bluuebunny/crossref_metadata_2025 huggingface dataset:
B‐mode and Contrast‐Enhanced Ultrasonographic Findings in Canine Prostatic Disorders
Authors: Russo M , Vignoli M , England GCW
Published: December 2012

Contents

Ultrasound is a common imaging technique used for examination of the dog's prostate gland that has proven to be successful for diagnosis of many clinical conditions. Whilst B‐mode ultrasound is useful for assessment of the prostate gland, there appears to be substantial variation in the appearance of some pathological conditions, although individual experienced ultrasonographers often have a high aptitude for making a correct diagnosis. Here, we report the normal volume and appearance of the canine prostate gland with B‐mode ultrasound and use thematic analysis to identify the categories described by experienced ultrasonographers when reporting both normal and abnormal findings. Four thematic categories were identified: background echotexture, parenchymal stippling, generalized appearance and focal changes. We found that a relatively narrow variety of thematic variables (descriptors) were used for reporting of prostatic pathology within these thematic categories. There was also poor association between these descriptors and the underlying pathology, with the only unique descriptors being ‘cysts’ for benign prostatic hyperplasia and ‘parenchymal mineralization’ for adenocarcinoma. In comparison with the limitations of B‐mode ultrasound, we document the value of measurement of vascular perfusion kinetics using contrast‐enhanced ultrasound and demonstrate how perfusion kinetics differ between benign and malignant prostatic disease.
Cellular Characteristics of Cultured Canine Trophoblasts
Authors: Sahlfeld L , Hazzard T , Kutzler M
Published: December 2012

Contents

Shallow trophoblast invasion is detrimental in human pregnancies, but represents normal endotheliochorial placentation in dogs. Factors regulating shallow trophoblast invasion into the canine decidua are not well described, but it is known that matrix metalloproteinases (MMPs) play a crucial role in trophoblast invasion in many species. Following the methods previously described for isolating human trophoblasts, canine trophoblasts were isolated using collagenase and trypsin digestions with Percoll density gradient centrifugation. In addition, placental pieces were cryopreserved prior to primary culture following methods previously described for human tissue. Expression of cytokeratin‐7, MMP2 and MMP9 was confirmed using fluorescent immunocytochemistry. Cellular morphology was similar to that reported for trophoblasts. More than 97% of the cells cultured expressed cytokeratin‐7. More cultured canine trophoblasts expressed MMP9 (54.7 ± 3.4%) compared with MMP2 (40.3 ± 1.8%) (p = 0.02). Although both MMPs were immunolocalized to the cytoplasm, MMP2 was found in large, coalescing granules, whereas MMP9 was more diffusely expressed throughout the cell. Cryogenic freezing of placental tissue prior to primary cell culture had no effect on cell proliferation (p = 0.37). This research has established a baseline for future studies investigating the canine placenta as a model for disorders of shallow trophoblast invasion in humans.
Generation and Characterization of Phage‐GnRH Chemical Conjugates for Potential Use in Cat and Dog Immunocontraception
Authors: Samoylov A , Cox N , Cochran A , Wolfe K , Donovan C , Kutzler M , Petrenko V , Baker H , Samoylova T
Published: December 2012

Contents

Overpopulation of cats and dogs is a serious worldwide problem that demands novel, safe and cost‐effective solutions. The objective of this study was to generate and characterize phage‐peptide conjugates with gonadotropin‐releasing hormone (GnRH) for potential use as an immunocontraceptive. A filamentous phage vector f5‐8 with wild‐type phage coat proteins was used as a carrier for construction of chemical conjugates with GnRH, a peptide that acts as a master reproductive hormone. In such conjugates, the phage body plays the role of a carrier protein, while multiple copies of GnRH peptide stimulate production of neutralizing anti‐GnRH antibodies potentially leading to contraceptive effects. To generate the constructs, four different GnRH‐based peptides were synthesized and conjugated to phage particles in a two‐step procedure: (i) peptides were reacted with phage to form a conjugate using 1‐ethyl‐3‐[3‐dimethylaminopropyl]carbodiimide hydrochloride chemistry (EDC) and (ii) the conjugates were separated from remaining free peptides by dialysis. Formation and specificity of phage‐GnRH conjugates were confirmed by three independent methods: spectrophotometry, electron microscopy and ELISA. When the conjugates were tested for interaction with sera collected from cats and dogs immunized with GnRH‐based vaccines in independent studies, strong specific ELISA signals were obtained, suggesting the potential use of the conjugates for cat and dog immunosterilization. The ability of the conjugates to stimulate production of anti‐GnRH antibodies in vivo was evaluated in mice. While optimization of dose, immunization route and adjuvant still requires investigation, our preliminary results demonstrated the presence of anti‐GnRH antibodies in sera of mice immunized with such conjugates. Fertility trials in cats and dogs will be needed to evaluate contraceptive potentials of the phage‐GnRH peptide chemical conjugates.
Early Canine Pregnancy – A Battle for Successful Growth and Angiogenesis
Authors: Schäfer‐Somi S
Published: December 2012

Contents

Currently, no early pregnancy marker has been identified in the bitch. However, significantly decreased concentrations of heat‐shock protein (HSP) 70 and increased activity of matrix metalloproteinases (MMP) 2,9 were detected in serum from bitches during the pre‐implantation period between days 5 and 13 after mating, that is, 2–3 days after ovulation as determined by the measurement of progesterone and vaginal cytology. Especially during the implantation period and thereafter (days 15–55), high serum concentrations of antibodies against desmin are present, which is believed to indicate or regulate decidualization. Pre‐implantation embryos express mRNA for enzymes and cytokines, known to promote and regulate trophoblast growth, and some intrauterine changings like the increased activity of MMP 2,9 in maternal endometrium are dependant on the presence of embryos. Some mechanisms that protect canine embryos from attack by the maternal immune system can also be identified. The embryos express CD4, a receptor known to interact with immune cells. They, furthermore, do not express MHC I and II, which might prevent them from being recognized as foreign antigen. Pre‐implantation embryos express FasL, which probably renders them able to destroy Fas‐bearing cytotoxic T cells. Furthermore, the uterus during pre‐implantation and implantation expresses cytokines that modulate the intrauterine milieu towards a predominance of Th2 cells. During pre‐implantation and implantation, an increased uterine expression of platelet activating factor (PAF) and PAFR, vascular endothelial growth factor (VEGF) and EGFR2 as well as epithelial growth factor (EGF) is characteristic. Towards placentation, the upregulation of leukaemia inhibiting factor (LIF) and at placentation the expression of insulin‐like growth factor(IGF)2 and granulocyte–macrophage colony‐stimulating factor (GM‐CSF) are striking. Progesterone receptor (PR) appears to be downregulated inside the uterus except at placentation sites, presumably where it is essential for maintenance of pregnancy. In addition, receptor‐bound P4 regulates the activity of MMP 2,9. Apoptosis seems to be a further regulatory mechanism. Expression of Fas and FasL mRNA in uterine tissue is maximum until implantation, both factors then decreased significantly. These changings might indicate increased endometrial apoptosis and defence against maternal cytotoxic T cells, probably promoting trophoblast invasion. In human decidual stromal cells, GnRH is involved in the regulation of apoptosis, which is proposed to be similar in pregnant bitches, as GnRH‐R is expressed at canine implantation sites. Our work investigating immunological changes in pregnant bitches has elucidated aspects of the complex physiology of implantation but raises important questions about the mechanisms involved.
Is Apoptosis a Regulatory Mechanism During Early Canine Pregnancy?
Authors: Schäfer‐Somi S , Sabitzer S , Klein D , Tomaszewski C , Kanca H , Beceriklisoy HB , Kucukaslan I , Agaoglu R , Kaya D , Ay S , Aslan S
Published: December 2012

Contents

Fas is a membrane‐bound protein which upon activation causes programmed cell death. Fas ligand (FasL) binds Fas on target cells. Both these factors are known to regulate apoptosis at implantation in different species and thus might be involved in the regulation of implantation in dogs. The aim of the study was to assess the expression of Fas and FasL in canine uterine tissue throughout pregnancy as well as in pre‐implantation embryos using RT‐PCR and RT‐qPCR. Uterine tissues was collected from of 21 healthy pregnant bitches (group I: days 10–12, n = 5; group II: days 18–25, n = 6; group III: days 28–45, n = 6) and from 4 non‐pregnant bitches (controls: days 10–12). Pregnancy stage was determined by days after mating, that is, 2–3 days after ovulation as determined by vaginal cytology and progesterone measurement. After ovariohysterectomy, uteri from group I bitches were flushed with PBS and the embryos washed and stored frozen at −80°. Tissues from the other groups were taken from the implantation and placentation sites, respectively, covered with Tissue Tek® and frozen at −80°. Extraction of RNA was performed with Trizol Reagent and RT‐qPCR using SYBR green probes. In pre‐implantation embryos, only FasL but not Fas could be detected. In all tissues from pregnant and non‐pregnant bitches, both parameters were detectable. Before implantation (group I) expression of FasL resembled that of non‐pregnant bitches in early dioestrus and decreased significantly during implantation and thereafter (p < 0.05). Expression of Fas did not change significantly until day 45. The relative expression of Fas exceeded that of FasL at each stage investigated, which is comparable to observations of other species; however, high standard deviations indicate high individual differences. These preliminary results point towards a regulatory function of the Fas/FasL system during early canine pregnancy.
Diseases of the Canine Uterus
Authors: Schlafer DH
Published: December 2012

Contents

Lesions most commonly found in the canine and feline uteri fall primarily into two categories: (i) abnormalities of endometrial growth and repair and (ii) uterine infections with associated endometritis. Neoplastic conditions of the tubular genitalia of the bitch, with the possible exception of leiomyomas (tumours that arise from smooth muscle cells), are uncommon. Congenital lesions involving the uterus are relatively rare. The primary object of this paper is to provide a summary of the most common uterine lesions found in the canine and feline uterus using a series of images that underscore the most prominent and important gross diagnostic features of each.
Feline Gonads Exhibit Tissue Specific Alternative Splicing of Oestrogen Receptor Alpha (ESR1)
Authors: Schoen J , Sharbati S , Ritter J , Jewgenow K
Published: December 2012

Contents

Male felids frequently show teratospermia. At least in the domestic cat model, teratospermia is accompanied by impaired regulation of testicular apoptosis. We hypothesize that this phenomenon is caused by dysregulations in oestrogen signalling pathways. Both classical oestrogen receptors (ESR1 and 2) are expressed in species and/or tissue‐specific manners and display different variants, inter alia, caused by alternative splicing. In vitro studies showed that exon deleted transcripts are translated into proteins and that some of the variants modify the effects of the full‐length ERs. It has been proposed that some of the functional and morphological dysregulations, for example, during spermatogenesis, could directly derive from this phenomenon. In the present basic study, we investigated the expression pattern of ESR1 splicing variants in the gonads of domestic cats. Testicular, epididymal as well as ovarian tissue samples were collected from routine castrations. ESR1 variants were detected by means of RT‐PCR using primers spanning one to three exons. We detected the variants Δ4 and Δ7 in all tissue samples investigated. Additionally, the testicular parenchyma expressed the variant Δ6 and double exon deletions of ESR1 (Δ4/6 and Δ6/7). Using an antiserum recognizing all previously identified ESR1 splicing variants, we revealed ESR1 proteins being expressed in nearly all cells of the testicular and ovarian parenchyma. ESR1 Δ6 protein, however, detected by an antiserum specifically raised against the Δ6 variant, was predominantly located in Sertoli cells. As the exon deletion variants are significantly expressed and show a distinct expression pattern, they could specifically modulate the cellular responsiveness to hormonal stimuli within the gonads.
Delay of Puberty and Reproductive Performance in Male Dogs Following the Implantation of 4.7 and 9.4 mg GnRH‐Agonist Deslorelin at an Early Pre‐pubertal Age
Authors: Sirivaidyapong S , Mehl NS , Trigg TE
Published: December 2012

Contents

Stray dogs are a significant problem in large cities. Contraception is an important and useful solution to control the growing population of these dogs. Early‐age neutering is an effective technique for canine population control; however, surgical neutering may not be possible in various situations. GnRH‐agonist implantation has been successful for long‐term reversible contraception in dogs. The efficacy of GnRH‐agonist implantation on long‐term suppression of reproductive performance was observed in male dogs. Eleven 4‐month‐old dogs were implanted with 4.7, 9.4 mg deslorelin or placebo. Sexual behaviour and testicular size were monitored every 2 months. Ejaculates were collected and evaluated at 8, 12, 15, 18, 24, 30, 32, 34 and 36 months of age. Dogs implanted with placebo were found to be healthy and in normal reproductive status. Most dogs (3/4) implanted with 4.7 mg deslorelin showed male sexual behaviour at age of 34 months old. From this group, two dogs had normal semen quality, while semen could not be collected from the other dog, and after castration, no sperm were obtained following epididymal flushing. One dog implanted 4.7 mg deslorelin and four dogs implanted with 9.4 mg deslorelin remained in the non‐pubertal reproductive status at 30–34 months. The delay to puberty was longer in dogs implanted with higher dose of GnRH agonist. Implantation of pre‐pubertal dogs with high doses of GnRH agonist will delay the onset of puberty and may be an effective strategy to reduce the number of unwanted breedings.
The Domestic Dog and Cat as Models for Understanding the Regulation of Ovarian Follicle Development In Vitro
Authors: Songsasen N , Comizzoli P , Nagashima J , Fujihara M , Wildt DE
Published: December 2012

Contents

The culture of ovarian follicles is an important tool for understanding the mechanisms controlling follicle development and differentiation of the oocyte. The benefit of recovering meiotically and developmentally competent oocytes from early stage follicles (primordial, primary, pre‐antral and early antral) also would be significant, ranging from rescue of genomes from endangered species to preserving fertility in women facing cancer treatments. This research field is at an early stage of scientific discovery. To‐date, live offspring from cultured primordial follicles that produced fertilizable oocytes has occurred only in the mouse. Progress in other more complex species has been limited because larger animals have longer durations of natural folliculogenesis, thereby requiring more culture time to generate fully grown follicles and oocytes. We believe the dog and cat are excellent models for understanding more about folliculogenesis in vitro. This review highlights what is known about this topic for these two species as well as future priorities. We have discovered that it is more challenging to maintain viability of primordial follicles within ovarian tissues in vitro in the dog than the cat. Nonetheless, it is possible to grow both isolated cat and dog pre‐antral follicles in culture. Although the follicles of both species have the capacity to increase in size and produce steroids, only cat oocytes appear morphologically normal. The reason for this striking difference between these two species is an area of high research priority. While much more fundamental data are required, we envision advanced technology that will allow harvesting oocytes from the vast, unused follicle stores sequestered within carnivore ovaries. These gametes have utility for reproducing genetically valuable dogs and cats that are ‘companions’ or biomedical models for investigating human disorders as well as for salvaging the genomes of rare canid and felid species that die before contributing to genetic management programs.
Cryopreservation of Canine Embryos and Resulting Pregnancies
Authors: Suzuki H
Published: December 2012

Contents

The assisted reproductive techniques used in dogs have strictly limited utility when compared with other mammals. Although successful somatic cell cloning has been reported, artificial insemination by frozen semen has been only readily available method for improved breeding for companion and working dogs. Recently, successful cryopreservation of embryos and subsequent embryo transfer with a non‐surgical technique in dog was reported. Application of embryo cryopreservation and transfer technology could contribute to breeding management in companion dogs, working dogs including guide dogs and drug‐detecting dogs and quarantine dogs. Such technology would also facilitate the transportation and storage of genetic materials and aid in the elimination of vertically transmitting diseases in the dog.
Laparoscopic Oviductal Embryo Transfer and Artificial Insemination in Felids – Challenges, Strategies and Successes
Authors: Swanson WF
Published: December 2012

Contents

Embryo transfer (ET) and artificial insemination (AI) are potentially invaluable techniques for the propagation and management of genetically valuable domestic cat and endangered nondomestic cat populations. Many of the challenges that impair the effective application of ET and AI in felids may be overcome by using laparoscopic oviductal (LO) approaches. LO‐ET and LO‐AI are minimally‐invasive procedures, requiring only two small skin incisions for insertion of a laparoscope and grasping forceps into the abdominal cavity to permit visualization and catheterization of the oviduct for embryo or semen deposition. With concurrent improvements in embryo culture systems and ovarian synchronization protocols, LO‐ET has proven effective over the past decade for propagation of laboratory cats, cat models of hereditary disease and nondomestic cats. To date, viable offspring have been produced following LO‐ET of non‐frozen and frozen‐thawed IVF‐derived embryos in eight cat hereditary disease models and two nondomestic cat species, the ocelot and sand cat. LO‐AI with low sperm numbers (c. 2–8 million motile) has shown similar efficacy to LO‐ET, resulting in high pregnancy percentages (50–70%) following insemination of gonadotropin‐treated domestic cats. Multiple kittens also have been produced in two hereditary disease models following LO‐AI with frozen semen, and both ocelot and Pallas' cat kittens have been born after LO‐AI with freshly‐collected semen. The application of LO‐ET and LO‐AI to felids has resulted in substantial improvement in the efficiency of assisted reproduction for genetic management of these invaluable domestic cat and wild cat populations.
Immunolocalization of Progesterone Receptors in the Canine Oviduct around Ovulation
Authors: Tahir MZ , Reynaud K , Mawa G , Thoumire S , Chastant‐Maillard S , Saint‐Dizier M
Published: December 2012

Contents

In the bitch, oocyte maturation, sperm storage, fertilization and early embryo development take place within the oviducts under high and increasing circulating progesterone concentrations. To investigate the potential effects of progesterone on the canine oviduct, nuclear progesterone receptors (PR) were localized. Oviducts were collected by ovariectomy from adult Beagle bitches during anestrus, after the onset of proestrus but prior to the Luteinizing Hormone (LH) peak (Pre‐LH), after the LH peak but before ovulation (Pre‐ov) and on Days 1, 4 and 7 after ovulation (n = 3 bitches per stage). The cellular distribution of PR was studied by immunohistochemistry (IHC) in the ampulla, isthmus and tubal part of the utero‐tubal junction (UTJ). Plasma progesterone and 17β‐oestradiol were assayed on the day of surgery. PR were specifically expressed in the nuclei of epithelial, stromal and muscular cells in the ampulla, isthmus and UTJ. The IHC scores did not vary from one oviductal region to another. However, the epithelium displayed higher scores than the stroma at anestrus, Pre‐ov, Days 4 and 7, and also higher scores than muscle at Days 4 and 7 (p < 0.05). Immunohistochemistry scores in the stroma and muscle decreased at Days 4 and 7 compared with previous stages (p < 0.05). Furthermore, muscular IHC scores were positively correlated with circulating 17β‐oestradiol concentrations and negatively correlated with circulating progesterone concentrations (p < 0.05). In conclusion, PR were identified in the canine oviduct, with differences in expression between tissues and times around ovulation, suggesting that progesterone may regulate tubal functions and reproductive events in this species.
Seasonal Aspects of Reproductive Physiology in Captive Male Maned Wolves (Chrysocyon brachyurus, Illiger 1815)
Authors: Teodoro LO , Melo‐Junior AA , Spercoski KM , Morais RN , Souza FF
Published: December 2012

Contents

The aim of the study was to evaluate the seasonality of andrological characteristics and hormonal profile of captive maned wolves (Chrysocyon brachyurus, Illiger 1811). Three adult males were evaluated from the Companhia Brasileira de Metalurgia e Mineração Scientific Breeding Center in Araxá, MG, Brazil, over 13 months. Semen was collected 2–3 times weekly and analysed. Scrotal circumference, biometrics and testicular volume were measured. Stool samples were collected 2–3 times weekly to analyse corticosteroid and testosterone metabolite concentrations. A success rate of 100% was achieved in the collection attempts during the breeding season (BS) and 77.8% during the non‐breeding season (NBS). The interval to achieve penile erection was 1–5 min in the BS and 6–10 in the NBS (p < 0.001). Of the ejaculates collected, 80.0% contained sperm during BS, while 28.6% did during the NBS. The ejaculate had only one fraction, was odourless, predominantly translucent (72.4%), with a watery appearance, pH 6.7 and osmolarity of 352.8 mOsmol. Seasonal influences were seen in ejaculate volume (1.3 ml vs 0.4 ml), number of spermatozoa per ejaculate (73.9 × 106 vs 6.1 × 106) and percentage of live sperm (82.0% vs 66.1%) between the BS and NBS (p < 0.05), respectively. A high percentage of major sperm defects were observed in both seasons (50.1% in BS; 65.7% in NBS). Testicular volume was larger (p < 0.05; right testicles 13.1 cm3 in BS vs 4.0 cm3 in NBS, while left testicles 12.9 cm3 in BS vs 5.3 cm3 in NBS) and testicular consistency increased in the BS. No difference was seen in the basal faecal metabolite concentrations of testosterone; however, the corticosteroid concentrations were higher in the BS. Based on these results, it is possible to conclude that the collection of semen is feasible in captive maned wolves without compromising libido, seminal characteristics and reproductive behaviour and that sperm production is influenced by seasonality; however, it appears that there is no seasonal influence on basal testosterone concentrations.
Birth of Kittens After the Transfer of Frozen–Thawed Embryos Produced by Intracytoplasmic Sperm Injection with Spermatozoa Collected from Cryopreserved Testicular Tissue
Authors: Tharasanit T , Buarpung S , Manee‐In S , Thongkittidilok C , Tiptanavattana N , Comizzoli P , Techakumphu M
Published: December 2012

Contents

The aim of this study is to produce live kittens from oocytes fertilized by intracytoplasmic sperm injection (ICSI) with frozen/thawed testicular spermatozoa. Spermatozoa were collected from thawed testicular tissue and subsequently injected into in vitro matured cat oocytes. At 24 h post‐ICSI, presumptive zygotes/cleaved embryos were treated with 10 μm forskolin for 24 h to reduce intracellular lipid content of embryos (delipidation). At 48 h after oocyte injection, cleaved embryos (2‐ to 8‐cell stage) were frozen in 10% (v/v) ethylene glycol‐based medium by a slow controlled rate method and stored in liquid nitrogen. To evaluate in vitro and in vivo developmental competence, frozen embryos were thawed and then cultured for 6 days (n = 155) or cultured for 2 h before transferred (n = 209) to hormonal (equine chorionic gonadotropin/hCG)‐treated cat recipients. Cleavage frequency at day 2 after ICSI with frozen/thawed testicular spermatozoa was ~30%. The percentages of frozen/thawed embryos that developed to morula and blastocyst stage (on day 3 and day 6 of in vitro culture, respectively) were significantly lower than that of fresh ICSI embryos (22.6 vs 45.2% and 21.3 vs 38.7%, respectively; p < 0.05). However, no difference was found in the number of blastomeres between frozen/thawed (242.5 ± 43.1) and fresh (320.2 ± 28.1) blastocysts. Three of seven cat recipients were pregnant and one pregnant cat delivered two healthy kittens. This is the first report of the birth of kittens after the transfer of frozen–thawed embryos produced by ICSI with frozen/thawed testicular sperm.
Effect of Storage Media and Storage Time on Histological and Ultrastructural Changes in Cat Epididymal Cells
Authors: Tittarelli CM , Jurado SB , Nuñez‐Favre R , Bonaura MC , de la Sota RL , Stornelli MA
Published: December 2012

Contents

The aim of this study was to assess the histological and ultrastructural changes in cat epididymides (n = 22) stored at 4°C in two different media [saline solution (SAL) or tris‐egg yolk (TEY)]. Our hypothesis was that epididymides stored in TEY would have delayed epithelial cell autolysis. Four epididymides were fixed and processed immediately, and the remaining 18 epididymides were stored at 4°C in SAL or TEY for 24, 48 or 72 h. In histological sections, the nuclear features and stereocilia morphology were scored from 0 to 3. Ultrastructurally, nuclear chromatin and stereocilia morphology were scored from 0 to 3. In addition, using transmission electron microscopy nuclear number, nuclear area, mitochondrial number and mitochondrial area were recorded. In the histological study, parameters changed with time and media (p < 0.01). A significant effect of time was observed (p < 0.01), and the morphological changes were greatest when the storage time increased. Morphological changes were higher in SAL compared with TEY (p < 0.01). In the ultrastructural study, nuclear chromatin and stereocilia morphology decreased with time and media as in the histological study (p < 0.01). In addition, nuclear number and nuclear area changed with time (p < 0.004; p < 0.001) but not with media. Conversely, mitochondrial number and mitochondrial area did not change with media or time (p > 0.05). In conclusion, these results show that TEY preserved epididymal epithelial cells better than SAL; this finding could help improve sperm quality of stored epididymides.
Spermatogenic Function in Cats
Authors: Tsutsui T , Oba H , Fujimoto S , Toyonaga M
Published: December 2012

Contents

There are limited data on feline sperm production. We exhausted epididymal spermatozoa (i.e. the number of ejaculated spermatozoa <5 × 106) by frequent semen collections using the artificial vagina method in five tomcats and determined the number of spermatozoa stored in the epididymis. We investigated the time (days) required for the number of epididymal spermatozoa to return to the pre‐exhaustion level and determined the number of spermatozoa produced per day. After spermatozoa were exhausted by frequent semen collection, 6 or more days were required to return to the pre‐exhaustion level. Based on the duration of resting (days) and total number of spermatozoa, the mean number of spermatozoa produced per day was 30 × 106.
Ovulation Compensatory Function After Unilateral Ovariectomy in Dogs
Authors: Tsutsui T , Hori T , Takahashi F , Concannon PW
Published: December 2012

Contents

As a step towards elucidation of the timing and mechanism of the determination of the number of ovulated ova in dogs, we excised one ovary 2, 5 and 8 days after the beginning of vulval bleeding and examined whether the lost ovulation function, assessed by estimating the number of ovulated oocytes, would be compensated for by the remaining ovary. The number of ovulated ova was maintained by the remaining ovary in the group that underwent unilateral ovariectomy 2 days after the beginning of vulval bleeding. However, in the groups ovariectomized 5 or 8 days after the beginning of vulval bleeding, no compensation for the number of ova that would have been ovulated from the lost ovary was observed; ova were ovulated only from the follicles 3 mm or greater in diameter observed in the remaining ovary at unilateral ovariectomy. Thus, in dogs, the number of ovulated ova is considered to be determined within 5 days after the beginning of vulval bleeding.
Ovulation Day After Onset of Vulval Bleeding in a Beagle Colony
Authors: Hori T , Tsutsui T , Amano Y , Concannon PW
Published: December 2012

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This study investigated the duration of the interval between the onset of vulval bleeding at pro‐oestrus and ovulation estimated from the plasma progesterone concentration in a large number of beagle bitches. The influence and association of individual variation, ageing and duration of the oestrous cycle were also investigated. The mean time of ovulation after the onset of vulval bleeding was 11.1 ± 0.2 days, but it widely ranged from 3 to 31 days. This timing was not influenced by age or duration of the oestrous cycle, and within‐individual variation was small. As there has been no previous report in which the ovulation day was investigated by the age, these data may be very valuable.
Intravaginal Artificial Insemination in Bitches Using Frozen/Thawed Semen after Dilution in Powdered Coconut Water (ACP‐106c)
Authors: Uchoa DC , Silva TFP , Mota Filho AC , Silva LDM
Published: December 2012

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The aim of this study was to evaluate powdered coconut water extender (ACP‐106c; ACP Serviços Tecnológicos Ltda, ACP Biotecnologia, Fortaleza, Ceará, Brazil) as a diluent for freezing dog semen and the fertility after vaginal insemination of semen frozen therein. Ten ejaculates were collected from five dogs, evaluated fresh, diluted in ACP‐106c, 10% egg yolk and 6% glycerol, cooled and frozen. In the first phase of the study, straws with frozen semen were thawed and immediately subjected to the same analysis as the fresh semen and, in addition, to Computer‐Assisted Semen Analysis (CASA). In phase 2, 10 bitches that had been subjected to natural breeding during a preceding oestrous cycle were vaginally inseminated with thawed semen that had been re‐diluted in ACP‐106c. After thawing, a mean of 77% sperm motility was obtained through subjective analysis and 77.3% through CASA. Following artificial insemination, a 60% pregnancy rate was observed, resulting in a 50% parturition rate and a mean litter size of 3.4 (SEM 0.6), with 47.1% males and 52.9% females. ACP‐106c can be successfully used for freezing canine semen, and vaginal deposition of such semen yields similar pregnancy rates to those reported in other studies.
Prenatal and Neonatal Adaptations with a Focus on the Respiratory System
Authors: Vannucchi CI , Silva LCG , Lúcio CF , Regazzi FM , Veiga GAL , Angrimani DS
Published: December 2012

Contents

Among the modifications that occur during the neonatal period, pulmonary development is the most critical. The neonate's lungs must be able to perform adequate gas exchange, which was previously accomplished by the placenta. Neonatal respiratory distress syndrome is defined as insufficient surfactant production or pulmonary structural immaturity and is specifically relevant to preterm newborns. Prenatal maternal betamethasone treatment of bitches at 55 days of gestation leads to structural changes in the neonatal lung parenchyma and consequently an improvement in the preterm neonatal respiratory condition, but not to an increase in pulmonary surfactant production. Parturition represents an important challenge to neonatal adaptation, as the uterine and abdominal contractions during labour provoke intermittent hypoxia. Immediately after birth, puppies present venous mixed acidosis (low blood pH and high dioxide carbon saturation) and low but satisfactory Apgar scores. Thus, the combination of physiological hypoxia during birth and the initial effort of filling the pulmonary alveoli with oxygen results in anaerobiosis. As a neonatal adaptation follow‐up, the Apgar analysis indicates a tachypnoea response after 1 h of life, which leads to a shift in the blood acid–base status to metabolic acidosis. One hour is sufficient for canine neonates to achieve an ideal Apgar score; however, a haemogasometric imbalance persists. Dystocia promotes a long‐lasting bradycardia effect, slows down Apgar score progression and aggravates metabolic acidosis and stress. The latest data reinforce the need to accurately intervene during canine parturition and offer adequate medical treatment to puppies that underwent a pathological labour.
Molecular Markers of Spermatogonial Stem Cells in the Domestic Cat
Authors: Vansandt LM , Pukazhenthi BS , Keefer CL
Published: December 2012

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Spermatogonial stem cells (SSCs) represent an exciting new avenue for assisted reproduction in endangered and genetically valuable species. Before this technology can be applied to wildlife, species‐specific markers are required to evaluate SSC enrichment strategies and monitor subsequent in vitro culture. This study was designed to evaluate six conserved SSC markers (THY1, GPR125, GFRalpha1, PLZF, UCHL1 and OCT4) in the cat. Testes from three juveniles and three adults were obtained following routine castrations and processed for mRNA extraction. RT‐PCR of whole testis and cell suspensions enriched for SSCs by differential plating confirmed that all six SSC markers are expressed in both the whole testis and SSC‐enriched cell fractions. The expression of all six putative SSC marker genes in the cat testis suggests conservation of SSC markers, and perhaps self‐renewal mechanisms, in felids.
Clinical Evaluation of Deslorelin to Induce Oestrus, Ovulation and Pregnancy in the Bitch
Authors: von Heimendahl A , Miller C
Published: December 2012

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The aim of this study was to evaluate the practical use of Deslorelin in a clinical setting to induce oestrus and ovulation and to assess pregnancy rates. Sixteen anoestrus bitches that were brought into the clinic were implanted subcutaneously in the ventral abdomen with Deslorelin. Implants were removed after ovulation was assumed to have occurred. All bitches came into oestrus and 13 ovulated. 11 of 16 bitches became pregnant (68.8%) and all continued to term. Average litter size was 7.6 puppies. On average, the implant was in place for 13.8 days.
Preservation of Primordial Follicles from Lions by Slow Freezing and Xenotransplantation of Ovarian Cortex into an Immunodeficient Mouse
Authors: Wiedemann C , Hribal R , Ringleb J , Bertelsen MF , Rasmusen K , Andersen CY , Kristensen SG , Jewgenow K
Published: December 2012

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Assisted reproductive technology (ART) is considered an important tool in the conservation of endangered species, but often the most limiting factor of ART is the availability of mature oocytes. The aim of the present study was to investigate the feasibility of preserving female germ cells from ovaries of female lions (Panthera leo). Good quality cumulus–oocyte complexes (COCs) were isolated and subjected to in vitro maturation (IVM). In addition, ovarian cortex was obtained and cut into pieces for culture and cryopreservation by slow freezing. The survival of ovarian follicles was assessed by histology. Frozen–thawed samples of ovarian cortex samples were xenotransplanted under the skin of ovariectomized immunodeficient mouse for 28 days. Overall, 178 intact COCs were obtained from 13 lions, but only 28.1% were matured in vitro indicating insufficient IVM conditions. In contrast, almost all follicles within the ovarian cortex survived culture when the original sample was from a young healthy lion collected immediately after euthanasia. Within the xenotransplants, the number of primordial follicles decreased after 28 days by 20%, but the relation between primordial and growing follicles changed in favour of follicular growth. Female gamete rescue from valuable felids may be performed by slow freeze cryopreservation of ovarian cortex. Although the IVM protocol for lions is not yet optimized, mature oocytes may be obtained after long‐term xenotransplantation and IVM and could potentially represent one way of salvage of endangered felid species in the future.
Litter Size Response to Oestrous Induction with Deslorelin (Ovuplant®) in Dogs
Authors: Wolf T , Meyer H , Kutzler M
Published: December 2012

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Four German Shorthair Pointer bitches each produced from two to five (total of 14) purebred litters in response to natural matings to either natural oestrus (n = 8) or oestrus induced by Ovuplant®, a sustained‐release implant containing 2.1 mg deslorelin (n = 6). All bitches initially produced litters from natural oestrous matings prior to producing litters from induced oestrus, and two of the bitches also produced natural litters subsequent to their Ovuplant® litters. Mean litter size was lower for Ovuplant® litters (5.4 vs 8.6 puppies; p < 0.001) and within each bitch every induced litter was smaller than any of her natural litters.
Testicular Steroids, Prolactin, Relaxin and Prostate Gland Markers in Peripheral Blood and Seminal Plasma of Normal Dogs and Dogs with Prostatic Hyperplasia
Authors: Wolf K , Kayacelebi H , Urhausen C , Piechotta M , Mischke R , Kramer S , Einspanier A , Oei CHY , Günzel‐Apel A
Published: December 2012

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Concentrations of 17β‐oestradiol (E2), testosterone (T), 5α‐dihydrotestosterone, prolactin (PRL) and relaxin (RLN) were determined in peripheral blood serum or plasma and prostatic secretion of 77 physically healthy intact male dogs (19 Rhodesian Ridgebacks/RR, 58 dogs of other breeds, 1–9 years of age). Furthermore, the concentrations of acid phosphatase in prostatic secretion and canine prostate‐specific esterase (CPSE) were measured in blood plasma. All dogs were submitted to a complete breeding soundness examination, including B‐mode sonography. Prostatic volume was larger, and blood plasma levels of CPSE were higher in ageing dogs and in dogs with benign prostatic hyperplasia (BPH) compared with young dogs and dogs with normal prostate. Furthermore, a higher E2/T ratio was found in dogs with BPH. Despite missing significant differences in PRL concentrations, the slight increases in PRL concentrations in the prostatic secretion observed both with increasing age and in dogs with BPH and the observed correlations between concentrations of PRL and testicular steroids may possibly indicate a role of PRL in the pathogenesis of canine BPH. Serum RLN concentrations were at similar level in all dogs. Regarding breed differences, an appreciably larger prostatic volume and higher concentration of CPSE were verified in RR than in other pure‐bred dogs, confirming our suspicion of a premature enlargement of the prostate gland, which may result from a genetic disposition for BPH in this breed.
Validation of a Model to Develop a Symptom Index for Benign Prostatic Hyperplasia in Dogs
Authors: Zambelli D , Cunto M , Gentilini F
Published: December 2012

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Benign prostatic hyperplasia (BPH) is a spontaneous and age‐related condition in humans and intact male dogs. A symptom index forBPHin men was created by theAmericanUrologicalAssociation. In this study, it has been developed and statistically validated as a model to assign an objective score to canineBPHseverity based on clinical signs observed and/or subjectively reported to the veterinarian by dog owners. The medical records of theAnimalReproductionUnit ofUniversity ofBologna (Italy) were used to select dogs with a clinical diagnosis ofBPH. A data set was built up, and the animals were included in the statistical analysis as dependent variables. A score of 1–3 was assigned to the disease severity of each case based on signs annotated, graded using a scale ranging from 1 to 4. Signs ofBHPwere entered as predictors while disease severity as dependent variable to generate the predictive model. The model was finally used to re‐classify each case of the data set, and the percentage of corrected predictions calculated. Overall, 373 subjects were entered in the model. Between them, 243, 107 and 23 animals have been represented based on medical records with aBPHseverity score of 1, 2 and 3, respectively. The model correctly predicted the response variable in 97.3% of the cases. In this study, aBPHsymptom index was created for the first time in dogs, which may be useful to standardizeBPHseverity with an objective score and to evaluate the necessity, the kind and the effectiveness of treatment.
Monophasic Teratoma of the Ovarian Remnant in a Bitch
Authors: Rota A , Tursi M , Zabarino S , Appino S
Published: December 2012

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An exploratory laparotomy on a mixed‐breed bitch of an estimated age of 5 years revealed that she had undergone ovariectomy in the past, but a cystic structure was present in the area of the right ovary and a whitish mass, approximately 3 cm in diameter, in the area of the left ovary. These structures were removed together with an apparently normal uterus. Histological examination of the cyst showed a thin layer of connective tissue, while the left ovarian mass revealed ovarian tissue and highly differentiated nervous tissue, confirmed through immunohistochemistry. A presumptive diagnosis of mature ovarian teratoma was made. Although teratomas generally contain recognizable elements from more than one of the three germ cell layers, they can also be monophasic, when there is only one germ layer component. Ovarian teratomas are rare in the dog and never before have been reported in an ovarian fragment.
Canine Herpesvirus During Pregnancy and Non‐Pregnant Luteal Phase
Authors: Ström Holst B , Hagberg Gustavsson M , Grapperon‐Mathis M , Lilliehöök I , Johannisson A , Isaksson M , Lindhe A , Axnér E
Published: December 2012

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Canine herpesvirus (CHV) is a widespread infection among dogs that typically get latently infected after exposure and can reactivate the infection after stress. The aim of the present study was to study the effects of latentCHVinfection during pregnancy on pregnancy outcome, and to study if there are signs of genital viral reactivation during pregnancy or during non‐pregnant luteal phase. Twelve mated bitches and eight control bitches were followed and sampled regularly during pregnancy or non‐pregnant luteal phase. Blood samples were taken for antibody analysis and vaginal swabs for real‐timePCRanalysis. Three of the pregnant bitches were vaccinated againstCHVduring pregnancy. All bitches had antibodies toCHV. Two pregnant bitches that were not vaccinated had a twofold or larger increase inCHVtitre, with no negative effects detected on pregnancy. Higher titres were not associated with smaller litters or with vaccination. There was no consistent variation in antibody titres due to pregnancy or non‐pregnant luteal phase. Vaginal excretion ofCHVwas not detected from any of the bitches.
Application of Vaginal Temperature Measurement in Bitches
Authors: Maeder B , Arlt S , Burfeind O , Heuwieser W
Published: December 2012

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Finding innovative, non‐invasive methods for continuously measuring body temperature minimizing human interference is important for accurate data collection. The objective of this study was to assess feasibility and accuracy of continuous body temperature measurements with loggers placed in the vaginal cavity of bitches. First, an in vitro experiment was performed to compare values obtained by temperature loggers (n = 26) to a calibrated liquid‐in‐glass thermometer. The mean differences between the two methods were low. Next, an in vivo experiment was performed using five healthy bitches, and values obtained by the vaginal loggers were compared to measurements collected rectally with digital thermometers. The results show that rectal and vaginal temperatures were correlated. The mean differences between rectal and vaginal temperatures were negligible. We conclude that the utilized temperature loggers provide accurate and reliable data.
Effect of Simulated Stages of the Canine Oestrous Cycle on Escherichia coli Binding to Canine Endometrium
Authors: Krekeler N , Lodge KM , Anderson GA , Browning GF , Charles JA , Wright PJ
Published: December 2012

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Pyometra, a prevalent infectious uterine disease that affects intact middle‐aged bitches, is typically associated with Escherichia coli. Our hypotheses were (i) that bacterial adhesion to canine endometrium differs between different stages of the oestrous cycle and (ii) that the adhesin FimH facilitates this adhesion. Twelve post‐pubertal, ovariectomized greyhound bitches were treated with exogenous hormones to simulate different stages of the oestrous cycle. Tissue samples from each uterus were incubated with a pathogenic E. coli strain carrying the fimH gene, but no other adhesin genes (P4‐wt)–or an E. coli strain in which fimH was insertionally inactivated (P4‐∆fimH::kan)–or with phosphate‐buffered saline as a negative control. After washing, tissue samples were homogenized for quantification of adherent bacteria. The differences in binding to canine endometrium at different stages of the oestrous cycle were not significant. However, the mean difference in binding of the P4‐wt and the P4‐∆fimH::kan across all stages of the simulated oestrous cycle was significant (p < 0.001 by paired t‐test on geometric means). Individual differences in numbers of P4‐wt bacteria bound between dogs might suggest genetic variations or epigenetic differences in FimH receptor expression by the endometrium, unrelated to the stage of the oestrous cycle.
Food Intake and Nutrition During Pregnancy, Lactation and Weaning in the Dam and Offspring
Authors: Fontaine Emmanuel
Published: December 2012

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In mammalian species, the connection between reproduction and nutrition is undeniable and illustrated by numerous publications: its role concerning regulation of hormonal secretions, fertility, pregnancy outcome, lactation and neonatal development indeed retains the attention of the scientific community. The interest on the influence of nutrition in dog and cat reproductive physiology is growing. Awareness on the key aspects of feeding during critical periods like pregnancy, lactation and weaning is essential to optimize the reproductive performances in these species.