Search Results (9,876)

Background: In the 1960s and 1970s, several authors reported that manipulation of diet by excluding certain additives and natural chemical substances led to apparent clinical benefit for patients with adverse food reactions involving the skin, gastrointestinal tract, respiratory tract, and central nervous system. Methods: Over the past 48 years, patients of all ages presenting to the Royal Prince Alfred Hospital Allergy Unit with adverse food reactions were offered an elimination diet low in natural salicylates, biogenic amines, and glutamate, and free of additives such as preservatives and colors, for two to six weeks. Those who became asymptomatic were offered testing with a “N-of-1” double-blind, placebo-controlled (DBPC) challenge protocol. Results: There were 14,119 patients who elected to undertake either the strict elimination diet (10,074) (71.4%), the moderate approach (2760) (19.5%), or the simple approach (1285) (9.1%). Of these, 6645 (47.1%) became asymptomatic within two to six weeks; 628 (4.4%) reported no improvement; and 6846 (48.5%) were lost to follow-up. Of those who improved, 4963 (35.2%) returned for interpretation of their challenges, and 2429 had undertaken open food challenges; 3326 had completed the DBPC capsule challenge protocol, and 1830 completed both DBPC capsule and food challenges. Although individual reactivity was idiosyncratic, a highly reproducible pattern was evident across the entire group. The proportion reacting to each of the active chemical challenges versus placebo was highly significant (p < 0.001). Conclusions: In many patients with food intolerance, idiosyncratic dietary triggers can be reliably identified with a standardized elimination diet and DBPC challenge protocol. Long-term symptom control can be achieved by appropriate dietary modification. Full article

Erdheim–Chester disease (ECD) is a rare, clonal non-Langerhans cell histiocytic neoplasm driven by activating mutations in the mitogen-activated protein kinase (MAPK) pathway, most notably the BRAF-V600E mutation. The discovery of these molecular alterations has shifted the therapeutic paradigm from non-specific immunosuppression to personalized targeted therapies, such as BRAF and MEK inhibitors, dramatically improving patient outcomes. Given its protean and insidious systemic manifestations, imaging plays an indispensable role in the personalized diagnostic and therapeutic pathway of ECD. Metadiaphyseal symmetrical osteosclerosis represents the skeletal hallmark, while extraskeletal involvement typically affects the cardiovascular, retroperitoneal, pulmonary, and central nervous systems, yielding pathognomonic imaging signatures such as the “coated aorta” and “hairy kidney.” Advanced cross-sectional techniques—including CT, MRI, and organ-specific mapping protocols—are crucial for baseline phenotype characterization, risk stratification, and guiding biopsies toward hypermetabolic sites for biomarker detection. Furthermore, molecular imaging with ¹⁸F-FDG PET/CT has emerged as the cornerstone for baseline systemic staging and quantitative metabolic response monitoring via PERCIST criteria, enabling early assessment of targeted treatment efficacy long before morphological changes occur. This review illustrates the multimodality imaging spectrum of ECD based on our institutional experience, emphasizing how precision imaging integrates with molecular profiling to guide personalized patient management. Full article

Background/Objectives. VHHs (Nanobodies^®) are camelid single-domain antibody fragments with broad applicability in central nervous system (CNS) research, including blood–brain barrier transport, modulation of pathological proteins and molecular detection. However, few VHHs have been developed for cell-type-specific targeting within the brain. Astrocytes play essential roles in CNS homeostasis and contribute to many neurological disorders, making them an appealing target for diagnostic and therapeutic approaches. Here, we report the development and validation of the first VHHs directed against the astrocyte-enriched glutamate transporters GLT-1 and GLAST-1. Methods. VHHs were generated from a camelid immune library using phage display. Target specificity and affinity were evaluated in cells overexpressing the mouse and human transporters, as well as in iPSC-derived astrocytes. Target engagement and astrocyte specificity was further assessed ex vivo on brain sections using immunohistochemistry and in dissociated mouse brain cell suspensions by flow cytometry. VHH in vivo binding was evaluated by stereotactic injections within the brain striatum. Results. One anti-GLT-1 VHH and four anti-GLAST-1 VHHs demonstrated high specificity and apparent nanomolar affinity when binding tocells overexpressing the mouse transporters. The anti-GLT-1 VHH was additionally cross-reactive with the human ortholog, as demonstrated in overexpressing cells and iPSC-derived astrocytes. All VHHs showed target engagement on mouse brain sections and astrocyte specificity in dissociated mouse brain cell suspensions. VHH direct brain delivery resulted in localized transporter-specific signal around the injection site, whereas the non-binding isotype control VHH produced no detectable signal. Conclusions. Here we present the first set of VHHs capable of selectively recognizing astrocyte-enriched glutamate transporters GLT-1 and GLAST-1 in vitro, ex vivo, and in vivo. These VHHs constitute promising tools for astrocyte detection in research applications and the cross-reactive anti-GLT-1 lead might hold potential for future diagnostic or therapeutic strategies, including cell-targeted payload delivery. Full article

Development of therapies and pharmacological management of central nervous system diseases is significantly hampered by the presence of the highly selective blood–brain barrier (BBB). Receptor-mediated transcytosis (RMT) has proven to be a robust mechanism for delivering a wide variety of therapeutic modalities across the BBB. The present study describes the use of a neurotensin-based pharmacodynamic model to evaluate the brain penetrating capacity of FC5, a well-characterized RMT shuttle. Here we produce fusion molecules of FC5 with human Fc and neurotensin and show that intact but not C-terminally clipped neurotensin proteins containing either FC5 (FC5Fc-neurotensin) or a non-BBB-crossing control (A20.1Fc-neurotensin) retain the ability to activate neurotensin receptor type 1. In in vivo studies, intravenous administration of FC5Fc-neurotensin, but not neurotensin alone or A20.1Fc-neurotensin, induced a drop in the core body temperature in both rats and mice. We further extended these findings to non-human primates, demonstrating the ability of FC5 to cross the BBB in this species. Collectively, our results further support the potential of FC5 in delivering payloads across the BBB. The neurotensin model of hypothermia is also validated as a pharmacodynamic approach for assessing brain delivery, albeit with some necessary considerations into the molecular characterization of fusion constructs before in vivo evaluation. Full article

Background/Objectives: Listeria monocytogenes is an uncommon but potentially life-threatening cause of central nervous system infection, predominantly affecting older adults and immunocompromised patients. Neurolisteriosis may present with nonspecific neurological manifestations, and the absence of classical meningeal signs may delay diagnosis. We report a case of L. monocytogenes meningoencephalitis in an elderly patient without known immunosuppression, with rapid cerebrospinal fluid (CSF) molecular testing facilitating early diagnosis and appropriate antimicrobial treatment. Case presentation: A 76-year-old man with significant cardiovascular comorbidities was admitted with a 3-day history of fever, headache, nausea, vomiting, fatigue, and gait instability. He was alert on admission, with only mild meningeal signs, and initial cranial computed tomography showed no acute intracranial abnormalities. CSF analysis revealed inflammatory changes. Empirical ceftriaxone therapy was supplemented with vancomycin, followed by the initiation of intravenous ampicillin and gentamicin after molecular identification of L. monocytogenes. The diagnosis was subsequently confirmed by CSF culture, while blood cultures remained negative. Clinical and laboratory parameters improved progressively, accompanied by marked improvement in CSF findings on follow-up examination. The patient was discharged in stable condition after 18 days of hospitalization. Conclusions: Neurolisteriosis should be considered in elderly patients presenting with fever and neurological symptoms, even in the absence of overt immunosuppression or prominent meningeal signs. This case highlights the diagnostic value of rapid CSF molecular testing, particularly when blood cultures are negative and the initial clinical presentation is nonspecific. Early recognition of L. monocytogenes infection and timely initiation of targeted antimicrobial therapy are essential to improving outcomes. Full article

Somatostatin receptor (SSTR)–targeted imaging and therapy are established approaches for neuroendocrine tumors, but accumulating evidence indicates that SSTRs are also expressed in various non-neuroendocrine tumors. This review aimed to summarize their expression patterns, imaging characteristics, and theranostic potential. Published evidence was reviewed for central nervous system tumors, head and neck cancers, thyroid carcinoma, lung cancer, breast cancer, lymphoma, and melanoma, with particular attention to the cellular localization of SSTR expression and the mechanisms underlying radiotracer uptake. Meningioma, non-keratinizing nasopharyngeal carcinoma, and selected estrogen receptor–positive breast cancers show relatively prominent tumor-cell SSTR expression. In several other malignancies, SSTR expression may predominantly involve tumor-associated macrophages, endothelial cells, fibroblasts, or other stromal components. Imaging findings are heterogeneous across and within tumor types. Preliminary studies suggest that peptide receptor radionuclide therapy may provide clinical benefit in selected patients with SSTR-positive non-neuroendocrine tumors, although the available evidence remains limited and largely non-prospective. Further studies integrating quantitative molecular imaging, spatial pathology, and molecular profiling are needed to clarify receptor biology, establish patient-selection criteria, and define the clinical role of SSTR-targeted theranostics beyond neuroendocrine tumors. Full article

Background: Neuropathic pain (NP) is a chronic pain condition caused by a lesion or disease of the somatosensory nervous system. Its development and persistence are closely associated with peripheral nociceptive input, central sensitization, neuroimmune dysregulation, and remodeling of the local metabolic microenvironment. Our previous studies demonstrated that combined treatment with ligustrazine and sinomenine ameliorates early sciatic nerve pathological damage and systemic metabolic disturbances in rats with chronic constriction injury (CCI). However, whether this combination modulates central sensitization in NP remains unclear. Objective: This study aimed to evaluate the effects of combined ligustrazine and sinomenine treatment on pain-related behaviors and motor function during the maintenance phase of CCI-induced neuropathic pain and to investigate the potential mechanisms underlying its attenuation of central sensitization, with particular emphasis on metabolic remodeling, neurotransmitter homeostasis, and associated regulatory molecules in the spinal dorsal horn. Methods: A rat model of CCI was established, and animals were treated with ligustrazine combined with sinomenine (12.5 + 12.5, 25 + 25, or 50 + 50 mg/kg) for 7 consecutive days. Mechanical withdrawal threshold, cold hypersensitivity, and gait behavior were assessed. A multiplex assay was used to profile 12 plasma inflammatory cytokines and pain-related mediators. Integrated transcriptomic and metabolomic analyses of the spinal dorsal horn were subsequently performed to identify key altered pathways. Oxylipin profiling, targeted neurotransmitter analysis, and quantification of metabolites associated with the arginine–polyamine pathway were further conducted to characterize alterations in the local metabolic microenvironment associated with central sensitization. Finally, quantitative real-time PCR (qRT-PCR) and Western blotting were used to evaluate molecules related to polyamine metabolism, redox homeostasis, and central sensitization. Results: Combined ligustrazine and sinomenine treatment improved pain-related behaviors and gait dysfunction in CCI rats and partially normalized systemic inflammatory dysregulation. The ipsilateral-to-contralateral maximum contact-area ratio decreased from 102.8 ± 9.0% in the Sham group to 53.1 ± 9.7% in the Model group and recovered to 85.5 ± 4.4% in the high-dose group. Metabolomic and transcriptomic analyses identified 54 differential metabolites and 130 differentially expressed genes, respectively, following treatment. Integrated multi-omics analysis highlighted arginine-related metabolic pathways, together with alterations in neurotransmitter, polyamine, oxylipin, and redox homeostasis. Full article

Neutrophils were long regarded as a uniform population of short-lived phagocytes that execute a stereotyped antimicrobial program before dying at the site of infection. Single-cell transcriptomics, mass cytometry, and spatial imaging have replaced this view with a picture of neutrophils as a developmental continuum, beginning with committed bone marrow precursors and extending through circulating, aged, suppressive, and tissue-imprinted states, each carrying a distinct transcriptional and functional signature. This review traces how granulopoiesis generates that continuum, how chemokine axes such as CXCR4-CXCL12 and CXCR2 govern bone marrow release and peripheral trafficking, and how infection reshapes the balance between protective and pathogenic neutrophil programs, including neutrophil extracellular trap formation, immunometabolic reprogramming, and tissue-specific adaptation in lung, gut, liver, and central nervous system compartments. Particular attention is given to the eye, where neutrophils confer early antiviral protection against herpes simplex virus type 1 keratitis yet also drive corneal neovascularization and nerve damage that contribute to vision loss, illustrating how a single tissue can showcase both faces of neutrophil biology. We also examine the comparatively underappreciated role of neutrophils and neutrophil extracellular traps in helminth and protozoan infection, where they can both restrict parasite burden and drive collateral tissue injury. We conclude by evaluating strategies to modulate specific neutrophil states, rather than neutrophils as a whole, as an emerging therapeutic direction across bacterial, viral, fungal, and parasitic diseases, and by identifying gaps that single-cell and spatial approaches are positioned to close. Full article

Recent advances in fetal neurosonography have markedly improved the detection of subtle central nervous system findings that were previously identified less frequently. Mild ventriculomegaly (MV), non-dilated ventricular asymmetry (NDVA), mega cisterna magna (MCM), and Blake’s pouch cyst (BPC) are increasingly encountered during routine prenatal ultrasound and, in the absence of associated genetic or infectious abnormalities, are commonly regarded as borderline conditions or anatomical variants. These findings frequently pose challenges in prognostic evaluation and in the counseling of expectant parents. The objective of this review is to summarize current evidence on the detection, characterization, and clinical management of these four entities and to provide a practical framework for evidence-based parental counseling. This narrative review examines the diagnostic approach to MV, NDVA, MCM, and BPC using both ultrasound and fetal magnetic resonance imaging (MRI). A uniform analytic framework is applied to each entity, addressing in turn etiology, sonographic assessment, the complementary role of MRI, genetic evaluation, and prognosis with counseling considerations. Representative ultrasound and fetal MRI images from a tertiary referral center are used for illustrative purposes. Across all four entities, fetal MRI provides complementary information in selected cases, while genetic testing and serological screening may help exclude chromosomal or infectious etiologies. Neurodevelopmental outcomes are broadly favorable when the finding is truly isolated, but the prognostic literature is limited by small samples, heterogeneous definitions of “isolated,” short follow-up, and unstandardized neurodevelopmental assessments. Common themes across these entities are synthesized into a staged framework intended to support evidence-based parental counseling regarding the expected neurodevelopmental outcomes of these fetuses. Full article

Background/Objectives: Leptomeningeal disease (LMD) is among the most severe complications of metastatic breast cancer, with median survival measured in weeks. Definitive diagnosis relies on neuroimaging and cerebrospinal fluid (CSF) cytology, which carries a false-negative rate of 40–50% at a single lumbar puncture. Improved tools for detecting and characterizing tumor cells across systemic and central nervous system (CNS) compartments are urgently needed. Methods: We applied an enrichment-free, fluorescence whole-slide imaging (fWSI)-based liquid biopsy platform to ante-mortem and post-mortem peripheral blood (PB) and CSF from five patients with metastatic breast cancer and confirmed or suspected LMD, collected through the City of Hope LEGACY warm autopsy program. Circulating tumor cells (CTCs) were classified by immunophenotype (cytokeratin [CK], vimentin [VIM], CD45/CD31) and morphology. Select cells underwent single-cell copy number variation (CNV) profiling by low-pass whole-genome sequencing. Results: Across 23 samples (11 PB, 12 CSF), we identified marked compartmental heterogeneity in CTC burden, phenotype, and genomic architecture. In the one patient (Patient 1) sampled across all four ventricular-to-cisternal CSF sites, CTC frequency declined from proximal to distal compartments (11.4% in the right lateral ventricle to 0.6% at the foramen magnum), a single-patient observation consistent with a proximal-to-distal gradient. Patients with active, untreated LMD showed high CSF CTC incidence with both epithelial and mesenchymal phenotypes, whereas the two patients receiving intrathecal therapy each showed low or absent CSF CTCs; given the small numbers involved and a plausible technical confound in a third patient, we interpret this association cautiously. Single-cell CNV profiling of CSF-derived CTCs across patients with active LMD revealed highly concordant chromosomal aberration profiles, 100% of CK+ CSF CTCs analyzed carried chromosomal aberrations, consistent with, though not formally establishing, expansion of a genomically evolved subclone within the leptomeningeal compartment; PB-derived cells showed substantially greater heterogeneity. In the two patients with paired post-mortem arterial and venous PB, arterial blood showed higher total rare cell counts in both, and higher CK-positive CTC incidence in one (1425.73 vs. undetectable epi.CTC/mL); the other showed the reverse for this specific metric. Conclusions: Enrichment-free, multi-compartment liquid biopsy reveals a regionally heterogeneous and biologically structured landscape in breast cancer LMD not resolvable by conventional cytology or single-site venous sampling. These findings are consistent with a hypothesis-generating model in which the leptomeningeal space acts as a selective niche for clonally convergent tumor expansion. They also point to the potential of regionally resolved CTC profiling for monitoring LMD treatment response in future, larger cohorts. Full article

Background/Objectives: Neurological manifestations in rheumatoid arthritis (RA) are common, yet the clinical relevance of central nervous system (CNS) abnormalities detected on neuroimaging remains uncertain. We aimed to determine the prevalence and patterns of cranial magnetic resonance imaging (MRI) findings in RA patients treated with rituximab (RTX) and to assess whether these abnormalities are associated with disease-specific mechanisms or with age- and comorbidity-related factors. Methods: In this retrospective observational study, 84 RA patients who received RTX and had available pre-treatment cranial MRI underwent systematic evaluation. Imaging findings were classified by an experienced neuroradiologist blinded to clinical data and dichotomized as normal or abnormal, and white matter hyperintensities were graded semiquantitatively using a Fazekas-type scale. Factors independently associated with abnormal MRI were assessed by multivariable logistic regression. Results: Cranial MRI abnormalities were identified in 60 patients (71.5%), predominantly as nonspecific white matter hyperintensities (48 patients, 57.1%). In multivariable analysis, older age (adjusted odds ratio [aOR] 1.16 per year, 95% CI 1.08–1.25) and the presence of comorbidities (aOR 5.36, 95% CI 1.49–19.23) were independently associated with abnormal imaging (p < 0.001 and p = 0.010, respectively), whereas no association was found with disease duration, seropositivity, or extra-articular involvement. Imaging findings remained stable over time in the majority of the 39 patients (46.4%) who underwent follow-up imaging. Conclusions: Cranial MRI abnormalities are highly prevalent in RA and were associated primarily with age and comorbidity burden rather than with RA-specific inflammation; however, in the absence of a control group, we cannot determine whether their prevalence exceeds that of a comparable non-RA population. No treatment-related demyelinating disease or progressive multifocal leukoencephalopathy was observed, but the study was not designed to assess drug safety, and these findings should be interpreted as the absence of an observed imaging signal rather than as evidence of the neurological safety of rituximab. Cranial neuroimaging in RA should therefore be interpreted cautiously and in clinical context. Full article

The glymphatic system is a brain-wide perivascular network that facilitates cerebrospinal-fluid–interstitial-fluid exchange and the clearance of metabolic waste. Growing experimental and clinical evidence suggests that glymphatic dysfunction may represent a convergent pathophysiological feature associated with aging, neuroinflammation, and neurodegeneration, although its causal position within these processes remains unresolved. In this narrative review, we synthesize human imaging, clinical, and translational evidence implicating glymphatic dysfunction across major neurodegenerative diseases and integrate these findings with emerging data supporting its role in chronic low-grade inflammation associated with aging (“inflammaging”). We further discuss how alterations in the gut microenvironment may remotely influence glymphatic function and contribute to neurodegeneration through the gut–brain axis. Although supported by extensive preclinical evidence, studies in humans increasingly demonstrate impaired glymphatic function in several neurodegenerative disorders, most assessed using diffusion tensor imaging along the perivascular space (DTI-ALPS) index. Glymphatic dysfunction is associated with cognitive decline, motor impairment, and disease progression. Aging-related alterations in astrocytic function, aquaporin-4 polarization, blood–brain barrier integrity, and perivascular fluid dynamics provide mechanistic links between inflammaging and glymphatic failure. Gut dysbiosis may further exacerbate these processes by promoting central nervous system inflammation and vascular dysfunction. Together, these findings identify the glymphatic system as a clinically relevant pathway in neurodegeneration that may be regulated by aging-associated neuroinflammatory mechanisms involving the gut–brain axis. Finally, we discuss physiological, behavioral, and pharmacologic modulators that influence glymphatic function and propose a unified framework positioning glymphatic dysfunction as a central integrator of impaired brain clearance, with potential implications for biomarker development, clinical assessment, and disease-modifying therapeutic strategies in neurodegenerative disorders. Full article

Background/Objectives: Blood–brain barrier (BBB) permeability prediction is important for central nervous system drug discovery. Many models rely mainly on chemical structure, omit transport information, and may report optimistic performance when related compounds occur across training and test sets without adequate leakage control. We developed MechBBB, a two-stage, transport-informed machine-learning framework that adds learned efflux, influx, and passive-permeability scores to conventional molecular features and evaluates their added value under explicit leakage control. Methods: In Stage 1, three LightGBM models were trained on separate efflux, influx, and passive-permeability datasets to generate transport-related scores. BBBP compounds were excluded from Stage 1 training by InChIKey matching. In Stage 2, a LightGBM classifier was trained on BBBP using a Murcko scaffold split with the three Stage 1 scores, ten physicochemical descriptors, and 2048-bit ECFP4 fingerprints. Performance was compared with a descriptor-plus-fingerprint baseline without transport scores. External evaluation used a strict B3DB set (n = 4080) with no InChIKey or nonempty scaffold overlap with BBBP and was performed without retraining, recalibration, or threshold adjustment. MechBBB was also compared with official SwissADME BOILED-Egg predictions on a chemically matched external subset. Results: On the BBBP scaffold test set, MechBBB achieved an AUROC of 0.932, an AUPRC of 0.983, an MCC of 0.737, and a balanced accuracy of 0.866. The AUROC improvement over the descriptor-plus-fingerprint baseline was small but statistically significant in the paired test (ΔAUROC = 0.0096; p = 0.0425). Scaffold-grouped cross-validation did not show a consistent ranking advantage from adding the Stage 1 scores. On strict B3DB, MechBBB achieved an AUROC of 0.894 and an AUPRC of 0.893, with no significant ranking advantage over the baseline. On the matched B3DB subset (n = 4043), MechBBB achieved higher accuracy (0.814 vs. 0.682), balanced accuracy (0.806 vs. 0.693), sensitivity (0.910 vs. 0.541), and MCC (0.631 vs. 0.401) than SwissADME BOILED-Egg, whereas BOILED-Egg had higher specificity (0.845 vs. 0.703). SHAP analysis identified TPSA, NumHDonors, and p_pampa among the leading contributors. Conclusions: MechBBB provides a leakage controlled and externally tested framework that combines competitive BBB permeability prediction with per-compound transport-related information. The Stage 1 scores produced a modest internal improvement but did not provide a consistent external ranking advantage, indicating that their main added value is transport-related context rather than a universal increase in predictive accuracy. MechBBB can be used to prioritize compounds for CNS drug discovery before experimental BBB testing and identify compounds that may warrant follow-up studies of passive permeability or active transport. The MechBBB web tool is freely available. Full article

Toxoplasma gondii persists in the central nervous system and may cause severe toxoplasmic encephalitis, particularly in immunocompromised hosts. Our previous studies demonstrated that T. gondii-induced neuropathology involves oxidative and nitrosative stress, apoptosis, glial activation, blood–brain barrier injury, and GMF-β- and SAP/NF-κB-associated responses. This study investigated the temporal immunoreactivity of CD68, S100B, Foxp3, and IL-5 during early and chronic experimental infection. Twenty-four female Swiss albino mice were assigned to one uninfected control group and three ME49-infected groups examined at 10, 30, and 60 days post-infection (n = 6/group). Marker immunoreactivity was evaluated by immunohistochemistry and quantitative image analysis. CD68 immunoreactivity was markedly increased at day 10, rose further by day 30, and remained high at day 60. In contrast, immunoreactivity for S100B, Foxp3, and IL-5 was limited at day 10 but increased markedly at days 30 and 60. Most notably, IL-5 continued to increase significantly from day 30 to day 60, whereas CD68, S100B, and Foxp3 remained at similarly high levels during this interval. These temporally divergent patterns demonstrate an early myeloid/phagolysosomal response followed by established astroglial, Foxp3-associated regulatory, and cytokine-associated tissue responses. The continued increase in IL-5, together with persistent neuropathological changes, supports the interpretation that late chronic toxoplasmic encephalitis is an active rather than quiescent neuroimmunopathological phase. Full article

HIV-associated neurocognitive disorders (HAND) can persist despite effective ART and are associated with persistent central nervous system (CNS) viral reservoirs in microglia, chronic neuroinflammation, synaptic dysfunction, and neuronal injury. Thus, human-relevant multicellular CNS models are needed to elucidate neuroHIV pathogenesis and ART neurotoxicity. Here, we establish and characterize a fully human induced pluripotent stem cell (hiPSC)-derived CNS tri-culture model comprising isogenic neurons, astrocytes, and microglia that remains viable and functionally active for up to 32 days in vitro. Using a microglia-tropic fluorescent HIV reporter virus, GFP-HIV-AD8, we demonstrate infection of IBA1+ microglia with no detectable infection of IBA1− cells or TUBB3+ neurons. HIV replication was quantified over 14 days post-infection and effectively suppressed by the first-generation integrase strand transfer inhibitor (INSTI)-containing regimen elvitegravir/tenofovir disoproxil fumarate/emtricitabine (EVG/TDF/FTC), and the second-generation INSTI-containing regimens dolutegravir/tenofovir disoproxil fumarate/emtricitabine (DTG/TDF/FTC) and bictegravir/tenofovir alafenamide/emtricitabine (BIC/TAF/FTC), at concentrations based on reported plasma Cmax values. HIV infection reduced microglial viability by approximately 70%, partially rescued by lower in vitro ART concentrations but worsened by the supratherapeutic 3× Cmax condition of DTG-containing regimens. In uninfected cultures, DTG induced microglial morphological changes consistent with a reactive or dysfunctional state. Calcium imaging revealed ART-specific neuronal effects: EVG/TDF/FTC reduced neuronal activity, whereas BIC/TAF/FTC enhanced it. In HIV-infected cultures, DTG shortened neuronal spike duration under viral suppression, identifying an HIV–DTG-associated electrophysiological phenotype. Overall, BIC/TAF/FTC showed the most favorable profile across the cellular and electrophysiological endpoints assessed in this donor line. Together, these findings establish an hiPSC-derived CNS tri-culture model for controlled studies of neuroHIV virology, ART effects, and neuronal function. Full article