Cellular Biotechnology Operations Support Systems: Human Renal Cortical Cell Differentiation and Hormone Production (CBOSS-01-02-Renal) - 07.14.16
Cells grown in microgravity grow and replicate into complex structures, unlike cells grown on Earth. To better understand the mechanisms that cause several kidney disorders, human renal cortical epithelial (kidney) cell lines were grown on ISS. Microgravity allows the cells to grow in three-dimensional structures which are similar to how they grow in the human body. The cells were returned to Earth and were used in studies of several kidney disorders. Science Results for Everyone
The Cellular Biotechnology Operations Support System (CBOSS) onboard the International Space Station supports various cell culture investigations. Problems with previous payloads in this hardware led to additional research and improvements. This investigation grew and examined human kidney cells to advance understanding of the mechanisms of several kidney disorders. Cultures of renal cortical cells in space and on the ground had similar immune reaction profiles for the different antibodies tested in both. This suggests that the techniques used can be generalized to other cell lines in future research and that an RNA stabilizing agent (RNAlater-Ambion) can be used for storage of proteins for long-duration investigations. Experiment Details
Timothy G. Hammond, M.B.B.S., Durham Veterans' Affairs Medical Center, Durham, NC, United States
Wyle, Integrated Science and Engineering, Houston, TX, United States
Sponsoring Space Agency
National Aeronautics and Space Administration (NASA)
Human Exploration and Operations Mission Directorate (HEOMD)
ISS Expedition Duration
August 2001 - June 2002
A previous version of the BSTC flew on STS-86/Mir and STS-90. BTR flew on STS-89 and STS-91. Cell-growth experiments have flown on the Shuttle since STS-70.
- CBOSS-01-02-Renal is one of seven in the CBOSS suite of experiments. The purpose of the CBOSS experiments is to support biotechnological research aboard ISS by providing a stable environment for growing cells.
- This self-contained apparatus is designed to allow multiple experiments studying various types of cells to operate simultaneously. It is a multi-component cell incubator intended to grow three-dimensional clusters of cells in microgravity.
- CBOSS-01-02-Renal will provide the cells that will be used for investigating human renal cells in order to understand the mechanisms in several kidney disorders.
The purpose of the Cellular Biotechnology Operations Support System (CBOSS) study was to support biotechnological research on board ISS by providing a stable environment in which to grow cells. The system was a multi-component cell incubator intended to grow three-dimensional clusters of cells in microgravity. A self-contained apparatus, CBOSS was designed to allow multiple experiments to be performed, thereby enabling scientists to study various types of cells operating simultaneously.
In the human body, cells normally grow within a scaffolding of protein and carbohydrate fibers that creates a three dimensional structure. But outside the body, cells tend to grow in flat sheets and are incapable of duplicating the structure they normally hold, which can make them behave differently in the laboratory than they would in the body. Past research has shown that cells grown in a microgravity environment arrange themselves into three-dimensional shapes that more closely duplicate how they would behave in the body. Cell culture in microgravity thus becomes a tool for studying cells in a state that is closer to that which occurs normally in the body.
To better understand the mechanisms that cause several kidney disorders, human renal cortical epithelial cell lines were grown on station. This experiment used kidney cells to study the mechanism by which the kidney reabsorbs proteins that are filtered from the blood. The goal of this ISS experiment was to again create three-dimensional growth of normal human renal cells, and to assess the production of erythropoietin and vitamin D3 while assessing the model for production of commercial applications.
Development of techniques to reliably cultivate organisms under controlled conditions is essential to understanding the effect of microgravity and radiation on living organisms and creating environmental conditioning sources for long term space flight.
In the human body cells normally grow within a scaffolding of protein and carbohydrate fibers that help create a three dimensional structure. This is how organs maintain their shape. Studying cells on Earth is difficult because outside the body cells tend to grow in flat sheets and are not capable of duplicating the structure they normally hold, which often makes them behave differently in the lab than they would in the body. Past research, however, has shown that cells grown in microgravity arrange themselves into three-dimensional shapes, more closely duplicating how they behave in the body. CBOSS, then, becomes very useful as a tool for studying cells in a state as close to that which occurs normally in the body. CBOSS-01-02-Renal may lead to improved techniques in treating renal injuries and renal disorders.
Operational Requirements and Protocols
BSTC and GSM were housed side by side in lockers 1 and 5, respectively, of EXPRESS Rack 4. At the end of Increment 3, BTR was transferred to EXPRESS Rack 4, as well. GSM does not use power or gas supplied by the EXPRESS Rack, but does interface with the Station computer via the EXPRESS Rack's Ethernet connection. BCSS-1 and -4, housed directly below BSTC and GSM in lockers 2 and 6, do not require EXPRESS Rack support. BTR is located in EXPRESS Rack 1, locker 6. It operates on 160 watts of continuous power and Ethernet connection provided by the rack.
After the CBOSS hardware was installed on ISS, the crew activated the experiment and monitored the status of the experiment and hardware. Crew members used a syringe to inject cells into fresh TCMs, by using the injection ports. They also added fresh media to growing cell lines and fixatives at the end of the experiment to halt growth. When the cultures were fixed, the TCMs were transferred to the BTR for storage. The crew performed periodic preventive maintenance on the CBOSS components. Video and data was downloaded to the CBOSS flight control team at the Johnson Space Center's Telescience Center. The CBOSS-01-02-Renal experiment TCM was returned to the principle investigator for in-depth analysis.
Decadal Survey Recommendations
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The CBOSS hardware supported six cell culture investigations with different detailed scientific objectives. There were problems in the growth and preservation of all of the cell lines grown on Expeditions 3 and 4. The PC12 and erythroleukemia cells did not survive well in long term culture, so no scientific results are expected from these experiments. It was found that there was more bubble formation than expected that may lead to cell death at the air-liquid interface. Although not well documented in this experiment, it was noted that poor mixing of cells/tissues and medium occurred in the other CBOSS payloads as well. Both the poor mixing and greater than expected bubble formation were important lessons learned that led to the addition of the CBOSS-Fluid Dynamics Investigation (CBOSS-FDI) to study mixing and bubble formation in microgravity on later Expeditions.
Renal cortical cells returned were treated with an RNA stabilizing agent (RNAlater-Ambion) that enabled analyses of both RNA and immunoreactive proteins. The space and ground control cell cultures exhibited similar immunoreactivity profiles for the antibodies tested. These data provide evidence that the techniques used can be generalized to other cell lines, and that RNAlater will provide long-term storage of proteins at 4 degrees C (39 degrees F) for long duration investigations (Hammond et al. 2006). (Evans et al. 2009)
Ground Based Results Publications
Wilson JW, Ott CM, Ramamurthy R, Porwollik S, McClelland M, Pierson DL, Nickerson CA. Low-Shear modeled microgravity alters the Salmonella enterica serovar typhimurium stress response in an RpoS-independent manner. Applied and Environmental Microbiology. 2002; 68(11): 5408-5416. DOI: 10.1128/AEM.68.11.5408-5416.2002.
Hammond TG, Benes E, O-Reilly KC, Wolf D, Linnehan RM, Taher A, Kaysen JH, Allen PL, Goodwin TJ. Mechanical culture conditions effect gene expression: gravity-induced changes on the space shuttle. Physiological Genomics. 2000 09/08/2000; 3(3): 163-173. PMID: 11015612.
Hammond TG, Hammond JM. Optimized suspension culture: the rotating-wall vessel. American Journal of Physiology: Renal Physiology. 2001; 281(1): F12-F25.
Jessup JM, Pellis NR. NASA biotechnology: cell science in microgravity.. In Vitro Cellular and Developmental Biology - Animal. 2001; 37(2): 2. PMID: 11332737.
NASA Fact Sheet, Expedition 3
NASA Fact Sheet, Expedition 4
Microscopy image of human renal cortical epithelial cells. Image courtesy of Marshall Space Center.
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NASA Image: ISS0003E5276 - Image of a Quad Tissue Culture Module Assembly (QTCMA) 7 on ISS Expedition 3 after activation of the cells. A syringe was used to inject cells into the pink nutrient growth media. The BSTC can hold eight of these QTCMAs, which will be used to grow human cells on ISS. When the samples completed their growth cycle, the crew transfered the QTCMAs from the BSTC to the BTR, where they were stored until they are examined at a ground-based laboratory.
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