Individual cell types have characteristic sizes, suggesting that size sensing mechanisms may coordinate transcription, translation, and metabolism with cell growth rates. Two types of size-sensing mechanisms have been proposed: spatial sensing of the location or dimensions of a signal, subcellular structure or organelle; or titration-based sensing of the intracellular concentrations of key regulators. Here we propose that size sensing in animal cells combines both titration and spatial sensing elements in a dynamic mechanism whereby microtubule motor-dependent localization of RNA encoding importin β1 and mTOR, coupled with regulated local protein synthesis, enable cytoskeleton length sensing for cell growth regulation. Size is a fairly uniform characteristic for any given cell type, and the reasons for this deceptively trivial observation have been vexing science since deep in the previous millennium . Size coEarly work in yeast and animal cells provided evidence for size sensing, with observations of non-linear growth rates and size-dependent fluctuations in growth duration between division points , 9. HoweDespite accumulating evidence for size sensing capability in different cell types, the molecular details of such a mechanism are not well understood. Yeast cells have been most intensively studied in this regard, and two classes of size-sensing models have been proposed—titration-based measurements versus spatial sensing. Titration-based mechanisms postulate that increases or decreases in levels of a key signal provide a critical checkpoint size signal. A recent study in fission yeast demonstrated size-dependent expression of the mitotic activator Cdc25, and suggested that size-dependent increases in Cdc25 levels trigger cell division upon reaching a threshold concentration . An alteIn the second class of models, subcellular localization of key signals provides size readouts to the cell. For example, in fission yeast the proteins Pom1 and Cdr2 have been proposed as components of such a mechanism, wherein Pom1 is transported to cell tips and diffuses from there to form longitudinal gradients along the cell, while Cdr2 is localized to large immobile structures at the plasma membrane in the cell middle, termed cortical nodes . ConflicCaenorhabditis elegans intestine demonstrated a direct proportionality of nucleolus size to both cell and whole-body size throughout worm development [Other types of spatial measurements might also provide proxies for size sensing, such as monitoring the sizes of key organelles within a cell. Nuclear size is the most well-studied example, and the karyoplasmic ratio describes the tight nuclear/cytoplasm size relationship in almost any cycling cell type . Intriguelopment . Centroselopment –39; thuselopment .Correlations of overall microtubule cytoskeleton dimensions with cell size raises the possibility of using cytoskeleton length as a proxy measurement for size sensing, thus simplifying the three-dimensional challenge of size sensing to the single dimension of length measurement . MicrotuWe looked into the possibility that active transport by the microtubule motors dynein and kinesin coordinates length sensing, using neuronal axon length as a model system. Axons comprise the largest compartment of a neuron; hence, axon length provides a proxy for overall neuronal size. Moreover, the distributed morphologies and large sizes of neurons can be advantageous in studies of compartmentalized signaling and size sensing . Dynein An experimental test of motor-dependent oscillatory signaling for axonal length sensing was suggested by simulations showing that reducing levels of either kinesin or dynein should slow frequency decay of the retrograde signal . If neurA follow-up study then examined a number of mouse mutants for axon-lengthening phenotypes similar to those observed upon microtubule motor knockdown, and identified such a phenotype in a mouse with an importin β1 3′ UTR deletion . Both adDisruption of the interaction between nucleolin and kinesin using AS1411, a nucleolin-targeted DNA aptamer, sequestered nucleolin from sensory axons and caused robust increases in axon growth . SimilarPerturbation of nucleolin or importin β1 localization shifts the balance of protein synthesis between the periphery and the center of the cell . This fiThe text above, summarized in Fig.


Salmonella and Escherichia coli O157:H7. These findings are worthy of further study, especially when coupled with the enhanced growth and performance observed with these products. Mechanistically, XPC appears to modulate these effects through the immune system and gut microbiome. Herein we further investigated this product and demonstrate that XPC mediates an enhancement of immunocyte killing of Salmonella in calves fed the product. Additionally, these studies reveal that XPC reduces the lymph node infiltration, invasiveness, and antibiotic resistance of Salmonella in dairy calves fed the product-consistent with findings observed in poultry and adult beef cattle. Furthermore, the reduction in invasiveness does not lead to a rebound hyperinvasive phenotype in Salmonella obtained from XPC-fed animals. In summary, these studies suggest that XPC reduces the invasion of Salmonella and may alter various phenotypic characteristics of the pathogen.Prior studies revealed that yeast fermentation products, specifically XPC™ and related products , serve as viable food safety tools across multiple food animal species including cattle and poultry. Providing this supplement in feed leads to reduced prevalence, load, virulence, and antibiotic resistance of foodborne pathogens such as  Salmonella enterica and shiga toxin-producing Escherichia coli serotypes such as O157:H7 (STEC), are serious concerns in food animal production management.The presence of important foodborne pathogens continues to prove a formidable challenge to overcome. Two important foodborne pathogens, Saccharomyces cerevisiae fermentation product designated as XPC™, appears to impact enteric pathogens across multiple livestock species  were fed daily doses of XPC plus SmartCare for 2 weeks as part of a previous study . The erythrocyte fraction was then removed and 120 μL of the buffy coat interface was collected and aliquoted into six separate tubes containing RPMI media, to which 107 colony-forming units of statically grown Salmonella  were added and the tubes were incubated at 37°C. After 1 h, extracellular  bacteria were killed by the addition of 50 μg/mL gentamicin. At 0, 1, 2, 4, 8, and 12 h post-killing, leukocytes were centrifuged and the gentamicin-containing media was removed and replaced with 50 μL of phosphate-buffered saline containing 1% Triton which lyses the leukocytes. Lysates were then plated on XLD agar that was incubated overnight at 37°C. The following day, black-centered colonies were enumerated and Salmonella survival/cell was calculated as number of recovered colonies/number of leukocytes per 20 μL blood . The strains used were gentamicin-susceptible S. Dublin SGI1 . Antibiogram assays focused on florfenicol resistance since this resistance is encoded on an integron present in the input strain. That is, the studies presented herein assessed the ability of XPC to reduce both the invasiveness and integron-mediated antibiotic resistance of Salmonella obtained from dairy calves from a prior study  of florfenicol divided by the number of colonies assayed. Percent containing SGI was derived as the number of colonies that were PCR(+) for the SGI1 integron divided by the number of colonies assessed.y calves  were suby calves , 3, i.e.y calves  and micror study . Studiesor study . InvasioSalmonella obtained from cattle . For data with a single measurement, an analysis of variance (ANOVA) was conducted with the Tukey's ad hoc test for multiple comparisons.For data with multiple sampling time points, a repeated measures analysis was used with the Bonferroni Salmonella fecal shedding and intestinal carriage in calves fed the products, when compared to calves fed the Control diet. To assess the possibility that the leukocytes from XPC-fed calves are clearing the Salmonella, peripheral blood cells were examined for the ability of the leukocytes to temporally reduce the presence of Salmonella. As shown in Salmonella was equivalent in both groups of calves  at the first time point but the numbers of Salmonella dropped significantly (50–75%) at each successive time point for leukocytes obtained from XPC-fed calves when compared to Control-fed calves. Importantly, this mechanism seems to be conserved across two divergent Salmonella enterica serovars- Typhimurium and Dublin.Our previous studies with SmartCare and XPC  revealedSalmonella to lymph nodes, peripheral lymph nodes  were excised at euthanasia and subjected to selective culture for Salmonella. As shown in Salmonella was significantly less abundant (~80% reduced) in lymph nodes from XPC-fed calves when compared to Control-fed calves.To address the possibility that these calves were more efficient at translocating the Salmonella recovered from the calves. As shown in hilA expression, antibiotic resistance, and prevalence of the antibiotic resistance element (SGI1) were all significantly diminished (~60–70% reductions) in Salmonella recovered from XPC-fed calves.Since the input strain used in the poultry studies  was alsoSalmonella and the microbe will return to the fully invasive state or even become hyperinvasive after exiting an XPC-fed animal, a series of temporal invasion assays were performed on isolates recovered from XPC-fed poultry and cattle. As shown in hila expression were ~70–80% reduced after recovery from animals (“Generation)” and then returned to baseline after three successive incursions (designated as “Generations”) into tissue culture cells. Neither the invasiveness nor the hila expression went above the baseline observed for Salmonella recovered from Control-fed animals or from the wild-type strain. Thus, it appears that, after exiting an XPC-fed animal, the Salmonella need to invade three additional and subsequent hosts before returning to the fully virulent state as per the wild-type strain. These data include Salmonella obtained from the experimentally infected dairy calves  then provided editorial comments that culminated in the final version of the manuscript.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.


Research in the field of relation between microbes and colorectal carcinogenesis has gained increasing interest in past years. Recently, link between microbial biofilm and carcinogenesis in colon was demonstrated by several authors indicating that biofilm not only is a key player in carcinogenesis, but also may contribute to the understanding of side-specific colon cancer—right sided colon cancer versus left sided. In this article, we briefly highlight the major findings of the research of biofilm and carcinogenesis and demonstrate our findings of colonic cancer tissue and colonic polyp examined for biofilm.Colonic cancer tissue from a patient with a right-sided colon cancer, and an adenoma tubular polyp were examined for biofilm formation by flourescens in situ hybridization. In cancer tissue we found biofilm formation on the surface epithelium but surprisingly also deep into the crypts. No biofilms were found in tubular polyp tissue.To our knowledge, this is the first-time biofilm formation deep into colonic crypts are demonstrated in a patient with right-sided colon cancer. This may indicate that bacterial biofilm may have a key role in carcinogenesis. Bacteroides fragilis (ETBF), Fusobacterium nucleatum, colibactin-producing Escherichia coli (CPEC) and Porphyromonas, damage mucosal barrier functions by degrading mucins, damaging intercellular junctions and cellular DNA. Direct bacterial actions and immunological responses may induce dysplasia and carcinogenesis [Interactions between the human host and the microbiota is in focus as accumulating data point towards important links in colorectal carcinogenesis. Experimental data show how potential colorectal cancer (CRC) driver bacteria such as enterotoxigenic ogenesis , 2.A recent study from Dejea et al. report colonization of aggregated bacteria (biofilms) consisting of ETBF and CPEC on the colonic epithelial surface in patients with familial adenomatous polyposis coli (FAP) . The adeDejea’s data suggest that the colonization with ETBF and CPEC, found in more than 50% of the FAP patients examined (in contrast to less than 25% of controls), degrades mucins, destroys cadherins, increases stem cell proliferation through the Wnt-pathway and thereby increasing risk of mutations and CRC in early life.The bi-layered mucus lining covering the colonic epithelium is the first line of defense separating microbes from epithelial cells and more importantly keeping them away from proliferating stem cells and progenitors (transit amplifying cells) in the lower crypt compartment. The mucin filling the colonic crypts contains antimicrobial peptides such as defensins, cathelicidins and IgA produced by crypt cells and submucosal immune cells. Furthermore, crypt entrances are guarded by sentinel goblet cells ready to flush microbes away by emptying their entire loads of mucins and cytoplasmic contents by exocytosis when the recognition receptors are triggered by pathogen-associated molecular patterns.The main constituent of the gel-forming mucus layer is the MUC2 protein secreted by goblet cells. Glycosylated MUC2 mucin molecules form a double-layered mucus membrane of multiple, large insoluble net-like structures. The innermost mucus layer covering the epithelial cell surfaces and filling the crypts is densely packed and impermeable to bacteria. This layer is anchored to the epithelium by goblet cells and by trans-membranous mucins on the apical surface of enterocytes .As mucin production pushes mucus luminally, it is exposed to proteolytic activities by the host and by commensal bacterial proteases and glycosidases. The mucus breaks up and expands 2–3 times in volume providing an ideal niche for bacterial colonization and at the same time serving as a giant glycan food source for microbes . This ouThus, during eubiosis the inner mucus layer and the colonic crypts are free from microorganisms. Bacterial biofilm formation in the inner mucus layer may down-regulate MUC2 mucins and production of defense peptides. This may result in direct contact between bacteria and epithelial cells, which is considered pathological .Dejea et al. report findings of bacterial biofilm formation on the surface of colonic epithelium in nearly all (89%) examined cases of right-sided colon cancer, whereas only a few percent (12%) of the left-sided cancers were biofilm positive. There was 100% concordance between biofilm-positivity on tumor tissue and the colon tissue at the resection margin. A subsequent metabolomics analysis revealed that biofilm formation in the colon significantly contributes to a pro-oncogenic state .Except from the stem cells and progenitors in the lower crypt compartment, differentiated crypt cells  migrate towards the surface where they are exfoliated. Stem cells and progenitors constitute a dynamic cell population exhibiting plasticity by transitioning between stem and non-stem cell function, heavily influenced by the micro-environment and tissue damage. Stem cells and progenitors can accumulate genetic and epigenetic alterations resulting in increased self-renewal, survival pressure and dysplasia.We examined colonic tissue from a patient with a right-sided colon cancer and as expected we found biofilm formation on the surface epithelium but surprisingly also deep into the crypts Fig. . In addiA conventional right-sided hemicolectomy was performed on a patient with an adenocarcinoma in the right colon. Tissue samples were obtained from resection margins. No preoperative oncological treatment or antimicrobial therapy. An adenoma tubular polyp from right colon was removed in routine colonoscopy. Samples were fixed in 4% buffered paraformaldehyde pH 7.4), embedded in paraffin and then cut, deparaffinized and stained with PNA-FISH-Tamra5-conjugated universal bacterial (BacUni) 16S rRNA probe . Slides were counter-stained with 3 µM 4′,6-diamidino-2-phenylindole (DAPI) , mounted  and cover-slipped . Slides were examined by CLSM . Images was captured using 405 nm (DAPI) and 561 nm (Tamra-5) lasers, as well as a 488 nm for capturing the green auto fluorescence contour of the surrounding tissue as described elsewhere [.4, embedFusobacterium Nucleatum can impact the efficacy of conventional chemotherapy and checkpoint inhibitors. Biofilms in the right colon may explain some of the differences in treatment outcomes between right-sided and left-sided colon cancer. Current research on biofilm inhibitors and quorum sensing inhibitors may prove valuable in developing new drugs for future prevention and treatment of CRC.To our knowledge, this is the first-time biofilm formation deep into colonic crypts are demonstrated in a patient with right-sided colon cancer. In theory, degradation of mucin by pathogens results in biofilm formation on the epithelial surface, invasion of potential driver bacteria into proliferating crypt compartments, adverse immune responses, accelerated proliferation and increased risk of deleterious mutations. Thus, biofilm invasion deep into colonic crypts would be expected to accelerate carcinogenesis. Also, bacteria like ETBF and


Research on the aryl hydrocarbon receptor (AhR) has largely focused on its activation by various environmental toxins. Consequently, only limited inferences have been made regarding its constitutive activity in the absence of an exogenous ligands. Evidence has shown that AhR is constitutively active in advanced prostate cancer cell lines which model castration resistant prostate cancer (CRPC). CRPC cells can thrive in an androgen depleted environment. However, AR signaling still plays a major role. Although several mechanisms have been suggested for the sustained AR signaling, much is still unknown. Recent studies suggest that crosstalk between constitutive AhR and Src kinase may sustained AR signaling in CRPC. AhR forms a protein complex with Src and plays a role in regulating Src activity. Several groups have reported that tyrosine phosphorylation of AR protein by Src leads to AR activation, thereby promoting the development of CRPC. This review evaluates reports that implicate constitutive AhR as a key regulator of AR signaling in CRPC by utilizing Src as a signaling intermediate. Androgens and androgen receptor (AR) signaling play a predominant role in male sexual development, growth of the prostate gland and progression of prostate cancer to CRPC2. In CRPC, AR signaling has been shown to be sustained by a variety of mechanisms including increased androgen uptake by prostate cancer cells, increased AR expression, AR gene mutation and activation by other transcription factors4. The aryl hydrocarbon receptor is a transcription factor that has been extensively studied for its role in mediating the toxic effects of a wide range of environmental toxins. However, recent evidence has shown that AhR possess intrinsic functions independent of activation by an exogenous ligand. Constitutively active AhR has been shown to interact with hormone receptors and may play a role in progression of hormone-related cancers. Particularly, constitutively active AhR interacts with AR and act as a functional transcription unit5, a mechanism that may lead to enhanced AR signaling in CRPC.Most men who die of prostate cancer present with castration resistant prostate cancer (CRPC)6. As an inactive complex, AhR is found in the cytosol where it interacts with tyrosine kinase c-Src as well as two molecules of HSP90, co-chaperone p23 and immunophilin-like AhR interacting protein (AIP/XAP2)8. In its active form, AhR disassociates from its chaperone proteins and dimerizes with the aryl hydrocarbon receptor nuclear translocator (ARNT). The heterodimer activates gene expression through interactions with related xenobiotic responsive elements (XREs) located on Ah- responsive gene promoters9. Studies concerned with the intrinsic functions of AhR have found that overexpression of the receptor may promote carcinogenesis in the absence of a exogenous ligand. AhR protein and mRNA expression is associated with phases of rapid proliferation and differentiation in certain tissues. Conversely, AhR-defective cell lines demonstrate a reduced proliferation rate10. Ectopic over expression of AhR in immortalized normal mammary epithelial cells induced a malignant phenotype with increased growth and acquired invasive capabilities11. A separate study using a constitutively active AhR construct lacking a ligand binding domain revealed that AhR acts as a transcriptional coregulator for the unliganded AR. These studies show that the endogenous AR along with the constitutively active AhR were recruited to androgen-responsive elements to initiate signaling in an androgen depleted environment5.Although AhR has been extensively researched for its role as a xenobiotic receptor; RNA interference, overexpression, and inhibition studies suggest a role for AhR in multiple tumor types beyond activation by environmental contaminants9 in both constitutive and ligand activated signaling. AhR ligands such as 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD), components of cigarette smoke such as benzo(a)pyrene (BaP), and a wide range of polycyclic aromatic hydrocarbons (PAH) exert their biological influences by binding directly to the AhR. Following ligand activation, AhR’s primary role is identified as the control of xenobiotic metabolism through cytochrome P450s, some of which are transcriptional targets of AhR13. Activation of AhR by PAHs has been reported to antagonize AR signaling. For example, TCDD has been shown to alter sex steroid hormone secretions. TCDD was also shown to block the androgen dependent proliferation of prostate cancer cells14. Simultaneous activation of AhR and AR with TCDD and an androgen derivative, respectively, decreased AR protein levels15. Thus, activation of AhR by a ligand results in decreased protein expression of both AhR and AR. This action may explain the anti- androgenic actions of a number of PAHs, is distinctive from the effects seen with constitutive AhR signaling that accompanies overexpression of the AhR protein and may be the result of enhanced activity of AhR chaperone protein, Src kinase.The aryl hydrocarbon locus includes AhR, ARNT, and the AhR repressor (AHRR), which are critical for regulation of AhR signaling16. The PAS domains facilitate interactions with other PAS domain proteins, such as the AhR binding partner, ARNT PAS-A is primarily responsible for protein-protein interaction and PAS-B also encompasses the ligand binding17. cSrc may interact directly with the AhR transactivation and PAS domains7. Western blot and FRET analysis confirm time- dependent phosphorylation of Src following activation of AhR which was blocked in the presence of a specific AhR antagonist18. Furthermore, coimmunopercipitation experiments revealed that AhR regulates Src activity by phosphorylating Src (Tyr 416) and dephosphorylating Src (Tyr527)19.The mammalian AhR protein contains four major structural motifs important in AhR’s interactions with other proteins and transcription factors. The N-terminal basic-helix-loop-helix is the site of DNA binding domain and also participates in dimerization and HSP90 binding. The transactivation domain spans from amino acid 490 to 805 and includes a central glutamine rich region. The two Per-Arnt-Sim (PAS) domains are named after their homology with the clock protein period (Per), the xenobiotic and oxygen sensing ARNT, and the neuronal cell lineage regulator single-minded (Sim) 16. The P20. There are also many reports that SFKs are abnormally activated in prostate cancer cells. SFKs are activated in response to numerous stimuli including neuroendocrine ligands, reactive oxygen species, cytokines and growth factors. These molecules have proven roles in cancer progression, including cell proliferation, adhesion, migration, and invasion21.The increased expression of Src and other Src family kinases (SFK) in a number of prostate cancer cell lines has suggested a role for Src in prostate cancer initiation and progression8. TCDD activation of AhR results in time- and dose-dependent phosphorylation of Src (Tyr416). Reported Src-mediated crosstalk between AhR and EGFR signaling pathways demonstrated an important link between AhR canonical function and TCDD-mediated tumor promotion23. Through direct physical interaction with AR, Src is able to phosphorylate AR and thereby induce ligand-independent activation of AR, a key mechanisms of CRPC24. Conversely, AR overexpression also plays a role in the oncogenic potential of wild-type Src. This suggests that crosstalk and activation of AR and Src is reciprocal25.Src and AhR coexist in a protein complex that also contains HSP90, AhR-interacting protein, and p23 that aides to maintain an inactive AhR complex26. As a phosphoprotein, AR has several serine/ threonine and tyrosine residues that are phosphorylated. Most of the phosphorylated residues are located in N-terminal domain that regulates AR cellular localization, stability and its transcriptional activity.AR contains at least sixteen phosphorylated residues. Several of the residues are phosphorylated after treatment of cells with androgen, antiandrogen, or reagents which activate other signaling pathways and alter transcriptional activity, cellular localization, and stability of AR28.Recently, several groups have reported that tyrosine phosphorylation of AR protein by non- receptor tyrosine kinases Src may have a role in AR activation in the low androgen environment, thereby promoting the development of CRPC. Src-mediated phosphorylation of AR at Y534 resulted in the activation of AR followed by nuclear translocation and DNA binding in the absence of androgens 26–28.29. In the presence of low levels of androgen, AR is normally localized to the cytosol in a complex with molecular chaperones, Hsp40, 70 and 90 in an inactive form. Upon androgen binding, the androgen induces conformational changes in the protein, forms a homodimer, and translocates to the nucleus30. The nuclear translocation results in binding of AR as a transcription factor to ARE’s in the regulatory region of target genes. AR signaling plays a critical role in prostate cancer cell proliferation, survival, and differentiation31.AR is a member of the steroid hormone receptor family and shares a similar domain organization with other members of the nuclear receptor (NR) which is primarily responsible for mediating the physiological effects of androgens by binding to AREs32.Several studies based on molecular cloning of AR cDNA, suggest that its transcriptional activity is critical for all stages of prostate cancer development and progression. Several neutral next- generation sequencing platforms have been used to pursue genomic characterization of prostate cancer at various stages. Results have consistently confirmed a critical role for AR activity in prostate cancer progression33. Whole genome sequencing of 11 early onset prostate cancers suggested that androgens, through AR, contribute in shaping somatic alterations34. These results are further supported by studies demonstrating that AR is known to stimulate the expression of TMPRSS2: ERG which is a common gene fusion associated with prostate cancer initiation via androgen-driven overexpression of the gene fusion products35. It is clear that AR signaling plays a critical role in the development and progression of prostate cancer and serves as a central component of progression to CRPC.While AR signaling is known to modulate the expression of genes associated with cell cycle regulation, survival and growth, its specific functions are not fully defined in prostate cancer38. Direct heterodimerization of AhR and AR has been shown to occur in cells and may partially explain the crosstalk between the two receptors5. Additionally, interaction can happen via coactivators. AhR and AR share a number of coactivator proteins such as SRC1 and p30039. Another possible mechanism AhR may utilize for AR activation is phosphorylation of AR by Src kinase40. Src was shown to mediate crosstalk between AhR and epidermal growth factor receptor in colon cancer cells19. Other studies have shown that Src kinase can promote AR transactivation in C4–2 cells prostate cancer cells. Consequently, inhibition of Src kinase function with a specific inhibitor resulted in decreased AR activation41. Coimmunoprecipitation experiments revealed that AhR forms a protein complex with Src and regulates Src activity by phosphorylating Src (Tyr416) and dephosphorylating Src (Tyr527)42. Immunoprecipitation assays also revealed the association of AR with Src, suggesting complex formation among them43.AhR signaling influences androgen signaling directly at the level of the protein and indirectly through actions on the endocrine system. AhR plays a significant role in sustained AR signaling and growth. However, the mechanism for this role is not clearly understood. One possible mechanism is direct interaction between AhR with AR. AhR has been reported to directly interact with a number of nuclear proteinsin vitro and in vivo studies have established a role for AhR in prostate and prostate cancer development. The development of AhR-null mice has revealed that the function of this receptor is not limited to mediating the effects of PAHs and other ligands44. AhR knockout mice exhibit decreased fertility, decreased liver size, and structural and functional deficits in several tissues45. These include reproductive tract problems such as decreased levels of mature follicles and formation of uric acid stones in the urinary bladder46.The precise molecular mechanism utilized by constitutive AhR signaling to activate AR signaling needs to be investigated further and could include induced activation via protein phosphorylation, direct heterodimerization and interacting via coactivators. Extensive 47. Ligand activation of AhR by TCDD is adequate to disrupt key stages of ductal morphogenesis. Significantly smaller prostate lobes were observed in rats and mice exposed to TCDD during fetalpubertal development. Indicating that TCDD-induced AhR activation delays prostate growth48. TCDD exposure also led to a decreased number of prostate ducts in monkeys49. AhR may serve as a key regulator of AR signaling in CRPC by utilizing Src as a signaling intermediate. Recent studies have revealed that simultaneous inhibition of AhR and Src is sufficient to abolish AR signaling in CRPC cells50. Activated Src is a common signaling intermediate in a number of pathways including AR signaling pathway and has increased activity in CRPC. Although several mechanisms have been suggested for the sustained AR signaling, tyrosine phosphorylation of AR protein by AhR- activated-Src may have a significant role in CRPC that also exhibit constitutive AhR signaling  and in normal, hyperplastic, and cancerous adult human prostate tissueignaling .


Subgroup analyses revealed that minocycline reduced depressive-like behavior in diseased, but not healthy, animal models. Finally, minocycline was found to reduce both immobility-based and anhedonia-based outcomes. These findings suggest that minocycline may be an effective treatment of core depressive symptoms, and that further investigation of minocycline treatment for clinically relevant depression in humans is warranted.Evidence from recent animal studies suggest that minocycline, a broad-spectrum antibiotic capable of regulating immune processes, may possess antidepressant properties. These studies, however, have yet to be comprehensively reviewed. Accordingly, this systematic review and meta-analysis summarizes the extant literature examining the effect of minocycline on depressive-like behavior in rodent models. PubMed, PsycINFO, and Web of Science databases were systematically searched for articles that met prespecified inclusion and exclusion criteria, and standardized mean differences (SMDs) were calculated for each continuous measure of depressive-like behavior. The overall effect of minocycline on depressive-like behavior was estimated using robust variance estimation meta-analysis. Separate subgroup analyses were conducted on diseased vs healthy animal models, different rodent species, and immobility-based vs anhedonia-based measures of depressive-like behavior. A total of 22 preclinical studies  were included. Overall, minocycline reduced depressive-like behavior in rodents (SMD = −1.07, 95% CI −1.41–−0.74,  The disorder is estimated to cost the United States over $200 billion per year in medical services and workplace-related losses3. Significantly, depression accounts for 7.5% of all years lived with disability worldwide—the greatest disability burden of any single disease2. It is also a significant source of mortality via depression-linked suicide, as well as through its association with ischemic heart disease4.Major depressive disorder is a debilitating form of mental illness that affects approximately 1 in 20 people worldwide5, but the magnitude of this effect was rather modest for nearly all the drugs evaluated. Simply put, many depressed patients do not respond adequately to conventional antidepressant therapy8, and even those who do will typically face a high risk of relapse following acute treatment10. Such findings highlight the continued need for development of novel efficacious depression interventions.Antidepressant medications are often employed as the primary treatment for depressive illness. A recent comprehensive meta-analysis found 21 different antidepressants to be significantly more efficacious than placebo12, and pro-inflammatory medications carry the potential to induce depressive symptoms in humans14. Preliminary evidence also suggests that some anti-inflammatory drugs may have antidepressant effects comparable to those of conventional pharmacological treatments15. Microglia are the primary immune cells within the central nervous system that mediate neuroinflammation16, and minocycline is capable of reducing inflammation both by directly suppressing microglia activation and by indirectly inhibiting microglia-induced inflammatory processes17. In doing so, minocycline also indirectly acts upon key monoaminergic systems implicated in depressive symptomatology, including the modulation of neurochemical cascades involving serotonin, dopamine, and norepinephrine11, as well as promotion of serotonin synthesis by reducing activation of the kynurenine pathway17. Finally, minocycline protects against oxidative stress and neuronal damage, two other harmful processes implicated in pathogenesis and maintenance of depression18.Minocycline, a second-generation antibiotic with pronounced inhibitory effects on the brain’s microglia, may represent one such innovative approach. A growing body of evidence suggests that minocycline beneficially targets several pathways of relevance to the pathophysiology of depression. For instance, inflammation is strongly associated with depression20. Additionally, an open-label study by Miyaoka et al.21 reported that minocycline, in combination with antidepressants, reduced depressive symptoms in individuals with psychotic depression. Only a few placebo-controlled clinical trials, however, have investigated minocycline as a treatment for depression in humans. Although a recent meta-analysis of three randomized controlled trials22 found support for an antidepressant effect of minocycline, it is premature to draw any substantive conclusions in light of the limited evidence available.Preliminary evidence also suggests that minocycline may have psychotropic effects in humans. The addition of minocycline to treatment with atypical antipsychotics has been shown to reduce negative symptoms, such blunted affect and anhedonia, in individuals with early-stage schizophreniaDespite the paucity of published clinical trials, there does exist a much larger preclinical  literature regarding the antidepressant potential of minocycline. However, to the best of our knowledge, these preclinical studies have not yet been comprehensively reviewed. Accordingly, the purpose of this study is to summarize and quantify existing preclinical studies of the effect of minocycline on depressive behaviors by means of a systematic review and meta-analysis.44 with 39 independent experimental groups and 816 rodent subjects were included in the meta-analysis . Two studies assessed olfactory bulbectomy-induced hyperactivity with the open field test, proposed to be a measure of depressive-like behavior in this specific rodent modelDoses of minocycline ranged from 10–160 mg/kg and the duration of treatment ranged from a single administration to 42 consecutive days. Minocycline was most commonly administered by either intraperitoneal injection or through drinking water.45. More detail was generally provided regarding attrition, reporting, and other forms of potential bias. Over half of the included studies had a low risk of attrition and other bias, while all included studies had a low risk of reporting bias. One study had a high risk of detection bias41, and five studies had a high risk of other bias42. A separate sensitivity analysis revealed that removal of these studies did not substantively impact our final results. Overall, however, the risk of bias for each individual study is unclear, which suggests that the observed treatment effects may be overestimated47.Risk of bias assessment for the included studies can be viewed in Table p < 0.001; Fig. p = 0.961), treatment duration , or treatment dose .Combining standardized mean differences (SMDs) for the 39 included experimental groups revealed a pooled SMD of −1.07 ; it had no significant impact among healthy animals, i.e., those naïve to experimental manipulation . Minocycline also significantly reduced depressive-like behavior in both rats  and mice . Separate analyses revealed that minocycline significantly reduced both immobility-based  and anhedonia-based  outcomes. An exploratory meta-regression indicated that both minocycline pretreatment and acute treatment in diseased animals resulted in a significant reduction in depressive-like behavior compared to healthy animals . Additionally, treatment following disease induction was associated with a significantly greater antidepressant effect than that of pretreatment .Subgroup analyses revealed that minocycline administration significantly reduced depressive-like behavior only among diseased animals . Together, these findings suggest that publication bias may be present in the included data, resulting in the omission of non-significant or opposing findings in the final analyses. There was also high heterogeneity across the 39 experimental groups (I2 = 75.7%), indicating a great deal of inconsistency across studies. High heterogeneity and funnel plot asymmetry, which have been previously identified as a significant problem among meta-analyses of animal research, weaken the generalizability of overall findings48. Inclusion of study characteristics, such as sample size, treatment differences, and subject species/strain as moderating variables failed to reduce heterogeneity. However, separate analysis of diseased and healthy animals did reduce heterogeneity to moderate levels , supporting recent proposals that the use of consistent and well-validated experimental models may aid in the translation of preclinical research findings49.Visual inspection of a funnel plot Fig.  revealedThis systematic review and meta-analysis included 22 preclinical studies, which encompassed a total of 39 independent experimental groups and 816 rodents. Minocycline administration significantly decreased depressive-like behavior among study animals in our primary analysis, with a robust pooled standardized mean difference (SMD) of −1.07 in comparison with placebo. Moreover, 19 of the 22 included studies reported a statistically significant minocycline effect on at least one salient outcome measure.39, provoked inflammation or sickness29, and olfactory bulbectomy34—are all associated with an increase in depressive-like behavior51. In other words, it is possible that minocycline only exerts significant antidepressant effects in the presence of elevated levels of depressive symptomatology. This conjecture is congruent with the finding that conventional antidepressants outperform placebo most consistently in more severe cases of depression53. Accordingly, future clinical trials might be advised to attend carefully to baseline depression severity among study patients, and perhaps to focus the investigation of minocycline effects upon the severe and very severe depression subtypes.Notably, however, minocycline only evidenced significant antidepressant effects among animals that were already experimentally stressed or diseased before they faced the depression assessment protocol—typically another acutely stressful procedure such as forced swimming or prolonged tail suspension. The drug had no significant impact upon animals that entered the challenging depression paradigm in an otherwise healthy state. This striking between-group contrast may perhaps best be explained by critical differences in depression symptom severity among the healthy and diseased rodents, as the induced disease states among study animals—chronic stress54, they also seem to protect against the onset of iatrogenic (treatment-induced) depression in humans. For example, prophylactic antidepressant use reduces the risk of depression among patients receiving depressogenic interferon-α treatment55. Additionally, patients administered an antidepressant prior to surgery or radiation therapy for cancer may become less likely to develop depression during treatment56. Because minocycline pretreatment likewise protects against disease-induced depressive-like behavior in rodents, it will be valuable to see future investigation of its prophylactic value in humans, especially for medically-induced depression in clinical settings. On the other hand, we did find that the therapeutic effect of minocycline was largest when administered following disease induction, which raises the possibility that the drug might be most useful as a treatment for existing depressive symptoms.In our secondary analyses, minocycline reduced depressive-like behavior in diseased animals regardless of whether the drug was administered as a pretreatment—prior to depressogenic challenge—or as an acute treatment of existing symptomatology. Such a finding has clear parallels in the clinical literature. Although antidepressants are most commonly prescribed to treat active depressive episodes57, although there is still some debate over the extent to which immobility challenge behaviors reflect a true depressive phenotype59. The sucrose preference test, on the other hand, is used to assess anhedonic behaviors59. Negative mood and anhedonia are fundamental features of depression; in fact, the current classification of major depressive disorder effectively requires at least one of these two symptoms to be present for diagnosis60. Anhedonia and dysphoria also both contribute substantially to psychosocial impairment in depression61, rendering them particularly important targets for antidepressant therapy. Inasmuch as minocycline may address both symptom domains, it may have potentially far-reaching clinical utility.It is also worth noting that minocycline improved performance on both immobility- and anhedonia-based measures of depressive-like behavior in study analyses. Immobility measures, such as the forced swim test and the tail suspension test, are widely used to screen potential antidepressants, and they are considered to represent negative affectivity44. Minocycline was also observed either to alter microglia function, including glial gene expression, or to reduce biomarkers of inflammation in 10 studies43. Finally, one reviewed study found that minocycline decreased depressive-like behavior while simultaneously altering the composition of the gut microbiota41—symbiotic microorganisms that play a key role in regulating immune processes62—thereby highlighting another possible route by which minocycline could target inflammation.The observed antidepressant-like effects of minocycline may derive in part from the drug’s established anti-inflammatory properties, including its ability to inhibit microglia-induced inflammatory processes in the brain. In our analysis, five of six studies found that minocycline had a meaningful effect on depressive-like behavior after the experimental induction of an inflammatory state63. Insufficient detail was provided for seven of the 10 domains enumerated in the influential SYRCLE risk-of-bias tool64, thereby raising concerns that observed results may be affected by biases related specifically to inadequate randomization, allocation concealment, and blinding procedures. Such biases can lead to an overestimation of treatment effects47, which can in turn hamper translational research efforts. In fact, only an estimated 10% of investigational drugs that begin clinical testing obtain final approval, and those that reach later phases of clinical trials frequently fail due to lack of efficacy65. Reducing bias in preclinical trials can help to decrease such attrition by facilitating more accurate predictions of clinical efficacy49, which may in turn decrease the financial and human burden associated with failed clinical trials. Future investigations of minocycline, accordingly, should take care to properly implement and report procedures to reduce selection, performance, and detection bias.One important interpretive caveat for the present work is the fact that the risk of bias for each reviewed study is unclear—a pervasive, well-documented limitation of much preclinical research66. It is possible that such resistance might diminish the antidepressive efficacy of minocycline, even if not used in an antibacterial capacity. Likewise, it may be useful to investigate the degree to which adjuvant medications designed to combat acquired resistance, such as loperamide67, potentiate the effects of minocycline in depressed patients.A major barrier facing the use of minocycline in clinical settings is acquired resistance. Similar to other forms of antibiotic, minocycline usage is linked to the development of resistant bacterial strains68, the present results should be interpreted with appropriate caution. A major source of this heterogeneity was simply the methodological variability across studies, as the mere separation of healthy and diseased animals for analytic purposes led to a substantial reduction in statistical heterogeneity.One important limitation of the present analysis concerns the high level of methodological heterogeneity present in the reviewed studies. Subject characteristics, animal models, treatment dose, and administration period differed significantly across studies. Because high heterogeneity reduces the predictive validity of meta-analyses69. They can also lead to publication bias, which our analyses suggest may be present in the minocycline preclinical literature; if so, it could further weaken the generalizability of study results69. Also, although 39 experimental groups—a reasonably large number for meta-analytic purposes—were included in the overall analysis, the number was roughly halved for our two key subgroup analyses . Because subgroup analysis can result in substantially reduced statistical power70, this study may not have been adequately powered to detect all statistically significant subgroup effects. Finally, our search strategy, limited to the inclusion of only mice and rat subjects and English-language articles, may have introduced bias71.Another limitation of the present work is the fact that sample sizes of included studies typically ranged between 10 to 30 animals. Such small sample sizes, common in preclinical studies, can negatively affect both the reliability and validity of study outcomesIn conclusion, minocycline was observed to significantly reduce depressive-like behavior on both immobility- and anhedonia-based measures. These findings support the desirability of additional clinical trials to investigate the antidepressant potential of this antibiotic among humans. Because minocycline had no effect on depressive-like behavior in healthy animals, it may prove optimal for future research to focus principally upon the treatment of clinically significant depression, as opposed to prophylaxis among healthy individuals. Finally, the anti-inflammatory effects of minocycline should be more extensively investigated as a promising candidate mechanism of antidepressant action.72. The study protocol was not preregistered and can be viewed at 10.17504/protocols.io.vege3bw.We adhered to CAMARADES guidelines for conducting systematic reviews and meta-analyses of animal studiesStudies were deemed eligible if they met the following inclusion criteria: 1) study subjects were either rats or mice; 2) minocycline was experimentally administered; 3) depressive-like behavior was a primary or secondary study outcome.Studies were omitted from review if they met one or more of the following exclusion criteria: 1) there was no matched control group; 2) the experimental subjects did not directly receive the administration of minocycline ; 3) descriptive outcome data  were not available for the measured depressive-like behavior; 4) the full text of the article was not available in English. Conference abstracts were omitted due to lack of necessary information.Potential studies were identified by searching PubMed, PsycINFO, and Web of Science databases, from the earliest record of the databases to September 2017. Search terms included minocycline OR probiotic AND depressive-like (the exemplar PubMed search strategy can be found in the Supplementary Information). All studies were screened and evaluated in a standardized manner by two independent reviewers (DR and EC). First, the title and abstract for each search result were reviewed to identify potential studies, after which full-texts were evaluated to determine study inclusion. Disagreements between reviewers were resolved by discussion, with final decisions made by consensus. The flow chart of study selection can be viewed in Fig. 72. Extracted data included subject information , intervention type , and outcome data . Different measures of depressive-like behavior were assumed to be equivalent. If depressive-like behavior was measured at multiple time points throughout the study, the final measurement of the interventional period was selected as the included data. Data presented only in graphical format were extracted using WebPlotDigitizer graph digitization software73, a program recommended for use in systematic reviews74. Potential study bias was evaluated using SYRCLE’s risk-of-bias tool64. All data and risk-of-bias evaluations were extracted by two independent reviewers (DR and EC), and disagreements during this process were resolved by discussion. Finally, 11 authors were contacted and asked to provide additional study details and data. Two authors responded and these studies were included in the final study selection. Extracted study characteristics can be viewed in Table Data collection was conducted using a custom form based on CAMARADES guidelines75. All analyses were pre-specified unless otherwise stated. A standardized mean difference  between minocycline-treated and matched control groups was calculated for all continuous measures of depressive-like behavior. A standard normal distribution was used to calculate confidence intervals for each SMD. Sample size, treatment duration, and treatment dose were assed as moderating variables in individual meta-regressions—study-level variables, rather than aggregated patient-level data, were chosen as moderators to avoid potential ecological bias76. Separate subgroup analyses were conducted on diseased  and healthy animals (those receiving only minocycline or placebo intervention prior to outcome measurement), as well as mouse and rat samples. An exploratory meta-regression was conducted to compare minocycline effects, respectively, among healthy animals, diseased animals given the drug prior to disease induction (pretreatment), and diseased animals given minocycline after disease induction (treatment). Results from immobility-based (i.e. forced swim test and tail suspension test) and anhedonia-based (i.e. sucrose preference test) measures of depressive-like behavior were also analyzed in separate analyses. Multiple rodent subgroups within a single study  were included as independent SMDs, provided that each treatment group had a separate, matched control group. Whenever multiple treatment groups were compared against the same control group, an SMD was calculated from the combined results of the different treatment groups77.The meta-analyses were performed with R 3.4.4 software78. In RVE meta-analysis, multiple outcomes from a single study are included as separate SMDs and share a single study weight. SMDs were weighted by their precision . Between-study heterogeneity was evaluated using the I2 statistic, with values of I2 more than 25%, 50%, and 75% selected to reflect low, moderate, and high heterogeneity, respectively79. Potential publication bias was assessed for using funnel plot asymmetry and an Egger test80.If multiple measures of depressive-like behavior were available for a single treatment group, then a separate SMD was calculated for each outcome. Summary SMDs were calculated using robust variance estimation (RVE) meta-analyses, a form of random-effects meta-analysis shown to address dependency between SMDs measured in the same sampleSupplementary InformationDataset 1


Objective: To present a case report highlighting a severe, yet avoidable, complication following small needle-scalpel treatment for cervical spondylosis.Introduction: The small needle-scalpel is a miniature surgical instrument used to create intense and invasive punctures at certain acupoints with a small latch needle. It has been increasingly gaining popularity among clinicians and patients all over the world during the past years. However, severe complications after small needle-scalpel treatment have not previously been reported.Methods: Here we report a 54-year-old man who recently suffered from cervical spondylosis and underwent small needle-scalpel treatment, which was performed by a rural doctor. While there were no new neurologic deficits, the patient experienced delayed functional deterioration until the onset of quadriplegia within 1 month of treatment. Magnetic resonance imaging demonstrated a C2–C7 dorsally placed extradural hematoma with severe cord compression and subcutaneous soft tissue hemorrhage.Results: The patient refused urgent corrective surgery and later died due to respiratory failure.Conclusions: Although small needle-scalpel therapy has many benefits, such as reducing pain, shorter expenditure, shorter period of therapy and better recovery of function, there are also many potentially severe risks, such as cervical extradural bleeding, which requires clinicians to pay more attention to avoid the complications. The small needle-scalpel is a miniature surgical instrument that creates intense and invasive punctures at certain acupoints with a small latch needle . The shaA 54-year-old Chinese man consulted a medical doctor with the complaint of recent painful neck and right upper limb without any clear causes. He had a medical history of left traumatic humeral fracture 10 years ago that did not significantly affect his recent daily life after surgery. The patient had no recent history of head or spinal trauma and did not take any related antiplatelet or anticoagulant drugs. In addition, the patient had no exposure to any toxic substances and no significant relevant family history. A clinical diagnosis of cervical spondylosis was originally considered based on X-rays of the cervical spine, and the patient received small needle-scalpel treatment for relieving pain relief, which was administered by a rural doctor. When no obvious improvement was observed 20 days later, the patient was admitted to our hospital for further diagnosis and treatment. The patient was experiencing progressive neurologic deterioration involving both upper and lower limbs. Upon neurological evaluation, he exhibited reduced response to light touch and pinprick, with a sensory level below the angle of the mandible. He had trouble during urination and significantly decreased muscle tension. He was subsequently incapable of antigravity power in his upper limbs  and had a significant loss of motor function in both lower extremities (the muscle strength was graded as 1/5). Urgent magnetic resonance imaging (MRI) of the cervical spine demonstrated a right spinal epidural hematoma located at the posterior spinal epidural space with severe cord compression and subcutaneous soft tissue hemorrhage, extending from the C2 to C7 spinal vertebral level . RoutineCervical extradural hematoma is an uncommon but highly disabling neurologic emergency in clinical practice that causes morbidity without early surgical intervention . The etiAlthough widely considered effective and safe, several potential side effects, such as neural tissue injury, skin infection, and bleeding adhesion, have been reported with the use of acupuncture, with having a 8.6% incidence of side effects , 19, 20.In conclusion, although there are many benefits, such as reducing pain, less expenditure, a shorter period of therapy, and better recovery of function, small needle-scalpel treatment may also have the potential to cause severe risks, such as cervical extradural bleeding which requires clinicians to pay more attention to avoid related complications. Precisely choosing the acupoints, standardized surgery and long-term follow-up may avoid related complications followed by small needle-scalpel treatment.As this is a case report without experimental intervention, no formal research ethics approval is required. The relatives of this patient provided written informed consent to the publication of the information and images related to this case report.JH drafted the manuscript. MY and QZ helped to prepare the figures and collected data. TJ performed a critical revision.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.


Neurodegenerative disorders constitute a group of multifaceted conditions characterized by the progressive loss of neurons and synaptic connections consequent to a combination of specific genetic predispositions and stochastic stressors. The neuropathologies observed in both Alzheimer’s and Parkinson’s disease are in part attributed to compounding intrinsic and extrinsic environmental stressors, which we propose may be limited by the administration of specific grape derived phytochemicals and their metabolized derivatives, specifically polyphenols isolated from grape botanicals. Current therapies for neurodegenerative disorders are limited as they solely target the final disease pathologies including behavioral changes, cognitive deficits, proteinopathies and neuronal loss; however, this strategy is not a sustainable approach toward managing disease onset or progression. This review discusses the application of grape derived polyphenols as an adjunctive treatment paradigm for the prevention of neuropathologies associated with Alzheimer’s disease, Parkinson’s disease and Chronic Traumatic Encephalopathy by simultaneously ameliorating two stochastic stressors that facilitate their disease pathologies: inflammation and oxidative stress. The biophysical attributes of grape-derived polyphenols buffer against redox potential dependent peripheral and neuroinflammation and down regulate the activation of inflammasomes in microglia and astrocytes, which could provide a novel mechanism through which grape-derived polyphenols simultaneously suppress risk factors across pathologically distinct neurodegenerative conditions. This approach therefore offers a prophylactic mode, not feasible through current pharmacological agents, to target activity dependent risk factors for neurodegenerative disorders that manifest over an individual’s lifetime. Neurodegeneration is the defining feature of a group of age-related neurological disorders characterized by the progressive atrophy of neurons in cortical and subcortical brain structures. The distinct etiological roots of neurodegeneration remain to be fully characterized; however, stochastic environmental stressors and toxins effectuating persistent inflammation and elevated oxidative stress increase the likelihood of developing neurodegenerative conditions and precede canonical neuropathologies commonly associated with neurodegeneration . IdentifAlzheimer’s Disease is an age-related neurodegenerative disease with treatment options limited to cholinesterase inhibitors for memory impairments. In later disease stages, AD incurs greater costs due to patient care in handling the progressive memory loss, nervousness, cognitive disability, and the eventual loss of bodily functions that ultimately lead to death. AD is the sixth leading cause of death in the United States affecting 5.4 million Americans and costing the healthcare system an estimated $236 billion: numbers which are expected to triple over the next 30 years . GloballThe remaining sporadic forms of AD are in part attributed to synergistic interplays between the accumulation of lifestyle dependent environmental stressors and genetic polymorphisms prevalent in patients with sporadic AD . ExposurParkinson’s Disease is the leading movement disorder and estimates suggest that more than 1 million people in the United States live with PD. More than 60,000 new PD cases are diagnosed each year; a prevalence expected to double in line with an aging population by 2040 . Like ADChronic Traumatic Encephalopathy (CTE) is a neurodegenerative disorder most prevalent in individuals who have experienced repetitive brain traumas or acute traumatic brain injury (TBI). Athletes who play contact intensive sports often resulting in concussions such as professional football players or soldiers exposed to military grade blasts are at an increased risk for developing CTE compared to the general population . TBI doePatients diagnosed with CTE initially present with insidious symptoms associated with cognitive dysfunction including irritability, impulsivity, depression and short-term memory loss . As the Neuropathological surveys of patients with advanced AD observe pervasive neurodegeneration and distinct neuropathological features including intercellular neurofibrillary tangles composed of hyperphosphorylated tau and intracellular senile plaques composed of Aβ42 . NeurodeElevated markers of inflammation and oxidative stress are pervasive in all brain regions in AD and serves as important biomarkers and potential pathological targets, as they promote Aβ deposition, microglia activation, and BBB permeability . The oxiin vivo imaging techniques  inhibited the motor deficits and improved neuronal survival to both the OHDA- and LPS-induced PD pathology indicating that inflammasome activation and its downstream pathways are critical determinates for PD pathologies , neurites, and glial tangles (GTs) in both cortical and subcortical structures. Tau deposition in CTE has distinguishable features from those in patients with AD . In CTE,Figure 1). Inflamm-aging was a special term coined by Francheschi et al. which suggested that age-related chronic low-grade inflammation is asymptomatic, but overtime the effects compound to promote altered cellular function  that compensate for acetylcholine deficits and loss of projection cholinergic neurons in the basal forebrain. Toxicity associated with upward titration and long-term use limits their prolonged clinical utility, and over time AD neuropathologies progress in spite of treatments with ChEI’s . From a As inflammation plays an important role in the etiology of AD pathologies, several anti-inflammatory therapies have been tested in clinical settings. For instance, long-term use of NSAIDs decreases AD development, slows disease progression, and improves cognitive functioning . LikewisSeveral antioxidant therapies have been tested in AD patients but with little success. Although supplementation with vitamin E and C in addition to cholinesterase inhibitors reduced levels of LPO in the blood, there were no changes in cognition . SimilarTherapeutic approaches under clinical development for AD target the neuroimmune system, microglia activation, or proteins in neuroinflammatory cascades. Mutations in the heterozygous triggering receptor expressed on myeloid cells (TREM2) represent an emerging genetic risk factor for AD  and implThe predominant symptomatic therapies for PD increase intrasynaptic concentrations of dopamine in degenerating dopamine nigrostriatal circuits. Dihydroxyphenylalanine (L-DOPA) is the molecular precursor of dopamine and its decarboxylation by tyrosine hydroxylase increases levels of intrasynaptic dopamine. Treatment of PD with formulated L-DOPA is the most common monotherapy for PD motor impairments . The breAlternative therapeutic approaches for treatment of PD focus on neuron and microglia metabolism and the reduction of synuclein plaques. Creatine monophosphate is an energy source during periods of cytosolic ATP depletion and has neuroprotective attributes in preclinical models of PD . HoweverOther novel therapeutic approaches under clinical development for PD target neuroinflammatory pathways. Gene polymorphisms of the LRRK2 protein are among the most prevalent causes of late-onset autosomal dominant and sporadic PD; the protein is highly expressed in macrophages, monocytes, and neutrophils and mutations result in hyperinflammatory responses from innate immune cells . The LRRTable 1); however, more work is required to define the specific bioactive components of these formulations and how they suppress the underlying disease physiologies. Our group has previously outlined the benefits of polyphenol-rich grape extracts toward the prevention of AD  containing 79.2 mg/g of proanthocyanidin dimers, 33.7 mg/g of catechin, 17 mg/g of epicatechin, and 22.3 mg/g of gallic acid . Using tin vitro . These raze test . In partaze test .Apart from Aβ aggregation, we have shown how GSPE attenuates the formation of tauopathies, another pervasive clinical manifestation in AD dementias. Non-covalent interactions between GSPE-derived polyphenols and tau residues were believed to be the mechanism of action for the prevention of tau peptide aggregation . Indeed,in vitro  toxic model of PD induces a state of chronic inflammation, mitochondrial dysfunction, and oxidative stress that contributes to disease onset and progression . When adN-methyl-4-phenyl-1,2,3,6-tetrahydropyridinium hydrochloride (MPP)+ + . Flavonoe (MPP)+ . Myricetl slices . Additiol slices .in vitro and in vivo PD models. In rats pre-treated for 15 days with resveratrol prior to a unilateral intrastriatal injection with 6-OHDA, there was a significant decrease in the levels of thiobarbituric acid reactive substances (TBARS), lipid peroxidation, and decreased activity of phospholipase A2. The investigators also reported a significant increase in expression and activities of antioxidants such as glutathione in the striatum , resveratrol and quercetin prevented apoptosis of the PC12 neurons in a manner dependent on the inhibition of microglial activation and consequent reduction of IL-1β and TNFα release , hydrogen peroxide (H2O2), hydroxyl radical (.OH), alkoxyl radicals (.OR), or peroxyl radicals (.OOR). In addition to activating inflammasomes, ROS readily react with cellular lipids, carbohydrates, proteins and nucleic acids to produce toxic secondary species. Polyphenol compounds are strong antioxidants due to their characteristic aromatic ring structure that always contains at least one substituent hydroxyl group. Polyphenols eliminate strong ROS such as hydroxyl radicals through homolytic cleavage of the hydroxyl group, which delocalizes the singlet electron into the π orbitals of phenol ring and forms energetically stable resonance derivatives. The relatively higher reduction potential of the new radical compounds prevent further propagation of new radicals. Polyphenol compounds are diverse in the number of aromatic rings and their conjugating moieties, which increase or decrease their antioxidative properties, respectively. For example, a greater number of para- and ortho- substituent electron donating groups on aromatic rings decrease the hydroxyl dissociation energy, while more conjugated π orbitals in the planar structure increase resonance effects . On a relative basis, it was previously reported that between the four polyphenol compounds mentioned, quercetin most readily sequesters the synthetic radical DPPH and the physiological peroxynitrate anion from solution followed by catechin, epicatechin and resveratrol (Polyphenol compounds have inherent antioxidant characteristics due to the electron structure of their phenolic rings. Reactive oxygen species (ROS) can be derived from the mitochondria, as seen following lysosomal rupture, osmotic disequilibrium, or metabolic stress , or can  effects . Abundanveratrol . SequestAs indicated throughout this discussion, inflammasome-mediated neuroinflammation may contribute to overt neuropathological signatures in neurodegenerative disorders. Inflammasomes are a class of multimeric protein complexes that are responsible for initiating and propagating inflammation in response to endogenous or exogenous insult signals, such as alterations to intracellular redox potentials. The NLRP3 inflammasome is of particular interest for neurodegenerative disorders because experimental evidence indicates that it gates neuroinflammatory responses through its unique kinetic properties, which is different from other inflammasomes . As neurNlrp3 gene, while a second activation signal promotes assembly of the multimeric complex and its activation (Section Polyphenol Compounds as Free Radical Scavengers identify them as potential inhibitors of ROS mediated NLRP3 activation. Several studies provide evidence that grape-derived polyphenols can suppress inflammasome activity and the production of deleterious inflammatory cytokines (Activation of the NLRP3 inflammasome requires a two-step paradigm whereby an initial stress signal promotes the transcription of the tivation . Upon activation . Importativation . Cholesttivation . Inhibitytokines . There iIn a subarachnoid hemorrhage model of TBI, the administration of resveratrol prevented cerebral inflammation and cortical apoptosis, events that were concomitant with reduced NLRP3 activity, microglia activation, and neutrophil infiltration into the brain . InhibitE. coli O157:H7 in epithelial cells by maintaining both mitochondrial integrity and by preventing the accumulation of mitochondrial ROS (Quercetin inhibited the induction of inflammation by the toxin rial ROS . In the rial ROS . Catechirial ROS . Similarrial ROS . The admrial ROS . In a rarial ROS .The procyanidins also possess anti-inflammatory activity, as illustrated through by their ability to inhibit inflammasome activation and reduce ROS formation in a LPS-induced model of human endothelial cell inflammation . SimilarActivation of inflammasomes parallels microglia reactivity. Studies were therefore conducted to determine if polyphenols simultaneously target NLRP3 inflammasome activation and microglia reactivity. One study found that intraperitoneal administration of dihydromyricetin attenuated the number of activated microglia in the hippocampus and cortex of APP/PS1 mice while reducing expression of NLRP3 inflammasome components and stimulating clearance of Aβ . Dual doin vitro and in vivo to have potent antioxidant and anti-inflammatory activity that improve neuropathological features and behavioral phenotypes in neurodegenerative diseases. In particular, a previously untapped mechanism of neuroinflammation involving inflammasome activation may be a potential pharmaceutical target of polyphenolic anti-neuroinflammatory effects in the context of neurodegeneration. The studies to date are encouraging as they support the development of an optimized grape-derived polyphenol therapeutic that through its anti-inflammatory and antioxidant activities, may delay the onset or slow the progression of neurodegenerative diseases.While current therapies for Alzheimer’s disease, Parkinson’s disease and CTE provide acute symptomatic relief, neurodegeneration proceeds exponentially in the brain. Both oxidative stress and neuroinflammation precede overt symptomology of neurodegenerative disorders and each promotes neurotoxic environments associated with neurodegeneration. Dietary botanicals, in particular grape-derived polyphenols, have the potential to serve as an alternative treatment mode for the prevention of neurodegenerative disorders due to their ability to limit activation of oxidative pathways and inflammation following the exposure to stochastic environmental stressors. As reviewed, individual polyphenols have been shown both FH and SW contributed equally to the preparation and writing of the manuscript. JB assisted in the writing and composing of the manuscript. GMP reviewed and edited the contents of the manuscript.The authors acknowledge that the contents of this study do not represent the views of the NCCIH, the ODS, the NIH, the U.S. Department of Veterans Affairs, or the United States Government.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.


It is very significant to explore the intrinsic differences in breast cancer subtypes. These intrinsic differences are closely related to clinical diagnosis and designation of treatment plans. With the accumulation of biological and medicine datasets, there are many different omics data that can be viewed in different aspects. Combining these multiple omics data can improve the accuracy of prediction. Meanwhile; there are also many different databases available for us to download different types of omics data. In this article, we use estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) to define breast cancer subtypes and classify any two breast cancer subtypes using SMO-MKL algorithm. We collected mRNA data, methylation data and copy number variation (CNV) data from TCGA to classify breast cancer subtypes. Multiple Kernel Learning (MKL) is employed to use these omics data distinctly. The result of using three omics data with multiple kernels is better than that of using single omics data with multiple kernels. Furthermore; these significant genes and pathways discovered in the feature selection process are also analyzed. In experiments; the proposed method outperforms other state-of-the-art methods and has abundant biological interpretations. Breast cancer is the most general cancer diagnosed in women, and it is also the main cause of cancer death behind lung cancer . There aRecent studies have been directed at molecular classification of breast cancer. With the development of high-throughput research techniques such as microarray, genotyping can not only reflect the clinical characteristics of distinct types of breast cancer, but also provide technical support for the further study of breast cancer genotyping. The concept of tumor molecular typing was proposed by the National Cancer Institute (NCI) in 1999. Perou et al. in Stanford University firstly reported the molecular classification of breast cancer in 2000, and concluded that there are four primary subtypes including luminal subtype, basal-like subtype, human epidermal growth subtype and normal breast-like subtype . In 2003The classification of breast cancer subtypes is the basis for the diagnosis, the prognosis, the choice of treatment methods, and the conduct of various studies. In this article, breast cancer was classified into five subtypes based on immunohistochemistry (IHC) markers which include estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2). These subtypes are: luminal A, luminal B, HER2 (+), basal-like/Triple-negative breast cancer (TNBC), and Unclear . The detThere are many studies on breast cancer subtypes classification. Brian D. Lehmann et al. used cluster analysis to identify TNBC subtypes based on gene expression . ThereseSupport Vector Machine (SVM) is a supervised learning model, and it has been generally used in classification problems. It effectively addresses non-linearly separable problems by introducing the kernel function to map the features of data to a higher dimension, where data are linearly separable. The classical kernel functions include linear kernel, polynomial kernel and gaussian kernel. Subsequently, we can use Multiple Kernel Learning (MKL) to improve the classification accuracy of SVM . MKL canSMO-MKL is an improved supervised method based on linear MKL framework fusing heterogeneous omics data of breast cancer from the Cancer Genome Atlas (TCGA) . The worhttp://cancergenome.nih.gov/), which is a public database containing thousand samples of cancer patients, which profiles the various omics data of about 10,000 patient samples across over 30 cancer subtypes [In this article, we collected breast cancer data from The Cancer Genome Atlas (TCGA)  problem, and it needs to consume lots of time and memory to solve the QP problem. Sequential minimization optimization (SMO) is an efficient and easily applicable optimization method. SMO can resolve large QP problem into a series of smallest QP problems. Each resolved QP problems only have two Lagrange multipliers, so this can reduce the complexity of the large QP problem .norm regularization can lead to a sparse solution, and MKL which is the dual of standard p-norm is differentiable, and it can lead to a dense solution on kernel weight. SMO-MKL used the dual of standard p-norm MKL, and it utilized SMO style co-ordinate ascent to optimize. SMO-MKL can get a better result than original MKL using l-norm regularization [Multiple Kernel Learning algorithms are extensively used in multi-omics data fusion to improve the performance and interpretation . In thisproblems . MKL usirization . The kerIn this part, we firstly showed the accuracy and the area under the curve (AUC) of classification on any two breast cancer subtypes. Then, we achieved multi-classification by integrating multiple binary classifiers and compared with random forest with our model base on same datasets. Finally, we analyzed some significant genes and pathways.We combined different types of omics data to classify any two subtypes of breast cancer including: (1) luminal A versus luminal B, (2) luminal A versus TNBC, (3) luminal A versus HER2 (+), (4) luminal A versus Unclear, (5) luminal B versus TNBC, (6) luminal B versus HER2 (+), (7) luminal B versus Unclear, (8) TNBC versus HER2 (+), (9) TNBC versus Unclear, (10) HER2 (+) versus Unclear. We used mean accuracies of 10-fold cross-validation and AUCs as measurements of MKL model. Additionally, we calculated accuracies and AUCs with combining different kernels and omics data. These classification performances were shown in We can get the result that the optimal performance is Luminal A & HER2 (+) in accuracy and AUC. The MKL accuracy of HER2 (+) with other subtypes were all better than single omics data. The accuracy of the Luminal A with HER2 (+) and Unclear are better than Luminal B with these two breast cancer subtypes. Meanwhile, several studies are shown that the expression of ER-related genes in patients with Luminal A are higher than patients with Luminal B ,25. We aWe also used this algorithm to predict breast cancer subtypes. Because the method we used is a binary classification algorithm, we achieved multi-classification by integrating multiple binary classifiers. There are two strategies for turning to multinomial classifiers: One vs rest (OvR) and One vs one (OvO). We used OvO to turn this method to multi-classification since its, low sensitivity to the imbalanced data. The theory of OvO is to train http://cbc.mc.vanderbilt.edu/tnbc/) [Lehmann et al. used K-means clustering to classify TNBC patients into six clusters using gene expression data, and then defined these six clusters as six TNBC subtypes . These Tu/tnbc/) , and we We compared SMO-MKL with Random Forest and Neural Network on the same datasets. The reasons we chose Random Forest as compared method was that we can use random forest to utilize different omics data indirectly through its tree structure, and RF often used in multiple biological data integration . The RanTo better compare the performance of Random Forest and Neural Network with SMO-MKL, we applied the same settings in our model to random forest and Neural Network, and the result was achieved by 10-fold cross-validation of these approaches. The accuracy and AUC were shown in In this part, Firstly, we displayed the heatmap depicted by top 10 differential genes, and analyzed these significant genes further. Secondly, we analyzed pathways enriched by genes. The top 10 genes are ranked by p-values in decreasing order in each omics data. Heatmap is utilized to show all top 10 differential genes in any two classifications between Luminal A, Luminal B, TNBC and HER2 (+). These heatmap results were shown in Additionally, some significant genes were also selected to make further interpretation. In the classification of Luminal A versus Luminal B, the MED1 gene was related to HER2 (+) status of cancer in tissue microarray analysis . FurtherTo gain more insights into the differences in breast cancer subtypes, we separately performed enrichment analysis on the differential genes screened out from the omics data in any two subtypes between Luminal A, Luminal B, TNBC and HER2 (+). The pathways we mainly used were from KEGG and REACTOME. Moreover, we used the P-value obtained from the hypergeometric distribution to sort the pathways and selected top 30 to analysis categories of these pathways see . AlthougBreast cancer has many distinct subtypes, and these subtypes have different biological, clinical and molecular characteristics ,39. We aIn our article, the mRNA, methylation and CNV data from TCGA can efficiently classify the breast cancer subtypes using ER, PR, HER2 defined. We also used Random Forest and Neural Network to compare the result with SMO-MKL, and we found that the performance of SMO-MKL was superior to Random Forest and Neural Network. We further applied the SMO-MKL model to predict breast cancer subtypes, and the result also showed that integrated multi-omics data can better separate breast cancer subtypes. In conclusion, the result of any two breast cancer subtypes indicated the multiple omics data can improve the accuracy and AUC in breast cancer subtypes classification using SMO-MKL.The parameters in polynomial kernel and gaussian kernel also could affect the performance of breast cancer subtypes prediction, which meant that we could improve the accuracy through rationally tuning parameters. As such, we used grid search to select the optimal parameters in polynomial kernel and Gaussian kernel individually in any two breast cancer subtypes.We searched many papers related to the genes which were have lower p-value in feature selection, and we found many genes related to breast cancer subtypes. and we also observed some selected genes shared by two or more classifications, like GATA3 and GRB7. GATA3 is overexpressed in luminal epithelial cells, and low expression of GATA3 is connected negative ER and PR, so it showed significantly different expression in Luminal A versus TNBC and Luminal B versus TNBC. GRB7 is associated with the status of Her2. We can clearly distinguish the status of Her2 in tumors under the protein expression of GRB7 . Moreover, we can further boost our model’s performance in classifying breast cancer subtypes which are based on ER, PR and Her2. We only simply classified breast cancer subtypes utilizing ER, PR and Her2, but they also have some other factors in the clinical diagnosis, like Ki-67 . TherefoThe purpose of this study was to classify breast cancer subtypes by using SMO-MKL models and multiple omics datasets from TCGA, and then analyze differences between distinct breast cancer subtypes in many aspects. The classification results showed that integrating multiple omics datasets can improve the accuracy than using single omics data through SMO-MKL models. Furthermore, we achieved multi-classification on breast cancer subtypes and TNBC subtypes by using SMO-MKL models, and the classification results also showed that the accuracy of using multiple omics data were higher than single omics data. Meanwhile, we tried to give some biological explanation for the differences between breast cancer subtypes by analyzing the significant genes and pathways, which also provide guidance for exploring the biological models on breast cancer subtypes.


Toxoplasma gondii is a successful protozoan parasite that cycles between definitive felid hosts and a broad range of intermediate hosts, including rodents and humans. Within intermediate hosts, this obligate intracellular parasite invades the small intestine, inducing an inflammatory response. Toxoplasma infects infiltrating immune cells, using them to spread systemically and reach tissues amenable to chronic infection. An intact immune system is necessary to control life-long chronic infection. Chronic infection is characterized by formation of parasite cysts, which are necessary for survival through the gastrointestinal tract of the next host. Thus, Toxoplasma must evade sterilizing immunity, but still rely on the host's immune response for survival and transmission. To do this, Toxoplasma exploits a central cost-benefit tradeoff in immunity: the need to escalate inflammation for pathogen clearance vs. the need to limit inflammation-induced bystander damage. What are the consequences of sustained inflammation on host biology? Many studies have focused on aspects of the immune response that directly target Toxoplasma growth and survival, commonly referred to as “resistance mechanisms.” However, it is becoming clear that a parallel arm of the immune response has evolved to mitigate damage caused by the parasite directly  or bystander damage due to the inflammatory response . These so-called “disease tolerance” mechanisms promote tissue function and host survival without directly targeting the pathogen. Here we review changes to host metabolism, tissue structure, and immune function that point to disease tolerance mechanisms during Toxoplasma infection. We explore the impact tolerance programs have on the health of the host and parasite biology. A successful immune response requires two distinct, but complementary components: “resistance mechanisms” and “tolerance mechanisms.” Resistance (or restriction) mechanisms directly target pathogens to limit microbial replication or dissemination. Examples of resistance mechanisms include antimicrobial peptides, complement, and degranulation by neutrophils or cytotoxic T cells. Tolerance mechanisms target host cell biology to improve tissue integrity and function in the setting of damage caused by the pathogen or the inflammatory response. Examples include extracellular matrix remodeling, the DNA damage response, antioxidant production, and shifts in cell metabolism  in peripheral tissues or in lymph nodes. Regulatory T cells (Tregs) play an important role in peripheral tolerance. Central and peripheral lymphocyte tolerance are included under the broader umbrella of so-called “disease tolerance” strategies because they are mechanisms of limiting inflammation and preventing auto-immunity but do not directly influence the pathogen. In this review, we refer to the broader definition of “disease tolerance” as mechanisms that support host fitness and survival during infection, not by directly targeting pathogen biology, but by shifting homeostasis to maintain tissue function during infection and inflammation  without increasing susceptibility to pathogens. Host survival during infection depends having both tolerance and resistance mechanisms, otherwise the host will succumb either to pathogen overgrowth or lethal immunopathology. As discussed in the following sections, it is also important to consider how promoting tolerance biology may benefit the life cycle of pathogens, particularly those that have evolved a strategy for long-term persistence within a host .Toxoplasma gondii is an obligate intracellular protozoan parasite that establishes life-long infection in a wide range of warm-blooded intermediate hosts, including rodents and humans. Three major haplotypes, types I–III, dominate Europe and North America which differ in virulence by several logs. Feline definitive hosts support Toxoplasma sexual recombination and shed millions of environmentally stable, highly infectious oocysts in feces. This confers a tremendous benefit for the parasite in terms of genetic diversity and dissemination potential, which suggests that rodents may be an important intermediate host for Toxoplasma given the predator-prey relationship between the two host organisms.Toxoplasma oocysts or tissue cysts, the so-called bradyzoite form, from a previously infected intermediate host. Toxoplasma invades the small intestine, converting to the rapidly dividing tachyzoite form and inducing inflammation  infection  during Toxoplasma infection  Toxoplasma infection. These casts which appeared to limited bacterial translocation into host circulation. Depleting neutrophils with an α-Gr-1 antibody increased mortality without increasing parasite burden, indicating that the immune-commensal axis also plays an important role in disease tolerance during Toxoplasma infection  which limited neutrophil-induced pathology. Administration of a PGE2 analog during infection was sufficient to reduce intestinal tissue pathology and immune infiltration in the absence of monocytes without affecting parasite burden  i.p. infected with 20 ME49 cysts succumbed by 13 days post-infection. No change in parasite burden was detected between knockout and wild type mice in peritoneal lavage fluid at 7 days post-infection, but knockout mice had elevated circulating IL-12, IFN-γ, increased inflammation and necrosis in the liver and lungs, and hyperactive and hyperproliferative CD4+ T cells in the spleen. Survival was partially rescued when CD4+ T cells were depleted  and tissue resident retinal cells expressed PD-L1, the ligand for the T cell inhibitory receptor PD-1. PD-L1 expressing retinal cells were able to suppress splenic T cell proliferation ex vivo using a co-culture system with Toxoplasma-antigen loaded dendritic cells , availability and storage (fat and glycogen), and detoxifies the blood. Liver resident macrophages called Kupffer cells screen incoming blood for pathogens and intestinal microbes that leak from the gut during intestinal inflammation, including during Toxoplasma infection (Molloy et al., Toxoplasma has been detected in the livers of mice at acute infection using bioluminescence, histology, and PCR in a number of studies using Type II strains (Silva et al., Toxoplasma has been observed replicating in hepatocytes near regions of inflammatory infiltrate and focal necrosis in both Swiss-Webster and BALB/c mice (Atmaca et al., Toxoplasma infection (Atmaca et al., Toxoplasma infection. However, an important caveat with these studies is that they were performed with lethal doses of the hypervirulent Type I strain RH raising the question of relevance to hepatic infection with strains that are more commonly found in mice or humans. Using clinically isolated Type II strains, Toxoplasma cysts were found in the livers of infected Swiss-Webster mice as late as 33 weeks post-infection, indicating that the liver may be a reservoir for chronic infection (Autier et al., Toxoplasma-negative transplant recipients who have developed toxoplasmosis after receiving livers from sera positive donors (Assi et al., Toxoplasma in the development of liver disease, approximately 30% of patients with chronic liver disease test sera-positive for Toxoplasma B1 compared to 6% in control populations (El-Sayed et al., Toxoplasma-infected patients had significantly elevated circulating ALT and AST, clinical markers of liver damage, compared to uninfected patients with liver disease. Although these data do not imply causality, they are consistent with the interpretation that chronic Toxoplasma infection may negatively impact host fitness in diseases of co-occurrence.Toxoplasma infection may also change the metabolic landscape of the liver. In a recent liver proteomics study, BALB/c mice intraperitoneally infected with 200 type II PYS tachyzoites had reduced signatures of fatty-acid oxidation proteins and an upregulation of cell death, inflammatory, and stress response pathways at 6 days post-infection (He et al., Toxoplasma occupies a liver niche in acute and chronic infection and may directly contribute to shifts in liver metabolic homeostasis.M. tuberculosis, T. brucei, and facultative pathogen strains of E. coli have been detected in this niche, indicating that it serves as a reservoir of infection for many pathogens (Neyrolles et al., Toxoplasma has been reported in the visceral fat following intraperitoneal infection and oral infection by bioluminescence assay and PCR (Di Cristina et al., Toxoplasma, and whether colonization of the fat is related to host metabolic shifts.Adipose tissue depots are another important site of calorie storage and immune regulation. Fat tissues have long been described as an anti-inflammatory environment that become inflamed in diseases of the adipose tissue, including obesity and diabetes. As better tools have become available to survey low pathogen loads in tissue, researchers have begun to appreciate the frequency of microbial translocation to and persistence in this nutrient-rich environment. Toxoplasma gondii has evolved sophisticated mechanisms to evade sterilizing immunity, yet activating a robust immune response is necessary to ensure host survival long enough for Toxoplasma encystation and transmission (Bohne et al., Toxoplasma infection, including immune-microbiota interactions, T cell-mediated responses and metabolic shifts fail to return to homeostasis in chronic infection. Emerging studies suggest that these shifts in homeostasis have sustained negative consequences for the host, including muscle wasting, and impaired responses to secondary immune stimuli (Arsenijevic et al., Toxoplasma because feline hosts facilitate genetic diversity and range expansion. In this way, Toxoplasma may benefit from promoting tolerance programs that ensure host survival during the acute phase of infection, during the tachyzoite to bradyzoite transition, but ultimately impair host fitness in the long term.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 advent of single-nucleus RNA-sequencing (snRNAseq) has allowed for the exploration of genetic signatures of the numerous cells in the brain. In particular, snRNAseq data can provide new insights into how many neurodegenerative diseases, such as Alzheimer’s Disease, alter cells in the brain. One major challenge with analyzing snRNAseq data is the lack of a systematic way to classify the various cell types across different datasets. To address this challenge, we developed a general classifier (“DeepSeq”) that uses state-of-the-art deep learning approaches. We trained our model on multiple snRNAseq datasets derived from post-mortem brain tissue in individuals with and without clinical diagnosis of Alzheimer’s Disease from the ROSMAP cohorts. The two snRNAseq datasets contained 70,064 nuclei and 170,275 nuclei. The two studies employed different clustering techniques, and identified 44 and 18 putative cell types. To map these disparate cluster identities across datasets, we extracted the most relevant genes and trained two separate networks, one on each dataset. We then validated each classifier separately on the holdout cells. The resulting classifier accuracy were 87% and 94%. To map clusters across datasets, we then applied each classifier to the other dataset. Both classifiers yielded mappings that reflected the overall biology, correctly categorizing the nuclei into broad and fine cell type classes. Although validation on additional datasets would expand the generality of this approach, our results show that DeepSeq is an easily implementable classification tool that can assign identity to nuclei in new snRNAseq datasets without the need for preprocessing or cross-batch alignment.