Philadelphia College of Osteopathic Medicine
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Investigating Internalization, Intracellular Tracking, and Localization of NGR-Tagged Rubredoxin in HT-1080 Fibrosarcoma Cells
No full textIntroduction Fibrosarcoma is a highly aggressive soft tissue sarcoma with limited treatment options and poor response to conventional therapies, necessitating the development of more targeted treatment strategies. CD13, a transmembrane metallopeptidase overexpressed in tumor vasculature, represents a promising target for tumor-selective drug delivery. This study investigates the internalization and subcellular trafficking of an NGR-tagged Rubredoxin holoprotein (NGR-Rb) in HT-1080 fibrosarcoma cells as a potential targeted therapeutic strategy. Rubredoxin, a small iron-containing redox protein derived from Pyrococcus furiosus, is particularly well-suited for this approach due to its extreme stability and ability to function in harsh environments, making it an ideal candidate for tumor-targeted protein therapeutics.
Methods A time-course internalization experiment will be conducted to assess the uptake of NGR-Rb at defined time points (0, 15 min, 45 min, 2 h, and 4 h). Internalization will be confirmed using a dynamin inhibitor, Dynasore, and a 4°C control to differentiate between receptor-mediated and passive uptake. Subcellular localization will be determined through fluorescence microscopy and co-localization studies with nuclear (DAPI), mitochondrial (MitoTracker), and lysosomal (LysoTracker) markers.
Results This study aims to provide insight into the mechanistic pathways governing NGR-Rb internalization and trafficking, offering a foundation for targeted protein-based drug delivery approaches in fibrosarcoma and other CD13-expressing malignancies. By elucidating these mechanisms, this work contributes to the development of innovative, tumor-selective therapeutics designed to improve treatment efficacy while minimizing off-target toxicity.
Discussion The findings of this research will inform the rational design of NGR-conjugated therapeutics, facilitating their integration with chemotherapeutics or immunotherapies for enhanced cancer treatment. Future studies will expand into in vivo models to confirm clinical viability, paving the way for the development of protein-based targeted therapies with reduced systemic toxicity in fibrosarcoma and other CD13-expressing malignancies. As this project advances, it is expected to contribute to the broader field of tumor-targeted therapy, offering a novel approach to drug delivery in fibrosarcoma
Neuropsychological prediction of Alzheimer’s disease and functional abilities using brief screeners
No full textIntroduction: Alzheimer’s disease (AD) is a neurodegenerative disease that is primarily characterized by memory impairment, deficits in other cognitive domains, and decline in functional status. According to the Alzheimer’s Association, approximately 6.7 million Americans, age 65 and older, were living with AD in 2023. Patients often undergo lengthy neuropsychological evaluations that creates testing fatigue and ultimately affects the validity of results. Extensive literature also suggests that sex differences affect appropriate AD diagnosis in that women are more likely to demonstrate more impairment in instrumental activities of daily living (IADLs).
Objective: The purpose of this study is to investigate the ability of the Mini-Mental State Examination (MMSE) with the addition of a semantic fluency measure compared to the Montreal Cognitive Assessment (MoCA) and the Alzheimer’s Disease Assessment Scale-13 item Cognitive subscale (ADAS-Cog-13) in predicting a diagnosis of AD. Additionally, the current study aims to examine the potential clinical use of the ADAS-Cog-13 in detecting changes in functional abilities using the Functional Abilities Questionnaire (FAQ).
Methods: Permission has been granted by the Alzheimer’s Disease Neuroimaging Initiative (ADNI) to use their de-identified archival data to explore the efficacy of these instruments in predicting a diagnosis of AD as well as the relationship between functional decline, executive functioning, and biological sex. Participants will include individuals aged 65 years or older that are cognitively intact and formally diagnosed with AD in an initial screening visit. Statistical analyses will then be used to determine significance at a p-level less than or equal to 0.05.
Results: It is hypothesized that the MMSE supplemented with a semantic fluency task will predict a diagnosis of AD, demonstrate similar predictive ability to the MoCA in identifying individuals with AD, and be more sensitive than the ADAS-Cog-13 in predicting AD diagnosis. It is postulated that higher ADAS-Cog-13 scores on an executive functioning item will correlate with greater impairments in IADLs as measured by the FAQ, and that women will exhibit greater impairments on both the ADAS-Cog-13 and FAQ than men.
Discussion: The current study will address the limitation of the MMSE lacking an executive function measure, which has made the MoCA historically more superior in detecting cognitive impairment. In regards to functional decline, the ADAS-Cog-13 will also provide clinical insight into its potential in assessing IADL impairment and sex differences. Findings will shed light on the potential benefits of various approaches to brief cognitive assessment in clinical practice among different disciplines including neuropsychology, primary care, psychiatry, and neurology. Such providers can use such tasks to grant earlier diagnoses and interventions
Investigation Into the Epicardial Requirement for Crk and CrkL Adaptor Proteins in the Mammalian Heart
No full textINTRODUCTION: Congenital heart defects occur in ~1% of the population and increase the risk of cardiovascular disease. During cardiac development, epicardial cells delaminate into the subepicardium where they undergo epithelial-mesenchymal transformation (EMT) to differentiate into mesenchymal cells that become epicardium-derived cells (EPDCs). These EPDCs form coronary vascular cells. These EPDCs secrete paracrine factors that are essential for fetal myocardial growth and coronary vessel patterning. Epicardial EMT is involved in cardiac repair and regeneration. The epicardium retains the ability to differentiate into various cardiac cell phenotypes following myocardial injury or ischemia, thus contributing to cardiac tissue regeneration. Crk and CrkL are adaptor proteins that mediate key signaling pathways in mammalian development, including heart valve formation, angiogenesis for vascular remodeling, and neural crest cell maturation necessary for pharyngeal arch artery development and outflow tract septation. Crk and CrkL are expressed during mouse heart development in the epicardium, myocardium, endocardium and coronary vessels. Global deletion of Crk-CrkL in mice causes embryonic lethality due to cardiac defects. However, the specific roles of Crk and CrkL in the epicardium and coronary vessels have not been identified.
OBJECTIVE: Our goal was to identify the expression requirement of Crk and CrkL adaptor proteins in the heart during epicardium and coronary vessel formation.
METHODS: We used a conditional knockout strategy to generate mouse embryos with Wt1-Cre-driven deletion of Crk and CrkL from the epicardium (Crk-CrkL epi+/-). This approach enabled us to bypass the embryonic lethality caused by global Crk-CrkL deletion. Embryos were isolated at embryonic day (E) 13.5, 15.5, and 17.5 to assess cardiovascular development. We used hematoxylin and eosin staining for histological evaluation of cardiac structure. Also, we used immunofluorescence staining with antibodies to evaluate markers for epicardial (Wt1), myocardial (cardiac actin), vascular smooth muscle (SM22α), and endothelial (VE-cadherin) cell lineages. RNA sequencing was conducted on control and Crk-CrkL deletion hearts to identify differentially expressed genes and potential pathways targeted by Crk-CrkL.
RESULTS: Our analysis of Wt1 expression in E13.5 Crk-CrkL epi+/- hearts versus controls indicated a decrease in EPDC migration from the epicardium and reduced invasion into the compact myocardium. This deficit was resolved by E15.5. Immunofluorescence for VE-cadherin in E13.5 Crk-CrkL epi+/- hearts revealed superficially displaced primitive coronary vessels that failed to embed properly into the compact myocardium. However, this was resolved by E15.5 when the VE-cadherin expression pattern was similar in control embryos. This suggests coronary vessel patterning was restored after a brief developmental delay. Expression of the vascular smooth muscle marker, SM22α, in coronary vessels was consistent with VE-cadherin expression by E15.5.
CONCLUSION: Our data suggest Crk and CrkL play a critical role in early epicardial formation and EPDC migration, particularly in guiding coronary vessel patterning. Coronary vascular developmental defects were transient. This implies compensation occurs to restore epicardium and coronary vessel formation by later stages of development. These findings enable us to understand mechanisms of Crk and CrkL function in cardiac biology and shed light on the role of the epicardium in cardiac repair and regeneration
Preparation of Xanthohumol (XN)-incorporated sub-5 ultrafine iron oxide nanoparticles: Investigation of their effects on multiple myeloma cell growth
No full textIntroduction
Multiple myeloma (MM) is a hematological cancer that affects B lymphocytes. MM cells can undergo a homing process within the bone marrow microenvironment (BMM). The BMM helps MM cells undergo proliferation through interactions with bone marrow stromal cells. The proliferation aids in MM’s continued growth and spread throughout the BMM and into neighboring organs by releasing interleukin-6 and vascular endothelial growth factor. Xanthohumol (XN) is categorized as a prenylated flavonoid and has exhibited anticancer effects in in-vitro models. XN has downregulated the NF-kappa B signaling pathway, which is activated by interleukin-6 and aids in MM cell proliferation. Major challenges limiting the use of XN are poor aqueous solubility and lack of targeted drug delivery. To address this, we formulated anti-biofouling polyethylene glycol-block-allyl glycidyl ether (PEG-b-AGE) polymer-coated ultrafine iron oxide nanoparticles (µIONPs) loaded with XN (µIONP/XN) to improve the delivery efficiency to MM cells. An RGD motif of arginine, glycine, and aspartic acid was conjugated to the µIONPs as the targeting ligand for the αvβ3 integrin overexpressed on MM cells. This study compared RGD-µIONP/XN, RGD-µIONP (vehicle control), and µIONP/XN (non-targeted drug control) for cytotoxic effects on MM RPMI 8226 cells in an in-vitro model.
Methods
Throughout this project, several experiments were conducted to gather quantitative data. Presto blue assays were used to analyze cell viability, western blots were used to evaluated the expression of apoptosis-related markers, and a reactive oxidative species assay (ROS) was used to determine the ROS production by the three nanoparticles in RPMI 8226 cells. The release of XN from RGD-µIONP/XN was measured by high-performance liquid chromatography with or without a short-wave radio frequency (RF) stimulation.
Results
XN induced a dose and time-dependent decrease in cellular viability of RPMI 8226 cells with an IC50 value of 12.28 µM at 48 hours and 7.54 µM at 72 hours. The data from the Western blot analysis showed an upregulation of cleaved-caspase 3 with increased XN concentrations. Interestingly, results indicated that the RGD-µIONP (vehicle control) also induced cytotoxic effects on the MM cells in a time dependent manner. This urged us to analyze the generation of ROS by the three nanoparticles. The preliminary ROS results indicated that all the nanoparticles are producing ROS in a time dependent manner. Upon a short-wave RF stimulation for 10 minutes, greater than 30% of XN was released.
Conclusion
The results indicate the potential mechanism of cell death induced by the RGD-µIONP/XN to be at least in part if not all, apoptosis. Additional experimentation will be performed to evaluate the cytotoxic effects of RGD-µIONP, XN-µIONP, and RGD-µIONP/XN on RPMI 8226 cells upon short-wave RF stimulation and the mechanism of another programed cell death, ferroptosis, that these nanoparticles can induce
Podium Talk: Pyogenic Granuloma at the Ileocecal Junction: Unusual Culprit of Chronic Anemia and Intussusception.
No full textPyogenic granulomas, benign vascular lesions typically found in the skin and mucous membranes are rarely implicated in gastrointestinal pathology. This case presentation highlights an unusual yet critical manifestation of a large pyogenic granuloma in the small intestine, resulting in chronic anemia. We explore this rare condition’s clinical implications, emphasizing its diagnostic challenges and the potential for long-term misdiagnosis. Furthermore, we discuss effective treatment modalities and underscore the importance of heightened awareness among healthcare providers regarding this atypical presentation. This case serves as a reminder of the diverse clinical presentations of pyogenic granulomas and the necessity of thorough evaluation in patients with unexplained anemia
Breaking the stereotype: small cell lung cancer in a never-smoking patient
Full text linkIn the United States, lung cancer is the second most common type of cancer whilst also accounting for 1 in 5 cancer deaths, making it also the deadliest. The vast majority of lung cancer is non-small cell lung cancer (NSCLC), contributing to 87% of all cases, while small cell lung cancer (SCLC) makes up about 13% of all lung cancers1. While SCLC is strongly related to smoking history, here we present a 65-year-old African-American male without a history of smoking tobacco presenting with a rapid progression of unilateral thoracic pain, dyspnea, and unintentional weight loss. The diagnostic work-up included concomitant chest radiography and a computed tomography (CT) scan without contrast, which revealed a dense mass in the right lung. The patient underwent oncologic lung resection followed by chemotherapy and radiation. Considering the close association of SCLC with tobacco and the reduced likelihood of African-American men developing SCLC compared to Caucasian men, here we present a rare and interesting case of SCLC in a never-smoker African-American man
Refining the smoothness index for reaching in ataxic mice using a curved model
Full text linkSmooth, goal-directed reaching movements require precise motor coordination, a function often impaired in cerebellar ataxia. Distinguishing between normal and ataxic movement requires a measure of smoothness of movement. Previously a linear smoothness index was developed, which relied on deviations from linear trajectories and acceleration profiles of reaching movement in the mouse forelimb. This linear smoothness index approach may not accurately capture the natural movement patterns of this reaching motion in the mouse model due to the more natural curve the mouse exhibits in this movement. In this project, a new index was introduced by incorporating a curved path, developed from prior work demonstrating the utility of curved path models in modeling human hand trajectories. This curve incorporated the reach patterns of multiple reaching movements to offer a flexible representation of movement that accounts for natural kinematic variability. The goal was to compare the new index of smoothness to the linear model to find if the curved model provides a higher level of discrimination between normal and ataxic movement.
This revised metric was applied to analyze reaching behavior in normal and transgenic mice (n=8). Mice were head-plated and placed on water restriction for at least two days preceding training for reaching behavior. They were head-fixed on a mount and trained to reach for water drops dispensed from a needle approximately 1 mm from the snout. Reaching movements were tracked using DeepLabCut, with trajectory data collected and analyzed in MATLAB. All reaches were averaged into a reference trajectory or averaged curve against which deviations were measured. Electrophysiological recordings from the ventromedial thalamus were obtained simultaneously to assess neural correlates of reaching behavior. Previous linear-derived smoothness index study compared smoothness indices between ataxic and normal mice using an unpaired t-test with 78 trials and possessed a p-value of 0.0275. Preliminary results using the newer curve-based smoothness index with 61 trials possessed a p-value of 0.0148, indicating a more significant degree of difference between the two indices and potentially suggesting a more definitive standard for determining smoothness.
These findings demonstrate the relevance of curved path models in analyzing fine motor impairments in a preclinical setting. Refining smoothness metrics with a curved path model may provide a more physiologically accurate measure of motor impairment in ataxic models. Future studies will further validate this approach and explore its implications for assessing and potentially modulating cerebellar motor control in combination with thalamic electrophysiology recordings
Development and validation of a low-cost tandem-lens mesoscope for wide field fluorescence imaging of neural activity
Full text linkAdvancements in mesoscopic imaging have enabled large field-of-view (FOV) fluorescence microscopy, facilitating the study of neural activity across cortical networks. Traditional two-photon and confocal microscopy techniques provide high spatial resolution but are limited in their ability to capture large-scale neuronal interactions due to their restricted FOV. Mesoscopic imaging bridges this gap by allowing simultaneous observation of widespread neural circuits, making it a powerful tool for studying distributed activity patterns, such as those involved in sensorimotor processing. This approach is particularly useful for investigating the cerebello-thalamo-cortical pathway, where coordinated activity across multiple brain regions is critical. Following the design outlined by Jose et al., a low-cost tandem-lens mesoscope was constructed. This study focuses on the assembly, calibration, and initial validation of the mesoscope using fluorescence microscopy techniques. The mesoscope was assembled using commercially available optical components in a tandem-lens configuration. The system consists of a Thorlabs CS505MU CMOS camera, MVL75M23 objective lens, and MVL50M23 imaging lens, allowing for two distinct FOVs: 12.6 × 10.5 mm (large FOV, 1.5 cm WD) and 6 × 5 mm (small FOV, 1.0 cm WD). Excitation was provided by a 470 nm LED (Thorlabs SOLIS 470C), with emission captured through a DMLP490L dichroic mirror and Chroma fluorescence filters optimized for GCaMP imaging. The dichroic mirror was positioned at a 45-degree angle to direct excitation light toward the sample while allowing fluorescence emission to pass through to the camera. The system was mounted on a 60 mm optical cage, incorporating a dovetail translation stage for fine z-axis adjustments. To validate imaging capability, a Snellen chart was placed underneath the scope. Images were acquired using ThorCam software and analyzed for spatial resolution and signal uniformity. The working distance (WD) was adjusted using the translational stage to optimize focus. Preliminary imaging of the Snellen chart demonstrated the ability to visualize structures with high contrast across both FOV configurations. The system effectively captured fine-scale details, confirming its suitability for widefield imaging. In future experiments, validation of the mesoscope will be tested using the natural fluorescence of pollen grains. Also, application of the mesoscope for in vivo imaging of mice brains during a reaching task will be performed, capturing neuronal activity in the thalamus using GCaMP-expressing mice. This will enable real-time investigation of the cerebello-thalamo-cortical pathway during skilled motor behavior, contributing to a better understanding of neural circuits involved in movement control
GLP1 Receptor Agonist Increases Fracture Risk In Patients with Obesity
Full text linkIntroduction: GLP-1 receptor agonist medication usage has seen a rapid increase in usage not only for diabetic patients but now for obese and overweight patients who are searching for alternative weight loss methods. Although these medications are proving useful for helping patients lose weight, there are notable side effects that patients need to consider.
Methods: A retrospective case-control study was conducted using data from the NIH All of Us database. Patients were included based on the diagnosis of overweight and obesity. Patients were excluded based on diagnosis of type 1 or type 2 diabetes mellitus, A1c \u3e 6.5 and diagnosis of conditions that would increase the risk of fragility fracture.
The correlations between use of GLP-1 agonist drugs and side effects, specifically risk of bone fractures, will be analyzed and discussed
Chronic traumatic encephalopathy and number of repetitive impacts in American football players
Full text linkBackground:
Chronic Traumatic Encephalopathy (CTE) is a degenerative brain disorder associated with repeated head injuries over the course of an individual\u27s lifetime. A definitive diagnosis of CTE requires a post-mortem examination to thoroughly identify pathological markers such as hyperphosphorylated tau protein and astrocytic abnormalities. This review seeks to investigate the correlation between the amount of repetitive head impacts in American Football careers and the likelihood of developing CTE.
Methods:
Data was sourced from PubMed and compiled using the search “((CTE) and (Football))”, between the years of 2014 to 2025. The data included focused on the estimated number of head impacts of American Football players and the diagnosis of CTE post-mortem as well as its severity. Articles were excluded for reasons such as drug abuse or existing neurological issues. The Downs and Black risk of bias assessment tool was used to rate the quality of the selected articles.
Results:
With the data from the 6 articles that were chosen for this review, the mean duration of play for Mild CTE cases in American Football players was calculated to be 11.98 years; and for severe CTE, 15.46 years. Along with this, 5 of the 6 studies concluded that there is a significant positive correlation between number of years played and risk of CTE neuropathology development. Furthermore, all 6 articles concluded that there is a significant positive correlation between the number of repeated head injuries and development of CTE post-mortem. According to Montenigro et al. (2017), the average amount of impacts in a season is 580.5. Using this value, we extrapolate the number of impacts on average for mild cases of CTE vs severe cases of CTE.
Conclusion:
This literature review compiled data from multiple studies to objectively quantify the average number of career impacts of post-mortem CTE positive Football players to further highlight the correlation between number of impacts and severity of diagnosis. Players with more impacts demonstrated more severe stages (3-4) of CTE, compared to those with less impacts demonstrating more moderate stages (1-2). These findings highlight the significant neurological risks associated with prolonged exposure to repetitive head trauma and support the need for continued research into position-specific CTE risks and protective strategies for athletes in high impact sports. Furthermore, the information from this review can be used as a framework to educating athletes of their risks of CTE development with increased career length.
The American Football players that were diagnosed with CTE were broken up into two separate classification groups mild CTE and severe CTE. Increased levels of abnormal tau protein were found in these brains post-mortem in increasing levels as the severity increased. However, the consensus as to whether the age of first exposure or the kinetics of the trauma experienced by the players has a stronger influence on CTE risk remains to be seen based on the articles available