TMEM41B Is an Interferon-Stimulated Gene That Promotes Pseudorabies Virus Replication.

Pseudorabies virus (PRV) is a double-stranded DNA virus that causes Aujeszky's disease and is responsible for economic loss worldwide. Transmembrane protein 41B (TMEM41B) is a novel endoplasmic reticulum (ER)-localized regulator of autophagosome biogenesis and lipid mobilization; however, the role of TMEM41B in regulating PRV replication remains undocumented. In this study, PRV infection was found to upregulate TMEM41B mRNA and protein levels both in vitro and in vivo. For the first time, we found that TMEM41B could be induced by interferon (IFN), suggesting that TMEM41B is an IFN-stimulated gene (ISG). While TMEM41B knockdown suppressed PRV proliferation, TMEM41B overexpression promoted PRV proliferation. We next studied the specific stages of the virus life cycle and found that TMEM41B knockdown affected PRV entry. Mechanistically, we demonstrated that the knockdown of TMEM41B blocked PRV-stimulated expression of the key enzymes involved in lipid synthesis. Additionally, TMEM41B knockdown played a role in the dynamics of lipid-regulated PRV entry-dependent clathrin-coated pits (CCPs). Lipid replenishment restored the CCP dynamic and PRV entry in TMEM41B knockdown cells. Together, our results indicate that TMEM41B plays a role in PRV infection via regulating lipid homeostasis. IMPORTANCE PRV belongs to the alphaherpesvirus subfamily and can establish and maintain a lifelong latent infection in pigs. As such, an intermittent active cycle presents great challenges to the prevention and control of PRV disease and is responsible for serious economic losses to the pig breeding industry. Studies have shown that lipids play a crucial role in PRV proliferation. Thus, the manipulation of lipid metabolism may represent a new perspective for the prevention and treatment of PRV. In this study, we report that the ER transmembrane protein TMEM41B is a novel ISG involved in PRV infection by regulating lipid synthesis. Therefore, our findings indicate that targeting TMEM41B may be a promising approach for the development of PRV vaccines and therapeutics.

The association of admission ionized calcium with outcomes of thrombolysed patients with anterior circulation ischemic stroke.

BACKGROUND AND PURPOSE: The relationship between ionized calcium and prognosis of ischemic stroke is controversial. We aim to determine the relationship of admission ionized calcium levels with acute ischemic stroke (AIS) after intravenous thrombolysis (IVT). METHODS: Consecutive anterior circulation AIS patients treated with recombinant tissue plasminogen activator (rt-PA) were retrospectively enrolled. According to ionized calcium quartiles, the patients were divided into four groups and clinical data were analyzed between groups. Ionized calcium was entered into logistic regression analysis in two models, separately: model 1, calcium as a continuous variable (per 1-mmol/L increase), and model 2, calcium as the four-categorized variable (being collapsed into quartiles: Q1-Q4). Early neurologic improvement (ENI) was defined as improvement of four or more points at 24 h after intravenous rt-PA, while long-term good outcome as the modified Rankin Scale (mRS) 0-1 at 90 days. RESULTS: A total of 546 patients met the study criteria (mean age was 63.51 ± 11.26 years and 365 [66.8%] were men). The median admission National Institute of Health Stroke Scale was 9 (range 4 to 15). When not adjusted, in model 1: ionized calcium was related to good outcome (odds ratio [OR] 69.061, 95%CI: 1.638-2911.111, p=0.027), but not ENI (OR 14.097, 95%CI: 0.133-1492.596, p=0.266); in model 2: compared with Q4, while good outcome was less common in Q1 (OR 0.623, 95%CI: 0.388-0.999, p=0.049). After adjusting for confounding factors, calcium in Q2 (OR 0.502, 95%CI: 0.253-0.997, p=0.049) was independently associated with ENI, but no matter as a continuous variable or categorized variable, ionized calcium displayed no association with a good outcome. CONCLUSION: The current results found that ionized calcium might be associated with early neurological improvement, but had no association with 3 months' outcome in anterior circulation AIS patients after IVT.

Pseudorabies Virus Inhibits Expression of Liver X Receptors to Assist Viral Infection.

Pseudorabies virus (PRV) is a contagious herpesvirus that causes Aujeszky's disease and economic losses worldwide. Liver X receptors (LXRs) belong to the nuclear receptor superfamily and are critical for the control of lipid homeostasis. However, the role of LXR in PRV infection has not been fully established. In this study, we found that PRV infection downregulated the mRNA and protein levels of LXRα and LXRß in vitro and in vivo. Furthermore, we discovered that LXR activation suppressed PRV proliferation, while LXR inhibition promoted PRV proliferation. We demonstrated that LXR activation-mediated reduction of cellular cholesterol was critical for the dynamics of PRV entry-dependent clathrin-coated pits. Replenishment of cholesterol restored the dynamics of clathrin-coated pits and PRV entry under LXR activation conditions. Interestingly, T0901317, an LXR agonist, prevented PRV infection in mice. Our results support a model that PRV modulates LXR-regulated cholesterol metabolism to facilitate viral proliferation.

EGCG Restricts PRRSV Proliferation by Disturbing Lipid Metabolism.

Porcine reproductive and respiratory syndrome virus (PRRSV) infection leads to late-term reproductive failure and respiratory illness that affect the global swine industry. Epigallocatechin gallate (EGCG) is a polyphenolic compound from green tea that exerts antiviral activity against diverse viruses. This study aimed to report an uncharacterized mechanism of how EGCG restricted PRRSV proliferation. EGCG showed no significant effects on cell viability, cell cycle progression, and apoptosis in porcine alveolar macrophages and MARC-145 cells. The treatment of cells with EGCG attenuated the replication of both highly pathogenic and less pathogenic PRRSV in vitro. The viral life cycle analysis demonstrated that EGCG affected PRRSV replication and assembly, but not viral attachment, entry, or release. Interestingly, EGCG treatment abrogated the increased lipid droplets formation and lipid content induced by PRRSV infection. We further demonstrated that EGCG blocked PRRSV-stimulated expression of the key enzymes in lipid synthesis. In addition, EGCG attenuated PRRSV-induced autophagy that is critical for PRRSV proliferation. The supplementation of oleic acid restored PRRSV replication and assembly under EGCG treatment. Together, our results support that EGCG inhibits PRRSV proliferation through disturbing lipid metabolism. IMPORTANCE Porcine reproductive and respiratory syndrome virus (PRRSV) is an enveloped single-positive-stranded RNA virus that causes acute respiratory distress in piglets and reproductive failure in sows, resulting in huge economic losses to the global swine industry. Several lines of evidence have suggested the crucial roles of lipids in PRRSV proliferation. Our previous report demonstrated that PRRSV activated lipophagy to facilitate viral replication through downregulating the expression of N-Myc downstream-regulated gene 1. The manipulation of lipid metabolism may be a new perspective to prevent PRRSV spread. In the present study, we reported that epigallocatechin-3-gallate (EGCG), the major component of green tea catechins, significantly attenuated PRRSV infection through inhibiting lipid synthesis and autophagy. Given that natural products derived from plants have helped in the prevention and treatment of various infectious diseases, EGCG has a great potential to serve as a safe and environmentally friendly natural compound to treat PRRSV infection.

An effective inactivant based on singlet oxygen-mediated lipid oxidation implicates a new paradigm for broad-spectrum antivirals.

Emerging viral pathogens cause substantial morbidity and pose a severe threat to health worldwide. However, a universal antiviral strategy for producing safe and immunogenic inactivated vaccines is lacking. Here, we report an antiviral strategy using the novel singlet oxygen (1O2)-generating agent LJ002 to inactivate enveloped viruses and provide effective protection against viral infection. Our results demonstrated that LJ002 efficiently generated 1O2 in solution and living cells. Nevertheless, LJ002 exhibited no signs of acute toxicity in vitro or in vivo. The 1O2 produced by LJ002 oxidized lipids in the viral envelope and consequently destroyed the viral membrane structure, thus inhibiting the viral and cell membrane fusion necessary for infection. Moreover, the 1O2-based inactivated pseudorabies virus (PRV) vaccine had no effect on the content of the viral surface proteins. Immunization of mice with LJ002-inactiviated PRV vaccine harboring comparable antigen induced more neutralizing antibody responses and efficient protection against PRV infection than conventional formalin-inactivated vaccine. Additionally, LJ002 inactivated a broad spectrum of enveloped viruses. Together, our results may provide a new paradigm of using broad-spectrum, highly effective inactivants functioning through 1O2-mediated lipid oxidation for developing antivirals that target the viral membrane fusion process.

Isolation and purification of Codonopsis pilosula polysaccharide and its anti-oxidant activity / 中草药

Objective: To isolate and purify the Codonopsis pilosula polysaccharide (CPP) and investigate its structure characterization and anti-oxidant activity. Methods: CPP was extracted by heating reflux, crud polysaccharide was refined and then isolated by hollow fiber ultrafiltration experimental device, the anti-oxidant activity of polysaccharides was studied by in vivo and in vitro methods. Results: CPP was purified and obtained three ingredients named CPP1, CPP2, and CPP3. The anti-oxidant activity in vitro showed that CPP3 had the strongest scavenging ability on DPPH, hydroxyl radical, and superoxide anion. In vivo study showed that high dose group of CPP3 had obvious protective effect on the mice induced by D-galactose. Conclusion: CPP has obvious anti-oxidant function. This study provides the theoretical basis for the development of CPP functional food.

Optimization of extraction process of Codonopsis pilosula polysaccharides and study on its composition / 中草药

Objective: To optimize the extraction process of polysaccharide from Codonopsis pilosula and determine the monosaccharide composition and molecular weight distribution, in order to provide the basis for further separation of C. pilosula polysaccharide. Methods: The content of polysaccharide in C. pilosula was determined by phenol sulfuric acid method, the extraction process of polysaccharide was optimized by orthogonal test. C. pilosula polysaccharides were prepared from crude polysaccharides by deproteinization, decoloration, dialysis, and lyophilization, then monosaccharide composition and mean molecular mass of C. pilosula polysaccharides were analyzed by high performance liquid chromatography (HPLC) and high performance gel permeation chromatography (HPGPC). Results: The extraction temperature was 85 ℃, the extraction time was 1.5 h per time, twice, and solid to liquid ratio was 1:12. Under these conditions, the yield of polysaccharides was 22.57%. The polysaccharides were consisted by glucuronic acid, aminogalactose, xylose, and small quantities of mannose, the average molecular mass was 21 498. Conclusion: This study provides a theoretical basis for the classification and activity of polysaccharide from C. pilosula.

The relationship between a nucleotide substitution of S100A8 gene and aggressive periodontitis / 中华口腔医学杂志

<p><b>OBJECTIVES</b>To screen polymorphisms in the upstream region of S100A8 gene and to detect whether the polymorphisms were associated with aggressive periodontitis.</p><p><b>METHODS</b>Thirty aggressive periodontitis patients and twenty-eight healthy controls were recruited for the study with informed consent. All subjects were of Chinese descent and systemically healthy. The regions about 800 bp upstream from the ATG start codon in exon 2 of the S100A8 gene of 10 patients and 8 controls were amplified by polymerase chain reaction (PCR) and analyzed by direct sequencing. A single nucleotide polymorphism (SNP) at 94 bp upstream from the ATG start codon was selected, and then the shorter regions (about 250 bp upstream from the ATG start codon) of the rest subjects were also amplified by PCR and analyzed by direct sequencing. The frequency of the SNP and the distribution of the genotype were detected and compared between the two groups.</p><p><b>RESULTS</b>A nucleotide substitution (A-->G) at 94 bp upstream from the ATG start codon was demonstrated in Chinese, which was in a cis-acting element, named gamma interferon response element (gamma-IRE) in intron 1 of S100A8 gene. All of the subjects that carried the polymorphism were heterozygous. There was no statistically significant difference in the frequency of allele 2 (corresponding to the nucleotide G) between patients and controls (11.7% vs. 17.9%, chi2 = 0.887, P > 0.05). The prevalence of the heterozygous genotype was 23.2% and 35.7% (chi2 = 1.07, P > 0.05) in patients and controls, respectively.</p><p><b>CONCLUSIONS</b>This is the first report that a nucleotide substitution of S100A8 gene was demonstrated in Chinese. The frequencies of allele 2 and heterozygous genotype were lower in patients, but there is no statistically significant difference between the aggressive periodontitis patients and healthy controls in this preliminary study.</p>