Topical tacrolimus in atopic dermatitis: Effects of ... - Helda - Helsinki.fi

Staphylococcus aureus colonization
Assessments of staphylococcal colonization took place at baseline, weeks one and 2,
and monthly thereafter. Agar plate lifts from the worst AD lesion of each patient for
bacterial culture came from application of a contact plate (NUNC, Roskilde, Denmark)
for 5 seconds. Cultures were incubated aerobically in a cystine-, lactose-, and
electrolyte-deficient agar medium at 35°C for 24 hours. The following criteria were
used to identify S. aureus: typical colony morphology on the contact plates and in pure
cultures on a blood agar plate, a positive catalase test result, positive DNAse test result,
or a positive coagulase tube test result. The density of S. aureus colonies was expressed
as colony-forming units per square centimeter (CFU/cm 2 ). Changes in CFU/cm 2 were
calculated from baseline to week one, month 6, and month 12 for each patient.
Skin collagen synthesis and skin thickness
Measurements of the tacrolimus patients and controls treated with conventional
treatment were done at baseline and after 12 months of treatment. Healthy subjects
underwent the measurement only at baseline. Measurements of the control groups were
performed during the same time period as in the primary study. Collagen synthesis was
assessed by measuring the level of aminoterminal propeptides of procollagen types I
and III (PINP and PIIINP, respectively) from suction blister fluid samples of abdominal
skin. The blisters were raised by a disposable suction blister device (Ventipress Oy,
Lappeenranta, Finland) that draws interstitial fluid between the epidermis and dermis
(Kiistala 1968). The fluid was aspirated from the blisters after approximately one hour
of suction at -100 to -300 mmHg and frozen at -20 degrees of Celsius. Concentrations
of PINP and PIIINP were determined by specific radioimmunoassays (Orion
Diagnostica, Oulunsalo, Finland) (Risteli et al. 1988, Melkko et al. 1990, Oikarinen et
al. 1992).
Skin thickness, defined as the distance between the surface of the stratum
corneum and the lower dermis, was measured digitally with a high-frequency
ultrasound device (DUB20-S, Taberna pro medicum, L�neburg, Germany) (Tan et al.
1982). As skin thickness differs markedly for different areas of the body, the
measurements were made in eight target regions: forehead, cheek, collarbone, thorax,
upper arm, inside elbow, forearm, and the abdomen 2 cm left of the navel,
independently of whether the skin was affected by AD or not. Measurements were taken
at a frequency of 30 MHz, and results were calculated as the median thickness of the
eight sites. Initially the plan was to measure skin thickness in all patients, but technical
problems with the ultrasound device prevented a complete analysis, and participation or
non-participation in skin thickness measurement was random.
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