Search

Leprosy, an infectious disease caused by Mycobacterium leprae, affects millions of people worldwide. However, little is known regarding its molecular pathophysiological mechanisms. In this study, a comprehensive assessment of human mRNA was performed on leprosy skin lesions by using DNA chip microarrays, which included the entire spectrum of the disease along with its reactional states. Sixty-six samples from leprotic lesions (10TT, 10BT, 10BB, 10BL, 4LL, 14R1, and 10R2) and nine skin biopsies from healthy individuals were used as controls (CC) (ages ranged from 06 to 83 years, 48 were male and 29 female). The evaluation identified 1,580 differentially expressed mRNAs [Fold Change (FC)≥2.0, p

Oxygen is critical for optimal bone regeneration. While axolotls and salamanders have retained the ability to regenerate whole limbs, mammalian regeneration is restricted to the distal tip of the digit (P3) in mice, primates, and humans. Our previous study revealed the oxygen microenvironment during regeneration is dynamic and temporally influential in building and degrading bone. Given that regeneration is dependent on a dynamic and changing oxygen environment, a better understanding of the effects of oxygen during wounding, scarring, and regeneration, and better ways to artificially generate both hypoxic and oxygen replete microenvironments are essential to promote regeneration beyond wounding or scarring. To explore the influence of increased oxygen on digit regeneration in vivo daily treatments of hyperbaric oxygen were administered to mice during all phases of the entire regenerative process. Micro-Computed Tomography (μCT) and histological analysis showed that the daily application of hyperbaric oxygen elicited the same enhanced bone degradation response as two individual pulses of oxygen applied during the blastema phase. We expand past these findings to show histologically that the continuous application of hyperbaric oxygen during digit regeneration results in delayed blastema formation at a much more proximal location after amputation, and the deposition of better organized collagen fibers during bone formation. The application of sustained hyperbaric oxygen also delays wound closure and enhances bone degradation after digit amputation. Thus, hyperbaric oxygen shows the potential for positive influential control on the various phases of an epimorphic regenerative response.

Invasion success by an alien species is dependent upon rate of reproduction, growth, mortality, physical characteristics of the environment, and successful competition for resources with native species. For sessile, epibenthic marine species, one critical resource is space. We examined competitive success in two invasive Indo-Pacific corals involved in competition for space in the northern Gulf of Mexico-Tubastraea coccinea and T. micranthus-on up to 13 offshore oil/gas platforms south of the Mississippi River. Still-capture photos of thousands of overgrowth interactions between the target corals and other sessile epibenthic fauna were analyzed from ROV videos collected at 8-183 m depth. T. micranthus was observed overgrowing >90% of all sessile epibenthic species which it encountered. Frequencies of competitive success varied significantly between platforms. T. coccinea was competitively superior to all competitors pooled, at the 60% level. There was little variability between T. coccinea populations. T. coccinea encountered the following species most frequently-the encrusting sponges Xestospongia sp. (with the commensal Parazoanthus catenularis), X. carbonaria, Dictyonella funicularis, Mycale carmigropila, Phorbas amaranthus, and Haliclona vansoesti-and was found to be, on average, competitively superior to them. Both T. micranthus and T. coccinea appear to be good competitors for space against these species in the northern Gulf of Mexico. Competitive success in T. micranthus was highest in the NE part of the study area, and lowest in the SW area near the Mississippi River plume. T. coccinea's competitive success peaked in the SW study area. This suggests that variation in competitive success both within and between populations of these species may be due to differences in local environmental factors.

Background. Literature data suggest that cells such as mast cells (MCs), are involved in angiogenesis. MCs can stimulate angiogenesis by releasing of several proangiogenic cytokines stored in their cytoplasm. In particular MCs can release tryptase, a potent in vivo and in vitro proangiogenic factor. Nevertheless few data are available concerning the role of MCs positive to tryptase in primary pancreatic cancer angiogenesis. This study analyzed MCs and angiogenesis in primary tumour tissue from patients affected by pancreatic ductal adenocarcinoma (PDAC). Method. A series of 31 PDAC patients with stage T2-3N0-1M0 (by AJCC for Pancreas Cancer Staging 7th Edition) was selected and then underwent surgery. Tumour tissue samples were evaluated by means of immunohistochemistry and image analysis methods in terms of number of MCs positive to tryptase (MCDPT), area occupied by MCs positive to tryptase (MCAPT), microvascular density (MVD), and endothelial area (EA). The above parameters were related to each other and to the main clinicopathological features. Results. A significant correlation between MCDPT, MCAPT, MVD, and EA group was found by Pearson’s t-test analysis (r ranged from 0.69 to 0.81; P value ranged from 0.001 to 0.003). No other significant correlation was found. Conclusion. Our pilot data suggest that MCs positive to tryptase may play a role in PDAC angiogenesis and they could be further evaluated as a novel tumour biomarker and as a target of antiangiogenic therapy.

BACKGROUND: Tryptase is a serin protease stored and released from mast cells (MCs) that plays a role in tumour angiogenesis. In this study we aimed to evaluate serum tryptase levels in colo-rectal cancer (CRC) patients before (STLBS) and after (STLAS) radical surgical resection. We also evaluated mast cell density positive to tryptase (MCDPT) and microvascular density (MVD) in primary tumour tissue. METHODS: A series of 61 patients with stage B and C CRC (according to the Astler and Coller staging system) were selected. Serum blood samples were collected from patients one day before and one day after surgery. Tryptase levels were measured using the UniCAP Tryptase Fluoroenzymeimmunoassay (Pharmacia, Uppsala, Sweden). Tumour sections were immunostained with a primary anti-tryptase antibody (clone AA1; Dako, Glostrup, Denmark) and an anti CD-34 antibody (QB-END 10; Bio-Optica Milan, Italy) by means of immunohistochemistry and then evaluated by image analysis methods. RESULTS: The mean ± s.d. STLBS and STLAS was 5.63±2.61 µg/L, and 3.39±1.47 µg/L respectively and a significant difference between mean levels was found: p = 0.000 by t-test. The mean ± s.d. of MCDPT and MVD was 8.13±3.28 and 29.16±7.39 respectively. A strong correlation between STLBS and MVD (r = 0.83, p = 0.000); STLBS and MCDPT (r = 0.60, p = 0.003); and MCDPT and MVD (r = 0.73; p = 0.001) was found. CONCLUSION: Results demonstrated higher STLBS in CRC patients, indicating an involvement of MC tryptase in CRC angiogenesis. Data also indicated lower STLAS, suggesting the release of tryptase from tumour-infiltrating MCs. Serum tryptase levels may therefore play a role as a novel bio-marker predictive of response to radical surgery. In this context tryptase inhibitors such as Gabexate and Nafamostat Mesilate might be evaluated in adjuvant clinical trials as a new anti-angiogenic approach.

Thrombophilic genetic factors PAI-1, MTHFRC677T, V Leiden 506Q, and Prothrombin 20210A were studied as risk factors in 235 Caucasian subjects: 85 patients with abdominal thrombosis (54 with portal vein thrombosis (PVT) and 31 with Budd-Chiari syndrome (BCS) without liver cirrhosis or hepatocellular carcinoma) and 150 blood bank donors. Seventy-five patients with PVT/BCS showed associated disease or particular clinical status (46 PVT/29 BCS): 37 myeloproliferative neoplasm (20 PVT/17 BCS), 12 abdominal surgery (10 PVT/2 BCS), 10 contraception or pregnancy (6 PVT/4 BCS), 7 abdominal acute disease (6 PVT/1 BCS), and 9 chronic disease (4 PVT/5 BCS); ten patients did not present any association (8 PVT/2 BCS). PAI-14G-4G, MTHFR677TT, and V Leiden 506Q were significantly frequent (OR 95% CI and χ2 test with P value) in abdominal thrombosis; in these patients PAI-14G-4G and MTHFR677TT distributions deviated from that expected from a population in the Hardy-Weinberg equilibrium (PAI-1: χ2=13.8, P

Herein, we performed microarray experiments in Schwann cells infected with live M. leprae and identified novel differentially expressed genes (DEG) in M. leprae infected cells. Also, we selected candidate genes associated or implicated with leprosy in genetic studies and biological experiments. Forty-seven genes were selected for validation in two independent types of samples by multiplex qPCR. First, an in vitro model using THP-1 cells was infected with live Mycobacterium leprae and M. bovis bacillus Calmette-Guérin (BCG). In a second situation, mRNA obtained from nerve biopsies from patients with leprosy or other peripheral neuropathies was tested. We detected DEGs that discriminate M. bovis BCG from M. leprae infection. Specific signatures of susceptible responses after M. leprae infection when compared to BCG lead to repression of genes, including CCL2, CCL3, IL8 and SOD2. The same 47-gene set was screened in nerve biopsies, which corroborated the down-regulation of CCL2 and CCL3 in leprosy, but also evidenced the down-regulation of genes involved in mitochondrial metabolism, and the up-regulation of genes involved in lipid metabolism and ubiquitination. Finally, a gene expression signature from DEG was identified in patients confirmed of having leprosy. A classification tree was able to ascertain 80% of the cases as leprosy or non-leprous peripheral neuropathy based on the expression of only LDLR and CCL4. A general immune and mitochondrial hypo-responsive state occurs in response to M. leprae infection. Also, the most important genes and pathways have been highlighted providing new tools for early diagnosis and treatment of leprosy.

Angiogenesis and lymphangiogenesis are the processes of neovascularization that evolve from preexisting blood and lymphatic vessels. There are few studies on angiogenesis and none on lymphangiogenesis in leprosy. Thus, the role of neovascularization in the pathophysiological mechanisms of the disease was studied across the spectrum of leprosy, its reactional states and its residual lesions.Seventy-six biopsies of leprosy skin lesions and seven healthy controls were selected. Fifty-five serum samples were used for the detection of CD105 by ELISA. Histological sections were stained with antibodies against CD31 (blood and lymphatic vessels), D2-40/podoplanin (lymphatic vessels), and CD105/endoglin (neovessels). Microvessels were counted in 100 high-power fields (400x) and the number of vessels was evaluated in relation to the extension of the inflammatory infiltrate (0-3), to the bacillary index (0-6) and to the clinical forms. Angiogenesis, as marked by CD31 and CD105, was observed across the leprosy spectrum, compared with the controls. Additionally, there was a positive correlation between these markers with extension of the infiltrate (p

Background. Angiogenesis is a complex process involved in both growth and progression of several human and animal tumours. Tryptase is a serin protease stored in mast cells granules, which plays a role in tumour angiogenesis. Mast cells (MCs) can release tryptase following c-Kit receptor (c-KitR) activation. Method. In a series of 25 gastric cancer patients with stage T3N2-3M0 (by AJCC for Gastric Cancer 7th Edition), immunohistochemistry and image analysis methods were employed to evaluate in the tumour tissue the correlation between the number of mast cells positive to tryptase (MCPT), c-KitR expressing cells (c-KitR-EC), and microvascular density (MVD). Results. Data demonstrated a positive correlation between MCPT, c-KitR-EC, and MVD to each other. In tumour tissue the mean number of MCPT was 15, the mean number of c-KitR-EC was 20, and the mean number of MVD was 20. The Pearson test correlating MCPT and MVD, c-KitR-EC and MVD was significantly (r=0.64, P=0.001; r=0.66, P=0.041, resp.). Conclusion. In this pilot study, we suggest that MCPT and c-KitR-EC play a role in gastric cancer angiogenesis, so we think that several c-KitR or tryptase inhibitors such as gabexate mesilate and nafamostat mesilate might be evaluated in clinical trials as a new antiangiogenetic approach.

Increases in Sea Surface Temperatures (SSTs) as a result of global warming have caused reef-building scleractinian corals to bleach worldwide, a result of the loss of obligate endosymbiotic zooxanthellae. Since the 1980's, bleaching severity and frequency has increased, in some cases causing mass mortality of corals. Earlier experiments have demonstrated that zooxanthellae in scleractinian corals from three families from the Great Barrier Reef, Australia (Faviidae, Poritidae, and Acroporidae) are more sensitive to heat stress than their hosts, exhibiting differential symptoms of programmed cell death - apoptosis and necrosis. Most zooxanthellar phylotypes are dying during expulsion upon release from the host. The host corals appear to be adapted or exapted to the heat increases. We attempt to determine whether this adaptation/exaptation occurs in octocorals by examining the heat-sensitivities of zooxanthellae and their host octocoral alcyonacean soft corals - Sarcophyton ehrenbergi (Alcyoniidae), Sinularia lochmodes (Alcyoniidae), and Xenia elongata (Xeniidae), species from two different families. The soft coral holobionts were subjected to experimental seawater temperatures of 28, 30, 32, 34, and 36°C for 48 hrs. Host and zooxanthellar cells were examined for viability, apoptosis, and necrosis (in hospite and expelled) using transmission electron microscopy (TEM), fluorescent microscopy (FM), and flow cytometry (FC). As experimental temperatures increased, zooxanthellae generally exhibited apoptotic and necrotic symptoms at lower temperatures than host cells and were expelled. Responses varied species-specifically. Soft coral hosts were adapted/exapted to higher seawater temperatures than their zooxanthellae. As with the scleractinians, the zooxanthellae appear to be the limiting factor for survival of the holobiont in the groups tested, in this region. These limits have now been shown to operate in six species within five families and two orders of the Cnidaria in the western Pacific. We hypothesize that this relationship may have taxonomic implications for other obligate zooxanthellate cnidarians subject to bleaching.

Overpopulation of urban areas results from constant migrations that cause disordered urban growth, constituting clusters defined as sets of people or activities concentrated in relatively small physical spaces that often involve precarious conditions. Aim. Using residential grouping, the aim was to identify possible clusters of individuals in São José do Rio Preto, Sao Paulo, Brazil, who have or have had leprosy. Methods. A population-based, descriptive, ecological study using the MapInfo and CrimeStat techniques, geoprocessing, and space-time analysis evaluated the location of 425 people treated for leprosy between 1998 and 2010. Clusters were defined as concentrations of at least 8 people with leprosy; a distance of up to 300 meters between residences was adopted. Additionally, the year of starting treatment and the clinical forms of the disease were analyzed. Results. Ninety-eight (23.1%) of 425 geocoded cases were located within one of ten clusters identified in this study, and 129 cases (30.3%) were in the region of a second-order cluster, an area considered of high risk for the disease. Conclusion. This study identified ten clusters of leprosy cases in the city and identified an area of high risk for the appearance of new cases of the disease.

In this work, the perception of affordances was analysed in terms of cognitive neuroscience during an interactive experience in a virtual reality environment. In particular, we chose a virtual reality scenario based on the Leap Motion controller: this sensor device captures the movements of the user’s hand and fingers, which are reproduced on a computer screen by the proper software applications. For our experiment, we employed a sample of 10 subjects matched by age and sex and chosen among university students. The subjects took part in motor imagery training and immersive affordance condition (a virtual training with Leap Motion and a haptic training with real objects). After each training sessions the subject performed a recognition task, in order to investigate event-related potential (ERP) components. The results revealed significant differences in the attentional components during the Leap Motion training. During Leap Motion session, latencies increased in the occipital lobes, which are entrusted to visual sensory; in contrast, latencies decreased in the frontal lobe, where the brain is mainly activated for attention and action planning.

The 3,000 oil/gas structures currently deployed in the northern Gulf of Mexico (GOM) provide hard substratum for marine organisms in a region where such has been rare since the Holocene. The major exception to this are the Flower Garden Banks (FGB). Corals are known to have colonized oil/gas platforms around the FGB, facilitating biogeographic expansion. We ask the question, what are the patterns of genetic affinity in these coral populations. We sampled coral tissue from populations of two species occurring on oil and gas platforms: Madracis decactis (hermatype) and Tubastraea coccinea (invasive ahermatype). We sampled 28 platforms along four transects from 20 km offshore to the continental shelf edge off 1) Matagorda Island, TX; 2) Lake Sabine, TX; 3) Terrebonne Bay, LA; and 4) Mobile, AL. The entire population of M. decactis was sampled between depths of 5 m and 37 m. T. coccinea populations were sub-sampled. Genetic variation was assessed using the PCR-based Amplified Fragment Length Polymorphisms (AFLPs). Data were analyzed via AFLPOP and STRUCTURE. Genetic connectivity among M. decactis platform populations was highest near the FGB and decreased to the east. Connectivity increased again in the eastern sector, indicating isolation between the populations from different sides of the Mississippi River (Transects 3 and 4). A point-drop in genetic affinity (relatedness) at the shelf edge south of Terrebonne Bay, LA indicated a population differing from all others in the northern GOM. Genetic affinities among T. coccinea were highest in the west and decreased to the east. Very low genetic affinities off Mobile, AL indicated a dramatic difference between those populations and those west of the Mississippi River, apparently a formidable barrier to larval dispersal.

The present study illustrates the procedures applied for the coordinate system conversion of the historical fumarole positions at La Fossa crater, to allow their comparison with newly acquired global positioning system (GPS) data. Due to the absence of ground control points in the field and on both the old Gauss Boaga and the new UTM WGS 1984 maps, we had to model the transformation errors between the two systems using differential GPS techniques. Once corrected, the maps show a residual Easting shifting, due to erroneous georeferencing of the original base maps; this is corrected by morphological comparative methods. The good correspondence between the corrected positions of the historical data and the results of the new GPS survey that was carried out in 2009 highlights the good quality of the old surveys, although they were carried out without the use of accurate topographical instruments.

Coral exposed to short periods of temperature stress (≥1.0°C above mean monthly maximum) and/or increased frequencies of high temperatures may bolster resilience to global warming associated with climate change. We compared Montastraea cavernosa (Linnaeus, 1767; Cnidaria, Scleractinia, Faviidae) from the Florida Keys National Marine Sanctuary (FKNMS) and the Flower Garden Banks National Marine Sanctuary (FGBNMS). Thermal stress has been reported frequently within the FKNMS; however, corals in the FGBNMS experience nominal exposures to similar stressors. Corals were exposed to three temperatures (27°C, 31°C, and 35°C) for 72 h. Colonies from the FKNMS lost significantly fewer viable and necrotic zooxanthellae under conditions of acute stress (35°C) than the FGBNMS colonies. This indicates that the FKNMS corals are less temperature-sensitive than those in the FGBNMS. The observed differences point to greater prior temperature exposure and adaptation in the former versus the latter site when correlated to previous years of thermal exposure.

Expansion of an unstable GAA*TTC repeat in the first intron of the FXN gene causes Friedreich ataxia by reducing frataxin expression. Structure formation by the repeat has been implicated in both frataxin repression and GAA*TTC instability. The GAA*TTC sequence is capable of adopting multiple non-B DNA structures including Y*R*Y and R*R*Y triplexes. Lower pH promotes the formation of Y*R*Y triplexes by GAA*TTC. Here we used the bacterial transposon Tn7 as an in vitro tool to probe whether GAA*TTC repeats can attract a well-characterized recombinase.Tn7 showed a pH-dependent preference for insertion into uninterrupted regions of a Friedreich ataxia patient-derived repeat, inserting 48, 39 and 14 percent of the time at pH 7, pH 8 and pH 9, respectively. Moreover, Tn7 also showed orientation and region specific insertion within the repeat at pH 7 and pH 8, but not at pH 9. In contrast, transposon Tn5 showed no strong preference for or against the repeat during in vitro transposition at any pH tested. Y*R*Y triplex formation was reduced in predictable ways by transposon interruption of the GAA*TTC repeat. However, transposon interruptions in the GAA*TTC repeats did not increase the in vitro transcription efficiency of the templates.We have demonstrated that transposon Tn7 will recognize structures that form spontaneously in GAA*TTC repeats and insert in a specific orientation within the repeat. The conditions used for in vitro transposition span the physiologically relevant range suggesting that long GAA*TTC repeats can form triplex structures in vivo, attracting enzymes involved in DNA repair, recombination and chromatin modification.

Trinucleotide repeat expansion is the genetic basis for a sizeable group of inherited neurological and neuromuscular disorders. Friedreich ataxia (FRDA) is a relentlessly progressive neurodegenerative disorder caused by GAA.TTC repeat expansion in the first intron of the FXN gene. The expanded repeat reduces FXN mRNA expression and the length of the repeat tract is proportional to disease severity. Somatic expansion of the GAA.TTC repeat sequence in disease-relevant tissues is thought to contribute to the progression of disease severity during patient aging. Previous models of GAA.TTC instability have not been able to produce substantial levels of expansion within an experimentally useful time frame, which has limited our understanding of the molecular basis for this expansion. Here, we present a novel model for studying GAA.TTC expansion in human cells. In our model system, uninterrupted GAA.TTC repeat sequences display high levels of genomic instability, with an overall tendency towards progressive expansion. Using this model, we characterize the relationship between repeat length and expansion. We identify the interval between 88 and 176 repeats as being an important length threshold where expansion rates dramatically increase. We show that expansion levels are affected by both the purity and orientation of the repeat tract within the genomic context. We further demonstrate that GAA.TTC expansion in our model is independent of cell division. Using unique reporter constructs, we identify transcription through the repeat tract as a major contributor to GAA.TTC expansion. Our findings provide novel insight into the mechanisms responsible for GAA.TTC expansion in human cells.

Making the correct choice between transcription elongation and transcription termination is essential to the function of RNA polymerase II, and fundamental to gene expression. This choice can be influenced by factors modifying the transcription complex, factors modifying chromatin, or signals mediated by the template or transcript. To aid in the study of transcription elongation and termination we have developed a transcription elongation reporter system that consists of tandem luciferase reporters flanking a test sequence of interest. The ratio of expression from the reporters provides a measure of the relative rates of successful elongation through the intervening sequence.Size matched fragments containing the polyadenylation signal of the human beta-actin gene (ACTB) and the human beta-globin gene (HBB) were evaluated for transcription termination using this new ratiometric tandem reporter assay. Constructs bearing just 200 base pairs on either side of the consensus poly(A) addition site terminated 98% and 86% of transcription for ACTB and HBB sequences, respectively. The nearly 10-fold difference in read-through transcription between the two short poly(A) regions was eclipsed when additional downstream poly(A) sequence was included for each gene. Both poly(A) regions proved very effective at termination when 1100 base pairs were included, stopping 99.6% of transcription. To determine if part of the increased termination was simply due to the increased template length, we inserted several kilobases of heterologous coding sequence downstream of each poly(A) region test fragment. Unexpectedly, the additional length reduced the effectiveness of termination of HBB sequences 2-fold and of ACTB sequences 3- to 5-fold.The tandem construct provides a sensitive measure of transcription termination in human cells. Decreased Xrn2 or Senataxin levels produced only a modest release from termination. Our data support overlap in allosteric and torpedo mechanisms of transcription termination and suggest that efficient termination is ensured by redundancy.

A poliquimioterapia/Organização Mundial da Saúde foi implantada efetivamente no Brasil em 1991, contribuindo drasticamente para redução da taxa de prevalência e cura da hanseníase. No entanto, a sua comprovada eficácia não tem impedido a ocorrência de recidiva da doença. Falha no tratamento, persistência bacilar ou resistência a drogas são fatores que podem ou não estarem associados a ela. O objetivo deste estudo foi verificar a ocorrência de recidiva e associá-la com a presença de cepas resistentes do Mycobacterium leprae entre 28 indivíduos que apresentaram suspeita clínica de recidiva após tratamento por monoterapia sulfônica, esquema da Divisão Nacional de Dermatologia Sanitária ou poliquimioterapia. Biópsias das lesões de pacientes multibacilares, com diagnóstico clínico de recidiva, atendidos por demanda espontânea, foram coletadas para avaliar resistência a drogas por meio da técnica de inoculação em pata de camundongo. Dentre as amostras avaliadas 42,8% apresentaram bacilos sensíveis à dapsona e rifampicina e 10,7% apresentaram resistência à dapsona; não foram isolados bacilos resistentes à rifampicina. A emergência de bacilos resistentes, especialmente à rifampicina, é um alerta para os programas de controle da hanseníase. Monitorar a disseminação destas cepas é importante, pois elas apresentam um sério obstáculo para a eliminação da doença, principalmente em países onde a hanseníase ainda é endêmica.