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1.
Front Plant Sci ; 15: 1445488, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-39381512

RESUMEN

The basic helix-loop-helix (bHLH) transcription factors play crucial roles in various processes, such as plant development, secondary metabolism, and response to biotic/abiotic stresses. Sophora flavescens is a widely used traditional herbal medicine in clinical practice, known for its abundant flavonoids as the main active compounds. However, there has been no comprehensive analysis of S. flavescens bHLH (SfbHLH) gene family reported currently. In this study, we identified 167 SfbHLH genes and classified them into 23 subfamilies based on comparative genomics and phylogenetic analysis. Furthermore, widespread duplications significantly contributed to the expansion of SfbHLH family. Notably, SfbHLH042 was found to occupy a central position in the bHLH protein-protein interaction network. Transcriptome analysis of four tissues (leaf, stem, root and flower) revealed that most SfbHLH genes exhibited high expression levels exclusively in specific tissues of S. flavescens. The integrated analysis of transcriptomics and metabolomics during pod development stages revealed that SfbHLH042 may play a central role in connecting SfbHLH genes, flavonoids, and key enzymes involved in the biosynthesis pathway. Moreover, we also checked the expression of 8 SfbHLH genes using RT-qPCR analysis to realize the expression profiles of these genes among various tissues at different cultivated periods and root development. Our study would aid to understand the phylogeny and expression profile of SfbHLH family genes, and provide a promising candidate gene, SfbHLH042, for regulating the biosynthesis of flavonoids in S. flavescens.

2.
Br J Pharmacol ; 2024 Oct 06.
Artículo en Inglés | MEDLINE | ID: mdl-39370580

RESUMEN

BACKGROUND AND PURPOSE: KCNT1 encodes a sodium-activated potassium channel (Slack channel), and its mutation can cause several forms of epilepsy. Traditional antiepileptic medications have limited efficacy in treating patients with KCNT1 mutations. Here, we describe one heterozygous KCNT1 mutation, M267T, in a patient with EIMFS. The pathological channel properties of this mutation and its effect on neuronal excitability were investigated. Additionally, this study aimed to develop a medication for effective prevention of KCNT1 mutation-induced seizures. EXPERIMENTAL APPROACH: Wild-type or mutant KCNT1 plasmids were expressed heterologously in Xenopus laevis oocytes, and channel property assessment and drug screening were performed based on two-electrode voltage-clamp recordings. The single-channel properties were investigated using the excised inside-out patches from HEK293T cells. Through in utero electroporation, WT and M267T Slack channels were expressed in the hippocampal CA1 pyramidal neurons in male mice, followed by the examination of the electrical properties using the whole-cell current-clamp technique. The kainic acid-induced epilepsy model in male mice was used to evalute the antiseizure effects of carvedilol. KEY RESULTS: The KCNT1 M267T mutation enhanced Slack channel function by increasing single-channel open probability. Through screening 16 FDA-approved ion channel blockers, we found that carvedilol effectively reversed the mutation-induced gain-of-function channel properties. Notably, the KCNT1 M267T mutation in the mouse hippocampal CA1 pyramidal neurons affected afterhyperpolarization properties and induced neuronal hyperexcitability, which was inhibited by carvedilol. Additionally, carvedilol exhibited antiseizure effects in the kainic acid-induced epilepsy model. CONCLUSION AND IMPLICATION: Our findings suggest carvedilol as a new potential candidate for treatment of epilepsies.

3.
Genomics ; 116(5): 110926, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-39178997

RESUMEN

During sunflower growth, cold waves often occur and impede plant growth. Therefore, it is crucial to study the underlying mechanism of cold resistance in sunflowers. In this study, physiological analysis revealed that as cold stress increased, the levels of ROS, malondialdehyde, ascorbic acid, and dehydroascorbic acid and the activities of antioxidant enzymes increased. Transcriptomics further identified 10,903 DEGs between any two treatments. Clustering analysis demonstrated that the expression of MYB44a, MYB44b, MYB12, bZIP2 and bZIP4 continuously upregulated under cold stress. Cold stress can induce ROS accumulation, which interacts with hormone signals to activate cold-responsive transcription factors regulating target genes involved in antioxidant defense, secondary metabolite biosynthesis, starch and sucrose metabolism enhancement for improved cold resistance in sunflowers. Additionally, the response of sunflowers to cold stress may be independent of the CBF pathway. These findings enhance our understanding of cold stress resistance in sunflowers and provide a foundation for genetic breeding.


Asunto(s)
Respuesta al Choque por Frío , Regulación de la Expresión Génica de las Plantas , Helianthus , Plantones , Transcriptoma , Plantones/metabolismo , Plantones/genética , Helianthus/genética , Helianthus/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Especies Reactivas de Oxígeno/metabolismo , Frío
4.
BMC Plant Biol ; 24(1): 792, 2024 Aug 22.
Artículo en Inglés | MEDLINE | ID: mdl-39169292

RESUMEN

BACKGROUND: With the progress of industrialization and urbanization, cadmium (Cd) pollution in farmland is increasingly severe, greatly affecting human health. Sunflowers possess high resistance to Cd stress and great potential for phytoremediation of Cd-contaminated soil. Previous studies have shown that humic acid (HA) effectively mitigates plant damage induced by Cd; however, its alleviating effects on sunflower plants under Cd stress remain largely unknown. RESULTS: We employed four different concentrations of HA (50, 100, 200, and 300 mg L-1) via foliar application to examine their ability to alleviate Cd stress on sunflower plants' growth, chlorophyll synthesis, and biochemical defense system. The results revealed that Cd stress not only reduced plant height, stem diameter, fresh and dry weight, and chlorophyll content in sunflower plants but also altered their chlorophyll fluorescence characteristics compared to the control group. After Cd stress, the photosynthetic structure was damaged and the number of PSII reactive centers per unit changed. Application of 200 mg L-1 HA promotes sunflower growth and increases chlorophyll content. HA significantly enhances antioxidant enzyme activities (SOD, POD, CAT, and APX) and reduces ROS content (O2 -, H2O2 and -OH). Totally, Application of 200 mg L-1 HA had the best effect than other concentrations to alleviate the Cd-induced stress in sunflower plants. CONCLUSIONS: The foliar application of certain HA concentration exhibited the most effective alleviation of Cd-induced stress on sunflower plants. It can enhance the light energy utilization and antioxidant enzyme activities, while reduce ROS contents in sunflower plants. These findings provide a theoretical basis for using HA to mitigate Cd stress in sunflowers.


Asunto(s)
Cadmio , Clorofila , Helianthus , Sustancias Húmicas , Clorofila/metabolismo , Helianthus/efectos de los fármacos , Helianthus/metabolismo , Helianthus/crecimiento & desarrollo , Hojas de la Planta/metabolismo , Hojas de la Planta/efectos de los fármacos , Estrés Fisiológico , Biodegradación Ambiental , Contaminantes del Suelo , Fotosíntesis/efectos de los fármacos , Antioxidantes/metabolismo , Especies Reactivas de Oxígeno/metabolismo
5.
Adv Sci (Weinh) ; 11(29): e2400560, 2024 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-38874331

RESUMEN

Intrinsic plasticity, a fundamental process enabling neurons to modify their intrinsic properties, plays a crucial role in shaping neuronal input-output function and is implicated in various neurological and psychiatric disorders. Despite its importance, the underlying molecular mechanisms of intrinsic plasticity remain poorly understood. In this study, a new ubiquitin ligase adaptor, protein tyrosine phosphatase receptor type N (PTPRN), is identified as a regulator of intrinsic neuronal excitability in the context of temporal lobe epilepsy. PTPRN recruits the NEDD4 Like E3 Ubiquitin Protein Ligase (NEDD4L) to NaV1.2 sodium channels, facilitating NEDD4L-mediated ubiquitination, and endocytosis of NaV1.2. Knockout of PTPRN in hippocampal granule cells leads to augmented NaV1.2-mediated sodium currents and higher intrinsic excitability, resulting in increased seizure susceptibility in transgenic mice. Conversely, adeno-associated virus-mediated delivery of PTPRN in the dentate gyrus region decreases intrinsic excitability and reduces seizure susceptibility. Moreover, the present findings indicate that PTPRN exerts a selective modulation effect on voltage-gated sodium channels. Collectively, PTPRN plays a significant role in regulating intrinsic excitability and seizure susceptibility, suggesting a potential strategy for precise modulation of NaV1.2 channels' function.


Asunto(s)
Endocitosis , Convulsiones , Animales , Ratones , Convulsiones/metabolismo , Convulsiones/genética , Endocitosis/fisiología , Endocitosis/genética , Ratones Transgénicos , Modelos Animales de Enfermedad , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Masculino , Ratones Noqueados
6.
Stem Cells Dev ; 33(15-16): 432-437, 2024 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-38801165

RESUMEN

The therapeutic potential of autologous stem cell transplantation for heart repair diminishes in the elderly due to stem cell aging. Rejuvenating aged stem cells to enhance their protective effects on injured cardiomyocytes is crucial for aging patients with heart failure. In this study, we aimed to investigate whether neuron-derived neurotrophic factor (NDNF) over-expression improves the protective effect of aged stem cells for injured cardiomyocytes and explore the underlying mechanism. Human bone marrow was collected from both young and old patients, and bone marrow mesenchymal stem cells (BMSCs) were cultured. Lentivirus expression vectors carrying NDNF genes were used to transfect aged BMSCs. Fatal hypoxia-induced injury in H9C2 cells served as an in vitro ischemia model. The conditioned medium from different BMSC groups was applied to assess the beneficial effects on hypoxia-induced damage in myocardial H9C2 cells. Results revealed that the conditioned medium of NDNF over-expressed old BMSCs increased H9C2 cell viability and reduced oxidative stress and apoptosis levels under fatal hypoxia. NDNF over-expressed old BMSCs exhibited an antiapoptotic role by upregulating the antiapoptotic gene Bcl-2 and downregulating the proapoptotic genes Bax. Additionally, the protective effects were mediated through the elevation of phosphorylated AKT. Our data support the promise of NDNF as a potential target to enhance the protective effects of autologous aged BMSCs on ischemic cardiomyocytes and then improve the curative effects of stem cell for ischemic heart injury in aged patients.


Asunto(s)
Apoptosis , Células Madre Mesenquimatosas , Miocitos Cardíacos , Estrés Oxidativo , Adulto , Anciano , Animales , Humanos , Masculino , Ratas , Apoptosis/genética , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/citología , Hipoxia de la Célula , Línea Celular , Supervivencia Celular , Células Cultivadas , Senescencia Celular/genética , Medios de Cultivo Condicionados/farmacología , Citoprotección/genética , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Miocitos Cardíacos/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética
7.
Molecules ; 29(7)2024 Mar 26.
Artículo en Inglés | MEDLINE | ID: mdl-38611748

RESUMEN

Stem cell-derived exosomes (SC-Exos) are used as a source of regenerative medicine, but certain limitations hinder their uses. The effect of hydrolyzed collagen oligopeptides (HCOPs), a functional ingredient of SC-Exos is not widely known to the general public. We herein evaluated the combined anti-aging effects of HCOPs and exosomes derived from human umbilical cord mesenchymal stem cells (HucMSC-Exos) using a senescence model established on human skin fibroblasts (HSFs). This study discovered that cells treated with HucMSC-Exos + HCOPs enhanced their proliferative and migratory capabilities; reduced both reactive oxygen species production and senescence-associated ß-galactosidase activity; augmented type I and type III collagen expression; attenuated the expression of matrix-degrading metalloproteinases (MMP-1, MMP-3, and MMP-9), interleukin 1 beta (IL-1ß), and tumor necrosis factor-alpha (TNF-α); and decreased the expression of p16, p21, and p53 as compared with the cells treated with HucMSC-Exos or HCOPs alone. These results suggest a possible strategy for enhancing the skin anti-aging ability of HucMSC-Exos with HCOPs.


Asunto(s)
Exosomas , Células Madre Mesenquimatosas , Humanos , Fibroblastos , Envejecimiento , Colágeno Tipo III , Cordón Umbilical
8.
Gene ; 910: 148336, 2024 Jun 05.
Artículo en Inglés | MEDLINE | ID: mdl-38447680

RESUMEN

DNA binding with one finger (Dof), plant-specific zinc finger transcription factors, can participate in various physiological and biochemical processes during the life of plants. As one of the most important oil crops in the world, sunflower (Helianthus annuus L.) has significant economic and ornamental value. However, a systematic analysis of H. annuus Dof (HaDof) members and their functions has not been extensively conducted. In this study, we identified 50 HaDof genes that are unevenly distributed on 17 chromosomes of sunflower. We present a comprehensive overview of the HaDof genes, including their chromosome locations, phylogenetic analysis, and expression profile characterization. Phylogenetic analysis classified the 366 Dof members identified from 11 species into four groups (further subdivided into nine subfamilies). Segmental duplications are predominantly contributed to the expansion of sunflower Dof genes, and all segmental duplicate gene pairs are under purifying selection due to strong evolutionary constraints. Furthermore, we observed differential expression patterns for HaDof genes in normal tissues as well as under hormone treatment or abiotic stress conditions by analyzing RNA-seq data from previous studies and RT-qPCR data in our current study. The expression of HaDof04 and HaDof43 were not detected in any samples, which implied that they may be gradually undergoing pseudogenization process. Some HaDof genes, such as HaDof25 and HaDof30, showed responsiveness to exogenous plant hormones, such as kinetin, brassinosteroid, auxin or strigolactone, while others like HaDof15 and HaDof35 may participate in abiotic stress resistance of sunflower seedling. Our study represents the initial step towards understanding the phylogeny and expression characterization of sunflower Dof family genes, which may provide valuable reference information for functional studies on hormone response, abiotic stress resistance, and molecular breeding in sunflower and other species.


Asunto(s)
Helianthus , Helianthus/genética , Helianthus/metabolismo , Filogenia , Familia de Multigenes , Estrés Fisiológico/genética , Genoma de Planta , Hormonas , Regulación de la Expresión Génica de las Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
9.
Adv Healthc Mater ; 13(10): e2304207, 2024 04.
Artículo en Inglés | MEDLINE | ID: mdl-38175149

RESUMEN

Myocardial infarction (MI) results in cardiomyocyte necrosis and conductive system damage, leading to sudden cardiac death and heart failure. Studies have shown that conductive biomaterials can restore cardiac conduction, but cannot facilitate tissue regeneration. This study aims to add regenerative capabilities to the conductive biomaterial by incorporating human endometrial mesenchymal stem cell (hEMSC)-derived exosomes (hEMSC-Exo) into poly-pyrrole-chitosan (PPY-CHI), to yield an injectable hydrogel that can effectively treat MI. In vitro, PPY-CHI/hEMSC-Exo, compared to untreated controls, PPY-CHI, or hEMSC-Exo alone, alleviates H2O2-induced apoptosis and promotes tubule formation, while in vivo, PPY-CHI/hEMSC-Exo improves post-MI cardiac functioning, along with counteracting against ventricular remodeling and fibrosis. All these activities are facilitated via increased epidermal growth factor (EGF)/phosphoinositide 3-kinase (PI3K)/AKT signaling. Furthermore, the conductive properties of PPY-CHI/hEMSC-Exo are able to resynchronize cardiac electrical transmission to alleviate arrythmia. Overall, PPY-CHI/hEMSC-Exo synergistically combines the cardiac regenerative capabilities of hEMSC-Exo with the conductive properties of PPY-CHI to improve cardiac functioning, via promoting angiogenesis and inhibiting apoptosis, as well as resynchronizing electrical conduction, to ultimately enable more effective MI treatment. Therefore, incorporating exosomes into a conductive hydrogel provides dual benefits in terms of maintaining conductivity, along with facilitating long-term exosome release and sustained application of their beneficial effects.


Asunto(s)
Quitosano , Exosomas , Células Madre Mesenquimatosas , Infarto del Miocardio , Humanos , Polímeros/metabolismo , Hidrogeles/farmacología , Hidrogeles/metabolismo , Pirroles , Exosomas/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Preparaciones de Acción Retardada/farmacología , Peróxido de Hidrógeno/metabolismo , Infarto del Miocardio/terapia , Materiales Biocompatibles/farmacología , Materiales Biocompatibles/metabolismo , Miocitos Cardíacos/metabolismo
10.
Elife ; 122024 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-38289338

RESUMEN

Quinidine has been used as an anticonvulsant to treat patients with KCNT1-related epilepsy by targeting gain-of-function KCNT1 pathogenic mutant variants. However, the detailed mechanism underlying quinidine's blockade against KCNT1 (Slack) remains elusive. Here, we report a functional and physical coupling of the voltage-gated sodium channel NaV1.6 and Slack. NaV1.6 binds to and highly sensitizes Slack to quinidine blockade. Homozygous knockout of NaV1.6 reduces the sensitivity of native sodium-activated potassium currents to quinidine blockade. NaV1.6-mediated sensitization requires the involvement of NaV1.6's N- and C-termini binding to Slack's C-terminus and is enhanced by transient sodium influx through NaV1.6. Moreover, disrupting the Slack-NaV1.6 interaction by viral expression of Slack's C-terminus can protect against SlackG269S-induced seizures in mice. These insights about a Slack-NaV1.6 complex challenge the traditional view of 'Slack as an isolated target' for anti-epileptic drug discovery efforts and can guide the development of innovative therapeutic strategies for KCNT1-related epilepsy.


Asunto(s)
Epilepsia , Canal de Sodio Activado por Voltaje NAV1.6 , Quinidina , Animales , Humanos , Ratones , Anticonvulsivantes/farmacología , Anticonvulsivantes/uso terapéutico , Homocigoto , Canal de Sodio Activado por Voltaje NAV1.6/genética , Proteínas del Tejido Nervioso/genética , Quinidina/farmacología , Sodio
11.
Stem Cells ; 42(5): 430-444, 2024 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-38253331

RESUMEN

It has been documented that the uterus plays a key cardio-protective role in pre-menopausal women, which is supported by uterine cell therapy, to preserve cardiac functioning post-myocardial infarction, being effective among females. However, whether such therapies would also be beneficial among males is still largely unknown. In this study, we aimed to fill in this gap in knowledge by examining the effects of transplanted uterine cells on infarcted male hearts. We identified, based on major histocompatibility complex class I (MHC-I) expression levels, 3 uterine reparative cell populations: MHC-I(neg), MHC-I(mix), and MHC-I(pos). In vitro, MHC-I(neg) cells showed higher levels of pro-angiogenic, pro-survival, and anti-inflammatory factors, compared to MHC-I(mix) and MHC-I(pos). Furthermore, when cocultured with allogeneic mixed leukocytes, MHC-I(neg) had lower cytotoxicity and leukocyte proliferation. In particular, CD8+ cytotoxic T cells significantly decreased, while CD4+CD25+ Tregs and CD4-CD8- double-negative T cells significantly increased when cocultured with MHC-I(neg), compared to MHC-I(mix) and MHC-I(pos) cocultures. In vivo, MHC-I(neg) as well as MHC-I(mix) were found under both syngeneic and allogeneic transplantation in infarcted male hearts, to significantly improve cardiac function and reduce the scar size, via promoting angiogenesis in the infarcted area. All of these findings thus support the view that males could also benefit from the cardio-protective effects observed among females, via cell therapy approaches involving the transplantation of immuno-privileged uterine reparative cells in infarcted hearts.


Asunto(s)
Infarto del Miocardio , Útero , Infarto del Miocardio/terapia , Infarto del Miocardio/patología , Masculino , Femenino , Animales , Útero/irrigación sanguínea , Ratones , Ratones Endogámicos C57BL , Antígenos de Histocompatibilidad Clase I/metabolismo
13.
iScience ; 26(11): 108112, 2023 Nov 17.
Artículo en Inglés | MEDLINE | ID: mdl-37860690

RESUMEN

Drought can adversely influence the crop growth and production. Accordingly, sunflowers have strong adaptability to drought; hence, we conducted analyses for sunflower seedlings with drought stress and rehydration drought acclimation through physiological measurements and transcriptomics. It showed that drought can cause the accumulation of ROS and enhance the activity of antioxidant enzymes and the content of osmolytes. After rehydration, the contents of ROS and MDA were significantly reduced concomitant with increased antioxidant activity and osmotic adjustment. Totally, 2,589 DEGs were identified among treatments. Functional enrichment analysis showed that DEGs were mainly involved in plant hormone signal transduction, MAPK signaling, and biosynthesis of secondary metabolites. Comparison between differentially spliced genes and DEGs indicated that bHLH025, NAC53, and SINAT3 may be pivotal genes involved in sunflower drought resistance. Our results not only highlight the underlying mechanism of drought stress and rehydration in sunflower but also provide a theoretical basis for crop genetic breeding.

14.
Plant Physiol ; 194(1): 511-529, 2023 Dec 30.
Artículo en Inglés | MEDLINE | ID: mdl-37757893

RESUMEN

Soil salinity is a major environmental factor constraining growth and productivity of highbush blueberry (Vaccinium corymbosum). Leaf Na+ content is associated with variation in salt tolerance among blueberry cultivars; however, the determinants and mechanisms conferring leaf Na+ exclusion are unknown. Here, we observed that the blueberry cultivar 'Duke' was more tolerant than 'Sweetheart' and accumulated less Na+ in leaves under salt stress conditions. Through transcript profiling, we identified a member of the high-affinity K+ transporter (HKT) family in blueberry, VcHKT1;1, as a candidate gene involved in leaf Na+ exclusion and salt tolerance. VcHKT1;1 encodes a Na+-preferential transporter localized to the plasma membrane and is preferentially expressed in the root stele. Heterologous expression of VcHKT1;1 in Arabidopsis (Arabidopsis thaliana) rescued the salt hypersensitivity phenotype of the athkt1 mutant. Decreased VcHKT1;1 transcript levels in blueberry plants expressing antisense-VcHKT1;1 led to increased Na+ concentrations in xylem sap and higher leaf Na+ contents compared with wild-type plants, indicating that VcHKT1;1 promotes leaf Na+ exclusion by retrieving Na+ from xylem sap. A naturally occurring 8-bp insertion in the promoter increased the transcription level of VcHKT1;1, thus promoting leaf Na+ exclusion and blueberry salt tolerance. Collectively, we provide evidence that VcHKT1;1 promotes leaf Na+ exclusion and propose natural variation in VcHKT1;1 will be valuable for breeding Na+-tolerant blueberry cultivars in the future.


Asunto(s)
Arabidopsis , Arándanos Azules (Planta) , Tolerancia a la Sal/genética , Arándanos Azules (Planta)/genética , Arándanos Azules (Planta)/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fitomejoramiento , Proteínas de Transporte de Membrana/metabolismo , Arabidopsis/metabolismo
15.
Plants (Basel) ; 12(16)2023 Aug 09.
Artículo en Inglés | MEDLINE | ID: mdl-37631118

RESUMEN

The Agrobacterium tumefaciens-mediated transformation for blueberries remains less efficient than is desirable. A new leaf callus regeneration and genetic transformation system was investigated in blueberries in this study. The leaf explants of cv. 'Legacy' and 'Northland' were used to establish the stable callus induction system when placed on the woody plant medium (WPM) supplemented with 1.0 mg·L-1 2, 4-D, 0.4 mg·L-1 6-BA for 30 d; then, the callus was sub-cultured in the proliferation medium supplemented with 1.5 mg·L-1 2, 4-D, 0.4 mg·L-1 6-BA in the darkness at 25 °C every 30 days. The co-cultivation of callus with A. tumefaciens was operated on WPM plus 100 µM acetosyringone for 4 days; then, the transferred callus was grown in WPM supplemented with 1.5 mg·L-1 2,4-D, 0.4 mg·L-1 6-BA, 50 mg·L-1 hygromycin, and 200 mg·L-1 cefotaxime. The VcCHS transgenic blueberry callus with both GFP signal and Hyg resistance was obtained from the transformed callus of cv. 'Northland'. The rate of GFP signal detected in the transformed callus was as high as 49.02%, which was consistent with the PCR assay. Collectively, this study provides a highly efficient genetic transformation system in blueberry callus and a powerful approach for the molecular breeding of blueberries.

16.
Drug Test Anal ; 15(11-12): 1410-1416, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37644676

RESUMEN

The issue of misjudgment in recombinant erythropoietin (rEPO) detection caused by the variant c.577del in human EPO gene has been found in recent years. Though the method of analyzing de-N-glycosylated erythropoietins (EPO) in blood samples was developed for identifying both EPO_p.Arg193AspfsTer28 (VAR-EPO) and rEPO, it cannot be applied without the evaluation of excreted samples. For this purpose, five heterozygous carriers of the variant c.577del were recruited in an administration study of rEPO. Urine and blood samples were collected at different times before and after subcutaneous injection with a single-dose of 50 IU/kg. The urine samples were analyzed for intact EPO, while the serum samples were analyzed for both intact and de-N-glycosylated EPO. A typical mixed band was detected in all blank and wash-out urine samples, which all displayed a similar result with rEPO abuse. For the analysis of intact EPO in serum samples, a typical mixed band was detected in the wash-out samples from day 1 to day 3, which could be identified as rEPO directly, while double-band was observed in other samples with inconclusive results. The result of de-N-glycosylated EPO in all serum samples showed two separated bands, and the ratioL/U decreased along with wash-out periods. Also, compared with the intact EPO analysis, a longer detection window without false positive results was obtained when analyzing de-N-glycosylated EPO. Analysis of de-N-glycosylated EPO is not only able to recognize the variant carriers directly but also able to detect rEPO abuse in the blood samples from the variant carriers with higher efficiency than the analysis of intact EPO.


Asunto(s)
Eritropoyetina , Humanos , Proteínas Recombinantes
18.
Int J Rheum Dis ; 26(4): 638-647, 2023 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-36789537

RESUMEN

BACKGROUND: Glucocorticoid-induced osteoporosis is the most common secondary cause of osteoporosis, which increases the risk of fracture. Long non-coding RNA telomerase RNA elements (TERC) has been proven to be closely related to osteoporosis. However, the role of TERC in glucocorticoid-induced osteoporosis and its underlying molecular mechanism remains unclear. METHODS: The in vitro model of osteoporosis was established after bone marrow mesenchymal stem cells (BMSCs) were exposed to dexamethasone (DEX). The cell viability, alkaline phosphatase (ALP) activity and mineralized nodules of BMSCs were evaluated. The messenger RNA and protein levels were detected by quantitative real-time polymerase chain reaction and Western blot. The interaction between TERC, enhancer of zeste homolog 2 (EZH2) and dickkopf-1 (DKK1) was confirmed by chromatin immunoprecipitation and RNA immunoprecipitation assays. RESULTS: Bone marrow mesenchymal stem cells were isolated, identified and induced osteogenic differentiation. The findings showed that the levels of osteogenic marker genes, including ALP, Runt-related transcription factor 2 (RUNX2) and osteocalcin (OCN) in BMSCs were increased dependent on the osteogenic induction time. Similarly, TERC was significantly increased, but DKK1 was significantly decreased during BMSC osteogenic differentiation. Functional research showed that TERC overexpression promoted cell viability, ALP activity and mineralized nodules of BMSCs and increased the levels of osteogenic differentiation-related genes (ALP, RUNX2 and OCN), and TERC overexpression increased EZH2 protein level. Moreover, the decrease of cell viability, ALP activity and mineralized nodules induced by DEX was reversed by TERC overexpression. Furthermore, TERC inhibited DKK1 expression by promoting the histone modification of DKK1, and TERC overexpression alleviated DEX suppressed osteogenic differentiation of BMSCs by interaction with EZH2 to regulate DKK1. CONCLUSION: Our findings illustrated that TERC overexpression alleviated DEX-induced osteoporosis by recruiting EZH2 to regulate DKK1. Our research provided a novel direction for the treatment of glucocorticoid-induced osteoporosis.


Asunto(s)
MicroARNs , Osteoporosis , ARN Largo no Codificante , Humanos , Glucocorticoides/efectos adversos , MicroARNs/genética , ARN Largo no Codificante/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Osteogénesis , Proteína Potenciadora del Homólogo Zeste 2/genética , Osteoporosis/inducido químicamente , Osteoporosis/genética , Diferenciación Celular , Células Cultivadas , Péptidos y Proteínas de Señalización Intercelular/genética
19.
Mol Cell Biochem ; 478(6): 1191-1204, 2023 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-36266491

RESUMEN

Human endometrial mesenchymal stem cells (hEMSCs) have been shown to promote neo-vascularization; however, its angiogenic function lessens with age. To determine the optimal conditions for maximizing hEMSC angiogenic capacity, we examined the effects of serial passaging on hEMSC activity. hEMSCs were cultured from passages (P) 3, 6, 9, and 12, and analyzed for proliferation, migration, differentiation and senescence, as well as their capacity to induce angiogenesis. The results showed that hEMSC proliferation and migration significantly decreased after P12. Furthermore, hEMSC differentiation into adipogenic and osteogenic lineages, as well as their proangiogenic capacity, gradually decreased from P9-12, while senescence only occurred after P12. Evaluation of angiogenic-related protein levels showed that both transforming growth factor ß2 and Tie-2 was significantly reduced in hEMSCs at P12, compared to P3, possibly serving as the basis behind their lowered angiogenic capacity. Furthermore, in vivo angiogenesis evaluation with Matrigel plug assay showed that the optimal hEMSC to HUVEC ratio, for maximizing vessel formation, was 1:4. This study showed that hEMSC passaging was associated with lowered cellular functioning, bringing them closer to a senescent phenotype, especially after P12, thereby defining the optimal time period for cultivating fully functional hEMSCs for therapeutic applications.


Asunto(s)
Fenómenos Fisiológicos Cardiovasculares , Células Madre Mesenquimatosas , Humanos , Diferenciación Celular , Neovascularización Fisiológica , Osteogénesis , Células Madre Mesenquimatosas/metabolismo , Células Cultivadas , Proliferación Celular
20.
Int J Mol Sci ; 23(24)2022 Dec 16.
Artículo en Inglés | MEDLINE | ID: mdl-36555693

RESUMEN

Saline-alkali stress is a widespread adversity that severely affects plant growth and productivity. Saline-alkaline soils are characterized by high salt content and high pH values, which simultaneously cause combined damage from osmotic stress, ionic toxicity, high pH and HCO3-/CO32- stress. In recent years, many determinants of salt tolerance have been identified and their regulatory mechanisms are fairly well understood. However, the mechanism by which plants respond to comprehensive saline-alkali stress remains largely unknown. This review summarizes recent advances in the physiological, biochemical and molecular mechanisms of plants tolerance to salinity or salt- alkali stress. Focused on the progress made in elucidating the regulation mechanisms adopted by plants in response to saline-alkali stress and present some new views on the understanding of plants in the face of comprehensive stress. Plants generally promote saline-alkali tolerance by maintaining pH and Na+ homeostasis, while the plants responding to HCO3-/CO32- stress are not exactly the same as high pH stress. We proposed that pH-tolerant or sensitive plants have evolved distinct mechanisms to adapt to saline-alkaline stress. Finally, we highlight the areas that require further research to reveal the new components of saline-alkali tolerance in plants and present the current and potential application of key determinants in breed improvement and molecular breeding.


Asunto(s)
Álcalis , Tolerancia a la Sal , Fitomejoramiento , Plantas , Estrés Salino , Solución Salina
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