A Review of Serotonin in the Developing Lung and Neonatal Pulmonary Hypertension.

Serotonin (5-HT) is a bioamine that has been implicated in the pathogenesis of pulmonary hypertension (PH). The lung serves as an important site of 5-HT synthesis, uptake, and metabolism with signaling primarily regulated by tryptophan hydroxylase (TPH), the 5-HT transporter (SERT), and numerous unique 5-HT receptors. The 5-HT hypothesis of PH was first proposed in the 1960s and, since that time, preclinical and clinical studies have worked to elucidate the role of 5-HT in adult PH. Over the past several decades, accumulating evidence from both clinical and preclinical studies has suggested that the 5-HT signaling pathway may play an important role in neonatal cardiopulmonary transition and the development of PH in newborns. The expression of TPH, SERT, and the 5-HT receptors is developmentally regulated, with alterations resulting in pulmonary vasoconstriction and pulmonary vascular remodeling. However, much remains unknown about the role of 5-HT in the developing and newborn lung. The purpose of this review is to discuss the implications of 5-HT on fetal and neonatal pulmonary circulation and summarize the existing preclinical and clinical literature on 5-HT in neonatal PH.

Waitlist-controlled trial of an online intervention to address mental health among older people living with HIV.

Background: Older people living with HIV (PLWH) often experience elevated levels of depression, anxiety, and loneliness.Methods: This waitlist-controlled trial examined the effectiveness of online audio mindfulness lessons in impacting these feelings among older PLWH.Results: Among 214 participants, the mean (SD) age was 60.4 (5.9) years, 89% were male, and 69% were white. After 25 days, the intervention group showed significant improvements versus the waitlist control group in symptoms of depression (20.3% improvement, p < .01) and symptoms of anxiety (22.4% improvement, p = .03), but not in loneliness as measured by a Daily Diary (12.9% improvement, p = .07) or the 3-Item Loneliness Scale (4.8% improvement, p = .27). Secondary analyses among participants with elevated baseline symptoms of depression showed a 26.3% improvement (p < .01), with a moderate effect size (Hedge's g = 0.69). Similarly, those with elevated baseline symptoms of anxiety showed a 25.6% improvement (p < .01), a moderate effect size (g = 0.54), while those with moderate or severely elevated loneliness showed an 18.9% improvement in daily loneliness (p < .01), a moderate effect size (g = 0.55).Conclusion: This waitlist-controlled trial is the first to show that a series of brief, online audio mindfulness lessons improves mental health outcomes among older PLWH. For many patients, this intervention may offer relief that is both accessible and affordable.

Vestibular rehabilitation in cerebellar ataxia with neuropathy and vestibular areflexia syndrome (CANVAS)- A case report.

Background and purpose: Cerebellar ataxia, neuropathy, and vestibular areflexia syndrome (CANVAS) is a neurodegenerative disease of the cerebellum. The disease progression is slow, with up to 25% of people diagnosed needing to use a wheelchair after 15 years from diagnosis. Vestibular symptoms arise from centrally-mediated ocular movement degradation and the reduced vestibular-ocular reflex functioning bilaterally. To date, no report has shown an improvement in VOR gain or gait outcome measures in someone with CANVAS after a course of vestibular physical therapy. Case description: A 65-year-old male, Patient X, first noticed symptoms in his fourth decade of life and was diagnosed with (CANVAS) in his seventh decade. Patient X reported numbness and tingling in his hands and feet, decreased ability to perform daily activities, and several falls. Intervention: Patient X completed a four-month course of vestibular physical therapy, including vestibular ocular reflex exercises, balance training, gait training, and the VestAid application for eye gaze compliance monitoring. The Vestaid application uses eyes and facial recognition software to record the percentage of time that the patient kept their eyes on the target. Outcomes: After vestibular therapy, Patient X had a clinically meaningful improvement in gait speed: from 1.02 m/s to 1.13 m/s and in the Functional Gait Assessment from 20/30 to 27/30. Patient X's eye gaze compliance improved from a median of 43% (range 25-68%) to a median of 67% (58-83%). Discussion: This case study demonstrates that vestibular rehabilitation improved eye gaze compliance and functional outcomes in a person living with CANVAS.

Deinterpenetration of IRMOF-9.

One of the iconic characteristics of metal-organic frameworks (MOFs) is the possesssion of guest-accessible pores. Increasing pore size has a direct and often beneficial impact on a MOF's adsorption and separation properties. However, as pore size increases, the resulting void spaces are often filled by interpenetrated frameworks, where one or more networks crystallize within the pore system of another identical network, reducing the MOF's free volume and pore size. Furthermore, due to the thermodynamic favorability of interpenetration during solvothermal synthesis, techniques to synthetically differentiate interpenetrated from non-interpenetrated MOFs are paramount. This study reports the synthesis of deinterpenetrated IRMOF-9 via halide mediated deinterpenetrative conversion of Zn4 O-derived IRMOF-9. IRMOF-9, when treated with ethylammonium bromide, is quasi-selectively etched, revealing the non-interpenetrated analogue, IRMOF-10 (deinterpenetrated IRMOF-9), which can be isolated prior to complete dissolution by the bromide solution. Dye adsorption, surface area and pore size distribution analysis, and powder X-ray diffraction are consistent with successful deinterpenetration.

Implementation Science and Practice-Oriented Research: Convergence and Complementarity.

Implementation science is the scientific study of methods to promote the uptake of research findings and other evidence-based practices in routine care, with the goal of improving the quality and effectiveness of health services (Bauer et al., 2015). In addition to this common goal, practice-oriented psychotherapy research (and researchers) and implementation science (and scientists) share a common focus on the people and the places where treatment happens. Thus, there exists strong potential for combining these two approaches. In this article, we provide a primer on implementation science for psychotherapy researchers and highlight important areas and examples of convergence and complementarity between implementation science and practice-oriented psychotherapy research. Specifically, we (a) define and describe the core features of implementation science; (b) discuss similarities and areas of complementarity between implementation science and practice-oriented psychotherapy research; (c) discuss a case example that exemplifies the integration of implementation science and practice-oriented research; and (d) propose directions for future research and collaborations that leverage both implementation science and practice-oriented research.

Agricultural Use of Insecticides Alters Homeostatic Behaviors and Cognitive Ability in Lymnaea stagnalis.

Lymnaea stagnalis is an ecologically important, stress-sensitive, freshwater mollusk that is at risk for exposure to insecticides via agricultural practices. We provide insight into the impact insecticides have on L. stagnalis by comparing specific behaviors including feeding, locomotion, shell regeneration, and cognition between snails collected at two different sites: one contaminated by insecticides and one not. We hypothesized that each of the behaviors would be altered in the insecticide-exposed snails and that similar alterations would be induced when control snails were exposed to the contaminated environment. We found no significant differences in locomotion, feeding, and shell regeneration of insecticide-exposed L. stagnalis compared with nonexposed individuals. Significant changes in feeding and shell repair were observed in nonexposed snails inhabiting insecticide-contaminated pond water. Most importantly, snails maintained and trained in insecticide-contaminated pond water did not form configural learning, but this cognitive deficit was reversed when these snails were maintained in insecticide-free pond water. Our findings conclude that insecticides have a primarily negative impact on this higher form of cognition in L. stagnalis. Environ Toxicol Chem 2023;42:2466-2477. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Platelets and ductus arteriosus closure in neonates.

Platelet plug formation is critically involved in murine ductus arteriosus closure and thrombocytopenia in preterm infants seems to negatively affect spontaneous and pharmacologically induced ductal closure. Furthermore, platelet dysfunction may contribute to ductal patency, especially in extremely immature infants. Neonatal platelets likely have multifaceted roles during ductal closure, such as secretion of several signaling molecules and facilitation of specific cell-cell interactions. The only available randomized-controlled trial on platelet transfusions in preterm infants with patent ductus arteriosus demonstrated that a liberal transfusion regimen did not promote ductal closure, but was associated with an increased rate of intraventricular hemorrhage. Herein, we discuss the available mechanistic evidence on the role of platelets in ductus arteriosus closure and their potential clinical implications in preterm infants. We further briefly outline future research directions aimed at a better understanding of platelet-endothelial interactions in neonatal health and disease.

Serotonin-deficient neonatal mice are not protected against the development of experimental bronchopulmonary dysplasia or pulmonary hypertension.

Serotonin (5-hydroxytryptamine, 5-HT) is a potent pulmonary vasoconstrictor and contributes to high pulmonary vascular resistance in the developing ovine lung. In experimental pulmonary hypertension (PH), pulmonary expression of tryptophan hydroxylase-1 (TPH1), the rate limiting enzyme in 5-HT synthesis, and plasma 5-HT are increased. 5-HT blockade increases pulmonary blood flow and prevents pulmonary vascular remodeling and PH in neonatal models of PH with bronchopulmonary dysplasia (BPD). We hypothesized that neonatal tph1 knock-out (KO) mice would be protected from hypoxia-induced alveolar simplification, decreased vessel density, and PH. Newborn wild-type (WT) and tph1 KO mice were exposed to normoxia or hypoxia for 2 weeks. Normoxic WT and KO mice exhibited similar alveolar development, pulmonary vascular density, right ventricular systolic pressures (RVSPs), and right heart size. Circulating (plasma and platelet) 5-HT decreased in both hypoxia-exposed WT and KO mice. Tph1 KO mice were not protected from hypoxia-induced alveolar simplification, decreased pulmonary vascular density, or right ventricular hypertrophy, but displayed attenuation to hypoxia-induced RVSP elevation compared with WT mice. Tph1 KO neonatal mice are not protected against hypoxia-induced alveolar simplification, reduction in pulmonary vessel density, or RVH. While genetic and pharmacologic inhibition of tph1 has protective effects in adult models of PH, our results suggest that tph1 inhibition would not be beneficial in neonates with PH associated with BPD.

A pilot randomized effectiveness trial of the unified protocol in trauma-exposed veterans.

BACKGROUND: The unified protocol (UP) is a promising transdisgnostic treatment for emotional disorders; limited data exists with trauma-exposed populations. This study compared effectiveness of the UP, presented centered therapy (PCT), and treatment as usual (TAU) in trauma-exposed veterans presenting to routine care. METHOD: Trauma-exposed veterans with one or more emotional disorder diagnoses participated in a pilot hybrid-1 effectiveness/preimplementation study. Thirty-seven male and female veterans were randomized to one of three conditions. RESULTS: Multilevel growth curve modeling demonstrated improvement over time across conditions with large effect sizes (range: -2.15 to -3.32), with the UP demonstrating the greatest change. The between group effect sizes for reductions in number of comorbid diagnoses were medium to small and statistically significant (TAU and UP, d = 0.49, p = .056; TAU and PCT d = 0.18, p = .166, UP and PCT d = 0.31, p = .229). Only the UP led to a decrease in the number of comorbid diagnoses (d = -0.71). Psychosocial functioning varied by group, with slight increases in impairment in PCT and TAU, and medium effect size reduction in the UP. Only the UP exhibited significant decreases in self-reported anxiety and depression. Between group differences for UP and PCT were medium to large and statistically significant for depression across two measures (d = -0.72 to d = -1.40). CONCLUSIONS: This represents the first trial examining effectiveness of the UP, PCT, and TAU in trauma-exposed veterans. Despite a small sample, large effect size differences demonstrated promising advantages for the UP. Trial Registration Number: NCT02944994.

Growth of the harmful benthic cyanobacterium Microseira wollei is driven by legacy sedimentary phosphorous.

Models for cyanobacterial harmful algae blooms (cHABs) in fresh waters are usually predicated on the relationship between cyanobacterial ecology and dissolved nutrients, particularly phosphorous. Here we show legacy sediment-associated phosphorous as the primary driver of a benthic cHAB, not phosphorous in the water column. Biogeographical surveys by teams of citizen science volunteers working with the University of South Carolina identified over 200 distinct mats of Microseira wollei in Lake Wateree, SC based on toxin characterization. In sum these were estimated to affect approximately 175 km of the lake's shoreline. This growth occurred under water quality conditions that were near or below the regulatory total maximum daily load for phosphorous and nitrogen. A series of established predictive models for cyanobacterial biomass growth were applied retroactively to match the measured growth with measured water quality parameters. The only component of the system that successfully predicted microbial biomass was sedimentary phosphorous. Concentrations of the Lyngbya wollei toxins (LWTs) 1, 4, 5, and 6 were determined at multiple sites over an 18-month period and a toxin inventory for the lake was calculated. Toxin profiles between sites differed at the 95% level of confidence, establishing each site as a unique mat. An empirical model of toxin production potential based on sedimentary phosphorous was developed.

A scoping review of frameworks in empirical studies and a review of dissemination frameworks.

BACKGROUND: The field of dissemination and implementation (D&I) research has grown immensely in recent years. However, the field of dissemination research has not coalesced to the same degree as the field of implementation research. To advance the field of dissemination research, this review aimed to (1) identify the extent to which dissemination frameworks are used in dissemination empirical studies, (2) examine how scholars define dissemination, and (3) identify key constructs from dissemination frameworks. METHODS: To achieve aims 1 and 2, we conducted a scoping review of dissemination studies published in D&I science journals. The search strategy included manuscripts published from 1985 to 2020. Articles were included if they were empirical quantitative or mixed methods studies about the dissemination of information to a professional audience. Studies were excluded if they were systematic reviews, commentaries or conceptual papers, scale-up or scale-out studies, qualitative or case studies, or descriptions of programs. To achieve aim 1, we compiled the frameworks identified in the empirical studies. To achieve aim 2, we compiled the definitions from dissemination from frameworks identified in aim 1 and from dissemination frameworks identified in a 2021 review (Tabak RG, Am J Prev Med 43:337-350, 2012). To achieve aim 3, we compile the constructs and their definitions from the frameworks. FINDINGS: Out of 6017 studies, 89 studies were included for full-text extraction. Of these, 45 (51%) used a framework to guide the study. Across the 45 studies, 34 distinct frameworks were identified, out of which 13 (38%) defined dissemination. There is a lack of consensus on the definition of dissemination. Altogether, we identified 48 constructs, divided into 4 categories: process, determinants, strategies, and outcomes. Constructs in the frameworks are not well defined. IMPLICATION FOR D&I RESEARCH: This study provides a critical step in the dissemination research literature by offering suggestions on how to define dissemination research and by cataloging and defining dissemination constructs. Strengthening these definitions and distinctions between D&I research could enhance scientific reproducibility and advance the field of dissemination research.

Diastereoselective and Enantioselective Synthesis of α-p-Methoxyphenoxy-ß-Lactones: Dependence on the Stereoelectronic Properties of the ß-Hydroxy-α-p-Methoxyphenoxycarboxylic Acid Precursors.

Treatment of ß-hydroxy-α-p-methoxyphenoxy carboxylic acids derived from the asymmetric glycolate aldol addition reaction with p-nitrobenzenesulfonyl chloride yielded divergent results depending on the nature of the ß-substituent of the carboxylic acid. Substrates bearing either alkyl substituents (R = -n-butyl, -n-octyl, -benzyl, isopropyl, -tert-butyl) or aryl systems bearing electron-withdrawing substituents (R = -p-C6H4Cl, -p-C6H4Br, -p-C6H4NO2) yielded ß-lactones. In contrast, α-p-methoxyphenoxy-ß-hydroxycarboxylic acids bearing electron-donating aryl groups or the sterically demanding 2-naphthyl group formed (Z)-alkenes.

Barriers to participation and lifestyle change among lower versus higher income participants in the National Diabetes Prevention Program: lifestyle coach perspectives.

Individuals from socioeconomically disadvantaged groups have lesser participation and success in the National Diabetes Prevention Program (NDPP). Barriers to NDPP participation and lifestyle change were examined from the perspective of Lifestyle Coaches serving lower versus higher income participants. Lifestyle Coaches (n = 211) who serve lower income (n = 82) or higher income (n = 129) participants reported on observed barriers to NDPP participation and lifestyle change and ranked the three most significant barriers to (a) NDPP participation and (b) lifestyle change. Group differences in number/type of barriers were examined using t-tests and chi-square analyses, and ranking differences were examined using multilevel cumulative logit models. Lifestyle Coaches of lower income (versus higher income) participants reported two additional barriers on average. Ranked barriers to participation were similar between groups, and notably included physical/emotional barriers. However, for lifestyle change, those serving lower income groups were more likely to rank lack of access to healthy grocery stores, but less likely to rank low motivation and lack of family support. Lifestyle Coaches of lower income participants were less likely to rank long wait period prior to enrollment as the most significant barrier to participation, and to rank lack of time off from work as the most significant barrier to lifestyle change. Despite more barriers observed among lower versus higher income participants, overlap in the most significant barriers highlights the potential utility of widely addressing common barriers among NDPP participants. In particular, physical and emotional barriers have been overlooked, yet deserve greater attention in future research and practice.

The National Diabetes Prevention Program (NDPP) has less successfully reached and changed the lifestyles of lower income (versus higher income) adults in the USA who are at high risk for type 2 diabetes. In a nationwide online survey, we asked Lifestyle Coaches who deliver the NDPP to identify up to 37 potential barriers to participation and success that they had observed among their participants. We then compared the number, type, and rankings of the most significant barriers to participation and success in the NDPP from the perspective of Lifestyle Coaches estimating the majority of their participants had lower versus higher incomes. Lifestyle Coaches delivering the NDPP to lower income participants reported an average of two additional barriers to participation and success than those delivering the program to higher income participants. The barriers ranked among the most significant to NDPP participation and lifestyle change were generally similar among Lifestyle Coaches working with lower versus higher income participants. Top-ranked barriers included physical/emotional symptoms (e.g., anxiety, depression) as well as barriers previously reported in studies focused on NDPP participants. It is critical that barriers be carefully evaluated and addressed to improve the nationwide impact of the NDPP.

Promises and Pitfalls of Dyads in the National Diabetes Prevention Program: Lifestyle Coach Perspectives.

PURPOSE: To describe Lifestyle Coach perceptions of dyads (i.e., family members and/or friends) in the National Diabetes Prevention Program (NDPP). DESIGN: Qualitative evaluation of cross-sectional survey responses. SETTING: Online. PARTICIPANTS: Lifestyle Coaches (n=253) with experience teaching at least one in-person year-long NDPP cohort at a CDC-recognized organization. MEASURES: Survey included items on background and experience with dyadic approach, as well as open-ended items on the benefits and challenges observed when working with dyads in the NDPP. ANALYSIS: Lifestyle Coach background and experience were analyzed descriptively in SPSS. Open-ended responses were content coded in ATLAS.ti using qualitative description, and then grouped into categories. RESULTS: Most Lifestyle Coaches (n=210; 83.0%) reported experience delivering the NDPP to dyads. Benefits of a dyadic approach included having a partner in lifestyle change, superior outcomes and increased engagement, and positive "ripple effects." Challenges included difficult relationship dynamics, differences between dyad members, negative "ripple effects," and logistics. CONCLUSION: Lifestyle Coaches described a number of benefits, as well as some challenges, with a dyadic approach to the NDPP. Given the concordance between close others in lifestyle and other risk factors for type 2 diabetes, utilizing a dyadic approach in the NDPP has the potential to increase engagement, improve outcomes, and extend the reach of the program.

Mental health clinician perspectives regarding factors impacting implementation of evidence-based psychotherapies in Veterans Health Administration community-based outpatient clinics.

OBJECTIVE: Uptake of Evidence-Based Psychotherapies (EBPs) by mental health (MH) clinicians, especially in community settings, remains highly variable. This formative pilot study aimed to understand the attitudes and practices of Veterans Health Administration community-based MH clinicians regarding EBPs and to identify multi-level factors that enable and hinder EBP implementation in this unique context. METHODS: Semi-structured interviews were conducted with MH clinicians (N = 40) working in community-based outpatient clinics (CBOCs) in metro/urban (n = 20) and non-metro/rural (n = 20) locations. Interviews were guided by the Consolidated Framework for Implementation Research and were analyzed using rapid content analysis. Results were organized by system-, clinician-, patient-, and innovation-levels. RESULTS: EBPs were consistently perceived as important to delivering quality MH care, with most clinicians having received training in at least one VHA EBP. However, limited EBP training and consultation opportunities, inadequate autonomy to schedule EBP sessions, high and complex caseloads, and feelings of isolation at CBOCs decreased EPB use. Social workers perceived disparities in EBP training access relative to psychologists. Some barriers were more salient in non-metro/rural settings (e.g., patient-level privacy concerns). CONCLUSIONS: Increased EBP training opportunities- particularly for social workers-, greater flexibility over schedules and caseloads, and more mechanisms for consultation and professional development may increase EBP uptake in community-based clinics.

Appetitive Behavior in the Social Transmission of Food Preference Paradigm Predicts Activation of Orexin-A producing Neurons in a Sex-Dependent Manner.

Orexin-producing cells in the lateral hypothalamic area have been shown to be involved in a wide variety of behavioral and cognitive functions, including the recall of appetitive associations and a variety of social behaviors. Here, we investigated the role of orexin in the acquisition and recall of socially transmitted food preferences in the rat. Rats were euthanized following either acquisition, short-term recall, or long-term recall of a socially transmitted food preference and their brains were processed for orexin-A and c-Fos expression. We found that while there were no significant differences in c-Fos expression between control and experimental subjects at any of the tested timepoints, females displayed significantly more activity in both orexinergic and non-orexinergic cells in the lateral hypothalamus. In the infralimbic cortex, we found that social behavior was significantly predictive of c-Fos expression, with social behaviors related to olfactory exploration appearing to be particularly influential. We additionally found that appetitive behavior was significantly predictive of orexin-A activity in a sex-dependent matter, with the total amount eaten correlating negatively with orexin-A/c-Fos colocalization in male rats but not female rats. These findings suggest a potential sex-specific role for the orexin system in balancing the stimulation of feeding behavior with the sleep/wake cycle.

Altering Perceived Context: Transportation Cues Influence Novelty-Induced Context Exploration.

Context is the milieu in which everything occurs. Many research studies consider context, or even explicitly manipulate it; yet it remains challenging to characterize. We know that a context surrounds and influences tasks; however, the boundaries of its influence are difficult to define. In behavioral science, context is often operationalized by the physical environment in which the experiment takes place, and the boundaries of the context are assumed to begin at the entrance to that of the room or apparatus. Experiences during transportation to the testing space have been shown to impact rodent behavior and memory, but transportation's relationship with novelty and physical environment is not fully understood. The current study explored how familiar vs. novel cues, both within a physical environment and preceding it, impact the perception of a context. We manipulated context on three levels: physical testing environment, object cues within that environment, and transportation cues preceding entrance to the testing environment. We found that novel transportation cues can change rats' perception of both familiar and novel contexts. The effects of transportation on perceived context may be affected by the length of the retention interval, testing environment, and behavioral range. These data suggest that context is a broad concept that includes cues across time and is sensitive to small differences in experience.

A Novel Approach to the Assessment of Fidelity to a Cognitive Behavioral Therapy for PTSD Using Clinical Worksheets: A Proof of Concept With Cognitive Processing Therapy.

Fidelity monitoring is a critical indicator of psychotherapy quality and is central to successful implementation. A major barrier to fidelity in routine care is the lack of feasible, scalable, and valid measurement strategies. A reliable, low-burden fidelity assessment would promote sustained implementation of cognitive behavioral therapies (CBTs). The current study examined fidelity measurement for cognitive processing therapy (CPT) for posttraumatic stress disorder (PTSD) using clinical worksheets. External raters evaluated patient worksheets done as a part of treatment, both guided by the therapist and completed independently as homework. Results demonstrated that fidelity ratings from CPT session worksheets were feasible and efficient. Notably, they were strongly correlated with observer ratings of the fidelity of CPT strategies that were present on the worksheets. Agreement among ratings conducted by individuals with a range of experience with CPT was acceptable to high. There was not a main effect of therapist-guided, in-session worksheet ratings on PTSD symptom change. However, patient competence in completing worksheets independently was associated with greater PTSD symptom decline and in-session, therapist-guided worksheet completion was associated with larger symptom decreases among patients with high levels of competence. With further research and refinement, rating of worksheets may be an efficient way to examine therapist and patient skill in key CPT elements, and their interactions, compared to the gold standard of observer ratings of therapy video-recordings. Additional research is needed to determine if worksheets are an accurate and scalable alternative to gold standard observer ratings in settings in which time and resources are limited.

Impact of DNA Adduct Size, Number, and Relative Position on the Toxicity of Aromatic Amines: A Molecular Dynamics Case Study of ANdG- and APdG-Containing DNA Duplexes.

Human exposure to aromatic amines (AAs) can result in carcinogenic DNA adducts. To complement previous work geared toward understanding the mutagenicity of AA-derived adducts, which has almost exclusively studied (monoadducted) DNA containing a single lesion, the present work provides the first in-depth comparison of the structure of monoadducted and diadducted DNA duplexes. Specifically, molecular dynamics (MD) simulations were initially performed on DNA containing the nonmutagenic single-ringed N-(deoxyguanosin-8-yl)-aniline (ANdG) or the mutagenic four-ringed N-(deoxyguanosin-8-yl)-1-aminopyrene (APdG) lesion at G1, G2, or G3 in the AA deletion hotspot (5'-G1G2CG3CC) in the anti or syn glycosidic orientation (B/S duplex conformation). Subsequently, diadducted strands were assessed that span each combination of damaged sites (G1G2 (nearest neighbors), G2G3 (next-nearest neighbors), and G1G3 (two intervening nucleotides)) and anti/syn lesion glycosidic orientations. Despite other N-linked C8-dG adducts exhibiting sequence dependence conformational heterogeneity, a single ANdG or APdG lesion induces helical conformational homogeneity that is exclusively controlled by aryl moiety size. However, the preferred damaged DNA conformation can change upon the addition of a second adduct depending on lesion separation, with neighboring lesions stabilizing a nonmutagenic conformation and next-nearest damaged sites stabilizing a promutagenic conformation regardless of adduct size. As a result, diadducted DNA is found to adopt conformations that are unfavored for the corresponding monoadducted system, pointing to differential replication and repair outcomes for diadducted DNA compared to those for monoadducted DNA. Thus, although the toxicity of monoadducted DNA is most significantly dictated by lesion size, the toxicity can increase or decrease upon a second damaging event depending on lesion size and relative position. Overall, our work adds the number of lesions and their spatial separation to the growing list of factors that determine the structure and biological outcomes of adducted DNA.