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The multifunctional enzyme fatty acid synthase (FASN) catalyzed the second step of the de novo biosynthesis of long-chain fatty acids in mammals. In this research, we investigated the variants in ketoacyl reductase (KR) and thioesterase (TE) domains in FASN by using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). As a result, two single nucleotide polymorphisms (SNPs) were detected and genotyped in 407 Chinese Holstein cows. The non-synonymous g.16024G>A, g.17924G>A in KR and TE domains respectively resulted in a non-conservative substitution of alanine by threonine. g.17924G>A was associated with milk fat percentage, and the milk fat percentage of homozygous genotype AA was significantly higher than the other genotypes. The polymorphism information content (PIC) analysis showed that g.17924G>A in the cows population belonged to the intermediate polymorphism level. The results preliminarily indicated that g.17924 G>A of FASN gene is a potential genetic marker for application of marker-assisted selection programmers to improve milk fat percentage in Chinese dairy cattle. [ABSTRACT FROM AUTHOR]

The Booroola fecunditymutation (FecB) in Small Tail Han sheep has been shown to enhance ovulation rates and litter sizes by affecting the hypothalamic-pituitary-gonadal (HPG) axis. Despite the pituitary's role in reproductive regulation, its involvement in FecB-induced ovulation remains understudied. Our study aimed to fill this gap by analyzing pituitary tissues from FecB homozygous (BB) and wildtype (WW) ewes during luteal and follicular phases using tandem mass tag-based protein quantification and the DIABLO framework for proteomic and transcriptomic data integration. Significant differences in 277 proteins were observed across estrus periods, with network analysis highlighting the voltagedependent calcium channel L-type alpha-1C as a key convergence point in oxytocin signaling and GnRH secretion pathways. The DIABLO method revealed a strong correlation (0.98) between proteomic and transcriptomic datasets, indicating a coordinated response in FecB ewes. Notably, higher expression levels of Follicle Stimulating Hormone Subunit Beta (FSHB) and Luteinizing Hormone Subunit Beta (LHB) were found in BB ewes during the follicular phase, potentially due to elevated E2 concentrations. Furthermore, our analysis identified genes related to the Gamma-aminobutyric acid type A receptor family (GABRA2, GABRG1, GABRB1) in the pituitary, with GABRB1 showing higher expression in BB ewes. This suggests a role for GABA in modulating GnRH and gonadotropin feedback loops, potentially contributing to the FecB mutation's effect on ovulation. This study provides novel insights into the pituitary's role in fertility among FecB sheep, identifying GABA as a potential regulatory factor within the HPG axis. The findings also open avenues for discovering new biomarkers in pituitary endocrinology for sheep breeding purposes. [ABSTRACT FROM AUTHOR]

Chickens are a major source of meat and eggs in human food and have significant economic value. Cadmium (Cd) is a common environmental pollutant that can contaminate feed and drinking water, leading to kidney injury in livestock and poultry, primarily by inducing the generation of free radicals. It is necessary to develop potential medicines to prevent and treat Cd-induced nephrotoxicity in poultry. Luteolin (Lut) is a natural flavonoid compound mainly extracted from peanut shells and has a variety of biological functions to defend against oxidative damage. In this study, we aimed to demonstrate whether Lut can alleviate kidney injury under Cd exposure and elucidate the underlying molecular mechanisms. Renal histopathology and cell morphology were observed. The indicators of renal function, oxidative stress, DNA damage and repair, NAD+ content, SIRT1 activity, and autophagy were analyzed. In vitro data showed that Cd exposure increased ROS levels and induced oxidative DNA damage and repair, as indicated by increased 8-OHdG content, increased γ-H2AX protein expression, and the over-activation of the DNA repair enzyme PARP-1. Cd exposure decreased NAD+ content and SIRT1 activity and increased LC3 II, ATG5, and particularly p62 protein expression. In addition, Cd-induced oxidative DNA damage resulted in PARP-1 over-activation, reduced SIRT1 activity, and autophagic flux blockade, as evidenced by reactive oxygen species scavenger NAC application. The inhibition of PARP-1 activation with the pharmacological inhibitor PJ34 restored NAD+ content and SIRT1 activity. The activation of SIRT1 with the pharmacological activator RSV reversed Cd-induced autophagic flux blockade and cell injury. In vivo data demonstrated that Cd treatment caused the microstructural disruption of renal tissues, reduced creatinine, and urea nitrogen clearance, raised MDA content, and decreased the activities or contents of antioxidants (GSH, T-SOD, CAT, and T-AOC). Cd treatment caused oxidative DNA damage and PARP-1 activation, decreased NAD+ content, decreased SIRT1 activity, and impaired autophagic flux. Notably, the dietary Lut supplement observably alleviated these alterations in chicken kidney tissues induced by Cd. In conclusion, the dietary Lut supplement alleviated Cd-induced chicken kidney injury through its potent antioxidant properties by relieving the oxidative DNA damage-activated PARP-1-mediated reduction in SIRT1 activity and repairing autophagic flux blockade. [ABSTRACT FROM AUTHOR]

Cadmium (Cd) is a common environmental pollutant and occupational toxicant that seriously affects various mammalian organs, especially the kidney. Iron ion is an essential trace element in the body, and the disorder of iron metabolism is involved in the development of multiple pathological processes. An iron overload can induce a new type of cell death, defined as ferroptosis. However, whether iron metabolism is abnormal in Cd-induced nephrotoxicity and the role of ferroptosis in Cd-induced nephrotoxicity need to be further elucidated. Sprague Dawley male rats were randomly assigned into three groups: a control group, a 50 mg/L CdCl2-treated group, and a 75 mg/L CdCl2-treated group by drinking water for 1 month and 6 months, respectively. The results showed that Cd could induce renal histopathological abnormalities and dysfunction, disrupt the mitochondria's ultrastructure, and increase the ROS and MDA content. Next, Cd exposure caused GSH/GPX4 axis blockade, increased FTH1 and COX2 expression, decreased ACSL4 expression, and significantly decreased the iron content in proximal tubular cells or kidney tissues. Further study showed that the expression of iron absorption-related genes SLC11A2, CUBN, LRP2, SLC39A14, and SLC39A8 decreased in proximal tubular cells or kidneys after Cd exposure, while TFRC and iron export-related gene SLC40A1 did not change significantly. Moreover, Cd exposure increased SLC11A2 gene expression and decreased SLC40A1 gene expression in the duodenum. Finally, NAC or Fer-1 partially alleviated Cd-induced proximal tubular cell damage, while DFO and Erastin further aggravated Cd-induced cell damage. In conclusion, our results indicated that Cd could cause iron deficiency and chronic kidney injury by interfering with the iron metabolism rather than typical ferroptosis. Our findings suggest that an abnormal iron metabolism may contribute to Cd-induced nephrotoxicity, providing a novel approach to preventing kidney disease in clinical practice. [ABSTRACT FROM AUTHOR]

Cadmium (Cd) can damage bone cells and cause osteoporosis. Osteocytes are the most numerous bone cells and also important target cells for Cd‐induced osteotoxic damage. Autophagy plays important role in the progression of osteoporosis. However, osteocyte autophagy in Cd‐induced bone injury is not well characterized. Thus, we established a Cd‐induced bone injury model in BALB/c mice and a cellular damage model in MLO‐Y4 cells. Aqueous Cd exposure for 16 months showed an increase in plasma alkaline phosphatase (ALP) activity and increase in urine calcium (Ca) and phosphorus (P) concentrations in vivo. Moreover, expression level of autophagy‐related microtubule‐associated protein 1A/1B‐light chain 3 II (LC3II) and autophagy‐related 5 (ATG5) proteins were induced, and the expression of sequestosome‐1 (p62) was reduced, along with Cd‐induced trabecular bone damage. In addition, Cd inhibited the phosphorylation of mammalian target of rapamycin (mTOR), protein kinase B (AKT), and phosphatidylinositol 3‐kinase (PI3K). In vitro, 80 μM Cd concentrations exposure upregulated LC3II protein expression, and downregulated of p62 protein expression. Similarly, we found that treatment with 80 μM Cd resulted in a reduction in the phosphorylation levels of mTOR, AKT, and PI3K. Further experiments revealed that addition of rapamycin, an autophagy inducer, enhanced autophagy and alleviated the Cd‐induced damage to MLO‐Y4 cells. The findings of our study reveal for the first time that Cd causes damage to both bone and osteocytes, as well as induces autophagy in osteocytes and inhibits PI3K/AKT/mTOR signaling, which could be a protective mechanism against Cd‐induced bone injury. [ABSTRACT FROM AUTHOR]

As a novel class of small RNAs, piRNAs are highly expressed in the animal gonads and their main known role is to inhibit transposon activity for ensuring the correctness and integrity of genome. In order to explore the characteristics of piRNAs in sheep testis and their possible regulatory roles on male reproduction, deep sequencing technology was used to sequence small RNAs and identify piRNAs in testes of sheep. The length of piRNAs in sheep testes showed a unimodal distribution between 26 and 31 nt, with a peak at 29 nt. These piRNAs exhibited obvious ping-pong signature and strand specificity. In the genome, they were mainly aligned to CDS, intron, repetitive sequence regions and unannotated regions. Furthermore, in transposon analysis, piRNAs were aligned predominantly to LINE, SINE, and LTR types of retrotransposon in sheep testes, and the piRNAs derived from each type showed obvious pingpong signature. The piRNA clusters identified in sheep testes were mainly distributed on chromosomes 3, 7, 15, 17, 18 and 20. The results combining semen determination with pathway enrichment analysis implied that differentially expressed piRNAs between the testes of rams with different fertility might participate in spermatogenesis by regulating multiple pathways closely related to stabilization of blood-testis barrier and renewal and differentiation of spermatogonial stem cell. Taken together, the study provided new insights into the characteristics, origin and expression patterns of piRNAs in sheep testes tissue, which would help us better understand the role of piRNAs in sheep reproduction. [ABSTRACT FROM AUTHOR]

The mammalian oviduct is functionally highly diverse during the estrus cycle. It provides a suitable milieu for oocyte maturation, sperm capacitation, fertilization, early embryo development and transportation. While there have been many studies of molecular mechanisms on the kidding number of goats, a systematic analysis by which the underlying circular RNAs (circRNAs) changes in the oviduct related to prolificacy traits is lacking. Herein, we present a comprehensive circRNA atlas of the oviduct among high- and low-fecundity goats in the follicular phase (FH vs. FL), luteal phase (LH vs. LL), and estrus cycle (FH vs. LH; FL vs. LL) to unravel their potential regulatory mechanisms in improving kidding number. We generated RNA sequencing data, and identified 4,078 circRNAs from twenty sampled Yunshang black goats. Many of these circRNAs are exon-derived and differentially expressed between each comparison group. Subsequently, eight differentially expressed (DE) circRNAs were validated by RT‒qPCR, which was consistent with the RNA-seq data. GO and KEGG enrichment analyses suggested that numerous host genes of DE circRNAs were involved in the hormone secretion, gamete production, fertilization, and embryo development processes. The competing endogenous RNA (ceRNA) interaction network analysis revealed that 2,673 circRNA–miRNA–mRNA axes (including 15 DE circRNAs, 14 miRNAs, and 1,699 mRNAs) were formed, and several target genes derived from the ceRNA network were associated with oviduct functions and reproduction, including SMAD1, BMPR1B, IGF1, REV1, and BMP2K. Furthermore, miR-15a-5p, miR-181b-5p, miR-23b-5p, miR-204-3p, and miR-145-5p might play important roles in reproduction. Finally, a novel circRNA, circIQCG, was identified as potentially involved in embryo development. Overall, our study provides a resource of circRNAs to understand the oviductal function and its connection to prolificacy trait of goats in the differentiation estrus cycle. [ABSTRACT FROM AUTHOR]

A pivotal clinical trial is often necessary to assess drug efficacy in the intended to use (IU) population. Ideally, patients should be enrolled based on a positive test result from a well-characterized companion diagnostic (CDx). However, the central challenge is that patients are instead recruited on the basis of a clinical trial assay (CTA) result. This challenge arises because, CTA is available at all local labs; the time delay to enable enrollment based on CDx could result in a significant proportion of patients being unable to participate, adversely affecting precision and/or bias. The difficulty is therefore that patients are recruited on the basis that their CTA result is positive (CTA+) but the goal is to assess the drug efficacy in patients positive by the companion diagnostic (CDx+). In this commentary, we will examine an apparent weakness of a variance formula that is proposed in the context of a sensitivity analysis. We will develop an alternative formula, and argue that this should be used instead. [ABSTRACT FROM AUTHOR]

Circular RNAs (circRNAs) are non-coding RNAs newly identified and play important roles in RNA regulation. However, little is known regarding photoperiods induced circRNAs in the thyroid gland. In this study, we performed a comprehensive analysis of circRNA profiles in the thyroid gland of OVX+E2 ewes at different photoperiods by whole transcriptome sequencing. A total of 37,470 novel circRNAs were detected in different photoperiods (42 days of short photoperiod treatment, SP42; 42 days of long photoperiod treatment, LP42; SP42 transfer to LP42, SPLP42), with a total of 817 circRNAs for SP42-LP42 (down: 132; up: 114), LP42-SPLP42 (down: 136; up: 112) and SP42-SPLP42 (down: 182; up: 141) having differentially expressed. Functional enrichment annotation analysis of DE-circRNAs for GO and KEGG by R package, features that influence photoperiod response in Sunite ewes through the Inositol phosphate metabolism, cGMP-PKG signaling pathway, Calcium signaling pathway, MAPK signaling pathway, and Oocyte meiosis. In addition, competitive endogenous RNA (ceRNA) network analysis revealed target binding sites for identified miRNAs in DE-cirRNAs such as oar-miR10b, oar-miR-200c, oar-miR-21, oar-miR-370-3p, oar-miR-377-3p, oar-miR181a, oar-miR-432, and oar-miR-495-3p. These results of this study will provide some new information for understanding circRNA function as well as the changes in the sheep thyroid gland under different photoperiods. [ABSTRACT FROM AUTHOR]

The fat tail/rump is considered as an adaptive selection under harsh challenges which serves as a fat store for the animal. However, the mechanism of fat deposition in tail is unclear. The polymorphisms of candidate sheep CEBPA, PRKAG3 and SREBF1 genes and their relationship with fat deposition between fat-tailed (rumped) and thin-tailed breeds were investigated. Two and one SNPs were identified for PRKAG3 and SREBF1 respectively. Genotyping method was used to analyze genotypes among Altay sheep (fat-rumped breed) and White Suffolk (thin-tailed breed) by Sequenom MassArray. For PRKAG3 gene, a c,1744C>T SNP and a c,1840C>T SNP have been genotyped. For SREBF1 gene, an unknown synonymous c.2878A>G SNP was detected. The genotype distributions in those two loci were significantly different between fat tail and thin tail breeds by chi-square test(P<0.05). [ABSTRACT FROM AUTHOR]

Follicle-stimulating hormone (FSH) is the pituitary hormone that regulates gonadal function by Hypothalamic-Pituitary-Gonadal Axis (HPGA) in vertebrates. FSHR has the function of transmitting follicle-stimulating growth biological information in animal breeding activities. In this study, we analyzed the structure of FSHR gene's promoter, expression regulation and the correlation between polymorphism with fecundity. The results show that it is highly possible that the promoter of FSHR gene is in the region of 2045-2295bp and 2038-2288bp, TATA box was also found there. The expression differences were found in oviduct during every stage of estrous cycle (P<0.05) between Small Tail Han (STH) sheep and Tan (T) sheep. There were significant differences in genotype distribution between STH sheep and T sheep at T1235C and AG2357-2358 CT sites (P<0.01). The results of haplotypes analysis indicated that H2, H7 were preponderant haplotype in high prolificacy sheep breed. The litter size ofTT genotype was significantly higher than that in CC genotype at T1235C loci in STH sheep (P<0.05). The current study provides evidence in sheep for genetic markers that might be used in breeding program. [ABSTRACT FROM AUTHOR]

A new phosphorus containing anhydride (DPO‐MA) curing agent is successfully synthesized via the addition reaction between diphenylphosphinoxid (DPO) and maleic anhydride (MA). It can improve the flame retardancy of epoxy resin (EP) without significantly deteriorating its mechanical properties. Due to the high efficiency of the gas phase flame‐retard mechanism, only 1.0 wt% of phosphorus is sufficient to increase the LOI value to 33.1% and UL‐94 to V0 rating. Cone calorimetry data shows that the peak of heat release decreases more than 42% than that of neat EP during combustion. Besides, the rigid DPO skeleton in the molecular chain can greatly improve the Tg value of the DPO‐MA/EP‐1.0, which is as high as 125°C, much higher than that of neat EP. Thus, this work exhibits a great potential value for EP materials. [ABSTRACT FROM AUTHOR]

The flammability and brittleness are two major shortages of epoxy resin (EP). Normally, adding flame retardants can deteriorate the mechanical property of EP. Herein, in this work, a reactive flame retardant (DPPO‐TES) containing both Si and P was synthesized and incorporated into EP. When the content of P is only 1.0 wt%, the LOI of the modified EP (DPPO‐TES/EP) reaches to 33.2%, UL‐94 passes V0, THR and pk‐HRR are 377.6 kw/m2 and 103.5 MJ/m2, respectively, which is attributed to the high flame‐retardant efficiency in both condense phase and gas phase. In addition, the flexible SiOSi chains in the structure can significantly improve the impact strength of DPPO‐TES/EP. Therefore, as a bifunctional modifier, DPPO‐TES has a wide application prospect for EP materials. [ABSTRACT FROM AUTHOR]

The pituitary is a vital endocrine organ that regulates animal seasonal reproduction by controlling the synthesis and secretion of the hormone. The change of photoperiod is the key factor affecting the function of the pituitary in animals, but the mechanism is unclear. Here, we studied the transcriptomic variation in pars distalis (PD) of the pituitary between short photoperiod (SP) and long photoperiod (LP) using RNA sequencing based on the OVX+E2 sheep. 346 differentially expressed (DE) lncRNAs and 186 DE-mRNA were found in the PD. Moreover, function annotation analysis indicated that the reproductive hormones and photoperiod response-related pathways including aldosterone synthesis and secretion, insulin secretion, thyroid hormone synthesis, and circadian entrainment were enriched. The interaction analysis of mRNA-lncRNA suggested that MSTRG.240648, MSTRG.85500, MSTRG.32448, and MSTRG.304959 targeted CREB3L1 and DUSP6, which may be involved in the photoperiodic regulation of the PD. These findings provide resources for further study on the seasonal reproductive in ewes. [ABSTRACT FROM AUTHOR]

The 2019 new coronavirus epidemic potentially induced by wild animals has drawn tremendous attention. Wild animal meat contamination and adulteration have become increasingly serious, particularly for highly cooked wild animal meats that are difficult to be detected. In this study, a highly specific polyclonal antibody targeting the cooked rat proteins was developed. The corresponding sandwich ELISA (swELISA) was developed and found highly sensitive and specific for cooked rat meat, while there are no cross-reactions to the cooked chicken, pork and beef meats. The limit of detection (LOD) is determined to be as low as 0.01 ug/L based OD values. The coefficient variation (CV) is 5% and 8% for intra and inter assays, respectively. The recovery efficiencies are between 90% and 110%. The sandwich ELISA can detect both raw and cooked rat meat and is also suitable for Swab test of rat contamination. The results indicated a highly reliable and robust ELISA-based assay for cooked rat meat identification and contamination. [ABSTRACT FROM AUTHOR]

A label-free and rapid detection of cooked rat meat-based molecular-imprinted polymer (MIP) conjugated with quantum weak measurement system (QWM) has been developed. Cooked rat protein-specific MIP was prepared by a modified reverse-phase microemulsion method and coated on the internal surface of measuring channels assembled into Mach-Zrhnder interferometer of an optical weak measurement system. The measurement is based on the optical phase shift due to the refractive index change, which is induced by the specific capture of rat protein at its MIP sites. The results can be characterized and calculated by the central wavelength shift of output spectra through weak value amplification. This MIP-WQM system was demonstrated with cooked rat protein showing linear dynamic range from 0 to 20 µg/L. The limit of detection is calculated as 0.1 µg/L for cooked rat meat protein. The system also showed high specificity to cooked rat meat protein, compared to cooked pork meat proteins. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the protective effect of naringin (Nar) on H2O2-induced apoptosis of nucleus pulposus-derived mesenchymal stem cells (NPMSC) and the potential mechanism in this process. Methods: Rat NPMSC were cultured in MSC culture medium or culture medium with different concentrations of H2O2. Nar or the combination of Nar and LY294002 was added into the culture medium to investigate the effects of Nar. Cell viability was evaluated by cell counting kit-8 (CCK-8) assay. The apoptosis rate was determined using Annexin V/PI dual staining and terminal deoxynucleotide transferase-mediated dUTP nick end labeling (TUNEL) assays. Additionally, the levels of reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were analyzed by flow cytometry. ATP level in NPMSC was analyzed via ATP detection kit. Mitochondrial ultrastructure change was observed through transmission electron microscope (TEM). Levels of apoptosis-associated molecules (cleaved caspase-3, Bax and Bcl-2) were evaluated via RT-PCR and western blot, respectively. Results: The cells isolated from NP met the criteria for MSC. H2O2 significantly promoted NPMSC apoptosis in a dose and time-dependent manner. Nar showed no cytotoxicity effect on NPMSC up to a concentration of 100 μM for 24 h. Nar exhibited protective effects against H2O2-induced NPMSC apoptosis including apoptosis rate, expressions of proapoptosis and antiapoptosis related genes and protein. Nar could also alleviate H2O2-induced mitochondrial dysfunction of increased mitochondrial ROS production, reduced MMP, decreased intracellular ATP and mitochondrial ultrastructure change. However, these protected effects were inhibited after LY294002 treatment. Conclusions: Our results demonstrated that Nar efficiently attenuated H2O2-induced NPMSC apoptosis and mitochondrial dysfunction. The activation of ROS-mediated PI3K/Akt pathway may be the potential mechanism in this process. [ABSTRACT FROM AUTHOR]

TAC3 gene is related to the regulation of animal reproductive function. Here we tried to find out the association of the expression and polymorphism of TAC3 gene with the estrus of sheep. TAC3 expression in uterus and pituitary gland increased in estrus of Small Tail Han sheep and Tan sheep. The TAC3 expression level in uterus might relate with estrus of sheep. Two mutations of TAC3 were found in intron 1 (C327T) and exon 2 (G1803A) separately. Although there were significant differences of C327T/G1803A genotype distributions in five different sheep breeds, it did not relate with the year-round estrus. The relationship between these polymorphisms of TAC3 and the litter size of Small Tail Han sheep was not found. [ABSTRACT FROM AUTHOR]

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a facultative intracellular pathogen that damages gastrointestinal tissue and causes severe diarrhoea. The mechanisms by which Salmonella disrupts epithelial barrier and increases the paracellular permeability are incompletely understood. Our present study aims to determine the role of Gli1, a transcription factor activated in the sonic hedgehog (Shh) pathway, in decreasing the levels of apical junction proteins in a Salmonella‐infected human colonic epithelial cancer cell line, Caco‐2, and in the intestinal tissue of Salmonella‐infected mice. Here, we report that S. Typhimurium increased the mRNA and protein levels of Gli1 and Snail, a downstream transcription factor that plays an important role in the epithelial‐to‐mesenchymal transition (EMT). S. Typhimurium also decreased the levels of E‐cadherin and three tight junction proteins (ZO‐1, claudin‐1, and occludin). Gli1 siRNA and GANT61, a Gli1‐specific inhibitor, blocked S. Typhimurium‐induced Snail expression, restored the levels of E‐cadherin and tight junction proteins, and prevented S. Typhimurium‐increased paracellular permeability. Further study showed that Gli1 was cross‐activated by the MAP and PI‐3 kinase pathways. S. Typhimurium devoid of sopB, an effector of the Type 3 secretion system (T3SS) responsible for AKT activation, was unable to induce Snail expression and to decrease the expression of apical junction proteins. Our study uncovered a novel role of Gli1 in mediating the Salmonella‐induced disruption of the intestinal epithelial barrier. [ABSTRACT FROM AUTHOR]

Motivation Single-cell RNA-sequencing (scRNA-seq) has become an important tool to unravel cellular heterogeneity, discover new cell (sub)types, and understand cell development at single-cell resolution. However, one major challenge to scRNA-seq research is the presence of 'drop-out' events, which usually is due to extremely low mRNA input or the stochastic nature of gene expression. In this article, we present a novel single-cell RNA-seq drop-out correction (scDoc) method, imputing drop-out events by borrowing information for the same gene from highly similar cells. Results scDoc is the first method that directly involves drop-out information to accounting for cell-to-cell similarity estimation, which is crucial in scRNA-seq drop-out imputation but has not been appropriately examined. We evaluated the performance of scDoc using both simulated data and real scRNA-seq studies. Results show that scDoc outperforms the existing imputation methods in reference to data visualization, cell subpopulation identification and differential expression detection in scRNA-seq data. Availability and implementation R code is available at https://github.com/anlingUA/scDoc. Supplementary information Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

Background: When conservative treatments do not work, TKA may be the best option for patients with knee osteoarthritis, although a relatively large proportion of individuals do not have clinically important improvement after TKA. Evidence also suggests that women are less likely to benefit from TKA than men, but the reasons are unclear. Widespread pain disproportionately affects women and has been associated with worse outcomes after joint arthroplasty, yet it is unknown if the effect of widespread pain on TKA outcomes differs by patient gender.Questions/purposes: (1) Does the association between widespread pain and no clinically important improvement in osteoarthritis-related pain and disability 2 years after TKA differ between men and women? (2) Does the use of pain medications 2 years after TKA differ between those with widespread pain and those without widespread pain before surgery?Methods: Osteoarthritis Initiative (https://nda.nih.gov/oai/) study participants were followed annually from March 2005 until October 2015. Participants who underwent TKA up to the 7-year follow-up visit with pain/disability assessment at the protocol-planned visit before TKA and at the second planned annual visit after surgery were included in the analysis. Among 4796 study participants, 391 had a confirmed TKA, including 315 with pain/disability assessment at the protocol-planned visit before TKA. Overall, 95% of participants (298) had the required follow-up assessment; 5% (17) did not have follow-up data. Widespread pain was defined based on the modified American College of Rheumatology criteria. Symptoms were assessed using the WOMAC pain (range 0 to 20; higher score, more pain) and disability (range 0 to 68; higher score, more disability) scores, and the Knee Injury and Osteoarthritis Outcome Score for pain (range 0 to 100; higher score, less pain). Improvements in pain and disability were classified based on improvement from established clinically important differences (decrease in WOMAC pain ≥ 1.5; decrease in WOMAC disability ≥ 6.0; increase in Knee Injury and Osteoarthritis Outcome Score for pain ≥ 9). At baseline, more women presented with widespread pain than men (45% [84 of 184] versus 32% [36 of 114]). Probability and the relative risk (RR) of no clinically important improvement were estimated using a logistic regression analysis in which participants with widespread pain and those without were compared. The analyses were done for men and women separately, then adjusted for depression and baseline outcome scores.Results: Among women, preoperative widespread pain was associated with an increased risk of no clinically important improvement 2 years after TKA, based on WOMAC pain scores (13.5% versus 4.6%; RR 2.93 [95% CI 1.18 to 7.30]; p = 0.02) and the Knee Injury and Osteoarthritis Outcome Score for pain (16.5% versus 4.9%; RR 3.39 [95% CI 1.34 to 8.59]; p = 0.02). Given the lower and upper limits of the confidence intervals, our data are compatible with a broad range of disparate associations between widespread pain and lack of clinically important improvement in WOMAC pain scores (RR 0.77 [95% CI 0.22 to 2.70]; p = 0.68) and the Knee Injury and Osteoarthritis Outcome Score for pain (RR 1.37 [95% CI 0.47 to 4.00]; p = 0.57) among men, as well as clinically important improvement in WOMAC disability scores among men (RR 0.72 [95% CI 0.20 to 2.55]; p = 0.61) and women (RR 1.98 [95% CI 0.92 to 4.26]; p = 0.08). Participants presenting with widespread pain before TKA were more likely than those without widespread pain to use medication for symptoms of knee osteoarthritis most days for at least 1 month 2 years after TKA (51% [61 of 120] versus 32% [57 of 178]; mean difference, 18.8 [95% CI 7.3 to 30.1]; p < 0.01).Conclusions: Widespread pain before TKA was associated with an increased risk of no clinically important improvement in knee pain 2 years postoperatively among women. Because of the small number of men with widespread pain in the sample, the results for men were inconclusive. In clinical practice, screening TKA candidates for widespread pain may be useful, and expectations of surgical outcomes may need to be tempered if patients have a concurrent diagnosis of widespread pain. Future studies should include more men with widespread pain and investigate if treatment of widespread pain before or concurrent with TKA surgery may improve surgical outcomes.Level Of Evidence: Level III, therapeutic study. [ABSTRACT FROM AUTHOR]

The expression characteristics of the hypothalamic–pituitary–gonadal (HPG) axis-related candidate genes, DIO2, EYA3, KISS1 and GPR54, were analyzed in year-round estrous rams (small-tail Han sheep, STH) and seasonally estrous rams (Sunite sheep, SNT) using qPCR. The results were as follows: DIO2 was mainly expressed in pituitary, and KISS1 was specifically expressed in hypothalamus in the two groups. However, EYA3 and GPR54 were widely expressed in the cerebrum, cerebellum, hypothalamus, pituitary, testis, epididymis, vas deferens and adrenal gland tissues in both breeds, with significant differences in the cerebellum, hypothalamus, pituitary, testis and vas deferens tissues. We speculated that DIO2 and KISS1 may have positive roles in different regions in ram year-round estrus. Moreover, the expression patterns of EYA3 and GPR54 suggested that they may regulate the estrous mode of ram via testis and vas deferens. This is the first study to systematically analyze the expression patterns of HPG axis-related genes in rams. [ABSTRACT FROM AUTHOR]

Data normalization is vital to single-cell sequencing, addressing limitations presented by low input material and various forms of bias or noise present in the sequencing process. Several such normalization methods exist, some of which rely on spike-in genes, molecules added in known quantities to serve as a basis for a normalization model. Depending on available information and the type of data, some methods may express certain advantages over others. We compare the effectiveness of seven available normalization methods designed specifically for single-cell sequencing using two real data sets containing spike-in genes and one simulation study. Additionally, we test those methods not dependent on spike-in genes using a real data set with three distinct cell-cycle states and a real data set under the 10X Genomics GemCode platform with multiple cell types represented. We demonstrate the differences in effectiveness for the featured methods using visualization and classification assessment and conclude which methods are preferable for normalizing a certain type of data for further downstream analysis, such as classification or differential analysis. The comparison in computational time for all methods is addressed as well. [ABSTRACT FROM AUTHOR]

To elucidate how expression characteristics of BMPR1B, BMP15, and GDF9 are associated with sheep reproduction, tissue-specific mRNA expression of these genes in Small Tail Han (STH) ewes (a polytocous breed) and Sunite (SNT) ewes (a monotocous breed) in the follicular phase were investigated using transcription profiling and quantitative real-time PCR (qPCR). Expression levels of the three genes were all highest in ovaries of the two sheep breeds, and BMPR1B and GDF9 expression in the ovarian tissue was significantly higher in STH sheep compared with SNT sheep (P < 0.01), whereas BMP15 expression in pituitary, ovarian, oviduct, and uterine tissues was significantly lower in STH sheep compared with SNT sheep (P < 0.01). This study revealed that BMPR1B, BMP15, and GDF9 may play important roles in the hypothalamic-pituitary-gonadal axis, especially in the ovary, which may affect sheep prolificacy. BMPR1B and GDF9 expression might increase litter size, whereas BMP15 expression may decrease litter size. These results provide preliminary information regarding molecular regulatory mechanisms underlying sheep prolificacy. [ABSTRACT FROM AUTHOR]

Like most seasonal domesticated species, sheep are short-day breeders, which means that the reproduction axis is activated by short days. The annual photoperiodic cycle affects the amount of daylength information that is transmitted to the hypothalamic-pituitary-gonadal (HPG) axis by regulating pulsatile secretion of gonadotrophinreleasing hormone from the hypothalamus. Kisspeptin, which is encoded by Kiss1, plays a major role in reproductive seasonality. Based on results from our previous Solexa sequencing data obtained from Tan (T) and Small Tail Han (STH) sheep during anoestrus and the breeding season, full-length mRNA information for ovine Kiss1 was obtained; 894 bp in T sheep and 1145 bp in STH sheep. Both encode 135 amino acids. Additionally, T and STH sheep have different transcription start sites of Kiss1. Kiss1 expression during oestrus was significantly higher than that during dioestrus, both in T and STH sheep (P<0.01). We also found a strong relationship between Kiss1 mRNA levels and histone H3 acetylation status in the 5' promoter region of ovine Kiss1. These data indicated that epigenetic modification occurs during reproduction in sheep and this is the first report that histone H3 deacetylation occurs in the hypothalamus of seasonal sheep breeders during the transition from dioestrus to oestrus. [ABSTRACT FROM AUTHOR]

Background: Animal domestication has been extensively studied, but the process of feralization remains poorly understood. Results: Here, we performed whole-genome sequencing of 99 sheep and identified a primary genetic divergence between 2 heterogeneous populations in the Tibetan Plateau, including 1 semi-feral lineage. Selective sweep and candidate gene analysis revealed local adaptations of these sheep associated with sensory perception, muscle strength, eating habit, mating process, and aggressive behavior. In particular, a horn-related gene, RXFP2, showed signs of rapid evolution specifically in the semi-feral breeds. A unique haplotype and repressed horn-related tissue expression of RXFP2 were correlated with higher horn length, as well as spiral and horizontally extended horn shape. Conclusions: Semi-feralization has an extensive impact on diverse phenotypic traits of sheep. By acquiring features like those of their wild ancestors, semi-feral sheep were able to regain fitness while in frequent contact with wild surroundings and rare human interventions. This study provides a new insight into the evolution of domestic animals when human interventions are no longer dominant. [ABSTRACT FROM AUTHOR]

Background: Alpha 1-antitrypsin (α1-AT) may affect the susceptibility of mastitis in dairy cattle for its possible role in the protection of lactoferrin from proteolytic degradation in the mammary. Milk somatic cell score (SCS) is a logarithmic transformation of the milk somatic cell count widely used as an index to evaluate mastitis. To study the relationships of α1-AT gene and SCS in Chinese Holstein cows, methods of PCR-SSCP, DNA sequencing, PCR-RFLP, and CRS-PCR technologies were used to detect single nucleotide polymorphisms sites in α1-AT gene. Results: Two polymorphic sites at G5503A and G5746C of α1-AT gene were found. AA (0.3633), AB (0.4644) and BB (0.1723) genotypes were detected at G5503A site, CC (0.3483), CD (0.4906) and DD (0.1611) genotypes were found at G5746C in Chinese Holstein cows. Least squares mean of SCS for individuals with BB genotype was significantly lower than that with AA and AB genotype (p < 0.01), and that with AB genotype was significantly lower than that with AA (p < 0.05). There was no significant difference among individuals with CC, CD and DD genotypes (p > 0.05). Least squares mean of SCS for individuals with BBDD genotype combination were significantly lower than those with AACC and AACD (p < 0.05). Conclusions: Statistical analysis indicated that B allele and BBDD genotype combination of α1-AT can improve mastitis resistance in dairy cattle. [ABSTRACT FROM AUTHOR]

Objective: To determine if severity of osteoarthritis-related knee pain is associated with a willingness to undergo total knee replacement (TKR) and whether this association is confounded or modified by components of socioeconomic status and health care coverage.Methods: Cross-sectional analysis was conducted among 3,530 Osteoarthritis Initiative study participants. Logistic regression models were used to assess the effect of knee pain severity (where 0 = none, 1-3 = mild, 4-7 = moderate, and 8-10 = severe) on willingness to undergo TKR. Stratified analyses were conducted to evaluate whether socioeconomic status and health care coverage modify the effect of knee pain severity on willingness.Results: Participants with severe knee pain, compared to participants without pain, were less willing to undergo TKR (odds ratio [OR] 0.73, 95% confidence interval [95% CI] 0.57-0.93). This association was attenuated when adjusted for age, sex, comorbidity, depression, health insurance coverage, prescription medicine coverage, health care source, education, income, employment, race, and marital status (adjusted OR 0.92, 95% CI 0.68-1.24). The odds of willingness to undergo TKR were significantly lower in those with the highest level of pain, compared to those without pain, among participants without health insurance (adjusted OR 0.08, 95% CI 0.01-0.56), but not among those with health insurance (adjusted OR 1.03, 95% CI 0.73-1.38), when adjusted for demographic, clinical, health care access, and socioeconomic factors (P = 0.015). However, <5% of participants were without health insurance.Conclusion: Among participants without health insurance, severe knee pain was paradoxically associated with less willingness to undergo TKR. Policies that improve access to quality health care may affect patient preferences and increase utilization of TKR surgery among vulnerable populations. [ABSTRACT FROM AUTHOR]

DIO2 was studied as a candidate gene for reproduction in sheep. Three pairs of primers were designed to detect single nucleotide polymorphisms (SNPs) of exon 2 of DIO2 in nonseasonal and seasonal reproduction breeds. The polymorphisms were detected using SeqMan II and analyzed by PCR-RFLP. The sheep DIO2 gene was cloned and its mutations were detected in five sheep breeds whose reproductive seasonality and litter sizes were different. Association analyses were performed to reveal the relationships between SNPs and reproductive seasonality as well as litter size in sheep. The results showed that a 1041-bp DNA fragment of the sheep DIO2 gene was obtained and a part of the coding sequence of 591 bp encodes 196 amino acids, having high homology with that of other mammals. Moreover, three mutations (G318T, G482C, and C885T) were detected. The test of genotype distribution in five sheep breeds showed that there were significant differences between nonseasonal and seasonal breeds for 318 and 482 loci. However, the associations between the three polymorphisms and sheep seasonal reproduction were not very distinct. Additionally, there was no significant associations between polymorphisms and litter sizes in Small Tail Han sheep. These results may provide the basic theory for further study. [ABSTRACT FROM AUTHOR]

The aims of study were to reveal the changes in genetic diversity in reproduction selection of sheep. Genetic polymorphisms of microsatellite GC101 locus and its linkage with the FecB gene were detected in Chinese local sheep (Small Tail Han and Hu sheep) and western sheep (Texel and Dorset) breeds. There was the same FecB mutation (A746G) in both Small Tail Han and Hu sheep as that in Booroola Merino ewes, but not in Texel and Dorset sheep. Seven alleles, from 192 bp to 210 bp, and 20 genotypes were detected at the GC101 locus in 447 individuals totally. The preponderant allele was 200 bp for Chinese sheep and 202 bp for western sheep. Linkage analysis showed that there was strong linkage disequilibrium between the 200-bp allele of the GC101 locus and the B allele of the FecB gene (r² = 0.639). These results preliminarily implied that both the 200-bp allele of the GC101 locus and the B allele of FecB had been reinforced in reproduction selection of Chinese sheep. The differentiation of GC101 allele distribution between Chinese sheep and western sheep also indicated that the FecB of Chinese sheep might be derived from Chinese local areas. [ABSTRACT FROM AUTHOR]

In our early work, one mutation (A898G, which caused Ser32Gly (S32G) change of mature peptide) was identified in the goat BMP15 gene and the allele G was associated with high litter size in Jining Grey goats (the goat breed with the highest fecundity in China). The aim of this research was to investigate the genetic structure of the prolific BMP15 gene in 18 goat breeds reared in China, including 12 native breeds, 3 introduced breeds, and 2 cultivated breeds. Genomic DNA of 1011 goats was screened for the S32G mutation. The results showed that this mutation existed in all 3 cultivated and 8 native goat breeds but in none of the 3 introduced goat breeds, which hinted that this mutation may originate from native goat breeds of China. Besides the Jining Grey goats, the BMP15 gene was also a potential prolificacy gene in Matou goats. Moreover, the structure prediction indicated that the S32G mutation might participate in the binding of BMP15 with receptors. [ABSTRACT FROM AUTHOR]

Background: Seasonal estrus is a critical limiting factor of animal fecundity, and it involves changes in both ovarian biology and hormone secretion in different seasons. Previous studies indicate that two classes of small RNAs (miRNAs and piRNAs) play important regulatory roles in ovarian biology. To understand the roles of small RNA-mediated post-transcriptional regulation in ovine seasonal estrus, the variation in expression patterns of ovarian small RNAs during anestrus and the breeding season were analyzed using Solexa sequencing technology. In addition, reproductive hormone levels were determined during ovine anestrus and the breeding season. Results: A total of 483 miRNAs (including 97 known, 369 conserved and 17 predicated novel miRNAs), which belong to 183 different miRNA families, were identified in ovaries of Tan sheep and Small Tail Han (STH) sheep. Compared with the three stages of the breeding season, 25 shared significantly differentially expressed (including 19 up- and six down-regulated) miRNAs were identified in ovine anestrus. KEGG Pathway analysis revealed that the target genes for some of the differentially expressed miRNAs were involved in reproductive hormone related pathways (e.g. steroid biosynthesis, androgen and estrogen metabolism and GnRH signaling pathway) as well as follicular/luteal development related pathways. Moreover, the expression of the differentially expressed miRNAs and most of their target genes were negatively correlated in the above pathways. Furthermore, the levels of estrogen, progesterone and LH in ovine anestrus were significantly lower than those in the breeding season. Combining the results of pathway enrichment analysis, expression of target genes and hormone measurement, we suggest that these differentially expressed miRNAs in anestrus might participate in attenuation of ovarian activity by regulating the above pathways. Besides miRNAs, a large and unexpectedly diverse set of piRNAs were also identified. Conclusions: The miRNA profiles of ovine ovaries in anestrus were presented for the first time. The identification and characterization of miRNAs that are differentially expressed between ovine anestrus and the breeding season will help understanding of the role of miRNAs in the regulation of seasonal estrus, and provides candidates for determining miRNAs which could be potentially used to regulate ovine seasonal estrus. [ABSTRACT FROM AUTHOR]

Single nucleotide polymorphisms of the 5' regulatory region of the FSHR gene were detected in 3 goat breeds (Jining Grey, Boer, and Inner Mongolia Cashmere) by the polymerase chain reaction-single-strand conformation polymorphism method, and its effects on litter size in Jining Grey goats were also evaluated. Concerning primer P1, 2 genotypes (AA and BB) were detected in Boer goats, while only the AA genotype was identified in Jining Grey and Inner Mongolia Cashmere goats. Sequencing revealed a T→A transversion at the 26th position and an A→C transversion at the 61st position of the amplified region in genotype BB in comparison to genotype AA. Regarding primer P2, 3 genotypes (CC, CD, and DD) were identified in the 3 goat breeds mentioned above. Sequencing revealed a T→A transversion at the 70th position and a G→C transversion at the 130th position of the amplified region in genotype DD compared with CC. Does with genotype CC had 0.46 (P < 0.05) or 1.03 (P < 0.05) kids more than those with genotype CD or DD, respectively, while does with CD had 0.57 (P < 0.05) kids more than those with DD in Jining Grey goats. These results preliminarily showed that allele C of the FSHR gene was a potential marker for improving litter size in goats. [ABSTRACT FROM AUTHOR]

The polymorphisms of arylalkylamine-N-acetyltransferase (AA-NAT) gene in high-prolificacy Jining Grey goat, medium-prolificacy Boer goat and low-prolificacy Liaoning Cashmere, Inner Mongolia Cashmere and Angora goats were detected to analyse their relationships with litter size. Primers (P1-P5) were designed to detect the polymorphisms by PCR-SSCP and PCR-RFLP. For P2, AA, AB and BB genotypes were detected. Sequencing revealed one silent mutation (T132C) of AA-NAT gene in BB in comparison to AA. For P3, CC and CD genotypes were detected and sequencing revealed one mutation (C265T) of AA-NAT gene in CD in comparison to CC, and this mutation resulted in an amino acid change of Arg→Cys (R89C). The Jining Grey does with genotype CD delivered by 0.56 kids (P<0.05) more than those with CC genotype. For P5, EE, EF and FF genotypes were detected and sequencing revealed one mutation (C586T) of AA-NAT gene in FF in comparison to EE. This mutation caused an amino acid change of Arg→Trp (R196W). For both P2 and P5, the differences in litter size among three genotypes were not significant in Jining Grey goats (P>0.05). These results preliminarily indicate that allele D at the C265T locus of AA-NAT gene is a potential marker in genetic improvement of litter size in goats. [ABSTRACT FROM AUTHOR]

The first description is presented of nucleotide sequence of exon 2 of caprine melatonin receptor 1b (MT2). No polymorphisms of MT2 gene were detected between high fertility and year-round estrous goat breeds and low fertility and seasonal estrous goat breeds. It is likely that exon 2 of MT2 gene is not associated with fertility or reproductive seasonality in goat breeds. The nucleotide sequence of exon 2 of MT2 gene of Jining Grey goats shows much closer phylogenetic relation to the MT2 of sheep (97%) and cattle (94%) than to that of pig (84%), human (80%) and mouse (74%). A rather high nucleotide identity (62-64%) with the melatonin receptor 1a (MT1) of goat, sheep, human and mouse was also found. The caprine MT2 contains the same NAXXY motif in transmembrane 7 as the other melatonin receptors. Both DRY and CYVCR motifs were detected just downstream from its third transmembrane domain (the same as in sheep and cattle) rather than NRY and CYICH found in other melatonin receptor groups. [ABSTRACT FROM AUTHOR]

The bone morphogenetic protein receptor IB (BMPR-IB) was studied as a candidate gene for the prolificacy of sheep. Nine pairs of primers (P1-P9) were designed to detect single nucleotide polymorphisms (SNPs) of exons 1-4 and 6-10 of the BMPR- IB gene in both high (Small Tail Han and Hu sheep) and low prolificacy breeds (Texel and Chinese Merino sheep) by polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP). Only the products amplified by primers P2, P5, P6, P7, P8 and P9 displayed polymorphisms. The present study identified 22 SNPs in partial coding regions of ovine BMPR- IB, in which 20 SNPs were reported for the first time. In total of the 22 mutations, 18 DNA variations were originated from the Hu breed, three were found in the Small Tail Han breed (two of them were found in other sheep breeds), three in the Chinese Merino breed, and none in the Texel breed. These results preliminarily demonstrated that BMPR-IB is a major gene affecting the hyperprolificacy in Small Tail Han and Hu sheep, and could be used as a molecular genetic marker for early auxiliary selection for hyperprolificacy in sheep. [ABSTRACT FROM AUTHOR]