Faculty Bibliography

Animals need daily intakes of three macronutrients: sugar, protein, and fat. Under fasted conditions, however, animals prioritize sugar as a primary source of energy. They must detect ingested sugar-specifically D-glucose-and quickly report its presence to the brain. Hypothalamic neurons that can respond to the caloric content in the gut regardless of the identity of macronutrient have been identified, but until now, the existence of neurons that can encode the specific macronutrients remained unknown. We found that a subset of corticotropin-releasing factor (CRF)-expressing neurons in the hypothalamic paraventricular nucleus (CRF^PVN) respond specifically to D-glucose in the gut, separately from other macronutrients or sugars. CRF^PVN neuronal activity is essential for fasted mice to develop a preference for D-glucose. These responses of CRF^PVN neurons to intestinal D-glucose require a specific spinal gut-brain pathway including the dorsal lateral parabrachial nuclei. These findings reveal the neural circuit that encodes the identity of D-glucose.

OBJECTIVE:This study examined racial and ethnic differences in percent total weight loss (%TWL) and glycemic improvement following sleeve gastrectomy (SG) and explored the role of socioeconomic and psychosocial factors in postsurgical outcomes. METHODS:This longitudinal study included patients who underwent SG between 2017 and 2020, with follow-up visits over 24 months. RESULTS:Non-Hispanic Black (NHB) participants had lower %TWL at 3, 12, and 24 months compared with Hispanic (H) and non-Hispanic White (NHW) participants. Fat mass index was initially lower in NHB, with smaller reductions over time and significant group differences persisting at 24 months. NHB participants had higher baseline fat-free mass index values; by 24 months, fat-free mass index values were lower in H participants. Hemoglobin A1c decreased across all groups but remained consistently higher in NHB and H compared with NHW at 24 months. NHB participants reported higher perceived discrimination, sleep disturbance, and perceived stress than H and NHW participants at all time points. Employment status predicted %TWL at 12 months. There was a significant interaction between race and ethnicity and employment status observed at 12 and 24 months, suggesting that employment-related disparities could impact surgical outcomes. CONCLUSIONS:NHB participants experienced less favorable outcomes following SG, emphasizing the need for tailored interventions addressing socioeconomic and psychosocial disparities.

Endosomal system dysfunction within neurons is a prominent early feature of Alzheimer's disease (AD) pathology. Multiple AD risk factors are regulators of endocytosis and are known to cause hyper-activity of the early-endosome small GTPase rab5, resulting in neuronal endosomal pathway disruption and cholinergic neurodegeneration. Adaptor protein containing Pleckstrin homology domain, Phosphotyrosine binding domain, Leucine zipper motif (APPL1), an important rab5 effector protein and signaling molecule, has been shown in vitro to interface between endosomal and neuronal dysfunction through a rab5-activating interaction with the BACE1-generated C-terminal fragment of amyloid precursor protein (APP-βCTF), a pathogenic APP fragment generated within endosomal compartments. To understand the contribution of APPL1 to AD-related endosomal dysfunction in vivo, we generated a transgenic mouse model over-expressing human APPL1 within neurons (Thy1-APPL1). Strongly supporting the important endosomal regulatory roles of APPL1 and their relevance to AD etiology, Thy1-APPL1 mice (both sexes) develop enlarged neuronal early endosomes and increased synaptic endocytosis due to increased rab5 activation. We demonstrated pathophysiological consequences of APPL1 overexpression, including functional changes in hippocampal long-term potentiation (LTP) and long-term depression (LTD), degeneration of large projection cholinergic neurons of the basal forebrain, and impaired hippocampal-dependent memory. Our evidence shows that neuronal APPL1 elevation modeling its functional increase in the AD brain induces a cascade of AD-related pathological effects within neurons, including early endosome anomalies, synaptic dysfunction, and selective neurodegeneration. Our in vivo model highlights the contributions of APPL1 to the pathobiology and neuronal consequences of early endosomal pathway disruption and its potential value as a therapeutic target.Significance Statement Neuronal endosome dysfunction appears early in Alzheimer's disease (AD) and is linked to memory loss. Genes and risk factors associated with AD often increase rab5 activity, a protein that disrupts endosomal signalling when hyperactivated. APPL1, a key rab5 partner, worsens this dysfunction via its interaction with APP-βCTF, a protein fragment associated with AD. To explore APPL1's role, we created a genetically modified mouse that overexpresses APPL1 in neurons. This model provides the first in vivo evidence that APPL1 overexpression triggers key AD-like effects: rab5 hyperactivation, enlarged early endosomes, loss of cholinergic neurons, reduced synaptic plasticity in memory-related brain regions, and memory deficits. These findings highlight APPL1's role in AD pathogenesis and its potential as a therapeutic target.

Allergic contact dermatitis (ACD) is a delayed-type IV hypersensitivity response driven by innate and adaptive immune cells. While specific immune regulations of these cell types are amply elucidated, their regulations by extracellular matrix (ECM) components and T cell mediated adaptive immunity in ACD remains unclear. Lumican and biglycan are ECM proteoglycans abundant in the dermis and lymph node, known to regulate innate immune myeloid cells, but have not been investigated in lymphoid cell regulations in ACD. By immunohistology we localized lumican and biglycan in skin biopsies of psoriatic patients. Using wild type (WT), lumican and biglycan knockout mice, we investigated CD4⁺T cell infiltration, activation and proliferation in the skin and draining lymph node (dLN) of CHS-challenged mice by immunohistochemistry and flow cytometry. We used the OT-II adoptive transfer model to test antigen specific CD4⁺T cell activation. We assessed interactions of the proteoglycans with LFA-1 on T cells by confocal microscopy. Compared to WTs, the knockouts showed severe ear inflammation, with increased CD4⁺T cells infiltration in the dermis. CHS-challenged knockout mice dLN showed increased T-bet, STAT1 and -STAT4 signaling, indicating enhanced Th1 commitment and proliferation. We found that WT lymph node fibroblastic reticular cells (FRCs) secrete lumican, biglycan and decorin, a related proteoglycan, while none are expressed by naive or activated T cells. Lumican and biglycan interact with LFA-1 on T cell surfaces, and in vitro all three proteoglycans suppress CD4⁺T cell activation. Secreted by dLN FRCs, lumican, biglycan, and possibly decorin interact with LFA-1 on CD4⁺T cells to restrict its activation and reduce dermatitis severity.

Targeted therapies and immunotherapy have improved treatment outcomes for many melanoma patients. However, patients whose melanomas harbor driver mutations in the neurofibromin 1 (NF1) tumor suppressor gene often lack effective targeted treatment options when their tumors do not respond to immunotherapy. In this study, we utilized patient-derived short-term cultures (STCs) and multiomics approaches to identify molecular features that could inform the development of therapies for patients with NF1 mutant melanoma. Differential gene expression analysis revealed that the epidermal growth factor receptor (EGFR) is highly expressed and active in NF1 mutant melanoma cells, where it hyper-activates ERK and AKT, leading to increased tumor cell proliferation, survival, and growth. In contrast, genetic or pharmacological inhibition of EGFR hindered cell proliferation and survival and suppressed tumor growth in patient-derived NF1 mutant melanoma models but not in NF1 wild-type models. These results reveal a connection between NF1 loss and increased EGFR expression that is critical for the survival and growth of NF1 mutant melanoma cells in patient-derived culture and xenograft models, irrespective of their BRAF and NRAS mutation status.

The intestinal secretory lineage is thought to comprise four distinct cell types derived from one Atoh1⁺ progenitor, but the mechanisms that distinguish Paneth and goblet cells are unclear. Bhattacharya et al.¹ argue that these cells are instead phenotypic manifestations of a common terminal Atoh1⁺ cell, actively shaped by niche-derived signals.

BACKGROUND:/calmodulin-dependent protein kinase II and selected for probe collection. RESULTS:This approach significantly reduced the amount of FFPE tissue needed for robust single population RNA-seq. We demonstrate ~20 identified L3 or L5 pyramidal neurons or one lamina-specific cortical ribbon from a single 5µm thick section is sufficient to generate robust RNA-seq reads. Bioinformatic analysis of neurons and ribbons showed notable similarities and differences reflective of the single neuron and multiple admixed cell types within the former and latter, respectively. Comparison with existing methods Protocols exist for DSP of postmortem human FFPE brain tissue. However, this new approach enables profiling small groups of ~14-21 pyramidal neurons using the GeoMx DSP platform. CONCLUSIONS:This optimized DSP assay provides high resolution RNA-seq data demonstrating utility and versatility of the GeoMx platform for individually characterized neurons and isolated cortical ribbons within postmortem FFPE human brain tissue for downstream analyses.

Alzheimer's disease (AD) is characterized by the deposition of full-length and truncated amyloid beta (Aβ) species within brain parenchyma and cerebral vessels. However, Aβ truncated species remain understudied. Here, we present a protocol for culture and characterization of a mouse monoclonal antibody targeting N-terminally truncated proteoforms starting at position 4. We describe a detailed procedure for hybridoma culture, antibody collection, and isolation via affinity chromatography. We then describe steps for target validation via dot blot, as well as potential applications. For complete details on the use and execution of this protocol, please refer to Cabrera et al. and Rostagno et al.¹

BACKGROUND:Synovial fluid (SF) biomarkers demonstrate time-dependent variation after acute knee injury, and it is postulated that persistently elevated inflammatory markers may mediate worse long-term outcomes. PURPOSE/OBJECTIVE:This study investigated the relationship between biomarkers in SF at the time of meniscectomy and long-term patient-reported outcomes in patients with acute versus chronic meniscal injuries. STUDY DESIGN/METHODS:Cohort study; Level of evidence, 3. METHODS:This retrospective analysis included patients who underwent knee SF aspiration on the day of arthroscopic meniscectomy between October 2011 and October 2020 with minimum 4-year follow-up. SF aspirated from the operative knee was analyzed for 10 pro- and anti-inflammatory biomarkers. Patients completed the visual analog scale for pain, Lysholm Knee Questionnaire, Tegner Activity Scale, and Knee injury and Osteoarthritis Outcome Score-Physical Function Short-form (KOOS-PS) before surgery and at follow-up. Patients were categorized as having acute (<6 weeks) or chronic (>1 year) symptoms. K-means clustering analysis was performed using biomarker levels to group patients into distinct cohorts. RESULTS:= .020) than the low-inflammation cohort. CONCLUSION/CONCLUSIONS:In patients with chronic meniscal injury, those with a more proinflammatory SF biomarker profile at the time of meniscectomy had worse outcomes than those who had a low inflammatory profile. In acute meniscal injuries, most patients demonstrate a high inflammatory profile, which was not associated with a difference in long-term outcomes.