Technical Reports

The use of antibodies against tumour necrosis factor (TNF) has markedly improved the treatment of Crohn's disease, but only certain patients respond to therapy. Here, Raja Atreya and colleagues have developed an approach using topical fluorescent antibodies to TNF and confocal laser endomicroscopy to evaluate the expression of transmembrane TNF (mTNF) in the intestinal mucosa of patients with active Crohn's disease in order to identify patients likely to respond to subsequent treatment with the anti-TNF therapy, adalimumab.

Relapses in malaria are caused by hypnozoites, the latent hepatic stage formed by species such as Plasmodium vivax and Plasmodium ovale. Drug discovery programs have been severely hampered by a lack of in vitro cultivation methods for malarial hypnozoites. Only one drug, primaquine, is currently available, but its use is limited in people with glucose-6-phosphate dehydrogenase deficiency. Here, Laurent Dembélé and colleagues offer a system that can be used to monitor the growth and development of Plasmodium cynomologi liver-stage forms, a model for P. vivax, for up to 40 d.

Diagnosis of bacterial infections is largely based on nonspecific criteria, with definitive diagnoses made only after biopsy or culture. Frank Hernandez and his colleagues demonstrate noninvasive imaging of Staphylococcus aureus infections in mice with an activatable fluorescent molecular imaging probe. The approach exploits the properties of micrococcal nuclease and uses short, synthetic oligonucleotides that are highly sensitive to micrococcal nuclease but are rendered resistant to mammalian serum nucleases by chemical modifications.

PET imaging using 2-deoxy-2-[¹⁸F]fluoro-d-glucose ([¹⁸F]FDG) has been widely used for the detection of plaque inflammation. Here, however, Nobuhiro Tahara and colleagues explore the possibility of using ¹⁸F-labeled mannose (2-deoxy-2-[¹⁸F]fluoro-d-mannose, [¹⁸F]FDM), a structural analog to FDG, as an alternative and potentially more specific PET tracer than FDG for assessing the risk of acute vascular events in patients. The approach targets the mannose receptor–bearing macrophages that are abundant in high-risk atherosclerotic plaques, with feasibility demonstrated in a rabbit model of atherosclerosis.

The measurement of metabolites such as creatine, which are involved in the tissue creatine kinase reaction, enable the study of the effects of energy deprivation on the heart. Haris and colleagues introduce a new magnetic resonance imaging technique that maps the distribution of creatine in the heart that does not use radiation or exogenous contrast agents and offers higher sensitivity compared to proton (¹H) magnetic resonance spectroscopy methods. Feasibility is demonstrated in vivo in infarcted swine myocardium using a standard clinical MRI scanner.

One of the most likely substrates for metabolic imaging of response to treatment in cancer is glucose, but until now, using hyperpolarized 13C-labelled glucose has been problematic because of the short lifetime of the hyperpolarization in this molecule. Using [U-¹³C, U-²H]glucose, Tiago Rodrigues et al. now show that they are able to image its glycolytic conversion to lactate in two mouse tumor models in vivo, and that in one model, flux is markedly reduced after treatment with the chemotherapeutic drug etoposide.

There are currently a paucity of approaches for the direct in vivo assessment of rates of hepatic mitochondrial oxidation and anaplerotic flux in humans. With this in mind, Douglas Befroy and colleagues have developed a new ¹³C-labeling strategy that they use in combination with ¹³C magnetic resonance spectroscopy, which should prove useful in determining the potential role of changes in hepatic mitochondrial fat oxidation in diseases such as nonalcoholic fatty liver disease and type 2 diabetes.

Matthias Hackl and his colleagues developed a new serial multiphoton microscopy approach to track migrating podocytes and parietal epithelial cells over time in vivo in the intact kidney. Use of this new approach is demonstrated in several mouse models and should help answer questions surrounding podocyte proliferation and migration to other (peri)glomerular regions, thus shedding light on the mechanisms of glomerular injury and regeneration.

There is an urgent need for quantitative magnetic resonance approaches for assessing brain development, as well as for studying the effects of drugs on neural tissue inflammation. Aviv Mezer and colleagues have developed a neuroimaging method for the quantification of local tissue volume and tissue-surface interaction, producing reliable quantitative measurements across a range of scanners. They apply their method to both the healthy brain and individuals with relapsing-remitting multiple sclerosis.

Progress in T cell receptor (TCR) gene therapy has been hampered by the lack of a rapid and efficient screening system for antigen-specific TCRs. Here, Kobayashi et al. have developed a direct single-cell TCR cloning system for cloning antigen-specific TCRs from peripheral blood in 10 d. The approach is used to clone and analyze Epstein-Barr virus–specific TCRs from healthy donors with latent Epstein-Barr virus infection, as well as TCRs from peptide-vaccinated patients with hepatocellular carcinoma.

The lack of robust and high-throughput technologies to analyze the human TCR repertoire has been a bottleneck in the analysis of human T cell responses. Linnemann and colleagues have addressed this issue by using a TCR gene capture technology that, because of its quantitative nature, allows the rapid identification of TCRab pairs from bulk populations of cells without the need for single-cell cloning. Such an approach should be useful in obtaining defined antigen-reactive TCRs for therapeutic purposes.

Thorek et al. describe an activatable imaging agent based on a radioactive decay signal through the Cerenkov luminescence effect, which is light produced by β-particle–emitting radionuclides, such as clinically available PET tracers. The approach offers a means of simultaneously investigating multiple disease-relevant biological activities in vivo—the radioluminescent readout providing a quantitative measure of enzymatic activation with reduced background.

Qiong Wang and colleagues introduce the AdipoChaser mouse, an in vivo tool to track the formation and turnover of adipocytes. They use this inducible mature adipocyte lineage-tracing system to monitor adipogenesis and follow the formation of white and beige adipocytes under different conditions: high-fat diet, cold exposure and β-adrenergic stimulation. The system produced some interesting findings on in vivo adipogenesis, including that beige adipocytes differentiate de novo from specialized precursors rather than by transdifferentiation of mature white adipocytes.

Hans Wehrl et al. use a multimodal approach to demonstrate the feasibility of measuring functional brain responses in activated and resting states with PET and MRI simultaneously. Using this method, which allows brain function studies to be conducted on multiple time and metabolic scales, they could tease apart the complex relationships between neural activity, blood flow and oxygenation that form the basis of the blood oxygen level–dependent (BOLD) effect.

There is a pressing need for techniques that can be used for the noninvasive assessment of response to therapy and staging of disease. As many pathological conditions are associated with disordered glucose metabolism, such as diabetes, stroke and cancer, Simon Walker-Samuel and his colleagues have developed a noninvasive MRI-based method for imaging glucose uptake in vivo termed glucose chemical exchange saturation transfer (glucoCEST). This potentially cost-effective approach does not require the use of radiolabeled glucose analogs or ionizing radiation and allows nonlabeled glucose to be imaged at physiological quantities.

Building on earlier work, Dekkers et al. describe the first application of their intestinal organoid culture technology to the study of human disease, in this case cystic fibrosis. These so called 'mini-guts', which recapitulate the essential in vivo intestinal tissue architecture in vitro, are used to develop a rapid and quantitative assay to measure mutant cystic fibrosis transmembrane conductance regulator (CFTR) function, as well as test the efficacy of correctors and potentiators of mutant CFTR.

Marsilius Mues et al. have overcome previous obstacles precluding the expression of genetically encoded calcium indicators (GECIs) in immune cells by developing a new fluorescence resonance energy transfer–based GECI for the functional calcium imaging of T cells in vivo. Using two-photon imaging, the group traced the real-time activation of T cells in peripheral lymph nodes after antigen application, as well as in the CNS during experimental autoimmune encephalomyelitis.

The work of Dmitri Lodygin and his colleagues focuses on the important issue of when and where autoaggressive T cells are activated within their target organ to trigger autoimmune disease. Using intravital two-photon imaging and a new molecular sensor—a genetically encoded fluorescent sensor of nuclear factor of activated T cells (NFAT) combined with a fluorescently tagged version of nuclear histone protein H2B—the group was able to pinpoint key T cell activation events in experimental autoimmune encephalomyelitis, a model for multiple sclerosis.

Building on their earlier work on heart and lung organ engineering, Jeremy Song and his colleagues have now adapted the technology of using decellularized scaffolds to develop bioengineered kidneys for transplantation. When reseeded with endothelial and epithelial cells, and after orthotopic transplantation in rats, the grafts became perfused by the recipient's circulation, produced urine and provided clearance of metabolites.