RESUMEN
The concentrations of eight heavy metals, V, Cr, Co, Ni, Cu, Zn, Cd, and Pb, were analyzed to explore the environmental pollution characteristics, ecological risk levels, and sources of heavy metals in the surface sediments of Luoyuan Bay in Fujian Province. The results indicated that the mean concentrations of V, Cr, Co, Ni, Cu, Zn, Cd, and Pb were 110.44, 97.76, 17.35, 41.99, 36.78, 137.26, 0.09, and 35.81 mgâ kg-1, respectively. The mean concentrations of Cr, Cu, and Zn exceeded the first category of the marine sediment quality standards, indicating a moderate regional comprehensive potential ecological hazard level. Due to variable hydrodynamic conditions, high concentrations of heavy metals were observed in the north and low concentrations were observed in the south, with high values generally recorded near steel plants and shipyard docks. Correlation and principal component analyses revealed that the V, Al, Co, Ni, and Pb originated from the weathering and erosion of rocks, and industrial wastewater discharge. Contrastingly, Cu, Zn, Cr, and Cd were mainly associated with the transportation and repair of ships, and marine aquaculture activities.
Asunto(s)
Metales Pesados , Contaminantes Químicos del Agua , Bahías , China , Monitoreo del Ambiente , Sedimentos Geológicos , Metales Pesados/análisis , Medición de Riesgo , Contaminantes Químicos del Agua/análisisRESUMEN
Allostery exploits the conformational dynamics of enzymes by triggering a shift in population ensembles toward functionally distinct conformational or dynamic states. Allostery extensively regulates the activities of key enzymes within biosynthetic pathways to meet metabolic demand for their end products. Here, we have examined a critical enzyme, 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (DAH7PS), at the gateway to aromatic amino acid biosynthesis in Mycobacterium tuberculosis, which shows extremely complex dynamic allostery: three distinct aromatic amino acids jointly communicate occupancy to the active site via subtle changes in dynamics, enabling exquisite fine-tuning of delivery of these essential metabolites. Furthermore, this allosteric mechanism is co-opted by pathway branchpoint enzyme chorismate mutase upon complex formation. In this study, using statistical coupling analysis, site-directed mutagenesis, isothermal calorimetry, small-angle X-ray scattering, and X-ray crystallography analyses, we have pinpointed a critical node within the complex dynamic communication network responsible for this sophisticated allosteric machinery. Through a facile Gly to Pro substitution, we have altered backbone dynamics, completely severing the allosteric signal yet remarkably, generating a nonallosteric enzyme that retains full catalytic activity. We also identified a second residue of prime importance to the inter-enzyme communication with chorismate mutase. Our results reveal that highly complex dynamic allostery is surprisingly vulnerable and provide further insights into the intimate link between catalysis and allostery.
Asunto(s)
3-Desoxi-7-Fosfoheptulonato Sintasa/química , Proteínas Bacterianas/química , Mutación Missense , Mycobacterium tuberculosis/enzimología , 3-Desoxi-7-Fosfoheptulonato Sintasa/genética , 3-Desoxi-7-Fosfoheptulonato Sintasa/metabolismo , Regulación Alostérica , Sustitución de Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Catálisis , Cristalografía por Rayos X , Mycobacterium tuberculosis/genéticaRESUMEN
Most proteins comprise two or more domains from a limited suite of protein families. These domains are often rearranged in various combinations through gene fusion events to evolve new protein functions, including the acquisition of protein allostery through the incorporation of regulatory domains. The enzyme 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (DAH7PS) is the first enzyme of aromatic amino acid biosynthesis and displays a diverse range of allosteric mechanisms. DAH7PSs adopt a common architecture with a shared (ß/α)8 catalytic domain which can be attached to an ACT-like or a chorismate mutase regulatory domain that operates via distinct mechanisms. These respective domains confer allosteric regulation by controlling DAH7PS function in response to ligand Tyr or prephenate. Starting with contemporary DAH7PS proteins, two protein chimeras were created, with interchanged regulatory domains. Both engineered proteins were catalytically active and delivered new functional allostery with switched ligand specificity and allosteric mechanisms delivered by their nonhomologous regulatory domains. This interchangeability of protein domains represents an efficient method not only to engineer allostery in multidomain proteins but to create a new bifunctional enzyme.
Asunto(s)
3-Desoxi-7-Fosfoheptulonato Sintasa/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Thermotoga maritima/metabolismo , 3-Desoxi-7-Fosfoheptulonato Sintasa/química , 3-Desoxi-7-Fosfoheptulonato Sintasa/genética , Regulación Alostérica , Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión Génica , Regulación Enzimológica de la Expresión Génica , Dominios Proteicos , Thermotoga maritima/genéticaRESUMEN
Neutrophils activated during acute lung injury (ALI) form neutrophil extracellular traps (NETs) to capture pathogens. However, excessive NETs can cause severe inflammatory reactions. Macrophages are classified as M1 macrophages with proinflammatory effects or M2 macrophages with anti-inflammatory effects. During ALI, alveolar macrophages (AMs) polarize to the M1 phenotype. This study tested the hypothesis that NETs may aggravate ALI or acute respiratory distress syndrome (ARDS) inflammation by promoting alveolar macrophage polarization to the M1 type. Our research was carried out in three aspects: clinical research, animal experiments and in vitro experiments. We determined that NET levels in ARDS patients were positively correlated with M1-like macrophage polarization. NET formation was detected in murine ALI tissue and associated with increased M1 markers and decreased M2 markers in BALF and lung tissue. Treatment with NET inhibitors significantly inhibitor NETs generation, downregulated M1 markers and upregulated M2 markers. Regardless of LPS pre-stimulation, significant secretion of proinflammatory cytokines and upregulated M1 markers were detected from bone marrow-derived macrophages (M0 and M2) cocultured with high concentrations of NETs; conversely, M2 markers were downregulated. In conclusion, NETs promote ARDS inflammation during the acute phase by promoting macrophage polarization to the M1 phenotype. We propose that NETs play an important role in the interaction between neutrophils and macrophages during the early acute phase of ALI.
Asunto(s)
Lesión Pulmonar Aguda/patología , Polaridad Celular , Trampas Extracelulares/metabolismo , Macrófagos Alveolares/patología , Síndrome de Dificultad Respiratoria/patología , Animales , Femenino , Lipopolisacáridos , Ratones Endogámicos C57BLRESUMEN
To explore the effect of double-stranded RNA-dependent kinase (PKR) in acute lung injury (ALI) and resultant acute respiratory distress syndrome (ARDS). A mouse model of lipopolysaccharide (LPS)-induced ALI was used to evaluate the levels of phosphorylated (p)-PKR and NLRP3 in lung tissue, and the protective effects of a PKR inhibitor on lung injury. And in vitro, macrophages were incubated with LPS, with or without PKR inhibitor pre-treatment. It was observed that the levels of p-PKR protein and NLRP3 protein were significantly increased compared with those in control tissues after LPS administration. Meanwhile, treatment with PKR inhibitor decreased inflammation, injury score, wet/dry weight ratio, bronchoalveolar lavage fluid (BALF) protein levels, neutrophil count in BALF, myeloperoxidase activity and expression of high-mobility group box1(HMGB1) and interleukin(IL)-1ß in the lungs of LPS-challenged mice. In vitro, we demonstrated that the levels of p-PKR and NLRP3, and cell mortality rate were increased in macrophages which were incubated with LPS compared with those without LPS administration, and PKR inhibitor significantly suppressed the level of NLRP3, caspase-1, HMGB1 and IL-1ß. These results indicate that PKR plays a key role in ALI through NLRP3-pyrotosis pathway and pharmacological inhibition of PKR may have potential therapeutic effects in the treatment of patients with ALI and ARDS.
Asunto(s)
Lesión Pulmonar Aguda/metabolismo , Modelos Animales de Enfermedad , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Piroptosis , eIF-2 Quinasa/metabolismo , Lesión Pulmonar Aguda/inducido químicamente , Lesión Pulmonar Aguda/patología , Animales , Lipopolisacáridos/administración & dosificación , Lipopolisacáridos/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína con Dominio Pirina 3 de la Familia NLR/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , eIF-2 Quinasa/antagonistas & inhibidoresRESUMEN
BACKGROUND/AIM: H2S is a novel vasoactivator. To verify the hypothesis that H2S may act as an endothelium-derived hyperpolarizing factor (EDHF) in the rat cerebrovasculature, the role of H2S in endothelium-derived relaxing factor (EDRF)-mediated responses was investigated. METHODS: Cystathionine-γ-lyase (CSE) was knocked down with an siRNA technique. Artery diameter, hyperpolarization and Ca2+-activated K+ (KCa) current were measured. RESULTS: CSE knockdown was indicated by a decrease in protein and mRNA expression in the rat middle cerebral artery (MCA) and cerebral basilar artery (CBA). Acetylcholine (ACh) induced significant hyperpolarization and vasodilation in endothelium-intact MCA and CBA. Removal of the endothelium abolished these responses. The nitric oxide (NO) synthase inhibitor L-NAME, but not the PGI2 production inhibitor indomethacin, significantly inhibited ACh-induced hyperpolarization and vasodilation in the CBA. In the presence of L-NAME and indomethacin, ACh-induced hyperpolarization and vasodilation in the MCA and CBA were attenuated. The non-NO/PGI2-mediated responses were abolished by the KCa channel blockers charybdotoxin and apamin. In the cerebral arteries from the CSE knockdown rat, non-NO/PGI2-mediated responses were significantly attenuated, and the remaining responses were abolished by charybdotoxin and apamin or the CSE inhibitor propargylglycine. CSE knockdown did not affect L-NAME-sensitive responses in the CBA. Sodium hydrosulfide (NaHS) augmented the KCa current in CBA vascular smooth muscle cells. CONCLUSION: EDHF-mediated responses in rat cerebral arteries were due to H2S activating the KCa channel.
Asunto(s)
Arteria Basilar/metabolismo , Factores Biológicos/metabolismo , Sulfuro de Hidrógeno/metabolismo , Arteria Cerebral Media/metabolismo , Vasodilatación , Animales , Arteria Basilar/efectos de los fármacos , Arteria Basilar/enzimología , Inhibidores de la Ciclooxigenasa/farmacología , Cistationina gamma-Liasa/genética , Cistationina gamma-Liasa/metabolismo , Relación Dosis-Respuesta a Droga , Epoprostenol/metabolismo , Masculino , Potenciales de la Membrana , Arteria Cerebral Media/efectos de los fármacos , Arteria Cerebral Media/enzimología , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Bloqueadores de los Canales de Potasio/farmacología , Canales de Potasio Calcio-Activados/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/administración & dosificación , Ratas Sprague-Dawley , Transducción de Señal , Vasodilatación/efectos de los fármacos , Vasodilatadores/farmacologíaRESUMEN
Orai1 is one of the key components of store-operated Ca(2+) entry (SOCE) involved in diverse physiological functions. Orai1 may associate with other proteins to form a signaling complex. In the present study, we investigated the interaction between Orai1 and small conductance Ca(2+)-activated potassium channel 3 (SK3). With the use of RNA interference technique, we found that the SOCE and its associated membrane hyperpolarization were reduced while Orai1 was knocked down by a specific Orai1 siRNA in guinea pig gallbladder smooth muscle. However, with the use of isometric tension measurements, our results revealed that agonist-induced muscle contractility was significantly enhanced after Orai1 protein was knocked down or the tissue was treated by SK3 inhibitor apamin, but not affected by larger conductance Ca(2+)-activated potassium channel inhibitor iberiotoxin or intermediate conductance Ca(2+)-activated potassium channel inhibitor TRAM-34. In addition, in the presence of apamin, Orai1 siRNA had no additional effect on agonist-induced contraction. In coimmunoprecipitation experiment, SK3 and Orai1 pulled down each other. These data suggest that, Orai1 physically associated with SK3 to form a signaling complex in gallbladder smooth muscle. Ca(2+) entry via Orai1 activates SK3, resulting in membrane hyperpolarization in gallbladder smooth muscle. This hyperpolarizing effect of Orai1-SK3 coupling could serve to prevent excessive contraction of gallbladder smooth muscle in response to contractile agonists.
Asunto(s)
Canales de Calcio/metabolismo , Vesícula Biliar/metabolismo , Músculo Liso/metabolismo , Canales de Potasio de Pequeña Conductancia Activados por el Calcio/metabolismo , Animales , Agonistas de los Canales de Calcio/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Canales de Calcio/química , Canales de Calcio/genética , Señalización del Calcio , Vesícula Biliar/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Cobayas , Técnicas In Vitro , Masculino , Potenciales de la Membrana , Complejos Multiproteicos/química , Complejos Multiproteicos/metabolismo , Contracción Muscular/efectos de los fármacos , Contracción Muscular/fisiología , Músculo Liso/efectos de los fármacos , Interferencia de ARN , Canales de Potasio de Pequeña Conductancia Activados por el Calcio/agonistas , Canales de Potasio de Pequeña Conductancia Activados por el Calcio/antagonistas & inhibidoresRESUMEN
Introduction: The pattern of extraocular muscle involvement in ocular myasthenia gravis varies across different reports, diverging from our own observations. Thus, we employed two novel tools to discern this pattern. Methods: A retrospective analysis was conducted to collect and organize clinical data from 43 patients diagnosed with ocular myasthenia gravis. Each patient underwent both the computerized diplopia test and the Ocular Motor Nerve Palsy Scale assessment to evaluate the involvement of extraocular muscles. Results: Among the patients, there were 30 male and 13 female individuals, with a total of 113 affected extraocular muscles identified. Among all the affected extraocular muscles, the involvement of the levator palpebrae superioris muscle accounted for 35.40%, medial rectus muscle 7.7%, lateral rectus muscle 16.81%, superior rectus muscle 13.27%, inferior rectus muscle 12.39%, superior oblique muscle 1.77%, and inferior oblique muscle 2.65% of the total affected extraocular muscles. The positivity rates of the Neostigmine test were 89.19%, AChR antibody detection was 59.38%, and repetitive nerve stimulation was 34.38%. The AChR antibody positive rate among patients with only diplopia was 100%; among those with only ptosis, it was 80%; and among those with both diplopia and ptosis, it was 86.67%. Conclusion: The involvement of the extraocular muscles is not uniform. The levator palpebrae superioris exhibits the highest incidence rate, followed by the four rectus muscles and two oblique muscles. The inferior oblique involvement typically occurs when four or more EOMs are affected. Moreover, the levator palpebrae superioris and medial rectus show a higher tendency for bilateral involvement compared with other extraocular muscles.
RESUMEN
Large capacitive loading of electrodes induces massive error current and imperfect settling in the electrochemical signal acquisition process, leading to inaccurate acquisition results. To efficiently mitigate this inaccuracy, this paper presents a current-and-voltage dual-mode acquisition technique in which a voltage front-end (VFE) is employed to acquire the electrode voltage error and compensate the nonlinearity induced by the electrode capacitive loading. Therefore, the gain and bandwidth requirements of the current front end (CFE) can be relaxed to reduce the complexity and power consumption. With a relieved gain requirement, an inverter-based capacitive trans-impedance amplifier (IB-CTIA) is adopted to boost the input transconductance for low-noise design. By reusing the supply current, the IB-CTIA effectively achieves a low input-referred current noise of 3.9 pArms and a dynamic range (DR) of 126 dB with only 18-µW static power. The prototype chip is fabricated in a 180-nm CMOS process. Interleukin-6 immunoassays (IL-6) are implemented to verify the chip's performance. With the proposed nonlinear error compensation, the correlation coefficient of the detection result is improved from 0.951 to 0.980 and the limit of detection (LoD) is reduced from 8.31 pg/mL to 6.90 pg/mL.
RESUMEN
The bacterial community in sediment serves as an important indicator for assessing the environmental health of river ecosystems. However, the response of bacterial community structure and function in river basin sediment to different land use/cover changes has not been widely studied. To characterize changes in the structure, composition, and function of bacterial communities under different types of land use/cover, we studied the bacterial communities and physicochemical properties of the surface sediments of rivers. Surface sediment in cropland and built-up areas was moderately polluted with cadmium and had high nitrogen and phosphorus levels, which disrupted the stability of bacterial communities. Significant differences in the α-diversity of bacterial communities were observed among different types of land use/cover. Bacterial α-diversity and energy sources were greater in woodlands than in cropland and built-up areas. The functional patterns of bacterial communities were shown that phosphorus levels and abundances of pathogenic bacteria and parasites were higher in cropland than in the other land use/cover types; Urban activities have resulted in the loss of the denitrification function and the accumulation of nitrogen in built-up areas, and bacteria in forested and agricultural areas play an important role in nitrogen degradation. Differences in heavy metal and nutrient inputs driven by land use/cover result in variation in the composition, structure, and function of bacterial communities.
Asunto(s)
Bacterias , Monitoreo del Ambiente , Sedimentos Geológicos , Ríos , China , Ríos/microbiología , Ríos/química , Sedimentos Geológicos/microbiología , Sedimentos Geológicos/química , Fósforo/análisis , Nitrógeno/análisis , Agricultura , Microbiota , Contaminantes Químicos del Agua/análisis , EcosistemaRESUMEN
Degenerative tendon injuries are common clinical problems associated with overuse or aging, and understanding the mechanisms of tendon injury and regeneration can contribute to the study of tendon healing and repair. As a transcription factor, Mohawk (Mkx) is responsible for tendons development, yet, the roles of which in tendon damage remain mostly elusive. In this study, using Mkx overexpressed mice on long treadmill as an in vivo model and MkxOE Achilles tenocytes stimulated by equiaxial stretch as an in vitro model, we anaylsed the effects of Mkx overexpression on the tendon. Mkx and tendon tension strength were decreased after the expose to excessive mechanical forces, and Mkx overexpression protected the tendon from damage. Moreover, we revealed that the Wnt/ß-catenin activation, inflammation, and Runx2 expression were increased at the injured Achilles tendon, upregulated Mkx significantly reversed the increased Wnt/ß-catenin pathway, Tnf-α, Il-1ß, and Il-6 levels, and reduced tendon cell damage. However, Wnt3a, IWR and BIO had not significantly affected the Mkx expression in achilles tenocytes. In conclusion, Mkx is involved in tendon healing and protects the tendon from damage through suppressing Wnt/ß-catenin pathway, suggesting Mkx/Wnt/ß-catenin pathway may be potential therapeutic targets for tendon damage.
RESUMEN
Heat shock protein 105/110-kDa (Hsp105/110), a member of the Hsp70 super family of molecular chaperones, serves as a nucleotide exchange factor for Hsc70, independently prevents the aggregation of misfolded proteins, and functionally relates to Hsp90. We investigated the roles of human Hsp105α, the constitutively expressed isoform, in the biogenesis and quality control of the cystic fibrosis transmembrane conductance regulator (CFTR). In the endoplasmic reticulum (ER), Hsp105 facilitates CFTR quality control at an early stage in its biosynthesis but promotes CFTR post-translational folding. Deletion of Phe-508 (ΔF508), the most prevalent mutation causing cystic fibrosis, interferes with de novo folding of CFTR, impairing its export from the ER and accelerating its clearance in the ER and post-Golgi compartments. We show that Hsp105 preferentially associates with and stabilizes ΔF508 CFTR at both levels. Introduction of the Hsp105 substrate binding domain potently increases the steady state level of ΔF508 CFTR by reducing its early-stage degradation. This in turn dramatically enhances ΔF508 CFTR cell surface functional expression in cystic fibrosis airway epithelial cells. Although other Hsc70 nucleotide exchange factors such as HspBP1 and BAG-2 inhibit CFTR post-translational degradation in the ER through cochaperone CHIP, Hsp105 has a primary role promoting CFTR quality control at an earlier stage. The Hsp105-mediated multilevel regulation of ΔF508 CFTR folding and quality control provides new opportunities to understand how chaperone machinery regulates the homeostasis and functional expression of misfolded proteins in the cell. Future studies in this direction will inform therapeutics development for cystic fibrosis and other protein misfolding diseases.
Asunto(s)
Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Fibrosis Quística/metabolismo , Retículo Endoplásmico/metabolismo , Proteínas del Choque Térmico HSP110/metabolismo , Pliegue de Proteína , Proteolisis , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Línea Celular , Fibrosis Quística/genética , Fibrosis Quística/patología , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Retículo Endoplásmico/genética , Retículo Endoplásmico/patología , Células Epiteliales/citología , Células Epiteliales/patología , Proteínas del Choque Térmico HSC70/genética , Proteínas del Choque Térmico HSC70/metabolismo , Proteínas del Choque Térmico HSP110/genética , Humanos , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Estabilidad Proteica , Transporte de Proteínas , Mucosa Respiratoria/citología , Mucosa Respiratoria/patologíaRESUMEN
The study focused on the transformation of coal fly ash to zeolite A (ZA) as a potential carrier for the slow release of urea. After being treated with HCl aqueous solution and NaOH successively, SiO2 and Al2O3 were converted into sodium silicoaluminate. The obtained silicoaluminate was then heated with NaAlO2 in an aqueous NaOH solution at 70-110 °C for 3-18 h and zeolite A was successfully prepared according to the X-ray diffraction measurements. By changing the hydrothermal temperature and time, ZA could reach 237.3 mmol/100 g in maximum cation exchange capacity. ZA impregnated with urea (ZA-U) at a mass ratio of more than 5:1 exhibited slow release of urea and the kinetics release mechanism of ZA-U was proposed. The plant growth test proved that the slow release of urea from ZA-U can promote the growth of maize seedling.
Asunto(s)
Ceniza del Carbón , Zeolitas , Dióxido de Silicio , Hidróxido de Sodio , Urea , Agua , AdsorciónRESUMEN
In clinical practice, repairing osteochondral defects presents a challenge due to the varying biological properties of articular cartilages and subchondral bones. Thus, elucidating how spatial microenvironment-specific biomimetic scaffolds can be used to simultaneously regenerate osteochondral tissue is an important research topic. Herein, a novel bioinspired double-network hydrogel scaffold produced via 3D printing with tissue-specific decellularized extracellular matrix (dECM) and human adipose mesenchymal stem cell (MSC)-derived exosomes is described. The bionic hydrogel scaffolds promote rat bone marrow MSC attachment, spread, migration, proliferation, and chondrogenic and osteogenic differentiation in vitro, as determined based on the sustained release of bioactive exosomes. Furthermore, the 3D-printed microenvironment-specific heterogeneous bilayer scaffolds efficiently accelerate the simultaneous regeneration of cartilage and subchondral bone tissues in a rat preclinical model. In conclusion, 3D dECM-based microenvironment-specific biomimetics encapsulated with bioactive exosomes can serve as a novel cell-free recipe for stem cell therapy when treating injured or degenerative joints. This strategy provides a promising platform for complex zonal tissue regeneration whilst holding attractive clinical translation potential.
Asunto(s)
Exosomas , Andamios del Tejido , Ratas , Humanos , Animales , Osteogénesis , Hidrogeles , Cartílago , Regeneración Ósea , Impresión TridimensionalRESUMEN
Meniscus is a complex and crucial fibrocartilaginous tissue within the knee joint. Meniscal regeneration remains to be a scientific and translational challenge. We clarified that mesenchymal stem cells (MSCs) participated in meniscal maturation and regeneration using MSC-tracing transgenic mice model. Here, inspired by meniscal natural maturational and regenerative process, we developed an effective and translational strategy to facilitate meniscal regeneration by three-dimensionally printing biomimetic meniscal scaffold combining autologous synovium transplant, which contained abundant intrinsic MSCs. We verified that this facilitated anisotropic meniscus-like tissue regeneration and protected cartilage from degeneration in large animal model. Mechanistically, the biomechanics and matrix stiffness up-regulated Piezo1 expression, facilitating concerted activation of calcineurin and NFATc1, further activated YAP-pSmad2/3-SOX9 axis, and consequently facilitated fibrochondrogenesis of MSCs during meniscal regeneration. In addition, Piezo1 induced by biomechanics and matrix stiffness up-regulated collagen cross-link enzyme expression, which catalyzed collagen cross-link and thereby enhanced mechanical properties of regenerated tissue.
Asunto(s)
Menisco , Células Madre Mesenquimatosas , Animales , Ratones , Menisco/metabolismo , Fibrocartílago/metabolismo , Células Madre Mesenquimatosas/metabolismo , Colágeno/metabolismo , Modelos Animales , Ratones Transgénicos , Canales Iónicos/metabolismoRESUMEN
OBJECTIVE: To compare the severity of cartilage degeneration after meniscal tears between juvenile and adult rabbits. DESIGN: This study included 20 juvenile rabbits (2 weeks after birth) and 20 adult rabbits (6 months after birth). Meniscal tears were prepared in the anterior horn of medial menisci of right knees. Rabbits were sacrificed at 1, 3, 6, and 12 weeks postoperatively. Cartilage degenerations in the medial femoral condyle and medial tibial plateau were evaluated macroscopically and histologically. The semiquantitative assessment of cartilage degeneration was graded by macroscopic Outerbridge scoring system and histological Osteoarthritis Research Society International (OARSI) scoring system. RESULTS: In juvenile rabbits, the morphologically intact cartilage and normal extracellular matrix architecture were observed at the first week postoperatively. Mild uneven cartilage surface and toluidine blue depletion in the medial femoral condyle were observed on histological assessment at 3 weeks postoperatively. The worsened cartilage deterioration demonstrating chondral fibrillation, prominent cell death, and glycosaminoglycan (GAG) release was observed at 6 and 12 weeks postoperatively. In adult rabbits, only mild cartilage degeneration was observed in the medial femoral condyle at 12 weeks postoperatively. The outcomes of Outerbridge and OARSI scores were consistent with the aforementioned findings in juvenile and adult rabbits. CONCLUSIONS: Our study validated that earlier and more severe cartilage degenerations were observed in juvenile rabbits after meniscal tears compared with adult rabbits. Moreover, the post-tear cartilage degeneration demonstrated regional specificity corresponded to the tear position. However, caution is warranted when extrapolating results of animal models to humans.
Asunto(s)
Enfermedades de los Cartílagos , Traumatismos de la Rodilla , Osteoartritis , Adulto , Humanos , Animales , Conejos , Articulación de la Rodilla/patología , Enfermedades de los Cartílagos/patología , Traumatismos de la Rodilla/cirugía , Meniscos Tibiales/cirugía , Meniscos Tibiales/patología , Tibia/patología , Osteoartritis/patologíaRESUMEN
FK506-binding protein 38 (FKBP38), a membrane-anchored, tetratricopeptide repeat (TPR)-containing immunophilin, associates with nascent plasma membrane ion channels in the endoplasmic reticulum (ER). It promotes the maturation of the human ether-à-go-go-related gene (HERG) potassium channel and maintains the steady state level of the cystic fibrosis transmembrane conductance regulator (CFTR), but the underlying mechanisms remain unclear. Using a combination of steady state and pulse-chase analyses, we show that FKBP38 knockdown increases protein synthesis but inhibits the post-translational folding of CFTR, leading to reduced steady state levels of CFTR in the ER, decreased processing, and impaired cell surface functional expression in Calu-3 human airway epithelial cells. The membrane anchorage of FKBP38 is necessary for the inhibition of protein synthesis but not for CFTR post-translational folding. In contrast, the peptidylprolyl cis/trans isomerase active site is utilized to promote CFTR post-translational folding but is not important for regulation of protein synthesis. Uncoupling FKBP38 from Hsp90 by substituting a conserved lysine in the TPR domain modestly enhances CFTR maturation and further reduces its synthesis. Removing the N-terminal glutamate-rich domain (ERD) slightly enhances CFTR synthesis but reduces its maturation, suggesting that the ERD contributes to FKBP38 biological activities. Our data support a dual role for FKBP38 in regulating CFTR synthesis and post-translational folding. In contrast to earlier prediction but consistent with in vitro enzymological studies, FKBP38 peptidylprolyl cis/trans isomerase plays an important role in membrane protein biogenesis on the cytoplasmic side of the ER membrane, whose activity is negatively regulated by Hsp90 through the TPR domain.
Asunto(s)
Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Retículo Endoplásmico/metabolismo , Proteínas de Unión a Tacrolimus/metabolismo , Línea Celular , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Proteínas HSP90 de Choque Térmico/genética , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Immunoblotting , Unión Proteica , Pliegue de Proteína , ARN Interferente Pequeño/genética , Proteínas de Unión a Tacrolimus/genéticaRESUMEN
Objectives: To explore independent risk factors with good and early predictive power for SAP severity and prognosis. Methods: Patients with SAP were enrolled at Central South University Xiangya Hospital between April 2017 and May 2021 and used as the training cohort. From June 2021 to February 2022, all patients with SAP were defined as external patients for validation. Patients were grouped by survival status at a 30-day posthospital admission and then compared in terms of basic information and laboratory tests to screen the independent risk factors. Results: A total of 249 patients with SAP were enrolled in the training cohort. The all-cause mortality rate at a 30-day postadmission was 25.8% (51/198). Blood urea nitrogen (BUN) levels were significantly higher in the mortality group (20.45 [interquartile range (IQR), 19.7] mmol/L) than in the survival group (6.685 [IQR, 6.3] mmol/L; P < 0.001). After propensity score matching (PSM), the BUN level was still higher in the mortality group than in the survival group (18.415 [IQR, 19.555] mmol/L vs. 10.63 [IQR, 6.03] mmol/L; P = 0.005). The area under the curve (AUC) of the receiver operating characteristic curve (ROC) of BUN was 0.820 (95% confidence interval, 0.721-0.870; P < 0.001). The optimal BUN level cut-off for predicting a 30-day all-cause mortality was 10.745 mmol/L. Moreover, patients with SAP were grouped according to BUN levels and stratified according to optimal cut-off value. Patients with high BNU levels were associated with significantly higher rates of invasive mechanical ventilation (before PSM: 61.8% vs. 20.6%, P < 0.001; after PSM: 71.1% vs. 32%, P = 0.048) and a 30-day all-cause mortality (before PSM: 44.9% vs. 6.9%, P < 0.001; after PSM: 60% vs. 34.5%, P = 0.032) than those with low BNU levels before or after PSM. The effectiveness of BUN as a prognostic marker was further validated using ROC curves for the external validation set (n = 49). The AUC of BUN was 0.803 (95% CI, 0.655-0.950; P = 0.011). It showed a good ability to predict a 30-day all-cause mortality in patients with SAP. We also observed similar results regarding disease severity, including the Acute Physiology and Chronic Health Evaluation II score (before PSM: 16 [IQR, 8] vs. 8 [IQR, 6], P < 0.001; after PSM: 18 [IQR, 10] vs. 12 [IQR, 7], P < 0.001), SOFA score (before PSM: 7 [IQR, 5] vs. 3 [IQR, 3], P < 0.001; after PSM: 8 [IQR, 5] vs. 5 [IQR, 3.5], P < 0.001), and mMarshall score (before PSM: 4 [IQR, 3] vs. 3 [IQR, 1], P < 0.001; after PSM: 5 [IQR, 2.5] vs. 3 [IQR, 1], P < 0.001). There was significant increase in intensive care unit occupancy in the high BUN level group before PSM (93.3% vs. 73.1%, P < 0.001), but not after PSM (97.8% vs. 86.2%, P = 0.074). Conclusions: Our results showed that BUN levels within 24 h after hospital admission were independent risk factors for a 30-day all-cause death in patients with SAP.
Asunto(s)
Pancreatitis , Enfermedad Aguda , Nitrógeno de la Urea Sanguínea , Humanos , Pancreatitis/diagnóstico , Pronóstico , Curva ROC , Estudios RetrospectivosRESUMEN
BACKGROUND: Acute respiratory distress syndrome (ARDS) is a severe complication among patients with severe acute pancreatitis (SAP), which may be associated with increased mortality in hospitalized patients. Thus, an effective model to predict ARDS in patients with SAP is urgently required. METHODS: We retrospectively analyzed the data from the patients with SAP who recruited in Xiangya Hospital between April 2017 and May 2021. Patients meeting the Berlin definition of ARDS were categorized into the ARDS group. Logistic regression models and a nomogram were utilized in the study. Descriptive statistics, logistic regression models, and a nomogram were used in the current study. RESULTS: Comorbidity of ARDS occurred in 109 (46.58%) of 234 patients with SAP. The SAP patients with ARDS group had a higher 60-day mortality rate, an increased demand for invasive mechanical ventilation, and a longer intensive care unit (ICU) stay than those without ARDS (p < .001 for all). Partial pressure of oxygen (PaO2): fraction of inspired oxygen (FiO2) < 200, platelets <125 × 109/L, lactate dehydrogenase >250 U/L, creatinine >111 mg/dL, and procalcitonin >0.5 ng/mL were independent risk variables for development of ARDS in SAP patients. The area under the curve for the model was 0.814, and the model fit was acceptable [p = .355 (Hosmer-Lemeshow)]. Incorporating these 5 factors, a nomogram was established with sufficient discriminatory power (C-index 0.814). Calibration curve indicated the proper discrimination and good calibration in the predicting nomogram model. CONCLUSION: The prediction nomogram for ARDS in patients with SAP can be applied using clinical common variables after the diagnosis of SAP. Future studies would be warranted to verify the potential clinical benefits of this model.
Asunto(s)
Pancreatitis , Síndrome de Dificultad Respiratoria , Enfermedad Aguda , Humanos , Oxígeno , Pancreatitis/complicaciones , Pancreatitis/diagnóstico , Pancreatitis/terapia , Síndrome de Dificultad Respiratoria/diagnóstico , Síndrome de Dificultad Respiratoria/epidemiología , Síndrome de Dificultad Respiratoria/terapia , Estudios RetrospectivosRESUMEN
Background: Synovium has been confirmed to be the primary contributor to meniscal repair. Particulated Juvenile Allograft Cartilage (PJAC) has demonstrated promising clinical effect on repairing cartilage. The synergistic effect of synovium and PJAC transplant on meniscal fibrocartilaginous repair is unclear. We hypothesize that the transplantation of synovium and PJAC synergistically facilitates meniscal regeneration and the donor cells within graft tissues still survive in the regenerated tissue at the last follow up (16 weeks postoperatively). Methods: The study included 24 mature female rabbits, which were randomly divided into experimental and control groups. A cylindrical full-thickness defect measuring 2.0 âmm was prepared in the avascular portion of the anterior horn of medial meniscus in both knees. The synovium and PJAC transplant were harvested from juvenile male rabbits (2 months after birth). The experimental group received synovium and PJAC transplant encapsulated with fibrin gel. The control groups received synovium transplant encapsulated with fibrin gel, pure fibrin gel and nothing. The macroscopic, imageological and histological evaluations of repaired tissue were performed at 8 weeks and 16 weeks postoperatively. The in situ hybridization (ISH) of male-specific sex-determining region Y-linked (SRY) gene was performed to detect the transplanted cells. Results: The regenerated tissue in experimental group showed superior structural integrity, superficial smoothness, and marginal integration compared to control groups at 8 weeks or 16 weeks postoperatively. More meniscus-like fibrochondrocytes filled the repaired tissue in the experimental group, and the matrix surrounding these cell clusters demonstrated strongly positive safranin O and type 2 collagen immunohistochemistry staining. By SRY gene ISH, the positive SRY signal of experimental group could be detected at 8 weeks (75.72%, median) and 16 weeks (48.69%, median). The expression of SOX9 in experimental group was the most robust, with median positive rates of 65.52% at 8 weeks and 67.55% at 16 weeks. Conclusion: The transplantation of synovium and PJAC synergistically facilitates meniscal regeneration. The donor cells survive for at least 16 weeks in the recipient. The translational potential of this article: This study highlighted the positive effect of PJAC and synovium transplant on meniscal repair. We also clarified the potential repair mechanisms reflected by the survival of donor cells and upregulated expression of meniscal fibrochondrocytes related genes. Thus, based on our study, further clinical experiments are needed to investigate synovium and PJAC transplant as a possible treatment to meniscal defects.