Cholestenoic acid as endogenous epigenetic regulator decreases hepatocyte lipid accumulation in vitro and in vivo.

Cholestenoic acid (CA) has been reported as an important biomarker of many severe diseases, but its physiological and pathological roles remain unclear. This study aimed to investigate the potential role of CA in hepatic lipid homeostasis. Enzyme kinetic studies revealed that CA specifically activates DNA methyltransferases 1 (DNMT1) at low concentration with EC50 = 1.99 × 10-6 M and inhibits the activity at higher concentration with IC50 = 9.13 × 10-6 M, and specifically inhibits DNMT3a, and DNMT3b activities with IC50= 8.41 × 10-6 M and IC50= 4.89 × 10-6 M, respectively. In a human hepatocyte in vitro model of high glucose (HG)-induced lipid accumulation, CA significantly increased demethylation of 5mCpG in the promoter regions of over 7,000 genes, particularly those involved in master signaling pathways such as calcium-AMPK and 0.0027 at 6 h. RNA sequencing analysis showed that the downregulated genes are affected by CA encoding key enzymes, such as PCSK9, MVK, and HMGCR, which are involved in cholesterol metabolism and steroid biosynthesis pathways. In addition, untargeted lipidomic analysis showed that CA significantly reduced neutral lipid levels by 60% in the cells cultured in high-glucose media. Administration of CA in mouse metabolic dysfunction-associated steatotic liver disease (MASLD) models significantly decreases lipid accumulation, suppresses the gene expression involved in lipid biosynthesis in liver tissues, and alleviates liver function. This study shows that CA as an endogenous epigenetic regulator decreases lipid accumulation via epigenetic regulation. The results indicate that CA can be considered a potential therapeutic target for the treatment of metabolic disorders.NEW & NOTEWORTHY To our knowledge, this study is the first to identify the mitochondrial monohydroxy bile acid cholestenoic acid (CA) as an endogenous epigenetic regulator that regulates lipid metabolism through epigenome modification in human hepatocytes. The methods used in this study are all big data analysis, and the results of each part show the global regulation of CA on human hepatocytes rather than narrow point effects.

Daisy-chain gene drives for the alteration of local populations.

If they are able to spread in wild populations, CRISPR-based gene-drive elements would provide new ways to address ecological problems by altering the traits of wild organisms, but the potential for uncontrolled spread tremendously complicates ethical development and use. Here, we detail a self-exhausting form of CRISPR-based drive system comprising genetic elements arranged in a daisy chain such that each drives the next. "Daisy-drive" systems can locally duplicate any effect achievable by using an equivalent self-propagating drive system, but their capacity to spread is limited by the successive loss of nondriving elements from one end of the chain. Releasing daisy-drive organisms constituting a small fraction of the local wild population can drive a useful genetic element nearly to local fixation for a wide range of fitness parameters without self-propagating spread. We additionally report numerous highly active guide RNA sequences sharing minimal homology that may enable evolutionarily stable daisy drive as well as self-propagating CRISPR-based gene drive. Especially when combined with threshold dependence, daisy drives could simplify decision-making and promote ethical use by enabling local communities to decide whether, when, and how to alter local ecosystems.

Designing DNA Nanotube Liquid Crystals as a Weak-Alignment Medium for NMR Structure Determination of Membrane Proteins.

Thirty percent of the human proteome is composed of membrane proteins that can perform a wide range of cellular functions and communications. They represent the core of modern medicine as the targets of about 50 % of all prescription pharmaceuticals. However, elucidating the structure of membrane proteins has represented a constant challenge, even in the modern era. To date, only a few hundred high-resolution structural models of membrane proteins are available. This chapter describes the emergence of DNA nanotechnology as a powerful tool for the structural characterization of membrane protein using solution-state nuclear magnetic resonance (NMR) spectroscopy. Here, we detail the large-scale synthesis of detergent-resistant DNA nanotubes that can be assembled into a dilute liquid crystal to be used as a weak-alignment media in solution NMR structure determination of membrane proteins.

Real-world economic value of a 21-gene assay in early-stage breast cancer.

OBJECTIVES: Value-based payment reforms shift cost-containment responsibilities to the physician. Although gene expression profiling (GEP) utilizing a 21-gene panel among patients with early-stage, axillary lymph node-negative, hormone receptor-positive, HER2/neu oncogene-negative breast cancer is able to identify a cohort that may achieve excellent outcomes without adjuvant chemotherapy, high up-front costs (list price, $4175) could dissuade usage. STUDY DESIGN: Retrospective review of consecutive patients with breast cancer treated at a single cancer center. METHODS: Chart review of 227 patients 70 years or younger with outpatient costs (ie, drug average sales price, reagent costs, physician charges) during first 6 months of treatment. RESULTS: Of these patients, 68% underwent GEP, with 52%, 43%, and 5% having low, intermediate, and high recurrence risk scores, respectively. Adjuvant chemotherapy was utilized less in genomically profiled cohorts (19% vs 29%; P = .08) and was consistent with recommendations of the recurrence scores. The mean 6-month outpatient costs were $24,955 with adjuvant chemotherapy and $2654 with hormonal therapy. Patients with stage II cancer undergoing GEP received adjuvant chemotherapy at a lower frequency (28.6% vs 86.7%), but patients with stage I cancer who underwent testing were slightly more likely to receive chemotherapy (15.8% vs 14%) because the test identified patients with higher-risk tumors. Universal GEP testing of patients with stage II cancer would have resulted in net savings of $11,494 per patient inclusive of test cost; stage I testing would have increased costs by $4505. Similar trends for grade 2/3 tumors (-$2394) and grade 1 tumors (+$6047) were noted. CONCLUSIONS: Universal GEP testing of women 70 years or younger with stage II or grade 2/3 lymph node-negative breast cancers would result in lower outpatient costs, inclusive of the diagnostic test, within the first 6-month episode of care.