A total of 79 aroma compounds were identified in a typical modern type xiaoqu Baijiu (M) and a typical traditional type xiaoqu Baijiu (T), 42 of them had OAV > 1 in both M and T samples. The main differences between the two samples were obtained for the concentration of 23 aroma-active compounds. A total of 22 samples made by different brewing processes were analyzed to confirm the differences. Partial least squares discriminant analysis confirmed that 20 compounds could be used as potential markers for discrimination between modern type xiaoqu Baijiu and traditional type xiaoqu Baijiu. Their difference in content is between 1.5 and 17.9 times for modern type xiaoqu Baijiu and traditional type xiaoqu Baijiu. The results showed the aroma characteristics of modern and traditional type xiaoqu Baijiu clearly and comprehensively, which will provide guidance for modern Baijiu quality control and evaluation.Predominant odorants in modern and traditional types of Chinese  Traditional fermented foods and beverages (TFFB) have been an important part of the human diet since the beginning of civilization . TFFB reBaijiu, Chinese liquor, is one of the most popular TFFB in China , which hxiaoqu Baijiu, respectively, by aroma extract dilution analysis and odor activity values (OAVs). However, the study was focused on only two samples and did not reveal the difference between the aroma compounds of the two types of xiaoqu Baijiu. The aroma is the most important characteristic of alcoholic beverages, and the production process is an important factor affecting the aroma profile [There are very few publications comparing modern type and traditional type Baijiu. Wang et al.  separate profile . So, theXiaoqu Baijiu, one of the oldest and most popular Baijiu in China, is the first and most successful one to which the fully mechanized modern brewing process has been applied. Therefore, it is necessary to understand fully the similarities and differences of aromatic compounds in the Baijiu manufactured by different production processes. The primary objectives of this study were: (1) to identify the major aroma-active compounds in traditional and modern type xiaoqu Baijiu using gas chromatography–olfactometry (GC–O) and gas chromatography–mass spectrometry (GC–MS), (2) to confirm the contribution of those aroma compounds by quantitative analysis and their OAVs, and (3) to verify the aroma difference between the two types of xiaoqu Baijiu using multivariate statistical techniques.xiaoqu Baijiu raw liquors were analyzed in this study, including 12 modern type of xiaoqu Baijiu and 12 traditional type xiaoqu Baijiu. All samples were manufactured by Jing Brand Co., Ltd.  in 2019. The representative modern and traditional type Baijiu samples were chosen by the sensory panel composed of five national Chinese Baijiu tasting judges in Jing Brand Co., Ltd. Among the obtained xiaoqu Baijiu samples, one modern type xiaoqu Baijiu sample (M) and one traditional type xiaoqu Baijiu sample (T) were selected for liquid–liquid extraction (LLE) and GC–O analysis. All the samples were stored at 4 °C until analysis. The brand name is mentioned here for clear labeling, not for advertising purposes. A total of 24 β-phenethyl alcohol, 1,1-dimethoxyethane, 2-methylpropanal, 3-methylbutanal, hexanal, decanal, 1,1,3-triethoxypropane, benzaldehyde, benzeneacetaldehyde, 2-pentanone, 2-octanone, acetophenone, 2-pentadecanone, acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, pentanoic acid, 4-methylpentanoic acid, hexanoic acid, octanoic acid, guaiacol, 4-methylguaiacol, 4-ethylguaiacol, 4-vinylguaiacol, 4-methylphenol, 4-ethylphenol, linalool, β-damascenone, geraniol, geranylacetone, β-ionone, 2-pentylfuran, furfural, 2-furan methanol, γ-nonanolactone, dimethyl trisulfide, 2-thiophenecarboxaldehyde, 2-phenylethyl acetate-D3 (IS1), n-hexyl-D13 alcohol (IS2), 2,2-dimethylpropanoic acid (IS3), 2-octanol (IS4), 2-methoxy-D3-phenol (IS5), and lactic acid were purchased from Sigma-Aldrich . A C6–C30n-alkanes mixture  was used for the determination of retention indices (RIs). Sodium chloride (NaCl), sodium carbonate (Na2CO3), anhydrous sodium sulfate (Na2SO4), and hydrochloric acid (HCl) were purchased from China National Pharmaceutical Group Corp. . Ethanol (high-performance liquid chromatography (HPLC) grade) was purchased from J&K Scientific Co., Ltd. . Dichloromethane  was distilled before use. Ultrapure water was obtained from a Milli-Q purification system .Chemical standards and internal standards (ISs) were supplied commercially at a high-purity grade . Ethyl acetate, isobutyl acetate, isoamyl acetate, pentyl acetate, ethyl propanoate, ethyl 2-methylpropanoate, ethyl butanoate, ethyl 2-methylbutanoate, ethyl 3-methylbutanoate, 3-methylbutyl butanoate, ethyl pentanoate, ethyl hexanoate, propyl hexanoate, hexyl hexanoate, ethyl heptanoate, ethyl octanoate, ethyl nonanoate, ethyl decanoate, ethyl lactate, diethyl succinate, ethyl benzoate, ethyl phenylacetate, 2-phenethyl acetate, ethyl 3-phenylpropanoate, ethyl dodecanoate, ethyl tetradecanoate, 1-propanol, 2-methylpropanol, 1-butanol, 2-butanol, 3-methylbutanol, 1-pentanol, 1-hexanol, 1-heptanol, 2-heptanol, 1-octanol, 3-octanol, 1-octen-3-ol, 1-nonanol, benzyl alcohol, 2Cl2 (100 mL each time) in a separatory funnel. The combined extracts were further separated into acidic/water-soluble and neutral/basic fractions with the following method.The extraction method was modified from a previous study . The M a2CO3  and then washed with 30 mL of saturated NaCl solution. The obtained organic phase, containing the neutral/basic aroma compounds, was named the NBF fraction. The combined aqueous phase was acidified to pH 2.0 with HCl (4.0 mol/L) and extracted three times with freshly distilled CH2Cl2 (50 mL each time). The combined extract (about 150 mL), containing the remaining acidic aroma compounds, was named the AF fraction. Afterward, each fraction was dried with anhydrous Na2SO4 overnight and concentrated to a final volume of 0.5 mL under a gentle stream of nitrogen. These concentrated NBF and AF were stored at −20 °C before the GC–O analysis.The combined extract (about 300 mL) was washed three times with NaGC–MS and GC–O analyses were performed on an Agilent 6890N GC equipped with an Agilent 5975 mass selective detector and a sniffing port . The sample was analyzed on both a DB-FFAP column  and a DB-5 column . High-purity helium (>99.999%) was used as the carrier gas at a constant flow rate of 2 mL/min. The concentrated extract (1 µL) was injected into the GC injector in splitless mode. The GC injector temperature was 250 °C. The oven temperature was held at 50 °C for 2 min, then raised to 230 °C at a rate of 6 °C/min and held for 15 min. The mass spectrometer (MS) was operated in electron ionization mode at 70 eV. The time of solvent delay was 5 min, and the temperature of the ion source was 230 °C. The mass range was set from 35 to 350 amu in full-scan mode.Four well-trained panelists  from the Laboratory of Brewing Microbiology and Applied Enzymology at Jiangnan University were selected for the GC–O study. During a GC run described above, a panelist placed his/her nose close to the sniffing port, responded to the aroma intensity of the stimulus, and recorded the retention time, descriptors, and intensities of the odor peak for each compound. The Osme values reflecting the aromatic intensity of the stimulus based on a six-point scale ranging from 0 to 5 was used for intensity judgment: 0 = none, 1 = very weak, 2 = weak, 3 = moderate, 4 = strong, and 5 = very strong. Every sample was sniffed twice by each panelist, if the aroma was detected six or more times on each column, an odor location was identified. The Osme value for aroma intensity was an average result of the four panelists.n-alkanes (C6–C30) in both the DB-FFAP and DB-5 columns under the same chromatographic conditions.The aroma compounds of the odor location were identified by comparing the aroma, mass spectrum (MS), and RIs to the pure standards (S). RIs were calculated based on the linear retention times of the xiaoqu Baijiu samples were quantitated using headspace solid-phase microextraction arrow (HS-SPME-Arrow) combined with GC–MS, as described previously [xiaoqu Baijiu sample (10 vol%) was transferred into a 20 mL headspace glass vial, saturated with 1.5 g NaCl, and spiked with 40 μL mixed ISs . Tightly capped with a Teflon-faced silicone septum, the sample was equilibrated at 45 °C for 5 min and extracted for 45 min at the same temperature under stirring of 250 rpm. After extraction, the fiber was immediately inserted into the injection port of the GC for desorption at 250 °C for 5 min. The samples were separated on a DB-FFAP column with splitless mode. The oven temperature was held at 40 °C for 2 min, increased at 4 °C/min to 150 °C, then held for 2 min, raised at 6 °C/min to 230 °C and held for 5 min.The aroma compounds in all eviously . An SPMExiaoqu Baijiu samples was added to the standard solutions, and then the working standard solutions were extracted using HS-SPME-Arrow and detected using GC–MS with the same conditions as for sample analysis. The standard curves were created by plotting the ratio of the peak area of the reference compounds to the corresponding IS against their concentration ratio. The analytical limit of detection (LOD) of aroma compounds was obtained from the lowest concentrations of the analyte standard solutions based on a signal-to-noise ratio of 3. The limits of quantitation (LOQ) was defined as the lowest concentration of the calibration curve based on a signal-to-noise ratio of 10. The accuracy is reported as the percentage recovery of a known amount of target analyte added to the xiaoqu Baijiu sample. All analyses were performed in triplicate.Every standard stock solution was prepared by dissolving a standard compound in the model synthetic solution  and then diluting to 10 different concentrations. The same amount of ISs as in the xiaoqu Baijiu samples in terms of their aroma compounds. To maximize the separation among samples and to identify the aroma compounds responsible for the separation, partial least squares discriminant analysis (PLS-DA) was used. PCA and PLS-DA were conducted using SIMCA version 14 software . The heatmap and hierarchical clustering analysis (HCA) were performed using HemI 1.0 .Principal component analysis (PCA) was performed to explore the clustering of the 2Cl2, an organic phase was obtained that exactly showed the typical xiaoqu Baijiu aroma when sniffed on a filter paper by the panelists. Thus, it was appropriate and reliable to be used to analyze the profile of aroma compounds in xiaoqu Baijiu. To facilitate GC–O analysis and compound identification, the extract obtained was further separated into AF and NBF fractions to reduce complexity as previously described [By extraction with CHescribed . To idenxiaoqu Baijiu [As shown in u Baijiu . Among tThe dominant group of odorants identified was esters, particularly ethyl esters. The most important esters identified were ethyl acetate, isoamyl acetate, ethyl 2-methylpropanoate, and ethyl octanoate with Osme values above 3.00 in both samples, contributing a strong fruity note. In general, the T sample had higher Osme values for most esters than the M sample, such as ethyl butanoate , ethyl pentanoate , ethyl hexanoate , and ethyl 3-phenylpropanoate . However, some other esters had higher Osme values in the M sample. They were ethyl acetate , ethyl 3-methylbutanoate , ethyl octanoate , ethyl decanoate , and ethyl phenylacetate .β-phenethyl alcohol showed the highest Osme values of 3.65, contributing whiskey, mushroom, and honey odors, respectively. However, 2-methylpropanol with the Osme value of 3.52 was the most important aroma contributor in the T sample.The second dominant group of aroma compounds identified were alcohols. Among the alcoholic compounds identified in the M sample, 3-methylbutanol, 1-octen-3-ol, and xiaoqu Baijiu, where 2-methylpropanal , 1,1-dimethoxyethane , hexanal , and benzeneacetaldehyde  had high Osme values above 3.00 and contributed to malty, fruity, green, and honey odors, respectively. In this study, four ketones were identified. Among them, 2-octanone with soap aroma showed the highest Osme values of 3.22 and 3.20 in M and T, respectively. All aldehydes and ketones had higher Osme values in the M sample than in the T sample except 1,1,3-triethoxypropane.Based on GC–O and GC–MS, eight aroma aldehydes were detected. Aromatic aldehydes seemed to be important in β-damascenone , and β-ionone  could be important to the aroma of xiaoqu Baijiu because of the high Osme values. Sulfur compounds are very important for Baijiu because of their low thresholds. They are mainly produced by the degradation of sulfur-containing amino acids [The important acids in M and T with high Osme values were acetic acid , 2-methylpropanoic acid , butanoic acid , 3-methylbutanoic acid , pentanoic acid , and hexanoic acid . Acids are important components in the quality and taste of alcoholic beverages. Most of the acids in Baijiu are produced by yeast during fermentation, followed by oxidation of alcohol and aldehyde . Acetic no acids . Two sulxiaoqu Baijiu, the 79 odorants detected using GC–O were quantitated by constructing standard curves for the M and T samples. The quantitative ion, IS, calibration curve, recovery, and LOQ of each aroma compound are shown in To gain a deeper insight into the aroma-active compounds of β-damascenone , dimethyl trisulfide , 1,1-dimethoxyethane , isoamyl acetate , ethyl hexanoate , ethyl pentanoate , and ethyl butanoate . These results revealed the important aroma contribution of esters, especially ethyl esters, for xiaoqu Baijiu liquors. Esters, which contribute a strong fruity odor, were also one of the most important aroma groups for other alcoholic beverages, such as wine [β-damascenone and dimethyl trisulfide was not high but the OAVs ranked at second and third places among all aroma compounds in xiaoqu Baijiu. This is because their thresholds are present at trace amounts in Baijiu [β-Damascenone giving a honey note was previously reported as a key odorant with high OAV in the light aroma type of Baijiu [β-ionone, ethyl lactate, 4-vinylguaiacol, 4-ethylphenol, and 2-phenethyl acetate, were other aroma contributors.It is known that the contribution to Baijiu depends not only on the concentration of an odorant but also its odor threshold. So, for further confirmation of the contribution of aroma-active compounds, their OAVs were calculated. The OAV was obtained from a compound concentration divided by its odor threshold . Herein, as wine , rice wi as wine , and oth as wine ,25,28. Bn Baijiu ,25. β-Daf Baijiu ,30. The xiaoqu Baijiu and traditional type xiaoqu Baijiu. To determine if the differences between M and T samples can also be found in an additional 22 different types of xiaoqu Baijiu samples, which were randomly selected from the modern and traditional workshops, the 79 aroma compounds were quantitated. Then, PCA and PLS-DA analyses were conducted based on the concentration of the aroma compounds in the 22 xiaoqu Baijiu samples.As concluded above, the odorant compositions of M and T infusions were similar, but the Osme value and OAV of each substance were different. However, these were just the results based on two samples, not enough to represent all the aroma characteristics of modern type xiaoqu Baijiu samples were well separated. PC1 clearly distinguished modern type xiaoqu Baijiu and traditional type xiaoqu Baijiu, traditional type xiaoqu Baijiu was on the positive side of PC1, while modern type xiaoqu Baijiu appeared on the negative side, indicating that there were obvious differences in aroma compounds’ features.PCA was performed to show a trend of intergroup separation on the scores plot of data obtained in both positive and negative modes. The PCA results showed that five principal components (PCs) with eigenvalues higher than one were obtained from the data sets, whose cumulative variance proportion was 74.5%. PC1 accounted for 46.7% of the total variance while PC2 explained 7.02%, which explained 55.1% of the total variance in the data set. As shown in xiaoqu Baijiu, a PLS-DA model was created for the 22 xiaoqu Baijiu samples based on the concentration data set. This type of supervised pattern recognition method provides a deeper analysis of the main matrix characteristics of samples and differs from PCA by the addition of grouping variables that indicate the category of samples. As shown in xiaoqu Baijiu samples were well distinguished by a dependable PLS-DA model. The explained variation (R2Y) and the predictive capability (Q2Y) for the model were 0.993 and 0.978, respectively, which indicated that the model was excellent because Q2Y was close to 1. In addition, permutation tests (n = 200) were performed to evaluate whether the discriminant models were overfitting the data [2Y up to 200 times. The low intercepts  were an indication of the validity of the original model. The R2 and Q2 values of the permutation test revealed that the initial model outperformed the randomly permuted models  indicated the concentration change from low to high. As shown in xiaoqu Baijiu than in modern type xiaoqu Baijiu, which included five ethyl esters , three phenols , pentanoic acid, β-ionone, and 2-butanol.To compare further the content of each substance in the two types of xiaoqu Baijiu were higher than those in modern type xiaoqu Baijiu, so the organic acids had a significant influence on the yield of aroma esters. Phenols, as an important trace component in Baijiu, play an important role in the aroma, taste, and stability of Baijiu [xiaoqu Baijiu and traditional type xiaoqu Baijiu are the same, therefore, it may be that the different types and abundance of microorganisms caused the higher content of phenols in traditional type xiaoqu Baijiu. β-Ionone with a floral aroma is also a very important aroma compound and bioactive substance in Baijiu [Saccharomyces cerevisiae and bacteria) during fermentation via catabolic and anabolic pathways [xiaoqu Baijiu is highly susceptible to climatic conditions, but the fermentation temperature of modern type xiaoqu Baijiu is controlled at a suitable temperature, which could effectively inhibit the production of higher alcohols [xiaoqu Baijiu than in traditional type xiaoqu Baijiu, which included three esters , guaiacol, γ-nonanolactone, β-phenethyl alcohol, β-damascenone, benzeneacetaldehyde, and 1-octen-3-ol. Not surprisingly, the acids content corresponding to these three esters in modern type xiaoqu Baijiu was also higher than in traditional type xiaoqu Baijiu. β-Phenethyl alcohol with a rose-like aroma is a very important aroma compound in Baijiu. The processes by which the key odorant is formed have been well-established and found to be influenced by various yeast species present during the fermentation of liquor from cereal and legume materials [β-Damascenone, presenting honey and floral aromas, was previously reported as an important odorant in whiskey, rum, brandy, and Baijiu [β-ionol) leading to the generation of β-damascenone during fermentation and/or distillation.The ethyl esters in Baijiu were mainly produced in the fermentation or aging stages through the esterification of acids and alcohols ,33. Thisf Baijiu . It was f Baijiu ,36. The n Baijiu . Besidespathways ,39. The alcohols . Clusteraterials . β-Damasd Baijiu ,41,42,43xiaoqu Baijiu and modern type xiaoqu Baijiu, multivariate analyses of the 22 xiaoqu Baijiu samples made by different brewing processes were applied, including PCA, PLS-DA, and HCA. Twenty odor-active compounds were confirmed as potential flavor markers for the differentiation of modern type xiaoqu Baijiu and traditional type xiaoqu Baijiu. The results in this study will help improve the flavor quality and processing technology of xiaoqu Baijiu.In summary, a total of 79 aroma compounds were identified using GC–O and GC–MS in both M and T samples. Among them, 42 aroma compounds were further recognized as the important aroma-active compounds in both M and T owing to their relatively high OAVs. Twenty-three aroma-active compounds might be the potential compounds responsible for the differences in the aroma profile between M and T samples because of a concentration ratio higher than 1.5. Moreover, to determine if the 23 aroma-active compounds can represent the difference in aroma compounds’ profile between traditional type


Aconitum, Delphinium, Consolida genera. Corals, Xenia, Okinawan/Clavularia, Alcyonacea  and marine sponges are rich sources of diterpenoids, despite the difficulty to access them and the lack of availability. Researchers have long been concerned with the potential beneficial or harmful effects of diterpenoid alkaloids due to their structural complexity, which accounts for their use as pharmaceuticals as well as their lousy reputation as toxic substances. Compounds belonging to this unique and fascinating family of natural products exhibit a broad spectrum of biological activities. Some of these compounds are on the list of clinical drugs, while others act as incredibly potent neurotoxins. Despite numerous attempts to prepare synthetic products, this review only introduces the natural diterpenoid alkaloids, describing ‘compounds’ structures and classifications and their toxicity and bioactivity. The purpose of the review is to highlight some existing relationships between the presence of substituents in the structure of such molecules and their recognised bioactivity.Diterpenoid alkaloids are natural compounds having complex structural features with many stereo-centres originating from the amination of natural tetracyclic diterpenes and produced primarily from plants in the  Diterpenoid alkaloids (DAs) are substances produced by various natural plants with significant thematic difficulties, bioactivity, and somewhat disreputable toxicity. To date, 1500 and more DAs have been isolated and characterised. Many remarkable DAs demonstrate different pharmacological properties such as neurotropic, antimicrobial, antitumour, hypotensive, analgesic, anti-inflammatory, muscle relaxant, antiarrhythmic, and local anaesthetic ,5,6,7,8.Delphinium, Aconitum and Consolida, are often distinctive and recognised as cytotoxic against cancer  a a1, 3, 4[59 (161  showed IMCC-7721 .153 (50 of 3.1 µM [112 (50 3.16 µM [DA-153  has a stf 3.1 µM ,149; fur µM [112  shows a  3.16 µM .51 [59 [161 (51–55 (Compounds 51  and 59 [51 [59  (Figure [59 [161  [101,1511 (51–55 b exerted19-DAs may play an essential role in cytotoxicity, especially substitutions in C1, C3, C6, and C8 [SAR of antitumor DAs indicates that the number and position of the hydroxyl and ester groups in C, and C8 ,152,153.Aconitum (165–167 in Three new bis-DAs derived from genus Aconitum, Delphinium, and Consolida genera.Over the past decade, more than 300 DAs were discovered and extracted from plants, particularly 18-DAs and C19-DAs groups has justified their development into new therapeutic drugs, except glycosidic DAs, which have additional sugar moieties in their structures that facilitate their water solubility unlike the other DA groups. This observation gives future hope to discovering new chemical compounds with low toxicity and useful bio-activity in the aqueous extracts of alkaloids with SAR similar to C19-DAs. The complex nature of the diterpenoid-alkaloids’ SAR suggests the need for an accurate knowledge of individual compound properties to discover further safe and valuable applications of novel bioactive compounds.Structurally, DAs derived from four isoprenyl ‘units’ condensation subdivide into more than 45 classes based on their central structure arrangement and different substituent. These compounds display a broad area of pleasant chemical properties and biological activity, such as analgesic, anti-inflammatory, antimicrobial, cytotoxic activity, and toxic effects. Their toxic effect is manifested in the nervous and cardiovascular systems, acting as potent neurotoxins and cardiotoxins. The toxicity of C20-DAs after a SAR analysis, displayed their chemical structure diversity and their little toxicity compared to C19-DAs. In addition, their classification into seven groups with different SARs facilitates the search for biologically active molecules and potential new drugs.The ‘researchers’ competition, turned to deeper study of C19-DAs and C20-DAs have noticeable effectiveness. The C20-hetisine class showed the highest possibilities with the lowest toxicity among the other DAs. For this reason, the hetisine compounds may be good starters for developing novel anticancer drugs using alkaloids.Many research efforts, oriented to studying the anti-inflammatory, analgesic, and anticancer activity of DAs, highlighted that numerous C


The coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), poses several challenges to clinicians, due to its unpredictable clinical course. The identification of laboratory biomarkers, specific cellular, and molecular mediators of immune response could contribute to the prognosis and management of COVID-19 patients. Of utmost importance is also the detection of differentially expressed genes, which can serve as transcriptomic signatures, providing information valuable to stratify patients into groups, based on the severity of the disease. The role of biomarkers such as IL-6, procalcitonin, neutrophil–lymphocyte ratio, white blood cell counts, etc. has already been highlighted in recently published studies; however, there is a notable amount of new evidence that has not been summarized yet, especially regarding transcriptomic signatures. Hence, in this review, we assess the latest cellular and molecular data and determine the significance of abnormalities in potential biomarkers for COVID-19 severity and persistence. Furthermore, we applied Gene Ontology (GO) enrichment analysis using the genes reported as differentially expressed in the literature in order to investigate which biological pathways are significantly enriched. The analysis revealed a number of processes, such as inflammatory response, and monocyte and neutrophil chemotaxis, which occur as part of the complex immune response to SARS-CoV-2. It was initially considered to be a respiratory infection; however, based on the clinical experience on COVID-19 acquired during the past 18 months, SARS-CoV-2 infection is now recognized as a multisystem disease, resulting in a wide range of clinical manifestations . Th. ThELANEand MMP9  38,84]  ELANE, O T cells .BLIMP-1 via STAT3 signaling [BLIMP-1 encodes PRDM1, which appears to be significantly activated at critical and convalescent phases [Multiple studies highlight elevated expression of a number of genes involved in B cell activation and differentiation to plasmablasts in patients with severe COVID-19. Parallel single cell RNA-seq (scRNAseq) and multiplex serum ELISA were performed on samples from 13 inpatients, a recovery control, and 14 healthy individuals, revealing that B cells were depleted, whereas proportions of PBs were elevated from the time of the infection until the early convalescent phase . This elignaling ,86. BLIMt phases . IFI27, IFI6, IFITM1) characterize the inflammatory response in every phase of the disease except critical [PIM2 and S100A4 (cell proliferation) remain elevated during the incremental phase of inflammation in patients with severe disease  and POU2AF1 in plasma cells, with disease severity [IRF4, together with BATF and CD27, are genes associated with T cell exhaustion, and, the more severe the disease is, the higher their expression in the blood samples [low cells expressing high and low levels of SELL and CCL5, respectively (particularly in the disease progression phase), and T_CD8_c10-MKI67-GZMK cells with upregulated STMN1, HMGB2, MKI67, and GZMK mainly in the convalescent phase [CD38, SLC1A4 involved in response to unfolded proteins, and ATP synthesis in mitochondria [CD44, IRF8, and CD244) or are characterized as immune-negative  or was promoted by signals that act as pro-inflammatory cytokines, like CCL3, CCL4, and the already mentioned IL-1 and TNF-α) [Increased PBs in severe versus mild disease, accompanied by the presence of IFN-I related gene products  . It shoud TNF-α) . Activatd TNF-α) .IRF3/7/8 and MAVS are suppressed [RPL32, RPL30, RPS12, and RPS3A, in hospitalized patients [IL-6, IL-1RA, KLF6, SPP1, CCL3, CCL2, and CCL8 [According to the results of the GO enrichment analysis, a number of differentially expressed genes was related to the GO terms type I interferon and interferon-gamma-mediated signaling pathways, indicating that IFN-I and II responses are predominant in patients with severe COVID-19 . Indeed,ppressed . Probablppressed , a fact patients . Overexppatients . Along wpatients . Contrarpatients . Blanco-patients  performepatients . Other oand CCL8 ; it shouand CCL8 . These fand CCL8 . Levels and CCL8 . On the and CCL8 . In geneand CCL8 . IFITM1, IFITM2, IFITM3, PKM, and ITGA2B (CD41) that are overexpressed in megakaryocytes (MKs) of individuals suffering severe disease and ODC1 and TGFB1, which are downregulated [IFITM1 overexpression in the plasma of severely ill is confirmed by Lee et al., who demonstrated that the expression of various ISGs, such as IFITM2, IFITM3, ISG20 and genes activated as a response to TNF/IL-1β activation, is elevated [IFITM1 and IFITM2 serve as gene signatures for severe disease as, together with H3F3B, SAT1, and S100A8 in neutrophils, they are highly expressed in severe patients compared to mild and uninfected [IFITM1, IFIT2, and MX1 is suppressed in critical patients, while IFNAR1, JAK1, and TYK2  are upregulated [IFI27 and IFITM3 between severe and mild samples [IFI27 was upregulated and the overexpression is correlated with high viral load [IFI27 levels in the upper respiratory tract  and the severity of the disease was reported [ISG15 and JAK3 which are elevated, and the downregulated RPS18 [IFN-I stimulated genes differentially expressed are egulated . IFITM1 elevated . Other sinfected ,88. The egulated . Of noteegulated . Other Iegulated . Regardiegulated . In cont samples  and a stral load . On the reported . This IFreported . Data deed RPS18 . In geneed RPS18 .The transcriptomic and KEGG pathway analysis of PBMCs of COVID-19 patients revealed that the main causes of lymphopenia might be the activation of apoptotic and P53 signaling pathways. Specifically, in the surface of T- and B cells, as well as in monocytes, in severe patients compared to the other groups, elevated expression of apoptotic signals is reported. Along with the suppression of expansion pathways, resulting in lower proliferating rates, these signals could be driving the lymphopenia ,96. ThisIRF1 and IRF9 expression is lower in severe patients compared to moderately ill, resulting in a deficient response of monocytes in IFN(-I) signaling, while on the other hand, elevated IRF7 expression in lymphocytes results in a more robust IFN-I response [PKM2) was observed [A condition of hypoxia is being established during different time points of the disease, each time accompanied by a different subset of genes with altered expression. Elevated levels of transcription factors, such as IRF1, STAT3, and the hypoxic signal HIF1A (hypoxia-inducible factor-1a), involved in inflammation and IFN signaling are also reported during active disease . Oppositresponse . In neutobserved ,98. The IL6R, SOCS3, and STAT3 genes induced by IL-6 is not reflected at the transcriptional level, as IL-6- mRNA was not detected in the PBMCs [The inflammatory response is the third biological process found to be enriched in the GO enrichment, while a large number of significantly differentially expressed genes are involved in the positive regulation of inflammatory response, cellular response to IL−1, and cellular response to tumor necrosis factor . Overexphe PBMCs . The samhe PBMCs . Elevatehe PBMCs  in severhe PBMCs ,71. The he PBMCs . Contrarhe PBMCs , other she PBMCs ,100, andhe PBMCs . Additiohe PBMCs . The comhe PBMCs ,102. Othhe PBMCs ,100, whihe PBMCs .It has been almost two years since the Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2) emerged and was designated as the causative agent of COVID-19, an infection which presents with great clinical variability, resulting in a great variety of distinctive features and extensive disease distribution, with extrapulmonary manifestations. Active research is necessary to identify all serological, clinical, epidemiological features of COVID-19, and characterize the immune response via transcriptome analysis. The identification of a number of hematological and immunological markers, as well as a panel of genes that have potential as biomarkers associated with disease progression from mild/moderate to severe/critical, will ensure the successful stratification of patients and predict immune responses, enabling effective management.As we can conclude from the study of a great number of published papers about COVID-19, hematological parameters, cell populations subsets and transcriptomic signatures could be used as biomarkers of disease severity of COVID-19. In particular, higher levels of CRP, PCT, LDH, and liver function indices are associated with severe disease and fatal outcome. In addition, concerning cell populations, elevated levels of immature monocytes, megakaryocytes, and B cells, and decreased levels of dendritic cells  and T cells  are mainly observed in severe COVID-19 patients and as a result could be considered as markers of severe disease ,56,67,69


Indeed, CD4+CD28- T lymphocytes produce higher levels of osteoclastogenic molecular mediators directly related to pathologic bone loss, such as tumor necrosis factor (TNF)-α, interleukin (IL)-17A, and receptor-activator of nuclear factor κB ligand (RANKL), as compared with regular CD4+CD28+ T lymphocytes. In addition, premature senescent CD8+CD28- T lymphocytes have been negatively associated with bone healing and regeneration by inhibiting osteoblast differentiation and mesenchymal stromal cell survival. Therefore, accumulated evidence supports the role of senescent T lymphocytes in osteoimmunology. Moreover, premature senescence of T-cells seems to be associated with the functional imbalance between the osteolytic T-helper type-17 (Th17) and bone protective T regulatory (Treg) lymphocytes, as well as the phenotypic instability of Treg lymphocytes responsible for its trans-differentiation into RANKL-producing exFoxp3Th17 cells, a key cellular phenomenon directly related to bone loss. Herein, we present a framework for the understanding of the pathogenic characteristics of T lymphocytes with a premature senescent phenotype; and particularly, we revise and discuss their role in the osteoimmunology of osteolytic diseases.Cellular senescence is a biological process triggered in response to time-accumulated DNA damage, which prioritizes cell survival over cell function. Particularly, senescent T lymphocytes can be generated prematurely during chronic inflammatory diseases regardless of chronological aging. These senescent T lymphocytes are characterized by the loss of CD28 expression, a co-stimulatory receptor that mediates antigen presentation and effective T-cell activation. An increased number of premature senescent CD4 Cellular senescence is defined as a biological process characterized by the arrest of the cell cycle, frequently accompanied by changes in cell phenotype, in response to accumulated DNA damage . Indeed,Senescent cells display molecular and morphological changes which translate into heterogeneous functional variations, and these are dependent on the type of affected cell and the type, intensity, and duration of the stimulus that provoked the DNA damage , 3. EvenFunctionally, senescent cells acquire pro-inflammatory properties, characterized by increased production of reactive oxygen species (ROS), derived from dysfunctional mitochondria, and the establishment of a senescence-associated secretory phenotype (SASP), responsible for the increased production of cytokines, chemokines, growth factors, and proteases , 8. The Notably, there are no actual unique markers for the identification of cellular senescence ; however- T lymphocytes, also called CD28null T lymphocytes, present common characteristics of the senescent cell phenotype, such as the upregulation of γ-H2Ax and p53, and a robust increment in the SA-βgal activity [- T lymphocytes are considered dysfunctional, oligoclonal, terminally differentiated, and with limited proliferative capacity, although with an increased capacity to produce pro-inflammatory cytokines [- T lymphocytes are highly resistant to apoptosis-triggering signals and accumulate in high quantities in the elderly [During the antigenic presentation, T lymphocytes are activated when their T-cell receptor (TCR) recognizes the antigen/MHC II complex and their CD28 receptor binds to the costimulatory molecules CD80 or CD86, expressed by antigen-presenting cells . The losactivity -18. CD28ytokines , 19 and ytokines , 21. In  elderly , 23.It is noteworthy that senescent T lymphocytes can also be generated prematurely during chronic diseases regardless of chronological aging , since c2 can trigger the loss of CD28 expression and induce the formation of CD28- T lymphocytes with senescent characteristics [Various pathways can lead to premature senescence of T cells. During repetitive T lymphocyte stimulation involving clonal expansion, the CD28 expression may be irreversibly lost by transcriptional silencing . In addieristics -38.+ T lymphocytes and only in 50% of CD8+ T lymphocytes in humans; thus, the loss of CD28 expression seems to be a good senescence marker for CD4+ T lymphocytes but less specific for CD8+ T lymphocytes [+ and CD8+ T lymphocytes. Indeed, senescent CD8+ T lymphocytes first lose the CD28 expression and, in a later stage, they lose the CD27 expression, while CD4+ T lymphocytes lose first the CD27 expression and then the CD28 expression [+CD27-CD28- and CD8+CD27-CD28- T lymphocytes downregulate the expression of molecules specifically involved in the TCR signaling pathway, such as Zap70 and Lck [+CD27-CD28- T lymphocytes, sestrins induce natural killer functions [+CD27-CD28- T lymphocytes, sestrins induce phosphorylation of distinct mitogen-activated protein kinases (MAPKs) [+ T-cell senescence; for instance, the activity of p38-MAPK has been associated with low telomerase activity, ERK with the activation of DDR and upregulation of γ-H2Ax, and JNK with the signalosome inhibition [While CD28 is expressed in most murine T lymphocytes, CD28 is expressed in 90% of CD4phocytes . In thispression . Both CD and Lck , 39. Int and Lck -42. Ther (MAPKs) . Thus, Mhibition .Osteoimmunology studies the relation and cross-communication between the immune and bone systems. Although osteoimmunology initially focused on the influence of the immune response on osteoclast activity, the key cell responsible for bone resorption, nowadays it also involves its effects over osteoblast activity and bone formation , 44. Hen+ osteoclasts, and this induction can be completely blocked by using neutralizing antibodies against the TNF-α receptor instead of OPG or RANK neutralizing antibodies [+ cathepsin-K+ DC-STAMP+ osteoclasts in the absence of exogenous RANKL [Osteoimmunological control of osteoclast function comprises the activity of a triad of proteins belonging to the superfamily of TNF ligands and receptors, which are the receptor-activator of nuclear factor κB ligand (RANKL), its functional receptor (RANK), and its soluble decoy receptor osteoprotegerin (OPG) . In genetibodies , 48. Simus RANKL , 50.At a cellular level, the osteoimmunological regulation of osteoclasts development and activity is mediated by T lymphocytes, particularly T-helper type-17 (Th17) and T regulatory (Treg) lymphocytes, which have opposite roles during immune surveillance, autoimmunity, and inflammation , 51. Th1+ T lymphocytes. Indeed, the increase in the number and activity of TGF-β1-producing Treg lymphocytes drives the assembly of the NFAT1/SMAD3 transcription complex in CD8+ T lymphocytes and Wnt10b production, and consequently, osteoblast differentiation and bone formation [Otherwise, osteoimmunological control of osteoblast function and bone formation depends on the Wnt signaling pathway. In osteoblasts precursors such as mesenchymal stromal cells, the binding of Wnt10b to its coreceptor, presumably low-density lipoprotein receptor-related protein 5 (LRP5), leads to β-catenin nuclear translocation, which in turn induces the expression of the transcription factors responsible for osteoblastogenesis and necessary for bone formation, such as runt-related transcription factor 2 (Runx2), distal-less homeobox 5 (Dlx5), and osterix (Osx) . At a ceormation , 57, 58.+CD28- T lymphocytes has become relatively common in rheumatoid arthritis, juvenile idiopathic arthritis, ankylosing spondylitis, osteopenia, osteoporosis, and osteomyelitis [+CD28- T cells have also been reported in HIV-infected individuals and lupus erythematosus-affected patients, in which their activity could explain the increased risk of developing osteoporosis observed in these diseases [Despite having different etiologies, chronic diseases characterized by manifesting pathological bone loss usually share osteoimmunological similarities that affect the target tissues. In some of these diseases, premature senescence of T lymphocytes has been related to increased Th17 lymphocyte activity and RANKL production, which induces osteoclast differentiation and provokes adverse effects on bone structure and function . In factmyelitis -64. Apardiseases -68.+CD28- T lymphocytes produced higher levels of RANKL as compared with CD4+CD28+ T lymphocytes, and these senescent CD4+CD28- T lymphocytes showed a greater capacity to induce the differentiation of TRAP+ osteoclasts and bone resorption when compared with non-senescent CD4+CD28+ T lymphocytes [+CD28- T lymphocytes isolated from the synovial fluid of rheumatoid arthritis-affected patients were capable of producing higher levels of IL-17A as compared with isolated CD4+CD28+ T lymphocytes [+CD28- T lymphocytes expressed higher levels of TNF-α and intensely induced the activation of TRAP in osteoclasts as compared with CD4+CD28+ T lymphocytes [+CD28- T lymphocytes as compared with RANKL or TNF-α blockade, which translated into an improvement in skeletal parameters and demonstrated the key role for IL-17A in the CD4+ T-cell senescence [+CD28- T lymphocytes was observed in bone tissue, which was associated with long-term activated T cells, CD11b overexpression, and enhanced cytotoxic ability, all characteristics of T-cell senescence [+CD28- T lymphocytes during periodontitis, the most prevalent osteolytic disease in humans, has not been reported, the continuous inflammatory signals and the persistent antigenic stimulus, provided by microbial dysbiosis and the destruction of periodontal tissues, could trigger the loss of CD28 expression and senescence of CD4+ T lymphocytes [In the case of rheumatoid arthritis, CD4phocytes . Also, Cphocytes , 70. Durphocytes . Besidesnescence . During nescence , 73. Altphocytes , 75.+ and CD8+ T lymphocytes, Treg lymphocytes can also lose the CD28 expression and acquire a senescent phenotype and related functions (+CD28- T lymphocytes has been reported, and these senescent Treg cells expressed lower levels of CD25, showed higher SA-βgal activity, and produced higher levels of TNF-α, IFN-γ, and IL-17A, as compared with Foxp3+CD4+CD28+ lymphocytes [+CD4+CD28- lymphocytes, suggesting a state of premature senescence rather than replicative senescence [Similar to effector CD4unctions . In rheuphocytes . Interesnescence .+ Treg lymphocytes show low proliferative capacity and impaired suppressive functions in vitro and in vivo, frequently losing the Foxp3 expression and reprogramming their activity towards a Th17 effector profile [In rheumatoid arthritis and periodontitis, the presence of unstable Treg lymphocytes that have lost the expression of Foxp3 and their regulatory capacities have been described , 77. The profile . In this profile , 80. Thu+ T lymphocytes has been negatively associated with bone formation and regeneration (+CD28-CD45RA+ CD57+ T lymphocytes inhibit osteoblast differentiation and human mesenchymal stromal cell survival, leading to a fewer number of osteoblast precursors [+CD28-CD45RA+CD57+ T lymphocytes have been identified as the main producers of pro-inflammatory cytokines, such as TNF-α and IFN-γ, during bone fracture healing failure [+CD28+ T lymphocytes decreases and associates with the downregulation of Wnt production [+CD28-CD45RA+CD57+ T lymphocytes is altered, which could also affect the process of bone formation. Additionally, CD8+ T lymphocytes must be in an anergic state to produce Wnt10b [+ T lymphocytes to an anergic state, affecting osteoblastogenesis and bone formation. Indeed, animal models with deficient Treg function show reduced proliferation, differentiation, and lifespan of osteoblasts, and consequently, less bone formation mediated by CD8+ T lymphocytes [The senescence of CD8neration . Senesceecursors . In addi failure . Indeed, failure . Furtheroduction ; howevere Wnt10b ; thus, sphocytes .+ and CD4+ T lymphocytes [+ T lymphocytes, p38-MAPK signaling governs SASP and regulates the production of SASP-related cytokines [Phosphorylation of p38-MAPK is increased in both senescent CD8phocytes , 85. In ytokines , 86. In ytokines . These cytokines , 82.+ T lymphocytes, the constitutive phosphorylation of p38-MAPK is determined by a sestrin-mediated intrasensory signaling pathway dependent on the DNA damage [+ T lymphocytes leads to intracellular STAT3 phosphorylation and Th17-type cytokine production [+CD28- T lymphocytes, including TNF-α, IL-17A, and RANKL, could also contribute to osteoclastogenesis and bone loss.In senescent CD4A damage , 85, 87.oduction . All the+ and CD4+ T lymphocytes that contributes to the explaining of the upregulation of SASP-related pro-inflammatory and pro-osteoclastogenic mediators, including RANKL, triggered by the inhibition of autophagy, accumulation of dysfunctional mitochondria, and increased ROS production (Taken together, the p38-MAPK phosphorylation could be a conserved mechanism in senescent CD8oduction .in vitro and in vivo models, including metformin, sestrin blockade, p38-MAPK inhibitors, and TNF-α1-receptor blockade [Nowadays, different therapeutic strategies have been proposed to specifically target cellular senescence. These emerging senotherapeutic approaches comprise senolytic strategies, which seek to eliminate senescent cells, and senomorphic strategies, which can partially reverse the senescent phenotype, including the establishment of SASP . Althougblockade , 88, 90.+ T lymphocytes obtained from elderly people, metformin rescued autophagy and mitochondrial function, inhibited the production of ROS, and downregulated the production of Th17-associated cytokines, IL-6, IL-17A, and IL-21 [+ T lymphocytes reduced the damaged DNA accumulation and enhanced the cell proliferative potential and telomerase activity, while its blockade in senescent CD8+ T lymphocytes negatively regulated natural killer functions [+ T lymphocytes significantly improved telomerase activity and cell survival after TCR activation, while its inhibition in senescent CD8+ T lymphocytes increased autophagic activity, restored mitochondrial function, and downregulated the TNF-α expression [+ T lymphocytes increased their proliferative potential, delayed the loss of CD28 expression, and improved telomerase activity [In CD4nd IL-21 . In addiunctions , 39. Simpression , 90. Finactivity . HoweverWith age, the inherent cellular mediated immunity flaws related to inflamm-aging increase the vulnerability of the immune system, which can lead to defective bone homeostasis. In addition, senescent T lymphocytes can also be generated prematurely during chronic inflammatory diseases regardless of chronological aging. In the context of osteoimmunology, the activity of T lymphocytes with senescent phenotype contributes to a novel understanding of the cross-communication between the immune response and bone metabolism. During osteolytic diseases, premature senescence of T cells can be crucially involved in the increased activity of RANKL-producing Th17 and exFoxp3Th17 lymphocytes, which cause the Th17/Treg imbalance and bone loss. These new findings uncover the need for the exploration of innovative therapeutic strategies focused on the control of immunosenescence related pathological bone loss. In fact, considering that the loss of CD28 expression and the SASP-related RANKL production are distinctive features of premature senescent T lymphocytes during osteolytic diseases, these could be a focus of therapeutic attention since these changes could potentially be reversible.


In recent years, there has been a growing interest in studying and exploring the potential health benefits of foods, mainly from vegetables and fruits from regular intake. The presence of secondary metabolites, namely polyphenols, carotenoids and terpenes, in certain food matrices seems to contribute to their functional properties, expressed through an increased prevention in the development of certain chronic diseases, namely coronary heart diseases, neurodegenerative diseases, cancer and diabetes. However, some foods’ volatile secondary metabolites also present important bioactive properties, although this is a poorly scientifically explored field. In this context, and in order to explore the potential bioactivity of volatile metabolites in different vegetables and fruits from regular consumption, the volatile composition was established using a green extraction technique, solid phase microextraction in headspace mode (HS-SPME), combined with gas chromatography tandem mass spectrometry (GC-MS). A total of 320 volatile metabolites, comprising 51 terpenic compounds, 45 organosulfur compounds, 31 aldehydes, 37 esters, 29 ketones, 28 alcohols, 23 furanic compounds, 22 hydrocarbons, 19 benzene compounds, 13 nitrogenous compounds, 9 carboxylic acids, 7 ethers, 4 halogenated compounds and 3 naphthalene derivatives, were positively identified. Each investigated fruit and vegetable showed a specific volatile metabolomic profile. The obtained results revealed that terpenic compounds, to which are associated antimicrobial, antioxidant, and anticancer activities, are the most predominant chemical family in beetroot (61%), orange carrot (58%) and white carrot (61%), while organosulfur compounds  are dominant in onion, garlic and watercress. Broccoli and spinach are essentially constituted by alcohols and aldehydes , while fruits from the Solanaceae family are characterized by esters in tamarillo and aldehydes in tomato. The exponential increase in the world’s population , combined with continuous climate changes and consequent decreasing of agricultural areas due to desertification, strengthened by the global crisis caused by COVID-19, constitutes societal and global concerns that challenge the food sector. Humanity depends heavily on plants, both as a food source and as a natural source of bioactive compounds with health-promoting properties. In the last few decades, the global interest in plant-based foods, namely fruits and vegetables, has increased due to the potential ability of bioactive compounds to scavenge free radicals (ROS and RNS) in addition to their antiproliferative, antimicrobial and anti-inflammatory activities ,7,8. HerThere are three major groups of secondary metabolites originated by the biosynthesis of primary metabolism: (i) the group of terpenes derived from malonic acid or pyruvate and 3-phosphoglycerate; (ii) the group of nitrogen compounds derived from aromatic amino acids, such as tryptophan and tyrosine and aliphatic amino acids such as ornithine and lysine; and finally (iii) the group of phenolic compounds  ,12.−1 to a few ng L−1, or even less), which means that an effective extraction technique and a sensitive methodology of analysis are required for their adequate characterization [Typically, these compounds are found in fruits and vegetables in small amounts, varying from species to species, and sometimes between varieties in its chemical nature . Accordirization . Solid prization . Moreoverization . This terization . HS-SPMErization ,30,31. Trization .In the present study, we report the use of HS-SPME/GC-qMS as a reliable methodology for the establishment of the volatile composition of fruits (tomato and tamarillo) and vegetables  from regular intake, as a useful strategy to identify volatile metabolites with potential benefit effects on human health. Although these effects are mainly attributed to other types of secondary metabolites—polyphenols, carotenoids, phytosterols, glucosinolates, among others—many volatile compounds are associated to health benefits according to reported in several studies ,34.DVB/CAR/PDMS/GC-qMS.The typical aroma of fruits and vegetables depends on a large number of volatiles and the synergic effects, the chemical nature, and its relative amount and odor threshold, which can be related to the fruit or vegetable composition as well as to ripening and processing conditions. A total of 320 volatile metabolites, including 51 terpenic compounds, 45 organosulfur compounds, 31 aldehydes, 37 esters, 29 ketones, 28 alcohols, 23 furanic compounds, 22 hydrocarbons, 19 benzene compounds, 13 nitrogenous compounds, 9 carboxylic acids, 7 ethers, 4 halogenated compounds and 3 naphthalene derivatives, were identified. Beta vulgaris L.) belongs to the Beta genus and to the Amaranthaceae family. A total of 61 volatile compounds were identified, being mainly characterized by terpenic compounds (61.0%), followed by furanic compounds (20.6%), carboxylic acids (5.6%) and benzene derivatives (5.2%). The principal volatile constituents of beetroot include terpinolene (89), γ-terpinene (75) and 5-hydroxymethylfurfural (261), representing 65.6% of the total volatile profile. In lower amounts are 1-methyl-2-(1-methylethyl)-benzene , 5-methylfurfural , β-pinene  and α-pinene . It is noteworthy that among the volatile compounds, it was possible to identify geosmin (267), one of the main characteristic compounds of the beetroot aroma. This sesquiterpene is often considered as an off flavor, being responsible for an earthy odor and flavor [Beetroot  and γ-terpinene (75) being the major volatiles found in white carrot (32.0%) and orange carrot (27.2%), respectively. Manach et al.  demonstrThe volatile profile of both (red and yellow) onion varieties is very distinct. Red onions are characterized by several chemical groups, namely aldehydes (26.7%), organosulfur compounds (19.6%) and carboxylic acids (15.3%), while yellow onions are characterized essentially by organosulfur compounds, which account for 73.8% of its total volatile fraction .The characteristic flavor and the bioactive properties of onions have been mainly attributed to the organosulfur volatile compounds , which aAmong the main organosulfur compounds identified in the studied onion varieties, dipropyl disulfide , dimethyl trisulfide  and dipropyl trisulfide  are predominant.Another class of volatile compounds commonly detected in fresh onion is volatile aldehydes , with 2-Like onions, garlic is rich in organosulfur compounds that are responsible for the characteristic odor and taste of this vegetable, as well as bioactive properties ,48. Of tDiallyl disulfide (175) is the major compound found in the investigated garlic samples, contributing to 70.4% of the total volatile composition. This is one of the most important organosulfur compounds present in garlic, being formed by the decomposition of allicin, an unstable and highly reactive molecule. It is the most common bioactive compound present in garlic and accounts for about 70% of the sulfur compounds family . AllicinDiallyl trisulfide (260) was also identified in a considerable percentage (12.4%), being the second most dominant volatile in garlic. Both diallyl disulfide and diallyl trisulfide, which are associated with anticancer properties ,54,55, aZ)-3-hexen-1-ol (136), hexanal (38), heptanal (62), nonanal (133) and pentanal (21) [Z)-3-Hexen-1-ol and hexanal contribute with 43.1% and 10.3%, respectively, for the total volatile composition. Broccoli is also characterized by organosulfur compounds, namely dimethyl disulfide (36) and methyl thiocyanate (99) formed from glucosinolates and amino acid precursors [It was possible to identify a total of 88 volatile metabolites in broccoli samples, of which 59% were alcohols and 26% aldehydes, the most representative chemical classes. The volatiles with the greatest influence on the volatile composition of the broccoli are  . (Z)-3-Hecursors ,58. As dAccording to Kebede et al. , the maiRegarding the characterization of the volatile profile of spinach, 57 volatile metabolites were identified, mainly 14 esters, 13 terpenic compounds, 8 alcohols and 7 aldehydes. Higher alcohols are the most dominant chemical group, accounting for 43.5% of the total volatile fraction, followed by esters (25.6%), aldehydes (16.5%) and terpenic compounds (10.9%).E)-2-hexen-1-ol (148), 2-hexenal (76) and 1-hexanol (126), which together account for 56.8% of the total volatile fraction. These volatile compounds are derived from C18 unsaturated fatty acids such as linoleic acid and/or α-linolenic via the lipoxygenase/hydroperoxide lyase pathways [C6 alcohols and aldehydes, with an odor similar to cut grass, are typical compounds produced by green leafy vegetables, such as spinach and esters . The majpathways . Once deZ)-3-hexenal. The latter is an unstable compound and is rapidly isomerized to (E)-2-hexenal by (3Z):(2E)-enal isomerase [E)-2-hexenal and (E)-2-hexen-1-ol volatiles have higher activity than hexanal saturated volatiles and 1-hexanol.C6 aldehydes responsible for the ‘herbaceous’ aroma are formed from 13-hydroperoxides and include hexanal and  is rich in folate, vitamin C, and potassium. Carotenoids, namely lycopene, the most prominent carotenoid, followed by β-carotene, γ-carotene and phytoene, as well as several minor carotenoids, are the most abundant phytonutrients, in addition to a wide variety of volatile compounds. Of the 77 identified volatiles, aldehydes and furanic compounds constitute the most predominant classes, representing about 71% of the total volatile profile. The remaining identified groups contributing to the aroma and taste of this sample were alcohols (6.6%), esters (5.9%), terpenic compounds (5.5%), carboxylic acids (4.7%), organosulfur compounds (2.9%) and ketones (1.6%). The remaining chemical families identified contribute less than 1% to the total volatile profile of tomato.Tomato (Z)-3-hexenal (49), 3-methyl-butanal (11), (E)-3-hexen-1-ol (128), β-ionone (289) and 2-isobutylthiazole (138), are considered the most important volatile components for defining the aroma of tomatoes, giving a fresh character to this matrix [The individual compounds identified in this study, hexanal (38), , it was possible to identify a total of 65 volatile metabolites, of which 20 were terpenic compounds, 17 esters, 7 alcohols, 5 benzene compounds, 4 aldehydes, 4 furan compounds and the remaining 7 miscellaneous compounds. Ethyl esters were the most important family (65.4%), followed by terpenic compounds (15.8%). Considering the individual metabolites , ethyl bThe biological activities and role of many volatile metabolites, so far identified, are partly known. Many of these compounds have proven to play a key role in the prevention and treatment of several diseases such as cancer , inflammAmong these metabolites, terpenic compounds are one of the most dominant chemical classes found in fruits and vegetables, with several biological effects including antimicrobial, antifungal, antiviral, anti-hyperglycemic, anti-inflammatory, and antiparasitic effects . Table 1An interesting study about the potential antioxidant activity of monoterpene and sesquiterpene compounds demonstrated that monoterpene hydrocarbons, those with strongly activated methylene groups in their structure such as terpinolene, α-terpinene, γ-terpinene and sabinene, were the most active compounds .Other studies have proved the anticancer properties of several terpenoid compounds acting on different stages of tumor development, such as inhibition of the early initiation and progression of tumorigenesis by inducing cell cycle arrest, tumor cell differentiation, and apoptosis, and in the late stages, the suppression of angiogenesis, invasion, and metastasis through the regulation of various intracellular signaling pathways .Another chemical family, identified in the investigated samples, with interesting biological properties is organosulfur compounds. These compounds are highly reactive phytochemical metabolites, usually found in cruciferous vegetables such as broccoli, cauliflower, brussels sprouts, garlic and onions . Organos−1 in n-hexane) supplied from Fluka .All reagents used were analytical quality. Sodium chloride  used to obtain the adequate ionic strength was supplied by Panreac . Ultrapure water (18 MΩ cm at 23 °C) was obtained from a Milli-Q system . The glass vials, SPME fiber coated with divinylbenzene/carboxen on polydimethylsiloxane (DVB/CAR/PDMS) and SPME holder for manual sampling were purchased from Supelco . The retention index (RI) was calculated through injection of a series of C8 to C20 straight-chain n-alkanes (concentration of 40 mg LTriplicates (1 kg) of different vegetables from regular consumption, namely tomato (Solanum lycopersicum), tamarillo (Solanum betaceum), watercress (Rorippa nasturtium-aquaticum), broccoli (Brassica oleracea), spinach (Spinacia oleracea), white carrot and orange carrot (Daucus carota), beetroot (Beta vulgaris), yellow onion and red onion (Allium cepa), and garlic (Allium sativum), were analyzed. The samples were purchased in a commercial area at Funchal, Madeira Island, Portugal. Vegetables with unpleasant appearance were discarded. Each sample was washed, and all inedible parts removed. After trituration to obtain the pulps, the samples were stored at −20 °C in the dark until analysis.The application of an effective HS-SPME method generally requires the optimization of the main experimental factors that influence the extraction process and the experimental response. HS-SPME was performed based on the conditions described by  and Peren = 3). It is important to note that the fiber was conditioned according to the manufacturer’s instructions before use and before the first daily extraction.The SPME fiber (DVB/CAR/PDMS) was exposed to the headspace for 45 min. After extraction, the fiber was withdrawn into the holder needle, removed from the vial and immediately introduced into GC injector port for 6 min at 250 °C for thermal desorption of the VOCs and SVOCs. All analyses were carried out in triplicate (−1 (column-head pressure: 13 psi). The injector temperature was fixed at 250 °C and a splitless injector equipped with an insert of 0.75 mm i.d. was used. The temperature program was set up as follows: initial temperature 40 °C for 1 min, 1.2 °C × min–1 ramp until 80 °C, held for 2 min, 3 °C × min–1 ramp until 150 °C, held for 2 min, 40 °C × min–1 ramp until 220 °C and then held isothermally at 220 °C for 10 min. MS detection was performed in full scan in an Agilent 5975 quadrupole inert mass selective detector, the ion energy used for the electron impact (EI) was 70 eV and the source temperature was 230 °C. The electron multiplier was set to the autotune procedure. The mass acquisition range, made in full scan mode, was 30–300 m/z.Chromatographic separations of VOCs and SVOCs from fruits and vegetables were performed using an Agilent Technologies 6890N  gas chromatography system equipped with a BP-20 fused silica capillary column (30 m × 0.25 mm i.d. × 0.25 μm film thickness) supplied by SGE  with helium  as carrier gas at a flow rate of 1 mL × minVolatile metabolite identification was achieved by comparing the retention times (RT) of the chromatographic peaks with those, when available, of authentic standards (ST) run under the same conditions. For metabolites whose reference substances were not available, the identification was performed by matching their retention indices (RI) determined relative to the retention time of a series of n-alkanes (C8–C20) with linear interpolation, with those of literature data, and by comparing their mass spectra using the National Institute of Standards and Technology (NIST) MS 05 spectral database .The characterization of the volatile composition of fruits and vegetables from regular consumption was carried out using the HS-SPME/GC-qMS methodology. A total of 320 volatile compounds, belonging to different chemical families, were identified. According to the results obtained, it was found that terpenic compounds were predominant in beetroot and in both carrot varieties (orange and white), whereas organosulfur compounds were found almost exclusively in garlic, watercress and in both onion varieties (red and yellow). These chemical compounds are responsible for the bioactive properties, such as antihypertensive, anti-inflammatory, antibacterial, and anticancer properties, among others, of numerous plants.The data presented here showed the potential of a solvent-free and high-throughput extraction technique to establish the volatile metabolomic pattern of different fruits and vegetables, allowing the establishment of its relationship with their bioactive activities. The knowledge of volatile composition is a valuable tool to improve the aroma quality and the nutritional value of the fruits and vegetables, as well as to identify which metabolites are biologically the most important and can be used for specific purposes as nutraceuticals. The study intends to draw attention to the importance of volatile compounds that constitute fruits and vegetables in human health. Although the vast majority of work on this thematic is directed to polyphenolic compounds, flavonoids and non-flavonoids, powerful bioactive effects are also attributed to volatile compounds, with a great impact on the prevention of certain pathologies such as cancer and cardiovascular and neurodegenerative diseases.


SOD2 (superoxide dismutase 2) as a potential CD44-downstream transcriptional target involved in BC metastasis. SOD2 gene belongs to the family of iron/manganese superoxide dismutase family and encodes a mitochondrial protein. SOD2 plays a role in cell proliferation and cell invasion via activation of different signaling pathways regulating angiogenic abilities of breast tumor cells. This review will focus on the findings supporting the underlying mechanisms associated with the oncogenic potential of SOD2 in the onset and progression of cancer, especially in BC and the potential clinical relevance of its various inhibitors.CD44, a cell-adhesion molecule has a dual role in tumor growth and progression; it acts as a tumor suppressor as well as a tumor promoter. In our previous work, we developed a tetracycline-off regulated expression of CD44’s gene in the breast cancer (BC) cell line MCF-7 (B5 clone). Using cDNA oligo gene expression microarray, we identified  Tumor cell invasion is the most defining and recurring event during the multistage process of metastasis. It involves the activation of a complex molecular network involving, involving, in particular, cell adhesion molecules (CAMs) ,2 which In order to better understand the exact function of the standard form of CD44 (CD44s) in BC and further elucidate the signaling mechanisms that underpin CD44-promoted BC invasion/metastasis, we have previously developed a tetracycline (Tet)-Off-regulated expression system of CD44s both in vitro  and in vSOD2 was significantly upregulated upon interaction of CD44 with its major ligand hyaluronan (HA), suggesting that SOD2 might be an additional novel transcriptional target of CD44-downstream signaling that underpins its role in promoting BC tumor cell invasion and metastasis.Among the remaining genes, SOD2 encodes the manganese superoxide dismutase (Mn-SOD), which is involved in regulating oxidative stress in the cell through the dismutation of superoxide radicals (O2−) to H2O2 and molecular oxygen [r oxygen ,16,17. Rr oxygen ,19. Damar oxygen . Alteratr oxygen .SOD2, and SOD3. While SOD1 is localized in the cytoplasm and the mitochondrial intermembrane space, SOD2 is exclusively present in the mitochondrial matrix, and SOD3 is extracellularly present [SOD2 have been found to be associated with many diseases, including neurodegeneration, mitochondrial dysfunction, premature aging, angiogenesis, and cancer [The SOD family encompasses three members, SOD1,  present . Geneticd cancer ,25,26,27SOD2 might be a novel transcriptional target of CD44-downstream signaling, which promotes BC cell invasion. We have discussed these lines of evidence and proposed a model of the signaling pathways linking CD44 activation by HA to the transactivation of SOD2 to promote breast tumor cell invasion.This review presents data from the literature supporting our hypothesis that SOD2 gene, located on the long arm of chromosome 6 , consists of 5 exons and 4 introns [SOD2 promoter lacks the typical TATA and CAAT boxes; it is, however, rich in GC sequences as well as binding sites for specificity protein (SP) and activator protein 2 (AP2) transcription factors [SOD2 expression. Sp1 maintains SOD2’s basal promoter activity, whereas Nf-kB increases SOD2 transcription mostly through a cis-element inside the second intronic enhancer region [SOD2 mRNA shows a conserved 41 bp translational enhancer region [The human  introns ,17 encod introns ,28,29. Tr region . Additior region .SOD2 protein is a homo-tetramer , with each of the four subunits having a molecular mass of around 23 kDa, and each including a manganese (Mn+3) cofactor [SOD2 starts with a 26 amino acid mitochondrial targeting sequence (MTS), which is vital for mitochondrial localization [SOD2 gene polymorphisms, which result in a substitution of Ala-9 to Val  [SOD2 MTS, and have been linked with susceptibility to various pathologies [The cofactor . The prelization . For exaVal9Ala) , or Ala-hologies ,33.SOD2 monomer consists of two unique domains, an N-terminal helical hairpin domain and a C-terminal a/b domain. The C-terminal a/b domain contains five-alpha helices and three-stranded antiparallel beta-sheets. Together, these domains encompass key residues , allowing the active site to interact with the Mn³+ cofactor and a water molecule [Each mature molecule , which aSOD2 is expressed in both normal and malignant cells, thus, the following sections will discuss the role of SOD2 in both contexts.Physiologically, SOD2 is ideally localized [SOD2 catalytic site triggers the enzyme to carry out its dismutase activity [While aerobic respiration produces low amounts of ROS, cells utilize SODs to prevent these molecules from causing damage. SODs provide such a mechanism, since most of the ROS is produced by the electron transport chain in the mitochondrial matrix of human cells, where ocalized . Integraactivity ,16, and activity ,37 and pactivity .SOD2 can act either as a tumor suppressor gene (TSG) or as an oncogene [Various lines of evidence lead to the conclusion that oncogene .SOD2 activity are found during different cell cycle stages [SOD2 expression in pancreatic cancer cells, as well as glioma cells, and SOD2 results showing significantly delayed tumor cell growth in nude mice xenograft studies [SOD2 expression enhances cell cycle progression (mitosis) by increasing O2•− levels [Variations in e stages ,41,42; a studies ,44. Such− levels , thus st− levels ,47.SOD2 in cancer. Mutations in the SOD2 gene promoter have been reported to impact the AP2 binding pattern in colorectal cancer cells, resulting in loss of SOD2 expression [SOD2 expression [SOD2 promoter methylation was reported in both transformed and cancer cells [SOD2 was found to result in reduced SOD2 expression [Different mechanisms have been proposed for the loss of pression . Some evpression ,41,42. Ser cells ,50, and pression . Such mepression ,50,51,52SOD2 expression is commonly reported in the early stages of tumor growth, some studies report elevated levels of SOD2 during metastatic progression [SOD2 was overexpressed in metastatic tumor lesions and aggressive tumor cell lines. Upregulated SOD2 levels enhance the oxidation of PTEN and PTP-N12, further activating Akt and p130 cas/Rac1 signaling pathways, respectively [SOD2 in the onset and progression of cancer.While reduced gression ,53,54. Sectively ; these pectively ,56,57,58SOD2 in different cancers.Accumulation of free radicals is associated with mitochondrial function and metastatic disease progression. Although, ROS play a role in tumor initiation and progression, the underlying mechanisms and causes of enhanced ROS accumulation during tumorigenesis still lie nascent.SOD2 to promote tumor cell invasion/metastasis  activates the transcription of tastasis .SOD2 expression increases cellular oxidant/antioxidant ratios, which are directly associated with tumor progression, angiogenesis and migration, and invasion [SOD2 expression is elevated during metastasis [SOD2 expression in metastatic cancer cell lines as well as in tumor tissues is associated with increased SOD2 activity, thus, indicating a functional role for SOD2 during metastatic progression and poor outcome, particularly in BC [SOD2 expression is associated with metastatic progression in estrogen receptor (ER) negative BCs [invasion ,87. In stastasis ,91,92,93ly in BC ,94,95,96SOD2 in BC [SOD2 promoter, in ER-negative BC cells (MDA-MB-231), the lack of DDB2 induced high levels of SOD2 [SOD2 expression was regulated by progestin [The transcriptional repressor DDB2 plays an important role in the dual nature of D2 in BC . While i of SOD2 . In the rogestin .SOD2 expression in a TNFα-dependent manner [SOD2 promoter, thus inducing SOD2 expression [SOD2 gene expression in cells containing high levels of CD44 (CD44H), which leads to the induction of epithelial-mesenchymal transition mechanism [Additionally, oxygen-free radicals and ionizing radiation have been shown to stimulate t manner ,99, as at manner . Brieflypression ,102. NF-echanism .SOD2 in tumor cells correlates with enhanced expression and activity of MMPs, leading to enhanced matrix degradation and a release of cytokines and growth factors, thereby promoting metastasis [SOD2 expression, and activated MMP2 in parallel [SOD2 in HT-1080 fibrosarcoma, as well as 253J bladder carcinoma cells, significantly promoted migration and invasion both in vitro and in vivo [SOD2 expression is a significant reduction in catalase activity, resulting in increased H2O2 production [Overexpression of tastasis . For exaparallel . Similar in vivo ,92,106. oduction , which hoduction , and migoduction .SOD2 [SOD2 is essential for the stimulation of MMP-1 as well as other MMP family members via the H2O2-dependent activation of MAPK signaling [Further, binding of Ets-1 to the MMP-1 promoter and sustained JNK signaling requires the enzyme activity of SOD2 ,110. Thuignaling ,110,111,ignaling .SOD2 was significantly upregulated upon the interaction of CD44 with its major ligand HA [SOD2 might be a novel transcriptional target of CD44-downstream signaling, and thus, have a critical role in promoting BC tumor cell invasion and metastasis.The above, along with our findings that igand HA ; prompteSOD2 is correlated with increased tumor cell invasion, metastasis, proliferation, and resistance to apoptosis [SOD2 as they are involved in common signaling pathways promoting metastasis, cell migration, cell invasion, and angiogenesis [SOD2 at RNA levels. More interestingly, inhibition of CD44 using specific siRNA significantly decreased RNA expression levels of SOD2. Ongoing functional, molecular, and pharmacological experiments aim to validate the physiological relevance of the CD44/SOD2 signaling pathway in BC cell invasion and further identify its intermediate molecular players. As a matter of fact, elevated content of poptosis . Thus, togenesis . Our iniThere is a dire need to develop anticancer therapeutics targeting ROS signaling, and hence, use of SOD mimetics that simulate endogenous SOD enzyme activity or inhibit oxidative injury—biomarkers of cell proliferation are being explored.SOD2 promoter methylation in the KAS6/1 human multiple myeloma cells [SOD2 expression by regulating histone methylation and acetylation in breast cancer cells [The epigenetic drug zebularine was found to effectively reverse ma cells . In addier cells ,114. Syner cells ,117,118.SOD2 adenoviral vectors in combination with BCNU significantly reduced breast cancer cell growth and increased nude mice survival [SOD2 and TRAIL-induced apoptosis inhibited colorectal tumor growth in xenograft models [Adenoviral vectors, in combination with chemotherapeutic agents have also been used to induce SOD expression. In breast tumor xenograft models, injection of Cu-ZnSOD or survival . Moreovet models .As mentioned above, elevated ROS stress in cancer cells is an unfavorable event associated with increased cancer progression and carcinogenesis. Recently, research has identified drugs capable of increasing cellular ROS accumulation to eliminate cancer cells ,122. SinSOD2 a good potential target. Various drugs are known to trigger ROS generation and accumulation; these include mitochondrial electron transport chain modulators , redox-cycling compounds (motexafin gadolinium), glutathione (GSH) depleting agents ), and inhibitors of SOD (2-methoxyestradiol) and catalase  [Promoting ROS accumulation in cancer cells to a toxic level is the focus of some emerging cancer therapies, which makes riazole) ,127,128.Rapamycin and its analogs are being used in clinical trials for certain types of cancer as rapamycin alone can inhibit cell proliferation and induce apoptosis ,130. MorSOD2 expression, resulting in a decrease of ROS-levels in cancer cells, thereby promoting angiogenesis and metastasis [FOXM1 in combination with oxidative stress is considered as an essential therapeutic strategy against cancer. Proteasome inhibitors (MG132) as well as thiazole antibiotics (siomycin-A) reduce FOXM1 expression and activity and induce apoptosis through DNA damaging agents, including doxorubicin and γ-irradiation [FOXM1 is a transcription factor that is over-expressed in the majority of human tumors. It can induce tastasis ,136. Taradiation ,138,139.In contrast to the above, the use of antioxidants has also shown some positive results. These often take the form of traditional therapeutics—natural occurring antioxidants. Natural products including curcumin ,141,142,SOD2 expression and delayed bile acid-enhanced esophageal injury and cancer progression [In one study, curcumin oil was found to maintain gression . It has gression .Astaxanthin, a xanthophyll, possesses anti-inflammatory properties and exerts its effect via ROS/reactive nitrogen species scavenging by inhibiting the expression of inflammatory cytokines  .The natural dietary element, ginger has both antioxidant and anti-carcinogenic properties: ginger supplementation enhanced SOD, CAT, GPx, glutathione-S-transferase, and glutathione reductase activities, thus reducing oxidative stress as well as cancer cell growth .SOD2 gene expression, glycolysis inhibition, and reduced tumor growth of transplanted human breast tumors in vivo [Oroxylin A, a flavonoid extracted from a Chinese medicinal plant, can induce  in vivo .One of the dietary combination supplements, protandim, was shown to be effective in inhibiting tumor progression as well lipid peroxidation . ProtandThis indicates the use of antioxidants to target ROS generation.SOD2-based therapeutic approaches.SOD2 as a factor in cancer progression. The ability to use SOD2 to modify the cellular redox environment can help with the identification of redox-responsive signaling events that stimulate malignancy, such as invasion, migration, and prolonged tumor cell survival. Additional studies of these redox-driven events will help in the development of targeted therapeutic strategies to efficiently restrict redox-signaling essential for malignant progression. In particular, results from our own work and others support our hypothesis that CD44-HA interaction can transactivate SOD2, ultimately leading to BC progression.Several studies have suggested the complicity of antioxidant enzymes in cancer development and identified high expression of


APOE4) is the primary genetic risk factor for the late-onset form of Alzheimer’s disease (AD). Although the reason for this association is not completely understood, researchers have uncovered numerous effects of APOE4 expression on AD-relevant brain processes, including amyloid beta (Aβ) accumulation, lipid metabolism, endosomal-lysosomal trafficking, and bioenergetics. In this study, we aimed to determine the effect of APOE4 allelic dosage on regional brain lipid composition in aged mice, as well as in cultured neurons. We performed a targeted lipidomic analysis on an AD-vulnerable brain region  and an AD-resistant brain region  from 14–15 month-old APOE3/3, APOE3/4, and APOE4/4 targeted replacement mice, as well as on neurons cultured with conditioned media from APOE3/3 or APOE4/4 astrocytes. Our results reveal that the EC possesses increased susceptibility to APOE4-associated lipid alterations compared to the PVC. In the EC, APOE4 expression showed a dominant effect in decreasing diacylglycerol (DAG) levels, and a semi-dominant, additive effect in the upregulation of multiple ceramide, glycosylated sphingolipid, and bis(monoacylglycerol)phosphate (BMP) species, lipids known to accumulate as a result of endosomal-lysosomal dysfunction. Neurons treated with conditioned media from APOE4/4 vs. APOE3/3 astrocytes showed similar alterations of DAG and BMP species to those observed in the mouse EC. Our results suggest that APOE4 expression differentially modulates regional neuronal lipid signatures, which may underlie the increased susceptibility of EC-localized neurons to AD pathology.Apolipoprotein E ε4 ( APOE) is the primary genetic risk factor for late-onset Alzheimer’s disease (AD). While early work investigating the association between APOE ε4 (APOE4) and AD focused on the ability of APOE4 to increase the aggregation and decrease the clearance of amyloid beta (Aβ) . It views by –14, a fupression –19. Impopression , and spepression . Moreovepression , 23.APOE isoform-related lipid alterations in plasma and serum  were elevated with increasing APOE4 alleles. Importantly, while BMP is exclusively found in the intraluminal vesicles (ILVs) of late endocytic compartments and is considered a bona fide marker of these structures [Specifically, we observe that in the EC, several neutral lipid species were affected by ion Fig. , includiructures , we haveAPOE4 expression, while BMP 34:2 was significantly increased , and which features are associated with a dominant APOE4 effect . To better understand the effect magnitude of each APOE4 allele on lipid signatures, we performed paired comparisons between the three APOE genotypes  and sphingolipids (SL) in the EC and PVC.The chemical structures of lipids are a determinant of their physical/chemical properties, which in turn determines the biological roles that they play in cells and tissues. While lipid categorization is largely determined by its polar head group, its acyl chain structure further contributes to membrane compositional diversity, which differs greatly between tissues and subcellular compartments . ImportaAPOE4 on decreasing the levels of short-length DAG species in the EC, no global impact was observed in the acyl chain length for DAG/GPL or for SL in APOE3/4 or APOE4/4 mice, as compared to APOE3/3 mice  in the EC [Despite the dominant effect of n the EC , our resAPOE4/4 vs. APOE3/3 expression in astrocytes (the primary source of apoE protein in the brain) on the lipid composition of wild-type, B6 cortical neurons. Specifically, ACM was collected from immortalized astrocytes generated from post-natal day 1-2 (P1-2) pups of APOE3/3 or APOE4/4 targeted replacement mice [APOE3/3 ACM, or APOE4/4 ACM (n = 6 wells per group) and incubated for 7 days, before harvesting for lipidomic analysis.A present limitation of performing whole tissue lipidomic analysis is the inability to discern the corresponding contribution of each cell-type present within that tissue. To overcome this constraint, we used the paradigm of in vitro conditioned media transfer to analyze the effect of humanized ent mice . PrimaryAPOE isoform  and TAG species  were increased  , Acyl-PG , and BMP , similar to what was observed in APOE4/4 brains, while BMP 34:1 levels were decreased  Fig. , whereas:6) Fig. , suggestACM Fig. . We alsosed Fig. . FinallyAPOE4, the primary genetic risk factor for late-onset AD, is itself a regulator of lipid transport and metabolism. Our findings suggest that the EC, one of the first brain regions to develop AD pathology, presents increased susceptibility to APOE4-associated lipid alterations, as many more lipids were altered by APOE4 expression in the EC than the PVC  [APOE4 also increases the risk for dementia in pure synucleinopathies and specifically exacerbates α-synuclein pathology [APOE4 carriers.Our data indicates that ns Figs. , 3, 4. Gisorders , 55. Notisorders , 56, 57.ase (PD) . Since Aathology , 60, we athology  and haveathology , underscAPOE4 in endosomal-lysosomal trafficking, we previously showed increased expression levels of Rab GTPase genes, such as Rab5b, Rab7 and Rab9, in the EC of aged APOE4 mice [APOE4 background, and their individual contributions to pathology or therapeutic potential remain elusive [APOE4 expression in exosomes release [WDR81, SNX1, CSTB, and GRN [APOE4 to endosomal-lysosomal dysfunction in contributing to increased susceptibility to AD and other forms of dementia.In keeping with a prominent effect of OE4 mice , similarOE4 mice . ImportaOE4 mice , Rab7b gOE4 mice , and altOE4 mice ; that sa elusive . Also, w elusive , possibl elusive , 69. Sin elusive , the inh release  may also and GRN . Thus, oAPOE4 on DAG levels, which was recapitulated when exposing neurons to APOE4 vs. APOE3 ACM  or co-existing gain-of-function effects  [APOE4 ACM on the accumulation of BMP, comparatively to APOE4 mice, may result from the interaction with endogenous neuronal murine apoE within the endolysosomal compartments.Another key finding we observed was a dominant effect of 3/3 mice , which ctrocytes . Intereslization , 74. Higlization , 76. As lization , decreas−/− mice , as wellablation . Howevergnaling) . The milAPOE4 ACM on increasing PA and decreasing PC and PCe levels in the ACM-treated neurons  breakdown in the hippocampus [APOE4 carriers [APOE4 are a cause or a consequence of BBB breakdown is still an intriguing unanswered question. Further caution is also warranted due to the observation that lipidome complexity increases from mice to humans [APOE lipid effects may diverge between mice and humans [While our results complement previous studies on pocampus , and BBBcarriers  and micecarriers . Whethero humans , and APOd humans .APOE4-associated lipid alterations and implicate endosomal-lysosomal lipid flux and lipid droplet regulation as potential factors in the increased risk of AD and other forms of dementia among APOE4 carriers. These findings support a role for lipid dyshomeostasis in selective vulnerability to AD pathology and highlight the therapeutic potential of lipid modulation in neurodegeneration.In summary, our findings demonstrate increased susceptibility of the EC to Summplementary InformationExtended Data File


Gastric cancer (GC) is a disease that threatens human health. It is thus crucial to clarify the mechanisms involved in GC development and discover diagnostic biomarkers and therapeutics. As a cancer stem cell marker, aldehyde dehydrogenase 1 (ALDH1) is involved in the development, progression, and treatment of GC. This review evaluated the prognostic value of ALDH1 and explored its mechanism of action in GC. Importantly, ALDH1 is an informative biomarker in clinical practice as it has specific relationships with indicators, such as metastasis and overall survival. Additionally, ALDH1 interacts with genes and exhibits properties that mimic stem cell characteristics amongst other mechanisms employed in the occurrence and progression of GC. Our results, therefore, provide evidence of possible clinical utility of ALDH1 as a GC therapeutic target. Gastric cancer (GC) ranks fifth in morbidity and is the third leading cause of death, with 1.0 million diagnosed cases and 783,000 deaths worldwide in 2018 . In a laγ-aminobutyric acid, which regulate cellular homeostasis. In addition, ALDHs also oxidize a series of endogenous and exogenous aldehydes to carboxylic acids, protecting them from oxidative stress in living organisms. In stem and progenitor cells, ALDH is derived from normal tissues and its expression is enhanced. Of note, ALDHs are involved in clone growth, self-renewal, drug resistance, and tumor-initiating capacity [Cancer stem cells (CSCs) are involved in the self-renewal, asymmetrical division, differentiation, progression, and resistance of cancer cells , 7. Notacapacity .As a vital stem-like cell marker, ALDH1 is involved in the oxidation and detoxification of malignant tumors . AlthougSince 2012, many researchers worldwide have focused on the relationship between ALDH1 expression and GC. In multiple retrospective studies, patients who were pathologically diagnosed with GC but were not receiving radiotherapy or chemotherapy were selected, and gastritis and para-cancer tissues were used as controls. In GC, para-cancer, and noncancerous tissues, the expression of ALDH1 was measured by immunohistochemistry. Different studies included approximately 36–1072 GC cases, and ALDH1 was expressed in 43.9%–100% of cases –19. ALDHSenel et al. found no statistically significant relationship between the expression of ALDH1 and clinicopathologic parameters, recurrence-free survival, or overall survival (OS) . In addiHelicobacter pylori infection, oncogenes, tumor suppressor genes, abnormal differentiation, unnatural cell cycle regulation, and abnormal signaling pathways [The pathogenesis of GC is overly complex and may be related to pathways . NotablyH. pylori infection. ALDH1 expression and the abundance of Helicobacter pylori were significantly increased in GC tissues than in normal gastric mucosa [H. pylori-positive cases compared to the H. pylori-negative cases and that there was a positive correlation between ALDH1 expression and positive H. pylori staining. Therefore, H. pylori may be associated with the upregulation of ALDH1 [H. pylori-associated gastritis, the number of ALDH1-positive cells increased and contributed to the progression of neoplasia in the gastric mucosa. The age-related increase in cancer stem/stem-like cells in the gastric mucosa may explain the increased incidence of GC during aging [Mao et al. concluded that ALDH1 is positively correlated with c mucosa . Zhao reof ALDH1 . In H. png aging .+ cell group, the colony/tumor formation rates in mice were higher, and the sensitivity to 5-fluorouracil was lower [Zhang reported that the positive rate of ALDH1 expression was 3.21%, 2.76%, and 1.89% in MKN-45, SGC7901, and MKN28 gastric carcinoma cells, respectively . Wang etas lower , 31.Overexpression of ALDH1A1 in MKN-28, a GC cell line, by the recombinant plasmid, pEGFP-N1-ALDH1A1, can increase the proliferation, colony formation, and invasion ability of cancer cells , 33. In ß-catenin in MKN-45 cells. Importantly, ALDH1A1 silencing can decrease cell viability and inhibit cell migration and invasion via regulation of the Wnt signaling pathway in MKN-45 cells [Using small interfering RNA interference, the expression of ALDH1A1 was downregulated in the GC cell line MKN-45. ALDH1A1 can inhibit the proliferation, colony formation, and invasion abilities of these cells and downregulate the expression of matrix metallopeptidases 2 and 9, Wnt1, Wnt5, and 45 cells , 36.These results support that, in GC, ALDH1+ cells have stem cell properties, and the growth of GC cells is repressed on inhibition of ALDH1 expression.Studies on GC have focused on abnormal cell energy metabolism, self-sufficient growth signals, genomic instability and mutation, insensitivity to anti-growth signals, avoidance of immune surveillance, tissue invasion and metastasis, resistance to cell death, continuous angiogenesis, unlimited replication capacity, and promotion of tumor inflammation . Many geβ, a hormone receptor in Choristoneura, is upregulated and promotes cell differentiation in GC, and in one study, ALDH1 and Wnt pathways were inhibited when Rorβ was highly expressed in 200 GC tissues [Ror tissues . Moreove tissues . Further tissues . Progest tissues . A trans tissues .β-catenin signaling by attenuating miR-216a-3p and promoted the stemness of GC cells [Bromodomain and extra-terminal domain protein 4 (BRD4) was dramatically augmented in GC tissues compared to normal adjacent tissues . One stuGC cells . Testis-GC cells . SilenciGC cells . miR-625GC cells . ALDH1+ GC cells . Taken tSurgical treatment after early diagnosis of GC has a good prognosis, but most GCs are diagnosed in the middle and late stages, and the opportunity for surgery is lost. Chemotherapy is currently the predominant therapy of advanced GC; however, there are patients with poor sensitivity to existing chemotherapy drugs. Therefore, it is necessary to identify new therapeutic drugs and treatment regimens that can be used clinically.in vitro. The number of ALDH+ cells dramatically reduced with 1 μM verteporfin treatment in MKN-45 tumor spheres [In MKN-45 tumor spheres, approximately 35% of cells expressed ALDH1, and this percentage was significantly suppressed gradually from 2 to 5 days of all-trans retinoic acid treatment. In diffuse and intestinal types of gastric carcinoma, all-trans retinoic acid appears to repress tumor growth and, more importantly, target GCSCs . Vertepo spheres . In adva spheres . GC cell spheres . 7-Diflu spheres . Taken tALDH1 is highly expressed in GC tissues and cell lines and may be related to differentiation, TNM stage, prognosis, cell growth, and invasion. As a CSC marker, the expression of ALDH1 is inhibited by nucleotides, proteins, or drugs, leading to the inhibition of GC cell growth by inhibiting the characteristics of GC stem cells. ALDH1 inhibitors may also be used in the clinical treatment of GC. Therefore, ALDH1 can be used as a therapeutic target to evaluate the prognosis of GC.