Influence of Dipeptidyl Peptidase IV Deficiency on the Immune Response in the Process of Wound Healing in Mice

Uvod: Dipeptidil-peptidaza IV odnosno molekula CD26 (DPP IV/CD26) ubikvitarna je proteaza i kostimulacijska molekula s važnom ulogom u fiziološkim i patološkim procesima. Dosadašnja istraživanja pokazuju da DPP IV/CD26 sudjeluje u procesima angiogeneze, adhezije, migracije te apoptoze stanica koji predstavljaju ključne procese u cijeljenju ranjenog tkiva. Međutim, uloga DPP IV/CD26 još uvijek nije dovoljno istražena. Pretpostavka ovog istraživanja je da DPP IV/CD26 ima značajnu ulogu u procesu cijeljenja rane te aktivaciji imunosnog odgovora u ranjenom tkivu kože, obzirom da je aktivnost mnogih biološki važnih molekula uključenih u upalne i imunosne mehanizme pod utjecajem DPP IV/CD26. Cilj istraživanja: Cilj ovog istraživanja bio je ispitati utjecaj nedostatka DPP IV/CD26 na makroskopske i mikroskopske promjene u procesu cijeljenja rane te na izražaj ciljnih stanica imunosnog sustava (limfocita T i makrofaga) tijekom navedenog procesa u definiranim vremenskim intervalima. Također, cilj nam je bio ispitati postoje li promjene u aktivnosti DPP IV/CD26 u serumu divljeg tipa miša tijekom procesa cijeljenja ranjenog tkiva. Materijal i metode: Pokusni model cijeljenja rana uspostavljen je u divljem tipu (C57BL/6) i CD26 deficijentnim (CD26-/-) miševima. Pokusne životinje su žrtvovane drugog, četvrtog, sedmog, desetog i petnaestog dana od induciranja rana. Proces cijeljenja rana praćen je makroskopskim i mikroskopskim analizama. Učinjena su histomorfometrijska, patohistološka i imunohistokemijska istraživanja. Određivao se stupanj obnavljanja koriuma te broj limfocita T i makrofaga. Serumska aktivnost DPP IV/CD26 kod divljeg tipa miša tijekom procesa cijeljenja rana određivana je spektrofotometrijski. Rezultati: Rezultati dobiveni ovim istraživanjem ukazuju na različitu dinamiku i intenzitet cijeljenja rane u uvjetima nedostatka DPP IV/CD26. Stupanj regeneracije koriuma bio je značajno veći kod CD26-/- životinja sedmog dana cijeljenja rana. Utvrđena je značajna razlika u izražaju limfocita T između ispitanih sojeva miševa desetog i petnaestog dana nakon induciranja rana, kada je uočen značajno veći broj limfocita T kod C57BL/6 miševa. Broj makrofaga četvrtog dana cijeljenja rana bio je gotovo dvostruko viši u uvjetima nedostatka DPP IV/CD26. Enzimska aktivnost serumske DPP IV/CD26 u divljem tipu životinja značajno je snižena četvrtog dana cijeljenja rana u usporedbi s fiziološkim uvjetima. Zaključak: Rezultati ovog istraživanja potvrđuju ulogu DPP IV/CD26 u procesu cijeljenja ranjenog tkiva. Potvrđena je pretpostavka o utjecaju DPP IV/CD26 na imunosni odgovor tijekom procesa cijeljenja rana, ali su potrebna daljnja istraživanja kako bi se utvrdili točni mehanizmi kojima molekula DPP IV/CD26 regulira ključne korake uključene u ove procese.Introduction: Dipeptidyl peptidase IV or CD26 molecule (DPP IV/CD26) is an ubiquitous protease and costimulatory molecule with an important role in physiological and pathological processes. It has been shown that DPP IV/CD26 is involved in the process of angiogenesis, cell adhesion, migration, and apoptosis, which are key events in healing of wounded tissues. However, the role of DPP IV/CD26 still remains insufficiently understood. The hypothesis of this research is that DPP IV/CD26 plays an important role in wound healing and in the activation of the immune response in injured skin tissue, since it regulates the activity of many biologically important molecules involved in the inflammatory and immune mechanisms. Aim: The aim of this study was to investigate the influence of DPP IV/CD26 on macroscopic and microscopic changes in the process of wound healing as well as on the expression of target cells of the immune system (T lymphocytes and macrophages) during this process at defined time intervals. Likewise, our aim was to examine changes in the activity of DPP IV/CD26 in the serum of wild-type mice during healing of the wounded tissue. Materials and Methods: An experimental model of wound healing was established in wild-type (C57BL/6) and CD26 deficient (CD26-/-) mice. Experimental animals were sacrificed the second, fourth, seventh, tenth and fifteenth day after wounding. The process of wound healing was monitored by macroscopic and microscopic analyzes. Histomorphometric, histopathological and immunohistochemical analyzes were done. The degree of the recovery of the corium and the number of T-lymphocytes and macrophages was determined. Serum DPP IV/CD26 activity in wild-type mice during the process of wound healing was determined spectrophotometrically. Results: Results obtained in this study indicate different dynamics and intensity of wound healing in conditions of DPP IV/CD26 deficiency. The rate of regeneration of the corium was significantly higher in CD26-/- animals the seventh day of wound healing. A significant difference in the expression of T lymphocytes was noticed between analyzed mice strains the tenth and fifteenth day after wounding, when a significantly higher number of T lymphocytes was determined in C57BL/6 animals. The number of macrophages on the fourth day of wound healing was almost twice as high in conditions of DPP IV/CD26 deficiency. Serum DPP IV/CD26 activity in wild-type animals was significantly lowered on the fourth day post-wounding compared to physiological conditions. Conclusion: Results of this study confirm the role of DPP IV/CD26 in the process of healing of the wounded tissue. The hypothesis about the influence of DPP IV/CD26 on the immune response during the process of wound healing is confirmed, but further research is needed to determine the exact mechanisms by which DPP IV/CD26 regulates key steps involved in these processes