Mice with a severe deficiency in protein C display prothrombotic and proinflammatory phenotypes and compromised maternal reproductive capabilities
Angelina J. Lay, Zhong Liang, Elliot D. Rosen, Francis J. Castellino
Angelina J. Lay, Zhong Liang, Elliot D. Rosen, Francis J. Castellino
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Research Article Hematology

Mice with a severe deficiency in protein C display prothrombotic and proinflammatory phenotypes and compromised maternal reproductive capabilities

Abstract

Anticoagulant protein C (PC) is important not only for maintenance of normal hemostasis, but also for regulating the host immune response during inflammation. Because mice with a designed total genetic deficiency in PC (PC–/– mice) die soon after birth, attempts to dissect PC function in various coagulation/inflammation-based pathologies through use of mice with less than 50% of normal PC levels have not been successful to date. In the current investigation, we have used a novel transgenic strategy to generate different mouse models expressing 1–18% of normal PC levels. In contrast to PC–/– mice, mice with only partial PC deficiency survived beyond birth and also developed thrombosis and inflammation. The onset and severity of these phenotypes vary significantly and are strongly dependent on plasma PC levels. Our findings additionally provide the first evidence that maternal PC is vital for sustaining pregnancy beyond 7.5 days postcoitum, likely by regulating the balance of coagulation and inflammation during trophoblast invasion. These low PC–expressing transgenic mouse lines provide novel animal models that can be used to elucidate the importance of PC in maintenance of the organism and in disease.

Authors

Angelina J. Lay, Zhong Liang, Elliot D. Rosen, Francis J. Castellino

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Figure 1

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Generation of low-PC transgenic mice. (A) Schematic diagram showing rele...
Generation of low-PC transgenic mice. (A) Schematic diagram showing relevant features of the low PC DNA construct. A 12.5-kb fragment of an inactivated (by partial promoter deletion FVII gene and an 18-kb fragment of an inactive (by exon 1 deletion) FX gene were cloned upstream and downstream, respectively, of the PC cDNA/polyA sequences. Expression of PC was driven by the FX promoter contained within the complete intergenic region (IGR) of the FVII-FX chromosomal segment. (B) Low-PC potential founders were identified by PCR analysis. The sense and antisense primers spanned exons 4 and 6 of the PC gene, respectively, amplifying a 497-bp fragment for the WT allele and a 284-bp fragment for the mPC cDNA that is derived from the transgene. (C) Low-PC transgenic mice from various lines were evaluated by Southern blot analysis of KpnI-digested genomic DNA. The Southern probe in A hybridized to a 2.7-kb fragment of the WT FVII gene in addition to various PC transgene fragments (asterisks), the sizes of which are dependent on sites of PC transgene integration. The 3.7-kb band shows that at least 1 tandem repeat of the transgene has occurred in each mouse line.