Methodology of asystolic donor heart’s condition study in an experiment using small laboratory animals

In transplantation, there has always been an acute problem of the discrepancy between the number of donor organs and the number of recipients, including donor hearts. There are various ways to increase the pool of donor organs, one of them is the use of asystolic or non-heart-beating donors. Due to poor myocardial tolerance of ischemia during the asystole period, as well as because of the difficulties in diagnosing cardiac diseases of the asystolic donor, which can be contraindication to transplantation. Therefore, an in-depth study of the state of the myocardium in asystolic donors is required. Currently, there is no generally accepted protocol for working with asystolic heart donors. This protocol should include methods of heart conditioning and assessing of myocardium state. For its development we need more experimental and preclinical studies. A protocol for such a study is proposed. The modeling of an asystolic donor using rats is described on the basis of experimental work carried out by a team of authors. The article describes the following technical aspects: anesthetic guidance, asystole detection criterion, maintaining the rat body temperature in accordance with the human body temperature during cardiac arrest, surgical aspects of performing the main experimental model. The Langendorff model of isolated cardiac perfusion was chosen as the main model for assessing the state of the myocardium of a small laboratory animal. Intra-left ventricular pressure, volume of coronary blood flow, heart rate and the presence of post-reperfusion arrhythmias were selected as criteria for assessing the state of donor hearts. Assessment of the volume of damage to the donor heart is carried out using triphenyltetrazolium chloride staining of the donor organ.

Longitudinal high-resolution imaging through a flexible intravital imaging window

Intravital microscopy (IVM) is a powerful technique that enables imaging of internal tissues at (sub)cellular resolutions in living animals. Here, we present a silicone-based imaging window consisting of a fully flexible, suture-less design that is ideally suited for long-term, longitudinal IVM of growing tissues and tumors. Crucially, we show that this window, without any customization, is suitable for numerous anatomical locations in mice using a rapid and standardized implantation procedure. This low-cost device represents a substantial technological and performance advance that facilitates intravital imaging in diverse contexts in higher organisms, opening new avenues for in vivo imaging of soft and fragile tissues.One-sentence summaryThis study presents a versatile, fully flexible imaging window that acts as an implantable transparent ‘second skin’ for small laboratory animal in vivo imaging.

Comparative Analysis of Genome of Ehrlichia sp. HF, a Model Bacterium to Study Fatal Human Ehrlichiosis

Background The genus Ehrlichia consists of tick-borne obligatory intracellular bacteria that can cause deadly diseases of medical and agricultural importance. Ehrlichia sp. HF, isolated from Ixodes ovatus ticks in Japan [also referred to as I. ovatus Ehrlichia (IOE) agent], causes acute fatal infection in laboratory mice that resembles acute fatal human monocytic ehrlichiosis caused by Ehrlichia chaffeensis. As there is no small laboratory animal model to study fatal human ehrlichiosis, Ehrlichia sp. HF provides a needed disease model. However, the inability to culture Ehrlichia sp. HF and the lack of genomic information have been a barrier to advance this animal model. In addition, Ehrlichia sp. HF has several designations in the literature as it lacks a taxonomically recognized name. Results We stably cultured Ehrlichia sp. HF in canine histiocytic leukemia DH82 cells from the HF strain-infected mice, and determined its complete genome sequence. Ehrlichia sp. HF has a single double-stranded circular chromosome of 1,148,904 bp, which encodes 866 proteins with a similar metabolic potential as E. chaffeensis. Ehrlichia sp. HF encodes homologs of all virulence factors identified in E. chaffeensis, including 23 paralogs of P28/OMP-1 family outer membrane proteins, type IV secretion system apparatus and effector proteins, two-component systems, ankyrin-repeat proteins, and tandem repeat proteins. Ehrlichia sp. HF is a novel species in the genus Ehrlichia, as demonstrated through whole genome comparisons with six representative Ehrlichia species, subspecies, and strains, using average nucleotide identity, digital DNA-DNA hybridization, and core genome alignment sequence identity. Conclusions The genome of Ehrlichia sp. HF encodes all known virulence factors found in E. chaffeensis, substantiating it as a model Ehrlichia species to study fatal human ehrlichiosis. Comparisons between Ehrlichia sp. HF and E. chaffeensis will enable identification of in vivo virulence factors that are related to host specificity, disease severity, and host inflammatory responses. We propose to name Ehrlichia sp. HF as Ehrlichia japonica sp. nov. (type strain HF), to denote the geographic region where this bacterium was initially isolated.

Primary Tree Shrew Dermis Fibroblasts Support Lytic Replication of Human Cytomegalovirus

As a universal pathogen leading to neonatal defects and transplant failure, Human cytomegalovirus (HCMV) has strict species specificity that the inability to using this virus in animals has hampered its pathogenesis study. However, the mechanism of cross-species barrier remains elusive that no non-human cell model has been established to fill this knowledge gap. We observed that primary dermis fibroblasts (TSDF) isolated from the Chinese tree shrew (Tupaia belangeri chinensis), a small laboratory animal with close affinity to primates, were permissive to HCMV replication. In TSDF infected with GFP-expressing HCMV, the green fluorescence and cytopathic effect were observed and the expression of 3 kinetic genes and replication of viral genome were detected. The cell-free viruses produced in TSDF reached 103 pfu/mL at 96 hpi, which were 10-fold lower than in primary human foreskin fibroblasts. Our results demonstrated that TSDF supported low level of lytic replication of HCMV. The TSDF model provides a useful platform for the mechanism study of species barrier of HCMV.

Approaches for the vaccination and treatment ofNeospora caninuminfections in mice and ruminant models

SUMMARYNeospora caninumis a leading cause of abortion in cattle, and is thus an important veterinary health problem of high economic significance. Vaccination has been considered a viable strategy to prevent bovine neosporosis. Different approaches have been investigated, and to date the most promising results have been achieved with live-attenuated vaccines. Subunit vaccines have also been studied, and most of them represented components that are functionally involved in (i) the physical interaction between the parasite and its host cell during invasion or (ii) tachyzoite-to-bradyzoite stage conversion. Drugs have been considered as an option to limit the effects of vertical transmission ofN. caninum.Promising results with a small panel of compounds in small laboratory animal models indicate the potential value of a chemotherapeutical approach for the prevention of neosporosis in ruminants. For both, vaccines and drugs, the key for success in preventing vertical transmission lies in the application of bioactive compounds that limit parasite proliferation and dissemination, without endangering the developing fetus not only during an exogenous acute infection but also during recrudescence of a chronic infection. In this review, the current status of vaccine and drug development is presented and novel strategies against neosporosis are discussed.