Bone marrow embolism: should it result from traumatic bone lesions? A histopathological human autopsy study.

Bone marrow embolism (BME) is likely a consequence of fractures in which pulmonary vessels are the most affected. However, some cases of BME were reported in the absence of trauma. Thus, a traumatic injury might not be necessary for developing BME. This study discusses BME cases in patients without signs of fractures or blunt trauma. The discussion addresses various possible mechanisms for the appearance of BME. Options include cancer in which bone marrow metastasis is a suggestive cause. Another proposal is the chemical theory where bone marrow fats are released via lipoprotein lipase in a pro-inflammatory state, resulting in vascular/pulmonary obstruction. Other cases discussed in this study are hypovolemic shock and drug-abuse related BME. All autopsy cases with BME were included regardless of the cause of death for a period of 2 years. Autopsies involved complete dissection with the macroscopic evaluation of the affected organs, including the heart, lungs, and brain. Tissues were also prepared for microscopic examination. Of the 11 cases, eight showed non-traumatic BME (72%). These findings conflict with theories in the literature that BME most commonly occurs after fractures or trauma. One of the eight cases exhibited mucinous carcinoma; one is presented with hepatocellular carcinoma; and two cases showed severe congestion. Lastly, one case was found to be associated with each of the following conditions: liposuction, drug abuse, pulmonary hypertension, and heart failure. Each case suggests a different pathophysiology for developing BME, yet the exact mechanisms are not fully understood. Further study of non-traumatic associated BME is recommended.

Household disinfection practices by women living in Egypt during the 2020 COVID-19 lockdown and the association of information sources and suspected bleach toxicity.

INTRODUCTION: The spread of contradictory health information was a hallmark of the early COVID-19 pandemic. Because of a limited understanding of the disease, its mode of transmission, and its pathogenicity, the public turned to easily accessible and familiar sources of information. Some of these sources included wrong or incomplete information that could increase health risks and incidents of toxicity due to improper information about the usage of disinfectants. The objective of this study was to assess the relationship between sources of information about the COVID-19 pandemic, the related household cleaning and disinfection practices among adult women living in Egypt, and the associated adverse effects of bleach toxicity during a national lockdown. METHODS: Through a self-administered online survey, 452 adult women (18 years and older) living in Egypt were recruited from 13 cities between 4 June 2022 and 4 July 2022 to answer the questionnaire. The questionnaire included (41) questions in Arabic and collected data about respondents' household cleaning and disinfection practices to prevent the spread of the SARS-CoV-2 virus and protect their families during the lockdown that started in Egypt in March 2020. RESULTS: The study found that 88.1% (n = 398) of participants reported increased use of disinfectants during the lockdown. Women who chose social media as their primary source of information to learn about disinfection practices reported an increased frequency of respiratory symptoms associated with bleach toxicity (correlation coefficient = 0.10, p-value = 0.03), followed by women who depended on relatives and friends as the primary source of information (correlation coefficient = 0.10, p-value = 0.02). CONCLUSION: This study showed that social media is an easily accessible, efficient and fast communication tool that can act as a primary source for individuals seeking medical information compared to other media platforms (e.g., websites, T.V., satellite channels). However, better regulations and monitoring of its content may help limit the harms caused by the misinformation and disinformation spread by these popular platforms, particularly in times of uncertainty and upheaval.

Reduced microRNA-503 expression augments lung fibroblast VEGF production in chronic obstructive pulmonary disease.

Alterations in microRNA (miRNA) expression may contribute to COPD pathogenesis. In COPD, lung fibroblast repair functions are altered in multiple ways, including extracellular mediator release. Our prior study revealed miR-503 expression is decreased in COPD lung fibroblasts, although the exact role played by miR-503 is undetermined. The current study examined a role of miR-503 in cytokine, growth factor and fibronectin production by lung fibroblasts from patients with and without COPD. Primary adult lung fibroblasts were isolated from patients with or without COPD. MiR-503 expression and interleukin (IL)-6, -8, PGE2, HGF, KGF, VEGF and fibronectin release were examined with or without inflammatory cytokines, IL-1ß and tumor necrosis factor (TNF)-α. MiR-503 expression was decreased in COPD lung fibroblasts. The expression of miR-503 was positively correlated with %FVC, %FEV1, and %DLco as well as IL-6, -8, PGE2, HGF, KGF, and VEGF in the absence or presence of IL-1ß/TNF-α. In addition, IL-8 and VEGF release from COPD lung fibroblasts were increased compared to those from control. Exogenous miR-503 inhibited VEGF release from primary adult and fetal lung fibroblasts but not IL-8 release. As expected, COPD fibroblasts proliferated more slowly than control fibroblasts. MiR-503 did not affect proliferation of either control or COPD lung fibroblasts. MiR-503 inhibition of VEGF protein production and mRNA was mediated by direct binding to the 3' untranslated region of VEGF mRNA. Endogenous miR-503 was differently regulated by exogenous stimulants associated with COPD pathogenesis, including IL-1ß/TNF-α, TGF-ß1 and PGE2. Endogenous miR-503 inhibition augmented VEGF release by IL-1ß/TNF-α and TGF-ß1 but not by PGE2, demonstrating selectivity of miR-503 regulation of VEGF. In conclusions, reduced miR-503 augments VEGF release from lung fibroblasts from patients with COPD. Since VEGF contributes to disturbed vasculature in COPD, altered miR-503 production might play a role in modulating fibroblast-mediated vascular homeostasis in COPD.

Effect of culture conditions on microRNA expression in primary adult control and COPD lung fibroblasts in vitro.

In vitro cell cultures, including lung fibroblasts, have been used to identify microRNAs (miRNAs) associated with chronic obstructive pulmonary disease (COPD) pathogenesis. However, culture conditions may affect miRNA expression. We examined whether miRNA expression in primary adult lung fibroblasts varies with cell density or passage in vitro and whether culture conditions confound the identification of altered miRNA expression in COPD lung fibroblasts. Primary adult control and COPD lung fibroblasts were cultured until passage 3 or 8, after which cells were further cultured for 3 or 7 d (low vs. high density). Then, cells at low density were cultured with serum-free media, and those at high density were cultured with serum-free media in the absence or presence of interleukin-1ß (IL-1ß) and tumor necrosis factor alpha (TNF-α) for 24 h. RNA was extracted to perform miRNA microarray from which 1.25-fold differential expression and 10% false discovery rate were applied to identify "invariant" and "variant" miRNA for the various culture conditions. Of the 2226 miRNAs evaluated, 39.0% for cell density, 40.7% for cell passage, and 29.4% for both conditions were identified as "invariant" miRNAs. Furthermore, 38.1% of the evaluated miRNAs were "invariant" for cell passage with IL-1ß and TNF-α. Differentially expressed miRNAs between control and COPD lung fibroblasts were identified with and without IL-1ß and TNF-α, and of these, 32 out of the 34 top-ranked miRNAs exceeded the differences due to culture conditions. Thus, culture conditions may affect miRNA expression of adult human lung fibroblasts. Nevertheless, in vitro cultures can be used to assess differential miRNA expression in COPD lung fibroblasts.

Prostaglandin E2 switches from a stimulator to an inhibitor of cell migration after epithelial-to-mesenchymal transition.

Epithelial-mesenchymal transition (EMT) is critical for embryonic development, and this process is recapitulated in adults during wound healing, tissue regeneration, fibrosis and cancer progression. Cell migration is believed to play a key role in both normal wound repair and in abnormal tissue remodeling. Prostaglandin E2 (PGE2) inhibits fibroblast chemotaxis, but stimulates chemotaxis in airway epithelial cells. The current study was designed to explore the role of PGE2 and its four receptors on airway epithelial cell migration following EMT using both the Boyden blindwell chamber chemotaxis assay and the wound closure assay. EMT in human bronchial epithelial cells (HBECs) was induced by TGF-ß1 and a mixture of cytokines (IL-1ß, TNF-α, and IFN-γ). PGE2 and selective agonists for all four EP receptors stimulated chemotaxis and wound closure in HBECs. Following EMT, the EP1 and EP3 agonists were without effect, while the EP2 and EP4 agonists inhibited chemotaxis as did PGE2. The effects of the EP2 and EP4 receptors on HBEC and EMT cell migration were further confirmed by blocking the expected signaling pathways. Taken together, these results demonstrate that PGE2 switches from a stimulator to an inhibitor of cell migration following EMT of airway epithelial cells and that this inhibition is mediated by an altered effect of EP2 and EP4 signaling and an apparent loss of the stimulatory effects of EP1 and EP3. Change in the PGE2 modulation of chemotaxis may play a role in repair following injury.

Fibroblasts that resist cigarette smoke-induced senescence acquire profibrotic phenotypes.

This study assessed the effect of extended exposure to cigarette smoke extract (CSE) on tissue repair functions in lung fibroblasts. Human fetal (HFL-1) and adult lung fibroblasts were exposed to CSE for 14 days. Senescence-associated ß-galactosidase (SA ß-gal) expression, cell proliferation, and tissue repair functions including chemotaxis and gel contraction were assessed. HFL-1 proliferation was inhibited by CSE and nearly half of the CSE-exposed cells were SA ß-gal positive after 14 days exposure, whereas 33% of adult lung fibroblasts were SA ß-gal positive in response to 10% CSE exposure. The SA ß-gal-positive cells did not proliferate as indicated by bromodeoxyuridine incorporation. In contrast, cells negative for SA ß-gal after CSE exposure proliferated faster than cells never exposed to CSE. These nonsenescent cells migrated more and contracted collagen gels more than control cells. CSE exposure stimulated TGF-ß1 production, and both inhibition of TGF-ß receptor kinase and TGF-ß1 siRNA blocked CSE modulation of fibroblast function. Extended exposure to CSE might induce two different fibroblast phenotypes, a senescent and a profibrotic phenotype. The fibroblasts that resist CSE-induced cellular senescence may contribute to the pathogenesis of idiopathic pulmonary fibrosis and could contribute to fibrotic lesions in chronic obstructive pulmonary disease acting through a TGF-ß1-mediated pathway. In contrast, the senescent cells may contribute to the pathogenesis of emphysema.

Reprogramming of COPD lung fibroblasts through formation of induced pluripotent stem cells.

Reprogramming somatic cells to induced pluripotent stem cells (iPSCs) eliminates many epigenetic modifications that characterize differentiated cells. In this study, we tested whether functional differences between chronic obstructive pulmonary disease (COPD) and non-COPD fibroblasts could be reduced utilizing this approach. Primary fibroblasts from non-COPD and COPD patients were reprogrammed to iPSCs. Reprogrammed iPSCs were positive for oct3/4, nanog, and sox2, formed embryoid bodies in vitro, and induced teratomas in nonobese diabetic/severe combined immunodeficient mice. Reprogrammed iPSCs were then differentiated into fibroblasts (non-COPD-i and COPD-i) and were assessed either functionally by chemotaxis and gel contraction or for gene expression by microarrays and compared with their corresponding primary fibroblasts. Primary COPD fibroblasts contracted three-dimensional collagen gels and migrated toward fibronectin less robustly than non-COPD fibroblasts. In contrast, redifferentiated fibroblasts from iPSCs derived from the non-COPD and COPD fibroblasts were similar in response in both functional assays. Microarray analysis identified 1,881 genes that were differentially expressed between primary COPD and non-COPD fibroblasts, with 605 genes differing by more than twofold. After redifferentiation, 112 genes were differentially expressed between COPD-i and non-COPD-i with only three genes by more than twofold. Similar findings were observed with microRNA (miRNA) expression: 56 miRNAs were differentially expressed between non-COPD and COPD primary cells; after redifferentiation, only 3 miRNAs were differentially expressed between non-COPD-i and COPD-i fibroblasts. Interestingly, of the 605 genes that were differentially expressed between COPD and non-COPD fibroblasts, 293 genes were changed toward control after redifferentiation. In conclusion, functional and epigenetic alterations of COPD fibroblasts can be reprogrammed through formation of iPSCs.

Vitamin D modulates prostaglandin E2 synthesis and degradation in human lung fibroblasts.

Vitamin D insufficiency has been increasingly recognized in the general population worldwide and has been associated with several lung diseases, including asthma, chronic obstructive pulmonary disease (COPD), and respiratory tract infections. Fibroblasts play a critical role in tissue repair and remodeling, which is a key feature of COPD and asthma. Fibroblasts modulate tissue repair by producing and modifying extracellular matrix components and by releasing mediators that act as autocrine or paracrine modulators of tissue remodeling. The current study was designed to investigate if vitamin D alters fibroblast release of key autocrine/paracrine repair factors. First, we demonstrated that human fetal lung (HFL)-1 cells express the vitamin D receptor (VDR) and that vitamin D, 25-hydroxyvitamin D [25(OH)D], or 1,25-dihydroxyvitamin D [1,25(OH)2D] induce VDR nuclear translocation and increase VDR-DNA binding activity. We next demonstrated that vitamin D, 25(OH)D, and 1,25(OH)2D significantly reduced prostaglandin (PG)E2 production by human lung fibroblasts (HFL-1) but had no effect on transforming growth factor ß1, vascular endothelial growth factor, or fibronectin production. Vitamin D, 25(OH)D, and 1,25(OH)2D significantly inhibited IL-1ß-induced microsomal PGE synthase (mPGES)-1 expression; in contrast, all three forms of vitamin D stimulated 15-hydroxy PG dehydrogenase, an enzyme that degrades PGE2. Cyclooxygenase-1 and -2 and the other two PGE2 synthases (mPGES-2 and cytosolic PGE synthase) were not altered by vitamin D, 25(OH)D, or 1,25(OH)2D. Finally, the effect of PGE2 inhibition by 25(OH)D was observed in adult lung fibroblasts. These findings suggest that vitamin D can regulate PGE2 synthesis and degradation and by this mechanism can modulate fibroblast-mediated tissue repair function.

Anti-inflammatory effects of budesonide in human lung fibroblast are independent of histone deacetylase 2.

OBJECTIVE AND DESIGN: Reduced expression of histone deacetylase 2 (HDAC2) in alveolar macrophages and epithelial cells may account for reduced response of chronic obstructive pulmonary disease (COPD) patients to glucocorticoids. HDAC2 expression and its role in mediating glucocorticoid effects on fibroblast functions, however, has not been fully studied. This study was designed to investigate whether HDAC2 mediates glucocorticoid effects on release of inflammatory cytokines and matrix metalloproteinases (MMPs) from human lung fibroblasts. METHODS: Human lung fibroblasts (HFL-1 cells) were stimulated with interleukin (IL)-1 ß plus tumor necrosis factor (TNF)-α in the presence or absence of the glucocorticoid budesonide. Cytokines (IL-6 and IL-8) were quantified by enzyme linked immunosorbent assay (ELISA) and MMPs (MMP-1 and MMP-3) by immunoblotting in culture medium. The role of HDAC2 was investigated using a pharmacologic inhibitor as well as a small interfering ribonucleic acid (siRNA) targeting HDAC2. RESULTS: We have demonstrated that budesonide concentration-dependently (10(-10)-10(-7) M) inhibited IL-6, IL-8, MMP-1, and MMP-3 release by HFL-1 cells in response to IL-1ß plus TNF-α. While an HDAC inhibitor significantly blocked the inhibitory effect of budesonide on human bronchial epithelial cells (HBECs) and monocytes (THP-1 cells), it did not block the inhibitory effect of budesonide on release of cytokines and MMPs from HFL-1 cells. Similarly, an HDAC2-siRNA blocked budesonide inhibition of cytokine release in HBECs, but it did not block the inhibitory effect of budesonide on HFL-1 cytokine and MMP release. Furthermore, budesonide significantly blocked release of cytokines and MMPs to a similar degree in normal and COPD lung fibroblasts as well as in HFL-1 cells exposed or not exposed to cigarette smoke extract. CONCLUSION: These findings suggest that, in contrast to airway epithelial cells and monocytes/macrophages, HDAC2 is not required for budesonide to inhibit MMP and cytokine release by lung fibroblasts and this inhibitory pathway appears to be intact in cultured fibroblasts from COPD patients. These results also suggest that budesonide has the potential to modulate fibroblast-mediated tissue remodeling following airway inflammation in COPD, which is mediated via an HDAC2 independent pathway.

Phosphodiesterase-4 inhibition augments human lung fibroblast vascular endothelial growth factor production induced by prostaglandin E2.

Lung fibroblasts are believed to be a major source of vascular endothelial growth factor (VEGF), which supports the survival of lung endothelial cells and modulates the maintenance of the pulmonary microvasculature. VEGF has been related to the pathogenesis of lung diseases, including chronic obstructive pulmonary disease (COPD). Prostaglandin E2 (PGE2) stimulates VEGF production from lung fibroblasts via the E-prostanoid (EP)-2 receptor. The EP2 signaling pathway uses cyclic adenosine monophosphate (cAMP) as a second messenger, and cAMP is degraded by phosphodiesterases (PDEs). This study investigates whether phosphodiesterase inhibition modulates the human lung fibroblast VEGF production induced by PGE2. Human fetal lung fibroblasts were cultured with PGE2 and PDE inhibitors. The PDE4 inhibitors roflumilast, roflumilast N-oxide, and rolipram with PGE2 increased VEGF release, as quantified in supernatant media by ELISA. In contrast, PDE3, PDE5, and PDE7 inhibitors did not affect VEGF release. Roflumilast increased VEGF release with either an EP2 or an EP4 agonist. Roflumilast augmented the cytosolic cAMP levels induced by PGE2 and VEGF release with other agents that use the cAMP signaling pathway. Roflumilast-augmented VEGF release was completely inhibited by a protein kinase A (PKA) inhibitor. Roflumilast with PGE2 increased VEGF mRNA levels, and the blockade of mRNA synthesis inhibited the augmented VEGF release. The stimulatory effect of roflumilast on VEGF release was replicated using primary healthy and COPD lung fibroblasts. These findings demonstrate that PDE4 inhibition can modulate human lung fibroblast VEGF release by PGE2 acting through the EP2 and EP4 receptor-cAMP/PKA signaling pathway. Through this action, PDE4 inhibitors such as roflumilast could contribute to the survival of lung endothelial cells.

Smad3 mediates cigarette smoke extract (CSE) induction of VEGF release by human fetal lung fibroblasts.

Cigarette smoke is the major cause of chronic obstructive pulmonary disease (COPD), yet pathogenic mechanisms are not fully understood. Vascular endothelial growth factor (VEGF) is one of the major regulators of endothelial cell survival and is believed to play a role in the pathogenesis of COPD. Fibroblasts are a significant source of VEGF in the lungs; however the effect of cigarette smoke exposure on VEGF release by fibroblasts is not fully understood. We hypothesized that cigarette smoke-induced disturbed VEGF release by human lung fibroblasts is a potential pathogenic mechanism that could contribute to COPD. Cigarette smoke extract (CSE) was prepared by modification of the methods of Carp and Janoff (American Review of Respiratory Disease, 1978). Human fetal lung fibroblasts (HFL-1) were exposed to different concentrations of CSE and for different durations. VEGF release into the media was measured using ELISA. TGF-ß1 receptor (TßR1)/Smad3 as a potential pathway for CSE modulated VEGF release was also investigated using biochemical analyses and siRNA inhibition of Smad3 and siRNA and pharmacologic inhibition of TßR1. CSE induced VEGF release by HFL-1 in concentration and time dependent manner. This was confirmed in two additional types of primary human fetal lung fibroblasts. CSE induced Smad3 phosphorylation and nuclear translocation in HFL-1 cells. Silencing of Smad3 by siRNA not only eliminated the stimulatory effect of CSE on VEGF release but also inhibited baseline VEGF production. Suppression of TßR1 by the pharmacological inhibitor (SB431542) markedly reduced VEGF release by HFL-1 in response to CSE and this effect was confirmed by TßR1 siRNA. In contrast, nicotine inhibited VEGF release by HFL-1 in a dose and time dependent manner. Our findings indicate that CSE stimulates Smad3-mediated VEGF release by lung fibroblasts. Nicotine does not account for the CSE stimulation of VEGF in HFL-1. The ability of lung fibroblasts to produce VEGF may play a role in pathogenesis of cigarette smoke induced lung disease.

Differential roles of JNK, ERK1/2, and p38 mitogen-activated protein kinases on endothelial cell tissue repair functions in response to tumor necrosis factor-α.

Tumor necrosis factor (TNF)-α can alter tissue repair functions in a variety of cells including endothelial cells. However, the mechanism by which TNF-α mediates these functional changes has not fully been studied. We investigated the role of mitogen-activated protein kinases (MAPKs) on mediating the regulatory effect of TNF-α on the tissue repair functions of human pulmonary artery endothelial cells (HPAECs). TNF-α protected HPAECs from undergoing apoptosis induced by serum and growth factor deprivation, augmented collagen gel contraction, and stimulated wound closure. TNF-α activated c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinases 1 and 2 (ERK1/2), and p38. Inhibitors of JNK (SP600125, 5 µM) or ERK1/2 (PD98059, 5 µM) significantly inhibited TNF-α-stimulated cell survival, contraction of collagen gels, and wound closure. In contrast, the p38 inhibitor SB203580 (5 µM) further amplified all of the TNF-α effects on HPAECs. TNF-α specifically activated p38α but not other p38 isoforms and suppression of p38α by an siRNA resulted in further amplification of the TNF-α effect. These results suggest that TNF-α stimulates tissue repair functions of HPAECs, and this may be mediated, at least in part, positively via JNK and ERK1/2, and negatively through p38α. MAPKs may play a role in endothelial cell-mediated tissue repair, especially in an inflammatory milieu where TNF-α is present.

Prostaglandin E2 stimulates the production of vascular endothelial growth factor through the E-prostanoid-2 receptor in cultured human lung fibroblasts.

Fibroblasts are the major mesenchymal cells present within the interstitium of the lung and are a major source of vascular endothelial growth factor (VEGF), which modulates the maintenance of pulmonary microvasculature. Prostaglandin E(2) (PGE(2)) acts on a set of E-prostanoid (EP) receptors that activate multiple signal transduction pathways leading to downstream responses. We investigated the modulation by PGE(2) of VEGF release by human lung fibroblasts. Human lung fibroblasts were cultured until reaching 90% confluence in tissue culture plates, after which the culture media were changed to serum-free Dulbecco's modified Eagle's medium, with or without PGE(2), and with specific agonists or antagonists for each EP receptor. After 2 days, culture media were assayed for VEGF by ELISA. The results demonstrated that PGE(2) and the EP2 agonist ONO-AE1-259-01 significantly stimulated the release of VEGF in a concentration-dependent manner. Agonists for other EP receptors did not stimulate the release of VEGF. The stimulatory effect of PGE(2) was blocked by the EP2 antagonist AH6809, but was not blocked by antagonists for other EP receptors. The protein kinase-A (PKA) inhibitor KT-5720 also blocked the stimulatory effect of PGE(2). The increased release of VEGF induced by PGE(2) was accompanied by a transient increase in the concentration of VEGF mRNA. These findings demonstrate that PGE(2) can modulate the release of VEGF by human lung fibroblasts through its actions in the EP2 receptor/PKA pathway. This activity may contribute to the maintenance of pulmonary microvasculature in the alveolar wall.

The p38 mitogen-activated protein kinases modulate endothelial cell survival and tissue repair.

OBJECTIVE AND DESIGN: This study is designed to investigate the role of p38 MAPK in modulating human pulmonary artery endothelial cells (HPAECs) survival and tissue repair functions. METHODS: HPAECs (passage 8-12) were used for all experiments. Cells were treated with IL-1ß (0.5 or 2 ng/ml) or p38 inhibitor (SB203580 or SB220025, 5 µM each). Cells were also transfected with 50 nM siRNAs. Cell length was measured using ImageJ software. Collagen gel contraction and wound close assay were performed to evaluate tissue repair functions. RESULTS: IL-1ß activated p38 MAPK and induced morphologic change of HPAECs. The p38 inhibitors further augmented IL-1ß-induced cell morphologic change, prevented cell death, and augmented collagen gel contraction. Suppression of p38α, γ, or δ, but not p38ß resulted in cell morphologic alteration, and suppressing any one of p38 isoforms by siRNAs increased cell survival. Suppression of p38α or δ augmented gel contraction. While p38α suppression stimulated cell migration, suppressing the rest of three isoforms inhibit cell migration. Nuclear factor p65-siRNA blocked IL-1ß-induced cell morphologic change, but did not affect p38 inhibitor-induced change. CONCLUSION: These findings suggest that p38 MAPK may negatively modulate tissue repair functions of endothelial cells via p65 independent pathway.

Inflammatory cytokines regulate endothelial cell survival and tissue repair functions via NF-κB signaling.

Inflammation contributes to the development of fibrotic and malignant diseases. We assessed the ability of inflammatory cytokines to modulate endothelial cell survival and functions related to tissue repair/remodeling. Treatment with interleukin (IL)-1ß or tumor necrosis factor (TNF)-α (2 ng/mL) led to human pulmonary artery endothelial cells becoming spindle-shaped fibroblast-like cells. However, immunoblot and DNA microarray showed no change in most endothelial and mesenchymal markers. In the presence of IL-1ß or TNF-α, cells were resistant to apoptosis induced by deprivation of serum and growth factor, and were more migratory. In addition, cells treated with IL-1ß or TNF-α contracted collagen gels more robustly. In contrast, transforming growth factor-ß1 did not induce these responses. RNA interference targeting nuclear factor (NF)-κB p65 blocked the effects of IL-1ß or TNF-α on cell morphologic change, survival, migration, and collagen gel contraction. These results suggest that endothelial cells may contribute to tissue repair/remodeling via the NF-κB signaling in a milieu of airway inflammation.

IL-4 induces differentiation of human embryonic stem cells into fibrogenic fibroblast-like cells.

BACKGROUND: Fibroblast heterogeneity is recognized, and fibroblasts from diseased tissues, including those of asthmatic subjects, have functional phenotypes that differ from normal tissue. However, progenitor-progeny relationships and the factors that control fibroblast differentiation are poorly defined. OBJECTIVE: We sought to determine whether IL-4 could alter the functional phenotype of fibroblasts during their differentiation from stem/progenitor cells. METHODS: Using a 3-dimensional collagen gel system, we obtained embryoid bodies derived from human embryonic stem cells and recovered spindle-shaped cells consistent with fibroblasts that had differentiated in the presence or absence of IL-4. RESULTS: IL-4-induced fibroblast-like cells were more active in contraction of collagen gels, migration, and production of fibronectin than control (without IL-4) cells. IL-4-induced cells demonstrated less expression of miR-155, which modulated contraction, migration, and fibronectin production. These differences persisted in culture without further addition of IL-4, suggesting the differentiated phenotype might be a permanent alteration. CONCLUSION: The current study demonstrates that IL-4 induces differentiation of stem/precursor cells into fibroblast-like cells that demonstrate a more fibrogenic phenotype, which is due to reduced expression of miR-155. These findings provide a novel mechanism for the persistent abnormalities in IL-4-related diseases and a novel target to regulate tissue remodeling by fibroblasts.

Prostaglandin E2 inhibits human lung fibroblast chemotaxis through disparate actions on different E-prostanoid receptors.

The migration of fibroblasts is believed to play a key role in both normal wound repair and abnormal tissue remodeling. Prostaglandin E (PGE)(2), a mediator that can inhibit many fibroblast functions including chemotaxis, was reported to be mediated by the E-prostanoid (EP) receptor EP2. PGE(2), however, can act on four receptors. This study was designed to determine if EP receptors, in addition to EP2, can modulate fibroblast chemotaxis. Using human fetal lung fibroblasts, the expression of all four EP receptors was demonstrated by Western blotting. EP2-selective and EP4-selective agonists inhibited both chemotaxis toward fibronectin in the blindwell assay and migration in a wound-closure assay. In contrast, EP1-selective and EP3-selective agonists stimulated cell migration in both assay systems. These results were confirmed using EP-selective antagonists. The role of both EP2 and EP4 receptors in mediating the PGE(2) inhibition of chemotaxis was also confirmed by small interfering RNA suppression. Furthermore, the role of EP receptors was confirmed by blocking the expected signaling pathways. Taken together, these results demonstrate that PGE(2) can act on multiple EP receptors in human lung fibroblasts, to exert disparate effects. Alterations in EP receptor expression may have the potential to alter PGE(2) action. Targeting specific EP receptors may offer therapeutic opportunities in conditions characterized by abnormal tissue repair and remodeling.