Circulating biomarkers in familial cerebral cavernous malformation.

BACKGROUND: Cerebral Cavernous Malformation (CCM) is a rare cerebrovascular disease, characterized by the presence of multiple vascular malformations that may result in intracerebral hemorrhages (ICHs), seizure(s), or focal neurological deficits (FND). Familial CCM (fCCM) is due to loss of function mutations in one of the three independent genes KRIT1 (CCM1), Malcavernin (CCM2), or Programmed Cell death 10 (PDCD10/CCM3). The aim of this study was to identify plasma protein biomarkers of fCCM to assess the severity of the disease and predict its progression. METHODS: Here, we have investigated plasma samples derived from n = 71 symptomatic fCCM patients (40 female/31 male) and n = 17 healthy donors (HD) (9 female/8 male) of the Phase 1/2 Treat_CCM trial, using multiplexed protein profiling approaches. FINDINGS: Biomarkers as sCD14 (p = 0.00409), LBP (p = 0.02911), CXCL4 (p = 0.038), ICAM-1 (p = 0.02013), ANG2 (p = 0.026), CCL5 (p = 0.00403), THBS1 (p = 0.0043), CRP (p = 0.0092), and HDL (p = 0.027), were significantly different in fCCM compared to HDs. Of note, sENG (p = 0.011), THBS1 (p = 0.011) and CXCL4 (p = 0.011), were correlated to CCM genotype. sROBO4 (p = 0.014), TM (p = 0.026) and CRP (p = 0.040) were able to predict incident adverse clinical events, such as ICH, FND or seizure. GDF-15, FLT3L, CXCL9, FGF-21 and CDCP1, were identified as predictors of the formation of new MRI-detectable lesions over 2-year follow-up. Furthermore, the functional relevance of ang2, thbs1, robo4 and cdcp1 markers was validated by zebrafish pre-clinical model of fCCM. INTERPRETATION: Overall, our study identifies a set of biochemical parameters to predict CCM progression, suggesting biological interpretations and potential therapeutic approaches to CCM disease. FUNDING: Italian Medicines Agency, Associazione Italiana per la Ricerca sul Cancro (AIRC), ERC, Leducq Transatlantic Network of Excellence, Swedish Research Council.

Location and condition based reconstruction of colon cancer microbiome from human RNA sequencing data.

BACKGROUND: The association between microbes and cancer has been reported repeatedly; however, it is not clear if molecular tumour properties are connected to specific microbial colonisation patterns. This is due mainly to the current technical and analytical strategy limitations to characterise tumour-associated bacteria. METHODS: Here, we propose an approach to detect bacterial signals in human RNA sequencing data and associate them with the clinical and molecular properties of the tumours. The method was tested on public datasets from The Cancer Genome Atlas, and its accuracy was assessed on a new cohort of colorectal cancer patients. RESULTS: Our analysis shows that intratumoural microbiome composition is correlated with survival, anatomic location, microsatellite instability, consensus molecular subtype and immune cell infiltration in colon tumours. In particular, we find Faecalibacterium prausnitzii, Coprococcus comes, Bacteroides spp., Fusobacterium spp. and Clostridium spp. to be strongly associated with tumour properties. CONCLUSIONS: We implemented an approach to concurrently analyse clinical and molecular properties of the tumour as well as the composition of the associated microbiome. Our results may improve patient stratification and pave the path for mechanistic studies on microbiota-tumour crosstalk.

GZMKhigh CD8+ T effector memory cells are associated with CD15high neutrophil abundance in non-metastatic colorectal tumors and predict poor clinical outcome.

CD8+ T cells are a major prognostic determinant in solid tumors, including colorectal cancer (CRC). However, understanding how the interplay between different immune cells impacts on clinical outcome is still in its infancy. Here, we describe that the interaction of tumor infiltrating neutrophils expressing high levels of CD15 with CD8+ T effector memory cells (TEM) correlates with tumor progression. Mechanistically, stromal cell-derived factor-1 (CXCL12/SDF-1) promotes the retention of neutrophils within tumors, increasing the crosstalk with CD8+ T cells. As a consequence of the contact-mediated interaction with neutrophils, CD8+ T cells are skewed to produce high levels of GZMK, which in turn decreases E-cadherin on the intestinal epithelium and favors tumor progression. Overall, our results highlight the emergence of GZMKhigh CD8+ TEM in non-metastatic CRC tumors as a hallmark driven by the interaction with neutrophils, which could implement current patient stratification and be targeted by novel therapeutics.

Characterization of skin surface and dermal microbiota in dogs with mast cell tumor.

The skin microbiota interacts with the host immune response to maintain the homeostasis. Changes in the skin microbiota are linked to the onset and the progression of several diseases, including tumors. We characterized the skin surface and dermal microbiota of 11 dogs affected by spontaneous mast cell tumor (MCT), using skin contralateral sites as intra-animal healthy controls. The microbial profile differed between healthy and tumor skin surfaces and dermis, demonstrating that the change in microbiota composition is related to the presence of MCT. The number of observed taxa between MCT and healthy skin surfaces was detected, showing a decrease in number and heterogeneity of taxa over the skin surface of MCT, at both inter- and intra-individual level. Preliminary data on bacterial population of MCT dermis, obtained only on three dogs, demonstrated an intra-individual reduction of taxa number when compared to the skin surface. Taxonomy reveals an increase of Firmicutes phylum and Corynebacteriaceae family in MCT skin surface when compared to the healthy contralateral. In conclusion, we demonstrate that microbial population of skin surface and dermis is related to mast cell tumor. Our study provides the basis for future investigations aiming to better define the interaction between mast cell tumors, microbiota and host immune response.

In-vitro effect of heat stress on bovine monocytes lifespan and polarization.

Heat stress (HS) has a negative impact on dairy cows' health, milk production, reproductive performance and immune defenses. Cellular and molecular responses to high temperatures in bovine polymorphonuclear cells and peripheral blood mononuclear cells (PBMCs) have been investigated so far. On the contrary, the effects of high temperatures on isolated monocytes remain almost undisclosed. The aim of this study was to unravel the in vitro effects of high temperatures, simulating a severe HS related body hyperthermia, on bovine lifespan and M1/M2 polarisation. The PBMCs were isolated from whole blood of 9 healthy dairy cattle. Monocytes were sorted by magnetic activated cell sorting and cultured over night at 39 °C (normothermia) or 41 °C (HS). Apoptotic rate and viability were assessed and mRNA abundance for heat shock proteins (HSPs), heat transcription factors (HSFs) and genes involved in monocyte/macrophage polarization (STAT1, STAT2, STAT3, STAT6, IL1ß, TGF1ß, IL-10, COX2) were quantified by qPCR. We found that apoptosis increased in monocytes exposed to 41 °C, as compared to control, while viability conversely decreased. HS increased the abundance of HSF1 and HSP70. The concomitant decrease of STAT1 and STAT2 and the increase of STAT6 genes abundance at 41 °C suggest, at transcriptional factors level, a polarization of monocytes from a classical activated M1 to a non-classically activated M2 monocytes. In conclusion, the exposure of bovine monocytes to high temperatures affects their lifespan as well as the abundance of genes involved in HS response and in monocyte/macrophages polarization phenotype, confirming that bovine immune response may be significantly affected by hyperthermia.

MicroRNA Expression in Formalin-Fixed, Paraffin-Embedded Samples of Canine Cutaneous and Oral Melanoma by RT-qPCR.

MicroRNAs (miRNAs) are a class of small, noncoding RNA that post-transcriptionally regulate protein expression. miRNAs are emerging as clinical biomarkers of many diseases, including tumors. The aim of this study was to investigate whether miRNA expression could vary in melanoma samples derived from formalin-fixed, paraffin-embedded (FFPE) tissues. The study included 4 groups: (1) 9 samples of oral canine malignant melanoma, (2) 10 samples of cutaneous malignant melanoma, (3) 5 samples of healthy oral mucosa, and (4) 7 samples of healthy skin. The expression levels of 6 miRNAs-miR-145, miR-146a, miR-425-5p, miR-223, miR-365, and miR-134-were detected and assessed by quantitative reverse transcription polymerase chain reaction (RT-qPCR) using TaqMan probes. Cutaneous canine malignant melanoma showed a decrease of the expression level of miR-145 and miR-365 and an increase of miR-146a and miR-425-5p compared to control samples. MiR-145 was also downregulated in oral canine malignant melanoma. The miRNAs with decreased expression may regulate genes involved in RAS, Rap1, and transforming growth factor ß (TGF-ß) signaling pathways, as well as upregulated genes associated with phosphatidylinositol signaling system, adherens junction, and RAS signaling pathways. In conclusion, miR-145, miR-365, miR-146a, and miR-425-5p were differentially expressed in canine malignant melanoma and healthy FFPE samples, suggesting that they may play a role in canine malignant melanoma pathogenesis.