Phenotypic Changes Research Articles

SFRP4 promotes autophagy and blunts FSH responsiveness through inhibition of AKT signaling in ovarian granulosa cells

BackgroundSecreted frizzled-related proteins (SFRPs) comprise a family of WNT signaling antagonists whose roles in the ovary are poorly understood. Sfrp4-null mice were previously found to be hyperfertile due to an enhanced granulosa cell response to gonadotropins, leading to decreased antral follicle atresia and enhanced ovulation rates. The present study aimed to elucidate the mechanisms whereby SFRP4 antagonizes FSH action.MethodsPrimary cultures of granulosa cells from wild-type mice were treated with FSH and/or SFRP4, and effects of treatment on gene expression were evaluated by RT-qPCR and RNAseq. Bioinformatic analyses were conducted to analyse the effects of SFRP4 on the transcriptome, and compare them to those of FSH or a constitutively active mutant of FOXO1. Additional granulosa cell cultures from wild-type or Sfrp4-null mice, some pretreated with pharmacologic inhibitors of specific signaling effectors, were used to examine the effects of FSH and/or SFRP4 on signaling pathways, autophagy and apoptosis by western blotting and TUNEL.ResultsTreatment of cultured granulosa cells with recombinant SFRP4 was found to decrease basal and FSH-stimulated mRNA levels of FSH target genes. Unexpectedly, this effect was found to occur neither via a canonical (CTNNB1-dependent) nor non-canonical WNT signaling mechanism, but was found to be GSK3β-dependent. Rather, SFRP4 was found to antognize AKT activity via a mechanism involving AMPK. This lead to the hypophosphorylation of FOXO1 and a decrease in the expression of a portion of the FSH and FOXO1 transcriptomes. Conversely, FSH-stimulated AMPK, AKT and FOXO1 phosphorylation levels were found to be increased in the granulosa cells of Sfrp4-null mice relative to wild-type controls. SFRP4 treatement of granulosa cells also induced autophagy by signaling via AKT-mTORC1-ULK1, as well as apoptosis.ConclusionsThis study identifies a novel GSK3β-AMPK-AKT signaling mechanism through which SFPR4 antagonizes FSH action, and further identifies SFRP4 as a novel regulator of granulosa cell autophagy. These findings provide a mechanistic basis for the phenotypic changes previously observed in Sfrp4-null mice, and broaden our understanding of the physiological roles of WNT signaling processes in the ovary.

Photothermal treatment-based heat stress regulates function of myeloid-derived suppressor cells

Photothermal therapy is an alternative cancer therapy that uses a photothermal agent with light irradiation to induce fatal hyperthermia in cancer cells. In a previous study, we found that ex vivo photothermal (PT) treatment induced expression of heat shock proteins (HSPs), such as HSP70, HSP27, and HSP90, in cancer cells; moreover, immunization with lysates from PT-treated tumor cells resulted in significant tumor growth inhibition in tumor-bearing mice. In this study, we hypothesized that sublethal PT treatment of antigen-presenting cells regulates their immunogenicity. We observed the upregulation of expression of intracellular HSP70 and surface activation markers, such as CD40, CD80, CD86, and MHC class II, in sublethal PT-treated cells. The protumoral activity of myeloid-derived suppressor cells (MDSCs) was reduced by sublethal hyperthermia. Furthermore, poorly immunogenic MDSCs were converted into immunogenic antigen-presenting cells by PT treatment. The differences in immunogenicity between MDSCs untreated or treated with the PT technique were evaluated using the Student’s t-test or Mann–Whitney rank sum test. Collectively, direct hyperthermic treatment resulted in phenotypic changes and the functional regulation of immune cells.

Aging human abdominal subcutaneous white adipose tissue at single cell resolution.

White adipose tissue (WAT) is a robust energy storage and endocrine organ critical for maintaining metabolic health as we age. Our aim was to identify cell-specific transcriptional aberrations that occur in WAT with aging. We leveraged full-length snRNA-Seq and histology to characterize the cellular landscape of human abdominal subcutaneous WAT in a prospective cohort of 10 younger (≤30 years) and 10 older individuals (≥65 years) balanced for sex and body mass index (BMI). The older group had greater cholesterol, very-low-density lipoprotein, triglycerides, thyroid stimulating hormone, and aspartate transaminase compared to the younger group (p < 0.05). We highlight that aging WAT is associated with adipocyte hypertrophy, increased proportions of lipid-associated macrophages and mast cells, an upregulation of immune responses linked to fibrosis in pre-adipocyte, adipocyte, and vascular populations, and highlight CXCL14 as a biomarker of these processes. We show that older WAT has elevated levels of senescence marker p16 in adipocytes and identify the adipocyte subpopulation driving this senescence profile. We confirm that these transcriptional and phenotypical changes occur without overt fibrosis and in older individuals that have comparable WAT insulin sensitivity to the younger individuals.

Spinal cord microglia drive sex differences in ethanol-mediated PGE2-induced allodynia

The mechanisms of how long-term alcohol use can lead to persistent pain pathology are unclear. Understanding how earlier events of short-term alcohol use can lower the threshold of non-painful stimuli, described as allodynia could prove prudent to understand important initiating mechanisms. Previously, we observed that short-term low-dose alcohol intake induced female-specific allodynia and increased microglial activation in the spinal cord dorsal horn. Other literature describes how chronic ethanol exposure activates Toll-like receptor 4 (TLR4) to initiate inflammatory responses. TLR4 is expressed on many cell types, and we aimed to investigate whether TLR4 on microglia is sufficient to potentiate allodynia during a short-term/low-dose alcohol paradigm. Our study used a novel genetic model where TLR4 expression is removed from the entire body by introducing a floxed transcriptional blocker (TLR4-null background (TLR4LoxTB)), then restricted to microglia by breeding TLR4LoxTB animals with Cx3CR1:CreERT2 animals. As previously reported, after 14 days of ethanol administration alone, we observed no increased pain behavior. However, we observed significant priming effects 3 hrs post intraplantar injection of a subthreshold dose of prostaglandin E2 (PGE2) in wild-type and microglia-TLR4 restricted female mice. We also observed a significant female-specific shift to pro-inflammatory phenotype and morphological changes in microglia of the lumbar dorsal horn. Investigations in pain priming-associated neuronal subtypes showed an increase of c-Fos and FosB activity in PKCγ interneurons in the dorsal horn of female mice directly corresponding to increased microglial activity. This study uncovers cell- and female-specific roles of TLR4 in sexual dimorphisms in pain induction among non-pathological drinkers.

Development and Progression of Polyneuropathy Over 5 Years in Patients With Type 2 Diabetes.

There is a need for knowledge regarding the natural course of diabetic polyneuropathy (DPN), a complication in type 2 diabetes (T2D). The aim of this study was to examine the development of DPN over time. Patients with newly diagnosed T2D, recruited from a national cohort, and controls without diabetes of similar age and sex, underwent sensory phenotyping in 2016-2018. The Toronto consensus criteria were used to classify patients into possible, probable, and confirmed DPN. For this 5-year, observational, follow-up, cohort study, all participants were invited to a reexamination combining bedside sensory examination, quantitative sensory testing (QST), nerve conduction studies (NCSs), and skin biopsies measuring intraepidermal nerve fiber density (IENFD) in order to compare phenotypic and diagnostic changes over time. Of the baseline 389 patients and 97 controls, 184 patients (median [interquartile range] diabetes duration 5.9 [4.1-7.4] years, mean hemoglobin A1c [HbA1c] 51 ± 11 mmol/mol at baseline) and 43 controls completed follow-up (46.9%). Confirmed DPN was present in 35.8% and 50.3%, probable DPN in 27.2% and 14.6%, possible DPN in 17.2% and 16.6%, and no DPN in 15.2% and 17.9% at baseline and follow-up, respectively. The estimated prevalence (95% CI) of confirmed DPN was 33.5% (24.9-42.1) compared with 22.7% (17.5-28.0) at baseline. During the follow-up period, 43.9% of patients with probable DPN developed confirmed DPN. Progression of neuropathy occurred in 16.5% and 24.7% and regression in 5.9% and 18.6% of patients based on NCS and IENFD, respectively. Progression based on NCS and/or IENFD was associated with higher baseline waist circumference and triglycerides, and regression with lower baseline HbA1c. Patients with at least probable DPN at baseline but neither patients without DPN nor controls developed increased spread of hyposensitivity, more hyposensitivity on QST and lower NCS z-scores at follow-up, and worsening of nerve parameters at follow-up correlated with higher baseline triglycerides. In patients with well-regulated T2D, the proportion of patients with confirmed DPN increased over 5 years driven by progression from probable DPN. A large proportion of patients progressed, and a smaller proportion regressed on nerve parameters. Higher triglycerides correlated with this progression and may constitute a risk factor.

Analysis of The Results of The Rhesus System Blood Group Phenotyping Examination in Routine Type O Blood Donors

The Rhesus (Rh) system is the second most important blood group system after ABO, has the highest immunogenicity and tends to produce alloantibodies that cause most transfusion reactions. In the Rh blood group system, there are more than 55 antigens, but the main ones are D, C, E, c, and e. Currently, only the D antigen is carried out without examining four other important Rh antigens. Still scarce, studies on the proportion of Rh antigens, except D, in Indonesia; in this study we have determined Rh antigens and phenotypes among a population of routine blood donors. The purpose of this study was to determine the phenotypic variation and frequency distribution of the main antigens of the Rhesus blood group system in routine blood type O blood donors at UDD PMI Depok City. This study was descriptive and observational with a cross-sectional design. Primary data are obtained from the Rhesus system antigen examination results on donor blood samples. The sample was 140 blood samples from 140 routine donors. A rhesus phenotyping examination was carried out using the gel method. The study was conducted on 140 routine donor blood samples at UDD PMI Depok City. The characteristics of the subjects were male (69.29%) and female (30.71%), with an average age of 38 years. Samples are analyzed for five major Rhesus antigens. Rhesus antigen test shows D antigen (100%), e antigen (97.14%), C antigen (95.71%), c antigen (36.43%) and E antigen (27.86%). Rhesus phenotype results were R1R1 (DCe/Dce) (62.86%), R1R2 (DCe/DcE) (22.86%), R1r (DCe/dce) (9.29%), R2R2 (DcE/DcE) (2.86%), R2r (DcE/dce) (1.43%), and R1RZ (DCe/DCE) (0.71%). In conclusion, the frequency of phenotypic changes in the Rh blood group system of regular blood donors at UDD PMI Depok City is classified as the most common types: DCe/DCe, DCe/DcE, DCe/dce, DcE/DcE, DcE/dce and DCe/DCE.

Age-Disturbed Vascular Extracellular Matrix Links to Abdominal Aortic Aneurysms.

Abdominal aortic aneurysm (AAA) is a common but life-threatening vascular condition in men at an advanced age. However, the underlying mechanisms of age-increased incidence and mortality of AAA remain elusive. Here, we performed RNA sequencing (RNA-seq) of mouse aortas from males (young: 3-month, n = 4 vs old: 23-month, n = 4) and integrated with the data sets of human aortas (young: 20-39, n = 47 vs old: 60-79 years, n = 92) from GTEx project and the data set (GSE183464) for AAA to search for age-shifted aortic aneurysm genes, their relevant biological processes, and signaling pathways. Angiotensin II-induced AAA in mice was used to verify the critical findings. We found 1001 genes transcriptionally changed with ages in both mouse and human. Most age-increased genes were enriched intracellularly and the relevant biological processes included mitochondrial function and translational controls, whereas the age-decreased genes were largely localized in extracellular regions and cell periphery and the involved biological processes were associated with extracellular matrix (ECM). Fifty-one were known genes for AAA and found dominantly in extracellular region. The common age-shifted vascular genes and known aortic aneurysm genes had shared functional influences on ECM organization, apoptosis, and angiogenesis. Aorta with angiotensin II-induced AAA exhibited similar phenotypic changes in ECM to that in old mice. Together, we present a conserved transcriptional signature for aortic aging and provide evidence that mitochondrial dysfunction and the imbalanced ribosomal homeostasis act likely as driven-forces for aortic aging and age-disturbed ECM is the substrate for developing AAA.

Substrate Curvature Influences Cytoskeletal Rearrangement and Modulates Macrophage Phenotype.

Inflammation serves as a critical defense mechanism against pathogens and tissue damage but can lead to chronic diseases, such as cardiovascular disease and diabetes, when dysregulated. Macrophages play a pivotal role in orchestrating inflammatory responses, transitioning from pro-inflammatory M1 to anti-inflammatory M2 phenotypes to resolve inflammation and promote tissue repair. Current approaches to modulate macrophage phenotype predominantly rely on biochemical cues, which may induce systemic side effects. Given the mechanosensitivity of macrophages, this study investigates biophysical cues, specifically substrate curvature, as a localized strategy to regulate macrophage phenotype and minimize systemic repercussions. We hypothesized that substrate curvature influences macrophage immunophenotype by modulating F-actin polymerization. To test this hypothesis, we fabricated spherical microgels with tunable curvatures and characterized their biophysical properties. Our findings indicate that macrophages adhere to microgel surfaces irrespective of curvature, but the curvature significantly alters F-actin dynamics. Furthermore, manipulating cytoskeletal dynamics via selective actin inhibition partially reversed curvature-induced changes in macrophage phenotype. These results underscore the pivotal role of substrate curvature in modulating macrophage behavior and immunophenotype. Overall, our study demonstrates that substrate curvature significantly influences macrophage cytoskeletal dynamics and resulting immunophenotype. This simple approach can be utilized as a localized immunomodulatory treatment for inflammatory diseases.

Exploratory phase I study of HF1K16 for the treatment of patients with refractory/recurrent advanced glioma.

TPS32 Background: Glioblastoma (GBM) is the most common primary and aggressive malignant brain tumor in adults. Almost all patients with glioblastoma eventually have disease relapse and the median overall survival (OS) remains at only 30 weeks for those recurrent GBM patients. However, very few therapies had been approved over the past two decades, emphasizing the need for novel treatments. It is now wildly acknowledged that the immune microenvironment in brain offers adequate opportunities to implement immunotherapy for the treatment. Since most recurrent tumors had previously been exposed to the genotoxic stress of irradiation and/or chemotherapy, it is predicted to resulted in a higher mutational load and more immunogenic. However, The limited success of T cell immunomodulatory approach for advanced glioma has prompted a deeper understanding of the brain tumor and the immune microenvironment. GBM tumors are well-appreciated to be a immunosuppressive and a hallmark of GBM immunosuppression is the appearance of circulating myeloid-derived suppressor cells (MDSCs) at higher levels. Our preliminary data suggests that HF1K16, a liposome ATRA (all trans retinoic acid) suspension, is capable of relieving immune suppression induced by MDSCs. Further, we recently completed a phase I dose escalation study which showed that HF1K16 is safe, well-tolerated with preliminary efficacy in brain tumor. Thus, we hypothesized that HF1K16 may reverse GBM-associated immune suppression and leading to an increase in TILs and improved survival. Methods: This is an advanced glioma-specific expansion arm for adult patients with prior confirmed brain tumor and failed standard treatment. We plan to enroll 20~30 evaluable patients. Key eligibility criteria include candidates age ≥ 18 years, Karnofsky performance status ≥ 60 with and at least one measurable lesion. The primary endpoint is determination of overall response rate (ORR), duration of response (DOR), disease control rate (DCR) and progression-free survival (PFS) according to RANO criteria. Secondary endpoints include longitudinal assessment of the peripheral immune response by changes in MDSC and T cell subsets and numbers. HF1K16 infusions were administered in 21-day cycles (q.o.d days 1-14). Prior to and during treatment, peripheral blood mononuclear cells were collected and analyzed with flow cytometry to monitor the changes in myeloid cell phenotype and T cell composition. The recruitment of patients is anticipated completed in 2024. Clinical trial information: NCT05388487 .

Evolution of phenotypic plasticity leads to tumor heterogeneity with implications for therapy.

Cancer is a significant global health issue, with treatment challenges arising from intratumor heterogeneity. This heterogeneity stems mainly from somatic evolution, causing genetic diversity within the tumor, and phenotypic plasticity of tumor cells leading to reversible phenotypic changes. However, the interplay of both factors has not been rigorously investigated. Here, we examine the complex relationship between somatic evolution and phenotypic plasticity, explicitly focusing on the interplay between cell migration and proliferation. This type of phenotypic plasticity is essential in glioblastoma, the most aggressive form of brain tumor. We propose that somatic evolution alters the regulation of phenotypic plasticity in tumor cells, specifically the reaction to changes in the microenvironment. We study this hypothesis using a novel, spatially explicit model that tracks individual cells' phenotypic and genetic states. We assume cells change between migratory and proliferative states controlled by inherited and mutation-driven genotypes and the cells' microenvironment. We observe that cells at the tumor edge evolve to favor migration over proliferation and vice versa in the tumor bulk. Notably, different genetic configurations can result in this pattern of phenotypic heterogeneity. We analytically predict the outcome of the evolutionary process, showing that it depends on the tumor microenvironment. Synthetic tumors display varying levels of genetic and phenotypic heterogeneity, which we show are predictors of tumor recurrence time after treatment. Interestingly, higher phenotypic heterogeneity predicts poor treatment outcomes, unlike genetic heterogeneity. Our research offers a novel explanation for heterogeneous patterns of tumor recurrence in glioblastoma patients.

Genetic background affects neutrophil activity and determines the severity of autoinflammatory osteomyelitis in mice.

The knowledge about the contribution of the innate immune system to health and disease is expanding. However, to obtain reliable results, it is critical to select appropriate mouse models for in vivo studies. Data on genetic and phenotypic changes associated with different mouse strains can assist in this task. Such data can also facilitate our understanding of how specific polymorphisms and genetic alterations affect gene function, phenotypes, and disease outcomes. Extensive information is available on genetic changes in all major mouse strains. However, comparatively little is known about their impact on immune response and in particular on innate immunity. Here, we analyzed a mouse model of chronic multifocal osteomyelitis (CMO), an autoinflammatory disease driven exclusively by the innate immune system, which is caused by an inactivating mutation in the Pstpip2 gene. We investigated how the genetic background of BALB/c, C57BL/6J, and C57BL/6NCrl strains alters the molecular mechanisms controlling disease progression. While all mice developed the disease, symptoms were significantly milder in BALB/c and partially also in C57BL/6J when compared to C57BL/6NCrl. Disease severity correlated with the number of infiltrating neutrophils and monocytes and with the production of chemokines attracting these cells to the site of inflammation. It also correlated with increased expression of genes associated with autoinflammation, rheumatoid arthritis, neutrophil activation, and degranulation, resulting in altered neutrophil activation in vivo. Together, our data demonstrate striking effects of genetic background on multiple parameters of neutrophil function and activity influencing the onset and course of the CMO disease.

A large-scale in vivo screen to investigate the roles of human genes in Drosophila melanogaster.

Understanding the signaling pathways in which genes participate is essential for discovering the etiology of diseases in humans. The model organism, Drosophila melanogaster, has been crucial in understanding the signaling pathways in humans, given the evolutionary conservation of a significant number of genes between the two species. Genetic screens using Drosophila are a useful way of testing large number of genes to study their function and roles within signaling pathways. We conducted a large-scale genetic screen to identify which human genes cause an alteration in the morphology of the Drosophila eye. The GMR-Gal4 was employed to activate a single UAS-human gene in the eye tissue. In total, we screened 802 UAS-human gene stocks, corresponding to 787 human protein-coding genes, for the ability to influence eye development. We found that overexpression of 64 human genes were capable of disrupting eye development, as determined by phenotypic changes in eye texture, size, shape, bristle morphology, and ommatidia organization. Subsequent analysis revealed that the fly genome encodes proteins that are homologous to a majority of the 64 human genes, raising the possibility that overexpression of these transgenes altered eye development by altering the activity of evolutionarily conserved developmental signaling pathways. Consistent with this hypothesis, a secondary screen demonstrated that overexpression of fly homologs produced phenotypes that mimicked those produced by overexpression of the human gene. Our screening has identified 64 human genes capable of inducing phenotypes in the fly, offering a foundation for ongoing research aimed at understanding functionally conserved pathways across species.

Rapid phenotypic and genotypic change in a laboratory schistosome population.

Genomic analysis has revealed extensive contamination among laboratory-maintained microbes including malaria parasites, Mycobacterium tuberculosis and Salmonella spp. Here, we provide direct evidence for recent contamination of a laboratory schistosome parasite population, and we investigate its genomic consequences. The Brazilian Schistosoma mansoni population SmBRE has several distinctive phenotypes, showing poor infectivity, reduced sporocysts number, low levels of cercarial shedding and low virulence in the intermediate snail host, and low worm burden and low fecundity in the vertebrate rodent host. In 2021 we observed a rapid change in SmBRE parasite phenotypes, with a ~10x increase in cercarial production and ~4x increase in worm burden. To determine the underlying genomic cause of these changes, we sequenced pools of SmBRE adults collected during parasite maintenance between 2015 and 2023. We also sequenced another parasite population (SmLE) maintained alongside SmBRE without phenotypic changes. While SmLE allele frequencies remained stable over the eight-year period, we observed sudden changes in allele frequency across the genome in SmBRE between July 2021 and February 2023, consistent with expectations of laboratory contamination. (i) SmLE-specific alleles rose in the SmBRE population from 0 to 41-46% across the genome between September and October 2021, documenting the timing and magnitude of the contamination event. (ii) After contamination, strong selection (s = ~0.23) drove replacement of low fitness SmBRE with high fitness SmLE alleles. (iii) Allele frequency changed rapidly across the whole genome, except for a region on chromosome 4 where SmBRE alleles remained at high frequency. We were able to detect contamination in this case because SmBRE shows distinctive phenotypes. However, this would likely have been missed with phenotypically similar parasites. These results provide a cautionary tale about the importance of tracking the identity of parasite populations, but also showcase a simple approach to monitor changes within populations using molecular profiling of pooled population samples to characterize fixed single nucleotide polymorphisms. We also show that genetic drift results in continuous change even in the absence of contamination, causing parasites maintained in different labs (or sampled from the same lab at different times) to diverge.

The protective mechanism of sevoflurane in pulmonary arterial hypertension via downregulation of TRAF6

Pulmonary arterial hypertension (PAH) is an obstructive vasculopathy that, if not promptly treated, culminates in right heart failure. Therefore, pre-clinical studies are needed to support and optimize therapeutic approaches of PAH. Here, we explore a prospective function of sevoflurane in experimental PAH through regulating TRAF6. Monocrotaline (MCT)-induced PAH rats were subjected to sevoflurane inhalation and intratracheal instillation of lentivirus overexpressing TRAF6. Platelet-derived growth factor (PDGF)-treated pulmonary artery smooth muscle cells (PASMCs) were exposed to sevoflurane and genetically manipulated for TRAF6 overexpression. It was found that MCT and PDGF challenge upregulated the levels of TRAF6 in rat lung tissues and PASMCs, but sevoflurane treatment led to reduced TRAF6 expression. Sevoflurane inhalation in MCT-induced rats resulted in alleviative pulmonary vascular remodeling, mitigated right ventricular dysfunction and hypertrophy, improved mitochondrial function and dynamics, and inactivation of NF-κB pathway. In vitro studies confirmed that exposure to sevoflurane repressed PDGF-induced proliferation, migration, and phenotype switching of PASMCs, and suppressed mitochondrial dysfunction and NF-κB activation in PDGF-stimulated PASMCs. The beneficial impact of sevoflurane on pathological changes of lung and cell phenotype of PASMCs were reversed by overexpression of TRAF6. In summary, our study suggested the protective properties of sevoflurane in targeting PAH by downregulating TRAF6 expression, providing a novel avenue for the management of PAH.

EnvZ/OmpR Controls Protein Expression and Modifications in Cronobacter sakazakii for Virulence and Environmental Resilience.

Cronobacter sakazakii is a notorious foodborne opportunistic pathogen, particularly affecting vulnerable populations such as premature infants, and poses significant public health challenges. This study aimed to elucidate the role of the envZ/ompR genes in environmental tolerance, pathogenicity, and protein regulation of C. sakazakii. An envZ/ompR knockout mutant was constructed and assessed for its impact on bacterial growth, virulence, environmental tolerance, and protein regulation. Results demonstrate that deletion of envZ/ompR genes leads to reduced growth rate and attenuated virulence in animal models. Additionally, the knockout strain exhibited compromised environmental tolerance, particularly in desiccation and oxidative stress conditions, along with impaired adhesion and invasion abilities in epithelial cells. Proteomic analysis revealed significant alterations in protein expression and phosphorylation patterns, highlighting potential compensatory mechanisms triggered by gene deletion. Furthermore, investigation into protein deamidation and glucose metabolism uncovered a link between envZ/ompR deletion and energy metabolism dysregulation. Interestingly, the downregulation of MalK and GrxC proteins was identified as contributing factors to altered desiccation tolerance and disrupted redox homeostasis, respectively, providing mechanistic insights into the phenotypic changes observed. Overall, this study enhances understanding of the multifaceted roles of envZ/ompR in C. sakazakii physiology and pathogenesis, shedding light on potential targets for therapeutic intervention and food safety strategies.

Microevolutionary hypothesis of the obesity epidemic.

The obesity epidemic represents potentially the largest phenotypic change in Homo sapiens since the origin of the species. Despite obesity's high heritability, it is generally presumed a change in the gene pool could not have caused the obesity epidemic. Here we advance the hypothesis that a rapid change in the obesogenic gene pool has occurred second to the introduction of modern obstetrics dramatically altering evolutionary pressures on obesity-the microevolutionary hypothesis of the obesity epidemic. Obesity is known to increase childbirth-related mortality several fold. Prior to modern obstetrics, childbirth-related mortality occurred in over 10% of women in their lifetime. After modern obstetrics, this mortality reduced to a fraction of a percent, thereby lifting a strong negative selection pressure. Regression analysis of data for ~ 190 countries was carried out to examine associations between 1990 lifetime maternal death rates (LMDR) and current obesity rates. Multivariate regression showed LMDR correlated more strongly with national obesity rates than GDP, calorie intake and physical inactivity. Analyses controlling for confounders via partial correlation show that LMDR explains approximately 11% of the variability of obesity rate between nations. For nations with LMDR above the median (>0.45%), LMDR explains 33% of obesity variance, while calorie intake, GDP and physical inactivity show no association with obesity in these nations. The microevolutionary hypothesis offers a parsimonious explanation of the global nature of the obesity epidemic.

Endothelin-1 (ET-1) contributes to senescence and phenotypic changes in brain pericytes in diabetes-mimicking conditions.

Diabetes mediates endothelial dysfunction and increases the risk of Alzheimer's disease and related dementias. Diabetes also dysregulates the ET system. ET-1-mediated constriction of brain microvascular pericytes (BMVPCs) has been shown to contribute to brain hypoperfusion. Cellular senescence, a process that arrests the proliferation of harmful cells and instigates phenotypical changes and proinflammatory responses in endothelial cells that impact their survival and function. Thus, we hypothesized that ET-1 mediates BMVPC senescence and phenotypical changes in diabetes-like conditions. Human BMVPCs were incubated in diabetes-like conditions with or without ET-1 (1 µmol/L) for 3 and 7 days. Hydrogen peroxide (100 µmol/L H2O2) was used as a positive control for senescence and to mimic ischemic conditions. Cells were stained for senescence-associated β-galactosidase or processed for immunoblotting and quantitative real-time PCR analyses. In additional experiments, cells were stimulated with ET-1 in the presence or absence of ETA receptor antagonist BQ-123 (20 μmol/L) or ETB receptor antagonist BQ-788 (20 μmol/L). ET-1 stimulation increased β-galactosidase accumulation which was prevented by BQ-123. ET-1 also increased traditional senescence marker p16 protein and pericyte-specific senescence markers, TGFB1i1, PP1CA, and IGFBP7. Furthermore, ET-1 stimulated contractile protein α-SMA and microglial marker ostepontin in high glucose suggesting a shift toward an ensheathing or microglia-like phenotype. In conclusion, ET-1 triggers senescence, alters ETA and ETB receptors, and causes phenotypical changes in BMVPCs under diabetes-like conditions. These in vitro findings need to be further studied in vivo to establish the role of ETA receptors in the progression of pericyte senescence and phenotypical changes in VCID.

Genetic analysis of platelet-derived growth factor receptor-like gene (PDGFRL) polymorphism and melanin traits in Lanping black-boned sheep (Ovis aries)

Abstract. In the intricate tapestry of Yunnan Province's biodiversity, the Lanping black-boned sheep (LPBB) emerges as a captivating enigma, distinguished by its profound melanin pigmentation adorning both its skin and its internal organs. Initially cataloged in the 1950s within the confines of Lanping County, this exceptional mammalian species presents a scarcity and uniqueness that extends beyond its geographic origins. Here, we collected 100 blood samples from Lanping black-boned sheep along with 50 samples each from Lanping normal sheep (LPN) and Huize normal sheep (HZN), all sourced from Yunnan Province. Our investigation focused on the association between the platelet-derived growth factor receptor-like gene (PDGFRL) polymorphism and the distinctive melanin characteristics observed in Lanping black-boned sheep. Utilizing UV–visible spectrophotometry, we assessed the melanin indexes present, such as tyrosinase activity and true melanin in the sheep blood, and the results demonstrated a significant elevation in melanin indexes for Lanping black-boned sheep compared to the control group (P&lt;0.05). We also identified three synonymous mutation sites within a partial 1128 bp exon fragment of the gene-encoding PDGFRL (EX2-G408A, EX5-T184C, and EX5-G222T). Notably, Lanping black-boned sheep, harboring genotypes GG, TT, and GG at these specific sites, showcased a pronounced surge in tyrosinase activity, eumelanin / total melanin ratios, and plasma colorimetric values when contrasted with the control group (P&lt;0.05). The discernment of GG, TT, and GG as the prevailing genotypes at their respective genetic loci in Lanping black-boned sheep heralds a breakthrough in our understanding of the genetic markers associated with black pigmentation. However, all three loci are silent mutations and do not alter the phenotypic changes. Whether they affect changes in melanin content through other metabolic pathways requires further study. In conclusion, the PDGFRL gene was silenced by mutations in our study and affected blood melanin levels. However, the gene did not undergo a missense mutation that altered the phenotypic changes, and the exact channel through which the changes in melanin content were affected needs to be further verified.

Hif-2α programs oxygen chemosensitivity in chromaffin cells.

The study of transcription factors that determine specialized neuronal functions has provided invaluable insights into the physiology of the nervous system. Peripheral chemoreceptors are neurone-like electrophysiologically excitable cells that link the oxygen concentration of arterial blood to the neuronal control of breathing. In the adult, this oxygen chemosensitivity is exemplified by type I cells of the carotid body, and recent work has revealed one isoform of the hypoxia-inducible transcription factor (HIF), HIF-2α, as having a nonredundant role in the development and function of that organ. Here, we show that activation of HIF-2α, including isolated overexpression of HIF-2α but not HIF-1α, is sufficient to induce oxygen chemosensitivity in adult adrenal medulla. This phenotypic change in the adrenal medulla was associated with retention of extra-adrenal paraganglioma-like tissues resembling the fetal organ of Zuckerkandl, which also manifests oxygen chemosensitivity. Acquisition of chemosensitivity was associated with changes in the adrenal medullary expression of gene classes that are ordinarily characteristic of the carotid body, including G protein regulators and atypical subunits of mitochondrial cytochrome oxidase. Overall, the findings suggest that, at least in certain tissues, HIF-2α acts as a phenotypic driver for cells that display oxygen chemosensitivity, thus linking 2 major oxygen-sensing systems.

Features of the neutrophil granulocyte phenotype in children with infectious mononucleosis

The course of infectious diseases caused by viruses, and their common outcome is determined by the activity of the inflammatory reaction which occurs both at the local and systemic levels. However, the features of neutrophil functions during inflammatory reaction are virtually unknown in the patients with infectious mononucleosis (IM), caused by Epstein–Barr virus (EBV). Hence, the aim of our study was to evaluate some characteristics of phenotypic spectrum of blood neutrophils in children with IM. Patients and methods. We examined 84 children aged 3 to 11 years with EBV infection with moderate or severe clinical course of the disease. All patients exhibited a positive test for EBV DNA in blood lymphocytes and appropriate serological markers of acute EBV infection. The control group consisted of 40 conditionally healthy children at the similar age range. The study of neutrophil phenotype was carried out by flow cytometry using direct immunofluorescence of whole peripheral blood samples. A study of the neutrophil phenotype with a combination of two functional antigens (CD64 and CD32) has revealed that in children with IM, regardless of age, the main fraction of blood neutrophils are double-negative cells, whereas in healthy children it consists of CD64-CD32+ neutrophils. The main fraction of neutrophils in the paired combination of CD64 and CD11b antigens in sick children aged 3-6 and 7-11 years was similar to the healthy controls (CD64-CD11b+), but with a change in the content of minor cell fractions. The number of CD64-CD15+ neutrophils (main fraction of cells in healthy children) proved to be significantly reduced in the IM patients of both age groups. However, we have revealed a marked increase in the level of double-negative cells for the CD64 and CD15 antigens. At the same time, the content of double-negative neutrophils for these markers was also increased in IM children of both age groups. The cells with CD11b-CD15+ and CD11b+CD15+ phenotypes comprised the main fractions in IM, as studied by a paired combination of CD11b and CD15 antigens; in healthy children – only CD11b+CD15+ neutrophils are detected. The phenotypic changes of neutrophils during IM suggest a decreased migratory ability of cells with high activity of proinflammatory functions. The established ontogenetic features of the neutrophil phenotype are significantly changed in the children with IM, probably, due to specific immunopathogenesis of the viral infection. The detected changes in phenotypic composition of neutrophils associated with IM may be caused both by the features of protective reaction of innate immune cells and pathogenic effects of the virus itself upon blood neutrophils.