The broad spectrum of the mechanism of action of immune-boosting natural compounds as well as the complex nature of the food matrices make researching the health benefits of various food products a complicated task. Moreover, many routes are involved in the action of most natural compounds that lead to the inhibition of chronic inflammation, which results in a decrease in the ability to remove a pathogen asymptomatically and is connected to various pathological events, such as cancer. A number of cancers have been associated with inflammatory processes. The current review strives to answer the question of whether plant-derived sulfur compounds could be beneficial in cancer prevention and therapy. This review focuses on the two main sources of natural sulfur compounds: alliaceous and cruciferous vegetables. Through the presentation of scientific data which deal with the study of the chosen compounds in cancer , the discussion of food processing’s influence on immune-boosting food content is presented. Additionally, it is demonstrated that there is still a need to precisely demonstrate the bioavailability of sulfur-containing compounds from various types of functional food, since the inappropriate preparation of vegetables can significantly reduce the content of beneficial sulfur compounds. Additionally, there is an urgent need to carry out more epidemiological studies to reveal the benefits of several natural compounds in cancer prevention and therapy.  Attempts to modulate the immune system started in the mid-1970s . The natBrassica oleracea var. italica), cabbage (Brassica oleracea var. capitata), cauliflower (Brassica oleracea var. botrytis), and other plants of the family brassicaceae are rich sources of bioactive compounds, including vitamin C and E, dietary fiber, and the glycosides of the flavonoids quercetin and kaempferol. However, the components that make cruciferous vegetables so beneficial to human health and set them apart from other plants are the sulfur-containing glycosides . Dietary GSLs are considered as potential anticancer agents [Cruciferous vegetables including broccoli (r agents . The bioAllium cepa) and garlic (Allium sativa) are some of the oldest cultivated plants in the world and are rich in flavonols and natural organosulfur compounds (OSCs). Their consumption was studied, as it was considered beneficial for heart diseases, tumors , and immunological processes, among others, especially due to the biological action of the sulfur-containing metabolic byproducts [Onion -N-hydroximinosulfate group and a variable side chain derived from a variable amino acid R-group . GSLs moO-sulfonate). Upon the cellular injury of plant tissues, GSLs are enzymatically degraded by myrosinase (a thioglucosidase) to produce a variety of different breakdown products, such as glucose, sulfate, isothiocyanates, epithionitriles, nitriles, indolic alcohols, oxazolidinethions, amines, and thiocyanate. It is believed that both GSLs and their breakdown products play an important role in the defense mechanism of plants against the attacks of fungi, insects, and microorganisms [GSLs are relatively stable compounds, but plant cells containing glucosinolates can be damaged due to the release of β-thioglucosidase or myrosinase enzymes during the cutting, mixing, chopping, and chewing of food . This enrganisms ,30. MoreIn order to preserve the beneficial properties of cruciferous plants, it is important to store and prepare them properly. The content of glucosinolates varies between cruciferous plant types, but the bioavailability of these compounds can be significantly increased by appropriate storage and preparation. The requirements for storage conditions vary in the available literature. Some reports indicate long-term storage at room temperature and in the refrigerator. Others suggest a significant decrease in GSLs after seven days of storage. An even greater decrease was noted after 6 h of storage for cruciferous vegetables that were previously chopped . In the 2/day) has a positive effect on the content of GSLs in broccoli. One of the newer and more promising methods that can be used in the treatment of cruciferous vegetables while maintaining a high GSL content is pulsed electric field (PEF). The advantage of this technique is the possibility to penetrate into the plant tissue without using a very high temperature and damaging the tissue mechanically [Several researchers have studied the thermostability and degradation of the GSLs contained in broccoli, red and white cabbage, and brussel sprouts. The results obtained indicate that the important determining factor is media type (food matrix or cooking water) . Florkieanically . With thAn important factor determining the bioavailability of GSLs is also human microflora. The most crucial role from the point of view of GSL metabolism is played by such enzymes as myrosinase and β-thioglucosidase. β-thioglucosidase is present in the human gut microbiome and influences human physiology and health, for example by regulating the metabolism of GSLs and their derivatives. In turn, the human homolog of myrosinase has not been described so far. The direct administration of isothiocyanates results in their higher bioavailability than that of GSLs, which indicates fundamental differences in such properties as lipophilicity. Therefore, a detailed knowledge of the factors determining the efficiency of the GSL to isothiocyanates conversion seems to be crucial . A commoIn addition to the scientific challenges, the creation of supplements from properly prepared cruciferous plants to increase the degree of GSL assimilation is a great opportunity, especially since heat treatment is one of the factors negatively affecting the degree of absorption. In the United States, there is a growing increase in the sale of dietary supplements from cruciferous plants, which are prepared in the form of extracts and powders and then sold in capsule form. This again confirms the need for a deeper analysis of many factors influencing the transformation and metabolism of the compounds they contain, not only in healthy people but also in the interactions with drugs taken or the possible therapeutic potential. The Food and Drug Administration (FDA) has also issued a number of warnings about the information on the labels of these types of dietary supplements that provide anti-cancer properties and other beneficial effects but which have not yet been confirmed by clinical trials . This isCurrently, little is known about how GSLs and their derivatives are metabolized in the human organism, as most reports relate to cell culture and animal studies. Data in humans are extremely sparse and mostly relate to the level of metabolites in the urine. With many compounds, it is the case that, despite the high level of absorption, the bioavailability is limited and reduced due to the high metabolic rate. With regard to the literature data on GSL metabolites, they are most often the result of the level of nitrogen compounds, isothiocyanates, and other derivatives. Additionally, the measurement of the level of metabolites in urine confirms that the hydrolysis reaction is an important criterion for the bioavailability of GSLs. The level of metabolites resulting from prior hydrolysis was six times higher than that of GSLs . The higThe phytochemicals in cruciferous plants can exert different effects that affect the immune system. Inflammation leads to a number of changes in the organism, such as increased cellular proliferation; the inhibition of apoptosis; and, as a result, an increased risk of various types of cancer. Biologically active substances that occur in cruciferous plants inhibit the development of inflammation by, for example, activating detoxification enzymes, removing reactive oxygen species, and stimulating natural defense mechanisms . SpecifiIn addition to those effects described above, SFN shows a wide range of impacts. It blocks apoptosis, inhibits the release of reactive oxygen species, and affects the expression of vascular cell adhesion molecule (VCAM)-1 or key molecules in the IL-1 signaling cascade. Other important properties of this isothiocyanate include the suppression of such inflammatory reaction markers as interferon regulatory factor 3 (IRF3), as well as macrophage migration inhibitory factor (MIF). These beneficial effects of SFN are the result of the activation of various signaling pathways and mechanisms, which involve, among other Toll-interleukin-1 receptor domains, Toll-like receptors (TLRs) and interferon-beta. Among the numerous activities of SFN, its chemopreventive potential cannot be overlooked. Indeed, SFN suppresses cyclooxygenase-2 (COX-2), which has immunoreactivity properties. These effects were again mainly NF-κB- and ERK -mediated. Another important compound of cruciferous vegetables with a significant impact on the immune system is berteroin (5 methylthiopentyl isothiocyanate). It is found in the largest quantities in vegetables, such as cabbage and rucola. It has been shown that its frequent consumption reduces edema by modulating COX-2, NF-κB, and ERK activity. There are also reports indicating the beneficial effect of cruciferous vegetables on the immune system parameters in healthy people. These concerns, in particular, the concentrations of circulating inflammatory biomarkers, such as IL-6, CRP, and sTNFRI. However, further more complex studies are needed to confirm that a cruciferous vegetable-rich diet reduces high levels of inflammatory markers in populations where this pathological condition persists chronically . Glucosinolates and their metabolites, such as isothiocyanates like sulforaphane (SFN), have several health-promoting activities . Sulfur-For example, SFN is one of the best known isothiocyanates, mainly due to its anticancer properties, but also because of its ability to reduce the secretion of inflammation markers by immune system cells, as well as nuclear factor kappa B (NF-κB), which plays an important role in the rapid regulation of immune reactions. In vitro studies have confirmed the anti-inflammatory properties of a diet rich in cruciferous vegetables. Among others, reduced serum concentrations of pro-inflammatory cytokines, such as IL-1β, TNFα, and IL-6, were observed . SeveralThe dietary consumption of GSLs and their metabolites may diminish the damaging impact of chemical carcinogenesis. A study reported that the risk of colon cancer is enhanced among individuals with a low consumption of broccoli, cabbage, and brussel sprouts, and reduced among those with a high consumption of these vegetables . These fThe consumption of broccoli sprouts or SFN could decrease oxidative stress in cardiovascular and kidney tissues. This might be due to the decreased protein nitrosation and high levels of glutathione, glutathione peroxidase, and glutathione reductase . In addiSimilar to cardiovascular and carcinogenesis disease, chronic inflammation and oxidative stress are the main cause of pathogenesis of the central nervous system. The administration of SFN could decrease the infarct size, brain edema, and cortical apoptosis, reducing the inflammation and tissue damage of the central nervous system. Mostly, the neuroprotective impacts of SFN are due to the activation of the transcription factor Nrf2 and the upregulation of different target genes . InteresSFN was also found to be effective in improving renal performance and reducing pathological changes in glomerulus. The velocity of neuron conduction, blood flow, and pain behavior were significantly improved to alleviate different metabolic disorders and improve protection against renal damage .Brassicaceae may play a significant role in the defence of the plant against fungal infection. However, the changes in disease resistance may depend on both the fungal pathogen species and the composition of the GSLs in the plant. This suggests that resistance against both fungi was due to a combination of the direct toxicity effect of GSLs and the activation of systemic acquired resistance. There is a strong interest in altering the levels of specific GSLs in order to obtain certain GSLs with desirable properties in terms of flavor; cancer prevention; and, in particular, pathogen protection [It is evident that the GSLs in otection . Howeverotection . In turnotection . GSL hydotection .A random double-blind placebo study in 103 Scandinavian patients diagnosed with type 2 diabetes mellitus showed that the daily consumption of broccoli extract for 12 weeks significantly reduces glucose and HbA1C levels. The most spectacular effect was observed in obese patients with high insulin resistance. Closer research by Iranian scientists has shown that the daily consumption of broccoli extract powder for 4 weeks reduces the plasma insulin levels in patients with the same type of diabetes. In addition, the beneficial effects of broccoli extracts have been demonstrated to minimize diabetes-related complications as well .There are also reports of the potential beneficial effects of cruciferous vegetable consumption on skin problems. In five volunteers with epidermolysis bullosa simplex who were given daily broccoli extract, the alleviation of skin lesions and induced expressions of keratins 16 and 6 were observed. It has also been suggested that products rich in SFN can reduce the risk of skin lesions caused by UV radiation, especially in high-risk patients. Nevertheless, further research is necessary to confirm the therapeutic potential of cruciferous plants in skin diseases .Allium plant family. Allium is a Latin word used for garlic, while the meaning of Allium sativum is cultivated garlic. The genus Allium belongs to the monocot flowering plants, which include almost 900 different species, such as cultivated onions, garlic, shallots, leeks, scallions, and chives [Alliaceous vegetables are considered to be the rich source of a large variety of beneficial bioactive compounds, such as oligosaccharides, arginine, flavonols, and OSCs. Alliaceous vegetables belong to the d chives . The lead chives . In gened chives . Much ofd chives ,86. Manyd chives ,83. Allium plants are consumed worldwide as vegetables, seasonings, and spices. Several types of Allium products, such as pastes, oils, salts, powders, and flakes, have been used in industries and homes as food seasonings and flavorings. These products can be prepared by different methods, including boiling, frying, baking, and drying. The differences in preparation methods can produce different amounts of bioactive sulfur compounds ,88. ThesAllium oil is produced by the distillation of minced vegetables . Juice extraction from milled alliaceous vegetables is conducted either by decantation or pressing. Allium extracts are extracted by the alcoholic extraction (methanol and ethanol) of raw materials. During extraction, nitrogen gas or liquid nitrogen can be added to isolate, concentrate, and trap the volatile compounds in the products [Allium sp [Alliaceous vegetables are processed to different products such as dietary supplements, oil macerates, essential oils, powders, and extracts. These are available in numerous fresh and dehydrated products and are utilized in a wide range of medicine and dishes . The dehproducts . The temproducts . This reproducts . The OSCproducts . High-teproducts ,96. Highproducts . On the products ,99. Moreproducts . Similarllium sp . TherefoAllium products. As stated earlier, heat treatment inactivates alliinase, thereby reducing the bioactive contents of the Allium product and diminishing the anticancer effects of these vegetable [Bioactive sulfur compounds may undergo undesirable degradation during processing—in particular, conventional thermal techniques may negatively impact the bioavailability and physiological characteristics of egetable . Additioegetable . There iegetable ,104. Thuegetable ,106. ThuS-alk(en)yl-l-cysteine sulfoxides (ACSOs) content in fresh garlic is 3–14 mg/g of the fresh weight [S-allylcysteine sulfoxide) is the major SACSO found in garlic. Another garlic compound that has been proven to induce apoptosis in tumor cells is the alliin thiosulfinate metabolite, allicin [Sulfur comprises approximately 1% of the dry weight of garlic, and the sulfur-rich compounds found in garlic correspond to its health benefits, especially in terms of cancer prevention and therapy . The S-ah weight . Among A allicin ,109. The allicin ,111. The allicin ,113. Mos allicin , but the allicin ,116. All allicin  was obse allicin ,118. The allicin . Recentl allicin .Allicin is highly unstable and is converted to lipid-soluble sulfides such as diallyl sulfide (DAS), diallyl disulfide (DADS), and diallyl trisulfide (DATS) . Those sAlso, garlic-derived water-soluble compounds, such as S-allyl cysteine (SAC) and S-allylmercaptocysteine (SAMC), were investigated with respect to cancer. By processing garlic, an aged garlic extracts rich in SAC (0.62 mg SAC/g product) and SAMC, among other things, could be obtained. SAC could be easily absorbed from the gastrointestinal tract in an unchanged form and have a 30 times lower toxicity than some other garlic bioactive components in studies conducted in animal models. SAC displays stronger antioxidant properties than fresh garlic, and it has strong anti-cancer properties not only at the early but also at the late stages of the disease. A thiol nucleophile group in SAC thiol is responsible for the scavenging of free radicals, as well as activating antioxidant enzymes such as catalase and glutathione peroxidase ,136,137.Some other epidemiological studies were poorly organized and did not reach statistical significance ,140. MorThere is some evidence that natural sulfur-containing phytochemicals could display anticancer properties. The purported compounds and mechanisms of action have been identified. The observed effects seem to depend on the concentration and bioavailability of the compounds. The biggest challenge is to determine the criteria for the storage and preparation or processing of alliaceous and cruciferous vegetables. Furthermore, conducting a number of clinical trials on a significantly large number of volunteers to optimize doses and learning the factors determining the bioavailability is necessary. Finally, the safety and potential effectiveness of treatment in specific diseases should be assessed. A metanalysis of the available trustworthy clinical records should be performed.


In addition, we also discuss another two programs relating to brachyury function: epithelial-to-mesenchymal transition (EMT) and cell cycle control, both of which implicate in the regulation of brachyury on biological behavior of tumor cells. This review will provide an overview of the function of master transcriptional factor brachyury, compare the similarities and differences of its role between embryonic development and carcinogenesis, and list the evidence on which brachyury-target therapies have the potential to help control advanced cancer populations.Transcription factor brachyury, with a DNA-binding T-domain, regulates posterior mesoderm formation and notochord development through binding with highly conserved palindromic consensus sequence in a variety of organisms. The absence of brachyury expression in majority of adult normal tissues and exclusive tumor-specific expression provides the potential to be developed into a novel and promising diagnostic and therapeutic target in cancer. As a sensitive and specific marker in the diagnosis of chordoma, brachyury protein has been verified to involve in the process of carcinogenesis and progression of chordoma and several epithelial carcinomas in various studies, but the mechanism by which brachyury promotes tumor cells migrate, invade and metastasis still remains less clear. To this end, we attempt to summarize the literature on the upstream regulatory pathway of brachyury transcription and downstream controlling network by brachyury activation, all of which involve in both the embryonic development and tumor progression. We present the respective correlation of brachyury expression with tumor progression, distant metastasis, survival rate and prognosis in several types of tumor samples , and various brachyury gain-of-function and loss-of-function experiments are summarized to explore its specific role in respective tumor cell line  The T-box genes encode a family of transcription factors, characterized by a highly conserved DNA-binding domain of about 180 amino acid residues, which is designated as T-domain , and areBrachyury (“short tail” from Greek) origins from the phenotype of this gene mutant mice, most striking defect with a truncated tail, which was first described by Dobrovolskaia-Zavadskaia in 1927  gene is required for the formation of posterior mesoderm and axial development. In all vertebrates, the gene is initially expressed throughout the presumptive mesoderm, and during later stage, the expression is gradually restricted to the developing notochord and tail bud  and fibroblast growth factor receptor (FGFR) signaling has been implicated in the patterning of mesoderm and activated pression –28. In X of Xbra , 29.Brachyury is expressed in a manner dependent on the FGF-mitogen-activated protein kinase kinase (MEK)-mitogen-activated protein kinase (MAPK)-Ets signaling pathway and on the intrinsic factors Zic and FoxA. Binding of Ets and ZicN at the 5′ upstream of Brachyury promoter region is required for FGF-responsive Brachyury gene activation in notochord precursor cells , tumor cell growth, and invasion.The study by Hu et al.  suggestsBrachyury expression in ascidian embryogenesis  has been reported to induce notochord formation and ogenesis . Activinogenesis –37 have Ciona, an invertebrate chordate, which is a commonly used model to study BRACHYURY function, over 50 validated genes have been identified to be controlled by BRACHYURY , an effector of the Hippo pathway and a master regulator of organ development , was rec cancers .BRACHYURY could be in the tumorigenesis of chordoma, gene duplication mutation and overexpression in samples verified by previous various studies suggest that BRACHYURY might be a crucial molecular driver in the initiation and propagation of chordoma  in any disease type. Nevertheless, BRACHYURY duplication is extremely rare in sporadic chordoma  either through polysomy of the entire chromosome 6 or structural rearrangements, which indicates that cCNGs of BRACHYURY are pathogenetically relevant in sporadic chordoma. Cho et al.  and overall survival rate in primary lung carcinoma samples 2 and MMP24  inhibitor  is the mediator of androgen activity in normal and malignant prostate cells. The BRACHYURY protein level in the nucleus of primary prostate cancer cells is statistically associated with the presence of AR , and theBRACHYURY/BRACHYURY has become a potential promising option in such a tricky scenario. A phase I/II trial (NCT03493945) testing a BRACHYURY-targeted antitumor vaccine has been performed in metastatic CRPC recently  develops and tumor progression occurs due to the induced epithelial-to-mesenchymal plasticity (EMP) and neuroendocrine transdifferentiation (NEtD) programs by androgen deprivation , 72, therecently .BRACHYURY/BRACHYURY on the tumorigenesis and progression of prostate cancer has been recognized, more details need to be further investigated and unveiled, for instance, the mechanism of BRACHYURY involved in NEtD, the biological significance of BRACHYURY binding with the regulatory elements of the marker genes  of prostate cancer.Although the specific role of BRACHYURY mRNA expression was found to elevate in tumors of the small intestine and in the majority of cell lines derived from the colon , including cytoplasmic and nuclear staining . BRACHYUThe process of EMT, converting immotile epithelial cells to migratory mesenchymal cells, is associated with enhancement of invasive and metastatic potential of tumor cells, as well as resistance to therapeutic interventions , 79. BRABRACHYURY 8, and transforming growth factor (TGF)-β1, among others. Various studies have attempted to investigate the correlation between the expressions of these genes and RACHYURY , 84, 85.RACHYURY , 87. FerRACHYURY  reportedRACHYURY , 82. In and Slug , 49, andchordoma . BRACHYUchordoma .BRACHYURY expression at mRNA and protein levels in primary and metastatic lung tumor tissues and associated with poor prognosis . BRACHYURY promotes tumor metastasis through modulating the EMT process and regulating cell cycle and closely correlates with patient poor prognosis. pression  but alsopression , 91, 92,All authors listed have made a substantial, direct, and intellectual contribution to the work and approved it for publication.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.


Plastid genomes typically display a circular, quadripartite structure with two inverted repeat regions, which challenges automatic assembly procedures. The correct assembly of plastid genomes is a prerequisite for the validity of subsequent analyses on genome structure and evolution. The average coverage depth of a genome assembly is often used as an indicator of assembly quality. Visualizing coverage depth across a draft genome is a critical step, which allows users to inspect the quality of the assembly and, where applicable, identify regions of reduced assembly confidence. Despite the interplay between genome structure and assembly quality, no contemporary, user-friendly software tool can visualize the coverage depth of a plastid genome assembly while taking its quadripartite genome structure into account. A software tool is needed that fills this void.We introduce ’PACVr’, an R package that visualizes the coverage depth of a plastid genome assembly in relation to the circular, quadripartite structure of the genome as well as the individual plastome genes. By using a variable window approach, the tool allows visualizations on different calculation scales. It also confirms sequence equality of, as well as visualizes gene synteny between, the inverted repeat regions of the input genome. As a tool for plastid genomics, PACVr provides the functionality to identify regions of coverage depth above or below user-defined threshold values and helps to identify non-identical IR regions. To allow easy integration into bioinformatic workflows, PACVr can be invoked from a Unix shell, facilitating its use in automated quality control. We illustrate the application of PACVr on four empirical datasets and compare visualizations generated by PACVr with those of alternative software tools.https://cran.r-project.org/package=PACVr.PACVr provides a user-friendly tool to visualize (a) the coverage depth of a plastid genome assembly on a circular, quadripartite plastome map and in relation to individual plastome genes, and (b) gene synteny across the inverted repeat regions. It contributes to optimizing plastid genome assemblies and increasing the reliability of publicly available plastome sequences. The software, example datasets, technical documentation, and a tutorial are available with the package at  The sequencing and comparison of complete plastid genomes has become a popular method in plant evolutionary research, rendering the precise genome assembly and its quality assessment of high importance. The plastid genomes of most photosynthetically active land plants display a circular, quadripartite structure and comprise two single copy (SC) regions separated by two identical inverted repeats (IR) . A totalDespite the development of assembly algorithms customized for plastid genomes, the plastome assembly process remains imperfect and often requires the verification, if not manual correction, of the assembly product. Concurrent with the surge in plastid genome sequencing, many new algorithms and pipelines specifically designed for the assembly of plastid genomes have been developed –23. MostSeveral measures have been used to indicate the quality of plastid genome assemblies, including contiguity metrics and the length and sequence equality of the IR regions, but sequencing coverage remains one of the most popular proxies for assembly quality. In genome research, the length of the shortest among all those contigs that cover at least 50% of a reference genome is often used as an indicator for the quality of a draft genome . The cloCurrently available software tools can generate either unpartitioned plots of plastome coverage depth or quadripartite plastome maps, but the simultaneous, user-friendly visualization of both aspects is presently unsupported. When employing currently available software tools, plant biologists must decide if they wish to visualize either plastome sequencing coverage as unpartitioned, often linear plots or, alternatively, the circular, quadripartite structure of a plastid genome. The assembly pipeline FastPlast , for exaGiven the plethora of complete plastid genomes generated in biological research each year , strong The input to PACVr consists of two different files of common file format which contain information on (A) genome sequence and structure, and (B) coverage depth. Information on genome sequence and structure, as well as the genes encoded by the sequence, is supplied via an input file in the GenBank flatfile format. GenBank flatfiles represent the default file type for sequence retrievals from NCBI Nucleotide  and contThe output of PACVr is a multi-layered, annotated plastome map with coverage information across the genome. Specifically, PACVr generates a circular, quadripartite map of the plastid genome in which coverage depth values are displayed as histogram bars, with bars below a predefined threshold highlighted in red and partition-wide average coverage values superimposed as horizontal, yellow lines. The map also displays positional information in regular intervals in the form of labeled tick marks as well as the location of all plastome genes, allowing the user to relate areas of low coverage depth to specific genome regions and genes. The map generated by PACVr is saved in PDF format to a user-defined output file.Coverage depth is calculated by PACVr via the application of user-defined window sizes with the software mosdepth . Window-Equality among the IR regions is evaluated by PACVr both directly and indirectly. The direct evaluation is done by the computational comparison of the sequences of IRa and IRb, the indirect evaluation via the numerical and visual comparison of number, length, and location of all genes contained in the IRs. Specifically, the software conducts a two-step procedure in which equality in sequence, sequence length, and the number of genes is confirmed across the two IR regions, and the equality then visualized by connecting the matching genes of the two IRs via blue connector lines. To that end, PACVr computationally extracts the two IR regions from the input sequence record, stores their sequences as well as the names, start and end positions of all IR genes in separate data frames, and compares the exact number, length, and location of the genes across both regions. Any difference in sequence, sequence length, or gene complement between the two IRs results in a warning message to the user. PACVr then visualizes the equality between the IRs by connecting genes with identical names across the regions using blue connector lines. The lines hereby originate and end at the central nucleotide of each gene shared between the two IRs. The start and end width of these connector lines can be set to be uniform or proportional to the length of the genes they connect. Any difference in name or location of the IR genes becomes visible through unequal or missing connector lines, thus enabling the visual assessment of equality among the IR regions regarding gene presence and synteny. This visualization of gene location and synteny contributes to the discovery of rules and patterns in genome orientation and rearrangements .PACVr employs RCircos  as the vycf1 and ycf2, which are located near the 5’ end of the SSC and the IRa, respectively, in most angiosperms and represent nearly 10% of the unit-genome length [rps12 in many angiosperms [The quadripartite structure of plastid genomes requires adjustments in the calculation of coverage depth and the visualization of IR equality compared to unpartitioned chromosomes. By default, PACVr calculates window-sized coverage depth values and, based on these, the region-wide average for each of the four plastome regions. However, PACVr would double-count the coverage of those windows that span across a region boundary, unless the coverage calculation included a special adjustment. Similarly, PACVr requires customization when visualizing the equality of gene position and synteny between the two IR regions. Natural expansions in IR size can cause genes located near the border of a single copy and an IR region to be displaced from one region into the other over time , 76. Wite length . A similiosperms ). Thus, https://cran.r-project.org/) using the R command install.packages(′PACVr′). It requires the presence of the R packages optparse [https://github.com/michaelgruenstaeudl/PACVr. The technical documentation and a user tutorial (vignette) is distributed as part of the R package. The vignette provides example commands for the installation and execution of PACVr as well as for the generation of BAM files under different read filtering settings.PACVr was written in R and can be installed via the Comprehensive R Archive Network  the window size for calculating coverage depth, with a default value of 250; (ii) the shell command to execute mosdepth, with a default command of mosdepth; (iii) the coverage depth threshold above which histogram bars are plotted in red as opposed to the default black, with a default value of 0.5; (iv) the selection if the threshold value is specified relative to the average genome-wide coverage depth as opposed to representing an absolute value, with the default set to true; (v) the selection if the coverage depth values are to be log-transformed prior to visualization, with the default set to false; (vi) the selection if and what type of connector lines to draw between matching genes of the IRs, with the default line type displaying a start and end width proportional to the length of the genes that the lines connect; (vii) the size of all text elements of the resulting visualization relative to the maximum font size, with the default set to 0.5; (viii) the decision to remove all temporary files generated during the coverage depth calculation, with the default set to true; and (ix) the name of, and file path to, the output file, with the output saved as ./PACVr_output.pdf by default. The software can be invoked either from within the R environment or directly from a Unix shell. A complete list of the short- and long-flag command-line (CLI) arguments available when invoking PACVr from the Unix shell is displayed via the shell command Rscript./inst/extdata/PACVr_Rscript.R -h/—help. In the framework of an automated workflow (and upon setting the location of PACVr to a shell variable with the same name), the following shell command can, for example, be used to execute PACVr on the empirical dataset co-supplied with the R package:Archidasyphyllum excelsum (Asteraceae) and Nuphar japonica (Nymphaeaceae) and display a quadripartite genome structure as well as a genome size typical for the majority of angiosperms [Pelargonium x hortorum  and thellaceae; ) and disllaceae;  and greellaceae; , respecttructure , 84. Dettocol of  and the tocol of ; the pla v.33.89 , which rNuphar japonica . Successful visualizations of coverage depth were also conducted for the plastid genomes of Pelargonium x hortorum . The threshold value for highlighting histogram bars was set to 100% of the average genome-wide coverage depth for both genomes to contrast this option with the visualizations of the newly generated plastid genomes. Second, the visualizations by PACVr indicated strong gene synteny across the IRs of those plastid genomes under study that possess a quadripartite genome structure. Specifically, the symmetric display of equal gene position and length via blue, variable-width connector lines between the IRs of a genome indicated IR gene synteny in Nuphar japonica, Archidasyphyllum excelsum, and Pelargonium x hortorum . Based on the resulting visualizations, several important observations were made. First, the visualizations indicated differences in region-wide average coverage depth and the presence of genomic areas with markedly lower coverage depth compared to other areas of the same genome in each of the plastid genomes under study. In the plastid genome of ica Fig. , for exaNuphar japonica, FastPlast generated a linear plot of coverage depth that indicated a higher depth in the area corresponding to the IRs than in the large and small SC regions Other requirements: R packages BiocGenerics, Biostrings, GenomicAlignments, genbankr, optparse; mosdepth (>= 0.2.5)License: BSD 3-ClauseAny restrictions to use by non-academics: noneAdditional file 1 Visualizations of coverage depth of the plastid genome of Archidasyphyllum excelsum (GenBank accession MH899017) as generated by the software tools FastPlast, IOGA, Circleator, and PACVr. (PDF 8211 kb)Additional file 2 Visualizations of coverage depth and IR equality of the plastid genome of Pelargonium x hortorum (GenBank accession NC_008454) and Prototheca cutis (GenBank accession NC_037480). (PDF 5540 kb)


Non-communicable diseases associated with immune system dysfunction are an increasing challenge for twenty-first century medicine. There is growing evidence that predisposition to many such conditions originate during early-life “programming events.” This term refers to critical points of developmental plasticity where changes in environmental factors have long-term effects on physiological development, including the development of the immune system.From birth, the neonatal immune system develops rapidly in response to external stimuli, primarily from the intestinal microbiota and nutrition, and this pattern of development is essential in adult immune functionality and competence , 2. NutrThe adult microbiota is largely stable and even substantial interventions have limited long term impact on the composition and biochemical activity of the microbiome. However, sustained shifts in microbial ecosystems can occur in early life, especially during the neonatal period and weaning. This can be in response to nutritional factors, such as malnutrition and breastfeeding, but also in response to events such as cesarean delivery and antibiotics. Shifts in the patterns of early microbial colonization can subsequently change the development and function of the host immune system and are likely to lead to developmental imprinting. Thus, the dynamic evolution of the gut microbiota during early-life may represent a window of opportunity in which to influence the development of immunity through nutritional interventions, or the administration of specific bacterial strains , 6.Azagra-Boronat et al. investigated the effect of one of the most active and abundant human milk oligosaccharides (HMOs), 2-fucosyllactose (2-FL) in a suckling rat model. 2-FL exhibited prebiotic activity and drove the development of the microbiota to a more Lactobacillus-rich phenotype with increased butyrate production, and promoted the maturation of immunity in early life. In addition, 2-FL enhanced gut barrier function, possibly through the generation of divergent bacteria- and host-derived metabolic profiles.This Research Topic explores prenatal and postnatal dietary strategies which differentially modify the maturation of the immune system and the microbiota, and may have physiological impact later in life. Perdijk et al. compared the effects of another important HMO, sialyllactose (SL) with galactooligosaccharides (GOS), a prebiotic enzymatically derived from lactose that is widely used in infant formulas, using multiple in vitro approaches. Both prebiotic compounds induced re-epithelialization in an in vitro epithelial wound repair assay and induced gene expression of alkaline phosphatase, a marker of epithelial cell differentiation. However, SL and GOS differentially modified microbiota composition and SCFA production.Taken together these studies provide further evidence that both HMOs and GOS drive important actions on the microbiota, immunity, and barrier development in early life. However, it should be noted that a single HMO cannot reflect the structure and function of the diverse oligosaccharide composition in breast milk. Breast milk provides many other important components such as cytokines, chemokines, hormones, and growth factors  which maGrases-Pintó et al. assessed the role of leptin and adiponectin, which are both present in breast milk, in neonatal rats. Early supplementation with both leptin and adiponectin influenced the composition of the gut microbiota at the end of the suckling period. In addition, innate immunity-associated molecules in the intestine, including mucin, were also affected by breast milk. Their study concluded that supplementation with adipokines had an impact on the phenotype of those immune cells in close contact with the intestinal milieu, the intraepithelial lymphocytes. This reinforces the idea that nutritional programming of the immune system by breast milk may be driven by factors other than oligosaccharides.Guadamuro et al. demonstrated that the introduction of intact CMP to infants with non-IgE mediated CMP allergy changed both immune mediators and the composition and metabolic end-products of the microbiota. The authors also identified that consumption of the probiotic L. rhamnosus GG during CMP allergy may have contributed to gut homeostasis by fine-tuning these profiles.One of the most prevalent food allergies during the first year of life is to cow's milk proteins (CMP) and the incidence is increasing rapidly. Christoforidou et al.. Early sex disparities were found to underlie both mucosal immune development and systemic responses to novel antigen. Crucially, significant effects on immune development of nutritional interventions with inulin or Bifidobacterium lactis were also sex dependent. The importance of sex-stratifying nutritional interventions, even in very young animals, could translate to personalized recommendations for human infants.Sexual dimorphism can also influence immunity in an out bred infant pig model as identified by Together, these results demonstrate that early nutrition can significantly modify the gut environment in infants and have positive impacts on immune, metabolic, and microbiota development during early life. Since the development of these systems is an important determining factor in later health, it follows that early nutrition could have a lasting impact on lifecourse health. This Research Topic therefore provides new insights to support the development of therapeutic dietary interventions to achieve optimal immune and metabolic health.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.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.


The current COVID-19 pandemic has resulted in over 54,800,000 SARS-CoV-2 infections worldwide with a mortality rate of around 2.5%. As observed in other airborne viral infections such as influenza and SARS-CoV-1, healthcare workers are at high risk for infection when performing aerosol-generating medical procedures (AGMP). Additionally, the threats of a global shortage of standard personal protective equipment (PPE) prompted many healthcare workers to explore alternative protective enclosures, such as the “aerosol box” invented by a Taiwanese anesthetist. Our study includes the design process of a protective barrier enclosure and its subsequent clinical implementation in the management of critically ill adults and children infected with SARS-CoV-2.  The barrier enclosure was designed for use in our tertiary care facility and named “SplashGuard CG” (CG for Care Givers). The device has been adapted using a multi- and interdisciplinary approach, with collaboration between physicians, respiratory therapists, nurses, and biomechanical engineers. Computer-aided design and simulation sessions throughout the entire process facilitated the rapid and safe implementation of the SplashGuard CG in different settings  during AGMPs such as bag-valve-mask ventilation, nasopharyngeal suctioning, intubation and extubation, and noninvasive ventilation. Indications for use and anticipatory precautions were communicated to all healthcare workers using the SplashGuard CG. The entire process was completed within one month.  The rapid design, development, and clinical implementation of a new barrier enclosure, the “SplashGuard CG,” was feasible in this time of crisis thanks to close collaboration between medical and engineering teams and the use of recurring simulation sessions to test and improve the initial prototypes. Following this accelerated process, it is necessary to maintain team skills, monitor any undesirable effects, and evaluate and continuously improve this new device. The current COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in over 54,800,000 cases of infection since its first description in December 2019 in Wuhan, China, and has already led to over 1,300,000 deaths worldwide . HoweverFor hospitalization of a COVID-infected patient, it is recommended to admit them to a negative-pressure room and for healthcare providers to wear standard personal protective equipment (PPE), namely, gown, gloves, and an N95 mask. Access to the standard PPE could become limited because of the high global demand, interrupted transportation lines, and altered deliveries in many countries including Canada . SeveralWe devised a similar protective barrier enclosure at our tertiary care facility and named it “SplashGuard CG,” CG for Care Givers . The SplIn this report, we aim to describe in detail the accelerated process of design and implementation of this new open innovation technology, with particular emphasis on the role of simulation in its development. We will also share the challenges encountered throughout this process, which took place at the very beginning of the COVID pandemic.The prototype design took place in the technical laboratory using computer aided design (CAD), with the collaboration of a pediatric intensivist and a biomechanical engineer.The simulation testing was performed in a simulated clinical environment. The prototype was tested using simulations of various clinical situations including an intrahospital patient transport, installation of noninvasive ventilation, and endotracheal intubation using pediatric and adult manikins. A select interdisciplinary team  to rapidly and efficiently train healthcare providers from the pediatric emergency, anesthesia, and pediatric critical care sectors. Rapid Cycle Deliberate Practice is a simulation-based instructional strategy that focuses on rapid acquisition of necessary skills when training for emergency clinical situations .During the final phase, we established cleaning and disinfection procedures  and obtaThe aim of the device development was to create a protective barrier enclosure for care givers, to be used in combination with standard PPE, for protection against contact and airborne transmission from pediatric and adult proven or suspected COVID patients. Its development and implementation used a living lab methodology including prototyping by engineers and testing in simulated conditions by end users see :The protThe development team included biomechanical engineers from Polytechnique Montreal, simulation specialists from the Mother-Child Simulation Center at the Centre Hospitalier Universitaire Sainte-Justine, physicians from the pediatric intensive care unit and the emergency and anesthesiology departments, as well as respiratory therapists and nurses. In addition, refining of this new technology required further collaboration with biomedical engineering, the infection prevention unit, and the ergonomic units at our institution (see SplashGuard CG Study Group).Starting on March 23, 2020, the SplashGuard CG was designed and developed at the Centre Hospitalier Universitaire Sainte-Justine in Montreal, Canada. The main objective during the development of the SplashGuard CG was to ensure safety and protection of healthcare workers managing a critically ill COVID patient, suspected or confirmed, who is in respiratory distress and requires aerosol-generating medical procedures, while equally respecting patient safety. Indications for use of the SplashGuard CG were established for hospitalized and bedridden patients with a highly contagious airborne infection, e.g., SARS-CoV-2. The use of the SplashGuard CG is suggested for (1) procedures at the head of the patient: intubation or extubation procedures, the placement of a oro- or nasogastric tube, oro- or nasopharyngeal aspiration, and installing the patient on high-flow nasal cannula or noninvasive ventilation and (2) patients admitted and supported by high-flow nasal cannula, noninvasive ventilation, or invasive ventilation (following the intubation procedure or tracheostomy), during transport or during their stay in one the various departments .Following simulation sessions, the SplashGuard CG was developed and manufactured in 2 sizes: a larger and a standard model Figures  and 2. TFollowing a first simulation session with the initial prototype, the size and height of the box were adjusted to the space needed for providers to perform different aerosol-generating medical procedures in a patient with a suspected or confirmed SARS-CoV-2 infection. We also adjusted the position of the access ports to ensure successful and safe assistance and completion of AGMPs. The vertical panel at the distal end of the SplashGuard CG, present in the first prototype, was changed for a plastic cover sheet attached to the SplashGuard CG to avoid patient neck or head injury when the patient wants to sit up.Anticipatory precautions when using the SplashGuard CG were also identified and further detailed during the whole process. There are theoretical risks of asphyxia, hypercarbia, impaired care during emergency situations, injury, and pressure wounds, which we will describe in detail in the following section.(1) Risk of asphyxia: the SplashGuard CG cannot be used in a unit where the patient is not under continuous surveillance because of the risk of acute asphyxia associated with the use of soft plastics close to the face. Thus, it is mandatory that the SplashGuard CG be used in a setting offering continuous patient monitoring, such as the intensive care unit, the emergency department, or the operating room. (2) Risk of hypercarbia: if the SplashGuard CG is used in a patient who is breathing spontaneously, a theoretical risk of hypercarbia exists if the gas flow entering the SplashGuard CG is insufficient. (3) Other risks attributable to emergency situations: there are conditions that require the immediate removal of the SplashGuard CG such as cardiac arrest, but access to the patient could be delayed by its presence. For that reason, a pair of scissors is attached to the top of the box at all time to cut the fixation ties if necessary. (4) Risk of injury: it is necessary to secure the SplashGuard CG to the hospital bed or stretcher, especially in cases of prolonged use, transport of a COVID-patient between units or when caring for agitated pediatric patients, by using the anchoring points designed for that purpose. (5) Risks related to pressure wounds: the possible risk of developing a pressure wound due to immobilization was also addressed in the instruction manual, with areas at risk identified as the scalp, shoulders, and arms of the patient.After elaborating the final prototype, the device was once again tested for use during a second simulation session . FollowiThe use of the SplashGuard CG is also recommended when invasive positive-pressure ventilation is maintained, at the discretion of the healthcare team, especially when considering the use of prone positioning. When use of an enclosed system for endotracheal suctioning is not possible, risks of aerosolization are increased because it can trigger cough, so the SplashGuard CG can also be used in this context.To ensure protection against the aerosolization of viral particles when no AGMPs are performed, six dispensable plastic obturators for the large circular access ports were created, in addition to the use of a plastic transparent shield on the open side of the device which allows easy access to the patient at all times. When using the SplashGuard CG in the intensive care unit, one or two of the obturators at the front panel contain an opening to connect to the wall suction to create a negative-pressure environment within the box and limit aerosolization of viral particles.The use of the SplashGuard CG for caregiver protection, when supporting patients with noninvasive positive-pressure ventilation or high-flow nasal cannula, requires the availability of continuous aspiration in the device, by using the obturators as described before. For short procedures , continuous aspiration is not recommended in order to allow the provider and those assisting him full access to the patient's head.Performance of a simulated cardiac arrest requiring cardiopulmonary resuscitation (CPR) demonstrated the limitations imposed by the SplashGuard CG as to proper hand positioning and posture during CPR and delays in defibrillator pad placement. A recommendation was added in the instruction manual to promptly remove the SplashGuard CG, by cutting the ties with the scissors, in case of an acute emergency or need for CPR.After validation by the infection prevention unit, an arbitrary recommendation was made to change the SplashGuard CG every 24 hours in cases requiring high-flow nasal cannula or noninvasive ventilation and every 48 hours in other cases to avoid the adherence of viral particles to the inner surface of the device. This recommendation provides an appropriate time window for disinfection of the device with 0.5% hydrogen peroxide .An instruction manual, including a full description of the various predefined clinical scenarios, was edited for education and training of all members of the emergency, critical care, and anesthesia healthcare teams.Prior to implementing the SplashGuard CG within the clinical environment, we organized several clinical-based simulation sessions and used Rapid Cycle Deliberate Practice to train healthcare providers from the pediatric emergency, anesthesia, and pediatric critical care departments. This training program consisted of various simulation scenarios see  for an ith, 2020, three weeks after the start of the innovation process. The PICU staff agreed to provide noninvasive ventilatory support and high-flow nasal cannula for COVID-suspected and COVID-positive patients, if the SplashGuard CG was available. This eliminated the need for systematic intubation in several patients. The SplashGuard CG has also been used routinely in the ED for COVID-suspected or COVID-confirmed patients with respiratory symptoms to transport patients from the ED to the PICU and in the operating room for multiple intubation and extubation procedures.The SplashGuard CG was first authorized for clinical use in the PICU, the OR, and the ED, on April 7Several clinical research studies, evaluating the safety and efficacy of the SplashGuard CG, were developed in April 2020 and are still ongoing, in addition to the continuous evaluation by the different medical teams.The rapid development and implementation of a secure protective barrier enclosure, including clinically relevant features, were accomplished by a diverse research team including engineers, caregivers, and several other experts, in the context of the COVID-19 pandemic. The well-elaborated plan, ensuring close collaboration between interprofessional teams from different departments, and using Rapid Cycle Deliberate Practice for intensive simulation training, contributed to the success of this process. A user manual, instructional videos, and both laboratory and in situ simulations made the rapid development and implementation of our SplashGuard CG successful in less than a month.We acknowledge that there were several challenges encountered during the process. Multidisciplinary and interdisciplinary discussions on a regular basis were a key component to making this accelerated process succeed. However, finding time to meet in times of crisis with the necessary social distancing despite every team member having a full schedule was a major challenge to the process. There definitely was a need for a team leader to facilitate the planning and coordination of this process and to encourage every team member to participate. In addition, the rapidly evolving knowledge on the characteristics of SARS-CoV-2, as well as the experience of other centers implementing similar barrier enclosures, had to be taken in account during the development process.In a recent commentary in the New England Journal of Medicine, Rosenblatt et al. criticized the implementation of the “aerosol box” barrier enclosure described by Canelli , 17. EquThe question remains as to what degree the SplashGuard CG can protect against viral aerosols in addition to droplet protection. By attaching the plastic shield at the open side of the device, we avoid aerosols and droplets being redirected to the distal end of the bed, but as the plastic shield must be moved partially at some point in time to allow adequate patient care, there may be some exposure to the provider manipulating the plastic shield. For this reason, we made it possible to aspirate air within the SplashGuard CG to reduce contamination risks, as recommended by the FDA since August 2020 . If the The need for cleansing and disinfection of the device after use is mandatory, as all material and equipment in a patient's room is considered contaminated. We do not see this as a limitation, but precautions must be taken during disinfection as disinfection can create aerosolization.This study demonstrates that the accelerated design, development, and rapid implementation of a new device is feasible in times of crisis through close collaboration between medical and engineering teams and using repeated simulation sessions to test and improve the initial prototypes. Once the final prototype of our barrier enclosure “SplashGuard CG” was developed, the rapid dissemination of knowledge to all healthcare workers involved simulation-based training, including Rapid Cycle Deliberate Practice, and the use of supplementary means such as written user manuals, a website, and training videos. This simulation-based development and implementation of a new device seems particularly crucial during a pandemic situation.It should, however, be highlighted that following this rapid process, particular attention must be paid to maintaining team skills, monitoring the global evaluations of users, and any undesirable effects, all of which will contribute to continuous improvement of the device.


Approaches to furthering our understanding of the bioeffects, behavioral changes, and treatment options following exposure to blast are a worldwide priority. Of particular need is a more concerted effort to employ animal models to determine possible sex differences, which have been reported in the clinical literature. In this review, clinical and preclinical reports concerning blast injury effects are summarized in relation to sex as a biological variable (SABV). The review outlines approaches that explore the pertinent role of sex chromosomes and gonadal steroids for delineating sex as a biological independent variable. Next, underlying biological factors that need exploration for blast effects in light of SABV are outlined, including pituitary, autonomic, vascular, and inflammation factors that all have evidence as having important SABV relevance. A major second consideration for the study of SABV and preclinical blast effects is the notable lack of consistent model design—a wide range of devices have been employed with questionable relevance to real-life scenarios—as well as poor standardization for reporting of blast parameters. Hence, the review also provides current views regarding optimal design of shock tubes for approaching the problem of primary blast effects and sex differences and outlines a plan for the regularization of reporting. Standardization and clear description of blast parameters will provide greater comparability across models, as well as unify consensus for important sex difference bioeffects. Traumatic brain injury (TBI) is a significant military health problem, with the Defense and Veterans Brain Injury Center reporting ~384,000 worldwide cases from the years 2000 to 2018 in the US forces . Blast-rThe increasing participation of women in the US military and the lifting of the Combat Exclusion Policy in 2013, officially giving women eligibility to participate in full combat operations, have placed women at greater risk of sustaining a military-related TBI, including blast-related TBI. The number of women using the Veterans Administration in the United States increased by 46% between 2005 and 2015 , confirmAnimal models of TBI, including models of blast injuries, have aided in our understanding of the pathophysiology and symptomology of brain injuries for decades . SinThe purpose of this review is to discuss TBI, particularly as inflicted by a blast event, in the context of sex as a biological variable (SABV). What are known about the functional consequences of military-acquired blast TBI are discussed, followed by a description of the present experimental approaches that have had utility in manipulating sex chromosomes and gonadal steroids as independent variables. Dependent variables, including pituitary, autonomic, vascular, and inflammation factors, are then discussed because these focus on the most reported systems that are perturbed by blast. Finally, relevant to the study of blast and SABV in preclinical studies, the review makes an appeal for investigators to apply the highest-quality experimental principles, because the study of SABV in this field is complex and requires the derivation of the uppermost-quality information for asking further questions and laying the groundwork for translational relevance. Suggestions and guidelines are provided for the use and reporting of sufficient information about blast animal models that will aid in the interpretation of data and generation of conclusions.Mild TBI (mTBI) as a result of an explosion often leads to symptoms that are well-studied in military populations. The symptoms are most often short-term, resolving within 7–10 days, and often include physical , cognitive , and behavioral  complaints , 42. HowIn addition to potential long-term symptoms following mTBI, there is a clear link between military-acquired TBI and PTSD e.g., , 45, 46], 4645, 4Blast Events), making the contributions of the primary blast wave to the subsequent outcomes difficult to clearly establish , vascular changes are noted in clinical and preclinical studies of blast effects. Sullivan et al.  to assess cerebral blood flow as a correlate of cerebral activity. Han et al.  found bln et al.  applied n et al. .Several trends, then, are gained from MRI. First, alterations in white matter integrity have been observed, and the findings suggest these cases exhibit diffuse changes and variability in the location of changes , 65, 66,Research on military health and blast-induced brain injuries has largely focused on males, as they have historically made up the majority of service members of the US military and have been more likely to occupy combat roles. However, ~300,000 female service members were deployed to Iraq and/or Afghanistan between 2001 and 2013 , and theOverall, following combat-related mTBI, women are likely to suffer the same symptom clusters as their male military peers, such as PCS, PTSD, psychiatric complaints , and somatic symptoms  –19, 45. In addition to depression, there is evidence that female service members may have increased susceptibility to anxiety disorders and/or PTSD following mTBI. The literature describing gender differences in PTSD symptoms in military personnel is relatively broad and reports mixed results, possibly as a result of variations in the definition of TBI or methodological differences . In an eBecause of the complexity of ascertaining relevant variables for blast mTBI etiologies, it is important to supplement the clinical literature by applying preclinical animal research. To further understand the role of sex-related variables as proximate causes for sex differences, animal modeling enables greater control of conditions and the ability to more invasively explore cellular response mechanisms from blast exposure. Following an overview of approaches to the study of SABV as an independent variable for blast TBI preclinical work, there is a summary of what is presently known concerning blast bioeffects on pituitary and the hypothalamic–pituitary–adrenal (HPA) axis, the autonomic nervous system, the vasculature, and inflammation.In addition to the NIH mandate regarding consideration of SABV in clinical and preclinical research, a second policy relates to scientific rigor by employment of preclinical experimental practices that derive valid and reliable findings to adequately address research gaps, set the stage for discovering important mechanisms underlying sex difference and properly modeling translational testing –83. AccoHPA Axis Dysfunction in Laboratory Animals After Blast. The review by Rubin and Lipton  or the paternal X chromosome (XP) is silenced by X chromosome inactivation (XCI) to partially rebalance the level of expression  an XY complement with the Sry gene for XY mice with testes; (2) an XY complement without Sry, resulting in XY mice with ovaries; (3) an Sry mouse with the gene incorporated into an autosome resulting in an XX mouse with testes; and (4) an XX mouse with no copy of Sry, resulting in XX mice with ovaries. The mice with similar gonadal forms then permit investigation of the sex chromosome complementation (XX vs. XY) in the context of gonad-related hormonal status , gonadal steroid action has been a main, classic focus for the study of sex differences. This is particularly relevant because there is strong evidence of neuroprotective roles of estrogen and progesterone after a range of brain injuries . Becker et al.  following gonadectomy in the first step as well as after hormone replacement in the second phase of the study. Experienced investigators suggest that the administration of gonadal steroids should be monitored to ensure the replacement procedure provides levels of steroid within the physiological range. Further studies can employ the same approach with compounds that block steroid synthesis or that disrupt steroid receptor effects, or to determine the role of clinically relevant intervening effects on steroid action such as contraceptives. Maintenance on the contraceptives, desogestrel and drospirenone, e.g., was found to reduce the severity of stroke neuropathology in ovariectomized mice . FinallyA second level of inquiry relates to blast-related mechanisms—the dependent variables—that may be differentially affected by sex differences. Outlined below are the most salient biological effects known to date for the impact of blast exposure relevant to SABV. The discussion for some effects begins with clinical descriptions, but some of the reportage concerns findings with non–blast-related methods that may help point to relevant effects, including sex-relevant differences of pituitary and HPA axis function, and blast effects on the autonomic nervous system function, the vasculature, and inflammation. Although researchers often focus on a single dimension of outcome, investigators have recognized that TBI consequences demonstrate it manifests as a systemic condition , 112. LiAlthough more studies of military-acquired, particularly blast-related, mTBI in women are required, a picture is emerging of a gender dichotomy in the stress response following mTBI. There are clear sex differences in non-TBI civilian populations in the lifetime susceptibility to depression and anxiety disorders , 114, asThe primary stress hormone is cortisol , which is released by the HPA axis when activated by a physical or psychological stressor. The stress response is characterized by release of corticotropin-releasing factor (CRF) from the paraventricular nucleus (PVN) of the hypothalamus, which binds to CRF receptors on the anterior pituitary gland. The anterior pituitary gland secretes adrenocorticotropic hormone (ACTH) into the bloodstream, where it reaches the adrenal cortex and binds to receptors to stimulate the synthesis and release of the steroid hormones, glucocorticoids , and mineralocorticoids . Steroid hormone receptors are located throughout the brain ,First, the authors employed a restraint-induced stress model and demonstrated that while blast injury increased the restraint-induced rise in CORT levels in males, the opposite effect was observed in female mice, with blast attenuating CORT levels in restrained animals compared to sham-treated mice. Blast did not alter CORT suppression in the dexamethasone-suppression test or affect the expression of pituitary or adrenal genes involved in ACTH or CORT synthesis or secretion, suggesting a central disruption in feedback, rather than a peripheral effect, as the more likely source of the sexually dimorphic response to injury. Examining potential central nervous system sources, it was first determined that there were no effects of blast injury in either males or females on mRNA expression of mineralocorticoid and glucocorticoid receptors at central feedback regulation sites: the PVN or other brain limbic structures . However, a restraint-induced increase in CRF neuron activation was differentially altered by blast injury in male and female mice: in males with restraint treatment, blast (compared to sham treatment) reduced CRF neuron activation in the PVN; in females, restraint-treated mice receiving blast treatment had increased levels of CRF neuron activation in the PVN. Retrograde tracing determined that there was a TBI-related decrease of CRF neurons in female mice primarily in preautonomic (non-neuroendocrine) neurons in the PVN, suggesting a decreased use of the preautonomic system in dealing with stressors, leading to a possible blast-induced disruption in CRF outputs to brainstem structures regulating autonomic function. There were no blast-induced changes in the percentage of activated CRF neurons that were endocrine projecting or preautonomic projecting in male mice, and the authors hypothesized that disruption in limbic structures of the HPA axis may result from blast-induced TBI.A second study was designed to measure changes in the expression of CRF receptor subtypes 1 and 2  in limbic structures following blast-induced brain injury in male and female mice, as well as to assess the sex-dependent effects of blast on anxiety-like behaviors . CRFR1 iDuring the acute period following TBI, systemic effects appear to result from excessive catecholamine release and subsequent autonomic dysfunction. In more severe cares, autonomic dysfunction leads to transient episodes of paroxysmal sympathetic hyperactivity (PSH), which includes tachycardia, hypertension, hyperthermia, spasticity, and tachypnea –147. A rEvidence for persistent cardiovascular changes after milder cases of TBI has been reported, with alterations in cardiac rhythm variability providing an overall, integrated indicator of autonomic function –156. In Autonomic Nervous System Function After TBI) and further trauma with leukocyte recruitment and the release of proinflammatory signals [, vascular alterations from blast exposure have received the greatest research attention. A particularly vulnerable organ to blast exposure is the pulmonary system, where more energetic shock waves result in significant lung contusions and accompanying autonomic dysregulation . The initiative got underway from a Workshop for the Classification of TBI for Targeted Therapies held in October 2007, by the National Institute of Neurological Disorders and Stroke, with the participation of representatives from other groups, including the Defense and Veterans Brain Injury Center and the National Institute on Disability and Rehabilitation Research. The working group initially focused on the limitations of diagnostic criteria and that a pathoanatomical classification system could be the springboard for addressing the heterogeneity of TBIs and for improved systemization for clinical studies and trials  and 41 elements in “Module 6,” specific to blast/shock studies  or compressed gas as the driver, must be recognized for its complexity of model application, the inherent pitfalls in each model, and the onus for understanding and communicating exposure metrics. A detailed description of the design of the experimental setting should be documented, including the dimensions of the free-field conditions or the shock/blast tube, for investigators to have a clear sense of the exposure conditions. A published description of the environment becomes a permanent record of intervening effects permitting meta-analyses through standard reporting. Full reportage also allows documentation of existing or potential experimental artifacts, such as reflective and blockage effects from the surroundings  and potential constraining factors such as shock tube dimensions, the size, location, orientation, fixation/restraint conditions of the study specimen(s), and exposure conditions of the test specimen(s) in relation to the overpressure source , 219. This ripe to get things right. As noted by the Traumatic Brain Injury Preclinical Working Group, preclinical blast research studies are presently an “immature research area” where the lack of more substantial clinical information as a guide to basic hypothesis-driven research is a challenge; the causal effects following blast are emerging but still uncertain, and there are no commonly accepted injury devices (For the role of sex in underlying biological responses, the field of laboratory blast studies  devices , as is s devices . Rigor aAll authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.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.


Enteroatmospheric fistula (EAF) after open abdomen adds difficulties to the management and increases the morbidity and mortality of patients. As an effective measurement, reconstructing gastrointestinal tract integrity not only reduces digestive juice wasting and wound contamination, but also allows expedient restoration of enteral nutrition and intestinal homeostasis. In this review, we introduce several technologies for the temporary isolation of EAF, including negative pressure wound therapy, fistuloclysis, fistula patch, surgical covered stent, three-dimensional (3D) printing stent, and injection molding stent. The manufacture and implantation procedures of each technique with their pros and cons are described in detail. Moreover, the approach in combination with finger measurement, x-ray imaging, and computerized tomography is used to measure anatomic parameters of fistula and design appropriate 3D printer-recognizable stereolithography files for production of isolation devices. Given the active roles that engineers playing in the technology development, we call on the cooperation between clinicians and engineers and the organization of clinical trials on these techniques. The open abdomen therapy can be chosen for severe intraabdominal infections and abdominal compartment syndrome (ACS) when no other perceived options exist. This strategy allows surgeons to carry out source control procedures on unexplored abdominal infections and reduce intraabdominal pressure for the prevention of visceral organ ischemia . Early cEAF requires comprehensive treatments: (1) nutritional support, among which the early total parenteral nutrition is beneficial for intestinal rest and spontaneous fistula closure; (2) somatostatin analogs, which reduce gastrointestinal (GI) secretions and allow fast fistula closure, but do not reduce the mortality. Most of the studies agree that the greatest benefit occurs in the first 10 days of treatment , 14; (3)NPWT has been widely used for TAC in clinical practice. Firstly, the skin necrosis and any other necrotic tissues need to be debrided. Secondly, an obligatory non-adherent polyvinyl alcohol membrane serves as the first layer of NPWT over the intestinal loops and a piece of white sponge as the second. The non-adherent layer can effectively prevent the adhesion between the intestinal loops and the sponge. It is worth noting that the sponge needs to be tailored slightly smaller than the size of abdominal wound to leave enough space for abdominal wall traction. If there are any skin folds, stoma and drainage tubes, the stoma pastes or silicone gels are required to make the entire negative pressure system sealed. Finally, the adhesive drape is placed over the sponge with the margins of intact skin. An external negative pressure is chosen ranging from −100 to −125 mmHg to drain the intestinal fluids according to their output, the number of EAFs, the amount of NPWT, and the process of wound healing . The whoAlthough treated with NPWT, the spontaneous closure of EAF is very rare and depends on the fistula location and output , 23. OncManagement of EAF requires sufficient nutritional and metabolic supports by the means of gradual transition from parenteral nutrition to enteral nutrition. Long-term parenteral nutrition may lead to complications such as the catheter-related bloodstream infections, liver injury, and intestinal dysfunction, while enteral nutrition can improve those conditions. As an enteral nutrition routine, fistuloclysis can be used to infuse enteral nutrients, formula, or proximal GI secretions to the distal limb of fistula in order to improve the nutritional status and maintain fluid/electrolyte homeostasis , 26. FigHowever, fistuloclysis has been less popular in recent years mainly due to technical and aesthetic concerns . Some EACompared with fistuloclysis, intraluminal occlusion of EAF is more advantageous since it supports the physical integrity of GI tract and ensures the pass of digestive contents with less leakage from EAF. Fistula patch is the first-generation device that addresses this issue. As shown in Notably, the patch can only be applied to the EAFs with mucosal protrusion and needs to be tailored in accordance with their anatomic characteristics. More insights have been put on the fistula stent that can be made tubularly and conforms to the shape of intestinal tract. As shown in 1, L2) and their resulting angle (β) in order to design a suitable stereolithography (STL) file based on the measurements  technology has shown higher sensitivity in distinguishing GI fistula from soft tissues , 41, andIn this review, we introduced several isolation approaches for the management of EAF after open abdomen including NPWT, fistuloclysis, fistula patch, surgical covered stent, 3D printing stent, and injection molding stent. The choice of these approaches should consider the condition of EAF, general body habitus, and the treatment purpose. The fistula stent is a new solution with promising functions in the maintenance of the GI tract physical integrity. The cooperation between surgeons and engineers is advocated to promote the improvement and application of these techniques.JR and XW conceived this review. JH organized literatures and wrote this review. HR organized the photographs and the table. YJ participated in the discussion. All authors contributed to the article and approved the submitted version.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.


Understanding the patterns of chemical exposure among biota across a landscape is challenging due to the spatial heterogeneity and complexity of the sources, pathways, and fate of the different chemicals. While spatially-driven relationships between contaminant sources and biota body burdens of a single chemical are commonly modelled, there has been little effort on modelling chemical mixtures across multiple wildlife species in the Canadian Oil Sands region. In this study, we used spatial principal components analysis (sPCA) to assess spatial patterns of the body burdens of 22 metals and Potentially Toxic Elements (PTEs) in 492 individual wildlife, including fur-bearing mammals, colonial waterbirds, and amphibians collected from the Canadian Oil Sands region in Canada. Spatial analysis and mapping both indicate that some of the complex exposures in the studied biota are distributed randomly across a landscape, which suggests background or non-point source exposures. In contrast, the pattern of exposure for seven metals and PTEs, including mercury, vanadium, lead, rubidium, lithium, strontium, and barium, exhibited a clustered pattern to the east of the open-pit mining area and in regions downstream of oil sands development which indicates point-source input. This analysis demonstrated useful methods for integrating monitoring datasets and identifying sources and potential drivers of exposure to chemical mixtures in biota across a landscape. These results can be used to support an adaptive monitoring program by identifying regions needing additional monitoring, health impact assessments, and possible intervention strategies. Th. Th38]. We then used the environmental fitting (envfit: vegan package ) to asseThe BCA results comparing the two data transformation methods (square-root and log base 10) in combination with the range-normalization method are shown in The visual patterns observed in The square-root transformed range-normalized data were fully interpreted using MEM to assess hotspots of complex metal exposure. The biplot from this analysis , and theThe first four components explained a total of 82.2% of the variance, which was attributed to positive spatial autocorrelation (self-similarity over space). In component 1, the positively loaded metals and PTEs were mercury, zinc, and thallium . The patThe positively loaded metals and PTEs in component 2 were rubidium and bismuth which had a dispersed pattern across the OSR . The negComponent 3 was positively loaded with mercury, vanadium, and cadmium. This distinct pattern is observed around Fort McMurray, Alberta and extends northward. West of the OSR, sites are negatively loaded with barium, silver, and bismuth . ComponeThe environmental fitting results in This paper demonstrates the application of methods for combining complex contaminants monitoring datasets comprised of multiple species and life stages. Range normalizing data is an important step that allows datasets to be combined and assessed for overarching spatial patterns of metal and PTE exposure, as demonstrated in Missing data is problematic for statistical methods, as it can limit analyses that can be used . The simThis paper also presents the application of a novel method for assessing the spatial patterns of multidimensional ecotoxicology data and builds on our previous work using the MEM method to assess spatial patterns of complex metal exposure in adult wood frogs in northern Alberta . The resThe results of this research demonstrate how the traditional PCA method can be expanded to include spatial structure into the analysis of ecotoxicological data. In this analysis, component 1 identified hotspots of mercury, rubidium, and thallium exposures. The mapped positive scores are similar to the locations of gull and amphibian data from northern Alberta. Thus, this spatial pattern may be related to a species difference, rather than differences in metal and PTE body burdens between sample locations. However, these scores also exhibit a latitudinal gradient, where the scores downstream of oil sands development are larger than scores upstream. This matches patterns of mercury exposures observed in the gull and tern eggs in northern Alberta , 11. FurHotspots of lead, vanadium, nickel, lithium, and strontium (component 2), and mercury, vanadium, and cadmium (component 3) clustered around the open-pit mining and upgrading operations. These elements also had similar directionality on the ordination plot with an environmental variable quantifying the percentage of mining in the local environment. Local enrichment of these metals and PTEs from oil sands operations has been widely reported , 20, 50.Some components had no apparent clustering pattern. The most highly loaded metals and PTEs in these components included arsenic and iron, and a variety of other crustal elements including bismuth, strontium, rubidium, cobalt, and lithium, The mapped components that exhibit a random pattern are likely indicative of background metal and PTE exposures. In the environmental fitting ordination plot, these elements clustered with intact landscapes and deciduous and mixed forests. Northern Alberta has very complex underlying natural geological and geochemical properties. For example, monitoring data shows that northern Alberta has arsenic levels up to 10 times higher, and iron levels up to 100 times higher than background concentrations in other regions of Alberta, which is due to the naturally occurring shale deposits , 52. TheThere are other sources that could contribute to the overall variance in the complex metal and PTE exposures in the biota. First, while the range normalization method attempted to address some of the issues associated with integrating contaminant data across different species and life stages, this method was not able to completely diminish the effect of species and life stage. This may be due to the bioaccumulative nature of some metals and PTEs with the age of the biota . In thisOil Sands development is changing how wildlife uses the landscape, population densities, and species interactions both during the process of development and reclamation , 55. ChaSpatial analyses of wildlife exposures can be used to identify priority metals and PTEs, determine sources, identify priority areas for further study, and help develop targeted strategies for additional monitoring or exposure reduction . HoweverIn this paper, we demonstrate the application of an sPCA method and address current limitations of spatially assessing landscape-level complex exposures across different species and life stages. The methods we used to transform and normalize the data diminished the effect of species and life-stage differences in contaminant burdens and is a useful pre-processing step for data integration. Results show that patterns observed in the sPCA were related to patterns of environmental exposures and not a difference in species-specific accumulation rates. The spatial analysis conducted in this study demonstrates that some metals and PTEs are being anthropogenically delivered and concentrated around oil sands development. For example, levels of mercury, vanadium, lead, rubidium, lithium, strontium, and barium were elevated in multiple species around the open-pit mining area as well as regions downstream of oil sands development. The techniques presented in this paper can be applied to monitoring programs in other areas undergoing development and anthropogenic change. We also highlight areas of further research, which include the need to evaluate a broader spectrum of contaminants, including PACs, and how anthropogenic changes to the landscape result in altered contaminant burdens in wildlife.S1 FigA comparison for each combination of transformation and normalization method and the effects of life stage differences.(TIF)Click here for additional data file.S2 FigThese data have been square-root transformed and range normalized.(TIF)Click here for additional data file.