Recent evidence suggests mesenchymal stem cells (MSCs) can downmodulate bleomycin-induced lung injury, and umbilical cord blood (UCB) is a promising source for human MSCs. This study examined whether intratracheal or intraperitoneal transplantation of human UCB-derived MSCs can attenuate hyperoxia-induced lung injury in immunocompetent newborn rats. Wild-type Sprague-Dawley rats were randomly exposed to 95% oxygen or air from birth. In the transplantation groups, a single dose of PKH26-labeled human UCBderived MSCs was administered either intratracheally (2 × 10 6 cells) or intraperitoneally (5 × 10 5 cells) at postnatal day (P) 5. At P14, the harvested lungs were examined for morphometric analyses of alveolarization and TUNEL staining, as well as the myeoloperoxidase activity, the level of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and transforming growth factor (TGF)-β mRNA, α-smooth muscle actin (SMA) protein, and collagen levels. Differentiation of MSCs to the respiratory epithelium was also evaluated both in vitro before transplantation and in vivo after transplantation. Despite one fourth dosage of MSCs, significantly more PKH26-labeled donor cells were recovered with intratracheal administration than with intraperitoneal administration both during normoxia and hyperoxia. The hyperoxia-induced increase in the number of TUNELpositive cells, myeloperoixdase activity, and the level of IL-6 mRNA were significantly attenuated with both intratracheal and intraperitoneal MSCs transplantation. However, the hyperoxia-induced impaired alveolarization and increased the level of TNF-α and TGF-β mRNA, α-SMA protein, and collagen were significantly attenuated only with intratracheal MSCs transplantation. MSCs differentiated into respiratory epithelium in vitro and a few PKH26-positive donor cells were colocalized with pro surfactant protein C in the damaged lungs. In conclusion, intratracheal transplantation of human UCB-derived MSCs is more effective than intraperitoneal transplantation in attenuating the hyperoxia-induced lung injury in neonatal rats.Key words: Hyperoxic lung injury; Transplantation; Animal; Newborn; Inflammation; Cell differentiation INTRODUCTION development of hyperoxic neonatal lung injury and BPD (42).Recently, the exogenous administration of bone marBronchopulmonary dysplasia (BPD) is a chronic pulmonary disease that usually occurs in premature infants row (BM)-derived mesenchymal stem cells (MSCs) significantly attenuated the bleomycin-induced lung injury receiving prolonged ventilatory support and oxygen supplementation. It is also an important cause of mortality by downmodulating the inflammatory responses and ameliorating their fibrotic effects (33). In addition, the transand long-term respiratory morbidity with few effective treatments (3,6). The histopathological characteristics of planted cells assumed the lung parenchymal cell phenotypes (34). The anti-inflammatory effect of MSCs has BPD include impaired alveolarization and interstitial fibrosis (30). Prolonged exposure of neonata...

I ntraventricular hemorrhage (IVH) is a serious complication of preterm birth, and the number of infants at high risk for developing IVH is increasing as a result of the rise in the absolute number and improved survival of very premature infants with recent advances in neonatal intensive care medicine. 1,2 Over half of infants with severe IVH (grade ≥3) die or develop posthemorrhagic hydrocephalus (PHH), which requires shunt surgery in up to 70% of cases.3 IVH is associated with brain damage, especially to the periventricular white matter, which is exacerbated by PHH, and finally results in increased mortality and long-term neurological morbidity, such as seizure, cerebral palsy, and developmental retardation in survivors. [4][5][6] Until now, however, there has not been any effective treatment to prevent PHH or ameliorate brain damage after severe IVH in preterm infants, so it remains a major problem of neonatal intensive care.Although the precise mechanism has not been completely delineated, the pathogenesis of communicating progressive posthemorrhagic ventricular dilatation has been explained by inflammation within subarachnoid spaces attributable to blood contact and deposition of blood products. 7,8 This obliterative arachnoiditis leads to dysfunction of arachnoid granulations, which reduces cerebrospinal fluid (CSF) resorption and increases intracranial pressure, resulting in venous infarction with decreased cerebral perfusion. 8 Moreover, inflammatory cytokines originating from blood products in the cerebral ventricles may injure the periventricular white matter. 9,10 Therefore, new therapeutic modalities with anti-inflammatory capabilities to treat PHH and brain damage after severe IVH would be of great value.Recent preclinical research reported that an anti-inflammatory agent, a cycloxygenase-2 inhibitor, noticeably reduced reactive gliosis and improved neurological impairment after IVH in a newborn rabbit model. This finding indicates that modulating inflammation could be a key factor in IVH therapy for preterm Background and Purpose-Severe intraventricular hemorrhage (IVH) in premature infants and the ensuing posthemorrhagic hydrocephalus cause significant mortality and neurological disabilities, and there are currently no effective therapies. This study determined whether intraventricular transplantation of human umbilical cord blood-derived mesenchymal stem cells prevents posthemorrhagic hydrocephalus development and attenuates brain damage after severe IVH in newborn rats. Methods-To induce severe IVH, 100 μL of blood was injected into each lateral ventricle of postnatal day 4 (P4) SpragueDawley rats. Human umbilical cord blood-derived mesenchymal stem cells or fibroblasts (1×10 5 ) were transplanted intraventricularly under stereotaxic guidance at P6. Serial brain MRI and behavioral function tests, such as the negative geotaxis test and rotarod test, were performed. At P32, brain tissue and cerebrospinal fluid were obtained for histological and biochemical analyses. Results-Intraventricular tran...

SummaryRecently, we demonstrated that intratracheal transplantation of human umbilical cord blood‐ derived mesenchymal stem cells (MSCs) attenuates Escherichia (E) coli‐ induced acute lung injury primarily by down‐ modulating inflammation and enhancing bacterial clearance iQn mice. This study was performed to elucidate the mechanism underlying the antibacterial effects of MSCs. The growth of E. coli in vitro was significantly inhibited only by MSCs or their conditioned medium with bacterial preconditioning, but not by fibroblasts or their conditioned medium. Microarray analysis identified significant up‐ regulation of toll‐ like receptors (TLR)‐ 2 and TLR‐ 4, and β‐ defensin 2 (BD2) in MSCs compared with fibroblasts after E. coli exposure. The increased BD2 level and the in vitro antibacterial effects of MSCs were abolished by specific antagonist or by siRNA‐ mediated knockdown of TLR‐ 4, but not TLR‐ 2, and restored by BD2 supplementation. The in vivo down‐ modulation of the inflammatory response and enhanced bacterial clearance, increased BD2 secretion and the resultant protection against E. coli‐ induced pneumonia observed only with MSCs, but not fibroblasts, transplantation in mice, were abolished by knockdown of TLR‐ 4 with siRNA transfection. Our data indicate that BD2 secreted by the MSCs via the TLR‐ 4 signalling pathway is one of the critical paracrine factors mediating their microbicidal effects against E. coli, both in vitro and in vivo. Furthermore, TLR‐ 4 from the transplanted MSCs plays a seminal role in attenuating in vivo E. coli‐ induced pneumonia and the ensuing acute lung injury through both its anti‐ inflammatory and antibacterial effects.

Melanoma skin cancer is one of the most dangerous skin cancers and the main cause of skin-cancer-related mortality. Hyaluronic acid (HA) has been used as an effective transdermal delivery carrier of chemical drugs and biopharmaceuticals. In this work, a nanographene oxide-HA conjugate (NGO-HA) was synthesized for photothermal ablation therapy of melanoma skin cancer using a near-infrared (NIR) laser. Confocal microscopy and ex vivo bioimaging clearly visualized the remarkable transdermal delivery of NGO-HA to tumor tissues in the skin of mice, which might be ascribed to highly expressed HA receptors and relatively leaky structures around tumor tissues, enabling the enhanced permeation and retention of nanoparticles. The NIR irradiation resulted in complete ablation of tumor tissues with no recurrence of tumorigenesis. The antitumor effect was confirmed by ELISA for caspase-3 activity and histological and immunohistochemical analyses with TUNEL assay for tumor apoptosis. Taken together, we could confirm the feasibility of transdermal NGO-HA for photothermal ablation therapy of melanoma skin cancers.

Background: Severe brain injury induced by neonatal stroke causes significant mortality and disability, and effective therapies are currently lacking. We hypothesized that human umbilical cord blood (UcB)-derived mesenchymal stem cells (MScs) can attenuate severe brain injury induced by permanent middle cerebral artery occlusion (McaO) in rat pups. Methods: after confirming severe brain injury involving more than 50% of the ipsilateral hemisphere volume at 1 h after McaO using diffusion-weighted magnetic resonance imaging (MRI) in postnatal day (P)10 rats, human UcB-derived MScs were transplanted intraventricularly. The brain MRI was evaluated periodically up to 28 d after McaO (P38). Sensorimotor function and histology in the peri-infarct tissues were evaluated at the end of the experiment. results: Severe brain injury induced by permanent McaO resulted in decreased survival and body weight gain, increased brain infarct volume as measured by MRI, impaired functional tests such as the rotarod and cylinder test, and histologic abnormalities such as increased terminal deoxynucleotidyl transferase nick-end labeling, reactive microglial marker, and glial fibrillary acidic protein-positive cells in the penumbra. all of these abnormalities were significantly improved by MSc transplantation 6 h after McaO. conclusion: These results suggest that human UcB-derived MScs are a promising therapeutic candidate for the treatment of severe perinatal brain injury including neonatal stroke.

Intratracheal transplantation of human umbilical cord blood (UCB)-derived mesenchymal stem cells (MSCs) attenuates the hyperoxia-induced neonatal lung injury. The aim of this study was to optimize the timing of MSCs transplantation. Newborn Sprague-Dawley rats were randomly exposed to hyperoxia (90% for 2 weeks and 60% for 1 week) or normoxia after birth for 21 days. Human UCB-derived MSCs (5×105 cells) were delivered intratracheally early at postnatal day (P) 3 (HT3), late at P10 (HT10) or combined early+late at P3+10 (HT3+10). Hyperoxia-induced increase in mortality, TUNEL positive cells, ED1 positive alveolar macrophages, myeloperoxidase activity and collagen levels, retarded growth and reduced alveolarization as evidenced by increased mean linear intercept and mean alveolar volume were significantly better attenuated in both HT3 and HT3+10 than in HT10. Hyperoxia-induced up-regulation of both cytosolic and membrane p47phox indicative of oxidative stress, and increased inflammatory markers such as tumor necrosis factor-α, interleukin (IL) -1α, IL-1β, IL-6, and transforming growth factor-β measured by ELISA, and tissue inhibitor of metalloproteinase-1, CXCL7, RANTES, L-selectin and soluble intercellular adhesion molecule-1 measured by protein array were consistently more attenuated in both HT3 and HT3+10 than in HT10. Hyperoxia-induced decrease in hepatocyte growth factor and vascular endothelial growth factor was significantly up-regulated in both HT3 and HT3+10, but not in HT10. In summary, intratracheal transplantation of human UCB derived MSCs time-dependently attenuated hyperoxia-induced lung injury in neonatal rats, showing significant protection only in the early but not in the late phase of inflammation. There were no synergies with combined early+late MSCs transplantation.

BackgroundHuman umbilical cord blood (UCB)-derived mesenchymal stem cells (MSCs) attenuate hyperoxic neonatal lung injury primarily through anti-inflammatory effects. We hypothesized that intratracheal transplantation of human UCB-derived MSCs could attenuate Escherichia coli (E. coli)-induced acute lung injury (ALI) in mice by suppressing the inflammatory response.MethodsEight-week-old male ICR mice were randomized to control or ALI groups. ALI was induced by intratracheal E. coli instillation. Three-hours after E. coli instillation, MSCs, fibroblasts or phosphate-buffered saline were intratracheally administered randomly and survival was analyzed for 7 days post-injury. Lung histology including injury scores, myeloperoxidase (MPO) activity, and protein levels of interleukin (IL)-1α, IL-1β, IL-6, tumor necrosis factor (TNF)-α, and macrophage inflammatory protein (MIP)-2 as well as the wet-dry lung ratio and bacterial counts from blood and bronchoalveolar lavage (BAL) were evaluated at 1, 3, and 7 days post-injury. Levels of inflammatory cytokines in the lung were also profiled using protein macroarrays at day 3 post-injury which showed peak inflammation.ResultsMSC transplantation increased survival and attenuated lung injuries in ALI mice, as evidenced by decreased injury scores on day 3 post-injury and reduced lung inflammation including increased MPO activity and protein levels of IL-1α, IL-1β, IL-6, TNF-α, and MIP-2 on day 3 and 7 post-injury. Inflammatory cytokine profiles in the lungs at day 3 post-injury were attenuated by MSC transplantation. MSCs also reduced the elevated lung water content at day 3 post-injury and bacterial counts in blood and BAL on day 7 post-injury.ConclusionsIntratracheal transplantation of UCB-derived MSCs attenuates E. coli-induced ALI primarily by down-modulating the inflammatory process and enhancing bacterial clearance.

We previously reported the role of vascular endothelial growth factor (VEGF) secreted by mesenchymal stem cells (MSCs) in protecting against neonatal hyperoxic lung injuries. Recently, the paracrine protective effect of MSCs was reported to be primarily mediated by extracellular vesicle (EV) secretion. However, the therapeutic efficacy of MSC-derived EVs and the role of the VEGF contained within EVs in neonatal hyperoxic lung injury have not been elucidated. The aim of the study was to determine whether MSC-derived EVs attenuate neonatal hyperoxic lung injury and, if so, whether this protection is mediated via the transfer of VEGF. We compared the therapeutic efficacy of MSCs, MSC-derived EVs with or without VEGF knockdown, and fibroblast-derived EVs in vitro with a rat lung epithelial cell line challenged with H2O2 and in vivo with newborn Sprague-Dawley rats exposed to hyperoxia (90%) for 14 days. MSCs (1 × 105 cells) or EVs (20 µg) were administered intratracheally on postnatal day 5. The MSCs and MSC-derived EVs, but not the EVs derived from VEGF-knockdown MSCs or fibroblasts, attenuated the in vitro H2O2-induced L2 cell death and the in vivo hyperoxic lung injuries, such as impaired alveolarization and angiogenesis, increased cell death, and activated macrophages and proinflammatory cytokines. PKH67-stained EVs were internalized into vascular pericytes (22.7%), macrophages (21.3%), type 2 epithelial cells (19.5%), and fibroblasts (4.4%) but not into vascular endothelial cells. MSC-derived EVs are as effective as parental MSCs for attenuating neonatal hyperoxic lung injuries, and this protection was mediated primarily by the transfer of VEGF.