Increased Gene Expression Research Articles

Parathyroid hormone promotes induction of beige adipocytes and reversibly ameliorates muscle quality and atrophy following chronic rotator cuff tear in a rat model

Fatty infiltration (FI) and muscle atrophy (MA) in the rotator cuff muscles following rotator cuff tears (RCT) persist postrepair, increasing the risk of re-tears. Brown adipocyte-like "beige adipocytes" are expected to have a therapeutic effect on intramuscular FI and MA due to their lipolytic activity and the muscle regenerative effects of their secreted factors. However, whether parathyroid hormone (PTH) ameliorates the already advanced FI and MA remains unknown. Therefore, this study aimed to clarify whether PTH promotes the expression of beige adipocytes and ameliorates advanced FI and MA following chronic RCT in rats. Supraspinatus muscles were harvested from rats with chronic RCT after 4 or 8weeks of PTH treatment and compared to those in the control group or to those at the start of treatment. FI was assessed by Oil Red O staining, and the staining area was evaluated as a percentage of the muscle cross-sectional area. MA was evaluated by measuring muscle wet weight and cross-sectional area of muscle fiber. Beige adipocyte expression was evaluated by immunostaining for uncoupling protein 1 (UCP1). Fibro-adipogenic progenitors (FAPs) were separated from muscle-injured mice. We assessed whether PTH could diminish fat droplet accumulation by promoting the differentiation of FAPs into beige adipocytes. After 4 weeks, PTH reduced the area fraction of FI in the rat supraspinatus muscle following chronic RCT compared with that at the beginning of treatment (P=.028). In addition, PTH increased wet muscle mass (P<.001), and muscle fiber cross-sectional area (P=.018) compared with measurements at the start of treatment. PTH administration promoted the expression of UCP1, a beige adipocyte marker, in the supraspinatus muscle (P=.019). PTH increased gene expression of beige adipocyte-related markers and suppressed fat droplet accumulation even after adipogenic differentiation of FAPs (P=.004) but did not reduce fat droplets that had already accumulated in invitro experiments. PTH facilitated beige adipocyte expression and reversibly ameliorated muscle quality and atrophy following chronic RCT by hindering fat droplet accumulation and facilitating muscle regeneration. Therefore, PTH may be a medical treatment for FI and MA following RCT, leading to expanded rotator cuff repair indications.

Investigating the interplay between LIGHT and HIF in inflammatory fibroblast responses

Introduction: Intestinal fibroblast differentiation into an inflammatory phenotype likely plays a key role in Inflammatory Bowel Disease (IBD). However, the mechanisms that control this process are poorly understood. The cytokine TNFSF14 (LIGHT) drives inflammatory fibroblast responses in other diseases such as Eosinophilic esophagitis. Additionally, hypoxia, defined as low cellular or tissue levels of oxygen, is a feature of IBD progression. The use of hydroxylase inhibiting drugs which exhibit hypoxia mimetic conditions have been shown in murine models to have protective effects in IBD. Here we hypothesize the role of LIGHT to be of central importance in intestinal fibroblast differentiation and test how this inflammatory phenotype may be affected by hypoxia mimetic drugs and conversely what effect LIGHT may have on the hypoxic condition. Methods: Primary human colonic fibroblasts (18CO), the hepatic stellate cell line (LX2) and the epithelial cell line (T84) were treated with LIGHT in time course studies. 18COs were also treated with hydroxylase inhibitors Dimethyloxalylglycine (DMOG), JNJ42041935 and Roxadustat. Responses were analysed using qPCR, flowcytometry and Immunoblot-analysis. Results: LIGHT-treated 18COs upregulated CCL2 (7-fold ± increase, p=&lt;0.001, n=5), CXCL10 (14-fold ± increase, p=&lt;0.0001, n=3) and IL34 (30-fold ± increase, p=&lt;0.05 n=5) at transcript level, and surface protein expression of ICAM1 (15% ± increase at 24 hours compared to vehicle, n=4) and VCAM1 (18% ± increase at 24 hours compared to vehicle, n=4) displaying distinct kinetics. This response was absent or to a much less significant degree in LX2s and T84s. DMOG decreased CCL2 (4-fold ± decrease compared to LIGHT treated samples, p=&lt;0.001, n=5) and CXCL10 (11-fold ± decrease compared to LIGHT treated samples, p=&lt;0.0001, n=3) gene expression in 18COs when subsequently treated with LIGHT for 8 hours but did not affect other targets tested such as CSF1, CXCL5, VCAM1 and ICAM1. Immunoblot analysis of 18COs showed a decreased HIF-1α stabilization when treated with hydroxylase inhibitors in the presence of LIGHT compared to hydroxylase inhibitors alone (3 different hydroxylase inhibiting drugs n=3-4, n=11 total). Preliminary data of HIF downstream activated genes VEGFA (14-fold ± increase, p=&lt;0.05 n=3) and GLUT1 (6-fold ± increase, p=&lt;0.05 n=3) show increased gene expression in 18COs when treated with DMOG with noticeable decreases when treated with a LIGHT DMOG combination. Conclusions: Our current studies show that LIGHT is a potential mediator of colonic inflammatory fibroblast differentiation and a possible inhibitor of HIF-1α stabilization. The effects of LIGHT are either significantly weaker or not displayed in colonic epithelial cells or hepatic stellate cells, indicating LIGHT-specific effects on intestinal stroma. Hydroxylase inhibitors show selective effects on the LIGHT driven response. Collectively our data reveals a potential LIGHT-hypoxia network that may be amenable for therapeutic interventions. SFI/IRC Pathway Fellows Award to Dr. Manresa, 21/PATH-S/9621. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Sex-Dependent Regulation of Mucin Gene Transcription and Goblet Cell Secretion Machinery Following Intra-Airway IL-13 in Mice with Conditional Loss of Club Cell Creb1

IL-13 is an important effector molecule in allergic asthma, converting secretoglobin-positive club cells into mucin-secreting goblet cells and promoting mucin hypersecretion. IL-13-mediated conversion of club cells to goblet cells requires decreased gene expression of forkhead box A2 ( FOXA2) and increased gene expression of SAM pointed domain containing ETS transcription factor ( SPDEF). In addition, IL-13-mediated mucin hypersecretion may include modulation of purinergic and muscarinic receptors that control basal and stimulated mucus secretion, respectively. We recently found that the cAMP response element-binding protein (CREB), a ubiquitous transcription factor, directly binds to FOXA2 and modulates mucus secretion mechanisms in mice. Here we tested the hypothesis that conditional loss of club cell Creb1 in murine airways modulates the pro-mucin effects of IL-13. Loss of club cell Creb1 augmented IL-13-mediated increases in gene expression of the major gel-forming secreted mucins Muc5ac and Muc5b in male murine airways, with no effect on IL-13-mediated increases in Muc5ac and Muc5b in female murine airways. Examination of mucin secretion mechanisms revealed that IL-13 decreased muscarinic 3 receptor ( M3R) mRNA expression in male murine airways, whereas loss of club cell Creb1 prevented this, resulting in a significant treatment by genotype interaction. Interestingly, in female airways, a main effect of genotype was observed, with loss of club cell Creb1 reducing M3R mRNA expression in the airway. Examination of the purinergic receptor P2Y ( P2ry2) mRNA revealed a main treatment effect for IL-13 to increase P2ry2 expression in male murine airways with no impact of loss of club cell Creb1. In the females, IL-13 increased P2ry2 mRNA in both genotypes, with loss of club cell Creb1 significantly blunting the effect of IL-13 on P2ry2 mRNA. Lastly, we examined goblet cell density and mucin secretion mechanisms using Alcian Blue-PAS staining post in vivo stimulation with a muscarinic agent. We found no major effect of treatment or genotype in the density of goblet cells containing acidic mucins in either male or female airways. In contrast, both males and females showed IL-13-mediated increases in the density of goblet cells containing neutral mucins that was not impacted by genotype. Our preliminary findings suggest that loss of club cell Creb1 decreases molecular mucin secretion mechanisms in male and female airways, and increases Muc5ac and Muc5b mRNA in male airways, though a corresponding genotype-dependent increase in goblet cell density post cholinergic stimulation was not observed. Thus, we conclude that loss of club cell Creb1 causes changes to molecules important for IL-13-mediated mucin hypersecretion, without apparent functional benefit or detriment to the murine airway. This work was supported by the National Institutes of Health (OD026582, HL152101), the Cystic Fibrosis Foundation (REZNIKO20I0, REZNIKO19I0), the Katie Rose Foundation (AWD08203), and the University of Florida McKnight Career Accelerator Award. Angelina Bonilla was funded by the National institutes of Health (R25GM115298). Pedro Trevizan Bau receives the University of Florida McKnight Brain Institute Gator NeuroScholar Fellowship. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

GPX4 Over-expression Is Associated with Increased Senescence Burden and Metabolic Dysfunction

Chronic inflammation is a hallmark of obesity, diabetes, and cardiovascular disease (CVD), contributing to morbidity and mortality worldwide. Adipose tissue inflammation is commonly observed in obesity-related diseases and is considered a critical driver of metabolic dysfunction. Cellular senescence, a state of cell cycle arrest, accumulates in adipose tissues in obesity and contributes to tissue inflammation through the production of pro-inflammatory factors known as the SASP (senescence associated secretory phenotype). Novel therapeutics targeting senescent cells have been shown to improve metabolic function in obesity, although the underlying mechanisms of senescent cell accumulation and clearance are not well known. Here, we compared a genetically modified mouse model that overexpresses glutathione peroxidase 4 (GPX4), with wildtype (WT) littermate mice. GPX4 is an antioxidant, selenoenzyme, and a key regulator of ferroptosis, an iron-dependent form of regulated cell death. Previous in vitro studies have shown that activation of ferroptosis through inhibition of GPX4 is associated with a decrease in senescence burden, suggesting that ferroptosis may serve as a novel senolytic pathway. We hypothesized that overexpression of GPX4 in the transgenic (TG) mice will reduce ferroptosis, increase senescence burden, and impair metabolic function compared to WT mice. To determine the effects of ferroptosis on metabolic function, GPX4 WT and TG mice were fed either a 60% high fat diet (HFD) or normal chow (NC) ad libitum for 8-10 weeks. We found higher body, liver, and white adipose tissue (WAT) mass in GPX4 TG compared to WT mice fed NC and HFD (Interaction all p&lt;0.0003, 0.0001, 0.0044, n=8-10/group). After HFD, T cell infiltration in the WAT was higher in GPX4 TG mice compared to WT (Interaction all p&lt;0.0253), and both CD8 and CD4 were significantly different from NC controls, contributing to metabolic dysfunction in these animals (p=0.027,0.207, n=4/group). Mice fed HFD demonstrated increased glucose intolerance compared to NC controls (p=0.029, p=0.0216, n=8-12/group), however, the TG HFD mice were trending towards higher glucose intolerance compared to WT HFD (p=0.17). Compared to NC and HFD fed WT mice, there was increased gene expression for the senescence markers, p16 (all p&lt;0.0041) and p21(all p&lt;0.002), as well as increased expression of SASP factors, CXCL2 (all p&lt;0.0055), TNFa (all p&lt;0.0159), and PAI1 (all p&lt;0.05, n=5-7/group) in the HFD fed TG mice, indicating increased senescence burden in the TG HFD mice. Malondialdehyde (MDA), a byproduct and marker of ferroptosis, was elevated in the HFD fed WT mice (p=0.0002) compared to all other groups, indicating increased ferroptosis. HFD fed TG mice also had increased expression of ferritin (all p&lt;0.0079, n=5/group) compared to all other groups, suggesting resistance to ferroptosis that may contribute to the increased senescence burden in the TG HFD group. Together these data demonstrate that increases in cellular senescence and impairments in metabolic function after HFD are exacerbated by increased GPX4 expression, suggesting that GPX4 may be a novel target to decrease senescence and improve metabolic function through induction of the ferroptosis pathway. Funded in part by awards from National Institute of Health Awards R01 AG 060395, R01 AG048366, T32 HL139451-05, T32 HL007576, R01 AG077751, Nora Eccles Treadwell Foundation (NEFT), and Veteran's Affairs Merit Review Award I01 BX004492. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

IL-17A-mediated JMJD3 regulation of Cxcl1 gene expression in diabetic keratinocytes increases neutrophil recruitment and impairs wound repair.

Abstract Diabetic keratinocytes display an overexpression of chemokines that contribute to prolonged inflammation and non-healing. Using sequencing, we identified that Th17 cells and an IL-17A signature predominate in human diabetic wound tissue compared to non-diabetic controls. The effects of increased IL-17A in diabetic skin on keratinocyte function is unknown. We subjected isolated murine keratinocytes to a 5hr rIL-17A stimulation and performed bulk RNA-seq. We observed increased neutrophil chemoattract genes Cxcl1, Cxcl3, Cxcl5 compared to their controls. We analyzed epigenetic enzymes known to increase gene expression and identified that JMJD3, an activating demethylase, was increased both in human and murine diabetic keratinocytes. IL-17A stimulation increased Jmjd3 expression in murine keratinocytes, and keratinocytes from mice with a keratinocyte-specific IL-17RA deficiency were unable to increase Jmjd3 with rIL-17A. Treatment with a JMJD3-specific inhibitor decreased chemokine genes in IL-17A-stimulated keratinocytes. Keratinocytes from mice with a keratinocyte-specific JMJD3-deficiency displayed decreased chemokines compared to their littermate controls with rIL-17A, suggesting that JMJD3 is relevant to IL-17A-mediated regulation of chemokines. We then analyzed immune cells recruited to the wounds of normal and diabetic mice via flow cytometry and found that Ly6G+ neutrophils were significantly increased in diabetic mice on day 5, a timepoint at which IL-17A is elevated.

Activin A Contribution to Diabetic Kidney Disease Injury and Effects of Activin A Antagonism In-Vitro

Diabetic kidney disease (DKD) is the worldwide leader in end-stage kidney failure and has limited treatment options. Cellular senescence abundance and inflammation contribute to DKD pathogenesis. We previously demonstrated that activin A, an inflammatory mediator of pro-fibrotic kidney injury and cellular senescence, is increased in DKD and correlates with DKD injury and kidney function. We hypothesized that activin A contributes to DKD injury causing pro-fibrotic, pro-inflammatory, and cellular senescence effects in vitro, and that antagonism of activin A is achieved with follistatin. An in vitro model of DKD injury was applied with high-glucose (HG) and transforming growth factor-β (TGFβ) exposure for 24 hours in U-937-derived macrophages and renal epithelial (HK2) cells. Prior to exposure, U-937 cells were converted to M0 macrophages with phorbol myristate acetate (PMA). To examine the potential of reducing activin A release by DKD injured cells, follistatin, an activin A antagonist, was added to the media of cells at the time of injury for 24 hours. Additionally, an activin A neutralizing antibody was added to the collected supernatant of DKD injured cells. Activin A, senescence (p16), pro-fibrotic (collagen I), and pro-inflammatory (monocyte chemoattractant protein [MCP]-1, tumor necrosis factor [TNF]-alpha, interleukin [IL]-6) markers were measured through qPCR, ELISA (activin A), and immunofluorescence staining (activin A). DKD injury in renal epithelial cells increased protein expression of activin A, increased gene expression of activin A, p16, and collagen I, but did not significantly increase MCP-1 expression at 24 hour exposure (Figure 1). Furthermore, DKD injury led to increased activin A release in conditioned medium vs. control renal epithelial cells. Administration of follistatin reduced activin A release by renal epithelial cells. Application of neutralizing antibody reduced activin A levels in collected medium after cell injury. After DKD injury in U-937 derived macrophages, activin A gene expression was increased. Injured macrophages had increased gene expression of TNF-alpha and IL-6, demonstrating a pro-inflammatory phenotype. In conclusion, diabetic kidney injury increases activin A release, inflammation markers, and senescence markers in kidney cells and macrophages, therein contributing to downstream effects that further propagate disease. As shown, follistatin antagonism reduces activin A release by injured cells. Additional studies are needed to explore activin A as a promising therapeutic target and follistatin as a possible treatment in DKD. NIH 5TL1TR002380-07, AG076537, DK123492, DK109134. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Expressions of CD80 and CD86 in Cancer Patients and Its Prognostic Significance

Background: Strategies for modulating the tumor microenvironment (TME) have opened up new treatment paths in a variety of malignancies, with dramatic but variable intertemporal success. Consequently, studying TME's molecular players may aid in understanding how tumor cells and TME interact. Tumor cells and infiltrative tumor lymphocytes express immune checkpoint proteins including Cluster of Differentiation 80 (CD80) and CD86 on their surface. CD80 and CD86 are members of the immunoglobulin superfamily (IgSF). The costimulatory protein CD28, which is present on the outermost portion of all T cells, and the inhibitory receptor CTLA-4 (cytotoxic T-lymphocyte antigen-4, also known as CD152) are ligands for CD86 and CD80. CD28 and CTLA-4 have essential yet opposing functions in T cell activation. T cell responses are stimulated when they bind to CD28, but they are suppressed when they connect to CTLA-4. Aim of the study: The goal of the study was to determine how much the CD80 and CD86 genes were expressed genetically. In five blood malignancies and ten solid tumors. Methods: Clinical specimen were collected from patient with different cancers attending Kirkuk oncology centre. Patients were categorised into two main groups: solid tumour group and Blood derived cancer group alongside with their control counterparts. The study investigated the genetic expression of the CD80 and CD86 genes using q RT-PCR technologies. Results: Using q RT- PCR we measured the expression of a gene CD80 /CD86. The results showed different levels of elevation in patient samples of solid tumors and hematological tumors and compared with the control group for this study First, evaluation of this marker CD80/CD86 in solid tumors showed a significant increase in patients with brain cancer compared to their counterparts in the control group. Secondly, the second solid tumor appears increased gene expression as ovarian cancer. The least expressed solid tumors are breast cancer. As for cancers that occur in the blood, lymphoma has an upregulation expression. And the least expression is CD80/CD86 were in ALL. Conclusion: There is evidence that several cancer types and immune cells have expressed CD80 and CD86. This investigation demonstrated that the cell surface markers CD80/CD86 have a role in progression of carcinomas of the brain.

Specific Immune Marker Associated With Coronavirus Disease 2019

Background: The infection that caused the pneumonia, the virus known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the source of coronavirus disease 2019 (COVID-19) is the source of the global pandemic. The pneumonia cases began in Wuhan, China in December 2019 and had no apparent reason. Essential inflammatory mediators Chemokines are necessary for an immune response to eliminate pathogens. However, the fundamental cause of hyperinflammation is their excessive release. Chemokines may be directly responsible for the acute respiratory illness syndrome in the current COVID-19 outbreak. Objective: The current study aimed to estimate hematological changes with immunogenic markers CD177, GNLY, and CXCR4 gene expression in COVID-19 patients, and compare these parameters among patients and healthy individuals. Materials and Methods: The study was conducted at Al-Yarmouk Teaching Hospital and Medical Teaching Laboratories. Finally, to detect the immune markers level in samples of patients, by RT-PCR. Results: The results revealed the gene expression results in patients with COVID-19 means Ct difference of CD177 (24.70) fold more than (10.5) elevated when compared with the mean of Ct control healthy group is (28.21) fold of gene expression is 1.00 of CD177 when our study showed increased gene expression the mean of GNLY Ct receptor was reached to (28.70 ng/L) with fold more than 15.0 in COVID-19 patients, while mean Ct in healthy control was (32.61 ng/L) with fold 1.00, the current study found mean CT of CXCR4 gene expression ( 31.2) with fold (1.1) compared with healthy control CT(31.3) fold of gene expression was (1.0). Conclusion: It was concluded that COVID-19 caused significant changes in many immunological parameters and found the titer of immune markers was higher in infection compared with control.

A Bidirectional Non-Coding RNA Promoter Mediates Long-Range Gene Expression Regulation.

Recent evidence suggests that human gene promoters display gene expression regulatory mechanisms beyond the typical single gene local transcription modulation. In mammalian genomes, genes with an associated bidirectional promoter are abundant; bidirectional promoter architecture serves as a regulatory hub for a gene pair expression. However, it has been suggested that its contribution to transcriptional regulation might exceed local transcription initiation modulation. Despite their abundance, the functional consequences of bidirectional promoter architecture remain largely unexplored. This work studies the long-range gene expression regulatory role of a long non-coding RNA gene promoter using chromosome conformation capture methods. We found that this particular bidirectional promoter contributes to distal gene expression regulation in a target-specific manner by establishing promoter-promoter interactions. In particular, we validated that the promoter-promoter interactions of this regulatory element with the promoter of distal gene BBX contribute to modulating the transcription rate of this gene; removing the bidirectional promoter from its genomic context leads to a rearrangement of BBX promoter-enhancer interactions and to increased gene expression. Moreover, long-range regulatory functionality is not directly dependent on its associated non-coding gene pair expression levels.

The Immune Response of OAS1, IRF9, and IFI6 Genes in the Pathogenesis of COVID-19.

COVID-19 is characterized by a wide range of clinical manifestations, where aging, underlying diseases, and genetic background are related to worse outcomes. In the present study, the differential expression of seven genes related to immunity, IRF9, CCL5, IFI6, TGFB1, IL1B, OAS1, and TFRC, was analyzed in individuals with COVID-19 diagnoses of different disease severities. Two-step RT-qPCR was performed to determine the relative gene expression in whole-blood samples from 160 individuals. The expression of OAS1 (p < 0.05) and IFI6 (p < 0.05) was higher in moderate hospitalized cases than in severe ones. Increased gene expression of OAS1 (OR = 0.64, CI = 0.52-0.79; p = 0.001), IRF9 (OR = 0.581, CI = 0.43-0.79; p = 0.001), and IFI6 (OR = 0.544, CI = 0.39-0.69; p < 0.001) was associated with a lower risk of requiring IMV. Moreover, TGFB1 (OR = 0.646, CI = 0.50-0.83; p = 0.001), CCL5 (OR = 0.57, CI = 0.39-0.83; p = 0.003), IRF9 (OR = 0.80, CI = 0.653-0.979; p = 0.03), and IFI6 (OR = 0.827, CI = 0.69-0.991; p = 0.039) expression was associated with patient survival. In conclusion, the relevance of OAS1, IRF9, and IFI6 in controlling the viral infection was confirmed.

Lack of mTORC2 signaling in CD11c+ myeloid cells inhibits their migration and ameliorates experimental colitis.

The mammalian target of rapamycin (mTOR) pathway plays a key role in determining immune cells function through modulation of their metabolic status. By specific deletion of Rictor in CD11c+ myeloid cells (referred to here as CD11cRicΔ/Δ), we investigated the role of mTOR complex 2 (mTORC2) signaling in dendritic cells (DCs) function in mice. We showed that upon dextran sulfate sodium-induced colitis, the lack of mTORC2 signaling CD11c+ cells diminishes the colitis score and abrogates DC migration to the mesenteric lymph nodes, thereby diminishing the infiltration of T helper 17 cells in the lamina propria and subsequent inflammation. These findings corroborate with the abrogation of cytoskeleton organization and the decreased activation of Rac1 and Cdc42 GTPases observed in CD11c+-mTORC2-deficient cells. Meta-analysis on colonic samples from ulcerative colitis patients revealed increased gene expression of proinflammatory cytokines, which coincided with augmented expression of the mTOR pathway, a positive correlation between the DC marker ITGAX and interleukin-6, the expression of RICTOR, and CDC42. Together, this work proposes that targeting mTORC2 on DCs offers a key to hamper inflammatory responses, and this way, ameliorates the progression and severity of intestinal inflammatory diseases.

Effect of metformin in hypothalamic astrocytes from an immunocompromised mice model

Astrocytes are glial cells that play key roles in neuroinflammation, which is a common feature in diabetic encephalopathy and aging process. Metformin is an antidiabetic compound that shows neuroprotective properties, including in inflammatory models, but astroglial signaling pathways involved are still poorly known. Interferons α/β are cytokines that participate in antiviral responses and the lack of their signaling increases susceptible to viral infections. Here, we investigated the effects of metformin on astrocytes from hypothalamus, a crucial brain region related to inflammatory processes. Astrocyte cultures were derived from interferon α/β receptor knockout (IFNα/βR−/−) and wild-type (WT) mice. Metformin did not change the expression of glial fibrillary acidic protein but caused an anti-inflammatory effect by decreasing pro-inflammatory cytokines (tumor necrosis factor-α and interleukin-1β), as well as increasing gene expression of anti-inflammatory proteins interleukin-10 and Nrf2 (nuclear factor erythroid derived 2 like 2). However, nuclear factor κB p65 and cyclooxygenase 2 were downregulated in WT astrocytes and upregulated in IFNα/βR−/− astrocytes. AMP-activated protein kinase (AMPK), a molecular target of metformin, was upregulated only in WT astrocytes, while sirtuin 1 increased in both mice models. The expression of inducible nitric oxide synthase was decreased in WT astrocytes and heme oxygenase 1 was increased in IFNα/βR−/− astrocytes. Although loss of IFNα/βR-mediated signaling affects some effects of metformin, our results support beneficial roles of this drug in hypothalamic astrocytes. Moreover, paradoxical response of metformin may involve AMPK. Thus, metformin can mediate glioprotection due its effects on age-related disorders in non-diabetic and diabetic encephalopathy individuals.

Placental senescence pathophysiology is shared between peripartum cardiomyopathy and preeclampsia in mouse and human.

Peripartum cardiomyopathy (PPCM) is an idiopathic form of pregnancy-induced heart failure associated with preeclampsia. Circulating factors in late pregnancy are thought to contribute to both diseases, suggesting a common underlying pathophysiological process. However, what drives this process remains unclear. Using serum proteomics, we identified the senescence-associated secretory phenotype (SASP), a marker of cellular senescence associated with biological aging, as the most highly up-regulated pathway in young women with PPCM or preeclampsia. Placentas from women with preeclampsia displayed multiple markers of amplified senescence and tissue aging, as well as overall increased gene expression of 28 circulating proteins that contributed to SASP pathway enrichment in serum samples from patients with preeclampsia or PPCM. The most highly expressed placental SASP factor, activin A, was associated with cardiac dysfunction or heart failure severity in women with preeclampsia or PPCM. In a murine model of PPCM induced by cardiomyocyte-specific deletion of the gene encoding peroxisome proliferator-activated receptor γ coactivator-1α, inhibiting activin A signaling in the early postpartum period with a monoclonal antibody to the activin type II receptor improved heart function. In addition, attenuating placental senescence with the senolytic compound fisetin in late pregnancy improved cardiac function in these animals. These findings link senescence biology to cardiac dysfunction in pregnancy and help to elucidate the pathogenesis underlying cardiovascular diseases of pregnancy.

Phenotypes of streptozotocin-induced gestational diabetes mellitus in mice.

Gestational diabetes mellitus (GDM) in human patients disrupts glucose metabolism post-pregnancy, affecting fetal development. Although obesity and genetic factors increase GDM risk, a lack of suitable models impedes a comprehensive understanding of its pathology. To address this, we administered streptozotocin (STZ, 75 mg/kg) to C57BL/6N mice for two days before pregnancy, establishing a convenient GDM model. Pregnant mice exposed to STZ (STZ-pregnant) were compared with STZ-injected virgin mice (STZ-virgin), citrate buffer-injected virgin mice (CB-virgin), and pregnant mice injected with citrate buffer (CB-pregnant). STZ-pregnant non-obese mice exhibited elevated blood glucose levels on gestational day 15.5 and impaired glucose tolerance. They also showed fewer normal fetuses compared to CB-pregnant mice. Additionally, STZ-pregnant mice had the highest plasma C-peptide levels, with decreased pancreatic islets or increased alpha cells compared to CB-pregnant mice. Kidneys isolated from STZ-pregnant mice did not display histological alterations or changes in gene expression for the principal glucose transporters (GLUT2 and SGLT2) and renal injury-associated markers. Notably, STZ-pregnant mice displayed decreased gene expression of insulin-receiving molecules (ISNR and IGFR1), indicating heightened insulin resistance. Liver histology in STZ-pregnant mice remained unchanged except for a pregnancy-related increase in lipid droplets within hepatocytes. Furthermore, the duodenum of STZ-pregnant mice exhibited increased gene expression of ligand-degradable IGFR2 and decreased expression of GLUT5 and GLUT12 (fructose and glucose transporters, respectively) compared to STZ-virgin mice. Thus, STZ-pregnant mice displayed GDM-like symptoms, including fetal abnormalities, while organs adapted to impaired glucose metabolism by altering glucose transport and insulin reception without histopathological changes. STZ-pregnant mice offer a novel model for studying mild onset non-obese GDM and species-specific differences in GDM features between humans and animals.

Seeds treatment with salicylic acid increases gene expression and activity of antioxidant enzymes in wheat plants under zinc or copper deficiency

The effect of wheat seeds treatment with salicylic acid (SA) on the expression of the TaCu/ZnSOD, TaFeSOD and TaCAT2 genes and the activity of antioxidant enzymes (superoxide dismutase (SOD) and catalase (CAT)) in leaves at the optimal content of zinc (2 µM) and copper (0.3 µM) in root environment or there deficiencies has been studied. It was shown for the first time that seeds treatment with SA leads to an increase of the number of genes transcripts compared to untreated plants, both under optimal conditions of mineral nutrition and under zinc or copper deficiency. The activity of enzymes, especially catalase, also increases. Judging by the MDA content, this allows one to avoid increasing the intensity of lipid peroxidation (LPO) and, accordingly, the development of oxidative stress. It is concluded that the discovered positive effect of seed treatment with SA on the activity of the main enzymes of antioxidant system may underlie the stimulating effect of this phytohormone on physiological processes in plants under microelements deficiency.

Induction of the Inflammasome Pathway by Tyrosine Kinase Inhibitors Provides an Actionable Therapeutic Target for Hepatocellular Carcinoma.

During the last decade, tyrosine kinase inhibitors (TKIs) sorafenib and regorafenib have been standard systemic treatments for advanced hepatocellular carcinoma (HCC). Previous data associated sorafenib with inflammasome activation. However, the role of the inflammasome in sorafenib and regorafenib signaling has not been described in liver cancer patients. For this purpose, we analyzed inflammasome-related transcriptomic changes in a murine HCC model. Our data confirmed inflammasome activation after both TKI treatments, sharing a similar pattern of increased gene expression. According to human database results, transcriptional increase of inflammasome genes is associated with poorer prognosis for male liver cancer patients, suggesting a sex-dependent role for inflammasome activation in HCC therapy. In biopsies of HCC and its surrounding tissue, we detected durable increases in the inflammasome activation pattern after sorafenib or regorafenib treatment in male patients. Further supporting its involvement in sorafenib action, inflammasome inhibition (MCC950) enhanced sorafenib anticancer activity in experimental HCC models, while no direct in vitro effect was observed in HCC cell lines. Moreover, activated human THP-1 macrophages released IL-1β after sorafenib administration, while 3D Hep3B spheres displayed increased tumor growth after IL-1β addition, pointing to the liver microenvironment as a key player in inflammasome action. In summary, our results unveil the inflammasome pathway as an actionable target in sorafenib or regorafenib therapy and associate an inflammasome signature in HCC and surrounding tissue with TKI administration. Therefore, targeting inflammasome activation, principally in male patients, could help to overcome sorafenib or regorafenib resistance and enhance the efficacy of TKI treatments in HCC.

Enteropathogenic Escherichia coli modulates the virulence and pathogenicity of Entamoeba dispar

Amoebiasis is a disease caused by Entamoeba histolytica, affecting the large intestine of humans and occasionally leading to extra-intestinal lesions. Entamoeba dispar is another amoeba species considered commensal, although it has been identified in patients presenting with dysenteric and nondysenteric colitis, as well as amoebic liver abscess. Amoebic virulence factors are essential for the invasion and development of lesions. There is evidence showing that the association of enterobacteria with trophozoites contributes to increased gene expression of amoebic virulence factors. Enteropathogenic Escherichia coli is an important bacterium causing diarrhea, with high incidence rates in the world population, allowing it to interact with Entamoeba sp. in the same host. In this context, this study aims to evaluate the influence of enteropathogenic Escherichia coli on ACFN and ADO Entamoeba dispar strains by quantifying the gene expression of virulence factors, including galactose/N-acetyl-D-galactosamine-binding lectin, cysteine proteinase 2, and amoebapores A and C. Additionally, the study assesses the progression and morphological aspect of amoebic liver abscess and the profile of inflammatory cells. Our results demonstrated that the interaction between EPEC and ACFN Entamoeba dispar strains was able to increase the gene expression of virulence factors, as well as the lesion area and the activity of the inflammatory infiltrate. However, the association with the ADO strain did not influence the gene expression of virulence factors. Together, our findings indicate that the interaction between EPEC, ACFN, and ADO Entamoeba dispar strains resulted in differences in vitro and in vivo gene expression of Gal/GalNAc-binding lectin and CP2, in enzymatic activities of MPO, NAG, and EPO, and consequently, in the ability to cause lesions.

Smoking enhances proliferation, inflammatory markers, and immunoglobulins in peripheral blood mononuclear cells from Graves' patients.

Graves' disease (GD) and Graves' ophthalmopathy (GO) are complex autoimmune diseases. This study delved into the impact of cigarette smoke extract (CSE), simvastatin, and/or diclofenac on peripheral blood mononuclear cells (PBMCs). Specifically, we explored alterations in IL-1B, IL-6, PTGS2 expression, B- and T-lymphocyte proliferation, and Immunoglobulin G (IgG) production. We also assessed IGF1's influence on B- and T-lymphocyte proliferation. PBMCs from Graves' patients were exposed to CSE with/without simvastatin and/or diclofenac. Gene and protein expression was compared with untreated PBMCs. B- and T-lymphocyte proliferation was assessed following IGF1 treatment. PBMCs exposed to CSE exhibited increased expression of IL-1B (6-fold), IL-6 (10-fold), and PTGS2 (5.6-fold), and protein levels of IL-1B (4-fold), IL-6 (16-fold) and PGE2 (3.7-fold) compared with untreated PBMCs. Simvastatin and/or diclofenac downregulated the expression of PTGS2 (0.5-fold), IL-6 (0.4-fold), and IL-1B (0.6-fold), and the protein levels of IL-1B (0.6-fold), IL-6 (0.6-fold), and PGE2 (0.6-fold) compared with untreated PBMCs. CSE exposure in PBMCs increased the proliferation of B and T lymphocytes by 1.3-fold and 1.4-fold, respectively, compared with untreated. CSE exposure increased IgG (1.5-fold) in supernatant from PBMCs isolated from Graves' patients. IGF1 treatment increased the proliferation of B and T lymphocytes by 1.6-fold. Simvastatin downregulated the proliferation of B and T lymphocytes by 0.7-fold. Our study shows that CSE significantly upregulated the expression and release of the inflammatory markers PTGS2, IL-6 and IL-1B,the IgG levels, and the proliferation of B and T lymphocytes. Additionally, IGF1 increased the proliferation of B and T lymphocytes. Finally, these effects were decreased by diclofenac and/or simvastatin treatment.

Alleviating effect of calcium sulfate on aluminum stress through enhanced malate release with increased gene expression of malate transporter and sulfate transporter

ABSTRACT Gypsum (calcium sulfate) are commonly used to improve subsoil acidity. Aluminum (Al) toxicity in acid soil reduces crop yields worldwide, especially in the tropical regions. In soil, sulfate decrease Al toxicity and improves plant growth and yield. Here, we aimed to investigate the effects of CaSO4 on Al stress of Al-tolerant accession Col-0 as well as the Al-sensitive accessions, Wei-0 and Ts-5, of in Arabidopsis thaliana using hydroponics. Our research indicates that CaSO4 supply has a significant effect on root growth and malate release compared to CaCl2 under Al stress, especially in Al-sensitive accessions. In response to Al stress, CaSO4 supply enhanced the expression of malate transporter gene AtALMT1 and sulfate transporter gene SULTR3;5, which were regulated by the Al-resistant transcription factor STOP1. Furthermore, knockout lines of SULTR3;5 and SULTR2;1, which are involved in sulfate uptake and translocation, were more Al-sensitive than the wild type under Al stress without CaSO4, and malate release was reduced with decreased AtALMT1 expression. In conclusion, one of the alleviative effects of gypsum application on Al rhizotoxicity is promoted by sulfate through organic acid release mechanism enhancing AtALMT1 expression, alongside the Ca2+ reduction Al3+ activity on the plasma membrane.

Immune-phenotyping and transcriptomic profiling of blood monocytes from patients with breast cancer under neoadjuvant chemotherapy

Introduction. Chemotherapy is a common treatment for breast cancer. Chemotherapeutic drugs effect blood monocytes, which are major contributors to cancer pathogenesis. However, to date, pro-tumor or anti-tumor programming by chemotherapy of monocytes is controversial.Aim. To characterize changes in phenotypic and transcriptomic profiles of monocytes of breast cancer patients before and after chemotherapeutic treatment.Materials and methods. In a cohort of 50 breast cancer patients, monocyte populations were identified based on their expression of CD14, CD16, CD163, and HLA-DR evaluated by flow cytometry before and after neoadjuvant chemotherapy. Bulk RNA sequencing was adopted to explore the transcriptomic profile of CD14+ monocytes before and after treatment. After treatment, we observed an increase in the activity of signaling pathways related to lipid metabolism and intracellular transport of vesicles from the endoplasmic reticulum, against the background of a decreased response to exposure to interferon γ and interferon α, and foreign molecules (exogenous nucleic acids, viruses and bacteria).Results. In breast cancer patients, neoadjuvant chemotherapy decreased in CD14+16+HLA-DR+ monocytes. Under cytostatic treatment, increased gene expression of MGLL, NR4A2, UCK1, YOD1, ABCA2, PAPSS2, ATP10 (log2FoldChange ≥0.8; false discovery rate (FDR) ≤0.01) and decreased gene expression of KPNA2, ERCC4, JAGN1, RUBCNL, SMYD4, B3GALT4 (log2FoldChange ≥0.8; FDR ≤0.01) were observed in monocytes of patients. Using discriminant analysis, the relative numbers of CD14+16–, CD14+16+, CD14-16+, CD14+16-HLA-DR+, CD14+16+HLA-DR+ and CD14–16+HLA-DR+ monocytes in the blood were found to be valuable in predicting response to neoadjuvant chemotherapy.Conclusion. Thus, association of blood monocytes with chemotherapeutic treatment in breast cancer was revealed.