Preliminary findings on the absence of PEPITEM release in B cells isolated from Saudi donors: implications for expanded population studies.

Am J Clin Exp Immunol 2024;13(5):215-225 www.ajcei.us /ISSN:2164-7712/AJCEI0159055 Original Article Preliminary findings on the absence of PEPITEM release in B cells isolated from Saudi donors: implications for expanded population studies Mohammed Alassiri1,6, Asma Alanazi2, Tlili Barhoumi4, Bahauddeen Alrfaei2,5, Maisa Alanazi5, Mamoon Rashid3, Aiman S Alhazmi1,6, Mohammed Alasseiri7, Abdulrahman AlMefleh8, Mohammad Boudjelal4, Hayat Shaibah4, Khawlah Almuhalhil4, Fatmah A Mansour4, Zeyad Alehaideb4, Bandar Alghanem4 Department of Basic Sciences, College of Science and Health Professions, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), Riyadh, KSA; 2 Department of Basic Medical Sciences, College of Medicine, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), King Abdulaziz Medical City (KAMC), Riyadh, KSA; 3Department of AI and Bioinformatics, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), King Abdulaziz Medical City, Ministry of National Guard Health Affairs (MNGHA), Riyadh, KSA; 4Medical Research Core Facility and Platforms, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), King Abdulaziz Medical City, Ministry of National Guard Health Affairs (MNGHA), Riyadh, KSA; 5Department of Cellular Therapy and Cancer Research, King Abdullah International Medical Research Center (KAIMRC), King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), King Abdulaziz Medical City, Ministry of National Guard Health Affairs (MNGHA), Riyadh, KSA; 6Department of Pathology and Laboratory Medicine, King Abdulaziz Medical City (KAMC), Riyadh, KSA; 7Department of Medical Laboratory Technology, Faculty of Applied Medical Sciences, Prince Fahad Bin Sultan Chair for Biomedical Research, University of Tabuk, Tabuk, KSA; 8Department of Radiology, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia 1 Received July 8, 2024; Accepted October 8, 2024; Epub October 25, 2024; Published October 30, 2024 Abstract: Background: Adiponectin (AQ) plays a role in regulating immune responses. Previous research indicates that B cells can affect T cell transmigration via the adiponectin-induced peptide PEPITEM in Caucasians. This study explores whether this mechanism is also applicable to Saudi populations, considering potential ethnic variations in immune response. Methods: We conducted unbiased peptidomic screen on B cells, NK cells, and monocytes isolated from the peripheral blood of male healthy Saudi donors. The cells were stimulated with AQ, and the secretion of PEPITEM and other peptides was assessed using liquid chromatography-mass spectrometry (LC-MS/MS). Flow cytometry was utilized to confirm the purity of isolated cell populations and to verify the expression of adiponectin receptors AR1 and AR2. Results: PEPITEM was not detected in the supernatants of AQ-stimulated B cells, NK cells, or monocytes. All three cell populations were isolated and purified with high purity, confirmed by flow cytometry showing AR1 and AR2 expression on the surface of these cells. Specifically, less than 47% of B cells expressed ARs, with AR1 at 12% and AR2 at 17%. AQ stimulation increased the number of identified peptides in B cells and monocytes but decreased peptide numbers in NK cells. Dimensionality reduction analysis demonstrated clear segregation of cell types, with strong reproducibility across technical replicates. Conclusion: The inability of B cells to release PEPITEM in response to AQ stimulation is an interesting finding and it needs more confirmatory tests and experiments, however; a hypothesis about the impact of predisposing factors, such as ethnicity could be formulated and tested in the future. Keywords: Adiponectin, PEPITEM, immunometabolism, T cell recruitment, inflammation Introduction The immune cell interface has various environmental conditions, which in some cases affect their metabolic regulation, leading to potential transcriptional reprogramming of immune cells; however, when reprogramming is impaired, the risk of autoimmune disorder development is increased [1]. Immune cell activation results in the acquisition of new immune functions, such as cytokine https://doi.org/10.62347/XNNO3661 Absence of PEPITEM release in B cells from Saudi donors production or proliferation, and immune cells are required to provide continuous metabolic adaptation to meet bioenergetics demands [2]. One of the activating molecules of immune cells is adiponectin (AQ), which stimulates B cells to release a naturally occurring mediator peptide, called PEPtide inhibitor of transendothelial migration (PEPITEM) that controls the recruitment of immune cells to inflamed tissues [3]. The role of cellular metabolism in immune cell function and differentiation has recently attracted a lot of attention. It has been shown that the treatment of NK cells with interleukin-2 and interleukin-15 increases glycolysis and oxidative phosphorylation (OXPHOS) pathways to improve their function [4]. There is highly regulated integration and interaction between the immune system and metabolism, as evident from accumulating data. For example, recent data indicate that T cell proliferation is influenced by nutrient uptake and utilization [5]. One mechanism by which immune cells respond to activating and proliferating signals is by modulating their metabolic activity; for example, T cells undergo metabolic reprogramming that leads to glucose and amino acid upregulation to improve their immune response [6, 7]. However, the mechanisms of intracellular metabolism and how these metabolic pathways modify the phenotype and activation of immune cells are poorly understood. The only aim of cellular metabolic reactions and pathways is biosynthesis and generation of ATP; however, recent discoveries and insights have shown that these reactions and pathways control and regulate immune cell function by metabolic reprogramming mechanisms [8]. Controlling leukocyte recruitment and transmigration plays a crucial role in inflammatory response initiation and resolution. In some cases, inappropriate leukocyte recruitment is prolonged, eventually leading to pathological conditions, such as diabetes, autoimmune diseases, and atherosclerosis [9-11]. Our previous work showed that B cells stimulated by AQ secrete PEPITEM, which effectively inhibits inappropriate and prolonged T cell transmigration, ameliorates the symptoms, and inhibits disease progression [3]. Moreover, exogenous treatment with PEPITEM restored PEPITEM control of T cell trafficking and inhibited T cell recruitment into inflamed tissues in animal models [3]. During pathological conditions, for 216 many various cellular metabolic modifications, the B cell-AQ (...truncated)