Regulatory T Lymphocytes (Treg): Modulation and Clinical Application

Treg cells CD4+CD25+FOXP3+ have a specific function in the tolerance of autoantigens and regulation of the immune response. Modulation of differentiation pathways and the use of Treg cells in cell therapy have been reported in autoimmune diseases, systemic lupus erythromatosis, autoimmune hepatitis, type 1 diabetes mellitus, multiple sclerosis, rheumatoid arthritis, graft-versus-host disease, bone marrow transplantation and solid organs. The expansion of Treg cells in vivo occurs through low-dose IL-2 treatment. However, because of the heterogeneity and variability of Treg cells, the isolation of peripheral blood cells, through the technique of leucopheresis by GMP (good manuring practice), for in vitro expansion is difficult, necessitating a large combination of specific and reliable cellular markers. Currently, two specific markers, Helios and neuropilin-1, are being studied to facilitate the differentiation of thymus Treg cells and peripheral Treg cells. However, Treg cells induced in vitro are unstable. Modulation of the FOXP3 gene in the CNS1 and CNS2 region is an alternative to maintaining the stability of expanded Treg cells in vitro

Hepcidin: SNP-Like Polymorphisms Present in Iron Metabolism and Clinical Complications of Iron Accumulation and Deficiency

The metabolism of iron is regulated by the peptide hormone hepcidin. Genetic alterations in the proteins involved in the signalling pathway and hepcidin transcription cause damage to the organism. Mutations and polymorphisms in the hepcidin antimicrobial peptide(HAMP), HFE, HJV, ferroportin and matriptase-2 genes influence serum hepcidin concentration. Genetic deficiency of hepcidin increases iron overload in tissues, leading to haemochromatosis. However, genetics changes in the TMPRSS6 gene promote an increase in serum hepcidin, with the development of severe anaemia and resistance to iron treatment, as observed in IRIDA. Making the flow and efflux of extracellular and intracellular iron is impossible. To date, no drug that works by inhibiting or enhancing hepcidin transcription is available, largely because of the cytotoxicity described in vitro models. The proposed therapeutic targets are still in the early stages of clinical trials, some are good candidates, such as heparin derivatives and mini-hepcidins

Influence of Hepcidin in the Development of Anemia

Anemia presents a global public health problem. It is related to several factors, ranging from deficiency in nutrients from food to genetic alterations in iron absorption and metabolism. In this context, hepcidin is a peptide molecule that regulates iron homeostasis. Hepcidin is synthesized, in part, by hepatocytes. In physiological conditions, increased serum transferrin, serum iron, inflammation, and erythropoiesis trigger stimuli that promote hepcidin antimicrobial peptide (HAMP) gene transcription and hepcidin synthesis. However, in pathological situations, an overexpression of hepcidin occurs, an increase in the plasma concentration that damages the organism. Hepcidin contributes to the pathogenesis of iron deficiency anemia, anemia of inflammation, in hemoglobinopathies. Then, there is a restriction of the availability of iron to the tissues and the formation of new erythroid precursors, with the consequent development of anemia

A influência da hepcidina em aspectos do metabolismo do ferro de indivíduos portadores do vírus HIV-1

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências da Saúde, Programa de Pós-Graduação em Farmácia, Florianópolis, 2018.A hepcidina-codificada pelo gene HAMP-atua no controle da absorção, circulação e armazenamento de ferro no organismo. Estudos sugerem que os estoques de ferro celular influenciam no curso e progressão da infecção pelo vírus HIV-1. Portanto, foi avaliado, a influência da hepcidina no metabolismo do ferro em 199 indivíduos portadores do vírus HIV-1; bem como, verificou-se a frequência das variantes, c.251A>G e c.212G>A do gene HAMP em 191 indivíduos HIV-1 e 100 indivíduos soronegativos para HIV-1, através da técnica de PCR em tempo real. A concentração de hepcidina foi determinada por ELISA. O valor de referência estabelecido para hepcidina foi correlacionado com biomarcadores do metabolismo do ferro, com o hemograma, leucograma e marcadores de infecção pelo HIV-1. A mediana da concentração de hepcidina em indivíduos saudáveis foi de 235,5 µg/L e o intervalo de referência estabelecido entre 114?477 µg/L. As concentrações séricas de hepcidina foram semelhantes entre o grupo de indivíduos soronegativo 235,5(114?477) µg/L e grupo ART-naive 274(128?409) µg/L, enquanto que nos grupos fazendo uso da TARV a concentração foi elevada, INNTR/INTR com 444 (124?580) µg/L e INTR/IP com 412(185?548) µg/L. No grupo ART-naive a concentração de hepcidina teve uma correlação com a concentração de ferro (r=0,278; pG foi de 47,7% em indivíduos portadores do vírus HIV-1 quando comparado aos indivíduos soronegativos para HIV-1, foi de 39,0%. O genótipo AG foi relacionado ao aumento da concentração sérica de hepcidina, tanto em indivíduos soronegativos quanto em indivíduos portadores do vírus HIV-1. De acordo com os resultados obtidos, a concentração sérica de hepcidina influencia no metabolismo do ferro de indivíduos HIV-1 e contribui para o desenvolvimento de alterações hematológicas.Abstract : The hepcidin protein, synthesized in the liver and encoded by the HAMP gene, acts to control the absorption, circulation and storage of iron in the body. Different studies have suggested that cellular iron stores influence the course and progression of human immunodeficiency virus type-1 (HIV-1) infection. In this context, the influence of hepcidin on iron metabolism in 199 HIV-1 virus carriers was evaluated; as well as the frequency of variants, c.251A> G and c.212G> A, of the HAMP gene in 191 HIV-1 individuals and 100 HIV-1 seronegative individuals were verified by real polymerase chain reaction. Serum hepcidin concentration was determined by enzyme immunoassay. A reference value for hepcidin was established in order to correlate the serum concentration of hepcidin with biomarkers of iron metabolism, with the blood count, leukogram and markers of HIV-1 infection. The median serum hepcidin concentration in healthy subjects was 235.5 µg /L and the reference range established was between 114-477 µg/L. Serum hepcidin concentrations were similar between the 235.5 (114-477) µg/L seronegative group and the ART-naive 274 (128-409) µg/L group, whereas in the groups taking antiretroviral therapy the concentration was elevated, NNRTI/INTR with 444 (124-580) µg/L and INTR/IP with 412 (185-548) µg/L. In the ART-naive group, the serum concentration of hepcidin had a correlation with iron concentration (r = 0.278, p G variant was 47.7% in HIV-1 virus-infected individuals compared to HIV-1 seronegative individuals, which was 39.0%. The AG genotype was related to increased serum hepcidin concentration, both in seronegative individuals and in HIV-1 virus carriers. According to the results, the serum concentration of hepcidin influences the iron metabolism of HIV-1 individuals and contributes to the development of hematological alterations

Characteristics of the action of oxysterols 7-ketocholesterol and cholestane-3,5,6-triol on ABCs and LRP transport proteins of mesenchymal stem cells derived from bone marrow of patients with acute myeloid leukemia

As proteínas ABCs e LRP (Lung resistance-related protein) fazem o transporte de lipídios e de xenobióticos, e estão associadas ao fenótipo de resistência a múltiplas drogas. As células-tronco mesenquimais (CTM) são células multipotentes presentes na medula óssea e fazem parte do nicho hematopoiético. Alterações fenotípicas em CTMs estão relacionadas a progressão da leucemia mieloide aguda (LMA). Os oxisteróis, 7-KC e Triol, são derivados do colesterol; e apresentam funções na sinalização, proliferação e morte celular. Sendo assim, este estudo avaliou a ação do 7-KC (30 M, 50 M e 70 M) e de Triol (20 M, 30 M e 40 M) nas proteínas ABC-A1, ABC-D4, ABC-G1, ABC-G2 e LRP e, em suas respectivas vias de modulação, LXR-/ e Sonic Hedgehog (SHh), das CTM derivadas de medula óssea de pacientes com LMA (CTM-D), sendo utilizado como controle CTM de indivíduos saudáveis (CTM-S). Primeiro, foi observado que a quantidade de LRP, ABC-A1 e do receptor LXR- diminuíram em CTM-D, enquanto houve aumento de glutationa oxidada e das proteínas relacionadas a resistência a apoptose, a proliferação e invasão celular: a survivina, o receptor SHh, a proteína SMO, Gli3 e do CD147 correceptor da proteína ABC-G2 respectivamente. Em seguida, após o tratamento com 7-KC por 24 horas, foi observado que em CTM-S ocorreu aumento de ABC-A1 na concentração de 30 M, e translocação da proteína SMO para o núcleo; com 70 M de 7-KC foi observado aumento de survivina, de glutationa oxidada e diminuição da proteína LRP. Esses efeitos não foram observados em CTM-D tratadas com 7-KC. Em ambas as linhagens, foi observado, que em nível molecular, o tratamento com 7-KC apresentou efeito no processo de fusão e fissão mitocondrial. O tratamento com Triol por 24 horas, diminuiu a quantidade de ABC-A1 e de seu modulador, o receptor LXR- em CTM-S. Esses resultados foram associados ao aumento da proteína SMO e, de sua translocação para o núcleo na contração de 20 M. Esse efeito não foi observado em CTM-D. Em ambas as linhagens tratadas com Triol ocorreu aumento de glutationa oxidada e diminuição de glutationa reduzida. Além disso, foi observado, em nível molecular, inversão no processo metabólico de fusão e fissão mitocondrial, enquanto em CTM-S houve aumento na expressão de genes relacionados fusão, em CTM-D houve aumento dos genes relacionados a fissão. Em CTM-D tratadas com Triol, foi observado aumento de ABC-D4 no citoplasma; e aumento de LRP apenas na concentração de 20 M de Triol, seguida de diminuição nas concentrações de 30 M e 40 M de Triol. Esses efeitos não foram observados em CTM-S. O tratamento com 40 M de Triol aumentou a quantidade de CD147 em CTM-D e diminuiu em CTM-S. Aqui, foi encontrado a proteína ABC-D4 presente no citoplasma e no núcleo das CTMs. Em suma, as linhagens CTM-S e CTM-D são diferentes fisiologicamente. E, nas mesmas concentrações utilizadas de 7-KC e de Triol em CTMs foi observado efeito diferente, e em partes, oposto sendo que essa diferença é decorrente da origem das células. Além disso, nossos resultados são característicos da ação metabólica citotóxica dos oxisteróis 7-KC e Triol nas CTMsThe ABCs and LRP (Lung resistance-related protein) proteins transport lipids and xenobiotics, and are associated with the multidrug resistance phenotype. Mesenchymal stem cells (MSC) are multipotent cells present in the bone marrow and are part of the hematopoietic niche. Phenotypic changes in MSCs are related to the progression of acute myeloid leukemia (AML). The oxysterols, 7-KC and Triol, are derived from cholesterol; and have functions in cell signaling, proliferation and death. Therefore, this study evaluated the action of 7-KC (30 M, 50 M and 70 M) and Triol (20 M, 30 M and 40 M) on proteins ABC-A1, ABC-D4, ABC-G1, ABC -G2 and LRP and, in their respective modulation pathways, LXR-/ and Sonic Hedgehog (SHh), from MSCs derived from bone marrow of patients with AML (MSC-D), being used as control MSCs in healthy individuals (MSC-S). First, it was observed that the amount of LRP, ABC-A1 and the LXR- receptor were decreased in MSC-D, while there was an increase in oxidized glutathione and proteins related to resistance to apoptosis, proliferation and cell invasion: survivin, the SHh receptor, the SMO protein, Gli3 and CD147 ABC-G2 protein co-receptor respectively. Then, after treatment with 7-KC for 24 hours, it was observed that in MSC-S there was an increase in ABC-A1 at a concentration of 30 M, and translocation of the SMO protein to the nucleus; with 70 M of 7-KC, an increase in survivin, oxidized glutathione and a decrease in LRP protein was observed. These effects were not observed in MSC-D treated with 7-KC. In both strains, it was observed that, at the molecular level, treatment with 7-KC influenced the process of mitochondrial fusion and fission. Treatment with Triol for 24 hours decreased the amount of ABC-A1 and its modulator, the LXR- receptor in MSC-S. These results were associated with an increase in SMO protein and its translocation to the nucleus in a 20 M contraction. This effect was not observed in MSC-D. In both strains treated with Triol there was an increase in oxidized glutathione and a decrease in reduced glutathione. Furthermore, at the molecular level, an inversion in the metabolic process of mitochondrial fusion and fission was observed, while in MSC-S there was an increase in the expression of genes related to fusion, in MSC-D there was increase in genes related to fission. In MSC-D treated with Triol, an increase of ABC-D4 in the cytoplasm was observed; and LRP increase only in the 20 M Triol concentration, followed by a decrease in the 30 M and 40 M Triol concentrations. These effects were not observed in MSC-S. Treatment with 40 M of Triol increased the amount of CD147 in MSC-D and decreased in MSC-S. Here, the ABC-D4 protein was found to be present in the cytoplasm and nucleus of MSCs. In summary, the MSC-S and MSC-D strains are physiologically different. And, in the same used concentrations of 7-KC and Triol in MSCs, a different effect was observed, and in parts, opposite, being that this difference is due to the origin of the cells. Furthermore, our results are characteristic of the cytotoxic metabolic action of the oxysterols 7-KC and Triol on MSC

Paraoxonase Role in Human Neurodegenerative Diseases

The human body has biological redox systems capable of preventing or mitigating the damage caused by increased oxidative stress throughout life. One of them are the paraoxonase (PON) enzymes. The PONs genetic cluster is made up of three members (PON1, PON2, PON3) that share a structural homology, located adjacent to chromosome seven. The most studied enzyme is PON1, which is associated with high density lipoprotein (HDL), having paraoxonase, arylesterase and lactonase activities. Due to these characteristics, the enzyme PON1 has been associated with the development of neurodegenerative diseases. Here we update the knowledge about the association of PON enzymes and their polymorphisms and the development of multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), Alzheimer’s disease (AD) and Parkinson’s disease (PD)

Effects of Oxysterols on Immune Cells and Related Diseases

Oxysterols are the products of cholesterol oxidation. They have a wide range of effects on several cells, organs, and systems in the body. Oxysterols also have an influence on the physiology of the immune system, from immune cell maturation and migration to innate and humoral immune responses. In this regard, oxysterols have been involved in several diseases that have an immune component, from autoimmune and neurodegenerative diseases to inflammatory diseases, atherosclerosis, and cancer. Here, we review data on the participation of oxysterols, mainly 25-hydroxycholesterol and 7α,25-dihydroxycholesterol, in the immune system and related diseases. The effects of these oxysterols and main oxysterol receptors, LXR and EBI2, in cells of the immune system (B cells, T cells, macrophages, dendritic cells, oligodendrocytes, and astrocytes), and in immune-related diseases, such as neurodegenerative diseases, intestinal diseases, cancer, respiratory diseases, and atherosclerosis, are discussed

Ferroptosis Mechanisms Involved in Neurodegenerative Diseases

Ferroptosis is a type of cell death that was described less than a decade ago. It is caused by the excess of free intracellular iron that leads to lipid (hydro) peroxidation. Iron is essential as a redox metal in several physiological functions. The brain is one of the organs known to be affected by iron homeostatic balance disruption. Since the 1960s, increased concentration of iron in the central nervous system has been associated with oxidative stress, oxidation of proteins and lipids, and cell death. Here, we review the main mechanisms involved in the process of ferroptosis such as lipid peroxidation, glutathione peroxidase 4 enzyme activity, and iron metabolism. Moreover, the association of ferroptosis with the pathophysiology of some neurodegenerative diseases, namely Alzheimer’s, Parkinson’s, and Huntington’s diseases, has also been addressed

7-Ketocholesterol Promotes Oxiapoptophagy in Bone Marrow Mesenchymal Stem Cell from Patients with Acute Myeloid Leukemia

7-Ketocholesterol (7-KC) is a cholesterol oxidation product with several biological functions. 7-KC has the capacity to cause cell death depending on the concentration and specific cell type. Mesenchymal stem cells (MSCs) are multipotent cells with the ability to differentiate into various types of cells, such as osteoblasts and adipocytes, among others. MSCs contribute to the development of a suitable niche for hematopoietic stem cells, and are involved in the development of diseases, such as leukemia, to a yet unknown extent. Here, we describe the effect of 7-KC on the death of bone marrow MSCs from patients with acute myeloid leukemia (LMSCs). LMSCs were less susceptible to the death-promoting effect of 7-KC than other cell types. 7-KC exposure triggered the extrinsic pathway of apoptosis with an increase in activated caspase-8 and caspase-3 activity. Mechanisms other than caspase-dependent pathways were involved. 7-KC increased ROS generation by LMSCs, which was related to decreased cell viability. 7-KC also led to disruption of the cytoskeleton of LMSCs, increased the number of cells in S phase, and decreased the number of cells in the G1/S transition. Autophagosome accumulation was also observed. 7-KC downregulated the SHh protein in LMSCs but did not change the expression of SMO. In conclusion, oxiapoptophagy (OXIdative stress + APOPTOsis + autophagy) seems to be activated by 7-KC in LMSCs. More studies are needed to better understand the role of 7-KC in the death of LMSCs and the possible effects on the SHh pathway

Paraoxonases (PON) 1, 2, and 3 Polymorphisms and PON-1 Activities in Patients with Sickle Cell Disease

(1) Background: Oxidative stress, chronic inflammation, vasoocclusion, and free iron are all features present in sickle cell disease. Paraoxonases (PON) are a family (PON-1, PON-2, PON-3) of antioxidant enzymes with anti-inflammatory action. Here, for the first time, we described PON-1 activities and PON-1, PON-2, PON-3 polymorphisms in patients with sickle cell disease, homozygous for HbSS, compared with healthy controls. (2) Methods: The groups were matched for age and gender. PON-1 activities (arylesterase and paraoxonase) were determined by enzymatic hydrolysis of phenylcetate and paraoxon, respectively. Polymorphisms were determined by Restriction Fragment Length Polymorphism- Polymerase Chain Reaction (RFLP-PCR). (3) Results: Plasma cholesterol and fractions, ApoA1 and ApoB levels were all decreased in sickle cell disease patients, while anti-oxidized low-density lipoprotein (LDL) antibodies and C-reactive protein were increased. Serum arylesterase activity was lower in sickle cell disease patients when compared with healthy controls. In patients, paraoxonase activity was higher in those with PON-1 RR Q192R polymorphism. In these patients, the increase of serum iron and ferritin levels and transferrin saturation were less pronounced than those observed in patients with QQ or QR polymorphism. No differences were observed with PON-1 L55M, and PON-2 and PON-3 polymorphisms. Multivariate regression analysis showed that transferrin and ferritin concentrations correlated with arylesterase and paraoxonase activities. (4) Conclusions: Both transferrin and ferritin were the main predictors of decreased arylesterase and paraoxonase activities in patients with sickle cell disease. LDL oxidation increased, and RR PON-1 Q192R polymorphism is likely to be a protective factor against oxidative damage in these patients