Expression Of Factor Research Articles

Abstract 890: DNA deaminase APOBEC3B regulates theresponse to PARP inhibitors of epithelial ovarian cancer stem cells

Abstract The most aggressive types of ovarian cancer is high-grade serous ovarian cancer (HGSOC), and the 5-year survival is less than 20%. Relapsed HGSOC patients often have enrichment of cancer stem cells (CSCs) with enhanced pro-survival and self-renewal capacity, suggesting a potential vulnerable population for effective targeted therapies with less toxicity. APOBEC3 proteins are a family of primate-specific DNA deaminases that catalyze cytidine to uracil (C-to-U) on single-strand DNA thereby introducing C-to-T/G point mutations. This important DNA modification is abundant in a wide array of cancer types including HGSOCs. However, the prevalence and significance of A3-induced DNA deamination in HGSOC CSC is currently unknown. We recently discovered that CSCs maintain a low A3-induced mutation burden. Using patient-sample derived spheroid culture and HGSOC cell lines, we found A3B was the only A3 members differentially expressed between CSC and non-CSC and the expression was significantly reduced in CSCs. In addition, inhibition of A3B leads to elevated frequency of CSCs and enhanced expression of the stemness factors. Important, our data suggest that DNA deaminase activity of A3B leads to induction of DNA damage thereby sensitizing HGSOC CSCs to PARPi. These novel insights suggest that DNA deamination plays a fundamental role in CSC biology and also raises the possibility that A3B activation may sensitize HGSOC CSCs to PARPi by promoting DNA damages. Citation Format: Jane isquith, Jessica Pham, Robert Phavong, Maria Rivera, Kathleen Fisch, Catriona Jamieson, Michael McHale, Ramez Eskander, Qingfei Jiang. DNA deaminase APOBEC3B regulates theresponse to PARP inhibitors of epithelial ovarian cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 890.

Abstract 3682: Quadruple negative breast tumors in African American women express factors associated with worse prognosis compared to triple negative tumors

Abstract Introduction: Quadruple negative breast cancer (QNBC) is defined by the absence of androgen (AR), estrogen (ER), progesterone (PR) receptors and human epidermal growth factor (HER2) expression. This subset of triple negative breast cancer (TNBC) has been described as unique to African American (AA) women and represents about 67-90% of all TNBC. Yet, there are limited clinical and therapeutic studies focused on this new subset of breast cancer (BCa) that is highly aggressive and portends a worse survival compared to AR positive Bca subtypes. The objectives of our study were to 1) determine the molecular and clinical characteristics of QNBC and compare them with other BCa subtypes and 2) determine the association of QNBC with clinicopathological factors and prognostic markers. Materials and Methods: Tissue arrays were constructed from FFPE tumor blocks of invasive ductal breast carcinomas in 202 AAs, diagnosed at Howard University from 2000 to 2010. Two separate tissue cores of IDC represented each surgical case in the TMA and was linked to an Institutional Review Board-approved database of demographic and clinical data. Using a microtome, 5-µm sections were cut from the TMA blocks and mounted onto Superfrost Plus microscope slides. Sections were stained with a mouse monoclonal antibody against AR (Clone AR 441, DAKO). Immunohistochemistry for AR was considered positive if ≥10% of tumor cells showed nuclear staining. Bivariate analysis was performed via χ2 analysis and survival data was calculated via Kaplan-Meier curves (SPSS v28.0.0). Results: The most prevalent breast cancer subtype was luminal A (ER+ or PR+, HER2−; 43.5%), followed by TNBC (33.7%). Of the 67 TNBCs on the array, 63 had AR data; 33 (52%) of the TNBCs were AR negative. Greater than 70% of all tumors were stage I and II; and Grade 3 comprised 67.4% of the tumors. QNBC tumors were more likely to be higher grade (p<0.001) and larger in size (p=0.05) compared to other subtypes. There was also a trend for increased metastases in QNBCs (Luminal and HER2 tumors=11.2%; TNBC=13.3%; AR negative= 18.8%; and QNBC=20.8% (p=0.15)). Compared to TNBCs, QNBCs were larger (p=0.048), more likely to express markers associated invasion (Fascin, p=0.004) and reduced survival (Vimentin, p=0.014). There was a trend for increased expression of markers associated with metastases: Mammaglobin (expressed in 30.8% of TNBCs compared to 52.1% of QNBCs; p=0.17) and CD105 (expressed in 6.7% of TNBCs compared to 27.1% of QNBCs; p=0.097). There was no association between QNBC and stage, recurrence-free survival, or overall survival. Conclusion: In our study of AAs, a statistically significant association was noted between QNBC tumors and poorer prognosis. The high prevalence of AR negativity of TNBCs in AAs could explain observed worse outcomes and supports the existence of a unique subtype of Bca known as QNBC, worthy of scientific attention. Citation Format: Oluwadamilola Oladeru, Yasmine Kanaan, Tammy Naab, Luisel J. Ricks-Santi. Quadruple negative breast tumors in African American women express factors associated with worse prognosis compared to triple negative tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3682.

Proteomic Profiling Identifies Co-Regulated Expression of Splicing Factors as a Characteristic Feature of Intravenous Leiomyomatosis.

Simple SummaryIntravenous leiomyomatosis is a rare form of smooth muscle tumour that has unique and distinct clinical features including growth within the uterine and pelvic veins. Here we use proteomics by mass spectrometry to show that this disease is distinct from uterine leiomyomas and other benign smooth muscle tumours due to the enrichment of components of the spliceosome machinery. In particular, we find that intravenous leiomyomatosis tumours harbour co-regulated expression of multiple splicing factors that are associated with biological processes including cell signalling. Our study demonstrates that intravenous leiomyomatosis is a distinct disease from other smooth muscle tumours and indicates a possible functional role for alternative splicing in disease initiation and progression.Intravenous leiomyomatosis (IVLM) is a rare benign smooth muscle tumour that is characterised by intravenous growth in the uterine and pelvic veins. Previous DNA copy number and transcriptomic studies have shown that IVLM harbors unique genomic and transcriptomic alterations when compared to uterine leiomyoma (uLM), which may account for their distinct clinical behaviour. Here we undertake the first comparative proteomic analysis of IVLM and other smooth muscle tumours (comprising uLM, soft tissue leiomyoma and benign metastasizing leiomyoma) utilising data-independent acquisition mass spectrometry. We show that, at the protein level, IVLM is defined by the unique co-regulated expression of splicing factors. In particular, IVLM is enriched in two clusters composed of co-regulated proteins from the hnRNP, LSm, SR and Sm classes of the spliceosome complex. One of these clusters (Cluster 3) is associated with key biological processes including nascent protein translocation and cell signalling by small GTPases. Taken together, our study provides evidence of co-regulated expression of splicing factors in IVLM compared to other smooth muscle tumours, which suggests a possible role for alternative splicing in the pathogenesis of IVLM.

The Specifically Androgen-Regulated Gene (SARG) Promotes Papillary Thyroid Carcinoma (PTC) Lymphatic Metastasis Through Vascular Endothelial Growth Factor C (VEGF-C) and VEGF Receptor 3 (VEGFR-3) Axis.

The papillary thyroid carcinoma (PTC) metastasizes through lymphatic spread, but the follicular thyroid cancer (FTC) metastasis occurs by following hematogenous spread. To date, the molecular mechanism underlying different metastatic routes between PTC and FTC is still unclear. Here, we showed that specifically androgen-regulated gene (SARG) was significantly up-regulated in PTC, while obviously down-regulated in FTC through analyzing the Gene Expression Omnibus (GEO) database. Immunohistochemistry assay verified that the PTC lymph node metastasis was associated with higher levels of SARG protein in clinical PTC patient samples. SARG-knockdown decreased TPC-1 and CGTH-W3 cells viability and migration significantly. On the contrary, SARG-overexpressed PTC cells possessed more aggressive migratory ability and viability. In vivo, SARG overexpression dramatically promoted popliteal lymph node metastasis of xenografts from TPC-1 cells mouse footpad transplanting. Mechanistically, SARG overexpression and knockdown significantly increased and decreased the expression of vascular endothelial growth factor C (VEGF-C) and VEGF receptor 3 (VEGFR-3), respectively, thereby facilitating or inhibiting the tube formation in HUVECs. The tube formation experiment showed that SARG overexpression and knockdown promoted or inhibited the number of tube formations in HUVEC cells, respectively. Taken together, we showed for the first time the differential expression profile of SARG between PTC and FTC, and SARG promotes PTC lymphatic metastasis via VEGF-C/VEGFR-3 signal. It indicates that SARG may represent a target for clinical intervention in lymphatic metastasis of PTC.

Modulatory Effects of Estradiol and Its Mixtures with Ligands of GPER and PPAR on MAPK and PI3K/Akt Signaling Pathways and Tumorigenic Factors in Mouse Testis Explants and Mouse Tumor Leydig Cells.

The present study was designed to evaluate how estradiol alone or in combination with G protein-coupled estrogen receptor (GPER) agonists and GPER and peroxisome proliferator-activated receptor (PPAR) antagonists alter the expression of tumor growth factor β (TGF-β), cyclooxygenase-2 (COX-2), hypoxia inducible factor 1-alpha (HIF-1α), and vascular endothelial growth factor (VEGF) in mouse testis explants and MA-10 mouse tumor Leydig cells. In order to define the hormone-associated signaling pathway, the expression of MAPK and PI3K/Akt was also examined. Tissue explants and cells were treated with estradiol as well as GPER agonist (ICI 182,780), GPER antagonist (G-15), PPARα antagonist (GW6471), and PPARγ antagonist (T00709072) in various combinations. First, we showed that in testis explants GPER and PPARα expressions were activated by the GPER agonist and estradiol (either alone or in mixtures), whereas PPARγ expression was activated only by GPER agonist. Second, increased TGF-β expression and decreased COX-2 expression were found in all experimental groups of testicular explants and MA-10 cells, except for up-regulated COX-2 expression in estradiol-treated cells, compared to respective controls. Third, estradiol treatment led to elevated expression of HIF-1α and VEGF, while their lower levels versus control were noted in the remaining groups of explants. Finally, we demonstrated the up-regulation of MAPK and PI3Kp85/Akt expressions in estradiol-treated groups of both ex vivo and in vitro models, whereas estradiol in mixtures with compounds of agonistic or antagonistic properties either up-regulated or down-regulated signaling kinase expression levels. Our results suggest that a balanced estrogen level and its action together with proper GPER and PPAR signaling play a key role in the maintenance of testis homeostasis. Moreover, changes in TGF-β and COX-2 expressions (that disrupted estrogen pathway) as well as disturbed GPER-PPAR signaling observed after estradiol treatment may be involved in testicular tumorigenesis.

“Near Cure” treatment of severe acute EAE in MIF-1-deficient female and male mice with a bifunctional MHCII-derived molecular construct

Our previous studies demonstrated increased serum levels of macrophage migration inhibitory factor (MIF-1) and its homologue, MIF-2, in males during MS progression; and that genetically high-MIF-expressing male subjects with relapsing multiple sclerosis (MS) had a significantly greater risk of conversion to progressive MS than lower-MIF-expressing males and females. However, female MS subjects with severe disease expressed higher levels of CD74, the common MIF-1/MIF-2 receptor, on blood cells. In the murine model of MS, experimental autoimmune encephalomyelitis (EAE), both male and female mice lacking MIF-1 and/or MIF-2 were clinically improved during development of moderately severe disease, thus implicating both homologs as co-pathogenic contributors. The current study using MIF-deficient mice with severe acute EAE revealed a highly significant reduction of EAE scores in MIF-1-deficient females, in contrast to only minor and delayed reduction of clinical signs in MIF-1-deficient males. However, clinical EAE scores and factor expression were strongly suppressed in males and further reduced in females after treatment of WT and MIF-1-, MIF-2- and MIF-1/2-DUAL-deficient female and male mice with a MHCII DRα1-MOG-35-55 molecular construct that competitively inhibits MIF-1 & MIF-2 signaling through CD74 as well as T cell activation. These results suggest sex-dependent differences in which the absence of the MIF-1 and/or MIF-2 genotypes may permit stronger compensatory CD74-dependent EAE-inducing responses in males than in females. However, EAE severity in both sexes could still be reduced nearly to background (a “near cure”) with DRα1-MOG-35-55 blockade of compensatory MIF and CD74-dependent factors known to attract peripheral inflammatory cells into the spinal cord tissue.

Effect of rottlerin on astrocyte phenotype polarization after trimethyltin insult in the dentate gyrus of mice

BackgroundIt has been demonstrated that reactive astrocytes can be polarized into pro-inflammatory A1 phenotype or anti-inflammatory A2 phenotype under neurotoxic and neurodegenerative conditions. Microglia have been suggested to play a critical role in astrocyte phenotype polarization by releasing pro- and anti-inflammatory mediators. In this study, we examined whether trimethyltin (TMT) insult can induce astrocyte polarization in the dentate gyrus of mice, and whether protein kinase Cδ (PKCδ) plays a role in TMT-induced astrocyte phenotype polarization.MethodsMale C57BL/6 N mice received TMT (2.6 mg/kg, i.p.), and temporal changes in the mRNA expression of A1 and A2 phenotype markers were evaluated in the hippocampus. In addition, temporal and spatial changes in the protein expression of C3, S100A10, Iba-1, and p-PKCδ were examined in the dentate gyrus. Rottlerin (5 mg/kg, i.p. × 5 at 12-h intervals) was administered 3–5 days after TMT treatment, and the expression of A1 and A2 transcripts, p-PKCδ, Iba-1, C3, S100A10, and C1q was evaluated 6 days after TMT treatment.ResultsTMT treatment significantly increased the mRNA expression of A1 and A2 phenotype markers, and the increased expression of A1 markers remained longer than that of A2 markers. The immunoreactivity of the representative A1 phenotype marker, C3 and A2 phenotype marker, S100A10 peaked 6 days after TMT insult in the dentate gyrus. While C3 was expressed evenly throughout the dentate gyrus, S100A10 was highly expressed in the hilus and inner molecular layer. In addition, TMT insult induced microglial p-PKCδ expression. Treatment with rottlerin, a PKCδ inhibitor, decreased Iba-1 and C3 expression, but did not affect S100A10 expression, suggesting that PKCδ inhibition attenuates microglial activation and A1 astrocyte phenotype polarization. Consistently, rottlerin significantly reduced the expression of C1q and tumor necrosis factor-α (TNFα), which has been suggested to be released by activated microglia and induce A1 astrocyte polarization.ConclusionWe demonstrated the temporal and spatial profiles of astrocyte polarization after TMT insult in the dentate gyrus of mice. Taken together, our results suggest that PKCδ plays a role in inducing A1 astrocyte polarization by promoting microglial activation and consequently increasing the expression of pro-inflammatory mediators after TMT insult.

Pharmacological Targeting of Senescence with ABT-263 in Experimental Heart Failure with Preserved Ejection Fraction

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Fundação Ciência e Tecnlogia HFpEF is the most common form of heart disease in the elderly and is associated with high morbidity and mortality. Our understanding of HFpEF pathophysiology is limited and development of efficient therapies that alter the clinical course of the disease has proved greatly challenging. Although aging is a risk factor of HFpEF1, the involvement of aging hallmarks such as cell senescent and immunosenescence is unclear. Recently, a link between endothelial senescence and HFpEF development was demonstrated in mice with accelerated aging [1], endorsing anti-aging pharmacologic as potential new therapeutic alternatives for HFpEF. In fact, the senolytic (drugs that selectively promote apoptosis of senescent cells) ABT-263 has already proven efficacy in the context of cardiovascular diseases. Further studies are needed to clarify the relevance of aging and of anti-aging therapies in the context of HFpEF. Using ZSF1 obese rats (ZSF1-Ob) as model of HFpEF we showed from 18 weeks signs of immunosenescence compared to ZSF1-Ln, namely an increased frequency of circulating myeloid cells and decreased frequency of T and B cells. Concomitantly, expression of pro-inflammatory factors (IL-6, IL-1, TGF-β, TNF-α) was upregulated in peripheral blood mononuclear cells (PBMCs) of ZSF1-Ob which also displayed characteristic features of cell senescence (p21 expression, lysosomal endogenous Beta-galactosidase (SA-B-Gal) senescence-associated secretory phenotype (SASP) and up-regulation of BCL-XL. Importantly, no signs of cellular senescence (SA-B-gal, pH2AX and senescence associated pathways) were found in the main hematopoietic organs (spleen and bone marrow). Alongside, these systemic alterations, an upsurge of cellular senescence was observed in myocardium of ZSF1-Ob rats, particularly in endothelial and hematopoietic cells. Serum of ZSF1-Ob rat was able to induce activation and cellular senescence of cardiac microvascular endothelial cells, indicating that systemic circulating factors may be the upstream mechanism of cellular senescence and dysfunction in HFpEF. Analysis of HFpEF patients and a control cohort adjusted to main co-morbidities further demonstrated accumulation of senescent monocytes in HFpEF patients. In these patients, the senescence marker SA-B-Gal correlated with plasmatic brain natriuretic peptide (BNP) levels and pulmonary artery systolic pressure. Targeting aging hallmarks through the treatment of ZSF1-Ob rat with ABT-263 resulted in a reduction in circulating senescent cells, decreased systemic and local inflammation, re-established immune proportions, restore the levels of circulating BNP and attenuated myocardial remodeling, particularly endothelial dysfunction and fibrosis. Collectively these findings support that premature cellular senescence contributes to the establishment of a deleterious pro-inflammatory environment in HFpEF and that senolytic agents hold promise for the treatment of this syndrome.

Sorafenib inhibits LPS-induced inflammation by regulating Lyn-MAPK-NF-kB/AP-1 pathway and TLR4 expression

Sorafenib is an anti-tumor drug widely used in clinical treatment, which can inhibit tyrosine kinase receptor on cell surface and serine/threonine kinase in downstream Ras/MAPK cascade signaling pathway of cells. Tyrosine kinase phosphorylation plays an important role in inflammatory mechanism, such as TLR4 tyrosine phosphorylation, MAPK pathway protein activation, and activation of downstream NF-кB. However, the effects of sorafenib on LPS-induced inflammatory reaction and its specific mechanism have still remained unknown. We found that sorafenib inhibited the phosphorylation of tyrosine kinase Lyn induced by LPS, thereby reducing the phosphorylation level of p38 and JNK, inhibiting the activation of c-Jun and NF-κB, and then inhibiting the expression of inflammatory factors IL-6, IL-1β, and TNF-α. Furthermore, sorafenib also decreased the expression of TLR4 on the macrophage membrane to inhibit the expression of inflammatory factors latterly, which may be related to the inactivation of Lyn. These results provide a new perspective and direction for the clinical treatment of sepsis.

Antifungal activity and potential mechanism of berberine hydrochloride against fluconazole-resistant Candida albicans.

Introduction. The emergence of resistance to fluconazole in Candida albicans has made the clinical treatment of this microbe difficult. A potential strategy to address this problem involves diminishing fungal resistance to antimicrobial drugs.Hypothesis. Berberine hydrochloride (BH), the primary active ingredient of the traditional Chinese medicine (TCM) Coptis, inhibits the growth of fluconazole-resistant C. albicans through its action on the high-osmolarity glycerol mitogen-activated protein kinase (HOG-MAPK) pathway.Aim. To examine the effect of BH on the HOG-MAPK pathway to assess the potential molecular mechanism by which BH inhibits fluconazole-resistant C. albicans.Methodology. The minimum inhibitory concentration (MIC) of BH to fluconazole-resistant C. albicans was measured using the broth microdilution approach to determine the concentration of effective drug intervention. Changes in physiological functions regulated by the HOG-MAPK pathway in response to BH treatment were measured, as well as the expression of central signalling pathway genes and key downstream factors by qRT-PCR and Western blotting, respectively.Results. BH inhibited fluconazole-resistant C. albicans and the sensitivity to fluconazole increased after BH treatment. At a concentration of 256 and 64 μg ml-1 BH may affect key downstream factors that regulate several physiological functions of C. albicans by upregulating the core genes expression of SLN1, SSK2, HOG1, and PBS2 in the HOG-MAPK pathway. Upregulation of GPD1, the key gene for glycerol synthesis, increased cell osmotic pressure. BH treatment increased the accumulation of reactive oxygen species by upregulating the expression of the key respiratory metabolism gene ATP11 and downregulating the expression of the superoxide dismutase gene SOD2. Furthermore, downregulation of mycelial-specific HWP1 hindered the morphological transformation of C. albicans and inhibition of the chitin synthase gene CHS3 and the β-(1,3) glucan synthase gene GSC1 impaired cytoderm integrity.Conclusion. BH affects multiple target genes in diminishing the resistance of C. albicans strains to fluconazole. This effect may be related to the action of BH on the HOG-MAPK pathway.

Effects of Long-Term Exposure to PM2.5 on Oxidative Stress Injury and Expression of Inflammatory Factors, NF-κB p65 and Cx43 in Bone Marrow of Mice

Objective: This study aims to explore the toxic effect of PM2.5 on the hematopoietic microenvironment of the bone marrow, and investigate the effect of PM2.5 on oxidative stress injury, the secretion of inflammatory cytokines and the expression of NF-κB p65 and C x 43 in the bone marrow of mice.Methods: A total of 615 mice were treated with normal saline, low dose (0.1 mg/ml) PM2.5 suspension, and high dose (0.2 mg/ml) PM2.5 suspension by intratracheal instillation, respectively. The ROS content, activity of total SOD (T-SOD), DNA injury, and the protein expression levels of IL-1 β, IL-6, TNF-α, NF-κB p65 and Cx43 in bone marrow cells (BMCs) of mice were detected in these three groups.Results: Compared with the normal saline control group, the expression level of reactive oxygen species (ROS) significantly increased in BMCs, while the activity of SOD enzymes significantly decreased in PM2.5 exposed mice. Furthermore, DNA injury significantly increased in PM2.5 exposed mice, and the expression of IL-1 β, IL-6, TNF-α and NF-κB p65 significantly increased, while the protein expression of Cx43 significantly decreased in the PM2.5 exposed mice. The differences were statistically significant (p < 0.05). There were significant differences in ROS content, T-SOD activity, DNA injury (TL, TD and TM), protein expression of IL-1β, IL-6, TNF-α and Cx43 between the high-dose PM2.5 exposure group and low-dose PM2.5 exposure group (p < 0.05), but there was no significant difference in the expression of NF-κB p65 between the high-dose and low-dose PM2.5 exposure groups (p > 0.05).Conclusion: PM2.5 has toxic effects on the bone marrow of mice. Oxidative stress injury, inflammatory reaction and abnormal intercellular communication may be the underlying mechanism.

Prenatal hypoxia alters the early ontogeny of dopamine neurons

Dopaminergic (DA) dysfunction is a significant feature in the pathophysiology of schizophrenia. Established developmental risk factors for schizophrenia such as maternal immune activation (MIA) or developmental vitamin D (DVD) deficiency, when modelled in animals, reveal the differentiation of early DA neurons in foetal brains is delayed suggesting this may be a convergent aetiological pathway. Here we have assessed the effects of prenatal hypoxia, another well-known developmental risk factor for schizophrenia, on developing DA systems. Pregnant mice were exposed to a hypoxic environment of 10% oxygen for 48 h from embryonic day 10 (E10) to E12. Embryonic brains were collected and the positioning of mesencephalic cells, expression of DA specification and maturation factors were examined along with the expression of factors that may govern the migration of these neurons. We show that prenatal hypoxia results in a decrease in dopaminergic progenitors retards early DA neuron lateral migration and reduces expression of the receptors known to govern this process. A second time-point, postnatal day 10 (P10) was also examined in order to assess whether prenatal hypoxia alters early presynaptic architecture in the developing striatum. We show reduced expression of tyrosine hydroxylase (TH) in the postnatal striatum along with increases in the density of high-probability DA release sites within TH varicosities. These findings add to the emerging literature showing that multiple epidemiologically validated environmental risk factors for schizophrenia may induce early alterations to develop DA systems. This may represent a possible convergent mechanism in the onset of presynaptic DA dysfunction in patients.

The role of hypoxia in the pathophysiology of chronic rhinosinusitis.

Chronic rhinosinusitis (CRS) is a chronic inflammatory disease of the nasal cavity characterized by excessive nasal mucus secretion and nasal congestion. The development of CRS is related to pathological mechanisms induced by hypoxia. Under hypoxic conditions, the stable expression of both Hypoxia inducible factor-1 (HIF-1) α and HIF-2α are involved in the immune response and inflammatory pathways of CRS. The imbalance in the composition of nasal microbiota may affect the hypoxic state of CRS and perpetuate existing inflammation. Hypoxia affects the differentiation of nasal epithelial cells such as ciliated cells and goblet cells, induces fibroblast proliferation, and leads to epithelial-mesenchymal transition (EMT) and tissue remodeling. Hypoxia also affects the proliferation and differentiation of macrophages, eosinophils, basophils, and mast cells in sinonasal mucosa, and thus influences the inflammatory state of CRS by regulating T cells and B cells. Given the multifactorial nature in which HIF is linked to CRS, this study aims to elucidate the effect of hypoxia on the pathogenic mechanisms of CRS.

Maternal nutrient restriction and over-feeding during gestation alter expression of key factors involved in placental development and vascularization.

Poor maternal nutrition can negatively affect fetal and placental growth and development. However, the mechanism(s) that contribute to altered placenta growth and function are not well understood. We hypothesized that poor maternal diet would impact signaling through the C-X-C motif chemokine ligand (CXCL) 12-CXCL4 axis and/or placental expression of the insulin-like growth factor (IGF) axis. Using our established sheep model of poor maternal nutrition, we examined the effects of restricted- and over-feeding on ewe placentome gene and protein expression. Specifically, ewes were fed a control (CON; 100%), restricted (RES; 60%), or over (OVER; 140%) diet beginning at day 30.2 ± 0.02 of gestation, and samples were collected at days 45, 90, and 135 of gestation, representing periods of active placentation, peak placental growth, and near term, respectively. Placentomes were separated into cotyledon and caruncle, and samples snap frozen. Protein was determined by western blot and mRNA expression by real-time PCR. Data were analyzed by ANOVA and significance determined at P ≤ 0.05. Ewes fed a RES diet had decreased CXCL12 and vascular endothelial growth factor (VEGF), and increased tumor necrosis factor (TNF)α protein compared with CON ewes in caruncle at day 45 (P ≤0.05). In day 45 cotyledon, CXCR7 protein was increased and mTOR was decreased in RES relative to CON (P ≤0.05). At day 90, CXCR4 and CXCR7 were reduced in RES caruncle compared with CON, whereas VEGF was reduced and mTOR increased in cotyledon of RES ewes relative to CON (P ≤0.05). In OVER caruncle, at day 45 CXCR4 and VEGF were reduced and at day 90 CXCR4, CXCR7, and TNFα were reduced in caruncle compared with CON (P ≤0.05). There was no observed effect of OVER diet on protein abundance in the cotyledon (P > 0.05). Expression of IGF-II mRNA was increased in OVER at day 45 and IGFBP-3 was reduced in RES at day 90 in caruncle relative to CON (P ≤0.05). Maternal diet did not alter placentome diameter or weight (P > 0.05). These findings suggest that restricted- and over-feeding negatively impact protein and mRNA expression of key chemokines and growth factors implicated in proper placenta development and function.

Changes in the expression of T-cell factor-1 in follicular helper T cells reflect the condition of systemic lupus erythematosus patients

ObjectiveThis study aimed to identify changes in T-cell factor-1 (TCF1) in circulating T-cell subsets of systemic lupus erythematosus (SLE) patients and to explore their significance in SLE. MethodsPeripheral blood was collected from 41 SLE patients and 45 healthy controls (HCs). TCF1 in follicular helper T (TFH), follicular regulatory T (TFR) and regulatory T (Treg) cells was analyzed by flow cytometry. Interleukin-21 (IL-21), programmed death receptor 1 (PD-1) and killer cell lectin-like receptor G1 (KLRG1) were detected and compared among TFH subsets sorted according to TCF1 and CD62L expression. Correlation analyses were conducted between TCF1-related TFH cell subsets and autoantibody levels, systemic lupus erythematosus disease activity index (SLEDAI), and plasmablast levels of SLE patients. ResultsAbsolute numbers of TCF1- TFH and TCF1+ Treg cells were increased in SLE patients. According to TCF1 and CD62L expression, circulating TFH cells and Tregs were sorted into four subsets. CD62L+TCF1+ TFH cell percentages were decreased, and CD62L-TCF1- TFH cells were increased. CD62L+TCF1+ Treg cell percentages were increased, and CD62L-TCF1- Treg cell percentages were decreased. KLRG1+ percentages in CD62L-TCF1-TFH were higher in SLE patients than in HCs. Functionally, CD62L+TCF1+ TFH cells had stronger IL-21 secretion, while CD62L-TCF1-TFH cells had weaker IL-21 secretion and lower PD-1 expression. TCF1- and CD62L-TCF1- TFH numbers were positively correlated with anti-ribosomal P, anti-dsDNA and SLEDAI. ConclusionThe increased TFH cells in SLE patients were mostly TCF1-negative subsets with weakened function. Changes in TCF1-related subsets can reflect the condition and abnormal humoral immunity of SLE patients.

Effects of Electroacupuncture on the Expression of Glial Fibrillary Acidic Protein and Vascular Endothelial Growth Factor and Cell Structure in the Gliavascular Net of Cerebral Ischemic Tissues in Hypertensive Rats

Introduction: This study aims to explore the effects of electroacupuncture on the expression of GFAP and VEGF and cell structure in the glia vascular net of cerebral ischemic tissues in hypertensive rats. Material and Methods: Stroke-prone renovascular hypertensive rat (RHRSP) models and middle cerebral artery occlusion (MCAO) were established. After two hours of ischemia and at 1, 7, 14 and 28 days after reperfusion, the expression of GFAP and VEGF in rat cerebral ischemic tissues was detected by immunohistochemistry and light microscopy. Results: Positive GFAP cells were found in the semidarkness area after two hours of ischemic and one day of reperfusion in the control group, and these increased after 14 days, decreased after 14 days, and significantly decreased after 28 days, when compared to the sham group (P < 0.01). Compared to the control group, GFAP-positive cells significantly increased after 7, 14 and 28 days of ischemic and reperfusion in the electric acupuncture group. Then, this significantly increased and reached the maximum after seven days of reperfusion, but decreased after 14 days. Contrarily, this significantly decreased after 30 day, but remained higher than that in sham-operated group (P < 0.05). Compared to the control group, the number of positive VEGF cells increased after 7, 14 and 28 days of ischemic and reperfusion in the electric acupuncture group (P < 0.01). Conclusions: Electric acupuncture can promote neurofunctional recovery in the cerebral ischemic area. The mechanism may be correlated with the regulation of the expression of GFAP and VEGF, and the rebuilding of the gliavascular net of cerebral ischemic tissues, thereby decreasing cerebral ischemic damage.

Bacterial Pathogens in Food: Modern Safety Criteria and Methods of Molecular Genetic Analysis.

The review is devoted to the issues of improving the food biosafety system by developing and implementing modern methods of microbiological and molecular genetic analysis and new safety standards for the main groups of food products. The results of fundamental and applied microbiological research in the field of food safety conducted by the Federal Research Center for Nutrition and Biotechnology are presented. We demonstrated the necessity of a detailed study of the ecology and features of survival of new types of pathogenic microorganisms and evaluation of the role of technological factors in the formation of altered properties of these pathogens in food production environment. It is noted that the most effective basis for establishing criteria for biosafety of food products is the use of a structural model of microbiological risk analysis (MRA), which implies assessment and integration of risks throughout the food chain and step-by-step study of microbial hazards, predicts the degree of danger to the health of certain foods contaminated by them and the risk and severity of adverse effects. Consideration of the variability of pathogens and the ability of MRA to forecast allows rationalizing resources in the most relevant areas when developing measures to protect the population. It is shown how, as a result of studying the biology of pathogenic bacteria of the genera Campylobacter, Salmonella, Cronobacter, and Listeria and mechanisms for regulating the expression of pathogenicity and resistance factors of these pathogens, improved schemes and modified methods have been developed over a number of years, allowing accelerated indication of pathogens in food products using combined schemes of bacteriological and molecular genetic analysis based on the selection of the most informative biochemical and immunological tests, and their genotyping. Accumulated scientific data in the field of biosafety indicate the need for further research in the prediction of the pathogen behavior in food, studying the mechanisms of regulation of the expression of pathogenicity genes in food-borne pathogens, the transmission of antibiotic resistance genes, and studying the interaction of the human with pathogens to calculate the biological response to microbial agents coming from food.

1447-P: The Snd1 Coregulator Controls ß-Cell Function

The main function of the pancreatic β-cell is glucose-stimulated insulin secretion (GSIS) which serves to maintain whole body glucose homeostasis. Under type 2 diabetic conditions, a subset of β-cells fail and lose expression of key transcription factors (TFs) required for GSIS. Among these TFs is Pdx1, which requires the recruitment of a distinct set of transcriptional coregulators to modulate its activity. Recently, Pdx1 has been shown, through and IP-Mass Spec approach, to interact with numerous coregulators, including Snd1 (Staphylococcal nuclease and Tudor domain-containing protein) , which has been shown to facilitate protein-protein interactions and transcriptional control through the recruitment of chromatin modifying enzymes. Here, we first confirmed Pdx1 and Snd1 interact in the rodent β-cell lines by Co-IP, and in primary mouse β-cells and human EndoC-βH1 β-cell lines using proximity ligation assay. Next, we utilized CRISPR-Cas9 technology to generate Snd1-deficient mouse β-cell lines (Snd1KO) . Bulk mRNA-sequencing performed on Control and Snd1KO β-cell lines identified numerous differentially expressed genes linked to insulin secretion and cell proliferation. WST-1 cell proliferation assays revealed impaired proliferation in Snd1KO cell lines and high-glucose stimulated insulin secretion was blunted in INS-1 832/13 cells following siRNA-mediated knockdown of Snd1. As Snd1 has been shown to facilitate protein-protein interactions in other tissues, we found Snd1 interacts with numerous nuclear enriched proteins in the β-cell by IP-Mass Spec, including Stat3, Pax6, Ruvbl2 and Trim28. Future efforts are underway to characterize how Snd1 interactions with these proteins impact gene expression in vitro. Moreover, the role of Snd1 in modulating gene expression and β-cell function in vivo is being evaluated in a novel β-cell-specific (Ins1-Cre) Snd1 knockout mouse model. Disclosure S.Kanojia: None. R.K.Davidson: None. J.Spaeth: None. N.Casey: None. Funding K01DK115633Wells Center/Pediatric startupCDMD Pilot feasibility grant

Efficacy and safety study of fotemustine and bevacizumab in patients with recurrent glioblastoma.

e14041 Background: Glioblastoma (GBM) is highly vascularized tumor with elevated expression of vascular endothelial growth factor (VEGF). Bevacizumab (Bz) is a humanized monoclonal antibody against VEGF, which alone or associated with chemotherapy has obtained optimal results. However, the most suitable combination with bevacizumab is yet unknown. The objective of our study is to evaluate the efficacy and safety of Bz and fotemustine (Ft) in patients (pts) with recurrent GBM of the Huelva area and to compare our results with those published in the phase II study of the Italian Association of Neuro-oncology (AINO). Methods: Observational and descriptive study of the combination of Ft-Bz in GBM at first relapse after standard radiotherapy and TMZ concomitant in the hospital area of Huelva during the years 2010-2021. We used the same therapeutic scheme of the phase II study of AINO and the IBM SPSS Statistics 22 program to analyze the variables. Results: 42 pts were treated, 16 women and 25 men. Median age 52 years. ECOG 0-1 in 85.4%. 65.9% and 92.7% received antiepileptics and corticosteroids, respectively. In 90.2%, methylation status of the promoter of O6-methylguanine-DNA methyltransferase (MGMT) was not evaluated. Previously, 56.1% of the pts underwent total/subtotal resection, 29.3% partial and 14.6% were not resected, followed by concomitant radiotherapy and temozolamide in 100% of cases. 35.7% relapsed in less than 3 months after adjuvant treatment. 61% presented unifocal relapse and 39% multifocal. 46.3% of the pts had stable disease, 24.4% partial response and 2.4% complete response. 22% presented grade 3-4 toxicity. The most frequent toxicities are listed in the table. The median progression-free survival (PFS) was 6 months (95% CI 4.18-7.82) and the median overall survival (OS) was 7 months (95% CI 4.22-9.77). Conclusions: The results obtained in our study were similar to those of other previously published studies of Bz in combination with Ft. Although PFS was slightly higher than the phase II study AINO, OS was lower, probably due to the inclusion of a high rate of rapid progressors pts. Treatment tolerance was similar to that in the phase II study AINO. An increase in hematologic adverse events stands out in our pts. Therefore, this scheme can be a valid alternative in the management of recurrent GBM.[Table: see text]

Bone Marrow Mesenchymal Cells (BMSCs) on Breast Cancer Cell Proliferation and Metastasis by Regulating Phosphatase and Tensin Homolog 12

Among the newly diagnosed cancers in women, breast cancer metastasis is a key factor contributing to the poor prognosis. BMSCs are critical components for the malignant microenvironment. Studies have shown that the interaction between tumor cells and BMSCs support breast cancer progression. However, BMSCs’ effect on breast cancer cells is not yet clear. BMSCs and breast cancer cell MCF-7 were co-cultured to analyze tumor cell proliferation and apoptosis along with analysis of E-cadherin and Vimentin expression by real-time PCR, interleukin-6 and matrix protease-2 and PTEN12 expression. Co-culture of BMSCs promoted breast cancer cell proliferation, decreased apoptosis-related Caspase 3 activity and downregulated the expression of EMT related factors, upregulated IL-6 secretion and MMP-2, and downregulated PTEN12 expression (P < 0.05). In conclusion, BMSCs can promote breast cancer cell proliferation and survival and affect breast cancer transformation possibly through inhibiting the expression of PTEN12.