Zanchi et al Cell Biochem Funct 2010; 28: 232–238.


New Member
Naturally this one caught my attention and you'll see why as you wade through it.

Experimental chronic low-frequency resistance training produces skeletal muscle hypertrophy in the absence of muscle damage and metabolic stress markers

Abstract:Volitional animal resistance training constitutes an important approach to modeling human resistance training. However, the lack of standardization protocol poses a frequent impediment to the production of skeletal muscle hypertrophy and the study of related physiological variables (i.e., cellular damage/inflammation or metabolic stress). Therefore, the purposes of the present study were: (1) to test whether a long term and low frequency experimental resistance training program is capable of producing absolute increases in muscle mass; (2) to examine whether cellular damage/inflammation or metabolic stress is involved in the process of hypertrophy. In order to test this hypothesis, animals
were assigned to a sedentary control (C, n=8) or a resistance trained group (RT, n=7). Trained rats performed 2 exercise sessions per week (16 repetitions per day) during 12 weeks. Our results demonstrated that the resistance training strategy employed was capable of producing absolute mass gain in both soleus and plantaris muscles (12%, p<0.05). Furthermore, muscle tumor necrosis factor (TNF-a) protein expression (soleus muscle) was reduced by 24% (p<0.01) in trained group when compared to sedentary one. Finally, serum creatine kinase (CK) activity and serum lactate concentrations were not affected in either group. Such information may have practical applications if reproduced in situations where skeletal muscle hypertrophy is desired but high mechanical stimuli of skeletal muscle and inflammation are not. Copyright 2010 John Wiley & Sons, Ltd.

Methods Synopsis:
A feeding apparatus was used which trained the rats to stand up to receive food. Rats then on Tuesday and Friday were subjected to receiving food while weighted. Each training session (twice per day) including eight reps. The weights used were 80–90–95% of MVSC (1 RM) for the first 3 reps, then 95% of MVSC for reps 4-7. The 8th repetition was used to access the MVSC. If the rat was successful, the load would be increased 2% until failure. MVSC procedure was performed only in the morning session when the rats were usually unable to perform the 10th repetition. If a lift was not properly performed, the load was decreased 2% in relation to MVSC. The unsuccessful lifts were discarded.

A rest interval of 4 h between sessions was used. In each session, the rats performed approximately 8 repetitions, which took approximately 30 min to be performed. The total number of repetitions performed daily was approximately 16. The rest interval between repetitions was 3 min.

Measurment Synopsis:

Serum creatine kinase (CK). Before the 5th resistance exercise session, blood samples (0.2 ml) from
six rats were obtained from the caudal vein. Additional blood samples were obtained 15 min, 24 h, and 48 h after the afternoon exercise session (6th resistance exercise session). Serum creatine kinase (CK) activity was measured spectrophotometrically at 378C, by using a standard kit (CK-NAC kinetic; Bioclin, Brazil), according to the manufacturer’s instructions.

Plasma lactate. In the 8th resistance exercise session, blood samples (0.2 ml) from four rats were
obtained from the caudal vein. The samples were collected before (pre-exercise) and 1 min after the last repetition. Lactate concentration was measured by using an automated lactate analyzer (YSI 2300, Yellow Springs, OH).

Regenerating fibers. In addition, the general skeletal muscle fiber morphology was assessed 24 h after the last exercise session through hematoxylin staining & eosin counter staining. Approximately 800 fibers from plantaris and soleus were analyzed, as previously described. Regenerating fibers were defined as those with centralized nuclei without other degenerative changes. Fiber counts were performed by two independent observers, and values were averaged.

tumor necrosis factor (TNF-a) immunoassay. 24 h after the last exercise session, the soleus was carefully rinsed in ice cold 0.9% NaCl to remove any blood contaminants. Samples were minced in 1 ml/100mg tissue of ice-cold RIPA buffer (phosphate buffer saline (PBS), pH 7.4, 1% Nonidet P40, 0.1% sodium dodecyl sulfate (SDS), 0.5% sodium deoxycholate, 100 mgml1 phenylmethylsulphonyl fluoride
(PMSF), 30 ml aprotinin, 100mM NaVO4) and homogenized with a Polytron homogenizer. The homogenate was centrifuged at 48C for 20 min at 15 000 xg. Clear supernatants were used. Protein concentration was determined by the method of Bradford. TNF-a was determined in tissue extracts using specific enzyme immunoassay (QuantiKine M, R&D Systems Inc., Minneapolis, MN, USA), following the manufacturer’s instructions.


External load

There was a progressive increase of 34.4% from the first to the last week.

Body and muscle morphologic parameters

Final body weight was not different between RT and C groups. The weight of the soleus and plantaris muscles was significantly greater in the RT than the C group.

Soleus muscle (mg)
Control 109.2 +/-3.06
Trained 123.4 +/-4.17

Plantaris muscle (mg)
Control 228.5 +/-11.06
Trained 259.7+/-4.52

Fiber CSA Changes

Soleus +29%
Plantaris +22.6%

Serum creatine kinase
Not changed at all time points

Centralized nuclei
There was no difference in the presence of central nuclei among C group (0.78 +/-0.13) compared to RT group (0.614 +/-0.43) in the plantaris muscle and the same pattern was observed in the soleus muscle.

C and RT groups exhibited similar low levels of degeneration/inflammation in the skeletal muscle fibers.

TNF-a immunoassay
This parameter was not increased in RT group, when compared to C group, suggesting the absence of
inflammation. Actually, a significantly diminished response of TNF-a in RT group when compared to C
(24.1%; p<0.01) was found.

Plasma lactate
Plasma lactate was slightly, but significantly increased ( p<0.05) after exercise (1.14+/-0.09 mmol L-1) when
compared to pre-exercise values (0.93+/-0.06 mmol L-1).


Many other studies have used 3-5X per week protocols and yet few have used only 2X week. In this study the 2X week (twice daily) at high loads with limited repetitions was capable of inducing absolute growth.

Here is another study (likeKomulainen Int J Sports Med 2000; 21(2): 107-112) in which not only can hypertrophy occur when using concentric only contractions but also when looking at muscle damage itself it can be seen under this protocol that damage was not present, although measured indirectly. Since they did not look at the sarcomere structure itself it is yet to be said if there was any structural changes occurring at that level. But, there was no centralized nuclei and inflammation was also unchanged.

Metabolic Accumulation
Finally, during resistance exercise, metabolic acidosis (via skeletal muscle lactate production) is still widely speculated to be involved in the response of increased plasma growth hormone, local secretion of growth factors such as IGF-1 within the muscle, and skeletal muscle hypertrophy. Although lactate did rise some 22% above baseline this small amount of increase was unlikely to be the cause of any hypertrophy seen. Yet they did not measure any endocrine hormone responses to analyze a relationship.

As can be seen in Denton (Journal of Strength and Conditioning Research, 2006, 20(3), 528–534), lactate increases are a response to mechanical work and even in Denton when intra-set rest was inserted lactate still increased in conjunction with the amount of work performed. Being that high forces used in this study would certainly impact the amount of metabolites needed to perform the work it's evident that some increases in lactate should have been seen.

So whether metabolic accumulation is a cause of hypertrophy or merely an effect of resistance training and muscle contraction is yet to be clearly defined.

So the take home message;

Reps do not have to be performed consecutively to be effective in creating hypertrophic change.
Metabolic acidosis (accumulation) in this study did not seem to be additive to signaling processes involved in hypertrophy.
Damage was not evident in this study.
Training frequency of 2 or 3 times per week may be equally effective in creating a hypertrophic response.