Ex vivo expanded human regulatory T cells modify neuroinflammation in a preclinical model of Alzheimer's disease.

BACKGROUND: Regulatory T cells (Tregs) play a neuroprotective role by suppressing microglia and macrophage-mediated inflammation and modulating adaptive immune reactions. We previously documented that Treg immunomodulatory mechanisms are compromised in Alzheimer's disease (AD). Ex vivo expansion of Tregs restores and amplifies their immunosuppressive functions in vitro. A key question is whether adoptive transfer of ex vivo expanded human Tregs can suppress neuroinflammation and amyloid pathology in a preclinical mouse model. METHODS: An immunodeficient mouse model of AD was generated by backcrossing the 5xFAD onto Rag2 knockout mice (5xFAD-Rag2KO). Human Tregs were expanded ex vivo for 24 days and administered to 5xFAD-Rag2KO. Changes in amyloid burden, microglia characteristics and reactive astrocytes were evaluated using ELISA and confocal microscopy. NanoString Mouse AD multiplex gene expression analysis was applied to explore the impact of ex vivo expanded Tregs on the neuroinflammation transcriptome. RESULTS: Elimination of mature B and T lymphocytes and natural killer cells in 5xFAD-Rag2KO mice was associated with upregulation of 95 inflammation genes and amplified number of reactive microglia within the dentate gyrus. Administration of ex vivo expanded Tregs reduced amyloid burden and reactive glial cells in the dentate gyrus and frontal cortex of 5xFAD-Rag2KO mice. Interrogation of inflammation gene expression documented down-regulation of pro-inflammatory cytokines (IL1A&B, IL6), complement cascade (C1qa, C1qb, C1qc, C4a/b), toll-like receptors (Tlr3, Tlr4 and Tlr7) and microglial activations markers (CD14, Tyrobp,Trem2) following Treg administration. CONCLUSIONS: Ex vivo expanded Tregs with amplified immunomodulatory function, suppressed neuroinflammation and alleviated AD pathology in vivo. Our results provide preclinical evidences for Treg cell therapy as a potential treatment strategy in AD.

LINC01315 promotes the aggressive phenotypes of papillary thyroid cancer cells by sponging miR-497-5p.

Dysregulation of the long intergenic noncoding RNA 01315 (LINC01315) has recently been demonstrated in cancer. However, the role of LINC01315 in papillary thyroid cancer (PTC) has not been determined. We attempted to determine the function of LINC01315 in PTC. The levels of LINC01315 were higher in thyroid carcinoma tissues and cell lines compared with that in noncancerous tissues or normal cells, respectively. LINC01315 knockdown significantly inhibited the in vitro colony formation and invasion of PTC cells. Upregulation of LINC01315 produced opposite effects. Bioinformatic analysis and luciferase reporter assays indicated direct binding of miR-497-5p to LINC01315. Gain- and loss-of-function assays indicated that miR-497-5p acts as a suppressive miRNA in PTC. Furthermore, LINC01315 facilitated the growth and invasion of PTC cells by sponging miR-497-5p. Our results demonstrated the critical role of the LINC01315-miR-497-5p axis in the growth and invasion of PTC cells.

Differences in TCR repertoire and T cell activation underlie the divergent outcomes of antitumor immune responses in tumor-eradicating versus tumor-progressing hosts.

BACKGROUND: Antitumor immunity is highly heterogeneous between individuals; however, underlying mechanisms remain elusive, despite their potential to improve personalized cancer immunotherapy. Head and neck squamous cell carcinomas (HNSCCs) vary significantly in immune infiltration and therapeutic responses between patients, demanding a mouse model with appropriate heterogeneity to investigate mechanistic differences. METHODS: We developed a unique HNSCC mouse model to investigate underlying mechanisms of heterogeneous antitumor immunity. This model system may provide a better control for tumor-intrinsic and host-genetic variables, thereby uncovering the contribution of the adaptive immunity to tumor eradication. We employed single-cell T-cell receptor (TCR) sequencing coupled with single-cell RNA sequencing to identify the difference in TCR repertoire of CD8 tumor-infiltrating lymphocytes (TILs) and the unique activation states linked with different TCR clonotypes. RESULTS: We discovered that genetically identical wild-type recipient mice responded heterogeneously to the same squamous cell carcinoma tumors orthotopically transplanted into the buccal mucosa. While tumors initially grew in 100% of recipients and most developed aggressive tumors, ~25% of recipients reproducibly eradicated tumors without intervention. Heterogeneous antitumor responses were dependent on CD8 T cells. Consistently, CD8 TILs in regressing tumors were significantly increased and more activated. Single-cell TCR-sequencing revealed that CD8 TILs from both growing and regressing tumors displayed evidence of clonal expansion compared with splenic controls. However, top TCR clonotypes and TCR specificity groups appear to be mutually exclusive between regressing and growing TILs. Furthermore, many TCRα/TCRß sequences only occur in one recipient. By coupling single-cell transcriptomic analysis with unique TCR clonotypes, we found that top TCR clonotypes clustered in distinct activation states in regressing versus growing TILs. Intriguingly, the few TCR clonotypes shared between regressors and progressors differed greatly in their activation states, suggesting a more dominant influence from tumor microenvironment than TCR itself on T cell activation status. CONCLUSIONS: We reveal that intrinsic differences in the TCR repertoire of TILs and their different transcriptional trajectories may underlie the heterogeneous antitumor immune responses in different hosts. We suggest that antitumor immune responses are highly individualized and different hosts employ different TCR specificities against the same tumors, which may have important implications for developing personalized cancer immunotherapy.

[Phenological responses of apple tree to climate warming in the main apple production areas in northern China]. / 中国北方苹果主产地苹果物候期对气候变暖的响应.

To reveal the spatio-temporal variation characteristics of apple's phenology and their critical response time period and intensity to the temperature change in the main production areas of northern China, we chose Fushan, Wanrong and Akesu to respresent the Bohai Gulf, the Loess Plateau and Xinjiang apple production areas, respectively. Apple's phenology data of buds opening (BO), first leaf unfolding (LU), first flowering (FF), fruit maturing (FM), end of leaf coloring (LC) and the end of leaf fall (LF) at the three stations during 1996-2018 were used to analyze the changes of phenological occurrence dates and different growth stage lengths. Partial least squares (PLS) regression was applied to identify the impacts of climate warming on different phenology events at daily resolution. Results showed that regional mean occurrence dates of apple's BO, LU and FF advanced by a rate of 0.36, 0.33 and 0.23 day per year, respectively. However, apple's LF postponed by 0.68 d·a-1. The FM and LC showed different trends among all the sites. The length of fruit growing period (FG) and that of tree growing period (TG) extended at average rates of 1.20 and 0.82 day per year. Apple's spring phenophases dates at all stations correlated negatively with mean temperature during early January to pre-phenophases date, with a 1 ℃ increase inducing an advancement of 3.70, 3.47 and 3.48 days for apple's BO, LU and FF, respectively. In contrast, apple's autumn phenophases correlated positively with mean temperature 21-72 days before the phenophases date, and its correlation with mean temperature was lower than the correlation for spring phenophases. Generally, the effect of temperature on spring phenophase was stronger than that of autumn phenophase, and the extension of FG and TG was mainly caused by the advance of spring phenophase. The responses of apple's phenophases to climate warming differed across all the stations. Temperature had the greatest impact on the development of apple industry in Akesu, less in Wanrong, and with the least influence in Fushan. Our results could provide theoretical basis for response to climate change for apple industry in different areas of China.

Site-specific, covalent immobilization of an engineered enterokinase onto magnetic nanoparticles through transglutaminase-catalyzed bioconjugation.

Enterokinase (EK) is one of the most popular enzymes for the in vitro cleavage of fusion proteins due to its high degree of specificity for the amino-acid sequence (Asp)4-Lys. Enzyme reusability is desirable for reducing operating costs and facilitating the industrial application of EK. In this work, we report the controlled, site-specific and covalent cross-linking of an engineered EKLC on amine-modified magnetic nanoparticles (NH2-MNPs) via microbial transglutaminase-catalyzed bioconjugation for the development of the oriented-immobilized enzyme, namely, EKLC@NH2-MNP biocatalyst. Upon the site-specific immobilization, approximately 90% EKLC enzymatic activity was retained, and the biocatalyst exhibited more than 85% of initial enzymatic activity regardless of storage or reusable stability over a month. The EKLC@NH2-MNP biocatalyst was further applied to remove the His tag-(Asp)4-Lys fusion partner from the His tag-(Asp)4-Lys-(GLP-1)3 substrate fusion protein, result suggested the EKLC@NH2-MNP possessed remarkable reusability, without a significant decrease of enzymatic activity over 10 cycles (P > 0.05). Supported by the unique properties of MNPs, the proposed EKLC@NH2-MNP biocatalyst is expected to promote the economical utilization of enterokinase in fusion protein cleavage.

[Vertical temperature distribution and its forecast for two tree structures of apple orchard during the blooming period in the Loess Plateau].

Temperature is the most sensitive environment factor for the blooming period of apple. Temperatures at different levels were measured by automatic micro-climatic gradient system in the blooming periods from 2011 to 2014, in two Fuji apple orchards with two different tree ages and structures [small canopy open center shape (SMCOCS) and freedom spindle shape (FSS)], respectively, which were typical in the Loess Plateau. Variations of the temperature gradient in both canopy and tree body were analyzed in sunny, overcast, cloudy, and rainy weather conditions, and a predicting model was established that could predict the temperature of the canopy (TL) according to the temperature observed in nearby meteorological station (TM). The results showed that the vertical distribution of canopy temperature and its difference to the outside of orchard was mainly due to the tree structure, rather than the weather condition. The average temperature and daily minimum temperature increased while the daily maximum temperature and the diurnal temperature range decreased from the bottom to the upper of the canopy. For SMCOCS, the diurnal temperature range reached its peak under the canopy in the clear days, and the diurnal temperature range was less than that for FSS in the middle and upper canopy in cloudy or overcast conditions. The daily variation of temperature difference between inside and outside the orchard behaved as a single peak-valley-peak for FSS but as a single peak for SMCOCS. The minimum temperature outside the orchard was closer to that in the middle of canopy, but higher than that in the bottom of the canopy. For SMCOCS, the minimum temperature in the bottom of its canopy was rather lower than that in the orchard outside, especially in cloudy or overcast day, while in the middle or upper canopy, the minimum temperature difference with the orchard outside was smaller than that for the FSS. The linear model was found to be able to predict the TL with absolute errors below 1 °C, and the best prediction was found for the FSS in rainy days.

Reversibly thermoswitchable two-dimensional periodic gratings prepared from tethered poly(N-isopropylacrylamide) on silicon surfaces.

In this study we used atom transfer radical polymerization to graft poly(N-isopropylacrylamide) (PNIPAAm) onto flat Si substrates. We then applied very large-scale integration and reactive ion etching sequentially to generate 200-nm-scale hole arrays of tethered PNIPAAm as two-dimensional periodic concave gratings (2DPCGs) on the Si surfaces. The hole array structures of tethered PNIPAAm could be created and erased reversibly at 25 and 40 °C, respectively, leading to significant changes in the effective refractive index (neff). The values of neff of the 2DPCGs were related to the depth of their holes generated after etching for various times, resulting in a color change from blue to red that could be observed by the naked eye at incident angles of 10-20°. Moreover, we used effective-medium theory to calculate the filling factors of air inside the 2DPCGs to verify the structural changes of the tethered PNIPAAm. Such designed 2DPCGs of thermorespective hydrogel films have potential applications in temperature-responsive optical devices [e.g., as antireflection structured surfaces (ARSs)] that operate at both visible and near-infrared wavelengths.