The mesozooplankton associated with the pelagic realm over the reef formation was characterized, considering the estuarine plume and oceanic influence. Vertical hauls were carried out using a standard plankton net with 200 μm mesh size during September 2014. An indicator index was applied to express species importance as ecological indicators in community. Information on functional traits was gathered for the most abundant copepod species. Overall, 179 zooplankton taxa were recorded. Copepods were the richest (92 species), most diverse and most abundant group, whereas meroplankton were rare and less abundant. Species diversity (>3.0 bits.ind-1) and evenness (>0.6) were high, indicating a complex community. Small holoplanktonic species dominated the zooplankton, and the total density varied from 107.98 ind. m-3 over the reef area to 2,609.24 ind. m-3 in the estuarine plume, with a significant difference between coastal and oceanic areas. The most abundant copepods were the coastal species ithona plumifera and Clausocalanus furcatus and early stages copepodites of Paracalanidae. The holoplanktonic Oikopleura, an important producer of mucous houses, was very abundant on the reefs. The indicator species index revealed three groups: (1) indicative of coastal waters under the influence of the estuarine plume ; (2) characterized coastal and oceanic conditions ; (3) characterized the reef system (O. plumifera). Two major copepods functional groups were identified and sorted according to their trophic strategy and coastal-oceanic distribution. The species that dominated the coastal area and the area over the rhodolith beds are indicators of the estuarine plume and are mixed with species of the North Brazil Current. These species practically disappear offshore, where occur oceanic species commonly found in other oligotrophic tropical areas. This ecosystem shows a mixture of estuarine, coastal and oceanic communities coexisting in the waters over the Amazon reefs, with no significant differences among these areas. However, the MDS clearly separated the communities along the salinity gradient in the plume.At the mouth of the Amazon River, a widespread carbonate ecosystem exists below the river plume, generating a hard-bottom reef (∼9500 km Coastal waters with a riverine influence are extremely productive, ecologically and societally valued ecosystems . These wGiven the overall importance of the coastal area of Amazonia and to understand the various complex interactions that link estuarine-costal-oceanic systems, two complementary multidisciplinary projects were proposed. NOVOBIOMA aimed to characterize the distribution, structure and biodiversity associated with the reef formations of the Brazilian North Coast  and the major physical, chemical and biological properties of the water column overlying this Brazilian biome, which has been little studied to date . The CamThe role of zooplankton in the food web as energy transfer, linking bottom–up  to top–down elements  make it an important indicator of ecological conditions , thus pr6 km2 that extends to the western tropical North Atlantic  that influence water masses in the marine milieu . The higAtlantic , generatAtlantic .In most tropical and subtropical marine waters, the zooplankton community is dominated by copepods, which occupy key trophic positions . The funThe available zooplankton data for the Amazon coast are very limited because this is one of the areas in the Atlantic equatorial region with little available information . The relWithin this context, our hypothesis is that the reef and oceanic zooplankton is modified by the Amazon estuarine plume. Thus, the present study aims to investigate the structure and function of the mesozooplankton community over the recently rediscovered reef under the influence of the Amazon estuarine plume and the role of the plume on the zooplankton from coastal to oceanic waters, including retroflection effects. We highlight the composition and abundance of mesozooplankton, ecological descriptors  in this unique carbonate reef system.The Amazonia Coast of Brazil extends from the mouth of the Parnaíba River to the mouth of the Oiapoque River. This coast has been subdivided into three sectors: the Guyanese Coast, Golfão Marajoára and Amazonian Eastern Coast . The widCoastal currents represent one of the most important mechanisms that play a role in the forcing of the ACS . The Nor5 m3 s-1 of fresh water onto the ACS . This zone contains a clear oxycline between the plume and sub-plume waters, with anaerobic metabolism in the plume and sulfate reduction/S oxidation in the sub-plume layers. (2) The central zone containing sand waves, where the same crustose calcareous algae and sponges occur. 3) The southern zone, characterized by low-relief patchy reefs with the dominance of scleractinian corals (The Amazon reef area presents three main zones. (1) The northern zone, composed by shelf-edge reefs with abundant crustose calcareous algae (including rhodoliths), and sponges . In this campaign, two field projects were developed: NOVOBIOMA and Camadas Finas V, on board the research vessel HOc. Cruzeiro do Sul – H38 (DHN/Brazilian Navy).The survey was performed during a single campaign carried out from September 24 to 29 of 2014, along the Brazilian North Coast   and Camadas Finas V (St. 1 to St. 17) were sampled using a ring plankton net (200 μm mesh size), which was hauled vertically to the surface from a depth of 5–7 m from the bottom in the nearshore area and 150 m offshore of the shelf break. A flowmeter  was fitted onto the opening of the net. Samples were collected and preserved in a 4% buffered formalin-seawater solution. Sestonic biomass was estimated via the wet-weight method . No signZooplankton species were identified using the manuals produced by Abiotic data  and chlorophyll-a were determined using a CTD. The thermohaline water mass were differentiated as follows: South Atlantic Central Water ; coastal water ; and tropical water .t-tests or the Mann–Whitney rank test when the data did not present equal variance and normality (Shapiro–Wilk)  between sampling areas  were performed with unpaired ro–Wilk) . The strro–Wilk) , and eveMultivariate analyses included the following: (i) non-metric multidimensional scaling (nMDS) based on the Bray-Curtis distance matrix of the ln (x + 1) transformed biological data; (ii) a permutational multivariate analysis of variance (PERMANOVA)  based onp < 0.05, and indicator value indices  of more than 60% were considered. To calculate IndVal, the program PC-ORD, Version 6.08, from the MJM software (1995–2011) was used. Because each IndVal is calculated independently of other species assemblages, comparisons of indicator values can be performed between taxonomically unrelated taxa, taxa in different functional groups, and taxa in different communities. Comparisons across taxa are sensitive to differences in abundance that may be due to differences in sampling methods  was performed to estimate the fidelity and specificity of the species in the coastal and oceanic groups. The statistical significance of the indicator values for the species was evaluated using a Monte Carlo permutation test (1000 permutations) . The ind methods .A literature review of the functional traits of marine copepods was carried out, resulting in a matrix for assessing functional traits .In this study, three water masses were recorded: coastal water (CW), characterized by high temperature and low salinity; tropical water (TW), which is relatively warm and salty; and South Atlantic Central Water (SACW), with low temperatures and salinities between 34.5 and 36. CW forms over the inner portion of the continental shelf as a result of non-isopycnal mixing between river runoff water, estuarine plumes, SACW and TW. Due to the influence of the continental discharge, CW exhibits low salinity (below 35.4), and its temperature is subject to spatial and seasonal variation, exhibiting values between 27°C and 30°C in the present study. CW was mainly restricted to the western side of the river mouth, at stations 10–16, in the 50 m of water depth, possibly being transported westward by the North Brazil Current (NBC). The presence of CW at stations 10–12 was probably due the retroflection of the NBC .TW was present mainly at stations 5–13, on outer continental shelf, was confined to depths between 50 and 200 m of water depth, and exhibited temperatures between 20°C and 27°C and salinities varying from 36 to 36.5. SACW was also found at stations 5 to 13, from 150 to 450 m, and displayed temperatures between 7°C and 18°C and salinities from 34.5 to 36 (-1 (St. 13) to 0.32 μg L-1 (St. 1), with average of 0.10 ± 0.07 μg L-1 . The general pattern observed was that high values occurred under the influence of the Amazon plume at coastal stations, while lower values were observed at oceanic stations.The chlorophyll-a concentration varied from 0.05 μg LTables 2), were identified among the zooplankton. Holoplankton contributed 75% of the zooplankton community, and the copepods were by far the most abundant taxa. A total of 92 species of copepods were recorded, 65% of which were calanoids.A total of 179 taxa, consisting of Radiozoa, Foraminifera, Cnidaria, Mollusca, Annelida, Nemertea, Crustacea, Bryozoa, Echinodermata, Chaetognatha, and Chordata  speciosus, Oncaea media and Farranula gracilis and the chaetognath Flaccisagitta enflata, the appendicularian Oikopleura (Vexillaria) dioica, and Hydrozoa and Polychaeta (larvae). These taxa comprised 5.8% of the mesozooplankton community. In the study area, 14.8% of the taxa in the community were frequent (45–74% frequency of occurrence), whereas 31.8% of the taxa in the community presented a frequency of occurrence between 15 and 44%. The rare taxa (<14% frequency of occurrence) accounted for 47.6% of the mesozooplankton community. Copepod species represented nearly 50% of the rare taxa.Common taxa with a wide distribution in the area (frequency of occurrence > 75%) were the copepods Figure 3B), varying from 2.09 bits ind-1 (St. 17) to 4.15 bits ind-1 (St. 14), with a mean value of 3.37 ± 0.56 bits ind-1. Higher values were registered in oceanic samples, with no significant differences among areas . Evenness was also very high, varying from 0.57 (NB6) to 0.86 (St. 16), and mean value of 0.72 ± 0.08, with no significant differences between oceanic and coastal samples .High copepod species diversity values were found in all stations (-3 (NB1) to 73.8 mg m-3 (NB2), with an average of 13 ± 18 mg m-3 . The highest biomass values were caused by high abundances of the appendicularians Oikopleura spp. and chaetognaths Flaccisagitta spp., followed by the copepods Oithona plumifera, Clausocalanus furcatus and Paracalanidae. Additionally, we observed a large amount of other elements in several samples, such as aggregates represented mainly by appendicularians houses , in addition to algae fragments, other detritus, microplastics and few fine sediments. In all samples with high appendicularian abundances, we detected high contribution of their houses.Total sestonic biomass varied from 5 mg m-3 (NB10) to 2,609.24 ind. m-3 (St. 17) with a mean of 780.46 ± 830.95 ind. m-3 . Significant differences were found between coastal and oceanic samples to the copepods Oithona plumifera, Clausocalanus furcatus and Paracanalidae (p = 0.03).The total zooplankton density varied from 107.98 ind. mOikopleura) comprised 63.1 and 28.9% of the community density, respectively.Copepods represented 60% of total community density, followed by appendicularians (24%), considering all stations together. At each station, copepods accounted for > 58%, except for St.16 (28.1%) and St.14 (49.4%). At these stations, appendicularians . A cluster group was formed by the non-reef samples from coastal zones, and another group was formed for oceanic and reef samples, which were separated into two subgroups (data not shown). A significant source of spatial variation in the taxonomic composition of the mesozooplankton community was suggested by the PERMANOVA result (p = 0.001). The observed differences were examined based on the average dissimilarity (SIMPER), and a total of 42 species contributed 70% of this spatial dissimilarity. These differences were caused by varying abundance among the following dominating species at each sampling site: Clausocalanus spp. (juvenile), Oikopleura (Vexillaria) dioica, Euterpina acutifrons, Undinula vulgaris, Parvocalanus crassirostris, Crustacea (nauplius) and Calocalanus pavo, contributing 18.6% to the observed dissimilarity.MDS showed clear separation of the oceanic and coastal samples, and in coastal samples, clear separation was observed between the reef area and other coastal areas (Table 3). The first was the strongest and was indicative of coastal waters under the influence of the estuarine plume, where the best indicator taxa were Euterpina acutifrons , Hydromedusae , Oikopleura (Vexillaria) dioica , Parvocalanus crassirostris  and Nauplius . These taxa highlight the great relevance of the riverine outflow in the area. The oceanic species group was characterized by Clausocalanus spp. (juvenile)  and the reef system group by Oithona plumifera .The Indicator Value index showed three groups of indicators . These groups were each subdivided into two subgroups, according to feeding and reproduction strategies, size, distance from the coast and migration.The trophic strategy was the major functional trait related to the sorting of the community, forming two main groups: an herbivorous-omnivorous group (group 1) and a carnivorous-detritivorous group (group 2) .Subgroup 1a was the largest group and was mostly comprised of oceanic or coastal copepods, filterers, broadcasters, non-migrant taxa and taxa with smaller sizes (500–1000 μm). This group consisted mainly of calanoids, such as those of the genera Candacia and Haloptilus genera, which are typically oceanic, epimesopelagic and relatively large in size (2500–6500 μm). Subgroup 2b was the second largest group and was divided in two groups: group 2.b.1 was mainly composed of cyclopids of the families Oncaeidae, Corycaeidae and Sapphirinidae, in addition to two important calanoids . These cyclopids are oceanic, omnivorous or carnivorous, cruiser, sac-spawner, non-migrant, epimesopelagic species. The two calanoids are filterers, broadcasters and non-migrants and are relatively small in size. Group 2.b.2 was composed of three common, but low-abundance copepods , all which are oceanic, omnivorous or carnivorous species that exhibit cruising or active ambush behavior, weak to strong vertical diel migration, an epipelagic habit and variable class sizes (1500–4500 μm).Subgroup 2a included only three species of calanoids belonging to the Oikopleura (Vexillaria) dioica], chaetognaths (Flaccisagitta enflata and Serratosagitta serratodentata), and decapods of the family Luciferidae (Belzebub faxoni). These groups were frequently found at high abundances. On the other hand, meroplankton was rare, and few groups were registered.The Amazon area showed a mixture of communities from estuary, coastal, reef and oceanic habitats, where the predominance of holoplanktonic species, mainly copepods (with 92 species), was the rule. The high species richness, diversity and evenness of copepods, with no significant differences between samples, can be explained by the habitat heterogeneity hypothesis e.g., , in whicEuterpina acutifrons, Oikopleura (Vexillaria) dioica and Parvocalanus crassirostris characterized the estuarine community, while Clausocalanus spp. (juvenile) and Oithona plumifera characterized oceanic and reef, respectively. Oithona plumifera was abundant, especially over the reef area and the region of retroflection, and predominates in oligotrophic waters of the tropical Atlantic  speciosus, Oncaea media and Farranula gracilis stood out among the zooplankton in terms of their frequency of occurrence, whereas in terms of abundance, the coastal copepod species C. furcatus, Parvocalanus crassirostris, O. hebes and Euterpina acutifrons were the most important. These last three species are characteristic of estuarine systems in Brazil  driven by trade winds prevailing in the region during almost the whole year  was found to be common in studies conducted in coastal areas of the Amazon . Group 2, in contrast, was represented by copepods with an oceanic distribution that were carnivorous and detritivorous, exhibiting cruising or active ambushing habits, larger sizes and weak to strong migration habits, behaving as sac-spawners or broadcasters .The copepod functional groups identified in the present study highlight the connections of species through their ecological functions. Two main functional groups of copepods occurred in the area, mainly in the reef one, where the Amazon plume dominates. The main factor separating the groups was their trophic strategies, which are commonly regulated by female weight, environmental temperature, and food . In termHowever, we found a large number of copepod species coexisting with apparently similar function traits in this reef area. Also in line with our findings, In our study, the general pattern of chlorophyll-a and sestonic biomass was of high values under the influence of the Amazon plume at coastal stations, including those under the reef area, while lower values were observed at oceanic stations. 8 tons per year  appears to be minimal in terms of mesozooplankton composition and structure , which are common according to previous studies elsewhere. However, our data shows a clear separation of the oceanic and coastal communities, and in the coastal community, a clear separation exists between the reef area and the other coastal areas, showing that the oceanic region also contributes with several species to the Amazon reef system. This ecosystem shows a mixture of estuarine, coastal and oceanic zooplanktonic communities, with a gradual separation among them along the salinity gradient in the plume.MA, FT, and RM coordinated the projects. SN-L, MA, DV, RS, and RM designed and coordinated the fieldwork. SN-L, PM, MM, and RS wrote the paper. SN-L, DV, RS, MM, PM, XD, AS, LF, RC, NM, AC, FT, and RM contributed with data and data analyses.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


A landmark study showed that APL can be cured without chemotherapy using arsenic trioxide (ATO) and all-v €6,500 per patient) from an Italian payer perspective.4 However, from a Canadian perspective, a combined ATO plus ATRA treatment is associated with incremental cost-effectiveness ratios of 50,193 Canadian dollars per quality-adjusted life-years.5 Moreover, in the United States, compared with the ATRA plus idarubicin regimen, first-line ATO plus ATRA treatment had incremental cost-effectiveness ratios of $4,512 per life-year saved and $5,614 per quality-adjusted life-year gained.6 These figures have to be put in context with the fact that 70% of countries in the world, which contribute to > 75% of the world’s population, have a gross national income per capita of < $10,000. Therefore, reducing the price of ATO and adopting an outpatient treatment model may overcome this problem.However, the high price of ATO in Western countries is a challenge to the health care system, limiting the maximization of its benefits. The price of ATO is approximately $676 in the United States, 530 Canadian dollars in Canada, and €393 in Europe . Three-yChina has taken important measures to improve the outcomes of patients with APL. First, the price of 10 mg of ATO is approximately $25 (¥160) and that of 10 mg of ATRA is roughly $0.2 (¥1.5) under the government price regulation policy. The total pharmacy cost of ATO and ATRA treatment is approximately $3,626 after the completion of an entire round of treatment using combined ATO and ATRA as a first-line treatment of non–high-risk patients with APL . The gross domestic product per capita of China was $7,603 in 2014, which makes the costs of the two drugs affordable even by out-of-pocket payment standards.Indigo naturalis formula) provided a similar outcome compared with intravenous ATO using a chemotherapy-included protocol in patients with APL7 and showed a reduced cost in the oral arsenic group .8Second, 70% to 85% of the costs of the two drugs are covered by three health insurance schemes , which cover > 95% of the population. Therefore, the direct economic burden of patients with APL is significantly reduced. Third, we propose a novel oral arsenic and ATRA–based outpatient treatment model to reduce the medical costs. We previously found that oral arsenic . Patients resumed their usual lifestyles during postremission therapy, and their quality of life was rated as nearly normal. A prospective, multicenter, randomized trial is underway in China , which has completed recruitment; preliminary results are awaited.We recently reported excellent, albeit preliminary, outcomes in 20 patients with non–high-risk APL treated with oral arsenic and ATRA via a largely home-based treatment protocol.Importantly, APL is a model of how targeted therapies on their own can trigger definitive cures and show the power of traditional medicine. This Chinese model of APL treatment is worth promoting, especially in undeveloped countries. With its low costs and the merits of oral administration, traditional medicine may bring hope to overcoming the high cost of the treatment of cancer.


To test the accuracy and reproducibility of ultrasound and computedtomography (CT) for the quantification of abdominal fat in correlation withthe anthropometric, clinical, and biochemical assessments.Using ultrasound and CT, we determined the thickness of subcutaneous andintra-abdominal fat in 101 subjects-of whom 39 (38.6%) were men and 62(61.4%) were women-with a mean age of 66.3 years (60-80 years). Theultrasound data were correlated with the anthropometric, clinical, andbiochemical parameters, as well as with the areas measured by abdominalCT.Intra-abdominal thickness was the variable for which the correlation with theareas of abdominal fat was strongest . We also tested the reproducibility of ultrasound and CT for theassessment of abdominal fat and found that CT measurements of abdominal fatshowed greater reproducibility, having higher intraobserver andinterobserver reliability than had the ultrasound measurements. There was asignificant correlation between ultrasound and CT, with a correlationcoefficient of 0.71.In the assessment of abdominal fat, the intraobserver and interobserverreliability were greater for CT than for ultrasound, although both methodsshowed high accuracy and good reproducibility. There isevidence of functional differences between subcutaneous fat and visceral fat; thevisualization and quantification of these compartments by imaging methods are usefulin building knowledge about these differences.There is a growing interest in the assessment of body fat distribution, especiallyfor research into metabolic syndrome, obesity and lipodystrophy studies, consideringits classifications and prognostic evaluations, as well as follow-up evaluations ofdietary and drug treatments. Knowledge about the correlation between body fatdistribution and the various anthropometric, biochemical, and functional parametersof anatomical data has increased steadily in recent years-7; various otherimaging methods, such as radiography, densitometry, and magnetic resonance imaging,have also been used for that purpose-10. Ultrasound andCT are both still prominent image methods, mainly due to their wide availability,and well as their relative ease of use and facility in obtaining measurements. It isof note that there is no consensus on which method should be used, mainly due to thedifficulty in validating their use in large-scale studies.Previous studies have shown the potential of ultrasound and computed tomography (CT)for evaluating the content of abdominal fatGiven that ultrasound and CT are the main methods used in order to evaluate body fat,especially abdominal fat, the objective of the present study was to evaluate thereproducibility and accuracy of ultrasound as a tool to study abdominal body fat,using CT as the reference. The hypothesis of the study is that, although bothmethods can be used for assessing abdominal fat, ultrasound would be preferable.This was an observational, cross-sectional study in which abdominal fat was analyzedby anthropometric and imaging methods (ultrasound and CT). The imaging exams wereevaluated by two observers who applied identical techniques. The study was approvedby the Research Ethics Committee of the Hospital das Clínicas da Faculdade deMedicina de Ribeirão Preto da Universidade de São Paulo (HCFMRP-USP),in the city of Ribeirão Preto, Brazil, and all participating subjects gavewritten informed consent.We randomly selected households of individuals residing in the region covered bythe Family Health Program of HCFMRP-USP, within a medium- to low-income areawith approximately 2000 inhabitants. Subjects between 60 and 80 years of age,living in the selected households, were invited to participate in the study. Atotal of 112 individuals agreed to participate and were referred to theHCFMRP-USP Department of Geriatrics and Radiodiagnosis. Individuals wereselected if they met all of the inclusion and exclusion criteria.The inclusion criteria were being 60-80 years of age and having cardiovasculardisease, diabetes, or another controlled chronic disease. The exclusion criteriawere as follows: being bedridden; having thyroid disease; being an alcoholic or a smoker; having undergone asurgical procedure, such as liposculpture or liposuction, in the last threemonths before the evaluation; having any incapacitating disease that wouldprevent the individual from submitting to the measurements and examinations; andusing corticosteroids, anabolic steroids, or hormone replacement therapy.Anamnesis and physical examination were performed. The study sample includedindividuals of both genders. Because of the randomized nature of the selectionprocess, individuals were selected without regard to gender, body mass index(BMI), and associated comorbidities, such as diabetes, hypertension,dyslipidemia, and metabolic syndrome. At the time of enrollment, the necessaryancillary examinations were requested. After those evaluations, 11 subjects wereexcluded because of insufficient data for the statistical analysis. Therefore,the study sample comprised 101 individuals, of whom 39 (38.6%) were men and 62(61.4%) were women. The mean age of the individuals evaluated was 66.3 ±4.2 years.2). Circumferences  were measured with a tape measure on bare skin.The presence of comorbidities was evaluated by the staff of the HCFMRP-USPGeriatric Outpatient Clinic. For all subjects, blood pressure was assessed afterfive minutes of rest. After a 12-hour fast, subjects were submitted tobiochemical evaluation to determine their lipid profile . The anthropometric evaluation,according to the criteria of the HCFMRP-USP Metabolic Clinic, included weight(in kg), height (in m), and the calculation of the BMI as weight in kilogramsdivided by height in meters squared , using a LOGIQ e ultrasound system  with multifrequency convex (3.5-5.0 MHz) and linear(7.5-10.0 MHz) transducers, as depicted in The thickness of subcutaneous fat was measured with a linear transducer at afrequency of 10.0 MHz. All subjects were evaluated in the dorsal decubitusposition, with the right arm elevated, after a 12-hour fast. The transducer waspositioned transversely at 1.0 cm above the umbilicus on the xiphoid-pubic linewithout exerting any pressure on the abdomen, to avoid underestimating thethickness. The anatomical limits for the measurement of subcutaneous fat werethe skin and the external  fascia of the rectus abdominis muscle,and the thickness was quantified in centimeters. The thickness of visceral fatwas measured with a convex transducer at a frequency of 4.0 MHz. The transducerwas positioned transversely at 1.0 cm above the umbilicus on the xiphoid-pubicline without exerting any pressure on the abdomen, to avoid underestimating thethickness The anatomical limits for the measurement of the visceral fat were theinternal (deep) fascia of the rectus abdominis muscle and the anterior wall ofthe aorta, during expiration, and the thickness was quantified incentimeters.Ultrasound examinations were performed by two evaluators  who used identical examination techniques.Each individual was evaluated on the same day, at different times, and eachobserver was blinded to the measurements obtained by the other.The subjects underwent CT of the abdomen, from the surface of the diaphragm tothe L4-L5 level of the lumbar spine. Scans were acquired with a helical CTscanner . The parameters were as follows:thickness, 10.0 mm; interval, 10.0 mm; voltage, 130 kV; current, 250 mA;acquisition time, 6 s; and matrix, 512 × 512 mm. The images were recordedin the supine position, during expiration, and after a 12-hour fast. Thesuperficial fascia defined the division between the superficial and deepsubcutaneous fat. The abdominal musculature, in continuity with the deep fasciaof the paraspinal muscles, was used in order to distinguish between subcutaneousand visceral fat. The quantity of subcutaneous fat was determined by calculatingthe difference between the total and visceral fat.® platform, version 7.0 . The area of subcutaneous fat in the L4-L5 slice was calculatedby manual segmentation of the appropriate anatomical limits , which delimits the entire abdominal area. Thesum of the areas of the fat density pixels (between -50 and -250 Hounsfieldunits) within the limits of the determined area  and the sum of the areas of pixels with adensity between -50 and -250 Hounsfield units, within the limits of thedetermined area . Each obStatistical analysis was performed using the Statistical Package for the SocialSciences, version 16.0 . The reproducibility of themeasurements was evaluated by determining the intraclass correlationcoefficients and the 95% confidence intervals.For the determination of intraobserver reproducibility, we considered the valuesof the two measurements made only by the first observer; for the determinationof interobserver reproducibility, we considered only the first measurement madeby each observer for each parameter.p ≤ 0.05.When the null hypothesis was rejected, the nonparametric Mann-Whitney test wasused. When normality was accepted, the Student’s t-test for independent sampleswas used. Student’s t-tests, Pearson’s correlation coefficient, and multipleregression analysis were used as appropriate. In the calculation of thePearson’s correlation coefficients, all correlations higher than 0.20 or lowerthan -0.20 were considered significant. For all statistical analyses, the levelof significance was set at 2 for the total abdominal area,396.6 ± 141.8 cm2 for the total area of abdominal fat, 220.8± 75.0 cm2 for the intra-abdominal area, and 136.3 ± 62.9cm2 for the area of intra-abdominal fat  were men and 6261.4%) were women. % were woinal fat .p < 0.001 andp = 0.001, respectively), indicating that the results did notdiffer among themselves in terms of the measurements, whether obtained by ultrasoundor by CT.The differences between the subcutaneous and visceral fat measurements did not differsignificantly between the two evaluators or between two measurements of a givenparameter made by the same evaluator  and 0.91 ,respectively. There is atendency in the literature to propose that ultrasound be used as a routine screeningmethod in the evaluation of visceral fat and repeated as necessary,8. However, some studies have shown that its reproducibility andobjectivity in such evaluations are highly debatable, because it is a method that ishighly dependent on the skill of the operator.The results of the present study indicate that ultrasound shows high reproducibility,with intraobserver and interobserver agreement rates of 0.94 and 0.94, respectively,for the ultrasound measurement of subcutaneous fat, compared with 0.99 and 0.98,respectively, for the ultrasound measurement of visceral fat. These findings are inagreement with those of various other studies, whocompared ultrasound with CT in a sample of 50 obese women. In that study, theauthors found that ultrasound showed a good correlation with CT , inferring, from their research, theapplicability of ultrasound in the evaluation of abdominal fat, as demonstrated in asubsequent study. In thepresent study, the thickness of the visceral fat measured by ultrasound correlatedwell with the area quantified by CT . In addition, using ultrasound, we were able to visualize and measurethe “distances” of the subcutaneous and visceral abdominal fat separately.Ultrasound offers a simple technique of abdominal fat measurement, which requiresonly the use of a 3.5-MHz transducer placed 1 cm from the umbilicus, whichcorresponds to the same area analyzed by CT. The technique was proposed in the 1990sby Armellini et al.. CT facilitatesthe differentiation between the subcutaneous and visceral adipose tissuecompartments.CT is considered the best imaging method for assessing body components because of itsreproducibility and because the fat mass thus obtained has shown correlationcoefficients above 0.90 when compared with the actual amount present in acadaver, who showed thatit is a site with a high percentage of body fat and that allows betterdifferentiation between subcutaneous tissue and intra-abdominal fat. Subsequently,Kvist et al. demonstrated thatthe L4-L5 level presents the best and highest correlation with abdominal fat in bothgenders. Another study showed that the area of visceral fat measured in a single CTslice, at the level of the umbilicus (L4-L5), correlates strongly with the totalvolume of visceral fat, which supports the use of this method for the diagnosis ofvisceral fat deposition. Inthat study, the reliability and reproducibility of the method was verified, becauseit demonstrated a maximum intraobserver variation of 1.0%, 1.1%, 5.5%, and 5.0%,respectively, between CT measurements of the abdominal area, abdominal fat area,intra-abdominal area, and intra-abdominal fat area, compared with 2.1%, 5.1%, 5.7%,and 6.0%, respectively, for the maximum interobserver variation. That indicates that CT can beconsidered the gold standard, not only for assessing abdominal fat tissue but alsofor measuring multiple compartments of the body. Chowdhury et al. showed that CT has high reproducibility by analyzing 28scans to determine the intraobserver reproducibility of abdominal fat quantificationby CT, showing an intraobserver variability of 0.4% for abdominal fat, compared with1.0% for the interobserver variability.The choice to perform the scanning in the umbilical region was initially proposed byBorkan et al.-15,19-21. However, it is necessary to consider the risksof the use of ionizing radiation for an evaluation whose clinical significance isrelative; that is, although CT provides greater reproducibility and correlatesbetter with the anthropometric data, its use does not seem justifiable in view ofthe individual risk-benefit ratio.In the present study, we evaluated how well ultrasound and CT data correlated withanthropometric data. We found that the CT data showed a stronger correlation withthe anthropometric data than did the ultrasound data. That result allows us to statethat CT is more appropriate than is ultrasound for the study of abdominal fat in thecharacterization of individual body composition, given that it shows a greatercorrelation with the various anthropometric parameters evaluated, as evidenced bythe Pearson’s correlation coefficients of 0.82, 0.77, 0.79, and 0.84, respectively,for BMI, waist circumference, hip circumference, and abdominal circumference. Thisleads us to agree with several authors who consider CT the reference for theevaluation of body fat,14.The results obtained, together with the advantages of the ultrasound method,including the fact that it is a rapid, easy to use method with good specificity andreproducibility, suggest that ultrasound is a potentially useful option for thestudy of visceral obesity in patients at high risk for metabolic syndrome, despitepresenting lower reproducibility and a weaker correlation with anthropometric datathan does CT. In the present study, we also found that ultrasound showed anexcellent correlation with certain measures, mainly BMI, serum cholesterol, andwaist circumference. Other authors have demonstrated an excellent correlationbetween ultrasound and anthropometric measures, stressing the advantages of it beingless costly than CT or MRI and more accurate than direct anthropometricmeasurementsIn the evaluation of abdominal fat, ultrasound showed good intraobserver andinterobserver agreement, as did CT to a greater degree. Therefore, both methodspresent acceptable accuracy and good reproducibility. Using CT as the reference, weobserved a correlation with ultrasound assessments of abdominal fat. CT correlatedbetter with anthropometric measures than did ultrasound, mainly for the measurementof total abdominal fat. Among the clinical and biochemical data, CT and ultrasoundboth correlated well with the serum cholesterol level.


In a multivariable logistic regression model, we identified overweight/obesity and late onset of menarche as independent risk factors for PDM. In contrast, age of first birth >27 years had a protective effect. With 16S rRNA gene sequencing, we confirmed bacterial infections were found in all PDM patients, but none of the control patients was positive on the gene expression of 16S rRNA. Our results also demonstrated significant increases of the IFN-γ and IL-12A expression in PDM, but there was no difference in IL-17A expression in these two groups. Taken together, this study suggests that reproductive factors and overweight/obesity are possible predisposing risk factors for PDM. Bacterial infection and the increased expression of some proinflammatory cytokines are associated with the pathogenesis of this disease. Periductal mastitis (PDM) is a prolonged inflammatory disease, but the cause of PDM is poorly understood. In the present case control study, 87 PDM and 87 healthy controls were enrolled and the results were evaluated to identify the significant risk factors for PDM. To investigate the roles of bacterial infection and critical cytokines expression, 16S rRNA gene sequencing and bacterial culturing were conducted. We also measured the levels of interferon- Periductal mastitis (PDM) is histopathologically defined as a chronic inflammation of the breast, with dilation of the mammary duct, plasma cell infiltration, and abscess formation. A variety of different terms, including mammary ductal ectasia and plasma cell mastitis, have been used for this condition , 2. The Thus far, the etiology of PDM is still unknown. Several factors have been associated with an increased risk for this disease, including smoking, obesity, and diabetes mellitus. However, all of these stated risks are identified based on small case series reports and have not been confirmed epidemiologically , 9. Alth Pseudomonas aeruginosa and Staphylococcus aureus [γ (IFN-γ) and IL-12; while the major cytokines featured in the Th2 subset are interleukin-4 (IL-4) and IL-10, respectively. The Th17 subset was described more recently and is simply defined by the production of IL-17. As reported, while Th1 subset controls intracellular bacterial infections, the Th2 subset controls parasitic infections and the Th17 subset controls fungal and extracellular bacterial infections, respectively [Etiologically, the exact role of bacterial infection in PDM has been debated for many years. Bacterial infection was thought to be a possible etiological factor of PDM. However, the current overall data are not conclusive as some studies also suggest sterile lesions in PDM patients without invasion of any bacteria . Among ts aureus . Althougs aureus , 16. Thes aureus . Th cellectively . Since PIn the present study, besides exploring and identifying the possible risk factors for PDM, we also investigate the relationship between bacterial infection and PDM and the significance of the expression of consequent Th1 and Th17 cytokines in this disease. Our results may help to further elucidate the pathogenesis of PDM. Furthermore, since there are very few well-designed investigations tending to explore the risk factors and possible causes for PDM in Chinese patients now, our results here may lay an important ground for the future epidemiological study on this issue in Chinese Han population.This was a retrospective nonmatched case control study. Eighty-seven patients with PDM were recruited from the Department of Breast Surgery at the Second Hospital of Shandong University. The study was conducted from January 2011 to March 2015. The inclusion criteria for patients with PDM were as follows: (I) newly diagnosed and histologically confirmed periductal mastitis; (II) Han ethnic group; and (III) no evidence or history of cancer. Eighty-seven healthy controls were collected from the Physical Examination Center of the Second Hospital of Shandong University from June 2015 to August 2015. The criteria for the control group were as follows: (I) normal results on breast physical examinations, breast ultrasound scans, and/or mammographic screening and without any possible benign breast diseases ; (II) Han ethnic group; and (III) no evidence or history of breast cancer. All eligible subjects were women in reproductive age and willing to participate in the study voluntarily. All procedures performed here involving human participants were approved by the Second Hospital of Shandong University Research Committee.Vacuum-assisted, incisional, or excisional biopsy was performed based on the clinical findings at the time of admission. Pathological results were reviewed by two pathologists. PDM patients were included in this study only if they had confirmed pathological changes. All patients with other possible causes of mammary inflammation were ruled out, such as breast tuberculosis, fat necrosis, and granulomatous lobular mastitis, as well as inflammation due to lactation or pregnancy.Possible etiologic factors were written down at the time of admission, including sociodemographic status, smoking status, alcohol consumption, reproductive history, and medical history. In addition, the following information was also obtained: patient age, marital status, menopause status, height, weight, body mass index (BMI), active and passive smoking, alcohol abuse, heart and other chronic disease history, history of allergies, history of benign breast disease, galactostasis, age at menarche, age of first child birth, parity, miscarriage history, and duration of breastfeeding. For individual patient, clinicopathologic characteristics were also collected, including major complaint on presentation, size of lesion, and histology findings.n = 12). Breast tissue samples were collected under aseptic conditions at the time of operation. Tissues were divided into two sections, with one section frozen immediately in liquid nitrogen and another section sent to the laboratory for bacterial culture. Fresh breast specimens were homogenized and incubated onto Columbia blood agar  and anaerobic blood culture bottles . The agar plates were incubated at 37°C and humidified incubator was supplied with 10% carbon dioxide. The colonies formed in the plates were picked and analyzed using a MicroScan Walkaway-96 system . Gram staining and a stain for acid-fast bacilli were conducted for all specimens. All samples were also analyzed by 16S rRNA gene sequencing. Genomic DNA was prepared using the bacterial DNA extraction kit  according to the manufacturer's recommendations. The hypervariable regions V1–V3 of the 16S rRNA gene were PCR-amplified using universal primers   [https://www.ncbi.nlm.nih.gov).For the specimen collection, patients who received any antibiotics before admission and had sinus or fistula or abscess incision drainage were ruled out from the study. Finally, 33 specimens from 31 patients, including 21 pus samples and 12 tissue samples, were collected in this study. Normal tissues adjacent to benign breast pathology  were used as control specimens (n = 31) for immunohistochemistry study of proinflammatory cytokines. Normal tissues adjacent to benign breast pathology  were used as control specimens (n = 22). Immunohistochemistry was performed on 4 μm thick sections. And then, the streptavidin-peroxidase-biotin (SP) immunohistochemical method was performed to detect the expression of IFN-γ, IL-12A, and IL-17A. Briefly, after deparaffinization and rehydration, the human tissue sections were incubated in 3% hydrogen peroxide in methanol to quench the endogenous peroxidase activity, followed by incubation with normal goat serum to block nonspecific binding. Samples were then incubated overnight at 4°C with rabbit anti-IFN-γ , anti-IL-12A , or anti-IL-17A . The secondary antibody was from the SP reagent kit . The color reaction was performed with 3,3′-Diaminobenzidine , counterstained with hematoxylin, dehydrated, treated with xylene, and then mounted. For negative controls, the antibodies were replaced with PBS. To determine the ratio of positive cells for these three cytokines, 5 fields (×400) were randomly examined per slide and scored by two pathologists who are blinded to the patients' information. The levels of IFN-γ, IL-12A, and IL-17A expression were semiquantitatively expressed using a visual grading system based on the extent of staining  and the intensity of staining . The combination of extent (E) and intensity (I) of staining was obtained by the product of E and I, with EI varying from 0 to 9 for each spot. Low expression was defined as EI ≤ 3, and others were considered high expression [For the patients who were tested for bacterial infection, tissue samples which are mentioned above were also embedded in paraffin (pression .t-test was used to compare the difference between the means of the continuous variables. Chi-square test (or Fisher exact test) was used to compare the differences between the proportions if appropriate. Risk factors for PDM were further assessed using both univariate and multivariable  logistic regression models. The continuous variables were evaluated according to the clinically relevant categories. The categories for BMI were based on the criteria of being overweight and obese for Chinese individuals  [P < 0.05 (two-tailed).Statistical analysis was performed with SPSS 17.0 software . The results were reported as counts (percentage) for the categorical variables, and mean ± standard deviation for the continuous variables. A two-sample Student's 8 kg/m2) . The cat8 kg/m2) , 23. OddP = 0.134). The majority of subjects in this study were married. There were no statistically significant differences between these two groups in terms of for the history of hypertension, type 2 diabetes mellitus, heart disease, or any history of allergy. Neither alcohol abuse nor active smoking was reported in the case and control groups. There was not any form of breast cancer in these patients of PDM confirmed by pathologic examination.To investigate the risk factors for PDM, 87 patients and 87 healthy control subjects were enrolled in this study. The characteristics of patients and controls are summarized in Most patients had unilateral breast disease, with only 8 patients exhibiting bilateral symptoms. Breast mass with or without pain was the most common complaint. The clinical presentations were summarized in P = 0.002), a late onset of menarche , having more than one child , history of benign breast disease , and nipple retraction  were statistically significant risk factors for PDM. Late age at first birth  had a protective effect for the disease. When stratified at 12 months, duration of breastfeeding was not significantly related to PDM.To examine the relationship between multiple factors and PDM, a univariate analysis was performed. The results were shown in P values < 0.2 were regarded as candidate predictors for a logistic regression model and a backward variable selection process was used. In the multivariable model (P = 0.008) and late onset of menarche  were independent predictors of PDM. In contrast, late age at first birth had a protective effect against PDM .All factors in the univariate analysis with le model , overwei Brevibacterium flavum and a rare Gram-positive bacterium, Bacillus cereus, with 16S rRNA gene sequencing. There was no anaerobic bacterial growth from any sample.To investigate the role of bacterial infection in the development of PDM, multiple microorganism detection method was employed in this study. Acid-fast bacilli staining was negative in both case and control groups. However, colony formation was found on only one pus specimen plate in bacterial culturing study and two distinct bacterial strains were further identified as Pseudomonas spp. (29.03%) , followed by single bacterial infection of(29.03%) . Enterocγ, IL-12A, and IL-17A, were involved in the disease progression with an IHC method  . HoweverObesity is related to low-grade chronic inflammation, which can disrupt immune function , 28. FurThe relationship between reproductive factors and PDM is very interesting. Inconsistent with the finding reported previously from literature in which parity was not found to be associated with the risk for duct ectasia , our resThe relationship between nipple retraction and PDM has been well-documented . Nipple  S. aureus was the predominant pathogen, followed by Bacteroides spp., Anaerobic strep., and P. aeruginosa. However, the relationship between bacteria and PDM was not confirmed in other report [ S. aureus was the most common bacterium in PDM or anaerobic bacteria was associated with this disease based on our current data. Interestingly, the bacteria identified here were those microorganisms which are often related to community-acquired infection, including Pseudomonas spp. and S. aureus. Collectively, these finding suggests that bacterial infection may only be acting as an induction factor and the immune system which are following the bacterial infection might play a major role in the development of PDM. However, more study is needed for the future to determine the common route in which the bacterial infection and the following immune response converge.Bacterial infection is considered to be another possible etiology of PDM, although it has been a long-standing controversial problem. Al Benwan et al.  found thr report . Here, wr report , we did γ and IL-12A are characteristic Th1 cytokines [γ and IL-12A were upregulated in breast stromal inflammatory cells of PDM patients. Recently, a lineage of CD4+ T cells producing IL-17A was described and accordingly named as Th17 cells. IL-17A has been shown to be important for host defense against pathogens [γ and IL-12A were detected in PDM tissue samples, there is a rationale to conclude that the Th1 immune response plays a role in this disease. The mechanism by which these cytokines contribute to the course of disease is still unknown and needs further investigation.IFN-ytokines . Th1 celytokines . In thisathogens . Howeverγ and IL-12A participated in the course of the disease.The strengths of the present study include the following: (a) a relatively large population of patients was included; (b) the assessment of tissue was done by two histopathologists experienced in breast inflammation. However, our study also had several limitations. First, this was a nonmatched case control study and there might be some uncontrolled biases which may influence the credibility of the results. Second, there was an issue with missing data, especially the data of reproductive factors, which may also lead to the bias of results. Finally, we have not investigated the mechanism by which IFN-γ and IL-12A, were also shown to be associated with PDM, which indicates that the Th1 immune response may closely relate to this disease. Therefore, PDM could be an inflammatory disease related to bacterial infection and consequent immune responses with increased cytokines. As far as we know, this is first report about the possible immune responses following bacterial infection which is involved in development of PDM disease, and more research is needed for the future to further investigate the pathogenesis of PDM on the mechanism by which bacterial infection and consequent immune responses affect the progression of the disease.The results of the present study indicated that overweight/obesity and later onset of menarche are independent risk factors for PDM. In contrast, age of first birth >27 years is a protective factor for PDM. The importance of these reproductive factors to the risk of PDM has not been previously reported and will provide important novel information to broaden the avenue of investigating the etiology of PDM in the future. We also confirm that bacterial infection with a mixture of different bacterial species is a closely associated with the pathogenesis of PDM. Moreover, proinflammatory cytokines, IFN-


Thrombocytopenia affects 25–55% of intensive care unit (ICU) patients . The reaEscherichia coli (STEC)-associated HUS have estimated incidences of 6 and up to 29 cases per million, respectively, and atypical HUS (aHUS) a prevalence of 0.2–0.4 cases per million [DIC is relatively common, developing in 9–19% of ICU patients, usually as a result of sepsis , with an million , making  million . Accordi million . These cSeveral diagnostic algorithms for TMA have been published , 8–10. HGiven the importance of differentiating DIC from TTP and HUS, we propose a concise algorithm based on existing guidance , 9, 11 aOnce DIC has been excluded, confirming the cause of the TMA is paramount for appropriate management. The two most concerning causes of TMA are TTP and HUS. TTP is caused by a deficiency in a disintegrin-like metalloproteinase with thrombospondin motif type 1 member 13 (ADAMTS13) and has 90% mortality without plasma exchange . HUS is Like any pragmatic guidelines, we chose to focus on the most common presentation as we considered this of most benefit. For comprehensive guidance on TMA diagnosis and management, we refer to other works, such as those of Scully et al. , CampistIt should also be noted that some of the tests required in the differential diagnosis  are not available at all institutions. If rapid ADAMTS13 testing is not possible, the PLASMIC score, a seven-component prediction tool that can accurately and reliably predict the probability of severe ADAMTS13 deficiency , can be Critically ill patients have a range of clinical problems, including multi-organ failure, sepsis, and shock , and ear


Hydroxyurea is a well-established inhibitor of ribonucleotide reductase that has a long history of scientific interest and clinical use for the treatment of neoplastic and non-neoplastic diseases. It is currently the staple drug for the management of sickle cell anemia and chronic myeloproliferative disorders. Due to its reversible inhibitory effect on DNA replication in various organisms, hydroxyurea is also commonly used in laboratories for cell cycle synchronization or generating replication stress. However, incubation with high concentrations or prolonged treatment with low doses of hydroxyurea can result in cell death and the DNA damage generated at arrested replication forks is generally believed to be the direct cause. Recent studies in multiple model organisms have shown that oxidative stress and several other mechanisms may contribute to the majority of the cytotoxic effect of hydroxyurea. This review aims to summarize the progress in our understanding of the cell-killing mechanisms of hydroxyurea, which may provide new insights towards the improvement of chemotherapies that employ this agent. Hydroxyurea , that binds to RNR in the G1 phase [Because RNR catalyzes the rate-limiting step in the biosynthesis of all four precursors for DNA replication, its activity is tightly regulated during the cell cycle, which generates a periodic fluctuation of the dNTP concentration in proliferating cells. As mentioned above, the enzyme’s allosteric specificity regulation controls the balanced concentrations of dNTPs. In mammalian cells, RNR activity in the G0/G1 phase is suppressed by transcriptional repression of the R2 gene and by anaphase-promoting complex Cdh1-dependent degradation of the R2 subunit in the M phase ,58. The G1 phase ,63. The G1 phase ,65,66,67G1 phase ,69.In the presence of HU, proliferating cells are arrested in S phase due to the decreased levels of dNTPs, which slows the DNA polymerase movement at replication forks. In eukaryotes, slowed forks activate the replication checkpoint, a highly conserved intracellular signaling pathway that is crucial for the maintenance of genome stability under replication stress ,71. The E. coli. Earlier studies showed that the cytotoxic effect of HU in both mammalian cells and E. coli appears to be linked to the accumulation of DNA strand breaks in HU-treated cells [2O2 and nitric oxide [S. pombe are highly sensitive to HU (see below) and the genetic backgrounds of the cell lines used in the earlier studies are unknown, the linkage between the DNA damage and the cell killing effect of HU may need to be reconsidered with caution.Due to the reversible S phase arrest, HU is generally considered to be cytostatic, particularly to non-cycling cells ,78,79,80ed cells ,92 or caed cells ,94,95,96ic oxide . HoweverROS is a collective term used to describe ions and free radicals containing derivatives of molecular oxygen that are more reactive than oxygen itself. The ROS formed inside living cells commonly includes superoxide anion, hydrogen peroxide, and hydroxyl radical . The norE. coli using systems-level analyses have revealed the genomic and physiological effects of HU treatment that lead to cell death [E. coli [2O2 alkyl hydroperoxide reductase [To explain the mechanisms of rapid cell killing in the S phase and the teratogenic effect of HU, DeSesso hypothesized in 1979 that HU may exert its cytotoxic effects through radical chain reactions initiated by its hydroxylamine group, and predicted that antioxidants should ameliorate the cytotoxic and teratogenic effects of HU . Subsequll death . It was ll death . Productll death . An accu[E. coli ,98. Conseductase , enhanceeductase . Intereseductase ,105.S. cerevisiae and S. pombe are relatively insensitive to HU. However, recent studies suggest that HU treatment may generate ROS in both species. In addition to the DNA damage and environmental stress responses, HU treatment activates the Yap and Aft regulons in S. cerevisiae that function in redox and iron homeostasis respectively [S. cerevisiae [S. pombe is Pap1. Similar to that in S. cerevisiae, depletion of Pap1 also moderately sensitized S. pombe to HU [S. pombe to various other agents such as staurosporine [Wild type yeasts such as ectively ,106,107.revisiae . The Yapbe to HU , suggestosporine , caffeinosporine , and berosporine  and to Dosporine . Scavengosporine , which dosporine . The actosporine ,115. Conosporine . Apd1 isosporine .S. cerevisiae to chronic treatment with HU [S. cerevisiae, E. coli cells devoid of YfaE protein, which contains an iron–sulfur cluster and is required for the diferric tyrosyl radical cofactor maintenance of RNR, are also sensitive to HU [The ROS generated by HU treatment can also alter the functions of proteins that contain iron–sulfur centers. For example, Dre2-Tah18 protein complex functions in cytosolic iron–sulfur protein biogenesis ,117 and  with HU ,120 and  with HU . Similarve to HU .Together, these studies show that HU may kill the cells by affecting the iron–sulfur clusters in proteins that function in the maintenance of the diferric tyrosyl radical center in RNRs or other cellular processes. Without a proper maintenance of diferric tyrosyl radical center in RNRs, the radical may leak into the cytoplasm and generate superoxide . InteresS. pombe that, similar to a previous study [hem13 mutant, in which the heme level is low due to the hypomorphic mutation of the enzyme coproporphyrinogen III oxidase in the heme biosynthesis pathway, the levels of ROS as well as protein carbonylation, an indicator of the oxidation of various macromolecules, were significantly increased in HU-treated cells (our unpublished data). Unlike the checkpoint mutants that usually die within 2 to 3 h or one cell cycle time in HU, the hem13 mutant is highly sensitive only to chronic treatment of HU. Similar chronic HU sensitivity was also observed in S. cerevisiae lacking Sod1, the enzyme that catalyzes the decomposition of superoxide [hem13 mutant can be suppressed by culturing the cells under anaerobic conditions, which inhibits aerobiosis and thus decreases the production of endogenous ROS. Like the S. cerevisiae cells lacking Sod1, increased RNR activity cannot rescue the HU sensitivity of the hem13 mutant, which is consistent with the notion that the hem13 mutant is killed by a mechanism that is unrelated to dNTP depletion.We have recently found in us study , the levperoxide , which sS. pombe that are sensitive to replication stress, we identified a new hypomorphic mutation, erg11-1, that dramatically sensitizes the cells to chronic, but not acute treatment with HU [erg11 is the enzyme sterol-14α-demethylase, which is required for ergosterol biosynthesis and a major target of antifungal agents. We found that, unlike wild type cells that are arrested in S phase, HU arrests the mutant cells mainly in cytokinesis. The HU-induced cytokinesis arrest is relatively stable and occurs at low doses of HU, which likely explains the remarkable HU sensitivity. HU hypersensitivity has also been observed in several erg mutants in S. cerevisiae, including erg10-1, which encodes the first enzyme in the ergosterol biosynthesis pathway acetolacetyl-CoA thiolase [erg3 that encodes C-5 sterol desaturase [S. cerevisiae mutants remains to be determined, HU may suppress cell division in the presence of sterol deficiency in diverse eukaryotic organisms.While screening for new mutants in  with HU   involved in cell division that can be unmasked by sterol deficiency and become druggable to HU  of HU [Since the cytokinesis arrest occurs at HU concentrations much lower (1–3 mM) than that required for replication arrest (more than 6 mM) in le to HU . As ments arrest . Genetics) of HU ,134,135.s) of HU ,137. IdeS. pombe erg11-1 mutant is clearly caused by a previously unknown mechanism [This review has focused upon the mechanisms by which HU exerts its cytotoxic effects. Clearly our knowledge is far from complete. For example, how are ROS generated in HU-treated cells? Fork collapse can clearly generate ROS ,20,22. Hechanism  and is cechanism ,53,54,55echanism ,56.


The term gossypiboma refers to a sponge that has been forgotten in the surgical field. It is the most common retained surgical item, and constitutes a continuing problem for surgical safety. We performed a hospital-based study to examine their incidence, root cause, and outcomes, as an effort toward improving prevention.This retrospective study covered 10 years (2006-2015) and included surgically confirmed cases of abdominal gossypibomas occurring after 45,011 abdominal and gynaecological operations in 2 public hospitals in Lome (Togo). Age, diagnosis, initial surgical procedure, evidence of textile count, and data related to the revision procedure were collected for descriptive analysis.Fifteen cases of gossypibomas (11 women and 4 men) were recorded. The mean age of the patients was 27 (range 21-55) years. Initial procedures were gynaecological in 11 patients and 5 cases involved an emergency surgery. Evidence of sponge counting was found in 6cases. Gossypiboma was an incidental finding in 1 patient. The average time to onset of symptoms after the initial procedure was 2 months. The gossypiboma was removed within 7 days to 4 years after the initial procedure. Postoperative complications included enterocutaneous fistula in 2 patients, incisional hernia in 2 patients, and wound sepsis in 1 patient. Death occurred in 2 patients (13.3%).Although rare, the incidence of gossypibomas is still unacceptably high and reveals failures regarding patient safety standards. The associated morbidity and mortality are significant, yet can be reduced by an early diagnosis in the immediate postoperative period. A systematic methodical count of sponges is the cornerstone of prevention, and introducing surgical safety protocols, such as the WHO Safe Surgery Saves Lives checklist, can enhance effectiveness. There is a crucial need for safety-focused policies, which may include a never event reporting system, elaboration of prevention strategies, interventions, and evaluation. The term gossypiboma refers to a sponge that has been forgotten in the surgical field . “GossypThis retrospective study covered 10 years  and included surgically confirmed cases of abdominal gossypibomas in the general/visceral surgery and gynaecological departments of two hospitals in Lome, Togo, West Africa . These two hospitals are the only surgical public facilities serving the urban demographic area of Lome.During the study period, 45,011 abdominal and pelvic procedures  were performed, including those in the gynaecological departments. At the time of the study, the two hospitals did not have an institutional policy regarding quality improvement, especially in the surgical safety area, and the WHO surgical checklist, or any surgical safety checklist, was not implemented. Furthermore, there were no institution-level specific protocols to reduce the incidence of never events, including RSIs. Although a sponge count was recommended, no standardized count practice was established. Radiopaque sponges were not always available, and the hospitals did not have specific equipment for an intra-operative X-ray search of a forgotten sponge.Using the operative registries, summaries of all gynaecological and abdominal procedures during the study period, including name, date, operative diagnosis, and procedure, were manually reviewed, as there was no coding system, or a specific reporting procedure. Reviewers were asked to select, for a thorough screening of the surgery reports, patients with the following terms indicated in the diagnosis box: foreign body, gossypiboma, and swab. Additionally, the surgeons and gynaecologists at the 2 hospitals, as well as others involved in the identified cases, were interviewed in order to complete data collection from the patient’s records. The data were reported on a form created in EPI Info 7 for descriptive analysis.And concerned the following parameters: age, diagnosis, initial surgical procedure, evidence of textile count, and data related to the revision procedure .Fifteen cases of gossypibomas  were recorded in the two hospitals. The overall incidence was 1/3030 procedures, with an incidence of 1/2656 (4 cases) and 1/3947 (11 cases) at the Lome Commune regional hospital and Sylvanus Olympio teaching hospital, respectively. The mean age of the patients was 27  years. Table Gossypiboma was an incidental finding in 1 patient during an elective re-operation. Signs presented by other patients are shown in Table The median date of detection (by surgery or preoperative workup) was 9 months after surgery . The gossypiboma was removed via a midline laparotomy in 14 patients and an inguinal incision in 1 patient. The removal was performed in 8 patients within the first year after the initial procedure. The gossypiboma was a small sponge (10×10cm) in 3 cases and a large laparotomy sponge (30×30cm) in 12 cases. The macroscopic appearance of the gossypiboma corresponded to a pseudo-tumour fibrous reaction in 3cases Fig. . There wGossypibomas are the most common RSI , 3. TheiRisk factors of gossypibomas that have been reported in the literature include emergency surgery, high body mass index, unplanned change in the type of surgery, and a complex surgical procedure , 3, 4. ANearly half of the gossypibomas were found more than a year after the initial operation in our study. Although some patients may remain totally asymptomatic for a long time , it is aThe surgical treatment of gossypibomas is unequivocal , 10, 19 Although rare, the incidence of gossypibomas is still unacceptably high regarding patient safety standards toward never events. They are evidence of failures in preserving the quality of care and safety around surgical cares in the present low-income setting. Morbidity and mortality associated with gossypibomas are high, yet can be significantly reduced by an early diagnosis in the immediate postoperative period. A systematic methodical sponge count is the cornerstone of prevention. Introducing surgical safety protocols, such as the WHO checklist, can be a part of focused policies to improve global surgical safety and prevent never events, including gossypibomas. A reporting system of these medical errors remains necessary to provide valuable insights for the elaboration of prevention strategies, interventions, and evaluation.


Background. Adalimumab is effective for the maintenance of remission in patients with moderate-to-severe ulcerative colitis (UC). Currently, biologic therapies are used in cases where patients fail conventional medical therapies. If biologic therapies are not available, patients often choose to remain in an unwell state rather than undergo colectomy. Objective. The aim of the study was to evaluate the cost-effectiveness of adalimumab in patients with UC where adalimumab was readily available compared to not available. Methods. A previously validated Markov model was used to simulate disease progression of patients with UC who are corticosteroid-dependent and/or did not respond to thiopurine therapy. Utility scores and transition probabilities between health states were determined by using data from randomized controlled trials and real-life observational studies. Costs were obtained from the Ontario Case Costing Initiative and the Alberta Health Schedule of Medical Benefits. Results. The incremental cost-effectiveness ratios for readily available adalimumab treatment of UC were $40,000 and $59,000 per quality-adjusted life year, compared with ongoing medical therapy in an unwell state, at 5-year and 10-year treatment time horizons, respectively. Conclusion. Considering real-life patient preferences to avoid colectomy, adalimumab is cost-effective according to a willingness-to-pay threshold of $80,000 for treatment of UC. Ulcerative colitis (UC) is a chronic, relapsing, and remitting inflammatory disease that is characterized by symptoms of diarrhea, rectal bleeding, urgency, and abdominal pain . Canada Due to the relapsing-remitting course of UC, treatment is focused on the induction and maintenance of clinical remission and endoscopic mucosal healing . MaintaiFor patients who fail to maintain remission on the above therapy, the options for treatment are limited to continuous corticosteroid use, colectomy, or biologic therapy. The chronic use of corticosteroids is associated with significant adverse effects and can leave the patient in a chronically unwell state . While cα), adalimumab, has been shown to be well tolerated and effective in inducing remission in patients with moderate-to-moderately severe active UC [The use of the antitumor necrosis factor alpha  over a 1-year time period [₤7190/QALY for 1 year of treatment [The cost of biologic therapy is significant, varying from $18,000 to $33,000 per patient per year . While te period . Similarreatment . In contreatment . The conIn this analysis, the costs and utility of patients receiving readily available adalimumab treatment for ulcerative colitis was compared to that where adalimumab was not readily available and thus the patient preference for a chronically unwell state, with or without corticosteroids, rather than immediate colectomy dominated –18.Our study replicated a previously validated Markov model in ulcerative colitis that was conducted by our center for another anti-TNF agent, infliximab, that calculates the difference in costs divided by the differences in utility between the study option and alternative intervention, the result being the incremental cost-effectiveness ratio (ICER) . In the A base-case analysis was used that consisted of a theoretical cohort of patients with moderate-to-severe active UC who are corticosteroid-dependent and either failed or are intolerant to thiopurine treatment. Approximately 60% were male, with an average age of 40 years old.A previous established Markov model was used to determine the ICER of two management strategies: (1) no adalimumab, which includes scenarios where adalimumab was not available and patients therefore remained in a chronically unwell state in order to avoid colectomy, and (2) adalimumab therapy, where adalimumab was readily available to induce and maintain clinical response. Patients in this group were modeled as being treated initially with 160 mg, 80 mg at week 2, followed by 40 mg every other week. The Markov model structure is displayed in The different health states used in the model were defined and verified by a panel of gastroenterologists and gastrointestinal surgeons with expertise and experience in the treatment of inflammatory bowel disease . Based oThe probabilities of moving on to subsequent health states were as follows: patients who received adalimumab therapy either experienced an induction response or became “nonresponders.” The patients who responded to adalimumab may continue to respond to treatment over time or they may experience a secondary loss of response. The patients who did not respond to the initial adalimumab treatment or who lost response to treatment returned to ongoing steroid therapy, where a portion of patients eventually underwent colectomy. Patients who experienced an adverse effect due to adalimumab therapy could sometimes be successfully treated for the complication. If they could not be treated for the complication, they were either taken off adalimumab and returned to ongoing steroid therapy or offered a colectomy. Patients who received a colectomy could develop complications associated with the surgery or could remain in a response state. If a patient develops chronic pouchitis due to surgery, they can be subsequently treated with adalimumab or return to steroid treatment. The possibility of patient mortality was considered for each health state.Our analysis follows the 2006 economic evaluation guidelines as set out by the Canadian Agency for Drugs and Health Technologies [The probabilities of patients moving between health states were derived from a review of the literature from both randomized controlled trials and real-life studies. Study results were weighted based on sample size. Loss of response to adalimumab was obtained from our center data in an attempt to replicate real-life clinical response. The weighted probabilities were then reviewed by the panel of gastroenterologists for face validity. Tables A literature search was administered to assess the costs of each health state per 3-month cycle. To estimate resource use, we included physician, hospital, and outpatient drug costs. Physicians' fees were obtained from the Alberta provincial fee schedules of Alberta Health and Wellness . HospitaIn order to calculate quality-adjusted life years (QALYs) for each treatment regimen, utilities for each health state were determined through a review of the literature and access to expert opinion. The utility value assigned to each health state is outlined in Tables u), and a gamma distribution was used.One-way sensitivity analyses with tornado diagrams (available on request) were conducted on all key parameters for 6 scenarios with a time horizon of 5, 10, or 15 years, and with a utility score of the response-to-adalimumab health state as 0.79 or 0.82. The probabilities and utility scores were varied between the lower and upper ends of 95% confidence interval, and the cost of each health state was varied by 25%, as shown in Tables As shown in In a retrospective cohort study from our expert IBD center, it was determined that dose escalation was required in 50% of UC patients after a mean time span of 59.3 (±70.5) weeks . Dose esThe significant amount of patients needing dose escalation and the high costs of dose escalation results in many centers attempting dose de-escalation. To determine the cost-effectiveness of dose de-escalation, we assumed that dose de-escalation was attempted in 54% of patients and was successful in 63% of those patients, based on data obtained by Baert et al. . When deCosts and utility scores were discounted annually at the rate of 5%.With a utility score of 0.79, the cost-utility analysis yielded a cost of $97,000 with 3.154 QALYs for a patient during a 10-year period.When adalimumab is readily available and patients opt for adalimumab treatment to induce and maintain response, the cost-utility analysis with a utility score of 0.79 yielded a cost of $107,000 with 3.321 QALYs for a patient during a 10-year period.The ICER at 10 years, when comparing readily available adalimumab treatment to ongoing medical therapy in an unwell state, was $59,000 per QALY gained when using a utility score of 0.79 measured by time trade-off and $53,000 per QALY gained when using a utility score of 0.82 measured by visual rating scale .Sensitivity analysis of probabilities, costs, and utility scores showed that the ICER varied from $37,000 to $81,000 (utility score 0.79) or from $33,000 to $72,000 (utility score 0.82) at the 10-year horizon . The mosSensitivity analyses were also performed with varying time horizons. The ICER of adalimumab therapy versus no adalimumab therapy ranged from $25,000 to $65,000 (utility score of 0.79) and from $22,000 to $58,000 (utility score of 0.82) at a 5-year horizon. At a 15-year horizon, the ICER ranged from $45,000 to $91,000 (utility score of 0.79) and from $40,000 to $81,000 (utility score of 0.82).Upon analyzing dose escalation response rates using Crohn's disease data, we estimated dose escalation ICERs to be $85,000 at 5 years, $102,000 at 10 years, and $113,000 at 15 years when using a utility score of 0.79. Analysis of dose escalation with a utility score of 0.82 revealed ICERs of $77,000 at 5 years, $92,000 at 10 years, and $102,000 at 15 years.The ICERs associated with dose de-escalation were $75,000 at 5 years, $93,000 at 10 years, and $105,000 at 15 years when applying a utility score of 0.79 to the response-to-adalimumab health state. Analysis of dose de-escalation with a utility score of 0.82 revealed ICERs of $63,000, $84,000, and $95,000, at 5, 10, and 15 years, respectively.Cost-effectiveness acceptability curves are presented in Currently, two multicenter, randomized, placebo-controlled trials have shown adalimumab to be well tolerated and effective in inducing and maintaining remission in patients with moderate-to-severe UC , 9, in aTo date, there is no willingness-to-pay threshold that is accepted universally throughout healthcare systems. A threshold value of $50,000 per QALY gained has been widely used in many studies and countries as a reference threshold since the 1970s, although its use is often debated as being too low. Grosse argued that the $50,000 per QALY threshold is an arbitrary decision rule that lacks theoretical and empirical justification and is outdated due to the failure to adjust the value for inflation or changing levels of income or healthcare budgets since its introduction . Also, iBased on the $50,000 willingness-to-pay threshold, it appears that adalimumab therapy is cost-effective compared to ongoing less effective medical therapies at a 5-year time horizon. Although the ICERs for 10-year and 15-year time horizons surpass that threshold, they are all considered to be cost-effective according to a willingness-to-pay threshold of $80,000 per QALY gained as per Rocchi et al. . These rBy using the previously validated Markov model validated for infliximab  as the mThe lower ICER for adalimumab compared to infliximab is likely due to the lower cost of adalimumab in Canada compared to infliximab per 3-month cycle. In addition, this difference may be larger than expected due to the lack of the infliximab model taking into consideration the indirect costs of infliximab administration. Infliximab administration requires patients to receive intravenous injections at an outpatient health center, opposed to adalimumab which can be administered by the patients subcutaneously. Examining the cost-effectiveness of adalimumab and infliximab in separate models is a limitation of our study, as in real-life practice, patients are able to switch between these two agents.An exploratory analysis of ICERs associated with dose escalation was conducted due to the high rate of patients who undergo dose escalation as a result of secondary loss of response. The average ICER associated with dose escalation was $85,000, $102,000, and $113,000 at 5, 10, and 15 years, respectively. This data clearly demonstrates that a need for dose escalation in patients who experience a loss of response to adalimumab results in additional costs, thus increasing the ICER above frequently used willingness-to-pay thresholds. Furthermore, dose de-escalation is common in patients who regain response after being dose escalated. Our analysis revealed ICERS of $75,000 at 5 years, $93,000 at 10 years, and $105,000 at 15 years, indicating that dose de-escalation has the potential to reduce costs after patients have regained response to adalimumab upon dose escalation.An important limitation of our study is the lack of our model taking into consideration the use of concomitant therapies, including immunosuppressants and methotrexate. Numerous studies have demonstrated a difference in the efficacy of anti-TNF agents and steroids with the use of combination therapy , 34. ForIn conclusion, this study demonstrates that, using response rates from real-life centers and real-life patient preference to avoid colectomy, readily available adalimumab treatment of ulcerative colitis is cost-effective according to willingness-to-pay thresholds of $80,000 per QALY compared with when adalimumab is not available and the patients elect for a chronic unwell state to avoid colectomy. Dose escalation will increase these costs.


Being considered an exclusive human structure for a long time, fibularis tertius (FT) is believed to have a secondary function of foot dorsiflexion and eversion. This study is an attempt to approach the issue from an anatomical perspective. A systematic literature search identified 35 studies (7601 legs) which met the inclusion criteria. The weighted results of FT presence were as follows: an “adult cadaveric” frequency of 93.2% and a clinical frequency of 80%. The most common FT origin and insertion sites were the distal half of fibula and the base of the 5th metatarsal, respectively. In 95% of cases, an accessory fibular muscle was detected when FT was lacking. We demonstrated that the discrepancy found between the adult cadaveric and clinical frequency values would point out a probable bias in interpreting previous kinesiological results. On an evolutionary level, comparative anatomy demonstrated a very low FT prevalence among monkeys while reaching a frequency of 30% in gorillas, the only non-human apes having an almost exclusive terrestrial locomotion. The consistent prevalence among humans and the presence of similar functional muscles when it is missing would support an essential role of FT during the phylogenetic development of the erect bipedal posture and probably during gait.  It is most likely that the first description of the PT muscle has been reported by Vesalius . The Digital Collections of the National Library of Medicine, All anatomical studies were eligible for inclusion, be it cadaveric, clinical, or radiological, with no restriction to the age of the sample. The primary outcomes were set to be the true or crude prevalence of FT. The true prevalence rate is defined as the number of legs affected compared to the number of legs available for study. The crude FT prevalence is the number of individuals who have either one or two FT compared to the number of individuals available for study. Secondary outcomes were defined as the ancestry-based, laterality-based, gender-based, and side-based frequencies, size of FT, and its origin and insertion variants. Data Extraction and Analysis. Data extracted included sample size, baseline demographic characteristics of the sample, and prevalence values. Size, origin, and insertion of PT were also extracted when available. Analysis was performed using StatsDirect v2.7.8 . Proportion meta-analysis was used to (a) calculate the overall true/crude pooled prevalence estimate (PPE), (b) perform subgroup analysis for side-based, gender-based, and ancestry-based PPEs. Odds ratio (OR) meta-analysis was used to establish potential associations with variables such as sex and side. Heterogeneity was investigated using the I2 statistic; whenever I2 > 50%, the random-effect estimate was reported. In case of subgroup analysis of five studies or less, the fixed-effect estimate was used regardless of I2 value. Sensitivity analyses were performed in two instances, when applicable: a subgroup analysis of studies having samples of more than 100 specimens and another two subgroup analyses of studies published before and after the year 1920.The electronic databases search yielded 164 hits. Fourteen duplicates were removed. Thirty-five studies were considered potentially relevant and twenty met the inclusion criteria. Reasons for exclusion were as follows: 9 reports of anatomical variation, 3 electrophysiological studies, and 3 traumatic clinical case reports. The search of old manuscripts revealed 14 relevant studies where 10 met inclusion criteria. Reference checking of the 30 studies meeting inclusion criteria yielded another 5 studies . In totaI2 = 62.2%). A sensitivity analysis of 9 studies including samples of more than 100 leg specimens with a total of 2517 specimens yielded a weighted frequency of 92.7% . Eight studies published before the year 1920 with a total of 1723 legs yielded a weighted frequency of 93.3% . Seventeen studies published after the year 1920 with a total of 1805 legs yielded a weighted frequency of 93.7% .Twenty-five studies , 37–52 iI2 = 56.8%).Eight studies , 49, 52 I2 = 68.3, p = 0.6). No significance was found in relation to the side.Thirteen studies , 51, 52 I2 = 0%, and p = 0.03) in favor of male sex.Nine studies , 49, 52 I2 = 0%).Thirteen “Caucasian” studies –47, 49 iI2 = 82.4%).Four Indian studies , 50–52 iI2 = 67.5%).Three South American studies , 32, 48 I2 = 36.5%).Two Japanese studies , 41 inclOne Turkish study , one AfrI2 = 0%).Four studies –35, 53 wI2 = 98.6%).Six studies , 54–57 wI2 = 97.5%).All clinical studies but Bourdon and Petitdant  with a tEleven studies , 47–52 rAll studies but that of Bhatt et al.  reportedSix studies –50, 52 wEleven studies , 51, 52 The study of Johnson  reportedJoshi et al.  reportedThis meta-analysis demonstrated that FT is highly prevalent in humans (93%). This value varied insignificantly when compared to those of the two subgroup sensitivity analyses: the one including samples of more than 100 specimens and the other comparing it to studies published before 1920. The weighted frequency of 93% is found to be higher than that reported in some reviews and textbooks. For instance, low absence frequencies were reported by Kimura and Takahashi  and WillInterestingly, cadaveric dissection revealed a higher frequency (93%) when compared to that obtained with clinical examination (80%). Thus, it seems that around 13% of FT could be missed by routine clinical tests. However, it has been demonstrated that FT tendon and/or FT belly are sometimes difficult to separate from the extensor digitorum longus . That co and 4th metatarsals as possible sites of insertion.On the other hand, origin and insertion sites as found in this study were not similar to those reported in many anatomy textbooks. The majority of FT in this study were found to originate from the distal half, rather from the usually reported distal third or fourth of the fibula , 66. MorInterestingly, this study showed that, in more than 95% of cases where FT is missing, an accessory fibular muscle or a tendinous slip from EDL was present. We believe that the findings of Johnson  of a conAdditionally, many of the included studies described FT insertion as broad and fan-shaped; Krammer et al.  compared Strepsirrhini and New World monkeys as stated earlier. Two single instances in Old World monkeys were reported in the pig tailed baboon (Papio ursinus) [ (Macaca sinica) [ (Macaca fascicularis). In great apes, FT has been found with higher frequencies. Loth [ chimpanzees. Kimura and Takahashi [ gorillas. It is very likely that the presence of PT is associated with the bipedal type of locomotion. The prevalent presence of this muscle in modern humans and gorillas, exclusively and mostly terrestrial, respectively, would point out an evolutionary acquisition related to “bipedalism” [ chimpanzees where their lesser presence is likely due to the fact that those apes are not exclusively terrestrial: being both, arboreal and terrestrial. In fact, foot inversion due to dorsiflexion is not counteracted by any muscle in arboreal apes [On an evolutionary level, FT is not found inursinus)  and in t sinica) . Additio sinica)  observedes. Loth  reportedakahashi  completeakahashi  and concedalism” . The sameal apes . When coeal apes  ascribedeal apes  labeled eal apes , 77. ThuThe evolutionary, functional, and morphological interpretations made in the light of our results concur to conclude that FT is an important functional anatomical structure. Its high frequency, the presence of other similar muscles when absent, and its broad insertion on the lateral aspect of the mid-foot would confer to this muscle an essential role in human bipedalism. By using quantitative evidence synthesis, the findings of this meta-analysis coupled with FT frequencies found in primates indicate that FT function would have had an essential role during the phylogenetic development of the erect bipedal posture, a statement which is in line with that made by Krammer et al. . These aThe interpretation of the meta-analytical and comparative results would point out the fact that such a recent and constant muscle might have an important function, or a fine-tuning function, related to human motion, be it during walking and/or during faster paces of motion such as running. Due to our “important gait muscle” argument and to the lack of reported individual analysis in human gait, further research is warranted to assess FT contribution during the different types of human locomotion.


HTRA1) gene in the pro-band, which may be a pathogenic factor according to the biological software analysis. The following SNP results revealed that the other family members also had the HTRA1 gene mutation. Thus, the CADASIL-like family disease may be caused by heterozygous HTRA1 gene mutation, which leads to autosomal dominant hereditary cerebral small vessel disease.The aim of this study was to find related pathogenic genes in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy in (CADASIL)-like patients. The direct sequencing and high-throughput multiplex polymerase chain reaction (PCR) was performed to screen for related genes. The clinical and imaging data of a CADASIL-like patient (the pro-band) and his family members were collected. At first, the known hereditary cerebral vascular genes of the pro-band were screened with direct sequencing to find candidate gene mutations. High-throughput multiplex PCR was then used to analyze the single nucleotide polymorphism of the candidate gene in the family members. The results showed that there was missense mutation of the high temperature requirement protease A1 ( Cerebral small vessel disease (CSVD) is a set of heterogeneous diseases due to the pathological changes of cerebral arterioles, venules and capillaries –3. StroHTRA1 allele mutations, which is also called HTRA1 homozygous mutations  stroke and progressive cognitive impairment as the major clinical characteristics; 2) negative Notch3. Stroke is an ischemic or infarct injury of the brain that results in neurological deficit and contralateral impairments in verbal, cognition, motor and sensory system. Stroke and cognitive impairment are inter-related.HTRA1, APP, CST3, and ITM2B, etc., and SNP genotyping assay were then conducted on family members after identifying the candidate gene.Blood samples of the pro-band were collected and used for screening the hereditary cerebrovascular disease related genes, such as NOTCH3, HTRA1, KRIT1 (CCM1), CCM2, PDCD10 (CCM3), GLA, HBB, CBS, MMADHC, MTHFR, MTR, MTRR, PRKAR1A, F5, APP, CST3, ITM2B, ENG, ACVRL1, SMAD4, ABCD1, ARSA PLP1, MLC1, ASPA, GFAP, and GALC.The genetic sequencing of the 27 related genes of hereditary cerebrovascular disease were conducted. The 27 genes included To extract DNA, EDTA was added to venous blood (2 mL) for anticoagulation, and the column method  was used for extracting genomic DNA from peripheral blood leukocytes. DNA was stored at 4°C.HTRA1 gene exon coding region, using 2 X PCR MasterMix polymerase (Tiangen) for PCR amplification . The PCR products from the specific primers were direct sequenced by Taiyuan Jinyu Clinical Inspection Ltd. using ABI3500 . Then, reference sequences (NM_002775.4 and NG_011554.1) were used for comparison to localize the possible gene mutation.Gene sequences were obtained from the human genome database GenBank. The Primer Premier 5.0 software was applied to design primers of the Sequencing results were analyzed by PolyPhen2 , SIFT (sorting intolerant from tolerant) and Mutation Taster to see whether there is a likelihood of pathogenesis by gene mutation. SIFT predicts whether an amino acid substitution can affect protein function and prediction is based on the degree of conservation of amino acid residues in sequence alignments derived from closely related sequences, collected through PSI-BLAST.Based on the pedigree chart of the pro-band , there wThe pro-band, with onset of the disease at the age of 35 years, had no hypertension, diabetes, smoking or other risk factors of cerebrovascular disease, and there was no family history of cerebrovascular disease. In 2006, he presented with dizziness and double lower extremity weakness. In 2008, 2009, and 2011, he presented with similar symptoms, and double lower extremity weakness appeared alternately. After cerebral infarction treatment, the symptoms were slightly reduced. In 2012, he presented with dizziness and double lower limb weakness that were more serious, and required hospitalization. There was no definite risk factor for cerebrovascular disease. On physical examination, findings were clear mind, slower verbal reaction, slightly dull, the tongue to the right, limb muscle strength of grade 5, hypermyotonia, hyperreflexia, tendon hyperreflexia, bilateral symmetry for sense of depth, and bilateral Babinski syndrome (+). In cranial MRI, findings were multiple lacunar infarction in bilateral basal ganglia, lateral ventricles, and semi-oval center, and changes of bilateral periventricular white matter demyelination.III6 was the sister of the pro-band, born in May 1955. In 2003, she was diagnosed with cerebral infarction for the first time, and it recurred once a year on average. In 2012, she presented with recurrent right limb numbness and weakness, and slurred speech. Symptoms aggravated in 4 days, and she required hospitalization. On physical examination, she was found to be clear-minded, with slurred speech, shallow right nasolabial fold, tongue to the right, positive right limb paresis test, and right Babinski syndrome (+). The imaging findings were similar to the pro-band.Head MRI imaging data -4 suggeThe pro-band score was 17 on the MMSE, which was low considering that he was a graduate of technical school , indicating moderate damage of neurologic function. The III6 score was 22, considered low as she had a bachelor degree, indicating slight damage of neurologic function. The III9 score was 28, and his educational level was bachelor degree, indicating he was normal.HTRA1, i.e., HTRA1 (NM_002775.4) Exon4 c.905G>A p. (Arg302Gln). The biological software analysis with PolyPhen-2, SIFT, and Mutation Taster suggested that this missense mutation very likely caused cerebrovascular disease. Subsequent SNP genotyping assay of the candidate gene in III6 and III9 indicated that the HTRA1 gene had the same single nucleotide polymorphism, strongly suggesting that this heterozygous missense mutation is the reason for family disease  encoding a series of serine protease and its encoding product is widely expressed in vivo  by cleaving the IGR binding protein. In addition, HTRA1 was reported to regulate cell proliferation activity (The  in vivo . This practivity .HTRA1 allele, whereas heterozygous mutations are not pathogenic (It is commonly recognized that the CARASIL disease is caused by the mutation of the thogenic . Howeverthogenic . The conHTRA1 missense could lead to the arginine to glutamine change for the encoded protein located at 302nd amino acids. The mutation mode leads to a change in the structure of the protein, which may lead to changes in the spatial structure, and then to the change of the protein physical and chemical properties, physiological characteristics etc. Differently than HGMD, ESP6500, and dbSNP databases, this mutation has not been included in databases. Therefore, the SNP of HTRA1 is considered not present in the normal population, but present in the family members who are either diagnosed with cerebrovascular disease or positive in clinical imaging. The clinical imaging of subject III6 indicated the presence of disease but there was no clear clinical symptoms yet, which still needs further follow-ups. This result suggested that the genetic missense mutation was first observed in clinical imaging, and then in clinical manifestations. At the same time, we have found that onset of the disease occurred in younger family members, which complies with genetic characteristics of the early occurrence.The current study found that heterozygous HTRA1 mutation is the cause of the disease, and the effects of the single gene can be enhanced in the genetic process. However, further research is needed to understand the physicochemical properties and biological activity of the encoding protein caused by gene mutation. In addition, it is not clear whether the SNP is present in the normal population. Lastly, we selectively performed partial gene screening of the pro-band rather than the entire sequencing, so whether the gene mutation acts individually or in combination with other types of mutation for the family members is unknown and needs further investigation.In summary, the heterozygous