Adhesion Molecule CD44 Research Articles

Expression of EGFRvIII and its co‑expression with wild‑type EGFR, or putative cancer stem cell biomarkers CD44 or EpCAM are associated with poorer prognosis in patients with hepatocellular carcinoma.

The aberrant expression of HER family members and cancer stem cells (CSCs) have been associated with tumour progression and resistance to therapy. At present, several HER inhibitors have been approved for the treatment of patients with a range of cancers but not for the treatment of patients with hepatocellular carcinoma (HCC). The present study investigated the co‑expression and prognostic significance of HER family members, type‑III deletion mutant EGFR (EGFRvIII), and the putative CSC biomarkers CD44 and epithelial cell adhesion molecule (EpCAM) in 43 patients with HCC. The relative expression of these biomarkers was determined using immunohistochemistry. At a cut off value of >5% of tumour cells stained for these biomarkers, 35% [wild‑type (wt)EGFR], 58% (HER‑2), 0% (HER‑3), 19% (HER‑4), 26% (EGFRvIII), 40% (CD44) and 33% (EpCAM) of patients were positive. In 23, 14 and 9% of the patients, wtEGFR expression was accompanied by co‑expression with HER‑2, EGFRvIII and HER‑2/EGFRvIII, respectively. EGFRvIII expression, membranous expression of CD44 and co‑expression of wtEGFR/EGFRvIII were associated with poor overall survival (OS). By contrast, cytoplasmic CD44 expression was associated with a longer OS time. The present study also investigated the effect of several agents targeting one or more members of the HER family, other growth factor receptors and cell signalling proteins on the proliferation of HCC cell lines. Among agents targeting one or more members of the HER family, the pan‑HER family blocker afatinib was the most effective, inhibiting the proliferation of three out of seven human liver cancer cell lines (LCCLs), while the CDK inhibitor dinacicilib was the most effective agent, inhibiting the proliferation of all human LCCLs tested. Taken together, the present results suggested that EGFRvIII expression and its co‑expression with wtEGFR or CD44 was of prognostic significance. These results also support further investigations of the therapeutic potential of drugs targeting EGFRvIII and other members of the HER family in patients with HCC.

Cell adhesion molecule CD44 is dispensable for reactive astrocyte activation during prion disease

Prion diseases are fatal, infectious, neurodegenerative disorders resulting from accumulation of misfolded cellular prion protein in the brain. Early pathological changes during CNS prion disease also include reactive astrocyte activation with increased CD44 expression, microgliosis, as well as loss of dendritic spines and synapses. CD44 is a multifunctional cell surface adhesion and signalling molecule which is considered to play roles in astrocyte morphology and the maintenance of dendritic spine integrity and synaptic plasticity. However, the role of CD44 in prion disease was unknown. Here we used mice deficient in CD44 to determine the role of CD44 during prion disease. We show that CD44-deficient mice displayed no difference in their response to CNS prion infection when compared to wild type mice. Furthermore, the reactive astrocyte activation and microgliosis that accompanies CNS prion infection was unimpaired in the absence of CD44. Together, our data show that although CD44 expression is upregulated in reactive astrocytes during CNS prion disease, it is dispensable for astrocyte and microglial activation and the development of prion neuropathogenesis.

Flow cytometric analysis of immune cell populations in the bronchial and mesenteric lymph nodes of the dromedary camel.

Dromedary camel is an important livestock species with special economic value in arid and semi-arid regions of the world. Given the limited data on detailed immune cell composition and cell marker expression in the dromedary camel lymph node tissue, the present study was undertaken to investigate the immune cell composition of bronchial and mesenteric lymph nodes from healthy dromedary camels using flow cytometry. In this study, we applied flow cytometry and multicolor immuno-fluorescence to phenotype the main populations of immune cells in the bronchial and mesenteric camel lymph nodes and compared them with separated peripheral blood mononuclear cells and granulocytes. We used antibodies to detect several cell surface molecules associated with camel T cells (CD4, WC1), B cells (MHCII, BAQ44A), monocytes/macrophages (CD172a, CD14, CD163), in addition to the pan-leukocyte marker CD45 and the cell adhesion molecules CD44 and CD18. Compared to blood mononuclear cells, camel lymph node cells contained a higher percentage of lymphoid cells with only a minor fraction of myeloid cells. In addition, the lower expression of CD44 and CD18 on lymph node lymphocytes compared to lymphocytes from peripheral blood indicates higher frequency of naïve lymphocytes in the lymph nodes. The frequency of CD4+ T cells, B cells and γδ T cells within camel lymph node lymphocytes compared to blood indicates a similar tissue distribution pattern of lymphocyte subsets in camel and bovine and supports previous reports on the similarity between the camel immune system and the immune system of other ruminants. Lymph node neutrophils were identified as CD45++ CD172a++, CD14+, MHCIIlow, BAQ44A+, CD44++, CD18++ cells. In conclusion, the present study is describing the employment of flow cytometric single-cell analysis and immunostaining for the analysis of the immune cell composition in the camel lymph node.

Porphyromonas gingivalis Strain W83 Infection Induces Liver Injury in Experimental Alcohol-Associated Liver Disease (ALD) in Mice

The liver plays a vital role in the defense against infections. Porphyromonas gingivalis (P. gingivalis), a dominant etiologic oral bacterium implicated in periodontal disease (PD), has been associated with various systemic diseases. This study aimed to investigate the influence of P. gingivalis on alcohol-associated liver diseases (ALD). Mice were fed a Lieber–DeCarli liquid diet containing 5% ethanol for 10 days after an initial adaptation period on a diet with lower ethanol content for 7 days. Two days before tissue sample collection, the mice were administered P. gingivalis strain W83 (Pg) through intraperitoneal injection (IP). Pair-fed mice with Pg infection (PF+Pg) exhibited an activated immune response to combat infections. However, alcohol-fed mice with Pg infection (AF+Pg) showed liver injury with noticeable abscess lesions and elevated serum alanine aminotransferase (ALT) levels. Additionally, these mice displayed liver infiltration of inflammatory monocytes and significant downregulation of proinflammatory cytokine gene expression levels; and AF+Pg mice also demonstrated increased intrahepatic neutrophil infiltration, as confirmed by chloroacetate esterase (CAE) staining, along with elevated gene expression levels of neutrophil cytosol factor 1 (Ncf1), neutrophilic inflammation driver lipocalin 2 (Lcn2), and complement component C5a receptor 1 (C5ar1), which are associated with neutrophilic inflammation. Interestingly, compared to PF+Pg mice, the livers of AF+Pg mice exhibited downregulation of gene expression levels of NADPH oxidase 2 (Cybb), the leukocyte adhesion molecule Cd18, and the Toll-like receptor adaptor Myd88. Consequently, impaired clearance of P. gingivalis and other bacteria in the liver, increased susceptibility to infections, and inflammation-associated hepatic necrotic cell death were observed in AF+Pg mice, which is likely to have facilitated immune cell infiltration and contributed to liver injury. Furthermore, in addition to the Srebf1/Fasn pathway induced by alcohol feeding, Pg infection also activated carbohydrate response element-binding protein (ChREBP) in AF+Pg mice. In summary, this study demonstrates that P. gingivalis infection, acting as a “second hit”, induces dysfunction of immune response and impairs the clearance of bacteria and infections in alcohol-sensitized livers. This process drives the development of liver injury.

Abstract 3068: CD146mediates intimate crosstalk between adipocytes and triple-negative breast cancer cells and drives tumor metastasis through lipid metabolic reprogramming

Abstract Background & Aim: In the tumor microenvironment, the adipocytes enhance the invasiveness of breast cancer (BC) by remodeling the lipid metabolism, especially in triple-negative breast cancer (TNBC). The cell adhesion molecule CD146 was highly expressed in TNBC and predicted poor distant metastasis free survival (DMFS) in patients. Meanwhile, CD146 was known to alter the obesity process by regulating the metabolic level of the body. In this study, we aimed to explore the molecular mechanism of CD146 in tumor metastasis mediating by lipid metabolic reprogramming between adipocytes and TNBC cells. Methods: CD146 knockout (KO) or overexpression cell models were constructed by CRISPR/Cas9 or lentivirus. Cells labeled with luciferase were injected in tail vein of Balb/c-nude mice and monitored lung metastasis by IVIS Kinetic system in vivo. Transwell and wound healing assays were analyzed cell aggressiveness in vitro. We synthesized anti-CD146 nanobodies, which significantly degraded CD146 protein and inhibitd its function. RNA-seq and Lipidomics in CD146 knockout models were performed to analyze the role of CD146 in TNBC cell metabolism. Immunofluorescence was used to detect the intracellular content of Lipid Droplet (LD) and protein localization. Finally, we used oleic acid (OA) and adipocyte-conditioned medium (CM) to mimic the tumor microenvironment. Results: CD146 was highly expressed in TNBC and predicted poor DMFS survival. Inhibition of CD146 suppressed tumor aggressiveness and lung metastasis in vitro and in vivo. According to the bioinformatics and RNA-Seq analysis, we showed that CD146 was intimately involved in the lipid metabolic pathway, particularly fatty acid metabolism, and revealed that inhibiting CD146 in TNBC elevated the intracellular triglyceride level through lipidomic assay. Meanwhile, knockout of CD146 promoted LD accumulation and decreased fatty acid oxidation (FAO) and mitochondrial ATP levels. In molecular mechanism, we found that adipocytes secrete high level of IL6 and fatty acid, which activated JAK/STAT3 signaling and FAO mediating by CD146 in TNBC cells. Blocking CD146 using its nanobody inhibited IL6/JAK/STAT3 activities and ACADM, leading to impairment of FAO and lung metastasis suppression in vitro and in vivo. Conclusions: In this study, we demonstrate the crucial role of CD146 in the crosstalk between tumor cells and adipocytes in the tumor microenvironment. CD146 promotes the utilization of fatty acids through fatty acid oxidation and activates the downstream inflammatory pathway response with adipocyte-secreted IL-6, thereby driving BC aggressiveness. Our study has important implications for the development of TNBC therapeutics to block CD146-driven FAO process, inflammatory response and tumor metastasis. Citation Format: Hexing Sun, Kaiyuan Huang, Bingmei Sun, Jiajia Lin, Ruiqian Yang, Jiawei Li, Shimeng Wang, De Zeng, Haoyu Lin, Yuanke Liang. CD146mediates intimate crosstalk between adipocytes and triple-negative breast cancer cells and drives tumor metastasis through lipid metabolic reprogramming [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3068.

Eosinophils exhibit a unique transcriptional signature and increased sensitivity to IL-3-induced activation in response to colorectal cancer cells.

Eosinophils have been mainly studied in allergic diseases and parasitic infections. Nonetheless, eosinophils accumulate in a variety of solid tumors, including colorectal cancer, where their presence is associated with improved prognosis. Eosinophils can promote antitumor immunity through various mechanisms, including direct cytotoxicity toward tumor cells and promoting T-cell activation. However, the mechanisms by which tumor cells regulate eosinophil activities are largely unknown. Herein, we characterized the potential interactions between eosinophils and colorectal cancer cells using an unbiased transcriptomic and proteomic analyses approach. Human eosinophils were stimulated with colorectal cancer cell conditioned media, containing tumor cell secreted factors from multiple cancer cell lines. RNA sequencing analysis identified a "core" signature consisting of 101 genes that characterize a baseline transcriptional program for the response of human eosinophils to colorectal cancer cells. Among these, the increased expression of IL-3Rα and its βc chain was identified and validated at the protein level. Secreted factors from tumor cells potentiated IL-3-induced expression of the adhesion molecule CD11a in eosinophils. Combining proteomics analysis of tumor cell secreted factors with RNA sequencing revealed potential ligand-receptor pairs between tumor cells and eosinophils and the potential involvement of the adhesion molecule CD18 and F2RL3/PAR4. Subsequent functional analyses demonstrated that F2RL3/PAR4 suppresses eosinophil migration in response tumor cell secreted factors. These findings add to the growing body of evidence that eosinophils are conditioned by their local microenvironment. Identifying mechanisms by which eosinophils interact with tumor cells could lead to the development of new immunotherapies for colorectal cancer and other solid tumors.

Engineered IRES-mediated promoter-free insulin-producing cells reverse hyperglycemia.

Endogenous insulin supplementation is essential for individuals with type 1 diabetes (T1D). However, current treatments, including pancreas transplantation, insulin injections, and oral medications, have significant limitations. The development of engineered cells that can secrete endogenous insulin offers a promising new therapeutic strategy for type 1 diabetes (T1D). This approach could potentially circumvent autoimmune responses associated with the transplantation of differentiated β-cells or systemic delivery of viral vectors. We utilized CRISPR/Cas9 gene editing coupled with homology-directed repair (HDR) to precisely integrate a promoter-free EMCVIRES-insulin cassette into the 3' untranslated region (UTR) of the GAPDH gene in human HEK-293T cells. Subsequently quantified insulin expression levels in these engineered cells, the viability and functionality of the engineered cells when seeded on different cell vectors (GelMA and Cytopore I) were also assessed. Finally, we investigated the therapeutic potential of EMCVIRES-based insulin secretion circuits in reversing Hyperglycaemia in T1D mice. Our results demonstrate that HDR-mediated gene editing successfully integrated the IRES-insulin loop into the genome of HEK-293T cells, a non-endocrine cell line, enabling the expression of human-derived insulin. Furthermore, Cytopore I microcarriers facilitated cell attachment and proliferation during in vitro culture and enhanced cell survival post-transplantation. Transplantation of these cell-laden microcarriers into mice led to the development of a stable, fat-encapsulated structure. This structure exhibited the expression of the platelet-endothelial cell adhesion molecule CD31, and no significant immune rejection was observed throughout the experiment. Diabetic mice that received the cell carriers reversed hyperglycemia, and blood glucose fluctuations under simulated feeding stimuli were very similar to those of healthy mice. In summary, our study demonstrates that Cytopore I microcarriers are biocompatible and promote long-term cell survival in vivo. The promoter-free EMCVIRES-insulin loop enables non-endocrine cells to secrete mature insulin, leading to a rapid reduction in glucose levels. We have presented a novel promoter-free genetic engineering strategy for insulin secretion and proposed an efficient cell transplantation method. Our findings suggest the potential to expand the range of cell sources available for the treatment of diabetes, offering new avenues for therapeutic interventions.

Cardiac tamponade due to advanced gastric cancer: a pathomorphological systematic review of clinical cases

Annotation. Pericardial involvement due to gastric cancer is uncommon, especially when it’s secondary to direct transdiaphragmatic tumor infiltration, and its manifestation in the form of cardiac tamponade is an exceptional situation. The aim of this review is to provide a pathomorphological overview of three rare clinical cases of cardiac tamponade, which was the first clinical manifestation of advanced gastric cancer. To realize the goal, a meta-analysis of 15 publications included in the Medline and Scopus scientific databases was conducted. The statistical substantiation of the research was carried out using Microsoft Excel 2016 and Statistica 12.0. According to the results of histological analysis, these three cases of gastric cancer were classified as: poorly differentiated adenocarcinoma of the antrum in a 52-year-old patient; sarcomatoid carcinoma of the fundus in a 49-year-old patient; and scirrhous carcinoma of the fundus in a 70-year-old patient. The tropism of gastric tumor infiltration into the pericardium is substantiated with the help of transforming growth factor, which increases the expression of CD44 adhesion molecules in tumor cells and stimulates the ability of the tumor to infiltrate the pericardium due to adhesion to the mesothelium. The hepatocyte growth factor, which enhances the invasive properties of the tumor, also played an important role: the C-Met receptors were determined on tumor cells in all cases. Thus, cardiac tamponade, which is a terminal condition, can occur in patients with various histological types of gastric cancer, including the three described. Furthermore, in the medical literature this over review is probably the only one attempt to analyze cardiac tamponade due to advanced gastric cancer with a comparison of different clinical cases.

The CNS Microenvironment Promotes Leukemia Cell Survival By Disrupting Tumour Suppression and Cell Cycle Regulation in Pediatric T-Cell Acute Lymphoblastic Leukemia

Although modern treatment has improved the overall survival of pediatric T-cell acute lymphoblastic leukemia (T-ALL) greatly, more than 20% of all children diagnosed with T-ALL relapse (Fattizzo et al., 2020). Approximately 30% of all relapse cases occur in the central nervous system (CNS) and CNS involvement is considered a major risk factor for treatment failure and increased mortality (Frishman-Levy & Izraeli, 2017). To date, CNS directed therapy is considered essential for complete cure to prevent relapse in the CNS, leading to severe neurotoxic effects and long-term adverse complications (Thastrup et al., 2022). Due to lack of knowledge regarding the mechanisms promoting leukemia survival in the CNS and absence of reliable biomarkers to predict CNS involvement, the development of new therapeutic drugs able to eradicate leukemia cells in the CNS is not progressing, therefor limiting the overall survival of pediatric T-ALL patients (Thastrup et al., 2022). To identify genetic alterations in CNS involved T-ALL we performed an RNA-sequencing analysis coupled with a gene set enrichment analysis (GSEA) studying pediatric T-ALL patient samples obtained from the TARGET initiative. We included 30 samples from patients at initial diagnosis subdivided by degree of CNS involvement, ranging from CNS1 (no CNS disease) to CNS3 (advanced CNS disease), as well as 13 samples from patients who had relapsed in the CNS. Our analysis identified distinct genetic alterations in CNS involved T-ALL and CNS relapse compared to diagnosis samples without CNS implications, with dysregulation of genes involved in tumour suppression, apoptosis and self-renewal. CNS relapsed patients were distinguished from diagnosis patients by a distinct gene expression pattern, with an upregulation of many genes, including cell cycle inhibitor CDKN2A and transcription factor HOXA3 (Figure 1A). We also found disruption of many signaling pathways, including enrichment of the p53 pathway in patients who relapsed in the CNS compared to non-CNS involved diagnosis patients. To further study the effects of CNS implications in pediatric T-ALL, we established a co-culture of pediatric T-ALL cell lines (n=4) and a glioblastoma cell line (U87-MG). Following co-culture, changes to mRNA expression was evaluated by RT-qPCR, revealing dysregulation of many genes, including an upregulation of cancer stem cell marker and cell adhesion molecule CD44 (Chen et al., 2018). Moreover, T-ALL cells grown in CNS co-culture displayed a downregulation of tumour suppressor TP53 and an upregulation of its negative regulator, proto-oncogene MDM2 when compared to cells grown in suspension (Figure 1B) (Hernandez Borrero & El-Deiry, 2021). Co-culture also induced pro-survival isoform switching of BCL2L1 (BCLX), part of the BCL family of apoptotic regulators (Daniel et al., 2013). Together, these results suggest an increased survival and loss of tumour suppression of T-ALL cells when grown in direct contact with CNS derived cells. To determine effects of the CNS microenvironment on therapy resistance, T-ALL cell lines grown in co-culture were exposed to 2µM methotrexate, a chemotherapeutic agent used during CNS directed therapy (Thastrup et al., 2022). Leukemia cells in co-culture displayed an increased survival compared to cells grown in suspension following 48 hours of methotrexate exposure, as well as dysregulation of several genes involved in cell survival, self-renewal and cell cycle regulation. Cyclins and CDKs involved in controlling cell cycle progression were differentially expressed in co-culture versus suspension, with a downregulation of CDK2 and cyclin A, crucial for S/G2 transition, as well as CDK1 and cyclin B, responsible for G2/M transition (Leal-Esteban & Fajas, 2020). These results indicate a shift towards cell cycle arrest and senescence of T-ALL cells during methotrexate exposure in CNS co-culture, potentially promoting leukemia survival and chemoresistance. Taken together, our findings indicate a differential gene expression pattern in CNS involved T-ALL, which could potentially be utilized as predicative biomarkers to properly identify patients at risk of CNS relapse. CNS co-culture revealed cell cycle alterations and loss of proliferation markers, promoting senescence and therapy resistance, which could serve as potential therapeutic targets.

Vascular Cell Adhesion Molecule-1 (VCAM-1) contributes to macular fibrosis in neovascular age-related macular degeneration through modulating macrophage functions

BackgroundNeovascular age-related macular degeneration (nAMD) is a major cause of blindness in the elderly. The disease is due to the growth of abnormal blood vessels into the macula, leading to the loss of central vision. Intravitreal injection of vascular endothelial growth factor (VEGF) inhibitors (e.g., anti-VEGF) is the standard of care for nAMD. However, nearly 50% of patients do not respond or respond poorly to the therapy. More importantly, up to 70% of nAMD patients develop macular fibrosis after 10 years of anti-VEGF therapy. The underlying mechanism of nAMD-mediated macular fibrosis is unknown although inflammation is known to play an important role in the development of abnormal macular blood vessels and its progression to fibro-vascular membrane. In this study, we measured the intraocular levels of adhesion molecule VCAM-1, ICAM-1, CD44, CD62L, and CD62P in nAMD patients with and without macular fibrosis and investigated the link between the levels of adhesion molecule and clinical features (e.g., visual improvement, retinal thickness, etc.). We further investigated the effect of VCAM-1 in macrophage function in vitro and the development of subretinal fibrosis in vivo using a two-stage laser-induced protocol.ResultsThe aqueous levels of ICAM-1, VCAM-1, CD44, and CD62L were significantly higher in nAMD patients compared to cataract controls. The aqueous level of VCAM-1 (but not other adhesion molecules) was significantly higher in patients with macular fibrosis than those without and the level correlated positively with the retinal thickness. VCAM-1 was highly expressed at the lesion site in the mouse model of subretinal fibrosis. Blocking VCAM-1 or its receptor VLA-4 significantly prevented macrophage infiltration and reduced subretinal fibrosis in vivo. VCAM-1 induced macrophage migration and upregulated the expression of Arg-1, Mmp12 and Il6 but down-regulated the expression of iNOS and Il1b in macrophages.ConclusionsVCAM-1 may contribute to the development of macular fibrosis in nAMD patients by modulating macrophage functions, including migration and profibrotic polarization.

Update on immune-based therapy strategies targeting cancer stem cells.

Accumulating data reveals that tumors possess a specialized subset of cancer cells named cancer stem cells (CSCs), responsible for metastasis and recurrence of malignancies, with various properties such as self-renewal, heterogenicity, and capacity for drug resistance. Some signaling pathways or processes like Notch, epithelial to mesenchymal transition (EMT), Hedgehog (Hh), and Wnt, as well as CSCs' surface markers such as CD44, CD123, CD133, and epithelial cell adhesion molecule (EpCAM) have pivotal roles in acquiring CSCs properties. Therefore, targeting CSC-related signaling pathways and surface markers might effectively eradicate tumors and pave the way for cancer survival. Since current treatments such as chemotherapy and radiation therapy cannot eradicate all of the CSCs and tumor relapse may happen following temporary recovery, improving novel and more efficient therapeutic options to combine with current treatments is required. Immunotherapy strategies are the new therapeutic modalities with promising results in targeting CSCs. Here, we review the targeting of CSCs by immunotherapy strategies such as dendritic cell (DC) vaccines, chimeric antigen receptors (CAR)-engineered immune cells, natural killer-cell (NK-cell) therapy, monoclonal antibodies (mAbs), checkpoint inhibitors, and the use of oncolytic viruses (OVs) in pre-clinical and clinical studies. This review will mainly focus on blood malignancies but also describe solid cancers.

Mechanotransduction via endothelial adhesion molecule CD31 initiates transmigration and reveals a role for VEGFR2 in diapedesis

Engagement of platelet endothelial cell adhesion molecule 1 (PECAM, PECAM-1, CD31) on the leukocyte pseudopod with PECAM at the endothelial cell border initiates transendothelial migration (TEM, diapedesis). We show, using fluorescence lifetime imaging microscopy (FLIM), that physical traction on endothelial PECAM during TEM initiated the endothelial signaling pathway. In this role, endothelial PECAM acted as part of a mechanotransduction complex with VE-cadherin and vascular endothelial growth factor receptor 2 (VEGFR2), and this predicted that VEGFR2 was required for efficient TEM. We show that TEM required both VEGFR2 and the ability of its Y1175 to be phosphorylated, but not VEGF or VEGFR2 endogenous kinase activity. Using inducible endothelial-specific VEGFR2-deficient mice, we show in three mouse models of inflammation that the absence of endothelial VEGFR2 significantly (by ≥75%) reduced neutrophil extravasation by selectively blocking diapedesis. These findings provide a more complete understanding of the process of transmigration and identify several potential anti-inflammatory targets.

Effects of Myrcia pubipetala Miq (Myrtaceae) extract on innate inflammatory response

Myrcia is a genus widespread in South America with many species presenting anti-inflammatory and biological properties. We investigated the anti-inflammatory activity of crude hydroalcoholic extract of Myrcia pubipetala leaves (CHE-MP) using macrophages (RAW 264.7), and the air pouch model in mice to evaluate leukocyte migration and mediator’s release. Adhesion molecule expression, CD49 and CD18, was evaluated in neutrophils. In vitro, the CHE-MP significantly reduced nitric oxide (NO), interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF) levels in the exudate and the supernatant culture. CHE-MP did not present cytotoxicity and modulated the percentage of positive neutrophils for CD18 and its expression per cell, without modifying the expression of CD49, which corroborated with significantly reduced neutrophil migration to inflammatory exudate and subcutaneous tissue. Taken together, the data demonstrate that CHE-MP presents a potential activity on innate inflammatory.

Association of high expression of CD44 in clinicopathological factors of endometrial cancer.

Clinical stages, histologic type, degree of cell differentiation, myometrial invasion, and lymph-vascular space invasion (LVSI) have been identified as clinicopathological factors that are predictive for endometrial cancer, however, further prognostic indicators are still required to account for the heterogeneity of this cancer. Adhesion molecule CD44, affects the invasion, metastasis, and prognosis of many forms of cancer. The purpose of this study is to examine the expression of CD44 in endometrial cancer and its correlation with established prognostic variables. A cross-sectional study was conducted on 64 samples of endometrial cancer from Wahidin Sudirohusodo Hospital and Hasanuddin University Hospital. Immunohistochemical analysis was used to detect CD44 expression using mouse anti-human CD44 monoclonal antibody. Differences in Histoscore were studied to determine the association between CD44 expression and clinicopathological factors of endometrial cancer. Of the overall sample, 46 samples were in the early stage, whereas 18 samples were in the advanced stage. High expression of CD44 was associated with advanced stage compare than early stage (P=0.010), poor differentiation compare than well-moderate differentiation (P=0.001), myometrial invasion ≥50% compare than myometrial invasion <50% (P=0.004), and positive LVSI compare than negative LVSI (P=0.043) in endometrial cancer, but not associated with histological type of endometrial cancer (P=0.178). High expression of CD44 may be considered as a poor prognostic marker and predictive marker for targeted therapy in endometrial cancer.

CD146‐targeted nuclear medicine imaging in cancer: State of the art

AbstractThe transmembrane glycoprotein adhesion molecule CD146 is overexpressed in a wide variety of cancers. CD146 expression constitutes a marker of poor prognosis and correlates with cancer progression. Through molecular imaging, a specific biomarker's expression and distribution can be viewed in vivo non‐invasively. Radionuclide‐labeled monoclonal antibodies or relevant fragments that target CD146 may find potential applications in cancer imaging, thereby offering tremendous value in cancer diagnosis, staging, prognosis evaluation, and prediction of drug resistance. This review discusses the recent developments of CD146‐targeted molecular imaging via nuclear medicine, especially in malignant melanoma, brain tumor, lung cancer, liver cancer, breast cancer, and pancreatic cancer. Many studies have proved that CD146 targeting may present a promising strategy for cancer theranostics.

&lt;i&gt;Mytilus edulis&lt;/i&gt; hydrolysate enhances proliferation and protects endothelial cells against hypochlorous acid-induced oxidative stress

BACKGROUND: Endothelial dysfunction underlies the pathogenesis of many socially significant diseases. The search for new original drugs for the treatment of this condition remains an important scientific and practical task. Anti-inflammatory, anticoagulant and antioxidant effects of bivalve mollusks from the family of mussels (Mytilus edulis) hydrolysate and its derivatives have been described in different model systems.&#x0D; AIM: The purpose of this study was to investigate the effect of M. edulis hydrolysate on the functional activity of EA.hy926 endothelial cell line.&#x0D; MATERIALS AND METHODS: The viability and metabolic activity of endothelial cells were studied in MTT-test. To investigate the proliferative activity, a test with staining of cells with crystal violet dye was used. The ability of the preparation to neutralize the toxic effect of HOCl and H2O2 was evaluated using fluorescent dyes and flow cytometry.&#x0D; RESULTS: It was found that the preparation did not have cytotoxicity and significantly increased the proliferation of endothelial cells in dilutions from 1:10 to 1:60. The preparation had a neutralizing effect against HOCl, and in all the studied dilutions significantly increased the viability of the endothelium. The preparation was not effective against H2O2, and increased H2O2 toxic effect in the maximal studied concentration. At the same time, the anti-inflammatory effect of M. edulis hydrolysate was not confirmed in this model system. The preparation had no effect on the IL-8 production and adhesion molecule CD54 (ICAM-1) and tissue factor CD146 the expression.&#x0D; CONCLUSIONS: The preparation of M. edulis hydrolysate enhances the proliferation of endothelial cells and is able to neutralize HOCl toxic effects.

INTERRELATION BETWEEN EXPRESSION OF BONE TISSUE REMODELING MARKERS AND MARKERS OF MOLECULAR PROFILE OF BREAST NEOPLASMS INVOLVED IN MECHANISMS OF INITIATION AND DEVELOPMENT OF BONE METASTASES

Summary. Literature data analysis proved that the expression of bone tissue remodeling proteins (osteopontin, osteonectin, BMP-7) and their impact on metastasis, particularly into bone tissue, are carried out due to direct or indirect involvement of molecular profile markers, namely steroid hormone receptors (estrogens, progesterone) and Ki-67, E-cadherin, N-cadherin, CD-44 adhesion molecules. With the appearance of malignant neoplasm, the absence or expression of each of these markers of the molecular profile reflects the orientation of the tumor process, and therefore its malignancy, and thereby determines the degree of involvement in metastatic spread of tumor cells to bones. Therefore, the most objective solution to the issue of determining the prognosis of the disease and justified treatment of patients with breast cancer can be accepted only at complex application of modern biomarkers of bone tissue remodeling, as well as a panel of markers of the molecular profi of neoplasms.

The analysis of the relationship between transferrin receptor 1 (TfR1) and clinical, morphological and immunophenotypic characteristics of breast cancer: retrospective cohort study

Background. Transferrin receptor 1 (TfR1) expression has been identified in a number of malignant tumors. It is noted that its overexpression gives growth advantages to cancer cells. Estimation of transferrin receptor expression in breast cancer (BC) might be an important component in disease prognosis, choice of treatment, also might be an attractive target for targeted therapy.&#x0D; Aim. To evaluate the expression of TfR1 by BC cells and to study its relationship with the clinical, morphological and immunophenotypic characteristics of the tumor.&#x0D; Materials and methods. This study included 82 patients with BC who received treatment at the Blokhin National Medical Research Center of Oncology (Moscow). The expression of TfR1 on primary tumor cells was studied, the relationship of TfR1 with clinical, morphological and immunophenotypic characteristics of BC was analyzed. Immunophenotyping of the primary tumor was performed by the immunohistochemical method (immunofluorescent staining) on cryostat sections. Antibodies to CD71, CD95, CD54, CD29, MUC1, Pgp170 were used. The reaction was evaluated using a luminescent microscope (AXIOSKOP, Germany). The study was dominated by patients with stage IIB 54% and IIIB 21%. Infiltrative ductal BC was diagnosed in 67% (n=55) of patients, infiltrative-lobular in 22% (n=18) of cases, other types in 11.0% (n=9).&#x0D; Results. BC cells expressed TfR1 in most cases (64.4%; n=61). A combination of TfR1 monomorphic expression with MUC1 monomorphic expression (74.4%; n=47) was noted. CD29 is presented both mosaic (38.7%) and monomorphic (51.6%). The Pgp170 antigen was monomorphically observed in 27.5% of cases. As the proportion of TfR+ cells increased, the expression frequency of the adhesion molecule CD54 increased from 10.5 to 33.3%, a positive correlation was established (r=0.293; p=0.008). In the group with TfR1 monomorphic expression, the frequency of tumors expressing the CD95 apoptosis molecule decreased: 25.0% vs 13% (p=0.042).&#x0D; Conclusion. BC cells overexpress TfR1. TfR1 expression is associated with tumor immunophenotype.

Cytokine Adsorber Use during DCD Heart Perfusion Counteracts Coronary Microvascular Dysfunction.

Microvascular dysfunction (MVD) in cardiac allografts is associated with an impaired endothelial function in the coronary microvasculature. Ischemia/reperfusion injury (IRI) deteriorates endothelial function. Hearts donated after circulatory death (DCD) are exposed to warm ischemia before initiating ex vivo blood perfusion (BP). The impact of cytokine adsorption during BP to prevent MVD in DCD hearts is unknown. In a porcine DCD model, we assessed the microvascular function of hearts after BP with (DCD-BPCytoS, n = 5) or without (DCD-BP, n = 5) cytokine adsorption (CytoSorb®). Microvascular autoregulation was assessed by increasing the coronary perfusion pressure, while myocardial microcirculation was measured by Laser-Doppler-Perfusion (LDP). We analyzed the immunoreactivity of arteriolar oxidative stress markers nitrotyrosine and 4-hydroxy-2-nonenal (HNE), endothelial injury indicating cell adhesion molecules CD54, CD106 and CD31, and eNOS. We profiled the concentration of 13 cytokines in the perfusate. The expression of 84 genes was determined and analyzed using machine learning and decision trees. Non-DCD hearts served as a control for the gene expression analysis. Compared to DCD-BP, relative LDP was improved in the DCD-BPCytoS group (1.51 ± 0.17 vs. 1.08 ± 0.17). Several pro- and anti-inflammatory cytokines were reduced in the DCD-BPCytoS group. The expression of eNOS significantly increased, and the expression of nitrotyrosine, HNE, CD54, CD106, and CD31, markers of endothelial injury, majorly decreased in the DCD-BPCytoS group. Three genes allowed exact differentiation between groups; regulation of HIF1A enabled differentiation between perfusion (DCD-BP, DCD-BPCytoS) and non-perfusion groups. CAV1 allowed differentiation between BP and BPCytoS. The use of a cytokine adsorption device during BP counteracts preload-dependent MVD and preserves the microvascular endothelium by preventing oxidative stress and IRI of coronary arterioles of DCD hearts.

Combined Genetic Ablation of CD54 and CD58 in CAR Engineered Cytotoxic Lymphocytes Effectively Averts Allogeneic Immune Cell Rejection

Allogeneic cell therapies hold promise to be cost effective with scaled manufacturing for multi-dosing and on-demand off-the-shelf availability. A critical consideration for allogeneic cell products is their ability to persist, maintain function and avoid rejection by the patient's immune system. Genetic knockout (KO) of beta-2-microglobulin (B2M) leads to complete loss of cell-surface human leukocyte antigen (HLA) class I expression and efficiently abrogates CD8+ T-cell reactivity. However, loss of HLA class I triggers NK cell-mediated missing-self recognition and manipulation of B2M must therefore be combined with other immune-modulating strategies to limit recipient NK cell reactivity. We hypothesized that rejection by the patient's immune system can be diminished in primary CAR T cells, iPSC-derived T (iT) and NK (iNK) cells by reverse-engineering common tumor escape mechanisms. The adhesion molecules CD54 and CD58 are both present at the target cell side of the immune synapse, and loss of either of these molecules have previously been reported to elicit immune escape. Here, we show that the combined deletion of CD54 and CD58 in allogeneic immune effector cells makes them resistant to rejection by recipient immune cells through unidirectional reduced synapse formation (Figure 1A). HLA class I down-regulation by B2M silencing in primary T and NK cells triggered potent cytotoxicity by resting allogeneic NK cells. This response was mostly driven by educated NK cells expressing either NKG2A or killer cell immunoglobulin-like receptors (KIR) binding to HLA-E and HLA-C, respectively. However, over-expression of HLA-E or single HLA-C ligands in a K562 screening model only shut down the specific response of the NK cell subset carrying the cognate inhibitory receptor, resulting in only partial resistance to NK cells at the bulk level. Notably, the introduction of HLA-E was particularly detrimental in donors with expanded NKG2C+ NK cell subsets, due to its stimulatory effect through the activating NKG2C receptor. In contrast, combined deletion of CD54 and CD58 in target cells uniquely decreased the response of all tested NK cell subsets and showed universal reduction across NK cell populations from 18 healthy donors (Figure 1B). To delineate the mechanisms behind the increased resistance of target cells carrying these edits, we studied NK cell-target cell interactions at the single cell level by confocal microscopy in microchips. Allogeneic NK cells formed fewer conjugates and failed to form productive immune synapses with CD54-/-CD58-/- target cells, supporting the notion that they are more resistant to NK-cell mediated killing by unidirectional altered adhesion. We next introduced these edits in primary B2M-/- T cells engineered to express a second generation CAR19 from the TRAC locus. Corroborating the K562 screen, CD54-/-CD58-/-B2M-/- CAR-T cells had a selective survival advantage over B2M-/- CAR T cells and HLA-E-over-expressing B2M-/- CAR T cells in conventional mixed lymphocyte reaction (MLR) assays in vitro. Furthermore, we established an in vivo model to probe the effect of different genetic edits on the persistence of allogeneic cell therapy products. To this end, a mixed population of B2M-/- CAR T cells additionally bearing either CD54 and/or CD58 KO, HLA-E over-expression, or no further edits were infused into mice harboring allogeneic healthy donor PBMC. We found that CD54-/-CD58-/-B2M-/- CAR T cells had significantly better in vivo persistence compared to both B2M-/- CAR T cells and HLA-E+B2M-/- CAR T cells in the presence of PBMC from healthy donors (Figure 1B). Although multiplexed editing is feasible in primary CAR T cells, the iPSC platform has an unmatched capacity for homogenously introducing multiple immune-evasion strategies for off-the-shelf cell therapy. Similar to primary CAR T cells, multiplexed edited CD54-/-CD58-/-B2M-/-CIITA-/- iNK cells showed normal growth kinetics and were resistant to rejection by activated allogeneic NK cells in MLR assays. Together, these data demonstrate that reverse-engineering of common tumor escape mechanisms, which render target cells less susceptible to immune synapse formation, is an effective strategy to avert immune rejection of allogeneic CAR T and iPSC-derived CAR NK cells. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal