Influence of enamel matrix derivative on primary CD4+ T-helper lymphocyte migration, CD25 activation, and apoptosis.
BACKGROUND
Enamel matrix derivative (EMD) has a low immunogenic potential. To the best of our knowledge, there are no studies on the influence of EMD on lymphocyte migration as a sensitive cellular reaction parameter. This study investigated the influence of EMD on primary T-lymphocyte migration, CD25 activation, and activation-induced cell death.


METHODS
After immunomagnetic-positive CD4+ lymphocyte separation from peripheral blood taken from three healthy volunteers per trial, the influence of EMD on cell locomotion was assessed in a three-dimensional collagen matrix migration model (CMMM). Direct CD4+ cell contact with EMD at concentrations of 25 and 100 microg/ml was mediated in a one-phase CMMM. We investigated the indirect influence of EMD in a two-phase CMMM: one collagen phase contained 25 and 100 microg EMD/ml, using the same concentrations, and a second adjacent phase contained T lymphocytes. After time-lapse videomicroscopy, the mean locomoting percentage of 30 randomly selected cells was analyzed. Using flow cytometry, CD25 receptor activation was assessed, and annexin V was used for apoptosis detection in lymphocytes challenged with 0, 1, 25, 50, and 100 microg EMD/ml.


RESULTS
The one-phase CMMM revealed a reduction and the two-phase CMMM showed a dose-dependent increase in the mean locomoting cell percentage (P <0.001). Increasing EMD concentrations resulted in dose-dependent enhanced T-cell CD25 receptor expression and in increasing apoptosis (P <0.001).


CONCLUSIONS
Our study showed immediate effects of EMD on primary CD4+ lymphocyte migration, CD25 activation, and apoptosis. CD4+ lymphocyte apoptosis may be a further possible background for uneventful early wound healing as seen clinically as the result of EMD application.
Gassmann
Georg
G
Department of Periodontology, Faculty of Dental Medicine, University of Witten/Herdecke, Witten, Germany. gassmann@uni-wh.de
Schwenk
Bettina
B
Entschladen
Frank
F
Grimm
Wolf-Dieter
WD
D000328:Adult; D017304:Annexin A5; D017209:Apoptosis / Q000187:drug effects*; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects; D002634:Chemotaxis, Leukocyte / Q000187:drug effects*; D003094:Collagen; D003746:Dental Enamel Proteins / Q000494:pharmacology*; D005260:Female; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives; D005456:Fluorescent Dyes; D006801:Humans; D007155:Immunologic Factors / Q000494:pharmacology*; D053645:Interleukin-2 Receptor alpha Subunit / Q000187:drug effects*; D008297:Male; D018715:Microscopy, Video; D008875:Middle Aged; D006377:T-Lymphocytes, Helper-Inducer / Q000187:drug effects*; D055815:Young Adult
D017304:Annexin A5; D003746:Dental Enamel Proteins; C492760:FITC-annexin A5; D005456:Fluorescent Dyes; C508017:IL2RA protein, human; D007155:Immunologic Factors; D053645:Interleukin-2 Receptor alpha Subunit; C109069:enamel matrix proteins; D003094:Collagen; D016650:Fluorescein-5-isothiocyanate