Basic FGF Research Articles

Contemporaneous Inflammatory, Angiogenic, Fibrogenic, and Angiostatic Cytokine Profiles of the Time-to-Tumor Development by Cancer Cells to Orchestrate Tumor Neovascularization, Progression, and Metastasis.

Cytokines can promote various cancer processes, such as angiogenesis, epithelial to mesenchymal transition (EMT), invasion, and tumor progression, and maintain cancer stem-cell-like (CSCs) cells. The mechanism(s) that continuously promote(s) tumors to progress in the TME still need(s) to be investigated. The data in the present study analyzed the inflammatory, angiogenic, fibrogenic, and angiostatic cytokine profiles in the host serum during tumor development in a mouse model of human pancreatic cancer. Pancreatic MiaPaCa-2-eGFP cancer cells were subcutaneously implanted in RAG2xCγ double mutant mice. Blood samples were collected before cancer cell implantation and every week until the end point of the study. The extracted serum from the blood of each mouse at different time points during tumor development was analyzed using a Bio-Plex microarray analysis and a Bio-Plex 200 system for proinflammatory (IL-1β, IL-10, IFN-γ, and TNF-α) and angiogenic and fibrogenic (IL-15, IL-18, basic FGF, LIF, M-CSF, MIG, MIP-2, PDGF-BB, and VEGF) cytokines. Here, we find that during cancer cell colonization for tumor development, host angiogenic, fibrogenic, and proinflammatory cytokine profiling in the tumor-bearing mice has been shown to significantly reduce host angiostatic and proinflammatory cytokines that restrain tumor development and increase those for tumor growth. The proinflammatory cytokines IL-15, IL-18, and IL-1β profiles reveal a significant host serum increase after day 35 when the tumor began to progress in growth. In contrast, the angiostatic cytokine profiles of TNFα, MIG, M-CSF, IL-10, and IFNγ in the host serum revealed a dramatic and significant decrease after day 5 post-implantation of cancer cells. OP treatment of tumor-bearing mice on day 35 maintained high levels of angiostatic and fibrogenic cytokines. The data suggest an entirely new regulation by cancer cells for tumor development. The findings identify for the first time how pancreatic cancer cells use host cytokine profiling to orchestrate the initiation of tumor development.

Circulating immunoglobulins and transient lymphocytopenia in a sub-study of CAPRISA 012B, testing HIV monoclonal antibodies in a phase 1 trial

Acute, transient lymphocytopenia, not clinically significant was observed in the CAPRISA 012B phase 1 clinical trial following administration of broadly neutralizing antibodies (bnAb)-CAP256V2LS alone or with VRC07-523LS. Lymphocytopenia was assigned upon a > 50% decline in absolute lymphocyte counts following bnAb administration. We posited that systemic immunoglobulins (Igs), and cytokine profiles of eight women who developed lymphocytopenia were different to the 12 women without lymphocytopenia. Plasma Ig subclasses (IgG)/isotypes (IgM/IgA), and 27 cytokines were measured at enrolment (prior to bnAbs) and at days 1, 7, 28, 56 post-bnAb administration. IgG subclasses, IgM and total lymphocyte counts were significantly lower prior to bnAbs in women with gradable lymphocytopenia than those without. Gradable lymphocytopenia compared to non-lymphocytopenia women had significantly higher MIP-1β from enrolment up to day 56. TNF-α was significantly lower in gradable lymphocytopenia compared to non-lymphocytopenia women for enrolment, days 7, 28 and 56 except for day 1. Within the gradable and within the non-lymphocytopenia women, from enrolment to day 1, significantly elevated IL-6, IL-8, IP-10, MCP-1, G-CSF and IL-1RA were found. Additionally, within the gradable lymphocytopenia women, 9 additional cytokines (TNF-α, MIP-1α, MIP-1β, RANTES, Basic FGF, eotaxin, IFN-γ, IL-17A and IL-4) were significantly elevated at day 1 post-bnAbs compared to enrolment. This sub study presents preliminary findings to support the monitoring of baseline immunological markers including lymphocyte counts for assessing the development of transient lymphocytopenia. In high-risk settings conducting clinical trials testing bnAbs for HIV prevention, understanding factors that could amplify rates of lymphocytopenia, even if transient, remain undefined.

Relationship between fibroblast growth factor in plasma and carotid plaque neovascularization: a pilot study.

Unstable atherosclerotic carotid plaques with intraplaque neovascularization (IPN) carry a substantial risk for ischemic stroke. Conventional ultrasound methods fall short in detecting IPN, where superb microvascular imaging (SMI) has emerged as a promising tool for both visualizing and quantification. High levels of fibroblast growth factor 23 (FGF-23) have, in observational studies, been suggested as related to cardiovascular morbidity and mortality. The association of FGF-23 to atherosclerotic carotid plaque instability remains relatively unexplored. A cohort of twenty-nine patients with ≥50% atherosclerotic carotid stenosis underwent conventional carotid ultrasound, SMI, and blood tests, including measurement of FGF-23 in plasma. Nineteen patients were characterized as symptomatic and ten as asymptomatic. Our major findings were: i) Higher FGF-23 levels were strongly correlated with increased SMI-assessed IPN. ii) Neo-vessel count recorded by quantitative SMI was positively correlated to increased FGF-23 levels, but not with basic FGF levels. (iii) In contrast, traditional risk factors for plaque instability exhibited no noteworthy associations with SMI-assessed IPN or with FGF-23 levels. This pilot study suggest the potential of FGF-23 as a valuable marker for neovascularization and atherosclerotic carotid plaque instability as a risk factor for ischemic stroke. Further research involving larger cohorts and prospective data is necessary to understand FGF-23's role in this context comprehensively.

Changes in Cytokine Levels in Patients with Severe Fever with Thrombocytopenia Syndrome Virus.

To characterize the cytokine profile of patients with severe fever with thrombocytopenia syndrome (SFTS) in relation to disease severity. 60 laboratory-confirmed SFTS patients and 12 healthy individuals from multi-centers in Shandong Province of China were included, and all patients were divided into fatal patients (9) and recovered patients (51) due to their final outcomes. Multiplex-microbead immunoassays were conducted to estimate levels of 27 cytokines in the sera of patients and controls. The results showed that levels of IL-2, IL-4, IL-6, IL-7, IL-8, IL-15, IL-1RA, G-CSF, GM-CSF, IFN-γ, TNF-α, basic FGF, PDGF-BB, RANTES, IP-10, MIP-1α, MIP-1β, MCP-1, and Eotaxin differed significantly among the SFTS fatal patients, recovered patients, and the healthy controls (all p<0.05). Compared to the healthy controls, the fatal patients and recovered patients had reduced levels of IL-2, IL-4, IL-7, PDGF-BB, RANTES, and Eotaxin, while the levels of PDGF-BB and RANTES were significantly lower in fatal patients compared to recovered patients. The increasing levels of IL-6, IL-8, IL-15, IL-1RA, G-CSF, GM-CSF, IFN-γ, TNF-α, basic FGF, IP-10, MIP-1α, MIP-1β, and MCP-1 were observed in fatal patients (all p<0.05), and the levels of IL-6, IP-10, MIP-1α, and MCP-1 were significantly higher than other two groups. The Spearman correlation analysis indicated a positive correlation between platelet count and PDGF-BB levels (p<0.05), while the white blood cell count had a negative correlation with MIP-1 level (p<0.05). The research exhibited that the SFTS virus (SFTSV) caused an atypical manifestation of cytokines. The levels of IL-6, IP-10, MIP-1α, and MCP-1 had been observed a positive association with the severity of the illness.

SARS-CoV-2 nucleocapsid protein, rather than spike protein, triggers a cytokine storm originating from lung epithelial cells in patients with COVID-19

PurposeThe aim of this study was to elucidate the factors associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that may initiate cytokine cascades and correlate the clinical characteristics of patients with coronavirus disease 2019 (COVID-19) with their serum cytokine profiles.MethodsRecombinant baculoviruses displaying SARS-CoV-2 spike or nucleocapsid protein were constructed and transfected into A549 cells and THP-1-derived macrophages, to determine which protein initiate cytokine release. SARS-CoV-2-specific antibody titers and cytokine profiles of patients with COVID-19 were determined, and the results were associated with their clinical characteristics, such as development of pneumonia or length of hospital stay.ResultsThe SARS-CoV-2 nucleocapsid protein, rather than the spike protein, triggers lung epithelial A549 cells to express IP-10, RANTES, IL-16, MIP-1α, basic FGF, eotaxin, IL-15, PDGF-BB, TRAIL, VEGF-A, and IL-5. Additionally, serum CTACK, basic FGF, GRO-α, IL-1α, IL-1RA, IL-2Rα, IL-9, IL-15, IL-16, IL-18, IP-10, M-CSF, MIF, MIG, RANTES, SCGF-β, SDF-1α, TNF-α, TNF-β, VEGF, PDGF-BB, TRAIL, β-NGF, eotaxin, GM-CSF, IFN-α2, INF-γ, and MCP-1 levels were considerably increased in patients with COVID-19. Among them, patients with pneumonia had higher serum IP-10 and M-CSF levels than patients without. Patients requiring less than 3 weeks to show negative COVID-19 tests after contracting COVID-19 had higher serum IP-10 levels than the remaining patients.ConclusionOur study revealed that nucleocapsid protein, lung epithelial cells, and IP-10 may be potential targets for the development of new strategies to prevent, or control, severe COVID-19.

The impact of prior SARS-CoV-2 infection on host inflammatory cytokine profiles in patients with TB or other respiratory diseases.

Tuberculosis (TB) and COVID-19 are the two leading causes of infectious disease mortality worldwide, and their overlap is likely frequent and inevitable. Previous research has shown increased mortality in TB/COVID-coinfected individuals, and emerging evidence suggests that COVID-19 may increase susceptibility to TB. However, the immunological mechanisms underlying these interactions remain unclear. In this study, we aimed to elucidate the impact of prior or concurrent COVID-19 infection on immune profiles of TB patients and those with other respiratory diseases (ORD). Serum and nasopharyngeal samples were collected from 161 Gambian adolescents and adults with either TB or an ORD. Concurrent COVID-19 infection was determined by PCR, while prior COVID-19 was defined by antibody seropositivity. Multiplex cytokine immunoassays were used to quantify 27 cytokines and chemokines in patient serum samples at baseline, and throughout treatment in TB patients. Strikingly, TB and ORD patients with prior COVID-19 infection were found to have significantly reduced expression of several cytokines, including IL-1β, TNF-α and IL-7, compared to those without (p<0.035). Moreover, at month-six of anti-TB treatment, seropositive patients had lower serum Basic FGF (p=0.0115), IL-1β (p=0.0326) and IL-8 (p=0.0021) than seronegative. TB patients with acute COVID-19 coinfection had lower levels of IL-8, IL-13, TNF-α and IP-10 than TB-only patients, though these trends did not reach significance (p>0.035). Our findings demonstrate that COVID-19 infection alters the subsequent response to TB and ORDs, potentially contributing to pathogenesis. Further work is necessary to determine whether COVID-19 infection accelerates TB disease progression, though our results experimentally support this hypothesis.

Trimethylamine-N-oxide (TMAO) and basic fibroblast growth factor (bFGF) are possibly involved in corticosteroid resistance in adult patients with immune thrombocytopenia

PurposeImmune thrombocytopenia (ITP) is an autoimmune disease characterized by accelerated platelet clearance. Gut dysbiosis was associated with its pathogenesis, but the underlying mechanisms have not been fully elucidated. Patients with ITP exhibit varying degrees of responsiveness to corticosteroid treatment. Therefore, prognostic indexes for corticosteroid responsiveness in ITP could offer valuable guidance for clinical practices. MethodsThe present study examined the signature of six types of gut-microbiota metabolites and forty-eight types of cytokines, chemokines, and growth factors and their clinical significance in patients with ITP. ResultsBoth patients with good and poor corticosteroid responsiveness exhibited significantly elevated/suppressed secretion of twenty-two cyto(chemo)kins/growth factors in comparison to healthy controls. Additionally, patients with ITP demonstrated a significant decrease in plasma levels of trimethylamine-N-oxide (TMAO), which was found to be negatively correlated to circulating platelet counts, and positively correlated with Interleukin (IL)-1β and IL-18. Notably, patients who exhibited poor response to corticosteroid treatment displayed elevated levels of TMAO and basic fibroblast growth factor (bFGF) in comparison to responders. Additionally, we found that the amalgamation of TMAO, bFGF and interleukin (IL)-13 could serve as a valuable prognostic tool for predicting CS responsiveness. ConclusionPatients with ITP were characterized overall by an imbalanced secretion of cyto(cheo)kins/growth factors and inadequate levels of TMAO. The varying degrees of responsiveness to corticosteroid treatment can be attributed to different profiles of basic FGF and TMAO that might be related to overburdened oxidative stress and inflammasome overactivation, and ultimately mediate corticosteroid resistance.

COVID-19 and candiduria: an investigation of the risk factors and immunological aspects.

Secondary fungal infections are frequently observed in COVID-19 patients. However, the occurrence of candiduria in these patients and its risk factors are underexplored. We evaluated the risk factors of candiduria in COVID-19 patients, including inflammatory mediators that could be used as prognostic markers. Clinical information, laboratory test results, and outcomes were collected from severely ill COVID-19 patients with and without candiduria. Candida species identification, antifungal susceptibility, and plasma inflammatory mediators' measurements were performed. Regression logistic and Cox regression model were used to evaluate the risk factors. A higher risk of longer hospitalization and mortality were observed in patients with candiduria compared to those with COVID-19 only. Candiduria was caused by Candida albicans, C. glabrata, and C. tropicalis. Isolates with intermediate susceptibility to voriconazole and resistant to caspofungin were identified. Classic factors such as the use of corticosteroids and antibacterials, the worsening of renal function, and hematological parameters (hemoglobin and platelets) were found to predispose to candiduria. The mediators IL-1β, IL-1ra, IL-2, CXCL-8, IL-17, IFN-γ, basic FGF, and MIP-1β were significantly increased in patients with COVID-19 and candiduria. Furthermore, IFN-γ, IL-1ra, and CXCL-8 were associated with the occurrence of candiduria in COVID-19 patients, whereas basic FGF, IL-1β, and CXCL-8 were associated with the risk of death in these patients. Classical and immunological factors were associated with worse prognosis among patients with COVID-19 and candiduria. Some mediators, especially CXCL-8, can be a reliable biomarker of fungal coinfection and may guide the diagnostic and the treatment of these patients.

Machine learning-based protein signatures for differentiating hypertensive disorders of pregnancy.

Hypertensive disorders of pregnancy (HDP) result in major maternal and fetal complications. Our study aimed to find a panel of protein markers to identify HDP by applying machine-learning models. The study was conducted on a total of 133 samples, divided into four groups, healthy pregnancy (HP, n = 42), gestational hypertension(GH, n = 67), preeclampsia (PE, n = 9), and ante-partum eclampsia(APE, n = 15). Thirty circulatory protein markers were measured using Luminex multiplex immunoassay and ELISA. Significant markers were screened for potential predictive markers by both statistical and machine-learning approaches. Statistical analysis found seven markers such as sFlt-1, PlGF, endothelin-1(ET-1), basic-FGF, IL-4, eotaxin and RANTES to be altered significantly in disease groups compared to healthy pregnant. Support vector machine(SVM) learning model classified GH and HP with 11 markers (eotaxin, GM-CSF, IL-4, IL-6, IL-13, MCP-1, MIP-1α, MIP-1β, RANTES, ET-1, sFlt-1) and HDP with 13 markers(eotaxin, G-CSF, GM-CSF, IFN-gamma, IL-4, IL-5, IL-6, IL-13, MCP-1, MIP-1β, RANTES, ET-1, sFlt-1). While logistic regression(LR) model classified PE with 13 markers(basic FGF, IL-1β, IL-1ra, IL-7, IL-9, MIP-1β, RANTES, TNF-alpha, nitric oxide, superoxide dismutase, ET-1, PlGF, sFlt-1) and APE by 12 markers (eotaxin, basic-FGF, G-CSF, GM-CSF, IL-1β, IL-5, IL-8, IL-13, IL-17, PDGF-BB, RANTES, PlGF). These markers may be used to diagnose the progression of healthy pregnant to a hypertensive state. Future longitudinal studies with large number of samples are needed to validate these findings.

Serum biomarkers for monitoring response to tuberculosis treatment: an assessment of the effect of different covariates among slow and fast treatment responders

Introduction The long duration of tuberculosis treatment, as well as the 2-year post-treatment follow-up period often required for predicting relapse, present a hindrance to drug development and treatment monitoring efforts. Therefore, there is need for treatment response biomarkers to inform treatment time shortening, clinical decision-making, and inform clinical trials. Objectives To assess the abilities of serum host biomarkers to predict treatment response among active PTB patients. Methods Active pulmonary TB patients (n = 53) as confirmed by sputum MGIT culture were enrolled at a TB treatment centre in Kampala, Uganda. We evaluated concentrations of 27 serum host biomarkers at baseline, month 2, and month 6 following the initiation of anti-tuberculosis treatment using the luminex platform for their ability to predict sputum culture status at month-2 post treatment initiation. Results There were significant differences in concentrations of IL1ra, IL1β, IL6, IP10, MCP-1, and IFNγ during treatment. A bio-signature comprising TTP, TNFα, PDGF-BB, IL9, and GCSF best predicted month 2 culture conversion with sensitivity and specificity of 82% (95% CI; 66 -92% and 57 -96% respectively). Slow anti-TB treatment responders had higher pro-inflammatory marker levels during treatment. The strongest correlation was observed between VEGF and IL12p70 (0.94), IL17A and basic FGF (0.92), basic FGF, and IL2 (0.88), and IL10 with IL17A (0.87). Conclusion We identified host biomarkers that predicted early response to PTB treatment, which may be valuable in future clinical trials and treatment monitoring. Similarly, strong correlations between biomarkers provide options for biomarkers substitutions during the development of treatment response monitoring tools or point of care tests.

Prognostic prediction of subjective cognitive decline in major depressive disorder based on immune biomarkers: a prospective observational study

ObjectiveSubjective cognitive decline (SCD) is highlighted in patients with major depressive disorder (MDD), which impairs objective cognitive performance and worsens the clinical outcomes. Immune dysregulation is supposed to be the potential mechanism of cognitive impairment. However, the peripheral immune biomarkers in patients troubled with MDD and SCD are not conventionally described.MethodsA prospective-observational study was conducted for 8 weeks. Subjective cognitive function was measured using the Chinese version of the 20-item perceived deficits questionnaire-depression (PDQ-D) and depression symptoms were evaluated with Hamilton Depression Rating Scale-17 (HDRS-17). Luminex assays were used to measure 48 immune cytokines in plasma at baseline. Integrating these results and clinicopathological features, a logistic regression model was used to develop a prognostic prediction.ResultsTotally, 114 patients were enrolled in this study. Among the patients who completed follow-up, 56% (N = 50) had residual subjective cognitive decline, and 44% (N = 50) did not. The plasma levels of FGF basic, INF-γ, IL-1β, MCP-1, M-CSF and SCF were increased and the levels of IL-9, RANTES and PDGF-BB were decreased in the SCD group. Additionally, Basic FGF, IFN-γ, IL-1β, and SCF were positively correlated and IL-9, RANTES, and PDGF-BB were negatively correlated with the PDQ-D scores after treatment. Notably, combinations of cytokines (SCF and PDGF-BB) and PDQ-D scores at baseline showed good performance (The area under the receiver operating characteristic curve = 0.818) in the prediction of subjective cognitive decline.ConclusionA prognostic model based on protein concentrations of SCF, PDGF-BB, and scores of PDQ-D showed considerable accuracy in predicting residual subjective cognitive decline in depression.

Role of Topical Ritodrine Hydrochloride in Experimentally Induced Hypertrophic Scar in Rabbits

Hypertrophic scars are fibroproliferative illnesses caused by improper wound healing, during that, excessive inflammation, angiogenesis, and differentiated human dermal fibroblast (HDF ) function contribute to scarring, whereas hyperpigmentation negatively affects scar quality. Over 100 million patients heal with a scar every year. To investigate the role of the beta 2 adrenergic receptor (β2AR); Ritodrine, in wound scarring, the ability of beta 2 adrenergic receptor agonist (β2ARag) to alter HDF differentiation and function, wound inflammation, angiogenesis, and wound scarring was explored in HDFs, zebrafish, chick chorioallantoic membrane assay (CAM), and a porcine skin wound model, respectively. A study identify a β2AR-mediated mechanism for scar reduction. β2ARag significantly reduced HDF differentiation, via multiple cAMP and/or fibroblast growth factor 2 or basic FGF (FGF2)-dependent mechanisms, in the presence of transforming growth factor betaβ1, reduced contractile function, and inhibited mRNA expression of a number of profibrotic markers. β2ARag also reduced inflammation and angiogenesis in zebrafish and CAMs in vivo, respectively. In Red Duroc pig full-thickness wounds, β2ARag reduced both scar area and hyperpigmentation by almost 50% and significantly improved scar quality. Indeed, mechanisms delineated in vitro and in other in vivo models were evident in the β2ARag-treated porcine scars in vivo. Both macrophage infiltration and angiogenesis were initially decreased, whereas DF function was impaired in the β2ARag-treated porcine wound bed. This data reveal the potential of β2ARag to improve skin scarring. The purpose of this study was to assess the therapeutic effect of topical Ritodrine hydrochloride on hypertrophic scars in rabbits. Thirty-two healthy male albino rabbits that divided in to 4 groups were included in the study (healthy; induced untreated hypertrophic scars; induced hypertrophic scars treated with 0.1% Triamcinolone acetonide (TAC) as a standard drug; and induced hypertrophic scars treated with 0.5% Ritodrine HCL gel twice daily for 21 days. Histopathology of skin sections, transforming growth factor beta1 TGFβ-1 level, and collagen III alpha1 in skin tissue were all used as outcome measures. Compared to the induced hypertrophic scar group; treatment with Ritodrine significantly reduced means of TGF β1 and collagen III (p ≤0.01); significantly reduce mean score of inflammation (p ≤0.001), significantly lowered scar size (P ≤ 0.001), and significantly lower mean scar height (P≤0.001), but no significant decrease in SEI (P&gt;0.05). Therapy of induced hypertrophic scar with topical Ritodrine was successfully effective in rabbits. It reduced the immunological score (TGF-β1, collagen III), inflammation, and scar size in a substantial way. This effect was comparable (except in terms of SEI) to topical Triamcinolone acetonide efficacy

Dynamic immune and exosome transcriptomic responses in patients undergoing psychostimulant methamphetamine withdrawal.

Methamphetamine (METH) addiction and withdrawal cause serious harm to both the immune system and nervous system. However, the pathogenesis remains largely unknown. Herein, we investigated the peripheral cytokines and exosomal transcriptome regulatory networks in the patients with METH use disorders (MUDs) undergoing withdrawal. Twenty-seven cytokines were simultaneously assessed in 51 subjects, including 22 at the acute withdrawal (AW) stage and 29 at the protracted withdrawal (PW) stage, and 31 age and gender-matched healthy controls (HCs). Compared to the HCs, significantly decreased levels of interleukin (IL)-1β, IL-9, IL-15, Basic FGF, and MIP1a, increased levels of IL-1rα, IL-6, Eotaxin IP-10, VEGF, and RANTES were identified in AW. These disturbances were mostly or partly restored to the baseline in PW. However, the cytokines IL-6, IL-7, and IL-12p70 were consistently increased even after one year of withdrawal. Besides, a significant decrease in CD3+T and CD4+T cell numbers was observed in AW, and the diminishment was restored to baseline in PW. Comparatively, there were no statistically significant changes in CD8+T, NK, and B cells. Furthermore, the exosomal mRNAs and long non-coding RNAs (lncRNA) were profiled, and the lncRNA-miRNA-mRNA networks were constructed and associated with METH AW and PW stages. Notably, the chemokine signaling was remarkably upregulated during AW. By contrast, the differentially expressed mRNAs/lincRNAs were significantly enriched in neurodegeneration-related diseases. Taken together, a group of METH withdrawal-related cytokines and exosomal mRNA/lncRNA regulatory networks were obtained, which provides a useful experimental and theoretical basis for further understanding of the pathogenesis of the withdrawal symptoms in MUDs.

EGFR Mutation-Harboring Lung Cancer Cells Produce CLEC11A with Endothelial Trophic and Tumor-Promoting Activities.

Simple SummaryTumor angiogenesis is an important step in the progression of solid tumors. Understanding the mechanisms involved in tumor vasculature formation is critical for improving anti-angiogenic strategies. In this study, we reported that EGFR mutation-containing lung cancer cells produced CLEC11A with endothelial trophic and tumor-promoting activities. CLEC11A could be a novel factor involved in tumor angiogenesis.The formation of new blood vessels in solid tumors is regulated by various endothelial trophic factors. We identified that CLEC11A, an extracellular C-type lectin, was over-expressed in lung cancer cell lines harboring mutated EGFR. CLEC11A expression was also frequently elevated in lung adenocarcinoma (LAC) tissues with EGFR mutation. CLEC11A-expressing H1299 cells formed larger tumors in nude mice than did the control cells. The CLEC11A-expressing tumors contained more CD31-positive cells, suggesting that they had a higher angiogenic activity. CLEC11A per se did not induce blood vessel formation, but enhanced angiogenesis triggered by VEGF-A or basic FGF in vivo. Additionally, the expression of small hairpin RNA against CLEC11A (shCLEC11A) in HCC827 LAC cells suppressed their tumorigenic ability. Purified CLEC11A exhibited a chemotactic ability, which is dependent on its integrin-binding RGD and LDT motifs, toward endothelial cells. This chemotactic activity was not affected by the presence of a VEGFR inhibitor. Conditioned medium produced by HCC827-shCLEC11A cells had diminished chemotactic ability toward endothelial cells. CLEC11A treatments increased the levels of active integrin β1 that were not associated with activation of focal adhesion kinases in endothelial cells. Our results indicated that CLEC11A was a factor of angiogenic potential and was involved in lung cancer tumorigenesis.

The effect of stromal vascular fraction and Platelet-Rich Plasma combination on basic Fibroblast Growth Factor serum level during anal trauma healing in a Wistar rat model

IntroductionStromal Vascular Fraction (SVF) and Platelet-Rich Plasma (PRP) application play important roles in the healing process by increasing basic Fibroblast Growth Factor (bFGF) secretion. This research assesses the effect of combined SVF and PRP local injection on bFGF levels, using an anal trauma model in Wistar rats. MethodTwenty-eight adult Wistar rats were divided into three groups. Groups A and B underwent modified surgical anal trauma and repair; Group A was treated with the SVF and PRP combination local injection, while Group B was treated with only normal saline. Subsequently, we examined bFGF levels in Groups A and B on days 1, 7, and 14. Group C consisted of healthy controls sacrificed on day 0 to obtain baseline data on bFGF levels. ResultsThe bFGF levels were higher in Group A than in Group B on every experimental day. The Repeated Measures test shows a significant increase in bFGF levels on day 1 (p = 0.000), day 7 (p = 0.000), and day 14 (p = 0.000). This test also indicates that the local injection combination of SVF and PRP increased bFGF levels by 96.2% compared to the placebo group. ConclusionThe combination of SVF and PRP can increase bFGF levels during anal trauma healing in the Wistar rat model. Basic FGF is an important factor throughout the anal trauma healing process.

The Role for miR-146b-5p in the Attenuation of Dermal Fibrosis and Angiogenesis by Targeting PDGFRα in Skin Wounds

As a candidate microRNA antifibrotic effector in skin wounds, miR-146b-5p was upregulated by basic FGF, and PDGFRα was identified as a direct target of miR-146b-5p in fibroblasts. The treatment of fibroblasts with a miR-146b-5p mimic markedly downregulated the expression of PDGFRα and collagen type I. miR-146b-5p mimic transfection in wounds markedly attenuated cutaneous fibrosis, whereas a miR-146b-5p inhibitor strongly promoted fibrosis, with increases in PDGFRα and collagen I levels. These results indicate the positive effects of miR-146b-5p for the suppression of fibrosis, possibly through the inhibition of PDGFRα. The miR-146b-5p inhibitor markedly increased CD34+ vessel numbers and CD34 expression in wounds. We found miR-146b-5p+ cells in close contact with S100+ adipocytes. Moreover, we discovered the specific colocalization of the exosome marker CD81 and miR-146b-5p in the adipose tissue cells of mimic-transfected wounds, with miR-146b-5p signals being detected in the FSP1+ fibroblastic cells of adipose tissues. Therefore, fibroblastic cells of adipose tissues, which may specifically pick up and contain miR-146b-5p by exosome after transfection, may play an important role in the suppression of fibrosis. In this process, the inhibition of PDGFRα in adipose tissue cells by miR-146b-5p may lead to the loss of their PDGFRα-induced profibrotic activities, thereby suppressing fibrosis.

Periostin gene expression in neu-positive breast cancer cells is regulated by a FGFR signaling cross talk with TGF\u03b2/PI3K/AKT pathways

BackgroundBreast cancer is a highly heterogeneous disease with multiple drivers and complex regulatory networks. Periostin (Postn) is a matricellular protein involved in a plethora of cancer types and other diseases. Postn has been shown to be involved in various processes of tumor development, such as angiogenesis, invasion, cell survival and metastasis. The expression of Postn in breast cancer cells has been correlated with a more aggressive phenotype. Despite extensive research, it remains unclear how epithelial cancer cells regulate Postn expression.MethodsUsing murine tumor models and human TMAs, we have assessed the proportion of tumor samples that have acquired Postn expression in tumor cells. Using biochemical approaches and tumor cell lines derived from Neu+ murine primary tumors, we have identified major regulators of Postn gene expression in breast cancer cell lines.ResultsHere, we show that, while the stromal compartment typically always expresses Postn, about 50% of breast tumors acquire Postn expression in the epithelial tumor cells. Furthermore, using an in vitro model, we show a cross-regulation between FGFR, TGFβ and PI3K/AKT pathways to regulate Postn expression. In HER2-positive murine breast cancer cells, we found that basic FGF can repress Postn expression through a PKC-dependent pathway, while TGFβ can induce Postn expression in a SMAD-independent manner. Postn induction following the removal of the FGF-suppressive signal is dependent on PI3K/AKT signaling.ConclusionOverall, these results reveal a novel regulatory mechanism and shed light on how breast tumor cells acquire Postn expression. This complex regulation is likely to be cell type and cancer specific as well as have important therapeutic implications.

Cytokine Analysis in 154 Patients with Transient Abnormal Myelopoiesis: Jccg JPLSG TAM-10 Clinical Study

Introduction:Transient abnormal myelopoiesis (TAM), also known as transient leukemia or transient myeloproliferative disorder, is a unique clonal myeloproliferation characterized by immature megakaryoblasts occurring in 10% of neonates with Down syndrome. Although most patients show spontaneously resolution of TAM without therapeutic interventions, approximately 20% of TAM cases result in early deaths, i.e., within 9 months, and approximately 20% of the survivors develop acute megakaryoblastic leukemia (AMKL) within 4 years. A somatic GATA1 gene mutation that leads to the exclusive expression of a truncated GATA1 protein is shared by both TAM and AMKL cells. According to previous reports, cytokine levels are associated with liver failure, which is a cause of early death. Here, we analyzed 154 DS patients with TAM enrolled in the TAM-10 prospective observational study conducted by the Japan Pediatric Leukemia/Lymphoma Study Group to determine the association between clinical characteristics and cytokine levels in such patients.Patients and Methods:A total of 167 neonates (89 boys and 78 girls) diagnosed with TAM were prospectively registered in the TAM-10 study between May 2011 and February 2014. We analyzed cytokine levels in 154 of the 167 enrolled patients whose samples were available. Somatic GATA1 gene mutations were confirmed in 151 (98%) of 154 patients using Sanger and/or next-generation sequencing. Using the Bio-Prex cytokine assay, serum concentrations of the following 27 cytokines were measured: interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), vascular endothelial growth factor (VEGF), platelet-derived growth factor-BB (PDGF-BB), basic fibroblast growth factor (bFGF), granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory proteins alpha and beta (MIP-1α and MIP-1β), eotaxin, interferon gamma-induced protein (IP-10), regulated upon activation, normal T-cell expressed and secreted, interleukin I receptor agonist (IL-RA), and 13 different interleukins (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-15, and IL-17). For all analyses, P values were two-tailed and a P value of <0.05 was considered statistically significant. Mann-Whitney test were used as appropriate for comparisons between groups. Moreover, the cumulative incidence for competing events was compared with the Gray test.Results:The median (range) white blood cell (WBC) count at diagnosis was 37.2 (2.4-478.7) × 10 9 cells/L. Forty-seven (31%) of 154 patients received low-dose cytarabine. When we compared 29 patients with a high WBC count (≥100 × 10 9 cells/L, known as a poor prognostic factor in TAM patients) to 125 patients without a high WBC count for 27 cytokine levels, the levels of 16 cytokines (IL-1b, IL-1ra, IL-6, IL-7, IL-8, IL-9, IL-10, IL-13, Eotaxin, PDGF-bb, basic FGF, G-CSF, GM-CSF, MCP-1b, and VEGF) were significantly higher in patients with a high WBC group. Early death occurred in 14 (9%) of 154 patients. Cytokine levels were compared between the early death group (n = 14) and remaining patients (n = 140) and it was observed that the levels of 10 cytokines (IL-1b [p = 0.016], IL-1ra [p < 0.001], IL-6 [p < 0.001], IL-7 [p = 0.009], IL-8 [p < 0.001], IL-10 [p = 0.014], IL-13 [p = 0.002], MCP-1 [p = 0.030], MIP-1b [p = 0.024], and TNF-α [p = 0.008]) were significantly higher in the early death group. When the patients were divided in two groups according to the median IL-1β concentration showing the lowest p-value, the early death rate in the IL-1β high group was significantly higher than that in the IL-1β low group (16% vs. 3%, p < 0.001). IL-1, IL-6, IL-8, and TNF-α are proinflammatory cytokines induced by MCP-1 and MIP1-b. Early death was strongly associated with hypercytokinemia, suggesting that therapeutic interventions (e.g., systemic steroid therapy) may be effective in patients with hypercytokinemia. However, there was no relation between the levels of 27 cytokines and leukemia development.Conclusion:Our findings suggested that measurement of cytokine levels may be a useful marker for predicting early death and an indicator of therapeutic interventions required in TAM patients. DisclosuresNo relevant conflicts of interest to declare.

Cytokine and Chemokine Profile Changes in Patients with Neovascular Age-Related Macular Degeneration After Intravitreal Ranibizumab Injection for Choroidal Neovascularization.

ObjectiveTo investigate the concentrations of cytokine and chemokines profiling in aqueous humor for choroidal neovascularization (CNV) due to neovascular age-related macular degeneration (nAMD) before and during Intravitreal injection of ranibizumab (IVR) and its relation with the disease’s active state.MethodsThe cytokine levels in aqueous humour were detected by the Bio-Plex® 200 System and the Bio-Plex™ Human Cytokine Standard 27-Plex, Group I. Aqueous humour samples of experimental group were collected from 19 patients diagnosed nAMD at baseline and at 1 month after IVR. Aqueous humour samples of control group were collected from 20 patients undergoing cataract surgery.ResultsAqueous humor levels of basic fibroblast growth factor (basic FGF) and RANTES were significantly lower in nAMD patients than in the control group (P=0.044 and P<0.001, respectively). Vascular endothelial growth factor-A (VEGF-A) was significantly higher in nAMD patients than in the control group (P < 0.001). The average Eotaxin levels were significantly higher in nAMD patients after IVR than before (P=0.03). Contrarily, the average VEGF-A levels were significantly lower in AMD patients after IVR than before (P < 0.001).ConclusionAngiogenic, growth factors and inflammatory are involved in the formation of neovascularization of AMD patients. IVR did not cause significant differences in any growth factors or inflammatory cytokines in nAMD patients with the exception of VEGF.

Abstract PO-094: Human pluripotent stem cells acquire malignancy under tumor microenvironment

Abstract Cancer stem cell (CSC) theory suggests that cancer tissue contains a subpopulation of stem-like cells with self-renewal and differentiation potential as normal stem cells maintain organs by self-renewing and differentiation. Even in both cases of malignancy and normal the stemness is affected by the niche surrounding the stem cells. In the initial stage no stem cells should be related with cancer. The most important question is “How come the stem cells acquire malignancy?” Here in this study, using human induced pluripotent stem cells (iPSCs) we tried to generate cancer stem cells in the presence of the conditioned medium (CM) derived from human liver cancer cell line Hep 3B cells and human breast cancer cell line BT459 cells. The CM from both cells was collected when the cells became confluent and sterilized by 0.22 micrometer filter. The colonies of human iPSCs were detached from a mitomycin treated fibroblast feeder layer by collagenase and manually dissected into small pieces which were transferred onto ultra-low-attachment dishes. After 7 days the formed embryoid bodies (EBs) of human iPSCs were plated on Matrigel coated plates and maintained in Repro FF2 medium containing 5 ng/ml of basic FGF. Just after 24 hours from plating on Matrigel, the EBs of human iPSCs were treated with CM mixed with Repro FF2, in the ratio of 1:1. The medium was changed every two days with CM for 4 weeks. EBs cultured in the complete Repro FF2 medium without CM were prepared as the control but did not survive for 4 weeks. The survived cells were analyzed by RT-qPCR analysis and Immunofluorescence (IF) staining to confirm the expression of stemness and CSC markers and evaluated the conversion of iPSCs into CSCs. Distinct expression of self-renewal and CSC markers, such as Nanog, SOX2, CD44 and EPCAM, were observed in the survived cells. Sphere formation assay further confirmed the potential of self-renewal in the survived cells. The survived cells possessed differentiation potential into vascular endothelial-like cells. Thus, we concluded the iPSCs were converted into CSCs (hiPS-CSCs). Through these experiments, 2 different conditions derived from 2 independent cell lines Hep3B and BT549 cells were successful to hiPS-CSCs as hiPS-Hep3Bcm and hiPS-BT549cm cells. Furthermore, being treated with paclitaxel, the hiPS-CSCs exhibited potential of chemo-resistance maintaining the intact clonogenic morphology when compared to hiPSC. This study tries to summarize human cancer disease by utilizing the tumor microenvironment in the form of conditioned medium without any genetic manipulation. This should pave the way for the establishment of effective therapeutic agents. Citation Format: Said M. Afify, Ghmkin Hassan, Akimasa Seno, Yoshiaki Iwasaki, Masaharu Seno. Human pluripotent stem cells acquire malignancy under tumor microenvironment [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-094.