CD5L as a promising biological therapeutic for treating sepsis.

Sepsis results from systemic, dysregulated inflammatory responses to infection, culminating in multiple organ failure. Here, we demonstrate the utility of CD5L for treating experimental sepsis caused by cecal ligation and puncture (CLP). We show that CD5L's important features include its ability to enhance neutrophil recruitment and activation by increasing circulating levels of CXCL1, and to promote neutrophil phagocytosis. CD5L-deficient mice exhibit impaired neutrophil recruitment and compromised bacterial control, rendering them susceptible to attenuated CLP. CD5L-/- peritoneal cells from mice subjected to medium-grade CLP exhibit a heightened pro-inflammatory transcriptional profile, reflecting a loss of control of the immune response to the infection. Intravenous administration of recombinant CD5L (rCD5L) in immunocompetent C57BL/6 wild-type (WT) mice significantly ameliorates measures of disease in the setting of high-grade CLP-induced sepsis. Furthermore, rCD5L lowers endotoxin and damage-associated molecular pattern (DAMP) levels, and protects WT mice from LPS-induced endotoxic shock. These findings warrant the investigation of rCD5L as a possible treatment for sepsis in humans.

Exosomes define a local and systemic communication network in healthy pancreas and pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC), a lethal disease, requires a grasp of its biology for effective therapies. Exosomes, implicated in cancer, are poorly understood in living systems. Here we use the genetically engineered mouse model (ExoBow) to map the spatiotemporal distribution of exosomes from healthy and PDAC pancreas in vivo to determine their biological significance. We show that, within the PDAC microenvironment, cancer cells establish preferential communication routes through exosomes with cancer associated fibroblasts and endothelial cells. The latter being a conserved event in the healthy pancreas. Inhibiting exosomes secretion in both scenarios enhances angiogenesis, underscoring their contribution to vascularization and to cancer. Inter-organ communication is significantly increased in PDAC with specific organs as most frequent targets of exosomes communication occurring in health with the thymus, bone-marrow, brain, and intestines, and in PDAC with the kidneys, lungs and thymus. In sum, we find that exosomes mediate an organized intra- and inter- pancreas communication network with modulatory effects in vivo.

Transcriptomic remodeling of gastric cells by Helicobacter pylori outer membrane vesicles.

BACKGROUND: Outer membrane vesicles (OMVs) are spontaneously released by Gram-negative bacteria and influence bacteria-host interactions by acting as a delivery system for bacterial components and by interacting directly with host cells. Helicobacter pylori, a pathogenic bacterium that chronically colonizes the human stomach, also sheds OMVs, and their impact on bacterial-mediated diseases is still being elucidated. MATERIALS AND METHODS: Transcriptomic profiling of the human gastric cell line MKN74 upon challenge with H. pylori OMVs compared to control and infected cells was performed using the Ion AmpliSeq™ Transcriptome Human Gene Expression Panel to understand the gene expression changes that human gastric epithelial cells might undergo when exposed to H. pylori OMVs. RESULTS: H. pylori OMVs per se modify the gene expression profile of gastric epithelial cells, adding another layer of (gene) regulation to the already complex host-bacteria interaction. The most enriched pathways include those related to amino acid metabolism, mitogen-activated protein kinase signaling, autophagy, and ferroptosis, whereas the cell cycle, DNA replication, and DNA repair were the most downregulated. The transcriptomic changes induced by OMVs were mostly similar to those induced by the parental bacteria, likely amplifying the effects of the bacterium itself. CONCLUSIONS: Our data provide a valuable portrayal of the transcriptomic remodeling of gastric cells induced by H. pylori OMVs. It demonstrates the breadth of cellular pathways and genes affected by OMVs, most previously unreported, which can be further dissected for the underlying molecular mediators and explored to understand the pathobiology of the full spectrum of H. pylori-mediated diseases.

Syndecan-4 is a maestro of gastric cancer cell invasion and communication that underscores poor survival.

Gastric cancer is a dominating cause of cancer-associated mortality with limited therapeutic options. Here, we show that syndecan-4 (SDC4), a transmembrane proteoglycan, is highly expressed in intestinal subtype gastric tumors and that this signature associates with patient poor survival. Further, we mechanistically demonstrate that SDC4 is a master regulator of gastric cancer cell motility and invasion. We also find that SDC4 decorated with heparan sulfate is efficiently sorted in extracellular vesicles (EVs). Interestingly, SDC4 in EVs regulates gastric cancer cell-derived EV organ distribution, uptake, and functional effects in recipient cells. Specifically, we show that SDC4 knockout disrupts the tropism of EVs for the common gastric cancer metastatic sites. Our findings set the basis for the molecular implications of SDC4 expression in gastric cancer cells and provide broader perspectives on the development of therapeutic strategies targeting the glycan-EV axis to limit tumor progression.

Fine-scale subpopulation detection via an SNP-based unsupervised method: A case study on the 1000 Genomes Project resources.

SNP-based information is used in several existing clustering methods to detect shared genetic ancestry or to identify population substructure. Here, we present a methodology, called IPCAPS for unsupervised population analysis using iterative pruning. Our method, which can capture fine-level structure in populations, supports ordinal data, and thus can readily be applied to SNP data. Although haplotypes may be more informative than SNPs, especially in fine-level substructure detection contexts, the haplotype inference process often remains too computationally intensive. In this work, we investigate the scale of the structure we can detect in populations without knowledge about haplotypes; our simulated data do not assume the availability of haplotype information while comparing our method to existing tools for detecting fine-level population substructures. We demonstrate experimentally that IPCAPS can achieve high accuracy and can outperform existing tools in several simulated scenarios. The fine-level structure detected by IPCAPS on an application to the 1000 Genomes Project data underlines its subject heterogeneity.

First insight into oral microbiome diversity in Papua New Guineans reveals a specific regional signature.

The oral microbiota is a highly complex and diversified part of the human microbiome. Being located at the interface between the human body and the exterior environment, this microbiota can deepen our understanding of the environmental impacts on the global status of human health. This research topic has been well addressed in Westernized populations, but these populations only represent a fraction of human diversity. Papua New Guinea hosts very diverse environments and one of the most unique human biological diversities worldwide. In this study we performed the first known characterization of the oral microbiome in 85 Papua New Guinean individuals living in different environments, using a qualitative and quantitative approach. We found a significant geographical structure of the Papua New Guineans oral microbiome, especially in the groups most isolated from urban spaces. In comparison to other global populations, two bacterial genera related to iron absorption were significantly more abundant in Papua New Guineans and Aboriginal Australians, which suggests a shared oral microbiome signature. Further studies will be needed to confirm and explore this possible regional-specific oral microbiome profile.

Extracellular Vesicles from Pancreatic Cancer Stem Cells Lead an Intratumor Communication Network (EVNet) to fuel tumour progression.

OBJECTIVE: Intratumor heterogeneity drives cancer progression and therapy resistance. However, it has yet to be determined whether and how subpopulations of cancer cells interact and how this interaction affects the tumour. DESIGN: We have studied the spontaneous flow of extracellular vesicles (EVs) between subpopulations of cancer cells: cancer stem cells (CSC) and non-stem cancer cells (NSCC). To determine the biological significance of the most frequent communication route, we used pancreatic ductal adenocarcinoma (PDAC) orthotopic models, patient-derived xenografts (PDXs) and genetically engineered mouse models (GEMMs). RESULTS: We demonstrate that PDAC tumours establish an organised communication network between subpopulations of cancer cells using EVs called the EVNet). The EVNet is plastic and reshapes in response to its environment. Communication within the EVNet occurs preferentially from CSC to NSCC. Inhibition of this communication route by impairing Rab27a function in orthotopic xenographs, GEMMs and PDXs is sufficient to hamper tumour growth and phenocopies the inhibition of communication in the whole tumour. Mechanistically, we provide evidence that CSC EVs use agrin protein to promote Yes1 associated transcriptional regulator (YAP) activation via LDL receptor related protein 4 (LRP-4). Ex vivo treatment of PDXs with antiagrin significantly impairs proliferation and decreases the levels of activated YAP.Patients with high levels of agrin and low inactive YAP show worse disease-free survival. In addition, patients with a higher number of circulating agrin+ EVs show a significant increased risk of disease progression. CONCLUSION: PDAC tumours establish a cooperation network mediated by EVs that is led by CSC and agrin, which allows tumours to adapt and thrive. Targeting agrin could make targeted therapy possible for patients with PDAC and has a significant impact on CSC that feeds the tumour and is at the centre of therapy resistance.

Projecting Ancient Ancestry in Modern-Day Arabians and Iranians: A Key Role of the Past Exposed Arabo-Persian Gulf on Human Migrations.

The Arabian Peninsula is strategic for investigations centered on the early structuring of modern humans in the wake of the out-of-Africa migration. Despite its poor climatic conditions for the recovery of ancient human DNA evidence, the availability of both genomic data from neighboring ancient specimens and informative statistical tools allow modeling the ancestry of local modern populations. We applied this approach to a data set of 741,000 variants screened in 291 Arabians and 78 Iranians, and obtained insightful evidence. The west-east axis was a strong forcer of population structure in the Peninsula, and, more importantly, there were clear continuums throughout time linking western Arabia with the Levant, and eastern Arabia with Iran and the Caucasus. Eastern Arabians also displayed the highest levels of the basal Eurasian lineage of all tested modern-day populations, a signal that was maintained even after correcting for a possible bias due to a recent sub-Saharan African input in their genomes. Not surprisingly, eastern Arabians were also the ones with highest similarity with Iberomaurusians, who were, so far, the best proxy for the basal Eurasians amongst the known ancient specimens. The basal Eurasian lineage is the signature of ancient non-Africans who diverged from the common European-eastern Asian pool before 50,000 years ago, prior to the later interbred with Neanderthals. Our results appear to indicate that the exposed basin of the Arabo-Persian Gulf was the possible home of basal Eurasians, a scenario to be further investigated by searching ancient Arabian human specimens.

Multi-Tissue Transcriptomic-Informed In Silico Investigation of Drugs for the Treatment of Dengue Fever Disease.

Transcriptomics, proteomics and pathogen-host interactomics data are being explored for the in silico-informed selection of drugs, prior to their functional evaluation. The effectiveness of this kind of strategy has been put to the test in the current COVID-19 pandemic, and it has been paying off, leading to a few drugs being rapidly repurposed as treatment against SARS-CoV-2 infection. Several neglected tropical diseases, for which treatment remains unavailable, would benefit from informed in silico investigations of drugs, as performed in this work for Dengue fever disease. We analyzed transcriptomic data in the key tissues of liver, spleen and blood profiles and verified that despite transcriptomic differences due to tissue specialization, the common mechanisms of action, "Adrenergic receptor antagonist", "ATPase inhibitor", "NF-kB pathway inhibitor" and "Serotonin receptor antagonist", were identified as druggable (e.g., oxprenolol, digoxin, auranofin and palonosetron, respectively) to oppose the effects of severe Dengue infection in these tissues. These are good candidates for future functional evaluation and clinical trials.

Field and Molecular Epidemiology: How Viral Sequencing Changed Transmission Inferences in the First Portuguese SARS-CoV-2 Infection Cluster.

Field epidemiology and viral sequencing provide a comprehensive characterization of transmission chains and allow a better identification of superspreading events. However, very few examples have been presented to date during the COVID-19 pandemic. We studied the first COVID-19 cluster detected in Portugal (59 individuals involved amongst extended family and work environments), following the return of four related individuals from work trips to Italy. The first patient to introduce the virus would be misidentified following the traditional field inquiry alone, as shown by the viral sequencing in isolates from 23 individuals. The results also pointed out family, and not work environment, as the primary mode of transmission.

Inhibition of CXCR2 plays a pivotal role in re-sensitizing ovarian cancer to cisplatin treatment.

cDNA microarray data conducted by our group revealed overexpression of CXCL2 and CXCL8 in ovarian cancer (OC) microenvironment. Herein, we have proven that the chemokine receptor, CXCR2, is a pivotal molecule in re-sensitizing OC to cisplatin, and its inhibition decreases cell proliferation, viability, tumor size in cisplatin-resistant cells, as well as reversed the overexpression of mesenchymal epithelium transition markers. Altogether, our study indicates a central effect of CXCR2 in preventing tumor progression, due to acquisition of cisplatin chemoresistant phenotype by tumor cells, and patients' high lethality rate. We found that the overexpression of CXCR2 by OC cells is persistent and anomalously confined to the cellular nuclei, thus pointing to an urge in developing highly lipophilic molecules that promptly permeate cells, bind to and inhibit nuclear CXCR2 to fight OC, instead of relying on the high-cost genetic engineered cells.

Dynamics of a Dual SARS-CoV-2 Lineage Co-Infection on a Prolonged Viral Shedding COVID-19 Case: Insights into Clinical Severity and Disease Duration.

A few molecularly proven severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cases of symptomatic reinfection are currently known worldwide, with a resolved first infection followed by a second infection after a 48 to 142-day intervening period. We report a multiple-component study of a clinically severe and prolonged viral shedding coronavirus disease 2019 (COVID-19) case in a 17-year-old Portuguese female. She had two hospitalizations, a total of 19 RT-PCR tests, mostly positive, and criteria for releasing from home isolation at the end of 97 days. The viral genome was sequenced in seven serial samples and in the diagnostic sample from her infected mother. A human genome-wide array (>900 K) was screened on the seven samples, and in vitro culture was conducted on isolates from three late samples. The patient had co-infection by two SARS-CoV-2 lineages, which were affiliated in distinct clades and diverging by six variants. The 20A lineage was absolute at the diagnosis (shared with the patient's mother), but nine days later, the 20B lineage had 3% frequency, and two months later, the 20B lineage had 100% frequency. The 900 K profiles confirmed the identity of the patient in the serial samples, and they allowed us to infer that she had polygenic risk scores for hospitalization and severe respiratory disease within the normal distributions for a Portuguese population cohort. The early-on dynamic co-infection may have contributed to the severity of COVID-19 in this otherwise healthy young patient, and to her prolonged SARS-CoV-2 shedding profile.

Shedding Light on the African Enigma: In Vitro Testing of Homo sapiens-Helicobacter pylori Coevolution.

The continuous characterization of genome-wide diversity in population and case-cohort samples, allied to the development of new algorithms, are shedding light on host ancestry impact and selection events on various infectious diseases. Especially interesting are the long-standing associations between humans and certain bacteria, such as the case of Helicobacter pylori, which could have been strong drivers of adaptation leading to coevolution. Some evidence on admixed gastric cancer cohorts have been suggested as supporting Homo-Helicobacter coevolution, but reliable experimental data that control both the bacterium and the host ancestries are lacking. Here, we conducted the first in vitro coinfection assays with dual human- and bacterium-matched and -mismatched ancestries, in African and European backgrounds, to evaluate the genome wide gene expression host response to H. pylori. Our results showed that: (1) the host response to H. pylori infection was greatly shaped by the human ancestry, with variability on innate immune system and metabolism; (2) African human ancestry showed signs of coevolution with H. pylori while European ancestry appeared to be maladapted; and (3) mismatched ancestry did not seem to be an important differentiator of gene expression at the initial stages of infection as assayed here.

Sub-Saharan African information potential to unveil adaptations to infectious disease.

Sub-Saharan Africa is the most promising region of the world to conduct high-throughput studies to unveil adaptations to infectious diseases due to several reasons, namely, the longest evolving time-depth in the Homo sapiens phylogenetic tree (at least two-third older than any other worldwide region); the continuous burden of infectious diseases (still number one in health/life threat); and the coexistence of populations practising diverse subsistence modes (nomadic or seminomadic hunter-gatherers and agropastoralists, and sedentary agriculturalists, small urban and megacity groups). In this review, we will present the most up-to-date results that shed light on three main hypotheses related with this adaptation. One is the hypothesis of coevolution between host and pathogen, given enough time for the establishment of this highly dynamic relationship. The second hypothesis enunciates that the agricultural transition was responsible for the increase of the infectious disease burden, due to the huge expansion of the sedentary human population and the cohabitation with domesticates as main reservoirs of pathogens. The third hypothesis states that the boosting of our immune system against pathogens by past selection may have resulted in maladaptation of the developed hygienic societies, leading to an increase of allergic, inflammatory and autoimmune disorders. Further work will enlighten the biological mechanisms behind these main adaptations, which can be insightful for translation into diagnosis, prognosis and treatment interventions.

Chitinase 3-like-1 and fibronectin in the cargo of extracellular vesicles shed by human macrophages influence pancreatic cancer cellular response to gemcitabine.

Tumour-associated macrophages have been implicated in pancreatic ductal adenocarcinoma (PDAC) therapy response and Extracellular vesicles (EVs) shed by macrophages might have a role in this process. Here, we demonstrated that large EVs released by anti-inflammatory human macrophages decreased PDAC cellular sensitivity to gemcitabine. Using proteomic analysis, chitinase 3-like-1 (CHI3L1) and fibronectin (FN1) were identified as two of the most abundant proteins in the cargo of macrophages-derived EVs. Overexpression of CHI3L1 and FN1, using recombinant human proteins, induced PDAC cellular resistance to gemcitabine through ERK (extracellular-signal-regulated kinase) activation. Inhibition of CHI3L1 and FN1 by pentoxifylline and pirfenidone, respectively, partially reverted gemcitabine resistance. In PDAC patient samples, CHI3L1 and FN1 were expressed in the stroma, associated with the high presence of macrophages. The Cancer Genome Atlas analysis revealed an association between CHI3L1 and FN1 gene expression, overall survival of PDAC patients, gemcitabine response, and macrophage infiltration. Altogether, our data identifies CHI3L1 and FN1 as potential targets for pharmacological inhibition in PDAC. Further pre-clinical in vivo work is warranted to study the possibility of repurposing pentoxifylline and pirfenidone as adjuvant therapies for PDAC treatment.

Gastric Microbiome Diversities in Gastric Cancer Patients from Europe and Asia Mimic the Human Population Structure and Are Partly Driven by Microbiome Quantitative Trait Loci.

The human gastrointestinal tract harbors approximately 100 trillion microorganisms with different microbial compositions across geographic locations. In this work, we used RNASeq data from stomach samples of non-disease (164 individuals from European ancestry) and gastric cancer patients (137 from Europe and Asia) from public databases. Although these data were intended to characterize the human expression profiles, they allowed for a reliable inference of the microbiome composition, as confirmed from measures such as the genus coverage, richness and evenness. The microbiome diversity (weighted UniFrac distances) in gastric cancer mimics host diversity across the world, with European gastric microbiome profiles clustering together, distinct from Asian ones. Despite the confirmed loss of microbiome diversity from a healthy status to a cancer status, the structured profile was still recognized in the disease condition. In concordance with the parallel host-bacteria population structure, we found 16 human loci (non-synonymous variants) in the European-descendent cohorts that were significantly associated with specific genera abundance. These microbiome quantitative trait loci display heterogeneity between population groups, being mainly linked to the immune system or cellular features that may play a role in enabling microbe colonization and inflammation.

The mitogenome portrait of Umbria in Central Italy as depicted by contemporary inhabitants and pre-Roman remains.

Umbria is located in Central Italy and took the name from its ancient inhabitants, the Umbri, whose origins are still debated. Here, we investigated the mitochondrial DNA (mtDNA) variation of 545 present-day Umbrians (with 198 entire mitogenomes) and 28 pre-Roman individuals (obtaining 19 ancient mtDNAs) excavated from the necropolis of Plestia. We found a rather homogeneous distribution of western Eurasian lineages across the region, with few notable exceptions. Contemporary inhabitants of the eastern part, delimited by the Tiber River and the Apennine Mountains, manifest a peculiar mitochondrial proximity to central-eastern Europeans, mainly due to haplogroups U4 and U5a, and an overrepresentation of J (30%) similar to the pre-Roman remains, also excavated in East Umbria. Local genetic continuities are further attested to by six terminal branches (H1e1, J1c3, J2b1, U2e2a, U8b1b1 and K1a4a) shared between ancient and modern mitogenomes. Eventually, we identified multiple inputs from various population sources that likely shaped the mitochondrial gene pool of ancient Umbri over time, since early Neolithic, including gene flows with central-eastern Europe. This diachronic mtDNA portrait of Umbria fits well with the genome-wide population structure identified on the entire peninsula and with historical sources that list the Umbri among the most ancient Italic populations.

Hypoxia and Macrophages Act in Concert Towards a Beneficial Outcome in Colon Cancer.

In colon cancer, the prognostic value of macrophages is controversial, and it is still unknown how hypoxia modulates macrophage-cancer cell crosstalk. To unravel this, co-cultures of human primary macrophages and colon cancer cells were performed at 20% and 1% O2, followed by characterization of both cellular components. Different colon cancer patient cohorts were analyzed for hypoxia and immune markers, and their association with patient overall survival was established. A positive correlation between HIF1A and CD68 in colon cancer patients was identified but, unexpectedly, in cases with higher macrophage infiltration, HIF1A expression was associated with a better prognosis, in contrast to breast, gastric, and lung cancers. Under hypoxia, co-cultures' secretome indicated a shift towards a pro-inflammatory phenotype. These alterations occurred along with increased macrophage phagocytic activity and decreased SIRPα expression. Cancer cells were more invasive and exhibited higher CD47 expression. We hypothesize that the better prognosis associated with HIF1AHighCD68High tumors could occur due to macrophagic pro-inflammatory pressure. Indeed, we found that tumors HIF1AHighCD68High expressed increased levels of CD8A, which is positively correlated with HIF1A. In conclusion, we show that in colon cancer, hypoxia drives macrophages into a pro-inflammatory phenotype, concomitant with increased infiltration of anti-tumor immune cells, favoring better disease outcome.

A panel of intestinal differentiation markers (CDX2, GPA33, and LI-cadherin) identifies gastric cancer patients with favourable prognosis.

BACKGROUND: Gastric cancer is the fifth most common cancer and the third cause of global cancer mortality. CDX2 is an intestinal differentiation marker with prognostic value in gastric cancer and transcriptionally regulates the expression of glycoprotein A33 (GPA33) and liver intestine cadherin (LI-cadherin). METHODS: This study evaluated the clinical significance of the combined expression of CDX2 and its targets GPA33 and LI-cadherin in gastric cancer by fluorescence-based multiplex immunohistochemistry together with digital image analysis and chromogenic immunohistochemistry in 329 gastric cancer samples arranged in tissue microarrays. Additionally, publicly available RNA-seq expression data from 354 gastric cancer samples from the TCGA database were used to validate the immunohistochemistry results. RESULTS: Expression of the three markers (CDX2, GPA33, and LI-cadherin) was strongly correlated, defining an intestinal differentiation panel. Low or negative protein expression of the intestinal differentiation panel identified patients with particularly poor overall survival, irrespective of the methodology used, and was validated in the independent series at the RNA-seq level. CONCLUSIONS: Expression of the intestinal differentiation panel (CDX2, GPA33, and LI-cadherin) defines a set of biomarkers with a strong biological rationale and favourable impact for prognostication of gastric cancer patients.

Helicobacter Pylori Targets the EPHA2 Receptor Tyrosine Kinase in Gastric Cells Modulating Key Cellular Functions.

Helicobacter pylori, a stomach-colonizing Gram-negative bacterium, is the main etiological factor of various gastroduodenal diseases, including gastric adenocarcinoma. By establishing a life-long infection of the gastric mucosa, H. pylori continuously activates host-signaling pathways, in particular those associated with receptor tyrosine kinases. Using two different gastric epithelial cell lines, we show that H. pylori targets the receptor tyrosine kinase EPHA2. For long periods of time post-infection, H. pylori induces EPHA2 protein downregulation without affecting its mRNA levels, an effect preceded by receptor activation via phosphorylation. EPHA2 receptor downregulation occurs via the lysosomal degradation pathway and is independent of the H.pylori virulence factors CagA, VacA, and T4SS. Using small interfering RNA, we show that EPHA2 knockdown affects cell-cell and cell-matrix adhesion, invasion, and angiogenesis, which are critical cellular processes in early gastric lesions and carcinogenesis mediated by the bacteria. This work contributes to the unraveling of the underlying mechanisms of H. pylori-host interactions and associated diseases. Additionally, it raises awareness for potential interference between H. pylori infection and the efficacy of gastric cancer therapies targeting receptors tyrosine kinases, given that infection affects the steady-state levels and dynamics of some receptor tyrosine kinases (RTKs) and their signaling pathways.