Protein Gene Expression Research Articles

Theranostics: aptamer-assisted carbon nanotubes as MRI contrast and photothermal agent for breast cancer therapy

Breast cancer is one of the leading causes of death among women globally, making its diagnosis and treatment challenging. The use of nanotechnology for cancer diagnosis and treatment is an emerging area of research. To address this issue, multiwalled carbon nanotubes (MWCNTs) were ligand exchanged with butyric acid (BA) to gain hydrophilic character. The successful functionalization was confirmed by FTIR spectroscopy. Surface morphology changes were observed using SEM, while TEM confirmed the structural integrity of the MWCNTs after functionalization. Particle size, zeta potential, and UV spectroscopy were also performed to further characterize the nanoparticles. The breast cancer aptamer specific to Mucin-1 (MUC-1) was then conjugated with the functionalized MWCNTs. These MWCNTs successfully targeted breast cancer cells (MDA-MB-231) as examined by cellular uptake studies and exhibited a reduction in cancer-induced inflammation, as evidenced by gene transcription (qPCR) and protein expression (immunoblotting) levels. Immunoblot and confocal-based immunofluorescence assay (IFA) indicated the ability of CNTs to induce photothermal cell death of MDA-MB-231 cells. Upon imaging, cancer cells were effectively visualized due to the MWCNTs’ ability to act as magnetic resonance imaging (MRI) contrast agents. Additionally, MWCNTs demonstrated photothermal capabilities to eliminate bound cancer cells. Collectively, our findings pave the way for developing aptamer-labeled MWCNTs as viable “theranostic alternatives” for breast cancer treatment.Graphical abstract

Heat Shock Protein 70 Genes Are Involved in the Thermal Tolerance of Hippodamia variegata.

Previous studies have shown that the survival and reproduction of Hippodamia variegata are increasingly harmed by progressive increases in temperature (from 32 °C to 35 °C and 38 °C). In this study, transcriptome sequencing analysis was performed on H. variegata, after being exposed to different temperatures (from 32 to 38 °C) for 24 h, using high-throughput sequencing technology. We found the largest number of differentially expressed genes (DEGs) in the 35 °C vs. 32 °C group (1151) followed by the 38 °C vs. 32 °C group (1054) and then the 38 °C vs. 35 °C group (901), indicating that H. variegata expressed the largest number of newly mobilized genes under medium-high temperature (35 °C). Gene functional analysis showed that a large number of DEGs were involved in "Catalytic activity", "Oxidoreductase activity", "Metabolic pathways", and "Longevity regulating pathway-multiple species" gene groups. We randomly selected nine DEGs for validation using qRT-PCR. The results of qRT-PCR were consistent with the transcriptome data, confirming their reliability. Finally, the RNAi results showed that adult survival, longevity, and fecundity were lower in the group in which gene expression of the heat shock proteins (Hsp70-01 and Hsp68) was suppressed than in the control group (injection ds-GFP) at all the experimental temperatures (32, 35, and 38 °C). Our results indicate the important role of the heat shock proteins (Hsp70-01 and Hsp68) in resistance to high-temperature stress in H. variegata and provide a molecular basis for analyzing its thermotolerance mechanism.

UID-Dual Transcriptome Sequencing Analysis of the Molecular Interactions between Streptococcus agalactiae ATCC 27956 and Mammary Epithelial Cells.

Streptococcus agalactiae ATCC 27956 is a highly contagious Gram-positive bacterium that causes mastitis, has a high infectivity for mammary epithelial cells, and becomes challenging to treat. However, the molecular interactions between it and mammary epithelial cells remain poorly understood. This study analyzed differential gene expression in mammary epithelial cells with varying levels of S. agalactiae infection using UID-Dual transcriptome sequencing and bioinformatics tools. This study identified 211 differentially expressed mRNAs (DEmRNAs) and 452 differentially expressed lncRNAs (DElncRNAs) in host cells, primarily enriched in anti-inflammatory responses, immune responses, and cancer-related processes. Additionally, 854 pathogen differentially expressed mRNAs (pDEmRNAs) were identified, mainly enriched in protein metabolism, gene expression, and biosynthesis processes. Mammary epithelial cells activate pathways, such as the ERK1/2 pathway, to produce reactive oxygen species (ROS) to eliminate bacteria. The bacteria disrupt the host's innate immune mechanisms by interfering with the alternative splicing processes of mammary epithelial cells. Specifically, the bacterial genes of tsf, prfB, and infC can interfere with lncRNAs targeting RUNX1 and BCL2L11 in mammary epithelial cells, affecting the alternative splicing of target genes and altering normal molecular regulation.

Evaluation of the ameliorative potency of spirulina platensis against cerebellar damage induced by methotrexate in male rats: histopathological, ultrastructural, molecular, and biochemical studies

BackgroundMethotrexate (MTX), a drug utilized in cancer and rheumatoid arthritis treatment, is associated with acute and chronic neurodegenerative alterations. Spirulina platensis (SP) has several important phytochemical substances that act as free radical scavengers or natural antioxidants. The current study investigated the possible effects of the blue-green alga Spirulina platensis on cerebellar damage in male rats exposed to methotrexate. Forty (40) adult male albino rats were randomly divided into 4 groups (n = 10) and treated for one week: GI, the control group; GII was orally given 1000 mg SP/kg/daily, GIII was given a single intraperitoneal injection of MTX 75 mg/kg at the first day, and continued under the normal condition without other treatment till the end of the experiment, and GIV received both SP and MTX together with the same previous doses and duration. Neurobehavioral, histopathological, histochemical, immunohistochemical, ultrastructural, molecular, and biochemical data were recorded.ResultsMTX caused severe cerebellar degeneration in 3 cortical layers, especially the Purkinje layer. The Purkinje layer displayed a disrupted monolayer arrangement with pyknotic nuclei, a significant decrease in cell number, and shrunken cells surrounded by empty spaces. The molecular and granular layers are degenerated with elevated immunoreactions and gene expression of the glial fibrillary acidic protein (GFAP), ionized calcium-binding adapter molecule 1 (Iba-1), and neurofilament light chain antibody (NFL). Moreover, MTX significantly increased malondialdehyde (MDA) and myeloperoxidase (MPO) while decreasing the levels of reduced glutathione (GSH), serotonin, superoxide dismutase (SOD), acetylcholinesterase (ACHE), norepinephrine, and dopamine. These insults were noticeably mitigated by concomitant treatment with spirulina.ConclusionSpirulina improves neurological function by modulating the cerebellar damage elicited by MTX. This improvement may be attributed to the anti-inflammatory and antioxidant properties of spirulina.

Hyperlipidemia negatively impacts implantation by dysregulating tight junction and Claudin-3 and Claudin-4 expression in the endometrium

Clinical observational studies have suggested hyperlipidemia may disturb embryo implantation through endometrium; however, the mechanism has been unclear. With its profound implications for reproductive health, the present study aims to investigate whether hyperlipidemia affects endometrial epithelial cell tight junctions for implantation failures. By constructing hyperlipidemia mice model, the number and distribution of embryo implantation status were investigated after both natural mating and in vitro fertilization and embryo transfer (IVF-ET). Transmission electron microscopy (TEM) was used to compare the ultrastructure of tight junctions in endometrial endothelial cells. Western blot and immunofluorescence were used to explore the expression and localization of tight junction proteins, such as Claudin (CDLN)3, CLDN4, occludin (OCLN), and zonula occludens-1 (ZO1). For women with reproductive failure, mid-luteal phase endometrial tissues were collected, and gene expression of tight junction proteins was investigated using RNA sequencing and qRT-PCR. In hyperlipidemic mice, the number of embryo implantation sites significantly decreased with uneven distribution after natural mating and IVF-ET. Disrupted tight junctions were found, characterized by a decreased number of tight junctions by TEM, downregulated expressions of CDLN4, OCLN, and ZO1, and an increased expression of CLDN3 by western blot. In hyperlipidemic women with reproductive failure, the dysregulated expression of CLDN3 and CLDN4 was also present in the luteal phase endometrium. In this study, evaluation of both animal models and infertile women in vivo demonstrated that hyperlipidemia reduced female fertility, accompanied by disruption of tight junction structures and dysregulation of CLDN3 and CLDN4 expression in the endothelial cells of luteal phase endometrium.

Killing of xenogenous and virally infected homogenous target cells by shrimp lymphocyte-like haemocytes

Haemocytes play a crucial role in the invertebrate's immune system. In our lab, five subpopulations of shrimp haemocytes were identified in the past: hyalinocytes, granulocytes, semi-granulocytes and two subpopulations of non-phagocytic cells. In the latter two subpopulations, their characteristics such as having small cytoplasmic rims and not adhering to plastic cell-culture plates are very similar to those of mammalian lymphocytes. Therefore, they were designated lymphocyte-like haemocytes. Although little is known about their function, we hypothesize, based on their morphology, that they may have a cytotoxic activity like natural killer cells, with the ability to recognize and kill target cells. In our study, K562 cells and Sf9 cells were used as xenogenous target cells to detect the cytotoxic activity of the shrimp non-adherent lymphocyte-like haemocytes. Non-adherent haemocytes were collected and mixed with K562 cells and Sf9 cells at a 5:1 ratio and the binding activity was examined under a microscope. The binding rate of non-adherent haemocytes to K562 cells and Sf9 cells reached 6.6 % and 2.4 % after 240 min of culture, respectively. Then, the killing activity of non-adherent haemocytes was detected by an EMA staining (fluorescence microscopy), which showed 3.75 % dead K562 cells and 1.025 % dead Sf9 cells, and by Sytox® blue staining (flow cytometry), which showed 4.97 % of dead K562 cells. Next, a killing assay was developed to visualize the killing activity of shrimp non-adherent haemocytes. Non-adherent haemocytes were pre-labeled in blue (CellTracker blue) and K562/Sf9 cells in green (CFSE); dead cells were differentially stained red with ethidium bromide. The cytotoxic activity increased and reached a level of 2.59 % in K562 cells and 0.925 % in Sf9 cells at 120 min after co-culture. Furthermore, in the co-cultures of non-adherent haemocytes with K562 cells and Sf9 cells, upregulation of the gene and protein expression of the cytotoxic molecules torso-like protein and granzyme B was observed by RT-qPCR at 240 min and western blotting at 180 min. Additionally, non-adherent haemocytes were co-cultured with WSSV-inoculated shrimp ovary and lymphoid organ cells to detect the cytotoxicity to homogenous target cells. The binding activity started at 60 min in both the ovary and lymphoid organ cultures and reached at 240 min 50.62 % and 40.7 %, respectively. The killing activity was detected by EMA staining and the percentage of dead ovary and lymphoid organ cells increased respectively from 10.84 % to 6.89 % at 0 min to 13.09 % and 8.37 % at 240 min. In conclusion, we demonstrated the existence of cytotoxic activity of shrimp lymphocyte-like haemocytes against xenogenous cells from mammals and insects and against WSSV-infected homogenous shrimp cells.

Inheritance and ecological effects of exogenous genes from transgenic Brassica napus to Brassica juncea hybrids

●With the rapid development of new breeding techniques, the ecological impacts of transgenic plants receive wide concern again, particularly for potential gene flow from transgenic crops to their relatives. The transgene insertion position, number of gene copies, and flanking sequence of exogenous genes integrated into the recipient genome affect the genetic stability and fitness of offspring.●We employed hybrids F1 and F2, six backcross generations BC1-BC6 and BC1F1 from transgenic Brassica napus with Bacillus thuringiensis (Bt) cry1Ac gene and its wild relative B. juncea through hand pollination. We detected exogenous gene copies, mRNA transcription, and protein expression by ddPCR, qRT-PCR and ELISA, and the fitness of hybrid and backcross generations.●Exogenous genes followed Mendelian segregation expectations in hybrid and backcrossed generations, with stable gene copies, mRNA transcription and protein concentration of exogenous genes in offspring. Exogenous gene copies had no significant effects on plant fitness, but had positive effects on mRNA transcription and protein concentration that varied with growth stages in hybrid and backcross generations.●Our study demonstrated inheritance and ecological effects of exogenous genes from transgenic plants to wild relatives, which will help ecological risk management of biotechnological plants released in the nature.

The down-regulation of salivary protein gene expression by etofenprox partially contributed to reducing the risk of increased fecundity in the brown planthopper

Etofenprox is a pyrethroid insecticide that acts on the nervous system of insects. Due to its low toxicity to aquatic animals, it is permitted for use in controlling insect pests in rice fields. The brown planthopper (BPH), Nilaparvata lugens, a significant piercing-sucking pest feeding on rice exclusively, secretes various salivary components when feeding. Salivary proteins are essential for BPH feeding, but their response to etofenprox is not well understood. The application of etofenprox down-regulated the expression of 21 salivary protein genes, among which 9 genes (NlShpa, Salivap 3, CA, NlSEF1, Nl12, NlHSC70–3, NlSP1, NlG14, and NlDNAJB9) showed significant differences. Most differentially expressed genes are found important for BPH physiological processes, except Nl12. Here we found that silencing Nl12 impeded ovary development, thereby inhibiting oocyte formation. The potential explanation was that Nl12 was highly expressed in both salivary gland and ovary, and the ovary development abnormality may be due to the direct effect from expression reduction in ovary and/or indirect influence from expression reduction in salivary gland. Altogether, our findings provide a new insight into the mechanism of action of etofenprox on insect pests and explain part of the reason why etofenprox does not stimulate reproduction in BPH.

Abstract P378: Impaired Diurnal Control of Blood Pressure and Sodium Excretion in Aged Hypertensive Male Sprague Dawley Rats

Hypothesis: The diurnal rhythmicity of blood pressure (BP) and sodium excretion is important to blood pressure regulation and is strongly influenced by the circadian clock genes. We hypothesized that the diurnal control of BP and natriuresis is impaired in aged male Sprague Dawley (SD) rats and may be associated with altered kidney expression of circadian clock gene proteins. Methods: Radiotelemetry (to record mean arterial BP (MAP)) and metabolic balance (to assess sodium excretion) was conducted in 3-month-old (3M) and 16-month-old (16M) male SD rats. On a separate day, 3M and 16M rats received an acute 900 μEq NaCl load by oral gavage at ZT0 and sodium excretion was assessed. Total renal cortex expression of Per1 and Bmal1 was assessed by immunoblotting (N=4-6/group ± SD). Results: 16M rats exhibit significantly increased MAP and 24-h sodium retention vs. 3M rats [MAP (mmHg) 3M 102±4 vs. 16M 127±5, P<0.05; 24h Na retention (mEq/24h) 3M 0.3±0.1 vs. 16M 0.8±0.2, P<0.05). 16M rats exhibit an attenuation of the circadian MAP pattern compared to 3M rats. Aged rats have a significant decrease in the blood pressure rise during the dark (active) vs. 3M rats (Dark phase increase in MAP (mmHg) 3M +13.6±2.1 vs. 16M +4.8±1.9, P<0.05). 3M rats exhibit a classical diurnal pattern of 24-h sodium excretion with increased natriuresis during the dark (active) phase – a response that is attenuated in 16M rats that exhibit a significant reduction in the increase in natriuresis in the dark phase (Light phase UNaV (μEq/12h) 3M 380±48 vs. 16M 410±36, Dark phase natriuresis 3M 620±31 vs 16M 502±27, P<0.05). 16M rats had an impaired natriuretic response to an acute oral NaCl load in the first 12h post-gavage (UNaV 12h (μEq/12h) 3M 681±42 vs. 16M 243±39, P<0.05). In 16M old rats there was no change in renal Per1 and decreased renal Bmal1 protein (Bmal1 fold change 16M vs. 3M, 0.65±0.7, P<0.05). Conclusions: We observed impaired circadian control of blood pressure and renal sodium excretion in aged male SD rats. These changes in renal sodium handling may reflect endogenous decreases in renal Bmal1 levels with age and contribute to the observed hypertensive phenotype in aged rats.

The transcription factor RUNT-like regulates pupal cuticle development via promoting a pupal cuticle protein transcription.

Holometabolous insects undergo morphological remodeling from larvae to pupae and to adults with typical changes in the cuticle; however, the mechanism is unclear. Using the lepidopteran agricultural insect Helicoverpa armigera, cotton bollworm, as a model, we revealed that the transcription factor RUNT-like (encoded by Runt-like) regulates the development of the pupal cuticle via promoting a pupal cuticle protein gene (HaPcp) expression. The HaPcp was highly expressed in the epidermis and wing during metamorphosis and was found being involved in pupal cuticle development by RNA interference (RNAi) analysis in larvae. Runt-like was also strongly upregulated in the epidermis and wing during metamorphosis. Knockdown of Runt-like produced similar phenomena, a failure of abdomen yellow envelope and wing formation, to those following HaPcp knockdown. The insect molting hormone 20-hydroxyecdysonen (20E) upregulated HaPcp transcription via RUNT-like. 20E upregulated Runt-like transcription via nuclear receptor EcR and the transcription factor FOXO. Together, RUNT-like and HaPCP are involved in pupal cuticle development during metamorphosis under 20E regulation.

Combination treatment with hyaluronic acid synthesis and Bcl-2 inhibitors induces senolytic elimination of oral squamous cell carcinoma cells in vitro

ObjectiveHyaluronic acid (HA) is a major glycosaminoglycan in the extracellular matrix and plays a vital role in cancer progression. HA interacts with cell membrane receptors, such as receptor for HA-mediated motility (RHAMM), to regulate the cell cycle and cell division. 4-Methylumbelliferone (4-MU) is a well-known HA synthesis inhibitor and a promising cancer therapy alternative. 4-MU is a coumarin derivative with no obvious side effects and has been studied in several cancers. However, little is known about its use in oral squamous cell carcinoma (OSCC). In this study, we evaluated the antitumor effects of 4-MU on OSCC cells. MethodsThe effects of 4-MU on HAS gene and HA-related protein expression were evaluated with RT-qPCR and Western blotting. The effects on HA secretion was confirmed by ELISA. The effects on cell proliferation were confirmed by MTS assay. Trypan blue exclusion assay was used for cell viability assay. Cell cycle analysis, cell-clock cell cycle assay, and apoptosis assay were used to determine the effects on cell cycle and apoptosis. SA-β-gal staining was used to evaluate senescence. Results4-MU induced senescence change in OSCC cells by overexpression of the anti-apoptotic protein Bcl-2 and markedly suppressed their proliferation. Furthermore, following administration of Bcl-2 inhibitor, ABT-263 known as a senescent cell eliminator can specifically induce apoptosis and eliminate the senescent OSCC cells. ConclusionsThese findings highlight the potential use of 4-MU in OSCC chemotherapy as a senescence-inducing agent that not only potently suppresses OSCC cell proliferation but also enhances the efficacy of Bcl-2 inhibitor.

Review Article on Molecular Basis of Zinc and Copper Interactions in Cancer Physiology.

Various clinical manifestations associated with measurable abnormalities of Zn and Cu in serum and tissue were determined in Cancer-Patients (CP), and therefore, these two metals are drawing more and more attention presently than ever before. Cancer is a disease of uncontrolled-abnormal-cell-division with invasion-potential which was exhibited to occur due to dys-regulation/dys-homeostasis of fundamental-biological-pathways (FBP) including antioxidant-enzyme-defense-system, anti-inflammatory and immune-systems, and DNA-damage-repair-system in the human-body resulting in generation of chronic-oxidative-stress induced DNA-damage and gene-mutations, inflammation and compromised immune-system, tumor-induced increased angiogenesis, and inhibition of apoptosis processes. Zn and Cu were recognized to be the most crucial components of FBP and imbalance in Zn/Cu ratios in CP asserted to generate chronic toxicity in human body through various mechanisms including increased chronic oxidative stress linked compromised DNA integrity and gene mutations due to malfunctioning of DNA damage repair enzymes; increased angiogenesis process due to Zn- and Cu-binding proteins metallothionein and ceruloplasmin-induced enhanced expression of tumor growth factors; and elevation in inflammatory response which was further shown to down/upregulate gene expression of multiple Zn transporter proteins leading to dys-homeostasis of intracellular Zn concentrations, and it was determined to disturb the equilibrium between cell growth and division, proliferation, differentiation, and apoptosis processes which lead to cancer progression. Moreover, Zn was reported to affect matrix metalloproteinase activity and influence immune system cells to respond differently to different cytokines and enhance immune-suppressive effects accelerating the angiogenesis, invasion, and metastasis potential in cancer. Further, the most significant use of serum Cu/Zn ratio was recommended in clinical diagnosis, prognosis, tumor stage, patient survival, and cancer follow-up studies which need further investigations to elucidate and explore their roles in cancer physiology for clinical perspective.

Mendelian randomization reveals potential causal relationships between cellular senescence-related genes and multiple cancer risks

Cellular senescence is widely acknowledged as having strong associations with cancer. However, the intricate relationships between cellular senescence-related (CSR) genes and cancer risk remain poorly explored, with insights on causality remaining elusive. In this study, Mendelian Randomization (MR) analyses were used to draw causal inferences from 866 CSR genes as exposures and summary statistics for 18 common cancers as outcomes. We focused on genetic variants affecting gene expression, DNA methylation, and protein expression quantitative trait loci (cis-eQTL, cis-mQTL, and cis-pQTL, respectively), which were strongly linked to CSR genes alterations. Variants were selected as instrumental variables (IVs) and analyzed for causality with cancer using both summary-data-based MR (SMR) and two-sample MR (TSMR) approaches. Bayesian colocalization was used to unravel potential regulatory mechanisms underpinning risk variants in cancer, and further validate the robustness of MR results. We identified five CSR genes (CNOT6, DNMT3B, MAP2K1, TBPL1, and SREBF1), 18 DNA methylation genes, and LAYN protein expression which were all causally associated with different cancer types. Beyond causality, a comprehensive analysis of gene function, pathways, and druggability values was also conducted. These findings provide a robust foundation for unravelling CSR genes molecular mechanisms and promoting clinical drug development for cancer.

Therapeutic Potential of PLK1, KIF4A, CDCA5, UBE2C, CDT1, SKA3, AURKB, and PTTG1 Genes in Triple-Negative Breast Cancer: Correlating Their Expression with Sensitivity to GSK 461364 and IKK 16 Drugs.

The treatment of triple-negative breast cancer (TNBC) has been associated with challenges due to the lack of expression of ER, PR, and HER2 receptors in tumor cells. This study aimed to identify genes with potential therapeutic targets in TNBC. Data from the cancer genome atlas regarding breast cancer (BC) were downloaded. After initial preprocessing, cancer samples were categorized into four groups: TNBC, HER2-positive, luminal A, and luminal B. Gene expression differences between these groups were calculated, focusing on genes that showed differential expression in TNBC. A protein-protein interaction network was conducted to identify hub genes among the candidate genes related to TNBC. The protein expression of candidate genes was assessed using immunohistochemistry data from the human protein atlas. Drug resistance and sensitivity associated with hub genes were identified using data from PharmacoDB. TNBC samples and the RT-qPCR method were used to confirm the results. Our findings revealed that eight genes, namely PLK1, KIF4A, CDCA5, UBE2C, CDT1, SKA3, AURKB, and PTTG1, had significant upregulation at the RNA level in TNBC subgroup compared to other subgroups and could be considered hub genes in TNBC. Compared to other subgroups, their expression level in TNBC samples had high sensitivity and specificity. RT-qPCR results also demonstrated a significant increase in levels of SKA3 and PTTG1 in the TNBC compared to healthy tissue and other subgroups. The protein expression of these genes was notably high in some BC samples. PharmacoDB data showed that some candidate genes were closely linked to drug sensitivity of GSK 461364 and IKK 16. The results of this study showed a significant increase in the expression level of PLK1, KIF4A, CDCA5, UBE2C, CDT1, SKA3, AURKB, and PTTG1 in TNBC compared to other BC subgroups. These genes show considerable promise as therapeutic targets for the TNBC subgroup.

Comparison of selenium-mediated regulation of heat shock protein and inflammation in-vitro and in-ovo for heat resistance enhancement in broiler

Selenium is a heat-stress-reducing substance that improves heat resistance and is being studied for its effective application in the broiler industry. However, research on feed additives is labor-intensive and time-consuming because of the need for feeding experiments. We aimed to compare the effects of selenium under heat stress in vitro and in ovo, specifically examining the gene expression of heat shock proteins (HSP) and inflammatory markers. Two groups were included in the in-vitro study: in-vitro control (TC; selenium 0 μg/ml) and in-vitro selenium (TS; selenium 5 μg/ml). The satellite cells were cultured at 42°C for 48 h after selenium treatment. The in-ovo study comprised 4 groups: in-ovo control and in-ovo selenium 1-3 (OC, OS1, OS2, and OS3; selenium 2.5, 5, and 10 μg/egg, respectively). Selenium was injected on the 18th day after hatching, and heat treatment was performed at 32-34°C from the 14th to the 21st day after hatching, and the leg muscles of the chicks were collected on the 21st day. The gene expression of heat shock proteins (HSP), caspase3, nuclear factor kappa light-chain enhancer of activated B cells (NF-kB), and IL-8 was analyzed in in-vitro and in-ovo experiments, respectively. In-vitro results showed significant increases in HSP90, HSP60, and HSP40 in TS compared to TC, with a significant decrease in HSP70. In the in-ovo study, HSP70, caspase3, NF-kB and IL-8 were significantly increased in OS1. HSP90, HSP60, HSP40, HSP27 and NF-kB were significantly decreased in in-ovo OS2 compared to in-vitro TS, implying a trend in ratio compared to control. Selenium appeared to enhance heat resistance in-vitro and in-ovo by modulating HSPs and inflammation. However, differences in mRNA expression were observed depending on the concentration of selenium. These findings suggest that selenium modulates heat resistance through different mechanisms in-vitro and in-ovo, likely due to the complexity of whole-organism interactions in-ovo compared to the single-cell-type environment in-vitro. Therefore, to directly apply in-vitro results to in-ovo, a concentration comparison study for each additive is necessary.

Peanut allergen characterization and allergenicity throughout development.

Peanut allergy (PA) in children is a major concern. There is a need for better biological material for both diagnosis and oral immunotherapy (OIT) treatments. The unique state of seeds at early reproductive stages may affect the allergenicity of storage proteins, and impact clinical diagnostic and OIT protocols. The objective of this study was to evaluate the major allergen content in sequential seed developmental stages and monitor allergenicity via specific IgE binding quantification and skin prick testing. Seeds were collected from peanut plants and sorted into five developmental stages: initial (S1), developing (S2), full-size without coloration (S3), full-size with coloration (S4), and fully mature (S5) seeds. Samples were characterized by RNA-Seq, ELISA, and immunohistochemistry. Lyophilized, ground preparations were used for evaluation of skin test responses in sixty challenge-proven PA children. Gene expression, protein content, and specific IgE binding of allergenic proteins increased throughout seed maturation and development. An expression bias towards the less allergenic A-genome copy of the major allergen Ara h 2 was found in earlier stages, especially in stage S2. Immunohistochemical staining showed that Ara h 2 is more dispersed in the cell and less accumulated within organized bodies at stage S2 versus stage S4. Significant differences were found in mean wheal responses between the commercial peanut extract (equivalent to stage S5) and stages S1 and S2, but not with stage S4, upon skin prick testing in subjects with PA. The observed decrease in peanut-specific IgE binding of immature peanut seeds may be a result not only of decreased amounts of allergenic proteins, but also of profound changes in seed composition and conformation. This may be significant for developing a safer and more effective peanut OIT protocol.

Expanding the repertoire of Antibody Drug Conjugate (ADC) targets with improved tumor selectivity and range of potent payloads through in-silico analysis.

Antibody-Drug Conjugates (ADCs) have emerged as a promising class of targeted cancer therapeutics. Further refinements are essential to unlock their full potential, which is currently limited by a lack of validated targets and payloads. Essential aspects of developing effective ADCs involve the identification of surface antigens, ideally distinguishing target tumor cells from healthy types, uniformly expressed, accompanied by a high potency payload capable of selective targeting. In this study, we integrated transcriptomics, proteomics, immunohistochemistry and cell surface membrane datasets from Human Protein Atlas, Xenabrowser and Gene Expression Omnibus utilizing Lantern Pharma's proprietary AI platform Response Algorithm for Drug positioning and Rescue (RADR®). We used this in combination with evidence based filtering to identify ADC targets with improved tumor selectivity. Our analysis identified a set of 82 targets and a total of 290 target indication combinations for effective tumor targeting. We evaluated the impact of tumor mutations on target expression levels by querying 416 genes in the TCGA mutation database against 22 tumor subtypes. Additionally, we assembled a catalog of compounds to identify potential payloads using the NCI-Developmental Therapeutics Program. Our payload mining strategy classified 729 compounds into three subclasses based on GI50 values spanning from pM to 10 nM range, in combination with sensitivity patterns across 9 different cancer indications. Our results identified a diverse range of both targets and payloads, that can serve to facilitate multiple choices for precise ADC targeting. We propose an initial approach to identify suitable target-indication-payload combinations, serving as a valuable starting point for development of future ADC candidates.

Identification of differentially expressed genes and screening for key genes involved in ovarian cancer prognosis: An integrated bioinformatics and network analysis approach

Objectives: Ovaries are important and essential organs of animals in producing and releasing eggs. Ovarian cancer (OvCa) is one of the most prevalent lethal gynecological malignancies with a lack of distinct biomarkers. Advances in high-throughput genomic data and the continued refinement of bioinformatics tools enable the identification of potential biomarkers. Leveraging these insights, we can employ systems biology approaches to enhance the accuracy of diagnosis and prognosis. Material and Methods: A comparative analysis was conducted between normal and tumor samples, employing bioinformatics software and tools. Differential expression analysis utilized fold-change statistics, while DAVID 6.8 software was used to perform gene ontology enrichment analysis and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. The protein-protein interaction (PPI) network was constructed differentially expressed genes (DEGs) using Search Tool for the Retrieval of Interacting Genes database, and Cytoscape 3.9.1, along with its Molecular Complex Detection and CytoHubba plugins, facilitated network visualization, analysis, and module detection. Hub gene expression and overall survival were explored through the Kaplan–Meier plotter, while Gene Expression Profiling Interactive Analysis 2 analyzed the tumor stage of OvCa patients. Hub genes protein expression was analyzed using the human protein atlas database through immunostaining results. The NetworkAnalyst program and Cytoscape were employed to analyze and visualize the transcription factor-hub gene associations. Subsequently, single-nucleotide variation, methylation, and pathway activity of hub genes were examined. Validation of hub genes messenger RNA expression was done using quantitative real-time polymerase chain reaction analysis. Results: 607 DEGs, including 248 upregulated and 359 downregulated genes, were identified. The top 20 candidate genes were screened out through PPI network analysis. We discovered that the genes BUB1 Mitotic Checkpoint Serine/Threonine Kinase B (BUB1B), Cyclin A2 (CCNA2), Mitotic Arrest Deficient 2 Like 1 (MAD2L1), Protein Regulator of Cytokinesis 1 (PRC1), Thyroid Hormone Receptor Interactor 13 (TRIP13), and ZW10 Interacting Kinetochore Protein (ZWINT) exhibited significant importance in OvCa prognosis. Conclusion: Six genes, BUB1B, CCNA2, MAD2L1, PRC1, TRIP13, and ZWINT (identified as functional hub genes), are probably playing tumor-promotive roles, except TRIP13. All genes product is functionally related to the cell cycle. These can be targeted in quest of potential therapeutics for OvCa treatment.

Regulation of arbuscular mycorrhizal fungi in citrus root hairs mediated by auxin efflux carrier protein PtPINs

Arbuscular mycorrhizal fungi (AMF) and auxins are critical in the development of root hairs, but the interaction mechanism by which they jointly regulating the development of citrus needs further clarification. This study investigated the influence of Funneliformis mosseae on the growth, root architecture, root hair traits, indole-3-acetic acid (IAA) content and flow rate, and auxins efflux carrier protein genes (PtPINs) expression in potted trifoliate orange (Poncirus trifoliata L.) over 4, 6, 8, 10, and 12 weeks of inoculation. Results indicated continuous increases in mycorrhizal colonization and soil hyphae length over time. Compared with the non-AMF, AMF significantly enhanced plant growth and biomass, and notably increased total root length, projected area, surface area, volume, and the number of 2nd-order lateral roots, with these parameters improving further over the treatment duration. Whereas root hair growth was inhibited at 4 w, 6 w and 8 w, it was promoted at 10 w and 12 w following AMF treatment. AMF also significantly increased the content of indole-3-acetic acid (IAA) over time. And the expression patterns of auxins flow rate and PtPINs vary across different treatment periods, showing a strong correlation with citrus growth and root hair development. Specifically, PtPINs were significantly correlated with total IAA flow rate and total root hair growth, with PtPIN6 potentially playing a positive role in root hair development. These findings suggested that auxins are involved in AMF-regulated citrus root hair growth, with PtPIN6 identified as a potentially key gene in auxin transport regulation.

XueBiJing injection improves the symptoms of sepsis-induced acute lung injury by mitigating oxidative stress and ferroptosis

Ethnopharmacological relevanceXBJ injection is approved by the China Food and Drug Administration for the adjunctive treatment of sepsis, and it is derived from the traditional Chinese medicine (TCM) prescription XuefuZhuyu Decoction. It consists of five Chinese herbal extracts: Carthamus tinctorius, Paeonia lactiflora, Salvia miltiorrhiza, Conioselinum anthriscoides ‘Chuanxiong’ and Angelica sinensis. Aim of the studyThe purpose of this study was to explore the relationship between ferroptosis and acute septic lung injury, and to evaluate the improvement effect of XBJ injection on acute lung injury in sepsis. Materials and methodsAcute lung injury was induced in rats by cecum ligation and puncture, and these rats were treated with XBJ injection. Oxidative stress and inflammation levels were assessed in serum and lung tissue, and tissue samples were collected for histological and protein analyses. To illustrate the mechanism of the improvement effect of XBJ on acute lung injury in sepsis, serum lipidomics was carried out to investigate whether XBJ prevents oxidative stress-induced lipid metabolism disorders. Furthermore, protein expression of ferroptosis-related genes was also examined. ResultsXBJ was shown to be effective in alleviating sepsis-induced ALI. XBJ also improves sepsis-induced acute lung injury by reducing lipid peroxidation and inflammation and modulating ferroptosis pathways. Specifically, compared with the sham group, XBJ downregulated the levels of Fe2+, MDA and GSSG, and reversed the decrease in the levels of GSH and GSH/GSSH in lung tissue. Metabolic pathways such as glycerophospholipid metabolism, phospholipid metabolism, and lipid metabolism associated with ferroptosis were obtained by lipidomic analysis of differential lipid metabolite enrichment, suggesting that ferroptosis occurs in septic rats, and that XBJ inhibits ferroptosis and thereby improves sepsis-induced ALI. Furthermore, XBJ optimises iron metabolism and lipid oxide metabolism by regulating the expression of a series of proteins that are closely related to ferroptosis, such as GPX4, ACSL4, x-CT, and FTH1. ConclusionsOur findings, initially, indicated that XBJ ameliorates sepsis-induced ALI by reducing oxidative stress and ferroptosis, revealing a previously unrecognised mechanism by which XBJ ameliorates sepsis-induced ALI.