Related Experiment Video
Updated: Aug 9, 2026
Intravital Imaging of Neutrophil Priming Using IL-1β Promoter-driven DsRed Reporter Mice
Published on: June 22, 2016
Production of both interleukin-1 alpha and beta by newborn mouse calvarial cultures
J A Lorenzo1, S L Sousa, S E Van den Brink-Webb
1Department of Medicine, Veterans Administration Medical Center, Newington, CT 06111.
Insights
Newborn mouse calvarial cultures release bioactive interleukin-1 alpha (IL-1 alpha), a key regulator of bone cell function, into conditioned medium, influencing bone growth and remodeling.
Area of Science:
- Immunology
- Developmental Biology
- Cell Biology
Background:
- Conditioned medium (CM) from newborn mouse calvarial cultures exhibits thymocyte comitogen proliferation activity.
- This activity is specifically inhibited by antiserum to murine interleukin-1 alpha (IL-1 alpha), not IL-1 beta.
- The release of this activity is independent of endotoxin and not linked to interleukin-2 (IL-2).
Purpose of the Study:
- To characterize the molecular nature of the thymocyte comitogen proliferation activity in CM from newborn mouse calvarial cultures.
- To determine the specific form(s) of IL-1 involved in this activity.
- To investigate the role of local IL-1 production in regulating bone cell function and bone remodeling.
Main Methods:
- Gel filtration chromatography to determine the molecular weight of the active component.
- Western immunoblotting using antisera against murine IL-1 alpha and IL-1 beta.
- Immunoprecipitation of [35S]methionine-labeled CM with anti-IL-1 alpha antisera.
Main Results:
- Gel filtration indicated an active component with a peak molecular weight of 16-18 kD.
- Western immunoblots detected a 33 kD band for IL-1 beta but no band for IL-1 alpha.
- Sensitive immunoprecipitation assays revealed 15-17 kD IL-1 alpha bands, indicating its presence.
- The study identified both 17 kD IL-1 alpha (major bioactive form) and 33 kD IL-1 beta (greater quantity) in the CM.
Conclusions:
- Newborn mouse calvarial cultures spontaneously release IL-1 alpha and IL-1 beta.
- IL-1 alpha appears to be the primary bioactive form regulating thymocyte proliferation in this context.
- Local IL-1 production likely plays a significant role in regulating bone cell function, bone growth, and remodeling.
Abstract:
The conditioned medium (CM) from 4-6 day newborn mouse calvarial cultures was found to contain thymocyte comitogen proliferation activity. This activity was blocked by an antiserum to murine interleukin-1 alpha (IL-1 alpha) but not by an antiserum to murine interleukin-1 beta. The release of thymocyte comitogen proliferation activity from the cultures did not appear dependent on endotoxin and was not associated with detectable interleukin-2 activity in the CM. Activity in the CM eluted from a gel filtration column with a peak Mr of 16-18 kD (the Mr of mature murine IL-1 alpha and beta is 17 kD). Western immunoblots of 100-fold concentrated CM demonstrated only a single 33 kD band with an antiserum to murine IL-1 beta and no bands with an antiserum to murine IL-1 alpha. However, this assay was relatively insensitive (limit of detection 1-10 ng compared with 1-10 pg for the thymocyte comitogen proliferation assay). Immunoprecipitation of [35S]methionine-labeled CM with three different anti-IL-1 alpha antisera, a more sensitive assay, demonstrated 15-17 kD bands in all cases. These results demonstrate that 4-6 day newborn mouse calvarial cultures spontaneously release 17 kD IL-1 alpha and 33 kD IL-1 beta into their conditioned medium. It appears that although 17 kD IL-1 alpha is the major bioactive form in the CM, 33 kD IL-1 beta is present in greater amounts. These results also suggest that local production of IL-1 can regulate bone cell function and may play a role in bone growth and remodeling.
