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Cancer, development, cellular growth and differentiation are governed by gene expression. Recent molecular and cellular advances to visualize and perturb the pathways of transcriptional regulation, nascent RNA processing, and protein trafficking at the single cell level have been developed. More recently, applications utilizing the green fluorescent marker (GFP) from Aequorea victoria have facilitated visualization of these molecular events in a living cell. Specifically, we will describe a novel approach to perturb cellular processes by labeling discrete cellular components of interest with GFP and subsequently altering/ablating them with a laser microbeam. ©2004 Copyright SPIE - The International Society for Optical Engineering.

Context: Alzheimer disease (AD) represents a major and increasing public health problem. If populations were identified with significantly lower or higher incidence rates of AD, the search for risk factors in the genesis of AD could be greatly enhanced.Objective: To compare incidence rates of dementia and AD in 2 diverse, elderly community-dwelling populations.Design: The Indianapolis-Ibadan Dementia Project, a longitudinal, prospective population-based study consisting of a baseline survey (1992-1993) and 2 subsequent follow-up waves after 2 years (1994-1995) and 5 years (1997-1998). Each wave followed a 2-stage design, with an in-home screening interview followed by a full diagnostic workup of a subsample of participants based on screening performance.Setting and Participants: A total of 2459 community-dwelling Yoruba residents of Ibadan, Nigeria, without dementia, and 2147 community-dwelling African American residents of Indianapolis, Ind, without dementia (all aged 65 years or older). The cohorts were followed up for a mean of 5.1 years and 4.7 years, respectively.Main Outcome Measures: Incident cases of dementia and AD in each of the 2 populations.Results: The age-standardized annual incidence rates were significantly lower among Yoruba than among African Americans for dementia (Yoruba, 1.35% [95% confidence interval [CI], 1.13%-1.56%]; African Americans, 3.24% [95% CI, 2.11%-4.38%]) and for AD (Yoruba, 1.15% [95% CI, 0.96%-1.35%]; African Americans, 2.52% [95% CI, 1.40%-3.64%]).Conclusion: This is the first report of incidence rate differences for dementia and AD in studies of 2 populations from nonindustrialized and industrialized countries using identical methods and the same group of investigators in both sites. Further explorations of these population differences may identify potentially modifiable environmental or genetic factors to account for site differences in dementia and AD. [ABSTRACT FROM AUTHOR]

Thyroid hormone receptor α1 (TRα1) undergoes nucleocytoplasmic shuttling and mediates gene expression in response to thyroid hormone (T3). In Resistance to Thyroid Hormone Syndrome α (RTHα), certain TRα1 mutants have higher affinity for nuclear corepressor 1 (NCoR1) and may form stable complexes that are not released in the presence of T3. Here, we examined whether NCoR1 modulates intranuclear mobility and nuclear retention of TRα1 or RTHα-associated mutants in transfected human cells, as a way of analyzing critical structural components of TRα1 and to further explore the correlation between mutations in TRα1 and aberrant intracellular trafficking. We found no significant difference in intranuclear mobility, as measured by fluorescence recovery after photobleaching, between TRα1 and select RTHα mutants, irrespective of NCoR1 expression. Nuclear-to-cytoplasmic fluorescence ratios of RTHα mutants, however, varied from TRα1 when NCoR1 was overexpressed, with a significant increase in nuclear retention for A263V and a significant decrease for A263S and R384H. In NCoR1-knockout cells, nuclear retention of A263S, A263V, P389R, A382P, C392X, and F397fs406X was significantly decreased compared to control (wild-type) cells. Luciferase reporter gene transcription mediated by TRα1 was significantly repressed by both NCoR1 overexpression and NCoR1 knockout. Most RTHα mutants showed minimal induction regardless of NCoR1 levels, but T3-mediated transcriptional activity was decreased for R384C and F397fs406X when NCoR1 was overexpressed, and also decreased for N359Y in NCoR1-knockout cells. Our results suggest a complex interaction between NCoR1 and RTHα mutants characterized by aberrant intracellular localization patterns and transcriptional activity that potentially arise from variable repressor complex stability, and may provide insight into RTHα pathogenesis on a molecular and cellular level., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Macrophages contribute to the induction and resolution of inflammation and play a central role in chronic low-grade inflammation in cardiovascular diseases caused by atherosclerosis. Human milk oligosaccharides (HMOs) are complex unconjugated glycans unique to human milk that benefit infant health and act as innate immune modulators. Here, we identify the HMO 3'sialyllactose (3'SL) as a natural inhibitor of TLR4-induced low-grade inflammation in macrophages and endothelium. Transcriptome analysis in macrophages revealed that 3'SL attenuates mRNA levels of a selected set of inflammatory genes and promotes the activity of liver X receptor (LXR) and sterol regulatory element binding protein-1 (SREBP1). These acute antiinflammatory effects of 3'SL were associated with reduced histone H3K27 acetylation at a subset of LPS-inducible enhancers distinguished by preferential enrichment for CCCTC-binding factor (CTCF), IFN regulatory factor 2 (IRF2), B cell lymphoma 6 (BCL6), and other transcription factor recognition motifs. In a murine atherosclerosis model, both s.c. and oral administration of 3'SL significantly reduced atherosclerosis development and the associated inflammation. This study provides evidence that 3'SL attenuates inflammation by a transcriptional mechanism to reduce atherosclerosis development in the context of cardiovascular disease.

Profound functional switch of key regulatory factors may play a major role in homeostasis and disease. Dysregulation of circadian rhythm (CR) is strongly implicated in cancer with mechanisms poorly understood. We report here that the function of REV-ERBα, a major CR regulator of the orphan nuclear receptor subfamily, is dramatically altered in tumors in both its genome binding and functional mode. Loss of CR is linked to a functional inversion of REV-ERBα from a repressor in control of CR and metabolic gene programs in normal tissues to a strong activator in different cancers. Through changing its association from NCoR/HDAC3 corepressor complex to BRD4/p300 coactivators, REV-ERBα directly activates thousands of genes including tumorigenic programs such as MAPK and PI3K-Akt signaling. Functioning as a master transcriptional activator, REV-ERBα partners with pioneer factor FOXA1 and directly stimulates a large number of signaling genes, including multiple growth factors, receptor tyrosine kinases, RASs, AKTs, and MAPKs. Moreover, elevated REV-ERBα reprograms FOXA1 to bind new targets through a BRD4-mediated increase in local chromatin accessibility. Pharmacological targeting with SR8278 diminishes the function of both REV-ERBα and FOXA1 and synergizes with BRD4 inhibitor in effective suppression of tumorigenic programs and tumor growth. Thus, our study revealed a functional inversion by a CR regulator in driving gene reprogramming as an unexpected paradigm of tumorigenesis mechanism and demonstrated a high effectiveness of therapeutic targeting such switch., Competing Interests: Competing interests statement:The authors declare no competing interest.

Vitamin D signals through the vitamin D receptor (VDR) to induce its end-organ effects. Hepatic stellate cells control development of liver fibrosis in response to stressors and vitamin D signaling decreases fibrogenesis. VDR expression in hepatocytes is low in healthy liver, and the role of VDR in hepatocyte proliferation is unclear. Hepatocyte-VDR null mice (hVDR) were used to assess the role of VDR and vitamin D signaling in hepatic regeneration. hVDR mice have impaired liver regeneration and impaired hepatocyte proliferation associated with significant differential changes in bile salts. Notably, mice lacking hepatocyte VDR had significant increases in expression of conjugated bile acids after partial hepatectomy, consistent with failure to normalize hepatic function by the 14-day time point tested. Real-time PCR of hVDR and control livers showed significant changes in expression of cell-cycle genes including cyclins D1 and E1 and cyclin-dependent kinase 2. Gene expression profiling of hepatocytes treated with vitamin D or control showed regulation of groups of genes involved in liver proliferation, hepatitis, liver hyperplasia/hyperproliferation, and liver necrosis/cell death. Together, these studies demonstrate an important functional role for VDR in hepatocytes during liver regeneration. Combined with the known profibrotic effects of impaired VDR signaling in stellate cells, the studies provide a mechanism whereby vitamin D deficiency would both reduce hepatocyte proliferation and permit fibrosis, leading to significant liver compromise., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

Hydrogenases catalyze hydrogen/proton interconversion that is normally electrochemically reversible (having minimal overpotential requirement), a special property otherwise almost exclusive to platinum metals. The mechanism of [NiFe]-hydrogenases includes a long-range proton-coupled electron-transfer process involving a specific Ni-coordinated cysteine and the carboxylate of a nearby glutamate. A variant in which this cysteine has been exchanged for selenocysteine displays two distinct changes in electrocatalytic properties, as determined by protein film voltammetry. First, proton reduction, even in the presence of H 2 (a strong product inhibitor), is greatly enhanced relative to H 2 oxidation: this result parallels a characteristic of natural [NiFeSe]-hydrogenases which are superior H 2 production catalysts. Second, an inflection (an S -shaped "twist" in the trace) appears around the formal potential, the small overpotentials introduced in each direction (oxidation and reduction) signaling a departure from electrocatalytic reversibility. Concerted proton-electron transfer offers a lower energy pathway compared to stepwise transfers. Given the much lower proton affinity of Se compared to that of S, the inflection provides compelling evidence that concerted proton-electron transfer is important in determining why [NiFe]-hydrogenases are reversible electrocatalysts.

Background: The incidence of cancer diagnosed during pregnancy is increasing. Data relating to investigation and management, as well as maternal and foetal outcomes is lacking in a United Kingdom (UK) population., Methods: In this retrospective study we report data from 119 patients diagnosed with cancer during pregnancy from 14 cancer centres in the UK across a five-year period (2016-2020)., Results: Median age at diagnosis was 33 years, with breast, skin and haematological the most common primary sites. The majority of cases were new diagnoses (109 patients, 91.6%). Most patients were treated with radical intent (96 patients, 80.7%), however, gastrointestinal cancers were associated with a high rate of palliative intent treatment (63.6%). Intervention was commenced during pregnancy in 68 (57.1%) patients; 44 (37%) had surgery and 31 (26.1%) received chemotherapy. Live births occurred in 98 (81.7%) of the cases, with 54 (55.1%) of these delivered by caesarean section. Maternal mortality during the study period was 20.2%., Conclusions: This is the first pan-tumour report of diagnosis, management and outcomes of cancer diagnosed during pregnancy in the UK. Our findings demonstrate proof of concept that data collection is feasible and highlight the need for further research in this cohort of patients., (© 2024. The Author(s).)

Despite numerous female contraceptive options, nearly half of all pregnancies are unintended. Family planning choices for men are currently limited to unreliable condoms and invasive vasectomies with questionable reversibility. Here, we report the development of an oral contraceptive approach based on transcriptional disruption of cyclical gene expression patterns during spermatogenesis. Spermatogenesis involves a continuous series of self-renewal and differentiation programs of spermatogonial stem cells (SSCs) that is regulated by retinoic acid (RA)-dependent activation of receptors (RARs), which control target gene expression through association with corepressor proteins. We have found that the interaction between RAR and the corepressor silencing mediator of retinoid and thyroid hormone receptors (SMRT) is essential for spermatogenesis. In a genetically engineered mouse model that negates SMRT-RAR binding (SMRT mRID mice), the synchronized, cyclic expression of RAR-dependent genes along the seminiferous tubules is disrupted. Notably, the presence of an RA-resistant SSC population that survives RAR de-repression suggests that the infertility attributed to the loss of SMRT-mediated repression is reversible. Supporting this notion, we show that inhibiting the action of the SMRT complex with chronic, low-dose oral administration of a histone deacetylase inhibitor reversibly blocks spermatogenesis and fertility without affecting libido. This demonstration validates pharmacologic targeting of the SMRT repressor complex for non-hormonal male contraception., Competing Interests: Competing interests statement:The authors disclose that the reviewer Y.S. jointly authored a commentary on an unrelated topic with RME in 2022 (PMID: 35344400).

Mitochondrial dysfunction is a characteristic trait of human and rodent obesity, insulin resistance and fatty liver disease. Here we show that high-fat diet (HFD) feeding causes mitochondrial fragmentation in inguinal white adipocytes from male mice, leading to reduced oxidative capacity by a process dependent on the small GTPase RalA. RalA expression and activity are increased in white adipocytes after HFD. Targeted deletion of RalA in white adipocytes prevents fragmentation of mitochondria and diminishes HFD-induced weight gain by increasing fatty acid oxidation. Mechanistically, RalA increases fission in adipocytes by reversing the inhibitory Ser637 phosphorylation of the fission protein Drp1, leading to more mitochondrial fragmentation. Adipose tissue expression of the human homolog of Drp1, DNM1L, is positively correlated with obesity and insulin resistance. Thus, chronic activation of RalA plays a key role in repressing energy expenditure in obese adipose tissue by shifting the balance of mitochondrial dynamics toward excessive fission, contributing to weight gain and metabolic dysfunction., (© 2024. The Author(s).)

Background: Cardiac metabolic dysfunction is a hallmark of heart failure (HF). Estrogen-related receptors ERRα and ERRγ are essential regulators of cardiac metabolism. Therefore, activation of ERR could be a potential therapeutic intervention for HF. However, in vivo studies demonstrating the potential usefulness of ERR agonist for HF treatment are lacking, because compounds with pharmacokinetics appropriate for in vivo use have not been available., Methods: Using a structure-based design approach, we designed and synthesized 2 structurally distinct pan-ERR agonists, SLU-PP-332 and SLU-PP-915. We investigated the effect of ERR agonist on cardiac function in a pressure overload-induced HF model in vivo. We conducted comprehensive functional, multi-omics (RNA sequencing and metabolomics studies), and genetic dependency studies both in vivo and in vitro to dissect the molecular mechanism, ERR isoform dependency, and target specificity., Results: Both SLU-PP-332 and SLU-PP-915 significantly improved ejection fraction, ameliorated fibrosis, and increased survival associated with pressure overload-induced HF without affecting cardiac hypertrophy. A broad spectrum of metabolic genes was transcriptionally activated by ERR agonists, particularly genes involved in fatty acid metabolism and mitochondrial function. Metabolomics analysis showed substantial normalization of metabolic profiles in fatty acid/lipid and tricarboxylic acid/oxidative phosphorylation metabolites in the mouse heart with 6-week pressure overload. ERR agonists increase mitochondria oxidative capacity and fatty acid use in vitro and in vivo. Using both in vitro and in vivo genetic dependency experiments, we show that ERRγ is the main mediator of ERR agonism-induced transcriptional regulation and cardioprotection and definitively demonstrated target specificity. ERR agonism also led to downregulation of cell cycle and development pathways, which was partially mediated by E2F1 in cardiomyocytes., Conclusions: ERR agonists maintain oxidative metabolism, which confers cardiac protection against pressure overload-induced HF in vivo. Our results provide direct pharmacologic evidence supporting the further development of ERR agonists as novel HF therapeutics., Competing Interests: Disclosures Dr Zhang is a cofounder of and consultant for Pelagos Pharmaceuticals Inc. The other authors report no conflicts of interest.

The nuclear receptor corepressor (NCoR) forms a complex with histone deacetylase 3 (HDAC3) that mediates repressive functions of unliganded nuclear receptors and other transcriptional repressors by deacetylation of histone substrates. Recent studies provide evidence that NCoR/HDAC3 complexes can also exert coactivator functions in brown adipocytes by deacetylating and activating PPARγ coactivator 1α (PGC1α) and that signaling via receptor activator of nuclear factor kappa-B (RANK) promotes the formation of a stable NCoR/HDAC3/PGC1β complex that coactivates nuclear factor kappa-B (NFκB)- and activator protein 1 (AP-1)-dependent genes required for osteoclast differentiation. Here, we demonstrate that activation of Toll-like receptor (TLR) 4, but not TLR3, the interleukin 4 (IL4) receptor nor the Type I interferon receptor, also promotes assembly of an NCoR/HDAC3/PGC1β coactivator complex. Receptor-specific utilization of TNF receptor-associated factor 6 (TRAF6) and downstream activation of extracellular signal-regulated kinase 1 (ERK1) and TANK-binding kinase 1 (TBK1) accounts for the common ability of RANK and TLR4 to drive assembly of an NCoR/HDAC3/PGC1β complex in macrophages. ERK1, the p65 component of NFκB, and the p300 histone acetyltransferase (HAT) are also components of the induced complex and are associated with local histone acetylation and transcriptional activation of TLR4-dependent enhancers and promoters. These observations identify a TLR4/TRAF6-dependent signaling pathway that converts NCoR from a corepressor of nuclear receptors to a coactivator of NFκB and AP-1 that may be relevant to functions of NCoR in other developmental and homeostatic processes., Competing Interests: Competing interests statement:C.K.G. is a cofounder, equity holder and member of the Scientific Advisory Board of Asteroid Therapeutics.

Introduction: Sustained-release intraocular implants provide a therapeutic option for open-angle glaucoma (OAG) and ocular hypertension (OHT) patients who are non-compliant with eyedrops. Currently, there are no published patient-reported outcome (PRO) measures that assess treatment satisfaction with intraocular implants. To address this gap, a new PRO instrument, the Allergan Satisfaction with Treatment Experience Questionnaire (ASTEQ), has been developed in accordance with Food and Drug Administration guidance., Methods: Qualitative research interviews were conducted among patients with OAG/OHT who had received three intraocular injections of a sustained-release bimatoprost (10 or 15 µg) implant within the clinical trial setting. A preliminary conceptual framework capturing treatment satisfaction concepts in glaucoma, as identified from the literature, was used to develop a semi-structured interview guide. A concept elicitation (CE) interview to identify aspects of the glaucoma treatment experience pertinent to intraocular implants provided content for a draft instrument. A cognitive debriefing (CD) interview to test the instrument's interpretability, relevance, and validity informed its subsequent refinement. Interview analysis followed a grounded theory approach to identify data patterns and relationships., Results: CE interviews (n = 19) indicated that participants' main considerations in rating satisfaction with implant treatment were physical comfort during preparation for the implant and implant administration, anxiety about the procedure, frequency of implant administration, possible side effects, convenience and accessibility of the implant, relationship with the clinician, and lifestyle fit. Draft ASTEQ revision based on CD interviews (n = 20) and readability tests yielded a nine-item ASTEQ instrument comprising satisfaction with overall implant experience and frequency of administration, occurrence/bother of immediate and long-term side effects, worry about implant administration and possible risks/side effects, and physical discomfort during preparation for the implant and implant administration., Conclusion: The ASTEQ instrument has demonstrated content validity in patients with OAG/OHT treated with a sustained-release bimatoprost implant. Further research is necessary to evaluate its psychometric properties., (© 2023. The Author(s).)