Exosomes in Osteoarthritis: A Review on Their Isolation Techniques and Therapeutic Potential.

Background: Exosomes are the smallest extracellular vesicles (30-150 nm) secreted by all cell types, including synovial fluid. However, because biological fluids are complex, heterogeneous, and contain contaminants, their isolation is difficult and time-consuming. Furthermore, the pathophysiology of osteoarthritis (OA) involves exosomes carrying complex components that cause macrophages to release chemokines and proinflammatory cytokines. This narrative review aims to provide in-depth insights into exosome biology, isolation techniques, role in OA pathophysiology, and potential role in future OA therapeutics. Methods: A literature search was conducted using PubMed, Scopus, and Web of Science databases for studies involving exosomes in the osteoarthritis using keywords "Exosomes" and "Osteoarthritis". Relevant articles in the last 15 years involving both human and animal models were included. Studies involving exosomes in other inflammatory diseases were excluded. Results: Despite some progress, conventional techniques for isolating exosomes remain laborious and difficult, requiring intricate and time-consuming procedures across various body fluids and sample origins. Moreover, exosomes are involved in various physiological processes associated with OA, like cartilage calcification, degradation of osteoarthritic joints, and inflammation. Conclusion: The process of achieving standardization, integration, and high throughput of exosome isolation equipment is challenging and time-consuming. The integration of various methodologies can be employed to effectively address specific issues by leveraging their complementary benefits. Exosomes have the potential to effectively repair damaged cartilage OA, reduce inflammation, and maintain a balance between the formation and breakdown of cartilage matrix, therefore showing promise as a therapeutic option for OA.

The role of monocytes in malaria infection.

Malaria remains one of the most common human infections worldwide. In endemic areas, malaria is a leading cause of morbidity and mortality and it imposes significant socioeconomic burdens on the people affected. Monocytes are part of the immune system controlling parasite burden and protecting the host against malaria infection. Monocytes play their protective roles against malaria via phagocytosis, cytokine production and antigen presentation. Though monocytes are crucial for clearance of malaria infection, they have also been shown to cause adverse clinical outcomes. In this review, we discuss recent findings regarding the role of monocytes in malaria via mechanisms such as parasite detection and clearance, pro-inflammatory activities, and activation of other immune components. We also highlight the role of different monocyte subsets, and other myeloid cells that are involved in malaria infection. However, more investigations are required in order to explore the exact roles of these monocytes in malaria infection.

Endothelial Microparticles as Potential Biomarkers in the Assessment of Endothelial Dysfunction in Hypercholesterolemia.

Background and Objectives: Endothelial microparticles (EMP) particularly CD31+/42−/AV+, CD144+/AV+ and CD62e+/AV+ have been reported as having increased in cardiovascular-related diseases, making them potential biomarkers for endothelial dysfunction. This study aimed to compare these EMPs in patients with hypercholesterolemia and healthy controls and to correlate their levels with endothelium-dependent vasodilation (EDV) assessed via pulse wave analysis (PWA); an established method of assessing endothelial function. Materials and Methods: EMPs from 88 subjects (44 hypercholesterolemia patients and 44 controls) were quantified from whole blood using flow cytometry analysis. Endothelial function was determined using PWA combined with pharmacological challenge. Results: CD31+/42−/AV+ (3.45 ± 4.74 count/µL vs. 1.33 ± 4.40 count/µL; p = 0.03), CD144+/AV+ (7.37 ± 12.66 count/µL vs. 1.42 ± 1.71 count/µL; p = 0.003) and CD62e+/AV+ (57.16 ± 56.22 count/µL vs. 20.78 ± 11.04 count/µL; p < 0.001) were significantly elevated in the hypercholesterolemic group compared with the controls, respectively. There was a significant inverse moderate correlation between all circulating EMPs and EDV: CD31+/42−/AV+ (r = −0.36, p = 0.001), CD144+/AV+ (r = −0.37, p = 0.001) and CD62e+/AV+ (r = −0.35, p = 0.002). Conclusions: All EMPs were raised in the patients with hypercholesterolemia, and these values correlated with the established method of assessing endothelial function.

Single Nucleotide Polymorphisms in XMN1-HBG2, HBS1L-MYB, and BCL11A and Their Relation to High Fetal Hemoglobin Levels That Alleviate Anemia.

Anemia is a condition in which red blood cells and/or hemoglobin (Hb) concentrations are decreased below the normal range, resulting in a lack of oxygen being transported to tissues and organs. Those afflicted with this condition may feel lethargic and weak, which reduces their quality of life. The condition may be manifested in inherited blood disorders, such as thalassemia and sickle cell disease, whereas acquired disorders include aplastic anemia, chronic disease, drug toxicity, pregnancy, and nutritional deficiency. The augmentation of fetal hemoglobin (HbF) results in the reduction in clinical symptoms in beta-hemoglobinopathies. Several transcription factors as well as medications such as hydroxyurea may help red blood cells produce more HbF. HbF expression increases with the downregulation of three main quantitative trait loci, namely, the XMN1-HBG2, HBS1L-MYB, and BCL11A genes. These genes contain single nucleotide polymorphisms (SNPs) that modulate the expression of HbF differently in various populations. Allele discrimination is important in SNP genotyping and is widely applied in many assays. In conclusion, the expression of HbF with a genetic modifier is crucial in determining the severity of anemic diseases, and genetic modification of HbF expression may offer clinical benefits in diagnosis and disease management.

Orchestrated Cytokines Mediated by Biologics in Psoriasis and Its Mechanisms of Action.

Psoriasis is an autoimmune disease mediated by disturbed T cells and other immune cells, and is defined by deep-red, well-demarcated skin lesions. Due to its varied etiologies and indefinite standard pathogenesis, it is challenging to consider the right treatment exclusively for each psoriasis patient; thus, researchers yearn to seek even more precise treatments other than topical treatment and systemic therapy. Using biologics to target specific immune components, such as upregulated cytokines secreted by activated immune cells, is the most advanced therapy for psoriasis to date. By inhibiting the appropriate pro-inflammatory cytokines, cellular signaling can be altered and, thus, can inhibit further downstream inflammatory pathways. Herein, the roles of cytokines with their mechanisms of action in progressing psoriasis and how the usage of biologics alleviates cellular inflammation are discussed. In addition, other potential pro-inflammatory cytokines, with their mechanism of action, are presented herein. The authors hope that this gathered information may benefit future research in expanding the discovery of targeted psoriasis therapy.

Global endothelial function assessment using pulse wave analysis in hypercholesterolemia

To compare global endothelial function assessed by pulse wave analysis (PWA) using the ratio of endothelium dependent vasodilatation (EDV) to endothelium independent vasodilatation (EIV) in patients with hypercholesterolemia and controls. 92 subjects [46 hypercholesterolemics, 46 controls] were studied at standardized conditions. Baseline augmentation index (AIx) was assessed followed by the administration of 0.5 mg sublingual nitroglycerine, an endothelium independent vasodilator. AIx was assessed and the maximum change in AIx after nitroglycerine was recorded as EIV. After a washout period of 30 minutes, 400 µg of inhaled salbutamol, an endothelium dependent vasodilator was administered. AIx was assessed again and the maximum change in AIx after salbutamol was recorded as EDV. Global endothelial function was calculated as EDV:EIV ratio. EDV and EIV in patients with hypercholesterolemia compared to controls were 2.97 ± 3.95 and 6.65 ± 3.80 (p<0.001); and 13.41 ± 4.57 and 15.88 ± 4.78 (p=0.01) respectively. EDV:EIV ratio was significantly reduced in patients with hypercholesterolemia compared to controls; 0.21 ± 0.38 and 0.44 ± 0.24 (p<0.001) respectively. EDV:EIV ratio was significantly reduced in patients with hypercholesterolemia compared to controls. PWA is a potential clinical tool to assess global endothelial function in patients with hypercholesterole

Mesenchymal Stem Cells: Current Concepts in the Management of Inflammation in Osteoarthritis.

Osteoarthritis (OA) has traditionally been known as a "wear and tear" disease, which is mainly characterized by the degradation of articular cartilage and changes in the subchondral bone. Despite the fact that OA is often thought of as a degenerative disease, the catabolic products of the cartilage matrix often promote inflammation by activating immune cells. Current OA treatment focuses on symptomatic treatment, with a primary focus on pain management, which does not promote cartilage regeneration or attenuate joint inflammation. Since articular cartilage have no ability to regenerate, thus regeneration of the tissue is one of the key targets of modern treatments for OA. Cell-based therapies are among the new therapeutic strategies for OA. Mesenchymal stem cells (MSCs) have been extensively researched as potential therapeutic agents in cell-based therapy of OA due to their ability to differentiate into chondrocytes and their immunomodulatory properties that can facilitate cartilage repair and regeneration. In this review, we emphasized current knowledge and future perspectives on the use of MSCs by targeting their regeneration potential and immunomodulatory effects in the treatment of OA.

Extracellular Vesicles from Mesenchymal Stem Cells as Potential Treatments for Osteoarthritis.

Osteoarthritis (OA) is a chronic degenerative disorder of the joint and its prevalence and severity is increasing owing to ageing of the population. Osteoarthritis is characterized by the degradation of articular cartilage and remodeling of the underlying bone. There is little understanding of the cellular and molecular processes involved in pathophysiology of OA. Currently the treatment for OA is limited to painkillers and anti-inflammatory drugs, which only treat the symptoms. Some patients may also undergo surgical procedures to replace the damaged joints. Extracellular vesicles (EV) play an important role in intercellular communications and their concentration is elevated in the joints of OA patients, although their mechanism is unclear. Extracellular vesicles are naturally released by cells and they carry their origin cell information to be delivered to target cells. On the other hand, mesenchymal stem cells (MSCs) are highly proliferative and have a great potential in cartilage regeneration. In this review, we provide an overview of the current OA treatments and their limitations. We also discuss the role of EV in OA pathophysiology. Finally, we highlight the therapeutic potential of MSC-derived EV in OA and their challenges.

Differential polarization and the expression of efferocytosis receptor MerTK on M1 and M2 macrophages isolated from coronary artery disease patients.

BACKGROUND: Differential polarization of macrophage into M1 and M2 mediates atherosclerotic plaque clearance through efferocytosis. Higher expression of Mer proto-oncogene tyrosine kinase (MerTK) on M2 macrophage helps in maintaining macrophage efferocytic efficiency. In healthy individuals, macrophage polarization into M1 and M2 occurs in tissues in concomitance with the acquisition of functional phenotypes depending on specific microenvironment stimuli. However, whether the macrophage differential polarization and MerTK expression vary in coronary artery disease (CAD) patients remain unknown. OBJECTIVE: This study aimed to elucidate the polarization of M1 and M2 macrophage from CAD patients as well as to investigate the expression of MerTK in these macrophage phenotypes. METHODS: A total of 14 (n) CAD patients were recruited and subsequently grouped into "no apparent CAD", "non-obstructive CAD" and "obstructive CAD" according to the degree of stenosis. Thirty ml of venous blood was withdrawn to obtain monocyte from the patients. The M1 macrophage was generated by treating the monocyte with GMCSF, LPS and IFN-γ while MCSF, IL-4 and IL-13 were employed to differentiate monocyte into M2 macrophage. After 7 days of polarization, analysis of cell surface differentiation markers (CD86+/CD80+ for M1 and CD206+/CD200R+ for M2) and measurement of MerTK expression were performed using flow cytometry. RESULTS: Both M1 and M2 macrophage expressed similar level of CD86, CD80 and CD206 in all groups of CAD patients. MerTK expression in no apparent CAD patients was significantly higher in M2 macrophage compared to M1 macrophage [12.58 ± 4.40 vs. 6.58 ± 1.37, p = 0.040]. CONCLUSION: Differential polarization of macrophage into M1 and M2 was highly dynamic and can be varied due to the microenvironment stimuli in atherosclerotic plaque. Besides, higher expression of MerTK in patients with the least coronary obstructive suggest its vital involvement in efferocytosis.

Apoptosis Activity of the Mouse Macrophage Cell Line J774A.1 Infected with a Recombinant BCG consisting the C-Terminus of Merozoite Surface Protein-1 of Plasmodium falciparum.

Macrophage apoptosis exerts an efficient mechanism in controlling intracellular infection during innate immune response against various pathogens including malaria parasites. This study was carried out to determine the apoptosis activity in mouse macrophage cell line J774A.1 infected with a Mycobacterium bovis bacille Calmette-Guerin (BCG) clone and a recombinant BCG clone expressing the C-terminus of merozoite surface protein-1 (BCG-MSP1C) of Plasmodium falciparum for 48 h. In this study, a parent BCG cells was used as a control. The nuclear staining with Hoechst 33342 showed that the BCG-MSP1C cells was capable of increasing the nuclear condensation and morphological stages of apoptosis in the infected cells compared to the BCG-infected cells and the lipopolysaccharide (LPS)-stimulated cells. The flow cytometric analysis using Annexin-V and Propidium iodide (PI) staining confirmed that the BCG-MSP1C cells significantly increased the percentage of early apoptotic activity in the infected macrophage higher than the one stimulated by the parent BCG cells and LPS. This apoptotic response corresponded with the reduction of the anti-apoptotic Bcl-2 protein expression and higher p53 expression. The colorimetric assay demonstrated that the BCG cells capable of stimulating higher production of caspase-1, -3, -8 and -9 while the BCG-MSP1C cells stimulated the expression of caspase-1 and -9 in the infected macrophages, suggesting the involvement of mitochondrial-mediated (intrinsic) pathway of apoptosis. In conclusion, both the BCG and BCG-MSP1C cells are capable of inducing macrophage apoptosis activity in the mouse macrophage cell line J774A.1. This mechanism is important for the elimination of pathogens such as malaria parasite during the phagocytosis activity of macrophage. However, the BCG-MSP1C cells showed higher apoptosis activity than those produced by the parent BCG cells.

Review: the Multiple Roles of Monocytic Microparticles.

Monocytic microparticles (mMP) are microparticles derived from human monocytes either under in vivo or in vitro conditions. The size of mMP is between 0.1 and 1.0 µm. Apart from the size range, mMPs are also identified based on phosphatidylserine and CD14 expression on their surface, though this is not always the case. Monocytic MP are critical players in inflammation, endothelial cell function, and blood coagulation. They exhibit dual function by either helping the progression of such conditions or limiting it, depending on certain factors. Furthermore, the numbers of mMP are elevated in some autoimmune diseases, infectious diseases, and metabolic disorders. However, it is unknown whether mMP play an active role in these diseases or are simply biomarkers. The mechanism of mMP modulation is yet to be identified. In this review, we highlight the mechanism of mMP formation and the roles that they play in inflammation, blood coagulation, and different disease settings.

Immunogenicity of recombinant Mycobacterium bovis bacille Calmette-Guèrin clones expressing T and B cell epitopes of Mycobacterium tuberculosis antigens.

Recombinant Mycobacterium bovis bacille Calmette-Guèrin (rBCG) expressing three T cell epitopes of Mycobacterium tuberculosis (MTB) Ag85B antigen (P1, P2, P3) fused to the Mtb8.4 protein (rBCG018) or a combination of these antigens fused to B cell epitopes from ESAT-6, CFP-10 and MTP40 proteins (rBCG032) were used to immunize Balb/c mice. Total IgG responses were determined against Mtb8.4 antigen and ESAT-6 and CFP-10 B cell epitopes after immunization with rBCG032. Mice immunized with rBCG032 showed a significant increase in IgG1 and IgG2a antibodies against ESAT-6 and MTP40 (P1) B cell epitopes and IgG3 against both P1 and P2 B cell epitopes of MPT40. Splenocytes from mice immunized with rBCG018 proliferated against Ag85B P2 and P3 T cell epitopes and Mtb8.4 protein whereas those from mice-immunized with rBCG032 responded against all Ag85B epitopes and the ESAT-6 B cell epitope. CD4⁺ and CD8⁺ lymphocytes from mice immunized with rBCG018 produced primarily Th1 type cytokines in response to the T cell epitopes. Similar pattern of recognition against the T cell epitopes were obtained with rBCG032 with the additional recognition of ESAT-6, CFP-10 and one of the MTP40 B cell epitopes with the same pattern of cytokines. This study demonstrates that rBCG constructs expressing either T or T and B cell epitopes of MTB induced appropriate immunogenicity against MTB.

Screening of the HLDA9 panel on peripheral blood dendritic cell populations.

Dendritic cells (DC) are a heterogeneous population of bone marrow derived leucocytes that are essential in the initiation of primary T lymphocyte responses. DC are identified as Lineage negative, HLA-DR(+) blood cells that can be further subdivided by CD11c to distinguish CD11c(+) DC and the CD11c(-) plasmacytoid DC. Plasmacytoid DC are the primary IFNα producing cells and express CD303, CD304 and CD123. The CD11c(+) myeloid DC can be divided into populations by CD1c, CD16 and CD141 expression. Despite DC being a functionally unique population, they share many cell surface antigens with myeloid lineage cells and B lymphocytes. We used flow cytometry to screen fresh human blood DC populations with the HLDA9 panel of 63 directly labelled mAb which included mAb specific for a number of B lymphocyte antigens. Of this panel, 23 mAb did not bind Lin(-)HLA-DR(+) DC and 10 bound all four populations. Eight mAb bound to the three CD11c(+) DC populations whilst no mAb tested bound to only pDC. Some of the mAb expected to bind to DC populations failed in this analysis. Overall, this screening highlighted similarities between the CD11c(+) DC subsets and the relatively immature state of peripheral blood DC.

The CD300 molecules regulate monocyte and dendritic cell functions.

The CD300 glycoproteins are a family of related leucocyte surface molecules that modulate a diverse array of cell processes via their paired triggering and inhibitory receptor functions. All family members have a single Ig-V like domain and they share a common evolutionary pathway. At least one member of the family has undergone significant positive selection (ranked second in the top 50) indicating a need to maintain some crucial function. Here we have reviewed the CD300 family members, and their expression on cells of the monocyte and dendritic cell lineages. The consequences of CD300 molecule expression by these leucocyte lineages are only now beginning to be understood. The ability to fine tune monocyte and dendritic cell function and immune responses highlights several potential options to exploit these molecules as therapeutic targets in chronic inflammatory diseases, allergy and other disease states.