bluuebunny/crossref_metadata_2025 huggingface dataset:
Modeling and Monitoring Tools to Assess Recovery Status and Convergence Rates between Restored and Undisturbed Coral Reef Habitats
Authors: Lirman Diego , Miller Margaret W.
Published: November 2003

Abstract Boating activities are an increasing source of physical damage to coral reefs worldwide. The damage caused by ship groundings can be significant and may result in a shift in reef structure and function. In this study we evaluate the status of two restoration projects established in 1995, 6 years after two freighters, the M/V Maitland and the M/V Elpis, ran aground on reefs of the Florida Keys National Marine Sanctuary. Our approach includes field monitoring in support of simulation model development to assess the effectiveness of the restoration efforts. A population model was developed for the coral Porites astreoides to project the convergence rates of coral abundance and population size structure between the restored and surrounding reference habitats. Coral communities are developing rapidly on the restoration structures. Species richness and abundance of the dominant coral, P. astreoides, were nearly indistinguishable between the restoration structures and reference habitats after only 6 years. However, although abundance and size structure of P. astreoides populations are rapidly approaching those of the reference habitats (a convergence in size structure within 10 years was simulated), maximum coral size will take twice as long to converge for this species. The sensitivity of the model to maximum recruitment rates highlights the importance of recruitment on the recovery rates of restored habitats, suggesting that special attention should be afforded to provide coral recruits with appropriate recruitment substrate at the time of restoration. Finally, the rates of convergence and, hence, the level of success of a restoration effort were shown to be influenced not only by the recruitment and survivorship rates of corals on the restoration structures but by the characteristics of the reference population as well. Accordingly, reference populations ought to be considered a “moving target” against which restoration success has to be measured dynamically. The simple, cost‐effective, monitoring–modeling approach presented here can provide the necessary tools to assess the current status of a restoration effort and to project the time required for coral populations to resemble those found on undamaged reference habitats
Rab23, a Negative Regulator of Hedgehog Signaling, Localizes to the Plasma Membrane and the Endocytic Pathway
Authors: Evans Timothy M. , Ferguson Charles , Wainwright Brandon J. , Parton Robert G. , Wicking Carol
Published: November 2003

The regulation of hedgehog signaling by vesicular trafficking was exemplified by the finding that Rab23, a Rab‐GTPase vesicular transport protein, is mutated in open brain mice. In this study, the localization of Rab23 was analyzed by light and immunoelectron microscopy after expression of wild‐type (Rab23‐GFP), constitutively active Rab23 (Rab23Q68L‐GFP), and inactive Rab23 (Rab23S23N‐GFP) in a range of mammalian cell types. Rab23‐GFP and Rab23Q68L‐GFP were predominantly localized to the plasma membrane but were also associated with intracellular vesicular structures, whereas Rab23S23N‐GFP was predominantly cytosolic. Vesicular Rab23‐GFP colocalized with Rab5Q79L and internalized transferrin‐biotin, but not with a marker of the late endosome or the Golgi complex. To investigate Rab23 with respect to members of the hedgehog signaling pathway, Rab23‐GFP was coexpressed with either patched or smoothened. Patched colocalized with intracellular Rab23‐GFP but smoothened did not. Analysis of patched distribution by light and immunoelectron microscopy revealed it is primarily localized to endosomal elements, including transferrin receptor‐positive early endosomes and putative endosome carrier vesicles and, to a lesser extent, with LBPA‐positive late endosomes, but was excluded from the plasma membrane. Neither patched or smoothened distribution was altered in the presence of wild‐type nor mutant Rab23‐GFP, suggesting that despite the endosomal colocalization of Rab23 and patched, it is likely that Rab23 acts more distally in regulating hedgehog signaling.
Development and use of virus-vectored immunocontraception
Authors: Williams C. K.
Published: November 2003

Virus-vectored immunocontraception (VVIC) is perceived to present biological
risks, real or baseless, which create social and political constraints to
deploying VVIC for managing vertebrate pests. Developing and deploying VVIC
must be justified and address biological, social and political risks. Future
needs for pest management will influence deployment of VVIC. Projections of
human society and pest impact on agriculture and conservation suggest
increasing need for cost-beneficial strategies. Best strategies are likely to
integrate various methods, possibly including forms of VVIC. Processes
identifying future pest impacts and roles for VVIC are shown using the rabbit
in Australia. Present research developing VVIC aims to test physiological
feasibility, to develop it for specific pests, and address aspects of
ecological feasibility. Minimizing biological risks through choosing
species-specific antigens and, if possible, viral vectors, is central and
overseen by regulatory authorities. International collaborators study related
valued species to develop protective strategies. Excellent science can
withstand legal or public challenge to safe and cost-beneficial VVIC if aided
by information needed by the public exposed to media distortion of scientific
debate, complex argument and concepts of probability and risk. Sound science
needs support from strategies for public processes to enable cost-beneficial
management of vertebrate pests.
Progress towards using recombinant myxoma virus as a vector for fertility control in rabbits
Authors: Robinson A. J. , Jackson R. , Kerr P. , Merchant J. , Parer I. , Pech R.
Published: November 2003

The history of myxoma virus, its use in Australia as a mortality agent and the
development of the virus as a vector for controlling fertility in wild rabbit
populations in Australia is reviewed. Myxoma virus recombinants have been
constructed to express model antigens. Four potential insertion sites in the
genome have been identified and two have been used to construct single and
double recombinant viruses expressing
Escherichia colienzymes β-galactosidase and
β-glucuronidase. Another recombinant expressing an influenza virus
haemagglutinin gene (A/PR8/34) induced high and sustained antibody
responses following intradermal inoculation in rabbits. To demonstrate the
potential of introducing a recombinant virus into wild rabbit populations, a
virus containing a natural deletion was released at four field locations.
Preliminary analysis of the data has shown that the introduced virus spread
well on 3 of the 4 locations. The steps being taken to address the ethical and
safety implications of the introduction of a recombinant virus into the field
are discussed.
Effects of passive immunization against oestradiol-17β and inhibin on the secretion of gonadotrophin in the cyclic golden hamster (Mesocricetus auratus)
Authors: Kishi Hisashi , Okada Toshiyuki , Kawazu Satoko , Otsuka Mariko , Taya Kazuyoshi , Watanabe Gen , Sasamoto Shuji
Published: November 2003

To investigate the physiological importance of oestradiol-17b and inhibin in
the regulation of gonadotrophin secretion in the cyclic golden hamster,
females were passively immunized against two hormones. When 200 µL
antiserum against oestradiol-17β (oestradiol-AS) was given on Day 3 (Day 1
= day of ovulation), the preovulatory gonadotrophin surge was
completely blocked for 24 h and the length of the oestrous cycle was also
prolonged for one day. In the group given 200µL oestradiol-AS on Day 3,
basal levels of follicle-stimulating hormone (FSH) and luteinizing hormone
(LH) increased slightly and superovulation (19·6 ± 0·8,
mean ± s.e.m.) occurred. When 200 µL antiserum against inhibin
(inhibin-AS) was given at 1100 hours on Day 3, a dramatic increase in plasma
FSH and a slight increase in LH were noted, resulting in superovulation
(38·2 ± 2·6) on the expected Day 1. The present study
indicates clearly that inhibin plays a major role in regulating the specific
ovulation rate in the hamster through the control of FSH secretion. Present
results also indicate that oestradiol-17β suppresses basal LH secretion.
Oestradiol-17β may act as an indicator of the follicular maturation, and
the high plasma concentration of oestradiol-17β noted from Day 3 to Day 4
may play an important role in determining the timing of initiation of the
preovulatory gonadotrophin surge.
Mitochondrial function and ram sperm fertility
Authors: Windsor D. P.
Published: November 2003

These experiments investigated the effect of freezing on mitochondrial
function in ram sperm, the effectiveness of current freezing procedures in
protecting mitochondria, and the role of mitochondrial respiration in cervical
penetration and transit by ram sperm. Only sperm with functioning mitochondria
(assessed by rhodamine 123 staining) after freezing and thawing were motile in
a viscous medium (P < 0· 05). A simplified
rhodamine 123 uptake assay was developed to monitor sperm mitochondrial
function. The results of this procedure were highly correlated
(r2 = 0 · 98) with
the proportion of damaged sperm in the semen sample. A semen freezing
procedure commonly used by industry was compared with newer methods, and with
freezing without cryoprotectants. None of the freezing protocols produced
sperm with higher post-thaw levels of mitochondrial integrity than unprotected
sperm. Merino ewes were inseminated with semen treated with metabolic
inhibitors. Glycolytic inhibition did not affect fertility. Mitochondrial
inhibition reduced fertility in cervically (P < 0
· 05), but not laparoscopically inseminated ewes. It is concluded that
mitochondrial respiration plays an important part in penetration of the cervix
by ram sperm. Mitochondrial injury during freezing is likely to be implicated
in the poor fertility of frozen ram semen used for cervical insemination.
Leukaemia inhibitory factor stimulates proliferation of prospermatogonial stem cells
Authors: Nikolova Diana B. , Martinova Yordanka S. , Seidensticker Martin , Bellvé Anthony R.
Published: November 2003

The effect of leukaemia inhibitory factor (LIF) on the proliferation of
prospermatogonial stem cells was testedin vitro. Pieces
of 3-day-old rat testis were cultured in the presence of a range of doses of
LIF alone or in combination with 1 ng mL -1 seminiferous
growth factor (SGF). Stimulation of the proliferative activity of quiescent
prospermatogonia was detected immunocytochemically with a cell proliferation
kit. After 24 h culture, LIF signicantly increased the percentage of labelled
prospermatogonia, with the peak of activity at 20 pg mL
-1 . The combination of LIF and SGF resulted in a
decrease in DNA synthetic activity of the germ cells. Hence, LIF and SGF play
a role in local regulation at the onset of spermatogenesis in the rat testis.
Angiotensin II as a semen extender component increases retention of spermatozoa within the uterus of the heifer
Authors: Grove K. L. , Speth R. C. , Senger P. L.
Published: November 2003

The effects of angiotensin II (Ang II) as a semen extender were studied. In
the first experiment, individual ejaculates from 10 bulls were split and
extended in egg yolk citrate in the absence or presence of varying
concentrations of Ang II (10-5
–10-10 M) to a final concentration of
35×106 sperm per mL. The percentage of intact
acrosomes and percentage motility were determined in all treatments for all
bulls at 0 h (immediately post thaw) and after incubation for 4 h at 37°C.
Extension of the semen with Ang II did not affect spermatozoal viability at
either time studied. In the second experiment, mixed breed virgin heifers were
induced into oestrus with intramuscular injections of prostaglandin
F2α on Days 0 and 3. Animals that stood to be
mounted were paired for bilateral intracornual insemination using a
0·5-mL French straw on each side approximately 8 h later. One of the
paired heifers received semen containing Ang II (10-5 M)
while the other received control semen. A 1-mL aspirate of vaginal mucus was
collected at hourly intervals for 8 h after insemination. Concentration of
spermatozoa was determined by haemocytometry. There was a significant
reduction in cumulative semen loss into the vagina of heifers inseminated with
Ang II extended semen (14·4%) compared with heifers inseminated
with control semen (19 ·7%). This suggests that Ang II, when
added to extended semen, may reduce retrograde sperm loss following
insemination without affecting sperm viability.
Isolation, characterization and radioimmunoassay of luteinizing hormone in the brushtail possum
Authors: Moore L. G. , Chie W. Ng , Lawrence S. Lun, S. B. , Heath D. A. , McNatty K. P.
Published: November 2003

Luteinizing hormone (LH) was purified from brushtail possum
(Trichosurus vulpecula) pituitary glands. The
purification procedure consisted of ammonium sulfate precipitation followed by
triazinyl-dye chromatography, hydrophobic interaction chromatography and gel
filtration. A yield of 10 µg LH g-1 pituitary with
a recovery of 20% was obtained from 1400 pituitary glands (20·3
g). Contamination with possum follicle-stimulating hormone (FSH) was
≤0.05%. The amino acid analysis and the
N-terminal sequencing for 10 cycles revealed close
homology with LH from other mammals. Minor amounts of LH that had been
truncated near the N-terminal were also detected. No contaminating proteins
were found by amino acid sequencing. The potency of possum LH was 20%
that of ovine LH in a receptor assay using possum testicular receptors and
4% that of ovine LH when bovine corpora lutea receptors were used.
Possum LH was able to stimulate production of cyclic adenosine 3′
,5′-monophosphate by bovine granulosa cells. A radioimmunoassay (RIA)
for possum LH using 125I-possum LH and an antiserum
raised against ovine LH was developed. The RIA has a sensitivity of
0·15 ng mL-1 , a 50% displacement of
1·9 ng mL-1 and a cross-reactivity of <0
· 02% against possum FSH. Plasma concentrations were 0·24
± 0· 04 ng mL-1 (n
= 8) and 0·39 ± 0·12 ng mL-1
(n = 8) in female and male possums respectively.
Administration of mammalian gonadotrophin-releasing hormone (GnRH) and chicken
GnRH II stimulated increases in plasma LH concentrations in male and female
possums. When comparing LH responses with administration of mammalian GnRH or
chicken GnRH II, plasma LH concentrations remained elevated for a longer
period of time in males than in females (P <
0· 01); plasma LH concentrations also remained elevated for longer
after mammalian GnRH than after chicken GnRH II (P <
0· 01). Gonadectomy stimulated an increase in plasma concentrations of
LH in both male (P < 0· 01) and female
(P < 0· 05) possums. The rate of increase in
plasma LH concentrations in males was faster than that in females. In summary,
we have purified, partially characterized, and developed a RIA for possum LH.
Excretion profiles of some reproductive steroids in the faeces of captive Nepalese red panda (Ailurus fulgens fulgens)
Authors: Spanner A. , Stone G. M. , Schultz D.
Published: November 2003

Faecal samples were collected up to once daily from three female Nepalese red
panda (Ailurus fulgens fulgens) prior to, during, and
after the expected 1995 breeding season. Radioimmunoassay of faecal progestins
and oestrogens showed hormone profiles that suggest that this species is a
seasonally polyoestrous, induced ovulator. Examination of faecal extracts
following high pressure liquid chromatography and using antisera of high
specificity, demonstrated at least five faecal progestins but only one major
oestrogen, probably oestradiol. Samples from males were collected up to once
weekly during the breeding season in 1996, and androgens measured by
radioimmunoassay. Faecal androgens fluctuated widely for all males.
Vitrication of in vitro produced bovine embryos at different ages using one- and three-step addition of cryoprotective additives
Authors: Saha S. , Suzuki T.
Published: November 2003

The effect of embryo age on development and ratio of live : dead cells after
vitrication and warming was examined. One-step and three-step addition of
cryoprotectants in vitrification solution (40% ethylene glycol,
0·3 M trehalose and 12% polyvinylpyrrolidone) were compared
usingin vitro produced (IVP) bovine blastocysts and
expanded blastocysts. Rates of development and hatching were 74
·2% and 41· 9% for Day 7, 57·8% and
23· 8% for Day 8, 33· 7% and 6·1%
for Day 9 embryos with one-step addition. In three-step addition, those rates
were 86·2% and 77·3% for Day 7,
72·3% and 39·0% for Day 8, 47·3% and
10·5% for Day 9 embryos. Day 7 embryos showed highest
(P < 0·01) development and hatching rates with
one exception. Hatching rate of Day 7 embryos with three-step addition was
higher (P < 0·01) than with one-step addition.
The ratio of live : dead cells differed between one-step (94%) and
three-step (97%) additions for Day 7 embryos (P < 0· 05). The
results indicate the higher resistance of younger IVP bovine embryos against
vitrication and the potential for three-step addition of cryoprotectants to
yield a higher survival rate after warming than with one-step addition.
Postnatal development and differentiation of myobres in functionally diverse porcine skeletal muscles
Authors: Harrison A. P. , Latorre R. , Dauncey M. J.
Published: November 2003

Marked changes in muscle function occur after birth, with the response being
dependent on developmental stage. Therefore, postnatal cellular ontogeny of
functionally distinct skeletal muscles was investigated in the pig, a large
mammal born at a relatively advanced stage of development. Assessment of
myofibre contractile (type I slow/type II fast) and metabolic
(oxidative/glycolytic) properties at Days 0, 2, 5 and 14 revealed
type-specific differences in hypertrophy and differentiation. Type I bre
proportions increased significantly in soleus and
diaphragm, especially between Days 0 and 5, and
rhomboideus showed a similar trend, but in
longissimus there was a slight decrease during Days 0-2.
Cytochrome oxidase activity was relatively high and similar among myofibres in
all muscles at birth, and fibres with low activity were not detected until Day
5. In contrast with previous reports, glycolytic fibres were present in all
muscles at birth; postnatal changes in α-glycerophosphate dehydrogenase
activity were both muscle- and myofibre-specific. Hence, although myosin
ATPase activity and metabolic properties of porcine myobres are well developed
at birth, they continue to mature postnatally. This suggests that postnatal
muscle development can be modulated by extrinsic factors, even in mammals born
at a relatively advanced stage of development.
Metabolic activity of hypotonically treated mature boar spermatozoa
Authors: Jones A. R.
Published: November 2003

Treatment of washed boar sperm with hypotonic phosphate buffer removed the
acrosome, disrupted the cytoplasmic membrane and almost completely separated
the heads from the mid piece-tail segment. As assessed by oxygen uptake
studies and their ability to oxidize14C-labelled
substrates to 14CO2,
hypotonically-treated cells exhibit low glycolytic activity yet mitochondrial
activity remains high. Both lactate and glycerol 3-phosphate underwent
oxidation and these substrates continued to be metabolized by this preparation
which had been stored for up to 10 days at 4°C. Such preparations may be
of assistance in the investigation of the biochemistry of boar sperm
mitochondria.
Oviducal structure in four species of gekkonid lizard differing in parity mode and eggshell structure
Authors: Girling J. E. , Cree A. , Jr L. J. Guillette,
Published: November 2003

Oviducal structure was analysed in vitellogenic females from four species of
gekkonid lizard exhibiting variation in parity mode and eggshell structure:
Hemidactylus turcicus (oviparous) which produces a hard,
calcareous eggshell; Saltuarius wyberba (oviparous)
which produces a soft, parchment-like eggshell; and
Hoplodactylus maculatus and
Hoplodactylus duvaucelii (both viviparous). Oviducts
were analysed by light, scanning electron and transmission electron
microscopy. The uterus exhibited differences among species that were directly
attributable to parity mode. H. turcicus and
S. wyberba (oviparous) had numerous uterine shell
glands; H. maculatus and
H. duvaucelii(viviparous) had very few. The uterus also
exhibited differences between the two oviparous species
(H. turcicusand S. wyberba) which
may be related to the type of eggshell produced. Variations were noted in the
staining properties of the uterine glandular and epithelial cells. The
structure of the infundibulum, uterine tube, isthmus and vagina also differed
among species, but differences could not be directly related to parity mode or
eggshell structure. Instead, the differences may be related to how prepared
the oviduct is for ovulation in individuals analysed from the different
species. This study confirms, in the Gekkonidae, aspects of oviducal structure
that have been associated with parity mode in other squamate taxa.
Increased prostaglandin E generation and enhanced nitric oxide synthase activity in the non-insulin-dependent diabetic embryo during organogenesis
Authors: Jawerbaum Alicia , Gonzalez Elida T. , Novaro Virginia , Faletti Alicia , Sinner Debora , Gimeno Martha A. F.
Published: November 2003

Embryonic development, prostaglandin E (PGE) generation and nitric oxide
synthase (NOS) activity during organogenesis were evaluated in an experimental
rat model of non-insulin- dependent diabetes (NIDD) generated by neonatal
administration of streptozotocin. Gross malformations were detected in
5% of NIDD embryos and these embryos were all non-viable; in the other
95%, growth was retarded but no congenital abnormalities were found.
Control embryos were all alive and not malformed. The NIDD 11-day embryos
secreted more PGE into the incubation medium than did controls. The NO donor
SIN–1 increased PGE production in both control and NIDD embryos. A NOS
inhibitor (L-NMMA) reduced PGE generation in both experimental groups,
suggesting a modulatory role of NO on embryonic PGE production. Activity of
NOS was higher in NIDD 11-day embryos than in controls. Treatment
in vivo of control and NIDD rats (Days 7–11 of
gestation) with a NOS inhibitor (L-NAME; 5 mg kg-1 i.p.)
reduced embryonic PGE production and induced a higher resorption rate and an
increase in neural-tube defects. The results suggest that NO modulates PGE
generation in the organogenetic embryo. In the NIDD model, overproduction of
NO is observed, this NO probably enhancing embryonic PGE production. The
relationship between PGE generation and the appearance of congenital
abnormalities is discussed.
Properties and uses of embryonic stem cells: prospects for application to human biology and gene therapy
Authors: Rathjen P. D. , Lake J. , Whyatt L. M. , Bettess M. D. , Rathjen J.
Published: November 2003

Embryonic stem cells are pluripotent cells derived from the early mouse embryo
that can be propagated stably in the undifferentiated state
in vitro. They retain the ability to differentiate into
all cell types found in an embryonic and adult mouse
in vivo, and can be induced to differentiate into many
cell types in vitro. Exploitation of ES cell technology
for the creation of mice bearing predetermined genetic alterations has
received widespread attention because of the sophistication that it brings to
the study of gene function in mammals. Analysis of cell differentiation
in vitro has also been of value, leading to the
identification of novel bioactive factors and the elucidation of cell
specification mechanisms. In this paper, we summarise the features of
pluripotent cell lines and their applications, foreshadowing the impact that
these systems may have on human biology. While the isolation of definitive
human pluripotent cell lines has not yet been achieved, potential applications
for these cells in the study of human biology, particularly cell
specification, can be envisaged. Of particular interest is the possibility
that human embryonic stem cells with properties similar to mouse embryonic
stem cells might provide a generic system for gene therapy.
Antiandrogens as environmental endocrine disruptors¤
Authors: Kelce W. R. , Gray L. E. , Wilson E. M.
Published: November 2003

Steroid hormone receptors control fundamental events in embryonic development
and sex differentiation through their function as ligand-inducible
transcription factors. The consequences of disrupting these processes can be
especially profound during development due to the crucial role hormones play
in controlling transient and irreversible developmental processes. Several
environmental chemicals, including metabolites of the fungicide vinclozolin
and the pesticide DDT, disrupt male reproductive development and function by
inhibiting androgen receptor mediated events. A variety of
in vitro and in vivo approaches
have been used to determine the molecular basis of environmental antiandrogen
toxicity. These chemicals commonly bind androgen receptor with moderate
affinity and act as antagonists by inhibiting transcription of androgen
dependent genes.
Loss of livestock breeding efficiency due to uncompensable sperm nuclear defects
Authors: Evenson D. P.
Published: November 2003

An important goal of modern analyses of semen is to elucidate the molecular
traits of mammalian sperm chromatin structural abnormalities, defined here as
‘uncompensable’, that lead to abnormalities in fertility,
pronuclear formation, early embryo quality and pregnancy outcome. Sperm with
uncompensable nuclear abnormalities are able to fertilize oocytes both
in vivo and in vitro; however, due
to the uncompensable trait(s), the embryo development may be abnormal.
Uncompensable nuclear traits can be experimentally induced in bull sperm by a
mild thermal insult to the testis. Sperm nuclear morphology abnormalities seen
in ejaculates 11-days post stress are likely related to molecular changes in
chromatin observed 3-days post stress by the flow cytometric sperm chromatin
structure assay (SCSA). The SCSA measures the susceptibility of sperm nuclear
DNA to denaturation in situ. This susceptibility has
been correlated with the presence of DNA strand breaks that may be derived in
part by oxidative stress and possibly by a unique, abortive apoptotic
mechanism. The extent of DNA denaturation is not significantly related to the
level of disulfide bonding between the chromatin protamines. The use of human
sperm with uncompensable nuclear traits for artificial reproductive techniques
is also discussed. The goal of this research is to remove from semen doses
those sperm with uncompensable nuclear traits and thereby increase male
fertility potential.
Extra key words: male fertility potential, sperm
chromatin structure assay (SCSA).
Cloning sheep from cultured embryonic cells
Authors: Wells D. N. , Misica P. M. , Day A. M. , Peterson A. J. , Tervit H. R.
Published: November 2003

The production of transgenic farm animals will be greatly enhanced with the
development of cultured cell lines that remain totipotent following nuclear
transfer. Here, data are presented that demonstrate the generation of both
male and female cloned lambs from two established embryonic cell lines.
Cytoplasts derived from in vivo oocytes resulted in
slightly greater development to blastocyst (24%
v. 17%) and survival to term (7%
v. 2%) compared with
in vitro oocytes. There was no advantage in co-culturing
cloned embryos with oviductal epithelial cells compared with synthetic
oviductal fluid medium in terms of development to blastocyst (18%
v. 31%) or survival to term (both 8%).
Although the survival of cloned embryos immediately after transfer was high
based on ‘biochemical’ pregnancy, 64–80% of embryos
failed over the attachment phase with in vivo
cytoplasts. Although the co-transfer of trophoblastic vesicles improved embryo
survival to Day 35 (45% v. 25%), there was
no difference at term. A high proportion of fetuses were lost during the last
trimester (43%), resulting in 11% of embryos transferred
developing to term using in vivo cytoplasts
(12/112). Five lambs have survived and two rams are fertile. The current
nuclear transfer process is inefficient and further research is needed to
improve the development of healthy fetuses.
Cytoplasmic control of nuclear assembly
Authors: Collas Philippe
Published: November 2003

The reconstitution of a replication-competent, transcriptionally active
nucleus following mitosis, fertilization or nuclear transplantation involves a
stepwise series of reactions, most (if not all) of which are controlled by the
cytoplasmic environment. This review discusses the nature of cytoplasmic
contributions to the development of the male pronucleus at fertilization, and
the effect of altering the cytoplasmic environment on nuclear assembly. The
system used to investigate these regulations consists of permeabilized sea
urchin sperm nuclei incubated under controlled conditions in a cell-free
extract of fertilized sea urchin eggs. (1) In egg
cytoplasmic extract, male pronuclear formation is initiated by the disassembly
of the sperm nuclear lamina as a result of lamin phosphorylation by a
cytosolic protein kinase C. (2) Sperm histones are
phosphorylated by an as yet unidentified soluble kinase.
(3) The conical sperm nucleus decondenses into a
spherical pronucleus in an ATP-and cytosolic pH-dependent manner.
(4) Chromatin decondensation is associated with the
replacement of sperm histones by maternal histones. (5)
Nuclear membranes form by ATP-dependent binding of vesicles to chromatin and
GTP-dependent fusion of these vesicles to one another.
(6) Three cytoplasmic vesicle populations with distinct
biochemical, chromatin-binding and fusion properties are required for nuclear
envelope assembly. (7) Targeting of the bulk of nuclear
membrane vesicles to chromatin is mediated by an integral membrane protein
similar to human lamin B receptor. (8) The last step of
male pronuclear formation, nuclear swelling, is promoted by the assembly of
nuclear pores, nuclear import of soluble lamins and growth of the nuclear
membranes. (9) Once inside the nucleus, lamin B
associates with lamin B receptors, presumably to tether the inner nuclear
membrane with the lamina. Overall, these processes are similar to those
characterizing nuclear reconstitution after mitosis in somatic cells or
nuclear remodeling following transplantation into oocytes or eggs. The
influence of the egg cytoplasmic environment on some aspects of nuclear
remodeling after nuclear transplantation is also discussed.
Large-scale applications of cloning technologies for agriculture: an industry perspective
Authors: Lewis I. M. , Peura T. T. , Trounson A. O.
Published: November 2003

The present costs and efficiencies of producing cloned embryos, pregnancies
and offspring using the simplified nuclear transfer techniques developed in
the authors’ laboratories are compared with those required for the
large-scale application of such cloning technologies in cattle. The current
costs in the laboratory of producing large numbers of genetically identical
cloned embryos for transfer is around $15.00 per blastocyst, which is
within the cost estimated to be commercially viable for cloned female dairy
embryos for transfer. However, the pregnancy and calving rates from the
transfer of such embryos are still well below that required for large-scale
commercial application for which ongoing pregnancy rates of at least
50% per recipient will be required. The current pregnancy rate
(30–40 days post-transfer) following the transfer of an average of three
cloned embryos per recipient is 37%, and the calving rate 17%,
representing high losses between pregnancy diagnosis and term. In the beef
industry and in some dairy situations the final product (cloned bulls for
natural mating) will have a much higher inherent value and different
parameters will therefore apply. Recent developments in the technologies that
are likely to increase the probabilities of large-scale application are
discussed, including recycling nuclear transfer embryos, somatic cell cloning,
new cryopreservation techniques and automated oocyte harvesting.
Post-natal growth and development of Simmental calves derived from in vivo or in vitro embryos
Authors: McEvoy T. G. , Sinclair K. D. , Broadbent P. J. , Goodhand K. L. , Robinson J. J.
Published: November 2003

Large fetuses arising from embryos produced in vitro
have been shown to exhibit altered organ development
in utero, but it is not known whether this persists post
natally. Post-natal growth and development was examined in 18 Simmental bulls
derived from in vivo frozen–thawed
(n = 6), in vitro
frozen– thawed (n = 6) or
in vitro fresh (n = 6)
embryos and reared together post weaning on an
ad libitum diet until slaughter at approximately 13
months old. Calves weighing less than 60 kg at birth (n
= 11) were classified as normal, and heavier calves
(n = 7; all from in vitro
embryos) as oversize. Lifetime growth rates and slaughter weights apparently
were unaffected by embryo source or birthweight. Mean (± s.e.m.) post
mortem liver and kidney weights were unaffected by embryo source, but hearts
of bulls from in vitro frozen embryos were heavier than those of bulls from
in vivo frozen embryos (2.7 ± 0.04
v. 2.3 ± 0.07 kg,
P<0.025). Heart weight per kilogram body weight at
slaughter for the 7 perinatally oversize males (4.01 ± 0.08 g) exceeded
that of the other 5 bulls from in vitro embryos (3.60
± 0.10 g kg −1 ;
P<0.04) and the 6 in vivo males
(3.56 ± 0.12 g kg −1 ;
P<0.02). Overall, one-third of the variation in heart
weight at slaughter (r 2 =
0.35; P = 0.01) was due to variation in
birthweight. This is the first study to demonstrate birthweight-related
developmental effects on post-natal organ weight following the transfer of
embryos produced in vitro.
Kinematic definition of ram sperm hyperactivation
Authors: Mortimer S. T. , Maxwell W. M. C.
Published: November 2003

Although it is known that ram spermatozoa exhibit hyperactivated motility
under capacitating conditions, quantitative analyses of the head and flagellar
movement of washed ram spermatozoa have not been published. Motile spermatozoa
were recovered from semen by swim-up into HSOF medium, and their movement in
30-µm-deep chambers was videorecorded. Spermatozoa of interest were
identified during tape playback (hyperactivated spermatozoa were identified by
visual assessment of flagellar movement) and sequential head and tail images
were traced onto overhead projector film attached to the video monitor. The
flagellar movement characteristics beat angle (FBA), beat envelope (FBE) and
curvature ratio (FCR) were determined by first principles, and head centroid
kinematics were determined using Cartesian methods. Hyperactivated spermatozoa
had significantly higher FBA and FBE and significantly lower FCR values than
non-hyperactivated spermatozoa (all P<0.0001). The
centroid kinematic values were also found to be significantly different, and
kinematic criteria for ram sperm hyperactivation were developed. These
criteria were refined by consideration of 60-Hz CASA-derived trajectories, and
ram sperm hyperactivation was defined by: VCL > 250.0 µm s
−1 and VSL ≤ 100.0 m s
−1 and LIN ≤ 30% and ALHmax ≥ 9.0
µm.
Effects of pre- and post-mating feed intake on blastocyst size, secretory function and glucose metabolism in Meishan gilts
Authors: Ashworth C. J. , Beattie L. , Antipatis C. , Vallet J. L.
Published: November 2003

This experiment was designed to determine the
effects of a nutritional regime, known to increase embryo survival, on
blastocyst development and function. Day 12 blastocysts were recovered from
Meishan gilts allocated in a 2¥2 factorial design to receive either a high
or a maintenance diet before or after mating (n =
4–6 gilts per group). The post-mating diet had no effect on individual
blastocyst size, cell number, secretion of oestradiol-17b or retinol binding
protein, glucose metabolism or on the within-litter variability in these
measures. Blastocysts recovered from gilts consuming the high pre-mating diet
had more cells (13.501 v. 13.006 log cells; SED =
0.23; P = 0.05), greater production of CO
2 from glucose (2.19 v. 1.23 log
pmol –1 blastocyst –1
3 h –1 , SED = 0.42;
P = 0.05) and a lower within-litter standard
deviation in blastocyst surface area (0.66 v. 1.18 log
mm 2 , SED = 0.24; P
= 0.04) compared with gilts fed the maintenance pre-mating diet.
Collectively, these data suggest that a nutritional strategy that increases
embryo survival is also associated with an increase in individual blastocyst
cell number and reduced within-litter variability in blastocyst size.
Normal fertility in ewes after cervical insemination with frozen - thawed spermatozoa supplemented with seminal plasma
Authors: Maxwell W. M. C. , Evans G. , Mortimer S. T. , Gillan L. , Gellatly E. S. , McPhie C. A.
Published: November 2003

The effect of seminal plasma (SP) on the motility, capacitation status,
penetration through cervical mucus and fertility of frozen–thawed ram
spermatozoa was examined. In the presence of SP, motility of
frozen–thawed spermatozoa was better (P<0.001)
and there were more uncapacitated and less acrosome-reacted cells in
comparison with controls (P<0.001).
Frozen–thawed spermatozoa were also better able to penetrate cervical
mucus after addition of SP. Addition of SP increased the percentage of ewes
pregnant after insemination of frozen–thawed (39/94, 41.5%
v. 51/92, 55.4%;
P<0.05) but not fresh spermatozoa (34/55,
61.8% v. 42/58, 72.4% for 0
v. 30% SP in the resuspension medium). Moreover,
SP improved pregnancy rates after cervical (14/50; 28%
v. 25/49; 51%;
P<0.05) but not intrauterine insemination (25/44;
56.8 v. 26/43; 60.5%) with
frozen–thawed spermatozoa. In a second experiment, pregnancy rates were
30/45 (66.7%), 9/37 (24.3%) and 24/40 (60.0%)
for ewes inseminated with frozen–thawed spermatozoa in the uterus
(control), cervix without SP and cervix after supplementation with SP,
respectively (P<0.01 for unsupplemented
v. supplemented spermatozoa). These experiments
demonstrate that impaired function of cryopreserved spermatozoa can be
overcome by addition of SP, resulting in normal fertility after cervical AI.
Intrauterine growth retardation is associated with reduced cell cycle activity, but not myofibre number, in ovine fetal muscle
Authors: Greenwood Paul L. , Slepetis Ramona M. , Bell Alan W. , Hermanson John W.
Published: November 2003

Cellular development of muscle was studied in
sheep fetuses at 85 days of gestation. Large and small fetuses were compared
at 100, 115 and 130 days, and an additional group of large 130-day fetuses
were studied following 7 days of maternal undernutrition. Myogenesis in the
peroneus longus muscle was completed between 100 and 115
days of gestation, and myofibre number did not differ between small and large
fetuses. The proportion of myofibre-related nuclei identified as entering
S-phase of the cell cycle was 1.7% per hour in 85-day fetuses. In large
fetuses, subsequent rates were relatively constant (=1.5% h
–1 ), whereas in small fetuses cell cycle activity
declined with age from 1.3 to 0.9% h –1 ,
and was 0.5 % h –1 in 130-day fetuses of
restricted ewes. The constant rate of cell cycle activity in large fetuses was
associated with an increasing estimated rate of muscle growth
(peroneus longus (mg) = 0.831 ¥ 10
(0.024 ¥ age [d]) ,
r 2 = 0.98), which
contrasted with slow and relatively constant muscle accretion in small fetuses
(8.4 mg day –1 ), and slower muscle accretion at
130 days in large fetuses from restricted ewes. Differences in DNA and RNA
content in the semimembranosus muscle increased with
age, large fetuses having 70% more muscle DNA, 108% more muscle
RNA and 104% larger muscles than small fetuses at 130 days (all
P<0.001). The results demonstrate that myonuclei
accumulation, but not myofibre number, is associated with fetal growth in
sheep and, therefore, with fetal nutrition during mid to late gestation.
Origin and ploidy of multipronuclear zygotes
Authors: Trounson Alan O. , Tarín Juan J. , Sathananthan Henry
Published: November 2003

Recently, several authors have proposed
strategies for correction of triploidy based on the removal of the extra
pronucleus at the zygote stage. In the present bioassay, the following were
analysed: (1) the different factors that can induce the
formation of multipronuclear zygotes in mammals; (2) the
different morphological patterns established according to the number of
pronuclei and polar bodies that can be observed at the zygote stage and used
to distinguish the origin of multipronuclear zygotes; and
(3) the pattern of chromosomal segregation during the
first mitotic division and ploidy status of the resulting preim-plantation
embryos. Such an analysis shows that the morphological criterion of counting
the number of pronuclei and polar bodies can be misleading and should not be
used for ascertaining the origin of tripronu-clear zygotes. In addition,
although monospermic digynic zygotes have a single sperm centromere, which
likely organizes a single bipolar spindle during the first mitotic division,
more data on chromosomal distribution and segregation of digynic tripronuclear
zygotes should be collected before strategies for correction of triploidy are
implemented in humans.
Fate and functional integrity of fresh and frozen - thawed ram spermatozoa following intrauterine insemination
Authors: Gillan L. , Skovgold K. , Watson P. F. , Maxwell W. M. C. , Evans G.
Published: November 2003

Ewes in a synchronized oestrus were
inseminated (intrauterine) with fresh and frozen–thawed spermatozoa and
the spermatozoa were either recovered from each section of the reproductive
tract after the animal was killed (Experiments 1a and 1b) or after they were
voided from the cervix (Experiment 2). In Experiment 1a, only 1.2 0.27%
of the original inseminate was recovered. Placing a ligature at the base of
the uterine horn in Experiment 1b led to the recovery of 3.0 0.33% of
the original inseminate, located mainly in each uterine horn (33.1
5.48%), and each isthmic and ampullary region of the oviduct (2.9
5.48% and 4.0 5.48%, respectively). A higher proportion of
spermatozoa recovered from the isthmus were uncapacitated when observed by
chlortetracycline staining than those recovered from the uterus (26.4
1.92% and 15.6 1.92%, respectively,
P<0.05). Experiment 2 showed that large proportions
of spermatozoa were voided from the tract through the vagina, with similar
numbers of fresh and frozen–thawed spermatozoa lost from the tract.
However, frozen–thawed spermatozoa were lost at a faster rate than fresh
(P<0.05) and with a more advanced membrane state
(66.8 1.30% and 53.2 1.30% were acrosome reacted respectively;
P<0.001). Large numbers of recovered spermatozoa had
lost their tails, with frozen–thawed spermatozoa more susceptible to
tail loss than fresh spermatozoa (55.0 0.96% and 45.5 0.96%
respectively; P<0.05).
Physiological β Cell Death Triggers Priming of Self-reactive T Cells by Dendritic Cells in a Type-1 Diabetes Model
Authors: Turley Shannon , Poirot Laurent , Hattori Masakazu , Benoist Christophe , Mathis Diane
Published: November 2003

The prelude to type-1 diabetes is leukocyte infiltration into the pancreatic islets, or insulitis. This process begins in pancreatic lymph nodes when T lymphocytes reactive to islet β cells encounter antigen-presenting cells (APCs) displaying peptides derived from β cell proteins. We show here that a ripple of physiological β cell death, which occurs at 2 wk of age in all mouse strains, precipitates the arrival of such APCs, and that the relevant APC is a dendritic cell of CD11c+CD11b+CD8α− phenotype. These findings have significant implications concerning the nature of the diabetes-provoking deficits in NOD mice, the identity of the primordial diabetogenic antigens, and our understanding of the balance between immunity and tolerance in a pathological context.
A Common Dominant TLR5 Stop Codon Polymorphism Abolishes Flagellin Signaling and Is Associated with Susceptibility to Legionnaires' Disease
Authors: Hawn Thomas R. , Verbon Annelies , Lettinga Kamilla D. , Zhao Lue Ping , Li Shuying Sue , Laws Richard J. , Skerrett Shawn J. , Beutler Bruce , Schroeder Lea , Nachman Alex , Ozinsky Adrian , Smith Kelly D. , Aderem Alan
Published: November 2003

Although Toll-like receptors (TLRs) are critical mediators of the immune response to pathogens, the influence of polymorphisms in this gene family on human susceptibility to infection is poorly understood. We demonstrated recently that TLR5 recognizes flagellin, a potent inflammatory stimulus present in the flagellar structure of many bacteria. Here, we show that a common stop codon polymorphism in the ligand-binding domain of TLR5 (TLR5392STOP) is unable to mediate flagellin signaling, acts in a dominant fashion, and is associated with susceptibility to pneumonia caused by Legionella pneumophila, a flagellated bacterium. We also show that flagellin is a principal stimulant of proinflammatory cytokine production in lung epithelial cells. Together, these observations suggest that TLR5392STOP increases human susceptibility to infection through an unusual dominant mechanism that compromises TLR5's essential role as a regulator of the lung epithelial innate immune response.
Corporate budget planning, control and performance evaluation in Bahrain
Authors: Joshi P. L. , Al‐Mudhaki Jawahar , Bremser Wayne G.
Published: November 2003

Examines budget planning; implementation and performance evaluation practices by utilizing a questionnaire survey of 54 medium and large sized companies located in Bahrain. Most of the companies prepare long‐range plans and operating budgets, and they follow a definite budget procedure and implementation methodology. Uses budget variances to measure a manager’s ability, for timely recognition of problems, and to improve the next period’s budget. While both the listed and non‐listed companies have reported many similar budget practices, the main differences were specific purposes served by budgets, degree of budget participation, periodicity of variance reporting, and purposes and authority to evaluate budget variance reports. In certain cases, firm size influences budgeting practices. Contributes toward filling a gap in the literature on the use of budgets as a planning and control tool in developing countries. Most prior studies were mainly confined to advanced countries. The study findings suggest the need for research on attitudes held by the budgetees towards the use of budget variances in the context of advanced management accounting techniques.
Public Investment and the Risk Premium for Equity
Authors: Grant Simon , Quiggin John
Published: November 2003

Analysis of the equity premium puzzle has focused on private‐sector capital markets. However, the existence of an anomalous equity premium raises important issues in the evaluation of public‐sector investment projects. These issues are explored below. We begin by formalizing the argument that an equity premium may arise from uninsurable systematic risk in labour income, and show that, other things being equal, increases in public ownership of equity will improve welfare, up to the point where the equity premium is eliminated. Finally, we consider policy implications and the optimal extent of public ownership.
A Semi‐nonparametric Approach to the Demand for UK Monetary Assets
Authors: Drake Leigh , Fleissig Adrian R. , Swofford James L.
Published: November 2003

We estimate an asymptotically ideal model of the demand for UK personal sector monetary assets. We use data that are consistent with utility‐maximizing behaviour, and find that UK monetary assets are generally substitutes in use. The estimated elasticities of substitution during the 1980s and the early 1990s indicate that a relatively broad monetary aggregate should be used in economic studies. The results also suggest that any policy based on interest or user cost elasticities of substitution between financial assets should be based on the Morishima elasticities, as the Allen–Uzawa calculation can give misleading results.