We conclude that the VPR tail-cuff method provides accurate blood pressure measurements over the physiological range of blood pressure in mice.

The With-No-Lysine (K) (WNK) kinases play a critical role in blood pressure regulation and body fluid and electrolyte homeostasis. Herein, we introduce the first orally bioavailable pan-WNK-kinase inhibitor, WNK463, that exploits unique structural features of the WNK kinases for both affinity and kinase selectivity. In rodent models of hypertension, WNK463 affects blood pressure and body fluid and electro-lyte homeostasis, consistent with WNK-kinase-associated physiology and pathophysiology.

Nephropathy is a major contributor to overall morbidity and mortality in diabetic patients. Early renal changes during diabetes include Na retention and renal hypertrophy. Tumor necrosis factor (TNF) is elevated during diabetes and is implicated in the pathogenesis of diabetic nephropathy. We tested the hypothesis that TNF contributes to Na retention and renal hypertrophy during diabetes. Rats with streptozotocin-induced diabetes exhibit increased urinary TNF excretion, Na retention, and renal hypertrophy through the first 20 days of diabetes. Administration of a soluble TNF antagonist (TNFR:Fc) to diabetic rats reduces urinary TNF excretion and prevents Na retention and renal hypertrophy. TNF stimulates Na uptake in distal tubule cells isolated from diabetic rats, providing a possible mechanism for TNF-induced Na retention. We conclude that urinary TNF contributes to early diabetic nephropathy and may serve as a valuable diagnostic marker. Furthermore, inhibition of TNF during diabetes may attenuate early pathological changes in diabetic nephropathy.

A large unmet medical need exists for safer antithrombotic drugs because all currently approved anticoagulant agents interfere with hemostasis, leading to an increased risk of bleeding. Genetic and pharmacologic evidence in humans and animals suggests that reducing factor XI (FXI) levels has the potential to effectively prevent and treat thrombosis with a minimal risk of bleeding. We generated a fully human antibody (MAA868) that binds the catalytic domain of both FXI (zymogen) and activated FXI. Our structural studies show that MAA868 traps FXI and activated FXI in an inactive, zymogen-like conformation, explaining its equally high binding affinity for both forms of the enzyme. This binding mode allows the enzyme to be neutralized before entering the coagulation process, revealing a particularly attractive anticoagulant profile of the antibody. MAA868 exhibited favorable anticoagulant activity in mice with a dose-dependent protection from carotid occlusion in a ferric chloride–induced thrombosis model. MAA868 also caused robust and sustained anticoagulant activity in cynomolgus monkeys as assessed by activated partial thromboplastin time without any evidence of bleeding. Based on these preclinical findings, we conducted a first-in-human study in healthy subjects and showed that single subcutaneous doses of MAA868 were safe and well tolerated. MAA868 resulted in dose- and time-dependent robust and sustained prolongation of activated partial thromboplastin time and FXI suppression for up to 4 weeks or longer, supporting further clinical investigation as a potential once-monthly subcutaneous anticoagulant therapy.

Non-tuberculous mycobacteria (NTM) cause pulmonary infections in patients with structural lung damage, impaired immunity, or other risk factors. Delivering antibiotics to the sites of these infections is a major hurdle of therapy because pulmonary NTM infections can persist in biofilms or as intracellular infections within macrophages. Inhaled treatments can improve antibiotic delivery into the lungs, but efficient nebulization delivery, distribution throughout the lungs, and penetration into biofilms and macrophages are considerable challenges for this approach. Therefore, we developed amikacin liposome inhalation suspension (ALIS) to overcome these challenges. Nebulization of ALIS has been shown to provide particles within the respirable size range that distribute to both central and peripheral lung compartments in humans. The in vitro and in vivo efficacy of ALIS against NTM has been demonstrated previously. The key mechanistic questions are whether ALIS penetrates NTM biofilms and enhances amikacin uptake into macrophages. We found that ALIS effectively penetrated throughout NTM biofilms and concentration-dependently reduced the number of viable mycobacteria. Additionally, we found that ALIS improved amikacin uptake by ∼4-fold into cultured macrophages compared with free amikacin. In rats, inhaled ALIS increased amikacin concentrations in pulmonary macrophages by 5- to 8-fold at 2, 6, and 24 h post-dose and retained more amikacin at 24 h in airways and lung tissue relative to inhaled free amikacin. Compared to intravenous free amikacin, a standard-of-care therapy for refractory and severe NTM lung disease, ALIS increased the mean area under the concentration-time curve in lung tissue, airways, and macrophages by 42-, 69-, and 274-fold. These data demonstrate that ALIS effectively penetrates NTM biofilms, enhances amikacin uptake into macrophages, both in vitro and in vivo, and retains amikacin within airways and lung tissue. An ongoing Phase III trial, adding ALIS to guideline based therapy, met its primary endpoint of culture conversion by month 6. ALIS represents a promising new treatment approach for patients with refractory NTM lung disease.

No abstract

Evidence from inbred strains of mice indicates that a quarter or more of the mammalian genome consists of chromosome regions containing clusters of functionally related genes. The intense selection pressures during inbreeding favor the coinheritance of optimal sets of alleles among these genetically linked, functionally related genes, resulting in extensive domains of linkage disequilibrium (LD) among a set of 60 genetically diverse inbred strains. Recombination that disrupts the preferred combinations of alleles reduces the ability of offspring to survive further inbreeding. LD is also seen between markers on separate chromosomes, forming networks with scale-free architecture. Combining LD data with pathway and genome annotation databases, we have been able to identify the biological functions underlying several domains and networks. Given the strong conservation of gene order among mammals, the domains and networks we find in mice probably characterize all mammals, including humans.

Recent developments in high-density genotyping and statistical analysis methods that have enabled genome-wide association studies in humans can also be applied to outbred mouse populations. Increased recombination in outbred populations is expected to provide greater mapping resolution than traditional inbred line crosses, improving prospects for identifying the causal genes. We carried out genome-wide association mapping by using 288 mice from a commercially available outbred stock; NMRI mice were genotyped with a high-density single-nucleotide polymorphism array to map loci influencing high-density lipoprotein cholesterol, systolic blood pressure, triglyceride levels, glucose, and urinary albumin-to-creatinine ratios. We found significant associations (P < 10−5) with high-density lipoprotein cholesterol and identified Apoa2 and Scarb1, both of which have been previously reported, as candidate genes for these associations. Additional suggestive associations (P < 10−3) identified in this study were also concordant with published quantitative trait loci, suggesting that we are sampling from a limited pool of genetic diversity that has already been well characterized. These findings dampen our enthusiasm for currently available commercial outbred stocks as genetic mapping resources and highlight the need for new outbred populations with greater genetic diversity. Despite the lack of novel associations in the NMRI population, our analysis strategy illustrates the utility of methods that could be applied to genome-wide association studies in humans.