Carlos Magno da Costa Maranduba

Intestinal Microbiota as Modulators of the Immune System and Neuroimmune System: Impact on the Host Health and Homeostasis

Many immune-based intestinal disorders, such as ulcerative colitis and Crohns disease, as well as other illnesses, may have the intestines as an initial cause or aggravator in the development of diseases, even apparently not correlating directly to the intestine. Diabetes, obesity, multiple sclerosis, depression, and anxiety are examples of other illnesses discussed in the literature. In parallel, importance of the gut microbiota in intestinal homeostasis and immunologic conflict between tolerance towards commensal microorganisms and combat of pathogens is well known. Recent researches show that the immune system, when altered by the gut microbiota, influences the state in which these diseases are presented in the patient directly and indirectly. At the present moment, a considerable number of investigations about this subject have been performed and published. However, due to difficulties on correlating information, several speculations and hypotheses are generated. Thus, the present review aims at bringing together how these interactions work—gut microbiota, immune system, and their influence in the neuroimmune system.

Toward Personalized Cell Therapies by Using Stem Cells: Seven Relevant Topics for Safety and Success in Stem Cell Therapy

Stem cells, both embryonic and adult, due to the potential for application in tissue regeneration have been the target of interest to the world scientific community. In fact, stem cells can be considered revolutionary in the field of medicine, especially in the treatment of a wide range of human diseases. However, caution is needed in the clinical application of such cells and this is an issue that demands more studies. This paper will discuss some controversial issues of importance for achieving cell therapy safety and success. Particularly, the following aspects of stem cell biology will be presented: methods for stem cells culture, teratogenic or tumorigenic potential, cellular dose, proliferation, senescence, karyotyping, and immunosuppressive activity.

Mesenchymal stem cells derived from human exfoliated deciduous teeth (SHEDs) induce immune modulatory profile in monocyte-derived dendritic cells.

Background Mesenchymal stem cells have prominent immune modulatory properties, which may have clinical applications; however their major source, bone marrow, is of limited availability. On the other hand, mesenchymal stem cells derived from human exfoliated deciduous teeth (SHEDs) are readily accessible, but their immune regulatory properties have not been completely investigated. This study was designed, therefore, to evaluate the SHEDs influence on DCs differentiation, maturation, ability to activate T cells and to expand CD4+Foxp3+ T cells. Methodology/Principal Findings The experiments were based in cellular co-culture during differentiation and maturation of monocyte derived-DCs (moDCs), with, or not, presence of SHEDs. After co-culture with SHEDs, (moDCs) presented lower expression of BDCA-1 and CD11c, in comparison to DC cultivated without SHEDs. CD40, CD80, CD83 and CD86 levels were also decreased in mature DCs (mDCs) after co-cultivation with SHEDs. To assess the ability of SHEDs-exposed moDCs to modulate T cell responses, the former were separated from SHEDs, and co-cultured with peripheral blood lymphocytes. After 5 days, the proliferation of CD4+ and CD8+ T cells was evaluated and found to be lower than that induced by moDCs cultivated without SHEDs. In addition, an increase in the proportion of CD4+Foxp3+IL-10+ T cells was observed among cells stimulated by mature moDCs that were previously cultivated with SHEDs. Soluble factors released during co-cultures also showed a reduction in the pro-inflammatory cytokines (IL-2, TNF-α and IFN-γ), and an increase in the anti-inflammatory molecule IL-10. Conclusion/Significance This study shows that SHEDs induce an immune regulatory phenotype in moDCs cells, evidenced by changes in maturation and differentiation rates, inhibition of lymphocyte stimulation and ability to expand CD4+Foxp3+ T cells. Further characterization and validation of this phenomenon could support the use of SHEDs, directly or indirectly for immune modulation in the clinical practice.

Expression of Extracellular Matrix Proteins in Human Dental Pulp Stem Cells Depends on the Donor Tooth Conditions

INTRODUCTION Stem cells are characterized by the ability to renew themselves through mitotic cell division and differentiating into a diverse range of specialized cell types. An important source of adult stem cells is the dental pulp. In dentistry, regenerative strategies are of importance because of hard dental tissue damage especially as result of caries lesions, trauma, or iatrogenic procedures. The regeneration of dental tissues relies on the ability of stem cells to produce extracellular (ECM) proteins encountered in the dental pulp tissue. Thus, the aim of this study was to analyze the expression and distribution of proteins encountered in dental pulp ECM (type I collagen, fibronectin, and tenascin) in stem cells. METHODS Human immature dental pulp stem cells (hIDPSCs) from deciduous (DL-1 and DL-4 cell lines) and permanent (DL-2) teeth were used. The distribution of ECM proteins was observed using the immunofluorescence technique. The gene expression profile was evaluated using reverse transcription polymerase chain reaction (RT-PCR) analysis. RESULTS Positive reactions for all ECM proteins were observed independently of the hIDPSCs analyzed. Type I collagen appeared less evident in DL-2 than in other hIDPSCs. Fibronectin and tenascin were less clear in DL-4. The RT-PCR reactions showed that type I collagen was lesser expressed in the DL-2 cells, whereas fibronectin and tenascin were similarly expressed in all hIDPSCs. CONCLUSIONS The distribution and expression of ECM proteins differ among the hIDPSCs. These differences seemed to be related to the donor tooth conditions (deciduous or permanent, retained or erupted, and degree of root reabsorption).

Low-intensity laser phototherapy enhances the proliferation of dental pulp stem cells under nutritional deficiency

Dental trauma in immature permanent teeth can damage pulp vascularization, which leads to necrosis and cessation of apexogenesis. Studies on tissue engineering using stem cells from human exfoliated deciduous teeth (SHEDs) have yielded promising results. Laser phototherapy (LPT) is able to influence the proliferation and differentiation of these cells, which could improve tissue engineering. SHEDs (eighth passage) were seeded into 96-well culture plates (103 cells/well) and were grown in culture medium supplemented with 15% defined fetal bovine serum (FBS) for 12 h. After determining the appropriate nutrition deficiency status (5% FBS), the cells were assigned into four groups: 1) G1 - 15% FBS (positive control); 2) G2 - 5% FBS (negative control); 3) G3 - 5% FBS+LPT 3 J/cm2; and 4) G4 - 5% FBS+LPT 5 J/cm2. For the LPT groups, two laser irradiations at 6 h intervals were performed using a continuous wave InGaAlP diode laser (660 nm, with a spot size of 0.028 cm2, 10 mW) in punctual and contact mode. Cell viability was assessed via an MTT reduction assay immediately after the second laser irradiation (0 h) and 24, 48, and 72 h later. We found that G3 and G4 presented a significantly higher cell growth rate when compared with G2 (p < 0.01). Moreover, G4 exhibited a similar cell growth rate as G1 throughout the entire experiment (p > 0.05). These findings indicate that LPT with 5 J/cm2 can enhance the growth of SHEDs during situations of nutritional deficiency. Therefore, LPT could be a valuable adjunct treatment in tissue engineering when using stem cells derived from the dental pulp of primary teeth.

Satellite Cells: Regenerative Mechanisms and Applicability in Muscular Dystrophy

The satellite cells are long regarded as heterogeneous cell population, which is intimately linked to the processes of muscular recovery. The heterogeneous cell population may be classified by specific markers. In spite of the significant amount of variation amongst the satellite cell populations, it seems that their activity is tightly bound to the paired box 7 transcription factor expression, which is, therefore, used as a canonical marker for these cells. Muscular dystrophic diseases, such as Duchenne muscular dystrophy, elicit severe tissue injuries leading those patients to display a very specific pattern of muscular recovery abnormalities. There have been works on the application of precursors cells as a therapeutic alternative for Duchenne muscular dystrophy and initial attempts have proven the cells inefficient; however later endeavours have proposed solutions for the experiments improving significantly the results. The presence of a range of satellite cells populations indicates the existence of specific cells with enhanced capability of muscular recovery in afflicted muscles.

Dental pulp stem cells express proteins involved in the local invasiveness of odontogenic myxoma

Little is known about the histogenesis of the odontogenic myxoma (OM). Dental pulp stem cells could be candidate precursors of OM because both OM and the dental pulp share the same embryological origin: the dental papilla. For the purpose of comparing OM and stem cells, this study analyzed the expression of two proteins related to OM invasiveness (MMP-2 and hyaluronic acid) in human immature dental pulp stem cells (hIDPSCs). Three lineages of hIDPSCs from deciduous and permanent teeth were used in this study. Immunofluorescence revealed positive reactions for MMP-2 and hyaluronic acid (HA) in all hIDPSCs. MMP-2 appeared as dots throughout the cytoplasm, whereas HA appeared either as diffuse and irregular dots or as short fibrils throughout the cytoplasm and outside the cell bodies. The gene expression profile of each cell lineage was evaluated using RT-PCR analysis, and HA was expressed more intensively than MMP-2. HA expression was similar among the three hIDPSCs lineages, whereas MMP-2 expression was higher in DL-1 than in the other cell lines. The expression of proteins related to OM invasiveness in hIDPSCs could indicate that OM originates from dental pulp stem cells.

Review articleMyeloproliferative neoplasms and the JAK/STAT signaling pathway: an overview

Myeloproliferative neoplasms are caused by a clonal proliferation of a hematopoietic progenitor. First described in 1951 as ‘Myeloproliferative Diseases’ and reevaluated by the World Health Organization classification system in 2011, myeloproliferative neoplasms include polycythemia vera, essential thrombocythemia and primary myelofibrosis in a subgroup called breakpoint cluster region-Abelson fusion oncogene-negative neoplasms. According to World Health Organization regarding diagnosis criteria for myeloproliferative neoplasms, the presence of the JAK2 V617F mutation is considered the most important criterion in the diagnosis of breakpoint cluster region-Abelson fusion oncogene-negative neoplasms and is thus used as a clonal marker. The V617F mutation in the Janus kinase 2 (JAK2) gene produces an altered protein that constitutively activates the Janus kinase/signal transducers and activators of transcription pathway and other pathways downstream as a result of signal transducers and activators of transcription which are subsequently phosphorylated. This affects the expression of genes involved in the regulation of apoptosis and regulatory proteins and modifies the proliferation rate of hematopoietic stem cells.

Myeloproliferative neoplasms and the JAK/STAT signaling pathway: an overview

Myeloproliferative neoplasms are caused by a clonal proliferation of a hematopoietic progenitor. First described in 1951 as ‘Myeloproliferative Diseases’ and reevaluated by the World Health Organization classification system in 2011, myeloproliferative neoplasms include polycythemia vera, essential thrombocythemia and primary myelofibrosis in a subgroup called breakpoint cluster region-Abelson fusion oncogene-negative neoplasms. According to World Health Organization regarding diagnosis criteria for myeloproliferative neoplasms, the presence of the JAK2 V617F mutation is considered the most important criterion in the diagnosis of breakpoint cluster region-Abelson fusion oncogene-negative neoplasms and is thus used as a clonal marker. The V617F mutation in the Janus kinase 2 (JAK2) gene produces an altered protein that constitutively activates the Janus kinase/signal transducers and activators of transcription pathway and other pathways downstream as a result of signal transducers and activators of transcription which are subsequently phosphorylated. This affects the expression of genes involved in the regulation of apoptosis and regulatory proteins and modifies the proliferation rate of hematopoietic stem cells.

The JAK2 gene as a protagonist in chronic myeloproliferative neoplasms

The identification of the association of a JAK2 gene mutation with chronic myeloproliferative neoplasms (cMPN) negative for BCR-ABL(1,2) has allowed significant advances in the understanding of this group of hematologic diseases. The JAK2 gene, located on chromosome 9p24, encodes the JAK2 protein which is a cytoplasmic tyrosine kinase that plays an important role in the signal transduction of various hematopoietic growth factors. The JAK2V617F mutation results in the substitution of the amino acid valine for phenylalanine in the pseudokinase domain (JH2) causing constitutive activation of the kinase domain (JH1) and hypersensitivity to protein growth factors. Among the BCR-ABL negative cMPN cases, the JAK2617F mutation occurs at a frequency of 96% in polycythemia vera (PV), and 50% of essential thrombocythemia (ET) and chronic idiopathic myelofibrosis (MF) patients(2). The association of this mutation with BCR-ABL negative cMPN has contributed to improve the diagnosis, classification and treatment of patients, in particular in respect to PV. Tefferi and Pardanani(3) suggested that an investigation of the JAK2V617F mutation in the peripheral blood should be integrated into the initial assessment of patients with suspected diagnosis of PV and of those of thrombocytosis of unknown cause, with thrombotic complications, including cerebral or abdominal thrombosis, and other clinical manifestations of myeloproliferative diseases. Other less frequent mutations were also found in the JAK2 gene in JAK2V617F-negative PV patients, as well as in other myeloproliferative neoplasms. Several studies report deletions, point mutations and duplications(4) mainly affecting the seven highly conserved amino acid residues (F537-F547) in the JAK2 protein. PV patients positive for these mutations are often heterozygous for the mutation and are characterized by the predominance of myelopoesis, serum erythropoietin levels below normal and lower age at diagnosis(3-5). The clinical evolution of these patients is similar to JAK2V617F-positive PV patients(5). The V617F mutation of the JAK2 gene triggers three clinical manifestations and there is evidence that genetic and additional epigenetic events contribute to the pathogenesis(6). In addition, other genes, such as the MPL, TET2 and ASXL1 genes, may also be mutated with the accumulation of mutations possibly explaining the different phenotypes observed in MPN. The MPL gene, located on chromosome 1p34, encodes the thrombopoietin receptor (cMPL). Its expression is important for growth and survival of megakaryocytes. Some mutations in this gene result in gain of function and have been associated with thrombocytosis, splenomegaly, myelofibrosis and an increased risk of thrombosis(7). Mutations in the transmembrane domain of cMPL were observed in nine patients negative for the JAK2V617F mutation (MPLW515L and MPLW515K); mutations were also detected in JAK2V617F-positive patients. The TET2 (4q24) gene has many mutations (frameshift, missense, nonsense) that are observed in JAK2V617F cMPN-positive (17%) and JAK2V617F-negative patients (7%), with mutational frequencies of approximately 16% in PV, 5% in ET, 17% in MF, 14% in post-PV MF, 14% in post-TE MF and 17% in blast-phase MPN(8). The main function of the TET2 protein is the conversion of 5-methyl-cytosine to 5-hydroxymethyl cytosine; it eventually affects the epigenetic regulation of transcription. The ASXL1 gene maps to chromosome 20q11.1 and belongs to the enhancer of trithorax and polycomb gene family. The function of this gene is believed to include dual activator/suppressor activity toward transcription and includes repression of retinoic acid receptor-mediated transcription. Mutations in this gene are associated with myelodysplastic syndromes (MDS) and chronic myelomonocytic leukemia (CML)(9). In a recent study of 300 patients with a spectrum of non-MPN myeloid malignancies, ASXL1 gene mutations were found in 62 patients (~21%): ~7% in MDS without excess blasts, 11-17% in MDS with ring sideroblasts, 31% in MDS with excess blasts, 23% in post-MDS acute myeloid leukemia (AML), 33% in CML and 30% in primary AML. It was observed that mutations of the ASXL1 gene occur in MPNs in both chronic and blast phases. In a study of 64 patients with ET (n = 35), MF (n = 11), PV (n = 10), blast-phase MPN (n = 5) and unclassifiable MPNs (n = 3), mutations of this gene in heterozygosis were identified in five JAK2V617F-negative patients (~ 8%; 3 MF, 1 ET and 1 blast-phase ET)(10). There are other reports in the literature associating gene mutations to BCR-ABL-negative cMPN. The knowledge about genotype-phenotype interaction could elucidate the molecular mechanisms and contribute to improvements in diagnosis, in the status and in the treatment of patients with ET, MF and PV.