Related Experiment Videos
From growth factor dependence to growth factor responsiveness: the genesis of an alveolar macrophage cell line
Abstract:
A rat pulmonary alveolar macrophage (PAM) cell line (NR8383) was initiated in culture in the presence of a gerbil lung cell conditioned medium (GLCM), and has been propagated continuously for over 36 mo. When examined at different times throughout this in vitro period, NR8383 exhibited characteristics typical of macrophages: (a) Zymosan ingestion was seen in 90 to 98% of the cells examined; (b) Pseudomonas aeruginosa phagocytosis in 50 to 80%; (c) Nonspecific esterase activity in greater than 95%. During the first 6 mo., the PAM replicated with doubling times approximating 15 to 20 d. Throughout this period, GLCM dependence was evident. After 27 wk in vitro, NR8383 replication increased markedly, and within 2 wk, the doubling time was less than 48 h. NR8383 was readily monitored by [3H]thymidine (TdR) blastogenesis assay. In the presence of GLCM uptake of [3H]TdR was fivefold greater than in control cultures. Adherence and growth kinetics were effectively controlled by modulation of GLCM or serum content in culture medium. It was demonstrated that PAM growth factor(s) is ubiquitous, not species-specific, and under certain conditions may be derived from "endogenous" sources of persisting non-PAM populations within the parent, uncloned line NR8383. Cloned progeny remain devoid of non-PAM "feeder" cells, but retain macrophage properties, including interleukin-1 secretion, Fc receptors, and H2O2 production.
Insights
A rat pulmonary alveolar macrophage (PAM) cell line, NR8383, was established and maintained in culture. This cell line demonstrated stable macrophage characteristics and growth factor dependence, with significant replication increases observed over time.
Area of Science:
- Cell Biology
- Immunology
- Macrophage Research
Background:
- Establishing and characterizing cell lines is crucial for studying cellular functions.
- Pulmonary alveolar macrophages (PAMs) play a key role in lung immunity.
- In vitro models are essential for understanding macrophage behavior.
Purpose of the Study:
- To establish and characterize a rat PAM cell line (NR8383) for long-term in vitro studies.
- To investigate the growth characteristics and dependence of the NR8383 cell line.
- To confirm the retention of macrophage-specific functions in the established cell line.
Main Methods:
- Initiation and continuous propagation of NR8383 rat PAM cell line in gerbil lung cell conditioned medium (GLCM).
- Assessment of macrophage characteristics including zymosan and Pseudomonas aeruginosa phagocytosis, and non-specific esterase activity.
- Monitoring cell replication and proliferation using [3H]thymidine (TdR) blastogenesis assay.
- Analysis of growth kinetics through modulation of GLCM and serum content.
Main Results:
- NR8383 cells consistently exhibited macrophage markers (zymosan/P. aeruginosa phagocytosis, esterase activity).
- Initial slow replication (15-20 day doubling time) dependent on GLCM, followed by a significant increase in replication rate (<48h doubling time) after 27 weeks.
- [3H]TdR uptake was significantly enhanced by GLCM, indicating growth factor presence.
- Growth factor(s) were found to be ubiquitous and potentially derived from endogenous sources within the uncloned line.
- Cloned NR8383 progeny retained macrophage functions like IL-1 secretion, Fc receptors, and H2O2 production.
Conclusions:
- The NR8383 cell line serves as a stable in vitro model for rat PAMs, retaining key macrophage functions.
- The study identified growth factor(s) essential for PAM proliferation, which are ubiquitous and can be sourced endogenously.
- The characterized NR8383 cell line provides a valuable tool for immunological and cellular studies of macrophages.