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1.
Cells ; 10(12)2021 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-34944106

RESUMO

Impairment of efferocytosis in apoptotic macrophages is a known determinant of the severity of atherosclerosis and the vulnerability of plaques to rupture. The precise mechanisms involved in impaired efferocytosis are unclear. Given the well-recognized role of the inflammatory cytokine cyclophilin A (Cyp A) in modulating several atherogenic mechanisms in high-glucose primed monocytes, we investigated the role of Cyp A in macrophage efferocytosis. The efficiency of efferocytosis in RAW 264.7 macrophages grown in vitro and primed with cyclophilin A was assessed using flow cytometry and confocal assays. Cholesterol content in cells was measured using cell-based cholesterol efflux assay. Proteomic analysis and bioinformatics tools were employed to decipher the link between cyclophilin A and the known ligand receptors involved in efferocytosis. Cyclophilin A was found to impair efferocytosis in apoptotic macrophages by reducing ABCA1-mediated cholesterol efflux in foam cells derived from macrophages. Cyclophilin A-primed macrophages showed an increase in expression of the don't-eat-me signal CD 47 and a decrease in the expression of the eat-me signal, calreticulin. Phagocytosis was restored upon silencing of cyclophilin A. New Zealand white rabbits were fed a high-fat diet, and lesions in their aortae were analyzed histologically for evidence of atherosclerosis and the expression of Cyp A, CD 47 and calreticulin, the ligand receptor involved in efferocytosis. Gene and protein expressions in aortae and macrophages were analyzed by real-time PCR and Western blotting. Cyclophilin A, via its effects on the expression of CD 47 and calreticulin, impairs efferocytosis in apoptotic macrophages. Together with its impact on cholesterol efflux from macrophages, these effects can amplify other mechanisms of Cyp A in accelerating the progression of atherosclerosis.


Assuntos
Aterosclerose/patologia , Antígeno CD47/metabolismo , Ciclofilina A/metabolismo , Fagocitose , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Animais , Apoptose , Calreticulina/metabolismo , Ciclofilina A/sangue , Dieta Hiperlipídica , Regulação para Baixo , Células Espumosas/metabolismo , Camundongos , Modelos Biológicos , Células RAW 264.7 , Coelhos
2.
PLoS One ; 16(12): e0261254, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34910788

RESUMO

Cancer is one of the most important public health problems in the world. The curative effect of traditional surgery, radiotherapy and chemotherapy is limited and has inevitable side effects. As a potential target for tumor therapy, few studies have comprehensively analyzed the role of CALR in cancers. Therefore, by using GeneCards, UALCAN, GEPIA, Kaplan-Meier Plotter, COSMIC, Regulome Explorer, String, GeneMANIA and TIMER databases, we collected and analyzed relevant data to conduct in-depth bioinformatics research on the CALR expression in Pan-cancer to assess the possibility of CALR as a potential therapeutic target and survival biomarker. We studied the CALR expression in normal human tissues and various tumors of different stages, and found that CALR expression was associated with relapse free survival (RFS). We verified the expression of CALR in breast cancer cell lines by vitro experiments. Mutations of CALR were widely present in tumors. CALR interacted with different genes and various proteins. In tumors, a variety of immune cells are closely related to CALR. In conclusion, CALR can be used as a biomarker for predicting prognosis and a potential target for tumor molecular and immunotherapy.


Assuntos
Calreticulina/genética , Biologia Computacional/métodos , Neoplasias/genética , Calreticulina/metabolismo , Linhagem Celular Tumoral , Bases de Dados Genéticas , Humanos , Prognóstico
3.
Int J Mol Sci ; 22(19)2021 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-34638846

RESUMO

Previous in vitro studies have suggested that calreticulin (CALR), which is responsible for the folding and quality control of glycoproteins, may be associated with decidualization. However, its precise role in regulating decidualization has not been explored in vivo. Here, we used pregnant rat models to examine endometrial CALR expression during the peri-implantation period. We also examined whether polypectomy, a procedure that could ameliorate infertility, alters the endometrial expression levels of CALR and several implantation factors in women diagnosed as infertile. In rats, uterine CALR was expressed at a high level at the implantation site, and a marked increase in CALR expression was observed in decidual cells of normal pregnancy. In addition, endometrial CALR expression was enhanced by either administration of estradiol-17ß in the delayed implantation rat model or the artificial induction of decidualization in the pseudopregnant rat. In cultured stromal cells, siRNA-mediated silencing of CALR inhibited the decidual stimulus-induced expression of prolactin, decidual/trophoblast prolactin-related protein, and connexin 43. In humans, the endometrial expression levels of the mRNAs encoding CALR and the implantation-related factor insulin-like growth factor binding protein (IGFBP)-7 tended to increase after polypectomy. The strongest positive correlation between expression levels before polypectomy was observed for IGFBP-7 and CALR, and the strength of this correlation increased after the surgery. Thus, endometrial CALR may play a role in the formation of decidua, and the polypectomy of infertile patients may result in the co-operative expression of endometrial factors, including CALR, that could enhance endometrial receptivity.


Assuntos
Blastocisto/metabolismo , Calreticulina/genética , Decídua/metabolismo , Implantação do Embrião , Endométrio/metabolismo , Animais , Blastocisto/citologia , Calreticulina/metabolismo , Endométrio/citologia , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Infertilidade Feminina/genética , Masculino , Gravidez , Interferência de RNA , Ratos , Células Estromais/citologia , Células Estromais/metabolismo , Fatores de Tempo , Útero/metabolismo
4.
Anticancer Res ; 41(10): 4837-4855, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34593432

RESUMO

BACKGROUND/AIM: The adenovirus vector- carrying reduced expression in immortalized cell (REIC) gene (Ad-REIC) increases endoplasmic reticulum stress chaperone GRP78/BiP expression and induces the JNK-mediated apoptotic pathway. We aimed to determine whether Ad-REIC-induced apoptotic cell death can trigger immunogenic cell death (ICD). MATERIALS AND METHODS: We examined the emission of damage-associated molecular patterns in vitro and the vaccination effect in vivo. We determined the immunological changes in the tumour microenvironment by putative ICD inducers and the combined effects of immune checkpoint blockade therapies. RESULTS: Ad-REIC induced the release of high-mobility group box 1 and adenosine triphosphate and the translocation of calreticulin in murine mesothelioma AB12 cells. The vaccination effect was elicited by Ad-REIC treatment in vivo. The effect of Ad-REIC was potentiated by anti-cytotoxic T-lymphocyte-associated protein 4 antibody treatment in a murine mesothelioma AB1-HA cell model. CONCLUSION: Ad-REIC induces ICD in malignant mesothelioma.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/administração & dosagem , Vacinas Anticâncer/administração & dosagem , Morte Celular Imunogênica/efeitos dos fármacos , Mesotelioma Maligno/terapia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Trifosfato de Adenosina/metabolismo , Adenoviridae/genética , Animais , Apoptose/efeitos dos fármacos , Antígenos CD8/metabolismo , Calreticulina/metabolismo , Vacinas Anticâncer/genética , Vacinas Anticâncer/imunologia , Terapia Combinada , Terapia Genética , Vetores Genéticos , Proteína HMGB1/metabolismo , Humanos , Inibidores de Checkpoint Imunológico/imunologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Mesotelioma Maligno/imunologia , Camundongos , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Cells ; 10(8)2021 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-34440731

RESUMO

Philadelphia chromosome-negative myeloproliferative neoplasms (MPN) are clonal disorders of a hematopoietic stem cell, characterized by an abnormal proliferation of largely mature cells driven by mutations in JAK2, CALR, and MPL. All these mutations lead to a constitutive activation of the JAK-STAT signaling, which represents a target for therapy. Beyond driver ones, most patients, especially with myelofibrosis, harbor mutations in an array of "myeloid neoplasm-associated" genes that encode for proteins involved in chromatin modification and DNA methylation, RNA splicing, transcription regulation, and oncogenes. These additional mutations often arise in the context of clonal hematopoiesis of indeterminate potential (CHIP). The extensive characterization of the pathologic genome associated with MPN highlighted selected driver and non-driver mutations for their clinical informativeness. First, driver mutations are enlisted in the WHO classification as major diagnostic criteria and may be used for monitoring of residual disease after transplantation and response to treatment. Second, mutation profile can be used, eventually in combination with cytogenetic, histopathologic, hematologic, and clinical variables, to risk stratify patients regarding thrombosis, overall survival, and rate of transformation to secondary leukemia. This review outlines the molecular landscape of MPN and critically interprets current information for their potential impact on patient management.


Assuntos
Transtornos Mieloproliferativos/patologia , Calreticulina/genética , Calreticulina/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Dioxigenases/genética , Dioxigenases/metabolismo , Transplante de Células-Tronco Hematopoéticas , Humanos , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Mutação , Transtornos Mieloproliferativos/metabolismo , Transtornos Mieloproliferativos/terapia , Prognóstico , Medição de Risco , Transdução de Sinais/genética
6.
Cells ; 10(6)2021 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-34073978

RESUMO

The pregnancy-specific syndrome preeclampsia is a major cause of maternal mortality throughout the world. The initial insult resulting in the development of preeclampsia is inadequate trophoblast invasion, which may lead to reduced maternal perfusion of the placenta and placental dysfunction, such as insufficient trophoblast syncytialization. Endoplasmic reticulum (ER) stress has been implicated in the pathology of preeclampsia and serves as the major risk factor. Our previous studies suggested critical roles of calreticulin (CRT), which is an ER-resident stress response protein, in extravillous trophoblast invasion and cytotrophoblast syncytialization. Here, we studied the mechanism by which ER stress exposes the placenta to the risk of preeclampsia. We found that CRT was upregulated in the serum samples, but not in the placental specimens, from preeclamptic women. By using BeWo cells, an established model of cytotrophoblasts that syncytialize in the presence of forskolin, we demonstrated that thapsigargin-induced ER stress caused extracellular release of CRT from BeWo cells and that the extracellular CRT suppressed forskolin-induced release of ß-human chorionic gonadotropin and altered subcellular localization of E-cadherin, which is a key adhesion molecule associated with syncytialization. Our results together provide evidence that induction of ER stress leads to extracellular CRT release, which may contribute to placental dysfunction by suppressing cytotrophoblast syncytialization.


Assuntos
Calreticulina/metabolismo , Estresse do Retículo Endoplasmático , Células Gigantes/metabolismo , Trofoblastos/metabolismo , Linhagem Celular , Humanos
7.
Sci Rep ; 11(1): 13086, 2021 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-34158584

RESUMO

While ATF6α plays a central role in the endoplasmic reticulum (ER) stress response, the function of its paralogue ATF6ß remains elusive, especially in the central nervous system (CNS). Here, we demonstrate that ATF6ß is highly expressed in the hippocampus of the brain, and specifically regulates the expression of calreticulin (CRT), a molecular chaperone in the ER with a high Ca2+-binding capacity. CRT expression was reduced to ~ 50% in the CNS of Atf6b-/- mice under both normal and ER stress conditions. Analysis using cultured hippocampal neurons revealed that ATF6ß deficiency reduced Ca2+ stores in the ER and enhanced ER stress-induced death. The higher levels of death in Atf6b-/- neurons were recovered by ATF6ß and CRT overexpressions, or by treatment with Ca2+-modulating reagents such as BAPTA-AM and 2-APB, and with an ER stress inhibitor salubrinal. In vivo, kainate-induced neuronal death was enhanced in the hippocampi of Atf6b-/- and Calr+/- mice, and restored by administration of 2-APB and salubrinal. These results suggest that the ATF6ß-CRT axis promotes neuronal survival under ER stress and excitotoxity by improving intracellular Ca2+ homeostasis.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Calreticulina/metabolismo , Neurônios/metabolismo , Animais , Encéfalo , Calreticulina/fisiologia , Morte Celular/fisiologia , Sobrevivência Celular/fisiologia , Retículo Endoplasmático/metabolismo , Estresse do Retículo Endoplasmático/fisiologia , Feminino , Hipocampo , Homeostase , Ácido Caínico/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/fisiologia
8.
Cancer Sci ; 112(7): 2803-2820, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34109710

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is one of the most chemoresistant cancers. An understanding of the molecular mechanism by which PDAC cells have a high chemoresistant potential is important for improvement of the poor prognosis of patients with PDAC. Here we show for the first time that disruption of heat shock protein 47 (HSP47) enhances the efficacy of the therapeutic agent gemcitabine for PDAC cells and that the efficacy is suppressed by reconstituting HSP47 expression. HSP47 interacts with calreticulin (CALR) and the unfolded protein response transducer IRE1α in PDAC cells. Ablation of HSP47 promotes both the interaction of CALR with sarcoplasmic/endoplasmic reticulum Ca2+ -ATPase 2 and interaction of IRE1α with inositol 1,4,5-triphosphate receptor, which generates a condition in which an increase in intracellular Ca2+ level is prone to be induced by oxidative stimuli. Disruption of HSP47 enhances NADPH oxidase-induced generation of intracellular reactive oxygen species (ROS) and subsequent increase in intracellular Ca2+ level in PDAC cells after treatment with gemcitabine, resulting in the death of PDAC cells by activation of the Ca2+ /caspases axis. Ablation of HSP47 promotes gemcitabine-induced suppression of tumor growth in PDAC cell-bearing mice. Overall, these results indicated that HSP47 confers chemoresistance on PDAC cells and suggested that disruption of HSP47 may improve the efficacy of chemotherapy for patients with PDAC.


Assuntos
Calreticulina/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Resistencia a Medicamentos Antineoplásicos , Endorribonucleases/metabolismo , Proteínas de Choque Térmico HSP47/metabolismo , Neoplasias Pancreáticas/metabolismo , /metabolismo , Animais , Antimetabólitos Antineoplásicos/uso terapêutico , Cálcio/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Caspases/metabolismo , Linhagem Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/uso terapêutico , Técnicas de Inativação de Genes , Inativação Gênica , Proteínas de Choque Térmico HSP47/genética , Xenoenxertos , Humanos , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Camundongos , NADPH Oxidases/metabolismo , Transplante de Neoplasias , Neoplasias Pancreáticas/tratamento farmacológico , Espécies Reativas de Oxigênio/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Resposta a Proteínas não Dobradas
9.
Prog Mol Subcell Biol ; 59: 1-11, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34050859

RESUMO

Calreticulin is well known as an ER-resident protein that serves as the major endoplasmic reticulum (ER) Ca2+ binding protein. This protein has been the major topic of discussion in an international workshop that has been meeting for a quarter of a century. In sharing information about this protein, the field also witnessed remarkable insights into the importance of the ER as an organelle and the role of ER Ca2+ in coordinating ER and cellular functions. Recent technological advances have helped to uncover the contributions of calreticulin in maintaining Ca2+ homeostasis in the ER and to unravel its involvement in a multitude of cellular processes as highlighted in this collection of articles. The continuing revelations of unexpected involvement of calreticulin and Ca2+ in many critical aspects of cellular function promises to further improve insights into the significance of this protein in the promotion of physiology as well as prevention of pathology.


Assuntos
Calreticulina , Retículo Endoplasmático , Calreticulina/genética , Calreticulina/metabolismo , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Homeostase , Cristalino/metabolismo
10.
Prog Mol Subcell Biol ; 59: 13-25, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34050860

RESUMO

Calreticulin (Calr) is an endoplasmic reticulum (ER) chaperone involved in protein quality control, Ca2+ regulation and other cellular processes. The structure of Calr is unusual, reflecting different functions of the protein: a proline-rich ß-hairpin arm and an acidic C-terminal tail protrude from a globular core, composed of a ß-sheet sandwich and an α-helix. The arm and tail interact in the presence of Ca2+ and cover the upper ß-sheet, where a carbohydrate-binding site gives the chaperone glycoprotein affinity. At the edge of the carbohydrate-binding site is a conserved, strained disulphide bridge, formed between C106 and C137 of human Calr, which lies in a polypeptide-binding site. The lower ß-sheet has several conserved residues, comprised of a characteristic triad, D166-H170-D187, Tyr172 and the free C163. In addition to its role in the ER, Calr translocates to the cell surface upon stress and functions as an immune surveillance marker. In some myeloproliferative neoplasms, the acidic Ca2+-binding C-terminal tail is transformed into a polybasic sequence.


Assuntos
Calreticulina , Retículo Endoplasmático , Sítios de Ligação/genética , Calreticulina/genética , Calreticulina/metabolismo , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Humanos , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Transtornos Mieloproliferativos
11.
Prog Mol Subcell Biol ; 59: 145-162, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34050865

RESUMO

The endoplasmic reticulum (ER) is an organelle that mediates the proper folding and assembly of proteins destined for the cell surface, the extracellular space and subcellular compartments such as the lysosomes. The ER contains a wide range of molecular chaperones to handle the folding requirements of a diverse set of proteins that traffic through this compartment. The lectin-like chaperones calreticulin and calnexin are an important class of structurally-related chaperones relevant for the folding and assembly of many N-linked glycoproteins. Despite the conserved mechanism of action of these two chaperones in nascent protein recognition and folding, calreticulin has unique functions in cellular calcium signaling and in the immune response. The ER-related functions of calreticulin in the assembly of major histocompatibility complex (MHC) class I molecules are well-studied and provide many insights into the modes of substrate and co-chaperone recognition by calreticulin. Calreticulin is also detectable on the cell surface under some conditions, where it induces the phagocytosis of apoptotic cells. Furthermore, mutations of calreticulin induce cell transformation in myeloproliferative neoplasms (MPN). Studies of the functions of the mutant calreticulin in cell transformation and immunity have provided many insights into the normal biology of calreticulin, which are discussed.


Assuntos
Sinalização do Cálcio , Sinalização do Cálcio/genética , Calnexina/genética , Calnexina/metabolismo , Calreticulina/genética , Calreticulina/metabolismo , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Sistema Imunitário , Dobramento de Proteína
12.
Prog Mol Subcell Biol ; 59: 163-180, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34050866

RESUMO

The endoplasmic reticulum (ER) performs key cellular functions including protein synthesis, lipid metabolism and signaling. While these functions are spatially isolated in structurally distinct regions of the ER, there is cross-talk between the pathways. One vital player that is involved in ER function is the ER-resident protein calreticulin (CALR). It is a calcium ion-dependent lectin chaperone that primarily assists in glycoprotein synthesis in the ER as part of the protein quality control machinery. CALR also buffers calcium ion release and mediates other glycan-independent protein interactions. Mutations in CALR have been reported in a subset of chronic blood tumors called myeloproliferative neoplasms. The mutations consist of insertions or deletions in the CALR gene that all cause a + 1 bp shift in the reading frame and lead to a dramatic alteration of the amino acid sequence of the C-terminal domain of CALR. This alters CALR function and affects cell homeostasis. This chapter will discuss how CALR and mutant CALR affect ER health and disease.


Assuntos
Retículo Endoplasmático , Calreticulina/genética , Calreticulina/metabolismo , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Genes Reguladores , Humanos , Mutação/genética , Transtornos Mieloproliferativos/genética
13.
Prog Mol Subcell Biol ; 59: 181-196, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34050867

RESUMO

The lectin chaperones calreticulin (CALR) and calnexin (CANX), together with their co-chaperone PDIA3, are increasingly implicated in studies of human cancers in roles that extend beyond their primary function as quality control facilitators of protein folding within the endoplasmic reticulum (ER). Led by the discovery that cell surface CALR functions as an immunogen that promotes anti-tumour immunity, studies have now expanded to include their potential uses as prognostic markers for cancers, and in regulation of oncogenic signaling that regulate such diverse processes including integrin-dependent cell adhesion and migration, proliferation, cell death and chemotherapeutic resistance. The diversity stems from the increasing recognition that these proteins have an equally diverse spectrum of subcellular and extracellular localization, and which are aberrantly expressed in tumour cells. This review describes key foundational discoveries and highlight recent findings that further our understanding of the plethora of activities mediated by CALR, CANX and PDIA3.


Assuntos
Retículo Endoplasmático , Neoplasias , Biologia , Calnexina/genética , Calnexina/metabolismo , Calreticulina/genética , Calreticulina/metabolismo , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Humanos , Lectinas/genética , Lectinas/metabolismo , Neoplasias/genética , Isomerases de Dissulfetos de Proteínas/genética
14.
Blood ; 137(14): 1848-1849, 2021 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-33830192
15.
Proc Natl Acad Sci U S A ; 118(15)2021 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-33833053

RESUMO

Copy number variation (CNV) at the 16p11.2 locus is associated with neuropsychiatric disorders, such as autism spectrum disorder and schizophrenia. CNVs of the 16p gene can manifest in opposing head sizes. Carriers of 16p11.2 deletion tend to have macrocephaly (or brain enlargement), while those with 16p11.2 duplication frequently have microcephaly. Increases in both gray and white matter volume have been observed in brain imaging studies in 16p11.2 deletion carriers with macrocephaly. Here, we use human induced pluripotent stem cells (hiPSCs) derived from controls and subjects with 16p11.2 deletion and 16p11.2 duplication to understand the underlying mechanisms regulating brain overgrowth. To model both gray and white matter, we differentiated patient-derived iPSCs into neural progenitor cells (NPCs) and oligodendrocyte progenitor cells (OPCs). In both NPCs and OPCs, we show that CD47 (a "don't eat me" signal) is overexpressed in the 16p11.2 deletion carriers contributing to reduced phagocytosis both in vitro and in vivo. Furthermore, 16p11.2 deletion NPCs and OPCs up-regulate cell surface expression of calreticulin (a prophagocytic "eat me" signal) and its binding sites, indicating that these cells should be phagocytosed but fail to be eliminated due to elevations in CD47. Treatment of 16p11.2 deletion NPCs and OPCs with an anti-CD47 antibody to block CD47 restores phagocytosis to control levels. While the CD47 pathway is commonly implicated in cancer progression, we document a role for CD47 in psychiatric disorders associated with brain overgrowth.


Assuntos
Transtorno Autístico/metabolismo , Encéfalo/metabolismo , Antígeno CD47/metabolismo , Transtornos Cromossômicos/metabolismo , Deficiência Intelectual/metabolismo , Adolescente , Adulto , Animais , Transtorno Autístico/patologia , Encéfalo/patologia , Antígeno CD47/antagonistas & inibidores , Antígeno CD47/genética , Calreticulina/genética , Calreticulina/metabolismo , Linhagem Celular , Células Cultivadas , Criança , Pré-Escolar , Deleção Cromossômica , Transtornos Cromossômicos/patologia , Cromossomos Humanos Par 16/metabolismo , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Deficiência Intelectual/patologia , Masculino , Camundongos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Células Precursoras de Oligodendrócitos/citologia , Células Precursoras de Oligodendrócitos/metabolismo
16.
Int J Mol Sci ; 22(7)2021 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-33806036

RESUMO

CALR mutations are a revolutionary discovery and represent an important hallmark of myeloproliferative neoplasms (MPN), especially essential thrombocythemia and primary myelofibrosis. To date, several CALR mutations were identified, with only frameshift mutations linked to the diseased phenotype. It is of diagnostic and prognostic importance to properly define the type of CALR mutation and subclassify it according to its structural similarities to the classical mutations, a 52-bp deletion (type 1 mutation) and a 5-bp insertion (type 2 mutation), using a statistical approximation algorithm (AGADIR). Today, the knowledge on the pathogenesis of CALR-positive MPN is expanding and several cellular mechanisms have been recognized that finally cause a clonal hematopoietic expansion. In this review, we discuss the current basis of the cellular effects of CALR mutants and the understanding of its implementation in the current diagnostic laboratorial and medical practice. Different methods of CALR detection are explained and a diagnostic algorithm is shown that aids in the approach to CALR-positive MPN. Finally, contemporary methods joining artificial intelligence in accordance with molecular-genetic biomarkers in the approach to MPN are presented.


Assuntos
Calreticulina/genética , Mutação , Transtornos Mieloproliferativos/genética , Algoritmos , Animais , Inteligência Artificial , Biomarcadores/metabolismo , Calreticulina/metabolismo , Análise Mutacional de DNA , Deleção de Genes , Hematologia , Humanos , Ligantes , Aprendizado de Máquina , Chaperonas Moleculares/metabolismo , Fenótipo , Prognóstico , Transdução de Sinais , Trombocitose/metabolismo
17.
Methods Mol Biol ; 2267: 207-215, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33786794

RESUMO

Mitotic catastrophe is a modality of cell death (or occasionally senescence) that occurs after cells enter, and fail to resolve, abnormal mitosis, for instance after DNA damage or perturbations of the cell cycle. Mitotic catastrophe can avoid the generation of neoplastic cells from premalignant precursors, yet may also occur in cancer cells as a result of radiotherapy or chemotherapy. Of note, vinca alkaloids and taxanes, which are both known for affecting the stability of microtubules, can trigger mitotic catastrophe. Such agents can also cause cancer cells to undergo immunogenic cell death (ICD), which allows therapeutic responses to last beyond treatment discontinuation due to the induction of an antitumor immune response. ICD is commonly characterized by the exposure of the endoplasmic reticulum protein calreticulin on the cell surface. Here we describe an immunofluorescence-based cytofluorometric technique to detect calreticulin exposure on tumor cells exposed to drugs that induce mitotic catastrophe.


Assuntos
Calreticulina/metabolismo , Citometria de Fluxo/métodos , Morte Celular Imunogênica , Mitose , Animais , Linhagem Celular Tumoral , Humanos , Moduladores de Tubulina/toxicidade
18.
Int J Mol Sci ; 22(4)2021 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-33672997

RESUMO

The Philadelphia-negative myeloproliferative neoplasms (MPNs) are malignancies of the hematopoietic stem cell (HSC) arising as a consequence of clonal proliferation driven by somatically acquired driver mutations in discrete genes (JAK2, CALR, MPL). In recent years, along with the advances in molecular characterization, the role of immune dysregulation has been achieving increasing relevance in the pathogenesis and evolution of MPNs. In particular, a growing number of studies have shown that MPNs are often associated with detrimental cytokine milieu, expansion of the monocyte/macrophage compartment and myeloid-derived suppressor cells, as well as altered functions of T cells, dendritic cells and NK cells. Moreover, akin to solid tumors and other hematological malignancies, MPNs are able to evade T cell immune surveillance by engaging the PD-1/PD-L1 axis, whose pharmacological blockade with checkpoint inhibitors can successfully restore effective antitumor responses. A further interesting cue is provided by the recent discovery of the high immunogenic potential of JAK2V617F and CALR exon 9 mutations, that could be harnessed as intriguing targets for innovative adoptive immunotherapies. This review focuses on the recent insights in the immunological dysfunctions contributing to the pathogenesis of MPNs and outlines the potential impact of related immunotherapeutic approaches.


Assuntos
Células-Tronco Hematopoéticas/imunologia , Imunoterapia/métodos , Inflamação/imunologia , Transtornos Mieloproliferativos/terapia , Linfócitos T/imunologia , Microambiente Tumoral/imunologia , Calreticulina/genética , Calreticulina/imunologia , Calreticulina/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Humanos , Inflamação/genética , Janus Quinase 2/genética , Janus Quinase 2/imunologia , Janus Quinase 2/metabolismo , Mutação/imunologia , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/imunologia , Cromossomo Filadélfia , Linfócitos T/metabolismo , Microambiente Tumoral/genética
19.
Int J Biol Macromol ; 178: 229-239, 2021 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-33647340

RESUMO

The construction of protein abundance profiles helps to interpret the clinical applications of stem cells. Dental pulp stem cells (DPSCs) and gingival mesenchymal stem cells (GMSCs) can be isolated from teeth and used as a highly convenient clinical potential material. Here, we aimed to explore commonalities and differences of DPSCs and GMSCs at the protein level. TMT-based quantitative proteomics and two-dimensional gel electrophoresis technology were used in combination to describe the protein profile of DPSCs and GMSCs extracted from the same donor. A total of 2821 proteins were identified by LC-MS/MS, of which 248 differentially abundant proteins (DAPs) were highly expressed in GMSCs while 782 proteins were highly expressed in DPSCs. The biological functions and molecular pathways of DAPs were annotated with GO enrichment and KEGG analysis. The relationship between molecular abundance and cell characteristics including source, proliferation, angiogenesis and inflammation were connected by WGCNA. Special markers, including Calreticulin (CALR), Annexin A5 (ANXA5) and Rho GDP dissociation inhibitor alpha (GDIR1), were proposed to distinguish DPSCs from GMSCs. Our results provide a molecular basis for in-depth understanding of the protein composition and special functions of dental stem cells, and promote the potential clinical application.


Assuntos
Calreticulina/metabolismo , Polpa Dentária , Gengiva , Células-Tronco Mesenquimais , Biomarcadores , Células Cultivadas , Polpa Dentária/citologia , Polpa Dentária/metabolismo , Gengiva/citologia , Gengiva/metabolismo , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo
20.
Carbohydr Res ; 502: 108273, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33713911

RESUMO

N-glycans are attached to newly synthesised polypeptides and are involved in the folding, secretion, and degradation of N-linked glycoproteins. In particular, the calnexin/calreticulin cycle, which is the central mechanism of the entry and release of N-linked glycoproteins depending on the folding sates, has been well studied. In addition to biological studies on the calnexin/calreticulin cycle, several studies have revealed complementary roles of in vitro chemistry-based research in the structure-based understanding of the cycle. In this mini-review, we summarise chemistry-based results and highlight their importance for further understanding of the cycle.


Assuntos
Calnexina/metabolismo , Calreticulina/metabolismo , Glicoproteínas/metabolismo , Polissacarídeos/metabolismo , Calnexina/química , Calreticulina/química , Configuração de Carboidratos , Glicoproteínas/química , Polissacarídeos/química
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