Effects of dietary triiodothyronine or dopamine on small intestinal oxygen consumption in chicks.

This study aimed to investigate the effects of triiodothyronine (T3)- or dopamine (Dp)-supplemented diets on oxygen consumption by Na+, K+-ATPase activity in broiler chicks. Five groups, each with twenty-four 6-day-old chicks, randomly received one of the five dietary treatments: (1) Basal diet (commercial broiler rations with 23.0% crude protein and 3,133 kcal metabolizable energy/kg) or CON, (2) basal diet plus 0.7 µmol Dp/kg diet or Dp0.7, (3) basal diet plus 2.4 µmol Dp/kg diet or Dp2.4, (4) basal diet plus 1.9 µmol T3/kg diet or T1.9, and (5) basal diet plus 3.8 µmol T3/kg diet or T3.8 from 6 to 14 days of age. There were four replicates per treatment and 120 birds in total. At 14 days of age, three chicks from each replicate of each treatment were pooled into a flock and fed commercial broiler diets until 7 weeks of age. Compared to CON group, birds fed with T3-supplemented diets had lower thyroid, abdominal fat pad, gizzard and pancreas weight, and heavier heart weight adjusted for fasted body weight. Chicks with T1.9 had lower ileal densities at 14 day old compared with those in Dp groups or CON. Chicks with T3.8 exhibited greater duodenal and jejunal O2 consumptions as well as ouabain-sensitive O2 consumptions of jejunum and small intestine (duodenum, jejunum, and ileum) by 46.5%, 58.3%, 40.6%, and 26.4% increases, than those in CON. Partial correlation analysis revealed that the weight and length of the small intestine were negatively correlated with body weight gain. Oxygen consumption in the various small intestinal segments was negatively correlated with their respective densities (mg/mm2). In conclusion, a greater oxygen requirement for maintaining ouabain-sensitive respiration (Na+-K+-ATPase) in the intestine limits energy availability to support gastrointestinal tract growth and, thereby, may result in lower body weight gain.

Glucocorticoid transiently upregulates mitochondrial biogenesis in the osteoblast.

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.

Effect of line and floor type on growth performance and feather characterization during the growth period of White Roman geese.

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<0.001). Afterwards, the body weight, back length, keel length, chest girth and main wing feather length among 3 farms showed no significance difference prior to 12 weeks. The CF group showed heavier body weight, shorter back length, longer keel length, shorter chest girth and shorter main wing feather length than the CSF group prior to 12 weeks. The down weight in the CF was heavier than the CSF group (57.1 g vs 41.8 g; p<0.01) prior to 13 weeks. CONCLUSION: The body weight showed the positive relations for dry feather weight (r = 0.59), down weight (r = 0.69), percent of the down weight of live body weight prior to 13 weeks (r = 0.61).

Different susceptibilities of osteoclasts and osteoblasts to glucocorticoid-induced oxidative stress and mitochondrial alterations.

Glucocorticoid-induced bone loss is the most common form of secondary osteoporosis. This toxic effect has not been efficiently managed, possibly due to the incomplete understanding of the extraordinarily diverse cellular responses induced by glucocorticoid treatment. Previous literatures revealed that high dose of exogenous glucocorticoid triggers apoptosis in osteocytes and osteoblasts. This cell death is associated with glucocorticoid-induced oxidative stress. In this study, we aimed to investigate the mechanisms of glucocorticoid-induced apoptosis in osteoblasts and examine the responses of osteoclasts to the synthetic glucocorticoid, dexamethasone. We demonstrated the biphasic effects of exogenous glucocorticoid on osteoblastic mitochondrial functions and elevated intracellular oxidative stress in a dose- and time-dependent manner. On comparison, similar treatment did not induce mitochondrial dysfunctions and oxidative stress in osteoclasts. The production of reactive oxygen/nitrogen species was decreased in osteoclasts. The differences are not due to varying efficiency of cellular antioxidant system. The opposite effects on nitrogen oxide synthase might provide an explanation, as the expression levels of nos2 gene are suppressed in the osteoclast but elevated in the osteoblast. We further revealed that glucocorticoids have a substantial impact on the osteoblastic mitochondria. Basal respiration rate and ATP production were increased upon 24 h incubation of glucocorticoids. The increase in proton leak and nonmitochondrial respiration suggests a potential source of glucocorticoid-induced oxidative stress. Long-term incubation of glucocorticoids accumulates these detrimental changes and results in cytochrome C release and mitochondrial breakdown, consequently leading to apoptosis in osteoblasts. The mitochondrial alterations might be other sources of glucocorticoid-induced oxidative stress in osteoblasts.

Prenatal transplantation of human amniotic fluid stem cells for spinal muscular atrophy.

PURPOSE OF REVIEW: 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. RECENT FINDINGS: 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. SUMMARY: 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.

Infusion of Porcine-Derived Amniotic Fluid Stem Cells for Treatment of Experimental Colitis in Mice.

Recently, stem cells have offered an alternative treatment for inflammatory bowel disease (IBD) or colitis to overcome the poor outcomes associated with current therapies. Amniotic fluid-derived stem cells (AFSCs) 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 AFSCs (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 weightloss 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 (IL-6) and interferon gamma (IFN-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 IBD or colitis.

Ear fibroblasts derived from Taiwan yellow cattle are more heat resistant than those from Holstein cattle.

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 (P<0.05) 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 (P<0.05) 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 (P<0.05) 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 (P<0.05) 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 (P<0.05) 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 (P<0.05) 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 (P<0.05) 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.

Artificial insemination of Holstein heifers with sex-sorted semen during the hot season in a subtropical region.

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.

Engraftment of mouse amniotic fluid-derived progenitor cells after in utero transplantation in mice.

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.

Establishment of a DsRed-monomer-harboring ICR transgenic mouse model and effects of the transgene on tissue development.

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.

Differentiation of Enhanced Green Fluorescent Protein-Labeled Mouse Amniotic Fluid-Derived Stem Cells into Cardiomyocyte-Like Beating Cells.

BACKGROUND: Amniotic fluid-derived stem cells (AFSCs) possess optimal differentiation potential and are a promising resource for cell therapy and tissue engineering. Mouse is a good model to be studied for pre-clinical research. METHODS: In this study, we successfully established enhanced green fluorescent protein mouse-derived amniotic fluid stem cells (EGFP-mAFSCs) and investigated whether EGFP-mAFSCs possess the ability to differentiate into cardiomyocytes by in vitro culture. We evaluated stem-cell differentiation using immunofluorescence. RESULTS: This study showed that EGFP-mAFSCs can give rise to spontaneously beating cardiomyocyte-like cells expressing the specific markers c-kit, myosin heavy chain, and cardiac troponin I. CONCLUSIONS: We demonstrated that mAFSCs have the in vitro propensity to acquire a cardiomyogenic phenotype and to a certain extent cardiomyocytes; however the process efficiency which gives rise to cardiomyocyte-like cells remains quite low (2 out of 10 were found). KEY WORDS: Amniotic fluid; Cardiomyocytes; In vitro differentiation; Stem cells.

Cell fusion phenomena detected after in utero transplantation of Ds-red-harboring porcine amniotic fluid stem cells into EGFP transgenic mice.

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 × 10(4) 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.

Using deep learning to accurately detect sow vulva size in a group pen with a single camera.

This paper presents a non-contact method for the detection of changes in sow vulva size in a group pen. The traditional approach to estrus detection is manually pressing down on the back of the sow to elicit standing responses; however, this method causes undue distress for sows not in estrus. When a sow is in estrus, the vulva is red and swollen due to the presence of endocrine. Monitoring changes in vulva size to detect estrus with as little impact on the sow as possible is the focus of this study. This is achieved using a single camera combined with a deep learning framework. Our approach comprises two steps: vulva detection and vulva size conversion. Images of sows of Yorkshire, Landrace, and Duroc breeds were collected in group housing, and the vulva was detected through artificial markers and the network architecture of YOLO v4. Based on the internal and external parameters of the camera, the detected size was converted into millimeters and the results of manual measurement (MM) and automatic calculation combined to calculate the size of the vulva. Analysis of the calculated size compared with MM indicates that the object recognition rate of the system exceeds 97.06%, with a size error of only + 1.70 to -4.47 mm and high-calculation efficiency (>2.8 frames/s). Directions for future research include the automatic detection of pig width.

The size of a sow's vulva is an important indicator of sow estrus. Non-contact means of monitoring size changes for estrus timing would represent a significant contribution to the field of pig farming. This paper thus focuses on development of a system for the automatic detection of sow vulva size using a single camera combined with a deep learning framework. Experiments showed that the object recognition rate of the system exceeds 97.06%, the vulva size error is +1.70 to −4.47 mm, and the calculation efficiency is high (>2.8 frames/s).

Effects of Antrodia cinnamomea solid culture mycelium by-products on growth performance and immune response in weaning black piglets.

This study evaluated the effects of supplementation with Antrodia cinnamomea mycelium by-product (ACBP) on growth performance and immune response in weaning piglets. Total available content and antioxidant capacity of ACBP were determined. Ninety-six black pigs were randomly distributed to 24 pens. Study compared four groups which were supplemented with ACBP at 0%, 2.5%, 5%, or 10% for 6 weeks after weaning at 4 weeks. Results showed that ACBP on total phenolic, total flavonoid, and total triterpenoids contents were 13.68 mg GAE/g DW, 1.67 µg QE/g DW, and 15.6 mg/g, respectively. Weaning piglets fed 2.5% ACBP showed a significant decreased body weight gain compared with those supplemented with 5% ACBP, 10% ACBP, and control groups. Results showed that all ACBP groups increased the villi height of jejunum significantly. Incidence of diarrhea in 11 weeks with supplementation with 5% and 10% ACBP diets were lower than in control group. The 10% ACBP group showed significantly lower expression of immune response genes (IL-1ß, IL-6, IL-8, TNF-α, and IFN-γ) than the 2.5% and 5% ACBP groups. Based on results, dietary supplementation with 10% ACBP did not significantly affect body weight but could decrease piglet diarrhea condition and expression of IL-1ß and IL-6 genes.

Identification of Molecular Profile of Ear Fibroblasts Derived from Spindle-Transferred Holstein Cattle with Ooplasts from Taiwan Yellow Cattle under Heat Stress.

Global warming has a significant impact on the dairy farming industry, as heat stress causes reproductive endocrine imbalances and leads to substantial economic losses, particularly in tropical-subtropical regions. The Holstein breed, which is widely used for dairy production, is highly susceptible to heat stress, resulting in a dramatic reduction in milk production during hot seasons. However, previous studies have shown that cells of cows produced from reconstructed embryos containing cytoplasm (o) from Taiwan yellow cattle (Y) have improved thermotolerance despite their nuclei (n) being derived from heat-sensitive Holstein cattle (H). Using spindle transfer (ST) technology, we successfully produced ST-Yo-Hn cattle and proved that the thermotolerance of their ear fibroblasts is similar to that of Y and significantly better than that of H (p < 0.05). Despite these findings, the genes and molecules responsible for the different sensitivities of cells derived from ST-Yo-Hn and H cattle have not been extensively investigated. In the present study, ear fibroblasts from ST-Yo-Hn and H cattle were isolated, and differentially expressed protein and gene profiles were compared with or without heat stress (hs) (42 °C for 12 h). The results revealed that the relative protein expression levels of pro-apoptotic factors, including Caspase-3, -8, and -9, in the ear fibroblasts from the ST-Yo-Hn-hs group were significantly lower (p < 0.05) than those from the H-hs group. Conversely, the relative expression levels of anti-apoptotic factors, including GNA14 protein and the CRELD2 and PRKCQ genes, were significantly higher (p < 0.05) in the ear fibroblasts from the ST-Yo-Hn-hs group compared to those from the H-hs group. Analysis of oxidative phosphorylation-related factors revealed that the relative expression levels of the GPX1 gene and Complex-I, Complex-IV, CAT, and PGLS proteins were significantly higher (p < 0.05) in the ear fibroblasts from the ST-Yo-Hn-hs group compared to those from the H-hs group. Taken together, these findings suggest that ear fibroblasts from ST-Yo-Hn cattle have superior thermotolerance compared to those from H cattle due to their lower expression of pro-apoptotic factors and higher expression of oxidative phosphorylation and antioxidant factors. Moreover, this improved thermotolerance is attributed, at least partially, to the cytoplasm derived from more heat-tolerant Y cattle. Hence, using ST technology to produce more heat-tolerant H cattle containing Y cytoplasm could be a feasible approach to alleviate the negative impacts of heat stress on dairy cattle in tropical-subtropical regions.

Let-7b-mediated suppression of basigin expression and metastasis in mouse melanoma cells.

Basigin (Bsg), also called extracellular matrix metalloproteinase inducer (EMMPRIN), is highly expressed on the surface of tumor cells and stimulates adjacent fibroblasts or tumor cells to produce matrix metalloproteinases (mmps). It has been shown that Bsg plays an important role in growth, development, cell differentiation, and tumor progression. MicroRNAs (miRNAs) are a class of short endogenous non-protein coding RNAs of 20-25 nucleotides (nt) that function as post-transcriptional regulators of gene expression by base-pairing to their target mRNAs and thereby mediate cleavage of target mRNAs or translational repression. In this study, let-7b, one of the let-7 family members, was investigated for its effect on the growth and invasiveness of the mouse melanoma cell line B16-F10. We have shown that let-7b can suppress the expression of Bsg in B16-F10 cells and also provided evidence that this suppression could result in the indirect suppression of mmp-9. The ability of B16-F10 cells transfected with let-7b to invade or migrate was significantly reduced. In addition, let-7b transfected B16-F10 cells displayed an inhibition of both cellular proliferation and colony formation. Furthermore, it was shown that the overexpression of let-7b in B16-F10 cells could reduce lung metastasis. Taken together, the present study identifies let-7b as a tumor suppressor that represses cancer cell proliferation and migration as well as tumor metastasis in mouse melanoma cells.

Deep Learning to Detect Triangular Fibrocartilage Complex Injury in Wrist MRI: Retrospective Study with Internal and External Validation.

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.

Effects of intracytoplasmic sperm injection timing and fertilization methods on the development of bovine spindle transferred embryos.

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.

The Exploration of miRNAs From Porcine Fallopian Tube Stem Cells on Porcine Oocytes.

Fallopian tube is essential to fertilization and embryonic development. Extracellular vesicles (EVs) from Fallopian tube containing biological regulatory factors, such as lipids, proteins and microRNAs (miRNAs) serve as the key role. At present, studies on oocytes from porcine oviduct and components from EVs remain limited. We aim to explore the effect of EVs secreted by porcine fallopian tube stem cells (PFTSCs) on oocyte. When the fifth-generation PFTSCs reached 80-90% of confluency, the pig in vitro maturation medium was utilized, and the conditioned medium collected for oocyte incubations. To realize the functions of EVs, several proteins were used to determine whether extracted EVs were cell-free. Field emission scanning electron microscope and nanoparticle tracking analyzer were used to observe the morphology. By next generation sequencing, 267 miRNAs were identified, and those with higher expression were selected to analyze the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment maps. The selected miR-152-3p, miR-148a-3p, miR-320a-3p, let-7f-5p, and miR-22-3p, were predicted to target Cepb1 gene affecting MAPK pathway. Of the five miRNAs, miR-320a-3p showed significant difference in maturation rate in vitro maturation. The blastocyst rate of pig embryos was also significantly enhanced by adding 50 nM miR-320a-3p. In vitro culture with miR-320a-3p, the blastocyst rate was significantly higher, but the cleavage rate and cell numbers were not. The CM of PFTSCs effectively improves porcine oocyte development. The miRNAs in EVs are sequenced and identified. miR-320a-3p not only helps the maturation, but also increases the blastocyst rates.

Effect of Forskolin on Body Weight, Glucose Metabolism and Adipocyte Size of Diet-Induced Obesity in Mice.

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.