DNA replication in early mammalian embryos is patterned, predisposing lamina-associated regions to fragility.

DNA replication in differentiated cells follows a defined program, but when and how it is established during mammalian development is not known. Here we show using single-cell sequencing, that late replicating regions are established in association with the B compartment and the nuclear lamina from the first cell cycle after fertilization on both maternal and paternal genomes. Late replicating regions contain a relative paucity of active origins and few but long genes and low G/C content. In both bovine and mouse embryos, replication timing patterns are established prior to embryonic genome activation. Chromosome breaks, which form spontaneously in bovine embryos at sites concordant with human embryos, preferentially locate to late replicating regions. In mice, late replicating regions show enhanced fragility due to a sparsity of dormant origins that can be activated under conditions of replication stress. This pattern predisposes regions with long neuronal genes to fragility and genetic change prior to separation of soma and germ cell lineages. Our studies show that the segregation of early and late replicating regions is among the first layers of genome organization established after fertilization.

Effect of ultrasound-assisted L-lysine treatment on pork meat quality and myofibrillar protein properties during postmortem aging.

This paper aimed to investigate the effects of ultrasound-assisted L-lysine treatment on meat quality and myofibrillar proteins (MPs) properties of pork longissimus dorsi during postmortem aging. The results revealed that the L-lysine (Lys) and/or ultrasound treatment significantly increased (p < 0.05) the water-holding capacity and tenderness of the pork during postmortem aging, while the ultrasound-assisted Lys treatment had the lowest cooking loss, pressurization loss, Warner-Bratzler shear force, and hardness. In addition, L-lysine and/or ultrasound treatment increased (p < 0.05) pH value, T21, and myofibrillar fragmentation index, while the ultrasound-assisted Lys treatment had the highest value. Meanwhile, the protein solubility was increased with Lys and/or ultrasound treatment during postmortem aging, and ultrasound-assisted Lys treatment had the highest solubility, reaching 88.19%, 92.98%, and 91.73% at 0, 1, and 3 days, respectively. The result of protein conformational characteristics showed that Lys and/or ultrasound treatment caused the unfolding of the α-helix structure, resulting in the exposure of more hydrophobic amino acids and buried sulfhydryl groups, ultimately enhancing MPs solubility. In summary, ultrasound-assisted Lys treatment altered the structure of MPs, resulting in the enhancement of the water-holding capacity and tenderness of the pork. PRACTICAL APPLICATION: This study showed that ultrasound-assisted L-lysine (Lys) treatment could enhance the water-holding capacity and tenderness of pork during postmortem aging. The results might provide a reference for the application of ultrasound-assisted Lys treatment on the improvement of pork meat quality. To facilitate practical applications in production, the development of medium and large-sized ultrasound equipment for conducting small-scale and pilot experiments is crucial for future research.

Permanent neonatal diabetes-causing insulin mutations have dominant negative effects on beta cell identity.

OBJECTIVE: Heterozygous coding sequence mutations of the INS gene are a cause of permanent neonatal diabetes (PNDM), requiring insulin therapy similar to T1D. While the negative effects on insulin processing and secretion are known, how dominant insulin mutations result in a continued decline of beta cell function after birth is not well understood. METHODS: We explored the causes of beta cell failure in two PNDM patients with two distinct INS mutations using patient-derived iPSCs and mutated hESCs. RESULTS: we detected accumulation of misfolded proinsulin and impaired proinsulin processing in vitro, and a dominant-negative effect of these mutations on beta-cell mass and function after transplantation into mice. In addition to anticipated ER stress, we found evidence of beta-cell dedifferentiation, characterized by an increase of cells expressing both Nkx6.1 and ALDH1A3, but negative for insulin and glucagon. CONCLUSIONS: These results highlight a novel mechanism, the loss of beta cell identity, contributing to the loss and functional failure of human beta cells with specific insulin gene mutations.

Characterization and Correlation of Dominant Microbiota and Flavor Development in Different Post-Mortem Processes of Beef.

Post-mortem aging could enhance the unique flavors of beef via several biochemical pathways. The microbiota is one of the important factors in the flavor development of aging beef, but their potential relationship has rarely been studied. This study characterized the apparent meat quality, flavor profiles, and microbial communities of beef during the different post-mortem processes, followed by the investigation of the correlations between the dominant microbiota and key volatile compounds. The results showed that wet-aged beef has a higher product yield and more stable color than dry-aged beef, as evidenced by the significantly lower value of aging loss and discoloration (ΔE). According to the odor activity value, 11 out of 65 compounds were categorized as aroma-active components, and 9 of them, including 1-pentanol, 1-octen-3-ol, hexanal, nonanal, heptanal, octanal, 2-nonenal, (E)-, 2-octenal, (E)- and 2-decenal, (E)-, were enriched in beef wet-aged for 7 d. Significant variances were found in the microbial communities of different aging beef. Of these, 20 microbiota (with 10 bacterial and 10 fungal genera) were recognized as the dominant genus. Partial least squares regression combined with a correlation network model revealed that five microbial genera, including Trichosporon, Prauserella, Rhodotorula, Malassezia, and Corynebacterium, constituted the functional microbiota responsible for flavor formation in aging beef and were positively associated with ≥7 key volatile compounds (p < 0.05, |ρ| > 0.7). This study suggests that the application of wet aging within 7 d on beef is better for meat quality and provides novel insights into the mechanisms of flavor formation in post-mortem aging beef via functional microbiota.

Permanent Neonatal diabetes-causing Insulin mutations have dominant negative effects on beta cell identity.

Heterozygous coding sequence mutations of the INS gene are a cause of permanent neonatal diabetes (PNDM) that results from beta cell failure. We explored the causes of beta cell failure in two PNDM patients with two distinct INS mutations. Using b and mutated hESCs, we detected accumulation of misfolded proinsulin and impaired proinsulin processing in vitro, and a dominant-negative effect of these mutations on the in vivo performance of patient-derived SC-beta cells after transplantation into NSG mice. These insulin mutations derange endoplasmic reticulum (ER) homeostasis, and result in the loss of beta-cell mass and function. In addition to anticipated apoptosis, we found evidence of beta-cell dedifferentiation, characterized by an increase of cells expressing both Nkx6.1 and ALDH1A3, but negative for insulin and glucagon. These results highlight both known and novel mechanisms contributing to the loss and functional failure of human beta cells with specific insulin gene mutations.

Replication stress in mammalian embryo development, differentiation, and reprogramming.

Duplicating a genome of 3 billion nucleotides is challenged by a variety of obstacles that can cause replication stress and affect the integrity of the genome. Recent studies show that replication fork slowing and stalling is prevalent in early mammalian development, resulting in genome instability and aneuploidy, and constituting a barrier to development in human reproduction. Genome instability resulting from DNA replication stress is a barrier to the cloning of animals and to the reprogramming of differentiated cells to induced pluripotent stem cells, as well as a barrier to cell transformation. Remarkably, the regions most impacted by replication stress are shared in these different cellular contexts, affecting long genes and flanking intergenic areas. In this review we integrate our knowledge of DNA replication stress in mammalian embryos, in programming, and in reprogramming, and we discuss a potential role for fragile sites in sensing replication stress and restricting cell cycle progression in health and disease.

Dose-Dependent Effect of Hyperoside on the Physicochemical and Gel Properties of Porcine Myofibrillar Proteins at Different NaCl Concentrations under Oxidative Stress.

The effects of HYP (10, 50, and 250 µM/g protein) on the physicochemical and gel properties of myofibrillar proteins (MPs) at different NaCl concentrations under oxidative stress were explored. The incorporation of HYP significantly reduced carbonyl content and decreased the loss of free amine groups in a dose-dependent manner, regardless of NaCl concentration. In addition, HYP induced a dose-dependent decrement in total sulfhydryl content regardless of NaCl concentration, which might result from the formation of thiol-quinone adducts via Michael addition. The surface hydrophobicity was significantly increased with HYP addition. Nevertheless, compared with samples treated with 50 µM/g HYP, 250 µM/g HYP caused a significant decrease in surface hydrophobicity, which might be due to the increase in the extent of MPs unfolding and the concomitant aggregation of MPs by hydrophobic interaction. Furthermore, HYP also showed a dose-dependent increment in the water-holding capacity (WHC) and gel strength of MPs gels, which might be due to more orderly crosslinks via fibrous filaments at 0.2 M NaCl and more regular and lamellar structures with smaller and more homogeneous pores at 0.6 M NaCl. In summary, HYP reduced the oxidation-mediated changes of physicochemical characteristics, preventing the oxidative damage of MPs and reinforcing the ordered crosslinks of MPs-MPs and MPs-HYP during thermal gelation, ultimately resulting in a better gel quality. These results provide a theoretical support for the practical application of HYP as a natural antioxidant in gel-type meat products.

DNA replication in early mammalian embryos is patterned, predisposing lamina-associated regions to fragility.

DNA replication in differentiated cells follows a defined program, but when and how it is established during mammalian development is not known. Here we show using single-cell sequencing, that both bovine and mouse cleavage stage embryos progress through S-phase in a defined pattern. Late replicating regions are associated with the nuclear lamina from the first cell cycle after fertilization, and contain few active origins, and few but long genes. Chromosome breaks, which form spontaneously in bovine embryos at sites concordant with human embryos, preferentially locate to late replicating regions. In mice, late replicating regions show enhanced fragility due to a sparsity of dormant origins that can be activated under conditions of replication stress. This pattern predisposes regions with long neuronal genes to fragility and genetic change prior to segregation of soma and germ line. Our studies show that the formation of early and late replicating regions is among the first layers of epigenetic regulation established on the mammalian genome after fertilization.

Replication stress impairs chromosome segregation and preimplantation development in human embryos.

Human cleavage-stage embryos frequently acquire chromosomal aneuploidies during mitosis due to unknown mechanisms. Here, we show that S phase at the 1-cell stage shows replication fork stalling, low fork speed, and DNA synthesis extending into G2 phase. DNA damage foci consistent with collapsed replication forks, DSBs, and incomplete replication form in G2 in an ATR- and MRE11-dependent manner, followed by spontaneous chromosome breakage and segmental aneuploidies. Entry into mitosis with incomplete replication results in chromosome breakage, whole and segmental chromosome errors, micronucleation, chromosome fragmentation, and poor embryo quality. Sites of spontaneous chromosome breakage are concordant with sites of DNA synthesis in G2 phase, locating to gene-poor regions with long neural genes, which are transcriptionally silent at this stage of development. Thus, DNA replication stress in mammalian preimplantation embryos predisposes gene-poor regions to fragility, and in particular in the human embryo, to the formation of aneuploidies, impairing developmental potential.

Huangkui Capsule Attenuates Lipopolysaccharide-Induced Acute Lung Injury and Macrophage Activation by Suppressing Inflammation and Oxidative Stress in Mice.

Background: Huangkui capsule (HKC) comprises the total flavonoid extract of flowers of Abelmoschus manihot (L.) Medicus. This study aimed to explore the effects of HKC on lipopolysaccharide- (LPS-) induced acute lung injury (ALI) in mice and LPS-stimulated RAW 264.7 cells. Methods: Enzyme-linked immunosorbent assay, histopathology, spectrophotometry, and quantitative real-time polymerase chain reaction were used for the assessments. Statistical analysis was performed using a one-way analysis of variance. Results: LPS significantly increased lung inflammation, neutrophil infiltration, and oxidative stress and downregulated lung miR-451 expression. Treatment with HKC dramatically, reduced the total cell count in the bronchoalveolar lavage fluid (BALF), and inhibited myeloperoxidase activity in the lung tissues 24 h after LPS challenge. Histopathological analysis demonstrated that HKC attenuated LPS-induced tissue oedema and neutrophil infiltration in the lung tissues. Additionally, the concentrations of tumour necrosis factor- (TNF-) α and interleukin- (IL-) 6 in BALF and IL-6 in the plasma reduced after HKC administration. Moreover, HKC could enhance glutathione peroxidase and catalase activities and upregulate the expression of miR-451 in the lung tissues. In vitro experiments revealed that HKC inhibited the production of nitric oxide, TNF-α, and IL-6 in LPS-induced RAW 264.7 cells and mouse primary peritoneal macrophages. Additionally, HKC downregulated LPS-induced transcription of TNF-α and IL-6 in RAW 264.7 cells. Conclusions: These findings suggest that HKC has anti-inflammatory and antioxidative effects that may protect mice against LPS-induced ALI and macrophage activation.