In the past few decades, interest in the therapeutic benefits of exosomes and extracellular vesicles (EVs) has grown exponentially. Exosomes/EVs are small particles which are produced and exocytosed ...by cells throughout the body. They are loaded with active regulatory and stimulatory molecules from the parent cell including miRNAs and enzymes, making them prime targets in therapeutics and diagnostics. Breast milk, known for years to have beneficial health effects, contains a population of EVs which may mediate its therapeutic effects. This review offers an update on the therapeutic potential of exosomes/EVs in disease, with a focus on EVs present in human breast milk and their remedial effect in the gastrointestinal disease necrotizing enterocolitis. Additionally, the relationship between EV miRNAs, health, and disease will be examined, along with the potential for EVs and their miRNAs to be engineered for targeted treatments.
Abstract Background Macrophages can be polarized into proinflammatory (M1) and anti-inflammatory (M2) subtypes. However, whether macrophage polarization plays a role in necrotizing enterocolitis ...(NEC) remains unknown. Materials and methods Macrophages were derived from the THP-1 human monocyte cell line. Apoptosis of human fetal small intestinal epithelial FHs-74 cells was determined by Annexin V/propidium iodide flow cytometry and by Western blotting to detect cleaved caspase-3. The effect of heparin-binding epidermal growth factor-like growth factor (HB-EGF) on macrophage polarization was determined by flow cytometry with M1/M2 markers and real time polymerase chain reaction. In vivo , experimental NEC was induced in mouse pups by repeated exposure to hypoxia, hypothermia, and hypertonic feedings. Intestinal histologic sections were subjected to immunohistochemical staining for the detection of M1 and M2 macrophages. Results In vitro , FHs-74 cell apoptosis was increased after coculture with macrophages and lipopolysaccharide (LPS). This apoptosis was increased by exposure to M1-conditioned medium and suppressed by exposure to M2-conditioned medium. HB-EGF significantly decreased LPS-induced M1 polarization and promoted M2 polarization via signal transducers and activators of transcription 3 activation. Addition of HB-EGF to LPS-stimulated macrophages suppressed the proapoptotic effects of the macrophages on FHs-74 cells. In vivo , we found enhanced intestinal macrophage infiltration in pups subjected to NEC, most of which were M1 macrophages. HB-EGF treatment of pups subjected to experimental NEC significantly reduced M1 and increased M2 polarization and protected the intestines from NEC. Conclusions M1 macrophages promote NEC by increasing intestinal epithelial apoptosis. HB-EGF protects the intestines from NEC by preventing M1 and promoting M2 polarization.
Necrotizing enterocolitis (NEC) is a leading cause of death in premature infants. Breast feeding decreases the incidence of NEC but, even with aggressive promotion of nursing in Neonatal Intensive ...Care Units, morbidity and mortality remain high. Previous studies from our laboratory have demonstrated that extracellular vesicles (EVs) purified from mouse and rat stem cells can protect the intestines from NEC. The aim of this study was to determine whether human breast milk (BM)-derived EVs could prevent NEC.
EVs were purified from human donor breast milk. NEC was induced in premature rat pups by exposure to asphyxia/hypothermia/hypercaloric feeds. Pups were randomized to: (1) breast fed, no injury, (2) NEC, (3) NEC + BM-derived EVs once intraperitoneally (IP), (4) NEC + BM-derived EVs enterally (PO) with each feed. Intestinal tracts were examined for histologic damage. Additionally, the effect of BM-derived EVs on intestinal epithelial cells (IEC) subjected to hypoxia/reoxygenation injury in vitro was examined.
NEC incidence was 0% in breast-fed pups and 62% in pups subjected to NEC. IP administration of BM-derived EVs decreased NEC incidence to 29% and enteral administration further decreased NEC incidence to 11.9%. (p < 0.05). BM-derived EVs significantly increased cell proliferation and decreased apoptosis in IEC in vitro.
Breast milk-derived EVs delivered either IP or enterally significantly decrease the incidence and severity of experimental NEC, protect IEC from injury in vitro, and may represent an innovative therapeutic option for NEC in the future.
Basic science study.
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Necrotizing enterocolitis (NEC) is a devastating disease of newborns, and despite years of research, there is no known cure. The mortality rate of infants with NEC remains as high as 20%–30%. Babies ...who survive NEC frequently have long term complications including short gut syndrome, developmental delays and neurological sequelae. Unfortunately, despite much research over the past years, the precise pathogenesis of the disease is still not completely understood.
Our laboratory has focused on identifying novel therapies to prevent the disease, including the use of stem cells (SC), heparin-binding epidermal growth factor-like growth factor (HB-EGF) and recently, stem cell derived-exosomes, a type of nanovesicle, to combat this illness.
We have outlined the major SC lines and data suggesting potential benefit as a curative or preventive approach for NEC as well as describing several new therapeutic strategies, including stem cell derived- exosomes and HB-EGF for decreasing the incidence and severity of this disease in rat models in our lab.
Overall, our lab has demonstrated that these different types of SC equivalently reduce the incidence and severity of NEC and equally preserve intestinal barrier function during NEC. We have previously demonstrated that AF-MSC can protect the intestines from intestinal injury and may therefore hold strong therapeutic potential for the prevention of NEC. Most recently, our work with stem cell derived-exosomes has shown them to be equivalent to their derived SC lines in decreasing the incidence of this disease.
Necrotizing enterocolitis (NEC) is associated with loss of neurons and glial cells in the enteric nervous system (ENS). Our goal was to determine whether enteric neural stem cell (NSC) ...transplantation, in conjunction with heparin-binding epidermal growth factor-like growth factor (HB-EGF), could protect against experimental NEC.
In vitro, HB-EGF on NSC proliferation and migration, and the effects of receptors utilized by HB-EGF to exert these effects, were determined. In vivo, mouse pups were exposed to experimental NEC and treated with NSC alone, HB-EGF alone, NSC+HB-EGF, or HB-EGF overexpressing NSC. NSC engraftment and differentiation into neurons in the ENS, intestinal injury, intestinal permeability, and intestinal motility were determined.
HB-EGF promoted NSC proliferation via ErbB-1 receptors and enhanced NSC migration via ErbB-1, ErbB-4, and Nardilysin receptors. HB-EGF significantly enhanced the engraftment of transplanted NSC into the ENS during NEC. NSC transplantation significantly reduced NEC incidence and improved gut barrier function and intestinal motility, and these effects were augmented by simultaneous administration of HB-EGF or by transplantation of HB-EGF overexpressing NSC.
HB-EGF promotes NSC proliferation and migration. HB-EGF and NSC reduce intestinal injury and improve gut barrier function and intestinal motility in experimental NEC. Combined HB-EGF and NSC transplantation may represent a potential future therapy to prevent NEC.
Abstract Purpose Treatment options for necrotizing enterocolitis (NEC) remain inadequate. Bone marrow-derived mesenchymal stem cells (BM-MSCs) can protect the intestines from NEC. Exosomes are ...nanoparticle-sized vesicles with important cell signaling capabilities. The objective of this study was to determine whether BM-MSC-derived exosomes can prevent NEC. Methods Rat pups were either breast fed (Group 1) or subjected to experimental NEC and randomized to receive either no treatment (Group 2) or an intraperitoneal (IP) injection of PBS (Group 3), BM-MSC (Group 4), or BM-MSC-derived exosomes (Group 5). Histologic injury grade and intestinal permeability were determined. The effect of BM-MSC-derived exosomes on IEC-6 intestinal epithelial cells in an in vitro scrape model of wound healing was also determined. Results Animals exposed to NEC that were either untreated or received PBS alone had an NEC incidence of 46% and 41%, respectively ( p = 0.61). Compared to untreated pups, the incidence of NEC was significantly lower in pups treated with either BM-MSC (9%, p = 0.0003) or MB-MSC-derived exosomes (13%, p = 0.0008). Similar results were found for intestinal permeability. Wound healing in IEC-6 cells was significantly increased by BM-MSC-derived exosomes. Conclusion BM-MSC-derived exosomes protect the intestines from NEC and may represent a novel, cell-free, preventative therapy for NEC in the future.
Improvements in patient care are directly affected by scientific discovery, and surgeons have historically played a vital role in this process. However, increasing clinical demands and ...incentivization for pure clinical productivity present challenges for promoting academic productivity. The objective of this work was to analyze the effects of adding an academic relative value unit (aRVU) scoring system to an existing work RVU (wRVU)-based incentivization plan on academic productivity in a Department of Pediatric Surgery.
Prior to 2012, incentive bonuses in our Department were mainly based on clinical wRVU activities. A weighted scoring system for 30 specific aRVUs was established in 2012. Incentivization for wRVUs vs. aRVUs was based on the clinical full-time equivalent (cFTE) of each faculty member. Academic activities incentivized included grant submissions/funding, peer reviewed publications, national presentations, Study Section participation, education and mentoring activities, receipt of research or teaching awards, initiation of Institutional Review Board (IRB) protocols, new academic society committee memberships/chairpersons, and patents. Academic progress was analyzed from 2012 to 2020.
During the study period, annual external federal funding increased from $750,168 to $5,768,243 (7.7-fold increase); annual peer-reviewed publications increased from 24 to 140 (5.8-fold increase); annual national presentations accepted for oral/poster presentations nearly doubled; and faculty members and their trainees received 41 competitive research awards including 8 American Pediatric Surgical Association Awards, 9 American Academy of Pediatrics Section on Surgery Awards, and 3 American College of Surgeons Awards. During the same study period, wRVUs increased by 8%.
Incentivization based on the addition of an aRVU system to a pre-existing wRVU system was associated with a significant increase in academic productivity, while still maintaining clinical productivity. Implementing an aRVU program is an important means of increasing academic productivity in Pediatric and other Surgery Departments.
Abstract This lecture will describe how the properties of heparin-binding EFG-like growth factor (HB-EGF) can be utilized to protect the intestines from necrotizing enterocolitis (NEC), to augment ...the effects of stem cells in the treatment of NEC, and to improve the production of tissue-engineered intestine. It will also explore the role of the enteric nervous system in the development of NEC.
Interstitial cells of Cajal (ICCs) are pacemaker cells in the intestine, and their function can be compromised by loss of C-KIT expression. Macrophage activation has been identified in intestine ...affected by Hirschsprung disease-associated enterocolitis (HAEC). In this study, we examined proinflammatory macrophage activation and explored the mechanisms by which it downregulates C-KIT expression in ICCs in colon affected by HAEC. We found that macrophage activation and TNF-α production were dramatically increased in the proximal dilated colon of HAEC patients and 3-week-old Ednrb-/- mice. Moreover, ICCs lost their C-KIT+ phenotype in the dilated colon, resulting in damaged pacemaker function and intestinal dysmotility. However, macrophage depletion or TNF-α neutralization led to recovery of ICC phenotype and restored their pacemaker function. In isolated ICCs, TNF-α-mediated phosphorylation of p65 induced overexpression of microRNA-221 (miR-221), resulting in suppression of C-KIT expression and pacemaker currents. We also identified a TNF-α/NF-κB/miR-221 pathway that downregulated C-KIT expression in ICCs in the colon affected by HAEC. These findings suggest the important roles of proinflammatory macrophage activation in a phenotypic switch of ICCs, representing a promising therapeutic target for HAEC.
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