The document discusses prostaglandin production in ovine placentas. It finds that PGHS-1 is expressed in trophoblast epithelial cells and weakly in maternal tissues, while PGHS-2 expression increases specifically in trophoblast cells near term and during labor. Glucocorticoid treatment and spontaneous labor both significantly increase PGHS-2 levels in trophoblasts, indicating its role in elevated prostaglandin production during labor. The study elucidates the cellular localization of PGHS isozymes in ovine placentas and how their expression changes with gestation and the onset of labor.
Histochemical Studies of Enzymes in the Adrenal Gland of Rat & Rabbit during ...paperpublications3
Abstract: Administration of ACTH stimulates adrenal secretion of progesterone as well as corticosterone (Resko, 19691; Feder et al., 1969; Feder et al., 1971; Piva et al., 1973). Progesterone is both an obligatory intra-adrenal substrate for corticosterone production and a steroid essential for maintenance of pregnancy. Thus, the regulation of adrenal steroidogenesis during pregnancy has two potentially important aspects: i.) Maintenance of optimal blood levels of corticosterone and ii.)Contributing significant amounts of progesterone to the total maternal pool. Since the extended luteotrophic function of ovary in rat & mice during pregnancy is related to the Peroxidase-Ascorbate system (Agrawal, P. & Laloraya, M.M. 1979). It appears likely that synthesis of progesterone under the action of ACTH during pregnancy may be controlled by a similar mechanism as reported for LH in the ovary, thus causing increased synthesis and secretion of the Progesterone and corticosteroids from the adrenal gland.
Biochemical Changes in Ascorbate and Peroxidase Activity in the Adrenal Gland...paperpublications3
Abstract: Rapid synthesis of progesterone under the action of ACTH may be controlled by a similar mechanism as reported for LH in the ovary, thus causing increased synthesis and secretion of the progesterone and corticosteroids from the adrenal gland. ACTH is also known to cause depletion of adrenal ascorbate and cholesterol in the hypophysectomized rat which is shown to occur within Minutes of ACTH injection and to exhibit a characteristic time sequence. Peroxidase mediated conversion of pregnenolone to progesterone stimulated in the presence of ascorbate in the rat and rabbit ovarian tissue had also been demonstrated.Since ascorbate is known to be a donor in peroxidase reaction, the possibility of peroxidase system being involved in the rapid depletion of ascorbate during the normal reproductive cycle .
Histochemical Studies of Enzymes in the Adrenal Gland of Rat & Rabbit during ...paperpublications3
Abstract: Administration of ACTH stimulates adrenal secretion of progesterone as well as corticosterone (Resko, 19691; Feder et al., 1969; Feder et al., 1971; Piva et al., 1973). Progesterone is both an obligatory intra-adrenal substrate for corticosterone production and a steroid essential for maintenance of pregnancy. Thus, the regulation of adrenal steroidogenesis during pregnancy has two potentially important aspects: i.) Maintenance of optimal blood levels of corticosterone and ii.)Contributing significant amounts of progesterone to the total maternal pool. Since the extended luteotrophic function of ovary in rat & mice during pregnancy is related to the Peroxidase-Ascorbate system (Agrawal, P. & Laloraya, M.M. 1979). It appears likely that synthesis of progesterone under the action of ACTH during pregnancy may be controlled by a similar mechanism as reported for LH in the ovary, thus causing increased synthesis and secretion of the Progesterone and corticosteroids from the adrenal gland.
Biochemical Changes in Ascorbate and Peroxidase Activity in the Adrenal Gland...paperpublications3
Abstract: Rapid synthesis of progesterone under the action of ACTH may be controlled by a similar mechanism as reported for LH in the ovary, thus causing increased synthesis and secretion of the progesterone and corticosteroids from the adrenal gland. ACTH is also known to cause depletion of adrenal ascorbate and cholesterol in the hypophysectomized rat which is shown to occur within Minutes of ACTH injection and to exhibit a characteristic time sequence. Peroxidase mediated conversion of pregnenolone to progesterone stimulated in the presence of ascorbate in the rat and rabbit ovarian tissue had also been demonstrated.Since ascorbate is known to be a donor in peroxidase reaction, the possibility of peroxidase system being involved in the rapid depletion of ascorbate during the normal reproductive cycle .
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Fertility Response Following Induction of Lactation in Infertile Dairy Cowsijtsrd
The fertility response following induction protocol in infertile dry cows was assessed in terms of the induction of oestrus in anoestrus cows and conception in repeat breeders cows in the study. There were four anoestrous and two repeat breeder cows in G-1 out of them two anoestrus became cyclic, one repeater animal conceived after treatment protocol. Where as in G-2, three anoestrus and three repeat breeder animals given induction protocol where two anoestrous cows became cyclic and two repeaters conceived. The analysis of data revealed higher fertility response in G-2 as compared to G-1 (66.67 Vs. 50 %, respectively) with the higher conception rate (50 Vs. 33.33 %, respectively). It indicates better fertility response in G-2 as compared to G-1. K. Kumar | S. N. Shukla | S. Bhandekar | S. K. Singh | P. Inwati"Fertility Response Following Induction of Lactation in Infertile Dairy Cows" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-2 | Issue-2 , February 2018, URL: http://www.ijtsrd.com/papers/ijtsrd9615.pdf http://www.ijtsrd.com/other-scientific-research-area/other/9615/fertility-response-following-induction-of-lactation-in-infertile-dairy-cows/k-kumar
Progesterone Profile of Mature Nubian Goats iosrjce
Two experiments were carried out to record the progesterone (P4) profile of Nubian goats during
oestrous cycle and postpartum. In experiment I, 8 cyclic does were i.m injected with 2 doses of 125µg of
prostaglandin F2α (PG F2α) 11 days apart to synchronize oestrous. Eleven serum samples, were collected from
each doe at an interval of 2 days after the commencement of behavioral oestrous sings (day 0) and assayed for
P4 levels. The mean P4 concentration (conc.) on day 0 was 0.12 ± 0.01 ng/ml, then it increased gradually to
reach a peak of 6.03 ± 0.25 ng/ml on day 10 and it assumed a plateau over days 12 to 16. A sharp decline in P4
conc. was recorded on day 18 (0.3 ± 0.01 ng/ml) and a further drop (0.19 ± 0.00 ng/ml) ended the cycle on day
20. In experiment II, 10 parturient does were employed to study P4 profile during postpartum. Twenty one milk
samples, collected from each doe at 4 days interval starting from day 3 postpartum, were assayed for P4 levels.
The milk P 4 level remained below 0.04 ng/ml until a mean of 45 days postpartum; thereafter it increased to
attain values ≥ 1.0 ng/ml after the commencement of the first oestrus postpartum. It is concluded that P4 profile
of the Sudan Nubian goats during oestrous cycle and postpartum follows the normal trend of the P4 profile of
other breeds of goats with very minute differences in P4 conc. and the timing of peak values.
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Fertility Response Following Induction of Lactation in Infertile Dairy Cowsijtsrd
The fertility response following induction protocol in infertile dry cows was assessed in terms of the induction of oestrus in anoestrus cows and conception in repeat breeders cows in the study. There were four anoestrous and two repeat breeder cows in G-1 out of them two anoestrus became cyclic, one repeater animal conceived after treatment protocol. Where as in G-2, three anoestrus and three repeat breeder animals given induction protocol where two anoestrous cows became cyclic and two repeaters conceived. The analysis of data revealed higher fertility response in G-2 as compared to G-1 (66.67 Vs. 50 %, respectively) with the higher conception rate (50 Vs. 33.33 %, respectively). It indicates better fertility response in G-2 as compared to G-1. K. Kumar | S. N. Shukla | S. Bhandekar | S. K. Singh | P. Inwati"Fertility Response Following Induction of Lactation in Infertile Dairy Cows" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-2 | Issue-2 , February 2018, URL: http://www.ijtsrd.com/papers/ijtsrd9615.pdf http://www.ijtsrd.com/other-scientific-research-area/other/9615/fertility-response-following-induction-of-lactation-in-infertile-dairy-cows/k-kumar
Progesterone Profile of Mature Nubian Goats iosrjce
Two experiments were carried out to record the progesterone (P4) profile of Nubian goats during
oestrous cycle and postpartum. In experiment I, 8 cyclic does were i.m injected with 2 doses of 125µg of
prostaglandin F2α (PG F2α) 11 days apart to synchronize oestrous. Eleven serum samples, were collected from
each doe at an interval of 2 days after the commencement of behavioral oestrous sings (day 0) and assayed for
P4 levels. The mean P4 concentration (conc.) on day 0 was 0.12 ± 0.01 ng/ml, then it increased gradually to
reach a peak of 6.03 ± 0.25 ng/ml on day 10 and it assumed a plateau over days 12 to 16. A sharp decline in P4
conc. was recorded on day 18 (0.3 ± 0.01 ng/ml) and a further drop (0.19 ± 0.00 ng/ml) ended the cycle on day
20. In experiment II, 10 parturient does were employed to study P4 profile during postpartum. Twenty one milk
samples, collected from each doe at 4 days interval starting from day 3 postpartum, were assayed for P4 levels.
The milk P 4 level remained below 0.04 ng/ml until a mean of 45 days postpartum; thereafter it increased to
attain values ≥ 1.0 ng/ml after the commencement of the first oestrus postpartum. It is concluded that P4 profile
of the Sudan Nubian goats during oestrous cycle and postpartum follows the normal trend of the P4 profile of
other breeds of goats with very minute differences in P4 conc. and the timing of peak values.
Studying in Queensland: Uncovering Motivations & Experiences of International...Lara Klestov
As a part of the work integrated learning program at the University of Queensland (UQ), The Department of Tourism, Major Events, Small Business and the Commonwealth Games (DTESB) engaged the University of Queensland to undertake a research study investigating the motivations and experiences of international students in Queensland. This presentation highlights the results from 100 face-to-face surveys that were conducted.
1 aedl - iywinc lite block presentation march 2015 engMichael Walsh
New Patented Super LED Hi/Low-Bay LED Lights.
130 lm P/W, Life 150,000+ hr. life.
Operating temperature: -45 Deg C to +50 Deg C.
IP 68. Our 140 Watt LED will replace a 400 Watt HID.
This paper presents a review & performs a comparative evaluation of few known machine learning
algorithms in terms of their suitability & code performance on any given data set of any size. In this paper,
we describe our Machine Learning ToolBox that we have built using python programming language. The
algorithms used in the toolbox consists of supervised classification algorithms such as Naïve Bayes,
Decision Trees, SVM, K-nearest Neighbors and Neural Network (Backpropagation). The algorithms are
tested on iris and diabetes dataset and are compared on the basis of their accuracy under different
conditions. However using our tool one can apply any of the implemented ML algorithms on any dataset of
any size. The main goal of building a toolbox is to provide users with a platform to test their datasets on
different Machine Learning algorithms and use the accuracy results to determine which algorithms fits the
data best. The toolbox allows the user to choose a dataset of his/her choice either in structured or
unstructured form and then can choose the features he/she wants to use for training the machine We have
given our concluding remarks on the performance of implemented algorithms based on experimental
analysis
A SURVEY ON SIMILARITY MEASURES IN TEXT MINING mlaij
The Volume of text resources have been increasing in digital libraries and internet. Organizing these text documents has become a practical need. For organizing great number of objects into small or minimum number of coherent groups automatically, Clustering technique is used. These documents are widely used for information retrieval and Natural Language processing tasks. Different Clustering algorithms require a metric for quantifying how dissimilar two given documents are. This difference is often measured by similarity measure such as Euclidean distance, Cosine similarity etc. The similarity measure process in text
mining can be used to identify the suitable clustering algorithm for a specific problem. This survey discusses the existing works on text similarity by partitioning them into three significant approaches; String-based, Knowledge based and Corpus-based similarities.
A Multi-Level Security for Preventing DDOS Attacks in Cloud Environmentsmlaij
Incredible and amazing growths in the meadow of extranet, internet, intranet and its users have developed an innovative period of great global competition and contention. Denial of service attack by several computers is accomplished of distressing the services of competitor servers. The attack can be done for various reasons. So it is a key threat for cloud environment. Distributed-Denial of Service (DDoS) is a key intimidation to network and cloud computing security. Cloud computing Network is a group of nodes that interrelate with each other for switch over the information. So security is the major issue. There are several security attacks in cloud computing. One of the major intimidations to internet examine is DDoS attack. It is a malevolent effort to suspending or suspends services to destination node. DDoS or DoS is an effort to create network resource or the machine is busy to its intentional user. Numerous thoughts are developed for avoid the DDoS or DoS. DDoS occur in two different behaviours they may happen obviously or it may due to some attackers.
It appears that you have provided information about the "Indo-American Journal of Agricultural and Veterinary Sciences" . This journal seems to be an international online publication in English, published quarterly. It emphasizes fast publication while maintaining a rigorous peer-review process of the published journals.
The onset of parturition, commonly known as labor, is a complex physiological process that marks the culmination of pregnancy and the initiation of the birthing process. This intricate sequence of events involves a series of hormonal, mechanical, and neurological changes that ultimately lead to the expulsion of the fetus from the mother's uterus. Understanding the onset of parturition requires a comprehensive exploration of the various stages and factors involved.
The process of parturition can be broadly categorized into three main stages: pre-labor, labor, and post-labor. The pre-labor stage encompasses the preparatory changes occurring in the days and weeks leading up to labor, while the labor stage involves the actual contractions and cervical dilation facilitating delivery. The post-labor stage involves the expulsion of the placenta and the initial postpartum adjustments.
The hormonal regulation of parturition is a crucial aspect of its onset. Throughout pregnancy, the placenta produces progesterone, a hormone that maintains the uterine environment and prevents premature contractions. As term approaches, the ratio of progesterone to estrogen changes, leading to a decline in progesterone levels and a subsequent increase in estrogen. This shift triggers a cascade of events, including the activation of uterine contractions and the initiation of cervical ripening.
The role of oxytocin, often referred to as the "love hormone" or "cuddle hormone," is paramount in the onset of labor. Produced by the hypothalamus and released by the pituitary gland, oxytocin stimulates uterine contractions. Additionally, oxytocin plays a crucial role in the positive feedback loop of labor – as contractions intensify, more oxytocin is released, further promoting labor progression.
Mechanical factors also contribute to the onset of parturition. The growing fetus applies pressure on the cervix and uterine walls, leading to the release of prostaglandins. Prostaglandins are lipid compounds that promote uterine contractions and cervical ripening. The combination of hormonal changes and mechanical pressure creates a synergistic effect, fostering the progression of labor.
The intricate interplay between the maternal-fetal unit and the surrounding environment further influences the onset of parturition. Maternal stress, for instance, can impact the release of corticotropin-releasing hormone (CRH), which, in turn, influences the production of other hormones involved in labor. Moreover, the fetus itself plays an active role in signaling its readiness for delivery through various molecular signals.
The onset of labor is often heralded by a set of common signs. These may include the engagement of the fetal head into the pelvis, the "bloody show" – a discharge of mucus mixed with blood resulting from cervical changes, and the rupture of the amniotic sac, leading to the release of amniotic fluid. These signs, in conjunction with regular and increasingly intense contractions.
Participation of the oviductal s100 calcium binding protein G in the genomic effect of estradiol that accelerates oviductal embryo transport in mated rats
Mariana Ríos1, Alexis Parada-Bustamante1, Luis A Velásquez2,3, Horacio B Croxatto2,3,4 and Pedro A Orihuela2,3*
By Luis Alberto Velasquez Cumplido
Biochemical changes induced by Bioneem (0.03%) formulation in chick embryogen...Agriculture Journal IJOEAR
Abstract— In ovo studies on the effect of 1,3,5, ppm Bioneem (0.03%) formulation on Biochemical aspect of chick embryo revealed that there was dose dependent total protein reduction in 96 hrs old embryo (treated at 24 hrs) as compared to the control. Also there was reduction in total protein concentration Liver, Brain and Heart of 15 day old chick embryo (treated with Bioneem at 96 hrs. stage) as compared to that of control. Protein carbonyl concentration of 96 hrs old embryo (treated at 24 hrs with Bioneem) and that of Liver, Brain and Heart of 15 day old chick embryo (treated with bioneem at 96 hrs) increased in dose dependent manner. Most affected organ was Liver and least affected organ was Heart. Blood analysis of 15 day old chick embryo (treated with Bioneem at 96 hrs) showed increased level of Blood urea, LDH, SGOT, SGPT, while Serum alkaline phosphatase and serum cholesterol were decreased in dose dependent manner as compared to the control. Thus Bioneem though ecofriendly pesticide can adversely affect vertebrate non target organisms and therefore should be carefully used in pest management programs.
The objective of this study was to evaluate the effect of the systematic use of a single amount of the prostaglandin F2α after one month of calving on the renewal of sexual activity of beef at Chad. (Sixty five (65) cows of local breeds from N'Djamena urban area were selected and divided Into two groups: Forty-three (43) cows were treated with Prostaglandin F2α, one month after calving and 22 cows as witnesses. More than half (60.46%) of the cows answered the treatment and 39.54% did not react. The answer varied with the parity (68%) for the first half of the cows against 32% for the multiparous but the body weight at calving did not-have an effect. Only, the acceptance of overlapping was Retained for the detection of return in heat, which took place on an average of 2.36 ± 0.14 days after treatment. The treatment made it feasible to-have-one year interval between calving (first heat of 32.36 ± 0.14 days.) Heat thus obtained, can be used as reference to the stockbreeders to program the reproduction of their herd in order to have one year interval calving-calving closed. This test sample gave better Indication in first half of the cows while waiting for confirming it in the station.
The relationship between progesterone and biochemical constituents of amnioti...Ali Olfati
Ali Olfati1, Gholamali Moghaddam1, Nasroallah Moradi Kor2*, Mitra Bakhtiari3
1Department of Animal Science, Faculty of Agriculture, University of Tabriz, Iran
2Department of Reproduction Physiologies, Iranian Society of Physiology and Pharmacology, Tehran, Iran
3Department of Anatomical Sciences, Faculty of Medicine, University of Medical Sciences, Kermanshah, Iran
2. 1996). Immunohistochemistry revealed an increase in the
content of PGHS-2 in mononuclear trophoblast cells of
the placenta after day 140 of gestation. It was concluded
that PGHS-2 is the enzyme responsible for placental
prostaglandin production at term delivery. However, the
study of Gibb et al. (1996) did not elucidate the cellular
localization of PGHS-1 expression nor did it address the
factors that regulate enzyme formation.
Glucocorticoids regulate prostanoid production in many
tissues. PGHS activity increases after glucocorticoid treat-
ment in mouse myeloid leukaemia (MI) cells (Honma et al.,
1980), fetal rat lungs (Tsai et al., 1983) and in Swiss 3T3 mouse
fibroblast cells (Chandrabose et al., 1980). Human amnion
cells in primary culture have an increased capacity to convert
exogenous arachidonic acid into PGE2
in response to
dexamethasone treatment (Smieja et al., 1993). As labour
onset in ewes is preceded by activation of the fetal
hypothalamic–pituitary–adrenal axis (HPA axis), it is
possible that the prepartum increase in fetal cortisol
production acts to modulate intrauterine PGHS expression.
McLaren et al. (1996) demonstrated that the placentome is
the principal site of induced PGHS expression at the time
of glucocorticoid-induced labour in sheep, although the
distribution of the two forms of the enzyme within the
placentome was not addressed. The aim of the present
study was to characterize the cellular distribution of ir-
PGHS-1 and ir-PGHS-2 in ovine placental tissue using
immunohistochemical staining. The primary sites of
enzyme production were determined in placentomes after
glucocorticoid-induced and spontaneous labour and were
compared with those of ewes in late gestation but not in
labour.
Materials and Methods
Animals
Sixteen pregnant Border Leicester–Merino crossbred ewes
of known gestational age were used in this study. The animal
experiments were approved by the Monash University
Standing Committee on Ethics in animal experimentation.
Ewes underwent surgery between 118 and 125 days of
gestation to implant electromyogram electrodes. Electrodes
were implanted into the myometrium to enable labour onset
to be identified (Harding et al., 1982).
A paired treatment–control experimental paradigm
was used in the first part of the study to determine the effects
of glucocorticoid on PGHS expression. Fetal sheep were
injected using guided ultrasonography (via maternal
transabdominal injection) with either glucocorticoid
(betamethasone; Celestone Chronodose, Schering Plough
Pharmaceuticals, Baulkham Hills, NSW; 5.7 mg ml–1
in 1 ml
total volume; n = 5) or an equivalent volume of sterile
isotonic saline (control; n = 5) on day 131 of gestation.
Animals that received betamethasone were killed by i.v.
barbiturate injection when labour was established. Labour
onset occurred 56.6 Ϯ 0.8 h after glucocorticoid injection and
was determined from increased uterine electromyogram
activity and clinical factors such as swelling of the maternal
udder and vulva. Control ewes treated with saline were
killed at the same time as the experimental ewes to obtain
age-matched control tissues. The procedure of intrafetal
injection of glucocorticoid has been characterized and
validated (McLaren et al., 1996).
Six sheep were used for the second part of the study to
examine PGHS-1 and PGHS-2 expression at spontaneous-
onset labour. Ewes were allowed to progress to term and
were killed by i.v. barbiturate injection when increased
uterine contractile activity (consistent with labour onset) had
been observed for 6–8 h. The mean age to labour onset in
these ewes was 149 days.
Tissue collection
After the animals were killed, a post mortem examination
was conducted and tissues were collected in the shortest
possible period. Placentomes (sliced vertically along a radial
axis of symmetry) were placed in cold Bouin’s fixative
solution for 24 h. Tissues were washed five times in 70%
ethanol before dehydration and embedding. Tissues were
dehydrated using an automatic tissue processor (Histokinette,
Thomas Optical and Scientific Co. Pty Ltd). Tissues were
incubated sequentially in 70%, 90% and 100% ethanol for
12 h (2 ϫ 2 h washes), cleared of dehydrating agents by
washing in Histosol (2 ϫ 2 h washes) and embedded in
paraffin wax. Tissue sections (7 µm) were prepared by
standard techniques and placed on glass slides coated with
poly-L-lysine (0.01%; Sigma Chemical Company, St Louis,
MO).
Immunohistochemistry
Immunohistochemistry was performed using the DAKO
ENVISION™
System (DAKO Corporation, Carpinteria, CA).
After paraffin wax was removed from the tissue sections,
they were incubated with 0.03% (w/v) H2
O2
for 5 min to
quench endogenous peroxidase activity. Tissue sections were
incubated for 1 h at room temperature with PGHS-1 or
PGHS-2 antibody at 1:1000 dilution. PGHS-1 antiserum was
raised in rabbits against ram seminal vesicle PGHS (McLaren
et al., 1996). PGHS used for injection (99% purity) was
purchased from the Oxford Biochemicals Company (Oxford,
MI). The crossreactivity of the antibody with the PGHS-2
isozyme was estimated to be < 0.1%, as determined from
laser densitometry (McLaren et al., 1996). The polyclonal
PGHS-2 antiserum, raised in rabbits against murine PGHS-2,
was purchased from the Cayman Chemical Company (Ann
Arbor, MI). There was no detectable crossreactivity of the
PGHS-2 antibody with PGHS-1 protein at amounts up to
2.5 mg (as determined from western blot analysis; McLaren
et al., 1996). After washing with Tris-buffered saline (TBS)
(pH 7.5), the sections were incubated with peroxidase-
labelled polymer conjugated to goat anti-rabbit IgG second
antibody for 30 min. Slides were rinsed once and the
substrate–chromogen solution was added for 10 min.
Specific immunostaining was identified using diamino-
benzidine. The sections were counterstained with Harris’
34 W. J. McLaren et al.
3. haematoxylin, dehydrated and mounted with DPX moun-
tant. Immunoreactivity in tissue sections was examined
using a Leitz Wetzlar Dialux 20 microscope.
Controls
Tissues from animals of each group were processed
simultaneously to allow direct comparison between staining
runs. The following negative controls (derived from the
same specimen) were included in every staining run to
monitor daily variations in the immunohistochemical
staining procedure and to verify the performance of the
reagents: (i) the PGHS-1 or PGHS-2 primary antibody
was substituted either by antibody dilution buffer or by non-
immune rabbit serum (1:1000 dilution); (ii) the peroxidase-
labelled secondary link antibody (goat anti-rabbit immuno-
globulins) was substituted with TBS (pH 7.5) wash buffer;
and (iii) the slide section was only incubated with TBS
(pH 7.5) diluent before the addition of the substrate–
chromogen solution.
If any of the control specimens demonstrated residual
background staining, the results of the staining run were
considered to be invalid.
Pre-absorption controls were also performed to verify that
the staining observed in tissue sections was due to binding of
the primary antibody to its target antigen. PGHS-2 antibody
(protein concentration: 33 µg µl–1
) was pre-absorbed with an
excess (4000 times greater protein concentration) of PGHS-2
antigen (protein concentration: 1.04 µg µl–1
) for 18 h at 4ЊC.
Similarly, the PGHS-1 antibody (protein concentration: 83 µg
µl–1
) was pre-absorbed with an excess (4000 times greater
protein concentration) of PGHS-1 antigen (protein concen-
tration: 1 µg µl–1
) for 18 h at 4ЊC.
After addition of precipitating solution (0.05 mol phos-
phate buffer l–1
, 0.25% BSA, 5% polyethylene glycol 6000,
0.125% (v/v) normal rabbit serum and 0.5% (v/v) goat anti-
rabbit IgGs), the mixtures were centrifuged at 1200 g for
30 min at 4ЊC. The supernatant was incubated with the tissue
section. The tissue sections used for the PGHS-1 and PGHS-2
pre-absorption control incubations were derived from a
betamethasone-injected animal in labour previously shown
to demonstrate positive staining for both isozymes. A serial
section of the same specimen was run concurrently with
every pre-absorbed antibody control tissue to act as a
positive control.
Quantitation of immunohistochemical staining
Quantitative analysis of positive staining for ir-PGHS-1
and ir-PGHS-2 in placental tissue sections was performed
using image analysis software (Zeiss KS 400, Version 3.0).
Five placentomes collected from the different animals within
each treatment group (saline injection and betamethasone
administration) and six placentomes collected from the
animals in the spontaneous labour group were examined for
immunoreactivity. The percentage area of positive staining
was calculated from five fields of view (ϫ 20 magnification)
for each tissue section examined. Two sections per placen-
tome were assessed for the degree of positive staining. The
results are expressed as the mean percentage area of positive
staining for PGHS-1 and PGHS-2.
Statistical analysis
Statistical analysis of the data was performed using a
commercially available statistical program (GraphPad Prism
Version 2.01). Data were first analysed by a univariate
homogeneity of variance test (Bartlett’s Box F test). If
significance was found for a particular parameter (that is the
raw data were non-homogeneous), the test was repeated
using log10
transformation and square root transformation.
The transform most closely attaining homogeneity was then
used for all subsequent statistical analyses. Significant
interactions occurring between two or more factors were
identified using multifactorial ANOVA. The post hoc test of
least significant difference (LSD) was used subsequent to
the ANOVA to identify significant differences between pairs
of mean values. A probability level of 5% (P < 0.05) was
specified as significant. The data are presented graphically in
an untransformed state. Values are expressed as mean Ϯ SEM.
Results
PGHS-1 immunoreactivity
The location and the percentage area of tissue expressing
PGHS-1 immunoreactivity in sheep placentomes obtained
after intrafetal saline injection, glucocorticoid-induced
labour and spontaneous parturition are shown (Figs 1
and 2, respectively). In all groups studied, PGHS-1 im-
munostaining was localized to mononuclear cells of the
trophoblastic epithelium. The binucleate cells were clearly
immunonegative. The endothelial cells of fetal vessels in the
chorionic villi demonstrated weak immunostaining (Fig. 1b,
arrow). Moreover, the fibroblasts of the fetal stroma were
weakly immunopositive for PGHS-1. Both the fetal and
maternal mesenchymal core demonstrated diffuse staining
for PGHS-1. There was no significant effect of spontaneous
parturition or treatment (saline injection or glucocorticoid-
induced labour) on the location and the percentage area of
positive staining for PGHS-1 in placental tissue sections.
When control incubations were performed with primary
PGHS-1 antibody that had been pre-absorbed with purified
PGHS-1 isolated from ram seminal vesicles, no staining was
observed (Fig. 3a). Similarly, no staining for ir-PGHS-1 was
observed when the primary antibody was substituted in the
staining procedure with normal non-immune rabbit serum
(Fig. 3b).
PGHS-2 immunoreactivity
PGHS-2 was present in placental cotyledon tissue sections
on day 133 of gestation, as demonstrated in the saline-
injected control animals (Fig. 4a). Staining was confined to
the trophoblastic mononuclear epithelial cells. Binucleate
PGHS-1 and PGHS-2 immunoreactivity in ovine placenta 35
4. cells were consistently immunonegative. In contrast to the
PGHS-1 isozyme, the endothelial cells of fetal vessels in the
chorionic villi and the fibroblasts of the fetal stroma did not
contain ir-PGHS-2. Furthermore, both the fetal and maternal
stroma were immunonegative for PGHS-2. The location of
staining for PGHS-2 did not alter in response to glucocorticoid-
induced (Fig. 4b) or spontaneous labour (Fig. 4c); however,
there was a significant increase in the percentage area of
positive staining for PGHS-2 at labour onset (Fig. 2; P < 0.05).
The amount of positive staining for PGHS-2 in placental
tissue collected after glucocorticoid-induced labour and
spontaneous parturition was significantly higher than that
after intrafetal saline injection.
When control incubations were performed with primary
PGHS-2 antibody that had been pre-absorbed overnight with
purified PGHS-2 isolated from sheep placenta (70% purity),
no staining was observed in cells that previously demon-
strated positive staining for the PGHS-2 isozyme (Fig. 3c).
Moreover, no positive immunoreactivity was observed in
tissue sections in which the primary antibody was substituted
with non-immune rabbit serum (Fig. 3d).
Discussion
This study has demonstrated the presence and localization of
ir-PGHS-1 and ir-PGHS-2 in sheep placental tissue after
glucocorticoid-induced labour and spontaneous parturition.
Animals administered an intrafetal injection of isotonic
saline on day 131 of gestation acted as non-labour controls.
In all groups studied, the PGHS-2 isozyme was localized
exclusively to trophoblastic epithelial cells; no detectable
36 W. J. McLaren et al.
(a)
(b)
(c)
Fig. 1. Patterns of immunostaining for immunoreactive prosta-
glandin G/H synthase 1 (ir-PGHS-1) in placental tissue collected
from sheep after (a) intrafetal saline injection, (b) glucocorticoid-
induced labour and (c) spontaneous parturition. PGHS-1
immunoreactivity was localized to mononuclear cells of the
trophoblastic epithelium and the fibroblasts of the fetal stroma.
Binucleate cells were clearly immunonegative. Maternal placental
tissue demonstrated weak staining for the PGHS-1 isozyme.
Endothelial cells of the fetal vessels demonstrated weak
immunoreactivity for PGHS-1 (arrow in left-hand corner of (b)).
BNC: binucleate cell; F: fetal stroma; M: maternal placenta; T:
trophoblastic epithelium. Scale bar represents 50 µm.
30
25
20
15
10
5
0
PGHS-1 PGHS-2
a
a a a
b
b
Areaofpositivestaining(%)
Fig. 2. Computer-assisted quantitation of positive staining for
immunoreactive prostaglandin G/H synthase 1 and 2 (ir-PGHS-1
and ir-PGHS-2) in placental tissue collected from sheep after
intrafetal saline injection (ᮀ), glucocorticoid-induced labour ()
and spontaneous parturition (). Bars with different letters are
significantly different (P 0.05).
5. staining was observed in the intervening maternal epi-
thelium or stroma of the cotyledons. Similarly, ir-PGHS-1
was present primarily in the trophoblast cells, although cells
in the maternal mesenchyme and epithelium were weakly
immunopositive for this enzyme. There were no cell-specific
alterations in PGHS-1 or PGHS-2 immunoreactivity in sheep
placentomes during the last 10–15 days of pregnancy.
Furthermore, localization of enzyme staining was not
influenced by labour-onset, whether spontaneous or
glucocorticoid-induced.
The placenta has been identified as the major site of
prostaglandin production in the uterus of pregnant sheep.
Risbridger et al. (1985) demonstrated that prostaglandin
synthesis by dispersed ovine cotyledonary cells is low
during early and mid-gestation but increases markedly from
day 110–120 of gestation to term. In that study, exogenous
administration of arachidonic acid to incubations of
cotyledonary cells did not significantly increase prosta-
glandin synthesis. This finding is consistent with the
suggestion that PGHS activity in placental trophoblast cells
increases only after day 110 of gestation. The results of the
present study indicate that the fetal trophoblast cells are the
primary site of prostaglandin formation during the last
10–15 days of pregnancy.
In the past, studies of placental endocrine function in vitro
have relied on the use of heterogeneous placental explants
(Hoffmann et al. 1979; Matt and MacDonald, 1984) or cell
suspensions (Branchaud et al., 1983; Shemesh et al., 1984a,b)
to elucidate possible sites of hormone formation. Although
these studies have contributed to our knowledge of placental
endocrinology, the contribution of individual types of cell
was not elucidated. In an attempt to delineate the principal
sites of prostaglandin output in sheep placenta, Mitchell and
Flint (1978) manually separated the fetal cotyledon from the
maternal caruncle. The prostaglandin synthesizing capacity
of the two sides of the placentome was then investigated
using a dispersed cell preparation. Since the fetal cotyledon
had a greater prostaglandin output than the maternal uterine
epithelium, it was suggested that PGHS activity was
primarily localized to the fetal side of the placentome.
However, in that study, no anatomical evidence was pre-
sented to support the completeness of separation of fetal and
PGHS-1 and PGHS-2 immunoreactivity in ovine placenta 37
(a) (b)
(c) (d)
Fig. 3. Control tissue sections for the prostaglandin G/H synthase 1 and 2 (PGHS-1 and PGHS-2) staining procedures. When the
primary PGHS-1 antibody was pre-absorbed with purified immunoreactive PGHS-1 (ir-PGHS-1) isolated from ram seminal vesicles
(a), no staining was observed. Similarly, when the primary PGHS-2 antibody was pre-absorbed with ir-PGHS-2 isolated from sheep
placenta (c), no positive staining was observed. When the primary antibodies were substituted in the immunohistochemical staining
procedure with non-immune rabbit serum, no staining was observed (b,d). The tissue used for the control staining procedures was
derived from an animal in labour after intrafetal injection of betamethasone. This tissue was previously shown to demonstrate
positive staining for the PGHS-1 and PGHS-2 isozymes. BNC: binucleate cell; F: fetal stroma; M: maternal placenta; T: trophoblastic
epithelium. Scale bars represent 50 µm.
6. maternal components. Given the high degree of
interdigitation of the chorionic villi with the crypts of the
uterine mucosa, complete separation of the ovine cotyledon
into fetal and maternal components is not technically
feasible. Thus, the results are potentially confounded by the
possibility of a non-homogeneous cell preparation.
Evans et al. (1982) measured PGE2
, PGF2α
and 6-keto-PGF1α
output by sheep cotyledons at different stages of pregnancy
using an in vitro cell culture system. The capacity of
dispersed placental cells to synthesize these prostaglandins
was higher on days 130 and 145 than on days 50 and 100 of
gestation. This finding is consistent with the suggestion of
increased PGHS expression; however, the localization of
enhanced PGHS production could not be characterized as no
histological assessment was made of the types of cell in the
preparation. Risbridger et al. (1985) used a similar dispersion
protocol and suggested that cells of the fetal trophoblast
were the major site of prostaglandin production during
pregnancy and parturition. Although these investigators
demonstrated that enzymatic digestion of the placental
tissue yielded binuclear and mononuclear cells, the
dispersion technique excluded syncytial cells from the final
preparation. Thus, the contribution of maternal epithelial
cells to prostaglandin output was not demonstrated.
The data presented in the present study clearly
demonstrate that PGHS-1 and PGHS-2 enzyme formation
predominantly localizes to the mononuclear cells of the
fetal trophoblast. Other studies have confirmed that the
subcellular locations of PGHS-1 and PGHS-2 are also the
same. PGHS-1 was first shown to be localized to the
endoplas-mic reticulum and nuclear membrane of kidney
tissue sections using immunofluorescence (Smith and
Wilkin, 1977; Smith and Bell, 1978). This result was later
confirmed by immunoelectron microscopy of cultured
mouse fibroblasts (Rollins and Smith, 1980). Regier et al.
(1993) demonstrated that PGHS-2 is associated with the
endoplasmic reticulum and nuclear envelope of mouse 3T3
fibroblast cells.
Unlike the mononuclear trophoblast cells of the fetal
syncytium, binucleate cells of the trophoblast were shown to
be immunonegative. This result is consistent with the
findings of Boshier et al. (1991) and Gibb et al. (1996) and
indicates that these cells are not important to placental
prostaglandin formation. Rather, binucleate cells have two
alternative functions that are important to the normal
progression of pregnancy: (i) to form the fetomaternal
syncytium essential for successful implantation and
subsequent placental growth; and (ii) to produce and secrete
protein and steroid hormones. During the last two-thirds of
pregnancy, placental lactogens are measurable in the
maternal and fetal circulations of sheep (Chan et al., 1978;
Martel and Lacroix, 1978), cattle (Wallace et al., 1985) and
goats (Currie et al., 1990). The binucleate cells are the sole
source of these hormones. Furthermore, binucleate cells of
sheep and cows are capable of considerable progesterone
production from endogenous sources (Reimers et al., 1985;
Ullman and Reimers, 1989; Wango et al., 1991).
Quantitation of immunoreactive staining in placental
tissue sections using image analysis software demonstrated
a significant increase in PGHS-2 in association with
glucocorticoid-induced and spontaneous-onset labour. This
result strongly supports previous reports of increased
PGHS-2 protein concentrations in sheep placenta after
glucocorticoid-induced labour using western blot analysis
38 W. J. McLaren et al.
(a)
(b)
(c)
Fig. 4. Patterns of immunostaining for immunoreactive prosta-
glandin G/H synthase 2 (ir-PGHS-2) in placental tissue collected
from sheep after (a) intrafetal saline injection, (b) glucocorticoid-
induced labour and (c) spontaneous labour. PGHS-2 immuno-
reactivity was localized to mononuclear cells of the trophoblastic
epithelium. Binucleate cells and fibroblasts of the fetal stroma were
immunonegative. Maternal placental tissue did not express the
PGHS-2 isozyme. BNC: binucleate cell; F: fetal stroma; M:
maternal placenta; T: trophoblastic epithelium. Scale bar represents
50 µm.
7. (McLaren et al., 1996). In saline-injected control animals,
expression of the PGHS-2 isozyme was low. PGHS-2 staining
patterns in placental tissue sections from animals in labour
reflect higher enzyme production rates by uninucleate
trophoblast cells rather than increased expression by a more
diverse group of cells.
In summary, co-localization of PGHS-1 and PGHS-2
implies that the source of arachidonic acid, the site of
prostanoid formation and the mechanism of product
transport from the inside to the outside of the cell are the
same for these two isozymes. The primary difference
between PGHS-1 and PGHS-2 lies in the differential
regulation of enzyme expression. PGHS-1 isozyme
formation is constitutive. Conversely, PGHS-2 expression is
induced by a variety of stimulatory factors including
glucocorticoid-induced enzymes and cytokines (Liggins and
Thorburn, 1994). Regulation of PGHS-2 formation is critical
to myometrial activation that results in birth.
The work described in this manuscript was supported by a
Project Grant from the National Health and Medical Research
Council of Australia (G. E. Rice). G. E. Rice is in receipt of an NH
MRC Principal Research Fellowship. The authors wish to thank Carl
Zeiss Pty Ltd for providing the imaging software for quantitative
analysis of the data.
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