VEGF Research Articles

Fabrication of curcumin-incorporated human amniotic membrane extracellular matrix-derived scaffold to enhance full-thickness wound healing in diabetic rats.

The multifactorial nature of diabetic wounds necessitates a mixed approach for successful treatment. Compensation of degenerated wound tissue extracellular matrix (ECM) and application of anti-inflammatory and antioxidant agents have been shown to be promising. Here, an attempt was made to fabricate a biocompatible wound dressing from curcumin-incorporated human amniotic membrane (HAM) ECM-derived scaffold to accelerate diabetic wound healing in rats. Therefore, after inducing diabetes, an excisional ischemic wound was created on rat skin, then treatments were administered for a period of 21days. The main groups were the diabetic animals that received an engraftment of HAM scaffold (HAMS group) and the curcumin-incorporated HAMS (HAMS/β/C group). Evaluation at post-wounding days 7, 14, and 21 indicated that the parameters related to regeneration, including wound closure, volume of new epidermis and dermis, proliferating cells, fibroblasts, blood vessels, collagen deposition, and tensile strength, as well as transcripts of Vegf, bFgf, and Tgf-β genes of the healed wound in both HAMS and HAMS/β/C groups were considerably greater than those of the diabetic group. Conversely, the presence of inflammatory cells, i.e., neutrophils and macrophages, and the transcripts of Tnf-α and Il-1β showed a dramatic decrease in the treated groups relative to the diabetic group. Finally, compared to the HAMS group, considerable differences were found with the HAMS/β/C group in almost all evaluated parameters. Overall, these results suggest that using the complementary or synergistic effects of curcumin and HAMS could be a promising approach to improve diabetic wound healing.

HDL Cholesterol-Associated Shifts in the Expression of Preselected Genes Reveal both Pro-Atherogenic and Atheroprotective Effects of HDL in Coronary Artery Disease.

The associations of high-density lipoprotein (HDL) level and functionality with lipid metabolism, inflammation, and innate immunity in coronary artery disease (CAD) remain controversial. The differential expression of a set of genes related to HDL metabolism (24 genes) and atherogenesis (41 genes) in peripheral blood mononuclear cells (PBMC) from CAD and control patients with varied HDL cholesterol (HDL-C) levels was compared. 76 male patients 40-60 years old with CAD diagnosed by angiography and 63 control patients were divided into three groups with low, normal (1.0-1.4 mM), and increased HDL-C levels. Transcript levels were measured by real-time PCR. The differentially expressed genes (DEGs) and associated metabolic pathways were analyzed for three groups, with prevalent CAD as an outcome. The common feature was the increased odds ratio values for liver X receptor (LXR) gene expression for three patient groups. CAD patients with low HDL-C possessed 24 DEGs with lower expression of genes involved in cholesterol efflux, and down-regulated SREBF1 and ABCG1 are suggested as gene signatures. CAD patients with normal HDL-C possessed nine DEGs with down-regulated ITGAM and ALB as gene signatures. CAD patients with increased HDL-C possessed 19 DEGs with down-regulated APOA1 and HMGCR as gene signatures. With gene expression signatures, one standard deviation higher average gene expressions were associated with 5.1-, 48.8-, and 38.9-fold fewer CAD cases for three patient groups. As HDL-C increased in CAD patients, the expression of ABCG1, CUBN, and HDLBP genes increased, while the expression of HMGCR and NPC2 genes, involved in cholesterol synthesis and trafficking, decreased. The expression of CD14, CD36, S100A8, S100A9, S100A12, TLR5, TLR8, and VEGFA genes, involved in angiogenesis and inflammation mainly via nuclear factor-κB (NF-κB), decreased. The increased accumulation of cholesteryl ester in PBMC from patients with low HDL-C was suggested. This assumption contrasts with the suggested accumulation of free cholesterol in PBMC from patients with increased HDL-C, concomitant with suppression of cholesterol synthesis and traffic to the plasma membrane, and with an inflammatory state controlled by depressed CD36-mediated and upregulated apoE-mediated immunometabolic signaling. Gene signatures may be used for the diagnosis, prognosis, and treatment of CAD in dependence on HDL-C levels.

Studying the effect of thymol-containing Nano emulsion extracted from thyme on angiogenesis controlling genes VEGF and VEGFR in a human liver cancer cell line

يحتوي زيت الزعتر على الثيمول (2-إيزوبروبيل-5-ميثيلفينول)، وهو مركب فينولي بلوري عديم اللون وذو رائحة مميزة. يعتبر مضاد للأكسدة. الثيمول ذو قابلية منخفضة للذوبان في الماء وقابلية ذوبان جيدة في الزيت. تم تحضير مستحلب الثيمول النانوي المذاب في زيت السمسم عن طريق تحويله إلى قطرات نانوية. حيث أن في الدراسات السابقة أجريت العديد من الدراسات البحثية لدراسة تأثير الثيمول على سرطان المعدة وتأثيره على خلايا الورم الأرومي الدبقي. لقد قاموا بدراسة تأثير VEGF على نمو وهجرة الخلايا السرطانية في الكبد. من منظور بيولوجي، يمكن للأسطح البيولوجية أن تمتص قطرات المستحلب النانوي بشكل فعال من أجل أنشطة بيولوجية أكثر كفاءة. المستحلبات النانوية هي جسيمات كروية تعمل كحامل لجزيئات الدواء، ويتراوح حجمها بين 10-1000 نانومتر. بسبب تأثير الثيمول المضاد للسرطان، يمكن استخدامه كعلاج مناسب لمنع تكوين الأوعية الدموية في سرطان الخلايا الكبدية. أجرينا تحقيقًا في السمية الخلوية باستخدام اختبار MTT، حيث تمت معالجة خط خلايا سرطان الكبد البشري بتركيزات مختلفة من مستحلب الثيمول النانوي. أشارت النتائج إلى أنه مع زيادة التركيز، انخفضت صلاحية الخلية. ثم تم استخراج الحمض النووي الريبي (RNA) وتم تصنيع C-DNA. تم بعد ذلك فحص مستويات التعبير عن جينات VEGFR وVEGF في الخلايا المعالجة بالنانوثيمول والثيمول الحر وفي الخلايا غير المعالجة. وأظهرت النتائج أن تعبير هذا الجين في الخلايا المعالجة بالثيمول الحر انخفض بمقدار 5.33 مرة مقارنة بالخلايا غير المعالجة، ولكن عندما عولجت الخلايا بالنانوثيمول انخفض مستوى التعبير الجيني بنحو 10.02 مرة.

Unveiling the therapeutic potential of anthocyanin/cisplatin-loaded chitosan nanoparticles against breast and liver cancers

BackgroundLiver and breast cancers are among the leading causes of cancer-related deaths worldwide, prompting researchers to seek natural anticancer agents and reduce chemotherapy side effects. Red beetroot (Beta vulgaris Linnaeus), rich in polyphenols and powerful antioxidants, has shown potential in cancer prevention. This study aimed to evaluate the impact of red beetroot-derived anthocyanin (Ant), Ant-loaded chitosan nanoparticles (Ant NPs), cisplatin (Cis), Cis-loaded chitosan (Cis NPs), and Cis + Ant-loaded chitosan NPs on human hepatoma HepG2 and breast adenocarcinoma MCF7 cell lines.MethodsNPs preparation was evaluated by zeta potential, FTIR, and SEM. The cytotoxic, apoptotic, antioxidant, anti-inflammatory, anti-metastatic, and anti-angiogenic effects were assessed by MTT assay, qPCR, AO/EB staining, and flow cytometry.ResultsTreatment with Ant, Ant NPs, Cis, Cis NPs, and Cis + Ant NPs caused cytotoxicity in HepG2 and MCF7 with best effect in Cis-treated cells. The anticancer effects were attributed to mitochondrial-dependent apoptosis (with high Bax and low Bcl2 expression), chromatin disintegration, and cell cycle arrest in G2/M and S phases. All treatments inhibited migration by downregulating the migration-related gene MMP9 and upregulating the anti-migratory gene TIMP1 and decreased the angiogenesis-related gene VEGF and the inflammatory gene TNFα with best results in Cis NPs-treated cells. Interestingly, Ant, Ant NPs, and Cis + Ant NPs increased the antioxidant status (high GSH and upregulated expression of Nrf2 and OH-1) and decreased drug resistance-related MAPK1 and MDR1 genes compared to Cis and Cis NPs-treated cells.ConclusionsAnthocyanin and cisplatin-loaded chitosan nanoparticles effectively combat breast and liver cancers by inducing cancer cell apoptosis, enhancing antioxidant defenses, and reducing inflammation. They also inhibit tumor spread and blood vessel formation through downregulation of MMP9 and VEGF, highlighting their therapeutic potential.

Mathematical model of liver cirrhosis formation during morphological and molecular-genetic preclinical studies

BACKGROUND: Currently, researchers describe challenges in developing new treatments for fibrosis and cirrhosis: poor quality of preclinical models, insufficient trial duration, and lack of markers of therapeutic response. A separate task is to standardize the process of liver cirrhosis formation in preclinical trials, which is necessary to obtain accurate quantitative estimates in a short timeframe. AIM: This study aimed to develop a mathematical model for the formation of liver cirrhosis during preclinical trials. MATERIALS AND METHODS: Liver fibrosis and cirrhosis were induced in male Wistar rats using freshly prepared thioacetamide solution for 17 weeks. The area of connective tissue was determined as a percentage of the image area. The area of interlobular veins was measured in µm2. The numbers of cells expressing the FAP marker and the α-SMA marker were counted. The level of mRNA expression of the Vegfa and Yap1 genes was assessed by real-time polymerase chain reaction. A mathematical model for classifying observations into stages was constructed using multiple logistic regression with stepwise selection of predictors, followed by calculation of sensitivity, specificity, and area under the curve with a 95% confidence interval based on ROC analysis. RESULTS: As a result of the analysis, a mathematical model of liver cirrhosis formation was developed. The model is based on the values of two indicators: FAP+ cells and Yap1 mRNA and demonstrated good quality. The resulting value of the area under the ROC curve of 0.883 suggests good results for classifying cases. CONCLUSIONS: The mathematical model makes it possible to differentiate the stage of liver cirrhosis from the stage of fibrosis during preclinical studies. It provides a foundation for studying the pathogenesis of liver fibrosis and cirrhosis, identifying new potential molecular targets for antifibrotic therapy, and reducing the number of expensive, labor-intensive laboratory tests.

Unveiling the antimicrobial, antivirulence, and wound-healing accelerating potentials of resveratrol against carbapenem-resistant Pseudomonas aeruginosa (CRPA)-septic wound in a murine model.

Pseudomonas aeruginosa is a repertoire of several virulence factors that create a frightening high pathogenicity level as well as high antimicrobial resistance toward commercially used antibiotics. Therefore, finding a new alternative to traditional antimicrobials is a must. Resveratrol is a very famous phytochemical that harbors many beneficial health properties by possessing antibacterial, anti-inflammatory, and antioxidant properties. The current study aimed to explore the antimicrobial efficacy of resveratrol against P. aeruginosa and explore its ability to accelerate wound healing in a murine model. The obtained results revealed the potent antimicrobial, antivirulence, and wound-healing accelerating potentials of resveratrol against carbapenem-resistant P. aeruginosa (CRPA)-septic wounds. It significantly lowered the transcript levels of P. aeruginosa virulent genes toxA, pelA, and lasB. Additionally, resveratrol significantly accelerated skin wound healing by shortening the inflammatory phase and promoting re-vascularization, cell proliferation, re-epithelialization, and collagen deposition. Furthermore, it increased the immunoexpression of αSMA along with a reduction of the mRNA levels of VEGF, IL-1β, and TNF-α genes. Resveratrol has high therapeutic potential for the treatment ofP. aeruginosawound infection and is a prospective and promising candidate for this problem.

Preparation of bimetallic calcium chromatite nanocomposite using Stevia rebaudiana leaf extract for inducing apoptosis in human cancer cell lines

Abstract Despite the widespread industrial use of chromium-based compounds, there are concerns regarding their toxicity in biomedical applications. Herein, bimetallic calcium chromatite nanocomposite (CC) was prepared through green synthesis using Stevia rebaudiana leaf extract. A porous nanorod of CC was obtained through calcinations at 800 °C. The formation of CC was confirmed by SEM (scanning electron microscopy), EDAX (Energy-dispersive x-ray spectroscopy), HRTEM (High-resolution transmission electron microscopy), TGA (Thermal gravimetric analysis), and BET (Brunauer–Emmett–Teller). The cytotoxicity of the CC was evaluated by the MTT method against cancer lines (HeLa, U87, A549) and primary cell line (HUVEC). The IC50 value of CC against cancer cell lines was lower than the primary cell lines. The calculated specificity index was greater than one, indicating that this CC might target cancer cells without the use of a targeting moiety. Gene expression studies revealed that CC upregulated the proapoptotic gene, BAX and downregulated the anti-apoptotic gene, Bcl-2, to induce apoptosis. The KDR and VEGF genes responsible for angiogenesis were down-regulated, indicating that CC decreases angiogenesis to promote apoptosis. The findings suggest CC prepared using phytoextract could be considered an adjuvant cancer therapy treatment.

Aortic valve stenosis augments long noncoding RNA H19 to govern angiogenic responses and phenotypes of valvular endothelial cell through endothelial to mesenchymal transition

Abstract Background Long noncoding RNAs (lncRNAs) have emerged as important regulators of cardiovascular disease, including aortic valve stenosis (AVS). Although several recent studies have reported that lncRNAs are dysregulated and might play a crucial role in (Endothelial to mesenchymal transition) EndMT, no specific role of lncRNA was reported in AVS pathogenesis. Here, we sought to investigate the role of H19 in EndMT during the pathogenesis of AVS. Methods and results High-throughput lncRNA sequencing of human aortic valve tissue explanted from patients with AVS due to severe stenosis during surgical aortic valve replacement (SAVR) demonstrated that certain lncRNAs [H19, AGAP2-AS1 (PUNISHER), GAS5, MALAT1] are significantly dysregulated compared to patients without AVS. To validate the RNA-seq data, 60 patients with AVS (n=30) or without AVS (n=30) were prospectively investigated. Among these, H19 (p=0.002), AGAP2-AS1 (p=0.002), and GAS5 (0.02) were significantly upregulated in patients with AVS, compared to non-AVS patients. Aortic valvular endothelial cells (VECs) were isolated from explanted valve tissue from AVS patients or obtained commercially. In vitro EndMT-promoting stimuli (TNFα or TGFß+IL1ß) were used for VEC stimulation for 5 days. RT-qPCR was used to quantify the expression of EndMT markers (EC markers, vWF, and NOS3, and mesenchymal markers, αSMA, SM22, Vimentin, and SNAI2). Loss of H19 led to dysregulation of the EC gene network and protein expression patterns that were analyzed by qRT-PCR, immunoblotting, and immunofluorescence, respectively. Further experiments show that silencing of H19 affects migration and proliferation, and promotes EndMT in EC. Functional assays suggested that H19 is a critical regulator of angiogenic properties. The molecular signature of angiogenesis was investigated using a RT-qPCR-based angiogenesis array to identify the regulation of angiogenesis with 84-angiogenesis-related genes. Mechanistically, the angiogenesis array revealed that H19 regulates Jagged Canonical Notch Ligand 1 (JAG1) (4.43-fold vs. control EC), an important regulator of NOTCH1 signaling, and pro-angiogenic factor in EC. Accordingly, EndMT marker expressions were dysregulated upon H19 depletion. A high-throughput luciferase reporter array demonstrated STAT3 activation upon H19 knockdown. Depletion of H19 increased the phosphorylation of STAT3 at TYR705 and pharmacological inhibition of STAT3 activation abolished the effects of H19 silencing on EndMT marker expression as well as on angiogenesis. Overexpression of H19 in VEC abrogated the H19-mediated promotion of JAG1- and VEGFA gene- and protein expression. Our data suggest that H19 might play a role in EC angiogenic gene expression and EndMT. The data reveal a pivotal role of H19 in controlling EndMT in AVS via regulation of the JAG1-NOTCH axis, which could potentially be therapeutically targeted to halt EndMT during AVS.

Extracellular vesicle-incorporated long noncoding RNA PUNISHER is increased in coronary artery disease and regulates endothelial cell function

Abstract Aims Long noncoding RNAs (lncRNAs) have emerged as biomarkers and regulators of cardiovascular disease. Circulating lncRNAs can be incorporated into extracellular vesicles (EVs). Our aim was to study the expression pattern of circulating EV-incorporated lncRNAs in patients with and without coronary artery disease (CAD). The regulation of the lncRNA PUNISHER was further assessed as an intercellular messenger in endothelial cell (EC) biology. Methods and results Circulating EVs were isolated from the plasma of patients using ultracentrifugation. Electron microscopy, western blot, and NTA were used to determine the size and origin of the EVs. PCR-based human lncRNA array analysis revealed that certain EV-lncRNAs are significantly different in patients with CAD compared to patients without CAD. To validate the lncRNA array results, 60 patients with (n=30) or without (n=30) CAD were prospectively studied. Four atherosclerosis-related lncRNAs (PUNISHER, GAS5, MALAT1, and H19) were quantified in the circulating EVs by using real-time quantitative PCR (RT-qPCR). Among these, PUNISHER (p=0.002) and GAS5 (0.02) were significantly increased in patients with CAD, compared to non-CAD patients. Analysis of RNA degradation revealed that circulating PUNISHER is mainly incorporated within EVs. In vitro, atherosclerotic stimuli consisting of oxLDL or TNF-a treatment caused an upregulation of PUNISHER levels in human coronary artery endothelial cells (HCAEC) and in the corresponding EVs. Labeling of EVs followed by RT-qPCR demonstrated that functional PUNISHER was transported into the recipient ECs, which accelerated cell migration, proliferation, and tube formation. Mechanistically, the RNA-binding protein hnRNPK was identified by an RNA immunoprecipitation (RIP) assay as an interaction partner of PUNISHER, regulating its loading into small EVs. Knockdown of PUNISHER abrogated the EV-mediated effects on EC migration, proliferation, and tube formation. PCR-based gene profiling showed that the expression of VEGF RNA was significantly increased in ECs by treatment with EVs. Protein stability and RNA-immunoprecipitation followed by RT-qPCR analysis indicated that the PUNISHER-hnRNPK axis regulates the stability and binding of VEGFA mRNA to hnRNPK. Knockdown of PUNISHER in EVs abrogated the EV-mediated promotion of VEGFA gene- and protein expression. Conclusion The circulating lncRNA PUNISHER is increased in CAD patients. Intercellular transfer of EV-incorporated PUNISHER promotes a pro-angiogenic phenotype via a VEGFA-dependent mechanism.

Human amniotic membrane hydrogel loaded with exosomes derived from human placental mesenchymal stem cells accelerate diabetic wound healing

Diabetic wound is one of the most common and costly complication in diabetic patients. Hence, numerous studies have been carried out to discover a suitable approach to enhance the process of wound healing. Biological hydrogels are commonly utilized for wound healing due to their suitable properties among different materials available. Herein we investigated whether human amniotic membrane hydrogel (hAMH) loaded with human placental mesenchymal stem cells (PlaMSCs)-derived exosomes could promote healing in diabetic rats. Sixty diabetic rats were randomly assigned into the control group, hAMH group, exosome group, and hAMH+Exosome group. According to the phases of wound healing, sampling was done on days 7, 14, and 21 for further assessments. Our findings showed a significant increase in wound contraction rate, new epidermal length, fibroblast and blood vessel count, collagen density, and the levels of antioxidative factors (GSH, SOD, and CAT) in the treatment groups compared to the control group, with more pronounced effects observed in the hAMH+Exosome group. Furthermore, the levels of bFGF and VEGF gene expression significantly increased in each treatment group when compared to the control group, with the highest levels observed in the hAMH+Exosome group. This occurred as the hAMH+Exosome group showed a greater decrease in neutrophil count, the expression of TNF-α and IL-1β genes, and the levels of an oxidative factor (MDA) compared to the other groups. In summary, the combination of hAMH and PlaMSCs-derived exosomes was determined to have a more significant effect on healing diabetic wounds.

Knockout of histone deacetylase 8 gene in breast cancer cells may alter the expression pattern of the signaling molecules

PurposeBreast cancer (BC) is the most common cancer diagnosed in the world and it is also the main leading cause of cancer deaths in women. Change in epigenetic mechanisms promotes BC initiation and progression. Histone deacetylase 8 (HDAC8) was found to act as a potential oncogene in different malignancies. For better understanding of the HDAC8 function in BC development, we investigated the effect of HDAC8 deletion on the expression of genes involved in signaling pathways. Materials and methodsIn this study, CRISPR technology was used to knockout the HDAC8 gene in MDA-MB-468, MDA-MB-231 and MCF-7 ​cell lines. For this purpose, two gRNAs were designed and cloned into the PX459 vector. The gRNA-containing vectors were transfected into the BC cell lines and then the effect of this deletion on the expression of genes involved in signaling pathway was determined using quantitative real-time PCR (qRT-PCR). ResultsAnalysis of qRT-PCR results showed a reduction in the expression of studied genes in BC cell lines after deletion of the HDAC8 gene compared to untreated controls. Although this decline was not significant for FGF2 and FGFR1 genes, however the mTOR, IGF1R, INSR, VEGFA and VEGFR2 genes showed statistically significant reduction in the studied BC cell lines. In addition, the down-regulation of PDGFC and PDGFRA genes were only significant in the TNBC cell lines. ConclusionOverall, our study showed that HDAC8 can exert its oncogenic effects by altering the expression level of molecules involved in some signaling pathways, and inhibiting HDAC8 can revert these effects.

Atezolizumab Plus Chemotherapy With or Without Bevacizumab in Advanced Biliary Tract Cancer: Clinical and Biomarker Data From the Randomized Phase II IMbrave151 Trial.

Biliary tract cancers (BTCs) harbor an immunosuppressed tumor microenvironment and respond poorly to PD-1/PD-L1 inhibitors. Bevacizumab (anti-vascular endothelial growth factor) plus chemotherapy can promote anticancer immunity, augmenting response to PD-L1 inhibition. This randomized, double-blind, proof-of-concept phase II study enrolled patients (n = 162) with previously untreated advanced BTC (IMbrave151; ClinicalTrials.gov identifier: NCT04677504). Patients were randomly assigned 1:1 to receive cycles of atezolizumab (1,200 mg) plus bevacizumab (15 mg/kg) or atezolizumab plus placebo once every 3 weeks until disease progression or unacceptable toxicity. All patients received cisplatin (25 mg/m2) plus gemcitabine (1,000 mg/m2; cisplatin plus gemcitabine [CisGem]) on days 1 and 8 once every 3 weeks for up to eight cycles. Stratification of patients was by disease status, geographic region, and primary tumor location. The primary end point was progression-free survival (PFS). No formal hypothesis testing was performed. Exploratory correlative biomarker analysis was undertaken using transcriptome analysis (n = 95) and mutation profiling (n = 102) on baseline tumor samples. Between February and September 2021, 162 patients were enrolled. Median PFS was 8.3 months in the bevacizumab arm and 7.9 months in the placebo arm (stratified hazard ratio [HR], 0.67 [95% CI, 0.46 to 0.95]). Median overall survival (OS) was 14.9 and 14.6 months in the bevacizumab and placebo arms, respectively (stratified HR, 0.97 [95% CI, 0.64 to 1.47]). The incidence of grade 3 or 4 adverse events was 74% in both arms. High VEGFA gene expression was associated with improved PFS (HR, 0.44 [95% CI, 0.23 to 0.83]) in the bevacizumab arm versus placebo. In unselected patients with advanced BTC, adding bevacizumab to atezolizumab plus CisGem modestly improves PFS but not OS. High VEGFA gene expression may represent a predictive biomarker of benefit from atezolizumab/bevacizumab, warranting further investigation.

Changes in Inflammatory Cytokines, Vascular Markers, Cell Cycle Regulators, and Gonadotropin Receptors in Granulosa Cells of COVID-19 Infected Women

Background: COVID-19 infection can negatively affect multiple organ systems, including the reproductive system. Previous research has indicated altered levels of inflammatory markers in the reproductive tissues of women with chronic diseases. This study aimed to assess the expression of inflammatory, vascular, cell cycle, and gonadotropin receptor genes in the granulosa cells and oocytes of women with recent COVID-19 infection undergoing Assisted Reproductive Technology (ART), compared to healthy controls. Materials and Methods: The study involved 15 women who had tested positive for COVID-19 within three months of ART treatment and 15 age-matched healthy women as controls. Granulosa cells were collected during oocyte retrieval, and RNA was isolated to analyze gene expression using quantitative real-time PCR. The evaluated genes included inflammatory cytokines (IL-1B, TNF-α, IL-6, IL-8), vascular genes (VEGF, ANGPT1), cell cycle regulators (FOXL2, Cyclin D1, Cyclin D2, KLF4), and gonadotropin receptors (LHCGR, FSHR). Results: Results showed significantly higher expression of inflammatory cytokines in the granulosa cells of COVID-19 positive women, including IL-1B (4.2-fold), TNF-α (3.8-fold), IL-8 (2.5-fold), and IL-6 (3.2-fold). Vascular genes VEGF and ANGPT1 were also overexpressed, while FOXL2 was downregulated and Cyclin D1/D2 were upregulated in the study group. However, LH and FSH receptor expression remained similar between both groups. Conclusion: The present study demonstrates altered gene expression of inflammatory cytokines, vascular factors and cell cycle regulators in granulosa cells and oocytes of COVID-19 positive women undergoing ART. The dysregulated molecular pathways could potentially impair folliculogenesis and oocyte development in SARS-CoV-2 infected individuals.

Association of Donor Vascular Endothelial Growth Factor Gene Polymorphism With Acute Renal Allograft Rejection.

Recipient's VEGF-A polymorphisms have been reported to be associated with the risk of acute allograft rejection. However, an association of the donor's VEGF-A gene polymorphism with rejection remained unelucidated till now. In this study, VEGF-A gene SNPs at nine loci were analyzed in 160 kidney donors and recipients with rejection (rejectors, n=80) and without rejection (non-rejectors, n=80). Blood VEGF-A mRNA, plasma VEGF level, and intragraft VEGF expression were also analyzed by RT-PCR, ELISA, and immunohistochemistry, respectively. On comparing between the donor and rejectors, the polymorphic genotypes of VEGF -634 C>G [GG genotype, p<0.0001; OR (95% CI) = 17.74 (5.16-60.96)]; VEGF -1154 G>A [AG genotype, p<0.0001, OR (95% CI)=16.07 (3.68-70.15)]; VEGF +936 C>T [CT genotype, p<0.0001, OR (95% CI)=178.64 (23.28-1370.9), and TT genotype, p<0.0001; OR (95% CI)=3149 (278.91-35553)]; VEGF -1455 T>C [CC genotype, p value=0.0464, OR (95% CI) = 3.13 (1.07-9.10)]; VEGF -2578 C>A [CA genotype, p=0.0426, OR (95% CI)=4.62 (1.03-20.59), and AA genotype, p value<0.0001, OR (95% CI) = 21.89 (4.94-97.04)]; VEGF -2549 18bp Insertion/Deletion [ID genotype, p value<0.0001, OR (95% CI)=27.27 (3.61-205.73) and DD genotype, p value<0.0001, OR (95% CI)=25.18 (3.30-191.89) were significantly associated with acute rejection risk. On comparing rejectors versus non-rejectors, GA genotype of VEGF -1190 G>A and TC genotype of VEGF -1455 T>C were associated with the risk of rejection. On comparing donor VEGF between rejectors and non-rejectors, CG genotype of VEGF -634 C>G, AG of VEGF -1154 G>A; GA of VEGF -1190 G>A were associated with rejection. The blood VEGF-A mRNA and plasma VEGF levels were higher in the rejectors group compared to non-rejectors. Besides the recipient's VEGF SNPS, the donor's VEGF SNPs are also associated with the risk of acute rejection and may be closely monitored in evaluation to determine the risk of rejection.

Serum MicroRNA-126-3p and Serum Vascular Endothelial Growth Factor for Vision Threatening Diabetic Retinopathy in Type 2 Diabetes Mellitus Patients

Purpose: To investigate the precise function of miRNA-126-3p in controlling the expression of the VEGF gene and the signalling cascade that it is linked to in patients with type 2 diabetes Mellitus (DM). Study Design: Cross-sectional analytical study. Place and Duration of Study: Prof Ngoerah General Hospital as a tertiary hospital in Bali, Indonesia from January-April 2022 Methods: This study involved 35 vision-threatening diabetic retinopathy (VTDR) and 30 non-VTDR patients of type 2 DM. After diagnosis of DR by vitreoretinal specialist, blood samples were drawn. Real-time polymerase chain reaction assays were performed to examine miRNA-126-3p and ELISA to detect VEGF. Continuous data were analyzed using median (minimum-maximum range) and categorical data were analyzed using percentages. The statistical analysis was conducted using SPSS for Windows version 25.0.Thedifference and relationship between the proportions were tested by the Chi-square test and logistic regression test. Results: In case group, there were 23 subjects (65.7%) with low serum miRNA-126-3p expression, while in the control group, there were 7 subjects (23.3%) with low serum miRNA-126-3p expression (OR=6.3; 95% CI 2.1-18.86). In the case group, 20 individuals (57.1%) had high serum VEGF levels of 71.6 ng/L, whereas in the control group, there were 6 individuals (20.0%) with high serum VEGF levels. There was a statistically significant difference between the two groups (OR=5.33; 95% CI 1.75-16.3). Conclusion: The miRNA-126-3p expression and serum VEGF levels are the risk factors for VTDR development in patients with type 2 diabetes.

The Potential of Human Pulmonary Mesenchymal Stem Cells as Vectors for Radiosensitizing Metallic Nanoparticles: An In Vitro Study.

Metallic nanoparticles (NPs) exhibit interesting radiosensitizing effects, and finding a way to accurately deliver them appears to be crucial. Due to their tumor tropism, mesenchymal stem cells (MSCs) represent a strategic approach. Therefore, we aimed to evaluate the impact of core-shell Fe3O4@Au NPs on the functionality of human pulmonary MSCs (HPMSCs). The results showed that 100 µg/mL Fe3O4@Au NPs, accumulated in HPMSCs (revealed by Prussian blue staining), did not alter cell viability as assessed by cell counting, MTT, and LDH assays. However, caspase 9 and Bcl2 gene expression, evaluated by RT-qPCR, was regulated 72 h after exposure to the NPs. Moreover, the NPs also decreased proinflammatory cytokine/chemokine secretions, except for CXCL8 (ELISA). These modulations were associated with the downregulation of AMPK gene expression at 24 h. In contrast, the NPs did not modulate VEGF, PI3K, or PDGF gene expression. Nevertheless, a decrease in VEGF secretion was observed after 24 h of exposure to the NPs. Interestingly, the Fe3O4@Au NPs did not modulate Nrf2 gene expression, but they did regulate the expression of the genes encoding Nox4 and HMOX-1. Additionally, the NPs increased ROS production, suggesting a redox imbalance. Finally, the Fe3O4@Au NPs did not affect the HPMSCs' viability or proangiogenic/tumorigenic markers. These findings are encouraging for investigating the effects of Fe3O4@Au NPs delivered by HPMSCs to tumor sites in combination with radiation.

Effect of lyophilized exosomes derived from umbilical cord stem cells on chronic anterior cruciate ligament cell injury

BackgroundFacilitating the healing process of injured anterior cruciate ligament (ACL) tissue is crucial for patients to safely return to sports. Stem cell derived exosomes have shown positive effects on enhancing the regeneration of injured tendons/ligaments. However, clinical application of exosomes in terms of storage and pre-assembly is challenging. We hypothesized that lyophilized exosomes derived from human umbilical cord stem cells (hUSC-EX) could enhance the cell activity of chronically injured ACL cells.Materials and methodsWe harvested the 8 weeks injured ACL cells from rabbit under IACUC (No. 110232) approval. The studied exosomes were purified from the culture medium of human umbilical cord stem cells (IRB approval No. A202205014), lyophilized to store, and hydrated for use. We compared exosome treated cells with non-exosome treated cells (control group) from the same rabbits. We examined the cell viability, proliferation, migration capability and gene expression of type I and III collagen, TGFβ, VEGF, and tenogenesis in the 8 weeks injured ACL cells after hUSC-EX treatment.ResultsAfter hydration, the average size of hUSC-EX was 84.5 ± 70.6 nm, and the cells tested positive for the Alix, TSG101, CD9, CD63, and CD81 proteins but negative for the α-Tubulin protein. After 24 h of treatment, hUSC-EX significantly improved the cell viability, proliferation and migration capability of 8 weeks injured ACL cells compared to that of no exosome treatment group. In addition, the expression of collagen synthesis, TGFβ, VEGF, and tenogenesis gene were all significantly increased in the 8 weeks injured ACL cells after 24 h hUSC-EX delivery.DiscussionLyophilized exosomes are easily stored and readily usable after hydration, thereby preserving their characteristic properties. Treatment with lyophilized hUSC-EX improved the activity and gene expression of 8 weeks injured ACL cells.ConclusionLyophilized hUSC-EX preserve the characteristics of exosomes and can improve chronically injured (8 weeks) ACL cells. Lyophilized hUSC-EX could serve as effective and safe biomaterials that are ready to use at room temperature to enhance cell activity in patients with partial ACL tears and after remnant preservation ACL reconstruction.

VEGFA gene variants are associated with breast cancer progression.

Angiogenesis is recognized as a hallmark of cancer, and vascular endothelial growth factor (VEGF) is a key regulator of the angiogenic process and is related to cancer progression. Anti-VEGF therapy has been tried but with limited success and without useful stratification for angiogenesis markers. Further, the landscape of VEGF single nucleotide polymorphisms (SNPs) in breast cancer and their clinical relevance is not well studied, and their relation to tissue-based angiogenesis markers has not been explored. Here, we studied a selection of VEGFA SNPs in nontumor lymph nodes from a population-based breast cancer cohort (n = 544), and their relation to clinicopathologic variables, vascular tissue metrics, and breast cancer-specific survival. Two of the SNP candidates (rs833068GA genotype and rs25648CC genotype) showed associations with angiogenesis tissue markers, and the VEGFA rs833068GA genotype was associated with breast cancer-specific survival among ER-negative cases. We also found trends of association between the rs699947CA genotype and large tumor diameter and ER-negative tumors, and between the rs3025039CC genotype and large tumor diameter. Our findings indicate some associations between certain VEGF SNPs, in particular the rs833068GA genotype, and both vascular metrics and patient survival. These findings and their potential implications need to be validated by independent studies.

In Silico Characterization of Inflammatory and Anti-Inflammatory Modulation in Diabetic Nephropathy: The Construction of a Genetic Panel

Diabetic Nephropathy (DN) stands as a primary cause of end-stage renal disease and its etiology remains unclear. Thus, this study aims to construct a genetic panel with potential biomarkers linked to the inflammatory pathway of DN associated with the pathology’s susceptibility. Through a systematic review and meta-analysis, we selected observational studies in English, Portuguese, and Spanish, selected from the PubMed, SCOPUS, Virtual Health Library, Web of Science, and EMBASE databases. Additionally, a protein–protein interaction network was constructed to list hub genes, with differential expression analysis by microarray of kidneys with DN from the GSE30529 database to further refine results. Seventy-two articles were included, and 54 polymorphisms in 37 genes were associated with the inflammatory pathway of DN. Meta-analysis indicated a higher risk of complication associated with SNPs 59029 G/A, −511 C/T, VNTR 86 bp, −308 G/A, and −1031 T/C. Bioinformatics analyses identified differentially expressed hub genes, underscoring the scarcity of studies on CCL2 and VEGF-A genes in relation to DN. This study highlighted the intrinsic relationship between inflammatory activity in the etiology and progression of DN, enabling the effective application of precision medicine in diabetic patients for potential prognosis of the complications and contributing to cost reduction in the public health system.

Genetic structure analysis of yak breeds and their response to adaptive evolution

Yaks are crucial genetic resources in the Tibetan Plateau and surrounding regions. Throughout the long process of domestication, natural and artificial selection pressures have enabled yaks to demonstrate adaptive characteristics to the environment in terms of physiological structure and genetic molecules, but no systematic cell analysis has been carried out on this phenomenon of yaks. Here, the population structure and genetic diversity of yak were studied by WGRS, and the genes related to yak adaptability were excavated. Combined with scRNA-seq method, the transcription map of yak lung tissue and skin tissue was constructed, which provided a new comprehensive insight into yak adaptability. The analysis of yak population structure showed that there was obvious genetic differentiation between TZ _ yak and other seven yak populations, while there was significant genetic exchange between PL _ yak and SB _ yak at high altitude. WGRS and scRNA-seq analysis revealed that the gene HIF1A related to high altitude adaptation was expressed in various cell types, while EPAS1 was predominantly expressed in epithelial and endothelial cells of yak lung tissue. Endothelial cells play a critical role in hypoxia-adapted VEGF signaling, which correlates closely with the high expression of KDR and VEGFA genes in endothelial cells and monocytes. Furthermore, in the selection signal of High _ yak vs Low _ yak, 19.8 % of the genes overlapped with the genes screened by skin scRNA-seq, including genes related to coat color such as RORA, BNC2, and KIT. Notably, BNC2 is a gene associated with melanin deposition and shows high expression levels in HS cells. Additionally, GRN in melanocytes and SORT1 in IRS play an important role in cell communication between melanocytes and IRS. These findings offer new insights into the natural polymorphism of yaks and provide a valuable reference for future research on high-altitude mammals, and potentially even human genetics.