Activin and NADPH-oxidase in preeclampsia: insights from in vitro and murine studies.

OBJECTIVE: Clinical management of preeclampsia has remained unchanged for almost 5 decades. We now understand that maternal endothelial dysfunction likely arises because of placenta-derived vasoactive factors. Activin A is one such antiangiogenic factor that is released by the placenta and that is elevated in maternal serum in women with preeclampsia. Whether activin has a role in the pathogenesis of preeclampsia is not known. STUDY DESIGN: To assess the effects of activin on endothelial cell function, we cultured human umbilical vein endothelial cells in the presence of activin or serum from normal pregnant women or pregnant women with preeclampsia, with or without follistatin, a functional activin antagonist or apocynin, a NADPH oxidase (Nox2) inhibitor. We also administered activin to pregnant C57Bl6 mice, with or without apocynin, and studied maternal and fetal outcomes. Last, we assessed endothelial cell Nox2 and nitric oxide synthase expression in normal pregnant women and pregnant women with preeclampsia. RESULTS: Activin and preeclamptic serum induced endothelial cell oxidative stress by Nox2 up-regulation and endothelial cell dysfunction, which are effects that are mitigated by either follistatin or apocynin. The administration of activin to pregnant mice induced endothelial oxidative stress, hypertension, proteinuria, fetal growth restriction, and preterm littering. Apocynin prevented all of these effects. Compared with normal pregnant women, women with preeclampsia had increased endothelial Nox2 expression. CONCLUSION: An activin-Nox2 pathway is a likely link between an injured placenta, endothelial dysfunction, and preeclampsia. This offers opportunities that are not novel therapeutic approaches to preeclampsia.

Activin A contributes to the development of hyperoxia-induced lung injury in neonatal mice.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is one of the leading causes of morbidity and mortality in babies born prematurely, yet there is no curative treatment. In recent years, a number of inhibitors against TGFß signaling have been tested for their potential to prevent neonatal injury associated with hyperoxia, which is a contributing factor of BPD. In this study, we assessed the contribution of activin A-a member of the TGFß superfamily-to the development of hyperoxia-induced lung injury in neonatal mice. METHODS: We placed newborn C57Bl6 mouse pups in continuous hyperoxia (85% O2) to mimic many aspects of BPD including alveolar simplification and pulmonary inflammation. The pups were administered activin A receptor type IIB-Fc antagonist (ActRIIB-Fc) at 5 mg/kg or follistatin at 0.1 mg/kg on postnatal days 4, 7, 10, and 13. RESULTS: Treatment with ActRIIB-Fc and follistatin protected against hyperoxia-induced growth retardation. ActRIIB-Fc also reduced pulmonary leukocyte infiltration, normalized tissue: airspace ratio and increased septal crest density. These findings were associated with reduced phosphorylation of Smad3 and decreased matrix metalloproteinase (MMP)-9 activity. CONCLUSION: This study suggests that activin A signaling may contribute to the pathology of bronchopulmonary dysplasia.

Human amnion epithelial cells modulate hyperoxia-induced neonatal lung injury in mice.

BACKGROUND AIMS: Human amnion epithelial cells (hAECs) prevent pulmonary inflammation and injury in fetal sheep exposed to intrauterine lipopolysaccharide. We hypothesized that hAECs would similarly mitigate hyperoxia-induced neonatal lung injury. METHODS: Newborn mouse pups were randomized to either normoxia (inspired O2 content (FiO2) = 0.21, n = 60) or hyperoxia (FiO2 = 0.85, n = 57). On postnatal days (PND) 5, 6 and 7, hAECs or sterile saline (control) was administered intraperitoneally. All animals were assessed at PND 14. RESULTS: Hyperoxia was associated with lung inflammation, alveolar simplification and reduced postnatal growth. Administration of hAECs to hyperoxia-exposed mice normalized body weight and significantly attenuated some aspects of hyperoxia-induced lung injury (mean linear intercept and septal crest density) and inflammation (interleukin-1α, interleukin-6, transforming growth factor-ß and platelet-derived growth factor-ß). However, hAECs did not significantly alter changes to alveolar airspace volume, septal tissue volume, tissue-to-airspace ratio, collagen content or leukocyte infiltration induced by hyperoxia. CONCLUSIONS: Intraperitoneal administration of hAECs to neonatal mice partially reduced hyperoxia-induced lung inflammation and structural lung damage. These observations suggest that hAECs may be a potential therapy for neonatal lung disease.

The Animal-Human Interface in Farm Animal Production: Animal Fear, Stress, Reproduction and Welfare.

A negative human-animal relationship (HAR) from the perspective of the animal is a limiting factor affecting farm animal welfare, as well as farm animal productivity. Research in farm animals has elucidated sequential relationships between stockperson attitudes, stockperson behaviour, farm animal fear behaviour, farm animal stress physiology, and farm animal productivity. In situations where stockperson attitudes to and interactions with farm animals are sub-optimal, through animal fear and stress, both animal welfare and productivity, including reproductive performance, can be compromised. There is a growing body of evidence that farm animals often seek and enjoy interacting with humans, but our understanding of the effects of a positive HAR on stress resilience and productivity in farm animals is limited. In this review, we explore the pathways by which stress induced by human-animal interactions can negatively affect farm animal reproduction, in particular, via inhibitory effects on the secretion of gonadotrophins. We also review the current knowledge of the stockperson characteristics and the nature of stockperson interactions that affect fear and physiological stress in farm animals. The contents of this review provide an insight into the importance of the HAR on farm animal welfare and reproduction while highlighting the gap in knowledge regarding the effects of a positive HAR on farm animals.

Human amnion epithelial cells as a treatment for inflammation-induced fetal lung injury in sheep.

OBJECTIVE: The purpose of this study was to determine whether human amnion epithelial cells (hAECs) can modulate the pulmonary developmental consequences of intrauterine inflammation in fetal sheep that are exposed to intraamniotic lipopolysaccharide (LPS) injection. STUDY DESIGN: At 117 days' gestation, fetal sheep (n=16) received intraamniotic LPS (20 mg). hAECs were delivered at 0, 6, and 12 hours into the fetal jugular vein (n=4), trachea (n=4), or both (n=4). Controls (n=6) received equivalent administration of saline solution. Lungs were collected at 124 days. RESULTS: Intraamniotic LPS caused pulmonary inflammation and altered lung structure and function. hAECs attenuated changes in lung function and structure that had been induced by LPS: lung volume, 40 cm H2O (P<.05, intravenous+intratracheal hAECs vs LPS), tissue-to-airspace ratio (P<.05, intravenous+intratracheal hAECs vs LPS), and septal crest density (P<.001, all hAEC groups vs LPS). Leukocyte infiltration of the lungs was not reduced by hAECs; however, inflammatory cytokines were reduced (tumor necrosis factor-α, P<.01, vs LPS; interleukin-1b, P<.01, vs LPS; interleukin-6, P<.01 vs LPS). Surfactant protein A and C messenger RNA was increased by LPS, although this was not statistically significant (P>.05 vs control); there were significant increases in all hAEC-treated animals (surfactant protein-A, P<.05 vs LPS; surfactant protein-C, P<.01 vs LPS). CONCLUSION: Human amnion epithelial cells attenuate the fetal pulmonary inflammatory response to experimental intrauterine inflammation and reduce, but (as administered in our study) do not prevent, consequent alterations in lung development.

Enrichment with Lucerne Hay Improves Sow Maternal Behaviour and Improves Piglet Survival.

This study investigated the effects of providing lucerne hay on the behaviour and the performance of sows housed in farrowing crates during farrowing and lactation. Seventy-two mixed parity sows received either 1 kg lucerne hay daily from entry into the farrowing crate (-2 d from expected farrowing date) until weaning at 17 d (lucerne group, n = 36), or received no additional enrichment (control group, n = 36). In the 18 h prior to farrowing, the sows in the lucerne treatment spent more time performing nest-building behaviour (14.8% lucerne vs 11.1% control, p = 0.0009) and less time sham-chewing (1.0% lucerne vs 1.9% control, p = 0.01) than control sows, and gave birth to fewer stillborn piglets/litter (0.1 lucerne vs 0.4 control, p = 0.027). After farrowing (Day 3), the control sows spent less time lying than the lucerne sows (26% control vs 43% lucerne, p < 0.05). The control sows also spent less time interacting with their piglets during early lactation compared to late lactation (25.5% Day 5 vs 47.3% Day 12, p < 0.05), suggesting reduced maternal behaviour in this group. The lucerne sows continued to interact with the lucerne throughout lactation, indicating that they still found the enrichment rewarding after the nesting period had ceased. Based on these results, lucerne enrichment was considered to improve sow welfare during farrowing and lactation and reduce the number of stillborn piglets.

Role of activin A in the pathogenesis of endothelial cell dysfunction in preeclampsia.

Circulating markers for endothelial activation such as endothelin-1 (ET-1), ICAM-1 and VCAM-1 are elevated in women with preeclampsia. Using human umbilical vein endothelial cells (HUVECs) as an in vitro model of the maternal vasculature, we show that activin A and preeclamptic serum upregulate ET-1, ICAM-1, and VCAM-1 in HUVECs. Further, we show that follistatin, a specific binding protein for activin, mitigates the upregulation of ET-1, ICAM-1 and VCAM-1 in HUVECs exposed to either activin A or preeclamptic serum. These data are consistent with activin A contributing to the pathophysiology of preeclampsia and suggest that therapies targeting activin signalling are worth exploring.

Human amnion epithelial cells repair established lung injury.

With a view to developing a cell therapy for chronic lung disease, human amnion epithelial cells (hAECs) have been shown to prevent acute lung injury. Whether they can repair established lung disease is unknown. We aimed to assess whether hAECs can repair existing lung damage induced in mice by bleomycin and whether the timing of cell administration influences reparative efficacy. In addition, we aimed to characterize the effect of hAECs on fibroblast proliferation and activation, investigating possible mechanisms of reparative action. hAECs were administered intraperitoneally (IP) either 7 or 14 days after bleomycin exposure. Lungs were assessed 7 days after hAEC administration. Bleomycin significantly reduced body weight and induced pulmonary inflammation and fibrosis at 14 and 21 days. Delivery of hAECs 7 days after bleomycin had no effect on lung injury, whereas delivery of hAECs 14 days after bleomycin normalized lung tissue density, collagen content, and α-SMA production, in association with a reduction in pulmonary leucocytes and lung expression of TGF-ß, PDGF-α, and PDGF-ß. In vitro, hAECs reduced proliferation and activation of primary mouse lung fibroblasts. Our findings suggest that the timing of hAEC administration in the course of lung disease may impact on the ability of hAECs to repair lung injury.

Human amnion epithelial cells prevent bleomycin-induced lung injury and preserve lung function.

Human amnion epithelial cells (hAECs) have attracted recent attention as a promising source of cells for regenerative therapies, with reports that cells derived from human term amnion possess multipotent differentiation ability, low immunogenicity, and anti-inflammatory properties. Specifically, in animal models of lung disease characterized by significant loss of lung tissue secondary to chronic inflammation and fibrosis, the transplantation of hAECs has been shown to reduce both inflammation and subsequent fibrosis. To further explore the mechanisms by which hAECs reduce pulmonary fibrosis and enhance lung regeneration, we utilized a bleomycin-induced model of pulmonary fibrosis and investigated the ability of hAECs to reduce fibrosis and thereby improve pulmonary function. We aimed to determine if hAECs, injected into the peritoneal cavity could migrate to the lung, engraft, and form functional lung epithelium, and whether hAECs could modulate the inflammatory environment in the bleomycin-injured lung. We demonstrated that, compared to bleomycin alone, IP administration of hAECs 24 h after bleomcyin, decreased gene expression of the proinflammatory cytokines TNF-α, TGF-ß, IFN-γ, and IL-6 and decreased subsequent pulmonary fibrosis with less pulmonary collagen deposition, reduced levels of α-smooth muscle actin and decreased inflammatory cell infiltrate. We also showed that hAECs are able to prevent a decline in pulmonary function associated with bleomycin-induced lung damage. We were unable to detect any significant engraftment of hAECs in injured, or uninjured, lung after administration. The findings from this study support the further investigation of hAECs as a potential cell therapy for inflammatory and fibrogenic diseases.

Amnion epithelial cell isolation and characterization for clinical use.

Human amnion epithelial cells (hAECs) are a heterologous population positive for stem cell markers; they display multilineage differentiation potential, differentiating into cells of the endoderm (liver, lung epithelium), mesoderm (bone, fat), and ectoderm (neural cells). They have a low immunogenic profile and possess potent immunosuppressive properties. Hence, hAECs may be a valuable source of cells for cell therapy. This unit describes an efficient and effective method of hAEC isolation, culture, and cryopreservation that is animal product-free and in accordance with current guidelines on preparation of cells for clinical use. Cells isolated using this method were characterized after 5 passages by analysis of karyotype, cell cycle distribution, and changes in telomere length. The differentiation potential of hAECs isolated using this animal product-free method was demonstrated by differentiation into lineages of the three primary germ layers and expression of lineage-specific markers analyzed by PCR, immunocytochemistry, and histology.