Flu biology: receptors, II

In the first part of this two parter we summarized some biology background to a new paper that appeared online ahead of print in the FASEB Journal, Yuo et al., "Avian influenza receptor expression in H5N1-infected and noninfected human tissues." The paper addresses an important gap in our knowledge. Are there cells in the human body with appropriately matched receptors for avian flu virus and if so, where are they? Sporadic results in the last few years suggested that cells deep in the human lung (so-called type II pneumocytes) and ciliated cells in the upper respiratory tract might have avian receptors. But the evidence was confusing. And what about cells in other organs? Some H5N1 patients had multi-organ system involvement, particularly the nervous system, but in one case, also the intestinal tract. The new paper looked for evidence of bird or human receptors on human cells in the upper and lower respiratory tracts, and cells in the brain, heart, kidneys, liver, spleen, intestines and placenta.

Before considering the results, consider first the technical difficulty. We want to know, for individual cells, whether they have a particular sugar connected to an underlying protein on their surface in one of two configurations: an α2,3Gal linkage or an α2,6Gal linkage (see our explanations here, here, here and here). In each case the constituent atoms and their amounts are identical. Only a slight change in where a hook-up is taking place differs. In the Yuo et al. paper this is done by using plant lectins specific for the two linkages. What's a lectin? A lectin is a protein that binds or sticks to a sugar, like the sialic acid sugar that make up the two different kinds of receptors on the cell surface. The virus has a lectin on its surface called hemagglutinin (it is the H part of H5N1). Depending upon whether it is a hemagglutinin on the surface of a bird virus or a hemagglutinin on the surface of a human adapted virus it sticks to a sialic acide with an α2,3Gal linkage or an α2,6Gal linkage, respectively. But the authors didn't use the viral lectin. They used instead lectins derived from plants, one designated MAA comes from a leguminous plant, Maackia amurensis (lectin II), and is said to be specific for the α2,3Gal (bird) linkage. The other comes the black elderberry, Sambucus nigra agglutinin, and is supposedly specific for the (human) α2,6Gal linkage. The researchers tagged each of the plant lectins (by attaching a protein called biotin) and used a fluorescent dye system to make it visible. Thus cells that have one or the other lectin attached to their surfaces could be visualized. There is more involved, of course. The cell types also had to be identified, and this was done with techniques derived from immunology (immunohistochemistry). Studies like this have a lot of moving parts. It's easy to describe the results but actually carrying them out is tricky and takes skill, patience and ingenuity. I mention this because we sometimes forget this and wonder why important information isn't obtained faster. Unfortunately it's not just a matter of throwing money at the problem. The most important problems are hard, because they wouldn't be considered important if they weren't (because we'd know the answer already).

This group, a collaboration between the Peking University in Beijing and Northwestern University in Chicago, examined the tissues of two deceased H5N1 patients from China and 14 control cases (none died of infectious respiratory disease). They made thin sections of the different organ tissues, stained them for the tagged plant lectins and looked to see which ones lit up (I'm omitting an important control maneuver they performed using pretreatment with neuraminidase to check for the sialic acid specificity of the plant lectins; however it doesn't seem this checked for linkage specificity).

One important finding was that endothelial cells in blood vessels have both receptors. Blood vessel endothelial cells are the inner lining of blood vessels and since almost all tissues have a blood supply (a few don't but these are unusual exceptions) this means there are potential receptors in almost every organ -- if the virus can get there. Then the question becomes whether a virus that can latch on to an endothelial cell can infect it productively and if so, whether the virus that is produced can infect other cells in the same tissue or travel via the blood stream to a tissue that can be infected. Avian receptors are also found on both red and white blood cells, but only T-cells, not B-cells (B-cells are the antibody making cells).

What other tissues have bird receptors in humans, at least by the methods of this study? They confirmed that the deep lung type II pneumocytes have α2,3Gal receptors as do non-ciliated (and some ciliated) cells elsewhere in the respiratory tract, including the upper tract. But no avian receptors were seen in the trachea, despite the fact viral genomic sequences can be found in tracheal cells (indicating infection) seems to meant the receptor is not obligatory. They observe also that α2,3Gal receptors are widespread in the deep lung but that in clinical material only some cells are infected. Bottom line: having an α2,3Gal receptor does not seem to be necessary to get you infected with an avian virus. Other factors are involved. On the other side of the question, since the human respiratory tract is well supplied with both receptors, the authors raise the question whether humans could also act as the kind of mixing vessel we thought the pig was, that is a venue for reassortment or recombination of human and avian viruses. Nice thought.

Also of interest is the demonstration that neurons and neural tissue have avian receptors, explaining perhaps the catastrophic central nervous system involvement seen in many cases. Neither human nor avian receptors were found on the cells of the intestine that line its surface, in contrast to another study, but the authors suggest that perhaps the receptors seen elsewhere were actually in neural tissue in the gut, not intestinal cells per se. this is just a reminder that we are just scratching the surface of the science here and there is much more that needs to be learned, reconfirmed and checked before we start concocting elaborate stories and theories about what is happening.

Avian receptors were also seen in some liver and kidney cells, although not abundantly. On the other hand, it is clear that there are cells with avian (α2,3Gal) receptors all over the body, in some tissues more than others, but the density of these cells does not seem to correlate closely with infection location in patients with H5N1.

So we've learned something and may have to unlearn other things. Isn't science wonderful?