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Updated: Jul 6, 2026
Mouse Naïve CD4+ T Cell Isolation and In vitro Differentiation into T Cell Subsets
Published on: April 16, 2015
Oridonin promotes CD4+/CD25+ Treg differentiation, modulates Th1/Th2 balance and induces HO-1 in rat splenic
1State Key Laboratory of Bioreactor Engineering & School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China.
Objective And Design:
Oridonin is an ent-kaurene diterpenoid extracted from Isodon Serra, and we have previously demonstrated its immunosuppressive effect. Our goal was to study how Oridonin impacts CD4+/CD25+ regulatory T cells (Tregs) and Th1/Th2 balance, as well as its effect on the anti-inflammatory target HO-1.
Material:
Splenic lymphocytes were prepared from male 6-8-week-old SD rats.
Treatment:
Cells were cultured in four groups as Oridonin-L (Oridonin 12.5 micromol/l), Oridonin-H (Oridonin 25 micromol/l), Cobalt protoporphyrin (Copp 50 micromol/l) and control (DMSO) with stimulation of ConA (5 microg/ml) for 48 h or with no stimulation for 12 h.
Method:
We set up a model of Th1 polarization in vitro using ConA stimulation; ratios of CD4+/CD25+ Tregs (confirmed by the expression of Foxp3) were measured by flow cytometry, and levels of IL-2, IFN-gamma, TGF-beta and IL-10 were measured by ELISA. In addition, HO-1 expression was measured without stimulation with ConA by RT-PCR and Western blotting, and HO-1 level in vitro was then measured by enzyme activity assay. p<0.05 (t-test) was taken as the level of statistical significance.
Result:
Oridonin promoted differentiation towards CD4+/CD25+ Tregs, inhibited IL-2 and IFN-gamma but induced TGF-beta and IL-10, thus rectified the Th1 polarization. Moreover, Oridonin induced the expression of HO-1 mRNA and protein, and HO-1 activity in vitro was enhanced accordingly.
Conclusion:
The results suggest that Oridonin has a distinct effect on promoting CD4+/CD25+ Treg differentiation and modulating Th1/Th2 balance, and this effect may be achieved via inducing the anti-inflammatory target HO-1.
