Nov 11, 1986 - Immunocytochemical studies of desmin and vimentin in pericapillary cells of chicken. Published by: http://www.sagepublications.com. On behalf ...
Journal of Histochemistry & Cytochemistry http://jhc.sagepub.com/
Immunocytochemical studies of desmin and vimentin in pericapillary cells of chicken. T Fujimoto and S J Singer J Histochem Cytochem 1987 35: 1105 DOI: 10.1177/35.10.3305702 The online version of this article can be found at: http://jhc.sagepub.com/content/35/10/1105
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0022-15
54/87/$3.30
Vol. 35, No.
The Journal of Histochemistry and Cytochemistry Copyright © 1987 by The Histochemical Society,
Original
Immunocytochemical in Pericapillary Cells TOYOSHI
Received
for
ofBiology,
University
publication
November
Article
Studies of Desmin of Chicke&
of California 11,
1986
at San Diego,
and
in revised
Lajolla,
form
The composition of intermediate filaments in pericytes was examined by immunofluorescent and immunoelectron microscopic labeling of frozen sections of various chicken microvascular beds in situ. Pericytes in capillaries of cardiac muscle, exocnine pancreas, and kidney (peritubular capillary) were found to contain both desmin and vimentin. In some capillaries where pericytes do not exist, cells apposed to endotheial cellsthe Ito cell in the hepatic sinusoid and the reticular cell in the splenic sinusoid -were shown to contaiii both ofthe intermediate filament proteins. In contrast, podocytes and mesangial cells around renal glomerular capil-
and
Cal:fornia
92093.
3, 1987;
accepted
March
March
17,
Pericyte; Ito cell; Perisinusoidal cell; Intermediate filaDesmin; Vimentin; Immunofluomescence microscopy; Immunoelectron microscopy. WORDS:
KEY
ment;
peatedly
One
on the cardiovascular
system
has been
on van-
among microvasculan beds. Morphological differentiation among endothelial cells has been revealed not only between large and small blood vessels (for meview see ref. 28) but also among differations
ent
microvasculatunes
that in the
Our
temmediate
desmin
only,
Although endothelial
on vimentin
ofpemicytes
ical
significance
hypotheses
ciety
is even
Supported
examined
of penicytes
is not
suggested.
GM-15971
Singer, the
to con-
vimentin on
in-
the
only, panticu-
(11).
Furthermore,
the
understood, Functions
to SJS, who
physiolog-
although that
several
have
Foundation
been
ne-
and by US Pub-
is an American
Cancer
of Medicine,
Department
So-
Professor.
address: Yoshida,
Kyoto University, Sakyo,
Kyoto
Faculty 606,
Japan.
3 Correspondence to: Sj. Singer, Department ofBiology B-022, sity of California at San Diego, La Jolla, California 92093.
presumed
the other hand, unambiguously cytes have been matory
Univem-
for
pericytes
are
as follows:
(a) contractility
cells, which are present around observed to contract in vivo (2).
the On
penicytes in avians and mammals have never been proved to do so; (b) phagocytosis (4,18,35): penobserved to take up foreign materials in inflam-
reactions;
penicytes broblasts,
and
are believed and other
(c) mesenchymal
to differentiate mesenchymal
plunipotentiality
(20,24):
into smooth muscle cells in adult animals.
cells, fiEven dif-
fenentiation into plasma cells has been postulated by some authors (20). Because of the lack of specific markers for penicytes, however, this
capability To advance
examined
at least limited information about about the molecular ultrastruc-
meager.
Grant
Sj. that
believed
of either depending
chicken
more
been
and was
previously
by a grant from the Weingart
Service
Present
of Anatomy,
desmin,
found
differentiation as of membrane
however,
cells,
plus
adult
the
have
Research 2
of
as well
composed
we therefore have cells, our knowledge
tune
lic Health
finding,
are actually
we have
show
(T. Fujimoto
ofendothelial
only,
lam vasculatume
I
intriguing
filaments
of vimentin
studies,
proteins
to the cytoskeleton most
own
vasculatures
of cytoskeletal
anchored
in press).
In our
cells ofdifferent
distribution
proteins
sist
(17,24).
endothelial
(6Ao902).
1987
1987)
(9,19,24,30,36): e.g., Rouget capillaries of frog eyes, were
ofreseanch
Vimentin
laries contained only vimentin. The presence ofdesmin supports the hypothesis that pericytes may have a contractile apparatus similar to that of vascular smooth muscle cells. Our results also revealed that even in microvascular beds where pericytes are not found, cells having both desmin and vimentin exist next to endotheial cells and may assume similar functions to pericytes. (JHiscochem Cywchem35:1105-1115,
Introduction focus
1105-1115, 1987 Panted in USA.
S. J. SINGER3
and
FUJIMO’102
Department
10. pp.
Inc.
has not been our understanding
some
aspects
convincingly ofpenicytes,
of their
demonstrated. in this study
macnomoleculam
we have
composition
and
ultnastructure. Our approach has been to conduct immunocytochemical studies ofthe cells in situ. Pemicytes are difficult to isolate and
place
tematic
in culture investigation
the tissues
containing
(5),
and
of their penicytes
ticularly by immunoelectmon used techniques ofsemi-thin immunolabeling developed
therefore
the only
cell biology
approach
is to investigate,
by immunofluonescence,
to sysin situ, and
pan-
microscopy. Rr this purpose, we have and ultra-thin frozen sectioning and in this laboratory (31,32).
In our initial studies, the protein composition ofthe intermediate filaments of penicytes was examined. Since the expression of the subunits is thought, results in situ were state
of pemicytes.
penicytes
has been
in general, to be tissue-specific (10,22,27), the expected to characterize the true physiological Although
a preliminary
presented
(5), since
report it is known
about that
cultured the compo1105
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muscle
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penicapillary the proposal
Materials
and
both
is the same
as that
in Ito cells
cells of the splenic
between such mm supports
beds.
contain
(1,12,21,25,26,29).
was found
the reticulan
microvascular examined
ofwhich
are present just exterior to endothelial cytes do not exist. The results therefore
‘,‘?_
in culture
of different
all the
the composition
plus
and
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that
vimentin,
desmin
sinusoid
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from
in vascular
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In addiof the
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cells
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of descells.
Methods
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the cells in situ. In addition, in view of the among endothelial cells (11), we wished to
indicate
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Immunochemical Reagents. All the primary and secondary antibodies used in this study were purified by affinity chromatography on columns made with the respective antigens. Primary antibodies employed were antichicken
gizzard
vimentin
desmin
(34)
(34)
raised
raised
in guinea
in rabbits
pigs.
and
anti-chicken
Anti-desmin
and
erythmocyte
anti-vimentin
anti-
bodies were cross-absorbed with the heterologous antigen to eliminate any possible cmossmeactivity (34). Secondary antibodies were goat anti-rabbit IgG and goat anti-guinea pig IgG, cross-absorbed against each other, conjugated with
rhodamine
femmitin
and/or
(16)
and/or
Preparation with
into
glutaraldehyde. into
pieces,
fixation
aldehyde
tissues
were
Cryosectioning
sm)
frozen
through
a pressure
of 100-150
under
and
immediately, at room
were pH
ism)
according
instrument
depolymemized
kept
7.4,
with
and
to the
temperature.
at 4C
for
cmyoattachment
formup
in liquid
ultra-thin method
0.1%
trimmed
in 0.5%
blocks were frozen
(0.5-1.0 out
a
(freshly
fixative pieces
anesthetized
formaldehyde
excised
buffer,
on copper
was carried
MT-2B
tissue
phosphate
for semi-thin
sections
a Sonvall
were perfused
3%
in the same The
with
were
heart of
mm.
pieces mounted
gen.
ofthe
and
microscopy.
Leghorn)
fixatives
a mixture
immersed
was 60
in 0.1 M sodium
week. Tissue
using
or
and
time
(White
electron
used were 3% formaldehyde
Perfusion-fixed
small
hens
microscopy
(7) for
thomacotomy,
the left ventricle
mmHg. The fixatives from parafommaldehyde)
Total
Adult
Mtem
inserted
(3) for fluorescence
(iron-dextran)
ofTissues.
pentobarbital.
cannula
fluomescein Imposil
to a
nitmo-
(0.05-0.
10
of Tokuyasu, (31,32).
Immunofluorescence Microscopy. Semi-thin frozen sections mounted on glass slides were immunolabeled according to methods described (34). ftir tissues fixed with glutaraldehyde, sections were treated with 0. 5 mg/mI sodium bomohydride to quench the fluorescence caused by the fixative (23). Sections
were
observed
with
a Zeiss
photomicroscope
III (Carl
York) equipped with epifluomescent illumination ski objective lens. Photography was carried out with Microscopy.
Immunoelectron using (31),
ferritinexcept
and that
Ultra-thin
Imposil-conjugated pre-treatment
sections
secondary of sections
Zeiss
Inc.,
and a 63 x NomanKodak Tmi-X film.
New
were
immunolabeled
antibodies
was performed
as described with
a mixture
of 1% normal
goat serum and 2% dextran (Mw 10,000). Labeled sections were adsomption-stained and embedded in methyl cellulose as described
4
Figure 1. (a) Nomarski and (b,c) double indirect immunofluorescence micrographs of cardiac muscle. Semi-thin frozen sections were doubly immunolabeled with rabbit anti-desmin(b)and guinea pig anti-vimentin(c). Pericytes(arrowheads) are labeled for both desmin and vimentin. In contrast, endothelial cells (arrows) and fibroblasts (double arrowhead) are positive for vimentin but not for desmin. Note that cardiac muscle cells are labeled for desmin alone, but not for vimentin. Original magnification x 1500. Bar = 10 m.
Downloaded from jhc.sagepub.com by guest on July 11, 2011
DESMIN
AND
IN PERICAPILLARY
VIMENTIN
CELLS
1107
‘j’c
1;
-
Figure 2. Double indirect immunoelectron microscopy of a capillary in cardiac muscle. Desmin and vimentin are labeled by ferritin (arrowheads) and Imposil (arrows), respectively. Whereas Imposil is seen in the cell body of both the endothelium (E) and the pericyte (P), ferritin is observed only in the pericyte. L indicates the lumen of the capillary where a portion of an erythrocyte is seen. Original magnification x 75,000. Bar 0.1 tm.
(31),
and
ated
at 60 or 80 kV.
were
observed
with
a Philips
EM-300
electron
microscope
oper-
tin,
sections
mowheads entities
were
ample,
Results cardiac
indirect direct scribed
the presence
muscle,
of desmin
exocrine
pancreas,
vimentin
kidney,
immunofluorescence immunoelectron
and
microscopy.
and spleen
liver,
microscopy
and
Results
in capillaries
double
for
each
of
by double or single
tissue
are
many
de-
By contrast,
capillary
ic) were
sections
found
muscle observed
closely
immunofluorescence
thin
was sectioned as possible by
apposed
transversely in
a given
Nomarski
to endothelial microscopic
in order field.
microscopy,
cells labeling
In
to observe
semi-thin
small
(Figure
entities
la).
for desmin
as
frozen were
In double and
vimen-
coexistence by
frozen
is not visible
double sections
of desmin indirect
ritin
designating
ing
vimentin, Because
2).
by the present a flat
Because
were
of the
desmin,
both
observed
the
sections
were
neither in
and
Figures
was conultra-
apposition
penicytes.
particles
Fer-
designat-
distributed
in methyl
and
patchy.
membrane
close
to identify
Imposil
lb
of
basement
to
studied.
was often
microscopy
to be densely embedded
seemed
in pemicytes
the
in
relative
in all pericytes (arrows
cx-
the cap-
arrowheads
method, used
for
from
estimate
vimentin
embedding
profile
were
Downloaded from jhc.sagepub.com by guest on July 11, 2011
fibroblasts,
and the staining
and
(an-
these
(double
immunoelectron
(Figure
and
cytes.
cells
only,
proteins
away
possible,
finding
endothelial
to endothelia particles
is not
was a common
for vimentin
labeled
only
an accurate
proteins
both
localization,
of penicytes;
vimentin
Although
two
This
Muscle capillaries
of the
for
oftheim
to the cells farther
for
lc).
predominate.
below.
processes
labeled
in-
The
Cardiac
likely
positively
ic). Because
correspond
lb and
amounts
labeled
lb and
most
lumen,
Figures
firmed
Cardiac
invariably
probably
illany
We studied
were
in Figures
cellulose
in pen(31),
FUJIMOTO,
1108
individual
intermediate
filaments
thy, however, that another, whereas
the markers other areas
of either
By contrast,
marker.
for vimentin
(Figure
Eocrine
were
not
visible.
SINGER
It is notewom-
were always seen in proximity of the penicyte section were endothelial
cells
to one devoid
were
labeled
were
also
only
2).
Pancreas
Penicytes
in
for both
capillaries
desmin
of the
and
exocnine
vimentin
pancreas
by immunofluomescence
labeled
microscopy
(arrowheads composition
in Figure 3). The endothelial cells showed a different ofintermediate filament subunits as compared to those
of the heart.
Endothelial
cells of the exocmine
pancreas
were
for both desmin and vimentin, whereas those of the labeled only for vimentin (Figures 3b and 3c compared lb and
ic) (11). Immunoelectmon
finding
that
pancreas
both
penicytes
contain
desmin
and
microscopy
confirmed
endothelial
cells
as well
as vimentin
labeled
heart were to Figures the above
in the
exocmine
(photograph
not
shown).
Kidney Penicytes
of the penitubulan
to contain
both
capillary
desmin
immunoelectmon
and
in the renal
vimentin
microscopy
cortex
by separate
(Figures
4a and
were
shown
indirect
4b,
single
respectively).
In
this capillary, endothelial cells lack vimentin and contain only desmm (11). In the renal medulla, pemicytes apparently contain both desmin labeling
and vimentin, (arrowheads
cells
in the cortex,
tam
only
however,
vimentin
cells around gial
on the basis of double immunofluorescent in Figures 5a-c). In contrast to endothelial (arrows
the renal
cells,
were
endothelial in
glomerulan
labeled
for
cells
Figures
On
capillaries,
vimentin
in the medulla
5a-c).
but
the
podocytes
and
negative
for
were
con-
other
hand,
mesandesmin
(arrowheads, Figures Sd-f). The endothelium of the glomerular capillary was not noticeably stained for either of the two antigens (small arrows, Figures Sd-f), except for occasional vague staining for vimentin
(large
arrows
in Figures
5d-f).
Liver In
the
hepatic
and
Kupffen
mm
only,
sinusoid,
cells. and
the
the
We have latter
lumen
vimentin
tween those sinusoidal lining cells, a penivascular space called with
many
stoning
microvilli
cells)
the space
are
(37).
and vimentin, in the penisinusoidal areas
also doubly
Considering
that only
to run
labeled
cells
contains
cell
des6). Be-
the hepatic panenchymal of Disse is present. Along processes
of Ito
cells
to the sinusoidal
immunofluonescence
(fat-
wall
micrognaphs
for
in des-
for both proteins were found in Figure 6). Small discrete
were distributed they
endothelial
(11; see also Figure
parallel
cell bodies positive space (arrowheads
theirlocalization,
with
the former
cells and the space
of hepatocytes, observed
In double
mm
is lined
reported
are most
along
the sinusoidal
probably
the
cell
wall. bodies
4
Figure a Exocrine pancreas. (a) Nomarski optics and double indirect immunofluorescence micrographs for (b) desmin and (C) vimentin. Pericytes (arrowheads) as well as endothelial cells (arrows) are labeled for both desmin and vimentin. Original magnification x 1700. Bar = 10 m.
Downloaded from jhc.sagepub.com by guest on July 11, 2011
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Figure 4. Indirect single immunoelectron microscopy of the peritubular capiiiary in the renal cortex. (a) Desmin and (b) vimentin are marked The pericyte (P) is labeled for both desmin and vimentin, whereas the endothelium (E) is labeled for desmin only. Arrows indicate clusters of the capillary. Original magnification x 76,000. Bar - 0.1 sm.
Downloaded from jhc.sagepub.com by guest on July 11, 2011
I.
.3
by ferritin of ferritin.
particles. L, lumen
1110
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-.
:l_#{216}.A%%\. N.
‘
DESMIN
and
cell
AND
VIMENTIN
processes
of Ito
munofluorescence the
cell
and
in the
microscopy
tween
Labeling
are
cells
labeled
cell
the
processes. clearly
and
either
for
CELLS
two
was comparable
showed
endothelial
of Disse
cells.
microscopy
body
electron
IN PERICAPILLARY
panenchymal
for desmin
(Figure
7a)
that
both
inference
by im-
both
single
in
immuno-
cell processes
hepatic
ume 7b), confirming the above ent in the Ito cells.
proteins
in intensity
Indirect
that
1111
located
cells
in the
bespace
or vimentin
(Fig-
proteins
are pres-
and
particularly
dothelial
sinusoid cells
soidal wall membrane, cells
(6).
(littoral
cells).
are
for desmin
positively
a wide
not
found
labeled
due
mm
only.
soidal Figure
in the
for
suggest
that
to the
lumen,
farther
from
the splenic
the
one
found
vimentin.
The
the
other
(small
arrows,
hand,
some
of the
two
en-
ago.
However,
position
cord and
cells,
are mostly
labeled
vimentin.
These
mespectively.
ably represent
A few
phagocytosed
Figure
8), is labeled
extending
sinu-
the
one
closer
leculan
cell,
and
the
other,
those
for vimentin blood
labeled
for
or some
debris
Cells
only cells
desmin
or for and
only
stained
in me-
Double
indirect
sinusoidal
penisinusoidal tin
(Figure
immunoelectnon
endothelial
meticular 9).
As in the
with
are positive
cells are labeled penicytes
cmeas, and kidney (photographs markers, femmitin and Imposil, to one another, mamkem.
microscopy
cells
large
for both
ofheant
showing
only,
desmin
(Figure
not shown, were distributed areas
decisively
for desmin
but
abundantly
ities
conventional
cell-culture
cytochemistry
of intact
and tissues
few possible approaches to learn tume and, from such information, tions
of these We have
cells
is difficult
biochemical containing
is that
by combining
and vimen-
muscle,
in smooth
Further
muscle
and
support
for
impro-
penicyte
be
indicate
cytoskeleton with
explanation
may
cells (8). Therefore,
results
consistent
possible
penicytes
(fat-stoning
with
either
by
Immunoof the
microscopy
were
cell processes
pan-
about their molecular ultmastructo obtain insight into the funcimmunofluomescence
cells.
we have
desmin,
a contractile
for these
precursors
of
that
the mo-
closely
resemble
function
phenotypic smooth
for
similar-
muscle
cells
cells)
not
wrap
invested cells
mm
also found
in liver
and
penisinusoidal
are apposed
around within
to contain
to endothelial
the capillary the same
neticular
cells
in the
that
despite
these
of intermediate
lumen
basement
are wrapped; Ito cells A; splenic penisinusoidal
filament
possible that Ito cells the same developmental
both
desmin
neticulan
showed
cells,
cytoskeletal in penicytes
renal
proteins
do; they
in which
splenic
cord.
differences
Our they
proteins
results
have
myosin,
endothelial vitawith
have
shown,
the same
compo-
as penicytes.
It is therefore
cells of spleen and are probably
tmopomyosin,
does are not
contain many fat droplets stoning reticular cells make a meshwork
of liven and neticular omigin as penicytes,
(actin,
vimen-
the cell body
as penicytes
membrane
and
to perform similar roles. To further characterize these cells, be determined whether they have the same isofonms
cells. shown,
contractile
present
Another
Ito cells
whereas
is one
the case;
cells contain
tin. Although they exist adjacent to endothelial cells, with the basement membrane between, they are not generally called pemicytes. They are different from penicytes in several points: although the
to investigate cells
be always
but does
cells. Cells for contrac-
cells in spleen
see ref. 1 1), two in close proximity
techniques. these
are
with,
and
(20,24).
sition
of pemicapillany
not
two pro-
desmin
are contractile differentiated
endothelial
of the
of smooth
other
biology
is consistent
immunocytochemical
components
pemicytes.
prob-
Discussion cell
they
the accumulated
however,
The
that
this may some
ofthe
that both amounts.
nonspe-
2), exocnine
no labeling
clear
that
teins in penicytes, including muscle (as well as non-muscle) isoforms of actin (13), myosin (15), and tropomyosin (14). We have also demonstrated (not shown) that the penicytes ofcardiac muscle capillanes contain large concentrations of a-actinin and filamin, two
lines,
cell.
hand,
(11) that
com-
from
the notion ofpenicyte contractility, however, comes from recent munocytochemical studies that detected several muscle-specific
proteins
are probably
spots
for desthe
quantitation
in pemicytes
shown
whose
in culture
it appears equivalent
that penicytes are generally
arrowheads in These results
to the reticulan
material
the lu-
precise
ofdesmin
On the other
of cells
was altered
oftissues in situ was required to estabresult. Although the double-labeling
do not allow
presence
examples
filaments
not prove, the postulate known to contain desmin
it is not
cells,
into
from
(double vimentin.
endothelial either
(marked
of endothelial
we used
recently
cifically.
that
theme are several
teins present in a single pemicyte, vimentin are present in roughly
tion.
was further 8b and 8c)
it bulges
labeled
corresponds
wall
line which in Figures
med pulp desmin and
to the
lumen,
since
ofintemmediate
The
doubly
cnyo-ultramicro-
A brief report (5) that pemicytes and vimentin appeared several years
sinu-
were stained
where cells
parallel
When that
sinusoidal
cytoplasm
recognizable
corresponds the
desmin
ticulan
fusifomm
vessel. lines
8b), but only the (single arrowheads
to the nucleus On
this
around
wall into the surrounding 8) were labeled for both
both
with
endothelium,
were
is unambiguously
men
the
two parallel
for desmin
also
which
Outside
lined
and viinentin,
by a large arrow in Figure removed from the lumen was
lumen
is supported by the circumferential rib-like basement which further interacts with the processes of meticulam
Pemicytes
labeled
has
with
in their intermediate filaments. in culture contain both desmin
methods splenic
microscopy
in the capillaries of cardiac muscle, contain both desmin and vimentin
in vivo (10,22), examination lish the significance of this
Spleen The
immunoelectron
tomy of tissues, that pemicytes exocnine pancreas, and kidney
etc.)
are of able
it should of other
as are found
(13,14,15).
Among the glomerulan
microvascular capillary
beds we examined in this was exceptional, in that cells
Figure 5. Kidney. (a,b) Nomarski optics and double indirect immunofluorescence micrographs for (b,e) desmin and (c,f) vimentin. are labeled for both desmin and vimentin (arrowheads), whereas endothelial cells (arrows) are labeled for vimentin only. Original 10 sm. (d-f) Renal glomerulus. Mesangial cells (m) as well as podocytes (arrowheads) are labeled only for vimentin. Endothelial for either of the antigens (arrows). A thick arrow indicates the capsule of the glomerulus. Original magnification x 1100. Bar
Downloaded from jhc.sagepub.com by guest on July 11, 2011
study the in the vi-
(a-c) Renal medulla. Pericytes magnification x 1300. Bar cells are not labeled intensely = 10 sm.
=
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. ..
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,
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-
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.,,-.,, .
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8a
Figure 6. Liver. (a) Nomarski optics and double indirect immunofluorescence micrographs for (b) desmin and (C) vimentin. Endothelial cells with a flat narrow profile (arrow) and Kupffer cells with an irregularly shaped cytoplasm protruding into the lumen (double arrows) are labeled for desmin only and vimentin only, respectively. Ito cells (fat-storing cells) in a recessed position beneath the sinusoidal lining cells (arrowheads) are labeled for both of the antigens. Small spots doubly stained are also seen along the sinusoidal wall and are thought to be processes of Ito cells. Original magnification x 1000. Bar = 10 sm. Figure 8. Spleen. (a) Nomarski optics and double indirect immunofluorescence micrographs for (b) desmin and (C) vimentin. Endothelial cells of the splenic sinusoid are labeled for desmin only (arrows). In contrast, reticular cells (arrowheads) are labeled for both desmin and vimentin. In the immunofluorescence micrograph for desmin (b), two parallel lines of labeling, one for the endothelium and the other for the reticular cell, are observed (large arrow). The cell body of reticular cells is often seen to extend into the splenic pulp (double arrowheads). Original magnification x 1700. Bar = 10 sm. Downloaded from jhc.sagepub.com by guest on July 11, 2011
,
.
,
.1
*
I
.
s
.-
-1-
#
-.
..
.i
.
.:
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-
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-
Figure 7. Indirect single immunoelectron microscopy ofthe hepatic sinusoid. Imposil was chosen to mark either desmin (a) or vimentin (b) because ofthe presence of endogeneous ferritin in this tissue. Sinusoidal lining endothelial cells (E) are labeled for desmin only, whereas processes of Ito cells (fat-storing cells, FS) are labeled for both proteins. Clusters oflmposil particles are shown by arrows. L, lumen ofthe sinusoid; MV, microvilli ofparenchymal liver cells. Original magnification x 67,000. Bar = 0.1 tm.
Downloaded from jhc.sagepub.com by guest on July 11, 2011
FUJIMOTO,
1114
9.
Figure
Double indirect immunoelectron microscopy ofthe splenic sinusoid. Desmin and vimentin are labeled by ferritin and Imposil, respectively. particles are observed both in the endothelium (E) and the perisinusoidal reticular cell (A), but Imposil particles (arrows) are seen virtually only portion of an erythrocyte is seen in the lumen of the sinusoid (L). Original magnification x 75,000. Bar = 0.1 tm.
cinity do not and vimentin. have have
vimentin desmin
have intermediate Podocytes and
alone. In this capillary, (11). These observations
capillary
merular
may
have
pared to other capillaries; ties may be unusual. Finally, smooth otubes not
carefully
pursued
direct immunoelectron and Imposil particles these
large
areas
findings
tin in individual
reflect
functional
that
cells also do not the renal gb-
characteristics
their
as com-
contractility
S. Sobin
Note
Added
In a report
proper-
presence
and in developing to result in both
individual this were
of desmin
with
penicytes,
marker
was present.
the co-polymerization
intermediate
filaments
but
ofpenicytes,
We have
in double
2 and
in-
enties
1.
whereas
It is probable
that
and vimenas in the other
Adams
and Mr Michael
McCaffery
for their
retinal
Cancilla
for
giving
our thanks
us invaluable
to Drs
advice.
this
article
was
was obtained
penicytes
U
in press
Cell
for the contractile
in collagen
lattices
Biol prop-
in vitro.
Cited PF, Frank E, Holtzem H, Somlyo AP: The intermediate of rabbit vascular smooth muscle: immunofluorescent
and vimentin. RR, Vimtnup
3. Brandtzaeg fluorochromes. phy. Scand
ofthe Ms Margie
evidence
lanes. Anat
4.
Acknowledgments
while
clear
of bovine
Bensley
We also wish to express
Tokuyasu
published 1987),
Bernen proteins
KT
Dense ferritin in the latter. A
in Proof
of desmin 2.
assistance. and
Literature
9), femnitin
together,
ofdesmin
in
muscle mybeing com-
filaments.
microscopy (e.g. , Figures often observed clustered
neither
vimentin
skeletal proteins
intermediate
point
and
cases.
We wish to thank
technical
Sidney
104:483,
the simultaneous
of the same
in other
special
endothelial indicate
in particular,
muscle cells (12,25) (33) has been shown
ponents
excellent
filaments made of both desmin mesangial cells around the capillary
SINGER
J Muscle
Res Cell Motil
BJ: On the nature
2:439,
of the Rouget
filament studies 1981
cells of capil-
Rec 39:37, 1928 P:
J
Conjugates of I. Characterization
Immunol
PA, Baker
central
1972
Downloaded from jhc.sagepub.com by guest on July 11, 2011
RN,
nervous
2:273,
Pollock
system
immunoglobulin by anionic-exchange
G
with different chromatogra-
1973
PS, Frommes
to exogenous
SP: Reaction
protein.
of penicytes
Lab Invest 26:376,
DESMIN
AND
VIMENTIN
IN
PERICAPILLARY
1115
CELLS
positive cells from the rat aortic arch to the level of the artemia iliaca communis. Differentiation 20:196, 1981
5. D’Amore PA, Edelman D, Stark D, Rmm DM: Tissue culture and charactemization of microvascular penicytes. J Cell Biol 93:334a, 1983 6.
Dc Bruyn PPH, Cho Y: Contractile structures in endothelial splenic sinusoids. J Ultrastnuct Res 49:24, 1974
cells of
7. Dutton
A, Tokuyasu KT, Singer SJ: Iron-dextran antibody conjugates. General method for simultaneous staining oftwo components in high resolution immunoelectron microscopy. Proc Nati Acad Sd USA
8.
76:3392,
1979
Feramisco
JR.
and 9.
10.
K: A rapid
from
of a-actinin,
smooth
J
muscle.
filamin Biol Chem
MS. Rennels
Am
Franke
WW,
ML, Nelson
J Anat
149:47,
Schmid
H, Jackson
of expression
25
E: Ultrastructure
in mouse
DL,
BW, Illmensee
Winter
S. Jarasch
ED,
K: Differentiation-related
of intermediate-size
Harbor
13.
1, Singer
SJ: Immunocytochemical
in tissues
Biol XLVI:431,
studies
Fujimoto T, Tokuyasu KT, Singer SJ: Direct tion of vimentin and desmin in the same vascular smooth muscle cells. J Submicrosc
R,
26.
morphological intermediate Cytol 19:1,
14. Joyce NC, Haire Immunoperoxidase 1985
MF, Palade GE: Contractile localization of tropomyosin.
proteins
15. Joyce
ME,
proteins
Haire
Immunocytochemical graded 16.
17.
concentrations.
G, Joris
20.
21.
Contractile
RA, Prockop conjugates
Mazanet pericytes
J
Biophys
and
in pericytes. I. Cell Biol 100:1379,
in penicytes.
1977:
GC, Nelson in atherogenesis.
CB,
a review.
Schwarts New York,
Biochem
R, Franzini-Armstrong in rat red muscle.
Cytol
11:571,
In Chandler CJ, Wessler
Plenum
Press,
1977,
29.
30.
in 31.
RA,
Franke
79:3452,
32.
33.
The 169
34.
of
Movat HZ, Fernando NVP: The fine structure of the terminal vascular bed. VI. The venules and perivascular cells (pericytes, adventitial cells). Exp Mol Pathol 3:98, 1964
a gradient
XLVI:413,
fila-
1982
and immunocytochemical tubulin-containing
struc-
1982 ofmammalian
venous
veins.
J Ultrastruct
Res 25:452,
capillaries,
WW:
Heteropolymer
filaments
venules
1968 of vimentin
and
muscle tissue and cultured baby hamster by chemical cmosslinking. Proc Nail Acad
1982
E, Osbom
M, Rungger-Bnandle
Distribution
ofvimentin
tissue of mammalian
E, Gabbiani
and
G, WeberK,
Franke
in smooth
mus-
Exp Cell Res 137:329,
1982
desmin
and avian aorta.
filaments
GS, Croop J, Fellini SA, Holtzer H, Franke WW: Differential location of different types of intermediatesized filaments in various tissues of the chicken embryo. DiffementiaSchmid
E, Tapscott
15:27,
S. Bennett
1979
Simionescu
M, Simionescu
N: Ultrastmucture
Small
JV, Sobieszek
muscle.
In Stephens NL, ed. Biochemistry Raton FL, CRC Press, 1983, 85
A: Contractile
and
Tilton RG, Kilo C, WilliamsonJR, contractile function in rat cardiac tunes. Microvasc Res 18:336, 1979 Tokuyasu KT: Immunochemistry 12:381 1980
Tokuyasu tissues.
KT: A technique 57:551,
J Cell Biol
of the microvascular
wall:
structural
proteins
of smooth
Munch DW: and skeletal
of smooth
muscle.
Differences in penicyte muscle microvascula-
frozen sections.
ofultrathin
Vol I.
Histochem
fila-
in desmin
35.
36.
37.
for ultracryotomy
of cell suspensions
and
1973
Tokuyasu KT, Maher PA, Singer, SJ: Distributions ofvimentin mm in developing chick myotubes in v#{241}’o. II. Immunoelectron scopic study. J Cell Biol 100:1157, 1985
and
Tokuyasu KT, Maher PA, Singer mm in developing chick myotubes
and
J Cell Biol 98:1961,
1961
C: Scanning electron microscopy Micmovasc Res 23:361, 1982
of intermediate
K: Intermediate
Biol
J
I. The effect of histaan electron microscopic
Osborn M, Caselitz J, Weber K: Heterogeneity ment expression in vascular smooth muscle cells:
collecting
in vascular smooth cells demonstrated
Schmid
Boca
II.
AB, Eu5, eds.
G, Weber
Quant
functional correlations. In Renkin E, MichelCC, Geiger SR, eds. Handbook of physiology. Section 2. The cardiovascular system. Vol IV. Bethesda, MD, Am Physiol Soc. 1984, 41
of
DJ: Two improved methods for electron microscopy. J Cell
Majno G, Palade GE: Studies on inflammation. mine and serotonin on vascular permeability: study.
19.
GE:
J
muscle
Symp
Ultnastructure
Quinlan
tion 28.
demonstrafilaments 1987
evidence for the presence of two isomyosins J Cell Biol 100:1387, 1985
I: Endothelium
renius K, McMillan thrombotic process 18.
Palade
Kishida Y, Olsen BR, Berg for preparing ferritin-protein Biol 64:331, 1981
Majno
.
dc
cells
G, Sharp
Harbor
Cell Biol 24:98,
RhodinjAG:
WW:
and 1982
ofendothelial
Herman IM, D’Amome PA: Microvasculam pemicytes contain nonmuscle actins. J Cell Biol 101:43, 1985
NC,
Meth
kidney
patterns
filaments
Symp Quant
Moll
N, Shaw
Spring
Osbom M, Weber K: Immunofluorescence procedures with affinity purified antibodies:
Sci USA
E, Schiller
of proteins
ofpemicytes
in vivo. I. The presence of desmin only, or of desmin plus vimentin, or vimentin only, in the endothelial cells ofdifferent capillaries of the adult chicken. J Cell Biol 103:2775, 1986 12.
Cold
and small
1977
cells. Cold Spring
Fujimoto
M, Geisler
menu.
tures.
27.
11.
Osbom
desmin
heart.
cultured
protein
purification
1980
Forbes
Denk
23.
24. Burridge
a 130,000-dalton
255:1194,
22.
SJ: Distributions
ofvimentin
in vivo. I. Immunofluorescence
IH,
Bumnside
cells.
Invest
Wisse E: An electron lining
of rat
liver
Downloaded from jhc.sagepub.com by guest on July 11, 2011
des-
study.
1984
van Deurs B: Observations on the blood-brain barrier rats, with particular reference to phagocytic pemicytes. 56:65, 1976 Wallow dothelial
des-
micro-
B: Actin
filaments
Ophthal
Vis Sci 19:1433,
in retinal
in hypertensive
J Ultrastruct
pemicytes 1980
and
microscope study of the fenestrated endothelial J Ultrastruct Res 31:125, 1970
sinusoids.
Res en-