Nuclease-resistant immunostimulatory phosphodiester CpG oligodeoxynucleotides as human Toll-like receptor 9 agonists

<p>Abstract</p> <p>Background</p> <p>Unmethylated cytosine-guanine (CpG) motif-containing oligodeoxynucleotides (ODNs) have been well characterized as agonists of Toll-like receptor 9 (TLR9). ODNs with a phosphorothioate (PTO) backbone have been studied as TLR9 agonists since natural ODNs with a phosphodiester (PD) backbone are easily degraded by a serum nuclease, which makes them problematic for therapeutic applications. However, ODNs with a PTO backbone have been shown to have undesirable side effects. Thus, our goal was to develop nuclease-resistant, PD ODNs that are effective as human TLR9 (hTLR9) agonists.</p> <p>Results</p> <p>The sequence of ODN2006, a CpG ODN that acts as an hTLR9 agonist, was used as the basic CpG ODN material. The 3'-end modification of ODN2006 with a PD backbone (PD-ODN2006) improved its potential as an hTLR9 agonist because of increased resistance to nucleolytic degradation. Moreover, 3'-end modification with oligonucleotides showed higher induction than modification with biotin, FITC, and amino groups. Further, enhancement of hTLR9 activity was found to be dependent on the number of CpG core motifs (GTCGTT) in the PD ODN containing the 3'-end oligonucleotides. In particular, ODN sequences consisting of two to three linked ODN2006 sequences with a PD backbone (e.g., PD-ODN2006-2006 and PD-ODN2006-2006-2006) acted as effective agonists of hTLR9 even at lower concentrations.</p> <p>Conclusions</p> <p>This study showed that PD-ODN2006-2006 and PD-ODN-2006-2006-2006 can be used as potentially safe agonists for hTLR9 activation instead of CpG ODNs with a PTO backbone. We propose these CpG ODNs consisting of only a PD backbone as a novel class of CpG ODN.</p

Effects of substitutions of glycine and asparagine for serine132 on activity and binding of human lipoprotein lipase to very low density lipoproteins

AbstractFor studying the role of Ser132 in the putative catalytic site of human lipoprotein lipase (LPL), mutant LPL cDNAs expressing LPLs with amino acid substitutions of Gly or Asn for Ser132 were obtained by site-directed mutagenesis, and were expressed in COS-1 cells. Considerable amounts of LPL enzyme protein mass were detected in the culture medium of COS-1 cells transfected with wild-type LPL, LPL-Gly132, or LPL-Asn132. LPL-Gly132 hydrolyzed Triton X-100-triolein and tributyrin as effectively as wild-type LPL, whereas LPL-Asn132 showed no activity. LPL-Asn132 bound to very low density lipoproteins as effectively as wild-type LPL

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

Removal of estrogenic activity of endocrine-disrupting genistein by ligninolytic enzymes from white rot fungi

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A Multifunctional Hybrid Nanocarrier for Non-Invasive siRNA Delivery to the Retina

Drug therapy for retinal diseases (e.g., age-related macular degeneration, the leading cause of blindness) is generally performed by invasive intravitreal injection because of poor drug delivery caused by the blood–retinal barrier (BRB). This study aimed to develop a nanocarrier for the non-invasive delivery of small interfering RNA (siRNA) to the posterior segment of the eye (i.e., the retina) by eyedrops. To this end, we prepared a hybrid nanocarrier based on a multifunctional peptide and liposomes, and the composition was optimized. A cytoplasm-responsive stearylated peptide (STR-CH2R4H2C) was used as the multifunctional peptide because of its superior ability to enhance the complexation, cell permeation, and intracellular dynamics of siRNA. By adding STR-CH2R4H2C to the surface of liposomes, intracellular uptake increased regardless of the liposome surface charge. The STR-CH2R4H2C-modified cationic nanocarrier demonstrated significant siRNA transfection efficiency with no cytotoxicity, enhanced siRNA release from endosomes, and effectively suppressed vascular endothelial growth factor expression in rat retinal pigment epithelium cells. The 2.0 mol% STR-CH2R4H2C-modified cationic nanocarrier enhanced intraocular migration into the retina after instillation into rat eyes