Tuesday, March 12, 2013

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.

  • 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.


Belgian Blue said...

The synergist ablation model in Mccarthy et al is deemed a maximally anabolic one, and the increase of a whopping 40% increase in muscle size in linear fashion with non-sattelite cell depleted animals up to that point makes it unlikely either group would have gained a whole lot more in subsequent weeks, or that a divergence would all of a sudden appear. I think it more likely that sattelite cells are not required for growth, but are required to maintain size in the absence of hypertrophic stimulus. All three studies that have conclusively shown that sattelite cells are not required for growth, were all models of continuous stimulation (myostatin deletion, Akt overexpression and synergist ablation), so while the scientific community at large has now accepted that sattelite cells are not required for muscle growth, they are most likely required to sustain that growth without stimulation (aka basal maintenance of current level).Instead of an army its more like an office building, where hard-working talented folks achieve great things (protein synthesis), but without maintenance and janitorial staff (sattelite cells) the place would fall apart during the time they aren't working.

I also think your last statement is worded wrong. The conclusion of said data is that IL-6 is required for growth, not Sattelite cells or IL-6 stimulation thereof.

Roy Nelson said...

I will see your acronyms and raise you some more...
IL-6 dare I say something about FO that zooms the IL-6 upwards?

Anonymous said...

All right Guys. Tell my it is so difficult to rebuild muscle mass after bone Marrow Transplantation? I lost 75lbs. 205 to 130. 5 years after, 150-155. I had 7 rounds of whole body radiation, can you say destruction of myogenic stem cells? During the course of my second treatment I received MTOR inhibitor drugs while I had cancer induced cachexia. Throw in 4.5 years of glucocortico steroids. Now I have CGVHD which means I have elevated TNF-a levels. I think the loss of stem cells is the key??

Adel aka Dr. Andro said...

probably the loss of stem cells and maybe still impaired generation of new ones could be a culprit. Also, in this particular case you will have lost muscle substance. That's very different from losing just protein from the muscle as you would if you were injured for a months or so. Therefore you are actually starting from zero (compared to a healthy rookie probably even the "negative zone"). Still, five years and 25lbs is a bit low of a gain. Did you have a comprehensive hormonal panel as of late?

Anonymous said...

Hi Dr Andro. Thank you for responding to my question. Now down to 148lbs
so only 18lb rebuild 4.5 years after transplant. My last hormone panel on 12/24/2012 results-
estradiol 29 upper limit for this lab 39pg/ml
total estrogen 148.3 high-115pg/ml upper limit
Testosterone 877 250-1100 ng/dl
Free test 162.2 35-155 pg/ml
Creatinine 1.38 1.2 upper limit. It has been like this since the transplant.
ast/sgot 42 15-41 u/l
alt/sgpt 68 17-63 ul Both a little higher than previous blood work. Heavy lifting prior to test?
Sed rate 14 0-13 mm/hr

On 4/6/2012
total testosterone was 1262 250-1100
free test 147.8 48-224
Bio avail 248.7 110-575

On 12/16/2011
Total testosterone 1411 250-1100
free testosterone 223.1 46-224 pg/ml
bioavailable t 429.8 110-575 ng/dl

Note I started 100mg test cyp about this time once per week. I made rapid gains for about 6 months 138lbs to 154lbs. Gains have actually reversed now as levels decline.


Anonymous said...

Loss of muscle substance? My MD said I was starting with less than I had as a teenager. Great I was 97lbs at 14. 205 at 47.

Adel aka Dr. Andro said...

nothing out of the ordinary at least in terms of hindering gains... but you should be aware that test doesn't really solve the problem of the loss of muscle substance. This is why you just "blew up" (=more protein per myonucleus) on T and do now lose everything afterwards. No structural changes, no persistent gains. I am yet not sure how you can solve that... did you ever have GH and IGF-1 tested?

Anonymous said...

Yes. I do not have the Numbers but I recall the Md saying it was very high. 222? Adel, could you explain what muscle substance is?

Adel aka Dr. Andro said...

check out my article on "growing beyond physiological limits" => http://suppversity.blogspot.de/2011/12/intermittent-thoughts-on-building_11.html

"substance" vs. amount of protein is like the number of balloons vs. the air in it; read the article, I guess afterwards you will have a better grasp of what I mean by that. Also take a look at the summary of the series and click yourself through the individual parts => http://suppversity.blogspot.de/2012/01/intermittent-thoughts-on-building_29.html

Anonymous said...

Got it, Endoplamic/sarcoplamic Mitochondrial proteins.

Great articles Dr Andro.