Abstract
BackgroundPeptides with cytoprotective functions, including antioxidants and anti-infectives, could be useful therapeutics. Carnosine, β-alanine-histidine, is a dipeptide with anti-oxidant properties. Tripeptides of Ala-His-Lys, Pro-His-His, or Tyr-His-Tyr are also of interest in this respect.ResultsWe synthesized several histidine-containing peptides including glycine or alanine, and tested their cytoprotective effects on hydrogen peroxide toxicity for PC12 cells. Of all these peptides (Gly-His-His, Ala-His-His, Ala-His-Ala, Ala-Ala-His, Ala-Gly-His, Gly-Ala-His (GAH), Ala-His-Gly, His-Ala-Gly, His-His-His, Gly-His-Ala, and Gly-Gly-His), GAH was found to have the strongest cytoprotective activity. GAH decreased lactate dehydrogenase (LDH) leakage, apoptosis, morphological changes, and nuclear membrane permeability changes against hydrogen peroxide toxicity in PC12 cells. The cytoprotective activity of GAH was superior to that of carnosine against hydrogen peroxide toxicity in PC12 cells. GAH also protected PC12 cells against damage caused by actinomycin D and staurosporine. Additionally, it was found that GAH also protected SH-SY5Y and Jurkat cells from damage caused by hydrogen peroxide, as assessed by LDH leakage.ConclusionThus, a novel tripeptide, GAH, has been identified as having broad cytoprotective effects against hydrogen peroxide-induced cell damage.
Highlights
Peptides with cytoprotective functions, including antioxidants and anti-infectives, could be useful therapeutics
We analyzed the cytoprotective functions of GAH, compared to the other tripeptides used in this study
GAH at 0.1 μg/μL, 1 μg/μL, and 5 μg/μL did not inhibit enzyme activity (Fig. 3). These results indicate that GAH did not directly block lactate dehydrogenase (LDH) activity, but most likely prevented its leakage into the media, caused by loss of membrane integrity
Summary
We incubated PC12 cell with 5000 μM hydrogen peroxide with 1 μg/μL of GHH, AHH, AHA, AAH, AGH, GAH, AHG, HAG, HHH, GHA, GGH, or carnosine for 4 h. The addition of GAH together with hydrogen peroxide increased the number of adherent cells and maintained the flat shape of the cell compared to the effect of hydrogen peroxide alone or after coculture with HAG, GHH, AHG, AGH, AAH, GHH, AHH, AHA, or carnosine (Fig. 4). The results showed that cell viability (as measured at OD 450 nm) was significantly increased in the GAH (0.625 ± 0.023)- and carnosine (0.711 ± 0.026)-treated groups relative to controls (0.144 ± 0.012, p < 0.01) (Fig. 8). GAH did prevent cell death induced by actinomycin D (p < 0.001) and staurosporine (p < 0.001)) similar to its effects on hydrogen peroxide toxicity (Fig. 9). At 1 μg/μL, GAH decreased the amount of LDH released from SH-SY5Y cells (p = 0.0039) (Fig. 10b) These findings indicate that GAH prevents Jurkat and SH-SY5Y cell membrane damage caused by hydrogen peroxide
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