Search

Background: Although the safety of mesenchymal stroma/stem cells (MSCs)-based therapies had already extensively demonstrated, non-clinical biodistribution studies are essential for predicting the safety and efficacy of these cells. Herein we assessed the biodistribution of human immature dental pulp stem cells (hIDPSCs), which has investigated as a candidate for the treatment of Huntington’s disease (HD). Method: For this, we intravenously transplanted hIDPSCs transfected with luciferase or labeled with magnetic nanoparticle in C57BL/6 mice and performed the bioluminescence image (BLI) or inductively coupled plasma mass spectrometer (ICP-MS) to quantity in vivo and ex vivo biodistribution after 4h, 24h, 3, 7, and 30 days of the hIDPSCs administration. Results: BLI’s results showed the presence of hIDPSCs in the chest, lungs, and head after 4h, 24 h, and 3 days of the cell transplantation. No bioluminescent signal was observed in the chest or head on days 7 and 30 days. The ICP-MS’s results showed that the hIDPSCs engraft into the liver, kidney, heart, and lungs. However, the number of hIDPSCs in these sites significantly reduced from the seventh day, being undetectable on the 30th day. By contrast, we observed that the hIDPSCs not only engrafted into the brain, but also remain in this organ for 30 days. Conclusion: These data provide evidence that the hIDPSCs successfully engraft and remain in the brain for until 30 days after the cell transplantation, demonstrating that these cells can migrate and homing to the brain, being a useful candidate for the treatment of neurodegenerative disorders, such as HD.

Crotamine is a rattlesnake-derived toxin that causes fast-twitch muscle paralysis. As a cell-penetrating polypeptide, crotamine has been investigated as an experimental anti-cancer and immunotherapeutic agent. We hypothesized that molecules targeting crotamine could be designed to study its function and intervene in its adverse activities. Here, we characterize synthetic crotamine and show that, like the venom-purified toxin, it induces hindlimb muscle paralysis by affecting muscle contraction and inhibits KCNA3 (Kv1.3) channels. Synthetic crotamine, labeled with a fluorophore, displayed cell penetration, subcellular myofiber distribution, ability to induce myonecrosis, and bind to DNA and heparin. Here, we used this functionally validated synthetic polypeptide to screen a combinatorial phage display library for crotamine-binding cyclic peptides. Selection for tryptophan-rich peptides was observed, binding of which to crotamine was confirmed by ELISA and gel shift assays. One of the peptides (CVWSFWGMYC), synthesized chemically, was shown to bind both synthetic and natural crotamine and to block crotamine-DNA binding. In summary, our study establishes a functional synthetic substitute to the venom-derived toxin and identifies peptides that could further be developed as probes to target crotamine. KEY MESSAGES: Synthetic crotamine was characterized as a functional substitute for venom-derived crotamine based on myotoxic effects. A combinatorial peptide library was screened for crotamine-binding peptides. Tryptophan-rich peptides were shown to bind to crotamine and interfere with its DNA binding. Crotamine myofiber distribution and affinity for tryptophan-rich peptides provide insights on its mechanism of action.

Huntington’s disease (HD) is a neurodegenerative disorder caused by the expansion of CAG repeats in the huntingtin gene. The disease causes the progressive degeneration of neurons affecting particularly the medium spiny neurons (MSNs) within the striatum. The mHtt inclusions promote neurodegeneration. However, the mHtt can spread to different brain areas through exosomes. For this reason, it is not surprising that HD causes motor, cognitive and neuropsychiatric dysfunctions. To date there is no treatment able to modify the natural history of the disease. In this sense, the advanced cellular therapy, based on the therapeutic use of mesenchymal stem cells (MSCs) emerges as a potential candidate for HD treatment. This is because, the MSCs produce many critical therapeutic molecules which act in multiple cellular and molecular targets. Moreover, in addition, advanced cell therapy is a unique approach that could provides neuroprotection and neuroregeneration. However, the current discovery that the MSC mechanism of action is mediated by exosomes, have encouraged scientist to explore the therapeutic potential of the cell-free therapy. Based on this, we revisited the HD pathophysiology, areas. Providing evidence that MSC and MSC-derived exosomes can be used to change the natural history of HD.

Crotamine is a lysine and cysteine rich 42 amino acids long bio-active polypeptide, isolated from the venom of a South American rattlesnake, that also can be used as cell penetrating peptide. A facile synthetic scheme for coupling cargo molecules like fluorophores (carboxyfluorescein) or MRI probes (Gd-DO3A-based macrocycle) is presented. The toxicity, cellular internalization and steady-state accumulation after long-term incubation for 18 h, as well as magnetic resonance relaxivities and cellular relaxation rates of crotamine based probes were evaluated and compared to its shorter synthetic fragment CyLoP-1. The longitudinal relaxivity (r1) of the conjugates of CyLoP-1 and crotamine is significantly lower in medium than in water indicating to the lower contrast enhancement efficacy of DO3A-based probes in biological samples. Carboxyfluorescein labeled crotamine did not exhibit toxicity up to a concentration of 2.5 µM. CyLoP-1 accumulated about four times better within the cells compared to crotamine. Fluorescence microscopy suggests different predominant uptake mechanisms for crotamine and CyLoP-1 in 3T3 cells. While crotamine is predominantly localized in vesicular structures (most likely endosomes and lysosomes) within the cell, CyLoP-1 is mainly homogeneously distributed in the cytosol. The cellular relaxation rate (R1, cell) of the crotamine based probe was not significantly increased whereas the corresponding CyLoP-1-derivative showed a slightly elevated R1, cell. This study indicates the potential of crotamine and in particular the shorter fragment CyLoP-1 to be useful for an efficient transmembrane delivery of agents directed to intracellular (cytosolic) targets. However, the applicability of the conjugates synthesized here as contrast agents in MR imaging is limited. Further improvement is needed to prepare more efficient probes for MRI applications, i.e., by replacing the DO3A- with a DOTA-based chelate.

Conditioned medium (CM) (cell secretome) is a cocktail of growth factors, cytokines, and other soluble mediators secreted by cells into a culture medium. These growth factors are fundamental in many cellular processes such as cell growth, differentiation, and others and the composition of these factors is individual for each cell type. Osteoclasts are large multinucleated cells that are responsible for bone resorption. Immune and cancer cells are known to produce different growth factors, which are able to induce or inhibit osteoclast differentiation. Herein, we evaluated the effect of CM obtained from the supernatant of activated and non-activated Jukart-E6 cells, as well as from one murine (B16-F10) and one human melanoma cell line (SK-MEL-28). To induce osteoclast differentiation, murine bone marrow mononuclear cells were cultured in the presence and absence of differentiation factors (DF), such as macrophage colony-stimulating factor, prostaglandin E2, receptor activator of nuclear factor-κB ligand, and CM. We measured the concentration of interleukin 6, tumor necrosis factor-α and interferon γ (IFN-γ) in CM that can inhibit or induce osteoclastogenesis. Our study demonstrated that CM obtained from each cell line suppresses or inhibits osteoclasts formation at early and intermediate stages of differentiation in the absence or presence of DF. CM obtained from activated Jurkat-E6 cells demonstrates a stronger effect when compared with CM from naïve Jurkat-E6 cells or human and murine melanoma cells. Moreover, CM obtained from activated Jurkat-E6 cells shows higher secretion of IFN-γ, which is an inhibitor of osteoclastogenesis, in comparison with CM obtained from the three other cell lines. On the other hand, CM derived from B16-F10 cells showed a smaller inhibitory effect when compared with CM derived from the other cells.

Cancer is one of the most important health problems and the second leading cause of death worldwide. Despite the advances in oncology, cancer heterogeneity remains challenging to therapeutics. This is because the exosome-mediated crosstalk between cancer and non-cancer cells within the tumor microenvironment (TME) contributes to the acquisition of all hallmarks of cancer and leads to the formation of cancer stem cells (CSCs), which exhibit resistance to a range of anticancer drugs. Thus, this review aims to summarize the role of TME-derived exosomes in cancer biology and explore the clinical potential of mesenchymal stem-cell-derived exosomes as a cancer treatment, discussing future prospects of cell-free therapy for cancer treatment and challenges to be overcome.

Facial paralysis can result in severe implications for the patients. However, stem cell biology has become an important field in regenerative medicine since the discovery and characterization of mesenchymal stem cells. Our aim was to evaluate the regeneration after facial nerve crush injury and application of human immature dental pulp stem cells (iDPSC). For this study 70 Wistar rats underwent a unilateral facial nerve crush injury and were divided into two groups: Group I (GI): Crushed; Group II (GII): Crushed and iDPSC, and distributed into study periods of 3, 7, 14, 21, and 42 postoperative days. Facial nerve regeneration was analyzed via functional recovery of whisker movement, histomorphometric analysis, and immunoblotting assay. The results show that GII had complete functional recovery at 14 days, while GI recovered after 42 days. Also, regarding the facial nerve trunk, GII presented histological improvement, evidencing better axonal and structural organization of the myelin sheath, and exhibited statistically higher values for the outer and inner perimeters and g-ratio. Nevertheless, GI exhibited statistically higher values for the thickness of myelin sheath. In the buccal branch, no differences were observed for all parameters between groups. At 42 days, both groups GI and GII were close to the levels observed for the control group. Concerning nerve growth factor expression, GII exhibited statistically greater values ( p < 0.05) compared with the control group at 7 days. In summary, a single injection of human iDPSC promoted a positive effect on regeneration of the facial nerve trunk after 14 days and provided an alternative to support regeneration following peripheral nerve injury.

Crotamine is a rattlesnake-derived toxin that causes fast-twitch muscle paralysis. As a cell-penetrating polypeptide, crotamine has been investigated as an experimental anti-cancer and immunotherapeutic agent. We hypothesized that molecules targeting crotamine could be designed to study its function and intervene in its adverse activities. Here, we characterize synthetic crotamine and show that, like the venom-purified toxin, it induces hindlimb muscle paralysis by affecting muscle contraction and inhibits KCNA3 (Kv1.3) channels. Synthetic crotamine, labeled with a fluorophore, displayed cell penetration, subcellular myofiber distribution, ability to induce myonecrosis, and bind to DNA and heparin. Here, we used this functionally validated synthetic polypeptide to screen a combinatorial phage display library for crotamine-binding cyclic peptides. Selection for tryptophan-rich peptides was observed, binding of which to crotamine was confirmed by ELISA and gel shift assays. One of the peptides (CVWSFWGMYC), synthesized chemically, was shown to bind both synthetic and natural crotamine and to block crotamine-DNA binding. In summary, our study establishes a functional synthetic substitute to the venom-derived toxin and identifies peptides that could further be developed as probes to target crotamine. KEY MESSAGES: Synthetic crotamine was characterized as a functional substitute for venom-derived crotamine based on myotoxic effects. A combinatorial peptide library was screened for crotamine-binding peptides. Tryptophan-rich peptides were shown to bind to crotamine and interfere with its DNA binding. Crotamine myofiber distribution and affinity for tryptophan-rich peptides provide insights on its mechanism of action.

Neurodegenerative disorders, such as Huntington’s disease (HD), Alzheimer’s disease (AD), and Parkinson’s disease (PD), are characterized by changes in the levels and activities of neurotrophic factors (NTFs), such as brain-derived neurotrophic factor (BDNF). Gain-of-function and loss-of-function experiments demonstrate in fact the linkage between wild-type huntingtin (HTT) and gene transcription and intracellular transport of BDNF. In the present chapter, we will analyze the involvement of BDNF in HD and other neurodegenerative diseases. We will discuss the current BDNF technologies focusing on stem cell therapies that induce BDNF upregulation, for instance, the method of autologous mesenchymal stem cell (MSC) culturing in the presence of cocktail of BDNF inducers and factors (MSC/BDNF), genetic engineering of MSC and their use as a vector for BDNF gene delivery, and combined method of establishment of embryonic stem cell (ESC)-derived BDNF-overexpressing neural progenitors, which is still at the preclinical stage. Clinical trial that uses MSC/BDNF is already in course, while genetic engineering of MSC/BDNF is in perspective to treat adult and juvenile HD. The potential application of these technologies is beyond HD. Other neurodegenerative disorders such as Alzheimer’s and Parkinson’s diseases also can be further included in the list of clinical trials that use MSC/BDNF or even ESC/BDNF-overexpressing neural progenitors.

Bothrops moojeni, a Brazilian lanced-head viper, presents a rich, but not well explored, venom composition. This venom is a powerful tool for the discovery of new molecular targets in many different biological processes. Osteoclasts (OC) are extremely important for bone maintenance, calcium physiology, and balance of tissue regeneration being involved in such diseases as osteoporosis and rheumatoid arthritis. The goal of our study was to evaluate the effect of Bothrops moojeni's venom and its fractions of human peripheral blood mononuclear cells derived OCs in vitro differentiation. After the induction of OCs differentiation, on day 4 the venom was added at different concentrations (5, 0.5, and 0.05 µg/mL), and the reduction of tartrate-resistant acid phosphatase positive (TRAP+) osteoclasts, which was more prominent at the concentration of 5 µg / mL were observed. In order to evaluate the effect of major venom’s components (metalloproteases and serinoproteases) on TRAP+ OCs, the EDTA, and aprotinin were used to inhibit the catalytic activity of both proteases. The results suggest that proteases were not crucial for the reduction of TRAP+ OCs. Phalloidin staining was used for morphological analyzes of F-actin rings integrity. Venom provoked F-actin ring disruption in treated versus control OCs. We obtain high molecular weight (HW) and low molecular weight (LW) venom fractions. Both fractions induced the reduction of TRAP+ OCs (HW fraction at a concentration of 5 µg / mL and LW fraction at -1 µg / mL, respectively). We performed a secretome analysis of OCs treated with venom and its fractions using mass spectrometry (LC-MS/IT-Tof). The data obtained demonstrate possible pathways and mechanisms involved in OCs reduction after the treatment. Example giving is catabolic mechanisms for HW and proteins correlated with genetic modifications for LW. New experiments are in progress, aiming to discover the molecules that possibly interfering in the osteoclasts differentiation.

Since the COVID-19 pandemic started, mesenchymal stromal cells (MSC) appeared as a therapeutic option to reduce the over-activated inflammatory response and promote recovery of lung damage. Most clinical studies use intravenous injection for MSC delivery, raising several concerns of thrombogenic risk due to MSC procoagulant activity (PCA) linked to the expression of tissue factor (TF/CD142). This is the first study that demonstrated procoagulant activity of TF+ human immature dental pulp stromal cells (hIDPSC, NestaCell® product) with the percentage of TF+ cells varied from 0.2% to 63.9% in plasma of healthy donors and COVID-19 heparin-treated patients. Thrombogenic risk of TF+ hIDPSCs was evaluated by rotational thromboelastometry (in vitro) and in critically ill COVID-19 patients (clinical trial). We showed that the thromboelastography is not enough to predict the risk of TF+ MSC therapies. Using TF-negative HUVEC cells, we demonstrated that TF is not a unique factor responsible for the cell's procoagulant activity. However, heparin treatment minimizes MSC procoagulant (in vitro). We also showed that the intravenous infusion of hIDPSCs with prophylactic enoxaparin administration in moderate to critically ill COVID-19 patients did not change the values of D-dimer, neither in the PT and PTT times. Our COVID-19 clinical study measured and selected the therapeutic cells with low TF (less than 25% of TF+ hIDPSCs). Our data indicate that the concomitant administration of enoxaparin and low TF-loaded is safe even for critically ill COVID-19 patients.

The aging population has contributed to the rapid rise in the global incidence of neurodegenerative diseases. Despite the medical advances, there are no effective treatments for these disorders. Therefore, there is an urgent need for new treatments for these diseases. In this sense, cell therapy has been recognized as the best candidate for treating incurable diseases, such as neurodegenerative disorders. However, the therapeutic use of these cells can be limited by several factors. Thus, there has been a rediscovery that extracellular vesicles, including exosomes, can be alternatively explored in the treatment of these diseases, overcoming the limits of cell-based therapy. In this sense, this review aims to revisit all areas from biology, including biogenesis and the content of exosomes, to biotechnology, proposing the minimal information required to isolate, characterize, and study the content of these vesicles for scientific and/or clinical purposes.

Crotamine is a basic, 42-residue polypeptide from snake venom that has been shown to possess cell-penetrating properties. Here we describe the preparation, purification, biochemical and biophysical analysis of venom-derived, recombinant, chemically synthesized, and fluorescent-labeled crotamine. We also describe the formation and characterization of crotamine-DNA and crotamine-RNA nanoparticles; and the delivery of these nanoparticles into cells and animals. Crotamine forms nanoparticles with a variety of DNA and RNA molecules, and crotamine-plasmid DNA nanoparticles are selectively delivered into actively proliferating cells in culture or in living organisms such as mice, Plasmodium, and worms. As such, these nanoparticles could form the basis for a nucleic acid drug-delivery system. We also describe here the design and characterization of crotamine-functionalized gold nanoparticles, and the delivery of these nanoparticles into cells. We also evaluated the viability of using the combination of crotamine with silica nanoparticles in animal models, aiming to provide slow delivery, and to decrease the crotamine doses needed for the biological effects. In addition, the efficacy of administering crotamine orally was also demonstrated.

Crotamine is a basic, 42-residue polypeptide from snake venom that has been shown to possess cell-penetrating properties. Here we describe the preparation, purification, biochemical and biophysical analysis of venom-derived, recombinant, chemically synthesized, and fluorescent-labeled crotamine. We also describe the formation and characterization of crotamine-DNA and crotamine-RNA nanoparticles; and the delivery of these nanoparticles into cells and animals. Crotamine forms nanoparticles with a variety of DNA and RNA molecules, and crotamine-plasmid DNA nanoparticles are selectively delivered into actively proliferating cells in culture or in living organisms such as mice, Plasmodium, and worms. As such, these nanoparticles could form the basis for a nucleic acid drug-delivery system. We also describe here the design and characterization of crotamine-functionalized gold nanoparticles, and the delivery of these nanoparticles into cells. We also evaluated the viability of using the combination of crotamine with silica nanoparticles in animal models, aiming to provide slow delivery, and to decrease the crotamine doses needed for the biological effects. In addition, the efficacy of administering crotamine orally was also demonstrated.

Background Although thromboembolism is a rarely reported event in mesenchymal stem cell (MSC) intravascular administration-based therapies, studies have been suggested that the tissue factor (TF/CD142) expression increases the risk for clot formation in a dose-dependent manner. For this reason, this study aimed to evaluate whether the expression levels of TF can be used as a predictive value to determine the risk for thromboembolism. Methods The thrombogenic potential of human immature dental pulp stem cells (hIDPSCs), in two proportional doses to clinically tested doses (1 × 106/Kg of body weight and 20 × 106/patient), and with different TF-positive cell percentages (0.2 to 60% of TF-positive cells), was evaluated using the rotational thromboelastometry (ROTEM). The percentual of TF-positive cells was correlated with the clotting time (CT). Results Our results showed that all cells, independently of the TF-positive cell percentages, reduced the CT when compared with the controls (plasma with saline), being not verified statistically significant differences among the CTs of cells expressing among 0.2 to 60% of TF. These results were also confirmed by the Pearson's correlation (R=0.06), which did not indicate a statistical correlation between the TF expression and CT. Additionally, we verified that HUVEC cells (TF-negative), in these even doses, had a CT statistically identical to the hIDPSCs. However, the CT of both HUVEC and hIDPSCs increased following the addition of enoxaparin. Conclusion Our results provide evidence that despite the TF is recognized as a key activator of extrinsic via of coagulation; the TF expression does not necessarily increase the risk for thromboembolism. Considering that previous studies suggesting that the TF expression can elicit the clot formation were based on ROTEM, and that they did not include a TF-negative cell as an additional control, we hypothesized that the CT reduction described can be elicited by the cell lysis, resulted from the rotational movement of the piston into the cup, leading to the exposure of phosphatidylserine phospholipids, which are the main activator of intrinsic via of coagulation cascade. Thus, the ROTEM's results do not reflect a predictive value for clinical studies. Despite this data, the prophylactic administration of anticoagulants such as enoxaparin reduces expressively clot formation.

Acquired aplastic anemia (AA) is a type of bone marrow failure (BMF) syndrome characterized by partial or total bone marrow (BM) destruction resulting in peripheral blood (PB) pancytopenia, which is the reduction in the number of red blood cells (RBC) and white blood cells (WBC), as well as platelets (PLT). The first-line treatment option of AA is given by hematopoietic stem cell (HSCs) transplant and/or immunosuppressive (IS) drug administration. Some patients did not respond to the treatment and remain pancytopenic following IS drugs. The studies are in progress to test the efficacy of adoptive cellular therapies as mesenchymal stem cells (MSCs), which confer low immunogenicity and are reliable allogeneic transplants in refractory severe aplastic anemia (SAA) cases. Moreover, bone marrow stromal cells (BMSC) constitute an essential component of the hematopoietic niche, responsible for stimulating and enhancing the proliferation of HSCs by secreting regulatory molecules and cytokines, providing stimulus to natural BM microenvironment for hematopoiesis. This review summarizes scientific evidences of the hematopoiesis improvements after MSC transplant, observed in acquired AA/BMF animal models as well as in patients with acquired AA. Additionally, we discuss the direct and indirect contribution of MSCs to the pathogenesis of acquired AA.

Melanoma is characterized by high heterogeneity and plasticity, most likely due to the presence of mutated melanocyte stem cells or immature progenitor cells in the skin that serves as precursors to melanoma. In the present study, for the first time, we identified rare cells in the murine melanoma B16F10, and human A2058 and SK-MEL-28 cell lines that express pluripotency markers, including Oct4, Nanog, Sox2 and a marker of melanoma cancer cells (ALDH1/2). These cells are very small with round morphology and they grow onto melanoma cells, thereby demonstrating feeder layer dependence similar to that of other pluripotent cells. These cells underwent self-renewal, symmetric and asymmetric division. We called these cells murine very small cancer stem cells (VSCSC). VSCSC were also found in B16F10-derived clones after 3–5 consecutive passages, where they occur as single cells or as small colonies, nevertheless, always using melanoma cells as feeders. These cells formed melanospheres enriched with Oct4-and ALDH1/2-positive cells. We also evaluated the possible effect of VSCSC that presented in the parental cell line (B16F10) and in clones based on their functional characteristics. We found that VCSCS present in the B16F10 cell line reappearing in their clones were required for continuous tumor growth and were responsible for melanoma cell heterogeneity and plasticity rather than directly affecting functional characteristics of melanoma cells. Our data, together with those of previous reports suggested the existence of melanoma-competent melanocyte stem cells, which corroborate the hypothesis of the existence of tumor-initiating cells and cancer stem cell hierarchies, at least in melanoma.

Acquired aplastic anemia (AA) is characterized by partial or total bone marrow (BM) destruction resulting in pancytopenia. Most of the acquired AA is the result of autoimmune condition the imbalance between T-regulatory cells (Treg), abnormal cytokines production and cytotoxic T cells activation, leading to the hematopoietic stem cells (HSCs) death. The first-line treatment is given by HSC transplant, but some patients did not respond to the treatment. Therefore, new technologies need to treat AA nonresponder patients. Studies are in progress to test the efficacy of stem cell-based therapeutic as mesenchymal stem cells (MSCs), which confer low immunogenicity and are reliable allogeneic transplants in refractory severe AA cases. Furthermore, MSCs comprise the BM stromal niche and have an important role in supporting hematopoiesis by secreting regulatory cytokines, providing stimulus to natural BM microenvironment. In addition, MSCs have immunomodulatory property and are candidates for efficient supporting AA therapy.

Background Huntington′s disease (HD) is a neurodegenerative condition caused by an increase in the CAG repeats of the huntingtin (HTT) gene at chromosome 4. It is a rare disease; whose prevalence varies from 0.4 to 5.7 cases per 100,000 habitants. The clinical manifestations usually start by the fourth decade of age with motor, cognitive and behavioral features, that typically evolve for 10 to 20 years until death. Currently, there is no curative treatment. Nestacell HD is an allogeneic cell therapy manufactured from the human immature dental pulp stem cells by the Brazilian company Cellavita. This study aimed at assessing the safety, tolerability, and preliminary data on the efficacy of intravenous injections of Nestacell HD. It was approved by the local IRB, CONEP, and ANVISA. Methods This is a first-in-human, uncontrolled, open-label, Phase I clinical trial with six HD participants. The study started following the patients for 4 months to collect baseline data. Then, they received 3 administrations with a 1-month interval of either 1 million cells per Kg (first 3 patients) or 2 million cells per kg (last 3 patients) by the intravenous route. The patients were then followed for 2 years. The safety was evaluated based on the adverse events (AV) incidence, severity, and causality. The efficacy was evaluated using the Unified Huntington‘s Disease Rating Scale – UHDRS. The immunological response was evaluated by CD4+ and CD8+ proliferation, and inflammatory markers (cytokines). Results All participants completed the treatment and the two-year follow-up period. The treatment was well tolerated. There were no treatment-related serious adverse events (SAE), nor any AE led to dropout. No patient increased the cytokines or the CD4/CD8. Five patients had improvement to the UHDRS motor domain, which started two weeks after the first administration and remained for six to nine months after the last administration. All patients were entitled to additional infusions to maintain clinical improvement. Conclusion The treatment with Nestacell HD was well tolerated and might have improved the HD motor manifestations; thus justifying further Phase 2 clinical trials.

Toxins have been shown to have many biological functions and to constitute a rich source of drugs and biotechnological tools. We focus on toxins that not only have a specific activity, but also contain residues responsible for transmembrane penetration, which can be considered bioportides-a class of cell-penetrating peptides that are also intrinsically bioactive. Bioportides are potential tools in pharmacology and biotechnology as they help deliver substances and nanoparticles to intracellular targets. Bioportides characterized so far are peptides derived from human proteins, such as cytochrome c (CYCS), calcitonin receptor (camptide), and endothelial nitric oxide synthase (nosangiotide). However, toxins are usually disregarded as potential bioportides. In this review, we discuss the inclusion of some toxins and molecules derived thereof as a new class of bioportides based on structure activity relationship, minimization, and biological activity studies. The comparative analysis of the amino acid residue composition of toxin-derived bioportides and their short molecular variants is an innovative analytical strategy which allows us to understand natural toxin multifunctionality in vivo and plan novel pharmacological and biotechnological products. Furthermore, we discuss how many bioportide toxins have a rigid structure with amphiphilic properties important for both cell penetration and bioactivity.

Cancer cells transformation into a normal state or into a cancer cell population which is less tumorigenic than the initial one is a challenge that has been discussed during last decades and it is still far to be solved. Due to the highly heterogeneous nature of cancer cells, such transformation involves many genetic and epigenetic factors which are specific for each type of tumor. Different methods of cancer cells reprogramming have been established and can represent a possibility to obtain less tumorigenic or even normal cells. These methods are quite complex, thus a simple and efficient method of reprogramming is still required. As soon as induced pluripotent stem cells (iPSC) technology, which allowed to reprogram terminally differentiated cells into embryonic stem cells (ESC)-like, was developed, the method strongly attracted the attention of researches, opening new perspectives for stem cell (SC) personalized therapies and offering a powerful in vitro model for drug screening. This technology is also used to reprogram cancer cells, thus providing a modern platform to study cancer-related genes and the interaction between these genes and the cell environment before and after reprogramming, in order to elucidate the mechanisms of cancer initiation and progression. The present review summarizes recent advances on cancer cells reprogramming using iPSC technology and shows the progress achieved in such field.

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Bovine papillomaviruses (BPVs) are recognized as causal agents of benign and malignant tumors in cattle. Thirteen types of BPVs have already been described and classified into 3 distinct genera. Divergences in the nucleotide sequence of the L1 gene are used to identify new viral types through the employment of PCR assays with degenerated primers. in the present study, a method for identifying BPVs based on PCR-RFLP and DNA sequencing allowed the identification of a new putative Deltapapillomavirus, designated JN/3SP (JQ280500.1). the analysis of the L1 gene showed that this strain was most closely related to the BPVs -1, -2, -13, and OaPV1 (71-73% genetic similarity). in this study, we describe the detection of this new putative Deltapapillomavirus type and verify its phylogenetic position within the genus. Inst Butantan, Lab Genet, São Paulo, Brazil Universidade Federal de São Paulo, Programa Posgrad Biol Estrutural & Func, São Paulo, Brazil Univ São Paulo, Programa Posgrad Interunidades Biotecnol, São Paulo, Brazil Univ Fed Pernambuco, Dept Bot, Recife, PE, Brazil Univ Fed Integracao Latino Amer, Dept Biol, Foz Do Iguacu, PR, Brazil Universidade Federal de São Paulo, Programa Posgrad Biol Estrutural & Func, São Paulo, Brazil CNPq: 402539/2011-7 CNPq: 554816/2006-7 Web of Science

Stem cells are capable of generating various cell lines and can be obtained from adult or embryonic tissues for clinical therapies. Stem cells from deciduous dental pulp are among those that are easily obtainable from adult tissues and have been widely studied because of their ability to differentiate into a variety of cell lines in the presence of various chemical mediators. We have analyze the expression of several proteins related to the differentiation and proliferative potential of cell populations that compose the tooth germ of human fetuses. We evaluate 20 human fetuses of both genders. After being paraffin-embedded, cap and bell stages of tooth germ development were subjected to immunohistochemistry for the following markers: Oct-4, Nanog, Stat-3 and Sox-2. The studied antibodies showed nuclear or cytoplasmic immunnostaining within various anatomical structures and with various degrees of expression, indicating the action of these proteins during tooth development. We conclude that the interrelationship between these transcription factors is complex and associated with self-renewal and cell differentiation. Our results suggest that the expression of Oct-4, Nanog, Sox-2 and Stat-3 are related to differentiation in ameloblasts and odontoblasts.

Stem cells are undifferentiated cells and can self-renew and differentiate into various cell types, besides having immuno-modulating properties and paracrine effects in response to tissue injury, and may therefore treat injuries and diseases or even replace damaged or lost cells. Adipose tissue is an attractive source of adult stem cells, since the human body has a large reserve that is obtained in large amounts by minimally invasive methods. Interest in these cells has been increasing steadily due to their properties and possible applications in regenerative medicine and cell therapy. A large part of these investigations are focused on cardiovascular diseases, which are a leading cause of morbidity and mortality worldwide. Although in recent years treatments have advanced in cardiology, the development of new therapies to recover the damaged tissue still remains one of the main goals of cardiac research. However, to achieve effective results, in vivo and in vitro animal models for preclinical studies and consequently for application in humans must be standardized. The development of preclinical models in large animals requires the use of well-characterized animal cell lines, similar to human cells, and the use of the porcine model represents a great advantage for preclinical translational research.

As células-tronco são células indiferenciadas, capazes de se autorrenovar e de se diferenciarem em diversos tipos celulares, além de apresentarem propriedades imunomoduladoras e efeitos parácrinos mediante injúria tecidual, podendo, dessa forma, tratar lesões e doenças ou ainda substituir células danificadas ou perdidas. Dentre as fontes de células-tronco adultas, o tecido adiposo é uma fonte atrativa, pois o organismo humano possui grande reserva desse tecido, que, por sua vez, é obtido em grandes quantidades por meio de métodos pouco invasivos. O interesse nessas células vem aumentando constantemente devido a suas propriedades e possíveis aplicações na medicina regenerativa e terapia celular. Grande parte dessas pesquisas está voltada para doenças cardiovasculares, que são a principal causa de morbidade e mortalidade em todo o mundo. Embora nos últimos anos, os tratamentos em cardiologia tenham avançado, o desenvolvimento de novas terapias que recuperem o tecido danificado ainda permanece como um dos objetivos principais das pesquisas cardíacas. Porém, para obter resultados eficazes, é necessária a padronização de modelos animais in vivo e in vitro para estudos pré-clínicos e, consequentemente, a aplicação em humanos. O desenvolvimento de modelos pré-clínicos em animais de grande porte exige o uso bem caracterizado de linhagens de células animais semelhantes aos seus equivalentes humanos. O modelo suíno representa uma grande vantagem para a investigação translacional pré-clínica. Stem cells are undifferentiated cells and can self-renew and differentiate into various cell types, besides having immunomodulating properties and paracrine effects in response to tissue injury, and may therefore treat injuries and diseases or even replace damaged or lost cells. Adipose tissue is an attractive source of adult stem cells, since the human body has a large reserve that is obtained in large amounts by minimally invasive methods. Interest in these cells has been increasing steadily due to their properties and possible applications in regenerative medicine and cell therapy. A large part of these investigations are focused on cardiovascular diseases, which are a leading cause of morbidity and mortality worldwide. Although in recent years treatments have advanced in cardiology, the development of new therapies to recover the damaged tissue still remains one of the main goals of cardiac research. However, to achieve effective results, in vivo and in vitro animal models for preclinical studies and consequently for application in humans must be standardized. The development of preclinical models in large animals requires the use of well-characterized animal cell lines, similar to human cells, and the use of the porcine model represents a great advantage for preclinical translational research.