it also gives rise to long - term handicap 
 giving negative influences on activities of daily living in stroke patients1 , 2 . 

there are a lot of stroke sequelae which have bad effects on quality of life in stroke 
 patients . among these aftereffects ,
hemiparesis defined as motor impairments of the limb 
 contralateral to unilateral brain damage3 , 4 .
the increased activation in unaffected 
 hemisphere could be a possible cause of a pathological inhibition in affected hemisphere as 
 well as motor impairments in affected side of the body5 , 6 .
a number of researches on 
 a physical activity reported that there is a close relationship between a performance of 
 exercise and an improvement of brain plasticity7 .
skilled reaching can be considered as task - specific training via 
 its purposeful characteristics , so it is used as a training program to rehabilitate stroke 
 patients with a motor disability in their upper limb8,9,10 .
treadmill exercise has been studied as one of conventional 
 treatments for a long time . in general , treadmill exercise is commonly applied to stroke 
 patients thanks to its beneficial effect on neuroprotection11 , 12 .
to date , the only 
 therapy approved by fda has been thrombolytic tissue plasminogen activator ( tpa ) although 
 approximately 80% of human strokes are classified into ischemic stroke13,14,15 .
it is suggested that continuous researches via 
 experimental ischemic stroke models should be performed to develop novel , proper , and 
 helpful solutions for patients with ischemic stroke . in this
regards , the present study 
 investigated effects of skilled reach training with affected forelimb and treadmill exercise 
 on the expression of neurotrophic factor following ischemia - induced brain injury in 
 rats .
thirty male sprague - dawley rats were divided into 3 groups randomly : namely , the control 
 sacrified 2 weeks after surgery ( con ) , skilled reach training with forepaw contralateral to 
 brain injury for 2 weeks ( sc ) , and treadmill exercise for 2 weeks ( te ) .
all the experiments 
 were performed in accordance with protocols approved by the animal experiment committee in 
 daegu university , based on the nih guidelines for the care and use of laboratory animals . 

transient focal cerebral ischemia was induced by intraluminal occlusion of the left middle 
 cerebral artery ( mca ) .
briefly , right common carotid artery ( cca ) was exposed and the 
 external carotid artery ( eca ) and the cca were ligated .
middle cerebral artery is occluded 
 by a 40 nylon monofilament coated with a silicone tip , the monofilament was inserted into 
 the internal carotid artery from the external carotid artery until mild resistance was felt . 

reperfusion was established by completely withdrawing the nylon monofilament after 120  min 
 of occlusion .
skilled reach training was conducted in a plexiglas chamber ( 45  cm in height , 
 15  cm in width , 40  cm in length ) containing of a 1  cm by 10  cm window in front wall allowed 
 for one of the rat s paw to reach via for a pellet .
animals were acclimated to the chamber 
 with 4.5  mg of sugar - flavored food pellets ( research diets , new brunswich , nj , usa ) placed 
 on the shelf placed in a small indentation 3  cm from the inside wall of the chamber and 
 trained in 30 minutes sessions administrated 6 days per week for 2 weeks based on previous 
 reports .
when a pellet was removed by a rat , the shelf was refilled for a time period of 30 
 minutes continuously .
a motor - driven treadmill was used for the treadmill exercise . before 
 treadmill exercise , rats in te experienced 1 days of adaptive running exercise at a speed of 
 69 m / min for 5  min .
the brains of each group were collected , washed twice in pbs , 
 and then homogenized and lysed with buffer ( 137  mm nacl , 8.1  mm na2hpo4 , 2.7  mm kcl , 1.5  mm 
 kh2po4 , 2.5  mm edta , 1  mm dithiothreitol , 0.1  mm pmsf , 10  g / ml leupeptin [ ph 7.5 ] ) for 
 30  min on ice .
the lysates were centrifuged for 10  min at 15,000  rpm at 4  c , and the 
 protein concentration was determined as described previously .
equal amounts of protein 
 ( 40  g ) were resolved via 10% sodium dodecyl sulfate - polyacrylamide gel electrophoresis 
 ( sds - page ) and transferred to nitrocellulose membranes .
the blots were washed with tbst 
 ( 10  mm tris  hcl [ ph 7.6 ] , 150  mm nacl , 0.05% tween 20 ) , blocked with 5% skim milk for 1 
 hour , then incubated with the appropriate primary antibodies at the dilutions recommended by 
 the suppliers .
the membranes were washed , and the primary antibodies were detected using 
 horseradish peroxidase - conjugated goat anti - rabbit igg or goat - anti mouse igg .
the bands 
 were then visualized via enhanced chemiluminescence ( amersham pharmacia biotech , piscataway , 
 nj , usa ) .
the expression of brain - derived neurotrophic factor ( bdnf ) and nerve growth factor ( ngf ) in 
 con , sc , and te were also investigated to identify the effect of skilled reach training with 
 contralesional forelimb and treadmill exercise on bdnf expression .
there were significant 
 differences in bdnf and ngf expression between con and sc , con and te ( p<0.05 ) .
there 
 were no significant differences in bdnf and ngf expression between sc and te 
 ( p>0.05 ) .
motor function in upper - extremity is closely related to quality of life in stroke patients 
 via having influences on activity and participation , and this is the reason to focus on the 
 recovery of upper limb motor function in stroke patients16,17,18 .
skilled reach training , one of task - specific exercises , has 
 frequently been mentioned as a treatment strategy for stroke patients in many 
 researches19 , 20 . in many studies ,
skilled reach training was applied to animal 
 models in order to investigate its effect on neurological diseases .
pagnussat et al . 
 displayed an effect of skilled reaching with affected forelimb on functional recovery in 
 their research21 . above this
, several 
 studies also reported that recovery of motor function can be induced by applying skilled 
 reach training with contralesional forelimb9 , 22 .
in addition , effects of treadmill 
 exercise have been studied in many researches for a long time .
it has several beneficial 
 effects such as motor function recovery and neuroprotection12 . in many studies investigating recovery of a damaged brain , plasticity is often 
 mentioned23 .
particularly , it is 
 reported that the activation of neurotrophic factors is able to facilitate plasticity via 
 inducing generation and differentiation of neuronal progenitor cell24,25,26 .
park et al . reported that neurotrophic factors have an 
 influence on neuroprotective function after brain injury27 .
for these reasons , the present study investigated the expression of bdnf and ngf via 
 western blot analysis to examine the effect of skilled reach training and treadmill exercise 
 on neurotrophic factor expression . from results , there were significant differences in bdnf 
 and ngf expression between con and sc , con and te ( p<0.05 ) .
although expression of bdnf 
 and ngf in sc slightly increased compared with in te , the difference was not statistically 
 significant .
( p>0.05 ) ( table 1table 1.expression of bdnf and ngf in each 
 group ( unit : pixel)consctebdnf2,046.5 
  108.74,240.1  72.5 * 4,173.3  
 65.1*ngf2,211.3  
 87.93,881.6  91.4 *
3,830.8  
 88.7*values are reported as the mean 
  sd.*p<0.05 vs. con ) .
these results 
 imply that both skilled reach training with affected forelimb and treadmill exercise can 
 induce expression of neurotrophic factors .
it is suggested that skilled reach training may 
 have more influence on them than treadmill exercise because of its repetitive and 
 task - specific characteristics .
values are reported as the mean 
  sd . there is the limitation of the present study .
since the results of the study only showed 
 expression of bdnf and ngf , it is not enough to explain the relationship between expression 
 of neurotrophic factors and recovery of function .
further study applying tests for 
 evaluating motor function is necessary to make the results of the present study more 
 obvious . in consequence ,
especially , skilled reach training with affected forelimb showed more 
 beneficial effects on neurotrophic factor expression than treadmill exercise with a moderate 
 intensity .
 [ purpose ] the aim of the present study was to investigate effects of skilled reach 
 training with affected forelimb and treadmill exercise on the expression of neurotrophic 
 factor following ischemia - induced brain injury in rats . [ subjects and methods ] thirty male 
 sprague - dawley rats were divided into 3 groups randomly : namely , the control sacrified 2 
 weeks after surgery , skilled reach training with forepaw contralateral to brain injury for 
 2 weeks , and treadmill exercise for 2 weeks . 
 transient focal cerebral ischemia was induced 
 by intraluminal occlusion of the left middle cerebral artery . 
 after that , skilled reach 
 training and treadmill exercise were conducted . 
 western blot analysis was performed to 
 investigate expressions of neurotrophic factors . 
 [ results ] there were significant 
 differences in brain - derived neurotrophic factor and nerve growth factor expression 
 between the control group and the experimental group . 
 there were no significant 
 differences in brain - derived neurotrophic factor and nerve growth factor expression 
 between the skilled reach training group and the treadmill exercise group . 
 [ conclusion ] 
 skilled reach training and treadmill exercise can affect the expression of neurotrophic 
 factors . 
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION