Acute myeloid leukemia and
head and neck squamous cell carcinomas are the major causes of mortality and morbidity in
Fanconi anemia (FA) patients.
Matrix metalloproteinases (
MMPs), particularly MMP-2 and MMP-9, have been implicated in
tumor invasion and
metastasis. Various
cytokines,
mitogens,
growth factors, inducers and inhibitors control
MMP activities. We investigated the roles of these in the regulation of MMP-2 and MMP-9 in human immortalized fibroblasts from FA. Human FA immortalized fibroblast cell lines FA-A:PD220 and FA-D2:PD20 were grown in minimum essential medium (MEM) supplemented with 15%
fetal bovine serum (FBS) and
antibiotics in 24-well tissue culture plates. At near confluence, the cells were washed with phosphate‑buffered saline (PBS) and incubated in
serum-free media with the following:
phorbol 12-myristate 13-acetate (PMA)
at 10-100 ng/ml;
tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) at 0.1-25 ng/ml;
lipopolysaccharide (LPS)
at 10-100 µg/ml;
epigallocatechin gallate (EGCG) and
doxycycline (Dox) at 10-100 µM without and with PMA; a nutrient mixture (NM) without and with PMA
at 10-1,000 µg/ml;
actinomycin-D and cyclohexamide at 2 and 4 µM;
retinoic acid and
dexamethasone at 50 µM. After 24 h, media were removed and analyzed for MMP-2 and MMP-9 by zymography. Both FA cell lines expressed only MMP-2 and responded similarly to
cytokines,
mitogens, inducers and inhibitors. PMA potently stimulated MMP-9 and had a moderate effect on MMP-2. TNF-α showed variable effects on MMP-2 and significantly enhanced MMP-9. IL-1β enhanced MMP-2 slightly and MMP-9 significantly. LPS had a moderate stimulatory effect on MMP-2 and no effect on MMP-9. EGCG, Dox and NM, without and with PMA, downregulated MMP-2 and MMP-9 expression.
Actinomycin-D,
retinoic acid and
dexamethasone also had inhibitory effects on MMP-2. Our results showed that
cytokines,
mitogens and inhibitors modulated FA fibroblast MMP-2 and MMP-9 expression, suggesting the clinical use of
MMP inhibitors, particularly such potent and non-toxic ones as the NM and its component EGCG in the management of FA
cancers.