The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

Design and Construction of pH-Selective Self-Lytic Liposome System

Liposomes are well-investigated drug or gene delivery vehicles for chemotherapy, used by taking advantage of their biocompatibility and biodegradability. A central question on the construction of intracellular liposomal delivery systems is to entrap the liposomes of interest in the highly acidic and proteolytic endosomal environment. In the other words, it is essential that the liposomes release a therapeutic drug into the cytosol before they are degraded in the endosome. As a strategy to enhance the endosome escape, the self-lytic liposomes with acidic pH-selective membrane active polypeptide are considered highly effective. Here, an acidic pH-selective membrane-lytic polypeptide (LPE) and its retro isomer (rLPE) were designed, and then their membrane-lytic activities against EggPC liposomes were determined. It was noticed that the rLPE polypeptide showed an increase in activity compared with the LPE polypeptide. Furthermore, the rLPE polypeptide was conjugated to liposomes via a flexible Gly-Gly-Gly-Gly linker to facilitate the pH-selective content release. The rLPE anchoring liposomes exhibited distinctly different contents release behavior at physiological and endosomal pHs, namely typical contents release from liposomes was positively observed at acidic pH range. The overarching goal of this paper is to develop efficient pH-selective therapeutic delivery systems by using our findings

Extending Whole Slide Imaging: Color Darkfield Internal Reflection Illumination (DIRI) for Biological Applications.

Whole slide imaging (WSI) is a useful tool for multi-modal imaging, and in our work, we have often combined WSI with darkfield microscopy. However, traditional darkfield microscopy cannot use a single condenser to support high- and low-numerical-aperture objectives, which limits the modality of WSI. To overcome this limitation, we previously developed a darkfield internal reflection illumination (DIRI) microscope using white light-emitting diodes (LEDs). Although the developed DIRI is useful for biological applications, substantial problems remain to be resolved. In this study, we propose a novel illumination technique called color DIRI. The use of three-color LEDs dramatically improves the capability of the system, such that color DIRI (1) enables optimization of the illumination color; (2) can be combined with an oil objective lens; (3) can produce fluorescence excitation illumination; (4) can adjust the wavelength of light to avoid cell damage or reactions; and (5) can be used as a photostimulator. These results clearly illustrate that the proposed color DIRI can significantly extend WSI modalities for biological applications

Comparison of Hausmann’s darkfield, white-LED DIRI, and three-color LED DIRI.

<p>Comparison of Hausmann’s darkfield, white-LED DIRI, and three-color LED DIRI.</p

Schematic of the whole slide imaging (WSI) system with darkfield internal reflection illumination (DIRI).

<p>DIRI was incorporated into the WSI system’s motorized stage. Three color light-emitting diodes (LEDs) illuminate the slide glass from the side, and the specimen scatters this light. The scattered light is then incident on the objective lens above the stage. The dichromatic mirror on the motorized turret of the microscope can be removed from the light path when acquiring darkfield images. A tube lens above the dichromatic mirror focuses the sample image onto the imaging device. A charge-coupled device camera then captures the image. A sharp cut-off filter is placed between the field diaphragm and the mirror.</p

DIRI with three-color-LED array.

<p>(a) The three-color-LED array and controller, (b) a sample illuminated with green-LED array, (c) a schematic diagram of the three-color LEDs and a slide glass sample, and (d) a sample illuminated with red (left), green (middle), and blue (right) LEDs in the <i>E gracilis</i> experiment.</p