Biology subjects

Bernardo, A.

Publications and source records attributed to Bernardo, A..

10 recordsLinked to original sources

bioRxiv2026

Nanomechanical phenotyping uncovers a non-canonical smooth muscle-like hypercontractility program in lung squamous cell carcinoma myofibroblasts

Myofibroblast-like cancer-associated fibroblasts (myCAFs) drive the stiff, tumor-promoting microenvironment, yet their mechanical phenotype in non-small cell lung cancer (NSCLC) remains undefined. myCAFs from major NSCLC subtypes (lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC)) were examined combining atomic force microscopy and traction force microscopy with molecular and signaling analyses. LUAD-myCAFs displayed canonical myofibroblast features, including elevated stiffness, traction forces, ROCK-MYPT1 signaling, and -SMA-rich stress fibers. Unexpectedly, LUSC-myCAFs amplified these nanomechanical features despite reduced ROCK-MYPT1 signaling and comparable -SMA levels. Instead, they exhibited a smooth muscle cell (SMC)-like contractile program characterized by enhanced SM-MHC(MYH11) expression colocalized with -SMA, ERK1/2-RLC hyperactivation, and DNA methylation reprogramming including SMC-contractility regulators. LUSC-myCAF hypercontractility was linked with reduced proliferative capacity and smaller intratumoral vessel lumens, consistent with the low CAF density and enhanced hypoxia patterns reported in LUSC. SMC markers were enriched and associated with adverse outcomes selectively in LUSC, highlighting the relevance of SMC-like myCAFs in LUSC progression. These findings unveil subtype-specific myCAF programs in NSCLC, and identify -SMA/SM-MHC(MYH11) myCAFs as key determinants of tumor mechanics, providing a rationale for the survival benefits of antihypertensive drugs targeting SMC hypercontractility. Moreover, they demonstrate that nanomechanical phenotyping can uncover translationally relevant myCAF phenotypes invisible to molecular profiling.

biophysics↗

bioRxiv2026

Mapping of Stripe Rust and Leaf Rust Resistance Genes in the Hard Red Winter Wheat Population Green Hammer/Lonerider

Stripe rust and leaf rust, caused by Puccinia striiformis f. sp. tritici and P. triticina, respectively, are the most destructive wheat diseases in the southern Great Plains. Green Hammer is a hard red winter wheat (HRWW) cultivar released by Oklahoma State University in 2018 and has demonstrated a stable adult plant resistance to stripe rust and race-specific seedling resistance to leaf rust. To identify and map rust resistance loci, 109 doubled haploid (DH) lines derived from the cross between Green Hammer and another HRWW cultivar, Lonerider, were developed. Lonerider showed adult plant resistance to stripe rust but was susceptible to multiple P. triticina races. The DH lines were evaluated for stripe rust at the adult plant stage in greenhouse and field environments across Oklahoma, Kansas, and Washington, and for leaf rust at the seedling stage against seven U.S. P. triticina races and at the adult plant stage in Oklahoma and Texas. Genotyping-by-sequencing generated 6,078 polymorphic single-nucleotide polymorphisms used for genetic mapping. Quantitative trait loci (QTL) analysis identified 14 stripe rust and 8 leaf rust resistance QTL. For stripe rust, a major QTL in Green Hammer, QYr.osughln-2AS, was identified in the proximity of the 2NvS translocation. Three other major stripe rust resistance QTL were identified in Lonerider on chromosomes 2AL (two QTL) and 2BS (one QTL). For leaf rust, QLr.osughln-1DS and QLr.osughln-2DS.1 were the two major QTL identified in Green Hammer and most likely correspond to the all-stage resistance genes Lr21 and Lr39, respectively. In this study, we identified previously characterized genes as well as unknown genes that can be utilized in wheat breeding programs to enhance resistance to leaf rust and stripe rust.

genetics↗

bioRxiv2025

Identification of stripe rust adult plant resistance genes in the hard winter wheat cultivar Bakers Ann

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is among the most destructive wheat diseases. Identifying resistance genes is crucial for the development of resistant cultivars. "Bakers Ann", a hard winter wheat cultivar developed by Oklahoma State University, has shown durable adult plant resistance to stripe rust. To dissect the genetic basis underlying stripe rust resistance in Bakers Ann, 125 doubled haploid lines, derived from the cross OK12D22004-016 x Bakers Ann, were evaluated at the adult plant stage in the greenhouse and in field environments in Oklahoma, Kansas, and Washington. This population was genotyped using genotyping-by-sequencing, which produced 7,268 single-nucleotide polymorphisms for genetic mapping. Quantitative trait loci (QTL) analysis identified six loci, four from Bakers Ann on chromosomes 2DL, 4BS, 4BL, and 7BL, and two from OK12D22004-016 on chromosomes 2AS and 2AL. Although OK12D22004-016 is susceptible in the US Great Plains, it was found to carry QYr.osu-2AS, which was linked to Yr17 on the 2NvS translocation and explained up to 30% of the phenotypic variation, but was effective in a single location in Washington. Two major QTL were identified in Bakers Ann, QYr.osu-2DL on chromosome 2DL that explained up to 57% of the phenotypic variation and mapped close to Yr54, and QYr.osu-4BL on chromosome 4BL that explained up to 15% of the phenotypic variation and mapped close to Yr62. Resistance in Bakers Ann resulted from additive effects of the four QTL. Two kompetitive allele-specific PCR markers were developed for QYr.osu-2DL to facilitate marker-assisted selection for stripe rust resistance.

genetics↗

bioRxiv2025

Protective effects of central leptin on whole-body energy homeostasis upon acute olanzapine exposure

Second-generation antipsychotic use is associated with severe metabolic side effects such as obesity and type 2 diabetes. Leptin is a hormone that is secreted by adipose tissue, and it acts on the brain to decrease body weight by reducing food intake and stimulating energy expenditure. Leptin also improves glucose and lipid metabolism. We examined the short-term impact of olanzapine, a commonly used second-generation antipsychotic, on the central leptin-mediated regulation of energy balance, lipid metabolism, and hypothalamic kinase activity. Male Sprague Dawley rats were given an acute intracerebroventricular (ICV, 3rd ventricle) injection of either leptin or vehicle, combined with subcutaneous olanzapine or vehicle. As expected, ICV leptin decreased food intake and importantly, olanzapine did not block this effect. Administration of leptin, olanzapine, or their combination reduced the average respiratory exchange ratio (RER) during the light cycle, which indicates that fat oxidation was increased. In the dark cycle, leptin decreased the average RER regardless of olanzapine administration, and in the presence of leptin, olanzapine did not affect the average RER. Leptin did not alter the olanzapine-induced increase in serum triglyceride concentrations. Olanzapine and central leptin treatment differentially activated hypothalamic kinases. In conclusion, regulation of food intake and fuel preference by central leptin is intact following acute olanzapine administration.

neuroscience↗

bioRxiv2024

Genetic modulation of yield and phenotypic plasticity of yield in winter wheat

For winter wheat in the US Central Great Plains, phenotypic plasticity of yield is agronomically adaptive, i.e., genotypes with higher plasticity have higher yield in high yielding environments with no tradeoff in stressful, low yielding environments. Using data from variety trials conducted between 2000 and 2022 and cultivars released between 1967 and 2022, we explored time trends in phenotypic plasticity and heritability of yield. We hypothesize that i) if yield plasticity is agronomically adaptive, then newer cultivars will have higher yield plasticity; ii) heritability of yield is declining in the time series; and iii) genomic regions associated with yield depend on the environment and do not fully overlap with those associated with phenotypic plasticity of yield. Breeding for yield and agronomic adaptation increased phenotypic plasticity of yield at 0.5% year -1; broad sense heritability of yield decreased from 0.23 in 1993 to 0.15 in 2017. Genome-wide-association analysis shows genomic regions associated with yield varied between high yielding and stressful environments and were partially independent of those associated with yield plasticity. Newer cultivars have a higher frequency of alleles associated with yield and its plasticity. We discuss implications for breeding and agronomy aimed to improve wheat phenotypes.

genetics↗

bioRxiv2024

Identification of leaf rust resistance loci in hard winter wheat using genome-wide association mapping

Leaf rust, caused by Puccinia triticina (Pt), is a serious constraint to wheat production. Developing resistant varieties is the best approach to managing this disease. Wheat leaf rust resistance (Lr) genes have been classified into either all-stage resistance (ASR) or adult-plant resistance (APR). The objectives of this study were to identify sources of leaf rust resistance in contemporary U.S. hard winter wheat (HWW) and to dissect the genetic basis underlying leaf rust resistance in HWW. A panel of 732 elite HWW genotypes was evaluated for response to U.S. Pt races at the seedling stage and at the adult plant stage in leaf rust nurseries in Oklahoma, Texas, and Kansas. Further, the panel was genotyped using Multiplex Restriction Amplicon Sequencing (MRA-Seq) and DNA markers linked to the known ASR genes Lr18, Lr19, Lr21, Lr24, Lr37, and Lr42 and APR genes Lr34, Lr46, Lr67, Lr68, Lr77, and Lr78. Single nucleotide polymorphism (SNP) markers derived from MRA-Seq, DNA markers linked to the known Lr genes, and the phenotypic data were used for genome-wide association study (GWAS) to identify markers associated with leaf rust response. Gene postulation based on leaf rust reactions, DNA markers, and GWAS suggested the presence of Lr1, Lr2a, Lr10, Lr14a, Lr16, Lr18, Lr19, Lr21, Lr24, Lr26, Lr34, Lr37, Lr39, Lr42, Lr46, Lr68, Lr77, and Lr78 in the HWW panel. The GWAS identified 59 SNPs significantly associated with leaf rust response, of which 20 were likely associated with novel resistance loci and can be used to enhance wheat leaf rust resistance.

genomics↗

bioRxiv2024

Positive Allosteric Modulation of the α5-GABAA receptors prevents neuronal atrophy and cognitive decline independently of tau tangle accumulation in the PS19 mouse model

BackgroundDysregulated Tau phosphorylation (p-Tau) is a hallmark of neurodegenerative disorders such as Alzheimers disease (AD) or frontotemporal dementia (FTD), resulting in neurofibrillary tangle accumulation, neuronal atrophy and cognitive impairment. Impaired somatostatin (SST) expression and reduced SST-expressing GABAergic neurons significantly contributes to AD-related pathophysiology and correlates with cognitive impairment. SST+ interneurons inhibit the dendrites of excitatory neurons in cortical layers and hippocampus, primarily through 5-GABA-A receptors, regulating cognitive function. Leveraging a newly developed small molecule that targets the 5-GABA-A receptors via positive allosteric modulation (5-PAM), we assessed its effects on p-Tau-related neuronal morphology, cognitive deficits and protein expression. MethodsIn the PS19 transgenic mouse model, we administered the 5-PAM, GL-II-73, either acutely or chronically at 3 and 6 months, corresponding to early and advanced stage of p-Tau accumulation. Golgi staining analyzed dendritic morphology and spine density in mice chronically exposed to 5-PAM. Western blotting was used to quantify p-tau and Tau expression. Spatial working memory was assessed using the Y-maze. ResultsChronic treatment at 3 and 6 months mitigated p-Tau-induced loss of spine density and reduced dendritic length. 5-PAM treatment did not affect p-tau levels. 5-PAM effectively reversed spatial working memory deficits induced by p-tau accumulation both acutely and chronically. Conclusions5-GABA-A receptor positive allosteric modulation displayed neurotrophic (spine and dendritic pathology) and procognitive (working memory) effects in the PS19 model, independently of p-Tau burden. This suggests a novel therapeutic strategy for p-Tau-related pathologies with both symptomatic and disease-modifying potential.

neuroscience↗

bioRxiv2022

Inhibition of hyperactive cyclin dependent kinase 5/p25 is protective in the 6-hydroxydopamine model of Parkinson's disease

BackgroundCyclin-dependent kinase 5 (CDK5) is a multifunctional enzyme involved in neuronal development, maturation and survival. CDK5 activity is tightly regulated by association with regulatory proteins p35 and p39. Upon neuronal insults, increased intracellular calcium activates calpain, cleaving p35 into p25, which has a higher affinity for CDK5. p25 hyperactivates CDK5, initiating apoptotic cascades that lead to significant dopaminergic (DAergic) loss that can leads to neurodegenerative disorders, such as Parkinsons disease (PD). ObjectiveThis study investigates hyperactivation of CDK5/p25 in the 6-hydroxydopamine (6-OHDA) rat model of PD and specific inhibition of CDK5/p25 by truncated peptide 5 (TP5). TP5 was investigated for amelioration of 6-OHDA induced behaviour impairments and significant protection of dopamine neurons through tyrosine hydroxylase (TH). Methods6-OHDA induced motor impairments and reduced TH. Motor assessments included locomotor activity, beam transversal, fixed speed rotarod and amphetamine-induced rotations. Immunohistochemistry investigated DAergic neurodegeneration using TH levels and immunoprecipitation and assay investigated CDK5 activity. ResultsPre-administration of TP5 maintained locomotor activity, preserved beam transversal scores, protected motor coordination and attenuated amphetamine induced rotations in 6-OHDA lesioned rats, all indicative of neuroprotection by TP5. 6-OHDA without pretreatment of TP increased CDK5 activation. CDK5 activity in TP5+6-OHDA animals was not significantly different from artificial cerebrospinal fluid (aCSF) treated sham surgery controls. Immunohistochemistry revealed significant TH protection within the substantia nigra (SN) of TP5 pretreated animals. Conclusions6-OHDA increases CDK5 activity. Hyperactive CDK5/p25 inhibition in the 6-OHDA model has neuroprotective capability, protecting against the development of a toxin-based induction of PD-like motor phenotypes and pathology. This supports CDK5/p25 specific inhibition as a target for further neuroprotective therapeutic development.

pharmacology and toxicology↗

bioRxiv2021

In-vitro cellular reprogramming to model gonad development and its disorders

During embryonic development, mutually antagonistic signaling cascades determine the fate of the bipotential gonad towards a testicular or ovarian identity. Errors in this process result in human Disorders of Sex Development (DSDs), where there is discordance between chromosomal, gonadal, and anatomical sex. The absence of an appropriate, accessible in-vitro system is a major obstacle in understanding mechanisms of sex-determination/DSDs. Here, we describe protocols for differentiation of mouse and human pluripotent cells towards gonadal progenitors. Transcriptomic analysis reveals that the in-vitro-derived murine gonadal cells are equivalent to E11.5 in-vivo progenitors. Using similar conditions, Sertoli-like cells derived from 46,XY human induced pluripotent stem cells (hiPSCs) exhibit sustained expression of testis-specific genes, secrete AMH, migrate and form tubular structures. The cells derived from a 46,XY DSD female hiPSCs, carrying a NR5A1 variant, show aberrant gene expression and absence of tubule formation. CRISPR/Cas9-mediated correction of the variant rescued the phenotype. This is a robust tool to understand mechanisms of sex-determination and model DSDs.

developmental biology↗

bioRxiv2021

Symptomatic and Disease-Modifying Effects of GABAA Receptor Positive Allosteric Modulation in a Mouse Model of Chronic Stress

Chronic stress is a risk factor for Major depressive disorder (MDD), and in rodents, it recapitulates human behavioral, cellular and molecular changes. In MDD and after chronic stress, neuronal dysfunctions and deficits in GABAergic signaling are observed and responsible for symptom severity. GABA signals predominantly through GABAA receptors (GABAA-R) composed of various subunit types that relate to downstream outcomes. Activity at 2-GABAA-Rs contributes to anxiolytic properties, 5-GABAA-Rs to cognitive functions, and 1-GABAA-Rs to sedation. Therefore, a therapy aiming at increasing 2- and 5-GABAA-Rs activity, but devoid of 1-GABAA-R activity, has potential to address several symptomologies of depression while avoiding side effects. This study investigated the activity profiles and behavioral efficacy of two molecules enantiomers of each other (GL-II-73 and GL-I-54), separately and as a racemic mixture (GL-RM), and potential disease-modifying effects on neuronal morphology. Results confirm GL-I-54 and GL-II-73 exert positive allosteric modulation at the 2-, 3-, 5-GABAA-Rs and 5-containing GABAA-Rs, respectively, and have anti-depressant and pro-cognitive effects independently. Using unpredictable chronic mild stress (UCMS) in male and female mice (n=12/group), we show that acute and chronic administration of GL-RM combined the anti-depressant and pro-cognitive effects of each enantiomer, although at lower doses avoiding sedation. Morphology studies showed reversal of spine density loss caused by UCMS after chronic GL-RM treatment at apical and basal dendrites of the PFC and CA1. Together, these results support using a racemic mixture with combined 2-, 3-, 5-GABAA-R profile to reverse chronic stress-induced mood symptoms, cognitive deficits, and with anti-stress neurotrophic effects.

neuroscience↗