Acute effects of one-leg standing on arterial stiffness in older women: Role of the vision condition and standing dose.

Objective: One-leg standing has been used exclusively for static balance testing and training purposes. We investigated the acute effects of one-leg standing with open or closed eyes on arterial stiffness in older women and explored the role of standing dose in arterial stiffness regulation. Methods: Eighteen older women (60 ± 2 years) underwent non-intervention control (CON), one-leg standing with open eyes for 2 × 3 min (SO2), and one-leg standing with closed eyes for 1 × 3 min (SC1), 2 × 3 min (SC2), and 3 × 3 min trials (SC3) in a randomized self-controlled crossover fashion. Arterial stiffness in the cardio-ankle vascular index (CAVI) was measured at baseline (BL), immediately (0 min), and 10 and 20 min after standing. CAVI changes from BL in the same trial (⊿CAVI) were used for analysis. Results: ⊿CAVI of the non-standing and standing side did not change with time in CON and SO2 trials. In SC1, SC2, and SC3 trials, ⊿CAVI of the standing side decreased significantly at 0 min compared to their corresponding BL (p < 0.01) and reverted gradually to the BL level afterward, with ⊿CAVI of the non-standing side undergoing no changes. At the time point of 0 min, only in the SC2 trial, ⊿CAVI of the standing side was significantly lower than that of CON (p < 0.01). Conclusion: One-leg standing with closed eyes, but not with open eyes, resulted in transient arterial stiffness improvement in older women. The improvement was restricted to standing leg, and the moderate standing dose had maximal benefit on arterial stiffness.

Identification of host cell proteins that interact with the M protein of Pigeon paramyxovirus type 1.

Pigeon paramyxovirus type 1 (PPMV-1) belongs to the avian paramyxovirus type 1 group of viruses, which can cause tremors, torticollis, and respiratory signs in domestic and wild pigeons. The M protein of PPMV-1 is a multifunctional structural protein. It not only helps in the assembly, budding, and positioning of the virus but also inhibits the host's immune response and promotes replication of the virus in the host. In this study, the GST pull-down method was used to screen host proteins that interact with PPMV-1 M protein, and then mass spectrometry (MS) was used to analyse the screened host proteins. Enrichment analysis of the differentially expressed genes showed that the 77 screened proteins were highly associated with the gene ontology categories: protein synthesis, metabolism, and cell signalling pathway transduction. We selected NIMA-related kinase 7 (NEK7) as the candidate protein for co-localization analysis and co-immunoprecipitation verification. The results revealed that PPMV-1 M protein interacts with NEK7 of the host cell. This interactome study of PPMV-1 M protein will serve to clarify its function during viral replication and will provide a crucial theoretical basis for studying the pathogenic mechanism of PPMV-1.

[Purification of mammalian orthoreovirus serotype 3 Dearing Sigma1 protein and preparation of polyclonal antibody].

Objective To prepare T3Dsigma1 polyclonal antibody with purified sigma1 fusion protein of mammalian orthoreovirus (MRV) serotype 3 Dearing (T3D). Methods The recombinant plasmid T3DS1-pET28a was transformed into Rosetta (DE3) competent cells, and the isopropyl-ß-D-thiogalactopyranoside (IPTG) was used to induce a large amount of target proteins which were subjected to purification by histidine-tagged Ni-IDA chromatography column to obtain the T3Dsigma1 fusion protein. New Zealand white rabbits were immunized with the purified protein to prepare polyclonal antibodies specific against sigma1 protein. The titer of polyclonal antibody was detected by indirect ELISA, and the specificity by Western blot analysis and indirect immunofluorescence assay (IFA). Results The relative molecular mass (Mr) of T3Dsigma1 fusion protein, mainly in the form of inclusion body, was about 32 000. The fusion protein was purified by denaturation and renaturation. The polyclonal antibody with the titer greater than 1:106 was prepared by immunizing New Zealand white rabbits and detected by Western blot analysis and IFA. Conclusion The T3Dsigma1 polyclonal antibody with high titer and sensitivity was prepared.

Analysis of the intestinal microbial community altered during rotavirus infection in suckling mice.

BACKGROUND: Rotavirus (RV) is a principal cause of diarrhea. However, there is a limited understanding regarding alteration of the gut microbial community structure and abundance during RV infection. This study was to characterize any potential associations between RV infection and the intestinal microbiota. METHODS: Suckling mice were divided into normal group (NC) and infected group (RV) randomly. All of the suckling mice were euthanized four days post-RV infection. The virus titer was counted as fluorescent focus assay, and viral load was quantified by QPCR. Five sucking mice were randomly selected from each RV group and NC group for sample collection and pathological analysis. Mixed intestinal contents of the colon and rectum were collected from all of the suckling mice. To investigate the detailed relationship between RV infection and intestinal microbiota, the composition and distribution of intestinal microbiota from suckling mice were first analyzed using 16S rRNA sequencing technology. RESULTS: The results of the pathological characteristics showed that vacuolar degeneration, vasodilation, hyperemia, and destruction of the intestinal epithelium were apparent in the RV group. Representative genera from Lactobacillus and Fusobacterium were enriched in the NC group, while the Enterococcus and Escherichia/Shigella genera were enriched in the RV group. Helicobacter, Alloprevotrlla, Brevundimonas, Paenibacillus, and Parabacteroides were completely undetectable in the RV group. The predicted intestinal flora metabolic function results showed that "carbohydrate metabolism" and "lipid metabolism" pathways were significantly enriched within the NC group. A significant difference has been observed in the gut microbiota composition between the two groups. CONCLUSIONS: Our results demonstrated a significant difference in the gut microbiota composition in RV-infected suckling mice as compared to the RV un-infected suckling mice group. This work may provide meaningful information regarding the bacterial genera changed during RV infection. Moreover, the changes in these bacteria may be related with the replication and pathogenesis of RV infection.

Preparation of a polyclonal antibody against the non-structural protein, NSs of SFTSV.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is newly discovered virus which is the member of the order Bunyavirales, family phenuiviridae, phlebovirus genus. Its genome is composed of 3 segments of negative-sense RNA L, M and S. NSs is a non structure protein encoded by S segment which is important for viral replication and virulence. NSs protein of SFTSV is only involved in the regulation of host innate immune responses and suppression of IFN-promoter activities. So, the exact functions of this protein need to be studied deeply. To understand the exact role of NSs from SFTSV in viral replication and host immune response, a qualified antibody against this protein is required. In this study, NSs gene of SFTSV, was cloned into a bacterial expression vector (pGEX-6P-1) and the recombinant plasmid was transformed into Escherichia coli BL21 (DE3) cells. The SFTSV NSs fusion protein was purified using Glutathione Sepharose 4B and utilized as an antigen to immunize rabbits and obtain an anti-SFTSV NSs polyclonal antibody. Proper expression of the fusion protein and polyclonal antibody specificity were confirmed by western blotting and immunofluorescence analyses. The polyclonal antibody recognized NSs from SFTSV specifically. This is the first report that NSs can form viroplasm-like structures not only in infected cells but also in transfected cells with NSs plasmids. This polyclonal antibody will be useful for future studies of NSs functions.

Characterization of a G9 group A rotavirus reassortant strain detected in Jinzhou, China, in 2018-2019.

A novel rotavirus A (RVA) strain (JZ) was detected in RVA-positive stool specimens from three pediatric patients in Jinzhou, Liaoning Province, in 2018-2019. The electrophoresis pattern of the JZ strain showed a long electropherotype. The genomic constellation G9-P[8]-I1-R1-C1-M1-A1-N1-T1-E2-H1 was detected, suggesting that a new inter-genogroup reassortment had occurred. Whole-genome sequencing showed that the JZ isolates were identical to each other. Phylogenetic analysis revealed that VP7 and VP4 clustered in lineages G9-VI and P[8]-3, respectively. JZ strain-specific amino acid substitutions were detected in VP7, VP4 and NSP4. This study provides information on the epidemiology and characteristics of G9 strains circulating in China.

The effects of autophagy on the replication of Nelson Bay orthoreovirus.

BACKGROUND: Nelson Bay orthoreovirus (NBV) was first isolated over 40 years ago from a fruit bat in Australia. Normally, NBV does not cause human diseases, but recently several NBV strains have been associated with human respiratory tract infections, thus attracting clinical attention. Autophagy, an evolutionarily conserved process in eukaryotic cells, degrades intracellular substrates, participates in multiple physiological processes, and maintains cellular homeostasis. In addition, autophagy is intimately involved in viral infection. METHODS: A new strain of NBV, isolated from a patient with a respiratory tract infection who returned to Japan from Bali, Indonesia, in 2007, was used in this study. NBV was rescued using a reverse genetics system involving cotransfection of BHK cells with 11 plasmids (pT7-L1 MB, pT7-L2 MB, pT7-L3 MB, pT7-M1 MB, pT7-M2 MB, pT7-M3 MB, pT7-S1 MB, pT7-S2 MB, pT7-S3 MB, pT7-S4 MB, and pcDNA3.1-T7), yielding NBV-MB. Recovered viruses were confirmed by immunofluorescence. The effect of NBV-MB on autophagy was evaluated by measuring the LC3-I/II proteins by immunoblot analysis after infection of BHK cells. Furthermore, after treatment with rapamycin (RAPA), 3-methyladenine (3-MA), chloroquine (CQ), or plasmid (GFP-LC3) transfection, the changes in expression of the LC3 gene and the amount of LC3-I/II protein were examined. In addition, variations in viral titer were assayed after treatment of BHK cells with drugs or after transfection with plasmids pCAGM3 and pCAGS3, which encode virus nonstructural proteins µNS and σNS, respectively. RESULTS: NBV-MB infection induced autophagy in host cells; however, the level of induction was dependent on viral replication. Induction of autophagy increased viral replication. By contrast, inhibiting autophagy suppressed NBV replication, albeit not significantly. The NBV-MB nonstructural protein µNS was involved in the induction of autophagy with viral infection. CONCLUSIONS: NBV-MB infection triggered autophagy. Also, the NBV nonstructural protein µNS may contribute to augmentation of autophagy upon viral infection.

[Purification of Nelson Bay virus S3 protein and preparation of rat polyclonal antibody].

Objective To prepare a polyclonal antibody against S3 of Nelson Bay virus (NBV). Methods The E.coli BL21 (DE3) competent cells were transfected with the constructed Pris His MB-S3 recombinant plasmid. Protein expression was induced by IPTG. The target protein was purified by Ni-NTA column affinity chromatography to obtain a large amount of fusion recombinant protein, which was tested and used as the antigen for producing the S3 polyclonal antibody in rats. Results The relative molecular mass (Mr) of Pris His MB-S3 protein was around 39 000, in the form of inclusion bodies. NBV S3 polyclonal antibody was successfully produced from the immunized rats. The titer of the antibody was up to 1:64 000 determined by indirect ELISA. The indirect immunofluorescence assay verified that the S3 protein was successfully expressed in cells and distributed in a granular form in the cytoplasm. Conclusion The highly reactive and specific S3 protein polyclonal antibody is successfully prepared.

Single-nucleotide polymorphisms and activity analysis of the promoter and enhancer of the pig lactase gene.

Lactose intolerance in northern Europeans is strongly associated with a single-nucleotide polymorphism (SNP) located 14 kb upstream of the human lactase gene: -13,910 C/T. We examined whether SNPs in the 5' flanking region of the pig lactase gene are similar to those in the human gene and whether these polymorphisms play a functional role in regulating pig lactase gene expression. The 5' flanking region of the lactase gene from several different breeds of pigs was cloned and analyzed for gene regulatory activity of a luciferase reporter gene. One SNP was found in the enhancer region (-797 G/A) and two were found in the promoter region (-308G/C and -301 A/G). The promoter C-308,G-301(Pro-CG) strongly promotes the expression of the lactase gene, but the promoter G-308,A-301(Pro-GA) does not. The enhancer A-797(Enh-A) genotype for Pro-GA can significantly enhance promoter activity, but has an inhibitory effect on Pro-CG. The Enhancer G-797(Enh-G) has a significant inhibitory effect on both promoters. In conclusion, the order of effectiveness on the pig lactase gene is Enh-A+Pro-GA>Enh-A/G+Pro-CG>Enh-G+Pro-GA.

Effect of degree and direction of rotation in egocentric mental rotation of hand: an event-related potential study.

We explored the neural mechanisms of mental rotation of hand, which may invoke a mental transformation of viewer's own hands. It was found that, when a hand picture was presented at an orientation rotated from the upright orientation, participants' performance in making left or right hand judgment was affected by the degree and direction of rotation, with the direction effect being implicated as the evidence for egocentric mental rotation. Our event-related potentials measure supported the idea that amplitude modulation in the parietal cortex is a psychophysiological marker of the mental rotation of hand. Furthermore, the rotation-direction-dependent modulation of a positive wave was identified as possible neural correlate for the egocentric nature of such mental rotation.