Introduction:The primary cilium is a sensory organelle that transmits environmental information to the cell interior. The PC has cytoplasmic accessory protein complexes such as the Bardet-Biedl family of proteins that may be involved in processing external stimuli to the cell such as oxidative status. Pleural effusion pathology often involves high oxidative stress and this is sparsely characterized with respect to the PC. We evaluated the BBSome gene(s) expression during oxidative stress using 100mM H2O2 stimulus in benign mesothelial cells in 2D and 3D spheroid models.

Methods:Benign mesothelial cells, MeT-5A were used in the study. Cells were grown either on standard petri dish surface or as spheroids by a hanging drop method using standard culture medium or media with 100?M H2O2 for 24 hours. Cells were lysed for RNA processing and cDNA was prepared followed by qPCR quantitation with gene specific primers for BBS1, BBS2, BBS4, BBS5, BBS7, BBS9 and BBS18 and ?actin. Log2 of 2-??Ct values were used to compare relative gene expression.

Results:We observed significant changes to BBS1 (Con mean±SEM;0.0±0.2, H2O2 1.23±0.49,p<0.05) and BBS7 (Con;0.0±0.08, H2O2 1.31±0.57,p<0.01), BBS5 was undetectable in 2D but observed in 3D irrespective of 100mM H2O2 stimulus. BBS9 in 3D was significantly down regulated (Con;0.0±0.2, H2O2 -1.11±0.35,p<0.01). Most BBS genes in 3D were negative values with no significant changes from peroxide stimulus.

Conclusions:Oxidative stress induces changes BBS gene expression in 2D and 3D culture configuration. Expression patterns differ qualitatively between 2D and 3D cell culture and this warrants further investigation.