Influenza (flu) is a contagious respiratory illness caused by flu viruses, leading to about 36,000 deaths every year in the United States alone, with the potential for at least a ten fold increase in epidemic and pandemic scenarios. During attachment of flu viruses to epithelial cells, hemagglutinin, one of its surface proteins, binds to sialic acid residues, initiating a series of events leading to activation of PKC, which in turn, down-regulates the activity of amiloride sensitive epithelial Na+ channels (ENaC) of tracheal and alveolar cells. It has been thought that these events are responsible for flu-induced rhinorrhea and life-threatening alveolar edema in humans. However, events occurring during the attachment of influenza virus to epithelial cells are likely to be transient and relatively few cells will be initially affected. We propose that M2, a transmembrane protein that plays a critical role in viral replication, enhances intracellular production of reactive oxygen-nitrogen species (RONS) which (i) oxidize and nitrate ENaC;and (ii) activate PKC(. Both processes enhance ENaC ubiquitination and subsequent destruction by the proteasome or lysosome systems. These hypotheses will be tested by completing the following comprehensive in vitro and in vivo studies listed in four specific aims: (1) Identify regions and specific amino acids of the influenza strain A/Udorn/72 M2 proton (H+) channel responsible for ENaC down-regulation in Xenopus oocytes microinjected with 1-,2-, and 3-ENaC. (2) Identify the mechanisms by which M2 decreases ENaC protein levels and function. We propose that M2 enhances intracellular production of reactive oxygen-nitrogen species (RONS) which (i) oxidize and nitrate ENaC;and (ii) activate PKC(. Both processes enhance ENaC ubiquitination and subsequent destruction by the proteasome or lysosome systems (3) Identify the mechanisms by which M2 inhibits amiloride sensitive Na+ currents in human airway (H441) and rat alveolar type II (rATII) cells, expressing native ENaC and (4) Establish the contribution of M2 in the inhibition of lung fluid clearance of mice infected by replicating flu viruses and identify the mechanisms involved. The results of our studies may provide the rational basis for the development of new therapeutic strategies, against a highly conserved region of the viral genome to knockdown M2 expression, and thus broadly and effectively decrease flu induced pulmonary edema and rhinorrhea. Due to the public health impact of influenza, there is a strong need to investigate and develop therapies that address the host response to viral infection, which may contribute to the morbidity and mortality of pathogenic respiratory viruses.

Public Health Relevance

The results of our studies will provide the rational basis for the development of new therapeutic strategies, such as administration of agents to decrease M2 expression, and thus broadly and effectively decrease the flu-induced rhinorrhea, alveolar edema and hypoxemia. Due to the public health impact of influenza, there is a strong need to investigate and develop therapies that address the host response to viral infection, which may contribute to the morbidity and mortality of pathogenic respiratory viruses.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL031197-27
Application #
8286342
Study Section
Lung Injury, Repair, and Remodeling Study Section (LIRR)
Program Officer
Harabin, Andrea L
Project Start
1987-09-01
Project End
2014-06-30
Budget Start
2012-07-01
Budget End
2014-06-30
Support Year
27
Fiscal Year
2012
Total Cost
$379,870
Indirect Cost
$101,277
Name
University of Alabama Birmingham
Department
Anesthesiology
Type
Schools of Medicine
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Lazrak, Ahmed; Jurkuvenaite, Asta; Ness, Emily C et al. (2014) Inter-?-inhibitor blocks epithelial sodium channel activation and decreases nasal potential differences in ?F508 mice. Am J Respir Cell Mol Biol 50:953-62
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Lazrak, Ahmed; Chen, Lan; Jurkuvenaite, Asta et al. (2012) Regulation of alveolar epithelial Na+ channels by ERK1/2 in chlorine-breathing mice. Am J Respir Cell Mol Biol 46:342-54
Lazrak, Ahmed; Jurkuvenaite, Asta; Chen, Lan et al. (2011) Enhancement of alveolar epithelial sodium channel activity with decreased cystic fibrosis transmembrane conductance regulator expression in mouse lung. Am J Physiol Lung Cell Mol Physiol 301:L557-67

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