Towards using 3D cellular cultures to model the activation and diverse functions of macrophages.

The advent of 3D cell culture technology promises to enhance understanding of cell biology within tissue microenvironments. Whilst traditional cell culturing methods have been a reliable tool for decades, they inadequately portray the complex environments in which cells inhabit in vivo. The need for better disease models has pushed the development of effective 3D cell models, providing more accurate drug screening assays. There has been great progress in developing 3D tissue models in fields such as cancer research and regenerative medicine, driven by desires to recreate the tumour microenvironment for the discovery of new chemotherapies, or development of artificial tissues or scaffolds for transplantation. Immunology is one field that lacks optimised 3D models and the biology of tissue resident immune cells such as macrophages has yet to be fully explored. This review aims to highlight the benefits of 3D cell culturing for greater understanding of macrophage biology. We review current knowledge of macrophage interactions with their tissue microenvironment and highlight the potential of 3D macrophage models in the development of more effective treatments for disease.

Leukocyte Tetraspanin CD53 Restrains α3 Integrin Mobilization and Facilitates Cytoskeletal Remodeling and Transmigration in Mice.

The importance of tetraspanin proteins in regulating migration has been demonstrated in many diverse cellular systems. However, the function of the leukocyte-restricted tetraspanin CD53 remains obscure. We therefore hypothesized that CD53 plays a role in regulating leukocyte recruitment and tested this hypothesis by examining responses of CD53-deficient mice to a range of inflammatory stimuli. Deletion of CD53 significantly reduced neutrophil recruitment to the acutely inflamed peritoneal cavity. Intravital microscopy revealed that in response to several inflammatory and chemotactic stimuli, absence of CD53 had only minor effects on leukocyte rolling and adhesion in postcapillary venules. In contrast, Cd53-/- mice showed a defect in leukocyte transmigration induced by TNF, CXCL1 and CCL2, and a reduced capacity for leukocyte retention on the endothelial surface under shear flow. Comparison of adhesion molecule expression in wild-type and Cd53-/- neutrophils revealed no alteration in expression of ß2 integrins, whereas L-selectin was almost completely absent from Cd53-/- neutrophils. In addition, Cd53-/- neutrophils showed defects in activation-induced cytoskeletal remodeling and translocation to the cell periphery, responses necessary for efficient transendothelial migration, as well as increased α3 integrin expression. These alterations were associated with effects on inflammation, so that in Cd53-/- mice, the onset of neutrophil-dependent serum-induced arthritis was delayed. Together, these findings demonstrate a role for tetraspanin CD53 in promotion of neutrophil transendothelial migration and inflammation, associated with CD53-mediated regulation of L-selectin expression, attachment to the endothelial surface, integrin expression and trafficking, and cytoskeletal function.

Tetraspanin CD53 modulates lymphocyte trafficking but not systemic autoimmunity in Lyn-deficient mice.

The leukocyte-restricted tetraspanin CD53 has been shown to promote lymphocyte homing to lymph nodes (LNs) and myeloid cell recruitment to acutely inflamed peripheral organs, and accelerate the onset of immune-mediated disease. However, its contribution in the setting of chronic systemic autoimmunity has not been investigated. We made use of the Lyn-/- autoimmune model, generating Cd53-/- Lyn-/- mice, and compared trafficking of immune cells into secondary lymphoid organs and systemic autoimmune disease development with mice lacking either gene alone. Consistent with previous observations, absence of CD53 led to reduced LN cellularity via reductions in both B and T cells, a phenotype also observed in Cd53-/- Lyn-/- mice. In some settings, Cd53-/- Lyn-/- lymphocytes showed greater loss of surface L-selectin and CD69 upregulation above that imparted by Lyn deficiency alone, indicating that absence of these two proteins can mediate additive effects in the immune system. Conversely, prototypical effects of Lyn deficiency including splenomegaly, plasma cell expansion, elevated serum immunoglobulin M and anti-nuclear antibodies were unaffected by CD53 deficiency. Furthermore, while Lyn-/- mice developed glomerular injury and showed elevated glomerular neutrophil retention above than that in wild-type mice, absence of CD53 in Lyn-/- mice did not alter these responses. Together, these findings demonstrate that while tetraspanin CD53 promotes lymphocyte trafficking into LNs independent of Lyn, it does not make an important contribution to development of autoimmunity, plasma cell dysfunction or glomerular injury in the Lyn-/- model of systemic autoimmunity.

The dendritic cell receptor Clec9A binds damaged cells via exposed actin filaments.

The immune system must distinguish viable cells from cells damaged by physical and infective processes. The damaged cell-recognition molecule Clec9A is expressed on the surface of the mouse and human dendritic cell subsets specialized for the uptake and processing of material from dead cells. Clec9A recognizes a conserved component within nucleated and nonnucleated cells, exposed when cell membranes are damaged. We have identified this Clec9A ligand as a filamentous form of actin in association with particular actin-binding domains of cytoskeletal proteins. We have determined the crystal structure of the human CLEC9A C-type lectin domain and propose a functional dimeric structure with conserved tryptophans in the ligand recognition site. Mutation of these residues ablated CLEC9A binding to damaged cells and to the isolated ligand complexes. We propose that Clec9A provides targeted recruitment of the adaptive immune system during infection and can also be utilized to enhance immune responses generated by vaccines.

Exploring the Interactions between Non-Medical Methamphetamine Use and Prescribed Buprenorphine or Naltrexone in Opioid Use Disorder Treatment Retention.

OBJECTIVES: Our objectives were to examine the impact of methamphetamine use on opioid use disorder (OUD) treatment retention in patients prescribed either buprenorphine/buprenorphine-naloxone (BUP-NX) or naltrexone/extended-release naltrexone (XR-NTX), while also exploring the role of other risk factors that may modify the impact of methamphetamine use. METHODS: We conducted an exploratory retrospective study examining OUD treatment retention in 127 patients in Ohio (USA). Patients were prescribed either BUP-NX or naltrexone/XR-NTX. Cox proportional hazard regression was used to compare time to dropout of treatment between patients positive and negative on screening for methamphetamines at intake, estimate the association between other risk factors and time to dropout, and test interactions between risk factors and methamphetamine status. RESULTS: Among patients prescribed naltrexone/XR-NTX, those positive for methamphetamines had almost three times the risk of treatment dropout (AHR = 2.89, 95% CI =1.11, 7.07), significantly greater (interaction p = .039) than the methamphetamine effect among those prescribed BUP-NX (AHR = 0.94, 95% CI = 0.51, 1.65). Early in treatment, being prescribed BUP-NX was strongly associated with a greater risk of treatment dropout (at baseline: AHR = 2.90, 95% CI = 1.33, 7.15), regardless of baseline methamphetamine use status. However, this effect decreased with time and shifted to greater risk of dropout among those prescribed naltrexone/XR-NTX (non-proportional hazard; interaction with time AHR = 0.66, 95% CI = 0.49, 0.86), with the shift occurring sooner among those positive for methamphetamine at baseline. CONCLUSIONS: Additional support should be provided to patients who use methamphetamines prior to starting OUD treatment.

Dendritic Cell Migration and Antigen Presentation Are Coordinated by the Opposing Functions of the Tetraspanins CD82 and CD37.

This study supports a new concept where the opposing functions of the tetraspanins CD37 and CD82 may coordinate changes in migration and Ag presentation during dendritic cell (DC) activation. We have previously published that CD37 is downregulated upon monocyte-derived DC activation, promotes migration of both skin and bone marrow-derived dendritic cells (BMDCs), and restrains Ag presentation in splenic and BMDCs. In this article, we show that CD82, the closest phylogenetic relative to CD37, appears to have opposing functions. CD82 is upregulated upon activation of BMDCs and monocyte-derived DCs, restrains migration of skin and BMDCs, supports MHC class II maturation, and promotes stable interactions between T cells and splenic DCs or BMDCs. The underlying mechanism involves the rearrangement of the cytoskeleton via a differential activation of small GTPases. Both CD37(-/-) and CD82(-/-) BMDCs lack cellular projections, but where CD37(-/-) BMDCs spread poorly on fibronectin, CD82(-/-) BMDCs are large and spread to a greater extent than wild-type BMDCs. At the molecular level, CD82 is a negative regulator of RhoA, whereas CD37 promotes activation of Rac-1; both tetraspanins negatively regulate Cdc42. Thus, this study identifies a key aspect of DC biology: an unactivated BMDC is CD37(hi)CD82(lo), resulting in a highly motile cell with a limited ability to activate naive T cells. By contrast, a late activated BMDC is CD37(lo)CD82(hi), and thus has modified its migratory, cytoskeletal, and Ag presentation machinery to become a cell superbly adapted to activating naive T cells.

Macrophage heterogeneity and renin-angiotensin system disorders.

Macrophages are heterogeneous innate immune cells which are important in both the maintenance of tissue homeostasis and its disruption, by promoting tissue inflammation and fibrosis. The renin-angiotensin system is central to the pathophysiology of a large suite of diseases, which are driven in part by large amounts of tissue inflammation and fibrosis. Here, we review recent advances in understanding macrophage heterogeneity in origin and function, and how these may lead to new insights into the pathogenesis of these chronic diseases.

New role for the (pro)renin receptor in T-cell development.

The (pro)renin receptor (PRR) was originally thought to be important for regulating blood pressure via the renin-angiotensin system. However, it is now emerging that PRR has instead a generic role in cellular development. Here, we have specifically deleted PRR from T cells. T-cell-specific PRR-knockout mice had a significant decrease in thymic cellularity, corresponding with a 100-fold decrease in the number of CD4(+) and CD8(+) thymocytes, and a large increase in double-negative (DN) precursors. Gene expression analysis on sorted DN3 thymocytes indicated that PRR-deficient thymocytes have perturbations in key cellular pathways essential at the DN3 stage, including transcription and translation. Further characterization of DN T-cell progenitors leads us to propose that PRR deletion affects thymocyte survival and development at multiple stages; from DN3 through to DN4, double-positive, and single-positive CD4 and CD8. Our study thus identifies a new role for PRR in T-cell development.

Tetraspanin CD37 Regulates ß2 Integrin-Mediated Adhesion and Migration in Neutrophils.

Deciphering the molecular basis of leukocyte recruitment is critical to the understanding of inflammation. In this study, we investigated the contribution of the tetraspanin CD37 to this key process. CD37-deficient mice showed impaired neutrophil recruitment in a peritonitis model. Intravital microscopic analysis indicated that the absence of CD37 impaired the capacity of leukocytes to follow a CXCL1 chemotactic gradient accurately in the interstitium. Moreover, analysis of CXCL1-induced leukocyte-endothelial cell interactions in postcapillary venules revealed that CXCL1-induced neutrophil adhesion and transmigration were reduced in the absence of CD37, consistent with a reduced capacity to undergo ß2 integrin-dependent adhesion. This result was supported by in vitro flow chamber experiments that demonstrated an impairment in adhesion of CD37-deficient neutrophils to the ß2 integrin ligand, ICAM-1, despite the normal display of high-affinity ß2 integrins. Superresolution microscopic assessment of localization of CD37 and CD18 in ICAM-1-adherent neutrophils demonstrated that these molecules do not significantly cocluster in the cell membrane, arguing against the possibility that CD37 regulates ß2 integrin function via a direct molecular interaction. Moreover, CD37 ablation did not affect ß2 integrin clustering. In contrast, the absence of CD37 in neutrophils impaired actin polymerization, cell spreading and polarization, dysregulated Rac-1 activation, and accelerated ß2 integrin internalization. Together, these data indicate that CD37 promotes neutrophil adhesion and recruitment via the promotion of cytoskeletal function downstream of integrin-mediated adhesion.

Tetraspanin CD37 contributes to the initiation of cellular immunity by promoting dendritic cell migration.

Previous studies on the role of the tetraspanin CD37 in cellular immunity appear contradictory. In vitro approaches indicate a negative regulatory role, whereas in vivo studies suggest that CD37 is necessary for optimal cellular responses. To resolve this discrepancy, we studied the adaptive cellular immune responses of CD37(-/-) mice to intradermal challenge with either tumors or model antigens and found that CD37 is essential for optimal cell-mediated immunity. We provide evidence that an increased susceptibility to tumors observed in CD37(-/-) mice coincides with a striking failure to induce antigen-specific IFN-γ-secreting T cells. We also show that CD37 ablation impairs several aspects of DC function including: in vivo migration from skin to draining lymph nodes; chemo-tactic migration; integrin-mediated adhesion under flow; the ability to spread and form actin protrusions and in vivo priming of adoptively transferred naïve T cells. In addition, multiphoton microscopy-based assessment of dermal DC migration demonstrated a reduced rate of migration and increased randomness of DC migration in CD37(-/-) mice. Together, these studies are consistent with a model in which the cellular defect that underlies poor cellular immune induction in CD37(-/-) mice is impaired DC migration.

The dualistic role of Lyn tyrosine kinase in immune cell signaling: implications for systemic lupus erythematosus.

Systemic lupus erythematosus (SLE, lupus) is a debilitating, multisystem autoimmune disease that can affect any organ in the body. The disease is characterized by circulating autoantibodies that accumulate in organs and tissues, which triggers an inflammatory response that can cause permanent damage leading to significant morbidity and mortality. Lyn, a member of the Src family of non-receptor protein tyrosine kinases, is highly implicated in SLE as remarkably both mice lacking Lyn or expressing a gain-of-function mutation in Lyn develop spontaneous lupus-like disease due to altered signaling in B lymphocytes and myeloid cells, suggesting its expression or activation state plays a critical role in maintaining tolerance. The past 30 years of research has begun to elucidate the role of Lyn in a duplicitous signaling network of activating and inhibitory immunoreceptors and related targets, including interactions with the interferon regulatory factor family in the toll-like receptor pathway. Gain-of-function mutations in Lyn have now been identified in human cases and like mouse models, cause severe systemic autoinflammation. Studies of Lyn in SLE patients have presented mixed findings, which may reflect the heterogeneity of disease processes in SLE, with impairment or enhancement in Lyn function affecting subsets of SLE patients that may be a means of stratification. In this review, we present an overview of the phosphorylation and protein-binding targets of Lyn in B lymphocytes and myeloid cells, highlighting the structural domains of the protein that are involved in its function, and provide an update on studies of Lyn in SLE patients.

A complementary role for the tetraspanins CD37 and Tssc6 in cellular immunity.

The cooperative nature of tetraspanin-tetraspanin interactions in membrane organization suggests functional overlap is likely to be important in tetraspanin biology. Previous functional studies of the tetraspanins CD37 and Tssc6 in the immune system found that both CD37 and Tssc6 regulate T cell proliferative responses in vitro. CD37(-/-) mice also displayed a hyper-stimulatory dendritic cell phenotype and dysregulated humoral responses. In this study, we characterize "double knockout" mice (CD37(-/-)Tssc6(-/-)) generated to investigate functional overlap between these tetraspanins. Strong evidence for a cooperative role for these two proteins was identified in cellular immunity, where both in vitro T cell proliferative responses and dendritic cell stimulation capacity are significantly exaggerated in CD37(-/-)Tssc6(-/-) mice when compared with single knockout counterparts. Despite these exaggerated cellular responses in vitro, CD37(-/-)Tssc6(-/-) mice are not more susceptible to autoimmune induction. However, in vivo responses to pathogens appear poor in CD37(-/-)Tssc6(-/-) mice, which showed a reduced ability to produce influenza-specific T cells and displayed a rapid onset hyper-parasitemia when infected with Plasmodium yoelii. Therefore, in the absence of both CD37 and Tssc6, immune function is further altered when compared with CD37(-/-) or Tssc6(-/-) mice, demonstrating a complementary role for these two molecules in cellular immunity.

Seeing your partner: Structural elucidation of the first C8 tetraspanin protein.

Tetraspanins are proteins that organize cell membranes via interactions with partner proteins mediated by their large ectodomain. In this issue of Structure, Lipper et al., 2022 have elucidated the structure of the first C8 tetraspanin and expand functional insight into how C8 tetraspanins regulate substrate specificity for the transmembrane protease ADAM10.

Tetraspanin CD82 restrains phagocyte migration but supports macrophage activation.

Phagocytes migrate into tissues to combat infection and maintain tissue homeostasis. As dysregulated phagocyte migration and function can lead to inflammation or susceptibility to infection, identifying molecules that control these processes is critical. Here, we show that the tetraspanin CD82 restrains the migration of neutrophils and macrophages into tissues. Cd82 -/- phagocytes exhibited excessive migration during in vivo models of peritoneal inflammation, superfusion of CXCL1, retinopathy of prematurity, and infection with the protozoan parasite L. mexicana. However, with the latter, while Cd82 -/- macrophages infiltrated infection sites at higher proportions, cutaneous L. mexicana lesions were larger and persisted, indicating a failure to control infection. Analyses of in vitro bone-marrow-derived macrophages showed CD82 deficiency altered cellular morphology, and impaired gene expression and metabolism in response to anti-inflammatory activation. Altogether, this work reveals an important role for CD82 in restraining phagocyte infiltration and mediating their differentiation in response to stimulatory cues.

Tetraspanin CD53 controls T cell immunity through regulation of CD45RO stability, mobility, and function.

T cells depend on the phosphatase CD45 to initiate T cell receptor signaling. Although the critical role of CD45 in T cells is established, the mechanisms controlling function and localization in the membrane are not well understood. Moreover, the regulation of specific CD45 isoforms in T cell signaling remains unresolved. By using unbiased mass spectrometry, we identify the tetraspanin CD53 as a partner of CD45 and show that CD53 controls CD45 function and T cell activation. CD53-negative T cells (Cd53-/-) exhibit substantial proliferation defects, and Cd53-/- mice show impaired tumor rejection and reduced IFNγ-producing T cells compared with wild-type mice. Investigation into the mechanism reveals that CD53 is required for CD45RO expression and mobility. In addition, CD53 is shown to stabilize CD45 on the membrane and is required for optimal phosphatase activity and subsequent Lck activation. Together, our findings reveal CD53 as a regulator of CD45 activity required for T cell immunity.

Discordance in STING-Induced Activation and Cell Death Between Mouse and Human Dendritic Cell Populations.

Stimulator of Interferon Genes (STING) is a cytosolic sensor of cyclic dinucleotides (CDNs). The activation of dendritic cells (DC) via the STING pathway, and their subsequent production of type I interferon (IFN) is considered central to eradicating tumours in mouse models. However, this contribution of STING in preclinical murine studies has not translated into positive outcomes of STING agonists in phase I & II clinical trials. We therefore questioned whether a difference in human DC responses could be critical to the lack of STING agonist efficacy in human settings. This study sought to directly compare mouse and human plasmacytoid DCs and conventional DC subset responses upon STING activation. We found all mouse and human DC subsets were potently activated by STING stimulation. As expected, Type I IFNs were produced by both mouse and human plasmacytoid DCs. However, mouse and human plasmacytoid and conventional DCs all produced type III IFNs (i.e., IFN-λs) in response to STING activation. Of particular interest, all human DCs produced large amounts of IFN-λ1, not expressed in the mouse genome. Furthermore, we also found differential cell death responses upon STING activation, observing rapid ablation of mouse, but not human, plasmacytoid DCs. STING-induced cell death in murine plasmacytoid DCs occurred in a cell-intrinsic manner and involved intrinsic apoptosis. These data highlight discordance between STING IFN and cell death responses in mouse and human DCs and caution against extrapolating STING-mediated events in mouse models to equivalent human outcomes.

The tetraspanin protein CD37 regulates IgA responses and anti-fungal immunity.

Immunoglobulin A (IgA) secretion by plasma cells in the immune system is critical for protecting the host from environmental and microbial infections. However, the molecular mechanisms underlying the generation of IgA(+) plasma cells remain poorly understood. Here, we report that the B cell-expressed tetraspanin CD37 inhibits IgA immune responses in vivo. CD37-deficient (CD37-/-) mice exhibit a 15-fold increased level of IgA in serum and significantly elevated numbers of IgA(+) plasma cells in spleen, mucosal-associated lymphoid tissue, as well as bone marrow. Analyses of bone marrow chimeric mice revealed that CD37-deficiency on B cells was directly responsible for the increased IgA production. We identified high local interleukin-6 (IL-6) production in germinal centers of CD37-/- mice after immunization. Notably, neutralizing IL-6 in vivo reversed the increased IgA response in CD37-/- mice. To demonstrate the importance of CD37-which can associate with the pattern-recognition receptor dectin-1-in immunity to infection, CD37-/- mice were exposed to Candida albicans. We report that CD37-/- mice are evidently better protected from infection than wild-type (WT) mice, which was accompanied by increased IL-6 levels and C. albicans-specific IgA antibodies. Importantly, adoptive transfer of CD37-/- serum mediated protection in WT mice and the underlying mechanism involved direct neutralization of fungal cells by IgA. Taken together, tetraspanin protein CD37 inhibits IgA responses and regulates the anti-fungal immune response.

Tetraspanins in cellular immunity.

Tetraspanins are a superfamily of integral membrane proteins involved in the organization of microdomains that consist of both cell membrane proteins and cytoplasmic signalling molecules. These microdomains are important in regulating molecular recognition at the cell surface and subsequent signal transduction processes central to the generation of an efficient immune response. Tetraspanins, both immune-cell-specific, such as CD37, and ubiquitously expressed, such as CD81, have been shown to be imp-ortant in both innate and adaptive cellular immunity. This is via their molecular interaction with important immune cell-surface molecules such as antigen-presenting MHC proteins, T-cell co-receptors CD4 and CD8, as well as cytoplasmic molecules such as Lck and PKC (protein kinase C). Moreover, the generation of tetraspanin-deficient mice has enabled the study of these proteins in immunity. A variety of tetraspanins have a role in the regulation of pattern recognition, antigen presentation and T-cell proliferation. Recent studies have also begun to elucidate roles for tetraspanins in macrophages, NK cells (natural killer cells) and granulocytes.

The C-type lectin Clec12A present on mouse and human dendritic cells can serve as a target for antigen delivery and enhancement of antibody responses.

We have cloned the mouse and human C-type lectin Clec12A, expressed both, and produced mAb recognizing both. Mouse Clec12A is highly expressed on splenic CD8(+) dendritic cells (DC) and plasmacytoid DC. A proportion of CD8(-)DC also expresses lower levels of Clec12A, as do monocytes, macrophages, and B cells. Human CLEC12A, like the mouse counterpart, is expressed on blood monocytes and DC, including pDC and BDCA-3(+)DC, the proposed equivalent of mouse CD8(+)DC. To determine whether Ag targeted to Clec12A could induce immune responses, mice were injected with a rat mAb recognizing Clec12A, or a control rat mAb, then production of anti-rat Ig was measured. Anti-Clec12A mAb alone produced only moderate responses, but these were amplified by coinjecting only small amounts of LPS as a DC activation agent. Furthermore, when OVA was conjugated to anti-Clec12A mAb, OVA-specific T cells were induced to proliferate. This Ag presentation to naive T cells was due to targeting conventional DC, because their ablation eliminated T cell activation. The potent Ab responses induced using microgram amounts of anti-Clec12A and minimal amounts of adjuvant demonstrate that this molecule can be used as an Ag-delivery target to enhance Ab responses to vaccines.

The relationship between dissociative symptoms and the medications used in the treatment of opioid use disorder.

Opioid use disorder has long been associated with psychiatric symptoms, including dissociative experiences. Medications used to treat opioid use disorder can potentially impact dissociative symptoms, but the existing literature has not explored this. We examined the relationship between dissociative symptoms and opioid use disorder using the Dissociative Experiences Scale (DES). We studied subjects who were taking prescribed methadone, buprenorphine, or naltrexone for opioid use disorder. We gave the DES, the Patient Health Questionairre-9 (PHQ-9), and the PTSD Checklist for DSM-5 (PCL-5) with Criterion A to subjects in three substance use treatment facilities in Ohio. We conducted Analysis of Variance (ANOVA) and Spearman's Rank Correlations to examine associations between the variables and outcomes. We developed three separate multiple linear regression models. We included 116 participants in our exploratory and naturalistic study. The majority of participants were female (51.7%), white (89.5%), ≤ 40 years of age (64.7%), and taking buprenorphine (55%). The average DES score was 16.1 (standard deviation = 14.9) and we considered 80.9% to have low dissociation (score < 30). Approximately 55% (n = 64) of participants were taking prescribed buprenorphine. Approximately 27% (n = 32) were taking prescribed methadone and approximately 18% (n = 21) were taking prescribed naltrexone (oral or extended release). There was a significant association between opioid medication type and log dissociative symptoms (p = .01). Participants taking prescribed buprenorphine had higher mean log dissociation symptom scores (2.7) compared to those taking prescribed methadone (2.2) and prescribed naltrexone (2.1). Log dissociation symptom scores were significantly associated with last use of any opiates (rs = -0.21; p = .02) and time on medication (rs = -0.228; p = .01). Compared to those taking buprenorphine, those taking both methadone (ß = -0.26; p = .01) and naltrexone (ß = -0.27; p = .006) had significantly lower dissociation scores, controlling for the other variables in the model. Dissociation scores were positively correlated with depression scores (r = 0.45; p < .0001) and with PCL-5 scores (r = 0.51; p < .0001). Our study highlights the importance of diagnosing and monitoring dissociative symptoms in individuals who are taking prescribed medications for opioid use disorder, especially since dissociative symptoms can interfere with substance use treatment.