The terms microparticles (MPs) and microvesicles (MVs) refer to large extracellular vesicles (EVs) generated from a broad spectrum of cells upon its activation or death by apoptosis. The unique ...surface antigens of MPs/MVs allow for the identification of their cellular origin as well as its functional characterization. Two basic aspects of MP/MV functions in physiology and pathological conditions are widely considered. Firstly, it has become evident that large EVs have strong procoagulant properties. Secondly, experimental and clinical studies have shown that MPs/MVs play a crucial role in the pathophysiology of inflammation-associated disorders. A cardinal feature of these disorders is an enhanced generation of platelets-, endothelial-, and leukocyte-derived EVs. Nevertheless, anti-inflammatory effects of miscellaneous EV types have also been described, which provided important new insights into the large EV-inflammation axis. Advances in understanding the biology of MPs/MVs have led to the preparation of this review article aimed at discussing the association between large EVs and inflammation, depending on their cellular origin.
The liver represents a unique organ biased toward a tolerogenic milieu. Due to its anatomical location, it is constantly exposed to microbial and food-derived antigens from the gut and thus equipped ...with a complex cellular network that ensures dampening T-cell responses. Within this cellular network, parenchymal cells (hepatocytes), non-parenchymal cells (liver sinusoidal endothelial cells and hepatic stellate cells), and immune cells contribute directly or indirectly to this process. Despite this refractory bias, the liver is capable of mounting efficient T-cell responses. How the various antigen-presenting cell (APC) populations contribute to this process and how they handle danger signals determine the outcome of the generated immune responses. Importantly, liver mounted responses convey consequences not only for the local but also to systemic immunity. Here, we discuss various aspects of antigen presentation and its consequences by the non-professional APCs in the liver microenvironment.
Radiofrequency ablation (RFA) is a potentially curative therapy for hepatocellular carcinoma (HCC). However, incomplete RFA can induce accelerated invasive growth at the periphery. The mechanisms ...underlying the RFA‐induced tumor promotion remain largely unexplored. Three human HCC cell lines were exposed to 45°C‐55°C for 10 minutes, simulating the marginal zone of RFA treatment. At 5‐12 days post‐treatment cell proliferation, parameters of epithelial‐mesenchymal transition (EMT), and activation of mitogen‐activated protein kinases were analyzed. Livers from patients with viral hepatitis without and with HCC (n = 114) were examined to confirm the relevance of altered kinase patterns. In vivo tumorigenic potential of heat‐treated versus untreated HCC cells was studied in nude mice. Heating to 55°C killed all HCC cells, whereas 65%‐85% of cells survived 48°C‐50°C, developing spindle‐like morphology and expressing CD133, cytokeratin (CK)7, CK19, procollagen‐α1(I), and Snail at day 5 after heat exposure, which returned to baseline at day 12. Heat‐exposed HCC cells showed enhanced proliferation and prominent activation of p46‐Shc (Src homology and collagen) and downstream extracellular signal‐related kinase (Erk)1/2. In patients, Shc expression correlated with malignant potential and overall survival. Blocking Erk1/2 reduced proliferation and EMT‐like changes of heat‐treated HCC cells. Implantation of heat‐exposed HEPG2 cells into nude mice induced significantly larger, more aggressive tumors than untreated cells. Conclusions: Sublethal heat treatment skews HCC cells toward EMT and transforms them to a progenitor‐like, highly proliferative cellular phenotype in vitro and in vivo, which is driven significantly by p46Shc‐Erk1/2. Suboptimal RFA accelerates HCC growth and spread by transiently inducing an EMT‐like, more aggressive cellular phenotype. (Hepatology 2013;58:1667–1680)
In recent years, tremendous progress has been made in understanding the roles of extracellular vesicles (EVs) in cancer. Thanks to advancements in molecular biology, it has been found that the ...fraction of EVs called exosomes or small EVs (sEVs) modulates the sensitivity of cancer cells to chemotherapeutic agents by delivering molecularly active non-coding RNAs (ncRNAs). An in-depth analysis shows that two main molecular mechanisms are involved in exosomal modified chemoresistance: (1) translational repression of anti-oncogenes by exosomal microRNAs (miRs) and (2) lack of translational repression of oncogenes by sponging of miRs through long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs). At the cellular level, these processes increase the proliferation and survival of cancer cells and improve their ability to metastasize and resist apoptosis. In addition, studies in animal models have shown enhancing tumor size under the influence of exosomal ncRNAs. Ultimately, exosomal ncRNAs are responsible for clinically significant chemotherapy failures in patients with different types of cancer. Preliminary data have also revealed that exosomal ncRNAs can overcome chemotherapeutic agent resistance, but the results are thoroughly fragmented. This review presents how exosomes modulate the response of cancer cells to chemotherapeutic agents. Understanding how exosomes interfere with chemoresistance may become a milestone in developing new therapeutic options, but more data are still required.
The high mortality rate of cholangiocarcinoma (CCA) is due, in part, to the lack of non‐invasive approaches able to accurately detect this silent tumour at early stages, when therapeutic options can ...be potentially curative or may at least increase the overall survival of patients. The fact that the majority of CCA tumours are not linked to any known aetiological factor highly compromises the monitoring of patients at risk for tumour development and also their early diagnosis. Combination of clinical/biochemical features, imaging techniques and analysis of non‐specific tumour biomarkers in serum are commonly used to help in the diagnosis of CCA, but tumour biopsy is usually required to confirm the diagnosis. Moreover, no prognostic biomarkers are currently used in the clinical setting, deserving more innovative research, and international validation and consensus. Important efforts have been made in the last few years to identify accurate non‐invasive biomarkers, by using innovative techniques and high‐throughput omics technologies. This review summarizes and discusses the advances in the investigation of novel diagnostic and prognostic biomarkers in CCA and envisions the future directions in this field of research.
Despite major advances in medicine, blood-borne biomarkers are urgently needed to support decision-making, including polytrauma. Here, we assessed serum-derived extracellular vesicles (EVs) as ...potential markers of decision-making in polytrauma.
Our Liquid Biopsy in Organ Damage (LiBOD) study aimed to differentiate polytrauma with organ injury from polytrauma without organ injury. We analysed of blood-borne small EVs at the individual level using a combination of immunocapture and high-resolution imaging.
To this end, we isolated, purified, and characterized small EVs according to the latest Minimal Information for Studies of Extracellular Vesicles (MISEV) guidelines from human blood collected within 24 h post-trauma and validated our results using a porcine polytrauma model.
We found that small EVs derived from monocytes CD14
and CD14
CD61
were significantly elevated in polytrauma with organ damage. To be precise, our findings revealed that CD9
CD14
and CD14
CD61
small EVs exhibited superior performance compared to CD9
CD61
small EVs in accurately indicating polytrauma with organ damage, reaching a sensitivity and a specificity of 0.81% and 0.97%, respectively. The results in humans were confirmed in an independent porcine model of polytrauma.
These findings suggest that these specific types of small EVs may serve as valuable, non-invasive, and objective biomarkers for assessing and monitoring the severity of polytrauma and associated organ damage.
Introduction
Gastrointestinal (GI) malignancies, such as cholangiocarcinoma, pancreatic carcinoma, and metastatic colorectal carcinoma, have a poor prognosis and effective therapeutic approaches are ...still challenging. Checkpoint inhibition with PD-1 or PDL-1 antibodies revealed promising results in different tumor entities; however, only few patients with GI tumors can potentially benefit from PD1/PDL1 inhibiting immunotherapy. Further immunotherapeutic strategies for GI malignancies are urgently needed. The aim of this study was to demonstrate that in vitro activation of the immune checkpoint CD40/CD40L can improve DC action towards bile duct, pancreas, and colorectal carcinoma.
Methods
Human DC were isolated from buffy coats from healthy donors, pulsed with tumor lysates and then transduced with adenoviruses encoding human CD40L (Ad-hCD40L). Using transwell assays, the effects of (m)CD40L on DC immunoactivation compared to (s)CD40L were analyzed. Surface marker and cytokine/chemokine expression were measured by flow cytometry, ELISA and cytokine arrays. Capacity of Ad-hCD40L-transduced DC to induce tumor-specific effector cells was tested using MTT proliferation assay and cytotoxicity assays. Apoptosis induction on tumor cells after culturing with supernatants of Ad-hCD40L-transduced DC was analyzed by flow cytometry.
Results
Ad-hCD40L transduction induced a high expression of (s)CD40L and (m)CD40L on DC and seemed to induce a strong cellular CD40/CD40L interaction among DC, leading to the formation of cell aggregates. Due to the CD40/CD40L interaction, a significant upregulation of DC maturation markers and a Th1-shift on cytokines/chemokines in the supernatant of DC were achieved. Interestingly, a pure Th1-shift was only achieved, when a cellular CD40/CD40L interaction among DC took place. (s)CD40L induced almost no upregulation of maturation markers and rather resulted in a Th2-cytokine expression, such as IL-10. Correspondingly, (m)CD40L-expressing DC led to significant proliferation and stimulation of tumor-specific effector cells with increased cytotoxicity towards pancreatic, bile duct and colorectal tumor cells. Supernatants of Ad-hCD40L-transduced DC could also induce apoptosis in the different tumor cells in vitro.
Conclusion
Stimulation of the immune checkpoint CD40L/CD40 by endogenous expression of (m)CD40L provokes a cellular interaction, which increases the immunomodulatory capacity of DC. A Th1 cytokine/chemokine expression is induced, leading to a significant proliferation and enabling cytotoxicity of effector cells towards human bile duct, pancreatic and colorectal tumor cells. The present data point to the promising approach for DC-based immunotherapy of gastrointestinal malignances by activating the CD40/CD40L immune checkpoint.
Dendritic cells (DC) as professional antigen presenting cells are able to prime T-cells against the tumor-associated antigen α-fetoprotein (AFP) for immunotherapy of hepatocellular carcinoma (HCC). ...However, a strong immunosuppressive tumor environment limits their efficacy in patients. The co-stimulation with CD40Ligand (CD40L) is critical in the maturation of DC and T-cell priming. In this study, the impact of intratumoral (i.t.) CD40L-expressing DC to improve vaccination with murine (m)AFP-transduced DC (Ad-mAFP-DC) was analyzed in subcutaneous (s.c.) and orthotopic murine HCC. Murine DC were adenovirally transduced with Ad-mAFP or Ad-CD40L. Hepa129-mAFP-cells were injected into the right flank or the liver of C3H-mice to induce subcutaneous (s.c.) and orthotopic HCC. For treatments, 106 Ad-mAFP-transduced DC were inoculated s.c. followed by 106 CD40L-expressing DC injected intratumorally (i.t.). S.c. inoculation with Ad-mAFP-transduced DC, as vaccine, induced a delay of tumor-growth of AFP-positive HCC compared to controls. When s.c.-inoculation of Ad-mAFP-DC was combined with i.t.-application of Ad-CD40L-DC synergistic antitumoral effects were observed and complete remissions and long-term survival in 62% of tumor-bearing animals were achieved. Analysis of the tumor environment at different time points revealed that s.c.-vaccination with Ad-mAFP-DC seems to stimulate tumor-specific effector cells, allowing an earlier recruitment of effector T-cells and a Th1 shift within the tumors. After i.t. co-stimulation with Ad-CD40L-DC, production of Th1-cytokines was strongly increased and accompanied by a robust tumor infiltration of mature DC, activated CD4+-, CD8+-T-cells as well as reduction of regulatory T-cells. Moreover, Ad-CD40L-DC induced tumor cell apoptosis. Intratumoral co-stimulation with CD40L-expressing DC significantly improves vaccination with Ad-mAFP-DC in pre-established HCC in vivo. Combined therapy caused an early and strong Th1-shift in the tumor environment as well as higher tumor apoptosis, leading to synergistic tumor regression of HCC. Thus, CD40L co-stimulation represents a promising tool for improving DC-based immunotherapy of HCC.
Liquid biopsies do promise a lot, but are they keeping it? In the past decade, additional novel biomarkers qualified to be called like that, of which, some took necessary hurdles resulting in FDA ...approval and clinical use. Some others are since a while around, well known and were once regarded to be a game changer in cancer diagnosis or cancer screening. But, during their clinical use limitations were observed from statistical significance and questions raised regarding their robustness, that eventually led to be dropped from associated clinical guidelines for certain applications including cancer diagnosis. The purpose of this review isn't to give a broad overview of all current liquid biopsy as biomarkers, weight them and promise a brighter future in cancer prevention, but rather to take a deeper look on two of those who do qualify to be called liquid biopsies now or then. These two are probably of greatest interest conceptually and methodically, and likely have the highest chances to be in clinical use soon, with a portfolio extension over their original conceptual usage. We aim to dig deeper beyond cancer diagnosis or cancer screening. Actually, we aim to review in depth extracellular vesicles (EVs) and compare with circulating tumour cells (CTCs). The latter methodology is partially FDA approved and in clinical use. We will lay out similarities as taking advantage of surface antigens on EVs and CTCs in case of characterization and quantification. But drawing readers' attention to downstream application based on capture/isolation methodology and simply on their overall nature, here apparently being living material eventually recoverable as CTCs are vs. dead material with transient effects on recipient cell as in case of EVs. All this we try to bring in perspective, compare and conclude towards which future direction we are aiming for, or should aim for. Do we announce a winner between CTCs vs EVs? No, but we provide good reasons to intensify research on them.
PDF
