Outpatient Factors Implicated in Timing of Delayed Elective Inguinal Hernia Repair in Premature Neonates: Single Center Analysis Brings Up a Touchy Issue and Balancing Metric.

INTRODUCTION: Among premature infants, the incidence of inguinal hernias (IH) has been reported to be as high as 10-30%. We performed this study to characterize the association between individual and systemic variables that may affect diagnosis to definitive operative repair of the premature neonatal IH in the outpatient setting. METHODS: A single center cohort retrospective review analyzing IH repair in the premature neonatal (<37 GA) population was performed. Data was collected between 2013 and 2022. The cohort was defined as patients who underwent repair before the age of 1 and excluded patients with major medical comorbidities or underwent simultaneous major abdominal surgeries. RESULTS: Of the 836 premature neonates who underwent IH repair, the majority (73%) were repaired electively. Patients were characterized into risk cohorts a-priori. High-risk patients (HR, n = 43) were more likely to have Government insurance (67%). There was a significant difference in HR patient time to surgery between Government versus Commercial insurance, 10.6 versus 4.7 days, respectively (95% CI -11.09 to -0.4396, p = 0.0345). HR patients were also seen more frequently (clinic or emergency department) prior to operative repair (2.51 vs 1.72 95% CI -1.296% to -0.289%, p = 0.0021). A multivariate linear regression model demonstrated that risk class (p = 0.0244), touches (p < 0.0001), GA (p < 0.001), and prior authorization (p < 0.0001) were significantly associated with time to hernia repair. CONCLUSIONS: Systemic variables such as insurance type may increase average wait time for elective outpatient IH repair. This increase in wait time is associated with an increased number of healthcare visits. Therefore, timely access to surgical care prevents potential harm and may decrease health system burden. TYPE OF STUDY: Retrospective comparative study/cohort study. LEVEL OF EVIDENCE: III.

In utero delivery of targeted ionizable lipid nanoparticles facilitates in vivo gene editing of hematopoietic stem cells.

Monogenic blood diseases are among the most common genetic disorders worldwide. These diseases result in significant pediatric and adult morbidity, and some can result in death prior to birth. Novel ex vivo hematopoietic stem cell (HSC) gene editing therapies hold tremendous promise to alter the therapeutic landscape but are not without potential limitations. In vivo gene editing therapies offer a potentially safer and more accessible treatment for these diseases but are hindered by a lack of delivery vectors targeting HSCs, which reside in the difficult-to-access bone marrow niche. Here, we propose that this biological barrier can be overcome by taking advantage of HSC residence in the easily accessible liver during fetal development. To facilitate the delivery of gene editing cargo to fetal HSCs, we developed an ionizable lipid nanoparticle (LNP) platform targeting the CD45 receptor on the surface of HSCs. After validating that targeted LNPs improved messenger ribonucleic acid (mRNA) delivery to hematopoietic lineage cells via a CD45-specific mechanism in vitro, we demonstrated that this platform mediated safe, potent, and long-term gene modulation of HSCs in vivo in multiple mouse models. We further optimized this LNP platform in vitro to encapsulate and deliver CRISPR-based nucleic acid cargos. Finally, we showed that optimized and targeted LNPs enhanced gene editing at a proof-of-concept locus in fetal HSCs after a single in utero intravenous injection. By targeting HSCs in vivo during fetal development, our Systematically optimized Targeted Editing Machinery (STEM) LNPs may provide a translatable strategy to treat monogenic blood diseases before birth.

Preexisting maternal immunity to AAV but not Cas9 impairs in utero gene editing in mice.

In utero gene editing (IUGE) is a potential treatment for inherited diseases that cause pathology before or soon after birth. Preexisting immunity to adeno-associated virus (AAV) vectors and Cas9 endonuclease may limit postnatal gene editing. The tolerogenic fetal immune system minimizes a fetal immune barrier to IUGE. However, the ability of maternal immunity to limit fetal gene editing remains a question. We investigated whether preexisting maternal immunity to AAV or Cas9 impairs IUGE. Using a combination of fluorescent reporter mice and a murine model of a metabolic liver disease, we demonstrated that maternal anti-AAV IgG antibodies were efficiently transferred from dam to fetus and impaired IUGE in a maternal titer-dependent fashion. By contrast, maternal cellular immunity was inefficiently transferred to the fetus, and neither maternal cellular nor humoral immunity to Cas9 impaired IUGE. Using human umbilical cord and maternal blood samples collected from mid- to late-gestation pregnancies, we demonstrated that maternal-fetal transmission of anti-AAV IgG was inefficient in midgestation compared with term, suggesting that the maternal immune barrier to clinical IUGE would be less relevant at midgestation. These findings support immunologic advantages for IUGE and inform maternal preprocedural testing protocols and exclusion criteria for future clinical trials.

Emergent Total Endovascular Arch Repair for Contained Aortic Arch Rupture: Another Tool in the Box.

To date, emergent total endovascular aortic arch repair has not been described in the literature. We present a 67-year-old female with a poorly differentiated posterior mediastinal sarcoma. Imaging obtained was concerning for intravascular extension of the tumor into the thoracic aorta. While awaiting radiation therapy, the patient complained of worsening chest and arm pain, vital signs demonstrating tachypnea and hypoxia. Subsequent imaging revealed an increase in vascular erosion, concerning for a contained rupture, with complete obliteration of the left mainstem bronchus. The patient was emergently taken for percutaneous endovascular repair of her aortic arch. A three-vessel physician modified fenestrated graft was created and deployed with concurrent stenting of the innominate, left carotid, and left subclavian arteries. Interval computed tomography angiography revealed patency in all stented vessels, with no endoleak and no evidence of pseudoaneurysm. The patient was able to undergo chemotherapy with favorable decrease in tumor burden. Total endovascular aortic arch repair, when planned carefully, is an attractive option in high-risk patients who are otherwise not ideally suited for open total arch replacement.

A case report of necrotizing fasciitis with growth of Actinomyces europaeus and Actinotignum schaalii.

Actinomyces europeaeus and Actinotignum schaalii are two facultative anaerobes that are common contaminants of human flora; namely the urinary tract, the female genital tract and the gastrointestinal tract. A. europeaeus has been linked with abscesses, decubitus ulcers and purulent urethritis, while A. schaalii has been associated with urinary tract infections, bacteremia and Fournier's gangrene. Here we present a case report of an 84-year-old female patient found to have a necrotizing soft tissue infection caused by A. europeaeus and A. schaalii. To our knowledge, this is the first case report that documents A. europeaeus as a causal agent of a necrotizing infection.

Membrane-associated human tyrosinase is an enzymatically active monomeric glycoprotein.

Human tyrosinase (hTyr) is a Type 1 membrane bound glycoenzyme that catalyzes the initial and rate-limiting steps of melanin production in the melanosome. Mutations in the Tyr gene are linked to oculocutaneous albinism type 1 (OCA1), an autosomal recessive disorder. Currently, the application of enzyme replacement therapy for a treatment of OCA1 is hampered by the absence of pure hTyr. Here, full-length hTyr (residues 1-529) was overexpressed in Trichoplusia ni larvae infected with a baculovirus, solubilized with detergent and purified using chromatography. Michaelis-Menten kinetics, enzymatic specific activity, and analytical ultracentrifugation were used to compare the hTyr in detergent with the soluble recombinant intra-melanosomal domain, hTyrCtr (residues 19-469). Active hTyr is monomeric in detergent micelles suggesting no stable interactions between protein molecules. Both, hTyr and hTyrCtr, exhibited similar enzymatic activity and ligand affinity in L-DOPA and L-Tyrosine reactions. In addition, expression in larvae is a scalable process that will allow high yield protein production. Thus, larval production of enzymatically active human tyrosinase potentially could be a useful tool in developing a cure for OCA1.

Oculocutaneous albinism type 1: link between mutations, tyrosinase conformational stability, and enzymatic activity.

Oculocutaneous albinism type 1 (OCA1) is an autosomal recessive disorder caused by mutations in the tyrosinase gene. Two subtypes of OCA1 have been described: severe OCA1A with complete absence of tyrosinase activity and less severe OCA1B with residual tyrosinase activity. Here, we characterize the recombinant human tyrosinase intramelanosomal domain and mutant variants, which mimic genetic changes in both subtypes of OCA1 patients. Proteins were prepared using site-directed mutagenesis, expressed in insect larvae, purified by chromatography, and characterized by enzymatic activities, tryptophan fluorescence, and Gibbs free energy changes. The OCA1A mutants showed very low protein expression and protein yield and are enzymatically inactive. Mutants mimicking OCA1B were biochemically similar to the wild type, but exhibited lower specific activities and protein stabilities. The results are consistent with clinical data, which indicates that OCA1A mutations inactivate tyrosinase and result in severe phenotype, while OCA1B mutations partially inactivate tyrosinase and result in OCA1B albinism.