JavaCyte, a novel open-source tool for automated quantification of key hallmarks of cardiac structural remodeling.

Many cardiac pathologies involve changes in tissue structure. Conventional analysis of structural features is extremely time-consuming and subject to observer bias. The possibility to determine spatial interrelations between these features is often not fully exploited. We developed a staining protocol and an ImageJ-based tool (JavaCyte) for automated histological analysis of cardiac structure, including quantification of cardiomyocyte size, overall and endomysial fibrosis, spatial patterns of endomysial fibrosis, fibroblast density, capillary density and capillary size. This automated analysis was compared to manual quantification in several well-characterized goat models of atrial fibrillation (AF). In addition, we tested inter-observer variability in atrial biopsies from the CATCH-ME consortium atrial tissue bank, with patients stratified by their cardiovascular risk profile for structural remodeling. We were able to reproduce previous manually derived histological findings in goat models for AF and AV block (AVB) using JavaCyte. Furthermore, strong correlation was found between manual and automated observations for myocyte count (r = 0.94, p < 0.001), myocyte diameter (r = 0.97, p < 0.001), endomysial fibrosis (r = 0.98, p < 0.001) and capillary count (r = 0.95, p < 0.001) in human biopsies. No significant variation between observers was observed (ICC = 0.89, p < 0.001). We developed and validated an open-source tool for high-throughput, automated histological analysis of cardiac tissue properties. JavaCyte was as accurate as manual measurements, with less inter-observer variability and faster throughput.

Malakoplakia in Thoracic Transplant Recipients.

Malakoplakia is a rare granulomatous disease characterized by the presence of Michaelis-Gutmann bodies on histopathologic analysis. Lesions manifest in a wide range of organs with cutaneous, gastrointestinal, and genitourinary systems being most common, and often result in significant comorbidities owing largely to misdiagnoses and the similar appearance to malignancy or granulomatous processes. Most patients are immunocompromised, including the solid-organ transplant population. Among organ recipients, malakoplakia is most commonly seen in renal transplantation, and only rarely reported in thoracic organ recipients. Herein we report 2 cases of malakoplakia in thoracic transplant patients that highlight the critical need for tissue diagnosis to avoid delay in management.

Promising new assays and technologies for the diagnosis and management of infectious diseases.

In the first decade of the 21st century, we have seen the completion of the human genome project and marked progress in the human microbiome project. The vast amount of data generated from these efforts combined with advances in molecular and biomedical technologies have led to the development of a multitude of assays and technologies that may be useful in the diagnosis and management of infectious diseases. Here, we identify several new assays and technologies that have recently come into clinical use or have potential for clinical use in the near future. The scope of this review is broad and includes topics such as the serum marker procalcitonin, gene expression profiling, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and nucleic acid aptamers. Principles that underlie each assay or technology, their clinical applications, and potential strengths and limitations are addressed.

Molecular characterization of a mannoprotein with homology to chitin deacetylases that stimulates T cell responses to Cryptococcus neoformans.

The fungus Cryptococcus neoformans is a major cause of morbidity and mortality in patients with impaired CD4(+) T cell function, particularly those with AIDS. To identify cryptococcal antigens that could serve as vaccine candidates by stimulating T cell responses, C. neoformans-reactive CD4(+) T cell hybridomas were generated by immunization of C57BL/6 mice and fusion of splenocytes with thymoma cells. The antigen that stimulated one of the hybridomas, designated P1D6, to produce IL-2 was purified to homogeneity by sequential anion exchange chromatography, hydrophobic interaction chromatography, and SDS/PAGE. Based on its apparent molecular mass of 98 kDa and mannosylation, the antigen of interest was named MP98. MP98 was N terminal-sequenced, and the gene encoding the protein was cloned and sequenced. Recombinant MP98, expressed in Saccharomyces cerevisiae, stimulated P1D6 to produce IL-2. Analysis of the derived 458-aa sequence of MP98 reveals an N-terminal cleavable signal sequence, a polysaccharide deacetylase domain found in fungal chitin deacetylases, and a serine/threonine-rich C-terminal region. Overall, there were 103 serine/threonine residues serving as potential O-linked glycosylation sites as well as 12 possible N-linked glycosylation sites. Thus, a C. neoformans mannoprotein has been characterized that stimulates T cell responses and has molecular properties of a chitin deacetylase.

Inhibition of T cell recruitment and cutaneous delayed-type hypersensitivity-induced inflammation with antibodies to monocyte chemoattractant protein-1.

Leukocytes express chemokine receptors that, upon ligand recognition, are believed to activate and induce the directed migration of these cells from the vasculature to sites of tissue injury. Previous investigations of human and animal inflammatory tissue have revealed that expression of chemokines can be increased in association with leukocyte infiltration. Monocyte chemotactic protein-1 (MCP-1) mediates monocyte chemotaxis in vitro and migration of monocytes to inflammatory sites in vivo. More recently T cell chemotaxis to MCP-1 has been observed in vitro, but the contribution of this protein to T cell migration in vivo and to lymphocyte-mediated inflammation has not been determined. In this report, we show that using a rat model of cutaneous delayed hypersensitivity, MCP-1 expression correlates spatially and kinetically with T cell and monocyte recruitment and that antibodies directed to MCP-1 when administered therapeutically to animals undergoing delayed hypersensitivity can almost completely abolish T cell migration and inflammatory sequelae. Moreover the concentration of antibody needed to inhibit T cell trafficking to inflammatory sites is almost on order of magnitude lower than that needed to impede monocyte recruitment. Therefore, MCP-1 is functionally relevant in the genesis of delayed hypersensitivity and may be a useful therapeutic target for diseases mediated in part by T lymphocytes.