-Cystic echinococcosis (CE), caused by infection with the larval stage of the cestode Echinococcus granulosus, is a chronic zoonosis, to which sheep are highly susceptible. Previously, we found that Kazakh sheep with different MHC haplotypes differed in CE infection. Sheep with haplotype MHCMvaIbc-SacIIab-Hin1Iab were resistant to CE infection, while their counterparts without this haplotype were not. MicroRNAs (miRNAs), a class of small non-coding RNAs, are key regulators of gene expression at the post-transcriptional level and play essential roles in fundamental biological processes such as development and metabolism. To identify microRNA controlling resistance to CE in the early stage of infection, microRNA profiling was conducted in the intestinal tissue of sheep with resistant and non-resistant MHC haplotypes after peroral infection with E. granulosus eggs. A total of 351 known and 186 novel miRNAs were detected in the resistant group, against 353 known and 129 novel miRNAs in the non-resistant group. Among these miRNAs, 83 known miRNAs were significantly differentially expressed, including 75 up-regulated and 8 down-regulated miRNAs. Among these known microRNAs, miR-21-3p, miR-542-5p, miR-671, miR-134-5p, miR-26b, and miR-27a showed a significantly higher expression in CE-resistant sheep compared to the CE-non-resistant library, with the FC > 3. Functional analysis showed that they were NF-kB pathway-responsive miRNAs, which are involved in the inflammation process. The results suggest that these microRNAs may play important roles in the response of intestinal tissue to E. granulosus.Key words: microRNA, Cystic Echinococcosis, Sheep, MHC haplotype. Résumé -Profilage des microARN du tissu intestinal des moutons Kazakh après infection expérimentale parEchinococcus granulosus, en utilisant une approche à haut débit. L'échinococcose kystique (EK), causée par infection par le stade larvaire du cestode Echinococcus granulosus, est une zoonose chronique, à laquelle les moutons sont particulièrement sensibles. Auparavant, nous avons constaté que les moutons Kazakh avec différents haplotypes de CMH différaient dans l'infection par EK. Les moutons avec l'haplotype MHC MvaIbc-SacIIabHin1Iab étaient résistants à l'infection par EK, tandis que leurs homologues sans cet haplotype ne l'étaient pas. Les microARN (miARN), une classe de petits ARN non-codants, sont des régulateurs-clés de l'expression des gènes au niveau post-transcriptionnel et jouent des rôles essentiels dans les processus biologiques fondamentaux tels que le développement et le métabolisme. Pour identifier les microARN contrôlant la résistance à EK dans le stade précoce de l'infection, un profilage des microARN a été réalisé dans le tissu de l'intestin des moutons d'haplotypes MHC résistants et non-résistants, après infection perorale par des oeufs d'E. granulosus. OPEN ACCESS RESEARCH ARTICLEplus élevée chez les moutons résistants à EK comparés à la bibliothèque des non-résistants, avec FC > 3. Une analyse fonctionnelle a montré que ce sont de...
The homology-independent targeted integration (HITI) strategy enables effective CRISPR/Cas9-mediated knockin of therapeutic genes in nondividing cells in vivo, promising general therapeutic solutions for treating genetic diseases like Xlinked juvenile retinoschisis. Herein, supramolecular nanoparticle (SMNP) vectors are used for codelivery of two DNA plasmids-CRISPR-Cas9 genome-editing system and a therapeutic gene, Retinoschisin 1 (RS1)-enabling clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (CRISPR/Cas9) knockin of the RS1 gene with HITI. Through small-scale combinatorial screenings, two SMNP vectors, with Cas9 and single guide RNA (sgRNA)plasmid in one and Donor-RS1 and green fluorescent protein (GFP)-plasmid in the other, with optimal delivery performances are identified. These SMNP vectors are then employed for CRISPR/Cas9 knockin of RS1/GFP genes into the mouse Rosa26 safe-harbor site in vitro and in vivo. The in vivo study involves intravitreally injecting the two SMNP vectors into the mouse eyes, followed by repeated ocular imaging by fundus camera and optical coherence tomography, and pathological and molecular analyses of the harvested retina tissues. Mice ocular organs retain their anatomical integrity, a single-copy 3.0-kb RS1/GFP gene is precisely integrated into the Rosa26 site in the retinas, and the integrated RS1/GFP gene is expressed in the retinas, demonstrating CRISPR/Cas9 knockin of RS1/GFP gene.
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