IADR Abstract Archives
The key role of salivary exosomes in the accessibility of curcumin to oral cancer cells
Objectives: Curcumin (Curcuma longa) is an antioxidant, anti-inflammatory and anticancer agent with hydrophobic properties. Exosomes are phospholipid membrane-bound, extracellular nano-vesicles abundantly present in body fluids, including saliva. We showed that salivary exosomes generate complexes with curcumin which facilitate the accessibility of curcumin to oral cancer cells and enable its cellular uptake.
Methods: One ml aliquots of pooled whole saliva of healthy individuals (N=25) were divided into A– whole clean saliva (basic centrifugation 3,000g); B– whole saliva after enhanced centrifugation (12,000g); C- exosomal pellets (ultracentrifugation 120,000g) resuspended in 1ml PBS. Control groups included: D- PBS and E- 90% ethanol. All groups were incubated with 0/80µM curcumin. Differences (Δ%) in the mean rates of absorption and fluorescence of curcumin, used to reflect its activation, were assessed at T0, after 24h and 48h. Pellets were also analyzed by scanning electron microscopy, atomic force microscopy and confocal microscopy. Finally, exosomal pellets loaded with 0/80µM curcumin were incubated with human oral squamous carcinoma cell line (SCC-25) cultures. Following immunofluorescence staining and using the natural fluorescence of curcumin we have visualized the cells by confocal microscopy.
Results: T0 absorption with 80µM curcumin was similar in groups A-C and E (optic density range 2.2-2.4) but 2-fold higher than in D. The ΔT48-T0 showed minor decline in absorbance in groups A-C (-8% to -15%), remarkable decrease in D (-38%) but remained nearly constant in E (-3%). T0 fluorescence with curcumin was 1.6-2.3-fold higher in groups A-C than in D. While in group D curcumin lost 20% of its fluorescence, in groups B and C it showed a trend of increase. All morphological methods highlighted formation of exosome-curcumin complexes. Confocal imaging showed curcumin uptake by SCC-25 cells.
Conclusions: Formation of exosome-curcumin complexes may facilitate prolonged retention of curcumin in saliva and enable its uptake into oral cancer cells.
Methods: One ml aliquots of pooled whole saliva of healthy individuals (N=25) were divided into A– whole clean saliva (basic centrifugation 3,000g); B– whole saliva after enhanced centrifugation (12,000g); C- exosomal pellets (ultracentrifugation 120,000g) resuspended in 1ml PBS. Control groups included: D- PBS and E- 90% ethanol. All groups were incubated with 0/80µM curcumin. Differences (Δ%) in the mean rates of absorption and fluorescence of curcumin, used to reflect its activation, were assessed at T0, after 24h and 48h. Pellets were also analyzed by scanning electron microscopy, atomic force microscopy and confocal microscopy. Finally, exosomal pellets loaded with 0/80µM curcumin were incubated with human oral squamous carcinoma cell line (SCC-25) cultures. Following immunofluorescence staining and using the natural fluorescence of curcumin we have visualized the cells by confocal microscopy.
Results: T0 absorption with 80µM curcumin was similar in groups A-C and E (optic density range 2.2-2.4) but 2-fold higher than in D. The ΔT48-T0 showed minor decline in absorbance in groups A-C (-8% to -15%), remarkable decrease in D (-38%) but remained nearly constant in E (-3%). T0 fluorescence with curcumin was 1.6-2.3-fold higher in groups A-C than in D. While in group D curcumin lost 20% of its fluorescence, in groups B and C it showed a trend of increase. All morphological methods highlighted formation of exosome-curcumin complexes. Confocal imaging showed curcumin uptake by SCC-25 cells.
Conclusions: Formation of exosome-curcumin complexes may facilitate prolonged retention of curcumin in saliva and enable its uptake into oral cancer cells.