BackgroundThe Signal Transducer and Activator of Transcription 1 (STAT1) has traditionally been regarded as a transmitter of interferon signaling and a pro-apoptotic tumour suppressor. Recent data have identified new functions of STAT1 associated with tumourigenesis and resistance to genotoxic stress, including ionizing radiation (IR) and chemotherapy. To investigate the mechanisms contributing to the tumourigenic functions of STAT1, we performed a combined transcriptomic-proteomic expressional analysis and found that STAT1 is associated with regulation of energy metabolism with potential implication in the Warburg effect.MethodsWe generated a stable knockdown of STAT1 in the SCC61 human squamous cell carcinoma cell line, established tumour xenografts in athymic mice, and compared transcriptomic and proteomic profiles of STAT1 wild-type (WT) and knockdown (KD) untreated or irradiated (IR) tumours. Transcriptional profiling was based on Affymetrix Human GeneChip® Gene 1.0 ST microarrays. Proteomes were determined from the tandem mass spectrometry (MS/MS) data by searching against the human subset of the UniProt database. Data were analysed using Significance Analysis of Microarrays for ribonucleic acid and Visualize software for proteins. Functional analysis was performed with Ingenuity Pathway Analysis with statistical significance measured by Fisher's exact test.ResultsKnockdown of STAT1 led to significant growth suppression in untreated tumours and radio sensitization of irradiated tumours. These changes were accompanied by alterations in the expression of genes and proteins of glycolysis/gluconeogenesis (GG), the citrate cycle (CC) and oxidative phosphorylation (OP). Of these pathways, GG had the most concordant changes in gene and protein expression and demonstrated a STAT1-dependent expression of genes and proteins consistent with tumour-specific glycolysis. In addition, IR drastically suppressed the GG pathway in STAT1 KD tumours without significant change in STAT1 WT tumours.ConclusionOur results identify a previously uncharacterized function of STAT1 in tumours: expressional regulation of genes encoding proteins involved in glycolysis, the citrate cycle and mitochondrial oxidative phosphorylation, with predominant regulation of glycolytic genes. STAT1-dependent expressional regulation of glycolysis suggests a potential role for STAT1 as a transcriptional modulator of genes responsible for the Warburg effect.

BackgroundTraditionally IFN/STAT1 signaling is connected with an anti-viral response and pro-apoptotic tumor-suppressor functions. Emerging functions of a constitutively activated IFN/STAT1 pathway suggest an association with an aggressive tumor phenotype. We hypothesized that tumor clones that constitutively overexpress this pathway are preferentially selected by the host microenvironment due to a resistance to STAT1-dependent cytotoxicity and demonstrate increased metastatic ability combined with increased resistance to genotoxic stress.Methodology/Principal FindingsHere we report that clones of B16F1 tumors grown in the lungs of syngeneic C57BL/6 mice demonstrate variable transcriptional levels of IFN/STAT1 pathway expression. Tumor cells that constitutively overexpress the IFN/STAT1 pathway (STAT1H genotype) are selected by the lung microenvironment. STAT1H tumor cells also demonstrate resistance to IFN-gamma (IFNγ), ionizing radiation (IR), and doxorubicin relative to parental B16F1 and low expressors of the IFN/STAT1 pathway (STAT1L genotype). Stable knockdown of STAT1 reversed the aggressive phenotype and decreased both lung colonization and resistance to genotoxic stress.ConclusionsOur results identify a pathway activated by tumor-stromal interactions thereby selecting for pro-metastatic and therapy-resistant tumor clones. New therapies targeted against the IFN/STAT1 signaling pathway may provide an effective strategy to treat or sensitize aggressive tumor clones to conventional cancer therapies and potentially prevent distant organ colonization.

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Introduction Reliable characterization of a hypertrophic scar (HTS) is integral to epidemiologic studies designed to identify clinical and genetic risk factors for HTS. The Vancouver Scar Scale (VSS) has been widely used for this purpose; however, no publication has defined what score on this scale corresponds to a clinical diagnosis of HTS. Methods In a survey of 1000 burn care providers, we asked respondents what VSS score indicates a HTS and asked them to score scar photos using the VSS. We used receiver-operating-characteristic (ROC) curves to evaluate VSS subscores and their combinations in diagnosis of HTS. Results Of 130 responses (13.5%), most were physicians (43.9%) who had worked in burn care for over 10 years (63.1%) and did not use the VSS in clinical practice (58.5%). There was no consensus as to what VSS score indicates a diagnosis of HTS. VSS height score (0–3) performed best for diagnosis of HTS; using a cut-off of ≥1, height score was 99.5% sensitive and 85.9% specific for HTS. Conclusions Burn clinicians do not routinely use the VSS and perceptions vary widely regarding what constitutes a HTS. When a dichotomous variable is needed, the VSS height score with a cut-off of ≥1 may be optimal. Our findings underscore the need for an objective tool to reproducibly characterize HTS across burn centers.

An siRNA screen targeting 89 IFN stimulated genes in 14 different cancer cell lines pointed to the RIG-I (retinoic acid inducible gene I)-like receptor Laboratory of Genetics and Physiology 2 (LGP2) as playing a key role in conferring tumor cell survival following cytotoxic stress induced by ionizing radiation (IR). Studies on the role of LGP2 revealed the following: (i) Depletion of LGP2 in three cancer cell lines resulted in a significant increase in cell death following IR, (ii) ectopic expression of LGP2 in cells increased resistance to IR, and (iii) IR enhanced LGP2 expression in three cell lines tested. Studies designed to define the mechanism by which LGP2 acts point to its role in regulation of IFNβ. Specifically (i) suppression of LGP2 leads to enhanced IFNβ, (ii) cytotoxic effects following IR correlated with expression of IFNβ inasmuch as inhibition of IFNβ by neutralizing antibody conferred resistance to cell death, and (iii) mouse embryonic fibroblasts from IFN receptor 1 knockout mice are radioresistant compared with wild-type mouse embryonic fibroblasts. The role of LGP2 in cancer may be inferred from cumulative data showing elevated levels of LGP2 in cancer cells are associated with more adverse clinical outcomes. Our results indicate that cytotoxic stress exemplified by IR induces IFNβ and enhances the expression of LGP2. Enhanced expression of LGP2 suppresses the IFN stimulated genes associated with cytotoxic stress by turning off the expression of IFNβ.innate immunity | cytoplasmic sensor | interferon beta | DHX58

The genetic determinants of post-burn hypertrophic scarring (HTS) are unknown, and melanocortin 1 receptor (MC1R) loss-of-function leads to fibrogenesis in experimental models. To examine the associations between self-identified race and MC1R single- nucleotide polymorphisms (SNPs) with severity of post-burn HTS, we conducted a prospective cohort study of burned adults admitted to our institution over 7 years. Subjects were evaluated using the Vancouver Scar Scale (VSS), asked to rate their itching, and genotyped for 8 MC1R SNPs. Testing for association with severe HTS (VSS>7) and itch severity (0-10) was based on multivariate regression with adjustment for known risk factors. Of 425 subjects analyzed, 77% identified as White. The prevalence of severe HTS (VSS>7) was 49%, and the mean itch score was 3.9. In multivariate analysis, Asian (prevalence ratio [PR] 1.54; 95% CI: 1.13-2.10), Black/African American (PR 1.86; 95% CI: 1.42-2.45), and Native American (PR 1.87; 95% CI: 1.48-2.35) race were independently associated with severe HTS. MC1R SNP R163Q was also significantly (P<0.001) associated with severe HTS. Asian race (linear regression coefficient 1.32; 95% CI: 0.23-2.40) but not MC1R SNP genotype was associated with increased itch score. We conclude that MC1R genotype may influence post-burn scarring.

BackgroundHypertrophic scarring (HTS) is hypothesized to have a genetic mechanism, yet its genetic determinants are largely unknown. The mitogen-activated protein kinase (MAPK) pathways are important mediators of inflammatory signaling, and experimental evidence implicates MAPKs in HTS formation. We hypothesized that single-nucleotide polymorphisms (SNPs) in MAPK-pathway genes would be associated with severity of post-burn HTS.MethodsWe analyzed data from a prospective-cohort genome-wide association study of post-burn HTS. We included subjects with deep-partial-thickness burns admitted to our center who provided blood for genotyping and had at least one Vancouver Scar Scale (VSS) assessment. After adjusting for HTS risk factors and population stratification, we tested MAPK-pathway gene SNPs for association with the four VSS variables in a joint regression model. In addition to individual-SNP analysis, we performed gene-based association testing.ResultsOur study population consisted of 538 adults (median age 40 years) who were predominantly White (76%) males (71%) admitted to our center from 2007–2014 with small-to-moderate-sized burns (median burn size 6% total body surface area). Of 2,146 SNPs tested, a rare missense variant in the PTPN5 gene (rs56234898; minor allele frequency 1.5%) was significantly associated with decreased severity of post-burn HTS (P = 1.3×10−6). In gene-based analysis, PTPN5 (P = 1.2×10−5) showed a significant association and BDNF (P = 9.5×10−4) a borderline-significant association with HTS severity.ConclusionsWe report PTPN5 as a novel genetic locus associated with HTS severity. PTPN5 is a MAPK inhibitor expressed in neurons, suggesting a potential role for neurotrophic factors and neuroinflammatory signaling in HTS pathophysiology.

Metabolomics can function as a serial, comprehensive, and potentially personalized tool to characterize metabolism after injury. A targeted metabolomics approach was associated with ongoing oxidative stress, impaired nucleotide synthesis, and initial suppression of protein metabolism followed by increased nitrogen turnover. This technique may provide new therapeutic and nutrition targets in critically injured patients.