Pigment Epithelium-derived Factor Overexpression Research Articles

Pigment epithelium‑derived factor inhibits proliferation, invasion and angiogenesis, and induces ferroptosis of extravillous trophoblasts by targeting Wnt‑β‑catenin/VEGF signaling in placenta accreta spectrum.

Placenta accreta spectrum (PAS) is one of the most dangerous complications in obstetrics, which can lead to severe postpartum bleeding and shock, and even necessitate uterine removal. The abnormal migration and invasion of extravillous trophoblast cells (EVTs) and enhanced neovascularization occurring in an uncontrolled manner in time and space are closely related to the abnormal expression of pro‑angiogenic and anti‑angiogenic factors. The pigment epithelium‑derived factor (PEDF) is a multifunctional regulatory factor that participates in several important biological processes and is recognized as the most efficient inhibitor of angiogenesis. The present study aimed to explore the effects of PEDF on EVT phenotypes and the underlying mechanisms in PAS. HTR‑8/SVneo cells were transfected to overexpress or knock down PEDF. Cell proliferation and invasion were assessed using Cell Counting Kit‑8, 5‑ethynyl‑2'‑deoxyuridine and Transwell assays. In vitro angiogenesis was analyzed using tube formation assays. The degree of ferroptosis was assessed by evaluating the levels of lipid reactive oxygen species, total iron, Fe2+, malondialdehyde and reduced glutathione using commercial kits. The expression levels of biomarkers of ferroptosis, angiogenesis, cell proliferation and Wnt signaling were examined by western blotting. PEDF overexpression decreased the proliferation, invasion and angiogenesis, and induced ferroptosis of EVTs. Activation of Wnt signaling with BML‑284 and overexpression of vascular endothelial growth factor (VEGF) reversed the PEDF overexpression‑induced suppression of cell proliferation, invasion and tube formation. PEDF overexpression‑induced ferroptosis was also decreased by Wnt agonist treatment and VEGF overexpression. It was predicted that PEDF suppressed the proliferation, invasion and angiogenesis, and increased ferroptosis in EVTs by decreasing Wnt‑β‑catenin/VEGF signaling. The findings of the present study suggested a novel regulatory mechanism of the phenotypes of EVTs and PAS.

Overexpressed pigment epithelium-derived factor alleviates pulmonary hypertension in two rat models induced by monocrotaline and SU5416/hypoxia

BackgroundPulmonary hypertension (PH) is a progressive and fatal cardiopulmonary disease characterized by vascular remodeling and is associated with endothelial-to-mesenchymal transition (EndoMT). The pigment epithelium-derived factor (PEDF), a secretory protein widely distributed in multiple organs, has been shown to demonstrate anti-EndoMT activity in cardiovascular diseases. In the present study, the role of PEDF in PH was investigated. MethodsFor PEDF overexpression, Sprague Dawley rats were infected with an adeno-associated virus through injection via the internal jugular vein. To establish PH models, the animals were subjected to monocrotaline or Sugen/hypoxia. Four weeks later, pulmonary artery angiography was performed, and hemodynamic parameters, right ventricular function, and vascular remodeling were evaluated. EndoMT and cell proliferation in the pulmonary arteries were assessed via immunofluorescence staining. Moreover, pulmonary artery endothelial cells (PAECs) isolated from experimental PH rats were cultured to investigate the underlying molecular mechanisms involved. ResultsPEDF expression was significantly downregulated in PAECs from PH patients and PH model rats. Overexpressed PEDF alleviated the development of PH by improving pulmonary artery morphology and perfusion, reducing pulmonary artery pressure, improving right ventricular function, and alleviating vascular remodeling. PEDF inhibits EndoMT and reduces excessive PAEC proliferation. Moreover, PEDF overexpression reduced EndoMT in cultured PAECs by competitively inhibiting the binding of wnt to LRP6 and downregulating phosphorylation at the 1490 site of LRP6. ConclusionsOur findings suggest that PEDF may be a potential therapeutic target for PH. We also found that PEDF can inhibit EndoMT in PAECs and may exert these effects by inhibiting the Wnt/LRP6/β-catenin pathway.

PEDF protects retinal pigment epithelium from ferroptosis and ameliorates dry AMD-like pathology in a murine model.

Age-related macular degeneration (AMD) is the leading cause of irreversible vision damage among elderly individuals. There is still no efficient treatment for dry AMD. Retinal pigment epithelial (RPE) degeneration has been confirmed to play an important role in dry AMD. Recent studies have reported that ferroptosis caused by iron overload and lipidperoxidation may be the primary causes of RPE degeneration. However, the upstream regulatory molecules of RPE ferroptosis remain largely unknown. Pigment epithelium-derived factor (PEDF) is an important endogenic protective factor for the RPE. Our results showed that in the murine dry AMD model induced by sodium iodate (SI), PEDF expression was downregulated. Moreover, dry AMD-like pathology was observed in PEDF-knockout mice. Therefore, the aim of this study was to reveal the effects and mechanism of PEDF on RPE ferroptosis and investigate potential therapeutic targets for dry AMD. The results of lipid peroxidation and transmission electron microscope showed that retinal ferroptosiswas significantly activated in SI-treated mice and PEDF-knockout mice. Restoration of PEDF expression ameliorated SI-induced retinal dysfunction in mice, as assessed by electroretinography and optical coherence tomography. Mechanistically, western blotting and immunofluorescence analysis demonstrated that the overexpression of PEDF could upregulate the expression of glutathione peroxidase 4 (GPX4) and ferritin heavy chain-1 (FTH1), which proved to inhibit lipidperoxidation and RPE ferroptosis induced by SI. This study revealed the novel role of PEDF in ferroptosis inhibition and indicated that PEDF might be a potential therapeutic target for dry AMD.

Tyrosine-Mutant AAV8 Vector Mediated Efficient and Safe Gene Transfer of Pigment Epithelium-Derived Factor to Mouse Lungs.

Recombinant adeno-associated viruses (rAAV) are an important tool for lung targeted gene therapy. Substitution of tyrosine with phenylalanine residues (Y-F) in the capsid have been shown to protect the AAV vector from ubiquitin/proteasome degradation, increasing transduction efficiency. We tested the mutant Y733F-AAV8 vector for mucus diffusion, as well as the safety and efficacy of pigment epithelium-derived factor (PEDF) gene transfer to the lung. For this purpose, Y733F-AAV8-PEDF (1010 viral genome) was administered intratracheally to C57BL/6 mice. Lung mechanics, morphometry, and inflammation were evaluated 7, 14, 21, and 28 days after injection. The tyrosine-mutant AAV8 vector was efficient at penetrating mucus in ex vivo assays and at transferring the gene to lung cells after in vivo instillation. Increased levels of transgene mRNA were observed 28 days after vector administration. Overexpression of PEDF did not affect in vivo lung parameters. These findings provide a basis for further development of Y733F-AAV8-based gene therapies for safe and effective delivery of PEDF, which has anti-angiogenic, anti-inflammatory and anti-fibrotic activities and might be a promising therapy for lung inflammatory disorders.

Knockdown of DAPK1 inhibits IL-1β-induced inflammation and cartilage degradation in human chondrocytes by modulating the PEDF-mediated NF-kB and NLRP3 inflammasome pathway.

Osteoarthritis (OA) is a common joint disease that is characterized by inflammation and cartilage degradation. Death-associated protein kinase 1 (DAPK1) is a multi-domain serine/threonine kinase and has been reported to be involved in the progression of OA. However, its role and mechanism in OA remain unclear. Here, we found the expression of DAPK1 in OA cartilage tissues was higher than that in normal cartilage tissues. The expression of DAPK1 in chondrocytes was up-regulated by IL-1β. Knockdown of DAPK1 promoted cell viability and anti-apoptotic protein expression, while it inhibited the apoptosis rate and pro-apoptotic protein expressions in IL-1β-induced chondrocytes. In addition, DAPK1 inhibition reduced the levels of inflammatory cytokines and expressions of matrix metalloproteinases (MMPs), and increased the expressions of collagen II and aggrecan. The data of mechanistic investigation indicated that the expression of pigment epithelium-derived factor (PEDF) was positively regulated by DAPK1. Overexpression of PEDF attenuated the effects of DAPK1 knockdown on IL-1β-induced cell viability, apoptosis, inflammation, and cartilage degradation. Furthermore, PEDF overexpression restored the activity of the NF-κB pathway and NLRP3 inflammasome after DAPK1 knockdown. Collectively, down-regulation of DAPK1 inhibited IL-1β-induced inflammation and cartilage degradation via the PEDF-mediated NF-κB and NLRP3 inflammasome pathways.

Overexpression of pigment epithelium-derived factor in breast cancer cell-derived exosomes induces M1 polarization in macrophages

M2 macrophages, the major component of tumor microenvironment, are recognized as important player in tumor progression. M2 macrophages mediate this effect by promoting tumor angiogenesis, tumor metastasis, and suppression of tumor immunity. Reprogramming of M2 macrophages can serve as a promising strategy in cancer immunotherapy. In this study, we constructed pigment epithelium-derived factor (PEDF) expressing vector and transfected MDA-MB-231 cells with this construct. Then, exosomes were isolated from transfected cells and M2 macrophages were incubated with isolated exosomes from transfected cell. The effect of isolated exosomes on macrophage polarization was examined by real-time PCR and ELISA. The results demonstrated reprogramming of M2 macrophages after incubation with isolated exosomes from PEDF transfected cells. M2-to-M1 repolarization of macrophages was confirmed by upregulation of CD80, IRF5, MCP1, and IL-1β and repression of CD206, Arg, TGM2, and TGF-β. Therefore, these findings revealed that introducing PEDF into exosomes by genetic manipulation of parent cells may be a potential approach for reprogramming of M2 macrophages in cancer.

PEDF is an antifibrosis factor that inhibits the activation of fibroblasts in a bleomycin-induced pulmonary fibrosis rat model

BackgroundIdiopathic pulmonary fibrosis (IPF) is a highly heterogeneous and fatal lung disease. In addition to dense fibrous tissue, abnormal angiogenesis is also an important feature of IPF. Pigment epithelium-derived factor (PEDF) is an angiogenesis inhibitor and a potential anti-fibrous factor. The purpose of this experiment is to observe the effect of PEDF on bleomycin (BLM)-induced pulmonary fibrosis in rats.MethodsIn vivo, pathological examination and detection of related factors were performed on pulmonary fibrosis induced by BLM in rats, and the temporal and spatial distribution of PEDF was investigated. Furthermore, lung gene delivery (PEDF-adeno-associated virus) was performed to investigate the effect of PEDF on pulmonary fibrosis. In vitro, lentiviral vectors were used to construct PEDF over-expression or knock out primary rat lung (PRL) fibroblasts. The effect of PEDF on fibroblast activation under TGF-β1 stimulation was evaluated, and the activation of TGF-β1/smad pathway and PPAR-γ expression (in the presence or absence of PPAR-γ inhibitors) were analyzed.ResultsIn vivo results showed that PEDF expression decreased during the inflammatory phase and increased during the fibrotic phase. PEDF could inhibit the progression of pulmonary fibrosis in rats. In vitro results showed that PEDF could effectively inhibit TGF-β1-stimulated fibroblast activation and reduce the production of α-SMA and collagen-I. PEDF could inhibit the TGF-β1/smad pathway by up-regulating the activity of PPAR-γ.ConclusionsPEDF can act as an anti-fibrotic factor, inhibit fibroblast activation by upregulating PPAR-γ activity and reduce BLM-induced pulmonary fibrosis in rats.

Pigment epithelium-derived factor, an anti-VEGF factor, delays ovarian cancer progression by alleviating polarization of tumor-associated macrophages.

Ovarian cancer (OC) is one of the most dangerous gynecological malignancies with no effective treatment so far. Pigment epithelium-derived factor (PEDF) has been reported to have ideal anti-tumor effects, but its relationship with the regulation of tumor-associated macrophage polarization is currently unclear. In this study, the mRNA expression of PEDF and macrophage markers were determined in OC tissues from clinic patients and five OC (A2780, SKOV3, CAOV3, OVCAR3, and OVCA433) cell lines through quantitative reverse transcription PCR. Afterwards, tumor growth, cell proliferation and apoptosis, and macrophage polarization in OC tumor-bearing mice with PEDF overexpression were recorded and investigated. Finally, the polarization of macrophages was explored in the presence of lentiviral PEDF overexpression, adipose triglyceride lipase (ATGL) and laminin receptor (LR) knockdown, and mitogen-activated protein kinase (MAPK) pathway inhibition. Our results suggest that PEDF mRNA level is significantly decreased in OC tissues and cells and has a significant negative correlation with OC progression and the level of tumor-related macrophage markers. Furthermore, OC tumors overexpressing PEDF show suppressed growth viability and increased apoptosis rate. The fluorescence activated cell sorting (FACS) analysis reveals that PEDF can promote macrophage polarization in OC tumors towards M1 subtype. Mechanistically, we found that ATGL and extracellular-regulated kinase 1/2 (ERK1/2) signaling are involved in the regulation of macrophage polarization in OC tumors by PEDF. Taken together, these data indicate that the role of PEDF in regulating the polarization of tumor-associated macrophages may make it a potential therapeutic strategy for the treatment of OC in the future.

Upregulated pigment epithelium-derived factor (PEDF) promotes trophoblast apoptosis and inhibits invasion in preeclampsia

Preeclampsia (PE) is a severe pregnancy-specific disorder. Previous findings indicated that pigment epithelium-derived factor (PEDF) was upregulated in placentas of women with PE. Here, we investigated the role of PEDF in trophoblast function, especially under hypoxia. The effects of hypoxia on the morphology of extravillous trophoblast (EVT)-derived HTR-8Svneo cells were observed under inverted microscope. Transfections with Lipofectamine LTX were performed according to the manufacturer's protocol. The expression of PEDF protein and mRNA were confirmed by immunofluorescence (IF) and quantitative real-time PCR (qPCR). Apoptosis was detected by transferase-mediated dUTP nick end labeling (TUNEL) assay, and proliferation of trophoblast was detected by CCK-8 method. The invasion capacity of trophoblast was assessed by Transwell assay. PEDF was expressed in HTR-8/SVneo under both normoxia and hypoxic stress. However, cells of hypoxia groups had higher expression level of PEDF, increased apoptosis and decreased invasion capability, as compared with normoxia group. Moreover, after transfection with plasmid expressing PEDF gene, overexpression of PEDF modulated trophoblast activities. In addition, PEDF expression was negatively associated with invasion while positively correlated with apoptosis.Our data suggest that PEDF is an important factor to maintain the biological function of trophoblast cells, thus representing a rational therapeutic target in PE.

PEDF-Mediated Mitophagy Triggers the Visual Cycle by Enhancing Mitochondrial Functions in a H2O2-Injured Rat Model.

Retinal degenerative diseases result from oxidative stress and mitochondrial dysfunction, leading to the loss of visual acuity. Damaged retinal pigment epithelial (RPE) and photoreceptor cells undergo mitophagy. Pigment epithelium-derived factor (PEDF) protects from oxidative stress in RPE and improves mitochondrial functions. Overexpression of PEDF in placenta-derived mesenchymal stem cells (PD-MSCs; PD-MSCsPEDF) provides therapeutic effects in retinal degenerative diseases. Here, we investigated whether PD-MSCsPEDF restored the visual cycle through a mitophagic mechanism in RPE cells in hydrogen peroxide (H2O2)-injured rat retinas. Compared with naïve PD-MSCs, PD-MSCsPEDF augmented mitochondrial biogenesis and translation markers as well as mitochondrial respiratory states. In the H2O2-injured rat model, intravitreal administration of PD-MSCsPEDF restored total retinal layer thickness compared to that of naïve PD-MSCs. In particular, PTEN-induced kinase 1 (PINK1), which is the major mitophagy marker, exhibited increased expression in retinal layers and RPE cells after PD-MSCPEDF transplantation. Similarly, expression of the visual cycle enzyme retinol dehydrogenase 11 (RDH11) showed the same patterns as PINK1 levels, resulting in improved visual activity. Taken together, these findings suggest that PD-MSCsPEDF facilitate mitophagy and restore the loss of visual cycles in H2O2-injured rat retinas and RPE cells. These data indicate a new strategy for next-generation MSC-based treatment of retinal degenerative diseases.

Overexpression of pigment epithelium-derived factor in placenta-derived mesenchymal stem cells promotes mitochondrial biogenesis in retinal cells

AbstractPigment epithelium-derived factor (PEDF) plays a role in protecting retinal pigment epithelial (RPE) cells from oxidative stress (OS), a causative factor of RPE cell death. Genetically modified mesenchymal stem cells (MSCs) can be used to treat critical and incurable retinal diseases. Here, we overexpressed PEDF in placenta-derived MSCs (PD-MSCsPEDF, PEDF+) using a nonviral gene delivery system and evaluated the characteristics of PD-MSCsPEDF and their potential regenerative effects on RPE cells damaged by H2O2-induced OS. PD-MSCsPEDF maintained their stemness, cell surface marker, and differentiation potential characteristics. Compared to naive cells, PD-MSCsPEDF promoted mitochondrial respiration by enhancing biogenesis regulators (e.g., NRF1, PPARGC1A, and TFAM) as well as antioxidant enzymes (e.g., HMOXs, SODs, and GPX1). Compared to OS-damaged RPE cells cocultured with naive cells, OS-damaged RPE cells cocultured with PD-MSCsPEDF showed PEDF upregulation and VEGF downregulation. The expression levels of antioxidant genes and RPE-specific genes, such as RPE65, RGR, and RRH, were significantly increased in RPE cells cocultured with PD-MSCsPEDF. Furthermore, OS-damaged RPE cells cocultured with PD-MSCsPEDF had dramatically enhanced mitochondrial functions, and antiapoptotic effects improved due to cell survival signaling pathways. In the H2O2-induced retinal degeneration rat model, compared to administration of the naive counterpart, intravitreal administration of PD-MSCsPEDF alleviated proinflammatory cytokines and restored retinal structure and function by increasing PEDF expression and decreasing VEGF expression. Intravitreal administration of PD-MSCsPEDF also protected retinal degeneration against OS by increasing antioxidant gene expression and regulating the mitochondrial ROS levels and biogenesis. Taken together, PEDF overexpression in PD-MSCs improved the mitochondrial activities and induced OS-damaged RPE cell regeneration by regulating the oxidative status and mitochondrial biogenesis in vitro and in vivo. These data suggest that genetic modification of PEDF in PD-MSCs might be a new cell therapy for the treatment of retinal degenerative diseases.Overexpression of pigment epithelium-derived factor (PEDF) in placenta-derived mesenchymal stem cells (PD-MSCs) improved the mitochondrial activities, and induced regeneration of oxidative stress-damaged RPE through regulating oxidative status and mitochondrial biogenesis. Therefore, genetic modification of PD-MSCs with PEDF might be a new cell therapy for treatment of retinal degenerative diseases.

PlGF silencing combined with PEDF overexpression: Modeling RPE secretionas potential therapy for retinalneovascularization.

Ocular neovascularization is a defining feature of several blinding diseases. We have previously described the effectiveness of long-term pigment epithelium-derived factor (PEDF) expression in the retina of diabetic mice in ameliorating some diabetic retinopathy hallmarks. In this study, we aimed to investigate if the antiangiogenic potential of PEDF overexpression was enhanced in combination with placental growth factor (PlGF) silencing. Human RPE cells were transfected with a self-replicating episomal vector (pEPito) for PEDF overexpression and/or a siRNA targeting PlGF gene. Conditioned media from PEDF overexpression, from PlGF inhibition and from their combination thereof were used to culture human umbilical vein endothelial cells, and their proliferation rate, migration capacity, apoptosis and ability to form tube-like structures were analyzed in vitro. We here demonstrate that pEPito-driven PEDF overexpression in combination with PlGF silencing in RPE cells does not affect their viability and results in an enhanced antiangiogenic activity in vitro. We observed a significant decrease in the migration and proliferation of endothelial cells, and an increase in apoptosis induction as well as a significant inhibitory effect on tube formation. Our findings demonstrate that simultaneous PEDF overexpression and PlGF silencing strongly impairs angiogenesis compared with the single approaches, providing a rationale for combining these therapies as a new treatment for retinal neovascularization.

Elevated pigment epithelium-derived factor induces diabetic erectile dysfunction via interruption of the Akt/Hsp90β/eNOS complex.

Diabetes mellitus erectile dysfunction (DMED) is a common complication of diabetes. The level of pigment epithelium-derived factor (PEDF) is significantly upregulated in the serum of individuals with obesity and diabetes. However, whether elevated PEDF levels contribute to DMED remains unknown. This study aimed to investigate the pathogenic role of PEDF and its related mechanism in DMED. We enrolled 65 men, of whom 20 were nondiabetic control participants, 21 participants with diabetes but without erectile dysfunction, and 24 with DMED. The International Index of Erectile Function (IIEF-5) questionnaire was administered to evaluate erectile function. Plasma PEDF in diabetic participants and streptozotocin (STZ)-induced diabetic animals was detected by ELISA. Erectile function was evaluated by measuring the intracavernous pressure (ICP) and the ICP/mean arterial pressure (MAP) ratio in STZ-induced diabetic rats treated with PEDF-neutralising antibody (PEDF-Ab), db/db mice treated with PEDF-Ab, and Pedf knockout mice with STZ-induced diabetes. The overexpression of PEDF was implemented by intraperitoneal injection of recombinant PEDF and intracavernous injection of PEDF-expressing adenovirus. A mechanistic study was performed by immunofluorescence staining, bimolecular fluorescence complementation (BiFC), immunoprecipitation and western blotting. We found that the plasma level of PEDF was significantly higher in participants with DMED compared with diabetic counterparts without erectile dysfunction and nondiabetic controls. Interestingly, PEDF levels were negatively correlated with plasma nitrite/nitrate levels and erectile function in DMED patients and STZ-induced diabetic rats. Furthermore, overexpression of PEDF significantly suppressed ICP and endothelial nitric oxide synthase (eNOS) phosphorylation in control rats. In contrast, the PEDF-Ab and Pedf knockout ameliorated ICP and eNOS phosphorylation in diabetic rats and mice. Mechanistically, PEDF promoted the membrane translocation of Hsp90β and directly bound to the amino acid residues 341-724 of Hsp90β on the endothelial cell surface, subsequently blocking intracellular Hsp90β/Akt/eNOS complex formation and downregulating eNOS phosphorylation. These results indicate that elevated PEDF levels contribute to impaired erectile function by suppressing Hsp90β-mediated eNOS phosphorylation and that PEDF may represent a novel therapeutic target for diabetic erectile dysfunction. Graphical abstract.

Malignant ascites: a source of therapeutic protein against ovarian cancer?

Ovarian cancer is the fifth leading cause of cancer-related death in the world. Some ovarian cancer patients present large amount of ascites at the time of diagnosis which may play an active role in tumor development. In earlier studies, we demonstrated that the acellular fraction of ascites can induce apoptosis of ovarian cancer cells. The current study identifies pigment epithelium derived factor (PEDF) as the molecule responsible for the apoptotic effect of ascites and evaluates the Sleeping Beauty transposon (SBT) system as a new tool for PEDF gene therapy against ovarian cancer. We utilize gel filtration, mass spectrometry, affinity column, cell viability assay, tumor development on chick chorioallantoic membrane and molecular biology techniques for these purposes. PEDF was thus identified as the agent responsible for the effects of ascites on ovarian cancer cell viability and tumor growth. Interestingly, the PEDF expression is decreased in ovarian cancer cells compared to healthy ovarian cells. However, the level of PEDF is higher in ascites than in serum of ovarian cancer patients suggesting that cells present in the tumor environment are able to secrete PEDF. We then used the SBT system to stably induce PEDF expression in ovarian cancer cells. The overexpression of PEDF significantly reduced the tumor growth derived from these cells. In conclusion, the results presented here establish that PEDF is a therapeutic target and that PEDF from ascites or SBT could be utilized as a therapeutic strategy for the treatment of ovarian cancer.

Effects of a combinatorial treatment with gene and cell therapy on retinal ganglion cell survival and axonal outgrowth after optic nerve injury.

After an injury, axons in the central nervous system do not regenerate over large distances and permanently lose their connections to the brain. Two promising approaches to correct this condition are cell and gene therapies. In the present work, we evaluated the neuroprotective and neuroregenerative potential of pigment epithelium-derived factor (PEDF) gene therapy alone and combined with human mesenchymal stem cell (hMSC) therapy after optic nerve injury by analysis of retinal ganglion cell survival and axonal outgrowth. Overexpression of PEDF by intravitreal delivery of AAV2 vector significantly increased Tuj1-positive cells survival and modulated FGF-2, IL-1ß, Iba-1, and GFAP immunostaining in the ganglion cell layer (GCL) at 4 weeks after optic nerve crush, although it could not promote axonal outgrowth. The combination of AAV2.PEDF and hMSC therapy showed a higher number of Tuj1-positive cells and a pronounced axonal outgrowth than unimodal therapy after optic nerve crush. In summary, our results highlight a synergistic effect of combined gene and cell therapy relevant for future therapeutic interventions regarding optic nerve injury.

Pigment Epithelium-Derived Factor Promotes Axon Regeneration and Functional Recovery After Spinal Cord Injury

Although neurons in the adult mammalian CNS are inherently incapable of regeneration after injury, we previously showed that exogenous delivery of pigment epithelium-derived factor (PEDF), a 50-kDa neurotrophic factor (NTF), promoted adult retinal ganglion cell neuroprotection and axon regeneration. Here, we show that PEDF and other elements of the PEDF pathway are highly upregulated in dorsal root ganglion neurons (DRGN) from regenerating dorsal column (DC) injury paradigms when compared with non-regenerating DC injury models. Exogenous PEDF was neuroprotective to adult DRGN and disinhibited neurite outgrowth, whilst overexpression of PEDF after DC injury in vivo promoted significant DC axon regeneration with enhanced electrophysiological, sensory, and locomotor function. Our findings reveal that PEDF is a novel NTF for adult DRGN and may represent a therapeutically useful factor to promote functional recovery after spinal cord injury.

A "Hibernating-Like" Viable State Induced by Lentiviral Vector-Mediated Pigment Epithelium-Derived Factor Overexpression in Rat Acute Ischemic Myocardium.

The failure to maintain the viability of ischemic myocardium is one of the mechanisms that causes ischemic heart dysfunction after revascularization. Hibernating myocardium is considered to be able to maintain long-term viability during chronic hypoperfusion. Pigment epithelium-derived factor (PEDF) decreases the contractility of hypoxic cardiomyocytes and protects cardiomyocytes against ischemic injury, which is strikingly similar to the pathophysiologic characteristics of hibernating myocardium. It was therefore postulated that PEDF may induce acute ischemic myocardium into a "hibernating-like" state to maintain its viability. Adult Sprague-Dawley rat models of acute myocardial infarction were surgically established. Lentiviral vectors carrying the PEDF gene (PEDF-LVs) were delivered into myocardium with infarction to overexpress PEDF locally. It was found that PEDF local overexpression significantly reduced myocardial infarct size and cardiomyocytes necrosis but did not improve cardiac function at rest. The contractile reserve assessed by low-dose dobutamine stress echocardiography and "perfusion-metabolism mismatch" assessed by positron emission tomography, which are the characteristics of viable myocardium in hibernation, were observed in the PEDF overexpressed ischemic heart. Ultrastructural changes observed by electron microscopy and glycogen deposition explored by Periodic acid-Schiff staining were similar to the histological characteristics of hibernating myocardium. Moreover, PEDF overexpression protected the cardiomyocytes against anoxic injury and retained their functional recovery potential after reoxygenation in vitro. PEDF local overexpression may induce acute ischemic myocardium into a "hibernating-like" state and maintain its viability. This novel effect of PEDF presents an important clinical approach to enhance functional recovery after revascularization therapy in acute myocardial infarction.

Pigment epithelium-derived factor inhibits lung cancer migration and invasion by upregulating exosomal thrombospondin 1

Exosomes are implicated in cancer cell development, migration and invasion. Pigment epithelium-derived factor (PEDF) is a secreted anticancer protein that can regulate lung cancer progression; however, the role of PEDF in non-small cell lung cancer (NSCLC), including metastasis and cancer cell-derived exosome secretion, is unclear. In this study, we analyzed the effects of PEDF on exosome-mediated migration, invasion, and tumorigenicity of cultured NSCLC cells. The results showed that PEDF overexpression significantly reduced NSCLC invasion and migration, while inducing cell aggregation, whereas PEDF knockdown had the opposite effects. Exosomes from NSCLC cells treated with recombinant PEDF had a significantly reduced ability to promote cancer cell motility, migration, and invasion compared to exosomes from untreated cells. Exosomes from PEDF-treated cells contained thrombospondin 1 (THBS1), which inhibited cytoskeletal remodeling and exosome-induced lung cancer cell motility, migration, and invasion. Furthermore, PEDF-overexpressing NSCLC cells formed smaller xenograft tumors with higher THBS1 expression compared to control tumors. Our findings indicate that PEDF decreases the metastatic potential of NSCLC cells through regulation of THBS1 release in cancer cell-derived exosomes, thus uncovering a new mechanism of lung cancer progression.

Pigment epithelium-derived factor inhibits metastasis and invasion of breast cancer via regulation of epithelial-mesenchymal transition

To investigate whether pigment epithelium-derived factor (PEDF) inhibits invasion and metastasis of breast cancer through regulation of epithelial-mesenchymal transition. The expressions of PEDF, vimentin, and E-cadherin were detected in 119 breast cancer tissues using immunohistochemistry. SK-BR-3 breast cancer cell models of PEDF knockdown and PEDF overexpression were established by transfecting the cells with a PEDF-siRNA vector and a lentivirusPEDF vector, respectively. Western blotting was used to detect the changes in the expressions of PEDF, vimentin, and E-cadherin in the cells, and the cell invasion and migration ability was assessed using scratch wound healing assay and transwell migration assay. PEDF positivity rate was significantly lowered in breast cancer tissues compared with the adjacent tissues. PEDF was positively correlated with the tumor size and the expression level of E-cadherin (r=0.473, P < 0.001), but was negatively correlated with vimentin expression (r=-0.412, P < 0.001). Transwell invasion experiment showed that PEDF interference enhanced the cell invasion and metastasis, while PEDF overexpression inhibited the invasion and migration of SKBR-3 cells. Western blotting showed that PEDF knockdown significantly decreased the expression of E-cadherin (P < 0.05) and increased vimentin expression in the cells (P < 0.05). PEDF is closely related with the metastasis of breast cancer cells through regulation of epithelial-mesenchy.

MiR-93-3p inhibition suppresses clear cell renal cell carcinoma proliferation, metastasis and invasion.

miRNA dysregulation is associated with many human diseases, including cancer. This study explored the effects of miR-93-3p on clear cell renal cell carcinoma (ccRCC). We found that miR-93-3p is upregulated an average of 38-fold in 138 ccRCC specimens compared to matched normal kidney tissues, which correlated with poor patient outcome. miR-93-3p inhibition reduced ccRCC cell growth, invasion, and migration in vitro and in a mouse xenograft model. A search of the TargetScan, miRanda, and PicTar databases revealed that miR-93-3p is predicted to regulate pigment epithelium-derived factor (PEDF). A direct PEDF-miR-93-3p interaction was confirmed via dual-luciferase reporter assays. Like miR-93-3p inhibition, PEDF overexpression induced cell apoptosis and inhibited migration and invasion. Additionally, co-transfection with PEDF siRNA reversed the effects of miR-93-3p inhibition in ccRCC cells. Thus, miR-93-3p is a likely ccRCC oncogene that acts by regulating PEDF. These results suggest that miR-93-3p may predict ccRCC patient clinical outcome and serve as a novel anti-ccRCC therapeutic target.