31 May, 2010

Goggles Goggles Goggles 1

Those are the beginning of my goggles. Looking back at them, I could probably have planned them out a bit better, but for what I had to work with I'm rather happy with how they're progressing. I even got out all the metal burrs out so that they don't poke little splinters into my face anymore! That being said, though, the steel is a bit uncomfortable to wear right up against my eye sockets, so I'm thinking some kind of padding is in order. I've been thinking maybe I ought to add some rolled flannel fabric around the rims, but then I'm not exactly certain how I'm going to get it to stick to the steel. Hot glue feels like cheating, but it may be my only option. The heat gun that I used to braze the steel ribbon to the steel pipe would set the fabric on fire, so unfortunately that's out.

There will be no nonfunctional gears on my goggles. Instead, I intend to add some iris mechanisms to both eyes (once I can bribe my way onto someone's CNC machine, anyone have some good cookie recipes they're willing to share?), green lenses, and at least 1 laser mounted on the side. I probably ought to also make a headband at some point.

How to make Really Heavy Glasses Frames:

1) Procure 1 3" steel pipe nipple, diameter to your specifications (I used 1.75"), with both ends threaded.

2) Sit down with calipers and devise needlessly complex formulas to make perfectly even angled cuts to get scalloped eye pieces.

3) Clamp the pipe to a firm surface.

4) Attempt to cut it with your Dremel, realize it will take about 7h to cut through it that way.

5) Get someone to teach you how to use the much more powerful angle grinder.

6) Attempt to use the angle grinder.

7) Observe that angle grinder has obliterated all planning of step 2.

8) Observe that angle grinder has kicked up a fine steel dust all over your face. Remove safety goggles and ponder your likeness to a raccoon for a couple minutes until you observe the twang that all the flying sparks left on the wall.

9) Continue angle grinding, creating several large burrs.

10) Finish cut.

11) Clamp down 1 of the eyepieces, burr side out. Change angle grinder head from cutting to grinding and apply to burrs. This may induce shrapnel. Eye protection is very important.

12) Get rid of all large burrs with angle grinder, then use Dremel to remove finer burrs.

13) Discover Dremel's polishing capabilities. This will set you back about an hour, but it won't get you much shinier.

14) Discover the 17 small burrs you missed. Go back over with a Dremel.

15) Cut slots for the 1/4" 14G steel ribbon. Thread precut lengths of steel ribbon through holes, manipulate with pliers into desired shape.

16) Rig up a vice on something fireproof.

17) Apply heatgun until goggles begin to glow, clamp down ribbon with long-handled pliers and apply some sort of binding flux*. You may have to do this sequentially, in which case it should be noted that the heatgun does not cool down rapidly.

18) Continue until all pieces are in place.

19) Be satisfied with progress for now.

20) Write self-deprecating blog post later about project.

*Yes, strangely enough this part where I was quite competent, but not with the angle grinder.

27 May, 2010


ResearchBlogging.orgWhen I came across this post "Medical Advice for Headbangers" on Boing Boing today, I couldn't help but click through to read the paper. What I found was a pun-fest of scholarly research, and I'm left intensely curious about who funded this research. Ironically enough, at the time I came across the post I was listening to an auto-swung version of Metallica's "Enter the Sandman" (songs run through a rather neat Python script to swing them*).

When Toaster was a young whelp in The Ozarks, it eventually came to that time in his life where he, like every young person in America, is contractually obliged to find something to annoy the hell out of their parents and stubbornly persist at it until they move out. Rather than drink illicit alcohol** and crash cars into trees, I chose to join a death metal band playing bass. As a direct result of this, I began listening to a lot more heavy metal music and going to concerts. I was never much for headbanging*** because being a wallflower is more fun, but I saw a lot of other people pursue it aggressively like a cocaine-addicted lab rat and I noted that over time these people gradually became a bit dimmer than they'd been when I met them. They were also almost always much less coordinated after they'd been to a concert.

This paper developed a mathematical model to explain why my peers were left so hammered after headbanging so much as they did. They analyzed the way people move when they headbang and developed an equation for a sinusoidal wave to describe peoples' head movements during headbanging and subsequently estimated the level of force experienced as described by the Head Injury Criterion (HIC) from the angular velocity of headbanging heads. Although the HIC projects serious injury is only likely to occur at rates of head movement/collision over 15m^2/s. However, since headbanging is a repetitive motion, they chose, with evidence, to evaluate any rate over 8m^2/s as potentially injurious. This means that the faster you headbang and the wider angle of motion your head travels through as you do so, the more likely you are to hurt your brain.

Now the methods get a bit strange. Not only did the researchers attend a couple metal concerts to observe headbangers in their natural element, but they also analyzed the way that the "legendary" headbanging duos Beavis and Butthead and Wayne and Garth headbanged. They concluded that Butthead was the one most likely to be injuring himself.

However, I drew some issue with the music they were calling metal. From the period of my youth described above, I considered such bands as In Flames, Dark Tranquility, Strapping Young Lad, and their doomy ilk to be metal. I'd never considered AC/DC or the Ramones to be music to headbang to because it was just hard rock and punk, respectively. It should be noted that metal comes in more flavors than anyone knows how to classify, but the primary ones are the American model (Static-X, Rammstein), the Scandinavian model (Hammerfall, Nightwish), and the experimental sort (Finntroll, Sleepytime Gorilla Museum).

In conclusion, they recommended that public health agencies issue headbanging warnings on headbanging-worthy CDs, issue neck braces to limit the range of neck motion, and advise that metal bands have tutorials before their concerts. This leads readers of the paper to conclude that the authors themselves have absolutely no metal cred, because everyone knows that the only way they'll ever get concert-goers to wear neck-braces is if they become a metal fashion statement.

*Swing is a musical concept that relates to the delay between upbeats and downbeats. Upbeats and downbeats refer, respectively, to beats 2+4 and 1+3 in 4/4 metered music. Much of rock has the space between all upbeats and downbeats even, but in jazz and some hiphop the space between them is staggered to create a "swing feel".
**See ***.
***OK, it was because I am and always have been a nerd and didn't want 1) to damage my brain or 2) lose my glasses. I knew what brain damage felt like, I was a clumsy kid and had been through a couple concussions--and headbanging felt far too similar to ever be enjoyable. Can't help but to feel a tinge of smug right now with science validating me choosing to almost entirely abstain from it.

Patton, D., & McIntosh, A. (2008). Head and neck injury risks in heavy metal: head bangers stuck between rock and a hard bass BMJ, 337 (dec17 2) DOI: 10.1136/bmj.a2825

24 May, 2010

T-Cell Receptors

I've been thinking about T-cell receptors (TCRs) a lot lately, primarily because they're a delicious, tangled little knot of wonderful complexity. Some (Sciliz, I'm looking at you) call them fickle, but no, they're far tastier than that. Synaesthetic metaphors aside, TCRs are the gate-keepers between strong and solemn anti-pathogen protection and a raging inferno of doomy autoimmunity. More specifically, the genes that encode TCR are divided up into Lego-block-like segments [V, D, and J segments] that are sorted more-or-less-at-random*. This happens in each developing T-cell, and the genes undergo some further random mutagenesis via the RAG proteins to generate completely unique receptor specificities. In turn, this means that each TCR is unique, like a precious little snowflake or puppy nose, and it guards the body against its cognate antigen vigorously**. However, much unlike a snowflake or puppy nose, the mechanism by which TCRs generate useful signals and screen out non-specific noise is absolutely badass. In fact, rather than naming battleships after dead guys, who are no longer badass, perhaps we'd be better suited to name warships after T-cell receptor clone lines: Jurkat, D0.110,...others?

Anyway, I've been thinking a lot about the applications of network information theory to TCRs lately. But that's not what I'm going to talk about today, because in thinking about TCRs like that I've recently come to appreciate just how absolutely unrefined, perhaps even crass, my initial understanding of TCRs was.

Back when I first took an immunology course through my university's medical school extension thing, this was the impression I got of TCRs. G-protein coupled receptors (GPCRs) had been beaten into our skulls with the Alberts' Molecular Biology of the Cell text and as such I figured it worked a lot like a GPCR. This turned out to be rather incorrect. In fact, I was also under the impression that almost everything in the world was some sort of GPCR***. Fortunately, this too turned out to be incorrect.

The CD4/8 molecule pictured above is a rather handy cell-surface marker that is expressed, respectively, on CD4+ and CD8+ T-cells, which in turn respectively recognize antigen in MHC-II and MHC-I proteins. If that didn't make any sense, don't worry about it, what's important here is that CD4 or CD8 help the T-cells see the antigen that elicits a signal. CD4 and CD8 are co-stimulatory molecules without which antigen-presenting cells cannot effectively transmit information.

Anywho, as evolution would have it, there are more proteins involved in TCR signal transduction.
At the time I learned of the existence of CD3 and CD28, I had absolutely no clue what they did, so I merrily accepted that they were part of the TCR complex and moved on. Why I didn't look further into them is currently beyond me, but the most likely answer is that I was distracted at the time by free cookies. As such, I thought that signal transduction just carried on with CD3 and CD28 as handsome bookends.

Little did I know of the strange superpowers these two molecules have. Eventually, because science enjoys knocking over the obelisks of my ignorance despite how hard we've labored to keep them intact****, I gained that knowledge.
I couldn't believe it. It was like when I found out that I couldn't eat clouds. Signal transduction WITHOUT antigen stimulation? What sort of stochastic heresy was this? This shattered the beautifully deterministic world-view that my undergraduate education had instilled, that every receptor has a ligand that can be defined and that the cell is a place of beautiful complacency and order. I had known that ligands::signals weren't always 1::1, and I had accepted that, but this, this was somehow darker and more malevolent, and the knowledge that TCRs could be so deftly manipulated and easily fooled was tantalizing. This assumption, also, turned out to be wrong. TCRs are stubborn little bastards.

Nonetheless, I set aside my beloved copy of Janeway's Immunobiology and leaped into the scientific literature face first. I read cell biology papers, I read biochemistry papers, I read biophysics papers, I read autoimmune pathogenesis papers, I read genetics papers, I may have read some enzymology papers, and I know I read a bunch of computational biology papers. I emerged with a mouthful of bistable switches and nonlinear dynamics, and I believe that my understanding of molecular biology as a whole was greatly enhanced by it. Cells weren't orderly little machines, they were complex and messy tiny mechanisms that had error and chaos built in as part of their very functions. And that chaos and nonlinear responses can be harnessed by evolution to increase the sensitivity, response range, and efficiency of cells was absolutely beautiful in its inhumanity. Humans build orderly structures, boxes and flat surfaces and well-engineered machines, but cells don't given a damn! Cells get by, thrive, and capture the very entropy we try so hard to scrub out of our lives, and that, that right there was something that I didn't appreciate until I wound up climbing up atop the tallest stuff at hand one night (Halloween, exactly) and laid sprawled out on top of it watching the sky. Seeing the moon reel about the sky and the stars wheel away above it while the earth beneath me tangibly spun made me feel so very small and tiny and insignificant in the face of such a vast expanse as the universe. But then, at the same time I knew that I was composed of billions of microscopic cells to the point where we are complex molecular galaxies unto ourselves and that, too, made me feel small and insignificant. And lastly, knowing that the stars in the sky and the cells in my body will get by without giving a damn about whether I know they exist or appreciate them was a sort of beautiful thought that has remained with me, probably as the closest thing to faith that I've found.

Epic literature reading aside, the mechanisms of TCR activation as I now envision them are rather complex to draw, so I am going to use art instead.

Apotheose der während des Befreiungskrieges für das Vaterland gefallenen französischen Helden, by Anne-Louis Girodet-Trioson

This isn't meant to be tongue-in-cheek about it being epic*****, but more that it is incredibly crowded and busy and spatial location matters a LOT. You see, resting T-cells have TCRs scattered across their cell surfaces more or less at random, and each of those TCRs is identical. But once 1 of those is conjugated with the triggering antigen at just the right specificity, they all suddenly (within a matter of mere minutes) condense over to the site of that triggering and form an immunological synapse.

The synapse polarizes the TCRs all to 1 side of the cell and clusters them together. Actin also gets rearranged. This provides a cell surface anchor point for intracellular proteins to begin forming a complex cascade. The TCR proteins themselves change shape and CD3's intracellular domains are released from the cellular membrane where they were held. CD3 can then serve as a scaffold for effector kinases such Lck and Fyn. Some JAK/STAT scaffolding proteins eventually transmit the signal from Lck/Fyn/Zap70 to ERK and JNK, which in turn then affect transcription of NF-kB and other response genes such as IL-1b. At the same time, there is a massive calcium flux across the cellular membrane.

Like this. I had to take considerable liberties in omitting details to make this fit on one slide.

However, the synapse is presently more interesting than its effector functions. The immunological synapse is actually composed of 2 parts, the central supramolecular activation complex (cSMAC) and the peripheral supramolecular activation complex (pSMAC). As you might have already imagined, the cSMAC is surrounded by the pSMAC. Within the synapse, many T-cell surface proteins are able to interact with their ligands on the surface of the antigen-presenting cell and vice versa. The SMAC structures are stable (relatively for T-cells) and allow the antigen-presenting cells to properly stimulate the T-cell with both its cognate antigen and some pro-proliferation cytokines such as IL-2.

Now here's where I really have no idea how to visually illustrate it. Within the cSMAC, TCRs are systematically obliterated as the activated, presumably phosphorylated?, TCRs get pulled into the cell and chewed through proteasomes to allow fresher TCRs to get at the antigen-presenting action. This degradation of activated TCRs from the cSMAC is essential for proper T-cell activation (if you block it the T-cells keel over and die, as they are wont to do). And this is what is really frackin' cool: bistable switch behavior in the activation kinetics of TCRs is mediated first by formation of the immunological synapse, and then sustained by endocytosis and degradation of activated TCRs.

Toaster, what the blugoon is a bistable switch?

Hysteresis can be found in a lot of situations, including magnetization, memristors, and a whole lot of very useful electronics and materials sciences.

A bistable switch exhibits the property of hysteresis, which is rather awesome. Hysteresis essentially means that a signal going 1 way across some response action (sensor) doesn't necessarily get the opposite response when the signal goes the other way.

Hystersis can be found in TCRs at several levels. First, it is part of the kinetic proofreading that occurs at TCRs prior to cSMAC formation. TCRs have to be extraordinarily sensitive to their cognate antigen and must be vigilant in screening out false positives. Therefore it takes a stronger signal to activate TCRs than it does to deactivate them. Second, use of hysteresis in this is an efficient way of screening out false positives; the formation of the cSMAC and subsequent degradation of activated TCRs both help to sustain hysteresis behavior. Condensation of TCRs into a cSMAC requires a stronger activating signal than deactivating signal, which means it will remain active at a signal weaker than that which activated it. Thirdly, degradation of activated TCRs helps sustain hysteresis by ensuring that the signal is propagated into the cell at a proper rate. Too fast and the T-cell's built-in anti-autoimmunity mechanisms will kill the whole cell, and too slowly risks death by anergy, while still the net signal from an antigen presenting cell must be higher to activate the T-cell than to deactivate it.

There are many finer details of TCRs that I did not cover here, and still more that I probably don't yet know about. T-cells are an essential part of the immune system, able to differentiate into several different useful phenotypes. CD8+ cytotoxic T-cells can circulate around the body and survey for viral infection or cancerous abnormalities. CD4+ Th1 cells secrete massive amounts of IFNg and help direct innate immune effector cells to combat bacterial infections. CD4+ Th2 cells secrete lots of IL-4 and are involved in battling away parasites or, more relevantly in the Western world, causing allergies and asthma. CD4+ CD25+ regulatory T-cells shut down the immune response by secreting IL-10 after the pathogens have been cleared away to prevent Th1 T-cells from tearing up the place (see: cytokine storm). Memory T-cells lurk around and provide lasting immunity to previously encountered pathogens. And Th17 T-cells do something, they seem to be involved in protection against autoimmunity but we're not quite sure yet. Each of these T-cell phenotypes relies upon the TCR to detect their cognate antigen and help defend the body against the invading microbes that find its squishy, nutrient-rich nooks so very appealing. At the population level, I sometimes think of T-cells as a library of exquisitely-finely-tuned peptide detection machinery that we all carry around with us, and I find that chaos they usefully harness in hysteresis to be beautiful and captivating. There may be other, as of yet, undiscovered T-cell phenotypes and functions despite an already broad and deep literature, and that is an exciting prospect. T-cells and the exact mechanisms of their TCRs remain a relatively open biological frontier, and the best we can do is to dive in face-first.

*Not quite, but explaining it goes beyond the scope here. Look here for more information.
**If it doesn't get killed off first, as the grand majority of developing lymphocytes do, due to anergy or too strong a reaction against self.
***I really liked imagining that there were little Ggamma subunits shuttling around everywhere, all the time, in absolutely everything making a "BlootablootalootaLOO!" burbling sound as they went, with tiny ADP bubbles behind them.
****This is an obligatory joke notification footnote.
*****Also to help break up large blocks of text.

04 May, 2010

Atari Punk Circuit

The Atari Punk Circuit was designed as a beginners' project by the venerable Forrest Mims, III. I built it as a prototype of a much grander project still to come, once I get around to making those flex resistor circuits for it. The Atari Punk circuit uses 2 555 integrated circuits (or 1 556), which is a timer circuit, and slaps resistors downstream of the inputs to vary the rate at which it pulses and capacitors on the outputs to modulate that pulse into a continuous and audible waveform. In this iteration I used 2 10K Ohm linear potentiometers as resistors. The 1st one modulates the frequency of the output wave, thus changing pitch; while the 2nd one is bridged across 2 input pins to provide destructive interference and noise upon the 1st wave, thus making buzzy distortion.

Shortly after running it in the video above, I connected this circuit to a power supply without checking what it was set to first, and promptly burnt one of the 555s out. Smoke and everything. Luckily, I have a replacement and will be putting it back together once I've wired up some flex bars in the place of the current linear potentiometers, even though linear potentiometers are dead sexy.

I have no idea how this relates to molecular biology. Yet.