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Assisted reproductive technology (ART) has been widely developed over the decades. This advanced technology has shown efficacy in the conception and breeding of an animal. However, several issues such as polyspermy, low maturation rate, and low development rate in vitro remain unresolved. Fallopian tube derived cells are proposed to promote the maturation and development of oocyte. This study aims to characterize porcine (PFTSC) and bovine fallopian tube stem cell (BFTSC) while comparing allogeneic and xenogeneic paracrine effects on porcine oocyte. FTSC of Taiwan yellow cattle (B. indicus) and porcine (Landrace x Yorkshire dam x Duroc) were isolated and identified. Conditioned media (Medium 199 or PZM-3) from porcine and bovine was collected and added to porcine cells during in vitro maturation (IVM) and in vitro culture (IVC). Both PFTSC and BFTSC expressed little CD44, CD105, and CD4. Both cells were induced to transform into chondrocytes, very few cells gave rise to osteocytes and adipocytes. IVM test showed a significant elevation of maturation rate in both groups (Porcine: 66.5 ± 3.5% 55.9 ± 1.7%, p .05; Bovine: 68.9 ± 2.3% 55.9 ± 1.7%, p .05). IVC test demonstrated markedly reduction of blastocyst in both groups. In a diluted conditioned medium with different concentration, 25% and 50% PFTSC showed a decrease in blastocyst rate which is significantly different, but BFTSC demonstrated no significant difference. PFTSC and BFTSC possessed properties of stem cells. Conditioned media from both PFTSC and BFTSC could improve maturation rate but not blastocyst rate in vitro.

Objective: To use deep learning to predict the probability of triangular fibrocartilage complex (TFCC) injury in patients’ MRI scans. Methods: We retrospectively studied medical records over 11 years and 2 months (1 January 2009–29 February 2019), collecting 332 contrast-enhanced hand MRI scans showing TFCC injury (143 scans) or not (189 scans) from a general hospital. We employed two convolutional neural networks with the MRNet (Algorithm 1) and ResNet50 (Algorithm 2) framework for deep learning. Explainable artificial intelligence was used for heatmap analysis. We tested deep learning using an external dataset containing the MRI scans of 12 patients with TFCC injuries and 38 healthy subjects. Results: In the internal dataset, Algorithm 1 had an AUC of 0.809 (95% confidence interval—CI: 0.670–0.947) for TFCC injury detection as well as an accuracy, sensitivity, and specificity of 75.6% (95% CI: 0.613–0.858), 66.7% (95% CI: 0.438–0.837), and 81.5% (95% CI: 0.633–0.918), respectively, and an F1 score of 0.686. Algorithm 2 had an AUC of 0.871 (95% CI: 0.747–0.995) for TFCC injury detection and an accuracy, sensitivity, and specificity of 90.7% (95% CI: 0.787–0.962), 88.2% (95% CI: 0.664–0.966), and 92.3% (95% CI: 0.763–0.978), respectively, and an F1 score of 0.882. The accuracy, sensitivity, and specificity for radiologist 1 were 88.9, 94.4 and 85.2%, respectively, and for radiologist 2, they were 71.1, 100 and 51.9%, respectively. Conclusions: A modified MRNet framework enables the detection of TFCC injury and guides accurate diagnosis.

Background High humidity and temperature in Taiwan have significant effects on the reproductivity of Holstein cattle, resulting in the occurrence of bovine ovarian follicular cyst (OFC). Because of economic loss from OFC, manual rupture and hormone injection have been advocated for the management of OFC. However, these incomplete treatments increase hormone resistance in cattle. Mesenchymal stem cells (MSCs) derived from placental stem cells (PSCs) demonstrate potential properties for the treatment of several diseases via promoting angiogenesis and immune modulation. Aim To establish the possibility of cattle placental stem cells (CPSCs) as a treatment modality for OFC of cows in Taiwan. Methods The cows with OFC were divided into three groups: control (BC1 and BC2), hormone (H1 and H2), and CPSC (PS1 and PS2) treatment groups. In the hormone treatment group, the cows were given gonadotrophin-releasing hormone (GnRH)-prostaglandin-GnRH intramuscular injection with or without drainage of follicular fluid. In the CPSC treatment group, CPSCs were isolated from the placenta after labor. With the identification of surface antigen on stem cells, the cows were administered ovarian injection of 1 × 106 or 6 × 106 CPSCs with drainage. In all groups, OFC was scanned by ultrasound once a week for a total of seven times. The concentrations of estradiol and progesterone in serum were tested in the same period. The estrus cycle was analyzed by food intake and activity. If estrus was detected, artificial insemination was conducted. Then the cow was monitored by ultrasound for confirmation of pregnancy. Results After 7 d of culture, CPSCs were successfully isolated from placental pieces. CPSCs significantly proliferated every 24 h and had high expression of MSC markers such as cluster of differentiation 44, as determined by flow cytometry. Ultrasound showed lower numbers of OFCs with drainage of follicular fluid. We achieved recovery rates of 0%, 50%, 50%, 75%, 75% and 75% in BC1, BC2, H1, H2, PS1, and PS2, respectively. Higher concentrations of progesterone were detected in the CPSC treatment groups. However, both hormone and CPSC treatment groups had no significant difference in the concentration of estradiol. The estrus rate was 0%, 100%, 25%, 75%, 75% and 75% in BC1, BC2, H1, H2, PS1, and PS2, respectively. The two fetuses were born in H2 and PS1. In brief, cows with CPSC injection achieved higher recovery, estrus, and inseminated conception rates. Conclusion CPSCs have efficacy in treating cows with OFC, and thus, may serve as an alternative treatment for reproductive disorders.

Objective: The purpose of this study was to investigate whether goose growth and feather characteristics are influenced by their line and feeding surroundings, inclusive of floor materials and types, since there are no reports regarding these factors.Methods: The 240 White Roman geese which were hatched and sex identified came from 3 commercial goose farms. They were randomly distributed to 24 pens depending on a completely random design. The study continued for 13 weeks and included 3 lines of commercial geese and 2 floor types (cement strip floor [CSF] or cement floor [CF]).Results: The day one gosling weight from A farm was lower than other two farms (96 g vs 107 and 115 g; p

Objective Bovine mastitis results in economic loss due to decrease in milk production. Antibiotic ointments are commonly used for treating. However, residue and anti-microbial resistance warranted attention progressively. Fortunately, stem cell anti-inflammatory properties and paracrine expression of cytokines accelerates wound healing and suppresses inflammatory reactions in mastitis. The objective of this study is to use the conditioned-Dulbecco's pluripotent stem cells (DPBS) from amniotic membrane stem cells (AMSCs) in treating bovine mastitis. Materials and methods The cows with mastitis were divided into two groups. In antibiotic control group, the cows were given tetraneomycin ointment. In conditioned-DPBS of AMSCs treatment group, amniotic membrane was collected for AMSCs after delivery. With expression of surface antigen and potential of tri-linage differentiation, AMSCs were injected into mammary glands. Then, milk was sampled every three days to monitor the effect of both treatments. The quality of milk was measured with pH, titratable acidity, free calcium ions and somatic cell count. Results Our results demonstrated the Bovine AMSCs expressed CD44, low levels of CD4 and no CD105. Bovine AMSCs demonstrated the differentiation capability in the tri-cell lineages. Mastitis treatment with conditioned-DPBS from AMSCs (experimental group) and conventional antibiotics (control group) showed insignificant difference in pH value and titratable acidity. The level of ionic calcium concentration in the conditioned-DPBS group decreased from 3rd day to 12th day, while the level in the antibiotic group decreased from 0 day to 12th day. The somatic cell number was similar in both groups, which meet the standard of Taiwan milk collection. Conclusion In conclusion, conditioned-DPBS from bovine AMSCs has the therapeutic potential to treat bovine mastitis and may replace antibiotics therapy in the future.

Glucocorticoid (GC)-induced bone loss is the most prevalent form of secondary osteoporosis. Previous studies demonstrated that long-term incubation of dexamethasone (DEX) induced oxidative stress and mitochondrial dysfunctions, consequently leading to apoptosis of differentiated osteoblasts. This DEX-induced cell death might be the main causes of bone loss. We previously described that DEX induced biphasic mitochondrial alternations. As GC affects mitochondrial physiology through several different possible routes, the short-term and long-term effects of GC treatment on mitochondria in the osteoblast have not been carefully characterized. Here, we examined the expression levels of genes that are associated with mitochondrial functions at several different time points after incubation with DEX. Mitochondrial biogenesis-mediated genes nuclear respiratory factor 1 (Nrf1) and Nrf2 were upregulated after 4-h incubation, and then declined after 24-h incubation, suggesting that mitochondrial biogenesis were transiently upregulated by DEX. In contrast, mitochondrial fusion gene optic atrophy 1 (Opa1) and mitofusin 2 (Mfn2) started to be elevated as the biogenesis started to decrease. Finally, the mitochondrial fission increased and apoptosis becomes prominent. Agree with the mitochondrial biphasic alterations hypothesis, the results suggested an early increase of mitochondrial activities and biogenesis upon DEX stimulation to the osteoblasts. The oxidative phosphorylation and inducible nitric oxide synthase levels increased results in oxidative stress accumulation, leading to mitochondrial fusion, and subsequently fission and triggering the apoptosis. Our results indicated that the primary effects of GC on mitochondria are promoting their functions and biogenesis. Mitochondrial breakdown and the activation of the apoptotic pathways appeared to be the secondary effect after long-term treatment.

Cytoplasmic replacement by spindle transfer (ST) technique can be applied to improve the developmental competence of poor qualitied or aged oocytes. In cattle, ST technology has not been well established for producing embryos and the calves successfully using intracytoplasmic sperm injection (ICSI) or in vitro fertilization (IVF). The objective of this study was to develop a novel procedure for producing bovine ST embryos, which could be fundamental to applying ST technology in other mammals. In the present study, the efficacies of performing ICSI before (ICSI-ST) or after (ST-ICSI) and IVF on the development of ST bovine embryos were investigated. Results indicated that the blastocyst rate of ST embryos produced by ICSI-ST (24.7%) was higher (P 0.05) than that produced by ST-ICSI (5.9%). On the other hand, ST-IVF had the highest fertilization rate (97.3%), polyspermy rate (24.7%), and lowest blastocyst rate (22.7%) when compared to denuded oocytes (DO), zona cut oocytes (ZC), and cumulus-oocyte complexes (COCs)-IVF groups. Finally, the in vitro development rates of ICSI-ST (24.5%) and ST-IVF (25.2%) were not significantly different (P 0.05). However, the pregnancy rate (46.7%) and birth rate (33.3%) of ST-IVF were higher (P 0.05) than those of ICSI-ST (6.3% and 0%, respectively). The percentage of mitochondrial DNA (mtDNA) heteroplasmy derived from donor karyoplasts of the 5 claves was between 2% and 18%. Taken together, performing ICSI prior to ST can improve the embryonic development of ST bovine embryos. Moreover, using IVF, instead of ICSI, for ST oocyte fertilization dramatically increased the pregnancy rate and birth rate of ST calves, in which mtDNA heteroplasmy derived from donor karyoplasts exists.

Spinal muscular atrophy (SMA) is a single gene disorder affecting motor function in uterus. Amniotic fluid is an alternative source of stem cell to ameliorate SMA. Therefore, this study aims to examine the therapeutic potential of Human amniotic fluid stem cell (hAFSC) for SMA. Our SMA model mice were generated by deletion of exon 7 of Smn gene and knock-in of human SMN2. A total of 16 SMA model mice were injected with 1 × 105 hAFSC in uterus, and the other 16 mice served as the negative control. Motor function was analyzed by three behavioral tests. Engraftment of hAFSC in organs were assessed by flow cytometry and RNA scope. Frequency of myocytes, neurons and innervated receptors were estimated by staining. With hAFSC transplantation, 15 fetuses survived (93.75% survival) and showed better performance in all motor function tests. Higher engraftment frequency were observed in muscle and liver. Besides, the muscle with hAFSC transplantation expressed much laminin α and PAX-7. Significantly higher frequency of myocytes, neurons and innervated receptors were observed. In our study, hAFSC engrafted on neuromuscular organs and improved cellular and behavioral outcomes of SMA model mice. This fetal therapy could preserve the time window and treat in the uterus.

Simple Summary Obesity has become a global pandemic involving an increase in the amount and size of fat cells in the body. Obesity is highly associated with insulin resistance and type 2 diabetes. Several medicinal plants have been reported to be useful in the prevention of obesity. Forskolin, a bioactive compound of Coleus forskohlii, can promote lipolysis in mature adipocytes. However, the effect of forskolin on body weight, glucose metabolism and adipocyte size of diet-induced obesity is still rarely investigated. In this study, the effects of forskolin on the high-fat diet-induced obese model were evaluated. Results showed that forskolin administration improves glucose metabolism and reduces fat cell diameter in the high-fat diet-fed mice. Forskolin also suppresses adipocyte differentiation of murine mesenchymal stem cells. Abstract The purpose of this study was to investigate the effects of forskolin on body weight, glucose metabolism and fat cell diameter in high-fat diet-induced obese mice. Four-week-old male mice (C57BL/6) were randomly assigned to 1 of 3 treatment groups: a high-fat diet plus 5% dimethyl sulfoxide (vehicle), high-fat diet plus 2 mg/kg of forskolin (dissolved in 5% dimethyl sulfoxide) and high-fat diet plus 4 mg/kg of forskolin (dissolved in 5% dimethyl sulfoxide). Forskolin or dimethyl sulfoxide was administered intraperitoneally every two days. The results indicated that no significant difference was observed in the body weight, feed intake and serum lipid parameters among groups at 20 weeks of age. The blood glucose levels were significantly reduced in the groups treated with 2 mg/kg of forskolin before glucose tolerance test. Forskolin administration linearly decreased blood glucose levels of high-fat diet-fed mice at 90 min and total area under curve (AUC) after insulin tolerance test. The subcutaneous adipocyte diameter was significantly reduced in the groups treated with 2 mg/kg of forskolin. Forskolin administration linearly reduced the gonadal adipocyte diameter of high-fat diet-fed mice. Forskolin significantly reduced the differentiation of murine mesenchymal stem cells into adipocytes and this was accompanied by a decrease in intracellular triglyceride content and an increase in glycerol concentration in the culture medium. The subcutaneous adipocyte diameter, gonadal adipocyte diameter and total AUC of insulin tolerance test were moderately negatively correlated with the concentration of forskolin in the high-fat diet-induced obese model. These results demonstrate that forskolin can regulate glucose metabolism and reduce fat cell diameter of high-fat diet-fed mice and inhibit the adipocyte differentiation of murine mesenchymal stem cells.

BackgroundType III spinal muscular atrophy (SMA) is a single gene disorder affecting motor function in uterus. Several types of stem cells were utilized to ameliorate SMA based on its capability of regeneration and differentiation. Amniotic fluid is an alternative source of stem cells and is safely sampled without ethical issues. Human amniotic fluid stem cell (hAFSC) shared common surface markers of mesenchymal stem cell. Therefore, this study aims to examine the therapeutic potential of hAFSC for SMA. MethodsOur SMA model mice were generated by deletion of exon 7 of Smn gene and knock-in of human SMN2. A total of 16 SMA model mice were injected with 1x105 hAFSC in uterus, and the other 16 mice served as the negative control. Motor function was analyzed by Rotarod maintenance test, tilting test and grasping test every two months. Twelve months after transplantation, all organs were extracted for post-mortem analysis. Engraftment of hAFSC in organs were assessed by flow cytometry and RNA scope. To observe the function of neuromuscular junction, frequency of myocytes, neurons and innervated receptors were estimated by H&E, methylene blue and immunocytochemistry staining. ResultsWith hAFSC transplantation, 15 fetuses from 5 dams survived (15 of 16, 93.75% survival) and showed better performance in all three motor function tests. Higher engraftment frequency in organs were observed in muscle and liver after hAFSC transplantation. Besides, the muscle of SMA mice with hAFSC transplantation expressed much laminin α and PAX-7. Significantly higher frequency of myocytes, neurons and innervated receptors were observed after hAFSC transplantation. ConclusionsIn our study, hAFSC engrafted on neuromuscular organs and improved cellular and behavioral outcomes of SMA model mice. This fetal therapy could preserve the time window and treat in the uterus to avoid irreversible damage.

Background Goslings in several Taiwanese farms experienced gosling feather loss disease (GFL) at 21-35 days and goose broke feather disease (GBF) at 42-60 days. The prevalence ranges from a few birds to 500 cases per field. It is estimated that about 12,000 geese have been infected, the morbidity is 70-80% and the mortality is 20-30%. Objectives This study aims to investigate the pathogens that cause GFL and GBF. Focus on the study of the correlation between goose circovirus (GoCV) and goose parvovirus (GPV) with the goose feather loss in southern Taiwan. Furthermore, a phylogenetic tree was established to align the differences between southern and northern Taiwan and compare with virus strains from China and Europe. Methods Samples were collected from animal hospitals. Molecular and microscopy diagnostics were used to examine 92 geese. Specific quantitative polymerase chain reaction (Q-PCR) assays are performed to evaluate GPV and GoCV viral loads and simultaneously evaluated the feather loss conditions in geese with the scoring method. Results High prevalence of GoCV and GPV infection in geese showing signs of GFL and GBF. Inclusion body was detected in the feather follicles and Lieberkuhn crypt epithelial cells. The Q-PCR showed the high correlation between feather loss and viruses during 3rd-5th week. However, the infection was not detected using the same test in 60 healthy geese. Conclusions Thus, GFL and GBF appear to be significantly closely related to GoCV and GPV. The geese feathers showed increasing recovery after being quarantined and disinfected.

Yogurt as a fermented dairy product has been providing several benefits for human health and appropriate for people with lactose intolerance. Recently, the consumption level of yogurt has significantly increased. Starch can be used as an alternative hydrocolloid in the manufacturing process of yogurt. Indonesian Canna edulis Ker contains 63.27% amylopectin that has the potential used as an alternative hydrocolloid. Amylopectin has a very high capability of the water holding capacity to increase the viscosity and maintain the stability of yogurt drink. The present research was to determine the additional effects of Canna starch as an alternative hydrocolloid on the physicochemical and sensory properties of the yogurt drink during storage. Concentrations of 0.1%, 0.2%, 0.3%, and 0.4% canna starch and 0.2% CMC (control) were added to make yogurt. All treatments were carried out with four replications. Samples were refrigerated at 4oC for 24 hours and then analyzed for pH, titratable acidity, viscosity, syneresis, sedimentable fraction, and sensory properties. Data were analyzed by one way ANOVA, and followed by Duncan's multiple range test (DMRT). Data from the pH, titratable acidity, syneresis, sedimentable fraction, viscosity and sensory analyses obtained from the present study indicated that concentration of 0.1–0.4% (w/v) Canna starch could be applicable in the manufacture of the yogurt drink. Furthermore, it was found that 0.1% (w/v) of Canna starch selected as the best concentration could be used in yogurt manufacture process that resulted in similar sensory quality compared with CMC as commercial hydrocolloid.

Amniotic fluid is a rich source of multipotent mesenchymal stem cells (MSCs). Amniotic fluid stem cells (AFSCs) have become a new source of stem cells; they have low immunogenicity and are easily harvested. For this reason, they may be useful in clinical tissue engineering. Moreover, AFSCs have anti-inflammatory properties and can repair tissues. This study evaluated the utility of AFSC injection to treat bilateral ovarian dystrophy in Holstein-Friesian cows. Bovine AFSCs (BAFSCs) were collected at slaughter from Holstein-Friesian cows during the third or fourth month of pregnancy and cultured in vitro. The BAFSCs began to show a fibroblast-like morphology. They were positive for β-integrin, CD44, CD73, CD106 and Oct4 and negative for CD34 and CD45. After induction, the cells differentiated into mesodermal lineages. Bilateral ovarian dystrophy was confirmed by ultrasonography in 16 lactating cows. The subsequent experiment lasted 15 weeks. Serum was collected weekly to analyse progesterone concentrations, and weekly ultrasonography recorded ovarian changes. Each cow was equipped with an automatic heat detection system to facilitate oestrus observation and breeding records. The progesterone concentration of two cows in the treatment group (25%) significantly increased during weeks 10-15. On ultrasonography, the treatment group demonstrated mature follicles after BAFSCs injection, and foetuses were visualized approximately 40 days after artificial insemination (AI). Oestrus rates in the control and treatment groups were 0% (0/8) and 50% (4/8), respectively; pregnancy rates were 0% (0/8) and 25% (2/8), respectively. Calves were successfully delivered in both cases of pregnancy. These results show that BAFSCs can alleviate bovine ovarian dystrophy and restore fertility.

We previously demonstrated that the cellular thermotolerance of cloned cattle produced by combination of ooplasm (o) derived from Taiwan native yellow cattle (Y) and the donor (d) nucleus derived from Holstein (H) cattle (Yo-Hd) transmits to their offspring (Yo-Hd-F1). In the present study, the responses of mitochondria in these cloned cattle and their offspring after heat shock were investigated to elucidate influence of cytoplasmic input (i.e., ooplasm) during the course of heat stress. After heat shock, oxidative phosphorylation proteins (Complex III and IV) of ear fibroblast cells with Y-originated cytoplasm (including Y, Yo-Hd, and Yo-Hd-F1 cattle) were significantly greater (P 0.05) than those of ear fibroblast cells with H-originated cytoplasm (including H, Ho-Hd, and Ho-Hd-F1 cattle). However, the expressions of Complex I and Complex II protein in heat shocked cells derived from Yo-Hd-F1 cattle were significantly (P 0.05) higher than those of cell derived from cattle with the H-cytoplasm. The catalase (CAT) expression in heat shocked ear fibroblast cells derived from Yo-Hd and Yo-Hd-F1 cattle were significantly (P 0.05) higher than that of cells derived from Ho-Hd-F1 cattle. However, the level of glutathione peroxidase (GPx) expression was higher (P 0.05) in ear fibroblast cells with Y-originated cytoplasm compared to cells with H-originated cytoplasm. In conclusion, the expression of proteins involved in mitochondrial oxidative phosphorylation and antioxidant enzymes after heat shock was increased in ear fibroblast cells from cattle with Y-originated cytoplasm, which can be transmitted to their offspring.

Objective Type 1 diabetes is an autoimmune disease that destroys islet cells and results in insufficient insulin secretion by pancreatic β-cells. Islet transplantation from donors is an approach used for treating patients with diabetes; however, this therapy is difficult to implement because of the lack of donors. Nevertheless, several stem cells have the potential to differentiate from islet-like cells and enable insulin secretion for treating diabetes in animal models. For example, placenta is considered a waste material and can be harvested noninvasively during delivery without ethical or moral concerns. To date, the differentiation of islet-like cells from cow-derived placental stem cells (CPSCs) has yet to be demonstrated. Materials and methods The investigation of potential differentiation of islet-like cells from CPSCs was conducted by supplementation with nicotinamide, exendin-4, glucose, and poly- d -lysine and was detected through reverse transcription polymerase chain reaction, dithizone staining, and immunocytochemical methods. Results Our results indicated that CPSCs are established and express mesenchymal stem cell surface antigen markers, such as CD73, CD166, β-integrin, and Oct-4, but not hematopoietic stem cell surface antigen markers, such as CD45. After induction, the CPSCs successfully differentiated into islet-like cells. The CPSC-derived islet-like cells expressed islet cell development-related genes, such as insulin, glucagon, pax-4, Nkx6.1, pax-6, and Fox. Moreover, CPSC-derived islet-like cells can be stained with zinc ions, which are widely distributed in the islet cells and enable insulin secretion. Conclusion Altogether, islet-like cells have the potential to be differentiated from CPSCs without gene manipulation, and can be used in diabetic animal models in the future for preclinical and drug testing trial investigations.

Our aim was to investigate insemination techniques in order to improve pregnancy rates of artificial insemination (AI) using sex-sorted semen (sexed AI) in cattle in tropical and subtropical (T/ST) regions. In T/ST regions, the pregnancy rates by sexed AI are reportedly the lowest in the hottest months of the year, with less than 15% in cows and 35–40% in heifers (PMID 24048822). We compared sexed AI by depositing the semen into the uterine body (UB-AI, n = 12) versus the unilateral uterine horn (UUH-AI, n = 14) of pre-ovulation heifers. The ovary and follicle were assessed by rectal ultrasound before AI. After insemination, pregnancy was determined by ultrasound at approximately 40 days and approximately 70 days. In the present study, we demonstrated that high pregnancy rates (>70%) by sexed AI in the hottest season in a subtropical region such as Taiwan can be achieved when heifers with pre-ovulation follicles are used. The overall pregnancy rates were 54% higher in the UUH-AI (71%) group than in the UB-AI (42%) group (P = 0.06), examined on approximately 40 days post-sexed AI. Surprisingly, however, the pregnancy outcome appeared to be higher in the hot season (62%) than in the cool season (46%) although this difference was not statistically significant. Based on the present study, we recommend that cattle breeders perform UUH-AI using sex-sorted semen for heifers with pre-ovulation follicles in order to achieve satisfactory pregnancy outcome in the hot seasons in T/ST regions.

The success of freezing chicken semen is desired for preserving Taiwan native chickens. Semen samples from three Taiwan roosters were collected using massage technique twice a week. Two Cryoprotectant Maltose concentrations, 0.6 and 1.2 g/ml, were added in Tris Aminomethane+20 % egg yolk as the basic semen extender. The purpose of this study was to observe the effect of Tris Aminomethane extender supplemented with maltose on post thawing quality (mass spermatozoa motility, spermatozoa motility, spermatozoa abnormality, and viability) following frozen storage on Taiwan Native Rooster spermatozoa. The experiment was conducted in Animal Embriology Engineering Laboratory of The National Pingtung University of Science and Technology, Taiwan. During the period of one month experiment, feed and water were provided ad libitum for the roosters. All semen samples were diluted (semen:diluent=1:10) with warmed diluents at 30ºC in a water bath. The diluted semen samples were gradually cooled to 4ºC for 4h. After cooling, spermatozoa motility was examined. The samples were packed in 0.25 ml straws and plugged with the straw powder. The straws were kept at 4ºC until frozen, which was carried out by placing the samples in liquid nitrogen vapor (-125 to -130ºC) for 20 minutes, and stored in liquid nitrogen (-196ºC) until examination after 3 hours. spermatozoa motility, abnormality and viability showed significant differences (P < 0.05) when preserved with cryoprotectant when compared to the control treatment. It can be concluded that maltose is a potential protector for improving spermatozoa quality following frozen storage.

We have previously demonstrated that the somatic cells from cattle cloned with Holstein (H) donor cells and Taiwan native yellow cattle (Y) ooplasm (Yo-Hd) had better thermotolerance than those from cattle cloned with both Holstein donor cells and ooplasm (Ho-Hd). The present study aimed to investigate whether the cellular thermotolerance of these cloned cattle is transmissible to their offspring (Ho-Hd-F1 and Yo-Hd-F1). Thermotolerance of ear fibroblasts derived from these cloned cattle and their offspring were analyzed. Polymorphisms in mitochondrial DNA (mtDNA) D-loop of ear fibroblasts derived from Yo-Hd and Yo-Hd-F1 indicated that the cytoplasm is originated from Bos indicus (Y). After heat shock, the apoptotic rates, B-cell lymphoma 2-associated X protein/B-cell lymphoma 2 ratios, and relative expression levels of cysteine-aspartic proteases (caspases)-3, -8, and -9 of ear fibroblasts with Y-originated cytoplasm (including Y, Yo-Hd, and Yo-Hd-F1) were lower (P

The use of livestock waste for the production of biogas and the application of biogas slurry to agricultural soil can resolve livestock waste problems and reduce synthetic fertiliser use. However, the migration of veterinary drugs to land and crops resulting from biogas slurry irrigation is a potential food safety concern. This study employed an ultra-performance liquid chromatography coupled with quadrupole-time of flight high-resolution mass spectrometry system for wide-scope suspect screening of pharmaceutically active substances on crop cultivated under biogas slurry irrigation. Briefly, a total of 22 pak choi samples were obtained from a greenhouse farmed in tropical south Taiwan between March 2019 and March 2020. Molecular spectra and fragmented ions (between m/z 70 and 1100) were acquired. Ion features were searched and matched with a library consisting of 1068 compounds. The matrixes in the crop production environment including soil, livestock wastewater, biogas slurry, and groundwater were included in this study to elucidate potential sources of the pharmaceutically active substances. Results demonstrated 23 suspects were matched with high mass accuracy (mass error within ±5.0 ppm) in pak choi. The detection of both bufexamac and nandrolone were confirmed using standards, where a new system of identification points was applied. Nandrolone was detected throughout the pak choi samples as well as livestock wastewater. Tetracycline, macrolide, and sulfonamide antibiotics were presented in biogas slurry and soil but not pak choi. This is the first study to reveal the presence of multiclass pharmaceutically active substances in a crop supplied as food. Such findings suggest that anabolics and antibiotics should be closely monitored in the corps irrigated by biogas slurry in future.

Side-channel attacks (SCA) are capable of bypassing the theoretical protection of cryptographic schemes and revealing secret information in hardware devices. They have become a serious threat to the hardware security of the cryptographic modules. Entering the Internet of Things (IoT) era, it is crucial to verify the IoT devices' ability to thwart potential attacks from stealing confidential data. In this work, we set out to design and implement a framework for efficient SCA resistance verification. By integrating trace measurement equipment and GPU-accelerated analyzing engines, we have successfully built an automatic system which greatly simplifies the test procedure and enables on-the-fly SCA tests. Experiment results on common IoT development boards show both promising strength and flexibility of our framework to meet different testing requirements smoothly.

To review the current medical and stem-cell therapy for spinal muscular atrophy (SMA) and prenatal transplantation of amniotic fluid stem cells in the future.SMA is an autosomal recessive inheritance of neurodegenerative disease, which is caused of the mutation in survival motor neuron. The severe-type SMA patients usually die from the respiratory failure within 2 years after birth. Recently, researchers had found that 3-methyladenine could inhibit the autophagy and had the capacity to reduce death of the neurons. The first food and drug administration-approved drug to treat SMA, Nusinersen, is a modified antisense oligonucleotide to target intronic splicing silencer N1 just recently launched. Not only medical therapy, but also stem cells including neural stem cells, embryonic stem cells, mesenchymal stem cells, and induced pluripotent stem cells could show the potential to repair the injured tissue and differentiate into neuron cells to rescue the SMA animal models. Human amniotic fluid stem cells (HAFSCs) share the potential of mesenchymal stem cells and could differentiate into tri-lineage-relative cells, which are also having the ability to restore the injured neuro-muscular function. In this review, we further demonstrate the therapeutic effect of using HAFSCs to treat type III SMA prenatally. HAFSCs, similar to other stem cells, could also help the improvement of SMA with even longer survival.The concept of prenatal stem-cell therapy preserves the time window to treat disease in utero with much less cell number. Stem cell alone might not be enough to correct or cure the SMA but could be applied as the additional therapy combined with antisense oligonucleotide in the future.

Background/Purpose Amniotic fluid-derived progenitor cells (AFPCs) are oligopotent and shed from the fetus into the amniotic fluid. It was reported that AFPCs express stem cell-like markers and are capable of differentiating into specific cell type in in vitro experiments. However, no study has fully investigated the potentiality and destiny of these cells in in vivo experiments. Methods Ds-red transgenic mice (on Day 13.5 of pregnancy) were transplanted in utero with enhanced green fluorescent protein-labeled mouse AFPC (EGFP-mAFPCs). After birth, baby mice were euthanized at 3-week intervals beginning 3 weeks postnatally, and the specimens were examined by polymerase chain reaction, histology, and flow cytometry. Results Our results demonstrate the transplantability of mAFPCs into all three germ layers and the potential of mAFPCs in the study of progenitor cell homing, differentiation, and function. Engraftment of EGFP-mAFPCs was detected in the intestine, kidney, muscle, skin, bladder, heart, stomach, etc., at 3 weeks after delivery. Conclusion This model using EGFP-mAFPCs injected in utero may provide an ideal method for determining the fate of transplanted cells in recipients and these findings may justify a clinical trial of in utero transplantation during gestation for patients who have inherited genetic disorders.

Acute myocardial infarction (MI) is a fatal event that causes a large number of deaths worldwide. MI results in pathological remodeling and decreased cardiac function, which could lead to heart failure and fatal arrhythmia. Cell therapy is a potential strategy to repair the damage through enhanced angiogenesis or by modulation of the inflammatory process via paracrine signaling. Amniotic fluid-derived progenitor cells (AFPCs) have been reported to differentiate into several lineages and can be used without ethical concerns or risk of teratoma formation. Since pigs are anatomically, physiologically, and genetically similar to humans, and pregnant pigs can be an abundant source of AFPCs, we used porcine AFPCs (pAFPCs) as our target cells. Intramyocardial injection of AFPCs has been shown to cure MI in animal models. However, intramuscular transplantation of cells has not been extensively investigated. In this study, we investigated the therapeutic potential of intramuscular injection of pAFPCs on acute MI. MI mice were divided into 1) PBS control, 2) medium cell dose (1 × 106 cells per leg; cell-M), and 3) high cell dose (4 × 106 cells per leg; cell-H) groups. Cells or PBS were directly injected into the hamstring muscle 20 min after MI surgery. Four weeks after MI surgery, the cell-M and cell-H groups exhibited significantly better ejection fraction, significantly greater wall thickness, smaller infarct scar sizes, and lower LV expansion index compared to the PBS group. Using in vivo imaging, we showed that the hamstring muscles from animals in the cell-M and cell-H groups had RFP-positive signals. In summary, intramuscular injection of porcine AFPCs reduced scar size, reduced pathological remodeling, and preserved heart function after MI.

Recently, stem cells have offered an alternative treatment for inflammatory bowel disease or colitis to overcome the poor outcomes associated with current therapies. Amniotic fluid-derived stem cells have the potential for the regeneration of impaired organs and the recovery of normal physiologic functions of damaged tissues without ethical concerns or risk of tumor formation. In this work, we aimed to examine the therapeutic effects of infusion of porcine amniotic fluid-derived stem cells (pAFSCs) in dextran sulfate sodium (DSS)-induced colitis in mice. Treatment with pAFSCs was shown to inhibit the shortening of the colon after induction of colitis and dramatically ameliorated the body weight-loss induced by the DSS treatment. In addition, pAFSCs could also reduce the extent of the inflamed area represented by epithelial mesenchymal transformation in the colitis mice. The levels of the inflammatory cytokines interleukin 6 and interferon-γ were also reduced in colitis mice transplanted with pAFSCs. In conclusion, pAFSCs can ameliorate experimental colitis in mice, suggesting that they may be a potential treatment for inflammatory bowel disease or colitis.

Milk, one of the important human foods, can not only be drunken directly but also be processed to produce more products. It is a critical issue for owners of ranches to effectively manage the pregnancy, reproduction, and milking of dairy cows. In current ranches, artificial insemination is performed to make dairy cows pregnant. Accurate estrus detection of dairy cows is essential for the success in artificial insemination. The traditional method to detect estrus of dairy cows is visual observation, whose success rate largely depends on the observers' experience and the frequency of observation. Therefore, several direct-contact monitor systems have been developed. But these sensors which directly contact the skin of dairy cows induce their discomfort. In this study, a non-contact video monitor system will be developed to avoid the contact-related discomfort. Infrared cameras will be used to record the imaging of dairy cows at night, when the frequency of estrus is the highest. Then the dairy cows with estrus-specific behaviors will be identified by imaging processing. Finally only simple judgments by owners are required to decide whether artificial insemination will be performed.

Objectives Amniotic fluid stem cells (AFSCs) are derived from the amniotic fluid of the developing fetus and can give rise to diverse differentiated cells of ectoderm, mesoderm, and endoderm lineages. Intrauterine transplantation is an approach used to cure inherited genetic fetal defects during the gestation period of pregnant dams. Certain disease such as osteogenesis imperfecta was successfully treated in affected fetal mice using this method. However, the donor cell destiny remains uncertain. Methods The purpose of this study was to investigate the biodistribution and cell fate of Ds-red-harboring porcine AFSCs (Ds-red pAFSCs) after intrauterine transplantation into enhanced green fluorescent protein-harboring fetuses of pregnant mice. Pregnant mice (12.5 days) underwent open laparotomy with intrauterine pAFSC transplantation (5 × 104 cells per pup) into fetal peritoneal cavity. Results Three weeks after birth, the mice were sacrificed. Several samples from different organs were obtained for histological examination and flow cytometric analysis. Ds-red pAFSCs migrated most frequently into the intestines. Furthermore, enhanced green fluorescent protein and red fluorescent protein signals were co-expressed in the intestine and liver cells via immunohistochemistry studies. Conclusion In utero xenotransplantation of pAFSCs fused with recipient intestinal cells instead of differentiating or maintaining the undifferentiated status in the tissue. © 2014 John Wiley & Sons, Ltd.

The decreasing of yogurt drink quality will occur during storage. Stabilizer that usually used for maintaining the yogurt quality is a chemical compound namely carboxymethyl cellulose (CMC). Canna (Canna edulis Ker) potentially used as a natural stabilizer, which is a local Indonesian tuber with high starch content and antioxidative properties. The purpose of this study is to investigate the capability of Canna starch to replace the used of CMC based on the physicochemical and antioxidant activity of yogurt drink during cold storage. The complete randomized factorial design will be used with two factor. The first factor was Canna starch/CMC levels divided into 5 groups, T0 (0.2%CMC) as a control, T1 (0.15% CMC + 0.025% canna), T2 (0.1% CMC + 0.05% canna), T3 (0.05% CMC + 0.075% canna), and T4 (0.1% Canna), respectively. The second factor was the storage time of yogurt drink divided into four groups, 1, 7, 14 and 21 days, to determine the quality change during stored in the refrigerator at 4°C. Data obtained from the results of subsequent studies analyzed by GLM (General Linear Model) and followed by Duncan's multiple range test (DMRT). This study resulted that Canna starch has the capability as a natural stabilizer for producing functional yogurt drink with potential health benefits related to the high antioxidant activity. The combination of 0.1% CMC and 0.05% Canna starch addition on yogurt drink manufacture showed the best physicochemical quality

Background Angiotensin II (AngII) reportedly enhances stem cell proliferation, and type 1 angiotensin II receptor (AT1R) antagonists reduce angiogenesis in a rodent hindlimb ischemic model. Whether AT1R antagonists can alter the angiogenic activity of bone-marrow mesenchymal stem cells (BMSCs) is unknown. The purpose of this study is to investigate whether AT1R antagonists can alter the angiogenic activity of BMSCs and explore the potential mechanism for such an action. Methods Mouse BMSCs were isolated and treated with AngII, an AT1R antagonist, and a type 2 angiotensin II receptor (AT2R) antagonist. Angiogenic activity of BMSCs was detected by vascular endothelial growth factor (VEGF) secretion and tube formation of human umbilical vein endothelial cells (HUVECs). BMSCs isolated from enhanced green fluorescent protein (eGFP)-transgenic mice were allografted into ischemic hindlimbs in mice. Results The BMSCs constitutively expressed AT1Rs and AT2Rs. AngII treatment significantly increased VEGF secretion by BMSCs. Valsartan (AT1R antagonist) but not PD123319 (AT2R antagonist) treatment attenuated the AngII-induced promotion of VEGF synthesis by BMSCs. The AngII and AngII receptor antagonist control of angiogenic activity of BMSCs were confirmed by tube formation of HUVECs. AngII treatment promoted phosphorylated Ser473 Akt abundance in BMSCs. RNA interference of an isoform of AT1R, valsartan, and wortmannin treatments attenuated AngII-induced Akt phosphorylation. Allograft of BMSCs significantly increased blood flow and VEGF expression in the gastrocnemius muscles of ischemic hindlimbs, which was attenuated after valsartan treatment. Conclusions AT1R antagonists, via AT-1R/PI3K/Akt pathways, impair the AngII-induced promotion of angiogenic activity of mouse BMSCs.

The objective of this study was to compare the thermotolerances of ear fibroblasts derived from Holstein (H) and Taiwan yellow cattle (Y) and their apoptosis-related protein expressions with (1, 3, 6, 12, and 24h) or without heat shock treatment. The results showed that the vaginal temperatures of Y (38.4-38.5°C) were (P0.05) lower than that of H (38.8°C) during the hot season. The apoptotic rates of ear fibroblasts derived from Y (6h: 1.1%; 12h: 1.6%; 24h: 2.6%) were lower (P0.05) than those of cells derived from H (6h: 1.8%; 12h: 4.0%; 24h: 6.9%), respectively, after heat shock (42°C). The expression level of apoptosis inducing factor (AIF) in ear fibroblasts derived from H was higher (P0.05) than those derived from Y after the heat shock treatment for 6h and 12h, respectively. The level of cytochrome c of ear fibroblasts derived from H was higher (P0.05) than those derived from Y after the heat shock treatment for 1-12h, respectively. The abundances of Caspase-3, Caspase-8 and Caspase-9 of ear fibroblasts derived from H were higher (P0.05) than those of cells derived from Y after 12h and 24h of heat shock, respectively; the Bcl-2/Bax ratios of ear fibroblasts derived from H were lower (P0.05) than those from Y-derived fibroblasts after heated for 1-24h. The expression level of HSP-70 of Y-derived ear fibroblasts was also higher (P0.05) than that from H after the same duration of heat shock treatments. Taken together, the thermotolerance of ear fibroblasts derived from Taiwan yellow cattle was better than that of cells derived from Holstein cattle.

Generally, the grating-lobe artifacts can be avoided with array pitch smaller than one half of wavelength; however, due to cost or physical limitations, such a condition is hard to be satisfied in multi-dimension or high frequency array transducers. Besides grating lobes, side-lobe signals also affect imaging contrast and quality. To suppress the side-lobes, the window method such as Tukey window and Hamming window is widely used. To suppress grating lobes and side-lobes associated with linear arrays, we propose a novel linear array beamformation method based on a concept of virtual sub-wavelength receiving elements. Each array element is virtually divided into multiple sub-elements with sub-wavelength pitch. The signal detected by each virtual sub-element contributes to that of its physical element at different arrival time. Therefore, the signals of the virtual sub-elements from the same physical elements are approximated using the same signal received by their physical element. Delay-and-sum beamformation on receive is then done with the approximated signals and corresponding delays of the virtual sub-elements instead of using those of physical elements. Such a beamformation synthesizes a non-uniform sampled receive aperture with its mean effective array pitch smaller than one wavelength; thus the grating lobes can be suppressed. The sub-wavelength mean effective array pitch implies that better receiving delay estimation, because of smaller effective receive elements, can be achieved, which plays the role of side-lobe suppression. Wire phantom experiments using a research ultrasound engine were performed to verify our idea. Our method provided at least 5-dB suppression of the grating lobe and sidelobes, and rivaled the window method. Our method works for coherent plane wave compounding as well. Optimal sub-wavelength dividing of each physical element will also be presented.

DsRed-monomer is an enhanced red fluorescent protein that may serve as a marker for studies in biotechnology and cell biology. Since the ICR mouse strain is a widely utilized outbred strain for oncology, toxicology, vaccine development and for aging studies, the objective of this study was to produce a DsRed-monomer transgenic mouse by means of pronuclear micro-injection of a vector driven by the cytomegalovirus (CMV) enhancer/chicken beta-actin promoter. Four transgenic mice were successfully produced, one of which expressed the DsRed-monomer protein in every tissue, although at varying levels. High expression levels were observed in the heart, pancreas and muscle. Moreover, amniotic fluid-derived progenitor cells, which also expressed the DsRed-monomer protein, could be collected from the DsRed-monomer- harboring ICR mice. As compared to wild-type mice, a few biochemical and histological dissimilarities were found in the DsRed-monomer transgenic mice, including the presence of intra-cytoplasmic eosinophilic threadlike materials in the acinar cells. Taken together, transgenic mice stably expressing DsRed-monomer can be produced using pronuclear micro-injection; however, expression of the DsRed-monomer gene or its insertion position may lead to minor influences.

Objective Liver fibrosis results from the wound healing response to chronic liver damage. Advanced liver fibrosis results in cirrhosis and liver failure, and liver transplantation is often the only option for effective therapy; however, the shortage of available donor livers limits this treatment. Thus, new therapies for advanced liver fibrosis are essential. Materials and methods Amniotic fluid contains an abundance of stem cells, which are derived from all three germ layers of the developing fetus. These cells do not induce teratomas in vivo and do not pose any ethical concerns. To generate liver fibrosis models, male ICR mice were treated with CCl 4 via oral gavage for 4 weeks, and the serum levels of glutamate oxaloacetate transaminase, glutamate pyruvate transaminase, and albumin were higher than in the control group following chemical induction. To assess the potential of amniotic-fluid-derived stem cells (mAFSCs) to ameliorate liver fibrosis in vivo , mAFSCs were isolated from amniotic fluid of 13.5-day-old transgenic mice, which globally express the fluorescent protein, enhanced green fluorescent protein (EGFP), for tracing purposes (EGFP-mAFSCs). Single cells were injected via the mesentery (1 × 10 6 cells/mouse) of transplanted mice with liver fibrosis. Results Four weeks after EGFP-mAFSC transplantation, the serum glutamate oxaloacetate transaminase, glutamate pyruvate transaminase, and albumin levels of recipient mice in the EGFP-mAFSC-injected group were significantly decreased when compared with mice in the saline-injected group. Additionally, fibrotic tissues were evaluated using Masson's trichrome staining 4 weeks after cell transplantation. Shrinkage of the fibrotic area was observed in the EGFP-mAFSC-injected group. The tissue-repair effects were also confirmed by hydroxyproline content analysis. Conclusion The possible repair mechanism from our data revealed that EGFP-mAFSCs may fuse with the recipient liver cells. Overall, EGFP-mAFSCs can ameliorate liver fibrosis in mice, thus providing insight into the future development of regenerative medicine.

Amniotic fluid stem cells (AFSCs) are derived from the amniotic fluid of the developing fetus and can give rise to diverse differentiated cells of ectoderm, mesoderm, and endoderm lineages. Intrauterine transplantation is an approach used to cure inherited genetic fetal defects during the gestation period of pregnant dams. Certain disease such as osteogenesis imperfecta was successfully treated in affected fetal mice using this method. However, the donor cell destiny remains uncertain.The purpose of this study was to investigate the biodistribution and cell fate of Ds-red-harboring porcine AFSCs (Ds-red pAFSCs) after intrauterine transplantation into enhanced green fluorescent protein-harboring fetuses of pregnant mice. Pregnant mice (12.5 days) underwent open laparotomy with intrauterine pAFSC transplantation (5 × 10(4) cells per pup) into fetal peritoneal cavity.Three weeks after birth, the mice were sacrificed. Several samples from different organs were obtained for histological examination and flow cytometric analysis. Ds-red pAFSCs migrated most frequently into the intestines. Furthermore, enhanced green fluorescent protein and red fluorescent protein signals were co-expressed in the intestine and liver cells via immunohistochemistry studies.In utero xenotransplantation of pAFSCs fused with recipient intestinal cells instead of differentiating or maintaining the undifferentiated status in the tissue.