Name: two methods for establishing primary human endometrial stromal cells from hysterectomy specimens
Uploaded: 2014-05-23T14:00:00
Description: Watch this Scientific Journal Video about Two Methods for Establishing Primary Human Endometrial Stromal Cells from Hysterectomy Specimens at JoVE.com

우리는 그들의 이미징 및 현미경 장비의 사용을 위해 박사 타오 댕와 그녀의 실험실의 구성원의 공동 노력을 주셔서 감사합니다. 우리는 또한 자궁 조직으로 우리를 제공하기위한 Biorepository 및 조직 연구 시설 (BTRF) 코어, 제프 하퍼, 버지니아 대학에서 주민 감사합니다. 우리는 도식 개요와 도움 카롤 Szlachta 감사합니다.

이 원고에 사용 된 자궁 절제술 표본 IRB-HSR # 14424 번호가 대학 IRB 승인 윤리 프로토콜과 일치에 수집되었다. 임상 자료 1. 샘플 수집 <ol><li> 시작하기 전에 정부와 기관 기반의 윤리 지침 및 승인 문서를 얻습니다. </li><li> 무균 조건의 모든 단계를 실시한다. </li><li> 샘플이 바로 문화에서 처리 할 수​​없는 경우 4 ° C에서 50 ML 튜브 미디어 (RPMI 또는 DMEM / 높은 포도당)의 환자...

<ol>
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M., Hampton, A. L., Riley, S. C., Findlay, J. K., Salamonsen, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Production+and+characterization+of+endothelin+released+by+human+endometrial+epithelial+cells+in+culture.">Production and characterization of endothelin released by human endometrial epithelial cells in culture.</a> J Clin Endocrinol Metab. 79, 1625-1631 (1994).</li><li>Siegfried, J. M., Nelson, K. G., Martin, J. L., Kaufman, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Histochemical+identification+of+cultured+cells+from+human+endometrium.">Histochemical identification of cultured cells from human endometrium.</a> In Vitro. 20, 25-32 (1984).</li><li>Rawdanowicz, T. J., Hampton, A. L., Nagase, H., Woolley, D. E., Salamonsen, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Matrix+metalloproteinase+production+by+cultured+human+endometrial+stromal+cells:+identification+of+interstitial+collagenase,+gelatinase-A,+gelatinase-B,+and+stromelysin-1+and+their+differential+regulation+by+interleukin-1+alpha+and+tumor+necrosis+factor-alpha.">Matrix metalloproteinase production by cultured human endometrial stromal cells: identification of interstitial collagenase, gelatinase-A, gelatinase-B, and stromelysin-1 and their differential regulation by interleukin-1 alpha and tumor necrosis factor-alpha.</a> J Clin Endocrinol Metab. 79, 530-536 (1994).</li><li>Dimitriadis, E., Robb, L., Salamonsen, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interleukin+11+advances+progesterone-induced+decidualization+of+human+endometrial+stromal+cells.">Interleukin 11 advances progesterone-induced decidualization of human endometrial stromal cells.</a> Mol Hum Reprod. 8, 636-643 (2002).</li><li>Sakai, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Involvement+of+histone+acetylation+in+ovarian+steroid-induced+decidualization+of+human+endometrial+stromal+cells.">Involvement of histone acetylation in ovarian steroid-induced decidualization of human endometrial stromal cells.</a> J Biol Chem. 278, 16675-16682 (2003).</li><li>Logan, P. C., Ponnampalam, A. P., Steiner, M., Mitchell, M. 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다른 그룹은 콜라게나 4,12,13,15-18를 활용 대부분의 자궁 내막 기질 문화의 제조 방법을 설명하고 적응. 이 논문에서, 우리는 경제적 인 이유와 트립신 및 / 또는 면도날의 편리한 이용 가능 여부 연구실로 활용 둘 중 두 가지 간단한 기본 자궁 내막 배양 방법에 대한 방법론과 증거를 제공했다. 우리의 두 가지 방법을 비교했을 때, 모두가 성공적으로 실행 가능한 차 문...

인간의 자궁 체부는 세 개의 층, perimetrium (또는 장막), 자궁 근층 및 자궁 내막으로 구성되어 있습니다. 이들 층의 각각을 구별하는 것은 내막 세포주를 확립하는 중요한 단계이다. perimetrium은 자궁의 가장 바깥 층과 얇은 장액 세포로 구성. 근육층은 두꺼운 중간 자궁의 층과 평활근 세포로 구성된다. 자궁 내막은 자궁의 내부 층으로 식별하고 상피 및 간질 세포 집단을 포함한다. 자궁 내막은 또한 그 줄기 세포 인구는 약 28 일마다 1 functionalis 층을 다시 채울 가설 basalis 층으로 구분된다. 인간의 자궁 내막의 functionalis 층 호르몬 순환에 대한 응답으로 중요한 생화학 적 및 형태 학적 변화를 겪는다. 이 호르몬은 뇌하수체와 난소에서 파생됩니다. 생식주기의 조정 생산 및 호르몬 결과의 릴리스. 생식주기는 잠재적 인 배아 주입 이벤트에 대한 내막을 준비하도록 설계되었습니다. 인간의 생식주기는 &quot;월경주기&quot;라고하며 다음과 같은 세 가지 단계로 구분 - 증식, 분비, 월경. 분비 단계가 functionalis 성숙에 의해 표시되는 반면 증식 단계는 functionalis 자궁 내막 층의 확산을 포함한다. 특히, 세포 변화, 분비물, 그리고 세포 분화는 잠재적 인 주입 신호. 주입이 분비 단계의 끝 이전에 발생하지 않는 경우, functionalis의 자궁 내막 층은 생리 단계 흘렸다. 생리 및 functionalis 층의 흘리기를 트리거하는 이벤트의 중요성은 여전히​​ 논의되고있다. 인간에서는, 생리 알려진 특정 미드 분비 기 감별 이벤트의 결과라고 제기되었다&quot;자연 탈락&quot;2 등. 이 논문에서는, 양쪽 자궁 내막 세포 분리 방법에 대한 상세한 방법을 제공하며, 이러한 방법의 효능을 입증하기 위하여 면역 형광 및 디지털 이미지의 조합을 사용한다. 또, 일반적으로 적용 자궁 내막 세포 격리을 확인하기 위하여 자발 탈락의 시험 관내 모델에서 사용.

<table border="0" cellpadding="0" cellspacing="0" width="713">
	<tbody>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">0.25 Trypsin or 0.05% Trypsin&#160;</td>
			<td dir="LTR" style="width:201px;">Hyclone&#160;</td>
			<td dir="LTR" style="width:216px;">SH3023602 or SH30004202&#160;&#160;</td>
			<td style="width:63px;"></td>
		</tr>
		<tr height="27">
			<td dir="LTR" height="27" style="height:27px;width:235px;">1.7 micro Centrifuge Tube&#160;&#160;</td>
			<td dir="LTR" style="width:201px;">Genesee Scientific&#160;</td>
			<td dir="LTR" style="width:216px;">22-272A&#160;</td>
			<td></td>
		</tr>
		<tr height="25">
			<td dir="LTR" height="25" style="height:25px;width:235px;">1&#181;l,20&#181;l, 200ml and 1000&#181;l Pipette&#160;&#160;</td>
			<td dir="LTR" style="width:201px;">Genesee Scientific&#160;</td>
			<td dir="LTR" style="width:216px;">24-401,24-402, 24-412, 24-430&#160;</td>
			<td></td>
		</tr>
		<tr height="22">
			<td dir="LTR" height="22" style="height:22px;width:235px;">15ml Conical Tube&#160;</td>
			<td dir="LTR" style="width:201px;">Hyclone&#160;</td>
			<td dir="LTR" style="width:216px;">339650</td>
			<td></td>
		</tr>
		<tr height="23">
			<td dir="LTR" height="23" style="height:23px;width:235px;">50ml Conical Tube&#160;</td>
			<td dir="LTR" style="width:201px;">Hyclone&#160;</td>
			<td dir="LTR" style="width:216px;">339652</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">6cm Cell Culture Dish&#160;</td>
			<td dir="LTR" style="width:201px;">Thermo scientific&#160;&#160;</td>
			<td dir="LTR" style="width:216px;">12-556-002&#160;</td>
			<td></td>
		</tr>
		<tr height="22">
			<td dir="LTR" height="22" style="height:22px;width:235px;">8 well Chambers&#160;</td>
			<td dir="LTR" style="width:201px;">Thermo Scientific&#160;</td>
			<td dir="LTR" style="width:216px;">AB-4162&#160;</td>
			<td></td>
		</tr>
		<tr height="23">
			<td dir="LTR" height="23" style="height:23px;width:235px;">Acetate&#160;</td>
			<td dir="LTR" style="width:201px;">Fisher scientific&#160;</td>
			<td dir="LTR" style="width:216px;">C4-100&#160;</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">AMV RT Enzyme/Buffer&#160;</td>
			<td dir="LTR" style="width:201px;">Bio Labs&#160;</td>
			<td dir="LTR" style="width:216px;">M077L&#160;</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">Bovine Serum Albumin (BSA)&#160;</td>
			<td dir="LTR" style="width:201px;">Fisher Scientific&#160;</td>
			<td dir="LTR" style="width:216px;">BP-1605-100&#160;</td>
			<td></td>
		</tr>
		<tr height="23">
			<td dir="LTR" height="23" style="height:23px;width:235px;">Buffered Zinc Formalin&#160;</td>
			<td dir="LTR" style="width:201px;">Thermo&#160;</td>
			<td dir="LTR" style="width:216px;">59201ZF&#160;</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">Charcoal strip FBS&#160;</td>
			<td dir="LTR" style="width:201px;">Fisher&#160;</td>
			<td dir="LTR" style="width:216px;">NC9019735&#160;&#160;</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">Chloroform&#160;</td>
			<td dir="LTR" style="width:201px;">Fisher Scientific&#160;</td>
			<td dir="LTR" style="width:216px;">BP1145-1</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">Cover slip&#160;</td>
			<td dir="LTR" style="width:201px;">Fisher Brand&#160;&#160;</td>
			<td dir="LTR" style="width:216px;">12-544D&#160;</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">Cyclic AMP (cAMP)&#160;</td>
			<td dir="LTR" style="width:201px;">Sigma&#160;</td>
			<td dir="LTR" style="width:216px;">B7880</td>
			<td></td>
		</tr>
		<tr height="24">
			<td dir="LTR" height="24" style="height:24px;width:235px;">DMEM/High Glucose&#160;</td>
			<td dir="LTR" style="width:201px;">Hyclone&#160;</td>
			<td dir="LTR" style="width:216px;">SH30243FS&#160;</td>
			<td></td>
		</tr>
		<tr height="25">
			<td dir="LTR" height="25" style="height:25px;width:235px;">dNTP&#160;</td>
			<td dir="LTR" style="width:201px;">Bioline&#160;</td>
			<td dir="LTR" style="width:216px;">BIO-39025&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td dir="LTR" height="21" style="height:21px;width:235px;">Donkey Anti Goat -TRITC&#160;</td>
			<td dir="LTR" style="width:201px;">Santa Cruz&#160;</td>
			<td dir="LTR" style="width:216px;">SC-3855&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td dir="LTR" height="21" style="height:21px;width:235px;">Donkey Serum&#160;</td>
			<td dir="LTR" style="width:201px;">Jackson&#8217;s lab&#160;</td>
			<td dir="LTR" style="width:216px;">&#160;017-000-002&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td dir="LTR" height="21" style="height:21px;width:235px;">E Cadherin Antibody&#160;&#160;</td>
			<td dir="LTR" style="width:201px;">Epitomics&#160;</td>
			<td dir="LTR" style="width:216px;">1702-1</td>
			<td></td>
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		<tr height="21">
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two methods for establishing primary human endometrial stromal cells from hysterectomy specimens

많은 노력이 체외 세포 배양 시스템의 구축에 전념하고 있습니다. 이러한 시스템은 생체 내 프로세스의 광대 한 번호를 모델로 설계되어 있습니다. 인간의 자궁 내막 샘플에서 발생하는 세포 배양 시스템도 예외는 아니다. 응용 프로그램은 정상적인 생리주기 프로세스에서 같은 부인과 암, 감염성 질환, 생식 결함으로 자궁 내막 병변의 범위. 여기, 우리는 수술 절제 자궁 적출술 검체에서 기본 자궁 내막 기질 세포를 확립하기위한 두 가지 방법을 제공합니다. 첫 번째 방법은 &quot;스크래핑 방법&quot;이라하고 두 번째 방법은 칭했다 반면 수술 또는 면도날을 사용하여 기계적 스크래핑 통합되는 &quot;트립신 방법.&quot;이 후자의 방법은 세포의 분리 및 일차 홍보 트립신 효소 활성을 사용 세포 가지. 우리는 디지털 이미지와 현미경을 통해 단계별 방법을 설명한다. 우리는 또한 제공하기양적 실시간 중합 효소 연쇄 반응 (qPCR에) 및 면역 (IF)를 통해 자궁 내막 기질 세포 라인의 유효성을 검사하는 전자의 예입니다.

프로토콜 섹션에 강조으로, 인간의 조직을 처리하고 준비 할 때 모든 기관 정부에서 방법, 윤리적 지침을 수행해야합니다. &quot;긁는 방법&quot;의 일반적인 워크 플로의 그림 (그림 1A) 및 기본 자궁 내막 문화를 설정하는 데 사용 &quot;트립신 방법&quot;(그림 1B)은이 원고에 포함되어 있습니다. 이들 방법은 프로토콜 섹션에서 ?...

Watch this Scientific Journal Video about Two Methods for Establishing Primary Human Endometrial Stromal Cells from Hysterectomy Specimens at JoVE.com

Two Methods for Establishing Primary Human Endometrial Stromal Cells from Hysterectomy Specimens

자궁 적출술 검체에서 기본 자궁 내막 기질 세포 배양 시스템을 구축하는 것은 이전의 연구 목표의 광대 한 배열을 추구하는 귀중한 생물 기술과 중요한 단계입니다. 여기서 우리는 인간의 환자의 수술 적 절제술 자궁 내막 조직에서 기질 문화를 설정하는 데 사용되는 두 가지 방법을 설명합니다.

자궁 적출술 검체에서 차 인간의 자궁 내막 기질 세포를 구축하는 두 가지 방법

Many efforts have been devoted to establish in vitro cell culture systems. These systems are designed to model a vast number of in vivo processes. Cell ...

Research

JoVE Journal

Medicine

Experimental Generation of Carcinoma-Associated Fibroblasts (CAFs) from Human Mammary Fibroblasts

Experimental Generation of Carcinoma-Associated Fibroblasts CAFs from Human Mammary Fibroblasts

인간 Mammary 섬유아 세포에서 암종 - 관련 섬유아 세포의 실험 세대 (CAFs)

Carcinomas are complex tissues comprised of neoplastic cells and a non-cancerous compartment referred to as the 'stroma'. The stroma consists of ...

연구

의학

Skin Punch Biopsy Explant Culture for Derivation of Primary Human Fibroblasts

인간의 기본 섬유 아세포의 유도를위한 피부 펀치 생검 절편 문화

Tissues and cell lines derived from an individual with disease are ideal sources to study disease-related cellular phenotypes. Patient-derived fibroblasts ...

Repair of a Critical-sized Calvarial Defect Model Using Adipose-derived Stromal Cells Harvested from Lipoaspirate

Lipoaspirate에서 수확 지방 - 파생 Stromal 셀을 사용하여 임계 크기 Calvarial 결함 모델의 수리

Craniofacial skeletal repair and regeneration offers the promise of de novo tissue formation through a cell-based approach utilizing stem cells. ...

Isolation, Characterization and Comparative Differentiation of Human Dental Pulp Stem Cells Derived from Permanent Teeth by Using Two Different Methods

인간 치과 펄프의 줄기 세포의 분리, 특성화 및 비교 차별화 두 가지 방법을 사용하여 영구 치아에서 파생

Developing wisdom teeth are easy-accessible source of stem cells during the adulthood which could be obtained by routine orthodontic treatments. Human ...

Improved Protocol For Laser Microdissection Of Human Pancreatic Islets From Surgical Specimens

수술 표본에서 인간 췌장 섬의 레이저 Microdissection을위한 개선 된 프로토콜

Laser microdissection (LMD) is a technique that allows the recovery of selected cells and tissues from minute amounts of parenchyma 1,2. The dissected ...

Transcriptomic Analysis of Human Retinal Surgical Specimens Using jouRNAl

저널을 사용하여 인간의 망막 수술 표본의 Transcriptomic 분석

Retinal detachment (RD) describes a separation of the neurosensory retina  from the retinal pigmented epithelium (RPE). The RPE is essential for normal  ...

Method for Obtaining Primary Ovarian Cancer Cells From Solid Specimens

고체 시편에서 기본 난소 암 세포를 얻는 방법

Reliable tools for investigating ovarian cancer initiation and progression are urgently needed. While the use of ovarian cancer cell lines remains a ...

Enrichment for Chemoresistant Ovarian Cancer Stem Cells from Human Cell Lines

인간 세포주에서 Chemoresistant 난소 암 줄기 세포에 대한 심화 학습

Cancer stem cells (CSCs) are defined as a subset of slow cycling and undifferentiated cells that divide asymmetrically to generate highly proliferative, ...

Studying Pancreatic Cancer Stem Cell Characteristics for Developing New Treatment Strategies

췌장암 새로운 치료 전략을 개발하기위한 전지 특성 줄기 공부

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor ...

Isolation and Culture of Primary Endothelial Cells from Canine Arteries and Veins

개 동맥과 정맥에서 차 내피 세포의 분리 및 문화

Cardiovascular disease is studied in both human and veterinary medicine. Endothelial cells have been used extensively as an in vitro model to study ...