Subjects:

          Ten female student girls from the faculty of physical
education (Grade 2) constituted subjects of this study. Written informed
consent was obtained from all participants according to the ethical approval
for this study conformed to the standards of the Declaration of Helsinki.  The Helwan University review board approved
the protocol before study initiation. Height, weight and age were recorded for
all investigated subjects. All participants were required to abstain from
non-steroidal anti-inflammatory use for at least 7 days prior the experimental
study day.

 

Training protocol:

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The subjects trained
for 12 weeks as 3 days/week in a basketball program. Each training session
consisted of 10 min. warm up followed by 60 – 80 min. of specific basketball
skills training program under specialized coach supervision, then positive cool
down for another 10 min. Exercise on a cycle ergometer to exhaustion was
performed on the sampling days.

 

Plasma sampling:

          Five milliliters of blood was collected under aseptic
conditions in standard anticoagulant (EDTA)-treated vacutainer tubes at
baseline (prior to exercise testing) and immediately post-exercise testing. All
blood samples were centrifuged at 2000 g for 10 min to pellet cellular elements
immediately after each blood draw. The supernatant plasma was then separated
and immediately frozen at –80?C till PCR assay.

 

miRNA extraction:

          Total
RNA extraction was performed using a MicroRNA Extraction Kit (Shanghai
ShineGene Molecular Biotech, Inc., Shanghai, China). Quantitative efficacy and
reproducibility of c-miRNA extraction was confirmed by extraction via serial
dilutions.

 

Quantification of
microRNA expression:

To quantify levels of
select c-miRNA, standard reverse transcription-quantitative (‘real time’)
polymerase chain reaction (RT-QPCR) was utilized. Specifically, reverse
transcription was performed to generate cDNA representing levels of mature
c-miRNA molecules.

 

Fold-change of miRNA 206
species was calculated using the formula specified by the manufacturing
company. Specifically, in this formula, Ct represents the ‘real time’
cycle number at which the increase in miRNA probe fluorescence is exponential.
As reference control for miRNA quantification, Ct values were then
subtracted from Ct values obtained from exogenously added miR-206, as
described above. Ct values were then compared (?Ct) with each athlete’s
own resting baseline at the pre-season time point (normalized to fold-change of
1).

Basketball skills
Measurements:

Basketball skills were assessed before and after the
program as follow:

A)  Ball defense/obstacle
skills for 10 times.

B)   Ball passing to/from
another player at 10 yards during running for 10 times.

C)   Ball shooting to the
basket from the shooting circle for 10 times.

D)  Shooting to the basket from the running and jumping position for 10
times.

Result of total 40 points were calculated and
compared before to after program.

 

Statistical analysis:

A Computer program (SPSS
version 17) was used to analyze obtained results. Student’s Mean, standard deviation of
players and t test were used to examine
the difference between baseline and after exercise measurements before and
after the training program. Probability
was assigned significant at ? 0.05.

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