Allele-Selective Suppression of Mutant Huntingtin in Primary Human Blood Cells

Post-transcriptional gene silencing is a promising therapy for the monogenic, autosomal dominant, Huntington\u27s disease (HD). However, wild-type huntingtin (HTT) has important cellular functions, so the ideal strategy would selectively lower mutant HTT while sparing wild-type. HD patients were genotyped for heterozygosity at three SNP sites, before phasing each SNP allele to wild-type or mutant HTT. Primary ex vivo myeloid cells were isolated from heterozygous patients and transfected with SNP-targeted siRNA, using glucan particles taken up by phagocytosis. Highly selective mRNA knockdown was achieved when targeting each allele of rs362331 in exon 50 of the HTT transcript; this selectivity was also present on protein studies. However, similar selectivity was not observed when targeting rs362273 or rs362307. Furthermore, HD myeloid cells are hyper-reactive compared to control. Allele-selective suppression of either wild-type or mutant HTT produced a significant, equivalent reduction in the cytokine response of HD myeloid cells to LPS, suggesting that wild-type HTT has a novel immune function. We demonstrate a sequential therapeutic process comprising genotyping and mutant HTT-linkage of SNPs, followed by personalised allele-selective suppression in a small patient cohort. We further show that allele-selectivity in ex vivo patient cells is highly SNP-dependent, with implications for clinical trial target selection

Study on Undertaking-starting of New Generation Migrant Workers

New generation migrant workers have become mainstay of China’s migrant workers and also major builders of China’s urbanization process. Compared with last generation migrant workers, new generation migrant workers have greater awareness of starting an undertaking. In the new trend, undertaking-starting is inevitable for new generation migrant workers. This study analyzed problems encountered by new generation migrant workers in the course of starting an undertaking. It reached conclusions that competent authorities should set up support mechanism in undertaking-starting training, undertaking-starting fund, service platform, and preferential policies, to encourage and support new generation migrant workers to start an undertaking

Unexpected Phylogenetic Position of Parapelophryne among Southeast Asian Bufonids as Revealed by Mitochondrial DNA Sequence (Amphibia, Anura, Bufonidae)

We estimated the phylogenetic relationships of an enigmatic small toad Parapelophryne scalpta from Hainan Island, China to nine other bufonid genera from Southeast and East Asia using ca. 2000 bp sequences of the mitochondrial DNA genes 12S rRNA, tRNAval, and 16S rRNA using maximum likelihood and Bayesian inference methods. The East and Southeast Asian bufonid genera formed a clade in which seven lineages with unresolved relationships to each other were recognized. Monophyly was supported only for (A) Parapelophryne and Bufo, (B) Phrynoidis and Pedostibes, and (C) Leptophryne and Ansonia. All genera were genetically divergent from each other and Parapelophryne, erected purely based on morphology, could be recognized as a distinct genus. On the other hand, it was found to be the sister genus of East Asian Bufo, an unexpected result given their great morphological difference and discontinuous distribution

Does the Mutant CAG Expansion in Huntingtin mRNA Interfere with Exonucleolytic Cleavage of its First Exon

BACKGROUND: Silencing mutant huntingtin mRNA by RNA interference (RNAi) is a therapeutic strategy for Huntington\u27s disease. RNAi induces specific endonucleolytic cleavage of the target HTT mRNA, followed by exonucleolytic processing of the cleaved mRNA fragments. OBJECTIVES: We investigated the clearance of huntingtin mRNA cleavage products following RNAi, to find if particular huntingtin mRNA sequences persist. We especially wanted to find out if the expanded CAG increased production of a toxic mRNA species by impeding degradation of human mutant huntingtin exon 1 mRNA. METHODS: Mice expressing the human mutant HTT transgene with 128 CAG repeats (YAC128 mice) were injected in the striatum with self-complementary AAV9 vectors carrying a miRNA targeting exon 48 of huntingtin mRNA (scAAV-U6-miRNA-HTT-GFP). Transgenic huntingtin mRNA levels were measured in striatal lysates after two weeks. For qPCR, we used species specific primer-probe combinations that together spanned 6 positions along the open reading frame and untranslated regions of the human huntingtin mRNA. Knockdown was also measured in the liver following tail vein injection. RESULTS: Two weeks after intrastriatal administration of scAAV9-U6-miRNA-HTT-GFP, we measured transgenic mutant huntingtin in striatum using probes targeting six different sites along the huntingtin mRNA. Real time PCR showed a reduction of 29% to 36% in human HTT. There was no significant difference in knockdown measured at any of the six sites, including exon 1. In liver, we observed a more pronounced HTT mRNA knockdown of 70% to 76% relative to the untreated mice, and there were also no significant differences among sites. CONCLUSIONS: Our results demonstrate that degradation is equally distributed across the human mutant huntingtin mRNA following RNAi-induced cleavage

Birds of the Ailao Mountains, Yunnan province, China

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Understanding the impact of wood type and moisture on the bonding strength of glued wood

The bonding strength of glued wood is strongly related to wood type and environmental conditions, and this relationship is a dynamic variable. It is thus important to investigate the effect of wood type and relative humidity (RH) on the bonding strength using dynamic tests. Two-layered plywood was lab produced, and two sides of the bonding interphase were latewood and earlywood. After conditioning the samples at 60%, 80%, and 95% RH, the bonding strength was measured with a universal mechanical test machine, and strain distribution was recorded simultaneously with digital image correlation (DIC). The results show that bonding strength and deformation increased with an increase in RH. The moisture content (MC) in the latewood was more than that in the earlywood. This decreased the difference in stiffness between latewood and earlywood and led to a more homogeneous strain distribution. This can protect earlywood from failure caused by larger local structural ruptures. Large strain in latewood contributed to high bonding strength. For all samples, failure occurred in the glue line or on the side with earlywood only. For samples failing in the glue line, structural ruptures occurred in the bonding interphase and the wood next to the bonding interphase

Rapid microwave activation of waste palm into hierarchical porous carbons for supercapacitors using biochars from different carbonization temperatures as catalysts

A rapid, simple and cost-effective approach to prepare hierarchical porous carbons (PCs) for supercapacitors is reported by microwave activation of abundant and low-cost waste palm, biochar (BC) and KOH. BCs from waste palm at different carbonization temperatures (300-700 °C), as catalysts and microwave receptors, were used here for the first time to facilitate the conversion of waste palm into hierarchical PCs. As a result, the high-graphitization PC obtained at a BC carbonization temperature of 300 °C (PC-300) possessed a high surface area (1755 m g), a high pore volume (0.942 cm g) and a moderate mesoporosity (37.79%). Besides their high-graphitization and hierarchical porous structure, the oxygen doping in PC-300 can also promote the rapid transport of electrolyte ions. The symmetric supercapacitor based on the PC-300 even in PVA/LiCl gel electrolyte exhibited a high specific capacitance of 164.8 F g at a current density of 0.5 A g and retained a specific capacitance of 121.3 F g at 10 A g, demonstrating a superior rate capacity of 73.6%. Additionally, the PC-300 supercapacitor delivered a high energy density of 14.6 W h kg at a power density of 398.9 W kg and maintained an energy density of 10.8 W h kg at a high power density of 8016.5 W kg, as well as an excellent cycling stability after 2000 cycles with a capacitance retention of 92.06%

Linking SNPs to CAG repeat length in Huntington\u27s disease patients

Allele-specific silencing using small interfering RNAs targeting heterozygous single-nucleotide polymorphisms (SNPs) is a promising therapy for human trinucleotide repeat diseases such as Huntington\u27s disease. Linking SNP identities to the two HTT alleles, normal and disease-causing, is a prerequisite for allele-specific RNA interference. Here we describe a method, SNP linkage by circularization (SLiC), to identify linkage between CAG repeat length and nucleotide identity of heterozygous SNPs using Huntington\u27s disease patient peripheral blood samples

Prophylactic nicotinamide mononucleotide (NMN) mitigates CSDS-induced depressive-like behaviors in mice via preserving of ATP level in the mPFC

Depression is a prevalent psychiatric disorder with accumulating evidence implicating dysregulation of extracellular adenosine triphosphate (ATP) levels in the medial prefrontal cortex (mPFC). It remains unclear whether facilitating endogenous ATP production and subsequently increasing extracellular ATP level in the mPFC can exert a prophylactic effect against chronic social defeat stress (CSDS)-induced depressive-like behaviors and enhance stress resilience. Here, we found that nicotinamide mononucleotide (NMN) treatment effectively elevated nicotinamide adenine dinucleotide (NAD+) biosynthesis and extracellular ATP levels in the mPFC. Moreover, both the 2-week intraperitoneal (i.p.) injection and 3-week oral gavage of NMN prior to exposure to CSDS effectively prevented the development of depressive-like behavior in mice. These protective effects were accompanied with the preservation of both NAD+ biosynthesis and extracellular ATP level in the mPFC. Furthermore, catalyzing ATP hydrolysis by mPFC injection of the ATPase apyrase negated the prophylactic effects of NMN on CSDS-induced depressive-like behaviors. Prophylactic NMN treatment also prevented the reduction in GABAergic inhibition and the increase in excitability in mPFC neurons projecting to the lateral habenula (LHb). Collectively, these findings demonstrate that the prophylactic effects of NMN on depressive-like behaviors are mediated by preventing extracellular ATP loss in the mPFC, which highlights the potential of NMN supplementation as a novel approach for protecting and preventing stress-induced depression in susceptible individuals