INTRODUCTION: The immunosuppressive roles of galectin-3 (Gal-3) in carcinogenesis make this lectin an attractive target for pharmacological inhibition in immunotherapy. Although current clinical immunotherapies appear promising in the treatment of solid tumors, their efficacy is significantly weakened by the hostile immunosuppressive tumor microenvironment (TME). Gal-3, a prominent TME modulator, efficiently subverts the elimination of cancer, either directly by inducing apoptosis of immune cells or indirectly by binding essential effector molecules, such as interferon-gamma (IFNγ). METHODS: N-(2-Hydroxypropyl)methacrylamide (HPMA)-based glycopolymers bearing poly-N-acetyllactosamine-derived tetrasaccharide ligands of Gal-3 were designed, synthesized, and characterized using high-performance liquid chromatography, dynamic light scattering, UV-Vis spectrophotometry, gel permeation chromatography, nuclear magnetic resonance, high-resolution mass spectrometry and CCK-8 assay for evaluation of glycopolymer non-toxicity. Pro-immunogenic effects of purified glycopolymers were tested by apoptotic assay using flow cytometry, competitive ELISA, and in vitro cell-free INFγ-based assay. RESULTS: All tested glycopolymers completely inhibited Gal-3-induced apoptosis of monocytes/macrophages, of which the M1 subtype is responsible for eliminating cancer cells during immunotherapy. Moreover, the glycopolymers suppressed Gal-3-induced capture of glycosylated IFNγ by competitive inhibition to Gal-3 carbohydrate recognition domain (CRD), which enables further inherent biological activities of this effector, such as differentiation of monocytes into M1 macrophages and repolarization of M2-macrophages to the M1 state. CONCLUSION: The prepared glycopolymers are promising inhibitors of Gal-3 and may serve as important supportive anti-cancer nanosystems enabling the infiltration of proinflammatory macrophages and the reprogramming of unwanted M2 macrophages into the M1 subtype.
- MeSH
- Acrylamides chemistry pharmacology MeSH
- Apoptosis drug effects MeSH
- Galectin 3 * antagonists & inhibitors MeSH
- Galectins MeSH
- Interferon-gamma * metabolism MeSH
- Blood Proteins MeSH
- Humans MeSH
- Macrophages drug effects MeSH
- Monocytes * drug effects MeSH
- Tumor Microenvironment drug effects MeSH
- Polymers * chemistry pharmacology MeSH
- Antineoplastic Agents * pharmacology chemistry MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
BACKGROUND: Leukemia is driven by complex interactions within the inherently hypoxic bone marrow microenvironment, impacting both disease progression and therapeutic resistance. Co-cultivation of leukemic cells with feeder cells has emerged as a valuable tool to mimic the bone marrow niche. This study explores the interplay between human commercial SD-1 and patient-derived UPF26K leukemic cell lines with feeders - human fibroblasts (NHDF) and mesenchymal stem cells (hMSCs) under normoxic and hypoxic conditions. RESULTS: Co-cultivation with feeders significantly enhances proliferation and glycolytic activity in the SD-1 cells, improving their viability, while this interaction inhibits the growth and glucose metabolism of the feeders, particularly NHDF. In contrast, UPF26K cells show reduced proliferation when co-cultivated with the feeders while this interaction stimulates NHDF and hMSCs proliferation and glycolysis but reduce their mitochondrial metabolism with hypoxia amplifying these effects. CONCLUSIONS: Cells that switch to glycolysis during co-cultivation, particularly under hypoxia, benefit most from these low oxygen conditions. Due to this leukemic cells' response heterogeneity, targeting microenvironmental interactions and oxygen levels is crucial for personalized leukemia therapy. Advancing co-cultivation models, particularly through innovations like spheroids, can further enhance in vitro studies of primary leukemic cells and support the testing of novel therapies.
- MeSH
- Fibroblasts * metabolism MeSH
- Glycolysis MeSH
- Cell Hypoxia MeSH
- Coculture Techniques methods MeSH
- Leukemia * pathology metabolism MeSH
- Humans MeSH
- Mesenchymal Stem Cells * metabolism MeSH
- Mitochondria metabolism MeSH
- Cell Line, Tumor MeSH
- Tumor Microenvironment MeSH
- Cell Proliferation MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Východiska: Signální dráha UPR (unfolded protein response, tj. odpověď na chybně složené proteiny) pomáhá myelomovým buňkám vyrovnat se se stresovými podmínkami vzniklými v důsledku nadměrné proteosyntézy, a představuje tak pro myelomové buňky prostředek umožňující jejich přežití. Extramedulární onemocnění je agresivnější forma mnohočetného myelomu, při které myelomové buňky ztrácí svoji závislost na mikroprostředí kostní dřeně a mohou infiltrovat jiné tkáně a orgány. Patogeneze vzniku extramedulárního onemocnění není dosud zcela objasněna. Cílem této studie bylo zjistit, zda existuje rozdíl v expresi genů spjatých s UPR mezi plazmatickými buňkami kostní dřeně od pacientů s mnohočetným myelomem a extramedulárním onemocněním. Materiál a metody: Pomocí reverzní transkripce ve spojení s kvantitativní polymerázovou řetězovou reakcí byla analyzována exprese šesti genů spjatých s UPR (ERN1, DDIT3, EIF2AK3, TUSC3, XBP1, HSPA5). Použito bylo celkem 76 vzorků plazmatických buněk kostní dřeně, z toho 44 bylo od pacientů s mnohočetným myelomem a 32 od pacientů s extramedulárním onemocněním. Výsledky: Byl pozorován statisticky významný rozdíl v expresi genů HSPA5, DDIT3, EIF2AK3 a ERN1 mezi skupinou mnohočetného myelomu a extramedulárního onemocnění; exprese byla ve všech případech vyšší u vzorků od pacientů s extramedulárním onemocněním. V případě genů XBP1 a TUSC3 nebyl pozorován statisticky významný rozdíl. Prokázáno bylo také několik statisticky významných korelací mezi hladinou exprese analyzovaných genů a klinickými daty pacientů. Závěr: Výsledky poukazují na možný význam signální dráhy UPR v patogenezi extramedulárního onemocnění. UPR se jeví jako vhodný směr dalšího výzkumu.
Background: The unfolded protein response (UPR) enables myeloma cells to overcome the stress conditions arising from excessive proteosynthesis and thus provides a survival advantage for myeloma cells. Extramedullary disease is a more aggressive form of multiple myeloma in which myeloma cells lose their dependence on the bone marrow microenvironment and are able to infiltrate other tissues and organs. The pathogenesis of extramedullary disease is not fully elucidated yet. The aim of this study was to determine whether there is a difference in the expression of UPR-related genes between bone marrow plasma cells from multiple myeloma and extramedullary disease patients. Materials and methods: Gene expression of six genes involved in UPR (ERN1, DDIT3, EIF2AK3, TUSC3, XBP1, HSPA5) was analyzed by quantitative reverse transcription polymerase chain reaction. In total, 76 bone marrow plasma cell samples were used, of which 44 were from patients with multiple myeloma and 32 from patients with extramedullary disease. Results: A statistically significant difference was observed between the multiple myeloma and extramedullary disease groups regarding the expression of HSPA5, DDIT3, EIF2AK3, and ERN1 genes. However, in the case of XBP1 and TUSC3 genes, no statistically significant difference in the expression was found. Several statistically significant correlations between the expression levels of the analyzed genes and the clinical data of the patients were observed as well. Conclusion: Our results suggest the importance of UPR in the pathogenesis of extramedullary disease. UPR appears to be a promising avenue for further research.
BACKGROUND: Chronic lymphocytic leukemia (CLL) is a common adult leukemia characterized by the accumulation of neoplastic mature B cells in blood, bone marrow, lymph nodes, and spleen. The disease biology remains unresolved in many aspects, including the processes underlying the disease progression and relapses. However, studying CLL in vitro poses a considerable challenge due to its complexity and dependency on the microenvironment. Several approaches are utilized to overcome this issue, such as co-culture of CLL cells with other cell types, supplementing culture media with growth factors, or setting up a three-dimensional (3D) culture. Previous studies have shown that 3D cultures, compared to conventional ones, can lead to enhanced cell survival and altered gene expression. 3D cultures can also give valuable information while testing treatment response in vitro since they mimic the cell spatial organization more accurately than conventional culture. METHODS: In our study, we investigated the behavior of CLL cells in two types of material: (i) solid porous collagen scaffolds and (ii) gel composed of carboxymethyl cellulose and polyethylene glycol (CMC-PEG). We studied CLL cells' distribution, morphology, and viability in these materials by a transmitted-light and confocal microscopy. We also measured the metabolic activity of cultured cells. Additionally, the expression levels of MYC, VCAM1, MCL1, CXCR4, and CCL4 genes in CLL cells were studied by qPCR to observe whether our novel culture approaches lead to increased adhesion, lower apoptotic rates, or activation of cell signaling in relation to the enhanced contact with co-cultured cells. RESULTS: Both materials were biocompatible, translucent, and permeable, as assessed by metabolic assays, cell staining, and microscopy. While collagen scaffolds featured easy manipulation, washability, transferability, and biodegradability, CMC-PEG was advantageous for its easy preparation process and low variability in the number of accommodated cells. Both materials promoted cell-to-cell and cell-to-matrix interactions due to the scaffold structure and generation of cell aggregates. The metabolic activity of CLL cells cultured in CMC-PEG gel was similar to or higher than in conventional culture. Compared to the conventional culture, there was (i) a lower expression of VCAM1 in both materials, (ii) a higher expression of CCL4 in collagen scaffolds, and (iii) a lower expression of CXCR4 and MCL1 (transcript variant 2) in collagen scaffolds, while it was higher in a CMC-PEG gel. Hence, culture in the material can suppress the expression of a pro-apoptotic gene (MCL1 in collagen scaffolds) or replicate certain gene expression patterns attributed to CLL cells in lymphoid organs (low CXCR4, high CCL4 in collagen scaffolds) or blood (high CXCR4 in CMC-PEG).
- MeSH
- Cell Culture Techniques methods MeSH
- Leukemia, Lymphocytic, Chronic, B-Cell * pathology metabolism MeSH
- Gels chemistry MeSH
- Collagen * chemistry pharmacology MeSH
- Humans MeSH
- Polyethylene Glycols * chemistry MeSH
- Receptors, CXCR4 metabolism MeSH
- Carboxymethylcellulose Sodium * chemistry pharmacology MeSH
- Cell Culture Techniques, Three Dimensional methods MeSH
- Tissue Scaffolds * chemistry MeSH
- Cell Survival drug effects MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Bone marrow stromal cells (BMSCs) play a significant role in bone metabolism as they can differentiate into osteoblasts, bone marrow adipocytes (BMAds), and chondrocytes. BMSCs chronically exposed to nutrient overload undergo adipogenic programming, resulting in bone marrow adipose tissue (BMAT) formation. BMAT is a fat depot transcriptionally, metabolically, and morphologically distinct from peripheral adipose depots. Reactive oxygen species (ROS) are elevated in obesity and serve as important signals directing BMSC fate. ROS produced by the NADPH oxidase (NOX) family of enzymes, such as NOX4, may be responsible for driving BMSC adipogenesis at the expense of osteogenic differentiation. The dual nature of ROS as both cellular signaling mediators and contributors to oxidative stress complicates their effects on bone metabolism. This review discusses the complex interplay between ROS and BMSC differentiation in the context of metabolic bone diseases.Special attention is paid to the role of NOX4-ROS in regulating cellular processes within the bone marrow microenvironment and potential target in metabolic bone diseases.
- Publication type
- Journal Article MeSH
- Review MeSH
Aspartate is a proteinogenic non-essential amino acid with several essential functions in proliferating cells. It is mostly produced in a cell autonomous manner from oxalacetate via glutamate oxalacetate transaminases 1 or 2 (GOT1 or GOT2), but in some cases it can also be salvaged from the microenvironment via transporters such as SLC1A3 or by macropinocytosis. In this review we provide an overview of biosynthetic pathways that produce aspartate endogenously during proliferation. We discuss conditions that favor aspartate uptake as well as possible sources of exogenous aspartate in the microenvironment of tumors and bone marrow, where most available data have been generated. We highlight metabolic fates of aspartate, its various functions, and possible approaches to target aspartate metabolism for cancer therapy.
- MeSH
- Aspartic Acid * metabolism MeSH
- Humans MeSH
- Tumor Microenvironment * MeSH
- Neoplasms * metabolism pathology MeSH
- Cell Proliferation MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Review MeSH
Extramedullary multiple myeloma (EMM) is an aggressive form of multiple myeloma (MM). This study represents the most comprehensive next-generation sequencing analysis of EMM tumors (N = 14) to date, uncovering key molecular features and describing the tumor microenvironment. We observed the co-occurrence of 1q21 gain/amplification and MAPK pathway mutations in 79% of EMM samples, suggesting that these are crucial mutational events in EMM development. We also demonstrated that patients with mutated KRAS and 1q21 gain/amplification at the time of diagnosis have a significantly higher risk of EMM development (HR = 2.4, p = 0.011) using data from a large CoMMpass dataset. We identified downregulation of CXCR4 and enhanced cell proliferation, along with reduced expression of therapeutic targets (CD38, SLAMF7, GPRC5D, FCRH5), potentially explaining diminished efficacy of immunotherapy. Conversely, we identified significantly upregulated EZH2 and CD70 as potential future therapeutic options. For the first time, we report on the tumor microenvironment of EMM, revealing CD8+ T cells and NK cells as predominant immune effector cells using single-cell sequencing. Finally, this is the first longitudinal study in EMM revealing the molecular changes from the time of diagnosis to EMM relapse.
- MeSH
- Bone Marrow pathology MeSH
- Middle Aged MeSH
- Humans MeSH
- Multiple Myeloma * genetics pathology MeSH
- Mutation MeSH
- Biomarkers, Tumor genetics MeSH
- Tumor Microenvironment * genetics MeSH
- Prognosis MeSH
- Aged MeSH
- High-Throughput Nucleotide Sequencing * MeSH
- Check Tag
- Middle Aged MeSH
- Humans MeSH
- Male MeSH
- Aged MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
BACKGROUND: The prognostic significance of mast cells and different phenotypes of macrophages in the microenvironment of hepatocellular carcinoma (HCC) following resection is unclear. We aimed in this study to assess the local distribution of infiltrating macrophages and mast cells of specific phenotypes in tissues of HCC and to evaluate their prognostic values for survival of post-surgical patients. METHODS: The clinicopathological and follow-up data of 70 patients with HCC, who underwent curative resection of tumor from 1997 to 2019, were collected. The infiltration of CD68+ and CD163+ macrophages and CD117+ mast cells was assessed immunohistochemically in representative resected specimens of HCC and adjacent tissues. The area fraction (AF) of positively stained cells was estimated automatically using QuPath image analysis software in several regions, such as tumor center (TC), inner margin (IM), outer margin (OM), and peritumor (PT) area. The prognostic significance of immune cells, individually and in associations, for time to recurrence (TTR), disease-free survival (DFS), and overall survival (OS) was evaluated using Kaplan-Meier and Cox regression analyses. RESULTS: High AF of CD68+ macrophages in TC and IM and high AF of mast cells in IM and PT area were associated with a longer DFS. High AF of CD163+ macrophages in PT area correlated with a shorter DFS. Patients from CD163TChigh & CD68TClow group had a shorter DFS compared to all the rest of the groups, and cases with CD163IMlow & CD68IMhigh demonstrated significantly longer DFS compared to low AF of both markers. Patients from CD68IMhigh & CD163PTlow group, CD117IMhigh & CD163PTlow group, and CD117PThigh & CD163PTlow group had a significantly longer DFS compared to all other combinations of respective cells. CONCLUSIONS: The individual prognostic impact of CD68+ and CD163+ macrophages and mast cells in the microenvironment of HCC after resection depends on their abundance and location, whereas the cumulative impact is built upon combination of different cell phenotypes within and between regions.
- MeSH
- Carcinoma, Hepatocellular * pathology MeSH
- Kaplan-Meier Estimate MeSH
- Humans MeSH
- Macrophages pathology MeSH
- Mast Cells pathology MeSH
- Tumor Microenvironment MeSH
- Liver Neoplasms * pathology MeSH
- Prognosis MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Obesity adversely affects bone and fat metabolism in mice and humans. Omega-3 polyunsaturated fatty acids (omega-3 PUFAs) have been shown to improve glucose metabolism and bone homeostasis in obesity. However, the impact of omega-3 PUFAs on bone marrow adipose tissue (BMAT) and bone marrow stromal cell (BMSC) metabolism has not been intensively studied yet. In the present study we demonstrated that omega-3 PUFA supplementation in high fat diet (HFD + F) improved bone parameters, mechanical properties along with decreased BMAT in obese mice when compared to the HFD group. Primary BMSCs isolated from HFD + F mice showed decreased adipocyte and higher osteoblast differentiation with lower senescent phenotype along with decreased osteoclast formation suggesting improved bone marrow microenvironment promoting bone formation in mice. Thus, our study highlights the beneficial effects of omega-3 PUFA-enriched diet on bone and cellular metabolism and its potential use in the treatment of metabolic bone diseases.
- MeSH
- Adiposity MeSH
- Bone and Bones metabolism MeSH
- Bone Marrow * metabolism MeSH
- Humans MeSH
- Disease Models, Animal MeSH
- Mice MeSH
- Obesity complications prevention & control metabolism MeSH
- Fatty Acids, Omega-3 * pharmacology metabolism MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Mice MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Macrophages represent heterogeneous cell population with important roles in defence mechanisms and in homoeostasis. Tissue macrophages from diverse anatomical locations adopt distinct activation states. M1 and M2 macrophages are two polarized forms of mononuclear phagocyte in vitro differentiation with distinct phenotypic patterns and functional properties, but in vivo, there is a wide range of different macrophage phenotypes in between depending on the microenvironment and natural signals they receive. In human infections, pathogens use different strategies to combat macrophages and these strategies include shaping the macrophage polarization towards one or another phenotype. Macrophages infiltrating the tumours can affect the patient's prognosis. M2 macrophages have been shown to promote tumour growth, while M1 macrophages provide both tumour-promoting and anti-tumour properties. In autoimmune diseases, both prolonged M1 activation, as well as altered M2 function can contribute to their onset and activity. In human atherosclerotic lesions, macrophages expressing both M1 and M2 profiles have been detected as one of the potential factors affecting occurrence of cardiovascular diseases. In allergic inflammation, T2 cytokines drive macrophage polarization towards M2 profiles, which promote airway inflammation and remodelling. M1 macrophages in transplantations seem to contribute to acute rejection, while M2 macrophages promote the fibrosis of the graft. The view of pro-inflammatory M1 macrophages and M2 macrophages suppressing inflammation seems to be an oversimplification because these cells exploit very high level of plasticity and represent a large scale of different immunophenotypes with overlapping properties. In this respect, it would be more precise to describe macrophages as M1-like and M2-like.
- MeSH
- Cell Differentiation MeSH
- Cytokines * MeSH
- Phenotype MeSH
- Humans MeSH
- Macrophages * MeSH
- Inflammation MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
