Results show that achieving a sustainable diet would entail a high reduction in the intake of meat and vegetable oils and a moderate reduction in cereals, roots and fish products and at the same time increased intake of legumes, nuts, seeds, fruits and vegetables. We identify several current data and research gaps that need to be filled in order to get more accurate results. Overall, our analysis underscores the need to consider multiple indicators while assessing the dietary sustainability and provides a template to conduct such studies in other countries and settings. Future efforts should focus on assessing the potential of different interventions and policies that can help transition the population from current to sustainable dietary patterns.Demand side interventions, such as dietary change, can significantly contribute towards the achievement of 2030 national sustainable development goals. However, most previous studies analysing the consequences of dietary change focus on a single dimension of sustainability  using a limited number of indicators and dietary scenarios. A multi-dimension and multi-indicator analysis can identify the potential trade-offs. Here, starting from the current food consumption data (year 2011), we first designed nine alternative dietary scenarios . Next we calculated three nutritional quality , five environmental , one economic (daily food expenditure) and one human health indicator (DALYs) for current and alternative diets. We found that transition towards a healthy diet following the guidelines of Swiss society of nutrition is the most sustainable option and is projected to result in 36% lesser environmental footprint, 33% lesser expenditure and 2.67% lower adverse health outcome (DALYs) compared with the current diet. On the other extreme, transition towards a meat or protein oriented diet can lead to large increases in diet related adverse health outcomes, environmental footprint, daily food expenditure and a reduction in intakes of essential nutrients . We found that shifting to the vegetarian and vegan diet scenarios might lead to a reduction in intakes of certain micronutrients currently supplied primarily by animal-sourced foods (Vitamin B The food system is highly related to the Sustainable Development Goals (SDGs) involving hunger, nutrition and health, climate change, natural resources, biodiversity and socioeconomics ,2. AgricPrevious studies have found that demand side interventions such as dietary shift  can have positive consequences for both human health and climate change ,11,12,13Recent evidence shows that different dietary scenarios can have widely varying consequences on different dimensions of sustainability . It is tStudies calculating more than one indicator of dietary sustainability are emerging ,24,25,26Recent years have seen development of new approaches to quantify the nutritional quality scores of individual diets taking into account the intake of essential and harmful nutrients and comparing it to their recommended levels ,27,28. AIn terms of various dietary change scenarios often employed, vegetarian and vegan diets are the most common ones. While the vegan diet excludes all animal-sourced foods, the vegetarian diet can include dairy products and an ovo-lacto vegetarian diet can include both egg and dairy products. Another variant, the pescatarian diet, excludes meat but can include fish. Much more relaxed than above, a flexitarian or semi-vegetarian diet, is a predominantly plant-based diet with the occasional inclusion of meat or fish .Few studies to date have integrated environment, nutrition, economic and human health indicators to assess the consequences of different dietary change scenarios. The aim of this study is to fill this research gap and demonstrate the calculation of multiple indicators of sustainability for a suite of dietary change scenarios through a case study of Switzerland and explore the potential synergies and trade-offs across different indicators.The current (year 2011) Swiss consumption of 82 food items was obtained from the Food and Agriculture Organization of the United Nations (FAO) food balance sheet . We call2eq per g of beef; from Springmann et al. . In. In2) orx) is the relative risk of disease for risk factor level x; (x) is the number of people in the population with risk factor level x in the baseline scenario; x in the counterfactual scenario.We estimated the reduced YLL and DALYs attributed to dietary and weight-related risk factors by calculating population impact fractions (PIFs) and applying age-specific and disease-specific mortality rate. PIF was the proportion of disease case that would be avoided when the risk exposure was changed from the counterfactual situation to the baseline situation, while the distribution of other risk factors in the population remained unchanged. By definition, PIF was calculated as follows:Here we regarded dietary and weight-related risk factors as discrete:We first calculated reduced death number (DN) by multiplying PIF with death rate (DR) and the population size (N) of the considered population group (Equation (3)), while YLL was the product of DN and predicted life expectancy (E)   by shifting from reference (current) diet to alternative scenarios for the whole Swiss population. For example for YLL in Equation (5), the impact due to each disease was the sum of the outcomes in different subgroups, which were gender- and age-specific. In Equations (3)–(5), index Here we adopted age- and gender-specific population and disease-specific mortality data from United Nation Population Division and Global Burden of Disease estimates of the Institute for Health Metrics and Evaluation (IHME), respectively. In our model, we estimated the health impact by shifting from reference diet to alternative scenarios for the average Swiss population.f = {redmeat, fruits, vegetables, legumes, fish, nuts & seeds} was calculated by raising the risk factor to the power of the consumption level over the serving size, f and disease/cause of death d was expressed as:For dietary risk factors, under a particular dietary scenario , we assumed that the whole population of a region is subjected to the dietary risk associated with the regional consumption level nbounded . We adopn et al. ). The asWe calculated the combined disease and mortality burden of changes in all six dietary risk factors by using equation:For the weight-related health impact assessment, we first estimated the prevalence  of overweight ( (8) and ) establiglobally .(8)Povew = {normal, overweigt, obese} and the relative risks RR are differentiated by disease d and weight category w; and the population fractions p are differentiated by weight category; they are indexed by ref and scn for their levels in the baseline and counterfactual scenario, respectively. For uncertainty analysis, we accounted for the major uncertainties by using error propagation. With the confidence intervals (CIs) of the relative risk parameters, we calculated uncertainty intervals associated with health impact indicators. See In Equations (8) and (9), We calculated three complementary indicators of the nutritional quality of a national average diet following the framework described in Gustafson et al.  and ChauWe calculated the nutrient balance score (0 < NBS < 100) of national diets using the intake values of calories, all 23 essential nutrients and their reference daily intake (RDI) values qikHere, 2000 represents the total daily energy needs of an individual  and As outlined by Fern et al. , if the The procedure to calculate the disqualifying nutrient score (DNS) is similar to that for NBS except that instead of essential  nutrients, the intake . We firAgain, 2000 kcal represents the total daily energy needs of an individual and  as  to estimate the amount of calorie and nutrients they supply. For nutrients like vitamin K, copper, selenium and choline, we imported data from United States Department of Agriculture (USDA) Food Composition Database [The 82 food items from FAO Food Balance Sheet were matched with the food commodities in Swiss Food Composition Database , we applied the estimated average requirement “cut-point” (EAR-CP) method to estimate the % of people with intakes of a particular nutrient above a demographically-weighted requirement threshold  taken from Arsenault et al. . When a For evaluating environmental impact associated with different dietary scenarios, we linked the consumption amounts of 82 food commodities (gram per capita per day) to commodity-specific footprints regarding GHG emission, freshwater use, cropland use and nitrogen and phosphorus application. We adopted the mean footprint values in the form of environmental impacts per gram of individual foods from Springmann et al. .They imported the GHG emission factors associated with non-carbon dioxide emissions of agriculture from existing studies on crops, livestock, fish and seafood ,56,57,582eq/g), followed by pork (~3 gCO2eq/g), eggs, milk, rice and palm oil (~1–2 gCO2eq/g). The freshwater footprint is high for animal-sourced products, sugar, legumes and rice (0.5–1 m3/kg) compared with other products. The cropland use is high for legumes, vegetable oils & oil crops, nuts & seeds and animal-sourced products (5–11 m2/kg). The nitrogen footprints were high for animal-sourced products, cereals , oil crops, nuts & seeds and fruits & vegetables (10–50 kgN/kg). The phosphorus footprints mainly followed the trends in nitrogen footprint values.Please note that the 95% confidence intervals around these footprint values were not available. Also, these values represent global average footprints as Swiss-specific values were not available for use in our analysis. This might under or overestimate the environmental footprint of diets depending upon whether the production system of a particular food item consumed in Switzerland is more intensive or extensive as compared to the global average. We acknowledge that the environmental impacts may vary a lot depending upon the region and production methods  and futuThere are many other studies providing the environmental footprints of individual or food groups mainly using the life cycle assessment (LCA) approach (see review in Reference ). HoweveTo sort out the issue of how much reduction in environmental footprint is required from current levels or how much increase in footprints is acceptable, the concept of environmental planetary boundaries provides an excellent framework ,7. The p2eq, 5.01 m2, 786 L, 27.4 gN and 6.4 gP, respectively (in per capita per day). Finally, we compared the individual environmental footprint under each diet with their per capita food-related planetary boundaries to understand which diets are acceptable from an environmental point of view.We divided the mean planetary boundaries calculated by Springmann et al.  with theMigros,www.leshop.ch). The cost of current and all alternative daily average diets were then calculated by multiplying the price of each food item with its daily intake amount (g/day). We also examined the correlation between the health based indicator (reduced DALYs), the three nutrition quality indicators , five environmental footprint indicators as well as the economic cost to understand synergies and trade-offs while shifting from current to alternative diets.We collected the price data of 82 food commodities (in Swiss Francs per kg) considered in this study from the online website of the largest food retailer in Switzerland .The health benefits of transitioning to the other vegetarian scenarios  are 25–50% of the above two diets and the estimated health benefit from TAX and HGD scenarios was very small. Due to the similarity between TAX scenario and current eating pattern, the estimated health and environmental benefits are not remarkable. We found that following the meat oriented (MTO) or the protein oriented (PTO) alternative diets with increased consumption of animal resource products would increase the risk of the four disease states considered in this study, associated with an additional 24,000 DALYs approximately .For example, under the RSN scenario, almost 34% of the health benefits could be attributed to the reduced risk of coronary heart disease (CHD) while 30% to the reduced risk of cancer. The beneficial impact of dietary change towards RSN on type 2 diabetes (T2D) risk was around 10% .As shown in We found that the RSN scenario will improve the nutrient balance score ) of Swiss diets by >5 (from current 93 to 99) while keeping the disqualifying nutrient score ) and the % population with adequate nutrition (PAN) score at almost the same levels as currently . In termCompared to the RSN scenario, the current Swiss diet is comprised of almost four times more meat (129 g currently vs. 33 g recommendation), three times higher vegetable oil, two times higher fish products and 1.5 times higher cereals  and roots. On the other hand, the intake of legumes, nuts & seeds is much lower than the recommended levels. The intake of fruits, vegetables, dairy and eggs is closer to the recommended levels .12, the adoption of the VGN scenario will lead to low intakes of these two essential nutrients because they are primarily supplied by the animal based products which are absent from VGN diet and the addition of fruits, vegetables & legumes do not compensate for these nutrients.All other eight alternative dietary scenarios considered here will lead to deterioration in one or the other indicator of nutrition quality . For exaWe found that the PAN score reduces by almost 10% under the VGN and HGD scenarios, meaning that just 87% of Swiss population will be achieving their daily essential nutrient intake goals as compared to 96% under current diet. The protein (PTO) and meat oriented (MTO) dietary scenario lead to reduction in DNS because of increased intake of disqualifying (harmful) nutrients such as cholesterol and total fats . The nut2eq, 590 L, 4.38 m2, 29.03 gN and 5.23 gP, respectively , vegetarian (VGT) and Pescatarian scenario leads to 83%, 63% and 64% reduction in GHG emissions from current levels, the freshwater use stays almost the same .Among the nine alternative diets in our analysis, the scenario following Swiss dietary guideline (RSN) is the cheapest in terms of economic cost based on the food product average retail prices in the largest Swiss supermarket (MIGROS). A shift to RSN scenario is projected to save more than one third of the daily food expenditure primarily because of replacement of expensive meat products with relatively cheaper fruits, vegetables, nuts and legumes . With laThe correlation between health & nutrition indicators and environmental footprints was negative and significant (ρ > 0.75) except for water footprint for which there was no statistically significant correlation. The five environmental footprints were positively correlated with each other except for greenhouse gas and the water footprint which were negatively correlated.N = 9), our preliminarily results are encouraging and show that the transition to a healthy and more sustainable diet in general would not increase economic pressure on consumers.Our multi-indicator quantitative comparison of current and nine alternative diets for the Swiss population revealed that transitioning towards a healthy diet following the guidelines of Swiss society of nutrition will bring the highest sustainability benefits as it is projected to reduce 36% of the environmental footprint, save one third of expenditure on food and lower the adverse health outcome by 2.67% compared to the current diet . AchieviOn the other extreme, our modelling results show that transitioning towards a meat or protein oriented diet is the least sustainable option out of all 10 diets considered here as it is projected to result in large increases in diet related adverse health outcomes, environmental footprint, daily food expenditure and low intakes in essential nutrient with risk of leading to deficiency  exceeds in the intake of harmful  nutrients such as fats, cholesterol and sugar . TherefoIn our environmental analysis, we found that the current greenhouse gas emission and nitrogen footprint of daily average Swiss diet exceeds the respective food related planetary boundaries, meaning that food items high in GHG emissions  and the or USDA  that we  or USDA .For the environmental emission factors, we used the global average values that were publicly available but future studies should use the country-specific emission factor values. Also, we could not consider the impacts of dietary change on other environmental categories such as biodiversity loss, ecosystem services ,67,68,69Third, our health impact estimates  which simply follows the guidelines of Willet at al. [Fourth, we used the average price of 82 food groups from the largest food retailer website in our cost analysis  but the Finally, we only included nine popular alternative dietary scenarios but one can also obtain a sustainable diet using an optimization algorithm that starting with the current diet, generates optimum intake amounts of different food items meeting multiple nutritional, environmental or other constraints .Overall, we estimated the impact of dietary shifts on environment, human health, nutrition quality and economic dimensions with multiple indicators and provide the magnitude of possible benefits for Switzerland if it decides to transition towards sustainable dietary patterns. Importantly, we found synergies among these four dimensions  meaning


The fundamental role that alpha-synuclein (aSyn) plays in the pathogenesis of neurodegenerative synucleinopathies, including Parkinson’s disease, dementia with Lewy bodies, and multiple system atrophy, is a well-accepted fact. A wealth of experimental evidence has linked this relatively small but ubiquitously expressed protein to a plethora of cytopathologic mechanisms and suggests that aSyn may be capable of seeding the progressive spread of synucleinopathy throughout the brain. Beyond the synucleinopathies, the abnormal deposition of aSyn is frequently seen in a variety of other neurodegenerative proteinopathies including Alzheimer’s disease. In spite of the fact that the frequency of concomitant aSyn pathology in these disorders is such that it can be considered the rule rather than the exception, the potential role that aSyn may have in these disorders has received relatively little attention.In this article we postulate that aSyn may in fact be a key protein in driving the pathogenic processes in neurodegenerative comorbidities. In addition to reviewing the frequency of concomitant deposition of aSyn in the neurodegenerative proteinopathies, we also consider our current understanding of the interaction of aSyn with other neurodegenerative disease-associated proteins, including tau, TDP-43, amyloid-β and prion protein, in the context of neuropathologic studies describing the anatomical sites of potential concomitant pathology. We conclude that a growing body of evidence, encompassing neuropathology studies in human brain, animal models of concomitant proteinopathies and studies employing sophisticated methods of probing protein-protein interaction, cumulatively suggest that aSyn is well positioned to exert a strong influence on the pathogenesis of the neurodegenerative comorbidities.We hope to stimulate research in this emerging field and consider that future studies exploring the contribution of aSyn to the pathogenic processes in neurodegenerative comorbidities may provide critical information pertaining to diagnosis and the development of vital disease modifying treatments for these devastating diseases. SNCA, the gene that encodes aSyn, is linked to an autosomal dominant early-onset form of Parkinson’s disease (PD) [Alpha synuclein (aSyn) is a 14 kDa protein ubiquitously expressed in the presynaptic terminals of the brain, where it has been estimated to account for up to 1% of all cytosolic proteins . Since tase (PD)  there haase (PD) , endoplaase (PD) , neuroinase (PD) , disruptase (PD)  as well ase (PD) . In addiase (PD) . CollectThere are several diseases in which pathologic aggregates of aSyn are a defining feature, collectively termed synucleinopathies. Primary synucleinopathies include Lewy body disorders, such as dementia with Lewy bodies (DLB), Parkinson’s disease (PD) PD with dementia (PDD) and pure autonomic failure (PAF). These are characterized by the predominance of intraneuronal cytoplasmic and neuritic deposits (Lewy bodies and Lewy neurites). The classification of these disorders is based on the clinical presentation and spatiotemporal development of aberrant aSyn pathology . A furthCurrently the pathological classification of neurodegenerative diseases is based on the predominant proteinopathy . HoweverPrior to considering any role aSyn may have in driving neuropathologic comorbidities it is important to appreciate the mechanisms by which aSyn has been shown to propagate synucleinopathy alone. This topic has been the focus of an impressive amount of research over the last decade inspired initially by the observation that aSyn pathology is evident in embryonic dopamine neurons grafted into the brains of human PD patients, leading to the hypothesis that aSyn may be capable of propagating synucleinopathy in a prion-like fashion . There iThe association of synucleinopathy with different neurodegenerative conditions can be discussed from two perspectives. First, when aSyn appears as comorbidity  in diveFirst, the classification of disorders with Lewy bodies awaits further crystallization. Diseases with Lewy bodies are grouped based on the early clinical presentation (movement disorder vs cognitive decline) . HoweverSecond, historical studies focused exclusively on the presence of Lewy bodies and their clinicopathological relevance. However, the spectrum of aSyn accumulations in Lewy body disorders is much broader than the mere presence of Lewy bodies and involves depositions in synapses, neurites, astrocytes, and oligodendrocytes , 32–34. The clinical subtypes associated with MSA pathology  cannot bDespite the relative frequency of concomitant neurodegenerative disorders, pure forms of proteinopathies are still apparent using current diagnostic methods. This supports the current protein-based molecular classification . The intTDP-43 pathology is clearly more frequent in Lewy body disorders  , while lOn the other hand, Lewy body pathology is reported in a wide range of neurodegenerative disorders and in unimpaired aging Fig. . StudiesTau. More recent studies in cultured cells, whilst supporting the ability of recombinant aSyn fibrils to induce aggregation of tau, however, have cast some doubt on the reciprocal ability of tau fibrils to cross-seed the aggregation of aSyn [Experimental studies suggest that aSyn and tau, including oligomeric forms, may directly interact with each other, furthermore, co-deposition of both proteins in Lewy bodies, Lewy neurites and tangles has been shown using antibodies against different epitopes of tau and aSyn in postmortem tissues (reviewed in ). The la of aSyn . These d of aSyn . This me of aSyn . Alterna of aSyn .The observation that individuals from the Contursi kindred, who express the aggregation prone A53T mutation in aSyn, also have tau filamentous amyloid inclusions, as do mice that overexpress A53T or E46K mutant aSyn, provides evidence that mutant aSyn can drive aggregation of tau in vivo , 65, 66.2)TDP-43Interestingly, in an experimental setting there is little evidence of overlap between tau and aSyn deposits in either cultured cells or in mice exposed to aSyn PFFs , 69. Fur3)Amyloid-βTDP-43 is frequently associated with tau and Aβ, yet to date only two recent studies, from the same group, have addressed the interaction between aSyn and TDP-43 at an experimental level. SHSY5Y cells, double transfected with both aSyn and a mutant TDP-43 lacking the nuclear localization signal (TDP-43dNLS) developed abundant aggregates of phosphorylated TDP-43 upon exposure to aSyn PFFs . These aIn 1993 Ueda and colleagues reported a study on an unrecognized component of amyloid in AD and tentatively named this 35-amino acid peptide NAC (non-Aβ component of AD amyloid) and its precursor NACP , which sIndeed, experimental studies provide a wealth of evidence thought to support the binding and coaggregation of Aβ and aSyn. In vitro, Aβ can co-seed the aggregation of aSyn in both cell free and cell-based systems –78 and r4)Prion proteinRecent studies have also suggested a further, albeit indirect mechanism by which aSyn may mediate deposition of Aβ. Following the observation that exposure to recombinant aSyn leads to increased levels of Aβ in cultured PC12 cells or primary hippocampal neurons , 86 a reC) has previously been suggested as a receptor for Aβ oligomers [C has also been described as a putative receptor that promotes the uptake of aSyn via binding to the N-terminal domain [C knockdown in murine neuroblastoma cells attenuated the uptake of recombinant aSyn oligomers with a similar effect noted when comparing aSyn uptake in mouse primary hippocampal neurons prepared from wildtype or PrPC knockout mice. In addition, PrPC knockout mice developed lower levels of aSyn aggregates in the cortex, striatum, thalamus and hippocampus following intrastriatal inoculation with aSyn fibrils, suggesting that PrPC may facilitate the uptake and aggregation of aSyn oligomers. These same authors provided evidence that the replication of scrapie prions was blocked by aSyn oligomers [C and aSyn oligomers [C knockout mice and wildtype mice were equally susceptible to aSyn oligomer-mediated toxicity and that PrPC expression was not a prerequisite for these toxic effects. It remains possible that these discrepancies are a result of methodological differences between the studies. Certainly, there are many different species of aSyn oligomers, which may have a differential binding capacity with PrPC and it remains possible that future studies could reveal that both PrPC -dependent and PrPC -independent pathways play a role in synucleinopathies [The cellular prior protein 4). The dIrrespective of which proteins are co-deposited, clearly, the degree of potential overlap is dependent on the temporal stage of each disease, with fewer overlapping regions predicted early in disease and a much higher degree of potential overlap in later stages, thus emphasizing the potential contribution of comorbid proteinopathies to pathogenicity particularly in later disease. It is also important to note that these figures highlight the potential brain regions where one would predict comorbid proteinopathies to coexist, but remain agnostic to the amount of pathology present, or which if any proteinopathy is predominant. Finally, we emphasize that these images Figs.  and 4 reOn an anatomical level the amygdala, a part of the limbic system located deep in the medial temporal lobe, suffers a high degree of overlapping aberrantly deposited proteins with pathology present in AD, synucleinopathies and tauopathies among others . This obGiven the frequency of occurrence, the impact of comorbid disease-associated proteins on disease pathogenesis in neurodegenerative proteinopathies has to date been relatively underexplored. It remains possible that the aggregation of multiple disease associated proteins in a given condition are unrelated to each other, perhaps the result of a common deleterious process in the brain, for example neuroinflammation or disturbances of the protein processing systems. However, emerging evidence suggests that the concomitant deposition of multiple disease-associated proteins may impact upon disease pathogenesis and have clinical significance. Based on neuropathological studies, as depicted in Fig. In this review we speculate that aSyn may be an as yet unrecognised potentiator of neurodegenerative diseases beyond those formally recognized as synucleinopathies. aSyn is bestowed with several characteristics that may allow this protein to impact the pathogenesis of multiple neurodegenerative proteinopathies. First, aSyn is abundantly expressed in the synapses of the brain and indeed, it is named after its localization on synaptic vesicles. All neurodegenerative diseases involve synapses (reviewed in ). Thus, In addition to Lewy bodies or abnormally aggregated aSyn, detectable with current immunohistochemical methods, aSyn may also undergo more subtle biochemical alterations, such as phosphorylation. These changes can occur in human brains with or without neurodegenerative proteinopathies and may also precede the development of Lewy bodies , 106. ThIt is important to note, however, that the evidence linking aSyn to other neurodegenerative proteinopathies is presently largely correlative. Indeed, it remains possible that, being an aggregation prone protein, biochemical and cellular changes in other neurodegenerative proteinopathies trigger coincidental aggregation of aSyn. Thus, until such studies have been undertaken as to provide direct evidence supporting that aSyn is the driving force for other neurodegenerative proteinopathies caution should be exercised in interpreting clinicopathologic observations.In conclusion, it is likely that future disease modifying therapies for neurodegenerative proteinopathies will entail a precision-based approach tailored to the molecular underpinning of disease in each patient. In order to achieve this goal we first need a better understanding of the interaction of different proteins in comorbid proteinopathies. This information will be vital to support effective stratification of patients for clinical trials and eventually for the successful application of disease appropriate therapeutic interventions.


Brain gliomas are the most common primary malignant brain tumors in adults and are graded according to the World Health Organization (WHO), from grade I to grade IV, with increasing malignancy. Recent studies in the field of molecular markers and genomic alterations conducted to a new molecular-based classification of gliomas . First iMolecular markers also provide prognostic information. A 1p19q codeletion is predictive of favorable prognostic and sensitivity to chemotherapy, and methylation of the methylguanine methytransferase (MGMT) promoter gene in glioblastomas is predictive of a better outcome after adjuvant chemotherapy. Also, IDH-mutant grades II and III diffuse gliomas have a more favorable prognosis than those with IDH gene wild type (IDHwt). In parallel, the advances in functional and physiologic magnetic resonance (MR) imaging techniques along with anatomic MR imaging allows us to not only improve diagnostic specificity but also to precisely localize the lesion, define the best surgical approach, and finally perform a longitudinal follow-up after surgery to assess the response to a specific treatment .In this presentation, we will review the role of different anatomic and advanced nonanatomic imaging techniques in the management of brain gliomas. In patients with a suspected brain tumor, anatomic imaging should include T1, T2, and fluid attenuation inversion recovery (FLAIR) MR sequences and T1 with contrast. Computed tomography (CT) may also be helpful by easily detecting calcifications and hemorrhage. Diffusion- and perfusion-weighted images are currently widely used and improve the diagnostic specificity by differentiation between different tumor subtypes and identifying signs of higher malignancy Figure


Clinical diagnosis of neonatal cholestasis is considered to be an extremely challenging process. Here we highlight the importance not only of the prompt distinction between extrahepatic and intrahepatic cholestasis forms, but also of the precise identification of the latter ones amongst the hotchpotch of recently discovered metabolic/genetic causes.Biliary atresia is considered a surgical emergency in a newborn infant. The rate of success in establishing the bile drainage is in fact a function of the early age when the hepato-portoenterostomy intervention is performed. Intrahepatic cholestasis is due to a broad and more and more puzzling variety of infectious, endocrine, genetic, metabolic and toxic disorders where Gamma-glutamyl transpeptidase serum levels may help for differential diagnosis. Recently established laboratory diagnostic techniques have allowed to discover new causes of neonatal cholestasis. Aim of the Commentary is to go through some of them and bring the focus particularly on the information deriving from the paper by Pinon et al. in this issue of the Journal, which paves the way to the inclusion of the hepatocyte nuclear factor-1-beta deficiency as a new condition to consider in the diagnostic process of the syndromic forms with paucity of intralobular bile ducts.Neonatal cholestasis poses diagnostic challenges in practice. Recent advances in the pathophysiology and in molecular genetics together with clinical features, histopathologic findings and careful reasoning remains paramount to put together the pieces of the jigsaw. Neonatal cholestasis affects approximately 1 in every 2500 term infants. It is defined as a reduced bile formation or flow leading to the retention of biliary substances which should instead be excreted into bile and eliminated through the intestinal lumen . On the Due to the broadness of this puzzling scenario, reaching a rapid aetiological diagnosis may represent an extraordinary clinical challenge. In any case, making an early diagnosis remains extremely important to determine a better outcome, and, therefore,the American Academy of Pediatrics and the North American and European Associations for Pediatric Gastroenterology, Hepatology and Nutrition recommend that infants with jaundice longer than 2–3 weeks should have tested both conjugated and total bilirubin, and be evaluated by an expert as soon as possible .. Relevant anamnestic elements should include history of maternal infection or cholestasis during pregnancy, which may be due to the carriage of Progressive Familial Intrahepatic Cholestasis (PFIC) genes. Delayed meconium emission may indicate cystic fibrosis, while a history of consanguinity may suggest a genetic/inherited metabolic disease. Timing of jaundice onset and changes in stool and urine pigmentation may offer clues to the differential diagnosis. Clinical elements to be sought after are the presence of hepatomegaly and liver hardness which may result increased in extrahepatic cholestasis. Splenomegaly may indicate storage disorders (e.g. Niemann-Pick type C disease), while cardiac murmurs and/or renal involvement may suggest either Tyrosinemia or Alagille Syndrome (ALGS). Neurologic signs and symptoms may bring into play some metabolic/mitochondrial disorders, sepsis or acute liver failure. Severe coagulopathy in neonates has been described as the consequence of sepsis or of acute liver failure especially within the historical group of metabolic diseases such as galactosemia, hereditary fructose intolerance and tyrosinemia , and some mitochondrial, peroxisomal and beta oxidation diseases. Thanks to the next generation sequencing, genetic tests for most of these metabolic/genetic conditions are now also commercially available  [Intrahepatic cholestasis is due to a broad and more and more puzzling variety of disorders. GGT serum levels may help in the differential diagnosis. A low or normal GGT can typically be found in PFIC types 1 and 2, or Bile Acids synthesis defects. As shown in Fig. r panel) .The hepatobiliary lesion in ALGS OMIM#118450) consists in paucity of intralobular bile ducts (PILBD). It is a rare multi-systemic autosomal dominant disorder, with mutations of the JAG1 (20p12) pleiotropic gene encoding a Notch signaling pathway ligand (ALGS type 1) or  of the NOTCH2 gene (1p12) (ALGS type 2) –8. Remar450 consifruste form of ALGS however has not JAG1 and NOTCH2 genes mutations, but has instead pathogenic HNF1β (hepatocyte nuclear factor-1-beta) gene mutations. HNF1β, a homeobox transcription factor, also plays a pivotal role in organogenesis during embryonic stage. This recently recognized syndromic disorder presents a variable multisystem phenotype known as Renal Cysts and Diabetes Syndrome . In spite of the name which is due to the characteristics of early reports, however, it is now evident that diabetes and renal cysts are not always present and instead it is associated to a large phenotypic spectrum including pancreatic, urogenital, neurological, parathyroid involvement and – although less typically- liver involvement as well. The latter is now known to encompass from an asymptomatic hypertransaminasemia to a cholestatic liver impairment, including neonatal or late-onset cholestasis [In this issue of the Italian Journal of Pediatrics, Pinon et al.  put the lestasis .In this regard, Pinon et al. , by accuClinical diagnosis of pediatric onset cholestasis all in all remains extremely challenging . One mus


Among the five basic taste qualities, much is known about the molecular receptors and corresponding taste receptor cell types in rodent taste buds for sweet, bitter, umami, sour, and low salt taste  in different parts of the rodent tongue and the palate, which are innervated by different peripheral nerves. There are three morphological types of taste receptor cells (TRCs) in taste buds  and anion (Cl−) are present in extracellular fluids, and the application of hypertonic/hypotonic stimuli to the entire extracellular space may induce non-physiological cellular changes. To circumvent these problems, Roebber et al.  but not a Type 3 TRC marker (anti-Car4). Second, TRCs with amiloride-insensitive high salt responses did not require external Ca2+, characteristic of Type 2 TRCs that utilize internal Ca2+ stores as opposed to Type 3 TRCs that express voltage-gated Ca2+ channels to allow entry of external Ca2+. Finally, sequential application of NaCl and other diagnostic tastants to the same taste bud showed a broad overlap in populations of cells that responded to both NaCl and to saccharin or a bitter mixture of cycloheximide and denatonium, which are known to activate Type 2 TRCs. This result is consistent with a previous study showing that bitter-sensing Type 2 TRCs also exhibit amiloride-insensitive high salt responses , which are known to be expressed in rodent taste buds (Huang et al., + interacts with this pathway.To explore the molecular mechanism underlying the amiloride-insensitive high salt responses in fungiform papilla, the authors substituted choline chloride for NaCl and found that choline chloride yielded Ca− anion. This work provides an important step toward the full picture of how different concentrations of salts are differentially encoded in the gustatory system. Together with prior findings that amiloride-insensitive high salt responses were observed in Type 3 TRCs of the circumvallate papillae (Ninomiya, In conclusion, Roebber et al.  demonstrVM and Y-CC contributed ideas and wrote the final manuscript.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.


This review considers a link between prematurity and autism by comparing symptoms, physiological abnormalities, and behavior. It focuses on the bidirectional signaling between the microbiota and the brain, here defined as the microbiota-gut-vagus-heart-brain (MGVHB) axis and its systemic disruption accompanying altered neurodevelopment. Data derived from clinical and animal studies document increased prevalence of gastrointestinal, cardiovascular, cognitive, and behavioral symptoms in both premature and autistic children and suggest an incomplete maturation of the gut-blood barrier resulting in a “leaky gut,” dysbiosis, abnormalities in vagal regulation of the heart, altered development of specific brain regions, and behavior. Furthermore, this review posits the hypothesis that common genetic variants link the abnormalities in the MGVHB axis in premature and autistic pathologies. This hypothesis is based on the recently identified common genetic variants: early B cell factor 1 (EBF1), selenocysteine tRNA-specific eukaryotic elongation factor (EEFSEC), and angiotensin II receptor type 2 (AGTR2), in the maternal and infant DNA samples, associated with risk of preterm birth and independently implicated in a risk of autism. We predict that the AGTR2 variants involved in the brain maturation and oxytocin-arginine-vasopressin (OXT-AVP) pathways, related to social behavior, will contribute to our understanding of the link between prematurity and autism paving a way to new therapies. This review considers an association between prematurity and autism supported by epidemiological data. It focusses on the disruption of the gut-brain communication and presents a new hypothesis of the common genetic link between prematurity and autism.The rise in global prematurity rates has been accompanied by an increase in prevalence of significant neurodevelopmental disorders including autism spectrum disorders (ASD) ; many prBoth genetic and environmental factors are implicated in preterm birth and autism; more than a few hundred genes associated with risk of autism have been identified, but the first genes associated with preterm birth risk were only recently reported . And whiThis review focusses on the bidirectional gut-brain communication, here defined as the microbiota-gut-vagal-heart-brain (MGVHB) axis. It examines a disruption of the MGVHB axis as a possible link between prematurity and autism by comparing symptoms, physiological abnormalities, and behavior in premature and autistic children. Indeed, clinical data have provided critical observations of increased prevalence of the gastrointestinal (GI) and cognitive problems and behavioral delays in the premature , 18 and The MGVHB axis encompasses the microbiota, the vagus nerve, sympathetic and parasympathetic pathways of the autonomic nervous system including the enteric nervous system, the neuroendocrine and neuroimmune system including the HPA axis, and the brain; more recently, the gut-vagal-heart-brain has been recognized , 26. TheEBF1), selenocysteine tRNA-specific eukaryotic elongation factor (EEFSEC), and angiotensin II receptor type 2 (AGTR2), associated with preterm birth were recently identified in maternal and infant DNA samples using genome-wide association analyses on 43,568 European-ancestry females and confirmed in three birth data sets from Nordic countries [Additionally, we present a new hypothesis that common genetic variants link the abnormalities in the MGVHB axis in premature and autistic pathologies. These common gene variants including early B cell factor 1  pathways, involved in major physiological processes and social behavior, may play an important role in the further understanding of the link between prematurity and autism paving a way to new therapies.Future studies should aim at identification of additional genes associated with prematurity as well as the reexamination of the autism susceptibility genes in the context of their function in the MGVHB axis, communication in premature infants, and neurodevelopmental pathologies, including autism. We tentatively predict that sequence variations in This article reviews the clinical and preclinical literature addressing the postulated association between prematurity and autism at the level of the microbiota, the gut, the vagus nerve, the heart and the brain and behavior, and in the context of newly identified gene variants associated with the risk of prematurity. Literature searches were performed using PubMed base; both original and review articles were selected by best match and the highest number of citations. Over 200 hundred papers were reviewed, out of which 140 were selected for discussion; over 80% of the reviewed papers were published within past 10 years.Gastrointestinal symptoms are often observed in both premature and autistic infants. Higher prevalence of sepsis and inflammatory bowel disease (IBD) and necrotizing enterocolitis (NEC) has been observed in prematurity , with upNormally developing human gut fully permeable in the fetus forms mucosal tight junctions towards the end of pregnancy and becomes less permeable shortly after birth in a process known as “gut closure.” In the premature neonate, the underdeveloped gut lining disrupts initial “gut closure” and the first bacterial colonization, which in turn affect the subsequent gut development  and the Furthermore, in term infants, the developing gut-associated lymphoid tissues (GALT) increasingly differentiate between commensal bacteria or food-derived antigens and the pathogens . In pretImportantly, there is evidence pointing to incomplete gut closure in autism resulting in increased gut permeability—the “leaky gut” , 32, whiGI symptoms, observed in both premature and autistic infants, are closely related to abnormal gut microbiota characterized by a decrease in biodiversity and beneficial bacterial species , 32, 39.Lactobacillus and Bifidobacterium, derived from mother’s vagina and feces. This first gut colonization is disrupted in premature infants [The gut is presumed by some  being st infants , especia infants . Further, and increased prevalence of IBD [The second gut colonization occurs around the time of weaning when the digestive system adjusts to the switch from mother’s milk to solids, and the gut is colonized by strict anaerobes. In the preterm infants, abnormalities in age-specific microflora may interfere with the maturation of epithelial gut-blood barrier and alter gut microbiota interplay  resultine of IBD . Animal e of IBD . Thus, ae of IBD .Gastrointestinal, cardiovascular, and psychiatric disorders  in both Vagus nerve is a primary communication channel between the gut microbiome and the brain and is the integral component of the MGVHB axis. It regulates metabolic homeostasis by controlling visceral functions as well as the innate immune response , 50 and VNA, measured in terms of heart rate variability HRV , is usedImpaired VNA innervation may be responsible for decreased cardiac baroreflex’s (BRS) control of heart rate in premature compared to term infants . AbnormaVagal activity is regulated by gut microbiota. This regulation is mediated through the gut-synthesized factors  and incrBoth prematurity and autism are associated with higher risk of cardiac abnormalities, such as heart failure in prematurity  and abnoOne method used to help diagnose cardiovascular disease  is the HRV, which reflects the autonomic heart regulation and the balance between the parasympathetic and sympathetic input. Preterm neonates show a less complex pattern of HRV  as compaThe occurrence of patent ductus arteriosus (PDA) is increased in the preterm infants  resultinCognitive and behavioral impairments are commonly manifested in both prematurity  and autiMRIs in premature infants subsequently diagnosed with autism have shown reduced volume of temporal, occipital, insular, and limbic regions that are typically involved in autistic pathology , 79. It Brain injury is common in preterm infants and includes intraventricular hemorrhage (IVH), cerebral white matter injury (WMI), and both cortical and deep gray matter involvement , 84. MRIThe cerebellum has emerged as a brain region involved in cognitive, executive, social, and emotional regulation in addition to its importance in motor learning . CerebelAdditionally, behavioral abnormalities in prematurity and autism may be directly related to an “emotion-processing network.” Clinical data suggest that the extent of social immaturity in adolescents born prematurely was related to gray matter changes in the fusiform gyrus and indirectly the orbitofrontal region, suggesting a link between preterm birth and social impairment . ImportaEBF1, EEFSEC, and AGTR2, are associated with gestational duration and preterm birth. Additional three genes, wingless-type MMTV integration site family member 4 (WNT4), adenylyl cyclase type 5 (ADCY5), and an oncogene (RAP2C), are associated with the gestational duration but not preterm birth. Lastly, our search for these genes in databases resulted in locating them among genes associated with increased risk of autism [The data cited above support the notion that prematurity and autism share abnormalities in MGVHB axis that may be linked to common factors regulating fetal development. This concept is supported by two recent studies. First, functional fMRI data have shown diminished functional connectivity in the left hemisphere of the fetal brain, which precedes the preterm birth , suggestf autism . A succiEBF1 gene plays an important early differentiation-promoting role of several neuronal cell types including Purkinje cells [EBF1 is involved in regulation of cell differentiation in the murine striatum [je cells . Alteredje cells  has alsoje cells . Cerebelje cells . Reductije cells . EBF1 isstriatum ; mutatiostriatum .EBF1 gene plays a crucial role in early development of the gut’s B cells, and thus immunoglobulins IgM and IgG, as evidenced by an absence of B cells in EBF1 knockout (KO) mice [KO) mice . AlteratKO) mice  and autiKO) mice .EBF1 has also been shown to be involved in cardiac ventricular conduction system (VCS) as demonstrated by slow VCS in the EBF1 KO mouse [EBF1 gene is involved in the control of blood pressure and carotid artery intima-media thickness. EBF1 variants could lead to bradycardia that is frequently observed in preterm infants. In turn, decreased blood flow to the brain may result in hypoxic-ischemic brain damage triggering neurodevelopmental abnormalities [EBF1 could potentially contribute to low baseline cardiac vagal parasympathetic activity observed in prematurity and autism [EBF1 gene could disrupt the MGVHB by affecting the gut, vagus nerve, the heart, and the brain in prematurity and autism.KO mouse . EBF1 gemalities . Geneticd autism . Thus, aEEFSEC gene is involved in regulation of selenium incorporation into antioxidant and anti-inflammatory proteins, both of which can affect the timing of birth [EEFSEC have been associated with decreased antioxidant capacity in immature infants [EEFSEC gene polymorphism may be compounded by the environmentally induced oxidative stress, and existing management practices in preterm infants, including interrupted nursing and the use of antibiotics [EEFSEC gene expression could thus impact brain development in both prematurity and autism.of birth . Genetic infants . During  infants  and sign infants . On the  infants –111. Oxi infants  as well  infants . Furtheribiotics . DisruptAGTR2 gene encodes angiotensin II receptor type 2 protein that is highly expressed in fetal brain [AGTR2 has been implicated in developmental brain processes [al brain  but restal brain . AGTR2 hrocesses , 118, nerocesses . It is arocesses .AGTR2 receptor subtypes are involved in the negative regulation of AVP by suppressing baseline systemic AVP levels [AGTR2 may thus have a profound impact on the OXT-AVP pathways; abnormalities in the OXT-AVP pathway associated with ASD have been previously reported [Importantly, P levels  but in tP levels . AGTR2 mreported . AGTR2 ireported .AGTR2 observed in prematurity have also been identified in human X-linked intellectual disability, with clinical features including seizures and autistic behavior [Genetic variants in behavior , 122 andbehavior  and coulWNT4 gene, a member of Wnt signaling pathway involved in the development of CNS, affects cell proliferation, differentiation, cell migration, axon guidance, and synapse formation in CNS. Wnt4 is involved in the retinoic acid-dependent neuronal differentiation [ntiation  and has ntiation . Importantiation , 127.ADCY5 gene is associated with birth weight [ADCY5 gene in the dorsal striatum of mouse brain produces autistic-like behaviors [h weight ; its dysehaviors .RAP2C gene, a member of the Ras GTPase superfamily, functions as a molecular switch of cellular proliferation, differentiation, and apoptosis. RAP2C has been associated with preterm delivery [RAP2C is involved in GI development and the regulation of selective permeability of the intestinal epithelial barrier [delivery . RAP2C i barrier . Its dys barrier  and NEC  barrier . Recent  barrier , consistWe have previously addressed the abnormalities in the gut-brain axis in autism , 32 and The key symptoms that emerged linking prematurity and autism are cognitive and behavioral delays and increased prevalence of gastrointestinal problems Table . AdditioThese common symptoms were related to the underlying physiological abnormalities that include the “leaky gut,” dysbiosis characterized by decreased biodiversity and increase in pathogenic species, lower vagal activity and decreased parasympathetic activity, abnormal heart activity reflected in the less complex HRV, and regional brain abnormalities related to cognitive and social delays and critical changes in the cerebellum , could explain higher risk of autism and/or autistic symptoms in premature infants (Fig. The coexpression of common genetic variants associated with risk of preterm birth and those associated with very high risk of autism , such asAGTR2 is in the regulation of circulation between placenta and uterus [AGTR2 is involved in the regulation of blood levels of AVP and OXT [The data discussed above support the notion that prematurity and autism share abnormalities in MGVHB axis that may be linked by common genes regulating fetal development. This concept is supported by the fMRI showing diminished functional connectivity in the left hemisphere of the fetal brain, which precedes the preterm birth , suggestd uterus  that is  and OXT , both re and OXT . Further and OXT . Altered and OXT  and prem and OXT , underscAGTR2 gene variants and their effects on the OXT-AVP pathways involved in major physiological processes, learning, and social behavior may contribute to further understanding of the link between prematurity and autism. The important outcome of such an association will allow for cross-application of therapies between premature and autistic population. Of special relevance and significance would be an application of probiotic and microbial transfer therapies.Clinical, physiological, and behavioral data support the link between the abnormalities in the MGVHB axis in prematurity and autism, suggested by the epidemiological data. This link is further supported by the identification of common genetic variants involved in MGVHB abnormalities in prematurity and autism. Future studies should aim at identification of additional genes involved in the regulation of the time of birth as well as the reexamination of the autism susceptibility genes in the context of their function in the of MGVHB axis. We tentatively predict that


We have used in silico modeling as an alternative approach to characterize the interactive dynamics of Phe and competing LNAAs (CL) in the neurovascular unit (NVU). Our results suggest that plasma Phe fluctuations can propagate into the NVU cells and change there the concentration of LNAAs, with the highest magnitude of this effect observed at low frequency and high amplitude-to-mean ratio of the plasma Phe concentration fluctuations. Our model further elucidates the effect of therapeutic LNAA supplementation in PKU, showing how abnormal concentrations of Phe and CL in the NVU move thereby toward normal physiologic levels.Phenylketonuria (PKU) is an inherited metabolic disease characterized by abnormally high concentrations of the essential amino acid L-phenylalanine (Phe) in blood plasma caused by reduced activity of phenylalanine hydroxylase (PAH). While numerous studies have shown association between high plasma Phe concentration and intellectual impairment, it is not clear whether increased Phe fluctuations also observed in PKU affect the brain as well. To investigate this, time-resolved  Phenylketonuria (PKU) is the most common disorder of amino acid (AA) metabolism, resulting from severely reduced activity of the liver enzyme phenylalanine hydroxylase (PAH), which leads to abnormal accumulation of the essential amino acid L-phenylalanine (Phe) in the blood plasma . Phe is The dysfunction of the PAH enzyme is also the cause of higher than normal fluctuations in Phe plasma concentration, where patterns greatly vary among patients in relation to age, diet, genotype and level of PAH defect . Neverth+-independent antiporter SLC7A5 (LAT1) in microvascular brain endothelial cells (MBEC) (+-independent antiporter SLC7A8 (LAT2) in astrocytes (+-dependent symporter SLC6A15 (B0AT2) in neurons  for transporters at NVU cell membranes . It has s (MBEC) , the Na+trocytes , and the neurons  are the  neurons . In vivoin silico how plasma Phe fluctuations influence LNAA concentrations in the NVU. In addition, we have quantified the variations in concentration of Phe and CL in NVU cells in relation to descriptors of plasma Phe fluctuation, namely mean, fundamental frequency and amplitude-to-mean ratio  and abluminal (abl)  membranes and astrocyte membrane , respectively; Michaelis-Menten binding constants in the individual NVU compartments (LAT1), which corresponds to the ratio between the absolute Michaelis-Menten binding constant for LAT1 in MBECs relative to the corresponding value in the ISF and in plasma  used in the model are taken from literature, and are reported in We employed a previously developed compartmental model of NVU-LNAA homeostasis in adult rats , to whic0AT2 . Fluxes 1P(ISF)) . The flu(T) [see  for deta(T) [see , at eachi  concentrations of Phe and CL in the individual NVU compartments We then prescribed fluctuating plasma Phe concentrations asf is a coefficient that can take on values between 0 and 1, and thereby scales the fluctuation amplitude (relative to mean), and where co is the fundamental frequency of Phe fluctuation, and an and bn are the Fourier coefficients.where n is an integer, fin silico investigation are shown as time series  normalized with respect to the corresponding values for normal physiologic conditions reported in f = 0; For presentation purposes, we initially considered three distinct plasma Phe fluctuation profiles referred to as cases c1–c3, with values of steady state concentration, frequency and relative amplitude as given in 0t), within realistic bounds (0.14 ≤ f0 ≤ 7 cycles per day and 0 ≤ cf ≤ 1)  of a purely sinusoidal signal oscillation,  cf ≤ 1) . We, simf = 0) and sinusoidally fluctuating plasma Phe concentration (0t), f = 0.5 and f0 = 1 cycles/day). We show the responses as time series (excluding transition from steady state) normalized with respect to the corresponding values of baseline physiologic conditions.Finally, we employed the model to investigate the impact of therapeutic supplementation of non-Phe LNAAs on the concentrations of Phe and CL in NVU cells. To this end, we considered the supplemented LNAAs (SL) as CL with the same kinetics, and prescribed it as constant input (representing the effective plasma concentration of SL) to the model. We then determined steady state and fluctuating responses in the NVU, considering, as model input, different values for the effective steady state concentrations of SL in the plasma  and of both steady state  is their low extracellular concentration in the brain interstitial fluid. Phe and CL concentrations in the other compartments are shown in in silico adult PKU rat brain model is qualitatively in line with experimental observations in a PKU mouse model  and To study how perturbations in Phe and CL concentrations in the NVU relate to plasma Phe fluctuation indices, we simultaneously varied fquencies . Finallyquencies .To elucidate the impact of non-Phe LNAA supplementation, we determined Phe and CL concentrations in neurons , MBECs , ISF Su and astrin silico model can serve as an alternative tool to probe these complex dynamics quickly and quantitatively.Failure of AA homeostasis in general negatively affects cell function . In cellin silico adult PKU rat brain model is qualitatively in line with experimental observations in a PKU mouse model (in silico experiments support LNAA supplementation as therapeutic strategy for reducing the propagation of plasma Phe concentration fluctuations into the brain. The model itself can also be seen as a potential stepping-stone for the development of a patient-specific LNAA supplementation treatment-planning tool.We first showed that with increasing Phe concentration in plasma, the steady state concentration of Phe in the NVU and in whole brain increases, while CL concentration decreases. The growth trend we determined for Phe in our se model . We thenThe computational model has been built with a number of simplifying assumptions. In particular, we focused on the pathways mediated by dominant transporters and thus disregarded pathways related to metabolism, diffusion and NVU transporters with low levels of expression, which have been shown to be of lesser importance . Additioin vivo investigations are not readily doable with current techniques. Our results show that plasma Phe fluctuations can propagate into the NVU and change there the concentration of LNAAs, with the magnitude of this effect largely dependent on the frequency and amplitude-to-mean ratio of the plasma concentration fluctuations. Finally, we quantified the therapeutic impact of supplementation of LNAAs in attenuating the disturbed fluctuating and steady state concentrations of Phe and CL in the cells of NVU in PKU disorder toward normal physiologic levels.Using a computational model of adult rat NVU-LNAA homeostasis, we investigated the effects of plasma Phe fluctuations on the dynamics of LNAAs in MBECs, brain ISF, astrocytes and neurons in PKU. Comparable MT implemented the computational model and performed the calculations with SB. FV and VK directed the research. All authors conceived and designed the study, analyzed the data, wrote the manuscript, and approved the final version.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.


Ischemic stroke is a major cause of death and morbidity worldwide. It has been suggested that polyunsaturated fatty acids (PUFAs) may be associated with a lower risk ischemic stroke, but this has been far less studied than their role for coronary heart disease. In this paper, we summarize the main findings from previous follow-up studies investigating associations between intake or biomarkers of the major PUFAs including alpha-linolenic acid (ALA), marine n-3 PUFAs and linoleic acid (LA) and the development of ischemic stroke. Several follow-up studies have suggested that marine n-3 PUFAs may be associated with a lower risk of ischemic stroke although results have not been consistent and limited knowledge exist on the individual marine n-3 PUFAs and ischemic stroke and its subtypes. The role of ALA is less clear, but most studies have not supported that ALA is appreciably associated with ischemic stroke risk. Some studies have supported that LA might be associated with a lower risk of total ischemic stroke, while limited evidence exist on PUFAs and ischemic stroke subtypes. The associations may depend on the macronutrients that PUFAs replace and this substitution aspect together with focus on dietary patterns represent interesting areas for future research. Ischemic stroke is characterized by an episode of neurological dysfunction attributed to cerebral infarction in the perfusion territory of an stenosed or occluded artery . IschemiAn important part of a healthy diet is believed to be an increased intake of polyunsaturated fatty acids (PUFAs). PUFAs are organic acids that naturally contains two or more double bonds in their carbon chain and are named according to the number, configuration and position of these double bonds . PUFAs aThe major dietary n-6 PUFA is linoleic acid LA, 18:2n-6) and together with ALA are the PUFAs consumed in largest amounts. The typical intake of LA in Western populations range between 10 to 17 g/d, while the average intake of ALA varies between 0.5 to 2.3 g/d :2n-6 and,10. LA aLC n-3 PUFAs has been ascribed several effects that might be beneficial in the prevention of ischemic stroke including lowering of triglycerides, inflammation, blood pressure and platelet aggregability, and improved endothelial function and atherosclerotic plaque stability, which likely are mediated by a combination of mechanisms ,15,16,17Several studies have suggested that LC n-3 PUFAs and LA may be associated with a lower risk of coronary heart disease (CHD) ,22,23,24The objective of this paper was to summarize the main findings from previous follow-up studies investigating associations between intake or biomarkers of exposure of the major PUFAs  and ischemic stroke.Few studies have investigated the association between ALA intake and ischemic stroke ,27,28,29The findings from previous follow-up and nested-case control studies using biomarkers of ALA to investigate associations with ischemic stroke have not been conclusive ,38,39,40Limited data are available on biomarkers of ALA and ischemic stroke subtypes . HoweverIn summary, previous studies evaluating either intake of ALA or content of ALA in blood components or adipose tissue have shown conflicting results, but most studies does not support that ALA exposure is inversely associated with ischemic stroke risk.Several studies have investigated the association between intake of LC n-3 PUFAs and total ischemic stroke ,44,45,46Some biomarker studies have investigated the associations between levels of LC n-3 PUFAs in blood compartments or adipose tissue and the risk of ischemic stroke ,39,40,48The associations between adipose tissue content of LC n-3 PUFAs and the risk of ischemic stroke and ischemic stroke subtypes were investigated based on data from the DCH cohort and no appreciable association was observed between total adipose tissue of LC n-3 PUFAs and the rate of total ischemic stroke . HoweverIn summary, several large prospective studies evaluating EPA and/or DHA intake or their content in blood components or adipose tissue have suggested an inverse association although results have not been consistent. The associations between LC n-3 PUFAs and ischemic stroke subtypes may differ with the most beneficial results observed in subjects with strokes of atherosclerotic etiology.Studies of the association between dietary intake of LA and the rate of ischemic stroke are sparse . In analSeveral prospective biomarker studies have supported that circulating or adipose tissue content of LA may be associated with a lower risk of ischemic stroke although results have not been consistent ,39,40,49In summary, limited evidence regarding LA intake and ischemic stroke exist, but LA intake might be associated with a lower risk of total ischemic stroke although the association may depend on macronutrient used to replace LA and the ischemic stroke subtype in question. Several biomarker studies have supported that a high content of LA in blood components or adipose tissue may be associated with a lower risk of total ischemic stroke.Several prospective follow-up studies have suggested that LC n-3 PUFAs perhaps in particular EPA and DHA may be associated with a lower risk of total ischemic stroke although results have not been consistent. A few studies have suggested that the associations between individual LC n-3 PUFAs may differ amongst subtypes of ischemic stroke with the most beneficial findings observed in ischemic strokes of presumed atherosclerotic origin. Regarding the role of ALA for development of ischemic strokes, most prospective studies have not supported that ALA may be appreciably associated with a lower risk of ischemic stroke. The associations between LA and ischemic stroke have been less studied than the role of n-3 PUFAs, but some biomarker studies have supported that LA might be associated with a lower risk of ischemic stroke. Limited evidence exist regarding LA intake and ischemic stroke, but the association may depend on the macronutrient used to replace LA.Biomarkers of PUFA exposure may represent useful complementary measures of exposure to investigate the associations with chronic diseases such as ischemic stroke . DietaryFinally, ischemic stroke is a heterogeneous condition and it is likely that the biological effects of individual LC n-3 PUFAs and LA may differ across subtypes of ischemic stroke of different etiology. Ischemic stroke due to large artery atherosclerosis is considered to be of atherosclerotic origin, whereas ischemic stroke due to small-vessel occlusion may develop as a result of atherosclerosis or lipohyalinosis affecting the smaller penetrating arteries in the brain ,53. In cIn addition, the associations may depend on the macronutrients used to replace PUFAs and such substitution aspects together—with focus on dietary patterns—represent interesting areas for future research.


Phenolic compounds as well as flavonoids are well-known as antioxidant and many other important bioactive agents that have long been interested due to their benefits for human health, curing and preventing many diseases. This review attempts to demonstrate an overview of flavonoids and other phenolic compounds as the interesting alternative sources for pharmaceutical and medicinal applications. The examples of these phytochemicals from several medicinal plants are also illustrated, and their potential applications in pharmaceutical and medical aspects, especially for health promoting e.g., antioxidant effects, antibacterial effect, anti-cancer effect, cardioprotective effects, immune system promoting and anti-inflammatory effects, skin protective effect from UV radiation and so forth are highlighted. Flavonoids and the other phenolic compounds are commonly known as plant secondary metabolites that hold an aromatic ring bearing at least one hydroxyl groups. More than 8000 phenolic compounds as naturally occurring substances from plants have been reported ,2. It isThis work aims to provide an overview of flavonoids and other phenolic phytochems as the potential sources of pharmaceutical and medical applications from the recent published studies as well as some interesting directions for future researches. The key word searches for flavonoids, phenolics, medicinal plant were performed on June, 2018 using Scopus, Google scholar and PubMed. The 351 resulted publications were found and carefully read, in order to find the more recent and non-redundant publications meeting the objective of this work with a few older publications to highlight some necessary points were also used. The 105 selected publications were employed in this review.During the production of adenosine triphosphate (ATP) to generate energy for the cells by using oxygen, reactive oxygen species (ROS) and reactive nitrogen species (RNS) are produced as the by-products from these cellular redox reaction. At the balance level, ROS and RNS are beneficial compounds for cellular functions and immune responses, but the unbalance concentration of ROS and RNS will lead to oxidative stress which can cause chronic and degenerative disorders ,13. The Ipomoea batatas (L.) Lam.), an edible and economic medicinal species in Japan by diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity; the obtained results showed the positive correlation between phenolic content and the activity of free-radical scavenging. In addition, chlorogenic acid was the phenolic compounds that acted as dominant DPPH radical-scavenger in “Miyanou-36” and “Bise” cultivars of I. batatas, whereas anthocyanins were the dominant DPPH radical-scavengers of “Ayamurasaki” and “Kyushu-132” cultivars  is an example of anticancer drugs originated from phytochemical compound for lymphomas and leukemia treatments  were analyzed together with their protective effects on cardiomyocytes treated with doxorubicin , a medicinal plant with extensively use as folk medicine in Central and South America, as an anti-inflammation agent without undesirable side effects from COX-1 inhibition. This work was conducted using in vitro free fatty acid-stimulated macrophage cell lines and combined molecular docking to investigate the interactions between the phenolic compounds and COX-2; the obtained results illustrated anti-inflammatory effects of phenolics from this medicinal plant to regulate macrophages by targeting COX-2 activity inhibition without any action on COX-1 activity [Vitex peduncularis Wall. ex Schauer, a herbal drug, were characterization together with their anti-inflammatory activity by the research group of Ferreres. They found high content of apigenin, C-rhamnosyl flavones and luteolin derivatives in this methanolic bark extract which reduced nitric oxide levels in macrophages and significantly inhibited the activity of phospholipase A2, a mediate enzyme in inflammatory processes [Smilax glabra Roxb and evaluated its anti-inflammatory effects in in vitro lipopolysaccharide-induced RAW264.7 macrophages. Their results pointed out that astilbin significantly suppressed nitric oxide production, tumor necrosis factor-α (TNF-α), mRNA expression of inducible nitric oxide synthase and TNF-α in the tested cells [S. glabra in China and its anti-inflammatory potential was also investigated by Dong and his group in in vivo complete Freund’s adjuvant-induced adjuvant arthritis rats (AA rats) model. Their results showed noteworthy inhibitory properties of astilbin on TNF-α, IL-1β as well as IL-6 mRNA expression; serum cytokine levels of TNF-α, IL-1β, and IL-6 were also decreased in treated AA rats. They also proved that oral treatment of astilbin daily at 5.3 mg/kg can reduce joint damage in hind paw of the animal model; this therapeutic properties of astilbin flavonoid on the inhibition of cytokines production and reduction of inflammatory response in in vivo AA rats model were effective as equal as leflunomide, the frequently used antirheumatic drug [n = 24) with ferulic acid 1000 mg daily and the control group (n = 24) with a placebo for six weeks; ferulic acid supplementation significantly decreased in the inflammatory markers with statistic different comparing with the control group [It is known that COX-2 syntheses prostaglandin E2 is an endogenous pain-producing substance, while COX-1 is a house-keeping enzyme. According to the molecular mechanism of some anti-inflammatory medicines which inhibit both cyclooxygenase-2 (COX-2) and COX-1 enzymes. Consequently, the medicines that inhibits both COX-1 and COX-2 concurrently can cause adverse side effects i.e., renal dysfunction or gastrointestinal bleeding. Therefore, the researchers have challenged to seek for the better candidate for drug development; some phenolics have been reported as the selective inhibiting compounds toward COX-2 expression. An interesting example is the study Ma and his team which aimed to validate the potential and mechanisms of polyphenols from inner bark of rocesses . Additioed cells . Recentltic drug . Moreoveol group .Overexposure to ultraviolet (UV) radiation can harm to skin. It induces extensive production of reactive oxygen species (ROS) and eventually causes skin damages . HoweverHypericum perforatum L. leaves [Ginkgo biloba L. extract (EGb 761) that contains a lot of quercetin derivatives had an ability to decrease sunburn symptoms UVB-induced skin in in vivo study using UVB irradiated-skin mice model; the results indicated that oral intake of EGb 761 may act as a protective and therapeutic agent [Silybum marianum (L.) Gaernt.) fruits contains silybin, a major active component [Apigenin is a major flavones with skin protective effect from UV light; this flavone can be found in many edible medicinal plants or plants-derived beverages e.g., red wine, beer and chamomile tea ,83. Quer. leaves . Topical. leaves . In addiic agent . Silymaromponent . The topomponent . Genisteomponent . Moreoveomponent . Equol iomponent ,92. An iomponent . Interesomponent .There is pros and cons between using synthetic chemical compounds and phytochemical substances in pharmacy and medicine. Synthetic substances or medicines are easy and quickly to produce in large scale of drug development process and modify as many forms of consumption for patients. Conversely, many non-natural  cause several undesirable side effects, particularly long-term treatments . Some syEstrogen is a sex hormone mainly responsible for reproductive functions and the menstrual cycle of women. In postmenopausal women, estrogen is depleted due to the failure of the response of ovary to pituitary. When the level of estrogen decreases, it leads to many postmenopausal symptoms including cardiovascular disease. In particular, postmenopausal woman who also has metabolic syndrome (MetS) will increase in the risk of cardiovascular disease . The res2. The 90 participant subjects were randomly and equally divided into three groups: Genistein, HRT and control group treating with placebo continuously for 1 year. The result indicated that using genistein in postmenopausal bone loss is more effective than HRT, because the undesirable side effects were not found in genistein group. Kruger and his team investigated the effects of bone turnover and the change in microflora between the groups of healthy New Zealand post-menopausal women who received daily isoflavone supplementation (daidzein and genistein) alone and those who consumed green kiwifruit combined with isoflavones for 4 months; their results indicated that the second group of post-menopausal women significantly improved bone health [Glycyrrhiza glabra L. or licorice plant showed no effect on bone loss in ovariectomized rats, an in vivo model of osteoporosis from estrogen deficiency in postmenopausal women [The decrease of estrogen leads to postmenopausal bone loss or osteoporosis. Morabito research team found that genistein can be used as hormone-replacement therapy (HRT) for preventing osteoporosis in postmenopausal women . This ree health . Neverthal women . The secal women ,95,96. MFor over four million years, flavonoids and other phenolic substances from medicinal herbs have been used or consumed by humans so as to live healthy and fight against undesirable diseases ,22,51. AThough many flavonoids and other phenolic compounds were examined from medicinal plant species, a large amount of native or endemic medicinal herbs are still waiting for being survey and observing their novel compounds from profiling works. These processes are an indispensable step to promote the progression in drug discovery and development using phytochemical compounds. Asian region is well-known as one of the greatest hotspot of plant biodiversity including medicinal species, especially Japan, China, Thailand and related areas in tropical and sub-tropical regions ,104,105.(1) The low cost of medicines and many other medical products is very important to allow all people to access to the drug. Consequently, the flavonoid and phenolic compounds which are abundant found in a large number of plant may possible be an interesting choice of molecules for drug and medical product development.(2) In the same species of medicinal plants, the different cultivars may provide different amount of flavonoid and phenolic compounds as well as the biological activities. Thus, the cultivars of medicinal plant should be taken into account for the future medical and pharmaceutical research studies.(3) The geographic areas of raw plant material should also be analyzed and compared in the future research. Since the environmental factors e.g., nutrients and mineral in soil are also effect on the quality and quantity of phytochemical compounds in some species of medicinal plant as discussed in this work.(4) Not only local medicinal plant species but also the wild or endemic species are interesting for the future studies, in order to discover the novel phytochemical compounds to increase the alternative sources of raw material for medical and pharmaceutical applications.(5) The molecular mechanism and signaling pathway of many known flavonoid and phenolic compounds are need to be done in the future, so as to apply this knowledge to the drug development processes.(6) The need of purified compounds to confirm data obtained with the plant extracts.(7) Epidemiological and in vitro studies are sometimes contradictory. In part because of the calculation of the intake based on general table estimated content without taking into account genetic variation among cultivars, geographic variations and so on. Also because of the possible need of metabolization by gut microflora for activation. Those of gut microflora are not taken into account, and quantification of the active circulating forms was not evaluated most of the time.To recapitulate, the use of phenolic compounds and flavonoids are the potential candidate of bioactive agents in pharmaceutical and medicinal sectors to promote human health, prevent and cure various diseases. In order to discover and progress these alternative choice of using phytochemical compounds, the survey of medicinal plants together with intense profiling research needs to be done. The targeted compounds should be employed in biomedical and pharmaceutical research ranging from in vitro, in vivo, and clinical trial step to evaluate the safety, efficacy and also the side effects of the tested candidate compounds.


A marked increase in the rate of glycolysis is a key event in the pathogenesis of hepatocellular carcinoma (HCC), the main type of primary liver cancer. Liver cirrhosis is considered to be a key player in HCC pathogenesis as it precedes HCC in up to 90% of patients. Intriguingly, the biochemical events that underlie the progression of cirrhosis to HCC are not well understood. In this study, we examined the expression profile of metabolic gene transcripts in liver samples from patients with HCC and patients with cirrhosis. We found that gene expression of glycolytic enzymes is up-regulated in precancerous cirrhotic livers and significantly associated with an elevated risk for developing HCC. Surprisingly, expression levels of genes involved in mitochondrial oxidative metabolism are markedly increased in HCC compared to normal livers but remain unchanged in cirrhosis. Our findings suggest that key glycolytic enzymes such as hexokinase 2 (HK2), aldolase A (ALDOA), and pyruvate kinase M2 (PKM2) may represent potential markers and molecular targets for early detection and chemoprevention of HCC. Hepatocellular carcinoma (HCC) is the main type of primary liver cancer and the second leading cause of cancer-related mortality worldwide, with more than 700,000 deaths every year . The incThe development of HCC has been closely linked to cirrhosis, an inflammatory liver condition in which the normal liver tissue is replaced by scar tissue and regenerative nodules after long-term damage induced by various etiologies including hepatitis B (HBV) or C (HCV) viral infection, chronic alcohol consumption or nonalcoholic fatty liver disease . In factin vivo. In addition, whether the aerobic glycolytic metabolism is operative in the early stage of the hepatic carcinogenesis particularly in cirrhosis and correlates with poor patient prognosis have not been completely elucidated. Such a background prompted us to investigate the expression level of enzymes required for the glycolytic and mitochondrial metabolism in liver samples from patients with cirrhosis and patients with HCC available from six open source data sets.Like many other cancers, HCC develops slowly after progressive accumulation of genetic and epigenetic alterations in liver cells accompanied by substantial changes in energy metabolism leading to unrestricted proliferation of mature hepatocytes. One of the most common metabolic changes observed in HCC cells is an increase in the rate of glycolysis with consequent lactate production . This meDifferential gene expression analyses involving multiple clinical samples were performed using different data sets available through Gene Expression Omnibus (GEO) database . IndepenU test, assigning a specific threshold . The analyses were performed in Prism GraphPad packages, as previously reported  (http://gepia.cancer-pku.cn/index.html (accessed on April/May 2018)  (accessed on August 2018) was used to generate Kaplan-Meyer plots of an additional liver HCC data set .For the differential gene expression analyses of TGCA-LIHC we used a user-friendly, interactive web resource for analyzing cancer transcriptome data accessible at ay 2018) . For theay 2018) . KM plotdata set . In bothA salient feature of HCC cells is that they adjust their metabolic profile to fulfill the bioenergetics and anabolic demands of the high rates of proliferation . Yet, liFigure 1A; Figure 1B, expression of the majority of glycolytic transcripts, including the rate limiting glycolytic enzyme HK2 [hexokinase 2], ALDOA , PFKL , GAPDH [glyceraldehyde 3-phosphate dehydrogenase], PKM2 [pyruvate kinase M2]) was significantly increased in HCC livers compared to their adjacent non-tumor tissues (P < 0.0001). An exception was the expression of PGAM1 transcripts that was still significantly higher (P = 0.0021) but with a less extent compared to other enzymes. LDHA [lactate dehydrogenase A] enzyme converts pyruvate into lactate in a reaction that generates NAD+, diverting glycolysis-derived pyruvate from the mitochondrial oxidative pathway . Similar results were also obtained in The Cancer Genome Atlas (TCGA) dataset consisting of 371 primary HCC tumors and 50 normal liver samples (LIHC cohort) .The glycolytic pathway consists of ten enzymatic reactions through which glucose is converted into pyruvate. Among the multiple isoforms of enzymes that catalyze each glycolytic reaction, we evaluated mRNA expression levels of those that are predominantly expressed in the liver ( pathway . As such cohort) . We obseFigure 2A; P < 0.0001) and normal livers (P < 0.0001) samples, respectively . Similar to G6PD, mRNA levels of the other two enzymes involved in the oxidative phase of the PPP, 6-phosphogluconolactonase (PGLS) and 6-phosphogluconate dehydrogenase (PGD), were higher in HCC samples compared to control tissue . Altogether these analyses are consistent with an increase in glycolysis and PPP pathways, leading to sustained ATP and cellular building blocks production both needed for abnormal hepatocytes proliferation  or normal livers (P < 0.0001), while in HCC samples the expression of the succinate dehydrogenase (SDHB), which converts succinate into fumarate in the TCA, was significantly lower than that in non-tumor tissue (P < 0.0001) or normal livers (P < 0.0001) . These observations are consistent with a recent study showing that decreased expression levels of SDHB in HCC promote the Warburg effect (P = 0.0010) from the GSE36376 data set, while IDH2 expression showed no changes in the TGA-LIHC data set (P = 0.7106) .Once in the mitochondria, pyruvate can be converted into acetyl-CoA by the pyruvate dehydrogenase (PDH) complex. Acetyl-CoA then enters into the TCA cycle to generate NADH and FADH2, which transfer their electrons to the electron transport chain to generate ATP through OXPHOS . In two g effect . Less clP = 0.4934) or significantly reduced  in HCC when compared to adjacent non-tumor tissue in the GSE36376 cohort . However, the expression levels of all three genes showed a significant increase in HCC samples compared to normal liver samples .Oxidative phosphorylation is coupled with the electron transport, which is organized into four large membrane-embedded proteins complexes I to IV , to geneFigures 3). Because HCC develops in the settings of cirrhosis, it is likely that surrounding non-tumor tissue is indeed a cirrhotic tissue with the presence of tumor nodules nearby . Similar trends were also detected for other glycolytic genes with some reaching sufficient significance  and very few not reaching significance  (data not shown). On the contrary, PFKL follows an inverse trend. High expression of PFKL was, indeed, inversely associated with poor overall survival, but this was not statistically significant (P = 0.420) . Moreover, although in our differential gene expression analyses LDHA mRNA expression was not sufficiently higher in HCC livers compared to healthy and surrounding non-tumor livers , it seems that LDHA expression has a prognostic value in HCC . This is in agreement with previous studies demonstrating that high level of serum LDH is associated with poor patient overall survival in several malignancies, including HCC  through the online tool GEPIA, a web server for cancer and normal gene expression profiling and interactive analyses . The cumding HCC .Figure 4B). Of note, although the expression of PGLS had no prognostic value , it seems that at least the rate-limiting enzyme G6PD of the oxidative phase of PPP has important clinical implications for patients with HCC. Surprisingly, metabolic genes involved in mitochondrial metabolism have no prognostic value in HCC as neither the expression of genes related to TCA cycle nor OXPHOS was associated with poor patient overall survival  despite their significant increases in HCC samples compared to normal livers . This could indicate that TCA and OXPHOS enzymes may be subject to a strict post-translational regulation that uncouple the mRNA levels from the actual enzyme function in HCC livers.In examining the individual genes functioning in the oxidative phase of the PPP, we found that the overall survival rate was significantly lower in the group of patients that expressed high levels of G6PD, the rate-limiting enzyme of the PPP, and PGD mRNA . Overall these data indicate that glycolytic genes have prognostic role in HCC.Interestingly, similar trend was obtained by analyzing survival of HCC patients by using a distinct data set via the KM plotter web server tool   expression of LDHB was also increased in cirrhotic liver samples compared to normal livers . On the contrary, expression levels of PFKL were significantly higher in cirrhotic livers compared to healthy specimens in the GSE25097 cohort (P = 0.0086) but found constant in the two other data sets (P = 0.1824 and P = 0.9888). Collectively, these results are consistent with those observed in HCC. Indeed, while the expression of HK2, ALDOA, and PKM2 transcripts is increased in HCC compared to normal livers and associated with poor overall patient survival, elevated levels of PFKL scored in HCC had no prognostic value . This suggests that the expression of these genes in cirrhotic livers resemble the gene expression signature of HCC .Liver cirrhosis is a precancerous state of HCC . Therefoin vivo . To bettFigure 5B). Notably, no significant increase of glycolytic and PPP gene expression has been observed in HCC compared to cirrhotic livers , suggesting that reprogramming of glucose metabolism may occur at pre-cancerous stages. For the TCA enzymes, the mRNA levels of PDHA1, IDH2, and SHDB were constant in both data sets of cirrhotic livers compared to normal livers . Similarly, transcripts levels of representative OXPHOS genes that were highly expressed in HCC were not found differentially expressed in cirrhotic livers compared to healthy livers . Collectively, these results suggest that, in contrast to a general increase in glycolysis genes, TCA and OXPHOS genes remained at the same level in cirrhosis. These observations suggest a shift toward aerobic glycolysis and PPP relative to oxidative phosphorylation in cirrhotic livers. This is also supported by the fact that, in contrast to HCC , cirrhotic livers exhibit elevated expression of LDHB compared to healthy livers , suggesting that pyruvate may be diverted away from the mitochondria for ATP production.We also evaluated the levels of transcripts encoding genes in the oxidative phase of PPP and found that while in the GSE25097 dataset there was a significant increase of the three PPP genes analyzed (including the rate limiting enzymes G6PD), no changes were observed in the other two data sets, except for PGLS, whose expression was significantly lower in cirrhotic livers  and high (n = 60) HCC risk based on the rates of patient survival and incidence of developing HCC , while PGLS and PGD show no differential expression . Finally, in line with results showed in Figure 6, no increase in expression of the metabolic genes in TCA cycle and OXPHOS was observed between the low and high HCC risk groups . On the contrary, the expression of IDH2, SDHB, and COXA5 were inversely correlated with poor prognosis . Thus, in cirrhotic livers high expression of glycolysis genes is associated with an elevated risk of developing HCC, suggesting that the glycolytic phenotype has a possible prognostic role in cirrhotic patients. Moreover, HK2, PFKL, ALDOA, PKM2, and LDHB levels show a weak differential expression in cirrhotic and HCC livers , indicating that the acquisition of a glycolytic phenotype occurs during both initiation and maintenance of HCC. Collectively, these results suggest that glycolytic enzymes may represent potential targets for HCC chemoprevention.For the oxidative phase of PPP enzymes, high G6PD mRNA expression levels were found to be associated with a progression of cirrhosis to HCC and reduced survival rate , fructose-1,6-bisphosphatase (FBP1), and PEP carboxykinase (PEPCK) are known to bypass the first, second and third irreversible reactions of the glycolytic pathway, respectively, resulting in slowing down the rate of glycolysis in normal hepatocytes (Recent works have established that the pyruvate kinase M2 isoform (PKM2), which catalyzes the synthesis of pyruvate and ATP, using phosphoenolpyruvate (PEP) and ADP as substrates, is a critical driver of the Warburg effect . Indeed,Figure 7), suggesting that glycolytic-derived pyruvate may not be all converted in lactate and is therefore channeled to other metabolic pathways .We indeed observed remarkable higher expression levels of PKM2 in cirrhotic and HCC livers compared to normal livers. Thus, although further studies are required, it is reasonable to hypothesize that the enhanced expression of PKM2 in cirrhotic livers may contribute to the apoptosis resistance of damaged (injured) hepatocytes that continue to survive, proliferate and accumulate until becoming malignant HCCs. Of note is also the observation that the expression of LDHA (or LDHB) (the glycolytic enzymes converting pyruvate in lactate) have no HCC risk associated in cirrhotic patients , rather than mitochondrial metabolism that contributes to the generation of ROS .Indeed, it has been suggested that in cancer cells the production of ROS by complex I, II, and III of the electron transport chain contributes to DNA damage and is required for cancer cell survival, Kras-induced cellular transformation and cancer metastasis . Thus, iPrevention of HCC is an unmet medical need . InteresAlthough there are potential limitations associated with the nature of the analyses carried out in this study, we sought to detect an early sign of metabolic changes in the development of HCC and show that HK2, ALDOA and PKM2 expression levels in cirrhotic livers could be used as new predictive biomarkers for HCC development. Early studies demonstrated that rat cirrhotic livers show a metabolic shift from OXPHOS to glycolysis, while normal rat hepatocytes continue to use OXPHOS . FurtherThe study involves the analyses of public available data sets using human subjects. Therefore, approval for the use of human subject has been granted to third parties that have deposited the public data sets, which have been properly cited in the text.NL, MC, CB, and SP performed the experiments. CB and SP are both senior authors, conceived the idea, chiefly carried out data analysis and interpretation, coordinate the study, and wrote the manuscript, which was commented by all the authors.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.