Question:

The tyrosine phosphatase SHP-2 plays an important role in growth factor signalling. Here we studied whether SHP-2 influences hypoxia induced endothelial cell proliferation and HIF-1α signalling.

Methods:

Overexpression of wild type (SHP-2 WT), catalytically inactive (SHP-2 CS) or constitutively active (SHP-2 E76A) SHP-2 in human microvascular endothelial cells (HMEC) was achieved by lentiviral transduction. Cells were exposed to hypoxia for 4h or 24h. Proliferation was assessed by MTT reduction. HIF-1α levels and ERK activity (Thr/Tyr-phosphorylation) were detected by western blot.

Results:

Compared to SHP-2 WT, expression of constitutively active SHP-2 enhanced proliferation both during normoxia (1.7-fold) and hypoxia (1.5-fold) (both p<0.05, n=8). Hypoxia inducible factor 1α (HIF-1α) protein levels (p<0.05, n=4) as well as the activity of the potential HIF-1α regulator MAPK ERK1/2 (n=3) were also enhanced. Importantly, the increased hypoxic proliferation was not only completely blocked upon HIF-1α (Echinomycin 10ng/ml, p<0.05, n=6) inhibition but also upon treatment with a MAPK-pathway inhibitor (GW4057, p<0.05, n=6). In contrast, expression of catalytically inactive SHP-2 impaired proliferation during normoxia (p<0.05, n=7) and hypoxia (p<0.05, n=6) compared to SHP-2 WT. In addition, hypoxic HIF-1α protein accumulation (p<0.05, n=4) and ERK1/2 activity (n=3) were reduced. However, the reduction in HIF-1α protein was rescued by treatment with proteasomal inhibitors (MG132 or Epoximicin, n=3).

Conclusions:

SHP-2 is important for both normoxic and hypoxic endothelial proliferation. During hypoxia SHP-2 influences endothelial cell proliferation through HIF-1α protein stabilisation, possibly by regulating ERK activity. Thus, induction of SHP-2 catalytic activity may be promising for angiogenesis induction in ischemic conditions.