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Despite the numerous advantages demonstrated by microfluidic mixing for RNA-loaded lipid nanoparticle (RNA-LNP) production over bulk methods, such as precise size control, homogeneous distributions, higher encapsulation efficiencies, and improved reproducibility, their translation from research to commercial manufacturing remains elusive. A persistent challenge hindering the adoption of microfluidics for LNP production is the fouling of device surfaces during prolonged operation, which significantly diminishes performance and reliability. The complexity of LNP constituents, including lipids, cholesterol, RNA, and solvent mixtures, makes it difficult to find a single coating that can prevent fouling. To address this challenge, we propose using an immobilized liquid lubricant layer of perfluorodecalin (PFD) to create an antifouling surface that can repel the multiple LNP constituents. We apply this technology to a staggered herringbone microfluidic (SHM) mixing chip and achieve >3 h of stable operation, a >15× increase relative to gold standard approaches. We also demonstrate the compatibility of this approach with a parallelized microfluidic platform that incorporates 256 SHM mixers, with which we demonstrate scale up, stable production at L/h production rates suitable for commercial scale applications. We verify that the LNPs produced on our chip match both the physiochemical properties and performance for both in vitroand in vivomRNA delivery as those made on chips without the coating. By suppressing surface fouling with an immobilized liquid lubricant layer, this technology not only enhances RNA-LNP production but also promises to transform the microfluidic manufacturing of diverse materials, ensuring more reliable and robust processes.

ABSTRACTPurposeTo compare the long-term outcomes of mucosal-sparing mechanical endoscopic dacryocystorhinostomy (MMED) for primary acquired nasolacrimal duct obstruction (PANDO) with or without silicone intubation.MethodsAn 11-year follow-up study of the Silicone intubation in Endoscopic Dacryocystorhinostomy (SEND) randomized controlled trial (RCT) was conducted at a university-affiliated dacryology clinic from December 2019 to March 2023. Questionnaires on symptoms, anterior segment examination, endoscopic examination with functional endoscopic dye test (FEDT) and FICI grading, and ostial size measurements using Image Jsoftware were performed by a masked ophthalmologist. The primary outcome was surgical success, defined by Munk’s score ≤1 and a positive fluorescein endoscopic dye test. Secondary outcomes included risk factors for failure and outcomes of revision surgeries.ResultsFifty-three of the original 118 patients were evaluated at 155 ± 21 (136–218) months postoperatively. Seventy-seven percent (46/60) ostia remained successful, including 70% (19/27) of unstented and 82% (27/33) of stented ostia (p = .3). Stented ostia had larger size (p = .003), but this did not confer higher success (p = .14). Successful ostia had higher FICI scores and better ostial dynamicity (p < .05). Ostium movement was the only parameter associated with surgical success on multivariate analysis (OR 13.1, p = .01). Four (1 stented) underwent revision MMED, intraoperative mitomycin-C, and 12-week intubation. All revision ostia were functional after 141 ± 43 months.ConclusionsSurgical success of MMED after 11-years was 77%, a notable reduction compared to 96% success at 1-year. Statistical advantage of silicone intubation for primary MMED was not demonstrated, though clinically, stented ostia had a higher success (82% vs 70%). The presence of a dynamic internal common opening was highly associated with long-term surgical success.

The N-demethylation of N,N'-dimethylphenobarbital by isolated rat hepatocytes was increased severalfold by pretreatment of animals with sodium phenobarbital (75 mg/kg/day ip for 3 days). The apparent extent of induction depended on the length of the cell incubation, being 3-fold when calculated after a 5-min incubation and 5-fold when calculated after a 60-min incubation. This difference was due to the fact that metabolism by "induced" hepatocytes more closely approached linearity over a 60-min period. Microsomal cytochrome P-450 levels were increased 3-fold upon phenobarbital treatment; during incubations of less than 30 min duration, increased N-demethylase activity could be entirely accounted for by this increase in cytochrome P-450. When protein levels were measured directly in microsomes isolated from hepatocyte homogenates, the levels in "induced" hepatocytes were approximately 45% higher than control. However, phenobarbital treatment had no significant effects on total cellular protein or on the protein distribution in various subcellular fractions when appropriate adjustments were made for apparent microsomal sedimentation with the 9000 g pellet. These data suggest that hepatic microsomes prepared from "induced" hepatocytes may sediment differently from those from untreated animals. The N-demethylation of N,N'-dimethylphenobarbital is an excellent model reaction to study in isolated hepatocytes because the primary metabolite, N-methylphenobarbital, is not further metabolized to an appreciable extent.

Duplication of methyl-CpG-binding protein 2 (MECP2) gene causes MECP2duplication syndrome (MDS). To normalize the duplicated MECP2in MDS, we developed a high-fidelity Cas13Y (hfCas13Y) system capable of targeting the MECP2(hfCas13Y-gMECP2) messenger RNA for degradation and reducing protein levels in the brain of humanized MECP2transgenic mice. Moreover, the intracerebroventricular adeno-associated virus (AAV) delivery of hfCas13Y-gMECP2in newborn or adult MDS mice restored dysregulated gene expression and improved behavior deficits. Notably, treatment with AAV9-hfCas13Y-gMECP2extended the median survival of MECP2transgenic mice from 156.5 to 226 d. Furthermore, studies with monkeys showed a single injection of AAV9-hfCas13Y-gMECP2was sufficient to drive robust expression of hfCas13Y in widespread brain regions, with MECP2knockdown efficiency reaching 52.19 ± 0.03% and significantly decreased expression of biomarker gene GDF11. Our results demonstrate that the RNA-targeting hfCas13Y-gMECP2system is an effective intervention for MDS, providing a potential strategy for treating other dosage-sensitive diseases.

Partial-thickness rotator cuff tears (PTRCTs), often considered less severe than full-thickness rotator cuff tears (FTRCTs), can be equally symptomatic. Despite the prevalence of PTRCTs, scholarly attention has predominantly focused on FTRCTs, resulting in a relative neglect of PTRCTs. This study aimed to assess the incidence of surgical repairs for PTRCTs in the United States (U.S.) using the Nationwide Ambulatory Surgery Sample (NASS) database and delineate practice patterns among a group of American Shoulder and Elbow Surgeons (ASES) members.

Procalcitonin is a rapid response biomarker specific for bacterial infection, which is not routinely used in the UK National Health Service. We aimed to assess whether using a procalcitonin-guided algorithm would safely reduce the duration of antibiotic therapy compared with usual care, in which C-reactive protein is the commonly used biomarker.