scholarly journals In Vitro Evaluation of Lipopolyplexes for Gene Transfection: Comparing 2D, 3D and Microdroplet-Enabled Cell Culture

Molecules ◽  
2020 ◽  
Vol 25 (14) ◽  
pp. 3277
Author(s):  
Juan L. Paris ◽  
Filipe Coelho ◽  
Alexandra Teixeira ◽  
Lorena Diéguez ◽  
Bruno F. B. Silva ◽  
...  
Keyword(s):  
Cell Culture ◽  
Transfection Efficiency ◽  
Gene Transfection ◽  
3D Culture ◽  
In Vitro Models ◽  
Great Promise ◽  
Cationic Polymers ◽  
Significant Gene ◽  
Biological Performance

Complexes combining nucleic acids with lipids and polymers (lipopolyplexes) show great promise for gene therapy since they enable compositional, physical and functional versatility to be optimized for therapeutic efficiency. When developing lipopolyplexes for gene delivery, one of the first evaluations performed is an in vitro transfection efficiency experiment. Many different in vitro models can be used, and the effect of the model on the experiment outcome has not been thoroughly studied. The objective of this work was to compare the insights obtained from three different in vitro models, as well as the potential limitations associated with each of them. We have prepared a series of lipopolyplex formulations with three different cationic polymers (poly-l-lysine, bioreducible poly-l-lysine and polyethyleneimine), and assessed their in vitro biological performance in 2D monolayer cell culture, 3D spheroid culture and microdroplet-based single-cell culture. Lipopolyplexes from different polymers presented varying degrees of transfection efficiency in all models. The best-performing formulation in 2D culture was the polyethyleneimine lipopolyplex, while lipoplexes prepared with bioreducible poly-l-lysine were the only ones achieving any transfection in microdroplet-enabled cell culture. None of the prepared formulations achieved significant gene transfection in 3D culture. All of the prepared formulations were well tolerated by cells in 2D culture, while at least one formulation (poly-l-lysine polyplex) delayed 3D spheroid growth. These results highlight the need for selecting the appropriate in vitro model depending on the intended application.

3D Culture Modelling: An Emerging Approach for Translational Cancer Research in Sarcomas

2020 ◽  
Vol 27 (29) ◽  
pp. 4778-4788 ◽  
Author(s):  
Victoria Heredia-Soto ◽  
Andrés Redondo ◽  
José Juan Pozo Kreilinger ◽  
Virginia Martínez-Marín ◽  
Alberto Berjón ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Cancer Biology ◽  
Soft Tissues ◽  
Three Dimensional ◽  
3D Culture ◽  
Resistance Mechanisms ◽  
In Vitro Models ◽  
Tumour Spheroids ◽  
Current Therapies

Sarcomas are tumours of mesenchymal origin, which can arise in bone or soft tissues. They are rare but frequently quite aggressive and with a poor outcome. New approaches are needed to characterise these tumours and their resistance mechanisms to current therapies, responsible for tumour recurrence and treatment failure. This review is focused on the potential of three-dimensional (3D) in vitro models, including multicellular tumour spheroids (MCTS) and organoids, and the latest data about their utility for the study on important properties for tumour development. The use of spheroids as a particularly valuable alternative for compound high throughput screening (HTS) in different areas of cancer biology is also discussed, which enables the identification of new therapeutic opportunities in commonly resistant tumours.

Non-Viral Vectors for Gene Delivery

2018 ◽  
Vol 9 (1) ◽  
pp. 4-11 ◽  
Author(s):  
Aparna Bansal ◽  
Himanshu
Keyword(s):  
Gene Therapy ◽  
Viral Vectors ◽  
Transfection Efficiency ◽  
Gene Transfection ◽  
Expression System ◽  
Target Cells ◽  
Specific Expression ◽  
Naked Dna

Introduction: Gene therapy has emerged out as a promising therapeutic pave for the treatment of genetic and acquired diseases. Gene transfection into target cells using naked DNA is a simple and safe approach which has been further improved by combining vectors or gene carriers. Both viral and non-viral approaches have achieved a milestone to establish this technique, but non-viral approaches have attained a significant attention because of their favourable properties like less immunotoxicity and biosafety, easy to produce with versatile surface modifications, etc. Literature is rich in evidences which revealed that undoubtedly, non–viral vectors have acquired a unique place in gene therapy but still there are number of challenges which are to be overcome to increase their effectiveness and prove them ideal gene vectors. Conclusion: To date, tissue specific expression, long lasting gene expression system, enhanced gene transfection efficiency has been achieved with improvement in delivery methods using non-viral vectors. This review mainly summarizes the various physical and chemical methods for gene transfer in vitro and in vivo.

scholarly journals The Role of Biomimetic Hypoxia on Cancer Cell Behaviour in 3D Models: A Systematic Review

Cancers ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 1334
Author(s):  
Ye Liu ◽  
Zahra Mohri ◽  
Wissal Alsheikh ◽  
Umber Cheema
Keyword(s):  
Systematic Review ◽  
Cellular Response ◽  
3D Culture ◽  
3D Models ◽  
In Vitro Models ◽  
Research Findings ◽  
Tissue Models

The development of biomimetic, human tissue models is recognized as being an important step for transitioning in vitro research findings to the native in vivo response. Oftentimes, 2D models lack the necessary complexity to truly recapitulate cellular responses. The introduction of physiological features into 3D models informs us of how each component feature alters specific cellular response. We conducted a systematic review of research papers where the focus was the introduction of key biomimetic features into in vitro models of cancer, including 3D culture and hypoxia. We analysed outcomes from these and compiled our findings into distinct groupings to ascertain which biomimetic parameters correlated with specific responses. We found a number of biomimetic features which primed cancer cells to respond in a manner which matched in vivo response.

Retracted Article: A multifunctional self-dissociative polyethyleneimine derivative coating polymer for enhancing the gene transfection efficiency of DNA/polyethyleneimine polyplexes in vitro and in vivo

2015 ◽  
Vol 6 (5) ◽  
pp. 780-796 ◽  
Author(s):  
Cheng Wang ◽  
Xiuli Bao ◽  
Xuefang Ding ◽  
Yang Ding ◽  
Sarra Abbad ◽  
...  
Keyword(s):  
Transfection Efficiency ◽  
Gene Transfection ◽  
Retracted Article ◽  
First Time

A novel coating polymer LPHF is developed for the first time to elevate the transfection efficiency of DP binary polyplexes in vitro and in vivo.

A rhabdomyosarcoma hydrogel model to unveil cell-extracellular matrix interactions

2021 ◽  
Author(s):  
Mattia Saggioro ◽  
Stefania D'Agostino ◽  
Anna Gallo ◽  
Sara Crotti ◽  
Sara D'Aronco ◽  
...  
Keyword(s):  
Cell Culture ◽  
Extracellular Matrix ◽  
Three Dimensional ◽  
3D Culture ◽  
3D Cell Culture ◽  
Culture Systems ◽  
Matrix Interactions ◽  
In Vitro Systems ◽  
Extracellular Matrix Interactions

Three-dimensional (3D) culture systems are progressively getting attention given their potential in overcoming limitations of the classical 2D in vitro systems. Among different supports for 3D cell culture, hydrogels (HGs)...

scholarly journals Interstitial Flow Recapitulates Gemcitabine Chemoresistance in A 3D Microfluidic Pancreatic Ductal Adenocarcinoma Model by Induction of Multidrug Resistance Proteins

2019 ◽  
Vol 20 (18) ◽  
pp. 4647 ◽  
Author(s):  
Bart Kramer ◽  
Luuk de Haan ◽  
Marjolein Vermeer ◽  
Thomas Olivier ◽  
Thomas Hankemeier ◽  
...  
Keyword(s):  
Cell Culture ◽  
Multidrug Resistance ◽  
Pancreatic Ductal Adenocarcinoma ◽  
3D Cell Culture ◽  
In Vitro Models ◽  
Ductal Adenocarcinoma ◽  
Interstitial Flow ◽  
Multidrug Resistance Proteins ◽  
And Function

Pancreatic Ductal Adenocarcinoma (PDAC) is one of the most lethal cancers due to a high chemoresistance and poor vascularization, which results in an ineffective systemic therapy. PDAC is characterized by a high intratumoral pressure, which is not captured by current 2D and 3D in vitro models. Here, we demonstrated a 3D microfluidic interstitial flow model to mimic the intratumoral pressure in PDAC. We found that subjecting the S2-028 PDAC cell line to interstitial flow inhibits the proliferation, while maintaining a high viability. We observed increased gemcitabine chemoresistance, with an almost nine-fold higher EC50 as compared to a monolayer culture (31 nM versus 277 nM), and an alleviated expression and function of the multidrug resistance protein (MRP) family. In conclusion, we developed a 3D cell culture modality for studying intratissue pressure and flow that exhibits more predictive capabilities than conventional 2D cell culture and is less time-consuming, and more scalable and accessible than animal models. This increase in microphysiological relevance might support improved efficiency in the drug development pipeline.

Computational profiling of hiPSC-derived heart organoids reveals chamber defects associated with Ebstein’s anomaly

2020 ◽  
Author(s):  
Wei Feng ◽  
Hannah Schriever ◽  
Shan Jiang ◽  
Abha Bais ◽  
Dennis Kostka ◽  
...  
Keyword(s):  
Heart Development ◽  
Cell Types ◽  
In Vitro Models ◽  
Ebstein’S Anomaly ◽  
Great Promise ◽  
Heart Cells ◽  
Ebstein's Anomaly ◽  
Disease States ◽  
Human Fetal Heart

AbstractHeart organoids have the potential to generate primary heart-like anatomical structures and hold great promise as in vitro models for cardiac disease. However, their properties have not yet been carefully studied, which hinders a wider spread application. Here we report the development of differentiation systems for ventricular and atrial heart organoids, enabling the study of heart disease with chamber defects. We show that our systems generate organoids comprising of major cardiac cell types, and we used single cell RNA sequencing together with sample multiplexing to characterize the cells we generate. To that end, we also developed a machine learning label transfer approach lever-aging cell type, chamber, and laterality annotations available for primary human fetal heart cells. We then used this model to analyze organoid cells from an isogeneic line carrying an Ebstein’s anomaly associated genetic variant, and we successfully recapitulated the disease’s atrialized ventricular defects. In summary, we have established a workflow integrating heart organoids and computational analysis to model heart development in normal and disease states.

scholarly journals Tailoring 3D cell culture templates with common hydrogels

2018 ◽  
Author(s):  
Aurélien Pasturel ◽  
Pierre-Olivier Strale ◽  
Vincent Studer
Keyword(s):  
Cell Culture ◽  
3D Cell Culture ◽  
In Vitro Models ◽  
Manufacturing Method ◽  
Biologically Relevant ◽  
Physiological Conditions ◽  
Subtractive Manufacturing ◽  
User Friendly

3D cell culture aims at reconciliating the simplicity of in vitro models with the human like properties encountered in vivo. Soft permeable hydrogels have emerged as user-friendly materials to grow cells in more physiological conditions. With the intent on turning these homogeneous substrates into biomimetic templates, we introduce a generic solution compatible with the most biologically relevant and often frail materials. Here we take control of the chemical environment driving generic radical reactions to craft common gels with patterned light. In a simple microreactor, we harness the well-known inhibition of radicals by oxygen to enable topographical photopolymerization. Strikingly, by sustaining an oxygen rich environment, we can also induce hydrogel photo-scission which turns out to be a powerful and generic subtractive manufacturing method. We finally introduce a flexible patterned functionalization protocol based on available photo-linkers. Using these common tools on the most popular hydrogels, we tailored soft templates where cells grow or self-organize into standardized structures. The platform we describe has the potential to set a standard in future 3D cell culture experiments.

scholarly journals Bioengineered Cystinotic Kidney Tubules Recapitulate a Nephropathic Phenotype

Cells ◽  
2022 ◽  
Vol 11 (1) ◽  
pp. 177
Author(s):  
Elena Sendino Garví ◽  
Rosalinde Masereeuw ◽  
Manoe J. Janssen
Keyword(s):  
Cell Culture ◽  
Proximal Tubule ◽  
Vesicle Trafficking ◽  
Severe Disease ◽  
In Vitro Models ◽  
Nephropathic Cystinosis ◽  
Kidney Tubules ◽  
Renal Fanconi Syndrome ◽  
Dimensional Cell

Nephropathic cystinosis is a rare and severe disease caused by disruptions in the CTNS gene. Cystinosis is characterized by lysosomal cystine accumulation, vesicle trafficking impairment, oxidative stress, and apoptosis. Additionally, cystinotic patients exhibit weakening and leakage of the proximal tubular segment of the nephrons, leading to renal Fanconi syndrome and kidney failure early in life. Current in vitro cystinotic models cannot recapitulate all clinical features of the disease which limits their translational value. Therefore, the development of novel, complex in vitro models that better mimic the disease and exhibit characteristics not compatible with 2-dimensional cell culture is of crucial importance for novel therapies development. In this study, we developed a 3-dimensional bioengineered model of nephropathic cystinosis by culturing conditionally immortalized proximal tubule epithelial cells (ciPTECs) on hollow fiber membranes (HFM). Cystinotic kidney tubules showed lysosomal cystine accumulation, increased autophagy and vesicle trafficking deterioration, the impairment of several metabolic pathways, and the disruption of the epithelial monolayer tightness as compared to control kidney tubules. In particular, the loss of monolayer organization and leakage could be mimicked with the use of the cystinotic kidney tubules, which has not been possible before, using the standard 2-dimensional cell culture. Overall, bioengineered cystinotic kidney tubules recapitulate better the nephropathic phenotype at a molecular, structural, and functional proximal tubule level compared to 2-dimensional cell cultures.

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