A complete system for studying viral function and replication
Cytokine storms | RNA Therapy | Glycomics | Viral infection | Virus production
Cytokine storms | RNA Therapy | Glycomics | Viral infection | Virus production
At Lena Biosciences we are doing our part by making our technologies and know-how available to viral researchers. In Perfused Organ Panel microphysiological system host cells can be grown at tissue-like cell densities providing high concentration of viral particles, and a concentrated secretome to detect cytokine storm like responses in vitro. Cells cultured in our system have higher metabolic activity and enhanced drug metabolism. Three key components of the system are the Blood Substitute that mimics the functions of hemoglobin, in-well perfusion that provides concentrated secretome, and SeedEZ three dimensional scaffold which allows growing cells at high densities.
SeedEZ
3D cell culture scaffold The scaffold supports high density cultures of host cells in a small footprint. Viral titers can be grown in relatively large quantities for studying the inhibition to viral entry, and extended viral life cycles to viral drug screens in a multiwell plate format.
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Perfused Organ Panel
Microphysiological system Perfused Organ Panel has PerfusionPal plates with SeedEZ scaffolds. In-well perfusion of dense 3D cell cultures in the scaffolds via hemoglobin-like Blood Substitute yields concentrated cytokines in the wells for time-course analyses.
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GradientEZ
3D cell migration assay system GradientEZ allows to study 3D migration of antigen-presenting cells, innate and adaptive immunity cells to the site of viral infection. It enables parallel testing of up to 6 immune modulators, or modulators of cell motility or invasion in a single well.
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RNA Therapy
One of the key hurdles in RNA therapy is ensuring targeted delivery of RNA to cells. Testing RNA therapies in vitro thus requires an accurate cellular organization and morphology in order to be predictive of success in vivo. Cells grown in 2D do not exhibit physiologically representative morphology and function. SeedEZ scaffolds facilitate 3D culture, restoring in vivo cell characteristics, including reception of and response to candidate RNA therapies. Such advantages are critical to the development of in vitro models that translate to humans.
Glycomics
The extraordinary complexity of the glycome makes it particularly difficult to study in vitro using standard 2D culture techniques. The artificial geometry of the cells and their tendency to de-differentiate has substantial effects on glycosylation pathways and the carbohydrate composition of the cell membrane. Culturing cells in 3D using SeedEZ scaffolds is a simple way to encourage in vivo geometry and function, restoring normal post-translational modification and other glycosylation pathways. The 3D cellular architecture ensures more in vivo-like, differentiated behavior, a key asset in glycomics research.
Viral Infection
3D cultures help bridge the gap between the use of cell lines (longevity in culture, but lack physiological parity) and organ cultures (physiologically-relevant, but can only survive short-term). Seeding primary or iPSC-derived target cells in SeedEZ scaffolds for viral infection studies generates a tissue-like construct that will more faithfully represent the in vivo environment in which the cells would be exposed to virus. Integration of the SeedEZ scaffolds into the Perfused Organ Panel extends the viability of the culture and models interstitial fluid flow to more accurately mimic the replication and spread of the virus. Since the cells are maintained in a low-stress environment, there are minimal artifacts that would impact (positively or negatively) the viral life cycle. These advantages are critical for accurate in vitro modeling of viral infection and replication.
Virus Production
Large-scale production of viruses in mammalian cells requires immense incubator real estate using 2D methods. Further, viral products need to be concentrated from a large, dilute pool. Culturing HEK293 cells in 3D using SeedEZ scaffolds provides 10X the surface area for cellular growth as compared to 2D, while keeping the volume of medium the same. This results in a concentrated virus yield using a fraction of the space. In addition to efficiency, 3D cultures ensure reproducibility and quality of viral production.
Brain effects (COVID-19)
As doctors and scientists work tirelessly to learn as much about COVID-19 as possible, biotechnology companies have joined the fight by adapting technologies to study the disease. Although initially thought to be solely a respiratory ailment, new research has shown that many other organ systems can be dramatically affected, particularly in the more serious cases. Recent reports have indicated that neurological symptoms linked to inflammation are more common than previously thought.
Lena Biosciences’ Perfused Organ Panel has been used to develop a model of human cortex to study inflammatory cascades. In a collaboration with Prof. Zhexing Wen at Emory University, an expert in cerebral organoids, and in association with the Marcus Center for Therapeutic Cell Characterization and Manufacturing at Georgia Tech, Prof. Michelle LaPlaca, we created a three-dimensional (3D) cortical model comprising human neurons, astrocytes, and microglia. Patient-derived, naïve human T cells were activated in vitro, and their conditioned medium was used to expose the cortical cultures to a physiological inflammatory stimulus. Cultures grown in the Perfused Organ Panel showed a significant increase in CXCL10 release in response to the activated T cell medium, whereas 3D control cultures showed an elevated, but not significant response. CXCL10 (interferon gamma-induced protein 10) is a chemokine that is integral to the T cell-mediated adaptive immune response. The ability to concentrate such molecules in vitro in a physiologically meaningful way is a critical tool in the fight to understand the mechanisms underlying the devastating effects of COVID-19.
Lung air-liquid interface cultures
The study of respiratory infection and disease in vitro is limited by the artificial cellular environment of lung epithelial cells in a dish. Air-liquid interface cultures offer a method by which epithelial cells are grown on a membrane such that their basal side is in contact with cell culture medium, while their apical side is exposed to air. This allows cells to undergo differentiation to the mature phenotypes of the airway epithelium. SeedEZ scaffolds are an ideal substrate for cellular growth and differentiation due to their hydrophilicity, porosity, and capacity to accept any matrix desired. Floating cell-seeded scaffolds on our gas-rich Blood Substitute will further support cellular differentiation and allow for additional gas exchange on the basal side of the culture, representing interstitial capillaries for a more complete model of lung epithelium in vitro.
Skin epithelial raft cultures
In vitro study of epidermal tissue using standard 2D methods limits cellular differentiation and fails to model physiological exposure to compounds. Epithelial raft cultures comprise keratinocytes seeded on collagen gels that are subsequently elevated to allow the apical surface contact with air. This allows cells to differentiate and stratify into an epidermis-like tissue. SeedEZ scaffolds are an ideal substrate for raft culture as their structure readily accepts matrices such as collagen upon which keratinocytes can be seeded. Floating these rafts on our gas-rich Blood Substitute further enhances differentiation and stratification by supplying the dividing cells on the basal side with additional gas-exchange, representing dermal capillaries for a more complete in vitro skin model.
In vitro study of epidermal tissue using standard 2D methods limits cellular differentiation and fails to model physiological exposure to compounds. Epithelial raft cultures comprise keratinocytes seeded on collagen gels that are subsequently elevated to allow the apical surface contact with air. This allows cells to differentiate and stratify into an epidermis-like tissue. SeedEZ scaffolds are an ideal substrate for raft culture as their structure readily accepts matrices such as collagen upon which keratinocytes can be seeded. Floating these rafts on our gas-rich Blood Substitute further enhances differentiation and stratification by supplying the dividing cells on the basal side with additional gas-exchange, representing dermal capillaries for a more complete in vitro skin model.
Cardiosphere-derived cells (cell therapy)
Cardiosphere-derived cells (CDCs) are a promising cellular therapeutic that have demonstrated efficacy in treating patients with heart disease, and more recently, those suffering from severe cases of COVID-19. Preliminary studies suggest that infusion of allogeneic CDCs into COVID-19 patients resulted in life-saving improvements to respiratory and cardiac symptoms. Perfused Organ Panel is an ideal platform for in vitro efficacy studies of CDCs across multiple organ systems, simultaneously. Lung and cardiac tissue in which inflammation is induced can be cultured alongside normal tissues (liver, gut, lymphocytes, etc) in a single plate, exposed to CDCs, and assessed for on-target efficacy and off-target inflammation. The gas-rich environment provides a more physiologically-relevant environment, making in vitro cell therapy studies more predictive.
Cardiosphere-derived cells (CDCs) are a promising cellular therapeutic that have demonstrated efficacy in treating patients with heart disease, and more recently, those suffering from severe cases of COVID-19. Preliminary studies suggest that infusion of allogeneic CDCs into COVID-19 patients resulted in life-saving improvements to respiratory and cardiac symptoms. Perfused Organ Panel is an ideal platform for in vitro efficacy studies of CDCs across multiple organ systems, simultaneously. Lung and cardiac tissue in which inflammation is induced can be cultured alongside normal tissues (liver, gut, lymphocytes, etc) in a single plate, exposed to CDCs, and assessed for on-target efficacy and off-target inflammation. The gas-rich environment provides a more physiologically-relevant environment, making in vitro cell therapy studies more predictive.
Cardiosphere-derived cells (exosomes)
Preliminary studies suggest that infusion of allogeneic cardiosphere-derived cells (CDCs) into COVID-19 patients resulted in life-saving improvements to respiratory and cardiac symptoms. The mechanism of action is through the release of exosomes, the cargo of which includes paracrine signaling molecules that can suppress cytokine-mediated inflammation. Perfused Organ Panel featuring SeedEZ 3D culture scaffolds is an ideal platform for CDC culture for the purpose of producing exosomes for use in in vitro efficacy studies. Perfusion-assisted exosome release from CDCs cultured in SeedEZ in Perfused Organ Panel produces a concentrated pool of exosomes that can be delivered directly to target tissues cultured alongside the CDC in the same plate, serving as an all-in-one production and testing platform.
Preliminary studies suggest that infusion of allogeneic cardiosphere-derived cells (CDCs) into COVID-19 patients resulted in life-saving improvements to respiratory and cardiac symptoms. The mechanism of action is through the release of exosomes, the cargo of which includes paracrine signaling molecules that can suppress cytokine-mediated inflammation. Perfused Organ Panel featuring SeedEZ 3D culture scaffolds is an ideal platform for CDC culture for the purpose of producing exosomes for use in in vitro efficacy studies. Perfusion-assisted exosome release from CDCs cultured in SeedEZ in Perfused Organ Panel produces a concentrated pool of exosomes that can be delivered directly to target tissues cultured alongside the CDC in the same plate, serving as an all-in-one production and testing platform.
Cancer research and cell therapies
Cytolytic T-cell Induction
There has been interest in using patient PBMCs to select T-cells to kill cancer cells. Activated T-cells can be selected, propagated to desired numbers and returned to patients for treatment. Cytolytic T-cell activation in the presence of patient-derived tumor cells can be greatly facilitated when using a perfused microphysiological system. Physiological oxygen levels and tumor spheroids mimic the in vivo environment to enhance active cytolytic selection. The Lena Biosciences PerfusionPal uses a hemoglobin analogue, Blood Substitute, that mimics the functions of red blood cells in vivo, that retains tumor structure, and maintains cultures for up to 4 weeks to maximize the successful selection and expansion of cytolytic T-cells.
Cancer Cell Line Establishment
Cancer patient cell line establishment is an important step in personalized medicine. Enhancing the metabolic activity of cells during the establishment and propagation stage are essential to generating data when the time is critical. The Lena Biosciences PerfusionPal uses its patented technology that activates the metabolic activity of cancer cells to accelerate cell growth in sufficient numbers.
Higher Assay Sensitivity
Chemotherapeutic screens for new drug candidates can be made more sensitive by elevating cytochrome P450 activity. Cytochrome P450 activity is greatly enhanced when the cells are cultured in 3D and perfused. Active perfusion with a hemoglobin analog greatly increases cytochrome P450 activity resulting in more sensitive cell-based assays when screening for chemotherapeutic efficacy.
Cytolytic T-cell Induction
There has been interest in using patient PBMCs to select T-cells to kill cancer cells. Activated T-cells can be selected, propagated to desired numbers and returned to patients for treatment. Cytolytic T-cell activation in the presence of patient-derived tumor cells can be greatly facilitated when using a perfused microphysiological system. Physiological oxygen levels and tumor spheroids mimic the in vivo environment to enhance active cytolytic selection. The Lena Biosciences PerfusionPal uses a hemoglobin analogue, Blood Substitute, that mimics the functions of red blood cells in vivo, that retains tumor structure, and maintains cultures for up to 4 weeks to maximize the successful selection and expansion of cytolytic T-cells.
Cancer Cell Line Establishment
Cancer patient cell line establishment is an important step in personalized medicine. Enhancing the metabolic activity of cells during the establishment and propagation stage are essential to generating data when the time is critical. The Lena Biosciences PerfusionPal uses its patented technology that activates the metabolic activity of cancer cells to accelerate cell growth in sufficient numbers.
Higher Assay Sensitivity
Chemotherapeutic screens for new drug candidates can be made more sensitive by elevating cytochrome P450 activity. Cytochrome P450 activity is greatly enhanced when the cells are cultured in 3D and perfused. Active perfusion with a hemoglobin analog greatly increases cytochrome P450 activity resulting in more sensitive cell-based assays when screening for chemotherapeutic efficacy.