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  Reactive Aldehyde Species (RASP) Inhibitors Sequester MAA-Adducts and Reduce Pro-Inflammatory Cytokines

  Duncan Works

  Background: Post-translational modifications of self-proteins have been implicated in the pathogenesis of Rheumatoid Arthritis (RA). One of these protein modifications termed malondialdehyde-acetaldehyde-adduct (MAA) has recently gained interest for its involvement in RA. This protein adduct contributes to inflammation by inducing immune cells to generate pro-inflammatory cytokines, T-cell specific responses, responses and circulating autoantibodies. Recently, novel reactive aldehyde species (RASP) inhibitors (ADX-629 and ADX-246) became available that have been shown to prevent the formation of MAA adducts by covalently binding and sequestering MDA and AA in a mouse model of alcoholic liver disease. These inhibitors prevented the release of key inflammatory cytokines and protected animals from progressive liver damage. While these experiments demonstrated the binding of RASP inhibitors to MDA and AA prior to MAA formation, they did not determine the capacity of these agents to scavenge pre-formed MAA protein adducts. Therefore, the purpose of this study was to determine if RASP inhibitors sequester MAA-adducts and block the subsequent cellular release of pro-inflammatory cytokines.
  
  Significance of the problem: MAA modified self-proteins may render harmful effects in patients with RA. Preventing inflammation and inflammation leading to fibrosis by inhibiting cellular binding of could represent significant advancement in the treatment of RA.
  
  Hypothesis: ADX-629 and -246 will prevent MAA protein adducts from binding to macrophage receptors and, as a result, prevent the release of pro-inflammatory cytokines.
  
  Experimental Design: Human monocytic cells (U-937 cell line) were activated to professional macrophages using phorbol 12-myristate 13-acetate (PMA) for 48 hours. Following pre-treatment with decreasing doses of ADX-629 or ADX-246 for 30 minutes, macrophages were incubated with 25µg/mL of fibrinogen (FIB) or MAA-modified fibrinogen (FIB-MAA) for 24 hours. Supernatants were collected for measurement of IL-6 and MCP-1 using commercially available kits. Cells were collected and tested for membrane integrity using a lactate dehydrogenase (LDH) assay.
  
  Results: In the absence of RASP inhibition, FIB-MAA stimulation of cells significantly increased the release IL-6 with mean concentration of approximately 30pg/mL compared to FIB alone 6-fold increase. (p<0.0001). IL-6 release was significantly reduced with only 1 µM of ADX-629 and fell to native FIB levels with drug concentrations exceeding 10 µM (Figure 1A). ADX-246 demonstrated similar results (Figure 1B). Likewise, similar patterns were observed for the release of MCP-1 (data not shown). LDH assays showed no evidence of cellular toxicity regardless of ADX dose.
  
  Conclusions: In addition to confirming the capacity of RASP inhibitors to sequester MAA, results of this study demonstrate that both ADX-629 and ADX-246 attenuate or prevent MAA-modified proteins from initiating the macrophage-mediated release of pro-inflammatory cytokines implicated in RA pathogenesis. These findings support the need for additional in vivo investigations of these RASP inhibitors as novel therapies in in the management of RA and possibly in other conditions wherein MAA adducts mediate tissue damage.

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  SGLT2i Use Potentiates Acute and Chronic Radiation-Induced Side Effects in Prostate Cancer Radiation Therapy

  Duncan Works

  Radiation therapy is a common treatment modality offered to patients with localized prostate cancer. It can be associated with early radiation-induced toxicities including dysuria, nocturia, frequency, urgency, spasm, and rarely hematuria. Early toxicities usually resolve once the treatment period has ended. Chronic toxicities are less common, and rarely, patients may experience radiation-induced hemorrhagic cystitis and hematuria months to years after radiation. We herein describe the case of a 65-year-old male with a past medical history of type 2 diabetes mellitus who experienced hemorrhagic cystitis for months following his radiation therapy. The patient was on Sodium-Glucose Cotransporter-2 inhibitor therapy (Jardiance), which we highlight as a potential risk factor for hemorrhagic cystitis. After cessation of Jardiance and initiation of Ozempic (GLP-1 agonist), his urinary symptoms significantly improved. To the best of our knowledge, this is the first such case reported.

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  Medication Mayhem: A Skin-teresting Consult

  Tiffany Truong, Jonathan H. Ryder, Clayton Mowrer, and Jasmine R. Marcelin

  Introduction: Adverse reactions to medications often present with involvement of the integument. They are characterized by the rapid change of skin appearance (erythema and dryness) and associated symptoms (pruritus) culminating in a visible rash. The challenge for physicians is to determine the etiology of such rashes in order to effectively treat them. Often, cessation of the offending agent resolves the rash. Case Description: A 77-year-old male with lymphedema and over 30 episodes of cellulitis started 250 mg penicillin VK BID for prophylaxis. Eleven days later, he developed a symmetric, erythematous, scaling rash on his buttocks and perineal region with associated pruritus and bleeding. He denied any fevers or chills. The patient tried multiple over the counter medications for the rash without relief. Further medical history included chronic kidney disease, heart failure, hypertension treated with amlodipine, and overall body xerosis. Skin examination demonstrated diffuse lichenified plaques with marked fissures, scaling, and crusting on the buttocks. Dermatology was consulted, and the patient’s symptoms were attributed to symmetrical drug-related intertriginous and flexural exanthem (SDRIFE), a systemic drug-related contact dermatitis characterized by symmetric well-demarcated patches of erythema on the buttocks. This condition is also known as Baboon Syndrome due to its characteristic rash similar to the markings of a baboon. This can be caused by agents such as penicillin, hydroxyzine, and cashews, all of which the patient was exposed to. The Infectious Disease team recommended the discontinuation of Penicillin VK and hydroxyzine. The patient was switched to triamcinolone 0.1% ointment BID and clobetasol 0.05% ointment BID to the affected area with petrolatum for xerosis. A follow-up appointment with Dermatology demonstrated marked improvement. Discussion: Erythema and pruritus following initiation of a new medication is often indicative of an adverse reaction. Rashes from penicillin and hydroxyzine in patients without a history of previous reactions are less common but must also be considered. In this case, correlation of the rash with the administration of penicillin and hydroxyzine, as well as the infrequent ingestion of cashews, prompted cessation of the offending agents. However, diagnosis was delayed until these physical signs became evident. Additionally, the paradoxical reaction of hydroxyzine is typically overlooked as a culprit for erythema and rash given its intended purpose of minimizing pruritus. This case underscores the value of a thorough history and physical in combination with a broad differential in the diagnosis of pruritic rash and highlights the value in understanding polypharmacy and medical reconciliation, rather than adding agents when symptoms continue to arise.

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  The Effect of Maternal Diet on Fetal Outcomes

  Tiffany Truong, Matthew Van Ormer, Tara Nordgren, and Ann Anderson-Berry

  Maternal diet is critical for a successful pregnancy, as well as fetal health outcomes. Recent investigations reveal that dietary fats, such as omega-3 fatty acids, serve as substrates for the biosynthesis of specialized pro-resolving lipid mediators (SPM), which have anti-inflammatory and immune-stimulating effects. However, the relationship between maternal omega-3 fatty acid intake and maternal and cord plasma SPM levels in normal weight versus obese pre-pregnancy body mass index (BMI) deliveries is unclear.

  Pre-pregnancy obesity is associated with serious adverse pregnancy outcomes, including an increased risk of miscarriage, caesarean section, pre-eclampsia, and thromboembolism. Along with maternal risk, these complications lead to a four-fold increase in neonatal mortality, attributed to prematurity and macrosomia. Obesity-associated inflammation in early development, from intrauterine, peri-partum, and early childhood insults, may have lifelong impacts on the offspring. Studies are needed to identify modifiable factors in the intrauterine environment and developing fetus that can reduce inflammation and limit the negative consequences of obesity during pregnancy. Recent studies reveal certain omega-3 fatty acid derivatives actively attenuate and resolve pro-inflammatory processes. These SPMs may be key to the beneficial effects of omega-3 fatty acids. While the association between inflammation and obesity is clear, the protective mechanisms of SPMs against complicated birth in maternal-fetal health are a gap in the field. Currently, it is known that SPM production is dependent on intermediates of the omega-3 fatty acid metabolic pathway. However, it is unknown how material SPM production is related to omega-3 fatty acid intake. In recent studies, the Anderson Berry Lab has found strikingly low intakes of omega-3 fatty acids in pregnant woman. Thus, understanding the therapeutic value of omega-3 fatty acid intake and the role of SPMs in maternal-fetal outcomes addresses an unmet need. We hope to achieve two specific aims: 1) to identify the relationship between maternal omega-3 fatty acid intake and maternal and cord plasma SPM levels in normal weight pre-pregnancy BMI and obese pre-pregnancy BMI deliveries and 2) to evaluate similarities and differences in intakes, food security, and transportation security. Dr. Anderson Berry will provide review of the pathophysiology of adverse pregnancy outcomes, teach and assist in a literature search for relevant manuscripts to study, and provide quality assurance for accuracy throughout the data collection process.

  Over a 10-week period, the recruitment of additional subjects to augment current samples was successfully performed. Subject recruitment required the collection of informed consent, preparation of maternal and cord blood, preparation of placental tissue samples, and administration of a validated food frequency questionnaire. Over 100 new subjects were successfully enrolled in the study in this manner. Preliminary evaluation of differences in intakes, food security, and transportation security between obese and normal weight groups was completed. Due to technical equipment challenges and timing inconsistencies in data analysis, utilization of a targeted lipidomics approach to measure SPMs and determine the association between maternal omega-3 fatty acid dietary intake and maternal and cord plasma SPMs is in progress with the mass spectroscopy coil and protocol being fine-tuned on other, less valuable samples.

  In the future, we hope to employ a targeted lipidomics approach to measure SPM levels and determine the association between maternal omega-3 fatty acid dietary intake and maternal and cord plasma SPM levels at the time of delivery. We plan to analyze 80 existing samples (40 mother-infant pairs) consisting of maternal, cord, placental, and neonatal blood and breast milk, 32% of which had a pre-pregnancy BMI >30. Clinical data from these subjects and dietary data measured via a validated food frequency questionnaire have been obtained. Dr. Nordgren will then determine SPM levels in plasma and placental samples via liquid chromatography-tandem mass spectrometry-mediated lipid identification. Key lipids and metabolites to be characterized will include 18- HEPE, 15-HETE, RvE1, RvD1, RvD2, RVD3, RvD5, 17(R)-RvD1, Maresin-1, and protectin-D1. This technique will also allow for determination of the association between maternal and cord serum concentrations of SPMs of obese pre-pregnancy BMI delivery. Levels and associations with clinical pregnancy outcomes will be analyzed. It is hypothesized that in the presence of obesity mediated inflammation, adequate omega-3 fatty acid intake provides a pool of substrates for increased SPM production, minimizing poor pregnancy outcomes.