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Abstract Previous studies have adopted an edge-centric framework to study fine-scale network dynamics in human fMRI. To date, however, no studies have applied this framework to data collected from model organisms. Here, we analyze structural and functional imaging data from lightly anesthetized mice through an edge-centric lens. We find evidence of “bursty” dynamics and events - brief periods of high-amplitude network connectivity. Further, we show that on a per-frame basis events best explain static FC and can be divided into a series of hierarchically-related clusters. The co-fluctuation patterns associated with each cluster centroid link distinct anatomical areas and largely adhere to the boundaries of algorithmically detected functional brain systems. We then investigate the anatomical connectivity undergirding high-amplitude co-fluctuation patterns. We find that events induce modular bipartitions of the anatomical network of inter-areal axonal projections. Finally, we replicate these same findings in a human imaging dataset. In summary, this report recapitulates in a model organism many of the same phenomena observed in previously edge-centric analyses of human imaging data. However, unlike human subjects, the murine nervous system is amenable to invasive experimental perturbations. Thus, this study sets the stage for future investigation into the causal origins of fine-scale brain dynamics and high-amplitude co-fluctuations. Moreover, the cross-species consistency of the reported findings enhances the likelihood of future translation.

Circulating tumor cells (CTCs) currently represent a revolutionary tool offering unique insights for the evaluation of cancer progression, metastasis, and response to therapies. Indeed, CTCs, upon detachment from primary tumors, enter the bloodstream and acquire a great potential for their use for personalized cancer management. In this review, we describe the current understanding of and advances in the clinical employment of CTCs. Although considered rare and fleeting, CTCs are now recognized as key players favoring the development of cancer metastasis and disease recurrence, particularly in malignant melanoma, lung, breast, and colorectal cancer patients. To date, the advancements in technology and the development of several successful approaches, also including immunomagnetic enrichment allow for a reliable and reproducible detection and characterization of CTCs. Those innovative methodologies improved the isolation, quantification, and characterization of CTCs from the blood of cancer patients, providing extremely useful evidence and new insights into the nature of the tumor, its epithelial/mesenchymal profile, and its potential resistance to therapy. In fact, in addition to their prognostic and predictive value, CTCs could serve as a valuable instrument for real-time monitoring of treatment response and disease recurrence, facilitating timely interventions and thus improving patient outcomes. However, despite their potential, several challenges hinder the widespread clinical utility of CTCs: (i) CTCs’ rarity and heterogeneity pose technical limitations in isolation and characterization, as well as significant hurdles in their clinical implementation; (ii) it is mandatory to standardize CTC detection methods, optimize the sample processing techniques, and integrate them with existing diagnostic modalities; and (iii) the need for the development of new techniques, such as single-cell analysis platforms, to enhance the sensitivity and specificity of CTC detection, thereby facilitating their integration into routine clinical practice. In conclusion, CTCs represent a potential extraordinary tool in cancer diagnostics and therapeutics, offering unprecedented opportunities for personalized medicine and precision oncology. Moreover, their ability to provide real-time insights into tumor biology, treatment response, and disease progression underlines a great potential for their clinical application to improve patients’ outcomes and advance our understanding of cancer biology.

Glaucoma is chronic optic neuropathy whose pathogenesis has been associated with the altered metabolism of Trabecular Meshwork Cells, which is a cell type involved in the synthesis and remodeling of the trabecular meshwork, the main drainage pathway of the aqueous humor. Starting from previous findings supporting altered ubiquitin signaling, in this study, we investigated the ubiquitin-mediated turnover of myocilin (MYOC/TIGR gene), which is a glycoprotein with a recognized role in glaucoma pathogenesis, in a human Trabecular Meshwork strain cultivated in vitro in the presence of dexamethasone. This is a validated experimental model of steroid-induced glaucoma, and myocilin upregulation by glucocorticoids is a phenotypic marker of Trabecular Meshwork strains. Western blotting and native-gel electrophoresis first uncovered that, in the presence of dexamethasone, myocilin turnover by proteasome particles was slower than in the absence of the drug. Thereafter, co-immunoprecipitation, RT-PCR and gene-silencing studies identified STUB1/CHIP as a candidate E3-ligase of myocilin. In this regard, dexamethasone treatment was found to downregulate STUB1/CHIP levels by likely promoting its proteasome-mediated turnover. Hence, to strengthen the working hypothesis about global alterations of ubiquitin-signaling, the first profiling of TMCs ubiquitylome, in the presence and absence of dexamethasone, was here undertaken by diGLY proteomics. Application of this workflow effectively highlighted a robust dysregulation of key pathways (e.g., phospholipid signaling, β-catenin, cell cycle regulation) in dexamethasone-treated Trabecular Meshwork Cells, providing an ubiquitin-centered perspective around the effect of glucocorticoids on metabolism and glaucoma pathogenesis.

Crohn’s disease (CD) is a type of inflammatory bowel disease (IBD) affecting the gastrointestinal tract that can also cause extra-intestinal complications. Following exposure to the mRNA vaccine BNT162b2 (Pfizer-BioNTech) encoding the SARS-CoV-2 Spike (S) protein, some patients experienced a lack of response to the biological drug Adalimumab and a recrudescence of the disease. In CD patients in progression, resistant to considered biological therapy, an abnormal increase in intestinal permeability was observed, more often with a modulated expression of different proteins such as Aquaporin 8 (AQP8) and in tight junctions (e.g., ZO-1, Claudin1, Claudin2, Occludin), especially during disease flares. The aim of this study is to investigate how the SARS-CoV-2 vaccine could interfere with IBD therapy and contribute to disease exacerbation. We investigated the role of the SARS-CoV-2 Spike protein, transported by extracellular vesicles (EVs), and the impact of various EVs components, namely, exosomes (EXOs) and microvesicles (MVs), in modulating the expression of molecules involved in the exacerbation of CD, which remains unknown.

Irritable bowel syndrome (IBS) involves low-grade mucosal inflammation. Among the various approaches capable of managing the symptoms, physical activity is still under investigation. Despite its benefits, it promotes oxidative stress and inflammation. Mitochondria impacts gut disorders by releasing damage-associated molecular patterns, such as cell-free mtDNA (cf-mtDNA), which support inflammation. This study evaluated the effects of a 12-week walking program on the cf-mtDNA and DNase in 26 IBS and 17 non-IBS subjects. Pro- and anti-inflammatory cytokines were evaluated by ELISA. Digital droplet PCR was used to quantify cf-mtDNA; DNase activity was assessed using a single radial enzyme diffusion assay. PCR-RFLP was used to genotype DNASE1 rs1053874 SNP. Significantly lower IL-10 levels were found in IBS than in non-IBS individuals. Exercise reduced cf-mtDNA in non-IBS subjects but not in IBS patients. DNase activity did not correlate with the cf-mtDNA levels in IBS patients post-exercise, indicating imbalanced cf-mtDNA clearance. Different rs1053874 SNP frequencies were not found between groups. The study confirms the positive effects of regular moderate-intensity physical activity in healthy subjects and its role in cf-mtDNA release and clearance. Walking alone might not sufficiently reduce subclinical inflammation in IBS, based on imbalanced pro- and anti-inflammatory molecules. Prolonged programs are necessary to investigate their effects on inflammatory markers in IBS.

Inherited defects in cytochrome c oxidase (COX) are associated with a substantial subset of diseases adversely affecting the structure and function of the mitochondrial respiratory chain. This multi-subunit enzyme consists of 14 subunits and numerous cofactors, and it requires the function of some 30 proteins to assemble. COX assembly was first shown to be the primary defect in the majority of COX deficiencies 36 years ago. Over the last three decades, most COX assembly genes have been identified in the yeast Saccharomyces cerevisiae, and studies in yeast have proven instrumental in testing the impact of mutations identified in patients with a specific COX deficiency. The advent of accessible genome-wide sequencing capabilities has led to more patient mutations being identified, with the subsequent identification of several new COX assembly factors. However, the lack of genotype–phenotype correlations and the large number of genes involved in generating a functional COX mean that functional studies must be undertaken to assign a genetic variant as being causal. In this review, we provide a brief overview of the use of yeast as a model system and briefly compare the COX assembly process in yeast and humans. We focus primarily on the studies in yeast that have allowed us to both identify new COX assembly factors and to demonstrate the pathogenicity of a subset of the mutations that have been identified in patients with inherited defects in COX. We conclude with an overview of the areas in which studies in yeast are likely to continue to contribute to progress in understanding disease arising from inherited COX deficiencies.

Oral squamous cell carcinoma (OSCC) is the most common and lethal type of head and neck cancer in the world. Variable response and acquisition of resistance to traditional therapies show that it is essential to develop novel strategies that can provide better outcomes for the patient. Understanding of cellular and molecular mechanisms of cell death control has increased rapidly in recent years. Activation of cell death pathways, such as the emerging forms of non-apoptotic programmed cell death, including ferroptosis, pyroptosis, necroptosis, NETosis, parthanatos, mitoptosis and paraptosis, may represent clinically relevant novel therapeutic opportunities. This systematic review summarizes the recently described forms of cell death in OSCC, highlighting their potential for informing diagnosis, prognosis and treatment. Original studies that explored any of the selected cell deaths in OSCC were included. Electronic search, study selection, data collection and risk of bias assessment tools were realized. The literature search was carried out in four databases, and the extracted data from 79 articles were categorized and grouped by type of cell death. Ferroptosis, pyroptosis, and necroptosis represented the main forms of cell death in the selected studies, with links to cancer immunity and inflammatory responses, progression and prognosis of OSCC. Harnessing the potential of these pathways may be useful in patient-specific prognosis and individualized therapy. We provide perspectives on how these different cell death types can be integrated to develop decision tools for diagnosis, prognosis, and treatment of OSCC.

Age-associated DNA methylation in blood cells convey information on health status. However, the mechanisms that drive these changes in circulating cells and their relationships to gene regulation are unknown. We identified age-associated DNA methylation sites in six purified blood-borne immune cell types (naive B, naive CD4+ and CD8+ T cells, granulocytes, monocytes, and NK cells) collected from healthy individuals interspersed over a wide age range. Of the thousands of age-associated sites, only 350 sites were differentially methylated in the same direction in all cell types and validated in an independent longitudinal cohort. Genes close to age-associated hypomethylated sites were enriched for collagen biosynthesis and complement cascade pathways, while genes close to hypermethylated sites mapped to neuronal pathways. In silico analyses showed that in most cell types, the age-associated hypo- and hypermethylated sites were enriched for ARNT (HIF1β) and REST transcription factor (TF) motifs, respectively, which are both master regulators of hypoxia response. To conclude, despite spatial heterogeneity, there is a commonality in the putative regulatory role with respect to TF motifs and histone modifications at and around these sites. These features suggest that DNA methylation changes in healthy aging may be adaptive responses to fluctuations of oxygen availability.

Background: Hepatocellular carcinoma (HCC) accounts for more than 75% of primary liver cancers, which are the second leading cause of cancer-related deaths. The GALAD (gender, age, AFP-L3, AFP, and des-carboxy-prothrombin) score is a diagnostic tool developed based on gender, age, alpha-fetoprotein, alpha-fetoprotein L3, and des-gamma-carboxy prothrombin, originally designed as a diagnostic tool for HCC in high-risk patients. Methods: We analyzed 212 patients with and without cirrhosis. The population study was divided into patients with liver cirrhosis without evidence of HCC at the time of serum sample collection for GALAD score determination and patients with liver cirrhosis and a confirmed diagnosis of HCC at the time of serum sample collection for GALAD score determination. Patients were followed up until death or liver transplantation. The association between variables and HCC mortality risk was performed, and the results were presented as hazard ratio (HR). The receiver operating characteristic (ROC) curve was used to assess the performance of the GALAD HCC diagnosis. The survival probability was explored using the non-parametric test, and the equality of survival amongst categories was assessed with the log-rank test. Results: Biomarkers were higher in the HCC group compared to cirrhosis. Kaplan–Meier survival probability analysis for individual GALAD categories revealed that a high GALAD level was associated with decreased survival during follow-up, and the difference between the curves was statistically significant (p = 0.01). Conclusions: Our findings suggest that the GALAD score has promise as a prognostic tool, with implications for improving patient management and treatment strategies for HCC.

Abstract The Gsα/cAMP signaling pathway mediates the effect of a variety of hormones and factors that regulate the homeostasis of the post-natal skeleton. Hence, the dysregulated activity of Gsα due to gain-of-function mutations (R201C/R201H) results in severe architectural and functional derangements of the entire bone/bone marrow organ. While the consequences of gain-of-function mutations of Gsα have been extensively investigated in osteoblasts and in bone marrow osteoprogenitor cells at various differentiation stages, their effect in adipogenically-committed bone marrow stromal cells has remained unaddressed. We generated a mouse model with expression of Gsα R201C driven by the Adiponectin (Adq) promoter. Adq-Gsα R201C mice developed a complex combination of metaphyseal, diaphyseal and cortical bone changes. In the metaphysis, Gsα R201C caused an early phase of bone resorption followed by bone deposition. Metaphyseal bone formation was sustained by cells that were traced by Adq-Cre and eventually resulted in a high trabecular bone mass phenotype. In the diaphysis, Gsα R201C , in combination with estrogen, triggered the osteogenic activity of Adq-Cre-targeted perivascular bone marrow stromal cells leading to intramedullary bone formation. Finally, consistent with the previously unnoticed presence of Adq-Cre-marked pericytes in intraosseous blood vessels, Gsα R201C caused the development of a lytic phenotype that affected both cortical (increased porosity) and trabecular (tunneling resorption) bone. These results provide the first evidence that the Adq-cell network in the skeleton not only regulates bone resorption but also contributes to bone formation, and that the Gsα/cAMP pathway is a major modulator of both functions.

The gut microbiota has gained increasing attention in recent years due to its significant impact on colorectal cancer (CRC) development and progression. The recent detection of bacterial DNA load in plasma holds promise as a potential non-invasive approach for early cancer detection. The aim of this study was to examine the quantity of bacterial DNA present in the plasma of 50 patients who have CRC in comparison to 40 neoplastic disease-free patients, as well as to determine if there is a correlation between the amount of plasma bacterial DNA and various clinical parameters. Plasma bacterial DNA levels were found to be elevated in the CRC group compared to the control group. As it emerged from the logistic analysis (adjusted for age and gender), these levels were strongly associated with the risk of CRC (OR = 1.02, p < 0.001, 95% C.I.: 1.01–1.03). Moreover, an association was identified between a reduction in tumor mass and the highest tertile of plasma bacterial DNA. Our findings indicate that individuals with CRC displayed a higher plasma bacterial DNA load compared to healthy controls. This observation lends support to the theory of heightened bacterial migration from the gastrointestinal tract to the bloodstream in CRC. Furthermore, our results establish a link between this phenomenon and the size of the tumor mass.

Background: Mitochondrial DNA copy number (mtDNAcn) in tissues and blood can be altered in conditions like diabetes and major depression and may play a role in aging and longevity. However, little is known about the association between mtDNAcn and personality traits linked to emotional states, metabolic health, and longevity. This study tests the hypothesis that blood mtDNAcn is related to personality traits and mediates the association between personality and mortality. Methods: We assessed the big five personality domains and facets using the Revised NEO Personality Inventory (NEO-PI-R), assessed depressive symptoms with the Center for Epidemiologic Studies Depression Scale (CES-D), estimated mtDNAcn levels from whole-genome sequencing, and tracked mortality in participants from the Baltimore Longitudinal Study of Aging. Results were replicated in the SardiNIA Project. Results: We found that mtDNAcn was negatively associated with the Neuroticism domain and its facets and positively associated with facets from the other four domains. The direction and size of the effects were replicated in the SardiNIA cohort and were robust to adjustment for potential confounders in both samples. Consistent with the Neuroticism finding, higher depressive symptoms were associated with lower mtDNAcn. Finally, mtDNAcn mediated the association between personality and mortality risk. Conclusions: To our knowledge, this is the first study to show a replicable association between mtDNAcn and personality. Furthermore, the results support our hypothesis that mtDNAcn is a biomarker of the biological process that explains part of the association between personality and mortality. Funding: Support for this work was provided by the Intramural Research Program of the National Institute on Aging (Z01-AG000693, Z01-AG000970, and Z01-AG000949) and the National Institute of Neurological Disorders and Stroke of the National Institutes of Health. AT was also supported by the National Institute on Aging of the National Institutes of Health Grant R01AG068093.

Nitrobindins (Nbs) are all-β-barrel heme proteins spanning from bacteria to Homo sapiens. They inactivate reactive nitrogen species by sequestering NO, converting NO to HNO2, and promoting peroxynitrite isomerization to NO3−. Here, the nitrite reductase activity of Nb(II) from Mycobacterium tuberculosis (Mt-Nb(II)), Arabidopsis thaliana (At-Nb(II)), Danio rerio (Dr-Nb(II)), and Homo sapiens (Hs-Nb(II)) is reported. This activity is crucial for the in vivo production of NO, and thus for the regulation of blood pressure, being of the utmost importance for the blood supply to poorly oxygenated tissues, such as the eye retina. At pH 7.3 and 20.0 °C, the values of the second-order rate constants (i.e., kon) for the reduction of NO2− to NO and the concomitant formation of nitrosylated Mt-Nb(II), At-Nb(II), Dr-Nb(II), and Hs-Nb(II) (Nb(II)-NO) were 7.6 M−1 s−1, 9.3 M−1 s−1, 1.4 × 101 M−1 s−1, and 5.8 M−1 s−1, respectively. The values of kon increased linearly with decreasing pH, thus indicating that the NO2−-based conversion of Nb(II) to Nb(II)-NO requires the involvement of one proton. These results represent the first evidence for the NO2 reductase activity of Nbs(II), strongly supporting the view that Nbs are involved in NO metabolism. Interestingly, the nitrite reductase reactivity of all-β-barrel Nbs and of all-α-helical globins (e.g., myoglobin) was very similar despite the very different three-dimensional fold; however, differences between all-α-helical globins and all-β-barrel Nbs suggest that nitrite reductase activity appears to be controlled by distal steric barriers, even though a more complex regulatory mechanism can be also envisaged.

Immunotherapy in oncology is replacing traditional therapies due to it specific action and limited side effects. Despite the high efficacy of immunotherapy, side effects such as bacterial infection have been reported. Bacterial skin and soft tissue infections represent one of the most important differential diagnoses in patients presenting with reddened and swollen skin and soft tissue. Among these infections, cellulitis (phlegmon) and abscesses are the most frequent. In most cases, these infections occur locally with possible contiguous spread, or as a multifocal manifestation, especially in immunocompromised patients. Herein, we report a case of pyodermitis in an immunocompromised district in a patient treated with nivolumab for non-small cell lung cancer. A 64-year-old, smoker male patient showed cutaneous lesions at a different evolution level in the left arm, all in a tattooed area, with one phlegmon and two ulcerated lesions. Microbiological cultures and gram staining revealed an infection caused by a methicillin-susceptible but erythromycin-resistant (ER-R), clindamycin-resistant (CL-R), and gentamicin-resistant (GE-R) Staphylococcus aureus strain. Despite immunotherapy becoming a milestone in oncologic treatment, more than the spectrum of immune-mediated toxicities of these agents needs to be investigated. This report highlights the importance of considering lifestyle and cutaneous background before starting immunotherapy for cancer treatment, with an emphasis on pharmacogenomics and the possibility of modified skin microbiota predisposing to cutaneous infections in patients treated with PD-1 inhibitors.

Paracoccidioidomycosis (PCM) is the main and most prevalent systemic mycosis in Latin America, that until recently, it was believed to be caused only by Paracoccidioides brasiliensis (P. brasiliensis). In 2006, researchers described three cryptic species: S1, PS2, PS3, and later, another one, PS4. In 2009, Paracoccidioides lutzii (Pb01-like) was described, and in 2017, a new nomenclature was proposed for the different agents: P. brasiliensis (S1), P. americana (PS2), P. restrepiensis (PS3), and P. venezuelensis (PS4). These species are not uniformly distributed throughout Latin America and, knowing that more than one cryptic species could coexist in some regions, we aimed to identify those species in patients’ biopsy samples for a better understanding of the distribution and occurrence of these recently described species in Botucatu region. The Hospital of Medical School of Botucatu—UNESP, which is a PCM study pole, is located in São Paulo State mid-west region and is classified as a PCM endemic area. Genotyping analyses of clinical specimens from these patients that have been diagnosed and treated in our Hospital could favor a possible correlation between genetic groups and mycological and clinical characteristics. For this, molecular techniques to differentiate Paracoccidioides species in these biopsies, such as DNA extraction, PCR, and sequencing of three target genes (ITS, CHS2, and ARF) were conducted. All the sequences were analyzed at BLAST to testify the presence of P. brasiliensis. The phylogenetic trees were constructed using Mega 7.0 software and showed that 100% of our positive samples were from S1 cryptic species, therefore P. brasiliensis. This is important data, demonstrating the predominance of this species in the São Paulo State region.

Abstract Crop genomics has seen dramatic advances in recent years due to improvements in sequencing technology, assembly methods, and computational resources. These advances have led to the development of new tools to facilitate crop improvement. The study of structural variation within species and the characterization of the pan-genome has revealed extensive genome content variation among individuals within a species that is paradigm shifting to crop genomics and improvement. Here, we review advances in crop genomics and how utilization of these tools is shifting in light of pan-genomes that are becoming available for many crop species.

Given the lack of advances in Oral Squamous Cell Carcinoma (OSCC) therapy in recent years, pharmacological strategies to block OSCC-related signaling pathways have gained prominence. The present study aimed to evaluate the therapeutic potential of Arsenic Trioxide (ATO) concerning its antitumoral effects and the inhibition of the Hedgehog (HH) pathway in OSCC. Initially, ATO cytotoxicity was assessed in a panel of cell lines. Cell viability, cell cycle, death patterns, and cell morphology were analyzed, as well as the effect of ATO on the expression of HH pathway components. After the cytotoxic assay, HSC3 cells were chosen for all in vitro assays. ATO increased apoptotic cell death and nuclear fragmentation in the sub-G1 cell cycle phase and promoted changes in cell morphology. In addition, the reduced expression of GLI1 indicated that ATO inhibits HH activity. The present study provides evidence of ATO as an effective cytotoxic drug for oral cancer treatment in vitro.

The interaction between malignant cells and the tumor microenvironment is critical for tumor progression, and the chemokine ligand/receptor axes play a crucial role in this process. The CXCR4/CXCL12 and CCR5/CCL5 axes, both related to HIV, have been associated with the early (epithelial–mesenchymal transition and invasion) and late events (migration and metastasis) of cancer progression. In addition, these axes can also modulate the immune response against tumors. Thus, antagonists against the receptors of these axes have been proposed in cancer therapy. Although preclinical studies have shown promising results, clinical trials are needed to include these drugs in the oncological treatment protocols. New alternatives for these antagonists, such as dual CXCR4/CCR5 antagonists or combined therapy in association with immunotherapy, need to be studied in cancer therapy.

Auxetic materials can be exploited for coupling different types of tissues. Herein, we designed a material where the microorganism metabolic activity yields the formation of buckled/collapsed bubbles within gelling silicone cylinders thus providing auxetic properties. The finite element model of such hollow auxetic cylinders demonstrated the tubular structure to promote worm-like peristalsis. In this scenario, the described hybrid auxetic structures may be applied to the longitudinal intestinal lengthening and tailoring procedure to promote enteral autonomy in short bowel syndrome. The presented material and analytical design synergistic approach offer a pioneering step for the clinical translation of hybrid auxetic materials.

The identification of genetic risk factors for non-syndromic oral clefts is of great importance for better understanding the biological processes related to this heterogeneous and complex group of diseases. Herein we applied whole-exome sequencing to identify potential variants related to non-syndromic cleft palate only (NSCPO) in the multiethnic Brazilian population. Thirty NSCPO samples and 30 sex- and genetic ancestry-matched healthy controls were pooled (3 pools with 10 samples for each group) and subjected to whole-exome sequencing. After filtering, the functional affects, individually and through interactions, of the selected variants and genes were assessed by bioinformatic analyses. As a group, 399 variants in 216 genes related to palatogenesis/cleft palate, corresponding to 6.43%, were exclusively identified in the NSCPO pools. Among those genes are 99 associated with syndromes displaying cleft palate in their clinical spectrum and 92 previously related to cleft lip palate. The most significantly biological processes and pathways overrepresented in the NSCPO-identified genes were associated with the folic acid metabolism, highlighting the interaction between LDL receptor-related protein 6 (LRP6) and 5-methyltetrahydrofolate-homocysteine methyltransferase (MTR) that interconnect two large networks. This study yields novel data on characterization of specific variants and complex processes and pathways related to NSCPO, including many variants in genes of the folate/homocysteine pathway, and confirms that variants in genes related to syndromic cleft palate and cleft lip-palate may cause NSCPO.

Intrahepatic cholangiocarcinoma (iCCA) is a highly aggressive cancer with limited therapeutic options and short overall survival. iCCA is characterized by a strong desmoplastic reaction in the surrounding ecosystem that likely affects tumoral progression. Overexpression of the Notch pathway is implicated in iCCA development and progression. Our aim was to investigate the effectiveness of Crenigacestat, a selective inhibitor of NOTCH1 signaling, against the cross-talk between cancer cells and the surrounding ecosystem in an in vivo HuCCT1-xenograft model. In the present study, a transcriptomic analysis approach, validated by Western blotting and qRT-PCR on iCCA tumor masses treated with Crenigacestat, was used to study the molecular pathways responsive to drug treatment. Our results indicate that Crenigacestat significantly inhibited NOTCH1 and HES1, whereas tumor progression was not affected. In addition, the drug triggered a strong immune response and blocked neovascularization in the tumor ecosystem of the HuCCT1-xenograft model without affecting the occurrence of fibrotic reactions. Therefore, although these data need further investigation, our observations confirm that Crenigacestat selectively targets NOTCH1 and that the desmoplastic response in iCCA likely plays a key role in both drug effectiveness and tumor progression.

Steroid-induced glaucoma is a severe pathological condition, sustained by a rapidly progressive increase in intraocular pressure (IOP), which is diagnosed in a subset of subjects who adhere to a glucocorticoid (GC)-based therapy. Molecular and clinical studies suggest that either natural or synthetic GCs induce a severe metabolic dysregulation of Trabecular Meshwork Cells (TMCs), an endothelial-derived histotype with phagocytic and secretive functions which lay at the iridocorneal angle in the anterior segment of the eye. Since TMCs physiologically regulate the composition and architecture of trabecular meshwork (TM), which is the main outflow pathway of aqueous humor, a fluid which shapes the eye globe and nourishes the lining cell types, GCs are supposed to trigger a pathological remodeling of the TM, inducing an IOP increase and retina mechanical compression. The metabolic dysregulation of TMCs induced by GCs exposure has never been characterized at the molecular detail. Herein, we report that, upon dexamethasone exposure, a TMCs strain develops a marked inhibition of the autophagosome biogenesis pathway through an enhanced turnover of two members of the Ulk-1 complex, the main platform for autophagy induction, through the Ubiquitin Proteasome System (UPS).

The administration of a ketogenic diet (KD) has been considered therapeutic in subjects with irritable bowel syndrome (IBS). This study aimed to investigate the molecular mechanisms by which a low-carbohydrate diet, such as KD, can improve gastrointestinal symptoms and functions in an animal model of IBS by evaluating possible changes in intestinal tissue expression of endocannabinoid receptors. In rats fed a KD, we detected a significant restoration of cell damage to the intestinal crypt base, a histological feature of IBS condition, and upregulation of CB1 and CB2 receptors. The diet also affected glucose metabolism and intestinal membrane permeability, with an overexpression of the glucose transporter GLUT1 and tight junction proteins in treated rats. The present data suggest that CB receptors represent one of the molecular pathways through which the KD works and support possible cannabinoid-mediated protection at the intestinal level in the IBS rats after dietary treatment.

Structural and functional properties of ferrous Mycobacterium tuberculosis (Mt-Nb) and human (Hs-Nb) nitrobindins (Nbs) were investigated. At pH 7.0 and 25.0 °C, the unliganded Fe(II) species is penta-coordinated and unlike most other hemoproteins no pH-dependence of its coordination was detected over the pH range between 2.2 and 7.0. Further, despite a very open distal side of the heme pocket (as also indicated by the vanishingly small geminate recombination of CO for both Nbs), which exposes the heme pocket to the bulk solvent, their reactivity toward ligands, such as CO and NO, is significantly slower than in most hemoproteins, envisaging either a proximal barrier for ligand binding and/or crowding of H2O molecules in the distal side of the heme pocket which impairs ligand binding to the heme Fe-atom. On the other hand, liganded species display already at pH 7.0 and 25 °C a severe weakening (in the case of CO) and a cleavage (in the case of NO) of the proximal Fe-His bond, suggesting that the ligand-linked movement of the Fe(II) atom onto the heme plane brings about a marked lengthening of the proximal Fe-imidazole bond, eventually leading to its rupture. This structural evidence is accompanied by a marked enhancement of both ligands dissociation rate constants. As a whole, these data clearly indicate that structural–functional relationships in Nbs strongly differ from what observed in mammalian and truncated hemoproteins, suggesting that Nbs play a functional role clearly distinct from other eukaryotic and prokaryotic hemoproteins.

Basal-like breast cancer (BLBC) is an aggressive and deadly subtype of human breast cancer that is highly metastatic, displays stem-cell like features, and has limited treatment options. Therefore, developing and characterizing preclinical mouse models with tumors that resemble BLBC is important for human therapeutic development. ATF3 is a potent oncogene that is aberrantly expressed in most human breast cancers. In the BK5.ATF3 mouse model, overexpression of ATF3 in the basal epithelial cells of the mammary gland produces tumors that are characterized by activation of the Wnt/β-catenin signaling pathway. Here, we used RNA-Seq and microRNA (miRNA) microarrays to better define the molecular features of BK5.ATF3-derived mammary tumors. These analyses showed that these tumors share many characteristics of human BLBC including reduced expression of Rb1, Esr1, and Pgr and increased expression of Erbb2, Egfr, and the genes encoding keratins 5, 6, and 17. An analysis of miRNA expression revealed reduced levels of Mir145 and Mir143, leading to the upregulation of their target genes including both the pluripotency factors Klf4 and Sox2 as well as the cancer stem-cell-related gene Kras. Finally, we show through knock-down experiments that ATF3 may directly modulate MIR145/143 expression. Taken together, our results indicate that the ATF3 mouse mammary tumor model could provide a powerful model to define the molecular mechanisms leading to BLBC, identify the factors that contribute to its aggressiveness, and, ultimately, discover specific genes and gene networks for therapeutic targeting.

Short bowel syndrome is a pathological condition resulting from extensive resection of the intestine, generally performed due to congenital abnormalities, Crohn’s disease, mesenteric ischemia, or neoplasms. The main consequence of this syndrome is a reduction of intestinal absorption, which causes malnutrition and dehydration. In the most severe cases, specific and complex surgical procedures are requested to manage the syndrome. Such procedures consist of the intestinal lengthening, with lead to an increase of absorptive mucosal surface and intestinal transit time and an overall enhancement of intestinal absorption. One of the most promising surgical procedures is spiral intestinal lengthening and tailoring, which consists of a spiral incision of the intestinal wall and in the elongation longitudinally of the intestine by sliding one flap over the other. The final intestinal lengthening is strictly dependent on a series of parameters, some of which are defined by the surgeon. The present paper proposes a mathematical model, based on patient specific anatomical data, which aims to help the surgeon in defining the optimal parameters for the intervention and in foreseeing its outcomes from the preoperative planning phase. Such a tool can assist the physician in the surgery room by improving the procedure and reducing surgical times.

Ferric nitrobindins (Nbs) selectively bind NO and catalyze the conversion of peroxynitrite to nitrate. In this study, we show that NO scavenging occurs through the reductive nitrosylation of ferric Mycobacterium tuberculosis and Homo sapiens nitrobindins (Mt-Nb(III) and Hs-Nb(III), respectively). The conversion of Mt-Nb(III) and Hs-Nb(III) to Mt-Nb(II)-NO and Hs-Nb(II)-NO, respectively, is a monophasic process, suggesting that over the explored NO concentration range (between 2.5 × 10−5 and 1.0 × 10−3 M), NO binding is lost in the mixing time (i.e., NOkon ≥ 1.0 × 106 M−1 s−1). The pseudo-first-order rate constant for the reductive nitrosylation of Mt-Nb(III) and Hs-Nb(III) (i.e., k) is not linearly dependent on the NO concentration but tends to level off, with a rate-limiting step (i.e., klim) whose values increase linearly with [OH−]. This indicates that the conversion of Mt-Nb(III) and Hs-Nb(III) to Mt-Nb(II)-NO and Hs-Nb(II)-NO, respectively, is limited by the OH−-based catalysis. From the dependence of klim on [OH−], the values of the second-order rate constant kOH− for the reductive nitrosylation of Mt-Nb(III)-NO and Hs-Nb(III)-NO were obtained (4.9 (±0.5) × 103 M−1 s−1 and 6.9 (±0.8) × 103 M−1 s−1, respectively). This process leads to the inactivation of two NO molecules: one being converted to HNO2 and another being tightly bound to the ferrous heme-Fe(II) atom.

Key Points High glucose quickly induces an atypical NF-kB pro-inflammatory program. CamKII phosphorylation of Rpt6 subunit of the proteasome stimulates IkBα turnover and p65-p50 release. Inhibition of either CamkII or proteasome blocks this pathway.

Yang et al. examine whether genetic ancestry is associated with genetic variation in fatty acid desaturases and plasma phospholipid levels of long chain polyunsaturated fatty acids (LC-PUFAs) in Hispanic Americans. They find strong associations between Amerind genetic ancestry and LC-PUFA levels; and report that the well-known FADS rs174537 variant was associated with several metabolic, inflammatory and anthropomorphic traits including circulating triglycerides.

Background: Oral cavity cancer is still an important public health problem throughout the world. Oral squamous cell carcinomas (OSCCs) can be quite aggressive and metastatic, with a low survival rate and poor prognosis. However, this is usually related to the clinical stage and histological grade, and molecular prognostic markers for clinical practice are yet to be defined. Heparanase (HPSE1) is an endoglycosidase associated with extracellular matrix remodeling, and although involved in several malignancies, the clinical implications of HPSE1 expression in OSCCs are still unknown.Methods: We sought to investigate HPSE1 expression in a series of primary OSCCs and further explore whether its overexpression plays a relevant role in OSCC tumorigenesis. mRNA and protein expression analyses were performed in OSCC tissue samples and cell lines. A loss-of-function strategy using shRNA and a gain-of-function strategy using an ORF vector targeting HPSE1 were employed to investigate the endogenous modulation of HPSE1 and its effects on proliferation, apoptosis, adhesion, epithelial–mesenchymal transition (EMT), angiogenesis, migration, and invasion of oral cancer in vitro.Results: We demonstrated that HPSE1 is frequently upregulated in OSCC samples and cell lines and is an unfavorable prognostic indicator of disease-specific survival when combined with advanced pT stages. Moreover, abrogation of HPSE1 in OSCC cells significantly promoted apoptosis and inhibited proliferation, migration, invasion, and epithelial–mesenchymal transition by significantly decreasing the expression of N-cadherin and vimentin. Furthermore, a conditioned medium of HPSE1-downregulated cells resulted in reduced vascular endothelial growth.Conclusion: Our results confirm the overexpression of HPSE1 in OSCCs, suggest that HPSE1 expression correlates with disease progression as it is associated with several important biological processes for oral tumorigenesis, and can be managed as a prognostic marker for patients with OSCC.

Many age-associated changes in the human hematopoietic system have been reproduced in murine models; however, such changes have not been as robustly explored in rats despite the fact these larger rodents are more physiologically similar to humans. We examined peripheral blood of male F344 rats ranging from 3 to 27 months of age and found significant age-associated changes with distinct leukocyte population shifts. We report CD25+ CD4+ population frequency is a strong predictor of healthy aging, generate a model using blood parameters, and find rats with blood profiles that diverge from chronologic age indicate debility; thus, assessments of blood composition may be useful for non-lethal disease profiling or as a surrogate measure for efficacy of aging interventions. Importantly, blood parameters and DNA methylation alterations, defined distinct juncture points during aging, supporting a non-linear aging process. Our results suggest these inflection points are important considerations for aging interventions. Overall, we present rat blood aging metrics that can serve as a resource to evaluate health and the effects of interventions in a model system physiologically more reflective of humans.

The Helicobacter pylori Neutrophil Activating Protein (HP-NAP) is endowed with immunomodulatory properties that make it a potential candidate for anticancer therapeutic applications. By activating cytotoxic Th1 responses, HP-NAP inhibits the growth of bladder cancer and enhances the anti-tumor activity of oncolytic viruses in the treatment of metastatic breast cancer and neuroendocrine tumors. The possibility that HP-NAP exerts its anti-tumor effect also by modulating the activity of innate immune cells has not yet been explored. Taking advantage of the zebrafish model, we examined the therapeutic efficacy of HP-NAP against metastatic human melanoma, limiting the observational window to 9 days post-fertilization, well before the maturation of the adaptive immunity. Human melanoma cells were xenotransplanted into zebrafish embryos and tracked in the presence or absence of HP-NAP. The behavior and phenotype of macrophages and the impact of their drug-induced depletion were analyzed exploiting macrophage-expressed transgenes. HP-NAP administration efficiently inhibited tumor growth and metastasis and this was accompanied by strong recruitment of macrophages with a pro-inflammatory profile at the tumor site. The depletion of macrophages almost completely abrogated the ability of HP-NAP to counteract tumor growth. Our findings highlight the pivotal role of activated macrophages in counteracting melanoma growth and support the notion that HP-NAP might become a new biological therapeutic agent for the treatment of metastatic melanomas.

Summary: Learning critically depends on the ability to rapidly form and store non-overlapping representations of the external world. In line with their postulated role in episodic memory, hippocampal place cells can undergo a rapid reorganization of their firing fields upon contextual manipulations. To explore the mechanisms underlying such global remapping, we juxtacellularly stimulated 42 hippocampal neurons in freely moving mice during spatial exploration. We found that evoking spike trains in silent neurons was sufficient for creating place fields, while in place cells, juxtacellular stimulation induced a rapid remapping of their place fields to the stimulus location. The occurrence of complex spikes was most predictive of place field plasticity. Our data thus indicate that plasticity-inducing stimuli are able to rapidly bias place cell activity, simultaneously suppressing existing place fields. We propose that such competitive place field dynamics could support the orthogonalization of the hippocampal map during global remapping. : Place cells can serve as a readout of hippocampal memory. Diamantaki et al. show that the activity of single place cells can be rapidly modified by single-cell stimulation in freely moving mice. This finding provides insights into the cellular mechanisms that support the rapid reorganization of hippocampal place maps.

(1) Background: Activation of the PI3K-AKT pathway controls most hallmarks of cancer, and the hedgehog (HH) pathway has been associated with oral squamous cell carcinoma (OSCC) development and progression. We hypothesized that fibroblast-derived insulin-like growth factor-1 (IGF-1) acts in oral squamous cell carcinoma (OSCC) cells, leading to the non-canonical activation of the HH pathway, maintaining AKT activity and promoting tumor aggressiveness. (2) Methods: Primary fibroblasts (MF1) were genetically engineered for IGF-1 overexpression (MF1-IGF1) and CRISPR/Cas9-mediated IGF1R silencing was performed in SCC-4 cells. SCC-4 cells were co-cultured with fibroblasts or incubated with fibroblast conditioned medium (CM) or rIGF-1 for functional assays and the evaluation of AKT and HH pathways. (3) Results: Gene expression analysis confirmed IGF-1 overexpression in MF1-IGF1 and the absence of IGF-1 expression in SCC-4, while elevated IGF1R expression was detected. IGF1R silencing was associated with decreased survival of SCC-4 cells. Ihh was expressed in both MF1 and MF1-IGF1, and increased levels of GLI1 mRNA were observed in SCC-4 after stimulation with CM-MF1. Activation of both PI3K-AKT and the HH pathway (GLI1, Ihh and SMO) were identified in SCC-4 cells cultured in the presence of MF1-IGF1-CM. rIGF-1 promoted tumor cell proliferation, migration, invasion and tumorsphere formation, whereas CM-MF1 significantly stimulated angiogenesis. (4) Conclusions: IGF-1 exerts pro-tumorigenic effects by stimulating SCC-4 cell proliferation, migration, invasion and stemness. AKT and HH pathways were activated by IGF-1 in SCC-4, reinforcing its influence on the regulation of these signaling pathways.

The present study provides new evidence that cationic porphyrins may be considered as tunable platforms to interfere with the structural “key code” present on the 20S proteasome α-rings and, by consequence, with its catalytic activity. Here, we describe the functional and conformational effects on the 20S proteasome induced by the cooperative binding of the tri-cationic 5-(phenyl)-10,15,20-(tri N-methyl-4-pyridyl) porphyrin (Tris-T4). Our integrated kinetic, NMR, and in silico analysis allowed us to disclose a complex effect on the 20S catalytic activity depending on substrate/porphyrin concentration. The analysis of the kinetic data shows that Tris-T4 shifts the relative populations of the multiple interconverting 20S proteasome conformations leading to an increase in substrate hydrolysis by an allosteric pathway. Based on our Tris-T4/h20S interaction model, Tris-T4 is able to affect gating dynamics and substrate hydrolysis by binding to an array of negatively charged and hydrophobic residues present on the protein surface involved in the 20S molecular activation by the regulatory proteins (RPs). Accordingly, despite the fact that Tris-T4 also binds to the α3ΔN mutant, allosteric modulation is not observed since the molecular mechanism connecting gate dynamics with substrate hydrolysis is impaired. We envisage that the dynamic view of the 20S conformational equilibria, activated through cooperative Tris-T4 binding, may work as a simplified model for a better understanding of the intricate network of 20S conformational/functional states that may be mobilized by exogenous ligands, paving the way for the development of a new generation of proteasome allosteric modulators.

Due to its importance in the pathogenesis of oral squamous cell carcinoma (OSCC), the Hedgehog (HH) pathway is considered a potential therapeutic target. We investigated the effects of GANT61, a GLI inhibitor, on HH gene expression, as well as on metastatic OSCC cell proliferation and death. Following culture in DMEM medium, cytotoxicity of GANT61 against different tumor and non-tumor cell types was assessed by alamarBlue assays. Cytotoxicity analysis revealed that the metastatic HSC3 cell line was the most sensitive (IC50: 36 µM) to the tested compound. The compound’s effects on the expression of HH pathways components were analyzed by qPCR and Western blot; cell viability was analyzed by trypan blue assay and flow cytometry were used to investigate cell cycle phase, morphology, and death patterns in HSC3 cells. A significant reduction in mRNA levels of the GLI1 transcription factor was found after 12 h of treatment withGANT61. Protein expression levels of other HH pathway components (PTCH1, SHH, and Gli1) and HSC3 cell viability also decreased after 24 h of treatment. Cell cycle analysis and death pattern evaluations revealed significantly increased nuclear fragmentation in sub-G1 phase, as well as cell death due to apoptosis. In conclusion, the significantly reduced GLI1 gene expression seen in response to the GLI inhibitor indicates diminished downstream activation in HH pathway components. GANT61 significantly reduced cell viability in the metastatic cell line of OSCC and promoted a significant increase in nuclear fragmentation and cell death by apoptosis.

The hydraulic and hydrogeological features of the Caposele aquifer have been investigated by using a numerical groundwater flow model. In particular, groundwater flow simulations were performed for a multilayered, unconfined aquifer in steady-state conditions for different thicknesses of the aquifer’s saturated zone. The Caposele groundwater model was carried out starting from a generic model drained by a unique spring outlet in accordance with the geo-hydrological features of the study area. The conceptual model was built considering hydrogeological features of spring catchment, and was then implemented with the MODFLOW numerical code. A combined 2D-3D approach was adopted, and the model was calibrated on borehole data available for the time period 2012–2019. Different thicknesses of the aquifer were set, and a reliable relationship was found between the hydraulic head, saturated zone and hydraulic conductivity of the aquifer. Using the MODPATH package, the mean travel time (Darcian) of groundwater was computed for five different scenarios, corresponding to the model’s depths; the analysis that was performed shows that the travel time is higher for a greater and lower for a smaller thickness of the aquifer’s saturated zone, respectively. The Caposele aquifer model was zoned in different sectors, named flow pipe areas, that play different roles in groundwater recharge-discharge processes. A vector analysis was also carried out in order to highlight the ascendant flow near the spring zone.

(1) Background: Colorectal cancer (CRC) is among the best examples of the relationship between inflammation and increased cancer risk. (2) Methods: To examine the effects of spontaneous low-grade chronic inflammation on the pathogenesis of CRC, we developed a new murine model of colitis-associated cancer (CAC) by crossing Mucin 2 mutated mice (Winnie) with ApcMin/+ mice. (3) Results: The resulting Winnie-ApcMin/+ model combines an inflammatory background with a genetic predisposition to small intestinal polyposis. Winnie-ApcMin/+ mice show an early occurrence of inflammatory signs and dysplastic lesions in the distal colon with a specific molecular signature. (4) Conclusion: The Winnie-ApcMin/+ model is a perfect model to demonstrate that chronic inflammation represents a crucial risk factor for the onset and progression of tumoral lesions in individuals genetically predisposed to CRC.

This article presents a case study about the impact that our mobile laboratory, the Rutgers Science Explorer bus, has had on the professional development of graduate students and content enrichment for the middle school communities in the state of New Jersey.

Molecular recognition reagents are key tools for understanding biological processes and are used universally by scientists to study protein expression, localisation and interactions. Antibodies remain the most widely used of such reagents and many show excellent performance, although some are poorly characterised or have stability or batch variability issues, supporting the use of alternative binding proteins as complementary reagents for many applications. Here we report on the use of Affimer proteins as research reagents. We selected 12 diverse molecular targets for Affimer selection to exemplify their use in common molecular and cellular applications including the (a) selection against various target molecules; (b) modulation of protein function in vitro and in vivo; (c) labelling of tumour antigens in mouse models; and (d) use in affinity fluorescence and super-resolution microscopy. This work shows that Affimer proteins, as is the case for other alternative binding scaffolds, represent complementary affinity reagents to antibodies for various molecular and cell biology applications.

Fibrin sealants derived from human blood can be used in tissue engineering to assist in the repair of bone defects. The objective of this study was to evaluate the support system formed by a xenograft fibrin sealant associated with photobiomodulation therapy of critical defects in rat calvaria. Thirty-six rats were divided into four groups: BC (n = 8), defect filled with blood clot; FSB (n = 10), filled with fibrin sealant and xenograft; BCPBMT (n = 8), blood clot and photobiomodulation; FSBPBMT (n = 10), fibrin sealant, xenograft, and photobiomodulation. The animals were killed after 14 and 42 days. In the histological and microtomographic analysis, new bone formation was observed in all groups, limited to the defect margins, and without complete wound closure. In the FSB group, bone formation increased between periods (4.3 ± 0.46 to 6.01 ± 0.32), yet with lower volume density when compared to the FSBPBMT (5.6 ± 0.45 to 10.64 ± 0.97) group. It was concluded that the support system formed by the xenograft fibrin sealant associated with the photobiomodulation therapy protocol had a positive effect on the bone repair process.

Neurons coding for head-direction are crucial for spatial navigation. Here we explored the cellular basis of head-direction coding in the rat dorsal presubiculum (PreS). We found that layer2 is composed of two principal cell populations (calbindin-positive and calbindin-negative neurons) which targeted the contralateral PreS and retrosplenial cortex, respectively. Layer3 pyramidal neurons projected to the medial entorhinal cortex (MEC). By juxtacellularly recording PreS neurons in awake rats during passive-rotation, we found that head-direction responses were preferentially contributed by layer3 pyramidal cells, whose long-range axons branched within layer3 of the MEC. In contrast, layer2 neurons displayed distinct spike-shapes, were not modulated by head-direction but rhythmically-entrained by theta-oscillations. Fast-spiking interneurons showed only weak directionality and theta-rhythmicity, but were significantly modulated by angular velocity. Our data thus indicate that PreS neurons differentially contribute to head-direction coding, and point to a cell-type- and layer-specific routing of directional and non-directional information to downstream cortical targets.

Gold nanoparticles-enabled intracellular surface-enhanced Raman spectroscopy (SERS) provides a sensitive and promising technique for single cell analysis. Compared with spherical gold nanoparticles, gold nanoflowers, i.e., flower-shaped gold nanostructures, can produce a stronger SERS signal. Current exploration of gold nanoflowers for intracellular SERS has been considerably limited by the difficulties in preparation, as well as background signal and cytotoxicity arising from the surfactant capping layer. Recently, we have developed a facile and surfactant-free method for fabricating hollow-channel gold nanoflowers (HAuNFs) with great single-particle SERS activity. In this paper, we investigate the cellular uptake and cytotoxicity of our HAuNFs using a RAW 264.7 macrophage cell line, and have observed effective cellular internalization and low cytotoxicity. We have further engineered our HAuNFs into SERS-active tags, and demonstrated the functionality of the obtained tags as trimodal nanoprobes for dark-field and fluorescence microscopy imaging, together with intracellular SERS.

We evaluated the genetic changes in bladder cancer biopsy by fluorescence in situ hybridization (FISH) and related them to stage and grade of the tumor, ploidy (FCM) and clinical outcome, to determine a simple method to identify tumors with a poorer prognosis. Using FISH the numerical aberrations of chromosomes 1,7,9,17 in tumor’s imprints of 70 patients with transitional cell cancer (TCC) were determined. First of all, the data demonstrated that the sensitivity of FISH in detecting quantitative DNA aberrations exceeds FCM’s sensitivity. The frequency of chromosome 1 and 9 aberrations did not show significant differences in diploid and aneuploid tumors in different stage and grade. On the contrary, the chromosome 7 and 17 aneusomy showed greater differences between pT1 and pT2-3 tumors (p

Abstract Background Melanoacanthoma (MA) has been described in the oral mucosa as a solitary lesion or, occasionally, as multiple lesions. MA mainly affects dark skinned patients and grows rapidly, showing a plane or slightly raised appearance and a brown to black color. The differential diagnosis includes oral nevi, amalgam tattoos, and melanomas. We report here the case of a 58-year-old black woman who presented multiple pigmented lesions on the hard palate. Case presentation Based on the differential diagnosis of melanoma, a punch biopsy (4 mm in diameter) was performed. The material was fixed in 10% formalin, embedded in paraffin, and stained with hematoxylin-eosin or submitted to immunohistochemical analysis. Immunohistochemistry using antibodies against protein S-100, melan-A, HMB-45, MCM-2, MCM-5, Ki-67 and geminin was performed. Immunohistochemical analysis revealed strong cytoplasmic immunoreactivity of dendritic melanocytes for proteinS-100, HMB-45 and melan-A.Positive staining for proliferative markers (MCM-2, MCM-5, Ki-67) was only observed in basal and suprabasal epithelial cells, confirming the reactive etiology of the lesion. The diagnosis was oral Melanoacanthoma (MA). Conclusion The patient has been followed up for 30 months and shows no clinical alterations. MA should be included in the differential diagnosis of pigmented lesions of the oral cavity.

Abstract Background With an abundant amount of microarray gene expression data sets available through public repositories, new possibilities lie in combining multiple existing data sets. In this new context, analysis itself is no longer the problem, but retrieving and consistently integrating all this data before delivering it to the wide variety of existing analysis tools becomes the new bottleneck. Results We present the newly released inSilicoMerging R/Bioconductor package which, together with the earlier released inSilicoDb R/Bioconductor package, allows consistent retrieval, integration and analysis of publicly available microarray gene expression data sets. Inside the inSilicoMerging package a set of five visual and six quantitative validation measures are available as well. Conclusions By providing (i) access to uniformly curated and preprocessed data, (ii) a collection of techniques to remove the batch effects between data sets from different sources, and (iii) several validation tools enabling the inspection of the integration process, these packages enable researchers to fully explore the potential of combining gene expression data for downstream analysis. The power of using both packages is demonstrated by programmatically retrieving and integrating gene expression studies from the InSilico DB repository [https://insilicodb.org/app/].

Abstract Background Huanglongbing (HLB, previously known as citrus greening), is associated with Candidatus Liberibacter species and is a serious threat to citrus production world-wide. The pathogen is a Gram negative, unculturable, phloem-limited bacterium with limited known genomic information. Expanding the genetic knowledge of this organism may provide better understanding of the pathogen and possibly develop effective strategies for control and management of HLB. Results Here, we report cloning and characterization of an additional 14.7 Kb of new genomic sequences from three different genomic regions of the Candidatus Liberibacter asiaticus (Las). Sequence variation analyses among the available Ca. Liberibacter species sequences as well as the newly cloned 1.5 Kb of rpoB gene from different Ca. Liberibacter strains have identified INDELs and SNPs. Phylogenetic analysis of the deduced protein sequences from the cloned regions characterizes the HLB-associated Candidatus Liberibacter as a new clade in the sub-division of the α-proteobacteria. Conclusion Comparative analyses of the cloned gene regions of Candidatus Liberibacter with members of the order Rhizobiales suggest overall gene structure and order conservation, albeit with minor variations including gene decay due to the identified pseudogenes. The newly cloned gene regions contribute to our understanding of the molecular aspects of genomic evolution of Ca. Liberibacter.

The main purpose of this study is to perform a nitrogen budget survey for the entire Brazilian Amazon region. The main inputs of nitrogen to the region are biological nitrogen fixation occurring in tropical forests (7.7 Tg.yr-1), and biological nitrogen fixation in agricultural lands mainly due to the cultivation of a large area with soybean, which is an important nitrogen-fixing crop (1.68 Tg.yr-1). The input due to the use of N fertilizers (0.48 Tg.yr-1) is still incipient compared to the other two inputs mentioned above. The major output flux is the riverine flux, equal to 2.80 Tg.yr-1 and export related to foodstuff, mainly the transport of soybean and beef to other parts of the country. The continuous population growth and high rate of urbanization may pose new threats to the nitrogen cycle of the region through the burning of fossil fuel and dumping of raw domestic sewage in rivers and streams of the region.

Riverine nitrogen distribution is increasingly controlled by anthropogenic activities in their watersheds, regardless of spatial scale, climate, and geographical zone. Consequently, modelling efforts to predict the export of nitrogen from rivers worldwide have used attributes such as population density, land use, urbanization and sanitation. These models have greatly enhanced our understanding of the sources and fate of nitrogen added to terrestrial systems and transported to rivers and streams, especially for developed countries of the North temperate zone. However, much of the world's population lives in developing countries of the tropics, where the effects of human activities on riverine N exports are still poorly understood. In an effort to close this gap, we compare riverine nitrogen data from 32 Brazilian rivers draining two contrasting regions in this tropical country in terms of economic development - the State of São Paulo and the Amazon. Our data include nitrogen in different dissolved forms, such as Dissolved Inorganic Nitrogen (DIN) and Dissolved Organic Nitrogen (DON). The results show that nitrogen concentrations decreased as river runoff increased in both study areas, and that concentrations were significantly higher in rivers draining the most economically developed region. The relationships between nitrogen concentrations and fluxes with demographic parameters such as population density were also determined and compared to those in temperate systems. In contrast to temperate watersheds, we found that nitrogen fluxes increased only after population densities were higher than 10 individuals per km².