μOrgano: A Lego?-Like Plug & Play System for Modular Multi-Organ-Chips

Human organ-on-a-chip systems for drug screening have evolved as feasible alternatives to animal models, which are unreliable, expensive, and at times erroneous. While chips featuring single organs can be of great use for both pharmaceutical testing and basic organ-level studies, the huge potential of the organ-on-a-chip technology is revealed by connecting multiple organs on one chip to create a single integrated system for sophisticated fundamental biological studies and devising therapies for disease. Furthermore, since most organ-on-a-chip systems require special protocols with organ-specific media for the differentiation and maturation of the tissues, multi-organ systems will need to be temporally customizable and flexible in terms of the time point of connection of the individual organ units. We present a customizable Lego^®-like plug & play system, μOrgano, which enables initial individual culture of single organ-on-a-chip systems and subsequent connection to create integrated multi-organ microphysiological systems. As a proof of concept, the μOrgano system was used to connect multiple heart chips in series with excellent cell viability and spontaneously physiological beat rates.     

Generation of Mouse Small Intestinal Epithelial Cell Lines That Allow the Analysis of Specific Innate Immune Functions

Cell lines derived from the small intestine that reflect authentic properties of the originating intestinal epithelium are of high value for studies on mucosal immunology and host microbial homeostasis. A novel immortalization procedure was applied to generate continuously proliferating cell lines from murine E19 embryonic small intestinal tissue. The obtained cell lines form a tight and polarized epithelial cell layer, display characteristic tight junction, microvilli and surface protein expression and generate increasing transepithelial electrical resistance during in vitro culture. Significant up-regulation of Cxcl2 and Cxcl5 chemokine expression upon exposure to defined microbial innate immune stimuli and endogenous cytokines is observed. Cell lines were also generated from a transgenic interferon reporter (Mx2-Luciferase) mouse, allowing reporter technology-based quantification of the cellular response to type I and III interferon. Thus, the newly created cell lines mimic properties of the natural epithelium and can be used for diverse studies including testing of the absorption of drug candidates. The reproducibility of the method to create such cell lines from wild type and transgenic mice provides a new tool to study molecular and cellular processes of the epithelial barrier.     

Isolation and Chimerization of a Highly Neutralizing Antibody Conferring Passive Protection against Lethal Bacillus anthracis Infection

Several studies have demonstrated that the passive transfer of protective antigen (PA)-neutralizing antibodies can protect animals against Bacillus anthracis infection. The standard protocol for the isolation of PA-neutralizing monoclonal antibodies is based upon a primary selection of the highest PA-binders by ELISA, and usually yields only few candidates antibodies. We demonstrated that by applying a PA-neutralization functionality-based screen as the primary criterion for positive clones, it was possible to isolate more than 100 PA-neutralizing antibodies, some of which exhibited no measurable anti-PA titers in ELISA. Among the large panel of neutralizing antibodies identified, mAb 29 demonstrated the most potent activity, and was therefore chimerized. The variable region genes of the mAb 29 were fused to human constant region genes, to form the chimeric 29 antibody (cAb 29). Guinea pigs were fully protected against infection by 40LD₅₀ B. anthracis spores following two separate administrations with 10 mg/kg of cAb 29: the first administration was given before the challenge, and a second dose was administered on day 4 following exposure. Moreover, animals that survived the challenge and developed endogenous PA-neutralizing antibodies with neutralizing titers above 100 were fully protected against repeat challenges with 40LD₅₀ of B. anthracis spores. The data presented here emphasize the importance of toxin neutralization-based screens for the efficient isolation of protective antibodies that were probably overlooked in the standard screening protocol. The protective activity of the chimeric cAb 29 demonstrated in this study suggest that it may serve as an effective immunotherapeutic agent against anthrax.     

The Risk of Rh Isoimmunization in Ruptured Tubal Pregnancy

In 9 (24%) out of 38 African women who had suffered a ruptured tubal pregnancy significant numbers of fetal erythrocytes (5 or more per 150,000 maternal cells) were found in the maternal circulation. This is a higher incidence than occurs after abortion and indicates that rupture of a tubal pregnancy is a potential source of Rh isoimmunization. The finding of fetal cells in the peritoneal cavity suggests that this is the main source of the fetal blood found in the maternal circulation. At operation on Rh-negative patients with ruptured tubal pregnancies, therefore, complete removal of the peritoneal blood should be attempted and the blood recovered should never be transfused into the patient, who should always receive prophylactic Rh immunoglobulin.     

Cyclic Nucleotides and the Release of Vasopressin from the Rat Posterior Pituitary Gland

Abstract: The effect of ATP, Mg²⁺, or MgATP on the release of luteinizing hormone-releasing hormone (LH-RH) from hypothalamic granules was examined under in vitro conditions. Granules, isolated from adult male hypothalami, were incubated at 37°C in a buffered (pH 7.8) medium containing 0.15 m -KCl. The addition of ATP to the incubation mixture did not stimulate the release of LH-RH. In contrast, the addition of MgATP stimulated the release of LH-RH, the release being 62% greater than control. The addition of Mg²⁺ to the incubated granules also stimulated the release of LH-RH. However, the magnitude of this Mg²⁺-stimulated release of LH–RH was significantly ( P < 0.01) lower than that of the MgATP-stimulated release, indicating that ATP stimulates LH-RH release in a Mg²⁺-dependent manner. As both MgATP and Mg²⁺ alone stimulated LH-RH release, we characterized further these two release processes by incubating the granules under one of the following conditions: incubation at 4°C in a buffered medium containing 0.15 m -KCl or incubation at 37°C in a medium that does not contain KCl. Under these two incubation conditions, the MgATP-stimulated release of LH-RH was not manifested, whereas the Mg²⁺-stimulated release of LH-RH was manifested. On the basis of these differences, we propose that two different processes can lead to the release of LH-RH from isolated hypothalamic granules: one process involves ATP and Mg²⁺ (MgATP) and another process involves Mg²⁺ alone.