Growth Factor Signaling Pathways Research Articles

#411 Magnesium deficiency abrogates the activation of Insulin-like growth factor-1 signaling pathway in the kidney induced by DPP4 inhibitor

Abstract Background and Aims Dipeptidyl peptidase-4 (DPP-4) inhibitor could attenuate cisplatin nephrotoxicity dependent on the glucagon-like peptide-1 (GLP-1) signaling pathway [1]. We had previously reported that the use of a DPP-4 inhibitor might also attenuate cisplatin nephrotoxicity in diabetic cancer patients [2], and its renoprotective effect was lost under magnesium (Mg) deficiency [3]. However, it remains to be investigated how the GLP-1 signaling pathway changes after the treatment of a DPP-4 inhibitor under Mg deficiency. The purpose of this study was to examine the changes in the downstream pathway of the GLP-1 signaling pathway after the treatment of DPP-4 inhibitor under Mg deficiency. Method Sprague Dawley rats received a Mg-deficient diet for 7 days to induce Mg deficiency (low Mg group). Control diet was given to the normal Mg group. Oral administration of a DPP-4 inhibitor was injected 1 day before sacrifice. The changes in serum active GLP-1, insulin, insulin-like growth factor (IGF-1), and the expression of their receptor in the kidney were examined. In addition, the phosphorylation of insulin/IGF-1 receptors in the kidney were investigated. Results Serum active GLP-1 increased equally in both groups with DPP-4 inhibitor administration. Renal GLP-1 receptor expression was not observed in the renal tubules. Serum IGF-1 did not change with DPP-4 inhibitor administration in the normal Mg group but increased in the low Mg group. In normal Mg group, renal IGF-1 receptor expression tended to increase with DPP-4 inhibitor administration, and its phosphorylation increased significantly. In contrast, the expression and phosphorylation of the IGF-1 receptor did not change with DPP-4 inhibitor administration in the low Mg group. The increase in serum insulin with the DPP-4 inhibitor was enhanced in the low Mg group, compared to the normal Mg group. The expression and phosphorylation of the insulin receptor did not change with DPP-4 inhibitor administration in both groups. Conclusion DPP-4 inhibitor activated the renal IGF-1 signaling pathway, but this effect disappeared under Mg deficiency. Further investigation is needed to determine whether the renal protective effect of DPP-4 inhibitor is dependent on the IGF-1 signaling pathway.

#1479 Single nucleus RNA-sequencing indicates functional heterogeneity of renal endothelial cells in chronic kidney disease

Abstract Background and Aims The burden of cardiovascular disease (CVD) in patients with chronic kidney disease (CKD) remains very high. Endothelial cell (EC) dysfunction is an important key player in the initiation and progression of both pathologies. More importantly, endothelial dysfunction occurs early in both disease courses and is potentially reversible, making it a promising target for effective preventive strategies. Recently, accumulating single-cell RNA sequencing-based evidence has highlighted EC heterogeneity in different pathophysiological contexts. However, while biological processes such as systemic inflammation, oxidative stress fibrosis are being defined as comm EC in CKD and CVD, a comprehensive understanding of at single-cell resolution is lacking. Here, we characterized renal EC subpopulations by single-nucleus RNA sequencing (snRNA-seq) to investigate alterations in CKD patients healthy controls and identify potential drivers of EC dysfunction in CKD. Method Droplet-based snRNA-seq (10X Genomics Chromium) was performed on cryopreserved kidney biopsy cores from 6 human patients with CKD (eGFR < 45 mL/min/1.73 m2) and 3 healthy controls. CKD patients included 2 patients with primary focal segmental glomerulosclerosis (FSGS) and 4 with secondary/maladaptive FSGS (2 patients with diabetes mellitus, 2 female patients in total); all patients had a Framingham 10-year cardiovascular risk score > 20, but no established CVD. Healthy control samples were pre-perfusion biopsies from patients undergoing living kidney donation (including 2 female patients). EC subclusters were annotated based on the expression of canonical EC marker genes. Differential gene expression analysis for patient samples vs. controls included all ECs and DEGs were filtered for log2FC > 0.25 or < −0.25 and adjusted P-value of < .05. Gene set enrichment analysis (GSEA) on the identified DEGs was performed using GO terms. Results A total of 4 081 ECs were present in the dataset, consisting of 5 EC subtypes in both the patient and control cohorts (Panel A). DEGs in CKD patients vs. controls included upregulated genes related to organogenesis and cell differentiation (MEF2C, MEIS2), vascular development (FLRT2), and angiogenesis regulators (VEGFC, PRCP, HDAC9) (Panel B). Notably, VEGFC and FLRT2, are involved in directing EC motility and vascular patterning. In accordance, GSEA showed enrichment for GO biological processes related to vascular endothelial growth factor signaling pathway, endothelial cell differentiation, endothelial development, regulation of angiogenesis, and endothelial cell migration (Panel C). This suggests a potential link between these genes and aberrant angiogenesis, analogous to previous findings in diabetic nephropathy. Some but not all of these changes were also reported for renal ECs of early diabetic nephropathy patients. Of note, the majority of patients studied in our cohort were non-diabetic, warranting further investigation into renal endothelial cell transcriptional alterations that are general to renal disease or specific to particular renal conditions such as diabetic nephropathy. Differential gene expression and GSEA analysis of the EC subclusters in CKD indicated involvement of the peritubular capillary and ascending vasa recta ECs in EC migration / angiogenesis-related responses, overlapping with the overall analysis described above. Glomerular capillary ECs showed upregulation of immune-related genes and processes, reflecting the inflammatory status of the kidney disease. Conclusion snRNA-seq analysis revealed heterogeneity and subtype-specific gene expression alterations of renal ECs from CKD patients compared to healthy controls. Differential functions include enhanced angiogenic (potentially compensatory) and immune-related responses. These findings suggest that specific biological functions may direct different roles of the renal EC subtypes in CKD and will be further validated for their cluster-specificity and for their therapeutic relevance.

#970 The impact of VEGF signalling pathway inhibitors and/or immune checkpoint inhibitors on kidney function over time

Abstract Background and Aims Treatment with vascular endothelial growth factor signalling pathway inhibitors (VSPI) and immune checkpoint inhibitors (ICI) has transformed outcomes in advanced kidney cancer. VSPI and ICI can both independently cause side effects that are risk factors for development or progression of chronic kidney disease (CKD). We analysed estimated glomerular filtration rate (eGFR) decline in people with kidney cancer following treatment with VSPI or ICI. Methods We linked routine healthcare databases in the West of Scotland, UK (spanning 2010–2020, population 1.4 million) to identify adults with advanced kidney cancer who received a VSPI and/or ICI. In the two years following therapy initiation, eGFR slope (defined as annualised rate of change in eGFR) was modelled using linear mixed-effects models for the whole group and subgroups of people who had nephrectomy, metastatic cancer and a baseline eGFR <60 mL/min/1.73 m² prior to systemic therapy. Additional outcomes were studied using competing risk regression considering the competing risk of death: 40% decline in eGFR, de-novo proteinuria (urine albumin creatinine ratio >3 mg/mmol), and progression to eGFR <15 mL/min/1.73 m2. The risk of all-cause mortality was estimated with Cox regression models. Results We studied 357 adults (62.5% male; median age 63.0 years, IQI 55.0-71.0) with kidney cancer who were treated with either VSPI and/or ICI. The median baseline eGFR before systemic therapy was 74.6 mL/min/1.73 m² (58.3-91.9) and most received VSPI monotherapy (86.0%). On average, eGFR remained relatively stable over time (average annual increase +1.03 mL/min/1.73 m²/year: 95% CI −1.64 to +3.70, p = 0.2). People who had nephrectomy prior to systemic therapy demonstrated the largest average increase in eGFR (+2.30 mL/min/1.73 m²/year: 95% CI −1.66 to + 6.26). People with baseline eGFR <60 mL/min/1.73 m² prior to systemic therapy had a small decline (−1.02 mL/min/1.73 m²/year: 95% CI −6.55 to +4.50) in eGFR (Fig. 1). A ≥40% decline in eGFR occurred in 82 people (23.0%) within one year of starting systemic therapy. People with diabetes were more likely to experience a ≥40% decline in eGFR (subdistribution HR 1.89: 95% CI 1.05-3.41, p = 0.04). A decline in eGFR to <15 mL/min/1.73 m² or kidney failure with replacement therapy was found in 4 (1.1%) people. Among the 75 people assessed for proteinuria, 30.2% developed de-novo microalbuminuria. Median overall survival was 1.36 years (IQI 1.11-1.75 years), but this was longer in people who had nephrectomy prior to systemic therapy: 2.13 years (IQI 1.82–2.57 years). A 40% acute or chronic decline in eGFR was not associated with increased hazards of death on univariable or multivariable analysis (adjusted HR: 1.11: 95% CI 0.85-1.46, p = 0.4). Conclusion Despite case series and prescribing guidelines highlighting adverse impact of VSPI/ICI therapy on kidney function, our results demonstrate that there is no significant impact on the average change in eGFR but highlights that people with diabetes are at higher risk of clinically significant renal events. With appropriate monitoring, more widespread use of these agents in patients with renal impairment may be warranted.

Profibrotic Inflammatory Cytokines and Growth Factors Are Predicted as the Key Targets of Uncaria gambir (Hunter) Roxb. in Keloids: An Epistatic and Molecular Simulation Approach

Keloid is characterized as the fibrotic tissue resulting from the increase of fibroblast activity. Uncaria gambir (Hunter) Roxb. possesses bioactive compounds that have potential as antifibrotic agents, while the mechanism of action in keloid has not yet been elucidated. The aim of this study was to investigate the interaction of gambir bioactive compounds with keloid target proteins using an epistatic and molecular simulation approach. The known bioactive compounds of gambir targets and keloid-related protein targets were screened using databases. The network was constructed and analyzed to obtain the core protein targets. The targets were enriched to describe the Gene Ontology (GO) and pathway related to the proteins. Eleven targets were defined as the main targets of gambir bioactive compounds related to keloid disease. Gambiriin C, Isogambirine, and Procyanidin B1 were identified as the most promising compounds with the highest binding energy to transforming growth factor beta 1 (TGFβ1), AKT serine/threonine kinase 1 (AKT1), and matrix metallopeptidase 1 (MMP1) as the target proteins. GO enrichment and pathway analysis found that gambir bioactive compounds may act on keloid-related target proteins to regulate cell proliferation, migration, transcription, and signal transduction activity via profibrotic cytokine and growth factor signaling pathways. This study provides a reference for potential targets, compounds, and pathways to explain the mechanism of gambir against keloid.

Sulforaphane Exposure Prevents Cadmium-Induced Toxicity and Mitochondrial Dysfunction in the Nematode Caenorhabditis elegans by Regulating the Insulin/Insulin-like Growth Factor Signaling (IIS) Pathway.

Cadmium (Cd) is a heavy metal that is highly toxic to humans and animals. Its adverse effects have been widely associated with mitochondrial alterations. However, there are not many treatments that target mitochondria. This study aimed to evaluate the impact of sulforaphane (SFN) pre-exposure against cadmium chloride (CdCl2)-induced toxicity and mitochondrial alterations in the nematode Caenorhabditis elegans (C. elegans), by exploring the role of the insulin/insulin-like growth factor signaling pathway (IIS). The results revealed that prior exposure to SFN protected against CdCl2-induced mortality and increased lifespan, body length, and mobility while reducing lipofuscin levels. Furthermore, SFN prevented mitochondrial alterations by increasing mitochondrial membrane potential (Δψm) and restoring mitochondrial oxygen consumption rate, thereby decreasing mitochondrial reactive oxygen species (ROS) production. The improvement in mitochondrial function was associated with increased mitochondrial mass and the involvement of the daf-16 and skn-1c genes of the IIS signaling pathway. In conclusion, exposure to SFN before exposure to CdCl2 mitigates toxic effects and mitochondrial alterations, possibly by increasing mitochondrial mass, which may be related to the regulation of the IIS pathway. These discoveries open new possibilities for developing therapies to reduce the damage caused by Cd toxicity and oxidative stress in biological systems, highlighting antioxidants with mitochondrial action as promising tools.

Metritis and the uterine disease microbiome are associated with long-term changes in the endometrium of dairy cows†.

Cows with metritis (uterine disease) during the first 1 to 2 wk postpartum have lower pregnancy rates when inseminated later postpartum (typically >10 wk). We hypothesized that metritis and the disease-associated uterine microbiome have a long-term effect on endometrial gene expression. Changes in gene expression may inform a mechanism through which disease lowers pregnancy rates. A total of 20 cows were enrolled at 1 to 2 wk postpartum to either metritis (clinical disease; n = 10) or healthy (control; n = 10) groups and randomly assigned to be slaughtered at approximately 80 d and 165 d postpartum (mid-lactation). The microbiome of the reproductive tract was sampled to confirm the presence of pathogens that are typical of metritis. In addition to the original clinical diagnosis, study cows were retrospectively assigned to uterine-disease and control groups based on the composition of their microbiome. There was no effect of early postpartum uterine disease on the uterine microbiome at mid-lactation (time of slaughter). Nonetheless, early postpartum metritis and the disease microbiome were associated with a large number of differentially-expressed genes at mid-lactation primarily in the caruncular compared with the inter-caruncular endometrium. Gene enrichment analysis identified oxidative phosphorylation as the primary pathway increased in caruncular endometrium of diseased cows whereas growth factor signaling pathways were reduced. The current study demonstrated that metritis and a uterine disease microbiome leave a sustained imprint on gene expression in the caruncular endometrium that may explain lower fertility in cows with postpartum uterine disease.

Computational insights into allosteric inhibition of focal adhesion kinase: A combined pharmacophore modeling and molecular dynamics approach

Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that modulates integrin and growth factor signaling pathways and is implicated in cancer cell migration, proliferation, and survival. Over the past decade various, FAK kinase, FERM, and FAT domain inhibitors have been reported and a few kinase domain inhibitors are under clinical consideration. However, few of them were identified as multikinase inhibitors. In kinase drug design selectivity is always a point of concern, to improve selectivity allosteric inhibitor development is the best choice. The current research utilized a pharmacophore modeling (PM) approach to identify novel allosteric inhibitors of FAK. The all-available allosteric inhibitor bound 3D structures with PDB ids 4EBV, 4EBW, and 4I4F were utilized for the pharmacophore modeling. The validated PM models were utilized to map a database of 770,550 compounds prepared from ZINC, EXIMED, SPECS, ASINEX, and InterBioScreen, aiming to identify potential allosteric inhibitors. The obtained compounds from screening step were forwarded to molecular docking (MD) for the prediction of binding orientation inside the allosteric site and the results were evaluated with the known FAK allosteric inhibitor (REF). Finally, 14 FAK-inhibitor complexes were selected from the docking study and were studied under molecular dynamics simulations (MDS) for 500 ns. The complexes were ranked according to binding free energy (BFE) and those demonstrated higher affinity for allosteric site of FAK than REF inhibitors were selected. The selected complexes were further analyzed for intermolecular interactions and finally, three potential allosteric inhibitor candidates for the inhibition of FAK protein were identified. We believe that identified scaffolds may help in drug development against FAK as an anticancer agent.

Abstract PO3-02-03: Ketogenic diet and letrozole to inhibit PI3K signaling in ER+ breast cancer

Abstract Obesity is associated with an increased risk of breast cancer recurrence, similar in magnitude to the reduction in risk seen with the use of adjuvant chemotherapy. However, whether and how obesity causes that increased risk of recurrence remains unknown. Plausible mechanisms include obesity-mediated increases in estrogen signaling, inflammatory conditions resulting from obesity, or effects on insulin or other growth factor signaling pathways resulting from the insulin-resistant state that often accompanies obesity. While phosphatidyl inositol 3-kinase (PI3K) signaling is the normal, biological effector of insulin signaling, abnormal activation of PI3K is of particular concern in breast cancer, where it is associated with resistance to endocrine therapies and HER2-targeted therapies. We hypothesize that at least part of the adverse outcome associated with obesity results from aberrant activation of PI3K signaling in tumors. This may confer resistance to established therapies, or directly stimulate tumor growth (particularly in tumors with existing PI3K pathway mutations), or both. We tested a dietary intervention designed to alter insulin resistance pathophysiology coupled with endocrine therapy in the well-established neoadjuvant treatment paradigm in breast cancer. This will allow us to study the effects of the dietary intervention directly on tumor biology. The primary objective of this neoadjuvant study was to assess feasibility and tolerability of 2 weeks of a very low-carbohydrate ketogenic diet in combination with letrozole for patients with early stage operable ER+ disease. Up to 36 patients will be enrolled for this pilot and feasibility study (24 in the treatment group and 12 controls). Baseline metabolic parameters will be measured and the treatment group will begin a dietitian-supervised 2-week diet to induce a ketogenic state, along with letrozole 2.5 mg daily. The control group will receive only letrozole. At the end of 2 weeks, patients will proceed with surgical treatment of their breast cancer. We will report the primary outcome measure of feasibility, assessed by determining 1) the proportion of patients in the diet group achieving ketosis, 2) adherence to the diet and 3) participant-reported measures of stress, fatigue, and emotional health. For both groups, cell proliferation will be measured in a tumor biopsy obtained from the surgical specimen and compared with the pre-treatment diagnostic biopsy. Correlative studies will evaluate tumor markers of insulin/PI3K signaling before and after the intervention and between diet and control. Citation Format: Brent Rexer, Vandana Abramson, Sara Nunnery, Sonya Reid, Laura Kennedy, Melinda Sanders, Jessica Bennett, Paula Gonzalez-Ericsson, Violeta Sanchez. Ketogenic diet and letrozole to inhibit PI3K signaling in ER+ breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-02-03.

New Directions for Advanced Targeting Strategies of EGFR Signaling in Cancer.

Epidermal growth factor (EGF)-EGF receptor (EGFR) signaling studies paved the way for a basic understanding of growth factor and oncogene signaling pathways and the development of tyrosine kinase inhibitors (TKIs). Due to resistance mutations and the activation of alternative pathways when cancer cells escape TKIs, highly diverse cell populations form in recurrent tumors through mechanisms that have not yet been fully elucidated. In this review, we summarize recent advances in EGFR basic research on signaling networks and intracellular trafficking that may clarify the novel mechanisms of inhibitor resistance, discuss recent clinical developments in EGFR-targeted cancer therapy, and offer novel strategies for cancer drug development.

Enhancement of southern flounder (Paralichthys lethostigma) sperm velocity and fertility by direct activation of sperm epidermal growth factor and adenylyl cyclase signaling pathways

Development of southern flounder commercial aquaculture is limited by poor sperm motility and fertility, but the underlying causes and mechanisms regulating sperm motility are unknown. In vitro treatment of southern flounder sperm with progestin hormones rapidly increases velocity (swimming speed), a major component of sperm motility, and also fertility through membrane progestin receptor alpha (mPRα) and increases in Acy (adenylyl cyclase)/cAMP signaling. The progestin-induced increase of Atlantic croaker sperm velocity, is also mediated through epidermal growth factor receptor (Egfr) transactivation resulting in extracellular-regulated kinase 1/2 (Erk1/2) signaling, but its role in regulating southern flounder sperm velocity is unknown. The hypotheses that Egfr transactivation and Erk1/2 signaling mediates progestin-induced increases in sperm velocity and calcium levels in southern flounder, and that direct stimulation of sperm Egfr/Erk1/2 and Acy/cAMP pathways increases sperm fertility in strip spawning trials were tested in the present study. Golf, Egfr, Erk1/2, and protein kinase A were detected on southern flounder sperm by immunocytochemistry. Involvement of Egfr/Erk1/2 signaling in increasing sperm velocity and calcium levels was investigated by preincubating sperm with two EGFR inhibitors, AG1478, and AG825, a matrix metalloproteinase inhibitor, Ilomastat, and two ERK1/2 inhibitors, PD98059, and U0126, for 30 min. Prior to 1 min. Treatment with progestins. Sperm swimming speed was observed under a microscope after activation with a hyperosmotic medium and recorded for 1 min. Whereas treatment with the inhibitors did not alter basal sperm velocity and calcium levels, all of them significantly attenuated velocity and calcium increases in response to progestins. Treatment in vitro for 1 min. With the Egfr agonist, recombinant human EGF (0.1 nM and 100 nM), mimicked the stimulatory effects of progestins on sperm velocity, the increase in calcium, and also fertility in a strip spawning trials with ovulated southern flounder eggs. The results suggest Egfr/Erk1/2 signaling is involved in sperm velocity and fertility responses to progestins in southern flounder. Direct in vitro treatment with the Acy activator, forskolin (10 μM) for 1 min. Mimicked the stimulatory action of progestins on sperm calcium levels, velocity, and fertility, suggesting that Acy/cAMP signaling also mediates increased sperm swimming speed and fertility. Importantly, both EGF and forskolin also directly stimulated southern flounder sperm velocity and fertility at the end of the reproductive season when sperm were no longer responsive to progestins. The results suggest that EGF and forskolin could potentially be used as pharmacological agents to enhance reproductive performance of male southern flounder broodstock.

Detoxification and Activating Blood Circulation Decoction Promotes Reendothelialization of Damaged Blood Vessels via VEGF Signaling Pathway Activation by miRNA-126.

The occurrence of in-stent restenosis (ISR) poses a significant challenge for percutaneous coronary intervention (PCI). Thus, the promotion of vascular reendothelialization is essential to inhibit endothelial proliferation. In this study, we clarified the mechanism by which Detoxification and Activating Blood Circulation Decoction (DABCD) promotes vascular reendothelialization to avoid ISR by miRNA-126-mediated modulation of the vascular endothelial growth factor (VEGF) signaling pathway. A rat model of post-PCI restenosis was established by balloon injury. The injured aortic segment was collected 14 and 28 d after model establishment. Our findings indicate that on the 14th and 28th days following balloon injury, DABCD reduced intimal hyperplasia and inflammation and promoted vascular reendothelialization. Additionally, DABCD markedly increased NO expression and significantly decreased ET-1 production in rat serum. DABCD also increased the mRNA level of eNOS and the protein expression of VEGF, p-Akt, and p-ERK1/2 in vascular tissue. Unexpectedly, the expression of miR-126a-5p mRNA was significantly lower in the aortic tissue of balloon-injured rats than in the aortic tissue of control rats, and higher miR-126a-5p levels were observed in the DABCD groups. The results of this study indicated that the vascular reendothelialization effect of DABCD on arterial intimal injury is associated with the inhibition of neointimal formation and the enhancement of vascular endothelial activity. More specifically, the effects of DABCD were mediated, at least in part, through miR-126-mediated VEGF signaling pathway activation.

Salvianic acid A sodium facilitates cardiac microvascular endothelial cell proliferation by enhancing the hypoxia-inducible factor-1 alpha/vascular endothelial growth factor signalling pathway post-myocardial infarction.

Cardiac microvascular endothelial cells (CMECs) are important cells surrounding the cardiomyocytes in the heart that maintain microenvironment homeostasis. Salvianic acid A sodium (SAAS) has been reported to prevent myocardial infarction (MI) injury. However, the role of SAAS on CMEC proliferation remains unclear. CEMCs exposed to oxygen glucose deprivation (OGD) were used to explore the angiogenic abilities of SAAS. In vivo, C57BL/6 mice were divided into three groups: sham, MI and SAAS + MI groups. Compared to OGD group, SAAS led to a reduction in the apoptotic rate and an increase of the proliferation in vitro. Additionally, SAAS increased the protein levels of Bcl2, HIF-1α and vascular endothelial growth factor (VEGF) with the reduction of Bax. In terms of the specific mechanisms, SAAS might inhibit HIF-1α ubiquitination and enhance the HIF-1α/VEGF signalling pathway to increase CMEC proliferation. Furthermore, SAAS increased the density of vessels, inhibited myocardial fibrosis and improved cardiac dysfunction in vivo. The present study has revealed that SAAS could potentially be used as an active substance to facilitate CMEC proliferation post-MI.

Antioxidant and anti-aging activities of Longan crude and purified polysaccharide (LP-A) in nematode Caenorhabditis elegans

Oxidative damage is an important cause of aging. The antioxidant and anti-aging activities of Longan polysaccharides, especially purified Longan polysaccharides, have not been thoroughly investigated. Therefore, this study aimed to investigate the antioxidant and anti-aging activities and mechanisms of crude polysaccharides and purified polysaccharides from Longan. A purified acidic Longan polysaccharide LP-A was separated from Longan crude polysaccharide LP. Subsequently, its structural characterization, anti-aging activity and mechanism were studied. The results showed that LP-A was an acidic heteropolysaccharide with an average molecular weight (Mw) of 4.606 × 104 Da which was composed of nine monosaccharides. The scavenging rate of ABTS free radical in vitro reached 99 %. In the nematode life experiment, 0.3 mg/mL LP group and LP-A group could prolong the average lifespan of nematodes by 9.31 % and 25.80 %, respectively. Under oxidative stress stimulation, LP-A group could prolong the survival time of nematodes by 69.57 %. In terms of mechanism, Longan polysaccharide can regulate insulin / insulin-like growth factor (IIS) signaling pathway, increase the activity of antioxidant enzymes, reduce lipid peroxidation, enhance the body's resistance to stress damage, and effectively prolong the lifespan of nematodes. In conclusion, LP-A has better anti-aging activity than crude polysaccharide LP, which has great potential for developing as an anti-aging drug.

Effects of moxibustion with wheat-grain size cone at "Zusanli" (ST 36) on vascular injury and oxidative stress in high-fat diet rats through mTOR/HIF-1α/VEGF signaling pathway.

To explore the effect mechanism of moxibustion with wheat-grain size cone at "Zusanli" (ST 36) on vascular injury and oxidative stress in hyperlipidemia through mammalian target of rapamycin (mTOR)/hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor (VEGF) signaling pathway. Forty healthy male SD rats with SPF grade were randomly divided into a normal group, a model group, a moxibustion group, and an inhibitor group, with 10 rats in each one. The hyperlipidemia model was established by feeding a high-fat diet for 8 weeks in rats of the model group, the moxibustion group and the inhibitor group. The moxibustion with wheat-grain size cone was delivered at bilateral "Zusanli" (ST 36) of each rat in the moxibustion group and the inhibitor group, with 3 cones on each acupoint in each intervention, once daily for 4 weeks. In the inhibitor group, before each intervention with moxibustion, rapamycin solution was injected intraperitoneally, 2.0 mg/kg. After modeling and intervention, using ELISA, the levels of total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in the serum of rats were determined. After intervention, with HE staining and oil red O staining adopted, the abdominal aortic morphology and peripheral lipid deposition were observed. Separately, using WST-1, TBA and micro-plate method, the superoxide dismutase (SOD) activity and the levels of malondialdehyde (MDA) and nitric oxide (NO) in the serum were detected. The protein expression of mTOR, HIF-1α and VEGF in abdominal aorta were measured by Western blot method. Compared with those in the normal group, the levels of TC, TG and LDL-C increased (P<0.01) and HDL-C decreased (P<0.01) in the serum of the rats in the model group, the moxibustion group and the inhibitor group after model establishment. When compared with the normal group after intervention, in the model group, the serum levels of TC, TG, LDL-C and MDA increased (P<0.01), HDL-C level, SOD activity and NO level were reduced (P<0.01); the cell structure of the abdominal arota was abnormal, the peripheral lipids deposited seriously; and the protein expression of mTOR, HIF-1α and VEGF of abdominal aorta was elevated (P<0.01, P<0.05). In comparison with the model group, the levels of TC, TG, LDL-C and MDA were reduced (P<0.01), HDL-C levels, SOD activities and NO levels elevated (P<0.01, P<0.05), as well as the protein expression of mTOR, HIF-1α and VEGF of abdominal aorta (P<0.01, P<0.05) in the moxibustion group and the inhibitor group; besides, the vascular structure was ameliorated and the lipid deposition reduced in the moxibustion group, while, the vascular structure was still abnormal and the lipid deposition declined in the inhibitor group. When compared with the inhibitor group, the serum SOD activity and NO level increased (P<0.05) and MDA decreased (P<0.05); and the protein expression of mTOR, HIF-1α and VEGF of abdominal aorta was elevated (P<0.01, P<0.05) in the moxibustion group. The vascular injury due to hyperlipidemia is repaired by moxibustion with wheat-grain size cone at "Zusanli" (ST 36) through ameliorating oxidative stress, which is associated potentially with the modulation of mTOR/HIF-1α/VEGF signaling pathway.

Deciphering Microbiome, Transcriptome, and Metabolic Interactions in the Presence of Probiotic Lactobacillus acidophilus against Salmonella Typhimurium in a Murine Model.

Salmonella enterica serovar Typhimurium (S. Typhimurium), a foodborne pathogen that poses significant public health risks to humans and animals, presents a formidable challenge due to its antibiotic resistance. This study explores the potential of Lactobacillus acidophilus (L. acidophilus 1.3251) probiotics as an alternative strategy to combat antibiotic resistance associated with S. Typhimurium infection. In this investigation, twenty-four BALB/c mice were assigned to four groups: a non-infected, non-treated group (CNG); an infected, non-treated group (CPG); a group fed with L. acidophilus but not infected (LAG); and a group fed with L. acidophilus and challenged with Salmonella (LAST). The results revealed a reduction in Salmonella levels in the feces of mice, along with restored weight and improved overall health in the LAST compared to the CPG. The feeding of L. acidophilus was found to downregulate pro-inflammatory cytokine mRNA induced by Salmonella while upregulating anti-inflammatory cytokines. Additionally, it influenced the expression of mRNA transcript, encoding tight junction protein, oxidative stress-induced enzymes, and apoptosis-related mRNA expression. Furthermore, the LEfSe analysis demonstrated a significant shift in the abundance of critical commensal genera in the LAST, essential for maintaining gut homeostasis, metabolic reactions, anti-inflammatory responses, and butyrate production. Transcriptomic analysis revealed 2173 upregulated and 506 downregulated differentially expressed genes (DEGs) in the LAST vs. the CPG. Functional analysis of these DEGs highlighted their involvement in immunity, metabolism, and cellular development. Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis indicated their role in tumor necrosis factor (TNF), mitogen-activated protein kinase (MAPK), chemokine, Forkhead box O (FOXO), and transforming growth factor (TGF-β) signaling pathway. Moreover, the fecal metabolomic analysis identified 929 differential metabolites, with enrichment observed in valine, leucine, isoleucine, taurine, glycine, and other metabolites. These findings suggest that supplementation with L. acidophilus promotes the growth of beneficial commensal genera while mitigating Salmonella-induced intestinal disruption by modulating immunity, gut homeostasis, gut barrier integrity, and metabolism.

The diverse roles of insulin signaling in insect behavior.

In insects and other animals, nutrition-mediated behaviors are modulated by communication between the brain and peripheral systems, a process that relies heavily on the insulin/insulin-like growth factor signaling pathway (IIS). Previous studies have focused on the mechanistic and physiological functions of insulin-like peptides (ILPs) in critical developmental and adult milestones like pupation or vitellogenesis. Less work has detailed the mechanisms connecting ILPs to adult nutrient-mediated behaviors related to survival and reproductive success. Here we briefly review the range of behaviors linked to IIS in insects, from conserved regulation of feeding behavior to evolutionarily derived polyphenisms. Where possible, we incorporate information from Drosophila melanogaster and other model species to describe molecular and neural mechanisms that connect nutritional status to behavioral expression via IIS. We identify knowledge gaps which include the diverse functional roles of peripheral ILPs, how ILPs modulate neural function and behavior across the lifespan, and the lack of detailed mechanistic research in a broad range of taxa. Addressing these gaps would enable a better understanding of the evolution of this conserved and widely deployed tool kit pathway.

Potential Mechanisms Involved in the Anticonvulsant Effect of Recombinant Human VEGF on Maximal Electroshock-induced Seizure

Background: Vascular endothelial growth factor (VEGF) signaling pathway plays an important role in the pathogenesis of seizure. The oxidant/antioxidant factors and miRNA expression in the brain are differentially regulated in seizure. Objectives: We aim to investigate the potential mechanism of action for the recombinant human VEGF (rhVEGF) in mice with maximal electroshock (MES)-induced seizure. Materials &amp; Methods: A total of 40 male mice (weight: 20-25 g) were treated intraperitoneally with normal saline, or rhVEGF (50, 100, and 150 µg/kg, daily for 4 consecutive days). One hour after the last injection, seizures were induced in each animal by MES. The latency for the onset of the first clonus and the duration of hind limb extension (HLE) were recorded. The levels of nitric oxide (NO), total antioxidant capacity (TAC), and micoRNA-142-5p expression were determined in the hippocampus of mice. Blood-brain barrier (BBB) permeability was also estimated by Evans blue dye extravasation method. Results: The administration of rhVEGF at all doses significantly reduced the HLE duration. However, latency for the seizure onset increased after administration of 50 and 150 μg/kg rhVEGF and decreased after administration of 100 μg/kg rhVEGF. In the brain, the NO level decreased, while TAC level and microRNA-142-5p expression increased by rhVEGF treatment in mice with MES-induced seizure. Pretreatment with rhVEGF at doses of 100 and 150 μg/kg reversed the increase in BBB leakage induced by MES-induced seizures. Conclusion: The rhVEGF administration can prevent MES-induced seizures by regulating NO, TAC, and miR-142-5 expression levels in the hippocampus and reducing BBB leakage in mice.

Identification of potential vascular endothelial growth factor receptor inhibitors via tree‐based learning modeling and molecular docking simulation

AbstractAngiogenesis, a crucial process in tumor growth, is widely recognized as a key factor in cancer progression. The vascular endothelial growth factor (VEGF) signaling pathway is important for its pivotal role in promoting angiogenesis. The primary objective of this study was to identify a powerful classifier for distinguishing compounds as active or inactive inhibitors of VEGF receptors. To build the machine learning model, compounds were sourced from the BindingDB database. A variety of common feature selection techniques, including both filter‐based and wrapper‐based methods, were applied to reduce dimensionality, subsequently, overfitting problem. Robust and accurate tree‐based classifiers were employed in the classification procedure. Application of the extra‐tree classifier using the MultiSURF* feature selection method provided a model with superior accuracy (83.7%) compared with other feature selection techniques. High‐throughput molecular docking followed by an accurate docking and comprehensive analysis of the results was performed to provide the best possible inhibitors of these receptors. Comprehensive analysis of the docking results revealed successful prediction of molecules with VEGFR1 and VEGFR2 inhibitory activity. These results emphasized that the performance of the extra‐tree model, coupled with MultiSURF* feature selection, surpassed other methods in identifying chemical compounds targeting specific VEGF receptors.

Formation of Inclusion Complex of Curcumin and Tetrahydrocurcumin Prevents Angiogenesis by Inhibiting VEGF Activity: An in-Silico Study

Curcumin and tetrahydrocurcumin (THC) are known for their anticancer properties, but limited solubility in water hinders their effectiveness against cancer. In this study, we conducted an in silico exploration of β-cyclodextrin's potential to form inclusion complexes with curcumin or THC. The aim of this study was to assess the potential of curcumin and THC inclusion complexes to inhibit vascular endothelial growth factor (VEGF) signaling pathway, a key element in carcinogenesis. The in silico analysis involved multiple stages, such as bioactive compound preparation, biological activity prediction, 3D structure retrieval of VEGF and VEGFR, protein-ligand docking, and visualization. The results of the study demonstrated that both the curcumin- and THC-inclusion complexes exhibit a lower requirement for binding free energy to interact with VEGFR compared to curcumin or THC molecules alone. When VEGFR binds with curcumin, the curcumin-inclusion complex, or the THC-inclusion complex before interacting with VEGF, there is a notable increase in the binding free energy for the VEGF-VEGFR interaction. Specifically, the presence of THC-inclusion complex demonstrates the highest binding free energy for the VEGF-VEGFR interaction. The molecular dynamic simulation study shows that when VEGFR binds with curcumin, curcumin-inclusion complex, or THC-inclusion complex, the fluctuation of amino acid residues in VEGFR decreases compared to the VEGFR protein structure before binding with these molecules. In conclusion, this study suggests that the formation of inclusion complexes holds considerable promise for enhancing the anticancer potential of curcumin and THC by augmenting their anti-angiogenic activity.

Ex Vivo Culture of Circulating Tumor Cells in the Cerebral Spinal Fluid from Melanoma Patients to Study Melanoma-Associated Leptomeningeal Disease.

Melanoma-associated leptomeningeal disease (M-LMD) occurs when circulating tumor cells (CTCs) enter into the cerebral spinal fluid (CSF) and colonize the meninges, the membrane layers that protect the brain and the spinal cord. Once established, the prognosis for M-LMD patients is dismal, with overall survival ranging from weeks to months. This is primarily due to a paucity in our understanding of the disease and, as a consequence, the availability of effective treatment options. Defining the underlying biology of M-LMD will significantly improve the ability to adapt available therapies for M-LMD treatment or design novel inhibitors for this universally fatal disease. A major barrier, however, lies in obtaining sufficient quantities of CTCs from the patient-derived CSF (CSF-CTCs) to conduct preclinical experiments, such as molecular characterization, functional analysis, and in vivo efficacy studies. Culturing CSF-CTCs ex vivo has also proven to be challenging. To address this, a novel protocol for the culture of patient-derived M-LMD CSF-CTCs ex vivo and in vivo is developed. The incorporation of conditioned media produced by human meningeal cells (HMCs) is found to be critical to the procedure. Cytokine array analysis reveals that factors produced by HMCs, such as insulin-like growth factor-binding proteins (IGFBPs) and vascular endothelial growth factor-A (VEGF-A), are important in supporting CSF-CTC survival ex vivo. Here, the usefulness of the isolated patient-derived CSF-CTC lines is demonstrated in determining the efficacy of inhibitors that target the insulin-like growth factor (IGF) and mitogen-activated protein kinase (MAPK) signaling pathways. In addition, the ability to intrathecally inoculate these cells in vivo to establish murine models of M-LMD that can be employed for preclinical testing of approved or novel therapies is shown. These tools can help unravel the underlying biology driving CSF-CTC establishment in the meninges and identify novel therapies to reduce the morbidity and mortality associated with M-LMD.