Because EVS circulate in the blood flow, they serve as shuttle mo

Because EVS circulate in the blood flow, they serve as shuttle modules and signaling transducers not only in their local environment find more but also at distance from their site of origin. Classification of membrane vesicles, protocols of their isolation and detection, molecular details of vesicular release, clearance

and biological functions are still under intense investigation. EVS have been identified in the blood circulation for a long time, and have been first considered as cell fragments. In fact, EVS are quite heterogeneous and at least two main distinct types have been identified: exosomes (EXS) and microparticles (MPS). Both EXS and MPS are detected in blood flow, and arose out of cells such as platelets, leukocytes and endothelial cells [20]. EXS are small (40–100 nm in diameter), spherical vesicles of endocytic origin that are secreted upon fusion of the limiting membrane of multivesicular bodies with the plasma membrane. Red blood cell (RBC)-derived vesicles (REVS) have

been also described in blood samples obtained from patients with many different diseases as well as a storage lesion from red blood cell Belnacasan preparations dedicated for transfusion [21] and [22]. EXS contain subproteome cytosolic proteins, mRNAs and miRNAs, and are involved in intercellular signaling. In contrast, MPS bud directly from the plasma membrane and their size ranges from 100 nm to 1 μm (Fig. 1) [23]. A model of MPS formation including translocases, lipid rafts, various protein STK38 modifications and irreversible membrane rearrangements has been proposed (Fig. 2) [24] and [25]. MPS are not cell fragments or “dust” without any biological function [26]. They play a role in various broad biological functions such as thrombosis and hemostasis [20], [27] and [28], inflammation [27] and [29] or immunosuppression [30] and [31]. However, numerous similarities exist between EXS and MPS with respect to their physical characteristics and

compositions. These similarities frequently hampered the separation and purification of these EVS in body fluids and brought confusion in the scientific literature. In this review, we will mainly focus on blood EVS, with a particular emphasis on platelet and RBC EVS, as well as on MPS released during storage of blood units. For clarity purposes, the term EVS will be used in the following sections, grouping both MPS and EXS. Quantification, proteomic analysis as well as the biology of RBC-derived EVS (REVS), platelet-derived EVS (PEVS), leukocyte-derived EVS (LEVS), and of endothelial cell-derived-EVS (EEVS) are different, even if they share many common determinants. This review will present proteomic data that are “specific” for each type of EVS and then, will give insights onto the physiology of the various forms of EVS that are normally present in the blood or in blood products.

That is, using ζ=0ζ=0, setting k   and m   according to the grid

That is, using ζ=0ζ=0, setting k   and m   according to the grid spacing and holding M2,f,νhM2,f,νh, and νvνv constant, the growth rates can be plotted purely as a function of N2N2. Furthermore, beginning with an initial state where Ri=0.25Ri=0.25, it is known a priori   that N2N2 must increase by a factor of 4 to reach the stable state of Ri=1Ri=1. Then the growth rates can be calculated for a discrete set of values of N2N2 between N02 and

4N02 to predict the SI-stable value of N2N2 that will be reached, and by extension the stable value of Ri  . Note that (23) and (24) require both M2M2 and N2N2 to be constant in space and time and mTOR inhibitor the perturbations to be small in amplitude, and are approximations to the instantaneous growth rate found by holding N2N2 fixed at each instant in time. The grid spacing ΔxΔx is varied from simulation to simulation to test the hypothesis that the amount of restratification depends on how well the SI modes are resolved. The pseudo-spectral numerical solver uses a PFT�� cost Two-Thirds Rule de-aliasing (Orszag, 1971) to prevent aliasing of high-wavenumber modes, making the shortest resolved wavelength in the model λ=3Δxλ=3Δx. The higher-resolution

simulations (subscripts 1 through 5) are meant to demonstrate that the restratification can be limited by the stratification and viscosity, not necessarily the model resolution. The lowest-resolution simulations do not resolve the most-restratifying mode, and demonstrate restratification that is limited (subscript 6) and completely negated (subscript 7) due to the model resolution. The dimensional width of the domain varies according to the choice of ΔxΔx for each individual simulation, but the depth of the mixed layer is set to be 300 m in all cases. A uniform grid of size (Ny,Nz)=(128,80)(Ny,Nz)=(128,80) points is used, with the vertical grid spacing set to a constant Δz=5Δz=5 m. Using this number of points in the horizontal ensures that the domain is wide

enough to resolve aminophylline multiple SI overturning cells in all cases, and that the largest SI modes will not be excluded even in the finest-resolution runs. The vertical diffusivity κv=1×10-6κv=1×10-6 m2 s−1 was set to be very small to prevent highly stratified fluid from diffusing up from the thermocline, and for simplicity in the stability analysis (Appendix A) the vertical viscosity was set to match this value. At higher values (i.e. κv⩾1×10-4κv⩾1×10-4 m2 s−1), diffusion caused the lowest parts of the mixed layer to become stabilized to SI before the instability became nonlinear. This effectively reduced the lengthscale of the gravest vertical mode and reduced the amount of restratification that could occur.

In our slices treatment with EtOH did not result in enhanced cyto

In our slices treatment with EtOH did not result in enhanced cytokine production. It seems likely that this treatment was not strong enough to induce an inflammatory cascade in the nbM. Indeed, B-Raf assay EtOH-induced inflammation in humans has been shown after chronic alcoholism and is not

a short time effect. In addition, cytokines found in the brains of individuals after heavy EtOH consume are originally produced by the liver cells (Crews and Nixon, 2009). Thus, any lack of direct EtOH on inflammation is in line with such a peripheral inflammatory process. In hippocampal–entorhinal brain slice cultures EtOH induced inflammatory gene expression (Zou and Crews, 2010), suggesting that this region may be more sensitive to EtOH-induced cytokine upregulation than the nbM. Further studies are necessary to investigate if the lack of inflammation in our slice model is area-related or a methodological limitation. Taken together, our data show that EtOH-induced a decline of cholinergic neurons in vitro, which was partly counteracted by NGF. Inhibition of MAPK p38 and NOS ameliorated the EtOH effect suggesting a role in the underlying mechanism of EtOH-mediated effects in vitro. In conclusion, the data may suggest that direct EtOH exposure to cholinergic nbM neurons may transiently

suppress the enzyme ChAT and may not induce cell Nivolumab mouse death of cholinergic neurons, but rather may reflect a form of neuronal plasticity in response to EtOH. Cholinergic neurons in organotypic brain slices were cultured, as described in detail previously (Humpel and Weis, 2002 and Weis et al., 2001). Briefly, the basal nucleus of Meynert (nbM) of postnatal day 10 (P10) rats Dapagliflozin was dissected under aseptic conditions. Further, 400 μm slices were cut with a tissue chopper (McIlwain, USA) and placed on 30 mm Millicell-CM 0.4 μm pore membrane culture plate inserts (6–8 slices per membrane). It needs to be pointed out that a single experiment included approximately

8–12 pups. In one dissection experiment 4 pups were dissected and all brain slices were randomly distributed on all 6-wells. An experiment was normally repeated 3 times on different groups, so that a single treatment contained at least slices from 9 different rat pups. Slices were cultured in 6-well plates at 37 °C and 5% CO2 with 1.2 ml/well of slice medium (50% MEM/HEPES (Gibco), 25% heat inactivated horse serum (Gibco/Lifetech, Austria), 25% Hanks’ solution (Gibco), 2 mM NaHCO3 (Merck, Austria), 6.5 mg/ml glucose (Merck), 2 mM glutamine (Merck), pH 7.2) including 10 ng/ml nerve growth factor (NGF) for 2 weeks. It is well established that the 400 μm brain slices become thinner during the 2 weeks of incubation resulting in a thickness of approx. 100 μm, which is a sign of healthy cultures. Slices, which did not flatten were immediately removed from the experiments.

1 1 1),

comp7073 (aminopeptidase N) (EC 3 4 11 2), comp12

1.1.1),

comp7073 (aminopeptidase N) (EC 3.4.11.2), comp12788 (pancreatic triacylglycerol lipase) (EC 3.1.1.3), and comp13347 (vitellogenin-A1) (Tables 2 and S6) are shown in Fig. 1. All of the contigs, except for comp13347 (vitellogenin-A1), were specifically expressed in salivary glands; transcript ratios were 3.7 × 102 − 1.9 × 106 times higher in salivary glands than in stomach and Malpighian tubules. Of the 13 contigs examined, only comp13347 (vitellogenin-A1) was similarly expressed in salivary glands, stomach, and Malpighian tubules, with relative expression levels 1.54:1:1.72) (Fig. 1). The expression patterns were surveyed using PCR amplification for 63 of the 76 contigs (contig IDs from comp13378 to comp13413 check details and comp13407 to comp13545 in Tables S6 and 2) using cDNAs of salivary glands, stomach, and Malpighian tubules that were subjected to qRT-PCR. As a result (data not shown), 56 contigs showed amplification almost specific to salivary glands and 40 of these showed no similarity PFT�� mw to known proteins. Seven contigs showed amplification in all tissues (salivary, stomach, and Malpighian tubules): comp12773 (protein disulfide-isomerase), comp13517 (40S ribosomal protein S15), comp13506 (transferrin), comp11878 (proactivator polypeptide), comp13359 (heat shock 70 kDa protein cognate 3), comp13270 (allergen Cr-PI),

and comp13610 (peptidyl-prolyl cis–trans isomerase B). Of the 76 most highly expressed putative secretory contigs, 68 were salivary gland-specific or at least -predominant transcripts and 48 of the 66 were unknown proteins. Many highly

expressed transcripts were salivary gland-specific and unknown, which suggests that the proteins have specifically evolved in the relationship between GRH and various poaceous host plants including rice. In a previous study, NcSP84 (comp13102) was detected as the most abundant protein in both secreted saliva and salivary gland extracts of GRH Oxymatrine (Hattori et al., 2012). This protein was predicted to have three EF hand motifs and was shown to exhibit calcium-binding activities (Hattori et al., 2012). The function of salivary calcium-binding protein is expected to be the binding of calcium ions that trigger the plugging response of wounded sieve tubes on insect feeding (Knoblauch et al., 2001). In addition, calcium-binding proteins are contained in the saliva of the pea aphid (Carolan et al., 2011), although proteins with similarity to NcSP84 have not been reported. Carboxylesterases are detoxification enzymes, as are cytochrome P450 monooxygenases (P450s) and glutathione S-transferases (GSTs) in insects (Després et al., 2007), and are considered to play important roles in insecticide resistance (Silva et al., 2012 and Jackson et al., 2013). However, their functions in the salivary gland remain unknown.


“Resect and discard” (RD) is a new paradigm for management


“Resect and discard” (RD) is a new paradigm for management of diminutive (< 6mm) polyps wherein histology is determined by real-time endoscopic imaging; GDC-0941 mw polyps are then resected and discarded rather than sent for histopathological review. The ASGE states that in order to be adopted, this approach should provide >90% agreement in assignment of post-polypectomy surveillance intervals when compared to decisions based on histopathologic

review of all polyps. 1) To compare post-polypectomy surveillance recommendations between a RD approach and standard care. 2) To determine accuracy of endoscopic prediction of polyp histology. This is a prospective, observational study conducted in a single outpatient endoscopy center over 12 months. Screening and surveillance colonoscopies were performed by four academic and two community gastroenterologists. All polyps < 6mm were endoscopically imaged and histology predictions (adenoma vs. non-adenomatous polyp) were made using high-definition white light and/or narrow-band imaging (NBI) at the discretion of the endoscopist. Confidence in histologic prediction

was assessed using a visual analog scale (VAS). Diagnostic performance and accordance of recommended surveillance intervals from endoscopic imaging were compared to histopathological review of the polyps. 606 diminutive polyps were found in 315 patients (mean age 62.4 ± 8.7 years, 49% female). Histological

Epothilone B (EPO906, Patupilone) prediction RG7204 mouse could be made in 95.7% of polyps, with high confidence on VAS in 74.3%. Surveillance interval recommendations could be made for 97.4% of patients based on predictions. The accordance for recommended surveillance intervals was 82.1% compared to histopathological review. Community and academic gastroenterologists were equally accurate in their predictions (80.2% vs. 76.3%, p=0.38) and had similar accordance in recommended surveillance intervals (83.6% vs. 81.7%, p=0.74). Overall sensitivity, specificity, and accuracy of histological predictions made with high confidence were 0.81, 0.36, and 77.1% (varying 67.9-91.4%). NBI was used in 64% of predictions and did not improve accuracy of predictions (73.9% overall). Prep quality (p=0.42) and location of polyps (p=0.69) did not influence accuracy of histological predictions. Prospective RD management of diminutive polyps was not supported by our surveillance interval accordance below the 90% threshold deemed acceptable by the ASGE. Diagnostic performance using optical imaging to predict histology was equal between community and academic endoscopists. NBI utilization at the discretion of the gastroenterologist did not improve endoscopic predictions in our study. “
“The learning curve for optical diagnosis of colorectal polyps with Narrow Band Imaging (NBI) is unknown.

mappocean org), and the federal government is working on a planni

mappocean.org), and the federal government is working on a planning process called the Pacific North Coast Integrated PD0332991 ic50 Management Area. In other areas, such as the west coast of Vancouver Island, MSP has been taking place via local community, First Nations and government partnerships (i.e., West Coast Aquatic, http://westcoastaquatic.ca/plans/). While these initiatives are promising, previous discussions about MSP have been slow to get started, which has significantly impeded progress to date [15], [16] and [17]. The British Columbia Marine Conservation Analysis (BCMCA) project emerged from the interest

of a multitude of stakeholders to set the stage for MSP in British Columbia. The BCMCA (www.bcmca.ca) is a collaborative project designed to provide resource managers, scientists, decision-makers, and those with a vested interest in the marine environment with a new set of resources to inform coast-wide integrated marine planning and management initiatives. Furthermore, the BCMCA project set out to spatially identify marine areas of high conservation value and areas important to human use in Canada’s Pacific Ocean. The BCMCA is not a planning process as it does not have the ability or mandate to implement management

actions, and it does not seek to replace OSI-744 in vivo planning initiatives that are underway or in preparation. Rather, the results are intended to inform and help advance marine planning initiatives in BC by providing collaborative analyses based on the best available ecological and human use spatial data at scales relevant to a BC coast-wide analysis. The BCMCA project is coordinated by a Project Team, comprised of representatives from the Canadian government, BC government, First Nations (self-defined as observers), academia, marine users and environmental organisations, which is responsible for coordinating, organising and implementing the project. The BCMCA project’s ecological objectives were to represent the diversity of BC’s marine ecosystems across their natural range of variation, maintain viable populations of native species, sustain ecological

and evolutionary processes within an acceptable range of variability, and build a conservation network that is resilient to environmental change. The history and approach of the project has been described by Ban et al. [18], and supporting documents can be found BCKDHA online (www.bcmca.ca). The purpose of this paper is to report the process and results of the multi-year BCMCA effort, and discuss its relevance to BC and beyond. With increasing global popularity of MSP, the impetus for the BCMCA project, an interest by a diversity of stakeholders to set the stage for MSP is likely emerging in many regions of the world. The experience of the BCMCA project has the potential to provide valuable guidance to those regions seeking to jump-start planning processes by collating spatial information and carrying out exploratory analyses.

, 2011) The crude synthetic peptide was dissolved at a protein c

, 2011). The crude synthetic peptide was dissolved at a protein concentration of 6 mM in 0.1 M Tris buffer pH 7.5 and then reduced with

20 mM Thiazovivin supplier DTT at RT for 1hr. The reduced peptide was added to the folding solution containing 0.1 M Tris buffer pH 7.5 and a mixture of 0.15 mM Cysteine and 1.5 mM Cystine at a final peptide concentration of 24.5 μM. Refolding and formation of the correct disulphide bridging pattern was achieved during 2 days at 40 °C. The refolded synthetic toxin was purified by reversed-phase HPLC on a semi-preparative C18 column (Phenomenex Jupiter, USA) using a 35 min gradient from 23% to 45% of 60% acetonitrile in 0.1% TFA. Refolding was confirmed by MALDI-TOF MS and bioassay. A chromatographic comparison of synthetic GTX1-15 with the samples of native, folded and reduced peptides by RP-HPLC (Shimadzu, Japan) using an analytical C18 column (Phenomenex, Kinetex, USA). RP-HPLC analysis was achieved within a 10 min linear gradient from 5% to 60%

acetonitrile (Fig. 2D, left). GsTx1-15 elutes in these conditions as a double peak on the HPLC chromatogram once in its native form or refolded synthetic form (which both contain 3 disulfide bridges as detected by MS analysis, see Section 3.2 for details), while eluted as a single peak in the reduced synthetic form. MS analysis of the two peaks show no detectable differences (data not shown) and the fact that the native and synthetic peptides behave in a very similar way suggests that the peptide is homogenous. In a recent paper (Chunxiao et al., 2011) describing GSK 3 inhibitor the synthesis of a mature protein, the authors have also observed a homogenous protein eluting as two

peaks in HPLC chromatograms, depending also on the solvents used. Crude peptide was weighted, dissolved in water and measure at 280 nm. The reducing of the peptide was carried out by DTT which was added to a final concentration of 20 mM and incubated at RT for 1 h. The reduced peptide was subjected to oxidative folding reaction in a 2 M Ammonium acetate buffer (pH = 7.0) containing 1 mM GSH, 0.1 mM GSSG and 1 mM EDTA. The reduced peptide was added to the solution drop wise in 6 portions to a final concentration of 10 μM. The solution was stirred at 24 °C for 120 h. The refolded material was purified either by a three-step purification procedure, containing a RP-HPLC and further ion-exchange chromatography: The RP-HPLC purification was carried out using Jupiter C18 column (Phenomenex, USA) by liner gradient using 60% acetonitrile containing 0.1% TFA as buffer B. The peak fractions were joined and lyophilized. Excess contamination were removed by ion-exchange chromatography using Luna SCX column (Phenomenex, USA) by 25 min liner gradient of 700 mM potassium chloride in potassium phosphate buffer (pH = 2.5) containing 25% acetonitrile as buffer B.

Glutathione has a diversity of crucial physiological roles, but i

Glutathione has a diversity of crucial physiological roles, but it principally serves as an endogenous antioxidant. It functions as a cofactor for GPx, the major defense mechanism

against potential toxic hydrogen peroxide and other peroxides [12]. During the detoxification process of peroxides, GSSG is formed. GSH is regenerated from GSSG by the NADPH-dependent enzyme glutathione reductase (GR) [13]. Additionally, glutathione S-transferase (GST) uses GSH as a substrate to form conjugates with electrophiles, resulting in more water soluble metabolites which are more readily excreted. Glutaredoxin (Grx) utilizes the reducing power of glutathione to catalyze disulfide reductions in the presence of NADPH and GR. Grx is involved in regulation of various cellular functions, including electron transport and protein folding [14]. Because little is known about the effects of AGEs on pancreatic beta cells, we

investigated Selleck Fluorouracil the effect of CML on pancreatic cell viability and determined the activity and expression of components belonging to the glutathione system. All chemicals were purchased from Sigma-Aldrich (Steinheim, Germany) unless stated otherwise. CML was obtained from SyMO-Chem BV (Eindhoven, Netherlands). Roswell Park Memorial Institute (RPMI) compound screening assay 1640 medium, Hank’s Balanced Salt Solution (HBSS), trypsin-EDTA (1x), non-heat inactivated fetal calf serum (FCS), and L-Glutamine were obtained from Gibco (Breda, Etomidate The Netherlands). The human pancreatic beta cell line 1.1E7 [15] was obtained from HPA Culture Collections. Cells were cultured in RPMI 1640 medium with 10% non-heat inactivated FCS and 2 mM L-glutamine. Cells were maintained in T75 flasks at 37 °C in a 5% CO2 atmosphere. Cells were seeded at a density of 5000 cells per well in a 96-well plate and after overnight attaching, medium was removed and cells were washed with HBSS. CML was added to the plate in different concentrations (0–1 mM). Subsequently, cells were incubated for 24 hours. After treatment, supernatant was removed and cells were washed with PBS. Next,

100 μl of MTT solution (0.5 mg/ml in culture medium) was added and cells were incubated for 1 hour at 37 °C. After incubation, the plate was washed with PBS and the formazan crystals were dissolved in 200 μl DMSO. Cells were incubated for 30 minutes after which the absorbance at 540 nm was measured spectrophotometrically using a microplate reader. Relative viability is expressed as a percentage relative to untreated cells. The production of intracellular reactive oxygen species was measured using 2,7-dichlorofluorescein diacetate (DCFH-DA) as described previously ([16] and [17]). Cells were seeded at a density of 5000 cells per well in a 96 well plate and after overnight attaching, medium was removed and cells were washed with HBSS. Cells were then incubated with 0.5 mM CML in the presence of 10 μM DCFH-DA.

However, maintenance of live colony is costly and sometimes diffi

However, maintenance of live colony is costly and sometimes difficult. Cryopreservation of germplasm circumvents the need for maintenance of live colony and genetic material would still be available for future use. In addition, up to now, many inbred mutant and genetically modified rat strains have not been readily available to investigators around the world [1], [28], [31] and [49]. Cryobanking of embryos, sperm, oocytes are becoming

very important both for reducing the maintenance cost and improving distribution of strains [1] and [36]. Cryopreservation of sperm provides a simpler and more economical alternative to cryopreservation of embryos, Compound Library price and reduces the cost and space needed for keeping a large number of rat strains having a single mutation [1] and [35]. Sperm preservation protocols vary among species due to their inherent characteristics. There are marked species differences

in spermatozoa size and morphology. In addition, there are also more subtle differences in membrane phospholipid composition and metabolism of spermatozoa [6]. Rat sperm are known to have extreme sensitivity to suboptimal conditions such as centrifugation, pipetting, chilling, osmotic stress [34], [46] and [51] freezing and thawing [25], [34] and [35] possibly due to unusually long tail, head shape and membrane composition [12], [20] and [24]. Thus, acceptable and repeatable rat sperm cryopreservation protocol has not been achieved [57]. Post-thaw BIBW2992 sperm quality is still unsatisfactory for intrauterine insemination Ergoloid or in vitro fertilization in rats with genetic modifications [34] and [57]. Despite species variation, there are common stages to any sperm freezing protocol. All protocols involve sperm collection and extension, addition of cryoprotective agents (CPA) and cooling above 0 °C, freezing below 0 °C, storage and thawing [11]. During all of these stages, spermatozoa are exposed a number of potentially damaging stresses such as the change in temperature, osmotic and toxic stresses presented by exposure to high molar concentrations

of CPA and the formation and dissolution of ice crystals in the extracellular space [54]. Success of cryopreservation depends on sperm endurance to these insults [45] and [54]. Extenders, CPA, optimal cooling and thawing rates play important role for successful cryopreservation of sperm [10], [20], [30] and [42]. Extender composition and cooling rate have significant effects on sperm viability and there is a strong interaction between extender and cooling rate [55]. If the cooling rate is slower or faster than optimum cooling rate, this may cause irreversible damage to sperm [13], [27] and [29]. An optimum cooling rate must be slow enough to permit water to leave the cells to avoid intracellular ice formation, and fast enough to avoid severe cell dehydration and cryo-injury due to the solution effect [29].

Also, all sequences have a terminal Lys, but we do not know if th

Also, all sequences have a terminal Lys, but we do not know if they are removed

after post-translational processing as occurs in crotamine. All sequences described exhibited the characteristics of the β-defensin family, namely the six conserved Cys motif, small size (about 5 kDa), positive net charge, and high hydrophobicity ( Table 4). We analyzed three data sets by maximum parsimony: intronic sequences only, exonic sequences only, and the whole genes. In the case of snake β-defensin-like sequences, the best phylogenetic signal was obtained selleck inhibitor using the concatenated exonic and intronic sequences. In contrast, Luenser et al. (2005) analyzed caprine and ovine β-defensin-like sequences and found a phylogenetic signal only when intronic sequences were used to construct the phylogenetic tree. Phylogenetic analyses were done using parsimony and probabilistic approaches obtaining

three topologies (Fig. 3, Fig. 4 and Fig. 5). The best substitution model obtained using TreeFinder resulted in two models, TVM for intron 1 and HKY for the other partitions (intron 2, exons 1, 2 and 3) and they were used for both maximum likelihood and Bayesian analyses. All topologies showed three branches including non-β-defensins and β-defensin-like sequences of Crotalus and Lachesis and two lineages of Bothrops. hypoxia-inducible factor pathway The lineages were jararaca (B.jararaca_defensinB_01 and _02, B.atrox_defensinB_01, B.erythromelas_defensinB_01, B.pauloensis_defensinB_01, B.diporus_defensinB_03) and jararacussu (B.jararacussu_defensinB_01, B.leucurus_defensinB_01, B.neuwiedi_defensinB_02, B.mattogrossensis_defensinB_02 and 03), and the β-defensin-like genes of ‘neuwiedi’ (B. erythromelas, B. pauloensis, B. diporus, B. neuwiedi and B. mattogrossensis) and ‘atrox’ (B. atrox and B. leucurus) groups were recovered in O-methylated flavonoid both branches. Maximum parsimony and Bayesian analyses recovered B.neuwiedi_defensinB_02 together with B.matogrossensis_defensinB_01 and 02, both of the ‘neuwiedi’ group, though without support. The lineage jararaca which showed polytomy in Bayesian analysis, had low support in other analyses. The two paralogous β-defensin-like genes jararaca_01 and jararaca_02 may

have duplicated before the speciation of the ‘neuwiedi’, ‘jararacussu’ and ‘jararaca’ groups. The sequences B.mattogrossensis_defensinB_02 and _03 seem to be polymorphic sequences and not duplicated genes. In all trees, the low support of branches was probably due to lack of sequence sampling from other snake species groups as well in the same species and due to gene duplications. Thus, an increase in the number of sequences of the same species, and also a larger sampling in β-defensin-like sequences from other snake species, may improve the tree topology and branch support in future studies. The great number of gaps and only one sequence in that gap did not seem to affect the parsimony or Bayesian analyses but it seemed to be spurious in likelihood analysis.