(947.10) GAL3 Excretion Regulates Smooth Muscle Cell Survival and Proliferation

Poster Board Number: E479

Stephen Haigh (Augusta Univeristy), Xi Li (Stanford University), Zsuzsanna Bordan (Augusta Univeristy), Hunter Sellers (Augusta Univeristy), Mary Louise Meadows (Augusta Univeristy), Scott Barman (Augusta Univeristy), David Fulton (Augusta Univeristy)

Pulmonary arterial hypertension (PAH) is a complex and fatal disorder characterized by an unrelenting increase in pulmonary arterial pressure.  Excessive proliferation of pulmonary artery smooth muscle cells (PASMC) leads to increased vascular resistance and eventually right ventricular failure and death.  Our laboratory has identified a key role for Galectin-3 (GAL-3) in PAH. However, the mechanisms by which GAL-3 alters PASMC behavior remains poorly understood.  GAL-3 contains a functional BH1 like domain that has an NWGR sequence with high homology to the anti-apoptotic protein, BCL2, and this domain has previously been shown to be functional in cancer cell lines. Our hypothesis is GAL-3 upregulation plays an important role in driving PASMC proliferation through repression of apoptosis via this NWGR domain.  Using adenoviral vector encoding the GAL-3 G182A mutation and isolated GAL-3 PASMCs from a GAL3 KO rat (RPASMC).  Our in vitro studies show that overexpression of GAL-3 G182A causes a decrease in viability upon treatment overnight with 0.5% serum containing media(plt;.001) compared to normal media (p=.840).  When exposed to apoptotic stimuli (TNFα, Cycloheximide) GAL-3 G182A overexpressing GAL-3 KO RPASMCs had significantly (plt;.001) decreased vitality.  Interestingly however, this domain also seems to regulate non-apoptotic related events such as cell attachment. Our previous work had shown that GAL-3 is readily excreted protein, and co-culture experiments as well as recombinant protein supplementation show that soluble GAL-3 is biologically active and induces SMC growth and proliferation(Plt;.05). Further experimentation also shows that inhibition of GAL-3 using blocking antibodies prevents SMC proliferation. (plt;.05) To test this hypothesis in-vivo we injected GAL-3 KO mice with a liver specific AAV overexpressing GAL-3, or GFP (control) and exposed them to sugen/hypoxia to induce PAH.  Surprisingly our initial findings indicate a slight protection with liver GAL-3 overexpression when compared to wt mice.(plt;.05) Also very interestingly, while liver overexpression of GAL-3 was very high there was no evidence of GAL3 in the lungs or plasma of GAL3KO mice indicating it may not be secreted from the liver in sufficient amounts unlike what was observed in other cell types. Overall our data suggests that the NWGR domain of GAL-3 is an important regulator of PASMC survivability and the blocking antibody suggests that secreted GAL-3 is capable of regulating SMC proliferation, however it does not appear that hepatocytes secrete sufficient GAL-3 for it to circulate and play a role in the lung vasculature.

Support or Funding Information

F31HL147428
R01HL125926-03

lt;pgt;F31HL147428lt;brgt; R01HL125926-03lt;/pgt;