PTD-fusion protein technology was used to transduce heat shock protein 27 (HSP27), an anti-apoptotic protein, into human lens epithelial cells (HLECs) (SRA01/04). The protein transduction domain (PTD) of the 11-amino acid YGRKKRRQRRR was tagged at the N-terminus of HSP27. The fusion protein was purified from bacteria transformed with a pKYB-PTD-HSP27 construct. The HLECs were incubated with PTD-HSP27-FITC and the fluorescence inside HLECs was found by fluorescence microscopic examination. To test the ability of PTD-HSP27 to pass through the corneas, PTD-HSP27-FITC was dropped onto the conjunctival sacs of rabbits; fluorescent labeled PTD-HSP27 was then observed in the rabbit aqueous humor. After being incubated with the PTD-HSP27 protein and irradiated with ultraviolet-B (UVB) light, HLECs was analyzed by flow cytometry, Hoechst 33258 staining and measurement of the potential of the mitochondrial transmembrane. HLECs incubated with PTD-HSP27 had a lower apoptotic rate and a higher mitochondrial membrane potential than the control cells. PTD-HSP27 appears to be sufficient to protect HLECs against UVB-induced apoptosis.
Keywords: HSP27; apoptosis; cataract; human lens epithelial cell (HLEC); protein transduction domain (PTD); transactivator of transcription (TAT).
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