J. Biomedical Science and Engineering, 2011, 4, 164-172 JBiSE
doi:10.4236/jbise.2011.43023 Published Online March 2011 (http://www.SciRP.org/journal/jbise/).
Published Online March 2011 in SciRes. http://www.scirp.org/journal/JBiSE
Angiogenesis in rat uterine scar after introduction after
autological mesenchymal stem cells of bone marrow origin
Igor Maiborodin, Natalia Yakimova, Vera Matveeva, Andrey Shevela, Elena Maiborodina,
Ekaterina Pekareva, Olesya Tkachuk
Center of New Medical Technologies Institute of Chemical Biology and Fundamental Medicine, Novosibirsk, Russia.
Received 24 August 2010; revised 10 September 2011; accepted 13 September 2010.
ABSTRACT
The results of injecting of autologic mesenchymal
stem cells of bone marrow origin (AMSCBMO), trans-
fected by the GFP gene, into the scar of rat uterine
horns were studied by methods of light microscopy.
After the in t roduction of AMSCBMO in to the formed
scar on the right (2 months after the ligation) large
groups of blood vessels with cellular elements inside
were present; groups like that were not found in the
opposite side. Studying unstained sections under re-
flected ultraviolet light the sufficient bright lumines-
cence in the endothelium and the external membrane
of scar vessels was found in uterine horn only on the
side of introduction of AMSCBMO. It was concluded
that after the introduction of AMSCBMO into the
scar tissue they form blood vessels by differentiation
into endotheliocytes and pericytes. GFP gene expres-
sion not only in endothelium of vessels, but also in
their external membrane indicates that differentia-
tion of AMSCMBO is possible in endothelial and in
pericytal directions.
Keywords: Uterine Scar; Mesenchymal Stem Cells;
Angiogenesis; Endo t h eliocy t e s ; Peric ytes
1. INTRODUCTION
According to modern concepts, the physiological regen-
eration of tissues of the adult organism and their repair
in case of damage occurs with direct participation of
low-differentiated progenitor cells or stem cells. The
main source of stem cells is bone marrow, capable, in
addition to its basic - hematopoietic function , to generate
precursors of cellular elements of a large number of
body tissues.
Bone marrow contains two basic types of stem cells:
hematopoietic and stromal (mesenchymal) [1]. It is be-
lieved that both of these types of cells are precursors
with the potential to transdifferentiation into cells of
different ph enotypes [2].
In addition to bone marrow, pluripotent stem cells
were also found in other tissues of the adult body - the
adipose, muscle, brain, as well as in peripheral blood and
blood from cord/placenta. Moreover, it is established
that depending on a microenvironment the stem cells are
able to get through haemopoetic/mesenchymal barrier as
possess to high plasticity in case of a differentiation and
a transdifferentiation.
J. M. Isner [3] isolated supposed endothelial progeni-
tor cells from the bone marrow, some surface antigens of
these cells were common to hematopoietic stem cells
(CD34, CD133, Flk-1, Tie-2). These endothelial pro-
genitor cells were injected systemically to immunodefi-
ciency rats with pre-performed occlusion of coronary
artery. Afterwards was discovered that donor cells were
accumulated mainly in the areas of neoangiogenesis
within the myocardium. Increasing the number of capil-
laries per unit volume associated with a significant im-
provement in function and parameters of the left ventri-
cle compared with the control group. Similar work with
similar results was performed on various models by
other researchers [4-7].
H. Kamihata et al. [8] showed that intensive neoan-
giogenesis after implantation of bon e marrow cells in the
area of myocardial infarction was caused not only by
direct participation of these cells, but also by significant
expression of angiogenesis factors such as vascular en-
dothelial growth factor, angiopoetin and others by these
cells. M. Takahashi et al. [9] also supported the idea of
paracrine mechanisms of the effect of bone marrow cells
on the myocardium by inflammatory cytokines.
Since coronary heart disease is characterized by myo-
cardial scarring, the expansion of the cavities and dete-
rioration of the work of ventricles, which can be pre-
vented only by full restoration of the myocardium, the
studies were undertaken to assess the possibility of its
restoration on the model of ischemic heart disease in

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 165
mice. Cardiac progenitor cells were implanted in scar
zone. Also local stimulation by hepatocyte growth factor
and insulin-like growth factor-1 directly of the scar area
was performed. These two factors have important char-
acteristics, it is known that the hepatocyte factor is
chemoattractant for cardiac progenitor cells. At the same
time, insulin-like factor causes a strong cellular prolif-
eration and increases the viability of these cells [10].
The results of this research showed decrease of the
scar area by 42% with both methods of effects on the
myocardium, as well as improved ventricular volume
configuration. The reduction of scar area was associated
with the formation of new myocardiocytes in which car-
diac progenitor cells produce matrix metalloproteinases.
In the analysis of chromosomes of myocardiocytes was
shown that in the process of recovery of myocardium
there is no “merger” of cells. As an alternative to intro-
duction of stem cells the authors propose using a com-
bination of growth factors [10].
The impoverishment of the population of circulating
bone marrow cells with age - an important limiting con-
dition for cellular therapy, as a preliminary preparation
for the treatment in such cases it is suggested to use
small molecules, polymers, growth factors or their com-
bination [11].
In experiments on rats were showed that autologic
mesenchymal stem cells of bone marrow origin (AM-
SCBMO) which hypoxically prepared within 24 hours
before the transplantation into peri-infarction zone in-
crease the expression of factors of vitality and vascu-
logenesis: hypoxia-inducible factor 1, angiopoietin-1,
vascular endothelial growth factor, as well as receptors -
Flk-1 , erythropoietin, Bcl-2 and Bcl-xL, that eventually
leads to improved angiogenesis through increased para-
crine mechanisms [12].
We can conclude that in the literature on treatment of
scars with the use of cellular technologies, there are 2
main hypotheses about the destiny of implanted AM-
SCBMO. The first is that AMSCBMO in the continua-
tion of their differentiation in hypoxic tissues are directly
involved in angiogenesis and revascularization [4-7].
According to other researchers, the effect of AMSCBMO
is mainly due to the expression of various factors of an-
giogenesis and other cytokines [8,9,12]. M. Rota et al.
[10] believe that there is a combination of direct in-
volvement of stem cells with paracrine mechanisms. So,
there is no common opinion and until there is remained
not clear what happens to AMSCBMO after implanta-
tion into the tissues and organs of the body.
2. AIM
These days inflammatory diseases of female genitals and,
as a result of them, adhesions, lead to reproductive dis-
orders in women. The most relevant in this regard is
infertility. Studying peculiarities of etiopathogenesis in
the development of adhesions and synechiae in the uter-
ine cavity allows justifying the search of new directions
of their treatment and prevention.
Due to the low efficiency of widely used techniques
for treating scars of myometrium the attempt to correct
the pathology with the use of cellular technologies was
made. In addition, cu rrently remains unexplored the des-
tiny of stem cells after the introduction one way or an-
other into the organism.
An attempt to use autologic mesenchymal stem cells
of bone marrow origin (AMSCBMO) to accelerate the
regeneration of scar of myometrium in the experiment
was made due to the numerous reports on the effective-
ness of cellular technologies in the treatment of co ronary
heart disease [3-10,12]. Cellular-mediated strategies in
the treatment of this pathology are based on the implan-
tation directly into the ischemic myocardium or the
coronary vessels of bone marrow cells. This serves two
purposes: myocardial revascularization and eliminating
the deficit of functional cellular elements in myocardium.
3. MATERIAL AND METHODS
In our research the female Wag-rats weighing 180-200 g
at the age of 6 months were used as models. In aseptic
conditions lower midline laparotomy was performed.
Uterine horns were taken out into the wound and care-
fully rounded by sterile gauze. At the end of each horn
near the corpus uteri were placed ligatures from catgut
and bandaged up in that part. Abdominal cavity was su-
tured tightly layer-by-layer.
Isolation of mesenchymal stem cells: AMSCBMO iso-
lated by washing out the bone marrow from femur
epiphysis that taken under ether anesthesia in male rats,
inbreeding line Wag. Obtained suspension of cells was
placed in plastic bottles (“Nunk”, Denmark), in 48 hours
after explantation of bone marrow the cells that did not
attach were removed. Attached cells were cultured in
α-MEM medium with supplement of 10% fetal calf se-
rum (“Biolot”, Russia) at 37˚C in СО2-incubator with
5% СО2 in saturated humidity. The culture medium was
changed every three days. Performing subculturing the
monolayer culture was dispersed with density of 1000-
5000 cells/sm2 (depending on the growth properties of
used fetal serum), were used standard solutions of Versene
and tripsin.
Transfection: Mesenchymal AMSCBMO from second
passage, obtained from rats of mentioned line, trans-
fected DNA of plasmid pEGFP-N1 (Clontech Laborato-
ries), which contained the gene of green fluorescent pro-
tein (GFP) under the control of cytomegalovirus pro-
moter and neomycin resistance gene under the control of

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 166
the promoter of the virus SV40, that necessary for the
subsequent selection using geneticyn G418 (pEGFP-N1;
Clontech Laboratories). Transfection was performed in
the presence of the reagent for transfection TurboFect
(Fermentas) as recommended by the manufacturer; the
protocol for transfection of cellular suspension was used.
Transfection was perf ormed using 1x106 cells in 1 ml of
the suspension, 4 mg plasmid DNA and 10 ml of reagent
for transfection (TurboFect). Cells after transfection
were cultured in α-MEM medium with supplement of
10% fetal calf serum (Biolot, Russia), using a selection
of G-418 (Sigma) at a concentration of 400 ng/ml.
As specific unique markers for AMSCBMO are un-
known, for identification of these cells were used physi-
cal, morphological and functional properties. Untrans-
fected and transfected cells (for demonstration that of
AMSCBMO properties after transfection remain without
changes) were characterized by the following physical,
morphological and functional properties which are pecu-
liar for mesenchymal stem cells cultured in vitro: at-
tachment to the surface of the cultivation, morphology,
proliferation, formation of colonies of fibroblast-like cells
and the ability to ind uced differentiation in bone tissue.
Using methods of light and fluorescent microscopy
and cytological staining methods was showed that the
cultured untransfected and transfected rat bone marrow
cells in vitro: were attached to the surface of cultivation,
had fibroblast-like morphology continued at all times of
cultivation, maintained in culture fo r several subcultures,
formed colonies of fibroblast-like cells when were dis-
persed at low density, in th e presence of a linear-specific
factors differentiated into cells of the bone tissue. For
research the untransfected cells of 0-3 passages and
transfected cells of 0-2 passage were used.
However, the physical, morphological and phenotypi-
cal characteristics are not unique criteria that can be used
for specific identification of AMSCBMO. AMSCBMO
ability for induced differentiation in vitro into bone,
adipose tissue and cartilage is the only critical requirement
for determining of prospective populations of stem cells.
Induction of osteogenic differentiation of mesenchy-
mal stem cells in vitro: For induction of osteogenic dif-
ferentiation the 0.1 μM desoximethasone, 50 μM ascor-
bic acid and 10 mM β-glycerophosphate (Sigma) was
used. As AMSCBMO have properties to differentiation
into cells of bone tissue, the cond itions of its implemen-
tation are most reproducible. Such differentiation is
typically used to characterize the cultures of stem cells
in vitro and is typical way by default for the majority of
AMSCBMO in culture.
Osteogenic differentiation was determined by two
markers: the alkaline phosphatase activity and miner-
alization of extracellular matrix by calcium ions: Cyto-
chemical detection of alkaline phosphatase was per-
formed by using nitrotetrazolium blue in the presence of
substrate for alkaline phosphatase of 5-bromo-4-chloro-
3-indolyl. Accumulation of calcium in the extracellular
matrix was recorded by alizarin red staining.
In 4 hours after the transfection the cells were diluted
with untransfected cells at a ratio of 1:2.5, respectively,
and 0.1 ml of the mixture was injected during procedure
of relaparotomy and after removal of residual unlysed
suture material into the region of formed scar within 2
months after ligation of right uterine horns of female rats
Wag.
Remaining cells after transplantation cultured for 10
days to evaluate the efficiency of transfection and stabil-
ity of gene expression. Expression of introduced gene
GFP by rat mesenchymal AMSCBMO was assessed
visually under a fluorescent microscope, directly looking
at culture in 48 hours after the transfection. The effi-
ciency of transfection was assessed as a percentage of
luminescent cells relative to all cells. The percentage of
transfected cells in the diluted culture was about 3%.
As in most cases, using technology based on the in-
troduction of plasmid DNA was observed only a tempo-
rary expression of the introduced GFP gene in rat AM-
SCBMO. Cultivation of cells, transfected by plasmid
pEGFP-N1, without selection (not using geneticyn
(G418, Sigma)) showed a decrease in the number of
cells synthesizing a green fluorescent protein, as a result
of their replacement by untransfected cells. Nevertheless,
after 1 week in culture of transfected cells of first pas-
sage, seeded with density of 5000 cells per 1 cm2, were
observed cells that synthesized green fluorescent protein.
Thus, as in most cases, using technology based on the
introduction of plasmid DNA with cationic lipids, with-
out subsequent selection for creation of stable clone, was
obtained culture of rat AMSCBMO temporarily ex-
pressin g gr e en fluore s cent protein.
Fragments of the uterine horns with scar tissues and
synechiae, biopsied after 1, 2, 3 and 4 weeks after the
introduction of AMSCBMO, were preserved in a 4%
paraformaldehyde on biphosphate buffer (pH 7.4) for at
least 24 hour, dehydrated in a gradien of ethanol, light-
ened in xylene and embedded in paraffin. Unpainted
microscopic sections 5-7 microns thick were studied
under a light microscope Axioimager M1 (Carl Zeiss,
Germany) with a magnification of up to 1500 times in
the luminescence mode with a filter Alexa 488. The scar
of left uterine horn in a corresponding time after the in-
troduction of a suspension AMSCBMO and myometrium
scar of animals 2 months after ligation of uterine horns
with no subsequent use AMSCBMO were used as the
controls. 6 animals were examined at each point of the
experiment.

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 167
All studies were performed in compliance with “Rules
for work using experimental animals”, all manipulations
were performed under general anesthesia by ether inha-
lation.
4. RESULTS
After 2 months there was detected a moderate develop-
ment of adhesions in the pelvic area without involve-
ment of the upper part of abdomen. There were discov-
ered ex pressed changes in the uterine horn s, which were
manifeste d by presence of hydrometra above the place of
ligation to the isthmic part of uterine tube, probably due
to lack of outflow into tube and further into the ab domi-
nal cavity. The content in the uterine horn was transpar-
ent, serous, without the presence of purulent elements.
At this time in place of ligation of rat uterine horns the
encapsulated suture materials were present, despite the
data on the timing of resorption of catgut in 15-20 days
in humans. It should be noted that we have already
drawn attention in previous pub lications at later dates of
resorption of “lyzed” suture material [13].
In the time of study of the uterine horn structure, using
magnification in 6-10 times, the formation of synechiae in
the site of ligation, hypertrophy and stretching of the
uterine wall, formation of complete obstruction were
discovered. In the area of narrowing of the uterine horns
the atrophy of the endometrium and myometrium was
found.
In 1 week after the introduction of AMSCBMO into
formed scar on the right side there were large groups of
blood vessels with cellular elements inside (Figure 1),
the such groups of vessels were no t found in the scar on
the left side.
Figure 1. Groups of blood vessels in a scar of right rat uterine
horn in 1 week after application of AMSCBMO, in the many
vessels the luminescence of accurately outlined endothelium
and an external membrane is present. Magnification 240 ×.
In the study of unstained sections of uterine scar in the
reflected ultraviolet light sufficiently the bright lumi-
nescence in the endothelium and the external membrane
of scar vessels of the right uterine horn was found (Fig-
ure 1). Also in the endometrium and myometrium the
slight edema and many small luminous objects, most
likely, vessels of capillary type were found (Figure 2).
In 2 weeks after the introduction of AMSCBMO the
very bright luminescence in the endothelium and adven-
titia of large vessels in scar of the right uterine horn was
discovered. The endothelium and the external membrane
of such vessels were represented in the form of bright,
clear, well defined lines (Figure 3). In the myometrium
Figure 2. In 1 week after introduction of AMSCBMO in en-
dometrium and myometrium of the right uterine horn the set of
small objects with luminescence (vessels of capillary type) and
the signs of interstitial edema are present. Magnification 240 ×.
Figure 3. In 2 weeks after injection of AMSCBMO the enough
bright luminescence is revealed in endothelium and adventitia
of large vessels in scar of the right uterine horn. Vessel mem-
branes with luminescence are presented by accurate, well lim-
ited lines. Magnification 630 ×.

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 168
and endometrium of the left uterine horn at this time the
luminescent structures were completely absent (Figure 4).
After 3 weeks only in the wall of some blood vessels
on the right was noted weak luminescence, almost at the
level of background (Figure 5).
By week 4 the luminescence intensity of the vascular
walls and the number of vessels in the structures, in
which was marked luminescence, was even more re-
duced (Figure 6).
On the opposite side for all periods of study the lumi-
nescence of the vascular walls were at the level of back-
ground, is necessary to repeat that large groups of ves-
sels were absent. In the myometrium and endometrium
small luminous objects were absent, but at the same time
Figure 4. Absence of structures with luminescence and signs
of edema in endometrium and myometrium of right uterine
horn in 2 weeks after use of AMSCBMO. Magnification 240 ×.
Figure 5. The weak luminescence, practically at background
level, is present in a wall of some blood vessels in 3 weeks
after introduction of AMSCBMO. Magnification 240 ×.
there were no signs of edema (Figures 7-11). In the scar
of the uterine horn of animals without the AMSCBMO
introduction (control) the luminescence objects were
absent in vessels, in the structure of the scar and in sur-
rounding tissues (Figure 12).
Unfortunately, after using AMSCBMO the more rapid
resorption of the scar and reduction of hydrometra were
not found. But, this pr eliminary study was conducted on
a small number of observations. For accurate conclu-
sions about the effectiveness of this procedure reg arding
the status of the scar there is necessary to repeat this
experiment with larger number of animals and to ob serve
longer periods after the introduction of AMSCBMO.
Figure 6. In 4 weeks after application of AMSCBMO the weak
luminescence remains in a wall of only individual vessels.
Magnification 240 ×.
Figure 7. In 1 week after introduction of AMSCBMO in a scar
of rat left uterine horn the number of vessels is much less, a
luminescence of their structures is practically at background
level. Magnification 320 ×.

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 169
Figure 8. In left uterine horn the objects with luminescence
and signs of edema in 1 week after injection of AMSCBMO
are absent. Magnification 240 ×.
Figure 9. The absence of signs of edema and structures with
luminescence in vessels of a scar of left uterine horn in 2
weeks after use of AMSCBMO. Magnification 240 ×.
5. DISCUSSIONS
Rat uterine horn is a thin tube, one end of which con-
nects with the uterine cavity, the other - with the fallo-
pian tube, opening into the peritoneal cavity near the
ovarian surface. Normally in mammals there is outflow
of content from fallopian tubes into the peritoneal cavity
with subsequent resorption by peritoneum. After ligation
of uterine horns in this section takes place ischemia fol-
lowed by necrosis, scar and hydrometra formation.
These changes are clearly seen in macropreparation with
a slight magnification.
Concerning the results of use of AMSCBMO it is nec-
essary to pay attention to the development of the vascu-
lar net at the injection site (Figures 1 and 3-5). The
proof, that these groups of vessels were formed precisely
as a result of AMSCBMO and from them, is the lumi-
nescence of walls of these vessels (Figures 1 and 3-5).
GFP gene, introduced in the DNA of AMSCBMO,
transfers without changes to daughter cells and cells of
next generations. These cells and structures formed from
them are also shone in the reflected ultraviolet light.
Until recently by prevailing dogma was that the an-
giogenic response, which develops in postnatal life, oc-
curs because of the growth of existing capillaries. How-
ever, now it is convincingly proven that a small, but
biologically significant portion of endothelial cells in-
volved in the formation of new capillaries, have bone
marrow ori gin [1 4- 1 6] .
Figure 10. In 3 weeks after injection of AMSCBMO the ob-
jects with luminescence are absent in structures of left uterine
horn and surrounding tissues. Magnification 180 ×.
Figure 11. Absence of a luminescence in vessels of scar of left
uterine horn in 4 weeks after introduction of AMSCBMO.
Magnification 480 ×.

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 170
Figure 12. A control animal without introduction of AM-
SCBMO. The objects with luminescence in scar structures of
uterine horn completely are absent. Magnification 480 ×.
Endothelial cells divide every three years, but in the
case of retinal vessels - every 14 years. However endo-
theliocytes can be induced to replication as response to
physiological or pathological stimulation. In some cases,
bone marrow populations of endothelial progenitor cells
can be completely replaced every five days. This is a real
advantage when the physiological circumstances require
increased blood supply, both in healing wounds, and in
preparation of a fertilized egg for th e implantation in the
richly vascularized endometrium [17].
In 1962 was first demonstrated the possibility of exis-
tence of circulating endothelial cells. It was shown that
on Dacron patch, which was isolated from contact with
the surrounding tissues and was located in the wall of th e
current vascular prosthesis of the aorta (experimental
prosthetics of the pig thoracic aorta), con nective tissue is
forming. After 14 days and later after implantation of
Dacron, there was determined a layer of endothelial and
other cells, including mononuclear cells, fibroblasts, as
well as giant cells of foreign bodies. Detected cells could
have only one source—the circulating blood [18,19].
In experiments on dogs was shown that in a few days
on the surfaces of endothelial impermeable vascular
prostheses the isolated islands of endothelial cells are
appeared, as when Dacron grafts was installed in inferior
vena cava and as in thoracic aorta. This also confirms
that the source of endothelial cells - circulating blood
[15]. These results were confirmed when after total irra-
diation of dogs the bone marrow transplantation from
animals of another kind was made, in which the donor
DNA of circulating endothelial cells were clearly differ-
ent from the DNA of recipient cells [16].
Blood vessels are forming by two interacting cellular
types. They are endothelial cells lining the inner surface
of the bloodstream and perivascular cells, named peri-
cytes, enveloping the outer surface of the vascular tubes
[20]. Pericytes are not only included in the hemody-
namic process, but also play an active role in the forma-
tion of blood vessels. Some researchers hypothesize that
endothelial cells and pericytes have common precursor
with hematopoietic cells. This hypoth esis is based on the
fact that developing hematopoietic and endothelial cells
have common surface markers and that hematopoietic
cells can be developed from embryonic cells of major
blood vessels [21-23].
Most commonly origin of pericytes associate with
mesenchymal stem cell [24]. After the initiation of dif-
ferentiation the progenitor of pericytes chemotactically
adhere to endothelial cells in the capillary plexus in the
time of beginning of angiogenesis [25]. In addition, there
are reports that pericytes can be generated from endothe-
lial cells during their transdifferentiation. This was dis-
covered in the posterior aorta [26], as well as in heart
valves [27].
The luminescence of the endothelium and the external
membrane of the vessels, discovered in our studies
(Figures 1 and 3-5), indicates that AMSCBMO directly
rather than indirectly (through cytokines or other cellular
signals), were involved in the formation of blood vessels,
possibly because of the presence of pluripotent cells
which already were stimulated to differentiate into en-
dothelial or pericytal directions. It is also possible that
tissue hypoxia, as a result of ligation of structures to-
gether with the vessels, stimulates the differentiation of
introduced AMSCBMO into endotheliocytes [12].
Additional evidence of angiogenesis, as a result of ap-
plication of AMSCBMO, is the groups of vessels in the
scar that were found only on the right – the side of
AMSCBMO introduction (Figures 1-6), but not in the
other side (Figures 7-11) and not in the control (Figure
12).
It is necessary to attract attention to another aspect of
the findings. Expression of introduced GFP gene not
only in the endothelium of blood vessels, but also in the
external membrane (Figures 1 and 3), is most likely a
sign that endotheliocytes and pericytes have the same
progenitor cells, or that differentiation of AMSCBMO is
possible as well as in endothelial and in pericytal direc-
tions. We have already pointed out that, in the opinion of
many researchers, endothelial cells and pericytes have
common precursor with hematopoietic cells [21-23].
Besides that, the unilateral development of blood ves-
sels and the presence of luminescence objects in walls
(Figures 1 and 3-5) indicate that in this case introduced
AMSCBMO do not migrate from the site of injection, do
not destroy, and they are not only “building material” or

I. Maiborodin et al. / J. Biomedical Science and Engineering 4 (2011) 164-172 171
“signal” to the beginning of angiogenesis for own cells
of the donor organism . In all these cases, of course, it is
possible that luminescence protein and transfected GFP
gene leave the destroyed AMSCBMO and then are ab-
sorbed by neighbor cells. However, proteins and DNA
fragments that got into cells by way of phagocytosis or
pinocytosis, degraded with the loss of ability to lumi-
nescence and insertion into the genome. So, when AM-
SCBMO are destroyed, intensity o f luminescence should
be minimal (the protein GFP leaves AMSCBMO) and
very quickly should entirely disappear; more so in this
case should not be clearly defined and limited luminous
structures.
The decrease of light intensity in the vessels of the
scar in the uterine horn after introduction of AMSCBMO
with time, probably due to the gradual restoration of
genome of transfected cells or the displacement of AM-
SCBMO by own cells.
Attention should b e paid, that the reduction of number
of objects, which can synthesize green fluorescent pro-
tein, was showed during cultivation of cells, transfected
by plasmid pEGFP-N1 without selection, as a result of
their replacement by untransfected cells.
In conclusion, it should be noted that after increasing
the number of vessels, especially “young” with thin
walls, metabolic processes in tissues with scar improve.
As a result of optimizing the conditions of life and func-
tioning of fibroblasts the exchange of components of
extracellular matrix of scar connective tissue may inten-
sify, rejuvenation of collagen and elastin fibers may oc-
cur, that will further manifest by appearance of thinner
structures, ordering of their location [28] and passable-
ness of uterine horns with formed adhesions can be re-
stored.
6. CONCLUSIONS
1) After introduction into uterine scar of AMSCBMO the
increase of number of vessels due to the processes of
neoangiogenesis was detected. In this case the AM-
SCBMO do not migrate and were not destroyed in the
site of introduction, but form the blood vessels by dif-
ferentiation into endotheliocytes and pericytes. In 1
week in rats after introduction of AMSCBMO the ves-
sels formed from these cells are already fully functional,
consist of all the structural walls and contain blood cells.
2) Expression of GFP gene not only in the endothe-
lium of blood vessels, but also in their outer membranes
indicates that differentiation of AMSCBMO in endothe-
lial and in pericytal directions is possible.
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