ANALYSES OF FOUNDER CELLS IN THE EPIBLAST AND SOMITES
ANALYSES OF FOUNDER CELLS IN THE EPIBLAST AND SOMITES
批准号:
2665706
负责人:
MINDY GEORGE-WEINSTEIN
金额:
$10.11万
依托单位国家:
美国
项目类别:
财政年份:
1998
资助国家:
美国
项目状态:
已结题
起止时间:
1998-07-01 至 2001-06-30
中文摘要
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英文摘要
DESCRIPTION: The George-Weinstein laboratory has demonstrated that cells
from the presomitic segmental plate mesoderm and epiblast are able to
differentiate into skeletal muscle and chondroblasts when isolated from
surrounding tissues of the embryo and cultured in hormone and protein free
medium. mRNA for MyoD and Pax-1, transcription factors that regulate
myogenesis and sclerotome development, are detectable in the segmental plate
by RT-PCR, and MyoD is detectable in the cells of the epiblast. Dr.
George-Weinstein will test the hypothesis that (1) stably programmed founder
cells for skeletal muscle and chondrogenic lineages are present in the chick
embryo prior to the initiation of gastrulation; and, (2) that myogenic and
chondrogenic founder cells recruit their uncommitted neighbors in the somite
to form the myotome and the sclerotome. This latter event happens perhaps
after receiving signals from surrounding structures such as the neural tube
and the notochord. The signals are probably secreted sonic hedgehog and
Wnts 1, 3, and 4. Existing data indicates that differentiation may be
inhibited by tissue interactions, direct cell-cell interactions, and by
nuclear proteins that affect transcription. For instance, when segmental
plates are cultured as single cell suspensions, differentiation occurs
readily, compared to when they are cultured as intact tissues.
In order to resolve much conflicting data and to answer questions concerning
the nature of uncommitted cells and the timing, location, and emergence of
stably committed precursor cells, Dr. George-Weinstein proposes four
specific aims. In aim 1, it will be determined if Pax-1, a factor specific
for chondroblast differentiation, is expressed in the epiblasts isolated
from prestreak stage embryos and from primitive streak stage embryos. This
will substantiate that founder cells for chondroblasts are localized in the
epiblast.
In specific aim 2, the numbers and locations of cells expressing MyoD and
Pax-1 mRNAs will be determined by in situ hybridization on tissue sectioned
from various stage embryos. These experiments are expected to be
informative because of the increased sensitivity (up to 100x) of probes
using fluorescently labeled dendrimers, a high complexity nucleic acid
signal network molecule developed by Polyprobe, Inc. An alternative approach
of increased sensitivity will be in situ PCR followed by probing with the
appropriate dentrimer.
For specific aim 3, the fate of subpopulations of epiblast and segmental
plate cells will be determined. The lab has previously developed 5
monoclonal antibodies that give distinct labeling patterns in unfixed
prestreak and primitive streak stage embryos, each binding to a limited
number of cells within the epiblast. The antibodies will be used to
determine if they are, in fact, markers for myogenic and chondrogenic cells
within the epiblast, and whether the epiblast cells that bind these
antibodies are responsible for establishing tissues within the somite. The
gold-labeled antibodies will be endocytosed by the (stage 2) embryos and
then the embryo will be cultured and sampled over the next 30 hours. A
second experiment involves the co-localization of antibodies with colloidal
gold and fluorescent dendrimers to MyoD and Pax-1.
Specific aim 4 proceeds similarly to aim 3, except those cells which bind
antibody will be ablated by complement mediated lysis. The treated whole
embryo will then be cultured up to 30 hours and the fate of tissue formation
will be analyzed morphologically for dermomyotome and sclerotome formation,
with antibodies to sarcomeric myosin, and by in situ hybridization with
probes for MyoD and Pax-1.
期刊论文(2)
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会议论文
Directing the Fate of Cells to Myogenic Lineages
-
批准号:7564681
-
项目类别:
-
资助金额:$27.63万
-
财政年份:2006
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Directing the Fate of Cells to Myogenic Lineages
-
批准号:7759114
-
项目类别:
-
资助金额:$27.36万
-
财政年份:2006
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Directing the Fate of Cells to Myogenic Lineages
-
批准号:7029248
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项目类别:
-
资助金额:$27.71万
-
财政年份:2006
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Directing the Fate of Cells to Myogenic Lineages
-
批准号:7215715
-
项目类别:
-
资助金额:$26.94万
-
财政年份:2006
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Directing the Fate of Cells to Myogenic Lineages
-
批准号:7555208
-
项目类别:
-
资助金额:$27.63万
-
财政年份:2006
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Origin and Fate of Myogenic Stem Cells
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批准号:6707004
-
项目类别:
-
资助金额:$19.64万
-
财政年份:2003
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Origin and Fate of Myogenic Stem Cells
-
批准号:6848295
-
项目类别:
-
资助金额:$19.64万
-
财政年份:2003
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Origin and Fate of Myogenic Stem Cells
-
批准号:6556168
-
项目类别:
-
资助金额:$23.24万
-
财政年份:2003
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Origin and Fate of Myogenic Stem Cells
-
批准号:7015604
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项目类别:
-
资助金额:$19.18万
-
财政年份:2003
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
Analyses of stem cells in the chick Embryo and Fetus
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批准号:6315756
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项目类别:
-
资助金额:$16.1万
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财政年份:2001
-
负责人:MINDY GEORGE-WEINSTEIN
-
依托单位:
DEVELOPMENTAL POTENTIAL OF CHICK EMBRYO EPIBLAST CELLS
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批准号:2206463
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项目类别:
-
资助金额:$9.97万
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财政年份:1995
-
负责人:MINDY GEORGE-WEINSTEIN
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依托单位:
COMMITMENT AND DEVELOPMENT OF SOMITIC MYOBLASTS
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批准号:3439705
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项目类别:
-
资助金额:$9.39万
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财政年份:1990
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负责人:MINDY GEORGE-WEINSTEIN
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依托单位:
海外基金