저자는 비판적으로 원고를 검토하고 증거 편집 재이든 스미스 감사합니다. 이 작품은 국립 다발성 경화증 학회 (NMSS, 부분적인 보조금 RG-4001-A1)에서지지를 받고있는, 이탈리아의 다발성 경화증 협회 (AISM가 2010/R/31을 부여), 이탈리아 보건부 (GR08-7), 생명, 방카 아그리콜라 POPOLARE 디 라 구사 (BAPR), 유럽 연구위원회 (ERC) ERC-2010-엑츄에 부여 계약에 따라 없음 260511-SEM_SEM과 유럽 공동체 (EC) 7 회 프레임 워크 프로그램 (FP7/2007-2013) 날개 보조금 협정 N * 내지 아래; 280772 - IONE.

<ol>
	<li>Ben-Hur, T., 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=Transplanted+multipotential+neural+precursor+cells+migrate+into+the+inflamed+white+matter+in+response+to+experimental+autoimmune+encephalomyelitis.">Transplanted multipotential neural precursor cells migrate into the inflamed white matter in response to experimental autoimmune encephalomyelitis.</a> Glia. 41, 73-80 (2003).</li><li>Pluchino, S., 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=Injection+of+adult+neurospheres+induces+recovery+in+a+chronic+model+of+multiple+sclerosis.">Injection of adult neurospheres induces recovery in a chronic model of multiple sclerosis.</a> Nature. 422, 688-694 (2003).</li><li>Chu, K., Kim, M., Jeong, S. W., Kim, S. U., Yoon, B. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+neural+stem+cells+can+migrate,+differentiate,+and+integrate+after+intravenous+transplantation+in+adult+rats+with+transient+forebrain+ischemia.">Human neural stem cells can migrate, differentiate, and integrate after intravenous transplantation in adult rats with transient forebrain ischemia.</a> Neurosci. Lett. 343, 129-133 (2003).</li><li>Bottai, D., Madaschi, L., Di Giulio, A. M., Gorio, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Viability-dependent+promoting+action+of+adult+neural+precursors+in+spinal+cord+injury.">Viability-dependent promoting action of adult neural precursors in spinal cord injury.</a> Mol. Med. 14, 634-644 (2008).</li><li>Pluchino, S., 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=Neurosphere-derived+multipotent+precursors+promote+neuroprotection+by+an+immunomodulatory+mechanism.">Neurosphere-derived multipotent precursors promote neuroprotection by an immunomodulatory mechanism.</a> Nature. 436, 266-271 (2005).</li><li>Einstein, O., 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=Intraventricular+transplantation+of+neural+precursor+cell+spheres+attenuates+acute+experimental+allergic+encephalomyelitis.">Intraventricular transplantation of neural precursor cell spheres attenuates acute experimental allergic encephalomyelitis.</a> Mol. Cell Neurosci. 24, 1074-1082 (2003).</li><li>Chu, K., 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=Human+neural+stem+cell+transplantation+reduces+spontaneous+recurrent+seizures+following+pilocarpine-induced+status+epilepticus+in+adult+rats.">Human neural stem cell transplantation reduces spontaneous recurrent seizures following pilocarpine-induced status epilepticus in adult rats.</a> Brain Res. 1023, 213-221 (2004).</li><li>Jeong, S. W., 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=Human+neural+stem+cell+transplantation+promotes+functional+recovery+in+rats+with+experimental+intracerebral+hemorrhage.">Human neural stem cell transplantation promotes functional recovery in rats with experimental intracerebral hemorrhage.</a> Stroke. 34, 2258-2263 (2003).</li><li>Aboody, K. S., 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=Neural+stem+cells+display+extensive+tropism+for+pathology+in+adult+brain:+evidence+from+intracranial+gliomas.">Neural stem cells display extensive tropism for pathology in adult brain: evidence from intracranial gliomas.</a> Proc. Natl. Acad. Sci. U.S.A. 97, 12846-12851 (2000).</li><li>Muller, F. J., Snyder, E. Y., Loring, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gene+therapy:+can+neural+stem+cells+deliver.">Gene therapy: can neural stem cells deliver.</a> Nat. Rev. Neurosci. 7, 75-84 (2006).</li><li>Martino, G., Pluchino, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+therapeutic+potential+of+neural+stem+cells.">The therapeutic potential of neural stem cells.</a> Nat. Rev. Neurosci. 7, 395-406 (2006).</li><li>Deckert-Schluter, M., Schluter, D., Hof, H., Wiestler, O. D., Lassmann, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+expression+of+ICAM-1,+VCAM-1+and+their+ligands+LFA-1,+Mac-1,+CD43,+VLA-4,+and+MHC+class+II+antigens+in+murine+Toxoplasma+encephalitis:+a+light+microscopic+and+ultrastructural+immunohistochemical+study.">Differential expression of ICAM-1, VCAM-1 and their ligands LFA-1, Mac-1, CD43, VLA-4, and MHC class II antigens in murine Toxoplasma encephalitis: a light microscopic and ultrastructural immunohistochemical study.</a> J. Neuropathol. Exp. Neurol. 53, 457-468 (1994).</li><li>Hemmer, B., Archelos, J. J., Hartung, H. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+concepts+in+the+immunopathogenesis+of+multiple+sclerosis.">New concepts in the immunopathogenesis of multiple sclerosis.</a> Nat. Rev. Neurosci. 3, 291-301 (2002).</li><li>Butcher, E. C., Picker, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lymphocyte+homing+and+homeostasis.">Lymphocyte homing and homeostasis.</a> Science. 272, 60-66 (1996).</li><li>Rampon, C., 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=Molecular+mechanism+of+systemic+delivery+of+neural+precursor+cells+to+the+brain:+assembly+of+brain+endothelial+apical+cups+and+control+of+transmigration+by+CD44.">Molecular mechanism of systemic delivery of neural precursor cells to the brain: assembly of brain endothelial apical cups and control of transmigration by CD44.</a> Stem Cells. 26, 1673-1682 (2008).</li><li>Pluchino, S., 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=Human+neural+stem+cells+ameliorate+autoimmune+encephalomyelitis+in+non-human+primates.">Human neural stem cells ameliorate autoimmune encephalomyelitis in non-human primates.</a> Ann. Neurol. 66, 343-354 (2009).</li><li>Martino, G., Pluchino, S., Bonfanti, L., Schwartz, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brain+regeneration+in+physiology+and+pathology:+the+immune+signature+driving+therapeutic+plasticity+of+neural+stem+cells.">Brain regeneration in physiology and pathology: the immune signature driving therapeutic plasticity of neural stem cells.</a> Physiol. Rev. 91, 1281-1304 (2011).</li><li>Aharonowiz, M., 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=Neuroprotective+effect+of+transplanted+human+embryonic+stem+cell-derived+neural+precursors+in+an+animal+model+of+multiple+sclerosis.">Neuroprotective effect of transplanted human embryonic stem cell-derived neural precursors in an animal model of multiple sclerosis.</a> PLoS ONE. 3, e3145 (2008).</li><li>Pluchino, S., 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=Immune+regulatory+neural+stem/precursor+cells+protect+from+central+nervous+system+autoimmunity+by+restraining+dendritic+cell+function.">Immune regulatory neural stem/precursor cells protect from central nervous system autoimmunity by restraining dendritic cell function.</a> PLoS One. 4, (2009).</li><li>Einstein, O., 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=Neural+precursors+attenuate+autoimmune+encephalomyelitis+by+peripheral+immunosuppression.">Neural precursors attenuate autoimmune encephalomyelitis by peripheral immunosuppression.</a> Ann. Neurol. 61, 209-218 (2007).</li><li>Gritti, A., 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=Multipotential+stem+cells+from+the+adult+mouse+brain+proliferate+and+self-renew+in+response+to+basic+fibroblast+growth+factor.">Multipotential stem cells from the adult mouse brain proliferate and self-renew in response to basic fibroblast growth factor.</a> J. Neurosci. 16, 1091-1100 (1996).</li><li>Furlan, R., Pluchino, S., Marconi, P. C., Martino, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cytokine+gene+delivery+into+the+central+nervous+system+using+intrathecally+injected+nonreplicative+viral+vectors.">Cytokine gene delivery into the central nervous system using intrathecally injected nonreplicative viral vectors.</a> Methods Mol. Biol. 215, 279-289 (2003).</li><li>Constantin, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Visualization+and+analysis+of+adhesive+events+in+brain+microvessels+by+using+intravital+microscopy.">Visualization and analysis of adhesive events in brain microvessels by using intravital microscopy.</a> Methods Mol. Biol. 239, 189-198 (2004).</li><li>Politi, L. S., 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=Magnetic-resonance-based+tracking+and+quantification+of+intravenously+injected+neural+stem+cell+accumulation+in+the+brains+of+mice+with+experimental+multiple+sclerosis.">Magnetic-resonance-based tracking and quantification of intravenously injected neural stem cell accumulation in the brains of mice with experimental multiple sclerosis.</a> Stem Cells. 25, 2583-2592 (2007).</li><li>Melzi, R., 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=Co-graft+of+allogeneic+immune+regulatory+neural+stem+cells+(NPC)+and+pancreatic+islets+mediates+tolerance,+while+inducing+NPC-derived+tumors+in+mice.">Co-graft of allogeneic immune regulatory neural stem cells (NPC) and pancreatic islets mediates tolerance, while inducing NPC-derived tumors in mice.</a> PLoS One. 5, (2010).</li><li>Amariglio, 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=Donor-derived+brain+tumor+following+neural+stem+cell+transplantation+in+an+ataxia+telangiectasia+patient.">Donor-derived brain tumor following neural stem cell transplantation in an ataxia telangiectasia patient.</a> PLoS Med. 6, (2009).</li><li>Ben-Hur, T., 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=Effects+of+proinflammatory+cytokines+on+the+growth,+fate,+and+motility+of+multipotential+neural+precursor+cells.">Effects of proinflammatory cytokines on the growth, fate, and motility of multipotential neural precursor cells.</a> Mol. Cell Neurosci. 24, 623-631 (2003).</li><li>Giusto, E., Donega, M., Cossetti, C., Pluchino, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuro-immune+interactions+of+neural+stem+cell+transplants:+From+animal+disease+models+to+human+trials.">Neuro-immune interactions of neural stem cell transplants: From animal disease models to human trials.</a> Exp. Neurol. , (2013).</li></ol>

저자가 공개하는 게 없다.

체세포 줄기 세포 기반 요법은 MS2 11과 같은 만성 염증성 CNS 장애 치료를위한 가장 유망한 전략의 하나로 부상하고있다. 자신의 치료 효과를 유지하는 메커니즘은 아직 완전히 규명 될 필요가 있지만, 신경 퇴행성 질환의 다양한 실험 모델에서 NPC 이식의 큰 영향은 세포가 곧 인간의 연구에 적용 할 수있다 줄기 다소 도발적인 믿음에 상승을 부여하고있다. 그러나, 우리는 몇 가지 주요 문?...

강력한 증거 CNS 장애 1-8의 동물 모델에서 체세포 신경 줄기 / 전구 세포 (NPC가)의 이식의 치료 효능을 입증 생체 내 연구에서 생겨났다. 이러한 실험 결과는 임상 응용 프로그램으로 변환하기 전에 그럼에도 불구하고, 호스트에 줄기 세포의 전달과 관련된 문제의 수는 신중한 검토가 필요합니다. 다 초점, 만성 염증성 뇌 질환 (nonhematopoietic) 원기를 회복시키는 줄기 세포 치료의 발전으로 특히 상당한 장애물은 NPC 주입의 최적 경로를 식별합니다. 대상 질병의 병태 생리의 확고한 이해 (초점 또는 다 초점, 기본 염증 또는 기본 퇴행성) 및 전달 기술과 관련된 가능성과 위험 문제의 신중한 분석은 줄기 세포 전달을위한 최적의 프로토콜을 식별합니다. 동안 초점 ( <em&gt; 예. 신경계의 실질에) 줄기 세포 이식 손상의 공간적으로 밀폐 된 공간을 특징으로 중추 신경계 질환의 치료에 대한 논리적 인 접근 방식 (예를 들면 파킨슨 병과 헌팅턴의 질병, 뇌와 척수 외상성 손상, 뇌졸중), 바로 그 방법을 증명할 수 예, 다 초점 만성, 공간적으로 전파 CNS 손상은 시간이 지남에 축적 MS, 같은 조건에서 실질적으로 가능하지합니다. 후자의 경우, 각각의 병변에 초점 세포 주사를 대상으로는 따라서 적게 침략 NPC 이식 타겟팅 CNS의 대안, 더 적절한 방법의 식별을 자극 CNS 실질 내 긴 거리를 통해 마이그레이션하는 이식의 NPC의 제한된 용량에 의해 방해된다 . CNS9 외부 혈관 내 주입했을 때 큰 약속은 생쥐의 NPC가 두개 내 종양 (예를 들면. 신경 교종)을 대상으로 관찰에서 나왔다. 이 정액 다음줄기 세포 pathotrophism 10의 생체 증거, 광범위한 데이터는 실험자가 면역 뇌척수염 (EAE)와 동물 실험의 가능성과의 NPC의 조직 이식의 치료 효과에 관한 축적 된 정맥 중 하나를 통해 염증 CNS 손상의 모델로 (IV) 또는 뇌 실내 (ICV). NPC 주입 1,2,5,6,8 .. 우리는 먼저이 대상을 입력 염증 CNS를, 이후 여러 세포 간 결합에 이식의 NPC의 능력에 의존하는 것으로 나타났습니다 생체 (11)의 특정 미세 환경 내에서 통신 프로그램. 특히 CNS를 대상으로하기 위해,의 NPC는 뇌척수액 (CSF) ICV 주입에 의한 순환, 또는 정맥 주사를 통해 혈류로 직접 전달됩니다. 일단 혈류 또는 CSF 중 하나를 입력, 이식의 NPC가 적극적으로 상호 작용혈액 뇌 (BBB)​​ 또는 혈액 뇌척수액 (BCSFB) 장애 및 중추 신경계의 실질을 입력합니다. NPC 이식하고 BBB (또는 BCSFB) 사이의 상호 작용은 NPC의 표면에 세포 부착 분자 (CAM들)의 특정 설정에 의해 규제 및 활성화 내피 / 뇌실막 세포 12-14에 CAM 카운터 - 리간드의 높은 수준의 발현에 의해 촉진된다. 이러한 캠 예는 히알루 론산에 대한 수용체, CD44, 및 세포 간 접착 분자 (ICAM) -1 리간드 늦게 항원 (VLA) -4 5,15,16 (즉, 백혈구의 활성화 뇌실막과의 상호 작용의 책임을 포함 내피 세포), 그리고 훨씬 낮은 정도의 림프구 기능 관련 항원 (LFA) -1 P-셀렉틴 당 단백질의 리간드 (PSGL) -1. NPC들도 CCR1, CCR2, CCR5, CXCR3 및 CXCR4 등의 케모카인 수용체의 넓은 범위를 표현하는 (그러나 CCR3 및 CCR7을 표명하지 아니합니다) 시험 관내 및 생체 5,16 모두 기능적으로 활성화되어있는. 따라서, systemica베드로 주입의 NPC는 염증 CNS의 수준에 축적, G-단백질 결합 수용체 (GPCR에)와 함께,이 캠 사용합니다. 반대로, NPC의 혈관이나 뇌척수액 공간 루트 2를 통해 CNS를 입력하지 않은 건강한 생쥐에 체계적으로 주입했다. 화학적으로 유도 된 뇌염의 모델로 조직의 사이토 카인 또는 lypopolisaccharide (LPS) 주입 다음 CNS의 염증, 또는 내피 / 뇌실막 세포 활성화는 뇌와 척수 2에 체계적으로 주입의 NPC의 축적을위한 필요가있다. 따라서, 조직 NPC 치료와 CNS의 성공 타겟팅은 뇌와 척수 환경이 축적의 NPC의 내피 마이그레이션에 도움이되는 기회 (우)의 질병 특정 윈도우의 식별에 따라 달라집니다. 이러한 조건은 일반적으로 급성 및 아 급성 염증 (17)의 상황에서 발생한다. 일단 미분화의 NPC를 이식, CNS를 입력 한마우스의 clinico 병리학 기능뿐만 아니라 EAE와 큰, 인간이 아닌 영장류를 개선하는 것으로 나타났다. 이 질문에 답변 최소한의 세포 교체 2 및 비 CNS 염증 영역 (19, 20) (예를 들어, 림프절) 대 혈관 주위 CNS 2,5,6,18 내 면역 조절 및 신경 주변 분비 인자의 놀라운 분비에서 의존하는 기술되었다 염증 세포의 신호는 면역 세포에게 5 침투에 의해 유발. 여기에서 우리는 만성 EAE의 마우스 모델에 체세포의 NPC의 전신 주사의 주요 방법 론적 측면을 설명합니다. 보다 구체적으로, 우리는 확장 및 성인 C57BL / 6 마​​우스의 subventricular 영역 (SVZ)에서 이식 체세포의 NPC를 준비, 우리는 (I)로 설정 한 프로토콜을 도출 정의; (II)는 마우스의 만성 EAE를 유도 그리고 (iii) 치료 효과가 전신 (IV 또는 ICV) NPC 이식을 수행하는 전n을 EAE 마우스.

<table border="0" cellpadding="0" cellspacing="0" width="1474">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
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			<td height="21" style="height:21px;width:753px;">Cell culture</td>
			<td style="width:300px;"></td>
			<td style="width:149px;"></td>
			<td style="width:272px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">EBSS</td>
			<td>Sigma</td>
			<td>E2888</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">L-Cystein</td>
			<td>SIGMA-ALDRICH CO LTD</td>
			<td>C7352</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Papain</td>
			<td>WORTHINGTON</td>
			<td>30H11965</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">EDTA</td>
			<td>Fisher scientific</td>
			<td>D/0700/50</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Mouse NeuroCult basal medium</td>
			<td>Stem Cell technologies</td>
			<td>05700</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">NeuroCult proliferation supplements</td>
			<td>Stem Cell technologies</td>
			<td>05701</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Heparin</td>
			<td>Sigma</td>
			<td>H3393</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Basic fibroblast growth factor</td>
			<td>Peprotech</td>
			<td>100-18B-1000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Epidermal growth factor</td>
			<td>Peprotech</td>
			<td>AF-100-15-1000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Pen/Strep</td>
			<td>Invitrogen</td>
			<td>1514012</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Matrigel (coating solution)</td>
			<td>BD biosciences</td>
			<td>354230</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">NeuroCult&#174; Differentiation Kit (Mouse)</td>
			<td>Stem cell technologies</td>
			<td>05704</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Accumax</td>
			<td>eBioscience</td>
			<td>00-4666-56</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dulbecco&#39;s PBS (DPBS) (10x) without Ca&#38; Mg</td>
			<td>PAA LABORATORIES LTD</td>
			<td>H15-011</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Myco trace</td>
			<td>PAA LABORATORIES LTD</td>
			<td>Q052-020</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dimethyl sulfoxide (DMSO)</td>
			<td>SIGMA</td>
			<td>D2650</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">immunofluorescence</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Normal goat serum</td>
			<td>PAA LABORATORIES LTD</td>
			<td>B11-035</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Polyethylene glycol p-(1,1,3,3-tetramethylbutyl)-phenyl ether</td>
			<td>SIGMA-ALDRICH CO LTD</td>
			<td>T8787</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Mouse anti Nestin</td>
			<td>Abcam</td>
			<td>ab11306</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rabbit anti GFAP</td>
			<td>DAKO</td>
			<td>203344</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Mouse anti Histone H3 (phospho S10)&#160;</td>
			<td>Abcam</td>
			<td>ab14955</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rabbit anti MAP-2</td>
			<td>Abcam</td>
			<td>ab32454</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rat anti MBP</td>
			<td>AbD SEROTEC</td>
			<td>MCA409S</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Anti-O4 Antibody, clone 81 | MAB345</td>
			<td>Millipore</td>
			<td>MAB345</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">DAPI</td>
			<td>Invitrogen</td>
			<td>D1306</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Mounting solution</td>
			<td>DAKO</td>
			<td>S3023</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">EAE</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Freund&#39;s Adjuvant Incomplete</td>
			<td>SIGMA-ALDRICH CO LTD</td>
			<td>F5506</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Mycobacterium tuberculosis&#160;</td>
			<td>DIFCO</td>
			<td>H37Ra</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">MOG(35&#8211;55)&#160;</td>
			<td>Espikem</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Pertussis toxin</td>
			<td>List Biological Laboratories</td>
			<td>181</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tissue processing</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Iris scissor straight</td>
			<td>Fine Sciences Tolls</td>
			<td>14060-09</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Blunt/bended forceps</td>
			<td>Fine Sciences Tolls</td>
			<td>11080-02</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Brain slicer</td>
			<td>Zivic instruments</td>
			<td>BSMAS005-1</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical blades</td>
			<td>Swann-Morton</td>
			<td>324</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">P200, P1000 pipettes</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ketamine (Vetalar)</td>
			<td style="width:300px;">Boehringer Ingelheim</td>
			<td>01LC0030&#160;&#160;</td>
			<td style="width:272px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Xylazine (Rompun)</td>
			<td>Bayer</td>
			<td>32371</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Stereotaxic frame</td>
			<td>KOPF</td>
			<td>Model 900</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Hamilton syringe</td>
			<td>Hamilton</td>
			<td>7762-04</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Paraformaldehyde (PFA)</td>
			<td>SIGMA</td>
			<td>158127</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">VECTASTAIN Elite ABC Kit</td>
			<td>vector laboratories</td>
			<td>PK-6100</td>
			<td></td>
		</tr>
	</tbody>
</table>

systemic injection of neural stem/progenitor cells in mice with chronic eae

신경 줄기 / 전구 세포 (NPC들)은 재생 신경과 뇌의 수리 또는 복원을 목표로 이식 방법에 대한 유망한 줄기 세포의 원천입니다. 이 지시문은 뇌 수리가 신경 질환의 여러 전임상 모델에서 초점 또는 전신 NPC 이식 후 달성하는 광범위한 증거에서 생겨났다. 이러한 실험 결과는 긴급 평가를 필요로 뇌 질환에 대한 회복 줄기 세포 치료의 주요 장애물 중 하나로서 세포 전달 경로를 발견했습니다. 뇌실질 내 줄기 세포 이식은 척수 손상과 파킨슨 병으로 격리와 접근 뇌 병변을 특징으로하는 병리에 대한 논리적 인 접근 방식을 나타냅니다. 불행하게도,이 원칙은 다발성 경화증 (MS)를 포함, 다 초점 염증 및 (시간과 공간 모두) 전파 특성에 의해 특징 조건에 제대로 적용 할 수있다. 예, 뇌 SYST에 의해 대상으로EMIC NPC 전달은 뇌 및 중추 신경계 (CNS)의 실험 만성 염증성 손상에 의해 영향을 설치류 및 영장류의 척수 세포를 제공하는 저 침습 치료 효과가 프로토콜이되고있다. 세포 전달이 대안적인 방법은, NPC의 pathotropism에 (ⅰ) 기능성 세포 부착 분자 및 염증성 사이토 카인 및 케모카인 수용체를 통해 환경을 감지하도록 특별히 타고난 용량 의존; (II) 누출 해부학 정맥 주사 후 장벽 (. IV) 또는 뇌 실내 (ICV) 주사를 건너; (III) 염증성 뇌와 척수 손상의 여러 혈관 주위 사이트 (들)의 수준에 축적; 및 (iv) 생체 내에서 다른 호스트 표적 세포 상 현저한 조직 영양 및 면역 조절 효과를 발휘한다. 여기에서 우리는 우리가 IV에 대한 개발 방법을 설명합니다. 및 &lt;EM&gt; ICV의 만성 CNS 염증성 탈수 초화의 모델로 실험자가 면역 뇌척수염 (EAE)와 생쥐의 동계의 NPC의 전달, 그리고 재생 신경의 염증이 뇌의 선택적 타겟팅에 대한 가치있는 기술과 조직의 줄기 세포 전달을 직시하다.

NPC 유도 및 특성 SVZ 해부는 기계 및 효소 분해 (그림 1A)에 의해 6~8주 된 C57BL / 6 마우스의 (N = 5-7 마우스 / 수영장) 풀에서 수행됩니다. CGM에서 배양 몇 일 후, 자유 부동을 neurospheres는 (그림 1A 및 1B)를 형성 시작합니다. 주요 분야는 수집 및 기계적으로 모든 4-5 DIV 계대 있습니다. 계대시, 라이브와 죽은 세포의 수는 확정 누적 세포 수는 성장 곡선 (그...

Watch this Scientific Journal Video about Systemic Injection of Neural Stem/Progenitor Cells in Mice with Chronic EAE at JoVE.com

Systemic Injection of Neural Stem/Progenitor Cells in Mice with Chronic EAE

신경 줄기 / 전구 세포 (NPC가)의 이식은 재생 신경의 큰 약속을 보유하고 있습니다. 의 NPC의 전신 배달은 뇌와 중추 신경계의 실험 만성 염증성 손상에 의해 영향을 설치류 및 영장류의 척수에 줄기 세포를 제공하는 효과적인 침략 낮은 및 치료 매우 효과적인 프로토콜로 설정되어 있습니다.

만성 EAE와 마우스의 신경 줄기 / 전구 세포의 전신 주입

Neural stem/precursor cells (NPCs) are a promising stem cell source for transplantation approaches aiming at brain repair or restoration in regenerative ...

systemic-injection-neural-stemprogenitor-cells-mice-with-chronic

Research

JoVE Journal

Immunology and Infection

17.6K 조회수.  University of Cambridge, UK.  신경 줄기 / 전구 세포 (NPC가)의 이식은 재생 신경의 큰 약속을 보유하고 있습니다. 의 NPC의 전신 배달은 뇌와 중추 신경계의 실험 만성 염증성 손상에 의해 영향을 설치류 및 영장류의 척수에 줄기 세포를 제공하는 효과적인 침략 낮은 및 치료 매우 효과적인 프로토콜로 설정되어 있습니다. 

비디오: Systemic Injection of Neural Stem/Progenitor Cells in Mice with Chronic EAE

Identifying DNA Mutations in Purified Hematopoietic Stem/Progenitor Cells

In recent years, it has become apparent that genomic instability is tightly related to many developmental disorders, cancers, and aging. Given that stem cells are responsible for ensuring tissue homeostasis and repair throughout life, it is reasonable to hypothesize that the stem cell population is critical for preserving genomic integrity of tissues. Therefore, significant interest has arisen in assessing the impact of endogenous and environmental factors on genomic integrity in stem cells and their progeny, aiming to understand the etiology of stem-cell based diseases.
LacI transgenic mice carry a recoverable &#955;&#160;phage vector encoding the LacI reporter system, in which the LacI gene serves as the mutation reporter. The result of a mutated LacI gene is the production of &#946;-galactosidase that cleaves a chromogenic substrate, turning it blue. The LacI reporter system is carried in all cells, including stem/progenitor cells and can easily be recovered and used to subsequently infect E. coli. After incubating infected E. coli on agarose that contains the correct substrate, plaques can be scored; blue plaques indicate a mutant LacI gene, while clear plaques harbor wild-type. The frequency of blue (among clear) plaques indicates the mutant frequency in the original cell population the DNA was extracted from. Sequencing the mutant LacI gene will show the location of the mutations in the gene and the type of mutation.
The LacI transgenic mouse model is well-established as an in vivo mutagenesis assay. Moreover, the mice and the reagents for the assay are commercially available. Here we describe in detail how this model can be adapted to measure the frequency of spontaneously occurring DNA mutants in stem cell-enriched Lin-IL7R-Sca-1+cKit++(LSK) cells and other subpopulations of the hematopoietic system.

Here we describe an in vivo mutagenesis assay for small numbers of purified hematopoietic cells using the LacI transgenic mouse model.&#160;The LacI gene can be isolated to determine the frequency, location, and type of DNA mutants&#160;spontaneously arisen or after exposure to genotoxins.

정제 된 조혈 줄기 / 전구 세포에 DNA 돌연변이를 확인

In recent years, it has become apparent that genomic instability is tightly related to many developmental disorders, cancers, and aging. Given that stem ...

연구

면역학 및 감염병학

Long Term Chronic Pseudomonas aeruginosa Airway Infection in Mice

A mouse model of chronic airway infection is a key asset in cystic fibrosis (CF) research, although there are a number of concerns regarding the model itself. Early phases of inflammation and infection have been widely studied by using the Pseudomonas aeruginosa agar-beads mouse model, while only few reports have focused on the long-term chronic infection in vivo. The main challenge for long term chronic infection remains the low bacterial burden by P. aeruginosa and the low percentage of infected mice weeks after challenge, indicating that bacterial cells are progressively cleared by the host.
This paper presents a method for obtaining efficient long-term chronic infection in mice. This method is based on the embedding of the P. aeruginosa clinical strains in the agar-beads in vitro, followed by intratracheal instillation in C57Bl/6NCrl mice. Bilateral lung infection is associated with several measurable read-outs including weight loss, mortality, chronic infection, and inflammatory response. The P. aeruginosa RP73 clinical strain was preferred over the PAO1 reference laboratory strain since it resulted in a comparatively lower mortality, more severe lesions, and higher chronic infection. P. aeruginosa colonization may persist in the lung for over three months. Murine lung pathology resembles that of CF patients with advanced chronic pulmonary disease.
This murine model most closely mimics the course of the human disease and can be used both for studies on the pathogenesis and for the evaluation of novel therapies.

Long Term Chronic Pseudomonas aeruginosa Airway Infection in Mice

We describe the agar-beads method to establish persistent long-term chronic Pseudomonas aeruginosa airway infection in the mouse model.

장기 만성<em&gt; 녹농균</em마우스의&gt;기도 감염

A mouse model of chronic airway infection is a key asset in cystic fibrosis (CF) research, although there are a number of concerns regarding the model ...

Myelin Oligodendrocyte Glycoprotein (MOG35-55) Induced Experimental Autoimmune Encephalomyelitis (EAE) in C57BL/6 Mice

Multiple sclerosis is a chronic neuroinflammatory demyelinating disorder of the central nervous system with a strong neurodegenerative component. While the exact etiology of the disease is yet unclear, autoreactive T lymphocytes are thought to play a central role in its pathophysiology. MS therapy is only partially effective so far and research efforts continue to expand our knowledge on the pathophysiology of the disease and to develop novel treatment strategies. Experimental autoimmune encephalomyelitis (EAE) is the most common animal model for MS sharing many clinical and pathophysiological features. There is a broad diversity of EAE models which reflect different clinical, immunological and histological aspects of human MS. Actively-induced EAE in mice is the easiest inducible model with robust and replicable results. It is especially suited for investigating the effects of drugs or of particular genes by using transgenic mice challenged by autoimmune neuroinflammation. Therefore, mice are immunized with CNS homogenates or peptides of myelin proteins. Due to the low immunogenic potential of these peptides, strong adjuvants are used. EAE susceptibility and phenotype depends on the chosen antigen and rodent strain. C57BL/6 mice are the commonly used strain for transgenic mouse construction and respond among others to myelin oligodendrocyte glycoprotein (MOG). The immunogenic epitope MOG35-55 is suspended in complete Freund's adjuvant (CFA) prior to immunization and pertussis toxin is applied on the day of immunization and two days later. Mice develop a "classic" self-limited monophasic EAE with ascending flaccid paralysis within 9-14 days after immunization. Mice are evaluated daily using a clinical scoring system for 25-50 days. Special considerations for care taking of animals with EAE as well as potential applications and limitations of this model are discussed.

Myelin Oligodendrocyte Glycoprotein MOG35-55 Induced Experimental Autoimmune Encephalomyelitis EAE in C57BL/6 Mice

Experimental autoimmune encephalomyelitis (EAE) is an established animal model of multiple sclerosis. C57BL/6 mice are immunized with myelin oligodendrocyte glycoprotein (MOG) peptide 35-55 (MOG35-55), resulting in an ascending flaccid paralysis caused by autoreactive immune cells in the central nervous system. Protocols for disease induction and monitoring will be discussed.

수초 희소 돌기 아교 세포 당 단백질 (MOG<sub&gt; 35-55</sub&gt;) C57BL / 6 쥐에 실험자가 면역 뇌척수염 (EAE) 유도

Multiple sclerosis is a chronic neuroinflammatory demyelinating disorder of the central nervous system with a strong neurodegenerative component. While ...

A Mouse Model for Pathogen-induced Chronic Inflammation at Local and Systemic Sites

Chronic inflammation is a major driver of pathological tissue damage and a unifying characteristic of many chronic diseases in humans including neoplastic, autoimmune, and chronic inflammatory diseases. Emerging evidence implicates pathogen-induced chronic inflammation in the development and progression of chronic diseases with a wide variety of clinical manifestations. Due to the complex and multifactorial etiology of chronic disease, designing experiments for proof of causality and the establishment of mechanistic links is nearly impossible in humans. An advantage of using animal models is that both genetic and environmental factors that may influence the course of a particular disease can be controlled. Thus, designing relevant animal models of infection represents a key step in identifying host and pathogen specific mechanisms that contribute to chronic inflammation.
Here we describe a mouse model of pathogen-induced chronic inflammation at local and systemic sites following infection with the oral pathogen Porphyromonas gingivalis, a bacterium closely associated with human periodontal disease. Oral infection of specific-pathogen free mice induces a local inflammatory response resulting in destruction of tooth supporting alveolar bone, a hallmark of periodontal disease. In an established mouse model of atherosclerosis, infection with P. gingivalis accelerates inflammatory plaque deposition within the aortic sinus and innominate artery, accompanied by activation of the vascular endothelium, an increased immune cell infiltrate, and elevated expression of inflammatory mediators within lesions. We detail methodologies for the assessment of inflammation at local and systemic sites. The use of transgenic mice and defined bacterial mutants makes this model particularly suitable for identifying both host and microbial factors involved in the initiation, progression, and outcome of disease. Additionally, the model can be used to screen for novel therapeutic strategies, including vaccination and pharmacological intervention.

Animal models have proven to be invaluable tools in defining host and pathogen specific mechanisms that contribute to the development of chronic inflammation. Here we describe a mouse model of oral infection with the human pathogen Porphyromonas gingivalis and detail methodologies to assess the progression of inflammation at local and systemic sites.

로컬 및 전신 사이트에서 병원체에 의한 만성 염증을위한 마우스 모델

Chronic inflammation is a major driver of pathological tissue damage and a unifying characteristic of many chronic diseases in humans including ...

Phenotypic and Functional Analysis of Activated Regulatory T Cells Isolated from Chronic Lymphocytic Choriomeningitis Virus-infected Mice

Regulatory T (Treg) cells, which express Foxp3 as a transcription factor, are subsets of CD4+ T cells. Treg cells play crucial roles in immune tolerance and homeostasis maintenance by regulating the immune response. The primary role of Treg cells is to suppress the proliferation of effector T (Teff) cells and the production of cytokines such as IFN-&#947;, TNF-&#945;, and IL-2. It has been demonstrated that Treg cells&#39; ability to inhibit the function of Teff cells is enhanced during persistent pathogen infection and cancer development. To clarify the function of Treg cells under resting or inflamed conditions, a variety of in vitro suppression assays using mouse or human Treg cells have been devised. The main aim of this study is to develop a method to compare the differences in phenotype and suppressive function between resting and activated Treg cells. To isolate activated Treg cells, mice were infected with lymphocytic choriomeningitis virus (LCMV) clone 13 (CL13), a chronic strain of LCMV. Treg cells isolated from the spleen of LCMV CL13-infected mice exhibited both the activated phenotype and enhanced suppressive activity compared with resting Treg cells isolated from na&#239;ve mice. Here, we describe the basic protocol for ex vivo phenotype analysis to distinguish activated Treg cells from resting Treg cells. Furthermore, we describe a protocol for the measurement of the suppressive activity of fully activated Treg cells.

Here, we describe a protocol to analyze the phenotype of regulatory T (Treg) cells isolated from na&#239;ve and chronic lymphocytic choriomeningitis virus-infected mice. In addition, we provide a process to evaluate the suppressive activity of the Treg cells.

표현형 및 만성 림프 구성 맥락 수막염 바이러스에 감염된 마우스에서 고립 활성화 규제 T 세포의 기능 분석

Regulatory T (Treg) cells, which express Foxp3 as a transcription factor, are subsets of CD4+ T cells. Treg cells play crucial roles in immune tolerance ...

Analysis of Lymphocyte Extravasation Using an In Vitro Model of the Human Blood-brain Barrier

Lymphocyte extravasation into the central nervous system (CNS) is critical for immune surveillance. Disease-related alterations of lymphocyte extravasation might result in pathophysiological changes in the CNS. Thus, investigation of lymphocyte migration into the CNS is important to understand inflammatory CNS diseases and to develop new therapy approaches. Here we present an in vitro model of the human blood-brain barrier to study lymphocyte extravasation. Human brain microvascular endothelial cells (HBMEC) are confluently grown on a porous polyethylene terephthalate transwell insert to mimic the endothelium of the blood-brain barrier. Barrier function is validated by zonula occludens immunohistochemistry, transendothelial electrical resistance (TEER) measurements as well as analysis of evans blue permeation. This model allows investigation of the diapedesis of rare lymphocyte subsets such as CD56brightCD16dim/- NK cells. Furthermore, the effects of other cells, cytokines and chemokines, disease-related alterations, and distinct treatment regimens on the migratory capacity of lymphocytes can be studied. Finally, the impact of inflammatory stimuli as well as different treatment regimens on the endothelial barrier can be analyzed.

Analysis of Lymphocyte Extravasation Using an In Vitro Model of the Human Blood-brain Barrier

Here, we describe a human blood-brain barrier model enabling to investigate lymphocyte transmigration into the central nervous system in vitro.

를 사용하여 림프구 혈관 외로 유출 분석<em&gt; 체외</em&gt; 인간의 혈액 - 뇌 장벽의 모델

Lymphocyte extravasation into the central nervous system (CNS) is critical for immune surveillance. Disease-related alterations of lymphocyte ...

A Chronic Autoimmune Dry Eye Rat Model with Increase in Effector Memory T Cells in Eyeball Tissue

Dry eye disease is a very common condition that causes morbidity and healthcare burden and decreases the quality of life. There is a need for a suitable dry eye animal model to test novel therapeutics to treat autoimmune dry eye conditions. This protocol describes a chronic autoimmune dry eye rat model. Lewis rats were immunized with an emulsion containing lacrimal gland extract, ovalbumin, and complete Freund&#39;s adjuvant. A second immunization with the same antigens in incomplete Freund&#39;s adjuvant was administered two weeks later. These immunizations were administered subcutaneously at the base of the tail. To boost the immune response at the ocular surface and lacrimal glands, lacrimal gland extract and ovalbumin were injected into the forniceal subconjunctiva and lacrimal glands 6 weeks after the first immunization. The rats developed dry eye features, including reduced tear production, decreased tear stability, and increased corneal damage. Immune profiling by flow cytometry showed a preponderance of CD3+ effector memory T cells in the eyeball.

This report describes a method to induce chronic experimental autoimmune dry eye in Lewis rats through immunization with an emulsion of rat lacrimal gland extract, ovalbumin, and complete Freund's adjuvant, followed by the injection of lacrimal gland extract and ovalbumin into the forniceal subconjunctiva and lacrimal glands six weeks later.

안구 조직에서 이펙터 기억 T 세포의 증가를 동반 한 만성자가 면역 건식 ​​쥐 모델

Dry eye disease is a very common condition that causes morbidity and healthcare burden and decreases the quality of life. There is a need for a suitable ...

Blocking Lymph Flow by Suturing Afferent Lymphatic Vessels in Mice

Lymphatic vessels are critical in maintaining tissue fluid balance and optimizing immune protection by transporting antigens, cytokines, and cells to draining lymph nodes (LNs). Interruption of lymph flow is an important method when studying the function of lymphatic vessels. The afferent lymphatic vessels from the murine footpad to the popliteal lymph nodes (pLNs) are well-defined as the only routes for lymph drainage into the pLNs. Suturing these afferent lymphatic vessels can selectively prevent lymph flow to the pLNs. This method allows for interference in lymph flow with minimal damage to the lymphatic endothelial cells in the draining pLN, the afferent lymphatic vessels, as well as other lymphatic vessels around the area. This method has been used to study how lymph impacts high endothelial venules (HEV) and chemokine expression in the LN, and how lymph flows through the adipose tissue surrounding the LN in the absence of functional lymphatic vessels. With the growing recognition of the importance of lymphatic function, this method will have broader applications to further unravel the function of lymphatic vessels in regulating the LN microenvironment and immune responses.

A protocol to block lymph flow by surgical suturing of afferent lymphatic vessels is presented.

마우스에서 흡개 성 림프혈관을 봉합하여 림프 흐름 차단

Lymphatic vessels are critical in maintaining tissue fluid balance and optimizing immune protection by transporting antigens, cytokines, and cells to ...

A Minimally Invasive, Accurate, and Efficient Technique for Intrathymic Injection in Mice

Intrathymic injection in mouse models is an important technique for studying thymic and immune function, including genetic and acquired T cell disorders. This requires methods for the direct deposition of reagents and/or cells into the thymus of living mice. Traditional methods of intrathymic injection include thoracic surgery or minimally invasive percutaneous blind injections, both of which have significant limitations. Ultra-high frequency ultrasound imaging devices have made image-guided percutaneous injections possible in mice, greatly improving the injection accuracy of the percutaneous injection approach and enabling the injection of smaller targets. However, image-guided injections rely on the utilization of an integrated rail system, making this a rigid and time-consuming procedure. A unique, safe, and efficient method for percutaneous intrathymic injections in mice is presented here, eliminating reliance on the rail system for injections. The technique relies on using a high-resolution micro-ultrasound unit to image the mouse thymus noninvasively. Using a free-hand technique, a radiologist can place a needle tip directly into the mouse thymus under sonographic guidance. Mice are cleaned and anesthetized before imaging. For an experienced radiologist adept at ultrasound-guided procedures, the learning period for the stated technique is quite short, typically within one session. The method has a low morbidity and mortality rate for the mice and is much faster than current mechanically assisted techniques for percutaneous injection. It allows the investigator to efficiently perform precise and reliable percutaneous injections of thymuses of any size (including very small organs such as the thymus of aged or immunodeficient mice) with minimal stress on the animal. This method enables the injection of individual lobes if desired and facilitates large-scale experiments due to the time-saving nature of the procedure.

The present protocol describes an interventional radiology procedure established for intrathymic injection in mice to avoid the risk of open surgery and improve the accuracy of blind percutaneous injections.

마우스에서 흉강내 주사를 위한 최소 침습적이고 정확하며 효율적인 기술

Intrathymic injection in mouse models is an important technique for studying thymic and immune function, including genetic and acquired T cell disorders. ...

Dual-Dye Live Cell Analysis를 사용한 NET 형성 정량화

Real-Time High Throughput Technique to Quantify Neutrophil Extracellular Traps Formation in Human Neutrophils

Neutrophils are myeloid-lineage cells that form a crucial part of the innate immune system. The past decade has revealed additional key roles that neutrophils play in the pathogenesis of cancer, autoimmune diseases, and various acute and chronic inflammatory conditions by contributing to the initiation and perpetuation of immune dysregulation through multiple mechanisms, including the formation of neutrophil extracellular traps (NETs), which are structures crucial in antimicrobial defense. Limitations in techniques to quantify NET formation in an unbiased, reproducible, and efficient way have restricted our ability to further understand the role of neutrophils in health and diseases. We describe an automated, real-time, high-throughput method to quantify neutrophils undergoing NET formation using a live cell imaging platform coupled with a membrane permeability-dependent dual-dye approach using two different DNA dyes to image intracellular and extracellular DNA. This methodology is able to help assess neutrophil physiology and test molecules that can target NET formation.

저자 스포트라이트: Quantifying Neutrophil Extracellular Traps in Disease and Drug Screening Using Dual-Color Live-Cell Imaging(이중 색상 생세포 이미징을 사용한 질병 및 약물 스크리닝에서 호중구 세포외 트랩 정량화)

We present an automated high-throughput method to quantify neutrophil extracellular traps (NETs) utilizing the live cell analysis system, coupled with a membrane permeability-dependent dual-dye approach.