Pneumocytes are distinguished by highly elevated expression of the ER stress biomarker GRP78, a co-receptor for SARS-CoV-2, in COVID-19 autopsies

Vaccinations are widely credited with reducing death rates from COVID-19, but the underlying host-viral mechanisms/interactions for morbidity and mortality of SARS-CoV-2 infection remain poorly understood. Acute respiratory distress syndrome (ARDS) describes the severe lung injury, which is pathologically associated with alveolar damage, inflammation, non-cardiogenic edema, and hyaline membrane formation. Because proteostatic pathways play central roles in cellular protection, immune modulation, protein degradation, and tissue repair, we examined the pathological features for the unfolded protein response (UPR) using the surrogate biomarker glucose-regulated protein 78 (GRP78) and co-receptor for SARS-CoV-2. At autopsy, immunostaining of COVID-19 lungs showed highly elevated expression of GRP78 in both pneumocytes and macrophages compared with that of non-COVID control lungs. GRP78 expression was detected in both SARS-CoV-2-infected and un-infected pneumocytes as determined by multiplexed immunostaining for nucleocapsid protein. In macrophages, immunohistochemical staining for GRP78 from deceased COVID-19 patients was increased but overlapped with GRP78 expression taken from surgical resections of non-COVID-19 controls. In contrast, the robust in situ GRP78 immunostaining of pneumocytes from COVID-19 autopsies exhibited no overlap and was independent of age, race/ethnicity, and gender compared with that from non-COVID-19 controls. Our findings bring new insights for stress-response pathways involving the proteostatic network implicated for host resilience and suggest that targeting of GRP78 expression with existing therapeutics might afford an alternative therapeutic strategy to modulate host-viral interactions during SARS-CoV-2 infections.     

FTIR determination of free fatty acids in fish oils intended for biodiesel production

Biodiesel is commonly derived from vegetable oils and animal (fish and livestock) fats by alkali- or lipase-catalyzed transesterification reactions. Since free fatty acid (FFA) content is a critical parameter in the conversion of fish oils to methyl esters, the performance of a Fourier transform infrared (FTIR) spectroscopic method was assessed as an alternative to the conventional AOCS titrimetric method. The FTIR method involves the simultaneous extraction of FFAs and their stoichiometric conversion to their salts using a weak base, sodium hydrogen cyanamide (NaHNCN) dissolved in methanol, followed by measurement of the carboxylate band, ν(COO^?), at 1573 cm^?1 relative to a baseline at 1820 cm^?1 in the differential spectrum of the methanol extract. With minor modifications, this method was found to be capable of responding linearly to oleic acid (0–6.5%) addition, producing a FFA calibration equation having a S.D. of ±0.014% FFA. FTIR and titrimetric analytical results were compared for samples prepared by standard addition as well as for fish oils extracted from salmon skin which had been stored up to 120 days at ?20 °C. Both methods responded in a comparable manner; however, the FTIR method was more reproducible and accurate as well as simpler to carry out and was deemed to be a better primary method than the titrimetric method. The FFA content of Atlantic salmon (Salmo salar) skin lipids increased linearly from ～0.6% to 4.5% within 120 days, likely as a result of autoxidation. It was concluded that the NaHNCN-based FTIR method is a flexible, viable instrumental alternative to the AOCS titrimetric procedure for the determination of FFA content of fish tissue lipids destined for biodiesel production.     

HaloTag7: A genetically engineered tag that enhances bacterial expression of soluble proteins and improves protein purification

Over-expression and purification of soluble and functional proteins remain critical challenges for many aspects of biomolecular research. To address this, we have developed a novel protein tag, HaloTag7, engineered to enhance expression and solubility of recombinant proteins and to provide efficient protein purification coupled with tag removal. HaloTag7 was designed to bind rapidly and covalently with a unique synthetic linker to achieve an essentially irreversible attachment. The synthetic linker may be attached to a variety of entities such as fluorescent dyes and solid supports, permitting labeling of fusion proteins in cell lysates for expression screening, and efficient capture of fusion proteins onto a purification resin. The combination of covalent capture with rapid binding kinetics overcomes the equilibrium-based limitations associated with traditional affinity tags and enables efficient capture even at low expression levels. Following immobilization on the resin, the protein of interest is released by cleavage at an optimized TEV protease recognition site, leaving HaloTag7 bound to the resin and pure protein in solution. Evaluation of HaloTag7 for expression of 23 human proteins in Escherichia coli relative to MBP, GST and His₆Tag revealed that 74% of the proteins were produced in soluble form when fused to HaloTag7 compared to 52%, 39% and 22%, respectively, for the other tags. Using a subset of the test panel, more proteins fused to HaloTag7 were successfully purified than with the other tags, and these proteins were of higher yield and purity.     

Effects of an exotic invasive macrophyte (tropical signalgrass) on native plant community composition,species richness and functional diversity

1. The issue of freshwater species being threatened by invasion has become central in conservation biology because inland waters exhibit the highest species richness per unit area, but apparently have the highest extinctions rates on the planet. 2. In this article, we evaluated the effects of an exotic, invasive aquatic grass (Urochloa subquadripara– tropical signalgrass) on the diversity and assemblage composition of native macrophytes in four Neotropical water bodies (two reservoirs and two lakes). Species cover was assessed in quadrats, and plant biomass was measured in further quadrats, located in sites where tropical signalgrass dominated (D quadrats) and sites where it was not dominant or entirely absent (ND quadrats). The effects of tropical signalgrass on macrophyte species richness, Shannon diversity and number of macrophyte life forms (a surrogate of functional richness) were assessed through regressions, and composition was assessed with a DCA. The effects of tropical signalgrass biomass on the likelihood of occurrence of specific macrophyte life forms were assessed through logistic regression. 3. Tropical signalgrass had a negative effect on macrophyte richness and Shannon and functional diversity, and also influenced assemblage composition. Emergent, rooted with floating stems and rooted submersed species were negatively affected by tropical signalgrass, while the occurrence of free‐floating species was positively affected. 4. Our results suggest that competition with emergent species and reduction of underwater radiation, which reduces the number of submersed species, counteract facilitation of free‐floating species, contributing to a decrease in plant diversity. In addition, homogenisation of plant assemblages shows that tropical signalgrass reduces the beta diversity in the macrophyte community. 5. Although our results were obtained at fine spatial scales, they are cause for concern because macrophytes are an important part of freshwater diversity.     

Biomarker assessment of organic matter sources and degradation in Canadian High Arctic littoral sediments

This paper presents an uncomplicated approach to improve estimates of groundwater nutrient load to a marine embayment. A two-dimensional chemical profile of shallow groundwater was analysed in a sandy beach in three seasons (early summer, late summer and mid winter) and an adjusted estimate of groundwater nutrient discharge was derived that accounts for a complex biogeochemical environment and non-conservative behaviour of nutrients in the pre-discharge beach groundwater. The study was conducted at Cockburn Sound, Western Australia, where there has been significant groundwater contamination and associated marine ecological degradation. Losses in nitrogen and increases in phosphorus were observed along the discharge pathway beyond that expected from mixing with marine water, and the changes were attributed to chemically and biologically mediated reactions. A slow groundwater velocity (0.14–0.18 m day^?1), high organic carbon (TOC = 0.35–4.9 mmol l^?1, DOC = 0.28–4.6 mmol l^?1) and low to sub-oxic conditions (DO = 0.4–24% saturation) were deemed suitable for chemically and biologically mediated reactions to occur and subsequently alter regional estimates of groundwater nutrient concentration. Accounting for this environment, groundwater loads were calculated that were 1–2 orders of magnitude less than previous regional-based estimates: 0.4–13 kg NO _x ^?  day^?1, 0.2–24 kg NH₄ ⁺ day^?1 and 0.004–0.8 kg FRP day^?1. This paper applies knowledge of recent research and presents scope to marine managers or modellers to account for groundwater inputs to the marine environment.     

Discovery of a series of indan carboxylic acid glycogen phosphorylase inhibitors

A series of carboxylic acid glycogen phosphorylase inhibitors, which have potential as oral antidiabetic agents, is described. Defining and applying simple physicochemical design criteria was used to assess the opportunity and to focus synthetic efforts on compounds with the greatest probability of success. The study led to compound 17, which exhibits a good balance of properties including potent inhibition of recombinant human liver glycogen phosphorylase in vitro, a good DMPK profile including excellent bioavailability and low clearance and good in vivo activity in a glucagon challenge model of diabetes in Zucker rats.     

Spirocyclic ureas: Orally bioavailable 11β-HSD1 inhibitors identified by computer-aided drug design

Structure-guided drug design led to the identification of a class of spirocyclic ureas which potently inhibit human 11β-HSD1 in vitro. Lead compound 10j was shown to be orally bioavailable in three species, distributed into adipose tissue in the mouse, and its (R) isomer 10j2 was efficacious in a primate pharmacodynamic model.     

长爪沙鼠线粒体DNA控制区全序列测定及分析

目的对长爪沙鼠线粒体DNA控制区全序列进行测定,并对其进行鉴定及进化分析。方法根据长爪沙鼠已知基因序列设计引物,采用PCR产物测序法,对所得的片段进行测序鉴定。结合已公布啮齿类动物D-loop区序列,分析其碱基组成、遗传距离、并基于最小进化法和UPGMA法构建系统进化树。结果获得长爪沙鼠D-loop区序列,其与家鼠、小家鼠和仓鼠平均同源性为58%;碱基组成分析显示,长爪沙鼠与啮齿类动物有相似的碱基组成和碱基偏离,其A-skew和G-skew分别为0.0047和-0.28。进化分析结果显示,长爪沙鼠与家鼠（0.35）、黑家鼠（0.38）和仓鼠（0.39）具有较近的遗传距离,其分化顺序为跳鼠、蔗鼠、长爪沙鼠、仓鼠、家鼠和小家鼠。结论本研究获得长爪沙鼠D-loop区全序列,确定了长爪沙鼠与仓鼠、家鼠、小家鼠及其它啮齿动物的进化关系,为长爪沙鼠进化研究、线粒体的结构和功能研究奠定基础。     

Efficacy and safety of combined prolonged-release oxycodone and naloxone in the management of moderate/severe chronic non-malignant pain: results of a prospectively designed pooled analysis of two randomised,double-blind clinical trials

Background  

Two randomised 12-week, double-blind, parallel-group, multicenter studies comparing oxycodone PR/naloxone PR and oxycodone PR alone on symptoms of opioid-induced bowel dysfunction in patients with moderate/severe non-malignant pain have been conducted.