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Pilot’s perception of coordinated turns : relevance of long-arm centrifuge training
No full textHuman sensory systems, including the vestibular and visual systems, evolved for terrestrial locomotion under Earth’s gravity, and therefore tend to generate spatial illusions when confronted with the unfamiliar forces and motions of flight. Spatial disorientation arises when a pilot’s sensed attitude does not correspond to the aircraft’s true attitude, potentially leading to dangerous misinterpretations of flight maneuvers or instrument indications. During coordinated turns the semicircular canals initially detect roll motion, whereas the otolith organs, unable to distinguish gravity from centrifugally generated inertia, continue to signal an upright posture. The ensuing conflict fosters somatogravic illusions in which the perceived and the actual horizon diverge.This thesis investigated spatial-orientation learning in dynamic flight-like environments, focusing on how pilots integrate vestibular cues with cognitive knowledge during aircraft and/or centrifuge turns. In all four studies, spatial orientation in the roll plane was quantified with the Subjective Visual Horizontal (SVH): in total darkness, participants rotated an illuminated line until it matched their perceived horizon, thereby yielding a direct measure of the perceived bank angle.Study I validated a long-arm centrifuge as a proxy for coordinated flight turns. Eight experienced fighter pilots produced closely matching SVH settings in the centrifuge and in the aircraft, supporting the centrifuge’s utility for certain spatialorientation studies.Study II examined how perceptual and cognitive factors contribute to SVH during simulated turns. Pearson correlation analyses revealed that accuracy was associated with each pilot’s knowledge of the G–bank-angle relationship, ability to sense sustained G-load, and ability to estimate and set verbally requested angles using a luminous line in darkness. Nevertheless, a systematic under-tilt and marked inter-individual differences remained.Study III assessed a three-week, multisensory centrifuge-training program. Feedback that combined G-load exposure with explicit tilt information significantly improved perception at larger bank angles (≈ 60°) in both centrifuge and aircraft, although the benefit had faded two years later, underscoring the need for refresher training.Study IV investigated whether increased G-awareness alone could improve rolltilt perception in nine flight cadets with no prior flight time. Five weeks of G-load exposure enhanced G-estimation but left roll-tilt perception unchanged, indicating that G-awareness by itself is insufficient.Taken together, Studies III and IV show that multisensory feedback and directed attention are necessary if perceptual training is to enhance pilots’ awareness of roll attitude. Such training can complement—but never replace—the ironclad reliability of flight instruments.Keywords: spatial disorientation; aviation; pilot; vestibular; cognition; spatial orientation learning; centrifuge training; subjective visual horizontal; perceptionList of scientific articlesThis thesis is based on the following four articles, which will be referred to by their Roman numerals:I. Visual measures of perceived roll tilt in pilots during coordinated flight and gondola centrifugation. Arne Tribukait, Eddie Bergsten, Andreas Brink and Ola Eiken. Journal of Vestibular Research. 2023; 33(1): 1-19. https://doi.org/10.3233/ves-220016II. Factors of significance for the ability of fighter pilots to visually indicate the magnitude of roll tilt during simulated turns in a centrifuge. Andreas Brink, Michail E. Keramidas, Arne Tribukait and Ola Eiken. Perception. 2024; 53(2): 75-92. https://doi.org/10.1177/03010066231209847III. Influence of spatial orientation training in a centrifuge on the ability of fighter pilots to assess the bank angle during flight without visual references. Andreas Brink, Michail E. Keramidas, Eddie Bergsten and Ola Eiken. Journal of Neurophysiology. 2024; 1;132(3):710-721. https://doi.org/10.1152/jn.00129.2024IV. A 5-week centrifuge-based G training with feedback on the magnitude of G force does not improve the perception of roll tilt during simulated coordinated turns. Michail E. Keramidas, Roger Kölegård, Andreas Brink and Ola Eiken. Journal of Neurophysiology. 2024, 1;132(5):1571-1576. https://doi.org/10.1152/jn.00311.2024</p
Delineating cellular heterogeneity of neuroblastoma for identifying therapeutically targetable vulnerabilities
No full textNeuroblastoma is a heterogeneous pediatric neuroendocrine tumor and the most common extracranial solid malignancy in children. Although neuroblastoma etiology is elusive, it is thought to originate from the neural crest lineage, which includes multipotent Schwann cell precursors (SCPs) that can give rise to sympatho-adrenal cells, including chromaffin cells and sympathoblasts. Neuroblastoma has a wide range of clinical outcomes. Some tumors regress spontaneously without treatment, while others respond well to chemotherapy. Approximately half of neuroblastoma cases are highly aggressive, leading to refractory and relapsed disease with poor prognosis. High-risk neuroblastoma is often diagnosed with metastases, preferentially spreading to the bone marrow. Common genetic aberrations associated with the high-risk group include MYCN- amplification, segmental gain of chromosome arms 1q, 2p, 17q and deletion of chromosome arms 1p and 11q.To study the cellular identity and clonal development of human neuroblastoma, we performed joint single-cell DNA and RNA sequencing and orthogonal validation by DNA-FISH with immunofluorescence staining in paper I. In addition to malignant adrenergic cells, we discovered aneuploid pre-malignant SCP-like cells in primary tumors. Clonal expansion was evident in both SCP-like and adrenergic subclones. Gain of chromosome 17 was a shared aberration of pre-malignant SCP-like cells across samples. Genetic analysis and phylogeny of tumor subclones suggest that migrating neural crest cells or multipotent SCPs, prone to aneuploidy, may represent putative tumor-initiating events in neuroblastoma. Abnormal SCP-like cells showed upregulated proliferation scores and downregulated antigen presentation via MHC molecule gene expression compared to non-malignant counterparts.Modeling highly heterogeneous tumors like neuroblastoma is challenging. Since MYCN is a potent oncogenic driver in high-risk neuroblastoma, the TH-MYCN transgenic mouse model is widely used in preclinical studies. However, the extent to which TH-MYCN tumors model the disease has not been explored. In paper II, we comprehensively characterized the single-cell transcriptional landscape of TH-MYCN mouse tumors across various ages, both sexes and genotypes. Joint alignment analysis of tumor cells with normal fetal adrenal gland exhibited resemblance with embryonic chromaffin cells and primarily sympathoblasts. Chromaffin to sympathoblast transitions were observed in tumors, consistent with normal murine developmental trajectories. Comparative analysis with human MYCN-amplified neuroblastoma confirmed similarities in the adrenergic tumor cell compartment. Additionally, inferred ligand-receptor analysis revealed potential therapeutic targets in the NCAM and NOTCH signaling pathways.Existing TH-MYCN cell lines lose the adrenergic identity of neuroblastoma and instead acquire a mesenchymal phenotype. To address this issue, we established novel ex vivo tumoroids that preserve PHOX2B expression and maintain the adrenergic cellular identity of the originating tumor. While ex vivo tumoroid cells demonstrated transcriptional resemblance with embryonic chromaffin cells and sympathoblasts, distinct adrenergic subclusters were enriched in culture. Ex vivo enriched gene expression profiles were associated with synaptic signaling, neuronal morphogenesis and metabolic processes. Some of the upregulated genes in the enriched subclusters were correlated with poor neuroblastoma survival.The major cause of cancer-related death is bone marrow metastasis. To examine the cellular and transcriptional shifts associated with neuroblastoma bone marrow metastasis, we compared the single-cell transcriptomes of non-metastatic and metastatic bone marrow biopsies in paper III. Metastatic tumor cells presented an adrenergic phenotype and acquired a transcriptional signature associated with poor neuroblastoma prognosis. We detected an immunosuppressive microenvironment in the metastatic samples encompassing B cell depletion and enriched regulatory T cell activity. We further identified cytotoxic T cells and CD56bright NK cells with upregulated expression of inhibitory receptors in the bone marrow metastatic niche. By flow cytometry analysis, we confirmed the presence of tumor cells and compositional shifts in B cell and T cell populations in matched bone marrow biopsies. Moreover, metastatic samples with enriched macrophages and mature neutrophils contributed to interactions with disseminated tumor cells via NOTCH signaling and other immunoregulatory interactions.Taken together, this thesis provides an overview of the tumor cell atlas of neuroblastoma and bone marrow metastatic niche remodeling, which may contribute to the identification of new therapeutic approaches. Furthermore, we highlight the translational potential of TH-MYCN in vivo and novel ex vivo models for the therapeutic testing of these new targets.List of scientific papersI. Olsen TK*, Otte J*, Mei S*, Embaie BT, Kameneva P, Cheng H, Gao T, Zachariadis V, Tsea I, Björklund Å, Kryukov E, Hou Z, Johansson A, Sundström E, Martinsson T, Fransson S, Stenman J, Fard SS, Johnsen JI, Kogner P, Adameyko I, Enge M, Kharchenko PV*, Baryawno N *. Joint single-cell genetic and transcriptomic analysis reveal pre- malignant SCP-like subclones in human neuroblastoma. Mol Cancer. 2024 Aug 31;23(1):180. https://doi.org/10.1186/s12943-024-02091-yII. Embaie BT*, Sarkar H*, Alchahin AM, Otte J, Olsen TK, Tümmler C, Kameneva P, Artemov AV, Akkuratova N, Adameyko I, Stukenborg JB, Wickström M, Kogner P, Johnsen JI, Mei S, Kharchenko PV, Baryawno N. Comparative Single-Cell Transcriptomics of Human Neuroblastoma and Preclinical Models Reveals Conservation of an Adrenergic Cell State. Cancer Res. 2025 Mar 14;85(6):1015-1034. https://doi.org/10.1158/0008-5472.can-24-1507III. Mei S*, Alchahin AM*, Embaie BT, Gavriliuc IM, Verhoeven BM, Zhao T, Li X, Jeffries NE, Pepich A, Sarkar H, Olsen TK, Wickström M, Stenman J, Reina-Bedoya O, Kharchenko PV, Saylor PJ, Johnsen JI, Sykes DB, Kogner P*, Baryawno N *. Single-cell analyses of metastatic bone marrow in human neuroblastoma reveals microenvironmental remodeling and metastatic signature. JCI Insight. 2024 Feb 15;9(6):e173337. https://doi.org/10.1172/jci.insight.173337*Contributed equally.</p
Improving wellbeing and quality of life in individuals with multiple health conditions and common mental health problems
No full textBackgroundAs the population ages, the number of adults living with multimorbidity- defined as having two or more chronic conditions-is increasing. The current healthcare system often focuses on treating one disease at a time, can fail to meet these individuals ' needs with risk of on the one hand unmet care needs and on the other risk of more visits to more healthcare providers and medications than benefits the individual. To better serve older individuals with multimorbidity, the healthcare system, particularly primary care, needs to adapt and evolve. This involves adopting a more person-centred and holistic approach to care, considering the individual' wants and needs, all health conditions, and overall well-being, rather than treating each disease separately. By doing so, the healthcare system can provide coordinated, person-centred care that effectively meets the complex needs of these individuals.AimThe aim of this thesis was to develop and pilot a complex intervention to improve wellbeing and quality of life in individuals with multiple health conditions and common mental health problems in Swedish primary care.MethodsThis thesis involves four studies involving quantitative, qualitative and mixed methods methodologies. Study 1 is a systematic literature review and qualitative synthesis of collaborative care interventions for individuals with multimorbidity and common mental health problems in primary care and community settings and a map of intervention components. Study 2 is a total population-based cohort study investigating the association between having multimorbidity with an increasing number of chronic diseases and potentially inappropriate prescribing of benzodiazepines in the total adult population in Stockholm, Sweden in two cohorts, one before and one during COVID-19. Study 3 is a qualitative study exploring GP's experiences of managing individuals with multimorbidity and common mental health problems in primary care in Stockholm, Sweden using reflexive thematic analysis. From the findings of study 1 and 3 the intervention Health and Life in Balance was developed for improving patient capacity for older people with multimorbidity. The intervention includes the collaborative care components of relational continuity, a shared care plan, and teamwork identified in study 1. Moreover, it uses a person-centred approach to care, minimally disruptive medicine, to assess patient capacity and workload in individuals with multimorbidity. This approach was identified in the literature sharing similarities with the findings from study 3. Study 4 is a pragmatic convergent mixed methods non- randomised pilot study. The study aimed to assess the acceptance, feasibility and further need of development of Health and Life in Balance.ResultsIn study 1, 12 Randomised controlled studies were included. Eleven studies with medium to high quality showed positive results in reducing depressive symptoms in individuals with multimorbidity involving depression and one chronic disease. The last study had medium to low quality and investigated the effect on anxiety and depressive symptoms. There was a lack of medium to high quality studies addressing collaborative care for individuals with multimorbidity and anxiety, and for individuals with more than two health conditions. The qualitative synthesis identified a map of shared content in the 11 studies of collaborative care models with medium to high quality. In study 2, an association between an increasing number of chronic conditions and potentially inappropriate prescribing of benzodiazepines was identified in both cohorts, before and during COVID-19. The logistic regression model was adjusted for age and socioeconomic status and stratified for sex. The association remained although decreased when adjusting for each of the clinical confounders: number of healthcare visits, polypharmacy, and having a psychiatric diagnosis. In study 3 two themes were generated: Unmet patient needs and fragmented care send patients and physicians off balance and Dancing with the patient individually and together with others leads to confident and satisfied patients and physicians. General practitioners found it hard to address common mental health problems in individuals with multimorbidity, limited by disease-specific guidelines and fragmentation. They highlighted that aspects of person-centred care, involving relational continuity, flexibility and teamwork, would help them better address these individuals. In study 4, Health and Life in Balance was acceptable and feasible regarding to have relational continuity to a district nurse addressing non-disease-specific issues for individuals in need of it. However, the intervention needs further development. The intervention needs to be targeted for individuals with the greatest care needs including aspects of frailty, deprived psychosocial situation and many healthcare providers. Further, it needs to be further developed to provide person-centred care in a busy primary care setting, including raising priority for individuals with multimorbidity involving non- disease-specific needs, addressing how to provide and implement a person- centred care, and addressing how to provide teamwork and cooperation with others.ConclusionsThis thesis identifies individuals with multiple health conditions to be at risk of potentially inappropriate prescribing of benzodiazepines and poorly addressed non-disease-specific issues in today's fragmented healthcare system and shows person-centred care as a potential solution to address these issues. Furthermore, it describes the development and piloting of Health and Life in Balance, a person-centred intervention aiming to address these risks. The findings from the pilot study identified the intervention to be acceptable and feasible when at-need individuals had relational continuity to a district nurse who could address non-disease-specific needs. Yet, the intervention needs further development to identify and target individuals with the greatest care needs, make these individuals a priority in primary care, and to ensure teamwork and delivery and implementation of person-centred care. The results from this thesis and included studies can inform policy makers and other stakeholders of the need of prioritising a shift from fragmented care to person- centred and holistic care for vulnerable individuals and a need of further intervention development to address these individuals' needs. Furthermore, the findings can be used in further intervention development. However, in future intervention development it is important to involve individuals with multimorbidity and other stakeholders in both the research group and in different phases of the intervention development.List of scientific papersI. Specific content of collaborative care: a systematic review of collaborative care interventions for patients with multimorbidity involving depression and/or anxiety in primary care. C Kappelin, A Carlsson, C Wachtler Fam Pract 2021 Sep 21;39(4):725-734. https://doi.org/10.1093/fampra/cmab079II. The association between number of chronic diseases and potentially inappropriate benzodiazepine prescribing: A total population-based cohort study in Region Stockholm, Sweden. Caroline Kappelin, Caroline Wachtler, Gunnar Ljunggren, Axel Carlsson. [Submitted]III. Dancing with the patient: a qualitative study of general practitioners' experiences of managing patients with multimorbidity and common mental health problems. Kappelin C, Sandlund C, Westman J, Wachtler C. BMC Prim Care. 2023 Apr 20;24:104. https://doi.org/10.1186/s12875-023-02056-yIV. Health and Life in Balance an intervention to improve patient capacity for older people with multimorbidity: A pragmatic mixed methods non-randomised pilot study. Caroline Kappelin, Klas Ytterbrink Nordenskiöld, Elisabeth Bos Sparén, Annica Lagerin, Caroline Wachtler. [Submitted]</p
Investigating the interplay between frailty and healthcare utilization in older adults
No full textFrailty, a syndrome characterized by physiological decline and below average resilience to stressors, often results in clinical complexity and poor quality of life. In this thesis, we examine how frailty has evolved over time in the older population, its association with acute clinical events, and how it relates to avoidable hospitalizations and late-life healthcare use. We used data from the longitudinal population-based Swedish National study on Aging and Care in Kungsholmen (SNAC-K), involving 3363 adults aged 60 and above.In Study I, frailty was measured with a data-driven 40 deficit frailty index (FI) among 2941 participants over the first six SNAC-K waves (median 9.6-year follow-up). Temporal trends in frailty state transitions were investigated, and life expectancies from age 60 in frailty states across five birth years (1900, 1910, 1920, 1930, 1940) were predicted. We found that those born more recently were less likely to transition from mild as well as moderate and severe frailty to death. Moreover, the number and proportion of years lived with frailty after age 60 expanded and occurred later in life in more recent years.Including 3146 SNAC-K participants, Study II explored the relationship between frailty trajectories, measured with a data-driven 40 deficit FI over five waves of SNAC-K, and acute clinical events, specifically MIs (myocardial infarctions), LRTIs (lower respiratory tract infections), and falls, resulting in hospitalization from five years before to twelve years after SNAC-K baseline (median follow-up 11 years). We found higher levels of frailty from age 75 to 95 with each additional acute event. The frailty trajectories differed most by the number of falls, with greater differences in the trajectories by number of MIs and LRTIs once we imputed frailty at death.Study III examined the association between physical frailty status and incident avoidable hospitalizations (median 7.6-year follow-up), defined as hospitalizations for specific chronic or acute conditions that could have been prevented through adequate outpatient care. Among the included 3168 SNAC-K participants, we found the relative and absolute risk of avoidable hospitalization to be higher among those who were pre-frail and frail. Moreover, we found this association to be greater among older adults aged Study IV included 1467 participants that died in the first 15 years of SNAC-K. Latent class trajectories of walking speed, as a proxy of frailty, were generated based on the first five waves of SNAC-K (median 11-year follow-up) and the associations of the different classes with healthcare utilization (total and avoidable hospitalizations, outpatient specialist care visits, days in hospital) in the last year of life were evaluated. We identified three classes: 1) 'faster speed and gradual decline', 2) rapidly declining', and 3) 'slower speed and gradual decline.' Inpatient hospitalizations and length of stay were less frequent in the slower speed and gradual decline class, avoidable hospitalization rates were higher in the rapidly declining class, and outpatient specialist care was less common in both, compared to the faster speed and gradual decline class. We found evidence that avoidable hospitalization rates were mitigated by informal and formal social care.With trends of living longer with frailty, preventing its progression at earlier stages is essential. Prevention of acute clinical events and frailty should be promoted, and provision of informal and formal social care could reduce excessive healthcare utilization among frail adults. Further investigations should explore whether and how the care of individuals with frailty can be optimized.Keywords: frailty, health trends, myocardial infarction, lower respiratory tract infections, falls, hospitalization, avoidable hospitalization, healthcare utilization, walking speed, end of lifeList of scientific papersI. Tazzeo C, Rizzuto D, Calderón-Larrañaga A, Dekhtyar S, Zucchelli A, Xia X, Fratiglioni L, Vetrano DL. Living longer but frailer? Temporal trends in life expectancy and frailty in older swedish adults. J Gerontol A Biol Sci Med Sci. 2024;79(1):glad212. https://doi.org/10.1093/gerona/glad212II. Tazzeo C, Gregorio C, Rizzuto D, Fratiglioni L, Maggi S, Welmer A-K, Zucchelli A, Calderón-Larrañaga A, Vetrano DL. Acute clinical events and trajectories of frailty after age 60: A population-based cohort study. [Manuscript]III. Tazzeo C, Rizzuto D, Calderón-Larrañaga A, Gentili S, Lennartsson C, Xia X, Fratiglioni L, Vetrano DL. Avoidable hospitalizations in frail older adults: The role of sociodemographic, clinical, and care-related factors. J Am Med Dir Assoc. 2024;25(11):105225. https://doi.org/10.1016/j.jamda.2024.105225IV. Tazzeo C, Gregorio C, Rizzuto D, Fratiglioni L, Gentili S, Welmer A-K, Calderón-Larrañaga A, Vetrano DL. Trajectories of walking speed after age 60 and healthcare utilization in the last year of life. [Manuscript].</p
Immunoprofiling the inflamed tissue in rheumatic diseases using single-cell technologies
No full textAutoimmune rheumatic diseases affect millions of people around the world, causing physical disability and poor quality of life. Current treatments ameliorate the symptoms, without curing the patients. Therefore, there is an urgent need to investigate further the diseases' mechanisms and to gain new knowledge. Here, we chose to work with three of these diseases: rheumatoid arthritis (RA), psoriatic arthritis (PSA) and idiopathic inflammatory myopathies (IIM). The overall aim of this thesis was to deeply characterise immune cells, primarily effector and memory T cells, that reside in the inflamed tissue and may contribute to the disease development and progression (Figure 1). To achieve this on transcriptomic and phenotypic levels, we used a combination of single-cell RNA sequencing (scRNA-seq) and flow cytometry.In RA, we isolated mononuclear cells from peripheral blood (PB) and synovial fluid (SF) from patients with or without anti-citrullinated protein antibodies (ACPA). We identified a G-protein coupled receptor 56 (GPR56) delineating peripheral helper T (TPH) cells in SF. TPH cells were also characterised by high expression of tissue resident memory and inhibitory receptors. GPR56 expression on CD4+ T cells was higher in ACPA+ RA, and these cells were clonally expanded (study I). Further investigating GPR56 expression, we also identified it on cytotoxic tissue resident memory and effector memory CD8+ T cells in SF of patients with RA and PsA (study II). Comparing the transcriptomic profile between ACPA+ and ACPA- patients revealed significantly higher type I interferon-stimulated gene (ISG) expression and increased interferon score in mononuclear cells in SF, but not PB, of ACPA- RA. This signature was derived from macrophages, conventional dendritic cells 2 (cDC2) and T cells, and was confirmed in synovial tissue of two ACPA- patients from an independent cohort. Interestingly, there were no differences in the IFN-a or -ß production in SF of the two patient groups (study III).In IIM, we isolated T cells from PB and skeletal muscle tissues of patients at time of diagnosis. We identified the presence of several T-cell populations infiltrating the muscle of IIM, including T cells with cytotoxic and tissue resident memory signatures, that were clonally expanded. In two patients we detected identical T-cell clones in a follow-up muscle biopsy after treatment suggesting their implication in IIM chronicity (study IV).In summary, our studies identify unique characteristics in the immune profile of inflamed tissues in RA, PsA and IIM, highlighting their importance in disease development and chronicity.List of scientific papersI. Single cell sequencing identifies clonally expanded synovial CD4+ TPH cells expressing GPR56 in rheumatoid arthritis Argyriou, A., M. H. Wadsworth, 2nd, A. Lendvai, S. M. Christensen, A. H. Hensvold, C. Gerstner, A. van Vollenhoven, K. Kravarik, A. Winkler, V. Malmström, and K. Chemin. 2022, Nat Commun, 13: 4046.https://doi.org/10.1038/s41467-022-31519-6II. G-protein coupled receptor 56 is expressed on tissue resident memory cytotoxic CD8 T cells in the synovial joint of patients with inflammatory arthritis Argyriou, A., B. Horuluoglu, C. Gerstner, L. Alemo Munters, F. Hjelm, C. Wennerstrom, M. Sharaf, V. Malmström, A. Hensvold, V. Oke, A. Antovic and Karine Chemin [Manuscript]III. Synovial immune cells exhibit a type I IFN gene signature in ACPA-negative rheumatoid arthritis Argyriou, A., M. H. Wadsworth, 2nd, C. Krishna, C. Gerstner, B. Horuluoglu, M. Sijbranda,, L. Rönnblom, M.L. Eloranta, M. Wahren- Herlenius, A. Hensvold, A. Winkler, V. Malmström and K. Chemin. [Submitted]IV. Single-cell profiling of muscle-infiltrating T cells in idiopathic inflammatory myopathiesArgyriou, A., B. Horuluoglu, A. S. Galindo-Feria, J. S. Diaz-Boada, M. Sijbranda, A. Notarnicola, L. Dani, A. van Vollenhoven, D. Ramsköld, I. Nennesmo, M. Dastmalchi, I. E. Lundberg, L. M. Diaz-Gallo, and K. Chemin. 2023., EMBO Mol Med, 15: e17240.https://doi.org/10.15252/emmm.202217240</p
Radiological aspects of spine diagnostics and surgery
No full textBackground: Intraoperative imaging in spine surgery, such as lateral radiographs and 2D fluoroscopy, was first used in the 1970s. Cone beam computed tomography (CBCT), which utilizes 3D technology, was introduced in the 1990s. Initially developed for dental imaging, CBCT has expanded into various fields, including spine surgery, interventional procedures and musculoskeletal radiology. In spine surgery CBCT helps spine surgeons to identify and correct malplaced pedicle screws before finalizing the procedure, potentially eliminating revision surgeries. However, this imaging technology raises radiation exposure concerns for patients and staff. Furthermore, while postoperative multidetector computed tomography (MDCT) scans remain standard for confirming results, the convergence in image quality between CBCT and MDCT creates opportunities to streamline workflows and reduce redundant imaging, decreasing radiation exposure and healthcare costs. Among individuals with cervical spine injuries requiring surgical fixation, traumatic vertebral artery injury (VAI) presents a significant and potentially dangerous complication. While detection of VAI has improved through routine CT angiography (CTA), modern hybrid operating rooms (OR) equipped with CBCT technology used both in spine surgery and interventional procedures, offer a promising advancement: the ability to diagnose and treat both spinal and vascular injuries in a single session. This integrated approach could streamline patient care and potentially lead to better clinical outcomes.Purpose: To investigate multiple aspects of spine surgery conducted within a hybrid OR. We evaluated occupational radiation exposure from various imaging equipment and different radiation protection shields (Paper I), compared the accuracy of CBCT with postoperative MDCT for detecting pedicle screw breaches (Paper II), assessed whether intraoperative CBCT provides equivalent image quality to postoperative MDCT to potentially reduce cumulative radiation exposure (Paper III), and examined the risk factors, incidence, outcomes, and imaging indicators of VAI in patients treated surgically for subaxial cervical spine injuries in trauma (Paper IV).Methods: In Paper I, scatter radiation was measured in a hybrid OR while imaging an anthropomorphic phantom with three systems: ceiling-mounted hybrid C-arm cone beam CT (hCBCT), mobile O-arm CBCT (oCBCT), and mobile 2D C-arm fluoroscopy. Measurements were taken at various room positions utilizing active personal dosimetry devices and an ionization chamber. Two different radiation protection shields were assessed. Paper II evaluated 260 pedicle screws in twenty spinal fixation patients using intraoperative CBCT and follow-up MDCT. Three neurosurgeons independently graded pedicle screw breach using the Gertzbein scale, with MDCT as reference standard. The diagnostic evaluation methods assessed sensitivity, specificity, and negative predictive value. In Paper III, twenty-seven spinal fixation cases were examined, comparing intraoperative CBCT with postoperative MDCT. Four neuroradiologists independently evaluated images using Likert scales, and visual grading analysis measured modality preferences. Observer agreement was quantified through ICC analysis while image quality was objectively measured via contrast- and signal-to-noise ratios (CNR, SNR). In Paper IV, traumatic subaxial injuries that were surgically treated during a twelve-year period were analyzed, with mortality and morbidity as primary outcomes. Propensity score matching was used to create comparable groups, survival analysis tracked outcomes over time, single- and multiple- variable analyses examined the clinical outcomes.Main Results: Paper I found that OR personnel were exposed to approximately 22% lower radiation when procedures utilized the hCBCT equipment compared to operations conducted with the oCBCT system. The scattered radiation doses emitted from the hCBCT were observed to be 27% less than those from the oCBCT when measured at 200 cm from the phantom. A single rotation for image acquisition utilizing hCBCT corresponded to 12 minutes of C-arm fluoroscopy, whereas the oCBCT corresponded to 16 minutes. The scatter dose decreased by over 90% behind radiation protection shielding, although this protective effect diminished slightly when measured at greater distances (60 cm), likely attributable to secondary radiation reflection from ceilings and walls. In Paper II intraoperative CBCT demonstrated a high negative predictive value of 99.6% in ruling out pedicle screw breaches. It also showed a sensitivity of 90.0% and a specificity of 97.6% in detecting screw accuracy compared to postoperative MDCT. In Paper III intraoperative CBCT was the superior modality in the thoracolumbar spine. Conversely, postoperative MDCT was preferred in cervical spine. The agreement was good for inter-observers and moderate in intra- observers. SNR and CNR were comparable in thoracolumbar imaging, while MDCT provided superior and more consistent image quality in the cervical spine. In Paper IV, our analysis revealed that VAI was predominantly associated with significant high-energy trauma, particularly collisions from motor vehicles, same-and high-level falls, with a demographic profile of male predominance with an age distribution centered at 64.4 years. Two-thirds of the cases with VAI also exhibited spinal cord trauma, with this injury combination linked to higher levels of neurological dysfunction. When adjusting for demographic variables our analysis revealed that VAI showed no significant impact on complications arising after surgery, immediate or extended recovery outcomes, or survival rates. Facet joint dislocation was a distinctive imaging indicator that predicted VAI.Conclusions: In Paper I, hCBCT reduced occupational radiation exposure compared to oCBCT in image-guided spine surgery. Staff can lower radiation exposure through optimal positioning relative to the radiation source, patient, walls and ceilings. This allows for the use of RPSs instead of lead aprons during imaging, improving comfort while reducing whole-body dose. Intraoperative CBCT imaging in Paper II proved to be reliable and equivalent to postoperative MDCT scanning in detecting malpositioned pedicle screws in the thoracolumbar spine, potentially eliminating the need for an additional postoperative MDCT examination. In Paper III, CBCT offered superior image quality for thoracolumbar imaging, while MDCT was better suited for cervical imaging. Intraoperative CBCT could potentially replace postoperative MDCT for thoracolumbar spine procedures, though postoperative MDCT remains essential for cervical spine evaluation. Paper IV found that VAI did not negatively affect the clinical results in surgically treated cervical spine injuries. Although multiple imaging features showed associations with VAI, facet joint dislocation was the only standalone predictor of VAI.List of scientific papersI. Radiation distribution in a hybrid operating room, utilizing different X-ray imaging systems: investigations to minimize occupational exposure. Cewe P, Vorbau R, Omar A, Elmi-Terander A, Edström E. J Neurointerv Surg. 2022 Nov;14(11):1139-1144. https://doi.org/10.1136/neurintsurg-2021-018220II. Intraoperative cone beam computed tomography is as reliable as conventional computed tomography for identification of pedicle screw breach in thoracolumbar spine surgery. Burström G*, Cewe P*, Charalampidis A, Nachabe R, Söderman M, Gerdhem P, Elmi-Terander A, Edström E. Eur Radiol. 2021 Apr;31(4):2349-2356. https://doi.org/10.1007/s00330-020-07315-5III. Image Quality Assessment in Spine Surgery: A Comparison of Intraoperative CBCT and Postoperative MDCT. Cewe P, Skorpil M, Fletcher-Sandersjöö A, VG El-Hajj, Grane P, Fagerlund M, Kaijser M, Elmi-Terander A, Edström E. Acta Neurochir. 167, 94 (2025). https://doi.org/10.1007/s00701-025-06503-wIV. Traumatic Vertebral Artery Injury After Subaxial Cervical Spine Injuries: Incidence, Risk Factors, and Long-Term Outcomes: A Population-Based Cohort Study. El-Hajj VG*, Habashy KJ*, Cewe P*, Atallah E, Singh A, Fletcher-Sandersjöö A, Bydon M, Fagerlund M, Jabbour P, Gerdhem P, Elmi-Terander A, Edström E. Neurosurgery. 2024 Sep 20. https://doi.org/10.1227/neu.0000000000003173</p
Mechanisms and targets in SARS-CoV-2 : from viral protein interactions to antiviral strategies
No full textSARS-COV-2 is an enveloped, positive-stranded RNA virus belonging to the ß- coronaviruses. The interplay between viral proteins and host factors is critical for viral propagation and immune evasion. Extensive virus-host interactome studies have revealed numerous host proteins that modulate SARS-CoV-2 replication, including the interaction between G3BP and the viral N protein.G3BP1, a key RNA-binding protein, is central to assembling SGs in response to various stressors, including viral infection. In Paper I, we demonstrate that the SARS-CoV-2 transiently induced SGs formation at early infection, then N protein binds to G3BP via a conserved ITFG motif within its intrinsically disordered region (N-IDR1), which triggers SG disassembly despite persistent PKR-elF2a activation. An engineered RATA virus lacking this G3BP-binding motif in the N protein induces sustained SG formation, resulting in reduced viral replication in vitro and complete attenuation of pathogenicity in K18-hACE2 mice, highlighting the importance of G3BP-N interaction in SARS-CoV-2 infection. Our data further reveal that N protein recruits G3BP1 to viral replication-transcription complex (RTC) through nsp3 scaffolding at double-membrane vesicle (DMV) pores during the early stage of infection; subsequently, G3BPI's RGG domain concentrates 40S ribosomes to facilitate localized viral mRNA translation. Collectively, these findings illustrate how SARS-CoV-2 strategically exploits G3BP1 to evade antiviral stress responses and enhance replication, offering promising avenues for therapeutic intervention.In Paper II, we employed drug repurposing to screen for antiviral agents by targeting conserved nucleotide-binding pockets (NBPs) across six viral proteins (nsp12, nsp13, nsp14, nsp15, nsp16, and N protein). These NBPs are evolutionarily constrained due to their essential roles in viral replication. Through structure- based screening of three compound libraries-the FDA-approved drug library, the natural product library (NPL), and the LOPAC library-we identified several promising candidates. Subsequent validation using isothermal titration calorimetry (ITC) and in vitro antiviral assays confirmed INCB28060, darglitazone sodium, and columbianadin as potent inhibitors of SARS-CoV-2 replication.In summary, Paper I clarifies a key host-pathogen interaction by demonstrating the complex role of the G3BP-N interaction in viral replication, thereby providing a conserved target for antiviral therapy; Paper II targets conserved viral features NBPs for therapeutic development, addressing both viral evasion mechanisms and clinical applicability.List of scientific papersThis thesis is based on the following publicationsI. SARS-CoV-2 N protein recruits G3BP to double membrane vesicles to promote translation of viral mRNAs. Long S; Guzyk M; Vidakovics LP; Han X; Sun R; Wang M; Panas MD; Urgard E; Coquet JM; Merits A; Achour A; Mcinerney GM. NATURE COMMUNICATIONS. 2024;15(1):10607. DOI: https://doi.org/10.1038/s41467-024-54996-3II. Multi-target direct-acting SARS-CoV-2 antivirals against the nucleotide- binding pockets of virus-specific proteins. Rani R; Long S; Pareek A; Dhaka P; Singh A; Kumar P; McInerney G; Tomar S. VIROLOGY. 2022;577:1-15. DOI: https://doi.org/10.1016/j.virol.2022.08.008</p
Studies on register-based family history of cardiovascular disease : from preclinical to recurrent disease
No full textBackgroundFamily history of coronary heart disease (CHD) is a known risk-factor for incident atherosclerotic cardiovascular disease (ASCVD). The underlying mechanisms are, however, poorly understood and the risk effects are seemingly independent of polygenic risk scores for ASCVD. Furthermore, the evidence of family history of CHD as a risk factor in specific settings, such as in the diagnostics of acute ASCVD events and in secondary prevention is conflicting, which may be partly due to misreporting of family history in studies relying on self-reported data.Methods and resultsIn order to find registered parents and siblings of index subjects, the index population of each study was linked to the Swedish Multi-Generation Register (MGR). Information on registered diagnoses and causes of death in relatives, including myocardial infarction (MI), any coronary revascularization procedure, and in some cases ischemic stroke, was retrieved from the Swedish National Patient register and the Cause of Death Register, along with the age of diagnosis. This data was consequently used to construct different definitions of register- based family history of CHD and ASCVD.Study I. The first consecutive visit of 28,188 adult patients seeking any of four emergency departments in Stockholm due to chest pain between 2013 and 2016 were included. Patients with no or only one parent identifiable in MGR were excluded. In multivariable logistic regression, a family history of early-onset CHD was associated with major adverse cardiac events, including MI and unstable angina, as well as cardiovascular deaths for those discharged from the emergency department, independently of traditional cardiovascular risk factors. The fully adjusted odds ratio for major adverse cardiac events was 1.62 (95% confidence interval [CI] 1.35 - 1.94) and family history had a larger effect on the outcome in subjects Study II. In total, 25,615 patients aged 18-76 years that attended the 1-year follow-up revisit after their first MI between 2005 and 2013 in the Swedish Web- system for Enhancement and Development of Evidence-based care in Heart disease Evaluated According to Recommended Therapies (SWEDEHEART) were included. Patients with no or only one parent identifiable in MGR were excluded. The hazard ratio (HR) for recurrent ASCVD (rASCVD) comparing patients with a family history of ASCVD to those without was estimated using Cox proportional- hazards regression, adjusted for cardiovascular risk factors and socioeconomic determinants. A family history of early-onset ASCVD was associated with an increased risk of rASCVD, with a fully adjusted HR of 1.22 (95% CI 1.05 - 1.42) during a median follow-up of 7.2 years. The addition of family history of early- onset ASCVD to a previously validated risk-model for rASCVD improved risk prediction significantly, albeit with small absolute increments.Study III. A total of 4,251 subjects free of known CHD, from the population-based Swedish Cardiopulmonary bioImage Study (SCAPIS), that had undergone coronary computed tomography angiography and proteomics analysis and with identifiable relatives in MGR were included. In the main analysis, 38 plasma proteins were associated with a family history of early-onset CHD using a false discovery rate of 0.05, whereas 27 remained significant after adjusting for metabolic risk factors. Follistatin (FST) was positively associated with family history and only slightly attenuated when adjusting for metabolic risk factors. Furthermore, 79 proteins were associated with the coronary atherosclerotic burden and differed across subgroups of family history status. In interaction analysis, 18 proteins showed steeper associations with the coronary atherosclerotic burden in subjects with family history as compared to those without such history. In two-sample Mendelian randomization, there was evidence of causal associations between three proteins and CHD; the proprotein convertase subtilisin/kexin type 9 (PCSK9) enzyme, platelet endothelial cell adhesion molecule 1 (PECAM-1) and FST.Study IV. We included 25,302 subjects from the SCAPIS cohort with both parents identifiable in MGR. The SCAPIS core questionnaire on family history of MI in parents and siblings, with and without information on age of onset, was compared to registered diagnoses and deaths due to MI in relatives from registers. The agreement for self-reported family history of MI in any parent or sibling as compared to register-based history was modest, with a Cohen's K = 0.491, 95% CI 0.480 - 0.503, a sensitivity of 57.6%, and specificity of 89.0%. Agreement measures for maternal and paternal history were similar, however, early-onset disease was generally less accurately reported. In subgroup analyses, female subjects consistently reported family history more accurately than male counterparts and having a university degree or a high income was indicative of slightly more accurate reporting.ConclusionsA family history of CHD and ASCVD is an important cardiovascular risk factor in both diagnostic and secondary preventive settings. Family history of particularly early-onset CHD is an independent risk factor for major adverse cardiac events in chest pain patients seeking emergency care, especially in those with otherwise low absolute risk, such as younger patients and those with low initial cardiac troponin values. Family history of early-onset ASCVD also increases the risk for disease recurrence, independently of traditional risk factors, and improves risk prediction in this setting. Furthermore, we present evidence of specific biological processes in familial CHD, as a number of circulating plasma proteins were associated with family history, independently of traditional risk factors. Furthermore, there was evidence of modifying effects of family history on proteins associated with the coronary atherosclerotic burden, indicating distinct effects of these proteins in subjects with family history. We found potentially causal associations for three proteins and CHD, of which FST is a novel marker. Lastly, self-reported family history only to a lesser extent captures registered diagnoses of MI in relatives, demonstrating that the use of register- based family history in research is more robust than self-reports. This enhanced understanding of family history as a risk factor for ASCVD could improve targeted therapies, as well as risk stratification for early diagnostics and treatments.List of scientific papersI. Family history of coronary artery disease is associated with acute coronary syndrome in 28,188 chest pain patients Agnes Wahrenberg, Patrik K.E. Magnusson, Andrea Discacciati, Lina Ljung, Tomas Jernberg, Mats Frick, Rickard Linder, Per Svensson European Heart Journal: Acute Cardiovascular Care. 2020; 9(7): 741-747. https://doi.org/10.1177/2048872619853521II. Cardiovascular family history increases the risk of disease recurrence after a first myocardial infarction Agnes Wahrenberg, Ralf Kuja-Halkola, Patrik K.E. Magnusson, Henrike Habel, Anna Warnqvist, Kristina Hambraeus, Tomas Jernberg, Per Svensson Journal of the American Heart Association. 2021; 10(23): e022264. https://doi.org/10.1161/jaha.121.022264III. Plasma protein profile associated with a family history of early- onset coronary heart disease Agnes Wahrenberg, Lars Lind, Natan Åberg, Henrike Häbel, Marika Ström, Anders Mälarstig, Patrik K.E. Magnusson, Ralf Kuja-Halkola, Göran Bergström, Gunnar Engström, Emil Hagström, Tomas Jernberg, Stefan Söderberg, Carl Johan Östgren, Per Svensson [Submitted]IV. Validation of self-reported family history of myocardial infarction using nationwide health care data Agnes Wahrenberg, Karin Leander, Henrike Häbel, Patrik K.E. Magnusson, Ralf Kuja-Halkola, Göran Bergström, Lars Lind, Emil Hagström, Gunnar Engström, Tomas Jernberg, Stefan Söderberg, Carl Johan Östgren, Per Svensson [Manuscript]</p
Delineating the molecular landscape of breast cancer and response to neoadjuvant systemic treatment
No full textBreast cancer represents a highly heterogeneous and systemic disease, shaped by tumor intrinsic characteristics, tumor microenvironment (TME) as well as host systemic immunity. Within this complex tumor ecosystem, clinical outcomes are determined by the interplay between the different components. Neoadjuvant systemic therapy (NAT) has become standard of care for high-risk early breast cancer and offers a unique opportunity to assess therapeutic sensitivity in vivo. However, not all patients respond to treatment, underscoring the need for predictive biomarkers and insights into resistance.This thesis delineates the molecular landscape of breast cancer and its evolution under therapeutic pressure. Leveraging comprehensive longitudinal tumor biopsies and blood samples from two prospective clinical trials, PROMIX (human epidermal growth factor receptor 2 (HER2)-negative, Paper I-II) and PREDIX HER2 (HER2-positive, Paper III-IV), we apply an integrated multi-omics approach to compute biomarkers reflecting tumor intrinsic features, TME components, and systemic immunity. We aim to identify those biologically driven biomarkers that can guide personalized treatment strategies and elucidate mechanisms of response and resistance to neoadjuvant systemic treatment.In Paper I, we established the immunometabolic phenotype of HER2-negative breast cancer and examined the interplay between tumor-intrinsic metabolic states and immune state. As expected, patients with immunologically "hot" tumors were more likely to achieve a pathologic complete response (pCR). Immunologically "cold" tumors were characterized by elevated tumor metabolic pathway activity, particularly in lipid and amino acid metabolism. We further validated several potential tumor-specific metabolic targets using in vitro co- culture models, supporting that modulating tumor metabolism may improve antitumor immunoregulation. Complementing tissue analysis, we evaluated serum thymidine kinase 1 (TK1) kinetics in the same trial (Paper II). While central Ki67 assessment at baseline provided prognostic value, we found that an increase in TK1 activity after two cycles of chemotherapy was independently associated with improved event-free and overall survival. In HER2-positive disease (Paper III), we dissected the determinants of response to the trastuzumab emtansine (T-DM1) versus dual HER2 blockade with chemotherapy. We found that T-DM1 sensitivity relies on efficient cellular transport activity, whereas resistance is driven by genomic instability and type 2 immune infiltration. Conversely, tumors with high "proteogenomic" ERBB2 expression derived greater benefit from standard dual- HER2 blockade. Finally, expanding to the systemic macroenvironment (Paper IV), we demonstrated that dynamic changes in circulating immune-related proteins, such as IL-33, correlate with local TME activation and pCR. By integrating plasma proteomics with tumor gene expression, we defined "tumor micro- /macroenvironment" (TMME) subtypes, with the immune-excluded subtype associated with poor response.In conclusion, these findings advance our understanding of the biological determinants of treatment response in breast cancer, and facilitate the development of more precise, biology-driven therapeutic strategies.List of scientific papersI. Wang K, Zerdes I, Johansson HJ, Sarhan D, Sun Y, Kanellis DC, Sifakis EG, Mezheyeuski A, Liu X, Loman N, Hedenfalk I, Bergh J, Bartek J, Hatschek T, Lehtio J, Matikas A, Foukakis T. Longitudinal molecular profiling elucidates immunometabolism dynamics in breast cancer. Nat Commun. 2024 May 7;15(1):3837. PMID: 38714665; PMCID: PMC11076527. https://doi.org/10.1038/s41467-024-47932-II. Matikas A, Wang K, Lagoudaki E, Acs B, Zerdes I, Hartman J, Azavedo E, Bjöhle J, Carlsson L, Einbeigi Z, Hedenfalk I, Hellström M, Lekberg T, Loman N, Saracco A, von Wachenfeldt A, Rotstein S, Bergqvist M, Bergh J, Hatschek T, Foukakis T. Prognostic role of serum thymidine kinase 1 kinetics during neoadjuvant chemotherapy for early breast cancer. ESMO Open. 2021 Apr;6(2):100076. Epub 2021 Mar 10. PMID: 33714010; PMCID: PMC7957142. https://doi.org/10.1016/j.esmoop.2021.100076III. Wang K, Zerdes I, Manikis G, Gaessler LF, Johansson HJ, Zhu YJ, Sifakis EG, Hu TB, Morales E, Tiklova K, Tsiknakis N, Salgkamis D, Zarrineh M, Chen Y, Harbers L, Crosetto N, Bergh J, Nilsson M, Lehtio J, Navin N, Hatschek T, Matikas A, Foukakis T. Multi-omic Profiling Reveals Predictive Biomarkers of Response to Antibody-Drug Conjugates in HER2-Positive Breast Cancer [Manuscript]IV. Zerdes I, Wang K, Manikis G, Zhu YJ, Salgkamis D, Morales E, Bernabé AM, Tzoras E, Mezheyeuski A, Tsiknakis N, Sifakis EG, Acs B, Hartman J, Östman A, Papakonstantinou A, Matikas A, Hatschek T, Bergh J, Foukakis T. Longitudinal macro- and microenvironment profiling predicts neoadjuvant treatment response in HER2-positive breast cancer: results from the randomized PREDIX HER2 trial [Manuscript]</p
The role of γδ T cells in haematopoietic stem cell transplantation and malignancies
No full textAllogeneic haematopoietic stem cell transplantation (aHCT) is a potentially curative immunotherapeutic approach for several lethal hematological disorders. However, morbidity and mortality rates remain high after aHCT due to complications such as relapse, graft-versus-host disease (GVHD), and infections. Successful aHCT requires rapid and effective immune reconstitution, particularly within the T-cell compartment, to protect against opportunistic infections and to eliminate residual tumor cells without aggravating GVHD during the immunocompromised period following transplantation. Early reconstitution of γδ T cells plays a substantial role in immune surveillance post-aHCT via mediating anti-infection and anticancer immunity, correlating with favorable clinical outcomes with less GVHD incidence. These unique features have attracted increasing attention towards harnessing γδ T cells as effector cells for cancer immunotherapy. This thesis presents three papers to bring further knowledge on the role of γδ T cells in the dimension of aHCT and their potential use in hematological malignancy therapy.Study I aimed to investigate long-term homeostatic steady-state γδ T cell reconstitution after aHCT, focusing on its associations with previous clinical outcomes. We performed an in-depth analysis of γδ T cell phenotypes, TCR-Y repertoire, and functional responses upon stimulation in 20 recipient/donor pairs using multiparametric flow cytometry and next-generation sequencing of the TCR-y chain. Results showed a comparable phenotypic profile between recipients and donors. Upon PMA/lonomycin stimulation, recipient γδ T cells secreted high levels of cytokines. Furthermore, the TRG repertoire in recipients was almost completely restored, with no significant differences in diversity, clonality, or gene segment usage compared to donors. However, we found an association between overrepresented donor-derived clonotypes and elevated HLA-DR expression in Vδ1 T cells with increased severity of chronic GVHD in some recipients.Study II focused on augmenting the antileukemic activity of expanded γδ T cells by CD34/CD3 bispecific T-cell engager (BTE) in vitro. We demonstrated that the CD34/CD3 BTE effectively activates and redirects γδ T cells (effector cells) against CD34-expressing leukemia cell lines (target cells), as evidenced by their specific cytotoxicity in a dose-dependent manner and high cytokine release. Furthermore, CD34/CD3 BTE induced γδ T cell-mediated killing of primary CD34+ AML blasts. In the presence of CD34/CD3, γδ T cells showed superior antileukemic activity compared to conventional aß T cells, while demonstrating no cytotoxic effects against CD34+ normal cells.In Study III, we aimed to further enhance the antileukemic activity of expanded yo T cells by sensitizing leukemia cells with thymoquinone (TQ) treatment. TQ is a phytochemical compound featuring epigenetic activity and immunomodulatory properties with growing evidence as a potent anticancer candidate. We observed that γδ T cells exhibited rapid and increased cytotoxicity when co-cultured with pre-treated leukemia cell lines with TQ compared to vehicle control and untreated conditions. This enhanced cytotoxicity could be attributed to the upregulated expression of NKG2D and DNAM-1 ligands on leukemia cells after TQ treatment. We also showed that TQ pretreatment in leukemia cell lines supports the condition of the γδ T cells-based CD34/CD3 approach.Altogether, this thesis investigated that long-term γδ T cell reconstitution reaches a homeostatic state with a normalized repertoire. Elevated HLA-DR expression on Vδ1 T cells in cGVHD recipients could be a potential therapeutic target, warranting further investigation. It also highlights the potential use of expanded γδ T cells in hematological malignancy therapy by targeting CD34 and sensitizing leukemia cell lines through thymoquinone for CD34/CD3 BTE treatment.List of scientific papersI. Alagrafi F, Stikvoort A, Gaballa A, Solders M, Ringden O, Poiret T, *Arruda LC, *Uhlin M. γδ T cell characterisation in the long term after haematopoietic stem cell transplantation and its impact on CMV control and cGVHD severity. Clin Transl Immunology. 2025 Mar 7;14(3):e70027. https://doi.org/10.1002/cti2.70027II. Al Agrafi F, Gaballa A, Hahn P, Arruda LCM, Jaramillo AC, Witsen M, Lehmann S, Önfelt B, Uhlin M, Stikvoort A. Selective lysis of acute myeloid leukemia cells by CD34/CD3 bispecific antibody through the activation of γδ T-cells. Oncoimmunology. 2024 Jul 27;13(1):2379063. https://doi.org/10.1080/2162402X.2024.2379063III. Alagrafi F, Stikvoort A, Gaballa A, Saher O, Bazaz S, Sundin M, *Poiret T, *Uhlin M. Enhancing γδ T cell cytotoxicity against leukemia cells through thymoquinone sensitization and CD34/CD3 bispecific antibody activation. [Manuscript]*Shared last authorship.</p