이 연구는 한국 건강 기술 R & D 프로젝트 통해는 한국 보건 산업 개발 연구소 (진흥원), 사역의 건강 및 복지, 한국에 의해 투자 되었다는에서 교부 금에 의해 지원 되었다 (허가 번호: HI16C1559).

프로토콜은 헬싱키의 선언에 따라 실시 하 고 프로토콜 차 분당 병원 기관 검토 위원회에 의해 승인 되었다.
1. 퇴행 성 힘 줄 조직 수확은 환자에서
<ol><li>의료 기록을 신체 검사 결과 사용 하 여 측면 염증 진단: 측면 뼈; 가까이 단 근원 부드러움 손목 관절의 저항된 확장에 의해 갖는 고통.</li>
	<li>다음 포함 기준을 사용 하 여: 18 세 이상; 최소 6 개월;에 대 한 측면 염증의 역사 비 수술 치료; 대 한 고집 불통 적어도 3 개월이 코르 티 코 스테로이드, 혈소판 리치 플라즈마, 또는 보 톨 리누스 독 소의 주입 이후</li>
	<li>다음 제외: 임신 여성; 자 궁 경관 등뼈 질병, 팔꿈치 관절염, rheumatologic 질병, 팔꿈치, 그리고 신경 함정 수사 증후군; 주위 수술의 역사를 가진 환자 조직 기증을 거부 하는 환자.</li>
	<li>
수술 기법 참고: 그림 124참조.<ol>
<li>Isoflurane endotracheal 삽 관 법에 의해 일반 마 취, 후 테이블에 팔을 넣고는 지 혈 대를 적용. 그런 다음 팔 전체에 무 균 수술 시트와 드 레이프 betadine 에이전트를 적용 합니다.</li>
		<li>15 번 외과 용 메스 블레이드 그냥 anteromedial 측면 뼈와 관절의 수준에 인접 된 3-5 cm 절 개를 통해 측면 팔꿈치를 노출 합니다.</li>
		<li>시각 신 근 피 radialis longus (ECRL) 및 신 근 aponeurosis 인터페이스를 식별 하 고 식별 된 인터페이스에서 15 번 외과 용 메스 블레이드와 ECRL 및 신 근 aponeurosis 사이 깊이에 분할 절 개 2-3 m m를 확인 합니다.</li>
		<li>추출 된 ECRL anteriorly,는 직접 보기에 아래 pathologic 신 근 피 radialis longus brevis (ECRB) 원산지를 가져올 것 이다.</li>
		<li>시각 특성 단조로운 잿 빛 색에 의해 pathologic 퇴행 성 조직 및 일반적으로 edematous 및 부서 조직을 식별 합니다.</li>
		<li>확인 후 신중 하 게 모든 퇴행 성 조직 급격히 15 번 외과 용 메스와 미니-블레이드를 사용 하 여 resect.</li>
		<li>버퍼링 하는 인산 염 (PBS)에 즉시 (약 1 c m3) 절제 조직을 포함 하 고 약 10 mL의 PBS로 신중 하 게 최대한 빨리 힘 줄 조직 제외 하 고 모든 주변 조직을 제거 합니다.</li>
		<li>측면 뼈에서 뼈 exostosis 경우, exostosis는 rongeur를 사용 하 여 제거 하 고는 강판으로 매끄럽게 합니다. 게 여러 드릴 구멍 (직경 1.6 m m, 약 6-8 구멍) 혈관 공급을 향상 시키기 위해 anterolateral 관절 돌기에. 이것은 선택 사항입니다.</li>
		<li>레이어, 레이어 비 흡수 봉합 사 물자를 사용 하 여 상처를 닫습니다.</li>
	</ol>
</li>
	<li>
건강 한 제어 tenocyte
	<ol>
<li>전방 십자 인 대 재건, 동안 자동 햄 스트링 또는 자동-슬 개 골 힘 줄의 남은 힘 줄에서 건강 한 tenocytes를 얻을 하 고 다음 실험25컨트롤로 사용 합니다. 참고:이 연구에 사용 된 샘플 수가 각 그룹에 3 했다.</li>
	</ol>
</li>
</ol>2입니다. 조직학
<ol><li>화학 증기 두건 또는 이와 동등한 중립 버퍼링 된 10% 포 르 말린에 수확된 조직의 절반을 저장 합니다.</li>
	<li>조직을 파라핀 블록 (4-5 mL)에 포함 하 고 coronally 4-5 µ m 섹션에 섹션.</li>
	<li>
제어 및 Hematoxylin 및 오신 얼룩 (H & E)와 2.5의 pH에 Alcian 블루 얼룩 퇴행 성 힘 줄에서 섹션을 얼룩. H & E 자동된 얼룩 기계에 얼룩이 지 다음과 같이 수행 합니다.
	<ol>
<li>크 실 렌을 사용 하 여 deparaffinize (3 시간, 5 분, 각각).</li>
		<li>물 (100% 알코올, 4 분, 100% 알코올, 3 분, 95% 알코올, 2 분, 70% 알코올, 1 분, 물, 2 분) 등급된 알콜에서 하이드 레이트.</li>
		<li>5 분 2 분 동안 물에 잘 세척에 대 한 되며에 얼룩.</li>
		<li>1% 산 알코올 차별화 (1% 70% 알코올에 HCl) 3 3 분 물에 잘 씻어 s..</li>
		<li>찍기 10는 0.5% 암모니아에 의해 HCl를 중화 s, 뒤에 3 분 물 세척.</li>
		<li>5 분 동안 Eosion에 얼룩.</li>
		<li>알콜을 통해 탈수 (80% 알코올, 30 100% 알코올 1 분, 100% 알코올 95% 알코올, 1 분; s 2 분).</li>
		<li>크 실 렌에 취소 (3 시간, 2 분, 2 분, 3 분, 각각).</li>
		<li>표지 슬라이드와 함께 탑재.</li>
	</ol>
</li>
	<li>
Immunohistochemitry (IHC) 사소한 수정 제조업체의 지침에 따라 채권 폴리머 수정 탐지 키트를 사용 하 여 수행 합니다.
	<ol>
<li>Deparaffinization 크 실 렌을 사용 하 여, 후 100 ° c.에 30 분 동안 본드 Epitope 검색 솔루션을 사용 하는 항 원 검색 절차를 수행 채권 Epitope 검색 솔루션 기반 시트르산 버퍼와 계면 활성 제 (1l, pH 5.9-6.1)에 포함 되어 있습니다.</li>
		<li>5 분 동안 과산화 수소와 조직 배양 하 여 생 peroxidases를 끄다.</li>
		<li>혈관 내 피 성장 인자 (VEGF) (1: 200), tenomodulin (1: 200), 또는 모호크 (1: 200)에 대 한 1 차 항 체와 온도 (21-22 ° C)에서 15 분에 대 한 섹션을 품 어.</li>
		<li>이차 항 체 고분자 biotin 무료 양 고추냉이 과산화 효소-링커 항 체 어원이 시스템 태그를 수행 합니다.<ol>
<li>안티 사용 보조 토끼 마우스 항 체를 지역화 하려면 Tris 버퍼 saline/0.09% 2-Methyl-4-isothiazolin-3-one 솔루션에서 10% (v/v) 동물 혈 청에서 IgG를 마우스.</li>
			<li>고분자 반대로 토끼 폴 리 HRP IgG를 사용 하 여 (< 25 µ g/mL) 10% (v/v) 동물 혈 청 지역화 토끼 항 체를 saline/0.09% Tris 버퍼에 포함 된.</li>
			<li>3, 3'-Diaminobenzidine tetrahydrochloride 수화물 갈색 침전을 통해 복잡 한 시각화를 안정제 솔루션에서 사용 합니다.</li>
			<li>와 함께 counterstain < 셀 핵 시각화를 0.1% 되며 (파란색).</li>
		</ol>
</li>
	</ol>
</li>
	<li>H 물 슬라이드 관찰 & E, Alcian 파랑 얼룩 및 IHC 가벼운 현미경 VEGF, tenomodulin, 및 모호크에 대 한 하나의 높은 전력 분야 (400 X)에서 각 슬라이드의 대표적인 미세한 이미지를 얻을.</li>
</ol>3. Tenocyte 문화
<ol><li>
조직 준비
	<ol>
<li>마이크로-가 위, 집게와 철저 하 게 모든 주변 조직을 제거 하 여 힘 줄 샘플을 준비 합니다.</li>
		<li>PBS로 세척 하 고 작은 조각 (약 1 m m3 이하로)으로 말하다.</li>
		<li>Dulbecco의 수정 독수리의 매체 (DMEM) 37 ° c.에 30 mg/mL 콜라 II와 보충의 30 mL에 1 시간에 대 한 조직을 소화합니다</li>
		<li>100 µ m 셀 스 트레이너를 통해 세포를 전달 합니다.</li>
		<li>소 태아 혈 청 (FBS)의 1 mL을 추가 하 여 콜라 II를 비활성화 합니다.</li>
		<li>5 분 196 x g에서 원심.</li>
	</ol>
</li>
	<li>
세포 배양
	<ol>
<li>100 mm 접시에 3 x 105 셀을 씨 고 DMEM 10 %FBS 1% 항생제 Antimycotic 솔루션 5% CO2 배양 기에서 37 ° C에서 1 주에 대 한 보완과 문화.</li>
		<li>문화 매체 마다 3 일 80% 합류 발생할 때까지 변경 합니다. 통로 4까지 건강 한 제어 및 퇴행 성 셀 샘플을 문화 및 실험을 위해 사용.</li>
	</ol>
</li>
</ol>4입니다. Immunocytochemistry (ICC)
<ol><li>챔버 슬라이드의 8 개의 우물을 교양된 셀 (2 x 105 셀)의 200 µ L를 전송 하 고 1 일에 대 한 신선한 미디어의 추가 함께 합류 하 셀 성장.</li>
	<li>PBS로 세포 세척 하 고 잘 당 4% 포름알데히드의 500 µ L로 수정.</li>
	<li>0.5%와 슬라이드를 품 어 세포 막의 permeabilization에 대 일 분에 대 한 PBS에 트라이 톤 X-100.</li>
	<li>PBS에서 1% 소 혈 청 알 부 민 및 0.05% 차단 트윈-20 모호크와 tenomodulin, 각각.</li>
	<li>마우스 방지 모호크 단일 클론 항 체 (1: 100 희석)와 4 ° C에서 염소 안티-tenomodulin 항 체 (1: 100 희석) 어둠 속에서 밤새 품 어.</li>
	<li>2 차 항 체 (알 렉 사 555-안티 염소 (1: 200) tenomodulin;에 대 한을 품 어 알 렉 사 488-안티 마우스 (1: 200) 모호크) 실 온에서 1 h에 대 한 어둠 속에서.</li>
	<li>형광 장착 매체의 0.5 µ L로 탑재 합니다.</li>
	<li>형광 현미경 (400 X)를 사용 하 여 분석 합니다.</li>
</ol>5. 확산 분석 결과
<ol><li>
셀 viabilities 수용 성 tetrazolium (서 부 표준시)-1 사용 하 여 측정 합니다.
	<ol>
<li>24, 72, 및 120 h 잘 당 2 x 103 셀의 밀도에서 96 잘 접시로 모두 건강 하 고 퇴행 성 셀 씨.</li>
		<li>부 화, 후 각 음을 10% 서 부 표준시-1 솔루션의 10 µ L를 추가 합니다.</li>
		<li>5% CO2에서 37 ° C에서 3 h에 대 한 번호판을 품 어.</li>
		<li>450에서 광학 밀도 측정 nm microplate 리더를 사용 하 여. 각 샘플에 대 한 모든 시간 포인트에서 세 번 이상 읽기를 가져가 라.</li>
	</ol>
</li>
	<li>
측정 세포질 DNA 양 BrdU 분석 결과 사용 하 여입니다.
	<ol>
<li>둘 다 건강 한 씨앗 하 고 96에 타락 셀 잘 잘 24, 72, 및 120 h 당 2 x 103 셀의 밀도에 격판덮개.</li>
		<li>부 화, 후 각 음을 10 %BrdU 솔루션의 10 µ L를 추가 합니다.</li>
		<li>5% CO2에서 37 ° C에서 3 h에 대 한 번호판을 품 어.</li>
		<li>100 µ L/잘 고정/Denaturing 솔루션의 추가 (60-70% 에탄올, 0.1-0.5%가 성소 다) 30 분 동안 실내 온도에 접시를 유지.</li>
		<li>1 시간에 대 한 탐지 항 체 솔루션 및 계속 실 온에서 플레이트 x 1을 준비 하는 100 µ L/잘 추가 합니다.</li>
		<li>솔루션을 제거 하 고 세 번 워시 버퍼 x 1와 접시 세척.</li>
		<li>100 µ L/잘 준비 1 x HRP 활용 된 이차 항 체 솔루션을 추가 하 고 30 분 동안 실내 온도에 접시를 유지.</li>
		<li>세 번 세척 버퍼 x 1와 솔루션 및 세척 접시를 제거 합니다.</li>
		<li>50 %TMB 기판의 100 µ L를 추가 합니다.</li>
		<li>실 온에서 30 분 동안 품 어.</li>
		<li>정지 솔루션의 100 µ L를 추가 합니다.</li>
		<li>450에서 광학 밀도 측정 nm microplate 리더를 사용 하 여. 참고: 걸릴 읽기 적어도 세 번 모든 시간 포인트 각 샘플에 대 한.</li>
	</ol>
</li>
</ol>6. 통계 분석
<ol><li>수집 하 고 SPSS를 사용 하 여 데이터를 분석.</li>
	<li>의미의 평균 ± 표준 오차에 의해 데이터를 묘사.</li>
	<li>사용만 휘트니 U 테스트 합니다.</li>
	<li>때 p 값이 0.05 보다 작으면 통계적 차이 고려 (p < 0.05).</li>
</ol>

<ol>
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J., Fong, G., Andersson, G., Scott, A., Danielson, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Substance+P+is+a+mechanoresponsive,+autocrine+regulator+of+human+tenocyte+proliferation.">Substance P is a mechanoresponsive, autocrine regulator of human tenocyte proliferation.</a> PLoS One. 6 (11), 27209 (2011).</li><li>Backman, L. J., Eriksson, D. E., Danielson, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Substance+P+reduces+TNF-alpha-induced+apoptosis+in+human+tenocytes+through+NK-1+receptor+stimulation.">Substance P reduces TNF-alpha-induced apoptosis in human tenocytes through NK-1 receptor stimulation.</a> Britich Journal of Sports Medicine. 48 (19), 1414-1420 (2014).</li><li>Rolf, C. G., Fu, B. 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저자는 공개 없다.

이전 연구의 숫자는 콜라 또는 kartogenin 주입, 실행, 디딜 방 아 등 더 많은26,27다른 절차를 사용 하 여 만성 tendinopathic 동물 모델을 만드는 방법을 보고 있다. 수많은 연구가 동물 모델에 따라 유망한 치료 효과 보여, 비록 인간의 퇴행 성 tenocyte를 사용 하 여 실험 것입니다 중요 한 tendinopathy의 분야에서 치료의 효능을 재생 하기 위하여. 이 문서에서는, 우...

Tendinopathy 신체의 다양 한 부분에서 개발 하 고 만성 퇴행 성 근 골격 계 상태 이다. 최근, tendinopathy의 케이스의 수에 참여 레크리에이션 스포츠 및 증가 평균 수명1,2성장으로 인해 선진국에서 크게 증가 했다. Tendinopathy의 원인 multifactorial 간주 됩니다 및 이러한 원인은 허 혈, 산소 자유 래 디 칼 부상, vasoconstrictor 및 vasodilator innervations, 내부 마이크로-눈물, 그리고 신경 레 귤 레이 션3에 변화 사이의 불균형 포함 ,,45,6,,78. Tendinopathy에 대 한 대부분의 치료만의 증상을 완화. 또한, 조직 재생 없이 치료 재활에 대 한 오랜 시간을 요구 하 고 의사9임상 과제 부과 부상된 힘 줄에서 제한 된 응답 달성.
현재 치료 옵션과 자체에 퇴행 성 힘 줄의 능력의 부족의 무는 대체 치료 전략을 탐구에 관심을 가져 리드 연구원. 최근, 성장 인자를 사용 하 여 tendinopathy 힘 줄의 치료 효능을 향상을 위한 많은 유망한 결과 보고 하는 새로운 연구, 줄기 세포 기반 치료, 그리고 유전자 치료10,,1112.
문학 검토를 통해 우리는 관련 된 연구의 분석 자료를 기반으로 두 가지 범주로 나눌 수 있습니다 발견: 쥐, 마우스, 또는 토끼; 같은 동물 모델 그리고 인간의 모델입니다. 동물 모델에 관해서는 현재 두 개의 인기 있는 기술이 있다 tendinopathy 생성 하: 부상 또는 모델을 오버 로드 하는 기계 화학 유도. 그러나, 각 동물 모델은 복잡 한 인간의 tendinopathy 병리학13,14재현에 제한 되었다.
대부분 논문 인간의 샘플을 사용 하 여 조직학 분석 했다 또는는 생체 외에서 퇴행 성 tenocyte15,16,,1718 대신 건강 한 인간의 tenocyte를 기반으로 하는 실험 , 19 , 20 , 21. 몇 논문만 보고 그들이 인간의 퇴행 성 tenocyte를 사용 하지만 그들은 않았다 하지 자세히 설명 인간의22,23에서 퇴행 성 tenocyte를 얻을 하는 데 사용 하는 프로토콜. 이러한 맥락에서 그것은 지적 되어야 인간의 효능 또는 효과 먹이 지 힘 줄 변성은 복잡 한 과정 때문에 병 인은 동물 모델 또는 건강 한 조직/tenocyte에서 성공적인 결과 반드시 예측 하지 수는 아직도 완전히 이해.
그것은 기증자에 게 부작용을 유발 하지 않고 인간의 조직에서 퇴행 성 tenocyte을 얻기 위한 표준 프로토콜을 설명 하는 데 필요한. 이 문서는 인간의 퇴행 성 tenocyte를 취득 하는 방법에 대 한 프로토콜을 설명 합니다. 프로토콜의 유효성을 검사 하려면 수확된 조직 조직학 분석 되었다. 그런 다음, 교양된 셀 immunocytochemistry (ICC), 정량 실시간 중 합 효소 연쇄 반응 (qRT-PCR), 생존 능력 분석 결과 사용 하 여 퇴행 성 tenocyte로 확인 되었다.

<table border="0" cellpadding="0" cellspacing="0" style="width:719px;" width="718">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Scalpel</td>
			<td style="width:199px;">Kisanbio</td>
			<td style="width:119px;">KS-Q0306-15</td>
			<td style="width:167px;">No. 15</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Mini-blade</td>
			<td style="width:199px;">Beaver</td>
			<td style="width:119px;">374769</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:235px;">Dulbecco&#39;s modified Eagle&#39;s medium (DMEM)</td>
			<td style="width:199px;">Gibco</td>
			<td style="width:119px;">11995065</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Collagenase &#8545;</td>
			<td style="width:199px;">Sigma-Aldrich</td>
			<td style="width:119px;">C6885</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">PBS</td>
			<td>Gibco</td>
			<td style="width:119px;">14190250</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">fetal bovine serum (FBS)</td>
			<td>Gibco</td>
			<td style="width:119px;">16000044</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">50 mM ascorbic acid-2-phosphate</td>
			<td style="width:199px;">Sigma-Aldrich</td>
			<td style="width:119px;">A5960</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Antibiotic-Antimycotic solution</td>
			<td>Gibco</td>
			<td style="width:119px;">15240062</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">4% formaldehyde</td>
			<td style="width:199px;">Bio-solution</td>
			<td style="width:119px;">BP031</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Triton X-100&#160;</td>
			<td style="width:199px;">Sigma-Aldrich</td>
			<td style="width:119px;">X100-100ml</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">BSA</td>
			<td style="width:199px;">Rdtech</td>
			<td style="width:119px;">C0082</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">TWEEN 20</td>
			<td style="width:199px;">Sigma-Aldrich</td>
			<td style="width:119px;">P9416-100ml</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">MKX (C-5)</td>
			<td style="width:199px;">Santa cruz biotechnology</td>
			<td style="width:119px;">sc-515878</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Tenomodulin (N-14)</td>
			<td style="width:199px;">Santa cruz biotechnology</td>
			<td style="width:119px;">sc-49325</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Fluorescence Mounting Medium</td>
			<td style="width:199px;">DAKO</td>
			<td style="width:119px;">S3023</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:235px;">DAPI (4&#39;,6-Diamidino-2-Phenylindole, Dihydrochloride)</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">D1306</td>
			<td></td>
		</tr>
		<tr height="46">
			<td height="46" style="height:46px;width:235px;">WST-1</td>
			<td style="width:199px;">Dojindo Molecular Technologies</td>
			<td style="width:119px;">CK04</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">BrdU Cell Proliferation Assay Kit</td>
			<td style="width:199px;">Cell Signaling Technology</td>
			<td style="width:119px;">#6813</td>
			<td></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:235px;">TRIzol Reagent</td>
			<td style="width:199px;">Invitrogen</td>
			<td style="width:119px;">15596018</td>
			<td></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:235px;">iScript cDNA Synthesis Kit&#160;</td>
			<td style="width:199px;">Bio-Rad</td>
			<td style="width:119px;">170-8891</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:235px;">TaqMan Gene Expression Master Mix</td>
			<td style="width:199px;">Applied Biosystems</td>
			<td style="width:119px;">4369016</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">GAPDH</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs02786624_g1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">COL3A1</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs00943809_m1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">ACTA2</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs00426835_g1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">TAC1</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs00243225_m1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">TACR1</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs00185530_m1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">PTGS2</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs00153133_m1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">ACTB</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">Hs99999903_m1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Cell Strainers (100 &#181;m)&#160;</td>
			<td style="width:199px;">Corning</td>
			<td style="width:119px;">352360</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">100mm culture dish</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">8188207</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">8-well Chamber Slide</td>
			<td style="width:199px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">154534</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:235px;">96 Well Clear Flat Bottom Polystyrene TC-Treated Microplates</td>
			<td style="width:199px;">Corning</td>
			<td style="width:119px;">3596</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Nikon Eclipse 50i Microscope&#160;</td>
			<td style="width:199px;">Nikon</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;">VERSA max microplate reader&#160;</td>
			<td style="width:199px;">Molecular Devices</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:235px;">CFX96 Real-Time PCR Detection System</td>
			<td style="width:199px;">Bio-Rad</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:235px;">Formalin solution, neutral buffered, 10%</td>
			<td style="width:199px;">Sigma-Aldrich</td>
			<td style="width:119px;">HT501128</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Paraffins</td>
			<td style="width:199px;">Leica Biosystems</td>
			<td style="width:119px;">3801340</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Ethanol</td>
			<td style="width:199px;">JUNSEI CHEMICAL</td>
			<td style="width:119px;">90303-2185</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Hematoxylin</td>
			<td style="width:199px;">DAKO</td>
			<td style="width:119px;">CS70030-2</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Eosin</td>
			<td style="width:199px;">DAKO</td>
			<td style="width:119px;">CS70130-2</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Alcian blue</td>
			<td style="width:199px;">DAKO</td>
			<td style="width:119px;">AR16011-2</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Citric acid</td>
			<td style="width:199px;">Sigma-Aldrich</td>
			<td style="width:119px;">251275</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Xylene</td>
			<td style="width:199px;">JUNSEI CHEMICAL</td>
			<td style="width:119px;">25165-0430</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Endogenous peroxidases&#160;</td>
			<td style="width:199px;">DAKO</td>
			<td style="width:119px;">S200380-2</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Canada balsam</td>
			<td style="width:199px;">JUNSEI CHEMICAL</td>
			<td style="width:119px;">23255-1210</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Microtome Blade</td>
			<td style="width:199px;">FEATHER</td>
			<td style="width:119px;">A35</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Slide glass</td>
			<td style="width:199px;">SUPERIOR</td>
			<td style="width:119px;">1000612</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Cover glass</td>
			<td style="width:199px;">Marienfeld-Superior</td>
			<td style="width:119px;">101050</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">VEGF</td>
			<td style="width:199px;">Santa cruz biotechnology</td>
			<td style="width:119px;">sc-7269</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">SPSS Software</td>
			<td style="width:199px;">IBM</td>
			<td style="width:119px;"></td>
			<td>Ver. 18.0</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:235px;">Multi-purpose Centrifuge</td>
			<td style="width:199px;">LABOGENE</td>
			<td style="width:119px;">1248R</td>
			<td></td>
		</tr>
	</tbody>
</table>

a protocol to acquire the degenerative tenocyte from humans

증가 신체 활동 및 더 긴 수명으로 인해 선진국에서 증가 Tendinopathy, 힘 줄 변성에 대 한 응답에서을 개발 하 고 고통 스러운 상태 이다. 그것의 증가 보급에도 불구 하 고 기본 병 인 아직도 불분명 하 게, 남아 있다 그리고 치료는 일반적으로 증상을 보인다. 최근, 성장 인자, 줄기 세포 및 유전자 치료 등 다양 한 치료 옵션은 퇴행 성 힘 줄의 치료 효능을 향상을 위해 조사 되었다. 그러나, 이러한 연구의 대부분은 동물 모델 또는 건강 한 인간 tenocytes에만 실시 되었다. 병 적인 tenocytes를 사용 하 여 일부 연구에도 불구 하 고 우리의 지식 최선을 다 해 현재는 인간의 퇴행 성 tenocytes을 구하는 방법을 설명 하는 아무 프로토콜. 이 연구의 목적은 인간의 퇴행 성 tenocytes 획득을 위한 표준 프로토콜을 설명 하는 것입니다. 처음에, 힘 줄 조직은 수술 하는 동안 측면 염증 환자에서 수확 했다. 다음 생 샘플 해당 수술 시 관찰 하는 구조적인 변화 하 신 근 피 radialis brevis 힘에서 촬영 됐다. 수확된 힘 줄의 모든 지루한, 회색, 어렵다면, 등장 하 고, edematous는 그들이 만든 건강 한 사람에서 시각적으로 뚜렷한. Tenocytes 배양 하 고 실험을 위해 사용 했다. 한편, 수확 조직의 절반 조직학, 분석 되었다 그리고 tendinopathy angiofibroblastic 형성 (증식) 같은 주요 기능을 공유 하는 그들은 보였다. Immunocytochemistry에 의해 2 차 분석 확인 배양된 세포 모호크와 tenomodulin 단백질에 대 한 긍정적인 얼룩을 갖는 셀의 대다수와 가진 tenocytes 했다. Tenocytes의 퇴행 성 자연의 자질 다음 확산 분석 결과 또는 qRT-PCR를 사용 하 여 건강 한 컨트롤 셀 비교 하 여 결정 했다. 퇴행 성 tenocyte 더 높은 확산 속도 일치 하는 이전 보고서 tendinopathy의 유사한 유전자 표현 패턴 표시. 전반적으로,이 새로운 프로토콜 tendinopathy의 미래 연구에 대 한 유용한 도구를 제공할 수 있습니다.

조직학 분석 공개 측면 염증에서 수확된 조직 tendinopathic 힘 줄의 특성을 했다. H & E 섹션 tendinopathy 퇴행 성 힘 줄의 극성 및 좋은 직선, 강하게 포장 병렬 섬유 구조 손실 무질서 콜라겐 번들을 밝혔다. 조직학 징후 높은 cellularity 등 전형적인 스핀 들 모양 없이 확대 핵 변성의 암시 샘플에서 일반적 이었다. 또한, 퇴 화 한 힘 줄의 콜라겐 번들 상대적으로 약한 오신 H & E 스?...

Watch this Scientific Journal Video about A Protocol to Acquire the Degenerative Tenocyte from Humans at JoVE.com

A Protocol to Acquire the Degenerative Tenocyte from Humans

Tendinopathy에 새로운 치료의 효능을 조사할 때 퇴행 성 tenocytes의 생체 외에서 사용은 필수적입니다. 그러나, 대부분의 연구 연구 동물 모델 또는 건강 한 tenocyte를 사용합니다. 우리는 수술 하는 동안 인간의 퇴행 성 tenocytes를 분리 하는 다음 프로토콜을 제안 합니다.

인간에서 퇴행 성 Tenocyte를 취득 하는 프로토콜

Tendinopathy, a painful condition that develops in response to tendon degeneration, is on the rise in the developed world due to increasing physical ...

a-protocol-to-acquire-the-degenerative-tenocyte-from-humans

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

7.3K 조회수.  CHA University School of Medicine. Tendinopathy에 새로운 치료의 효능을 조사할 때 퇴행 성 tenocytes의 생체 외에서 사용은 필수적입니다. 그러나, 대부분의 연구 연구 동물 모델 또는 건강 한 tenocyte를 사용합니다. 우리는 수술 하는 동안 인간의 퇴행 성 tenocytes를 분리 하는 다음 프로토콜을 제안 합니다.

비디오: A Protocol to Acquire the Degenerative Tenocyte from Humans

The Trier Social Stress Test Protocol for Inducing Psychological Stress

This article demonstrates a psychological stress protocol for use in a laboratory setting. Protocols that allow researchers to study the biological pathways of the stress response in health and disease are fundamental to the progress of research in stress and anxiety.1 Although numerous protocols exist for inducing stress response in the laboratory, many neglect to provide a naturalistic context or to incorporate aspects of social and psychological stress. Of psychological stress protocols, meta-analysis suggests that the Trier Social Stress Test (TSST) is the most useful and appropriate standardized protocol for studies of stress hormone reactivity.2 In the original description of the TSST, researchers sought to design and evaluate a procedure capable of inducing a reliable stress response in the majority of healthy volunteers.3 These researchers found elevations in heart rate, blood pressure and several endocrine stress markers in response to the TSST (a psychological stressor) compared to a saline injection (a physical stressor).3 Although the TSST has been modified to meet the needs of various research groups, it generally consists of a waiting period upon arrival, anticipatory speech preparation, speech performance, and verbal arithmetic performance periods, followed by one or more recovery periods. The TSST requires participants to prepare and deliver a speech, and verbally respond to a challenging arithmetic problem in the presence of a socially evaluative audience.3 Social evaluation and uncontrollability have been identified as key components of stress induction by the TSST.4 In use for over a decade, the goal of the TSST is to systematically induce a stress response in order to measure differences in reactivity, anxiety and activation of the hypothalamic-pituitary-adrenal (HPA) or sympathetic-adrenal-medullary (SAM) axis during the task.1 Researchers generally assess changes in self-reported anxiety, physiological measures (e.g. heart rate), and/or neuroendocrine indices (e.g. the stress hormone cortisol) in response to the TSST. Many investigators have adopted salivary sampling for stress markers such as cortisol and alpha-amylase (a marker of autonomic nervous system activation) as an alternative to blood sampling to reduce the confounding stress of blood-collection techniques. In addition to changes experienced by an individual completing the TSST, researchers can compare changes between different treatment groups (e.g. clinical versus healthy control samples) or the effectiveness of stress-reducing interventions.1

This article describes a protocol for inducing psychological stress in participants, which enables researchers to measure psychological, physiological and neuroendocrine responses to stress within single participants or between groups.

심리적 스트레스를 유발을위한 모금 사회 스트레스 테스트 프로토콜

This article demonstrates a psychological stress protocol for use in a laboratory setting.  Protocols that allow researchers to study the biological ...

연구

의학

Spectral Karyotyping to Study Chromosome Abnormalities in Humans and Mice with Polycystic Kidney Disease

Conventional method to identify and classify individual chromosomes depends on the unique banding pattern of each chromosome in a specific species being analyzed 1, 2. This classical banding technique, however, is not reliable in identifying complex chromosomal aberrations such as those associated with cancer. To overcome the limitations of the banding technique, Spectral Karyotyping (SKY) is introduced to provide much reliable information on chromosome abnormalities.
SKY is a multicolor fluorescence in-situ hybridization (FISH) technique to detect metaphase chromosomes with spectral microscope 3, 4. SKY has been proven to be a valuable tool for the cytogenetic analysis of a broad range of chromosome abnormalities associated with a large number of genetic diseases and malignancies 5, 6. SKY involves the use of multicolor fluorescently-labelled DNA probes prepared from the degenerate oligonucleotide primers by PCR. Thus, every chromosome has a unique spectral color after in-situ hybridization with probes, which are differentially labelled with a mixture of fluorescent dyes (Rhodamine, Texas Red, Cy5, FITC and Cy5.5). The probes used for SKY consist of up to 55 chromosome specific probes 7-10.
The procedure for SKY involves several steps (Figure 1). SKY requires the availability of cells with high mitotic index from normal or diseased tissue or blood. The chromosomes of a single cell from either a freshly isolated primary cell or a cell line are spread on a glass slide. This chromosome spread is labeled with a different combination of fluorescent dyes specific for each chromosome. For probe detection and image acquisition,the spectral imaging system consists of sagnac interferometer and a CCD camera. This allows measurement of the visible light spectrum emitted from the sample and to acquire a spectral image from individual chromosomes. HiSKY, the software used to analyze the results of the captured images, provides an easy identification of chromosome anomalies. The end result is a metaphase and a karyotype classification image, in which each pair of chromosomes has a distinct color (Figure 2). This allows easy identification of chromosome identities and translocations. For more details, please visit Applied Spectral Imaging website (<a href="http://www.spectral-imaging.com/" target="_blank">http://www.spectral-imaging.com/</a>).
SKY was recently used for an identification of chromosome segregation defects and chromosome abnormalities in humans and mice with Autosomal Dominant Polycystic Kidney Disease (ADPKD), a genetic disease characterized by dysfunction in primary cilia 11-13. Using this technique, we demonstrated the presence of abnormal chromosome segregation and chromosomal defects in ADPKD patients and mouse models 14. Further analyses using SKY not only allowed us to identify chromosomal number and identity, but also to accurately detect very complex chromosomal aberrations such as chromosome deletions and translocations (Figure 2).

Spectral Karyotyping (SKY) is an advanced cytogenetics technique to identify genomic and chromosomal aberrations. This technique takes advantage of chromosome painting probes, which allow classification of all chromosomes. SKY can also identify complex chromosome aberrations and segregation defects in mice and humans with various diseases, including polycystic kidney disease.

Polycystic 신장 질환과 함께 인간과 마우스의 염색체 이상을 공부하는 스펙트럼 Karyotyping

Conventional method to identify and classify individual chromosomes depends on the unique banding pattern of each chromosome in a specific species being ...

Heterotopic Renal Autotransplantation in a Porcine Model: A Step-by-Step Protocol

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality of life when compared to dialysis. Although the last few decades have seen major improvements in patient outcomes following kidney transplantation, the increasing shortage of available organs represents a severe problem worldwide. To expand the donor pool, marginal kidney grafts recovered from extended criteria donors (ECD) or donated after circulatory death (DCD) are now accepted for transplantation. To further improve the postoperative outcome of these marginal grafts, research must focus on new therapeutic approaches such as alternative preservation techniques, immunomodulation, gene transfer, and stem cell administration.
Experimental studies in animal models are the final step before newly developed techniques can be translated into clinical practice. Porcine kidney transplantation is an excellent model of human transplantation and allows investigation of novel approaches. The major advantage of the porcine model is its anatomical and physiological similarity to the human body, which facilitates the rapid translation of new findings to clinical trials. This article offers a surgical step-by-step protocol for an autotransplantation model and highlights key factors to ensure experimental success. Adequate pre- and postoperative housing, attentive anesthesia, and consistent surgical techniques result in favorable postoperative outcomes. Resection of the contralateral native kidney provides the opportunity to assess post-transplant graft function. The placement of venous and urinary catheters and the use of metabolic cages allow further detailed evaluation. For long-term follow-up studies and investigation of alternative graft preservation techniques, autotransplantation models are superior to allotransplantation models, as they avoid the confounding bias posed by rejection and immunosuppressive medication.

Porcine models of organ transplantation provide an important platform to study mechanisms of organ preservation. This article describes a heterotopic porcine renal autotransplantation model, which allows investigating new approaches to improve the outcome of transplantation using marginal kidney grafts.

돼지 모델에서 이소성 신장자가 이식 : 단계별 프로토콜

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality ...

A Model to Simulate Clinically Relevant Hypoxia in Humans

In case of apnea, arterial partial pressure of oxygen (pO2) decreases, while partial pressure of carbon dioxide (pCO2) increases. To avoid damage to hypoxia sensitive organs such as the brain, compensatory circulatory mechanisms help to maintain an adequate oxygen supply. This is mainly achieved by increased cerebral blood flow. Intermittent hypoxia is a commonly seen phenomenon in patients with obstructive sleep apnea. Acute airway obstruction can also result in hypoxia and hypercapnia. Until now, no adequate model has been established to simulate these dynamics in humans. Previous investigations focusing on human hypoxia used inhaled hypoxic gas mixtures. However, the resulting hypoxia was combined with hyperventilation and is therefore more representative of high altitude environments than of apnea. Furthermore, the transferability of previously performed animal experiments to humans is limited and the pathophysiological background of apnea induced physiological changes is poorly understood. In this study, healthy human apneic divers were utilized to mimic clinically relevant hypoxia and hypercapnia during apnea. Additionally, pulse-oximetry and Near Infrared Spectroscopy (NIRS) were used to evaluate changes in cerebral and peripheral oxygen saturation before, during, and after apnea.

Hypoxia simulation in humans has usually been performed by inhaling hypoxic gas mixtures. For this study, apneic divers were used to simulate dynamic hypoxia in humans. Additionally, physiological changes in desaturation and re-saturation kinetics were evaluated with non-invasive tools such as Near-Infrared-Spectroscopy (NIRS) and peripheral oxygenation saturation (SpO2).

A 모델은 인간의 임상 관련 저산소증을 시뮬레이션하기

In case of apnea, arterial partial pressure of oxygen (pO2) decreases, while partial pressure of carbon dioxide (pCO2) increases. To avoid damage to ...

A Clinical Trial Assessing the Safety, Efficacy, and Delivery of Olive-Oil-Based Three-Chamber Bags for Parenteral Nutrition

Limited evidence exists to precisely estimate efficacy and safety differences between parenteral nutrition (PN) prepared using olive-oil-based three-chamber bags (3CBs) and soybean-oil-based compounded bags (CoBs) in hospitalized adult patients. We designed a multicenter, randomized, prospective, open-label, noninferiority protocol to compare the efficacy, safety, and distribution of olive-oil-based 3CBs and soybean-oil-based CoB formulations in adult Chinese patients requiring PN during surgical intervention. Subjects were randomized to receive either one of the study treatments using an interactive voice or web-based recognition system in accordance with the randomization code. Randomization was further stratified based on the study site and surgical category. Both treatment groups received similar amounts of calories and protein. In addition, the two study treatments contained a similar composition of the amino-acid component. The only difference between the two PN formulations was the lipid constitution. The duration of administration of study treatments was a minimum of 5 days up to a maximum of 14 days after the surgical procedure. The primary efficacy endpoint was serum prealbumin levels on day 5 of the study. Noninferiority was proved if the anti-log of the lower bound of the 95% confidence interval (CI) of the treatment difference was at least 0.80. Other efficacy measures included treatment preparation time; duration to achieve tolerability of oral nutrition; associated infectious complications; length of hospitalization; and laboratory assessment of markers of nutrition, inflammation, metabolism, and oxidative stress. A total of 458 patients were enrolled in the study. The results showed that olive-oil-based 3CBs were non-inferior to soybean-based CoBs, besides being well tolerated. The infection rate was found to be significantly lower in the olive-oil-based 3CB group. Thus, this study may be used as a reference for future research on lipid emulsion and 3CBs.

Here, we present a protocol for a study comparing the efficacy, safety, and delivery of olive-oil-based 3CB and soybean-oil-based CoB formulations in adults requiring parenteral nutrition. The results revealed that olive-oil-based 3CBs is non-inferior and well tolerated compared to soybean formulations.

Limited evidence exists to precisely estimate efficacy and safety differences between parenteral nutrition (PN) prepared using olive-oil-based ...

Conducting Maximal and Submaximal Endurance Exercise Testing to Measure Physiological and Biological Responses to Acute Exercise in Humans

Regular physical activity has a positive effect on human health, but the mechanisms controlling these effects remain unclear. The physiologic and biologic responses to acute exercise are predominantly influenced by the duration and intensity of the exercise regimen. As exercise is increasingly thought of as a therapeutic treatment and/or diagnostic tool, it is important that standardizable methodologies be utilized to understand the variability and to increase the reproducibility of exercise outputs and measurements of responses to such regimens. To that end, we describe two different cycling exercise regimens that yield different physiologic outputs. In a maximal exercise test, exercise intensity is continually increased with a greater workload resulting in an increasing cardiopulmonary and metabolic response (heart rate, stroke volume, ventilation, oxygen consumption and carbon dioxide production). In contrast, during endurance exercise tests, the demand is increased from that at rest, but is raised to a fixed submaximal exercise intensity resulting in a cardiopulmonary and metabolic response that typically plateaus. Along with the protocols, we provide suggestions on measuring physiologic outputs that include, but are not limited to, heart rate, slow and forced vital capacity, gas exchange metrics, and blood pressure to enable the comparison of exercise outputs between studies. Biospecimens can then be sampled to assess cellular, protein, and/or gene expression responses. Overall, this approach can be easily adapted into both short- and long-term effects of two distinct exercise regimens.

To assess the influence of exercise intensity on physiologic and biologic responses, two different exercise testing protocols were utilized. Methods outlining exercise testing on a cycle ergometer as an incremental maximal oxygen consumption test and endurance, steady state submaximal endurance test are described.

최대한 하 고 Submaximal 지구력 운동 인간에서 급성 운동 생리 및 생물학 응답을 측정 하기 위해 테스트를 실시

Regular physical activity has a positive effect on human health, but the mechanisms controlling these effects remain unclear. The physiologic and biologic ...

A Proinflammatory, Degenerative Organ Culture Model to Simulate Early-Stage Intervertebral Disc Disease.

Symptomatic intervertebral disc (IVD) degeneration (IDD) is a major socioeconomic burden and is characterized by inflammation and tissue degradation. Due to the lack of causative therapies, there is an urgent need for innovative experimental organ culture models to study the mechanisms involved in the progression of the disease, find therapeutic targets, and reduce the need for animal models. We here present a novel, three-dimensional organ culture model protocol mimicking the proinflammatory and catabolic microenvironment, which is present during IDD. 
Initially, bovine caudal IVDs were dissected, cleaned, and cultured in the tissue culture medium. Dynamic physiologic or pathologic loading was applied in a custom-made bioreactor for 2 hours per day. IVDs were assigned to a control group (high glucose medium, physiological loading, phosphate-buffered saline injection) and a pathological group (low glucose medium, pathological loading, tumor necrosis factor-alpha injection) for four days. Gene expression analysis from collected nucleus pulposus cells of the IVDs and enzyme-linked immunosorbent assay of the conditioned organ culture media was performed. 
Our data revealed a higher expression of inflammatory markers and reduced disc heights after loading in the pathological group compared to the control group. This protocol is reliable to simulate IVD inflammation and degeneration and can be further expanded to broaden its application scope.

This protocol presents a novel experimental model of proinflammatory, degenerative bovine organ culture to simulate early-stage intervertebral disc degeneration.

초기 단계 무척추 디스크 질환을 시뮬레이션하는 선동적이고 퇴행성 장기 문화 모델.

Symptomatic intervertebral disc (IVD) degeneration (IDD) is a major socioeconomic burden and is characterized by inflammation and tissue degradation. Due ...

Retinal Pigment Epithelium Transplantation in a Non-human Primate Model for Degenerative Retinal Diseases

Retinal pigment epithelial (RPE) transplantation holds great promise for the treatment of inherited and acquired retinal degenerative diseases. These conditions include retinitis pigmentosa (RP) and advanced forms of age-related macular degeneration (AMD), such as geographic atrophy (GA). Together, these disorders represent a significant proportion of currently untreatable blindness globally. These unmet medical needs have generated heightened academic interest in developing methods of RPE replacement. Among the animal models commonly utilized for preclinical testing of therapeutics, the non-human primate (NHP) is the only animal model that has a macula. As it shares this anatomical similarity with the human eye, the NHP eye is an important and appropriate preclinical animal model for the development of advanced therapy medicinal products (ATMPs) such as RPE cell therapy.
This manuscript describes a method for the submacular transplantation of an RPE monolayer, cultured on a polyethylene terephthalate (PET) cell carrier, underneath the macula onto a surgically created RPE wound in immunosuppressed NHPs. The fovea-the central avascular portion of the macula-is the site of the greatest mechanical weakness during the transplantation. Foveal trauma will occur if the initial subretinal fluid injection generates an excessive force on the retina. Hence, slow injection under perfluorocarbon liquid (PFCL) vitreous tamponade is recommended with a dual-bore subretinal injection cannula at low intraocular pressure (IOP) settings to create a retinal bleb. 
Pretreatment with an intravitreal plasminogen injection to release parafoveal RPE-photoreceptor adhesions is also advised. These combined strategies can reduce the likelihood of foveal tears when compared to conventional techniques. The NHP is a key animal model in the preclinical phase of RPE cell therapy development. This protocol addresses the technical challenges associated with the delivery of RPE cellular therapy in the NHP eye.

The non-human primate (NHP) is an ideal model for studying human retinal cellular therapeutics due to the anatomical and genetic similarities. This manuscript describes a method for submacular transplantation of retinal pigment epithelial cells in the NHP eye and strategies to prevent intraoperative complications associated with macular manipulation.

퇴행성 망막 질병을 위한 비인간 영장류 모형에 있는 망막 안료 상피 이식

Retinal pigment epithelial (RPE) transplantation holds great promise for the treatment of inherited and acquired retinal degenerative diseases. These ...

Quantification of Optic Nerve Cross Sectional Area on MRI: A Novel Protocol using Fiji Software

Optic nerve assessment is an important aspect of glaucoma diagnosis and follow-up. This project describes a protocol for a unified methodology of optic nerve cross-sectional assessment and quantification using 3 T MRI for image acquisition and ImageJ's Fiji software for image processing quantification. Image acquisition was performed using 3 T MRI, with proper instructions for the patient to ensure straight fixation during imaging. A T2-weighted fat suppressed sequence was used. A coronal cut taken 3 mm behind the globe and perpendicular to the optic nerve axis should be uploaded to the software. Using the threshold function, the white matter area of the optic nerve is selected and quantified, thus, eliminating inter-individual measurement bias. We also described the normal limits for the optic nerve cross-sectional area according to age, based on previously published literature. We used the described protocol to assess optic nerve of a suspected glaucoma patient. The optic nerve cross-sectional area was found to be within the normal limits, a finding further confirmed via optical coherence tomography of the optic nerve.

We provided a detailed protocol for a standardized method of optic nerve assessment and quantification using MRI, utilizing a widely available imaging sequence, and open access software for image analysis. Following this standardized protocol would provide meaningful data for comparison between different patients and different studies.

MRI에 시신경 단면 영역의 정량화: 피지 소프트웨어를 사용 하 여 새로운 프로토콜

Optic nerve assessment is an important aspect of glaucoma diagnosis and follow-up. This project describes a protocol for a unified methodology of optic ...

A Real-World High-Intensity Interval Training Protocol for Cardiorespiratory Fitness Improvement

High-Intensity Interval Training (HIIT) has emerged as an interesting time-efficient approach to increase exercise adherence and improve health. However, few studies have tested the efficiency of HIIT protocols in a &#34;real world&#34; setting, e.g., HIIT protocols designed for outdoor spaces without specialized equipment. This study presents a &#34;real world&#34; training protocol, named &#34;beep training&#34;, and compares the efficiency of a HIIT regiment versus a traditional long-duration Moderate-Intensity Continuous Training (MICT) regiment using this beep training protocol on VO2 max of overweight untrained men. Twenty-two subjects performed outdoor running with MICT (n = 11) or HIIT (n = 11). Cardiorespiratory fitness was assessed before and after training protocols using a metabolic analyzer. Both training protocols were performed 3 days a week for 8 weeks using the Beep Test results. The MICT group performed the exercise program at 60%-75% of the maximum speed of the 20 m shuttle test (Vmax) and with a progression of the distance of 3,500-5,000 m. The HIIT group performed the interval exercise with 7-10 bouts of 200 m at 85%-100% of the maximum speed of the 20 m shuttle test (Vmax), interspersed with 1 min of passive recovery. Although the HIIT group presented a significantly lower training volume than the MICT group (p &#60; 0.05) after 8 weeks of beep training, HIIT was superior to MICT in improving VO2 max (MICT: ~4.1%; HIIT: ~7.3%; p &#60; 0.05). The &#34;real world&#34; HIIT regiment based on beep training protocol is a time-efficient, low-cost, and easy-to-implement protocol for overweight untrained men.

This study presents a low-cost and easy-to-implement "real world" high-intensity interval training (HIIT) protocol for scientific research and discusses its efficiency for cardiorespiratory fitness.

심장 호흡기 피트니스 개선을위한 실제 고강도 간격 훈련 프로토콜

High-Intensity Interval Training (HIIT) has emerged as an interesting time-efficient approach to increase exercise adherence and improve health. However, ...