Despite the treatment of lung adenocarcinoma (LUAD) having partially improved in recent years, LUAD patients still have poor prognosis rates. Therefore, it is especially important to explore effective biomarkers and exploit novel therapeutic developments. High-throughput technologies are widely used as systematic approaches to explore differences in expressions of thousands of genes for both biological and genomic systems. Recently, using big data analyses in biomedicine research by integrating several high-throughput databases and tools, including The Cancer Genome Atlas (TCGA), cBioportal, Oncomine, and Kaplan–Meier plotter, is an important strategy to identify novel biomarkers for cancer therapy. Here, we used two different comprehensive bioinformatics analysis and revealed protein tyrosine phosphatase non-receptor type (PTPN) family genes, especially PTPN1 and PTPN22, were downregulated in lung cancer tissue in comparison with normal samples. The survival curves indicated that LUAD patients with high transcription levels of PTPN5 were significantly associated with a good prognosis. Meanwhile, Gene Ontology (GO) and MetaCore analyses indicated that co-expression of the PTPN1, PTPN5, and PTPN21 genes was significantly enriched in cancer development-related pathways, including GTPase activity, regulation of small GTPase-mediated signal transduction, response to mechanical stimuli, vasculogenesis, organ morphogenesis, regulation of stress fiber assembly, mitogen-activated protein kinase (MAPK) cascade, cell migration, and angiogenesis. Collectively, this study revealed that PTPN family members are both significant prognostic biomarkers for lung cancer progression and promising clinical therapeutic targets, which provide new targets for treating LUAD patients. Lung cancer is the leading cause of cancer deaths worldwide, accounting for 18.0% of total cancer deaths . Lung adThe tyrosine phosphorylation of proteins plays a key role in controlling several signaling pathways involved in cell proliferation, apoptosis, migration, and invasion . The proIn the present study, we utilized large-scale bioinformatics databases to identify the expression status of different PTPN members in LUAD patients ,21,22,23The Cancer Genome Atlas (TCGA) level 3 RNA-sequencing (RNA-Seq) together with clinical data acquired from around 30 different types of cancer are combined in UALCAN ,26,27,28The CCLE project was employed to further examine expression level patterns of all PTPN members on a broader scale. This web-based application gave users access to more than 1000 human cancer cell lines and 130 different datasets that are both genetically and pharmacologically characterized ,30,31,32p value of <0.05 was as previously described [The InteractiVenn tool was selected to generate a one-way Venn diagram that presents the overlap and number of genes linked with the expression of PTPN target genes across the two given datasets obtained from TCGA database , and together with the MetaCore platform was employed to map the intersection between these two sets of data in terms of related pathways and involved networks. A escribed ,35,36,37https://kmplot.com/ (accessed on 14 April 2021) which prominently features in assessing target genes in survivors of more than 20 different types of cancer. This tool was subsequently leveraged to expand on LUAD prognosis-related issues, including OS, first progression (FP), and post-progression survival (PPS) [p value [The KM database is an integrated online database available at al (PPS) . The ind[p value ,39,40.PTPN members in various cancers and normal tissue samples  and PTPN21 (6%)  A.PTPN1, PTPN5, and PTPN21  cascade, cell migration involved in sprouting angiogenesis, and positive regulation of angiogenesis. These items may be related to the regulation of cell differentiation and cell migration of LUAD. In the CC group, the most significantly highly enriched items included cytoplasm, focal adhesion, intracellular, cytoskeleton, extracellular matrix, cell cortex, lamellipodium, transforming growth factor beta receptor homodimeric complex, and nuclear outer membrane. As for MFs, changes were mainly enriched in GTPase activator activity, heparin binding, protein binding, calcium ion binding, and lipid binding.Next, we input the genetic data of PTPNs from cBioPortal into Venny 2.0 and calculated the overlap rate of each group. Results revealed that there were 56 genes in common among d PTPN21 B. Moreovd PTPN21 –F. We diPTPN1, PTPN5, and PTPN21, and further analyzed their functions. As shown in PTPN1, PTPN5, and PTPN21 in terms of physical interactions, co-expression, predictions, co-localization, pathways, genetic interactions, and shared protein domains. The top five genes significantly associated with PTPN1 were NOX4 (NADPH oxidase 4), CTH (cystathionine gamma-lyase), JAK2 (Janus kinase 2), TRPV6 , and FCGR2A (Fc fragment of IgG receptor II-a). A further functional analysis showed that these genes were involved in the JAK-STAT pathway, and it was indicated that PTPN1 was related to cell proliferation and cell differentiation , PTPRN (protein tyrosine phosphatase receptor type N), GRIN1 (glutamate ionotropic receptor NMDA type subunit 1), GRIN2B (glutamate ionotropic receptor NMDA type subunit 2B), and MAPK3 (mitogen-activated protein kinase 3). From results of the functional analysis, we discovered that PTPN5 was related to the transmission process of protein kinase signaling (KIF1C (kinesin family member 1C), SRC , NRG3 (neuregulin 3), BMX (BMX non-receptor tyrosine kinase), and PARD6B (par-6 family cell polarity regulator beta) were genes highly associated with PTPN21. According to the functional analysis, we observed that PTPN21 was primarily correlated with the activity of protein tyrosine kinase and cell migration  signaling” (PTPN6 were correlated with “Immune response_T cell co-signaling receptors, schema”, “Breakdown of CD4+ T cell peripheral tolerance in type 1 diabetes mellitus”, and “Chemotaxis_SDF-1/CXCR4-induced chemotaxis of immune cells” (PTPN13 were correlated with “Immune response_HMGB1 release from the cell”, “Putative pathways of hormone action in neurofibromatosis type 1”, and “Immune response_HMGB1/RAGE signaling pathway” (Genes co-expressed with  system” . Genes cgnaling” . Genes ce cells” . Genes cpathway” . Genes cpathway” . Meanwhipathway” , and valpathway”  and GSEApathway” .Lung cancer is composed of SCLC and NSCLC. NSCLC comprises around 85% of lung cancer cases, and it is further classified into three subtypes, squamous-cell carcinoma, adenocarcinoma, and large-cell carcinoma. LUAD is the main subtype of NSCLC, accounting for 40% of all cases of lung cancer. Therefore, it is especially important to explore effective biomarkers and exploit novel therapeutic developments ,43,44,45A previous study demonstrated that PPDPF induces hyperactive STAT3 by interfering with STAT3-PTPN1 interactions in LUAD . The divPTPN3, PTPN7, PTPN12, and PTPN14 were expressed at higher levels, while expressions of PTPN1, PTPN5, PTPN6, PTPN9, PTPN12, PTPN13, PTPN18, PTPN21, and PTPN22 were downregulated in lung cancer. Survival curves revealed that LUAD patients with high transcription levels of PTPN1, PTPN5, PTPN6, PTPN13, and PTPN21 were associated with longer OS. These data suggest that PTPN1, PTPN5, PTPN6, PTPN13, and PTPN21 are associated with clinicopathological features of LUAD.Our bioinformatics analysis indicated that PTPN gene expressions were found to be involve in tumor multi-stage progression. According to various database analyses, PTPN1, PTPN5, and PTPN21 were significantly highly enriched in the positive regulation of GTPase activity, regulation of small GTPase-mediated signal transduction, response to mechanical stimuli, vasculogenesis, response to estrogen, organ morphogenesis, regulation of stress fiber assembly, MAPK cascade, cell migration involved in sprouting angiogenesis, and positive regulation of angiogenesis. The MetaCore analysis of lung cancer patients also suggested that PTPN3 signaling was correlated with “Cell adhesion_Tight junctions”, PTPN5 signaling was correlated with “Development_Transcriptional regulation of megakaryopoiesis”, PTPN6 signaling was correlated with “Development_Transcriptional regulation of megakaryopoiesis”, PTPN13 signaling was correlated with “Immune response_HMGB1 release from the cell”, and PTPN21 signaling was correlated with “Signal transduction_IGF-1 receptor signaling pathway”.Meanwhile, GO indicated that genes co-expressed with These results were consistent with previous findings. However, there are a few limitations of our research. The data we analyzed in this study were derived from high throughput databases. Therefore, further cellular studies are required to verify our findings and investigate possible mechanisms, molecular interactions, and clinical applications of various PTPN family genes involved in cancer development.As previous research was yet to elaborate the roles of PTPN family genes in LUAD, this study may serve as the first and foremost work that specifically examined roles of PTPN individuals in this disease, prior to giving a more extensive and incisive understanding of potential therapeutic and prognostic values in view of the benefits they offer to LUAD patients.


Apocynum venetum is a semi-shrubby perennial herb that not only prevents saline–alkaline land degradation but also produces leaves for medicinal uses. Although physiological changes during the seed germination of A. venetum in response to salt stress have been studied, the adaptive mechanism to salt conditions is still limited. Here, the physiological and transcriptional changes during seed germination under different NaCl treatments (0–300 mmol/L) were examined. The results showed that the seed germination rate was promoted at low NaCl concentrations (0–50 mmol/L) and inhibited with increased concentrations (100–300 mmol/L); the activity of antioxidant enzymes exhibited a significant increase from 0 (CK) to 150 mmol/L NaCl and a significant decrease from 150 to 300 mmol/L; and the content of osmolytes exhibited a significant increase with increased concentrations, while the protein content peaked at 100 mmol/L NaCl and then significantly decreased. A total of 1967 differentially expressed genes (DEGs) were generated during seed germination at 300 mmol/L NaCl versus (vs.) CK, with 1487 characterized genes  classified into 11 categories, including salt stress (29), stress response (146), primary metabolism (287), cell morphogenesis (156), transcription factor , bio-signaling (173), transport (144), photosynthesis and energy (125), secondary metabolism (58), polynucleotide metabolism (21), and translation (286). The relative expression levels (RELs) of selected genes directly involved in salt stress and seed germination were observed to be consistent with the changes in antioxidant enzyme activities and osmolyte contents. These findings will provide useful references to improve seed germination and reveal the adaptive mechanism of A. venetum to saline–alkaline soils. Apocynum venetum L. (Apocynaceae) is a semi-shrubby perennial herb that is widely distributed in saline–alkaline soils and plays a critical role in preventing the land from degradation [radation ,2. The lradation ,4,5.A. venetum in response to abiotic stresses. Specifically, salt and drought stress significantly inhibited seed germination [In natural habitats, the plants have to face abiotic stresses, such as drought, extreme temperatures, and high salinity . In salimination ,12,13,14mination ; a prolomination ,15; and mination . For themination ,17,18. Mmination .Halimodendron halodendron and Alhagi sparsifolia was significantly affected by the NaCl treatments [Halogeton glomeratus and Cannabis sativa was inhibited by increased salt concentrations [Broyssonetia papyrifera and Pogostemon cablin gradually increased with increased salt concentrations [Arabidopsis under salt stress [Previous studies have found that there are distinct resistances for seed germination among different psammophytes under salt stress. For example, the seed germination of eatments ; the seetrations ,22. Extetrations ,24. For trations ,26. Hundtrations . For exat stress ,29,30. TA. venetum has been found, and the physiological changes have also been primarily conducted [A. venetum to salt stress in this study, we examined the changes in the rate of seed germination, the activity of antioxidant enzymes, the content of osmolytes, and the expression levels of genes in A. venetum at the seed germination stage under different NaCl treatments.To date, a low-salt environment improving the seed germination of onducted ,17,18, wp < 0.05), with an increase at the low NaCl concentration (50 mmol/L), while there was a gradual decrease with increased NaCl concentrations (from 100 to 300 mmol/L) compared with CK.As shown in As shown in As shown in In order to find far more genes involved in salt tolerance, transcriptomic analysis was conducted on a higher salt concentration (300 mmol/L) instead of a lower salt concentration (50 mmol/L) vs. CK. After data filtering, 51.43 and 65.36 million high-quality reads were generated, and 42.12 and 53.26 million unique reads with 0.81 and 0.80 million multiple reads were mapped for CK and NaCl (300 mmol/L), respectively .Coffea arabica, Carica papaya, Coffea eugenioides, Pistacia vera, Actinidia chinensis, Vitis vinifera, Citrus clementine, Theobroma cacao, Olea europaea, and Citrus sinensis , Eukaryotic Orthologous Groups of proteins , NCBI non-redundant protein , and SwissProt  databases . Using tsinensis . Using tsinensis .Among the 1967 DEGs, 1626 genes were UR and 341 genes were DR at 300 mmol/L NaCl vs. CK A,B, baseOf these 1967 DEGs, 1729 genes were identified from the KEGG, KOG, SwissProt, or GO databases A. Among ALDH7B4, ALDH10A8, and ANN2) (At2g40140) to 18.54-fold (IQM4) UR for eight genes and 0.47- (PCC13) and 0.50-fold (WSD1) DR for two genes at the 300 mmol/L NaCl vs. CK  were found to be directly associated with salt stress  . The expl vs. CK . Meanwhil vs. CK .SODCP, FSD2, and SODA), POD , and CAT  (CAT1) to 18.34-fold (SODA) UR for 11 genes and 0.71-fold (poxN1) DR at 300 mmol/L NaCl vs. CK  directly associated with antioxidant enzymes, including SOD (nd PNC1) , were scnd PNC1) . The expl vs. CK . Meanwhil vs. CK .G6pc2, G6PDC, and GAPB), sucrose , fructose , galactose (GOLS1 and GOLS2), trehalose (TPS6 and TPS7), fucose (OFUT19 and OFUT39), and starch , and the 38 genes associated with protein metabolism included PSMD2, RPN1A, RPN10, etc. (ALEU) to 12.13-fold (GOLS1) UR for 22 genes and 0.57-fold (Cys) DR at 300 mmol/L NaCl vs. CK  directly associated with osmolytes, including soluble sugar (41 genes) and protein (38 genes), were screened from the 287 DEGs involved in primary metabolism; specifically, the 41 genes associated with soluble sugar metabolism included glucose  (At5g23350) to 14.48-fold (F16) UR at 300 mmol/L NaCl vs. CK  directly associated with seed germination   were scrnd ROH1) . The expl vs. CK . Meanwhi350 gene .MYB (MYB73 and MYB1R1), WRKY , NAC , BZIP44, TCP14, and UNE10 (JA2L) to 12.13-fold (MYB73) UR at 300 mmol/L NaCl vs. CK  directly associated with stress response and seed germination, including nd UNE10 , were scnd UNE10 . The expl vs. CK . Meanwhil vs. CK .GASA2), IAA , CTK , JA (JOX2 and TIFY6B), ABA , and ETH  response (JOX2) to 13.36-fold (PP2CA) UR for eight genes and 0.39- (ETR2) to 0.98-fold (ERF013) DR for four genes at 300 mmol/L NaCl vs. CK  directly associated with hormone response, including GA  (CAX1) to 12.41-fold (KEA2) UR for nine genes and 0.78-fold (ZIP1) DR at 300 mmol/L NaCl vs. CK  directly associated with ion transport   were scrd CLC-B) . The expl vs. CK . Meanwhil vs. CK .A. venetum in response to salt stress.There are 490 DEGs involved in other biological functions, including photosynthesis and energy , the content of osmolytes , and the expression levels of genes associated with salt-stress response and seed germination.Under both natural and agricultural conditions, plants are frequently exposed to unfavorable environments such as salt stress, extreme temperatures, and drought stress, which restrict plant distribution and reproduction, while plant adaptations that are genetically determined confer stress resistance . Salt stA. venetum decreases sharply under salt stress [Extensive experiments have demonstrated that stress resistance in response to salt stress is involved in the changes in physiological parameters , antioxidant enzyme activities , and osmolyte contents  ,38. For t stress ,12,13,14t stress ,42,43,44t stress ,24,45.A. venetum by adjusting the antioxidant enzyme activities and osmolyte contents. Actually, salt response is not only involved in the changes in phenotypes, antioxidant enzymes, and osmolytes but also in transcriptional alternations [A. venetum at 300 mmol/L NaCl vs. CK, with 11 categories classified, including response to salt, stress response, primary metabolism, cell morphogenesis, TFs, bio-signaling, transport, photosynthesis and energy, secondary metabolism, polynucleotide metabolism, and translation.In this study, we found that a low NaCl concentration (50 mmol/L) could promote the seed germination of rnations ,47. HereALDH7B4 is activated by high salinity, dehydration, and ABA in a tissue-specific manner [ANNs) in Arabidopsis is regulated by exposure to salt, drought, and extreme temperature conditions [RD22 is induced by salt stress and water deficit during the early and middle stages of seed development [CAMBP25 is induced in Arabidopsis seedlings exposed to high salinity, dehydration, and low temperature [PCC13 is abundantly expressed in dried leaves and abscisic acid-treated dried callus and is involved in the response to salt stress [RBG7 plays a role in altering the germination and seedling growth of Arabidopsis under various stresses [IQM4 plays a key role in modulating the responses to salt, ABA, and osmotic stress during seed germination and post-germination growth [NFD4 mediates the transport of Na+ and K+ into the vacuole, influences plant development, and contributes to salt tolerance [WSD1 is involved in the accumulation of wax esters in response to salt, drought, and ABA [At2g40140 plays important roles in modulating the tolerance of Arabidopsis to salt stress [A. venetum in saline–alkaline soils.Specifically, a total of 29 genes were directly associated with salt stress. For example, c manner ; annexinnditions ; and RD2elopment . CAMBP25perature . PCC13 it stress ; RBG7 plstresses ; IQM4 pln growth ; NFD4 meolerance ; WSD1 is and ABA ; and At2t stress . In thisThe primary effect of salt stress is hyperionic and hyperosmotic stresses, as well as, in severe cases, oxidative stress in plants. Oxidative stress is responsible for the generation of ROS, which are deleterious to plants. To minimize the effect of oxidative stress, plants have developed an efficient system of antioxidant enzymes  that can protect plants from damage ,59,60,61TPS) gene plays a critical role in Citrullus lanatus’ response to salt stress [OsGolS1 gene is significantly up-regulated in Oryza sativa ssp. japonica under salt stress [MsTRX in tobacco induced the upregulation of beta-amylase 1 (BAM1) under salt stress [G6PDC is involved in the synthesis of D-ribulose 5-phosphate from D-glucose 6-phosphate [GAPA1 is involved in the synthesis of pyruvate from D-glyceraldehyde 3-phosphate [PGMP is involved in both the breakdown and synthesis of glucose [UGPA is involved in the UDP-glucose metabolic process [INVE participates in cleaving sucrose into glucose and fructose and is associated with the assimilation of nitrogen to control the sucrose to hexose ratio [SUS2 is involved in providing UDP-glucose and fructose for various metabolic pathways [FKFBP is involved in the synthesis and degradation of fructose 2,6-bisphosphate [F16P2 and FBA3 are involved in the fructose 1,6-bisphosphate metabolic process [GOLS1 is involved in the synthesis of raffinose family oligosaccharides and promotes plant stress tolerance [TPS6 is involved in the trehalose biosynthetic process [OFUT39 is involved in the fucose metabolic process [DPE2 is involved in the starch catabolic process and an intermediate on the pathway by which starch is converted to sucrose [AMY3 is involved in stress-induced starch degradation [BAM1 plays a role in the starch degradation and maltose metabolism [GWD3 is involved in starch degradation and mediates the incorporation of phosphate into starch-like phospho-alpha-glucan [RPN1A is involved in the ATP-dependent degradation of ubiquitinated proteins [PCS1 is involved in proteolysis and possesses peptidase activity toward casein [Cys is involved in the protein catabolic process and shows the highest affinity for Pro [DEGP1 is involved in the degradation of damaged proteins [PBE2 is involved in the proteasomal protein catabolic process and shows an ATP-dependent proteolytic activity [SBT2.5 and ALEU are involved in proteolysis and show serine-type peptidase activity [Saline and alkaline ions can reduce the soil water potential, which makes it difficult for plants to absorb water, and then causes osmotic stress . Previout stress ; the OsGt stress ; and thet stress . In thishosphate ; GAPA1 ihosphate ; PGMP is glucose ; UGPA is process ; INVE pase ratio ; SUS2 ispathways ; FKFBP ihosphate ; F16P2 a process ; GOLS1 iolerance ; TPS6 is process ; OFUT39  process ; DPE2 is sucrose ; AMY3 isradation ; BAM1 pltabolism ; and GWDa-glucan . The othproteins ; PCS1 isd casein ,81; Cys  for Pro ; DEGP1 iproteins ,84; PBE2activity ; and SBTactivity ,85. In tF16 is involved in seed trichome differentiation and expressed in the early phase of cotton fiber development [KAI2 is involved in seed germination and seedling development [ROH1 is involved in seed coat development [For the biological functions of the six genes directly associated with seed germination, elopment ; GEM-likelopment ; KAI2 iselopment ; and ROHelopment . Thus, tFvMYB82 gene probably plays an important role in the response to salt and cold stresses in Arabidopsis thaliana by regulating downstream related genes [ThNAC4 gene of Tamarix hispida is involved in salt and osmotic stress tolerance [MYB73 is involved in the salt-stress response [WRKY TFs (WRKY24 and WRKY71) play critical regulatory roles in the plant alkaline metal (sodium) salt-stress response [NAC083 is involved in xylem vessel formation and mediates signaling crosstalk between the salt-stress response and leaf-aging process [JA2L with MYC2 forms a transcription module that regulates wounding-responsive genes [BZIP44 is involved in the positive regulation of seed germination through MAN7 gene activation [TCP14 is involved in the regulation of seed germination and regulates the activation of embryonic growth potential during seed germination [UNE10 negatively regulates seed germination [A. venetum under salt stress.TFs are emerged as key regulators in various signaling networks and play significant roles by improving the growth and development of plants under stress conditions . Previoued genes ; and NAColerance . In thisresponse ; WRKY TFresponse ; NAC083  process ; and JA2ve genes . For seetivation ; TCP14 imination ; and UNEmination . Here, tSmGASA4 was found to be positively regulated by Gibberellin (GA) and significantly enhanced plant resistance to salt, drought, and paclobutrazol (PBZ) stress in Salvia miltiorrhiza [OsIAA24 and OsIAA20 are up-regulated in rice under high salt stress [Tamarix ramosissima under NaCl stress [GASA2 plays a role in the promotion of GA responses during seed germination, flowering, and seed maturation [ARF2A, IAA9, and TET4 act as auxin-responsive genes and regulate vegetative growth, lateral root formation, and flower organ senescence [CISZOG2 may regulate active versus storage forms of cytokinin (CTK) and can have an impact on seed growth [AHK3 acts as a redundant positive regulator of CTK signaling that regulates many developmental processes such as seed germination, cell division, and shoot promotion [JOX2 prevents the over-accumulation of jasmonate (JA) under stress responses [TIFY6B acts as a repressor of JA responses and is involved in the regulation of defense responses [PYL3 is involved in ABA signaling during seed germination and abiotic stress responses [PP2CA acts as a major negative regulator of ABA responses during seed germination and cold acclimation [ETR2 acts as a redundant negative regulator of ethylene (ETH) signaling [ERF013 may be involved in the regulation of gene expression by stress factors and by the components of stress signal transduction pathways [A. venetum.As is known, endogenous hormones play critical roles in plant growth and development. Previous studies have demonstrated that genes related to hormone signaling play critical roles in response to salt stress in other plants. For example, iorrhiza ; Aux/IAAt stress ; genes rl stress . In thisturation ; ARF2A, nescence ,106; CISd growth ; AHK3 acromotion ; JOX2 presponses ; TIFY6B esponses ; PYL3 isesponses ; PP2CA alimation ; ETR2 acignaling ; and ERFpathways . Their d+ is the main toxicity ion due to the similarity in size of the hydrated ionic radii of Na+ and K+, which makes them difficult to discriminate [+/H+ antiporter activity during salt stress [CLC-c is involved in the response to salt stress tolerance and seed germination in Gossypium hirsutum [+ efflux transporters (KEAs) were expressed under abiotic stress  in Cajanus cajan [Zip) gene family participates in plant growth and development and the ability to cope with outside environment stresses, which may potentially regulate seed germination and stress resistance in Miscanthus sinensis [CSC1 acts as an osmosensitive calcium-permeable cation channel and is activated by hyperosmotic shock after NaCl treatment [CLC-C is involved in ion transmembrane transport [RAN1 is involved in copper import into the cell and acts by delivering copper to create functional hormone receptors [KEA2 transports K+ and Cs+ preferentially relative to Na+ or Li+ [OEP16 is involved in ion transport and acts as a voltage-dependent high-conductance channel [PPI1 is involved in proton transport and promotes AHA1 plasma membrane ATPase activity by binding to a site different from the 14-3-3 binding site [NHX2 is involved in the vacuolar ion compartmentalization necessary for cell volume regulation and cytoplasmic Na+ detoxification [CAX1 translocates Ca2+ and other metal ions into vacuoles using the proton gradient formed by H+-ATPase and H+-pyrophosphatase [VHA-a2 is involved in vacuolar nutrient storage and in the tolerance to some toxic ions, as well as catalyzing the translocation of protons across membranes [ZIP1 mediates zinc uptake from the rhizosphere and may also transport copper and cadmium ions [A. venetum under salt stress.Saline and alkaline stress can induce ion toxicity in plants when too many toxic ions enter plant cells, which can harm the plant cytoplasm and organelles; among them, Nariminate . The salt stress . Previouhirsutum ; K+ efflus cajan ; the Zipsinensis . In thisreatment ,121; CLCransport ; RAN1 iseceptors ; KEA2 tr+ or Li+ ; OEP16 i channel ; PPI1 ising site ; NHX2 isfication ; CAX1 trsphatase ; VHA-a2 embranes ; and ZIPium ions . Their dA. venetum is proposed  is generated; subsequently, the genes related to salt-stress responses , soluble sugar metabolism , protein metabolism , TFs , and ion transport  are up-regulated, which promotes the antioxidant enzyme activities  and osmolyte contents  that protect the cells from salt-stress-induced membrane injury; finally, these changes are in favor of building the cell morphogenesis, enhancing seed germination, and conferring the ability to adapt to the saline–alkaline environment.Based on the above studies on the physiological and transcriptional changes, a model of the low-salt-stress-enhanced seed germination of proposed . BrieflyApocynum venetum L. were collected from a resource nursery located at Alxa Left Banner, China  in September 2020. Seeds were air dried at room temperature, stored at 4 °C in air-tight bags in the dark for two months, then cleaned with tap water and successively immersed in 50 °C water for 15 min and sanitized in 70% ethanol for 20 s. After being rinsed with sterile water 5 times, 30 seeds were sown in a Petri dish  containing 10 mL of NaCl solution with 0 (CK), 50, 100, 150, 200, and 300 mmol/L. These doses were selected based on the published literature [Mature seeds of terature ,16,17. EAfter germination in a dark growth chamber at 22 °C for 3 days, the seed germination rate was measured. Three seeds were randomly collected from the 40 independent biological replicates for each NaCl treatment.The activities of 4 antioxidant enzymes including SOD, POD, CAT, and APX in seed embryos were determined using a spectrometer . Briefly, SOD activity was determined based on the ability to inhibit the photochemical reduction of nitro blue tetrazolium chloride: extracts (100 μL) were added to the reaction, and the absorbance reading was taken at 560 nm . POD actThe contents of 4 osmolytes including soluble sugar, protein, MDA, and Pro in seed embryos were determined using a spectrometer . Briefly, the soluble sugar content was determined by the phenolsulfuric acid method: extracts (1000 μL) were added to the reaction and the absorbance reading was taken at 485 nm . ProteinTotal RNA samples of CK and 300 mmol/L NaCl treatments with 3×40 biological replicates were extracted using an RNA kit . The RNA amount was quantified using a NanoDrop ND1000 spectrophotometer , and the quality was determined using an Agilent 2100 Bioanalyzer . One cDNA library was constructed from the total RNA samples of CK and 300 mmol/L NaCl treatments, and the processes of enrichment, fragmentation, reverse transcription, the synthesis of the second-strand cDNA, and the purification of cDNA fragments were conducted as previously described . Reads w2(fold-change)| > 1. The function of DEGs was annotated using BLAST against the databases, including Nr, KEGG, KOG, SwissProt, and GO, with an e-value ≤ 10−5 as a threshold [Raw reads were filtered using a FASTQ system to obtain high-quality clean reads by removing reads containing adapters, more than 10% of unknown nucleotides (N), and more than 50% of low-quality  bases . Clean rhreshold .Actin gene was used as a reference control [−CtΔΔ method  according to the following formula [The primer sequences  were des control . The REL formula . The valp < 0.05 considered statistically significant.SPSS 22.0 software was used for statistical analysis, and one-way analysis of variance (ANOVA) by Duncan’s test was performed for statistical comparisons with A. venetum. The seed germination of A. venetum is promoted at low NaCl concentrations such as 50 mmol/L, and significant changes in antioxidant enzyme activities, osmolyte contents, and genes expression levels in A. venetum play critical roles in regulating seed germination under different salt stresses. These findings indicate that A. venetum plants can adapt to salt stress in saline–alkaline soils by integrating physiological and transcriptional responses. The specific roles of key genes in conferring the ability to salt resistance will require further investigation.This research reveals that salt stress affects the seed germination of


This review describes the existing literature on bioprospecting and exploration of secondary metabolites from Chilean coasts.Chile is in the extreme southwestern part of America, and it has an extreme length, of approximately 4300 km that increases to 8000 km considering the Chilean Antarctic Territory. Despite the large extent of its coastal territory and the diversity of geographic environments and climates associated with Chilean coasts, the research on marine resources in Chile has been rather scarce. From marine organisms found in Chilean coastal waters, algae have been the most studied, since they contain a wide range of interesting secondary metabolites that have some structural traits that make them unique and uncharacteristic. Thus, a wide structural variety of natural products including terpenoids , furanones, and C Marine natural products have served as a rich source of new bioactive agents ,2,3. TheMarine macroalgae have been used in a number of important areas including the food industry , agricul1)   (Petrosia contignata ..39) and Tetrapygus niger embryos, and 43 was identified as the most active compound. However, its activity could be considered mild as compared to that shown by stypoldione, another active marine compound with ED50 = 1.1 µg/mL. Nevertheless, this test seems to be a reasonable prescreen to determine which substances merit further evaluation for antineoplastic properties [All of these compounds were assayed for inhibition of cytokinesis in the sea urchin operties .40), and two semisynthetic derivatives, 45 and 46 , andDictyota has been presented as a powerful resource for diterpenic compounds with novel chemical structures. Cyclic diterpenes are produced by many members of this genus, just like typical diterpenes with a 6-methyl-5-hepten-2-yl side chain A. Three types of main carbon skeletons have been reported: dolabellanes (including dolastanes), xenicanes, and extended sesquiterpenes [Diterpenoids are found in higher plants, insects, fungi, and marine organisms. Several of these compounds present antimicrobial, antitumor, cytotoxic, anti-inflammatory, antifungal, molluscicide, antifeedant, and antifouling activities . With moterpenes .Dictyota crenulata, collected at Vaihú, Easter Island, allowed chromatographic isolation of five diterpenes, 53–57 and 58  was assayed as well [Compounds  as well .59 . These diterpenes are frequently isolated in the ta genus .60 and 61  [Finally, two new plastoquinone diterpenes of mixed biogenesis, 2 and 63 , were isro Prat) .Meroterpenoids are prenylated aromatic compounds of mixed biogenesis combining acyclic, monocyclic, and bicyclic terpenes with aromatic or substituted aromatic groups possessing different degrees of oxidation. Many meroterpenoids from seaweeds have interesting biological activities such as antibacterial, antiviral, and antifeeding properties . Plants 64–80 (65) structure was fully revised, and a meroditerpenoid containing an unusual two fused six-membered rings forced into the twist-boat conformation was demonstrated [Seventeen meroterpenoids, 64–80 , were isnstrated .67 and 68   , it was ulations .76) (Stypopodium species. The absolute configurations of (−)-taondiol diacetate (74) and (+)-epitaondiol diacetate (71) isolated previously were determined using vibrational circular dichroism (VCD). For verification, their relative stereochemistry was determined by X-ray diffraction. Additionally, the crystal stereo structure of meroditerpenoid (79) (Taondiol (76)  has beenoid (79)  was repo65, 66 and 71  for agents active against HIV (killing of T lymphocytes by HIV) [66  . The inhHIV) [66  were stu77), isoepitaondiol (64), and epitaondiol (65) exhibited gastroprotective activity in mice [The meroditerpenoids stypodiol ( in mice .65 and sargaol (78) were tested on HCl/ethanol-induced gastric lesions in mice and compared with lansoprazole. Both 65 and 78 showed gastroprotective activity with ED50 values between 35 mg/kg and 40 mg/kg. The results suggest that 65 and 78 protect the gastric mucosa in the HCl/EtOH model in mice [Compound  in mice .43) (79) and epitaondiol (65)  , stypotriol (65)  were asstrophils .65, 68, 71, 77, 79 and 80  at 3 m depth, led to isolation and identification of fifteen metabolites belonging to a single biosynthetic class [81–86), magellenediol (87), magellenone (88) and ptiloninol (89) . It is worth noting that ptilonines present an unusual halogenation substitution pattern, which may confer evolutionary advantages to P. magellanica, for which a biogenetic origin is proposed [Studies carried out on the red alga ic class ,98. Ptilnol (89)  are nove4 and 89  were tesproposed .94) [91–93) and pyranosylmagellanicus D–E (95 and 96) (P. magellanica [90 (Previously reported γ-pyrone (94)  and five and 96) , were obellanica ,98. Thesnica [90 .91), was determined by treatment of 91 with (R)- and (S)-α-methoxy-α-phenylacetic acids (MPA). Compounds 91–93 were tested for their antimicrobial activity, but no activity was found [The absolute configuration of the known pyranosylmagellanicus A (as found .97) -13-epipinnatifidenyne (97) , a new Canalysis .®, HDMF, also 4-hydroxy-2,5-dimethyl-2,3-dihydrofuran-3-one) was identified for the first time in 1960, as a product of the Maillard reaction or nonenzymatic browning [2-Furanone is a heterocyclic organic compound. It is also known as γ-crotonolactone (GCL), as it is formally the lactone derived from γ-hydroxyisocrotonic acid. 4-Hydroxy-2,5-dimethyl-3(2H)-furanone (Furaneolbrowning . Previoubrowning .98) -furanone has been known since 1929, but it has not been reported previously from any natural sources [99) (99 was identified as a trimer of 2-methyl-3(2H)-furanone [Chilenone A (98)  was obta sources . Chilenoces [99)  was obtafuranone .100 , 31 are monoterpenes (8–38), 14 sesquiterpenes (39–52), 11 diterpenes (53–63), 17 meroterpenoids (64–80), 16 acetogenins (81–89 and 91–97), and three furanones (98–100). Among the biological activities studied in these compounds are the following: insecticidal activity of 27 and 28 against Aphis fabae, 53 and 55 against tomato moth (Tuta absolute), 65 against Spodoptera frugiperda; cytotoxicity of 31 against colon and cervical adenocarcinoma cells; inhibition of cytokinesis by 43 against Tetrapygus niger; toxicity against Schizaphis graminum; gastroprotective activity of 65 and 78 in mice; topical anti-inflammatory activity of 65 related to inhibition of leukocyte accumulation and human recombinant synovial phospholipase A2 activity; and inhibition of degranulation response by 40. Compounds 65, 68, 71, 77, 79 and 80 were tested for their cell proliferation inhibitory activity in five cell lines: Caco-2, SH-SY5Y, RBL-2H3, RAW.267 and V79. Antimicrobial activity by 77, 79 and 80, and antibacterial activity by 89 against K. pneumoniae were determined.A first review of investigations carried out with marine algae from the Chilean coasts was published in 1989. Herein, we have updated the information by compiling all works, published up to mid-2019, on natural products isolated from algae collected along the coasts of Chile. The main species of algae that have been studied are Plocamium is striking. The characteristic metabolites of this genus have been isolated from algae of the genera Microcladia, Shotera, Pantoneura and Ceramium. All of them belong to different families. So far, no explanation has been given for this phenomenon. On the other hand, it has been proposed that in P. violaceum, there would be two chemotypes depending on whether the monoterpenes are cyclic or lineal. An analogous situation can be described for P. cartilagineum; however, the description of oxygenated monoterpenes suggests that a possible chemotaxonomy of this species could become even more confusing.The situation observed in the genus Thus, from a chemical point of view, Chilean algae are characterized by the unique structures of some of their metabolites. However, considering the great variety of Chilean alga species, the number of works that have been published is relatively scarce. The extensive Chilean coasts are bathed by the cold Humboldt current, resulting in very cold waters in the extreme south and much warmer waters in the extreme north. Easter Island is a special case because it is in the middle of the ocean, far from the influence of this current and from any other kind of external influence. For these reasons, the marine biodiversity of the Chilean coast can be considered as an important source of new bioactive marine natural products that could be the basis for the development of new drugs but that have been poorly studied and exploited to date.


Adult-type diffuse gliomas are malignant primary brain tumors characterized by very poor prognosis. Dendritic cells (DCs) are key in priming antitumor effector functions in cancer, but their role in gliomas remains poorly understood.In this study, we characterized tumor-infiltrating DCs (TIDCs) in adult patients with newly diagnosed diffuse gliomas by using multi-parametric flow cytometry and single-cell RNA sequencing.We demonstrated that different subsets of DCs are present in the glioma microenvironment, whereas they are absent in cancer-free brain parenchyma. The largest cluster of TIDCs was characterized by a transcriptomic profile suggestive of severe functional impairment. Patients undergoing perioperative corticosteroid treatment showed a significant reduction of conventional DC1s, the DC subset with key functions in antitumor immunity. They also showed phenotypic and transcriptional evidence of a more severe functional impairment of TIDCs.Overall, the results of this study indicate that functionally impaired DCs are recruited in the glioma microenvironment. They are severely affected by dexamethasone administration, suggesting that the detrimental effects of corticosteroids on DCs may represent one of the mechanisms contributing to the already reported negative prognostic impact of steroids on glioma patient survival. Gliomas represent 75% of malignant primary brain tumors in adults, and still remain among the most difficult cancers to treat . Their sin vivo and partial benefit on overall and progression-free-survival rates  that play a central role in the activation and regulation of all immune responses . DC-lineBeyond their belonging to one of these subsets, the behaviour of DCs depends also on their state of activation that is in turn affected by stimuli provided by the tissue microenvironment where DCs reside or are recruited. Upon exposure to inflammatory stimulation, DCs up-regulate the expression of MHC and costimulatory molecules, secrete pro-inflammatory cytokines, and present antigens to T cells in fully stimulatory conditions. On the other hand, DCs exposed to immunosuppressive environment express low levels of MHC and costimulatory molecules, up-regulate the expression of inhibitory molecules, secrete immunosuppressive cytokines, and present antigens to T cells in tolerogenic conditions . AccordiThe identification of DC subsets in the TME, together with the characterization of their activatory/tolerogenic profile, has been hampered so far by the low number of DCs in the TME and the lack of DC-specific markers. The recent implementation of high-dimensional single-cell technologies is making possible to define DC features at an unprecedented definition, both at the phenotypic and transcriptomic levels. Accordingly, DCs have started to be deeply characterized in the TME of different types of tumors, providing evidence that tumor immune evasion involves crippling normal DC functions, and that DC heterogeneity and states are conserved across various solid human cancers , 22. In The study was conducted on 27 newly diagnosed, non-relapsing adult patients with diffuse glioma undergoing surgical resection at the unit of Neuro-Oncology of Humanitas Research Hospital, Rozzano, Milan, Italy. Clinical patient information is provided in Peripheral blood samples were collected from patients and controls in K2 EDTA BD vacutainer tubes  and stained with an 18-color DC-dedicated flow cytometry panel of monoclonal antibodies (mAbs) as previously reported . 500 μL 2+ and Mg2+, Lonza, with the addition of 2% FBS), and centrifuged at 290 rcf for 7 min. The samples were then incubated with FACS buffer and Myelin Removal Beads II (Milteny Biotec) and passed through LS Columns (Milteny Biotec) according to manufacturer’s instructions. The samples were stained with the same DC-dedicated flow cytometry panel used for peripheral blood samples.Brain tissue samples obtained during surgery were collected, stored at 4°C in supplemented Dulbecco’s Modified Eagle Medium (DMEM) high glucose (Lonza) added with 1% Penicillin/Streptomycin and 1% L-Glutamine and digested within 2 hours from excision with type IV Collagenase (1.6 mg/mL) (Merck KGaA) and type I DNase (0.4 mg/mL) (Merck KGaA) in Roswell Park Memorial Institute (RPMI) 1640 medium (Euroclone SpA) at 37°C for 1 hour. Homogenates were then smashed on a 70 μm filter (BD Biosciences), washed with RPMI with the addition of 2% fetal bovine serum (FBS) (Lonza), and collected in 50 mL collection tubes. Samples were then centrifuged at 290 rcf for 7 min, and the pelleted cells were incubated for 2 min with 1 mL of ACK 1X to lyse erythrocytes. Samples were then washed with FACS buffer  or CD45br (tissue samples) mononuclear cells, DC-lineage DCs were identified within the gate of lineage −/CD14−/CD16−/HLA-DR+ cells. Gated on these cells, pDCs were identified as CD123+/CD11c− cells; cDCs were identified as CD11c+/CD123− cells, and further divided into cDC1s and cDC2s based on the expression of CD141 and CD1c, respectively. Inflammatory DCs were identified as lin−/HLA-DR+/CD11c+ cells that could be positive or negative for CD14 and CD16 expression. They included slanDCs that expressed M-DC8, and moDCs that expressed CD1a. The activation state of each DC subset was examined by assessing the expression of the activatory molecules CD40, CD80 and CD86, and the inhibitory molecules PD-L1, ILT2 and TIM-3. The compensated data were further imported into FlowJo software version 10.7.1 and visualised with a uniform manifold approximation and projection (UMAP). For the UMAP analysis, 2 different concatenated files were created, containing the same number of live CD45+/lin-/HLA-DR+ cells derived respectively from whole blood of untreated patients (n=12) and whole blood of dex-treated patients (n=11). A unique computational barcode was assigned to each concatenated file. These files were then concatenated in a single file for further visualization in UMAP dot plots , based on the expression of the following markers: CD45, CD14, CD16, HLA-DR, CD11c, CD123, CD141, CD1c, M-DC8, CD1a, CD40, CD80, CD86, PD-L1, ILT2, TIM-3. The same analysis was applied also to the cells derived from the tumor, where 2 different concatenated files were created, containing the same number of live CD45br/lin-/HLA-DR+ cells derived respectively from tumour tissue of untreated patients (n=5) and tumor tissue of dex-treated patients (n=3).All data were acquired on a FACSymphony™ A5 flow cytometer (BD Biosciences). Flow Cytometry Standard (FCS) 3.0 files were imported into FlowJo software version 9.9.6 (FlowJo LLC), and data were compensated by using single-stained antibody-capture beads  as previously described –25. Theshttps://zenodo.org/record/6046299#.YgZ6bpbSKN4) and analyzed with R (v3.5.1) toolkit Seurat (v3.0.2). For each sample, Seurat objects were created from feature-barcode matrices. Cells containing > 200 genes and ≤ 10% mitochondrial genes were kept for downstream analysis. Gene expression matrices were then log-normalized with a scale factor of 10,000.Feature-barcode matrices generated by Savino et al. were down-loaded from Zenodo Repository, where the original data have been deposited . Cluster annotation was performed in silico using SingleR. The cell cluster enriched in DCs (cluster 19) was manually identified based on literature data obtained with scRNA-seq analyses of sorted DC subsets (adj<0.05) of cluster 19 were then identified by using the ‘FindAllMarkers’ function in Seurat, with the parameter ‘test.use=wilcox’ used by default. The aggregated expression scores of these DEGs were calculated on single-cell base using the ‘AddModuleScore’ function in Seurat. The distribution of DC subsets across different clusters at resolution 0.5 was investigated by analyzing the expression of genes characteristics of classical DC subsets and other subsets recently described on the basis of their transcriptomic profile, including preDCs, migDCs, cDC2A and cDC2B  and Canonical Pathways (CPs) tools on IPA software. Only statistically significant DFs and CPs characterized by p<0.05 and |z-score|>1.5 were considered. Each gene identifier was mapped to its corresponding gene object in the Ingenuity Pathway Knowledge Base (IPKB).In order to investigate whether the cluster distribution of TIDCs may reflect DC functional state, we analyzed cell clusters at resolution 0.3 using IPA software program (Qiagen), which analyzes gene expression patterns using a built-in scientific literature-based database. DEGs that were characterized by pStatistical analysis of flow cytometric results was performed using GraphPad Prism software, version 9.0.0. The normal distribution of data was tested by using Shapiro-Wilk’s test. The t-test was used for comparisons between samples. All statistical analyses assumed a two-sided significance level of 0.05.We first analysed PBDCs by using a high-dimensional flow cytometry panel that allows the identification of five distinct DC subsets, namely pDCs, cDC1s, cDC2s, slanDCs, and moDCs , 23. Ourvs untreated patients). The frequency of all circulating DC subsets, including pDCs, cDC1s, cDC2s and slanDCs, was significantly lower in dex-treated compared with untreated patients  was labelled as WHO grade 3. In order to investigate whether the reduction of PBDCs was associated with the histopathological diagnosis of gliomas, we analysed PBDC subsets in untreated patients (to avoid the confounding effect of dexamethasone) further stratified according to their histopathology and observed that, among patients with IDH-wildtype gliomas, the frequency of circulating pDCs, cDC1s and cDC2s was significantly lower in patients with a histopathological diagnosis of glioblastoma compared with those with anaplastic astrocytoma .br/lin-/HLA-DR+ cells obtained from down-sampled and concatenated files of all tissue samples . The Seurat integration procedure was used to remove batch effects. Based on their transcriptomes, unsupervised graph-based clustering partitioned 36,237 cells into 28 distinct clusters. Clusters 25, 26 and 27 were filtered-out because of their small size (less than 20 cells) and excluded from the analysis. We identified cluster 19 as the putative cluster of DCs based on previously reported DC transcriptomic signatures (www.proteinatlas.org), we verified that all the 50 DEGs composing the signature were expressed by human DCs and, in particular, 22 of them were enriched in myeloid and/or plasmacytoid DCs , whereas these cells were a minority (11%) in cluster 0. Among 967 DEGs between the two clusters, 576 were down-regulated and 391 were up-regulated. By further setting a threshold on |log2FC|>0.58 (corresponding to a 1.5-fold change), we selected 531 down-regulated and 362 up-regulated DEGs in cluster 2 compared with cluster 0 . The patients/participants provided their written informed consent to participate in this study.Data curation and Resources by CC, SF, AC, ST, MS, PP, LB, MN, FP, PK, CP, RB, BS, ML, SDB and DM. Conceptualization and Methodology by CC, SF, AC, ST, MS, PP, LB, MN, FP, PK, CP, RB, BS, ML, SDB and DM. Investigation by CC, SF, AC, ST, PK, CP, BS and Formal Analysis by CC, SF, AC, ST, SDB, PK, JM, FC. Writing – Original Draft by CC, SDB, DM. Writing – Review and Editing by CC, SF, AC, ST, MS, PP, LB, MN, FP, PK, CP, SDB, JM, FC, RB, BS, ML, SDB, DM. Funding Acquisition by ML, DM, SDB. Supervision by MS, LB, ML, SDB, DM. All authors contributed to the article and approved the submitted version.


Osteoarthritis (OA) is a complicated disease with both hereditary and environmental causes. Despite an increase in reports of possible OA risk loci, it has become clear that genetics is not the sole cause of osteoarthritis. Epigenetics, which can be triggered by environmental influences and result in transcriptional alterations, may have a role in OA pathogenesis. The majority of recent research on the epigenetics of OA has been focused on DNA methylation, histone modification, and non-coding RNAs. However, this study will explore epigenetic regulation in OA at the present stage. How genetics, environmental variables, and epigenetics interact will be researched, shedding light for future studies. Their possible interaction and control processes open up new avenues for the development of innovative osteoarthritis treatment and diagnostic techniques. Osteoarthritis (OA) is a widespread joint disease that affects around 15% of the world’s population . OA was Epigenetics is an essential gene-environment interaction process. Epigenetic modification, unlike genomic modifications, is more versatile and reversible. Regulations of epigenetics vary by cell type and gene. Epigenetic phenomena including DNA methylation, genomic imprinting, maternal effects, post-translational modifications of histones, RNA (controlled by non-coding regulatory RNAs), and epigenetic chromatin remodeling  have been well studied. . With th. CpG dinucleotides are distributed unevenly throughout the human genome, with the bulk of CpG islands found in the promoter and exon regions of genes 300 to 3000bp in length. As a result, the methylation level of CpGs can be dynamically controlled by DNMTs and TETs to regulate transcription (The most fully studied methylation is 5-methylcytosine (5 mC) . CpG dincription  (Rice etcription .ADAMTS 4) and matrix metalloproteinase (MMPs) 2 (gelatinase A), MMP3 (stromelysin 1), MMP9 (gelatinase B), and MMP13 (collagenase 3)  . DespiteCOL2A1, COL9A1, COL10A1, ACAN, MMP2, MMP3, MMP9, MMP13, ADAMTS-4  . Eilis H. et al. investigated DNA methylation at 850,000 locations across the genome using a MethylationEPIC BeadChip (850K chip) in samples from the Understanding Society UK Household Longitudinal Study (UKHLS)  . They diCOL11A2)  (ReynardCOL11A2) . Further defects . These f defects .DNMT3b , the de novo methyltransferase, can result in OA-like phenotypes in knee joints; whereas DNMT3b overexpression in articular chondrocytes can delay OA development after meniscus ligament injury  promoter region. According to a recent study, patients after bariatric surgery have lower serum leptin levels and a different methylation profile in the lep and lepr gene promoter regions. These findings back up previous studies in rat models, which indicated that a high-energy diet could alter lep methylation patterns  increased MMP13 activity in patient chondrocytes in vitro. MMP-13 was elevated after leptin’s epigenetic reactivation, and small interfering RNA against lep inhibited it directly  are histone and protein deacetylases dependent on nicotinamide dinucleotide (NAD+). SIRTs and SIRT-dependent epigenetic control have been widely reported to play vital roles in DNA repair, inflammation, and aging-related illnesses (SIRT6-deficient mice from the mesenchyme (SIRT1), like SOX9, is essential for maintaining cartilage homeostasis  and Istodax (Romidepsin) have been approved for the treatment of cutaneous T-cell lymphoma. Histone modification medicines for osteoarthritis, on the other hand, have not yet been developed.Studies concentrating on deacetylase/acetylases still have significant limitations because nt genes . HDAC inTypically, histone methylation is more stable than histone acetylation. Histone methylation is performed by histone methylation transferase (HMT) on lysine and arginine residues. Common locations of methylation include Lys 4, 9, 27, 36, 79 and Arg 2, 17, 26 on H3, Lys 20 and Arg 3 on H4, and Lys 4, 9, 27, 36, 79 and Arg 2, 17, 26 on H3 and H4. Studies have shown that arginine methylation on histone is a relatively dynamic marker. Arginine methylation is associated with gene activation, whereas arginine methylation loss in H3 and H4 is linked to gene silence . In contDot1l polymorphism was found to be associated with hip joint cartilage thickness and OA risk in research involving 6,532 people , HEK27Me3 demethylase, was found at increased level in cartilage from the tibial plateau of the OA knee. Its inhibition in vitro by GSK4, significantly increases the expression of OA-related genes MMP13 and PTGS2.Similar to histone acetylation, changes in histone methylation are frequently related to altered gene expression and signaling pathways in chondrocytes. For instance, H3k79 methylation was reduced in OA and RA patients , while H2 people . Accordiertrophy . EZH2 is pathway . Overexpreakdown . In addiMature miRNAs bind to complementary messenger RNA (mRNA) sequences of target genes via RNA-induced silencing complexes (RISCs) or block gene expression directly. Interactions between miRNAs and highly complementary targets lead to mRNA degradation, while incomplete interactions between miRNAs and target transcripts usually lead to translation suppression. In 2008, Dimitrios et al. found that in the chondrocytes of arthroplasty patients, miR-483, miR-22, miR-377, miR-103, miR-16, miR-223, miR-2b, miR-23b, and miR509 were elevated, and miR-29a, miR-140, miR-25, miR-337, miR-210, miR-26a, and miR-373 were decreased. The genetic signatures of these 17 miRNAs clearly distinguish OA chondrocytes from normal chondrocytes, and 17 miRNA-protein pairs that may be involved in the progression of osteoarthritis were revealed by matching microRNAs and proteomics. In this study, miR-22 and miR-103 expression were favorably connected with BMI, but miR-25, and miR-337, while miR-29a expression was inversely correlated with BMI . Since tUnlike microRNAs that rely mostly on RNA sequence complementary pairing to suppress target genes, lncRNAs operate in a significantly more sophisticated manner. lncRNAs have a unique role in a variety of gene expression regulation mechanisms, including epigenetic regulation, transcriptional regulation, and post-transcriptional regulation .Current research has revealed the way of axial regulation of the highly expressed lncRNA in OA tissue to the appropriate miRNA, which subsequently influences the OA target genes. HOTTIP, for instance, was engaged in the proliferation and death of OA chondrocytes via the miR-663a/FRK axis . Other lDNMT3b facilitated the downregulation of miR-29 by increasing its promoter methylation. This, in turn, led to the overexpression of PTHLH, a process strikingly similar to that observed in tumor disorders  proteins, and other variables traditionally associated with OA are all sensitive to epigenetic control to varying degrees. The combination of these genetic and epigenetic variables provides more insight into the pathophysiology of osteoarthritis. Epigenetic control can link a range of genetic and environmental factors, which could provide a holistic understanding of the etiology of osteoarthritis (OA) and guide treatment.At present, the epigenetics of osteoarthritis focuses primarily on chondrocytes, with only two published works addressing the epigenetic manifestations of subchondral bone . Zhang YFor future approaches to epigenetics in the realm of osteoarthritis, it would be prudent to investigate refined single-cell research. In cartilage tissue isolated from OA samples with variable degrees of injury and degeneration, there exist chondrocytes, macrophages, fibroblasts, and numerous other types of tissue. This heterogeneity can compromise the accuracy of current knowledge of gene function and epigenetic regulation. In the case of osteoarthritic cartilage, single-cell investigations provide more precise subtyping of diseased tissues. Embryonic cell research has demonstrated the validity of single-cell methylation investigations . Using E


Ulva rigida, Ulva sp. and Codium tomentosum within the Chlorophyta phylum. It was reported that seaweed derived lipids suppress inducible nitric oxide synthase and cyclooxygenase-2 expression and reduce nuclear factor κB p100 and myeloid differentiation primary response 88 protein levels leading to the downregulation of the production of several pro-inflammatory cytokines and nitric oxide. Further investigations are required to unravel the complex mechanisms underlying their preventive action against chronic inflammation and their potential use as a new functional food ingredient and/or health supplement.Chronic, low-grade inflammation is linked to the development of non-communicable diseases, including cancer, cardiovascular disease, obesity, insulin resistance, diabetes, and others which together contribute to more than 50% of deaths globally. Modulation of inflammatory responses may be a promising strategy, and n-3 long chain polyunsaturated fatty acids (n-3 LC-PUFA) may offer a new therapeutic option in inflammatory conditions. Seaweeds are characterised by high nutritional quality and are a good source of many bioactive compounds, including n-3 LC-PUFA. This review addresses the potential anti-inflammatory properties of seaweed derived lipids, and their immunomodulating mechanisms in order to identify the possible applications of seaweed as an anti-inflammatory functional food ingredient or dietary supplement. A few studies have evaluated the anti-inflammatory activity of seaweed lipids using crude lipid extracts, lipid fractions and isolated complex lipids from several seaweeds belonging to the Ochrophyta and Rhodophyta phyla, with only three  Inflammation is the natural protective response of the immune system of living organisms to infection and other pathogenic factors. It involves a cascade of cellular and microvascular reactions to eliminate pathogens and stimulate tissue repair processes resulting in the restoration of homeostasis at infected and/or injured sites ,2,3. TheNormally, inflammation is beneficial to the host and the inflammatory response is self-limiting resolving rapidly when the triggering insult has been eliminated, the infection is cleared, and damaged tissue is repaired. However, if the inflammatory response becomes dysregulated the activated immune system can cause irreparable damage to the host tissues and organs resulting in adverse health outcomes . Many liThe application of drugs targeting inflammatory pathways appears to be a promising strategy in type 2 diabetes and CVD treatment ,19. HoweTaking into account anti-inflammatory effects, n-3 LC-PUFAs may offer an important therapeutic option in inflammatory diseases. EPA and DHA can be derived from alpha-linolenic acid (ALA) through multiple enzymatic elongation and desaturation reactions catalysed by delta-5 and delta-6 desaturases. However, the conversion rate of ALA to DHA in humans is very low and depends on a number of external and internal factors ,32. TherThe global supply of n-3 LC-PUFAs from all known traditional sources, including capture fisheries and aquaculture, is insufficient to meet human nutritional requirements . Thus, aThis review addresses the potential anti-inflammatory properties of crude lipid extracts, lipid fractions, and isolated lipids from seaweeds and their immunomodulating mechanisms in order to identify the possible application of seaweed as an anti-inflammatory functional food ingredient or dietary supplement. In addition, this review provides a comprehensive overview of fatty acids composition, including the n-6:n-3 ratio, of different seaweed species to assess their usefulness as a dietary source of polyunsaturated fatty acids.This review is based on the literature search performed in the scientific databases of PubMed, Web of Science, Scopus and Science Direct, using the combination of the following keywords: (1) macroalgae or seaweed or algae and (2) lipids or lipid extracts or complex lipids or fatty acids and (3) anti-inflammatory or immunomodulatory. Only original research articles using lipid extracts, lipid fractions and/or isolated lipids of seaweeds were included in this review. In total 13 publications were included .Ulva, Codium, Chaetomorpha, and Cladophora. There are around 1800 species of brown seaweeds, with two main orders Fucales and Laminariales, and less than 1% are found in freshwater [Seaweeds are marine photosynthetic macroalgae that grow in various types of water and can be found in any climatic zone. They can be attached to rocks, pebbles, and other substrata or freely floating primitive plants that lack true root, stem, and leaves . Seaweedeshwater ,51.Porphyra/Pyropia sp.), wakame (Undaria sp.), kombu (Saccharina/Laminaria sp.), and dulse  [Seaweeds have been harvested and used as food since ancient times in Asian countries, such as Korea, China, and especially Japan . There apalmata) ,54,55. Ipalmata) . In Westpalmata) ,40,41. Ipalmata) ,56. Edibpalmata) ,42.Seaweeds are characterized by high nutritional quality and are a good dietary source of fat- and water-soluble vitamins and essential minerals , with some seaweeds containing 10–100 more minerals and vitamins than terrestrial plants or animal food products ,42,57,58They are also a good source of proteins , essential amino acids and fibre (36–60% of dry seaweed mass). Lipid content of seaweeds is low, but rich in EPA and DHA ,40,46. TLipid content in macroalgae is considered low and ranges from 0.5 to 8.0% of dry weight ,68, withGracilaria changii (48.36 ± 6.76)%. Both n-6 and n-3 PUFAs are essential for human health and the recommended ratio between n-6:n-3 is below 1:1–4:1 to reduce the risk of cardiovascular, inflammatory and neurological conditions [Despite the low-fat content, seaweeds are recognised as a rich source of biologically active lipids with a high proportion of unsaturated fatty acids, including n-3 LC-PUFA. In seaweeds, fatty acids are mainly found as polar lipids, including GL, PL, and betaine lipids . The fatnditions . The avenditions .In addition, seaweeds are also the source of other bioactive lipids including squalene and phytosterols such as brassicasterol, chondrillasterol, fucosterol and isofucosterol ,76 with Ulva rigida, Ulva sp. and Codium tomentosum within the Chlorophyta phylum [in chemico [The anti-inflammatory activity of seaweed derived lipids has been studied using crude lipid extracts, lipid fractions and isolated complex lipids from several seaweeds belonging to the Ochrophyta and Rhodophyta phyla ,112,113,a phylum ,90  and U. rigida (87.9 ± 0.1%), followed by P. dioica (83.6 ± 8.1%) and C. tomentosum (82.3 ± 2.2%), Da Costa et al. [G. turturu had a slightly lower potential to inhibit COX-2 activity reaching 50% inhibitory effect at a concentration of 33 µg/mL. A significantly lower inhibitory effect was observed for lipid extracts of F. vesiculosus with 34.6 ± 7.1% of COX-2 inhibition, and G. gracilis showed no inhibition potential of COX-2 activity. It should be noted that the lipid extracts of G. gracilis and F. vesiculosus were characterised by a higher content of AA and n-6, and lower levels of n-3 fatty acids than other studied seaweeds, which may explain their lower anti-inflammatory potential [COX-2 activity and NO levels were the most commonly assessed parameters in the studies evaluating anti-inflammatory properties of seaweed derived lipids. COX-2 is one of the key mediators of inflammatory responses that converts arachidonic acid to prostaglandins and is activated by extracellular and intracellular stimuli, including LPS, cytokines, growth factors, and other pro-inflammatory molecules . COX-2 ua et al.  reportedotential . Previouotential .Sargassum ilicifolium obtained from Ujung Genteng Beach–Sukabumi (UGB) and Awur Bay–Jepara (AB) regions and Gracilaria sp. were shown to significantly suppress NO production of LPS induced RAW 246.7 cells [Lobophora sp. displayed a strong NO inhibitory effect, with the non-polar lipid fraction showing the strongest activity (IC50 of 52.10  ±  4.43), followed by total lipid and polar lipid with IC50 of 61.09  ±  6.06 and 66.21  ±  6.24 µg/mL, respectively [Palmaria palmata and EPA, AA and galactolipids from Chondrus crispus possessed strong and dose-dependent NO inhibitory activity through down-regulation of inducible nitric oxide synthase (iNOS). However, it was observed that galactolipids have stronger NO inhibitory activity as compared to free polyunsaturated fatty acids, including EPA and AA, suggesting that the entire polar lipid structure may be essential for potent anti-inflammatory action [Fucus spiralis inhibit NO release by LPS stimulated RAW 264.7 macrophages. However, the monoacylglycerol, composed of a glycerol moiety linked to oleic acid, demonstrated slightly lower capacity to inhibit NO production by macrophages than the 1:1 mixture of the MGDGs, including a glycerol moiety linked to a galactose unit and eicosapentaenoic acid combined with octadecatetraenoic acid or linolenic acid . The NO inhibitory activity was also observed for several other seaweeds including Ecklonia radiata, Hormosira banksia, Myriogloea sciurus, Phyllospora comosa, Solieria robusta, and Ulva sp., with the NO inhibition being greatest in the nonpolar, lipid-rich DCM extracts , followed by the intermediate polarity ethyl acetate extracts , with the lowest activity observed in the polar butanol extracts [Several in vitro studies assessed NO production in LPS stimulated murine macrophage RAW 264.7. NO is a signaling molecule that plays a crucial role in inflammation and host defense mechanisms. It may act as anti-inflammatory agent under normal physiological conditions, but NO is a potent proinflammatory mediator in chronic inflammation ,119,120..7 cells ,111. Simectively . Moreoveectively ,110 repoy action . On the y action . Similary action  who obseextracts .Cytokines are potent signaling molecules that regulate immune responses and are secreted by the cells of innate and adaptive immunity in response to various stimuli . TherefoP. palmata lipid extract reduced the secretion of IL-8 and IL-6. However, interestingly, C. crispus lipid extract significantly increased production of TNF-α. It should be noted that C. crispus is characterised by high content of ARA, which is recognised as a pro-inflammatory fatty acid. In addition, lipid extracts of P. palmata, C. crispus, and P. dioica promoted the downregulation of mRNA of Toll-like receptor 1 (TLR1) and Toll-like receptor 8 (TLR8), and TNF receptor associated factor 5 (TRAF5). In addition, P. dioica extract downregulated nine other pro-inflammatory genes including TLR4 and signal transducer and activator of transcription 3 (STAT3). However, lipid extracts of P. palmata and P. dioica upregulated NOS2 and PTGER1 gene expression. Another study reported that the lipid extract of Macrocystis pyrifera attenuated lipoteichoic acid (LTA) and LPS induced inflammatory response in THP-1 monocytes through suppression of MCP-1, IL-1β and IL-8, and MCP-1 secretion, respectively [M. pyrifera lipids reduced mRNA expression of MCP-1, IL-1β and IL-8 and myeloid differentiation primary response 88 (MYD88) and NFκB2/p100 protein levels and the phosphorylation of RelA/p65 in LTA-activated monocytes, which indicates the extract inhibits both canonical and non-canonical NFκB pathways [The use of crude lipid extracts from seaweeds promoted down regulation of cytokines production in LPS stimulated THP-1 cells. Robertson et al.  observedectively . Moreovepathways .Ishige okamurae demonstrated anti-inflammatory activity by reduction of phorbol 12-myristate 13-acetate (PMA) induced oedema and erythema in a mouse model [Undaria pinnatifida in a BALB/c mice mode. SA and EPA significantly decreased PMA induced mouse ear inflammation by reducing oedema, erythema, and blood flow. Similar effects were observed for U. pinnatifida derived AA at low concentrations, but AA doses of more than 243 µg per ear demonstrated pro-inflammatory activity.The anti-inflammatory potential of seaweed lipids has also been evaluated in in vivo models (BALB/c mice). The 7-methoxy-9-methylhexadeca-4,8-dienoic acid (MMHDA) isolated from the brown seaweed se model . The obsse model , which ise model  Khan et se model  tested aPalmaria palmata, Chondrus crispus, and Porphyra doicia from Rhodophyta phylum. It is worth noting that red seaweeds are charcaterised by the highest content of EPA and DHA when compared to other phyla (This review highlights that seaweeds are a rich, sustainable, and underexplored source of bioactive lipids with potent anti-inflammatory activity. The global burden of chronic, non-communicable diseases, including type 2 diabetes, hypertension, obesity, some types of cancer, and CVD put enormous pressure on public health services around the world. The evidence indicates that targeting inflammatory pathways could be a new strategy in the treatment and/or prevention of these chronic metabolic complications. Seaweeds are characterised by high nutritional value and their lipids can offer a number of bioactive compounds, including n-3 LC-PUFA, which may modulate the inflammatory response and reduce the risk of several NCDs. n-3 LC-PUFA are considered one of the most important anti-inflammatory components present in diets . Thus, ter phyla . The antAs the majority of work focused on the evaluation of several pro-inflammatory mediators only there is limited evidence on the complex anti-inflammatory mechanisms of seaweed lipids . The lacChronic inflammatory conditions contribute to more than 50% of all deaths globally , and sea


NRF2, TXN, and HMOX1) in oral cancer cells rather than S–G cells. Fucoidan showed DNA damage-inducible effects (γH2AX and 8-hydroxy-2-deoxyguanosine) in oral cancer cells but not in S–G cells. Accordingly, these preferential changes in oral cancer but not in non-malignant cells contribute to the preferential antiproliferation mechanism of fucoidan. Furthermore, these changes were reverted by pretreatment with the antioxidant N-acetylcysteine. Therefore, for the first time, this study provides a detailed understanding of the preferential antiproliferation effects and mechanisms of fucoidan in oral cancer cells.Fucoidan is a dietary brown algae-derived fucose-rich polysaccharide. However, the anticancer effects of fucoidan for oral cancer treatment remain unclear, particularly in terms of its preferential antiproliferation ability and oxidative-stress-associated responses. This study first evaluated the effects and mechanisms of the preferential antiproliferation of fucoidan between oral cancer and non-malignant oral cells (S–G). In a 48 h MTS assay, fucoidan showed higher antiproliferation in response to five types of oral cancer cells, but not S–G cells, demonstrating preferential antiproliferation of oral cancer cells. Oral cancer cells (Ca9-22 and CAL 27) showing high sensitivity to fucoidan were selected to explore the antiproliferation mechanism compared to S–G cells. Fucoidan showed subG1 accumulation and an annexin V increase in apoptosis, accompanied by caspase 8, 9, and 3 activations in oral cancer cells, but not in S–G cells. Fucoidan increased reactive oxygen species and mitochondrial superoxide levels and decreased cellular glutathione in oral cancer cells compared with S–G cells. These oxidative stress effects were attributed to the downregulation of antioxidant signaling genes ( Oral cancer is the most common subtype of head and neck malignancy, with a high disease burden . BesidesFucus vesiculosus, Fucus evanescens, Alaria esculenta, Ascophyllum nodosum, Macrocystis pyrifera, Sargassum polycystum, and Laminaria japonicia [Dietary natural products are commonly non-toxic. Marine algae are popular dietary foods as nutritional and functional resources with non-cytotoxicity . Marine aponicia .Fucoidan exhibits anti-inflammatory , anticoaFucoidan-induced apoptosis is validated to depend on oxidative stress in the example of bladder cancer cells . BesidesThis investigation focused on evaluating the preferential antiproliferation impact of fucoidan against oral cancer cells but not normal cells. The detailed mechanism of fucoidan-exposed oral cancer cells was explored, particularly for preferential apoptosis, oxidative stress, antioxidant signaling, and DNA damage.Fucus vesiculosus was obtained from Carbosynth . Oxidative stress inhibitor N-acetylcysteine (NAC) [Fucoidan derived from ne (NAC) ,22 , and Ca9-22  oral cancer cell lines were included. The OC-2  and OECM3/well/96-well plate for 48 h viability assays, and 2, 2, and 3 × 103/well/96-well plate for 72 h viability assays, respectively. The cells were then incubated overnight before drug treatment. After drug treatment, cells were reacted with an MTS reagent for 1 h. Cell viability is proportional to colored formazan dye and measured by an ELISA reader at 490 nm [A tetrazolium-based cell viability detecting MTS kit , was applied. Ca9-22, CAL 27, and S–G cells were seeded at a density of 4, 4, and 6 × 10t 490 nm .Cells were fixed and then mixed with 7-aminoactinmycin D (7AAD) (1 μg/mL)  for a 30 min staining reaction [Cells were double-stained with annexin V (1:1000) and 7AAD (1 μg/mL)  (Strong Caspase (Cas) 3, 8, and 9 activations are commonly examined to detect the involvements of the apoptosis executor and the extrinsic and intrinsic apoptosis signaling. Peptide-based kits for flow cytometry  were applied to detect Cas 3, 8, and 9 ,33. Cell2DCFDA)   [ROS production, MitoSOX generation, and GSH depletion were used to examine oxidative stress changes, which were detected by 2′,7′-dichlorodihydrofluorescein diacetate (H 30 min) , MitoSOX 30 min) , for ROSNRF2; NFE2L2), thioredoxin (TXN), and heme oxygenase 1 (HMOX1)  gene.RNA was extracted for cDNA conversion by Trizol solvent   and an O (HMOX1) , were inDNA damage markers γH2AX and 8-OHdG  were exaANOVA and a post hoc test  were chosen for statistical evaluation. The post hoc connecting letters are shown in each treatment. Different connecting lowercase letters represent significant differences for multi-comparison. Examples are provided in each figure legend to elucidate the results.The 48 h cell viability (%) was dose-dependently decreased by fucoidan in oral cancer cells  but was maintained in non-malignant oral cells (S–G) A. SimilaAfter fucoidan incubation, the cell cycle histograms in oral cancer and non-malignant oral cells (S–G) were generated A. For doFurthermore, the impact of oxidative stress on cell cycle progression was concerning, due to the NAC effect on rescuing viability in fucoidan-treated oral cancer cells B. The anAnnexin V detects phosphatidylserine flipflopping to the outer plasma membrane, proportional to apoptosis. Based on the dose and time experiments for annexin V/7ADD analysis, fucoidan increased the annexin V (+) populations in oral cancer cells (Ca9-22 and CAL 27) more than in non-malignant oral cells (S–G) A,B, indiFurthermore, the impact of oxidative stress in regulating apoptosis was of concern. The antioxidant NAC reversed the fucoidan-induced annexin V increment in oral cancer cells B, suggesCas 3, 8, and 9 activation is detected by peptide cleavage to generate fluorescence, proportional to the activation of these apoptosis signals as detected by flow cytometry. Flow cytometry density plots are presented in Furthermore, the impact of oxidative stress on Cas 3, 8, and 9 activations was of concern. The antioxidant NAC reversed the fucoidan-induced Cas 3, 8, and 9 increments in oral cancer cells B,D,F, suOxidative stress (ROS and MitoSOX) and the cellular antioxidant GSH were detected. Based on the dose A,C and tFurthermore, the impact of oxidative stress and GSH changes was of concern. The antioxidant NAC reversed the fucoidan-induced ROS and MitoSOX increments B,D. NAC NRF2, TXN, and HMOX1, was detected. Fucoidan dramatically downregulated these antioxidant genes in oral cancer cells (Ca9-22 and CAL 27) but not in non-malignant oral cells (S–G)  . These aDNA damage, such as γH2AX and 8-OHdG, was detected. Fucoidan increased the populations of γH2AX (+) and 8-OHdG (+) in oral cancer cells (Ca9-22 and CAL 27) but not in non-malignant oral cells (S–G) A, indicaFurthermore, the impact of oxidative stress on DNA damage was of concern. The antioxidant NAC reversed the fucoidan-induced γH2AX and 8-OHdG increments in oral cancer cells B, suggesThe anticancer functions of fucoidan in oral cancer cells have not been thoroughly investigated. How antioxidant signaling and the DNA damage response are associated with oxidative stress in fucoidan-treated oral cancer cells remains unclear. Moreover, most studies on the anticancer properties of fucoidan focused on the antiproliferation effects without considering their impacts on normal cells. The present investigation used both oral cancer and normal cells to confirm that fucoidan exhibited preferential antiproliferation of oral cancer cells without changing non-malignant oral cells. The detailed preferential antiproliferation mechanisms were discussed, as follows.Fucoidan has been applied to several oral cancer studies. For example, fucoidan-based nanoparticles were developed to deliver the phosphoinositide 3-kinase α (PI3Kα) inhibitor BYL719 to head and neck squamous cell carcinoma (HNSCC) cells . Fucoida50 values of fucoidan for oral cancer cells (SCC15 and SCC25) were 218.2 and 508.5 μg/mL in a 24 h MTT assay [50 values of fucoidan for oral cancer cells  were 856.1, 1096.9, 1193.9, and 1190.6 μg/mL, respectively, in a 48 h MTS assay, but the viability of non-malignant oral cells was unchanged. Accordingly, the present study found that fucoidan exhibited preferential antiproliferation in oral cancer cells but not in non-malignant oral cells.Recently, antiproliferation studies were reported in oral cancer cells. For example, ICTT assay ,42. More50 values of 5.6 and 1.20 μg/mL in oral cancer Ca9-22 cells at the 24 h  were downregulated in fucoidan-treated oral cancer cells but not in non-malignant oral cells  and oxidative DNA damage (8-OHdG) ,71. FucoMoreover, for the first time, we further found that fucoidan induced oxidative 8-OHdG DNA damage in oral cancer cells . MoreoveIn addition to DNA damage, oxidative stress may act on other macromolecules such as lipids and proteins , causingFucoidan arrests the cell cycle at G1 in colorectal cancer cells (HCT116)  and breaThe several responses triggered by oxidative stress warrant a detailed investigation of the role of oxidative stress in fucoidan-treated oral cancer cells by using the oxidative stress inhibitor NAC. NAC has been reported to revert fucoidan-induced apoptosis and downregulations of telomerase reverse transcriptase (TERT), c-myc, Sp1 transcription factor, and AKT expressions in bladder cancer cells . HoweverIn the present investigation, we provided a different mechanism for fucoidan and validated the effects of NAC in oral cancer cells. NAC reversed fucoidan-induced preferential impacts on antiproliferation , subG1 aFucoidan shows preferential antiproliferation in oral cancer cells. However, the in vivo role of fucoidan against oral cancer was not examined in the present study. A detailed future assessment of fucoidan using the orthotopic nude mouse model of oral cancer cells is warranted ,76.Therefore, the expected next step is to examine fucoidan in preclinical and clinical trials. As recently reviewed, clinical trials have been carried out in other cancer types ,77. The Two clinical trials underway are on its safety and biodistribution ,79, and Fucoidan is a brown algae-derived fucose-rich polysaccharide. The anticancer effects of fucoidan on oral cancer cells have rarely been reported, especially with regard to its preferential antiproliferation ability and oxidative-stress-associated responses. The present investigation examined oxidative stress, DNA damage, apoptosis, and antioxidant signaling in fucoidan-treated oral cancer cells. Using oral cancer and non-malignant cells, the preferential antiproliferation effects of fucoidan on oral cancer cells were validated for the first time in the present study. Our findings demonstrate that fucoidan changed the cell cycle progression, triggered apoptosis, turned on intrinsic and extrinsic apoptosis signaling, stimulated oxidative stress, inhibited antioxidant signaling, and improved DNA damage in oral cancer cells. At the same time, non-malignant cells showed a low level of these events. These preferential changes between oral cancer and non-malignant cells contribute to the preferential antiproliferation mechanism of fucoidan . Moreover, the action of this antiproliferation mechanism was suppressed by the antioxidant NAC, indicating that fucoidan-induced antiproliferation in oral cancer cells is dependent on oxidative stress. Therefore, the contributions of the present study shed light on our understanding of preferential antiproliferation effects and the associated mechanism of fucoidan in oral cancer cells.