Zero to Three

Since the mid-1990s the changing electricity generation technology mix has been associated with electricity sector carbon dioxide emissions that are 32 percent lower than their 2005 levels. What role have wholesale power markets played in enabling this decarbonization? We examine this question using annual state-level data for 1990 to 2020. The institutional complexity of the history of wholesale power markets and regulatory restructuring informs our block treatment window difference-in-differences empirical strategy, in which we compare generation in restructured states to different control states across different time periods. Our results suggest that lower-carbon generating sources appear to be built in greater numbers in wholesale markets relative to the control states up to 2012, largely but not entirely due to the substitution of natural gas for other fossil fuel generation, namely coal. In more recent years, restructured states and control states look more similar in their generation mix. Our event study results support this conclusion. JEL Classification: L51, Economics of Regulation; L94, Electric Utilities; Q35, Hydrocarbon Resources; Q48, Energy: Government Policy; Q53, Air Pollution; Water Pollution; Noise; Hazardous Waste; Solid Waste; Recycling

Abstract The adage that "the easy oil is gone" underscores the need for continuous advancements in drilling technologies. Among these innovations is slim hole drilling, which has gained relevance across various applications, including casing drilling and mineral exploration. The objective of this paper is to identify and assess operational strategies for safely detecting and circulating gas influxes from a borehole to support the development of effective and reliable well-control procedures tailored to slim hole environments. The scenario in question involves a mineral exploration rig equipped with slick 70 mm (2¾ in.) OD drill rods drilling a 76 mm (3 in.) ID borehole to a depth of 1,600 m. Transient flow simulation tools were employed to evaluate the hydraulics and well-control phenomena associated with such a slim-hole configuration. Influx scenarios included fractured formations combined with lost circulation and the risks of encountering small high-pressure cavities in impermeable rock. Sensitivity analyses were performed to evaluate the effects of varying kick size and intensity. The reduced annular clearance between drill string and borehole introduces substantial differences from conventional drilling hydraulics and well-control principles. Simulations using conventional oilfield drilling fluids revealed that non-Newtonian fluids led to excessive equivalent circulating densities (ECD), making the use of solids-free Newtonian water-based fluids essential. Rotary speeds typical of slim-hole mineral exploration operations, up to 300 rpm, were shown to contribute an additional 0.36 SG (3 ppg) to ECD. Kick simulations revealed that standard well-control practices, such as immediate shut-in followed by the Driller's Method for circulating out a kick, were not optimal in slim-hole conditions. Depending on annular geometry and influx characteristics, a dynamic kill approach offered safer and more efficient control. The scenarios are based on a mineral exploration rig equipped with a 150 mm (6 in.) ID annular blowout preventer venting into a mud gas separator. Despite its limitations, transient simulations demonstrated that such minimal equipment is capable of managing the envisaged well control events. The findings supported the development of tailored well-control procedures, improving operational safety and reliability in such a complex environment. The use of transient flow simulations to evaluate well-control operations and equipment for slim-hole wells is described. The examples highlight how local geology dictates the well-control measures that need to be implemented. The findings demonstrate how modeling can guide equipment selection, procedure development, and training programs, enhancing preparedness for slim-hole drilling operations.

Vascular smooth muscle cell (VSMC) diversification drives atherosclerotic coronary artery disease (CAD). Mechanisms governing these cell state transitions remain unclear. We applied multiomic single-cell profiling, epitope mapping, and spatial transcriptomics across 27 human coronary arteries, identifying fibroblast activation protein (FAP) as a marker of modulated VSMCs. Lineage tracing in mice indicated that FAP+ cells originate from Myh11+ VSMCs, and FAP PET imaging in CAD patients showed plaque uptake. FAP+ cells states resided in the macrophage-rich neo-intima. Therapeutically, we developed an anti-FAP bispecific T-cell engager, which reduced plaque burden and remodeled the stromal-immune microenvironment through T-cell clonal expansion. Our study delivers a single-cell and spatial atlas of human CAD, establishes FAP as a marker of modulated VSMCs, and highlights immunotherapy for lipid-independent targets.

There remain large variations in cochlear implant auditory-speech perception ability, which are poorly explained by patient history factors. This study aims to better understand how combinations of speech processing limitations, identified using psychophysical and functional brain imaging measures, are associated with long-term speech understanding outcomes of new cochlear implant recipients. Forty-three cochlear implant recipients (44 ears) were recruited to participate in a longitudinal study to evaluate limitations at various locations along the auditory processing pathway. At the periphery, intra-cochlear neural health was estimated using focused psychophysical thresholds. Higher-order across-electrode intensity discrimination ability was assessed using psychophysical "spectral tilt." At the cortical level, cross-modal activation of the auditory cortex was assessed using functional near-infrared spectroscopy. Speech understanding outcomes were evaluated in competing multi-talker babble noise at 1-year post-device switch-on. We found that greater abilities to discriminate smaller across-electrode stimulation intensities ( R2 = 0.138, p = 0.047) and smaller cross-modal activations of the auditory cortex ( R2 = 0.216, p = 0.01) were significantly associated with better speech understanding outcomes in cochlear implant recipients. Furthermore, both measures contributed significantly to better predict speech understanding outcomes in a multiple regression model (Adj R2 = 0.312, p = 0.009). The results suggest that recipient-specific psychophysical and functional brain imaging metrics contribute significantly toward individual differences in speech understanding outcomes. Different recipients can be affected by different limitations, which can be identified using these tests, and therefore lead to potential patient-specific methods to improve their speech understanding.

High‐field magnetic resonance imaging (MRI) scanners have been the primary choice for high‐quality soft tissue imaging, but use remains limited by high costs, complex siting requirements, and infrastructure demands. In contrast, preclinical MRI scanners with a low‐field permanent magnet are increasingly being adopted in research due to their lower cost, portability, and ease of installation. However, their reduced intrinsic signal‐to‐noise ratio (SNR) and limited pulse sequence availability constrain their use for high‐quality ex vivo soft tissue characterization. To overcome these limitations, we established an optimized imaging protocol using a compact 1‐Tesla permanent‐magnet preclinical MRI system for high‐quality ex vivo soft tissue imaging. A 25‐mm (length) × 23‐mm (diameter) coil was employed to image both human and animal soft tissue samples. Protocol optimization focused on four key parameters, namely the number of excitations (NEX), repetition time (TR), echo time (TE), and slice thickness, using a 3D gradient‐echo (3D‐GRE) acquisition sequence. Increasing the NEX enhances the SNR through signal averaging, extending the TR further improves the SNR, reducing the TE provides improved tissue contrast, and thinner slices improve image resolution. A typical 3D‐GRE MRI scan using the imaging protocol takes 13 hr. The approach also incorporates standardized ex vivo sample preparation, including staining with gadolinium diethylenetriaminepentaacetic acid to enhance tissue contrast, and background proton signal suppression and sample immobilization with Fluorinert or agarose. Collectively, these optimizations substantially improve the performance of low‐field permanent‐magnet MRI scanners for high‐quality soft tissue imaging. This set of protocols provides a practical framework for researchers to expand the capabilities of low‐field MRI systems, enabling more widespread access to advanced tissue characterization methods that support disease research and therapeutic development. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC.Basic Protocol 1: Tissue sample preparation with Gd‐DTPA and FluorinertAlternate Protocol: Tissue sample preparation with Gd‐DTPA and agaroseBasic Protocol 2: High‐quality sample imaging with 1‐Tesla compact MRI

Immunoheterogeneity is a potential hindrance to the maximum applicability of mesenchymal stromal cells (MSCs) in biotherapeutics and regenerative medicine. The primary causes of this heterogeneityare donor variation, diversity of tissue sources, and the culture conditionsunder which MSCs clonally propagate. Immunoheterogeneity could manifest as functional heterogeneity within these stem cells, affecting their ability to differentiate into adult tissue lineages. The impacted healthy human third molar tooth is a rich source of MSCs, which are isolatable both from the dental pulp and the dental follicle. Dental pulp stem cells (DPSCs), though widely characterized for their multipotential lineage differentiation, demonstrate considerable immunoheterogeneity, which demands the enrichment of these stem cells post-isolation. The aim of this study is to enrich and understand the functional differences in sub-populations of MSCs towards a specific lineage. In this protocol, we have highlighted detailed, stepwise methods for sorting and culturing post-sort and optimized methods for quantifying the differentiation of sorted DPSCs towards the osteogenic lineage. Our workflow is designed with additional considerations for cell sorter availability, maintenance of cell viability during long-distance transport to a state-of-the-art facility, and optimization of post-sorting cell culture and downstream experiments. This protocol can be universally followed to understand functional differences within MSCs isolated from different tissue sources and their implications for multi-lineage differentiation. © 2025 Wiley Periodicals LLC. Basic Protocol 1: Assessing multiparametric expression of MSC surface markers and single cell sorting for enrichment of stem cells Support Protocol 1: Antibody panel design Support Protocol 2: MSC culture, preparation of cell suspension, and sample staining Support Protocol 3: Setting up controls Support Protocol 4: Analysis and data interpretation Basic Protocol 2: Quantitative and qualitative assessment of osteogenic differentiation of post-sorted DPSCs Support Protocol 5: Differentiation of MSCS (pre-sort and post-sort considerations) Support Protocol 6: Cytochemical staining and quantification Support Protocol 7: Immunofluorescence staining Support Protocol 8: RNA isolation and qPCR-based characterization.

Aim: Representing about 15% of lung cancers, small cell lung cancer (SCLC) is an extremely aggressive disease characterized by rapid growth and early spread, leading to dismal clinical outcomes. In this study, we aimed to investigate the dual roles of exosomal long non-coding RNA (lncRNA) LYPLAL1-DT (LYPLAL1 divergent transcript) in both tumor cells and vascular endothelial cells. Methods: The circulating levels of LYPLAL1-DT were measured using real-time polymerase chain reaction in 13 SCLC patients and 21 normal controls. Exosomes from the supernatant of cell culture medium or serum were extracted through ultracentrifugation and dyed with PKH67 green fluorescent cell linker to identify internalization. Migration and invasion assay, colony formation, Cell Counting Kit-8 (CCK-8), and tube formation assays were used to assess the malignant effects of extracellular RNAs (exRNAs) LYPLAL1-DT in exosomes. Results: Exosomal LYPLAL1-DT is upregulated in SCLC patients and plays a dual role in promoting tumor cell aggressiveness and enhancing pro-angiogenic behavior in endothelial cells, thereby accelerating SCLC progression. Mechanistically, LYPLAL1-DT functions as a competing endogenous RNA, exerting its effects through the miR-204-5p/profilin-2, miR-204-5p/B-cell lymphoma 2 and miR-204-5p/sirtuin 1 regulatory axes. These pathways underscore the pleiotropic effects of exosomal LYPLAL1-DT and underscore its value as a promising therapeutic target. Conclusion: In the current study, we investigated the bidirectional communication mediated by exRNA LYPLAL1-DT between SCLC and endothelial cells, while also exploring its potential regulatory targets. This research provides a potential circulating biomarker for the diagnosis, prognosis, and treatment of SCLC.

Antibodies have emerged as central components of therapeutic strategies against viral infectious diseases, functioning as key effectors in both prevention and treatment. While traditional antibody discovery has relied heavily on high-throughput screening, the field is now shifting toward rational antibody design, which requires integrative insights into sequence-structure-function relationships. However, existing resources provide a valuable foundation but remain limited in scope, highlighting the need for a standardized and well-annotated antibody database that integrates multidimensional features to further support systematic exploration, cross-pathogen comparison, and rational antibody design. Here, we introduce the Multidimensional Antiviral Antibody Database (MAAD; http://www.raabmd.org/raab/index), a curated platform dedicated to antibody, nanobody and single-chain variable fragment targeting three high-impact RNA virus families, Coronaviridae (SARS-CoV-1, SARS-CoV-2, MERS-CoV), Orthomyxoviridae (influenza virus), and Pneumoviridae (respiratory syncytial virus, human metapneumovirus), which were selected due to the large, high-quality datasets accumulated in recent years. MAAD further incorporates a suite of interactive analysis modules, including CDR and germline annotation, similarity-based sequence analysis, sequence-based clustering and structure-guided identification of antigen-antibody interface residues, complemented by per-site entropy and mutation rate profiling. These features enable in-depth exploration of antibody sequence characteristics, thereby facilitating functional and structural insights for rational antibody design. Together, by bridging antibody sequence, structure and function, MAAD offers an open and standardized platform that advances comparative antiviral research and supports therapeutic antibody discovery.

This article mainly describes a fermentation and purification method for expressing recombinant collagen protein in Escherichia coli. The method comprises constructing engineered bacteria expressing human type III collagen and adopting a strategy of feeding in batches for high-density fermentation. The rapid proliferation of bacterial cells is promoted at 37°C, and then the culture is inoculated into a fermentation tank with different carbon sources for growth. When glycerol is used as the main carbon source, the yield of recombinant collagen protein can reach 0.25 to 0.40 g/L. The method allows exploration of the differences in recombinant collagen production with different carbon sources in order to identify the most suitable fermentation medium component. The human type III collagen produced by the method has the typical structure of collagen, with high cell adhesion and the stability of tissue structure. Therefore, it can be used as the raw material for various collagen products, especially facial fillers, dressings, freeze-dried fibers, and gels. © 2025 Wiley Periodicals LLC. Basic Protocol: Fermentation and purification of recombinant human type III collagen expressed in Escherichia coli.

Abstract Synthetic multicellular inter‐communication systems are essential for understanding multicellular behaviors and developing cell‐based therapeutics. Synthetic receptors that allow programming of multicellular intercommunications through spatiotemporal control with light or chemical ligands are highly desirable but remain challenging. Here we propose a new strategy for designing multicellular inter‐communication systems using a synthetic conformation‐inducible kinase receptor (synCIKER). The synCIKER system is developed by engineering the extracellular domain of a tyrosine kinase receptor with domains responsive to inputs of chemical ligand, light or soluble protein, and rewiring the endogenous intracellular signaling pathways for prescribed transgene expression. The synCIKER system is demonstrated to sense these inputs, and generate customized secretory proteins for programming receiver cell behaviours such as surface receptor degradation and T cell‐mediated lysis in target cells. We also demonstrate the ability of synCIKER to construct cascade systems and reconfigure intelligent intercellular Boolean logic gates of OR, AND and INHIBIT for activating gene editing in receiver cells. The synCIKER platform is further exploited to modulate bi‐directional polarization of macrophages in a multi‐cellular system, highlighting its potential for designing synthetic intercellular communication systems for programming multicellular behaviors.