Pub Date : 2024-09-14DOI: 10.1101/2024.09.10.612264
Lei Yang, Anqi Di, Lishuang Song, Xuefei Liu, Di Wu, Song Wang, Zhengting Hao, Lige Bu, Chunling Bai, Guanghua Su, Zhuying Wei, Li Zhang, Zhonghua Liu, Shaorong Gao, Guangpeng Li
Haploid androgenetic stem cells (haSCs) are revolutionary resources for studying animal traits and greatly impact livestock breeding, but livestock haSCs have yet to be obtained. Here, we report the derivation of cattle and sheep haSCs using a novel FACE medium, and these cells harbor formative pluripotency features. Notably, knockout cattle and sheep can be efficiently obtained upon injection of the protaminized haSCs with targeted gene mutations into oocytes.
单倍体雄性干细胞（haSCs）是研究动物性状的革命性资源，对家畜育种有重大影响，但家畜haSCs尚未获得。在这里，我们报告了利用新型 FACE 培养基衍生牛羊单倍体雄性干细胞的情况，这些细胞具有形成性多能特征。值得注意的是，将带有目标基因突变的原氨基化造血干细胞注射到卵母细胞中，可以有效地获得基因敲除的牛羊。
{"title":"Rapid production of modified cattle and sheep from spermatid-like stem cells","authors":"Lei Yang, Anqi Di, Lishuang Song, Xuefei Liu, Di Wu, Song Wang, Zhengting Hao, Lige Bu, Chunling Bai, Guanghua Su, Zhuying Wei, Li Zhang, Zhonghua Liu, Shaorong Gao, Guangpeng Li","doi":"10.1101/2024.09.10.612264","DOIUrl":"https://doi.org/10.1101/2024.09.10.612264","url":null,"abstract":"Haploid androgenetic stem cells (haSCs) are revolutionary resources for studying animal traits and greatly impact livestock breeding, but livestock haSCs have yet to be obtained. Here, we report the derivation of cattle and sheep haSCs using a novel FACE medium, and these cells harbor formative pluripotency features. Notably, knockout cattle and sheep can be efficiently obtained upon injection of the protaminized haSCs with targeted gene mutations into oocytes.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"12 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256031","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-14DOI: 10.1101/2024.09.09.611631
Manik Bansal, Bingrui Wang, Susannah Waxman, Fugiang Zhong, Yi Hua, Yuankai Lu, Juan Reynaud, Brad Fortune, Ian A Sigal
Purpose. IOP-induced mechanical insult on retinal ganglion cell axons within the optic nerve head (ONH) is believed to be a key factor in axonal damage and glaucoma. However, most studies focus on tissue-level mechanical deformations, overlooking that axons are long and thin, and that their susceptibility to damage likely depends on the insult's type (e.g. stretch/compression) and orientation (longitudinal/transverse). We propose an axon-centric approach to quantify IOP-induced mechanical insult from an axon perspective. Methods. We used optical coherence tomography (OCT) scans from a healthy monkey eye along with histological images of cryosections to reconstruct the axon-occupied volume including detailed lamina cribrosa (LC) pores. Tissue-level strains were determined experimentally using digital volume correlation from OCT scans at baseline and elevated IOPs, then transformed into axonal strains using axon paths estimated by a fluid mechanics simulation. Results. Axons in the LC and post-LC regions predominantly experienced longitudinal compression and transverse stretch, whereas those in the pre-LC and ONH rim mainly suffered longitudinal stretch and transverse compression. No clear patterns were observed for tissue-level strains. Conclusions. Our approach allowed discerning axonal longitudinal and transverse mechanical insults, which are likely associated with different mechanisms of axonal damage. The technique also enabled quantifying insult along individual axon paths, providing a novel link relating the retinal nerve fiber layer and the optic nerve through the LC via individual axons. This is a promising approach to establish a clearer connection between IOP-induced insult and glaucoma. Further studies should evaluate a larger cohort.
研究目的眼压对视神经头（ONH）内视网膜神经节细胞轴突造成的机械损伤被认为是轴突损伤和青光眼的关键因素。然而，大多数研究都侧重于组织层面的机械变形，忽略了轴突又长又细，而且它们对损伤的敏感性可能取决于损伤的类型（如拉伸/压缩）和方向（纵向/横向）。我们提出了一种以轴突为中心的方法，从轴突的角度量化眼压诱导的机械损伤。方法。我们利用健康猴眼的光学相干断层扫描（OCT）扫描结果和冷冻切片的组织学图像来重建轴突占据的体积，包括详细的板层孔（LC）。组织级应变是通过基线和眼压升高时的 OCT 扫描数字体积相关性实验确定的，然后通过流体力学模拟估算的轴突路径转换为轴突应变。结果LC和LC后区域的轴突主要受到纵向压缩和横向拉伸，而LC前和ONH边缘的轴突主要受到纵向拉伸和横向压缩。组织级应变没有观察到明显的模式。结论我们的方法可以分辨轴突的纵向和横向机械损伤，这可能与轴突损伤的不同机制有关。该技术还能量化单个轴突路径上的损伤，通过单个轴突提供了视网膜神经纤维层和视神经之间的新联系。这是一种很有前途的方法，可以在眼压引起的损伤和青光眼之间建立更清晰的联系。进一步的研究应该对更大的群体进行评估。
{"title":"Proposing a methodology for axon-centric analysis of IOP-induced mechanical insult","authors":"Manik Bansal, Bingrui Wang, Susannah Waxman, Fugiang Zhong, Yi Hua, Yuankai Lu, Juan Reynaud, Brad Fortune, Ian A Sigal","doi":"10.1101/2024.09.09.611631","DOIUrl":"https://doi.org/10.1101/2024.09.09.611631","url":null,"abstract":"Purpose. IOP-induced mechanical insult on retinal ganglion cell axons within the optic nerve head (ONH) is believed to be a key factor in axonal damage and glaucoma. However, most studies focus on tissue-level mechanical deformations, overlooking that axons are long and thin, and that their susceptibility to damage likely depends on the insult's type (e.g. stretch/compression) and orientation (longitudinal/transverse). We propose an axon-centric approach to quantify IOP-induced mechanical insult from an axon perspective. Methods. We used optical coherence tomography (OCT) scans from a healthy monkey eye along with histological images of cryosections to reconstruct the axon-occupied volume including detailed lamina cribrosa (LC) pores. Tissue-level strains were determined experimentally using digital volume correlation from OCT scans at baseline and elevated IOPs, then transformed into axonal strains using axon paths estimated by a fluid mechanics simulation. Results. Axons in the LC and post-LC regions predominantly experienced longitudinal compression and transverse stretch, whereas those in the pre-LC and ONH rim mainly suffered longitudinal stretch and transverse compression. No clear patterns were observed for tissue-level strains. Conclusions. Our approach allowed discerning axonal longitudinal and transverse mechanical insults, which are likely associated with different mechanisms of axonal damage. The technique also enabled quantifying insult along individual axon paths, providing a novel link relating the retinal nerve fiber layer and the optic nerve through the LC via individual axons. This is a promising approach to establish a clearer connection between IOP-induced insult and glaucoma. Further studies should evaluate a larger cohort.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"202 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256035","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Biomimetic cues from the extracellular matrix (ECM) are essential for optimizing cell microenvironments and biomaterials. While native ECM proteins or synthetic peptides offer potential solutions, challenges such as production cost, solubility, and conformational stability limit their use. Here, we present the development of virus-like particles (VLPs) derived from the AP205 RNA phage displaying peptides from key ECM proteins and evaluate their biological activity in a variety of assays. We show that our engineered VLPs can effectively stimulate cell adhesion, migration, proliferation and differentiation. By comparing focal adhesions formed by RGD VLPs with their parent protein, fibronectin, we elucidate both similarities and differences in cell interactions. In addition, we construct heterodimeric particles co-expressing RGD with differentiation peptides and demonstrate retention of bioactivity in a multi-peptide context. This study establishes AP205 VLPs as versatile nanoscale platforms capable of tuning cell functions, with promising applications in nanomedicine and biomaterials.
{"title":"Biomimetic Virus-Like Particles to control cell functions","authors":"Hasna Maayouf, Thomas Dos Santos, Alphonse Boche, Rayane Hedna, Kaspars Tars, Isabelle Brigaud, Tatiana Petithory, Franck Carreiras, Carole Arnold, Ambroise Lambert, Laurent Pieuchot","doi":"10.1101/2024.09.14.612851","DOIUrl":"https://doi.org/10.1101/2024.09.14.612851","url":null,"abstract":"Biomimetic cues from the extracellular matrix (ECM) are essential for optimizing cell microenvironments and biomaterials. While native ECM proteins or synthetic peptides offer potential solutions, challenges such as production cost, solubility, and conformational stability limit their use. Here, we present the development of virus-like particles (VLPs) derived from the AP205 RNA phage displaying peptides from key ECM proteins and evaluate their biological activity in a variety of assays. We show that our engineered VLPs can effectively stimulate cell adhesion, migration, proliferation and differentiation. By comparing focal adhesions formed by RGD VLPs with their parent protein, fibronectin, we elucidate both similarities and differences in cell interactions. In addition, we construct heterodimeric particles co-expressing RGD with differentiation peptides and demonstrate retention of bioactivity in a multi-peptide context. This study establishes AP205 VLPs as versatile nanoscale platforms capable of tuning cell functions, with promising applications in nanomedicine and biomaterials.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"22 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142255987","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-14DOI: 10.1101/2024.09.10.612317
Ushasi Pramanik, Anirban Das, Elise M. Brown, Heather L. Struckman, Huihao Wang, Samuel Stealey, Macy L. Sprunger, Wasim Abdul, Jonathan Fascetti, Jagannath Mondal, Jonathan R. Silva, Silviya P. Zustiak, Meredith E. Jackrel, Jai S. Rudra
Peptides and peptidomimetics that self-assemble via LLPS have recently emerged as building blocks for fabricating functional biomaterials due to their unique physicochemical properties and dynamic nature. One of life's most distinctive signatures is its selectivity for chiral molecules and, to date, coacervates comprised of D-amino acids have not been reported. Here, we demonstrate that histidine-rich repeats of (GHGXY)4 (X=L/V/P) and their enantiomers undergo LLPS opening new avenues for enhancing coacervate stability. Through a series of biophysical studies, we find that LLPS kinetics, droplet size, fusion, and encapsulation efficiency are dictated by the primary sequence. Further, these coacervates can encapsulate therapeutic cargo which are then internalized via endocytic mechanisms. Finally, we show that the coacervates enhance antigen presentation to CD4+ and CD8+ T cells resulting in robust proliferation and production of functional cytokines. Collectively, our study describes the development and characterization of enantiomeric peptide coacervates as attractive vaccine delivery vehicles with tunable physicochemical properties.
{"title":"Enantiomeric histidine-rich peptide coacervates enhance antigen delivery to T cells","authors":"Ushasi Pramanik, Anirban Das, Elise M. Brown, Heather L. Struckman, Huihao Wang, Samuel Stealey, Macy L. Sprunger, Wasim Abdul, Jonathan Fascetti, Jagannath Mondal, Jonathan R. Silva, Silviya P. Zustiak, Meredith E. Jackrel, Jai S. Rudra","doi":"10.1101/2024.09.10.612317","DOIUrl":"https://doi.org/10.1101/2024.09.10.612317","url":null,"abstract":"Peptides and peptidomimetics that self-assemble via LLPS have recently emerged as building blocks for fabricating functional biomaterials due to their unique physicochemical properties and dynamic nature. One of life's most distinctive signatures is its selectivity for chiral molecules and, to date, coacervates comprised of D-amino acids have not been reported. Here, we demonstrate that histidine-rich repeats of (GHGXY)4 (X=L/V/P) and their enantiomers undergo LLPS opening new avenues for enhancing coacervate stability. Through a series of biophysical studies, we find that LLPS kinetics, droplet size, fusion, and encapsulation efficiency are dictated by the primary sequence. Further, these coacervates can encapsulate therapeutic cargo which are then internalized via endocytic mechanisms. Finally, we show that the coacervates enhance antigen presentation to CD4+ and CD8+ T cells resulting in robust proliferation and production of functional cytokines. Collectively, our study describes the development and characterization of enantiomeric peptide coacervates as attractive vaccine delivery vehicles with tunable physicochemical properties.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"16 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256033","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-14DOI: 10.1101/2024.09.10.612339
Biswajoy Ghosh, Krishna Agarwal
Cell migration is a fundamental biological process, yet the mechanisms underlying how cells sense and navigate complex environments remain poorly understood. In this study, we developed a system of randomly oriented microgrooves, designed at cellular length scales, to explore motility intelligence in response to varied topographies. These microgrooves allowed cells to freely choose their migratory paths, revealing key insights into how cells sense and adapt to topological cues. Using fibroblast cells migrating over these grooved substrates, we examined cellular processes such as actin cytoskeleton remodeling, cell adhesion dynamics, and the impact of groove alignment on migration speed and directionality. Our results demonstrate that cells align their cytoskeletal structures to groove geometries, forming actin-rich anchors that enhance migration in groove-aligned environments. Cells migrating in grooves aligned with their intrinsic polarity exhibited faster, more directed migration compared to those in misaligned or control conditions. This work advances our understanding of cell-topology interaction and provides new perspectives for tissue engineering applications in cancer therapy and wound healing.
{"title":"Randomly oriented microgrooved hydrogel guides cellular motility, modulates speed, and governs directionality of cellular spread","authors":"Biswajoy Ghosh, Krishna Agarwal","doi":"10.1101/2024.09.10.612339","DOIUrl":"https://doi.org/10.1101/2024.09.10.612339","url":null,"abstract":"Cell migration is a fundamental biological process, yet the mechanisms underlying how cells sense and navigate complex environments remain poorly understood. In this study, we developed a system of randomly oriented microgrooves, designed at cellular length scales, to explore motility intelligence in response to varied topographies. These microgrooves allowed cells to freely choose their migratory paths, revealing key insights into how cells sense and adapt to topological cues. Using fibroblast cells migrating over these grooved substrates, we examined cellular processes such as actin cytoskeleton remodeling, cell adhesion dynamics, and the impact of groove alignment on migration speed and directionality. Our results demonstrate that cells align their cytoskeletal structures to groove geometries, forming actin-rich anchors that enhance migration in groove-aligned environments. Cells migrating in grooves aligned with their intrinsic polarity exhibited faster, more directed migration compared to those in misaligned or control conditions. This work advances our understanding of cell-topology interaction and provides new perspectives for tissue engineering applications in cancer therapy and wound healing.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256032","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The study of the protein corona, the immediate and evolving biomolecular coating that forms on the surface of nanoparticles when exposed to a biological environment, is a crucial area in nanomedicine. This phenomenon significantly influences the behavior, functionality, and biological interactions of nanoparticles with biosystems. Until now, conclusions regarding the role of the protein corona in specific biological applications have been based on establishing correlation rather than causation. By understanding causality, researchers can predict how changes in nanoparticle properties or biological conditions will affect protein corona composition, in turn affecting the nanoparticle interactions with the biosystems and their applications. This predictive capability is essential for designing nanoparticles with specific characteristics tailored for therapeutic and diagnostic nanomedicine applications. Here, we explore the concept of actual causality (by Halpern and Pearl) to mathematically prove how various small molecules, including metabolites, lipids, vitamins, and nutrients, spiked into plasma can induce diverse protein corona patterns on identical nanoparticles. This approach significantly enhances the depth of plasma proteome profiling. Our findings reveal that among the various spiked small molecules, phosphatidylcholine was the actual cause of the observed increase in the proteomic depth of the plasma sample. By considering the concept of causality in the field of protein corona, the nanomedicine community can substantially improve their ability to design safer and more efficient nanoparticles for both diagnostic and therapeutic purposes.
蛋白质电晕是纳米粒子暴露于生物环境时在其表面形成的直接和不断演变的生物分子涂层，对蛋白质电晕的研究是纳米医学的一个重要领域。这一现象极大地影响了纳米粒子的行为、功能以及与生物系统的生物相互作用。到目前为止，有关蛋白质电晕在特定生物应用中的作用的结论都是基于建立相关性而非因果关系。通过了解因果关系，研究人员可以预测纳米粒子特性或生物条件的变化将如何影响蛋白质电晕的组成，进而影响纳米粒子与生物系统的相互作用及其应用。这种预测能力对于设计具有特定特性的纳米粒子以用于治疗和诊断纳米医学应用至关重要。在这里，我们探索了实际因果关系的概念（由 Halpern 和 Pearl 提出），以数学方法证明了各种小分子（包括代谢物、脂类、维生素和营养物质）如何通过添加到血浆中诱导相同纳米粒子上的不同蛋白质电晕模式。这种方法大大提高了血浆蛋白质组分析的深度。我们的研究结果表明，在各种添加的小分子中，磷脂酰胆碱是导致血浆样本蛋白质组深度增加的真正原因。通过考虑蛋白质电晕领域的因果关系概念，纳米医学界可以大大提高设计更安全、更高效的纳米粒子的能力，从而达到诊断和治疗的目的。
{"title":"Causality Analysis of Protein Corona Composition: Phosphatidylcholine-Enhances Plasma Proteome Profiling by Proteomics","authors":"Arshia Rafieioskouei, Kenneth Rogale, Amirata Saei Dibavar, Morteza Mahmoudi, Borzoo Bonakdarpour","doi":"10.1101/2024.09.10.612356","DOIUrl":"https://doi.org/10.1101/2024.09.10.612356","url":null,"abstract":"The study of the protein corona, the immediate and evolving biomolecular coating that forms on the surface of nanoparticles when exposed to a biological environment, is a crucial area in nanomedicine. This phenomenon significantly influences the behavior, functionality, and biological interactions of nanoparticles with biosystems. Until now, conclusions regarding the role of the protein corona in specific biological applications have been based on establishing correlation rather than causation. By understanding causality, researchers can predict how changes in nanoparticle properties or biological conditions will affect protein corona composition, in turn affecting the nanoparticle interactions with the biosystems and their applications. This predictive capability is essential for designing nanoparticles with specific characteristics tailored for therapeutic and diagnostic nanomedicine applications. Here, we explore the concept of actual causality (by Halpern and Pearl) to mathematically prove how various small molecules, including metabolites, lipids, vitamins, and nutrients, spiked into plasma can induce diverse protein corona patterns on identical nanoparticles. This approach significantly enhances the depth of plasma proteome profiling. Our findings reveal that among the various spiked small molecules, phosphatidylcholine was the actual cause of the observed increase in the proteomic depth of the plasma sample. By considering the concept of causality in the field of protein corona, the nanomedicine community can substantially improve their ability to design safer and more efficient nanoparticles for both diagnostic and therapeutic purposes.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"100 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256030","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-13DOI: 10.1101/2024.09.09.611468
Xinpei Song, Shreya Udani, Mengxing Ouyang, Mehmet Akif Sahin, Dino Di Carlo, Ghulam Destgeer
Microparticle-templated droplets or dropicles have recently gained interest in the fields of diagnostic immunoassays, single-cell analysis, and digital molecular biology. Amphiphilic particles have been shown to spontaneously capture aqueous droplets within their cavities upon mixing with an immiscible oil phase, where each particle templates a single droplet. Here, we fabricated an amphiphilic microparticle with four discrete hydrophilic patches embedded at the inner corners of a square-shaped hydrophobic outer ring of the particle (4C particle). 3D computational fluid dynamics simulations predicted droplet formation dynamics and differing equilibrium conditions depending on the patterning configuration. Experiments recapitulated equilibrium conditions enabling tunable dropicle configurations with reproducible volumes down to ~200pL templated by the amphiphilic particles. The dropicle configurations depended predominantly on the size of the hydrophilic patches of the 4C particles. This validated modeling approach can inform the design of dropicles with varying volumes and numbers per particle, which can be harnessed in new amplified bioassays for greater sensitivity, dynamic range, and statistical confidence.
{"title":"Tunable picoliter-scale dropicle formation using amphiphilic microparticles with patterned hydrophilic patches","authors":"Xinpei Song, Shreya Udani, Mengxing Ouyang, Mehmet Akif Sahin, Dino Di Carlo, Ghulam Destgeer","doi":"10.1101/2024.09.09.611468","DOIUrl":"https://doi.org/10.1101/2024.09.09.611468","url":null,"abstract":"Microparticle-templated droplets or dropicles have recently gained interest in the fields of diagnostic immunoassays, single-cell analysis, and digital molecular biology. Amphiphilic particles have been shown to spontaneously capture aqueous droplets within their cavities upon mixing with an immiscible oil phase, where each particle templates a single droplet. Here, we fabricated an amphiphilic microparticle with four discrete hydrophilic patches embedded at the inner corners of a square-shaped hydrophobic outer ring of the particle (4C particle). 3D computational fluid dynamics simulations predicted droplet formation dynamics and differing equilibrium conditions depending on the patterning configuration. Experiments recapitulated equilibrium conditions enabling tunable dropicle configurations with reproducible volumes down to ~200pL templated by the amphiphilic particles. The dropicle configurations depended predominantly on the size of the hydrophilic patches of the 4C particles. This validated modeling approach can inform the design of dropicles with varying volumes and numbers per particle, which can be harnessed in new amplified bioassays for greater sensitivity, dynamic range, and statistical confidence.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"52 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256040","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-13DOI: 10.1101/2024.09.09.612104
Carolyn K Jons, Alexander N Prossnitz, Noah Eckman, Changxin Dong, Eric Appel
Protein therapeutics, like peptides and antibodies, have become critical to healthcare. Despite their exceptional potency and specificity, biopharmaceuticals are prone to aggregation, often necessitating low formulation concentrations as well as cold storage and distribution to maintain stability. Yet, high doses are required to treat many diseases. To achieve these doses, most approved protein drug products are administered intravenously, imposing excessive burdens on patients and the healthcare system. New approaches are needed to formulate proteins at high concentrations to enable less burdensome subcutaneous injection, preferably in an autoinjector format. To address this challenge, we report a subcutaneously injectable biotherapeutic delivery platform composed of spray-dried protein microparticles suspended in a non-solvent liquid carrier. These microparticles contain only active biopharmaceutical agent and a high glass transition temperature polyacrylamide-derived copolymer excipient affording several key benefits over traditional excipients, including: (i) improved stabilization of biopharmaceuticals through the spray drying process, and (ii) improved morphology and properties of the spray-dried particles, enhancing suspension injectability. Experiments with albumin and human IgG demonstrate that this technology enables ultra-high-concentration protein formulations (exceeding 500 mg/mL) that are injectable through standard needles with clinically relevant injection forces. Additionally, experiments in mice show these ultra-high-concentration formulations reduce required injection volumes without altering pharmacokinetics. This approach could double the number of commercial protein drugs amenable to subcutaneous administration, dramatically reducing burden and improving access to these critical biopharmaceuticals.
{"title":"Glassy Surfactants Enable Ultra-High Concentration Biologic Therapeutics","authors":"Carolyn K Jons, Alexander N Prossnitz, Noah Eckman, Changxin Dong, Eric Appel","doi":"10.1101/2024.09.09.612104","DOIUrl":"https://doi.org/10.1101/2024.09.09.612104","url":null,"abstract":"Protein therapeutics, like peptides and antibodies, have become critical to healthcare. Despite their exceptional potency and specificity, biopharmaceuticals are prone to aggregation, often necessitating low formulation concentrations as well as cold storage and distribution to maintain stability. Yet, high doses are required to treat many diseases. To achieve these doses, most approved protein drug products are administered intravenously, imposing excessive burdens on patients and the healthcare system. New approaches are needed to formulate proteins at high concentrations to enable less burdensome subcutaneous injection, preferably in an autoinjector format. To address this challenge, we report a subcutaneously injectable biotherapeutic delivery platform composed of spray-dried protein microparticles suspended in a non-solvent liquid carrier. These microparticles contain only active biopharmaceutical agent and a high glass transition temperature polyacrylamide-derived copolymer excipient affording several key benefits over traditional excipients, including: (i) improved stabilization of biopharmaceuticals through the spray drying process, and (ii) improved morphology and properties of the spray-dried particles, enhancing suspension injectability. Experiments with albumin and human IgG demonstrate that this technology enables ultra-high-concentration protein formulations (exceeding 500 mg/mL) that are injectable through standard needles with clinically relevant injection forces. Additionally, experiments in mice show these ultra-high-concentration formulations reduce required injection volumes without altering pharmacokinetics. This approach could double the number of commercial protein drugs amenable to subcutaneous administration, dramatically reducing burden and improving access to these critical biopharmaceuticals.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"24 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256039","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-13DOI: 10.1101/2024.09.07.611801
Nadia Boroumand
Purpose: Epidemiological studies show that radon and cigarette smoke interact in inducing lung cancer, but the contribution of nicotine in response to alpha radiation emitted by radon is not well understood. Materials and methods: Bronchial epithelial BEAS 2B cells were either pre-treated with 2 uM nicotine during 16 h, exposed to radiation, or the combination. DNA damage, cellular and chromosomal alterations, oxidative stress as well as inflammatory responses were assessed to investigate the role of nicotine in modulating responses. Results: Less γH2AX foci were detected at 1 h after alpha radiation exposure (1 2 Gy) in the combination group versus alpha radiation alone, whereas nicotine alone had no effect. Comet assay showed less DNA breaks already just after combined exposure, supported by reduced p-ATM, p DNA PK, p p53 and RAD51 at 1 h, compared to alpha radiation alone. Yet the frequency of translocations was higher in the combination group at 27 h after irradiation. Although nicotine did not alter G2 arrest at 24 h, it assisted in cell cycle progression at 48 h post radiation. A slightly faster recovery was indicated in the combination group based on cell viability kinetics and viable cell counts, and significantly using colony formation assay. Pan-histone acetyl transferase inhibition using PU139 blocked the reduction in p p53 and γH2AX activation, suggesting a role for nicotine-induced histone acetylation in enabling rapid DNA repair. Nicotine had a modest effect on reactive oxygen species induction but tended to increase alpha particle induced pro inflammatory IL 6 and IL 1β (4 Gy). Interestingly, nicotine did not alter gamma radiation induced γH2AX foci. Conclusions: This study provides evidence that nicotine modulates alpha radiation response by causing a faster but more error prone repair, as well as rapid recovery, which may allow expansion of cells with genomic instabilities. These results hold implications for estimating radiation risk among nicotine users.
目的：流行病学研究表明，氡和香烟烟雾在诱发肺癌方面具有相互作用，但尼古丁对氡发出的α辐射的影响尚不十分清楚。材料和方法：将支气管上皮 BEAS 2B 细胞用 2 uM 尼古丁预处理 16 小时、暴露于辐射或两者结合。评估DNA损伤、细胞和染色体改变、氧化应激以及炎症反应，以研究尼古丁在调节反应中的作用。结果显示α射线照射（1 2 Gy）1小时后，联合组检测到的γH2AX病灶少于单独照射组，而单独使用尼古丁则没有影响。彗星试验显示，与单独使用α射线相比，联合照射后 1 小时，p-ATM、p DNA PK、p p53 和 RAD51 减少，DNA 断裂减少。然而，在照射后 27 小时，联合组的易位频率更高。虽然尼古丁不会改变辐射后24小时的G2停滞，但却有助于辐射后48小时的细胞周期进展。根据细胞存活动力学和存活细胞计数以及菌落形成检测，联合组的恢复速度稍快。使用 PU139 抑制泛组蛋白乙酰转移酶可以阻止 p p53 和 γH2AX 活化的减少，这表明尼古丁诱导的组蛋白乙酰化在快速修复 DNA 方面发挥了作用。尼古丁对活性氧的诱导作用不大，但往往会增加α粒子诱导的促炎 IL 6 和 IL 1β（4 Gy）。有趣的是，尼古丁并未改变伽马射线诱导的γH2AX病灶：本研究提供的证据表明，尼古丁可通过导致更快但更容易出错的修复以及快速恢复来调节α辐射反应，从而使基因组不稳定的细胞得以扩增。这些结果对估计尼古丁使用者的辐射风险具有一定的意义。
{"title":"Nicotine interacts with DNA lesions induced by alpha radiation which may contribute to erroneous repair in human lung epithelial cells","authors":"Nadia Boroumand","doi":"10.1101/2024.09.07.611801","DOIUrl":"https://doi.org/10.1101/2024.09.07.611801","url":null,"abstract":"Purpose: Epidemiological studies show that radon and cigarette smoke interact in inducing lung cancer, but the contribution of nicotine in response to alpha radiation emitted by radon is not well understood. Materials and methods: Bronchial epithelial BEAS 2B cells were either pre-treated with 2 uM nicotine during 16 h, exposed to radiation, or the combination. DNA damage, cellular and chromosomal alterations, oxidative stress as well as inflammatory responses were assessed to investigate the role of nicotine in modulating responses. Results: Less γH2AX foci were detected at 1 h after alpha radiation exposure (1 2 Gy) in the combination group versus alpha radiation alone, whereas nicotine alone had no effect. Comet assay showed less DNA breaks already just after combined exposure, supported by reduced p-ATM, p DNA PK, p p53 and RAD51 at 1 h, compared to alpha radiation alone. Yet the frequency of translocations was higher in the combination group at 27 h after irradiation. Although nicotine did not alter G2 arrest at 24 h, it assisted in cell cycle progression at 48 h post radiation. A slightly faster recovery was indicated in the combination group based on cell viability kinetics and viable cell counts, and significantly using colony formation assay. Pan-histone acetyl transferase inhibition using PU139 blocked the reduction in p p53 and γH2AX activation, suggesting a role for nicotine-induced histone acetylation in enabling rapid DNA repair. Nicotine had a modest effect on reactive oxygen species induction but tended to increase alpha particle induced pro inflammatory IL 6 and IL 1β (4 Gy). Interestingly, nicotine did not alter gamma radiation induced γH2AX foci.\u0000Conclusions: This study provides evidence that nicotine modulates alpha radiation response by causing a faster but more error prone repair, as well as rapid recovery, which may allow expansion of cells with genomic instabilities. These results hold implications for estimating radiation risk among nicotine users.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"42 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256042","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Purpose: We aim to implement a 3D DWI sequence and show its usage on patients with new ischemic lesions. Materials and Methods: The proposed 3D DWI sequence was implemented by integrating second-order gradient moment nulling (M2) and cardiac motion synchronization (Sync). All data were acquired on a 3T MAGNETOM Prisma scanner (Siemens Healthcare, Erlangen, Germany) using a 64 channel head and neck coil. 21 healthy volunteers underwent 3D DWI scans at 0.9 mm isotropic resolution using four motion compensation methods for comparison: no compensation (M0), M2 only, Sync only and the proposed M2+Sync method. 2D phase variation maps with different motion compensation methods were also acquired for one subject to illustrate the mechanism of the proposed method. A ghost-to-signal ratio (GSR) and blurring index was defined and compared among the four methods with repeated measures ANOVA and Tukey's test. 3D DWI was compared with 2D DWI for ADC quantification. Image quality and ischemic lesion conspicuity were evaluated with 12 patients after endovascular treatment. Results: Whole brain 3D DWI was achieved at 0.9 mm isotropic resolution within 5 minutes using the proposed sequence. M2+Sync achieved the lowest level of GSR and blurring along the slice direction. ADC quantification showed no statistically significant difference between M2+Sync compared to 2D DWI. 3D DWI showed similar image quality, higher lesion conspicuity and counts compared to 2D DWI. Conclusion: Direct 3D DWI can be achieved by the combination of second order gradient moment nulling and cardiac synchronization.
{"title":"Towards genuine three-dimensional diffusion imaging with physiological motion compensation","authors":"Yishi Wang, Dehe Weng, Jieying Zhang, Tianyi Qian, Wenzhang Liu, Kun Zhou, Yanglei Wu, Baogui Zhang, Qing Li, Jing Jing, Zhe Zhang","doi":"10.1101/2024.09.08.611927","DOIUrl":"https://doi.org/10.1101/2024.09.08.611927","url":null,"abstract":"Purpose: We aim to implement a 3D DWI sequence and show its usage on patients with new ischemic lesions. Materials and Methods: The proposed 3D DWI sequence was implemented by integrating second-order gradient moment nulling (M2) and cardiac motion synchronization (Sync). All data were acquired on a 3T MAGNETOM Prisma scanner (Siemens Healthcare, Erlangen, Germany) using a 64 channel head and neck coil. 21 healthy volunteers underwent 3D DWI scans at 0.9 mm isotropic resolution using four motion compensation methods for comparison: no compensation (M0), M2 only, Sync only and the proposed M2+Sync method. 2D phase variation maps with different motion compensation methods were also acquired for one subject to illustrate the mechanism of the proposed method. A ghost-to-signal ratio (GSR) and blurring index was defined and compared among the four methods with repeated measures ANOVA and Tukey's test. 3D DWI was compared with 2D DWI for ADC quantification. Image quality and ischemic lesion conspicuity were evaluated with 12 patients after endovascular treatment. Results: Whole brain 3D DWI was achieved at 0.9 mm isotropic resolution within 5 minutes using the proposed sequence. M2+Sync achieved the lowest level of GSR and blurring along the slice direction. ADC quantification showed no statistically significant difference between M2+Sync compared to 2D DWI. 3D DWI showed similar image quality, higher lesion conspicuity and counts compared to 2D DWI. Conclusion: Direct 3D DWI can be achieved by the combination of second order gradient moment nulling and cardiac synchronization.","PeriodicalId":501308,"journal":{"name":"bioRxiv - Bioengineering","volume":"193 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142256041","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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