more r igf 1 info

Here is Cy Wilson (T-Mag) responding to a question about IGF-1.

Looks like he is against the use of IGF-1.... BUt you wanted info.... and this is info......

IGF-1: Worst Bodybuilding Drug Ever?

Q: What ever happened to IGF-1? It was talked about in 'roid books in the early '90s but you don't hear much about it now, except for a few sleazy supplement companies who are using the name.

A: IGF-1 can allow for hypertrophy of muscle. Will it do such a thing when administered to humans? Yes. However, the gains seen really aren’t spectacular. More often than not, they don’t even come close to gains seen using androgens.

For the most part, people should realize that IGF-1 is primarily responsible for GH’s anabolic effects in skeletal muscle as well as cell proliferation, leading to enlarged internal organs and increasing the risk for cancer dramatically. Oh, and this most certainly includes Long R3 IGF-I as I know some people will try to argue that it's much safer.

Well, in order to give you the total picture, I’m going to go over some basic molecular biology as well as list the direct evidence we have concerning the side effects of IGF-1 and yes, that includes Long R3 IGF-I.

First, people should understand that in the human cell cycle, growth requires growth factors in general. Seems simple enough. The next thing people need to understand is that for a normal cell, death is something that'll inevitably occur via loss of telomerase or apoptosis (programmed cell death). Again, I can’t overemphasize enough that the default pathway in humans is death, not growth. (Reassuring, isn't it?)

Now, when you hear of cancer, malignant cancer, people tend to think of uncontrolled cell division. Essentially though, these transformed cancerous cells are immortalized. Now, many changes are required for this to occur (i.e. increased telomerase, increased bcl-2, increased myc and decreased p53). In the development of cancer, we tend to think of carcingogens consisting of both initiators and promoters. For instance, some initiators are UV radiation and tobacco smoke, usually causing DNA damage or mutation, whereas promoters tend to stimulate cell division. A few examples are phorbol esters, hormones (e.g. estrogens) and yes, growth factors.

Now, keep in mind both events, initiation and promotion, are required for the development of malignant cells. As a side note, viral infection can also lead to the two events, but I digress. Anyhow, normally a cell serves its purpose and then dies via apoptosis. However, malignant cells don’t undergo apoptosis. They are, as I said before, immortal. The normal triggers to apoptosis are DNA damage, loss of cell-matrix contact, loss of cell to cell contact, and last but most certainly not least, lack of growth factors.

When you introduce growth factors, you’re providing the catalyst for cancer formation, so to speak. Let’s say, for instance, you get many sunburns during your lifetime. Now, let’s say that one cell has its DNA damaged or altered. This, in and of itself, isn’t too much of a concern as this is only one part of the equation, the iniation. The second part is the promoter (including growth factors).

Well, let’s imagine we introduce growth factors to the cell which has damaged or mutated DNA and it then begins to divide at a more and more rapid rate until it won’t stop. Voila, you have a tumor, which is now capable of even faster growth as well as being invasive (able to invade surrounding tissues) and metastatic (able to cause growth in completely unrelated and distant tissues) in regard to other tissues.

In other words, you now have a malignant tumor, which we commonly refer to as cancer. The fact is, cancer stems from just one cell, just one cell, which begins to divide uncontrollably. People often talk about GH and the side effects thereof, but what most don’t realize is that many of those side effects aren't necessarily mediated by growth hormone but by IGF-1.

Many people may go their whole lives with some DNA damage (or mutation rather) and never have cancer, but with the addition of growth factors, you’re asking for trouble. Even more specifically, you can increase the risk of developing rare forms of cancer, like sarcomas, which are tumors commonly found in connective tissues (i.e. muscle, bone, cartilage, etc.)

Okay, now on to the more cosmetic side effects. With Long R3 IGF-I, it was shown to stimulate growth of the gastrointestinal tract. IGF-1 actually had no effect on body weight and wet tissue weight of the small and large intestine, whereas Long R3 IGF-I resulted in a 20% increase in the weight of the small and large intestine. This is what's causing a "GH gut" although using Long R3 IGF-I is much, much worse than using GH.

Something else to keep in mind is that Long R3 IGF-I was shown to be even more potent than IGF-1 in inhibiting apoptosis and thus its potential for causing cancer is many times greater.

Another idea is that IGF-1 may also keep telomerase activity high, which as we noted previously is a contributing factor for the loss of regulation in terms of cell division. In other words, it again can substantially increase the risk for developing cancer. Long R3 IGF-I was shown to increase telomerase activity in human prostate cancer cells, whereas IGF-1 had no effect.

So, when I tell you to stay away from IGF-1, I’m actually referring to Long R3 IGF-I as it’s what's most commonly circulated and used. Although both aren't something a person should use, Long R3 IGF-1 is probably the worst choice you can make.

So, unless you’re an IFBB pro who consistently places in the top ten at popular contests, you should forget about using IGF-1, or specifically the analogue of IGF-1 called Long R3 IGF-I. It’s really not worth the risk. This, out of all the compounds that bodybuilders may use, is probably the worst in terms of potential side effects.

If you want a true distended belly and increased risk of cancer, be my guest. (47-52)


References Cited

1. Tuzcu A, et al. "Insulin sensitivity and hyperprolactinemia." J Endocrinol Invest. 2003 Apr;26(4):341-6

2. Freemark M, et al. "Body weight and fat deposition in prolactin receptor-deficient mice." Endocrinology. 2001 Feb;142(2):532-7

3. Wasada T, Kawahara R, Iwamoto Y. "Lack of evidence for bromocriptine effect on glucose tolerance, insulin resistance, and body fat stores in obese type 2 diabetic patients." Diabetes Care. 2000 Jul;23(7):1039-40

4. Pijl H, et al. "Bromocriptine: a novel approach to the treatment of type 2 diabetes." Diabetes Care. 2000 Aug;23(8):1154-61

5. Ohem N, Holzl J. Some new investigations on Ilex paraguariensis - Flavonoids and triterpenes. Planta Med 1988; 54: 576

6. Bisset NG, ed. Herbal Drugs and Phytopharmaceuticals (Wichtl M, ed., German edition). Stuttgart: Medpharm, 1994

7. Duke JA. Handbook of Medicinal Herbs. Boca Raton: CRC, 1985

8. Martindale: The Extra Pharmacopoeia, 29th edn. (Reynolds JEF, ed.). London: The Pharmaceutical Press, 1989

9. The Merck Index. An Encyclopedia of Chemicals, Drugs and Biologicals, 11th edn. Rahway, NJ: Merck, 1989

10. Carani C, et al. "Role of oestrogen in male sexual behaviour: insights from the natural model of aromatase deficiency." Clin Endocrinol (Oxf). 1999 Oct;51(4):517-24

11. Scordalakes EM, Imwalle DB, Rissman EF. "Oestrogen's masculine side: mediation of mating in male mice." Reproduction. 2002 Sep;124(3):331-8

12. Brady BM, et al. "Demonstration of progesterone receptor-mediated gonadotrophin suppression in the human male." Clin Endocrinol (Oxf) 2003 Apr;58(4):506-12

13. Bauer ER, et al. "Characterisation of the affinity of different anabolics and synthetic hormones to the human androgen receptor, human sex hormone binding globulin and to the bovine progestin receptor." APMIS. 2000 Dec;108(12):838-46

14. Markiewicz L, Gurpide E. "Estrogenic and progestagenic activities of physiologic and synthetic androgens, as measured by in vitro bioassays. Methods Find Exp Clin Pharmacol. 1997 May;19(4):215-22

15. Breithaupt-Grogler K, Niebch G, Schneider E et al: Dose-proportionality of oral thioctic acid - coincidence of assessments via pooled plasma and individual data. Eur J Pharm Sci 1999; 8(1):57-65

16. Hermann R, Niebch G, Borbe HO et al: Enantioselective pharmacokinetics and bioavailability of different racemic alpha-lipoic acid formulations in healthy volunteers. Eur J Pharm Sci 1996; 4:167-174

17. Gleiter CH, Schug BS, Hermann R et al: Influence of food intake on the bioavailability of thioctic acid enantiomers. Eur J Clin Pharmacol 1996; 50(6):513-514

18. Fachinformation: Pleomix-Alpha(R), alpha-Liponsaeure. Illa Health Care GmbH, Geretsried, 1996

19. Streeper RS, Henriksen EJ, Jacob S et al: Differential effects of lipoic acid steroisomers on glucose metabolism in insulin-resistant skeletal muscle. Am J Physiol 1997; 273(1 pt 1):E185-E191

20. Khamaisi M, Potashnik R, Tirosh A et al: Lipoic acid reduces glycemia and increases muscle GLUT4 content in streptozotocin-diabetic rats. Metabolism 1997; 46(7):763-768

21. Henriksen EJ, Jacob S, Streeper RS et al: Stimulated by alpha-lipoic acid of glucose transport activity in skeletal muscle of lean and obese Zucker rats. Life Sci 1997; 61(8):805-812

22. Wickramasinghe SN & Hasan R: In vitro effects of vitamin C, thioctic acid and dihydrolipoic acid on the cytotoxicity of post-ethanol serum. Biochem Pharmacol 1992; 43(3):407-411

23. Dimpfel W, Spueler M, Pierau F-K et al: Thioctic acid induces dose-dependent sprouting of neurites in cultured rat neuroblastoma cells. Dev Pharmacol Ther 1990; 14(3):193-199

24. Prehn JHM, Karkoutly C, Nuglisch J et al: Dihyrolipoate reduces neuronal injury after cerebral ischemia. J Cereb Blood Flow Metab 1992; 12(1):78-87

25. Burkart V, Koike T, Brenner HH et al: Dihydrolipoic acid protects pancreatic islet cells from inflammatory attack. Agents Actions 1993; 38(1-2):60-65

26. Bauer A, Harrer T, Peukert M et al: Alpha-lipoic acid is an effective inhibitor of human immuno-deficiency Virus (HIV-1) replication. Klin Wochenschr 1991; 69(15):722-724

27. Suzuki YJ, Aggarwal BB & Packer L: alpha-Lipoic acid is a potent inhibitor of NF-kappaB activation in human T cells. Biochem Biophys Res Commun 1992; 189(3):1709-1715

28. Bierhaus A, Chevion S, Chevion M et al: Advanced glycation end product-induced activation of NF-kappaB is suppressed by alpha-lipoic acid in cultured endothelial cells. Diabetes 1997; 46(9):1481-1490

29. Constantinescu A, Tritschler H & Packer L: alpha-Lipoic acid protects against hemolysis of human erythrocytes induced by peroxyl radicals. Biochem Mol Biol Int 1994; 33(4):669-679

30. Greene EL, Nelson BA, Robinson KA, Buse MG. "alpha-Lipoic acid prevents the development of glucose-induced insulin resistance in 3T3-L1 adipocytes and accelerates the decline in immunoreactive insulin during cell incubation." Metabolism 2001 Sep;50(9):1063-9

31. Maddux BA, et al. "Protection against oxidative stress-induced insulin resistance in rat L6 muscle cells by mircomolar concentrations of alpha-lipoic acid." Diabetes 2001 Feb;50(2):404-10

32. Weinstein RB, Tritschler HJ, Henriksen EJ. "Antioxidant alpha-lipoic acid and protein turnover in insulin-resistant rat muscle." Free Radic Biol Med 2001 Feb 15;30(4):383-8

33. Yaworsky K, Somwar R, Ramlal T, Tritschler HJ, Klip A. "Engagement of the insulin-sensitive pathway in the stimulation of glucose transport by alpha-lipoic acid in 3T3-L1 adipocytes." Diabetologia 2000 Mar;43(3):294-303

34. Jacob S, et al. "Oral administration of RAC-alpha-lipoic acid modulates insulin sensitivity in patients with type-2 diabetes mellitus: a placebo-controlled pilot trial." Free Radic Biol Med 1999 Aug;27(3-4):309-14

35. Estrada DE, et al. "Stimulation of glucose uptake by the natural coenzyme alpha-lipoic acid/thioctic acid: participation of elements of the insulin signaling pathway." Diabetes 1996 Dec;45(12):1798-804

36. Hundal RS, et al. "Mechanism by which metformin reduces glucose production in type 2 diabetes." Diabetes 2000 Dec;49(12):2063-9

37. Anderwald C, et al. "Inhibition of glucose production and stimulation of bile flow by R (+)-alpha-lipoic acid enantiomer in rat liver." Liver 2002 Aug;22(4):355-62

38. Saengsirisuwan V, Perez FR, Kinnick TR, Henriksen EJ. "Effects of exercise training and antioxidant R-ALA on glucose transport in insulin-sensitive rat skeletal muscle." J Appl Physiol 2002 Jan;92(1):50-8

39. Evans JL, Goldfine ID. "Alpha-lipoic acid: a multifunctional antioxidant that improves insulin sensitivity in patients with type 2 diabetes." Diabetes Technol Ther 2000 Autumn;2(3):401-13

40. Saengsirisuwan V, Kinnick TR, Schmit MB, Henriksen EJ. "Interactions of exercise training and lipoic acid on skeletal muscle glucose transport in obese Zucker rats." J Appl Physiol 2001 Jul;91(1):145-53

41. Konrad T. "alpha-Lipoic acid treatment decreases serum lactate and pyruvate concentrations and improves glucose effectiveness in lean and obese patients with type 2 diabetes." Diabetes Care 1999 Feb;22(2):280-7

42. Jacob S, et al. "Enhancement of glucose disposal in patients with type 2 diabetes by alpha-lipoic acid." Arzneimittelforschung 1995 Aug;45(8):872-4

43. Jacob S, Rett K, Henriksen EJ, Haring HU. "Thioctic acid—effects on insulin sensitivity and glucose-metabolism." Biofactors 1999;10(2-3):169-74

44. Jacob S, Henriksen EJ, Tritschler HJ, Augustin HJ, Dietze GJ. "Improvement of insulin-stimulated glucose-disposal in type 2 diabetes after repeated parenteral administration of thioctic acid." Exp Clin Endocrinol Diabetes 1996;104(3):284-8

45. Bigsby RM, Caperell-Grant A, Madhukar BV. "Xenobiotics released from fat during fasting produce estrogenic effects in ovariectomized mice." Cancer Res. 1997 Mar 1;57(5):865-9

46. McCurdy CE, Davidson RT, Cartee GD. "Brief calorie restriction increases Akt2 phosphorylation in insulin-stimulated rat skeletal muscle." Am J Physiol Endocrinol Metab. 2003 Oct;285(4):E693-E700.

47. Steeb CB, Trahair JF, Read LC. "Administration of insulin-like growth factor-I (IGF-I) peptides for three days stimulates proliferation of the small intestinal epithelium in rats." Gut. 1995 Nov;37(5):630-8

48. Wetterau LA, Francis MJ, Ma L, Cohen P. "Insulin-like growth factor I stimulates telomerase activity in prostate cancer cells." J Clin Endocrinol Metab. 2003 Jul;88(7):3354-9

49. Devi GR, Graham DL, Oh Y, Rosenfeld RG. "Effect of IGFBP-3 on IGF- and IGF-analogue-induced insulin-like growth factor-I receptor (IGFIR) signalling." Growth Horm IGF Res. 2001 Aug;11(4):231-9

50. Nickerson T, Huynh H, Pollak M. "Insulin-like growth factor binding protein-3 induces apoptosis in MCF7 breast cancer cells." Biochem Biophys Res Commun. 1997 Aug 28;237(3):690-3

51. Vink-van Wijngaarden T, Pols HA, Buurman CJ, Birkenhager JC, van Leeuwen JP. "Inhibition of insulin- and insulin-like growth factor-I-stimulated growth of human breast cancer cells by 1,25-dihydroxyvitamin D3 and the vitamin D3 analogue EB1089." Eur J Cancer. 1996 May;32A(5):842-8

52. Wetterau LA, Francis MJ, Ma L, Cohen P. "Insulin-like growth factor I stimulates telomerase activity in prostate cancer cells." J Clin Endocrinol Metab. 2003 Jul;88(7):3354-9

Here's what Dave Palumbo had to say about igf1 in Muscular Development May 2002. " Oh, it works. I would be so bold as to say 10 mcg of IGF1 would be the equivalent of taking almost 50 ius of GH(wow!). It's a very potent hormone. People take too much, it's not a miracle drug-nothing is. IGF greatly stimulates the metabolic rate, which means you must eat a lot more. Plus, people may be taking fake IGF1.
So anyone try taking only 10mcgs/day? Considering it's only $250 for 1000mcgs, this dose would last a good 6 months @30day on/30 off...

More info from another user from another site:

R3(long) IGF-1

1: Type- IGF-1 Long R3 (Anything else is not as effective, and if the person providing it for you doesn't know anything about it, you are asking for trouble.)
2. Storage- the most popular (and most effective) way to store, transport, preserve IGF is by suspending it in sterile BA in a sterile vial.
This will keep your IGF 99% potent for many months at a time in just about ANY indoor storage, I.E.-closet, drawer, etc. (Take it from me, I stored mine because I wasn't ready to use it for about 6 months in my closet... I had fears about its potency, then I started my first week, and BAM I practically cleaned out the fridge.
3. Use- Usage should not exceed 4-5 weeks, and an OFF period should be about the same. Daily dosages work best (split up into 2 seems to make little difference in the Long R3 version) Most people see results at about 40mcg/day, some use as low as 30mcg/day, and some folks even use 80-100mcg. I SUGGEST to ALL first time users no matter what level, to start at about 40-50mcg/day.
4. Administration- I believe in IM injections over sub q, but either seems to be effective. I like IM better because IM using a slin pin is probably the least painful thing one could imagine, even at two times per day. Also, sub Q shots that contain BA, even diluted BA, can leave little nodules that you may not want to feel on your stomach.
5. Mixing- Most IGF comes suspended in BA. Hopefully it is @ 500mcg/ml or even 333mcg/ml (that would be at 2ml/mg and 3ml/mg respectively) Draw out your desired amount and back load a slin pin. Add enough Bacteriostatic Water to fill the U100 syringe completely.
Some inject immediately before training, while others choose to do 2 shots spread throughout the day... THEY BOTH WORK WELL. Try both; see which method makes your muscles pop out of your skin.
6. Add plenty of protein, and don’t shy away from carbs immediately after training. I used up to 100g of carbs after training, and my body fat went down, all without cardio.

Peace,
P

Another user:

One more thought on IGF-1 Long-R. It cutting properties are truly amazing but I think it would be a waste of ammo to use it in a cutter. I think adding it to the last weeks of a bulking cycle is the way to best utilize it. In another post I've read, someone said it helps to solidify gains. Well if you are like me, the lbs at the end of a bulking cycle are the hardest to come by. Also the end is where I put on the most fat.

I'm coming to the end of a bulk cycle this Sat. I started IGF in week 5 (of an 8 week cycle). I originally posted I added 750 calories a day. Well going back to check my training log I was a little off. I added 600 calories in week 5 (as soon as I started to lose weight) and another 300 in week 6. I weighed in at the beginning of week 8 and I dropped 3 more lbs. I'll weigh in again Sat for the final tally. At the end of week 4 I had put 1 1/4 inches on my waist. As of the beginning of week 8 the number dropped by 3/4 of an inch, down to a 1/2 inch waist gain. Strength has increased each week of this cycle. Now before you read too much into all these figures I was off for 2 1/2 months with the achilles injury. So alot of this can be attributed to rebound. I did set new PB's on delts and are very close on other bodyparts. I'm 44 yrs old too, so that plays into recovery abilities (not for the good).

I do have a 25 yr old friend who uses it as a bridge between cycles. This guy has a hard time keeping gains. He does follow all the post-cycle regime to a "T". It has allowed him to keep more post cycle than he has in the past.

Here is some info from animal's board

Here's a link to the pdf,complete with graphs and pics.It's about 4.5 megs,so just a heads up.
http://seroudelab.biology.queensu.ca/pdf/musaro.pdf

also the text,

Localized Igf-1 transgene expression sustains hypertrophy and regeneration in senescent skeletal muscle

Antonio Musarò1, 2, Karl McCullagh1, Angelika
Paul1, Leslie Houghton1, Gabriella Dobrowolny2, Mario Molinaro2, Elisabeth R. Barton3, H. L Sweeney3 & Nadia Rosenthal1

1. Cardiovascular Research Center, Massachusetts General
Hospital-East, Charlestown, Massachusetts, USA.
2. Department of Histology and Medical Embryology, University of Rome "La Sapienza", Rome, Italy.
3. Department of Physiology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA.
Correspondence should be addressed to N Rosenthal. e-mail: [email protected]

Aging skeletal muscles suffer a steady decline in mass and
functional performance, and compromised muscle integrity as fibrotic invasions replace contractile tissue, accompanied by a characteristic loss in the fastest, most powerful muscle fibers1, 2. The same programmed deficits in muscle structure and function are found in numerous neurodegenerative syndromes and disease-related cachexia3. We have generated a model of persistent, functional myocyte hypertrophy using a tissue- restricted transgene encoding a locally acting isoform of insulin-like growth factor-1 that is expressed in skeletal muscle (mIgf-1). Transgenic embryos developed normally, and postnatal increases in muscle mass and strength were not accompanied by the additional pathological changes seen in other Igf-1 transgenic models. Expression of GATA-2, a transcription factor normally undetected in skeletal muscle, marked hypertrophic myocytes that escaped age-related muscle atrophy and retained the proliferative response to muscle injury characteristic of younger animals. The preservation of muscle architecture and age-independent regenerative capacity through localized mIgf-1 transgene expression suggests clinical strategies for the treatment of age or disease-related muscle frailty.

Experimental models of muscle growth and regeneration have
implicated Igf-1 as an important mediator of anabolic pathways in skeletal muscle cells4, 5. Although liver is the principle source of circulating Igf-1, targeted ablation of Igf1 demonstrated that liver-derived Igf-1 is not required for postnatal body growth6. Administration of Igf-1 protein to cultured muscle cells elicits a biphasic response, first stimulating cell proliferation and subsequently enhancing myogenic differentiation7, 8, a sequence of events ideally timed for the repair of damaged tissue.

The fact that Igf-1 can act either as a hormone or as a local growth factor has complicated analysis of animal models in which transgenic Igf synthesized in extrahepatic tissues was released into the circulation9-12. The Igf1 locus encodes multiple isoforms generated by differential promoter usage and alternative transcript splicing, producing variable amino-terminal signal peptides and multiple carboxy-terminal E peptides (Fig. 1a). Restricting the action of supplemental Igf-1 to the tissue of origin by choice of the appropriate isoform allowed us to assess its autocrine/paracrine role in skeletal muscle throughout the lifespan of the animal, exclusive of possible endocrine effects on other tissues.

The Igf-1 isoform used here (referred to as mIgf-1) is normally synthesized in skeletal and cardiac muscle13. We generated transgenic mice with a rat mIgf-1 cDNA driven by skeletal muscle-specific regulatory elements from the rat myosin light chain (MLC)-1/3 locus. Two MLC/mIgf-1 transgenic lines were selected for characterization due to high transgene expression levels. Expression of the MLC/mIgf-1 transgene in adult mice was restricted to skeletal muscle and predominated in muscles enriched in fast fibers, such as the triceps or gastrocnemius. Transgene expression was reduced in slow muscles such as the soleus, where the MLC regulatory cassette is characteristically expressed at very low levels14 (Fig. 1b).

Embryonic development of MLC/mIgf-1 transgenic animals proceeded normally. Endogenous MLC1/3, myosin heavy chain (MyHC), muscle creatine kinase (MCK) and Igf binding protein- 5 (IgfBP5) expression were unaffected during development up to six weeks (data not shown), indicating that expression of the mIgf-1 transgene does not alter steady state levels of these genes as it does in cultured myocytes15. Skeletal muscle hypertrophy in MLC/mIgf-1 transgenic neonates was first detectable at 10 days (Fig. 2a), increasing into adulthood, when the transgenic mice showed hypertrophy in trunk and limb musculature with little or no body fat, compared with their age- matched wild-type sibs (Fig. 2b). No instances of cardiac hypertrophy were detected in the transgenic lines (data not shown), suggesting that persistent expression of mIgf-1 in skeletal muscle does not lead to systemic perturbations in these animals.

The fast-fiber-specific effects of the MLC/mIgf-1 transgene
were reflected in the average wet muscle weights of transgenic and wild-type adults, where muscles rich in fast fibers such as thigh triceps or gastrocnemious had the most increase (Table 1). That these changes were due to fiber hypertrophy in the MLC/mIgf-1 muscles was documented by the overall increase in the cross-sectional area of fibers in thigh as opposed to soleus, and in the increased diameter specifically of fast fibers in a mixed-bed muscle (Fig. 3a,b). The relative distribution of slow type I and fast type IIa fibers were unaltered by MLC/mIgf-1 expression, whereas the cross-sectional diameter of type IIb fibers was increased in MLC/mIgf-1 transgenic muscles compared with that of their wild-type counterparts (Fig. 3c).

Increased muscle mass in MLC/mIgf-1 transgenic EDL muscle was associated with increased force generation compared with age-matched wild-type littermates (Fig. 3d). A quantitative analysis of fiber composition in EDL muscles of wild-type and MLC/mIgf-1 transgenic adult (6 months) mice revealed only a modest shift in favor of the fastest IIb fibers (Fig. 3e).

Previous characterization of Igf-1 action in skeletal muscle cell cultures16, 17 implicated calcineurin signaling, which also underlies cardiomyocyte hypertrophy18. To test the involvement of calcineurin in MLC/mIgf-1 transgenic animals, we administered cyclosporin, a potent inhibitor of calcineurin activity and myocyte hypertrophy in cardiac and skeletal muscle16-19, by means of a subcutaneous pump. After three weeks, no reduction of fast fiber hypertrophy was evident; rather, slow muscle fibers were preferentially hypertrophied, perhaps through ectopic activation of the fast-fiber-specific MLC/Igf transgene (Fig. 4a). Induction of slow fiber hypertrophy in the MLC/mIgf-1 background is not unreasonable in the light of studies demonstrating a critical role for calcineurin determining in the slow fiber phenotype20-22, in which cyclosporin inhibited calcineurin activity in slow fibers and converted them to a fast phenotype. A slow-to-fast fiber conversion mediated by cyclosporin may activate ectopic MLC/ mIgf-1 transgene expression. Although cyclosporin can suppress differentiation and Igf-1-mediated hypertrophy in cell culture by a blockade in calcineurin signalling16, 17, 23, it may be less effective in vivo because other Igf-1-responsive pathways, such as those mediated by phosphoinositol (PI3) kinase15, or calmodulin (CaM) kinase24 may compensate.

Forced expression of either mIgf-1 or activated calcineurin in
muscle cultures also induces expression of GATA-2, a transcription factor that accumulates exclusively in hypertrophic myotube nuclei complexed with calcineurin and NFATc1, a target of calcineurin phosphatase activity16. Both GATA-2 transcripts and protein were activated in MLC/mIgf-1 transgenic muscles, but were undetectable in the muscles of wild-type sibs (Fig. 4b,c), establishing GATA-2 as a molecular marker of skeletal muscle hypertrophy in vivo.

Examination of older mice revealed that expression of the mIgf-1 transgene was protective against normal loss of muscle mass during senescence. Wet weights of MLC/mIgf-1 transgenic muscles remained unchanged as late as 20 months, whereas their wild-type littermates underwent muscle atrophy (Table 1 and Fig. 5a). Older transgenic mice occasionally contained centralized nuclei (Fig. 5b), which were already absent from wild-type sibs after 7-12 months of age (data not shown). Fibers with centralized nuclei induced expression of the MyHC neonatal isoform (Fig. 5c), another hallmark of regenerating myofibers. Thus, mIgf-1 may act as a survival factor by prolonging the regenerative potential of younger muscle.

We therefore analyzed the proliferative response of senescent transgenic muscle tissue to injury. Induction of an nls-lacZ transgene driven by the desmin promoter, normally silent in senescent muscle, was used as a marker of satellite cell mobilization, as desmin is activated early during the regeneration process25. Single intramuscular injections of cardiotoxin (CDX) in desmin/nls-lacZ mice at 22 months failed to activate the transgene at any time after injury (Fig. 6c, and data not shown). In contrast, injections of bigenic desmin/nls- lacZMLC/mIgf-1 mice at 22 months induced LacZ expression by day 2 post-injury, in both undamaged and damaged areas (Fig. 6d,e). Activation of proliferating cell nuclear antigen (PCNA; Fig. 6e, inset), MyoD and endogenous desmin (data not shown) marked proliferating cells. After six days, LacZ- positive nuclei were present in newly formed myofibers, concomitant with the activation of neonatal MyHC (Fig. 6f,g). Muscle fibers at day 12 post-injury were characterized by the peripheralization of Lac-Z positive nuclei (Fig. 6h), indicating the endpoint of the regeneration program. Thus Igf-1 promotes muscle regeneration in senescent muscle, leading to muscle reconstitution characteristic of young animals.

Our study demonstrates that the integrity and regenerative capacity of total body skeletal musculature is safely extended by local mIgf-1 supplementation into senescence. Even when expressed under the control of strong muscle regulatory elements, the mIgf-1 protein appears to remain in the muscle bed and does not enter the circulation, thereby avoiding hypertrophic effects on distal organs such as the heart, and eliminating risk of possible neoplasms induced by inappropriately high levels of circulating Igf-1. This is in contrast to previous reports in which another Igf-1 isoform expressed pan-fiber specifically in skeletal muscle and heart was detected in the circulation at all ages, causing first physiologic then pathologic cardiac hypertrophy10, 12.

Prolongation of functional regeneration in senescent MLC/ mIgf-1 muscle also reflects increased satellite cell proliferative capacity. We previously found that local delivery of mIgf-1 to individual mouse muscles by AAV virus-mediated gene transfer blocks age-related loss of muscle size and strength26, which is at least partially attributable to satellite cell activity27. As cell- based therapies aimed at prolongation of skeletal muscle strength in aging and neuromuscular disease are limited by the small number of muscle stem cells isolated from normal muscle, viable therapeutic strategies based on autologous material will depend on the development of methods to increase the muscle stem cell population. Expanding mature stem cell populations by administration of mIgf-1 genes may provide a clinically relevant avenue for the prevention, attenuation or reversal of age and disease-related muscle frailty.

Methods

Generation of MLC/mIgf-1 transgenic mice. The MLC/mIgf-1
expression vector has been described15. Briefly, we subcloned the mIgf-1 cDNA into the pMex plasmid containing the 1,500-bp fragment of the MLC promoter, an 840-bp fragment of SV40 poly(A), and a 900-bp fragment from the 3' end of the MLC1f/3f gene, which acts as an enhancer14. FVB mice (Jackson Laboratories) were used as embryo donors. The transgene insert was excised and purified to avoid introduction of vector sequences into the transgenes. We generated transgenic mice by standard methods. Positive founders were subsequently bred to FVB wild-type mice and MLC/mIgf-1 transgenic mice were selected by PCR using tail digests. The animals were housed in a temperature-controlled (22 °C) room with a 12:12 h light-dark cycle.

Histological and immunofluorescence analysis. Mice were anesthetized before cervical dislocation and muscle tissue was separated from bone and most connective tissue and immediately frozen in liquid nitrogen. Segments of quadriceps muscle from wild-type and MLC/mIgf-1 transgenic mice were embedded in TBS-tissue freezing medium and snap frozen in nitrogen-cooled isopentane. Frozen sections (7 m) were stained with hematoxylin and eosin and analyzed morphologically. For immunofluorescence analysis, frozen sections (7 m) were fixed in 4% paraformaldehyde for 10 min on ice, washed with PBS and then pre-incubated in PBS containing 1% BSA, 1:100 goat serum for 1 h at RT, and processed as described15. We used the following antibodies: monoclonal antibodies specific for myosin heavy chain (MyHC) I, IIa and IIb, MyHC-embryonic and GATA-2. Muscle fiber nuclei were visualized using Hoechst staining.

RNA preparation and northern-blot analysis. We obtained total RNA from wild-type and MLC/mIgf-1 transgenic muscles by RNA-TRIZOL extraction (Gibco-BRL). Total RNA (15 g) was fractionated by electrophoresis on 1.3% agarose gels and hybridized as described15.

Cyclosporin treatment. Animals were anesthetized with an intraperitoneal injection of avertin. Osmotic pumps (Alzet; model 1002) containing cyclosporin (CsA; Sandoz, 50 mg/ml) or PBS were implanted subcutaneously on 3-week MLC/mIgf-1 transgenic mice and wild-type littermates. The pumps delivered CsA at a dose of 18 mg/kg body weights/day or PBS alone for 14 d. Animals were monitored and remained in good health, as determined by the 50% increase in body weights at the end of treatment. Mice were killed at two weeks, blood samples were taken by cardiac puncture and muscles were frozen in preparation for sectioning and staining as discussed above. Blood samples were analyzed by the Massachusetts General Hospital biochemistry laboratory and all mice had cyclosporin blood levels >1,000 ng/ml.

Mechanical measurements. Mice, age 6-8 months, were killed by cervical dislocation under anesthesia, and EDL muscles were removed for isolated muscle force measurements as described26. The tendons were attached to a rigid post and to an isometric force transducer in a bath of Ringers solution gas-equilibrated with 95% O2/5% CO2. Optimum length of the muscle was determined by twitch force from supramaximal stimulation. Tetanic force was measured at 20, 50, 80, 100 and 120 Hz using a 500-ms pulse delivered through two parallel platinum plate electrodes. Maximal tetanic force was determined at the 120 Hz stimulation frequency. After force measurements were completed the muscle were removed from the bath, blotted and weighed. They were then pinned at optimum length, surrounded by embedding compound (TissueTek) and rapidly frozen in melting isopentane. Muscles were stored at -80 °C for subsequent analysis.

Myosin heavy chain composition. Immunohistochemistry was used to determine myosin heavy chain composition. Primary antibody dilutions were as follows: type I myosin (BA- F8), 1:50; type IIa (SC-71), 1:10; type IIb (BF-F3), 1:3. FTC- conjugated, goat anti-mouse IgM antibodies and donkey anti- mouse IgG (H+L) (Jackson) were used as secondary antibodies. Microscopy was performed on a Leitz DMR microscope (Leica). Image acquisition and analysis was carried out using a Micro MAX digital camera system (Princeton Instruments) and imaging software (IPLAB, Signal Analytics).

Muscle regeneration. Tibialis anterior (TA) muscle from 22- month Desmin/nls-LacZ (ref. 25) and MLC/Igf-1Desmin/nls- LacZ mice were injected with 30 l of 10 M cardiotoxin, diluted in PBS. The TA muscles were collected 2, 6 and 12 d following the injection of cardiotoxin. Frozen sections (7 m) of TA muscles from both Desmin/nls-LacZ and MLC/Igf-1Desmin/ LacZ mice were fixed in 4% paraformaldehyde 10 min on ice and then stained with X-gal. For immunohistochemistry, serial sections were fixed with 4% paraformaldehyde, incubated with 10% normal goat serum for 1 h at RT to reduce background staining, and incubated overnight at 4 °C with either mouse monoclonal PCNA (Santa Cruz), or mouse monoclonal MyHC- neonatal primary antibodies. The section were washed with PBS, incubated with secondary antibodies for 1 h at RT and mounted under coverslips.

Received 16 August 2000; Accepted 30 November 2000.

REFERENCES

Brooks, S. & Faulkner, J. Contractile properties of skeletal
muscles from young, adult and aged mice. J. Physiol. 404, 71-82 (1988). | PubMed | ISI |
Musaro, A. et al. Enhanced expression of myogenic regulatory factors in aging skeletal muscle. Exp. Cell Res. 221, 241-248 (1995). | Article | PubMed | ISI |
Nelson, K. The cancer anorexia-cachexia syndrome. Semin. Oncol. 27, 64-68 (2000). | PubMed | ISI |
Florini, J., Ewton, D. & Coolican, S. Growth hormone and the insulin-like growth factor system in myogenesis. Endocrine Rev. 17, 481-517 (1996). | ISI |
Stewart, C. & Rotwein, P. Growth, differentation, and survival: multiple physiological functions for insulin-like growth factors. Physiol. Rev. 76, 1005-1026 (1996). | PubMed | ISI |
Sjogren, K. et al. Liver-derived IGF-1 is the principal source of IGF-1 in blood but is not required for postnatal body growth in mice. Proc. Natl. Acad. Sci. USA 96, 7088-7092 (1999). | Article | PubMed | ISI |
Rosenthal, S. & Cheng, Z.Q. Opposing early and late effects of insulin-like growth factor I on differentiation and the cell cycle regulatory retinoblastoma protein in skeletal myoblasts. Proc. Natl. Acad. Sci. USA 92, 10307-10311 (1995). | PubMed | ISI |
Engert, J., Berglund, E.B. & Rosenthal, N. Proliferation precedes differentiation in IGF-1 stimulated myogenesis. J. Cell Biol. 135, 431-440 (1996). | PubMed | ISI |
Mathews, L. et al. Growth enhancement of transgenic mice expressing human insulin-like growth factor-I. Endocrinology 123, 2827-2833 (1988). | PubMed | ISI |
Coleman, M. et al. Myogenic vector expression of insulin- like growth factor I stimulate myocyte differentiation and myofiber hypertrophy in transgenic mice. J. Biol. Chem. 270, 12109-12116 (1995). | Article | PubMed | ISI |
Reiss, K. et al. Overexpression of insulin-like growth factor-I in the heart is coupled with myocyte proliferation in transgenic mice. Proc. Natl. Acad. Sci. USA 93, 8630-8635 (1996). | Article | PubMed | ISI |
Delaughter, M.C., Taffet, G.E., Fiorotto, M.L., Entman, M.L. & Schwartz, R.J. Local insulin-like growth factor I expression induces physiologic, then pathologic cardiac hypertrophy in transgenic mice. FASEB J. 13, 1923-1929 (1999). | PubMed | ISI |
Adamo, M. et al. Structure, expression and regulation of the IGF-1 gene. Adv. Exp. Med. Biol. 343, 1-11 (1993). | PubMed |
Grieshammer, U., Sassoon, D. & Rosenthal, N. A transgene target for positional regulators marks early rostrocaudal specification of myogenic lineages. Cell 69, 79-93 (1992). | PubMed | ISI |
Musaro, A. & Rosenthal, N. Maturation of the myogenic program is induced by post-mitotic expression of IGF-1. Mol. Cell. Biol. 19, 3115-3124 (1999). | PubMed | ISI |
Musaro, A., McKullagh, K.J.A., Naya, F.J., Olson, E.N. & Rosenthal, N. IGF-1 induces skeletal myocyte hypertrophy through calcineurin in assocation with GATA- 2 and NF-ATc1. Nature 400, 581-585 (1999). | Article | PubMed | ISI |
Semsarian, C. et al. Skeletal muscle hypertrophy is mediated by a Ca2+-dependent calcineurin signalling pathway. Nature 400, 576-581 (1999). | Article | PubMed | ISI |
Molkentin, J. et al. A calcineurin-dependent transcriptional pathway for cardiac hypertrophy. Cell 93, 215-228 (1998). | PubMed | ISI |
Lim, H. et al. Reversal of cardiac hypertrophy in transgenic disease models by calcineurin inhibition. J. Mol. Cell. Cardiol. 32, 697-709 (2000). | Article | PubMed | ISI |
Chin, E. et al. A calcineurin-dependent transcriptional pathway controls skeletal muscle fiber type. Genes Dev. 12, 2499-2509 (1998). | PubMed | ISI |
Naya, F. et al. Stimulation of slow skeletal muscle fiber gene expression by calcineurin in vivo. J. Biol. Chem. 275, 4545-4548 (2000). | Article | PubMed | ISI |
Bigard, X. et al. Calcineurin co-regulates contractile and metabolic components of slow muscle phenotype. J. Biol. Chem. 275, 19653-19660 (2000). | Article | PubMed | ISI |
Abbott, K.L., Friday, B.B., Thaloor, D., Murphy, T.J. & Pavlath, G.K. Activation and cellular localization of the cyclosporine A-sensitive transcription factor NF-AT in skeletal muscle cells. Mol. Biol. Cell 9, 2905-2916 (1998). | PubMed | ISI |
Passier, R. et al. CaM kinase singaling induces cardiac hypertrophy and activates the MEF2 transcription factor in vivo. J. Clin. Invest. 105, 1395-1406 (2000). | PubMed | ISI |
Lescaudron, L., Creuzet, S.E., Li, Z., Paulin, D. & Fontaine- Perus, J. Desmin-lacZ transgene expression and regeneration within skeletal muscle transplants. J. Muscle Res. Cell. Motil. 18, 631-641 (1997). | Article | PubMed | ISI |
Barton-Davis, E., Shoturma, D.I., Musaro, A., Rosenthal, N. & Sweeney, H.L. Viral mediated expression of IGF-1 blocks the aging-related loss of skeletal muscle function. Proc. Natl. Acad. Sci. USA 95, 15603-15607 (1998). | Article | PubMed | ISI |
Barton-Davis, E.R., Shoturma, D.I. & Sweeney, H.L. Contribution of satellite cells to IGF-1 induced hypertrophy of skeletal muscle. Acta Physiol. Scand. 167, 301-305 (1999). | Article | PubMed | ISI |
LeRoith, D. & Roberts, C.T., Jr. At the cutting edge. Insulin- like growth factor I (IGF-1): a molecular basis for endocrine versus local action. Mol. Cell. Endocrinol. 77, C57-C61 (1991). | PubMed | ISI |
McKoy, G. et al. Expression of insulin growth factor-I splice variants and structural genes in rabbit skeletal muscle induced by stretch and stimulation. J. Physiol. 516, 583-592 (1999). | PubMed | ISI |

correct i understand all this, this has been gone over before, but the qestion is can you obtain migf-1 from anywhere rosenthal group? or is the shit not availible yet?

One more user's experience:

Originally posted by napjy44
Ok here goes on the info...
Dosage... 40mcg eod (basically every training day)
Other gear.... None (for the first 1000mcg)
Ht-5'9"
Wt-202lbs
BF% 13.4-14 (Calipers)
Start date (4/5/03)
Training Style- Usually 2on2off (1/2 body trained each workout)
Dorian style sets, lots of Rest Pause though and negatives.
Diet- I eat about 6 times per day.
M1-Egg Whites, 1 piece of cheese, and 1 whole wheat english muffin.
M2- Usually protein leftovers, I.E.-chicken, pork, or 10oz of 1% cottage cheese
M3- Lunch- 12-16ozs grilled chicken, some rice or potatoes
M4-similar to meal 2
Snack- 30g protein, with some pre workout boosters, various Aminos etc.
Postworkout Shake- Approx 50-60g Protein and 60-80g Dextrose
M5-Protein- Meat, chicken, pork, etc. With small serving of starch source (dinner) some greens like organic salad.
M6-similar to meals 2 & 4
Before bed- 25g protein, in milk, with GABA or Taurine.

Supplements- I take in lots of EFA's and Olive oil. ALA included. 3mg a day of yohimbe, or topical yo for love handles.

More to come guys... I'm a pretty busy guy, not important, just busy.

So far though (day 3) my pumps are better, I AM very hungry for quite some time after injection, and I think I'll be adding more total protein..

Here are the finals:
Super Supps- NONE
Finish date- 5/7/03
Bdy wT- 209 (+7)
BF%-11.6 (calipers) (-1.8%)
Training style-2on -2off, Dorian style or Rest Pause
Some numbers from trainingall #'s w/out a date are from before cycle)
ALl reps are done with an 8 count negative, an explosive positive, and only fifteen breaths between RP sets.
Incline DB Press- 100lbs Rest Pause- 6,5,4
5/05/03 Inc DB Press- 115lbs RP 6,5,4
Neutral Pull up- BdyWt RP- 6,4,3
5/5/03 Neutral Pull-up- BdyWt- 7,6,5
Hammer Shoulder- 200lbs- 7,4,4
5/5/03 240lbs- 6,4,4
Cable Row- 200lbs 7,5,5
5/5/03 230lbs 6,5,5
Leg Ext 190lbs 7,5,4
5/7/03 220 10,9,8
Leg curl 90 7,5,4
5/7/03 120 9,8,7

*some numbers sre from a week or two before the start of my program b/c of the splits I use... I have 6 separate workouts, 3of one split, and 3 of another, hitting all six in a matter of 14 days. Over the five weeks+ time I have only done the rotation 3.5 times.

Lots more, too tedious and boring for you folks to read...

1. When I started taking 40mcg every day my gains took off.
2. Upping protein above 325g day made a bid difference in recovery.
3. Received numerous compliments on hardness and more definition/size.
4. Definitely setting up the next experiment with other items.
5. More protein than ever next time.
6. Any q's post away...

koz215 posted the other info that I was going to put up but if I find any more I will put it up. Hope this helps everyone.

Westsider

damn my eyes are tired from reading all this info now, good night guys..LOL