IL-6 - True Muscle Builder or Just a Measure of Workout Intensity? Plus: If Testosterone Does Not Matter, Why Does the Androgen Receptor Density Make a Difference?

No matter how close we look, the influence of previous, i.e. GH, IGF-1 and testosterone, as well as novel, i.e. IL-6 and AR expression, potentially growth promoting suspects remains elusive.

SuppVersity readers know, there is more to "inflammation" than the average mass media article will make you believe. The same "bad cytokines" that will decrease your insulin sensitivity, make you sick and obese, when they leak from your "inflamed" beer belly, are actually the good guys, when they are released in response to a workout from the musculature. "Myo-", not "cytokines", that's how researchers refer to them (Pedersen. 2007); and their role in (exercise) metabolism and immunity is until now still not fully understood.

The results the scientists from the McMaster University probably won't add much to our understanding of the systemic effects of IL-6 and other myokines. What certainly do, though, is to support the notion that "inflammation" can have profound and very far-reaching effects on our physiology (learn more).

Does IL-6 build muscle?

The mere idea that IL-6, a molecule of which most people think that it was a good measure of how messed up your health actually is, could be a promoter, or at least a measure, of skeletal muscle growth in response to a workout sounds about as logical (or illogical, if you will) as the previously established fact that the allegedly anabolic hormones testosterone, IGF-1 and growth hormone don't show the slightest correlation with the exercise induced skeletal muscle growth (West. 2011; learn more in "Anabolic Workouts Revisited").

Figure 1: Relative increase in muscle CSA change for androgen receptor density and p70S6K expression (left) and individual correlation of endocrine changes and muscle CSA (right), data based on Mitchel. 2013)

What was to be expected, though, was the correlation between the skeletal muscle hypertrophy response to the 16 week / 4x per week exercise program the 23 previously untrained study participants had to endure, on the one hand, and the exercise induced increase in p70S6K, on the other hand. The signalling protein p70S6K is after all something like the "protein pump"-gauge in the mTOR cascade.

Strength training can increase the androgen receptor density

Less well-known, but also not really new is another observation Mitchel et al. made: The resistance training lead to an allegedly subject specific and overall insignificant increase in the density (number per area unit) of androgen receptors; and though the overall increase may not have been statistical significant, the correlation of the the increase in androgen receptor density and the lean muscle gains of the subjects was.

New myonuclei (blue) are necessary to keep growing (learn more)

What about the muscle structure? What I am missing in this study is a measure of the myonuclei and domain sizes. We know that inflammation plays a major role in the restructuring process of the musculature that's necessary to maintain myonuclear domain size and thus the capacity for muscle protein synthesis (Bamman. 2001). It does furthermore seem likely that the any increase in myonuclei number would go hand in hand with increases in the number of androgen receptors. The corresponding data could thus help us to answer some of the questions we still have about the immediate (mTOR) and the chronic (endocrine and immune) contributions to skeletal muscle hypertrophy.

Similar effects on the androgen receptor density have been described by Willoughby & Taylor in 2004, already. In the pertinent study from the Baylor University, this increase did yet go hand in hand with increases in total testosterone and the free androgen index, of which the authors say that they were brought about by persistent increases in testosterone - increases in testosterone that were not observed in the more recent study by Cameron Mitchel, where the testosterone levels dropped (albeit non significantly) in response to the 2x2 upper-/lower-body split training.

"Our study corroborates previous findings that mean AR protein expression was not increased following resistance training; however, the response had marked heterogeneity, with some subjects showing a marked (1.5 - 2.5 fold) increase in AR protein content [...] Despite no statistically significant change in AR receptor protein content, there was a correlation between AR protein content with fibre hypertrophy. Our results suggest that changes in AR content may be part of a muscle-specific response present to a greater degree in responders and responsible for some (~25%) of the variation in muscle fibre hypertrophy." (Mitchel. 2013)

With regard to the correlation (I want to emphasis that we don't have enough evidence to do anything, but speculate about causative effects, here), it may be worth mentioning that the statistical significant association between androgen receptor density and the changes in muscle cross-sectional area was more pronounced in the "strength type" fast twitch (type II) than in the "endurance type" slow twich muscle fibers (0.6, p = 0.002 vs. 0.47, p= 0.023).

For IL-6, on the other hand,  it was the exact opposite. Unlike the number of androgen receptors per muscle area, the amount of interleukin 6 that was released or, as Mitchel et al. say, "filtered" from the muscle into the blood stream in response to the workout exerted a minimally more pronounced effect on the slow twitch "endurance type" fibers (type II).

So is IL-6 anabolic? It is counter-intuitive and would be premature to say that "IL-6 plays a causal role in skeletal muscle hypertrophy". This is particularly true in view of the fact that high baseline IL-6 levels that would be indicative of chronic vs. acute inflammation were "inversely correlated with fibre hypertrophy" and that IL-6 per se "is associated with both muscle protein breakdown and JAK/STAT signalling in satellite cells" (Mitchel. 2013).

Acutely (green circle) IL-6 will help build muscle, chronically it will make you sick (Muñoz-Cánoves. 2013).

It does thus appear to be more likely to assume that the increased IL-6 response is - just as the previously observed increase in cortisol in West et al. (2011), by the way - a marker of the workout induced strain, which does - in a non-overtraining scenario! - predict the adaptive response and thus the actual muscle gains.

The fact that the inclusion of IL-6 in a model prediction of the exercise induced hypertrophy response did not increase its accuracy would also point towards a corollary, not a causal involvement of a cytokine, the local (=in the muscle) production of which has also been implicated in the protective effect of exercise against insulin resistance, as well as increases in lipolysis and fatty acid oxidation (Pedersen. 2007).

In the end I have to admit that the study at hand does not really offer the material that would be necessary to formulate something like practical implications. What the results Mitchel et al. present in their paper can do is to reaffirm that skeletal muscle hypertrophy occurs in a(n at least) bi-phasic process with (1) an acute "inflammatory" phase where P70S6K (part of the mTOR cascade) determines and IL-6 (and thus inflammation) correlates with an increase in lean muscle mass and (2) a long-lasting "recovery" phase where the increase expression of androgen receptors could play an important role in the maintenance of the immediate gains in skeletal muscle protein. Your training should thus provide for both: An intense hypertrophy stimulus on the training days and lots of time and nutrients to recover on your off days.

References: 

• Bamman MM, Shipp JR, Jiang J, Gower BA, Hunter GR, Goodman A, McLafferty CL Jr, Urban RJ. Mechanical load increases muscle IGF-I and androgen receptor mRNA concentrations in humans. Am J Physiol Endocrinol Metab. 2001 Mar;280(3):E383-90
• McKay BR, De Lisio M, Johnston AP, O'Reilly CE, Phillips SM, Tarnopolsky MA, Parise G. Association of interleukin-6 signalling with the muscle stem cell response following muscle-lengthening contractions in humans. PLoS One. 2009 Jun 24;4(6):e6027. 
• Mitchell CJ, Churchward-Venne TA, Bellamy L, Parise G, Baker SK, et al. Muscular and Systemic Correlates of Resistance TrainingInduced Muscle Hypertrophy. PLoS ONE. 2013; 8(10): e78636
• Muñoz-Cánoves P, Scheele C, Pedersen BK, Serrano AL. Interleukin-6 myokine signaling in skeletal muscle: a double-edged sword? FEBS J. 2013 Sep;280(17):4131-48. doi: 10.1111/febs.12338. Epub 2013 Jun 18.
• Pedersen BK, Akerström TC, Nielsen AR, Fischer CP. Role of myokines in exercise and metabolism. J Appl Physiol (1985). 2007 Sep;103(3):1093-8. Epub 2007 Mar 8. Review.
• West DW, Phillips SM. Associations of exercise-induced hormone profiles and gains in strength and hypertrophy in a large cohort after weight training. Eur J Appl Physiol. 2012 Jul;112(7):2693-702. doi: 10.1007/s00421-011-2246-z. Epub 2011 Nov 22.
• Willoughby DS, Taylor L. Effects of sequential bouts of resistance exercise on androgen receptor expression. Med Sci Sports Exerc. 2004 Sep;36(9):1499-506.

Understanding Muscle Hypertrophy - Study Sheds More Light on Process of Satellite Cell Recruitement: SRF, IL-6, STAT3, COX2, IL4 + More Funky Acronyms With Important Roles in the Structural Component of Muscle Growth.

No pain inflammation, no gain? In the long(er) run this could in fact be true.

All of you who followed my advice to "like" the SuppVersity Facebook page and are thus keeping up with the numerous additional news I am posting there, should actually have seen the news item on the non-significance of the exercise-induced interleukin-6 (IL6) response for the exercise induced improvements in glucose metabolism (read more). The mere fact that the glucose metabolism of IL6(-) mice, which are mice who simply cannot express IL6, is still improved by "working out" does yet by no means preclude that the demonized cytokine does play a fundamental role in the exercise-induced systemic and local benefits. In fact, an even more recent rodent study would suggest that a certain degree of inflammation and the respective increase in IL6 immediately after a workout is even essential for persistent skeletal muscle hypertrophy.

As you may remember from the Intermittent Thoughts on Building Muscle Series there is more to skeletal muscle hypertrophy than the simple messages such as "increases protein synthesis by X%" that are printed in shiny letters on the boxes of hundreds of the currently available "natural muscle builders" on the real and digital shelves of the supplement vendors. One of these "mores" is the recruitement of satellite cells, muscle stem cells that are incorporated into the musculature to replace damaged myonuclei or increase the myonuclear density to allow for greater protein accretion (learn more).

Decreasing domain sizes = better function + higher growth propensity

"Hold on those are rodents and rodent studies are not relevant!" While it is a good thing to critically assess whether the results of a certain study can be species specific. The contemporary practice to question all rodent studies which are not part of your own cherry picked arsenal is getting onto my nerves. So, please check out the pretty analogues (short term unfortunately) human study by McCay from 2009 (McCay. 2009), before you stop reading after spotting the word "rat" in this article.

It is this process of satellite cell activation and incorporation of which Gwenaelle Begue and her colleagues from the University of Montpelier have now confirmed that it depends on the activation of the IL-6/STAT1/STAT3 signaling pathway in a prolonged 10 weeks resistance training scenario. In the course of the latter,  36 male Wistar rats were randomly assigned to one out of the following six groups:

• CTL2, CTL4, CTL10 (CTL = non-training controls, n = 6 in each group) and 
• TR2, TR4 and TR10, which were rats trained for 2, 4 and 10 weeks. 

The rodents in the TR-X groups were supposed to climb an apparatus with initially 50% later up to 210% of their body weight strapped to their back, five times a week. The load was increased every two days, if the rodents still managed to do "10 reps" = climb 10 steps and reached quite impressive levels of 120% of the body weight after two, 150% of the body weight after four and 210% after ten weeks of training.

Where is the rodent squat machine?

No rodent squat in the study at hand, but the "stair climbing" is a better full-body workout, anyways.

Now, this may not be as "realistic" a program as the rodent squat Aguiar et al. used in their 2012 study, but is is - and this is interesting - very similar to a test that has been done by many researchers with myostatin negative mice. As you will remember from the Intermittent Thoughts on Building Muscle Series (click here to read the pertinent part) those heavily muscled mice are unable to lift their own bodyweight, mainly because of the fact that the myonuclear domains within their muscle grew beyond a threshold where they absence of an adequate number of myonuclei per volume unit enders the muscle useless.

"Healthy" muscle growth does therefore require both, protein synthesis (increase in volume), as well as structural adaptations, so that the domain size does remain constant - at least!

"10 weeks of resistance training did not affect the myonuclear domain"

Against that background the last subheading, which is in fact a direct citation from the full text of the Begue paper is - contrary to what a non-SuppVersity reader could believe - good news. Very good news, to be precise:

Figure 1: Changes in fiber type ratios (left), cross sectional diameter according to fiber type (middle) and  fiber area per myonucleus (right; Begue. 2013)

As you can see in figure 1 (right hand side), there was even a small, yet statistically non-significant decrease in the fiber area each myonucleus had to control and that despite quite impressive increases of 77%, 92% and 100% in the cross-section of the type-I, type-IIa and type-IIx fibers of the animals (figure 1, middle).

Satellite cell recruitment, necessary of optional if you want to get big?

In this context, Begue et al. speficially point out that the "recruitment of additional nuclei derived from SC incorporated into muscle fibers" occurs parallel to the better known "resistance training induced enhancement of protein synthesis" that occurs "after the training session and last[s] up to 24–48 h in humans" (Bengue.2013). 

"Indeed, several works in humans have evidenced an increase in the number of myonuclei per fiber when fiber size increases approximately more than 25% (Kadi. 2004; Petrella. 2008). Thus, the myonuclear domain (i.e. the theoretical amount of cytoplasm supported by a single myonucleus in a muscle fiber) remained constant although a large increase in fiber CSA via the addition of SC-derived nuclei occurs." (Begue. 2013)

Since estrogen plays an important role in the regeneration of the satellite cell pool, it's pretty likely that you can literally "SERM your growth potential away" (learn more)

Notwithstanding the heavily quoted results of the 2011 study by McCarthy et al. in which the reasearchers were able to demonstrate that rodent muscle can grow even when it is satellite cell depleted, my personal conviction is that the latter process, i.e. the incorporation of new (not just even the replacement of damaged myonuclei is an obligatory prerequisite for persistent gains.

With +40% increased domain sizes, after only two weeks, it would have been interested to see how things would have developed in the subsequent weeks. I bet(!), the normal mice would have kept growing while their satellite cell depleted peers would have hit a plateau, where their own body woul have pulled the emergency brake aka myostatin (in this context, it's also interesting to remark that myostatin stops the proliferation of satellite cells and does thus indirectly divert the existing ones towards differentiation and incorporation into the muscle, cf. figure 2)

---

Bottom line: The study at hand delivers further evidence for the intimate connection between "inflammation" or rather the expression of the still demonized inflammatory cytokine interleukin-6 and the incorporation of "fresh" satellite cells into the muscle. With the latter being a necessary prerequisite to keep the domain sizes within functionally optimal limits while the cross section of the fibers is expanding (the muscle is growing), it is likely an (I want to emphasis that!) not yet disproven that continuous muscle growth requires satellite cell recruitment.

Basically you can think of it like the Army. While it is (or at least has historically been) relatively easy to find any recruits (=increase protein synthesis), people who are qualified to become officers and coordinate the actions of the rank and file are hard to find and without an adequate number of them you will end up with a chaotic mess instead of a powerful army. That's actually pretty much what happens to the myostatin negative mice, who may be able to recruit officers,... ah, I mean to recruit satellite cells, but simply outgrow the maximal pace of satellite cell incorporation.

Figure 2: IL-6 is the first myokine you should remember, it "wakes" the quiescent satellite cells up, he COX-2 activated IL-4 is myokine #2 and initiates the differentiation / incorporation process which will eventually result in the formation of a new nucleus. .

What, oh yes, of course! I had almost forgotten the unfortunately quite complicated connection to IL-6. If you take a parting look at the figure on the right, you will realize that a diagram explains things much better than I could. In fact, the "motor" of the whole growth business is the contraction induced expression of serum responsive factor, of which Guerci et al. have found in 2012 that it is the previously missing link between muscular contractions on the one hand and the expression of myokines, who happen to be the same molecules we know as "inflammatory cytokines" in other contexts. Il-6 and the COX-2 activated IL-4 are then getting things rolling (Guerci. 2012)... what? No, I cannot tell you whether taking antioxidants will block that, but I can promise you that you will learn more about this tie-in within the next 7 days, so stay tuned ;-)

What I can tell you in advance, though, is that strength and size gains of IL-6(-) mice are compromised (Serrano. 2008). So even if I would have to qualify my previous statement that satellite cells are necessary for continuous growth - one thing is sure: Their activation by IL-6 is necessary for optimal growth.

References:

• Begue G, Douillard A, Galbes O, Rossano B, Vernus B, Candau R, Py G. Early Activation of Rat Skeletal Muscle IL-6/STAT1/STAT3 Dependent Gene Expression in Resistance Exercise Linked to Hypertrophy. PLoS One. 2013;8(2):e57141. 
• Guerci A, Lahoute C, Hébrard S, Collard L, Graindorge D, Favier M, Cagnard N, Batonnet-Pichon S, Précigout G, Garcia L, Tuil D, Daegelen D, Sotiropoulos A. Srf-dependent paracrine signals produced by myofibers control satellite cell-mediated skeletal muscle hypertrophy. Cell Metab. 2012 Jan 4;15(1):25-37.
• Kadi F, Schjerling P, Andersen LL, Charifi N, Madsen JL. The effects of heavy resistance training and detraining on satellite cells in human skeletal muscles. J Physiol. 2004; 558: 1005–1012.
• McCarthy JJ, Mula J, Miyazaki M, Erfani R, Garrison K. Effective fiber hypertrophy in satellite cell-depleted skeletal muscle. Development. 2011; 138: 3657–3666
• McKay BR, De Lisio M, Johnston AP, O'Reilly CE, Phillips SM, Tarnopolsky MA, Parise G. Association of interleukin-6 signalling with the muscle stem cell response following muscle-lengthening contractions in humans. PLoS One. 2009 Jun 24;4(6):e6027. doi: 10.1371/journal.pone.0006027.
• Petrella JK, Kim JS, Mayhew DL, Cross JM, Bamman MM. Potent myofiber hypertrophy during resistance training in humans is associated with satellite cell-mediated myonuclear addition: a cluster analysis. J Appl Physiol. 2008. 104: 1736–1742
• Serrano AL, Baeza-Raja B, Perdiguero E, Jardí M, Muñoz-Cánoves P. Interleukin-6 is an essential regulator of satellite cell-mediated skeletal muscle hypertrophy. Cell Metab. 2008 Jan;7(1):33-44.

Omega 3 Attenuates Exercise Induced Rise in Inflammatory Markers, BUT is This Necessarily a Good Thing?

I want to take the results of a recent study (Bakhtyar. 2011) published in the Clinical Journal of Sport Medicine as an opportunity to readdress the question of whether or not the Omega 3 induced suppression of inflammation must be considered a good or a bad thing, both in view of athletic performance, as well from a health and longevity perspective.

Those of you, who listened to my interview on Carl Lenore's Super Human Radio show, will know that my understanding of "inflammation" is somewhat different from the mass market "explanation" of "a fire that causes damage". To be precise inflammation, or what scientists generally measure, is the release of signals (inflammatory markers) that tell immune cells to do their jobs. So, saying that inflammation is the root of all disease would be like saying that someone who calls the firefighters is to blame for the fire - but I am digressing from the topic at hand...

After administering 1.5g/day of an omega 3 supplement (experimental group) to every third of 45 previously untrained volunteers, Bakhtyar et al. found the subjects' "inflammatory" response to eccentric exercise to be modified:

The experimental group showed less elevation in TNF-α and PGE₂ immediately, 24, and 48 hours after exercise, when compared with the other groups. Significantly less elevation was shown in the concentration of IL-6, CK, and Mb for the experimental group at 24 and 48 hours after exercise. The experimental group also demonstrated a significant trend toward reduction in the plasma concentration of LDH immediately, 24, and 48 hours after the exercise program.

Now, what does this tell us about the training effect and health outcomes of the exercise regimen?

• With inflammation being a not yet fully understood prerequisite for muscular repair and hypertrophy, it would warrant further investigations like muscle biopsies and consistent training regimens with continuously monitored strength and muscle gains to conclude that omega 3 supplementation is beneficial in terms of physical performance.
• In view of the conclusions Pedersen draws in a recent review (Pedersen. 2011) of the role of exercise induced myokines, i.e. inflammatory markers released by muscles (myo- = muscle-), in chronic disease, blunting of muscular IL-6 release, which has been linked to muscular AMPK activation, increased glucose uptake and fat oxidation, omega 3 supplementation is probably counter-indicated in a health oriented exercise regimen, anyway.

I hope more scientists such as Pedersen will begin to question the current "anti-inflammatory" paradigm, so that major players in the medial landscape will be forced to take on their findings and stop portraying long chain polyunsaturated fatty acids in general and fish oil in particular as the savior of the fat and diabetic. Instead they should encourage people to finally get their asses off their sofas to induce exactly that amount of healthy, exercise-induced inflammation omega 3 supplementation appeared to suppress in the aforementioned study.