Intratumoral expression of IL-17 and its prognostic role in gastric adenocarcinoma patients

In this study, we characterized the intratumoral expression of IL-17 and CD8(+) TILs in gastric adenocarcinoma patients after resection and determined the correlation between the survival probability of gastric adenocarcinoma patients and the expression of IL-17 in tumor. Expression of IL-17 and CD8 was assessed by immunohistochemistry, and the prognostic effects of intratumoral IL-17 expression and CD8(+) TILs were evaluated by Cox regression and Kaplan-Meier analysis. Immunohistochemical detection revealed the presence of IL-17 and CD8(+) cells in gastric adenocarcinoma tissue samples (90.6%, 174 out of 192 patients and 96.9%, 186 out of 192 patients, respectively). We have also found that intratumoral IL-17 expression was significantly correlated with age (p=0.004) and that the number of CD8(+)TILs was significantly correlated with UICC staging (p=0.012) and the depth of tumor invasion (p=0.022). The five-year overall survival probability among patients intratumorally expressing higher levels of IL-17 was significantly better than those expressing lower levels of IL-17 (p=0.036). Multivariate Cox proportional hazard analyses revealed that intratumoral IL-17 expression (HR: 0.521; 95% CI: 0.329-0.823; p=0.005) was an independent factor affecting the five-year overall survival probability. We conclude that low levels of intratumoral IL-17 expression may indicate poor prognosis in gastric adenocarcinoma patients.     

Soil Respiration and Organic Carbon Dynamics with Grassland Conversions to Woodlands in Temperate China

Soils are the largest terrestrial carbon store and soil respiration is the second-largest flux in ecosystem carbon cycling. Across China''s temperate region, climatic changes and human activities have frequently caused the transformation of grasslands to woodlands. However, the effect of this transition on soil respiration and soil organic carbon (SOC) dynamics remains uncertain in this area. In this study, we measured in situ soil respiration and SOC storage over a two-year period (Jan. 2007–Dec. 2008) from five characteristic vegetation types in a forest-steppe ecotone of temperate China, including grassland (GR), shrubland (SH), as well as in evergreen coniferous (EC), deciduous coniferous (DC) and deciduous broadleaved forest (DB), to evaluate the changes of soil respiration and SOC storage with grassland conversions to diverse types of woodlands. Annual soil respiration increased by 3%, 6%, 14%, and 22% after the conversion from GR to EC, SH, DC, and DB, respectively. The variation in soil respiration among different vegetation types could be well explained by SOC and soil total nitrogen content. Despite higher soil respiration in woodlands, SOC storage and residence time increased in the upper 20 cm of soil. Our results suggest that the differences in soil environmental conditions, especially soil substrate availability, influenced the level of annual soil respiration produced by different vegetation types. Moreover, shifts from grassland to woody plant dominance resulted in increased SOC storage. Given the widespread increase in woody plant abundance caused by climate change and large-scale afforestation programs, the soils are expected to accumulate and store increased amounts of organic carbon in temperate areas of China.     

Symbiosis-promoting and deleterious effects of NopT, a novel type 3 effector of Rhizobium sp. strain NGR234

Establishment of symbiosis between certain host plants and nitrogen-fixing bacteria ("rhizobia") depends on type 3 effector proteins secreted via the bacterial type 3 secretion system (T3SS). Here, we report that the open reading frame y4zC of strain NGR234 encodes a novel rhizobial type 3 effector, termed NopT (for nodulation outer protein T). Analysis of secreted proteins from NGR234 and T3SS mutants revealed that NopT is secreted via the T3SS. NopT possessed autoproteolytic activity when expressed in Escherichia coli or human HEK 293T cells. The processed NopT exposed a glycine (G50) to the N terminus, which is predicted to be myristoylated in eukaryotic cells. NopT with a point mutation at position C93, H205, or D220 (catalytic triad) showed strongly reduced autoproteolytic activity, indicating that NopT is a functional protease of the YopT-AvrPphB effector family. When transiently expressed in tobacco plants, proteolytically active NopT elicited a rapid hypersensitive reaction. Arabidopsis plants transformed with nopT showed chlorotic and necrotic symptoms, indicating a cytotoxic effect. Inoculation experiments with mutant derivatives of NGR234 indicated that NopT affected nodulation either positively (Phaseolus vulgaris cv. Yudou No. 1; Tephrosia vogelii) or negatively (Crotalaria juncea). We suggest that NopT-related polymorphism may be involved in evolutionary adaptation of NGR234 to particular host legumes.     

Isolation of Chinese hamster ovary cell lines producing Man3GlcNAc2 asparagine-linked glycans.

Chinese hamster ovary lines with two mutations, one causing accumulation of Man5GlcNAc2-P-P-dolichol and a second resulting in defective N-acetylglucosaminyltransferase I activity, synthesize asparagine-linked glycans with the structure Man3GlcNAc2. As a result, the asparagine-linked glycans produced by these lines are smaller and less heterogeneous than those produced by other currently available animal cell lines.     

Assessment of hepatoprotective potential of Radix Fici Hirtae on alcohol-induced liver injury in Kunming mice

ObjectiveThe objective of the present study was to investigate the hepatoprotective role of Radix Fici Hirtae on acute alcohol-induced liver injury in mice.MethodsThe component of Radix Fici Hirtae was extracted using petroleum ether, chloroform, ethyl acetate and n-butanol and divided into three dose groups of high, medium and low according to the clinical man's normal dose of the 50 g crude drug/d (0.83 g/kg body weight). Saline in concentration of 10 mg/mL, 5 mg/mL and 2.5 mg/mL and a dose of mouse lavage (0.2 mL/10 g mouse body weight) were added to the solution. Histopathlogical analysis of liver was performed. Finally, liver protection was validated by examining the effect of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AKP), and lactate dehydrogenase (LDH) on the hepatic function of mice in alcohol-induced liver injury model.ResultsExcept for group with saturated n-butyl alcohol, for the rest of the groups, pathological changes of hepatic lipid and inflammatory cells infiltration were alleviated and liver sinus was normal. As compared to model group, the concentrations of AST, ALT, AKP and LDH in chloroform groups and ethyl acetate groups were significantly decreased.ConclusionsExtracts of Radix Fici Hirtae are effective for the prevention of alcohol-induced hepatic damage in mice. The results revealed that extracts of Radix Fici Hirtae could be used as hepatoprotective agent.     

Single‐Ion Conducting Electrolyte Based on Electrospun Nanofibers for High‐Performance Lithium Batteries

Herein, a novel electrospun single‐ion conducting polymer electrolyte (SIPE) composed of nanoscale mixed poly(vinylidene fluoride‐co‐hexafluoropropylene) (PVDF‐HFP) and lithium poly(4,4′‐diaminodiphenylsulfone, bis(4‐carbonyl benzene sulfonyl)imide) (LiPSI) is reported, which simultaneously overcomes the drawbacks of the polyolefin‐based separator (low porosity and poor electrolyte wettability and thermal dimensional stability) and the LiPF₆ salt (poor thermal stability and moisture sensitivity). The electrospun nanofiber membrane (es‐PVPSI) has high porosity and appropriate mechanical strength. The fully aromatic polyamide backbone enables high thermal dimensional stability of es‐PVPSI membrane even at 300 °C, while the high polarity and high porosity ensures fast electrolyte wetting. Impregnation of the membrane with the ethylene carbonate (EC)/dimethyl carbonate (DMC) (v:v = 1:1) solvent mixture yields a SIPE offering wide electrochemical stability, good ionic conductivity, and high lithium‐ion transference number. Based on the above‐mentioned merits, Li/LiFePO₄ cells using such a SIPE exhibit excellent rate capacity and outstanding electrochemical stability for 1000 cycles at least, indicating that such an electrolyte can replace the conventional liquid electrolyte–polyolefin combination in lithium ion batteries (LIBs). In addition, the long‐term stripping–plating cycling test coupled with scanning electron microscope (SEM) images of lithium foil clearly confirms that the es‐PVPSI membrane is capable of suppressing lithium dendrite growth, which is fundamental for its use in high‐energy Li metal batteries.     

Large-scale expansion of Vγ9Vδ2 T cells with engineered K562 feeder cells in G-Rex vessels and their use as chimeric antigen receptor–modified effector cells

Vγ9Vδ2 T cells are a minor subset of lymphocytes in the peripheral blood that has been extensively investigated for their tolerability, safety and anticancer efficacy. A hindrance to the broad application of these cells for adoptive cellular immunotherapy has been attaining clinically appropriate numbers of Vγ9Vδ2 T cells. Furthermore, Vγ9Vδ2 T cells exist at low frequencies among cancer patients. We, therefore, sought to conceive an economical method that allows for a quick and robust large-scale expansion of Vγ9Vδ2 T cells. A two-step protocol was developed, in which peripheral blood mononuclear cells (PBMCs) from healthy donors or cancer patients were activated with Zometa and interleukin (IL)-2, followed by co-culturing with gamma-irradiated, CD64-, CD86- and CD137L-expressing K562 artificial antigen-presenting cells (aAPCs) in the presence of the anti-CD3 antibody OKT3. We optimized the co-culture ratio of K562 aAPCs to immune cells, and migrated this method to a G-Rex cell growth platform to derive clinically relevant cell numbers in a Good Manufacturing Practice (GMP)-compliant manner. We further include a depletion step to selectively remove αβ T lymphocytes. The method exhibited high expansion folds and a specific enrichment of Vγ9Vδ2 T cells. Expanded Vγ9Vδ2 T cells displayed an effector memory phenotype with a concomitant down-regulated expression of inhibitory immune checkpoint receptors. Finally, we ascertained the cytotoxic activity of these expanded cells by using nonmodified and chimeric antigen receptor (CAR)–engrafted Vγ9Vδ2 T cells against a panel of solid tumor cells. Overall, we report an efficient approach to generate highly functional Vγ9Vδ2 T cells in massive numbers suitable for clinical application in an allogeneic setting.     

Cigarette smoke extract alters genome‐wide profiles of circular RNAs and mRNAs in primary human small airway epithelial cells

As a novel kind of non‐coding RNA, circular RNAs (circRNAs) were involved in various biological processes. However, the role of circRNAs in the developmental process of chronic obstructive pulmonary disease (COPD) is still unclear. In the present study, by using a cell model of COPD in primary human small airway epithelial cells (HSAECs) treated with or without cigarette smoke extract (CSE), we uncovered 4,379 previously unknown circRNAs in human cells and 903 smoke‐specific circRNAs, with the help of RNA‐sequencing and bioinformatic analysis. Moreover, 3,872 up‐ and 4,425 down‐regulated mRNAs were also identified under CSE stimulation. Furthermore, a putative circRNA‐microRNA‐mRNA network was constructed for in‐depth mechanism exploration, which indicated that differentially expressed circRNAs could influence expression of some key genes that participate in response to pentose phosphate pathway, ATP‐binding cassette (ABC) transporters, glycosaminoglycan biosynthesis pathway and cancer‐related pathways. Our research indicated that cigarette smoke had an influence on the biogenesis of circRNAs and mRNAs. CircRNAs might be involved in the response to CSE in COPD through the circRNA‐mediated ceRNA networks.     

Estimation of protein and metabolite release rates from damaged mammalian cells by using GFP as a marker molecule

The quantification of metabolite leakage from damaged mammalian cells to the surrounding medium is of high interest for the processing of samples for metabolomic analysis. It is also of relevance to know the typical time span which is required for a promoted metabolite release through a selectively permeabilized cell membrane. The real-time observation of such a process is difficult since small metabolites cannot be observed directly by optical methods and other more indirect assays can disturb the metabolite concentration itself. However, the diffusion based loss of metabolites from the cytoplasm can be predicted on the basis of reference measurements taken from an easy-to-detect molecule with known diffusion coefficient. In this work, we use green fluorescent protein (GFP) as a marker and model its release from damaged cells using the finite-element method. A correlation between the disrupted membrane area fraction, A _d , the distribution of membrane ruptures and the rate of GFP efflux, k _e , has been established. k _e has been determined experimentally for Chinese hamster ovary cells, which have been damaged mechanically by passage through a micronozzle geometry in a microfluidic system. The immediate GFP release downstream of the micronozzles has been observed in real-time and the corresponding membrane damage has been predicted. On this basis, we calculated the expected times required for the drainage of freely diffusable cytosolic glucose and found a loss of ??90% within 1 s for a disrupted membrane area fraction of ??5%. Hence, even minimal membrane damage would lead to a rapid loss of cytosolic metabolites by diffusion unless membrane resealing processes take place.