School of Medicine

Acute respiratory distress syndrome (ARDS) is an often fatal critical illness where lung epithelial injury leads to intrapulmonary fluid accumulation. ARDS became widespread during the COVID-19 pandemic, motivating a renewed effort to understand the complex etiology of this disease. Rigorous prior work has implicated lung endothelial and epithelial injury in response to an insult such as bacterial infection; however, the impact of microorganisms found in other organs on ARDS remains unclear. Here, we use a combination of gnotobiotic mice, cell culture experiments, and re-analyses of a large metabolomics dataset from ARDS patients to reveal that gut bacteria impact lung cellular respiration by releasing metabolites that alter mitochondrial activity in lung epithelium. Colonization of germ-free mice with a complex gut microbiota stimulated lung mitochondrial gene expression. A single human gut bacterial species, Bifidobacterium adolescentis, was sufficient to replicate this effect, leading to a significant increase in mitochondrial membrane potential in lung epithelial cells. We then used genome sequencing and mass spectrometry to confirm that B. adolescentis produces L -lactate, which was sufficient to increase mitochondrial activity in lung epithelial cells. Finally, we found that serum lactate was significantly associated with disease severity in patients with ARDS from the Early Assessment of Renal and Lung Injury (EARLI) cohort. Together, these results emphasize the importance of more broadly characterizing the microbial etiology of ARDS and other lung diseases given the ability of gut bacterial metabolites to remotely control lung cellular respiration. Our discovery of a single bacteria-metabolite pair provides a proof-of-concept for systematically testing other microbial metabolites and a mechanistic biomarker that could be pursued in future clinical studies. Furthermore, our work adds to the growing literature linking the microbiome to mitochondrial function, raising intriguing questions as to the bidirectional communication between our endo- and ecto-symbionts.

BACKGROUND: The COVID-19 pandemic exacerbated existing disparities in healthcare access and outcomes, particularly among underserved communities. As one site participating in the NIH-funded Community Engagement Alliance Against COVID-19, our focus was to address COVID-19 disparities by training immigrant and refugee communities to advocate for their needs by increasing capacity to campaign for policy-level changes. OBJECTIVE: To evaluate the impact of a train-the-trainer policy advocacy program for ethnically-based community leaders within San Diego County using a mixed-methods evaluation. METHODS: We partnered with a non-profit social change, intermediary organization to adapt a five-session, 4-hour per session training that was conducted over five weeks. A baseline survey, pre- and post-training surveys, and ethnographic documentation were employed during each session. RESULTS: Among participants (n = 16), 50% were Latino(a), 25% were Somali Bantu, and 25% were Karen. Training results were relatively stable with slight variations in perceptions within and between sessions. The first session showed a slight decrease in confidence by the training participants, while sessions 3, 4, and 5 showed increases in confidence. Ethnographic documentation revealed that engagement patterns evolved over time, with the Latino(a) participants having the highest levels of engagement initially but with more equitable engagement across participants by the final session. CONCLUSION: These findings provide valuable feedback which will aid in the improvement of the training sessions for future use. This study also underscores the potential for community leaders to effectively advocate for policy changes and offers insights for future empowerment initiatives.

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Insulin is an important regulator of whole-body glucose homeostasis. In insulin sensitive tissues such as muscle and adipose, insulin induces the translocation of glucose transporter 4 (GLUT4) to the cell membrane, thereby increasing glucose uptake. However, insulin also signals in tissues that are not generally associated with glucose homeostasis. In the human reproductive endocrine axis, hyperinsulinemia suppresses the secretion of gonadotropins from gonadotrope cells of the anterior pituitary, thereby linking insulin dysregulation to suboptimal reproductive health. In the mouse, gonadotropes express the insulin receptor which has the canonical signaling response of IRS, AKT, and mTOR activation. However, the functional outcomes of insulin action on gonadotropes are unclear. Here, we demonstrate through use of an optimized cell fractionation protocol that insulin stimulation of the LβT2 gonadotropic cell line results in the unexpected translocation of GLUT1 to the plasma membrane. Using our high purity fractionation protocol, we further demonstrate that though Akt signaling in response to insulin is intact, insulin-induced translocation of GLUT1 occurs independently of Akt activation in LβT2 cells.

Extrachromosomal circular DNA (ecDNA) has been found in most types of human cancers, and ecDNA incorporating viral genomes has recently been described, specifically in human papillomavirus (HPV)-mediated oropharyngeal cancer (OPC). However, the molecular mechanisms of human-viral hybrid ecDNA (hybrid ecDNA) for carcinogenesis remains elusive. We characterize the epigenetic status of hybrid ecDNA using HPVOPC cell lines and patient-derived tumor xenografts, identifying HPV oncogenes E6/E7 in hybrid ecDNA are flanked by previously unrecognized somatic DNA enhancers and HPV L1 enhancers, with strong cis-interactions. Targeting of these enhancers by clustered regularly interspaced short palindromic repeats interference or hybrid ecDNA by bromodomain and extra-terminal inhibitor reduces E6/E7 expression, and significantly inhibites in vitro and/or in vivo growth only in ecDNA(+) models. HPV DNA in hybrid ecDNA structures are associated with previously unrecognized somatic and HPV enhancers in hybrid ecDNA that drive HPV ongogene expression and carcinogenesis, and can be targeted with ecDNA disrupting therapeutics.

BACKGROUND: Interleukin-18 (IL-18), or interferon (IFN)-γ-inducing factor, potentiates T helper 1 and natural killer cell activation as well as CD8+ T-cell proliferation. Recombinant IL-18 has displayed limited clinical efficacy in part due to the expression of the decoy receptor, IL-18 binding protein (IL-18BP). A series of IL-18 variants that are devoid of IL-18BP binding, termed DR18 (decoy-resistant IL-18), was developed via directed evolution. We tested DR18 using oncolytic adenovirus (oAd) as a platform for delivery in syngeneic mouse tumor models. METHODS: oAd harboring wild-type IL-18 or DR18 (oAdDR18) was constructed by inserting IL-18 mutant into modified oAd backbone with Ad5/3 chimeric fiber. The delivery effect and IFN-γ induction were determined by ELISA. The antitumor efficiency of oAdDR18 was tested in CT26, B16BL6 and 4T1 tumor-bearing mice, or athymic nude mice and compared with recombinant DR18 protein (rDR18). 4T1 lung metastasis model was used to evaluate the antitumor efficiency of local and distant tumors. Antitumor memory and synergistic effect with an anti-programmed cell death protein-1 (PD-1) antibody was evaluated. The phenotypes of the immune cells in tumor microenvironment were analyzed by flow cytometry and immunohistochemistry. RESULTS: Mice received oAdDR18 maintained stable production of IL-18 and IFN-γ compared with those received rDR18. Intratumoral delivery of oAdDR18 significantly reduced tumor growth across several tumor models, but not in the athymic nude mouse model. Mice that had tumor remission showed antitumor memory. The antitumor effect was associated with intratumor infiltration of CD4+ and CD8+ T cells. DR18 delivered by oAd demonstrated long-lasting and enhanced antitumor activities against local and distant tumors compared with that received rDR18 or wild-type IL-18 delivered by oAd (oAdwtIL-18). oAdDR18 treatment also reduced 4T1 lung metastasis. In addition, combination of this virotherapy with immune checkpoint inhibitors (ICIs)like the anti-PD-1 antibody further enhanced the antitumor activity as compared with respective monotherapy. CONCLUSIONS: oAdDR18 demonstrates enhanced antitumor activities through the induction of stronger local and system immunities and modulation of the tumor microenvironment compared with those of oAdwtIL-18 and rDR18. A combination of oncolytic virotherapy with cytokine engineering would lead to cytokine-based therapeutics for cancer and other diseases.

PURPOSE: Patients with invasive lobular carcinoma (ILC) face high rates of positive margins and completion mastectomy, which can be improved with the use of specific techniques, such as oncoplastic surgery. However, prior studies have shown that type of breast cancer surgery performed is also associated with patient factors such as elevated body mass index (BMI). Thus, this study investigates whether BMI impacts the type of surgical interventions in patients with ILC. METHODS: A retrospective analysis of 705 patients with stage I-III ILC from an institutional database was conducted. Patients were stratified by BMI (underweight, normal weight, overweight, obese). Pearsons Chi-square, ANOVA, and multivariable logistic regression were used to evaluate the relationship between BMI and surgical procedures. RESULTS: Breast-conserving surgery (BCS) was the initial operation in 60% of patients, with no significant difference by BMI. Among those undergoing BCS, patients with obese BMI were significantly more likely to undergo oncoplastic surgery (46.9% vs. 7.7%, 37.3%, and 33.6% for underweight, normal, and overweight, respectively, p = 0.032). Obese BMI patients undergoing mastectomy were less likely to have reconstruction compared to those with underweight, normal weight, and overweight BMI (44.2% vs. 50%, 71.1%, and 64.1%, p = 0.002). CONCLUSION: Overweight/obese BMI patients with ILC underwent different surgical interventions compared to those with lower BMI. While initial BCS rates were similar, overweight/obese patients had higher oncoplastic surgery rates in BCS and lower reconstruction rates in mastectomy. Further research is needed to understand BMIs impact on surgical decisions and outcomes in ILC.

Significance

BACKGROUND: Historically, the clinical characteristics and treatment pathways for patients with uterine fibroids and heavy menstrual bleeding have differed between White and Black women. OBJECTIVE: To provide a contemporary comparison of patient characteristics and treatment patterns among White and Black women with uterine fibroids and heavy menstrual bleeding in the United States. STUDY DESIGN: This retrospective cohort study included administrative claims data from 46,139 White and 17,297 Black women with uterine fibroids and heavy menstrual bleeding from the Optum Clinformatics database (January 2011-December 2020) and 7353 White and 16,776 Black women from the IBM MarketScan Multi-State Medicaid Insurance database (January 2010-December 2019). Patients were indexed at their initial uterine fibroid diagnosis claim and were required to have a claim for heavy menstrual bleeding and ≥12 months of continuous enrollment pre- and postindex. Patients were followed until the earliest of death, disenrollment, hysterectomy date, or end of study database. Outcomes were stratified by race and included patient demographics, clinical characteristics, pharmacologic treatment patterns, and surgeries/procedures. Pearsons Chi-square test for categorical variables and Students t-test for continuous data were used to evaluate differences in baseline characteristics. Descriptive statistics were used to characterize treatment pathways for hormonal contraceptive use in women with ≥24 months of follow-up. Kaplan-Meier survival analysis was used to estimate time until hysterectomy, with log-rank testing to assess between-group differences. RESULTS: The mean (standard deviation) duration of follow-up was 44.6 (27.9) and 41.0 (24.9) months in the commercial and Medicaid databases, respectively. Mean (standard deviation) age at uterine fibroid diagnosis was lower for Black than White women in both databases (commercial: 42.3 [6.5] vs 44.4 [6.3] years; P<.0001; Medicaid: 39.6 [7.1] years vs 40.2 [7.2] years; P<.0001). Anemia was more prevalent in Black vs White women in both databases (commercial: 5.9% [1028/17,297] vs 3.6% [1648/46,139]; P<.0001; Medicaid: 7.0% [1180/16,776] vs 4.5% [331/7353]; P<.0001). In the commercial database, approximately one-half of women had claims for ≥1 bulk symptom, with no significant differences between groups. In the Medicaid database, significantly more White than Black women had claims for bulk symptoms (77.0% [5665/7353] vs 68.4% [11,477/16,776]; P<.0001). Approximately 40% of all patients received hormonal drug therapies as initial treatment, most commonly hormonal contraceptives. However, discontinuation of hormonal contraceptive therapy was nearly universal, with one-half discontinuing within a median treatment duration of ∼5 months. Most women stopped treatment after 1 or 2 agents (commercial: White, 89.9% [9757/10,857]; Black, 90.0% [3594/3993]; Medicaid: White, 92.2% [1635/1773]; Black, 94.2% [4454/4726]). Hysterectomy was the most common procedure, and was more common among White vs Black women (commercial: 43.9% [20,235/46,139] vs 37.8% [6536/17,297]; Medicaid: 46.8% [3444/7353] vs 32.0% [5364/16,776]). CONCLUSIONS: Black women with UF-HMB were diagnosed at a younger age than White women, and White women had higher hysterectomy rates than Black women, representing a shift from earlier researched treatment patterns. Patients with UF-HMB were also highly reliant on hormonal contraceptives, followed by nearly universal therapeutic discontinuation.