A simple method is described to establish primary cultures of kidney proximal tubule cells (PTC) on membranes. The permeable membranes represent a unique culture surface, allowing a high degree of differentiation since both apical and basolateral membranes are accessible for medium. Proximal tubule (PT) segments from collagenase-digested mouse renal cortices were grown for 7 days, by which time cells were organized as a confluent monolayer. Electron microscopic evaluation revealed structurally polarized epithelial cells with numerous microvilli, basolateral invaginations, and apical tight junctions. Immunoblotting for markers of distinct parts of the nephron demonstrated that these primary cultures only expressed PT-specific proteins. Moreover immunodetection of distinct components of the receptor-mediated endocytic pathway and uptake of FITC-albumin indicated that these cells expressed a functional endocytotic apparatus. In addition, primary cultures possessed the PT brush-border enzymes, alkaline phosphatase, and gamma-glutamyl-transferase, and a phloridzin-sensitive sodium-dependent glucose transport at their apical side. Electrophysiological measurements show that the primary cultured cells have a low transepithelial resistance and high short-circuit current that was completely carried by Na(+) similar to a leaky epithelium like proximal tubule cells. This novel method established well-differentiated PTC cultures.

Multiple sclerosis is a chronic inflammatory disease of the central nervous system. Myelin and oligodendrocytes are considered the major targets of injury caused by a cell-mediated immune response. There is circumstantial evidence that proinflammatory cytokines like tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) could have disease-promoting roles in multiple sclerosis (MS). In the present study, the cytotoxic effects of IFN-gamma and TNF-alpha on the human oligodendroglial cell lines human oligodendroglioma (HOG) and MO3.13 were analyzed. When the oligodendroglial cell lines were cultured in the presence of IFN-gamma or TNF-alpha, apoptotic cell death was observed in both cell lines after>24 hr incubation. Apoptosis was evidenced by a decrease in cell viability, apoptotic changes in cell and nucleus morphology, and disruption of the membrane asymmetry. Our data show that TNF-alpha and IFN-gamma induce apoptosis in a dose-dependent fashion in both oligodendroglial cell lines and that their synergistic effect results in enhanced cell death. Understanding the regulation of cell death pathways in oligodendrocytes is critical for protecting myelin-producing cells and their associated axons during injury in patients with MS.

Mesenchymal stem cells (MSCs) are one of the most promising stem cell types due to their availability and relatively simple requirements for in vitro expansion and genetic manipulation. Besides the well-characterized MSCs derived from bone marrow, there is growing evidence suggesting that dental pulp and the umbilical cord matrix both contain a substantial amount of cells having properties similar to those of MSCs. In order to assess the potential of dental pulp-derived MSCs (DPSC) and umbilical cord-derived MSCs (UCSC) in future clinical applications, it is essential to gain more insight into their differentiation capacity and to evaluate the tissues formed by these cells. In the present study, the morphological and ultrastructural characteristics of DPSC and UCSC induced towards osteogenic, adipogenic, and chondrogenic lineages were investigated. Cultured DPSC and UCSC showed a similar expression pattern of antigens characteristic of MSCs including CD105, CD29, CD44, CD146, and STRO-1. Under appropriate culture conditions, both DPSC and UCSC showed chondrogenic and osteogenic potential. Adipogenesis could be only partially induced in DPSC resulting in the de novo expression of fatty acid binding protein (FABP), whereas UCSC expressed FABP combined with a very high accumulation of lipid droplets in the cytoplasm. Our results demonstrate, at the biochemical and ultrastructural level, that DPSC display at least bilineage potential, whereas UCSC, which are developmentally more primitive cells, show trilineage potential. We emphasize that transmission electron microscopical analysis is useful to elucidate detailed structural information and provides indisputable evidence of differentiation. These findings highlight their potential therapeutic value for cell-based tissue engineering.

While many efforts have been made to pave the way toward human space colonization, little consideration has been given to the methods of protecting spacefarers against harsh cosmic and local radioactive environments and the high costs associated with protection from the deleterious physiological effects of exposure to high-Linear energy transfer (high-LET) radiation. Herein, we lay the foundations of a roadmap toward enhancing human radioresistance for the purposes of deep space colonization and exploration. We outline future research directions toward the goal of enhancing human radioresistance, including upregulation of endogenous repair and radioprotective mechanisms, possible leeways into gene therapy in order to enhance radioresistance via the translation of exogenous and engineered DNA repair and radioprotective mechanisms, the substitution of organic molecules with fortified isoforms, and methods of slowing metabolic activity while preserving cognitive function. We conclude by presenting the known associations between radioresistance and longevity, and articulating the position that enhancing human radioresistance is likely to extend the healthspan of human spacefarers as well.

In multiple sclerosis (MS), damage to oligodendrocytes is believed to be caused by an aberrant immune response initiated by autoreactive T cells. Increasing evidence indicates that these T cells are not exclusively detrimental but might also exert protective effects. We report for the first time that myelin-reactive T-cell clones from eight MS patients (6/19) and five healthy controls (4/11) produce leukemia inhibitory factor (LIF), a member of the neuropoietic family of neurotrophins. In addition, T-cell clones specific for tetanus toxoid, CD4(+) and CD8(+) T cells, and monocytes, but not B cells, secreted LIF. LIF-producing T lymphocytes and macrophages were also identified immunohistochemically in both active and chronic-active MS lesions. We further demonstrated dose-dependent protective effects of LIF on tumor necrosis factor-alpha-induced apoptosis of oligodendrocytes. In conclusion, our data demonstrate that peripheral and CNS-infiltrating T cells from MS patients produce LIF, a protective factor for oligodendrocytes. This study emphasizes that secretion of LIF may contribute to the neuroprotective effects of autoreactive T cells.

Despite the generalized use of photon-based radiation (i.e., gamma rays and X-rays) to treat different cancer types, particle radiotherapy (i.e., protons and carbon ions) is becoming a popular, and more effective tool to treat specific tumors due to the improved physical properties and biological effectiveness. Current scientific evidence indicates that conventional radiation therapy affects the tumor immunological profile in a particular manner, which in turn, might induce beneficial effects both at local and systemic (i.e., abscopal effects) levels. The interaction between radiotherapy and the immune system is being explored to combine immune and radiation (including particles) treatments, which in many cases have a greater clinical effect than any of the therapies alone. Contrary to localized, clinical irradiation, astronauts are exposed to whole body, chronic cosmic radiation, where protons and heavy ions are an important component. The effects of this extreme environment during long periods of time, e.g., a potential mission to Mars, will have an impact on the immune system that could jeopardize the health of the astronauts, hence the success of the mission. To this background, the purpose of this mini review is to briefly present the current knowledge in local and systemic immune alterations triggered by particle irradiation and to propose new lines of future research. Immune effects induced by particle radiation relevant to clinical applications will be covered, together with examples of combined radiotherapy and immunotherapy. Then, the focus will move to outer space, where the immune system alterations induced by cosmic radiation during spaceflight will be discussed.

The use of charged-particle beams, such as carbon ions, is becoming a more and more attractive treatment option for cancer therapy. Given the precise absorbed dose-localization and an increased biological effectiveness, this form of therapy is much more advantageous compared to conventional radiotherapy, and is currently being used for treatment of specific cancer types. The high ballistic accuracy of particle beams deposits the maximal dose to the tumor, while damage to the surrounding healthy tissue is limited. In order to better understand the underlying mechanisms responsible for the increased biological effectiveness, we investigated the DNA damage and repair kinetics and cell cycle progression in two p53 mutant cell lines, more specifically a prostate (PC3) and colon (Caco-2) cancer cell line, after exposure to different radiation qualities. Cells were irradiated with various absorbed doses (0, 0.5, and 2 Gy) of accelerated 13C-ions at the Grand Accélérateur National d’Ions Lourds facility (Caen, France) or with X-rays (0, 0.1, 0.5, 1, 2, and 5 Gy). Microscopic analysis of DNA double-strand breaks showed dose-dependent increases in γ-H2AX foci numbers and foci occupancy after exposure to both types of irradiation, in both cell lines. However, 24 h after exposure, residual damage was more pronounced after lower doses of carbon ion irradiation compared to X-irradiation. Flow cytometric analysis showed that carbon ion irradiation induced a permanent G2/M arrest in PC3 cells at lower doses (2 Gy) compared to X-rays (5 Gy), while in Caco-2 cells the G2/M arrest was transient after irradiation with X-rays (2 and 5 Gy) but persistent after exposure to carbon ions (2 Gy).

Reactive astrocytes in active MS lesions re-express alpha-SMA and nestin. We suggest that the in vivo re-expression might be under regulation of TGF-beta1. These results further clarify the regulation of astrocyte activity after CNS injury, which is important for the astroglial adaptation to pathological situations.