Tuesday, August 31, 2004

Wounding maggots

I'm always on the lookout for simple new preps for developmental biology labs, and this paper by Galko and Krasnow describes an interesting possibility: studying wound healing in Drosophila larvae. It seems easy to do—punch a small hole in the little maggot, and watch as it repairs itself—and it's a process relevant to all of us. I'm already planning to have the students poison developing zebrafish embryos and observe the formation of teratological defects, so I think sending 'em off to stab baby flies would fit right in with the program.

The Galko and Krasnow paper also does a good job of delivering background, and in particular discussing the similarities and differences between mammalian and insect larva wound healing. I'm always conscious of the fact that when you're discussing model systems, you have to be extremely careful to separate out what are thought to be the common properties from the derived features.

Similarities:

• Clot formation
• Epidermal cell migration into the wound/clot
• Extension by lamellipodia to form a continuous cellular mesh
• Redifferentiation
• Recruitment of inflammatory cells
• Remodeling and degradation of the clot

Differences:

• Different coagulogens
• Instead of proliferating, local cells in Drosophila form a syncytium
• Migrating Drosophila cells do not detach from the basal lamina
• Drosophila have relatively little recruitment of cells to the wound site

As an added bonus, wound healing in the fly involves the Fos/Jun pathway via Jun N-terminal kinase (JNK), so there's some interesting molecular shenanigans going on; Fos and Jun are ubiquitous transcription factors and oncogenes that are also involved in cancer metastasis, so it's not surprising to see them active in migrating, dedifferentiating cells.

 

I'll have to see what I can work up in the class. Right now I emphasize early embryogenesis, but regeneration and repair are also important subjects in development.

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Galko MJ, Krasnow MA (2004) Cellular and Genetic Analysis of Wound Healing in Drosophila Larvae. PLoS Biol 2(8): e239.

Monday, August 30, 2004

Fetus toys!

Not only is it my daughter's birthday today, but it is the first day of classes, and I'm going to be teaching developmental biology this term. In a pleasant synchronicity, Skatje has found this very weird but embryologically relevant site, Fetopia, which sells beads and magnets and clay figures in the shape of cute little fetuses. They also sell Feto Soap, which sounds horribly gross, but no, it isn't made from fetuses, it just has a little fetus figurine embedded in it. There is even a Fetopia LiveJournal. I think I'm going to have to order some of these things.

I agree with a sentiment found in the Feto FAQ:

Fetos were not meant to be symbolic of the pro-life, nor the pro-choice campaign. We at FeTO just think fetos are neat. Fetos are meant to represent neither living nor dead fetos. They are clay.

Very cool. We don't have to take sides in the abortion debate to agree that development and embryology and fetuses are neat.

An iconic image gets used again

I like it.

Friday, August 27, 2004

Clone Bone

This is kind of cool: Doctors Grow New Jaw Bone in Man's Back. It's not exactly an elegant use of stem cell technology (and there are reasons to avoid calling it a stem cell technique at all), but hey, if it works...

According to this week's issue of The Lancet medical journal, the German doctors used a mesh cage, a growth chemical and the patient's own bone marrow, containing stem cells, to create a new jaw bone that fit exactly into the gap left by the cancer surgery.
Tests have not been done yet to verify whether the bone was created by the blank-slate stem cells and it is too early to tell whether the jaw will function normally in the long term. But the operation is the first published report of a whole bone being engineered and incubated inside a patient's body and transplanted.

Here are a few more details on the technique:

Warnke and his group began by creating a virtual jaw on a computer, after making a three-dimensional scan of the patient's mouth.
The information was used to create a thin titanium micro-mesh cage. Several cow-derived pure bone mineral blocks the size of sugar lumps where then put inside the structure, along with a human growth factor that builds bone and a large squirt of blood extracted from the man's bone marrow, which contains stem cells.

Oral cancer is what hit my grandfather, and he had most of his jaw removed (not pretty). Then he had a chunk of rib carved into shape and stuck in there (still not pretty, but better), which was never very satisfactory—it was fragile and more of a placeholder than anything, and he was on a liquid diet for the rest of his life. This procedure sounds like it would have produced something much more satisfactory.

The article alludes to a few pending problems. Your jaw is more than just a piece of bone with a specific shape: it also has a particularly intricate internal structure, delicate struts and ribs of bone, that give it strength in appropriate directions and minimizes weight. That structure is partly a consequence of patterns of use, which the cultured bone hasn't experienced, so the new jaw is almost certainly not as functional as the original. Still, it's a clever way to build an approximation of the original living structural tissue.

Smacking both sides of the stem cell issue

Maybe you consider the right-wing ranter an absurd caricature, and that nobody could be that insane…but then I saw this, which comes awfully close.

As for the left-winger, man, am I ever sick of wimpy, mealy-mouthed liberals and the cowardly, bullying extremists of the right who whine about being victimized and abused every time a liberal raises a finger in objection.

Ediacaran fossils from Newfoundland

I told you I liked fossils, and here are some more that put yesterday's Junggarsuchus to shame in both age and weirdness. Everyone has heard of the Cambrian 'explosion', but there are also collections of pre-Cambrian fossil animals that have always been rather enigmatic—they just don't seem to correspond well to the morphology of Cambrian, or modern, forms.

Some new specimens from the pre-Cambrian have been described in a paper titled, "Modular construction of early Ediacaran complex life forms." They have been collected from 560 million year old rocks in Newfoundland, Canada. The focus of the paper is on patterns of organization: as the title says, these organisms appear to be modular in form, but it isn't the segmental modularity we see today. Instead, these pre-Cambrian animals were built on a fractal branching plan, repeated iterations of a structure called the "rangeomorph frondlet". The result was a creature that looked feathery or fern-like, and when described, it's hard to avoid using terms we usually associate with plants, like "stalk" and "leaf-like". But don't be confused, these are not plants, nor are they anything like modern animals, such as sea-pens, which have also adopted this kind of morphology.

All taxa at the new Spaniard's Bay locality, and most of the 20 to 30 other taxa of the Mistaken Point assemblage elsewhere, are composed of varying constructions of the same architectural building block—a centimeter-scale module herein termed a "rangeomorph frondlet". Each frondlet consists of inflated, self-similar branches that are indistinguishable from the "fractal pneus" defined by Seilacher. Pneus pass from the midline at an acute angle in an alternate pattern, but variable orientations of rangeomorph frondlets can give the appearance of asymmetric branch lengths or even branching on only one side of the frondlet. The pattern of branching is remarkably self-similar over three orders of fracticality: Major branches with diameters ranging from 1 to 5 mm are composed of minor branches 0.3 to 0.6 mm in diameter, which, in the best preserved specimens, appear to be composed of tertiary branches <150 µm in diameter.

Here's what a rangeomorph frondlet looks like:

And here are some examples of the kinds of creatures made up of these frondlets:

An accompanying review by Brasier and Antcliffe tries to give a bit more perspective on the Ediacaran fauna as a whole. As you can see from this diagram, the animals of the Ediacara had an almost 40 million year reign, during which they were the biggest, most complex forms on the planet.

The review makes another interesting point: that we may be able to better understand the evolution of the Ediacaran fauna, given enough samples, by taking a developmental approach and analyzing life cycles. The bottom part of the diagram illustrates that point. Some of the different forms may represent different stages in the life history of the organisms. At the same time, evolution often works by shifting the timing of development, a process called heterochrony, so as you look at different fossils over long periods of geological time, you may also be seeing shifts in form that are consequences of shifts in development. The authors presented no data on this idea (which is unfortunate, and a little odd), but I agree that the evo-devo approach is a promising one.

The Narbonne paper summarizes what we do know right now about the pre-Cambrian biota.

The morphologic features described above provide strong support for the view that rangeomorphs represent a single clade, a high-level taxon that went extinct in the terminal Proterozoic. It is probable that the Ediacara biota included stem groups for the Cambrian explosion of animals, but there are no obvious analogs for rangeomorph architecture and construction among modern taxa. Recent comparison of rangeomorph structure with the radial canals of fossil and recent ctenophores seems remote from a morphological standpoint and is also inconsistent with the internal organic skeletons documented above. It would be tempting to regard the two-dimensional view of a rangeomorph frondlet as representing the bases of an array of open tubes that housed polyps or other filter-feeding organisms, but this is not consistent with partially overturned specimens that show that both sides are identical; the very small diameter of the secondary (300 to 600 µm) and tertiary (<150 µm) pneus would also specifically rule out cnidarian polyps as the originators of the tubes.
These modular constructions effectively partitioned food resources: Spindle-shaped rangeomorph recliners lay on the sea floor, whereas bush-, plume-, comb-, and frond-shaped rangeomorphs were elevated above the sea floor and fed from different levels within the water column. Rangeomorphs dominated the earliest Ediacaran ecosystems, with no evidence for burrowing, mobile, or metameric megascopic organisms among the taxa of the Mistaken Point assemblage. Rangeomorphs occur only sporadically in younger Ediacaran assemblages, perhaps as a result of competition with early animals, and have not been reported from any Phanerozoic assemblage including fossil Lagerstätten such as the Burgess Shale. It is difficult to relate rangeomorphs to any modern group of macroscopic organisms, and they appear to represent a "forgotten" architecture and construction that characterized early stages in the terminal Neoproterozoic evolution of complex multicellular life.

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Brasier M, Antcliffe J (2004) Decoding the Ediacaran enigma. Science 305:1115-1117.

Narbonne, GM (2004) Modular construction of early Ediacaran complex life forms. Science 305:1141-1144.

Thursday, August 26, 2004

Mindbending McGurk

First, watch and listen to this short video (180K, QuickTime) of a man making baby-noises. But then, close your eyes and listen; the sounds will shift in an interesting way.

It's a very nifty example of the McGurk Effect, in which multiple modalities modify our perception of a stimulus. I won't bother to say how it works here, you'll just have to go read this nice explanation of how the video was made and how it works on the Freethought Forum.

Junggarsuchus

I'm a sucker for fossils, so here's one:

It's a new Jurassic sphenosuchid, or a sort of proto-crocodile, named Junggarsuchus sloani. The nifty thing about it is two-fold: 1) its skull has similarities to modern crocodilian skulls (those of us who have taken a comparative anatomy course know how strange crocodile skulls are—thick and bony, and the cruel TA expected us to be able to sort out all those bones fused together in a mish-mash), and 2) the rest of the skeleton was cursorially adapted, that is, it was better suited to running on land then modern crocodiles. Here's Clark et al.'s phylogenetic analysis:

They place it as the closest relative to the crocodyliformes, which suggests that modern crocodiles and their specialized skulls evolved from more terrestrial forms. Before you start imagining giant ferocious crocodiles galloping across the plains, though, I should mention that these were little guys, less than a few feet long total, and that skull is only about 6 inches long.

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Clark JM, Xu X, Forster CA, Wang Y (2004) A Middle Jurassic 'sphenosuchian' from China and the origin of the crocodylian skull. Nature 430:1021-1024.

Wednesday, August 25, 2004

Tangled Bank #10

We have the tenth Tangled Bank online…stop on by Wolverine Tom's place and see what's up.

If you'd like to be represented in the 11^th edition in two weeks, send your links to John McKay of Archy. And we're always looking for new people to host the biweekly affair, so if you're interested in that, send a line to pzmyers@pharyngula.org.

Tuesday, August 24, 2004

Hurry up!

Last call, everyone…send those science links to Wolverine Tom as soon as possible if you hope to make it in time for tomorrow's Tangled Bank.

Monday, August 23, 2004

The Loom is back in business

Carl Zimmer is posting again, and he's got an interesting and ominous (to certain traditionalists) interpretation of human evolutionary data:

Scientists have proposed that humans have a history of polygyny before (our sperm, for example, looks like the sperm of polygynous apes and monkeys, for example). But with these new DNA results, the Arizona researchers have made a powerful case that polygyny has been common for tens of thousands of years across the Old World. It's possible that polygyny was an open institution for much of that time, or that secret trysts made it a reality that few would acknowledge. What's much less possible is that monogamy has been the status quo for 50,000 years.

A history of human polygyny? Uh-oh. I hope my wife isn't going to tell me now that tradition demands that I share.

Aaiieee! Don't ever do this!!

This is a horrific story of an undergraduate student doing a science demonstration, which is inexplicably labeled as "humor." I didn't find it at all funny, I'm afraid. It starts reasonably enough, and then escalates slowly into something truly dreadful.

That may have been the point of no return. I, as is traditionally my role, answered that the nitrogen evaporates at the surface of the table, which provides a cushion of air for the drop to sit on, and thermally insulates the drop to minimize further evaporation. So you see a drop dance around without boiling away, and without interacting with the table and getting slowed down or smeared out. Then, I continued... I mentioned that the same principle makes it possible to dip a wet hand into molten lead, or to drink liquid nitrogen without injury.
I had done the latter several years earlier in a cryogenics lab, and remembered the physics behind how it worked. Naturally, people around me were skeptical. "You can't drink the stuff... it'll freeze your whole body... Remember 'Terminator 2?' " But I was sure of myself. I had done it before, and I believed in the physics behind it. So, naturally, I poured myself a glass and took a shot.

At this point, I could feel my entire gastro-intestinal tract going into sympathetic spasm. Ick. For a change, I will spare you the grisly details—read it yourself if you must—but basically the short story is the guy swallowed liquid nitrogen with catastrophic consequences. Here's the "happy" ending.

So... the recovery... they were impressed with my recuperative skills. I could breathe on my own completely after a few days. I could sit up in bed after a week, and was walking in two. About that time, I began to eat again as well. After four weeks, I was up and about again. Now, something like eight weeks, I'm virtually healed, with the exception of a number of unsightly scars.

I think you can surmise from the fact that the guy is relieved to be able to breathe and walk and is covered with scars that the rest of the story isn't exactly pretty.

(via Bastard Sword)

Better bug killing through Science

I spent part of my weekend reading a very nice, detailed paper by Soderlund and Knipple (thanks for the recommendation, Nick!) on the distribution and mechanisms of mutations that confer pesticide resistance on insects. The main message is that we have been using potent pyrethroid poisons that have a common mechanism, targeting the highly conserved sodium channel of the nervous system, and that similar mutations that reduce the affinity of the channel for the pyrethroid are popping up in many insect species.

First, a little elementary neurobiology. Signals are shuttled around the nervous system by pulses of ion flux that are primarily mediated by two ions, potassium (K⁺) and sodium (Na⁺). Neurons are typically sitting at a negative voltage at rest; a signal, or action potential, is a result of a transient opening of Na⁺ channels so that positive sodium ions rush in, depolarizing (or reducing the voltage difference towards zero) the cell. So, the resting state of a neuron is negative, and a change in state is signalled by short, sharp flicks towards 0.

Normally, these signals are very brief. There are two general reasons for that: one is that the sodium channel is kept on a tight leash, and after opening for that inward rush of positive sodium ions, it tends to automatically inactivate after a short period. The other is that there is an opposing current, the K⁺ current, that consists of positive K⁺ ions rushing outwards, balancing the sodium influx. You can see the two currents diagrammed here, the outward potassium current (I_K) in black, and the inward sodium current (I_Na) in purple. You can also see how fast these events occur, on the order of a few milliseconds.

What pyrethroid poisons do is mess up that purple line; they bind to the open sodium channel and prevent it from closing, so that the inward sodium flux continues for a longer period of time. This is not good for a finely-tuned, functional nervous system—it means the signals flick towards zero, and then stay near there longer than they should. It's a subtle difference, but since depolarizing a cell a little bit makes it more likely to send a signal, it means the whole nervous system is made hypersensitive, and storms of activity can go thundering through the whole thing. The poor bugs hit with this poison are taken out fast as the nervous system locks up, a process called knockdown.

You can knockdown lots of different kinds of insects with one poison because the sodium channel, the target of the pyrethroid, is highly conserved. The channel protein has a characteristic structure consisting of four domains that cluster in the membrane, forming a pore between them. The channel must have a characteristic shape and arrangement of charged amino acids in its interior to confer specificity: only Na⁺ is allowed through, not Cl^- or Ca⁺⁺ or K⁺ or any of the other ions wafting through the fluid-filled space around the cells. The molecule also has specific bits that mediate gating, and make it sensitive to the transmembrane voltage. It's not surprising that the sodium channel is highly conserved—it's an intricate apparatus with a lot of critical bits and pieces. There is variation, and the differences increase with increasing phylogenetic distance, but the overall structure of four domains forming a pore is found everywhere in animals, not just in insects, but in us mammals as well.

This diagram therefore applies everywhere in the animal kingdom. The sodium channel is made up of four domains (I-IV), each with 6 membrane spanning strands (S1-S6), that assemble to form the pore.

One of the useful things the Soderlund and Knipple paper does is put in one place a list of all the insect mutations that confer knockdown resistance. The idea is that what these mutations do is interfere with the binding of the pyrethroid to the channel, so mutations that block that binding are likely to be changes near the binding site. It's a way to map the location of the pyrethroid binding site, and here's the summary diagram:

They may look a bit scattered, but keep in mind that in the 3-dimensional structure of the protein, the four domains are folded together, and it's S5 and S6 that all line the interior of the pore, so these are less widely dispersed than the initial impression might give. The site labeled L1014 (S, H, and F are 3 different variants at the same site) is the most common target of mutations that confer knockdown resistance.

The M918 site is also interesting. All insect species happen to have a methionine residue at that particular spot in the loop between S4 and S5 in domain II; animals that mutate that particular amino acid acquire a profound resistance to pyrethroids. As it turns out, all known mammalian sequences contain an isoleucine at that point, and all mammals are resistant to pyrethroid toxicity (which is why it's valuable as a relatively selective poison). Converting that isoleucine to methionine in a rat makes the rat 10 times as sensitive to pyrethroid poisons.

To me, the virtue of the paper is in the elucidation of mechanisms and the discussion of variations in the sodium channel, but hey, you know that one of the goals has to be better, more effective ways of killing bugs (I used to work in an insect lab, and know a number of insect neurobiologists, and one of the unfortunate facts of life in that field is that it helps get grant money to mention something about finding new ways to enhance arthropod neurotoxins.) The authors tout one of the techniques used here, the heterologous expression assay, as a great way to characterize resistant mutants and identify poisons that circumvent the mutations. The heterologous expression assay involves extracting the sodium channel RNA from the insect, expressing it in frog oocytes, and measuring its properties and responses to toxins there. Frog oocytes are a great tool for measuring channel properties, because they are huge, with wide expanses of membrane, making it relatively easy to measure the ion fluxes of a particular, artificially expressed channel type. The idea is that we could refine a suite of toxins that blast different common insect knockdown resistance mutations, and then design a rational pesticide rotation strategy that kills the typical variants as they emerge from each cycle of selection.

That's one of the dilemmas of insect research, I guess, that you may love this nifty little critter you study, but one of your jobs is to find more thorough ways to eradicate them.

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Soderlund DM, Knipple DC (2003) The molecular biology of knockdown resistance to pyrethroid insecticides. Insect Biochemistry and Molecular Biology 33: 563-577.

Looking for more Tangled Bank entries....

It's Tangled Bank time again. I've sent a link to some science blogging to Tom of Wolverine Tom…have you?

Sunday, August 22, 2004

A distinguished early morning visitor

I got up this morning and went out the door, and there, waiting for me, was this huge beautiful creature clinging to the corner column.

I briefly entertained the idea that he was a contract killer on assignment for the Vespid Mafia, but nah...he was much too calm and polite. I'm going to pretend he's a dignified old ambassador from the Class Insecta, sent to grant forgiveness.

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