Selective expansion of allogeneic regulatory T cells by hepatic stellate cells: role of endotoxin and implications for allograft tolerance

Hepatic stellate cells (HSCs) may play an important role in hepatic immune regulation by producing numerous cytokines/chemokines and expressing Ag-presenting and T cell coregulatory molecules. Due to disruption of the endothelial barrier during cold-ischemic storage and reperfusion of liver grafts, HSCs can interact directly with cells of the immune system. Endotoxin (LPS), levels of which increase in liver diseases and transplantation, stimulates the synthesis of many mediators by HSCs. We hypothesized that LPS-stimulated HSCs might promote hepatic tolerogenicity by influencing naturally occurring immunosuppressive CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs). Following their portal venous infusion, allogeneic CD4(+) T cells, including Tregs, were found closely associated with HSCs, and this association increased in LPS-treated livers. In vitro, both unstimulated and LPS-stimulated HSCs upregulated Fas (CD95) expression on conventional CD4(+) T cells and induced their apoptosis in a Fas/Fas ligand-dependent manner. By contrast, HSCs induced Treg proliferation, which required cell-cell contact and was MHC class II-dependent. This effect was augmented when HSCs were pretreated with LPS. LPS increased the expression of MHC class II, CD80, and CD86 and stimulated the production of IL-1α, IL-1β, IL-6, IL-10 and TNF-α by HSCs. Interestingly, production of IL-1α, IL-1β, IL-6, and TNF-α was strongly inhibited, but that of IL-10 enhanced in LPS-pretreated HSC/Treg cocultures. Adoptively transferred allogeneic HSCs migrated to the secondary lymphoid tissues and induced Treg expansion in lymph nodes. These data implicate endotoxin-stimulated HSCs as important immune regulators in liver transplantation by inducing selective expansion of tolerance-promoting Tregs and reducing inflammation and alloimmunity.     

When minerals become complex: an elementary introduction to superspace crystallography to describe natural-occurring incommensurately modulated structures

Abstract  Aperiodic crystals are compounds with perfect long-range order of their atomic structures, but without three-dimensional translational symmetry. In the last thirty years, incommensurate crystal structures have been discovered in many compounds from synthetic inorganic and organic compounds to proteins. With the introduction of the superspace theory it was realized that any crystal structure requiring more than three integers to index its diffraction pattern can be described as a periodic object in superspace with dimension equal to the number of the required integers. The structure observed in physical space is a three-dimensional intersection of the structure described as periodic in superspace. In recent years, the use of CCD and imaging plate systems considerably changed the sensitivity of data collection for modulated structures and, therefore, there was a need for further improvement of the methods. Today, several computer programs are able to solve and refine incommensurately modulated structures using the superspace approach. In nature, it is uncommon to find minerals having strong and sharp incommensurate satellites that could be used for a multidimensional refinement. As a very clear example, the case of natural ?kermanite, Ca₂MgSi₂O₇, which belongs to the melilite-group of minerals, will be shown in detail. The five-dimensional refinement carried out on this crystal shows that the displacive modulation of the atoms is mainly related to a variation of the Ca cation coordination. Keywords Superspace crystallography, Incommensurate structure, Modulation vector, Modulated minerals, Melilite Subject codes: G24009, P25056, P25099     

Activation of Store-Operated Calcium Entry in Airway Smooth Muscle Cells: Insight from a Mathematical Model

Intracellular dynamics of airway smooth muscle cells (ASMC) mediate ASMC contraction and proliferation, and thus play a key role in airway hyper-responsiveness (AHR) and remodelling in asthma. We evaluate the importance of store-operated entry (SOCE) in these dynamics by constructing a mathematical model of ASMC signaling based on experimental data from lung slices. The model confirms that SOCE is elicited upon sufficient depletion of the sarcoplasmic reticulum (SR), while receptor-operated entry (ROCE) is inhibited in such conditions. It also shows that SOCE can sustain agonist-induced oscillations in the absence of other influx. SOCE up-regulation may thus contribute to AHR by increasing the oscillation frequency that in turn regulates ASMC contraction. The model also provides an explanation for the failure of the SERCA pump blocker CPA to clamp the cytosolic of ASMC in lung slices, by showing that CPA is unable to maintain the SR empty of . This prediction is confirmed by experimental data from mouse lung slices, and strongly suggests that CPA only partially inhibits SERCA in ASMC.     

Ovalbumin sensitization of guinea-pigs reduces fMLP-induced calcium signal in alveolar macrophages

In this study we analyzed the N-formyl-Met-Leu-Phe (fMLP)-induced calcium signal in alveolar macrophages (AM) isolated from ovalbumin-sensitized (OA-sensitized AM) and naive (naive AM) guinea-pigs. Guinea-pigs were sensitized by subcutaneous injection of OA and AM were isolated by bronchoalveolar lavage 6 weeks thereafter. On the following day, we measured in resting and fMLP-stimulated cells: intracellular calcium concentration by fura-2 imaging analysis, forskolin-induced cyclic AMP production and superoxide dismutase inhibitable superoxide anion release of adherent AM. Resting calcium was 82+/-5.0 nM (n=217) and 144+/-9.3 nM (n=213, P<0.001) in naive and OA-sensitized AM respectively. fMLP (10(-11)-10(-7)M) induced a dose-dependent calcium increase, 10(-8)M being the maximal effective dose in both naive and OA-sensitized AM. However, at all doses tested, this fMLP effect was lower in OA-sensitized than in naive AM. While in resting condition 10(-5)M forskolin increased cyclic AMP both in naive and OA-sensitized AM, in fMPL-stimulated AM forskolin was effective only in OA-sensitized AM. Superoxide anion release measured 10 min after fMLP stimulus was higher in naive than in sensitized AM. These data suggest that the fMLP-induced intracellular signal is different in OA-sensitized AM compared to naive cells.     

Simple sequence repeat analysis of a clonally propagated species: a tool for managing a grape germplasm collection.

The USDA germplasm repositories help to preserve the genetic variability of important crop species by collecting and maintaining representative cultivars and related germplasm. Simple sequence repeat markers with high allelic diversity were used to type 41 grapevines from 40 accessions. All vines were either seedless table grape cultivars or cultivars with names similar to table grape cultivars. The proportion of shared alleles was selected as the most appropriate statistical measure of genetic distance for this population. In conjunction with morphological traits, known synonyms were confirmed and a previously unknown synonym was discovered. An alleged synonym in the literature was disproved by the DNA data. The data were consistent with known parentage, where such data were available. Two mislabeled vines in the USDA collection were identified. UPGMA grouped the cultivars loosely into three groups: a group of nine mostly Middle Eastern cultivars, a group of 22 accessions mostly from Russia and Afghanistan that were morphologically similar to 'Thompson Seedless', and a third very loose group of 11 accessions consisting mostly of eastern European wine grape cultivars. The limitations and usefulness of this type of analysis are discussed.