Modifications to the hemagglutinin cleavage site control the virulence of a neurotropic H1N1 influenza virus.

A key determinant of influenza virus pathogenesis is mutation in the proteolytic cleavage site of the hemagglutinin (HA). Typically, low-pathogenicity forms of influenza virus are cleaved by trypsin-like proteases, whereas highly pathogenic forms are cleaved by different proteases (e.g., furin). Influenza virus A/WSN/33 (WSN) is a well-studied H1N1 strain that is trypsin independent in vitro and has the ability to replicate in mouse brain. Previous studies have indicated that mutations in the neuraminidase (NA) gene allow the recruitment of an alternate protease (plasminogen/plasmin) for HA activation. In this study we have identified an additional mutation in the P2 position of the WSN HA cleavage site (S328Y) that appears to control virus spread in a plasmin-dependent manner. We reconstructed recombinant WSN viruses containing tyrosine (Y), phenylalanine (F), or serine (S) in the P2 position of the cleavage site. The Y328 and F328 viruses allowed plaque formation in the absence of trypsin, whereas the S328 virus was unable to form plaques under these conditions. In mice, Y328 and F328 viruses were able to efficiently spread following intracranial inoculation; in contrast, the S328 virus showed only limited infection of mouse brain. Following intranasal inoculation, all viruses could replicate efficiently, but with Y328 and F328 viruses showing a limited growth defect. We also show that wild-type HA (Y328) was more efficiently cleaved by plasmin than S328 HA. Our studies form the foundation for a more complete understanding of the molecular determinants of influenza virus pathogenesis and the role of the plasminogen/plasmin system in activating HA.

Maternally administered dexamethasone at 0.7 of gestation suppresses maternal and fetal pituitary and adrenal responses to hypoxemia in sheep.

Women who are at risk of preterm delivery are treated with antenatal steroids to facilitate fetal lung maturation. During this period, there is a potential for fetal or maternal hypoxemia to occur. Fetal responses to hypoxemia in sheep are well documented. However, less is known regarding maternal responses to hypoxemia. Therefore, we determined the effects of dexamethasone (DM) on maternal and fetal responses to hypoxemia in sheep. Ewes received four i.m. injections of DM or saline at 12-h intervals beginning at 103 d of gestation. Samples for ACTH, cortisol, and glucose were collected at 0900 h. At 105 d of gestation, hypoxemia was induced for 1 h by maternal nitrogen gas inhalation. Samples for ACTH, cortisol, and glucose were collected at 15-min intervals before, during, and after the hypoxemia challenge. Fluorescent microspheres were administered to the mother and the fetus before and during hypoxemia to measure organ perfusion. DM suppressed basal fetal and maternal cortisol and ACTH concentrations but increased glucose levels. DM also increased fetal but not maternal blood pressure. In control subjects, hypoxemia elevated fetal and maternal cortisol and ACTH concentrations. These responses were obliterated by DM. Hypoxemia increased blood pressure in DM-exposed fetuses but not in control subjects. In addition, hypoxemia decreased fetal adrenal vascular resistance in saline but not DM fetuses or ewes from either treatment group. In summary, maternal administration of a low dose of DM at 0.7 of gestation suppresses maternal and fetal adrenal function and changes fetal responses to hypoxemic stress to resemble those observed later in gestation.