A cytokine-like factor 1 homolog acts as a macrophage chemoattractant in grass carp.

Cytokine-like factor 1 (CYTL1) is a small cytokine and has diverse biological functions in mammals. However, whether CYTL1 exists in lower vertebrates is not clear. In this study, we identified cytl homologs in fish and characterized the immune functions in a teleost species, grass carp (Ctenopharyngodon idella). Fish CYTL1 homologs share conserved molecular features with their mammalian counterparts, including 6 cysteine residues in the mature peptide, genomic organization and synteny. Gene expression analysis revealed that cytl1 was constitutively expressed in tissues of grass carp, with the highest expression detected in the heart. Upon infection with Aeromonas hydrophila (A. hydrophila), cytl1 was downregulated in the hindgut, head kidney, skin, and spleen. In the primary head kidney leukocytes (HKLs), stimulation with inactivated A. hydrophila, LPS, poly(I:C), IL-22, IFN-a or IFN-γrel resulted in downregulation of cytl1 expression. Recombinant grass carp CYTL1 protein produced in the HEK293-F cells was potent to induce il-10 expression, but had little effect on the expression of il-1ß and il-6. In vivo experiments revealed that CYTL1 was effective to recruit macrophages to the muscle injected with cytl expression plasmids. Taken together, our results indicate that CYTL1 is a potent chemokine for recruitment of macrophages in fish.

Multi-omics Analysis Identifies Hypoxia Subtypes and S100A2 as an Immunosuppressive Factor in Cervical Cancer.

Cervical cancer is a common gynecological oncology. Growing evidence indicates hypoxia plays an important role in tumor progression and immunity. However, no study has examined the hypoxia landscape in cervical cancer. In this study, using hierarchical clustering, we identified three hypoxia subtypes in cervical cancer samples from The Cancer Genome Atlas dataset according to formerly described hypoxia-related genes. The overall survival time, hypoxic features, genomics, and immunological characteristics of these subtypes existed distinct differences. We also created a hypoxia score by principle component analysis for dimension reduction. The hypoxiaScore was an effective prognostic biomarker validated by GSE44001 and was associated with immunotherapy response. Furthermore, combined with single-cell RNA-sequence (scRNA-seq) and experiments, S100A2 was identified as an immunosuppressive factor induced by hypoxia and regulated expression of PD-L1. S100A2 also served as an oncogene promoting the proliferation and migration of cervical cancer cells. These findings depicted a new hypoxia-based classification and identified S100A2 as a potential therapeutic target for cervical cancer, thereby advancing the understanding of immunotherapy resistance mechanisms and cervical cancer genetic markers.

A2BAR Antagonism Decreases the Glomerular Expression and Secretion of Chemoattractants for Monocytes and the Pro-Fibrotic M2 Macrophages Polarization during Diabetic Nephropathy.

Some chemoattractants and leukocytes such as M1 and M2 macrophages are known to be involved in the development of glomerulosclerosis during diabetic nephropathy (DN). In the course of diabetes, an altered and defective cellular metabolism leads to the increase in adenosine levels, and thus to changes in the polarity (M1/M2) of macrophages. MRS1754, a selective antagonist of the A2B adenosine receptor (A2BAR), attenuated glomerulosclerosis and decreased macrophage-myofibroblast transition in DN rats. Therefore, we aimed to investigate the effect of MRS1754 on the glomerular expression/secretion of chemoattractants, the intraglomerular infiltration of leukocytes, and macrophage polarity in DN rats. Kidneys/glomeruli of non-diabetic, DN, and MRS1754-treated DN rats were processed for transcriptomic analysis, immunohistopathology, ELISA, and in vitro macrophage migration assays. The transcriptomic analysis identified an upregulation of transcripts and pathways related to the immune system in the glomeruli of DN rats, which was attenuated using MRS1754. The antagonism of the A2BAR decreased glomerular expression/secretion of chemoattractants (CCL2, CCL3, CCL6, and CCL21), the infiltration of macrophages, and their polarization to M2 in DN rats. The in vitro macrophages migration induced by conditioned-medium of DN glomeruli was significantly decreased using neutralizing antibodies against CCL2, CCL3, and CCL21. We concluded that the pharmacological blockade of the A2BAR decreases the transcriptional expression of genes/pathways related to the immune response, protein expression/secretion of chemoattractants, as well as the infiltration of macrophages and their polarization toward the M2 phenotype in the glomeruli of DN rats, suggesting a new mechanism implicated in the antifibrotic effect of MRS1754.

High expression of S100A2 predicts poor prognosis in patients with endometrial carcinoma.

BACKGROUND: S100A2, a member of the S100 protein family, is abnormally expressed and plays a vital role in multiple cancers. However, little is known about the clinical significance of S100A2 in endometrial carcinoma. METHODS: Clinicopathological data were obtained from The Cancer Genome Atlas (TCGA), Genotype-Tissue Expression (GTEx), Gene Expression Omnibus (GEO), and Clinical Proteomic Tumor Analysis Consortium (CPTAC). First, the expression and prognostic value of different S100 family members in endometrial carcinoma were evaluated. Subsequently, the Kaplan-Meier plotter and Cox regression analysis were used to assess the prognostic significance of S100A2, while the association between S100A2 expression and clinical characteristics in endometrial carcinoma was also analyzed using logistic regression. A receiver operating characteristic (ROC) curve and a nomogram were constructed. The putative underlying cellular mechanisms were explored using Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis and gene set enrichment analysis (GSEA). RESULTS: Our results revealed that S100A2 expression was significantly higher in endometrial carcinoma tissue than in non-cancerous tissue at both the mRNA and protein levels. Analysis of Kaplan-Meier plotter data revealed that patients with high S100A2 expression had shorter overall survival (OS) and disease specific survival (DSS) compared with those of patients with low S100A2 expression. Multivariate Cox analysis further confirmed that high S100A2 expression was an independent risk factor for OS in patients with endometrial carcinoma. Other clinicopathologic features found to be related to worse prognosis in endometrial carcinoma included age, clinical stage, histologic grade, and tumor invasion. Importantly, ROC analysis also confirmed that S100A2 has a high diagnostic value in endometrial carcinoma. KEGG enrichment analysis and GSEA revealed that the estrogen and IL-17 signaling pathways were significantly upregulated in the high S100A2 expression group, in which estrogen response, JAK-STAT3, K-Ras, and TNFα/NF-κB were differentially enriched. CONCLUSIONS: S100A2 plays an important role in endometrial carcinoma progression and may represent an independent diagnostic and prognostic biomarker for endometrial carcinoma.

SepA Enhances Shigella Invasion of Epithelial Cells by Degrading Alpha-1 Antitrypsin and Producing a Neutrophil Chemoattractant.

Shigella spp. are highly adapted pathogens that cause bacillary dysentery in human and nonhuman primates. An unusual feature of Shigella pathogenesis is that this organism invades the colonic epithelia from the basolateral pole. Therefore, it has evolved the ability to disrupt the intestinal epithelial barrier to reach the basolateral surface. We have shown previously that the secreted serine protease A (SepA), which belongs to the family of serine protease autotransporters of Enterobacteriaceae, is responsible for the initial destabilization of the intestinal epithelial barrier that facilitates Shigella invasion. However, the mechanisms used by SepA to regulate this process remain unknown. To investigate the protein targets cleaved by SepA in the intestinal epithelium, we incubated a sample of homogenized human colon with purified SepA or with a catalytically inactive mutant of this protease. We discovered that SepA targets an array of 18 different proteins, including alpha-1 antitrypsin (AAT), a major circulating serine proteinase inhibitor in humans. In contrast to other serine proteases, SepA cleaved AAT without forming an inhibiting complex, which resulted in the generation of a neutrophil chemoattractant. We demonstrated that the products of the AAT-SepA reaction induce a mild but significant increase in neutrophil transepithelial migration in vitro. Moreover, the presence of AAT during Shigella infection stimulated neutrophil migration and dramatically enhanced the number of bacteria invading the intestinal epithelium in a SepA-dependent manner. We conclude that by cleaving AAT, SepA releases a chemoattractant that promotes neutrophil migration, which in turn disrupts the intestinal epithelial barrier to enable Shigella invasion. IMPORTANCEShigella is the second leading cause of diarrheal death globally. In this study, we identified the host protein targets of SepA, Shigella's major protein secreted in culture. We demonstrated that by cleaving AAT, a serine protease inhibitor important to protect surrounding tissue at inflammatory sites, SepA releases a neutrophil chemoattractant that enhances Shigella invasion. Moreover, SepA degraded AAT without becoming inhibited by the cleaved product, and SepA catalytic activity was enhanced at higher concentrations of AAT. Activation of SepA by an excess of AAT may be physiologically relevant at the early stages of Shigella infection, when the amount of synthesized SepA is very low compared to the concentration of AAT in the intestinal lumen. This observation may also help to explain the adeptness of Shigella infectivity at low dose, despite the requirement of reaching the basolateral side to invade and colonize the colonic epithelium.

Single-cell transcriptomics reveals heterogeneous progression and EGFR activation in pancreatic adenosquamous carcinoma.

Pancreatic adenosquamous carcinoma (PASC) - a rare pathological pancreatic cancer (PC) type - has a poor prognosis due to high malignancy. To examine the heterogeneity of PASC, we performed single-cell RNA sequencing (scRNA-seq) profiling with sample tissues from a healthy donor pancreas, an intraductal papillary mucinous neoplasm, and a patient with PASC. Of 9,887 individual cells, ten cell subpopulations were identified, including myeloid, immune, ductal, fibroblast, acinar, stellate, endothelial, and cancer cells. Cancer cells were divided into five clusters. Notably, cluster 1 exhibited stem-like phenotypes expressing UBE2C, ASPM, and TOP2A. We found that S100A2 is a potential biomarker for cancer cells. LGALS1, NPM1, RACK1, and PERP were upregulated from ductal to cancer cells. Furthermore, the copy number variations in ductal and cancer cells were greater than in the reference cells. The expression of EREG, FCGR2A, CCL4L2, and CTSC increased in myeloid cells from the normal pancreas to PASC. The gene sets expressed by cancer-associated fibroblasts were enriched in the immunosuppressive pathways. We demonstrate that EGFR-associated ligand-receptor pairs are activated in ductal-stromal cell communications. Hence, this study revealed the heterogeneous variations of ductal and stromal cells, defined cancer-associated signaling pathways, and deciphered intercellular interactions following PASC progression.

Non-Genetic Diversity in Chemosensing and Chemotactic Behavior.

Non-genetic phenotypic diversity plays a significant role in the chemotactic behavior of bacteria, influencing how populations sense and respond to chemical stimuli. First, we review the molecular mechanisms that generate phenotypic diversity in bacterial chemotaxis. Next, we discuss the functional consequences of phenotypic diversity for the chemosensing and chemotactic performance of single cells and populations. Finally, we discuss mechanisms that modulate the amount of phenotypic diversity in chemosensory parameters in response to changes in the environment.

McpT, a Broad-Range Carboxylate Chemoreceptor in Sinorhizobium meliloti.

Chemoreceptors enable the legume symbiont Sinorhizobium meliloti to detect and respond to specific chemicals released from their host plant alfalfa, which allows the establishment of a nitrogen-fixing symbiosis. The periplasmic region (PR) of transmembrane chemoreceptors act as the sensory input module for chemotaxis systems via binding of specific ligands, either directly or indirectly. S. meliloti has six transmembrane and two cytosolic chemoreceptors. However, the function of only three of the transmembrane receptors have been characterized so far, with McpU, McpV, and McpX serving as general amino acid, short-chain carboxylate, and quaternary ammonium compound sensors, respectively. In the present study, we analyzed the S. meliloti chemoreceptor McpT. High-throughput differential scanning fluorimetry assays, using Biolog phenotype microarray plates, identified 15 potential ligands for McpTPR, with the majority classified as mono-, di-, and tricarboxylates. S. meliloti exhibited positive chemotaxis toward seven selected carboxylates, namely, α-ketobutyrate, citrate, glyoxylate, malate, malonate, oxalate, and succinate. These carboxylates were detected in seed exudates of the alfalfa host. Deletion of mcpT resulted in a significant decrease of chemotaxis to all carboxylates except for citrate. Isothermal titration calorimetry revealed that McpTPR bound preferentially to the monocarboxylate glyoxylate and with lower affinity to the dicarboxylates malate, malonate, and oxalate. However, no direct binding was detected for the remaining three carboxylates that elicited an McpT-dependent chemotaxis response. Taken together, these results demonstrate that McpT is a broad-range carboxylate chemoreceptor that mediates chemotactic response via direct ligand binding and an indirect mechanism that needs to be identified. IMPORTANCE Nitrate pollution is one of the most widespread and challenging environmental problems that is mainly caused by the agricultural overapplication of nitrogen fertilizers. Biological nitrogen fixation by the endosymbiont Sinorhizobium meliloti enhances the growth of its host Medicago sativa (alfalfa), which also efficiently supplies the soil with nitrogen. Establishment of the S. meliloti-alfalfa symbiosis relies on the early exchange and recognition of chemical signals. The present study contributes to the disclosure of this complex molecular dialogue by investigating the underlying mechanisms of carboxylate sensing in S. meliloti. Understanding individual steps that govern the S. meliloti-alfalfa molecular cross talk helps in the development of efficient, commercial bacterial inoculants that promote the growth of alfalfa, which is the most cultivated forage legume in the world, and improves soil fertility.

CXCR4/CXCL12-mediated entrapment of axons at the injury site compromises optic nerve regeneration.

Regenerative failure in the mammalian optic nerve is generally attributed to axotomy-induced retinal ganglion cell (RGC) death, an insufficient intrinsic regenerative capacity, and an extrinsic inhibitory environment. Here, we show that a chemoattractive CXCL12/CXCR4-dependent mechanism prevents the extension of growth-stimulated axons into the distal nerve. The chemokine CXCL12 is chemoattractive toward axonal growth cones in an inhibitory environment, and these effects are entirely abolished by the specific knockout of its receptor, CXCR4 (CXCR4-/-), in cultured regenerating RGCs. Notably, 8% of naïve RGCs express CXCL12 and transport the chemokine along their axons in the nerve. Thus, axotomy causes its release at the injury site. However, most osteopontin-positive α-RGCs, the main neuronal population that survives optic nerve injury, express CXCR4 instead. Thus, CXCL12-mediated attraction prevents growth-stimulated axons from regenerating distally in the nerve, indicated by axons returning to the lesion site. Accordingly, specific depletion of CXCR4 in RGC reduces aberrant axonal growth and enables long-distance regeneration. Likewise, CXCL12 knockout in RGCs fully mimics these CXCR4-/- effects. Thus, active CXCL12/CXCR4-mediated entrapment of regenerating axons to the injury site contributes to regenerative failure in the optic nerve.

miR-325-3p Promotes the Proliferation, Invasion, and EMT of Breast Cancer Cells by Directly Targeting S100A2.

This study was designed to investigate the precise mechanisms of miR-325-3p/S100A2 axis in breast cancer (BC). In this study, we found that the level of miR-325-3p was dramatically increased in BC tissues and cell lines, and the expression of S100A2 was significantly decreased. Also, the high level of miR-325-3p was closely associated with low expression of S100A2 in BC tissues. Moreover, introduction of miR-325-3p significantly promoted proliferation, invasion, and EMT of BC cells. Bioinformatics analysis predicted that the S100A2 was a potential target gene of miR-325-3p. Luciferase reporter assay demonstrated that miR-325-3p could directly target S100A2. In addition, miR-325-3p overexpression had similar effects with knockdown of S100A2 on BC cells. Overexpression of S100A2 in BC cells partially reversed the promoted effects of miR-325-3p mimic. Overexpression of miR-325-3p promoted cell proliferation, invasion, and EMT of BC cells by directly downregulating S100A2 expression.

S100A2 Silencing Relieves Epithelial-Mesenchymal Transition in Pulmonary Fibrosis by Inhibiting the Wnt/ß-Catenin Signaling Pathway.

Pulmonary fibrosis (PF) is a progressive and lethal disease with poor prognosis. S100A2 plays an important role in the progression of cancer. However, the role of S100A2 in PF has not yet been reported. In this study, we explored the potential role of S100A2 in PF and its potential molecular mechanisms. Increased expression of S100A2 was first observed in lung tissues of PF patients. We found that downregulation of S100A2 inhibited the transforming growth factor-ß1 (TGF-ß1)-induced epithelial-mesenchymal transition (EMT) in A549 cells. Mechanically, TGF-ß1 upregulated ß-catenin and the phosphorylation of glycogen synthase kinase-3ß, which was blocked by silencing S100A2 in vitro. Furthermore, lithium chloride (activator of the Wnt/ß-catenin signaling pathway) effectively rescued S100A2 knockdown-mediated inhibition of EMT in PF. In conclusion, these findings demonstrate that downregulation of S100A2 alleviated PF through inhibiting EMT. S100A2 is a promising potential target for further understanding the mechanism and developing a strategy for the treatment of PF and other EMT-associated diseases.

Inactivation of the core cheVAWY chemotaxis genes disrupts chemotactic motility and organised biofilm formation in Campylobacter jejuni.

Flagellar motility plays a central role in the bacterial foodborne pathogen Campylobacter jejuni, as flagellar motility is required for reaching the intestinal epithelium and subsequent colonisation or disease. Flagellar proteins also contribute strongly to biofilm formation during transmission. Chemotaxis is the process directing flagellar motility in response to attractant and repellent stimuli, but its role in biofilm formation of C. jejuni is not well understood. Here we show that inactivation of the core chemotaxis genes cheVAWY in C. jejuni strain NCTC 11168 affects both chemotactic motility and biofilm formation. Inactivation of any of the core chemotaxis genes (cheA, cheY, cheV or cheW) impaired chemotactic motility but did not affect flagellar assembly or growth. The ∆cheY mutant swam in clockwise loops, while complementation restored normal motility. Inactivation of the core chemotaxis genes interfered with the ability to form a discrete biofilm at the air-media interface, and the ∆cheY mutant displayed reduced dispersal/shedding of bacteria into the planktonic fraction. This suggests that while the chemotaxis system is not required for biofilm formation per se, it is necessary for organized biofilm formation. Hence interference with the Campylobacter chemotaxis system at any level disrupts optimal chemotactic motility and transmission modes such as biofilm formation.

Identification of cisplatin-resistant factor by integration of transcriptomic and proteomic data using head and neck carcinoma cell lines.

Cisplatin is an important drug for the treatment of head and neck squamous cell carcinoma (HNSCC). Determining chemoresistant factors prior to treatment will lead to great benefits for clinicians and patients. Here, we evaluated chemoresistant factors by integrating proteomic and transcriptomic data using HNSCC cell lines to identify a more precise chemoresistant factor in HNSCC. We used four HNSCC cell lines: cisplatin-sensitive, acquired cisplatin resistance, naturally cisplatin-resistant, and acquired 5-FU resistance. Proteomic analysis was performed using iTRAQ, tandem mass spectrometry, and liquid chromatography-electrospray ionization-tandem mass spectrometry. Transcriptomic analysis was performed using microarrays. By integrating these independent data, common factors were addressed and functional analysis was performed using small interfering RNAs (siRNAs) to change the chemosensitivity. Using iTRAQ analysis, 7 proteins were identified as specific for cisplatin chemoresistance factors. Transcriptomic analysis revealed hundreds of potential candidate factors. By combining and integrating these data, S100A2 was identified as a potential cisplatin-specific chemoresistance factor. Functional analysis with siRNA revealed that the expression of S100A2 was reduced and cisplatin sensitivity recovered in the acquired and naturally cisplatin-resistant cell lines, but not in the cisplatin-sensitive cell lines. S100A2 was identified as a cisplatin-specific chemoresistance factor by integrating the transcriptomic and proteomic results obtained using HNSCC cell lines. This is a novel technique that allows for a precise identification, also known as a comprehensive analysis. Our findings indicate that these proteins could be used as biomarkers of HNSCC treatments, providing physicians with new treatment strategies for patients with HNSCC, showing chemoresistance.

Expression Profile of S100A2 and its Clinicopathological Significance in Renal Cell Carcinoma.

BACKGROUND/AIM: Dysregulated expression of S100 protein family members, including S100A2, has been reported in various types of human malignant tumors; however, the functional role of S100A2 in renal cell carcinoma (RCC) remains unknown. The objective of this study was to assess the S100A2 expression pattern and its clinicopathological significance in RCC. MATERIALS AND METHODS: This study investigated the expression profiles of S100A2 mRNA, S100A2 protein and p53 mRNA in addition to S100A2 DNA methylation levels in 3 human RCC cell lines and 81 surgically resected RCC specimens. These findings were analyzed according to several clinicopathological parameters. RESULTS: In all RCC cell lines, both S100A2 mRNA and protein were barely detectable, and the S100A2 promoter was considered to be methylated. S100A2 mRNA expression levels in RCC tissues were markedly lower than those in normal kidney tissues. The rate in clear cell RCC (ccRCC) specimens with higher expression of S100A2 mRNA was significantly lower than that in non-ccRCC specimens (29.9 versus 71.4%, respectively). Furthermore, S100A2 expression in ccRCC specimens was significantly correlated with p53 mRNA expression, but not conventional clinicopathological parameters. CONCLUSION: These findings suggest limited roles of S100A2 in the progression of RCC, particularly ccRCC.

Mutually exclusive dendritic arbors in C. elegans neurons share a common architecture and convergent molecular cues.

Stress-induced changes to the dendritic architecture of neurons have been demonstrated in numerous mammalian and invertebrate systems. Remodeling of dendrites varies tremendously among neuron types. During the stress-induced dauer stage of Caenorhabditis elegans, the IL2 neurons arborize to cover the anterior body wall. In contrast, the FLP neurons arborize to cover an identical receptive field during reproductive development. Using time-course imaging, we show that branching between these two neuron types is highly coordinated. Furthermore, we find that the IL2 and FLP arbors have a similar dendritic architecture and use an identical downstream effector complex to control branching; however, regulation of this complex differs between stress-induced IL2 branching and FLP branching during reproductive development. We demonstrate that the unfolded protein response (UPR) sensor IRE-1, required for localization of the complex in FLP branching, is dispensable for IL2 branching at standard cultivation temperatures. Exposure of ire-1 mutants to elevated temperatures results in defective IL2 branching, thereby demonstrating a previously unknown genotype by environment interaction within the UPR. We find that the FOXO homolog, DAF-16, is required cell-autonomously to control arborization during stress-induced arborization. Likewise, several aspects of the dauer formation pathway are necessary for the neuron to remodel, including the phosphatase PTEN/DAF-18 and Cytochrome P450/DAF-9. Finally, we find that the TOR associated protein, RAPTOR/DAF-15 regulates mutually exclusive branching of the IL2 and FLP dendrites. DAF-15 promotes IL2 branching during dauer and inhibits precocious FLP growth. Together, our results shed light on molecular processes that regulate stress-mediated remodeling of dendrites across neuron classes.

GnRH neurogenesis depends on embryonic pheromone receptor expression.

Gonadotropin-releasing hormone (GnRH) neurons control mammalian reproduction and migrate from their birthplace in the nasal placode to the hypothalamus during development. Despite much work on the origin and migration of GnRH neurons, the processes that control GnRH lineage formation are not fully understood. Here, we demonstrate that Nhlh genes control vomeronasal receptor expression in the developing murine olfactory placode associated with the generation of the first GnRH neurons at embryonic days (E)10-12. Inactivation of ß2-microglobulin (ß2-m), which selectively affects surface expression of V2Rs, dramatically decreased the number of GnRH neurons in the Nhlh2 mutant background, preventing rescue of fertility in female Nhlh2 mutant mice by male pheromones. In addition, we show that GnRH neurons generated after E12 fail to establish synaptic connections to the vomeronasal amygdala, suggesting the existence of functionally specialized subpopulations of GnRH neurons, which process pheromonal information.

Precision Oncology in Surgery: Patient Selection for Operable Pancreatic Cancer.

OBJECTIVE: We aimed to define preoperative clinical and molecular characteristics that would allow better patient selection for operative resection. BACKGROUND: Although we use molecular selection methods for systemic targeted therapies, these principles are not applied to surgical oncology. Improving patient selection is of vital importance for the operative treatment of pancreatic cancer (pancreatic ductal adenocarcinoma). Although surgery is the only chance of long-term survival, 80% still succumb to the disease and approximately 30% die within 1 year, often sooner than those that have unresected local disease. METHOD: In 3 independent pancreatic ductal adenocarcinoma cohorts (total participants = 1184) the relationship between aberrant expression of prometastatic proteins S100A2 and S100A4 and survival was assessed. A preoperative nomogram based on clinical variables available before surgery and expression of these proteins was constructed and compared to traditional measures, and a postoperative nomogram. RESULTS: High expression of either S100A2 or S100A4 was independent poor prognostic factors in a training cohort of 518 participants. These results were validated in 2 independent patient cohorts (Glasgow, n = 198; Germany, n = 468). Aberrant biomarker expression stratified the cohorts into 3 distinct prognostic groups. A preoperative nomogram incorporating S100A2 and S100A4 expression predicted survival and nomograms derived using postoperative clinicopathological variables. CONCLUSIONS: Of those patients with a poor preoperative nomogram score, approximately 50% of patients died within a year of resection. Nomograms have the potential to improve selection for surgery and neoadjuvant therapy, avoiding surgery in aggressive disease, and justifying more extensive resections in biologically favorable disease.

Proteomics of Galápagos Marine Iguanas Links Function of Femoral Gland Proteins to the Immune System.

Communication between individuals via molecules, termed chemosignaling, is widespread among animal and plant species. However, we lack knowledge on the specific functions of the substances involved for most systems. The femoral gland is an organ that secretes a waxy substance involved in chemical communication in lizards. Although the lipids and volatile substances secreted by the femoral glands have been investigated in several biochemical studies, the protein composition and functions of secretions remain completely unknown. Applying a proteomic approach, we provide the first attempt to comprehensively characterize the protein composition of femoral gland secretions from the Galápagos marine iguana. Using samples from several organs, the marine iguana proteome was assembled by next-generation sequencing and MS, resulting in 7513 proteins. Of these, 4305 proteins were present in the femoral gland, including keratins, small serum proteins, and fatty acid-binding proteins. Surprisingly, no proteins with discernible roles in partner recognition or inter-species communication could be identified. However, we did find several proteins with direct associations to the innate immune system, including lysozyme C, antileukoproteinase (ALP), pulmonary surfactant protein (SFTPD), and galectin (LGALS1) suggesting that the femoral glands function as an important barrier to infection. Furthermore, we report several novel anti-microbial peptides from the femoral glands that show similar action against Escherichia coli and Bacillus subtilis such as oncocin, a peptide known for its effectiveness against Gram-negative pathogens. This proteomics data set is a valuable resource for future functional protein analysis and demonstrates that femoral gland secretions also perform functions of the innate immune system.

CUX1 and IκBζ (NFKBIZ) mediate the synergistic inflammatory response to TNF and IL-17A in stromal fibroblasts.

The role of stromal fibroblasts in chronic inflammation is unfolding. In rheumatoid arthritis, leukocyte-derived cytokines TNF and IL-17A work together, activating fibroblasts to become a dominant source of the hallmark cytokine IL-6. However, IL-17A alone has minimal effect on fibroblasts. To identify key mediators of the synergistic response to TNF and IL-17A in human synovial fibroblasts, we performed time series, dose-response, and gene-silencing transcriptomics experiments. Here we show that in combination with TNF, IL-17A selectively induces a specific set of genes mediated by factors including cut-like homeobox 1 (CUX1) and IκBζ (NFKBIZ). In the promoters of CXCL1, CXCL2, and CXCL3, we found a putative CUX1-NF-κB binding motif not found elsewhere in the genome. CUX1 and NF-κB p65 mediate transcription of these genes independent of LIFR, STAT3, STAT4, and ELF3. Transcription of NFKBIZ, encoding the atypical IκB factor IκBζ, is IL-17A dose-dependent, and IκBζ only mediates the transcriptional response to TNF and IL-17A, but not to TNF alone. In fibroblasts, IL-17A response depends on CUX1 and IκBζ to engage the NF-κB complex to produce chemoattractants for neutrophil and monocyte recruitment.

Predicting Angiogenesis by Endothelial Progenitor Cells Relying on In-Vitro Function Assays and VEGFR-2 Expression Levels.

Clinical trials have demonstrated the safety and efficacy of autologous endothelial progenitor cell (EPC) therapy in various diseases. Since EPCs' functions are influenced by genetic, systemic and environmental factors, the therapeutic potential of each individual EPCs is unknown and may affect treatment outcome. Therefore, our aim was to compare EPCs function among healthy donors in order to predict blood vessel formation (angiogenesis) before autologous EPC transplantation. Human EPCs were isolated from the blood of ten volunteers. EPCs proliferation rate, chemoattractant ability, and CXCR4 mRNA levels were different among donors (p < 0.0001, p < 0.01, p < 0.001, respectively). A positive correlation was found between SDF-1, CXCR4, and EPCs proliferation (R = 0.736, p < 0.05 and R = 0.8, p < 0.01, respectively). In-vivo, blood vessels were counted ten days after EPCs transplantation in a subcutaneous mouse model. Mean vessel density was different among donors (p = 0.0001); nevertheless, donors with the lowest vessel densities were higher compared to control (p < 0.05). Finally, using a linear regression model, a mathematical equation was generated to predict blood vessel density relying on: (i) EPCs chemoattractivity, and (ii) VEGFR-2 mRNA levels. Results reveal differences in EPCs functions among healthy individuals, emphasizing the need for a potency assay to pave the way for standardized research and clinical use of human EPCs.