Deficiency of plasminogen activator inhibitor‐2 results in accelerated tumor growth

BackgroundUpregulation of the plasminogen activation system, including urokinase plasminogen activator (uPA), has been observed in many malignancies, suggesting that co‐opting the PA system is a common method by which tumor cells accomplish extracellular matrix proteolysis. PAI‐2, a serine protease inhibitor, produced from the SERPINB2 gene, inhibits circulating and extracellular matrix‐tethered uPA. Decreased SERPINB2 expression has been associated with increased tumor invasiveness and metastasis for several types of cancer. PAI‐2 deficiency has not been reported in humans and PAI‐2‐deficient (SerpinB2−/−) mice exhibit no apparent abnormalities.ObjectivesWe investigated the role of PAI‐2 deficiency on tumor growth and metastasis.MethodsTo explore the long‐term impact of PAI‐2 deficiency, a cohort of SerpinB2−/− mice were aged to >18 months, with spontaneous malignancies observed in 4/9 animals, all of apparently vascular origin. To further investigate the role of PAI‐2 deficiency in malignancy, SerpinB2−/− and wild‐type control mice were injected with either B16 melanoma or Lewis lung carcinoma tumor cells, with markedly accelerated tumor growth observed in SerpinB2−/− mice for both cell lines. To determine the relative contributions of PAI‐2 from hematopoietic or nonhematopoietically derived sources, bone marrow transplants between wild‐type C57BL/6J and SerpinB2−/− mice were performed.Results and ConclusionsOur results suggest that PAI‐2 deficiency increases susceptibility to spontaneous tumorigenesis in the mouse, and demonstrate that SerpinB2 expression derived from a nonhematopoietic compartment is a key host factor in the regulation of tumor growth in both the B16 melanoma and Lewis lung carcinoma models.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/163438/2/jth15054_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163438/1/jth15054.pd

The endothelial-specific regulatory mutation, Mvwf1, is a common mouse founder allele

Mvwf1 is a cis-regulatory mutation previously identified in the RIIIS/J mouse strain that causes a unique tissue-specific switch in the expression of an N-acetylgalactosaminyltransferase, B4GALNT2, from intestinal epithelium to vascular endothelium. Vascular B4galnt2 expression results in aberrant glycosylation of von Willebrand Factor (VWF) and accelerated VWF clearance from plasma. We now report that 13 inbred mouse strains share the Mvwf1 tissue-specific switch and low VWF phenotype, including five wild-derived strains. Genomic sequencing identified a highly conserved 97-kb Mvwf1 haplotype block shared by these strains that encompasses a 30-kb region of high nucleotide sequence divergence from C57BL6/J flanking B4galnt2 exon 1. The analysis of a series of bacterial artificial chromosome (BAC) transgenes containing B4galnt2 derived from the RIIIS/J or C57BL6/J inbred mouse strains demonstrates that the corresponding sequences are sufficient to confer the vessel (RIIIS/J) or intestine (C57BL6/J)-specific expression patterns. Taken together, our data suggest that the region responsible for the Mvwf1 regulatory switch lies within an approximately 30-kb genomic interval upstream of the B4galnt2 gene. The observation that Mvwf1 is present in multiple wild-derived strains suggests that this locus may be retained in wild mouse populations due to positive selection. Similar selective pressures could contribute to the high prevalence of von Willebrand disease in humans

Genetic Modifiers of Hemostasis.

Factor V serves as the nonenzymatic cofactor for Factor Xa to produce thrombin. A Factor V variant called Factor V Leiden (FVL) is the most common known inherited risk factor for venous thrombosis, though incomplete penetrance is observed in people inheriting FVL. FVL is resistant to degradation by Activated Protein C (APC), resulting in increased thrombin. In vitro studies suggest that wildtype FV can also serve as a cofactor for APC in the inactivation of Factor VIIIa. We used mouse models of FV and FVL to demonstrate the in vivo relevance of the wildtype FV APC cofactor activity on thrombosis. Additionally, we used a synthetic lethal genetic interaction between FVL homozygosity (FVQ/Q) and heterozygous tissue factor pathway inhibitor deficiency (TFPI+/-) in mice as a phenotype in a sensitized ENU mutagenesis screen to identify dominant modifier genes contributing to FVL penetrance. As conceptual proof, it was demonstrated that: [1] loss of one tissue factor allele or [2] a suite of dominantly acting genes in the DBA mouse strain suppress the lethal phenotype. In the mutagenesis screen, analysis of 6677 G1 offspring identified 92 FVQ/Q TFPI+/- mice surviving to weaning, with 13 exhibiting successful transmission of a putative suppressor mutation. These findings have important implications for the genetic regulation of hemostatic balance. While characterizing gene-targeted mice deficient in SerpinB2, we observed mice which were small at birth (sml). Sml was initially attributed to SerpinB2 deficiency. However, analysis of SerpinB2 deficient mice derived from two additional, independent ES cell clones revealed no growth abnormalities. Using recombination observed between sml and the SerpinB2 locus, sml was mapped to a 2.78 Mb interval ~12 cM (30 Mb) proximal to SerpinB2. Sequencing of candidate genes revealed a nonsense mutation in insulin receptor substrate 1, Irs1. Analysis of ES cell DNA demonstrated that the Irs1 mutation arose spontaneously in the original cell culture used for gene targeting. This study suggests that new mutations arising during ES cell culture and linked to the targeted allele could result in incorrect assignment of phenotype and may account for a subset of discordant results for experiments independently targeting the same gene.Ph.D.Human GeneticsUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/62388/1/westricr_1.pd