Background: For several antibiotics an immunomodulatory effect has been described. Although
fungal infections play a central role in severely ill and immunosuppressed patients, immunomodulatory
effects of antifungals have not been sufficiently investigated. The present study sets out to establish
an in-vitro human whole blood aspergillosis model and to assess the effect of different antimycotics on
cytokine levels in blood stimulated with mold.
Material/methods: Establishment of an in-vitro aspergillosis model: Aspergillus fumigatus
ATCC204305 was used to develop inflammation in whole blood obtained from 6 healthy volunteers invitro.
Different concentrations of conidia (106/mL, 107/mL and 108/mL) as well as hyphal fragments
(autoclaved/inactivated and alive) were investigated for induction of cytokine levels quantified by
commercially available ELISA.
Immunomodulation by antimycotics: Based on the described experiments autoclaved hyphal
suspension (retrieved from a prior conidia count of 5mL x 107 conidia/mL cultured 2-4 days on
Sabouraud Dextrose agar) demonstrated the most potent cytokine release and was used for the main
part of the study.
Hyphal suspension or saline solution as control was added to whole blood from volunteers (n=5).
Experiments were performed at concentrations according to clinically achieved maximal serum
concentrations for conventional and liposomal amphotericin B (both final concentration 5mg/L),
fluconazole (20mg/L), voriconazol (5mg/L), posaconazol (2mg/L) or control. All samples were
incubated at 37°C over 4hours at a slow rotation rate imitating physiologic conditions. At predefined
time points selected samples were centrifuged to obtain serum for determination of concentration-time
profiles of IL6, IL8 and TNF-alpha.
Results: For all investigated samples cytokine levels were higher after 4 than after 2hours, indicating
that the experiment captured the dynamic phase of the stimulation. Compared to baseline cytokine
levels increased by addition of hyphal suspension over 4hours approx. 60-fold for IL-6, 1500-fold for
IL-8 and 200-fold for TNF-alpha. While conventional amphotericin B further increased the cytokine
levels, especially the release of IL-6, in our in-vitro aspergillosis model, this was not the case for its
liposomal formulation. Fluconazole and to a smaller extend the other two azoles reduced the cytokine
increase compared to samples without antifungal agent. Whether the compared to the two other
azoles higher potency of fluconazole may be ascribed to stronger intrinsic activity or to the in analogy
to the clinically observed higher plasma values higher simulated concentrations in-vitro remains to be
Conclusions: We were able to successfully establish an in-vitro aspergillus inflammation model. Our
data indicate significant differences in the immunomodulatory potency of different classes of
antimycotics. In our aspergillosis model fluconazole had the highest anti-inflammatory potential while
conventional amphotericin even increased cytokine release. This preliminary information might have
clinical implication, since cytokine dysregulation plays a major role in the pathogenesis and outcome of
fungal infections. Clinical studies are warranted to confirm our findings.
Full conference title:
- ECCMID 27th (2017)