Immunocytochemical detection of cytokines and chemokines in Langerhans cells and in vitro derived dendritic cells.
We have developed a direct immunocytochemical technique to identify cytokine and chemokine production in epidermal Langerhans cells (LC) and in vitro derived CD14-, CD1a+, CD83+, CD40+ dendritic cells (DC) at the single cell level. Formaldehyde fixation combined with saponin permeabilization preserved cellular morphology and generated a characteristic juxtanuclear staining signal due to the accumulation of cytokine to the Golgi organelle. This approach was used for the assessment of TNF-alpha, IL-6, IL-8, IL-10, IL-12, GM-CSF, MIP-1alpha, MIP-1beta and RANTES producing cells. In contrast, a diffuse cytoplasmic staining was evident for IL-1ra, IL-1alpha and IL-1beta production. IL-1ra and IL-1alpha were expressed in 10-25% of unstimulated cultured cells, while all the other cytokines were undetectable. IL-1ra, IL-1alpha and IL-1beta were also the dominating cytokines, expressed in up to 85% of the DC, after 3 h of LPS stimulation. A significantly lower number of cells (0-5%) synthesized TNF-alpha, IL-6, IL-10, IL-12 and GM-CSF. The incidence of chemokine producing cells (IL-8, RANTES, MIP-1alpha, MIP-1beta) peaked 10 h after LPS stimulation in up to 60% of the DC. Both immature CD83- and mature CD83+ DC as well as LC had a similar cytokine production pattern. Thus, in comparison to monocytes, LPS stimulation of DC generated a lower incidence of TNF-alpha, IL-6, IL-10 and IL-12 producing cells while IL-1 was expressed in a comparable number of cells.
Loré
K
K
Department of Immunology, Microbiology, Pathology and Infectious Diseases, Karolinska Institute, Huddinge University Hospital, Sweden. knlo@labd01.hs.sll.se
Sönnerborg
A
A
Spetz
A L
AL
Andersson
U
U
Andersson
J
J
D018925:Chemokines / Q000096:biosynthesis*; D016207:Cytokines / Q000096:biosynthesis*; D003713:Dendritic Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D007801:Langerhans Cells / Q000166:cytology / Q000378:metabolism*
D018925:Chemokines; D016207:Cytokines