Bioscience Reports

Preeclampsia (PE), new onset hypertension after 20 weeks of gestation, affects 10% of all pregnancies in the U.S. and it is associated with progesterone deficiency, chronic inflammation, elevated angiotensin II type 1 receptor agonistic autoantibody (AT1-AA) and endothelial dysfunction. Progesterone, through its receptors, stimulates an anti- inflammatory protein called Progesterone Induced Blocking Factor (PIBF) which decreases during various pregnancy disorders. Therefore, this study was designed to test the hypothesis that a progestogen, in the form of 17-hydroxyprogesterone caproate, stimulates PIBF, lowers vasoactive mechanisms which reduces maternal blood pressure in women with early-onset preeclampsia (EOPE). PE women received 17-OHPC (250 mg, I.M.) and blood draws were collected before and after 17-OHPC supplementation. Placentas were collected at the delivery. 17-OHPC prolonged time of delivery beyond 72h on average and maternal blood pressure was significantly decreased in PE+17- OHPC. Progesterone and PIBF levels were reduced in PE group vs. NP group. Importantly, 17-OHPC increased PIBF and decreased vasoactive mechanisms and markers of inflammation. In conclusion, 17-OHPC or progesterone supplementation improves maternal outcomes in response to EOPE without causing further harm to the fetus.

Congenital disorders of glycosylation and deglycosylation are rare, serious, and lethal disorders afflicting humans. CDGs and CDDGs result in loss of function enzymes which fail to build or break down oligosaccharides on proteins. This can produce protein aggregates and, in turn, reactive oxygen species that harm the cell eventually leading to autophagy and apoptosis. Because sperm contain high concentrations of polyunsaturated fatty acids, they are especially sensitive to these effects, which is understood as one of the leading factors in human male infertility. Sperm are developed in zebrafish similarly to humans and are useful models to examine human reproductive health, as well as genetic disorders. The combination of these advantages makes the analysis of sperm from zebrafish with heterozygous ALG1 or DPAGT1 CDGs or the NGLY1 CDDG suitable. Analysis of sperm concentration, motility, status, viability, and hypoosmotic swelling demonstrated the effects of these disorders on sperm quality. Results showed a significant decrease in sperm concentration, motility, and hypoosmotic swelling for all mutant zebrafish compared to the wild type. This suggests that CDGs and CDDGs influence the amount of sperm produced, the percentage of sperm cells that are mobile, and the integrity of the plasma membrane.

Synthetic 6-Br-Q0C10 has been shown to have a partial electron transfer activity of native coenzyme Q in the isolated mitochondria. It reduces energy coupling efficiency by 30 %, suggesting that it may be useful in the management of obesity. The effect of 6-Br-Q0C10 on cell growth has been confirmed by several cell lines. Whether or not it behaves in the same way in the animal, however, has not yet been tested. Recently, we investigated the effect of 6-Br-Q0C10 on growth of rats. When 6-Br-Q0C10 was dissolved in different media, such as carboxymethyl cellulose, ethanol, and mixture of oil and butter and then feed to rats It shows no toxicity and little negative effects on growth as measured body weight gains over a period of time. When higher concentration (0.5 mg) of 6-Br-Q0C10 was given to each rat in 0.3 mL of oil/butter (70%/30%) mixture via intragastric injection daily for a period, a significant reduction in body weight gains was observed. These results validate the earlier observation that 6-Br-Q0C10 reduces the growth (30-60%) of all cell lines tested, in a time- and concentration dependent manner. These results strengthen the idea of using 6-Br-Q0C10 to manage obesity. It is also implying that 6-BrQ0C10 may slow the growth rate of cancer cells and thus prolong life. (This study was partially funded by NIH grants AA030221 and DA046258 to S.L.Chang.)

Thymosin {beta}4 (T{beta}4) is a conserved acidic polypeptide with 43-amino acids participating in multiple pathophysiological processes. In this study in vivo effects of T{beta}4 on liver regeneration are investigated in carbon-tetrachloride (CCL4) induced rodent animal liver jury models. Results illustrate that exogenous T{beta}4 treatment significantly reduced CCL4-rendered liver necrosis around central vein. At 48 hours after CCL4 insults hepatocytes proliferation occur mainly around the periportal area, while hepatocytes proliferation around the necrosis area is prominently increased by exogenous T{beta}4 treatment. The holistic proliferation level of liver tissues are also enhanced by exogenous T{beta}4. Hepatocyte proliferation activities negatively correlate with the necrosis extent of the liver tissue. These results suggested firstly exogenous T{beta}4 treatment could enhance liver regeneration and exhibit prosperous potential for application in clinical conditions such as liver transplantation.

The most prevalent RNA alteration in the mammalian genome is N-6-methylenediosine (m6A). There is mounting evidence linking dysregulation of m6A regulatory factors and alterations in m6A levels to the development, course, or prognosis of ovarian cancer. Genes having prognostic value were screened using the univariate, multifactorial, and Least Absolute Shrinkage Selection Operator (LASSO) Cox regression analyses. Important genes' m6A expression in clinical material was verified by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR). In present study, all 23 regulators were significantly differentially expressed in ovarian cancer tissues. LASSO regression analysis screened for 10 key genes associ-ated with both autophagy and m6A. A risk score was constructed and nomogram was developed to forecast the prognosis of ovarian cancer patients. Additionally, individuals with ovarian cancer were classified as high-risk or low-risk; and the low-risk group might be more likely to benefit from im-munotherapy. RT-qPCR was used for the bioinformatics study of human ovarian cancer and normal tissues. Lastly, PLK2 and LEPR were confirmed to be associated with tumorigenesis in scRNA-seq. The risk score established by m6A and autophagy can be used to predict prognosis and susceptibility to anticancer drugs in patients with ovarian cancer.

Embryo implantation is early and complex stage of pregnancy begins when competent blastocyst makes a physiological attachment to receptive endometrium. Expression of numerous molecules are essential for initiation of pregnancy. leukemia inhibitory factor (LIF) is essential cytokines required for priming uterus to make it receptive for implantation. In mice, the ovarian estrogen regulated expression of LIF is absolutely required for implantation. Golden hamster showed ovarian estrogen independent process of embryo implantation. Hence, the regulation of LIF in uterus of golden hamster during early pregnancy is still ambiguous. In this study, we explored the possible regulation of LIF by uterine factor and their spatio-temporal localization and expression in the uterus of golden hamster during early pregnancy and pseudopregnancy. We further demonstrated their ability to activate prostaglandin synthesizing enzymes to achieve successful pregnancy. We used immunohistochemistry, quantitative and semiquantitative PCR to achieve the objectives. We observed the expression of LIF in all the day of early pregnancy and pseudopregnancy in the uterus of hamster. Their m-RNA was found to be upregulated around the day of implantation and decidualization. LIF showed high expression in D3 pseudopregnancy. LIF was found to be regulated by estrogen in ovariectomized uterus and significantly reduced expression of LIF was observed in letrozole treated uterine horn. Downregulated expression of prostaglandin synthesizing enzymes was observed in anti-LIF antibody treated uterus. Together, these findings highlights that uterine factor regulated LIF mediate their action via activating prostaglandin synthesizing enzymes to make uterus receptive for successful early pregnancy in hamster. HighlightO_LIExpression of LIF in uterus during pregnancy in golden hamster is independent from the presence of blastocyst C_LIO_LILIF is regulated by estrogen in ovariectomized hamster C_LIO_LIExpression of LIF mRNA is downregulated in letrozole treated uterine horn in day 5 of pregnancy indicating the possibility of their regulation by uterine estrogen in golden hamster C_LIO_LIProstaglandin synthesizing enzyme and LIF might be associated with the activation of inflammatory signals which are essential for successful establishment of early pregnancy in golden hamster. C_LI

Purpose To determine the frequency of echocardiographic abnormalities in women with preeclampsia with severe features. To describe the spectrum and types of echocardiographic abnormalities associated with preeclampsia with severe features. Method This is a Prospective observational study conducted in Vani Vilas hospital attached to Bangalore Medical College and Research Institute, Bangalore from January 2023 to December 2025. 560 pregnant women diagnosed with severe preeclampsia(SPE) were included in the study. Chronic hypertension without superimposed preeclampsia, underlying cardiac diseases and previous history of peripartum cardiomyopathy were excluded from the study. Transthoracic echocardiography-TTE (2D ECHO) was done to evaluate cardiac structure and function. Echocardiographic abnormalities identified during the study were documented and analysed using descriptive statistical methods. Results Abnormalities in ECHO was noted in 23.03%. A unique finding was the documentation of elevated pulmonary artery systolic pressures (PASP) suggestive of Pulmonary Hypertension (PH) (PASP >35 mm HG) among 20.25% of the participants. It was also the commonest abnormality on ECHO. Mild PH was the commonest (15.71%), moderate PH was seen in 3.92% and severe PH in 0.71% of cases. Next most frequent abnormality was moderate to severe valvular regurgitation (10%), followed by left ventricular hypertrophy (5.53%). Diastolic dysfunction (DD) was seen in 3.92%, systolic dysfunction(SD) in 3.57%, chamber dilatation in 3.57% and LV global hypokinesia in 3.03% cases of SPE Conclusion Preeclampsia with severe features (SPE) is associated with 23.03% abnormalities on echocardiography. SPE is associated with systolic dysfunction, diastolic dysfunction, chamber dilatation, valvular regurgitation, left ventricular hypertrophy and pulmonary hypertension.

Background: Endothelial progenitor cells (EPCs) contribute to endothelial repair and neovascularization, and EPC dysfunction is closely associated with oxidative stress-related vascular injury. Forkhead box O3a (FoxO3a) regulates cellular stress responses, whereas miR-34a has been implicated in endothelial dysfunction, senescence, and apoptosis. However, the relationship between FoxO3a and miR-34a-3p in oxidatively injured EPCs remains incompletely defined. Objective: This study investigated the role of FoxO3a in H2O2-induced EPC dysfunction and examined whether miR-34a-3p directly interacts with the FoxO3a 3' untranslated region (3'UTR). Methods: Human umbilical cord blood-derived EPCs were identified by DiI-ac-LDL uptake, FITC-UEA-1 binding, and the expression of EPC-related markers. Oxidative stress was induced by H2O2. Cell viability, apoptosis, and angiogenic capacity were evaluated using CCK-8 assay, Annexin V/7-AAD flow cytometry, and Matrigel tube formation assay, respectively. FoxO3a expression was modulated using adenoviral overexpression or knockdown vectors, and miR-34a was modulated using mimics or antagomir. FoxO3a and miR-34a expression levels were detected by Western blot and qPCR. A dual-luciferase reporter assay was used to verify the interaction between hsa-miR-34a-3p and the FoxO3a 3'UTR. Results: H2O2 reduced EPC viability, increased apoptosis, and impaired tube formation in a concentration-dependent manner. H2O2 increased FoxO3a protein abundance and miR-34a expression, whereas FoxO3a mRNA did not change markedly. FoxO3a overexpression aggravated, whereas FoxO3a knockdown partially alleviated, H2O2-induced EPC dysfunction. Similarly, miR-34a mimics further suppressed EPC viability and tube formation, while miR-34a antagomir exerted a protective effect. Dual-luciferase reporter analysis showed that hsa-miR-34a-3p significantly reduced the activity of the wild-type FoxO3a 3'UTR reporter, while mutation of the predicted binding site abolished this suppression. Conclusion: FoxO3a and miR-34a participate in oxidative stress-induced EPC dysfunction. The dual-luciferase data demonstrate that hsa-miR-34a-3p directly targets the FoxO3a 3'UTR, suggesting the presence of miR-34a-3p-mediated post-transcriptional feedback within the FoxO3a-related stress-response network in EPCs.

Objectives; The role of nuclear receptor subfamily 4, group A, member 3 (NR4A3) in hepatic steatosis, inflammation, and insulin resistance (IR) within the context of metabolic dysfunction-associated steatotic liver disease (MASLD) remains largely underexplored. Consequently, this study aimed to examine NR4A3's impact on MASLD and the potential underlying mechanisms. Methods; We aimed to elucidate the functional role of NR4A3 in MASLD through its knockdown in cell culture and animal models. To establish the cell culture model of MASLD, LO2 cells were treated with free fatty acids (FFAs), while male C57BL/6 mice were fed a high-fat diet (HFD) to create the animal model. NR4A3 knockdown was achieved using specific short hairpin RNA (NR4A3-shRNA) in the mice model and three small interfering RNAs (NR4A3-siRNAs) in the cell culture model. The lipids content, fatty acid synthesis, inflammatory factors, and IR were then assessed with and without NR4A3 knockdown. Furthermore, the underlying mechanism through which NR4A3 exerts its influence was explored by analyzing the interaction between NR4A3 and activating transcription factor 3 (ATF3). Results: In the cell culture experiments, the knockdown of NR4A3 significantly decreased the lipids content, fatty acid synthesis, and inflammatory factors in the LO2 cells treated with FFAs in the NR4A3-shRNA group compared with those in the NC-shRNA control group. In the animal model experiments, NR4A3 knockdown in the HFD male C57BL/6 mice significantly ameliorated HFD-induced hepatic steatosis, inflammation, and IR. Mechanistically, the knockdown of NR4A3 downregulated the expression and transcriptional activity of ATF3, resulting in an impaired ATF3 function. ATF3 overexpression significantly reversed lipid accumulation decline and reduced inflammation after NR4A3 knockdown. Conclusion: The downregulation of NR4A3 alleviates MASLD by modulating ATF3, suggesting this may be a promising therapeutic target.

BackgroundAlcohol-induced osteonecrosis of the femoral head (AIONFH) is an orthopedic disorder from chronic alcohol abuse, characterized by disrupted femoral head blood supply, osteocyte death and structural collapse. Current hip-preserving therapy is unsatisfactory, and most patients eventually require total hip arthroplasty. Panax Notoginseng Saponins (PNS), the core active component of Panax notoginseng, exerts pro-angiogenic and anti-osteocyte apoptosis effects, but its specific therapeutic mechanism remains unclear. ObjectiveThis study used network pharmacology, molecular dynamics simulation and animal experiments to identify PNSs active components, core targets and key pathways for AIONFH, verify its in vivo efficacy, and provide a scientific basis for clinical application. MethodsPNS active components, their targets and AIONFH-related targets were screened from databases; intersection targets constructed an interaction network, core targets were screened by three machine learning algorithms, with concurrent GO and KEGG analysis. Molecular docking was performed between core targets and PNS components; Gromacs 2022 conducted 100 ns simulation to evaluate complex stability. AIONFH rat models were grouped with 4-week intragastric intervention; pathology, immunofluorescence and PCR were used for detection. Results and DiscussionNetwork pharmacology identified 127 PNS targets and 18 intersections with 672 AIONFH targets. Six core targets (including FGF2, HSD11B1) were screened; KEGG indicated VEGF pathway as key. Ginsenoside Re bound HSD11B1 with the lowest binding energy (-12.4 kcal/mol), and 100 ns simulation confirmed complex stability. Animal experiments showed PNS improved trabecular structure and regulated osteocyte activity. PNS treats AIONFH via multi-component, multi-target mode, core mechanism being osteocyte apoptosis inhibition. Results and DiscussionNetwork pharmacology screening identified 127 potential targets of PNS, and 18 potential intersection targets were obtained by overlapping with 672 AIONFH-related targets. Six core targets including FGF2 and HSD11B1 were screened out by machine learning, and KEGG analysis indicated that the VEGF pathway and other pathways were the key signaling pathways for PNS action. Molecular docking showed that Ginsenoside Re had the lowest binding energy with HSD11B1 (-12.4 kcal/mol), and 100 ns molecular dynamics simulation confirmed the stable conformation of this complex. Animal experiments demonstrated that PNS could improve trabecular bone structure and regulate osteocyte activity. In summary, PNS exerts a therapeutic effect on AIONFH through a multi-component, multi-target and multi-pathway mode, with the core mechanism of inhibiting osteocyte apoptosis.

It has been previously shown that chronic social defeat stress caused by paired agonistic interactions between male mice is accompanied by the development of depression-like state and immune deficiency. The aim of this study was to investigate changes in the expression of C1qtnf superfamily genes (encoding the complement component related with tumor necrosis factor) in the hypothalamus, thymus and lungs against the background of the Lewis lung adenocarcinoma growth. In the experiments, on the 5th day of social stress, male mice were injected with tumor cells into the tail vein. Chronic social stress continued for the next two weeks. The transcriptomes of the hypothalamus, thymus and lungs of mice were sequenced at the Genoanalytica Collective Center (http://genoanalytica.ru/, Moscow). Changes in the expression of the C1qtnf genes in the tissues of stressed mice were studied compared with the control and mice that were additionally injected with tumor cells. Overall, significant correlations were found between expression of most genes in each tissue of the experimental groups. In the hypothalamus of stressed animals, when tumor cells were introduced, an increase in the expression of the genes C1qtnf1, C1qtnf2, C1qtnf3, C1qtnf6 and C1qtnf7 was observed compared to controls. In the thymus of these animals, tumor cell injection increased expression of the C1qtnf1, C1qtnf5, and C1qtnf6 genes. In the lung of tumor-injected stressed mice, expression of the C1qtnf1, C1qtnf2, C1qtnf7, and C1qtnf9 genes was decreased relative to controls and non-tumor-injected depressed mice, reaching near-zero levels in some mice. Analysis of C1qtnf superfamily gene expression in the all tissues revealed negative correlations between the expression of the C1qtnf1, C1qtnf2, and C1qtnf7 genes in the hypothalamus and lungs indicating synchronization of processes against the background of social stress and Levis lung adenocarcinoma.

BackgroundWe previously performed a strain intercross between atherosclerosis resistant AKR Apoe-/-mice and atherosclerosis sensitive DBA/2 Apoe-/- mice and identified the Ath28 quantitative trait locus (QTL) on the distal end of chromosome 2. Congenic strain fine mapping identified the Ath28.1 QTL atherosclerosis modifying subregion, encompassing 217 Kb, containing for only three protein-coding genes, Zbp1, Pck1, and Pmepa1, encoding respectively, Z-DNA binding protein 1, phosphoenolpyruvate carboxykinase 1, and prostate transmembrane protein androgen induced 1. MethodsThe effect of macrophage-specific knockout of Pck1 (KO) was tested using the AAV2 transduced proprotein convertase subtilisin kexin type 9 (PCSK9) overexpression mouse model of hyperlipidemia and atherosclerosis. ResultsUnexpectedly, macrophage Pck1 deficiency lowered body weight, liver weight, and HDL-cholesterol levels in both sexes, while total and non-HDL cholesterol levels were only decreased in male mice. Aortic root lesion area and necrotic lesion area were unchanged in KO mice of both sexes. ConclusionPck1 was not confirmed as an atherosclerosis modifier gene.

Background Menstrual cycle disturbances have been increasingly reported after COVID-19 vaccination, raising questions about their prevalence and clinical significance among women of reproductive age. Objective This study aimed to investigate the incidence and types of menstrual cycle alterations following different doses of COVID-19 vaccines among Iranian women of reproductive age. Methods A cross-sectional survey was conducted among vaccinated women who reported their menstrual cycle status before and after each vaccine dose. Data on cycle regularity, flow characteristics, and specific menstrual disorders were collected and analyzed. Results Menstrual cycle alterations were reported by 28.8%, 25.4%, 30.3%, and 68.4% of participants after the first, second, third, and fourth vaccine doses, respectively. The most common changes were oligomenorrhea after the first and second doses (8.9% and 5.6%), menorrhagia after the third dose (5.3%), and hypomenorrhea after the fourth dose (8.3%). Comparisons with international studies revealed a wide variation in prevalence (ranging from 25% to 78%), which may be explained by differences in methodology, population characteristics, vaccine types, and pre-vaccination health status. Conclusion A considerable proportion of Iranian women experienced menstrual alterations following COVID-19 vaccination, most commonly oligomenorrhea, menorrhagia, and hypomenorrhea. While generally self-limiting, these findings highlight the need to integrate menstrual health into post-vaccination monitoring and patient counseling. Future research should explore the underlying immune-endocrine mechanisms and long-term clinical implications of these changes.

Protein homeostasis - i.e., proteostasis - is the biological process by which the qualitative and quantitative balance of the proteome is conducted, either by preserving functionally relevant proteins or by degrading unnecessary ones. Stress conditions can modulate cellular proteostasis in order to promote cytoprotection and preserve the viability of living organisms. Among the cellular pathways already described that can play an important role in preserving biological functions by modulating proteostasis are the heat shock response and the ribosome quality control pathways. In this work, we show that the Rqc1p protein is necessary for the thermoadaptation of S. cerevisiae to heat shock, as RQC1-deficient yeast is sensitive to elevated temperatures. In silico approaches - such as multiple sequence alignment, structural analysis, and molecular dynamics simulations - confirmed earlier predictions that Rqc1p shares characteristics with the bHLH family of proteins. We also verified, through computational prediction of sub-cellular localization, that S. cerevisiae Rqc1p contains nuclear localization signals, suggesting that this protein can potentially be translocated toward the nucleus, thereby broadening its current range of recognized biological functions in this organism. Also, analysis of yeast transcriptomes subjected to heat shock showed that Rqc1p mRNA levels do not fluctuate in response to heat shock, suggesting that cellular concentrations of Rqc1p are already at optimal levels to elicit a rapid and effective response during thermal stress in S. cerevisiae.

The chemokine-like peptide GPR15LG is a known agonist of G protein-coupled receptor 15 (GPR15), a ligand-receptor pair primarily implicated in mammalian mucosal immunity and lymphocyte homing. However, the evolutionary origin and phylogenetic distribution of this signaling system remain poorly understood due to the extreme sequence diversity of GPR15LG orthologs. In this study, we identified GPR15LG orthologs in several fish species for the first time according to their conserved gene synteny, genomic organization, and amino acid sequence features. A representative ortholog from the spotted gar (Lepisosteus oculatus), termed Lo-GPR15LG, was recombinantly prepared and functionally characterized using NanoLuc Binary Technology (NanoBiT)-based {beta}-arrestin recruitment assay and homogenous ligand-receptor binding assay. Our results demonstrated that Lo-GPR15LG directly binds to and efficiently activates its cognate receptor, Lo-GPR15, with a dissociation constant (Kd) of approximately 60 nM and an EC50 value of approximately 10 nM. Functional assays further revealed that receptor activation is critically dependent on the conserved C-terminal residues. Notably, human and fish orthologs exhibited no cross-species activity, consistent with their high sequence divergence. These findings reveal that the GPR15LG-GPR15 signaling system originated in ancient fish ancestors and has remained a conserved signaling axis throughout vertebrate evolution, suggesting a fundamental role in immunity across all vertebrate lineages.

Background: Anemia during pregnancy remains a major public health concern, particularly in low- and middle-income countries, where it contributes substantially to maternal and neonatal morbidity and mortality. Identifying women at increased risk is essential for timely intervention and improved pregnancy outcomes. Objective: This study aimed to identify the significant predictors of anemia among pregnant women using logistic regression and to evaluate the association between selected clinical and sociodemographic characteristics and anemia. Methods: A cross-sectional study was conducted using secondary data obtained from a community maternity care facility in the Suame Municipality of the Ashanti Region, Ghana. Pregnant women who attended antenatal care during the study period and had complete information on hemoglobin concentration and relevant predictor variables were included. Women with missing hemoglobin measurements at registration or delivery were excluded. Logistic regression analysis was performed to identify independent predictors of anemia. Additional analyses examined the effects of age and weight, as well as the relationship between sickle cell status and blood group. Results: Logistic regression identified diastolic blood pressure, height, hemoglobin concentration at registration, maternal weight and gestational age as significant predictors of anemia during pregnancy (p < 0.05). Although employment status was statistically significant in the model, its direct association with anemia was relatively weak. Maternal age was not significantly associated with anemia. Pregnant women with sickle cell disease had a significantly higher likelihood of anemia. Blood group did not demonstrate a significant relationship with anemia. Effect sizes and confidence intervals were not available in the dataset. Conclusion: Diastolic blood pressure, height, hemoglobin concentration at registration, maternal weight, gestational age, sickle cell status and employment status were identified as important predictors of anemia during pregnancy. These findings highlight the importance of incorporating both clinical and sociodemographic characteristics into antenatal risk assessment and screening programs. Further prospective studies with larger sample sizes and more comprehensive clinical measurements are recommended to validate these findings and strengthen predictive models for anemia during pregnancy.

Cav1.2 is an L-type voltage-gated Ca2+ channel (VGCC) that supports Ca2+ influx in response to membrane depolarization. Ca2+ entering via Cav1.2 alters gene expression, activates Ca2+-dependent enzymes and has been implicated in synaptic plasticity. ORL-1 is a Gi/o-coupled G protein-coupled receptor (GPCR) that is expressed in the peripheral and central nervous systems. Both Cav1.2 and ORL-1 are expressed in the hippocampus, where they have been implicated in learning and memory. It is well-documented that ORL-1 interacts with another VGCC, Cav2.2. However, less is known about potential interactions between Cav1.2 and ORL-1. Here, we examine the interplay between Cav1.2 (Cav1c, Cav2{delta}-1, Cav{beta}1) and ORL-1 co-expressed in tsA-201 cells by using biochemical, electrophysiological and confocal imaging analysis. Co-immunoprecipitations revealed that ORL-1 independently interacts with Cav1c and Cav2{delta}-1 subunits of the Cav1.2 channel complex. Electrophysiological recordings revealed that co-expression with ORL-1 reduced Cav1.2 peak current density without altering its biophysical properties. Acute perfusion with the ORL-1 receptor agonist nociceptin (1 M) did not alter Cav1.2 current density. Confocal imaging experiments revealed that ORL-1 significantly decreases Cav1.2 plasma membrane expression by disrupting forward trafficking. Interestingly, ORL-1 did not affect Cav1.2 endocytosis. Overall, our results demonstrate a previously unrecognized interaction between ORL-1 and Cav1.2 that alters Cav1.2 membrane expression without affecting biophysical properties.

Background: Chronic and non-healing wounds remain a major clinical challenge with limited therapeutic options. Angiogenesis and inflammation are central to tissue repair, and mesenchymal stem cells (MSC) contribute to these processes through their trophic and immunomodulatory secretome. Cannabidiol (CBD) exhibits antioxidant and immunomodulatory properties. However, whether CBD-rich Cannabis sativa extract stimulate MSC toward a pro-angiogenic secretome remains unclear. Purpose: This study aims to determine whether purified CBD or a phytochemically CBD-rich full spectrum extract stimulate umbilical cord-derived human MSC (UC-hMSC) to secrete pro-angiogenic factors and enhance endothelial responses relevant to wound healing. Methods: UC-hMSC were preconditioned with either purified CBD or a CBD-rich full-spectrum extract. Transcriptional changes were assessed by qPCR. The functional impact of the resulting secretome was evaluated in vitro using HUVEC-based proliferation and tube formation assays, and in vivo through the chick chorioallantoic membrane assay. To explore underlying mechanisms, we examined HIF-1 stabilization and VEGFA release in UC-hMSC, and VEGFR-2/ERK signaling in HUVEC. Results: Purified CBD and full-spectrum CBD extract preconditioned UC-hMSC secretomes, increased HUVEC proliferation, tube formation, and enhanced vascular branching in the CAM assay. Mechanistic analyses indicated activation of the HIF-1/VEGF axis in UC-hMSC, and ERK1/2 activation in HUVEC that was sensitive to VEGFR-2 blockade. Conclusion: Purified CBD and CBD-rich full-spectrum extract prime UC-hMSC toward a pro-angiogenic secretome that promotes endothelial activation and neovascularization. These findings suggest that cannabinoid-based preconditioning of UC-hMSC involves the HIF-1/VEGF axis and VEGFR-2/ERK signaling pathways in endothelial cells, supporting further investigation of this approach in wound healing and regenerative therapies.

Delta-like protein 1 (DLK1) is a transmembrane protein involved in the regulation of cellular differentiation and stem cell maintenance in several tissues, including the pituitary gland. Although DLK1 expression has been reported in the adult pituitary, its spatiotemporal distribution during mouse pituitary development remains incompletely characterized. The aim of this study was to analyse the distribution of DLK1 during embryonic and postnatal development of the mouse pituitary gland and to characterize its relationship with hormone-producing cell populations. Immunohistochemistry was performed in Swiss albino mice from embryonic day 9.5 (e9.5) to postnatal day 15 (p15). Double immunofluorescence was used at e18.5 and p15 to examine the association of DLK1 immunoreactivity with ACTH-, TSH-, GH-, FSH- and PRL-producing cells. DLK1 immunoreactivity was detected from the earliest stages of pituitary development in Rathkes pouch and the ventral diencephalon. During embryonic development, DLK1-ir cells were widely distributed throughout adenohypophyseal and neurohypophyseal primordia and subsequently became progressively regionalized within the anterior, intermediate and tuberal lobes, as well as in the median eminence and posterior lobe. Cells displaying overlapping immunoreactivity for DLK1 and all hormone-producing cell populations analysed were observed at late embryonic and postnatal stages. Semiquantitative analysis at p15 indicated that approximately 32% of adenohypophyseal cells were DLK1-immunoreactive. These findings provide a detailed description of the spatiotemporal distribution of DLK1 during mouse pituitary ontogeny and reveal its association with differentiating endocrine cell populations throughout pituitary development.

BackgroundReactive Oxygen Species (ROS) are the small, unstable and highly reactive species, having DNA oxidizing ability. Oxidation of the DNAs purine and pyrimidine bases can lead to single or double strands in this macromolecule. In this situation, the ATM molecule, a serine-threonine kinase, targets several proteins for phosphorylation, which causes the cell cycle to stop and the DNA damage repair begins. It has previously been proven that natural polyphenols have the cancer inhibiting properties due to their high efficacy and low side effects. Silibinin is the main herbal and medical ingredient in Milk Thistle (Silybum marianum) is a polyphenol flavonolignan, which has been widely considered as an antioxidant and anticancer agent. The purpose of the present study was to investigate the ATM gene expression and measurement of reactive oxygen species (ROS) in SKBR3 cell line, treated with Silibinin. Materials and MethodsAt first, the SKBR3 cell line was cultured in RPMI1640 culture medium and MTT assay was carried out to evaluate the Silibinin cytotoxicity. Flow Cytometry was carried out for cell cycle analysis, apoptotic induction, and ROS detection. While, Real Time PCR was used to evaluate the ATM gene expression in the Silibinin-treated and un-treated SKBR3 cells. ResultsPresent results have shown that 150 {micro}M Silibinin had the most significant cytotoxicity and apoptotic induction influence after the treatment period of 48 h. Flow cytometry data have shown that Silibinin induced considerable amount of apoptosis and caused cell cycle arrest at G1/S phase and induced production of ROS. Real-time PCR results have revealed that Silibinin increased the ATM expression in SKBR3 cell line. ConclusionSilibinin causes increased ATM gene expression by inducing ROS production, which initiates cell cycle arrest and apoptotic induction in SKBR3 cells line.