Financial outcomes of high-flow nasal cannula use for bronchiolitis on the general pediatric floor.

High-flow nasal cannula (HFNC) is an increasingly common treatment utilized for bronchiolitis on general pediatric hospital floors. This could present a financial burden for hospitals if reimbursement has not accounted for the increased costs associated with increased HFNC use. Pediatric Health Information System and Revenue Management Program data set discharges from 2018 to 2019 were utilized to calculate the ratio of reimbursements to costs as a cost coverage ratio (CCR). The CCR was compared by HFNC use, the severity of illness, and payor type. The CCR was highest at 1.68 for children with high severity and HFNC use but varied greatly by payor. The lowest CCR at 0.77 was seen for children with low severity and public insurance and indicated potential financial losses for these patients. This was lower than low-severity patients on HFNC with private payors (1.68) and high-severity patients on HFNC with public payors (1.18).

Financial Implications of Short Stay Pediatric Hospitalizations.

BACKGROUND: Observation status (OBS) stays incur similar costs to low-acuity, short-stay inpatient (IP) hospitalizations. Despite this, payment for OBS is likely less and may represent a financial liability for children's hospitals. Thus, we described the financial outcomes associated with OBS stays compared to similar IP stays by hospital and payer. METHODS: We conducted a retrospective cohort study of clinically similar pediatric OBS and IP encounters at 15 hospitals contributing to the revenue management program in 2017. Clinical and demographic characteristics were described. For each hospitalization, the cost coverage ratio (CCR) was calculated by dividing revenue by estimated cost of hospitalization. Differences in CCR were evaluated using Wilcoxon rank sum tests and results were stratified by billing designation and payer. CCR for OBS and IP stays were compared by institution, and the estimated increase in revenue by billing OBS stays as IP was calculated. RESULTS: OBS was assigned to 70 981 (56.9%) of 124 789 hospitalizations. Use of OBS varied across hospitals (8%-86%). For included hospitalizations, OBS stays were more likely than IP stays to result in financial loss (57.0% vs 35.7%). OBS stays paid by public payer had the lowest median CCR (0.6; interquartile range [IQR], 0.2-0.9). Paying OBS stays at the median IP rates would have increased revenue by $167 million across the 15 hospitals. CONCLUSIONS: OBS stays were significantly more likely to result in poor financial outcomes than similar IP stays. Costs of hospitalization and billing designations are poorly aligned and represent an opportunity for children's hospitals and payers to restructure payment models.

Tracking and quantifying polymer therapeutic distribution on a cellular level using 3D dSTORM.

We used a single-molecule localization technique called direct stochastic optical reconstruction microscopy (dSTORM) to quantify both colocalization and spatial distribution on a cellular level for two conceptually different N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer conjugates. Microscopy images were acquired of entire cells with resolutions as high as 25nm revealing the nanoscale distribution of the fluorescently labeled therapeutic components. Drug-free macromolecular therapeutics consisting of two self-assembling nanoconjugates showed slight increase in nanoclusters on the cell surface with time. Additionally, dSTORM provided high resolution images of the nanoscale organization of the self-assembling conjugates at the interface between two cells. A conjugate designed for treating ovarian cancer showed that the model drug (Cy3) and polymer bound to Cy5 were colocalized at an early time point before the model drug was enzymatically cleaved from the polymer. Using spatial descriptive statistics it was found that the drug was randomly distributed after 24h while the polymer bound dye remained in clusters. Four different fluorescent dyes were used and two different therapeutic systems were tested to demonstrate the versatility and possible general applicability of dSTORM for use in studying drug delivery systems.

Super-Resolution Imaging and Quantitative Analysis of Membrane Protein/Lipid Raft Clustering Mediated by Cell-Surface Self-Assembly of Hybrid Nanoconjugates.

Super-resolution imaging was used to quantify organizational changes in the plasma membrane after treatment with hybrid nanoconjugates. The nanoconjugates crosslinked CD20 on the surface of malignant B cells, thereby inducing apoptosis. Super-resolution images were analyzed by using pair-correlation analysis to determine cluster size and to count the average number of molecules in the clusters. The role of lipid rafts was investigated by pre-treating cells with a cholesterol chelator and actin destabilizer to prevent lipid raft formation. Lipid raft cluster size correlated with apoptosis induction after treatment with the nanoconjugates. Lipid raft clusters had radii of ∼ 200 nm in cells treated with the hybrid nanoconjugates. Super-resolution images provided precise molecule location coordinates that could be used to determine density of bound conjugates, cluster size, and number of molecules per cluster.

Multimodality imaging of coiled-coil mediated self-assembly in a "drug-free" therapeutic system.

Two complementary coiled-coil peptides CCE/CCK are used to develop a "drug free" therapeutic system, which can specifically kill cancer cells without a drug. CCE is attached to the Fab' fragment of anti-CD20 1F5 antibody (Fab'-CCE), and CCK is conjugated in multiple grafts to poly[N-(2-hydroxypropyl)methacrylamide] (P-(CCK)x ). Two conjugates are consecutively administered: First, Fab'-CCE coats peptide CCE at CD20 antigen of lymphoma cell surface; second, CCE/CCK biorecognition between Fab'-CCE and P-(CCK)x leads to coiled-coil formation, CD20 crosslinking, membrane reorganization, and ultimately cell apoptosis. To prove that two conjugates can assemble at cell surface, multiple fluorescence imaging studies are performed, including 2-channel FMT, 3D confocal microscopy, and 4-color FACS. Confocal microscopy shows colocalization of two fluorescently labeled conjugates on non-Hodgkin's lymphoma (NHL) Raji cell surface, indicating "two-step" targeting specificity. The fluorescent images also reveal that these two conjugates can disrupt normal membrane lipid distribution and form lipid raft clusters, leading to cancer cell apoptosis. This "two-step" biorecognition capacity is further demonstrated in a NHL xenograft model, using fluorescent images at whole-body, tissue and cell levels. It is also found that delaying injection of P-(CCK)x can significantly enhance targeting efficacy. This high-specificity therapeutics provide a safe option to treat NHL and other B cell malignancies.

Immunogenicity of coiled-coil based drug-free macromolecular therapeutics.

A two-component CD20 (non-internalizing) receptor crosslinking system based on the biorecognition of complementary coiled-coil forming peptides was evaluated. Exposure of B cells to Fab'-peptide1 conjugate decorates the cell surface with peptide1; further exposure of the decorated cells to P-(peptide2)x (P is the N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer backbone) results in the formation of coiled-coil heterodimers at the cell surface with concomitant induction of apoptosis. The aim of this study was to determine the potential immunogenicity of this therapeutic system that does not contain low molecular weight drugs. Enantiomeric peptides (L- and D-CCE and L- and D-CCK), HPMA copolymer-peptide conjugates, and Fab' fragment-peptide conjugates were synthesized and the immunological properties of peptide conjugates evaluated in vitro on RAW264.7 macrophages and in vivo on immunocompetent BALB/c mice. HPMA copolymer did not induce immune response in vitro and in vivo. Administration of P-peptide conjugates with strong adjuvant resulted in antibody response directed to the peptide. Fab' was responsible for macrophage activation of Fab'-peptide conjugates and a major factor in the antibody induction following i.v. administration of Fab'-conjugates. There was no substantial difference in the ability of conjugates of D-peptides and conjugates of L-peptides to induce Ab response.

Ultrasonic gene and drug delivery using eLiposomes.

eLiposomes are liposomes encapsulating emulsions and therapeutics for targeted delivery. By applying ultrasound to eLiposomes, emulsion droplets can transform from liquid to gas and rupture the lipid bilayer of the eLiposome to release a drug or plasmid. In this study, perfluoropentane (PFC5) emulsions were encapsulated inside folated eLiposomes carrying a model drug (calcein) or a model GFP plasmid to examine the effects of a folate ligand, PFC5 emulsion and various ultrasonic acoustic parameters in drug delivery and gene transfection into HeLa cells. Confocal microscopy was used to quantify drug delivery and the level of plasmid transfection into HeLa cells. The results showed that drug delivery or transfection was minimal without incorporation of internal PFC5 emulsions and folate ligand on the eLiposome surface. It was also shown that application of ultrasound greatly enhanced the drug delivery and plasmid transfection. Delivery of these therapeutics appears to be to the cytosol, indicating that the expansion of the emulsion droplets disrupted both the eLiposomes and the endosomes.

Encapsulating nanoemulsions inside eLiposomes for ultrasonic drug delivery.

An eLiposome is a liposome encapsulating an emulsion nanodroplet and can be used for drug delivery. For example, therapeutic agents are encapsulated inside the eLiposomes, and the application of ultrasound can cause the emulsion droplet to change from liquid to gas, thus increasing the volume inside the vesicle and causing rupture and the release of the drug. In this research, two different methods were used to prepare eLiposomes. In the first method, emulsion droplets were made of perfluorohexane or perfluoropentane and stabilized with 1,2-dipalmitoyl-sn-glycero-3-phosphate. A layer of 1,2-dimyristoyl-sn-glycero-3-phosphocholine was dried in a round-bottomed flask. Then the emulsion suspension was added to the flask. As the suspension hydrated the phospholipids, they formed liposomes around the emulsions. In the second method, emulsions and liposomes were made separately, and then they were mixed using ultrasound. The advantage of this second method compared to the previous one is that eLiposomes can be made with fewer restrictions because of incompatible combinations of surfactants. Dynamic light scattering and transmission electron microscopy were used to measure the size of the emulsions, liposomes, and eLiposomes. The size of eLiposomes is appropriate for extravasation into tumors with malformed capillary beds. We hypothesize that ultrasound breaks open these eLiposomes. Both types of eLiposomes were constructed with folate attached via a poly(ethylene glycol) tether to induce endocytosis of the eLiposome. The latter eLiposomes were successfully used to deliver calcein as a model drug to HeLa cells.

Preliminary results of combining low frequency low intensity ultrasound and liposomal drug delivery to treat tumors in rats.

Ultrasound is a convenient trigger for site-specific drug delivery in cancer therapy. Nano-sized liposomes formulated from soy phosphatidyl choline, cholesterol, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[carboxy(polyethylene glycol)-2000] and alpha-tocopherol were loaded with Doxorubicin (Dox) using a pH gradient. The liposomal suspension was infused through the tail vein of BDIX rats possessing bilateral intradermal DHD/K12 tumors on their hind legs. Then 20-kHz ultrasound was applied to only one of the tumors for 15 minutes. This therapy was repeated weekly for 4 weeks. The results showed that in five of six rats, the tumors regressed to non-measurable size within 4 weeks. A paired comparison of the normalized size of the insonated and non-insonated tumors in the same rat indicated that the insonated tumors were smaller (p < 0.0001, n = 6 rats, 21 pairs). This observation has significant potential for non-invasive site-specific therapy of solid tumors.