GWAS in Mice Maps Susceptibility to HIV-Associated Nephropathy to the Ssbp2 Locus.

BACKGROUND: To gain insight into the pathogenesis of collapsing glomerulopathy, a rare form of FSGS that often arises in the setting of viral infections, we performed a genome-wide association study (GWAS) among inbred mouse strains using a murine model of HIV-1 associated nephropathy (HIVAN). METHODS: We first generated F1 hybrids between HIV-1 transgenic mice on the FVB/NJ background and 20 inbred laboratory strains. Analysis of histology, BUN, and urinary NGAL demonstrated marked phenotypic variation among the transgenic F1 hybrids, providing strong evidence for host genetic factors in the predisposition to nephropathy. A GWAS in 365 transgenic F1 hybrids generated from these 20 inbred strains was performed. RESULTS: We identified a genome-wide significant locus on chromosome 13-C3 and multiple additional suggestive loci. Crossannotation of the Chr. 13 locus, including single-cell transcriptomic analysis of wildtype and HIV-1 transgenic mouse kidneys, nominated Ssbp2 as the most likely candidate gene. Ssbp2 is highly expressed in podocytes, encodes a transcriptional cofactor that interacts with LDB1 and LMX1B, which are both previously implicated in FSGS. Consistent with these data, older Ssbp2 null mice spontaneously develop glomerulosclerosis, tubular casts, interstitial fibrosis, and inflammation, similar to the HIVAN mouse model. CONCLUSIONS: These findings demonstrate the utility of GWAS in mice to uncover host genetic factors for rare kidney traits and suggest Ssbp2 as susceptibility gene for HIVAN, potentially acting via the LDB1-LMX1B transcriptional network.

In vitro performance characteristics of valved holding chamber and spacer devices with a fluticasone metered-dose inhaler.

STUDY OBJECTIVE: To compare the in vitro aerosol deposition characteristics of several commercially available valved holding chamber (VHC) and spacer devices used with a fluticasone metered-dose inhaler (MDI). DESIGN: In vitro aerosol deposition study SETTING: University-affiliated research center. DEVICES: Seven VHC devices: BreatheRite, E-Z Spacer, EasiVent, AeroChamber, InspirEase, OptiChamber, and Space Chamber. Six spacer devices: OptiHaler, Aerosol Cloud Enhancer (ACE), Gentle-Haler, MediSpacer, Ellipse, and a 6-inch tube (1-inch inside diameter). INTERVENTION: The respirable dose (aerosol particles 1-5 microm) of fluticasone was determined by sampling 10 220-microg actuations from five runs with each spacer or VHC plus MDI combination, by using a well-established in vitro cascade impactor method. MEASUREMENTS AND MAIN RESULTS: Fluticasone aerosol was washed from the impactor with methanol and quantified by means of high-performance liquid chromatography. Differences among outcomes were determined with analysis-of-variance testing. Among spacers, Ellipse had the highest respirable dose (104 microg, p < 0.01). Respirable doses for the 6-inch tube (74.3 microg), Gentle-Haler (81.7 microg), and MediSpacer (82.6 microg) were no different from that of the MDI (p > 0.05), whereas respirable doses of OptiHaler (44.6 microg) and ACE (47.2 microg) were less than those of all other spacers (p < 0.001). Among VHC devices, respirable doses from EasiVent (35.6 microg), AeroChamber (47.0 microg), InspirEase (52.7 microg), OptiChamber (53.1 microg), and Space Chamber (58.3 microg) were not different (p > 0.05), whereas BreatheRite (13.1 microg) and E-Z Spacer (27.3 microg) respirable doses were less than those of the other VHC devices (p < 0.05). CONCLUSION: Spacers and VHC devices available in the United States do not demonstrate equivalent in vitro performance with the fluticasone MDI. The difference between highest and lowest respirable doses in each device category would likely lead to clinically relevant differences in the quantity of fluticasone delivered to a patient.

Differences in inhaled fluticasone bioavailability between holding chambers in children with asthma.

STUDY OBJECTIVE: To determine if differences between holding chambers in previous in vitro aerosol studies would agree with the bioavailability of inhaled fluticasone propionate between the same holding chambers in children with asthma. DESIGN: Double-blind, randomized, crossover study SETTING: University of Florida Clinical Research Center. PATIENTS: Eight children (aged 5-9 yrs) with stable asthma. INTERVENTION: Under observation, the children inhaled two 110-microg puffs of fluticasone twice/day through the InspirEase or E-Z Spacer holding chamber. MEASUREMENTS AND MAIN RESULTS: Blood samples were collected at baseline and then at 0.5, 1, 1.5, 2, 4, and 6 hours after the last dose of fluticasone. Primary outcome measures were peak fluticasone steady-state plasma concentration (Cmax) and area under the fluticasone plasma concentration-time curve from 0-6 hours (AUC0-6). The fluticasone plasma concentrations were determined by high-performance liquid chromatography-mass spectrometric assay. Mean +/- SD Cmax from InspirEase (245 +/- 77 pg/ml) was 18% higher than that after E-Z Spacer (199 +/- 58 pg/ml, p < 0.05). Mean +/- SD AUC0-6 from InspirEase (1103 +/- 305 pg x hr(-1) x ml(-1) was 22% higher than that delivered from E-Z Spacer (863 +/- 258 pg x hr(-1) x ml(-1), p = 0.06). CONCLUSION: Differences in inhaled fluticasone bioavailability between these holding chambers in children with asthma corroborate differences reported in earlier in vitro aerosol studies.

Performance of a corticosteroid inhaler with a spacer fashioned from a plastic cold-drink bottle: effects of changing bottle volume.

Some clinicians advocate using metered-dose inhaler (MDI) spacers prepared from common household bottles. We evaluated the in vitro aerosol characteristics of fluticasone from the MDI alone and through spacers fashioned from 237-, 500-, 1000-, and 1500-mL polyethylene terephlate plastic cold drink bottles using a cascade impactor. Ten 110-microg actuations of fluticasone were sampled into the impactor during each of five runs with each configuration. The primary outcomes were the respirable dose (i.e., quantity aerosol 1.1-4.7 microm in size) and the quantity trapped in the oropharynx. The mean fluticasone respirable dose from the MDI alone (46.2 microg/actuation) was no different (p > 0.05) compared with the same MDI mated to any of the bottle spacers, regardless of volume. The mean quantity of fluticasone trapped in the oropharynx from the MDI alone (39.2 microg/actuation) was greater (p < 0.001) than the same MDI mated with any bottle spacer (range 1.5-3.5 microg/actuation). These data suggest that any of the bottle spacers tested would be an acceptable method to decrease the quantity of fluticasone that would deposit onto the oropharynx and thereby diminish the risk of topical adverse effects. Among the range of volumes tested, there was no relationship between spacer volume and respirable dose of fluticasone.

In vitro deposition of fluticasone aerosol from a metered-dose inhaler with and without two common valved holding chambers.

BACKGROUND: Previous in vitro aerosol deposition experiments indicate that the corticosteroid respirable dose from a metered-dose inhaler (MDI) can vary by threefold depending on the specific valved holding chamber (VHC) MDI combination. OBJECTIVE: We compared in vitro aerosol deposition from a fluticasone propionate MDI (Flovent, GlaxoSmithKline, Research Triangle Park, NC) to that of the same MDI used in combination with two VHCs (EasiVent, Dey, Napa, Ca; and AeroChamber-Plus, Monaghan Medical Corp, Plattsburgh, NY) to evaluate how these VHCs affect the respirable dose of fluticasone. METHODS: The respirable dose (aerosol particles 1 to 5 microm in size) of fluticasone was determined by sampling 5 x 110-microg actuations from each configuration (MDI alone, MDI plus AeroChamber-Plus, and MDI plus EasiVent) in multiples of ten using a well established, in vitro cascade impactor method. Fluticasone aerosol was washed from individual impactor stages with 50% methanol and quantified via ultraviolet high-pressure liquid chromatography. Differences among outcomes were determined using analysis of variance. RESULTS: Mean respirable dose from AeroChamber-Plus (47.9 +/- 6.9 microg/actuation) was not different (P > 0.05) from that produced by the MDI alone (50.3 +/- 2.2 microg/actuation). EasiVent respirable dose (27.0 +/- 3.6 microg/actuation) was less than that produced by either the AeroChamber-Plus or the MDI alone (P < 0.001). CONCLUSIONS: VHCs do not display equivalent in vitro performance with a fluticasone MDI. If a patient needs a VHC, clinicians should use available in vitro performance information to aid in selecting the best device.