Dexmedetomidine attenuates isoflurane-induced neuroapoptosis through the miR-137/GSK-3ß pathway in the developing rat hippocampus.

Long-term isoflurane inhalation has been reported to induce hippocampal apoptosis in young animals, whereas dexmedetomidine (DEX) can reduce isoflurane-induced neuronal apoptosis. The neuroprotective effect of miR-137 has been reported before, however, the effect of on isoflurane triggered neuronal apoptosis, and whether miR-137 is involved in the neuroprotection of DEX remain unclear. To investigate these doubts, we established an isoflurane exposure model in postnatal day 7 (P7) Sprague‒Dawley rats and the PC12 cells, containing a control group (CON), isoflurane group (ISO), DEX group (DEX) and DEX pretreatment group (DEX + ISO). We first confirmed that DEX attenuates isoflurane-induced hippocampal apoptosis. And we found DEX increased miR-137 and attenuated GSK-3ß levels in the DEX and DEX + ISO groups in the hippocampus and PC12 cells. In addition, the regulative relationship of miR-137 and GSK-3ß was confirmed using the TargetScan tool and dual-luciferase reporter assay. Moreover, miR-137 overexpression inhibited GSK-3ß and increased its downstream gene ß-catenin, whereas knockdown of miR-137 changed the GSK-3ß and ß-catenin expression oppositely. Upregulation of miR-137 increased the apoptosis-related genes and decreased the anti-apoptosis gene; however, knockdown of miR-137 produced the opposite results. This study suggested that DEX attenuated isoflurane-induced neuroapoptosis by upregulating the miR-137 mediated GSK-3ß/ß-catenin pathway in the developing rat hippocampus.

Molecular epidemiological characteristics and genetic evolutionary relationships of methicillin-resistant Staphylococcus aureus of different avian origins in Qingdao, China, using whole-genome sequencing.

Introduction: To understand the prevalence of avian methicillin-resistant Staphylococcus aureus (MRSA) and the current status of drug resistance in Qingdao, a comprehensive molecular epidemiological investigation and analysis of evolutionary relationships of MRSA isolates from broiler and layer chickens and waterfowl was conducted. Material and Methods: One hundred and two avian MRSA strains were identified by multi-locus sequence typing, staphylococcal protein A (spa) and staphylococcal cassette chromosome mec (SCCmec) typing, and whole-genome sequencing. Results: The sequence type (ST) 9-t899-SCCmec IVb type represented the highest proportion of avian-derived MRSA strains (71.57%), with ST398 type strains occasionally observed in broilers and waterfowl. The poultry-derived MRSA strains were all resistant to eight or more antimicrobials. Avian-derived MRSA strains carried 20 resistance genes, 109 virulence genes and 10 plasmids. Strains carrying the cfr oxazolidinone resistance gene were occasionally seen in broiler- and layer-derived MRSA. Single nucleotide polymorphism (SNP) core genome evolution and locus difference analysis showed that the closest strains were all of ST9-t899 type (to which also affiliated the highest number of strains) and this type occurred on all three kinds of poultry farm, but the SNP difference loci between strains of the same type ranged from 0 to 1472. Conclusion: The dominant type of MRSA from different poultry sources in Qingdao is ST9-t899-SCCmec IVb, which is commonly resistant to a variety of antimicrobial drugs and carries a variety of resistance genes and a large number of virulence genes. Sequence type 9-t899 type is widely spread among the three kinds of poultry investigated, but there are differences in affiliations.

Aflatoxin B1 disrupts testicular development via the cell cycle-related Ras/PI3K/Akt signaling in mice and pig.

Aflatoxins B1 (AFB1), a type I carcinogen widely present in the environment, not only poses a danger to animal husbandry, but also poses a potential threat to human reproductive health, but its mechanism is still unclear. To address this question, multi-omics were performed on porcine Sertoli cells and mice testis. The data suggest that AFB1 induced testicular damage manifested as decreased expression of GJA1, ZO1 and OCCLUDIN in mice (p < 0.01) and inhibition of porcine Sertoli cell proliferation. Transcriptomic analysis suggested changes in noncoding RNA expression profiles that affect the cell cycle-related Ras/PI3K/Akt signaling pathway after AFB1 exposure both in mice and pigs. Specifically, AFB1 caused abnormal cell cycle of testis with the characterization of decreased expressions of CCNA1, CCNB1 and CDK1 (p < 0.01). Flow cytometry revealed that the G2/M phase was significantly increased after AFB1 exposure. Meanwhile, AFB1 downregulated the expressions of Ras, PI3K and AKT both in porcine Sertoli cell (p < 0.01) and mice testis (p < 0.01). Metabolome analysis verified the alterations in the PI3K/Akt signaling pathway (p < 0.05). Moreover, the joint analysis of metabolome and microbiome found that the changes of metabolites were correlated with the expression of flora. In conclusion, we have demonstrated that AFB1 impairs testicular development via the cell cycle-related Ras/PI3K/Akt signaling.

[Applied anatomy of the stylomastoid foramina and its relation to the surrounding bone structures].

PURPOSE: To study the relationship between the stylomastoid foramen and surrounding bony structures, enrich anatomical data and provide reference for clinical surgery. METHODS: A total of 62 intact and dry adult skulls were selected. The shape of the stylomastoid foramen was observed, the diameter of the stylomastoid foramen, the distances from the posterolateral point and the anterior medial point to the surrounding bony structures were measured with a vernier caliper. SPSS 25.0 software package was used to analyze the data. RESULTS: There were four shapes of stylomastoid foramen, i.e., circular (61.29%), oval (29.84%), irregular (8.06%) and triangular (0.81%). The circular diameter was (2.80±0.61) mm, the oval long and short diameters were (4.43±0.96) and (2.79±0.60) mm. Distances from the posterolateral and anterior medial points of the stylomastoid foramen to the posterolateral point of the external opening of the carotid canal, the anterior medial point of the jugular foramen, the midline, the most anterior point of the foramen magnum, the posterior point of the great palatine foramen, the posterolateral point of the foramen lacerated, the foramen ovale, the posterolateral point of the foramen spinosum, the anterior point of the styloid process root, the outermost point of the tympanomastoid fissure and the tip of the mastoid process were (16.10±2.81), (24.01±2.65), (44.95±3.24), (45.10±2.71), (61.66±4.14), (35.56±4.35), (32.26±2.85), (29.12±3.40), (10.39±3.25), (9.49±2.24) and (12.01±2.79) mm; (12.80±2.41), (21.56±2.51), (42.96±3.97), (42.91±2.76), (58.97±3.97), (32.98±4.14), (29.20±2.77), (25.80±2.87), (7.37±2.33), (11.42±2.00) and (15.41±2.57) mm, respectively. Statistical analysis showed that there was no significant difference in the apertures and distances between the left and right side(P＞0.05). CONCLUSIONS: Most of the stylomastoid foramen are round and oval, understanding the distance between the foramen and surrounding bony structures is helpful for guiding clinical operations and enriching anatomical knowledge.

Characterization and genome analysis of Pseudomonas aeruginosa phage vB_PaeP_Lx18 and the antibacterial activity of its lysozyme.

A lytic Pseudomonas aeruginosa phage, vB_PaeP_Lx18 (Lx18), was isolated from the sewage of a dairy farm. Biological characterization revealed that Lx18 was stable from 40 °C to 60 °C and over a wide range of pH values from 4 to 10. It was able to lyse 63.6% (21/33) of the P. aeruginosa strains tested and was able to reduce and disperse biofilms, with a biofilm reduction rate of 76.8%. Whole-genome sequencing showed that Lx18 is a dsDNA virus with a genome of 42,735 bp and G+C content of 62.16%. The genome contains 54 open reading frames (ORFs), 28 of which have known functions, including DNA replication and modification, transcriptional regulation, structural and packaging proteins, and host cell lysis. No virulence or tRNA genes were identified. Phylogenetic analysis showed that phage Lx18 belongs to the genus Phikmvvirus. The lysozyme of Lx18, Lys18, was cloned and expressed. The combined action of Lys18 and ethylenediaminetetraacetic acid (EDTA) had antibacterial activity against Pseudomonas aeruginosa. The study of phage Lx18 and its lysozyme will provide basic information for further research on the treatment of Pseudomonas aeruginosa infections.

A C-terminally truncated mouse Best3 splice variant targets and alters the ion balance in lysosome-endosome hybrids and the endoplasmic reticulum.

The Bestrophin family has been characterized as Cl(-) channels in mammals and Na(+) channels in bacteria, but their exact physiological roles remian unknown. In this study, a natural C-terminally truncated variant of mouse Bestrophin 3 (Best3V2) expression in myoblasts and muscles is demonstrated. Unlike full-length Best3, Best3V2 targets the two important intracellular Ca stores: the lysosome and the ER. Heterologous overexpression leads to lysosome swelling and renders it less acidic. Best3V2 overexpression also results in compromised Ca(2+) release from the ER. Knocking down endogenous Best3 expression in myoblasts makes these cells more excitable in response to Ca(2+) mobilizing reagents, such as caffeine. We propose that Best3V2 in myoblasts may work as a tuner to control Ca(2+) release from intracellular Ca(2+) stores.

Di (2-ethylhexyl) Phthalate Exposure Impairs Growth of Antral Follicle in Mice.

Di (2-ethylhexyl) phthalate (DEHP) is a widely used plastic additive. As an environmental endocrine disruptor, it has been shown to be harmful to the mammalian reproductive system. Previous studies indicated that DEHP inhibited the development of mouse ovarian follicles. However, the mechanisms by which DEHP affects ovarian antral follicle development during the pre-puberty stage are poorly understand. Thus, we investigated the effects of direct DEHP exposure on antral follicle growth in pre-pubescent mice by use of intraperitoneal injection. Our results demonstrated that the percentage of large antral follicles was significantly reduced when mice were exposed to 20 or 40 µg/kg DEHP every 5 days from postnatal day 0 (0 dpp) to 15 dpp. In 20 dpp, we performed microarray of these ovaries. The microarray results indicated that mRNA levels of apoptosis related genes were increased. The mRNA levels of the apoptosis and cell proliferation (negative) related genes Apoe, Agt, Glo1 and Grina were increased after DEHP exposure. DEHP induced the differential gene expression of Hsp90ab1, Rhoa, Grina and Xdh which may play an important role in this process. In addition, TUNEL staining and immunofluorescence showed that DEHP exposure significantly increased the number of TUNEL, Caspase3 and γH2AX positive ovarian somatic cells within the mouse ovaries. Flow cytometer analyses of redox-sensitive probes showed that DEHP caused the accumulation of reactive oxygen species. Moreover, the mRNA expression of ovarian somatic cell antioxidative enzymes was down-regulated both in vivo and in vitro. In conclusion, our data here demonstrated that DEHP exposure induced oxidative stress and ovarian somatic cell apoptosis, and thus may impact antral follicle enlargement during the pre-pubertal stage in mice.

Cloning and in vitro function analysis of codon-optimized FatI gene.

Currently, n-3 polyunsaturated fatty acids (n-3 PUFAs) have attracted great attention because of their biological significance to organisms. In addition, PUFAs show an obvious impact on prevention and treatment of various diseases. Because n-3 PUFAs cannot be endogenously synthesized by mammals, mammals have to rely on a dietary supplement for sufficient supply. The finding and application of the fatty acid dehydrogenase I (FatI) gene are expected to change the current situation because it can convert n-6 polyunsaturated fatty acids (n-6 PUFAs) to n-3 PUFAs. Meanwhile, the gradual maturation of transgenic technology makes it possible to produce transgenic animals that can synthesize n-3 PUFAs by themselves. In this study, the DNA coding sequence of FatI was synthesized by a chemical method after codon optimization according to the mammal's codon bias. The synthesized DNA sequence was introduced into Boer goat fetal fibroblasts by the constructed recombinant eukaryotic expression vector pcDNA3.1(+)-FatI. Boer goat fetal fibroblasts were transfected by electroporation, and the stable transfected cell lines were obtained by G418 selection. Genomic DNA PCR and Southern blot were applied to verify that the foreign gene FatI was integrated into the genome of the Boer goat fibroblasts. RT-PCR results showed the expression of FatI gene at the mRNA level. The fatty acid profile of cells carrying the FatI gene revealed an increase in total n-3 PUFAs (from 0.61 to 0.95), but a decrease in n-6 PUFAs (from 10.34 to 9.85), resulting in a remarkable increase in the n-3:n-6 ratio (from 0.059 to 0.096). The n-3:n-6 ratio had a 63.49 percent increase, which is a precursor of the response of n-3 desaturase activity of the FatI gene. The study may provide a practical tool for producing transgenic animals that can produce n-3 PUFAs by themselves, and we hope that the application will lay the foundation for animals producing n-3 PUFAs, which will benefit human nutrition and wellness.

Anti-senescence effect of FatI gene in goat somatic cells.

The fatty acid dehydrogenase I (FatI) is able to express in mammalian cells and convert n-6 polyunsaturated fatty acids (PUFAs) to n-3 PUFAs. n-3 PUFA is an important component of the cell membrane and plays an important role in the prevention and control of a variety of human diseases. However, n-3 PUFAs cannot be endogenously synthesized by mammals because they lack the dehydrogenase that converts n-6 to n-3 PUFA. For the time being, gradually matured transgenic technology makes it possible to produce transgenic animals that are able to synthesize n-3 PUFAs by themselves. However, the transgenic technology itself may bring negative impacts. In this study, the eukaryotic expression vector pcDNA3.1-FatI was introduced into the genome of Boer goat fetal fibroblasts cultured in vitro, and the influence of biological characteristics of the fetal fibroblast was studied via overexpression of FatI. The results showed that the proliferation and apoptosis of cultured fetal fibroblast were not affected significantly by the overexpression of FatI using BrdU and TUNEL staining methods, respectively. Moreover, the overexpression of FatI significantly inhibited the senescence of somatic cells compared with enhanced green fluorescent protein (EGFP) transgenic cells (P < 0.01). Quantitative PCR revealed that the mRNA expression of P16 and P53 in the FatI transgenic cell group was significantly lower than that in the EGFP transgenic cell group (P < 0.01). In conclusion, the senescence of goat somatic cells was inhibited by the overexpression of the FatI gene.