Development and characterization of a continuous cell line (EL) from the liver of European eel Anguilla anguilla

In the present study, a new hepatic tissue‐origin cell line from European eel Anguilla anguilla has been developed and characterized. This cell line designated EL has been maintained in Leibovitz L‐15 supplemented with 10% fetal bovine serum over 72 months, and subcultured more than 90 times. The EL cell line consisted predominantly of fibroblast‐like cells, which could survive over 100 days in vitro, and could grow at 15–32°C. The optimum temperature for growth was 27°C. The chromosome analysis revealed a modal diploid karyotype of 2n = 38. The origin of this cell line was confirmed by the 18S recombinant (r)RNA sequencing. The susceptibility test indicated significant cytopathic effects in the EL cells with regard to the Rana grylio virus and the Herpesvirus anguillae. The viral replication was confirmed by transmission electron microscopy and polymerase chain reaction analysis. Following poly (I:C) exposure, the expression levels of the immune‐related molecules interferon regulatory factor‐7 (irf7) and transforming growth factor‐β (TGF‐β) were downregulated in EL cells, whereas the expression levels of the rf3 and the cytochrome P450 (CYP450) were upregulated. All four genes were significantly upregulated following inflammation by lipopolysaccharide (LPS). These data suggested the application of EL cell line for viral identification, as well as for immunodiagnosis and pharmacological targeting.     

Reduced expression of CYLD promotes cell survival and inflammation in gefitinib‐treated NSCLC PC‐9 cells: Targeting CYLD may be beneficial for acquired resistance to gefitinib therapy

The application of tyrosine kinase inhibitors (TKIs) to the epidermal growth factor receptor (EGFR) has been proven to be highly effective for non‐small‐cell lung cancer (NSCLC). However, patients often evolve into acquired resistance. The secondary mutations in EGFR account for nearly half of the acquired resistance. While the remaining 50% of patients exhibit tolerance to EGFR‐TKIs with unclear mechanism(s). Cylindromatosis (CYLD), a deubiquitinase, functions as a tumor suppressor to regulate cell apoptosis, proliferation, and immune response, and so on. The role of CYLD in NSCLC EGFR‐TKI resistance remains elusive. Here, we found CYLD was upregulated in PC‐9 cells, whereas downregulated in PC‐9 acquired gefitinib‐resistant (PC‐9/GR) cells in response to the treatment of gefitinib, which is consistent with the results in the Gene Expression Omnibus database. Overexpression of CYLD promoted a more apoptotic death ratio in PC‐9/GR cells than that in PC‐9 cells. In addition, silencing the expression of CYLD resulted in an increase of the expression level of interleukin‐6, transforming growth factor‐β and tumor necrosis factor‐α, which may contribute to acquired resistance of PC‐9 cells to gefitinib. Taken together, our data in vitro demonstrate that PC‐9/GR cells downregulated CYLD expression, enhanced subsequent CYLD‐dependent antiapoptotic capacity and inflammatory response, which may provide a possible target for acquired gefitinib‐resistant treatment in NSCLC.     

MicroRNA‐93‐5p expression in tumor tissue and its tumor suppressor function via targeting programmed death ligand‐1 in colorectal cancer

This work aimed to investigate miR‐93‐5p expression in tumor tissue and its in vitro effects in colorectal cancer (CRC) by targeting programmed death ligand‐1 (PD‐L1). MiR‐93‐5p and PD‐L1 expression was detected in CRC and adjacent normal tissues by quantitative real‐time polymerase chain reaction and immunohistochemistry. The correlation between miR‐93‐5p and PD‐L1 was validated by a dual‐luciferase reporter assay. HCT116 and SW480 cells were divided into blank, miR‐NC, miR‐93‐5p mimics, miR‐93‐5p inhibitor, PD‐L1 small interfering RNA (siRNA) and miR‐93‐5p inhibitor + PD‐L1 siRNA groups, and wound‐healing and transwell assays were performed to detect cell migration and invasion, respectively. Protein expression was measured by western blotting. The secretion of cytokines was detected in the CRC cell/T coculture models. MiR‐93‐5p was downregulated in CRC tissues with upregulated PD‐L1. In PD‐L1‐negative patients, miR‐93‐5p expression was increased compared with that in PD‐L1‐positive patients. MiR‐93‐5p and PD‐L1 expression levels were associated with the tumor differentiation, lymphatic metastasis, TNM, Duke's stage, and prognosis of CRC. PD‐L1 siRNA weakened the migration and invasion abilities via decreased expression of matrix metalloproteinase‐1 (MMP‐1), ‐2, and ‐9, and these effects were abolished by the miR‐93‐5p inhibitor. Additionally, anti‐PD‐L1 upregulated the expressions of interleukin‐2 (IL‐2), tumor necrosis factor‐α (TNF‐α), and interferon γ (IFN‐γ) in the coculture of T cells with CRC cells, but downregulated the expressions of IL‐1β, IL‐10, and TGF‐β. However, these changes were partially reversed by miR‐93‐5p inhibition. miR‐93‐5p is expected to be a novel target for CRC treatment since it decreases the migration and invasion, as well as the immune evasion, of CRC cells via targeting PD‐L1.     

Factors Affecting the Mixotrophic Flagellate Poterioochromonas malhamensis Grazing on Chlorella Cells

Grazing behaviour between protozoa and phytoplankton exists widely in planktonic ecosystems. Poterioochromonas malhamensis is a well‐known and widespread mixotrophic flagellate, which is recognized to play an important role within marine and freshwater planktonic ecosystems and regarded as the greatest contamination threat for mass algal cultures of Chlorella. In this study, a comprehensive range of factors, including morphological characters, biochemical compositions, and specific growth rate of ten species or strains of Chlorella, were evaluated for their effect on the feeding ability of P. malhamensis, which was assessed by two parameters: the clearance rate of P. malhamensis on Chlorella spp. and the specific growth rate of P. malhamensis. The results showed that the clearance rate of P. malhamensis was negatively correlated with cell wall thickness and specific growth rate of Chlorella spp., while the specific growth rate of P. malhamensis was positively correlated with carbohydrate percentage and C/N ratio and negatively correlated with protein, lipid percentage, and nitrogen mass. In conclusion, the factors influencing feeding selectivity include not only the morphological character and chemical composition of Chlorella, but also its population dynamics. Our study provides useful insights into the key factors that affect the feeding selectivity of P. malhamensis and provides basic and constructive data to help in screening for grazing‐resistant microalgae.     

Population diversity of Cistopus indicus inferred from mitochondrial DNA (Cytochrome b) variation from China and Vietnam

ABSTRACT

The octopus Cistopus indicus is an important target of cephalopod fisheries in China. It is widely distributed in the South Pacific and tropical Indian Ocean, from the South China Sea, the Philippines, Malaysia, to Indian and Pakistan seas. We collected specimens from five sites in China and Vietnam (Zhoushan, Wenzhou, Shacheng, Zhanjiang and Mangjie). A fragment of 675bp of cytochrome b (Cytb) was amplified from 95 individuals. A total of 27 haplotypes and 78 variable nucleotide sites was observed. High haplotype diversity and low nucleotide diversity were observed in all populations. The phylogenetic analysis separated these populations into two clades; one was composed of three populations (Zhoushan, Wenzhou and Shacheng), the other of two (Zhanjiang, Mangjie). AMOVA analysis detected that 4.67% of the genetic variation occurred within populations and 95.33% occurred among populations. FST values ranged from 0.014 to 0.993, highlighting the high genetic variation among the populations. Assuming a molecular clock with a rate of 2.15–2.6%/Ma for the Cytb gene, the two clades may have diverged 2.88–3.49 million years ago (Pliocene). Neutral evolution tests and mismatch distribution analysis suggested recent population expansion. The present results revealed valuable information for genetic assessment, management and conservation of this species.     

Traits mediate drought effects on wood carbon fluxes

CO₂ fluxes from wood decomposition represent an important source of carbon from forest ecosystems to the atmosphere, which are determined by both wood traits and climate influencing the metabolic rates of decomposers. Previous studies have quantified the effects of moisture and temperature on wood decomposition, but these effects were not separated from the potential influence of wood traits. Indeed, it is not well understood how traits and climate interact to influence wood CO₂ fluxes. Here, we examined the responses of CO₂ fluxes from dead wood with different traits (angiosperm and gymnosperm) to 0%, 35%, and 70% rainfall reduction across seasonal temperature gradients. Our results showed that drought significantly decreased wood CO₂ fluxes, but its effects varied with both taxonomical group and drought intensity. Drought‐induced reduction in wood CO₂ fluxes was larger in angiosperms than gymnosperms for the 35% rainfall reduction treatment, but there was no significant difference between these groups for the 70% reduction treatment. This is because wood nitrogen density and carbon quality were significantly higher in angiosperms than gymnosperms, yielding a higher moisture sensitivity of wood decomposition. These findings were demonstrated by a significant positive interaction effect between wood nitrogen and moisture on CO₂ fluxes in a structural equation model. Additionally, we ascertained that a constant temperature sensitivity of CO₂ fluxes was independent of wood traits and consistent with previous estimates for extracellular enzyme kinetics. Our results highlight the key role of wood traits in regulating drought responses of wood carbon fluxes. Given that both climate and forest management might extensively modify taxonomic compositions in the future, it is critical for carbon cycle models to account for such interactions between wood traits and climate in driving dynamics of wood decomposition.     

Reforestation and surface cooling in temperate zones: Mechanisms and implications

Land‐use/cover change (LUCC) is an important driver of environmental change, occurring at the same time as, and often interacting with, global climate change. Reforestation and deforestation have been critical aspects of LUCC over the past two centuries and are widely studied for their potential to perturb the global carbon cycle. More recently, there has been keen interest in understanding the extent to which reforestation affects terrestrial energy cycling and thus surface temperature directly by altering surface physical properties (e.g., albedo and emissivity) and land–atmosphere energy exchange. The impacts of reforestation on land surface temperature and their mechanisms are relatively well understood in tropical and boreal climates, but the effects of reforestation on warming and/or cooling in temperate zones are less certain. This study is designed to elucidate the biophysical mechanisms that link land cover and surface temperature in temperate ecosystems. To achieve this goal, we used data from six paired eddy‐covariance towers over co‐located forests and grasslands in the temperate eastern United States, where radiation components, latent and sensible heat fluxes, and meteorological conditions were measured. The results show that, at the annual time scale, the surface of the forests is 1–2°C cooler than grasslands, indicating a substantial cooling effect of reforestation. The enhanced latent and sensible heat fluxes of forests have an average cooling effect of ?2.5°C, which offsets the net warming effect (+1.5°C) of albedo warming (+2.3°C) and emissivity cooling effect (?0.8°C) associated with surface properties. Additional daytime cooling over forests is driven by local feedbacks to incoming radiation. We further show that the forest cooling effect is most pronounced when land surface temperature is higher, often exceeding ?5°C. Our results contribute important observational evidence that reforestation in the temperate zone offers opportunities for local climate mitigation and adaptation.     

The impacts of warming and nitrogen addition on competitive ability of native and invasive populations of Plantago virginica

全球变化因子(如增温和氮沉降)可能会影响生物入侵,但是这些因子如何影响入侵物种的表现并进一步调节入侵物种与本地竞争者之间的相互作用仍不清楚。本文通过为期五个月的温室实验,研究了增温(开顶式增温箱,+0.62°C)和氮添加(4.2 g N m⁻²)对入侵物种北美 车前(Plantago virginica)原产地和入侵地种群与本地车前草(Plantago asiatica)竞争的影响。实验结果表明,在增温及其与氮添加处理(W × N) 的相互作用下,P. virginica的入侵种群(PV-In)和原产地种群(PV-Na)在与本地竞争者P. asiatica竞争时具有不同的生物量分配策略。其中,PV-Na在与P. asiatica竞争时增加了地下生物量,而PV-In增加了地上生物量。我们还发现,P. virginica对增温和氮添加比P. asiatica的反应更强 烈。增温显著降低了P. virginica的竞争能力,这表明P. virginica比P. asiatica对增温的响应更为敏感。同样,在竞争条件下,氮添加及 其和增温交互作用减少了PV-In地下生物量,但增加了PV-Na地上和总生物量。这些发现表明,P. virginica在入侵过程中改变了生物量分配 策略,PV-In展示出更具弹性的竞争能力以适应环境变化(特别是增温)。这些发现可能有助于我们预测气候变化下的植物入侵并制定相应的 管理策略。     

The asymmetric relationships of the distribution of conspecific saplings and adults in forest fragments

随着土地利用方式变化的加剧,生境片段化已成为影响植物多样性的主要因子之一。通常,当成年树个体的密度越高,其周边同种幼树个体的存活率可能会下降,从而为其它物种提供了空间和资源,进而可以维持较高的局域物种多样性。因此,同种成年树和幼树个体的空间分布格局关系和作用强度可以调节植物多样性。然而,对于在片段化森林中,同种成年树和幼树个体空间分布关系的研究却很少报道,迄今尚不清楚片段化景观中同种个体的空间分布关系与物种多样性之间的联系。本研究选择千岛湖陆桥岛屿系统中的27个岛屿,基于岛屿上幼树和成年树个体的空间分布数据,利用混合效应模型分析它们之间的作用强度。同种幼树和成年树个体的空间作用强度越大,说明它们之间的负相互作用越强,即幼树和成年树个体空间分布越分散。此外,本研究分析了岛屿属性(岛屿面积、与大陆的距离和与最近岛屿的距离)与同种个体空间作用强度及物种多样性之间的关系。结果表明,同种个体的空间作用强度随着与最近岛屿距离的增加而增加。同时,物种多样性随着同种个体的空间作用强度的增加而显著增加,且岛屿面积和同种个体的空间作用强度分别解释了岛屿间物种多样性差异的26%和6%,共同解释了8%。耐阴种和非常见种比非耐阴种和常见种的同种幼树和成年树的空间分布更为分散。本研究表明,同种个体的空间分布可能会影响多度较低物种在片段化森林中的生存,反映了生物相互作用对于维持片段化森林中的植物多样性具有重要作用。 本研究也强调在检验同种密度制约时应考虑森林之间的连接度。     

Do innate killing mechanisms activated by inflammasomes have a role in treating melanoma?

Melanoma, as for many other cancers, undergoes a selection process during progression that limits many innate and adaptive tumor control mechanisms. Immunotherapy with immune checkpoint blockade overcomes one of the escape mechanisms but if the tumor is not eliminated other escape mechanisms evolve that require new approaches for tumor control. Some of the innate mechanisms that have evolved against infections with microorganisms and viruses are proving to be active against cancer cells but require better understanding of how they are activated and what inhibitory mechanisms may need to be targeted. This is particularly so for inflammasomes which have evolved against many different organisms and which recruit a number of cytotoxic mechanisms that remain poorly understood. Equally important is understanding of where these mechanisms will fit into existing treatment strategies and whether existing strategies already involve the innate killing mechanisms.