Unit:
Κατεύθυνση Κοσμητολογία - ΔερματοφαρμακολογίαLibrary of the School of Science
Author:
Grammatikaki Ntiana-Stefania
Supervisors info:
Λιβανίου Ευαγγελία, Ερευνήτρια Α΄, ΙΠΡΕΤΕΑ, ΕΚΕΦΕ «Δημόκριτος»
Κλέτσας Δημήτριος, Ερευνητής Α΄ - Διευθυντής ΙΒΕ, ΕΚΕΦΕ «Δημόκριτος»
Original Title:
Πεπτίδια με πιθανή επουλωτική δράση
Translated title:
Peptides with potential healing effect
Summary:
The aim of the present study was, first, the laboratory synthesis, purification and analytical characterization of acetylSer-Asp-Lys-Pro-OH (AcSDKP) and acetylAla-Asp-Lys-Pro-OH (AcADKP) peptides and, second, the in vitro biological evaluation of these peptides, using specific cell lines which are involved in the dermal healing process. The AcSDKP and AcADKP peptides are identical with the N-terminal tetrapeptide of two longer bioactive peptides, the 43-peptides thymosin β4 (Tβ4) and thymosin β10 (Tβ10), respectively. Tβ4 and Tβ10 exhibit high structural homology and belong to a family of endogenous peptides, the β-thymosins. They are expressed in almost all human cells and their biological activity appears to be important. According to literature data, both Tβ4 and AcSDKP (which is formed by Tβ4 proteolysis) have a positive effect on dermal wound healing, while less is known about Tβ10 and its fragments.
The synthesis of the tetrapeptides AcSDKP and AcADKP was performed with the Fmoc - solid phase peptide synthesis technique. Purification of AcSDKP and AcADKP was performed with semi-preparative RP-HPLC and the final product purity was evaluated with analytical RP-HPLC (in which commercially available AcSDKP was used as an internal standard). The purity of the peptides was excellent and the overall synthesis yield (~ 30%) was very satisfactory. Pure peptides were identified by Electrospray Ionization – Mass Spectrometry (ESI-MS).
The in vitro biological evaluation of AcSDKP and AcADKP was performed as follows: Initially, the potential cytotoxic effect of AcSDKP and AcADKP on human dermal fibroblasts (AG01523c) and keratinocytes (HaCaT) was tested by the MTT assay (in which commercially available AcSDKP was used in parallel, for comparison reasons). Also, was tested the effect of AcSDKP and AcADKP on the proliferation of dermal fibroblasts, by quantifying the DNA synthesis with the tritiated thymidine assay. The effect of AcSDKP and AcADKP tetrapeptides on fibroblast’s collagen synthesis was also investigated, as well as, their effect on keratinocyte migration, using the scratch assay. The above in vitro cell assays did not show any statistically significant effect of the AcSDKP and AcADKP tetrapeptides on the biological parameters studied, at least under the experimental conditions used. However, the experimental results of the present study show that both AcSDKP and AcADKP can be prepared in the laboratory in sufficient quantities (mg amounts) and in high purity, following specific synthesis and purification protocols. These peptides are non toxic (at concentrations of 10-10 to 10-7 M) when incubated with dermal fibroblasts or keratinocytes. Thus, the above peptides may be prepared in the laboratory and used in a series of specially selected biological assays, in order to further study their effect on dermal wound healing.
Main subject category:
Science
Other subject categories:
Pharmaceutical technology
Keywords:
peptides, dermal wound healing, thymosins, β-thymosins, acSDKP, acADKP, Solid Phase Peptide Synthesis, keratinocytes, skin fibroblasts
