Relationship between the transcriptional expression of PIM1 and local control in patients with head and neck squamous cell carcinomas treated with radiotherapy.

PURPOSE: Proviral integration site for Moloney murine leukemia virus (PIMs) are proto-oncogenes encoding serine/threonine kinases that phosphorylate a variety of substrates involved in the regulation of cellular processes. Elevated expression of PIM-1 has been associated with poor prognosis in several types of cancer. There are no studies that have analyzed the response to radiotherapy in patients with head and neck squamous cell carcinoma (HNSCC) according to the expression of PIM-1. The aim of our study was to analyze the relationship between the transcriptional expression of PIM-1 and local response to radiotherapy in HNSCC patients. METHODS: We determined the transcriptional expression of PIM-1 in 135 HNSCC patients treated with radiotherapy, including patients treated with chemoradiotherapy (n = 65) and bioradiotherapy (n = 15). RESULTS: During the follow-up, 48 patients (35.6%) had a local recurrence of the tumor. Patients with local recurrence had a higher level of PIM-1 expression than those who achieved local control of the disease (P = 0.017). Five-year local recurrence-free survival for patients with a high expression of PIM-1 (n = 43) was 44.6% (95% CI 29.2-60.0%), and for patients with low expression (n = 92) it was 71.9% (95% CI 62.5-81.3%) (P = 0.007). According to the results of multivariate analysis, patients with a high PIM-1 expression had a 2.2-fold increased risk of local recurrence (95% CI 1.22-4.10, P = 0.009). CONCLUSION: Patients with elevated transcriptional expression levels of PIM-1 had a significantly higher risk of local recurrence after radiotherapy.

Presence of Ceramidase Activity in Electronegative LDL.

Electronegative low-density lipoprotein (LDL(-)) is a minor modified fraction of human plasma LDL with several atherogenic properties. Among them is increased bioactive lipid mediator content, such as lysophosphatidylcholine (LPC), non-esterified fatty acids (NEFA), ceramide (Cer), and sphingosine (Sph), which are related to the presence of some phospholipolytic activities, including platelet-activating factor acetylhydrolase (PAF-AH), phospholipase C (PLC), and sphingomyelinase (SMase), in LDL(-). However, these enzymes' activities do not explain the increased Sph content, which typically derives from Cer degradation. In the present study, we analyzed the putative presence of ceramidase (CDase) activity, which could explain the increased Sph content. Thin layer chromatography (TLC) and lipidomic analysis showed that Cer, Sph, and NEFA spontaneously increased in LDL(-) incubated alone at 37 °C, in contrast with native LDL(+). An inhibitor of neutral CDase prevented the formation of Sph and, in turn, increased Cer content in LDL(-). In addition, LDL(-) efficiently degraded fluorescently labeled Cer (NBD-Cer) to form Sph and NEFA. These observations defend the existence of the CDase-like activity's association with LDL(-). However, neither the proteomic analysis nor the Western blot detected the presence of an enzyme with known CDase activity. Further studies are thus warranted to define the origin of the CDase-like activity detected in LDL(-).

Circulating microRNAs modulating glycolysis as non-invasive prognostic biomarkers of HNSCC.

BACKGROUND: The identification of prognostic non-invasive biomarkers is a priority for cancer patients' care. Circulating microRNA (miRNAs) have been described in numerous human malignancies as diagnostic, prognostic, and therapeutic cancer biomarkers. The aim of our study was to analyze the expression profile of a set of miRNAs, involved in the modulation of the glycolytic pathway, as prognostic factors in human head and neck squamous cell carcinomas (HNSCC). METHODS: Serum samples of 54 patients with untreated HNSCC were obtained at the time of diagnosis. The prognostic value of circulating miR-26b, miR-124, miR-155 and miR-375 was evaluated towards disease-free survival. RESULTS: We found that there were optimal miRNAs cut-off values for lower risk of recurrence in HNSCC patients. Kaplan-Meier curves showed that higher levels of miR-26b and lower levels of miR-155 were associated with better disease-free survival rates. In the multivariate analysis, patients with serum miR-26b > 0.062 and miR-155 < 0.159 presented more than 2.9 times lower risk of poor outcome. CONCLUSION: Our results suggest that two miRNAs that modulate the glycolytic pathway, miR-26b and miR-155, are independently associated with the risk of recurrence in patients with HNSCC. The overall results in this study supports the evidence that the glucose homeostasis may be a target to improve the outcomes for patients with HNSCC. LEVEL OF EVIDENCE: Individual retrospective cohort study (2b).

Role of the Scavenger Receptor CD36 in Accelerated Diabetic Atherosclerosis.

Diabetes mellitus entails increased atherosclerotic burden and medial arterial calcification, but the precise mechanisms are not fully elucidated. We aimed to investigate the implication of CD36 in inflammation and calcification processes orchestrated by vascular smooth muscle cells (VSMCs) under hyperglycemic and atherogenic conditions. We examined the expression of CD36, pro-inflammatory cytokines, endoplasmic reticulum (ER) stress markers, and mineralization-regulating enzymes by RT-PCR in human VSMCs, cultured in a medium containing normal (5 mM) or high glucose (22 mM) for 72 h with or without oxidized low-density lipoprotein (oxLDL) (24 h). The uptake of 1,1'-dioctadecyl-3,3,3',3-tetramethylindocarbocyanine perchlorate-fluorescently (DiI) labeled oxLDL was quantified by flow cytometry and fluorimetry and calcification assays were performed in VSMC cultured in osteogenic medium and stained by alizarin red. We observed induction in the expression of CD36, cytokines, calcification markers, and ER stress markers under high glucose that was exacerbated by oxLDL. These results were confirmed in carotid plaques from subjects with diabetes versus non-diabetic subjects. Accordingly, the uptake of DiI-labeled oxLDL was increased after exposure to high glucose. The silencing of CD36 reduced the induction of CD36 and the expression of calcification enzymes and mineralization of VSMC. Our results indicate that CD36 signaling is partially involved in hyperglycemia and oxLDL-induced vascular calcification in diabetes.

Enhanced endoplasmic reticulum and mitochondrial stress in abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) is a degenerative vascular disease with a complex aetiology that remains to be fully elucidated. Clinical management of AAA is limited to surgical repair, while an effective pharmacotherapy is still awaited. Endoplasmic reticulum (ER) stress and mitochondrial dysfunction have been involved in the pathogenesis of cardiovascular diseases (CVDs), although their contribution to AAA development is uncertain. Therefore, we aimed to determine their implication in AAA and investigated the profile of oxysterols in plasma, specifically 7-ketocholesterol (7-KC), as an ER stress inducer.In the present study, we determined aortic ER stress activation in a large cohort of AAA patients compared with healthy donors. Higher gene expression of activating transcription factor (ATF) 6 (ATF6), IRE-1, X-binding protein 1 (XBP-1), C/EBP-homologous protein (CHOP), CRELD2 and suppressor/enhancer of Lin-12-like (SEL1L) and greater protein levels of active ATF6, active XBP1 and of the pro-apoptotic protein CHOP were detected in human aneurysmatic samples. This was accompanied by an exacerbated apoptosis, higher reactive oxygen species (ROS) production and by a reduction in mitochondrial biogenesis in the vascular wall of AAA. The quantification of oxysterols, performed by liquid chromatography-(atmospheric pressure chemical ionization (APCI))-mass spectrometry, showed that levels of 7-KC were significantly higher while those of 7α-hydroxycholesterol (HC), 24-HC and 27-HC were lower in AAA patients compared with healthy donors. Interestingly, the levels of 7-KC correlate with the expression of ER stress markers.Our results evidence an induction of ER stress in the vascular wall of AAA patients associated with an increase in circulating 7-KC levels and a reduction in mitochondrial biogenesis suggesting their implication in the pathophysiology of this disease.

Prognostic value of CD45 transcriptional expression in head and neck cancer.

INTRODUCTION: Tumor-infiltrating lymphocytes (TILs) have a recognized antitumor activity in head and neck squamous cell carcinoma (HNSCC). CD45 is one of the most highly expressed proteins in lymphocytes. We carry out a study to assess the prognostic value of transcriptional expression of CD45 in HNSCC. MATERIAL AND METHODS: We determined the transcriptional expression of CD45 in 160 consecutive HNSCC patients and compared the TIL values according to the CD45 expression. RESULTS: Five-year disease-free survival for patients with a high transcriptional expression of CD45 (n = 107) was 62.4% and for patients with a low expression (n = 53) it was 36.2% (P = 0.003). Patients with a high expression of CD45 had a better local recurrence-free survival and disease-specific survival. The results of a multivariate analysis showed that patients with a low expression of CD45 had 2.0-fold high risk of recurrence (95% CI 1.2-3.2, P = 0.003). In oropharyngeal carcinomas, HPV-positive tumors showed a higher transcriptional CD45 expression than HPV-negative tumors. Tumors with high CD45 expression had immunohistochemical TIL scores significantly higher than those with low CD45 expression. CONCLUSION: According to our results, CD45 expression is a potential marker for tumor outcome in HNSCC patients.

Increased Peak Wall Stress, but Not Maximum Diameter, Is Associated with Symptomatic Abdominal Aortic Aneurysm.

OBJECTIVE: Maximum diameter (MD) is the established rupture predictor for abdominal aortic aneurysm (AAA). However, biomechanical markers from finite element analysis (FEA) could be more accurate predictors for these patients. In this study, the association between peak wall stress (PWS) and MD with symptoms of AAA was evaluated. METHODS: Patients diagnosed with infrarenal non-ruptured AAA at the centre between 2009 and 2015 were included. Clinical data, morphological variables (including MD), and the biomechanical variables PWS and diameter normalised PWS (dnPWS) in symptomatic (sAAA) and asymptomatic AAA patients (aAAA) were included. RESULTS: A total of 170 patients were analysed, 153 aAAA and 17 sAAA. MD was significantly greater in sAAA patients than in aAAA patients (70.4 mm, 95% CI 66.4-86.0 vs. 59.1 mm, 95% CI 53.7-67.8, respectively; p = .002). PWS was also significantly higher in the sAAA group (324.6 kPa, 95% CI 217.4-399.5 vs. 199.2 kPa, 95% CI 165.6-239.5; p < .01). No differences in MD were found in patients with an AAA ≥ 65 mm (43 aAAA and 14 sAAA); however, both PWS (327.4 kPa, 95% CI 239.0-473.3 vs. 229.4 kPa, 95% CI 210.0 to 289.4; p = .020) and dnPWS (4.3, 95% CI 3.17-4.67 vs. 3.03, 95% CI 2.8-3.49; p = .004) were higher in sAAA than in aAAA. CONCLUSIONS: This study suggests that MD and the biomechanical parameters obtained by finite element analysis are greater in sAAA than in aAAA. However, considering patients with MD ≥ 65 mm alone, only PWS, and particularly dnPWS, were able to differentiate sAAA from aAAA.

Prostacyclin-synthase expression in head and neck carcinoma patients and its prognostic value in the response to radiotherapy.

Prostacyclin (PGI2 ) plays a role in cancer progression but the mechanism is currently poorly understood. Additionally, no data are available about the prognostic value of the PGI2 pathway in head and neck squamous cell carcinoma (HNSCC) therapy. We evaluated the expression of the PGI2 pathway in HNSCC patients. PGI2 production and PGI synthase (PGIS) expression, in terms of mRNA (RT-PCR) and protein (immunoblotting), were lower in tumour samples than in non-tumoural mucosa, whereas, as expected, COX-2 expression was increased in HNSCC tumour samples. Using local control of the tumour after radiotherapy or chemoradiotherapy as a dependent variable, patients were classified into two categories of PGIS transcript levels. The high-PGIS group had a significantly lower frequency of local and distant failure than the low-PGIS group, and the 5-year cancer-specific survival was higher [90.2% (95% CI 81.0-99.4%) versus 60.5% (95% CI 44.4-76.6%)]. None of the four HNSCC cell lines analysed expressed PGIS and therefore they did not produce PGI2 . However, HNSCC-conditioned media enhanced PGI2 production in endothelial cells (ECs). The stable analogue of PGI2 , carbaprostacyclin (cPGI2 ), exerted little effect on HNSCC cell line migration, and no effect on cell cycle distribution or proliferation rate after radiation injury was observed. Nevertheless, cPGI2 promoted EP-4-dependent in vitro angiogenesis. Von Willebrand factor expression (EC marker) and capillary density were significantly higher in the group of patients with high expression of PGIS. Our results indicate that PGIS expression was associated with radiotherapy efficiency. Although we do not provide direct evidence of a relationship between tumour vascularization and radiotherapy efficiency, our results suggest that the effect of PGI2 is related to its ability to promote vascularization. These results also support the concept that co-adjuvant therapy with PGIS enhancers, such as retinoids, could have therapeutic value for HNSCC treatment.

Expression of the CXCL12/CXCR4 chemokine axis predicts regional control in head and neck squamous cell carcinoma.

Expression of the CXCL12/CXCR4 chemokine axis has been related with the appearance of metastatic recurrence survival, including regional and distant recurrence, in patients with head and neck squamous cell carcinoma (HNSCC). RT-PCR was used to determine mRNA expression levels of CXCL12 and CXCR4 in biopsy tumor samples in 111 patients with HNSCC. Five-year regional recurrence-free survival for patients with low CXCR4 expression (n = 39, 31.5 %) was 97.4 %, for patients with high CXCR4/high CXCL12 expression (n = 22, 19.8 %) it was 94.7 %, and for patients with high CXCR4/low CXCL12 expression (n = 50, 45.0 %) it was 63.3 %. We found significant differences in the regional recurrence-free survival according to CXCR4/CXCL12 expression values (P = 0.001). HNSCC patients with high CXCR4 and low CXCL12 expression values had a significantly higher risk of regional recurrence and could benefit from a more intense treatment of lymph node areas in the neck.

Active smoking increases microsomal PGE2-synthase-1/PGE-receptor-4 axis in human abdominal aortic aneurysms.

BACKGROUND: The cyclooxygenase- (COX-) 2/microsomal PGE-synthase- (mPGES-) 1/PGE-receptor- (EP-) 4 axis could play a key role in the physiopathology of abdominal aortic aneurysm (AAA) in humans. In this study, we investigated the influence of cardiovascular risk factors on the expression of the PGE2 pathway in human AAA. METHODS: Aortic (n = 89) and plasma (n = 79) samples from patients who underwent AAA repair were collected. Patients were grouped according to risk factors. COX-isoenzymes, mPGES-1, EPs, α-actin, and CD45 and CD68 transcripts levels were quantified by QRT-PCR and plasma PGE2 metabolites by EIA. RESULTS: Current smoking (CS) patients compared to no-CS had significantly higher local levels of mPGES-1 (P = 0.009), EP-4 (P = 0.007), and PGE2 metabolites plasma levels (P = 0.008). In the multiple linear regression analysis, these parameters remained significantly enhanced in CS after adding confounding factors. Results from association studies with cell type markers suggested that the increased mPGES-1/EP-4 levels were mainly associated with microvascular endothelial cells. CONCLUSIONS: This study shows that elements of the PGE2 pathway, which play an important role in AAA development, are increased in CS. These results provide insight into the relevance of tobacco smoking in AAA development and reinforce the potential of mPGES-1 and EP-4 as targets for therapy in AAA patients.

Predictive capacity for local disease control of neogenin-1 (NEO1) transcriptional expression in patients with head and neck squamous cell carcinoma.

PURPOSE: To analyze the predictive capacity for local disease control of the transcriptional expression of neogenin-1 (NEO1) gene in patients with head and neck squamous cell carcinoma (HNSCC). METHODS/PATIENTS: A retrospective study was performed on tumor biopsies from 107 patients with HNSCC treated surgically. The transcriptional expression of NEO1 was determined by RT-PCR. NEO1 transcriptional expression value was categorized according to local disease control by recursive partitioning analysis. RESULTS: Lower NEO1 transcriptional expression was associated with worse local control after surgical treatment. Patients with lower NEO1 expression (n = 25, 23.4%) had a 5-year local recurrence-free survival of 61.8% (95% CI: 42.1-81.5%), while patients with higher NEO1 expression (n = 82, 76.6%) had a 5-year local recurrence-free survival of 85.6% (95% CI: 77.6-93.6%), (P = 0.003). According to the result of multivariable analysis, patients with lower NEO1 expression had a 2.7-fold increased risk of local tumor recurrence (95% CI: 1.0-7.0, P = 0.043) compared to patients with higher NEO1 expression. CONCLUSIONS: HNSCC patients with a lower transcriptional expression of NEO1 have a significantly higher risk of local recurrence after surgical treatment.

Microvascular COX-2/mPGES-1/EP-4 axis in human abdominal aortic aneurysm.

We investigated the prostaglandin (PG)E2 pathway in human abdominal aortic aneurysm (AAA) and its relationship with hypervascularization. We analyzed samples from patients undergoing AAA repair in comparison with those from healthy multiorgan donors. Patients were stratified according to maximum aortic diameter: low diameter (LD) (<55 mm), moderate diameter (MD) (55-69.9 mm), and high diameter (HD) (≥70 mm). AAA was characterized by abundant microvessels in the media and adventitia with perivascular infiltration of CD45-positive cells. Like endothelial cell markers, cyclooxygenase (COX)-2 and the microsomal isoform of prostaglandin E synthase (mPGES-1) transcripts were increased in AAA (4.4- and 1.4-fold, respectively). Both enzymes were localized in vascular cells and leukocytes, with maximal expression in the LD group, whereas leukocyte markers display a maximum in the MD group, suggesting that the upregulation of COX-2/mPGES-1 precedes maximal leukocyte infiltration. Plasma and in vitro tissue secreted levels of PGE2 metabolites were higher in AAA than in controls (plasma-controls, 19.9 ± 2.2; plasma-AAA, 38.8 ± 5.5 pg/ml; secretion-normal aorta, 16.5 ± 6.4; secretion-AAA, 72.9 ± 6.4 pg/mg; mean ± SEM). E-prostanoid receptor (EP)-2 and EP-4 were overexpressed in AAA, EP-4 being the only EP substantially expressed and colocalized with mPGES-1 in the microvasculature. Additionally, EP-4 mediated PGE2-induced angiogenesis in vitro. We provide new data concerning mPGES-1 expression in human AAA. Our findings suggest the potential relevance of the COX-2/mPGES-1/EP-4 axis in the AAA-associated hypervascularization.

NAD+ Precursors: A Physiological Reboot?

In this Editorial, we comment on a series of recent articles featured in the Special Issue "Emerging Benefits of Vitamin B3 Derivatives on Aging, Health and Disease: From Basic Research to Translational Applications" in Nutrients [...].

Predictive capacity of FAT1 transcriptional expression in patients with head and neck squamous cell carcinomas treated with radiotherapy.

OBJECTIVE: To analyze the predictive capacity at the primary location of the tumor of the FAT1 transcriptional expression in patients with head and neck squamous cell carcinoma treated with radiotherapy. MATERIAL AND METHODS: We conducted a retrospective study from biopsies of the primary location of the tumor in 82 patients with head and neck squamous cell carcinoma treated with radiotherapy. The transcriptional expression of FAT1 was determined by RT-PCR. The level of FAT1 transcriptional expression was categorized according to the local control after radiotherapy using a recursive partitioning analysis. RESULTS: Elevated FAT1 transcriptional expression was associated with an increased risk of local recurrence after radiotherapy. Patients with a high expression level of FAT1 (n=18; 22.0%) had a 5-year local recurrence-free survival of 42.1% (95% CI: 18.6%-65.6%), whereas for patients with a low expression (n=64; 78.0%) it was 72.4% (95% CI: 61.5%-83.3%) (p=0.002). According to the result of a multivariate analysis, patients with a high FAT1 expression category had a 2.3-fold increased risk of local recurrence (95% CI: 1.0-5.2; p=0.043). CONCLUSIONS: Elevated FAT1 transcriptional expression was associated with a significantly increased risk of local recurrence in patients with head and neck squamous cell carcinoma treated with radiotherapy.

Relationship between transcriptional expression of pyruvate dehydrogenase and local control of disease in patients with oral cavity carcinomas.

BACKGROUND: The altered cellular metabolism is one of the hallmarks of the cancer cells, favoring the process of aerobic glycolysis, known as the Warburg effect. The pyruvate dehydrogenase (PDH) complex is one of the elements involved in this metabolic process. The present study aims to evaluate the relationship between the transcriptional expression of PDHB and the risk of local recurrence in patients with oral cavity carcinomas. METHODS: We determined the transcriptional expression of PDHB in biopsies from 41 patients with oral cavity carcinomas treated with surgery. The PDHB expression was categorized according to the local control of the disease with a recursive partitioning analysis. RESULTS: During the follow-up period 13 patients (31.7%) had a local recurrence of the tumor. Considering local disease control as the dependent variable, the recursive partitioning analysis classified the patients in two categories according to high (n=16, 39.0%) or low (n=25, 61.0%) PDHB expression. Five-year local recurrence-free survival for patients with high PDHB expression was 84.8% (95% CI: 65.2-100%), and for patients with low expression it was 54.3% (95% CI: 34.3-74.2 %) (P=0.034). The results of multivariate analysis showed that patients with a low PDHB expression had a 4.90 times higher risk of local recurrence of the tumor (95% CI: 1.02-22.68, P=0.042). CONCLUSION: There is a relationship between the metabolic characteristics of the tumor and its aggressiveness. According to our results, patients with oral cavity carcinomas with low transcriptional expression levels of PDHB have a significantly higher risk of local tumor recurrence.

Properties and cellular uptake of photo-triggered mixed metallosurfactant vesicles intended for controlled CO delivery in gas therapy.

The scientific relevance of carbon monoxide has increased since it was discovered that it is a gasotransmitter involved in several biological processes. This fact stimulated research to find a secure and targeted delivery and lead to the synthesis of CO-releasing molecules. In this paper we present a vesicular CO delivery system triggered by light composed of a synthetized metallosurfactant (TCOL10) with two long carbon chains and a molybdenum-carbonyl complex. We studied the characteristics of mixed TCOL10/phosphatidylcholine metallosomes of different sizes. Vesicles from 80 to 800 nm in diameter are mainly unilamellar, do not disaggregate upon dilution, in the dark are physically and chemically stable at 4 °C for at least one month, and exhibit a lag phase of about 4 days before they show a spontaneous CO release at 37 °C. Internalization of metallosomes by cells was studied as function of the incubation time, and vesicle concentration and size. Results show that large vesicles are more efficiently internalized than the smaller ones in terms of the percentage of cells that show TCOL10 and the amount of drug that they take up. On balance, TCOL10 metallosomes constitute a promising and viable approach for efficient delivery of CO to biological systems.

Electronegative LDL Is Associated with Plaque Vulnerability in Patients with Ischemic Stroke and Carotid Atherosclerosis.

Owing to the high risk of recurrence, identifying indicators of carotid plaque vulnerability in atherothrombotic ischemic stroke is essential. In this study, we aimed to identify modified LDLs and antioxidant enzymes associated with plaque vulnerability in plasma from patients with a recent ischemic stroke and carotid atherosclerosis. Patients underwent an ultrasound, a CT-angiography, and an 18F-FDG PET. A blood sample was obtained from patients (n = 64, 57.8% with stenosis ≥50%) and healthy controls (n = 24). Compared to the controls, patients showed lower levels of total cholesterol, LDL cholesterol, HDL cholesterol, apolipoprotein B (apoB), apoA-I, apoA-II, and apoE, and higher levels of apoJ. Patients showed lower platelet-activating factor acetylhydrolase (PAF-AH) and paraoxonase-1 (PON-1) enzymatic activities in HDL, and higher plasma levels of oxidized LDL (oxLDL) and electronegative LDL (LDL(-)). The only difference between patients with stenosis ≥50% and <50% was the proportion of LDL(-). In a multivariable logistic regression analysis, the levels of LDL(-), but not of oxLDL, were independently associated with the degree of carotid stenosis (OR: 5.40, CI: 1.15-25.44, p < 0.033), the presence of hypoechoic plaque (OR: 7.52, CI: 1.26-44.83, p < 0.027), and of diffuse neovessels (OR: 10.77, CI: 1.21-95.93, p < 0.033), indicating that an increased proportion of LDL(-) is associated with vulnerable atherosclerotic plaque.

Nicotinamide Prevents Diabetic Brain Inflammation via NAD+-Dependent Deacetylation Mechanisms.

This study investigated the effect of nicotinamide (NAM) supplementation on the development of brain inflammation and microglial activation in a mouse model of type 1 diabetes mellitus. C57BL/6J male mice, which were made diabetic with five consecutive, low-dose (55 mg/kg i.p.) streptozotocin (STZ) injections. Diabetic mice were randomly distributed in different experimental groups and challenged to different doses of NAM (untreated, NAM low-dose, LD, 0.1%; NAM high-dose, HD, 0.25%) for 25 days. A control, non-diabetic group of mice was used as a reference. The NAD+ content was increased in the brains of NAM-treated mice compared with untreated diabetic mice (NAM LD: 3-fold; NAM HD: 3-fold, p-value < 0.05). Immunohistochemical staining revealed that markers of inflammation (TNFα: NAM LD: -35%; NAM HD: -46%; p-value < 0.05) and microglial activation (IBA-1: NAM LD: -29%; NAM HD: -50%; p-value < 0.05; BDKRB1: NAM LD: -36%; NAM HD: -37%; p-value < 0.05) in brains from NAM-treated diabetic mice were significantly decreased compared with non-treated T1D mice. This finding was accompanied by a concomitant alleviation of nuclear NFκB (p65) signaling in treated diabetic mice (NFκB (p65): NAM LD: -38%; NAM HD: -53%, p-value < 0.05). Notably, the acetylated form of the nuclear NFκB (p65) was significantly decreased in the brains of NAM-treated, diabetic mice (NAM LD: -48%; NAM HD: -63%, p-value < 0.05) and inversely correlated with NAD+ content (r = -0.50, p-value = 0.03), suggesting increased activity of NAD+-dependent deacetylases in the brains of treated mice. Thus, dietary NAM supplementation in diabetic T1D mice prevented brain inflammation via NAD+-dependent deacetylation mechanisms, suggesting an increased action of sirtuin signaling.

Genome-wide association meta-analysis identifies risk loci for abdominal aortic aneurysm and highlights PCSK9 as a therapeutic target.

Abdominal aortic aneurysm (AAA) is a common disease with substantial heritability. In this study, we performed a genome-wide association meta-analysis from 14 discovery cohorts and uncovered 141 independent associations, including 97 previously unreported loci. A polygenic risk score derived from meta-analysis explained AAA risk beyond clinical risk factors. Genes at AAA risk loci indicate involvement of lipid metabolism, vascular development and remodeling, extracellular matrix dysregulation and inflammation as key mechanisms in AAA pathogenesis. These genes also indicate overlap between the development of AAA and other monogenic aortopathies, particularly via transforming growth factor ß signaling. Motivated by the strong evidence for the role of lipid metabolism in AAA, we used Mendelian randomization to establish the central role of nonhigh-density lipoprotein cholesterol in AAA and identified the opportunity for repurposing of proprotein convertase, subtilisin/kexin-type 9 (PCSK9) inhibitors. This was supported by a study demonstrating that PCSK9 loss of function prevented the development of AAA in a preclinical mouse model.

Interaction between head and neck squamous cell carcinoma cells and fibroblasts in the biosynthesis of PGE2.

Prostaglandin (PG)E(2) is relevant in tumor biology, and interactions between tumor and stroma cells dramatically influence tumor progression. We tested the hypothesis that cross-talk between head and neck squamous cell carcinoma (HNSCC) cells and fibroblasts could substantially enhance PGE(2) biosynthesis. We observed an enhanced production of PGE(2) in cocultures of HNSCC cell lines and fibroblasts, which was consistent with an upregulation of COX-2 and microsomal PGE-synthase-1 (mPGES-1) in fibroblasts. In cultured endothelial cells, medium from fibroblasts treated with tumor cell-conditioned medium induced in vitro angiogenesis, and in tumor cell induced migration and proliferation, these effects were sensitive to PGs inhibition. Proteomic analysis shows that tumor cells released IL-1, and tumor cell-induced COX-2 and mPGES-1 were suppressed by the IL-1-receptor antagonist. IL-1α levels were higher than those of IL-1ß in the tumor cell-conditioning medium and in the secretion from samples obtained from 20 patients with HNSCC. Fractionation of tumor cell-conditioning media indicated that tumor cells secreted mature and unprocessed forms of IL-1. Our results support the concept that tumor-associated fibroblasts are a relevant source of PGE(2) in the tumor mass. Because mPGES-1 seems to be essential for a substantial biosynthesis of PGE(2), these findings also strengthen the concept that mPGES-1 may be \a target for therapeutic intervention in patients with HNSCC.