Identification of the primary outcomes that result from deficient spiral arterial modification in pregnant mice

Abstract

Pre-eclampsia, an acute complication of human pregnancy, is associated within complete physiological modification of decidual spiral arteries. This is thought to promote oxidative stress from perfusion/reperfusion of the placenta and to restrict placental and fetal growth. Alymphoid (genotype Rag2(-/-)/Il2rg(-/-)) mice, sufficient in dendritic and myeloid cell functions, lack spiral arterial modification with individual spiral arteries having ~1.7x the vascular resistance and 0.66x the blood velocity of +/+ mice. Their placentae are measurably hypoxic yet neither placental growth nor fetal survival is impaired and gestational hypertension is not seen. Thus, lymphocytes rather than vascular adaptations appear to be the pivotal contributors to the clinical complications of pre-eclampsia.

Figure 1

Photomicrograph of mids aggital sections of placenta and deciduas basalis of (A) +/+ and (B) Rag2 ^−/−/Il2rg^−/− on gd12. Mesometrial aspect is to the top of each image. Panels (C, D) show at higher magnification, the typically observed differences in the decidua basalis between (A) modified and (B) unmodified spiral arteries, respectively. The differences in wall thicknesses and lumen diameters can be quantified morphometrically. From a series of comparisons between these genotypes on the BALB/c background, it was estimated that vascular resistance is 1.7x greater in an unmodified artery at gd12. Stained using DBA lectin. DB, decidua basalis; MLAp, mesometrial lymphoid aggregate of pregnancy P, placenta; SA, spiral artery; TGC, trophoblast giant cells; uNK + arrows, indicates several of the very abundant DBA+ uNK cell population. Bars indicate magnifications.

Figure 2

A model for the pathogenesis of pre-eclampsia in which endothelial cell dysfunction (ie activation, insufficient availability of viability and growth factor molecules, pro-coagulant state etc) is seen as the pivotal systemic event.

Figure 3

(A) Data are shown for mean ± SEM of viable fetuses in BALB/c+/+ (open bars) and Rag2 ^−/−/Il2rg^−/− (solid bars) mice mated and euthanized within our colony. Either 3 or 4 litters were studied per histogram bar. Many more litters would need to be sacrificed to obtain an accurate estimation of fetal success for these genotypes. These preliminary data show however that viability of Rag2 ^−/−/Il2rg^−/− is not disadvantaged compared with +/+. (B) Data are shown for mean± SEM cross sectional areas of placentae. Errors are too small for visualization of the error bars. A total of 135 tissues sections were scanned for each mean. At gd10, 12 and 14, there were no significant differences in placenta size between the genotypes. At gd 16, +/+ placentae were significantly larger (6.16mm² ± 0.17 versus 5.4 mm² ± 0.14 P<0.001) and there was no subsequent placental growth (+/+ placental size at gd18 was 5.99 mm² ± 0.29). Rag2 ^−/−/Il2rg^−/− placentae continued to grow and had a peripartum gd18 mean size of 7.02 mm² ± 0.26. This was statistically larger (P<0.01) than in the +/+ females.

Figure 4

Postnatal growth (mean weight ±SEM) of BALB/c +/+ (○) and Rag2 ^−/−/Il2rg^−/− (●) from birth to 16 wks of age. All Rag2 ^−/−/Il2rg^−/− data are from our Queen’s University laboratory with 6–9 animals weighed/time point. BALB/c+/+ data are from mixed sources. BALB/c birth weights were published by Pal et al., (n=9) and Buzas et al., (n=11). Data for BALB/c growth to 8 wks of age (#) are from the Jackson Laboratory Mouse Phenome Database (n=40; www.jax.org). Data for BALB/c+/+ growth after 8 wks is from our Queen’s University laboratory. Because husbandry and diet may influence weight measurements, statistics are not presented for measures made before wk 10, which was 2 wk after receipt and conditioning of BALB/c +/+ at our colony. For both genotypes, data collected before weaning (<4 wk), include both sexes. Wk 4–16 data are for females only. *P<0.01 compared with BALB/c+/+ females.

Figure 5

Change (Δ) in 24 hr mean arterial pressures (MAP) across pregnancy in BALB/c +/+ (○; n=5) and Rag2 ^−/−/Il2rg^−/− (●; n=9) for females delivering live born offspring. Because no changes from a 4 day pre-conception baseline were found during pre-implantation gestation in these mice or C57Bl/6J, gd0–3 mean 24 hr MAP was used for subsequent daily 24hr mean statistical comparisons. Both strains showed a decline in ΔMAP shortly after implantation that continued until the time of spiral arterial modification and opening of the placental circulation (~gd9–10). The decline was statistically significant from the pre-implantation baseline for Rag2 ^−/−/Il2rg^−/−. A gain in ΔMAP followed and baseline values were achieved a few days later. These remained relatively stable, without hypertension until parturition (P). A pattern of late gestational hypertensive (not shown) was seen in two Rag2 ^−/−/Il2rg^−/− females that gave birth to litters of stillborn pups.

Figure 6

A model for the pathogenesis of pre-eclampsia in which lymphocyte dysfunction triggers the pregnancy complications that arise from absence of spiral arterial modification. Although immune activation has been documented by others as a component of the pre-eclampsia syndrome, our studies suggest that it is exclusively the changes in lymphocytes and not those in antigen presenting or myeloid-lineage immune cells such as macrophages, neutrophils and mast cells that are central to pathogenesis.

Figure 7

A model for pathways that lymphocytes may utilize to drive pregnancy complications including intrauterine growth restriction and fetal death.