Objective Cryptosporidium spp. are prevalent globally and sheep are an important zoonotic reservoir. Little data regarding the rates of Cryptosporidium infections in ovines in China are available. This study assessed ...Objective Cryptosporidium spp. are prevalent globally and sheep are an important zoonotic reservoir. Little data regarding the rates of Cryptosporidium infections in ovines in China are available. This study assessed the prevalence of Cryptosporidium spp. in pre-weaned ovines from Aba Tibetan and Qiang Autonomous Prefecture in the Sichuan province of China. Methods A total of 213 fecal samples were collected from pre-weaned ovines and were examined microscopically （following modified acid fast staining）. In addition, 18S rRNA genetic sequences were amplified from fecal samples by nested PCR and phylogenetically analyzed. Results The prevalence of Cryptosporidium in the collected samples was at 14.6% （31/213） and four isolates identified by PCR belonged to the Cryptosporidium cervine genotype （Cryptosporidium ubiquiturn） demonstrating that this species was the primary sheep species found in sheep in China. Conclusion The present study suggested that the high incidence of Cryptosporidium in sheep poses a significant public health threat and that surveillance practices must be established to prevent zoonotic disease of humans.展开更多
Background Selenium(Se)is an essential soil mineral that can be incorporated into animal feedstuffs.Because of a lack of soil Se in some regions,organic or inorganic supplementation strategies must be implemented to p...Background Selenium(Se)is an essential soil mineral that can be incorporated into animal feedstuffs.Because of a lack of soil Se in some regions,organic or inorganic supplementation strategies must be implemented to prevent deficiencies and promote optimal ovine health.Therefore,the objectives of this study were to assess how inorganic versus organic Se supplementation influenced ewe Se status and immune function during late gestation and postpartum.Dorset Rideau ewes(n=110)were fed a Se deficient diet from gestation d 110 through postpartum d 49 and received one of four daily oral Se treatments diluted in 5 mL of sugar water:0 mg Se,0.3 mg inorganic Se,0.3 mg organic Se,and 0.6 mg organic Se.Throughout the trial,the ewes received various immune challenges,including intramuscular immunizations with a novel antigen(ovalbumin;OVA)on trial d 0 and 10,an intradermal OVA challenge on d 20,and a lipopolysaccharide(LPS)endotoxin challenge on trial d 49.Results The organic Se treatment groups had higher serum Se concentrations on most trial days compared to the 0.3 mg inorganic and control groups(P<0.05).No significant treatment differences were found for the dermal hypersensitivity response to OVA,anti-OVA antibody response,glutathione peroxidase activity,cytokine response,cortisol response,or rectal temperature(P>0.05).However,4 h post-LPS injection,the serum albumin concentration was significantly lower in the 0.3 mg inorganic group compared to both organic Se groups,potentially indicating a higher degree of inflammation in the ewes supplemented with the inorganic Se.Conclusions The results of this study indicate that organic Se supplementation can promote a higher Se status in ewes over time,but Se supplementation during this study period did not affect tested immunological parameters.This lack of difference in immune responsiveness between groups may be due to an absence of true serum Se deficiencies in the Se-deficient group or the levels of Se supplementation being insufficient to significantly improve immunocompetence.展开更多
Ovine bones are the major by-products after slaughtered. The present study was conducted to extract and characterize acid soluble collagens （ASC） and pepsin soluble collagens （PSC） from ovine bones （Ujumuqin shee...Ovine bones are the major by-products after slaughtered. The present study was conducted to extract and characterize acid soluble collagens （ASC） and pepsin soluble collagens （PSC） from ovine bones （Ujumuqin sheep）. Ovine bones collagen were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis （SDS-PAGE） and liquid chromatography-tan- dem mass spectrometry （LC-MS/MS） as type I collagen. The results of Fourier transform infrared （FTIR） spectra analysis testified the existence of triple superhelical structure in both ASC and PSC, showing pepsin did not disrupt the triple helical structure of ovine bones collagen. Glycine, accounting for one-third of total amino acids, was the major amino acid for ovine bones collagen. Higher imino acid content was responsible for higher thermal denaturation temperature of ovine bones collagen compared to fish collagens. The isoelectric point of ASC was lower than PSC due to the higher content of acidic amino acids. Therefore, this study provides the potential reference for collagen extraction and application of ovine bones by-procduct.展开更多
Phosphorylation post-translational modification plays an important role in postmortem muscle quality traits. Adenosine triphosphate(ATP) is an energy source and a key substrate of phosphorylation which provides the ph...Phosphorylation post-translational modification plays an important role in postmortem muscle quality traits. Adenosine triphosphate(ATP) is an energy source and a key substrate of phosphorylation which provides the phosphatase groups to proteins in the presence of protein kinases. However, in postmortem muscle, the effects of ATP content on phosphorylation are poorly studied. The study investigated the effect of ATP on protein phosphorylation and degradation in postmortem ovine muscle. The ground muscle with/without additional ATP were treated/control groups and stored at 25 and 4℃, respectively. The ATP content led to different changes of p H value between the ATP-treated and control groups. The phosphorylation level of myofibrillar proteins was higher(P<0.05) in ATP-treated group compared to the control group at both temperatures, which suggested that ATP played a vital role in postmortem protein phosphorylation. A slower degradation rate of μ-calpain, desmin and troponin T was observed in the ATP-treated group which showed that there was a negative relationship between ATP level and the degradation of proteins. These observations clearly highlighted the role of ATP on the development of meat quality by regulating the phosphorylation and degradation of myofibrillar proteins in postmortem ovine muscle.展开更多
Endogenous beta retroviruses (enJSRV) are highly homologous with Jaagsiekte sheep retrovirus (exJSRV),this exogenous retrovirus is the aetiological agent of ovine pulmonary adenocarcinoma (OPA). The aim of this study ...Endogenous beta retroviruses (enJSRV) are highly homologous with Jaagsiekte sheep retrovirus (exJSRV),this exogenous retrovirus is the aetiological agent of ovine pulmonary adenocarcinoma (OPA). The aim of this study was to clarify the function of enJSRV and the immunological mechanisms of its corresponding antibody, that is undetectable in JSRV-infected ovine serum. The expression of enJSRV envelope protein and Hyal-2 mRNA in immune organs and lungs of ovine fetuses and lambs were analyzed by Real-Time reverse transcription PCR and In Situ Hybridization using specific probes. In Situ Hybridization results indicated that the enJSRV envelope protein and Hyal-2 mRNA were expressed in thymus, spleen, mesenteric lymph nodes and lungs at different times, while no positive signals were detected in the negative controls. On the other hand, results from Real-Time reverse transcription PCR analysis showed that in 130d fetuses and 3d newborn lambs the enJSRV mRNA levels were much higher in organs associated with the immune system than that in lungs, especially in the thymus and spleen, but levels of Hyal-2 mRNA expression was not significantly different in all collected tissue. These results provided evidence from an immunology point of view to understand why the circulating antibodies against exJSRV are undetectable in JSRV-infected ovine, and will help to unravel the pathogenesis of JSRV-infected ovine.展开更多
Background:Heavy metal cadmium(Cd)is a widespread environmental contaminant with a potential toxicity that might negatively affect female reproduction and fertility.It has been reported that Cd exposure impaired the q...Background:Heavy metal cadmium(Cd)is a widespread environmental contaminant with a potential toxicity that might negatively affect female reproduction and fertility.It has been reported that Cd exposure impaired the quality of oocytes and led to a defective maturation and fertilization,through oxidative stress induction.Resveratrol(Res)is a natural polyphenol with strong antioxidant properties that exhibited protective role in preventing oocyte redox homeostasis disruption and quality decline.Here,we explored whether the addition of Res to in vitro maturation(IVM)medium might act as a protection against Cd-induced toxicity on ovine oocyte maturation and fertilization.Firstly,we evaluated the effect of supplementing IVM medium with two different Res concentrations(1and 2μmol/L)on nuclear maturation and fertilization of oocytes matured under CdCl2(2μmol/L)exposure.Therefore,the concentration of 1μmol/L Res was selected to analyse the effects of this compound on intracellular ROS levels,mitochondrial(mt)distribution and activity,chromatin configuration,cytoskeleton morphology,cortical granules(CGs)distribution and mRNA expression of genes associated with cellular response to oxidative stress(i.e.SIRT1,SOD 1,GPX1,GSR,CAT)in Cd-exposed in vitro matured oocytes.Results:We found that 1μmol/L Res restored the reduced oocyte meiotic competence induced by Cd exposure as well as,Res sustained oocyte ability to be normally fertilized and decreased polyspermic fertilization at both tested concentrations.Moreover,we demonstrated that 1μmol/L Res mitigated Cd-induced alterations of oocyte cytoplasmic maturation by reducing reactive oxygen species(ROS)accumulation,preventing mt dysfunction,maintaining the correct meiotic spindle and cortical F-actin assembly and the normal cortical granule distribution as well as up-regulating SIRT1,SOD1 and GPX1 genes.Conclusions:Taken together,our findings highlighted the beneficial influence exerted by Res in preventing Cdinduced disturbance of nuclear and cytoplasmic maturation and subsequent fertilization in ovine oocytes.Res treatment may help to establish defence strategies counteracting Cd-induced toxicity on the female gamete.展开更多
ISG20 is an interferon-inducible exonuclease that inhibits virus replication.Although ISG20 is thought to degrade viral RNA,the antiviral mechanism and specificity of ISG20 remain unclear.In this study,the antiviral r...ISG20 is an interferon-inducible exonuclease that inhibits virus replication.Although ISG20 is thought to degrade viral RNA,the antiviral mechanism and specificity of ISG20 remain unclear.In this study,the antiviral role of ovine ISG20(o ISG20)in bluetongue virus(BTV)infection was investigated.It was found that BTV infection upregulated the transcription of ovine ISG20(o ISG20)in a time-and BTV multiplicity of infection(MOI)-dependent manner.Overexpression of o ISG20 suppressed the production of BTV genome,proteins,and virus titer,whereas the knockdown of o ISG20 increased viral replication.o ISG20 was found to co-localize with BTV proteins VP4,VP5,VP6,and NS2,but only directly interacted with VP4.Exonuclease defective o ISG20 significantly decreased the inhibitory effect on BTV replication.In addition,the interaction of mutant o ISG20 and VP4 was weakened,suggesting that binding to VP4 was associated with the inhibition of BTV replication.The present data characterized the anti-BTV effect of o ISG20,and provides a novel clue for further exploring the inhibition mechanism of double-stranded RNA virus by ISG20.展开更多
In order to amplify the complete genome of enJSRV from the strain of Inner Mongolia (enJSRV-NM), we used enJSRV-specific and JSRV-specific DNA probes in dot blot hybridization. Seven pairs of primers were designed bas...In order to amplify the complete genome of enJSRV from the strain of Inner Mongolia (enJSRV-NM), we used enJSRV-specific and JSRV-specific DNA probes in dot blot hybridization. Seven pairs of primers were designed based on Genbank sequences. Seven fragments were obtained by PCR and were cloned into the PMD19-T vectors. The recombinant plasmids were sequenced and analyzed. The results showed that the genome was 7 942 bp in length and contained four overlapping open reading frames corresponding to the gag, pro, pol and env genes as well as an additional open reading frame (orf-x) that overlaps the 3' end of the pol gene. The nucleotide acid sequences of the enJSRV-NM loci were compared with the sequences of South Africa enJS56A1 strain (Accession No. AF153615) and USA JSRV21 strain (Accession No. AF105220). The nucleotide acid identities were 99.2% and 92.3% respectively. Two zinc fingers were found in the NC region in the predicted amino acid sequence. However, the YXXM motif, which is a reliable molecular marker for the infectious exogenous virus, was not found in the TM region. It was found that the enJSRV-NM region was 90%-98% identical at the amino acid level to its exogenous infectious counterparts in most of the retroviral genome. This is the first nucleotide sequence of enJSRV reported in P.R China. The resource work has provided a wide range of information useful not only for expression genomics and annotation of genomic DNA sequence, but also for further research on the clinical diagnosis of OPA.展开更多
Background:To advance the use of embryo vitrification in veterinary practice,we developed a system in which embryo vitrification,warming and dilution can be performed within a straw.Ovine in vitro produced embryos(IVE...Background:To advance the use of embryo vitrification in veterinary practice,we developed a system in which embryo vitrification,warming and dilution can be performed within a straw.Ovine in vitro produced embryos(IVEP)were vitrified at either early(EBs:n=74)or fully expanded blastocyst stage(FEBs:n=195),using a new device named"E.Vit",composed by a 0.25-m L straw with a 50-μm pore polycarbonate grid at one end.Embryos at each stage(EBs and FEBs)were vitrified by either Two-step(TS)or Multi-step(MS;6 different concentrations of vitrification solutions)protocol.Non-vitrified embryos(n=102)were maintained in in vitro culture as a control.Warming consisted of placing the straws directly into 1.5 m L tubes containing a TCM-199 solution with three decreasing concentrations of sucrose.Blastocyst re-expansion,embryo survival and hatching rate were evaluated at2,24 and 48 h post warming.The number of apoptotic cells was determined by TUNEL assay.Results:Blastocyst re-expansion(2 h)after warming was higher(P<0.05)in FEBs group,vitrified with the MS and TS methods(77.90%and 71.25%,respectively)compared with the EBs group(MS:59.38%and TS:48.50%,respectively).Survival rates of vitrified FEBs after 24 h IVC were higher(P<0.001)in both methods(MS and TS)than vitrified EBs(MS:56.25%;TS:42.42%)and was higher(P<0.05)in the MS method(94.19%)compared with those in TS(83.75%).After 48 h of culture the hatching rate for FEBs vitrified in MS system(91.86%)was similar to control(91.89%),but higher than FEB TS(77.5%)and EBs vitrified in MS(37.5%)and TS(33.33%).Number of apoptotic cells were higher in EBs,irrespective of the system used,compared to FEBs.The number of apoptotic cells in FEBs vitrified with MS was comparable to the control.Conclusions:A high survival rate of IVP embryos can be achieved by the new"E.Vit"device with hatching rates in vitro comparable with control fresh embryos.This method has the potential for use in direct embryo transfer in field conditions.展开更多
This study aimed to examine changes in phosphorylation of sarcoplasmic and myofibrillar proteins from longissimus lumborum,semitendinosus,and psoas major muscles during postmortem ageing for 5 d.These sarcoplasmic and...This study aimed to examine changes in phosphorylation of sarcoplasmic and myofibrillar proteins from longissimus lumborum,semitendinosus,and psoas major muscles during postmortem ageing for 5 d.These sarcoplasmic and myofibrillar proteins were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with phosphorous and protein specific stains.Myofibril fragmentation index,pH,the content of lactic acid and the relative activity of μ-calpain in three ovine muscles were measured.These results showed that the relative phosphorylation level of sarcoplasmic and myofibrillar proteins of psoas major muscle were lower compared with longissimus lumborum and semitendinosus muscles(P<0.05).The pH of psoas major muscle was the lowest at 0.5 h postmortem,and the highest after 12 h postmortem(P<0.05).In addition,the relative activity of μ-calpain was higher within 5 d postmortem and myofibril fragmentation index was higher after 1 d postmortem in psoas major muscle than those of longissimus lumborum and semitendinosus muscles(P<0.05).The sarcoplasmic protein phosphorylation may regulate the rate of pH decline to influence the μ-calpain activity and then proteolysis of proteins consequently.This study gives a new perspective of the mechanism of postmortem meat tenderization.展开更多
Background Recent evidence suggests important roles for progesterone(P4)and interferon tau in the regulation of calcium,phosphate,and vitamin D signaling in the uteri of pregnant sheep.However,the effects of P4 and es...Background Recent evidence suggests important roles for progesterone(P4)and interferon tau in the regulation of calcium,phosphate,and vitamin D signaling in the uteri of pregnant sheep.However,the effects of P4 and estradiol(E2),with respect to the expression of their receptors PGR and ESR1,respectively,in uterine epithelia on mineral signaling during the estrous cycle has not been investigated.Estrous cycles of mature Suffolk ewes were synchronized,prostaglandin F2αwas administered,and ewes were observed for estrus(designated as Day 0)in the presence of vasectomized rams.On Days 1,9,or 14 of the estrous cycle,hysterectomies were performed.Results 25-hydroxyvitamin D was more abundant in plasma from ewes on Day 14 than Day 1(P<0.05).Expression of fibroblast growth factor receptor 2(FGFR2),a disintegrin and metalloprotease 17(ADAM17),and parathyroid hormone-related protein(PTHrP)mRNAs was greater in endometria on Day 9 compared to Days 1 and 14(P<0.01).Similarly,expression of transient receptor potential cation channel subfamily V member 6(TRPV6)mRNA was greater in endometria on Day 9 than Day 1(P<0.05).ATPase plasma membrane Ca^(2+)transporting 4(ATP2B4)and S100 calcium binding protein G(S100G)mRNA expression was greater in endometria on Day 14 than on Days 1 and 9(P<0.01).In contrast,endometrial expression of vitamin D receptor(VDR)mRNA was lower on Days 9 and 14 than Day 1(P<0.01).Expression of klotho(KL)(P<0.05)and cytochrome P450 family 24 subfamily A member 1(CYP24)(P<0.01)mRNAs was lower on Day 14 than Days 1 and 9.ADAM17,FGF23,CYP2R1,CYP27B1,KL,and VDR proteins immunolocalized to the uterine myometrium,blood vessels,and uterine luminal(LE),superficial glandular(sGE),and glandular(GE)epithelia.S100A9 protein was weakly expressed in the uterine myometrium,LE,sGE,and GE.Immunoreactivity of CYP2R1 and KL proteins in uterine LE and sGE was less on Day 1 than on Days 9 and 14.In contrast,S100G protein was expressed exclusively by GE,and immunoreactive S100G protein was less on Day 9.S100A12 protein localized to stromal cells of the uterine stratum spongiosum and blood vessels,but not by uterine epithelial cells.Conclusion Collectively,these results implicate E2,P4,and PGR in the regulation of phosphate,calcium,and vitamin D signaling in cyclic ewes.展开更多
<span style="font-family:Verdana;">The successful application of assisted reproductive techniques (ARTs) in ovine as in other mammal species relies on many factors among which the quality of the semen ...<span style="font-family:Verdana;">The successful application of assisted reproductive techniques (ARTs) in ovine as in other mammal species relies on many factors among which the quality of the semen used. After collection, semen samples are generally processed for storage (liquid storage or cryoconservation) before being used for insemination or </span><i><i><span style="font-family:Verdana;">in vitro</span></i><span></span></i><span style="font-family:Verdana;"> embryo production. During the liquid storage process, sperm cells are exposed to artificial conditions which lead to oxidative stress—the imbalance between pro-oxidants and antioxidants (AO), following overproduction of reactive oxygen species (ROS)</span><span style="font-family:Verdana;">—resulting in ultrastructural, biochemical and functional damages of spermatozoa. Especially, viability, motility, mitochondrial activity, membrane integrity, and</span><span style="font-family:;" "=""> </span><span style="font-family:Verdana;">acrosome integrity are reduced while morphological abnormalities, DNA fragmentation, and lipid peroxidation (LPO) are increased, affecting the fertilizing ability and subsequent early embryonic development when using standard extenders. Indeed, an optimal semen extender must not only regulate and support an environment of adequate pH and buffering capacity to protect spermatozoa from osmotic and cooling stresses, but, also prevent the generation and/or scavenge excess ROS. To improve ram semen liquid storage, several methods have been developed with the supplementation of extenders with antioxidants or antioxidant like-compounds (enzymes, amino-acids, vitamins, plant extracts), seminal plasma, sugars, fatty acids, and nanoparticles being a relevant approach. Promising results have been registered with the supplementation of extenders with these compounds, confirming they can be used to preserve ram semen quality and fertility. Therefore, the present review provides an updated overview of the damages and associated mechanisms that ram spermatozoa undergo during liquid storage. Moreover, the supplementation of extenders with different compounds as a tool to improve semen storage is also discussed as well as their efficiency to reduce and/or prevent sperm damages during storage.</span>展开更多
High energy bone fractures resulting from impact trauma are often accompanied by subcutaneous soft tissue injuries, even if the skin remains intact. There is evidence that such closed soft tissue injuries affect the h...High energy bone fractures resulting from impact trauma are often accompanied by subcutaneous soft tissue injuries, even if the skin remains intact. There is evidence that such closed soft tissue injuries affect the healing of bone fractures, and vice versa. Despite this knowledge, most impact trauma studies in animals have focussed on bone fractures or soft tissue trauma in isolation. However, given the simultaneous impact on both tissues a better understanding of the interaction between these two injuries is necessary to optimise clinical treatment. The aim of this study was therefore to develop a new experimental model and characterise, for the first time, the healing of a complex fracture with concurrent closed soft tissue trauma in sheep. A pendulum impact device was designed to deliver a defined and standardised impact to the distal thigh of sheep, causing a reproducible contusion injury to the subcutaneous soft tissues. In a subsequent procedure, a reproducible femoral butterfly fracture (AO C3-type) was created at the sheep’s femur, which was initially stabilised for 5 days by an external fixator construct to allow for soft tissue swelling to recede, and ultimately in a bridging construct using locking plates. The combined injuries were applied to twelve sheep and the healing observed for four or eight weeks (six animals per group) until sacrifice. The pendulum impact led to a moderate to severe circumferential soft tissue injury with significant bruising, haematomas and partial muscle disruptions. Posttraumatic measurements showed elevated intra-compartmental pressure and circulatory tissue breakdown markers, with recovery to normal, pre-injury values within four days. Clinically, no neurovascular deficiencies were observed. Bi-weekly radiological analysis of the healing fractures showed progressive callus healing over time, with the average number of callus bridges increasing from 0.4 at two weeks to 4.2 at eight weeks. Biomechanical testing after sacrifice showed in- creasing torsional stiffness between four and eight weeks healing time from 10% to 100%, and increasing ultimate torsional strength from 10% to 64% (relative to the contralateral control limb). Our results demonstrate the robust healing of a complex femur fracture in the presence of a severe soft tissue contusion injury in sheep and demonstrate the establishment of a clinically relevant experimental model, for research aimed at improving the treatment of bone fractures accompanied by closed soft tissue injuries.展开更多
Recently the use of biologic materials as dura mater repair patches has been increasing. The purpose of this study is to assess the basis for efficacy and safety of using a novel fish derived acellular dermis (Kerecis...Recently the use of biologic materials as dura mater repair patches has been increasing. The purpose of this study is to assess the basis for efficacy and safety of using a novel fish derived acellular dermis (Kerecis Omega3 DuraTM). In an ovine model a craniotomy under general anaesthesia was performed. A defect was produced in the dural covering of approximately 1 × 2 cm and closed with an onlay technique, with Kerecis Omega3 Dura. The bone defect was covered with the bony flap and the overlying tissues closed in layers. At 2, 5, 8 and 11 weeks the sheep underwent MRI scanning followed by euthanasia, necropsy and histological assessment. MRI images taken at 2, 5, 8 and 11 weeks showed initially moderate inflammatory response, which diminished over time, and at 11 weeks no evidence of inflammation existed. There was evidence of cerebrospinal fluid leakage at no time point. Necropsy revealed some adhesions at 5 and 8 weeks, in particular at 5 weeks, but at 11 weeks there were no adhesions found. From 2 - 11 weeks, there was evidence of initially an inflammatory reaction followed by neodura formation at the defect site through cellular ingrowth and remodeling of the acellular fish skin. Histology showed a histiocytic foreign body reaction initially that subsided over time. As early as 8 weeks there was evidence of neodura formation and by 11 weeks there was a minimal inflammatory response with an intact neodura formed. In this pilot study the Kerecis Omega3 Dura patches performed in a safe and efficacious manner. This new material needs to be fully assessed and compared with other products that are currently on the market in a larger scale animal study.展开更多
The study investigated the effects of epidermal growth factor（EGF） and insulin-like growth factor 1 （ IGF-I）, alone or together, on the in vitro maturation and cleavage of ovine oocytes, aimed to optimize the in v...The study investigated the effects of epidermal growth factor（EGF） and insulin-like growth factor 1 （ IGF-I）, alone or together, on the in vitro maturation and cleavage of ovine oocytes, aimed to optimize the in vitro maturation conditions for ovine oocytes. The results showed that the maturation and cleavage rates were 71.2% and 45.5% respectively when the medium was supplemented with 50 ng/mL EGF alone, which was significantly higher than other EGF supplemented groups （0, 10, 20, 30, and 40 ng/mL） （P 〈0.05）. The highest maturation and cleavage rates were 72.9% and 45. 7% when the EGF concentration reached 100 ng/mL. The maturation and cleavage rates were 70. 7% and 58. 5% with 40 ng/mL IGF-I supplemented, which were significantly higher than other treatments （0,10,20,60,80, and 100 ng/mL） （P 〈0.05）. The lowest maturation and cleavage rates were 38.8% and 20.0% when the IGF-I concentration reached 100 ng/mL （ P 〈 0.05 ） When 50 ng/mL EGF and 40 ng/mL IGF-I were used concomitantly,the maturation and cleavage rates were 85. 6% and 61. 0% respectively, which were significantly higher than the treatments with EGF or IGF-I alone （ P 〈 0.05 ）.展开更多
Objective:To determine the ovine ovarian histomorphology and follicular staging at various age periods in Awassi breed.Methods:Ovaries were collected from prenatal fetuses[gestational age(95±5)days],neonatal(day ...Objective:To determine the ovine ovarian histomorphology and follicular staging at various age periods in Awassi breed.Methods:Ovaries were collected from prenatal fetuses[gestational age(95±5)days],neonatal(day 0),and prepubertal ewe lambs(two and four months of age);each age group included six animals.Ovaries(n=12,each group)were dissected and processed for hematoxylin and eosin staining.Stained sections(n=24,each group)were imaged and utilized for histomorphology assessment,follicle measurement,and classification.Results:Prenatal ovaries were mainly enriched with primordial follicles accompanied by a lower proportion of primary follicles.In addition to primordial and primary follicles,neonatal ovaries demonstrated a proportion of centrally located multilayered and antral follicles.In comparison with neonatal ovaries,the proportion of multilayered and antral follicles was significantly higher in the ovaries of two-month-old lambs;conversely,the proportion of peripherally situated primordial follicles dramatically declined compared to that of earlier age of lamb.Although there was no statistical variation in the sizes of primordial follicles across groups,the mean diameter of the primary follicle in the prenatal ovaries was substantially smaller than in postnatal ovaries.Compared to the neonatal ovaries,the size of the multilayered and antral follicles in the prepubertal ovaries was substantially larger.Conclusions:The earliest follicular developmental stages were established prenatally whereas the advanced growth stages started in the neonatal period and greatly increased in the prepubertal period.展开更多
Background: In recent decades, there has been a growing interest in the impact of insults during pregnancy on postnatal health and disease. It is known that changes in placental development can impact fetal growth and...Background: In recent decades, there has been a growing interest in the impact of insults during pregnancy on postnatal health and disease. It is known that changes in placental development can impact fetal growth and subsequent susceptibility to adult onset diseases;however, a method to collect sufficient placental tissues for both histological and gene expression analyses during gestation without compromising the pregnancy has not been described. The ewe is an established biomedical model for the study of fetal development. Due to its cotyledonary placental type, the sheep has potential for surgical removal of materno-fetal exchange tissues, i.e., placentomes. A novel surgical procedure was developed in well-fed control ewes to excise a single placentome at mid-gestation.Results: A follow-up study was performed in a cohort of nutrient-restricted ewes to investigate rapid placental changes in response to undernutrition. The surgery averaged 19 min, and there were no viability differences between control and sham ewes. Nutrient restricted fetuses were smaller than controls(4.7 ± 0.1 kg vs. 5.6 ± 0.2 kg;P < 0.05), with greater dam weight loss(-32.4 ± 1.3 kg vs. 14.2 ± 2.2 kg;P < 0.01), and smaller placentomes at necropsy(5.7 ± 0.3 g vs. 7.2 ± 0.9 g;P < 0.05). Weight of sampled placentomes and placentome numbers did not differ.Conclusions: With this technique, gestational studies in the sheep model will provide insight into the onset and complexity of changes in gene expression in placentomes resulting from undernutrition(as described in our study),overnutrition, alcohol or substance abuse, and environmental or disease factors of relevance and concern regarding the reproductive health and developmental origins of health and disease in humans and in animals.展开更多
Objective To investigate the development and characterizations of the hepatocytes isolated from fetal ovine and to determine the effect of hypoxia on their growth and metabolism.Methods Fresh hepatocytes were isolated...Objective To investigate the development and characterizations of the hepatocytes isolated from fetal ovine and to determine the effect of hypoxia on their growth and metabolism.Methods Fresh hepatocytes were isolated from the liver of fetal ovine at late gestation, cultured in specific media, and exposed to normoxia(21% O2) or hypoxia(2% O2).The cellular characteristics and population purity were identified by immunocytochemistry and flow cytometry(FCM).The effects of hypoxia on cell cycle and apoptosis of the hepatocytes were evaluated by FCM, whereas the cellular ultrastructure changes were examined with a transmission electron microscope.Results The cell purity of hepatocytes was over 95%.Under hypoxia exposure, the hepatocytes showed a gradual increase in proportion at the S phase and in proliferative index, followed with a compatible increase in apoptosis and progressively decreased cell viability.Additionally, the organelles of the hepatocytes demonstrated dramatic changes, including swelling of mitochondria, disorder in cristae arrangement, expansion of endoplasmic reticulum, and a large number of circular lipid droplets emerging in the cytoplasm.Conclusion Fetal ovine hepatocytes could be primarily cultured in a short-term culture system with a high purity of over 95% and with their preserved original characteristics.Hypoxia could induce changes in ultrastructural and inhibit the proliferation of cultured fetal ovine hepatocytes through apoptotic mechanisms.展开更多
This study is aimed to investigate the effect of luteinizing hormone （LH） on maturation and fertilization in vitro of ovine oocytes. The ovine oocytes were cultured in vitro in medium with or without LH, and then ch...This study is aimed to investigate the effect of luteinizing hormone （LH） on maturation and fertilization in vitro of ovine oocytes. The ovine oocytes were cultured in vitro in medium with or without LH, and then checked by confocal laser scanning microscope to observe the distribution of cortical granules stained with FITC-LCA during different maturation periods. Similarly, some in vitro matured oocytes were also fertilized in vitro for analysis of their developmental potentiality further. After being cultured in vitro for 4 h, there were significant differences about the rate of germinal vesicle break down （GVBD） between the treatment （with LH） and the control groups （without any hormones） （36.76% vs 18%, P〈0.05）. Further, there were also significant differences of the cleavage and blastocyst rates between these two groups （67.15% vs 42.37%, 21.9% vs 12.71%, P〈0.05, respectively）. The distribution of cortical granules appeared to spread from the edges to the central site of sheep oocytes following their delaying durations of maturation in vitro. It can be concluded that LH may play a role to delay the occurence of GVBD, prolong the maturation duration of cytoplasm, and enhance the nuclear and cytoplasm synchronization of ovine oocytes matured in vitro and finally improve their in vitro developmental potentiality.展开更多
基金supported by grants from the Chinese Special Program for Scientific Research of Public Health (200802012)Chinese National Key Program for Infectious Diseases of China (2009ZX10004‐201, and 2008ZX10004‐002)
文摘Objective Cryptosporidium spp. are prevalent globally and sheep are an important zoonotic reservoir. Little data regarding the rates of Cryptosporidium infections in ovines in China are available. This study assessed the prevalence of Cryptosporidium spp. in pre-weaned ovines from Aba Tibetan and Qiang Autonomous Prefecture in the Sichuan province of China. Methods A total of 213 fecal samples were collected from pre-weaned ovines and were examined microscopically （following modified acid fast staining）. In addition, 18S rRNA genetic sequences were amplified from fecal samples by nested PCR and phylogenetically analyzed. Results The prevalence of Cryptosporidium in the collected samples was at 14.6% （31/213） and four isolates identified by PCR belonged to the Cryptosporidium cervine genotype （Cryptosporidium ubiquiturn） demonstrating that this species was the primary sheep species found in sheep in China. Conclusion The present study suggested that the high incidence of Cryptosporidium in sheep poses a significant public health threat and that surveillance practices must be established to prevent zoonotic disease of humans.
基金Ontario Ministry of Agriculture,Food and Agribusiness(OMAFA)Natural Sciences and Engineering Research Council of Canada(NSERC)+1 种基金AdisseoOntario Sheep Farmers。
文摘Background Selenium(Se)is an essential soil mineral that can be incorporated into animal feedstuffs.Because of a lack of soil Se in some regions,organic or inorganic supplementation strategies must be implemented to prevent deficiencies and promote optimal ovine health.Therefore,the objectives of this study were to assess how inorganic versus organic Se supplementation influenced ewe Se status and immune function during late gestation and postpartum.Dorset Rideau ewes(n=110)were fed a Se deficient diet from gestation d 110 through postpartum d 49 and received one of four daily oral Se treatments diluted in 5 mL of sugar water:0 mg Se,0.3 mg inorganic Se,0.3 mg organic Se,and 0.6 mg organic Se.Throughout the trial,the ewes received various immune challenges,including intramuscular immunizations with a novel antigen(ovalbumin;OVA)on trial d 0 and 10,an intradermal OVA challenge on d 20,and a lipopolysaccharide(LPS)endotoxin challenge on trial d 49.Results The organic Se treatment groups had higher serum Se concentrations on most trial days compared to the 0.3 mg inorganic and control groups(P<0.05).No significant treatment differences were found for the dermal hypersensitivity response to OVA,anti-OVA antibody response,glutathione peroxidase activity,cytokine response,cortisol response,or rectal temperature(P>0.05).However,4 h post-LPS injection,the serum albumin concentration was significantly lower in the 0.3 mg inorganic group compared to both organic Se groups,potentially indicating a higher degree of inflammation in the ewes supplemented with the inorganic Se.Conclusions The results of this study indicate that organic Se supplementation can promote a higher Se status in ewes over time,but Se supplementation during this study period did not affect tested immunological parameters.This lack of difference in immune responsiveness between groups may be due to an absence of true serum Se deficiencies in the Se-deficient group or the levels of Se supplementation being insufficient to significantly improve immunocompetence.
基金funded by the emarked fund for China Agriculture Research System (CARS-39)the National Agricultural Science and Technology Innovation Program
文摘Ovine bones are the major by-products after slaughtered. The present study was conducted to extract and characterize acid soluble collagens （ASC） and pepsin soluble collagens （PSC） from ovine bones （Ujumuqin sheep）. Ovine bones collagen were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis （SDS-PAGE） and liquid chromatography-tan- dem mass spectrometry （LC-MS/MS） as type I collagen. The results of Fourier transform infrared （FTIR） spectra analysis testified the existence of triple superhelical structure in both ASC and PSC, showing pepsin did not disrupt the triple helical structure of ovine bones collagen. Glycine, accounting for one-third of total amino acids, was the major amino acid for ovine bones collagen. Higher imino acid content was responsible for higher thermal denaturation temperature of ovine bones collagen compared to fish collagens. The isoelectric point of ASC was lower than PSC due to the higher content of acidic amino acids. Therefore, this study provides the potential reference for collagen extraction and application of ovine bones by-procduct.
基金financial support from the National Natural Science Foundation of China (31771995)the earmarked fund for China Agriculture Research System (CARS-38)the Agricultural Science and Technology Innovation Program, Chinese Academy of Agricultural Sciences (CAAS-ASTIP-IFST)。
文摘Phosphorylation post-translational modification plays an important role in postmortem muscle quality traits. Adenosine triphosphate(ATP) is an energy source and a key substrate of phosphorylation which provides the phosphatase groups to proteins in the presence of protein kinases. However, in postmortem muscle, the effects of ATP content on phosphorylation are poorly studied. The study investigated the effect of ATP on protein phosphorylation and degradation in postmortem ovine muscle. The ground muscle with/without additional ATP were treated/control groups and stored at 25 and 4℃, respectively. The ATP content led to different changes of p H value between the ATP-treated and control groups. The phosphorylation level of myofibrillar proteins was higher(P<0.05) in ATP-treated group compared to the control group at both temperatures, which suggested that ATP played a vital role in postmortem protein phosphorylation. A slower degradation rate of μ-calpain, desmin and troponin T was observed in the ATP-treated group which showed that there was a negative relationship between ATP level and the degradation of proteins. These observations clearly highlighted the role of ATP on the development of meat quality by regulating the phosphorylation and degradation of myofibrillar proteins in postmortem ovine muscle.
基金The Chinese National Natural Science Foundation(No.30960271 and No.31160493)Inner Mongolia technological innovation fund (20101808)the Fund of innovation research team(NDPYTD2010-6)
文摘Endogenous beta retroviruses (enJSRV) are highly homologous with Jaagsiekte sheep retrovirus (exJSRV),this exogenous retrovirus is the aetiological agent of ovine pulmonary adenocarcinoma (OPA). The aim of this study was to clarify the function of enJSRV and the immunological mechanisms of its corresponding antibody, that is undetectable in JSRV-infected ovine serum. The expression of enJSRV envelope protein and Hyal-2 mRNA in immune organs and lungs of ovine fetuses and lambs were analyzed by Real-Time reverse transcription PCR and In Situ Hybridization using specific probes. In Situ Hybridization results indicated that the enJSRV envelope protein and Hyal-2 mRNA were expressed in thymus, spleen, mesenteric lymph nodes and lungs at different times, while no positive signals were detected in the negative controls. On the other hand, results from Real-Time reverse transcription PCR analysis showed that in 130d fetuses and 3d newborn lambs the enJSRV mRNA levels were much higher in organs associated with the immune system than that in lungs, especially in the thymus and spleen, but levels of Hyal-2 mRNA expression was not significantly different in all collected tissue. These results provided evidence from an immunology point of view to understand why the circulating antibodies against exJSRV are undetectable in JSRV-infected ovine, and will help to unravel the pathogenesis of JSRV-infected ovine.
基金funded by Fondazione Banco di Sardegna,FDS 2016(CUP J86C18000780005 and J86C18000810005)。
文摘Background:Heavy metal cadmium(Cd)is a widespread environmental contaminant with a potential toxicity that might negatively affect female reproduction and fertility.It has been reported that Cd exposure impaired the quality of oocytes and led to a defective maturation and fertilization,through oxidative stress induction.Resveratrol(Res)is a natural polyphenol with strong antioxidant properties that exhibited protective role in preventing oocyte redox homeostasis disruption and quality decline.Here,we explored whether the addition of Res to in vitro maturation(IVM)medium might act as a protection against Cd-induced toxicity on ovine oocyte maturation and fertilization.Firstly,we evaluated the effect of supplementing IVM medium with two different Res concentrations(1and 2μmol/L)on nuclear maturation and fertilization of oocytes matured under CdCl2(2μmol/L)exposure.Therefore,the concentration of 1μmol/L Res was selected to analyse the effects of this compound on intracellular ROS levels,mitochondrial(mt)distribution and activity,chromatin configuration,cytoskeleton morphology,cortical granules(CGs)distribution and mRNA expression of genes associated with cellular response to oxidative stress(i.e.SIRT1,SOD 1,GPX1,GSR,CAT)in Cd-exposed in vitro matured oocytes.Results:We found that 1μmol/L Res restored the reduced oocyte meiotic competence induced by Cd exposure as well as,Res sustained oocyte ability to be normally fertilized and decreased polyspermic fertilization at both tested concentrations.Moreover,we demonstrated that 1μmol/L Res mitigated Cd-induced alterations of oocyte cytoplasmic maturation by reducing reactive oxygen species(ROS)accumulation,preventing mt dysfunction,maintaining the correct meiotic spindle and cortical F-actin assembly and the normal cortical granule distribution as well as up-regulating SIRT1,SOD1 and GPX1 genes.Conclusions:Taken together,our findings highlighted the beneficial influence exerted by Res in preventing Cdinduced disturbance of nuclear and cytoplasmic maturation and subsequent fertilization in ovine oocytes.Res treatment may help to establish defence strategies counteracting Cd-induced toxicity on the female gamete.
基金the National Key Research and Development Program of China(2021YFD18005022017YFD0502304)+2 种基金National Natural Science Foundation of China(31672562)NBCITS(CARS-37)ASTIP(CAAS-ASTIP-2016-LVRI)。
文摘ISG20 is an interferon-inducible exonuclease that inhibits virus replication.Although ISG20 is thought to degrade viral RNA,the antiviral mechanism and specificity of ISG20 remain unclear.In this study,the antiviral role of ovine ISG20(o ISG20)in bluetongue virus(BTV)infection was investigated.It was found that BTV infection upregulated the transcription of ovine ISG20(o ISG20)in a time-and BTV multiplicity of infection(MOI)-dependent manner.Overexpression of o ISG20 suppressed the production of BTV genome,proteins,and virus titer,whereas the knockdown of o ISG20 increased viral replication.o ISG20 was found to co-localize with BTV proteins VP4,VP5,VP6,and NS2,but only directly interacted with VP4.Exonuclease defective o ISG20 significantly decreased the inhibitory effect on BTV replication.In addition,the interaction of mutant o ISG20 and VP4 was weakened,suggesting that binding to VP4 was associated with the inhibition of BTV replication.The present data characterized the anti-BTV effect of o ISG20,and provides a novel clue for further exploring the inhibition mechanism of double-stranded RNA virus by ISG20.
基金National Natural Science Foundation ofChina (Grant No. 30560108)
文摘In order to amplify the complete genome of enJSRV from the strain of Inner Mongolia (enJSRV-NM), we used enJSRV-specific and JSRV-specific DNA probes in dot blot hybridization. Seven pairs of primers were designed based on Genbank sequences. Seven fragments were obtained by PCR and were cloned into the PMD19-T vectors. The recombinant plasmids were sequenced and analyzed. The results showed that the genome was 7 942 bp in length and contained four overlapping open reading frames corresponding to the gag, pro, pol and env genes as well as an additional open reading frame (orf-x) that overlaps the 3' end of the pol gene. The nucleotide acid sequences of the enJSRV-NM loci were compared with the sequences of South Africa enJS56A1 strain (Accession No. AF153615) and USA JSRV21 strain (Accession No. AF105220). The nucleotide acid identities were 99.2% and 92.3% respectively. Two zinc fingers were found in the NC region in the predicted amino acid sequence. However, the YXXM motif, which is a reliable molecular marker for the infectious exogenous virus, was not found in the TM region. It was found that the enJSRV-NM region was 90%-98% identical at the amino acid level to its exogenous infectious counterparts in most of the retroviral genome. This is the first nucleotide sequence of enJSRV reported in P.R China. The resource work has provided a wide range of information useful not only for expression genomics and annotation of genomic DNA sequence, but also for further research on the clinical diagnosis of OPA.
基金supported by Regione Autonoma della Sardegna.-L.R.7-MIGLIOVINGENSAR ProjectBando competitivo Fondazione di Sardegna–2016,CUP J86C18000800005“Progetto FAR2019LEDDAS Una Tantum 2019,University of Sassari”.
文摘Background:To advance the use of embryo vitrification in veterinary practice,we developed a system in which embryo vitrification,warming and dilution can be performed within a straw.Ovine in vitro produced embryos(IVEP)were vitrified at either early(EBs:n=74)or fully expanded blastocyst stage(FEBs:n=195),using a new device named"E.Vit",composed by a 0.25-m L straw with a 50-μm pore polycarbonate grid at one end.Embryos at each stage(EBs and FEBs)were vitrified by either Two-step(TS)or Multi-step(MS;6 different concentrations of vitrification solutions)protocol.Non-vitrified embryos(n=102)were maintained in in vitro culture as a control.Warming consisted of placing the straws directly into 1.5 m L tubes containing a TCM-199 solution with three decreasing concentrations of sucrose.Blastocyst re-expansion,embryo survival and hatching rate were evaluated at2,24 and 48 h post warming.The number of apoptotic cells was determined by TUNEL assay.Results:Blastocyst re-expansion(2 h)after warming was higher(P<0.05)in FEBs group,vitrified with the MS and TS methods(77.90%and 71.25%,respectively)compared with the EBs group(MS:59.38%and TS:48.50%,respectively).Survival rates of vitrified FEBs after 24 h IVC were higher(P<0.001)in both methods(MS and TS)than vitrified EBs(MS:56.25%;TS:42.42%)and was higher(P<0.05)in the MS method(94.19%)compared with those in TS(83.75%).After 48 h of culture the hatching rate for FEBs vitrified in MS system(91.86%)was similar to control(91.89%),but higher than FEB TS(77.5%)and EBs vitrified in MS(37.5%)and TS(33.33%).Number of apoptotic cells were higher in EBs,irrespective of the system used,compared to FEBs.The number of apoptotic cells in FEBs vitrified with MS was comparable to the control.Conclusions:A high survival rate of IVP embryos can be achieved by the new"E.Vit"device with hatching rates in vitro comparable with control fresh embryos.This method has the potential for use in direct embryo transfer in field conditions.
基金funded by the National Agricultural Science and Technology Innovation Program in China
文摘This study aimed to examine changes in phosphorylation of sarcoplasmic and myofibrillar proteins from longissimus lumborum,semitendinosus,and psoas major muscles during postmortem ageing for 5 d.These sarcoplasmic and myofibrillar proteins were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with phosphorous and protein specific stains.Myofibril fragmentation index,pH,the content of lactic acid and the relative activity of μ-calpain in three ovine muscles were measured.These results showed that the relative phosphorylation level of sarcoplasmic and myofibrillar proteins of psoas major muscle were lower compared with longissimus lumborum and semitendinosus muscles(P<0.05).The pH of psoas major muscle was the lowest at 0.5 h postmortem,and the highest after 12 h postmortem(P<0.05).In addition,the relative activity of μ-calpain was higher within 5 d postmortem and myofibril fragmentation index was higher after 1 d postmortem in psoas major muscle than those of longissimus lumborum and semitendinosus muscles(P<0.05).The sarcoplasmic protein phosphorylation may regulate the rate of pH decline to influence the μ-calpain activity and then proteolysis of proteins consequently.This study gives a new perspective of the mechanism of postmortem meat tenderization.
基金supported by Agriculture and Food Research Initiative Competitive Grant 2016-67015-24958 from the USDA National Institute of Food and Agriculture.
文摘Background Recent evidence suggests important roles for progesterone(P4)and interferon tau in the regulation of calcium,phosphate,and vitamin D signaling in the uteri of pregnant sheep.However,the effects of P4 and estradiol(E2),with respect to the expression of their receptors PGR and ESR1,respectively,in uterine epithelia on mineral signaling during the estrous cycle has not been investigated.Estrous cycles of mature Suffolk ewes were synchronized,prostaglandin F2αwas administered,and ewes were observed for estrus(designated as Day 0)in the presence of vasectomized rams.On Days 1,9,or 14 of the estrous cycle,hysterectomies were performed.Results 25-hydroxyvitamin D was more abundant in plasma from ewes on Day 14 than Day 1(P<0.05).Expression of fibroblast growth factor receptor 2(FGFR2),a disintegrin and metalloprotease 17(ADAM17),and parathyroid hormone-related protein(PTHrP)mRNAs was greater in endometria on Day 9 compared to Days 1 and 14(P<0.01).Similarly,expression of transient receptor potential cation channel subfamily V member 6(TRPV6)mRNA was greater in endometria on Day 9 than Day 1(P<0.05).ATPase plasma membrane Ca^(2+)transporting 4(ATP2B4)and S100 calcium binding protein G(S100G)mRNA expression was greater in endometria on Day 14 than on Days 1 and 9(P<0.01).In contrast,endometrial expression of vitamin D receptor(VDR)mRNA was lower on Days 9 and 14 than Day 1(P<0.01).Expression of klotho(KL)(P<0.05)and cytochrome P450 family 24 subfamily A member 1(CYP24)(P<0.01)mRNAs was lower on Day 14 than Days 1 and 9.ADAM17,FGF23,CYP2R1,CYP27B1,KL,and VDR proteins immunolocalized to the uterine myometrium,blood vessels,and uterine luminal(LE),superficial glandular(sGE),and glandular(GE)epithelia.S100A9 protein was weakly expressed in the uterine myometrium,LE,sGE,and GE.Immunoreactivity of CYP2R1 and KL proteins in uterine LE and sGE was less on Day 1 than on Days 9 and 14.In contrast,S100G protein was expressed exclusively by GE,and immunoreactive S100G protein was less on Day 9.S100A12 protein localized to stromal cells of the uterine stratum spongiosum and blood vessels,but not by uterine epithelial cells.Conclusion Collectively,these results implicate E2,P4,and PGR in the regulation of phosphate,calcium,and vitamin D signaling in cyclic ewes.
文摘<span style="font-family:Verdana;">The successful application of assisted reproductive techniques (ARTs) in ovine as in other mammal species relies on many factors among which the quality of the semen used. After collection, semen samples are generally processed for storage (liquid storage or cryoconservation) before being used for insemination or </span><i><i><span style="font-family:Verdana;">in vitro</span></i><span></span></i><span style="font-family:Verdana;"> embryo production. During the liquid storage process, sperm cells are exposed to artificial conditions which lead to oxidative stress—the imbalance between pro-oxidants and antioxidants (AO), following overproduction of reactive oxygen species (ROS)</span><span style="font-family:Verdana;">—resulting in ultrastructural, biochemical and functional damages of spermatozoa. Especially, viability, motility, mitochondrial activity, membrane integrity, and</span><span style="font-family:;" "=""> </span><span style="font-family:Verdana;">acrosome integrity are reduced while morphological abnormalities, DNA fragmentation, and lipid peroxidation (LPO) are increased, affecting the fertilizing ability and subsequent early embryonic development when using standard extenders. Indeed, an optimal semen extender must not only regulate and support an environment of adequate pH and buffering capacity to protect spermatozoa from osmotic and cooling stresses, but, also prevent the generation and/or scavenge excess ROS. To improve ram semen liquid storage, several methods have been developed with the supplementation of extenders with antioxidants or antioxidant like-compounds (enzymes, amino-acids, vitamins, plant extracts), seminal plasma, sugars, fatty acids, and nanoparticles being a relevant approach. Promising results have been registered with the supplementation of extenders with these compounds, confirming they can be used to preserve ram semen quality and fertility. Therefore, the present review provides an updated overview of the damages and associated mechanisms that ram spermatozoa undergo during liquid storage. Moreover, the supplementation of extenders with different compounds as a tool to improve semen storage is also discussed as well as their efficiency to reduce and/or prevent sperm damages during storage.</span>
文摘High energy bone fractures resulting from impact trauma are often accompanied by subcutaneous soft tissue injuries, even if the skin remains intact. There is evidence that such closed soft tissue injuries affect the healing of bone fractures, and vice versa. Despite this knowledge, most impact trauma studies in animals have focussed on bone fractures or soft tissue trauma in isolation. However, given the simultaneous impact on both tissues a better understanding of the interaction between these two injuries is necessary to optimise clinical treatment. The aim of this study was therefore to develop a new experimental model and characterise, for the first time, the healing of a complex fracture with concurrent closed soft tissue trauma in sheep. A pendulum impact device was designed to deliver a defined and standardised impact to the distal thigh of sheep, causing a reproducible contusion injury to the subcutaneous soft tissues. In a subsequent procedure, a reproducible femoral butterfly fracture (AO C3-type) was created at the sheep’s femur, which was initially stabilised for 5 days by an external fixator construct to allow for soft tissue swelling to recede, and ultimately in a bridging construct using locking plates. The combined injuries were applied to twelve sheep and the healing observed for four or eight weeks (six animals per group) until sacrifice. The pendulum impact led to a moderate to severe circumferential soft tissue injury with significant bruising, haematomas and partial muscle disruptions. Posttraumatic measurements showed elevated intra-compartmental pressure and circulatory tissue breakdown markers, with recovery to normal, pre-injury values within four days. Clinically, no neurovascular deficiencies were observed. Bi-weekly radiological analysis of the healing fractures showed progressive callus healing over time, with the average number of callus bridges increasing from 0.4 at two weeks to 4.2 at eight weeks. Biomechanical testing after sacrifice showed in- creasing torsional stiffness between four and eight weeks healing time from 10% to 100%, and increasing ultimate torsional strength from 10% to 64% (relative to the contralateral control limb). Our results demonstrate the robust healing of a complex femur fracture in the presence of a severe soft tissue contusion injury in sheep and demonstrate the establishment of a clinically relevant experimental model, for research aimed at improving the treatment of bone fractures accompanied by closed soft tissue injuries.
文摘Recently the use of biologic materials as dura mater repair patches has been increasing. The purpose of this study is to assess the basis for efficacy and safety of using a novel fish derived acellular dermis (Kerecis Omega3 DuraTM). In an ovine model a craniotomy under general anaesthesia was performed. A defect was produced in the dural covering of approximately 1 × 2 cm and closed with an onlay technique, with Kerecis Omega3 Dura. The bone defect was covered with the bony flap and the overlying tissues closed in layers. At 2, 5, 8 and 11 weeks the sheep underwent MRI scanning followed by euthanasia, necropsy and histological assessment. MRI images taken at 2, 5, 8 and 11 weeks showed initially moderate inflammatory response, which diminished over time, and at 11 weeks no evidence of inflammation existed. There was evidence of cerebrospinal fluid leakage at no time point. Necropsy revealed some adhesions at 5 and 8 weeks, in particular at 5 weeks, but at 11 weeks there were no adhesions found. From 2 - 11 weeks, there was evidence of initially an inflammatory reaction followed by neodura formation at the defect site through cellular ingrowth and remodeling of the acellular fish skin. Histology showed a histiocytic foreign body reaction initially that subsided over time. As early as 8 weeks there was evidence of neodura formation and by 11 weeks there was a minimal inflammatory response with an intact neodura formed. In this pilot study the Kerecis Omega3 Dura patches performed in a safe and efficacious manner. This new material needs to be fully assessed and compared with other products that are currently on the market in a larger scale animal study.
文摘The study investigated the effects of epidermal growth factor（EGF） and insulin-like growth factor 1 （ IGF-I）, alone or together, on the in vitro maturation and cleavage of ovine oocytes, aimed to optimize the in vitro maturation conditions for ovine oocytes. The results showed that the maturation and cleavage rates were 71.2% and 45.5% respectively when the medium was supplemented with 50 ng/mL EGF alone, which was significantly higher than other EGF supplemented groups （0, 10, 20, 30, and 40 ng/mL） （P 〈0.05）. The highest maturation and cleavage rates were 72.9% and 45. 7% when the EGF concentration reached 100 ng/mL. The maturation and cleavage rates were 70. 7% and 58. 5% with 40 ng/mL IGF-I supplemented, which were significantly higher than other treatments （0,10,20,60,80, and 100 ng/mL） （P 〈0.05）. The lowest maturation and cleavage rates were 38.8% and 20.0% when the IGF-I concentration reached 100 ng/mL （ P 〈 0.05 ） When 50 ng/mL EGF and 40 ng/mL IGF-I were used concomitantly,the maturation and cleavage rates were 85. 6% and 61. 0% respectively, which were significantly higher than the treatments with EGF or IGF-I alone （ P 〈 0.05 ）.
文摘Objective:To determine the ovine ovarian histomorphology and follicular staging at various age periods in Awassi breed.Methods:Ovaries were collected from prenatal fetuses[gestational age(95±5)days],neonatal(day 0),and prepubertal ewe lambs(two and four months of age);each age group included six animals.Ovaries(n=12,each group)were dissected and processed for hematoxylin and eosin staining.Stained sections(n=24,each group)were imaged and utilized for histomorphology assessment,follicle measurement,and classification.Results:Prenatal ovaries were mainly enriched with primordial follicles accompanied by a lower proportion of primary follicles.In addition to primordial and primary follicles,neonatal ovaries demonstrated a proportion of centrally located multilayered and antral follicles.In comparison with neonatal ovaries,the proportion of multilayered and antral follicles was significantly higher in the ovaries of two-month-old lambs;conversely,the proportion of peripherally situated primordial follicles dramatically declined compared to that of earlier age of lamb.Although there was no statistical variation in the sizes of primordial follicles across groups,the mean diameter of the primary follicle in the prenatal ovaries was substantially smaller than in postnatal ovaries.Compared to the neonatal ovaries,the size of the multilayered and antral follicles in the prepubertal ovaries was substantially larger.Conclusions:The earliest follicular developmental stages were established prenatally whereas the advanced growth stages started in the neonatal period and greatly increased in the prepubertal period.
文摘Background: In recent decades, there has been a growing interest in the impact of insults during pregnancy on postnatal health and disease. It is known that changes in placental development can impact fetal growth and subsequent susceptibility to adult onset diseases;however, a method to collect sufficient placental tissues for both histological and gene expression analyses during gestation without compromising the pregnancy has not been described. The ewe is an established biomedical model for the study of fetal development. Due to its cotyledonary placental type, the sheep has potential for surgical removal of materno-fetal exchange tissues, i.e., placentomes. A novel surgical procedure was developed in well-fed control ewes to excise a single placentome at mid-gestation.Results: A follow-up study was performed in a cohort of nutrient-restricted ewes to investigate rapid placental changes in response to undernutrition. The surgery averaged 19 min, and there were no viability differences between control and sham ewes. Nutrient restricted fetuses were smaller than controls(4.7 ± 0.1 kg vs. 5.6 ± 0.2 kg;P < 0.05), with greater dam weight loss(-32.4 ± 1.3 kg vs. 14.2 ± 2.2 kg;P < 0.01), and smaller placentomes at necropsy(5.7 ± 0.3 g vs. 7.2 ± 0.9 g;P < 0.05). Weight of sampled placentomes and placentome numbers did not differ.Conclusions: With this technique, gestational studies in the sheep model will provide insight into the onset and complexity of changes in gene expression in placentomes resulting from undernutrition(as described in our study),overnutrition, alcohol or substance abuse, and environmental or disease factors of relevance and concern regarding the reproductive health and developmental origins of health and disease in humans and in animals.
基金funded by National Natural Science Foundation [81370719 and 81671535]Jiangsu Science Foundation [BE2015642]+3 种基金Jiangsu Key Discipline of Human Assisted Reproduction Medicine Foundation [FXK201749]Jiangsu Provincial Medical Talent of the Project of Invigorating Healthcare through Science,Technology and Education [ZDRCA2016044]and Chinese Medical Association Clinical Medicine Research Reproductive Medicine [17020270696]The Priority Academic Program Development of the Jiangsu Higher Education Institutes(PAPD)
文摘Objective To investigate the development and characterizations of the hepatocytes isolated from fetal ovine and to determine the effect of hypoxia on their growth and metabolism.Methods Fresh hepatocytes were isolated from the liver of fetal ovine at late gestation, cultured in specific media, and exposed to normoxia(21% O2) or hypoxia(2% O2).The cellular characteristics and population purity were identified by immunocytochemistry and flow cytometry(FCM).The effects of hypoxia on cell cycle and apoptosis of the hepatocytes were evaluated by FCM, whereas the cellular ultrastructure changes were examined with a transmission electron microscope.Results The cell purity of hepatocytes was over 95%.Under hypoxia exposure, the hepatocytes showed a gradual increase in proportion at the S phase and in proliferative index, followed with a compatible increase in apoptosis and progressively decreased cell viability.Additionally, the organelles of the hepatocytes demonstrated dramatic changes, including swelling of mitochondria, disorder in cristae arrangement, expansion of endoplasmic reticulum, and a large number of circular lipid droplets emerging in the cytoplasm.Conclusion Fetal ovine hepatocytes could be primarily cultured in a short-term culture system with a high purity of over 95% and with their preserved original characteristics.Hypoxia could induce changes in ultrastructural and inhibit the proliferation of cultured fetal ovine hepatocytes through apoptotic mechanisms.
基金supported in part by the grants fromthe National Natural Science Foundation of China(30871836) the Key Fund of Natural Science from Beijing Municipal Government, China (Type B,KZ200510020013)
文摘This study is aimed to investigate the effect of luteinizing hormone （LH） on maturation and fertilization in vitro of ovine oocytes. The ovine oocytes were cultured in vitro in medium with or without LH, and then checked by confocal laser scanning microscope to observe the distribution of cortical granules stained with FITC-LCA during different maturation periods. Similarly, some in vitro matured oocytes were also fertilized in vitro for analysis of their developmental potentiality further. After being cultured in vitro for 4 h, there were significant differences about the rate of germinal vesicle break down （GVBD） between the treatment （with LH） and the control groups （without any hormones） （36.76% vs 18%, P〈0.05）. Further, there were also significant differences of the cleavage and blastocyst rates between these two groups （67.15% vs 42.37%, 21.9% vs 12.71%, P〈0.05, respectively）. The distribution of cortical granules appeared to spread from the edges to the central site of sheep oocytes following their delaying durations of maturation in vitro. It can be concluded that LH may play a role to delay the occurence of GVBD, prolong the maturation duration of cytoplasm, and enhance the nuclear and cytoplasm synchronization of ovine oocytes matured in vitro and finally improve their in vitro developmental potentiality.
