지구성 운동에 의하여 증가된 PPARβ/δ는 NRF-1 promoter 활성을 통하여 생쥐 근육내 GLUT4 및 미토콘드리아 효소의 발현을 유도한다

고진호 ( Jin Ho Koh ),김기진 ( Ki Jin Kim ),정수련 ( Su Ryun Jung ),김상현 ( Sang Hyun Kim ) 한국운동생리학회 2015 운동과학 Vol.24 No.2

PURPOSE: The propose of this study was to evaluate whether PPARβ/δ can increase mitochondrial enzymes and GLUT4 through NRF-1 promoter activity. METHODS: To evaluate PPARβ/δ and NRF-1 protein expression in mouse skeletal muscle, TA muscle were dissected at 24 h after 10-day swimming exercise (Ex). To evaluate PPARβ/δ effects on gene regulation, PPARβ/δ was overexpressed in mouse TA muscle using electrical pulse-mediated gen transfer (electroporation; EPO) method. We conducted chip and promoter reporter assay to confirm that PPARβ/δ can bind NRF-1 promoter and activates NRF-1 promoter in mouse muscle cell. RESULTS: PPARβ/δ and NRF-1 protein expression in skeletal muscle of 10-day Ex group were increased 2.2 and 2.7 fold, respectively, when compare to non-exercise muscle. PGC-1α protein expression in PPARβ/δ overexpression skeletal muscle were not increased, but NRF-1, MEF2A, GLUT4, ATP syn and Cytp C expression were increased 2.2, 2.3, 2.1, 2 and 1.7 fold, respectively, when compare to empty vector (EV) treated muscle. Our ChiP and promoter activity assay result show that NRF-1 promoter has a PPARβ/δ binding site (PPRE) and PPARβ/δ can activates NRF-1 promoter through the PPRE. CONCLUSIONS: The increased PPARβ/δ by endurance exercise can control MEF2A-GLUT4 and mitochodrial enzymes such as ATP syn and Cytp C through NRF-1 promoter activity.

PPARβ / δ 과발현은 PGC-1α ubiquitination의 감소를 통해 PGC-1α 단백질을 증가시킨다

고진호 ( Jin Ho Koh ),김홍수 ( Hong Soo Kim ),김기진 ( Ki Jin Kim ) 한국운동생리학회 2015 운동과학 Vol.24 No.4

PURPOSE: The aim of this study was to find mechanism with which PPARβ/δ control PGC-1αprotein stability in mouse skeletal muscle. METHODS: Tibialis anterior (TA) muscles of rat were dissected at 18h after 2 weeks swimming exercise. PPARβ/δ was over-expressed in mouse tibialis anterior (TA) muscle using electrical pulse-mediated gen transfer (electroporation; EPO) method. To evaluate, PGC-1α mRNA and protein, PPARβ/δ, ubiquitin and mitochondrial enzyme protein expression in mouse skeletal muscle, TA muscle were dissected at 2wk and 4wk after EPO. To evaluate binding between PGC-1α and ubiquitin, we conducted V5-PGC-1α Immunoprecipitation. RESULTS: PPARβ/δ and PGC-1α in swimming skeletal muscle for 2 weeks were increased 2.1 and 2.5 fold respectively than sedentary muscle. PGC-1α mRNA in PPARβ/δ over-expression skeletal muscle were not different at 2 wk and 4wk when compared to an empty vector (EV) treated group. Endogenous and exogenous PPARβ/δ in PPARβ/δ over-expression skeletal muscle were increased 2.5 and 3.8 fold respectively when compared to an EV treated group. We identified binding between PGC-1α and ubiquitin, and ubiquitin were decreased 60% in PPARβ/δ transfected C2C12 when compared to an EV treated group. PGC-1α were increased 3 fold in muscle that ubiquitin were decreased 60% when compared to an EV treated muscle and COXI and CYTO C were increased 2.27 and 1.75 fold respectively in muscle when compared to an EV treated muscle. CONCLUSIONS: PPARβ/δ overexpression increases PGC-1α protein through the decreased PGC-1α ubiquitination without the increased PGC-1α mRNA in mouse skeletal muscle.

원문 : PGC-1α단백질이 생쥐 근육세포의 글리코겐 분해 및 해당효소의 발현을 조절한다

고진호 ( Jin Ho Koh ),김기진 ( Ki Jin Kim ),김상현 ( Sang Hyun Kim ),정수련 ( Su Ryun Jung ) 한국운동생리학회(구 한국운동과학회) 2014 운동과학 Vol.23 No.4

본 연구는 골격근 세포인 C2C12 myotube에 ppargc1a(PGC-1α 유전자) mRNA를 silence시키는 기법으로 PGC-1α의 발현을 부분적으로 억제하여 글리코겐 분해 및 해당 효소의 발현에 어떠한 영향을 미치는지 확인하는 것이다. C2C12골격근 세포에 PGC-1α의 과발현을 유도하기 위하여 Ad-PGC-1α adenovirus를 처치하였으며, PGC-1α의 발현을 억제하기 위하여ppargc1a mRNA를 silence 시키는 기법을 이용하였다. 골격근 세포에 PGC-1α의 과발현을 유도한 결과, PFK의 발현이 empty vector를 처치한 그룹보다 0.4배 감소한 것으로 나타났다. 골격근 세포에 PGC-1α의 발현을 부분적으로 억제시킨 결과 phosphorylase kinase, glycogen phosphorylase 및 PFK는 empty vector를 처치한 그룹보다 각각 2배, 1.4배, 1.5배 증가한 것으로 나타났다. 본 연구의 결과는 단기간 지구성 운동 후 빠르게 증가하는 PGC-1α가 골격근 세포 내 글리코겐 분해효소 및 해당효소를 간접적으로 조절하여 글리코겐 절약효과를 유도할 수 있음을 증명하였다. The purpose of this study was to evaluate the role of PGC-1α on the expression of glycolytic and glycogenolytic enzyme in C2C12 mouse skeletal muscle cell. PGC-1α overexpression in C2C12 myotube was achieved by PGC-1α encoded adenovirus. When PGC-1α was overexpressed, PFK expression was suppressed 0.4 fold compared to control, empty vector treated cell. Contrary to PGC-1α overexpression, silencing ppargc1a mRNA (suppressed PGC-1α protein expression) induced 2, 1.4 and 1.5 fold increase in phosphorylase kinase, glycogen phosphorylase, and PFK protein expression respectively. In summary, down regulation of expression of the glycogenolytic and glycolytic enzymes in muscle that mediates a slowing of muscle glycogen depletion appears to be through the PGC-1α expression.

장기간 지구성 훈련은 생쥐 골격근의 Slow Twitch Myosin Heavy Chain 발현과 미토콘드리아 생합성을 증가시킨다

고진호 ( Jin Ho Koh ),서현규 ( Hyun Gyu Suh ),김기진 ( Ki Jin Kim ) 한국운동생리학회 2016 운동과학 Vol.25 No.1

PURPOSE: The purpose of this study was to identify spatial distribution of type I myosin heavy chain among different muscle fibers by endurance exercise for 8 week. METHODS: The animal were trained for 8 week a on treadmill. To avoid acute exercise effect of training, Tibialis anterior (TA) muscles were dissected 18 hours after the last bout of exercise. To evaluate different type myosin heavy chain and mitochondria expression, we performed themulticolor immunofluorescence analysis and western-blotting. RESULTS: Type IIb and IIx fiber in the 8-week trained group decreased trained skeletal muscle for 8 weeks were decreased 46% and 51% respectively than the sedentary group. Type IIa and I in the 8-week trained group increased 2.2 and 1.7 folds respectively than the sedentary group. ATP synthase, cytochrome C, NADH and SERCA2a in the 8-week trained increased 2.5, 2.2, 2.8, and 2.5 folds respectively than the sedentary group. TA muscles showed increased in the hybrid fiber type distribution. Type I and lla MHC increased at all type II fiber the trained group than the sedentary group. CONCLUSIONS: The results of the study raised the possibility that the muscle fiber type conversion may occur from all type II muscle fibers to type IIa and I MHC through 8 weeks of long-term endurance exercise, instead of going through the step-by-step conversion process.

원문 : PPARδ가 생쥐 골격근의 미토콘드리아 생합성에 미치는 영향

고진호 ( Jin Ho Koh ),김상현 ( Sang Hyun Kim ),정수련 ( Su Ryun Jung ),김기진 ( Ki Jin Kim ) 한국운동생리학회(구 한국운동과학회) 2014 운동과학 Vol.23 No.4

본 연구는 골격근에 PPARδ의 과발현이 미토콘드리아 효소의 발현에 어떠한 영향을 미치는지 확인하는 것이다. 전기자극유전자 전이기법을 이용하여 생쥐의 골격근에 PPARδ의 과발현을 유도하였으며, PGC-1α가 증가하기 전과 증가된 시점에서 전자전달계 효소인 NADH dehydrogenase(NADH), cytochrome c oxidase subunit Ⅳ(COXⅣ) 및 succinate-ubiquinone oxidoreductase(SUO)와 구연산회로의 효소인 citrate synthase(CS)의 발현을 분석하였다. 생쥐의 골격근에 PPARδ가 2주 또는 4주간 과발현된 근육은 PPARδ가 대조군보다 각각 2.1배 2.8배 증가하였다. PPARδ가 2주간 과발현된 근육은 PGC-1α, COXⅣ, CS가 대조군과 차이가 없는 것으로 나타났으나, NADH와 SUO는 대조군보다 각각 1.5배 2.4배 증가하였다. PPARδ가 4주간 과발현된 근육은 PGC-1α, COXⅣ, CS, NADH 그리고 SUO가 대조군보다 각각 1.6배, 2.1배, 1.8배, 2배 그리고 2.5배 증가하였다. 본 연구는 PPARδ가 PGC-1α의 단백질 발현을 증가시키거나 또는 다른 기전을 통하여 미토콘드리아 효소의 발현을 증가시킬 수 있음을 증명하였다. The purpose of this study was to evaluate the role of PGC-1α and PPARδ on the mitochondrial biogenesis in mouse skeletal muscle. PPARδ overexpression in mouse skeletal muscle was achieved by electroporation (EPO). To evaluate mitochondrial biogenesis, such as NADH dehydrogenase (NADH), cytochrome c oxidase subunit Ⅳ (COXⅣ), citrate synthase (CS) were analyzed 2 and 4 weeks post PPARδ overexpression by EPO. Two and four weeks after PPARδ EPO, PPARδ expression in muscle was increased 2.1 and 2.8 fold respectively when compared to empty vector (EV) treated muscle. Two weeks after PPARδ EPO in muscle, PGC-1α, COXⅣ, and CS were not increased, but NADH and SUO were increased 1.5 and 2.4 fold respectively when compared to EV treated muscle. Four weeks after PPARδ EPO in muscle, PGC-1α, COXⅣ, CS, NADH, and SUO were increased 1.6, 2.1, 1.8, 2, and 2.5 fold respectively when compared to EV treated muscle. Thus, this study demonstrated that PPARδ can increase PGC-1α by posttranscriptional mechanism causing mitochondrial biogenesis, and PPARδ can affect directly or indirectly mitochodrial biogenesis without PGC-1α.

PPARδ의 활성 또는 과발현은 PGC-1α 단백질의 분해를 지연시킨다

고진호 ( Jin Ho Koh ),김홍수 ( Hong Soo Kim ),안나영 ( Na Young Ahn ),김기진 ( Ki Jin Kim ) 한국운동생리학회 2015 운동과학 Vol.24 No.1

PURPOSE: The purpose of this study was to evaluate whether PPARδattenuates PGC-1α protein degradation in mouse skeletal muscle after a single bout of swimming exercise (Ex). METHODS: PPARδactivation in mouse skeletal muscle was achieved by high fat diet (HFD) for 2 weeks. PPARδoverexpression in mouse skeletal muscle was achieved by electroporation (EPO). To evaluate PGC-1α, cytochrome c oxidase subunit Ⅳ (COXⅣ) and succinate- ubiquinone oxidoreductase (SUO) protein in mouse skeletal muscle, tibialis anterior (TA) muscle were dissected at 24 h or 54 h after a single bout of swimming exercise. RESULTS: PGC-1α and mitochondrial enzymes in all exercised muscle were significantly increased by a single bout of swimming exercise when compared to non-exercised muscle. PGC-1α expression in 54 h post-Ex Chow group was decreased 60% when compared to 24 h post-Ex Chow group, but 54 h post-Ex HFD group was not significantly decrease when compared to 24 h post-Ex HFD. PGC-1αexpression in 54 h post-Ex HFD group was higher 1.4 fold than 54 h post-Ex Chow group. PGC-1α expression in 54 h post-Ex EV group was decreased 75% when compared to 24 h post-Ex Ev group, but 54 h post-Ex PPARδ group was decreased 1% when compare to 24 h post-Ex PPARδ. PGC-1α expression in 54 h post-Ex PPARδ group was 1.9 fold higher than 54 h post-Ex EV group. COXⅣ와 SUO expression in 54 h post-Ex PPARδ group were 1.4 and 1.94 fold higher than 54 h post-Ex EV group. CONCLUSIONS: This study demonstrated that PPARδ activation or overexpression attenuates PGC-1α protein degradation in skeletal muscle after a single bout of swimming exercise that causes a more increase mitochondrial biogenesis.

지구성 운동에 의한 AMPK 활성은 MEF2A 통하여 PPARβ/δ의 발현을 조절한다

고진호 ( Jin-ho Koh ),안나영 ( Na-young Ahn ),김기진 ( Ki-jin Kim ) 한국운동생리학회(구 한국운동과학회) 2016 운동과학 Vol.25 No.2

PURPOSE: The aim of this study was to identify whether the increased PPARβ/δ gene expression induced by exercise is mediated by activation of AMPK and MEF2A. METHODS: To analyze PPARβ/δ mRNA, rat triceps muscles were dissected immediately after a single bout of swimming exercise, and constitutively active (CA) AMPK over expressed L6 rat skeletal muscle cells were harvested. MEF2A activation by AMPK was analyzed by a Gal4 and green fluorescence system in the L6 cells. Dominant negative AMPK was over expressed in rat tibialis anterior (TA) using electrical pulse-mediated gen transfer (electroporation [EPO]) method before 2 weeks swimming exercise. RESULTS: PPARβ/δ mRNA in rat TA muscle after exercise was increased by 5.5 fold compared with the sedentary muscle, and was increased by 6.7 fold in the CA-AMPK over expressed cells compared with the empty vector (EV) treated cells. MEF2A-Gal4 was activated by a 1.9 fold in the L6 cells transfected with CA-AMPK constructs. AMPK and HDAC5 phosphorylation, and PPARβ/δ expression were increased 3.2, 2.8, and 2.1 fold, respectively, by exercise in EV treated muscle, but were not increased by exercise in DNAMPK over expressed muscle. CONCLUSIONS: MEF2A activation by AMPK increases the PPARβ/δ expression in endurance exercised muscle.

아동의 비만지표에 따른 심혈관계 질환 및 인슐린 저항성의 위험성 선별 및 예측능력 비교

고진호(Jin Ho Koh),김홍수(Hong Soo Kim),김기진(Ki Jin Kim) 한국발육발달학회 2014 한국발육발달학회지 Vol.22 No.2

This study was to analyze the selection and predictive capability of risk of cardiovascular disease and insulin resistance according to obese indices in childhood. We studied 569 2~5 grade elementary school students who were divided into 2 groups(obese & normal) according to body mass index(BMI) and degree of obesity(DO). Subjects were measured for cardiovascular disease factors and insulin. The statistics were analyzed through t-test, correlation, R-square, and regression of cardiovascular disease and insulin resistance according to obese indices. Both obese indices were significant( p<.05) in correlation with cardiovascular disease factors and insulin concentration. All cardiovascular disease factors, except total cholesterol(TC) concentration, including insulin concentration and HOMA-IR, between the normal and obese groups, according to BMI, were significant( p<.05) difference. But the normal and obese groups according to degree of obesity(DO), were significant except glucose and TC concentrations. The R square of BMI(31.1%) was higher than DO(27.8%). the area under curve(AUC) of BMI(0.78) by ROC curve was higher than DO(0.68). Cutoff point of insulin resistance risk according to BMI index for childhood was 1.975, in which sensitivity and specificity were respectively 70.1% and 71.5%. obesity decision over the 95 percentile in childhood BMI according to age and gender yielded a stronger prediction & usefulness on risk of cardiovascular disease and insulin resistance than DO. HOMA-IR over the 95 percentile in childhood BMI according to age and gender yielded a high risk probability of 1.975.

운동 중 골격근 내 PGC-1α 농도는 PPARβ/δ를 통한 후전사 기전에 의하여 조절된다: PPARβ/δ silence가 ubiquitin을 통한 PGC-1α 안정성에 미치는 영향

정수련 ( Su Ryun Jung ),김기진 ( Ki Jin Kim ),고진호 ( Jin Ho Koh ) 한국운동생리학회 2015 운동과학 Vol.24 No.3

PURPOSE: The aim of this study was to identified which PPARβ/δ control PGC-1α protein stability in rat skeletal muscle by long-term endurance exercise. METHODS: shPPARβ/δ was over-expressed in rat epitrochlearis (Epi) muscle using electrical pulse-mediated gen transfer (electroporation; EPO) method. Then animal underwent 2 wk long swimming exercise. To evaluate PPARβ/δ, PGC-1α ubiquitination, PGC-1α and mitochondrial enzyme expression in rat skeletal muscle, Epi muscles were dissected 18 h post final bout of swimming exercise. RESULTS: PPARβ/δ, PGC-1α, COX I and NADH protein expression in shPPARβ/δ over-expression sedentary (Sed) skeletal muscle were significantly decreased over 55%, but PGC-1α ubiquitination was significantly increased 2.6 fold when compared to scramble (Scr) treated Sed muscle. PPARβ/δ, PGC-1α, COX I and NADH protein expression in 2 wks swimming exercise with Scr gene treated muscle were significantly increased over 2 fold, but PGC-1α ubiquitination was significantly decreased 66% when compared to Scr gene treated Sed muscle. PPARβ/δ, PGC-1α, COX I and NADH protein expression in 2 wks swimming exercise with shPPARβ/δ gene treated muscle were not increased, but PGC-1α ubiquitination was significantly increased 1.96 fold when compared to Scr gene treated Sed muscle. CONCLUSIONS: Our results indicate that PPARβ/δ controls PGC-1α protein stability in rat skeletal muscle following long-term endurance exercise.

일회성 저강도 지구성운동 시 MEF2A를 통한 PPARδ의 GLUT4 생합성 조절

김상현 ( Sang Hyun Kim ),김기진 ( Ki Jin Kim ),정수련 ( Su Ryun Jung ),고진호 ( Jin Ho Koh ) 한국운동생리학회 2015 운동과학 Vol.24 No.1

PURPOSE: There is increasing evidence that the regulator of skeletal muscle metabolism, PPARδ is associated with GLUT4 biogenesis. But, whether acute low-intensity endurance exercise affects PPARδ and the mechanism of GLUT4 biogenesis has not been clearly identified. Even Though GLUT4 expression is increased by acute low-intensity endurance exercise. PPARδ expression by acute low-intensity endurance exercise is unclear. PPARδ was shown to induce the expression of PGC-1α in skeletal muscle. Since both are increased with acute low-intensity endurance exercise, it is difficult to determine whether one or both, induce GLUT4 biogenesis in skeletal muscle. METHODS: This study measured PPARδ expression in skeletal muscle by acute low-intensity endurance exercise and confirmed the effect of PPARδ on GLUT4 biogenesis in PGC-1α KO mice. Sixteen rats were randomly assigned into 2 groups(Sedentary, Sed & 1 day exercise, 1d Ex). Exercise groups performed a 6 hours low-intensity swimming program. RESULTS: The expressions of GLUT4, PGC-1α and PPARδ were increased in the 1d Ex group from the Sed group. PPARδ was overexpressed using electric pulse-mediated gene transfer technique in PGC-1α KO mice to confirm effects of PPARδ for GLUT4 biogenesis without PGC-1α. The expressions of GLUT4 and MEF2A were increased in PPARδ overexpressed muscles. CONCLUSIONS: These findings suggest that PPARδ-driven GLUT4 biogenesis through MEF2A by acute low-intensity endurance exercise was independent of PGC-1α.