Pregnancy Alters Platelet Counts and Enhances Platelet Reactivity in Mice
Authors: Hobbs Linzi M , Brake Marisa A , Redshaw Dakota R , Barbarich Lauren C , Hobbs Kelsey A , Rodionov Roman N , Westrick Randal J 
Published: November 2020

Background: Pregnancy is a major risk factor for venous thromboembolism (VTE) and is the leading cause of maternal death during pregnancy and the immediate post-partum period. Although plasma levels of clotting factors and fibrinolysis inhibitors are known to increase during pregnancy peaking during the third trimester as parturition nears, little is known about whether platelets are affected in response to pregnancy. Platelets are derived from megakaryocytes and have a circulating half-life of 5-10 days. They are essential for blood coagulation, delivering procoagulant molecules directly to sites of vascular injury. Platelets contain a spliceosome that processes pre-mRNA, providing them with the necessary machinery to respond to external stimuli. Thus, platelets can be affected by diseases such as cancer, which can alter platelet mRNA profiles and function. The plasticity of platelets in these contexts suggests that they could also change in response to pregnancy as an adaptation to preventing blood loss during parturition. However, little is known about the transcriptional and/or functional changes that could occur in platelets during the course of pregnancy.

Aims: We aimed to analyze mouse platelet function at early, middle, late gestation, and postpartum timepoints in order to identify pregnancy related functional changes.

Methods: C57BL/6J female mice were mated with C57BL/6J male mice to generate pregnant mice. Pregnancy was confirmed by the presence of a vaginal plug, which was used to estimate embryonic day 0.5. Pregnant mice were randomly assigned to one of four groups based on gestational stages corresponding to human trimesters and postpartum. The stages are early (days 7-9), middle (days 11-14), late (days 16-20), and postpartum (days 22-24). The maximum volume of mouse whole blood was drawn into a 1:9 volume of buffered sodium citrate via cardiac puncture. A 20ul aliquot was used for complete blood counts using the Advia 2120 with settings optimized for C57BL/6 mouse blood. The remaining blood was reserved for platelet aggregation studies using the Roche Multiplate Aggregometer. ADP and type 1 collagen were used as the aggregating agents with the operator blinded to the gestational groups. Studies were initiated at the same time of day in order to control for the circadian rhythm and the gestational age.

Results: The maximum blood volume we were able to draw from each mouse was significantly increased for all groups (early, middle and late gestational stages and postpartum) compared to the non-pregnant control group (p ≤ 0.03, n ≥ 9). Platelet counts (per microliter of blood) were statistically unchanged among the control and the three gestational groups. However, postpartum platelet counts were significantly increased relative to all groups (control, early, middle and late gestational stages) (p ≤ 0.02, n ≥ 9). There was a more vigorous platelet aggregation response in the late gestational group compared to the non-pregnant control and early gestational groups (areas under the curve with ADP (p ≤ 0.004, n ≥ 7) and collagen (p ≤ 0.028, n ≥ 7).

Conclusions and future directions: Our results demonstrate significant changes in blood volume and a dramatic increase in postpartum platelet counts. Our results are consistent with those observed in pregnant humans, indicating that platelet counts have a similar response to pregnancy in both humans and mice. Our results demonstrating a late gestational increase in platelet reactivity suggest that pregnancy induced platelet alterations could play a role in the increased risk of VTE during pregnancy and the immediate post-partum period. We are currently performing additional analyses including RNA sequencing, to determine the mechanisms responsible for the phenotypic changes that we have observed here. The results of our studies could lead to enhanced treatment strategies for pregnancy associated VTE.

Disclosures

No relevant conflicts of interest to declare.