Psoriasisis an inflammatory disease of the epidermis based onan immunological mechanism involving Langerhans cells and T lymphocytesthat produce pro-inflammatory cytokines. Genetic factors, environmentalfactors, and improper nutrition are considered triggers of the disease.Numerous studies have reported that in a high number of patients,psoriasis is associated with obesity. Excess adipose tissue, typicalof obesity, causes a systemic inflammatory status coming from theinflammatory active adipose tissue; therefore, weight reduction isa strategy to fight this pro-inflammatory state. This study aimedto evaluate how a nutritional regimen based on a ketogenic diet influencedthe clinical parameters, metabolic profile, and inflammatory stateof psoriasis patients. To this end, 30 psoriasis patients were subjectedto a ketogenic nutritional regimen and monitored for 4 weeks by evaluatingthe clinical data, biochemical and clinical parameters, NMR metabolomicprofile, and IL-2, IL-1β, TNF-α, IFN-γ, and IL-4concentrations before and after the nutritional regimen. Our datashow that a low-calorie ketogenic diet can be considered a successfulstrategy and therapeutic option to gain an improvement in psoriasis-relateddysmetabolism, with significant correction of the full metabolic andinflammatory status. Psoriasis is associated with morbidity and mortality.2 Generally, significant differences among individualsfrom various ethnic groups and geographical location have been recorded,with an increased incidence in individuals living at high latitudes.3 From a sex viewpoint, some studies reported differencesbetween males and females.5 Patients with psoriasishave a decreased quality of life,6 withanxiety and depression. Psoriasis is a disorder of multifactorialetiology with both genetic and trigging factors. Numerous studieshave reported the identification of genetic loci, in particular, 10loci as susceptibility regions. The factors identified as triggersfor the development of psoriasis are trauma, obesity, infections,medications, sunlight, stress, alcohol, smoking, and endocrine factors.7Psoriasis is a chronicinflammatory and multifaceted disease. Thiscondition affects approximately 2–3% of the world’spopulation8Histologically, this disease is characterizedby hyperproliferativekeratinocytes and the infiltration of prominent T cells, dendriticcells, and neutrophils in the dermis. The diagnosis is usually clinical,including an examination of the primary lesion and affected areas.General presentations include plaque, inverse, guttate, erythrodermic,and pustular forms, with cutaneous manifestations and nail, scalp,and joint abnormalities. The psoriasis area and severity index (PASI),body surface area (BSA), and dermatology life quality index (DLQI)are tools that are commonly used for the classification of plaquepsoriasis.11 Moreover, numerous studies have reported evidenceof a causal relationship between obesity and psoriasis, investigatingthe connection between body mass index (BMI) and psoriasis.13A systematic review andmeta-analysis summarize the epidemiologicalassociation between psoriasis and obesity, indicating a higher prevalenceand incidence of obesity in psoriasis patients, compared with thegeneral population.12 especially adherence to a low-calorie dietary regimen.17 Fatty tissue is an active endocrine tissue and causes a pro-inflammatorystate in obese patients.18 The clinicaleffect of weight loss is the reduction of adipose tissue as a sourceof pro-inflammatory cytokines. From this perspective, a very-low-caloriediet with adequate protein content is responsible for weight lossand the reduction of visceral fat mass.22A few studies of weight-lossinterventions have been shown to improvepsoriasis and to increase the response to treatment,23 At a biochemical level, the ketogenicdiet induces a switch to ketone metabolism, causing a reduction inblood glucose and an increase in blood ketones and mitochondrial function.24 Recent scientific studies have shown the therapeuticpotential of ketogenic diets in many diseases, such as diabetes, polycysticovary syndrome, acne, neurological diseases, cancer, and the ameliorationof respiratory and cardiovascular disease risk factors.30 Moreover, the ketogenic diet has been identified as an effectiveremedy for obesity and psoriasis, with a significant reduction ininflammatory components that are possibly localized in visceral adiposetissue.22The ketogenic diet is anutritional regimen characterized by areduction in carbohydrates and a relative increase in protein andfat.31 In this novel study, we performed an NMR-based metabolomicanalysis of the sera of psoriasis patients subjected to a ketogenicnutritional regimen for 4 weeks. Metabolomic data analyzed with theaid of univariate and multivariate statistical methods were correlatedwith the biochemical and clinical parameters, including IL-2 and IL-1βcytokines. Metabolomic profiles of psoriasis patients compared tothose of healthy controls before and after a 4 week ketogenic dietprovide preliminary indications to identify candidate biomarkers usefulin the theranostic control of psoriasis. Results of the metabolicpathway analysis reveal the therapeutic potential of a dietary regimenand provide new insights into the etiopathogenesis of psoriasis.In recent years, metabolomic studieshave played a significantrole in revealing biomarkers, identifying the biochemical pathwaysinvolved in many diseases, and in providing information related topathway perturbations. Nuclear magnetic resonance (NMR) spectroscopyrepresents a robust and suitable technique for metabolomic approaches:low-molecular-mass compounds can be concurrently qualitatively andquantitatively detected in biological samples.The study was conducted at the NutriKeto_LAB,Azienda Ospedaliera “San Giuseppe Moscati”, Avellino,Italy, between October 1, 2018, and March 1, 2019. Consecutive participantswere recruited from hospital wards. Demographics and clinical informationare reported in Participants eligiblefor inclusion criteria in the psoriasis group were overweight patients,aged 18–65 years old, with plaque psoriasis: 35 patients werescreened, and 30 patients were recruited and completed the interventionstudy. The exclusion criteria were pregnancy; breastfeeding; insulintreatment; heart, kidney, or liver disease; obesity due to hypothyroidism;neoplastic disease; intentional or unintentional weight loss of morethan 5 kg up to 3 months beforethe study; active treatment or treatment in the past 2 weeks withtopical drugs or retinoic acid, ultraviolet light therapy or systemictherapy (in the recent 4 weeks), or biological preparations in thepast 12 weeks; and recent history of drug addiction or alcohol abuse.Blood serum samples were collected from30 healthy subjects used as controls and 30 subjects diagnosed withpsoriasis according to the mentioned psoriasis diagnostic criteria.The institutional ethical committee of Azienda Ospedaliera “SanGiuseppe Moscati”, Avellino, Italy, approved the study protocol,which followed the 1964 Declaration of Helsinki and its later amendments,and all subjects gave written informed consent.2) kg/m2. A flexible plastic tape was used to assess waist and hipcircumferences. Blood samples were analyzed in the clinical laboratoryusing automated analyzers and available commercial kits. Quantitativeevaluation of the following clinical parameters was performed: hemoglobin,total lymphocyte count, creatinine, uric acid, glucose, insulin, C-peptide,glycated hemoglobin, growth hormone (GH), total cholesterol, high-densitylipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol,triglycerides, apolipoproteins A1 and B ,albumin, cholinesterase, serum aspartate aminotransferase (AST), alanineaminotransferase, gamma glutamyl transferase (γGT), lactatedehydrogenase, sodium, potassium, magnesium, calcium, phosphorus,the homeostasis model assessment insulin resistance (HOMA-IR), bilirubin,hematocrit, prothrombin activity, cortisol, vitamin B12, folic acid,azotemia, insulin, and homocysteine. For the assessment of visceraladipose tissue (VAT), ultrasound measurement of the aortomesentericfat thickness (AMFT) was performed according to a previous procedure.32The diagnosis of plaque psoriasiswas made by dermatologists based on clinical characteristics. PASI,BSA, the DLQI, and the VAS for itch ratings were employed to measurepsoriasis severity. All the participants had height and body weightmeasured by calibrated flat scales equipped with a telescopic verticalsteel stadiometer . BMI was calculatedas the weight (kg) divided by the height squared  and after 4 weeks (T1). The control group  adopted a conventional diet and was instructed to eat ordinaryfood according to the national guidelines for a healthy diet.l-arginine, α-ketoglutarate, l-ornithine, l-carnitine, l-glutamine, taurine, l-citrulline, l-cysteine, and vitamin B6. Other dietarysupplements were alkalizing substances and herbal remedies , such as garcinia (Garciniacambogia), hawthorn (Crataegus oxyacantha), java tea (Orthosiphon stamineus), dandelion , thistle fruit extract (Silybum marianum), a multivitamin /multimineral supplement, and 10g of hydrolyzed collagen powder. Ashwagandha (Withaniasomnifera) and bacopa (Bacopa monnieri) were also administered for psychophysical balance, and Triphala was administered to implement the correctintestinal function. Patients were advised to drink at least 2 L ofbicarbonate-rich alkaline water per day (not tea or coffee) and tonot use table salt but to salt their food with potassium chloride.All treatments with hypoglycemic agents and diuretics were interrupted.Diet recommendations for psoriasis patients included the consumptionof a very-low-calorie  protein-based diet providing10–20 g of carbohydrates ,20–30 g of lipids, and 1.4 g per kg of ideal body weight  of protein per day. Half of thisprotein dosage is sufficient to supply 12 g of 90% whey protein, withthe addition of The primary outcomewas PASI, an index of psoriasis severity: the PASI was measured atbaseline (T0) and at 4 weeks (T1), along with BSA assessment. Thesecondary outcome was DLQI to determine the quality of life, the reductionin BSA, the improvement in itch severity, and weight loss.33 To obtain the blood serum, the whole blood was collected into tubesnot containing anticoagulant and allowed to clot at room temperaturefor 30 to 120 min. After centrifugation at 12,000 g, the blood serum was aliquoted and stored at −80 °Cin Greiner cryogenic vials before NMR spectroscopy measurements. Beforebeing transferred to a 5 mm heavy-walled NMR tube, samples were thawedat room temperature. NMR samples were prepared by mixing 300 μLof blood sera with 200 μL of phosphate buffer, including 0.075M Na2HPO4·7H2O, 4% NaN3, and H2O. Trimethylsilyl propionic-2,2,3,3-d4 acid, sodium salt (0.1% TSP in D2O) was usedas an internal reference for the alignment and quantification of NMRsignals; the mixture, homogenized by vortexing for 30 s, was transferredto a 5 mm NMR tube (Bruker NMR tubes) before analysis started.31NMR sample preparationand NMR spectra acquisition were performed as previously reported.z-gradient CryoProbe.TOPSPIN, version 3.0, was used for spectrometer control and data processing. For nonfiltered biofluids,low-mass metabolites coexist with high-mass biomolecules, such aslipids, proteins, and lipoproteins; therefore, to selectively observesmall-molecule components in solutions, Carr–Purcell–Meiboom–Gill(CPMG) experiments were performed. 1D 1H pulse-sequenceCPMG experiments comprised a spectral width of 7 kHz, 32,000 datapoints, water presaturation applied over 3.5 s of relaxation delay,and a spin-echo delay of 80 ms.34 The pulsesequence used included an excitation sculpting routine for the suppressionof the water signal.35 Due to the effectof excitation sculpting on the signal height of resonances in theregion close to the water resonance,37 the metabolitesthat have resonances close to this region  were quantified using resonances from those metabolitesin other spectral regions. A weighted Fourier transform was appliedto the time domain data with a 0.5 Hz line-broadening followed bya manual phase and baseline correction in preparation for targetedprofiling analysis.NMR experimentswere carried outon a Bruker DRX600 MHz spectrometer  equippedwith a 5 mm triple-resonance 38 Briefly, the Chenomx profiler is linked to the Human MetabolomeDatabase (HMDB), containing more than 250 metabolite NMR spectralsignatures encoded at different 1H spectrometer frequencies,including 600 MHz (http://www.hmdb.ca). A comparison of the spectral data obtained for each serum samplewith the Chenomx metabolite library results in a list of compoundstogether with their respective concentrations based on the known concentrationof the added internal reference compound TSP-d4 (5.8 mM).NMR spectrawere manually phasedand baseline-corrected. The quantification of serum metabolites wasachieved using Chenomx NMR-Suite v8.0 .http://www.metaboanalyst.ca/). The performance of the PCA andPLS-DA model was evaluated using the coefficient Q2 and coefficient R,2 definingthe variance predicted and explained by the model, respectively. Theloading plot was used to identify significant metabolites responsiblefor maximum separation in the PLS-DA score plot, and these metaboliteswere ranked according to their variable influence on projection (VIP)scores. VIP scores are weighted sums of squares of the PLS-DA weights,which indicate the importance of the variable.Multivariatestatistical analysis,principal component analysis (PCA), and partial least-squares discriminantanalysis (PLS-DA) were conducted with normalized metabolomics datausing MetaboAnalyst 4.0 ,Serum concentrations of thecytokines IL-2, IL-1β, TNF-α, IFN-γ, and IL-4 weredetermined by enzyme-linked immunosorbent assay (ELISA) commercialkits ) according to themanufacturer’s instructions. All tests were performed in duplicate.The ranges of the sensitivity standard curve of the ELISA kits were31.2–1000 pg/mL for IL-2, 15.6–500 pg/mL for IL-1β,25–800 pg/mL for TNF-α, 12.5–400 pg/mL for IFN-γ,and 1.1–35 pg/mL for IL-4.Standard diluents, the captureantibody, and the detection antibody were obtained as a complete kitfor each cytokine. Standard diluent–serum samples were usedto obtain a Cedex standard curve. The samples in the multiwell platewere mixed by repeated aspirations and ejections, taking care notto scratch the inner surfaces. Freeze-dried control vials were alsoreconstituted with the most appropriate standard diluent to the sampleto gain a solution at the concentration stated on the vial. Biotinylatedanticytokines, biotinylated secondary antibody, and streptavidin-HRPwere also prepared according to the manufacturer’s protocol.The absorbance value of each well was read on a Thermo ScientificMultiskan GO spectrophotometer using 450 nm as the primary wavelengthand 620 nm as the reference wavelength (610 to 650 nm is acceptable).U test to assess theirsignificance and P value. The VIP score was calculatedusing the R package to identify the variables that discriminated betweenthe two groups. The variables with VIP scores >1 were consideredsignificantfor the analysis. The VIP score values for selected variables concerningpsoriasis patients before and after the ketogenic diet are given in 44Clinical data for 30 patients withpsoriasis before and after the ketogenic diet were statistically analyzedusing the Mann–Whitney 46 Clinical data identified by the VIP score also included severalbiochemical and clinical parameters derived from blood analysis. Inparticular, the concentrations of folic acid, vitamin B12, AST, cortisol,calcium, and direct bilirubin were higher in patients at T1 than atT0. LDL cholesterol, total cholesterol, and HOMA-IR levels were lowerin patients at T1 than at T0  and normalizationstrategy were applied. The PRICONA and normalization strategy arebased on the assumption that, since concentration differences resultin proportional variations of spectral intensities, nonproportionalchanges most likely can be attributed to the effects of the disease.The proportional variations are described by the normalization factor R, which must be calculated to compare the intensities ofthe T0 and T1 signals. When the first principal component (PC) explainsthe major part of the variance of a spectral data set, it representsthe function shape of the sum of a group of proportional peaks , and its scores represent the proportionality constants . Once the spectra are normalized, peak intensities canbe directly compared. According to the normalization procedure describedabove, signals that showed significant variations were quantified.At the same time, an opportune strategy based on the PRICONA is performedto achieve a reliable NMR metabolomic analysis. The normalizationalgorithm is based on PCA with some advantages: it allows simultaneousnormalization of data sets of spectra by identifying signals affectedby the agent  and quantitativemeasurement of variations. In this way, the differences in peak intensitiesare excluded, and the real differences do not depend on differencesin sample concentration. All extraneous sample-to-sample variationsand those within each metabolite are removed. After spectral normalization,all differences related to metabolites can be used to identify potentialcandidate biomarkers. 2 (fold change); the differences characterizedby P < 0.05 are considered significant. An analysisof the data indicated a significant increase in l-serine,dimethyl sulfone, and hydroxybutyric acid and a decrease in malonate,choline, and pyruvic acid.To quantitativelyevaluate the 1H NMR data collectedin 1D-1H-CPMG,34 were analyzedaccording to multivariate statistical analysis using MetaboAnalyst4.0.47 Multivariate analysis (MVA) wasperformed to identify the metabolic profile of psoriasis patients.The original matrix included the sera from 30 subjects with psoriasisand the sera from 30 healthy controls. The data matrix, after normalizationby sum, log transformation, and Pareto scaling, was analyzed by PLS-DA.47 In Matrices,including metabolitesand their concentrations as derived from y PLS-DA2. To minl-tryptophan, l-tyrosine, l-lysine, l-histidine, l-methionine, l-arginine, l-ornithine, and l-glutamine in psoriasis patients.The creation of separate clusters indicates a different metabolomecharacterizing patients with psoriasis and healthy controls. Thisevidence is confirmed by applying VIP score analysis 3. Accord48 The data matrix, after normalizationby sample median and log transformation, was analyzed by PCA  and PLS-DA . To obtain robuststatistical models and to calculate the P value,the Mann–Whitney U test was applied.49 For each sample, 38 metabolites were identifiedand quantified.The multivariate analysis was repeated, taking into accountpatientsbefore ketogenic diet (T0) and after the 4 week ketogenic diet (T1).Original matrices were normalized according to the concentration rangesof HMDB.l-leucine, pyruvic acid, and cholineas metabolites that significantly discriminate patients at baselinefrom those after 4 weeks of the diet. This evidence is confirmed throughthe application of VIP score analysis . Inparticular, the graph reported in l-leucine, pyruvicacid, choline, l-alanine, and acetoacetate and a lower concentrationof 3-hydroxybutyrate and acetone than after the diet.An inspection of the PLS-DA score scatter plot4A and loanalysis 5  controls against psoriasis patients before the ketogenic dietand (ii) psoriasis patients before the ketogenic diet (T0) againstpsoriasis patients after 4 weeks from the ketogenic diet (T1). P value and false discoveryrate (FDR)50 are reported to confirm thesignificance of the pathways. We also reported the most discriminatingmetabolite (P < 0.05) belonging to the pathwayand detected through the KEGG database.51To gainmeaningful insight from these data, we applied metabolicpathway analysis using MetaboAnalyst 4.0.The comparisonbetween controls and patient at T0 identified anamino acid dysmetabolism related to different pathways, in particular,arginine and proline metabolism; histidine metabolism; arginine biosynthesis;cysteine and methionine metabolism; alanine, aspartate, and glutamatemetabolism; glycine, serine, and threonine metabolism; and phenylalanine,tyrosine, and tryptophan biosynthesis. In addition, the comparisonbetween psoriasis patients at T0 and psoriasis patients at T1 highlightedthe involvement of pathways related to ketogenic diet, in particular,synthesis and degradation of ketone bodies and butanoate metabolism,which are not disregulated in comparison with previous clusters. Also,in the comparison between psoriasis patients at T0 psoriatic patientsat T1, there is an amino acid dysmetabolism related to glycine, serine,and threonine metabolism; arginine and proline metabolism; alanine,aspartate, and glutamate metabolism; and valine, leucine, and isoleucinedegradation and biosynthesis.Table S4, Supporting Information). The graph in 52 Amino acid dysmetabolism was confirmed by a change in l-leucine, l-isoleucine, and l-valine levels,which decreased after the diet,53 and bythe dysfunction of the urea cycle.53In enrichment analysis, by increasingthe number of reference metabolitesand evaluating concentrations of metabolites present in our matrix,it is possible to identify a change in the metabolome of psoriasispatients before and after the diet regimen. All dysmetabolism identifiedin the pathway analysis was confirmed by the enrichment analysis . Pathway analysis by Reactome confirmedall dysmetabolism individuated to pathway analysis by MetaboAnalyst4.0 and has shown a possible link to SLC6A14 genevariations that may be associated with obesity. Several studies haveconfirmed the correlation between obesity and psoriasis.13A more accurate analysis of the metabolic pathwaysinvolved inpsoriasis was carried out by Reactome analysisvs controls atT1 is lower than the mean concentration difference in the patients vs controls at T0. The metabolites formate, l-histidine,methionine, l-arginine, choline, l-ornithine, pyruvicacid, and l-alanine are the most discriminant.To assess the benefitsof ketogenic diet for patients with psoriasis,the average of the concentrations of metabolites with VIP score >1 was calculated in the following groups: (i) healthy controls, (ii)patients with psoriasis at T0 (before the ketogenic diet), and (iii)patients with psoriasis at T1 (after 4 weeks of diet). P = 0.04) and IL-1βin patients between T0 and T1 (P = 0.006) (55 The graph in P = 0.47), IFN-γ (P = 0.17), and IL-4 (P = 0.65).ELISAs performed onserum samples toevaluate the levels of IL-2, IL-1β, TNF-α, IFN-γ,and IL-4 showed that there were significant differences in the levelsof the cytokines IL-2 (= 0.006) 8. The si1H NMR analysis. Asa preliminary step, we identified the psoriatic patients’ metabolomicprofile and the healthy controls’ metabolomic profile.1 Based on preliminary data previously obtainedin our laboratory and for reasons related to the restrictions imposedfrom the nutritional regimen, the patients were subjected to 4 weektreatment. As the parallel evaluation of ketogenic diet effects inpsoriatic and healthy subjects was not possible, we considered thehealthy controls’ metabolomic profile as our experimental control.By following the indications of the Italian Society of Endocrinology(ISE),56 the enrollment of healthy subjectsfor the low-calorie ketogenic nutritional regimen is not recommendedas the low-calorie ketogenic diet causes alteration of the metabolicprofile in healthy subjects, with potentially harmful implicationsin blood pH equilibrium, calcium homeostasis, and lean mass balance.Therefore, we considered the therapeutic effect of ketogenic dietsby comparing the metabolomic profile of psoriatic patients at T1 withhealthy subjects’ metabolomic profile at T0.Inthe present study, we evaluated the effect of a ketogenic dieton psoriasis disease progression. Thirty psoriasis patients were subjectedto a ketogenic nutritional regimen and monitored by evaluating (i)the clinical symptoms, (ii) the blood biochemical parameters, includingIL-2, IL-1β, TNF-α, IFN-γ, and IL-4, and (iii) themetabolomic profile, as derived from P < 0.05). In particular, the PASI showed a reductionof approximately 50%.Data resultingfrom the clinical evaluation showed caloric restriction-inducedpsoriasis disease regression after 4 weeks with a significant reductionin the DLQI, PASI, VAS pain, and VAS pruritus clinical scores . The weight loss was associated with a significantincrease in ketone bodies at T1, as shown in the NMR-based metabolomicanalysis. This effect was the main physiological effect of the ketogenicdiet. The basic principle of the low-calorie ketogenic diet is tolimit the availability of carbohydrates, forcing the consumption offats as the primary energy source, with a resulting increase in fattyacids, ketone bodies, and pyruvic acid. However, weight loss and theincrease in ketone bodies were not associated with any alterationof those biochemical parameters that initially were in the averageconcentration range, thus proving the safety of the proposed dietaryintervention.The ketogenic diet resulted in weight loss at T1, correspondingto ∼10% of the initial body weight. Other significant modificationsof anthropometric measurements were waist circumference and hip circumference. A decrease in HDL was also observed due to the drop in total cholesterol. Last, the reduction in the aortomesenteric fat thickness(AMFT) proved a substantial decrease in VAT.In contrast, biochemical and clinical parameterspreviously foundin abnormal concentrations and related to carbohydrate and lipid metabolism,such as glucose, total cholesterol, LDL, Apo A1 and B, AST, γGT,insulin, and HOMA-I, returned to healthy average ranges  level byactivating the transcriptase for adipogenesis. Conversely, GH phosphorylatesPPAR and inhibits adipogenesis. As a result, reduced insulin concentrationfavors a lipolytic route with the mobilization of visceral localizedfat deposits.64 Confirming this evidence,the VIP analysis (l-isoleucine, although decreasing, do not fall into the physiologicalrange of healthy subjects, perhaps due to the short-term treatment.Earlier scientific studies have identifiedlactic acid as a possiblebiomarker of psoriasis.analysis 3 indicat65 and these concentrations were found in the ranges of the healthycontrols at T1, suggesting a rebalancing of the metabolome after theketogenic regimen.65The confirmation that the ketogenic diet induces a correction ofthe dysmetabolic condition related to psoriasis disease results fromthe NMR metabolomics study performed on psoriasis patients’blood sera. NMR metabolomics data on the patient sera collected beforethe diet indicated abnormal concentrations of metabolites that arerelated to the condition of psoriasis,l-alanine and l-leucine and alower concentration of l-glutamine were previouslyidentified as biomarkers of psoriasis.53 VIP score analysis based on the multivariate statistical analysisof the NMR metabolomics data shows a decrease in l-leucineand l-alanine and an increase in glutamine and glutamateat T1.Abnormally highconcentrations of l-arginine, l-phenylalanine, l-aspartic acid, and l-prolineconcentrations at T1 were also observable.53 An interpretation of these metabolic changes according to the pathwayand enrichment analysis (53Moreover, significant variations in analysis 6 and 7 i52NMR metabolomics data showed that choline levels were decreasedin psoriasis patients at T1 compared to T0. The decrease in cholineand nicotine concentrations is additional evidence that the regressionof psoriasis corresponds to the regression of the inflammatory process:previous data showed high choline and nicotine levels in psoriasispatients due to mast cell degranulation.67 we found decreased IL-2 and IL-1βconcentrations at T1 compared to T053 (Additionally, we measured the concentration of the cytokinesIL-4,TNF-α, INF-γ, IL-2, and IL-1β that are consideredmarkers of inflammatory status. In agreement with previous scientificstudies pointing to altered interleukin serum concentrations in psoriasispatients, to T053 3. The coTaken together, our data suggest that a low-calorie ketogenic dietcan be considered a successful strategy and therapeutic option forthe management of psoriasis. IL-2 and IL-1β, together with theconcentrations of leucine, alanine, glutamine, glutamate, and choline,can be considered promising biomarkers for the early diagnosis andcorrect prognosis of psoriasis patients.l-leucine, l-alanine, l-glutamine, l-glutamate,and choline, can be considered promising biomarkers for the earlydiagnosis and correct prognosis of psoriasis patients. The dietaryprogram is feasible, with high compliance, and safe. The main effectsdepend on reducing VAT, disrupting the inflammatory environment, andthe source of inflammatory cytokines.Our data suggest thata low-calorie ketogenic diet can be considereda successful strategy and therapeutic option for psoriasis management.IL-2 and IL-1β, together with the concentrations of


An 84-year-old man presented to the emergency department for acute vomiting associated with rotational vertigo and a sudden right sensorineural hearing loss. A left peripheral vestibular nystagmus was highlighted. The patient was afebrile, without respiratory signs or symptoms. Blood sampling at admission showed lymphopenia, thrombopenia and neutrophil polynucleosis, without elevation of C reactive protein. Cerebral magnetic resonance imaging eliminated a neurovascular origin. Vestibule, right semicircular canals and cochlear FLAIR hypersignals were highlighted, leading to the diagnosis of right labyrinthitis. A nasopharyngeal swab sampled at admission returned positive for SARS CoV2 by polymerase chain reaction. The etiologic investigation, including syphilitic and viral research, was otherwise negative. An oral corticotherapy (prednisone 70 mg daily) was introduced, followed by a progressive clinical recovery. Although acute otitis media have already been highlighted as an unusual presentation of COVID-19, radiology-proven labyrinthitis had to our knowledge, never been described to date. In mid-November 2020, at the epidemic peak of the first COVID-19 wave in France, an 84-year-old man presented to the emergency department for acute vomiting associated with rotational vertigo in the standing position. He also reported a sudden right sensorineural hearing loss. The patient did not complain of any abdominal pain or pulmonary symptoms.His medical history included arterial hypertension treated by LOSARTAN 50 mg per day and an aneurysm of the abdominal aorta and of iliac arteries, treated by endovascular stent and for which a long-term anticoagulant (WARFARINE 2 mg/day) treatment was required.On arrival, the patient was afebrile, with a blood pressure of 110/78 mm Hg, and a pulse of 84 beats/minute. The room air oxygen saturation was at 99% and a respiratory rate of 20/min without respiratory signs or symptoms. Lung auscultation was normal.3/mm3 with lymphopenia at 0.4 × 103/mm3/mm3 normal neutrophil polynucleosis count at 2.57 × 103/mm3/mm3, and platelet count at 118 × 103/mm3. C reactive protein was measured at 20 mg/L. Liver and kidney functions were both within the normal ranges.Laboratory tests at admission showed a white blood cell count at 2.8 × 10We performed a complete initial physical examination, looking for specific symptoms that would point us towards one of the possible causes of vertigo . We highBrain magnetic resonance imaging excluded neurovascular neoplastic disease. However, vestibule, semicircular canals and cochlear on the right appeared hyperintense on FLAIR images  and on dViral research including HSV, VZV, HIV, CMV, hepatitis B, hepatitis C, as well as syphilitic serology, were negative. Clinical examination excluded otitis media or meningitis as a cause of secondary suppurative bacterial labyrinthitis. Anti-nuclear antibodies, anticytoplasmic neutrophil antibodies, anti-MPO, anti-PR3, anti-MBG antibodies were also negative.® SARS-CoV-2 Assay, Hologic, Marlborough, MA, USA).Finally, a nasopharyngeal swab sampled at admission returned positive for SARS CoV2 by reverse transcription polymerase chain reaction  ,3. LabyrThe patient was admitted at the epidemic peak of COVID-19, presented a brutal asthenia one week before and regressive lymphopenia and thrombopenia at admission, suggesting that acute labyrinthitis occurred at the acute phase of COVID-19 . A compaIn the absence of respiratory symptoms, no specific COVID-19 treatment was recommended. Because of the labyrinthitis, a one-week oral corticosteroid therapy (prednisone 70 mg daily) was started, associated with physiotherapy rehabilitation, followed by a progressive clinical recovery. Corticosteroid has proven efficiency in acute vestibular vertigo . However


The past five decades have seen significant progress in our understanding of human hematopoiesis. This has in part been due to the unprecedented development of advanced technologies, which have allowed the identification and characterization of rare subsets of human hematopoietic stem and progenitor cells and their lineage trajectories from embryonic through to adult life. Additionally, surrogate in vitro and in vivo models, although not fully recapitulating human hematopoiesis, have spurred on these scientific advances. These approaches have heightened our knowledge of hematological disorders and diseases and have led to their improved diagnosis and therapies. Here, we review human hematopoiesis at each end of the age spectrum, during embryonic and fetal development and on aging, providing exemplars of recent progress in deciphering the increasingly complex cellular and molecular hematopoietic landscapes in health and disease. This review concludes by highlighting links between chronic inflammation and metabolic and epigenetic changes associated with aging and in the development of clonal hematopoiesis. Human development and aging are associated with significant changes in functional hematopoietic cell outputs from specific anatomical regions and hematopoietic tissues ,2,3,4. IAlthough temporally different and not an exact recapitulation, murine studies have often been used as a model for human hematopoiesis ,2,11. FrHere and with these inherent biases in mind, we review recent molecular advances in single-cell (sc) omics analyses of human HSPCs, set against huge historical efforts spanning more than five decades that have aimed to identify and characterize human HSCs, their lineage-committed progeny during development and aging, and that provide mechanistic insights. Additional sophisticated technological advances over these decades include the discovery of monoclonal antibodies, the development of flow cytometry and cell sorting, of enhanced in situ imaging and single-cell capture technologies for the immunophenotypic identification and isolation of specific human HSPC subsets, of single-cell barcoding, lineage tracing, fate mapping and gene editing, and of sophisticated gene regulatory and three-dimensional genome organizational analyses, coupled with surrogate models in vivo and/or in vitro to assess the function of HSCs and their progeny, or following transplantation into human recipients as exemplified in some of our own and other studies ,37,38,39Murine studies have provided considerable insight into the changes in hematopoiesis during development and aging ,40,41. TThe first wave of primitive hematopoiesis originates in the murine yolk sac from about E7 , thus giving rise to nucleated erythroid cells, macrophages and megakaryocytes ,44,45,46The third wave of murine hematopoiesis originates in the AGM region of the embryo proper, with immature or pro-HSCs emerging from distinct HE before maturing into definitive long-term repopulating (LT) HSCs (via type I and II pre-HSC) by around E11.5 ,64,65,66+CD11b+CD5+ B-1a B cells of the peritoneal and pleural cavities, and mucosa [-Sca-1+Kit+ (LSK) CD150+CD48– LT HSCs failed to regenerate B-1a lymphoid cells in adult murine transplant models, in which the adult microenvironment had been shown to be conducive to B-1a B cell reconstitution [+CD48–FLT3– single fetal liver LT HSC transplants only inefficiently reconstituted B-1a cells in adult mice. Further fate mapping studies defined a transient or developmentally restricted KSL CD50lo/- fetal HSC subset that could give rise to B-1a lymphoid cells and that differed from the previously defined LSK CD150+ LT HSC subset described above, but with these two HSC subsets possessing differential abilities to persist into adulthood [The contribution of yolk sac EMPs versus HSCs to initial lymphoid development is a matter of some debate . For exad mucosa , and pard mucosa ,88,89,90d mucosa ,87. An Ltitution ,92. Thistitution ,94, the titution ,96. Barctitution , althougdulthood ,91,98. Cdulthood  examineddulthood . Subsequdulthood . Ghosn edulthood ,96 that Although these studies support the concept of a layered immune and hematopoietic system, divergent lineage trajectories during different waves of murine hematopoiesis have added increased complexity that is not yet fully resolved.Although studies on human hematopoietic ontogeny are much more limited than in murine models, human hematopoiesis is reported to commence at approximately Carnegie Stage (CS) 7–8 of embryonic development (16–18.5 days post-conception (dpc)) in the secondary extra-embryonic yolk sac and in close association with yolk sac endothelial cells, termed the yolk sac blood islands ,102,103.+ macrophages become evident in the fetal liver rudiment, which is then seeded by CD34+CD45+ cells from CS13, and finally by definitive AGM-derived HSCs between CS13 and CS17 (27–42 dpc) [Definitive human hematopoiesis arises around CS13 27 dpc) from HE in the AGM (aorta-gonad-mesonephros) region of the embryo proper, for example from the ventral wall of the dorsal aorta, with the generation of HPCs and then importantly the appearance of immature HSCs ,111,112.–42 dpc) ,114. Her7 dpc fro–42 dpc) ,119,120.Different hematological malignancies are more prevalent in different age groups, with some subtypes originating in utero, and with pediatric hematological malignancies generally differing from those in the adult in terms of their etiology and molecular characteristics ,122. ThiSurveillance, Epidemiology, and End Results (SEER) Program of the National Cancer Institute and the American Cancer Society , and they update those described by Marcotte and colleagues [Incidence rates for hematological malignancies by age at diagnosis in the United States of America, from 2014 to 2018, are illustrated in lleagues . As indiSEER (2014–2018) (https://seer.cancer.gov) and the International Incidence of Childhood Cancer-3 (IICC-3) (2001–2010) studies reporting that the leading hematological cancers in infants and children aged < 10 years of age are leukemias, particularly acute lymphoblastic leukemia (ALL), although in adolescents  and young adults, lymphomas are more prevalent [In contrast, acute leukemia is the most common pediatric malignancy, with both the revalent .https://seer.cancer.gov) of 1.8 and 7.5 cases per 100,000 individuals per annum, respectively. Data from the MRC-UKALL clinical trials also show that a peak incidence of ALL, predominantly B cell precursor ALL, occurs at 2 to 5 years of age [As shown in s of age ,124. Hows of age .Transcriptomic and next-generation sequencing technologies, together with cytogenetic analyses, have contributed to the 2016 and current updates in progress of the World Health Organization classification of hematological malignancies and serve to highlight the heterogeneity of pediatric acute leukemias ,129,130.ETV6/RUNX1 fusion gene. A similar proportion (70–80%) of infant B ALLs (<1 years of age) carry KMT2A (MLL) gene (encoding lysine methyltransferase 2A and located on chromosome 11q23) rearrangements with at least 94 partner genes identified [KMT2A/AFF1 (MLL/AF4) fusion gene. Smaller incidences of high hyperdiploidy and t/TCF3/PBX1 gene fusions are also observed [Around 75–80% of pediatric ALL cases are B cell precursor ALL ,124,135.entified ,139,140,observed ,138,139.observed .ETV6/RUNX1 gene fusion) occurs in utero [BCR/ABL1 and TCF3/PBX1 gene fusions and KMT2A rearrangements, including the t/KMT2A/AFF1 fusion gene [ETV6-RUNX1 gene fusions in approximately 1 in 10,000 B lymphoid lineage cells  without overt B cell precursor ALL developing in the vast majority of these children, and with predisposing factors for development of B cell precursor ALL post-natally, including environmental factors and additional mutations [Because of its prevalence compared to T ALL and AML, there has been a particular interest in understanding the origin and development of pediatric B cell precursor ALL. Risk factors have recently been reviewed in detail elsewhere ,142,143.in utero . This coin utero . Other pion gene ,143. Simion gene ,147,148.utations ,147,148.utations ,124,143.For childhood B cell precursor ALL, various hypotheses have been proposed as causal factors in disease progression after birth, including population mixing, infection and delayed infection ,151,152.KMT2A rearrangements (particularly KMT2A/AFF1 gene fusions) in utero appear to be sufficient for the onset of leukemia before or shortly after birth [KMT2A rearrangements [FLT3, as well as abnormal DNA methylation patterns [KMT2A rearrangements in utero always develop overt infant ALL remains to be fully determined.In infants with B ALL (<1 year of age), initial er birth ,139, alter birth . Overallngements ,157, altpatterns ,160,161.More recently, there has been a surge of interest in using increasingly advanced omics technologies to define more clearly human hematopoiesis and hematopoietic lineage trajectories during embryonic and fetal development. As the initiating events leading to infant and childhood B ALL are thought to mostly originate in utero, the sequence of human B lymphoid development during human embryonic, fetal and early post-natal life has generated intense interest in the quest to define the cell of origin of these childhood leukemias. Progress in defining human fetal B cell development has recently been reviewed , and thi+CD10+CD19+ Pro B-progenitors in adult human bone marrow and CD34+CD10-CD19+ PrePro B-progenitors in UCB and second trimester human liver [+CD10−CD34+ PrePro B-progenitors were the earliest B-lymphoid restricted progenitors  detected and were positioned upstream of CD19+CD10+CD34+ Pro B-progenitors and downstream from fetal specific Lin-CD34+CD127++/IL-7Rα++CD10–CD19– ELPs with the ability to give rise to B, T, NK and some myeloid cells. O’Byrne et al. [+ cells in the latter part of the second trimester. While complete VH-DH-JH rearrangements occur in Pro B-progenitors, only partial DH-JH IgH rearrangements occur in fetal ELP and PrePro B-progenitors [Based on the detection of phenotypically defined HSC, MPP and lymphoid primed multipotent progenitors (LMPP), and of oligopotent fetal-specific early lymphoid progenitor (ELP) cells ,163,164,an liver ,169,170,an liver  comprehee et al.  used matgenitors . Notablygenitors . These sgenitors ,119.+CD27+CD43+CD70– cells in UCB and adult peripheral blood [+CD38hi plasmablast and preplasmablast precursors, it remains possible that the human fetal B-1 B cell frequency has been over estimated [+CD20+CD27+CD43+CD38lo/int, while lacking CD3, CD4 and CD8 T cell markers, making them distinct from B-2 lymphoid cells, which are predominantly found in adult tissues [Other studies have examined and set out to better characterize human B-1 and B-2 lymphoid development. As indicated earlier, innate-like B-1 (B-1a and B-1b) and conventional B-2 cells constitute two main branches of the murine B cell population. During embryonic and fetal development, murine B-1a subsets are proposed to arise from multiple sites including yolk sac, P-Sp/AGM and fetal liver from HSC independent or dependent precursors, and to a lesser degree from adult bone marrow HSCs ,87. Althal blood , and in al blood . As thisstimated ,177,178. tissues ,177,178. tissues .ETV6/RUNX, TCF3/PBX1, CRLF2 and ERG B ALLs are more likely to originate from B-1-like cells, while BCR/ABL1, hyperdiploid, and KTM2A rearranged B ALLs derive from B-2-like B cells [KMT2A/AFF1) and childhood B cell precursor ALL (ETV6/RUNX1) yet are predicted from the studies above to arise in different B lymphoid lineages, the former in conventional B 2-like and the latter in innate B 1-like B cell lineages.There have been suggestions that at least a proportion of infant and childhood B ALLs arise from B-1 B cells or their precursors . Given t B cells ,181. The B cells ,181. Two B cells  are of sKMT2A/AFF1 infant B ALL, fetal PrePro B-progenitors express several genes implicated in infant ALL, while KMT2A-rearranged B ALL clones, similar to PrePro B progenitors, are CD10- and carry partial DH-JH rearrangements [KMT2A rearranged infant B ALL [KMT2A rearranged infant and childhood ALLs originate in utero  [KMT2A (MLL)-rearranged infant ALL is initiated and maintained by co-operation between fetal specific gene expression programs or environment and the rearranged KMT2A gene. Using KMT2A/AFF1 (MLL/AF4), the most common infant ALL rearrangement as a model, their studies demonstrated that KMT2A/AFF1 infant ALL did maintain a designated fetal specific gene expression profile, while KMT2A/AFF1 childhood ALL did not [KMT2A/AFF1 translocation; they were shown to drive fetal specific and infant ALL molecular programs and to recapitulate clinical characteristics of the human disease (including treatment resistance and CNS disease) in an NSG xenograft model of infant ALL [In relation to ngements . These ont B ALL . Conside course) . Rice et course)  hypothes did not ,183. Fur- and CD127+ (IL-7Rα+) ELPs generate human lymphoid cells, with both CD127 subsets arising independently from multipotent CD34hiCD45RA+ Lympho-Mono-Dendritic cell progenitors (LMDPs) [+ ELPs generated NK, ILC and B cells, but not T cell subsets, CD127- ELPs gave rise to T cells, ILCs, and NK and marginal zone B cells [+ ELPs closely resemble the fetal bone marrow ELPs that are putative PrePro B cell precursors [KMT2A rearranged infant B ALL with previously published fetal human bone marrow transcriptomes [KMT2A rearranged infant B ALL resembles a fetal ELP state as defined above [KMT2A/AFF1 fusion gene, or whether the leukemic/preleukemic event occurs in another progenitor type that then arrests at the ELP stage, and, if so, if this has any bearing on the development of infant versus childhood ALL. In this respect, assessment of the phylogenetic origin of a rare case of lineage switching from KMT2A rearranged infant B ALL to KMT2A rearranged childhood AML has led these same researchers to suggest that, in this specific case, the KMT2A rearrangement may have occurred before gastrulation and hematopoietic specification [Other studies, which have included an analysis of human fetal bone marrow (13–29 pcw) ELP subsets, have proposed a “two-family” model for lymphopoiesis, in which CD127 (LMDPs) . While C B cells . Except  B cells , the forecursors , perhapsecursors  comparinriptomes  have coned above ,184. Givfication .ETV6/RUNX1 B cell precursor ALL, Boiers and colleagues [-IL-7R+/CD127+ (Lin-CD19-CD34+CD38+CD45RA+IL-7R+KIT+) cells, which during ontogeny (CS17 to CS20), transition from a myeloid-primed to a lymphoid-primed program, as the earliest human B lymphoid progenitors. Additionally, a proportion of CD19-IL-7R+/CD127+ progenitor showed evidence of DH-JH rearrangements and myeloid  cells in vitro [-IL-7R+ progenitors were also identified when human iPS cells were differentiated in an OP9/MS5 co-culture system, with day 10 differentiated human iPS cells resembling CS17 fetal liver CD19-IL-7R+ progenitors and day 31 differentiated human iPS cells resembling CS17 and CS20 fetal liver CD19-IL-7R+ progenitors (mixed myeloid and lymphomyeloid primed) [ETV6/RUNX1 fusion gene at physiological levels in, and differentiation of, these human iPS cells led to an expansion of the CD19-IL-7R+ progenitor cells, a partial B lymphoid lineage commitment block, and the generation of proB cells aberrantly co-expressing myeloid gene signatures and potential, and thus potentially recapitulating the ETV6/RUNX1 preleukemic state [-IL-7R+ human B lymphoid progenitors to B-1 and B-2 B cells is yet to be established, although it has been reported that fetal liver and adult human pro B cells that express both CD27 (IL-7Rα) and LIN28B preferentially mature to a B1-like B cells [In other studies involving lleagues  identifiin vitro . Transcr primed) ,187. Subic state . The rel B cells .Thus, these studies, while incomplete and still ongoing, have substantially increased our understanding of human fetal B lymphoid development and progenitor cells, and the development of pediatric B ALL.Ras signaling pathways [Myeloid leukemias occurring neonatally or in early childhood include infant AML , juvenilpathways ,190, andpathways . As wellpathways ,141. SinKMT2A/MLLT3, t/PML/RARA, t/RUNX1/RUNX1T1, and inv(16)/t/CBFB/MYH11 /KMT2A/MLLT3 (MLL/AF9) fusion gene, the incidence of which declines during childhood and adolescence [PML/RARA, t/RUNX1/RUNX1T1, inv(16)/t/CBFB/MYH11, and trisomy 8 karyotypic subtypes [GATA2, FLT3 and CBL, and recurrent mutations in KRAS, NRAS, KIT, WT1 and MYC-ITD [KMT2A/AFF1 fusion gene, which as described above, commonly presents as pediatric B cell precursor ALL, may also demonstrate an infant B/myeloid mixed phenotype or relapse with the original clone switching to AML [Over 20 different subtypes of AML have been defined based on the 2016 revision of the WHO classification of myeloid neoplasms and acute leukemias , with ape et al.  and othee et al. ,192,193)lescence . Of furtsubtypes . Both cosubtypes ,127. Gen MYC-ITD . Lineageg to AML ,198,199.PML/RARA and t/RUNX1/RUNX1T1 fusion genes in neonatal blood spots or UCB of children or adolescents subsequently developing AML, indicating that these as a minimum can be initiated in utero [https://clinicaltrials.gov/ct2/show/NCT05014165; accessed 28 November 2021). In utero, preleukemic events would of course also include such genetic or inherited predispositions as Noonan and CBL Syndromes and Neurofibromatosis type-1 (NF-1), which place such children at higher risk of developing, although not necessarily exclusively, myeloid leukemias [RAS pathway genes, are predisposed to JMML [RAS pathway genes , not all develop JMML. About 5% develop a transient myeloproliferative disorder, which spontaneously resolves, although in some cases, this will progress to JMML [RAS mutations in PTPN11, NRAS and KRAS, who account for a significant proportion of JMML cases [RAS pathway mutations  in 38% of children (n = 34) without Noonan Syndrome but presenting with JMML at a median age of 1.5 years [Although significantly fewer than for pediatric ALL, backtracking studies have detected t/in utero ,202,203.eukemias ,204,205..5 years . These c.5 years . This suKMT2A/MLLT3 fusion gene on AML development [KMT2A/MLLT3 AML xenograft model, using human UCB CD34+ HSPCs and CRISPR/Cas9 genome editing technologies, which also reported that the developmental age and the genetic background of the human CD34+ HSPCs, as well as the microenvironmental niche in surrogate murine models of hematopoiesis influenced AML progression [KMT2A/MLLT3 fusion gene prior to transplantation into NS-SGM3 [KMT2A/MLLT3 fusion gene generated both AML and ALL in NSG mice, whereas similarly transduced adult bone marrow HSPCs generated LT hematopoietic engraftment with a myeloid bias, which did not progress to AML. More recently, using a retroviral based KMT2A/MLLT3 model derived from human UCB CD34+ HSPCs, in which the genetic background of the UCB donor was defined by next generation sequencing prior to the introduction of the KMT2A/MLLT3 fusion gene, Milan et al. [KMT2A/MLLT3 fusion gene require additional signals, possibly from the bone marrow niche, for leukemic transformation. Consistent with this, Hyrenius-Wittsten et al. [KMT2A/MLLT3 and N676KFLT3 in human CD34+ UCB HSPCs mainly resulted in the development of AML in NSG mice, concluding that constitutively active signaling mutations within the transduced cell could replace exogenous factors and promote AML.Given its importance in infant AML, murine and human model systems, combined with single-cell omic assays ,208,209,elopment . These igression . Similar NS-SGM3 , Horton  NS-SGM3  showed tn et al.  also conn et al.  demonstrKMT2A/MLLT3 AML, as well as defining the individual functions of the KMT2A and MLLT3 in normal human hematopoiesis. As an example, MLLT3 is a key regulator of human HSC self-renewal and engraftment, potentially acting as an HSC maintenance factor by protecting the stemness program as HSC divide, and a critical regulator of early human erythroid and megakaryocyte fate [HOX gene transcriptional activation, while the KMT2A/MLLT3 fusion protein forms part of the disruptor of telomere silencing 1-like  complex (DOTCOM), the effects of which are reviewed elsewhere [KMT2A/MLLT3 AML may be initiated in utero [KMT2A/MLLT3 preleukemic event occurs and determine if additional mutations and/or the fetal or early neonatal hematopoietic microenvironment differentially influence progression to AML in infants as opposed to older children. Such an approach has recently been taken by one of us in the context of infant KMT2A/AFF1-driven ALL and AML [Importantly, significant progress is being made in understanding the molecular basis for the initiation and progression of pediatric yte fate ,217,218.lsewhere ,220,221.in utero , it woul and AML ,222,223.RAS pathway , with approximately 35% being somatic PTPN11 (gain of SHP-2 function) exon 3 or 13 mutations [PTPN11, NRAS and KRAS subgroups and with a median age of 2.2 years) in immunodeficient NSG and NSG-SGM3 mice. This was further investigated by Louka et al. [−CD34+CD38−CD90+CD45RA− HSC, Lin−CD34+CD38+CD123+ CD45RA+ GMP and as a novel JMML specific double-positive Lin−CD34+CD38−CD90+CD45RA+ myeloid progenitor subset) into NSG mice, demonstrating that each HSPC subset propagated JMML in vivo. Single-cell index sorting of Lin−CD34+ HSPCs and colony genotyping revealed that the JMML HSPC compartment was clonally heterogeneous, containing both clonally dominant RAS pathway mutations and subclones bearing other mutations , with all somatic mutations being backtracked to the phenotypically defined HSC subset [RAS pathway mutations followed both linear and branching patterns of clonal evolution [HMGA2, CNN3 and VNN2, and overexpression of HOPX, which encodes a non-DNA binding homeodomain protein involved in primitive hematopoiesis has been demonstrated in JMML HSCs, supporting a putative JMML embryonic or fetal origin [HOPX, SPINX2, CLERC9A) was present in both JMML/HSC and JMML myeloid progenitors, with gene regulatory networks/regulons  in these JMML subsets being reminiscent of fetal HSC/MPP [JMML, which accounts for 1% of pediatric leukemias, has a median age of onset of 2 years ,224,225.utations ,228,229.utations ,228,229.utations ,235,236 utations ,238,239.l origin ,241. Add HSC/MPP .As described above, sequential waves of human HPCs and HSCs are generated from such tissues as the yolk sac and the AGM, coupled with their expansion in the fetal liver in order to meet the needs of the developing embryo and fetus. The shift of this hematopoiesis from the fetal liver to the fetal bone marrow from 10–12 pcw represents the first step toward establishing the bone marrow as the predominant site for adult human hematopoiesis. Recently, single-cell multi-omics of human fetal liver and fetal, pediatric and adult bone marrow have highlighted the significant changes in the composition and function of HSPCs from fetal to adult life ,116. Nothttps://www.who.int; accessed 28 November 2021). The aging process coincides with an increase in HSC numbers and adipogenesis in adult human bone marrow; this is accompanied by a loss of clonal HSC functional heterogeneity, decreased regenerative capacity and a reduction in lymphopoiesis, characterized by a shift from lymphopoiesis to myelopoiesis [At the other end of the spectrum, the aging hematopoietic system of the adult bone marrow not only shows increased susceptibility to certain hematopoietic disorders but is also linked to the development of other diseases that include stroke and cardiovascular diseases ,243. Theopoiesis ,249,250.opoiesis ,251.DNMT3A, PPM1D and V617FJAK2 originated either in utero or in childhood [DNMT3A mutations ranged from 8 pcw to 7.6 years of age, and of the V617FJAK2 mutations from at least 33 pcw to 10.8 years of age, while a PPM1D mutation was estimated to occur at 5.8 years of age [V617FJAK2, the MPNs were diagnosed 11 to 54 years later [V617FJAK2 mutation occurred decades before MPN diagnosis , with the HSC carrying the mutation having a selective fitness or growth advantage.Somatic mutations are prevalent in the highly proliferative hematopoietic system and accumulate during development (as indicated earlier) and, to a greater extent, with aging ,252,253.hildhood . Althougs of age . In the rs later . Lineagers later  additionDNMT3A , TET2 (ten-eleven translocation dioxygenase 2) and ASXL1  [TET2 loss of function also skewing hematopoietic differentiation toward the myelomonocytic lineage as reviewed recently elsewhere [In adults, HSPC-associated somatic mutations can lead to clonal hematopoiesis, which increases with age, occurring in more than 10% and possibly as many as 15–20% of individuals aged 70 years or older, and defined as somatic genomic changes in cells of the hematopoietic lineage of individuals with no evidence of hematological malignancy ,257,258.lator 1) . Each oflsewhere . Similarlsewhere . Follow lsewhere . M-CHIP lsewhere . Previoulsewhere ,257. Addlsewhere . Other slsewhere ,271,272.Dicer1 gene was deleted in murine osteoprogenitor cells [+ HSPC proliferation in long term in vitro cultures has also been reported [Although not the focus of this manuscript, alterations in the aged bone marrow microenvironment  may lead to changes  in HSPCs and support their evolution to hematological malignancies. The studies, which support these conclusions ,276,277,or cells . Culturior cells . In contor cells ,282. Stuor cells , using aor cells , while areported . More rereported  have demreported ,288,289,reported  identifireported . A variereported .DNMT3A, an epigenetic writer, and TET2, an epigenetic eraser, both of which, as indicated above, play significant roles in CHIP development.A cell autonomous increase in human adult bone marrow HSC self-renewal has been considered as the main driving force for clonal hematopoiesis, rather than neutral genetic drift . AlthougMetabolism controls HSPC function, in part by providing energy (ATP) and tricarboxylic acid (TCA) cycle metabolites ,294. ATPDNMT3A and TET2 are early drivers of age-related clonal hematopoiesis. DNMT3A, one of three mammalian DNA methyltransferases, catalyzes the de novo methylation of DNA principally by converting cytosine residues to 5-methylcytosine; its major methyl donor is S-adenylmethionine, a product of mitochondrial one-carbon metabolism [Dnmt3a-/- HSCs exhibit extensive self-renewal ability as well as a reduced ability to differentiate [Dnmt3a-/- (but not Dnmt3a+/+) HSCs and MPPs by promoting widespread changes in global methylation and a reduction in stress-related apoptosis [DNMT3A mutations are spread across the gene, with a significant proportion (74%) being loss-of-function mutations, which in 50% of cases, exhibit a reduction in DNMT3A protein stability and increased protein degradation that correlates with enhanced clonal expansion [The role of different classes of epigenetic regulators in hematopoiesis and age-related clonal hematopoiesis has been recently reviewed ,313,314.tabolism ,315. Surrentiate ,318,319.poptosis ,321. Recxpansion .++ and ascorbate, and is inhibited by 2-hydroxyglutarate, fumarate and succinate, with increases in glycolysis limiting α-ketoglutarate availability and with upregulation of oxidative phosphorylation enhancing α-ketoglutarate levels [Tet2-/- murine HSCs demonstrate significant changes to DNA methylation of lineage specific transcription factor binding motifs, with resultant disruption to transcriptional priming [TET2 loss-of-function mutations also increase secretion of proinflammatory cytokines by mutant TET2 hematopoietic cells, which can alter HSPC fitness and enhance clonal expansion [Tet2 is deleted in hematopoietic cells, allow bacteria to enter the blood stream provoking a microbial-dependent pro-inflammatory response and resulting in further survival and proliferative advantages to the Tet2-/- HSPC clones [DNMT3A and TET2 loss-of-function hematological clones in older individuals.In contrast to DNMT3A, TET2 dioxygenase demethylates DNA by oxidizing 5-methylcytosine to 5-hydroxy-methylcytosine . Fle levels ,294,302.e levels , while s priming . TET2 loxpansion . In thisC clones ,327. ThuC clones  and metaIn this review, we have attached a special importance to more fully understanding normal human hematopoiesis at each end of the age spectrum, and to highlight evidence for certain premalignant chromosomal rearrangements/translocations or driver mutations arising in utero or in early post-natal life, yet with the malignant hematological disease not manifesting itself clinically for weeks to many decades later. We have further sought to describe the importance of better defining hematopoietic lineage hierarchies and trajectories in tissues such as the human fetal liver and fetal bone marrow to not only identify both the cell of origin in which driver mutations and chromosomal alterations initially arise or are expressed but also to more fully decipher the mechanisms that subsequently promote the development of hematopoietic malignancies in the few selected individuals who will eventually be diagnosed with these diseases. For example, if the initiating mutations for clonal hematopoiesis and CHIP arise in utero and are expressed in a fetal HSC subset that is more highly proliferative and possesses higher self-renewal capacity and multipotentiality than do the HSCs of aging adult bone marrow, then these mutated clonal HSCs may have a significant competitive growth advantage over the quiescent HSCs that normally reside in post-natal and aging bone marrow. There is thus a need to more fully understand the mechanisms that provide such cells with a competitive advantage for progression to CHIP, and whether this relates to a genetic predisposition, the specific subtype of initiating mutation, further gene mutations, or differences in HSC fate determination that are controlled epigenetically, by inflammatory states or by the nutrient signaling and sensing hubs of the mitochondria and lysosomes. Of further importance has been the increasing complexity of the chromosomal alterations or mutations that have been determined by using advanced high throughput sequencing platforms and which have led to the identification and more accurate classification and risk stratification of many malignant hematopoietic subtypes. It is clear that our understanding of human hematopoiesis at each end of the age spectrum is much more limited than it is in murine model systems. A detailed analysis of the broader experimental approaches to defining the effects of aging on hematopoiesis and of the many unknowns still facing us is presented in  and is n


Gallinula melanops, a species of Rallidae found in Brazil, is among the few species studied cytogenetically, but only with conventional staining and repetitive DNA mapping, showing 2n=80. Thus, in order to understand the karyotypic evolution and phylogeny of this group, the present study aimed to analyze the karyotype of G. melanops by classical and molecular cytogenetics, comparing the results with other species of Gruiformes. The results show that G. melanops has the same chromosome rearrangements as described in Gallinula chloropus (Clade Fulica), including fission of ancestral chromosomes 4 and 5 of Gallus gallus (GGA), beyond the fusion between two of segments resultants of the GGA4/GGA5, also fusions between the chromosomes GGA6/GGA7. Thus, despite the fact that some authors have suggested the inclusion of G. melanops in genus Porphyriops, our molecular cytogenetic results confirm its place in the Gallinula genus.Although Rallidae is the most diverse family within Gruiformes, there is little information concerning the karyotype of the species in this group. In fact,  Gruiformes is an avian order showing great heterogeneity of habits, habitats and morphology and a wide geographic distribution . BecauseWithin the core Gruiformes, Rallidae is the family with the highest number of species, around 152, distributed in 33 to 40 genera, comprising 85% of the order diversity . The phyGallinula melanops), are distributed from northeastern to southern Brazil, with occurrences in Bolivia, Paraguay and Argentina southward (G. melanops (2n=80) is one of the six Rallidae species with known karyotypes, together with Fulica atra (2n=92), Gallinula chloropus (2n=78), Aramides cajaneus (2n=78), Porzana albicollis (2n=72), and Porphyrio porphyrio (2n=80) , Aramides  and Porphyrio (genus Porphyrio) have been analyzed by classical or molecular cytogenetics.Rallidae shows a huge geographic distribution and taxonomic complexity . Severalouthward . G. mela (2n=80) . Hence,  (2n=80) , only spGallus gallus-GGA), stone-curlew (Burhinus oedicnemus-BOE), white hawk , griffon vulture (Gyps fulvus-GFU) and eared dove (Zenaida auriculata-ZAU) , following TM -II 5% and penicillin (PNS) 1% then incubated at 37 ºC. Afterwards, metaphase arrest was obtained by adding colcemid  followed by incubation for 1 hour at 37 ºC, and hypotonic solution treatment  for 15 minutes. Finally, the suspensions were fixed using Carnoy’s fixative methanol: acetic acid (3:1 v/v). The experiments followed ethical protocols approved by the Ethics Committee nº018/2014 (UNIPAMPA) and SISBIO: 44173-3.Fibroblast cultures obtained from wing skin biopsies of five female specimens of Gallus gallus (GGA) chromosome probes from 1 to 14 obtained by flow-sorting and labeled with biotin-16-dUTP or digoxigenin-11-dUTP  by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR) . At least 10 metaphases were analyzed to confirm the hybridizations signals. Final editing of images was performed using Adobe Photoshop CC software. For chromosomal evolution inferences, we used chromosome painting data from Fulica atra, Gallinula chloropus and Aramides cajaneus (DOP-PCR)  were hybcajaneus , also thcajaneus , to clarG. gallus or Taeniopygia guttata (Zebra finch) for the microchromosomes GGA17-28, following The bacterial artificial chromosomes (BACs) were selected from the genome library of Gallinula melanops (2n=80) is composed of 11 pairs of macrochromosomes, including the Z and W, and 29 pairs of microchromosomes, which corroborates previous findings , analyzed herein  can be explained by extensive fission of microchromosomes.Furthermore, it was possible to confirm that each chicken or Zebra finch BAC was conserved as a distinct element in each microchromosome of melanops . Microchmelanops . DespiteG. melanops should be included in the genus Porphyiro. However, the chromosome morphology data of P. porphyrio, the only species from this genus with a known karyotype, do not show many similarities with the karyotype of G. melanops. For example, in G. melanops the macrochromosomes 1-5 are biarmed and 6-10 are telocentrics, whereas in P. porphyrio chromosomes 1-7 are biarmed and 8-10 are acrocentrics , based on mitochondrial DNA (mtDNA), suggest that this group is paraphyletic , performed by G. melanops (2n=80) was recovered as the sister clade to G. chloropus (2n=78). However, these species share the same chromosome rearrangements, which could indicate that their common ancestor would contain the fission into GGA4 and GGA5, aside from the association between GGA4/GGA5 and GGA6/GGA7.The phylogenetic relationships within the ‘Clade phyletic . In the G. melanops in a more basal position within Clade Fulica   , consisthloropus .Aramides would be sister group to clade Fulica, despite the species A. cajaneus (Clade Aramides) not showing the fission into GGA4 and GGA5, or the fusions between GGA4/GGA5 and GGA6/GGA7 . The results illustrate the value of comparative chromosome painting and BAC mapping in phylogenetic studies.In conclusion, the comparative chromosome painting reveals that


Luteolin (LUTN), well-known as an active phytochemical agent, acts as a strong antioxidant. This research was designed to investigate whether LUTN exerts protective effects against AFB1-induced hepatotoxicity and explore the possible molecular mechanism in mice. A total of forty-eight mice were randomly allocated following four treatment groups (n = 12): Group 1, physiological saline (CON). Group 2, treated with 0.75 mg/kg BW aflatoxin B1 (AFB1). Group 3, treated with 50 mg/kg BW luteolin (LUTN), and Group 4, treated with 0.75 mg/kg BW aflatoxin B1 + 50 mg/kg BW luteolin (AFB1 + LUTN). Our findings revealed that LUTN treatment significantly alleviated growth retardation and rescued liver injury by relieving the pathological and serum biochemical alterations  under AFB1 exposure. LUTN ameliorated AFB1-induced oxidative stress by scavenging ROS and MDA accumulation and boosting the capacity of the antioxidant enzyme . Moreover, LUTN treatment considerably attenuates the AFB1-induced apoptosis in mouse liver, as demonstrated by declined apoptotic cells percentage, decreased Bax, Cyt-c, caspase-3 and caspase-9 transcription and protein with increased Bcl-2 expression. Notably, administration of LUTN up-regulated the Nrf2 and its associated downstream molecules  at mRNA and protein levels under AFB1 exposure. Our results indicated that LUTN effectively alleviated AFB1-induced liver injury, and the underlying mechanisms were associated with the activation of the Nrf2 signaling pathway. Taken together, LUTN may serve as a potential mitigator against AFB1-induced liver injury and could be helpful for the development of novel treatment to combat liver diseases in humans and/or animals.Aflatoxin B Aspergillus flavus and Aspergillus parasiticus and usually found in agricultural environments and food commodities [1 (AFB1) is the most potent liver carcinogen among the aflatoxins, and it has been categorized as a Class I carcinogen by the International Agency for Research on Cancer (IARC) [1 is reported to cause severe health issues, including growth retardation, hepatotoxicity, neurotoxicity, teratogenicity, mutagenicity and immunotoxicity in humans and animals [1 toxicity, a major metabolizing and detoxifying organ in the body [1 in the liver [1 toxicity. Natural active biological compounds derived from plants have recently received much attention due to their low toxicity. Herbal metabolites are effective alternatives for tackling the hazardous effects of AFB1 [Aflatoxins are one of the most dangerous mycotoxins, produced mainly by modities . Accordimodities ,3. Curremodities ,5. Aflatr (IARC) . Additio animals ,8,9. Thethe body . Previouhe liver . ROS ovehe liver . Therefo of AFB1 ,14.1-induced liver damage in broiler chicks has been associated with dysregulating the Nrf2 signaling pathway [1-induced liver damage and underlying mechanisms have still not been explored. Therefore, the current research was designed to investigate whether LUTN exerts protective effects against AFB1-induced hepatotoxicity and explore the possible molecular mechanism in mice. Presumably, this is the first study to highlight the protective role of luteolin against AFB1-induced hepatic damage in mice.Flavonoids are bioactive compounds primarily present in plants with a wide range of pharmacological and health-promoting properties . Luteoli pathway . Therefo pathway ,25. Alth pathway . Moreove pathway ,27. Howe1  used in the present study was supplied by Sigma-Aldrich . The ELISA assay kits for malondialdehyde , reactive oxygen species , glutathione peroxidase , total antioxidant capacity , catalase  and total superoxide dismutase  were from . Apoptosis detection kit annexin V-FITC/PI (#KGA-108) was supplied by . A test kit for the bicinchoninic acid assay was provided by Thermo Fisher Scientific . Chemiluminescence WesternBright ECL substrate kit (# ab65623) was supplied by . The primary antibodies, heme oxygenase-1 , glutamate-cysteine ligase catalytic subunit , quinone oxidoreductase 1 , nuclear factor erythroid 2-related factor 2 , Bcl-2-associated X protein , caspase-3 (#A2156) and B-cell lymphoma 2  were procured from . Secondary antibodies, anti-mouse IgG (#4410), anti-rabbit IgG (#4414), and β-actin (#3700), were obtained from .The luteolin  was bought from . Aflatoxin BFour weeks old male C57BL/6 mice were bought from Wuhan University . As an adoption period, mice were housed in separate cages and given an appropriate environment one week before the start of the experiment. Standard feed pellets and freshwater were accessible to the animal’s ad libitum. All mice were housed under laboratory conditions, light-dark period (12 h light/12 h dark), relative humidity of 45–60%, and the temperature of 22 ± 2 °C. Huazhong Agricultural University permitted the animal experiments under the Laboratory Animals Care and Ethics Committee (Permit No. HZAUMO-2018-07). Besides this, the health of experimental mice was closely monitored, and necessary measures were taken to assure the maximum welfare of the animals.n = 12):Group 1, (CON) received physiological saline.1), treated with 0.75 mg/kg BW aflatoxin B1.Group 2, , treated with 50 mg/kg BW luteolin.1 + Luteolin), treated with 0.75 mg/kg BW aflatoxin B1 + 50 mg/kg BW luteolin.Group 4,  was used to quantify the amounts of globulin and albumin, as well as GGT, ALP, AST, and ALT in serum samples following the manufacturer’s suggested protocol.The H&E staining was conducted as reported in our prior study . Liver sThe 10% tissue homogenates were prepared from collected liver samples following our previously reported procedure . The bicSingle-cell suspensions were prepared to detect the apoptosis rate in the liver of mice following our previously described procedure ,32. Brie−ΔΔCt formula was used to quantify with the β-actin as a reference gene [The transcription levels of pertaining genes used in the present study were determined by qRT-PCR following the method previously mentioned in our study . The prince gene . The pronce gene ,35. The p-value < 0.05.The experimental data were analyzed for significance by using SPSS  software. A one-way analysis of variance (ANOVA) was used for statistical analysis, followed by the least significant difference (LSD) test. The results are presented as mean ± SD. The significance level of data was set at 1 (AFB1) are depicted in 1 recorded the (p < 0.05) lowest ADG and ADFI in the comparison of CON, LUTN and AFB1 + LUTN groups, respectively. Contrastingly, LUTN therapy considerably improved growth performance in mice, as evidenced by increased ADG and ADFI (p < 0.05) compared to the AFB1 group. Furthermore, LUTN treatment significantly reduced liver coefficient (p < 0.05) increased by AFB1 (1 on the growth of mice.The protective effects of luteolin (LUTN) on the growth of mice exposed to aflatoxin B by AFB1 C. These 1 are summarized in 1 exposure considerably (p < 0.05) increased serum liver enzymes activities such as ALP, ALT, AST, and GGT, while decreased globulin and albumin content. On the other hand, LUTN treatment considerably reversed AFB1-induced alterations on the biochemical profile of mice. Moreover, histological analysis revealed that AFB1 exposure damaged the liver structure of mice, as evident by microvesicular appearance of the lipid droplets with small and large area of blood infiltration were observed in the AFB1 treated group (p < 0.05) ameliorated and restored liver injury induced by AFB1, indicating that LUTN could protect the liver from AFB1-induced hepatotoxicity.The effects of LUTN treatment on the biochemical profile of mice exposed to AFBed group B. Striki1 revealed a significant (p < 0.05) increase in ROS and MDA levels. At the same time, the activities of T-AOC, CAT, GSH-Px and T-SOD were significantly reduced as compared to the CON, LUTN and AFB1 + LUTN groups, respectively. Contrastingly, LUTN treatment significantly (p < 0.05) reversed AFB1-induced alterations in the oxidative stress markers and antioxidant variables in the liver of mice, as evident by decreased ROS and MDA levels by 38% and 20%, respectively, while increased antioxidant enzyme activities of CAT, T-SOD, GSH-Px, and T-AOC by 36.85, 30.27, 27.26, and 40% respectively, as compared to the AFB1 treated group  the proportion of apoptotic cells relative to the CON, LUTN, and AFB1 + LUTN groups, respectively. However, LUTN administration dramatically reduced (p < 0.05) the proportion of apoptotic cells in comparison to the AFB1 group.Apoptosis rate in the hepatocytes was measured by flow cytometry . Apoptosp < 0.05) increased, while Bcl-2 was decreased under AFB1 exposure. Contrastingly, LUTN administration significantly reversed the transcripts levels of these genes as comparative to the AFB1 exposed group. Moreover, we investigated the protein expression of Bcl-2, Bax and caspase-3 in the liver tissue of mice by western blotting  down-regulated, while Bax and caspase-3 protein expression was (p < 0.05) up-regulated as compared to the CON group. Conversely, LUTN treatment significantly alleviated the alterations in the Bcl-2, Bax and caspase-3 protein levels induced by AFB1.Mitochondrial apoptosis-associated transcription and protein expressions were detected by qRT-PCR and western blotting. As depicted in blotting F,G. Inte1 is associated with Nrf2 signaling pathway activation, the transcript levels and protein expression of Nrf2 and downstream targets were detected. As depicted in 1 exposed group showed a significant (p < 0.05) decrease in the gene expression of Nrf2, HO-1, GCLC, NQO1 and SOD1 compared to the CON and other experimental groups. In contrast, LUTN treatment considerably improved the transcript expressions of Nrf2, HO-1, GCLC, NQO1 and SOD1 altered by AFB1 (p < 0.05). Further, the protein expression of Nrf2 and its target genes were detected by western blotting and revealed the same tendency (p < 0.05) down-regulated in the AFB1 challenged group. However, LUTN treatment considerably up-regulated the Nrf2, NQO1, HO-1 and GCLC protein expressions as down-regulated by AFB1 (p < 0.05).To confirm our hypothesis that LUTN promotes the antioxidant capacity and alleviates hepatotoxicity in mice induced by AFBtendency F,G. Simi1 exposure decreased ADG and ADFI in mice. The observed growth retardation may have resulted from anorexia, suppression of lipogenesis, and protein synthesis induced by AFB1 [1 poisoning. We found that AFB1 generated clinical and pathological symptoms of liver injury in mice, as evident from the increased haptic enzymes AST, ALT, ALP, GGT, and decreased globulin and albumin content as well as fatty droplets and hepatocytes infiltration with macro vesicles in the liver of mice. These findings imply that AFB1 can cause hepatotoxicity, consistent with previous findings [1. The current results align with a previous study, which has reported that LUTN supplementation prevented acetaminophen-induced hepatic injury in rats [1-induced hepatic damage in mice. However, special clinical studies are warranted to know the hepatoprotective effects of LUTN on ongoing or established AFB1 toxicity in humans or animals.Aflatoxins, threatening mycotoxins, are commonly found in cereals and animal forages and pose major health and economic risks to humans and animals . Growth  by AFB1 . The livfindings ,38,39. I in rats . The pre1-induced hepatotoxicity and apoptosis in mice [1 exposure generated ROS and MDA accumulation and inhibited antioxidant enzyme activities such as T-SOD, CAT, GAH-Px and T-AOC, and induced apoptosis in the liver of mice. LUTN is considered a potent ROS scavenger, protecting cells from ROS accumulation and apoptosis induced by oxidative stress [1 as evidenced by decreased ROS and MDA accumulation, strengthened antioxidant defense system , and reduced apoptosis rate in the liver of mice.Oxidative stress is considered to be critical molecular process underlying cell damage . Oxidati in mice ,30,42. Se stress . We foun1 harmful effects in causing apoptosis [1 exposure could cause apoptosis in various tissues and cells and the mechanism was linked with the mitochondrial apoptosis pathway [1. These findings demonstrated that AFB1-induced excessive apoptosis in the liver of mice is linked with the mitochondrial apoptosis pathway. Notably, LUTN treatment increased the Bcl-2 expression, inhibited mitochondrial Cyt-c release and reduced the activated caspase-3 and caspase-9 expression in the liver of mice under AFB1 exposure. Furthermore, mitochondria are the primary target of ROS attacks, and superfluous ROS production can lead to oxidative stress and mitochondrial malfunctioning [1. As a result, we hypothesized that LUTN reduces AFB1-induced excessive apoptosis in mouse liver either directly or through the suppression of oxidative stress.Mitochondria have been identified as the primary targets for AFBpoptosis . The mitpoptosis . Bcl-2 ipoptosis ,47. The poptosis ,49. Prev pathway ,50,51,52ctioning . We obse1-induced cytotoxicity [1 exposure inhibited the expression of Nrf2 and its associated-target genes such as HO-1, NQO1, SOD1, and GCLC in the liver of mice. However, LUTN dramatically rescued these effects induced by AFB1. Previously, LUTN prevents the progression of liver fibrosis induced by carbon tetrachloride-(CCL4) through targeting AKT/mTOR/p70S6K and TGFβ/Smad signaling pathways [1 exposure suppressed Nrf2 nuclear translocation [1-induced toxicity in human and/or animal.Nrf2 is a transcription factor that plays a significant role in the process of AFBtoxicity ,54. Undetoxicity . Nrf2 antoxicity . The prepathways . The curlocation ,54, and location ,58,59. H1-induced hepatotoxicity in mice. LUTN effectively rescued liver injury, as evident by the amelioration of toxic effects on serum biochemical profile and pathological alterations induced by AFB1. LUTN attenuates AFB1-induced excessive apoptosis by inhibiting the mitochondria-dependent apoptosis pathway. Additionally, LUTN suppressed AFB1-induced oxidative stress by scavenging ROS accumulation and enhancing antioxidant enzymes capacity via regulation of Nrf2 signaling. The current study suggested that the key mechanisms underlying the LUTN hepatoprotective effects were associated with the activation of the Nrf2 signaling pathway  against AFB pathway . The pre


The outbreak of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly evolved into a global pandemic. Recent findings indicate an increased risk for kidney involvement, including progressive acute kidney injury (AKI) during COVID-19 infection, specifically in critically ill patients, and associated with high mortality rates. As no specific treatment options exist for AKI secondary to COVID-19, intensive care is largely supportive with a frequent need for renal replacement therapy (RRT). Convalescent plasma (CP) has been approved as an emergency investigational drug with clinical benefits in observational studies. We described a first case of a patient with severe COVID-19 and AKI who had remarkable improvement in his respiratory status and in kidney function following CP therapy. Our findings demonstrate important therapeutic implications of effective multimodality therapy including CP when treating patients with COVID-19 and AKI.  Although pulmonary involvement and respiratory failure are the main features of coronavirus disease 2019 (COVID-19) infection, recent findings indicate an increased risk for acute kidney injury (AKI) during the course of COVID-19 infection , 4, 5. AAn 81-year-old Caucasian man with a history of hypertension, type 2 diabetes mellitus, cardiovascular disease, hyperlipidemia, and benign prostate hypertrophy presented with a fall at home without witnessed syncope or seizures. His medications consisted of clopidogrel, enalapril, finasteride, gabapentin, glimepiride, sitagliptin, metformin, and tamsulosin. 3/µL; hemoglobin, 12.2 g/dL; thrombocytes, 89 × 103/µL; glucose, 235 mg/dL; sodium, 134 mEq/L; potassium 4.6 mEq/L; bicarbonate, 16 mEq/L; calcium, 8.5 mEq/L; serum creatinine (Scr) 1.1 mg/dL; blood urea nitrogen (BUN), 29 mg/dL; creatine phosphokinase (CPK), 9,128 IU/L; prothrombin time, 13.4 s; partial thromboplastin time, 25.3 s. Urinalysis and urinary microscopy demonstrated glucose, erythrocytes, and trace protein and no casts. A chest X-ray showed clear lungs. COVID-19 infection was diagnosed based on a reverse transcriptase-polymerase chain reaction test that detected acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a throat swab sample. Initial treatment was supportive, with nasal cannula oxygen, doxycycline, and thrombosis prophylaxis with apixaban. At presentation at the Mount Sinai Medical Center, Brooklyn, New York, in September 2020, the patient complained of weakness, fatigue, and frequent urinations. His body temperature was 39.0 °C, and blood pressure was 150/95 mm Hg, with a pulse rate of 98 beats/minute and oxygen saturation rate of 98%. He was alert and oriented, with unremarkable lung and cardiovascular examination. Laboratory findings showed leukocytes, 6.6 × 102 78 mmHg, Pa CO2 31 mmHg, and HCO3 13 meq/L. Chest X-ray showed diffuse patchy bilateral infiltrates. The patient was transferred to the Intensive Care Unit (ICU), started BiPAP therapy, intravenous steroids, and was given convalescent plasma. Over the next 48 hours, his respiratory status started improving, with Scr decreasing to 1.2 mg/dL, and CPK to 280 IU/L  . Over thAcute respiratory distress syndrome due to diffuse alveolar damage has been associated with highest morbidity and mortality, but it is now being described that involvement of the other organs, such as the heart, kidney, and nervous system, may occur and influence the clinical course and patient outcome. Renal involvement with proteinuria, hematuria, and changes in kidney function might be present in up to 75% of cases . AKI is The exact mechanism of kidney involvement is likely to be multifactorial with direct tubular and endothelial damage through an angiotensin-converting enzyme 2 (ACE2)-dependent pathway causing mitochondrial dysfunction, acute tubular necrosis, collapsing glomerulopathy, and protein leakage in Bowman’s capsule . Other iEarly recognition of kidney involvement in COVID-19 and use of preventive and therapeutic measures to limit progression of AKI are crucial to reduce morbidity and mortality. As no specific treatment options exist for AKI secondary to COVID-19, intensive care is largely supportive. Current approaches to prevention and management of AKI, and identification of potential indications for use of RRT are based largely on clinical experience, and AKI strategies are adjusted empirically to patients with COVID-19 , 10. RemCP therapy, a classic adaptive immunotherapy, has been applied to the prevention and treatment of many bacterial and viral diseases since the 1880s. The use of CP was recommended as an empirical treatment during the outbreak of Ebola virus in 2014 and as a protocol for treatment of sever acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS) . On MarcIn conclusion, we presented a case of a patient with severe COVID-19 and AKI who had rapid and substantial improvement in his respiratory status and in kidney function following CP therapy. The in-hospital mortality of patients with COVID-19 with AKI is extremely high, and our findings may have a significant impact on therapeutic approach in management of these patients. Therefore, we recommend that physicians carefully monitor kidney function from the early period and administer effective multimodality therapy including CP when treating patients with COVID-19 and AKI. There was no support/funding for this report. There is no potential conflict of interest.