Partridge NC, Brown RJ, Fiacco GJ.

ASGSB Bull. 1995 Oct; 9: 28.

Dept. of Pharmacological and Physiological Science, St. Louis Univ. Medical School, MO, USA.

As a preliminary to possibly flying an experiment as part of the NASA/NIH-C.4 cell culture project, we tested the feasibility of using the CellMax automated culture system with the rat osteoblastic, osteosarcoma cell line, UMR 106-01. First, we determined that UMR cells grow extremely well in polypropylene bioreactors in the CellMax. Since direct microscopic examination is impossible, growth rates were assessed by glucose consumption and lactate production. Next, we cultured in the CellMax UMR cells stably transfected with the rat collagenase promoter (6500bp of sequence 5' to the transcriptional start site of the gene) ligated to chloramphenicol acetyltransferase (CAT, a reporter gene). These cells and the transfected gene were shown to be PTHrP-responsive in parallel tissue culture plates. When the cells reached maximal numbers (21 days in the artificial capillary module), they were treated with PTHrP (10(-7) M) or control medium for 24 h. The media were then assayed by ELISA for secreted collagenase and the cells lysed to measure CAT activity. Both parameters showed stimulation by PTHrP when measured immediately. A portion of each of the media and lysate samples was incubated at 37 degrees C for 13 days to mimic flight conditions which could possibly be experienced using the Space Tissue Loss Apparatus (STL-A, the flight equivalent of a CellMax). Collagenase in the media was stable at 37 degrees C over 13 days. However, CAT activity and intracellular proteins were unstable under these conditions, degrading to 20% of initial values. Thus, to do the experiment as proposed we need either refrigeration to be provided as part of the STL-A equipment and/or a cocktail of protease inhibitors to prevent breakdown of intracellular proteins.

Publication Types:

• Meeting Abstracts

Keywords:

• Animals
• Base Sequence
• Cats
• Cell Culture Techniques
• Chloramphenicol O-Acetyltransferase
• Collagenases
• Laboratory Techniques and Procedures
• Osteosarcoma
• Parathyroid Hormone-Related Protein
• Promoter Regions (Genetics)
• Rats
• Research Design
• genetics
• organization & administration
• NASA Discipline Musculoskeletal
• NASA Discipline Number 40-40
• NASA Program Space Biology
• Non-NASA Center

Other ID:

• 97613882

UI: 102222691

From Meeting Abstracts