Kenneth W. Liechty

Interleukin-10-mediated regenerative postnatal tissue repair is dependent on regulation of hyaluronan metabolism via fibroblast-specific STAT3 signaling.

The cytokine IL‐10 has potent antifibrotic effects in models of adult fibrosis, but the mechanisms of action are unclear. Here, we report a novel finding that IL‐10 triggers a signal transducer and activator of transcription 3(STAT3)–dependent signaling pathway that regulates hyaluronan (HA) metabolism and drives adult fibroblasts to synthesize an HA‐rich pericellular matrix, which mimics the fetal regenerative wound healing phenotype with reduced fibrosis. By using cre‐lox‐mediated novel, inducible, fibroblast‐, keratinocyte‐, and wound‐specific STAT3‐knockdown postnatal mice—plus syngeneic fibroblast cell‐transplant models—we demonstrate that the regenerative effects of IL‐10 in postnatal wounds are dependent on HA synthesis and fibroblast‐specific STAT3‐dependent signaling. The importance of IL‐10‐induced HA synthesis for regenerative wound healing is demonstrated by inhibition of HA synthesis in a murine wound model by administering 4‐methylumbelliferone. Although IL‐10 and STAT3 signaling were intact, the antifibrotic repair phenotype that is induced by IL‐10 overexpression was abrogated in this model. Our data show a novel role for IL‐10 beyond its accepted immune‐regulatory mechanism. The opportunity for IL‐10 to regulate a fibroblast‐specific formation of a regenerative, HA‐rich wound extracellular matrix may lead to the development of innovative therapies to attenuate postnatal fibrosis in organ systems or diseases in which dysregulated inflammation and HA intersect.—Balaji, S., Wang, X., King, A., Le, L. D., Bhattacharya, S. S., Moles, C. M., Butte, M. J., de Jesus Perez, V.A., Liechty, K. W., Wight, T. N., Crombleholme, T. M., Bollyky, P. L., Keswani, S. G. Interleukin‐10‐mediated regenerative postnatal tissue repair is dependent on regulation of hyaluronan metabolism via fibroblast‐specific STAT3 signaling. FASEB J. 31, 868–881 (2017). www.fasebj.org

Mechanisms of mesenchymal stem cell correction of the impaired biomechanical properties of diabetic skin: The role of miR-29a

Diabetic skin has impaired wound healing properties following injury. We have further shown that diabetic skin has weakened biomechanical properties at baseline. We hypothesize that the biomechanical properties of diabetic skin decline during the progression of the diabetic phenotype, and that this decline is due to the dysregulation of miR‐29a, resulting in decreased collagen content. We further hypothesize that treatment with mesenchymal stem cells (MSCs) may improve diabetic wound healing by correction of the dysregulated miR‐29a expression. We analyzed the biomechanical properties, collagen gene expression, collagen protein production, and miR‐29a levels in skin harvested from 6 to 18 week old mice during the development of the diabetic phenotype. We also examined the correction of these impairments by both MSC treatment and the inhibition of miR‐29a. Diabetic skin demonstrated a progressive impairment of biomechanical properties, decreased collagen content, and increased miR‐29a levels during the development of the diabetic phenotype. MSC treatment decreased miR‐29a levels, increased collagen content, and corrected the impaired biomechanical properties of diabetic skin. Additionally, direct inhibition of miR‐29a also increased collagen content in diabetic skin. This decline in the biomechanical properties of diabetic skin during the progression of diabetes may increase the susceptibility of diabetic skin to injury and miR‐29a appears to play a key role in this process.

SCF increases in utero–labeled stem cells migration and improves wound healing

Diabetic skin wounds lack the ability to heal properly and constitute a major and significant complication of diabetes. Nontraumatic lower extremity amputations are the number one complication of diabetic skin wounds. The complexity of their pathophysiology requires an intervention at many levels to enhance healing and wound closure. Stem cells are a promising treatment for diabetic skin wounds as they have the ability to correct abnormal healing. Stem cell factor (SCF), a chemokine expressed in the skin, can induce stem cells migration, however the role of SCF in diabetic skin wound healing is still unknown. We hypothesize that SCF would correct the impairment and promote the healing of diabetic skin wounds. Our results show that SCF improved wound closure in diabetic mice and increased HIF‐1α and vascular endothelial growth factor (VEGF) expression levels in these wounds. SCF treatment also enhanced the migration of red fluorescent protein (RFP)‐labeled skin stem cells via in utero intra‐amniotic injection of lenti‐RFP at E8. Interestingly these RFP+ cells are present in the epidermis, stain negative for K15, and appear to be distinct from the already known hair follicle stem cells. These results demonstrate that SCF improves diabetic wound healing in part by increasing the recruitment of a unique stem cell population present in the skin.

Mesenchymal stem cells correct impaired diabetic wound healing by decreasing ECM proteolysis

Impaired diabetic wound healing is associated with a dermal extracellular matrix protein profile favoring proteolysis; within the healing diabetic wound, this is represented by an increase in activated matrix metalloproteinase (MMPs). Treatment of diabetic wounds with mesenchymal stem cells (MSCs) has been shown to improve wound healing; however, there has not yet been an assessment of their ability to correct dysregulation of MMPs in diabetic wounds. Furthermore, there has been no prior assessment of the role of microRNA29b (miR-29b), an inhibitory regulatory molecule that targets MMP-9 mRNA. Using in vitro models of fibroblast coculture with MSCs and in vivo murine wound healing models, we tested the hypothesis that MSCs correct dysregulation of MMPs in a microRNA-29b-dependent mechanism. In this study, we first demonstrated that collagen I and III protein content is significantly reduced in diabetic wounds, and treatment with MSCs significantly improves collagen I content in both nondiabetic and diabetic wounds. We then found that MMP-9 gene expression and protein content were significantly upregulated in diabetic wounds, indicating elevated proteolysis. Treatment with MSCs resulted in a decrease in MMP-9 gene expression and protein content level in diabetic wounds 3 and 7 days after wounding. Zymographic analysis indicated that MSC treatment also decreased the amount of activated MMP-9 present in diabetic wounds. Furthermore, miR-29b expression was inversely associated with MMP-9 gene expression; miR-29b expression was decreased in diabetic wounds and diabetic fibroblast. Following treatment of diabetic wounds with MSCs, as well as in diabetic fibroblasts cocultured with MSCs, miR-29b was significantly increased. These findings suggest a potential mechanism through which MSCs enhance diabetic wound healing by improving collagen I content in diabetic wounds through decreasing MMP-9 expression and increasing miR-29b expression.

Twin pregnancy complicated by esophageal atresia, duodenal atresia, gastric perforation, and hypoplastic left heart structures in one twin: a case report and review of the literature

BackgroundThe antenatal diagnosis of a combined esophageal atresia without tracheoesophageal fistula and duodenal atresia with or without gastric perforation is a rare occurrence. These diagnoses are difficult and can be suspected on ultrasound by nonspecific findings including a small stomach and polyhydramnios. Fetal magnetic resonance imaging adds significant anatomical detail and can aid in the diagnosis of these complicated cases. Upon an extensive literature review, there are no reports documenting these combined findings in a twin pregnancy. Therefore we believe this is the first case report of an antenatal diagnosis of combined pure esophageal and duodenal atresia in a twin gestation.Case presentationWe present a case of a 30-year-old G1P0 white woman at 22-week gestation with a monochorionic-diamniotic twin pregnancy discordant for esophageal atresia, duodenal atresia with gastric perforation, hypoplastic left heart structures, and significant early gestation maternal polyhydramnios. In this case, fetal magnetic resonance imaging was able to depict additional findings including area of gastric wall rupture, hiatal hernia, dilation of the distal esophagus, and area of duodenal obstruction and thus facilitated the proper diagnosis. After extensive counseling at our multidisciplinary team meeting, the parents elected to proceed with radiofrequency ablation of the anomalous twin to maximize the survival of the normal co-twin. The procedure was performed successfully with complete cessation of flow in the umbilical artery and complete cardiac standstill in the anomalous twin with no detrimental effects on the healthy co-twin.ConclusionsPrenatal diagnosis of complex anomalies in twin pregnancies constitutes a multitude of ethical, religious, and cultural factors that come into play in the management of these cases. Fetal magnetic resonance imaging provides detailed valuable information that can assist in management options including possible prenatal intervention. The combination of a cystic structure with peristalsis-like movement above the diaphragm (for example, “the upper thoracic pouch sign”), polyhydramnios, and progressive distention of the stomach and duodenum should increase suspicion for a combined pure esophageal and duodenal atresia.

Impact of Objective Echocardiographic Criteria for Timing of Congenital Diaphragmatic Hernia Repair

Objective To assess the impact of specific echocardiographic criteria for timing of congenital diaphragmatic hernia repair on the incidence of acute postoperative clinical decompensation from pulmonary hypertensive crisis and/or acute respiratory decompensation, with secondary outcomes including survival to discharge, duration of ventilator support, and length of hospitalization. Study design The multidisciplinary congenital diaphragmatic hernia management team instituted a protocol in 2012 requiring the specific criterion of echocardiogram‐estimated pulmonary artery pressure ≤80% systemic blood pressure before repairing congenital diaphragmatic hernias. A retrospective review of 77 neonatal patients with Bochdalek hernias repaired between 2008 and 2015 were reviewed: group 1 included patients repaired before protocol implementation (n = 25) and group 2 included patients repaired after implementation (n = 52). Results The groups had similar baseline characteristics. Postoperative decompensation occurred less often in group 2 compared with group 1 (17% vs 48%, P = .01). Adjusted analysis accounting for repair type, liver herniation, and prematurity yielded similar results (15% vs 37%, P = .04). Group 2 displayed a trend toward improved survival to 30 days postoperatively, though this did not reach statistical significance (94% vs 80%, P = .06). Patient survival to discharge, duration of ventilator support, and length of hospitalization were not different between groups. Conclusions The implementation of a protocol requiring echocardiogram‐estimated pulmonary arterial pressure ≤80% of systemic pressure before congenital diaphragmatic hernia repair may reduce the incidence of acute postoperative decompensation, although there was no difference in longer‐term secondary outcomes, including survival to discharge.

Use of Cerium Oxide Nanoparticles Conjugated with MicroRNA-146a to Correct the Diabetic Wound Healing Impairment

BACKGROUNDnDiabetic wounds have become one of the most challenging public health issues of the 21st century, yet there is no effective treatment available. We have previously shown that the diabetic wound healing impairment is associated with increased inflammation and decreased expression of the regulatory microRNA miR-146a. We have conjugated miR-146a to cerium oxide nanoparticles (CNP-miR146a) to target reactive oxygen species (ROS) and inflammation. This study aimed to evaluate the consequences of CNP-miR146a treatment of diabetic wounds.nnnSTUDY DESIGNnEight-millimeter wounds were created on the dorsal skin of Db/Db mice and treated with PBS or differing concentrations of CNP-mir146a (1; 10; 100; or 1,000 ng) at the time of wounding. Rate of wound closure was measured until the wounds were fully healed. At 4 weeks post-healing, a dumbbell-shaped skin sample was collected, with the healed wound in the center, and an Instron 5942 testing unit was used to measure the maximum load and modulus.nnnRESULTSnOur data showed that diabetic wounds treated with PBS or 1 ng CNP-miR146a took 18 days to heal. Treatment with 10, 100, or 1,000 ng of CNP+miR-146a effectively enhanced healing, and wounds were fully closed at day 14 post-wounding. The healed skin from the CNP-miR146a-treated group showed a trend of improved biomechanical properties (increased maximum load and modulus), however it did not reach significance.nnnCONCLUSIONSnWe found that a 100-ng dose of CNP-miR146a improved diabetic wound healing and did not impair the biomechanical properties of the skin post-healing. This nanotechnology-based therapy is promising, and future studies are warranted to transfer this therapy to clinical application.

Cardiac Progenitor Cell Recruitment Drives Fetal Cardiac Regeneration by Enhanced Angiogenesis

BACKGROUNDnIn contrast to adults, the fetal response to myocardial infarction (MI) is regenerative, requiring the recruitment of cardiac progenitor cells to replace infarcted myocardium. Macrophage contribution to tissue repair depends on their phenotype: M1 are proinflammatory and initiate angiogenesis; M2a are profibrotic and contribute to blood vessels maturation; and M2c are proremodeling and proangiogenesis. The goal of the present study was to expand on this work by examining cardiac progenitor cells recruitment, and the role of macrophages in promoting angiogenesis and cardiac regeneration in the fetal heart after MI.nnnMETHODSnFetal and adult sheep underwent MI and were sacrificed 3 or 30 days after MI. Some fetal hearts received stromal cell-derived factor-1α-inhibitor treatment. The microvasculature was evaluated by micro-computed tomography, gene expression was evaluated by real-time polymerase chain reaction, and vessels counts were evaluated by immunohistochemistry.nnnRESULTSnMicro-computed tomography analysis showed restoration of microvasculature in fetal hearts after MI. Vascular endothelial growth factor-α increased, and the expression of tissue markers associated with the M1, M2a, and M2c macrophage phenotypes were elevated at day 3 after MI, but returned to baseline by 30 days after MI. In contrast, adult hearts after MI exhibited low vascular endothelial growth factor-α and persistent upregulation of all macrophage markers, consistent with prolonged inflammation, fibrosis, and remodeling. Inhibition of stromal cell-derived factor-1α in fetal infarcts prevented angiogenesis, decreased vascular endothelial growth factor-α, and was associated with a sustained increase in M1, M2a, and M2c markers afterxa0MI.nnnCONCLUSIONSnChanges in angiogenesis and macrophage phenotype-related gene expression after MI are important for the fetal regenerative response to MI and are mediated at least in part by cardiac progenitor cells recruitment.

Modified Hysterotomy Closure Technique for Open Fetal Surgery

Objective: We reviewed our experience with open fetal surgical myelomeningocele repair to assess the efficacy of a new modification of the hysterotomy closure technique regarding hysterotomy complication rates at the time of cesarean delivery. Methods: A modification of the standard hysterotomy closure was performed on all patients undergoing prenatal myelomeningocele repair. The closure consisted of an interrupted full-thickness #0 polydioxanone (PDS) retention suture as well as a running #0 PDS suture to re-approximate the myometrial edges, and the modification was a third imbricating layer resulting in serosal-to-serosal apposition. A standard omental patch was placed per our routine. Both operative reports and verbal descriptions of hysterotomy from delivering obstetricians were reviewed. Results: A total of 49 patients underwent prenatal repair of myelomeningocele, 43 having adequate follow-up for evaluation. Of those, 95.4% had completely intact hysterotomy closures, with only 1 partial dehiscence (2.3%) and 1 thinned scar (2.3%). There were no instances of uterine rupture. Discussion: In patients undergoing this modified hysterotomy closure technique, a much lower than expected complication rate was observed. This simple modified closure technique may improve hysterotomy healing and reduce obstetric morbidity.

Case report of myometrial window following fetoscopic treatment of twin-twin transfusion syndrome: indications of underlying collagen vascular disease?

Intraperitoneal amniotic fluid leak is a known complication of fetoscopic procedures that usually resolves spontaneously with expectant management. Intraperitoneal amniotic fluid leak may persist after fetoscopic procedures due to a myometrial window as well as to persistent chorioamniotic membrane disruption, which may be amenable to surgical repair.