Gene Level Research Articles

Abstract 4133122: Integrative Metabolomic Analysis of Triglyceride-Independent Lipoprotein Lipase Pathway Mechanisms for Coronary Artery Disease

Background: Strong genetic evidence supports the associations between key genes in triglyceride (TG) metabolism via the lipoprotein lipase (LPL) pathway (i.e., LPL , APOC3 , and ANGPTL3 ) and coronary artery disease (CAD). Given sparse human genetic or clinical trial evidence for other TG-modulating pathways, it is unclear if CAD risk reduction is due to reductions in TG levels alone or a secondary mechanism of action. Aim: To determine if there are TG-independent mechanisms of CAD risk reduction specific to the LPL pathway using genomics and metabolomics. Methods: We assessed the genetic effects of LPL pathway genes (67, 46, and 17 SNPs for LPL , APOC3 , and ANGPTL3 , respectively) and non-LPL pathway genes (3462 SNPs) for TG levels and 168 metabolites. We identified metabolites with associated effects that were greater than the associated effects with TG levels for LPL pathway genes and showed heterogeneity compared to the effects for non-LPL pathway genes. Cox proportional-hazards models were used to evaluate the association of candidate metabolites measured at baseline with incident CAD in participants of the UK Biobank. Models were adjusted for age, sex, TG levels, BMI, prevalent type 2 diabetes, statin, and ten PCs of genetic ancestry. Results: Among 233,719 UK Biobank participants free of prevalent CAD and with metabolite measurements, the mean (SD) age was 57.1 (8.0) years, 105,111 (45.0%) were male, and all self-reported as white. Over a median (IQR) follow-up of 11.2 (10.5-11.9) years, there were 7992 (3.4%) incident CAD cases. There were 30, 6, and 93 metabolites with significantly greater genetic effects for LPL , APOC3 , and ANGPTL3 than what was observed with TG levels, respectively. Among 20 metabolites with heterogeneity for ≥2 LPL pathway genes compared to non-LPL pathway genes, multi-variable Cox regression revealed 19 significantly associated with CAD (P<3x10 -4 ). Conclusion: LPL pathway genes have convergent and distinct impacts across metabolites. While LPL and ANGPTL3 have large genetic effects on VLDL metrics, APOC3 may have genetic effects beyond lipoprotein metabolism that impact CAD risk. The extent to which these metabolites influence CAD risk beyond TG alterations requires further study.

Examining The impact of Chitosan and Nano-Chitosan Addition on The expression Levels of SOD and CAT Genes in Two Lines of Japanese Quail

Examining The impact of Chitosan and Nano-Chitosan Addition on The expression Levels of SOD and CAT Genes in Two Lines of Japanese Quail

Efficacy of two different microbial consortia on salinity tolerance in chickpea: an in-planta evaluation on biochemical, histochemical, and genomic aspects.

This study aimed to identify and characterize actinobacteria and rhizobia with plant growth-promoting (PGP) traits from chickpea plants. Out of 275 isolated bacteria, 25 actinobacteria and 5 chickpea rhizobia showed 1-aminocyclopropane-1-carboxylate deaminase (ACCd) activity. Selected chickpea rhizobia were tested for their nodulating capacity under sterile and non-sterile soil conditions. Further screening on salinity and PGP traits identified three promising isolates: Nocardiopsis alba KG13, Sinorhizobium meliloti KGCR17, and Bacillus safensis KGCR11. These three isolates were analyzed for their compatibility and made into a consortium (Consortium 1). This along with another consortium made from our salinity-tolerant lab strains Chryseobacterium indologenes ICKM4 and Stenotrophomonas maltophilia ICKM15 (Consortium 2) was compared in planta studies. Trials revealed that Consortium 2 showed significant (p < 0.05) tolerance and on above-ground, below-ground traits and yield components than Consortium 1. Moreover, both consortia induced nodulation in saline-stressed plants, alleviated electrolyte leakage (2.3 vs. 0.4 in ICCV 2; 1.8 vs. 0.6 in JG 11), and increased chlorophyll content. Histochemical staining indicated reduced oxidative stress and lipid peroxidation in consortium-treated plants under salinity stress. Further, gene expression studies revealed mixed patterns, with up-regulation of antioxidant and transporter genes observed in consortium-treated plants, particularly in Consortium 2. Overall, Consortium 2 showed better gene expression levels for antioxidant and transporter genes, indicating its superior efficacy in mitigating salinity stress in chickpea plants. This study provides valuable insights into the potential use of these microbial isolates in improving chickpea productivity by enhancing salinity tolerance.

The arylbipyridine platinum (II) complex increases the level of ROS and induces lipid peroxidation in glioblastoma cells.

Here we present the biological properties of the arylbipyridine platinum (II) complex (arylbipy-Pt) and describe the potential mechanism of its antitumor action which differs from that of the well-known cisplatin. Leading to the oxidative stress and lipid peroxidation, the arylbipyridine platinum (II) complex showcases the significant cytotoxicity against the glioblastoma cells as shown by the MTT test. Using the 5-ethyl-2-deoxyuridine we study the proliferative activity of glioblastoma cells to affirm that arylbipyridine platinum (II) complex does not impede cell division or DNA replication. Staining by the MitoCLox dye and 2',7'-dichlorodihydrofluorescein diacetate demonstrates that the glioblastoma cells treated with arylbipy-Pt exhibit a strong increase of the lipid peroxidation and the stimulation of the reactive oxygen species formation. The hypothesis that arylbipy-Pt promotes oxidative death of tumor cells is confirmed by control experiments using N-acetyl-L-cysteine as an antioxidant. Further evidence for the oxidative mechanism of action is provided by real-time PCR, which shows high expression levels for genes associated with the heat shock proteins HSP27 and HSP70, which can be used as markers of tumor cell ferroptosis. To elucidate the chemical nature of the arylbipy-Pt complex activity, we perform 195Pt NMR spectroscopy and cyclic voltammetry measurements under biologically relevant conditions. The results obtained clearly indicate the structural transformation of the arylbipy-Pt complex in the DMSO-saline mixture, which is crucial for its further antitumor activity via the oxidative pathway. The found correlation between the molecular structure of arylbipy-Pt and its effect on the tumor cell cycle paves the way for the rational design of Pt complexes possessing the alternative mechanism of antitumor activity as compared to DNA intercalation, providing possible solutions to the major problems such as toxicity and drug resistance.

Human Umbilical Cord Mesenchymal Stem Cell Exosome-derived miR-335-5p Alleviated Lipopolysaccharide-induced Acute Lung Injury by Regulating the m6A Level of ITGβ4 Gene.

Acute lung injury (ALI) is a serious complication that may accompany severe pneumonia in children. Derived from human umbilical cord mesenchymal stem cell exosome (HucMSC-Exo) can contribute to the regeneration of damaged lung tissue. This study aims to investigate the impact of HucMSC-Exo on ALI and its potential mechanisms. Firstly, RT-qPCR was performed to assess the expression of miR-335-5p. Subsequently, Pearson correlation analysis was performed to examine the correlation between METTL14 and miR-335-5p, as well as the correlation between METTL14 and ITGβ4, while RNA immunoprecipitation (RIP) was used to determine the m6A modification level of ITGβ4. Additionally, molecular biology techniques were employed to evaluate the expression of glycolysis-related factors. Definitively, an LPS-induced ALI model was established to investigate the effect of miR-335-5p on mice lung tissue. miR-335-5p was found to be highly expressed in HucMSC-Exo. Transfection with miR-335-5p mimics resulted in increased glucose uptake. Pearson correlation analysis revealed a negative correlation between METTL14 and miR-335-5p, as well as between METTL14 and ITGβ4. The m6A level of ITGβ4 was elevated in ALI. Overexpression of METTL14 was found to reduce the expression of ITGβ4 and glucose levels, while overexpression of ITGβ4 reversed the effects of METTL14 overexpression. In vivo, results demonstrated that miR-335-5p could improve the extent of lung tissue lesions and reduce glycolytic levels. This study revealed the mechanism by which miR-335-5p derived from HucMSC-Exo could alleviate LPS-induced ALI by regulating the m6A modification of ITGβ4, providing a new direction for the treatment of ALI.

A basally active cGAS-STING pathway limits SARS-CoV-2 replication in a subset of ACE2 positive airway cell models.

Host factors that define the cellular tropism of SARS-CoV-2 beyond the cognate ACE2 receptor are poorly defined. Here we report that SARS-CoV-2 replication is restricted at a post-entry step in a number of ACE2-positive airway-derived cell lines due to tonic activation of the cGAS-STING pathway mediated by mitochondrial DNA leakage and naturally occurring cGAS and STING variants. Genetic and pharmacological inhibition of the cGAS-STING and type I/III IFN pathways as well as ACE2 overexpression overcome these blocks. SARS-CoV-2 replication in STING knockout cell lines and primary airway cultures induces ISG expression but only in uninfected bystander cells, demonstrating efficient antagonism of the type I/III IFN-pathway in productively infected cells. Pharmacological inhibition of STING in primary airway cells enhances SARS-CoV-2 replication and reduces virus-induced innate immune activation. Together, our study highlights that tonic activation of the cGAS-STING and IFN pathways can impact SARS-CoV-2 cellular tropism in a manner dependent on ACE2 expression levels.

Noise robustness and metabolic load determine the principles of central dogma regulation.

The processes of gene expression are inherently stochastic, even for essential genes required for growth. How does the cell maximize fitness in light of noise? To answer this question, we build a mathematical model to explore the trade-off between metabolic load and growth robustness. The model provides insights for principles of central dogma regulation: Optimal protein expression levels for many genes are in vast overabundance. Essential genes are transcribed above a lower limit of one message per cell cycle. Gene expression is achieved by load balancing between transcription and translation. We present evidence that each of these regulatory principles is observed. These results reveal that robustness and metabolic load determine the global regulatory principles that govern gene expression processes, and these principles have broad implications for cellular function.

Bacterial cross-feeding can promote gene retention by lowering gene expression costs.

Gene loss is expected in microbial communities when the benefit of obtaining a biosynthetic precursor from a neighbor via cross-feeding outweighs the cost of retaining a biosynthetic gene. However, gene cost primarily comes from expression, and many biosynthetic genes are only expressed when needed. Thus, one can conversely expect cross-feeding to repress biosynthetic gene expression and promote gene retention by lowering gene cost. Here we examined long-term bacterial cocultures pairing Escherichia coli and Rhodopseudomonas palustris for evidence of gene loss or retention in response to cross-feeding of non-essential adenine. Although R. palustris continued to externalize adenine in long-term cultures, E. coli did not accumulate mutations in purine synthesis genes, even after 700 generations. E. coli purine synthesis gene expression was low in coculture, suggesting that gene repression removed selective pressure for gene loss. In support of this explanation, R. palustris also had low transcript levels for iron-scavenging siderophore genes in coculture, likely because E. coli facilitated iron acquisition by R. palustris. R. palustris siderophore gene mutations were correspondingly rare in long-term cocultures but were prevalent in monocultures where transcript levels were high. Our data suggests that cross-feeding does not always drive gene loss, but can instead promote gene retention by repressing costly expression.

Competitive Effect of Overexpressed C-terminal of Snail-1 (CSnail) in Control of the Growth and Metastasis of Melanoma Cells.

Epithelial-to-mesenchymal transition (EMT) plays a role in the invasion and metastasis of cancer cells. During this phenomenon, Snail can promote tumor progression by upregulating mesenchymal factors and downregulating the expression of pro-apoptotic proteins. Therefore, interventions on the expression rate of Snails may show beneficial therapeutic applications. In this study, the C-terminal region of Snail1, capable of binding to E-box genomic sequences, was subcloned into the pAAV-IRES-EGFP backbone to make complete AAV-CSnail viral particles. B16F10 as a metastatic melanoma cell line, with a null expression of wild type TP53 was transduced by AAV-CSnail. Moreover, the transduced cells were analyzed for in vitro expression of apoptosis, migration, and EMT-related genes, and in vivo inhibition of metastasis. In more than 80% of the AAV-CSnail transduced cells, the CSnail gene expression competitively reduced the wild-type Snail functionality and consequently lowered the mRNA expression level of EMT-related genes. Furthermore, the transcription level of cell cycle inhibitory factor p21 and pro-apoptotic factors were promoted. The scratch test showed a decrease in the migration ability of AAV-CSnail transduced group compared to control. Finally, metastasis of cancer cells to lung tissue in the AAV-CSnail-treated B16F10 melanoma mouse model was significantly reduced, pointing out to prevention of EMT by the competitive inhibitory effect of CSnail on Snail1 and increased apoptosis of B16F10 cells. The capability of this successful competition in reducing the growth, invasion, and metastasis of melanoma cells indicates that gene therapy is a promising strategy for the control of the growth and metastasis of cancer cells.

A Six-gene Prognostic Model Based on Neutrophil Extracellular Traps (NETs)-related Gene Signature for Lung Adenocarcinoma.

Lung adenocarcinoma (LUAD) is one of the most common malignant cancers. Neutrophil extracellular traps (NETs) have been discovered to play a crucial role in the pathogenesis of LUAD. We aimed to establish an innovative prognostic model for LUAD based on the distinct expression patterns of NETs-related genes. The TCGA LUAD dataset was utilized as the training set, while GSE31210, GSE37745, and GSE50081 were undertaken as the verification sets. The patients were grouped into clusters based on the expression signature of NETs-related genes. Differentially expressed genes between clusters were identified through the utilization of the random forest and LASSO algorithms. The NETs score model for LUAD prognosis was developed by multiplying the expression levels of specific genes with their corresponding LASSO coefficients and then summing them. The validity of the model was confirmed by analysis of the survival curves and ROC curves. Additionally, immune infiltration, GSEA, mutation analysis, and drug analysis were conducted. Silencing ABCC2 in A549 cells was achieved to investigate its effect. We identified six novel NETs-related genes, namely UPK1B, SFTA3, GGTLC1, SCGB3A1, ABCC2, and NTS, and developed a NETs score signature, which exhibited a significant correlation with the clinicopathological and immune traits of the LUAD patients. High-risk patients showed inhibition of immune-related processes. Mutation patterns exhibited variability among the different groups. AZD3759, lapatinib, and dasatinib have been identified as potential candidates for LUAD treatment. Moreover, the downregulation of ABCC2 resulted in the induction of apoptosis and suppression of migration and invasion in A549 cells. Altogether, this study has identified a novel NET-score signature based on six novel NET-related genes to predict the prognosis of LUAD and ABCC2 and has also explored a new method for personalized chemo-/immuno-therapy of LUAD.

LncRNA FAM83H-AS1 Contributes to the Radio-resistance and Proliferation in Liver Cancer through Stability FAM83H Protein.

Liver cancer (LC) is one of China's most common malignant tumors, with a high mortality rate, ranking third leading cause of death after gastric and esophageal cancer. Recent patents propose the LncRNA FAM83H-AS1 has been verified to perform a crucial role in the progression of LC. LncRNA FAM83H-AS1 has been verified to perform a crucial role in the progression of LC. However, the concrete mechanism remains to be pending further investigation. This study aimed to explore the embedding mechanism of FAM83H-AS1 molecules in terms of radio sensitivity of LC and provide potentially effective therapeutic targets for LC therapy. Quantitative real-time PCR (qRT-PCR) was conducted to measure the transcription levels of genes. Proliferation was determined via CCK8 and colony formation assays. Western blot was carried out to detect the relative protein expression. A xenograft mouse model was constructed to investigate the effect of LncRNA FAM83H-AS1 on tumor growth and radio-sensitivity in vivo. The levels of lncRNA FAM83H-AS1 were remarkably increased in LC. Knockdown of FAM83H-AS1 inhibited LC cell proliferation and colony survival fraction. Deletion of FAM83H-AS1 increased the sensitivity of LC cells to 4 Gy of X-ray radiation. In the xenograft model, radiotherapy combined with FAM83H-AS1 silencing significantly reduced tumor volume and weight. Overexpression of FAM83H reversed the effects of FAM83H-AS1 deletion on proliferation and colony survival fraction in LC cells. Moreover, the over-expressing of FAM83H also restored the tumor volume and weight reduction caused by the knockdown of FAM83H-AS1 or radiation in the xenograft model. Knockdown of lncRNA FAM83H-AS1 inhibited LC growth and enhanced radiosensitivity in LC. It has the potential to be a promising target for LC therapy.

RNA-sequencing reveals a shared neurotranscriptomic profile in the medial preoptic area of highly social songbirds and rats.

Rough-and-tumble play in juvenile rats and song in flocks of adult songbirds outside a breeding context (gregarious song) are two distinct forms of non-sexual social behavior. Both are believed to play roles in the development of sociomotor skills needed for later life-history events, including reproduction, providing opportunities for low-stakes practice. Additionally, both behaviors are thought to be intrinsically rewarded and are associated with a positive affective state. Given the functional similarities of these behaviors, this study used RNA-sequencing to identify commonalities in their underlying neurochemical systems within the medial preoptic area. This brain region is implicated in multiple social behaviors, including song and play, and is highly conserved across vertebrates. DESeq2 and rank-rank hypergeometric overlap analyses identified a shared neurotranscriptomic profile in adult European starlings singing high rates of gregarious song and juvenile rats playing at high rates. Transcript levels for several glutamatergic receptor genes, such as GRIN1, GRIN2A, and GRIA1, were consistently upregulated in highly gregarious (i.e., playful/high singing) animals. This study is the first to directly investigate shared neuromodulators of positive, non-sexual social behaviors across songbirds and mammals. It provides insight into a conserved brain region that may regulate similar behaviors across vertebrates.

Enhanced bone matrix formation through a dense lamellar scaffold of chitosan, collagen type I, and hyaluronic acid

This in vitro study aimed to assess the biocompatibility and osteogenic inducing properties of a new scaffold composed of chitosan, collagen type I, and hyaluronic acid. For in vitro assays, pre-osteoblastic immortalized cells were cultivated in standard medium (SD) and osteogenic medium (OM) in the following groups: a) Control (C) – cells cultured directly on the polystyrene plate, and b) Chitosan + Collagen type I + Hyaluronic Acid (CH + COL + HA) - cells cultured in a scaffold produced by the association of type I collagen, chitosan and hyaluronic acid. Cells in CH+COL+ HA scaffold presented metabolic activity similar to the control group. After 14 days, the CH+ COL+ HA scaffold induced a higher mineral nodule deposition compared than the control group, regardless of the cultured condition (SD or OM medium). In addition, the CH + COL + HA scaffold itself increased of the alkaline phosphatase activity and mRNA levels for Runx2 and Ocn genes, which occurred earlier than control group. Based on the results, it is possible to conclude that the dense lamellar scaffold composed of CH/COL (type I)/HA stimulated osteogenic phenotype maturation of cells and can be a promising material for future bone regenerative approaches.

Methionine supplementation regulates eggshell quality and uterine transcriptome in late-stage broiler breeders

This study aimed to compare the effects of dietary methionine (Met) and 2-hydroxy-4-(methylthio)-butanoate (HMTBA) on the eggshell quality of broiler breeder hens and elucidate the mechanism of Met in improving eggshell quality from the perspectives of eggshell microstructure and shell gland physiological function. A total of 720 WOD188 broiler breeder hens at 40 wk old were assigned to 3 groups, with 8 replicates per group and 30 birds per replicate. Over 7 weeks, birds were fed a basal diet or the same diet supplemented with 0.15% Met or 0.17% HMTBA. Our findings revealed significant improvements in the Met group for egg shape index, shell thickness, breaking strength, and fracture toughness (P < 0.05), whereas the HMTBA group showed no significant improvements (P > 0.05). Met supplementation increased calcium and phosphorus levels in both serum and shell gland tissue (P < 0.05), and enhanced Ca2+ ATPase activity in shell gland tissue (P < 0.05). Histomorphological changes included enhanced mucosal fold dimensions and increased epithelial height in the shell gland (P < 0.05). Met also improved eggshell ultrastructure, resulting in a thicker effective layer and broader mammillae with fewer type B structures (P < 0.05). The mRNA levels for genes regulating eggshell ultrastructure, such as OC-116, CALB1, and ITM2C, were significantly upregulated in the Met group (P < 0.05). Transcriptome analysis identified 248 differentially upregulated genes in the Met group, primarily linked to the non-canonical Wnt/Ca2+ signaling pathway, crucial for calcium ion transport and cellular proliferation. This research highlights that Met supplementation improves eggshell quality by enhancing calcium transport and cellular proliferation in uterine function, particularly through the modulation of WNT11 and CALB1, influencing calcium deposition and ultrastructural development.

Synonymous variant at the terminal nucleotide in exon 3 of F7 causes abnormal splicing: A case report.

Synonymous variants are non-pathogenic due to non-substitution of amino acids. However, synonymous exonic terminal nucleotide substitutions may affect splicing. Splicing variants are easily analyzed at RNA level for genes expressed in blood cells. Minigene analysis provides another method for splicing variant analysis of genes that are poorly or not expressed in peripheral blood. Whole exome sequencing was performed to screen for potential pathogenic mutations in the proband, which were validated within the family by Sanger sequencing. The pathogenicity of the synonymous mutation was analyzed using the minigene technology. The proband harbored the compound heterogeneous variants c. [291G >A; 572-50C >T] and c.681 + 1G >T in F7, of which the synonymous variant c.291G >A was located at the terminal position of exon 3. Minigene analysis revealed exon3 skipping due to this mutation, which may have subsequently affected protein sequence, structure, and function. Our finding confirmed the pathogenicity of c.291G >A, thus extending the pathogenic mutation spectrum of F7, and providing insights for effective reproductive counseling.

Co-exposure to microplastic and plastic additives causes development impairment in zebrafish embryos

Since the run off of microplastic and plastic additives into the aquatic environment through the disposal of plastic products, we investigated the adverse effects of co-exposure to microplastics and plastic additives on zebrafish embryonic development. To elucidate the combined effects between microplastic mixtures composed of microplastics and plastic additives in zebrafish embryonic development, polystyrene (PS), bisphenol S (BPS), and mono-(2-ethylhexyl) phthalate (MEHP) were chosen as a target contaminant. Based on non-toxic concentration of each contaminant in zebrafish embryos, microplastic mixtures which is consisted of binary and ternary mixed forms were prepared. A strong phenotypic toxicity to zebrafish embryos was observed in the mixtures composed with non-toxic concentration of each contaminant. In particular, the mixture combination with ≤ EC10 values for BPS and MEHP showed a with a strong synergistic effect. Based on phenotypic toxicity to zebrafish embryos, change of transcription levels for target genes related to cell damage and thyroid hormone synthesis were analyzed in the ternary mixtures with low concentrations that were observed non-toxicity. Compared with the control group, cell damage genes linked to the oxidative stress response and thyroid hormone transcription factors were remarkably down-regulated in the ternary mixture-exposed groups, whereas the transcriptional levels of cyp1a1 and p53 were significantly up-regulated in the ternary mixture-exposed groups (P < 0.05). These results demonstrate that even at low concentrations, exposure to microplastic mixtures can cause embryonic damage and developmental malformations in zebrafish, depending on the mixed concentration-combination. Consequently, our findings will provide data to examine the action mode of zebrafish developmental toxicity caused by microplastic mixtures exposure composed with microplastics and plastic additives.

The pentatricopeptide repeat protein TCD6 functions RNA editing and cleavage of ndhA and is required for chloroplast development in early rice seedlings

Pentatricopeptide repeat (PPR) proteins compose one of the largest protein families in higher plants and play a role in regulating organellar gene expression. In this study, we discovered that a new rice mutant tcd6 exhibited albino phenotype and aberrant chloroplast before the three-leaf (autotrophic) seedling stage. Through Map-based cloning and complementation tests, it was shown that TCD6 encodes a chloroplast-located PPR protein, with 14 PPR motifs and an atypical DYW-like motif. In addition, the disruption of TCD6 hindered the nuclear-encoded polymerase (NEP)-dependent transcript levels for plastid genes and led to defects in the cleavage and editing of ndhA (encoding NDH subunit) in early tcd6 mutant seedlings. Taken together, our results indicate that TCD6 is indispensable for chloroplast development and involves in RNA editing and cleavage of ndhA during early seedling (autotrophic) growth of rice.

Suppression of GATA3 promotes epithelial-mesenchymal transition and simultaneous cellular senescence in human extravillous trophoblasts

The regulatory mechanism of the transcription factor GATA3 in the differentiation and maturation process of extravillous trophoblasts (EVT) in early pregnancy placenta, as well as its relevance to the occurrence of pregnancy disorders, remains poorly understood. This study leveraged single-cell RNA sequencing data from placental organoid models and placental tissue to explore the dynamic changes in GATA3 expression during EVT maturation. The expression pattern exhibited an initial upregulation followed by subsequent downregulation, with aberrant GATA3 localization observed in cases of recurrent miscarriage (RM). By identifying global targets regulated by GATA3 in primary placental EVT cells, JEG3, and HTR8/SVneo cell lines, this study offered insights into its regulatory mechanisms across different EVT cell models. Shared regulatory targets among these cell types and activation of trophoblast cell marker genes emphasized the importance of GATA3 in EVT differentiation and maturation. Knockdown of GATA3 in JEG3 cells led to repression of GATA3-induced epithelial-mesenchymal transition (EMT), as evidenced by changes in marker gene expression levels and enhanced migration ability. Additionally, interference with GATA3 accelerated cellular senescence, as indicated by reduced proliferation rates and increased activity levels for senescence-associated β-galactosidase enzyme, along with elevated expression levels for senescence-associated genes. This study provides comprehensive insights into the dual role of GATA3 in regulating EMT and cellular senescence during EVT differentiation, shedding light on the dynamic changes in GATA3 expression in normal and pathological placental conditions.

Effects of inactivated Streptococcus iniae, Edwardsiella tarda, and Poly I:C on mRNA Expression Levels of CXCL-10 and CXCL-9 Genes in Japanese Flounder

The present study accomplished the successful cloning and sequencing of the JfCXCL9_L and JfCXCL10_L genes found in the spleen cDNA of the Japanese flounder (Paralichthys olivaceus). The tissue distribution of these two genes was determined before any stimuli administration at the zero-hour mark. The qRT-PCR analysis demonstrated that the expression of JfCXCL10_L closely mirrored that of IL1-β, displaying an upregulation following the application of Poly I:C (Viral mimic) and formalin-killed Edwardsiella tarda (Gram-Negative mimic), while showing a downregulation after the application of formalin-killed Streptococcus iniae (Gram-Positive mimic) treatment. These findings strongly suggest a role for JfCXCL10_L in the immune response to viral and gram-negative bacterial stimuli. Regarding JfCXCL9_L, mRNA levels were found to be significantly downregulated after FKCET and FKCSI treatments, though to varying extents. Interestingly, at specific time points, JfCXCL9_L levels were even lower compared to Poly I:C treatment. These intriguing findings shed valuable light on the roles of both JfCXCL9_L and JfCXCL10_L in potential functions of immune response mechanisms of the Japanese flounder'.

Phytol and α-Bisabolol Synergy Induces Autophagy and Apoptosis in A549 Cells and Additional Molecular Insights through Comprehensive Proteome Analysis via Nano LC-MS/MS.

Non-Small Cell Lung Cancer (NSCLC) is a malignancy with a significant prevalence and aggressive nature, posing a considerable challenge in terms of therapeutic interventions. Autophagy and apoptosis, two intricate cellular processes, are integral to NSCLC pathophysiology, each affecting the other through shared signaling pathways. Phytol (Phy) and α-bisabolol (Bis) have shown promise as potential anticancer agents individually, but their combined effects in NSCLC have not been extensively investigated. The present study was to examine the synergistic impact of Phy and Bis on NSCLC cells, particularly in the context of autophagy modulation, and to elucidate the resulting differential protein expression using LCMS/ MS analysis. The A549 cell lines were subjected to the patented effective concentration of Phy and Bis, and subsequently, the viability of the cells was evaluated utilizing the MTT assay. The present study utilized real-time PCR analysis to assess the expression levels of crucial apoptotic genes, specifically Bcl-2, Bax, and Caspase-9, as well as autophagy-related genes, including Beclin-1, SQSTM1, Ulk1, and LC3B. The confirmation of autophagy marker expression (Beclin-1, LC3B) and the autophagy-regulating protein SQSTM1 was achieved through the utilization of Western blot analysis. Differentially expressed proteins were found using LC-MS/MS analysis. The combination of Phy and Bis demonstrated significant inhibition of NSCLC cell growth, indicating their synergistic effect. Real-time PCR analysis revealed a shift towards apoptosis, with downregulation of Bcl-2 and upregulation of Bax and Caspase-9, suggesting a shift towards apoptosis. Genes associated with autophagy regulation, including Beclin-1, SQSTM1 (p62), Ulk1, and LC3B, showed significant upregulation, indicating potential induction of autophagy. Western blot analysis confirmed increased expression of autophagy markers, such as Beclin-1 and LC3B, while the autophagy-regulating protein SQSTM1 exhibited a significant decrease. LC-MS/MS analysis revealed differential expression of 861 proteins, reflecting the modulation of cellular processes. Protein-protein interaction network analysis highlighted key proteins involved in apoptotic and autophagic pathways, including STOML2, YWHAB, POX2, B2M, CDA, CAPN2, TXN, ECHS1, PEBP1, PFN1, CDC42, TUBB1, HSPB1, PXN, FGF2, and BAG3, emphasizing their crucial roles. Additionally, PANTHER pathway analysis uncovered enriched pathways associated with the differentially expressed proteins, revealing their involvement in a diverse range of biological processes, encompassing cell signaling, metabolism, and cellular stress responses. The combined treatment of Phy and Bis exerts a synergistic inhibitory effect on NSCLC cell growth, mediated through the interplay of apoptosis and autophagy. The differential protein expression observed, along with the identified proteins and enriched pathways, provides valuable insights into the underlying molecular mechanisms. These findings offer a foundation for further exploration of the therapeutic potential of Phy and Bis in the management of NSCLC.