Levels Of PDGF-BB Research Articles

Regular aerobic exercise activates PDGF-BB/PDGFR-β signaling and modulates the inflammatory-anti-inflammatory balance in diet-induced obese mice

ObjectiveRegular aerobic exercise induces cardioprotection by counteracting the obesity-associated inflammatory response, dyslipidemia. PDGF-BB/PDGFR-β signaling is established as a crucial mechanism of endothelial cell-cardiomyocyte communication and cardioprotection, but its physiological roles in response to obesity and regular aerobic exercise are unknown. MethodsThirty C57BL/6 mice were divided into three groups: a normal diet group, a high-fat diet group, and a high-fat diet plus aerobic exercise group. Glucose metabolic parameters, inflammation-related indicators, and blood lipids indicators were detected. In addition, gene expression levels of the inflammatory factors, PDGF-BB, PDGFR-β, PI3K, Akt, eNOS, and P53 in cardiac tissue were quantified. Morphological analysis was also used to quantify the magnitude of inflammation. ResultsHigh-fat diet (HFD) feeding resulted in adiposity, dyslipidemia, and low levels of cardioprotective factors such as APN and eNOS (P < 0.05), which were improved significantly by 8 weeks of aerobic exercise (P < 0.05). HFD feeding increased the gene expression levels of proinflammatory cytokines and decreased the gene expression levels of anti-inflammatory cytokines in cardiac tissue (P < 0.05), which was reversed by regular aerobic exercise (P < 0.05). In addition, HFD feeding suppressed the levels of the cardioprotective factors PDGF-BB and eNOS through PDGF-BB/PDGFR-β/PI3K/Akt/eNOS signaling in cardiac tissue, while regular aerobic exercise activated PDGF-BB/PDGFR-β/PI3K/Akt/eNOS signaling. ConclusionRegular aerobic exercise improved adiposity, dyslipidemia induced by HFD feeding. Regular aerobic exercise exerted a prominent role in modulating the inflammatory-anti-inflammatory balance and activating the levels of the cardioprotective factors eNOS and PDGF-BB through PDGF-BB/PDGFR-β signaling.

Pneumonitis in patients with thymoma and Good's syndrome.

e20595 Background: The association between thymoma (T) and immunological dysregulations is well acknowledged. Good’s syndrome (GS), which occurs in approximately 6% to 11% of T patients, is characterized by hypogammaglobulinemia, few or absent B-cells (CD19+), CD4 T cells lymphopenia, abnormal CD4/CD8 T ratio and impaired T cell mitogenic response. Patients with GS have increased susceptibility to bacterial, opportunistic and viral infections related to both humoral and cell-mediated immunodeficiency. The recent Sars-Cov-2 pandemic drew attention to the clinical condition of fatal viral pneumonitis and cytokines storm. Here we reported our monocentric experience of pneumonitis in patients with T and GS before the Sars-Cov-2 pandemic. Methods: We conducted a retrospective analysis of T patients with associated GS referred to the Rare Tumours Reference Center of University Federico II of Naples over a 10-year period (from 2009 to 2019). All the patients with radiological and/or clinical pneumonitis diagnosis were evaluated for this report. Immunological features, histopathological diagnosis and clinical outcome were registered. Results: A total of 41 patients with T and GS were identified, including 17 patients with local disease (stage I-II according to Masaoka-Koga) and 24 with advanced disease (stage III-IV). The majority (56.3%) had B2 T diagnosis. Radiological and/or clinical pneumonitis diagnosis was assessed in 23 cases (56%). Viral pneumonitis was detected in 8 patients: 3 patients with H1N1 infection, 3 patients with CMV infection, 2 patients with EBV infection. Bacterial pneumonitis was diagnosed in 9 patients (3 patients with K. pneumoniae, 4 patients with S. aureus, 2 patients with H. influenza). Opportunistic pneumonitis was found in 2 patients with Aspergillosis infection. In 2 cases no pathogenic agent was identified. The immunophenotyping, assessed in 4 patients with viral pneumonitis, displayed very low/undetectable levels of B cells, with median % value of CD3+T cells and NKT of respectively 9.8% and 0.4% of the total leukocytes. The median % of Treg was 4.7%. CD4+/CD8+ ratio was variable, ranging from 1,2 to 0,6. Interestingly the number of B cells was extremely low, independent of CD4+/CD8+ ratio. Blood levels of cytokines, chemokines and growth factors revealed elevated IL-4, Eotaxin, CCL2 / MCP-1 and CCL5 / RANTES ad strong reduction of IL-10. PDGF-BB levels were also elevated. 15 patients required admission to intensive care Six patients died for fatal pneumonitis. Conclusions: The management of T patients with GS is extremely challenging. Clear diagnostic algorithms do not yet exist and immune-profiling and quantitative immunoglobulins should be considered a part of diagnostic search in these patients. The coexistence of cancer, infections and immunosuppression may trigger life-threatening conditions, such as fatal pneumonitis, which often require intensive care and multidisciplinary approach.

Light rare earth elements hinder bone development via inhibiting type H vessels formation in mice

Light rare earth elements (LREEs) are widely used in medical, industrial, and agricultural fields. Wide application of light rare earth and exposure to these elements in human society leads to increasing accumulation of LREE in human skeletal system. However, the effects of LREEs on human bone health is not clear. In this study, we found that LREE reduced CD31highEmcnhigh endothelial cell mediated type H vessels formation at the metaphyseal sites, resulting in reduced bone mass and low bone quality in mouse bone development. To explore the underlying mechanism, we induced bone marrow macrophages (BMMs) to preosteoclasts (pOCs) with exposure of LREE (Pr3+, Nd3+, Sm3+). The cytotoxicity of LREE was evaluated by CCK-8. Platelet-derived growth factor (PDGF-BB) is the cytokine secreted by pOCs that most responsible for inducing Type H vessel formation. We used ELISA kit to determine the PDGF-BB level in pOC supernatant, and mouse serum finding that the PDGF-BB level was reduced by LREEs treatment. Then we tested the ability of migration and tube formation of HUVECs using condition medium from pOCs. The migration and tube formation ability of HUVECs were both suppressed with LREEs pretreatment. We concluded that LREEs hinder mouse bone development by suppressing type H vessels associated bone formation. Data and materials availabilityAll data generated or analyzed during this study are included in this article. Please contact the corresponding author for unique material requests. Some material used in the reported research may require requests to collaborators and agreements with both commercial and non-profit institutions, as specified in the paper. Requests are reviewed by Third Military Medical University to verify whether the request is subject to any intellectual property or confidentiality obligations. Any material that can be shared will be released via a Material Transfer Agreement.

The role of the primary cilium in pulmonary arterial hypertension

Introduction Vascular remodeling in pulmonary arterial hypertension (PAH) is characterized by endothelial dysfunction and smooth muscle cell proliferation and hypertrophy ultimately causing right heart failure and death. Up to now, the molecular mechanisms of PAH pathogenesis have not been adequately resolved, preventing the development of novel therapies specifically targeting vascular remodeling in PAH. Based on increased expression levels of PDGF-BB and TGF-β in both patients and animal models with PAH, these signaling pathways have been proposed to play a critical role as drivers of lung vascular remodeling. Among multiple other effects, PDGF-BB and TGF-β cause shortening of the primary cilium – an antenna-like organelle functioning as a flow-sensor and signaling hub on most eukaryotic cells. Loss or shortening of cilia is characteristically associated with cell proliferation and promotes PDGF signaling, thus potentially establishing a positive feedback loop. We thus hypothesized that PAH is associated with a loss or shortening of primary cilia in pulmonary artery endothelial (PAECs) and smooth muscle cells (PASMCs) which in turn drives cell proliferation and thus, remodeling in the pulmonary arterial wall. Methods Pulmonary artery tissue from PH-patients and non-PH donors was fixed and stained for acetylated α-tubulin to determine the number of primary cilia. PAECs and PASMCs from PAH patients and non-PAH donors were exposed in vitro to three different characteristic stimuli or mediators, respectively, of PAH, namely PDGF-BB, TGF-β1, or hypoxia (1%). Cells were fixed and stained for measurement of primary cilia length. In parallel, donor and PAH-SMCs were compared functionally in migration and proliferation assays. IFT88 - an essential structural protein of the primary cilium - was knocked down by siRNA in PAECs and PASMCs to assess the effect of cilium loss on migration and proliferation. Results In pulmonary arteries of PAH patients, the number of primary cilia per area was reduced by 80% as compared to donor lungs. PAH-SMCs were found to have shorter cilia and similar to IFT88 knock-down ECs and SMCs showed increased migration and proliferation as compared to healthy control cells. Additionally, primary cilia of PAECs, PASMCs, and PAH-SMCs stimulated with PDGF-BB, TGF-β1, or hypoxia were shorter as compared to controls. Lithium, which has been shown to elongate cilia in other cell types, was able to rescue cilia length and reduced migration and proliferation in PAH-SMCs. Conclusions Here, we demonstrate that PAH is associated with a loss of cilia in vivo and reduced cilium length in vitro. Primary cilium loss or shortening may promote progression of vascular remodeling as indicated by the fact that cilium loss causes proliferation/migration in PAH-SMCs, and their parallel reversal by lithium. Cilium loss may as such be an important propagator of vascular remodeling in PAH and may present a target for novel therapeutic interventions such as lithium.

Decreased Platelet Counts and Serum Levels of VEGF-A, PDGF-BB, and BDNF in Extremely Preterm Infants Developing Severe ROP

Introduction: Thrombocytopenia has been identified as an independent risk factor for retinopathy of prematurity (ROP), although underlying mechanisms are unknown. In this study, the association of platelet count and serum platelet-derived factors with ROP was investigated. Methods: Data for 78 infants born at gestational age (GA) <28 weeks were included. Infants were classified as having no/mild ROP or severe ROP. Serum levels of vascular endothelial growth factor A, platelet-derived growth factor BB, and brain-derived neurotrophic factor were measured in serum samples collected from birth until postmenstrual age (PMA) 40 weeks. Platelet counts were obtained from samples taken for clinical indication. Results: Postnatal platelet counts and serum concentrations of the 3 growth factors followed the same postnatal pattern, with lower levels in infants developing severe ROP at PMA 32 and 36 weeks (p < 0.05–0.001). With adjustment for GA, low platelet counts and low serum concentrations of all 3 factors at PMA 32 weeks were significantly associated with severe ROP. Serum concentrations of all 3 factors also strongly correlated with platelet count (p < 0.001). Conclusion: In this article, we show that ROP, platelet counts, and specific pro-angiogenic factors correlate. These data suggest that platelet-released factors might be involved in the regulation of retinal and systemic angiogenesis after extremely preterm birth. Further investigations are needed.

Angiokines Associated with Targeted Therapy Outcomes in Patients with Non-Clear Cell Renal Cell Carcinoma.

Biomarkers are needed in patients with non-clear cell renal cell carcinomas (NC-RCC) to inform treatment selection but also to identify novel therapeutic targets. We thus sought to profile circulating angiokines in the context of a randomized treatment trial of everolimus versus sunitinib. ASPEN (NCT01108445) was an international, randomized, open-label phase II trial of patients with metastatic papillary, chromophobe, or unclassified NC-RCC with no prior systemic therapy. Patients were randomized to everolimus or sunitinib and treated until disease progression or unacceptable toxicity. The primary endpoint was radiographic progression-free survival (PFS) defined by RECIST 1.1. Plasma angiokines were collected at baseline, cycle 3, and progression and associated with PFS and overall survival (OS). We enrolled 108 patients, 51 received sunitinib and 57 everolimus; of these, 99 patients had evaluable plasma for 23 angiokines. At the final data cutoff, 94 PFS and 64 mortality events had occurred. Angiokines that were independently adversely prognostic for OS were osteopontin (OPN), TIMP-1, thrombospondin-2 (TSP-2), hepatocyte growth factor (HGF), and VCAM-1, and these were also associated with poor-risk disease. Stromal derived factor 1 (SDF-1) was associated with improved survival. OPN was also significantly associated with worse PFS. No statistically significant angiokine-treatment outcome interactions were observed for sunitinib or everolimus. Angiopoeitin-2 (Ang-2), CD-73, HER-3, HGF, IL6, OPN, PIGF, PDGF-AA, PDGF-BB, SDF-1, TGF-b1-b2, TGFb-R3, TIMP-1, TSP-2, VCAM-1, VEGF, and VEGF-R1 levels increased with progression on everolimus, while CD-73, ICAM-1, IL6, OPN, PlGF, SDF-1, TGF-b2, TGFb-R3, TIMP-1, TSP-2, VEGF, VEGF-D, and VCAM-1 increased with progression on sunitinib. In patients with metastatic NC-RCC, we identified several poor prognosis angiokines and immunomodulatory chemokines during treatment with sunitinib or everolimus, particularly OPN.

Serum Cytokine Profiles of Melanoma Patients and Their Association with Tumor Progression and Metastasis.

Purpose Previous studies have shown that melanoma cells produce excessive levels of cytokines, which have various biological roles during melanoma development. The aim of this study was to expand the profile of serum cytokines, chemokines, growth factors, and angiogenic factors that are associated with melanoma, to find more cytokines with abnormal concentrations in melanoma patients, to identify whether the level of cytokines correlated with prognostic variants, such as Breslow thickness and BRAF mutation, and, finally, to find out the cytokines that play important roles during melanoma development. Materials and Methods Multiplex immunobead assay technology and 45-plex immunoassays ProcartaPlex™ kits were used to simultaneously compare the levels of cytokines, growth factors, angiogenic factors, and chemokines between the serum of healthy patients (n = 30) and those with melanoma (n = 72). Data were analyzed according to the clinical characteristics of the designated patient subgroups. Results Compared to the control group, melanoma patients had higher levels of VEGF-A, PDGF-BB, IL-1RA, PIGF-1, IFN-γ, TNF-α, MIP-1α, and SCF, but lower levels of BDNF, SDF-1α, MCP-1, Eotaxin, EGF, and IL-7. Furthermore, the levels of TNF-α (P=0.320, r = 0.019), IFN-γ (P=0.311, r = 0.023), VEGF-A (P=0.014, r = 0.337), and BDNF (0.004, r = −0.391) showed a significant correlation with Breslow thickness. IL-7 was of lower levels in patients harboring BRAF mutants. Melanoma patients with high levels of MIP-1α and MCP-1 showed the poorest overall survival. Conclusions We found that the levels of VEGF-A and PDGF-BB in the serum of both primary and metastatic melanoma patients are elevated. TNF-α, IFN-γ, and VEGF-A presented a positive correlation with Breslow thickness, whereas BDNF showed a negative association. MIP-1α and MCP-1 correlated negatively with survival. In addition, lower levels of IL-7 were found in patients harboring BRAF mutants. These findings indicate that these cytokines may play critical roles in the progression of melanoma.

Measurement of 45 cytokine, chemokine and growth factors in established cell culture supernatants and autologous serum from advanced melanoma patients.

Melanoma is one of the most aggressive forms of human cancer and its incidence has significantly increased worldwide over the last decades. This neoplasia has been characterized by the release of a wide variety of soluble factors, which could stimulate tumor cell proliferation and survival in an autocrine and paracrine manner. Consequently, we sought to evaluate the pattern of soluble factors produced by pre-metastatic and metastatic melanoma established cultures, and to determine whether these factors can be detected in the autologous serum of malignant melanoma patients. Our results showed that both melanoma cultures had a common profile of 27 soluble factors mainly characterized by the high expression of VEGF-A, IL-6, MCP-1, IL-8, and SDF-1. In addition, when we compared supernatants, we observed significant differences in VEGF-A, BDNF, FGF-2, and NGF-β concentrations. As we found in melanoma cultures, serum samples also had their specific production pattern composed by 21 soluble factors. Surprisingly, PDGF-BB and EGF were only found in serum, whereas IL-2, IL-4, IL-8, IL31, FGF2, and GRO-α were only expressed in the supernatant. Significant differences in PDGF-BB, MIP-1β, HGF, PIGF-1, BDNF, EGF, Eotaxin, and IP-10 were also found after comparing autologous serum with healthy controls. According to this, no correlation was found between culture supernatants and autologous serum samples, which suggests that some factors may act locally, and others systemically. Nonetheless, after validation of our results in an independent cohort of patients, we concluded that PDGF-BB, VEGF-A, and IP-10 serum levels could be used to monitor different melanoma stages.

Evaluation of Platelet Rich Fibrin Obtained Using Different Centrifugation Parameters as a Tool for Regenerative Medicine.

Platelet-rich fibrin (PRF) is a biomaterial widely used in the field of regenerative medicine. The purpose of this work was to analyze the structure and biomolecular characteristics of PRF through nine centrifugation parameters (CP) for its preparation, using a pool of blood samples of five volunteers. The PRF obtained was analyzed by morphological and histological characteristics, as well as electronic and atomic force microscopy and growth factors determinations. A longer time of centrifugation showed taller clots and denser mesh fibrin in comparison with a short time (p < 0.05). The protocols with higher speed of centrifugation showed higher levels of PDGF-BB and VEGF. Higher levels of TGFβ1 were found in protocols with a shorter centrifuge time. The mean platelet count (916.05 ± 23.73 cells x 103 cells x cm3) and its roughness (Ra) (616.5 ± 45.2 nm) did not show significant differences between different CP (p > 0.05). A significant correlation between fibrin density and levels of PDGF (r = 0.57) and VEGF (r = 0.52) was found. Additionally, the size of the clot had a significant correlation (r = -0.47) with TGFβ1 levels. Different centrifugation parameters to obtain PRF have been reported. These results indicate that changes in the conditions to obtain PRF have a significant impact on their fibrin structure, cellular distribution, and biomolecular content, which can be decisive for its choice in the different clinical applications to be used. It is necessary to use a standardized centrifuge and protocol to guarantee high-quality PRF and clinical outcomes with less variability.

IL-1RA in the supernatant of QuantiFERON-TB Gold In-Tube and QuantiFERON-TB Gold Plus is useful for discriminating active tuberculosis from latent infection

IntroductionThe new-generation QuantiFERON (QFT)-TB Gold Plus (QFT-Plus) is expected to be useful because it includes a new peptide that is supposed to induce a CD8+ T cell response. There is a need for a new marker with sensitivity higher than that of the conventional IFN-γ release assays owing to false-negative results in the latter. This study aimed to compare cytokines in QFT-plus and QuantiFERON-Gold In-Tube (QFT-GIT) supernatants to discriminate between active tuberculosis and latent tuberculosis infection (LTBI). MethodsA cross-sectional study was conducted at Tokyo National Hospital, wherein 21 LTBI patients and age and sex-matched active TB patients were randomly selected. The levels of various cytokines were measured and compared using the MAGPIX System, and receiver operating characteristic (ROC) curves were generated. ResultsIL-1RA, IFN-γ, CXCL10/IP-10, and CCL4/MIP-1β levels were higher in the active TB group than in the LTBI group in QFT-GIT antigen (GIT Ag) tubes. In QFT-plus tubes, IL-1RA was higher in TB1 and TB2 tubes, while CCL2/MCP-1 was higher only in TB2 tubes. In Nil tubes, CCL5/RANTES, TNF-α, PDGF-BB, and IL-2 levels were significantly higher in the active TB group. IL-1RA in GIT Ag tubes showed the highest area under the curve of 0.8367. The sensitivity and specificity of IL-1RA were 66.7% (95% confidence interval [CI]: 43.0–85.4) and 90.5% (95% CI: 69.6–98.8), respectively, which were the highest among the cytokines. ConclusionsIL-1RA level in the QFT-GIT supernatant can be a good marker for discriminating active TB from LTBI.

Effect of Cryo-Processing on Platelet-Rich Autoplasma Preparations.

Platelet-derived growth factor (PDGF) plays an important role in angiogenesis, affects activation of migration and proliferation of mesenchymal stem cells, fibroblasts, smooth muscle cells, osteoblasts; activation of migration of monocytes, macrophages, and neutrophils.The aim of the investigation was to study the effect of cryo-processing on the qualitative properties of platelet-rich autoplasma (PRP) at different time intervals.Materials and Methods.Autologous plasma preparations were obtained from the blood of 31 donors. The biological material was prepared by double centrifugation according to the protocol for obtaining P-PRP and L-PRP. Platelet count and the concentration of growth factors (PDGF-AA and PDGF-BB) were studied in fresh PRP preparations. In frozen PRP samples, the concentration of PDGF-AA and PDGF-BB was determined 2 weeks after cryo-processing and 2 months after cryo-processing at –35 °С. P-PRP and L-PRP samples activated with 10% CaCl2 solution and those non-activated were studied.Results.L-PRP preparations are significantly superior to P-PRP preparations: the concentration of platelets is 1.7 times higher in them. The level of PDGF-AA in non-activated L-PRP is 1.8 times higher than in non-activated P-PRP (p<0.05). The level of PDGF-AA is 1.5 times higher in activated L-PRP than in activated P-PRP (p<0.05). The level of PDGF-BB is 2.9 times higher in non-activated L-PRP than in non-activated P-PRP and 1.8 times higher in activated L-PRP than in activated P-PRP (p<0.05). The concentration of PDGF-BB in non-activated P-PRP sharply increases in the 2nd week after freezing and remains at the same level after 2 months (p<0.05). The concentration of PDGF-BB in activated plasma does not change (p>0.05).Conclusion.Cryo-processing of non-activated autologous L-PRP allows preserving and subsequently enhancing the properties of plasma concentrate, which makes it possible to apply it in clinical practice.

Total Flavonoids of Rhizoma Drynariae Enhances Angiogenic-Osteogenic Coupling During Distraction Osteogenesis by Promoting Type H Vessel Formation Through PDGF-BB/PDGFR-β Instead of HIF-1α/ VEGF Axis.

Background: Total flavonoids of Rhizoma Drynariae (TFRD), extracted from the kidney-tonifying traditional Chinese medicine Rhizoma Rrynariae, has been proved to be effective in treating osteoporosis, bone fractures and defects. However, pharmacological effects of TFRD on type H vessels, angiogenic-osteogenic coupling in distraction osteogenesis (DO) and the mechanism remain unclear. This study aims at investigating whether type H vessels exist in the DO model, effects of TFRD on angiogenic-osteogenic coupling and further elucidating the underlying mechanism. Methods: Rats models of DO and bone fracture (FR) were established, and then were separately divided into TFRD and control subgroups. Imageological and histological analyses were performed to assess bone and vessel formation. Immunofluorescent staining of CD31 and endomucin (Emcn) was conducted to determine type H vessel formation. Matrigel tube formation, ALP and Alizarin Red S staining assays were performed to test the effects of TFRD on angiogenesis or osteogenesis of endothelial precursor cells (EPCs) or bone marrow-derived mesenchymal stem cells (BMSCs). Additionally, expression levels of HIF-1α, VEGF, PDGF-BB, RUNX2 and OSX were determined by ELISA, qPCR or western blot, respectively. Results: The in vivo results indicated more formed type H vessels in DO groups than in FR groups and TFRD obviously increased the abundance of type H vessels. Moreover, groups with higher abundance of type H vessels showed better angiogenesis and osteogenesis outcomes. Further in vitro experiments showed that TFRD significantly promoted while blocking PDGF-BB remarkably suppressed the angiogenic activity of EPCs under stress conditions. The levels of p-AKT and p-ERK1/2, downstream mediators of the PDGF-BB pathway, were up-regulated by TFRD but blocked by function blocking anti-PDGF-BB antibody. In contrast, the activated AKT and ERK1/2 and corresponding tube formation were not affected by the HIF-1α inhibitor. Besides, blocking PDGF-BB inhibited the osteogenic differentiation of the stretched BMSCs, but TFRD enhanced the osteogenic activity of BMSCs and ameliorated the inhibition, with more calcium nodes, higher ALP activity and mRNA and protein levels of RUNX2 and OSX. Conclusion: Type H vessels exist in the DO model and TFRD enhances angiogenic-osteogenic coupling during DO by promoting type H vessel formation via PDGF-BB/PDGFR-β instead of HIF-1α/VEGF axis.

RETRACTED: Neointimal hyperplasia after carotid transection and anastomosis surgery is associated with degradation of decorin and platelet-derived growth factor signaling

ObjectiveIntimal hyperplasia (IH) is the expansion of the vascular intimal region after intervention, which can lead to stenosis and eventual failure of vascular grafts or interventional procedures such as angioplasty or stent placement. Our goals were to investigate the development of IH in a rabbit open surgical model and to evaluate the associated pathophysiologic processes involving decorin and the platelet-derived growth factor-BB/platelet-derived growth factor receptor-β/mitogen-activated protein kinase (PDGF-BB/PDGFR-β/MAPK) pathway. MethodsWe conducted carotid transection and primary anastomosis on five New Zealand white rabbits to induce IH and examined the associated pathophysiologic changes. Tissue was obtained for histological and protein analysis on postoperative day 21 using the contralateral vessel as a control. Intimal medial thickness (IMT) was calculated to measure IH and compared with the unoperated side. Western blot analysis was performed on tissue lysates to determine the expression of decorin core protein, PDGF-BB, PDGFR-β, and phosphorylated-MAPK (ph-MAPK). Immunofluorescence microscopy was used to assess tissue distribution of matrix metalloproteinase-2 (MMP-2) and ph-PDGFR-β. ResultsBilateral carotid arteries were harvested on postoperative day 21. We compared the IMT in operated with unoperated specimens. IMT was significantly elevated in operated arteries vs unoperated arteries in all five animals (148.6 μm ± 9.09 vs 103.40 μm ± 7.08; 135.2 μm ± 8.30 vs 92.40 μm ± 2.35; 203.1 μm ± 30.23 vs 104.00 μm ± 4.52; 236.2 μm ± 27.22 vs 141.50 μm ± 9.95; 226.9 μm ± 11.12 vs 98.8 μm ± 3.78). Western blot analysis revealed degradation of decorin protein in the operated tissue, including loss of a 50 kDa band and the appearance of a cleaved fragment at 10 kDa. Decorin and MMP-2 were observed, via immunofluorescence microscopy, in the neointima of the operated vessels. Western blot analysis also revealed increased PDGF-BB, PDGFR-β, and ph-MAPK levels in operated tissue. Immunofluorescent staining for ph-PDGFR-β primarily localized to the neointima, indicating increased signaling through PDGF in this region. ConclusionsCarotid transection and primary reanastomosis in rabbits induced IH that was associated with MMP-2 activation, degradation of decorin, and activation of the PDGF/PDGFR-β/MAPK pathway. The findings in this study should lead to further mechanistic evaluation of these pathways to better understand the potential to modify the intimal hyperplastic response to surgery. (JVS–Vascular Science 2021;2:2-12.) Clinical RelevanceIntimal hyperplasia remains a significant challenge to the vascular surgeon in open and interventional procedures. Basic science studies have made headway into understanding the process, but this has not translated into many therapeutic options particularly for primary prevention after a procedure. Decorin is gaining popularity as an important mediator of various pathophysiologic processes involving the extracellular matrix. We sought to determine the possible role of decorin in the development of neointimal hyperplasia in an open surgical model. This study provides a replicable model for the development of intimal hyperplasia and potential therapeutic targets going forward.

Effect of rat bone marrow stromal stem cells overexpressing platelet-derived growth factor and bone morphogenic protein into platelet-rich plasma gels on osteogenic differentiation

The aim of this study was to improve osteoblast function by incorporating rat bone marrow stromal stem cells (rBMSCs) overexpressing platelet-derived growth factor (PDGF-BB) and bone morphogenic protein (BMP-2) into platelet-rich plasma (PRP) gels. rBMSCs were isolated, cultured, and identified. The rBMSCs were subsequently co-transfected with two recombinant adenoviruses delivering PDGF-BB-GFP and BMP-2-GFP. PDGF-BB and BMP-2 expression levels in transduced BMSCs were detected, and a post-transfection analysis of the osteogenic differentiation trend of rBMSCs was performed. Autologous PRP gels were constructed and optimized, and the levels of growth factor in PRP were detected. The optimal growth conditions of the genetically-modified rBMSCs in the scaffolds were established, and the effects of tissue engineering materials and PRP gel construction on the osteogenic differentiation of rBMSCs were assessed. The results revealed that high-purity rBMSCs were obtained, and high levels of BMP-2 and PDGF-BB were secreted by the transduced cells. Furthermore, PRP promoted the proliferation and osteogenic differentiation of rBMSCs overexpressing PDGF-BB and BMP-2. Collectively, the results of the present study revealed that genetically modified rBMSCs incorporated into PRP gels enhanced osteogenic differentiation.

Transcriptome analysis of primary podocytes reveals novel calcium regulated regulatory networks.

Podocytes are pivotal in establishing the selective permeability of the glomerular filtration barrier. Recently, we showed that an increase of the intracellular calcium ion concentration [Ca2+ ] causes a rapid and transient actin reset (CaAR) measurable through live imaging microscopy using lifeact-mCherry as an actin dye in different cell types including the podocyte. This and other studies show the critical role [Ca2+ ] and the actin cytoskeleton play in podocyte homeostasis. To further investigate the role of [Ca2+ ] and the actin cytoskeleton in podocytes, we used a double fluorescent reporter mouse model to establish a primary podocyte culture system. We treated these podocytes temporarily with a Calcium Ionophore and facultatively with Latrunculin A, an inhibitor of actin polymerization. Unbiased genome wide transcriptional analysis identified a transcriptional response in podocytes to elevated [Ca2+ ] levels, affecting mRNA levels of PDGF-BB, RICTOR, and MIR17HG as mediators of Ca2+ -signaling. Comparison of the ex vivo transcriptional response from the primary podocyte culture with glomerular transcripts across a wide spectrum of CKD disease confirmed co-regulation of transcript sets, establishing the disease relevance of the model system. Our findings demonstrate novel [Ca2+ ] regulated gene networks in podocytes deepening our understanding of podocyte biology and disease.

Viral Dynamics and Immune Correlates of Coronavirus Disease 2019 (COVID-19) Severity.

BackgroundKey knowledge gaps remain in the understanding of viral dynamics and immune response of SARS-CoV-2 infection.MethodsWe evaluated these characteristics and established their association with clinical severity in a prospective observational cohort study of 100 patients with PCR-confirmed SARS-CoV-2 infection (mean age 46 years, 56% male, 38% with comorbidities). Respiratory samples (n=74) were collected for viral culture, serum samples for measurement of IgM/IgG levels (n=30), and plasma samples for levels of inflammatory cytokines and chemokines (n=81). Disease severity was correlated with results from viral culture, serologic testing, and immune markers.Results57 (57%) patients developed viral pneumonia, of whom 20 (20%) required supplemental oxygen including 12 (12%) invasive mechanical ventilation. Viral culture from respiratory samples was positive for 19 of 74 patients (26%). No virus was isolated when the PCR cycle threshold (Ct) value was >30 or >14 days after symptom onset. Seroconversion occurred at a median of 12.5 days (IQR 9-18) for IgM and 15.0 days (IQR 12-20) for IgG; 54/62 patients (87.1%) sampled at day 14 or later seroconverted. Severe infections were associated with earlier seroconversion and higher peak IgM and IgG levels. Levels of IP-10, HGF, IL-6, MCP-1, MIP-1α, IL-12p70, IL-18, VEGF-A, PDGF-BB and IL-1RA significantly correlated with disease severity.ConclusionWe found virus viability was associated with lower PCR Ct value in early illness. A stronger antibody response was associated with disease severity. The overactive proinflammatory immune signatures offers targets for host-directed immunotherapy which should be evaluated in randomised controlled trials.

A combination of circulating chemokines as biomarkers of obesity-induced insulin resistance at puberty.

In obesity adipose tissue undergoes structural re-modelling leading to a chronic low-grade inflammatory state linked to insulin resistance (IR). We aimed to develop a clinically relevant biomarker model for stratifying IR in adolescents with obesity. Cytokines [tumour cell derived factor 1α, monocyte chemoattract protein (MCP) 1, eotaxin and fractalkine], growth factors [brain-derived neurotrophic factor, pro-fibrotic platelet-derived growth factor (PDGF-BB) and insulin-like growth factor 1] and biochemical/metabolic factors were analysed in serum of 143 pubertal patients with obesity (50% IR; 50% non-IR) and 33 controls. Factor analysis, correlation, binary logistic regression and receiver operating characteristic analysis were used to evaluate combinations of these biomarkers as possible diagnostic tools for IR. Two biomarker IR models combining levels of triglycerides (TG)/HDL, eotaxin, MCP-1 and PDGF-BB in pubertal patients with obesity of both sexes were defined. Altered levels of MCP-1, eotaxin, and PDGF-BB constitute a main component that determines 27.7% of the variance explaining IR. Growth and inflammatory factors comprise two other components linked to the first, together accounting for 59.2% of the variance determining IR. PDGF-BB, MCP-1, eotaxin, TG and cholesterol concentrations constitute a solid panel of biomarkers associated with IR in pubertal children with obesity that could be useful in their stratification in a clinical setting for stratification.

Severity of intervertebral disc herniation regulates cytokine and chemokine levels in patients with chronic radicular back pain

The contributions of intervertebral disc disease and subject-specific covariates to systemic inflammation in low back pain are unknown. We examined the effects of symptomatic disc herniation (DH) and MRI herniation severity on serum cytokine levels in clinical subjects. Cytokine levels from lumbar DH subjects (N=78) were compared to control subjects (N=57) accounting for effects of DH, age, body mass index (BMI) and gender. Effect of DH severity on cytokine levels was analyzed on subsets of subjects with acute or chronic pain. Serum cytokines were also analyzed in a subset of patients between pre- and 3 months post-surgery. Cytokine levels were elevated in the serum of patients with symptomatic DH, and the covariates age, BMI and gender significantly contributed to levels of some cytokines. Severity of herniation was a significant contributor to pain intensity (VAS), serum levels of HMGB1, PDGFbb, and IL-9. The relationship between DH severity and cytokine levels was confirmed in subjects with chronic, but not acute symptoms. Serum levels of macrophage migration inhibitory factor (MIF) decreased, whereas levels of CCL3, CCL11, CXCL1, and CXCL10 were significantly elevated post surgery. This study is the first to show that DH severity is coordinately associated with changes in serum levels of inflammatory cytokines in chronic pain subjects. HMGB1, PDGFbb and IL-9 are novel mediators of increasing DH severity, indicative of cellular damage, neuro-inflammation and angiogenesis. Resolution of inflammation was observed with decrease in MIF post surgery. However, elevated chemokine levels indicate ongoing remodeling and wound healing at 3-month time point.

Analysis of Platelet Counts and Platelet-Derived Growth Factor-Bb Levels in Platelet Rich Plasma Produced with Edta as Anticoagulant in Three Different Centrifugation Methods

Background: Platelet rich plasma (PRP) is an autologous product made of whole blood through centrifugation process producing high platelet concentrate in a small volume of plasma. This high platelet concentrate can lead to high growth factor levels that play important roles in the nature of thrombosis, hemostasis and wound healing. PRP has been widely used in clinical setting, however standardized procedure of PRP production has been lacking, mainly the procedure related to duration and speed of centrifugation, and also anticoagulant used. Aim: To analyze the difference between platelet counts and PDGF-BB levels in PRP produced from various preparation methods (centrifugation speed and duration), using Ethylenediaminetetraacetic Acid /EDTA as anticoagulant. Method: This study used experimental laboratory design, involving 34 healthy volunteers that met inclusion criteria. These subjects were divided into three groups with different centrifugation methods that adopted from previous study. Results: The platelet counts was found in group 1, 2, and 3 with an increase of, 2,69, 3,69, and 2,53 times from the initial platelet counts respectively and there was significant difference between platelet counts before and after treatment on three groups that used EDTA (p=0,000, p=0,003, and p=0,002 for group 1, 2 and 3 respectively). PDGF-BB levels in this group was also higher than the remaining groups. Conclusion: The highest platelet counts and PDGF-BB levels was found in PRP with EDTA produced from first spin at 2800 RPM in five minutes, and second spin at 3800 RPM in seven minutes. This study suggests further research on qualitative assessments of platelets and PDGF-BB resulted from various preparation methods (centrifugation and anticoagulant usage).

Quantification of angiogenic factors and their clinicopathological associations in breast cancer.

Breast cancer (BC) is one of the top three common cancers in women, responsible for nearly one-third of all new cancer diagnoses. Angiogenesis plays a crucial role in BC progression. In this study, we aimed to measure the serum concentrations of eight angiogenic factors in BC patients and healthy controls and to assess their correlation with clinicopathological variables. In a case-control study, 62 pathologically confirmed BC patients as well as 54 age-matched controls were recruited. A bead-based immunoassay was used to measure serum levels of VEGF-A, ANG-2, PDGF-AA, PDGF-BB, EGF, TGF-α, HGF, and bFGF. We observed a significant elevation in serum levels of VEGF-A, EGF, and PDGF-AA in BC patients compared with the controls (P < 0.05). Patients with grade III had higher ANG-2 levels compared with those with grades I (P = 0.007) and II of the disease (P = 0.003). In addition, estrogen-positive and progesterone-positive BC patients had higher levels of TGF-α (P < 0.05). The significant elevation of VEGF-A, EGF, and PDGF-AA serum levels in BC patients suggests these cytokines might have diagnostic value as potential biomarkers in BC. Further large-scale studies are needed to generalize these results to all BC patients.