Pathway Genes Research Articles

Photoinduction of Ferroptosis and cGAS-STING Activation by a H2S-Responsive Iridium(III) Complex for Cancer-Specific Therapy.

Triggering ferroptosis represents a promising anticancer therapeutic strategy, but the development of a selective ferroptosis inducer for cancer-specific therapy remains a great challenge. Herein, a H2S-responsive iridium(III) complex NA-Ir has been well-designed as a ferroptosis inducer. NA-Ir could selectively light up H2S-rich cancer cells, primarily localize in mitochondria, intercalate into mitochondrial DNA (mtDNA), and induce mtDNA damage, exhibiting higher anticancer activity under light irradiation. Mechanistic studies showed that NA-Ir-mediated PDT triggered lipid peroxidation and glutathione peroxidase 4 downregulation through ROS production and GSH depletion, resulting in ferroptosis through multiple pathways. Moreover, the intense mtDNA damage can activate the cyclic GMP-AMP synthase-stimulator of the interferon gene (cGAS-STING) pathway, leading to ferritinophagy and further ferroptosis. RNA-sequencing analysis showed that NA-Ir-mediated PDT mainly affects the expression of genes related to ferroptosis, autophagy, and cancer immunity. This study demonstrates the first cancer-specific example with ferroptosis and cGAS-STING activation, which provides a new strategy for multimodal synergistic therapy.

Changes in primary metabolism and associated gene expression during host-pathogen interaction in clubroot resistance of Brassica napus.

The role of primary metabolism during Brassica napus-Plasmodiophora brassicae interaction leading to clubroot resistance has not yet been investigated thoroughly. In this study, we investigated some of the primary metabolites and their derivatives as well as expression of the genes involved in their biosynthesis to decipher this host-pathogen interaction. For this, two sets (clubroot resistant and susceptible) of canola lines were inoculated with P. brassicae pathotype 3A to investigate the endogenous levels of primary metabolites at 7-, 14-, and 21-days after inoculation (DAI). The associated pathways were curated, and expression of the selected genes was analyzed using qRT-PCR. Our results suggested the possible involvement of polyamines (spermidine and spermine) in clubroot susceptibility. Some of the amino acids were highly abundant at 7- or 14-DAI in both resistant and susceptible lines; however, glutamine and the amino acid derivative phenylethylamine showed higher endogenous levels in the resistant lines at later stages of infection. Organic acids such as malic, fumaric, succinic, lactic and citric acids were abundant in the susceptible lines. Conversely, the abundance of salicylic acid (SA) and the expression of benzoate/salicylate carboxyl methyltransferase (BSMT) were higher in the resistant lines at the secondary stage of infection. A reduced disease severity index and gall size were observed when exogenous SA (1.0 mM) was applied to susceptible B. napus; this further supported the role of SA in clubroot resistance. In addition, a higher accumulation of fatty acids and significant upregulation of the pathway genes, glycerol-3-phosphate dehydrogenase (GPD) and amino alcohol phosphotransferase (AAPT) were observed in the resistant lines at 14- and 21-DAI. In contrast, some of the fatty acid derivatives such as phosphatidylcholines represented a lower level in the resistant lines. In conclusion, our findings provided additional insights into the possible involvement of primary metabolites and their derivatives in clubroot resistance.

Genome-wide analysis of DNA methylation and transcriptional changes associated with overwintering memory in Brassica rapa L. grown in the field

BackgroundWinter rapeseed, the sole overwintering oilseed crop in northern China, emphasizes winter resilience, yet epigenetic regulatory mechanisms governing overwintering memory remain poorly understood.ResultsIn this study, the root collar tissues from the robust cold-resistant variety Longyou-7 were sampled during the pre-winter period (S1), overwintering periods (S2–S5), and re-greening period (S6), to analyze overall genomic DNA methylation levels using high-performance liquid chromatography (HPLC). The result showed that DNA methylation level exceeded 80% in the S1 stage. Throughout the overwintering periods, methylation levels displayed a decreasing trend in S3, followed by an increase in S5, and a pronounced decrease in S6. Consequently, S1, S3, S5, and S6 periods were chosen for whole-genome bisulfite sequencing analyses to elucidate the overwintering memory mechanisms of Longyou-7. The result revealed that DNA methylation primarily occurs in the CG context in Longyou-7. However, methylation of mC sites is most prevalent in the CHH type, gradually decreasing during overwintering periods. Analysis of methylation patterns in specific genomic regions of Longyou-7 showed that the highest methylation levels in the intergenic region. Moreover, mC sites in repeats and transposon elements are distributed differently across the three contexts. Subsequently, differentially methylated regions and promoters of Longyou-7 were identified during various periods compared to the S1 stage, followed by joint analysis with transcriptome sequencing. Functional enrichment analysis highlighted the involvement of most overlapping genes in the MAPK signaling pathway, plant hormone signal transduction, and starch and sucrose metabolism pathways. Changes in candidate gene expression within these three pathways correlated closely with DNA methylation levels.ConclusionsOur findings underscored the critical role of DNA methylation in regulating the expression of overwintering memory genes in winter rapeseed. These results offer a comprehensive insights into the epigenetic regulatory mechanisms governing winter rapeseed's overwintering memory, while identified overwintering memory genes served as crucial genetic resources for multifaceted breeding of winter-resistant varieties.Graphical

Notch signaling mediated repressive effects of resveratrol in inducing caspasedependent apoptosis in MCF-7 breast cancer cells.

Resveratrol, a potent anticancer bioactive compound, has been shown to trigger apoptosis in numerous cancer cells. Although Notch signaling promotes breastcancer apoptosis, it is unclear whether resveratrolinduces apoptosis in MCF-7cells via influencing the Notch pathway. This study aimed to evaluate the effect of resveratrolon modulating Notch signaling targetsand provide critical information for employing resveratrolin breastcancer therapy. Thus, in this study, we have deciphered the effect of resveratrol against three potent genes (Notch1, Jagged1, and DLL4) of the notch signaling pathway. For mechanistic studies, in silico, and in vitro analysis was executed to investigate the apoptotic-inducing potential of resveratrol against three selected oncogenes involved in the progression of breast cancer. Docking analysis revealed the inhibitory potential of resveratrol against all three selected targets of the Notch pathway (Notch1: -5.0; Jagged-1: -5.9; DLL4: -5.8). In vitro, findings further displayed a significant reduction in cell viability in resveratrol-treated MCF-7 cancer cells, which were concomitantly related to the downregulation of Notch-1, Jagged-1, and DLL4. Moreover, the antiproliferative efficacy of resveratrol was correlated with apoptosis and modulation in the expression of Bax, Bcl-2, cyclin D1, CDK4, p21, and caspase-3 activation. Taken together, these experimental findings suggested that apoptotic inducing potential of resveratrol was mediated through a novel mechanism involving suppression of the Notch signaling pathway.

Distinct Variations in Gene Expression and Cell Composition across Lichen Planus Subtypes.

Lichen planus (LP) is a highly prevalent inflammatory skin disease. While various clinical subtypes have been defined, detailed comparisons of these variants are lacking. This study aimed to elucidate differences in gene expression and cellular composition across LP subtypes. Lesional skin biopsies from 28 LP patients (classical, oral, genital, and lichen planopilaris) and seven non-diseased skin controls (NDC) were analyzed. Gene expression profiling of 730 inflammation-related genes was conducted using NanoString. Immune cell compositions were assessed by multiplex immunohistochemistry. Gene expression profiles revealed unique inflammatory signatures for each LP subtype. Lichen planopilaris exhibited the most divergence, with downregulated gene expression and upregulation of complement pathway genes (C5-7), along with elevated M2 macrophages. Oral and genital LP demonstrated similar profiles with strong upregulation of TNF-related and Toll-like receptor-associated genes. Oral LP showed the highest upregulation of cytotoxicity-associated genes, as well as high numbers of CD8+ IL-17A+ (Tc17) cells (8.02%). Interferon gene signatures were strongly upregulated in oral and classical LP. The study highlights distinct differences in inflammatory gene expression and cell composition across LP subtypes, emphasizing the need for tailored therapeutic approaches.

Mycorrhizal and non-mycorrhizal perennial ryegrass roots exhibit differential regulation of lipid and Ca2+ signaling pathways in response to low and high temperature stresses

Lipids and Ca2+ are involved as intermediate messengers in temperature-sensing signaling pathways. Arbuscular mycorrhizal (AM) symbiosis is a mutualistic symbiosis between fungi and terrestrial plants that helps host plants cope with adverse environmental conditions. Nonetheless, the regulatory mechanisms of lipid- and Ca2+-mediated signaling pathways in mycorrhizal plants under cold and heat stress have not been determined. The present work focused on investigating the lipid- and Ca2+-mediated signaling pathways in arbuscular mycorrhizal (AM) and non-mycorrhizal (NM) roots under temperature stress and determining the role of Ca2+ levels in AM symbiosis and temperature stress tolerance in perennial ryegrass (Lolium perenne L.) Compared with NM plants, AM symbiosis increased phosphatidic acid (PA) and Ca2+ signaling in the roots of perennial ryegrass, increasing the expression of genes associated with low temperature (LT) stress, including LpICE1, LpCBF3, LpCOR27, LpCOR47, LpIRI, and LpAFP, and high temperature (HT) stress, including LpHSFC1b, LpHSFC2b, LpsHSP17.8, LpHSP22, LpHSP70, and LpHSP90, under LT and HT conditions. These effects result in modulated antioxidant enzyme activities, reduced lipid peroxidation, and suppressed growth inhibition caused by LT and HT stresses. Furthermore, exogenous Ca2+ application enhanced AM symbiosis, leading to the upregulation of Ca2+ signaling pathway genes in roots and ultimately promoting the growth of perennial ryegrass under LT and HT stresses. These findings shed light on lipid and Ca2+ signal transduction in AM-associated plants under LT and HT stresses, emphasizing that Ca2+ enhances cold and heat tolerance in mycorrhizal plants.

Nano-alumina induced developmental and neurobehavioral toxicity in the early life stage of zebrafish, associated with mTOR

The study aims to investigate the effects of nano-alumina (AlNPs) on the early development and neurobehavior of zebrafish and the role of mTOR in this process. After embryos and grown-up larvae exposed to AlNPs from 0 to 200 μg/mL, we examined the development, neurobehavior, AlNPs content, and mTOR pathway genes. Moreover, embryos were randomly administered with control, negative control, mTOR knockdown, AlNPs, and mTOR knockdown + AlNPs, then examined for development, neurobehavior, oxidative stress, neurotransmitters, and development genes. As AlNPs increased, swimming speed and distance initially increased and then decreased; thigmotaxis and panic-avoidance reflex substantially decreased in the high-dose AlNPs group; aluminum and nanoparticles considerably accumulated in the 100 μg/mL AlNPs group; AlNPs at high dose decreased mTOR gene and protein levels, stimulating autophagy via increasing ULK1 and ULK2. mTOR knockdown exacerbated the harm to normal development rate, eye and body length, and neurobehavior induced by AlNPs through raising ROS, SOD, and ACH levels but decreasing AchE activity and development genes. Therefore, AlNPs suppress neurobehavior through downregulating mTOR, and mTOR knockdown further aggravates their early development and neurobehavior loss, suggesting mTOR could be a potential target for the toxicity of AlNPs.

EjWRKY6 Is Involved in the ABA-Induced Carotenoid Biosynthesis in Loquat Fruit during Ripening.

The yellow-fleshed loquat is abundant in carotenoids, which determine the fruit's color, provide vitamin A, and offer anti-inflammatory and anti-cancer health benefits. In this research, the impact of abscisic acid (ABA), a plant hormone, on carotenoid metabolism and flesh pigmentation in ripening loquat fruits was determined. Results revealed that ABA treatment enhanced the overall content of carotenoids in loquat fruit, including major components like β-cryptoxanthin, lutein, and β-carotene, linked to the upregulation of most genes in the carotenoid biosynthesis pathway. Furthermore, a transcription factor, EjWRKY6, whose expression was induced by ABA, was identified and was thought to play a role in ABA-induced carotenoid acceleration. Transient overexpression of EjWRKY6 in Nicotiana benthamiana and stable genetic transformation in Nicotiana tabacum with EjWRKY6 indicated that both carotenoid production and genes related to carotenoid biosynthesis could be upregulated in transgenic plants. A dual-luciferase assay proposed a probable transcriptional control between EjWRKY6 and promoters of genes associated with carotenoid production. To sum up, pre-harvest ABA application could lead to carotenoid biosynthesis in loquat fruit through the EjWRKY6-induced carotenoid biosynthesis pathway.

Unveiling Endogenous Serum Peptides as Potential Biomarkers for Hepatocellular Carcinoma in Patients with Liver Cirrhosis.

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide, mainly associated with liver cirrhosis. Current diagnostic methods for HCC have limited sensitivity and specificity, highlighting the need for improved early detection and intervention. In this study, we used a comprehensive approach involving endogenous peptidome along with bioinformatics analysis to identify and evaluate potential biomarkers for HCC. Serum samples from 40 subjects, comprising 20 HCC cases and 20 patients with liver cirrhosis (CIRR), were analyzed. Among 2568 endogenous peptides, 67 showed significant differential expression between the HCC vs CIRR. Further analysis revealed three endogenous peptides (VMHEALHNHYTQKSLSLSPG, NRFTQKSLSLSPG, and SARQSTLDKEL) that showed far better performance compared to AFP in terms of area under the receiver operating characteristic curve (AUC), showcasing their potential as biomarkers for HCC. Additionally, endogenous peptide IAVEWESNGQPENNYKT that belongs to the precursor protein Immunoglobulin heavy constant gamma 4 was detected in 100% of the HCC group and completely absent in the CIRR group, suggesting a promising diagnostic biomarker. Gene ontology and pathway analysis revealed the potential involvement of these dysregulated peptides in HCC. These findings provide valuable insights into the molecular basis of HCC and may contribute to the development of improved diagnostic methods and therapeutic targets for HCC.

Bioinformatics Analysis Screening and Identification of Key Biomarkers and Drug Targets in Human Glioblastoma.

Glioblastoma is the most common type of brain cancer, with a prognosis that is unfortunately poor. Despite considerable progress in the field, the intricate molecular basis of this cancer remains elusive. The aim of this study was to identify genetic indicators of glioblastoma and reveal the processes behind its development. The advent and integration of supercomputing technology have led to a significant advancement in gene expression analysis platforms. Microarray analysis has gained recognition for its pivotal role in oncology, crucial for the molecular categorization of tumors, diagnosis, prognosis, stratification of patients, forecasting tumor responses, and pinpointing new targets for drug discovery. Numerous databases dedicated to cancer research, including the Gene Expression Omnibus (GEO) database, have been established. Identifying differentially expressed genes (DEGs) and key genes deepens our understanding of the initiation of glioblastoma, potentially unveiling novel markers for diagnosis and prognosis, as well as targets for the treatment of glioblastoma. This research sought to discover genes implicated in the development and progression of glioblastoma by analyzing microarray datasets GSE13276, GSE14805, and GSE109857 from the GEO database. DEGs were identified, and a function enrichment analysis was performed. Additionally, a protein-protein interaction network (PPI) was constructed, followed by module analysis using the tools STRING and Cytoscape. The analysis yielded 88 DEGs, consisting of 66 upregulated and 22 downregulated genes. These genes' functions and pathways primarily involved microtubule activity, mitotic cytokinesis, cerebral cortex development, localization of proteins to the kinetochore, and the condensation of chromosomes during mitosis. A group of 27 pivotal genes was pinpointed, with biological process analysis indicating significant enrichment in activities, such as division of the nucleus during mitosis, cell division, maintaining cohesion between sister chromatids, segregation of sister chromatids during mitosis, and cytokinesis. The survival analysis indicated that certain genes, including PCNA clamp-associated factor (PCLAF), ribonucleoside- diphosphate reductase subunit M2 (RRM2), nucleolar and spindle-associated protein 1 (NUSAP1), and kinesin family member 23 (KIF23), could be instrumental in the development, invasion, or recurrence of glioblastoma. The identification of DEGs and key genes in this study advances our comprehension of the molecular pathways that contribute to the oncogenesis and progression of glioblastoma. This research provides valuable insights into potential diagnostic and therapeutic targets for glioblastoma.

Gene expression changes in Brodmann’s Area 46 differentiate epidermal growth factor and immune system interactions in schizophrenia and mood disorders

How early in life stress-immune related environmental factors increase risk predisposition to schizophrenia remains unknown. We examined if pro-inflammatory changes perturb the brain epidermal growth factor (EGF) system, a system critical for neurodevelopment and mature CNS functions including synaptic plasticity. We quantified genes from key EGF and immune system pathways for mRNA levels and eight immune proteins in post-mortem dorsolateral prefrontal (DLPFC; Brodmann’s Area (BA) 46) and orbitofrontal (OFC; BA11) cortices from people with schizophrenia, mood disorders and neurotypical controls. In BA46, 64 genes were differentially expressed, predominantly in schizophrenia, where attenuated expression of the MAPK-ERK, NRG1-PI3K-AKT and mTOR cascades indicated reduced EGF system signalling, and similarly diminished immune molecular expression, notably in TLR, TNF and complement pathways, along with low NF-κB1 and elevated IL12RB2 protein levels were noted. There was nominal evidence for altered convergence between ErbB-PI3K-AKT-mTOR and TLR pathways in BA46 in schizophrenia. Comparatively minimal changes were noted in BA11. Overall, distinct pathway gene expression changes may reflect variant pathological processes involving immune and EGF system signalling between schizophrenia and mood disorder, particularly in DLPFC. Further, the abnormal convergence between innate immune signalling and candidate EGF signalling pathways may indicate a pathologically important interaction in the developing brain in response to environmental stressors.

Metabolic engineering of Saccharomyces cerevisiae for enhanced taxadiene production

BackgroundMetabolic engineering enables the sustainable and cost-efficient production of complex chemicals. Efficient production of terpenes in Saccharomyces cerevisiae can be achieved by recruiting an intermediate of the mevalonate pathway. The present study aimed to evaluate the engineering strategies of S. cerevisiae for the production of taxadiene, a precursor of taxol, an antineoplastic drug.ResultSCIGS22a, a previously engineered strain with modifications in the mevalonate pathway (MVA), was used as a background strain. This strain was engineered to enable a high flux towards farnesyl diphosphate (FPP) and the availability of NADPH. The strain MVA was generated from SCIGS22a by overexpressing all mevalonate pathway genes. Combining the background strains with 16 different episomal plasmids, which included the combination of 4 genes: tHMGR (3-hydroxy-3-methylglutaryl-CoA reductase), ERG20 (farnesyl pyrophosphate synthase), GGPPS (geranyl diphosphate synthase) and TS (taxadiene synthase) resulted in the highest taxadiene production in S. cerevisiae of 528 mg/L.ConclusionOur study highlights the critical role of pathway balance in metabolic engineering, mainly when dealing with toxic molecules like taxadiene. We achieved significant improvements in taxadiene production by employing a combinatorial approach and focusing on balancing the downstream and upstream pathways. These findings emphasize the importance of minor gene expression modification levels to achieve a well-balanced pathway, ultimately leading to enhanced taxadiene accumulation.

AMPK-mediated regulation of cardiac energy metabolism: Implications for thermotolerance in Argopecten irradians irradians

AMPK is a key regulator of metabolism in eukaryotes across various pathways related to energy regulation. Although extensive investigations of AMPK have been conducted in mammals and some model organisms, research on AMPK in scallops is comparatively limited. In this study, three AMPK family genes (AiAMPKα, AiAMPKβ and AiAMPKγ) in scallop Argopecten irradians irradians were identified through genome scanning. Structure prediction and phylogenetic analyses of AiAMPKs were performed to determine their structural features and evolutionary relationships. Spatiotemporal expression patterns of AiAMPKs at different developmental stages and in healthy adult tissues were analyzed to elucidate the function of AiAMPKs in bay scallops’ growth and development. The spatiotemporally specific expression of AiAMPKs implied their important roles in growth and development of bay scallops. Heat stress experiment was performed to determine the regulations of AiAMPKs in four kinds of thermosensitive tissues. Expression profiles revealed distinct molecular mechanisms of AiAMPKs in different tissues in response to heat stress: significant down-regulations in mobile hemocytes, but dominant up-regulations occurring in stationary gills, mantles and hearts. Functional verification including knock-down of AiAMPKα and inhibition of AiAMPK was separately conducted in the thermotolerant tissue heart at the post-transcription and translation levels. The thermotolerant index Arrhenius break temperature (ABT) showed a significant decrease and the rate-amplitude product (RAP) peaked earlier in the individuals after RNAi targeting AiAMPKα, displaying an earlier transition to anaerobic metabolism under heat stress, indicating an impairing ability of aerobic metabolism. After AiAMPK inhibition, widespread down-regulations of genes in key energy metabolism pathways, RNA polymerase II-mediated transcription, and aminoacyl-tRNA synthesis pathways were obviously observed, revealing the post-translational inhibition of AiAMPK hindered cardiac energy metabolism, basal transcription and translation. Overall, our findings provide evidences for exploring the molecular mechanisms of energy regulation in thermotolerant traits in bay scallops under ongoing global warming.

Transcriptomic landscape of quiescent and proliferating human corneal stromal fibroblasts

This study analyzed the transcriptional changes in primary human corneal stromal fibroblasts (hCSFs) grown under quiescent (serum-free) and proliferating (serum-supplemented) culture conditions to identify genes, pathways, and protein‒protein interaction networks influencing corneal repair and regeneration. Primary hCSFs were isolated from donor human corneas and maintained in serum-free or serum-laden conditions. RNA was extracted from confluent cultures using Qiagen kit and subjected to RNA sequencing (RNAseq) analysis. Differential gene expression (DGE) and pathway enrichment analyses were conducted using DESeq2 and Gene Set Enrichment Analysis (GSEA), respectively. Protein‒protein interaction (PPI) networks were created exploiting the STRING database and analyzed with Cytoscape and the cytoHubba plugin. RNA-seq revealed 5,181 genes that were significantly differentially expressed/changed among the 18,812 annotated genes (p value ˂0.05). A cutoff value of a log2-fold change of ±1.5 or greater was used to identify 674 significantly upregulated and 771 downregulated genes between quiescent and proliferating hCSFs. Pathway enrichment analysis revealed significant changes in genes linked to cell cycle regulation, inflammatory, and oxidative stress response pathways, such as E2F Targets, G2M Checkpoint, and MYC Targets, TNFA signaling via NF-kB, and oxidative phosphorylation. Protein-protein interaction network analysis highlighted critical hub genes. The FGF22, CD34, ASPN, DPT, LUM, FGF10, PDGFRB, ECM2, DCN, VEGFD, OMD, OGN, ANGPT1, CDH5, and PRELP were upregulated, whereas genes linked to cell cycle regulation and mitotic progression, such as BUB1, TTK, KIF23, KIF11, BUB1B, DLGAP5, NUSAP1, CCNA2, CCNB1, BIRC5, CDK1, KIF20A, AURKB, KIF2C, and CDCA8, were downregulated. The RNA sequences and gene count files have been submitted to the Gene Expression Omnibus (accession # GSE260476). Our study provides a comprehensive information on the transcriptional and molecular changes in hCSFs under quiescent and proliferative conditions and highlights key pathways and hub genes.

Distinct FLT3 Pathways Gene Expression Profiles in Pediatric De Novo Acute Lymphoblastic and Myeloid Leukemia with FLT3 Mutations: Implications for Targeted Therapy.

Activating FLT3 mutations plays a crucial role in leukemogenesis, but identifying the optimal candidates for FLT3 inhibitor therapy remains controversial. This study aims to explore the impacts of FLT3 mutations in pediatric acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) and to compare the mutation profiles between the two types to inspire the targeted application of FLT3 inhibitors. We retrospectively analyzed 243 ALL and 62 AML cases, grouping them into FLT3-mutant and wild-type categories, respectively. We then assessed the associations between FLT3 mutations and the clinical manifestations, genetic characteristics, and prognosis in ALL and AML. Additionally, we compared the distinct features of FLT3 mutations between ALL and AML. In ALL patients, those with FLT3 mutations predominantly exhibited hyperdiploidy (48.6% vs. 14.9%, p < 0.001) and higher FLT3 expression (108.02 [85.11, 142.06] FPKM vs. 23.11 [9.16, 59.14] FPKM, p < 0.001), but lower expression of signaling pathway-related genes such as HRAS, PIK3R3, BAD, MAP2K2, MAPK3, and STAT5A compared to FLT3 wild-type patients. There was no significant difference in prognosis between the two groups. In contrast, AML patients with FLT3 mutations were primarily associated with leucocytosis (82.90 [47.05, 189.76] G/L vs. 20.36 [8.90, 55.39] G/L, p = 0.001), NUP98 rearrangements (30% vs. 4.8%, p = 0.018), elevated FLT3 expression (74.77 [54.31, 109.46] FPKM vs. 34.56 [20.98, 48.28] FPKM, p < 0.001), and upregulated signaling pathway genes including PIK3CB, AKT1, MTOR, BRAF, and MAPK1 relative to FLT3 wild-type, correlating with poor prognosis. Notably, internal tandem duplications were the predominant type of FLT3 mutation in AML (66.7%) with higher inserted base counts, whereas they were almost absent in ALL (6.3%, p < 0.001). In summary, our study demonstrated that the forms and impacts of FLT3 mutations in ALL differed significantly from those in AML. The gene expression profiles of FLT3-related pathways may provide a rationale for using FLT3 inhibitors in AML rather than ALL when FLT3 mutations are present.

Low-Dose Metformin and Profibrotic Signature in Central Centrifugal Cicatricial Alopecia

Central centrifugal cicatricial alopecia (CCCA) is a scarring alopecia predominantly affecting Black female individuals. Current conventional treatments target inflammation but not the underlying fibrotic processes, often leading to permanent hair loss. To investigate the associations of low-dose oral metformin, an antidiabetic medication with antifibrotic properties, with clinical symptoms and scalp gene expression patterns in patients with CCCA. This retrospective clinical case series and transcriptomic analysis included patients treated at a single tertiary academic medical center between January 2023 and March 2024. All patients had biopsy-confirmed CCCA refractory to standard treatments. Transcriptomic analysis was performed on patients with previously banked, paired scalp biopsies before and after treatment with adjuvant metformin for at least 6 weeks. Extended-release metformin, 500 mg, once daily was added to participants' baseline CCCA treatment regimens. Clinical assessments included pruritus, inflammation, scalp resistance, and hair regrowth. Gene expression profiling via bulk RNA sequencing analysis evaluated differential gene expression and pathway enrichment. A total of 12 Black female participants were included in the study, and transcriptomic analysis was performed in 4 participants. After at least 6 months of metformin treatment, 9 participants experienced improvement in disease, including scalp pain, inflammation, and/or pruritus, and 6 demonstrated clinical evidence of hair regrowth. The addition of metformin led to reversal of many prominent gene pathways previously identified in CCCA. Transcriptomic analysis revealed upregulation of pathways and genes (keratin-associated proteins [KRTAPs]) involved in keratinization, epidermis development, and the hair cycle (absolute log2-fold change > 4), with concomitant downregulation of fibrosis-related pathways and genes (eg, MMP7, COL6A1) (fold change >1.5; all false discovery rate <.05). Gene set analysis showed reduced expression of helper T cell 17 and epithelial-mesenchymal transition pathways and elevated adenosine monophosphate kinase signaling and KRTAPs after metformin treatment. In this case series of patients with treatment-refractory CCCA, low-dose oral metformin was associated with symptomatic improvement and dual modulation of gene expression, stimulating hair growth pathways while suppressing fibrosis and inflammation markers. These findings provide a rationale for future clinical trials studying metformin as a targeted therapy for CCCA and other cicatricial alopecias.

Delayed Reproduction, Injury, and Regeneration of Testes in Out-of-Season Breeding of Largemouth Bass (Micropterus nigricans).

Out-of-season breeding is an effective method for addressing seasonal shortages of fry in aquaculture species such as largemouth bass (LMB) for year-round production. Off-season breeding of LMB can be achieved by subjecting breeding LMB to prolonged low-temperature conditions; however, this can alter reproductive rhythms, affecting the quality of their sperm and leading to a decrease in reproductive efficiency. Therefore, it is crucial to investigate issues such as the damage to the testes and the related mechanisms caused by low-temperature stress during out-of-season breeding. In this experiment, we assessed the changes in the testes during this time in LMB by comparing reproductive rhythms, testicular histomorphology, ultrastructure, antioxidant capacity and apoptosis. We synthesized measurements of LMB from three identically treated cement ponds and fish exposed to water temperatures of 13-16 °C to assess the changes in the testes. The results showed that (1) out-of-season reproduction delayed sperm production and promoted sperm redevelopment in LMB, various hormone levels have changed over time (e.g., LH, FSH, and T). (2) The head plasma membrane of LMB spermatozoa was separated, and the middle mitochondria were swollen. (3) The expression levels of antioxidant enzymes (cat, sod, and gpx) were upregulated, and oxidative stress occurred in LMB. (4) The expression levels of apoptosis genes (e.g., bax, bcl2, and caspase3) were upregulated, and apoptosis occurred in LMB due to off-season breeding. Moreover, important genes of the mitochondrial apoptosis pathway (bid, CYT-C) were upregulated, indicating that spermatozoan apoptosis in LMB was probably achieved through the mitochondrial apoptosis pathway. These results suggest the delays, damage, and regeneration of LMB testes. Our findings provide new insights into the molecular mechanisms that trigger changes in sperm quality during out-of-season breeding in fish.

Development of a high-yield recombinant clone for the enhanced production of a long-acting Glucagon like peptide-1 analogue with improved biological efficacy

Glucagon-like peptide-1 (GLP-1) analogues have demonstrated greater efficacy and safety in treating type 2 diabetes mellitus (T2DM) due to their strong ability to regulate hypoglycemia. The short plasma half-life of native GLP-1 has prompted the development of novel strategies, such as the conjugation of a C-16 fatty acid to lysine in the GLP-1 analogue sequence, to enhance its longevity. With the escalating burden of T2DM, there is a pressing need for the development of innovative GLP-1 analogues with enhanced efficacy and increased production capacities. In this study, we engineered three recombinant DNA-based clones by incorporating distinct upstream fusion tags, enzyme cleavage sites, and charge-based additional amino acid sequences. These constructed plasmids were subsequently transformed into E. coli BL21 DE3 hosts to enhance solubility and streamline downstream protein purification and were evaluated for yield, purity, and biological effectiveness. A high-yield clone was selected, and the peptide was purified using column chromatography, yielding 150–180 mg per Liter of fermentation culture. The purified C-16 fatty acid-conjugated GLP-1 analogue peptide was analysed for its biological activity, particularly cAMP generation in pancreatic β-cells. Results revealed a concentration of 10.58 pmol/mL of generated cAMP and an average 1.5-fold up-regulation of genes in the cAMP downstream pathway compared to the innovator standard. Based on these findings, we conclude that the developed clone, featuring the enterokinase cleavage site with a sequence of modified TrxA tag with five additional hydrophobic amino acids, not only enhances yield but also preserves the biological efficacy of the final product.

Influence of exogenous 24-epibrassinolide on improving carotenoid content, antioxidant capacity and gene expression in germinated maize seeds.

Carotenoids have various physiological functions, such as immune regulation and cancer prevention. Germination could further improve the content of carotenoids in maize seeds. In this study, yellow maize seeds (Suyu 29) were soaked and germinated with different concentrations of 24-epibrassinolide. The changes of germination percentage, sprout length, bioactive components, antioxidant capacity and carotenoid content of the maize seeds were analyzed. Additionally, the relative expression of key genes in the carotenoid synthesis pathway was investigated. The results showed that the sprout length, germination percentage, soluble protein, free amino acids, proline, endogenous abscisic acid, vitamin C, total phenolics and carotenoids displayed a significant increasing trend compared with the control group (P < 0.05). The activity of superoxide dismutase and peroxidase increased by 55.1% and 58.5% versus the control group, and the antioxidant capacity of 2,2-diphenyl-1-picrylhydrazyl, 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) and ferric reducing antioxidant power was 19.8%, 13.4% and 44.1% higher than that of the control group (P < 0.05). Compared with the control group, the expression of genes was significantly up-regulated (P < 0.05). Under the treatment of 0.1 mg L-1 of 24-epibrassinolide, carotenoid content reached the highest value. The carotenoids showed a positive correspondence with antioxidant enzyme activity, antioxidant capacity and total phenolics content (P < 0.05). This study showed that 0.1 mg L-1 of exogenous 24-epibrassinolide promoted the accumulation of carotenoids and improved the antioxidant capacity and the quality of germinated maize seeds. It could provide a method for the development of germinated maize products enriched in carotenoids. © 2024 Society of Chemical Industry.

The Aqueous Extract of Hemerocallis citrina Baroni Improves the Lactation-Promoting Effect in Bovine Mammary Epithelial Cells through the PI3K-AKT Signaling Pathway.

Insufficient milk supply is a widespread issue faced by women globally and associated with a higher risk of health problems in infants and mothers. Hemerocallis citrina Baron, commonly known as daylily, is a perennial edible plant often used in traditional Asian cuisine to promote lactation. However, the active compound(s) and mechanism of its lactation-promoting effect remain unclear. This study aimed to confirm the traditional use of daylily in promoting lactation and investigate its potential active components and underlying molecular mechanisms. Our results showed that the aqueous extracts of H. citrina Baroni (HAE) significantly enhanced milk production, and the serum levels of lactation-related hormones, and promoted mammary gland development in lactating rats, as well as increased the levels of milk components in bovine mammary epithelial cells (BMECs) (p < 0.05). UHPLC-Q-Exactive Orbitrap-MS analysis revealed that hexamethylquercetin (HQ) is the representative flavonoid component in HAE, accounting for 42.66% of the total flavonoids. An integrated network pharmacology and molecular docking analysis suggested that HQ may be the potential active flavonoid in HAE that promotes lactation, possibly supporting lactation by binding to key target proteins such as STAT5A, PIK3CA, IGF1R, TP53, CCND1, BCL2, INS, AR, and DLD. Cell experiments further demonstrated that HQ could promote cell proliferation and the synthesis of milk proteins, lactose, and milk fat in BMECs. Transcriptomic analysis combined with a quantitative reverse transcription polymerase chain reaction (RT-qPCR) revealed that both HAE and HQ exert a lactation-promoting function mainly through regulating the expression of key genes in the PI3K-Akt signaling pathway.