They also showed an increase in the leukocytes diapedesis duration and that this effect disappears with annexin 1 antagonists. Although we did not perform experiments by using antagonists and did not measure the annexin 1 production, we suggest that DEXA could mediate the leukocytes migration activated directly by B. jararacussu components by endogenous mediators, decreasing systemic responses produced by cell damage, keeping the cells on the blood stream without stopping their recruitment

from the bone marrow. In other way we propose that the EP extract seems to reduce the local leukocytes presence in the injection site probably by diminishing the venom initial activities or its cytotoxic effect, therefore reducing find more the bone marrow cell recruitment. It is known that Bothrops venoms are able to activate leukocyte

oxidative stress and this property is due to the presence of MPO enzyme in these cells ( Zamuner selleck chemicals et al., 2001; Elifio-Esposito et al., 2011). The local activity measurement of this enzyme is an indirect indicator of activated neutrophil presence that in excess can cause tissue damage ( Klebanoff, 2005; Davies, 2011). Our data have shown that mouse EDL muscles exposed to the perimuscular injection with B. jararacussu had an increase on the MPO activity. Treatment with DEXA and EP extract reduced the MPO activity in these muscles, corroborating the reduction found in the inflammatory cells count in the same muscle.

DEXA has a known anti-inflammatory property ascribed to the ability of inhibiting the expression of pro-inflammatory factors, consequently reducing the inflammatory reaction and the cell damage ( Euzger et al., 1999; Clark, 2007). Even though some investigators argue about the importance of inflammatory cells in the acute tissue damage caused by venoms (Teixeira et al., 2003 and Teixeira et al., 2005) our results strongly suggest that the local infiltration of these cells is significantly involved in the muscle tissue injury and is correlated with data previously described (Farsky et al., 1997; Barros et al., 1998; acetylcholine Araujo et al., 2007; Carneiro et al., 2002 and Carneiro et al., 2008). In conclusion, inflammation seems to play an important role in the local muscle damage induced by these Bothrops venoms once the use of the anti-inflammatory drug DEXA protected the muscle tissue from the venom myonecrotic effect. Our data also confirm and reproduced the antiophydic effects of EP crude extract and added that EP has an anti-inflammatory effect itself. Finally we have to look forward to investigate the role of inflammation on the Bothrops snakebites and find anti-inflammatory drugs that can help the antivenom in venom neutralization and prevent the late disabilities.

10). Wasps reduced the duration of ventilation movements at higher temperatures (Fig. 9). Total duration of respiration movement events was up to tenfold longer than in honeybees (42.2 vs. 4.8 s at 20 °C, 27.8 vs. 2.3 s at 25 °C; mean values, honeybee data from Kovac et al., 2007). It seems that resting yellow jackets gain their efficient gas exchange to a considerable extent via the length of respiration movements per respiratory cycle. Therefore, they manage a considerably higher RMR (see Käfer et al., 2012) with a similar respiration frequency as honeybees (see Fig. 4). The high respiration volume and efficiency might be responsible for the rather high transition temperature

from discontinuous to cyclic respiration. Despite an overall high level JAK assay Entinostat mw and a steep increase of resting metabolism with increasing ambient temperature (high Q10), resting yellow jackets maintain DGC at comparably high ambient temperatures. They breathe more ‘efficiently’ than other insects, achieving more CO2 emission per

respiration cycle at comparable respiration frequencies. Abdominal ventilation movements at rest were not uniform pumping movements but also included movements of legs antennae and wings, and lateral flipping of the abdomen. Results suggest that respiration efficiency was increased by long duration of these ventilation movements. The research was funded by the Austrian Science Fund (FWF): P20802-B16, P25042-B16. We greatly appreciate the help with electronics by G. Stabentheiner and with data evaluation by M. Bodner, M. Brunnhofer, M. Fink, P. Kirchberger, A. Lienhard, L. Mirwald and A. Settari. We

also thank W. Schappacher for his help in clarifying some quirks with data conversion, two anonymous reviewers for helpful comments and the editor D.L. Denlinger. “
“The defense response to infection in insets is in part mediated by the hemocytes. This cellular response includes phagocytosis, hemocyte aggregation around the invader (nodulation), and formation of a multicellular capsule involving Adenylyl cyclase the invader (encapsulation). The cellular response is often accompanied by a humoral response which relies on enzyme cascades for hemolymph coagulation, activation of the phenoloxidase system in hemolymph leading to melanization and production of cytotoxic reactive oxygen species and reactive nitrogen species. In addition, several antibacterial peptides induced by infection in the hemocytes and fat body are secreted into the hemolymph (as reviewed by Gillespie et al., 1997 and Marmaras and Lampropoulou, 2009). The limitations of the immune response due to its physiological cost have been described in insects; indeed, mobilizing available resources to combat infection often comes at the expense of other needs (Schmid-Hempel, 2005). For example, Drosophila females exposed to dead bacteria lay fewer eggs, presumably because resources for egg production are redirected to synthesizing defense molecules ( Zerofsky et al., 2005).

A imunorreatividade «para»

Hep-par-1 revelou-se inconclusiva. O restante parênquima hepático apresentava aspetos diagnósticos de cirrose, com depósitos de cirrose. Foram consideradas as seguintes hipóteses de diagnóstico: colangiocarcinoma, find more hepato-colangiocarcinoma, CHC esclerosante. O caso foi apresentado em reunião de grupo oncológico hepatobiliar, que propôs tratamento cirúrgico da lesão. Em março de 2009 foi realizada laparotomia exploradora e, per-operatoriamente, uma ecografia hepática, que revelou nódulos hepáticos dispersos. Foram realizadas biopsias da lesão hepática com cerca de 10 cm, dos segmentos vii e viii e de 2 outros pequenos nódulos, sem outras intervenções. No exame histológico observaram-se características sobreponíveis às identificadas na biopsia percutânea. O caso foi enviado para consulta em centro de referência internacional (Centro Médico da Universidade de Chicago, EUA), cujo relatório descreve, no exame histológico, um parênquima hepático cirrótico, infiltrado por neoplasia maligna constituída por células dispostas em ninhos e cordões, com estruturas glandulares ocasionais, apresentando as células neoplásicas citoplasma eosinofílico e ligeiramente granular.

No exame imunocitoquímico observou-se positividade das células tumorais «para» o click here CK7 e CK19 e negatividade para o Hepar-1, CD10 e glipicano 3, sugerindo-se o diagnóstico de colangiolocarcinoma, que, pelas características morfológicas, poderá corresponder à entidade recentemente descrita como colangiolocarcinoma4. A evolução clínica após a cirurgia foi desfavorável, com descompensação da insuficiência hepática e rápida progressão da doença, tendo o doente falecido um mês depois. A hemocromatose é uma anomalia hereditária da população caucasiana, na qual a incidência da expressão da doença é de um em 300-4005. O gene da hemocromatose foi identificado no braço curto do cromossoma 6 e, 80-90% dos doentes são homozigotos para a mutação C282Y6. Complicações major da HH são a cirrose e o carcinoma hepatocelular. O carcinoma primário do fígado é responsável por até 45% das mortes em doentes com HH.

O risco relativo para o desenvolvimento duma neoplasia primária do fígado em doentes com HH e cirrose é cerca www.selleck.co.jp/products/AG-014699.html de 200 x superior ao da população em geral. A maior parte dos tumores hepáticos, na HH, corresponde ao CHC clássico. No entanto, raros casos isolados de colangiocarcinoma (CC) foram relatados. A incidência de tumores hepáticos com diferenciação colangiolar permanece por esclarecer 2. O colangiolocarcinoma (CLC) é um tumor maligno primário do fígado, que é responsável por menos de 1% de todos os cancros primários do fígado, sendo portanto muito raro4. O CLC é categorizado pela Organização Mundial de Saúde (OMS) como um subtipo de hepato-colangiocarcinoma com características de células estaminais3. Esta entidade engloba 3 subtipos: o subtipo típico de células estaminais, o subtipo de células intermédias e o subtipo de colangiolocarcinoma3.

A smaller study (N = 39) by the same group reported no differences in patient survival, graft survival, or BPAR incidence between patients receiving SRL/standard TAC and those receiving SRL/reduced TAC (Table 1) [49]. However, 38% and 6% of patients on standard TAC were discontinued due to TAC nephrotoxicity and thrombotic microangiopathy, respectively. Several factors may have contributed to the apparent increased nephrotoxicity, including the study population (79% black), use of kidneys from deceased

donors, and high incidence of delayed graft function (59%). Two-year data compared similar regimens in 132 live donor renal allotransplant patients [50]. The efficacy outcomes were patient survival and graft

survival, BPAR incidence, and graft function. At 2 years, renal function TGF-beta inhibitor was significantly improved with the TAC-free regimen (SRL/MMF), compared with Ixazomib the SRL/TAC-sparing regimen, as measured by serum creatinine level and calculated GFR (both p < 0.05; Table 1). In addition, the rate of acute rejection was numerically lower in the TAC-free group (13.5% vs 18.5%; p = ns). Three-year results from a long-term study (N = 150) comparing SRL/TAC, MMF/TAC, and SRL/CsA are also available [51]. At 3 years, patient survival, graft survival, and BPAR incidence did not differ significantly among the 3 groups (Table 1), although the latter showed a trend in favor of MMF/TAC (p = 0.07). Although renal function (as measured by creatinine) was acceptable in each of the 3 groups, the MMF/TAC group was statistically more favorable when compared with SRL/CsA at 12, 24, and 36 months

(p = 0.02, p = 0.05, ALOX15 and p = 0.04, respectively) and SRL/TAC at 24 months (p = 0.05). Rates of NODM by year 3 were lowest with MMF/TAC (11% vs 27–31% in other groups). Longer-term follow-up of the same study (median of 8 years) showed significant differences or trends with respect to the above endpoints that consistently favored MMF/TAC over the other regimens [52]. Viral infections and need for antilipid therapy were significantly lower with MMF/TAC versus the other regimens combined (p < 0.05), and the incidence of NODM was numerically lower with MMF/TAC (Table 1). Similar long-term findings were reported by Chhabra and colleagues [53]. In their study, 82 renal transplant recipients were followed for up to a mean of 8.5 years. MMF/TAC provided better efficacy and safety than SRL/TAC, with significant differences seen for graft survival and GFR (Table 1). In summary, results to date are derived mainly from single-center studies, and thus more robust data are needed to confirm the preliminary findings. Two small-scale studies compared reduced-dose TAC versus standard-dose TAC, when used in combination with SRL [47] and [49].

Additionally we found that the positive associations between fat mass and bone size and the negative associations between fat mass and volumetric density persisted after adjustment for lean mass, suggesting find more that the relationships were not mediated by muscle mass. The emerging evidence that fat is not an inert tissue, but a highly active endocrine organ, yields some additional possible explanations. Adipocytes produce leptin, a

peptide hormone involved in the regulation of fat metabolism and appetite through hypothalamic mechanisms [19]. Recent work in animals has suggested that the primary effect of leptin on bone formation is negative via hypothalamic action on the sympathetic nervous system [20]. How this relates to mechanisms in humans is as yet unclear. Conversely leptin may push mesenchymal stem cells towards differentiation to osteoblasts rather than adipocytes [21] and [22] and leptin receptors have been found on osteoblasts, chondrocytes and bone marrow stromal cells [23]. Thus it is possible that leptin may explain some of the relationship between fat mass and bone, both positive and negative. Adiponectin is another hormone released by adipocytes;

in contrast to leptin it is negatively related to overall fat mass. Adiponectin is associated with increased insulin sensitivity and improved glucose tolerance. A recent study from a large UK cohort related adiponectin, measured Selleck Cyclopamine at 9.9 years, cross-sectionally to bone indices measured by DXA, and longitudinally to those measured by pQCT at 15.5 years [24]. The direct relationships between fat mass and volumetric density were not reported but total fat mass was negatively related to adiponectin concentration, which in turn negatively predicted volumetric density at 15.5 years. It seems unlikely, therefore, that adiponectin could explain negative relationships between fat mass and volumetric bone density. Insulin has been shown to have positive effects on bone in animal studies [25], with insulin resistance (and higher levels of insulin, as might be found in obesity)

associated with increased BMD [26], [27] and [28] and reduced fracture risk in humans [29]. Finally, recent work has suggested a role for peroxisome proliferator-activated receptors (PPARs) in the regulation of bone mass; reduced osteoblast Silibinin function [30] and [31], increased osteoclastogenesis [32] and altered adipocyte/osteoblast differentiation [33] have been demonstrated in animal studies; thiazolidinedione drugs, which activate PPAR-gamma, have been shown to increase fracture risk [34]. Subtypes of these nuclear receptors also have a role in regulating insulin sensitivity and lipid metabolism [35], and thus are likely to relate to obesity, but there are currently insufficient data to allow detailed conclusions regarding any bone-related role in humans to be made.

In contrast, the droplet culture requires less than 1 week because temporal observations are possible for evaluating cell growth. In addition to growth improvement, the number of colonies formed in droplet

culture was approximately 70% whereas that in solid culture was less than 10% of the number of cells before culture. Therefore, we concluded that micro-compartmentalized droplet cultivation of S. elongatus was successfully conducted using dodecane as the Volasertib organic solvent phase. Cell growth was evaluated for cyanobacteria cultured under conditions of 1 cell/droplet using the droplet culture method. S. elongatus was cultured in the presence or absence of chloramphenicol. A concentration of 15 μg/mL chloramphenicol was used; this concentration is sufficient for arresting cell growth in test tube cultures. Fig. 5 shows the population Fluorouracil of compartmentalized cells within each droplet. Approximately 30% of droplets contained single cells. The percentage of droplets containing zero, two, or three cells was 8, 23, and 18%, respectively. After culturing droplets for two and four days, cell growth was evaluated using fluorescence microscopy. In cultures without chloramphenicol, we could confirm growth from single cells. We observed changes in the cell population for each droplet. After two days of culturing, 48% of droplets contained five or more

cells. After four days of culturing, this number further increased and approximately 72% of droplets contained more than five cells. On the other hand, little growth was observed for cultures grown with chloramphenicol. Following the addition of antibiotics, changes in the cell population for each droplet indicated that the droplet cultivation method could be applied to mutant screening after transformation. Furthermore, daughter cells were observed to divide near parent cells ( Fig. 4 and Fig. 5). Therefore, even if all droplets did not contain single cell, cell growth could be continuously observed under the microscope. In this study, droplet cultures were constructed using dodecane as an oil phase with little observed cytotoxicity. The

oil phase resulted Rebamipide in an increased CO2 supply to the droplet medium, and specific growth rates were higher compared to those observed for liquid cultures grown under normal air conditions. We anticipate that droplet culture can be applied to high-throughput screening for the acquisition of useful mutants, such as high-growth strains and strains resistant to specific metabolic products. In addition to these applications, we hope this method can be applied to single colony isolation for other microalgae that are able to fix CO2 and are difficult to grow on agar plates due to drying. This research was supported in part by the Japan Science and Technology Agency (JST), CREST, entitled by “Bioalcohol production using synthetic pathway in cyanobacteria”. We would like to express gratitude to Dr. M.

The cytotoxic effect of P. motoro venom, mucus and bacterial culture supernatants on human epithelial cells (HEp-2) was determined by the MTT method which measures the viability of cells in terms of their mitochondrial metabolic rate. Accordingly, MS-275 purchase 100 μL of DMEM (Dulbecco’s Modified Eagle’s Medium) containing 106 cells was added to each well of 96 well cell culture plates and incubated for 24 h at 37 °C in a 5% CO2 incubator. After incubation, the medium was discarded and either

100 μL of different concentrations of tissue extract (5 mg, 1 mg, 0.5 mg and 0.1 mg), 100 μL of mucus (v/v) or 100 μL of bacterial culture previously grown for 18 h in DMEM were added to the plates and incubated overnight at 37 °C in a 5% CO2 incubator. After incubation the supernatant was discarded and 20 μL of a 5% solution of MTT in PBS was then added into each well and the plates were incubated for 2 h at 37 °C. One hundred microliters of Triton (1%) was used as positive control. Subsequently, 100 μL/well of methanol (100%) was added to the plate and then incubated for further 10 min. After incubation, the absorbance of each sample was determined at 570 nm in a Spectronic 20 Genesys 1 spectrophotometer. Results were expressed as mean ± SD. Single criterion ANOVA followed

by Bonferroni’s test was used to analyze the data, using SigmaStat 3.0 software. Values with p < 0.05 were considered statistically significant. In order to determine the species of bacteria present in the mucus of P. motoro SB203580 rays or environmental water, 89 bacterial strains obtained either from the mucus of P. motoro rays

(n = 24) or from the Alto Paraná river water were isolated and identified. The results showed that only 3.4% of all isolates were Gram positive and they were found only in the mucus. A total of fifteen different species of Gram-negative bacteria were identified, however, Acinetobacter spp., P. aeruginosa, Klebsiella pneumoniae, Klebsiella oxytoca, Serratia spp., Shigella spp. and Enterobacter spp. were encountered only in the mucus whereas Plesiomonas shigelloides and Citrobacter koseri were found only in the water. Six bacterial species, A. hydrophila, Aeromonas sobria, Pseudomonas putida, C. freundii, E. coli and Enterobacter mafosfamide cloacae were encountered in both, water and mucus samples ( Table 1). The API 20E and 2API 20NE kits, casein agar and erythrocyte hemolysis assays were utilized to determine the ability of all Gram-negative bacterial isolates to produce gelatinase, caseinase and hemolysin respectively. The results showed that all A. sobria, A. hydrophila and P. aeruginosa strains produced gelatinase. All A. sobria and to a lesser extent, other Gram-negative strains produced hemolysin. Caseinase was produced only by A. sobria, A. hydrophila, P. aeruginosa and C. freundii strains ( Table 2). The antimicrobial profile of each Gram-negative bacterial isolate was determined by the standard disk diffusion method.

5% saponin for 15 minutes with repeated pipetting. A total of 10 μL of 1:10 dilution ALK inhibitor cancer rows were plated on horse serum agar plates. For heat inactivation, bacteria were kept at 56°C for 1 hour. To test inflammatory stimuli, organoids were incubated with medium containing the following substances in the final concentration: lipopolysaccharide (LPS) from Escherichia coli (1

μg/mL; Invivogen), recombinant human tumor necrosis factor (TNF)α (10 ng/mL; BD Pharmingen), recombinant human interleukin (IL)1β (100 ng/mL; Sigma-Aldrich), CpG oligodeoxynucleotide (ODN) 1668 (1 μg/mL; Enzo), and flagellin from Salmonella typhimurium (100 ng/mL; Invivogen). The reader is referred to the Supplementary Materials and Methods section for fluorescence-activated cell sorting, polymerase chain reaction (PCR) and microarray, cell viability assay, karyotyping, histology, and imaging. To generate a culture system for human gastric epithelium, we isolated gastric glands from human gastric corpus tissue www.selleckchem.com/products/CAL-101.html (Figure 1A) and observed their growth under different culture conditions. We started from the conditions for mouse gastric epithelium, 4

containing EGF, noggin, R-spondin1, Wnt, FGF10, and gastrin (ENRWFG). Isolated glands from human donors could form organoids in these conditions with very low efficiency and with a limited lifespan in vitro. We then tested a panel of growth factors and inhibitors for organoid-forming efficiency, phenotype of the organoids, and longevity of the human gastric cultures. TGFβ inhibitor,

p38 inhibitor, GSK2β inhibitor, and PGE2 were chosen because of the relevance of these respective pathways in cancer. IGF is expressed in normal gastric tissue.10 Nicotinamide suppresses sirtuin activity.19 Similar to human intestine,17 nicotinamide increased the number of human gastric organoids formed (Figure 1B and Supplementary Figure 1A). It therefore was included in the subsequent culture condition. IGF, p38 inhibitor, GSK3β inhibitor, and TGFβ inhibitor all induced budding structures in a concentration-dependent manner ( Supplementary Figure 1B) and had a positive effect on the lifespan of the organoids ( Figure 1C). PGE2 induced growth of large cysts and also prolonged the lifespan of the cultures. Addition of TGFβ inhibitor increased Nabilone the lifespan to a maximum of half a year ( Figure 1C), whereas all other factors had no such effect. We therefore only added TGFβ inhibitor to the ENRWFG culture medium. To analyze the importance of the single factors, we then withdrew each of the components from the medium. Without EGF, noggin, R-spondin1, or Wnt, organoid formation was strongly reduced and cultures deteriorated within 1–3 weeks ( Figure 1D and Supplementary Figure 1C). Removal of FGF10, gastrin, or TGFβ inhibitor allowed growth for 10–20 weeks. Removal of nicotinamide increased the lifespan of the cultures ( Figure 1D).

3 currents in human T lymphocytes ( Fig. 4B). The dose-response relationships of OcyTx2 for the inhibition of both Shaker-B and Kv1.3 channels, obtained from experiments as in A & B, are presented as the Lineweaver–Burk reciprocal-plot in Fig. 4C. The dissociation constants obtained from the corresponding slopes are 93.5 nM and 18.0 nM for Shaker-B and Kv1.3, respectively. The direct dose-response relationships are shown in Fig. 4D for the inhibition of the Shaker-B and Kv1.3 currents by OcyTx2. Fitting the Hill equation to the data points ( Fig. 4D, solid lines) yielded Kd = 96.6 nM, nH = 1.00

and Kd = 17.7 nM, nH = 1.10, respectively, in close agreement with the values obtained with the double-reciprocal plot of the points, which indicates that the toxin binds to channels with a this website 1:1 stoichiometry. Fig. 4E shows the current-voltage relationship obtained for Kv1.3 using a voltage-ramp protocol, thereby allowing the determination of the activation threshold of the Kv1.3 current in control solution and GSK2118436 datasheet in the presence of OcyTx2, Fig. 4E shows that the activation threshold of Kv1.3 does not change upon treatment with 20 nM OcyKTx2, and confirms that this peptide does not affect the voltage-dependence of the activation gating of the channel. Thus, the reduction

of the peak currents in the presence of OcyKTx2 is a consequence of blockage of the K+ current rather than an overt shift in the voltage-dependence of gating. Herein we have described the functional characterization of OcyKTx2, a 34 amino acid long peptide with four disulfide bridges and a molecular weight of 3807 Da. OcyKTx2 is the second KTx that has been purified and characterized from O. cayaporum scorpion venom. Based on sequence alignment, identity and click here phylogenetic tree analysis we propose that OcyKTx2 belongs to the KTx6 family of scorpion toxins and thus its systematic name is α-KTx6.17. It is interesting to note that all KTx6 peptides were identified in non-Buthidae scorpions, and since Buthidae scorpions are mostly studied because of their medical importance, it seems that KTx6

peptides are restricted to the Iurida (suborder) and to the superfamily Scorpionoidea, which includes the Bothriuridae, Liochelidae, Scorpionidae, and Urodacidae families. Except for α-KTx6.11 (IsTX from O. madagascariensis) and α-KTx6.16 (OcyC12, a putative sequence described in the cDNA library of O. cayaporum), all other α-KTx6 peptides were included in the same branch in the phylogenetic tree (Fig 3). In this branch were also included Vm23 and Vm24, purified from Vaejovis mexicanus smithi, two peptides belonging to α-KTx7 family from Pandinus imperator (UniProtKB P55927 and P55928), and Parabutoxin-3 (α-KTx1.10 from Parabuthus transvaalicus, UniProtKB P83112). The last one is the only peptide belonging to a Buthidae scorpion included in this branch. Most scorpion KTxs are three disulfide-bounded peptides. All members of α-KTx6 subfamily possess four S-S bridges.

The most well characterized of these are the de novo methylases, DNMT3A, and DNMT3B, which symmetrically methylate cytosines in the dinucleotide Cytosine-phosphate-Guanine

(CpG) on both strands of unmethylated DNA, and DNMT1, a so-called maintenance methylase, that adds a methyl group to the symmetric CpG on the unmethylated strand of DNA after DNA replication. From the time of the discovery that silent embryonic and fetal β-type globin genes are methylated and that the cytidine analog, 5-azacytidine, inhibits the processive methylation of hemimethylated DNA after replication, many studies have focused on DNMT1 as PI3K inhibitors in clinical trials a target for reversing globin gene silencing. Initial studies in animal models42 were followed by clinical interventions that demonstrated increased HbF expression in patients with both sickle cell anemia and β-thalassemia who were treated with 5-azacytidine.43, 44 and 45 The mechanism by which 5-azacytidine actually induces increased human fetal gamma globin gene expression has been debated, and mechanisms such as generalized cytotoxicity and induced erythroid cellular stress have been proposed.13, 46, 47, 48, 49 and 50 Nonetheless GSK2118436 nmr in well-characterized primate and human β-globin gene locus-bearing transgenic

mouse models, disruption of DNA methylation appears to MYO10 be a major mechanism of relieving ɣ-globin gene silencing, although perhaps indirectly in part.51, 52, 53, 54, 55 and 56 Despite the development of more specific inhibitors of DNMT1,

such as decitabine, which, unlike 5-azacytidine, lacks effects on RNA metabolism, concerns about the safety of this class of agents have limited clinical application in β-hemoglobinopathies. However, a recent study of low dose decitabine in β-thalassemia patients reported an increase in HbF without detectable short-term cytotoxicity or genotoxicity.57 The readers of DNA methylation are a group of proteins that preferentially bind to DNA containing symmetrically methylated CpG dinucleotides. The largest family of these are the methylcytosine-binding domain (MBD) proteins, which include MBD1, MBD2, MECP2, and MBD4.58 Of these, the role of MBD2 in regulating embryonic/fetal β-type globin gene silencing in adult erythroid cells is the most well characterized. MBD2 binds preferentially to DNA containing a high density of methylated CpGs. MBD2 has been shown to bind directly to the avian embryonic ρ-globin gene, and knockdown of MBD2 derepresses the gene in adult erythroid cells in culture.59 Knockdown of MBD2 has also been shown to induce a large increase in expression of the silent human ɣ-globin gene in human β-globin locus–bearing transgenic mice53 and in human primary CD34 precursor–derived adult erythroid cells.