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Importance of lipid droplets to immunosuppression in Coxiella burnetii-infected alveolar macrophages.
No full textCoxiella burnetii, the known causative agent of Q fever, is an obligate intracellular bacterium capable of infecting host alveolar macrophages and residing within these cells for several years. While causing mild symptoms at initial infection, C. burnetii retains the ability to survive long-term by disseminating and causing serious complications such as culture-negative endocarditis. Previous studies in endocarditis patients demonstrated that C. burnetii suppresses the host immune response via production of prosta-glandin E2 (PGE2) and modulation of the anti-inflammatory cytokine IL-10 and proin-flammatory nitric oxide (NO). C. burnetii is also found in lipid droplet-rich foam cells suggesting the importance of host cell immune response and lipid metabolism during bacterial infection. Previous studies in our lab revealed that C. burnetii manipulates host cell lipid storage organelles, lipid droplets (LDs) via its type 4 secretion system (T4SS) which secrets effector proteins into the host cell cytoplasm to modulate cellular processes. Further, we observed that breakdown of LDs is essential for C. burnetii growth suggesting the importance of LD-derived lipids during bacterial infection. LD breakdown releases arachidonic acid, a PGE2 precursor. Since PGE2 plays an immu-nosuppressive role during C. burnetii infection, we hypothesize that LD-derived PGE2 production leads to modulation of IL-10 and NO. To assess this, we determined IL-10 and NO production enzyme iNOS gene expression in mouse alveolar macrophages by quantitative Real Time PCR (RT-qPCR). Compared to uninfected cells, iNOS gene expression decreased while IL-10 expression remained unchanged. Further we per-formed ELISA to determine IL-10 protein level and identify the contribution of LDs con-tribute IL-10 production. Compared to uninfected cells, IL-10 production in C. burnetii-infected cells remained unchanged. Surprisingly, compared to untreated cells, blocking LD formation and breakdown with specific inhibitors resulted in increased IL-10 produc-tion. This suggests that LDs play an important role in IL-10 production in C. burnetii-infected cells. However, if IL-10 contributes to immunosuppression is yet to be deter-mined. Ongoing studies are identifying the importance of LDs in NO production. Future studies will determine the changes in the overall cytokine profile in the presence/absence of LD breakdown
The Role of Notum in Bone’s Response to Mechanical Loading and Aging
No full textOsteoporosis is a silent, dangerous, pathology frequently undiagnosed until patients experience a major fracture. Fractures from osteoporosis lead to a 20% increase of mortality, revealing a need for novel anabolic therapies. Recently, mice lacking Notum protein expression were reported to have increased endocortical bone formation and strength. Notum is an inhibitory protein of the Wnt pathway, a major growth pathway in mammalians. The hypothesis for this work is Notum expression is affected by mechanical loading and aging. The goal of these studies is to investigate factors that could affect Notum expression, with the future direction of identifying factors that when pharmacologically modulated could promote anabolic bone growth. After establishing No-tum expression in the osteocyte cell line, MLOY4s, Notum expression was monitored in response to mechanical loading, mimicked via orbital shear stress for 15 min, 1hr, and 2hr. In addition, Notum expression was analyzed in aged bone. Our preliminary results suggest mechanical loading decreases Notum protein expression, while aging increases the expression of the Notum gene (p\u3c0.05). . With these results our hypothesis is supported, and future studies repressing Notum expression in bone as novel approach to promoting anabolic bone growth will be conducted
Diagnosis and Management of Postpartum Eclampsia in the ED
No full textPostpartum preeclampsia refers to hypertension and proteinuria after delivery, and the occurrence of a seizure within six weeks after delivery is called postpartum eclampsia. Etiology is unknown but theorized to involve abnormal placental formation. Major risk factors include preeclampsia, multiple gestations, obesity, and 1st pregnancy. First pregnancy was the only known risk factor in this case. A 24-year-old Hispanic female (Gravida 1, two weeks postpartum) presented to the ED after a tonic clonic seizure, witnessed by her husband. Except for acquiring ß-hemolytic strep, she had a normal pregnancy and full-term vaginal delivery. Past medical and family history of seizures were negative. Her preeclampsia during pregnancy was closely monitored and did not require medication. She reported feeling short of breath prior to the seizure, which led to consideration of other causes of the seizure. The differential included postpartum eclampsia, new onset seizure, head trauma, intracranial hemorrhage, PE, and syncope. Patient was placed in high acuity on a cardiac monitor to track her BP and kept on seizure precautions. A magnesium sulfate drip was started since eclampsia was the most likely differential. Upon attempting a 6 gm bolus, she developed tachycardia so the drip was reduced to 2 gm/hr. Head CT, chest CT, and EKG ruled out the head bleed, PE, and syncope, respectively. Patient had a BP of 140/99, elevated LFTs, and proteinuria. Based on the patient’s eclamptic episode within the six week window, she was diagnosed with postpartum eclampsia. While the overall prevalence of postpartum preeclampsia has remained steady, incidence of postpartum eclampsia has fallen due to improved prenatal care and identification of preeclampsia. Despite these improvements, this case highlights the importance of providing women additional counseling on postpartum preeclampsia and hypertension management as well as the challenges in early identification of postpartum eclampsia in the ED
Implementing Fluid Education and Adherence with Hemodialysis Patients
Get PDFBackground: Outpatient hemodialysis (HD) requires patient compliance and adherence to their prescribed treatment, including fluid and dietary restrictions. Without compliance and adherence, patients are at an increased risk of morbidity and mortality. Regardless, hemodialysis patients continue to struggle with compliance and continue to have increased hospitalizations and mortality. Purpose: The purpose of this project was to examine whether individualized education on the importance of fluid and dietary restrictions is done with hemodialysis patients will increase compliance. Methods: An educational intervention was done with hemodialysis patients who were given a pre-assessment questionnaire to assess baseline knowledge before education. After education, a post-assessment questionnaire was completed to assess if the individualized education increased their knowledge. Implementation Plan: Implementation of this project included pre-and post-assessment of hemodialysis patients\u27 knowledge of fluid and dietary restrictions. Comparison or patients\u27 pre and post-intervention fluid weight gain was assessed, documented, and compared. The findings were shared with the dialysis unit staff and patients. Conclusions: The post-assessment showed that there was an increase of knowledge among patients who participated. There, was no significant correlation between understanding of fluid gain and patients\u27 demographics. Patients\u27 who adhered to their fluid restrictions had decreased fluid weight gain between their treatments and reported feeling better overall. As with some hemodialysis patients, there were outlying factors that went beyond the control of the patient, including family involvement, living environment, and multiple chronic diseases which had conflicting dietary needs
Just because it has been there for years doesn’t mean it is not important: Koenen Tumor and Tuberous Sclerosis
No full textFracture Healing in Aging: Improving Angiogenesis
No full textBACKGROUND: With an aging world population, there is an increased risk of fracture and impaired fracture healing. Enhancement of angiogenesis is one potential mechanism for improved fracture healing. The question then becomes: How do therapeutic targets improve these angiogenic and vasculitic conditions to facilitate successful fracture union? The current therapy, bone morphogenetic protein 2 (BMP-2), while successful in promoting fracture union, presents with several potential side effects, including excess bone growth and an increased risk of developing cancer. We hypothesize that thrombopoietin (TPO) may serve as a novel therapeutic modality for improved angiogenesis and fracture union. Here we characterize and stimulate endothelial cells (ECs) with TPO in vitro. OBJECTIVE: We hypothesize that thrombopoietin (TPO) may serve as a novel therapeutic modality for improved angiogenesis and fracture union. METHODS: Five young (3 month-old) and 5 old (22-24 month-old) C57BL/6 mice were euthanized and bone marrow was collected from the tibia and humeri for EC isolation. Additionally, five young mice underwent a 2 mm femoral diaphyseal defect surgery. Three weeks post-surgery mice were euthanized and bone marrow from the ipsilateral and contralateral tibia as well as the humeri were collected and ECs isolated. All ECs were assessed for vessel-like formation and proliferation. All in vitro studies were completed at least 3 times with triplicate samples. Statistical analyses were performed using a two-way ANOVA with Tukey’s post-hoc testing. All procedures were approved by the Indiana University IACUC. RESULTS: In vitro characterization of ECs showed impaired vessel-like formation, branch length, number of nodes, mesh area, and proliferation in old ECs compared to young ECs. ECs isolated from the ipsilateral tibia of mice undergoing a SBD surgery had impaired ECs vessel-like parameters and proliferation compared to ECs from the contralateral tibia. Finally, treatment of old ECs with TPO improved vessel-like parameters and proliferation compared to untreated cells. CONCLUSIONS: Although preliminary, our results confirm that EC proliferation and vessel formation are impaired in aging and in the ipsilateral tibia of mice, which underwent a femoral SBD surgery. Although BMP-2 is used by orthopaedic surgeons for bone healing, its use is not without side effects, leaving researchers to investigate alternative treatments. We have previously shown that TPO is effective for bone healing. Here we show that TPO improves vessel formation in aged ECs, and therefore, TPO may improve angiogenesis and fracture healing in old mice, which is currently under investigation
Add It Up: Using Data to Make Informed Spatial Decisions
Get PDFIn 2016, Marian University’s library began collecting data on how patrons utilized our building. We conducted student surveys, held focus groups, and collected spatial data using Suma, an open-source assessment toolkit.
In 2019, in response to the student surveys, the library began upgrades to our building including new furniture and additional technology. We utilized results from Suma studies to make evidence-based decisions about where to focus our improvements.
This poster covers using SUMA for assessment. It covers what SUMA is, what sort of data can be collected, and how it can be tailored to your library’s needs
IBD Clinical Trials Competitive Landscape
No full textBackground: Inflammatory Bowel Disease (IBD) is a chronic relapsing and remitting inflammatory disease that affects the GI tract. IBD has two major types: Crohn’s disease and Ulcerative Colitis. IBD occurs when the immune system attacks the intestinal lining, which results in prolonged inflammation and damage to the tissue. An IBD pa-tient’s quality of life is often reduced with symptoms of fatigue, urgency, abdominal pain and persistent diarrhea. With the advent of biologics and anti-TNF therapy, treatment options for IBD have vastly improved, yet only about one fifth of patients are in remis-sion one year after diagnosis. Objective: One goal of this project is to assess the competitive environment of biologics clinical trials in IBD. A secondary goal is to summarize mirikizumab’s phase II trial results. Method: This project acts as a secondary analysis and review of the ongoing and completed randomized placebo and active comparator clinical trials for IBD with biologics. Trial data was obtained from clinicaltrials.gov and trialtrove, and focuses on phase II through phase IV trials. The drugs of interest were upadacitinib, filgotinib, tofacitinib, risankizumab, brazikumab, mirikizumab, guselkumab, ustekinumab, vedolizumab, etrolizumab, etrasimod and ozanimod. This analysis compares the key endpoints, endoscopic response, endoscopic remission, PROs and CDAI remission, of RCTs in Ulcerative Colitis and Crohn’s Disease. Results: 24 trials are in progress for Ulcerative Colitis, while 25 trials are in progress for Crohn’s Disease. The mechanisms of action include JAK inhibitors, IL-23 p19 inhibi-tors, IL-12/23 p40 inhibitors, integrin antagonists and sphingosine phosphate receptor agonists. The majority (80% and 84% respectively) of trials were phase III. In UC… • 5 trials are completed with positive results • 5 trails are active, not recruiting • 14 trials are recruiting In CD… • 7 trails are completed with positive results • 1 trial is active, not recruiting • 16 trials are recruiting or planned Conclusion: These trials represent new treatment options for primary nonresponders and refractory patients that previously had no other treatment options. With new medications on the horizon, it is essential to consider which patients will be benefiting most from these treatments. More completed clinical trials are needed to compare these biologics head-to-head for specific patient populations so that clinicians can accurately choose the most effective treatment. Mirikizumab’s phase III trial aims to compare its effectiveness with ustekinumab, an IL-12/23 p40 inhibitor. Inhibition of IL-23 may offer a lower risk of infections compared to anti-TNF therapy
Role of peroxisome proliferator-activated receptor gamma (PPAR-γ) in Coxiella burnetii infection
No full textPeroxisome proliferator-activated receptors (PPARs) are important nuclear receptors in regulating macrophage lipid metabolism and modulating cellular inflammatory responses. In recent years, host cell PPARs have been linked to bacterial pathogenesis and macrophage infections. Specifically, bacteria-induced manipulation of lipid droplets (LD), host lipid storage organelles, are directly associated with PPAR-γ activation. For example, the intracellular pathogens Mycobacterium bovis and M. leprae breakdown the macrophage LDs to release free fatty acids. These free fatty acids subsequently activate PPAR-γ to induce an anti-inflammatory immune response, thus sustaining Mycobacterial intracellular growth. Our previously published studies report that the intracellular bacterium Coxiella burnetii induces host cell LD breakdown to support its growth in macrophages. Coxiella is known to cause non-culturable endocarditis that can present up to 20 years post initial infection, suggesting its ability to survive long-term in the host by subverting the host immune response. Our overall goal is to identify the strategies that Coxiella employs to manipulate the host immune response. This could ultimately aid in developing therapeutic targets to inhibit Coxiella’s intracellular growth. Since LD breakdown is important for intracellular Coxiella growth through the release of free fatty acids (PPAR-γ agonists), we hypothesize that Coxiella infection activates host cell PPAR-γ to induce an anti-inflammatory environment which supports bacterial growth and infection. To test this hypothesis, we quantified PPAR-γ gene expression in Coxiella-infected macrophages using quantitative Real Time PCR (qRT-PCR) at different times post-infection. Compared to uninfected cells, infected macro-phages demonstrated increased PPAR-γ gene expression suggesting that Coxiella induces PPAR-γ activation. Ongoing studies are focused on identifying PPAR-γ tran-scriptional activity in Coxiella-infected cells and the downstream targets. Future studies will determine if PPAR-γ activity is dependent on host cell LD metabolism and identify potential PPAR-γ antagonists as therapeutic targets to inhibit Coxiella intracellular growth
Further Elucidation of the Role of Protein Kinase-C and Rho Kinase in the Mechanism of Action of Bitter Melon Extract on Human Metastatic Breast Cancer Cells
No full textIntroduction: It is well documented in the literature that Bitter Melon Extract (BME) inhibits the growth and proliferation of MCF-7 breast cancer cells. While the exact mechanism of this inhibition is unknown, it is possible that Protein Kinase C (PKC) and Rho Kinase (ROK) play a role in this mechanistic pathway due to their known critical roles in cell migration, division, and survival. Previous research has shown that ROK inhibition results in decreased cell viability and decreased MYPT1 (Myosin phosphatase targeting subunit-1) expression, while PKC inhibition alone does not have a significant effect. Aims: The aim of this research is to investigate the cytotoxic mechanism of action enacted by BME on MCF-7 cells and how PKC and ROK might be involved in this mechanism by building on past research to create more robust data and a better understanding of the metabolic state of MCF-7 cells under the set conditions. Methods: Glucose assays were run via a GM-100 glucometer to test for cell metabolism as a measure of cell viability on BME dose (0%, 0.5%, 1%, 2%, 5%, and 10% BME (v/v)) study samples, PKC inhibitor (GF109203x; 0.5 uM) samples, and ROK inhibitor (H-1152; 1 uM) samples. The samples used in each of the three conditions were secured from past research. Results & Discussion: BME dose-dependently increased glucose remaining in the culture media. The sample sizes for different doses of BME were as follows; at 0% (n=4), 0.5% (n=5), and 1% (n=5), while 2% (n=2), 5% (n=3), and 10% (n=3). The data suggests two possibilities; (i) with increasing BME dose, there is an increasing effect on the MCF-7 cell’s metabolism of glucose, and that this decrease in ability to metabolize glucose when in the presence of increasing [BME] could play a role in the cytotoxic mechanism of BME on MCF-7 cells. (ii) The BME kills cells and hence glucose is not used as much thus leaving plenty of glucose in the medium. Our results further showed that neither PKC inhibitor samples (n= 7) nor ROK inhibitor samples (n=11) showed significant differences in [glucose]. This suggests that the measured effect in glucose metabolism in the BME dose-dependent study samples is independent of PKC and ROK mechanistic pathways