Dienstag, 25. März 2014

Does reduced GH/IGF signalling extend lifespan? A possible answer.

Berryman et al. (Kopchick lab) recently published a paper (1) that I sort of disagree with. I'd like to take the opportunity to discuss a bigger issue: how can we know the true effects of GH-inihibtion, at least in mouse models? Isn't the data conflicting as Sonntag et al. have shown?

Let's start with a brief introduction to long-lived dwarf mice (1):
"[T]here appears to be a preferential accumulation of excess fat in the subcutaneous depots, which is thought to have metabolically beneficial effects in ... long-lived mice and that may contribute to their counterintuitive association between obesity and life span."
Then the authors say that:
"Not all mouse strains with reduced GH or IGF-1 action show improvements in life span. One example is GH receptor antagonist (GHA) transgenic mice, which were generated in our laboratory more than 20 years ago (22–24)...
The GHA expressed in these mice contains a lysine in place of glycine at position 119 (22–24) in the bGH protein. This single substitution results in the production of a protein that competes with endogenous GH for GHR [Growth Hormone Receptor] binding..."
However, I'd like to contest their claim that these mice fail to show lifespan (LS) extension. I don't think their data is able to show one way or another.

Consider the problem of sample size and power. GHA mice weigh more than classical dwarf mice, e.g. GHR -/- mice, which is due to their incomplete inhibition of GH-signalling. Female GHA mice weigh up to 33% less than controls (ref. 1), males are even heavier. GHR -/- weigh closer to 60-70% less compared to controls! But this also implies that the effects on lifespan will be diminished in GHA mice, albeit non-zero. I think the IGF-1 data also highlights this striking difference:
"In the GHA mice, [serum IGF-1] levels were reduced to about 75–80% the level of controls (P 0.05). In the GHR [KO] mice, levels were reduced to about 20% the level of controls (P 0.0001)." (2)

Lorenzini et al. recently showed that subtle differences in LS due to modest changes in GH/IGF signalling may require n ~ 100 per group (3). In contrast: Coschigano et al."only" used n~50 per group. Normally, a very respectable sample size, but Lorenzinin and Co needed a larger sample size even though their mice have lower IGF1-levels, i.e. ~50% of controls.
In fact, in the study by Coschigano (2), numerically, the median, mean and maximum lifespan is higher in both genders.

Secondly, males show a smaller numerical improvement in lifespan (2):
Although there was a tendency for the GHA mice to live longer than their controls, especially for the females, this difference did not reach statistical significance for either gender.
First of all, this is consistent with other studies also showing a gender disparity. In addition, given their new data (1) it seems that some idiosyncratic side-effect of GH-antagonism or GH/gentoype interaction might be harming male mice.
"...male GHA mice had significantly lower body weights than WT males until 52 weeks, but thereafter, no statistical difference was observed. In contrast, female GHA mice had significantly lower body weights (ranging from 66% to 81% of female littermate controls) throughout the study...40% and 33% of the total body weight of male and female GHA mice, respectively, was attributed to fat by 68 weeks of age compared with 17% for both control males and females of the same age...  
male GHA mice became significantly more obese with advancing age...The mass of all adipose depots was higher in male GHA mice than their littermate controls at 82 weeks of age; however, there was only a significant increase in the mass of subcutaneous and retroperitoneal fat pads (p = 1.0 × 10−6 and p = .04, respectively) "
So either excess fat is harming them, or perhaps, GH antagonism is not maintained throughout the whole lifespan. However, I am not very satisfied with either explanation, since Berryman et al. also showed beneficial changes in male mice, i.e. increased adiponectin and subcutaneous adiposity with low-normal liver triacylglycerol content. There was also a slight uptick in late-life insulin levels and an increase of the leptin-adiponectin ratio which is more consistent with harmful effects.

Third, as much as I hate to say it, but their study (2) is not good enough, not rigorous enough. Their controls on the black 6 background have mean and median lifespans below 800d, while 900d would be expected. Maximum lifespans also appear to be well below the ~1200d expected from a healthy lab mouse.
Overall this weakens conclusions we can draw based on this study, though, it should not introduce much systematic bias. In fact, low LS is often seen as contributing to exaggerated lifespan effects. In particular, with interventions that offset some harmful aspect of questionable husbandry.

Fourth, GH-toxicity might be a real thing biasing low-IGF1 models, which often show compensatory increases in GH levels (5). I do wonder if the GHA-model (Kopchick lab) and the IGF1 hypomorphic mice also show these compensatory increases?
Increased GH-levels have been shown for the liver IGF1 deficient mice (5) and apparently for "Liver-specific GH receptor gene disrupted (LiGHRKO) mice" as well (6), also by Kopchick.

The solution
A meta-analysis  - not a narrative review, not a systematic review - could in principle overcome the issues I raised above, especially sample size and power. Husbandry effects and study quality could be considered as well.

Is the data too heterogenous for analysis? Perhaps, but I don't think so. Meta-analyses of the effects of dietary restriction have been performed, some even comparing different taxa (c.f. 4.a to d).

So far I'd side with the idea that almost all healthy, well-husbanded models of low GH/IGF1 signalling will show delayed aging if there's no frank toxicity. Let's not forgot that toxicity (extrinsic mortality) shouldn't be confused with intrinsic aging - which is what we'd like to study. It's quite possible that things like CVD-toxicity ultimately limit the efficacy of GH/IGF1-signalling reduction.

References

1. A Dwarf Mouse Model With Decreased GH/IGF-1 Activity That Does Not Experience Life-Span Extension: Potential Impact of Increased Adiposity, Leptin, and Insulin With Advancing Age. Berryman DE, Lubbers ER, Magon V, List EO, Kopchick JJ.J Gerontol A Biol Sci Med Sci. 2014 Feb;69(2):131-41. doi: 10.1093/gerona/glt069. Epub 2013 May 21

2. Coschigano KT, Holland AN, Riders ME, List EO, Flyvbjerg A, Kopchick JJ.
Deletion, but not antagonism, of the mouse growth hormone receptor results in severely decreased body weights, insulin, and insulin-like growth factor I levels and increased life span. Endocrinology. 2003; 144: 3799–3810.

3. J Gerontol A Biol Sci Med Sci. 2013 Jul 20. [Epub ahead of print]Mice Producing Reduced Levels of Insulin-Like Growth Factor Type 1 Display an Increase in Maximum, but not Mean, Life Span. Lorenzini A, Salmon AB, Lerner C, Torres C, Ikeno Y, Motch S, McCarter R, Sell C.

4.a. Aging Cell. 2013 Jun;12(3):410-4. doi: 10.1111/acel.12061. Epub 2013 Mar 27.Dietary restriction of rodents decreases aging rate without affecting initial mortality rate -- a meta-analysis. Simons MJ1, Koch W, Verhulst S.

4.b. Dietary restriction in rats and mice: a meta-analysis and review of the evidence for genotype-dependent effects on lifespan.Swindell WR.Ageing Res Rev. 2012 Apr;11(2):254-70. doi: 10.1016/j.arr.2011.12.006. Epub 2011 Dec 23. Review.

4.c. Mol Biosyst. 2012 Apr;8(4):1339-49. doi: 10.1039/c2mb05255e. Epub 2012 Feb 10.A meta-analysis of caloric restriction gene expression profiles to infer common signatures and regulatory mechanisms.Plank M1, Wuttke D, van Dam S, Clarke SA, de Magalhães JP.

4.d. Int J Cancer. 2003 Sep 20;106(5):766-70.Energy restriction and the risk of spontaneous mammary tumors in mice: a meta-analysis.Dirx MJ1, Zeegers MP, Dagnelie PC, van den Bogaard T, van den Brandt PA.

5. Reductions in serum IGF-1 during aging impair health span.Gong Z, Kennedy O, Sun H, Wu Y, Williams GA, Klein L, Cardoso L, Matheny RW Jr, Hubbard GB, Ikeno Y, Farrar RP, Schaffler MB, Adamo ML, Muzumdar RH, Yakar S.Aging Cell. 2013 Dec 17. doi: 10.1111/acel.12188. [Epub ahead of print]

6. Endocrinology. 2014 Feb 11:en20132086. [Epub ahead of print]Liver-specific GH receptor gene disrupted (LiGHRKO) mice have decreased endocrine IGF-1, increased local IGF-1 as well as altered body size, body composition and adipokine profiles.List EO1, Berryman DE, Funk K, Jara A, Kelder B, Wang F, Stout MB, Zhi X, Sun L, White TA, Lebrasseur NK, Pirtskhalava T, Tchkonia T, Jensen EA, Zhang W, Masternak MM, Kirkland JL, Miller RA, Bartke A, Kopchick JJ.