Allergies! Type I Hypersensitivity: When More Isn’t Better

Our last article discussed various hay fever inducing allergens encountered throughout the year. We learned that even for some of the most allergenic pollens, like birch and ragweed, only certain antigens derived from the pollen actually induce an allergic response. While the differences in the structure of these primary antigens can partially explain why some are allergenic and others are not, it really boils down to how the antigen interacts with an individual’s immune system. Some molecules make better allergens than others because they interact with the major player in Type I hypersensitivity, immunoglobulin E (IgE).

Interestingly, IgE earned its name based on the fact that it reacted with the ragweed pollen antigen E, now known as the primary ragweed antigen “Amb a 1”. In 1921, scientists K. Prausnitz and H. Kustner identified a serum component that was responsible for allergic reaction. It wasn’t until 1966 that T. and K. Ishikawa identified IgE as the serum component. Everyone has a small amount of this potent antibody circulating the blood; IgE accounts for less than 0.05-0.2% (0.1-0.4 μg/mL) of the circulating antibodies in non-atopic individuals. Some, but not all, atopic individuals have higher levels of circulating IgE, up to 0.79%.

Even Sabrina Fairchild knew that “More isn’t always better…sometimes it’s just more.”

In individuals without allergies, an IgE-mediated immune response occurs as a defense against parasitic infections. In this case, the resulting physiological changes clear the parasite and protect the body against further damage caused by the parasite. However, in individuals with allergies, the IgE-mediated response is classified as a Type I hypersensitivity.

Let’s follow a pollen grain on its first journey in an allergic individual. The first encounter of an allergen sensitizes the individual to that specific allergen, but symptoms are not experienced. Initially, the pollen particle encounters the peripheral defenses, nasal hairs, eyelids, and beating cilia in the throat. These hairs prevent most particles from entering the airway or sinuses. Pollen particles must be extremely tiny (about 1x10^-6 meters) to pass through this initial barrier. Upon reaching the nasal mucosa, enzymes in mucous secretions break down the tough outer shell of the pollen (the exine), releasing the allergenic substance.

 

Antigen presenting cells engulf the allergenic substance, process it with enzymes, and display the antigen on the cell surface within a cradle-like protein called the class II major histocompatibility complex (MHC). Another type of immune cell, called T-helper, or Th, cells bind the presented antigen. Th2 cells release molecules called cytokines, which communicate to naive B cells to begin dividing and maturing. Some B cells differentiate into plasma cells, which produce and secrete a specific class of antibodies, or immunoglobulin (Ig). Humans produce 5 circulating antibody isotypes:  IgG, IgM, IgA, IgD and IgE. Particularly, Th2 cells produce the cytokines interleukin (IL)-4 and IL-13, which stimulate B cells to produce IgE. The allergic response appears to be localized, as plasma cells secreting IgE are 1000 times greater in nasal mucosa than in circulation.

In addition to producing the correct isotype, the plasma cells also produce highly specific antibodies that will bind the antigen tightly. Through the process of clonal selection and clonal expansion, a specific IgE molecule with high affinity for the antigen is produced en masse, creating an army like the clone troopers.

Although the army of IgE clones may not be as large as the clone troopers, it's every bit as powerful in wreaking immune havoc.

The circulating IgE has a specific receptor that allows it to bind tissue mast cells and blood basophils. At this point, the body is considered “sensitized” to the allergen. Additionally, memory B cells are formed in preparation for the second encounter of the antigen.

Nothing happens yet, but the body essentially lays in wait to encounter the allergen again. Upon second exposure, the allergenic antigen binds two IgE molecules that are already situated on the mast cells and basophils. These crosslinked IgE molecules are much more stable and can continue sending signal for weeks. The signal, as allergy sufferers know all too well, is a massive inflammatory response mediated by various pharmacologically active molecules contained within and produced by mast cells and basophils. These cells store the inflammatory molecules, like histamine, in granules or inner pockets. When the antigen binds IgE, mast cells and basophils undergo degranulation, releasing large amounts of chemical mediators like the histamine targeted by most antihistamine allergy medications.

The antigen acts like Wile E. Coyote, detonating the IgE fuse, causing the mast cell bomb to explode and release clouds of histamine. Histamine, in turn, damages only our tissues, never touching the elusive (and harmless) Roadrunner allergen.

Mast cells quickly synthesize additional mediators, including leukotriene and prostaglandin. These mediators signal certain physiological changes, including vasodilation (nasal blockage), smooth muscle contraction (coughing), increased mucus secretion (runny nose), and increased vascular permeability (inflammation). Sensory nerves are stimulated, resulting in sneezing and itching. This early phase, or immediate hypersensitivity reaction, happens so rapidly that symptoms are noticed within minutes of exposure to the allergen.

Although histamine is probably the most well-known pharmacologically active molecule, it is actually not the most potent or the longest acting player. Rather, it is the first molecule released in the allergic reaction. Following degranulation, mast cells and basophils produce and release other mediators called prostaglandins and leukotrienes. Initially, contraction of bronchial and tracheal muscles is mediated by histamine, but shortly after, further contraction occurs as a result of prostaglandin and leukotrienes. Leukotrienes are 10 times more potent than histamine at causing bronchoconstriction than histamine.

How an antigen, say pollen, triggers an allergic response.

About 50% of the time, 4 to 8 hours after the early phase reaction, the late phase begins. Other cytokines, particularly IL-5, attract other inflammatory cells, including eosinophils. The symptoms of the late phase reaction resemble those of the early phase, but tend to be characterized by less sneezing and itching and more congestion and mucus production. The inflammatory response from the late phase can damage tissues and last for days.

So why do some people endure the suffering of hay fever and others do not? Tune in next time to find out the genetic and environmental factors that contribute to allergic rhinitis.

 

 

Contributed by Julia van Rensburg, PhD
Follow Julia on Twitter.

Ishizaka K, Ishizaka T, & Hornbrook MM (1966). Physico-chemical properties of human reaginic antibody. IV. Presence of a unique immunoglobulin as a carrier of reaginic activity. Journal of immunology (Baltimore, Md. : 1950), 97 (1), 75-85 PMID: 4162440

Kasaian MT, Meyer CH, Nault AK, & Bond JF (1995). An increased frequency of IgE-producing B cell precursors contributes to the elevated levels of plasma IgE in atopic subjects. Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, 25 (8), 749-55 PMID: 7584687

Verstraelen, S., Bloemen, K., Nelissen, I., Witters, H., Schoeters, G., & Heuvel, R. (2008). Cell types involved in allergic asthma and their use in in vitro models to assess respiratory sensitization Toxicology in Vitro, 22 (6), 1419-1431 DOI: 10.1016/j.tiv.2008.05.008

Takhar P, Smurthwaite L, Coker HA, Fear DJ, Banfield GK, Carr VA, Durham SR, & Gould HJ (2005). Allergen drives class switching to IgE in the nasal mucosa in allergic rhinitis. Journal of immunology (Baltimore, Md. : 1950), 174 (8), 5024-32 PMID: 15814733

From Herb Garden To Medicine Cabinet: Developing A New Drug for Malaria

We live on a lush planet filled with over 290,000 species of plants. Herbs are a particular type of plant that lack a wooden stem, and humans have often sampled them with hopes of finding a new food or flavoring. Sometimes ingestion of an herb produces unwanted effects, such as death. But other herbs have medicinal qualities, such as the alleviation of fever.

Dichroa febrifuga, a medicinal herb that has been historically used to treat fever, is named for its active ingredient, febrifugine.

Dichroa febrifuga is one of the most important herbs in traditional Chinese medicine, used for millennia to treat ailments such as malaria. Malaria is caused by a unicellular parasite called Plasmodium that is transmitted by mosquitoes, and a high fever is one of the trademark symptoms.

 

Malaria has a complex life cycle. After the parasites (sporozoites) are injected via mosquitoes, they travel to the liver (merozoites) and then infect red blood cells. In blood cells, they gobble up the hemoglobin as a nutrient source for replication and development into sexual stages (gametocytes) that can be taken up by another mosquito, thereby spreading the parasite to a new victim.

Malaria continues to be a devastating disease, killing up to 1 million people each year, most of whom are children under the age of five in sub-Saharan Africa. There is an urgent need for new treatments since the parasite has developed resistance to most of our anti-malaria drugs.

While effective against malaria, febrifugine is not tolerated well. What is needed is a better understanding of how febrifugine works:  how does it kill the malaria parasite? If the natural product’s mechanism of action against malaria could be identified, it would pave the way for the development of refined derivatives that are more specific against the parasite and less detrimental to patients. Alas, this is not an easy task. Over 2000 years in the making, scientists have now identified an enzyme in the parasite that is inhibited by febrifugine. That enzyme is called prolyl-tRNA synthetase.

Prolyl-tRNA synthetase is critical for the production of proteins in a cell, a process known as translation. As shown in the figure below, messenger RNA (mRNA), which serves as the “middle man” conveying the information in genes to build proteins, is read by molecular machines called ribosomes. Another type of RNA molecule called transfer RNA (tRNA) recognizes specific nucleotide sequences in the mRNA, bringing the corresponding amino acid to the ribosome so it can be added to a growing protein sequence.

The production of proteins in the cell. Proteins are composed of amino acids (the colored balls) that are connected together in a specific order, as directed by the gene coding for it. The chain of amino acids then typically folds into a three-dimensional shape so that the protein can do its job in the cell.

Aminoacyl-tRNA synthetase enzymes are needed to “charge” the tRNA; in other words, they attach the correct amino acid to the correct tRNA. When prolyl-tRNA synthetase is blocked by febrifugine, the amino acid proline does not get attached to tRNA. This leads to a buildup of “uncharged” tRNA, which is interpreted as a sign of starvation by the cell (or by the single-celled malaria parasite in this case). Proline is a common amino acid needed to build many proteins, and when prolyl-tRNA synthetase isn’t able to do its job, protein production grinds to a halt.

Even better, this enzyme is required in multiple stages of the parasite’s life cycle, knocking out both the liver and the blood forms. But as mentioned above, humans do not tolerate febrifugine very well, probably because we also have a version of prolyl-tRNA synthetase and perhaps other proteins that febrifugine poisons. Having identified this drug target is helping researchers develop derivatives of febrifugine, such as halofuginol, that act more strongly against the parasite’s prolyl-tRNA synthetase with less toxicity in humans.

Halofuginol is chemically similar to febrifugine (see above), having potent activity against malaria but less adverse effects on the host.

So how did scientists figure out that febrifugine targets prolyl-tRNA synthetase? There are several ways to identify the molecular mechanism of drug activity. In this case, the group cultured malaria in the presence of drug, forcing the parasites to evolve or die. Those that lived were less sensitive to febrifugine, meaning that they accrued a genetic change (one or more mutations in their DNA) that allowed them to persist despite the presence of the drug. This process is very analogous to the development of penicillin-resistant bacteria.

Parasites that were able to grow better in febrifugine had their genomes sequenced. Such a feat would have taken years and millions of dollars not long ago, but today it has become routine. The genome sequence of the febrifugine-resistant parasites contained a common mutation in the gene encoding prolyl-tRNA synthetase, which signaled that this enzyme plays a critical role in the drug’s action. Understanding how the parasite develops resistance also helps scientists design compounds that act on the target differently. As you may surmise, we are in a constant “arms race” with these insidious microbes, but this discovery is a step towards a victory for us.

 

Two independent parasite lines that were resistant to febrifugine, HFGR I and II, contained mutations in their prolyl-tRNA synthetase gene. In drug-sensitive parasites (Dd2), an amino acid called leucine (leu) is present at position 1444, but in the mutant parasites, a DNA change led to a different amino acid that conferred resistance to the drug.

 

Contributed by:  Bill Sullivan
Follow Bill on Twitter.

Herman JD, Pepper LR, Cortese JF, Estiu G, Galinsky K, Zuzarte-Luis V, Derbyshire ER, Ribacke U, Lukens AK, Santos SA, Patel V, Clish CB, Sullivan WJ Jr, Zhou H, Bopp SE, Schimmel P, Lindquist S, Clardy J, Mota MM, Keller TL, Whitman M, Wiest O, Wirth DF, & Mazitschek R (2015). The cytoplasmic prolyl-tRNA synthetase of the malaria parasite is a dual-stage target of febrifugine and its analogs. Science translational medicine, 7 (288) PMID: 25995223

I Am Groot! Plants Are More “Alive” Than We Think

So who saw Guardians of the Galaxy this summer? Awesome, wasn’t it? How could a movie with a talking raccoon and a 1970s-based soundtrack go wrong? Oh, and then there’s Groot, the beloved walking, talking tree-like creature who sprouted the catch phrase, “I AM GROOT!” Until Groot, the only sentient plants children probably knew of were the Evil Trees hurling apples at Dorothy in The Wizard of Oz. But Groot was a “Giving Tree” incarnate.    



Groot shatters the evil living plant stereotype and saves the day in Guardians of the Galaxy.

 

Groot was so endearing, he has given my kids a new reason not to eat their veggies – they don’t like the idea of slaughtering plants. When I reminded the kids that plants are only sentient in the movies, they gave me the look of skepticism. I know it well…I taught it to them. This inspired me to leaf through some botany literature. Much to my surprise, my kids might have a point! Plants are more “alive” than we ever imagined.

Exhibit A.  Plants know when they are being eaten alive! And they fight back…

There is a plant that scientists typically use as a model to study in the lab called Arabidopsis (thale cress, similar to broccoli but tastes even worse). Researchers recently discovered that when they expose these plants to vibrations that mimic those produced by a hungry caterpillar, the plants increase production of glucosinolate and anthocyanin defenses. These are mustard oils that don’t sit well with caterpillars. In other words, the plants can tell when they are being chewed on and release oils to deter the predator.

Another sophisticated defense system used by plants comes from studies on tobacco. When caterpillars attack, these tobacco plants produce “green leaf volatiles”, compounds that act as a distress call by attracting insects that devour caterpillars!

The Hungry Caterpillar: Adorable children's tale or a horror story of gruesome predation?

 

Exhibit B. Plants have a memory and can be trained.

Everyone is familiar with the story of Pavlov’s dog, the famous experiment that demonstrated classical conditioning. Dogs salivate when presented with meat. If you ring a bell before presenting the meat, the dogs become conditioned to salivate at just the sound of a bell. Plants do a similar thing when exposed to light. Researchers have shown that when a plant is exposed to a certain wavelength of light, and then infected with a plant pathogen, the plant “learns” to build up resistance to that pathogen when it “sees” that particular wavelength of light once again. Plants that were infected and then exposed to the light developed no protective response. Plants must possess some sort of biochemical nervous system and memory in order to execute this kind of behavior.

Plants don’t have a brain, but they do behave as if they can think.

 

Exhibit C. Plants protect their young.

Seeds are the equivalent of a plant’s babies and plants have evolved a variety of fascinating ways to take care of their young. Consider serotinous plants, which keep some of their seeds inside the plant body instead of releasing them into the environment where they can be eaten or destroyed by weather. These plants can hold onto their seeds and release them when the time is most favorable for them to survive.

In the 1986 film, “Little Shop of Horrors”, the carnivorous plant Audrey II demonstrated a terrifying new way plants could protect their kin.

 

In another striking example, scientists studying a plant called sea rocket (Cakile edentula) noticed that when grown in a pot with a different member of its species, its roots grew wildly so to soak up more water and nutrients from its competitor. However, if the sea rocket was put into the same pot as its offspring, this competition did not take place!

Still not convinced that plants are more alive than we give them credit for? Check out this video by Michael Pollan.
 

 

While the evidence above isn’t sufficient to conclude that plants on Earth are like Groot, it is clear they are capable of some level of feeling and response. But don’t tell your vegetarian friends…what else would they eat?

Contributed by:  Bill Sullivan

Follow Bill on Twitter.

 

 

Appel HM, & Cocroft RB (2014). Plants respond to leaf vibrations caused by insect herbivore chewing. Oecologia, 175 (4), 1257-66 PMID: 24985883

Allmann, S., & Baldwin, I. (2010). Insects Betray Themselves in Nature to Predators by Rapid Isomerization of Green Leaf Volatiles Science, 329 (5995), 1075-1078 DOI: 10.1126/science.1191634

Karpiński S, & Szechyńska-Hebda M (2010). Secret life of plants: from memory to intelligence. Plant signaling & behavior, 5 (11), 1391-4 PMID: 21051941

Dudley SA, & File AL (2007). Kin recognition in an annual plant. Biology letters, 3 (4), 435-8 PMID: 17567552

The Friday Five

Highlighting some of the coolest science news we’ve seen lately.

1. Summer is the time for BBQ. Learn the science of BBQ in this video by It’s Okay To Be Smart.

2. Ever wonder how they get the caffeine out of coffee? Ever wonder what they do with it once it is extracted?

3. Want to know how Tylenol works? Well, so do scientists!

4. The hills might have eyes, but plants have ears! Scientists claim that plants can even hear themselves being eaten alive…much like a graduate student at a thesis defense.

5. How do you turn someone on (or off)? Stimulate their claustrum. Their what?!

 

Science quote of the week:

"The future belongs to science and those who make friends with science." --Jawaharlal Nehru 

Contributed by:  Bill Sullivan

Follow Bill on Twitter: @wjsullivan


Koubeissi, M., Bartolomei, F., Beltagy, A., & Picard, F. (2014). Electrical stimulation of a small brain area reversibly disrupts consciousness Epilepsy & Behavior, 37, 32-35 DOI: 10.1016/j.yebeh.2014.05.027