Platelet-derived Growth factor-BB Research Articles

Downregulation of LILRB4 Promotes Human Aortic Smooth Muscle Cell Contractile Phenotypic Switch and Apoptosis in Aortic Dissection.

Aortic dissection (AD) is a severe vascular disease with high rates of mortality and morbidity. However, the underlying molecular mechanisms of AD remain unclear. Differentially expressed genes (DEGs) were screened by bioinformatics methods. Alterations of histopathology and inflammatory factor levels in β-aminopropionitrile (BAPN)-induced AD mouse model were evaluated through Hematoxylin-Eosin (HE) staining and Enzyme-linked immunosorbent assay (ELISA), respectively. Reverse transcription quantitative real-time polymerase chain reaction was performed to detect DEGs expression. Furthermore, the role of LILRB4 in AD was investigated through Cell Counting Kit-8 (CCK-8), wound healing, and flow cytometry. Western blotting was employed to assess the phenotypic switch and extracellular matrix (ECM)-associated protein expressions in platelet-derived growth factor-BB (PDGF-BB)-stimulated in vitro model of AD. In the AD mouse model, distinct dissection formation was observed. TNF-α, IL-1β, IL-8, and IL-6 levels were higher in the AD mouse model than in the controls. Six hub genes were identified, including LILRB4, TIMP1, CCR5, CCL7, MSR1, and CLEC4D, all of which were highly expressed. Further exploration revealed that LILRB4 knockdown inhibited the cell vitality and migration of PDGF-BB-induced HASMCs while promoting apoptosis and G0/G1 phase ratio. More importantly, LILRB4 knockdown promoted the protein expression of α-SMA and SM22α, while decreasing the expression of Co1, MMP2, and CTGF, which suggested that LILRB4 silencing promoted contractile phenotypic transition and ECM stability. LILRB4 knockdown inhibits the progression of AD. Our study provides a new potential target for the clinical treatment of AD.

Platelet-derived growth factor signaling in pericytes promotes hypothalamic inflammation and obesity.

Pericytes are a vital component of the blood-brain barrier, and their involvement in acute inflammation was recently suggested. However, it remains unclear whether pericytes contribute to hypothalamic chronic inflammation and energy metabolism in obesity. The present study investigated the impact of pericytes on the pathophysiology of obesity by focusing on platelet-derived growth factor (PDGF) signaling, which regulates pericyte functions. Tamoxifen-inducible systemic conditional PDGF receptor β knockout mice (Pdgfrb∆SYS-KO) and Calcium/calmodulin-dependent protein kinase type IIa (CaMKIIa)-positive neuron-specific PDGF receptor β knockout mice (Pdgfrb∆CaMKII-KO) were fed a high-fat diet, and metabolic phenotypes before and 3 to 4weeks after dietary loading were examined. Intracellular energy metabolism and relevant signal transduction in lipopolysaccharide- and/or platelet-derived growth factor-BB (PDGF-BB)-stimulated human brain pericytes (HBPCs) were assessed by the Seahorse XFe24 Analyzer and Western blotting. The pericyte secretome in conditioned medium from HBPCs was studied using cytokine array kit, and its impact on polarization was examined in bone marrow-derived macrophages (BMDMs), which are microglia-like cells. Energy consumption increased and body weight gain decreased after high-fat diet loading in Pdgfrb∆SYS-KO mice. Cellular oncogene fos (cFos) expression increased in proopiomelanocortin (POMC) neurons, whereas microglial numbers and inflammatory gene expression decreased in the hypothalamus of Pdgfrb∆SYS-KO mice. No significant changes were observed in Pdgfrb∆CaMKII-KO mice. In HBPCs, a co-stimulation with lipopolysaccharide and PDGF-BB shifted intracellular metabolism towards glycolysis, activated mitogen-activated protein kinase (MAPK), and modulated the secretome to the inflammatory phenotype. Consequently, the secretome showed an increase in various proinflammatory chemokines and growth factors including Epithelial-derived neutrophil-activating peptide 78 (C-X-C motif chemokine ligand (CXCL)5), Thymus and activation-regulated chemokine (C-C motif chemokine (CCL)17), Monocyte chemoattractant protein 1 (CCL2), and Growth-regulated oncogene α (CXCL1). Furthermore, conditioned medium from HBPCs stimulated the inflammatory priming of BMDMs, and this change was abolished by the C-X-C motif chemokine receptor (CXCR) inhibitor. Consistently, mRNA expression of CXCL5 was elevated by lipopolysaccharide and PDGF-BB treatment in HBPCs, and the expression was significantly lower in the hypothalamus of Pdgfrb∆SYS-KO mice than in control Pdgfrbflox/flox mice (FL) following 4 weeks of HFD feeding. PDGF receptor β signaling in hypothalamic pericytes promotes polarization of macrophages by changing their secretome and contributes to the progression of obesity.

Abstract A085: Stromal PDGFRβ hyperactivation modulates the myeloid lineage in the brain metastatic microenvironment

Abstract Breast cancer is the most common cancer diagnosis worldwide, and an unfortunately high number of women die every year due to a lack of effective therapeutic strategies for metastatic breast cancer. Patients with brain metastases in particular have a significantly poorer prognosis than women with metastasis to other sites (e.g., lung, liver, bone). Previous studies by our group identified a ligand-receptor complex in the tumor microenvironment (TME) that promotes metastasis of breast cancer to the brain. This interaction between platelet-derived growth factor-BB (PDGFB), produced by cancerous breast epithelial cells, and its receptor, platelet-derived growth factor receptor beta (PDGFRβ), expressed by mesenchymal cells, is also prognostic of breast cancer metastasis to the brain in patients. The mechanism of how the PDGFB-PDGFRβ pathway mediates brain metastasis is still unclear and is the focus of our current work. To further analyze the biological effects of PDGFB, our group evaluated gene expression changes between primary human breast tumors expressing high PDGFB versus those expressing low PDGFB and found significantly more immune signaling in the low-expressing tumors. These results led us to hypothesize that the PDGFB-PDGFRβ pathway creates an immunosuppressive microenvironment, allowing for increased metastasis of cancerous cells. To test this, flow cytometry was completed on the brains, spleens, lungs, and livers of control mice and mutant mice expressing stromal-specific hyperactive PDGFRβ following intracardiac injection of PDGFB-expressing mammary tumor cells. No significant changes in immune cell populations were observed in non-tumor-bearing mice. However, in the brains of tumor-bearing mutant mice, there are significant changes to the myeloid lineage. Specifically, analysis at day 10 post-injection (prior to detectable metastatic lesions) revealed diminished CD45+CD11b+ populations. Segregating by CD45hi/CD45lo confirmed a profound decrease in CD45loCD11b+F4/80+ microglia, likely a tumor-suppressive population. Looking at day 16 post-injection (detectable metastatic lesions), there is a shift towards an increase in CD45+ CD11b+GR1+ infiltrating myeloid-derived suppressor cells (MDSCs) in experimental mice, likely a tumor-promoting population. Minimal changes were observed in the lungs and livers at any time point. Confirmatory immunofluorescence staining showed a decrease in tumor-infiltrating microglia (Iba1+) and a significant increase in tumor-infiltrating peripheral macrophages (F4/80+), mirroring the flow data. This experiment was then repeated to test the contribution of the ligand (PDGFB), where a significant increase of infiltrating microglia (Iba1+) was observed upon tumor cell knockdown of PDGFB. Together, these results suggest that the PDGFB-PDGFRβ pathway creates an immunosuppressive environment in the brain TME at least in part through reorchestration of the myeloid populations. Citation Format: Alexis A Mossing, Johnathon G Schiebel, Rebecca L Packard, Amrendra Kumar, Sarah A Steck, Nathaniel S Grabinski, Katie A Thies, Bri Wasik, Tasneem Ariswala, Gary Tozbikian, Johnathan Godbout, Steven T Sizemore, Paul R Lockman, Anna E Vilgelm, Gina M Sizemore. Stromal PDGFRβ hyperactivation modulates the myeloid lineage in the brain metastatic microenvironment [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A085.

Investigating the molecular mechanism of traditional Chinese medicine for the treatment of placental syndromes by influencing inflammatory cytokines using the Mendelian randomization and molecular docking technology.

Placental syndromes, which include pregnancy loss, preterm birth, gestational diabetes mellitus (GDM), and hypertensive disorders in pregnancy (HDP), have a strong association with disorder inflammatory reactions. Nonetheless, the exact causal relationship has not been established. This study aims to investigate the causal relationship between placental syndromes and inflammatory cytokines utilizing Mendelian randomization (MR). Additionally, we examined the interaction between small molecular compounds derived from traditional Chinese medicine and inflammatory cytokines using molecular docking method. After obtaining the data of inflammatory cytokines and placental syndromes, as well as establishing single nucleotide polymorphisms (SNPs), we employed the inverse variance weighted (IVW) method to assess the causal relationship. We also accessed the heterogeneity and the horizontal pleiotropy of these data. The "ClusterProfiler" R package was utilized for Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) term analyses. The protein-protein interaction (PPI) network was constructed using STRING database. AutoDock Vina software was used for molecular docking, and Discovery Studio 2019 was used for visualization purposes. We found that the growth regulated oncogene A (GROA) and interleukin-9 (IL-9) were associated with the development of pregnancy hypertension, whereas interleukin-10 (IL-10) and hepatocyte growth factor (HGF) were linked to the occurrence of preeclampsia. Moreover, there were correlations observed between interleukin-18 (IL-18), IL-10, macrophage colony-stimulating factor (MCSF), and platelet-derived growth factor BB (PDGFbb) in cases of chronic hypertension combined with pregnancy (CHP). Additionally, macrophage migration inhibitory factor (MIF) exhibited a connection with GDM, and TNF related apoptosis inducing ligand (TRAIL) demonstrated a causal relationship with preterm birth. It is plausible to suggest that interleukin-1β (IL-1β) might contribute to the promotion of pregnancy loss. All of the binding free energy values of small molecular compounds with inflammatory cytokines were below -5.0 kcal/mol. Furthermore, all of the RMSD values were less than 2. GROA, IL-1β, IL-9, IL-10, IL-18, MIF, MCSF, HGF, PDGFbb and TRAIL were found to be causally associated with placental syndromes. Molecular docking analysis revealed that small molecular compounds, such as puerarin, magnolol, atractylenolide I, paeoniflorin, tumulosic acid and wogonin, are closely bound to these inflammatory cytokines.

Neonatal stress disrupts the glymphatic system development and increases the susceptibility to Parkinson's disease in later life.

Neonatal stress disrupts brain development and increases the risk of neurological disorders later in life. However, the impact of neonatal stress on the development of the glymphatic system and susceptibility to Parkinson's disease (PD) remains largely unknown. Neonatal maternal deprivation (NMD) was performed on mice for 14 consecutive days to model chronic neonatal stress. Adeno-associated virus expressing A53T-α-synuclein (α-syn) was injected into the substantia nigra to establish PD model mice. Glymphatic activity was determined using invivo magnetic resonance imaging, exvivo fluorescence imaging and microplate assay. The transcription and expression of aquaporin-4 (AQP4) and other molecules were evaluated by qPCR, western blotting, and immunofluorescence. Animal's responses to NMD and α-syn overexpression were observed using behavioral tests. Glymphatic activity was impaired in adult NMD mice. AQP4 polarization and platelet-derived growth factor B (PDGF-B) signaling were reduced in the frontal cortex and hippocampus of both young and adult NMD mice. Furthermore, exogenous α-syn accumulation was increased and PD-like symptoms were aggravated in adult NMD mice. The results demonstrated that NMD could disrupt the development of the glymphatic system through PDGF-B signaling and increase the risk of PD later in life, indicating that alleviating neonatal stress could be beneficial in protecting the glymphatic system and reducing susceptibility to neurodegeneration.

Hsa_circ_0032389 Enhances Proliferation and Migration in PDGF-BB-Induced Human Aortic Vascular Smooth Muscle Cells.

Circular RNA (circRNAs) has been confirmed to participate in atherosclerosis (AS) progression. However, the role and mechanism of hsa_circ_0032389 in AS process still need to be further revealed. This study evaluates the role and mechanism of hsa_circ_0032389 in AS process. Platelet-derived growth factor-BB (PDGF-BB) was used to induce human aortic vascular smooth muscle cells (HA-VSMCs). The expression levels of hsa_circ_0032389, microRNA (miR)-513a-5p, and fibroblast growth factor receptor substrate 2 (FRS2) were examined by quantitative real-time PCR. Cell proliferation and migration were analyzed using cell counting kit 8 assay, flow cytometry, EdU assay, transwell assay, and wound healing assay. Protein expression was assessed using western blot analysis. Dual-luciferase reporter and RIP assays were used to confirm RNA interaction. Hsa_circ_0032389 was overexpressed in PDGF-BB-induced HA-VSMCs, and its downregulation inhibited HA-VSMC viability, cell cycle, EdU positive cell rate, migratory cell number, and wound closure rate under PDGF-BB treatment. The luciferase activity of hsa_circ_0032389wt could be reduced by miR-513a-5p mimic, and both hsa_circ_0032389 and miR-513a-5p were enriched in anti-Ago2, confirming that miR-513a-5p could be sponged by hsa_circ_0032389. MiR-513a-5p inhibitor reversed the effect of hsa_circ_0032389 knockdown on PDGF-BB-induced HA-VSMC viability, cell cycle, EdU positive cell rate, migratory cell number, and wound closure rate. Moreover, the luciferase activity of FRS2wt was reduced by miR-513a-5p mimic, and both FRS2 and miR-513a-5p were enriched in anti-Ago2, verifying that FRS2 was targeted by miR-513a-5p. MiR-513a-5p suppressed PDGF-BB-induced HA-VSMC viability, cell cycle, EdU positive cell rate, migratory cell number, and wound closure rate by targeting FRS2. Our results indicated that hsa_circ_0032389 enhanced PDGF-BB-induced HA-VSMC proliferation and migration via regulating miR-513a-5p/FRS2 axis.

Effect of Platelet-Rich Plasma at Different Initiation Times on Healing of the Bone-Tendon Interface of the Rotator Cuff in a Mouse Model.

Platelet-rich plasma (PRP) has demonstrated beneficial effects on healing of the bone-tendon interface (BTI). To determine the optimal initiation time for PRP application after rotator cuff repair in an animal model. Controlled laboratory study. A total of 136 C57BL/6 mice were included; 40 mice were used to prepare PRP, while 96 mice underwent acute supraspinatus tendon (SST) repair. The animals were randomly divided into 4 groups: a control group and 3 groups in which PRP was injected into the injury interface immediately after surgery, on the 7th postoperative day (PRP-7d), and on the 14th postoperative day. At 4 and 8 weeks postoperatively, the animals were sacrificed, blood was collected by eyeball removal, and samples of the SST-humerus complex were collected. Histological, imaging, immunological, and biomechanical data were compared among the groups using 1-way analysis of variance with the Bonferroni post hoc test. Histological analysis revealed that the fibrocartilage layer at the BTI was larger in the PRP-7d group compared to the other groups at both 4 and 8 weeks postoperatively. Moreover, the PRP-7d group exhibited improved proteoglycan content and distribution compared to the other groups. Enzyme-linked immunosorbent assay results demonstrated that at 4 weeks postoperatively, higher concentrations of transforming growth factor-β1 and platelet-derived growth factor-BB (PDGF-BB) were seen in the PRP-7d group versus the PRP-14d and control gruops (P < .05), and at 8 weeks postoperatively, the concentration of PDGF-BB was higher in the PRP-7d group versus the control group (P < .05). Biomechanical testing at 4 weeks postoperatively revealed that the failure load and ultimate strength of the SST-humerus complex were superior in the PRP-7d group compared to the other groups (P < .05), at 8 weeks, PRP-7d group was superior to the control group (P < .05). Additionally, at 8 weeks postoperatively, the PRP-7d group exhibited a greater trabecular number and trabecular thickness at the BTI compared to the PRP-14d and control gruops (P < .05). PRP promoted healing of the BTI after a rotator cuff injury at an early stage. A PRP injection on the 7th postoperative day demonstrated superior therapeutic effects compared with injections at other time points.

The effect of bta-miR-1296 on the proliferation and extracellular matrix synthesis of bovine mammary fibroblasts.

Mammary fibrosis in dairy cows is a chronic condition caused by mastitis, and can lead to serious culling of dairy cows resulting in huge economic losses in the dairy industry. MicroRNAs (miRNAs) exert an important role in regulating mammary gland health in dairy cows. This study investigated whether exosomal miRNAs in mammary epithelial cells can regulate the proliferation of bovine mammary fibroblasts (BMFBs) in mastitis. Liposome transfection technology was used to construct a cellular model of the overexpression and inhibition of miRNAs. The STarMir software, dual luciferase reporter gene test, real-time quantitative PCR (qRT-PCR), a Cell Counting Kit-8 (CCK-8), and a Western Blot and plate clone formation test were used to investigate the mechanism by which bta-miR-1296 regulates the proliferation of BMFBs. Target gene prediction results revealed that glutamate-ammonia ligase was a direct target gene by which bta-miR-1296 regulates cell proliferation. It was found that bta-miR-1296 significantly inhibited the proliferation of BMFBs. After BMFBs were transfected with a bta-miR-1296 mimic, mRNA expression in the extracellular matrix (ECM), α-smooth muscle actin (α-SMA), collagen type I alpha 1 chain (COL1α1) and collagen type III alpha 1 chain (COL3α1), and various cell growth factors (basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), platelet-derived growth factor-BB (PDGF-BB), and transforming growth factor-β1 (TGF-β1)) were down-regulated, and the expressions of α-SMA, COL1α1, COL3α1, phospho-extracellular regulated protein kinases, phospho-protein kinaseB, TGF-β1, and phospho-Smad family member3 proteins were inhibited. In conclusion, bta-miR-1296 can inhibit the proliferation of BMFBs and the synthesis of ECM in BMFBs, thus affecting the occurrence and development of mammary fibrosis in dairy cows and laying the foundation for further studies to clarify the regulatory mechanism of mammary fibrosis.

Developmental morphogens direct human induced pluripotent stem cells toward an annulus fibrosus-like cell phenotype.

Therapeutic interventions for intervertebral disc herniation remain scarce due to the inability of endogenous annulus fibrosus (AF) cells to respond to injury and drive tissue regeneration. Unlike other orthopedic tissues, such as cartilage, delivery of exogenous cells to the site of annular injury remains underdeveloped, largely due to a lack of an ideal cell source and the invasive nature of cell isolation. Human induced pluripotent stem cells (iPSCs) can be differentiated to specific cell fates using biochemical factors and are, therefore, an invaluable tool for cell therapy approaches. While differentiation protocols have been developed for cartilage and fibrous connective tissues (e.g., tendon), the signals that regulate the induction and differentiation of human iPSCs toward the AF fate remain unknown. iPSC-derived sclerotome cells were treated with various combinations of developmental signals including transforming growth factor beta 3 (TGF-β3), connective tissue growth factor (CTGF), platelet derived growth factor BB (PDGF-BB), insulin-like growth factor 1 (IGF-1), or the Hedgehog pathway activator, Purmorphamine, and gene expression changes in major AF-associated ECM genes were assessed. The top performing combination treatments were further validated by using three distinct iPSC lines and by assessing the production of upregulated ECM proteins of interest. To conduct a broader analysis of the transcriptomic shifts elicited by each factor combination, and to compare genetic profiles of treated cells to mature human AF cells, a 96.96 Fluidigm gene expression array was applied, and principal component analysis was employed to identify the transcriptional signatures of each cell population and treatment group in comparison to native AF cells. TGF-β3, in combination with PDGF-BB, CTGF, or IGF-1, induced an upregulation of key AF ECM genes in iPSC-derived sclerotome cells. In particular, treatment with a combination of TGF-β3 with PDGF-BB for 14 days significantly increased gene expression of collagen II and aggrecan and increased protein deposition of collagen I and elastin compared to other treatment groups. Assessment of genes uniquely highly expressed by AF cells or SCL cells, respectively, revealed a shift toward the genetic profile of AF cells with the addition of TGF-β3 and PDGF-BB for 14 days. These findings represent an initial approach to guide human induced pluripotent stem cells toward an AF-like fate for cellular delivery strategies.

S-propargyl-cysteine promotes the stability of atherosclerotic plaque via maintaining vascular muscle contractile phenotype.

Introduction: Plaque rupture in atherosclerosis contributes to various acute cardiovascular events. As a new sulfide-containing donor, S-propargyl-cysteine (SPRC) has been reported to play a beneficial role in cardioprotection, potentially through its anti-inflammatory, anti-oxidative and anti-atherogenic activities. Our previous study observed an increase in eNOS phosphorylation in endothelial cells. However, it remains unclear whether SPRC influences vascular smooth muscle cells (VSMCs) within the plaque and if this effect contributes to plaque stabilization. Methods: An atherosclerotic unstable plaque mouse model was established by subjecting ApoE-/- mice to tandem stenosis of the right carotid artery along with a Western diet. Daily SPRC administration was conducted for 13weeks. Plaque morphology and stability were assessed using MRI scanning and histopathological staining. In our in vitro studies, we stimulated human artery vascular smooth muscle cells (HAVSMCs) with platelet-derived growth factor-BB (PDGF-BB), both with and without 100μM SPRC treatment. Cell phenotype was assessed using both Western blot and Real-time PCR. Cell proliferation was assessed using the BrdU cell proliferation kit and immunofluorescence of Ki-67, while cell migration was measured using scratch wound healing and transwell assay. MiR-143-3p overexpression and knockdown experiments were used to investigate whether it mediates the effect of SPRC on VSMC phenotype. Results and Discussion: SPRC treatment reduced plasma lipid levels, increased collagen content and decreased cell apoptosis in atherosclerotic plaques, indicating improved plaque stability. Both in vivo and in vitro studies elucidated the role of SPRC in preserving the contractile phenotype of VSMCs through up-regulation of miR-143-3p expression. Furthermore, SPRC suppressed the pro-proliferation and pro-migration effects of PDGF-BB on HAVSMCs. Overall, these findings suggest that the inhibitory effect of SPRC on phenotype switch from contractile to synthetic VSMCs may contribute to its beneficial role in enhancing plaque stability.

Causal associations between circulating inflammatory cytokines and blinding eye diseases: a bidirectional Mendelian randomization analysis.

Previous studies have explored the associations between circulating inflammatory cytokines and blinding eye diseases, including glaucoma, cataract and macular degeneration. However, the causality of these associations remains controversial. This study employs a bidirectional Mendelian randomization (MR) study to investigate the causal relationships between 41 circulating inflammatory cytokines and these blinding eye diseases. Summary data for glaucoma, cataract, macular degeneration and 41 circulating inflammatory cytokines were publicly available. The inverse variance weighted (IVW) method was employed as the main analysis method. Additionally, various sensitivity tests, including MR-Egger regression, weighted median, weight mode, Cochran's Q test, MR pleiotropy Residual Sum and Outlier test, and leave-one-out test, were conducted to evaluate sensitivity and stability of results. The IVW analysis identified six circulating inflammatory cytokines causally associated with the risk of blinding eye diseases: Monokine induced by interferon-gamma (MIG) for glaucoma, interleukin-1 receptor antagonist (IL-1ra), IL-6, IL-10, and platelet derived growth factor BB (PDGFbb) for cataract, and MIG and hepatocyte growth factor (HGF) for macular degeneration. However, it is noteworthy that none of these associations remained significant after Bonferroni correction (p < 0.0004). Reverse MR analyses indicated that cataract may lead to a decrease in vascular endothelial growth factor (VEGF) levels (OR: 3.326 × 10-04, 95% CI: 5.198 × 10-07 - 2.129 × 10-01, p = 0.0151). This study highlights the potential roles of specific inflammatory cytokines in the development of glaucoma, cataract and macular degeneration. Moreover, it suggests that VEGF is likely to be involved in cataract development downstream. These findings offer insights for early prevention and novel therapeutic strategies for these blinding eye diseases.

Characterization of platelet rich plasma in feline immunodeficiency virus-infected cats: Cell, and PDGF-BB and TGF-ß1 growth factor analysis

Autologous platelet-rich plasma (PRP) contains growth factors (GFs) that modulate the expression of inflammatory cells; thus, these products could be considered a good strategy to favor tissue regeneration in feline immunodeficiency (FIV) positive cats. However, there is no scientific documentation on obtaining PRP in FIV-positive cats.Authors hypothesized that PRP can be obtained in FIV cats following the PRGF®-Endoret® methodology. The objectives of this study were to compare the platelet, erythrocyte, and leukocyte concentration between whole blood (WB) and the PRP; and determine the concentration of platelet-derived growth factor BB (PDGF-BB) and transforming growth factor β1 (TGF-β1) in FIV-positive cats. Sixteen adults FIV-positive asymptomatic cats were included in the study. WB samples were drawn and the PRP was obtained by centrifugation at 265g for 10 min. Erythrocyte and leukocyte, platelets, and mean platelet volume (MPV) were determined both in WB and in PRP. PDGF-BB and TGF-β1 concentrations were additionally determined in PRP.Platelet concentration increased 1.1 times in PRP fraction compared to WB, but no significant differences were reported. MPV was statistically higher in WB than in PRP (p = 0.001). Erythrocytes and leukocytes counts were decreased by 99% and 92%, respectively in the PRP fraction (p < 0.001). Regarding TGF-ß1, a higher concentration was shown in the PRP (p < 0.02). Although the product obtained could not be classified as PRP according to the PRGF®-Endoret® methodology, based on the drastic reduction of RBC and WBC, the PLT concentrate is of high purity.

WNTA5-mediated miR-374a-5p regulates vascular smooth muscle cell phenotype transformation and M1 macrophage polarization impacting intracranial aneurysm progression

miR-374a-5p expression and localization in intracranial aneurysm (IA) tissues were detected, and its correlation with vascular smooth muscle cells (VSMCs) and macrophage markers was analyzed. Using platelet-derived growth factor-BB (PDGF-BB) induced VSMC model, elastase-induced IA rat model. Subsequently, miR-374a-5p was knocked down or overexpressed. We investigated the effects of miR-374a-5p on phenotypic conversion, and in vivo experiments were also carried out to verify the findings. The targeted relationship between miR-374a-5p and WNTA5 was analyzed. The effect of WNT5A inhibition on VSMC phenotypic transformation and THP-1-derived macrophage polarization was explored. Clinical studies have shown that miR-374a-5p was upregulated in IA patients. miR-374a-5p was negatively correlated with SM22α, α-SMA, CD206, and positively correlated with CD86. In vitro experiments showed that knocking down miR-374a-5p reversed the promotion of SM22α and α-SMA expression by PDGF-BB, while overexpression of miR-374a-5p had the opposite effect. In addition, knocking down miR-374a-5p also reversed the decrease in Calponin, TIMP3, TIMP4, and IL-10 levels caused by PDGF-BB, and further reduced the levels of MMP1, MMP3, MMP9, IL-1β, IL-6, and TNF-α. These findings were further validated in vivo. In IA rats, there were notable increases in both systolic and diastolic blood pressure, along with an elevated M1/M2 ratio and the occurrence of vascular lesions. However, these symptoms were improved after knocking down miR-374a-5p. Furthermore, miR-374a-5p could target the WNT signals (WNT2B, WNT3, and WNT5A). miR-374a-5p regulated the VSMC phenotypic conversion and M1 macrophage polarization by targeting WNT5A, thereby impacting the progression of IA.

2-N, 6-O sulfated chitosan evokes periosteal stem cells for bone regeneration

Musculoskeletal injuries and bone defects represent a significant clinical challenge, necessitating innovative approaches for effective bone tissue regeneration. In this study, we investigated the potential of harnessing periosteal stem cells (PSCs) and glycosaminoglycan (GAG)-mimicking materials for in situ bone regeneration. Our findings demonstrated that the introduction of 2-N, 6-O sulfated chitosan (26SCS), a GAG-like polysaccharide, enriched PSCs and promoted robust osteogenesis at the defect area. Mechanistically, 26SCS amplifies the biological effect of endogenous platelet-derived growth factor-BB (PDGF-BB) through enhancing the interaction between PDGF-BB and its receptor PDGFRβ abundantly expressed on PSCs, resulting in strengthened PSC proliferation and osteogenic differentiation. As a result, 26SCS effectively improved bone defect repair, even in an osteoporotic mouse model with lowered PDGF-BB level and diminished regenerative potential. Our findings suggested the significant potential of GAG-like biomaterials in regulating PSC behavior, which holds great promise for addressing osteoporotic bone defect repair in future applications.

A bilayer bioengineered patch with sequential dual-growth factor release to promote vascularization in bladder reconstruction.

Bladder tissue engineering holds promise for addressing bladder defects resulting from congenital or acquired bladder diseases. However, inadequate vascularization significantly impacts the survival and function of engineered tissues after transplantation. Herein, a novel bilayer silk fibroin (BSF) scaffold was fabricated with the capability of vascular endothelial growth factor (VEGF) and platelet derived growth factor-BB (PDGF-BB) sequential release. The outer layer of the scaffold was composed of compact SF film with waterproofness to mimic the serosa of the bladder. The inner layer was constructed of porous SF matrix incorporated with SF microspheres (MS) loaded with VEGF and PDGF-BB. We found that the 5% (w/v) MS-incorporated scaffold exhibited a rapid release of VEGF, whereas the 0.2% (w/v) MS-incorporated scaffold demonstrated a slow and sustained release of PDGF-BB. The BSF scaffold exhibited good biocompatibility and promoted endothelial cell migration, tube formation and enhanced endothelial differentiation of adipose derived stem cells (ADSCs) in vitro. The BSF patch was constructed by seeding ADSCs on the BSF scaffold. After in vivo transplantation, not only could the BSF patch facilitate the regeneration of urothelium and smooth muscle, but more importantly, stimulate the regeneration of blood vessels. This study demonstrated that the BSF patch exhibited excellent vascularization capability in bladder reconstruction and offered a viable functional bioengineered patch for future clinical studies.

Tetramethylpyrazine regulates angiogenesis of endothelial cells in cerebral ischemic stroke injury via SIRT1/VEGFA signaling pathway

This study aims to investigate whether tetramethylpyrazine(TMP) can stimulate angiogenesis in cerebral microvascular endothelial cells and alleviate cerebral ischemic stroke(CIS) and to explore the underlying mechanisms. In the animal study, adult Sprague-Dawley rats(n=15) were assigned into sham surgery(sham), middle cerebral artery occlusion/reperfusion(MCAO/R), and MCAO/R+TMP(intraperitoneal injection of 20 mg·kg~(-1)) groups. The neurological function was evaluated by the Z-Longa method. The cerebral infarction volume was detected by TTC staining. Enzyme-linked immunosorbent assay(ELISA) was employed to detect the expression of vascular endothelial growth factor(VEGF), angiopoietin(Ang), and platelet-derived growth factor(PDGF). Immunofluorescence staining was employed to detect Ki67 and the expression of vascular endothelial growth factor A(VEGFA) and slient information regulator 1(SIRT1). Western blot was employed to determine the expression levels of VEGFA, SIRT1, angiopoietin-2(Ang-2), and platelet-derived growth factor B(PDGFB). In the cell study, mouse brain-derived endothelial cells(Bend.3) were cultured, and the optimal concentration of TMP was determined. Then, VEGF, Ang, and PDGF were detected by ELISA after the addition of cabozantinib. Western blot was employed to measure the expression of VEGFA, Ang-2, and PDGFB. Immunofluorescence staining was used to detect CD31, CD34, and Ki67, and the proliferation, migration, and tube formation ability of Bend.3 cells were observed in vitro. Western blot and immunofluorescence staining were performed to measure the expression of SIRT1 and VEGFA after addition of the SIRT1-specific inhibitor selisistat(EX-527). The results showed that compared with the sham group, the MCAO/R group had severe neurological function damage, increased infarction volume, up-regulated expression of VEGF, VEGFA, Ang, Ang-2, PDGF, and PDGFB, and down-regulated expression of Ki67 and SIRT1(P&lt;0.01). Compared with the MCAO/R group, the MCAO/R+TMP group presented alleviated neurological function damage, reduced infarction volume, and activated expression of VEGF, VEGFA, Ang, Ang-2, PDGF, PDGFB, Ki67, and SIRT1(P&lt;0.01). The cell experiments showed that compared with the normal group, Bend.3 cells were activated by oxygen glucose deprivation/reoxygenation(OGD/R) treatment(P&lt;0.05, P&lt;0.01). Compared with the OGD/R group, the OGD/R+TMP group upregulated the expression levels of VEGF, VEGFA, Ang, Ang-2, PDGF, PDGFB, SIRT1, Ki67, CD31, and CD34, enhanced the angiogenic ability of Bend.3 cells without being inhibited by BMS or EX-527(P&lt;0.05, P&lt;0.01, P&lt;0.001). The results suggest that TMP can activate the SIRT1/VEGFA signaling pathway to stimulate angiogenesis and alleviate CIS injury.

MiR-411-5p Promotes Vascular Smooth Muscle Cell Phenotype Switch by Inhibiting Expression of Calmodulin Regulated Spectrin-Associated Protein-1.

When stimulated, vascular smooth muscle cells (VSMCs) change from a differentiated to a dedifferentiated phenotype. Dedifferentiated VSMCs have a key activity in cardiovascular diseases such as in-stent restenosis. MicroRNAs (miRNAs) have crucial functions in conversion of differentiated VSMCs to a dedifferentiated phenotype. We investigated the activity of miR-411-5p in the proliferation, migration, and phenotype switch of rat VSMCs.Based on a microRNA array assay, miR-411-5p expression was found to be significantly increased in cultured VSMCs stimulated by platelet-derived growth factor-BB (PDGF-BB). A CCK-8 assay, transwell assay, and scratch test were performed to measure the effect of miR-411-5p on the proliferation and migration of PDGF-BB-treated VSMCs. MiR-411-5p promoted expression of dedifferentiated phenotype markers such as osteopontin and tropomyosin 4 in PDGF-BB-treated VSMCs. Using mimics and inhibitors, we identified the target of miR-411-5p in PDGF-BB-treated VSMCs and found that calmodulin-regulated spectrin-associated protein-1 (CAMSAP1) was involved in the phenotypic switch mediated by PDGF-BB.By inhibiting expression of CAMSAP1, miR-411-5p enhanced the proliferation, migration, and phenotype switch of VSMCs.Blockade of miR-411-5p interaction with CAMSAP1 is a promising approach to treat in-stent restenosis.

CircRNA_001373 promotes liver fibrosis by regulating autophagy activation in hepatic stellate cells via the miR-142a-5p/Becn1 axis.

The purpose of this study was to investigate the regulatory role and underlying mechanisms of circRNA_001373 in the hepatic stellate cell (HSC) activation. Quantitative real-time polymerase chain reaction was used to detect the expression of circRNA_001373, miR-142a-5p and Becn1. The viability of JS-1 cells was measured by Cell Counting Kit-8. The targeting relationship between miR-142a-5p and CircRNA_001373, as well as between miR-142a-5p and Becn1 was predicted using CircInteractome and TargetScan databases, respectively, and validated by dual-luciferase reporter assay. Western blot was utilized to determine the expression levels of proteins related to autophagy and the activation if HSCs in JS-1 cells. After activation by platelet-derived growth factor-BB, an increase was observed in the expression of collagen I and α-smooth muscle actin proteins. The expression of CircRNA_001373 was up-regulated in the activated HSCs. Knockdown of CircRNA_001373 significantly inhibited cell viability and activation of JS-1 cells, as well as autophagy in the activated HSCs. CircRNA_001373 could sponge miR-142a-5p in the activated HSCs, which in turn elevated the Becn1 expression. Concurrent knockdown of both CircRNA_001373 and miR-142a-5p reversed the inhibitory effects of the knockdown of CircRNA_001373 alone on cell viability and autophagy in activated JS-1 cells. CircRNA_ 001373 promotes cell viability and autophagy as well as the activation of JS-1 cells by regulating the miR-142a-5p/Becn1 axis.

Sequential Human Histology Results of the Subperiosteal Minimally Invasive Aesthetic Ridge Augmentation Technique (SMART): A Chronologic Wound Healing Proof-of-Principle Study.

Traditional GBR procedures have been associated with frequent complications and compromised peri-implant esthetics. Tunneling techniques have been proposed as a promising alternative in this regard. More recently, a subperiosteal minimally invasive aesthetic ridge augmentation technique (SMART) was reported to have been clinically successful in a prospective case series. This technique includes the use of a bone graft/recombinant human platelet-derived growth factor-BB combination delivered to the site by a tunneling method. However, published histologic information regarding the nature of the regenerated tissue has been limited. The current study evaluated the histologic and histomorphometric findings of four human specimens harvested at 2, 5, 9, and 14 months after ridge augmentation using the SMART method. Evaluations of the wound healing and bone regeneration sequence over time found that the ridge augmentation was the result of extensive new bone formation that progressed through the woven bone to lamellar bone stages, with remodeling of the xenogeneic graft material and replacement by patient bone. This is the first study utilizing sequential human specimens to histologically examine the chronology of wound healing following alveolar ridge augmentation.

The effect of titanium-platelet rich fibrin on periodontal intrabony defects: A randomized controlled split-mouth clinical study.

This study aimed to determine the contribution of titanium prepared platelet-rich fibrin (T-PRF) with open flap debridement (OFD) on clinical, biochemical and radiographic measurements of periodontal regeneration. Twenty periodontitis patients with bilateral intrabony defects and stage III grade A periodontitis were included in the study. A total of 40 defects were randomly selected for OFD alone (control group, n = 20) or combined OFD+ T-PRF (test group, n = 20). Clinical and radiographic parameters (at baseline and nine months after surgery), and growth factor levels in gingival crevicular fluid (at baseline and at two, four, six, and twelve weeks after surgical treatment) were also evaluated. Considering the clinical parameters, alterations in probing pocket depth, gingival marginal level and clinical endpoint in the test regions treated with T-PRF significantly improved (P<0.05). Fibroblast growth factor-2 and platelet-derived growth factor-BB levels between the two groups in the second and fourth weeks were also significantly different (P<0.05). Furthermore, the receptor activator of nuclear factor κB ligand/osteoprotegerin ratio between the groups was significantly different in the second, fourth, sixth, and twelfth weeks (P<0.05). The bone-filling rate was also significantly greater in the test group than in the control group (P <0.001). Compared with OFD alone, combining T-PRF with the procedure was more successful with regards to clinical, radiographic, and biochemical measurements of periodontal regeneration.