In vivo, ELDL represents one of the many forms of modified LDLs present in the atherosclerotic vessel. Phenotypic changes of macrophages and SMCs brought about by the uptake of modified LDLs overlap significantly in an atherosclerotic milieu, making it practically impossible to differentiate between the effects from oxidized LDL, ELDL, and other LDL modification. By studying in vitro-generated modifications of LDL, we were able to demonstrate marked differences in the transcriptome of human coronary artery SMCs (HCASMCs) upon uptake of ELDL, OxLDL, and native LDL, indicating that specific modifications of LDL in atherosclerotic plaques may determine the biology and functional consequences in vasculature. Enzyme-modified non-oxidized LDL (ELDL) induces calcification of SMCs and this is associated with reduced mRNA levels for genes protective for calcification  and upregulation of osteoblastic genes. A second focus of this review is on the synergy between hyperlipidemia and accelerated calcification In vivo in a mouse models with transgenic expression of human S100A12. We summarize mechanisms of S100A12/RAGE mediated vascular inflammation promoting vascular and valve calcification in vivo.Arterial calcification is a feature of atherosclerosis and shares many risk factors including diabetes, dyslipidemia, chronic kidney disease, hypertension, and age. Although there is overlap in risk factors, anti-atherosclerotic therapies, including statins, fail to reduce arterial, and aortic valve calcifications. This suggests that low density lipoprotein (LDL) may not be the main driver for aortic valve disease and arterial calcification. This review focuses on modified LDLs and their role in mediating foam cell formation in smooth muscle cells (SMCs), with special emphasis on enzyme modified non-oxidized LDL (ELDL).  Two main types of vascular calcification have been reported in adults: intimal calcification and medial calcification . Intimal calcification occurs in the setting of lipid accumulation, foam cell formation, and macrophage infiltration, and is often present in advanced atherosclerotic plaques. In contrast, medial calcification occurs without intimal plaque formation and localizes to elastin fibers and smooth muscle cells (SMCs); foam cells and infiltrating macrophages are not essential to formation of medial calcification. Medial calcification can and does occur independent of intimal plaque formation in some cases, but the processes are not always separate and medial calcification can be found in patients that also have intimal calcification of plaques. For example, calcification of coronary arteries occurs mainly in the intima and is strongly associated with increased risk of future myocardial infarction. Hence, coronary artery calcification (CAC) on computed tomography is associated with cardiovascular morbidity and mortality . ScreeniIn contrast, thoracic aortic calcification (TAC) preferentially involves the media and provides limited incremental value for cardiovascular risk, and current data do not support screening for TAC . TAC is Another example of medial calcification is the “tram-track sign” that can be seen on routine mammography X ray for breast cancer screening in 60–70% of women>70 years of age or with chronic kidney disease. This represents calcification along the circumference of the internal mammary artery. Calcification of the internal mammary artery is associated with cardiovascular disease with the highest hazard ratio for heart failure (HR 1.52 [1.18–1.98]) and to a lesser degree with coronary artery disease (HR 1.29 [1.01–2.05]) . Th. Th44). in vivo. In cultured SMCs isolated from the aorta of young S100A12 transgenic mice, S100A12 transgene expression alone was not sufficient to induce an osteoblast-like cell transformation and/or spontaneous calcification of SMCs cultured in regular DMEM medium, but gained osteoblast-like features when hyperlipidemic serum from ApoE null mice was added to the cell culture medium. This strongly suggests that S100A12 primes SMCs to switch to osteoblast-like cells in the presence of lipoprotein cholesterol , and Runx-2 in the aorta of S100A12/ApoE null mice supporting a role for S100A12 to induce a phenotypic switch that drives cells to an osteoblast-like cell lesterol .+/+ mice and Figure B depicting a hyperlipidemic ApoE−/− mice. Taken together, these findings suggest that cholesterol lipoproteins have profound effects on SMCs by altering their function with gain in osteoblast-like phenotype. We speculate that the pro-inflammatory milieu mediated by activation of the S100/RAGE axis might modify lipoproteins and render them more atherogenic. Our lab recently reported on mechanisms of transition of SMCs into foam cells with osteoblast-like differentiation in response to enzyme-modified LDLs as outlined below.In summary, transgenically-expressed S100A12 mediates vascular calcification and pathological remodeling of the blood vessel, but this process is dependent on environmental cues with hyperlipidemia being an important modulator of inflammation-driven vascular calcification in this model. This is illustrated by comparing the amount of arterial calcification between Figures The uptake of cholesterol lipoproteins by cells leads to morphological changes visible by light microscopy, and this was termed “foaming,” or “foam cell formation.” Foam cells are a pathological hallmark of atherosclerosis. Since foam cells express several macrophage markers, it has long been thought that foam cells arise primarily from monocyte-derived macrophages. Rong et al. demonstrated in 2003 that SMCs in response to cholesterol loading downregulate expression of typical SMC markers such as smooth muscle actin and upregulate markers commonly seen in macrophages (CD68) . Recent ex vivo, the products could differ significantly, depending on the fatty acid composition and antioxidant status of the starting LDL preparation. The diversity of oxidized LDL particles is reviewed by Levitan et al.  or in combination with organic phosphate beta Glycerophosphate (10–30 mM) did not promote Alizarin Red staining of HCASMC when cultured up to 7 days, however, the addition of inorganic phosphate at low concentrations from 0.5 to 1.5 mM significantly induced mineralization of ELDL-treated cells as early as the second day of incubation Figure . mRNA foOther models support the hypothesis that cholesterol metabolism in SMCs effects profoundly processes relevant to blood vessel remodeling, atherosclerosis and calcification. For example, low-density lipoprotein receptor-related protein 1 (LRP1) protects against cholesterol intracellular accumulation  and LDLRin vitro (Recently, there has been growing interest in understanding the role of sortilin in vascular disease after the initial GWAS studies found the 1p13 locus harboring the SORT 1 gene encoding the protein sortilin to be associated with LDL, myocardial infarction, abdominal aortic aneurysms and CAC –85. Sortin vitro . CollectHyperphosphatemia is associated with vascular calcification and this has been attributed to increases in spontaneous formation of calcium-phosphate crystals, induction of BMP-2 and other osteochodrogenic regulators, and downregulation of alpha smooth muscle actin .Hydroxyapatite is the main calcium phosphate crystal found in bone and calcified tissue and is formed through intermediaries of phosphate and calcium. The high glycine content of extracellular matrix proteins collagen type 1 and elastic fibers favors the binding of calcium non-ionically, via charge neutralization to glycine. This enables phosphate to bind the calcium in a configuration that favors apatite formation. Pyrophosphate is a potent inhibitor of calcium crystallization and deposition. Pyrophosphate has been used industrially for this purpose and is the active ingredient in plaque-preventing toothpaste. Pyrophosphate acts by avidly binding to nascent hydroxyapatite crystals and inhibits hydroxyapatite crystal formation completely at micromolar concentrations.in vitro (in vivo (Extracellular pyrophosphate is a potent physicochemical inhibitor of hydroxyapatite crystal formation in vitro  and in v(in vivo –95. Pyro(in vivo . Lack of(in vivo .ENPP1 was shown to be causal for infantile arterial calcification, and infants born with this recessive disease develop arterial calcification in the first months of life (ENPP1 gene had been previously linked to ectopic calcification through a mouse model known as the “tiptoe walking mouse” or tww (Pyrophosphate deficiency is the basis for vascular calcification in several genetic disorders , 99. ENP of life –100. Mor” or tww .Villa Bellosta et al. have shown that pyrophosphate synthesis increases 10 times more than pyrophosphate degradation during phosphate-induced calcification , 102. HeENPP1 and its role in ectopic and vascular calcification is likely to expand, studies implicate RAGE, the receptor for advanced glycation end products, as one pathway through which ENPP1 may act (ENPP1 null mice in combination with loss of RAGE developed less arterial calcification and intra-arterial chondrogenic differentiation compared to ENPP1 null mice with intact RAGE signaling, indicating an important role for oxidative stress and chronic inflammation as a modifier of phosphate-mediated calcification (ENPP1 gene expression is intriguing and suggests a possible key mechanism of ELDL-mediated calcification of SMCs.While our understanding of the biology of  may act . ENPP1 nfication . Our finin vitro, also reduces atherosclerosis and vascular calcification in ApoE null mice with transgenic S100A12 expression, suggesting that that S100A12 is a potential pharmacological target (There is substantial evidence for RAGE as a mediator of vascular disease , and impl target , 35. S10l target , and mayl target . Howeverin vitro-generated ELDL supports the hypothesis that ELDL could be a critical mechanistic link between chronic inflammation and accelerated vascular calcification.While there is no doubt of the importance of pyrophosphate as an physiologic inhibitor of vascular calcification , strategIn summary, vascular calcification is complex and is driven by genetic as well as environmental exposures. While our understanding of this complex disorder has significantly progressed over the last two decades, much work remains in order to clarify mechanisms, particularly given that vascular calcification varies phenotypically, and may actually represent variations on a pathobiological theme. Prevention of calcification would be ideal, but reversing or limiting vascular calcification, once initiated, is another key strategy to mitigate the morbidity and mortality that occur as a result of vascular calcification. Given our current understanding of vascular calcification, strategies of prevention are more likely to translate into clinical practice than the more complex task of reversing vascular calcification. Therefore, any visualization of subclinical vascular calcification, either on a screening mammogram or on computed tomography should be seen as an opportunity to investigate risk factors and initiate preventive strategies. So perhaps the greatest power in screening mammography lies in the potential to prevent breast cancer and coronary artery disease from developing in the first place. Breast cancer and vascular calcification share several modifiable risk factors: diabetes mellitus, smoking, physical inactivity, poor diet, and obesity. If women were counseled each time they had a mammogram that controlling their risk factors could help them avoid two of the deadliest conditions for women, then the message of prevention could gain considerable strength .All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Non-coding RNA species contribute more than 90% of all transcripts and have gained increasing attention in the last decade. One of the most recent members of this group are long non-coding RNAs (lncRNAs) which are characterized by a length of more than 200 nucleotides and a lack of coding potential. However, in contrast to this simple definition, lncRNAs are heterogenous regarding their molecular function—including the modulation of small RNA and protein function, guidance of epigenetic modifications and a role as enhancer RNAs. Furthermore, they show a highly tissue-specific expression pattern. These aspects already point towards an important role in cellular biology and imply lncRNAs as players in development, health and disease. This view has been confirmed by numerous publications from different fields in the last years and has raised the question as to whether lncRNAs may be future therapeutic targets in human disease. Here, we provide a concise overview of the current knowledge on lncRNAs in both glomerular and tubulointerstitial kidney disease. Most of the human genome is actively transcribed but less than 2% contains protein coding transcripts (mRNA). The other transcripts produced show no or low coding potential and have, therefore, been summarized in the large group of non-coding RNAs (ncRNA). This group contains the long known ribosomal RNAs (rRNAs) and transfer RNAs (tRNAs) involved in protein synthesis as well as two very diverse classes of ncRNAs mainly divided by their length: the class of small non-coding RNAs containing transcripts with lengths of less than 200 nucleotides consists of small interfering RNAs (siRNAs), small nuclear RNAs (snRNA), small nucleolar RNAs (snoRNAs), PIWI-interacting RNAs (piRNAs) and microRNAs (miRNAs). The role of miRNAs in kidney disease has been investigated extensively and reviewed multiple times over the last decades ,2, thereDue to their generally low expression levels the vast majority of the 215,008 annotated human lncRNAs  was only discovered within the last decade . Whilst As to glomerular disease by far most publications have analyzed the role of lncRNAs in diabetic nephropathy . ConsequPVT1 locus [With the incidence of diabetes mellitus rising and diabetic nephropathy (DN) being a leading cause of end-stage renal disease in Western societies ,12, it iT1 locus  all of wT1 locus . InhibitRegarding the involvement of specific lncRNAs we will focus on genes that have been implicated in DN by evidence from several publications.PVT1 had been linked to diabetic nephropathy by the finding that variants in this gene are associated with the development of end-stage renal disease (ESRD) in both type 1 and 2 diabetes mellitus [The plasmacytoma variant translocation gene mellitus ,21. Soonmellitus , providimellitus . Zhang emellitus .MALAT1 is another important example of an lncRNA involved in DN. MALAT1 is induced in the streptozocin-induced diabetic nephropathy mouse model . Based oMitochondrial dysfunction is one of the hallmarks in DN . The modBesides, several lines of evidence point towards a functionally important involvement of the lncRNA NEAT1 in diabetic nephropathy. NEAT1 is induced in a streptozocin-mediated diabetic rat model and murine mesangial cells treated with high-glucose ,31. IncrCYP4B1-PS1-001 was first described in diabetic nephropathy in a microarray based-screen for dysregulated lncRNAs in the db/db mouse model . Whilst More than 20 additional lncRNAs have been implicated in diabetic nephropathy by single publications. Here, we summarize the most important findings. Several of these lncRNAs interact directly with miRNAs and inhibit their function. As examples, LINC01619 induces oxidative podocyte damage by serving as a sponge for miR-27a , Gm6135 In comparison to DN, little is known about the contribution of lncRNAs to other glomerular disease entitites. A Pubmed search for lncRNAs AND glomerulus revealed more than 80% of these publications to deal with DN. Fewer than 10 studies report results regarding different types of glomerulonephritis and focal-segmental glomerulosclerosis (FSGS). Due to the scarcity of data, all of these studies are discussed in the following paragraph independent from impact and approach. LncRNA LOC105375913 was found to be increased in tubular cells of 5 FSGS patients  and thisTo date, the majority of lncRNA studies on tubulointerstitial kidney disease were performed in the context of acute kidney injury (AKI) . ResearcAKI is a central problem in nephrology considering the high and increasing incidence associated with the demographic changes in Western societies and the lack of specific preventive and therapeutic strategies . ConsequMost evidence for an lncRNA to play a role in AKI has been presented for metastasis-associated lung adenocarcinoma transcript 1 (MALAT1). Initially identified as the most highly induced lncRNA gene in kidney and testis of hypoxic mice, MALAT1 was proposed to be hypoxia-inducible factor (HIF)-2 activated and postulated to function in renal proximal tubuli . MALAT1 Nuclear enriched abundant transcript 1 (NEAT1) was detected as significantly upregulated in the serum of patients with sepsis-induced as well as in patients with ischemia-induced AKI ,72. UsinTUG1) [MEG3) [TapSAKI) [Similar studies link the lncRNAs HOX transcript antisense RNA (HOTAIR) , taurineTUG1) , materna) [MEG3)  and tranTapSAKI)  to a ranHOTAIR—known to play an important role in apoptosis—was upregulated in rats with sepsis caused by urinary tract infection . The autTo unravel the role of the lncRNA TUG1, detected at lower levels in the serum of patients suffering from sepsis-associated AKI in comparison to healthy controls, a LPS based in vitro model was used . DespiteIn two individual studies MEG3 was found to be upregulated after LPS or hypoxia treatment, respectively and in both cases the authors report the sequestration of miRNAs by MEG3 ,63. YangTapSAKI, initially discovered as a circulating lncRNA upregulated in the plasma of AKI patients and their kidney tissue was proposed as a biomarker with predictive value for the survival of AKI patients . RecentlIn the section below we discuss lncRNAs which so far have only a single study connecting them to acute kidney injury—plasmacytoma variant translocation 1 (PVT1) , psoriasAn involvement of PVT1 in kidney disease has been described previously for diabetic nephropathy  and is dThe expression and secretion of RANTES/CCL5  is known to recruit circulating leukocytes to sites of injury and to reinforce inflammatory reactions. In their report, Yu et al. showed that PRINS, a HIF-1α regulated lncRNA is potentially involved in RANTES production . The stuThe hypoxia-responsive lncRNA GAS5 was reported to be upregulated in IRI treated mice by Geng et al. . The incThe study conducted on HK-2 and primary renal proximal tubular epithelial cells (RPTEC) by Mimura et al. compared lncRNA expression patterns between hypoxic and normoxic cells . DARS-ASTian et al. observed an upregulation of LINC00520 in rat kidney tissue following IRI treatment . Using HUC.173 belongs to a group of lncRNAs transcribed from an ultra-conserved region (T-UCR) shared between human, rat and mouse genomes and was initially reported as downregulated in lead-exposed human populations and animal models . Qin et In this section we give an overview of recent advances with regard to the function of lncRNAs in tubulointerstitial diseases other than acute kidney injury. We discuss the role of lncRNAs in 5/6 nephrectomy, autosomal dominant polycystic kidney disease (ADPKD) and kidney injury by crystal-formation .To date, two lncRNAs have been associated with the transcriptional response in rat models of 5/6 nephrectomy ,79. LINCPkd1 and Pkd2 knockout mice, a genetic model of ADPKD, a downregulation of the mouse ortholog was observed. In a cell model using mIMCD3, the authors showed that knockout of Hoxb3os leads to mTORC1 activation and increase in mitochondrial respiration and that re-expression of Hoxb3os rescues this phenotype.To the best of our knowledge, so far only one lncRNA has been shown to have functional implications in ADPKD. Aboudehen et al. investigated the role of Hoxb3os, an lncRNA abundantly expressed in the kidney and evolutionary conserved with the human ortholog HOXB-AS1 . The expRecent findings have also shown that lncRNAs are associated with renal injury mediated by crystal formation. The studies - mainly based on HK-2 cell exposure to calcium oxalate monohydrate (COM), the major constituent of kidney stones , range fRegarding glomerular disease there are no reports on circulating lncRNAs as potential biomarkers. For membranous nephropathy lncRNA XIST has been hypothesized to be a potential urinary biomarker . HoweverLncRNAs provide a fascinating new layer to pathophysiological studies and the search for novel therapeutic strategies in kidney disease. However, apart from few examples such as diabetic nephropathy and acute kidney injury, little is known about their impact in renal pathologies and most studies have remained primarily descriptive. Future work will need to close this gap in order to increase our understanding of this new class of non-coding RNAs in nephrology and to eventually elucidate how we can exploit the potential of lncRNA modulation for patients suffering from kidney disease. The inhibition of lncRNAs in vivo is technically no more challenging than the inhibition of messenger RNAs; therefore, with an increasing number of antisense oligonucleotide therapies coming into clinical use at the moment  this concept could be of high interest for targeting lncRNAs in kidney disease ,89,90. R


Crimean-Congo hemorrhagic fever virus (CCHFV) is a widely distributed hemorrhagic fever virus and the cause of hemorrhagic disease in Africa, Southern and Eastern Europe, the Middle East, India and Asia. Recent emergence of CCHFV into Spain indicates that the geographic range of this virus is expanding and the presence of its tick vector in several countries without reported disease suggest that CCHFV will continue to spread. Research into CCHFV was historically limited by a lack of suitable animal models and tools to study viral pathogenesis. However, in the past few years the toolset for studying CCHFV has expanded with small animal and non-human primate models for CCHFV being developed along with a reverse genetics system that allows for investigation of viral determinants of disease. These tools have been utilized to understand how CCHFV antagonizes host restriction factors and to develop novel vaccine candidates that may help limit the substantial morbidity and mortality in humans caused by CCHFV. Nairoviridae family within theBunyavirales order of viruses. CCHFV contains three genomic segments: small and medium, which encode for the nucleoprotein and glycoproteins, respectively, and a large segment encoding the RNA-dependent RNA-polymerase. CCHF as a disease was first described in humans in the 1940s when soldiers re-occupying abandoned farmland in the Crimea became ill with a hemorrhagic disease1. In the late 1960s, it was discovered that the causative agent of this hemorrhagic disease in the Crimea was similar to the causative agent of hemorrhagic disease in the Belgian Congo (current Democratic Republic of the Congo)2, and the name “Crimean–Congo hemorrhagic fever virus” was ascribed to the pathogen. The main vector and reservoir of CCHFV are hard-body ticks principally of theHyalomma genus, although there is limited evidence that other species of ticks such asRhipicephalus andDermacentor species may be vectors3. Vertebrate hosts such as domestic livestock and wild animals such as hares likely serve as amplifying hosts of CCHFV, with uninfected ticks becoming infected during feeding on viremic animals or during co-feeding with infected ticks6  is a negative-sense RNA virus in theticks6 . TheHya8  and aspartate aminotransferase (AST) are common predictors of poor outcome14  during the viremic phase of disease. RT–qPCR can also determine viral load, which often is correlated with disease outcome. An important consideration for these assays is the substantial genetic diversity of CCHFV; however, assays that can recognize a multitude of CCHFV genotypes have been developed5, yet humans seem to be uniquely susceptible to severe or lethal disease. Even within humans, CCHFV infection may result in a mild or subclinical infection9, and why some patients develop a serious or fatal disease whereas others develop an asymptomatic infection is unknown.Curiously, extensive serological evidence indicates that CCHFV can productively infect numerous vertebrate species, including both wild and domestic animals25 suggests that innate immune responses in vertebrate hosts play a substantial role in limiting CCHFV pathogenesis.In vitro studies have identified retinoic acid-inducible gene I (RIG-I) as an innate immune sensor of CCHFV26, although CCHFV may process its viral RNA to avoid RIG-I sensing27. However, it is likely that more innate immune sensors contribute to sensing of CCHFV. Retrospective studies in humans have identified correlates between polymorphisms in Toll-like receptors (TLRs) and severity of disease, suggesting that TLRs may be an important immune-sensing pathway in the control of CCHFV. Polymorphisms in TLR7, 8, 9, and 10 have been found to be correlated with disease severity and outcome in infected patients in Turkey30. Polymorphisms in nuclear factor-kappa B have also been found to correlate with an increased risk of CCHF31. Whether similar polymorphisms are associated with CCHF and case severity in populations in other geographic regions remains to be determined. Host apoptotic pathways may restrict CCHFV replication, as CCHFV replication can induce apoptosis33, and caspase 3, activated during apoptosis, can cleave the CCHFV nucleoprotein and inhibit viral replication35.The ability of CCHFV to cause severe or lethal disease in mice deficient in the type I interferon system but not wild-type (WT) mice36. The CCHFV OTU domain can also cleave interferon-stimulated gene 15 (ISG15) modifications38. As ISG15 modifications have been implicated in direct antiviral effects and as ISG15 itself has been found to regulate the host innate immune response to viral infection39, the de-ISGylation activity of the CCHFV OTU domain may be important for viral pathogenesis. Recombinant CCHFV encoding an OTU deficient in deubiquitinase activity or deficient in both deubiquitinase and de-ISGylation activity had reduced viral growth kinetics compared with WT CCHFV in interferon-competent but not interferon-incompetent cells, demonstrating that the OTU domain is important in overcoming interferon responses36. Intriguingly, the CCHFV OTU domain has reduced affinity for mouse ISG15 compared with human ISG15, suggesting that mouse ISG15 may be an important barrier to CCHFV infection of mice41. The CCHFV OTU domain also has higher affinity for human ISG15 than the OTU domains of relatedNairoviridae members that are not associated with disease as severe as CCHF, suggesting that the OTU domain may be an important virulence factor for CCHFV40.CCHFV also antagonizes innate immune signaling. The L segment of CCHFV encodes an ovarian tumor-like deubiquitinase (OTU) domain that recently has been shown to suppress innate immune responses likely by deubiquitinating proteins involved in innate immune signaling pathways14, and levels of antibody may serve as a predictor of disease outcome. However, whether antibody responses contribute to the control of primary CCHFV infection is unknown, and neutralizing antibody responses even in survivors are typically low13. Epitope mapping using linear peptides and serum from CCHFV survivors in Turkey and South Africa identified antibody responses toward epitopes in the mucin-like domain, GP38, and Gn protein42 regions that are not likely to result in neutralizing antibodies43. However, non-neutralizing antibodies were protective against lethal CCHFV challenge43, demonstrating that antibodies can be protective via mechanisms other than neutralization. Studies evaluating a modified vaccinia virus-based vaccine for CCHFV found protection to require both cellular and humoral responses, suggesting that T cells may be an important component of vaccine-mediated protection against CCHFV45. The role for T cells in controlling primary CCHFV infection is unclear. Levels of circulating CD3+CD8+ T cells in peripheral blood were found to positively correlate with fatal outcome46, and human CCHF survivors have been shown to exhibit long-lived CD8+ T-cell responses to CCHFV47. Human leukocyte antigen alleles have also been found to correlate with protection and susceptibility to CCHFV48. A humanized mouse model of CCHFV demonstrated that T cells were activated following CCHFV infection and CD8+ T cells had elevated levels of perforin, a marker of cytolytic activity49. However, further studies are needed to understand how T-cell responses contribute to resolution of the infection.The role for adaptive immune responses against CCHFV in human pathogenesis is less clear. Low-to-absent anti-CCHFV antibody responses have been found to correlate with severe disease and death+ and CD8+ T cells, although these cells were depleted from the spleen by day 3 post-infection (PI) and mice succumbed prior to detectable antibody responses25. In a recent study using mice treated with an interferon blockade antibody, mice deficient in adaptive immune responses, but not WT mice, supported persistent viral replication for at least 2 weeks PI, demonstrating that adaptive immune responses can control CCHFV in mice50. This model also demonstrated that adaptive immune responses, such as cytolytic T-cell activity, were not necessary for hepatic injury following CCHFV infection50. These data suggest that CCHFV is directly capable of causing liver damage independent of the host adaptive immune response’s attempts to control viral replication. Furthermore, the recent development of a cynomolgus macaque model of CCHF51 may provide insight into the role of the adaptive immune response and disease outcome. In this model, neither antibody titers nor neutralizing activity of the antibodies correlated with disease outcome51. However, studies evaluating the contribution of the T-cell response to disease outcome in this model are still needed.Understanding the role of the adaptive immune response in the control of primary CCHFV infection has been limited by the lack of suitable animal models. Type I interferon-deficient mice exhibit a rapid-onset lethal disease prior to the development of adaptive immune responses, limiting the suitability of this model for studying adaptive immune responses. However, studies in STAT1-deficient mice demonstrated early activation of CD452. Currently, the standard mouse model uses mice deficient in type I or both type I and type II interferon responses25 , and several vaccines have shown promise in pre-clinical trials. A modified vaccinia virus expressing the glycoproteins of CCHFV was shown to provide 100% protection to lethally challenged mice44. DNA-based vaccination or virus-like particle vaccination has also been shown to confer protection against lethal CCHFV challenge63. Mice fed transgenic plants expressing the CCHFV glycoproteins developed antibodies against the glycoproteins64, although the efficacy of this platform against CCHFV challenge was not tested. Interestingly, although subunit vaccines containing the glycoproteins are immunogenic in mice66, they may not induce protective immune responses to CCHFV challenge65. A formalin-inactivated preparation of CCHFV was also found to be protective against CCHFV infection in mice55.Currently, the only vaccine for CCHFV is an inactivated preparation of virus grown in neonatal mouse brains68 or in combination platforms that include both the nucleoprotein and the glycoproteins63. Notably, vaccination with an adenovirus expressing the nucleoprotein of CCHFV provided substantial protection against lethal CCHFV challenge in mice67, demonstrating that immune responses directed against the nucleoprotein can be protective independently of responses to the glycoproteins. However, a modified vaccinia virus expressing the nucleoprotein of CCHFV, though immunogenic, failed to protect against lethal CCHFV challenge68, suggesting that protection afforded by nucleoprotein-based vaccines may be incomplete. Nevertheless, the greater sequence conservation of the nucleoprotein, even among divergent CCHFV strains, suggests that the nucleoprotein may be worth including in vaccine preparations to generate broadly protective immune responses.In addition to the glycoproteins, the nucleoprotein of CCHFV, encoded by the S segment, has been targeted by vaccines either on its own71. Notably, a placebo-controlled study failed to identify a clinical benefit of ribavirin treatment in patients with CCHF72. A recent meta-analysis demonstrated that ribavirin treatment needs to be started soon after symptom onset (<48 hours) to reduce odds of death73. In the recent emergence of CCHFV in Spain, although treatment of an infected nurse with ribavirin had mutagenic effects on CCHFVin vivo and a reduction in viral titers was coincident with treatment start74, ultimately ribavirin treatment was discontinued because of suspected hemolytic anemia75, a potential complication of ribavirin treatment77. The inconsistent data on the clinical benefit of ribavirin for the treatment of CCHFV and the potential for adverse events with ribavirin treatment have caused significant debate in the field81. Ethical considerations of placebo-controlled studies will likely make further studies of this type difficult82, preventing definitive conclusions on the efficacy of ribavirin in patients with CCHF.In addition to vaccines, investigations of antivirals against CCHFV have been conducted. Ribavirin, a nucleoside analog, is suggested by the World Health Organization for the treatment of CCHFV. However, clinical data supporting the use of ribavirin to treat CCHF are inconsistent; some studies report benefits whereas others report no benefit, and meta-analyses of multiple studies suggest that the efficacy of ribavirin is poor or inconclusive54. However, another study showed that ribavirin could protect against lethal disease following CCHFV strain 10200 infection when administered early and that protection diminished when treatment was delayed or challenge dose was increased25. However, results from our lab showed that although early ribavirin treatment extended the mean time to death, ribavirin was unable to prevent death in 10200-infected mice54. The reason for the distinct outcomes in ribavirin-treated strain 10200-infected mice seen between our study54 and that by Benteet al.25 is unknown but could be due to differences in mouse strain (IFNAR−/− versus STAT1−/−), challenge dose, or the time treatment was started after infection. Cumulatively, data in humans and mice suggest that while ribavirin may have limited clinical benefit in patients with CCHF, treatment likely needs to be started early in the course of disease to have clinical benefit. This may prove difficult, as the early symptoms of CCHF are non-specific and can progress rapidly to severe, hemorrhagic manifestations8 and therefore patients may not present to health-care providers until exhibiting the more serious symptoms of CCHF.Studies in mouse models have also shown inconsistent efficacy of ribavirin. Two studies have shown that even early treatment with ribavirin (<6 hours PI) was unable to prevent lethal disease following infection with two distinct clinical isolates of CCHFV83 but has shown promise against other highly pathogenic RNA viruses, including Ebola84 and Lassa86. Two studies have evaluated favipiravir against CCHFVin vivo. In a study by Oestereichet al., favipiravir treatment was effective in suppressing viral replication and preventing mortality following CCHFV infection, even when treatment was started 48 hours PI53. Similarly, work by our group has shown that favipiravir treatment could be delayed until 6 days PI, a time point at which mice were exhibiting advanced disease, including death, and still offer significant clinical benefit to CCHFV-infected mice54. These data suggest that favipiravir may be an effective antiviral for the treatment of advanced CCHF. Furthermore, Oestereichet al. demonstrated that favipiravir and ribavirin could synergistically inhibit CCHFVin vitro, allowing lower doses of both drugs to be usedin vivo with clinical efficacy, suggesting that combination therapies in humans may be effective in treating CCHF while reducing unwanted side effects53. A similar approach has been used in Lassa fever cases87. In addition, a high-throughput screen using recombinant CCHFV identified a compound, 2′-deoxy-2′-fluorocytidine, with inhibitory activity superior to that of favipiravir or ribavirinin vitro88.In vivo animal studies will be needed to evaluate how this compound performs in animal models of CCHF. Lastly, monoclonal antibodies have shown efficacy against CCHFVin vivo43, and several clones were shown to neutralize divergent CCHFV strains89, suggesting that they may have promise for the treatment of CCHF.Favipiravir is approved in Japan for the treatment of influenza virus infectionsIn conclusion, much remains to be understood about the pathogenesis of CCHFV. However, the toolset for studying CCHFV has been steadily improving in recent years with the development of mouse and non-human primate models to a reverse genetics system for CCHFV that will facilitate dissection of the host and viral determinants of CCHFV pathogenesis. These tools will also allow the development and evaluation of novel therapies that reduce or prevent CCHFV-induced morbidity and mortality. Hopefully, the collaboration of multiple institutions in countries around the world toward the development of vaccines against CCHFV will lead to safe and effective vaccines for CCHFV being deployed in populations at risk for CCHFV infection. Lastly, the role of the tick vector in transmission and pathogenesis needs more attention, as does the role of livestock and other animal species in maintaining and transmitting CCHFV. This could lead to more effective measures to block CCHFV transmission.


The conferences focused on the complex interplay of factors that contribute to cancer susceptibility, development, and resistance, providing an excellent discussion forum on the key challenges and the latest advancements that mostly promise to propel this field forward, hosting high-profile speakers.The Medical Research Council, Toxicology Unit and CDDAshok Venkitaram  showed that toxic agents such as formaldehyde and acetaldehyde, pervasively found in the environment but also endogenously accumulated in certain tissues as byproducts of cellular metabolism, are able to lower the abundance of BRCA2. As a consequence, these agents induce haploinsufficiency, particularly in individuals carrying BRCA2 heterozygous mutations, in which the threshold of a functional protein is lower than in wt carriers, thereby promoting spontaneous mutations and tumorigenesis. This explains why BRCA2 loss affects specific tissues, setting the bases for possible chemopreventive strategies in mutation carriers, and impacts on public health and safety issues alerting on the detrimental effects of widespread environmental and dietary aldehydes.Vishva Dixit , who was awarded the CDD Juerg Tschopp Prize 2018, discussed a more recently identified cancer predisposing gene, the BAP1 tumor suppressor. BAP1 germline mutation predisposes carriers to the development of mesothelioma, uveal, and cutaneous melanoma and various other tumors, a condition known as BAP1 cancer syndrome. BAP1 depletion, however, triggers cell death in many cell types, which could seem paradoxical considering its tumor suppressor function. In his engaging Keynote Lecture Dixit clued in on this seeming paradox showing that BAP1, which encodes a deubiquitinase, functions in concert with ring finger protein 2 (RNF2) to fine tune the activity of histone H2A. While RNF2, through H2A monoubiquitination, silences the BCL2 and MCL1 prosurvival genes and thereby induces apoptosis in some cell types, this does not occur in melanocytes, suggesting that BAP1 promotes tumorigenesis in cells that do not engage such RNF2 apoptotic programme.Karen Vousden , who was awarded the CDD Award 2018, retraced the long road leading to her key discoveries of p53 functions. She highlighted how the identification of the p53 target gene TIGAR led to define p53 role in regulating cell metabolism, showing that p53 can also protect cells rather than killing them upon stress conditions. Vousden showed how the ability of p53 to rewire cell metabolism, limiting ROS and providing nutrient sources, can be detrimental favoring cancer cell survival. Indeed, some tumor derived p53 mutants selectively retain these functions. She laid the ground for the development of novel tailored nutritional approaches, which might aid conventional cancer treatments.Moving to the p53 tumor suppressor, Ivano Amelio  showed that p53 mutants can interact with the hypoxia inducible transcription factor HIF1 promoting the expression of a gene set that generates a pro-tumorigenic extracellular microenvironment in lung cancer.Similarly, Xin Lu  showed how p53 transcriptional activity can be modulated by the direct binding of iASPP, which affects specific p53 targets. Interestingly iASPP, mostly known as an oncogene able to inhibit the p53 apoptotic function, was also found to act as a tumor suppressor in certain microenvironmental contexts.Carol Prives  received the 2019 CDD Award presenting her work on the p53 control of the mevalonate pathway via the sterol regulatory element-binding protein 2, which represents an early event for hepatocellular carcinoma (HCC) development. She also discussed the role of MDM2 in promoting ferroptosis independently of p53, likely through PPARα.Finally, Michael Karin  presented a novel substrate of caspase 2 promoting steatosis and nonalcoholic steatohepatitis progression, identifying in caspase 2 a new target to prevent and treat these diseases.Still on HCC, Tak Mak  provided an insightful and compelling argument about the current state of cancer therapy, summarizing collective translational efforts and the lesson learned so far from common experience. He showed how crucial is the activation of T cells to mount an immune response against cancer cells and how this can be boosted with immunotherapy approaches based on immunecheckpoint inhibitors ; he finally discussed his most recent work on choline acetyltransferase-expressing T cells that are required to control chronic viral infection.Scott W. Lowe : a rational combination of drugs can be used to counteract lung cancer by inducing cell senescence and, in particular, a senescence associated secretory phenotype that is able to invoke the attack of NK cells within the microenvironment stimulating the antitumoral immune surveillance.A new approach based on the stimulation of the immune system was proposed by Doug Green  showed how targeting autophagy can be used as another strategy to enhance anticancer immunity. LC3-associated phagocytosis in the myeloid compartment can tackle tumor growth via the upregulation of the cGAS-STING pathway.Yufang Shi  discussed the role of mesenchymal stromal/stem cells (MSCs) in cancer. MSCs have the ability to modulate the microenvironment and control tumor growth and metastasis orchestrating both innate and adaptive, local, and systemic immune responses.Ying Wang  presented recently identified data on how macrophages, key players of both the innate and adaptive immune responses, can be trained to become anti-inflammatory. She showed in particular that IGF2 can program macrophages during their maturation determining a metabolic commitment for OXPHOS, which shapes their anti-inflammatory competences. Adoptive transfer of such reprogrammed macrophages functioned in a mouse model of autoimmune disease and promises to be relevant in other inflammatory conditions.Charles Swanton  presented his studies on cancer evolution discussing the importance of mutations within tumours and how the targeting of trunk mutations that are communal to the main bulk of the tumour cells originate the subsequent heterogeneity in the resistant cancer cells. Intratumoral heterogeneity and genomic instability are finely tuned allowing cancer cells to evolve during the disease course evading immune surveillance and resisting to therapies. Targeting such heterogeneity and new clonal antigenic architecture might help to reduce the treatment failure associated with targeted monotherapies.Paul Sorenson : cancer cells, upon stress cues, initiate a translational reprogramming at the mRNA level that inhibits overall translation activity to preserve energy and nutrients and induces at the same time the expression of major stress adaptor proteins  that allow them not only to survive but also to evolve to more aggressive phenotypes.Another key mechanism allowing tumor cells to adapt to environmental stress factors such as hypoxia, starvation, oxidative, or genotoxic stress was discussed by Pierre Close  proposed a new strategy to counteract melanoma cell survival and resistance to therapy by acting on the protein synthesis rewiring mediated by wobble tRNA modification enzymes, which are required by cancer cells for specific decoding during mRNA translation.Conversely, Xiadong Wang  was awarded the CDD Juerg Tschopp Prize 2019. He introduced the audience to a new mechanism leading to apoptosis execution. He showed that estrogen and its related steroid hormones at the high concentration reached in the developing placenta, induce apoptosis by binding to phosphodiesterase 3A (PDE3A), which recruits and stabilizes the Schaflen 12 (SLFN12) protein increasing its levels. SLFN12 then binds to ribosomes impairing the engagement of signal recognition particles thereby blocking the translation of antiapoptotic BCL2 and MCL1, the decrease of which triggers apoptosis. This process guarantees the dynamic cell turnover occurring in the development of placenta that is required for a successful implantation.For his pioneering studies defining key components of cell death and necroptotic pathways, Vishva Dixit presenting data from a catalytic dead caspase 8 knock-in mouse, which revealed an unexpected cross talk between apoptosis, necroptosis, and other death pathways; Andrew Oberst  showing a recent strategy devised to induce necroptosis in cancer cells acting on the RIP kinases to achieve tumor clearance through the induction of antitumor immunity unleashed by dying cells; Peter Vandenabeele  reporting a thorough comparative analysis of immunogenic cell death triggered by necroptosis, apoptosis, and ferroptosis; Pascal Meier  showing how the ubiquitin signaling system impacts on TNF-induced cell death regulating RIPK1 activity and how such modulation could be used to limit inflammation in clinical applications based on TNF administration.Cell death pathways were discussed further with other engaging talks including: Greg Winter  provided a delightful end to the 2019 meeting describing his successful approach to generate a diversity of antibodies through the phage display method inspired by the principle of genetic change and selection to evolve new proteins. Spanning from the first humanized antibodies to the latest bicycles peptides, he described all the steps that have led so far to the development of a wide range of successful therapeutics, highlighting pros and cons of each class as well as challenges for future developments.Finally, the 2018 Nobel Laureate Despite the intense schedule of inspiring talks, the meetings provided many opportunities for discussions and interactions with leading scientists from around the globe in a very pleasant atmosphere. During the conferences the Editorial Boards of the three Cell Death journals gathered to discuss editorial strategies along with Springer Nature representatives who showed the performance of the various journals. Gerry Melino, founder and chief editor of CDD, now in its 25th anniversary, encouraged all editors to keep working to the highest publishing standards providing a high quality and timely service to the whole scientific community to which the journals belong.


Innate immunity activates the corresponding immune response relying on multiple pattern recognition receptors (PRRs) that includes pattern recognition receptors (PRRs), like NOD-like receptors (NLRs), RIG-I-like receptors (RLRs), and C-type lectin receptors (CLRs), which could accurately recognize invasive pathogens. In particular, NLRs belong to a large protein family of pattern recognition receptors in the cytoplasm, where they are highly correlated with activation of inflammatory response system followed by rapid clearance of invasive pathogens. Among the NLRs family, NLRC5, also known as NOD4 or NOD27, accounts for a large proportion and involves in immune responses far and wide. Notably, in the above response case of inflammation, the expression of NLRC5 remarkably increased in immune cells and immune-related tissues. However, the evidence for higher expression of NLRC5 in immune disease still remains controversial. It is noted that the growing evidence further accounts for the participation of NLRC5 in the innate immune response and inflammatory diseases. Moreover, NLRC5 has also been confirmed to exert a critical role in the control of regulatory diverse signaling pathways. Together with its broad participation in the occurrence and development of immune diseases, NLRC5 can be consequently treated as a potential therapeutic target. Nevertheless, the paucity of absolute understanding of intrinsic characteristics and underlying mechanisms of NLRC5 still make it hard to develop targeting drugs. Therefore, current summary about NLRC5 information is indispensable. Herein, current knowledge of NLRC5 is summarized, and research advances in terms of NLRC5 in characteristics, biological function, and regulatory mechanisms are reviewed. The innate immune system has always been a striker against pathogenic microorganisms such as bacteria, fungi, and viruses . Innate At present, there are 23 NLR genes in the human genome and at least 34 NLR genes in the mouse genome that are expressed widely in various cells and tissues . It is cThere is growing evidence that members of the NLRs family play a key role in antiviral responses. In fact, NLRC5 was demonstrated to be the last prosecutor of NLRs in the presence of virus invasion . It was NLRs could recognize intracellular pathogens and promote the expression of pro-inflammatory cytokines. A great number of studies have suggested that NLRC5 is widely expressed in immune tissues, lymphocytes, and macrophages/monocytes . FurtherIt is well known that genetic background has an effect on gene expression . The NLRNLRC5 is widely expressed in the liver tissues and has recently been recognized as an active regulator of NF-κB signaling pathways . It has Pattern recognition receptors (PRRs) are innate immunity sensors that performed a pivotal role in the modulation of tumor immunity. They could identify danger signals, clean the profitless cells, and balance the quantity of the host flora to thereby modulate the tumor immunity. In addition, PPRs were also found to facilitate tumorigenesis by reducing the population of regulatory cell and inducing the immunosuppressive cytokines . A lot oAccumulating evidence showed that persistent inflammation leads to liver injury and continuous activation of hepatic stellate cells (HSC) will make liver injury worsen. It is generally known that HSC can be activated and transformed into myofibroblast-like cells. Damage to hepatocytes and Kupffer are commonly acknowledged as the first-line effects during the activation of quiescent HSC . It was A large number of studies have shown that hepatic steatosis is a key event in the process of liver injury induced by EtOH. Alcoholic steatosis is one of the major causes of alcoholic liver disease . AlcoholIschemia reperfusion injury (IRI) of liver is an unavoidable clinical event during liver surgery, such as hepatectomy, hypovolemic shock, and liver transplantation . A largeRenal fibrosis is considered to be a representative pathological character in the development of chronic renal disease, which is a worldwide public health event with high mortality . It is cin vitro. Han et al. found that NLRC5 is highly expressed in HK-2 cell of H/R injury model. It has also been shown that silencing NLRC5 obviously increased the viability of H/R injury HK-2 cell, inhibited the apoptosis of renal I/R injury cell and promoted the recovery of renal I/R injury  injury . TubularR injury . QuanxinR injury . In addiDiabetic nephropathy (DN) is one of the major causes of advanced nephropathy and has become a worldwide public health problem . PatholoRheumatoid arthritis (RA) is a chronic autoimmune disease, which could eventually cause joint deformities and disability . FibroblCardiac fibrosis is one of the most obvious pathological characteristics of myocardial remodeling in heart diseases, which has a high morbidity and mortality . BecauseThe gene of human NLRC5 is highly expressed in lung and spleen . NLRC5 iIt has been reported that DNA methylation plays an important role in genomic dynamics, which is closely related with development of various diseases. It may change at any one site, and thus affect many biological processes of human disease . An accuThe formation of NLRC5 inflammasome is of great importance to recognize pathogens, activate inflammatory caspases, and modulate host defence response . Davis eNF-κB is considered to be one of the most important transcription factors associated with inflammatory diseases. It is known that NF-κB and type I interferon signaling pathways could be directly regulated by themselves through the modulation of feedback loops . NLRC5 pAs reviewed above, the diverse characters of NLRC5 have been introduced and summarized. In addition, recent advance about immanent characteristics, biological function of NLRC5 and molecular mechanisms of NLRC5-mediated immune diseases are also discussed. In the past decades of years, researchers have made much progress in characterizing the NLRC5 crystal structure and detecting the expression of NLRC5 in tissues and cells of immune disease. The fact that NLRC5 plays an essential role in the control of the regulatory mechanism has made it a promising therapeutic target for immune disease. The role of NLRC5, however, still remains controversial in innate immunity and relevant signaling pathways. In a word, the relevant pathways of NLRC5 in innate immunity have not yet been elucidated clearly so far and more experimental investigations about how NLRC5 performs in the host defense against pathogens and transmits immune signals are urgently needed in the future. Moreover, there are few studies on the role of NLRC5 in human immune diseases. Therefore, it is necessary to put on a deep insight on the relationship between the immune function of NLRC5 and human immune diseases. Besides, given that the swift development of molecular docking technology, more precise characterization of crystal deserved to be explored specifically. In summary, these promising properties of NLRC5 make it an appealing therapeutic target through regulating the expression of NLRC5 and NLRC5-dependent signaling pathways.J-QW, Y-RL, and JLi wrote the main manuscript text. QX and JLia prepared The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Aortic valve replacement has stood the test of time but is no longer an operation that is exclusively approached through a median sternotomy using only sutured prostheses. Currently, surgical aortic valve replacement can be performed through a number of minimally invasive approaches employing conventional mechanical or bioprostheses as well as sutureless valves. In either case, the direct surgical access allows inspection of the valve, complete excision of the diseased leaflets, and debridement of the annulus in a controlled and thorough manner under visual control. It can be employed to treat aortic valve pathologies of all natures and aetiologies. When compared with transcatheter valves in patients with a high or intermediate preoperative predictive risk, conventional surgery has not been shown to be inferior to transcatheter valve implants. As our understanding of sutureless valves and their applicability to minimally invasive surgery advances, the invasiveness and trauma of surgery can be reduced and outcomes can improve. This warrants further comparative trials comparing sutureless and transcatheter valves. With the publication of the Cavalier trial2 of the Perceval sutureless aortic valve and the Placement of Aortic Transcatheter Valves (PARTNER) 2 randomised controlled trial4, the spectrum of treatment of aortic valve disease has definitely increased. In this review, we will focus mainly on the most important recent advances in the surgical field, including the incoming surgical valve technologies, such as the sutureless devices, and surgical accesses to the aortic valve, such as minimally invasive surgery via hemisternotomy or right minithoracotomy.The realm of aortic valve replacement (AVR) is quickly changing. The increasing use of transcatheter techniques and the advancement of sutureless valve are contributing to a change in both the indications and the operative strategies for AVR, particularly for patients with aortic stenosis (AS). AVR is indicated in symptomatic patients with severe stenosis (mean pressure gradient of at least 40 mm Hg or maximum velocity of at least 4 m/s) or in asymptomatic patients with impaired left ventricular ejection fraction or low surgical risk5. However, incoming manufacturing technologies such as the one used for the new Resilia Inspiris valve  are meant to significantly increase bioprosthesis durability and the results of the Prospective, non-randomized, Multicenter Clinical Evaluation of Edwards Pericardial Bioprostheses With a New Tissue Treatment Platform (COMMENCE) trial are eagerly awaited6.Standard aortic valve replacement (SAVR) is the classic commonly used approach for aortic valve surgery and is performed through a median sternotomy with cardiopulmonary bypass with excellent outcomes. Following transverse or hockey-stick aortotomy, the pathological valve leaflets are excised and the annulus is debrided. A series of interrupted sutures (with or without pledgets) or continuous sutures are placed under direct vision to anchor a biological valve, whether stented or stentless, or mechanical valve. Whilst mechanical prostheses suffer extremely rarely from structural valve deterioration and have excellent durability, the rates of freedom from structural valve failure in stented bioprostheses are 70 to 90% at 10 years and 50 to 80% at 15 years7. Also, SAVR remains the only option in several hostile conditions such as endocarditis, anomalies of coronary origin, bicuspidy or redo surgery after homograft implantation in the congenital population.Given the outstanding short- and long-term outcomes, SAVR is deemed to be the gold standard operation for aortic valve disease and represents the benchmark against which new therapies are compared8, suggesting that this category still constitutes the greatest part of patients undergoing treatment. When the operative mortality was stratified by using the Society of Thoracic Surgeons (STS) predictive risk classification, actual in-hospital mortality was significantly lower in all patients (2.5% versus 2.95%) both in the overall population and within each risk category. Of note, there was a notable increase in the percentage of high-risk (STS of more than 8%) and intermediate-risk (STS of 4 to 8%) patients undergoing surgery from the earlier to the latter years .Improvements in the perioperative management of critically ill elderly patients with multiple comorbidities have widened the range of patients eligible for surgery. Interestingly, in a 2015 review of 141,905 patients undergoing isolated first-time SAVR between 2002 and 2010, the majority of patients were in a surgical low-risk group However, certain subgroups of patients with significant comorbidities  and deemed at elevated risk might not be considered suitable candidates for SAVR and are currently the main point of discussion at heart team meetings. In addition to significant comorbidities and excessive preoperative risk, SAVR may be declined in otherwise-fit patients presenting anatomical features that determine particular intra-operative challenges . These shortfalls in SAVR have stimulated the development of alternative interventions in the form of sutureless aortic valve replacement (SuAVR), minimally invasive aortic valve replacement (MIAVR) and, more recently, transcatheter aortic valve implantation/transcatheter aortic valve replacement (TAVI/TAVR).9) as in the traditional fashion while still allowing complete excision of the diseased native valve and cleaning of aortic annulus of calcified debris or infected material.Whilst conventional surgical AVR performed by median sternotomy still represents the standard of care in the treatment of aortic disease, less invasive approaches are progressively gaining consensus by providing effective results with a reduced interventional burden on the patient. Sutureless (rapid deployment) valves are bioprostheses that can be surgically implanted without the need of anchoring sutures  and the balloon-expandable Intuity (Edwards Lifesciences) sutureless valve. Sutureless valves are amenable to be implanted by median sternotomy or minimally invasive accesses such as ministernotomy (MIS) and right anterior thoracotomy (RAT)11. These approaches have been shown to reduce bleeding and blood transfusions, atrial fibrillation, wound infection, ventilation times, and time to return to normal activities13.Two main types of sutureless aortic prostheses categorised by implantation mechanism are available on the market14. Sutureless valves can be applied routinely but are particularly pertinent in patients with multiple comorbidities or those in need of multiple procedures by its ability to reduce cross-clamp time. Hanedanet al. showed better haemodynamic outcomes and shorter ischaemic times in elderly, high-risk patients who underwent multiple cardiac surgical procedures when implanted with sutureless valves rather than conventional AVR15. The significant reduction in time of implantation makes the sutureless valve a valuable adjunct in the case of double-valve procedures or need for concomitant atrial fibrillation ablation, as shown recently by Baranet al.16.Indications for sutureless valve implantation equate to those for surgical AVR17.An International Expert Consensus Panel recommends sutureless valves as first choice of valve prosthesis for patients who require concomitant procedures or who have a small aortic annulus. Further indications are for the patient who requires a redo operation or who has a delicate aortic wall condition18.The main drawback of sutureless valves concerns paravalvular leaks and the need for pacemaker implantation. The Surgical Treatment of Aortic Stenosis With a Next Generation Surgical Aortic Valve (TRITON) study with the Edwards Intuity prosthesis demonstrated a paravalvular leak rate at 1 year of 2.3% 20, worse than standard AVR (3.0%). Other significant complications include neurological events (transient ischemic attack or disabling stroke), myocardial infarction, kidney failure, and surgical site infections21.The reported incidence of pacemaker implantation ranges from 5.6 to 9.1% in the literature22. Little difference was found in the rate of pacemaker implantation: 6% in the Perceval S group and 6.8% in the Intuity group, respectively; similar early clinical and haemodynamic outcomes were reported. Perceval S was associated with shorter aortic cross-clamp and cardiopulmonary bypass times , but transaortic peak and mean gradients were lower in the Intuity group 22.Another recent multi-centre retrospective study compared the outcomes of Perceval S and Intuity valves in a propensity-matched analysis23. Two techniques are available and amenable to be paired with the use of sutureless valves: MIS and RAT11.MIAVR is defined as an AVR procedure that, as opposed to conventional full sternotomy, is performed through a small chest wall incisionThe first approach entails a “J” sternotomy performed at the level of the third or fourth intercostal space which provides direct access to the aorta and, in the majority of the cases, to the right atrium, permitting the cardiopulmonary bypass to be established centrally via the ascending aorta and right atrium.et al., accurate planning of this procedure with the use of multi-slice computed tomography is required with the aim to evaluate the anatomical relationship among intercostal spaces, the ascending aorta and aortic valve24. Also, transverse aortotomy is normally performed 2 cm higher than conventional aortotomy; the rest of the procedure is carried out in accordance with the instructions of the manufacturer. Briefly, suture guidance is used to accompany the prosthesis and then the valve is deployed or balloon-expanded according to the model used25.The second approach foresees a skin incision of 5 to 7 cm placed at the level of the second intercostal space to access the chest cavity. Care is taken to avoid rib spreading, although some reports describe the need for rib resection. Direct aortic cannulation can be performed with a flexible cannula while venous drainage is achieved via the femoral vein by using a multi-stage cannula positioned into the right atrium with the Seldinger technique and under transoesophageal echocardiographic guidance. As described by Glauber12. The benefits in the postoperative management of these patients further translate to a quicker return to daily activities with fewer costs related to rehabilitation resources. Minimally invasive AVR can be performed via MIS or RAT; the latter produces more evident benefits26. Sutureless valve technology seems the best fit in this context where the risk profile of the intervention meets the technical challenge of restricted access with more complex angles of chest entrance and field visualisation.A number of meta-analyses have revealed several advantages in using a minimally invasive approach, including reduction of bleeding and transfusion requirement and postoperative complications, such as atrial fibrillation, wound infection and ventilation time, leading to an overall shortening of the length of stay in hospital27. In a meta-analysis27 of small retrospective studies, no significant differences in in-hospital mortality and stroke  were detected when comparing minimally invasive approaches with the aortic valve and conventional sternotomy in re-operative settings. Similarly, no significant difference was found in the length of hospital stay or rates of pacemaker implantation, renal failure, re-operation for bleeding, and hospital stay between the two groups. Volaet al. showed the feasibility of using minimally invasive SuAVR in three patients with degenerated small 19 mm aortic bioprostheses with no mortality and an average implantation time of 10.3 minutes28. However, the current body of evidence relies mainly on outcomes from experienced centres with scarce possibility of diffuse worldwide reproducibility of these results.Minimally invasive approaches to the aortic valve have also been used in the redo setting. Besides the expected technical challenges of redo surgery though a minimal access, the major concern relies in the possibility to deliver an adequate myocardial protection strategy. Also, in the case of redo after coronary surgery, the limited surgical field might render the isolation and control internal thoracic artery (ITA) grafts prior to clamping extremely challenging24. Also, it has been demonstrated that RAT might be a valid adjunct in the treatment of octogenarians or elderly patients and a more expeditious and effective alternative to full sternotomy AVR. In a report by Gilmanovet al., RAT was associated with lower postoperative stroke incidence, earlier extubation and shorter hospital stay29.Several studies comparing the outcomes of RAT with median sternotomy demonstrated lower incidence of postoperative atrial fibrillation, blood transfusion, and shorter ventilation time and hospital length of stay in the RAT group30. In another retrospective study, by Beckmannet al., implantation of sutureless valves in small aortic annulus patients achieved effective orifice areas comparable to patients receiving root enlargement surgery and conventional AVR31. However, there was no difference in 30-day mortality and survival rates and no statistically significant increase in the incidence of patient–prosthesis mismatch31. These findings are pertinent, especially when the risk–benefit balance of performing AVR in high-risk or geriatric patients with small annuli is considered.In a large retrospective study with RAT, sutureless valves outperformed standard suturable valves through the same approach. Although 1-year mortality, incidence of postoperative strokes and pacemaker implantation rate were similar in the two cohorts, cardiopulmonary bypass and cross-clamping times were significantly shorter in the sutureless group and postoperative duration of mechanical ventilation was also reduced. Interestingly, a larger prosthesis could be implanted in the sutureless valve groupet al.32 published a multi-centre randomised trial comparing MIAVR using the Edwards Intuity valve versus full sternotomy SAVR in 46 and 48 patients, respectively. Sutureless valve replacement was associated with significantly lower cross-clamp durations (41.3 versus 54 minutes), mean transvalvular gradients (8.5 versus 10.3 mm Hg) and prevalence of patient–prosthesis mismatch (0% versus 15.0%) at 3 months. This study was underpowered to investigate differences in mortality or morbidity; however, no clear differences in early clinical outcomes, including quality-of-life measures, were found. Pacemaker implantation rates were higher in the sutureless cohort but this was not statistically significant (4.3% versus 0%). Previous non-randomised studies confirmed shorter procedural times, which did not translate to better outcomes, and showed comparable in-hospital mortality and perioperative stroke rates33. Some investigators reported lower rates of blood transfusions, shorter intensive care unit (ICU) and intubation times, and lower incidences of postoperative atrial fibrillation and respiratory insufficiency with SuAVR and this translated to significant overall cost reductions that were attributed mainly to reduced overall hospital stay and diagnostics33. Despite this positive evidence in favour of MIS, a comprehensive meta-analysis in 2017 failed to show a significant advantage in clinical outcomes with the exception of a reduction in postoperative stay and blood consumption and concluded with the need for randomised evidence to elucidate this point34.Borger35.The results of two randomised trials have recently been published. The Mini-Stern trial was a randomised clinical trial comparing full sternotomy with MIS for AVR. The trial failed to show shorter hospital stay and faster recovery or improved survival and was not cost-effective. It was concluded that the MIS approach is not superior to full sternotomy for performing AVR36. Evidently, the benefit of MIS over full sternotomy is still debated and the adoption of one or the other approach is driven mainly by the experience of each centre.More recently, the MAVRIC —a single-centre, single-blind randomised study—compared AVR via manubrium-limited ministernotomy using a 5 to 7 cm midline incision (intervention) and conventional median sternotomy and had postoperative red cell transfusion as the primary outcome. MIS was associated with higher cardiopulmonary bypass time and reduced drain losses but this difference did not translate in a significant reduction in blood transfusion. Additionally, conventional SAVR was found to be more cost-effective 12; however, more randomised evidence is needed to be the final word on this debate.Generally, one of the main obstacles to the wide adoption of minimally invasive AVR is the association with increased operative times, technical difficulty and steep learning curves. Interestingly, a meta-analysis revealed a weighted mean difference of 7.9 additional minutes of cross-clamp time with minimally invasive AVR11. Interestingly, a 2015 report from Hassanet al. examining the cost-effectiveness and logistic balance in MIS and RAT-AVR from eight large studies in the US—while showing no significant inter-group differences in 30-day mortality, conversion to sternotomy, neurologic events, arrhythmia, wound infection, or postoperative bleeding among the two groups—demonstrated slightly better outcomes in terms of blood transfusion and hospital stay37. A subsequent cost-effectiveness analysis demonstrated that, given a volume of 50 cases per year, the added operative costs per case were US$ 4,254 for RAT-AVR and US$ 290 for MIS-AVR. The added costs per case, assuming 200 cases per year, were US$ 4,209 and US$ 290, respectively. A RAT-AVR programme performing 50 cases per year adds US$ 1,063,665 of operative costs over five years compared with US$ 72,500 for a MIS-AVR programme. Unlike the results of previously reported studies, these results suggest that the clinical benefits of MIS-AVR are comparable to or better than those of RAT-AVR and cost less, prompting careful consideration in healthcare delivery organisations when developing minimally invasive surgical valve replacement programmes37.Similarly, RAT outperformed MIS in terms of postoperative complication and length of stay in hospitalet al.38, TAVR has experienced a progressive expansion in indications and a rapid technological development. TAVR was initially targeted at patients who have severe AS and are unfit for conventional surgery42, and in our previous report43, we highlighted the procedural techniques and important trials in this field. The most recent follow-up of these initial trials suggested a progressive shift in indications, including intermediate-risk categories.Since its first application in humans by Cribier44 and PARTNER 2A3—have indeed demonstrated non-inferiority of TAVI versus SAVR for treatment of severe AS in patients at intermediate surgical risk. Compared with SAVR, the SURTAVI also showed that percutaneous technology produced better haemodynamics and significantly lower rates of all stroke at 30 days, acute kidney injury and atrial fibrillation.Two large multi-centre trials—Surgical Replacement and Transcatheter Aortic Valve Implantation (SURTAVI)45.Finally, in low-risk patients, the latest results of the PARTNER study series demonstrated lower rates of death or stroke and new-onset atrial fibrillation in TAVR than surgery at 30-day follow-up, and the composite of death, stroke or rehospitalisation at 1 year significantly favoured TAVR over surgery. Also, no significant differences in major vascular complications, new permanent pacemaker insertions, or moderate or severe paravalvular regurgitation were found among the two groups22 (reported as ranging from 13.2 to 17.1%)47. Lastly, this randomised evidence arises from the comparison of the most modern TAVR technology with old-fashioned conventional AVR through median sternotomy. The recent literature has been intrigued by the idea of comparing TAVR with the use of a similarly advanced technology on the surgical side  within the context of a minimally invasive approach. Sutureless valves with a minimally invasive approach would still have the advantage to remove the diseased valve, thus achieving an adequate calcium debridement of the annulus with potential to implant more haemodynamically performing valves and reduce complications such as paravalvular leaks, one of the main predictors of poor survival48, constituting a valid surgical alternative to TAVI49. Three meta-analyses focusing on comparisons among sutureless valves and TAVI and conventional prosthesis have been performed51. A significant reduction in mortality and complications in the perioperative period was found in the sutureless valve group. The low- and intermediate-risk population benefitted from a reduction of at least 30% in 30-day mortality and in the risk for paravalvular leak51, and Takagiet al. confirmed better survival in sutureless valve–AVR over TAVI after combining the results of direct-comparison and adjusted indirect-comparison meta-analyses50. However, when a comparison with conventional AVR was made, outcomes were burdened by similar mortality but a reduced rate of permanent pacemaker implantation51. These results confirm the findings of a previous meta-analysis including 5,000 high-risk patients in which sutureless valve–AVR produced a reduction in early mortality and postoperative paravalvular leak52 and echo the data from Biancariet al. in intermediate-risk patients53. One of the most recent reports eventually demonstrated better perioperative mortality, 1- and 2-year survival, and paravalvular leak occurrence and similar ICU length of stay, pacemaker implantation need, and kidney failure in the sutureless group25. The SuAVR group was troubled by increased transfusion requirements, whereas TAVR was hampered by major vascular complications25. Interestingly, when the results of these studies comparing MIS sternotomy sutureless AVR with TAVR were specifically examined, better rates of paravalvular leakage and improvement in survival at 2 years could be demonstrated in the surgical group, suggesting that minimally invasive sutureless valve–AVR could be considered as the first-line treatment for high-risk patients in the “grey zone” between TAVR and conventional surgery54. Similarly, RAT sutureless valve–AVR showed a significant reduction in paravalvular leaks and a trend towards improved immediate- and mid-term outcomes and survival when compared with TAVR55.These results should be weighted against the still-unknown long-term durability of TAVR. Patient preference and individual patient factors, such as age and small left ventricular outflow tract, and other factors not normally included in the currently used surgical scoring systems play a significant role. Additionally, compared with TAVR, SAVR continues to have absolute lower rates of residual paravalvular leakage, major vascular complications and new permanent pacemakers56. However, at a later stage, the results of the longer-term follow-up on TAVR allowed a more permissive view on the applicability of TAVR, even on intermediate low-risk candidates59. Again, these results derive from a comparison of TAVR and conventional AVR. The results of a head-to-head randomised comparison between the most modern transcatheter technology and access and the relative counterpart in the surgical field are eagerly awaited.In terms of cost-effectiveness, the literature initially produced discouraging data regarding the reimbursement of TAVR showing a lack of economic advantage in its clinical useGiven the latest evidence regarding the progressive expansion of indications for TAVI, the choice between transcatheter and surgical AVR is continuing to incite debate among heart team members. In this context, SUV-AVR provides an interesting alternative, especially for high-risk patients with borderline indications or contraindications for TAVI.60. However, there is a paucity of long-term durability data in contrast to conventional stented bioprostheses and mechanical valves, which still represent the gold standard for the surgical treatment of aortic valve disease. New incoming technologies combined with minimally invasive approaches will surely achieve more importance in surgical practice, but SAVR remains a cornerstone in “non-conventional” conditions such as redo surgery after homograft or autograft implantation, congenital structural abnormalities, bicuspidy and endocarditis.The face of AVR is rapidly changing in both the interventional and surgical fields. In the surgical realm, the introduction of sutureless valve technologies, obviating the need for anchoring sutures, has been shown to reduce operative time and duration of the cardiopulmonary bypass, being amenable to be applied to combined cardiac surgery procedures . The use of these devices also simplifies minimally invasive approaches and is a valid adjunct in patients with small aortic annulus or fragile aortic wall or requiring redo operations. Increased use of these valves in current surgical practice should be considered by the heart team and encouragedAS, aortic stenosis; AVR, aortic valve replacement; ICU, intensive care unit; MIAVR, minimally invasive aortic valve replacement; MIS, ministernotomy; PARTNER, Placement of Aortic Transcatheter Valves; RAT, right anterior thoracotomy; SAVR, standard aortic valve replacement; STS, Society of Thoracic Surgeons; SuAVR, sutureless aortic valve replacement; SURTAVI, Surgical Replacement and Transcatheter Aortic Valve Implantation; TAVI, transcatheter aortic valve implantation; TAVR, transcatheter aortic valve replacement


Pain is an essential protective mechanism meant to prevent tissue damages in organisms. On the other hand, chronic or persistent pain caused, for example, by inflammation or nerve injury is long lasting and responsible for long-term disability in patients. Therefore, chronic pain and its management represents a major public health problem. Hence, it is critical to better understand chronic pain molecular mechanisms to develop innovative and efficient drugs. Over the past decades, accumulating evidence has demonstrated a pivotal role of glutamate in pain sensation and transmission, supporting glutamate receptors as promising potential targets for pain relieving drug development. Glutamate is the most abundant excitatory neurotransmitter in the brain. Once released into the synapse, glutamate acts through ionotropic glutamate receptors (iGluRs), which are ligand-gated ion channels triggering fast excitatory neurotransmission, and metabotropic glutamate receptors (mGluRs), which are G protein-coupled receptors modulating synaptic transmission. Eight mGluRs subtypes have been identified and are divided into three classes based on their sequence similarities and their pharmacological and biochemical properties. Of note, all mGluR subtypes (except mGlu6 receptor) are expressed within the nociceptive pathways where they modulate pain transmission. This review will address the role of mGluRs in acute and persistent pain processing and emerging pharmacotherapies for pain management. Patients suffering from chronic pain not only experience exacerbated responses to both painful  and non-painful stimuli   are ligand-gated ion channels involved in the fast synaptic response to glutamate. Metabotropic glutamate receptors (mGluRs) are G protein-coupled receptors that are responsible for the slow neuromodulatory response to glutamate. Eight mGluRs have been identified so far. They are named mGlu1 to mGlu8 receptors by chronological order of discovery. Later, based on their sequence homology, signalization and pharmacology, they were subdivided in three groups. Group I mGluRs (mGlu1 and 5) are canonically coupled to Gαq/11 and lead to phospholipase C (PLC) activation that promotes neuronal excitability and are mostly expressed postsynaptically. In contrast, group II (mGlu2 and 3) and group III  mGluRs are predominantly coupled to Gαi/o triggering adenylate cyclase (AC) inhibition. Group II and III mGluRs also regulate neuronal excitability and synaptic transmission through Gβγ subunits, which notably inhibit voltage-sensitive calcium channels and activate potassium channels. Both group II and group III mGluRs are mainly localized on presynaptic terminals. Both iGluRs and mGluRs (except mGlu6 receptor) are expressed all along the pain neuraxis where they shape the transmission of pain information Figure . This phActing on the molecular mechanisms of glutamatergic transmission may, therefore, be a way of developing future analgesics counteracting chronic pain. However, even if iGluR selective antagonists have proven efficacious in releasing several pain states, drastically inhibiting glutamatergic transmission via iGluR blocking inevitably induces numerous side effects, notably hallucinations, ataxia and sedation . They are also referred to as competitive ligands. In mGluRs, the glutamate-binding pocket is located in the extracellular domain of the receptor. Due to the high degree of conservation of the glutamate-binding pocket among the mGluRs, the identification of subtype selective ligands is highly challenging. Therefore, many orthosteric ligands are selective for a specific group but do not discriminate between receptors within the group. The typical specific group I, II or III mGluRs agonists are S-3, 5-DHPG, LY379268 and L-AP4, respectively, and have been used in many preclinical studies. Recently, selective orthosteric ligands have been generated, LY2794193 for mGlu3 receptor  or potentiate (positively modulate) the activity of a co-binding orthosteric ligand at a target receptor and so can act as negative or positive allosteric modulators, respectively. Moreover, neutral allosteric ligands capable of inhibiting the action of either positive or negative allosteric modulators but devoid of activity by themselves have also been described . Negative allosteric modulators (NAM) act as non-competitive antagonists and can have inverse agonist properties, meaning that they can inhibit the constitutive activity of the receptor. Interestingly, due to their non-competitive mode of action, the action of NAMs is less dependent on the concentration of endogenous ligands. Positive allosteric modulators (PAM) can enhance either the potency or the efficacy, or both, of orthosteric agonists. Therefore, in contrast to agonists that maintain the receptor active, pure PAMs potentiate the cellular response resulting from the action of the endogenous ligand. Some PAMs can also directly activate the receptor, referred to as agoPAMs, although such activity is usually partial.The first described allosteric modulators of mGluRs were CPCCOEt, BAY36-7620 and MPEP, which display inverse agonist activity on mGlu1 and mGlu5 receptors  to their projections into the dorsal horn of the spinal cord. Spinal neurons then project to higher centers in the brain which process the sensory information. After nerve injury or inflammation, a number of dysregulations occur in sensory neurons affecting activity, properties or gene expression, driving an increased sensitivity to both non-noxious and noxious stimuli with or without ectopic activities. Because the primary afferents are the first relay of nociceptive transmission and can trigger the chronicization of pain, they represent an interesting target for the development of analgesic drugs.Early evidence of a glutamate role in nociceptive transmission at the periphery derived from the observation of thermal and mechanical hypersensitivity following subcutaneous injection of glutamate into naive rat hind paw  is the first relay in the transmission of sensory information from the periphery to the brain. It is submitted to control from peripheral inputs, interneurons within the SC and both inhibitory and excitatory descending pathways from supraspinal regions. This network makes the SC an important site for the modulation of signals generated at the periphery. Any alteration in neurons from the SC network can imbalance spinal relay and lead to chronic pain conditions.The dorsal horn (DH) of the SC which receives nociceptive inputs is organized into different laminae, from the superficial laminae I to the deep laminae V. Most nociceptive fibers (Aδ- and C-fibers) superficially innervate laminae I-III and, to a lesser extent, laminae V, whereas low-threshold Aβ-fibers mainly project into laminae III–VI. Early studies have demonstrated that glutamate is released from primary afferent neurons into the DH in response to both acute and persistent painful stimuli, highlighting a role of the glutamatergic system in nociceptive transmission  expressed mGlu7 receptors and their role in pain have been investigated. The DS is connected to the descending pain modulatory systems, including to the RVM. When locally administrated in the DS of sham animals, an mGlu7 PAM enhanced pain and simultaneously stimulates ON cells and inhibits OFF cells in the RVM. Whereas, in nerve-injured animals, the mGlu7 PAM has an anti-hyperalgesic effect in addition to increasing RVM OFF cell firing. This opposite effect of an mGluR7 PAM in acute or chronic pain conditions is assumed to be due to the recruitment of different pain pathways (Marabese et al., The first strong evidence of supraspinal mGlu4 involvement in pain processing is thanks to the recent development of an mGlu4 photoswitchable PAM allowing the time resolved control of endogenous receptors in freely behaving animals. Strikingly, dynamic modulation of mGlu4 receptor activation in the amygdala by the photoswitchable PAM reverses, in a light dependent manner, both inflammatory pain-related sensory and affective symptoms (Zussy et al., In vitro, neuroinflammatory factors trigger an opposite regulation in the gene expression of the two predominant mGluR subtypes found in astrocytes and microglia, namely an upregulation of mGlu3 and a downregulation mGlu5 (Berger et al., in vitro yields two opposite effects in cultured microglia, mGlu2 activation enhancing neurotoxicity whilst mGlu3 activation promotes neuroprotection (Taylor et al., in vivo (Fazio et al., Beside neurons, mGluRs are also widely expressed in glial cells, noteworthy in microglia, astrocytes, and oligodendrocytes (for a recent review, see Spampinato et al., The growing number of selective compounds for the different mGluRs has significantly improved our understanding of the specific role of each subtype in nociception. Numerous evidences tend to suggest these receptors are promising targets for the treatment of chronic pain. However, at doses proven to be analgesic, mGlu1 antagonists are associated with motor and cognitive impairment (El-Kouhen et al., Given the analgesic effects observed after targeting peripheral mGluRs, peripherally restricted molecules may have satisfying analgesic effectiveness while decreasing the central-associated side effects. Furthermore, the use of new pharmacological tools such as photoswitchable or caged ligands, which allow the spatiotemporal tuning of mGluRs, could reduce off-target effects related to the modulation of the glutamatergic system outside the pain neuraxis.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


TP53, CCND1 and KRAS) were common to all three types of cancer. The three up-regulated miRNAs were potentially targeted by 18, 15 and 10 genes associated with all three types of cancer. Our current bioinformatics analysis suggests the potential influence of several miRNAs on the development of cancer in SCI. In general, these data may provide novel information regarding potential molecular mechanisms involved in the development of cancer among individuals with SCI. Further studies aiming at understanding how miRNAs contribute to the development of the major cancers that affect patients after SCI may help elucidate the role of these molecules in the pathophysiology of the disease.Patients with spinal cord injury (SCI) have an increased risk of developing esophageal, bladder and hematologic malignancies compared with the normal population. In the present study, we aimed to identify, through in silico analysis, miRNAs and their target genes related to the three most frequent types of cancer in individuals with SCI. In a previous study, we reported a pattern of expression of miRNAs in 17 sedentary SCI males compared with 22 healthy able-bodied males by TaqMan OpenArray. This list of miRNAs deregulated in SCI patients was uploaded to miRWALK2.0 to predict the target genes and pathways of selected miRNAs. We used Cytoscape software to construct the network displaying the miRNAs and their gene targets. Among the down-regulated miRNAs in SCI, 21, 19 and 20 miRNAs were potentially associated with hematological, bladder and esophageal cancer, respectively, and three target genes ( Spinal cord injury (SCI) is an insult that commonly occurs because of trauma resulting in loss or impairment of motor/sensory function below the injury level. According to the 2014 National SCI Statistical Center (NSCISC) annual report .MicroRNAs (miRNAs) are small molecules of highly conserved non-coding RNAs, which promote post-transcriptional regulation of gene expression, and play an important role in both physiological and pathological conditions . SeveralIn a previous study, we reported for the first time a specific pattern of circulating miRNAs expression in individuals with chronic SCI compared with healthy controls . Given tWe previously evaluated 17 sedentary SCI (SCI-S) males with 1 year or more of SCI and 22 apparently healthy able-bodied males . Twenty-Potential target genes regulated by the differentially expressed miRNAs between SCI-S and healthy individuals were predicted using the miRWalk 2.0 online tool . To undeTP53, CCND1 and KRAS genes were common to all three types of cancer  resulted in 18 target genes associated with hematologic cancer, 10 with bladder cancer and 15 with esophageal cancer . The comf cancer .In the present study, a bioinformatics analysis was performed to identify potential miRNAs and their target genes related to the three most frequent types of cancer in individuals with SCI. For this purpose, we analyzed the differentially deregulated circulating miRNAs in individuals with SCI from a previous study of our group  and showTP53 [CCND1 [VEGFA [in silico analysis showed that these genes might be potential targets of down-regulated miRNAs in SCI individuals.Increased risk of bladder cancer has been reported, ranging from 16 to 28 times in patients with SCI compared with the able-bodied population, even in patients without neurogenic bladder ,9. Sever3 [CCND1  and VEGF1 [VEGFA  in bladdin-silico analysis suggested five potential gene targets regulated by this miRNA . Furthermore, miR-25, an up-regulated miRNA in our sample of SCI individuals, may promote migration and invasion of esophageal squamous cell carcinoma cells by suppressing the expression of the CDH1 gene [in silico analysis pointed toward CDH1 as a potential target for this miRNA.A greater risk of hematologic and esophageal cancer has been reported in patients with SCI , but theDH1 gene . Consistin silico analysis, we observed 21 down-regulated miRNAs in SCI individuals that may regulate the expression of 69 potential gene targets associated with hematologic cancer, including acute myeloid leukemia and chronic myeloid leukemia. The miR-15/16 cluster, miR-146a, miR-328 and miR-125b have been associated with development of hematologic cancer [in silico analysis identified several potential targets genes regulated for these miRNAs, including the oncogene PIM1 [in silico analysis revealed nine target genes as potential targets of this miRNA associated with hematological disease, including tumor suppressor genes such as TP53, STAT3 and TCF7 [In our c cancer . In the ene PIM1 , while mand TCF7 .TP53, CCND1 and KRAS) were common to all three types of cancer. When considering the up-regulated miRNAs in SCI patients, three distinct target genes  were common to all three types of cancer. Our in silico analysis showed that down-regulation of miR-15b-5p, miR-30e-3p and miR-374b-5p in SCI individuals may have KRAS as one of the targets gene presents in the three most frequent types of cancer related to SCI. Thus, we speculate that the KRAS gene, may be an attractive target for further studies evaluating oncogenesis in SCI individuals.Notably, three target genes (in silico analysis alone is not enough to elucidate the role of these molecules in the pathophysiology of cancer in SCI, suggesting that further in vitro and in vivo studies should be performed to confirm the role of these miRNAs in the development of cancer associated with SCI.The difficulty in obtaining tumor samples in patients who have developed cancer after SCI in addition to the fact that circulating miRNAs do not always originate from tumor tissue  are limiTo date, we are unaware of any study that has evaluated the association of miRNA expression pattern with any type of cancer in SCI patients. The aim of our study was to investigate whether miRNAs that have been already related to cancer development in the general population might be also potentially related to cancer in SCI individuals. Since our study is a hypothesis-generating research, these findings might help to elucidate the role of these miRNAs in the increased prevalence of cancer in patients with SCI.Understanding the regulatory roles of miRNAs has already been demonstrated in several types of cancer, however few studies have demonstrated the influence of miRNAs on the development of cancer in SCI individuals to date. Our current bioinformatics analysis suggests the potential influence of several of miRNAs on the development of cancer in individuals with SCI. Further studies aiming at understanding how miRNAs contribute to the development of the major cancers that affect patients after SCI may help elucidate the role of these molecules in the pathophysiology of the disease.