Abstract
Domestic cats, an important companion animal, can be infected with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). This has aroused concern regarding the ability of domestic ...cats to spread the virus that causes coronavirus disease 2019. We systematically demonstrated the pathogenesis and transmissibility of SARS-CoV-2 in cats. Serial passaging of the virus between cats dramatically attenuated the viral transmissibility, likely owing to variations of the amino acids in the receptor-binding domain sites of angiotensin-converting enzyme 2 between humans and cats. These findings provide insight into the transmissibility of SARS-CoV-2 in cats and information for protecting the health of humans and cats.
Domestic cats are susceptible to SARS-CoV-2 and have attenuated transmissibility after serial passaging.
B cells are a class of professional antigen-presenting cells that produce antibodies to mediate humoral immune response and participate in immune regulation. m
A modification is the most common RNA ...modification in mRNA; it involves almost all aspects of RNA metabolism and can affect RNA splicing, translation, stability, etc. This review focuses on the B-cell maturation process as well as the role of three m
A modification-related regulators-writer, eraser, and reader-in B-cell development and B-cell-related diseases. The identification of genes and modifiers that contribute to immune deficiency may shed light on regulatory requirements for normal B-cell development and the underlying mechanism of some common diseases.
Melatonin is a pleiotropic molecule with a variety of biological functions, which include its immunoregulatory action in mammals. Brucellosis is a worldwide endemic zoonotic disease caused by the ...Brucella, which not only causes huge economic losses for the livestock industry but also impacts human health. To target this problem, in current study, two marker‐free transgenic sheep overexpressing melatonin synthetic enzyme ASMT (acetylserotonin O‐methyltransferase) gene were generated and these melatonin enrich transgenic sheep were challenged by Brucella infection. The results showed that the serum melatonin concentration was significantly higher in transgenic sheep than that of wild type (726.92 ± 70.6074 vs 263.10 ± 34.60 pg/mL, P < .05). Brucella challenge test showed that two thirds (4/6) of the wild‐type sheep had brucellosis, while none of the transgenic sheep were infected. Whole‐blood RNA‐seq results showed that differential expression genes (DEGs) were significantly enriched in natural killer cell‐mediated cytotoxicity, phagosome, antigen processing, and presentation signaling pathways in overexpression sheep. The DEGs of toll‐like receptors (TLRs) and NOD‐like receptors (NLRs) families were verified by qPCR and it showed that TLR1, TLR2, TLR7, CD14, NAIP, and CXCL8 expression levels in overexpression sheep were significantly higher and NLRP1, NLRP3, and TNF expression levels were significantly lower than those of wild type. The rectal feces were subjected to 16S rDNA amplicon sequencing, and the microbial functional analysis showed that the transgenic sheep had significantly lower abundance of microbial genes related to infectious diseases compared to the wild type, indicating overexpression animals are likely more resistant to infectious diseases than wild type. Furthermore, exogenous melatonin treatment relieved brucellosis inflammation by upregulating anti‐inflammatory cytokines IL‐4 and downregulating pro‐inflammatory IL‐2, IL‐6, and IFN‐γ. Our preliminary results provide an informative reference for the study of the relationship between melatonin and brucellosis.
The yield efficiency of transgenic animal generation is relatively low
1
. To improve its efficiency has become a priority task for researchers
2
. Melatonin (N-acetyl-5-methoxytryptamine, MT) is a ...potent-free radical scavenger and antioxidant to protect mitochondria, lipids, protein and DNA from oxidative stress
3
. In this study, we observed that improving the quality of both donor and recipient cells by giving physiological concentration (10
−7
M) of MT significantly increase the sheep transgenic embryo development in the in vitro condition. MT promotes the donor cell viability, proliferation, efficiency of monoclonal formation and the electrotransferring efficiency of fetal fibroblast cells (FFCs). The mechanistic exploration indicates that MT has the capacity for the synchronization of cell division cycle, reduction of cellular oxidative stress, apoptosis, and the increase of mitochondrial number and function. All of these render MT's ability to increase the efficiency of animal transgenic processes such as somatic cell nuclear transfer (SCNT) and electroporation. The outcomes are the increased cleavage rate and blastocyst rate of the transgenic sheep embryos after MT treatment. These beneficial effects of MT on transgenic embryo development are worth to be tested in the in vivo condition in the future.
Melatonin plays an important role in mammalian reproductive activities, to further understand the effects of endogenous melatonin on functions of ovary, the transgenic sheep with overexpression of ...melatonin synthetic enzyme gene ASMT in ovary were generated. The results showed that total melatonin content in follicular fluid of transgenic sheep was significantly greater than that in the wild type. Accordingly, the follicle numbers of transgenic sheep were also significantly greater than those in the WT. The results of follicular fluid metabolites sequencing showed that compared with WT, the differential metabolites of the transgenic sheep were significantly enriched in several signaling pathways, the largest number of metabolites was lipid metabolism pathway and the main differential metabolites were lipids and lipoid molecules. SMART-seq2 were used to analyze the oocytes and granulosa cells of transgenic sheep and WT sheep. The main differential enrichment pathway was metabolic pathway, in which lipid metabolism genes accounted for the majority. In conclusion, this is the first report to show that ovary overexpression of ASMT increased local melatonin production and follicle numbers. These results may imply that ASMT plays an important role in follicle development and formation, and melatonin intervention may be a potential method to promote this process.
•Overexpression of ASMT in ovary does not affect the health of sheep.•Overexpression of ASMT in ovary can increase the level of melatonin in follicular fluid of ewes.•Overexpression of ASMT in ovary increases the number of follicles in ewes.•The difference of ovarian metabonomics between transgenic sheep and wild type sheep is enriched in lipid metabolic pathway.•The ovarian differential genes of transgenic sheep and wild-type sheep are mainly concentrated in lipid metabolism.
Improving livestock and poultry growth rates and increasing meat production are urgently needed worldwide. Previously, we produced a myostatin (MSTN) and fibroblast growth factor 5 (FGF5) ...double-knockout (MF
) sheep by CRISPR Cas9 system to improve meat production, and also wool production. Both MF
sheep and the F1 generation (MF
) sheep showed an obvious "double-muscle" phenotype. In this study, we identified the expression profiles of long noncoding RNAs (lncRNAs) in wild-type and MF
sheep, then screened out the key candidate lncRNAs that can regulate myogenic differentiation and skeletal muscle development. These key candidate lncRNAs can serve as critical gatekeepers for muscle contraction, calcium ion transport and skeletal muscle cell differentiation, apoptosis, autophagy, and skeletal muscle inflammation, further revealing that lncRNAs play crucial roles in regulating muscle phenotype in MF
sheep. In conclusion, our newly identified lncRNAs may emerge as novel molecules for muscle development or muscle disease and provide a new reference for MSTN-mediated regulation of skeletal muscle development.
To explore the anti-inflammatory activity of endogenous produced melatonin, a melatonin-enriched animal model (goat) with AANAT transfer was successfully generated with somatic cell nuclear transfer ...(SCNT) technology. Basically, a pIRES2-EGFP-AANAT expression vector was constructed and was transferred into the female fetal fibroblast cells (FFCs) via electrotransfection and then the nuclear of the transgenic FFC was transferred to the eggs of the donor goats. The peripheral blood mononuclear cells (PBMCs) of the transgenic offspring expressed significantly higher levels of AANAT and melatonin synthetic function than those PBMCs from the wild-type (WT) animals. After challenge with lipopolysaccharide (LPS), the transgenic PBMCs had increased autophagosomes and LC3B expression while they exhibited suppressed production of the proinflammatory cytokines, IL1B and IL12 (IL12A-IL12B/p70), compared to their WT. The mechanistic analysis indicated that the anti-inflammatory activity of endogenous melatonin was mediated by MTNR1B (melatonin receptor 1B). MTNR1B stimulation activated the MAPK14 signaling pathway to promote cellular macroautophagy/autophagy, thus, suppressing the excessive inflammatory response of cellular. However, when the intact animals challenged with LPS, the serum proinflammatory cytokines were significantly higher in the transgenic goats than that in the WT. The results indicated that endogenous melatonin inhibited the MAPK1/3 signaling pathway and ROS production, subsequently downregulated gene expression of BECN1, ATG5 in PMBCs and then suppressed the autophagy activity of PBMCs and finally elevated levels of serum proinflammatory cytokines in transgenic animals, Herein we provided a novel melatonin-enriched animal model to study the potential effects of endogenously produced melatonin on inflammatory responses and autophagy activity.
•PKZ and PKR were up-regulated by poly I:C in a time-dependent manner.•Both PKZ and Caspase-3 mRNA came with a high degree of consistency in CIK cells either after treatment with poly I:C or ...transfection with siRNA against PKZ gene.•Overexpression of wild type PKZ but not mutant K198R in CIK cells resulted in higher apoptotic rate and a significantly reduced viability rate.•Overexpression of wild type eIF2α induced apoptosis, whereas phosphodeficient eIF2α (S51A) blocked induction of apoptosis.•Mutant of eIF2α (S51A) would block the induction of apoptosis and PKZ could elicit apoptosis via phosphorylating eIF2α at Ser51 excluding the effects of endogenous PKZ/PKR largely.
PKZ, protein kinase containing Z-DNA binding domains, is a novel member of eIF2α kinase in fish. PKZ can be up-regulated in response to Poly I:C or heat stress, and it has a typical eIF2α kinase activity. However, the relationship between fish PKZ and apoptosis remains unclear. In the present study, effects of PKZ on apoptosis were explored. Initially, we found that PKZ and PKR were up-regulated by poly I:C in a time-dependent manner. Q-PCR analysis showed that the change of Caspase-3 mRNA was consistent with that of PKZ under the stimulation of poly I:C or transfection with PKZ siRNA in CIK cells. It suggested that PKZ might mediate the induction of apoptosis. Knockdown and overexpression assays indicated that a significant increase of apoptotic cell number was observed in the wild type PKZ (PKZ-wt) transfected PKZ-deficient cells, while mutant PKZ-K198R lost this ability. MTT also showed that overexpression of PKZ-wt in CIK cells resulted in a striking decrease of cell viability rate to about 32.5%, whereas the viability of cell with PKZ-K198R was about 88.1%. Likewise, when transfected eIF2α-wt or phosphomimetic eIF2α (S51D), CIK cells displayed higher apoptotic rate than the controls. In contrast, overexpression of phosphodeficient eIF2α (S51A) blocked induction of apoptosis. In addition, levels of eIF2α phosphorylation were significantly related to apoptosis in these CIK cells. Furthermore, when mutant eIF2α (S51A) was transiently transfected into the PKZ/PKR knockdown cells that transfected with wildtype PKZ previously, the apoptotic rates and levels of eIF2α phosphorylation were dramatically decreased. Overall, these results suggested that PKZ could facilitate apoptosis via eIF2α phosphorylation.
Melatonin as a potent antioxidant exhibits important nutritional and medicinal values. To produce melatonin‐enriched milk will benefit the consumers. In this study, a sheep bioreactor which generates ...melatonin‐enriched milk has been successfully developed by the technology that combined CRISPR/Cas9 system and microinjection. The AANAT and ASMT were cloned from pineal gland of Dorper sheep (Ovis aries). The in vitro studies found that AANAT and ASMT were successfully transferred to the mammary epithelial cell lines and significantly increased melatonin production in the culture medium compared to the nontransgenic cell lines. In addition, the Cas9 mRNA, sgRNA, and the linearized vectors pBC1‐AANAT and pBC1‐ASMT were co‐injected into the cytoplasm of pronuclear embryos which were implanted into ewes by oviducts transferring. Thirty‐four transgenic sheep were generated with the transgenic positive rate being roughly 35% which were identified by Southern blot and sequencing. Seven carried transgenic AANAT, two carried ASMT, and 25 carried both of AANAT and ASMT genes. RT‐PCR and Western blot demonstrated that the lambs expressed these genes in their mammary epithelial cells and these animals produced melatonin‐enriched milk. This is the first report to show a functional AANAT and ASMT transgenic animal model which produce significantly high levels of melatonin milk compared to their wild‐type counterparts. The advanced technologies used in the study laid a foundation for generating large transgenic livestock, for example, the cows, which can produce high level of melatonin milk.
Spermatogenesis in mammalian testes is essential for male fertility, ensuring a continuous supply of mature sperm. The testicular microenvironment finely tunes this process, with retinoic acid, an ...active metabolite of vitamin A, serving a pivotal role. Retinoic acid is critical for various stages, including the differentiation of spermatogonia, meiosis in spermatogenic cells, and the production of mature spermatozoa. Vitamin A deficiency halts spermatogenesis, leading to the degeneration of numerous germ cells, a condition reversible with retinoic acid supplementation. Although retinoic acid can restore fertility in some males with reproductive disorders, it does not work universally. Furthermore, high doses may adversely affect reproduction. The inconsistent outcomes of retinoid treatments in addressing infertility are linked to the incomplete understanding of the molecular mechanisms through which retinoid signaling governs spermatogenesis. In addition to the treatment of male reproductive disorders, the role of retinoic acid in spermatogenesis also provides new ideas for the development of male non-hormone contraceptives. This paper will explore three facets: the synthesis and breakdown of retinoic acid in the testes, its role in spermatogenesis, and its application in male reproduction. Our discussion aims to provide a comprehensive reference for studying the regulatory effects of retinoic acid signaling on spermatogenesis and offer insights into its use in treating male reproductive issues.
PDF
