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(1) Background: Many studies have shown that microgravity experienced by astronauts or long-term bedridden patients results in increased oxidative stress and bone loss. Low-molecular-weight chondroitin sulfates (LMWCSs) prepared from intact chondroitin sulfate (CS) have been demonstrated to possess good antioxidant and osteogenic activities in vitro. This study aimed to assess the antioxidant activity of the LMWCSs in vivo and evaluate their potential in preventing microgravity-induced bone loss. (2) Methods: we used hind limb suspension (HLS) mice to simulate microgravity in vivo. We investigated the effects of LMWCSs against oxidative stress damage and bone loss in HLS mice and compared the findings with those of CS and a non-treatment group. (3) Results: LMWCSs reduced the HLS-induced oxidative stress level, prevented HLS-induced alterations in bone microstructure and mechanical strength, and reversed changes in bone metabolism indicators in HLS mice. Additionally, LMWCSs downregulated the mRNA expression levels of antioxidant enzyme- and osteogenic-related genes in HLS mice. The results showed that overall effect of LMWCSs was better than that of CS. (4) Conclusions: LMWCSs protect against the bone loss caused by simulated microgravity, which may be related to their ability to reduce oxidative stress. LMWCSs can be envisaged as potential antioxidants and bone loss protective agents in microgravity.

Abstract Background Litter size is an important index of mammalian prolificacy and is determined by the ovulation rate. The ovary is a crucial organ for mammalian reproduction and is associated with follicular development, maturation and ovulation. However, prolificacy is influenced by multiple factors, and its molecular regulation in the follicular phase remains unclear. Methods Ten female goats with no significant differences in age and weight were randomly selected and divided into either the high-yielding group (n = 5, HF) or the low-yielding group (n = 5, LF). Ovarian tissues were collected from goats in the follicular phase and used to construct mRNA and miRNA sequencing libraries to analyze transcriptomic variation between high- and low-yield Yunshang black goats. Furthermore, integrated analysis of the differentially expressed (DE) miRNA-mRNA pairs was performed based on their correlation. The STRING database was used to construct a PPI network of the DEGs. RT–qPCR was used to validate the results of the predicted miRNA-mRNA pairs. Luciferase analysis and CCK-8 assay were used to detect the function of the miRNA-mRNA pairs and the proliferation of goat granulosa cells (GCs). Results A total of 43,779 known transcripts, 23,067 novel transcripts, 424 known miRNAs and 656 novel miRNAs were identified by RNA-seq in the ovaries from both groups. Through correlation analysis of the miRNA and mRNA expression profiles, 263 negatively correlated miRNA-mRNA pairs were identified in the LF vs. HF comparison. Annotation analysis of the DE miRNA-mRNA pairs identified targets related to biological processes such as “estrogen receptor binding (GO:0030331)”, “oogenesis (GO:0048477)”, “ovulation cycle process (GO:0022602)” and “ovarian follicle development (GO:0001541)”. Subsequently, five KEGG pathways (oocyte meiosis, progesterone-mediated oocyte maturation, GnRH signaling pathway, Notch signaling pathway and TGF-β signaling pathway) were identified in the interaction network related to follicular development, and a PPI network was also constructed. In the network, we found that CDK12, FAM91A1, PGS1, SERTM1, SPAG5, SYNE1, TMEM14A, WNT4, and CAMK2G were the key nodes, all of which were targets of the DE miRNAs. The PPI analysis showed that there was a clear interaction among the CAMK2G, SERTM1, TMEM14A, CDK12, SYNE1 and WNT4 genes. In addition, dual luciferase reporter and CCK-8 assays confirmed that miR-1271-3p suppressed the proliferation of GCs by inhibiting the expression of TXLNA. Conclusions These results increase the understanding of the molecular mechanisms underlying goat prolificacy. These results also provide a basis for studying interactions between genes and miRNAs, as well as the functions of the pathways in ovarian tissues involved in goat prolificacy in the follicular phase.

AbstractUnusual rainfall is the primary cause of the failure of the tailing dams, and overtopping is the most representative model of the tailing dam failure. The upstream tailing dam was selected as the research object to study the whole process of breach extension and the overtopping dam-failure mechanism under the full-scale rainfall condition. The results showed that the significant size grading phenomenon in the front, middle, and end of the tailing pond was obvious due to the flow separation effect, and its average particle diameter was D50. At different moments of rainfall, the height of the infiltration line at different positions of the dam body was different; at the rainfall of 3600 s, the height of the infiltration line lagged behind the height of the tailing pond, and this phenomenon from the tail of pond to the outside of the dam slope became more obvious. After the rainfall of 3600 s, the height of the infiltration line lagging behind the water level in the pond basically disappeared, and the rate of infiltration line rise kept pace with the rate of water level. The process of overtopping dam-failure experienced dam overtopping (gully erosion), formation of a multistepped small “scarp,” breach rapid expansion, formation of large “scarp,” and burst (fan-shaped formation). The width and depth of the breach showed a positive correlation, and the widening rate of the breach was 3 to 8 times of the deepening rate, especially in the middle of the dam break, widening behavior occupied the dominant factor. The shape of the dam body after failure was parabolic, and the dam body had obvious elevation changes. These results provide the theoretical guidance and engineering application value for improving the theory and early warning model of the upstream tailing dam.

IGF1, a member of the insulin-like growth factor (IGF) superfamily, is also known as the growth-promoting factor (somatomedin C). IGF1 is involved in vertebrate growth and development, immunity, cell metabolism, reproduction, and breeding. However, there are relatively few studies on the relationship between IGF1 and goat reproduction. In this study, a new transcription factor SP1 bound to the IGF1 g. 64943050T>C promoted granulosa cell (GC) proliferation. A mutation g.64943050T>C located in the promoter region of IGF1 was identified. Association analysis revealed that the kidding number in the first and second litters and the average number of first three litters of the CC genotype (2.206 ± 0.044, 2.254 ± 0.056, and 2.251 ± 0.031) were significantly higher than those in the TC genotype (1.832 ± 0.049, 1.982 ± 0.06, and 1.921 ± 0.034) and TT genotype (1.860 ± 0.090, 1.968 ± 0.117, and 1.924 ± 0.062) (p < 0.05). The kidding number in the third litter of the CC genotype (2.355 ± 0.057) was significantly higher than that in the TT genotype (2.000 ± 0.107) (p < 0.05). Then, the function of this mutation was validated by the dual-luciferase reporter assay and EMSA. The results showed that the luciferase activity of IGF1-mutant-C was significantly higher than that of IGF1-Wild-T (p < 0.05). The EMSA also showed that the binding ability of IGF1-mutant-C was higher than that of IGF1-Wild-T (p < 0.05). Subsequently, the transcription factor SP1 was predicted to bind to the mutation of IGF1 (g.64943050T>C). Overexpression of SP1 promotes the expression of IGF1 in the primary granulosa cells (GCs). The results of the CCK-8 assay and the expression of GC proliferation factors (CDK4, cyclin D1, and cyclin D2) demonstrated that SP1 promoted GC proliferation by regulating IGF1 expression. Our results suggested that the IGF1 g.64943050T>C was significantly associated with the kidding number of Yunshang black goats, and SP1 as a transcription factor of IGF1 binding to the mutation T>C regulated the expression of IGF1. Furthermore, SP1 promoted goat GC proliferation by regulating the expression of IGF1, which provides a new insight for the goat fertility trait.

The granulosa cell (GC) is the basic functional unit of follicles, and it is important for promoting follicle growth and sex hormones, as well as growth factor secretion in the process of reproduction. A variety of factors influence granulocyte proliferation, yet there are still many gaps to be filled in target and non-coding RNA regulation. In our study, the differentially expressed (DE) mRNAs and miRNAs were detected by using RNA-seq, and we constructed a mRNA–miRNA network related to goat prolificacy. Then, the goat primary GCs were isolated from the follicle for the function validation of candidate genes and their regulator miRNAs. A total of 2,968 DE mRNAs and 99 DE miRNAs were identified in the high- and low-prolificacy goat by RNA-seq, of which there were 1,553 upregulated and 1,415 downregulated mRNAs, and 80 upregulated and 19 downregulated miRNAs, respectively. JAK3 was identified as highly expressed in the low-prolificacy goats (3 times higher than high-prolificacy goats), and the integrated analysis showed that chi-miR-493-3p was a potential regulator of JAK3. The analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) showed that JAK3 was involved in the PI3K-Akt signaling pathway, the Jak-STAT signaling pathway, and signaling pathways regulating pluripotency of stem cells. In particular, the PI3K-Akt signaling pathway was a typical pathway for cell proliferation, differentiation, apoptosis, and migration. We found that the chi-miR-493-3p targets JAK3 directly via RT-qPCR, dual fluorescence assays, and Western blot. Furthermore, the expression of JAK3 was significantly decreased by the chi-miR-493-3p mimic and increased by the chi-miR-493-3p inhibitor. The CCK-8 assay showed that overexpression of JAK3 promoted cell proliferation, while inhibiting JAK3 had the opposite effect. The expression of cell proliferation markers CDK4 and cyclin D2 also showed the same results. Moreover, the enzyme-linked immunosorbent assay showed that steroid hormones E2 and PROG were increased by overexpressing JAK3 and decreased by inhibiting JAK3. Therefore, our results identified a chi-miR-439-3p-JAK3 regulatory pathway, which provided a new insight into the GC proliferation and prolificacy of goat.

To solve the problems of image threshold segmentation based on weak continuous constraint theory, the running time is long, and the two parameters need to be selected manually, and therefore a fast single-parameter energy function thresholding for image segmentation based on region information (FSEFTISRI) is proposed in this paper. The proposed FSEFTISRI algorithm uses simple linear iterative clustering (SLIC) technology to pre-block the image, extract the image super-pixels, and then map the image super-pixels to the interval type-2 fuzzy set (IT2FS), so as to construct the single-parameter energy function to search the optimal threshold, and adaptively select the penalty parameters in the energy function through the class uncertainty theory. On a non-destructive testing (NDT) database and Berkeley segmentation datasets and benchmarks (BSDS), the proposed FSEFTISRI is compared with five related algorithms. The average misclassification error (ME) of the proposed FSEFTISRI algorithm on NDT and BSDS are 0.0466 and 0.0039, respectively. The results show that the proposed FSEFTISRI has acquired more satisfactory results in visual effect and evaluation index, and the running time of the proposed FSEFTISRI algorithm is shorter, which shows the effectiveness of the proposed FSEFTISRI.

The pituitary gland is an important organ. It is a complex area of the brain involved in endocrine function and reproductive regulation. However, the function of the pituitary in goat reproduction is still unclear. Herein, RNA sequencing was used to explore the expression patterns of circle RNAs (circRNAs) in the pituitary of Yunshang black goats during the various estrus phases. Then the host genes of the circRNAs were predicted, and a competing endogenous RNA (ceRNA) network was constructed. The results showed a total of 6,705 circRNAs in the pituitary of Yunshang black goats, among which 388 differentially expressed (DE) circRNAs (214 were upregulated, while 174 were downregulated) were identified between high- and low-yield Yunshang black goats in the follicular phase (HF vs. LF); moreover, 361 DE circRNAs (136 were upregulated, while 225 were downregulated) were identified between high- and low-yield Yunshang black goats in the luteal phase (HL vs. LL). There were 65 DE circRNAs targeting 40 miRNAs in the HF vs. LF comparison and 46 DE circRNAs targeting 31 miRNAs in the HL vs. LL comparison. We identified chi_circ_0030920, chi_circ_0043017, chi_circ_0008353, chi_circ_0041580, and chi_circ_0016478 as the key circRNAs through functional enrichment analysis. The ceRNA network analysis showed that chi_circ_0031209 and chi_circ_0019448 might play an important role in reproduction by influencing the expression of prolactin receptor (PRLR) in high- and low-yielding goats during the luteal phase, whereas chi_circ_0014542 regulates the expression of WNT5A during the follicular phase. Our study provided the overall expression profiles of circRNAs in the goat pituitary during the estrus phase, which provides new insight into the mechanism of high-yield goats, which can be helpful to guide goat breeding.

Granulosa cell (GC) proliferation provides essential conditions for ovulation in animals. A previous study showed that DENND1A plays a significant role in polycystic ovary syndrome. However, the modulation of DENND1A in GCs remains unclear. Our previous integrated analysis of miRNA–mRNA revealed that the 3'-untranslated region of DENND1A could be a target of chi-miR-324-3p. In this study, we used quantitative reverse transcription polymerase chain reaction (RT-qPCR) to investigate DENND1A expression in ovarian tissues of high- and low-yielding goats. Furthermore, dual-fluorescent reporter vector experiments, Cell Counting Kit-8 (CCK-8) assay, and RT-qPCR were used to elucidate the regulatory pathway of chi-miR-324-3p-DENND1A in GCs. The results revealed an opposite tendency between the expressions of chi-miR-324-3p and DENND1A in the ovaries of high- and low-yielding goats. The CCK-8 assay indicated that chi-miR-324-3p overexpression significantly suppressed GC proliferation, whereas chi-miR-324-3p inhibition promoted GC proliferation. In addition, the expressions of GC proliferation markers LHR, Cylin D2, and CDK4 showed the same tendency. The dual-fluorescent reporter assay revealed that chi-miR-324-3p directly targeted DENND1A, and the RT-qPCR results revealed that DENND1A expression was inhibited by chi-miR-324-3p. In summary, chi-miR-324-3p inhibited the proliferation of GCs by targeting DENND1A.

Pefloxacin (PFLX) is a type of fluoroquinolone (FQ) that has been widely used in aquaculture and animal husbandry. It is also a new kind of pseudo-persistent pollutant, which has some toxic effects. In this study, zebrafish (Danio rerio) embryos were used as exposure models to analyze the toxicity of PFLX. The results showed that PFLX was absorbed on the outer membrane surface at the beginning, and then a few PFLX slowly entered the cytoplasm. The adsorption complied with the Langmuir isotherm adsorption model. The surface morphology of the embryonic membrane was changed. PFLX exposure had no effect on fetal autonomous movement, but it significantly inhibited the embryo hatching rate. After PFLX exposure, the embryo and larval survival rates showed a clear dose–effect relationship. Though PFLX exposure did not cause teratogenic toxicity, it still affected cardiovascular function. The potential toxicity mechanism and developmental toxicity of PFLX to zebrafish were revealed. These results could be important for studies concerning FQ toxicology.

Screening for candidate genes and genetic variants associated with litter size is important for goat breeding. The aim of this study was to analyze the relationship between single nucleotide polymorphisms (SNPs) in PPP2R5C and SLC39A5 and litter size in Yunshang black goats. KASP genotyping was used to detect the SNP genetic markers in the PPP2R5C and SLC39A5 in a population of 569 Yunshang black goats. The results show that there were two SNPs in the PPP2R5C and SLC39A5 promoter regions. Association analysis revealed that the polymorphisms PPP2R5C g.65977743C>T and SLC39A5 g.50676693T>C were significantly associated with the litter size of the third parity of Yunshang black goats (p < 0.05). To further explore the regulatory mechanism of the two genes, the expression of different genotypes of PPP2R5C and SLC39A5 was validated by RT-qPCR and Western blotting. The expression of PPP2R5C was significantly higher in individuals with the TT genotype than in those with the TC and CC genotypes (p < 0.05). The expression of SLC39A5 was also significantly higher in individuals with the TT genotype than in TC and CC genotypes (p < 0.05). Dual luciferase reporter analysis showed that the luciferase activity of PPP2R5C-C variant was significantly higher than that of PPP2R5C-T variant (p < 0.05). The luciferase activity of SLC39A5-T variant was significantly higher than that of SLC39A5-C variant (p < 0.05). Software was used to predict the binding of transcription factors to the polymorphic sites, and the results show that SOX18, ZNF418, and ZNF667 and NKX2-4 and TBX6 might bind to PPP2R5C g.65977743C>T and SLC39A5 g.50676693T>C, respectively. These results provide new insights into the identification of candidate genes for marker-assisted selection (MAS) in goats.

Retinol-binding protein 4 (RBP4), a member of the lipocalin family, is a specific carrier of retinol (vitamin A) in the blood. Numerous studies have shown that RBP4 plays an important role in mammalian embryonic development and that mutations in RBP4 can be used for the marker-assisted selection of animal reproductive traits. However, there are few studies on the regulation of reproduction and high-prolificacy traits by RBP4 in goats. In this study, the 5′ flanking sequence of RBP4 was amplified, and a G>C polymorphism in the promoter region -211 bp (g.36491960) was detected. An association analysis revealed that the respective first, second and third kidding number and mean kidding number of nanny goats with CC and GC genotypes (2.167 ± 0.085, 2.341 ± 0.104, 2.529 ± 0.107 and 2.189 ± 0.070 for CC and 2.052 ± 0.047, 2.206 ± 0.057, 2.341 ± 0.056 and 2.160 ± 0.039 for GC) were significantly higher (p < 0.05) than those with the GG genotype (1.893 ± 0.051, 2.027 ± 0.064, 2.107 ± 0.061 and 1.74 ± 0.05). The luciferase assay showed that luciferase activity was increased in C allele individuals compared with that in G allele individuals. A competitive electrophoretic mobility shift assay (EMSA) showed that individuals with the CC genotype had a stronger promoter region binding capacity than those with the GG genotype. In addition, transcription factor prediction software showed that the RBP4 g.36491960G>C mutation added a novel binding site for transcription factor DP-1 (TFDP1). RT–qPCR results showed that the expression of TFDP1 was significantly higher in the high-prolificacy group than in the low-prolificacy group, and the expression of RBP4 was higher in both the CC and GC genotypes than that in the GG genotype. TFDP1 overexpression significantly increased the expression of RBP4 mRNA (p < 0.05) and the expression of the cell proliferation factors cyclin-D1, cyclin-D2 and CDK4 (p < 0.05). The opposite trend was observed after interference with TFDP1. Both the EdU and CCK-8 results showed that TFDP1 expression could regulate the proliferation of goat ovarian granulosa cells. In summary, our results showed that RBP4 g.36491960G>C was significantly associated with fecundity traits in goats. The g.36491960G>C mutation enhanced the transcriptional activity of RBP4 and increased the expression of RBP4, thus improving the fertility of Yunshang black goats.

Due to its essential use as a fertilizer, ammonia synthesis from nitrogen and hydrogen is considered to be one of the most important chemical processes of the last 100 years. Since then, an enormous amount of work has been undertaken to investigate and develop effective catalysts for this process. Although the catalytic synthesis of ammonia has been extensively studied in the last century, many new catalysts are still currently being developed to reduce the operating temperature and pressure of the process and to improve the conversion of reactants to ammonia. New catalysts for the Haber–Bosch process are the key to achieving green ammonia production in the foreseeable future. Herein, the history of ammonia synthesis catalyst development is briefly described as well as recent progress in catalyst development with the aim of building an overview of the current state of ammonia synthesis catalysts for the Haber–Bosch process. The new emerging ammonia synthesis catalysts, including electride, hydride, amide, perovskite oxide hydride/oxynitride hydride, nitride, and oxide promoted metals such as Fe, Co, and Ni, are promising alternatives to the conventional fused‐Fe and promoted‐Ru catalysts for existing ammonia synthesis plants and future distributed green ammonia synthesis based on the Haber–Bosch process.

Background: MicroRNA-133b (miR-133b) has been shown to play a critical role in spinal cord regeneration. The aim of this study was to investigate the cellular role of miR-133b in neural cells. Methods: PC12 cells and primary cortical neurons (PCNs) were transfected with lenti-miR-133b, lenti-miR-133b inhibitor, plasmid-shRNA-RhoA, plasmid-RhoA and their negative controls. After 48 hours of transfection, the levels of proteins and mRNA or miRNA were evaluated by Western blotting and qRT-PCR, respectively. Moreover, the neurite outgrowth was analyzed by Image J. For pharmacological experiments, inhibitors of MEK1/2 kinase (PD98059), phosphoinositide-3 kinase (PI3K) (LY294002) and ROCK (Y27632) were added into the culture medium. Results: Overexpression of miR-133b in PC12 cells enhanced neurite outgrowth. Conversely, inhibition of miR-133b reduced neurite length. We further identified RhoA as a target and mediator of mir-133b for neurite extension by Western blot and knockdown experiment. Moreover, overexpression of RhoA could attenuate the neurite growth effects of miR-133b. Also, we observed that miR-133b activated MEK/ERK and PI3K/Akt signaling pathway by targeting RhoA. Finally, in PCNs, miR-133b also increased axon growth and attenuated axon growth restrictions from chondroitin sulfate proteoglycans (CSPG). Conclusions: In summary, our study suggested that miR-133b regulated neurite outgrowth via ERK1/2 and PI3K/Akt signaling pathway by RhoA suppression.

Objective: This study aimed to explore the best parameters of buckwheat noodle composite preformed powder. Methods: Single factor experiment, multi-index comprehensive evaluation method and orthogonal experiment were used to optimize the special composite preformed powder recipe of buckwheat noodles. Results: The best parameters of buckwheat noodle composite preformed powder were 45% of buckwheat flour, 8% of modified starch, 1.2% of compound phosphate and 3% of gluten powder. In this situation, the tensile area of composite flour dough was 3 378.6 g·mm; the stabilization time was 7.23 min, the content of wet gluten was 23.71%; the sensory score was 89.2; the comprehensive score of multiple indexes was 0.926 5. Compared to the groups of single factor test and orthogonal test, the quality of composite powder in each test group was significantly improved (P＜0.05), and its application in non-fried instant noodles had a good effect. Conclusion: The optimized composite powder can be used as the raw powder of various kinds of noodles, such as buckwheat coarse grain noodles, semi-dry noodles and new-type non-fried coarse cake.

Herein, a novel fluorescence probe Fla-DNP based on flavonol has been designed and synthesized for rapid, specific detection of H2S. With the addition of H2S, Fla-DNP triggered thiolysis and released Fla displaying the "turn-on" fluorescence response at 566 nm, which is consistent with the reaction site predicted by calculating Electrostatic potential and ADCH charges. As an easily available H2S probe, Fla-DNP has the advantages of high selectivity, anti-interference, low detection limit (0.834 μM), short response time (6 min), and large Stokes shift (124 nm). The sensing mechanism of H2S was determined by HRMS analysis and DFT calculation. Moreover, Fla-DNP processes a wide range of multiple applications, including the detection of H2S in environmental water samples with good recovery rates ranging from 89.6% to 102.0%, as well as tracking the production of H2S during food spoilage. Meanwhile, the probe exhibits superior biocompatibility and can not only be available used for H2S detection in living cells but be further designed as an H2S-activated CO photoreleaser, based on which it can be developed as a targeted anti-cancer drug. A novel fluorescence probe Fla-DNP was synthesized utilizing 4-dimethylaminobenzoxanthone fluorescent dye (Fla) as the fluorophore, 2, 4-dinitrobenzenether group (DNP) as the recognition group, which can rapidly respond to H2S with high selectivity, anti-interference, low detection limit (0.834 μM), short response time (6 min), and large Stokes shift (124 nm) characteristics. The practical applications of Fla-DNP were further explored in water, foodstuffs samples and living cells. It is reflected that Fla-DNP can not only track H2S in complex environment water, but also can detect H2S produced during foodstuffs spoilage to monitor food freshness. More importantly, Fla-DNP can be available used for H2S detection in living cells and utilize the properties of the photoinduced release of CO from flavonols to be designed as a bifunctional platform for H2S detection and CO release. It is demonstrated that H2S-activated CO photoreleaser Fla-DNP has promise for development as an anti-cancer drug. [ABSTRACT FROM AUTHOR]