CircSSBP2 acts as a MiR-2400 sponge to promote intramuscular preadipocyte proliferation by regulating NDRG1.

Intramuscular fat (IMF) is a critical factor in beef quality. IMF is mainly distributed between muscle fibres and its accumulation can affect the marbling and meat quality of beef. IMF formation and deposition is a complex process and in recent years a group of non-coding RNAs (ncRNAs), known as circRNAs, have been discovered to play an important role in regulating intramuscular fat deposition. CircRNAs form a covalent loop structure after reverse splicing of precursor mRNAs. They can act by adsorbing miRNAs, thereby reducing their repressive effects on downstream target genes. Based on high-throughput sequencing of circRNAs in intramuscular fat of Qinchuan and Japanese black cattle, we identified a novel circSSBP2 that is differentially expressed between the two species and associated with adipogenesis. We show that circSSBP2 knockdown promotes bovine intramuscular preadipocyte proliferation, whereas overexpression inhibits bovine intramuscular preadipocyte proliferation. We also show that circSSBP2 can act as a molecular sponge for miR-2400 and that miR-2400 overexpression promotes bovine intramuscular preadipocyte proliferation. Furthermore, N-myc downstream-regulated gene 1 (NDRG1) was identified as a direct target gene of miR-2400, and NDRG1 interference promoted the proliferation of bovine intramuscular preadipocytes. In conclusion, our results suggest that circSSBP2 inhibits the proliferation of bovine intramuscular preadipocytes by regulating the miR-2400/NDRG1 axis.

Separation of bufadienolides from Helleborus thibetanus Franch. by a combination approach involving macroporous resin column chromatography and gradient countercurrent chromatography.

In this study, a combination approach involving macroporous resin (MR) column chromatography and gradient countercurrent chromatography (CCC) was employed to enrich and purify bufadienolides from the roots and rhizomes of Helleborus thibetanus Franch. Initially, a D101 MR-packed column chromatography was utilized for fractionation and enrichment of the bufadienolides, which were effectively eluted from the column using a 60% ethanol solution. CCC was subsequently introduced to separate the enriched product using the ethyl acetate/n-butanol/water (EBuWat, 4:1:5, v/v) and EBuWat (5:0:5, v/v) solvent systems in a gradient elution mode. As results, five bufadienolides, including 6.1 mg of hellebrigenin-3-O-ß-D-glucoside (1), 2.2 mg of tigencaoside A (2), 8.3 mg of deglucohellebrin (3), 3.5 mg of 14 ß-hydroxy-3ß-[ß-D-glucopyranosyl-(1→6)-(ß-D-glucopyranosyl)oxy]-5α-bufa-20,22-dienolide (4), and 3.0 mg of 14ß-hydroxy-3ß-[(ß-D-glucopyranosyl)oxy]-5α-bufa-20,22-dienolide (5), were effectively separated from 300 mg of the enriched product. The respective high-performance liquid chromatography purities were as follows: 95.2%, 75.8%, 85.7%, 82.3%, and 92.8%. This study provides valuable insights for the efficient enrichment and separation of bufadienolides from Helleborus thibetanus Franch.

Eletrocarboxylation Reactions Using CO2 Both as Promoter and Carboxylative Reagent.

Electrocarboxylation reaction, which employs organic electrosynthesis to achieve the utilization of CO2 as a carboxylative reagent, provides a powerful and efficient tool for the preparation of organic carboxylic acid. In some electrocarboxylation reactions, CO2 also acts as a promoter to facilitate the desired reaction. This concept mainly highlights recent CO2 -promoted electrocarboxylation reactions via CO2 ⋅- intermediate or transiently protective carboxylation of active intermediate with CO2 .

Electroreductive Ring-Opening Carboxylation of Cycloketone Oxime Esters with Carbon Dioxide.

Electroreductive ring-opening carboxylation of cycloketone oxime esters with atmospheric carbon dioxide is reported. This reaction proceeded under simple constant current conditions in an undivided cell using glassy carbon as the cathode and magnesium as the sacrificial anode, providing substituted γ- and δ-cyanocarboxylic acids in moderate to good yields. Electrochemically generated cyanoalkyl radicals and cyanoalkyl anion are proposed as the key intermediates.

The role of BBS2 in regulating adipogenesis and the association of its sequence variants with meat quality in Qinchuan cattle.

The BBS2 gene plays a vital role in human obesity and fat deposition. However, little is known about it in beef cattle. Therefore, this study investigates the function of BBS2 in the fat deposition of beef cattle and screens the effective SNPs marker for meat quality traits in cattle breeding. The expression of BBS2 is negatively correlated with marbling ratios of beef cattle. Moreover, the knockdown of BBS2 promoted adipogenesis and lipid accumulation of bovine preadipocytes by stimulating PPARγ, FABP4, and FASN expression (P < 0.01). Four novel SNPs in the exons of BBS2 in Chinese Qinchuan cattle were identified and of which the g.24226239C > T (Q527), g.24223562G > A (V441I), and g.24227851A > G (Q627R) were significantly associated with the meat quality of Qinchuan cattle (P < 0.01, P < 0.05). The findings suggested that BBS2 could be used as a candidate gene for meat quality improvement in Qinchuan cattle. Furthermore, these genotypes can be exploited as molecular markers in future beef breeding projects.

Long Non-Coding RNA BNIP3 Inhibited the Proliferation of Bovine Intramuscular Preadipocytes via Cell Cycle.

The intramuscular fat (or marbling fat) content is an essential economic trait of beef cattle and improves the flavor and palatability of meat. Several studies have highlighted the correlation between long non-coding RNAs (lncRNAs) and intramuscular fat development; however, the precise molecular mechanism remains unknown. Previously, through a high-throughput sequencing analysis, we found a lncRNA and named it a long non-coding RNA BNIP3 (lncBNIP3). The 5' RACE and 3' RACE explored 1945 bp total length of lncBNIP3, including 1621 bp of 5'RACE, and 464 bp of 3'RACE. The nucleoplasmic separation and FISH results explored the nuclear localization of lncBNIP3. Moreover, the tissue expression of lncBNIP3 was higher in the longissimus dorsi muscle, followed by intramuscular fat. Furthermore, down-regulation of lncBNIP3 increased the 5-Ethynyl-2'- deoxyuridine (EdU)-EdU-positive cells. The flow cytometry results showed that the number of cells in the S phase was significantly higher in preadipocytes transfected with si-lncBNIP3 than in the control group (si-NC). Similarly, CCK8 results showed that the number of cells after transfection of si-lncBNIP3 was significantly higher than in the control group. In addition, the mRNA expressions of proliferative marker genes CyclinB1 (CCNB1) and Proliferating Cell Nuclear Antigen (PCNA) in the si-lncBNIP3 group were significantly higher than in the control group. The Western Blot (WB) results also showed that the protein expression level of PCNA transfection of si-lncBNIP3 was significantly higher than in the control group. Similarly, the enrichment of lncBNIP3 significantly decreased the EdU-positive cells in the bovine preadipocytes. The results of flow cytometry and CCK8 assay also showed that overexpression of lncBNIP3 inhibited the proliferation of bovine preadipocytes. In addition, the overexpression of lncBNIP3 significantly inhibited the mRNA expressions of CCNB1 and PCNA. The WB results showed that the overexpression of lncBNIP3 significantly inhibited the expression of the CCNB1 protein level. To further explore the mechanism of lncBNIP3 on the proliferation of intramuscular preadipocytes, RNA-seq was performed after interference with si-lncBNIP3, and 660 differentially expressed genes (DEGs) were found, including 417 up-regulated DEGs and 243 down-regulated DEGs. The KEGG pathway analysis showed that the cell cycle was the most significant pathway for the functional enrichment of DEGs, followed by the DNA replication pathway. The RT-qPCR quantified the expression of twenty DEGs in the cell cycle. Therefore, we speculated that lncBNIP3 regulated intramuscular preadipocyte proliferation through the cell cycle and DNA replication pathways. To further confirm this hypothesis, the cell cycle inhibitor Ara-C was used to inhibit DNA replication of the S phase in intramuscular preadipocytes. Herein, Ara-C and si-lncBNIP3 were simultaneously added to the preadipocytes, and the CCK8, flow cytometry, and EdU assays were performed. The results showed that the si-lncBNIP3 could rescue the inhibitory effect of Ara-C in the bovine preadipocyte proliferation. In addition, lncBNIP3 could bind to the promoter of cell division control protein 6 (CDC6), and down-regulation of lncBNIP3 promoted the transcription activity and the expression of CDC6. Therefore, the inhibitory effect of lncBNIP3 on cell proliferation might be understood through the cell cycle pathway and CDC6 expression. This study provided a valuable lncRNA with functional roles in intramuscular fat accumulation and revealed new strategies for improving beef quality.

Direct and Selective Electrocarboxylation of Styrene Oxides with CO2 for Accessing ß-Hydroxy Acids.

Highly selective and direct electroreductive ring-opening carboxylation of epoxides with CO2 in an undivided cell is reported. This reaction shows broad substrate scopes within styrene oxides under mild conditions, providing practical and scalable access to important synthetic intermediate ß-hydroxy acids. Mechanistic studies show that CO2 functions not only as a carboxylative reagent in this reaction but also as a promoter to enable efficient and chemoselective transformation of epoxides under additive-free electrochemical conditions. Cathodically generated α-radical and α-carbanion intermediates lead to the regioselective formation of α-carboxylation products.

MiR-145 reduces the activity of PI3K/Akt and MAPK signaling pathways and inhibits adipogenesis in bovine preadipocytes.

Adipose tissue is the largest metabolic organ because of adipogenesis controlled by numerous miRNAs. MiR-145 is classified into the same cluster with famous miR-143. However, few studies have investigated the role of miR-145 in adipogenesis. In the current study, we observed that the expression of miR-145 was downregulated during bovine adipogenesis in vivo and in vitro. The results of RNA-Seq analysis showed that miR-145 mainly disturb the PI3K/Akt and MAPK signaling pathways in bovine preadipocytes. MiR-145 inhibited bovine preadipocyte differentiation and downregulated phosphorylation level of Akt and ERK1/2 proteins. Furthermore, insulin, as a powerful inducer initiating adipogenesis and an activator of the PI3K/Akt and MAPK signaling pathways, was able to rescue the downregulation of Akt and ERK1/2 phosphorylation levels caused by miR-145. Taken together, our findings suggest that miR-145 is a potent inhibitor of adipogenesis that may function by reducing the activity of PI3K/Akt and MAPK signaling pathways.

Carboxylative Cyclization of 2-Butenoates with Carbon Dioxide: Access to Glutaconic Anhydrides.

Cyclic anhydrides are versatile synthons and functional comonomers. Herein, we reported an organic base-promoted carboxylative cyclization of 2-butenoates with carbon dioxide to produce important glutaconic anhydrides in good yields. This metal-free reaction showed broad substrate scopes and proceeded under mild reaction conditions.

Transcriptional regulation of bovine elongation of very long chain fatty acids protein 6 in lipid metabolism and adipocyte proliferation.

The elongation of very long chain fatty acids protein 6 (ELOVL6) gene encodes a key enzyme that plays a role in lipogenesis through the catalytic elongation of both saturated and monounsaturated fatty acids. Previous studies have described the high expression of bovine ELOVL6 in adipose tissues. However, transcriptional regulation and the functional role of ELOVL6 in lipid metabolism and adipocyte proliferation remain unexplored. Here, a 1.5 kb fragment of the 5'-untranslated region promoter region of ELOVL6 was amplified from the genomic DNA of Qinchuan cattle and sequenced. The core promoter region was identified through unidirectional 5'-end deletion of the promoter plasmid vector. In silico analysis predicted important transcription factors that were then validated through site-directed mutation and small interfering RNA interference with an electrophoretic mobility shift assay. We found that the binding of KLF6 and PU.1 transcription factors occurred in the region -168/+69. Both perform a vital regulatory function in the transcription of bovine ELOVL6. Overexpression of ELOVL6 significantly upregulated the expression of peroxisome proliferator activated receptor γ (PPARγ), but inhibited the expression of fatty acid-binding protein 4 (FABP4), while silencing of ELOVL6 negatively regulated the messenger RNA expression level of PPARγ, FABP4, ACSL, and FATP1. In addition, ELOVL6 promotes adipocyte proliferation by regulating the cell-cycle genes' expression. Taken together, these findings provide useful information about the transcriptional regulation and functional mechanisms of bovine ELOVL6 in lipid metabolism and adipocyte proliferation in Qinchuan cattle.

Characterization of the promoter region of bovine SIX4: Roles of E-box and MyoD in the regulation of basal transcription.

The sine oculis homeobox 4 (SIX4) gene belongs to the Six gene family and encodes an evolutionarily conserved transcription factor. Previous studies have demonstrated that SIX4 plays an essential role in proper muscle regeneration. However, the mechanisms regulating SIX4 transcription remain elusive. In the present study, we determined that bovine SIX4 was highly expressed in the longissimus thoracis and in undifferentiated Qinchuan cattle muscle cells (QCMCs) and that its protein localizes to both the cytoplasm and the nucleus. To elucidate the bovine the molecular mechanisms of SIX4 regulation, 1.3 kb of the 5'-regulatory region was obtained. MyoD and Ebox recognition sites were identified in the core promoter region at -522/-193 of the bovine SIX4 using a series of 5' deletion promoter plasmids in luciferase reporter assays. An electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay in combination with site-directed mutation and siRNA interference demonstrated that MyoD binding occurs at MyoD and Ebox recognition sites through direct and indirect mechanisms and play important roles in the transcriptional regulation of the bovine SIX4 promoter. Taken together, these interactions provide insight into the regulatory mechanisms of SIX4 transcription in mediating skeletal muscle growth in cattle.

Identification of Potential Key Genes Associated with Adipogenesis through Integrated Analysis of Five Mouse Transcriptome Datasets.

Adipose tissue is the most important energy metabolism and secretion organ, and these functions are conferred during the adipogenesis process. However, the cause and the molecular events underlying adipogenesis are still unclear. In this study, we performed integrated bioinformatics analyses to identify vital genes involved in adipogenesis and reveal potential molecular mechanisms. Five mouse high-throughput expression profile datasets were downloaded from the Gene Expression Omnibus (GEO) database; these datasets contained 24 samples of 3T3-L1 cells during adipogenesis, including 12 undifferentiated samples and 12 differentiated samples. The five datasets were reanalyzed and integrated to select differentially expressed genes (DEGs) during adipogenesis via the robust rank aggregation (RRA) method. Functional annotation of these DEGs and mining of key genes were then performed. We also verified the expression levels of some potential key genes during adipogenesis. A total of 386 consistent DEGs were identified, with 230 upregulated genes and 156 downregulated genes. Gene Ontology (GO) analysis showed that the biological functions of the DEGs primarily included fat cell differentiation, lipid metabolic processes, and cell adhesion. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that these DEGs were mainly associated with metabolic pathways, the peroxisome proliferator-activated receptor (PPAR) signaling pathway, regulation of lipolysis in adipocytes, the tumor necrosis factor (TNF) signaling pathway, and the FoxO signaling pathway. The 30 most closely related genes among the DEGs were identified from the protein⁻protein interaction (PPI) network and verified by real-time quantification during 3T3-L1 preadipocyte differentiation. In conclusion, we obtained a list of consistent DEGs during adipogenesis through integrated analysis, which may offer potential targets for the regulation of adipogenesis and treatment of adipose dysfunction.

1,3-Dipolar Cycloaddition of Nitrile Imine with Carbon Dioxide: Access to 1,3,4-Oxadiazole-2(3H)-ones.

Efficient synthesis of 1,3,4-oxadiazole-2(3H)-one was achieved by CsF/18-crown-6 mediated 1,3-dipolar cycloaddition of nitrile imine and 2.0 MPa of CO2. CsF/18-crown-6 played a key role in enhancing the reactivity of CO2 as a 1,3-dipolarophile. The practical utility of this transition-metal-free approach to 1,3,4-oxadiazole-2(3H)-one is highlighted by the convenient synthesis of a commercial herbicide Oxadiazon and a MAO B inhibitor.

Access to α-Arylglycines by Umpolung Carboxylation of Aromatic Imines with Carbon Dioxide.

A straightforward and transition-metal-free approach for the efficient synthesis of α-arylglycine derivatives from aromatic imines and carbon dioxide was enabled by an umpolung carboxylation reaction. Various substituted diphenylmethimines underwent the carboxylation smoothly with carbon dioxide in the presence of potassium tert-butoxide and 18-crown-6 to give the corresponding carboxylated products in good to high yields. Besides the enhancement of the solubility of potassium tert-butoxide in THF, 18-crown-6 also plays key roles in suppressing the reverse protonation or 1, 3-proton shift isomerization as well as by stabilizing the carboxylated intermediate.

CO2-Mediated Formation of Chiral Carbamates from meso-Epoxides via Polycarbonate Intermediates.

Carbon dioxide has attracted broad interest as a renewable C1 feedstock for efficient transformation into value-added organic chemicals; nevertheless, far less attention was paid to its stereochemically controlled catalytic fixation/conversion processes. Here, we report a new strategy for the selective synthesis of chiral carbamates from carbon dioxide via polycarbonate intermediates, which are formed by the desymmetric copolymerization of meso-epoxides using enantiopure dinuclear Co(III) catalyst systems with 99% enantioselectivity. Subsequent degradation reaction of the resultant polycarbonates with various primary or secondary amine nucleophiles can afford optically active carbamates, with the complete configuration retention of the two chiral carbon centers. Our accomplishment reported here opens up a new route to prepare a wide range of CO2-based carbamate scaffolds with excellent yields and 99% enantiomeric excess.

Enantioselective rhodium-catalyzed isomerization of 4-iminocrotonates: asymmetric synthesis of a unique chiral synthon.

An enantioselective isomerization of 4-iminocrotonates catalyzed by a rhodium(I)/phosphoramidite complex is described. This reaction uses widely available amines to couple with 4-oxocrotonate to provide a convenient access to a central chiral building block in good yield and high enantioselectivity. Although the mechanism of this new transformation remains unclear, both Rh and the phosphoramidite play a central role.

DBU-promoted carboxylative cyclization of o-hydroxy- and o-acetamidoacetophenone.

The carboxylative cyclization of o-hydroxy- and o-acetamidoacetophenone with carbon dioxide promoted by the organic base 1,8-diazabicycloundec-7-ene (DBU) is reported. This reaction provides convenient access to the biologically important compounds 4-hydroxy-2H-chromen-2-one and 4-hydroxy-2(1H)-quinolinone in moderate to good yields using carbon dioxide as the carboxylation reagent. An acyl migration from nitrogen to carbon is observed in the reaction of o-acetamidoacetophenone.

Long-term outcome of IgA nephropathy patients with recurrent macroscopic hematuria.

BACKGROUND/AIMS: The long-term renal outcomes of patients with IgA nephropathy (IgAN) who present with recurrent macroscopic hematuria (RMH) have not been described in previous studies. METHODS: Patients with biopsy-proven primary IgAN in Jinling Hospital were divided into three groups according to different patterns of macroscopic hematuria (MH): RMH, isolated MH (IMH), and those without a history of MH (NMH). RESULTS: A total of 1,155 patients were enrolled in the study (158 in the RMH group, 256 in the IMH group, and 741 in the NMH group). At biopsy, patients with RMH were younger, had lower median proteinuria, a lower incidence of hypertension, and a higher estimated glomerular filtration rate than those in the NMH group. Pathologically, patients with RMH had a lower level of mesangial hypercellularity and segmental glomerulosclerosis as well as less tubular atrophy than those with NMH. The demographic and clinical features of patients with IMH fell between patients with RMH and those with NMH. During a median follow-up of 7.9 years, the 5-, 10- and 20-year cumulative renal survival after biopsy, as calculated by K-M methods, were 98, 91, and 91% in the RMH group, 95, 89, and 64% in the IMH group, and 95, 79, and 57% in the NMH group. The renal survival in patients with RMH was significantly better than patients with NMH or IMH. CONCLUSIONS: The long-term prognosis of patients who present with RMH is significantly better than patients with NMH or IMH.

An improved point cloud denoising method in adverse weather conditions based on PP-LiteSeg network.

Reliable point cloud data (PCD) generated by LiDAR are crucial to perceiving surroundings when autonomous driving systems are a concern. However, adverse weather conditions can impact the detection range of LiDAR, resulting in a significant amount of noisy data that substantially deteriorates the quality of PCD. Point cloud denoising algorithms used for challenging weather conditions suffer from poor accuracy and slow inferences. The manuscript proposes a Series Attention Fusion Denoised Network (SAFDN) based on a semantic segmentation model in real-time, called PP-LiteSeg. The proposed approach provides two key components to the model. The insufficient feature extraction issue in the general-purpose segmentation models is first addressed when dealing with objects with more noise, so the WeatherBlock module is introduced to replace the original layer used for feature extraction. Hence, this module employs dilated convolutions to enhance the receptive field and extract multi-scale features by combining various convolutional kernels. The Series Attention Fusion Module (SAFM) is presented as the second component of the model to tackle the problem of low segmentation accuracy in rainy and foggy weather conditions. The SAFM sequentially applies channel and spatial attention mechanisms to enhance the model's sensitivity to crucial features. Furthermore, weighted feature fusion is employed to enhance the model's efficiency in integrating low-level and high-level feature information configurations. Experimental evaluations were conducted on the publicly available DENSE dataset. The results demonstrate that the improved model achieved an 11.1% increase in denoising accuracy measured by MIOU and an inference speed of 205.06 FPS when compared to the PP-LiteSeg model. As a result, the noise recognition accuracy and denoising capability in real-time are enhanced.