Irradiated Fibroblasts Research Articles

Plasma-Derived Exosomes Boost the Healing of Irradiated Wound by Regulating Cell Proliferation and Ferroptosis.

Ionizing radiation (IR) therapy for malignant tumors can damage adjacent tissues, leading to severe wound complications. Plasma-derived exosome treatment has recently emerged as a safe and impactful cell-free therapy. Herein, we aimed to determine whether plasma-derived exosomes could improve the healing of post-radiation wound. Rat plasma-derived exosomes (RP-Exos) were locally injected on cutaneous wounds created on the backs of irradiated rats and boosted the healing process as well as the deposition and remodeling of the extracellular matrix with collagen formation. Subsequently, the effects of RP-Exos were further evaluated on irradiated fibroblasts in vitro. The results suggested that exosomes promoted fibroblast proliferation, migration, cell cycle progression, and cell survival. Moreover, transcriptome sequencing analysis and quantitative polymerase chain reaction validation were performed to identify potential mechanisms. RPExos enhanced the expression of cell proliferation and radioresistance-related genes, and yet downregulated ferroptosis pathway in irradiated fibroblasts. Inhibition of ferroptosis by RP-Exos was further confirmed through colorimetric assay, fluorescence probe and flow cytometry in ferroptosis-induced fibroblasts. Our results suggest that RP-Exos regulate cell proliferation and ferroptosis in irradiated fibroblasts, thereby boosting the healing of irradiated wound. These findings support plasma-derived exosomes as a potential therapeutic method for post-radiation wound complications.

MiR-21 is upregulated, promoting fibrosis and blocking G2/M in irradiated rat cardiac fibroblasts.

BackgroundRadiation exposure of the thorax is associated with a greatly increased risk of cardiac morbidity and mortality even after several decades of advancement in the field. Although many studies have demonstrated the damaging influence of ionizing radiation on cardiac fibroblast (CF) structure and function, myocardial fibrosis, the molecular mechanism behind this damage is not well understood. miR-21, a small microRNA, promotes the activation of CFs, leading to cardiac fibrosis. miR-21 is overexpressed after irradiation; however, the relationship between increased miR-21 and myocardial fibrosis after irradiation is unclear. This study was conducted to investigate gene expression after radiation-induced CF damage and the role of miR-21 in this process in rats.MethodsWe sequenced irradiated rat CFs and performed weighted correlation network analysis (WGCNA) combined with differentially expressed gene (DEG) analysis to observe the effect on the expression profile of CF genes after radiation.ResultsDEG analysis showed that the degree of gene changes increased with the radiation dose. WGCNA revealed three module eigengenes (MEs) associated with 8.5-Gy-radiation—the Yellow, Brown, Blue modules. The three module eigengenes were related to apoptosis, G2/M phase, and cell death and S phase, respectively. By blocking with the cardiac fibrosis miRNA miR-21, we found that miR-21 was associated with G2/M blockade in the cell cycle and was mainly involved in regulating extracellular matrix-related genes, including Grem1, Clu, Gdf15, Ccl7, and Cxcl1. Stem-loop quantitative real-time PCR was performed to verify the expression of these genes. Five genes showed higher expression after 8.5 Gy-radiation in CFs. The target genes of miR-21 predicted online were Gdf15 and Rsad2, which showed much higher expression after treatment with antagomir-miR-21 in 8.5-Gy-irradiated CFs. Thus, miR-21 may play the role of fibrosis and G2/M blockade in regulating Grem1, Clu, Gdf15, Ccl7, Cxcl1, and Rsad2 post-irradiation.

Radiation-induced DNA damage and repair effects on 3D genome organization

The three-dimensional structure of chromosomes plays an important role in gene expression regulation and also influences the repair of radiation-induced DNA damage. Genomic aberrations that disrupt chromosome spatial domains can lead to diseases including cancer, but how the 3D genome structure responds to DNA damage is poorly understood. Here, we investigate the impact of DNA damage response and repair on 3D genome folding using Hi-C experiments on wild type cells and ataxia telangiectasia mutated (ATM) patient cells. We irradiate fibroblasts, lymphoblasts, and ATM-deficient fibroblasts with 5 Gy X-rays and perform Hi-C at 30 minutes, 24 hours, or 5 days after irradiation. We observe that 3D genome changes after irradiation are cell type-specific, with lymphoblastoid cells generally showing more contact changes than irradiated fibroblasts. However, all tested repair-proficient cell types exhibit an increased segregation of topologically associating domains (TADs). This TAD boundary strengthening after irradiation is not observed in ATM deficient fibroblasts and may indicate the presence of a mechanism to protect 3D genome structure integrity during DNA damage repair.

Conditional reprogramming culture conditions facilitate growth of lower-grade glioma models.

The conditional reprogramming cell culture method was developed to facilitate growth of senescence-prone normal and neoplastic epithelial cells, and involves co-culture with irradiated fibroblasts and the addition of a small molecule Rho kinase (ROCK) inhibitor. The aim of this study was to determine whether this approach would facilitate the culture of compact low-grade gliomas. We attempted to culture 4 pilocytic astrocytomas, 2 gangliogliomas, 2 myxopapillary ependymomas, 2 anaplastic gliomas, 2 difficult-to-classify low-grade neuroepithelial tumors, a desmoplastic infantile ganglioglioma, and an anaplastic pleomorphic xanthoastrocytoma using a modified conditional reprogramming cell culture approach. Conditional reprogramming resulted in robust increases in growth for a majority of these tumors, with fibroblast conditioned media and ROCK inhibition both required. Switching cultures to standard serum containing media, or serum-free neurosphere conditions, with or without ROCK inhibition, resulted in decreased proliferation and induction of senescence markers. Rho kinase inhibition and conditioned media both promoted Akt and Erk1/2 activation. Several cultures, including one derived from a NF1-associated pilocytic astrocytoma (JHH-NF1-PA1) and one from a BRAF p.V600E mutant anaplastic pleomorphic xanthoastrocytoma (JHH-PXA1), exhibited growth sufficient for preclinical testing in vitro. In addition, JHH-NF1-PA1 cells survived and migrated in larval zebrafish orthotopic xenografts, while JHH-PXA1 formed orthotopic xenografts in mice histopathologically similar to the tumor from which it was derived. These studies highlight the potential for the conditional reprogramming cell culture method to promote the growth of glial and glioneuronal tumors in vitro, in some cases enabling the establishment of long-term culture and in vivo models.

Exosomal MiR-769-5p Exacerbates Ultraviolet-Induced Bystander Effect by Targeting TGFBR1.

Exosomal microRNAs have been investigated in bystander effect, but it is unclear whether microRNA works in ultraviolet radiation–induced bystander effects (UV-RIBEs) and what the underlying mechanism could be. Exosomes from ultraviolet (UV)–irradiated human skin fibroblasts (HSFs) were isolated and transferred to normal HSFs, followed by the detection of proliferation rate, oxidative damage level, and apoptosis rate. Exosomal miRNAs were evaluated and screened with miRNA sequencing and quantitative reverse transcriptase–polymerase chain reaction method. MiRNA shuttle and bystander photodamage reactions were observed after transfection of miR-769-5p. MiR-769-5p targeting gene transforming growth factor-β1 (TGFBR1), and TGFBR1 mRNA 3′-untranslated region (UTR) was assessed and identified by Western blotting and dual-luciferase reporter assay. Bystander effects were induced after being treated with isolated exosomes from UV-irradiated HSFs. Exosomal miR-769-5p expression was significantly upregulated. Human skin fibroblasts showed lower proliferation, increasing oxidative damage, and faster occurrence of apoptosis after transfection. Exosome-mediated transfer of miR-769-5p was observed. Upregulation of miR-769-5p induced bystander effects, whereas downregulation of miR-769-5p can suppress UV-RIBEs. In addition, miR-769-5p was found to downregulate TGFBR1 gene expression by directly targeting its 3′-UTR. Our results demonstrate that exosome-mediated miR-769-5p transfer could function as an intercellular messenger and exacerbate UV-RIBEs. MiR-769-5p inhibits the expression of TGFBR1 by targeting TGFBR1 mRNA 3′-UTR.

Collagen and soy peptides attenuate contractile loss from UVA damage and enhance the antioxidant capacity of dermal fibroblasts.

Wrinkles and extracellular matrix (ECM) loss are common signs of skin aging and are thought to be the result of damage caused by reactive oxygen species (ROS); ROS induces an imbalance between ECM degradation and production. In this study, we evaluate soy peptides (SP) and collagen peptides (CP), alone and in combination, for their ability to inhibit ROS formation and increase ECM gene expression in order to ameliorate the signs of skin aging. Using tert-Butyl hydroperoxide (t-BuOOH)-treated dermal fibroblasts, we explored the potential of CP and SP to inhibit ROS formation by flow cytometry, as well as their effect on ECM component genes by real-time quantitative PCR. In addition, we examined the effect of CP and SP on UVA irradiated fibroblasts in a 3D collagen lattice model that measured contractility. The results showed that the combination of CP and SP synergistically reduces ROS formation. This combination also increased expression of collagen I, collagen II, elastin, and fibronectin in t-BuOOH-treated or untreated dermal fibroblasts. In the UVA-treated 3D collagen lattice model, the results show that CP and SP significantly improved fibroblast contractility when compared to UVA control (P<0.05). In conclusion, CP and SP attenuate the loss of contractility due to UVA damage, inhibit t-BuOOH-induced ROS formation, and improve expression of ECM component genes.

OC-0081: Immuno-regulatory properties of irradiated tumor fibroblasts

OC-0081: Immuno-regulatory properties of irradiated tumor fibroblasts

Activated Beta-Catenin Signaling Ameliorates Radiation-Induced Skin Injury by Suppressing Marvel D3 Expression.

Radiation-induced skin injury remains a serious concern for cancer radiotherapy, radiation accidents and occupational exposure, and the damage mainly occurs due to apoptosis and reactive oxygen species (ROS) generation. There is currently no effective treatment for this disorder. The β-catenin signaling pathway is involved in the repair and regeneration of injured tissues. However, the role of the β-catenin signaling pathway in radiation-induced skin injury has not been reported. In this study, we demonstrated that the β-catenin signaling pathway was activated in response to radiation and that its activation by Wnt3a, a ligand-protein involved in the β-catenin signaling pathway, inhibited apoptosis and the production of ROS in irradiated human keratinocyte HaCaT cells and skin fibroblast WS1 cells. Additionally, Wnt3a promoted cell migration after irradiation. In a mouse model of full-thickness skin wounds combined with total-body irradiation, Wnt3a was shown to facilitate skin wound healing. The results from RNA-Seq revealed that 24 genes were upregulated and 154 were downregulated in Wnt3a-treated irradiated skin cells, and these dysregulated genes were mainly enriched in the tight junction pathway. Among them, Marvel D3 showed the most obvious difference. We further found that the activated β-catenin signaling pathway stimulated the phosphorylation of JNK by silencing Marvel D3. Treatment of irradiated cells with SP600125, a JNK inhibitor, augmented ROS production and impeded cell migration. Furthermore, treatment with Wnt3a or transfection with Marvel D3-specific siRNAs could reverse the above effects. Taken together, these findings illustrate that activated β-catenin signaling stimulates the activation of JNK by negatively regulating Marvel D3 to ameliorate radiation-induced skin injury.

3105 – ENHANCED OUTPUT OF PRIMITIVE HEMATOPOIETIC CELL PHENOTYPES IN TERATOMAS GENERATED FROM HUMAN INDUCED PLURIPOTENT STEM CELLS (HIPSCS) IN MORE PERMISSIVE IMMUNODEFICIENT MICE

Investigation of human hematopoietic stem cells (HSCs) has been revolutionized by advances in cell separation and clonal tracking of cells produced in transplanted patients and immunodeficient mice. However, methods able to expand them significantly remain elusive. iPSCs are of continuing interest as an alternative source, although robust conditions that support iPSC generation of human HSC either in vitro or in vivo have also not yet been identified. To determine whether the use of a more compatible in vivo host environment might offer an improvement, we compared the outputs of teratomas generated from human iPSC transplanted (± irradiated mouse fibroblasts engineered to produce human FLT3-L, SCF and IL3) into immunodeficient NOD-Rag1-null- IL2Rgc-null mice with a c-kit deficiency with or without a transgenic source of human IL3, GM-CSF and SCF (NRG-W41± 3GS mice). 6-8 week teratomas generated in the NRG-W41-3GS mice contained 40-fold more human CD34+CD45+ cells than those produced in NRG-W41 mice with yields up to 7 × 106 CD45+ cells. Co-injection of human growth factor-producing mouse fibroblasts further increased the production of human CD34+CD45+ ∼6-fold in the NRG-W41-3GS mice that extended to the phenotypically early GPI80+ and CD49f+ cells within the CD45+CD34+CD38-CD45RA-CD90+ subset. However, despite their display of short-term granulopoietic and erythroid differentiation potential in vitro, no evidence of in vivo engrafting potential was detected. These experiments suggest a persisting failure of conditions in the teratomas to support the acquisition of certain functional properties that primitive human cells require to engraft and set the stage for their future identification.

Glycosyl flavones from Humulus japonicus suppress MMP-1 production via decreasing oxidative stress in UVB irradiated human dermal fibroblasts

Exposure to Ultraviolet (UV) light induces photoaging of skin, leading to wrinkles and sunburn. The perennial herb Humulus japonicus, widely distributed in Asia, is known to have anti-inflammatory, antimicrobial, and antioxidant effects. However, the physiological activities of isolated compounds from H. japonicus have rarely been investigated. This study focused on the isolation of active compounds from H. japonicus and the evaluation of their effects on photoaging in UVB-irradiated human fibroblast (Hs68) cells. When the extract and four fractions of H. japonicus were treated respectively in UVB-irradiated Hs68 cells to investigate anti-photoaging effects, the ethyl acetate (EtOAc) fraction showed the strongest inhibitory effect on MMP-1 secretion. From EtOAc fraction, we isolated luteolin-8-C-glucoside (1), apigenin-8-C-glucoside (2), and luteolin-7-O-glucoside (3). These compounds suppressed UVB-induced MMP-1 production by inhibiting the phosphorylation of the mitogen-activated protein kinases (MAPKs) and activator protein-1 (AP-1). When the antioxidant activity of the compounds were estimated by conducting western blot, calculating the bond dissociation energies of the O-H bond (BDE) at different grade, and measuring radical scavenging activity, we found luteolin-8-C-glucoside (1) showed the strongest activity on the suppression of UVB-induced photoaging. These results demonstrate the inhibitory effect of three flavone glycosides derived from H. japonicus on MMP-1 production, MAPK and AP-1 signaling, and oxidative stress; this could prove useful in suppressing UVB induced photoaging.

Conditional reprogramming: Modeling urological cancer and translation to clinics.

Patient‐derived models, including cell models (organoids and conditionally reprogrammed cells [CRCs]) and patient‐derived xenografts, are urgently needed for both basic and translational cancer research. Conditional reprogramming (CR) technique refers to a co‐culture system of primary human normal or tumor cells with irradiated murine fibroblasts in the presence of a Rho‐associated kinase inhibitor to allow the primary cells to acquire stem cell properties and the ability to proliferate indefinitely in vitro without any exogenous gene or viral transfection. Considering its robust features, the CR technique may facilitate cancer research in many aspects. Under in vitro culturing, malignant CRCs can share certain genetic aberrations and tumor phenotypes with their parental specimens. Thus, tumor CRCs can promisingly be utilized for the study of cancer biology, the discovery of novel therapies, and the promotion of precision medicine. For normal CRCs, the characteristics of normal karyotype maintenance and lineage commitment suggest their potential in toxicity testing and regenerative medicine. In this review, we discuss the applications, limitations, and future potential of CRCs in modeling urological cancer and translation to clinics.

Effect of different irradiation doses on human and murine fibroblasts used as feeder layer for keratinocytes’ culture

Background & Aim The establishment of human keratinocytes’ culture (KTs) to obtain human Tissue Engineered Skin Substitutes is under research nowadays. Best results have been achieved culturing KTs on a feeder layer of irradiated 3T3 murine fibroblasts (i3T3s). Nevertheless, molecular effects of murine cells on KTs is unknown and for this reason, irradiated human fibroblasts (iHFs) have been studied. The objective of this study is to evaluate the effect of different irradiation doses on cell reducing power and cell proliferation of both fibroblasts. Methods, Results & Conclusion Methods 3T3s were obtained from a commercial vial of 3T3-Swiss Albino. HFs were isolated from human skin biopsies. After culturing and final recovery, both cell types were irradiated to doses of 50 and 100 Gy to constitute the feeder. Non-irradiated cells were used as control. Each condition was cultured 15 days in two 24-well plates (2 cm2/well). In each plate, cell densities cultured in triplicate were: 7,500 cells/cm2 and 10,000 cells/cm2. One plate was used for reducing power analysis; using a dilution of PrestoBlue™ (10%). The other plate was used for cell proliferation analysis; using Trypan Blue solution. N for each plate was 3 and a value of p Results Analysis of each type of feeder (Figs. 1-2) revealed significant differences at Days 10 and 15, between both irradiation doses and non-irradiated cells. In the case of iHFs, irradiation dose had a significant effect on cell reducing power, mainly at Day 15. Analysis of i3T3s revealed that the effect of irradiation on reducing power was extremely significant from Day 3 to 15, but no dose-dependent. A comparative analysis (Fig. 3) indicated that the number of non-irradiated cells after 15 days was higher in the case of 3T3s than HFs (644,620 cells vs 143,740 cells), but cell reducing power was lower in the case of murine fibroblasts (72.33% vs 80.67%). In the case of irradiated cells, iHFs were more resistant to irradiation in both, cell reducing power (27.5% vs 4.92%) and cell proliferation (5,962 cells vs 1,351 cells) at Day 15. Conclusion No differences exist considering initial cell density or irradiation dose in both feeders. Comparative analysis revealed that irradiation process is more effective in 3T3s than in HFs. This study needs to be completed with the incorporation of KTs on the different feeder conditions analyzed. Supported by PI17/02083, SAS PI-0458-2016 and PhD fellowship FI18/00269–Alvaro Sierra-Sanchez).

Antioxidative photochemoprotector effects of cerium oxide nanoparticles on UVB irradiated fibroblast cells

The Ultraviolet-B radiation (UVB) might induce cellular redox imbalance which plays an important role in the development of skin disorders. Thus, the search for photochemoprotective alternatives with antioxidant efficacy would be a safe aspect towards prevention of skin diseases. Cerium oxide nanoparticles (CNPs) have antioxidant properties, that are mostly related to CNPs catalase and superoxide dismutase (SOD)-like antioxidative mimetic activity. Considering that, we investigated whether CNPs induce photochemoprotection against UVB-induced cellular damages on L929 fibroblasts. Our results showed that CNPs prevented UVB mediated L929 cell oxidative damage by reestablishing the oxidative balance through ameliorating the reactive oxygen species (ROS) level and enhancing the antioxidant enzyme activities.

Therapeutic Reversal of Radiotherapy Injury to Pro-fibrotic Dysfunctional Fibroblasts In Vitro Using Adipose-derived Stem Cells.

To investigate cellular changes resulting in the fibrotic side-effects of RTx, cultured normal human dermal fibroblasts (NHDF) were irradiated (10Gy), then studied using functional assays that reflect key fibroblast functions, and compared with unirradiated controls. RNA-Seq and targeted microarrays (with specific examination of TGFβ) were performed to elucidate altered gene pathways. Finally, conditioned-media from ADSC was used to treat irradiated fibroblasts and model fat graft surgery. RTx altered NHDF morphology, with cellular functional changes reflecting transition into a more invasive phenotype: increased migration, adhesion, contractility, and disordered invasion. Changes in genes regulating collagen and MMP homeostasis and cell-cycle progression were also detected. However, TGFβ was not identified as a key intracellular regulator of the fibroblast response. Finally, treatment with ADSC-conditioned media reversed the RTx-induced hypermigratory state of NHDF. Our findings regarding cellular and molecular changes in irradiated fibroblasts help explain clinical manifestations of debilitating RTx-induced fibrosis. ADSC-secretome-mediated reversal indicated that these constituents may be used to combat the devastating side-effects of excessive unwanted fibrosis in RTx and other human fibrotic diseases.

Nectandra cuspidata fraction and the isolated polyphenols protect fibroblasts and hairless mice skin from UVB-induced inflammation and oxidative stress

Excessive exposure to UVB radiation can lead to oxidative and inflammatory damage that compromises the cutaneous integrity. The application on the skin of photochemoprotective products is considered a relevant approach for the prevention of oxidative damage. In this study the in vitro and in vivo photochemoprotective effects of antioxidant plant materials obtained from the leaves of Nectandra cuspidata Nees following UVB irradiation were evaluated. The cytoprotective effect, reactive oxygen species (ROS) production and lipid peroxidation (LPO) were assessed in L-929 fibroblasts treated with the ethyl acetate fraction (EAF) or isolated compounds (epicatechin, isovitexin and vitexin) before or after irradiation with UVB (500 mJ/cm2). EAF substantially reduced the dead of cells and inhibited the UVB-induced ROS production and LPO in both treatments, compared with the irradiated untreated fibroblasts, presenting effects similar or better than pure compounds. The in vivo photochemoprotective effects of a topical emulsion containing 1% EAF (F2) were evaluated in hairless mice exposed to UVB. F2 improved all evaluated parameters in the skin of animals, inhibited ROS production, increased antioxidant defenses by decreasing reduced glutathione (GSH) and catalase depletion, reduced the activities of metalloproteinases (MMP-2 and MMP-9) and myeloperoxidase, decreased epidermal thickness and skin edema, and inhibited the appearance of sunburn cells as well as the recruitment of neutrophils and mast cell inflammatory infiltrates. These findings show that EAF presents high photochemoprotective effects, and that a topical formulation containing it may have potential for skin care.

Abstract P6-03-14: Conditional reprogrammed cells enable us to examine the drug resistance for recurrent metastatic hormone receptor-positive, human epidermal receptor 2-negative breast cancer

Abstract Objectives: Various new drugs have been developed for treating recurrent hormone receptor-positive/human epidermal receptor 2-negative breast cancer. However, directly identifying effective drugs remains difficult. In this study, we elucidated the clinical relevance of cultured cells derived from patients with recurrent hormone receptor-positive/human epidermal receptor 2-negative metastatic breast cancer. Methods: We generated conditional reprogrammed cells from primary and metastatic breast cancer. CR cells were co-cultured with irradiated mouse fibroblast feeder cells in the presence of the Rho kinase inhibitor Y-27632. We performed microarray analysis using a SurePrint G3 Human GE microarray kit Ver. 2.0, and DNA target sequence using QIAseq Targeted DNA Panels to analyze the genetic variants of the 93 most commonly mutated genes in human breast cancer samples. We examined the pathological features by xenograft model. For drug screening assay, we used Cambridge Cancer Compound Library. Results: The results of microarray analysis, DNA target sequencing and xenograft experiments indicated that the mutation status and pathological features were preserved in CR cells, whereas RNA expression was different from that in the primary tumor cells, especially with respect to cell adhesion-associated pathways. The results of drug sensitivity assays involving the use of primary breast cancer CR cells were consistent with the result of Oncotype Dx. We performed drug screening assays using liver metastases, which were sensitive to 66 drugs. The result reflected the actual clinical course of this patient, resistant to taxanes and sensitive to fulvestrant plus palbociclib. Conclusion: These results supported the use of CR cells obtained from the metastatic lesions of patients with HR+/HER2− breast cancer for predicting the clinical drug efficacy. Citation Format: Rei Mimoto, Satomi Yogosawa, Atsushi Fushimi, Hiroko Nogi, Tadashi Asakura, Kiyotsugu Yoshida, Hiroshi Takeyama. Conditional reprogrammed cells enable us to examine the drug resistance for recurrent metastatic hormone receptor-positive, human epidermal receptor 2-negative breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-03-14.

IgE deficient patients have no IgE-encoding B cells in the periphery – a pilot study

While we learn more about different clinical presentations of IgE-deficient patients (IgE<2.5 kU/L), the mechanism resulting in IgE-deficiency is unknown. Through this small pilot study, we sought to investigate the frequency of IgE-encoding B cells in the peripheral blood of IgE-deficient individuals. Peripheral blood mononuclear cells (PBMCs) were collected from 5 IgE-deficient and 5 non-IgE deficient patients with different allergy-like symptoms. Cells were counted and assessed for viability before culturing in the presence of murine anti-human kappa/lambda, irradiated murine fibroblasts expressing human CD40-ligand, B-cell activating factor and IL-21. The cells were screened for IgE secretion after 6 days of incubation. All IgE-deficient patients had undetectable IgE levels (<2 kU/L). The median IgE level in non-IgE deficient patients was 1615 kU/L (range: 260-4340 kU/L). IgE-deficient patients had allergic rhinoconjunctivitis-like symptoms, two patients had associated asthma, and one drug allergy. The non-IgE deficient patients had asthma, food allergies and atopic dermatitis. The median PBMCs cultured was similar between IgE-deficient (16 cells x106, range: 8-16 cells x106) and non-IgE deficient individuals (24 cells x106, range: 8-24 cells x106, p=0.2). However, while the median frequency of IgE-encoding B cells was 2.9 cells/107 PBMCs (range: 2.5-4.8 cells/107 PBMCs) in non-IgE deficient patients, the IgE-deficient patients had zero IgE-encoding B cells. The abscence of IgE-encoding B cells in the serum of IgE-deficient patients suggests a defect in either these cells' survival, or in the IgE-class switching process. Further research is needed to elucidate the exact mechanism resulting in absence of serum IgE in certain individuals.

Ionizing radiation induces cutaneous lipid remolding and skin adipocytes confer protection against radiation-induced skin injury

BackgroundRadiation-induced skin injury is a serious concern during radiotherapy and radiation accidents. Skin fat represents the dominant architectural component of the human skin. However, the interplay between skin fat and the progression of radiation-induced skin injury remains largely unexplored. ObjectiveThis study aims to elucidate the interplay between skin fat and the progression of radiation-induced skin injury. MethodsSD rats were irradiated with an electron beam. mRNA profiles were determined by RNA-Seq. The skin lipid mass was monitored by magnetic resonance imaging (MRI) and lipid profiles were measured by liquid chromatography-mass spectrometry (LC–MS). Human mature adipocytes isolated from dermal and subcutaneous white adipose tissues (WATs) were co-cultured with human keratinocytes (HaCaT) and skin fibroblasts (WS1) in the transwell culture system. Cell migration ability was measured by migration assay. ResultsRadiation modulated cutaneous lipid metabolism by downregulating multiple pathways. Moreover, radiation decreased skin fat mass with altered lipid metabolite profiles. The rats fed with a high-fat diet showed resistance to radiogenic skin injury compared with that with a control diet, indicating that skin lipid plays a radioprotective role. Mature adipocytes promoted the migration but not the proliferation of co-cultured skin keratinocytes and fibroblasts. Palmitic acid, the most abundant fatty acid in skin tissues, facilitated the migration of WS1 cells. Moreover, fatty acid-binding protein 4 (FABP4) could be incorporated into skin cells and promote DNA damage repair in irradiated skin fibroblasts. ConclusionRadiation induces cutaneous lipid remolding, and skin adipocytes confer a protective role against radiation-induced skin injury.

Photochemoprotective effects of Spirulina platensis extract against UVB irradiated human skin fibroblasts

Ultraviolet B (UVB) irradiation is an important cause of a number of harmful side effects such as sunburn, photoaging, pigmentation and skin cancer. Photoprotection, which is used to prevent UVB interaction and to attenuate UVB related damage could be used to prevent the effects of UVB irradiation. Recently, the use of natural products as skin photoprotective agents has been in increasing demand. In this study, we are proposing a panel of in vitro methods for the evaluation of Spirulina platensis extract for its ability to absorb the UVB spectra by determination of its spectrophotometric Solar Protection Factor (SPF), and by attenuating its biological protective effects against UVB-irradiated human skin fibroblast cultures. Our data revealed that S. platensis extract contained high amounts of phenolic compounds (23,465.71 ± 167.72 mg GAE/kg extract) and produced a high degree of absorbance within a wavelength range of 290–320 nm yielding an SPF value of 30.39 ± 3.37 (3 mg/ml), all of which collectively confirmed the photoprotective activity of S. platensis polyphenols. S. platensis extract has no cytotoxicity on UVB irradiation of human skin fibroblasts, peripheral blood mononuclear cells and red blood cells. It was also found to exert antioxidant activities both in vitro and by way of intracellular reactive oxygen species in UVB irradiation fibroblast cells. The induction of interleukin- 6 and -8 secretions by UVB irradiation was dramatically aborted after S. platensis extract treatment. Moreover, the S. platensis extract significantly inhibited tyrosinase activity. Our above results indicated the photochemoprotective effects of S. platensis extract against UVB irradiation in human skin fibroblast cells. Therefore, S. platensis extract could be considered a prospective source of UVB-absorbers with antioxidant capacity. It could also serve as an additive in photoprotective cosmetic formulations which could prevent the harmful effects of UVB radiation.

Cytoprotective activity of carrot and tomato callus extracts and the ex‐ pression of cytokines in UV‐B irradiated fibroblast cells

Studies have suggested that both carrot (Daucus carota L.) and tomato (Solanum lycopersicum L.) callus extracts contain antoxidant compounds that might have the potental to protect cells from free radicals such as H2O2 that contribute to cell damage. The other sources of free radical exposure in human cells, such as UV‐B, should also be examined. UV‐B exposure can trigger increased expression of inflammatory cytokines such as cyclooxygenase‐2 (COX‐2) and tumor necrosis factor‐α (TNF‐α) and the antinflammatory cytokine interleukin‐10 (IL‐10), which causes photoaging. This study was conducted to investgate the cytoprotectve actvity of carrot and tomato callus aqueous extracts by observing cell viability using the MTT assay. Immunocytochemistry methods were used to examine the effects of carrot and tomato callus aqueousextracts on the expression of COX‐2, TNF‐α, and IL‐10 in human dermal fbroblast adult (HDFa) cells exposed to UV‐B light. Carrot and tomato callus aqueous extracts were obtained by the maceraton method using aqua bidistlled solvent. Results showed that both carrot and tomato callus aqueous extracts at 0.5 mg/mL exhibited the highest cytoprotectve effect in HDFa cells compared to that at other concentratons. Both carrot and tomato callus aqueous extracts could also decrease the expression of COX‐2 and TNF‐α, whereas carrot callus aqueous extract increased the expression of the ant‐inflammatory cytokine IL‐10 in HDFa cells.