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1.
Front Neurol ; 13: 823882, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35557619

RESUMO

The understanding of brain structural abnormalities across different clinical forms of dystonia and their contribution to clinical characteristics remains unclear. The objective of this study is to investigate shared and specific gray matter volume (GMV) abnormalities in various forms of isolated idiopathic dystonia. We collected imaging data from 73 isolated idiopathic dystonia patients and matched them with healthy controls to explore the GMV alterations in patients and their correlations with clinical characteristics using the voxel-based morphometry (VBM) technique. In addition, we conducted an activation likelihood estimation (ALE) meta-analysis of previous VBM studies. Our study demonstrated widespread morphometry alterations in patients with idiopathic dystonia. Multiple systems were affected, which mainly included basal ganglia, sensorimotor, executive control, and visual networks. As the result of the ALE meta-analysis, a convergent cluster with increased GMV was found in the left globus pallidus. In subgroup VBM analyses, decreased putamen GMV was observed in all clinic forms, while the increased GMV was observed in parahippocampal, lingual, and temporal gyrus. GD demonstrated the most extensive GMV abnormalities in cortical regions, and the aberrant GMV of the posterior cerebellar lobe was prominent in CD. Moreover, trends of increased GMV regions of the left precuneus and right superior frontal gyrus were demonstrated in the moderate-outcome group compared with the superior-outcome group. Results of our study indicated shared pathophysiology of the disease-centered on the dysfunction of the basal ganglia-thalamo-cortical circuit, impairing sensorimotor integration, high-level motor execution, and cognition of patients. Dysfunction of the cerebello-thalamo-cortical circuit could also be involved in CD especially. Finally, the frontal-parietal pathway may act as a potential marker for predicting treatment outcomes such as deep brain stimulation.

2.
Front Vet Sci ; 9: 848833, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35573408

RESUMO

Activation of human immune T-cells by swine leukocyte antigens class I (SLA-I) and class II (SLA-II) leads to xenograft destruction. Here, we generated the GGTA1, B2M, and CIITA (GBC) triple-gene-modified Diannan miniature pigs, analyzed the transcriptome of GBC-modified peripheral blood mononuclear cells (PBMCs) in the pig's spleen, and investigated their effectiveness in anti-immunological rejection. A total of six cloned piglets were successfully generated using somatic cell nuclear transfer, one of them carrying the heterozygous mutations in triple genes and the other five piglets carrying the homozygous mutations in GGTA1 and CIITA genes, but have the heterozygous mutation in the B2M gene. The autopsy of GBC-modified pigs revealed that a lot of spot bleeding in the kidney, severe suppuration and necrosis in the lungs, enlarged peripulmonary lymph nodes, and adhesion between the lungs and chest wall were found. Phenotyping data showed that the mRNA expressions of triple genes and protein expressions of B2M and CIITA genes were still detectable and comparable with wild-type (WT) pigs in multiple tissues, but α1,3-galactosyltransferase was eliminated, SLA-I was significantly decreased, and four subtypes of SLA-II were absent in GBC-modified pigs. In addition, even in swine umbilical vein endothelial cells (SUVEC) induced by recombinant porcine interferon gamma (IFN-γ), the expression of SLA-I in GBC-modified pig was lower than that in WT pigs. Similarly, the expression of SLA-II DR and DQ also cannot be induced by recombinant porcine IFN-γ. Through RNA sequencing (RNA-seq), 150 differentially expressed genes were identified in the PBMCs of the pig's spleen, and most of them were involved in immune- and infection-relevant pathways that include antigen processing and presentation and viral myocarditis, resulting in the pigs with GBC modification being susceptible to pathogenic microorganism. Furthermore, the numbers of human IgM binding to the fibroblast cells of GBC-modified pigs were obviously reduced. The GBC-modified porcine PBMCs triggered the weaker proliferation of human PBMCs than WT PBMCs. These findings indicated that the absence of the expression of α1,3-galactosyltransferase and SLA-II and the downregulation of SLA-I enhanced the ability of immunological tolerance in pig-to-human xenotransplantation.

3.
J Neurosurg ; : 1-11, 2022 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-35523258

RESUMO

OBJECTIVE: Functional connectivity shows the ability to predict the outcome of subthalamic nucleus deep brain stimulation (DBS) in Parkinson disease (PD). However, evidence supporting its value in predicting the outcome of globus pallidus internus (GPi) DBS remains scarce. In this study the authors investigated patient-specific functional connectivity related to GPi DBS outcome in PD and established connectivity models for outcome prediction. METHODS: The authors reviewed the outcomes of 21 patients with PD who received bilateral GPi DBS and presurgical functional MRI at the Ruijin Hospital. The connectivity profiles within cortical areas identified as relevant to DBS outcome in the literature were calculated using the intersection of the volume of tissue activated (VTA) and the local structures as the seeds. Combined with the leave-one-out cross-validation strategy, models of the optimal connectivity profile were constructed to predict outcome. RESULTS: Connectivity between the pallidal areas and primary motor area, supplementary motor area (SMA), and premotor cortex was identified through the literature as related to GPi DBS outcome. The similarity between the connectivity profile within the primary motor area, SMA, pre-SMA, and premotor cortex seeding from the VTA-GPi intersection from an out-of-sample patient and the constructed in-sample optimal connectivity profile predicts GPi DBS outcome (R = 0.58, p = 0.006). The predictions on average deviated by 13.1% ± 11.3% from actual improvements. On the contrary, connectivity profiles seeding from the GPi (R = -0.12, p = 0.603), the VTA (R = 0.23, p = 0.308), the VTA outside the GPi (R = 0.12, p = 0.617), or other local structures were found not to be predictive. CONCLUSIONS: The results showed that patient-specific functional connectivity seeding from the VTA-GPi intersection could help in GPi DBS outcome prediction. Reproducibility remains to be determined across centers in larger cohorts stratified by PD motor subtype.

4.
Front Vet Sci ; 9: 871289, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35433903

RESUMO

Cryopreservation of embryos has been confirmed to cause oxidative stress as a factor responsible for impaired developmental competence. Currently, astaxanthin (Ax) raises considerable interest as a strong exogenous antioxidant and for its potential in reproductive biology. The present study aimed to investigate the beneficial effects of Ax supplementation during in vitro culture of vitrified porcine zygotes and the possible underlying mechanisms. First, the parthenogenetic zygotes were submitted to vitrification and then cultured in the medium added with various concentrations of Ax (0, 0.5, 1.5, and 2.5 µM). Supplementation of 1.5 µM Ax achieved the highest blastocyst yield and was considered as the optimal concentration. This concentration also improved the blastocyst formation rate of vitrified cloned zygotes. Moreover, the vitrified parthenogenetic zygotes cultured with Ax exhibited significantly increased mRNA expression of CDX2, SOD2, and GPX4 in their blastocysts. We further analyzed oxidative stress, mitochondrial and lysosomal function in the 4-cell embryos and blastocysts derived from parthenogenetic zygotes. For the 4-cell embryos, vitrification disturbed the levels of reactive oxygen species (ROS) and glutathione (GSH), and the activities of mitochondria, lysosome and cathepsin B, and Ax supplementation could fully or partially rescue these values. The blastocysts obtained from vitrified zygotes showed significantly reduced ATP content and elevated cathepsin B activity, which also was recovered by Ax supplementation. There were no significant differences in other parameters mentioned above for the resultant blastocysts. Furthermore, the addition of Ax significantly enhanced mitochondrial activity and reduced lysosomal activity in resultant blastocysts. In conclusion, these findings revealed that Ax supplementation during the culture period improved subsequent embryonic development and quality of porcine zygotes after vitrification and might be used to ameliorate the recovery culture condition for vitrified embryos.

5.
Anim Biotechnol ; : 1-10, 2022 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-35404767

RESUMO

Interspecies somatic cell nuclear transfer (iSCNT) has an immense potential to rescue endangered animals and extinct species like mammoths. In this study, we successfully established an Asian elephant's fibroblast cell lines from ear tissues, performed iSCNT with porcine oocytes and evaluated the in vitro and in vivo development of reconstructed embryos. A total of 7780 elephant-pig iSCNT embryos were successfully reconstructed and showed in vitro development with cleavage rate, 4-cell, 8-cell and blastocyst rate of 73.01, 30.48, 5.64, and 4.73%, respectively. The total number of elephant-pig blastocyte cells and diameter of hatched blastocyte was 38.67 and 252.75 µm, respectively. Next, we designed species-specific markers targeting EDNRB, AGRP and TYR genes to verify the genome of reconstructed embryos with donor nucleus/species. The results indicated that 53.2, 60.8, and 60.8% of reconstructed embryos (n = 235) contained elephant genome at 1-cell, 2-cell and 4-cell stages, respectively. However, the percentages decreased to 32.3 and 32.7% at 8-cell and blastocyst stages, respectively. Furthermore, we also evaluated the in vivo development of elephant-pig iSCNT cloned embryos and transferred 2260 reconstructed embryos into two surrogate gilts that successfully became pregnant and a total of 11 (1 and 10) fetuses were surgically recovered after 17 and 19 days of gestation, respectively. The crown-rump length and width of elephant-pig cloned fetuses were smaller than the control group. Unfortunately, none of these fetuses contained elephant genomes, which suggested that elephant embryos failed to develop in vivo. In conclusion, we successfully obtained elephant-pig reconstructed embryos for the first time and these embryos are able to develop to blastocyst, but the in vivo developmental failure needs further investigated.

6.
Front Endocrinol (Lausanne) ; 13: 844351, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35273576

RESUMO

Background: It is of great clinical importance to assess the microstructure of the articular cartilage and cortical bone of the human knee joint. While quantitative susceptibility mapping (QSM) is a promising tool for investigating the knee joint, however, previous QSM studies using conventional gradient recalled echo sequences or ultrashort echo time (UTE) sequences only focused on mapping the magnetic susceptibility of the articular cartilage or cortical bone, respectively. Simultaneously mapping the underlying susceptibilities of the articular cartilage and cortical bone of human in vivo has not been explored and reported. Method: Three-dimensional multi-echo radial UTE sequences with the shortest TE of 0.07 msec and computed tomography (CT) were performed on the bilateral knee joints of five healthy volunteers for this prospective study. UTE-QSM was reconstructed from the local field map after water-fat separation and background field removal. Spearman's correlation analysis was used to explore the relationship between the magnetic susceptibility and CT values in 158 representative regions of interest of cortical bone. Result: The susceptibility properties of the articular cartilage and cortical bone were successfully quantified by UTE-QSM. The laminar structure of articular cartilage was characterized by the difference of susceptibility value in each layer. Susceptibility was mostly diamagnetic in cortical bone. A significant negative correlation (r=-0.43, p<0.001) between the susceptibility value and CT value in cortical bone was observed. Conclusion: UTE-QSM enables simultaneous susceptibility mapping of the articular cartilage and cortical bone of human in vivo. Good association between susceptibility and CT values in cortical bone suggests the potential of UTE-QSM for bone mapping for further clinical application.


Assuntos
Cartilagem Articular , Cartilagem Articular/diagnóstico por imagem , Osso Cortical/diagnóstico por imagem , Humanos , Imageamento Tridimensional , Articulação do Joelho/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Estudos Prospectivos
7.
Cell Biosci ; 12(1): 26, 2022 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-35255981

RESUMO

BACKGROUND: Mice with humanized livers are important models to study drug toxicology testing, development of hepatitis virus treatments, and hepatocyte transplantation therapy. However, the huge difference between mouse and human in size and anatomy limited the application of humanized mice in investigating human diseases. Therefore, it is urgent to construct humanized livers in pigs to precisely investigate hepatocyte regeneration and human hepatocyte therapy. CRISPR/Cas9 system and somatic cell cloning technology were used to generate two pig models with FAH deficiency and exhibiting severe immunodeficiency (FAH/RAG1 and FAH/RAG1/IL2RG deficiency). Human primary hepatocytes were then successfully transplanted into the FG pig model and constructed two pigs with human liver. RESULTS: The constructed FAH/RAG1/IL2RG triple-knockout pig models were characterized by chronic liver injury and severe immunodeficiency. Importantly, the FG pigs transplanted with primary human hepatocytes produced human albumin in a time dependent manner as early as 1 week after transplantation. Furthermore, the colonization of human hepatocytes was confirmed by immunochemistry staining. CONCLUSIONS: We successfully generated pig models with severe immunodeficiency that could construct human liver tissues.

8.
Front Neurorobot ; 16: 848746, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35295674

RESUMO

Objective: Our study aimed to develop an approach to improve the speed and resolution of cerebral-hemisphere and lesion modeling and evaluate the advantages and disadvantages of robot-assisted surgical planning software. Methods: We applied both conventional robot planning software (method 1) and open-source auxiliary software (FreeSurfer and 3D Slicer; method 2) to model the brain and lesions in 19 patients with drug-resistant epilepsy. The patients' mean age at implantation was 21.4 years (range, 6-52 years). Each patient received an average of 12 electrodes (range, 9-16) between May and November 2021. The electrode-implantation plan was designed based on the models established using the two methods. We statistically analyzed and compared the duration of designing the models and planning the implantation using these two methods and performed the surgeries with the implantation plan designed using the auxiliary software. Results: A significantly longer time was needed to reconstruct a cerebral-hemisphere model using method 1 (mean, 206 s) than using method 2 (mean, 20 s) (p < 0.05). Both methods identified a mean of 1.4 lesions (range, 1-5) in each patient. Overall, using method 1 required longer (mean, 130 s; range, 48-436) than using method 2 (mean, 68.1 s; range, 50-104; p < 0.05). In addition, the clarity of the model based on method 1 was lower than that based on method 2. To devise an electrode-implantation plan, it took 9.1-25.5 min (mean, 16) and 6.6-14.8 min (mean, 10.2) based on methods 1 and 2, respectively (p < 0.05). The average target point error of 231 electrodes amounted to 1.90 mm ± 0.37 mm (range, 0.33-3.61 mm). The average entry point error was 0.89 ± 0.26 mm (range, 0.17-1.67 mm). None of the patients presented with intracranial hemorrhage or infection, and no other serious complications were observed. Conclusions: FreeSurfer and 3D Slicer-assisted SEEG implantation is an excellent approach to enhance modeling speed and resolution, shorten the electrode-implantation planning time, and boost the efficiency of clinical work. These well-known, trusted open-source programs do not have explicitly restricted licenses. These tools, therefore, seem well suited for clinical-research applications under the premise of approval by an ethics committee, informed consent of the patient, and clinical judgment of the surgeon.

9.
Vet Sci ; 9(3)2022 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-35324845

RESUMO

Considerable improvements in sheep multiple ovulation and embryo transfer (MOET)protocols have been made; however, unlike for cattle, MOET is poorly developed in sheep, and thus has not been broadly applicable as a routine procedure. The tightly folded nature of the ewe cervix, the inconsistent ovarian response to various superovulatory treatments, and the requirement of labor to handle animals, particularly during large-scale production, has limited the implementation of successful MOET in sheep. Moreover, several extrinsic factors (e.g., sources, the purity of gonadotrophins and their administration) and intrinsic factors (e.g., breed, age, nutrition, reproductive status) severely limit the practicability of MOET in sheep and other domestic animals. In this review, we summarize the effects of different superovulatory protocols, and their respective ovarian responses, in terms of ovulation rate, and embryo recovery and transfer. Furthermore, various strategies, such as inhibin immunization, conventional superovulation protocols, and melatonin implants for improving the ovarian response, are discussed in detail. Other reproductive techniques and their relative advantages and disadvantages, such as artificial insemination (AI), and donor embryo recovery and transfer to the recipient through different procedures, which must be taken into consideration for achieving satisfactory results during any MOET program in sheep, are also summarized in this article.

10.
Front Neurol ; 13: 797649, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35211082

RESUMO

BACKGROUND: Fragile X premutation carriers (55-200 CGG triplets) may develop a progressive neurodegenerative disorder, fragile X-associated tremor/ataxia syndrome (FXTAS), after the age of 50. The neuroradiologic markers of FXTAS are hyperintense T2-signals in the middle cerebellar peduncle-the MCP sign. We recently noticed abnormal T2-signals in the globus pallidus in male premutation carriers and controls but the prevalence and clinical significance were unknown. METHODS: We estimated the prevalence of the MCP sign and pallidal T2-abnormalities in 230 male premutation carriers and 144 controls (aged 8-86), and examined the associations with FXTAS symptoms, CGG repeat length, and iron content in the cerebellar dentate nucleus and globus pallidus. RESULTS: Among participants aged ≥45 years (175 premutation carriers and 82 controls), MCP sign was observed only in premutation carriers (52 vs. 0%) whereas the prevalence of pallidal T2-abnormalities approached significance in premutation carriers compared with controls after age-adjustment (25.1 vs. 13.4%, p = 0.069). MCP sign was associated with impaired motor and executive functioning, and the additional presence of pallidal T2-abnormalities was associated with greater impaired executive functioning. Among premutation carriers, significant iron accumulation was observed in the dentate nucleus, and neither pallidal or MCP T2-abnormalities affected measures of the dentate nucleus. While the MCP sign was associated with CGG repeat length >75 and dentate nucleus volume correlated negatively with CGG repeat length, pallidal T2-abnormalities did not correlate with CGG repeat length. However, pallidal signal changes were associated with age-related accelerated iron depletion and variability and having both MCP and pallidal signs further increased iron variability in the globus pallidus. CONCLUSIONS: Only the MCP sign, not pallidal abnormalities, revealed independent associations with motor and cognitive impairment; however, the occurrence of combined MCP and pallidal T2-abnormalities may present a risk for greater cognitive impairment and increased iron variability in the globus pallidus.

11.
Artigo em Inglês | MEDLINE | ID: mdl-35170794

RESUMO

OBJECTIVES: Cushing's disease (CD) is most common endogenous Cushing's syndrome. This study aimed to assess iron alternations in deep grey matter in CD. DESIGN: A cross-sectional study was performed. PATIENTS: In this study, 48 active CD patients, 39 remitted CD patients and 52 healthy control (HC) subjects underwent magnetic resonance imaging. MEASUREMENTS: Quantitative susceptibility mapping (QSM). RESULTS: Decreased susceptibility values were found in the bilateral putamen, caudate, red nucleus, subthalamic nucleus and pulvinar nuclei of the thalamus (TL-PLV) in active and remitted patients with CD compared with HCs. Interestingly, in remitted patients with CD, altered susceptibility values were significantly correlated with altered brain volumes in TL-PLV, while TL-PLV may play an essential role as a general regulatory hub for adaptive and flexible cognition. CONCLUSION: Chronic exposure to hypercortisolism may be related to iron distribution and significantly correlated with altered brain volumes and clinical features in patients with CD.

12.
Magn Reson Imaging ; 88: 89-100, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35124180

RESUMO

PURPOSE: This study developed a data-driven optimization to improve the accuracy of deep learning QSM quantification. METHODS: The proposed deep learning QSM pipeline consisted of two projections onto convex set (POCS) models designed to decouple trainable network components with the spherical mean value (SMV) filters and dipole kernel in the data-driven optimization. They were a background field removal network (named POCSnet1) and a dipole inversion network (named POCSnet2). Both POCSnet1 and POCSnet2 were the unrolled V-Net with iterative data-driven optimization to enforce the data fidelity. For training POCSnet1, we simulated phantom data with random geometric shapes as the background susceptibility sources. For training POCSnet2, we used geometric shapes to mimic the QSM. The evaluation was performed on synthetic data, a public COSMOS (N = 1), and clinical data from a Parkinson's disease cohort (N = 71) and small-vessel disease cohort (N = 26). For comparison, DLL2, FINE, and autoQSM, were implemented and tested under the same experimental setting. RESULTS: On COSMOS, results from POCSnet1 were more similar to that of the V-SHARP method with NRMSE = 23.7% and SSIM = 0.995, compared with the NRMSE = 62.7% and SSIM = 0.975 for SHARQnet, a naïve V-Net model. On COSMOS, the NRMSE and HFEN for POCSnet2 were 58.1% and 56.7%; while for DLL2, FINE, and autoQSM, they were 62.0% and 61.2%, 69.8% and 67.5%, and 87.5% and 85.3%, respectively. On the Parkinson's disease cohort, our results were consistent with those obtained from VSHARP+STAR-QSM with biases <3% and outperformed the SHARQnet+DeepQSM that had biases of 7% to 10%. The sensitivity of cerebral microbleed detection using our pipeline was 100%, compared with 92% by SHARQnet+DeepQSM. CONCLUSION: Data-driven optimization improved the accuracy of QSM quantification compared with that of naïve V-Net models.


Assuntos
Aprendizado Profundo , Algoritmos , Encéfalo/diagnóstico por imagem , Mapeamento Encefálico/métodos , Humanos , Processamento de Imagem Assistida por Computador/métodos , Imageamento por Ressonância Magnética/métodos
13.
Mol Med Rep ; 25(2)2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35014689

RESUMO

Triple­negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, and it often becomes resistant to paclitaxel (PTX) therapy. Autophagy plays an important cytoprotective role in PTX­induced tumor cell death, and targeting autophagy has been promising for improving the efficacy of tumor chemotherapy in recent years. The aim of the present study was to clarify the mechanism of PTX inducing autophagy in TNBC cells to provide a potential clinical chemotherapy strategy of PTX for TNBC. The present study reported that PTX induced both apoptosis and autophagy in MDA­MB­231 cells and that inhibition of autophagy promoted apoptotic cell death. Furthermore, it was found that forkhead box transcription factor O1 (FOXO1) enhanced PTX­induced autophagy through a transcriptional activation pattern in MDA­MB­231 cells, which was associated with the downstream target genes autophagy related 5, class III phosphoinositide 3­kinase vacuolar protein sorting 34, autophagy related 4B cysteine peptidase, beclin 1 and microtubule associated protein 1 light chain 3ß. Knocking down FOXO1 attenuated the survival of MDA­MB­231 cells in response to PTX treatment. These findings may be beneficial for improving the treatment efficacy of PTX and to develop autophagic targeted therapy for TNBC.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proteína Forkhead Box O1/metabolismo , Paclitaxel/farmacologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismo , Adenina/análogos & derivados , Adenina/farmacologia , Proteína 5 Relacionada à Autofagia/metabolismo , Proteína Beclina-1/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Classe III de Fosfatidilinositol 3-Quinases/metabolismo , Cisteína Endopeptidases/metabolismo , Resistencia a Medicamentos Antineoplásicos/fisiologia , Proteína Forkhead Box O1/genética , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo
14.
Transgenic Res ; 31(1): 59-72, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34741281

RESUMO

Leptin is a well-known adipokine that plays critical role in adiposity. To further investigate the role of leptin in adiposity, we utilized leptin overexpressing transgenic pigs and evaluated the effect of leptin on growth and development, fat deposition, and lipid metabolism at tissue and cell level. Leptin transgenic pigs were produced and divided into two groups: elevated leptin expression (leptin ( +)) and normal leptin expression group (control). Results indicated that leptin ( +) pigs had elevated leptin protein and mRNA expression levels and exhibited sluggish growth and development followed by decreased subcutaneous fat thickness, low serum triglycerides, saturated, unsaturated fatty acids and high cholesterol esters (p < 0.05). There were differences in the lipid metabolism related genes at different fat depots, including upregulation of PPARγ, AGPAT6, PLIN2, HSL and ATGL in subcutaneous, PPARγ in perirenal, and FAT/CD36 and PLIN2 in mesenteric adipose tissues and downregulation of AGPAT6 and ATGL in perirenal and AGPAT6 in mesenteric adipose tissues (p < 0.05). Additionally, in-vitro cultured leptin ( +) preadipocytes exhibited upregulation of PPARγ, FAT/CD36, ACACA, AGPAT, PLIN2, ATGL and HSL as compared to control (p < 0.05). These findings suggested that homeostasis imbalance in lipolysis and lipogenesis at adipose tissue and adipocytes levels led to low subcutaneous fat depots in leptin overexpression pigs. These pigs can act as model for obesity and related metabolic disorder.


Assuntos
Leptina , PPAR gama , Tecido Adiposo/metabolismo , Animais , Leptina/genética , Leptina/metabolismo , Lipólise , Obesidade/genética , PPAR gama/genética , PPAR gama/metabolismo , PPAR gama/farmacologia , Suínos/genética , Triglicerídeos/genética
15.
Cell Res ; 32(4): 383-400, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-34848870

RESUMO

Pig epiblast-derived pluripotent stem cells are considered to have great potential and broad prospects for human therapeutic model development and livestock breeding. Despite ongoing attempts since the 1990s, no stably defined pig epiblast-derived stem cell line has been established. Here, guided by insights from a large-scale single-cell transcriptome analysis of pig embryos from embryonic day (E) 0 to E14, specifically, the tracing of pluripotency changes during epiblast development, we developed an in vitro culture medium for establishing and maintaining stable pluripotent stem cell lines from pig E10 pregastrulation epiblasts (pgEpiSCs). Enabled by chemical inhibition of WNT-related signaling in combination with growth factors in the FGF/ERK, JAK/STAT3, and Activin/Nodal pathways, pgEpiSCs maintain their pluripotency transcriptome features, similar to those of E10 epiblast cells, and normal karyotypes after more than 240 passages and have the potential to differentiate into three germ layers. Strikingly, ultradeep in situ Hi-C analysis revealed functional impacts of chromatin 3D-spatial associations on the transcriptional regulation of pluripotency marker genes in pgEpiSCs. In practice, we confirmed that pgEpiSCs readily tolerate at least three rounds of successive gene editing and generated cloned gene-edited live piglets. Our findings deliver on the long-anticipated promise of pig pluripotent stem cells and open new avenues for biological research, animal husbandry, and regenerative biomedicine.


Assuntos
Camadas Germinativas , Células-Tronco Pluripotentes , Animais , Diferenciação Celular/genética , Linhagem Celular , Suínos , Transcriptoma
16.
Annu Int Conf IEEE Eng Med Biol Soc ; 2021: 3676-3679, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34892034

RESUMO

Deep brain nuclei are closely related to the pathogenesis of neurodegenerative diseases. Automatic segmentation for brain nuclei plays a significant role in aging and disease-related assessment. Quantitative susceptibility mapping (QSM), as a novel MRI imaging technique, attracts increasing attention in deep gray matter (DGM) nuclei-related research and diagnosis. This paper proposes DeepQSMSeg, a deep learning-based end-to-end tool, to segment five pairs of DGM structures from QSM images. The proposed model is based on a 3D encoder-decoder fully convolutional neural network. For concentrating network on the target regions, spatial and channel attention modules are adopted in both encoder and decoder stages. Dice loss is combined with focal loss to alleviate the imbalance of ROI classes. The result shows that our method can segment DGM structures from QSM images precisely, rapidly and reliably. Comparing with ground truth, the average Dice coefficient for all ROIs in the test dataset achieved 0.872±0.053, and Hausdorff distance was 2.644±2.917 mm. Finally, an age-related susceptibility development model was used to confirm the reliability of DeepQSMSeg in aging and disease-related studies.Clinical Relevance-Accurate and automatic segmentation tool for sub-cortical regions in QSM can significantly alleviate the pressure of radiologists. It can also accelerate the progress of related research and clinical translation.


Assuntos
Aprendizado Profundo , Processamento de Imagem Assistida por Computador , Substância Cinzenta , Imageamento por Ressonância Magnética , Reprodutibilidade dos Testes
17.
Life (Basel) ; 11(12)2021 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-34947951

RESUMO

The base editing 3 (BE3) system, a single-base gene editing technology developed using CRISPR/Cas9n, has a broad range of applications for human disease model construction and gene therapy, as it is highly efficient, accurate, and non-destructive. P53 mutations are present in more than 50% of human malignancies. Due to the similarities between humans and pigs at the molecular level, pig models carrying P53 mutations can be used to research the mechanism of tumorigenesis and improve tumor diagnosis and treatment. According to pathogenic mutations of the human P53 gene at W146* and Q100*, sgRNAs were designed to target exon 4 and exon 5 of the porcine P53 gene. The target editing efficiencies of the two sgRNAs were 61.9% and 50.0%, respectively. The editing efficiency of the BE3 system was highest (about 60%) when C (or G) was at the 5th base. Puromycin screening revealed that 75.0% (21/28) and 68.7% (22/32) of cell colonies contained a P53 mutation at sgRNA-Exon5 and sgRNA-Exon4, respectively. The reconstructed embryos from sgRNA-Exon5-5# were transferred into six recipient gilts, all of which aborted. The reconstructed embryos from sgRNA-Exon4-7# were transferred into 6 recipient gilts, 3 of which became pregnant, resulting in 14 live and 3 dead piglets. Sequencing analyses of the target site confirmed 1 P53 monoallelic mutation and 16 biallelic mutations. The qPCR analysis showed that the P53 mRNA expression level was significantly decreased in different tissues of the P53 mutant piglets (p < 0.05). Additionally, confocal microscopy and western blot analysis revealed an absence of P53 expression in the P53 mutant fibroblasts, livers, and lung tissues. In conclusion, a porcine cancer model with a P53 point mutation can be obtained via the BE3 system and somatic cell nuclear transfer (SCNT).

18.
Front Cell Dev Biol ; 9: 712224, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34616727

RESUMO

Porcine-induced pluripotent stem cells (piPSCs) could serve as a great model system for human stem cell preclinical research. However, the pluripotency gene network of piPSCs, especially the function for the core transcription factor estrogen-related receptor beta (ESRRB), was poorly understood. Here, we constructed ESRRB-overexpressing piPSCs (ESRRB-piPSCs). Compared with the control piPSCs (CON-piPSCs), the ESRRB-piPSCs showed flat, monolayered colony morphology. Moreover, the ESRRB-piPSCs showed greater chimeric capacity into trophectoderm than CON-piPSCs. We found that ESRRB could directly regulate the expressions of trophoblast stem cell (TSC)-specific markers, including KRT8, KRT18 and CDX2, through binding to their promoter regions. Mutational analysis proved that the N-terminus zinc finger domain is indispensable for ESRRB to regulate the TSC markers. Furthermore, this regulation needs the participation of OCT4. Accordingly, the cooperation between ESRRB and OCT4 facilitates the conversion from pluripotent state to the trophoblast-like state. Our results demonstrated a unique and crucial role of ESRRB in determining piPSCs fate, and shed new light on the molecular mechanism underlying the segregation of embryonic and extra-embryonic lineages.

19.
Cell Cycle ; 20(21): 2264-2277, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34583621

RESUMO

Considerable advancements have recently been achieved in porcine somatic cell nuclear transfer (SCNT), but the efficiency remains low. Donor cell size might play an important role in SCNT, but its effects in pigs remain unclear. This study aimed to evaluate the efficiency of porcine SCNT by selecting donor cells of suitable size. Porcine fetal fibroblasts (PFFs) were divided into three groups, group S (small, d ≤ 13 µm), group M (medium, 13 µm 18 µm), and their biological characteristics were analyzed. Next, SCNT was performed using PFFs of different sizes to evaluate the developmental potential of reconstructed embryos. The data showed that PFFs in groups S, M and L accounted for 17.5%, 47.7% and 34.8% of cells, respectively. Morphologically, cells in group S exhibited clear and regular cell membranes and nuclei, whereas cells in groups M and L displayed varying degrees of cell membrane protuberance, karyo-pyknosis, autophagy and mitochondrial abnormalities. In addition, the growth status and proliferation capabilities of cells in group S were significantly better than those of group M and group L. The percentage of cells at G0/G1 in group S and M were significantly greater than group L. The senescence rate of group S was lower than group M and group L. The apoptosis rate of group S was significantly lower than that of group L but comparable to that of group M . The cleavage rate of group S was also significantly greater than that of group M but comparable to that of group L . The blastocyst rate of group S was significantly greater than that of group M and group L. The blastocyst cell numbers of group S were also significantly greater than those of group M and group L. These findings suggested that small PFFs with a diameter of less than 13 µm are more suitable donor cells for SCNT in pigs.Abbreviations: DMEM: Dulbecco's modified Eagle's medium; FBS: Fetal bovine serum; PBS: Phosphate buffer saline; PFFs: Porcine fetal fibroblast cells; SCNT: Somatic cell nuclear transfer.


Assuntos
Blastocisto , Técnicas de Transferência Nuclear , Animais , Blastocisto/metabolismo , Tamanho Celular , Clonagem de Organismos , Embrião de Mamíferos , Desenvolvimento Embrionário , Feminino , Feto , Fibroblastos/metabolismo , Gravidez , Suínos
20.
Front Neurosci ; 15: 731109, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34557069

RESUMO

BACKGROUND: Emerging evidence indicates that iron distribution is heterogeneous within the substantia nigra (SN) and it may reflect patient-specific trait of Parkinson's Disease (PD). We assume it could account for variability in motor outcome of subthalamic nucleus deep brain stimulation (STN-DBS) in PD. OBJECTIVE: To investigate whether SN susceptibility features derived from radiomics with machine learning (RA-ML) can predict motor outcome of STN-DBS in PD. METHODS: Thirty-three PD patients underwent bilateral STN-DBS were recruited. The bilateral SN were segmented based on preoperative quantitative susceptibility mapping to extract susceptibility features using RA-ML. MDS-UPDRS III scores were recorded 1-3 days before and 6 months after STN-DBS surgery. Finally, we constructed three predictive models using logistic regression analyses: (1) the RA-ML model based on radiomics features, (2) the RA-ML+LCT (levodopa challenge test) response model which combined radiomics features with preoperative LCT response, (3) the LCT response model alone. RESULTS: For the predictive performances of global motor outcome, the RA-ML model had 82% accuracy (AUC = 0.85), while the RA-ML+LCT response model had 74% accuracy (AUC = 0.83), and the LCT response model alone had 58% accuracy (AUC = 0.55). For the predictive performance of rigidity outcome, the accuracy of the RA-ML model was 80% (AUC = 0.85), superior to those of the RA-ML+LCT response model (76% accuracy, AUC = 0.82), and the LCT response model alone (58% accuracy, AUC = 0.42). CONCLUSION: Our findings demonstrated that SN susceptibility features from radiomics could predict global motor and rigidity outcomes of STN-DBS in PD. This RA-ML predictive model might provide a novel approach to counsel candidates for STN-DBS.

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