Factor Activity Research Articles

Abstract C121: Racial differences in expression of the oxygen-sensor BACH1 in breast cancer

Abstract Hypoxia is an important hallmark of aggressive solid tumors and a key driver of metastasis and resistance to therapy. Hypoxic stress induces the activation of various factors, such as hypoxia- inducible factors (HIFs), which facilitate cellular adaptation and promote tumorigenesis. Given the complexity of this mechanism, identifying novel targets that sensitize hypoxic tumors to therapy could have significant clinical impact. We demonstrate HIF-independent induction of BACH1 within multiple TNBC (Triple-Negative Breast Cancer) cell lines and patient-derived organoids under hypoxic conditions. Utilizing ChIP-seq, bulk, single-cell and spatial transcriptomics across diverse BACH1 knockout tumor models, our study revealed that BACH1 selectively and directly modulates the transcriptional response associated with hypoxia and stress in TNBC. TGCA analysis reveals racial disparities in BACH1 mRNA expression in TNBC BACH1 mRNA and activity across all breast cancer subtypes (Basal, Her, LumA, LumB). Our findings suggest that BACH1 is an oxygen-sensitive mediator of hypoxia and a promising therapeutic target for mitigating hypoxia-induced pro-metastatic signaling and treatment resistance. These results also highlight the necessity of accounting for racial differences in BACH1 expression and function as a means of developing targeted therapy that takes into account racial disparities in TNBC. Citation Format: Long C. Nguyen, Christopher Dann, Dongbo Yang, Madeline Henn, Emily Shi, Thomas Li, Kent Schechter, Letícia Stock, Wenchao Liu, Margarite Matossian, Andrea Valdespino, Yoo Jane Han, Jing Zhang, Geetha Priyanka Yerradoddi, Yan Li, Mitsuyo Matsumoto, Olufunmilayo I Olopade, Kazuhiko Igarashi, Marsha R Rosner. Racial differences in expression of the oxygen-sensor BACH1 in breast cancer [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr C121.

Abstract C089: Characterization of the breast tumor immune microenvironment in women of African descent using single-nucleus RNA- and ATAC-sequencing

Abstract Women of African descent are at an increased risk of developing and dying from aggressive subtypes of breast cancer. A connection between aggressive disease and Western Sub-Saharan African ancestry has been postulated, but it remains largely unknown to what extent breast cancer in Africa is reminiscent of breast cancer in U.S. African American (AA) women who experience disproportionately high mortality rates. We performed ATAC- and RNA-sequencing on 9 human triple-negative breast cancer cell lines of U.S. origin and discovered that African ancestry influences the chromatin landscape, leading to disparate transcription factor (TF) activity and downstream gene expression patterns indicative of an aggressive tumor biology. Here, we describe an ambitious study that employs single-nucleus (sn) ATAC- and RNA-sequencing (snMultiome) of frozen breast tumors to characterize chromatin accessibility and gene expression patterns with single-cell resolution in AA (n=33), Kenyan (n=25), and European American (EA, n=24) women in relation to genetic ancestry, risk factor exposures, clinical characteristics, and 5-year survival. To achieve this, we successfully isolated intact, high-quality single nuclei from archival frozen breast tumor tissue through an optimized combination of enzymatic digestion and automated tissue homogenization. We performed snMultiome sequencing of 82 tumors using the 10x Genomics platform. Following filtering, normalization (SCT for snRNA; LSI for snATAC), peak calling (MACS2), and integration (Harmony), our dataset includes a total of 296,557 nuclei. Cancerous (163,419 nuclei) and non-cancerous (133,138 nuclei) cells were distinguished based on DNA copy number (CopyKat). Within the microenvironment, 11 major immune, epithelial, and stromal cell types were successfully annotated, exhibiting distinct patterns by population group (e.g. AA tumors showed markedly increased abundance of myeloid and T-cells, while Kenyan tumors showed increased abundance of pericytes and fibroblasts, relative to EA). A large number of enriched TFs within each cell type varied significantly by population group, suggesting distinct chromatin accessibility patterns related to genetic ancestry. CD45+, EPCAM- cells were further extracted, clustered, and manually annotated for subpopulations based on known marker genes. We detected 26 distinct immune subpopulations across our samples, including 2 inflammatory macrophage, 6 immunosuppressive macrophage, 1 fibronectin+/thrombospondin+ monocyte, 6 T-cell, 1 natural killer cell, 2 B-cell, 1 mast cell, and 4 dendritic cell subpopulations. Differential gene expression analyses of each immune cell subpopulation, adjusted for age, BMI, and tumor subtype, revealed ancestry-specific gene expression patterns, which could provide novel insights into functional differences that may impact tumor immune response. Current efforts focus on in-depth molecular characterization of ancestry-related differences in the tumor immune environment and distinct signatures present in lethal disease. Citation Format: Alexandra R. Harris, Huaitian Liu, Brittany D. Jenkins, Tiffany H. Dorsey, Francis Makokha, Shahin Sayed, Gretchen Gierach, Stefan Ambs. Characterization of the breast tumor immune microenvironment in women of African descent using single-nucleus RNA- and ATAC-sequencing [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr C089.

Transcriptomic and Phenotypical Analysis of the Physiological Plasticity of Chamaecyparis hodginsii Roots under Different Nutrient Environments and Adjacent Plant Competition.

Chamaecyparis hodginsii seedlings undergo significant changes during growth due to different nutrient environments and adjacent plant competition, which is evident in the physiological plasticity changes in their roots. Therefore, in this experiment, 20 one-year-old elite C. hodginsii family seedlings were selected as the test objects, and the different nutrient environments and adjacent plant competition environments in nature were artificially simulated. Four nutrient environments (N heterogeneous nutrient environment, P heterogeneous nutrient environment, K heterogeneous nutrient environment, and homogeneous environment) and three planting patterns (single plant, conspecific neighbor, and heterospecific neighbor) were set up to determine the differences in root physiological indexes and plasticity of different family seedlings, and the families and treatment combinations with higher comprehensive evaluation were selected. The transcriptome sequencing of fine roots of C. hodginsii under different treatments was performed to analyze the differentially expressed genes. The results showed that the root activity, antioxidant enzyme activity, and nutrient element content of C. hodginsii seedlings in the N and P heterogeneous environments were higher than those in the homogeneous nutrient environment, while there was no significant difference between the K heterogeneous nutrient environment and the homogeneous environment, but MDA content was higher than that in other nutrient environments. The root activity and antioxidant enzyme activity in the competitive patterns were generally higher than those in the single plant and reached the peak in the heterospecific neighbor. The root physiological plasticity index of line 490 was the highest, but the comprehensive evaluation of root physiological indexes of lines 539 and 535 was better. The pattern with the highest comprehensive evaluation score was P heterogeneous nutrient environment × heterospecific neighbor. The effects of the N and P heterogeneous nutrient environments on root transcriptome genes were similar, which significantly increased DNA transcription and regulatory factor activity, while K heterogeneous nutrient environment focused on the regulation of root enzyme activity. The heterogeneous nutrient environment induces the conduction of hormone signals in the roots of C. hodginsii and induces the synthesis of phenylpropanone. The biosynthesis of phenylpropanone in the roots of C. hodginsii will increase significantly under competitive patterns. In summary, the N and P heterogeneous nutrient environments and the heterospecific neighbor can improve the root physiological indexes of C. hodginsii families, and the root physiological indexes of lines 539 and 535 are the best. The nutrient environment and competition pattern mainly affect the root system to transmit hormone signals to regulate enzyme activity.

Transcriptome analysis of inflorescence embryogenesis in Festuca Glauca

In this study, RNA-seq was employed for transcriptome sequencing at four developmental stages of normal (L) and embryogenic (X) Festuca glauca ‘Elijah Blue’ inflorescences to analyze and identify the metabolic pathways and regulatory genes associated with inflorescence embryogenesis, thereby facilitating the understanding of the molecular mechanisms of inflorescence embryogenesis in Festuca glauca. The results revealed a total of 50,733 differentially expressed genes (DEGs) between the control (L) and embryogenic (X) samples at different developmental stages. Among them, 19,640 (38.71 %) were upregulated and 31,093 (61.29 %) were downregulated. A total of 2585 DEGs were expressed in both stage 1 (L1-vs-X1) and stage 4 (L4-vs-X4). Gene Ontology (GO) analysis revealed that the DEGs in these stages were mainly enriched in processes related to photosynthetic membranes, chloroplasts, activity of DNA-binding transcription factors, components of ribosomal structure, reactions involving oxidized compounds, and photosynthesis; while Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the DEGs in these stages were mainly enriched in pathways such as plant-pathogen interactions, plant hormone signal transduction, phenylpropanoid biosynthesis, ribosomes, and galactose metabolism. The top families containing differentially expressed transcription factors (DETFs) in these stages included ERF, bHLH, MYB-related, NAC, and WRKY. A total of 39 and 29 DETFs associated with embryogenesis were identified in the L1-vs-X1 and L4-vs-X4 stages, respectively. Additionally, 79 and 110 embryogenesis-related genes were identified in the plant hormone signal transduction metabolic pathway in the L1-vs-X1 and L4-vs-X4 stages, respectively.

The mitochondrial Na+/Ca2+ exchanger NCLX is implied in the activation of hypoxia-inducible factors

Eukaryotic cells and organisms depend on oxygen for basic living functions, and they display a panoply of adaptations to situations in which oxygen availability is diminished (hypoxia). A number of these responses in animals are mediated by changes in gene expression programs directed by hypoxia-inducible factors (HIFs), whose main mechanism of stabilization and functional activation in response to decreased cytosolic oxygen concentration was elucidated two decades ago. Human acute responses to hypoxia have been known for decades, although their precise molecular mechanism for oxygen sensing is not fully understood. It is already known that a redox component, linked with reactive oxygen species (ROS) production of mitochondrial origin, is implied in these responses. We have recently described a mechanism by which the mitochondrial sodium/calcium exchanger, NCLX, participates in mitochondrial electron transport chain regulation and ROS production in response to acute hypoxia.Here we show that NCLX is also implied in the response to hypoxia mediated by the HIFs. By using a NCLX inhibitor and interference RNA we show that NCLX activity is necessary for HIF-α subunits stabilization in hypoxia and for HIF-1-dependent transcriptional activity. We also show that hypoxic mitochondrial ROS production is not required for HIF-1α stabilization under all circumstances, suggesting that the basal cytosolic redox state or other mechanism(s) could be operating in the NCLX-mediated response to hypoxia that operates through HIF-α stabilization. This finding provides a link between acute and medium-term responses to hypoxia, reinforcing a central role of mitochondrial cell signalling in the response to hypoxia.

Moringa oleifera hydroalcoholic leaf extracts mitigate valproate-induced oxidative status in the extraorbital lacrimal gland in a rat model

Dysfunction of the extraorbital lacrimal gland (ELG) can lead to loss of vision due to damage to the epithelium of cornea. The broad-spectrum anti-epileptic drug sodium valproate (SV) has numerous side effects. Moringa oleifera (M.oleifera) is widely used as a food and in folk medicine. The effects of orally administered SV and M. oleifera hydroalcoholic leaf extract on rat ELG were investigated in this study by analysing both antioxidant and oxidant parameters. Additionally, boron level and tissue factor (TF) activity were determined. Protein changes were detected by sodium dodecyl sulfate gel electrophoresis (SDS-PAGE). Significantly lower values of glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and total antioxidant status (TAS) were observed in the SV group compared to the control group. Treatment with Moringa extract significantly increased SOD, CAT and TAS values in the Moringa given SV group (SVM). While no significant differences were observed between the sialic acid values of the groups, lipid peroxidation (LPO), nitric oxide (NO) and total oxidant status (TOS) values were significantly elevated in the SV group compared to the control group. Due to the effect of Moringa extract, LPO, NO and TOS levels were significantly decreased in the SVM group compared to the SV group. TF activity was not meaningfully altered between groups. Compared to control rats, oxidative stress index (OSI) level significantly increased, whereas the boron level decreased in the SV group. Moringa extract treatment noticeably reduced OSI in the SVM group. According to SDS-PAGE, decreases in the density of protein bands with molecular weights of 51, 83, and 90 kDa were observed in SV given rats compared to the other groups. These decreases were reversed by the administration of Moringa extract. Moringa extract has shown protective properties arising from antioxidant potential, especially with its very low OSI value. Individuals undergoing SV treatment and having ELG complications might consider using Moringa extract to mitigate valproate induced damage.

Spatial and signaling overlap of growth factor receptor systems at clathrin-coated sites.

Cellular communication is regulated at the plasma membrane by the interactions of receptor, adhesion, signaling, exocytic,and endocytic proteins. Yet, the composition and control of these complexes in response to external cues remain unclear. We use high-resolution and high-throughput fluorescence imaging to map the localization of growth factor receptors and related proteins at single clathrin-coated structures in human squamous HSC3 cells. We find distinct protein signatures between control cells and cells stimulated with growth factors. Clathrin sites at the plasma membrane are preloaded with some receptors but not others. Stimulation with epidermal growth factor induces capture and concentration of epidermal growth factor, fibroblast growth factor1, and low-density lipoprotein receptor (EGFR, FGFR1, and LDLR). Regulatory proteins including ubiquitin ligase Cbl, the scaffold Grb2, and the mechanoenzyme dynamin2 are also recruited. Disrupting FGFR1 or EGFR activity with drugs prevents the recruitment of both EGFR and FGFR1. EGF was able to activate FGFR1 phosphorylation. Our data reveal novel coclustering and activation of receptors and regulatory factors at clathrin-coated sites in response to stimulation by a single growth factor, EGF or FGF. This behavior integrates growth factor signaling and allows for complex responses to extracellular cues and drugs at the plasma membrane of human cells.

Pharmacological restriction of genomic binding sites redirects PU.1 pioneer transcription factor activity

Transcription factor (TF) DNA-binding dynamics govern cell fate and identity. However, our ability to pharmacologically control TF localization is limited. Here we leverage chemically driven binding site restriction leading to robust and DNA-sequence-specific redistribution of PU.1, a pioneer TF pertinent to many hematopoietic malignancies. Through an innovative technique, ‘CLICK-on-CUT&Tag’, we characterize the hierarchy of de novo PU.1 motifs, predicting occupancy in the PU.1 cistrome under binding site restriction. Temporal and single-molecule studies of binding site restriction uncover the pioneering dynamics of native PU.1 and identify the paradoxical activation of an alternate target gene set driven by PU.1 localization to second-tier binding sites. These transcriptional changes were corroborated by genetic blockade and site-specific reporter assays. Binding site restriction and subsequent PU.1 network rewiring causes primary human leukemia cells to differentiate. In summary, pharmacologically induced TF redistribution can be harnessed to govern TF localization, actuate alternate gene networks and direct cell fate.

From single-cell to spatial transcriptomics: decoding the glioma stem cell niche and its clinical implications.

Gliomas are aggressive brain tumors associated with a poor prognosis. Cancer stem cells (CSCs) play a significant role in tumor recurrence and resistance to therapy. This study aimed to identify and characterize glioma stem cells (GSCs), analyze their interactions with various cell types, and develop a prognostic signature. Single-cell RNA sequencing data from 44 primary glioma samples were analyzed to identify GSC populations. Spatial transcriptomics and gene regulatory network analyses were performed to investigate GSC localization and transcription factor activity. CellChat analysis was conducted to infer cell-cell communication patterns. A GSC signature (GSCS) was developed using machine learning algorithms applied to bulk RNA sequencing data from multiple cohorts. In vitro and in vivo experiments were conducted to validate the role of TUBA1C, a key gene within the signature. A distinct GSC population was identified, characterized by high proliferative potential and an enrichment of E2F1, E2F2, E2F7, and BRCA1 regulons. GSCs exhibited spatial proximity to myeloid-derived suppressor cells (MDSCs). CellChat analysis revealed an active MIF signaling pathway between GSCs and MDSCs. A 26-gene GSCS demonstrated superior performance compared to existing prognostic models. Knockdown of TUBA1C significantly inhibited glioma cell migration, and invasion in vitro, and reduced tumor growth in vivo. This study offers a comprehensive characterization of GSCs and their interactions with MDSCs, while presenting a robust GSCS. The findings offer new insights into glioma biology and identify potential therapeutic targets, particularly TUBA1C, aimed at improving patient outcomes.

The effect of intravenous calcium gluconate on the prevention of ovarian hyperstimulation syndrome. (A randomized clinical trial)

ObjectiveOral Cabergoline and intravenous Calcium have the potential to prevent Ovarian Hyperstimulation Syndrome (OHSS) in assisted reproductive technology by regulating the activity of the Vascular Endothelial Growth Factor (VEGF) receptor. The purpose of this study was to examine the effect of Cabergoline with intravenous Calcium versus oral Cabergoline alone on the overall rate of OHSS. MethodsThis study is a randomized clinical trial which was carried out in Milad Infertility Center affiliated with Mashhad University of Medical Sciences, Mashhad, Iran between April 2016 and January 2018. A total of 192 patients were randomly assigned into two groups. The control group received oral Cabergoline and the intervention group received Calcium gluconate in addition to Cabergoline. A total rate of OHSS, moderate and severe OHSS were measured in both groups. ResultsThe demographic characteristics of the participants and the types of drugs used showed homogeneity between the intervention and control groups (P > 0.05). Furthermore, there was no significant difference between the two groups in terms of the number of the follicle, oocytes obtained, metaphase II oocytes, the number of embryos, and the rate of fertilization. Regarding the incidence of OHSS, 26.2 % of participants in the control group experienced OHSS, while the occurrence rate was 15.7 % in the intervention group (P = 0.401). The incidence of severe OHSS in the control group and intervention group was 7.1 % and 3.6 %, respectively. ConclusionIntravenous injection of Calcium gluconate can be effective in preventing Ovarian Hyperstimulation Syndrome.

Cardiovascular prevention in old age

The enormous potential of cardiovascular prevention in terms of expanding the life span and health span is presently nowhere near being realized. The five classical cardiovascular risk factors body mass index (BMI), systolic blood pressure, non-high-density lipoprotein (non-HDL) cholesterol, tobacco smoking, and diabetes mellitus account for more than half of the cases of incident cardiovascular diseases. Cardiovascular prevention is also effective and adequate in seemingly healthy individuals aged 70years or above, although the association of several cardiovascular risk factors with cardiovascular diseases is less pronounced in old age. The cardiovascular risk of seemingly healthy persons aged 70years or above can validly be determined using the Systematic COronary Risk Evaluation-Older Persons (SCORE2-OP), leading to risk-adjusted clear treatment recommendations. National and international guidelines advocate individualized cardiovascular prevention in several domains including diet, physical activity and risk factor management through to old age.

Effects of fasting on golden pompano Trachinotus ovatus: Physiological and biochemical responses

In practice, golden pompano (Trachinotus ovatus) reared in offshore cages may experience short-term fasting during extreme weather conditions, yet their physiological and biochemical responses remain poorly understood. To investigate the responses of this species to fasting, golden pompano weighing 138.0 ± 2.1 g were deprived of food for 0, 7, 14, 21 and 28 days, followed by assessments of body mass, somatic parameters, proximate composition, muscle glycogen, intestinal digestive enzymes, muscle amino acid and fatty acid profiles, nutritional value, and muscle texture and water holding capacity. Lower condition factor, hepatosomatic index, viscerosomatic index, hepatic glycogen, and intestinal activities of amylase, lipase and trypsin were found in the fasted fish than in the non-fasted fish, while a sharp decline in these parameters occurred on the first week of fasting. Fresh mass, dry mass, whole body and muscle contents of crude protein and lipid tended to decrease with prolonged fasting, while no statistical differences were found in these parameters between fish fasted for 7 days and the non-fasted fish. Furthermore, the decline rate of whole body crude lipid was consistently higher than that of whole body crude protein throughout the different stages of the 28-day fasting period, with the exception of the second week of fasting. No statistical differences were found in the total essential amino acids (∑EAA), non-essential amino acids (∑NEAA), ratio of ∑EAA to∑NEAA, chemical score, amino acid score, and essential amino acid index in the fasted fish compared to the non-fasted fish of group 0d. Fasting altered the muscle profile of SFA, MUFA and PUFA, sometimes reducing muscle springiness (groups 7d and 14d) and hardness (group 21d). Compared to non-fasted fish, fish fasted for 7 and 14 days exhibited statistically lower or equal drip loss, water loss rate, storage losses, and frozen exudate rate, whereas higher cooked meat rate. The above results indicate that during the 28-day fasting period, the visceral fat and hepatic glycogen are initially metabolized, followed by body proteins and lipids, to maintain the normal metabolism in fasted fish. Fasting has beneficial effects on the nutritional value of amino acids and fatty acids in muscle, although it modifies the muscle SFA, MUFA and PUFA composition and sometimes has negative effects on muscle texture. Considering the balance between body weight loss and flesh quality improvement, the duration of fasting for cultured golden pompano prior to harvesting should not exceed 14 days.

Abstract C045: Targeting metabolic heterogeneity in pancreatic ductal adenocarcinoma

Abstract Pancreatic ductal adenocarcinoma (PDAC) is the third leading cause of cancer-related death in the US. This poor prognosis is related to the complex tumor microenvironment and tumor heterogeneity that drives resistance to chemotherapy. Here the heterogeneity of the stromal, immune, and malignant epithelial cell populations enables signaling and metabolic crosstalk mechanisms that drive immune suppression and resistance to therapy. Recently, our group unveiled that multiple distinct metabolic subclasses of cancer cells coexist within individual PDAC tumors. Among these, two PDAC clonal populations can engage in metabolic symbiosis that supports mitochondrial metabolism. One subtype has a constitutively high integrated stress response (ISR), enabling resistance to mitochondrial inhibitors. Further, these cells release asparagine that supports the mitochondrial metabolism of the other subtype that are more sensitive to mitochondrial inhibition. Given these observations, we are developing approaches to disrupt this metabolic symbiosis. Clonal PDAC cells with constitutively active ISR exhibit increased sensitivity to inhibition of the redox activity of Redox factor 1/Apurinic-apyrimidinic endonuclease 1 (Ref-1). Importantly, this can be leveraged to disrupt the support of the mitochondrial metabolism of clonal symbiotic pairs. Moreover, polyclonal PDAC cell lines representative of the heterogeneity seen in PDAC patients show a highly synergistic response to the combination of Ref-1 redox inhibitors with mitochondrial inhibitors. These data suggest this treatment combination will be synergistic in polyclonal tumors that exist in most PDAC patients. Overall, these data have the potential to lead to new avenues to directly target heterogenous populations of cancer cells to treat pancreatic cancer- an urgent clinical need. Citation Format: Taryn Morningstar, Cecily Anaraki, Lorenzo Scipioni, Giulia Tedeschi, Michelle Digman, Aimee Edinger, Claus Jorgensen, Melissa Fishel, Christopher J Halbrook. Targeting metabolic heterogeneity in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr C045.

Tiny but Mighty: Vipera ammodytes meridionalis (Eastern Long-Nosed Viper) Ontogenetic Venom Variations in Procoagulant Potency and the Impact on Antivenom Efficacies.

The Eastern Long-Nosed Viper (Vipera ammodytes meridionalis) is considered one of the most venomous snakes in Europe. However, it is unknown whether ontogenetic variation in venom effects occurs in this subspecies and how this may impact antivenom efficacy. In this study, we compared the procoagulant activities of V. a. meridionalis venom on human plasma between neonate and adult venom phenotypes. We also examined the efficacy of three antivenoms-Viperfav, ViperaTAb, and Inoserp Europe-across our neonate and adult venom samples. While both neonate and adult V. a. meridionalis venoms produced procoagulant effects, the effects produced by neonate venom were more potent. Consistent with this, neonate venom was a stronger activator of blood-clotting zymogens, converting them into their active forms, with a rank order of Factor X >> Factor VII > Factor XII. Conversely, the less potent adult venom had a rank order of FXII marginally more activated than Factor VII, and both much more so than Factor X. This adds to the growing body of evidence that activation of factors besides FII (prothrombin) and FX are significant variables in reptile venom-induced coagulopathy. Although all three examined antivenoms displayed effective neutralization of both neonate and adult V. a. meridionalis venoms, they generally showed higher efficacy on adult venom than on neonate venom. The ranking of antivenom efficacy against neonate venom, from the most effective to the least effective, were Viperfav, Inoserp Europe, ViperaTAb; for adult venom, the ranking was Inoserp Europe, Viperfav, ViperaTAb. Our data reveal ontogenetic variation in V. a meridionalis, but this difference may not be of clinical concern as antivenom was effective at neutralizing both adult and neonate venom phenotypes. Regardless, our results highlight a previously undocumented ontogenetic shift, likely driven by the documented difference in prey preference observed for this species across age classes.

Exploring herbal preconditioning strategies to improve adipose tissue stem cell therapy efficacy

The application of mesenchymal stem cells (MSCs), specifically those sourced from adipose tissue, has become a focal point in regenerative medicine, showing potential for effective cell therapy. Adipose-derived stem cells (ASCs) offer hope in treating various conditions, yet their viability and integration rates are hindered by the harsh post-transplantation environment marked by inflammation and oxidative stress at the injury site. Lawsonia inermis (henna), Zizyphus spina-christi (Christ's Thorn), and Glycyrrhiza glabra (licorice) are three plants known for their potent antioxidant properties primarily due to their rich content of phenolic compounds and flavonoids. Their potential health benefits make them valuable in both traditional medicine and modern therapeutic applications.This research delves into investigating and contrasting the effects of hydroalcoholic extracts from Lawsonia inermis, Zizyphus spina-christi, and Glycyrrhiza glabra on human adipose tissue stem cells as a pre-conditioning method to alleviate oxidative stress in cell therapy. ASCs were isolated from the pelvic cavity and abdomen, characterized using flow cytometry, and prompted to differentiate into adipogenic and osteogenic lineages. They were then subjected to different concentrations of the aforementioned herbal extracts at varying time frames, followed by MIC and MTT analysis. The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay and the Minimum Inhibitory Concentration (MIC) assay are both used to evaluate the effects of substances on cell viability and cell growth receptively. RNA extraction from ASCs and evaluation of antioxidant gene expression were conducted via Real-time PCR. Flow cytometry data confirmed the presence of MSC markers in the isolated cells from ASCs. Analysis of the herbal extract concentrations revealed that 50 ng/ml had a toxic effect on ASCs, establishing a toxicity reference point for adipose-derived MSCs. Real-time PCR results showcased a notable increase in the expression of antioxidant genes post-preconditioning with hydroalcoholic extracts. This study underscores, for the first time, the safety and lack of toxicity of preconditioning with these extracts for h.ASCs, enhancing their resilience and acclimatization under oxidative stress conditions, potentially boosting the therapeutic potential of h.ASCs. The antioxidant properties of Lawsonia inermis, Zizyphus spina-christi, and Glycyrrhiza glabra may enhance the expression of antioxidant genes (Sod, Gpx and Cata) through several mechanisms, including the activation of transcription factors like Nrf2 (Nuclear factor erythroid 2-related factor 2), which regulates antioxidant gene expression; modulation of signaling pathways such as MAPK (Mitogen-Activated Protein Kinase) and PI3K/Akt (Phosphoinositide 3-kinase/Protein Kinase B); and influencing epigenetic modifications like histone changes or DNA methylation that can alter gene expression. However, further research is necessary to clarify the specific mechanisms by which these plant extracts enhance antioxidant gene expression and to explore their potential therapeutic applications in combating oxidative stress-related diseases. Future proposals should consider more gene investigation and in vivo applications, as well as the long-term effects of pre-conditioning.

DNA breathing integration with deep learning foundational model advances genome-wide binding prediction of human transcription factors.

It was previously shown that DNA breathing, thermodynamic stability, as well as transcriptional activity and transcription factor (TF) bindings are functionally correlated. To ascertain the precise relationship between TF binding and DNA breathing, we developed the multi-modal deep learning model EPBDxDNABERT-2, which is based on the Extended Peyrard-Bishop-Dauxois (EPBD) nonlinear DNA dynamics model. To train our EPBDxDNABERT-2, we used chromatin immunoprecipitation sequencing (ChIP-Seq) data comprising 690 ChIP-seq experimental results encompassing 161 distinct TFs and 91 human cell types. EPBDxDNABERT-2 significantly improves the prediction of over 660 TF-DNA, with an increase in the area under the receiver operating characteristic (AUROC) metric of up to 9.6% when compared to the baseline model that does not leverage DNA biophysical properties. We expanded our analysis to in vitro high-throughput Systematic Evolution of Ligands by Exponential enrichment (HT-SELEX) dataset of 215 TFs from 27 families, comparing EPBD with established frameworks. The integration of the DNA breathing features with DNABERT-2 foundational model, greatly enhanced TF-binding predictions. Notably, EPBDxDNABERT-2, trained on a large-scale multi-species genomes, with a cross-attention mechanism, improved predictive power shedding light on the mechanisms underlying disease-related non-coding variants discovered in genome-wide association studies.

Energy Deficit and Factors Associated with Energy Balance during a Combat Deployment in U.S. Army Special Operation Forces Soldiers.

Forty-six SOF Soldiers (age: 30.1 ± 3.5 yrs, body mass index: 27.7 ± 4.1 kg/m2) completed surveys on demographic data, mission activity characteristics, gastrointestinal issues, ration consumption, resilience, mood state, and dietary intake using a 127-question food frequency questionnaire at the end of a six-month deployment. EE was estimated using a SOF-specific prediction equation with a physical activity factor of 2.1. A paired t-test compared reported energy intake (EI) with estimated energy expenditure (EE). Pearson correlations identified significant variables associated with energy balance, which were then incorporated into a multiple linear regression model. The regression analysis included Profile of Mood States (POMS) anger and POMS depression as predictor variables to determine their influence on energy balance. Reported mean EI was 2512 ± 1059 kcal·d-1, while estimated mean EE was 5272 ± 525 kcal·d-1. The mean energy imbalance was -2854 kcal/d (95% CI: -2655 to -3055, p < 0.001), with all participants in negative energy balance (range: -492 to -3813 kcal/d). POMS depression (r = 0.517, p < 0.01) and POMS anger (r = 0.363, p = 0.020) were associated with energy balance. The regression model was significant (R2 = 0.23, F (2, 38) = 7.02, p < 0.01), with POMS depression significantly predicting energy balance (β = 50.76, p = 0.011). Deployed SOF Soldiers reported high EE and limited EI, which may negatively impact performance. Higher POMS depression scores were associated with lower energy deficits. Future studies should investigate the relationship between mood and energy balance, using direct measures of EI and EE.

Osteoarthritis induced by monosodium iodoacetate model

Objectives: this study aimed to highlight the evolution of the osteoarthritis (OA) model induced by monosodium iodoacetate (MIA) and its collaboration with the knowledge of the pathophysiology of inflammation, hypernociception and cartilage degeneration. Methods: PubMed and Science Direct were used with the descriptors: knee osteoarthritis or osteoarthritis of ankle) and (rat or osteoarthritis induced by monoiodoacetate) and (animal model of osteoarthritis or joint nociception). Studies containing rats, mice, and monoiodoacetate-induced osteoathritis were included. Exclusion criteria were: Animal models of osteoarthritis induced by destabilization; meniscectomy-induced osteoarthritis; non-invasive animal models of osteoarthritis induced by fracture, compression and tibial overload; animal models of osteoarthritis with large animals and models of osteoarthritis by papain or bacterial collagenase. We summarized studies that used MIA to induce knee or ankle OA in rats or mice and the evolution of the inflammatory markers. Results: a total of 38 original manuscripts met the inclusion criteria and were considered in the study. The model of OA induced by MIA is well-established and explored with several methodologies and has been widely used in different species. MIA induces cell death, progressive loss of chondrocytes and histological changes that mimics human OA. Studies with this model demonstrated inflammation, neurogenic hyperalgesia, release of cytokines and matrix metalloproteinases. More recently, molecular biology data confirm the activation of nuclear transcription factors, which modulates the expression of citokines in apoptotic chondrocytes. Conclusion: the MIA-induced OA has been useful in predicting several characteristics for the elucidation of osteoarthritis pathophysiology.

Yak DEFB123 alleviates lung injury caused by Klebsiella pneumoniae through MAPKs signaling pathway

Beta-defensins, such as β-defensin 123 (DEFB123), are vital components of the immune system's defense against infections due to their strong antimicrobial properties and capacity for modulating the body's immunological responses. In this study, we successfully cloned and analyzed the yak DEFB123 gene sequence. Subsequently, we obtained recombinant protein DEFB123 (rDEFB123) through prokaryotic expression. Our results demonstrate that rDEFB123 effectively inhibits the growth of Escherichia coli, Klebsiella pneumoniae, and Staphylococcus aureus. Furthermore, rDEFB123 enhances the phagocytic activity of macrophages by regulating specific factors. In a mouse model infected with Klebsiella pneumoniae, the administration of rDEFB123 showed significantly lower levels of serum ALT and AST compared to the control group. Moreover, IFN-γ and IgG were significantly increased in the rDEFB123-treated groups, indicating an enhanced immune response. In the MAPKs signaling pathway of the infected mouse lungs, the expressions of JNK, TRAF2, TRAF6, MIF, and IL-1β genes were downregulated in the rDEFB123-treated groups. Moreover, the levels of p-JNK protein were significantly decreased in these groups as well. Klebsiella pneumoniae caused systemic infection with organ damage in mice. However, the administration of rDEFB123 suppressed the expressions of inflammatory factors, thereby mitigating organ injury and regulating the activity of apoptosis-related factors to enhance immunity. These findings provide valuable theoretical data for future exploration of the functionality and potential applications of DEFB123 in yak.

Minthostachys verticillata essential oil modulates cytokine synthesis and Staphylococcus aureus internalization in MAC-T cells at least through TLR4/MyD88/NFkB pathway.

The aim of this study was to evaluate the activation pathway(s) triggered by Minthostachys verticillata essential oil (EO) in bovine mammary epithelial cells (MAC-T) challenged with a strain of bovine Staphylococcus aureus. MAC-T cells were stimulated with EO, S. aureus or pre-treated with EO and then challenged with S. aureus. Cytokine's release was measured by ELISA. The mRNA for TLR2, TLR4, NOD2, MyD88 and NFκB was quantified by RT-qPCR. S. aureus adherence and internalization was also evaluated. MAC-T cells stimulated with S. aureus synthesized high levels of IL-1ß and IL-6 were kept up to 48 h, while IL-4 levels were not altered.Cells pre-treated with EO for 2 and 6h and then challenged with S. aureus showed a significant increase of IL-1β and IL-6. However, in these cells, a decrease in IL-1ß and IL-6 levels and an increase of IL-4 values was observed from 24 h. No significant increase in the expression levels of TLR2 or NOD2 was detected in all stimulated cells. However, the expression of TLR4, MyD88 and NFκB was increased in cells stimulated with S. aureus at 2 and 6 h as well as in cells pre-treated with EO between 2 and 6h and then challenged with S. aureus. The NFκB expression levels was similar to control at 24h in all stimulated cells, although pro-inflammatory cytokine levels and TLR4 and MyD88 expression levels remained high in cells stimulated with S. aureus. This results suggested the activation of other pathways independent of MyD88 by the pathogen that involucrated the activation of others transcription factors. Pre-treatment with EO during 2, 6 and 24h did not affect S. aureus adherence but decreased its internalization. In conclusion, pre-treatment with EO increased the IL-1β and IL-6 synthesis during the first hours post-challenged with S. aureus up-regulating TLR4/MyD88/NFκB pathway. Furthermore, EO increased the IL-4 levels from 6 to 24 h down-regulating the NFκB and possibly other transcription factors activated by the pathogen, which decreased its internalization into MAC-T cells.