Citations

Background: Tendon-bone healing after rotator cuff injury is, or remains, a clinical challenge. Purpose: To examine the role of intraoperative use of purified exosome products (PEPs) in the treatment of rotator cuff injuries and to assess the effect of PEP on supraspinatus tendon-bone healing after rotator cuff repair in a rat model. Study Design: Controlled laboratory study. Methods: This study used 120 Sprague-Dawley rats that underwent an acute supraspinatus tendon transection from the insertion, followed by repair using a single transosseous suture. They were divided into 3 groups: a control group that received only sutures, a TISSEEL group that used a commercially available fibrin sealant patch (TISSEEL) of approximately 0.5 cm 2 , and a PEP-TISSEEL group that used a similarly sized TISSEEL patch combined with PEP. Autopsies were performed at 1, 3, 6, and 8 weeks after surgery. Analyses were performed using molecular biological, biomechanical, and histological techniques. Results: At 6 weeks, the PEP-TISSEEL group achieved greater failure load (mean, 27.5 ± 5.3 N) than the control (mean, 16.4 ± 4.2 N; P = .012) and TISSEEL (mean, 23.4 ± 5.0 N; P = .041) groups, with superiority persisting at 12 weeks (27.2 ± 6.5 N vs 13.1 ± 2.6 N [ P = .002] for the control group and 21.1 ± 5.8 N [ P = .037] for the TISSEEL group). DigiGait showed earlier recovery of stride length and paw angle. Histology revealed organized collagen and reduced scarring, while molecular assays demonstrated early inflammatory modulation with subsequent upregulation of collagen type 1 or 3 and TGF-β, supporting accelerated tendon-bone healing. Conclusion: Intraoperative application of PEP-TISSEEL enhanced tendon-bone healing in a rat model by improving biomechanical strength, promoting enthesis-specific histological regeneration, and accelerating functional recovery as evidenced by gait analysis. Clinical Relevance: The intraoperative application of PEP combined with TISSEEL significantly enhances rotator cuff tendon healing by modulating early inflammation, promoting collagen synthesis, and improving biomechanical properties in a rat model. These findings suggest that PEP-based therapies could offer a promising adjunct in surgical repair, potentially reducing retear rates and improving functional outcomes in patients with rotator cuff injuries. Further clinical studies are warranted to translate these benefits into human applications.

Abstract Microglia are essential for maintaining brain homeostasis, including the sleep-wake cycle, but the underlying mechanisms remain unclear. Here, using in vivo two-photon imaging in both head-fixed and freely moving mice with simultaneous electroencephalogram–electromyogram (EEG–EMG) recording, we investigated microglial process dynamics and their interactions with locus coeruleus (LC) noradrenergic (NE) axons in the frontal cortex across sleep-wake states. During sleep or chemogenetic suppression of LC neurons, microglia enhanced their surveillance and increased interactions with NE axons. Correlative electron microscopy at nanometer resolution identified structural contacts between microglial process endings and NE boutons. Mechanistically, microglia-bouton interaction is primed by NE–β2-adrenergic receptor (β2AR) signaling, leading to enhanced Ca²⁺ activity in NE axons. These findings suggest that microglial interactions with NE axons promote cortical norepinephrine transmission and maintain stable wakefulness.

Abstract Background More than 85% of Alzheimer's disease (AD) donor brains exhibit some degree of cerebral amyloid angiopathy (CAA), which is characterized predominantly by Aβ40 deposits in the brain vasculature and can cause blood brain barrier (BBB) leakage. CAA is associated with risk of infarcts, cerebral hemorrhages, and cognitive decline. Brain cell type‐specific mechanisms that influence CAA pathology, as well as related measures such as Aβ40 biochemical measures and tight junction proteins, an indicator of BBB integrity, are still elusive. Method We performed single‐nucleus RNA sequencing (snRNAseq) of temporal cortex tissue from 79 AD donors, with varying levels of CAA co‐pathology. Measurements of soluble, insoluble and membrane‐associated fractions of brain Aβ40 and tight junction protein (claudin‐5, occludin) levels were available for these donors. We correlated these measures with cell proportions. We also conducted differential gene expression and analyses for intercellular communications and regulatory networks. Result We identified 25 clusters and annotated them with major brain cell types. There were 3 oligodendrocyte, 7 excitatory and 7 inhibitory neuronal, 2 microglial, 2 astrocytic, and 1 each for endothelial, pericytic, fibroblast, and oligodendrocyte precursor cell clusters. We found that higher CAA and Aβ40 levels correlate with reduced neuronal and increased microglial, astrocytic and vascular cell proportions. Increased tight junction proteins correlate with increased inhibitory neuronal and reduced microglial, astrocytic and vascular cell proportions. Among all clusters, one microglial, both astrocytic and the pericytic clusters have greater number of genes significantly associated with CAA, Aβ40 and tight junction protein levels. Most significant genes have negative associations with CAA and membrane‐bound Aβ40 and positive with tight junction protein measures. Using these genes as input for intercellular communications analysis, we prioritized ligand‐target interactions from microglia or astrocytes to pericytes. Some of the prioritized targets are transcription factors in pericytes. Using regulon analysis, we identified genes downstream of these transcription factors that are also significantly associated with lower CAA and membrane‐bound Aβ40 and higher tight junction protein levels. Conclusion We revealed glia‐to‐pericytes intercellular interactions and pericytic gene regulatory networks that are potentially perturbed by CAA and BBB breakdown. Future directions include analyzing external snRNAseq datasets and performing functional validations.

ABSTRACT Tendon adhesion is a major complication following tendon reconstruction surgery in the hand. Previous studies showed that a combination of hyaluronic acid (HA) and gelatin, covalently bound to the tendon surface, can provide a durable gliding surface to reduce adhesions in animal models both in‐vitro and in‐vivo. We have developed a similar product using good manufacturing practice (GMP) standards, in preparation for use in humans. This study aimed to examine the frictional properties and elastic modulus of the Manufactured Tendon Lubricant (MTL) when applied to an extrasynovial canine peroneus longus (PL) tendon in‐vitro. Twenty PL tendons along with the 2nd and 5th digits from adult canine forepaws, were obtained (mean age: 12 months, weight: 35 kg) from animals sacrificed in Institutional Animal Care and Use Committee (IACUC) approved protocols. Tendons were treated either with the MTL or saline solution, as a control. Eight tendons from each group (MTL vs saline‐treated) underwent friction testing, using a previously described protocol, followed by indentation testing to assess tendon elastic modulus (a marker of successful crosslinking). Two tendons from each group were not tested and were sent for scanning electron microscopy (SEM). The MTL group had significantly ( p < 0.001) lower friction and gliding resistance as well as higher elastic modulus of the PL tendons, compared to the saline group. This study demonstrates that MTL provides lubricating effects and fixation to the tendon surface and may be suitable for human testing.

ABSTRACT Chimeric antigen receptor T-cell (CART) and T-cell engager (TCE) therapies targeting B-cell maturation antigen (BCMA) are transforming the treatment landscape for relapsed multiple myeloma (MM). However, despite impressive initial response rates, most patients eventually relapse. To investigate this unmet medical need, we applied whole-genome sequencing (WGS) to MM cells from cohorts of 102 relapsed patients treated with anti-BCMA CART and TCE therapies. Several genomic alterations were associated with clinical outcomes, particularly primary refractoriness, including high genomic complexity and mutations in genes regulating plasma cell identity, which predicted resistance to therapy. Single-cell RNA sequencing further revealed that MM cells from refractory patients exhibited high proliferation signatures and reduced expression of TNFRSF17 (encoding BCMA), while were less enriched for plasma cell–associated transcriptional programs, a phenomenon we term “plasma cell identity escape.” This profile was strongly associated with immune dysregulation of CD8 T cells including increased activation and exhaustion. This evolution of MM toward a more proliferative and lineage-divergent state, refractory to the anti-BCMA T-cell redirecting therapies, was functionally validated in preclinical MM mouse models. Collectively, our results comprehensively define the cellular and molecular mechanisms underlying primary resistance to anti-BCMA therapies.

Abstract Background Standardizing beam modeling in the treatment planning system is challenging due to machine‐specific configuration variations. Because of the proton therapy system (PTS) complexity and continuous development, there is no vendor's representative (“golden”) beam data like in LINACs at this stage. Purpose The primary goal of this study is to create a PTS‐specific repository of source model parameters and beam commissioning data for scanning proton therapy treatment planning systems and establish a set of reference values with associated bands of variations for this data. Methods We conducted a survey of 31 PTSs at 27 proton therapy centers to gather beam commissioning source model parameters used in their treatment planning system, ensuring a diverse selection of PTSs from major vendors: Hitachi, IBA, Varian, and Mevion. For each proton therapy center, we collected (i) beam meterset calibration in protons per MU, (ii) in‐air spot size at isocenter in terms of standard deviation in x‐ (crossline, σ x ) and y‐ (inline, σ y ) directions, (iii) in‐air angular spread at isocenter in x‐direction (θ x ) and y‐direction (θ y ) and (iv) distal falloff. For each PTS, we plotted each parameter as a function of nominal energy and performed polynomial regression analysis with optimal model selection based on Akaike Information Criterion. We calculated intraclass correlation coefficients (ICC) to quantify inter‐PTS variability and generated uncertainty bands representing standard deviations across PTSs for each parameter. Additionally, we computed percentage deviations of each individual PTS from the overall multi‐vendor mean to identify systematic vendor‐specific patterns and outliers across the 31 total PTSs analyzed. Results Multi‐vendor beam parameter analysis across 31 PTSs revealed systematic vendor‐specific patterns in inter‐PTS consistency and energy dependencies. Beam meterset calibration demonstrated excellent inter‐PTS consistency across most vendors (ICC: Varian ProBeam ≈ 0, IBA Proteus ONE = 0.006, Mevion S250i with HYPERSCAN = 0.015), with IBA Proteus PLUS systems showing moderate variation (ICC = 0.156 for the dedicated nozzle, ICC = 0.248, for the universal nozzle). Spot size and angular spread variability were highly vendor‐dependent, ranging from excellent consistency in IBA Proteus ONE to substantial variation in IBA Proteus PLUS with the universal nozzle (angular spread ICC = 0.566). Distal falloff maintained excellent consistency across all vendors (ICC < 0.023), indicating reliable energy spread characteristics. Uncertainty band analysis revealed characteristic energy‐dependent patterns, with most systems showing larger uncertainties at lower energies for spot size and angular spread, while IBA Proteus PLUS with the universal nozzle exhibited the largest uncertainty bands overall. Percentage deviation analysis demonstrated vendor‐specific clustering patterns, with some systems showing systematic positive or negative deviations from the multi‐vendor mean. Conclusions The reference data provided in this work will help to cross‐check the beam model during treatment planning system commissioning. Furthermore, this information is useful for Monte Carlo applications and research, where beam characteristics are required to set up phase space parameters, thereby avoiding the need to model the entire PTS. Finally, the results of this study will have indispensable educational value by providing valuable insights into the characteristics and capabilities of different PTSs.

Abstract Background Adjuvant 5-FU monotherapy was previously the standard treatment for stage II/III rectal cancer. This study compared adjuvant FOLFIRI, FOLFOX, and 5-FU/LV. Methods Eligible patients had T3-T4 N0 or Tany N1-3 rectal adenocarcinoma ≤12 cm from the anal verge and received pre-operative or postoperative 5-FU chemoradiotherapy (CRT). Patients were randomized to adjuvant FOLFIRI (8 cycles), FOLFOX (8 cycles), or 5-FU/LV weekly (6/8 weeks; 3 cycles). The trial planned to enroll 3,150 patients but was closed early following activation of an alternative adjuvant rectal cancer study including bevacizumab (E5204). The primary endpoint was overall survival (OS); secondary endpoints included disease-free survival (DFS), sphincter preservation, tolerability, and quality of life (QOL). Results The trial enrolled 225 patients (179 randomized) before early closure. Grade 3/4 toxicity occurred in 59%, with neutropenia, leukopenia, and diarrhea being the most common. At a median follow-up of 9.7 years, no significant OS or DFS differences were observed. The rate of sphincter preservation was numerically higher with FOLFIRI and FOLFOX versus 5-FU/LV, but the difference was not statistically significant. Conclusion FOLFOX and FOLFIRI can be safely administered after CRT in rectal cancer patients safely with expected toxicities. Due to early trial closure and small sample size, the survival analysis was underpowered to detect a significant difference between regimens. ClinicalTrials.gov identifier NCT00068692

Abstract Introduction Microdissection testicular sperm extraction (MicroTESE) is a procedure traditionally performed in the operating room for men with nonobstructive azoospermia. Sperm are extracted for use in IVF-ICSI with thawed eggs harvested from female partners. Our practice has evolved to largely perform microTESE in the office with an oral anxiolytic and spermatic cord block, thereby avoiding the risk of general anesthesia and the increased cost of OR utilization. Objective To report sperm retrieval outcomes, embryo development outcomes, and live birth rate amongst patients undergoing office-based fresh microTESE for nonobstructive azoospermia. Methods A review was conducted of all patients who underwent office-based microTESE for nonobstructive azoospermia with a partner that who underwent concurrent oocyte retrieval for an intended in-vitro fertilization (IVF) intracytoplasmic injection (ICSI) cycle 2022-2025. All male partners underwent Y chromosome microdeletion testing, karyotype analysis, and hormonal testing preoperatively. Patients with a diagnosis of hypogonadism were offered medical therapy and those diagnosed with a varicocele were offered varicocelectomy prior to microTESE. Office-based microTESE was performed using a surgical operating microscope, spermatic cord block, and oral anxiolytic. Female partners underwent oocyte retrieval with oocyte cryopreservation prior to microTESE. Oocyte warming and ICSI was performed only following the collection of motile sperm from the fresh microTESE samples. Relevant demographic and clinical data were collected for patients and their partners. Descriptive data analysis was performed and reported as mean ± standard deviation. Results Nineteen patients underwent 20 office-based microTESE procedures. One patient had repeat microTESE after an initial successful sperm extraction and live birth with the intention of having a second child. Overall sperm retrieval rate was 53% for first time microTESE with mean search time 174±74 min. Amongst couples with successful sperm retrieval, normal fertilization rate was 40%. Early cleavage arrest was seen in 84% of normally fertilized oocytes with a 16% blastocyst rate. Embryo development arrested entirely in three couples, while the remaining seven couples had at least one blastocyst on day 5 and underwent embryo transfer. Of these, five resulted in a clinical pregnancy (71%) and all five of these couples had a successful live birth. Conclusions Our small series over 3 years demonstrates high sperm retrieval rates from office-based microTESE. Fertilization rates are comparable with those reported in the literature, and viable pregnancy rate and live birth rate amongst those with successful blastocyst development is high (71%). Given the financial cost associated with microTESE and IVF/ICSI for nonobstructive azoospermia patients, office-based microTESE may be a solution to make the process more financially accessible for couples. Disclosure No

Abstract Background: For large language models (LLMs) to reach their potential as information technology tools that make medication use safer, clinically relevant benchmarks capable of automated grading and designed specifically to measure the performance of LLMs for medication tasks are required. The purpose of this study was to design a suite of benchmarking tests reflective of Comprehensive Medication Management (CMM; the standard of care for medication optimization) and quantify the baseline performance of the latest LLMs. Methods: We established six benchmarks representing critical stages of the CMM process: drug formulation matching, drug order (sig) generation, drug route matching, drug-drug interaction identification, renal dose identification, and drug-indication matching. For each benchmark, we curated a clinician-annotated dataset comprising 250 standardized input-output pairs including both inpatient and outpatient medications. We evaluated the clinical knowledge retrieval capabilities of three LLMs: GPT-4o-mini, MedGemma-27B, and LLaMA3-70B. We employed a zero-shot prompting strategy, excluding in-context examples, to assess the models’ internal clinical knowledge rather than their few-shot learning potential. To check reliability, each model was run three times using a temperature of 0.7 (a mid-range value of an LLM setting controlling text generation randomness). Performance was assessed using task-specific evaluation metrics including precision (positive predictive value), recall (sensitivity), F1-score, accuracy, and correctness consistency across trials. Results: Across six benchmarks, LLaMA3-70B demonstrated the highest performance in four tasks: drug-formulation matching (F1, 54.0% [95 CI: 50.1-58]), drug-order generation (accuracy, 88.0%), drug-route identification (F1, 74.3% [95 CI: 71-78]), and drug-indication identification (accuracy, 97.6% [95 CI: 95.6-99.2]). In the drug–drug interaction task, GPT-4o-mini achieved the highest overall accuracy (70.4% [95 CI: 64.8-75.7]). For renal dose–adjustment identification, GPT-4o-mini demonstrated the highest F1 score (83.3% [95 CI: 77.6-88]). Correctness-consistency scores ranged from 8.0% to 97.6% across benchmarks, with no model exhibiting uniformly superior consistency. Conclusions: Model performance varied substantially across medication-related tasks. LLaMA3-70B demonstrated promising baseline performance in tasks involving formulation, ordering, route, and indication. GPT-4o-mini showed potential advantages in drug–drug interaction detection and renal dose adjustment. These findings underscore the need for task-specific evaluation when deploying models for medication-focused clinical decision support.

Abstract Immune checkpoint inhibitor–associated immune thrombocytopenia (ICI-ITP) has been described in case reports and small case series, but comprehensive data on its incidence, risk factors, clinical features, treatment, and outcomes are lacking. We reviewed medical records of all adults initiating ICI therapy between 2016 and 2023 at 29 US hospitals across 7 major cancer centers to identify patients with ICI-ITP. Multivariable logistic regression was used to identify risk factors, and Cox modeling was performed to assess the association between ICI-ITP, its severity, and mortality. Among 86 467 patients, ICI-ITP occurred in 214 (0.25%). Independent risk factors included lower baseline platelet count, combination ICI therapy, stage IV cancer, and additional immune-related adverse events. ICI-ITP occurred at a median of 8 weeks (interquartile range [IQR], 4-18) after ICI initiation, with a median nadir platelet count of 41 × 109/L (IQR, 17 × 109/L to 64 × 109/L). Patients were treated with glucocorticoids (n = 106 [49.5%]), immune globulin (n = 39 [18.2%]), and thrombopoietin receptor agonists (n = 29 [13.6%]). Recovery occurred in 161 patients (75.2%) at a median of 2.3 weeks (IQR, 1.0-5.3). Of 76 patients rechallenged with ICIs, 23 (30.3%) developed recurrent ICI-ITP. ICI-ITP and its severity were associated with higher all-cause mortality, with a nearly threefold increase in risk among patients with severe ICI-ITP than those without ICI-ITP (adjusted hazard ratio, 2.96 [95% confidence interval, 2.14-4.08]). These findings establish ICI-ITP as a rare but clinically significant complication of ICI therapy, provide, to our knowledge, the first large-scale description of its risk factors and clinical course, and underscore the importance of timely recognition and management.