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1.
Br J Haematol ; 2024 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-39087522

RESUMEN

Acute myeloid leukaemia (AML) is a heterogeneous disease characterized by complex molecular and cytogenetic abnormalities. Pro-oxidant cellular redox status is a common hallmark of AML cells, providing a rationale for redox-based anticancer strategy. We previously discovered that auranofin (AUF), initially used for the treatment of rheumatoid arthritis and repositioned for its anticancer activity, can synergize with a pharmacological concentration of vitamin C (VC) against breast cancer cell line models. In this study, we observed that this drug combination synergistically and efficiently killed cells of leukaemic cell lines established from different myeloid subtypes. In addition to an induced elevation of reactive oxygen species and ATP depletion, a rapid dephosphorylation of 4E-BP1 and p70S6K, together with a strong inhibition of protein synthesis were early events in response to AUF/VC treatment, suggesting their implication in AUF/VC-induced cytotoxicity. Importantly, a study on 22 primary AML specimens from various AML subtypes showed that AUF/VC combinations at pharmacologically achievable concentrations were effective to eradicate primary leukaemic CD34+ cells from the majority of these samples, while being less toxic to normal cord blood CD34+ cells. Our findings indicate that targeting the redox vulnerability of AML with AUF/VC combinations could present a potential anti-AML therapeutic approach.

2.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 53(2): 194-200, 2024 Apr 25.
Artículo en Inglés, Zh | MEDLINE | ID: mdl-38268403

RESUMEN

OBJECTIVES: To isolate a potassium ion channel Kv4.1 inhibitor from centipede venom, and to determine its sequence and structure. METHODS: Ion-exchange chromatography and reversed-phase high-performance liquid chromatography were performed to separate and purify peptide components of centipede venom, and their inhibiting effect on Kv4.1 channel was determined by whole-cell patch clamp recording. The molecular weight of isolated peptide Kv4.1 channel inhibitor was identified with matrix assisted laser desorption ionization-time-of-flight mass spectrometry; its primary sequence was determined by Edman degradation sequencing and two-dimensional mass spectrometry; its structure was established based on iterative thread assembly refinement online analysis. RESULTS: A peptide SsTx-P2 was separated from centipede venom with the molecular weight of 6122.8, and its primary sequence consists of 53 amino acid residues NH2-ELTWDFVRTCCKLFPDKSECTKACATEFTGGDESRLKDVWPRKLRSGDSRLKD-OH. Peptide SsTx-P2 potently inhibited the current of Kv4.1 channel transiently transfected in HEK293 cell, with 1.0 µmol/L SsTx-P2 suppressing 95% current of Kv4.1 channel. Its structure showed that SsTx-P2 shared a conserved helical structure. CONCLUSIONS: The study has isolated a novel peptide SsTx-P2 from centipede venom, which can potently inhibit the potassium ion channel Kv4.1 and displays structural conservation.


Asunto(s)
Secuencia de Aminoácidos , Venenos de Artrópodos , Canales de Potasio Shal , Animales , Humanos , Venenos de Artrópodos/química , Venenos de Artrópodos/farmacología , Datos de Secuencia Molecular , Péptidos/farmacología , Péptidos/aislamiento & purificación , Péptidos/química , Bloqueadores de los Canales de Potasio/farmacología , Bloqueadores de los Canales de Potasio/aislamiento & purificación , Bloqueadores de los Canales de Potasio/química , Canales de Potasio Shal/antagonistas & inhibidores , Quilópodos/química
3.
Biomacromolecules ; 24(4): 1662-1674, 2023 04 10.
Artículo en Inglés | MEDLINE | ID: mdl-36913719

RESUMEN

Numerous metabolic reactions and pathways use adenosine 5'-triphosphate (ATP) as an energy source and as a phosphorous or pyrophosphorous donor. Based on three-dimensional (3D)-printing, enzyme immobilization can be used to improve ATP regeneration and operability and reduce cost. However, due to the relatively large mesh size of 3D-bioprinted hydrogels soaked in a reaction solution, the lower-molecular-weight enzymes cannot avoid leaking out of the hydrogels readily. Here, a chimeric adenylate-kinase-spidroin (ADK-RC) is created, with ADK serving as the N-terminal domain. The chimera is capable of self-assembling to form micellar nanoparticles at a higher molecular scale. Although fused to spidroin (RC), ADK-RC remains relatively consistent and exhibits high activity, thermostability, pH stability, and organic solvent tolerance. Considering different surface-to-volume ratios, three shapes of enzyme hydrogels are designed, 3D bioprinted, and measured. In addition, a continuous enzymatic reaction demonstrates that ADK-RC hydrogels have higher specific activity and substrate affinity but a lower reaction rate and catalytic power compared to free enzymes in solution. With ATP regeneration, the ADK and ADK-RC hydrogels significantly increase the production of d-glucose-6-phosphate and obtain an efficient usage frequency. In conclusion, enzymes fused to spidroin might be an efficient strategy for maintaining activity and reducing leakage in 3D-bioprinted hydrogels under mild conditions.


Asunto(s)
Adenilato Quinasa , Fibroínas , Adenilato Quinasa/química , Adenilato Quinasa/metabolismo , Hidrogeles , Adenosina Trifosfato/química , Catálisis
4.
Prep Biochem Biotechnol ; 53(8): 914-922, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36573266

RESUMEN

Due to their advantages in structural stability and versatility, cysteine-rich peptides, which are secreted from the venom glands of venomous animals, constitute a naturally occurring pharmaceutical arsenal. However, the correct folding of disulfide bonds is a challenging task in the prokaryotic expression system like Escherichia coli due to the reducing environment. Here, a secretory expression plasmid pSE-G1M5-SUMO-HWTX-I for the spider neurotoxin huwentoxin-I (HWTX-I) with three disulfides as a model of cysteine-rich peptides was constructed. By utilizing the signal peptide G1M5, the fusion protein 6 × His-SUMO-HWTX-I was successfully secreted into extracellular medium of BL21(DE3). After enrichment using cation-exchange chromatography and purification utilizing the Ni-NTA column, 6 × His-SUMO-HWTX-I was digested via Ulp1 kinase to release recombinant HWTX-I (rHWTX-I), which was further purified utilizing RP-HPLC. Finally, both impurities with low and high molecular weights were completely removed. The molecular mass of rHWTX-I was identified as being 3750.8 Da, which was identical to natural HWTX-I with three disulfide bridges. Furthermore, by utilizing whole-cell patch clamp, the sodium currents of hNav1.7 could be inhibited by rHWTX-I and the IC50 value was 419 nmol/L.


Asunto(s)
Venenos de Araña , Arañas , Animales , Neurotoxinas/química , Neurotoxinas/farmacología , Cisteína/metabolismo , Arañas/química , Arañas/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Venenos de Araña/genética , Venenos de Araña/química , Venenos de Araña/metabolismo , Péptidos/metabolismo , Disulfuros/metabolismo
5.
Zhongguo Dang Dai Er Ke Za Zhi ; 24(2): 132-140, 2022 Feb 15.
Artículo en Inglés, Zh | MEDLINE | ID: mdl-35209977

RESUMEN

OBJECTIVES: To investigate the incidence of extrauterine growth retardation (EUGR) and its risk factors in very preterm infants (VPIs) during hospitalization in China. METHODS: A prospective multicenter study was performed on the medical data of 2 514 VPIs who were hospitalized in the department of neonatology in 28 hospitals from 7 areas of China between September 2019 and December 2020. According to the presence or absence of EUGR based on the evaluation of body weight at the corrected gestational age of 36 weeks or at discharge, the VPIs were classified to two groups: EUGR group (n=1 189) and non-EUGR (n=1 325). The clinical features were compared between the two groups, and the incidence of EUGR and risk factors for EUGR were examined. RESULTS: The incidence of EUGR was 47.30% (1 189/2 514) evaluated by weight. The multivariate logistic regression analysis showed that higher weight growth velocity after regaining birth weight and higher cumulative calorie intake during the first week of hospitalization were protective factors against EUGR (P<0.05), while small-for-gestational-age birth, prolonged time to the initiation of total enteral feeding, prolonged cumulative fasting time, lower breast milk intake before starting human milk fortifiers, prolonged time to the initiation of full fortified feeding, and moderate-to-severe bronchopulmonary dysplasia were risk factors for EUGR (P<0.05). CONCLUSIONS: It is crucial to reduce the incidence of EUGR by achieving total enteral feeding as early as possible, strengthening breastfeeding, increasing calorie intake in the first week after birth, improving the velocity of weight gain, and preventing moderate-severe bronchopulmonary dysplasia in VPIs.


Asunto(s)
Recien Nacido Prematuro , Recién Nacido de muy Bajo Peso , Femenino , Retardo del Crecimiento Fetal , Edad Gestacional , Hospitalización , Humanos , Incidencia , Lactante , Recién Nacido , Estudios Prospectivos , Factores de Riesgo
6.
Exp Cell Res ; 383(2): 111539, 2019 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-31369751

RESUMEN

Desminopathies are a type of myofibrillar myopathy resulting from mutations in DES, encoding the intermediate filament protein desmin. They display heterogeneous phenotypes, suggesting environment influences. Patient muscle proteins show oxidative features linking oxidative stress, protein aggregation, and abnormal protein deposition. To improve understanding of redox balance in desminopathies, we further developed cellular models of four pathological mutants localized in 2B helical domain (the most important region for desmin polymerization) to explore desmin behavior upon oxidative stress. We show that the mutations desQ389P and desD399Y share common stress-induced aggregates, desR406W presents more scattered cytoplasmic aggregative pattern, and pretreatment with N-acetyl-l-cysteine (NAC), an antioxidant molecule, prevents all type of aggregation. Mutants desD399Y and desR406W had delayed oxidation kinetics following H2O2 stress prevented by NAC pretreatment. Further, we used AAV-injected mouse models to confirm in vivo effects of N-acetyl-l-cysteine. AAV-desD399Y-injected muscles displayed similar physio-pathological characteristics as observed in patients. However, after 2 months of NAC treatment, they did not have reduced aggregates. Finally, in both models, stress induced some post-translational modifications changing Isoelectric Point, such as potential hyperphosphorylations, and/or molecular weight of human desmin by proteolysis. However, each mutant presented its own pattern that seemed to be post-aggregative. In conclusion, our results indicate that individual desmin mutations have unique pathological molecular mechanisms partly linked to alteration of redox homeostasis. Integrating these mutant-specific behaviors will be important when considering future therapeutics.


Asunto(s)
Cardiomiopatías/genética , Cardiomiopatías/metabolismo , Desmina , Músculo Esquelético/metabolismo , Distrofias Musculares/genética , Distrofias Musculares/metabolismo , Oxidación-Reducción , Sustitución de Aminoácidos/genética , Animales , Antioxidantes/metabolismo , Cardiomiopatías/patología , Células Cultivadas , Desmina/genética , Desmina/metabolismo , Modelos Animales de Enfermedad , Homeostasis/genética , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Musculares/metabolismo , Músculo Esquelético/patología , Distrofias Musculares/patología , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Estrés Oxidativo/genética , Procesamiento Proteico-Postraduccional/genética
7.
Anal Chem ; 91(22): 14561-14568, 2019 11 19.
Artículo en Inglés | MEDLINE | ID: mdl-31638767

RESUMEN

The quantification of cellular deoxyribonucleoside triphosphate (dNTP) levels is important for studying pathologies, genome integrity, DNA repair, and the efficacy of pharmacological drug treatments. Current standard methods, such as enzymatic assays or high-performance liquid chromatography, are complicated, costly, and labor-intensive, and alternative techniques that simplify dNTP quantification would present very useful complementary approaches. Here, we present a dNTP assay based on isothermal rolling circle amplification (RCA) and rapid time-gated Förster resonance energy transfer (TG-FRET), which used a commercial clinical plate reader system. Despite the relatively simple assay format, limits of detection down to a few picomoles of and excellent specificity for each dNTP against the other dNTPs, rNTPs, and dUTP evidenced the strong performance of the assay. Direct applicability of RCA-FRET to applied nucleic acid research was demonstrated by quantifying all dNTPs in CEM-SS leukemia cells with and without hydroxyurea or auranofin treatment. Both pharmacological agents could reduce the dNTP production in a time- and dose-dependent manner. RCA-FRET provides simple, rapid, sensitive, and specific quantification of intracellular dNTPs and has the potential to become an advanced tool for both fundamental and applied dNTP research.


Asunto(s)
Desoxirribonucleótidos/análisis , Transferencia Resonante de Energía de Fluorescencia/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , Auranofina/farmacología , Secuencia de Bases , Línea Celular Tumoral , Inhibidores Enzimáticos/farmacología , Humanos , Hidroxiurea/farmacología , Límite de Detección , Prueba de Estudio Conceptual , Ribonucleótido Reductasas/antagonistas & inhibidores , Sensibilidad y Especificidad , Reductasa de Tiorredoxina-Disulfuro/antagonistas & inhibidores
8.
Plasmid ; 105: 102431, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-31319110

RESUMEN

Pinene is a monoterpene with wide industrial applications, especially as a promising high energy-density jet fuel. Traditional production of pinene on an industrial scale is material consumptive and has a low yield. As an alternative, microbial organisms have been engineered though advanced synthetic biological techniques to produce a variety of heterologous products, including pinene. Here, we investigated the stability of genetic circuits encoding the pinene producing pathway during fermentation and its relationship to the pinene titer. By replacing scar sequences in the genetic elements and modifying the genome of E. coli strain MG1655, plasmid loss caused by serious metabolic burden was eliminated, generating a remarkable increase in the pinene titer. Furthermore, the heterologous mevalonate pathway was analyzed by overexpression of enzymes and intermediates monitoring. Optimized pathway plasmids and strains were combined to increase the pinene titer to 104.6 mg/L.


Asunto(s)
Vías Biosintéticas/genética , Monoterpenos/metabolismo , Plásmidos/genética , Recombinación Genética/genética , Escherichia coli/genética , Fermentación , Plásmidos/metabolismo
9.
Biotechnol Lett ; 41(10): 1147-1154, 2019 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-31428906

RESUMEN

OBJECTIVES: Developing a dynamic regulation strategy is an essential step in establishing an automatic control system for manipulating metabolic fluxes and cellular behaviors. To broaden the extent of the application, a system that can generally control any gene of interest is demanded. RESULTS: Through characterization and optimization, the strategy repressed the immediate expression incrementally from 0 to 90% during culturing. Moreover, by changing single base pair in the lux box of the Plux promoter, the degree of repression of the target genomic gene was tuned to a difference of 70%. This strategy is expected to control metabolic flux without disrupting cell growth. CONCLUSIONS: We engineered bacterial small RNA to develop a pathway-independent strategy that can dynamically repress the expression of any gene at the posttranscription level.


Asunto(s)
Escherichia coli/genética , Escherichia coli/metabolismo , Regulación Bacteriana de la Expresión Génica , Percepción de Quorum , ARN Bacteriano/biosíntesis , ARN Pequeño no Traducido/biosíntesis
10.
Int J Mol Sci ; 20(16)2019 Aug 09.
Artículo en Inglés | MEDLINE | ID: mdl-31405050

RESUMEN

Compelling evidence supports a tight link between oxidative stress and protein aggregation processes, which are noticeably involved in the development of proteinopathies, such as Alzheimer's disease, Parkinson's disease, and prion disease. The literature is tremendously rich in studies that establish a functional link between both processes, revealing that oxidative stress can be either causative, or consecutive, to protein aggregation. Because oxidative stress monitoring is highly challenging and may often lead to artefactual results, cutting-edge technical tools have been developed recently in the redox field, improving the ability to measure oxidative perturbations in biological systems. This review aims at providing an update of the previously known functional links between oxidative stress and protein aggregation, thereby revisiting the long-established relationship between both processes.


Asunto(s)
Estrés Oxidativo , Agregación Patológica de Proteínas/metabolismo , Proteínas/metabolismo , Enfermedad de Alzheimer/metabolismo , Animales , Humanos , Enfermedad de Parkinson/metabolismo , Enfermedades por Prión/metabolismo , Agregado de Proteínas
11.
Apoptosis ; 23(7-8): 377-387, 2018 08.
Artículo en Inglés | MEDLINE | ID: mdl-29926313

RESUMEN

In the early stage of acute pancreatitis (AP), abundant cytokines induced by local pancreatic inflammation enter the bloodstream, further cause systemic inflammatory response syndrome (SIRS) by "trigger effect", which eventually leads to multiple organ dysfunction syndrome (MODS). During SIRS and MODS, the intestinal barrier function was seriously damaged accompanied by the occurrence of gut-derived infection which forms a "second hit summit" by inflammatory overabundance. Gastrointestinal microecology, namely the biologic barrier, could be transformed into a pathogenic state, which is called microflora dysbiosis when interfered by the inflammatory stress during AP. More and more evidences indicate that gastrointestinal microflora dysbiosis plays a key role in "the second hit" induced by AP gut-derived infection. Therefore, the maintenance of gastrointestinal microecology balance is likely to provide an effective method in modulating systemic infection of AP. This article reviewed the progress of gastrointestinal microecology in AP to provide a reference for deeply understanding the pathogenic mechanisms of AP and identifying new therapeutic targets.


Asunto(s)
Apoptosis/fisiología , Insuficiencia Multiorgánica/patología , Pancreatitis/patología , Sepsis/patología , Enfermedad Aguda , Animales , Citocinas/metabolismo , Humanos , Sepsis/complicaciones
12.
J Membr Biol ; 250(3): 315-325, 2017 06.
Artículo en Inglés | MEDLINE | ID: mdl-28597209

RESUMEN

Tarantula toxins compose an important class of spider toxins that target ion channels, and some are known to interact with lipid membranes. In this study, we focus on a tarantula toxin, Jingzhaotoxin-III (JZTx-III) that specifically targets the cardiac voltage-gated sodium channel Na[Formula: see text]1.5 and is suspected to be able to interact with lipid membranes. Here, we use an all-atom model and long-term molecular dynamics simulations to investigate the interactions between JZTx-III and lipid membranes of different compositions. Trajectory analyses show that JZTx-III has no substantial interaction with the neutral 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipids, but binds to membranes containing negatively charged 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG). The most intriguing observations in our simulation are the different interactions between the toxin and the membrane in the mixed and pure POPG membrane systems. The POPC/POPG mixed membrane undergoes a phase transition to a rippled phase upon binding of the toxin, while the pure POPG membrane has no apparent change. Moreover, the binding of JZTx-III to both of the mixture and the pure POPG membrane systems induce small conformational changes. The sequence alignment shows that JZTx-III may not partition into the lipid bilayer due to the mutations of a C-terminal hydrophobic residue and some charged residues that affect toxin orientation. Taken together, JZTx-III and lipid membranes have unique effects on each other that may facilitate the specific binding of JZTx-III to Na[Formula: see text]1.5. This computational study also enriches our understanding of the potential complex interactions between spider toxins and lipid membranes.


Asunto(s)
Membrana Dobles de Lípidos/química , Lípidos de la Membrana/química , Simulación de Dinámica Molecular , Venenos de Araña/química , Potenciales de la Membrana , Fosfatidilcolinas/química
13.
Prep Biochem Biotechnol ; 47(2): 158-162, 2017 Feb 07.
Artículo en Inglés | MEDLINE | ID: mdl-27249514

RESUMEN

In the present study, we used Escherichia coli to produce recombinant Hainantoxin-III (rHNTX-III), a 33-amino acid peptic toxin from the tarantula spider Haplopelma hainanum. The toxin has three pairs of disulfide bonds. A pET-HS-HNTX-III vector was constructed and transformed into the E. coli strain SHuffleTM. rHNTX-III was expressed using auto-induction medium. After using a Ni-NTA column, the expressed fusion protein was digested using SUMO protease (ULP1) to remove the HIS-SUMO tag, and then RP-HPLC and ultrafiltration were used for further purification. Then the rHNTX-III was identified by MALDI-TOF/TOF mass spectrometry. The purified rHNTX-III was further analyzed using a whole-cell patch-clamp assay. It was shown that the rHNTX-III was able to block currents generated by human Nav1.7 (hNav1.7) at an IC50 of 225 nM and also have high selectivity for different voltage-gated sodium channels. Therefore, it has very similar activity to the natural one.


Asunto(s)
Escherichia coli/genética , Venenos de Araña/genética , Cromatografía de Afinidad , Electroforesis en Gel de Poliacrilamida , Células HEK293 , Humanos , Técnicas de Placa-Clamp , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Venenos de Araña/aislamiento & purificación , Ultrafiltración
14.
J Huazhong Univ Sci Technolog Med Sci ; 35(1): 54-57, 2015 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-25673193

RESUMEN

The purpose of this study was to observe the expression of LINGO-1 after cerebral ischemia, investigate the effects of retinoic acid (RA) on the expression of LINGO-1 and GAP-43, and the number of synapses, and to emplore the repressive effect of LINGO-1 on neural regeneration after cerebral ischemia. The model of permanent focal cerebral ischemia was established by the modified suture method of middle cerebral artery occlusion (MCAO) in Sprague-Dawley (SD) rats. The expression of LINGO-1 was detected by Western blotting and that of GAP-43 by immunohistochemistry. The number of synapses was observed by transmission electron microscopy. The SD rats were divided into three groups: sham operation (sham) group, cerebral ischemia (CI) group and RA treatment (RA) group. The results showed that the expression level of LINGO-1 at 7th day after MCAO in sham, CI and RA groups was 0.266 ± 0.019, 1.215 ± 0.063 and 0.702 ± 0.081, respectively (P<0.01). The number of Gap-43-positive nerve cells at 7th day after MCAO in sham, CI and RA group was 0, 59.08 ± 1.76 and 76.20 ± 3.12 per high power field, respectively (P<0.05). The number of synapses at 7th day after MCAO was 8.42 ± 0.13, 1.74 ± 0.37 and 5.39 ± 0.26 per µm², respectively (P<0.05). It is concluded that LINGO-1 expression is up-regulated after cerebral ischemia, and RA inhibits the expression of LINGO-1, promotes the expression of GAP-43 and increases the number of synapses. It suggests that LINGO-1 may be involved in the pathogenesis of cerebral ischemia, which may provide an experimenal basis for LINGO-1 antogonist, RA, for the treatment of cerebral ischemia.


Asunto(s)
Isquemia Encefálica/metabolismo , Expresión Génica/efectos de los fármacos , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Tretinoina/farmacología , Animales , Western Blotting , Proteína GAP-43/genética , Proteína GAP-43/metabolismo , Masculino , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Ratas , Ratas Sprague-Dawley
15.
Mater Today Bio ; 24: 100915, 2024 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-38188648

RESUMEN

Objective: Abdominal wall hernias are common abdominal diseases, and effective hernia repair is challenging. In clinical practice, synthetic meshes are widely applied for repairing abdominal wall hernias. However, postoperative complications, such as inflammation and adhesion, are prevalent. Although biological meshes can solve this problem to a certain extent, they face the problems of heterogeneity, rapid degradation rate, ordinary mechanical properties, and high-cost. Here, a novel electrospinning mesh composed of polylactic acid and silk fibroin (PLA-SF) for repairing abdominal wall hernias was manufactured with good physical properties, biocompatibility and low production cost. Materials and methods: FTIR and EDS were used to demonstrate that the PLA-SF mesh was successfully synthesized. The physicochemical properties of PLA-SF were detected by swelling experiments and in vitro degradation experiments. The water contact angle reflected the hydrophilicity, and the stress‒strain curve reflected the mechanical properties. A rat abdominal wall hernia model was established to observe degradation, adhesion, and inflammation in vivo. In vitro cell mesh culture experiments were used to detect cytocompatibility and search for affected biochemical pathways. Results: The PLA-SF mesh was successfully synthesized and did not swell or degrade over time in vitro. It had a high hydrophilicity and strength. The PLA-SF mesh significantly reduced abdominal inflammation and inhibited adhesion formation in rat models. The in vitro degradation rate of the PLA-SF mesh was slower than that of tissue remodeling. Coculture experiments suggested that the PLA-SF mesh reduced the expression of inflammatory factors secreted by fibroblasts and promoted fibroblast proliferation through the TGF-ß1/Smad pathway. Conclusion: The PLA-SF mesh had excellent physicochemical properties and biocompatibility, promoted hernia repair of the rat abdominal wall, and reduced postoperative inflammation and adhesion. It is a promising mesh and has potential for clinical application.

16.
Metallomics ; 16(5)2024 05 02.
Artículo en Inglés | MEDLINE | ID: mdl-38744662

RESUMEN

Iron-sulfur (Fe-S) clusters are an essential and ubiquitous class of protein-bound prosthetic centers that are involved in a broad range of biological processes (e.g. respiration, photosynthesis, DNA replication and repair and gene regulation) performing a wide range of functions including electron transfer, enzyme catalysis, and sensing. In a general manner, Fe-S clusters can gain or lose electrons through redox reactions, and are highly sensitive to oxidation, notably by small molecules such as oxygen and nitric oxide. The [2Fe-2S] and [4Fe-4S] clusters, the most common Fe-S cofactors, are typically coordinated by four amino acid side chains from the protein, usually cysteine thiolates, but other residues (e.g. histidine, aspartic acid) can also be found. While diversity in cluster coordination ensures the functional variety of the Fe-S clusters, the lack of conserved motifs makes new Fe-S protein identification challenging especially when the Fe-S cluster is also shared between two proteins as observed in several dimeric transcriptional regulators and in the mitoribosome. Thanks to the recent development of in cellulo, in vitro, and in silico approaches, new Fe-S proteins are still regularly identified, highlighting the functional diversity of this class of proteins. In this review, we will present three main functions of the Fe-S clusters and explain the difficulties encountered to identify Fe-S proteins and methods that have been employed to overcome these issues.


Asunto(s)
Proteínas Hierro-Azufre , Proteínas Hierro-Azufre/metabolismo , Proteínas Hierro-Azufre/química , Proteínas Hierro-Azufre/genética , Oxidación-Reducción
17.
J Inorg Biochem ; 255: 112535, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38527404

RESUMEN

Human mitoNEET (mNT) and CISD2 are two NEET proteins characterized by an atypical [2Fe-2S] cluster coordination involving three cysteines and one histidine. They act as redox switches with an active state linked to the oxidation of their cluster. In the present study, we show that reduced glutathione but also free thiol-containing molecules such as ß-mercaptoethanol can induce a loss of the mNT cluster under aerobic conditions, while CISD2 cluster appears more resistant. This disassembly occurs through a radical-based mechanism as previously observed with the bacterial SoxR. Interestingly, adding cysteine prevents glutathione-induced cluster loss. At low pH, glutathione can bind mNT in the vicinity of the cluster. These results suggest a potential new regulation mechanism of mNT activity by glutathione, an essential actor of the intracellular redox state.


Asunto(s)
Proteínas Mitocondriales , Humanos , Cisteína/metabolismo , Glutatión/metabolismo , Homeostasis , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Oxidación-Reducción , Compuestos de Sulfhidrilo
18.
Carcinogenesis ; 34(4): 760-9, 2013 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-23239746

RESUMEN

Peroxiredoxin (Prx) 1 is a member of the thiol-specific peroxidases family and plays diverse roles such as H2O2 scavenger, redox signal transducer and molecular chaperone. Prx1 has been reported to be involved in protecting cancer cells against various therapeutic challenges. We investigated how modulations of intracellular redox system affect cancer cell sensitivity to reactive oxygen species (ROS)-generating drugs. We observed that stable and transient Prx1 knockdown significantly enhanced HeLa cell sensitivity to ß-lapachone (ß-lap), a potential anticancer agent. Prx1 knockdown markedly potentiated 2 µM ß-lap-induced cytotoxicity through ROS accumulation. This effect was largely NAD(P)H:quinone oxidoreductase 1 dependent and associated with a decrease in poly(ADP-ribose) polymerase 1 protein levels, phosphorylation of JNK, p38 and Erk proteins in mitogen-activated protein kinase (MAPK) pathways and a decrease in thioredoxin 1 (Trx1) protein levels. Trx1 serves as an electron donor for Prx1 and is overexpressed in Prx1 knockdown cells. Based on the fact that Prx1 is a major ROS scavenger and a partner of at least ASK1 and JNK, two key components of MAPK pathways, we propose that Prx1 knockdown-induced sensitization to ß-lap is achieved through combined action of accumulation of ROS and enhancement of MAPK pathway activation, leading to cell apoptosis. These data support the view that modulation of intracellular redox state could be an alternative approach to enhance cancer cell sensitivity to ROS-generating drugs or to overcome some types of drug resistance.


Asunto(s)
Naftoquinonas/farmacología , Neoplasias/tratamiento farmacológico , Peroxirredoxinas/genética , Especies Reactivas de Oxígeno/metabolismo , Apoptosis/fisiología , Línea Celular Tumoral , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Células HeLa , Humanos , Peróxido de Hidrógeno/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Sistema de Señalización de MAP Quinasas , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Neoplasias/metabolismo , Peroxirredoxinas/metabolismo , Fosforilación , Poli(ADP-Ribosa) Polimerasa-1 , Poli(ADP-Ribosa) Polimerasas/biosíntesis , Poli(ADP-Ribosa) Polimerasas/metabolismo , Interferencia de ARN , ARN Interferente Pequeño , Inhibidores de la Transcriptasa Inversa/farmacología , Tiorredoxinas/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo
19.
J Biol Chem ; 287(7): 4552-61, 2012 Feb 10.
Artículo en Inglés | MEDLINE | ID: mdl-22170048

RESUMEN

Glutathione (GSH) has several important functions in eukaryotic cells, and its intracellular concentration is tightly controlled. Combining mathematical models and (35)S labeling, we analyzed Saccharomyces cerevisiae sulfur metabolism. This led us to the observation that GSH recycling is markedly faster than previously estimated. We set up additional in vivo assays and concluded that under standard conditions, GSH half-life is around 90 min. Sulfur starvation and growth with GSH as the sole sulfur source strongly increase GSH degradation, whereas cadmium (Cd(2+)) treatment inhibits GSH degradation. Whatever the condition tested, GSH is degraded by the cytosolic Dug complex (composed of the three subunits Dug1, Dug2, and Dug3) but not by the γ-glutamyl-transpeptidase, raising the question of the role of this enzyme. In vivo, both DUG2/3 mRNA levels and Dug activity are quickly induced by sulfur deprivation in a Met4-dependent manner. This suggests that Dug activity is mainly regulated at the transcriptional level. Finally, analysis of dug2Δ and dug3Δ mutant cells shows that GSH degradation activity strongly impacts on GSH intracellular concentration and that GSH intracellular concentration does not affect GSH synthesis rate. Altogether, our data led us to reconsider important aspects of GSH metabolism, challenging notions on GSH synthesis and GSH degradation that were considered as established.


Asunto(s)
Ligasas de Carbono-Nitrógeno/metabolismo , Dipeptidasas/metabolismo , Glutatión/metabolismo , Homeostasis/fisiología , Complejos Multienzimáticos/metabolismo , Péptido Hidrolasas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Cadmio/farmacología , Ligasas de Carbono-Nitrógeno/genética , Dipeptidasas/genética , Eliminación de Gen , Glutatión/genética , Semivida , Homeostasis/efectos de los fármacos , Complejos Multienzimáticos/genética , Péptido Hidrolasas/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Azufre/metabolismo
20.
Blood ; 117(6): 1917-27, 2011 Feb 10.
Artículo en Inglés | MEDLINE | ID: mdl-21139082

RESUMEN

Chronic lymphocytic leukemia (CLL) is characterized by a clonal accumulation of mature neoplastic B cells that are resistant to apoptosis. Aiolos, a member of the Ikaros family of zinc-finger transcription factors, plays an important role in the control of mature B lymphocyte differentiation and maturation. In this study, we showed that Aiolos expression is up-regulated in B-CLL cells. This overexpression does not implicate isoform imbalance or disturb Aiolos subcellular localization. The chromatin status at the Aiolos promoter in CLL is defined by the demethylation of DNA and an enrichment of euchromatin associated histone markers, such as the dimethylation of the lysine 4 on histone H3. These epigenetic modifications should allow its upstream effectors, such as nuclear factor-κB, constitutively activated in CLL, to gain access to promoter, resulting up-regulation of Aiolos. To determine the consequences of Aiolos deregulation in CLL, we analyzed the effects of Aiolos overexpression or down-regulation on apoptosis. Aiolos is involved in cell survival by regulating the expression of some Bcl-2 family members. Our results strongly suggest that Aiolos deregulation by epigenetic modifications may be a hallmark of CLL.


Asunto(s)
Epigénesis Genética , Factor de Transcripción Ikaros/genética , Leucemia Linfocítica Crónica de Células B/genética , Adulto , Anciano , Anciano de 80 o más Años , Apoptosis/genética , Apoptosis/fisiología , Linfocitos B/metabolismo , Linfocitos B/patología , Secuencia de Bases , Supervivencia Celular/genética , Supervivencia Celular/fisiología , Cromatina/genética , Cromatina/metabolismo , Islas de CpG , Metilación de ADN , Cartilla de ADN/genética , Femenino , Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Factor de Transcripción Ikaros/antagonistas & inhibidores , Factor de Transcripción Ikaros/metabolismo , Técnicas In Vitro , Leucemia Linfocítica Crónica de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/patología , Masculino , Persona de Mediana Edad , Modelos Biológicos , FN-kappa B/antagonistas & inhibidores , FN-kappa B/metabolismo , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Neoplásico/genética , ARN Neoplásico/metabolismo , ARN Interferente Pequeño/genética , Fracciones Subcelulares/metabolismo
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