Name: intrasplenic transplantation of hepatocytes after partial hepatectomy in nod.scid mice
Uploaded: 2018-02-10T13:00:00
Description: Watch this Scientific Journal Video about Intrasplenic Transplantation of Hepatocytes After Partial Hepatectomy in NOD.SCID Mice at JoVE.com

이 작품은 학과 생명 공학, 면역학, 뉴델리의 국립 연구소에 인도 정부에서에서 받은 코어 그랜트에 의해 지원 되었다. 박사 Bhattacharjee의 현재 주소는 사단의 위장, 간장 학과 및 영양, 아동 병원 로스 앤젤레스.

이 프로토콜에서 제시 하는 절차는 면역학, 뉴델리의 국립 연구소의 기관 동물 윤리 위원회에 의해 승인 되었습니다. 승인의 직렬 참조 번호는 IAEC #319/13.
참고: 일반 외과 절차20 및 설치류 수술21에 대 한 특정 프로토콜에 우수한 자원 있습니다. 그는 처음으로 동물 수술을 하 고,이 좋습니다 광범위 하 게 연습을 인형에 수술 동물에 ?...

<ol>
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부분 hepatectomy 조사 간 재생에 대 한 설립된 기술 이며 과도 한 hepatectomy AHF 모델을 모방 하기 위해 보고. AHF의 동물 모델 중 특히 쥐, 설치류는 가장 연구 모델입니다. 최대 70 %hepatectomy 쥐에서 간 상해 모델을 얻기 위해 좋은 생존 율25,26보고 되었습니다. 그러나 누드 및 다른 immunodeficient 마우스, 70 %hepatectomy 치명적으로 알려졌다 고 동물27</su...

현재, hepatocyte 이식 데 심한 간 장애 환자 치료에 대 한 전체 장기 이식의 대 안으로 제시 된다. 그것은 전체 장기 이식1환자 다리 수 믿 었 다. Allogenic hepatocytes2뿐만 아니라 xenogenic hepatocytes3 및 줄기 세포4 에서 파생 된 hepatocytes는 또한 되 고 조사 동물 모델에서. 이 컨텍스트, 추적 및 engraftment 받는 사람에서 이식된 세포의 잠재력 급성 간장 장애 (AHF)에 따라 셀 치료에 대 한 중요 한 기준입니다.
조사 hepatocytes 또는 hepatocyte 같은 셀5의 이식, AHF 수술6 또는 약리학7 절차, 뒤에 세포 이식 동물 모델에서 만들어집니다. 약리학 적인 시 약, 많은 hepatotoxins d galactosamine8, 아 세트 아미노 펜9, 사염화 탄소10, thioacetamide11, Concanavalin A12, lipopolysaccharide13 AHF 동물 모델을 , 등., 사용 되었습니다. 이 목록에서 모든 시 약 AHF, 독특한 기능 집합이 생성 하지만 불행히도 아무 단일 시 약 인간의 AHF 모방. 또한, hepatotoxins에 의해 유도 된 AHF는 시간이 오래 걸립니다, 만성 스트레스는 동물을 박 았, 그리고 재현성 결과 얻기 어렵다.
다른 한편으로, 부분 hepatectomy (PHx)의 수술은 기술 종속, 고 재현성 결과 필요한 능력 개발 후 얻을 쉽습니다. 혼자 외 과적으로 AHF을 유도, 간 70% 이상의 절제가 필요; 그러나, 70 %hepatectomy 여전히 engraftment의 확산을 공부 하 고 이용 될 수 있다 보다는 더 적은 셀 간 손상14동안 그들의 치료 용량을 분석 하기 위한 간 이식. Hepatocytes의 이식 복15, 꼬리 정 맥16,17간 정 맥 또는 비장18통해 수행된 게시물 hepatectomy 되었습니다. 현재, 간 정 맥 주입 및 hepatocytes의 intrasplenic 이식으로 재현 하기 쉬운 기본 절차입니다.
이 문서에 우리는 30% 부분 hepatectomy 끄 덕에 대 한 절차를 설명 했습니다. SCID (끄 덕 임. CB17-Prkdcscid/J) 마우스의 왼쪽된 엽 excised입니다. 그것은 이식의 0.2 백만 GFP 표현 마우스 (C57BL/6-Tg (UBC-GFP) 30Scha/J) hepatocytes 인간의 기원 NeoHep19 비장에 옵니다. 이 절차에에서 이식된 세포의 engraftment 이끌어 낸다. 이 절차는 최소한 침략 적 및 최소한 고통 스러운 기술.

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			<td>Local Procurement</td>
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			<td>Mercian ;&#160; England&#160;</td>
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			<td>Dispo Van; India</td>
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			<td height="21" style="height:21px;">1 ml syringe with 26 G * 1/2 needle</td>
			<td>BD ; US&#160;</td>
			<td>REF 303060</td>
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			<td style="width:255px;">Mighty ; India</td>
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			<td style="width:255px;">Ethicon, Johnson &#38; Johnson, India</td>
			<td style="width:119px;">NW 5047</td>
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			<td>Sutures India Pvt. Ltd; India</td>
			<td>SN 5048</td>
			<td>Sterilised Surgical Needled Suture Catgut Chromic</td>
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			<td style="width:255px;">3M India ; India</td>
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		<tr height="42">
			<td height="42" style="height:42px;width:411px;">Automated analyser</td>
			<td>Tulip, Alto Santracruz, India</td>
			<td style="width:119px;">Screen Maaster 3000</td>
			<td>Biochemical analyser for liver functional test</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Flow Cytometer</td>
			<td style="width:255px;">BD ; US&#160;</td>
			<td style="width:119px;">BD FACSverse</td>
			<td>Assesment of presence of cells post transplantation</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Veet hair removal cream&#160;</td>
			<td>Reckitt Benckiser , India</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">FORANE</td>
			<td style="width:255px;">Abbott ; US</td>
			<td style="width:119px;"></td>
			<td>isoflurane USP 99.9%&#160;</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Taxim</td>
			<td style="width:255px;">AlKem ; India</td>
			<td style="width:119px;"></td>
			<td>cefotaxime sodium injection</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Povidone-Iodine solution&#160;</td>
			<td style="width:255px;">Win-Medicare;&#160; India</td>
			<td style="width:119px;"></td>
			<td>Betadine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Paraformaldehyde</td>
			<td style="width:255px;">Himedia; India</td>
			<td style="width:119px;">GRM 3660</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:411px;">Iscove&#39;s Modified Dulbecco&#39;s Medium (IMDM)</td>
			<td style="width:255px;">Life technologies, Thermo Fisher scientific ; US</td>
			<td style="width:119px;">12200-036</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Sucrose</td>
			<td style="width:255px;">Sigma ; US</td>
			<td style="width:119px;">S0389</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Tissue-Tek</td>
			<td style="width:255px;">Sakura; US</td>
			<td>25608-930</td>
			<td>O.C.T compound</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:411px;">DAPI</td>
			<td style="width:255px;">Himedia; India</td>
			<td style="width:119px;">MB 097</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">anti-Albumin goat Polyclonal</td>
			<td style="width:255px;">Thermo Scientific,Pierce, US</td>
			<td style="width:119px;">PA126081</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">anti-connexin 32/GJB1 Polyclonal</td>
			<td style="width:255px;">abcam, UK</td>
			<td style="width:119px;">ab64609-500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">antiGFP rabbit polyclonal&#160;</td>
			<td style="width:255px;">Santa Cruz biotechnology; US</td>
			<td style="width:119px;">SC 8334</td>
			<td></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:411px;">Alexa Fluor 594 donkey anti-goat</td>
			<td style="width:255px;">&#160;Molecular Probes , Thermo Fisher Scientific ;&#160; US</td>
			<td style="width:119px;">A11058</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:411px;">Alexa Fluor 488 donkey anti-sheep</td>
			<td style="width:255px;">&#160;Molecular Probes , Thermo Fisher Scientific ;&#160; US</td>
			<td style="width:119px;">A11015</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:411px;">Alexa Fluor 594 chicken anti rabbit</td>
			<td style="width:255px;">&#160;Molecular Probes , Thermo Fisher Scientific ;&#160; US</td>
			<td style="width:119px;">A21442</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:411px;">Goat anti rabbit IgG HRP</td>
			<td style="width:255px;">Invitrogen, Thermo Fisher Scientific; US&#160;</td>
			<td>65-6120</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">anti-Ki67 antibody</td>
			<td style="width:255px;">abcam, UK</td>
			<td style="width:119px;">ab15580</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Antigen Unmasking Solution, Citric acid base</td>
			<td style="width:255px;">Vector laboratories, US</td>
			<td style="width:119px;">H-3300</td>
			<td></td>
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		<tr height="42">
			<td height="42" style="height:42px;">ProLong Diamond antifade mountant</td>
			<td style="width:255px;">Life technologies, Thermo Fisher scientific ; US</td>
			<td style="width:119px;">P36966</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">SGOT (ASAT) KIT</td>
			<td style="width:255px;">Coral Clinical System, India</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">SGPT (ALAT) KIT</td>
			<td style="width:255px;">Coral Clinical System, India</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Alkaline Phosphatase Kit (DEA)</td>
			<td style="width:255px;">Coral Clinical System, India</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Hematoxylin Solution, Mayer&#39;s</td>
			<td style="width:255px;">Sigma ; US</td>
			<td style="width:119px;">MHS16</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Eosin Y solution, alcoholic</td>
			<td style="width:255px;">Sigma ; US</td>
			<td style="width:119px;">HT110132</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">DPX Mountant&#160;</td>
			<td style="width:255px;">Sigma ; US</td>
			<td align="right" style="width:119px;">6522</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">Melonex (Pain Killer)</td>
			<td style="width:255px;">Intas Pharmaceuticals Ltd; India</td>
			<td style="width:119px;"></td>
			<td>Meloxicam injection&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:411px;">DAB enhanced liquid substrate system tetrahydrochloride</td>
			<td style="width:255px;">Sigma ; US</td>
			<td style="width:119px;">D3939</td>
			<td></td>
		</tr>
	</tbody>
</table>

intrasplenic transplantation of hepatocytes after partial hepatectomy in nod.scid mice

부분 hepatectomy 간 재생 및 다양 한 병 적인 조건에서 세포 기반 치료제의 효과 공부 하는 다양 하 고 재현할 수 방법입니다. 또한 부분 hepatectomy에 간 쪽으로 가속 neovascularization 및 셀에 의해 증가 engraftment와 이식된 세포의 증식 촉진 한다. 여기, 우리 immunodeficient 끄 덕을 결합 하 여 비만 아닌 당뇨병/심한의 비장에 30 %hepatectomy 이식 세포의 수행에 대 한 간단한 프로토콜을 설명 합니다. SCID (끄 덕 임. CB17-Prkdcscid/J) 마우스.
이 절차에서는 두 개의 작은 절 개 만들어집니다. 첫 번째 절 개 노출 하 고, 간 좌 엽을 resect 하는 것입니다 그리고 또 다른 작은 절 개 비장 세포의 intrasplenic 이식에 대 한 폭로 이루어집니다. 이 절차는 어떤 전문된 외과 기술이 필요 하지 않습니다 그리고 그것은 5-7 분 더 적은 스트레스와 고통, 빠른 복구, 및 더 나은 생존에에서 완료 될 수 있다. 우리 마우스 (유전자 변형 C57BL/6-Tg (UBC-GFP) 30Scha/J), 뿐만 아니라 부분적으로 hepatectomized 끄 덕에에서 인간의 기원 (NeoHep)의 세포 처럼 hepatocyte 표현 녹색 형광 단백질 (GFP)에서 절연 hepatocytes의 이식 설명 했다. SCID 쥐입니다.

30% 부분 hepatectomy 후 hepatocyte 확산: 간 30 %hepatectomy 후 남은 hepatocytes의 확산 immunohistochemical (IHC) 세포 확산 표식 기-67에 대 한 얼룩에 의해 시험 되었다. 하루 게시물 hepatectomy 쥐 했다 안락사, 나머지 간 엽 excised 했다, 및 파라핀 섹션 가져온. 섹션은 뒤에 양 고추냉이 과산화 효소 (HRP) 활용 된 이차 항 체와 라벨 기-67 항 체와 스테인드 했다. 디 아미노 Benzidine (한 덩어리)으로 사용 되었?...

Watch this Scientific Journal Video about Intrasplenic Transplantation of Hepatocytes After Partial Hepatectomy in NOD.SCID Mice at JoVE.com

Intrasplenic Transplantation of Hepatocytes After Partial Hepatectomy in NOD.SCID Mice

우리는 끄 덕의 부분 hepatectomy (PHx) 및 비장을 통해 세포 이식 수행을 위한 프로토콜을 설명 했습니다. SCID (끄 덕 임. CB17-Prkdcscid/J) 쥐. 이 프로토콜에서 절 개 노출 하 고 셀의 intrasplenic 이식에 대 한 또 다른 절 개에 의해 따라 간 좌 엽 resect 이루어집니다.

끄 덕 임에서 부분 Hepatectomy 후 Hepatocytes의 intrasplenic 이식 SCID 마우스

Partial hepatectomy is a versatile and reproducible method to study liver regeneration and the effect of cell based therapeutics in various pathological ...

Research

JoVE Journal

Medicine

Surgical Procedures for a Rat Model of Partial Orthotopic Liver Transplantation with Hepatic Arterial Reconstruction

간 간선 재건과 부분 Orthotopic 간 이식의 쥐 모델에 대한 외과 절차

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as ...

연구

의학

Heterotopic Cervical Heart Transplantation in Mice

마우스의 이소성 자궁 심장 이식

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies ...

Visualization of Vascular and Parenchymal Regeneration after 70% Partial Hepatectomy in Normal Mice

일반 마우스의 70 % 부분 간 절제술 후 혈관 및 실질 재생의 시각화

A modified silicone injection procedure was used for visualization of the hepatic vascular tree. This procedure consisted of in-vivo injection of the ...

Re-Arterialized Rat Partial Liver Transplantation with an in vivo Vessel-Oriented 70% Hepatectomy

Re-Arterialized Rat Partial Liver Transplantation with an in vivo Vessel-Oriented 70% Hepatectomy

다시는 vivo에서 선박 지향 70% 쥐 부분 간 이식 Arterialized Hepatectomy

Split liver transplantation and living liver donor liver transplantation were developed in the clinic to utilize liver organs in a more efficient manner. ...

Transplantation of Adipose Tissue-Derived Stem Cell Sheet to Reduce Leakage After Partial Colectomy in A Rat Model

쥐 모델에 부분 Colectomy 후 누설을 줄이기 위해 지방 조직 유래 줄기 세포의 이식

Anastomotic leakage is a disastrous complication after colorectal surgery. Although current methods for leakage prevention have different levels of ...

Lentiviral Vector-mediated Gene Therapy of Hepatocytes Ex Vivo for Autologous Transplantation in Swine

Lentiviral Vector-mediated Gene Therapy of Hepatocytes Ex Vivo for Autologous Transplantation in Swine

돼지에서 자가 이식 Hepatocytes 전 비보 의 lentiviral 벡터 중재 된 유전자 치료

Gene therapy is an ideal choice to cure many inborn errors of metabolism of the liver. Ex-vivo, lentiviral vectors have been used successfully in the ...

Generation of a Rat Model of Acute Liver Failure by Combining 70% Partial Hepatectomy and Acetaminophen

70% 부분 간 절제술과 아세트아미노펜을 결합하여 급성 간 기능 부전의 쥐 모델 생성

Acute liver failure (ALF) is a clinical condition caused by various etiologies resulting in the loss of metabolic, biochemical, synthesizing, and ...

Reduced Complications after Arterial Reconnection in a Rat Model of Orthotopic Liver Transplantation

직교 간 이식의 쥐 모형에 있는 동맥 재연결 후에 감소된 합병증

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation ...

Intramyocardial Transplantation of MSC-Loading Injectable Hydrogels after Myocardial Infarction in a Murine Model

Murine 모델에서 심근 경색 후 MSC 로딩 주사용 하이드로겔의 자궁 내 이식

One of the major issues facing current cardiac stem cell therapies for preventing postinfarct heart failure is the low retention and survival rates of ...