Expression Levels Of CD44 Research Articles

Significance of CD47 and Its Association With Tumor Immune Microenvironment Heterogeneity in Ovarian Cancer.

BackgroundIt was reported that tumor heterogeneity and the surrounding tumor microenvironment (TME) in ovarian cancer affects immunotherapy efficacy and patient outcomes. And the TME of ovarian cancer is intrinsically heterogeneous. CD47 plays vital roles in cell functional behavior and immune homeostasis relating to cancer prognosis. But how it affects TME and its contribution to heterogeneity in ovarian cancer has not been fully illustrated. Therefore, we aimed to identify a prognostic biomarker which may help explain tumor immune microenvironment heterogeneity of ovarian cancer.MethodsCancer single-cell state atlas (CancerSEA) was used to evaluate functional role of CD47. Several bioinformatics database including Oncomine, Gene Expression Profiling Interaction Analysis (GEPIA), Tumor Immune Estimation Resource (TIMER), The Human Protein Atlas (HPA), Ualcan and Kaplan-Meier plotter (KM plotter) were applied to illustrate correlation of CD47 with ovarian cancer prognosis and immune infiltration. Tumor Immune Single-cell Hub (TISCH) single cell database was employed to evaluate correlation of CD47 with tumor microenvironment. GeneMANIA was implemented to identify regulation networks of CD47. Differentially expressed genes (DEGs) between CD47 high and low expression groups were analyzed with R package DESeq2. Kyoto encyclopedia of genes and genomes (KEGG) and Gene Set Enrichment Analysis (GSEA) were utilized to explore how CD47 affect the immune related cell signaling pathway.ResultsCD47 expression was upregulated and connected to worse OS and PFS in ovarian cancer. Close relation was found between CD47 expression level and immune infiltration in ovarian cancer, especially with Treg cells, Monocytes, Macrophages and T cell exhaustion (P<0.05). The CD47 expression level was relatively low in plasma cells, dendritic cells and Mono/Macro cells of OV_GSE115007, in myofibroblasts, fibroblasts and endothelial cells of OV_GSE118828, compared to malignant cells of OV_GSE118828 dataset. The cell components and distribution in primary and metastatic ovarian cancer are quite distinct, which may lead to TME heterogeneity of ovarian cancer.ConclusionOur results indicated that CD47 is closely correlated to ovarian cancer immune microenvironment and might induce ovarian cancer heterogeneity. Therefore, CD47 may be used as a candidate prognostic biomarker and provide us with new insights into potential immunotherapy in ovarian cancer patients.

Chimeric antigen receptor T cells targeting CD147 for non-small cell lung cancer therapy

Non-small cell lung cancer (NSCLC) is a highly malignant tumor, with a significant mortality and morbidity. With the development of tumor immunotherapy, chimeric antigen receptor T cells (CART) gets increasingly attention and achieves prominent contributions in the treatment of hematologic malignancies. However, CART therapy for NSCLC proceeds slowly and further researches need to be investigated. In our study, we performed bioinformatics analysis to evaluate the significant role of CD147 in NSCLC. The expression level of CD147 was detected in human NSCLC cell lines and NSCLC tissues. Meanwhile, CD147-CART was constructed and identified. Cell cytotoxicity and cytokine secretion were performed to evaluate the efficacy of CD147-CART. We also constructed cell-derived xenograft (CDX) model and patient-derived xenograft (PDX) model, which was used to further investigate the safety and efficacy of CD147-CART in vivo. Our observations show that CD147 is a specific tumor antigen of NSCLC and plays an essential role in NSCLC progression, which can be used as a target for CART therapy in NSCLC. CD147-CART cells exhibit robust cytotoxicity and cytokine production in vitro, suggesting a strong anti-tumor activity against NSCLC tumor cells. Importantly, CD147-CART cells have strong anti-tumor activity against NSCLC cells in vivo in both CDX and PDX models and no adverse side effects. Our findings show that CD147-CART immunotherapy for NSCLC is safe and effective, which is an ideal and promising medical patch for treating NSCLC.

Metformin Promotes the Hair-Inductive Activity of Three-Dimensional Aggregates of Epidermal and Dermal Cells Self-Assembled in vitro

Introduction: Although it has been reported that the antidiabetic drug metformin has multiple extra-hypoglycemic activities, such as anti-oxidation, antiaging, and even antitumor, topical metformin also can induce hair regeneration, but the precise mechanism involved in that process is still unclear. Objectives: The aim of this study was to assess the effect of metformin on hair growth in a mouse hair-follicle reconstitution model generated by in vitro self-assembled three-dimensional aggregates of epidermal and dermal cells (DCs) (3D aggregates). Methods: Epidermal cells and DCs were isolated and cultured from the mouse skin of 50 C57BL/6 mouse pups (1-day-old). For tracing the distribution of DCs during the self-assembly process of 3D aggregates, the DCs were labeled with Vybrant Dil Cell-Labeling Solution and mixed with epidermal cells at a 1:1 ratio. Formed 3D aggregates were treated with 10 mM metformin and then were grafted into recipient BALB/c nude mice. The biomarkers (hepatocyte growth factor [HGF], prominin-1 [CD133], alkaline phosphatase [ALP], β-catenin, and SRY-box transcription factor 2 [SOX2]) associated with the hair-inductive activity of DCs were detected in the grafted skin tissues and in cultured 3D aggregates treated with metformin using immunofluorescent staining, quantitative real-time RT-PCR (qRT-PCR), and Western blotting. Furthermore, the expression levels of CD133 were also examined in DCs with different passage numbers using qRT-PCR and Western blotting. Results: Metformin directly stimulates the activity of ALP of cultured 3D aggregates, upregulates both the protein and mRNA expression levels of molecular markers (HGF, CD133, ALP, β-catenin, and SOX2), and improves the survival rate of reconstituted hair follicles. Moreover, we also found that metformin increases the expression of CD133 in DCs thus maintaining their trichogenic capacity that would normally be lost by serial subculture. Conclusions: These results suggest that metformin can promote hair follicle regeneration in vitro through upregulation of the hair-inductive capability of DCs, warranting further evaluation in the clinical treatment of male or female pattern hair loss.

IMMUNOHISTOCHEMICAL FEATURES IN SOFT TISSUE SARCOMAS

Sarcomas are relatively rare, accounting for only 1% of all adult malignancies and 15% of childhood malignancies [2]. According to the World Health Organization (WHO), the group of soft tissue sarcomas includes more than 100 different histological subtypes [3]. According to the recommmendations of the WHO to improve the accuracy of diagnosis in soft tissue sarcoma, the traditional histopathological examination should be supplemented with immunohistochemical and molecular methods [1]. It was carried out immunohistochemical research of CD31 receptor of Vascular Endothelial Growth Factor (VEGF) in 129 patients with STS. The age range of patients is on average 14-77 years with a peak in the age of 50-59 years. The most common reason for initial visit to a medical institution was palpable tumor (100%). It was studied the following correlative interdependences: It was determined that histodifferentiation of tumour is in inverse correlation with expression CD31 receptor. Distinctions of expression level of CD31 are shown, depending on the degree of tumor differentiation, presence or absence of its spontaneous necrotization, limphoid infiltration of pathological nidus and amount of vessels in tumor tissue. That is as high the degree of histological differentiation of tumour as low the level of expression CD31 receptor (χ2=35,4; p&lt;0,001; р=-0,322). In FHTof ST is exposed inverse dependence between expression CD31 endotelial cells with spontaneous necrosis and lymphoid infiltration. As more the size and foci of spontaneous necrosis (χ2=67,1; p&lt;0,001; р=-0,473) and lymphoid infiltration (χ2=46,1; p&lt;0,001; р=-0,346), as low the level of expression CD31 receptor. It was found that the high level of espression CD31 receptor is in direct correlation with metastasis (χ2=18,6; р=0,42; p&lt;0,001) and recurrence (χ2=9,43; р=0,30; p&lt;0,01) of the process. Studied modern approaches to the diaqnostics and treatment of malignancies. One such approach is ”Oncology Control”, which means minimizing the risk of recurrence at the local and systemic level. Following treatment, follow-up should include history and physical examination accompanied by imaging (Ultrasound or MRI) every 3 to 6 month for 2 to 3 years, and then every 6 to 12 months thereafter to asses for recurrence. So in high positivity of CD31 receptor is exposed unfavourable prognosis in patients with STS. Summarizing before said we have come to the conclusion that immune-histo-chemical investigation of CD31 receptor of VEGF has important prognostical meaning in patients STS.

Multiplex Single-Cell Analysis of Cancer Cells Enables Unbiased Uncovering Subsets Associated with Cancer Relapse: Heterogeneity of Multidrug Resistance in Precursor B-ALL.

Earlier detection of biomarkers responsible for cancer relapse facilitates more rational cancer treatment regimens to be designed. Herein, we develop a mass cytometry-based strategy for unbiased mining of cell subsets that potentially contribute to cancer recurrence through panoramic examination of the immunophenotypic features and multidrug resistance characteristics. The incorporation of metal tags enables multiplexed information of single cells to be interrogated based on metal fingerprint. Using acute lymphoblastic leukemia (B-ALL) as a showcase, we show overexpressed multidrug resistance biomarkers, i. e., BCRP, Bcl-2, MRP1, and P-gp in B-ALL cells compared with healthy control, and a positive correlation among different multidrug resistance biomarkers. Different cell subsets with multidrug resistance are well-defined, featured with CD34+ CD38+ CD10- and CD34+ CD38+/int CD10+ . Importantly, we uncovered that CD34 expression level is positively correlated to multidrug resistance, indicative of a higher potential of immature cells to induce B-ALL relapse. In addition, the cell subsets positively expressing CD73 and CD304 (CD34+ CD10+ CD304+ ; CD34+ CD38+/int CD10+ CD73+ ) also overexpress multidrug resistance biomarkers, suggesting that they may serve as additional new biomarkers for B-ALL stratification and prognosis. Our data provide the first evidence that highly expressed multidrug resistance biomarkers in certain cell subpopulations with specific immunophenotypes may potentially induce B-ALL recurrence. The incorporation of multidrug resistance features with cell phenotypes using mass cytometry proposed in this study provides a general strategy for risk assessment and the prediction of recurrence of different types of cancers.

Relevance of VEGF and CD147 in different SARS-CoV-2 positive digestive tracts characterized by thrombotic damage.

Several evidence suggests that, in addition to the respiratory tract, also the gastrointestinal tract is a main site of severe acute respiratory syndrome CoronaVirus 2 (SARS-CoV-2) infection, as an example of a multi-organ vascular damage, likely associated with poor prognosis. To assess mechanisms SARS-CoV-2 responsible of tissue infection and vascular injury, correlating with thrombotic damage, specimens of the digestive tract positive for SARS-CoV-2 nucleocapsid protein were analyzed deriving from three patients, negative to naso-oro-pharyngeal swab for SARS-CoV-2. These COVID-19-negative patients came to clinical observation due to urgent abdominal surgery that removed different sections of the digestive tract after thrombotic events. Immunohistochemical for the expression of SARS-CoV-2 combined with a panel of SARS-CoV-2 related proteins angiotensin-converting enzyme 2 receptor, cluster of differentiation 147 (CD147), human leukocyte antigen-G (HLA-G), vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 was performed. Tissue samples were also evaluated by electron microscopy for ultrastructural virus localization and cell characterization. The damage of the tissue was assessed by ultrastructural analysis. It has been observed that CD147 expression levels correlate with SARS-CoV-2 infection extent, vascular damage and an increased expression of VEGF and thrombosis. The confirmation of CD147 co-localization with SARS-CoV-2 Spike protein binding on gastrointestinal tissues and the reduction of the infection level in intestinal epithelial cells after CD147 neutralization, suggest CD147 as a possible key factor for viral susceptibility of gastrointestinal tissue. The presence of SARS-CoV-2 infection of gastrointestinal tissue might be consequently implicated in abdominal thrombosis, where VEGF might mediate the vascular damage.

Human umbilical cord mesenchymal stem cells regulate CD54 and CD105 in vascular endothelial cells and suppress inflammation in Kawasaki disease

ObjectiveThe objective was to evaluate the expression levels of CD31+CD54+ and CD31+CD105+ endothelial microparticles (EMPs) before and after intravenous immunoglobulin (IVIG) treatment of Kawasaki disease (KD). To explore the role of human umbilical cord mesenchymal stem cells (hucMSCs) in inhibiting endothelial inflammation in KD, the effects of hucMSCs on the expression of CD54 and CD105 in endothelial cells in KD were analyzed in vivo and in vitro. MethodsThe concentrations of IL-1β and VEGF in the peripheral blood of KD or healthy children were detected, and the distributions of CD31+CD54+ and CD31+CD105+ EMPs in platelet-poor plasma (PPP) were analyzed by flow cytometry. Human umbilical vein endothelial cells (HUVECs) were first cocultured with the patients’ peripheral blood mononuclear cells (PBMCs). Next, HUVECs were cocultured with hucMSCs after stimulation with inactivated serum from patients. Cell proliferation and migration activities were assessed, and the expression of CD54, CD105 and IL-1β was analyzed. In an in vivo study, hucMSCs were transplanted into KD mice. The locations and expression levels of CD54, CD105 and IL-1β in the heart tissues of mice were analyzed. ResultsThe levels of IL-1β and CD31+CD54+ EMPs were significantly higher before IVIG treatment and 2 weeks after treatment in KD patients (P < 0.01). However, the levels of VEGF and CD31+CD105+ EMPs increased significantly in KD only after IVIG treatment (P < 0.01). KD-inactivated serum stimulation combined with cocultivation of PBMCs can activate inflammation in HUVECs, leading to reduced cell proliferation and migration activities. Cocultivation also increased the expression of CD54 and decreased the expression of CD105 (P < 0.001). Cocultivation with hucMSCs can reverse these changes. Additionally, hucMSC transplantation downregulated the expression of IL-1β and CD54 and significantly upregulated the expression of CD105 in KD mice. ConclusionThe expression levels of CD31+CD54+ and CD31+CD105+ EMPs showed inconsistent changes at different KD statuses, providing potential markers for clinical application. HucMSCs suppress inflammation and regulate the expression levels of CD54 and CD105 in vascular endothelial cells in KD, possibly providing a new basis for stem cell therapy for KD.

MiR-199 Reverses the Resistance to Gemcitabine in Pancreatic Cancer by Suppressing Stemness through Regulating the Epithelial-Mesenchymal Transition.

Purpose: the present study aims to investigate the function of miR-199 on gemcitabine (GEM)-resistance in pancreatic cancer, as well as the underlying mechanism. Methods: the GEM-resistant SW1990 cell line (SW1990/SZ) was established. The CCK-8 assay was used to detect the cell viability. The self-renewal of SW1990/SZ cells was evaluated by sphere formation and the colony formation assay. The apoptosis was detected by flow cytometry and the migration ability was measured by the transwell assay. The dual-luciferase gene reporter assay was utilized to confirm the binding between miR-199 and Snail. The expression level of CD44, ALDH1, Nanog, E-cadherin, Vimentin, β-catenin, and Snail was determined by the Western blotting assay. Results: the cell sphere formation rate, number of spheres, and expression level of CD44, ALDH1, and Nanog in GEM-treated SW1990/SZ cells were significantly suppressed by miR-199, accompanied by declined proliferation ability, an increased apoptotic rate, inhibited migration ability, and suppressed EMT progression. The binding site between miR-199 and 3′-UTR of Snail was predicted and confirmed. The inhibitory effect of miR-199 on self-renewal of SW1990/GZ cells and the faciliating property of miR-199 on the inhibitory effect of GEM against the proliferation ability, migration ability, and EMT progression were abolished by overexpressing Snail. Conclusion: MiR-199 reversed the resistance to GEM in pancreatic cancer by suppressing stemness through regulating the EMT.

Effects of CD133 expression on chemotherapy and drug sensitivity of adenoid cystic carcinoma.

The cellular resistance of tumors is a major obstacle for successful tumor therapy. Cluster of differentiation (CD)133 plays an important role in the regulation of drug resistance in gastric and colon cancers. However, its effect on chemotherapeutic sensitivity in adenoid cystic carcinoma (ACC) has not been fully explored. The present study discussed the specific role of CD133 in ACC drug-resistant sensitive cells. KOA-1 cells were treated with 5-fluorouracil (5-FU) and pingyangmycin (PYM) to form drug-resistant cell lines. A Cell Counting Kit-8 assay was used to detect the cell survival rate. Cell invasion was measured using a Transwell assay. The expression levels of CD133 were detected by reverse transcription-quantitative (RT-q) PCR. The expression levels of drug-resistant mRNAs and proteins were detected by RT-qPCR and immunofluorescence analyses, respectively. The CD133 were inhibited by small interfering RNA technology. The survival rate and invasive ability of KOA-1 cells were increased following the induction of drug resistance. The expression levels of CD133, multidrug resistance protein (MDR)1 and multidrug resistance-associated protein (MRP)1 were significantly increased in drug-resistant cell lines. Knockdown of CD133 expression in the resistant cell lines, KOA-1/5-FU and KOA-1/PYM, decreased the survival rate and invasive ability. The expression levels of MDR1 and MRP1 were also significantly decreased. Knockdown of CD133 expression in ACC drug-resistant cells could inhibit the viability and invasion of tumors and enhance the sensitivity of drug-resistant cells to chemotherapeutic drugs.

CD73 Severed as a Potential Prognostic Marker and Promote Lung Cancer Cells Migration via Enhancing EMT Progression.

To investigate the expression levels and prognostic value of CD73 in lung cancer. And moreover, to identify the effect and potential mechanism of CD73 on lung cancer cells proliferation and migration. CD73 expression levels in lung cancer were analyzed base on GEPIA2 and GEO database. GEPIA2 and Kaplan-Meier Plotter (KM Plotter) was used to analyzed the correlation between CD73 expression and prognosis. GEO dataset were analyzed via GEO2R. CD73 overexpression cell model was construction via recombinant lentivirus transfection into A549 and NCI-H520 cells. CCK8 assay were used to investigate cells proliferation. Migration and invasion ability were evaluated by scratch and transwell methods. Base on GEPIA2, GSE32683, GSE116959 and GSE37745 dataset, we found that CD73 expression were significant higher in tumor tissues of lung adenocarcinoma (LUAD) compared with that in non-tumor normal tissues and in lung squamous cell carcinoma (LUSC), while there were no significant difference of CD73 expression between LUSC and normal control tissues. Interestingly, a high CD73 level predict poor overall survival (OS) of LUSC. However, GEPIA2 and KM plotter showed the opposite conclusion of prognostic value of CD73 in LUAD. By using cell experiments, we found that CD73 overexpression promoted proliferation and migration of LUAD A549 cells. However, there was no significant effect of CD73 overexpression on LUSC NCI-H520 cells. Furthermore, CD73 overexpression facilitates epithelial to mesenchymal transition (EMT) progression of A549 cells. In conclusion, our results indicated that CD73 expression were increased in LUAD and might be an poor prognostic marker for LUSC patients. CD73 play an important role in LUAD cells proliferation and migration. These data allowed to support CD73 as a therapeutic target for LUAD.

Radiomics features based on T2-weighted fluid-attenuated inversion recoveryMRI predict the expression levels of CD44 and CD133 in lower grade gliomas.

Objective: To verify the association between CD44 and CD133 expression levels and the prognosis of patients with lower grade gliomas (LGGs) and constructing radiomic models to predict those two genes'expression levels before surgery. Materials & methods: Genomic data of patients with LGGand the corresponding T2-weighted fluid-attenuated inversion recovery images were downloaded from The Cancer Genome Atlasand The Cancer Imaging Archive, which were utilized for prognosis analysis, radiomic featureextraction and modelconstruction, respectively. Results & conclusion: CD44 and CD133 expression levels in LGG can significantly affect the prognosis of patients with LGG. Based on the T2-weighted fluid-attenuated inversion recovery images, the radiomic features can effectively predict the expression levels of CD44 and CD133 before surgery.

Antiproliferative activity of CD44 siRNA-PEI-PEG nanoparticles in glioblastoma: involvement of AKT signaling.

Background and purpose:Glioblastoma multiforme (GBM) is the most invasive type of cancer which starts inside the brain. GBM cells were found to have similar properties to glioblastoma cancer stem cells. CD44 can be used as a marker of the cancer stem cells in a subset of glioblastoma tumor cells. Recent studies showed that CD44 is involved in developing cancer cells via the protein kinase B (PKB or AKT) signaling pathway. Therefore, this study aimed to investigate the CD44 mRNA silencing effects on the glioblastoma cell cycle via AKT signaling pathway.Experimental approach:To determine CD44 expression in the samples of the patients with GBM, we used the analysis of data extracted from TCGA database. qRT-PCR and western blotting were used to evaluate the expression level of genes and proteins. Different cell cycles were evaluated by DAPI staining and flow cytometry.Findings/Results:Bioinformatics results showed that CD44 expression level in GBM tumor samples is higher than in normal samples. Effects of poly (ethylene imine)-polyethylene glycol (PEI-PEG)-loaded CD44 siRNA in cell cycle showed that CD44 silencing could inhibit cell cycle in G0-G1 phase by more than 20% compared to the negative control (P < 0.05). Furthermore, PEI-PEG-loaded CD44 siRNA reduces the expression of cyclin D1 and CKD-4. According to our findings, this structure also prevented AKT phosphorylation at Thr-308 and Ser-473.Conclusion and implications:Our results suggest that PEI-PEG-loaded CD44 siRNA may attenuate the cell cycle by suppressing AKT signaling pathway.

Single-Cell Transcriptomic Analysis Revealed a Critical Role of SPP1/CD44-Mediated Crosstalk Between Macrophages and Cancer Cells in Glioma.

High-grade glioma is one of the most lethal human cancers characterized by extensive tumor heterogeneity. In order to identify cellular and molecular mechanisms that drive tumor heterogeneity of this lethal disease, we performed single-cell RNA sequencing analysis of one high-grade glioma. Accordingly, we analyzed the individual cellular components in the ecosystem of this tumor. We found that tumor-associated macrophages are predominant in the immune microenvironment. Furthermore, we identified five distinct subpopulations of tumor cells, including one cycling, two OPC/NPC-like and two MES-like cell subpopulations. Moreover, we revealed the evolutionary transition from the cycling to OPC/NPC-like and MES-like cells by trajectory analysis. Importantly, we found that SPP1/CD44 interaction plays a critical role in macrophage-mediated activation of MES-like cells by exploring the cell-cell communication among all cellular components in the tumor ecosystem. Finally, we showed that high expression levels of both SPP1 and CD44 correlate with an increased infiltration of macrophages and poor prognosis of glioma patients. Taken together, this study provided a single-cell atlas of one high-grade glioma and revealed a critical role of macrophage-mediated SPP1/CD44 signaling in glioma progression, indicating that the SPP1/CD44 axis is a potential target for glioma treatment.

Red Blood Cells from COVID-19 Patients Show Evidence of Increased Oxidative Stress and Increased Lactate Influx

Red Blood Cells from COVID-19 Patients Show Evidence of Increased Oxidative Stress and Increased Lactate InfluxCorona Disease 19 (COVID-19) is caused by SARS-CoV-2, a novel, highly infectious, single stranded RNA virus. In severe cases, excess oxidative stress produced by a ‘cytokine storm’ may generate excess reactive oxygen species (ROS) and lead to tissue damage in the lungs and elsewhere. As the potential role of RBCs in the pathophysiology of COVID-19 remains controversial (1), we investigated for evidence of increased oxidative stress and increased thrombotic tendency in RBCs from patients with COVID-19 infection.Following ethical approval and written informed consent, we used flow cytometry (BD FACSCanto II) to measure baseline RBC ROS following incubation with 2‘-7‘-dichlorofluorescein diacetate (DCF). RBC ROS were also measured following pre-incubation with hydrogen peroxide (H2O2) (2mM) +/- antioxidant N-acetyl cysteine (NAC) (0.6mM). We also measured RBC surface expression of adhesion molecules CD44, CD47 and CD242, as well as CD147. Results were expressed as mean +/- standard deviation (SD).RBC ROS were measured in 22 COVID-19 positive patients and in 10 age matched healthy controls. One patient died from respiratory failure, whilst only 3 others required ITU admission for continuous positive airway pressure (CPAP) or intubation. There was no statistical difference in mean basal RBC DCF mean fluorescence intensity (MFI) levels between COVID-19 positive patients and controls. However, mean increase in RBC DCF MFI following H2O2 incubation was significantly higher in the COVID-19 positive group (1105.7+/-336.3) compared to the control group (843.4+/-256.7)( p= 0.042). The increase in RBC DCF MFI in the COVID-19 positive group correlated with CRP (p=0.014) but not with D-dimer, serum ferritin or any complete blood count (CBC) parameters. Incubation of RBC with 0.6 mM NAC for 30 minutes prior to H2O2 exposure caused a mean reduction in DCF MFI of 26.7% in the COVID-19 positive group. RBC expression of CD44, CD47, CD242 and CD147 were measured In a separate cohort of COVID-19 positive patients (n=32), and in 22 age matched controls. There were no statistically significant differences in mean expression levels of CD44, CD47 and CD242 between the 2 groups. However, mean RBC CD147 MFI expression was higher in the COVID-19 group (1319.64+/-374.76) compared to controls (1061.59+/-253.33) (p=0.018). There was no significant correlation between RBC CD147 MFI and D-dimer, CRP, serum ferritin or any CBC parameters in the COVID-19 positive group. However, 21 of the 32 COVID-19 positive patients had blood lactate levels measured and there was a positive correlation between CD147 MFI expression and blood lactate (R=0.56, p=0.0077).Induction of oxidative stress by H2O2 resulted in a greater increase in ROS in RBCs from COVID-19 patients compared to controls and with correlation to CRP, despite the fact that there were very few patients with severe disease in the study. This suggests a role for oxidative stress in disease pathogenesis. Pre-incubation with NAC attenuated this increase in ROS, suggesting a possible role for antioxidants in therapy. Increased RBC cell surface expression of adhesion molecules CD44, CD47 and CD242 can facilitate RBC interaction with platelets and/or endothelial cells, potentially contributing to thrombosis. We found no increase in their expression in COVID-19 patients compared to controls although RBCs may contribute to thrombosis in COVID-19 infection by other means (1). CD147 is tightly associated with and enables proper expression of monocarboxylate transporter 1, the lactate transporter for RBCs. We found increased surface expression of CD147 on RBCs of COVID-19 patients, whilst CD147 expression showed a moderate correlation with serum lactate levels, suggesting that RBCs in COVID-19 infection may be acting as a lactate sink to protect against lactic acidosis.In summary, our study suggests that COVID-19 infection causes increased oxidative stress and increased lactate influx in RBCs. Further studies are warranted into the role of RBCs in COVID-19 infection.Reference:(1) Murphy P, Glavey S, Quinn J. Anemia and red blood cell abnormalities in COVID-19. Leuk Lymphoma 2021;62:1539 DisclosuresQuinn: Takeda: Honoraria. Glavey: Abbvie: Research Funding; Celgene and BMS company: Research Funding; Janssen: Honoraria, Research Funding; Amgen: Honoraria, Research Funding.

Engineered Hypoimmune Allogeneic CAR T Cells Exhibit Innate and Adaptive Immune Evasion Even after Sensitization in Humanized Mice and Retain Potent Anti-Tumor Activity

Engineered Hypoimmune Allogeneic CAR T Cells Exhibit Innate and Adaptive Immune Evasion Even after Sensitization in Humanized Mice and Retain Potent Anti-Tumor Activity

CELLULAR AND MOLECULAR EVALUATION OF IN VITRO QUALITY PARAMETERS OF HUMAN UMBILICAL CORD BLOOD-DERIVED MESENCHYMAL STEM CELLS

Introduction: Establishing a reproducible adult stem cell culture system, such as mesenchymal stem cells (MSCs), is critical for elucidating the function of molecular markers associated with these cells' undifferentiated state. In this study, we describe some important parameters to be considered for a successful isolation, culture, and characterization of MSCs from human umbilical cord blood (hUCB). Methods: Five hUCB samples were collected from healthy female subjects who were free from infectious diseases and genetic disorders. Mononuclear cells (MNCs) were counted, and viability was determined using MTT assay. MNCs were cultured in DMEM supplemented with 10% fetal bovine serum and enriched through culture and characterized using morphometric, and molecular analysis. Results: The minimum number of cells was 12.5 million and highest number of cells was 20.6 million from all ve samples. In initial culture of MSCs from hUCB, various morphologic phenotypes were seen, although the cells eventually developed a homogeneous broblast-like shape at day 14 showing &gt;80% conuency. Spindle-shaped clonogenic MSCs expressed a high level of CD90, CD105, and CD73, while negative expression of CD34. Our study provided evidence of expansion of enriched MSCs in culture from day 1 to day 21 as supported by data of CD90, CD105 and CD73 expression levels in a time-dependent manner. Conclusion: Our results suggest that the expanded hUCB harbor an enriched source of MSCs that express pluripotent stem cell markers and lack hematopoietic markers after culture and forms the basis for using hUCB as eminent source of MSCs, which can be used for different therapeutic applications.

Influence of Hyaluronan Density on the Behavior of Breast Cancer Cells with Different CD44 Expression.

Molecular gradients are common in biosystems and play an essential role in physiological and pathological processes. During carcinogenesis, for example, hyaluronan (HA) homeostasis is dysregulated by cancer cells and the altered synthesis and degradation processes result in the formation of HA gradients within the tumor microenvironment. Herein, a platform is developed to study the biological role of HA gradient in breast cancer cells. Cells with different aggressiveness and expression of CD44-the main HA receptor usually overexpressed in breast cancers, are selected for this study. The developed platform is compatible with several imaging modalities and allows assessment of cell density, morphology, CD44 expression, and cell motility in a function of HA density. Using high-throughput analysis, it is shown that cells that do not express CD44 do not change along the gradient, while CD44 positive cells respond differently to the HA gradient depending on the level of CD44 expression and HA density. This different response is associated with the activation of different signaling pathways by the CD44-HA interactions.

Investigation of the Expression of Inflammatory Markers in Oral Biofilm Samples in Patients with Systemic Scleroderma and the Association with Clinical Periodontal Parameters-A Preliminary Study.

Background: Systemic scleroderma (SSc) has multiple orofacial effects. The aim of this study was to analyze the expression of inflammatory mediators in biofilm samples. It was hypothesized that different expression levels and clinical associations might be drawn. Methods: A total of 39 biofilm samples from group 1 = SSc and group 2 = healthy control were examined for the expression levels of interleukin (IL)-2,-6, and -10; matrix metalloprotease (MMP)-9; and surface antigens CD90 and CD34 by quantitative real-time PCR and clinical parameters. Relative quantitative (RQ) gene expression was determined using the ∆∆CT method. Results: The mean bleeding on probing values (p = 0.006), clinical attachment loss (CAL) (p = 0.009), gingival recession (p = 0.020), limited mouth opening (p = 0.001) and cervical tooth defects (p = 0.011) were significantly higher in group 1. RQ expressions of IL-2 and CD34 were significantly lower, IL-6, MMP-9, and CD90 were significantly higher. There was a significant positive correlation of IL-6/MMP-9 and negative correlation of mouth opening/CAL and IL-6/CAL. Conclusion: Different expression levels of IL-2, IL-6, MMP-9, CD34 and CD90 were detected in biofilm samples from patients with SSc compared to control. An immunological correlation to the clinical parameters of mouth opening and CAL was shown; thus, we conclude that SSc might have an impact on periodontal tissues.

Relationship between CD 163 Tumor-Associated Macrophages and Colorectal-Cancer Stem Cell Markers

BACKGROUND: Colorectal-cancer stem cells (CR-CSCs) represent a specific subpopulation of colorectal cancer (CRC) cells, which are characterized by the expression of CD133 and CD166. Tumor-associated macrophages (TAMs), found near CSCs may represent polarized macrophages, which are characterized by CD163 expression. In most tumors, TAMs may promote aggressive tumor development, leading to poor prognoses. AIM: The aim of this study was to determine whether any association exists between CD163 expression in TAMs and CD133 and CD166 expression in CR-CSCs. METHODS: This study used a cross-sectional design that was conducted at the General Hospital and affiliates in Medan, from September 2018 to July 2019. CRC tissues were collected from colonoscopy biopsies and surgical resections performed on CRC patients, who fulfilled all necessary inclusion and exclusion criteria and provided informed consent. Subjects were divided into high- and low-CD163-level groups. We analyzed the expression levels of CD163, CD133, and CD166 using immunohistochemical (IHC) assays. RESULTS: A total of 118 CRC patients were enrolled in this study, of whom 58.5% were male. No significant differences in hemoglobin, leukocyte, or platelet levels were observed between high- and low-level CD163 expression. We didn’t find any significant association of CD163 TAM with CRC histological grade and TNM stagings. Significant associations were found between the CD 163 expression level and the CD133 expression level (p &lt; 0.001) and between the CD 163 expression level and the CD166 expression level (p&lt; 0.001). Increased TAM levels of CD163 was associated with 2.770-fold and 2.616-fold increased risks of elevated CD133 and CD166 levels, respectively. CONCLUSION: An association was found between the expression levels of CD163 in TAMs and the expression levels of CD133 and CD166 in CR-CSCs.

Low expression of CD24 is associated with poor survival in colorectal cancer

In this study we analyzed expression of CD24 in a cohort of colorectal cancer patients using immunohistochemistry staining of CD24. We found a significant association between absence or low expression of CD24 (10% of membranous and 55% of cytoplasmic staining) and shortened patient survival. Protein localization played a crucial role in the prognosis: membranous form was the major and prognostic one in primary tumors, while cytoplasmic expression was elevated in liver metastases compared to the primary tumors and contained prognostic information. Then, using The Cancer Genome Atlas Colon Adenocarcinoma (TCGA-COAD) RNA-seq data, we showed that CD24 mRNA level was two-fold decreased in primary colorectal cancers compared to adjacent normal mucosa. Like the protein staining data, ten percent of patients with the lowest mRNA expression levels of CD24 in primary tumors had reduced survival compared to the ones with higher expression. To explain these findings mechanistically, shRNA-mediated CD24 knockdown was performed in HT-29 colorectal cancer cells. It resulted in the increase of cell migration in vitro, no changes in proliferation and apoptosis, and a slight decrease in cell invasion. As increased cell migration is a hallmark of metastasis formation, this finding corroborates the association of a decreased CD24 expression with poor prognosis. Differential gene expression analysis revealed upregulation of genes involved in cell migration in the group of patients with low CD24 expression, including integrin subunit α3 and α3, β3 subunits of laminin 332. Further co-expression analysis identified SPI1, STAT1 and IRF1 transcription factors as putative master-regulators in this group.