Whole-transcriptome sequencing analysis to identify key circRNAs, miRNAs, and mRNAs in the development of yak testes.

BACKGROUND: The Testis is an important reproductive organ in male mammals and the site for spermatogenesis, androgen synthesis, and secretion. Non-coding RNAs (ncRNAs) play an important regulatory role in various biological processes. However, the regulatory role of ncRNAs in the development of yak testes and spermatogenesis remains largely unclear. RESULT: In this study, we compared the expression profiles of circular RNAs (circRNAs), microRNAs (miRNAs), and messenger RNAs (mRNAs) in yak testicular tissue samples collected at 6 months (Y6M), 18 months (Y18M), and 4 years (Y4Y). Using RNA sequencing (RNA-Seq), we observed a significant difference in the expression patterns of ncRNAs in the samples collected at different testicular development stages. Twenty-two differentially expressed (DE) circRNAs, 69 DE miRNAs, and 64 DE mRNAs were detected in Y6M, Y18M, and Y4Y testicular samples, respectively. The results of gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that the source genes of DE circRNAs, predicted target genes of DE miRNAs, and DE mRNAs were specifically associated with signaling pathways and GO terms that were related to sperm synthesis, sperm vitality, and testicular development, such as cell cycle, Wnt signaling pathway, MAPK signaling pathway, GnRH signaling pathway, and spermatogenesis. The analysis of the circRNA-miRNA-mRNA network revealed that some DE ncRNAs, including miR-574, miR-449a, CDC42, and CYP11A1, among others, may be involved in testicular spermatogenesis. Concurrently, various circRNA-miRNA interaction pairs were observed. CONCLUSION: Our findings provide a database of circRNAs, miRNAs, and mRNAs expression profiles in testicular tissue of yaks at different developmental stages and a detailed understanding of the regulatory network of ncRNAs in yak testicular development and provide data that can help elucidate the molecular mechanisms underlying yak testicular development.

The Combined Analysis of GC-IMS and GC-MS Reveals the Differences in Volatile Flavor Compounds between Yak and Cattle-Yak Meat.

In order to investigate the composition and differences in volatile organic compounds (VOCs) in yak and cattle-yak meat and determine the key metabolites and metabolic pathways related to flavor formation. In this study, the VOCs and non-volatile metabolites in Longissimus dorsi muscle of two groups of samples were detected and analyzed by gas chromatography-ion migration spectrometry (GC-IMS) and gas chromatography-mass spectrometry (GC-MS). The results showed that 31 VOCs were identified by GC-IMS, including 5 alcohols, 5 ketones, 5 esters, 3 aldehydes, 2 furans, 2 hydrocarbons, 1 amine, 1 acid, 1 thiazole, 1 pyrazine, and 5 others. Most of them were alcohols, ketones, esters, and aldehydes. A total of 75 non-volatile metabolites with significant differences were obtained by GC-MS screening, among which amino acid contents such as serine, glycine, phenylalanine, and aspartic acid were significantly up-regulated in cattle-yak, and glutamic acid and tyrosine were significantly up-regulated in yak. The non-volatile differential metabolites in the two groups were significantly enriched in the metabolic pathways of arginine biosynthesis and oxidative phosphorylation. By combining GC-IMS and GC-MS, this study comprehensively and intuitively reflected the differences in VOCs between yak and cattle-yak meat, and clarified the metabolomic reasons for the differences in VOCs, so as to provide a theoretical basis for meat quality improvement.

Electrochemically Enabled Hydroxyphosphorylation of 1,3-Enynes to Access Phosphinyl-Substituted Propargyl Alcohols.

Catalytic difunctionalization with the direct activation of (O)P-H bonds has been recently established as a potentially robust platform to generate valuable organophosphorus compounds. In terms of 1,3-enynes, despite of the various catalytic methods developed for hydrophosphorylation, the radical-mediated hetero-functionalization of two different atoms has been less explored. In this study, we disclosed an electrochemically induced hydroxyphosphorylation of 1,3-enynes for the construction of phosphinyl-substituted propargyl alcohols. The system involves the direct activation of both arylphosphine oxides and oxygen in ambient air with no external metal or additive needed. The use of electrochemistry ensures the regioselective, atom-economic and eco-friendly for the difunctionalization process. This strategy highlights the advantages of mild reaction conditions, readily available starting materials and broad substrate scope, showing its practical synthetic value in organic synthesis.

Case Report: Assessment of primary myxofibrosarcoma in the left atrium using multimodal ultrasonography.

Primary myxofibrosarcoma of the heart, a rare cardiac malignancy, was diagnosed in a middle-aged female patient exhibiting progressive dyspnea following transthoracic echocardiography and pathological analysis. Postoperatively, the patient underwent chemotherapy and Lenvatinib mesylate therapy, with regular check-ups confirming her survival. After 10 months the patient is still alive and well.

Comprehensive analysis of differentially expressed mRNAs, circRNAs, and miRNAs and their ceRNA network in the testis of cattle-yak, yak, and cattle.

Cattle-yak is a hybrid offspring resulting from the crossbreeding of yak and cattle, and it exhibits substantial heterosis in production performance. However, male sterility in cattle-yak remains a concern. Reports suggest that noncoding RNAs are involved in the regulation of spermatogenesis. Therefore, in this study, we comprehensively compared testicular transcription profiles among cattle, yak, and cattle-yak. Numerous differentially expressed genes (DEGs), differentially expressed circRNAs (DECs), and differentially expressed miRNAs (DEMs) were identified in the intersection of two comparison groups, namely cattle versus cattle-yak and yak versus cattle-yak, with the number of DEGs, DECs, and DEMs being 4968, 360, and 59, respectively. The DEGs in cattle-yaks, cattle, and yaks were mainly associated with spermatogenesis, male gamete generation, and sexual reproduction. Concurrently, GO and KEGG analyses indicated that DEC host genes and DEM source genes were involved in the regulation of spermatogenesis. The construction of a potential competing endogenous RNA network revealed that some differentially expressed noncoding RNAs may be involved in regulating the expression of genes related to testicular spermatogenesis, including miR-423-5p, miR-449b, miR-34b/c, and miR-15b, as well as previously unreported miR-6123 and miR-1306, along with various miRNA-circRNA interaction pairs. This study serves as a valuable reference for further investigations into the mechanisms underlying male sterility in cattle-yaks.

Access Polyarylbipyrazoles via Palladium-Catalysis and Visible-Light-Driven C(sp3)-P(V) Cleavage Relay Strategy.

An unprecedented palladium-catalyzed and visible-light-driven relay reaction of allenylphosphine oxide with in situ generated nitrile imines is presented for the direct synthesis of highly valuable polyarylbipyrazole skeletons. This one-pot strategy involves double 1,3-dipolar cycloaddition and C(sp3)-P(V) bond cleavage under photocatalyst-free and mild reaction conditions. The approach features simple operation, a high step economy, and a broad substrate scope, affording the corresponding products in moderate to excellent yields.

Pd-catalyzed divergent regioselective annulation of phosphinyl allenes accessing polyarylfurans and 2H-chromenes.

A novel and efficient regioselective annulation of phosphinyl allenes with 2-bromophenol or 1-bromo-2-naphthol is achieved by palladium catalysis. The divergent pathway delivers structurally diverse polyarylfurans and 2H-chromene skeletons via two different Heck-type annulations. This protocol represents regioselectivity-tunable transformation of allenes into functionalized O-containing heterocycles with excellent group compatibility.

E/e' Ratio and Diastolic Function Between Deep Vein Catheterization and Internal Fistula Treatment in Patients with Chronic Kidney Disease.

Objective: To examine the relationship between diastolic function and the ratio of early diastolic mitral inflow to early diastolic mitral annular velocity (E/e') in patients with chronic renal disease who had deep vein catheterization and internal fistula. Methods: The clinical data of 50 uremia patients treated at The Affiliated Dongyang Hospital of Wenzhou Medical University from January 2020 to January 2022 were retrospectively analyzed. To assess the differences in E/e' ratio and patients' diastolic function between the two groups, they were split into two teams according to the various therapy modalities: the internal fistula team (n = 42) and the deep vein catheterization team (n = 8). Results: After treatment, the left ventricular end-diastolic diameter (LVd), E peak, a peak and E/A value, the volume and area of four chambers of the left ventricle (LV), the volume and area of two chambers of LV in both groups were significantly lower than those before treatment (P < .001). After treatment, the LVd left ventricular end-systolic diameter (LVs), the four-chamber volume of LV, and the two-chamber volume and area of LV in patients with internal fistula were significantly lower than those in patients with deep vein catheterization (P < .001). After treatment, E peak, A peak and E/A value, e' interventricular septum, E/e' value of interventricular septum, e' lateral wall, and E of lateral wall in patients with internal fistula group. Conclusion: Both deep vein catheterization and internal fistula treatment can improve the diastolic function and reduce the pulmonary pressure of uremic patients to a certain extent, but internal fistula treatment is better than deep vein catheterization in reducing LVd, LVs, LV four-chamber volume, LV two-chamber volume and area, and the effects of both in improving the E/e ratio of patients are not obvious.

"Boomerang" Strategy in Carbohydrate Chemistry: Diastereoselective Synthesis of C-Glycosylated Benzothiazoles from ortho-Isocyanophenyl Thioglycosides.

This paper reveals a novel "boomerang" strategy in the expedient and diastereoselective synthesis of C-nucleoside analogues. Bench-stable ortho-isocyanophenyl thioglycosides can be converted to glycosyl radicals through rapid and efficient C-S bond homolysis when they are irradiated by visible light. The glycosyl radicals are subsequently trapped by the corresponding leaving group or other radical acceptors to provide diverse C-nucleoside analogues under mild conditions.

Comparison of the 2009 Institute of Medicine and 2021 Chinese guidelines for gestational weight gain: A retrospective population-based cohort study.

OBJECTIVE: To analyze the associations between gestational weight gain (GWG) and perinatal outcomes based on the GWG guidelines of the Chinese Nutrition Society (CNS) and the Institute of Medicine (IOM). METHODS: This was a retrospective study with 9075 low-risk singleton pregnant women. Logistic regression model was used to analyze associations between GWG categories and perinatal outcomes. Sensitivity analyses were performed based on pre-pregnancy body mass index (calculated as weight in kilograms divided by the square of height in meters). RESULTS: Excessive GWG as defined by the two guidelines was associated with a higher risk of adverse perinatal outcomes. Inadequate GWG was associated with higher risks of small for gestational age (adjusted odds ratio [aOR] 1.34, 95% confidence interval [CI] 1.10-1.64) and preterm birth (aOR 1.70, 95% CI 1.22-2.36), but a lower risk of large for gestational age (LGA) (aOR 0.77, 95% CI 0.63-0.95) according to the IOM guidelines. When using the CNS guidelines, inadequate GWG was associated with only a lower risk of preterm birth (aOR 1.80, 95% CI 1.19-2.70). Sensitivity analyses suggested that excessive GWG was associated with a higher risk of LGA in underweight women. CONCLUSIONS: Both guidelines could demonstrate the relationship between GWG and adverse perinatal outcomes. The CNS guidelines were more suitable for the Chinese population with underweight or normal weight before pregnancy, whereas IOM was more suitable for pregnant women with inadequate GWG.

Identification and Characterization of Circular RNAs (circRNAs) Using RNA-Seq in Two Breeds of Cashmere Goats.

Circular RNA (circRNA) is a type of non-coding RNA generated from back-splicing the reactions of linear RNA. It plays an important role in various cellular and biological processes. However, there are few studies about the regulatory effect of circRNAs on cashmere fiber traits in cashmere goats. In this study, the expression profiles of circRNAs in skin tissue were compared between Liaoning cashmere (LC) goats and Ziwuling black (ZB) goats, with a significant difference in cashmere fiber yield, cashmere fiber diameter, and cashmere fiber color, using RNA-seq. A total of 11,613 circRNAs were expressed in the caprine skin tissue, and their type, chromosomal distribution, and length distribution were characterized. A total of 115 up-regulated circRNAs and 146 down-regulated circRNAs in LC goats were screened compared to ZB goats. The authenticity of 10 differentially expressed circRNAs was validated by detecting their expression levels and the head-to-tail splice junction using RT-PCR and DNA sequencing, respectively. The parent genes of differentially expressed circRNA were mainly enriched in some Gene Ontology (GO) terms and pathways related to cashmere fiber traits, such as the canonical Wnt signaling pathway, which is involved in the regulation of cell promotion, stem cell proliferation, Wnt signaling pathway regulation, epithelial morphogenesis, MAPK signaling pathway, and cell adhesion molecules pathway. Eight differentially expressed circRNAs were further selected to construct a circRNA-miRNA network, and some miRNAs that were previously reported as related to fiber traits were found in the network. This study provides a deep understanding of the roles of circRNAs in the regulation of cashmere fiber traits in cashmere goats and the involvement of differential splicing in phenotypic expression according to breed and region.

Clinical Study on Different Delivery Methods of Twin Pregnancy.

OBJECTIVE: To investigate the effect of different methods of delivery on the outcome of twin pregnancies. STUDY DESIGN: This is a retrospective cohort review of 627 twin pregnancies with delivery from January 2016 to December 2019. According to the clinical guidelines for cesarean section, the cesarean section and vaginal delivery groups were determined. Finally, the baseline information, delivery method, pregnancy outcome, and maternal and infant complications of the two groups were compared. RESULTS: For different delivery methods, the incidence of preeclampsia was significantly higher in the cesarean section group than in the vaginal delivery group (X 2 = 4.405, p < 0.05). There were 23 fetal growth ratios (FGR) in the cesarean section group, which were significantly higher than the vaginal delivery group (X 2 = 4.740, p < 0.05). However, the incidence of preterm premature rupture of membranes (PPROM) in the vaginal delivery group was significantly higher than in the cesarean section group (X 2 = 5.235, p < 0.05). In addition, the volume of postpartum bleeding in the vaginal delivery group was significantly higher than in the cesarean section group (t = 4.723, p < 0.001). The neonatal weights and 5-minute Apgar scores of the vaginal delivery group were lower than the cesarean section group, and the difference was statistically significant. In the vaginal delivery group, 48 and 26 neonates were transferred to the intensive care and neonatal units, respectively, which were significantly higher than in the cesarean section group (X 2 = 5.001, p < 0.05). CONCLUSION: The major complications of a twin pregnancy are gestational diabetes mellitus and PPROM. Cesarean section can reduce the rate of neonatal asphyxia in twins and improve the pregnancy outcome. KEY POINTS: · Twin pregnancy.. · Delivery methods.. · Pregnancy outcomes..

Development and validation of an ultrasound-based radiomics nomogram for predicting the luminal from non-luminal type in patients with breast carcinoma.

Introduction: The molecular subtype plays a significant role in breast carcinoma (BC), which is the main indicator to guide treatment and is closely associated with prognosis. The aim of this study was to investigate the feasibility and efficacy of an ultrasound-based radiomics nomogram in preoperatively discriminating the luminal from non-luminal type in patients with BC. Methods: A total of 264 BC patients who underwent routine ultrasound examination were enrolled in this study, of which 184 patients belonged to the training set and 80 patients to the test set. Breast tumors were delineated manually on the ultrasound images and then radiomics features were extracted. In the training set, the T test and least absolute shrinkage and selection operator (LASSO) were used for selecting features, and the radiomics score (Rad-score) for each patient was calculated. Based on the clinical risk features, Rad-score, and combined clinical risk features and Rad-score, three models were established, respectively. The performances of the models were validated with receiver operator characteristic (ROC) curve and decision curve analysis. Results: In all, 788 radiomics features per case were obtained from the ultrasound images. Through radiomics feature selection, 11 features were selected to constitute the Rad-score. The area under the ROC curve (AUC) of the Rad-score for predicting the luminal type was 0.828 in the training set and 0.786 in the test set. The nomogram comprising the Rad-score and US-reported tumor size showed AUCs of the training and test sets were 0.832 and 0.767, respectively, which were significantly higher than the AUCs of the clinical model in the training and test sets (0.691 and 0.526, respectively). However, there was no significant difference in predictive performance between the Rad-score and nomogram. Conclusion: Both the Rad-score and nomogram can be applied as useful, noninvasive tools for preoperatively discriminating the luminal from non-luminal type in patients with BC. Furthermore, this study might provide a novel technique to evaluate molecular subtypes of BC.

MicroRNA-200c Affects Milk Fat Synthesis by Targeting PANK3 in Ovine Mammary Epithelial Cells.

Milk fat is the foremost nutrient of milk and a vital indicator in evaluating milk quality. Accumulating evidence suggests that microRNAs (miRNAs) are involved in the synthesis of milk fat. The miR-200c is closely related to lipid metabolism, but little is known about its effect on the synthesis of milk fat in MECs of ewes. Herein, the effect of miR-200c on the proliferation of ovine mammary epithelial cells (MECs) and its target relationship with a predicted target gene were investigated. The regulatory effects of miR-200c on the expression of the target genes and the content of triglycerides in ovine MECs were further analyzed. The results revealed that the expression level of miR-200c was differentially expressed in both eight tissues selected during lactation and in mammary gland tissues at different physiological periods. Overexpression of miR-200c inhibited the viability and proliferation of ovine MECs, while inhibition of miR-200c increased cell viability and promoted the proliferation of ovine MECs. Target gene prediction results indicated that miR-200c would bind the 3'UTR region of pantothenate kinase 3 (PANK3). Overexpression of miR-200c reduced the luciferase activity of PANK3, while inhibition of miR-200c increased its luciferase activity. These findings illustrated that miR-200c could directly interact with the target site of the PANK3. It was further found that overexpression of miR-200c reduced the expression levels of PANK3 and, thus, accelerated the synthesis of triglycerides. In contrary, the inhibitor of miR-200c promoted the expression of PANK3 that, thus, inhibited the synthesis of triglycerides in ovine MECs. Together, these findings revealed that miR-200c promotes the triglycerides synthesis in ovine MECs via increasing the lipid synthesis related genes expression by targeting PANK3.

Basic research investment, innovation capability improvement, and economic growth efficiency.

This study explores the relationship between the proportion of basic research investment in R&D expenditure and regional economic growth efficiency and its influence path. A panel of data from 31 China's provincial was analysed by the fixed-effects model over 2009 to 2019. Empirical results highlight that: (1) the increase in the proportion of basic research investment has a significant promoting effect on regional economic growth efficiency, but it presents an "inverted U-shaped" impact path. Meanwhile, the threshold effect model test results reveal that basic research investment plays a more significant role in promotion when the per capita income level exceeds a certain threshold. (2) The test results of the mediating effect model indicate that the improvement of the level of human capital is the intermediate channel through which the investment in basic research affects regional economic growth efficiency, while the mediating effect of the increase of knowledge storage on the process of the investment in basic research affecting regional economic growth efficiency is not supported by data. Potential policy measures are discussed.

Clinical analysis on the influencing factors related to preterm twin pregnancy.

OBJECTIVE: The aim of this study was to explore the influencing factors of preterm twin pregnancy. METHODS: In total, 602 twin-pregnant women delivered from February 2016 to February 2020 were analyzed retrospectively. According to whether the pregnant women were preterm or not, they were divided into preterm group (n=363) and term group (n=239). Baseline information, such as maternal age, address, and education level of the pregnant women, were collected. The clinical information of pregnant women, such as chorionic, preeclampsia, gestational diabetes, premature rupture of membranes, abnormal fetal position, and fetal weight, were analyzed. Logistic regression analysis was used to analyze the risk factors. p-value <0.05 was considered statistically significant. RESULTS: In the preterm group, monochorionic diamniotic comorbidities were significantly higher compared with the control group (p<0.05). Higher risks of preterm group have lower education (p<0.05). Multiple logistic regression analysis demonstrated that education, preeclampsia, and premature rupture of membranes were risk factors for preterm twin pregnancy. CONCLUSIONS: Preterm birth in twin pregnancy is associated with many risk factors, such as education, preeclampsia, and premature rupture of membranes. Pregnancy supervision and prenatal guidance for twin pregnancy should be strengthened. Furthermore, early detection and diagnosis of comorbidities can improve maternal and neonatal outcomes.

Integration of ultrasound radiomics features and clinical factors: A nomogram model for identifying the Ki-67 status in patients with breast carcinoma.

Objective: The aim of this study was to develop and validate an ultrasound-based radiomics nomogram model by integrating the clinical risk factors and radiomics score (Rad-Score) to predict the Ki-67 status in patients with breast carcinoma. Methods: Ultrasound images of 284 patients (196 high Ki-67 expression and 88 low Ki-67 expression) were retrospectively analyzed, of which 198 patients belonged to the training set and 86 patients to the test set. The region of interest of tumor was delineated, and the radiomics features were extracted. Radiomics features underwent dimensionality reduction analysis by using the independent sample t test and least absolute shrinkage and selection operator (LASSO) algorithm. The support vector machine (SVM), logistic regression (LR), decision tree (DT), random forest (RF), naive Bayes (NB) and XGBoost (XGB) machine learning classifiers were trained to establish prediction model based on the selected features. The classifier with the highest AUC value was selected to convert the output of the results into the Rad-Score and was regarded as Rad-Score model. In addition, the logistic regression method was used to integrate Rad-Score and clinical risk factors to generate the nomogram model. The leave group out cross-validation (LGOCV) method was performed 200 times to verify the reliability and stability of the nomogram model. Results: Six classifier models were established based on the 15 non-zero coefficient features. Among them, the LR classifier achieved the best performance in the test set, with the area under the receiver operating characteristic curve (AUC) value of 0.786, and was obtained as the Rad-Score model, while the XGB performed the worst (AUC, 0.615). In multivariate analysis, independent risk factor for high Ki-67 status was age (odds ratio [OR] = 0.97, p = 0.04). The nomogram model based on the age and Rad-Score had a slightly higher AUC than that of Rad-Score model (AUC, 0.808 vs. 0.798) in the test set, but no statistical difference (p = 0.144, DeLong test). The LGOCV yielded a median AUC of 0.793 in the test set. Conclusions: This study proposed a convenient, clinically useful ultrasound radiomics nomogram model that can be used for the preoperative individualized prediction of the Ki-67 status in patients with BC.

Screening high-risk population of persistent postpartum hypertension in women with preeclampsia using latent class cluster analysis.

BACKGROUND: A significant proportion of women with preeclampsia (PE) exhibit persistent postpartum hypertension (PHTN) at 3 months postpartum associated with cardiovascular morbidity. This study aimed to screen patients with PE to identify the high-risk population with persistent PHTN. METHODS: This retrospective cohort study enrolled 1,000 PE patients with complete parturient and postpartum blood pressure (BP) profiles at 3 months postpartum. The enrolled patients exhibited new-onset hypertension after 20 weeks of pregnancy, while those with PE superimposed upon chronic hypertension were excluded. Latent class cluster analysis (LCCA), a method of unsupervised learning in machine learning, was performed to ascertain maternal exposure clusters from eight variables and 35 subordinate risk factors. Logistic regression was applied to calculate odds ratios (OR) indicating the association between clusters and PHTN. RESULTS: The 1,000 participants were classified into three exposure clusters (subpopulations with similar characteristics) according to persistent PHTN development: high-risk cluster (31.2%), medium-risk cluster (36.8%), and low-risk cluster (32.0%). Among the 1,000 PE patients, a total of 134 (13.4%) were diagnosed with persistent PHTN, while the percentages of persistent PHTN were24.68%, 10.05%, and 6.25% in the high-, medium-, and low-risk clusters, respectively. Persistent PHTN in the high-risk cluster was nearly five times higher (OR, 4.915; 95% confidence interval (CI), 2.92-8.27) and three times (OR, 2.931; 95% CI, 1.91-4.49) than in the low- and medium-risk clusters, respectively. Persistent PHTN did not differ between the medium- and low-risk clusters. Subjects in the high-risk cluster were older and showed higher BP, poorer prenatal organ function, more adverse pregnancy events, and greater medication requirement than the other two groups. CONCLUSION: Patients with PE can be classified into high-, medium-, and low-risk clusters according to persistent PHTN severity; each cluster has cognizable clinical features. This study's findings stress the importance of controlling persistent PHTN to prevent future cardiovascular disease.

MicroRNA-199a-3p regulates proliferation and milk fat synthesis of ovine mammary epithelial cells by targeting VLDLR.

In our previous study, microRNA (miR)-199a-3p was found to be the most upregulated miRNA in mammary gland tissue during the non-lactation period compared with the peak-lactation period. However, there have been no reports describing the function of miR-199a-3p in ovine mammary epithelial cells (OMECs) and the biological mechanisms by which the miRNA affects cell proliferation and milk fat synthesis in sheep. In this study, the effect of miR-199a-3p on viability, proliferation, and milk fat synthesis of OMECs was investigated, and the target relationship of the miRNA with very low-density lipoprotein receptor (VLDLR) was also verified. Transfection with a miR-199a-3p mimic increased the viability of OMECs and the number of Edu-labeled positive OMECs. In contrast, a miR-199-3p inhibitor had the opposite effect with the miR-199a-3p mimic. The expression levels of three marker genes were also regulated by both the miR-199a-3p mimic and miR-199-3p inhibitor in OMECs. Together, these results suggest that miR-199a-3p promotes the viability and proliferation of OMECs. A dual luciferase assay confirmed that miR-199a-3p can target VLDLR by binding to the 3'-untranslated regions (3'UTR) of the gene. Further studies found a negative correlation in the expression of miR-199a-3p with VLDLR. The miR-199a-3p mimic decreased the content of triglycerides, as well as the expression levels of six milk fat synthesis marker genes in OMECs, namely, lipoprotein lipase gene (LPL), acetyl-CoA carboxylase alpha gene (ACACA), fatty acid binding protein 3 gene (FABP3), CD36, stearoyl-CoA desaturase gene (SCD), and fatty acid synthase gene (FASN). The inhibition of miR-199a-3p increased the level of triglycerides and the expression of LPL, ACACA, FABP3, SCD, and FASN in OMECs. These findings suggest that miR-199a-3p inhibited milk fat synthesis of OMECs. This is the first study to reveal the molecular mechanisms by which miR-199a-3p regulates the proliferation and milk fat synthesis of OMECs in sheep.

Tissue-Specific Expression of Circ_015343 and Its Inhibitory Effect on Mammary Epithelial Cells in Sheep.

Circular RNAs (circRNAs) are a kind of non-coding RNA that have an important molecular function in mammary gland development and lactation of mammals. In our previous study, circ_015343 was found to be highly expressed in the ovine mammary gland tissue at the peak-lactation period by using RNA sequencing (RNA-seq). In the present study, the authenticity of circ_015343 was confirmed by using reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and Sanger sequencing. The circ_015343 was derived from the complete 10 exons of aminoadipic semialdehyde synthase (AASS), ranging from exon 2 to exon 11 and mainly located in cytoplasm of ovine mammary epithelial cells. The circRNA was found to be expressed in eight ovine tissues, with the highest expression level in the mammary gland and the least expression in Longissimus dorsi muscle. The circ_015343 had a lower level of expression in a sheep breed with higher milk yield and milk fat content. The disturbed circ_015343 increased the viability and proliferation of the ovine mammary epithelial cells. The inhibition of circ_015343 also increased the expression levels of three milk fat synthesis marker genes: acetyl-coenzyme A carboxylase alpha (ACACA), fatty acid-binding protein 4 (FABP4), and sterol regulatory element-binding protein 1 (SREBP1), as well as three proliferation-related genes: cyclin dependent kinase 2 (CDK2), cyclin dependent kinase 4 (CDK4) and proliferating cell nuclear antigen (PCNA), but decreased the expression level of its parent gene AASS. A circRNA-miRNA-mRNA interaction network showed that circ_015343 would bind some microRNAs (miRNAs) to regulate the expression of functional genes related to the development of mammary gland and lactation. This study contributes to a better understanding of the roles of circ_015343 in the mammary gland of sheep.