Terapia prechirurgica della fibromatosi uterina - FedOA - Università ...

polyglactic acid sutures (Vicryl; Ethicon, Somerville, NJ). Vasoactive drugs were not used. Surgeons
were blinded to the pre-surgical medical treatment.
For each patient, we evaluated the myoma diameter, the total operating time (from skin incision to
closure) and the intraoperative blood loss. The blood loss was estimated by weighing swabs and
adding the estimated volume of blood to that removed by suction. The volume of lactated Ringer’s
solution used to irrigate the pelvis was estimated precisely and then subtracted from the fluid collected
in the suction unit. The amount of blood loss during abdominal wall opening was not calculated.
At the end of the intervention, surgeons were asked to state if they promptly identified the cleavage
planes between the leiomyoma and the surrounding myometrium or not.
In addition to the patients enrolled in the study, we also considered paraffin embedded samples
obtained in a retrospective manner from twenty premenopausal patients (ten treated with GnRH-a and
ten untreated patients) submitted to abdominal hysterectomy due to uterine fibromatosis. These
patients were randomly selected from the database of our Pathology Unit, in order to evaluate the
features of the border between myometrium and leiomyoma using an hematoxylin and eosin staining.
For each leiomyoma, three paraffin blocks of representative areas were selected and 4 μm thick serial
sections were cut from each block. One section was stained with hematoxylin and eosin to confirm the
original diagnosis, excluding the areas of necrosis.
Sections from leiomyomas were pre-treated with a heat-induced antigen retrieval technique, with
incubation in a 650W microwave oven (three sequential steps of 4 min each in citrate buffer (pH 6.0)
10 mM) for antigen unmasking and then incubated for 20 min at room temperature with 0.3%
hydrogen peroxide in methanol, to quench the endogenous peroxidases.
Further incubation with non-immune mouse serum (1:20, Dakopatts, Hamburg, Germany) diluted in
PBS-bovine serum albumin (1%) for 25 min was performed to prevent non-specific immunostaining.
After three washing with TRIS saline buffer, immunostaining was performed for the study of the
expression of PCNA using an anti-PCNA primary antibody (PC10, monoclonal, 1:200, DBA- Milan,
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