Cross-Generational Reproductive Fitness Enforced by Microchimeric Maternal Cells.

Exposure to maternal tissue during in utero development imprints tolerance to immunologically foreign non-inherited maternal antigens (NIMA) that persists into adulthood. The biological advantage of this tolerance, conserved across mammalian species, remains unclear. Here, we show maternal cells that establish microchimerism in female offspring during development promote systemic accumulation of immune suppressive regulatory T cells (Tregs) with NIMA specificity. NIMA-specific Tregs expand during pregnancies sired by males expressing alloantigens with overlapping NIMA specificity, thereby averting fetal wastage triggered by prenatal infection and non-infectious disruptions of fetal tolerance. Therefore, exposure to NIMA selectively enhances reproductive success in second-generation females carrying embryos with overlapping paternally inherited antigens. These findings demonstrate that genetic fitness, canonically thought to be restricted to Mendelian inheritance, is enhanced in female placental mammals through vertically transferred maternal cells that promote conservation of NIMA and enforce cross-generational reproductive benefits.

Fetal bradycardia in open versus fetoscopic prenatal repair of spina bifida.

OBJECTIVE: To compare the occurrence of fetal bradycardia in open versus fetoscopic fetal spina bifida surgery. METHODS: This is a single-institution retrospective cohort study of patients undergoing open (n = 25) or fetoscopic (n = 26) spina bifida repair between 2017 and 2022. From October 2017 to June 2020, spina bifida repairs were performed via an open classical hysterotomy, and from November 2020 to June 2022 fetoscopic repairs were performed following transition to this technique. Fetal heart rate (FHR) in beats per minute (bpm) was recorded via echocardiography every 15 min during the procedure. Cohort characteristics, fetal bradycardia and maternal physiologic parameters were compared between the groups. RESULTS: Fetuses undergoing an open repair more frequently developed bradycardia defined as <110 bpm (32% vs. 3.8%, p = 0.008), and a trend was observed for FHR decreases more than 25 bpm from baseline (20% vs. 3.8%, p = 0.073). Profound bradycardia less than 80 bpm was rare, occurring in only three operations (two in open, one in fetoscopic repair) with two fetuses (one in each group) requiring emergency cesarean delivery. CONCLUSION: When compared to open fetal surgery, fetal bradycardia occurred less frequently in fetoscopic surgery despite a significantly greater anesthetic exposure and the use of the intraamniotic carbon dioxide insufflation.

Needle Fetal Thoracoscopy: A Technique to Assist with Ultrasound-Guided Placement of Challenging Thoracoamniotic Shunts.

INTRODUCTION: Fetal thoracoamniotic shunts are common lifesaving interventions but frequently require replacement. Needle fetal thoracoscopy is a technique that uses standard thoracoamniotic shunt introducer sheaths to permit direct visualization and even instrument manipulation during shunt deployment to facilitate optimal positioning and primary shunt function in the most challenging cases. CASE PRESENTATION: In this study, 5 patients who underwent needle fetal thoracoscopy-assisted thoracoamniotic shunt placement were reviewed. Three patients with large, macrocystic congenital pulmonary airway malformations (CPAMs) with evidence of worsening mediastinal shift and/or hydrops and 2 patients with large chylothorax with fetal hydrops were treated. Four cases had previous shunts that failed due to poor sonographic visualization during initial placement, cyst septations, shunt obstruction, or dislodgment. Needle fetal thoracoscopy was used to disrupt cyst walls and septations, clear hematoma, and confirm the optimal initial position of the shunt. In this series, 1 severe CPAM patient with a short cervix developed preterm labor postoperatively resulting in neonatal demise. The remaining 4 patients experienced resolution of hydrops and progressed to successful delivery with excellent neonatal outcomes. CONCLUSION: Needle fetal thoracoscopy is a procedure that may be selectively deployed in challenging thoracoamniotic shunt cases impacted by recurrent failure, poor sonographic windows, and challenging fetal positioning.

Nonparasitic Splenic Cysts in Children: Outcomes after Sclerotherapy or Surgical Resection as Initial Management.

Management of splenic cysts in children remains undefined. Sclerotherapy is an innovative, less invasive treatment. This study examined the safety and preliminary effectiveness of sclerotherapy for splenic cysts in children compared with those of surgical treatment. A retrospective review of pediatric patients treated for nonparasitic splenic cysts from 2007 to 2021 was performed at a single institution. Posttreatment outcomes for patients who underwent either expectant management, sclerotherapy, or surgery were reviewed. Thirty patients aged between 0 and 18 years met the inclusion criteria. Cysts in 3 of 8 patients who underwent sclerotherapy were either unresolved or recurred. Patients who underwent sclerotherapy and required surgery for residual symptomatic cyst had an initial cyst diameter of >8 cm. Symptoms resolved in 5 of 8 patients who underwent sclerotherapy, with a significantly reduced cyst size compared with that in patients with continued symptoms who underwent sclerotherapy (61.4% vs 7.0%, P = .01). Sclerotherapy is an effective treatment for splenic cysts, particularly those measuring <8 cm. However, surgical excision may be preferable for large cysts.

Selective Fetoscopic Laser Photocoagulation or Expectant Management for Stage I Twin-Twin Transfusion: A Cost-Effectiveness Analysis.

BACKGROUND: Selective fetoscopic laser photocoagulation (SFLP) is the preferred intervention for stage II-IV twin-twin transfusion syndrome (TTTS); however, there is no consensus on whether SFLP or expectant management (EM) is the preferred strategy to manage Quintero stage I TTTS. OBJECTIVE: The objective of this study is to estimate whether SFLP or EM is the cost-effective strategy for management of Quintero stage I TTTS. STUDY DESIGN: A decision-analysis (DA) model compared SFLP to EM for 1,000 pregnant people with monochorionic-diamniotic twins affected by stage I TTTS. All subjects were assumed to be appropriate candidates for either SFLP or EM. Probabilities, costs, and utilities were derived from the literature. The DA was conducted from a healthcare payor perspective, and the analytic horizon was over the course of an offspring's lifetime, with primary outcomes of survivorship (i.e., no intrauterine fetal demise or neonatal death) and long-term neurodevelopmental impairment. The model incorporated Markov processes with 4-week cycles throughout pregnancy. Incremental cost-effectiveness ratios (ICER) for each strategy were calculated and compared to estimate marginal cost effectiveness. An ICER of USD 100,000 per quality-adjusted life year was used to define the cost-effectiveness threshold. One-way sensitivity and Monte Carlo analyses (MCA), as well as microsimulations, were performed. RESULTS: For base-case estimates, SFLP was found to be cost-effective compared to EM in the management of stage I TTTS. In one-way sensitivity analysis, varying each variable along pre-specified ranges did not result in changes in the conclusion. MCA projects SFLP as the cost-effective strategy in 100% of runs. CONCLUSIONS: With base-case estimates, SFLP is estimated to be the cost-effective strategy for the treatment of Quintero stage I TTTS when compared with EM. This remained true across a wide range of inputs.

Immunosuppressive CD71+ erythroid cells compromise neonatal host defence against infection.

Newborn infants are highly susceptible to infection. This defect in host defence has generally been ascribed to the immaturity of neonatal immune cells; however, the degree of hyporesponsiveness is highly variable and depends on the stimulation conditions. These discordant responses illustrate the need for a more unified explanation for why immunity is compromised in neonates. Here we show that physiologically enriched CD71(+) erythroid cells in neonatal mice and human cord blood have distinctive immunosuppressive properties. The production of innate immune protective cytokines by adult cells is diminished after transfer to neonatal mice or after co-culture with neonatal splenocytes. Neonatal CD71(+) cells express the enzyme arginase-2, and arginase activity is essential for the immunosuppressive properties of these cells because molecular inhibition of this enzyme or supplementation with L-arginine overrides immunosuppression. In addition, the ablation of CD71(+) cells in neonatal mice, or the decline in number of these cells as postnatal development progresses parallels the loss of suppression, and restored resistance to the perinatal pathogens Listeria monocytogenes and Escherichia coli. However, CD71(+) cell-mediated susceptibility to infection is counterbalanced by CD71(+) cell-mediated protection against aberrant immune cell activation in the intestine, where colonization with commensal microorganisms occurs swiftly after parturition. Conversely, circumventing such colonization by using antimicrobials or gnotobiotic germ-free mice overrides these protective benefits. Thus, CD71(+) cells quench the excessive inflammation induced by abrupt colonization with commensal microorganisms after parturition. This finding challenges the idea that the susceptibility of neonates to infection reflects immune-cell-intrinsic defects and instead highlights processes that are developmentally more essential and inadvertently mitigate innate immune protection. We anticipate that these results will spark renewed investigation into the need for immunosuppression in neonates, as well as improved strategies for augmenting host defence in this vulnerable population.

Prenatal Allogeneic Tolerance in Mice Remains Stable Despite Potent Viral Immune Activation.

Transplanting stem cells before birth offers an unparalleled opportunity to initiate corrective treatment for numerous childhood diseases with minimal or no host conditioning. Although long-term engraftment has been demonstrated following in utero hematopoietic cellular transplantation during immune quiescence, it is unclear if prenatal tolerance becomes unstable with immune activation such as during a viral syndrome. Using a murine model of in utero hematopoietic cellular transplantation, the impact of an infection with lymphocytic choriomeningitis virus on prenatal allospecific tolerance was examined. The findings in this report illustrate that established mechanisms of donor-specific tolerance are strained during potent immune activation. Specifically, a transient reversal in the anergy of alloreactive lymphocytes is seen in parallel with the global immune response toward the virus. However, these changes return to baseline following resolution of the infection. Importantly, prenatal engraftment remains stable during and after immune activation. Collectively, these findings illustrate the robust nature of allospecific tolerance in prenatal mixed chimerism compared with models of postnatal chimerism and provides additional support for the prenatal approach to the treatment of congenital benign cellular disease.

Prenatal Allospecific NK Cell Tolerance Hinges on Instructive Allorecognition through the Activating Receptor during Development.

Little is known about how the prenatal interaction between NK cells and alloantigens shapes the developing NK cell repertoire toward tolerance or immunity. Specifically, the effect on NK cell education arising from developmental corecognition of alloantigens by activating and inhibitory receptors with shared specificity is uncharacterized. Using a murine prenatal transplantation model, we examined the manner in which this seemingly conflicting input affects NK cell licensing and repertoire formation in mixed hematopoietic chimeras. We found that prenatal NK cell tolerance arose from the elimination of phenotypically hostile NK cells that express an allospecific activating receptor without coexpressing any allospecific inhibitory receptors. Importantly, the checkpoint for the system appeared to occur centrally within the bone marrow during the final stage of NK cell maturation and hinged on the instructive recognition of allogeneic ligand by the activating receptor rather than through the inhibitory receptor as classically proposed. Residual nondeleted hostile NK cells expressing only the activating receptor exhibited an immature, anergic phenotype, but retained the capacity to upregulate inhibitory receptor expression in peripheral sites. However, the potential for this adaptive change to occur was lost in developmentally mature chimeras. Collectively, these findings illuminate the intrinsic process in which developmental allorecognition through the activating receptor regulates the emergence of durable NK cell tolerance and establishes a new paradigm to fundamentally guide future investigations of prenatal NK cell-allospecific education.

Management of fetal teratomas.

Fetal teratomas are the most common tumors diagnosed prenatally. The majority of these tumors are benign and cured by complete resection of the mass during the neonatal period. Prenatal diagnosis has improved the perinatal management of these lesions and especially for the teratomas that might benefit from fetal intervention. A comprehensive prenatal evaluation including conventional ultrasounds, Doppler, echocardiography and fetal MRI, is essential for an effective counseling and perinatal management. Antenatal counseling helps the parents to better understand the natural history, fetal intervention, and perinatal management of these tumors, which differ dramatically depending on their size and location. Fetal surgical debulking improves survival in cases of sacrococcygeal teratoma with cardiac decompensation. Additionally, the use of an EXIT procedure reduces the morbidity and mortality if a complicated delivery in cases of cervical and mediastinal teratomas. Here, we offer an overview of all fetal teratomas and their recommended management, with emphasis on in utero treatment options.

Fetal lung growth represented by longitudinal changes in MRI-derived fetal lung volume parameters predicts survival in isolated left-sided congenital diaphragmatic hernia.

OBJECTIVE: The aim of this study was to evaluate fetal lung growth rate for isolated left-sided congenital diaphragmatic hernia (CDH) using serial magnetic resonance imaging (MRI)-based volumetric measures. METHODS: Early and late gestational (22-30 and >30 weeks' gestation) lung volumetry was obtained by fetal MRI in 47 cases of isolated left-sided CDH. At both of these time points, lung volume indices, including total lung volume (TLV), observed to expected TLV (o/e TLV), and percentage of predicted lung volume (PPLV) as well as their change rates (Δ) and relative Δ during gestation were calculated and analyzed in regard to their capacity to predict neonatal survival. RESULTS: TLV, o/e TLV, and PPLV had various changes during gestation. Late TLV, early and late o/e TLV, and late PPLV were predictive of neonatal survival. Non-survivors had lower ΔTLV and more negative relative ΔPPLV than survivors (1.18 vs 1.85 mL/week, P = 0.004 and -4.15%/week vs -1.95%/week, P = 0.002, respectively). CONCLUSIONS: The severity of pulmonary hypoplasia is dynamic and can worsen in the third trimester. MRI lung volumetry repeated in late gestation can provide additional information on individual lung growth that may facilitate prenatal counseling and focus perinatal management.

Expanded intrathoracic space in fetal cases of isolated congenital diaphragmatic hernia contributes to disparity between percent predicted lung volume and observed to expected total lung volume.

OBJECTIVE: The aim of this study was to determine whether fetal lung volume and visceral herniation are associated with changes in intrathoracic space in congenital diaphragmatic hernia(CDH). METHODS: We retrospectively examined the relationship between magnetic resonance imaging-derived measurements of intrathoracic space [predicted lung volume (PLV)] and residual lung volume or visceral herniation among isolated left-sided CDH fetuses. RESULTS: Data from fetal magnetic resonance imaging studies of 60 isolated left-sided CDH cases were analyzed. The median PLV of the CDH fetuses was found to be much greater than the expected total lung volume (eTLV) of a normal fetus at the same gestational age. Surprisingly, liver herniation and observed TLV(oTLV) were positively correlated with the PLV. Although the PPLV was consistently less than the o/eTLV, both indices were greater in survivors than in non-survivors, whereas no significant difference was seen in the PLV/eTLV ratio in regard to survivorship. CONCLUSION: The intrathoracic domain available for lungs and viscera is expanded in CDH fetuses and positively affected by the lung volume and the presence of liver herniation, leading to the difference in the PPLV and o/eTLV. Future study of intrathoracic space as it relates to the growth of the lung and herniated viscera is needed to better characterize the relationship between these parameters.

Differential patterns of prenatal ipsilateral and contralateral lung growth in cases of isolated left-sided congenital diaphragmatic hernia.

OBJECTIVE: The aim of this research was to compare the impact of varying degrees of visceral herniation on the growth rates of the contralateral and ipsilateral fetal lungs in cases of isolated left-sided congenital diaphragmatic hernia (CDH). METHODS: Data were retrieved from 58 fetuses with isolated left-sided CDH undergoing magnetic resonance imaging studies at both mid-gestation (20-30 weeks) and late-gestation (>30 weeks) time points. The growth of the right and left lungs (ΔLV-R and ΔLV-L) was calculated. The impact of the degree of visceral herniation on the growth disparity between the right and left lungs was then compared. RESULTS: Measurable growth occurred in both lungs between the mid-gestation and late-gestation time points in each group. The ΔLV-R exhibited a strong correlation with ΔLV-L. However, the right lung grew significantly faster than the left lung (ΔLV-R = 1.36 vs ΔLV-L = 0.17 mL/week, P < 0.001). A higher degree of visceral herniation appeared to decrease the growth rate disparity by progressive limitation of the growth of the right lung. CONCLUSION: The contralateral lung retains the potential to grow faster than the ipsilateral lung during the third trimester. A higher degree of visceral herniation places progressive limitations on contralateral lung growth thereby diminishing the growth rate disparity between the right and left lungs.

Comparison of interleukin 10 homologs on dermal wound healing using a novel human skin ex vivo organ culture model.

BACKGROUND: Anti-inflammatory cytokine interleukin (IL)-10 has been shown to induce regenerative healing in postnatal wounds. A viral homolog of IL-10 produced by human cytomegalovirus (CMV IL-10) similarly generates potent immunoregulatory effects, but its effects on wound healing have not been investigated. Currently, there are limited cost-effective methods of screening vulnerary therapeutics. Taken together, we aim to develop and validate a novel human ex vivo dermal wound model and hypothesize that CMV IL-10 will enhance dermal wound healing. METHODS: Full-thickness circular (6-mm) explants were taken from surgical skin samples and 3-mm full-thickness wounds were created. Explants were embedded in collagen I matrix and maintained in specially formulated media with the epidermis at air-liquid interface, and treated with human IL-10 or CMV IL-10 (200 ng/mL). The viability of cultured explants was validated by histology and lactate dehydrogenase (LDH) activity. Epithelial gap, epithelial height, basal keratinocyte migration, vascular endothelial growth factor levels, and neovascularization were measured at days 3 and 7 to determine IL-10 effects on wound healing. RESULTS: Culture explants at day 7 appeared similar to fresh skin in morphology, cell, and vessel density. By day 14, the epidermis separated from the dermis and the cell density diminished. Day 7 wounds appeared viable with advancing epithelial and basal keratinocyte migration with no evidence of necrosis. Cytotoxicity analysis via the quantification of LDH revealed no differences between controls and treated groups. There was a slight increase in the quantity of LDH in media at day 3; however, this decreased at day 5 and continued to decline up to day 21. CMV IL-10 treatment resulted in a significant decrease in the epithelial gap and an increase in epithelial height. There were no differences in the rates of basal keratinocyte migration at day 7 between treated and control groups. Interestingly, human IL-10 increased vascular endothelial growth factor expression and neovascularization compared with controls. CONCLUSIONS: The human ex vivo wound model provides a simple and viable design to study dermal wound healing. Both IL-10 homologs demonstrate vulnerary effects. The viral homolog demonstrates enhanced effects on wound closure compared with human IL-10. These data represent a novel tool that can be used to screen therapeutics, such as CMV IL-10, before preclinical studies.

Adenoviral-mediated gene transfer of insulin-like growth factor 1 enhances wound healing and induces angiogenesis.

BACKGROUND: Chronic wounds are characterized by a wound healing and neovascularization deficit. Strategies to increase neovascularization can significantly improve chronic wound healing. Insulin-like growth factor (IGF)-1 is reported to be a keratinocyte mitogen and is believed to induce angiogenesis via a vascular endothelial growth factor (VEGF)-dependent pathway. Using a novel ex vivo human dermal wound model and a diabetic-impaired wound healing murine model, we hypothesized that adenoviral overexpression of IGF-1 (Ad-IGF-1) will enhance wound healing and induce angiogenesis through a VEGF-dependent pathway. METHODS: Ex vivo: 6-mm full-thickness punch biopsies were obtained from normal human skin, and 3-mm full-thickness wounds were created at the center. Skin explants were maintained at air liquid interface. Db/db murine model: 8-mm full-thickness dorsal wounds in diabetic (db/db) mice were created. Treatment groups in both human ex vivo and in vivo db/db wound models include 1×10(8) particle forming units of Ad-IGF-1 or Ad-LacZ, and phosphate buffered saline (n=4-5/group). Cytotoxicity (lactate dehydrogenase) was quantified at days 3, 5, and 7 for the human ex vivo wound model. Epithelial gap closure (hematoxylin and eosin; Trichrome), VEGF expression (enzyme-linked immunosorbent assay), and capillary density (CD 31+CAPS/HPF) were analyzed at day 7. RESULTS: In the human ex vivo organ culture, the adenoviral vectors did not demonstrate any significant difference in cytotoxicity compared with phosphate buffered saline. Ad-IGF-1 overexpression significantly increases basal keratinocyte migration, with no significant effect on epithelial gap closure. There was a significant increase in capillary density in the Ad-IGF-1 wounds. However, there was no effect on VEGF levels in Ad-IGF-1 samples compared with controls. In db/db wounds, Ad-IGF-1 overexpression significantly improves epithelial gap closure and granulation tissue with a dense cellular infiltrate compared with controls. Ad-IGF-1 also increases capillary density, again with no significant difference in VEGF levels in the wounds compared with control treatments. CONCLUSIONS: In two different models, our data demonstrate that adenoviral-mediated gene transfer of IGF-1 results in enhanced wound healing and induces angiogenesis via a VEGF-independent pathway. Understanding the underlying mechanisms of IGF-1 effects on angiogenesis may help produce novel therapeutics for chronic wounds or diseases characterized by a deficit in neovascularization.

Pseudotyped adeno-associated viral vectors for gene transfer in dermal fibroblasts: implications for wound-healing applications.

BACKGROUND: Cell-specific gene transfer and sustained transgene expression are goals of cutaneous gene therapy. Pseudotyping strategy with adeno-associated viral (AAV) vectors has the potential to confer unique cellular tropism and transduction efficiency. We hypothesize that pseudotyped AAV vectors have differential tropism and transduction efficiency under normal and wound conditions in dermal fibroblasts. MATERIALS AND METHODS: We packaged AAV2 genome with green fluorescent protein reporter in capsids of other serotypes, AAV5, AAV7, and AAV8, producing pseudotyped vectors AAV2/5, AAV2/7, and AAV2/8, respectively. Murine and human dermal fibroblasts were transduced by the different pseudotypes for 24 h at multiplicities of infection 10(2), 10(3), 10(4), and 10(5). We assessed transduction efficiency at days 3 and 7. Experiments were repeated in a simulated wound environment by adding 10 ng/mL platelet-derived growth factor-B to culture media. RESULTS: Transduction efficiency of the pseudotyped AAV vectors was dose dependent. Multiplicity of infection 10(5) resulted in significantly higher gene transfer. Under normal culture conditions, the pseudotyping strategy conferred differential transduction of dermal fibroblasts, with significantly enhanced transduction of murine cells by AAV2/5 and AAV2/8 compared with AAV2/2. Adeno-associated virus 2/8 was more efficacious in transducing human cells. Under wound conditions, transduction efficiency of AAV2/2, 2/5, and 2/8 was significantly lower in murine fibroblasts. At day 3 under wound conditions, all vectors demonstrated similar transduction efficiency, but by day 7, the three pseudotyped vectors transduced significantly more murine cells compared with AAV2/2. However, in human cells, there was no significant difference in the transduction efficiency of each pseudotype between normal and wound conditions at both 3 and 7 d. CONCLUSIONS: The AAV pseudotyping strategy represents a gene transfer technology that can result in differential transduction of dermal fibroblasts. The differences in transduction efficiency in murine and human dermal fibroblasts in both the normal and wound environment highlight issues with translatability of gene transfer techniques. These data provide a template for using pseudotyped AAV vectors in cutaneous applications.

The pediatric surgeon's road to research independence: utility of mentor-based National Institutes of Health grants.

BACKGROUND: The current research environment for academic surgeons demands that extramural funding be obtained. Financial support from the National Institutes of Health (NIH) is historically the gold standard for funding in the biomedical research community, with the R01 funding mechanism viewed as indicator of research independence. The NIH also supports a mentor-based career development mechanism (K-series awards) in order to support early-stage investigators. The goal of this study was to investigate the grants successfully awarded to pediatric surgeon-scientists and then determine the success of the K-series award recipients at achieving research independence. METHODS: In July 2012, all current members of the American Pediatric Surgery Association (APSA) were queried in the NIH database from 1988-2012 through the NIH Research Portfolio Online Reporting Tools. The following factors were analyzed: type of grant, institution, amount of funding, and funding institute or center. RESULTS: Among current APSA members, there have been 83 independent investigators receiving grants, representing 13% of the current APSA membership, with 171 independent grants funded through various mechanisms. Six percent currently have active NIH funding, with $7.2 million distributed in 2012. There have been 28 K-series grants awarded. Of the recipients of expired K08 awards, 39% recipients were subsequently awarded an R01 grant. A total of 63% of these K-awarded investigators transitioned to an independent NIH award mechanism. CONCLUSIONS: Pediatric surgeon-scientists successfully compete for NIH funding. Our data suggest that although the K-series funding mechanism is not the only path to research independence, over half of the pediatric surgeons who receive a K-award are successful in the transition to independent investigator.

Interleukin-10 regulates the fetal hyaluronan-rich extracellular matrix via a STAT3-dependent mechanism.

BACKGROUND: The midgestational fetus is capable of regenerative healing. We have recently demonstrated a novel role for the anti-inflammatory cytokine interleukin 10 (IL-10) as a regulator of hyaluronan (HA) in the extracellular matrix. The signaling pathway of IL-10 has been studied in monocytes but is unknown in dermal fibroblasts. We hypothesized IL-10 signals through its primary receptor, IL-10R1, to activate STAT3, resulting in HA synthesis. METHODS: Murine midgestational (E14.5) fetal fibroblasts were evaluated in vitro. Pericellular matrix was quantified using a particle exclusion assay. STAT3 levels and cellular localization were evaluated by Western blot/band densitometry and immunocytochemistry/confocal microscopy. HA levels were quantified by enzyme-linked immunosorbent assay. The effects of IL-10R1 signal blockade by a neutralizing antibody and STAT3 inhibition were evaluated. An ex vivo midgestation fetal forearm culture incisional wound model in control and transgenic IL-10-/- mice was used to evaluate the role of STAT3 on the extracellular matrix. RESULTS: Fetal fibroblasts produce a robust hyaluronan-rich pericellular matrix that is IL-10R1 and STAT3 dependent. Inhibition of IL-10R1 signaling results in decreased phosphorylated STAT3 levels and inhibition of nuclear localization. Inhibition of STAT3 results in decreased HA production. At day 3, midgestation fetal wounds have efficient re-epithelialization, which is significantly slowed in IL-10-/- wounds at the same gestation and with inhibition of STAT3. CONCLUSIONS: Our data demonstrate that IL-10 regulates HA synthesis through its primary receptor IL-10R1 and STAT3 activation. This supports a novel nonimmunoregulatory mechanism of IL-10 in its role in fetal regenerative wound healing.

Ex-Utero Intrapartum Treatment-to-Airway for Obstructing Fetal Neck Masses: A Singular Methodology for Monochorionic and Dichorionic Twin Pregnancies.

Fetal airway obstruction in one twin of a diamniotic pregnancy presents unique challenges. Very few cases of ex-utero-intrapartum-treatment (EXIT) procedures for twin pregnancy have been reported and only in dichorionic pregnancies. We report a singular methodology for EXIT-to-airway procedures in two pregnancies involving monochorionic and dichorionic twins. Two cases of EXIT-to-airway in twin pregnancies were performed in 2018 and 2019 at a regional fetal treatment center. Case 1 involved a giant cervical teratoma in a monochorionic-diamniotic twin pregnancy with preterm labor at 29 weeks. Case 2 involved a dichorionic-diamniotic pregnancy with a large cervical lymphatic malformation with preterm labor at 36 weeks. In each case, the polyhydramnios caused the affected twin's amniotic sac to be the presenting sac for the surgical approach. Bronchoscopy and successful intubation was completed after 22 and 10 minutes of uteroplacental bypass, respectively. The bystander twins were delivered second without intubation and resuscitated without perinatal distress. EXIT-to-airway appears to be a reasonable option for twins including monochorionic pregnancies, via delivery of the affected twin first followed by delivery of the bystander twin. Thoughtful preparation and counseling by an experienced multidisciplinary team permits an EXIT-to-airway approach for twin pregnancies even in an emergent setting.

CD4 and IL-2 mediated NK cell responses after COVID-19 infection and mRNA vaccination in adults.

A detailed understanding of protective immunity against SARS-CoV-2 is incredibly important in fighting the pandemic. Central to protective immunity is the ability of the immune system to recall previous exposures. Although antibody and T cell immunity have gained considerable attention, the contribution of the NK cell compartment to immune recall and protection from SARS-CoV-2 has not been explored. In this study, we investigate the NK cell responses to stimulation with SARS-CoV-2 in previously exposed and non-exposed individuals. We show that NK cells demonstrate an enhanced CD4+ T cell dependent response when re-exposed to SARS-CoV-2 antigen. The enhanced response is dependent on T cells and correlates with the number of SARS-CoV-2 specific CD4 T cells. We find that IL-2 is a critical mediator of NK cell function. These findings suggest that NK cells contribute to the protective responses against SARS-CoV-2 through a cooperation with antigen-specific CD4 T cells and have significant implications on our understanding of protective immunity in SARS-CoV-2.

The interplay between prenatal liver growth and lung development in congenital diaphragmatic hernia.

Objective: Liver herniation is a known risk factor for increased severity in CDH and is associated with clinically significant pulmonary hypoplasia and pulmonary hypertension. Better studies are needed to understand the growth of the herniated liver compared to the liver that remains in the abdomen and how this liver growth then affects lung development. Serial hi-resolution fetal MRI enables characterization of liver growth throughout gestation and examination of macroscopic features that may regulate liver growth. Here, we hypothesized that the nature of liver herniation affects liver growth and, in turn, affects lung growth. Methods: Clinical data were retrospectively collected from consecutive cases of prenatally diagnosed isolated left-sided or right-sided CDH from June 2006 to August 2021. Only those cases with MRI lung volumetry for both mid-gestation and late-gestation time points were recruited for analysis. Cases with fetal chromosomal abnormalities and other major structural abnormalities were excluded. Fractional liver volume and liver growth was indexed to estimated fetal weight and compared to lung growth. Results: Data was collected from 28 fetuses with a left liver-down CDH (LLD), 37 left liver-up CDH (LLU) and 9 right liver-up CDH (RLU). Overall, RLU fetuses had greater overall and fractional (intra-thoracic vs. intra-abdominal) liver growth when compared to LLD and LLU fetuses. Additionally, intra-thoracic liver growth was consistently slower than intra-abdominal liver growth for either right- or left-sided CDH. When the liver was not herniated, a positive correlation was seen between liver growth and lung growth. However, when the liver was herniated above the diaphragm, this positive correlation was lost. Conclusion: Right-sided CDH fetuses exhibit greater liver growth compared to left-sided CDH. Liver herniation disrupts the normal positive correlation between liver and lung growth that is seen when the liver is entirely within the abdomen.