α-Ketoglutarate & Lifespan, Really?

So this paper came across my desk this weekend (yes it was published a month ago so I’m a bit late to the game here, shoot me). I’m having a hard time believing the central result isn’t due to a simple artifact.

The paper describes the ability of α-ketoglurate, an intermediate in the Krebs’ cycle, to extend lifespan in C. elegans. The authors let the worms swim around in a solution of 8 mM α-KG and they lived longer.  There are some interesting knockout experiments showing the importance of various downstream mediators (mTOR, ATPsynthase) in the effects, but the key problem here seems to be that the authors may have overlooked a potential alternative mechanism…


Yeah, so side-stepping the issue that just about every small molecule has at some point in the past few years been tagged as an antioxidant, there might actually be some decent proof here, in the form of a chemical mechanism. You see, a few years ago I reviewed (and was subsequently asked to write an editorial on) a paper in Free Radical Biology & Medicine, showing that α-keto acids (e.g., α-KG, pyruvate, oxaloacetate) can be non-enzymatically decarboxylated in free solution by reaction with hydrogen peroxide.  The reaction is as follows:

α-KG + H2O2 → succinate + CO2

As outlined in our editorial, what this amounts to is a short-cut in the TCA cycle… you can get from α-KG to succinate without α-KGDH, so if the enzyme is inhibited (coincidentally this may occur due to oxidative stress such as H2O2), there’s a way to bypass the inhibition and allow the cycle to keep turning. Furthermore, this property of α-keto acids is well known in experimental circles… if you’ve ever made up a solution of pyruvate you know it goes off pretty quickly. That’s oxidative decarboxylation in action (the product in this case would be acetate). We even showed that this reaction (oxaloacetate → malonate) could occur in perfused hearts and may play a role in ischemic preconditioning. So, it happens in real mammalian tissues and is not just a theoretical test-tube reaction of the type often wheeled out to show that something is a biologically relevant antioxidant.

So, overall it just strikes me as not a very big deal that adding 8 mM of a known antioxidant to a plate of worms has an effect on lifespan.

The dose response shows an effect on lifespan in the 1-10 mM range with an EC50 of ~3 mM, which is right in line with a generic non-enzymatic effect, and not really compatible with the KM values for most of the enzymes that handle these metabolites (typical KM for α-KGDH is 150 μM in the presence of Ca2+, only going into the mM range in the complete absence of Ca2+ which never happens in-vivo). Moreover, Fig. 2 of the paper shows that many of the effects which are proposed to account for the lifespan phenotype at the whole organism level occur at concentrations of α-KG in the 100-200 μM range, i.e. 5-fold lower than the level used in live worms). Oh, and it’s probably worth mentioning that some of the proposed downstream mechanisms might be sensitive to oxidative stress.

Given all the kerfuffle in recent years about TCA cycle intermediates as hot sexy new signaling molecules, it’s hardly surprising that this study made it into Nature. However, it’s frustrating that some simple controls were not included… (i) Do other α-keto acids prolong lifespan? (ii) Does adding α-KG to media and letting it sit for a few days at room temp’ on the shelf diminish the effect? (ii) Are lifespan-extending effects of other antioxidants additive to the effect of α-KG? (iv) Are effects of α-KG greater in worms known to generate more H2O2?

I left a comment on this at PubMedCommons, and also at PubPeer. The latter notifies the authors automatically, so hopefully we can get some feedback on this.

Summer Happenings

It’s been a while since I updated the lab blog, so here’s a run-down of what’s new in the past few months…

  • Marcin Karcz, a resident in the Department of Anesthesiology, got his fellowship grant funded by FAER (The Foundation for Anesthesia Education and Research), and will start in the lab shortly.
  • Our paper with Michael O’Reilly on mitochondrial DNA double strand breaks and ATM activation was finally published in FRBM.
  • The publications page is finally updated to include our 2013 and YTD 2014 papers.
  • We have a summer undergraduate intern, Ling Yang from biomedical engineering, who is designing/making circuits for electrical stimulation of isolated cardiomyoyctes.
  • Our dual-PI R01 application with Keith Nehrke on the mitochondrial unfolded protein response received a 13th percentile score. Not fundable (NHLBI payline is 11%) but hopefully close enough to fly on the next cycle.
  • Chad Galloway got an impact score of 30 for his K01 application. No percentiles given, so it’s fingers crossed for the October council meeting at NIDDK.
  • Chad also got his paper on mito’ morphology in NASH published in AJP Endo (accepted but not online yet).
  • AHA Grants have been flying out the door… pre-doc’ from Owen Smith, and lots of letters/references for pre-doc’, post-doc’, grant-in-aid and others. It looks like their fall review cycle is going to be packed!
  • Andrew and Sergiy went to the AHA Basic Cardiovascular Sciences meeting in Las Vegas NV, where allegedly Sergiy won a whole 5 dollars (and promptly lost it again).  Oh, and Andrew won a travel award.

Outside the lab’ there’s plenty happening too.  Summer vacations being taken, home improvement projects being done, happenings in the world of PPPR such as this great editorial at PubPeer, trying to get letters/paperwork together for my promotion, playing with my 3D printer, and of course the usual shenanigans over on Twitter.

Sensationalism in science reporting

Being British, I have a certain reverence for the BBC, which was arguably at some point in history the finest news organization in the world. That’s certainly no longer the case, especially in science reporting.

Being a cardiac biologist, my interest was piqued by the following headline… “Mini hearts grown to study disease“, but the story itself is less actual science reporting, and more lab/university PR.  Such material does not science news make. A better headline would be “Dude from obscure Scottish university figures out how to do something already done“.

The first red flag that this news piece is nothing more than PR, is no link to a report of the underlying science in a peer reviewed journal. The most fundamental bar for something being newsworthy in science is journal publication. How did this even get past the BBC proof readers?

Well, maybe the link to the authors’ University webpage* can help? (oddly it was put at the bottom of the piece, not embedded in the text of the article itself).  Nope, nothing about cardiac stem cells there.**

What about PubMed?  Nope. 5 papers published since 2007, none of them about hearts.

Then there’s this statement… “They are indeed human cells, which physiologically are the same as human hearts, in this case the size does not matter”. Ever heard of pre-load?  Afterload? Frank-Starling? Oxygen tension?  Does anyone in the BBC science department even understand the meaning of the word “physiology”?  Resisting my temptation to make a puerile joke that size DOES matter, let’s just say a ball of cells is about as far removed from a living beating human heart, as a naked mole rat penis.

The kicker is this tag-line at the end… “We can work now, in one experiment, with 1,000 human hearts and test large amounts of compounds, which you can’t do in animals”.  While ignoring that you actually CAN do this in animals (you just need 1000 animals, which isn’t a lot for a mouse lab), the key word here is “now”.  NOW we can do this.  Before we couldn’t do this.  Everything hinges on novelty, but as the link above to the pioneering work of Chuck Murry shows (as does a quick search for “heart on a chip”), this is far from novel.

None of the above is to detract from the actual work of Dr. Zhelev. I’ve never met him, and his work is probably awesome. Hey, he got cardiac stem cells to grow in a dish, which is more than I’ve ever done with stem cells, so hats off to him!  But please BBC science news, try and recognize when you’re being used for blatant PR with zero underlying content.  As a scientist, I like a little more meat behind my science reading.


*Who even knew there was such a thing as the University of Abertay? I come from the other end of the Sceptred Isle and had never heard of it before. I’m sure it’s a great place, but maybe someone in their science PR office needs to lay off the caffeine for a while.

**Lordy! Those are some freaky cold lookin’ folks at the top of the page. The dude in the middle looks like he just got out of a knife fight. That one with the big eye in the magnifier is giving me the fear!


Administrative Bloat

This is not so much a rant, but a post intended to give some idea of the enormous amount of administrative burden a typical PI at a research university has to deal with every day. For the first 3 days of this week I accomplished absolutely zero science whatsoever – no experiments, no paper writing, no data analysis, nothing. Instead, I spent 3 solid days (8am-5:30pm) trying to rid my email in-box of administrative tasks…

  • Wrote a letter of recommendation for someone I’ve only ever met once and who didn’t send me her CV when requesting the letter. Typically do at least 2 such letter a month.
  • Submitted 2 animal protocol modifications via what is quite possibly the most arcane online system imaginable (Granite/Topaz – only works in IE, frequently crashes or just doesn’t load at all).
  • Submitted protocols again when they came back with questions. Then submitted them a 3rd time just now.
  • Bought plane tickets for a visiting lectureship I have to do next week. Admin’ assistant at the other university had left, and her replacement didn’t contact me to arrange anything. Still haven’t received any itinerary or idea of who I’m meeting with during my visit. They want me to take a cab from airport to hotel when I arrive at 1pm (seriously, there’s no-one available to meet with?)
  • Complied powerpoint slides for the above.
  • Processed paperwork for getting student reimbursed for a trip he made to a collaborator’s lab. Apparently students have to use a different form than the one everyone else uses. Does it say “not for students” on the form we used first time? Like hell it does.
  • Drafted 2 thank-you letters for the reviewers and readers of a grant panel that I co-chair for a charity.
  • Submitted an NIH progress report via the delightful new RPPR system. This is the replacement for eSNAP, and hey why use a simple 2 page form when 7 pages with multiple tabs will do?
  • Related to this, 2 weeks ago I submitted manuscripts via the NIHMS system, to make sure they’re in compliance with the public access mandate. Apparently it takes up to 6 weeks for each manuscript to be assigned a PubMedCentral ID. The business admin’ at my institution contacted me at 5:30pm the night before the progress report deadline, claiming they could not submit the report due to non-compliance. Some advance notice of this might have helped! (Thankfully they submitted anyway, but now there’ll be a delay in issuance of the NOA).  All-told over the past 2 weeks this progress report involved >30 emails back and forth.
  • Just to confuse matters further, the NIHMS website, the RPPR site, and the MyNCBI site (used to enter papers into NIHMS), all use the exact same login as NIH eRA commons. As such, at one point I was logged into all 4 websites simultaneously with the same login ID. If this isn’t a major security flaw I don’t know what is. Could they make it any more complicated?
  • Had an annual evaluation meeting with my lab’ tech. This year Human Resources replaced the simple 1 page form with a new 6-page PDF, full of really insightful questions (Q. How does the employee behave toward patients and customers? A. They don’t because we’re researchers with no patient contact!  Q. How does the employee add value to the institution’s mission? A. Maybe if the institution didn’t re-write their mission statement every 6 months I’d be able to answer).  The PDF “lost” some information during submission (different versions of Acrobat), so my Dept’ admin’ had to print it out and bring it to me to fill in the missig info’ and sign it manually.
  • In addition to the 6 page forms (one each for me and the employee), there was a job description form, an education record, a HIPAA statement, an “ICARE” employee values commitment statement, and an “age specific competency statement”.  6 items in total, more than half of which was about patient care duties, for a research lab’ tech with no patient contact whatsoever.
  • To get it out of the way (to make room for more admin!), I mistakenly decided to do my Mandatory In-Service exam. This is something all employees have to do to prove they understand various rules (what “code blue/pink/orange” on the intercom means, etc). The course is administered in BlackBoard (vomit). The first part of the test is an assessment of roles to determine which test you have to take. Despite qualifying as having no patient contact, my test (65 questions, 58 correct required to pass) included questions about wearing lifting gear to transfer heavy patients, how to deal with patients with guns, how to find an interpreter, etc. Who designs these things? I passed, but seriously what a waste of everyone’s time.
  • Until recently I was paid 5% off a grant that involved human subjects research, and even though I had no involvement in that part of the project, all members of the team were required to have human subjects protection training. I’m no longer involved with the project, but got a notice to renew my certification last week. The test is administered via a 3rd party (CITI). Given that I hadn’t done the test in about 4 years, the first step was to recover my password for the site.  1st attempt – nothing after 15 minutes. 2nd attempt – nothing. 3rd attempt – bingo there’s the email!  Reset my password and took the test. It was littered with grammatical errors, dead links, obtusely worded questions. I had to stop half-way to attend to something else. At 11pm (6 hours after first logging in), I got 2 more emails for password reset. When I tried logging in the next day it wouldn’t let me (because of the password reset requests – doh!) so I had to reset twice more and then resume the test – quel surprise – some of my answers had been lost.
  • Processed paperwork for getting RAP’s professional society membership refunded from a discretionary account. But account was overdrawn…
  • Last fall an equipment purchase resulted in the account being charged twice and going $5k overdrawn. Finally (5 months after being requested!) accounts payable got their act together and refunded, so now the account is in the black it can be un-frozen for me to actually spend on.
  • Reviewed 2 de novo manuscripts for journals, and a 3rd that was a resubmission.

There was a bunch of other stuff interspersed too – follow ups from lab’ safety inspection, scheduling of June NIH study section and assignment of proposals to review, lots of dealing with journal editors in my ongoing activities in the area of scientific integrity, plus minor stuff like responding to emails, signing bits of paper.

But where’s the science?  During the past 3 days (well actually a week because I was out of town reviewing grants Thu/Fri last week) I’ve done nothing that I’m actually paid to do. No experiments at the bench, no reading papers (except on the bus), no discussing data with my lab, no thinking about future ideas for experiments, no writing exam questions or updating teaching materials.  Just hour after hour of mind-numbing attempts to get my email inbox cleared of this constant barrage of administrative clutter…  If it’s not paper review requests then it’s grant reviews, travel arrangements, departmental paperwork, grant submissions, society work, personnel and HR stuff, committees, legal/compliance/oversight paperwork, newfangled forms, accounting errors, chasing up lost orders, etc. It never ends. Never.

I did it in the end (got down to 3 emails, all of them from collaborators and talking actual science). This catharsis is supposed to make me feel accomplished, serene, refreshed, ready to do and talk science. But instead all I feel is anger at the wasted time, depressed at how this is going to happen all over again in a few days. Administrative bloat is sucking my will to live. It is slowly killing me and everyone around me, one email at a time.

P.S. required reading for anyone enraged by this topic

Is it spring yet?

It’s been a long and cold winter in Rochester, but there are still lots of interesting things going on in the lab’, so we don’t have to think about the weather too much…

– Jimmy Zhang (MD/PhD student who rotated in the lab’ last year) has decided to come back and pursue his PhD with us, starting this June.

Marcin Karcz, a resident in the Department of Anesthesiology, wrote a fellowship grant proposal to FAER (The Foundation for Anesthesia Education and Research), so if that gets funded then he’ll be joining us for a year of research starting this July.

– Isaac Fisher, a Pharm/Phys graduate student, is just now completing his rotation project, doing some drug screening for regulators of cardiac metabolism.

– Paul’s paper entitled “Internet publicity of data problems in the bioscience literature correlates with enhanced corrective action” was accepted at Peer J, so should be in press by the end of March. The title pretty much says it all, but I’ll post more details once it’s out.

– Paul gave an interview to Science Careers, about how to walk the fine line between being a whistle-blower and maintaining an academic career.

– Andrew Wojtovich resubmitted his K99/R00 application, and Chad Galloway resubmitted his K01 application. Also Andrew’s review on optogenetic control of ROS came out in the new SFRBM open access journal Redox Biology.

– On March 12th URMC is hosting Nicholas Steneck, a guru in the area of research integrity (having been involved in the establishment of ORI guidelines on responsible conduct of research), for the annual bioethics visiting lectureship. This is part of a one day CTSI workshop about ethics in translational research.

– Graduate student Owen Smith is off to Yale for a week, to work with our collaborator ‘Liz Jonas on mitochondrial ion channels.

– Most of the lab will be headed to Michigan in May, for the wedding of former graduate student David Hoffman, who now works at Cayman Chemical.

– Now if we could just understand our metabolomics data…..