Cellular Biotechnology Operations Support Systems: Evaluation of Ovarian Tumor Cell Growth and Gene Expression (CBOSS-01-Ovarian) - 01.09.14
Science Objectives for Everyone Cells grown in microgravity grow and replicate into complex structures, unlike cells grown on Earth. To better understand the mechanisms that cause the differentiation of cells in microgravity, a human ovarian tumor cell line was grown on ISS. The cells were returned to Earth and were used in the studies to define mechanisms of ovarian cancer with the goal of developing new ovarian cancer treatments.
Science Results for Everyone
LN1 ovarian tumor cells grown on the International Space Station did not survive long-term, but generated reduced amounts of cytokines (small secreted proteins) compared to those grown on the ground. These were vimentin, an intermediate filament protein in embryonic cells that plays a role in abnormal tissue growth, and EMA, a human milk fat globulemembrane protein reactive against many types of gland tumors. The data indicates the presence of these proteins in stabilized LN1 cells after long-duration storage at 39 degrees F, with similar profiles at different times in flight and on the ground. This confirms that techniques used here can be generalized to other cell lines for long-duration investigations.
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 - December 2001
Previous ISS Missions
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-Ovarian is one of seven in the CBOSS suite of experiments. The purpose of CBOSS 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-Ovarian examines growth and development of LN1 human ovarian tumor cells.
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.
The goal of this study was to characterize the complex three-dimensional development of the human Muellerian ovarian (LN1) tumor cell line to characterize morphological changes that occur during three-dimensional growth. This study also sought to determine accompanying alterations in cell cycle kinetics, cell cycle proteins, and cellular oncoproteins. Cells were preserved in RNAlater, a fixative that allows cells to remain stable at refrigerator temperatures (4 degrees C, 39 degrees F) for up to 30 days. Following return to Earth after three months on ISS, the cells were analyzed for antigenic stability after removal of RNA using the RNAqueous kit. Knowledge gained from this experiment could help define mechanisms in tumor cell development that can be targeted for treatment of patients with ovarian cancer.
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. Its benefits include applications in cancer research.
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 CBOSS-01-Ovarian 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 the growing cell line 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-Ovarian experiment's TCM was returned to the principle investigators for in-depth analysis.
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.
The LN1 ovarian cell cultures on board station did not survive in long-term culture. However, the cells grown on ISS were found to have produced reduced amounts of cytokines (small secreted proteins that mediate and regulate immunity, inflammation, and hematopoiesis) compared to the ground controls. The proteins were recovered after the RNA had been removed from the cells via filtration. The novel proteins, vimentin and epithelial membrane antigen (EMA) proteins, were extracted from filtrate of the RNA extraction. Vimentin is the main intermediate filament protein in embryonic cells. It plays an important role in the differential diagnosis of undifferentiated neoplasms (abnormal tissue growths). EMA, which belongs to a family of proteins known as human milk fat globulemembrane proteins, is considered a broad spectrum antibody that is reactive against many types of adenocarcinomas. The data obtained from the protein extraction indicate the presence of the antigenic proteins, vimentin and EMA, in RNA-stabilized LN1 cells following long-duration storage at 4 degrees C (39 degrees F). The vimentin and EMA proteins showed similar profiles at different times between the flight and ground samples. These data provide confirmation 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. 2005). (Evans et al. 2009)
Ground Based Results Publications
Jessup JM, Pellis NR. NASA biotechnology: cell science in microgravity.. In Vitro Cellular and Developmental Biology - Animal. 2001; 37(2): 2.
Scanning Electron Micrograph of an human muellerian ovarian cancer cell nurtured in microgravity conditions. The three-dimensional structure shown is much closer in true size and form to natural tumor cells found in cancer patients. Courtesy image of Marshall Space Flight Center.
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NASA Image: ISS003E5285 - Image of a Quad Tissue Culture Module Assembly (QTCMA) 6 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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