Synthesis and Expression of a Targeted, Ferritin-Based Tracer for PET Imaging of Kidney Glomeruli.

Cationic ferritin (CF) has been developed as a multimodal, targeted imaging tracer to directly detect and map nephrons in the kidney in vivo. Direct detection of functional nephrons provides a unique, sensitive biomarker to predict or monitor kidney disease progression. CF has been developed to map functional nephron number from magnetic resonance imaging (MRI) or positron emission tomography (PET). Previous preclinical imaging studies have used non-human-derived ferritin and commercial formulations that must still be developed for translation to clinical use. Here we describe the reproducible formulation of CF (either derived from horse or from human recombinant ferritin) optimized for intravenous injection and radiolabeling by PET. The human recombinant heteropolymer ferritin is spontaneously assembled in liquid culture (Escherichia coli, E. coli) and modified to form human recombinant cationic ferritin (HrCF) to mitigate potential immunologic reactions for use in humans.

Measuring the intrarenal distribution of glomerular volumes from histological sections.

Glomerular volume is an important metric reflecting glomerular filtration surface area within the kidney. Glomerular hypertrophy, or increased glomerular volume, may be an important marker for renal stress. Current stereological techniques report the average glomerular volume (AVglom) within the kidney. These techniques cannot assess the spatial or regional heterogeneity common in developing renal pathology. Here, we report a novel "unfolding" technique to measure the actual distribution of individual glomerular volumes in a kidney from the two-dimensional glomerulus profiles observed by optical microscopy. The unfolding technique was first developed and tested for accuracy with simulations and then applied to measure the number of glomeruli (Nglom), AVglom, and intrarenal distribution of individual glomerular volume (IVglom) in the oligosyndactyl (Os/(+)) mouse model compared with wild-type (WT) controls. The Os/(+) mice had fewer and larger glomeruli than WT mice: Nglom was 12,126 ± 1,658 (glomeruli/kidney) in the WT mice and 5,516 ± 899 in the Os/(+) mice; AVglom was 2.01 ± 0.28 × 10(-4) mm(3) for the WT mice and 3.47 ± 0.35 × 10(-4) mm(3) for the Os/(+) mice. Comparing the glomerular volume distributions in Os/(+) and WT kidneys, we observed that the Os/(+) distribution peaked at a higher value of IVglom than the WT distribution peak, and glomeruli with a radius greater than 55 µm were more prevalent in the Os/(+) mice (3.4 ± 1.6% of total glomeruli vs. 0.6 ± 1.2% in WT). Finally, the largest profiles were more commonly found in the juxtamedullary region. Unfolding is a novel stereological technique that provides a new quantitative view of glomerular volume distribution in the individual kidney.

Use of Cationized Ferritin Nanoparticles to Measure Renal Glomerular Microstructure with MRI.

Magnetic resonance imaging (MRI) is becoming important for whole-kidney assessment of glomerular morphology, both in vivo and ex vivo. MRI-based renal morphological measurements can be made in intact organs and allow direct measurements of every perfused glomerulus. Cationic ferritin (CF) is used as a superparamagnetic contrast agent for MRI. CF binds to the glomerular basement membrane after intravenous injection, allowing direct, whole-kidney measurements of glomerular number, volume, and volume distribution. Here we describe the production, testing, and use of CF as an MRI contrast agent for quantitative glomerular morphology in intact mouse, rat, and human kidneys.

Phenotyping by magnetic resonance imaging nondestructively measures glomerular number and volume distribution in mice with and without nephron reduction.

Reduced nephron mass is strongly linked to susceptibility to chronic renal and cardiovascular diseases. There are currently no tools to identify nephropenia in clinical or preclinical diagnostics. Such new methods could uncover novel mechanisms and therapies for chronic kidney disease (CKD) and reveal how variation among traits can affect renal function and morphology. Here we used cationized ferritin (CF)­enhanced MRI (CFE-MRI) to investigate the relationship between glomerular number (Nglom) and volume (Vglom) in kidneys of healthy wild-type mice and mice with oligosyndactylism (Os/+), a model of congenital nephron reduction. Mice were injected with CF and perfused, and the resected kidneys were imaged with 7T MRI to detect CF-labeled glomeruli. CFE-MRI was used to measure the intrarenal distribution of individual glomerular volumes and revealed two major populations of glomeruli distinguished by size. Spatial mapping revealed that the largest glomeruli were located in the juxtamedullary region in both wild-type and Os/+ mice and the smallest population located in the cortex. Os/+ mice had about a 50% reduction and 35% increase of Nglom and Vglom, respectively, in both glomerular populations compared with wild type, consistent with glomerular hypertrophy in the Os/+ mice. Thus, we provide a foundation for whole-kidney, MRI-based phenotyping of mouse renal glomerular morphology and provide new potential for quantitative human renal diagnostics.

Principles and emerging applications of nanomagnetic materials in medicine.

The development of nanometer-scale magnetic materials for biomedical applications spans the interface between the physical sciences and biology. Applications of these materials are rapidly becoming important in medicine and enable targeted therapies and diagnostics. At the same time, specific applications add focus to the development of novel magnetic materials and facilitate a deeper understanding of the physical mechanisms behind their function. This review presents a broad, nontechnical overview of the basis of magnetism in materials at the nanometer scale and describes how these materials are created, characterized, and used. Specific emerging applications in medical diagnostics and therapies are discussed, including cancer cell targeting for thermal ablation, tissue engineering, and three-dimensional noninvasive molecular imaging. Challenges in these fields are discussed, including the toxicity and delivery of magnetic nanomaterials and the sensitivity of imaging and therapeutic techniques. The development of novel nanomagnetic nanomaterials should continue to accelerate as new applications are identified and researchers uncover new mechanisms to increase and modulate magnetism at the nanometer scale.

Multiple, novel biologically active endophytic actinomycetes isolated from upper Amazonian rainforests.

Microbial biodiversity provides an increasingly important source of medically and industrially useful compounds. We have isolated 14 actinomycete species from a collection of approximately 300 plant stem samples from the upper Amazonian rainforest in Peru. All of the cultured isolates produce substances with inhibitory activity directed at a range of potential fungal and bacterial pathogens. For some organisms, this activity is very broad in spectrum while other organisms show specific activity against a limited number of organisms. Two of these organisms preferentially inhibit bacterial test organisms over eukaryotic organisms. rDNA sequence analysis indicates that these organisms are not equivalent to any other cultured deposits in GenBank. Our results provide evidence of the untapped biodiversity in the form of biologically active microbes present within the tissues of higher plants.

Bioactive endophytes warrant intensified exploration and conservation.

BACKGROUND: A key argument in favor of conserving biodiversity is that as yet undiscovered biodiversity will yield products of great use to humans. However, the link between undiscovered biodiversity and useful products is largely conjectural. Here we provide direct evidence from bioassays of endophytes isolated from tropical plants and bioinformatic analyses that novel biology will indeed yield novel chemistry of potential value. METHODOLOGY/PRINCIPAL FINDINGS: We isolated and cultured 135 endophytic fungi and bacteria from plants collected in Peru. nrDNAs were compared to samples deposited in GenBank to ascertain the genetic novelty of cultured specimens. Ten endophytes were found to be as much as 15-30% different than any sequence in GenBank. Phylogenetic trees, using the most similar sequences in GenBank, were constructed for each endophyte to measure phylogenetic distance. Assays were also conducted on each cultured endophyte to record bioactivity, of which 65 were found to be bioactive. CONCLUSIONS/SIGNIFICANCE: The novelty of our contribution is that we have combined bioinformatic analyses that document the diversity found in environmental samples with culturing and bioassays. These results highlight the hidden hyperdiversity of endophytic fungi and the urgent need to explore and conserve hidden microbial diversity. This study also showcases how undergraduate students can obtain data of great scientific significance.