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1.
J Am Chem Soc ; 145(3): 1593-1606, 2023 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-36626587

RESUMO

Gene regulation via chemically induced dimerization (CID) is useful for biomedical research. However, the number, type, versatility, and in vivo applications of CID tools remain limited. Here, we demonstrate the development of proteolysis-targeting chimera-based scalable CID (PROTAC-CID) platforms by systematically engineering the available PROTAC systems for inducible gene regulation and gene editing. Further, we show orthogonal PROTAC-CIDs that can fine-tune gene expression at gradient levels or multiplex biological signals with different logic gating operations. Coupling the PROTAC-CID platform with genetic circuits, we achieve digitally inducible expression of DNA recombinases, base- and prime-editors for transient genome manipulation. Finally, we package a compact PROTAC-CID system into adeno-associated viral vectors for inducible and reversible gene activation in vivo. This work provides a versatile molecular toolbox that expands the scope of chemically inducible gene regulation in human cells and mice.


Assuntos
DNA , Recombinases , Humanos , Camundongos , Animais , Dimerização , DNA/metabolismo , Recombinases/metabolismo , Edição de Genes , Genoma , Sistemas CRISPR-Cas , Mamíferos/genética , Mamíferos/metabolismo
2.
Artigo em Inglês | MEDLINE | ID: mdl-36708027

RESUMO

BACKGROUND: Diabetes and obesity are associated with muscle atrophy that reduces life quality and lacks effective treatment. Mesenchymal stromal cell (MSC)-based therapy can ameliorate high fat-diet (HFD) and immobilization (IM)-induced muscle atrophy in mice. However, the effect of MSCs on muscle atrophy in type 2 diabetes mellitus (T2DM) and the potential mechanism is unclear. Here, we evaluated the efficacy and explored molecular mechanisms of human umbilical cord MSCs (hucMSCs) and hucMSC-derived exosomes (MSC-EXO) on diabetes- and obesity-induced muscle atrophy. METHODS: Diabetic db/db mice, mice fed with high-fat diet (HFD), mice with hindlimb immobilization (IM), and C2C12 myotubes were used to explore the effect of hucMSCs or MSC-EXO in alleviating muscle atrophy. Grip strength test and treadmill running were used to measure skeletal muscle strength and performance. Body composition, muscle weight, and muscle fibre cross-sectional area (CSA) was used to evaluate muscle mass. RNA-seq analysis of tibialis anterior (TA) muscle and Western blot analysis of muscle atrophy signalling, including MuRF1 and Atrogin 1, were performed to investigate the underlying mechanisms. RESULTS: hucMSCs increased grip strength (P = 0.0256 in db/db mice, P = 0.012 in HFD mice, P = 0.0097 in IM mice), running endurance (P = 0.0154 in HFD mice, P = 0.0006 in IM mice), and muscle mass (P = 0.0004 in db/db mice, P = 0.0076 in HFD mice, P = 0.0144 in IM mice) in all models tested, with elevated CSA of muscle fibres (P < 0.0001 in db/db mice and HFD mice, P = 0.0088 in IM mice) and reduced Atrogin1 (P = 0.0459 in db/db mice, P = 0.0088 in HFD mice, P = 0.0016 in IM mice) and MuRF1 expression (P = 0.0004 in db/db mice, P = 0.0077 in HFD mice, P = 0.0451 in IM mice). MSC-EXO replicated all these hucMSC-mediated changes (P = 0.0103 for grip strength, P = 0.013 for muscle mass, P < 0.0001 for CSA of muscle fibres, P = 0.0171 for Atrogin1 expression, and P = 0.006 for MuRF1 expression). RNA-seq revealed that hucMSCs activated the AMPK/ULK1 signalling and enhanced autophagy. Knockdown of AMPK or inhibition of autophagy with 3-methyladenine (3-MA) diminished the beneficial anti-atrophy effects of hucMSCs or MSC-EXO. CONCLUSIONS: Our results suggest that human umbilical cord mesenchymal stromal cells mitigate diabetes- and obesity-induced muscle atrophy via enhancing AMPK/ULK1-mediated autophagy through exosomes, with implications of applying hucMSCs or hucMSC-derived exosomes to treat muscle atrophy.

3.
Pest Manag Sci ; 2023 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-36693803

RESUMO

BACKGROUND: Although matrine is widely used, the knowledge of its absorption and transport mechanisms in crops remains unexplored. In this study, three methods including foliar application, hydroponics, and seed immersion were used to investigate whether matrine molecules could enter into plants through different channels, and to further resolve its transport characteristics. In addition, the systemic activity of matrine was also evaluated. RESULT: Matrine was quickly absorbed and transported downward after the leaves of wheat or peppers were treated, also accumulated and transmitted upward by roots. It was not only absorbed by seeds, but also appeared continuously in young roots and leaves in both plants for nearly 20 days. There were some differences in the uptake and conduction of matrine between pepper and wheat, mainly concentrating that matrine in pepper upper leaves delivering downward to roots was less than wheat, and also matrine in pepper lower leaves transducing upward to upper leaves was less than wheat. Matrine had systemic activity, with LC50 of 361.99 µg·mL-1 and 904.24 µg·mL-1 against Rhopalosiphum padib on wheat and Myzus persicae (Sulzer) on pepper plants at 48 h, separately. CONCLUSION: Matrine can be absorbed by the roots, seeds, and leaves of plants and transmitted bidirectionally to any organs, presenting satisfactory systemic poisoning activity against aphids. It is of great significance to develop new formulation products of matrine and promote its commercialized value. This article is protected by copyright. All rights reserved.

4.
Free Radic Biol Med ; 195: 219-230, 2023 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-36587924

RESUMO

The function of mitochondrial fusion and fission is one of the important factors causing ischemia-reperfusion (I/R) injury in diabetic myocardium. Aldehyde dehydrogenase 2 (ALDH2) is abundantly expressed in heart, which involved in the regulation of cellular energy metabolism and stress response. However, the mechanism of ALDH2 regulating mitochondrial fusion and fission in diabetic myocardial I/R injury has not been elucidated. In the present study, we found that the expression of ALDH2 was downregulated in rat diabetic myocardial I/R model. Functionally, the activation of ALDH2 resulted in the improvement of cardiac hemodynamic parameters and myocardial injury, which were abolished by the treatment of Daidzin, a specific inhibitor of ALDH2. In H9C2 cardiomyocyte hypoxia-reoxygenation model, ALDH2 regulated the dynamic balance of mitochondrial fusion and fission and maintained mitochondrial morphology stability. Meanwhile, ALDH2 reduced mitochondrial ROS levels, and apoptotic protein expression in cardiomyocytes, which was associated with the upregulation of phosphorylation (p-PI3KTyr458, p-AKTSer473, p-mTOR). Moreover, ALDH2 suppressed the mitoPTP opening through reducing 4-HNE. Therefore, our results demonstrated that ALDH2 alleviated the ischemia and reperfusion injury in diabetic cardiomyopathy through inhibition of mitoPTP opening and activation of PI3K/AKT/mTOR pathway.


Assuntos
Diabetes Mellitus , Cardiomiopatias Diabéticas , Traumatismo por Reperfusão Miocárdica , Ratos , Animais , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Aldeído-Desidrogenase Mitocondrial/genética , Aldeído-Desidrogenase Mitocondrial/metabolismo , Cardiomiopatias Diabéticas/genética , Cardiomiopatias Diabéticas/metabolismo , Dinâmica Mitocondrial/genética , Miócitos Cardíacos/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Isquemia/metabolismo , Apoptose , Diabetes Mellitus/metabolismo
5.
Chem Asian J ; 2023 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-36655414

RESUMO

A series of furoxan derivatives with N-nitroso groups were synthesized in good yields by TBN initiated radical sp3 C-N bond cleavage of 1-nitromethyl-N-aryltetrahydroisoquinolines. This reaction grafts the biologically important furoxan skeleton and N-nitroso group into on molecule, greatly improving the molecular complexity in one step transformation. The mechanistic study shows that this reaction is mediated by the in situ generated α-carbonyl nitrile oxide, which is afforded by TBN promoted C-N bond cleavage.

6.
Med Phys ; 2023 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-36609788

RESUMO

BACKGROUND: Breast cancer is one of the most prevalent malignancies diagnosed in women. Mammogram inspection in the search and delineation of breast tumors is an essential prerequisite for a reliable diagnosis. However, analyzing mammograms by radiologists is time-consuming and prone to errors. Therefore, the development of computer-aided diagnostic (CAD) systems to automate the mass segmentation procedure is greatly expected. PURPOSE: Accurate breast mass segmentation in mammograms remains challenging in CAD systems due to the low contrast, various shapes, and fuzzy boundaries of masses. In this paper, we propose a fully automatic and effective mass segmentation model based on deep learning for improving segmentation performance. METHODS: We propose an effective transformer-based encoder-decoder model (TrEnD). Firstly, we introduce a lightweight method for adaptive patch embedding (APE) of the transformer, which utilizes superpixels to adaptively adjust the size and position of each patch. Secondly, we introduce a hierarchical transformer-encoder and attention-gated-decoder structure, which is beneficial for progressively suppressing interference feature activations in irrelevant background areas. Thirdly, a dual-branch design is employed to extract and fuse globally coarse and locally fine features in parallel, which could capture the global contextual information and ensure the relevance and integrity of local information. The model is evaluated on two public datasets CBIS-DDSM and INbreast. To further demonstrate the robustness of TrEnD, different cropping strategies are applied to these datasets, termed tight, loose, maximal, and mix-frame. Finally, ablation analysis is performed to assess the individual contribution of each module to the model performance. RESULTS: The proposed segmentation model provides a high Dice coefficient and Intersection over Union (IoU) of 92.20% and 85.81% on the mix-frame CBIS-DDSM, while 91.83% and 85.29% for the mix-frame INbreast, respectively. The segmentation performance outperforms the current state-of-the-art approaches. By adding the APE and attention-gated module, the Dice and IoU have improved by 6.54% and 10.07%. CONCLUSION: According to extensive qualitative and quantitative assessments, the proposed network is effective for automatic breast mass segmentation, and has adequate potential to offer technical assistance for subsequent clinical diagnoses.

7.
Neural Regen Res ; 18(5): 1076-1083, 2023 May.
Artigo em Inglês | MEDLINE | ID: mdl-36254996

RESUMO

Studies have shown that gut microbiota metabolites can enter the central nervous system via the blood-spinal cord barrier and cause neuroinflammation, thus constituting secondary injury after spinal cord injury. To investigate the correlation between gut microbiota and metabolites and the possible mechanism underlying the effects of gut microbiota on secondary injury after spinal cord injury, in this study, we established mouse models of T8-T10 traumatic spinal cord injury. We used 16S rRNA gene amplicon sequencing and metabolomics to reveal the changes in gut microbiota and metabolites in fecal samples from the mouse model. Results showed a severe gut microbiota disturbance after spinal cord injury, which included marked increases in pro-inflammatory bacteria, such as Shigella, Bacteroides, Rikenella, Staphylococcus, and Mucispirillum and decreases in anti-inflammatory bacteria, such as Lactobacillus, Allobaculum, and Sutterella. Meanwhile, we identified 27 metabolites that decreased and 320 metabolites that increased in the injured spinal cord. Combined with pathway enrichment analysis, five markedly differential amino acids (L-leucine, L-methionine, L-phenylalanine, L-isoleucine and L-valine) were screened out, which play a pivotal role in activating oxidative stress and inflammatory responses following spinal cord injury. Integrated correlation analysis indicated that the alteration of gut microbiota was related to the differences in amino acids, which suggests that disturbances in gut microbiota might participate in the secondary injury through the accumulation of partial metabolites that activate oxidative stress and inflammatory responses. Findings from this study provide a new theoretical basis for improving the secondary injury after spinal cord injury through fecal microbial transplantation.

8.
BMC Endocr Disord ; 22(1): 325, 2022 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-36539773

RESUMO

BACKGROUND: Thyrotropin-secreting pituitary neuroendocrine tumors (PitNETs) are rare pituitary adenomas that are occasionally accompanied by hypersecretion of other anterior pituitary hormones, such as growth hormone (GH) and prolactin (PRL). The clinical, biochemical, and pathological characteristics may represent diverse circumstances. CASE PRESENTATION: In this report, a 33-year-old female diagnosed with a TSH PitNET co-secreting GH presented no obvious clinical symptoms. The main characteristics were elevated thyroid-stimulating hormone (TSH), free tri-iodothyronine (FT3), and free thyroxine (FT4) levels accompanied by slightly elevated GH and insulin-like growth factor-1 (IGF-1) levels. Magnetic resonance imaging (MRI) detected a pituitary macroadenoma (18 × 16 × 16 mm) with cavernous sinus and suprasellar invasion. Immunohistochemistry revealed diffuse positivity for TSH, strong immunoreactivity for GH, and sporadic positivity for PRL. The electron microscope and double immunofluorescence staining confirmed a plurimorphous plurihormonal adenoma producing TSH, GH, and PRL. After preoperative somatostatin receptor ligand (SRL) treatment and transsphenoidal surgery, the patient achieved temporary clinical and biochemical remission. However, 3 months after surgery, the patient was suspected of having Hashimoto's thyroiditis due to higher thyroglobulin antibody (TGAb), thyroid peroxidase antibody (TPOAb), and thyroid receptor antibody (TRAb) and an enlarged thyroid nodule. During follow-up, thyroid function and TSH slowly transformed from transient hyperthyroidism to hypothyroidism. They were maintained in the normal range by L-T4. CONCLUSION: In the TSH PitNET, the positive immunohistochemistry for TSH, GH, and PRL translated into hormonal overproduction with TSH and GH.


Assuntos
Adenoma , Hormônio do Crescimento Humano , Hipertireoidismo , Neoplasias Hipofisárias , Feminino , Humanos , Adulto , Hipertireoidismo/complicações , Hipertireoidismo/diagnóstico , Neoplasias Hipofisárias/patologia , Adenoma/complicações , Adenoma/diagnóstico , Adenoma/cirurgia , Tireotropina , Hormônio do Crescimento , Prolactina
9.
Front Oncol ; 12: 1040679, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36479063

RESUMO

Background: Ocular graft-versus-host disease (oGVHD) is one of the complications after allogeneic hematopoietic stem cell transplantation (HSCT), which impairs the quality of life and may indicate poor prognosis. In this retrospective study, the aim was to investigate the characteristics of ocular surface after HSCT, and analyze the risk factors related to the severity of ocular surface lesions. Methods: 248 post-HSCT patients were enrolled in this retrospective study. Subjects were divided into no lesion group, mild lesion group and severe lesion group, according to the severity of ocular surface lesions. The correlations between grades of ocular surface lesions and gender, age, primary disease, donor source, human leukocyte antigen (HLA) type, kinship, donor-recipient relationship, blood type, source of stem cell and systemic GVHD were analyzed. Results: The median scores of corneal epitheliopathy, lid margin lesions and meibomian gland loss were 3, 6 and 2 points, respectively. The grade of corneal epitheliopathy was related to donor source (P<0.001), kinship (P=0.033), HLA-matching (P<0.001), and systemic GVHD (P=0.007), especially oral GVHD (P<0.001) and liver GVHD (P=0.002). The grade of lid margin lesions was related to donor source (P=0.019), HLA-matching (P=0.006), and systemic GVHD (P=0.013), especially skin GVHD (P=0.019) and oral GVHD (P=0.019). The grade of meibomian gland loss was related to age (P=0.035) and gastrointestinal GVHD (P=0.007). The grade of corneal epitheliopathy after HSCT was related to the lid margin lesion score (P<0.001). Conclusions: The occurrence and development of ocular GVHD are mostly accompanied by the history of systemic GVHD. While in few cases, ocular surface lesions related to GVHD can be observed prior to the rejection of other tissues and organs. Severe corneal epitheliopathy occurs in patients with severe lid margin lesions in ocular GVHD. The lesions of corneal epithelium and lid margin are milder in HLA partially matching transplantation.

10.
Adv Sci (Weinh) ; : e2204018, 2022 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-36504449

RESUMO

Closure of the neural tube represents a highly complex and coordinated process, the failure of which constitutes common birth defects. The serine/threonine kinase p21-activated kinase 2 (PAK2) is a critical regulator of cytoskeleton dynamics; however, its role in the neurulation and pathogenesis of neural tube defects (NTDs) remains unclear. Here, the results show that Pak2-/- mouse embryos fail to develop dorsolateral hinge points (DLHPs) and exhibit craniorachischisis, a severe phenotype of NTDs. Pak2 knockout activates BMP signaling that involves in vertebrate bone formation. Single-cell transcriptomes reveal abnormal differentiation trajectories and transcriptional events in Pak2-/- mouse embryos during neural tube development. Two nonsynonymous and one recurrent splice-site mutations in the PAK2 gene are identified in five human NTD fetuses, which exhibit attenuated PAK2 expression and upregulated BMP signaling in the brain. Mechanistically, PAK2 regulates Smad9 phosphorylation to inhibit BMP signaling and ultimately induce DLHP formation. Depletion of pak2a in zebrafish induces defects in the neural tube, which are partially rescued by the overexpression of wild-type, but not mutant PAK2. The findings demonstrate the conserved role of PAK2 in neurulation in multiple vertebrate species, highlighting the molecular pathogenesis of PAK2 mutations in NTDs.

11.
Opt Express ; 30(26): 47672-47689, 2022 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-36558690

RESUMO

Achieving high-quality surface profiles under strong ambient light is challenging in fringe projection profilometry (FPP) since ambient light inhibits functional illumination from exhibiting sinusoidal stripes with high quantization levels. Conventionally, large-step phase shifting approaches are presented to enhance the anti-interference capability of FPP, but the image acquisition process in these approaches is highly time-consuming. Inspired by the promising performance of deep learning in optical metrology, we propose a deep learning-enabled anti-ambient light (DLAL) approach that can help FPP extract phase distributions from a single fringe image exposed to unbalanced lighting. In this work, the interference imposed by ambient light on FPP is creatively modeled as ambient light-induced phase error (ALPE). Guided by the ALPE model, we generate the dataset by precisely adjusting the stripe contrast before performing active projection, overcoming the challenge of collecting a large sample of fringe images with various illumination conditions. Driven by the novel dataset, the generated deep learning model can effectively suppress outliers among surface profiles in the presence of strong ambient light, thereby implementing high-quality 3D surface imaging. Experimentally, we verify the effectiveness and adaptability of the proposed DLAL approach in both indoor and outdoor scenarios with strong irradiation.

12.
Dev Cell ; 57(24): 2761-2775.e6, 2022 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-36495874

RESUMO

Spinal cord development is precisely orchestrated by spatiotemporal gene regulatory programs. However, the underlying epigenetic mechanisms remain largely elusive. Here, we profiled single-cell chromatin accessibility landscapes in mouse neural tubes spanning embryonic days 9.5-13.5. We identified neuronal-cell-cluster-specific cis-regulatory elements in neural progenitors and neurons. Furthermore, we applied a novel computational method, eNet, to build enhancer networks by integrating single-cell chromatin accessibility and gene expression data and identify the hub enhancers within enhancer networks. It was experimentally validated in vivo for Atoh1 that knockout of the hub enhancers, but not the non-hub enhancers, markedly decreased Atoh1 expression and reduced dp1/dI1 cells. Together, our work provides insights into the epigenetic regulation of spinal cord development and a proof-of-concept demonstration of enhancer networks as a general mechanism in transcriptional regulation.


Assuntos
Cromatina , Epigênese Genética , Animais , Camundongos , Cromatina/genética , Sequências Reguladoras de Ácido Nucleico , Medula Espinal , Expressão Gênica , Elementos Facilitadores Genéticos/genética
13.
Alzheimers Res Ther ; 14(1): 185, 2022 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-36514127

RESUMO

OBJECTIVE: To investigate the characteristics and associations of MRI-visible perivascular spaces (PVS) with clinical progression and longitudinal cognitive decline across the Alzheimer's disease spectrum. METHODS: We included 1429 participants (641 [44.86%] female) from the Alzheimer's Disease Neuroimaging Initiative (ADNI) database. PVS number and grade in the centrum semiovale (CSO-PVS), basal ganglia (BG-PVS), and hippocampus (HP-PVS) were compared among the control (CN), mild cognitive impairment (MCI), and Alzheimer's disease (AD) groups. PVS were tested as predictors of diagnostic progression (i.e., CN to MCI/AD or MCI to AD) and longitudinal changes in the 13-item Alzheimer's Disease Assessment Scale-cognitive subscale (ADAS-Cog 13), Mini-Mental State Examination (MMSE), memory (ADNI-MEM), and executive function (ADNI-EF) using multiple linear regression, linear mixed-effects, and Cox proportional hazards modeling. RESULTS: Compared with CN subjects, MCI and AD subjects had more CSO-PVS, both in number (p < 0.001) and grade (p < 0.001). However, there was no significant difference in BG-PVS and HP-PVS across the AD spectrum (p > 0.05). Individuals with moderate and frequent/severe CSO-PVS had a higher diagnostic conversion risk than individuals with no/mild CSO-PVS (log-rank p < 0.001 for all) in the combined CN and MCI group. Further Cox regression analyses revealed that moderate and frequent/severe CSO-PVS were associated with a higher risk of diagnostic conversion (HR = 2.007, 95% CI = 1.382-2.914, p < 0.001; HR = 2.676, 95% CI = 1.830-3.911, p < 0.001, respectively). A higher CSO-PVS number was associated with baseline cognitive performance and longitudinal cognitive decline in all cognitive tests (p < 0.05 for all). CONCLUSIONS: CSO-PVS were more common in MCI and AD and were associated with cognitive decline across the AD spectrum.


Assuntos
Doença de Alzheimer , Disfunção Cognitiva , Feminino , Humanos , Masculino , Doença de Alzheimer/diagnóstico , Disfunção Cognitiva/diagnóstico , Testes Neuropsicológicos , Imageamento por Ressonância Magnética , Testes de Estado Mental e Demência
14.
Plant J ; 2022 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-36564995

RESUMO

Carnation (Dianthus caryophyllus L.) is one of the most famous and ethylene-sensitive cut flowers worldwide, but how ethylene interacts with other plant hormones and factors to regulate petal senescence in carnation is largely unknown. Here we found that a gene encoding WRKY family transcription factor, DcWRKY33, was significantly upregulated upon ethylene treatment. Silencing and overexpression of DcWRKY33 could delay and accelerate the senescence of carnation petals, respectively. Abscisic acid (ABA) and H2 O2 treatments could also accelerate the senescence of carnation petals by inducing the expression of DcWRKY33. Further, DcWRKY33 can bind directly to the promoters of ethylene biosynthesis genes (DcACS1 and DcACO1), ABA biosynthesis genes (DcNCED2 and DcNCED5), and the reactive oxygen species (ROS) generation gene DcRBOHB to activate their expression. Lastly, relationships are existed between ethylene, ABA and ROS. This study elucidated that DcWRKY33 promotes petal senescence by activating genes involved in the biosynthesis of ethylene and ABA and accumulation of ROS in carnation, supporting the development of new strategies to prolong the vase life of cut carnation.

15.
Artigo em Inglês | MEDLINE | ID: mdl-36579716

RESUMO

The COVID-19 outbreak has created turbulence and uncertainty into multiple aspects of life in countries around the world. In China, the pandemic continues to pose a great challenge to the nature of traditional in-class education in schools. Chinese education has faced the difficult decision of whether to resume in-person teaching in an unprecedented and time-pressured manner. To ensure the quality of teaching and learning during this time, this study aims to explore the effectiveness of an "online + in-person" hybrid teaching model with a new three-part approach to the hybrid teaching lab, where students prepare for the in-person lab using virtual simulated experiments and learning modules and debrief their learning afterwards online as well. This approach not only enhances the efficiency during the in-person lab but also strongly reinforces concepts and laboratory skills by providing a "practice run" before physically attending the lab. A total of 400 medical undergraduates from Dalian Medical University in China were recruited for this study. In an undergraduate molecular biology laboratory course, we observed 200 students in a hybrid teaching model. We evaluated the learning outcomes from the "online + in-person" hybrid teaching model with a questionnaire survey and assessed the quality of experiment execution, report writing, and group collaboration. Moreover, the 200 students from the hybrid group were evaluated during an annual science competition at the university and compared to 200 students from the competition cohort who had no experience with a hybrid learning model. The comparison data were analyzed using a student's t-test statistical analysis. The students in the hybrid learning group demonstrated a strong enthusiasm for the model, high amount of time utilizing the online system, and high scores on laboratory evaluation assignments. Approximately 98% of the hybrid learning students reported that they preferred mixed teaching to the traditional teaching mode, and all students scored above 96% on the online laboratory report. Teachers of the course observed that the hybrid group had a noticeably higher level of proficiency in lab skills compared to the previous students. At the Dalian Medical University annual science competition, where we compared our hybrid group to a traditional learning group, scores for both the objective and subjective items showed that the students instructed with the hybrid lab model had superior performance (p < 0.05). In the context of the COVID-19 pandemic, we developed a new three-part molecular biology laboratory course that strongly improved students' laboratory skills, knowledge retention, and enthusiasm for the course using online learning to improve their learning efficiency and expedite the in-person laboratory experience. We found that these students performed at a higher level in a combined theoretical/practical science competition compared to the students in traditional in-person lab courses. Additionally, our model subjectively fostered enthusiasm and excellence in both teachers and students. Further, cultivation of the students' independent learning and creative problem solving skills were emphasized. The exploration of an effective teaching model, such as the one described here, not only provides students with a solid foundation for their future medical studies and career development but also promotes more efficient in-person laboratory time.

16.
Dev Comp Immunol ; 139: 104589, 2022 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-36403789

RESUMO

In mammals, type II interferon (IFN; i.e. IFN-γ) signalling transduces through its specific receptors IFN-γR1 and IFN-γR2. In an osteoglossiform fish, the arapaima Arapaima gigas, three type II IFNs, IFN-γ-like, IFN-γ and IFN-γrel, and their four possible receptor subunits IFN-γR1-1, IFN-γR1-2, IFN-γR2-1 and IFN-γR2-2 were identified in this study. The three type II IFN genes are composed of four exons and three introns, and they all contain IFN-γ signature motif and signal peptide, with the presence of potential nuclear localization signal (NLS) in IFN-γ-like and IFN-γ. The IFN-γR1-1, IFN-γR1-2, IFN-γR2-1 and IFN-γR2-2 are composed of seven exons and six introns, with predicted IFN-γR1-1 and IFN-γR1-2 proteins containing JAK1 and STAT1 binding sites, and IFN-γR2-1 and IFN-γR2-2 containing JAK2 binding sites. Gene synteny analysis showed that the type II IFN and their receptor loci are duplicated in arapaima. All these genes were expressed constitutively in all organs/tissues examined, and responded to the stimulation of polyI:C. The prokaryotic recombinant IFN-γ-like, IFN-γ and IFN-γrel proteins can significantly induce the upregulation of immune-related genes in trunk kidney leucocytes. The ligand-receptor relationship analyses revealed that recombinant IFN-γ-like, IFN-γ, and IFN-γrel transduce downstream signalling through IFN-γR1-1/IFN-γR2-1, IFN-γR1-2/IFN-γR2-2, and IFN-γR1-1, respectively, in xenogeneic cells with the overexpression of original or chimeric receptors. In addition, tyrosine (Y) 366 and Y377 in the intracellular region may be essential for the function of IFN-γR1-2 and IFN-γR1-1, respectively. The finding of type II IFN system in A. gigas thus provides different knowledge in understanding the diversity and evolution of type II IFN ligand-receptor relationships in vertebrates.

17.
Front Pharmacol ; 13: 899628, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36386186

RESUMO

REV-ERB agonists have shown antifibrotic effects in the heart and other organs. The function of REV-ERB in the cardiac fibroblasts remains unstudied. Here, we characterize the functional difference of REV-ERB in mouse embryonic fibroblasts and cardiac fibroblasts using genetic deletion of REV-ERBα and ß in vitro. We show that REV-ERB α/ß double deleted cardiac fibroblasts have reduced viability and proliferation, but increased migration and myofibroblasts activation. Thus, REV-ERB α/ß has essential cell-autonomous role in cardiac fibroblasts in maintaining them in a healthy, quiescent state. We also show that existing REV-ERB agonist SR9009 strongly suppresses cardiac fibroblasts activation but in a REV-ERB-independent manner highlighting the need to develop novel REV-ERB agonists for treating cardiac fibrosis.

18.
Drug Deliv ; 29(1): 3317-3327, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36369759

RESUMO

Meloxicam (MLX) is a commonly used drug in the clinical treatment of osteoarthritis, but it is associated with gastrointestinal adverse reactions. Therefore, in this study, we developed a sustained-release microsphere formulation of MLX for topical administration of knee joint. The MLX-loaded PLGA microspheres (MLX-MS) were prepared by emulsion solvent evaporation method with optimization of formulation using orthogonal experimental design. Physicochemical characterization results show MLX-MS were spherical with a smooth surface, the particle size was about 100 µm, drug loading was 30%, and encapsulation efficiency was 76.8%. In addition, the in vivo pharmacokinetics, tissue distribution, and pharmacodynamics were evaluated in rats by intra-articular administration of MLX. The microspheres showed a typical long-term sustained release pattern with a low initial burst release. In contrast to oral administration, local injection of MLX-MS produced a much higher value of elimination half-life time(T1/2) and peak time (Tmax) in plasma, while the intestinal drug distribution was significantly decreased. MLX-MS could also cause a greater reduction in the body level of IL-6 and TNF-α, which was positively correlated with R2=0.981. A good linear relationship (R2 = 0.9945) between the in vitro and in vivo drug release from MLX-MS could be observed, bivariate correlation analysis. All the findings demonstrated that local administration of MLX-MS can prolong the action time of MLX and reduce side effects, thus would be a promising preparation for the treatment of arthritis.


Assuntos
Sistemas de Liberação de Medicamentos , Ratos , Animais , Microesferas , Meloxicam , Preparações de Ação Retardada/química , Sistemas de Liberação de Medicamentos/métodos , Tamanho da Partícula , Injeções Intra-Articulares
19.
J Org Chem ; 87(22): 15139-15151, 2022 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-36398528

RESUMO

Using tris(4-bromophenyl)aminium hexachloroantimonate as a "waste-utilized"-type initiator, the aerobic oxidation of the sp3 C-H bond of proline esters was realized via C-H activation relay, giving a series of halogenated pyrroles in high yields. The mechanistic study revealed that the counterion, SbCl6-, was involved in the radical chlorination process, which provides a new way to understand the role of the counterions.


Assuntos
Halogenação , Compostos Organometálicos , Pirróis , Compostos Organometálicos/química , Compostos de Amônio Quaternário/química
20.
Microbiol Spectr ; : e0317322, 2022 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-36416550

RESUMO

Porcine reproductive and respiratory syndrome virus (PRRSV) is an Arterivirus that has been devastating the swine industry worldwide since the late 1980s. Severe interstitial pneumonia is the typical pathological characteristic of PRRSV-infected swine. Accumulating evidence has suggested that hypoxia-inducible factor 1α (HIF-1α) plays vital roles in the development of inflammation and the viral life cycle. However, the role and the underlying mechanism of HIF-1α in PRRSV infection remain elusive. Here, we found that PRRSV infection elevated HIF-1α expression. Furthermore, overexpression of HIF-1α increased PRRSV replication, whereas knockdown of HIF-1α inhibited PRRSV infection. Our further mechanistic analysis revealed that PRRSV-encoded nonstructural protein 1ß (nsp1ß) promoted HIF-1α transcription via its N-terminal nuclease activity and degraded the polyubiquitin chain of HIF-1α via its C-terminal deubiquitylation (DUB) enzyme activity, collectively stabilizing HIF-1α. Meanwhile, nsp1ß interacted with both HIF-1α and von Hippel-Lindau tumor suppressor (pVHL) to form a ternary complex, which may have hindered pVHL-mediated ubiquitination degradation of HIF-1α by impairing the interaction between HIF-1α and pVHL. Interestingly, pVHL also stabilized nsp1ß via K63-linked ubiquitination, forming a positive feedback loop to stabilize HIF-1α. Taken together, these results indicate that PRRSV infection stabilizes HIF-1α to facilitate viral proliferation and that viral nsp1ß plays a vital role in enhancing the expression and stabilization of HIF-1α. The regulation of HIF-1α may have great therapeutic potential for the development of novel drugs against PRRSV. IMPORTANCE Porcine reproductive and respiratory syndrome virus (PRRSV) has devastated the swine industry worldwide for over 30 years and shows no signs of slowing down. In this study, we found that PRRSV infection elevated hypoxia-inducible factor 1α (HIF-1α) expression. In addition, overexpressed HIF-1α contributed to PRRSV replication, whereas knockdown of HIF-1α reduced PRRSV growth. The PRRSV-encoded nonstructural protein 1ß (nsp1ß) exerted a stabilizing effect on HIF-1α through its nuclease protease and papain-like cysteine protease enzymatic domains. PRRSV nsp1ß also interacted with von Hippel-Lindau tumor suppressor (pVHL) and HIF-1α, whereby nsp1ß impaired the interaction between HIF-1α and pVHL. This work deepens our understanding of the molecular mechanisms involved in PRRSV infection and provides new insights for the development of HIF-1α-based anti-PRRSV therapies.

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