Identification of human exTreg cells as CD16+CD56+ cytotoxic CD4+ T cells.

In atherosclerosis, some regulatory T (Treg) cells become exTreg cells. We crossed inducible Treg and exTreg cell lineage-tracker mice (FoxP3eGFP-Cre-ERT2ROSA26CAG-fl-stop-fl-tdTomato) to atherosclerosis-prone Apoe-/- mice, sorted Treg cells and exTreg cells and determined their transcriptomes by bulk RNA sequencing (RNA-seq). Genes that were differentially expressed between mouse Treg cells and exTreg cells and filtered for their presence in a human single-cell RNA-sequencing (scRNA-seq) panel identified exTreg cell signature genes as CST7, NKG7, GZMA, PRF1, TBX21 and CCL4. Projecting these genes onto the human scRNA-seq with CITE-seq data identified human exTreg cells as CD3+CD4+CD16+CD56+, which was validated by flow cytometry. Bulk RNA-seq of sorted human exTreg cells identified them as inflammatory and cytotoxic CD4+T cells that were significantly distinct from both natural killer and Treg cells. DNA sequencing for T cell receptor-ß showed clonal expansion of Treg cell CDR3 sequences in exTreg cells. Cytotoxicity was functionally demonstrated in cell killing and CD107a degranulation assays, which identifies human exTreg cells as cytotoxic CD4+T cells.

Immunodominant MHC-II (Major Histocompatibility Complex II) Restricted Epitopes in Human Apolipoprotein B.

BACKGROUND: CD (cluster of differentiation) 4+ T-cell responses to APOB (apolipoprotein B) are well characterized in atherosclerotic mice and detectable in humans. CD4+ T cells recognize antigenic peptides displayed on highly polymorphic HLA (human leukocyte antigen)-II. Immunogenicity of individual APOB peptides is largely unknown in humans. Only 1 HLA-II-restricted epitope was validated using the DRB1*07:01-APOB3036-3050 tetramer. We hypothesized that human APOB may contain discrete immunodominant CD4+ T-cell epitopes that trigger atherosclerosis-related autoimmune responses in donors with diverse HLA alleles. METHODS: We selected 20 APOB-derived peptides (APOB20) from an in silico screen and experimentally validated binding to the most commonly occurring human HLA-II alleles. We optimized a restimulation-based workflow to evaluate antigenicity of multiple candidate peptides in HLA-typed donors. This included activation-induced marker assay, intracellular cytokine staining, IFNγ (interferon gamma) enzyme-linked immunospot and cytometric bead array. High-throughput sequencing revealed TCR (T-cell receptor) clonalities of APOB-reactive CD4+ T cells. RESULTS: Using stringent positive, negative, and crossover stimulation controls, we confirmed specificity of expansion-based protocols to detect CD4+ T cytokine responses to the APOB20 pool. Ex vivo assessment of AIM+CD4+ T cells revealed a statistically significant autoimmune response to APOB20 but not to a ubiquitously expressed negative control protein, actin. Resolution of CD4+ T responses to the level of individual peptides using IFNγ enzyme-linked immunospot led to the discovery of 6 immunodominant epitopes (APOB6) that triggered robust CD4+ T activation in most donors. APOB6-specific responding CD4+ T cells were enriched in unique expanded TCR clonotypes and preferentially expressed memory markers. Cytometric bead array analysis detected APOB6-induced secretion of both proinflammatory and regulatory cytokines. In clinical samples from patients with angiographically verified coronary artery disease, APOB6 stimulation induced higher activation and memory phenotypes and augmented secretion of proinflammatory cytokines TNF (tumor necrosis factor) and IFNγ, compared with patients with low coronary artery disease. CONCLUSIONS: Using 3 cohorts, each with ≈20 donors, we discovered and validated 6 immunodominant, HLA-II-restricted APOB epitopes. The immune response to these APOB epitopes correlated with coronary artery disease severity.

Long COVID following Omicron wave in Eastern India-A retrospective cohort study.

Long coronavirus disease (COVID) or postacute sequelae of coronavirus disease of 2019 (COVID-19) is widely reported but the data of long COVID after infection with the Omicron variant is limited. This study was conducted to estimate the incidence, characteristics of symptoms, and predictors of long COVID among COVID-19 patients diagnosed during the Omicron wave in Eastern India. The cohort of COVID-19 patients included were adults (≥18 years) diagnosed as severe acute respiratory syndrome coronavirus 2 positive with Reverse Transcription Polymerase Chain Reaction. After 28 days of diagnosis; participants were followed up with a telephonic interview to capture data on sociodemographic, clinical history, anthropometry, substance use, COVID-19 vaccination status, acute COVID-19 symptoms, and long COVID symptoms. The long COVID symptoms were self-reported by the participants. Logistic regression was used to determine the predictors of long COVID. The median follow-up of participants was 73 days (Interquartile range; 67-83). The final analysis had 524 participants' data; among them 8.2% (95% Confidence Interval [CI]: 6%-10.9%) self-reported long COVID symptoms. Fatigue (34.9%) was the most common reported symptom followed by cough (27.9%). In multivariable logistic regression only two predictors were statistically significant-number of acute COVID-19 symptoms ≥ five (Adjusted odds ratio (aOR) = 2.95, 95% CI: 1.30-6.71) and past history of COVID-19 (aOR = 2.66, 95% CI: 1.14-6.22). The proportion of self-reported long COVID is considerably low among COVID-19 patients diagnosed during the Omicron wave in Eastern India when compared with estimates during Delta wave in the same setting.

Kindlin-3 recruitment to the plasma membrane precedes high-affinity ß2-integrin and neutrophil arrest from rolling.

Integrin-mediated neutrophil adhesion starts by arrest from rolling. Activation of integrins involves conformational changes from an inactive, bent conformation to an extended conformation (E+) with high affinity for ligand binding (H+). The cytoplasmic protein kindlin-3 is necessary for leukocyte adhesion; mutations of kindlin-3 cause leukocyte adhesion deficiency type 3. Kindlin-3 binds the ß2-integrin cytoplasmic tail at a site distinct from talin-1, but the molecular mechanism by which kindlin-3 activates ß2-integrins is unknown. In this study, we measured the spatiotemporal dynamics of kindlin-3 and ß2-integrin conformation changes during neutrophil and HL-60 cell rolling and arrest under flow. Using high-resolution quantitative dynamic footprinting microscopy and kindlin-3-fluorescent protein (FP) fusion proteins, we found that kindlin-3 was recruited to the plasma membrane in response to interleukin-8 (IL-8) before induction of the H+ ß2-integrin conformation. Intravital imaging revealed that EGFP-kindlin-3-reconstituted, kindlin-3-knockout neutrophils arrest in vivo in response to CXCL1. EGFP-kindlin-3 in primary mouse neutrophils was also recruited to the plasma membrane before arrest. Upon arrest, we found small clusters of high-affinity ß2-integrin molecules within large areas of membrane-proximal kindlin-3 FP. Deletion of kindlin-3 or its pleckstrin homology (PH) domain in neutrophil-like HL-60 cells completely abolished H+ ß2-integrin induction. IL-8 also triggered recruitment of the isolated kindlin-3 PH domain to the plasma membrane before arrest. In summary, we showed that the kindlin-3 PH domain is necessary for recruitment to the plasma membrane, where full-length kindlin-3 is indispensable for the induction of high-affinity ß2-integrin.

Evidence-Based Interventions to Improve Opioid Prescribing in Primary Care: a Qualitative Assessment of Implementation in Two Studies.

BACKGROUND: The TOPCARE and TEACH randomized controlled trials demonstrated the efficacy of a multi-faceted intervention to promote guideline-adherent long-term opioid therapy (LTOT) in primary care settings. Intervention components included a full-time Nurse Care Manager (NCM), an electronic registry, and academic detailing sessions. OBJECTIVE: This study sought to identify barriers, facilitators, and other issues germane to the wider implementation of this intervention. DESIGN: We conducted a nested, qualitative study at 4 primary care clinics (TOPCARE) and 2 HIV primary care clinics (TEACH), where the trials had been conducted. APPROACH: We purposively sampled primary care physicians and advanced practice providers (hereafter: PCPs) who had received the intervention. Semi-structured interviews explored perceptions of the intervention to identify unanticipated barriers to and facilitators of implementation. Interview transcripts were analyzed through iterative deductive and inductive coding exercises. KEY RESULTS: We interviewed 32 intervention participants, 30 physicians and 2 advanced practice providers, who were majority White (66%) and female (63%). Acceptability of the intervention was high, with most PCPs valuing didactic and team-based intervention elements, especially co-management of LTOT patients with the NCM. Adoption of new prescribing practices was facilitated by proximity to expertise, available behavioral health care, and the NCM's support. Most participants were enthusiastic about the intervention, though a minority voiced concerns about the appropriateness in their particular clinical environments, threats to the patient-provider relationship, or long-term sustainability. CONCLUSION: TOPCARE/TEACH participants found the intervention generally acceptable, appropriate, and easy to adopt in a variety of primary care environments, though some challenges were identified. Careful attention to the practical challenges of implementation and the professional relationships affected by the intervention may facilitate implementation and sustainability.

Combined protein and transcript single-cell RNA sequencing in human peripheral blood mononuclear cells.

BACKGROUND: Cryopreserved peripheral blood mononuclear cells (PBMCs) are frequently collected and provide disease- and treatment-relevant data in clinical studies. Here, we developed combined protein (40 antibodies) and transcript single-cell (sc)RNA sequencing (scRNA-seq) in PBMCs. RESULTS: Among 31 participants in the Women's Interagency HIV Study (WIHS), we sequenced 41,611 cells. Using Boolean gating followed by Seurat UMAPs (tool for visualizing high-dimensional data) and Louvain clustering, we identified 50 subsets among CD4+ T, CD8+ T, B, NK cells, and monocytes. This resolution was superior to flow cytometry, mass cytometry, or scRNA-seq without antibodies. Combined protein and transcript scRNA-seq allowed for the assessment of disease-related changes in transcriptomes and cell type proportions. As a proof-of-concept, we showed such differences between healthy and matched individuals living with HIV with and without cardiovascular disease. CONCLUSIONS: In conclusion, combined protein and transcript scRNA sequencing is a suitable and powerful method for clinical investigations using PBMCs.

Inpatient Addiction Medicine Consultation Service Impact on Post-discharge Patient Mortality: a Propensity-Matched Analysis.

BACKGROUND: Inpatient addiction medicine consultation services (AMCS) have grown rapidly, but there is limited research of their impact on patient outcomes. OBJECTIVE: To examine whether AMCS is associated with all-cause mortality and hospital utilization post-discharge. DESIGN: This was a propensity-score-matchedcase-control study from 2018 to 2020. PARTICIPANTS: The intervention group included patients referred to the AMCS from October 2018 to March 2020. Matched control participants included patients hospitalized from October 2017 to September 2018 at an urban academic hospital with a large suburban and rural catchment area. MAIN MEASURES: The effect of treatment was estimated as the difference between the proportion of subjects experiencing the event (7-day and 30-day readmission, emergency department visits, and mortality within 90 days) for each group in the matched sample. KEY RESULTS: There were 711 patients in the intervention group and 2172 patients in the control group. The most common substance use disorders among the intervention group were primary alcohol use disorder (n=181; 25.5%) and primary opioid use disorder (n=175, 24.6%) with over a third with polysubstance use (n=257, 36.1%). Intervention patients showed a reduction in 90-day mortality post-hospital discharge (average treatment effect [ATE]: -2.35%, 95% CI: -3.57, -1.13; p-value <0.001) compared to propensity-matched controls. We found a statistically significant reduction in 7-day hospital readmission by 2.15% (95% CI: -3.65, -0.65; p=0.005) and a nonsignificant reduction in 30-day readmission (ATE: -2.38%, 95% CI: -5.20, 0.45; p=0.099). There was a statistically significant increase in 30-day emergency department visits (ATE: 5.32%, 95% CI: 2.19, 8.46; 0.001) compared to matched controls. CONCLUSIONS: There was a reduction in 90-day all-cause mortality for the AMCS intervention group compared to matched controls, although the impact on hospital utilization was mixed. AMCS are systems interventions that are effective tools to improve patient health and reduce all-cause mortality.

Sex Differences in Coronary Artery Disease and Diabetes Revealed by scRNA-Seq and CITE-Seq of Human CD4+ T Cells.

Despite the decades-old knowledge that males and people with diabetes mellitus (DM) are at increased risk for coronary artery disease (CAD), the reasons for this association are only partially understood. Among the immune cells involved, recent evidence supports a critical role of T cells as drivers and modifiers of CAD. CD4+ T cells are commonly found in atherosclerotic plaques. We aimed to understand the relationship of CAD with sex and DM by single-cell RNA (scRNA-Seq) and antibody sequencing (CITE-Seq) of CD4+ T cells. Peripheral blood mononuclear cells (PBMCs) of 61 men and women who underwent cardiac catheterization were interrogated by scRNA-Seq combined with 49 surface markers (CITE-Seq). CAD severity was quantified using Gensini scores, with scores above 30 considered CAD+ and below 6 considered CAD-. Four pairs of groups were matched for clinical and demographic parameters. To test how sex and DM changed cell proportions and gene expression, we compared matched groups of men and women, as well as diabetic and non-diabetic subjects. We analyzed 41,782 single CD4+ T cell transcriptomes for sex differences in 16 women and 45 men with and without coronary artery disease and with and without DM. We identified 16 clusters in CD4+ T cells. The proportion of cells in CD4+ effector memory cluster 8 (CD4T8, CCR2+ Em) was significantly decreased in CAD+, especially among DM+ participants. This same cluster, CD4T8, was significantly decreased in female participants, along with two other CD4+ T cell clusters. In CD4+ T cells, 31 genes showed significant and coordinated upregulation in both CAD and DM. The DM gene signature was partially additive to the CAD gene signature. We conclude that (1) CAD and DM are clearly reflected in PBMC transcriptomes, and (2) significant differences exist between women and men and (3) between subjects with DM and non-DM.

A TNF-p100 pathway subverts noncanonical NF-κB signaling in inflamed secondary lymphoid organs.

Lymphotoxin-beta receptor (LTßR) present on stromal cells engages the noncanonical NF-κB pathway to mediate RelB-dependent expressions of homeostatic chemokines, which direct steady-state ingress of naïve lymphocytes to secondary lymphoid organs (SLOs). In this pathway, NIK promotes partial proteolysis of p100 into p52 that induces nuclear translocation of the RelB NF-κB heterodimers. Microbial infections often deplete homeostatic chemokines; it is thought that infection-inflicted destruction of stromal cells results in the downregulation of these chemokines. Whether inflammation per se also regulates these processes remains unclear. We show that TNF accumulated upon non-infectious immunization of mice similarly downregulates the expressions of these chemokines and consequently diminishes the ingress of naïve lymphocytes in inflamed SLOs. Mechanistically, TNF inactivated NIK in LTßR-stimulated cells and induced the synthesis of Nfkb2 mRNA encoding p100; these together potently accumulated unprocessed p100, which attenuated the RelB activity as inhibitory IκBδ. Finally, a lack of p100 alleviated these TNF-mediated inhibitions in inflamed SLOs of immunized Nfkb2-/- mice. In sum, we reveal that an inhibitory TNF-p100 pathway modulates the adaptive compartment during immune responses.

Confronting COVID, racism, and addiction: The association of multidisciplinary education and research in substance use and addiction (AMERSA).

The COVID-19 pandemic, the ongoing opioid epidemic, rise in substance use, and social and political unrest in the US and globally has impacted how substance use-related health needs are addressed. These issues were driving forces in planning AMERSA's 44th annual conference. True to the multidisciplinary spirit, and with diversity goals and advocacy at the forefront of mind, "together we rise" became the beacon for the AMERSA 2020 conference. This commentary provides an overview of the conference proceedings, topics that were highly relevant for clinicians, educators, researchers, and advocates for change.

Pain management in patients with chronic kidney disease and end-stage kidney disease.

PURPOSE OF REVIEW: This review evaluates current recommendations for pain management in chronic kidney disease (CKD) and end-stage kidney disease (ESKD) with a specific focus on evidence for opioid analgesia, including the partial agonist, buprenorphine. RECENT FINDINGS: Recent evidence supports the use of physical activity and other nonpharmacologic therapies, either alone or with pharmacological therapies, for pain management. Nonopioid analgesics, including acetaminophen, topical analgesics, gabapentinoids, serotonin-norepinephrine reuptake inhibitors, and TCA may be considered based on pain cause and type, with careful dose considerations in kidney disease. NSAIDs may be used in CKD and ESKD for short durations with careful monitoring. Opioid use should be minimized and reserved for patients who have failed other therapies. Opioids have been associated with increased adverse events in this population, and thus should be used cautiously after risk/benefit discussion with the patient. Opioids that are safer to use in kidney disease include oxycodone, hydromorphone, fentanyl, methadone, and buprenorphine. Buprenorphine appears to be a promising and safer option due to its partial agonism at the mu opioid receptor. SUMMARY: Pain is poorly managed in patients with kidney disease. Nonpharmacological and nonopioid analgesics should be first-line approaches for pain management. Opioid use should be minimized with careful monitoring and dose adjustment.

Disagreement and Uncertainty Among Experts About how to Respond to Marijuana Use in Patients on Long-term Opioids for Chronic Pain: Results of a Delphi Study.

BACKGROUND: Marijuana use is common among patients on long-term opioid therapy (LTOT) for chronic pain, but there is a lack of evidence to guide clinicians' response. OBJECTIVE: To generate expert consensus about responding to marijuana use among patients on LTOT. DESIGN: Analysis from an online Delphi study. SETTING/SUBJECTS: Clinician experts in pain and opioid management across the United States. METHODS: Participants generated management strategies in response to marijuana use without distinction between medical and nonmedical use, then rated the importance of each management strategy from 1 (not at all important) to 9 (extremely important). A priori rules for consensus were established, and disagreement was explored using cases. Thematic analysis of free-text responses examined factors that influenced participants' decision-making. RESULTS: Of 42 participants, 64% were internal medicine physicians. There was consensus that it is not important to taper opioids as an initial response to marijuana use. There was disagreement about the importance of tapering opioids if there is a pattern of repeated marijuana use without clinical suspicion for a cannabis use disorder (CUD) and consensus that tapering is of uncertain importance if there is suspicion for CUD. Three themes influenced experts' perceptions of the importance of tapering: 1) benefits and harms of marijuana for the individual patient, 2) a spectrum of belief about the overall riskiness of marijuana use, and 3) variable state laws or practice policies. CONCLUSIONS: Experts disagree and are uncertain about the importance of opioid tapering for patients with marijuana use. Experts were influenced by patient factors, provider beliefs, and marijuana policy, highlighting the need for further research.

Utilizing a Faculty Development Program to Promote Safer Opioid Prescribing for Chronic Pain in Internal Medicine Resident Practices.

OBJECTIVE: To implement a skills-based faculty development program (FDP) to improve Internal Medicine faculty's clinical skills and resident teaching about safe opioid prescribing. DESIGN: An FDP for Internal Medicine attendings that included a one-hour didactic presentation followed immediately by an Objective Structured Clinical Examination (OSCE) that focused on assessing and managing opioid misuse risk, opioid treatment outcomes (benefits and harms), and aberrant opioid use behaviors. The evaluation compared pre- and three-months-post-FDP changes in faculty's safe opioid prescribing knowledge, attitudes, confidence (clinical and teaching), and self-reported resident teaching. RESULTS: The 25 Internal Medicine faculty participants had a mean of 13 years in clinical practice, including 10 years precepting residents. During the three months post-FDP, faculty treated a mean of 22 patients with chronic pain on long-term opioids and precepted a mean of seven residents caring for patients on long-term opioids. At three months post-FDP, there were significant improvements in correct responses to knowledge questions (68% to 79% P = 0.008), "high-level" confidence in safer opioid prescribing clinical practice (43.5% to 82.6% P = 0.007) and resident teaching (45.8% to 83.3%, P = 0.007), and improvements in alignment of desired attitudes toward safer opioid prescribing. There were nonsignificant increases in self-reported safe opioid prescribing resident teaching. CONCLUSIONS: A skills-based faculty development program that includes a lecture followed by an OSCE can improve Internal Medicine faculty safe opioid prescribing knowledge, attitudes, and clinical and teaching confidence. Improving resident teaching may require additional training in safe opioid prescribing teaching skills.

Managing Concerning Behaviors in Patients Prescribed Opioids for Chronic Pain: A Delphi Study.

BACKGROUND: Current guideline-recommended monitoring of patients prescribed long-term opioid therapy (LTOT) for chronic pain will likely result in increased identification of behaviors of concern for misuse and addiction, but there is a dearth of empiric evidence about how these behaviors should be managed. OBJECTIVE: To establish expert consensus about treatment approaches for common and challenging concerning behaviors that arise among patients on LTOT. DESIGN: We used a Delphi approach, which allows for generation of consensus. PARTICIPANTS: Participants were clinical experts in chronic pain and opioid prescribing recruited from professional societies and other expert groups. MAIN MEASURES: The Delphi process was conducted online, and consisted of an initial brainstorming round to identify common and challenging behaviors, a second round to identify management strategies for each behavior, and two rounds to establish consensus and explore disagreement/uncertainty. KEY RESULTS: Forty-two participants completed round 1, 22 completed round 2, 30 completed round 3, and 28 completed round 4. Half of round 1 participants were female (52%), and the majority were white (83%). Most (71%) were physicians, and most participants practiced in academic primary (40%) or specialty care (19%).The most frequently cited common and challenging behaviors were missing appointments, taking opioids for symptoms other than pain, using more opioid medication than prescribed, asking for an increase in opioid dose, aggressive behavior, and alcohol and other substance use. Across behaviors, participants agreed that patient education and information gathering were important approaches. Participants also agreed that stopping opioids is not important initially, but if initial approaches do not work, tapering opioids and stopping opioids immediately may become important approaches. CONCLUSIONS: This study presents clinical expert consensus on how to manage concerning behaviors among patients on LTOT. Future research is needed to investigate how implementing these management strategies would impact patient outcomes, practice and policy.

Correction to: Managing Concerning Behaviors in Patients Prescribed Opioids for Chronic Pain: a Delphi Study.

In this paper, the author degrees were presented incorrectly. The correct listing is above.

Breaking tolerance: the autoimmune aspect of atherosclerosis.

Atherosclerotic cardiovascular disease (ASCVD) is a chronic inflammatory disease of the arterial walls and is characterized by the accumulation of lipoproteins that are insufficiently cleared by phagocytes. Following the initiation of atherosclerosis, the pathological progression is accelerated by engagement of the adaptive immune system. Atherosclerosis triggers the breakdown of tolerance to self-components. This loss of tolerance is reflected in defective expression of immune checkpoint molecules, dysfunctional antigen presentation, and aberrations in T cell populations - most notably in regulatory T (Treg) cells - and in the production of autoantibodies. The breakdown of tolerance to self-proteins that is observed in ASCVD may be linked to the conversion of Treg cells to 'exTreg' cells because many Treg cells in ASCVD express T cell receptors that are specific for self-epitopes. Alternatively, or in addition, breakdown of tolerance may trigger the activation of naive T cells, resulting in the clonal expansion of T cell populations with pro-inflammatory and cytotoxic effector phenotypes. In this Perspective, we review the evidence that atherosclerosis is associated with a breakdown of tolerance to self-antigens, discuss possible immunological mechanisms and identify knowledge gaps to map out future research. Rational approaches aimed at re-establishing immune tolerance may become game changers in treating ASCVD and in preventing its downstream sequelae, which include heart attacks and strokes.

Developmental delay in a community setting: Role of a primary care physician.

The early part of childhood especially the first 1000 days plays an essential role in the growth and development of the child. Various internal and external factors affect the child's development, including genetic factors, socioeconomic status, sociocultural environment, maternal mental health, and the parenting environment. There is a high prevalence of developmental delay 17.6% globally, whereas in India, it is around 6.6%. Numerous screening tools are available to detect developmental delay in the child early. Early identification and intervention are crucial because we can have a better outcome for the child if intervention is performed on time. The children can be identified during the postnatal follow-up period. Literature has shown that few parents take their children for regular developmental assessment after delivery. Identifying the developmental impairment early from a primary care physician's point of view is essential. In India under the Rashtriya Bal Swasthya Kariyakram (RBSK), the children are screened at home, Anganwadi centers, and schools to detect at-risk children under 4D's, so that early intervention can be planned by linking them to District Early Intervention Center.

Identification of apolipoprotein B-reactive CDR3 motifs allows tracking of atherosclerosis-related memory CD4+T cells in multiple donors.

Introduction: Atherosclerosis is a major pathological condition that underlies many cardiovascular diseases (CVDs). Its etiology involves breach of tolerance to self, leading to clonal expansion of autoreactive apolipoprotein B (APOB)-reactive CD4+T cells that correlates with clinical CVD. The T-cell receptor (TCR) sequences that mediate activation of APOB-specific CD4+T cells are unknown. Methods: In a previous study, we had profiled the hypervariable complementarity determining region 3 (CDR3) of CD4+T cells that respond to six immunodominant APOB epitopes in most donors. Here, we comprehensively analyze this dataset of 149,065 APOB-reactive and 199,211 non-reactive control CDR3s from six human leukocyte antigen-typed donors. Results: We identified 672 highly expanded (frequency threshold > 1.39E-03) clones that were significantly enriched in the APOB-reactive group as compared to the controls (log10 odds ratio ≥1, Fisher's test p < 0.01). Analysis of 114,755 naïve, 91,001 central memory (TCM) and 29,839 effector memory (TEM) CDR3 sequences from the same donors revealed that APOB+ clones can be traced to the complex repertoire of unenriched blood T cells. The fraction of APOB+ clones that overlapped with memory CDR3s ranged from 2.2% to 46% (average 16.4%). This was significantly higher than their overlap with the naïve pool, which ranged from 0.7% to 2% (average 1.36%). CDR3 motif analysis with the machine learning-based in-silico tool, GLIPHs (grouping of lymphocyte interactions by paratope hotspots), identified 532 APOB+ motifs. Analysis of naïve and memory CDR3 sequences with GLIPH revealed that ~40% (209 of 532) of these APOB+ motifs were enriched in the memory pool. Network analysis with Cytoscape revealed extensive sharing of the memory-affiliated APOB+ motifs across multiple donors. We identified six motifs that were present in TCM and TEM CDR3 sequences from >80% of the donors and were highly enriched in the APOB-reactive TCR repertoire. Discussion: The identified APOB-reactive expanded CD4+T cell clones and conserved motifs can be used to annotate and track human atherosclerosis-related autoreactive CD4+T cells and measure their clonal expansion.