Circuit Design Features of a Stable Two-Cell System.

Cell communication within tissues is mediated by multiple paracrine signals including growth factors, which control cell survival and proliferation. Cells and the growth factors they produce and receive constitute a circuit with specific properties that ensure homeostasis. Here, we used computational and experimental approaches to characterize the features of cell circuits based on growth factor exchange between macrophages and fibroblasts, two cell types found in most mammalian tissues. We found that the macrophage-fibroblast cell circuit is stable and robust to perturbations. Analytical screening of all possible two-cell circuit topologies revealed the circuit features sufficient for stability, including environmental constraint and negative-feedback regulation. Moreover, we found that cell-cell contact is essential for the stability of the macrophage-fibroblast circuit. These findings illustrate principles of cell circuit design and provide a quantitative perspective on cell interactions.

Natural Killer Cells Control Tumor Growth by Sensing a Growth Factor.

Many tumors produce platelet-derived growth factor (PDGF)-DD, which promotes cellular proliferation, epithelial-mesenchymal transition, stromal reaction, and angiogenesis through autocrine and paracrine PDGFRß signaling. By screening a secretome library, we found that the human immunoreceptor NKp44, encoded by NCR2 and expressed on natural killer (NK) cells and innate lymphoid cells, recognizes PDGF-DD. PDGF-DD engagement of NKp44 triggered NK cell secretion of interferon gamma (IFN)-γ and tumor necrosis factor alpha (TNF-α) that induced tumor cell growth arrest. A distinctive transcriptional signature of PDGF-DD-induced cytokines and the downregulation of tumor cell-cycle genes correlated with NCR2 expression and greater survival in glioblastoma. NKp44 expression in mouse NK cells controlled the dissemination of tumors expressing PDGF-DD more effectively than control mice, an effect enhanced by blockade of the inhibitory receptor CD96 or CpG-oligonucleotide treatment. Thus, while cancer cell production of PDGF-DD supports tumor growth and stromal reaction, it concomitantly activates innate immune responses to tumor expansion.

Complement Component 3 Adapts the Cerebrospinal Fluid for Leptomeningeal Metastasis.

We molecularly dissected leptomeningeal metastasis, or spread of cancer to the cerebrospinal fluid (CSF), which is a frequent and fatal condition mediated by unknown mechanisms. We selected lung and breast cancer cell lines for the ability to infiltrate and grow in CSF, a remarkably acellular, mitogen-poor metastasis microenvironment. Complement component 3 (C3) was upregulated in four leptomeningeal metastatic models and proved necessary for cancer growth within the leptomeningeal space. In human disease, cancer cells within the CSF produced C3 in correlation with clinical course. C3 expression in primary tumors was predictive of leptomeningeal relapse. Mechanistically, we found that cancer-cell-derived C3 activates the C3a receptor in the choroid plexus epithelium to disrupt the blood-CSF barrier. This effect allows plasma components, including amphiregulin, and other mitogens to enter the CSF and promote cancer cell growth. Pharmacologic interference with C3 signaling proved therapeutically beneficial in suppressing leptomeningeal metastasis in these preclinical models.

Sclerotome-derived PDGF signaling functions as a niche cue responsible for primitive erythropoiesis.

Primitive erythropoiesis serves a vital role in embryonic development, generating primitive red blood cells responsible for transportation of oxygen throughout the body. Although diverse niche factors are known to function in definitive hematopoiesis, the microenvironment contributing to primitive hematopoiesis remains largely elusive. Here, we report that platelet-derived growth factor (PDGF) signaling is required for erythroid progenitor differentiation in zebrafish. Ablating pdgfαa (also known as pdgfaa) and pdgfαb (also known as pdgfab) or blocking PDGF signaling with an inhibitor impairs erythroid progenitor differentiation, thus resulting in a significant decrease in the number of erythrocytes. We reveal that pdgfαb is expressed in sclerotomal cells, and that its receptor genes, pdgfra and pdgfrb, are expressed in the adjacent erythroid progenitor cells. Sclerotome-specific overexpression of pdgfαb effectively restores primitive erythropoiesis in pdgfαa-/-;pdgfαb-/- mutant embryos. In addition, we have defined ERK1/2 signaling as a downstream pathway of PDGF signaling during embryonic erythropoiesis. Taken together, our findings indicate that PDGF signaling derived from sclerotome functions as a niche cue for primitive erythropoiesis.

MDA-9/Syntenin in the tumor and microenvironment defines prostate cancer bone metastasis.

Bone metastasis is a frequent and incurable consequence of advanced prostate cancer (PC). An interplay between disseminated tumor cells and heterogeneous bone resident cells in the metastatic niche initiates this process. Melanoma differentiation associated gene-9 (mda-9/Syntenin/syndecan binding protein) is a prometastatic gene expressed in multiple organs, including bone marrow-derived mesenchymal stromal cells (BM-MSCs), under both physiological and pathological conditions. We demonstrate that PDGF-AA secreted by tumor cells induces CXCL5 expression in BM-MSCs by suppressing MDA-9-dependent YAP/MST signaling. CXCL5-derived tumor cell proliferation and immune suppression are consequences of the MDA-9/CXCL5 signaling axis, promoting PC disease progression. mda-9 knockout tumor cells express less PDGF-AA and do not develop bone metastases. Our data document a previously undefined role of MDA-9/Syntenin in the tumor and microenvironment in regulating PC bone metastasis. This study provides a framework for translational strategies to ameliorate health complications and morbidity associated with advanced PC.

Functional and clinical roles of stromal PDGF receptors in tumor biology.

PDGF receptors play pivotal roles in both developmental and physiological processes through the regulation of mesenchymal cells involved in paracrine instructive interactions with epithelial or endothelial cells. Tumor biology studies, alongside analyses of patient tissue samples, provide strong indications that the PDGF signaling pathways are also critical in various types of human cancer. This review summarizes experimental findings and correlative studies, which have explored the biological mechanisms and clinical relevance of PDGFRs in mesenchymal cells of the tumor microenvironment. Collectively, these studies support the overall concept that the PDGF system is a critical regulator of tumor growth, metastasis, and drug efficacy, suggesting yet unexploited targeting opportunities. The inter-patient variability in stromal PDGFR expression, as being linked to prognosis and treatment responses, not only indicates the need for stratified approaches in upcoming therapeutic investigations but also implies the potential for the development of PDGFRs as biomarkers of clinical utility, interestingly also in settings outside PDGFR-directed treatments.

Endothelial PDGF-D contributes to neurovascular protection after ischemic stroke by rescuing pericyte functions.

Ischemic stroke induces neovascularization of the injured tissue as an attempt to promote structural repair and neurological recovery. Angiogenesis is regulated by pericytes that potently react to ischemic stroke stressors, ranging from death to dysfunction. Platelet-derived growth factor (PDGF) receptor (PDGFR)ß controls pericyte survival, migration, and interaction with brain endothelial cells. PDGF-D a specific ligand of PDGFRß is expressed in the brain, yet its regulation and role in ischemic stroke pathobiology remains unexplored. Using experimental ischemic stroke mouse model, we found that PDGF-D is transiently induced in brain endothelial cells at the injury site in the subacute phase. To investigate the biological significance of PDGF-D post-ischemic stroke regulation, its subacute expression was either downregulated using siRNA or upregulated using an active recombinant form. Attenuation of PDGF-D subacute induction exacerbates neuronal loss, impairs microvascular density, alters vascular permeability, and increases microvascular stalling. Increasing PDGF-D subacute bioavailability rescues neuronal survival and improves neurological recovery. PDGF-D subacute enhanced bioavailability promotes stable neovascularization of the injured tissue and improves brain perfusion. Notably, PDGF-D enhanced bioavailability improves pericyte association with brain endothelial cells. Cell-based assays using human brain pericyte and brain endothelial cells exposed to ischemia-like conditions were applied to investigate the underlying mechanisms. PDGF-D stimulation attenuates pericyte loss and fibrotic transition, while increasing the secretion of pro-angiogenic and vascular protective factors. Moreover, PDGF-D stimulates pericyte migration required for optimal endothelial coverage and promotes angiogenesis. Our study unravels new insights into PDGF-D contribution to neurovascular protection after ischemic stroke by rescuing the functions of pericytes.

PDGF-D-PDGFRß signaling enhances IL-15-mediated human natural killer cell survival.

The axis of platelet-derived growth factor (PDGF) and PDGF receptor-beta (PDGFRß) plays prominent roles in cell growth and motility. In addition, PDGF-D enhances human natural killer (NK) cell effector functions when binding to the NKp44 receptor. Here, we report an additional but previously unknown role of PDGF-D, whereby it mediates interleukin-15 (IL-15)-induced human NK cell survival but not effector functions via its binding to PDGFRß but independent of its binding to NKp44. Resting NK cells express no PDGFRß and only a low level of PDGF-D, but both are significantly up-regulated by IL-15, via the nuclear factor κB signaling pathway, to promote cell survival in an autocrine manner. Both ectopic and IL-15-induced expression of PDGFRß improves NK cell survival in response to treatment with PDGF-D. Our results suggest that the PDGF-D-PDGFRß signaling pathway is a mechanism by which IL-15 selectively regulates the survival of human NK cells without modulating their effector functions.

A vasculature niche orchestrates stromal cell phenotype through PDGF signaling: Importance in human fibrotic disease.

SignificanceTissue fibrotic diseases, for example of the liver and lung, represent a huge unmet medical need. In this study, using single-cell RNA sequencing, cytometry by time of flight (CyTOF), tissue imaging, and functional assays, we identify a complex vascular niche in Dupuytren's disease (DD), a common localized fibrotic condition of the palm, where early-disease-stage tissue can be accessed readily. We uncover a population of myofibroblast precursors within the pericyte compartment and demonstrate that the endothelium instructs the differentiation of functionally distinct stromal cells, thereby orchestrating discrete microenvironments in the fibrotic milieu. Together, these findings provide a basis for the concept of targeting blood vessel signaling to control the progression of human fibrosis.

A 3D atlas of the human developing pancreas to explore progenitor proliferation and differentiation.

AIMS/HYPOTHESIS: Rodent pancreas development has been described in great detail. On the other hand, there are still gaps in our understanding of the developmental trajectories of pancreatic cells during human ontogenesis. Here, our aim was to map the spatial and chronological dynamics of human pancreatic cell differentiation and proliferation by using 3D imaging of cleared human embryonic and fetal pancreases. METHODS: We combined tissue clearing with light-sheet fluorescence imaging in human embryonic and fetal pancreases during the first trimester of pregnancy. In addition, we validated an explant culture system enabling in vitro proliferation of pancreatic progenitors to determine the mitogenic effect of candidate molecules. RESULTS: We detected the first insulin-positive cells as early as five post-conceptional weeks, two weeks earlier than previously observed. We observed few insulin-positive clusters at five post-conceptional weeks (mean ± SD 9.25±5.65) with a sharp increase to 11 post-conceptional weeks (4307±152.34). We identified a central niche as the location of onset of the earliest insulin cell production and detected extra-pancreatic loci within the adjacent developing gut. Conversely, proliferating pancreatic progenitors were located in the periphery of the epithelium, suggesting the existence of two separated pancreatic niches for differentiation and proliferation. Additionally, we observed that the proliferation ratio of progenitors ranged between 20% and 30%, while for insulin-positive cells it was 1%. We next unveiled a mitogenic effect of the platelet-derived growth factor AA isoform (PDGFAA) in progenitors acting through the pancreatic mesenchyme by increasing threefold the number of proliferating progenitors. CONCLUSIONS/INTERPRETATION: This work presents a first 3D atlas of the human developing pancreas, charting both endocrine and proliferating cells across early development.

Effects of acetazolamide combined with remote ischemic preconditioning on risk of acute mountain sickness: a randomized clinical trial.

BACKGROUND: We aimed to determine whether and how the combination of acetazolamide and remote ischemic preconditioning (RIPC) reduced the incidence and severity of acute mountain sickness (AMS). METHODS: This is a prospective, randomized, open-label, blinded endpoint (PROBE) study involving 250 healthy volunteers. Participants were randomized (1:1:1:1:1) to following five groups: Ripc (RIPC twice daily, 6 days), Rapid-Ripc (RIPC four times daily, 3 days), Acetazolamide (twice daily, 2 days), Combined (Acetazolamide plus Rapid-Ripc), and Control group. After interventions, participants entered a normobaric hypoxic chamber (equivalent to 4000 m) and stayed for 6 h. The primary outcomes included the incidence and severity of AMS, and SpO2 after hypoxic exposure. Secondary outcomes included systolic and diastolic blood pressure, and heart rate after hypoxic exposure. The mechanisms of the combined regime were investigated through exploratory outcomes, including analysis of venous blood gas, complete blood count, human cytokine antibody array, ELISA validation for PDGF-AB, and detection of PDGF gene polymorphisms. RESULTS: The combination of acetazolamide and RIPC exhibited powerful efficacy in preventing AMS, reducing the incidence of AMS from 26.0 to 6.0% (Combined vs Control: RR 0.23, 95% CI 0.07-0.70, P = 0.006), without significantly increasing the incidence of adverse reactions. Combined group also showed the lowest AMS score (0.92 ± 1.10). Mechanistically, acetazolamide induced a mild metabolic acidosis (pH 7.30 ~ 7.31; HCO3- 18.1 ~ 20.8 mmol/L) and improved SpO2 (89 ~ 91%) following hypoxic exposure. Additionally, thirty differentially expressed proteins (DEPs) related to immune-inflammatory process were identified after hypoxia, among which PDGF-AB was involved. Further validation of PDGF-AB in all individuals showed that both acetazolamide and RIPC downregulated PDGF-AB before hypoxic exposure, suggesting a possible protective mechanism. Furthermore, genetic analyses demonstrated that individuals carrying the PDGFA rs2070958 C allele, rs9690350 G allele, or rs1800814 G allele did not display a decrease in PDGF-AB levels after interventions, and were associated with a higher risk of AMS. CONCLUSIONS: The combination of acetazolamide and RIPC exerts a powerful anti-hypoxic effect and represents an innovative and promising strategy for rapid ascent to high altitudes. Acetazolamide improves oxygen saturation. RIPC further aids acetazolamide, which synergistically regulates PDGF-AB, potentially involved in the pathogenesis of AMS. TRIAL REGISTRATION: ClinicalTrials.gov NCT05023941.

Platelet-derived growth factor-stimulated pulmonary artery smooth muscle cells regulate pulmonary artery endothelial cell dysfunction through extracellular vesicle miR-409-5p.

Platelet-derived growth factor (PDGF)-induced changes in vascular smooth muscle cells (VSMCs) stimulate vascular remodeling, resulting in vascular diseases such as pulmonary arterial hypertension. VSMCs communicate with endothelial cells through extracellular vesicles (EVs) carrying cargos, including microRNAs. To understand the molecular mechanisms through which PDGF-stimulated pulmonary artery smooth muscle cells (PASMCs) interact with pulmonary artery endothelial cells (PAECs) under pathological conditions, we investigated the crosstalk between PASMCs and PAECs via extracellular vesicle miR-409-5p under PDGF stimulation. miR-409-5p expression was upregulated in PASMCs upon PDGF signaling, and it was released into EVs. The elevated expression of miR-409-5p was transported to PAECs and led to their impaired function, including reduced NO release, which consequentially resulted in enhanced PASMC proliferation. We propose that the positive regulatory loop of PASMC-extracellular vesicle miR-409-5p-PAEC is a potential mechanism underlying the proliferation of PASMCs under PDGF stimulation. Therefore, miR-409-5p may be a novel therapeutic target for the treatment of vascular diseases, including pulmonary arterial hypertension.

Microvascular smooth muscle cells exhibit divergent phenotypic switching responses to platelet-derived growth factor and insulin-like growth factor 1.

OBJECTIVE: Vascular smooth muscle cell (VSMC) phenotypic switching is critical for normal vessel formation, vascular stability, and healthy brain aging. Phenotypic switching is regulated by mediators including platelet derived growth factor (PDGF)-BB, insulin-like growth factor (IGF-1), as well as transforming growth factor-ß (TGF-ß) and endothelin-1 (ET-1), but much about the role of these factors in microvascular VSMCs remains unclear. METHODS: We used primary rat microvascular VSMCs to explore PDGF-BB- and IGF-1-induced phenotypic switching. RESULTS: PDGF-BB induced an early proliferative response, followed by formation of polarized leader cells and rapid, directionally coordinated migration. In contrast, IGF-1 induced cell hypertrophy, and only a small degree of migration by unpolarized cells. TGF-ß and ET-1 selectively inhibit PDGF-BB-induced VSMC migration primarily by repressing migratory polarization and formation of leader cells. Contractile genes were downregulated by both growth factors, while other genes were differentially regulated by PDGF-BB and IGF-1. CONCLUSIONS: These studies indicate that PDGF-BB and IGF-1 stimulate different types of microvascular VSMC phenotypic switching characterized by different modes of cell migration. Our studies are consistent with a chronic vasoprotective role for IGF-1 in VSMCs in the microvasculature while PDGF is more involved in VSMC proliferation and migration in response to acute activities such as neovascularization. Better understanding of the nuances of the phenotypic switching induced by these growth factors is important for our understanding of a variety of microvascular diseases.

PDGF-BB accelerates TSCC via fibroblast lactates limiting miR-26a-5p and boosting mitophagy.

The tumor microenvironment and cancer-associated fibroblasts (CAFs) play crucial roles in tumor development, and their metabolic coupling remains unclear. Clinical data showed a positive correlation between PDGF-BB, CAFs, and glycolysis in the tumor microenvironment of oral tongue squamous cell carcinoma patients. In vitro, CAFs are derived from hOMF cells treated with PDGF-BB, which induces their formation and promotes aerobic glycolysis. Mitophagy increased the PDGF-BB-induced formation of CAF phenotypes and aerobic glycolysis, while autophagy inhibition blocked PDGF-BB-induced effects. Downregulation of miR-26a-5p was observed in CAFs; upregulation of miR-26a-5p inhibited the expression of mitophagy-related proteins ULKI, Parkin, PINK1, and LC3 and aerobic glycolysis in PDGF-BB-induced CAFs. PDGF-BB-induced CAFs promoted tumor cell proliferation, invasion, metastasis, NF-κB signaling pathway activation, and PDGF-BB secretion. Thus, PDGF-BB is associated with lactate-induced CAF formation and glucose metabolism reprogramming. These findings indicate potential therapeutic targets in oral tongue squamous cell carcinoma.

Investigation of the impact of bromodomain inhibition on cytoskeleton stability and contraction.

BACKGROUND: Injury to contractile organs such as the heart, vasculature, urinary bladder and gut can stimulate a pathological response that results in loss of normal contractility. PDGF and TGFß are among the most well studied initiators of the injury response and have been shown to induce aberrant contraction in mechanically active cells of hollow organs including smooth muscle cells (SMC) and fibroblasts. However, the mechanisms driving contractile alterations downstream of PDGF and TGFß in SMC and fibroblasts are incompletely understood, limiting therapeutic interventions. METHODS: To identify potential molecular targets, we have leveraged the analysis of publicly available data, comparing transcriptomic changes in mechanically active cells stimulated with PDGF and TGFß. Additional Analysis of publicly available data sets were performed on SMC and fibroblasts treated in the presence or absence of the MYC inhibitor JQ1. Validation of in silico findings were performed with qPCR, immunoblots, and collagen gel contraction assays measure the effect of JQ1 on cytoskeleton associated genes, proteins and contractility in mechanically active cells. Likelihood ratio test and FDR adjusted p-values were used to determine significant differentially expressed genes. Student ttest were used to calculate statistical significance of qPCR and contractility analyses. RESULTS: Comparing PDGF and TGFß stimulated SMC and fibroblasts identified a shared molecular profile regulated by MYC and members of the AP-1 transcription factor complex. Additional in silico analysis revealed a unique set of cytoskeleton-associated genes that were sensitive to MYC inhibition with JQ1. In vitro validation demonstrated JQ1 was also able to attenuate TGFß and PDGF induced changes to the cytoskeleton and contraction of smooth muscle cells and fibroblasts in vitro. CONCLUSIONS: These findings identify MYC as a key driver of aberrant cytoskeletal and contractile changes in fibroblasts and SMC, and suggest that JQ1 could be used to restore normal contractile function in hollow organs.

SCARB1-encoded circ _0029343 induces p73 splicing to promote growth and metastasis of hepatocellular carcinoma via miR-486-5p/SRSF3 axis.

Circular RNAs (circRNAs) exhibit essential regulation in the malignant development of hepatocellular carcinoma (HCC). This study aims to investigate the physiological mechanisms of circ_0029343 encoded by scavenger receptor class B member 1 (SCARB1) involved in the growth and metastasis of HCC. Differentially expressed mRNAs in HCC were obtained, followed by the prediction of target genes of differentially expressed miRNAs and gene ontology and kyoto encyclopedia of genes and genomes analysis on the differentially expressed mRNAs. Moreover, the regulatory relationship between circRNAs encoded by SCARB1 and differentially expressed miRNAs was predicted. In vitro cell experiments were performed to verify the effects of circ_0029343, miR-486-5p, and SRSF3 on the malignant features of HCC cells using the gain- or loss-of-function experiments. Finally, the effects of circ_0029343 on the growth and metastasis of HCC cells in xenograft mouse models were also explored. It was found that miR-486-5p might interact with seven circRNAs encoded by SCARB1, and its possible downstream target gene was SRSF3. Moreover, SRSF3 was associated with the splicing of various RNA. circ_0029343 could sponge miR-486-5p to up-regulate SRSF3 and activate PDGF-PDGFRB (platelet-derived growth factor and its receptor, receptor beta) signaling pathway by inducing p73 splicing, thus promoting the proliferation, migration, and invasion and inhibiting apoptosis of HCC cells. In vivo, animal experiments further confirmed that overexpression of circ_0029343 could promote the growth and metastasis of HCC cells in nude mice. circ_0029343 encoded by SCARB1 may induce p73 splicing and activate the PDGF-PDGFRB signaling pathway through the miR-486-5p/SRSF3 axis, thus promoting the growth and metastasis of HCC cells.

Curcumol reduces lower limb arteriosclerosis in rats by inhibiting human arterial smooth muscle cell activity.

Cardiovascular diseases, particularly those involving arterial stenosis and smooth muscle cell proliferation, pose significant health risks. This study aimed to investigate the therapeutic potential of curcumol in inhibiting platelet-derived growth factor-BB (PDGF-BB)-induced human aortic smooth muscle cell (HASMC) proliferation, migration and autophagy. Using cell viability assays, 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays and Western Blot analyses, we observed that curcumol effectively attenuated PDGF-BB-induced HASMC proliferation and migration in a concentration-dependent manner. Furthermore, curcumol mitigated PDGF-BB-induced autophagy, as evidenced by the downregulation of LC3-II/LC3-I ratio and upregulation of P62. In vivo experiments using an arteriosclerosis obliterans model demonstrated that curcumol treatment significantly ameliorated arterial morphology and reduced stenosis. Additionally, curcumol inhibited the activity of the KLF5/COX2 axis, a key pathway in vascular diseases. These findings suggest that curcumol has the potential to serve as a multi-target therapeutic agent for vascular diseases.

Cathepsin K inhibition alleviates periodontal bone resorption by promoting type H vessel formation through PDGF-BB/PDGFR-ß axis.

OBJECTIVES: To explore the effects of cathepsin K (CTSK) inhibition on type H vessel formation and alveolar bone resorption within periodontitis. METHODS: Conditioned media derived from preosteoclasts pretreated with the CTSK inhibitor odanacatib (ODN), ODN supplemented small interfering RNA targeting PDGF-BB (si-PDGF-BB), or PBS were prepared, to assess their proangiogenic effects on endothelial cells (HUVECs). A series of angiogenic-related assays were conducted to evaluate HUVEC proliferation, migration, and tube formation abilities in vitro. In addition, qRT-PCR and Western blot assays were employed to examine the expression levels of genes/proteins related to PDGF-BB/PDGFR-ß axis components. A mouse periodontitis model was established to evaluate the effects of CTSK inhibition on type H vessel formation. RESULTS: CTSK inhibition promoted PDGF-BB secretion from preosteoclasts and proliferation, migration, and tube formation activities of HUVECs in vitro. However, the conditioned medium from preosteoclasts pretreated by si-PDGF-BB impaired the angiogenic activities of HUVECs. This promoted angiogenesis function by CTSK inhibition may be mediated by the PDGF-BB/PDGFR-ß axis. Functionally, in vivo studies demonstrated that CTSK inhibition significantly accelerated type H vessel formation and alleviated bone loss within periodontitis. CONCLUSION: CTSK inhibition promotes type H vessel formation and attenuates alveolar bone resorption within periodontitis via PDGF-BB/PDGFR-ß axis.

The Effect of a Rotating Magnetic Field on the Regenerative Potential of Platelets.

Platelets are actively involved in tissue injury site regeneration by producing a wide spectrum of platelet-derived growth factors such as PDGF (platelet-derived growth factor), IGF-1 (insulin-like growth factor), TGF-ß1 (transforming growth factor ß), FGF (fibroblast growth factor), etc. A rotating magnetic field (RMF) can regulate biological functions, including reduction or induction regarding inflammatory processes, cell differentiation, and gene expression, to determine the effect of an RMF on the regenerative potential of platelets. The study group consisted of 30 healthy female and male volunteers (n = 15), from which plasma was collected. A portion of the plasma was extracted and treated as an internal control group. Subsequent doses of plasma were exposed to RMF at different frequencies (25 and 50 Hz) for 1 and 3 h. Then, the concentrations of growth factors (IGF-1, PDGF-BB, TGF-ß1, and FGF-1) were determined in the obtained material by the ELISA method. There were statistically significant differences in the PDGF-BB, TGF-ß1, IGF-1, and FGF-1 concentrations between the analyzed groups. The highest concentration of PDGF-BB was observed in the samples placed in RMF for 1 h at 25 Hz. For TGF-ß1, the highest concentrations were obtained in the samples exposed to RMF for 3 h at 25 Hz and 1 h at 50 Hz. The highest concentrations of IGF-1 and FGF-1 were shown in plasma placed in RMF for 3 h at 25 Hz. An RMF may increase the regenerative potential of platelets. It was noted that female platelets may respond more strongly to RMF than male platelets.

[Sinomenine inhibits PDGF/PDGFR signaling pathway to reduce RA-FLS migration induced by NETs].

This study aims to decipher the mechanism of sinomenine in inhibiting platelet-derived growth factor/platelet-derived growth factor receptor(PDGF/PDGFR) signaling pathway in rheumatoid arthritis-fibroblast-like synoviocyte(RA-FLS) migration induced by neutrophil extracellular traps(NETs). RA-FLS was isolated from the synovial tissue of 3 RA patients and cultured. NETs were extracted from the peripheral venous blood of 4 RA patients and 4 healthy control(HC). RA-FLS was classified into control group, HC-NETs group, RA-NETs group, RA-NETs+sinomenine group and RA-NETs+sinomenine+CP-673451 group. RNA-sequencing(RNA-seq) was conducted to identify the differentially expressed genes between HC-NETs and RA-NETs groups. Sangerbox was used to perform the Gene Ontology(GO) function and the Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment. Cytoscape was employed to build the protein-protein interaction(PPI) network. AutoDock Vina and PyMOL were used for molecular docking of sinomenine with PDGFß and PDGFRß. The cell proliferation and migration were determined by the cell counting kit-8(CCK-8) and cell scratch assay, respectively. Western blot was employed to determine the protein level of PDGFRß. Real-time quantitative polymerase chain reaction(RT-qPCR) was carried out to determine the mRNA levels of matrix metalloproteinases(MMPs). The results revealed that neutrophils in RA patients were more likely to produce NETs. Compared with HC-NETs group, RA-NETs group showed up-regulated expression of PDGFß and PDGFRß. Compared with control group, RA-NETs group showed increased cell proliferation and migration and up-regulated protein level of PDGFRß and mRNA levels of PDGFß, PDGFRß, MMP1, MMP3, and MMP9(P<0.05). Compared with RA-NETs group, RA-NETs+sinomenine group presented decreased cell proliferation and migration and down-regulated protein and mRNA level of PDGFRß and mRNA levels of MMP1, MMP3, and MMP9(P<0.05). Compared with RA-NETs+sinomenine group, the proliferation ability of RA-NETs+sinomenine+CP-673451 group decreased(P<0.05). The findings prove that sinomenine reduces the RA-NETs-induced RA-FLS migration by inhibiting PDGF/PDGFR signaling pathway, thus mitigating RA.