The generation of reactive oxygen species (ROS) and subsequent mitochondrial and DNA damage in bovine somatic cell nuclear transfer (SCNT) embryos were examined. Bovine enucleated oocytes were electrofused with donor cells and then activated by a combination of Ca-ionophore and 6-dimethylaminopurine culture. The H2O2 and ˙OH radical levels, mitochondrial morphology and membrane potential (ΔΨ), and DNA fragmentation of SCNT and in vitro fertilized (IVF) embryos at the zygote stage were analyzed. The H2O2 (35.6 ± 1.1 pixels/embryo) and ˙OH radical levels (44.6 ± 1.2 pixels/embryo) of SCNT embryos were significantly higher than those of IVF embryos (19.2 ± 1.5 and 23.8 ± 1.8 pixels/embryo, respectively, p < 0.05). The mitochondria morphology of SCNT embryos was diffused within the cytoplasm. The ΔΨ of SCNT embryos was significantly lower (p < 0.05) than that of IVF embryos (0.95 ± 0.04 vs. 1.21 ± 0.06, red/green). Moreover, the comet tail length of SCNT embryos was longer than that of IVF embryos (515.5 ± 26.4 µm vs. 425.6 ± 25.0 µm, p < 0.05). These results indicate that mitochondrial and DNA damage increased in bovine SCNT embryos, which may have been induced by increased ROS levels.
cattle; cellular damage; DNA fragmentation; ROS generation; somatic cell nuclear transfer
We analyzed alcoholic extracts of herbs possessing anti-neosporal activity against Neospora (N.) caninum. To identify the chemical components of Sophora (S.) flavescens and Torilis (T.) japonica associated with anti-neosporal activity, specific fractions were isolated by high-performance liquid chromatography (HPLC). In vitro activity of the fractions against N. caninum was then assessed. Gas chromatography/mass spectrometry (GC/MS) was used to identify and quantify specific anti-neosporal molecules in the herbal extracts. Almost all HPLC fractions of S. flavescens and T. japonica had higher levels of anti-neosporal activity compared to the not treated control. Active constituents of the extracts were sophoridane, furosardonin A, and tetraisopropylidene-cyclobutane in S. flavescens; 5,17-β-dihydroxy-de-A-estra-5,7,9,14-tetraene, furanodiene, and 9,12-octadecadienoic acid (Z,Z)-(CAS,1) in T. japonica.
GC/MS; HPLC; Neospora caninum; Sophora flavescens; Torilis japonica
This study was conducted to investigate whether administration of IH901, a ginseng intestinal metabolite, ameliorates exercise-induced oxidative stress while preserving antioxidant defense capability in rat skeletal muscles and lung. Eight adult male Sprague-Dawley rats per group were randomly assigned to the resting control, exercise control, resting with IH901 (25, 50, and 100 mg/kg) consumption (R/IH901), or exercise with IH901 (25, 50, and 100 mg/kg) consumption (E/IH901) group. The trained groups ran 35 min 2 days/week for 8 weeks. To analyze the IH901-training interaction, serum biochemical analysis, lipid peroxidation, citrate synthase, protein oxidation, antioxidant and superoxide dismutase in skeletal muscles and lung tissue were measured. Compared to the exercise control group, animals that consumed IH901 had significantly increased exercise endurance times (p < 0.05) and decreased plasma creatine kinase and lactate dehydrogenase levels (p < 0.05), while those in the E/IH901 groups had increased citrate synthase and anti-oxidant enzymes and decreased lipid peroxidation and protein oxidation (p < 0.05). In conclusion, IH901 consumption in aging rats after eccentric exercise has beneficial effects on anti-inflammatory and anti-oxidant activities through down-regulation of pro-inflammatory mediators, lipid peroxidation, and protein oxidation and up-regulation of anti-oxidant enzymes.
anti-oxidant activity; eccentric exercise; IH901; lipid peroxidation; rat
Vitamin D3 up-regulated protein 1 (VDUP1) is a potent growth suppressor that inhibits tumor cell proliferation and cell cycle progression when overexpressed. In a previous study, we showed that VDUP1 knockout (KO) mice exhibited accelerated liver regeneration because such animals could effectively control the expression of cell cycle regulators that drive the G1-to-S phase progression. In the present study, we further investigated the role played by VDUP1 in initial priming of liver regeneration. To accomplish this, VDUP1 KO and wild-type (WT) mice were subjected to 70% partial hepatectomy (PH) and sacrificed at different times after surgery. The hepatic levels of TNF-α and IL-6 increased after PH, but there were no significant differences between VDUP1 KO and WT mice. Nuclear factor-κB (NF-κB), c-Jun-N-terminal kinase (JNK), and signal transducer and activator of transcription 3 (STAT-3) were activated much earlier and to a greater extent in VDUP1 KO mice after PH. A single injection of TNF-α or IL-6 caused rapid activation of JNK and STAT-3 expression in both mice, but the responses were stronger and more sustained in VDUP1 KO mice. In conclusion, our findings provide evidence that VDUP1 plays a role in initiation of liver regeneration.
c-Jun-N-terminal kinase; NF-κB; partial hepatectomy; signal transducer and activator of transcription 3; Vitamin D3 up-regulated protein 1
Verocytotoxic Escherichia (E.) coli strains are responsible for swine oedema disease, which is an enterotoxaemia that causes economic losses in the pig industry. The production of a vaccine for oral administration in transgenic seeds could be an efficient system to stimulate local immunity. This study was conducted to transform tobacco plants for the seed-specific expression of antigenic proteins from a porcine verocytotoxic E. coli strain. Parameters related to an immunological response and possible adverse effects on the oral administration of obtained tobacco seeds were evaluated in a mouse model. Tobacco was transformed via Agrobacteium tumefaciens with chimeric constructs containing structural parts of the major subunit FedA of the F18 adhesive fimbriae and VT2e B-subunit genes under control of a seed specific GLOB promoter. We showed that the foreign Vt2e-B and F18 genes were stably accumulated in storage tissue by the immunostaining method. In addition, Balb-C mice receiving transgenic tobacco seeds via the oral route showed a significant increase in IgA-positive plasma cell presence in tunica propria when compared to the control group with no observed adverse effects. Our findings encourage future studies focusing on swine for evaluation of the protective effects of transformed tobacco seeds against E. coli infection.
Agrobacterium tumefaciens; edible vaccines; Escherichia coli; pig; verocytotoxins
We previously determined that AKR/J mice housed in a low-dose-rate (LDR) (137Cs, 0.7 mGy/h, 2.1 Gy) γ-irradiation facility developed less spontaneous thymic lymphoma and survived longer than those receiving sham or high-dose-rate (HDR) (137Cs, 0.8 Gy/min, 4.5 Gy) radiation. Interestingly, histopathological analysis showed a mild lymphomagenesis in the thymus of LDR-irradiated mice. Therefore, in this study, we investigated whether LDR irradiation could trigger the expression of thymic genes involved in the DNA repair process of AKR/J mice. The enrichment analysis of Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes pathways showed immune response, nucleosome organization, and the peroxisome proliferator-activated receptors signaling pathway in LDR-irradiated mice. Our microarray analysis and quantitative polymerase chain reaction data demonstrated that mRNA levels of Lig4 and RRM2 were specifically elevated in AKR/J mice at 130 days after the start of LDR irradiation. Furthermore, transcriptional levels of H2AX and ATM, proteins known to recruit DNA repair factors, were also shown to be upregulated. These data suggest that LDR irradiation could trigger specific induction of DNA repair-associated genes in an attempt to repair damaged DNA during tumor progression, which in turn contributed to the decreased incidence of lymphoma and increased survival. Overall, we identified specific DNA repair genes in LDR-irradiated AKR/J mice.
AKR/J mice; DNA repair genes; low-dose-rate radiation; thymic lymphoma
To investigate the effects of gossypol acetic acid (GA) on proliferation and apoptosis of the macrophage cell line RAW264.7 and further understand the possible underlying mechanism responsible for GA-induced cell apoptosis, RAW264.7 cells were treated with GA (25~35 µmol/L) for 24 h and the cytotoxicity was determined by MTT assay, while apoptotic cells were identified by TUNEL assay, acridine orange/ethidium bromide staining and flow cytometry. Moreover, mitochondrial membrane potential (ΔΨm) with Rhodamine 123 and reactive oxygen species (ROS) with DCFH-DA were analyzed by fluorescence spectrofluorometry. In addition, the expression of caspase-3 and caspase-9 was assessed by Western Blot assay. Finally, the GA-induced cell apoptosis was evaluated by flow cytometry in the present of caspase inhibitors Z-VAD-FMK and Ac-LEHD-FMK, respectively. GA significantly inhibited the proliferation of RAW264.7 cells in a dose-dependent manner, and caused obvious cell apoptosis and a loss of ΔΨm in RAW264.7 cells. Moreover, the ROS production in cells was elevated, and the levels of activated caspase-3 and caspase-9 were up-regulated in a dose-dependent manner. Notably, GA-induced cell apoptosis was markedly inhibited by caspase inhibitors. These results suggest that GA-induced RAW264.7 cell apoptosis may be mediated via a caspase-dependent mitochondrial signaling pathway.
apoptosis; gossypol acetic acid; mitochondrial signaling pathway
A recombinant hemagglutinin-neuraminidase (rHN) protein from Newcastle disease virus (NDV) with hemagglutination (HA) activity was expressed in Spodoptera frugiperda cells using a baculovirus expression system. The rHN protein extracted from infected cells was used as an antigen in a hemagglutination inhibition (HI) test for the detection and titration of NDV-specific antibodies present in chicken sera. The rHN antigen produced high HA titers of 213 per 25 µL, which were similar to those of the NDV antigen produced using chicken eggs, and it remained stable without significant loss of the HA activity for at least 12 weeks at 4℃. The rHN-based HI assay specifically detected NDV antibodies, but not the sera of other avian pathogens, with a specificity and sensitivity of 100% and 98.0%, respectively, in known positive and negative chicken sera (n = 430). Compared with an NDV-based HI assay, the rHN-based HI assay had a relative sensitivity and specificity of 96.1% and 95.5%, respectively, when applied to field chicken sera. The HI titers of the rHN-based HI assay were highly correlated with those in an NDV-based HI assay (r = 0.927). Overall, these results indicate that rHN protein provides a useful alternative to NDV antigen in HI assays.
antibody detection; hemagglutinin-neuraminidase protein; hemagglutination inhibition; Newcastle disease virus
The sequences of the ccrAB genes from bovine-, canine- and chicken-originating methicillin-resistant Staphylococcus (S.) epidermidis (MRSE) and bovine methicillin-resistant Staphylococcus (S.) aureus (MRSA) were compared to investigate the frequency of intra-species horizontal transfer of the staphylococcal cassette chromosome mec (SCCmec) complex. Nineteen MRSE strains were isolated from bovine milk, chickens, and dogs, and their genetic characteristics were investigated by multilocus sequence typing and SCCmec typing. Among the animal MRSE strains, the most frequent SCCmec type was type IV, which consisted of the type B mec complex and ccrAB type 2. The ccrA2 and ccrB2 genes were sequenced from the bovine, chicken and canine MRSE strains and compared with those of the bovine MRSA strains. The sequences generally clustered as MRSA and MRSE groups, regardless of the animal source. Additionally, no bovine MRSE sequence was associated with the bovine MRSA groups. Although most of the bovine MRSE and MRSA isolates possessed SCCmec type IV sequences, our results suggest that the intra-species gene transfer of the SCCmec complex between bovine S. aureus and bovine S. epidermidis strains is not a frequent event.
ccrAB; methicillin-resistant Staphylococcus aureus; methicillin-resistant Staphylococcus epidermidis; SCCmec
The glycoprotein 3 (GP3) of type II porcine reproductive and respiratory syndrome virus has the characteristic domains of a membrane protein. However, this protein has been reported to be retained in the endoplasmic reticulum (ER) rather than transported to the plasma membrane of the cell. In this study, we performed confocal laser scanning microscopy analysis of variants of GP3 and foundthat the signal sequence of the GP3 led to confinement of GP3 in the ER, while the functional ortransmembrane domain did not affect its localization. Based on these results, we concludedthat the signal sequence of GP3 contains the ER retention signal, which might play an important role in assembly of viral proteins.
endoplasmic reticulum; glycoprotein 3; porcine reproductive and respiratory syndrome virus; retention signal; signal sequence
This study was conducted to establish an in vitro maturation (IVM) system by selection of efficient porcine serum during porcine in vitro production. To investigate the efficient porcine serum (PS), different types of PS [newborn pig serum, prepubertal gilt serum (PGS), estrus sow serum, and pregnancy sow serum] were used to supplement IVM media with or without gonadotrophin (GTH) and development rates of parthenogenetic activation (PA) and in vitro fertilization (IVF) embryos were then compared. The maturation rates of the PGS group was significantly higher when GTH was not added. Additionally, during development of PA embryos without GTH, the PGS group showed significantly higher cleavage and blastocyst formation rates. Moreover, the cleavage rates of IVF embryos were significantly higher in the PGS group, with no significant differences in the blastocyst formation. However, when GTH was supplemented into the IVM media, there were no significant differences among the four groups in the cleavage rates, development rates of the blastocyst, and cell number of the blastocyst after PA and IVF. In conclusion, PGS is an efficient macromolecule in porcine IVM, and GTH supplementation of the IVM media is beneficial when PS is used as macromolecule, regardless of its origin.
gonadotropin; in vitro fertilization parthenogenesis; porcine; serum
Transrectal Doppler sonography was used to evaluate uterine blood flow during the first two weeks after parturition in six primiparous Simmental cows. The uterine blood flow was evaluated on the day of parturition (Day 0), once daily from Days 1 to 8 and then every other day until Day 14. Blood flow was quantified by determining the diameter (D), the time-averaged maximum velocity (TAMV), the pulsatility index (PI) and the blood flow volume (BFV) of the uterine arteries ipsilateral and contralateral to the formerly pregnant uterine horn. During the first four days after calving D, TAMV and BFV declined (ipsilateral: TAMV 70%, BFV 87%, contralateral: D 47%, BFV 84%; p < 0.05), while PI increased (ipsilateral 158%, contralateral 100%; p < 0.05) distinctly. Between Days 4 and 14 only the ipsilateral D (12%) and the BFV of both arteries (ipsilateral 5%, contralateral 8%) decreased (p < 0.05). Blood flow variables were very strongly correlated with each other (r > ±0.75, p < 0.05), with negative correlations with PI and positive correlations with all other investigated factors. Overall, this study revealed characteristic changes in uterine perfusion during the first two weeks after parturition in cows that were pronounced during the first four days postpartum.
involution; puerperal; uterus
Here, percutaneous spinal cord injury (SCI) methods using a balloon catheter in adult rats are described. A balloon catheter was inserted into the epidural space through the lumbosacral junction and then inflated between T9-T10 for 10min under fluoroscopic guidance. Animals were divided into three groups with respect to inflation volume: 20 µL (n = 18), 50 µL (n = 18) and control (Fogarty catheter inserted but not inflated; n = 10). Neurological assessments were then made based on BBB score, magnetic resonance imaging and histopathology. Both inflation volumes produced complete paralysis. Gradual recovery of motor function occurred when 20 µL was used, but not after 50 µL was applied. In the 50 µL group, all gray and white matter was lost from the center of the lesion. In addition, supramaximal damage was noted, which likely prevented spontaneous recovery. This percutaneous spinal cord compression injury model is simple, rapid with high reproducibility and the potential to serve as a useful tool for investigation of pathophysiology and possible protective treatments of SCI in vivo.
balloon compression; laminectomy-free; magnetic resonance imaging; percutaneous spinal cord injury; rat
The present study was conducted to compare the effects of xenogenic bovine fetal demineralized bone matrix (DBM), commercial DBM, omentum, omentum-calf fetal DBM, cortical autograft and xenogenic cartilage powder on the healing of tibial defects in a dog model to determine the best material for bone healing. Seven male adult mongrel dogs, weighing 26.2 ± 2.5 kg, were used in this study. Seven holes with a diameter of 4-mm were created and then filled with several biomaterials. Radiographs were taken postoperatively on day 1 and weeks 2, 4, 6, 8. The operated tibias were removed on the 56th postoperative day and histopathologically evaluated. On postoperative days 14, 42 and 56, the lesions of the control group were significantly inferior to those in the other group (p < 0.05). On the 28th postoperative day, the autograft group was significantly superior to the control and omentum groups (p < 0.05). Moreover, calf fetal DBM was significantly superior to the control group. There was no significant difference between the histopathological sections of all groups. Overall, the omentum and omentum-DBM groups were superior to the control group, but inferior to the autograft, commercial-DBM, calf fetal DBM and calf fetal cartilage groups.
autograft; bone healing; calf fetal-demineralized bone matrix; commercial demineralized bone matrix; omentum
The effects of black rice anthocyanidins (BRACs) on retinal damage induced by photochemical stress are not well known. In the present study, Sprague-Dawley rats were fed AIN-93M for 1 week, after which 80 rats were randomly divided into two groups and treated with (n = 40) or without BRACs (n = 40) for 15 days, respectively. After treatment, both groups were exposed to fluorescent light (3,000 ± 200 lux; 25℃), and the protective effect of dietary BRACs were evaluated afterwards. Our results showed that dietary BRACs effectively prevented retinal photochemical damage and inhibited the retinal cells apoptosis induced by fluorescent light (p < 0.05). Moreover, dietary BRACs inhibited expression of AP-1 (c-fos/c-jun subunits), up-regulated NF-κB (p65) expression and phosphorylation of IκB-α, and decreased Caspase-1 expression (p < 0.05). These results suggest that BRACs improve retinal damage produced by photochemical stress in rats via AP-1/NF-κB/Caspase-1 apoptotic mechanisms.
apoptosis; black rice anthocyanidins; Caspase-1; photochemical damage; retina
This study was conducted to assess the ability of two-dimensional tissue tracking (2DTT) to evaluate changes in left ventricular (LV) myocardial function associated with sustained high electrical pacing. Pacemakers were implanted at the right ventricular (RV) apex of five female Beagles, and sustained high electrical pacing of 250 beats per minute (bpm) was performed for three consecutive weeks. Conventional echocardiography and 2DTT were performed at baseline, and at every week for three weeks with pacing. The baseline parameters were then compared to those of weeks 1, 2, and 3. Three weeks of pacing resulted in significant reduction of radial and circumferential global strains (p < 0.001). Regional analysis revealed reduction of segmental strains in both radial and circumferential directions, as well as increased dyssynchrony after three weeks of pacing in the radial direction (p = 0.0007). The results of this study revealed the ability of 2DTT to measure radial and circumferential strains in dogs with sustained high-electrical pacing, and allowed assessment of global and regional myocardial function and the degree of dyssynchrony.
cardiology; dog; echocardiography; experimental animal model; myocardial failure
Our investigation was conducted in order to verify a recent severe epidemic at several swine farms in northern China that indicated a newly emerging disease. Evidence confirmed that the epidemic was caused by a virulent Pseudorabies virus infection in swine herds.
fatal infection; Northern China; pseudorabies virus; swine
Tendinitis of the superficial digital flexor tendon (SDFT) is a significant cause of lameness in horses; however, recent studies have shown that stem cells could be useful in veterinary regenerative medicine. Therefore, we isolated and characterized equine umbilical cord blood mesenchymal stem cells (eUCB-MSCs) from equine umbilical cord blood obtained from thoroughbred mares during the foaling period. Horses that had tendinitis of the SDFT were treated with eUCB-MSCs to confirm the therapeutic effect. After eUCB-MSCs transplantation, the core lesion in the SDFT was found to decrease. These results suggest that transplantation using eUCB-MSCs could be another source of cell treatment.
cell transplantation; equine; mesenchymal stem cells; umbilical cord blood
Despite the development of new technologies, new challenges still remain for large scale proteomic profiling when dealing with complex biological mixtures. Fractionation prior to liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis is usually the preferred method to reduce the complexity of any biological sample. In this study, a gel LC-MS/MS approach was used to explore the stage specific proteome of Cryptosporidium (C.) parvum. To accomplish this, the sporozoite protein of C. parvum was first fractionated using SDS-PAGE with subsequent LC-MS/MS analysis. A total of 135 protein hits were recorded from 20 gel slices (from same gel lane), with many hits occurring in more than one band. Excluding all non-Cryptosporidium entries and proteins with multiple hits, 33 separate C. parvum entries were identified during the study. The overall goal of this study was to reduce sample complexity by protein fractionation and increase the possibility of detecting proteins present in lower abundance in a complex protein mixture.
bioinformatics; Cryptosporidium; proteomics; SDS-PAGE; tandem mass spectrometry
The high genetic diversity of porcine reproductive and respiratory syndrome virus (PRRSV) has been an obstacle to developing an effective vaccine for porcine reproductive and respiratory syndrome (PRRS). This study was performed to assess the degree of genetic diversity among PRRSVs from Korean pig farms where wasting and respiratory syndrome was observed from 2005 to 2009. Samples from 786 farms were tested for the presence of PRRSV using reverse transcription PCR protocol. A total of 117 farms were positive for type 1 PRRSV while 198 farms were positive for type 2. Nucleotide sequences encoding the open reading frame (ORF) 5 were analyzed and compared to those of various published PRRSV isolates obtained worldwide. Sequence identity of the ORF 5 in the isolates was 81.6~100% for type 1 viruses and 81.4~100% for type 2 viruses. Phylogenetic analysis of the ORF 5 sequences showed that types 1 and 2 PRRSVs from Korea were mainly classified into three and four clusters, respectively. The analyzed isolates were distributed throughout the clusters independent of the isolation year or geographical origin. In conclusion, our results indicated that the genetic diversity of PRRSVs from Korean pig farms is high and has been increasing over time.
Korea; open reading frame 5; phylogenetic analysis; porcine reproductive and respiratory syndrome virus
The purpose of this study was to evaluate the effect of meloxicam (MEL) on selected immune parameters of bovine CD25highCD4+, CD25lowCD4+, and CD25-CD4+ cells. Peripheral blood mononuclear cells (PBMCs) collected from 12-month-old heifers were treated with MEL at a concentration corresponding to the serum level of this medication following administration at the recommended dose (MEL 5 × 10-6 M) and at a concentration 10 times lower (MEL 5 × 10-7 M). After 12 and 24 h of incubation with the drug, the percentage of CD25highCD4+ cells decreased; however, this disturbance was quickly reversed. Furthermore, the absolute number of CD25highCD4+ cells in the PBMC populations treated with MEL 5 × 10-6 M for 48 and 168 h was increased. Prolonged (168 h) exposure to the drug increased the percentage of Foxp3+ cells in the CD25highCD4+ cell subpopulation. The higher dose of MEL was found to significantly increase the percentage of IFN-γ+ cells among the CD25-CD4+ cells. These results indicated that MEL does not exert an immunosuppressive effect by depleting CD4+ cells and suppression of IFN-γ+ production by these cells. Furthermore, IL-10 and TGF-β production was not changed following exposure to MEL.
cattle; CD4+ cells; cytokines; Foxp3; meloxicam
Germanium biotite (GB) is an aluminosilicate mineral containing 36 ppm germanium. The present study was conducted to better understand the effects of GB on immune responses in a mouse model, and to demonstrate the clearance effects of this mineral against Porcine reproductive and respiratory syndrome virus (PRRSV) in experimentally infected pigs as an initial step towards the development of a feed supplement that would promote immune activity and help prevent diseases. In the mouse model, dietary supplementation with GB enhanced concanavalin A (ConA)-induced lymphocyte proliferation and increased the percentage of CD3+CD8+ T lymphocytes. In pigs experimentally infected with PRRSV, viral titers in lungs and lymphoid tissues from the GB-fed group were significantly decreased compared to those of the control group 12 days post-infection. Corresponding histopathological analyses demonstrated that GB-fed pigs displayed less severe pathological changes associated with PRRSV infection compared to the control group, indicating that GB promotes PRRSV clearance. These antiviral effects in pigs may be related to the ability of GB to increase CD3+CD8+ T lymphocyte production observed in the mice. Hence, this mineral may be an effective feed supplement for increasing immune activity and preventing disease.
germanium biotite; immune enhancement; porcine reproductive and respiratory syndrome virus
Monoclonal antibody (mAb, NVRQS-DON) against deoxynivalenol (DON) was prepared. DON-Ag coated enzyme linked immunosorbent assay (ELISA) and DON-Ab coated ELISA were prepared by coating the DON-BSA and DON mAb. Quantitative DON calculation ranged from 50 to 4,000 ng/mL for DON-Ab coated ELISA and from 25 to 500 ng/mL for DON-Ag coated ELISA. 50% of inhibitory concentration values of DON, HT-2, 15-acetyl-DON, and nivalenol were 23.44, 22,545, 5,518 and 5,976 ng/mL based on the DON-Ab coated ELISA. Cross-reactivity levels of the mAb to HT-2, 15-acetyl-DON, and nivalenol were 0.1, 0.42, and 0.40%. The intra- and interassay precision coefficient variation (CV) were both <10%. In the mAb-coated ELISA, mean DON recovery rates in animal feed (0 to 1,000 µg/kg) ranged from 68.34 to 95.49% (CV; 4.10 to 13.38%). DON in a buffer solution (250, 500 and 1,000 ng/mL) was isolated using 300 µg of NVRQS-DON and 3 mg of magnetic nanoparticles (MNPs). The mean recovery rates of DON using this mAb-MNP system were 75.2, 96.9, and 88.1% in a buffer solution spiked with DON (250, 500, and 1,000 ng/mL). Conclusively we developed competitive ELISAs for detecting DON in animal feed and created a new tool for DON extraction using mAb-coupled MNPs.
deoxynivalenol; ELISA; magnetic nanoparticles; monoclonal antibody
Recent studies have shown that mesenchymal stem cells (MSCs) are able to differentiate into multi-lineage cells such as adipocytes, chondroblasts, and osteoblasts. Amniotic membrane from whole placenta is a good source of stem cells in humans. This membrane can potentially be used for wound healing and corneal surface reconstruction. Moreover, it can be easily obtained after delivery and is usually discarded as classified waste. In the present study, we successfully isolated and characterized equine amniotic membrane-derived mesenchymal stem cells (eAM-MSCs) that were cultured and maintained in low glucose Dulbecco's modified Eagle's medium. The proliferation of eAM-MSCs was measured based on the cumulative population doubling level (CPDL). Immunophenotyping of eAM-MSCs by flow cytometry showed that the major population was of mesenchymal origin. To confirm differentiation potential, a multi-lineage differentiation assay was conducted. We found that under appropriate conditions, eAM-MSCs are capable of multi-lineage differentiation. Our results indicated that eAM-MSCs may be a good source of stem cells, making them potentially useful for veterinary regenerative medicine and cell-based therapy.
amnion; amniotic membrane; equine; isolation; mesenchymal stem cells
To compare an injectable progesterone (MAD-4) with an intravaginal device (IPD), and natural O17 with synthetic oestradiol (OB) in a synchronisation protocol, 51 cows were divided into four groups. Each group was treated with one of the two sources of progesterone and one of the two oestradiol formulations. Oestrus behaviour, follicle diameter, and pregnancy rates were evaluated. Oestrus behaviour (p = 0.902), numbers of cows in oestrus (p = 0.917), follicle diameter (p = 0.416), and pregnancy rates (p = 0.873) were similar among the four groups. More cows in the group treated with the IPD and OB scored > 200 oestrus behaviour points compared to the other groups (p = 0.038). A longer interval between the end of treatment and oestrus was observed among cows treated with MAD-4 than cows given the IPD (p = 0.030), but no differences were found between animals receiving the two oestradiol formulations (OB and O17). While the use of MAD-4 requires further testing, similar responses to natural oestradiol observed in the present study could allow the use of this formulation in reproductive protocols because it is not associated with the potential human health risks of OB.
dairy cows; oestrus behaviour; synchronisation