Dermal fibroblasts were cultured from the skin tissue as
previously described [27]. Briefly, dermal specimens were
minced into small pieces of less than 0.5 mm in any dimension
in Dulbecco’s modified Eagle’s medium (DMEM) (Sigma–
Aldrich, St. Louis, MO) with 10% fetal bovine serum (FBS)
(Sigma–Aldrich, St. Louis, MO). The tissue fragments were then
washed with medium and distributed into 60 mm 15 mm
Petri dishes. A sterile glass coverslip attached to the dish with a
drop of sterile silicone lubricant was used to immobilize the
tissue fragments. 3 mL of DMEM containing 10% FBS was added
to each dish and they were incubated at 37 8C in an atmosphere
of 5% CO2 and 95% relative humidity. The medium was replaced
every 5 days. After 4 weeks, the fibroblasts were released from
the dishes after treatment with trypsin (0.25%, w/v), transferred
to 75 cm2 culture flasks, and sub-cultured for experiments.
Dermal fibroblasts were cultured from the skin tissue aspreviously described [27]. Briefly, dermal specimens wereminced into small pieces of less than 0.5 mm in any dimensionin Dulbecco’s modified Eagle’s medium (DMEM) (Sigma–Aldrich, St. Louis, MO) with 10% fetal bovine serum (FBS)(Sigma–Aldrich, St. Louis, MO). The tissue fragments were thenwashed with medium and distributed into 60 mm 15 mmPetri dishes. A sterile glass coverslip attached to the dish with adrop of sterile silicone lubricant was used to immobilize thetissue fragments. 3 mL of DMEM containing 10% FBS was addedto each dish and they were incubated at 37 8C in an atmosphereof 5% CO2 and 95% relative humidity. The medium was replacedevery 5 days. After 4 weeks, the fibroblasts were released fromthe dishes after treatment with trypsin (0.25%, w/v), transferredto 75 cm2 culture flasks, and sub-cultured for experiments.
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