Light and Electron Microscopic Studies on the Effect of a Contraceptive Drug on the Stomach of Mouse
Fatma A. Elhumeed
Wafaa B. Yousif
The present study was an investigation of the effect of the contraceptive drug, Nordette, on the stomach of the mouse when administered daily at a recommended therapeutic dose rate of 0.0026 mg kg-1 for 30 days. Extensive light and electron microscopic changes were noticed. The drug caused enlargement in the all types of cells. The oxyntic cells appeared hypertrophied with irregular cell boundaries, enlarged nuclei and faintly stained cytoplasm. Their cytoplasm contained irregularly distributed mitochondria with dense matrix, decreased rER, obviously increased sER, disorganized intracellular canaliculi and some lysosomes. The peptic cells appeared enlarged and contained hypertrophied rough endoplasmic reticulum and an increase amount of ribosomes and secretory granules. There was an increase in the amount of the secretory granules in the lumen of the gastric gland. The mucus cells at the upper region of the gastric gland were greatly decreased. Smooth muscle fibers showed enlargement and degeneration. The submucosa and lamina propria showed vacuolation. The most pathological effects were restricted to the obvious decrease of the lymphoid cells in the submucosa and lamina propria. Dilatation and congestion of the blood vessels and blood capillaries were noticed. Blood capillaries lined by enlarged endothelial cells containing enlarged heterochromatic nuclei.
Estrogen has various physiological functions such
as growth and differentiation, via not only endocrine but autocrine or
paracrine fashion (Simpson et al., 1999). Though the ovary is a
major source of systemic estrogen, estrogen is produced also in extraovarian
tissues including, adipose tissue (Ackerman et al., 1981), skin
(Leshin et al., 1981), osteoblasts (Purohit et al., 1992),
testis (Brodie and Inkster, 1993), brain (Lephart, 1996), vascular smooth
muscle cells (Harada et al., 1999), T cells in the spleen (Samy
et al., 2001). These extraovarian tissues, however probably do
not produce a sufficient quantity of estrogen to affect the circulatory
level. Recently, Ueyama et al. (2006) stated that under normal
physiological condition a large quantity of estrogen equivalent to ovary
is secreted from the unidentified region of the gastrointestinal organs
into the portal vein and most of it is then trapped in the liver, but
in the case of increased portal systemic shunting it overflows into the
systemic circulation resulting in the elevation of systemic estrogen concentration
to cause estrogen-excess signs associated with liver cirrhosis. Patients
with liver cirrhosis often show an increase of serum estradiol (E2)/testosterone
ratio, which is supposedly caused by an abnormal estrogen metabolism in
the liver. Gene expression of aromatase, estrogen synthetase, was demonstrated
in the gastric mucosa of male and female adult rats, Parietal cells exhibited
strong signals for aromatase mRNA and immunoreactive protein by in
situ hybridization histochemistry and immunohistochemistry. These
findings indicate that gastric parietal cells play a potent endocrine
role in secreting estrogen that may function as a regulator of the gastro-hepatic
axis. These data showed that the gastric mucosa can produce a large quantity
of estrogen equivalent to that in the ovary (Ueyama et al., 2006).
Previous studies showed that experimental portal-systemic shunting in
adult male rats resulted in a marked increase of systemic estrogen concentration
(Farrell et al., 1988). Matsuyama et al. (2002) studied
expression Estrogen Receptor (ER) beta in human 29 stomach adenocarcinoma
and stated that the effects of estrogen in stomach cancer, as well as
those in normal stomach may be mediated by ER beta.
Jiang et al. (2006) stated that exposure to estrogens
is associated with an increase risk of developing breast, cervical and
liver cancer. Estrogens strongly induce the human granzyme B inhibitor,
proteinase inhibitor 9 (PI9). Cytotoxic T lymphocytes (CTLs) and Natural
Killer (NK) cells use the granzyme pathway to induce apoptosis of target
cells. Yousif (1997) and Yousefi et al. (1988) reported that sex
hormones including estrogen, progesterone and testosterone are known to
have adverse effect on immune system and particularly on the proliferative
Humans are routinely exposed to bisphenol-A (BPA), an
estrogenic compound that leaches from dental material, food and beverage
container and other plastic consumer products. Vandenberg et al.
(2007) showed that BPA induced adverse effect in mammary glands development
in the embryo. Correspondence to Stallard et al. (2000), hormone
replacement therapy is being used increasingly. Although it is known that
the risk of developing breast cancer is slightly increased with long term
use (Gambrell, 1992).
Li et al. (2006) studied how the estrogen modulates
the rigidity of the cytoskeleton in the epithelial cells; estrogen depletion
decreased and treatment with 17β-estradiol increased deformability
of cervical-vaginal epithelial cells. Estrogen also induced redistribution
of nonmuscle myosin II-B(NMM-II-B); lesser interaction of NMM-II-B with
actin; increased phosphorylation of NMM-II-B heavy chains and decreased
filamentation of NMM-II-B in vitro. Similarly, Gorodeski (2007)
tested the hypothesis that estrogen control epithelial paracellular resistance
through modulation of myosin and stated that estrogen modulates nonmuscular
myosin (NMM)-II-B, the main component of cortical actomyosin in human
epithelial cervical cells.
This study aims to examine the histopathological and
the ultrastructural changes in the stomach of mice treated with oral therapeutic
doses of a contraceptive drug.
MATERIALS AND METHODS
The experiment was conducted at the Faculty of Science
laboratory, Alexandria University, Egypt. Female Swiss mice (Mus musculus)
(from the animal house of the High Health Institute) of 8 weeks old and
20-22 g in weight were used in this study. Food and water were provided
ad libitum. Nordette tablets (Levonorgestrel 0.15 mg and Ethyl
estradiol 0.03 mg, manufactured by The Nile Company for Pharmaceutical,
Cairo) were uniformly suspended in distilled water. The animals were treated
daily intragastrically via a bent stainless steel feeding tube with a
therapeutic dose of 0.0026 mg kg-1 of Nordette for 30 days.
The dose used in the present study was the recommended therapeutic dose.
A group of female mice of similar age and weight was as control and received
no treatment. Small slices of stomach were immediately fixed in 2.5% glutaraldehyde
buffered with phosphate buffer (pH 7.6) for 1 h and post fixed in 1% osmium
tetraoxide (in the same buffer) for 1-2 h at 4°C. Specimens were dehydrated
through graded series of ethyl alcohol and embedded in Araldite-Epon mixture.
Semithin (1 μm) and ultrathin (50 nm) sections from selected areas
were cut with a glass knife on LKB ultramicrotome. Semithin section were
stained with Toluidine Blue (TB) and examined with light microscopy, while
ultrathin section were double stained with uranyl acetate and lead citrate
and examined by Jeol 100 CX Electron.
Light microscopy of control stomach: The gastric mucosa consists
of a surface epithelium that invaginated to varying extend into the lamina
propria, forming pits. Emptying into the gastric pits are long straight,
branched tubular glands. The lamina propria of the stomach, which extend
between the glands is composed of loose connective tissue interspersed
with blood and lymph vessels, smooth muscle fibers and lymphoid cells
(Fig. 1- 4). The lamina propria supported
the gastric glands (Fig. 1- 4). The
submucosa is relatively loose and distensible and the larger blood vessels
and a lymphoid aggregate lie in it (Fig. 1, 2).
Separating the mucosa from the underlying submucosa is a continuous layer
of smooth muscle, muscularis mucosa (Fig. 2). The stomach
has a well developed muscularis mucosa. Muscularis mucosa lying immediately
beneath the base of gastric glands clearly demarcating them from the underlying
submucosa (Fig. 2). Stips of muscularis mucosa may closely
associated with gastric glands and the contraction
of stomach of control mouse, showing oxyntic cells (Ox) peripherally
in the gland, small peptic cells (P), smooth muscle fibers (mf),
lymphoid cells in submucosa (sm) and lamina propria (arrow), nerve
plexuses (*) toluidine blue (TB) (x400)
of stomach of control mouse, showing peptic cells (P) with basal
nucleus and apical refractile secretory granules, oxyntic cells
(Ox), lymphoid cells in submucosa (sm) and lamina propria (lp),
muscularis mucosa (arrow) TB (x1000)
of stomach of control mouse, showing blood capillaries (bc) under
thick, well developed mucus cell (Mc) layer, oxyntic cells (Ox),
peptic cells (P), lymphoid cells (arrow), smooth muscle fibers (head
arrow) TB (x400)
of stomach of control mouse, showing many large blood capillaries
(bc) under surface mucus cell (Mc), oxyntic cells (Ox), few secretory
granules in gastric gland (arrow), lymphoid cells (head arrow) TB
of these smooth muscle cells may help gastric glands express their secretion
(Fig. 3). The muscularis consists of an outer thin longitudinal
and inner thick circular smooth muscle layers, it is composed of bundles
of smooth muscle fibers held together by loose connective tissue. Nerve
plexuses are seen between the outer and inner muscle layers (Fig.
The gastric mucosal coat appear thick due to the presence
of gastric gland. The gastric glands contain a mixed population of endocrine
cells and different type of exocrine cells, mucus cells, oxyntic cells
and peptic cells.
Mucous-secreting cells:The epithelium covering the surface and
lining the pit is a simple columnar mucus cells, all the cells secrete
mucin (Fig. 3, 4). The Mucous surface
cells are the most superficial cells in the gland. They extend a short
distance into the gastric gland. Mucus-secreting cells of the stomach
are not of the goblet form found in other parts of the body. Other mucus
secreting cells in the necks and ischmus portions of the gastric glands,
called neck mucous cells they secrete a less viscous mucus which may protect
the gland duct from autodigestion. Mucus neck cells are slightly different
cells, their mucus has different chemical properties from the mucus secreted
by mucous surface cells. Neck mucous cells are smaller and less regular
in shape than surface mucous cells, mainly because they are compressed
and distorted by adjacent cells. They have basal nucleus and finely granular
cytoplasm due to the presence of small mucin vacuoles (Fig.
Oxyntic (parietal) cells: Oxyntic cells are distributed along
the length of the glands but tend to be most heavily concentrated in the
upper and middle portions of gastric glands, where they are mixed with
peptic cells and stem cells. They are larger than the other cells of the
gastric glands. They can be recognized by their characteristic fried egg
appearance. They are located peripherally in the gland so their broad
bases appear to bulge into the underlying lamina propria. Oxyntic cells
vary slightly in their shape and size (Fig. 1- 4).
The much larger oxyntic cells are pyramidal, oval, or rounded cells, have
centrally located large spherical nucleus of low density and pale eosinophilic
cytoplasm which often appears vacuolated, particularly around the nucleus.
Their attachment to the basal membrane is broad, but their luminal aspect
is narrow being compressed between adjacent cells (Fig. 1-
4). The eosinophilic cytoplasm is due to the numerous mitochondria
(seen by electron microscope) which are a feature of highly metabolically-active
Peptic (chief) cells: Peptic cells are the principle cell types
in the basal third of the gastric glands. These cells appear small, irregular
pyramidal, or cuboidal in shape. Peptic cells are recognized by their
condensed basally located large nuclei and strongly basophilic basal cytoplasm.
They contain apical refractile large cytoplasmic granules (Fig.
stem cells are the precursor cells of all epithelial
cells of the gastric mucosa. They are small cells with oval basal large
nuclei and small amount of cytoplasm and show no cytoplasmic specialization
when completely undifferentiated. Occasionally, the enteroendocrine cells
can be identify in the section, characterized by their triangular shape.
Because of increase of the array of the subsurface capillaries, it seems
certain that virtually all surface mucus cells will be in close proximity
to this capillary net work (Fig. 3, 4).
Progressing towards the gastric lumen the amount of the blood capillaries
increased and become obviously enlarged, there was a notable decrease
in the amount of connective tissue between the basal lamina of the endothelium
and that of the adjacent gastric gland mucus cells (Fig.
Electron microscopy of control stomach:
Oxyntic cells: Oxyntic cell nuclei contain relatively few moderately
electron-dense hetrochromatin, many nuclear pores and obvious nucleoli
(Fig. 5). They have an elaborate system of apical surface
invaginations called intracellular canaliculi, consist of a branching
system of passages within the cell, opening at the free surface of the
cell to communicate either directly or via an intercellular canaliculus
with the lumen of the gland i.e., the luminal plasma membrane forms deep,
branching canaliculi which extend throughout the cytoplasm and between
adjacent cells, numerous short microvilli project into the lumina of intracellular
canaliculi greatly increasing the surface area. The canaliculi are closely
associated with mitochondria and an abundance of smooth endoplasmic reticulum.
These features facilitate the active transport of hydrogen ions across
a large concentration gradient in the resting cell. In survey micrographs
the ovoid, large mitochondria are the dominant feature of the cytoplasm
The cytoplasm possess a great number of mitochondria with closely packed
cristae. The mitochondria appear to be separated into peripheral group
and a central perinuclear group by a ring-like lighter zone of the cytoplasm
containing the intracellular canaliculi. The cytoplasm of the oxyntic
cells have a discrete, small Golgi apparatus near the cell base and some
rough endoplasmic reticulum (Fig. 5).
Peptic cells: Peptic cells predominate in the
lower region of the gastric glands. Their basophilia is due to the abundant
rough endoplasmic reticulum. Peptic cells have
of stomach of control mouse, showing oxyntic cell contain nucleus
(N) with moderately dense heterochromatin, nucleolus (Nu), nuclear
pore (head arrow), intracellular canaliculi (IC), microvilli (arrow),
mitochondria (M), rER, few sER, part of peptic cell with moderate
amount of rER (x7.500)
of stomach of control mouse, showing thin smooth muscle fibers contain
contractile protein, nucleus (N), mitochondria (M), thin and regular
endmycium (arrow) (x5000)
large basal nucleus containing large nucleolus and moderate amount of
heterochromatin. They contain large membrane bounded cytoplasmic secretory
granules and a rich rough endoplasmic reticulum, a prominent Golgi apparatus
and a usual number of mitochondria (Fig. 5). Smooth muscle
fibers were thin, containing elongated nucleus with moderately electron-dense
heterochromatin, contractile proteins, some mitochondria and thin endmycium
Light and electron microscopy of Nordette-treated stomach:
The changes induced by Nordette administration were
confined to gastric gland cells, smooth muscle fibers, connective tissue.
The connective tissue manifested obvious symptoms of lesions. These
stomach of Nordette-treated mouse, showing submucosa (sm) devoid
of lymphoid cells, oxyntic cells (Ox), peptic cells (P), degeneration
(arrow) in smooth muscle fibers (mf). TB (x400)
of the above micrograph, showing peptic cells (P) which highly basophilic
basally, an increase in the apical refractile secretory granules,
blood capillaries (bc) muscularis mucosa (arrow). TB (x1000)
pathological symptoms included marked vacuolation of the submucosa and
marked decrease in their cellularity. The most pathological effects were
restricted to the absence of the lymphoid cells in the submucosa and lamina
propria (Fig. 7- 10). Although, there
were many lymphoid cells in the submucosa and lamina propria of the stomach
of control animals the submucosa and lamina propria of the treated stomach
were nearly devoid of lymphoid cells. Smooth muscle layer of the treated
stomach showed an increased in their thickness. The muscle fibers appeared
enlarged and contained enlarged nuclei and some of the muscle fibers showed
obvious degeneration (Fig. 9, 10).
The light microscopic examination revealed an obvious change in the blood
vessels and blood capillaries including dilatation and congestion and
lined by enlarged endothelial cells
of stomach of Nordette-treated mouse, showing degeneration in smooth
muscle fibers (arrow), nerve plexuses (*), vacuolated submucosa
(sm). TB (x400)
of stomach of Nordette-treated mouse, showing degenerated smooth
muscle fibers (arrow), enlarged nucleus (N), submucosa (sm). TB
(Fig. 11). The histopathological changes of the stomach
oxyntic cells of treated animals showed some changes. Oxyntic cells appeared
hypertrophied with enlarged nuclei and faintly stained cytoplasm (Fig.
11- 13). Some cells possess ill definite cell boundaries.
Some oxyntic cells and peptic cells appeared irregular in outline. Peptic
cells appeared hypertrophid, with highly basophilic basal cytoplasm and
an increase in the amount of the refractile secretory granules in the
apical cytoplasm (Fig. 8). The gastric gland lumen contained
an increase amount of the secretory granules especially in the upper region
of the gland (Fig. 11- 13). The mucus
cell layer at the apical region of the gastric gland was greatly decreased
(compare Fig. 3 of the control and Fig.
11 of the treated).
Significant ultrastructural changes were revealed by
electron microscope. Several unusual structural features were observed
in oxyntic cells including some nuclei were
section of stomach of Nordette-treated mouse, showing oxyntic cells
(Ox), obvious decrease in mucus cells (Mc), dilated and congested
blood capillaries (bc), secretory granules at the gastric pit (arrow)
section of stomach of Nordette-treated mouse, showing oxyntic cells
(Ox), increase of secretory granules (arrow) in the gland lumen
and discharge of it TB (x1000)
section of stomach of Nordette-treated mouse, showing transverse
section of the gastric gland, oxyntic cells (Ox), peptic cells (P),
increase of secretory granules (arrow) in the gland lumen. TB (x1000)
section of stomach of Nordette-treated control mouse, showing oxyntic
cell with irregular cell boundaries contain irregular nucleus (N)
with highly electron-dense heterochromatin, peripheral dense nucleolus
(Nu), hypertrophied sER, increased, mitochondria (M) with dense
matrix, peripheral cytoplasmic vacuoles (arrow) underneath a very
thin diffusion distances between capillaries (bc) and adjacent oxyntic
section of stomach of Nordette-treated mouse, showing oxyntic cell,
nucleus (N), nucleolus (Nu), mitochondria (M) with dense matrix,
lysosome (L), intracellular canaliculi (IC) with irregular microvilli,
Note endothelial cells (E) with heterochromatic nucleus, vacuolated
basal lamina (arrow), (x7.500)
irregular, with increase in nuclear pores, contained dense heterochromatin
and peripherally located dense nucleoli (Fig. 14). The
mitochondria were irregularly distributed in the cytoplasm and not appeared
to be separated into peripheral group and a central perinuclear group.
The mitochondria increased and in some cells appeared smaller in size
and contained denser matrix. The most characteristic features in the oxyntic
cells were the pronounced increased and dilatation of smooth
section of stomach of Nordette-treated mouse, showing oxyntic cell,
nucleus (N) with peripheral nucleolus (Nu), mitochondria (M), sER
intracellular canaliculi (IC), hypertrophied Golgi apparatus (G),
lysosomes (L). peptic cell contain increased rER and large secretory
granules (arrow) (x5000)
of stomach of Nordette-treated mouse, showing hypertrophied peptic
cells, nucleus (N), increase in rER, ribosomes (R) and secretory
granules (arrow) (x7.500)
endoplasmic reticulum and an obvious decrease of the rough endoplasmic
reticulum (Fig. 14). The cytoplasm contained some lysosomes
(Fig. 15, 16); lysosomes were not
noticed in the oxyntic cells of the control animals. The Golgi apparatus
showed hypertrophy and increased activity (Fig. 15,
16). The intracellular canaliculi of some cells were
not clearly noticed as in the case of the control and in other cells appeared
increased (Fig. 16). The cytoplasm of oxyntic cells
beside the blood capillaries contained cytoplasmic vacuoles (Fig.
15). The peptic cells of the treated stomach appeared enlarged and
contained hypertrophied rough endoplasmic reticulum and an increase amount
of secretory granules in the apical cytoplasm (Fig. 16,
17). Blood capillaries lined by enlarged endothelial
cells containing heterochromatic, enlarged nuclei. Endothelial cell cytoplasm
section of stomach of Nordette-treated mouse, showing hypertrophied
smooth muscle fibers, hypertrophied heterochromatic nucleus (N)
with three electron-dense nucleoli (Nu), many ribosomes in degenerated
muscle fibers (arrow) (x5000)
lysosomes and ribosomes (Fig. 14, 15).
The smooth muscle fibers appeared enlarged and contained enlarged heterochromatic
nuclei which contained many nucleoli. Some of the muscle fibers showed
degeneration in their contractile proteins and contained numerous ribosomes
In the present study, the administration of Nordette
induced histopathological and ultrastructural alterations into stomach
of mice. These pathological symptoms included alteration in gastric gland
cells, blood vessels, lymphoid cells and smooth muscle fibers.
No work has been reported on possible histopathological
effects of the contraceptive drug on the stomach. Many previous reports
demonstrated histopathological effects of the contraceptive drug on some
other body organs including immune organs (Yousif, 1997), uterus (Hassanein,
2000) and pancreas (El Rawi and Yousif, 2006). Results of the present
work revealed that the histopathological changes of the stomach oxyntic
cells of treated animals included increase in size, weaker stainability
of the cytoplasm. Ultrastructural alterations included an irregular distribution
of the mitochondria in the cytoplasm, an obvious decrease of rough endoplasmic
reticulum and an increased and dilatation of smooth endoplasmic reticulum,
the Golgi apparatus showed hypertrophy and increased activity and the
cytoplasm contained some lysosomes, the cytoplasm of oxyntic cells beside
the blood capillaries contained cytoplasmic vacuoles. The intracellular
canaliculi of some cells were not clearly noticed as in the case of the
control. In the present study, peptic cells showed an increase in rER,
ribosomes, zymogen granules. Similarly, Hassanein (2000) stated that the
administration of oestradiol benzoate stimulated the secretory activity
of the uterine luminal epithelial cells as evidenced by the development
of rough endoplasmic reticulum, Golgi complexes and the abundance of secretory
granules and provokes other alterations including mitochondria were large
and swollen, most of their cristae got destructed and many lysosomes appeared
in the cytoplasm. Also the administration of Nordette caused increase
in rER, ribosomes, zymogen granules in the pancreatic acinar cells (El
Rawi and Yousif, 2006). In previous report, Yousif (1997) concluded that
the hormonal contraceptive drug caused hyperactivity of lymphocyte function,
this has been noticed by increase in the RNA and protein content in the
cells. The administration of androgenic steroids in fish leads to a general
increase in protein synthesis (Lone and Ince, 1983) and enhanced the proteolytic
activity of the digestive tract (Lone and Matty, 1981). In mammals, anabolic
steroids increase protein synthesis and activate the secretion (Michelsen
et al., 1982). On the other hand, GH promotes both increased size
of the cells and protein synthesis by the ribosomes, this is caused by
a direct effect on ribosomes, stimulate the transcription of DNA causing
formation of large quantities of RNA (Mountcastle, 1980). Sakai et
al. (2004) stated that ghrelin was recently identified as an endogenous
ligand for GH secretagogue receptor; in this study, they investigated
the effects of ovariectomy on the numbers of ghrelin-immunopositive and
-expressing cells, ghrelin mRNA levels and plasma ghrelin concentrations
in female rats. Three days after ovariectomy, the number of ghrelin cells,
plasma ghrelin level and the ghrelin mRNA level significantly increased.
These responses were reversed by 17β-estradiol replacement. they
also found that ghrelin-immunopositive cells express estrogen receptor.
These results suggested that estrogen is involved in the regulation of
In the present study the amount of the mucus cells at
the apical region of the gastric gland of the Nordette-treated animals
was greatly decreased. The major functions of the gastric epithelium are
the secretion of acid and digestive enzymes. It also secretes mucus to
lubricate ingested food and to protect itself from the corrosive effects
of the acid and enzymes. The mucosal surface of the stomach has a thick
protective coating of mucus (Junqueira et al., 1995). In the present
study, it has been noticed that Nordette cause damage to the mucus cell
layer. Confirming the present study, Williams and Tumberg (1981) stated
that a pH gradient across the mucus layer thereby established and acts
as a protective barrier between the luminal acid and the gastric epithelium,
therefore any damage to the mucus cells destruct the protective barrier.
On the other hand, recent evidence suggests that tight junctions around
surface and pit cells also form a part of barrier to acid. Stress and
other psychosomatic factors or such substances as aspirin that cause gastric
irritation can disrupt this epithelial layer and lead to ulceration (Junqueira
et al., 1995).
In the present study, concerning the blood vessels, it
appeared that an obvious adverse changes were observed in the blood vessels
and blood capillaries including dilatation and congestion especially in
the apical region of the gastric glands. Blood capillaries lined by enlarged
endothelial cells containing enlarged nuclei with highly condensed heterochromatin.
Endothelial cell cytoplasm contained lysosomes and ribosomes. Immunoblotting
studies on the estrogens phosphorylate endothelial NOS (NOS III), suggesting
that NOS III is the target for estrogen-induced NOS activity (Jaubert
et al., 2007). Cells of the gastric epithelium are responsible
for secretory and protective functions. To subserve these functions, an
intimate relationship between these cells and the mucosal microcirculation
might be expected. Physiological studies indeed show a direct relationship
between acid secretion and gastric mucosal blood flow (Lanciault and Jacobson,
1976); abundant evidence implicates a distributed mucosal microcirculation
in the pathogenesis of acute gastric ulceration (Moody, 1971). Progressing
toward the gastric lumen there was a notable decrease in the amount of
connective tissue between the basal lamina of the endothelium and that
of the adjacent gastric gland cells. The capillaries situated immediately
below the surface mucus cells were in close proximity to the basal laminae
of these cells. Diffusion distances between capillaries and adjacent parietal
cells of the gastric glands were usually even smaller (Gannon et al.,
1982). In agreement with the above observations, the cytoplasm of oxyntic
cells beside the blood capillaries contained many cytoplasmic vacuoles.
This observation suggested that there is a disturbance in the diffusion
of the small molecules between the capillary and the oxyntic cells.
The most pathological effects in the present study were
restricted to the extensible decrease of the lymphoid cells in the submucosa
and lamina propria. Although, there were many lymphoid cells lies in the
submucosa and lamina propria of the stomach of the control animals the
submucosa and lamina propria of the treated stomach were obviously decrease
of lymphoid cells. Also, Yousif (1997) demonstrated that administration
of a daily therapeutic dose of hormonal contraceptive drug for 30 days
cause an obvious decrease of lymphocytes in the thymus and the spleen
of the mice. Confirm to the above study, Jiang et al. (2006) showed
that exposure to estrogens is associated with an increased risk of developing
breast, cervical and liver cancer. Estrogens strongly induce the human
granzyme B inhibitor, proteinase inhibitor 9 (PI-9). Because cytotoxic
T lymphocytes (CTLs) and natural killer (NK) cells use the granzyme pathway
to induce apoptosis of target cells, Jiang et al. (2006) tested
the ability of activated CTLs and the human NK cell line, YT cells, to
lyse human liver cells. Estrogen induction of PI-9 protected the liver
cells against CTL and NK cell-mediated, granzyme-dependent, apoptosis.
Knockdown of PI-9 by RNA interference blocked the protective effect of
estrogen. This study demonstrates that estrogens can act on target cells
to control their destruction by immune system cells and shows that induction
of PI-9 expression can inhibit both CTL and NK cell-mediated apoptosis.
Estrogen induction of PI-9 may reduce the ability of cytolytic lymphocytes-mediated
immune surveillance to destroy newly transformed cells, possibly providing
a novel mechanism for an estrogen-mediated increase in tumor incidence
(Jiang et al., 2006).
On the other hand, in the present study Nordette cause
irregularity in cell boundaries of gastric gland cells. Recently, Li et
al. (2006) stated that estrogen depletion decreased and treatment
with 17β-estradiol increased deformability of cervical-vaginal epithelial
cells. Estrogen also induced redistribution of nonmuscle myosin II-B (NMM-II-B);
lesser interaction of NMM-II-B with actin; increased phosphorylation of
NMM-II-B-heavy chains at threonine and serine residues and decreased filamentation
of NMM-II-B in vitro. Augmented phosphorylation of NMM-II-B can
block filamentation and induce disassociation of the myosin from the cortical
actin and disruption of the actomyosin ring can increase cell deformability,
we confirm these observations.
Ueyama et al. (2006) showed that gastic parietal
cells produce and secrete a substantial amount of estrogen (E2) into portal
vein, using biochemical and morphological step-up of E2 concentration
in the portal vein, compared with that in the artery, which was not seen
after gastrectomy and also an increase of arterial E2 concentration after
portal-systemic shunting. This indicates that the stomach, but not other
organs, is the main origin of portal venous E2. In the physiological tissue
culture system, gastric mucosa produced testosterone and E2 from androstenetione.
These data indicate the presence of aromatase in gastric mucosa. Because
gastric mucosa contains a mixed population of cells. The histochemical
data indicate that parietal cells in gastric mucosa have aromatase mRNA
and its protein. Taken together, Ueyama et al. (2006) further showed
that the signals for ERmRNA and protein were located in hepatocytes and
mRNA levels for hepatic ER were regulated by E2 concentration in the blood
flowing into the liver. Ueyama et al. (2006) proposed an expanded
role of gastric parietal cells beyond the well-known exocrine function
of secreting hydrochloric acid and intrinsic factor into gastric juice,
that the gastric parietal cells serve an endocrine function, whereby estrogen
is synthesized and secreted into the portal vein, the consequences of
which may be of clinical significance in the case of portal-systemic shunting.
Gastric parietal cells are unique, in that they secrete E2 and can produce
its precursor, testosterone. Le Goascogne et al. (1995) reported
that gastric parietal cells may use circulating progesterone derived from
adrenal cortex and gonads as the first precursor for successive steps,
or they can also use circulating testosterone preferentially as the substrate
of aromatase. Gastric parietal cells are also endowed with a large number
of mitochondria and an elaborate tubulovesicular system, similar to a
smooth-surfaced endoplasmic reticulum. These structural characteristics
seem to be common in adrenal cortical cells and testicular interstitial
cells. A functional role for gastric estrogen has still to be elucidated.
As one possibility, gastric estrogen may act as a local regulator of the
gastro-hepatic axis, because ERα mRNA and immunoreactive protein
were expressed in hepatocytes and mRNA levels of hepatic ERα were
regulated by E2 concentration in the blood flowing into the liver. Estrogen
might affect gastric motility (Bond et al., 1998), because a low
level of ERα mRNA was expressed in the muscularis externa of stomach.
Gastric estrogen might also act directly on parietal cells in an autocrine
or intracrine fashion, because the protein and mRNA expression of ERα
and ERβ was reported in gastric parietal cells (Campbell-Thompson
et al., 2001).
By immunohistochemistry the expression of ER beta was
studied by Matsuyama et al. (2002) in 29 stomach adenocarcinomas.
In this study, the expression of ER beta in stomach adenocarcinomas has
been investigated for the first time. All adenocarcinomas, demonstrated
clear ER beta nucleus staining. Lymphocytes, venous endothelial cells,
smooth muscle and non-cancerous stomach glands also showed strong ER beta
staining, Equivalent ER beta protein levels in cancerous and non-cancerous
tissues, which was consistent with the results of immunohistochemical
staining. The results imply that the effects of estrogen in stomach cancer,
as well as those in normal stomach, may be mediated by ER beta (Matsuyama
et al., 2002). In the present study, the microscopic changes observed
in Lymphocytes, endothelial cells of the capillaries and smooth muscle
cells confirm the above observations.
From the present study, we can concluded that the
contraceptive drug induced extensive light and electron microscopic changes
and these changes may lead to disturbance in the stomach functions, including
disturbance in the secretory function of the parietal cells, the interaction
between the blood and oxyntic cells and the stomach mobility. The important
characteristic features is the obvious decrease in the mucus cell layer
in the treated stomach. On the other hand, the contraceptive drug obviously
induce adverse effect in the immune system cells by decrease the amount
of the lymphoid cells in the stomach. These changes, which noticed in
gastric gland cells, lymphocytes, endothelial cells and smooth muscle
fibers in the stomach may be mediated by ER. Contraceptive drug cause
histolpathogical changes in the stomach and it may be toxic to the stomach
when used for long period.
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