RESUMEN
INTRODUCTION: Chronic hyperglycemia affects neutrophil functions, leading to reduced pathogen killing and increased morbidity. This impairment has been directly linked to increased glycemia, however, how this specifically affects neutrophils metabolism and their differentiation in the bone marrow is unclear and difficult to study. RESEARCH DESIGN AND METHODS: We used high-resolution respirometry to investigate the metabolism of resting and activated donor neutrophils, and flow cytometry to measure surface CD15 and CD11b expression. We then used HL-60 cells differentiated towards neutrophil-like cells in standard media and investigated the effect of doubling glucose concentration on differentiation metabolism. We measured the oxygen consumption rate (OCR), and the enzymatic activity of carnitine palmitoyl transferase 1 (CPT1) and citrate synthase during neutrophil-like differentiation. We compared the surface phenotype, functions, and OCR of neutrophil-like cells differentiated under both glucose concentrations. RESULTS: Donor neutrophils showed significant instability of CD11b and OCR after phorbol 12-myristate 13-acetate stimulation at 3 hours post-enrichment. During HL-60 neutrophil-like cell differentiation, there was a significant increase in surface CD15 and CD11b expression together with the loss of mitochondrial mass. Differentiated neutrophil-like cells also exhibited higher CD11b expression and were significantly more phagocytic. In higher glucose media, we measured a decrease in citrate synthase and CPT1 activities during neutrophil-like differentiation. CONCLUSIONS: HL-60 neutrophil-like differentiation recapitulated known molecular and metabolic features of human neutrophil differentiation. Increased glucose concentrations correlated with features described in hyperglycemic donor neutrophils including increased CD11b and phagocytosis. We used this model to describe metabolic features of neutrophil-like cell differentiation in hyperglycemia and show for the first time the downregulation of CPT1 and citrate synthase activity, independently of mitochondrial mass.
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Diferenciación Celular , Hiperglucemia , Neutrófilos , Humanos , Neutrófilos/metabolismo , Células HL-60 , Hiperglucemia/metabolismo , Hiperglucemia/patología , Antígeno CD11b/metabolismo , Glucosa/metabolismo , Carnitina O-Palmitoiltransferasa/metabolismo , Consumo de Oxígeno , Antígeno Lewis X/metabolismo , Citrato (si)-Sintasa/metabolismoRESUMEN
BACKGROUND: An elevated neutrophil-to-lymphocyte ratio (NLR) in peripheral blood is an independent prognostic indicator of various cancers. AIMS: In this study, we aimed to investigate the prognostic relevance of the intratumoral immune cell balance in gastric cancer. METHODS AND RESULTS: The study included 82 patients who underwent curative resection for gastric cancer. The intratumoral cluster of differentiation (CD) 15- and CD8-positive cells were evaluated using immunohistochemical staining. Additionally, clinicopathological factors and prognoses were analyzed. Patients with high intratumoral CD15/CD8 ratios had significantly lower overall survival (OS) and relapse-free survival (RFS) compared to those with low CD15/CD8 ratios (p = .0026 and p < .0001, respectively). Additionally, a high CD15/CD8 ratio was associated with lymph node metastasis (p = .019). Patients with high NLR had a significantly lower RFS than those with low NLR (p = .0050). Multivariate analysis revealed that the intratumoral CD15/CD8 ratio, NLR, and venous invasion were independent prognostic indicators of RFS (CD15/CD8 ratio: p < .001, hazard ratio (HR) = 14.7, 95% confidence interval (CI) = 3.8-56.8; NLR: p = .010, HR = 5.4, 95% CI = 1.5-19.6; venous invasion: p = .005, HR = 7.4, 95% CI = 1.8-29.7). CONCLUSION: In summary, we found that the intratumoral CD15/CD8 ratio is an independent prognostic factor following gastric cancer resection and its increase is associated with lymph node metastasis and microscopic lymph vessel invasion. Immunological evaluation with additional aspects of innate immunity may be useful in predicting cancer prognosis.
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Linfocitos T CD8-positivos , Recurrencia Local de Neoplasia , Neutrófilos , Neoplasias Gástricas , Humanos , Neoplasias Gástricas/cirugía , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/mortalidad , Neoplasias Gástricas/patología , Masculino , Femenino , Neutrófilos/inmunología , Neutrófilos/patología , Linfocitos T CD8-positivos/inmunología , Persona de Mediana Edad , Anciano , Recurrencia Local de Neoplasia/inmunología , Recurrencia Local de Neoplasia/patología , Pronóstico , Antígeno Lewis X/análisis , Antígeno Lewis X/metabolismo , Adulto , Anciano de 80 o más Años , Gastrectomía , Metástasis Linfática/patología , Linfocitos Infiltrantes de Tumor/inmunología , Estudios Retrospectivos , Supervivencia sin EnfermedadRESUMEN
Men who have sex with men (MSM) with HIV are at high risk for squamous intraepithelial lesion (SIL) and anal cancer. Identifying local immunological mechanisms involved in the development of anal dysplasia could aid treatment and diagnostics. Here, we studied 111 anal biopsies obtained from 101 MSM with HIV, who participated in an anal screening program. We first assessed multiple immune subsets by flow cytometry, in addition to histological examination, in a discovery cohort. Selected molecules were further evaluated by immunohistochemistry in a validation cohort. Pathological samples were characterized by the presence of resident memory T cells with low expression of CD103 and by changes in natural killer cell subsets, affecting residency and activation. Furthermore, potentially immunosuppressive subsets, including CD15+CD16+ mature neutrophils, gradually increased as the anal lesion progressed. Immunohistochemistry verified the association between the presence of CD15 in the epithelium and SIL diagnosis for the correlation with high-grade SIL. A complex immunological environment with imbalanced proportions of resident effectors and immune-suppressive subsets characterized pathological samples. Neutrophil infiltration, determined by CD15 staining, may represent a valuable pathological marker associated with the grade of dysplasia.
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Neoplasias del Ano , Infecciones por VIH , Antígeno Lewis X , Humanos , Masculino , Infecciones por VIH/inmunología , Infecciones por VIH/complicaciones , Infecciones por VIH/patología , Neoplasias del Ano/patología , Neoplasias del Ano/inmunología , Adulto , Persona de Mediana Edad , Antígeno Lewis X/metabolismo , Homosexualidad Masculina , Lesiones Intraepiteliales Escamosas/patología , Canal Anal/patología , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Antígenos CD/metabolismo , Neutrófilos/inmunología , Neutrófilos/patología , Neutrófilos/metabolismo , Biopsia , Inmunohistoquímica , Cadenas alfa de Integrinas/metabolismoRESUMEN
Peripheral T-cell lymphomas (PTCL) are morphologically and biologically heterogeneous and a subset expresses CD30, including anaplastic large cell lymphomas (ALCL) and a minority of PTCL, not otherwise specified (PTCL, NOS). ALCL with ALK translocations (ALCL, ALK+) are readily identified by routine diagnostic methods, but differentiating ALCL without ALK translocation (ALCL, ALK-) and PTCL, NOS expressing CD30 (PTCL CD30+) can be challenging. Furthermore, rare PTCL co-express CD30 and CD15 (PTCL CD30+CD15+); some resemble ALCL, ALK- while others resemble classic Hodgkin lymphoma. To explore the relationship between PTCL CD30+CD15+ and ALCL, ALK-, we analysed 19 cases of PTCL with CD30 expression, previously diagnosed as ALCL, ALK- (nine cases) and PTCL CD30+CD15+ (10 cases) for DUSP22/IRF4 rearrangements, coding RNA expression and selected transcriptome analysis using the NanoString nCounter gene expression analysis platform. Unsupervised clustering showed no clear segregation between ALCL, ALK- and PTCL CD30+CD15+. Three cases previously classified as PTCL CD30+CD15+ showed DUSP22/IRF4 rearrangements, favouring a diagnosis of ALCL, ALK-. Our results suggest that cases previously designated PTCL CD30+CD15+, likely fall within the spectrum of ALCL, ALK-; additionally, a subset of ALCL, ALK- with DUSP22/IRF4 rearrangement expresses CD15, consistent with previous reports and expands the immunophenotypic spectrum of this lymphoma subgroup.
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Quinasa de Linfoma Anaplásico , Antígeno Ki-1 , Antígeno Lewis X , Linfoma Anaplásico de Células Grandes , Linfoma de Células T Periférico , Femenino , Humanos , Masculino , Quinasa de Linfoma Anaplásico/genética , Quinasa de Linfoma Anaplásico/metabolismo , Fosfatasas de Especificidad Dual/genética , Reordenamiento Génico , Factores Reguladores del Interferón/genética , Factores Reguladores del Interferón/metabolismo , Antígeno Ki-1/metabolismo , Antígeno Ki-1/genética , Antígeno Ki-1/análisis , Antígeno Lewis X/análisis , Antígeno Lewis X/metabolismo , Linfoma Anaplásico de Células Grandes/genética , Linfoma Anaplásico de Células Grandes/patología , Linfoma Anaplásico de Células Grandes/diagnóstico , Linfoma de Células T Periférico/genética , Linfoma de Células T Periférico/metabolismo , Linfoma de Células T Periférico/patología , Linfoma de Células T Periférico/diagnóstico , Fosfatasas de la Proteína Quinasa Activada por Mitógenos/genéticaRESUMEN
Melanoma incidence and mortality rates are historically higher for men than women. Although emerging studies have highlighted tumorigenic roles for the male sex hormone androgen and its receptor (AR) in melanoma, cellular and molecular mechanisms underlying these sex-associated discrepancies are poorly defined. Here, we delineate a previously undisclosed mechanism by which androgen-activated AR transcriptionally upregulates fucosyltransferase 4 (FUT4) expression, which drives melanoma invasiveness by interfering with adherens junctions (AJs). Global phosphoproteomic and fucoproteomic profiling, coupled with in vitro and in vivo functional validation, further reveal that AR-induced FUT4 fucosylates L1 cell adhesion molecule (L1CAM), which is required for FUT4-increased metastatic capacity. Tumor microarray and gene expression analyses demonstrate that AR-FUT4-L1CAM-AJs signaling correlates with pathological staging in melanoma patients. By delineating key androgen-triggered signaling that enhances metastatic aggressiveness, our findings help explain sex-associated clinical outcome disparities and highlight AR/FUT4 and its effectors as potential prognostic biomarkers and therapeutic targets in melanoma.
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Melanoma , Molécula L1 de Adhesión de Célula Nerviosa , Humanos , Masculino , Femenino , Melanoma/metabolismo , Andrógenos , Molécula L1 de Adhesión de Célula Nerviosa/metabolismo , Antígeno Lewis X/metabolismo , Glicosilación , Receptores Androgénicos/genética , Receptores Androgénicos/metabolismo , Línea Celular Tumoral , Fucosiltransferasas/genética , Fucosiltransferasas/metabolismoRESUMEN
The expression of one or more of a small number of molecules, typically cell surface-associated antigens, or transcription factors, is widely used for identifying pluripotent stem cells (PSCs) or for monitoring their differentiation. However, none of these marker molecules are uniquely expressed by PSCs and all are expressed by stem cells that have lost the ability to differentiate. Consequently, none are indicators of pluripotency, per se. Here we summarize the nature and characteristics of several markers that are in wide use, including the cell surface antigens, stage-specific embryonic antigen (SSEA)-1, SSEA-3, SSEA-4, TRA-1-60, TRA-1-81, GCTM2, and the transcription factors POUF5/OCT4, NANOG, and SOX2, highlighting issues that must be considered when interpreting data about their expression on putative PSCs.
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Células Madre Pluripotentes , Células Madre Pluripotentes/metabolismo , Antígeno Lewis X/metabolismo , Diferenciación Celular , Factores de Transcripción/genética , Antígenos de Superficie/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/metabolismoRESUMEN
Inflammation is a hallmark of cancer1. In patients with cancer, peripheral blood myeloid expansion, indicated by a high neutrophil-to-lymphocyte ratio, associates with shorter survival and treatment resistance across malignancies and therapeutic modalities2-5. Whether myeloid inflammation drives progression of prostate cancer in humans remain unclear. Here we show that inhibition of myeloid chemotaxis can reduce tumour-elicited myeloid inflammation and reverse therapy resistance in a subset of patients with metastatic castration-resistant prostate cancer (CRPC). We show that a higher blood neutrophil-to-lymphocyte ratio reflects tumour myeloid infiltration and tumour expression of senescence-associated mRNA species, including those that encode myeloid-chemoattracting CXCR2 ligands. To determine whether myeloid cells fuel resistance to androgen receptor signalling inhibitors, and whether inhibiting CXCR2 to block myeloid chemotaxis reverses this, we conducted an investigator-initiated, proof-of-concept clinical trial of a CXCR2 inhibitor (AZD5069) plus enzalutamide in patients with metastatic CRPC that is resistant to androgen receptor signalling inhibitors. This combination was well tolerated without dose-limiting toxicity and it decreased circulating neutrophil levels, reduced intratumour CD11b+HLA-DRloCD15+CD14- myeloid cell infiltration and imparted durable clinical benefit with biochemical and radiological responses in a subset of patients with metastatic CRPC. This study provides clinical evidence that senescence-associated myeloid inflammation can fuel metastatic CRPC progression and resistance to androgen receptor blockade. Targeting myeloid chemotaxis merits broader evaluation in other cancers.
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Antagonistas de Receptores Androgénicos , Antineoplásicos , Quimiotaxis , Resistencia a Antineoplásicos , Células Mieloides , Neoplasias de la Próstata Resistentes a la Castración , Humanos , Masculino , Quimiotaxis/efectos de los fármacos , Progresión de la Enfermedad , Inflamación/tratamiento farmacológico , Inflamación/patología , Antígeno Lewis X/metabolismo , Células Mieloides/efectos de los fármacos , Células Mieloides/patología , Metástasis de la Neoplasia , Próstata/efectos de los fármacos , Próstata/metabolismo , Próstata/patología , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/patología , Receptores Androgénicos/metabolismo , Antagonistas de Receptores Androgénicos/farmacología , Antagonistas de Receptores Androgénicos/uso terapéutico , Antineoplásicos/farmacología , Antineoplásicos/uso terapéuticoRESUMEN
Immune checkpoint blockade (ICB) benefits some patients with triple-negative breast cancer, but what distinguishes responders from non-responders is unclear1. Because ICB targets cell-cell interactions2, we investigated the impact of multicellular spatial organization on response, and explored how ICB remodels the tumour microenvironment. We show that cell phenotype, activation state and spatial location are intimately linked, influence ICB effect and differ in sensitive versus resistant tumours early on-treatment. We used imaging mass cytometry3 to profile the in situ expression of 43 proteins in tumours from patients in a randomized trial of neoadjuvant ICB, sampled at three timepoints (baseline, n = 243; early on-treatment, n = 207; post-treatment, n = 210). Multivariate modelling showed that the fractions of proliferating CD8+TCF1+T cells and MHCII+ cancer cells were dominant predictors of response, followed by cancer-immune interactions with B cells and granzyme B+ T cells. On-treatment, responsive tumours contained abundant granzyme B+ T cells, whereas resistant tumours were characterized by CD15+ cancer cells. Response was best predicted by combining tissue features before and on-treatment, pointing to a role for early biopsies in guiding adaptive therapy. Our findings show that multicellular spatial organization is a major determinant of ICB effect and suggest that its systematic enumeration in situ could help realize precision immuno-oncology.
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Inmunoterapia , Linfocitos T , Neoplasias de la Mama Triple Negativas , Humanos , Linfocitos B/inmunología , Biopsia , Linfocitos T CD8-positivos/inmunología , Granzimas/metabolismo , Antígenos de Histocompatibilidad Clase II/inmunología , Antígeno Lewis X/metabolismo , Terapia Neoadyuvante , Medicina de Precisión , Pronóstico , Ensayos Clínicos Controlados Aleatorios como Asunto , Linfocitos T/inmunología , Neoplasias de la Mama Triple Negativas/inmunología , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/terapiaRESUMEN
Induced pluripotent stem cells (iPSCs) can differentiate into all types of cells and can be used in livestock for research on biological development, genetic breeding, and in vitro genetic resource conservation. The Bactrian camel is a large domestic animal that inhabits extreme environments and holds value in the treatment of various diseases and the development of the local economy. Therefore, we transferred four mouse genes (Oct4, Sox2, Klf4, and c-Myc) into Bactrian camel fetal fibroblasts (BCFFs) using retroviruses with a large host range to obtain Bactrian camel induced pluripotent stem cells (bciPSCs). They were comprehensively identified based on cell morphology, pluripotency gene and marker expression, chromosome number, transcriptome sequencing, and differentiation potential. The results showed the pluripotency of bciPSCs. However, unlike stem cells of other species, late formation of stem cell clones was observed; moreover, the immunofluorescence of SSEA1, SSEA3, and SSEA4 were positive, and teratoma formation took four months. These findings may be related to the extremely long gestation period and species specificity of Bactrian camels. By mining RNA sequence data, 85 potential unique pluripotent genes of Bactrian camels were predicted, which could be used as candidate genes for the production of bciPSC in the future. Among them, ASF1B, DTL, CDCA5, PROM1, CYTL1, NUP210, Epha3, and SYT13 are more attractive. In conclusion, we generated bciPSCs for the first time and obtained their transcriptome information, expanding the iPSC genetic information database and exploring the applicability of iPSCs in livestock. Our results can provide an experimental basis for Bactrian camel ESC establishment, developmental research, and genetic resource conservation.
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Células Madre Pluripotentes Inducidas , Animales , Ratones , Camelus/genética , Diferenciación Celular/genética , Animales Domésticos/metabolismo , Antígeno Lewis X/metabolismo , Proteínas de Complejo Poro Nuclear/metabolismo , Citocinas/metabolismoRESUMEN
BACKGROUND: Expression of cluster of differentiation (CD) antigens changes according to disease status and inflammation. Profiles of CD antigens expression in gastric cancer patients are different based on the status of H. pylori infection. AIMS: We conducted this study to profile CD antigen markers in gastric adenocarcinoma cells (AGS cell line) infected with distinct cytotoxin-associated gene A (cagA) genotypes of H. pylori clinical isolates. METHODS: The AGS cells were infected with H. pylori isolates with different cagA genotypes, and CD antigens expression was determined using DotScan™ antibody microarray. Formation of "hummingbird" phenotype was determined, and the percentage was calculated. RESULTS: H. pylori strains harboring cagA upregulated the expression of CD antigen involved in cancer stem cell formation (CD55), but downregulated CD antigens involved in immune regulation (CD40 and CD186) and cell adhesion (CD44). CD54 (neutrophil adhesion) and CD71 (iron transfer) were highly downregulated in the gastric cells infected with Western cagA isolates compared with East Asian isolates. CD antigen expression was different in the cells infected with H. pylori harboring different CagA EPIYA (Glu-Pro-Ile-Tyr-Ala) numbers, in which higher repression of CD54 and CD15 (Lewis x antigen) were observed in the isolate with the highest number of EPIYA motif. Furthermore, higher downregulation of CD15 was observed in the infected gastric cells with high percentage of "hummingbird" phenotype than that of low percentage of "hummingbird" phenotype. CONCLUSION: Our study demonstrated the critical roles of CD antigens in the CagA pathogenesis and should be investigated further.
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Infecciones por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Secuencias de Aminoácidos , Antígenos Bacterianos/genética , Antígenos Bacterianos/metabolismo , Antígenos CD/metabolismo , Antígenos de Superficie , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Carcinogénesis , Citotoxinas/metabolismo , Infecciones por Helicobacter/patología , Helicobacter pylori/genética , Helicobacter pylori/metabolismo , Humanos , Hierro/metabolismo , Antígeno Lewis X/metabolismoRESUMEN
Sialyl 6-sulfo Lewis X (6-sulfo sLeX) and its derivative sialyl 6-sulfo N-acetyllactosamine (LacNAc) are sialylated and sulfated glycans of sialomucins found in the high endothelial venules (HEVs) of secondary lymphoid organs. A component of 6-sulfo sLeX present in the core 1-extended O-linked glycans detected by the MECA-79 antibody was previously shown to exist in the lymphoid aggregate vasculature and bronchial mucosa of allergic and asthmatic lungs. The components of 6-sulfo sLeX in pulmonary tissues under physiological conditions remain to be analyzed. The CL40 antibody recognizes 6-sulfo sLeX and sialyl 6-sulfo LacNAc in O-linked and N-linked glycans, with absolute requirements for both GlcNAc-6-sulfation and sialylation. Immunostaining of normal mouse lungs with CL40 was performed and analyzed. The contribution of GlcNAc-6-O-sulfotransferases (GlcNAc6STs) to the synthesis of the CL40 epitope in the lungs was also elucidated. Here, we show that the expression of the CL40 epitope was specifically detected in the mesothelin-positive mesothelium of the pulmonary pleura. Moreover, GlcNAc6ST2 (encoded by Chst4) and GlcNAc6ST3 (encoded by Chst5), but not GlcNAc6ST1 (encoded by Chst2) or GlcNAc6ST4 (encoded by Chst7), are required for the synthesis of CL40-positive glycans in the lung mesothelium. Furthermore, neither GlcNAc6ST2 nor GlcNAc6ST3 is sufficient for in vivo expression of the CL40 epitope in the lung mesothelium, as demonstrated by GlcNAc6ST1/3/4 triple-knock-out and GlcNAc6ST1/2/4 triple-knock-out mice. These results indicate that CL40-positive sialylated and sulfated glycans are abundant in the pleural mesothelium and are synthesized complementarily by GlcNAc6ST2 and GlcNAc6ST3, under physiological conditions in mice.
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Antígeno Lewis X , Sulfatos , Animales , Epitelio/metabolismo , Epítopos/metabolismo , Antígeno Lewis X/metabolismo , Ratones , Oligosacáridos/metabolismo , Pleura/metabolismo , Polisacáridos/metabolismo , Antígeno Sialil Lewis XRESUMEN
N-glycans are diversified by a panel of glycosyltransferases in the Golgi, which are supposed to modify various glycoproteins in promiscuous manners, resulting in unpredictable glycosylation profiles in general. In contrast, our previous study showed that fucosyltransferase 9 (FUT9) generates Lewis X glycotopes primarily on lysosome-associated membrane protein 1 (LAMP-1) in neural stem cells. Here, we demonstrate that a contiguous 29-amino acid sequence in the N-terminal domain of LAMP-1 is responsible for promotion of the FUT9-catalyzed Lewis X modification. Interestingly, Lewis X modification was induced on erythropoietin as a model glycoprotein both in vitro and in cells, just by attaching this sequence to its C-terminus. Based on these results, we conclude that the amino acid sequence from LAMP-1 functions as a "Lewis X code", which is deciphered by FUT9, and can be embedded into other glycoproteins to evoke a Lewis X modification, opening up new possibilities for protein engineering and cell engineering.
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Fucosiltransferasas , Antígeno Lewis X , Fucosiltransferasas/genética , Glicoproteínas/metabolismo , Glicosilación , Antígeno Lewis X/genética , Antígeno Lewis X/metabolismo , Polisacáridos/metabolismoRESUMEN
Structural changes known as airway remodeling characterize chronic/severe asthma and contribute to lung dysfunction. We previously reported that neonatal SSEA-1+ pulmonary stem/progenitor cells (PSCs) ameliorated airway inflammation in asthmatic mice. However, the molecular mechanisms by which endogenous SSEA-1+ PSC of adult mice afford beneficial effects in alveolar homeostasis and lung repair after allergen challenge remain incompletely understood. To analyze the expression profile and clarify the biological significance of endogenous adult lung SSEA-1+ cells in asthmatic mice. Lung SSEA-1+ cells and circulating SSEA-1+ cells in peripheral blood were determined by confocal microscopy and cytometric analysis. GFP chimeric mice were used to trace cell lineage in vivo. The roles of circulating SSEA-1+ cells were verified in ovalbumin-induced and house dust mite-induced allergic asthmatic models. In asthmatic mice, endogenous lung SSEA-1+ cells almost disappeared; however, a unique population of circulating SSEA-1+ cells was enriched after the challenge phase. In asthmatic mice, adoptive transfer of circulating SSEA-1+ cells had a specific homing preference for the lung in response to inhaled antigen through upregulating CXCR7-CXCL11 chemokine axis. Circulating SSEA-1+ cells can transdifferentiate in the alveolar space and ameliorate lung inflammation and structural damage through inhibiting the infiltration of inflammatory cells into peribronchovascular and goblet cell hyperplasia areas, reducing the thickened smooth muscle layers and PAS-positive mucus-containing goblet cells. Reinforcing bone marrow-derived circulating SSEA-1+ cells from peripheral blood into lung tissue which create a rescue mechanism in maintaining alveolar homeostasis and tissue repair to mediate lung protection for emergency responses after allergen challenge in asthmatic conditions.
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Asma , Antígeno Lewis X , Remodelación de las Vías Aéreas (Respiratorias) , Alérgenos/metabolismo , Alérgenos/farmacología , Animales , Líquido del Lavado Bronquioalveolar , Modelos Animales de Enfermedad , Inflamación/metabolismo , Antígeno Lewis X/metabolismo , Pulmón/metabolismo , Ratones , Ratones Endogámicos BALB C , Ovalbúmina/metabolismo , Células Madre/metabolismoRESUMEN
Infection with the SARS-CoV2 virus can vary from asymptomatic, or flu-like with moderate disease, up to critically severe. Severe disease, termed COVID-19, involves acute respiratory deterioration that is frequently fatal. To understand the highly variable presentation, and identify biomarkers for disease severity, blood RNA from COVID-19 patient in an intensive care unit was analyzed by whole transcriptome RNA sequencing. Both SARS-CoV2 infection and the severity of COVID-19 syndrome were associated with up to 25-fold increased expression of neutrophil-related transcripts, such as neutrophil defensin 1 (DEFA1), and 3-5-fold reductions in T cell related transcripts such as the T cell receptor (TCR). The DEFA1 RNA level detected SARS-CoV2 viremia with 95.5% sensitivity, when viremia was measured by ddPCR of whole blood RNA. Purified CD15+ neutrophils from COVID-19 patients were increased in abundance and showed striking increases in nuclear DNA staining by DAPI. Concurrently, they showed >10-fold higher elastase activity than normal controls, and correcting for their increased abundance, still showed 5-fold higher elastase activity per cell. Despite higher CD15+ neutrophil elastase activity, elastase activity was extremely low in plasma from the same patients. Collectively, the data supports the model that increased neutrophil and decreased T cell activity is associated with increased COVID-19 severity, and suggests that blood DEFA1 RNA levels and neutrophil elastase activity, both involved in neutrophil extracellular traps (NETs), may be informative biomarkers of host immune activity after viral infection.
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Biomarcadores/sangre , COVID-19/diagnóstico , Neutrófilos/metabolismo , SARS-CoV-2/genética , Adulto , COVID-19/patología , COVID-19/virología , Femenino , Humanos , Unidades de Cuidados Intensivos , Antígeno Lewis X/metabolismo , Masculino , Persona de Mediana Edad , Activación Neutrófila , Neutrófilos/citología , Neutrófilos/inmunología , Elastasa Pancreática/sangre , ARN Viral/química , ARN Viral/metabolismo , Receptores de Antígenos de Linfocitos T/genética , SARS-CoV-2/aislamiento & purificación , Sensibilidad y Especificidad , Análisis de Secuencia de ARN , Índice de Severidad de la Enfermedad , alfa-Defensinas/genéticaRESUMEN
The phenotype of glioma-initiating cells (GIC) is modulated by cell-intrinsic and cell-extrinsic factors. Phenotypic heterogeneity and plasticity of GIC is an important limitation to therapeutic approaches targeting cancer stem cells. Plasticity also presents a challenge to the identification, isolation, and propagation of purified cancer stem cells. Here we use a barcode labelling approach of GIC to generate clonal populations over a number of passages, in combination with phenotyping using the established stem cell markers CD133, CD15, CD44, and A2B5. Using two cell lines derived from isocitrate dehydrogenase (IDH)-wildtype glioblastoma, we identify a remarkable heterogeneity of the phenotypes between the cell lines. During passaging, clonal expansion manifests as the emergence of a limited number of barcoded clones and a decrease in the overall number of clones. Dual-labelled GIC are capable of forming traceable clonal populations which emerge after as few as two passages from mixed cultures and through analyses of similarity of relative proportions of 16 surface markers we were able to pinpoint the fate of such populations. By generating tumour organoids we observed a remarkable persistence of dominant clones but also a significant plasticity of stemness marker expression. Our study presents an experimental approach to simultaneously barcode and phenotype glioma-initiating cells to assess their functional properties, for example to screen newly established GIC for tumour-specific therapeutic vulnerabilities.
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Antígenos CD/inmunología , Neoplasias Encefálicas/inmunología , Glioma/inmunología , Células Madre Neoplásicas/inmunología , Microambiente Tumoral/inmunología , Antígeno AC133/inmunología , Antígeno AC133/metabolismo , Antígenos CD/metabolismo , Biomarcadores de Tumor/inmunología , Biomarcadores de Tumor/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Células Cultivadas , Células Clonales/inmunología , Células Clonales/metabolismo , Citometría de Flujo , Glioma/metabolismo , Glioma/patología , Humanos , Receptores de Hialuranos/inmunología , Receptores de Hialuranos/metabolismo , Inmunofenotipificación , Antígeno Lewis X/inmunología , Antígeno Lewis X/metabolismo , Microscopía Confocal , Células Madre Neoplásicas/clasificación , Células Madre Neoplásicas/metabolismoRESUMEN
Lewis x functions as an adhesion molecule in glycolipids and glycoproteins since it mediates homophilic and heterophilic attachment of normal and tumoral cells. During malignancy, altered glycosylation is a frequent event; accumulating data support the expression of Lewis x in tumors although controversial results have been described including its relationship with patient survival. This report has been developed as an introduction to the relationship between Lewis x expression and breast cancer and head and neck squamous cell carcinoma (HNSCC). Results obtained in our laboratory are presented in the context of the literature.
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Neoplasias de la Mama/inmunología , Neoplasias de Cabeza y Cuello/inmunología , Antígeno Lewis X/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Biomarcadores de Tumor/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Glicosilación , Humanos , Análisis de SupervivenciaRESUMEN
The conservation of Taiwan Country chicken (TCC) is important due to concerns for the local breed's adaptability to the area and disease resistance. Furthermore, the genetic resource base of native chickens can be used to improve egg and meat production efficiency in commercial TCC. As the embryonic stem cells (ESCs) hold great potential for regenerative medicine and species conservation, the aims of this study were to isolate and characterize ESCs of TCC. The blastodermal cells (BCs) were isolated from the zona pellucida of stage X chicken embryos and cultured in conditioned medium for the proliferation and maintenance of BCs in vitro. The quantitative real-time polymerase chain reaction (qPCR) results showed that POUV, SOX2 and NANOG were expressed in the putative ESCs. In addition, the expression of pluripotent markers, SSEA-1 and SSEA-4, was detected. The DiI-stained ESCs were injected into the dorsal aorta of the E3.5 recipient fetuses soon after staining and the injected embryos were continuously incubated and checked on day 7 of incubation. It was shown that some DiI-positive cells were found in the 7-d-old chimeric embryos. The results demonstrated that some pluripotent cells existed in the cultured BCs for the production of germline chimeric embryos from TCC.
Asunto(s)
Blastodermo , Pollos , Animales , Diferenciación Celular , Embrión de Pollo , Quimera , Medios de Cultivo Condicionados/metabolismo , Células Madre Embrionarias/metabolismo , Antígeno Lewis X/metabolismo , TaiwánRESUMEN
The difficulty in treatment of glioblastoma is a consequence of its natural infiltrative growth and the existence of a population of therapy-resistant glioma cells that contribute to growth and recurrence. To identify cells more likely to have these properties, we examined the expression in tumor specimens of several protein markers important for glioma progression including the intermediate filament protein, Nestin (NES), a glucose transporter (Glut1/SLC2A1), the glial lineage marker, glial fibrillary acidic protein, and the proliferative indicator, Ki-67. We also examined the expression of von Willebrand factor, a marker for endothelial cells as well as the macrophage/myeloid markers CD163 and CD15. Using a multicolor immunofluorescence and hematoxylin and eosin staining approach with archival formalin-fixed, paraffin embedded tissue from primary, recurrent, and autopsy IDH1 wildtype specimens combined with high-resolution tissue image analysis, we have identified highly proliferative NES(+)/Glut1(-) cells that are preferentially perivascular. In contrast, Glut1(+)/NES(-) cells are distant from blood vessels, show low proliferation, and are preferentially located at the borders of pseudopalisading necrosis. We hypothesize that Glut1(+)/NES(-) cells would be naturally resistant to conventional chemotherapy and radiation due to their low proliferative capacity and may act as a reservoir for tumor recurrence.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Transportador de Glucosa de Tipo 1/metabolismo , Nestina/metabolismo , Microambiente Tumoral , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/genética , Antígenos de Diferenciación Mielomonocítica/metabolismo , Neoplasias Encefálicas/patología , Glioblastoma/patología , Transportador de Glucosa de Tipo 1/genética , Humanos , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Antígeno Lewis X/genética , Antígeno Lewis X/metabolismo , Nestina/genética , Neuroglía/metabolismo , Neuroglía/patología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Células Tumorales Cultivadas , Macrófagos Asociados a Tumores/metabolismoRESUMEN
Introduction: Protective host responses in those exposed to or infected with tuberculosis (TB) is thought to require a delicate balance between pro-inflammatory and regulatory immune responses. Myeloid-derived suppressor cells (MDSCs), regulatory cells that dampen T-cell function, have been described in cancer and other infectious diseases but there are limited data on their role in TB. Methods: Peripheral blood was obtained from patients with active pulmonary TB and participants with presumed latent TB infection (LTBI) from Cape Town, South Africa. MDSC frequency was ascertained by flow cytometry. Purified MDSCs were used to assess (i) their suppressive effect on T-cell proliferation using a Ki67 flow cytometric assay and (ii) their effect on mycobacterial containment by co-culturing with H37Rv-infected monocyte-derived macrophages and autologous pre-primed effector T-cells with or without MDSCs. Mycobacterial containment was measured by plating colony forming units (CFU). Results: MDSCs (CD15+HLA-DR-CD33+) had significantly higher median frequencies (IQR) in patients with active TB (n=10) versus LTBI (n= 10) [8.2% (6.8-10.7) versus 42.2% (27-56) respectively; p=0.001]. Compared to MDSC-depleted peripheral blood mononuclear and effector T cell populations, dilutions of purified MDSCs isolated from active TB patients suppressed T-cell proliferation by up to 72% (n=6; p=0.03) and significantly subverted effector T-cell-mediated containment of H37Rv in monocyte-derived macrophages (n=7; 0.6% versus 8.5%; p=0.02). Conclusion: Collectively, these data suggest that circulating MDSCs are induced during active TB disease and can functionally suppress T-cell proliferation and subvert mycobacterial containment. These data may inform the design of vaccines and immunotherapeutic interventions against TB but further studies are required to understand the mechanisms underpinning the effects of MDSCs.
Asunto(s)
Granulocitos/inmunología , Tuberculosis Latente/inmunología , Viabilidad Microbiana/inmunología , Mycobacterium tuberculosis/genética , Células Supresoras de Origen Mieloide/inmunología , Tuberculosis Pulmonar/inmunología , Adulto , Proliferación Celular , Técnicas de Cocultivo , Femenino , Antígenos HLA-DR/metabolismo , Humanos , Hidrolasas/inmunología , Tuberculosis Latente/sangre , Tuberculosis Latente/epidemiología , Tuberculosis Latente/microbiología , Antígeno Lewis X/metabolismo , Macrófagos/inmunología , Masculino , Persona de Mediana Edad , Mycobacterium tuberculosis/aislamiento & purificación , Datos Preliminares , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismo , Sudáfrica/epidemiología , Linfocitos T/inmunología , Tuberculosis Pulmonar/sangre , Tuberculosis Pulmonar/epidemiología , Tuberculosis Pulmonar/microbiologíaRESUMEN
PURPOSE: The identification of prognostically and therapeutically relevant molecular markers is fundamental to the further development of personalised therapies in brain tumours. Current therapeutic options for the treatment of gliomas rely mainly on surgical resection and the inhibition of tumour cell proliferation by irradiation and chemotherapy. Glioma stem cells are a subpopulation of proliferating tumour cells that have self-renewal capacity and can give rise to heterogeneous cells that comprise the tumour and are thought to play a role in the resistance of gliomas to therapy. The aim of this study was to evaluate the expression of markers of glioma stem cells and differentiated glial cells in proliferating glioma cells in comparison to the overall expression of the respective markers in the tumour tissue. METHODS: Tissue microarrays were assembled from specimen of pilocytic astrocytoma, diffuse astrocytoma, anaplastic astrocytoma, glioblastoma, oligodendroglioma, anaplastic oligodendroglioma, ependymoma, and anaplastic ependymoma. These were immunohistochemically double stained with antibodies against the proliferation-associated antigen Ki67 and marker proteins for glioma stem cells (CD133, Nestin, Musashi, CD15, CD44), and differentiated glioma cells (GFAP, MAP2c). RESULTS: The expression of both glial and glioma stem cell markers differs between proliferating and non-proliferating glioma cells. Furthermore, the proliferating cells in the different glial tumour entities show a different expression profile. CONCLUSION: Further analysis of marker expression in proliferating glioma cells and correlation with clinical outcome and susceptibility to irradiation and chemotherapy might help establish new biomarkers and therapies for glioma.