The rockefeller university

Methods for describing and reporting the clinical and histologic characteristics of cutaneous tissue samples from patients with hidradenitis suppurativa (HS) are not currently standardized, limiting clinicians' and scientists' ability to uniformly record, report, and communicate about the characteristics of tissue used in translational experiments. A recently published consensus statement outlined morphological definitions of typical HS lesions, but no consensus has been reached regarding clinical characterization and examination of HS tissue samples. In this study, we aimed to establish a protocol for reporting histopathologic and clinical characteristics of HS tissue specimens. This study was conducted from May 2023 to August 2023. Experts in clinical care, dermatopathology, and translational research were recruited, and a modified Delphi technique was used to develop a protocol for histologic reporting and clinical characterization of submitted tissue specimens from patients with HS. A total of 27 experts participated (14 dermatologists, 3 fellowship-trained dermatopathologists, 3 plastic surgeons, 3 general surgeons, and 4 research scientists) in creating and reviewing protocols for the clinical and histopathological examination of HS tissue specimens. The protocols were formatted as a synoptic report and will help to consistently classify specimens in biobanks on the basis of histologic features and more accurately report and select samples used in translational research projects.

Multidrug resistance-associated protein 2 (MRP2) is an ATP-powered exporter important for maintaining liver homeostasis and a potential contributor to chemotherapeutic resistance. Using cryogenic electron microscopy (cryo-EM), we determine the structures of human MRP2 in three conformational states: an autoinhibited state, a substrate-bound pre-translocation state, and an ATP-bound post-translocation state. In the autoinhibited state, the cytosolic regulatory (R) domain plugs into the transmembrane substrate-binding site and extends into the cytosol to form a composite ATP-binding site at the surface of nucleotide-binding domain 2. Substrate displaces the R domain, permitting conformational changes necessary for transport. These observations suggest that the R domain functions as a selectivity gauge, where only at sufficiently high concentrations can the substrate effectively initiate transport. Comparative structural analyzes of MRP2 bound to various substrates, as determined in this study and others, reveal how MRP2 recognizes a diverse array of compounds, supporting its role in multidrug resistance.

BackgroundAdvanced practice nursing (APN) and advanced practice midwifery (APM) in Kenya are critical in healthcare system strengthening by expanding access to quality care, particularly in underserved regions. Globally, the International Council of Nurses (ICN) defines APN as a level of nursing practice that requires a minimum of a master's degree, along with advanced clinical training and competency. Similarly, APM involves midwives with advanced training to provide specialized maternal and neonatal care. In Kenya, the APN and APM concepts were launched in 2020 and are still nascent, and their practice is determined by the context as provided by the country credentialing the practice. Despite the strides made in developing the advanced roles, there is an eminent gap in understanding stakeholders' perceptions of the roles, scope of practice, and regulation.AimTo explore stakeholders' perceptions of the roles and regulations of APN and APM in Kenya.DesignAn exploratory qualitative study was undertaken as part of a larger formative research conducted between January 2022 and November 2023, utilizing a multimethod design that included a gap analysis of the scopes of practice for APN and APM in Kenya. Data collection was conducted among purposively sampled participants using a total of 7 focus group discussions (FGDs), with three of them conducted among 14 APN and APM students and 4 FGDs among 16 members of the Kenyan-based nursing and midwifery associations. Seven key informant interviews (KIIs) were also held with 2 medical doctors (OBGYN), two faculty, and 3 students (practicing APN). A thematic analysis approach was employed, and reporting was done according to the Standards for Reporting Qualitative Research.FindingsParticipant's perspectives were reflected in 5 themes: definition of APN/APM and scope; entry qualifications into APN/APM practice; roles of APN/APM; barriers to enacting APN/APM roles; the national policy and regulatory landscape for advanced practice roles.ConclusionThere were varied viewpoints on how APNs/APMs are defined in the Kenyan context. Similarly, views on the entry level qualifications into APN/APM were diverse. Many participants emphasized the importance of licensure, autonomy of practice, ability to manage patients independently, training, and competency. Well-defined policies and regulations can create an enabling environment for practice.

Fascin cross-links actin filaments (F-actin) into bundles that support tubular membrane protrusions including filopodia and stereocilia. Fascin dysregulation drives aberrant cell migration during metastasis, and fascin inhibitors are under development as cancer therapeutics. Here, we use cryo-EM, cryo-electron tomography coupled with custom denoising and computational modeling to probe human fascin-1's F-actin cross-linking mechanisms across spatial scales. Our fascin cross-bridge structure reveals an asymmetric F-actin binding conformation that is allosterically blocked by the inhibitor G2. Reconstructions of seven-filament hexagonal bundle elements, variability analysis and simulations show how structural plasticity enables fascin to bridge varied interfilament orientations, accommodating mismatches between F-actin's helical symmetry and bundle hexagonal packing. Tomography of many-filament bundles and modeling uncover geometric rules underlying emergent fascin binding patterns, as well as the accumulation of unfavorable cross-links that limit bundle size. Collectively, this work shows how fascin harnesses fine-tuned nanoscale structural dynamics to build and regulate micron-scale F-actin bundles.

Background While significant progress has been made in understanding the molecular characteristics of inflammatory skin diseases, their systemic impact warrants further investigation. A comprehensive large-scale study that examines systemic proteomic expression across the most common inflammatory skin diseases is currently lacking. Methods Serum from 38 alopecia areata/AA, 41 atopic dermatitis/AD, 21 psoriasis, 18 hidradenitis suppurativa/HS, and 25 vitiligo patients was analyzed using OLINK high-throughput multiplex assay explore panel and compared to 49 healthy-matched controls. Differentially expressed proteins/DEPs were defined using the criteria of fold change/|FCH| > 1.2, FDR < 0.1. Spearman analysis was also conducted to identify correlations between biomarkers and disease severity. Gene ontology of DEPs was performed using enrichr. Results Our results reveal both distinct and shared patterns of systemic dysregulation across the spectrum of inflammatory diseases. HS exhibited the highest level of dysregulation compared to healthy serum, followed by AA, AD, psoriasis, and vitiligo. The DEPs identified in HS showed the greatest overlap with those in psoriasis, while AA and AD also displayed significant similarity. HS and/or psoriasis primarily showed upregulation of markers belonging to T-cell activation/migration (interleukin/IL-2RA/CD40LG), innate immunity (IL-6/CXCL8/IL-8), Th1 (TNF/CXCL9/CXCL10), and Th17/22 (IL-17A/IL-20/CXCL1/LCN2) compared to normal. AA and/or AD were characterized by upregulation of markers of general inflammation (MMP12), T-cell activation/migration (IL-15/IL-16), T Helper 1/Th1 (IFNGR1/CXCL10), Th2 (IL-4R/CCL26/CCL27), Th17/22 (IL-19/IL-20/PI3) compared to controls. HS showed the largest dysregulation in cardiovascular/CV/atherosclerosis markers (PDGFA/SELP/MMP9) compared to controls (all FDR < 0.05). Spearman analysis captured multiple positive correlations between key immune markers (IL-4R/OX40/TNFRSF4/IL-17A/TNF) and respective clinical severity scores (e.g., SALT, SCORAD, IHS4, PASI). Conclusions Various inflammatory skin diseases show shared and distinct systemic protein immune activation. Overlap across conditions encourages the investigation of shared therapeutic approaches.

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, remains the deadliest human pathogen. Treatment is hampered by drug resistance and the persistence of slow-growing or non-replicating populations. Rifampicin, a cornerstone of first-line therapy, inhibits transcription during promoter escape, but resistance mutations undermine efficacy and drive resistance spread. We revisited the transcription cycle as an antibacterial target by characterizing AAP-SO2, an RNA polymerase inhibitor with whole-cell activity against Mtb. AAP-SO2 slows the nucleotide addition cycle, disrupting elongation and termination. Rifampicin-resistant mutations impose fitness costs by perturbing the balance of these steps, creating exploitable weaknesses. Inhibition of transcription with AAP-SO2 reduced the evolution of rifampicin resistance and was especially effective against the most common resistant mutant. Combination treatment with rifampicin and AAP-SO2 synergistically killed non-replicating Mtb in an ex vivo rabbit granuloma model. These findings show that exploiting functional vulnerabilities of the transcription cycle can counter rifampicin resistance and improve clearance of recalcitrant Mtb populations.

T cell-based immunotherapies have demonstrated effectiveness in treating diffuse large B cell lymphoma (DLBCL) and follicular lymphoma (FL) but predicting response and understanding resistance remains a challenge. To address this, we developed syngeneic models reflecting the genetics, epigenetics, and immunology of human FL and DLBCL. We show that EZH2 inhibitors reprogram these models to re-express T cell engagement genes and render them highly immunogenic. EZH2 inhibitors do not harm tumor-controlling T cells or CAR-T cells. Instead, they reduce regulatory T cells, promote memory chimeric antigen receptor (CAR) CD8 phenotypes, and reduce exhaustion, resulting in a decreased tumor burden. Intravital 2-photon imaging shows increased CAR-T recruitment and interaction within the tumor microenvironment, improving lymphoma cell killing. Therefore, EZH2 inhibition enhances CAR-T cell efficacy through direct effects on CAR-T cells, in addition to rendering lymphoma B cells immunogenic. This approach is currently being evaluated in two clinical trials, NCT05934838 and NCT05994235, to improve immunotherapy outcomes in B cell lymphoma patients.

Increased antibody affinity over time after vaccination, known as affinity maturation, is a prototypical feature of immune responses. Recent studies have shown that a diverse collection of B cells, producing antibodies with a wide spectrum of different affinities, is selected into the plasma cell (PC) pathway. How affinity-permissive selection enables PC affinity maturation remains unknown. We found that PC precursors (prePCs) expressing high-affinity antibodies received higher levels of T follicular helper cell (T-FH cell)-derived help and divided at higher rates compared with their lower-affinity counterparts once they left the germinal center. Our findings indicate that differential cell division by selected prePCs accounts for how diverse precursors develop into a PC compartment that mediates serological affinity maturation.

The thrombolytic protease tissue plasminogen activator (tPA) is expressed in the CNS, where it regulates diverse functions including neuronal plasticity, neuroinflammation, and blood-brain-barrier integrity. However, its role in different brain regions such as the substantia nigra (SN) is largely unexplored. In this study, we characterize tPA expression, activity, and localization in the SN using a combination of retrograde tracing and beta-galactosidase tPA reporter mice. We further investigate tPA's potential role in SN pathology in an alpha-synuclein mouse model of Parkinson's disease (PD). To characterize the mechanism of tPA action in alpha-synuclein-mediated pathology in the SN and to identify possible therapeutic pathways, we performed RNA-seq analysis of the SN and used multiple transgenic mouse models. These included tPA deficient mice and two newly developed transgenic mice, a knock-in model expressing endogenous levels of proteolytically inactive tPA (tPA Ala-KI) and a second model overexpressing proteolytically inactive tPA (tPA Ala-BAC). Our findings show that striatal GABAergic neurons send tPA+ projections to dopaminergic (DA)-neurons in the SN and that tPA is released from SN-derived synaptosomes upon stimulation. We also found that tPA levels in the SN increased following alpha-synuclein overexpression. Importantly, tPA deficiency protects DA-neurons from degeneration, prevents behavioral deficits, and reduces microglia activation and T-cell infiltration induced by alpha-synuclein overexpression. RNA-seq analysis indicates that tPA in the SN is required for the upregulation of genes involved in the innate and adaptive immune responses induced by alpha-synuclein overexpression. Overexpression of alpha-synuclein in tPA Ala-KI mice, expressing only proteolytically inactive tPA, confirms that tPA-mediated neuroinflammation and neurodegeneration is independent of its proteolytic activity. Moreover, overexpression of proteolytically inactive tPA in tPA Ala-BAC mice leads to increased neuroinflammation and neurodegeneration compared to mice expressing normal levels of tPA, suggesting a tPA dose response. Finally, treatment of mice with glunomab, a neutralizing antibody that selectively blocks tPA binding to the N-methyl-D-aspartate receptor-1 (NMDAR1) without affecting NMDAR1 ion channel function, identifies the tPA interaction with NMDAR1 as necessary for tPA-mediated neuroinflammation and neurodegeneration in response to alpha-synuclein-mediated neurotoxicity. Thus, our data identifies a novel pathway that promotes DA-neuron degeneration and suggests a potential therapeutic intervention for PD targeting the tPA-NMDAR1 interaction.