Liver-specific transgenic expression of human NTCP in rhesus macaques confers HBV susceptibility on primary hepatocytes.

Hepatitis B virus (HBV) poses a significant global health challenge, necessitating the urgent development of curative therapeutics. However, this progress is impeded by the lack of robust, immunocompetent preclinical animal models due to HBV's strict species specificity. We previously showed that vector-mediated expression of the HBV entry receptor, human sodium-taurocholate cotransporting polypeptide (hNTCP), renders macaques fully susceptible to HBV infection.

Generation of Rhesus Macaque Embryos with Expanded CAG Trinucleotide Repeats in the Gene.

Huntington's disease (HD) arises from expanded CAG repeats in exon 1 of the () gene. The resultant misfolded HTT protein accumulates within neuronal cells, negatively impacting their function and survival. Ultimately, HTT accumulation results in cell death, causing the development of HD.

A Multifunctional and Highly Adaptable Reporter System for CRISPR/Cas Editing.

CRISPR/Cas systems are some of the most promising tools for therapeutic genome editing. The use of these systems is contingent on the optimal designs of guides and homology-directed repair (HDR) templates. While this design can be achieved in silico, validation and further optimization are usually performed with the help of reporter systems.

The history, use, and challenges of therapeutic somatic cell and germline gene editing.

The advent of directed gene-editing technologies now over 10 years ago ushered in a new era of precision medicine wherein specific disease-causing mutations can be corrected. In parallel with developing new gene-editing platforms, optimizing their efficiency and delivery has been remarkable. With their development, there has been interest in using gene-editing systems for correcting disease mutations in differentiated somatic cells ex vivo or in vivo or for germline gene editing in gametes or 1-cell embryos to potentially limit genetic diseases in the offspring and in future generations.

CRISPR/Cas9 editing of the MYO7A gene in rhesus macaque embryos to generate a primate model of Usher syndrome type 1B.

Mutations in the MYO7A gene lead to Usher syndrome type 1B (USH1B), a disease characterized by congenital deafness, vision loss, and balance impairment. To create a nonhuman primate (NHP) USH1B model, CRISPR/Cas9 was used to disrupt MYO7A in rhesus macaque zygotes. The targeting efficiency of Cas9 mRNA and hybridized crRNA-tracrRNA (hyb-gRNA) was compared to Cas9 nuclease (Nuc) protein and synthetic single guide (sg)RNAs.

Rapid, accurate mapping of transgene integration in viable rhesus macaque embryos using enhanced-specificity tagmentation-assisted PCR.

Genome engineering is a powerful tool for research and the creation of novel model organisms and has growing clinical applications. Randomly integrating vectors, such as lentivirus- or transposase-based methods, are simple and easy to use but carry risks arising from insertional mutagenesis. Here we present enhanced-specificity tagmentation-assisted PCR (esTag-PCR), a rapid and accurate method for mapping transgene integration and copy number.

Metabolomics analysis of follicular fluid coupled with oocyte aspiration reveals importance of glucocorticoids in primate periovulatory follicle competency.

Gonadotropin administration during infertility treatment stimulates the growth and development of multiple ovarian follicles, yielding heterogeneous oocytes with variable capacity for fertilization, cleavage, and blastocyst formation. To determine how the intrafollicular environment affects oocyte competency, 74 individual rhesus macaque follicles were aspirated and the corresponding oocytes classified as failed to cleave, cleaved but arrested prior to blastulation, or those that formed blastocysts following in vitro fertilization. Metabolomics analysis of the follicular fluid (FF) identified 60 unique metabolites that were significantly different between embryo classifications, of which a notable increase in the intrafollicular ratio of cortisol to cortisone was observed in the blastocyst group.

TET family regulates the embryonic pluripotency of porcine preimplantation embryos by maintaining the DNA methylation level of .

The ten-eleven translocation (TET) family (TET1/2/3) initiates conversion of 5-methylcytosine to 5-hydroxymethylcytosine, thereby orchestrating the DNA demethylation process and changes in epigenetic marks during early embryogenesis. In this study, CRISPR/Cas9 technology and a TET-specific inhibitor were applied to elucidate the role of TET family in regulating pluripotency in preimplantation embryos using porcine embryos as a model. Disruption of unexpectedly resulted in the upregulation of and transcripts, although there was no change to the level of DNA methylation in the promoter of .

Use of gene-editing technology to introduce targeted modifications in pigs.

Pigs are an important resource in agriculture and serve as a model for human diseases. Due to their physiological and anatomical similarities with humans, pigs can recapitulate symptoms of human diseases, making them a useful model in biomedicine. However, in the past pig models have not been widely used partially because of the difficulty in genetic modification.

CRISPR/Cas9-Mediated Gene Targeting during Embryogenesis in Swine.

Ability to disrupt genes is essential in elucidating gene function. Unlike rodents or amphibians, it has been difficult to generate gene-targeted embryos in large animals. Therefore, studies of early embryo development have been hampered in large animals.

Biallelic modification of IL2RG leads to severe combined immunodeficiency in pigs.

Background: Pigs with SCID can be a useful model in regenerative medicine, xenotransplantation, and cancer cell transplantation studies. Utilizing genome editing technologies such as CRISPR/Cas9 system allows us to generate genetically engineered pigs at a higher efficiency. In this study, we report generation and phenotypic characterization of IL2RG knockout female pigs produced through combination of CRISPR/Cas9 system and SCNT.

Anti-Inflammatory Effects of Artemisia Leaf Extract in Mice with Contact Dermatitis In Vitro and In Vivo.

The leaves of Artemisia argyi Lev. et Vant. and A.

Alpha-1,3-galactosyltransferase-deficient miniature pigs produced by serial cloning using neonatal skin fibroblasts with loss of heterozygosity.

Objective: Production of alpha-1,3-galactosyltransferase (GT)-deficient pigs is essential to overcome xenograft rejection in pig-to-human xenotransplantation. However, the production of such pigs requires a great deal of cost, time, and labor. Heterozygous GT knockout pigs should be bred at least for two generations to ultimately obtain homozygote progenies.

Increased and prolonged human norovirus infection in RAG2/IL2RG deficient gnotobiotic pigs with severe combined immunodeficiency.

Application of genetically engineered (GE) large animals carrying multi-allelic modifications has been hampered by low efficiency in production and extended gestation period compared to rodents. Here, we rapidly generated RAG2/IL2RG double knockout pigs using direct injection of CRISPR/Cas9 system into developing embryos. RAG2/IL2RG deficient pigs were immunodeficient, characterized by depletion of lymphocytes and either absence of or structurally abnormal immune organs.

Swept source digital holographic phase microscopy.

We propose a swept source-based digital holographic phase microscopy technique. By scanning source wavelength, a series of on-axis interferograms can be obtained for accurate determination of the sample phase using spectral domain interferometry. With these sample spectra, sources of undesirable interference artifacts, often significant in holographic systems, can be identified and avoided by placing the sample signal at a spectral frequency with a clean background.

Anti-inflammatory activity of Kochia scoparia fruit on contact dermatitis in mice.

The mature fruit of Kochia scoparia (L.) Schrad. is widely administered in China and Korea as a medicinal herb for treatment of skin diseases, diabetes mellitus and rheumatoid arthritis.

Anti-inflammatory Effects of Sanguisorbae Radix on Contact Dermatitis Induced by Dinitrofluorobenzene in Mice.

Objective: To investigate the anti-inflflammatory effects of Sanguisorbae Radix on contact dermatitis (CD).

Methods: Mice were sensitized by painting 30 µL of 1-fluoro-2,4-dinitrofluorobenzene (DNFB) onto each ear for 3 days. Four days later, mice were challenged by painting with 50 µL of DNFB onto the shaved dorsum every 2 days.

Pig oocyte activation using a Zn²⁺ chelator, TPEN.

Artificial oocyte activation is a critical step during SCNT. Most current activation protocols focus on inducing an increase in the intracellular free Ca(2+) concentration of the oocyte. Here, we have used a zinc chelator, TPEN, to enhance the efficiency of oocyte activation during SCNT.

Targeted disruption of Ataxia-telangiectasia mutated gene in miniature pigs by somatic cell nuclear transfer.

Ataxia telangiectasia (A-T) is a recessive autosomal disorder associated with pleiotropic phenotypes, including progressive cerebellar degeneration, gonad atrophy, and growth retardation. Even though A-T is known to be caused by the mutations in the Ataxia telangiectasia mutated (ATM) gene, the correlation between abnormal cellular physiology caused by ATM mutations and the multiple symptoms of A-T disease has not been clearly determined. None of the existing ATM mouse models properly reflects the extent to which neurological degeneration occurs in human.

Flytrap-inspired robot using structurally integrated actuation based on bistability and a developable surface.

The Venus flytrap uses bistability, the structural characteristic of its leaf, to actuate the leaf's rapid closing motion for catching its prey. This paper presents a flytrap-inspired robot and novel actuation mechanism that exploits the structural characteristics of this structure and a developable surface. We focus on the concept of exploiting structural characteristics for actuation.