Inhibition of Nur77 expression and translocation by compound B6 reduces ER stress and alleviates cigarette smoke-induced inflammation and injury in bronchial epithelial cells

Chronic obstructive pulmonary disease (COPD) is a leading cause of death worldwide with inflammation and injury in airway epithelial cells. However, few treatment options effectively reduce severity. We previously found that Nur77 is involved in lipopolysaccharide-induced inflammation and injury of lung tissue. Here, we established an in vitro model of COPD-related inflammation and injury in 16-HBE cells induced by cigarette smoke extract (CSE). In these cells, Nur77 expression and localization to the endoplasmic reticulum (ER) increased following CSE treatment, as did ER stress marker (BIP, ATF4, CHOP) expression, inflammatory cytokine expression, and apoptosis. The flavonoid derivative, named B6, which was shown to be a modulator of Nur77 in previous screen, molecular dynamics simulation revealed that B6 binds strongly to Nur77 through hydrogen bonding and hydrophobic interactions. Treating CSE-stimulated 16-HBE cells with B6 resulted in a reduction of both inflammatory cytokine expression and secretion, as well as attenuated apoptosis. Furthermore, B6 treatment resulted in a decrease in Nur77 expression and translocation to the ER, which was accompanied by a concentration-dependent reduction in the expression of ER stress markers. Meanwhile, B6 played a similar role in CSE-treated BEAS-2B cells. These combined effects suggest that B6 could inhibit inflammation and apoptosis in airway epithelial cells after cigarette smoke stimulation, and support its further development as a candidate intervention for treating COPD-related airway inflammation

Improved CAR internalization and recycling through transmembrane domain optimization reduces CAR-T cytokine release and exhaustion

BackgroundAnti-CD19 chimeric antigen receptor T (CAR-T) cell therapy has proven effective for treating relapsed or refractory acute B cell leukemia. However, challenges such as cytokine release syndrome, T cell dysfunction, and exhaustion persist. Enhancing CAR-T cell efficacy through changing CAR internalization and recycling is a promising approach. The transmembrane domain is the easiest motif to optimize for modulating CAR internalization and recycling without introducing additional domains, and its impact on CAR internalization and recycling has not yet been thoroughly explored. In this study, we aim to enhance CAR-T cell function by focusing on the solely transmembrane domain design.MethodsUtilizing plasmid construction and lentivirus generation, we get two different transmembrane CAR-T cells [19CAR-T(1a) and 19CAR-T(8α)]. Through co-culture with tumor cells, we evaluate CAR dynamic change, activation levels, exhaustion markers, mitochondrial function, and differentiation in both CAR-T cells. Furthermore, immunofluorescence microscopy analysis is performed to reveal the localization of internalized CAR molecules. RNA sequencing is used to detect the transcriptome of activated CAR-T cells. Finally, a mouse study is utilized to verify the anti-tumor efficacy of 19CAR-T(1a) cells in vivo.ResultsOur findings demonstrate that 19CAR-T(1a) has lower surface CAR expression, faster internalization, and a higher recycling rate compared to 19CAR-T(8α). Internalized 19CAR(1a) co-localizes more with early and recycling endosomes, and less with lysosomes than 19CAR(8α). These features result in lower activation levels, less cytokine release, and reduced exhaustion markers in 19CAR-T(1a). Furthermore, CAR-T cells with CD1a transmembrane domain also exhibit a superior anti-tumor ability and reduced exhaustion in vivo.ConclusionOverall, we demonstrate that the transmembrane domain plays a critical role in CAR-T cell function. An optimized transmembrane domain can alleviate cytokine release syndrome and reduce CAR-T cell exhaustion, providing a direction for CAR design to enhance CAR-T cell function

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Formation and Evolution of sp2 Hybrid Conjugate Structure of Polyacrylonitrile Precursor during Stabilization

The generated sp2 hybrid conjugate structure of a C atom, which resulted from the chemical reaction affected by temperature and time, is an effective six-membered ring planar structure of the final carbon fiber. This kind of hybrid conjugate structure determined the formation of the final structure and mechanical properties of carbon fiber. In this paper, the formation and evolution of sp2 hybrid conjugated structures of PAN precursor during thermal stabilization were investigated by Raman, UV-vis and 13C-NMR methods. The results indicated that with the increase of stabilization temperature, the degree of the sp2 hybrid conjugated structure of stabilized PAN fiber increases “linearly”, while the content of the sp2 hybrid carbon atoms increases with “S-type”. The final sp2 hybrid conjugated ring structure is mainly composed of single-ring, double-ring, triple-ring, and double-bond structures. Compared with the time factor, the temperature effect plays a decisive role in the formation of the sp2 hybrid conjugate structure.</jats:p

Formation and Evolution of sp2 Hybrid Conjugate Structure of Polyacrylonitrile Precursor during Stabilization

The generated sp2 hybrid conjugate structure of a C atom, which resulted from the chemical reaction affected by temperature and time, is an effective six-membered ring planar structure of the final carbon fiber. This kind of hybrid conjugate structure determined the formation of the final structure and mechanical properties of carbon fiber. In this paper, the formation and evolution of sp2 hybrid conjugated structures of PAN precursor during thermal stabilization were investigated by Raman, UV-vis and 13C-NMR methods. The results indicated that with the increase of stabilization temperature, the degree of the sp2 hybrid conjugated structure of stabilized PAN fiber increases &ldquo;linearly&rdquo;, while the content of the sp2 hybrid carbon atoms increases with &ldquo;S-type&rdquo;. The final sp2 hybrid conjugated ring structure is mainly composed of single-ring, double-ring, triple-ring, and double-bond structures. Compared with the time factor, the temperature effect plays a decisive role in the formation of the sp2 hybrid conjugate structure

Synchrotron study on the evolution of the radial structural distribution of carbon fiber monofilaments during heat treatment process

Evolution of the radial distribution of the crystalline structure within PAN monofilaments during carbonization was investigated via the method of synchrotron diffraction and micro-scanning Raman spectroscopy.</p

Practices and effectiveness of building information modelling in construction projects in China

Based on an investigation of 106 projects involving the use of building information modelling (BIM), this paper examines current BIM practices in China, and assesses how various practices alter their effectiveness. The results reveal that in current practice BIM is principally employed as a visualization tool, and how it is implemented is significantly associated with project characteristics. BIM use in the majority of the surveyed projects is seen to have positive outcomes, with the benefits of improved task effectiveness being more substantial than those related to efficiency improvement. The results also provide evidence that project characteristics significantly influence the success of BIM use; however, more substantial contributing factors to BIM effectiveness are the extent of integrated use and client/owner support. While indicating that current BIM practices involve both technological and organizational problems, the findings also provide insights into how the potential for BIM could be better exploited within the industry

Droj2 Facilitates Somatosensory Neurite Sculpting via GTP-Binding Protein Arf102F in Drosophila

Developmental remodeling of neurite is crucial for the accurate wiring of neural circuits in the developing nervous system in both vertebrates and invertebrates, and may also contribute to the pathogenesis of neuropsychiatric disorders, for instance, autism, Alzheimer&rsquo;s disease (AD), and schizophrenia. However, the molecular underpinnings underlying developmental remodeling are still not fully understood. Here, we have identified DnaJ-like-2 (Droj2), orthologous to human DNAJA1 and DNAJA4 that is predicted to be involved in protein refolding, as a developmental signal promoting dendrite sculpting of the class IV dendritic arborization (C4da) sensory neuron in Drosophila. We further show that Arf102F, a GTP-binding protein previously implicated in protein trafficking, serves downstream of Droj2 to govern neurite pruning of C4da sensory neurons. Intriguingly, our data consistently demonstrate that both Droj2 and Arf102F promote the downregulation of the conserved L1-type cell-adhesion molecule Neuroglian anterior to dendrite pruning. Mechanistically, Droj2 genetically interacts with Arf102F and promotes Neuroglian downregulation to initiate dendrite severing. Taken together, this systematic study sheds light on an unprecedented function of Droj2 and Arf102F in neuronal development

Responses of Issatchenkia terricola WJL-G4 upon Citric Acid Stress

This study aimed to elucidate the responses of a novel characterized Issatchenkia terricola WJL-G4 against citric acid stress by performing physiological analysis, morphology observation, and structural and membrane fatty acid composition analysis. The results showed that under citric acid stress, the cell vitality of I. terricola WJL-G4 was reduced. The cell morphology changed with the unclear, uncompleted and thinner cell wall, and degraded the cell structure. When the citric acid concentration was 20 g/L, I. terricola WJL-G4 could tolerate citric acid and maintain the cell structure by increasing the intracellular pH, superoxide dismutase activity, and contents of unsaturated fatty acids. As the citric acid concentration was &ge;80 g/L, the stress has exceeded the cellular anti-stress ability, causing substantial cell damage. The cell membrane permeability, the content of membrane lipids, malondialdehyde and superoxide anion increased, but the intracellular pH and superoxide dismutase activities decreased, accompanying the increase of citric acid concentrations. The findings of this work provided a theoretical basis for the responsive mechanism of I. terricola WJL-G4 under high concentrations of citric acid, and can serve as a reference for biological acid reduction in fruit processing