세포 외 pH 변화가 인간 골수 기질세포의 증식 및 분화에 미치는 영향

임예현,장재석,남태석,김정화,이강식,이동호,윤준오 대한골대사학회 2012 대한골대사학회지 Vol.19 No.1

Objectives: The purpose of this study is to identify whether the change of pH affects the proliferation and the differentiation of human bone marrow stem cells (hBMSCs) and what mechanism is underlied. Methods: To achieve objective of this study, hBMSCs were cultivated in the conditioned media adjusted to potential of hydrogen (pH) ranging from 6.4 to 8.0 using addition of hydrochloric acid (HCl) and sodium hydroxide (NaOH). The ratio of proliferation of hBMSCs according to the change of pH was measured for 24 h, 48 h, and 72 h using water-soluble tetrazolium salt (WST)-8 method. To elucidate the mechanism involved, hBMSCs was subjected to blocking extracellular signal-regulated kinases (ERK) and calcium sensing receptor (CaSR) activation. The Osteogenicrelated genes and alkaline phosphatase (ALP) activity were tested under the conditioned media. Results: The proliferation of hBMSCs was promoted under extracellular alkali conditions (pH 7.6~8.0) via CaSR/ERK pathway. On the other hand, the differentiation was inhibited/delayed via decreased ALP activity besides gene expression at pH 8.0. Conclusion: Extracellular alkali or acidic surrounding according to pH alteration can play a crucial role in hBMSC behavior including the proliferation and the differentiation. 연구목적: 본 연구는 세포 외 pH 변화에 따른 골수 기질세포의 증식 및 분화에 어떠한 영향을 미치는 지와 그와 관련된기전을 알아보았다. 연구방법: 본 연구의 가설을 검증하기 위해 HCl과 NaOH를 이용하여 배양액의 pH를 6.4에서 8.0의 다양한 범위를 유도하여 골수 기질세포의 증식 및 분화를 확인하였다. 또한, 관련 기전을 알아보기 위해 CaSR 및 ERK의 활성 억제를 통해골수 기질세포의 증식을 관찰하였다. 또한, 분화능과 관련하여 골 형성 관련 유전자인 Runx2, ALP, 및 OCN의 mRNA 발현 양상과 ALP 활성을 관찰하였다. 결 과: 골수 기질세포의 증식은 pH 7.6~8.0에서 통계적으로 유의하게 증가함을 보였으며, 이러한 변화는 CaSR/ERK pathway와 관련이 있음을 보였다. 반면에, pH 7.6 혹은 8.0에서는 감소된 ALP 및 OCN mRNA 발현 감소와 ALP 활성의감소가 관찰되었다. 결 론: 골수 기질세포의 증식 및 분화 능력은 다양한 요인에 의해 변화될 수 있는데, 그 중 세포 외 pH 변화에 따른알칼리 및 산성 환경도 하나의 중요한 요인으로 작용될 수 있음을 시사한다.

탈진적인 1회성 수영운동과 단시간 금식 복합 조건이 쥐의 심장근과 골격근의 GLUT-4와 GRP-78 발현에 미치는 영향

임예현(Im Ye Hyeon),조준용(Jo Jun Yong),정국현(Jeong Gug Hyeon),이규성(Lee Gyu Seong),오윤선(O Yun Seon) 한국사회체육학회 2003 한국사회체육학회지 Vol.19 No.2

This study investigates the distribution and heart and skeletal levels of glucose regulated protein(GRP-78) and glucose transport protein(GLUT-4) in response to starvation(48hr) and swimming exercise in the rats. Results demonstrate that GRP-78 is strongly expressed within the heart, plantaris and soleus muscle in responce to starvation(48hr) and swimming exercise and glucose transport protein(GLUT-4) is also expressed with in the soleus and plantaris muscle in responce to swimming exercise and starvation & swimming exercise in the rats GRP78 levels are highest during fasting in both the heart and plantaris muscle whereas are highest during fasting & swimming exercise in only soleus muscle and GLUT-4 levels are highest during swimming exercise in both the soleus and plantaris muscle, considering the tissue-specific pattern of GRP-78 and GLUT-4 expression and change during exposing to varous stress condition.

카테킨 섭취와 운동수행에 따른 obese Zucker rat 골격근의 GLUT-4 VAMP-2와 Syntaxin-4 단백질 발현의 변화

김철현,임예현,조인호,김성찬,방상식,조준용,이규성 한국운동과학회 2003 운동과학 Vol.12 No.4

김철현, 임예현, 조인호, 김성찬, 방상식, 조준용, 이규성, 카테킨 섭취와 운동수행에 따른 obese Zucker rat 골격근의 GLUT-4, VAMP-2와 Syntaxin-4 단백질 발현의 변화. 운동과학, 제12권 제4호, 663-678, 2003. 골격근에서 인슐린의 자극은 GLUT4를 세포막으로 전좌하도록 활성화하여 세포내로 글루코스를 공급할 수 있도록 한다. GLUT4의 전좌는 특이적인 액포(vesicle)결합 단백질인 v-SNARE가 표적 세포막에 위치한 상동성이 큰 t-SNAHE(Syntaxin-4)단백질을 특이적으로 인식하고 상호작용하여 GLUT-4를 세포막 표면으로 가역적 전좌(translocation)를 시켜 글루코스 섭취 능력을 증가시킨다. 본 연구는 v-와 t-SNARE 단백질의 상호 신호전달 작용 및 GLUT-4 작용 기전을 규명하고, 운동과 녹차 성분인 카테킨 섭취에 따른 골격근의 인슐린 저항성에 미치는 영향을 구명하는데 그 목적이 있다. 이를 위해, 본 연구는 비만동물로 확립된 obese Zucker rat(fa/fa)를 사용하여 실험을 수행하였다. 집단은 비교집단, 카테킨 섭취, 운동집단, 운동과 카테킨 섭취 집단(4집단)으로 구분하여 8주간 실험을 실시하였다. 실험결과 카테킨 섭취는 운동수행은 독립적으로 obese Zucker rat의 골격근내 인슐린 조절 단백질인 GLUT-4와 Vamp-2, Syntaxin-4 단백질 발현을 증가시킨 것으로 나타났다. 이는 운동 혹은 카테킨은 각각 독립적으로 골격근의 글루코스 섭취 능력 및 인슐린 저항성을 개선시키는데 도움이 되는 것을 확인할 수 있었다. 즉, 운동 영양학 측면에서 카테킨과 같은 항당뇨 관련 보조물 섭취는 운동과 함께 비만과 당뇨병 개선 효과를 극대화시키는데 도움이 될 것으로 나타났다. 그러나 운동에 의한 GLUT-4의 세포막 이동 외에 카테킨 섭취가 GLUT-4의 세포막으로 이동에 있어서 또 다른 기전에 관여할 가능성이 있어 카테킨 섭취에 따른 GLUT-4 이동의 기전을 다시 확인하기 위한 후속 연구가 필요하다. Kim, C.H., Leem, Y.H., Cho, I.H., Kim, S.C., Bang, S.S., Cho, J.Y.,Lee, K.S. The effect of exercise training and catechins supplementation on skeletal muscle GLUT-4, VAMP-2 and Syntaxin-4 protein expression in obese Zucker rats. Exercise Science, 12(4): 653-670, 2003. Studies have suggested an important role for synaptobrevin and syntaxin homologues in this event, particularly the v-soluble N-ethylmaleimide attachment protein receptors(SNAREs)cellubrevin and vesicle-associated memo r ane protein-2(VAMP-2) and the syntaxin-4(t-SNARE) but the expression of these proteins has not been studied in insulin resistant tissues. There fore, we examined SNARE protein content in skeletal muscle from obese Zucker rats and determined the effect of exorcise and/or catechin treatment on these proteins. GLUT4 levels in skeletal muscle from exorcise and/or catechin treatments were elevated compared with control group. VAMP-2, and syntaxin 4 protein levels were elevated in skeletal muscle from obese Zuker rats compared with controls Restoration of normo-glycemia and normo-insulinemia in exorcised obese Zuker rats improved VAMP-2, and syntaxin-4 protein to levels approaching normal in lean animals. These data show that elevated v- and t-SNARE protein levels are associated with insulin resistance in skeletal muscle and that these increase may be achieved by exorcise and/or catechin treatments concomitant with a restoration of glycemic control. in summary, we have demonstratedthat v- and 1- SNAHE concentration as well as muscle GLUT-4 protein and mRNA can increase significantly after prolonged exorcise and/or catechin, and confirmed that this increase in GLUT-4 protein is controlled at both the pre-translational and translational levels. The observation that improved glucose storage suggests that the increased expression of GLUT-4 protein expression can serve as a molecular yardstick for exorcise-dependent glycogen deposition.

The effect of exercise training on Amyloid-β and learning and memory behaviors in NSE/PS2m Alzheimer`s disease model mice

Cho, Joon Yong,Leem, Yea Hyun,Lee, Kyu Sung,Hwang, Dae Hyun,Kim, Yong Kyu 한국운동과학회 2002 운동과학 Vol.11 No.2

조준용, 임예현, 이규성, 장대연, 김용규. The effect of exercise training on Amyloid-β and learning and memory behaviors in NSE/PS_2m Alzheimer's disease model mice. 운동과학, 제11권 제2호, 441-452, 2002. 이 연구는 hPS_(2m)을 neuron-specific enolase (NSE) 유전자의 프로모터 조절하에 연결시킨 융합 유전자를 이식한 12개월된 알츠하이머 질환 모델동물을 대상으로 hPS_(2m) 유전자의 조절이 뇌조직에 특이적으로 발현되는지를 조사한 결과, hPS_(2m) mRNA의 발현은 뇌, 근육, 폐, 심장과 간 순서로 나타났으며 신장과 소장에서는 볼 수 없었다 12주간 트레드밀 운동 수행에 따른 hPS_(2m) 형질전환 생쥐의 인지적·행동적 결함을 개선시키는지를 수중미로검사 (water maze test) 를 통하여 검증하고, 뇌조직의 AB_42 발현량 및 골격근의 AB_42을 분석하고 뇌조직의 Presenllin-2 발현 및 지질 성분을 관찰한 결과 다음 같은 결과를 얻었다. 먼저 수중미로검사를 통하여 속도 (escape velocity), 거리 (escape distance), 시간 (escape latency) 을 관찰한 결과, Tg 비교집단에 비해 Tg 트레드밀 운동집단이 향상된 것으로 나타났으며 뇌조직에서, AB_42 발현량은 Tg 비교집단에 비해 Tg 트레드밀 운동집단이 상대적으로 적은 것으로 나타났다. 또한 골격근에서도 AB_42 발현량은 Tg 비교집단에 비해 Tg 트레드밀 운동집단이 상대적으로 감소된 것으로 나타났다. 12주간 트레드밀 운동수행에 따른 혈청 지질 성분을 비교한 결과, Tg 비교집단에 비해 Tg 트레드밀 운동집단이 HDL-C는 높고, TC, TG, LDL-C는 낮은 것으로 것으로 나타났다. 결론적으로 12주간 트레드밀 런닝은 뇌조직과 골격근에서, AB_42의 침적을 감소시키고, 혈청 지질 성분을 감소시킨다는 측면에서 볼 때, 12주간 트레드밀 운동은 알츠하이머 질환을 어느 정도 지연시키는데 도움을 준다는 것을 확인하였다. Cho, J.Y, Leem, Y.H., Lee, K.S., Hwang D.H., Kim, Y.K. Elevation of Aβ_42 has been confirmed in the skeletal muscle of Alzheimer's Disease (AD) patients. Exercise Science, 11(2): 441-452, 2002. We used the transgenic model of AD that express a human mutant PS_2 (N1411) transgene (hPS_2m) under the regulation of the rat neuron-specific enolase (NSE) promoter. These transgenic mice have behavioral deficit at 12 months of age that are accompanied by increasing levels of Aβ_42 Unexpectively, mice expressed skeletal muscle-specific expression of hPS_2m gene that had previously been confined to the brain. We examined the protective effect of exercise on A/2 deposits in the skeletal muscle, brain, behavioral deficit, and serum cholesterol of transgenic mice respectively. To investigate this, the transgenic mice were subjected to treadmill exercise during the course of the experiment. Upon treatment, the levels of the Aβ_42 deposits were significantly reduced in both the skeletal muscle and the brain, and also behavioral deficits were improved. In addition, TG and LDL levels were significantly reduced The results suggest that treadmill exercise in transgenic mice resulted in reduction at the low levels of cholesterol, thereby reducing Aβ_42 secretion. Thus, these transgenic mice are useful experimental system in which to study the protective effect of exercise on improving behavioral function and reducing Aβ_42 deposition in the skeletal muscle and brain of transgenic mice. Treadmill exercise may represent practical strategy applying for human subjects with Alzheimer's Disease in the preventive management on the basis of these scientific evidences.

운동수행에 따른 쥐의 골격근 형태별 UCP-3 mRNA의 변화

양대승,김철현,임예현,김문희,이승국,방상식,오유성,조준용,이규성 한국운동과학회 2003 운동과학 Vol.12 No.4

양대승, 김철현, 임예현, 김문희, 이승국, 방상식, 오유성, 조준용, 이규성. 운동과학, 제12권 제4호, 653-662, 2003. 운동수행에 따른 쥐의 골격근 형태별 UCP-3 mRNA의 변화. 이 연구는 비만 모델동물인 Obese Zucker Rat을 활용하여 12주간 지구성 운동이 골격근형태별 UCP-3 mRNA 발현, citrate synthase 활성도 및 혈청 지질 수준에 영향을 미치는가를 분자생물학적·생화학적 수준에서 평가한 결과, 장기간 지속적인 지구성 운동에 대한 골격근의 적응은 UCP-3 mRNA 발현량 감소 및 citrate synthase 활성도 증가와 혈청 지질(혈청 콜레스테롤, 중성지방, 저밀도 지단백 감소) 수준을 감소시키는 것으로 나타났다. 그러나 골격근의 UCP-3 유전자의 발현은 추위, 반응 산소종, 식이, 당뇨병 등과 같은 여러가지 요인과도 상관이 있기 때문에 운동과 관련된 보다 많은 연구들이 차후에 수행되어야 할 것이다. Yang, D.S., Kim, C.H, Leem, Y.H., Kim M.H., Lee, S.K,Bang, S.S., Oh, Y.S., Cho,J.Y., Lee, K.S. Thechanges of UCP-3 mRNA of obese Zucker rats during exercise training, Exercise Science, 12(4): 653-662, 2003. The factors that regulate gene expression of uncoupling protein-3 (UCP-3) in skeletal muscle are poorly understood, but uncoupling protein-3 gene is clearly responsive to the metabolic state of organism. There fore, We tested the hypothesis that chronic endurance exercise would alter UCP-3 gene expression. After 12wk endurance treadmill exercise, UCP-3 mRNA level of trained obese Zucker rat were decreased [vs. lean-type Zucker rat (100%) vs. obese Zucker rat control] in ail two tissues. in addition, cholesterol, triglyceride and LDL-C levels were significantly reduced in trained rats, whereas citrate synthase activites were significantly increased in trained rats. The results of this study show trot a need for a higher metabolic efficiency is associated with decreased mRNA expression of the uncoupling protein-3 (UCP-3) which would decrease energy dissipation in these tissues.

Acid sphingomyelinase inhibition alleviates muscle damage in gastrocnemius after acute strenuous exercise

이영익,임예현 한국운동영양학회 2019 Physical Activity and Nutrition (Phys Act Nutr) Vol.23 No.2

[Purpose] Strenuous exercise often induces skeletal muscle damage, which results in impaired performance. Sphingolipid metabolism contributes to various cellular processes, including apoptosis, stress response, and inflammation. However, the relationship between exercise-induced muscle damage and ceramide (a key component of sphingolipid metabolism), is rarely studied. The present study aimed to explore the regulatory role of sphingolipid metabolism in exercise-induced muscle damage. [Methods] Mice were subjected to strenuous exercise by treadmill running with gradual increase in intensity. The blood and gastrocnemius muscles (white and red portion) were collected immediately after and 24 h post exercise. For 3 days, imipramine was intraperitoneally injected 1 h prior to treadmill running. [Results] Interleukin 6 (IL-6) and serum creatine kinase (CK) levels were enhanced immediately after and 24 h post exercise (relative to those of resting), respectively. Acidic sphingomyelinase (A-SMase) protein expression in gastrocnemius muscles was significantly augmented by exercise, unlike, serine palmitoyltransferase-1 (SPT-1) and neutral sphingomyelinase (N-SMase) expressions. Furthermore, imipramine (a selective A-SMase inhibitor) treatment reduced the exercise-induced CK and IL-6 elevations, along with a decrease in cleaved caspase-3 (Cas-3) of gastrocnemius muscles. [Conclusion] We found the crucial role of A-SMase in exercise-induced muscle damage.

Effects of exercise intensity on the reactive astrocyte polarization in the medial prefrontal cortex

어수주,임예현 한국운동영양학회 2023 Physical Activity and Nutrition (Phys Act Nutr) Vol.27 No.2

[Purpose] Physical exercise contributes to neuroplasticity by promoting cognitive functions, such as learning and memory. The astrocytic phenotype is closely associated with synaptic plasticity. This study aimed to determine whether astrocyte polarization and synaptic alterations in the medial prefrontal cortex (mPFC) are affected differently by high- and moderate-intensity exercise. [Methods] Mice were subjected to moderate- (MIE) and high-intensity treadmill running (HIE). Memory capacity was assessed using the novel object recognition and modified Y-maze tests. For immunohistochemistry, c-Fos-positive cells were counted in the mPFC. Using western blot analysis, astrocyte phenotype markers were quantified in whole-cell lysates, and synaptic molecules were determined in the synaptosomal fraction. [Results] Exercise lengthened the approach time to novel objects regardless of intensity in the NOR test, whereas MIE only improved spatial memory. Exercise induced c-Fos expression in the anterior cingulate cortex (ACC) and c-Fos-positive cells were higher in MIE than in HIE in the ACC area. In the prelimbic/infralimbic cortex region, the number of c-Fos-positive cells were enhanced in MIE and decreased in HIE mice. The A1 astrocyte marker (C3) was increased in HIE mice, while the A2 astrocyte markers were enhanced in exercised mice, regardless of the intensity. In the synaptosomal fraction, synaptic proteins were elevated by exercise regardless of intensity. [Conclusion] These results suggest that exercise intensity affects neuronal plasticity by modulating the reactive state of astrocytes in the mPFC.

The effects of peripherally-subacute treatment with irisin on hippocampal dendritogenesis and astrocyte-secreted factors

김문희,임예현 한국운동영양학회 2019 Physical Activity and Nutrition (Phys Act Nutr) Vol.23 No.4

[Purpose] Fibronectin type III domain containing 5 (FNDC5)/irisin is an exercise-induced myokine, which contributes to cognitive functions. However, the relationship between the neuroprotective effects of FNDC5/irisin and hippocampal dendritic remodeling and astrocyte-secreted factors remains unclear. Therefore, we explored whether subchronic recombinant irisin treatment affected hippocampal morphology and some astrocyte-derived molecules. [Methods] Mice were intraperitoneally injected with irisin (0.5 µg/kg/day) for seven days, followed by their sacrifice two days later. Hippocampal morphometric parameters were analyzed and pgc-1a, fndc5, bdnf,and some astrocyte-derived factors mRNA levels were measured. [Results] Dendritic length, arborization, and spine density were enhanced by irisin regimen in hippocampal CA1 and CA3 areas. Hippocampal pgc-1a, fndc5, and bdnf mRNA levels were significantly increased by irisin treatment. Moreover, hevin mRNA levels were significantly enhanced, whereas tgf-b1 levels downregulated by irisin treatment. [Conclusion] FNDC5/irisin has dendritogenic activity probably through hevin induction and TGF-β1 suppression.

Neurogenic effect of exercise via the thioredoxin-1/ extracellular regulated kinase/β-catenin signaling pathway mediated by β2-adrenergic receptors in chronically stressed dentate gyrus

김문희,임예현 한국운동영양학회 2019 Physical Activity and Nutrition (Phys Act Nutr) Vol.23 No.3

[Purpose] Chronic stress is a precipitating factor for depression, whereas exercise is beneficial for both the mood and cognitive process. The current study demonstrates the anti-depressive effects of regular exercise and the mechanisms linked to hippocampal neurogenesis. [Methods] Mice were subjected to 14 consecutive days of restraint, followed by 3 weeks of treadmill running, and were then subjected to behavioral tests that included the forced swimming and Y-maze tests. Protein levels were assessed using western blot analysis and newborn cells were detected using 5-bromo-2′-deoxyuridine (BrdU). [Results] Three weeks of treadmill running ameliorated the behavioral depression caused by 14 days of continuous restraint stress. The exercise regimen enhanced BrdU-labeled cells and class III β-tubulin levels in the hippocampal dentate gyrus, as well as those of thioredoxin-1 (TRX-1) and synaptosomal β2-adrenergic receptors (β2-AR) under stress. In vitro experiments involving treatment with recombinant human TRX-1 (rhTRX-1) augmented the levels of phospho-extracellular signal-regulated kinases 1 and 2 (ERK1/2), nuclear β-catenin, and proliferating cell nuclear antigens, which were previously inhibited by U0216 and FH535 (inhibitors of ERK1/2 and β-catenin/T cell factor-mediated transcription, respectively). The hippocampal neurogenesis elicited by a 7-day exercise regimen was abolished by a selective inhibitor of β2-AR, butoxamine. [Conclusion] These results suggest that TRX-1-mediated hippocampal neurogenesis by β2-AR function is a potential mechanism underlying the psychotropic effect of exercise.

The potential role of creatine supplementation in neurodegenerative diseases

장혁기,임예현 한국운동영양학회 2023 Physical Activity and Nutrition (Phys Act Nutr) Vol.27 No.4

[Purpose] The maintenance of energy balance in the body, especially in energy-demanding tissues like the muscles and the central nervous system, depends on creatine (Cr). In addition to improving muscle function, Cr is necessary for the bioenergetics of the central nervous system because it replenishes adenosine triphosphate without needing oxygen. Furthermore, Cr possesses anti-oxidant, anti-apoptotic, and anti-excitotoxic properties. Clinical research on neurodegenerative illnesses has shown that Cr supplementation results in less effective outcomes. With a brief update on the possible role of Cr in human, animal, and in vitro experiments, this review seeks to offer insights into the ideal dosage regimen. [Methods] Using specified search phrases, such as “creatine and neurological disorder,” “creatine supplementation and neurodegenerative disorders,” and “creatine and brain,” we searched articles in the PubMed database and Google Scholar. We investigated the association between creatine supplementation and neurodegenerative illnesses by examining references. [Results] The neuroprotective effects of Cr were observed in in vitro and animal models of certain neurodegenerative diseases, while clinical trials failed to reproduce favorable outcomes. [Conclusion] Determining the optimal creatinine regime for increasing brain creatinine levels is essential for maintaining brain health and treating neurodegeneration.