation. This finding is important for understanding the mechanism by which SS regulates VSMC proliferation, and should be helpful in developing strategies to prevent flow-initiated vascular disease formation.
Asunto(s)
Proliferación Celular , Canales de Potasio de Gran Conductancia Activados por el Calcio/fisiología , Músculo Liso Vascular/fisiología , Miocitos del Músculo Liso/fisiología , Estrés Mecánico , Animales , Células CHO , Calcio/metabolismo , Cricetinae , Cricetulus , Replicación del ADN , Expresión Génica , Canales de Potasio de Gran Conductancia Activados por el Calcio/genética , Canales de Potasio de Gran Conductancia Activados por el Calcio/metabolismo , Masculino , Potenciales de la Membrana , Músculo Liso Vascular/citología , Miocitos del Músculo Liso/metabolismo , Antígeno Nuclear de Célula en Proliferación/genética , Antígeno Nuclear de Célula en Proliferación/metabolismo , Ratas , Ratas Sprague-Dawley , Imagen de Colorante Sensible al VoltajeRESUMEN
Nanoscale materials (such as TiO2, hydroxyapatite nanoparticles) have gained much concern in the coating of implants for cell adhesion and growth to improve the osteoconductivity. However, due to attrition and corrosion, the wear particles would be generated from the joint in living organism, and influence the physiological function of synovial membranes in joint cavity. In this study, the potential cytotoxicity of anatase TiO2 nanoparticles (TiO2 NPs) on rat synovial cell line 364 (RSC-364) was investigated. After treatment with different concentrations of TiO2 NPs (0, 3, 30, 300 microg/ml), the viability of RSC-364 cells were decreased in a dose-dependent manner. TiO2 NPs exposure could disrupt the integrity of cell plasma membrane, leading to the increased leakage of lactate dehydrogenase (LDH) into the culture medium. TiO2 NPs were uptaken by RSC-364 cells. The ultrastructure of RSC-364 cells was changed such as nuclear shrinkage and mitochondrial swelling. The reactive oxygen species (ROS) was over-produced especially in the cells exposed to 30 and 300 microg/ml TiO2 NPs. The activities of endogeneous antioxidant enzymes, superoxide dismutase (SOD) and catalase (CAT), were significantly decreased. The increased lipid peroxidation product (malondialdehyde, MDA) suggests the oxidative damage in cells. The flow cytometry detected that the cell cycle was blocked in G0/G1 phase, inhibiting the cell proliferation. These preliminary results indicate the oxidative stress injury and cytotoxicity of anatase TiO2 NPs on rat synovial cells. The reasonable and safe application of nanomaterials in artificial implants needs further study.
Asunto(s)
División Celular/efectos de los fármacos , Membrana Sinovial/efectos de los fármacos , Titanio/farmacología , Animales , Ciclo Celular , Línea Celular , Ratas , Membrana Sinovial/citologíaRESUMEN
The increased proliferation of vascular smooth muscle cells (VSMCs) contributes to the pathogenesis of vascular diseases. The intermediate conductance calcium-activated potassium (IKCa ) channel plays a critical role in VSMC proliferation by raising the intracellular calcium concentration ([Ca2+ ]i ), but the underlying mechanism is still not unclear. Here we investigated the cooperation between IKCa and transient receptor potential canonical 1 (TRPC1) channels in mediating extracellular Ca2+ entry, which in turn activates downstream Ca2+ signalling in the regulation of VSMC proliferation using serum-induced cell proliferation model. Serum-induced cell proliferation was accompanied with up-regulation of IKCa expression and an increase in [Ca2+ ]i . Serum-induced cell proliferation and increase in [Ca2+ ]i were suppressed by IKCa inhibition with TRAM-34 or IKCa knockdown. Serum-induced cell proliferation was strongly reduced by the removal of extracellular Ca2+ with EGTA or intracellular Ca2+ with BAPTA-AM and, additionally, by TRPC1 knockdown. Moreover, the increase in [Ca2+ ]i induced by serum or by IKCa activation with 1-EBIO was attenuated by TRPC1 knockdown. Finally, serum induced ERK1/2 activation, which was attenuated by treatment with TRAM-34 or BAPTA-AM, as well as TRPC1 knockdown. Consistently, serum-induced cell proliferation was suppressed by ERK1/2 inhibition with PD98059. Taken together, these results suggest that the IKCa and TRPC1 channels cooperate in mediating Ca2+ influx that activates the ERK1/2 pathway to promote cell proliferation, thus providing new mechanistic insights into VSMC proliferation.
Asunto(s)
Músculo Liso Vascular , Canales de Potencial de Receptor Transitorio , Músculo Liso Vascular/metabolismo , Canales de Potencial de Receptor Transitorio/metabolismo , Calcio/metabolismo , Sistema de Señalización de MAP Quinasas , Proliferación Celular , Canales Catiónicos TRPC/metabolismo , Miocitos del Músculo Liso/metabolismoRESUMEN
Developing new antimicrobial agents has become an urgent task to address the increasing prevalence of multidrug-resistant pathogens and the emergence of biofilms. Cationic antimicrobial peptides (AMPs) have been regarded as promising candidates due to their unique non-specific membrane rupture mechanism. However, a series of problems with the peptides hindered their practical application due to their high toxicity and low bioactivity and stability. Here, inspired by broadening the application of cell-penetrating peptides (CPPs), we selected five different sequences of cationic peptides which are considered as both CPPs and AMPs, and developed a biomimetic strategy to construct cationic peptide-conjugated liposomes with the virus-like structure for both enhancements of antibacterial efficacy and biosafety. The correlation between available peptide density/peptide variety and antimicrobial capabilities was evaluated from quantitative perspectives. Computational simulation and experimental investigations assisted to identify the optimal peptide-conjugated liposomes and revealed that the designed system provides high charge density for enhanced anionic bacterial membrane binding capability without compromised cytotoxicity, being capable of enhanced antibacterial efficacy of bacteria/biofilm of clinically important pathogens. The bio-inspired design has shown enhanced therapeutic efficiency of peptides and may promote the development of next-generation antimicrobials.
Asunto(s)
Antiinfecciosos , Péptidos de Penetración Celular , Liposomas/metabolismo , Plancton , Membrana Celular/metabolismo , Bacterias , Péptidos Catiónicos Antimicrobianos/farmacología , Péptidos Catiónicos Antimicrobianos/química , Antiinfecciosos/metabolismo , Antibacterianos/farmacología , Antibacterianos/química , Péptidos de Penetración Celular/farmacología , Péptidos de Penetración Celular/metabolismo , Biopelículas , Pruebas de Sensibilidad MicrobianaRESUMEN
The neuromuscular junction (NMJ) is a peripheral synaptic connection between presynaptic motor neurons and postsynaptic skeletal muscle fibers that enables muscle contraction and voluntary motor movement. Many traumatic, neurodegenerative, and neuroimmunological diseases are classically believed to mainly affect either the neuronal or the muscle side of the NMJ, and treatment options are lacking. Recent advances in novel techniques have helped develop in vitro physiological and pathophysiological models of the NMJ as well as enable precise control and evaluation of its functions. This paper reviews the recent developments in in vitro NMJ models with 2D or 3D cultures, from organ-on-a-chip and organoids to biohybrid robotics. Related derivative techniques are introduced for functional analysis of the NMJ, such as the patch-clamp technique, microelectrode arrays, calcium imaging, and stimulus methods, particularly optogenetic-mediated light stimulation, microelectrode-mediated electrical stimulation, and biochemical stimulation. Finally, the applications of the in vitro NMJ models as disease models or for drug screening related to suitable neuromuscular diseases are summarized and their future development trends and challenges are discussed.
Asunto(s)
Sistemas Microfisiológicos , Unión Neuromuscular , Unión Neuromuscular/fisiología , Neuronas Motoras , Fibras Musculares Esqueléticas , Contracción Muscular , Músculo EsqueléticoRESUMEN
Binding of selectins to P-selectin glycoprotein ligand-1 (PSGL-1) mediates tethering and rolling of leukocytes on the endothelium during inflammation. Previous measurements obtained with a flow-chamber assay have shown that mutations of three tyrosines at the PSGL-1 N-terminus (Y46, Y48, and Y51) increase the reverse rates and their sensitivity to the force of bonds with P- and L-selectin. However, the effects of these mutations on the binding affinities and forward rates have not been studied. We quantified these effects by using an adhesion frequency assay to measure two-dimensional affinity and kinetic rates at zero force. Wild-type PSGL-1 has 2.2- to 8.5-fold higher binding affinities for P- and L-selectin than PSGL-1 mutants with two of three tyrosines substituted by phenylalanines, and 9.6- to 49-fold higher affinities than the PSGL-1 mutant with all three tyrosines replaced. In descending order, the affinity decreased from wild-type to Y48/51F, Y46/51F, Y46/48F, and Y46/48/51F. The affinity differences were attributed to major changes in the forward rate and minor changes in the reverse rate, suggesting that these tyrosines regulate the accessibility of PSGL-1 to P- and L-selectin via electrostatic interactions, which is supported by molecular-dynamics simulations. Our results provide insights into the structure-function relationship of receptor-ligand binding at a single-residue level.
Asunto(s)
Sustitución de Aminoácidos , Membrana Celular/metabolismo , Selectina L/metabolismo , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/metabolismo , Selectina-P/metabolismo , Tirosina , Animales , Células CHO , Cricetinae , Cricetulus , Humanos , Cinética , Glicoproteínas de Membrana/genética , Simulación de Dinámica Molecular , Unión Proteica , Estructura Terciaria de Proteína , Electricidad EstáticaRESUMEN
BACKGROUND: The risk factors for cardiovascular and cerebrovascular diseases in young and middle-aged people have not yet been determined. We conducted a meta-analysis to find the risk factors for cardiovascular and cerebrovascular diseases, in order to provide guidance for the prevention of diseases in the young and middle-aged population. METHODS: We searched PubMed, Embase, Cochrane Library from the establishment of the database to Mar 2022. We included case-control or cohort studies reporting risk factors for cardiovascular and cerebrovascular disease in young and middle-aged adults. We excluded repeated publication, research without full text, incomplete information or inability to conduct data extraction and animal experiments, reviews and systematic reviews. STATA 15.1 was used to analyze the data. RESULTS: The pooled results indicated that increased systolic blood pressure was significantly associated with increased risk of any stroke, ischemic stroke and hemorrhagic stroke. Body Mass Index (BMI), current smoking, hypertension, and diabetes were significantly associated with increased risk of any stroke and ischemic stroke. Atrial fibrillation was only significantly associated with increased risk of any stroke. Increased total cholesterol was significantly associated with an increased risk of ischemic stroke, whereas increased triglycerides were significantly associated with a decreased risk of ischemic stroke. In addition, increased hypertension was also significantly associated with an increased risk of acute coronary syndrome. CONCLUSION: Our pooled results show that BMI, current smoking, atrial fibrillation, hypertension, systolic blood pressure, and total cholesterol can be used as risk factors for cardiovascular and cerebrovascular diseases in young people, while triglycerides can be used as protective factors for cardiovascular and cerebrovascular diseases in young and middle-aged adults.
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Fibrilación Atrial , Trastornos Cerebrovasculares , Hipertensión , Accidente Cerebrovascular Isquémico , Accidente Cerebrovascular , Humanos , Trastornos Cerebrovasculares/epidemiología , Trastornos Cerebrovasculares/etiología , Factores de Riesgo , Hipertensión/epidemiología , ColesterolRESUMEN
Compared with normal cells, tumor cells mainly obtain energy through aerobic glycolysis. Hexokinase 2 (HK2) plays a key role in the regulation of tumor cell aerobic glycolysis, and targeting HK2 has become a new strategy for cancer treatment. However, little is known about the role of HK2 in colon cancer and the regulation of its targeted inhibitors. In this study, we found that the expression of HK2 in colorectal cancer tissues was significantly higher than that in adjacent tissues, and the expression level of HK2 in metastatic colorectal cancer was further increased. Meanwhile, the expression level of HK2 was closely related to clinical TNM stage and outcome of colorectal cancer patients. We provide here evidence that HK2 inhibitor 3-Bromopyruvate acid (3-BP) can significantly inhibit the survival and proliferation of colon cancer cells, and induce apoptosis through mitochondrial apoptosis signaling pathway. In addition, we found that 3-BP can also induce endoplasmic reticulum stress in colon cancer cells, the mechanism may be through the increase of intracellular calcium concentration. In vitro and in vivo experiments showed that inhibition of endoplasmic reticulum stress could further increase the proliferation inhibition and apoptosis induced by 3-BP. Collectively, our results show that HK2 is highly expressed in colorectal cancer. 3-BP, an inhibitor of HK2, can induce apoptosis and endoplasmic reticulum stress in colon cancer cells. Endoplasmic reticulum stress plays a protective role in cell death induced by 3-BP. This result suggested that targeting HK2 and endoplasmic reticulum stress may be a valuable strategy in targeted and combination therapy of colon cancer.
Asunto(s)
Neoplasias del Colon , Hexoquinasa , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Neoplasias del Colon/tratamiento farmacológico , Combinación de Medicamentos , Estrés del Retículo Endoplásmico , Regulación Neoplásica de la Expresión Génica , Glucólisis/fisiología , Hexoquinasa/genética , Hexoquinasa/metabolismo , HumanosRESUMEN
Objective@#To compare the efficacy of two bioceramic materials, iRoot BP Plus and mineral trioxide aggregate (MTA) in the preservation of vital pulp of mature permanent teeth with exposed pulp of caries origin, so as to provide insights into appropriate selection of pulp capping agents in clinical process.@*Methods@#Vital pulp therapy were performed on 120 mature permanent teeth with carious exposure at the Department of Stomatology of Hangzhou Traditional Chinese Medicine Hospital Affiliated to Zhejiang Chinese Medical University. The teeth were randomly divided into two groups which were treated respectively by iRoot BP Plus (iRoot group) and MTA (MTA group). The clinical efficacy was evaluated by clinical examinations and imaging examinations.@*Results@#There were 60 cases in iRoot group, including 23 males and 37 females, 27 cases affected premolars and 33 cases affected molars, and 8 cases of Class I and 52 cases of Class II cavity type, with a median age of 41 (interquartile range, 12) years. There were 60 cases in MTA group, including 29 males and 31 females, 21 cases affected premolars and 39 cases affected molars, and 10 cases of Class I and 50 cases of Class II cavity type, with a median age of 39 (interquartile range, 14) years. There were no significant differences in gender, age, affected tooth location and cavity type between the two groups (P>0.05). The success rate of iRoot group at 12 months was 91.67% while the MTA group was 88.33%, and the pulp infection rate of iRoot group at 12 months was 8.33% while the MTA group was 11.67%. There were no significant differences in success rate and pulp infection rate between the two groups (P>0.05). The rate of crown discoloration in MTA group was 61.67%, while there was no discoloration in iRoot group.@*Conclusions@#During 12 months, iRoot BP Plus and MTA can both achieve great effects in the treatment of mature permanent teeth with carious pulp exposure, but there is a problem of tooth discoloration after pulp covering using MTA. The long-term clinical effects of the two materials need to be further studied.