The immunomodulatory role of heme oxygenase-1 during late pregnancy in mice

Abstract

The inducible enzyme heme oxygenase-1 (HO-1) plays a regulatory role in a number of inflammatory processes. It catabolizes the degradation of heme and acts as an anti-oxidant and cytoprotectant through its products biliverdin, carbon monoxide and free iron. In the placenta, HO-1 is highly expressed and assumed to mediate pregnancy-protective effects, but its function throughout pregnancy is not well understood. Our objective was to examine if a decreased HO-1 expression, e.g., upon stress challenges during late murine pregnancy or in heterozygous HO-1 knockout mice (further referred to as Hmox+/-) affect fetal development and maternal immune adaptation. DBA/2J-mated BALB/c females were exposed to 24 hour sound stress on gestation days (gd) 12.5 and 14.5. On gd 16.5 placental tissue and uterus-draining lymph nodes (LN) were collected, fetal weight was documented. Placental HO-1 expression was analysed by immunohistochemistry and quantitative real time-PCR. Hmox+/- mice were syngeneically mated. On gd 16.5 tissue was collected and fetal HO-1 genotype and weight was recorded. Placental tissue sections were stained with Masson’s trichrome to study placental function. Phenotype and frequency of maternal immune cells harvested from uterus-draining LN were identified by flow cytometry. For statistical analyses, continuous outcomes were checked for normal distribution. If data was normally distributed, one-way analyses of variance were carried out. For not normally distributed data, Mann-Whitney U-tests were used. Level of significance was set at a p-value of 0.05. Stress challenges led to a reduced fetal weight. Further, stress challenge lowered HO-1 tissue expression at the feto-maternal interface and significantly decreased HO-1 expression largely in T and dendritic cells among maternal leucocytes from LN, along with an increase in CD8+ T cells. The increased frequency of CD8+ T cells inversely correlated with a reduced expression of CD122 on CD8+ T cells in stressed mice, compared to control. CD8+CD122+ T cells are presumed to have regulatory, pregnancy-protective functions. Similarly, an increased frequency of CD8+ T cells and a reduction of CD122+ could be detected on HO-1+/- dams, compared to wild type (wt) controls. Lower fetal weight was observed in Hmox+/- and -/- offspring, compared to wt fetuses of the same litter. Placental size was slightly decreased in +/- offspring when compared to wt. This was accompanied by altered ratios of functional placental areas (labyrinth/junctional zone). Our results show that reduced HO-1 expression induced by stress or present in Hmox+/- mice results in an impaired immune adaptation to pregnancy, characterized by increased CD8+ T cells and decreased CD8+ regulatory T cells, which are pregnancy-protective functions and of which generation is presumably induced by placental HO-1. This coincides with altered placental function and signs for fetal growth restriction. Reduced HO-1 expression could thereby have detrimental short- and/or long-term effects on offspring development and health. To prevent these effects pharmacological interventions could target HO-1 to modulate immune tolerance during pregnancy.