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Visualised genotyping assay with oral swabs in a closed tube by nested invasive reaction assisted with gold nanoparticle probes 封闭管内口腔拭子可视化基因分型分析,巢式侵入性反应辅助金纳米颗粒探针
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-03-10 DOI: 10.1049/nbt2.12123
Yijun Li, Wei Wei, Yi Ma, Jingwen Shan, Yanan Chu, LiKun Zhang, Danni Liu, Xueping Ma, Guohua Zhou, Haiping Wu

Single nucleotide polymorphism (SNP) typing is crucial for drug dosage and disease progression. Therefore, a simple and convenient genotyping assay is essential for personalised medicine. Herein, we developed a non-invasive, closed-tube, and visualised method for genotyping. In this method, oral swabs were lysed to directly perform PCR coupled with nested invasive reaction and visualisation based on gold nanoparticle probes in a closed tube. The strategy for genotyping assay depends on the single base recognition property of invasive reaction. This assay allowed quick and simple sample preparation and the detection of 25 copies/μL of CYP2C19*2 and 100 copies/μL of CYP2C19*3 within 90 min. Further, 20 oral swab samples for CYP2C19*2 and CYP2C19*3 were correctly typed, which agreed with pyrosequencing, indicating that this method has great potential for SNP typing in source-limited regions to guide personalised medicine.

单核苷酸多态性(SNP)分型对药物剂量和疾病进展至关重要。因此,一种简单方便的基因分型检测方法对于个性化医疗至关重要。在此,我们开发了一种非侵入性、闭管和可视化的基因分型方法。在这种方法中,口腔拭子被裂解,直接进行PCR,结合巢式侵入反应和基于封闭管中的金纳米颗粒探针的可视化。基因分型分析的策略取决于侵袭性反应的单碱基识别特性。该方法可快速简便地制备样品,在90 min内检测到CYP2C19*2的25拷贝/μL和CYP2C19*3的100拷贝/μL。此外,20份CYP2C19*2和CYP2C19*3的口腔拭子样品正确分型,与pyrosequencing一致,表明该方法在来源有限的地区具有很大的SNP分型潜力,可指导个性化用药。
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引用次数: 0
Preparation, characterisation, and in vitro cancer-suppression function of RNA nanoparticles carrying miR-301b-3p Inhibitor 携带miR-301b-3p抑制剂的RNA纳米颗粒的制备、表征及其体外抑癌功能
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-03-09 DOI: 10.1049/nbt2.12120
Junjie Zhao, Niu Niu, Fan Yang, Haitao Liu, Weibo Qi

Background

Multidrug resistance is the biggest barrier on the way to chemotherapy for lung adenocarcinoma (LUAD). For some LUAD patients with cisplatin (DDP) resistance and poor prognoses, the authors put forward RNA nanoparticles (NPs) carrying miR-301b-3p Inhibitor.

Methods

The NPs were composed of miR-301b-3p, A549 aptamer (A549apt), and Cyanine 5 in a bottom-up manner with a 3-way-junction (3WJ) structure. Diameter, assembly process, and morphology of NPs were observed by Dynamic Light Scattering, Native-Polyacrylamide Gel Electrophoresis, and Atomic Force Microscopy. Cell internalisation, toxicity, proliferation, migration, invasion, and apoptosis were assayed by confocal laser scanning microscope, CCK8, colony formation assay, Transwell, western blot, and flow cytometry.

Results

3WJ-apt-miR was evenly distributed, with diameter of 19.61 ± 0.49 nm and triangular branching structures. The accurate delivery of this NP in vivo was ensured by A549 aptamer featuring specific targeting, with smaller side effects than traditional chemotherapy. Such nanomaterials were effectively internalized by cancer cells, with normal cell activity intact. Cancer cell proliferation, invasion, and migration were suppressed, and DDP sensitivity was enhanced, causing DNA damage and facilitating apoptosis of DDP-resistant cells.

Conclusion

Based on RNA self-assembling, the authors researched the effect of miRNA on DDP sensitivity in LUAD regarding gene regulation. 3WJ-apt-miR paves the way for clinical tumour therapy.

多药耐药是肺腺癌(LUAD)化疗的最大障碍。对于部分顺铂(DDP)耐药且预后不良的LUAD患者,作者提出了携带miR-301b-3p抑制剂的RNA纳米颗粒(NPs)。方法NPs由miR-301b-3p、A549适配体(A549apt)和Cyanine 5以自下而上的方式组成,具有3路结(3WJ)结构。通过动态光散射、原生聚丙烯酰胺凝胶电泳和原子力显微镜观察纳米粒子的直径、组装过程和形态。采用共聚焦激光扫描显微镜、CCK8、集落形成实验、Transwell、western blot和流式细胞术检测细胞内化、毒性、增殖、迁移、侵袭和凋亡。结果3WJ-apt-miR分布均匀,直径为19.61±0.49 nm,呈三角形分支结构。A549适体具有特异性靶向性,保证了这种NP在体内的准确递送,其副作用比传统化疗小。这种纳米材料被癌细胞有效地内化,正常细胞活性完好无损。抑制了癌细胞的增殖、侵袭和迁移,增强了DDP敏感性,造成DNA损伤,促进DDP耐药细胞凋亡。结论基于RNA自组装,从基因调控角度研究了miRNA对LUAD患者DDP敏感性的影响。3WJ-apt-miR为临床肿瘤治疗铺平了道路。
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引用次数: 1
Evaluation of the parameters affecting the loading of anticancer drug Paclitaxel on coated gold nanoparticles for breast cancer treatment 影响抗癌药物紫杉醇在包覆金纳米颗粒上的负载的参数评价
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-02-27 DOI: 10.1049/nbt2.12121
Afrooz kharazmi, Neda Attaran

The purpose of this study is the design and synthesis of gold nanoparticles (GNPs) conjugated with paclitaxel and to investigate the parameters affecting the stability of synthesised nanoparticles with drug delivery capability. Here, synthesised GNPs were coated with polyethylene glycol. Then these particles were conjugated with paclitaxel under different conditions and the physical and structural characteristics, as well as the factors affecting the loading of paclitaxel on nanoparticles, were evaluated by ultraviolet spectrophotometer, fourier transform infrared spectroscopy, transmission electron microscopy, dynamic light scattering and zeta potential apparatus. It was found that pegylated GNPs have a limited loading capacity at the time of 24 h of incubation and the Paclitaxel loading was observed to be pH dependent. The use of these particles in the treatment of breast cancer (MCF7) was also investigated using the MTT test. It was determined that the survival percentage of MCF7 cells in the presence of paclitaxel-bound nanoparticles decreases to about 55% at the maximum measured concentration (690 μM).

本研究的目的是设计和合成与紫杉醇偶联的金纳米颗粒(GNPs),并研究影响合成纳米颗粒稳定性和给药能力的参数。在这里,合成的GNPs被聚乙二醇包裹。利用紫外分光光度计、傅里叶变换红外光谱、透射电镜、动态光散射和zeta电位仪等研究了纳米粒子与紫杉醇在不同条件下的共轭,考察了纳米粒子的物理结构特性以及影响紫杉醇负载的因素。结果发现,聚乙二醇化GNPs在孵育24 h时具有有限的负载能力,并且观察到紫杉醇负载与pH有关。这些颗粒在治疗乳腺癌(MCF7)中的应用也通过MTT试验进行了研究。结果表明,在最大浓度(690 μM)下,紫杉醇结合纳米颗粒对MCF7细胞的存活率降低至55%左右。
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引用次数: 0
Anionic liposomes prepared without organic solvents for effective siRNA delivery 无有机溶剂制备的阴离子脂质体用于siRNA的有效递送
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-02-14 DOI: 10.1049/nbt2.12117
Xiu Han, Yan Lu, Zhaoluo Xu, Yanan Chu, Xueping Ma, Haiping Wu, Bingjie Zou, Guohua Zhou
Abstract Currently, organic solvents are necessary for the preparation of anionic liposomes for siRNA delivery. The removal of organic solvent is time‐consuming and the residual organic solvent is not only a hidden danger, but also affects the stability of anionic liposomes. Glycerol, which is physiologically compatible and does not need to be removed, is used to promote the dispersion of lipids and the formation of anionic liposomes. Additionally, the preparation process is simple and not time‐consuming. The results showed that anionic liposomes, which were typically spherical with a particle size of 188.9 nm were successfully prepared with glycerol. And with the help of Ca2+, siRNA was encapsulated in anionic liposomes. The highest encapsulation efficiency at 2.4 mM Ca2+ reached 91%. And the formation of calcium phosphate could promote the endosomal escape of siRNA effectively. The results from cell viability showed that the anionic liposomes had no obvious cytotoxicity. It was also verified that anionic liposomes could improve the resistance of siRNA against degradation. Additionally, siRNA delivered by anionic liposomes could play an effective role in knockout. Therefore, anionic liposomes prepared with glycerol will be a safe and effective delivery platform for siRNA and even other nucleic acid drugs.
目前,有机溶剂是制备用于siRNA递送的阴离子脂质体所必需的。有机溶剂的去除耗时长,有机溶剂的残留不仅是一个隐患,而且还会影响阴离子脂质体的稳定性。甘油是生理相容的,不需要去除,用来促进脂质分散和阴离子脂质体的形成。此外,制备过程简单,不耗时。结果表明,用甘油制备的阴离子脂质体具有典型的球形结构,粒径为188.9 nm。在Ca2+的帮助下,siRNA被包裹在阴离子脂质体中。2.4 mM Ca2+的包封效率最高,达到91%。磷酸钙的形成能有效促进siRNA的内体逃逸。细胞活力测定结果表明,阴离子脂质体无明显的细胞毒性。研究还证实阴离子脂质体可以提高siRNA的抗降解能力。此外,阴离子脂质体传递的siRNA可能在敲除中发挥有效作用。因此,用甘油制备的阴离子脂质体将成为siRNA甚至其他核酸药物安全有效的递送平台。
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引用次数: 2
Chitosan nanoparticle loaded by epidermal growth factor as a potential protein carrier for wound healing: In vitro and in vivo studies 表皮生长因子负载壳聚糖纳米颗粒作为伤口愈合的潜在蛋白质载体:体外和体内研究
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-02-03 DOI: 10.1049/nbt2.12116
Samaneh Montazeri, Ali Rastegari, Zohreh Mohammadi, Mahboobeh Nazari, Maryam Yousefi, Fatemeh Yazdi Samadi, Somayeh Najafzadeh, Mehdi Aghsami

Epidermal growth factor (EGF) can be efficiently used in wound healing process; but the main obstacle of its clinical use is its susceptibility to proteolysis and maintaining its effective concentration in the site of action. In this study, chitosan nanoparticles containing EGF is formulated using a simple method to increase its stability in physiological pH as well as protect its biological activity and effectiveness in wound healing process. Nanoparticles with different ratios of chitosan/EGF were prepared and evaluated in vitro and in vivo. Obtained results showed nanoparticles with 2:1 ratio of chitosan/EGF were able to release 80% of encapsulated protein after 12 h. Cell proliferation study demonstrated that prepared nanoparticles could protect EGF functionality in physiological pH. In vivo results showed that nanoparticles with 2:1 ratio of chitosan/EGF could significantly accelerate the wound closure-rate, re-epithelialisation and collagen deposition. In conclusion, the designed nanoparticles in optimal ratio can be considered as a potential vehicle for EGF delivery to wounds with the aim of improving healing process.

表皮生长因子(EGF)在创面愈合过程中发挥着重要作用;但其临床应用的主要障碍是其对蛋白水解的易感性和在作用部位维持其有效浓度。本研究采用简单的方法制备了含有EGF的壳聚糖纳米颗粒,以提高其在生理pH值中的稳定性,并保护其在伤口愈合过程中的生物活性和有效性。制备了壳聚糖/表皮生长因子不同配比的纳米颗粒,并对其进行了体外和体内评价。结果表明,壳聚糖/EGF比例为2:1的纳米颗粒在12 h后可释放80%的被包被蛋白。细胞增殖研究表明,制备的纳米颗粒在生理ph下可保护EGF的功能。体内实验结果表明,壳聚糖/EGF比例为2:1的纳米颗粒可显著加速伤口愈合率、再上皮化和胶原沉积。综上所述,所设计的最佳比例纳米颗粒可以作为表皮生长因子输送到伤口的潜在载体,以改善伤口愈合过程。
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引用次数: 0
scRNA-seq data analysis method to improve analysis performance scRNA-seq数据分析方法,提高分析性能
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-02-02 DOI: 10.1049/nbt2.12115
Junru Lu, Yuqi Sheng, Weiheng Qian, Min Pan, Xiangwei Zhao, Qinyu Ge

With the development of single-cell RNA sequencing technology (scRNA-seq), we have the ability to study biological questions at the level of the individual cell transcriptome. Nowadays, many analysis tools, specifically suitable for single-cell RNA sequencing data, have been developed. In this review, the currently commonly used scRNA-seq protocols are discussed. The upstream processing flow pipeline of scRNA-seq data, including goals and popular tools for reads mapping and expression quantification, quality control, normalization, imputation, and batch effect removal is also introduced. Finally, methods to evaluate these tools in both cellular and genetic dimensions, clustering and differential expression analysis are presented.

随着单细胞RNA测序技术(scRNA-seq)的发展,我们有能力在单个细胞转录组水平上研究生物学问题。目前,已经开发出许多专门适用于单细胞RNA测序数据的分析工具。本文对目前常用的scRNA-seq协议进行了综述。本文还介绍了scRNA-seq数据的上游处理流程,包括目标和常用的reads映射和表达量化、质量控制、归一化、imputation和批次效应去除的工具。最后,介绍了在细胞和遗传维度、聚类和差异表达分析方面评估这些工具的方法。
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引用次数: 1
Synergistic effects of 3D chitosan-based hybrid scaffolds and mesenchymal stem cells in orthopaedic tissue engineering 三维壳聚糖复合支架与间充质干细胞在骨科组织工程中的协同作用
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-01-28 DOI: 10.1049/nbt2.12103
Ping Qi, Zhaohui Ning, Xiuju Zhang

Restoration of damaged bone and cartilage tissue with biomaterial scaffolds is an area of interest in orthopaedics. Chitosan is among the low-cost biomaterials used as scaffolds with considerable biocompability to almost every human tissue. Considerable osteoconductivity, porosity, and appropriate pore size distribution have made chitosan an appropriate scaffold for loading of stem cells and a good homing place for differentiation of stem cells to bone tissue. Moreover, the similarity of chitosan to glycosaminoglycans and its potential to be used as soft gels, which could be lasting more than 1 week in mobile chondral defects, has made chitosan a polymer of interest in repairing bone and cartilage defects. Different types of scaffolds using chitosan in combination with mesenchymal stem cells (MSCs) are discussed. MSCs are widely used in regenerative medicine because of their regenerative ability, and recent line evidence reviewed demonstrated that the combination of MSCs with a combination of chitosan with different materials, including collagen type 1, hyaluronic acid, Poly(L-lacticacid)/gelatin/β-tricalcium phosphate, gamma-poly[glutamic acid] polyelectrolyte/titanium alloy, modified Poly(L-Lactide-co-Epsilon-Caprolactone), calcium phosphate, β-glycerophosphate hydrogel/calcium phosphate cement (CPC), and CPC-Chitosan-RGD, can increase the efficacy of using MSCs, and chitosan-based stem cell delivery can be a promising method in restoration of damaged bone and cartilage tissue.

生物材料支架修复受损骨和软骨组织是骨科研究的热点。壳聚糖是一种低成本的生物支架材料,对几乎所有人体组织都具有相当的生物相容性。壳聚糖具有良好的骨导电性、孔隙率和合适的孔径分布,是干细胞装载的合适支架,也是干细胞向骨组织分化的良好载体。此外,壳聚糖与糖胺聚糖的相似性及其作为软凝胶的潜力,可在移动软骨缺损中持续1周以上,使壳聚糖成为修复骨和软骨缺损的聚合物。讨论了壳聚糖与间充质干细胞(MSCs)复合制备不同类型的支架。MSCs因其具有再生能力而被广泛应用于再生医学,最近的证据表明,MSCs与壳聚糖与不同材料的组合,包括1型胶原蛋白、透明质酸、聚乳酸/明胶/β-磷酸三钙、γ -聚谷氨酸聚电解质/钛合金、改性聚乳酸-co- epsilon -己内酯、磷酸钙、β-甘油磷酸水凝胶/磷酸钙水泥(CPC)、和cpc -壳聚糖- rgd可以提高MSCs的使用效果,壳聚糖干细胞递送是修复受损骨和软骨组织的一种很有前景的方法。
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引用次数: 3
Fabrication of nano-encapsulated angelica (Heracleum persicum) essential oil for enriching dairy dessert: Physicochemical, rheological and sensorial properties 乳品甜点用纳米胶囊化当归精油的制备:理化、流变和感官特性
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-01-27 DOI: 10.1049/nbt2.12112
Narin Mhemmedamin Nanakali

In this study, the nanoemulsions containing angelica essential oil (AEO) was used as a novel nano-carrier for enrichment of dairy dessert. Firstly, oil-in-water nanoemulsions were prepared by different levels of GE (1%, 5%, 10%, and 15%) as the dispersed phase, Tween 80 as surfactant with a constant surfactant to essential oil ratio (1:1), and distillated water as a continuous phase. Droplet size, free radical scavenging capacity, antimicrobial activity against gram-positive (Staphylococcus aureus (25923 ATCC)) and gram-negative (Escherichia coli H7 O157 (700728 ATCC)) were evaluated for produced nanoemulsions. The mean droplet size of nanoemulsion increased from 75 to 95 nm and antioxidant capacity also enhanced from 15.4% to 30.2% by increasing AEO level from 1% to 15%. Antimicrobial analysis by disk diffusion methods for nanoemulsions containing different levels of AEO cleared that nanoemulsions with high levels of AEO showed the stronger antimicrobial activity against both used bacteria and especially more activity against Staphylococcus aureus. The results of the total count and yeast and mould count show that the nanoemulsions with different levels of AEO have been effective on the number of microorganisms, particularly during storage. The incorporation of pure essential oil and nanoemulsions with different levels of AEO did not affect significantly the pH of different dessert samples however, they affected the dry matter and free radical scavenging capacity. Adding of nanoemulsions with different levels of AEO to the desserts had a considerable effect on the rheological properties including apparent viscosity, Gʹ, G", Tan δ and complex viscosity and all samples showed shear-thining behaviour. Results from organoleptic characteristics (taste, odour colour, mouthfeel and total acceptance) showed that enriched samples by nanoemulsions, particularly with higher level of AEO had higher sensorial scores. In general, samples containing free AEO (not encapsulated) had the lower scores in all organoleptic characteristics.

本研究以当归精油纳米乳为载体,对乳制品甜点进行浓缩。首先,以不同浓度的GE(1%、5%、10%、15%)为分散相,Tween 80为表面活性剂,表面活性剂与精油的比例恒定(1:1),蒸馏水为连续相制备水包油纳米乳。对制备的纳米乳液进行液滴大小、自由基清除能力、对革兰氏阳性(金黄色葡萄球菌(25923 ATCC))和革兰氏阴性(大肠杆菌H7 O157 (700728 ATCC))的抑菌活性评价。当AEO含量从1%增加到15%时,纳米乳液的平均粒径从75 nm增加到95 nm,抗氧化能力从15.4%提高到30.2%。采用圆盘扩散法对不同AEO含量的纳米乳进行抗菌分析,结果表明,AEO含量高的纳米乳对所用细菌均有较强的抗菌活性,尤其是对金黄色葡萄球菌的抗菌活性更强。结果表明,不同AEO含量的纳米乳对微生物数量有显著的影响,尤其是在贮藏过程中。不同AEO含量的纯精油和纳米乳液的掺入对不同甜点样品的pH值没有显著影响,但对干物质和自由基清除能力有影响。加入不同AEO水平的纳米乳对甜点的表观粘度、G′、G′、Tan δ和复合粘度等流变性能均有显著影响,且所有样品均表现出剪切变薄行为。感官特征(味觉、气味、颜色、口感和总接受度)的结果表明,纳米乳液富集的样品,特别是具有较高水平的AEO的样品具有较高的感官得分。一般来说,含有游离AEO(未封装)的样品在所有感官特征中得分较低。
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引用次数: 2
The inhibition of ORMDL3 prevents Alzheimer's disease through ferroptosis by PERK/ATF4/HSPA5 pathway ORMDL3通过PERK/ATF4/HSPA5途径抑制铁下垂预防阿尔茨海默病
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-01-20 DOI: 10.1049/nbt2.12113
Yankun Shao, Yilin Xu, Huang Di, Xinxiu Shi, Yingying Wang, Hongyu Liu, Lina Song

Alzheimer's disease (AD) is a neurodegenerative disease with high incidence and widespread attention. There is currently no clear clarification of the pathogenesis. However, ORMDL3 causes ferroptosis in AD, and the potential mechanisms remain unclear. So, this study explore the function of ORMDL3 on ferroptosis in AD and its potential regulatory mechanisms. APPswe/PS1dE9 mice and C57BL/6 mice were induced into the mice model. The murine microglial BV-2 cells also were induced into the vitro model. In serum samples of AD patients, ORMDL3 mRNA expression levels were upregulated. The serum ORMDL3 levels expression was positively related to the ADL score or MoCA score in AD patients. The serum ORMDL3 expression level was positively related to MMSE score or Hcy levels in AD patients. The mRNA expression of ORMDL3 in the hippocampal tissue of the mice model of AD was upregulated at one, four and eight months. The protein expression of ORMDL3 was upregulated in the mice model of AD. ORMDL3 promoted Alzheimer's disease, and increased oxidative response and ferroptosis in a model of AD. PERK/ATF4/HSPA5 pathway is one important signal pathway for the effects of ORMDL3 in a model of AD. Collectively, these data suggested that ORMDL3 promoted oxidative response and ferroptosis in a model of AD by the PERK/ATF4/HSPA5 pathway, which might be a novel target spot mechanism of ferroptosis in AD and may serve as a regulator of AD-induced ferroptosis.

阿尔茨海默病(Alzheimer's disease, AD)是一种发病率高、受到广泛关注的神经退行性疾病。目前还没有明确的发病机制。然而,ORMDL3导致AD中的铁下垂,其潜在机制尚不清楚。因此,本研究探讨ORMDL3在AD患者铁下垂中的作用及其可能的调控机制。将APPswe/PS1dE9小鼠和C57BL/6小鼠分别诱导成小鼠模型。小鼠小胶质细胞BV-2也被诱导成体外模型。在AD患者的血清样本中,ORMDL3 mRNA表达水平上调。AD患者血清ORMDL3水平表达与ADL评分或MoCA评分呈正相关。AD患者血清ORMDL3表达水平与MMSE评分或Hcy水平呈正相关。在1个月、4个月和8个月时,AD模型小鼠海马组织中ORMDL3 mRNA表达上调。ORMDL3蛋白在AD小鼠模型中表达上调。ORMDL3促进阿尔茨海默病,并增加AD模型中的氧化反应和铁吊。PERK/ATF4/HSPA5通路是ORMDL3在AD模型中发挥作用的重要信号通路。综上所述,这些数据表明ORMDL3通过PERK/ATF4/HSPA5途径促进AD模型中的氧化反应和铁下垂,这可能是AD中铁下垂的一种新的靶点机制,可能是AD诱导的铁下垂的调节剂。
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引用次数: 2
Optically driven microtools with an antibody-immobilised surface for on-site cell assembly 带有抗体固定表面的光学驱动微工具,用于现场细胞组装
IF 2.3 4区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-01-16 DOI: 10.1049/nbt2.12114
Shuntaro Mori, Takumi Ito, Hidekuni Takao, Fusao Shimokawa, Kyohei Terao

To enable the accurate reproduction of organs in vitro, and improve drug screening efficiency and regenerative medicine research, it is necessary to assemble cells with single-cell resolution to form cell clusters. However, a method to assemble such forms has not been developed. In this study, a platform for on-site cell assembly at the single-cell level using optically driven microtools in a microfluidic device is developed. The microtool was fabricated by SU-8 photolithography, and antibodies were immobilised on its surface. The cells were captured by the microtool through the bindings between the antibodies on the microtool and the antigens on the cell membrane. Transmembrane proteins, CD51/61 and CD44 that facilitate cell adhesion, commonly found on the surface of cancer cells were targeted. The microtool containing antibodies for CD51/61 and CD44 proteins was manipulated using optical tweezers to capture HeLa cells placed on a microfluidic device. A comparison of the adhesion rates of different surface treatments showed the superiority of the antibody-immobilised microtool. The assembly of multiple cells into a cluster by repeating the cell capture process is further demonstrated. The geometry and surface function of the microtool can be modified according to the cell assembly requirements. The platform can be used in regenerative medicine and drug screening to produce cell clusters that closely resemble tissues and organs in vivo.

为了实现器官在体外的精确繁殖,提高药物筛选效率和再生医学研究,需要将具有单细胞分辨率的细胞进行组装,形成细胞簇。然而,组装这种形式的方法还没有开发出来。在本研究中,开发了一种在微流体装置中使用光学驱动微工具进行单细胞水平现场细胞组装的平台。利用SU-8光刻技术制备微工具,并将抗体固定在微工具表面。微工具通过微工具上的抗体和细胞膜上的抗原之间的结合来捕获细胞。目标是癌细胞表面常见的促进细胞粘附的跨膜蛋白CD51/61和CD44。使用光学镊子对含有CD51/61和CD44蛋白抗体的微工具进行操作,以捕获放置在微流体装置上的HeLa细胞。通过对不同表面处理的黏附率的比较,显示了抗体固定化微工具的优越性。通过重复细胞捕获过程,进一步演示了将多个细胞组装成集群。微工具的几何形状和表面功能可以根据单元装配要求进行修改。该平台可用于再生医学和药物筛选,以产生与体内组织和器官非常相似的细胞团。
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引用次数: 1
期刊
IET nanobiotechnology
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