iPSC-Based Modeling of RAG2 Severe Combined Immunodeficiency Reveals Multiple T Cell Developmental Arrests

RAG2 severe combined immune deficiency (RAG2-SCID) is a lethal disorder caused by the absence of functional T and B cells due to a differentiation block. Here, we generated induced pluripotent stem cells (iPSCs) from a RAG2-SCID patient to study the nature of the T cell developmental blockade. We observed a strongly reduced capacity to differentiate at every investigated stage of T cell development, from early CD7−CD5− to CD4+CD8+. The impaired differentiation was accompanied by an increase in CD7−CD56+CD33+ natural killer (NK) cell-like cells. T cell receptor D rearrangements were completely absent in RAG2SCID cells, whereas the rare T cell receptor B rearrangements were likely the result of illegitimate rearrangements. Repair of RAG2 restored the capacity to induce T cell receptor rearrangements, normalized T cell development, and corrected the NK cell-like phenotype. In conclusion, we succeeded in generating an iPSC-based RAG2-SCID model, which enabled the identification of previously unrecognized disorder-related T cell developmental roadblocks.In this article, Mikkers

Differential immune-response of congenic mice to ultraviolet-treated major histocompatibility complex class-II incompatible skin-grafts

The influence of ultraviolet (UVB) irradiation on the survival of H-2 class II-disparate skin grafts was studied in congenic mouse strains. Isolated skin was UVB irradiated in vitro at a dose of 40 mJ/cm2 from both sides to remove Ia immunogenicity. Immediately after irradiation the skin was transplanted onto the flank of allogeneic mice. When B10.AQR grafts were transplanted onto B10.T(6R) recipients, a significant prolongation of the survival time was observed, while 50% of the UVB-treated grafts were not rejected at all. However, in the opposite direction--i.e., B10.T(6R) grafts onto B10.AQR recipients, no significant prolongation of the survival was observed. To test whether this effect was due to a difference in susceptibility of the donor skin to UVB irradiation or to a different immune response in the recipients, (B10.T(6R) x B10.AQR) grafts were transplanted onto the parent strains. Similar results were obtained, in that UVB-treated grafts did not show a prolonged survival in B10.AQR recipients, whereas a significant prolongation (50% of the grafts survived more than 100 days) was observed in B10.T(6R) recipients. UVB-treated (B10.T(6R) x B10.AQR)F1 grafts were also transplanted onto (B10.T(6R) x C57B1/10)F1, (B10.AQR x C57B1/10)F1, (B10.T(6R) x Balb/c)F1 and (B10.AQR x Balb/c)F1 recipients--but in none of these combinations was a prolonged survival time observed. These data suggest that, in contrast to all in vitro experiments, the abrogation of the immune response by UVB treatment of the stimulator cells is, in vivo, not a general phenomenon. The genetic constitution of the responder mice seems to play an important role in determining whether or not an immune response takes place

A method for dendrochronological assessment of medium-term gully erosion rates

A method based on dendrochronology to estimate gully erosion rates was developed as an alternative of traditional methods for assessing medium-term gully retreat rates, such as field monitoring of headcuts or aerial-photo interpretation of gully retreat. The method makes use of trees or parts of a tree affected by gully erosion revealing information on the history of the erosion process by datable deviations of their normal growth pattern, hence defined as 'datable objects'. These include roots exposed by erosion; browsing scars made by ungulates on exposed roots or on above-ground parts of fallen trees; exposed and dead root ends; root suckers; stems, branches and leading shoots of fallen trees; and a sequence of trees within a gully. The method is based on the differentiation between three main conditions depending on the relation between the dynamics of the datable object (part of the tree) and the development of the gully. The first condition implies that the datable object was created before erosion of the gully volume to be dated, e.g. exposed tree roots. According to the second condition, the datable object developed as an immediate consequence of the erosion event, e.g. growth reactions of a fallen tree. The third condition implies that the datable object was created some time after the erosion event took place, e.g. trees colonising the gully bed. Each principle has consequences for the accuracy and the correct interpretation of the estimated erosion rate, i.e. whether the true erosion rate is underestimated, exact or overestimated. In spite of methodological limitations and dendrochronological dating problems, the method was successfully applied in southeast Spain. Conservative estimations of gully-head retreat rate resulted in an average medium-term (3-46 years) value of 6 m(3) year(-1) (n = 9). For gully sidewall processes, the average minimum erosion rate per unit sidewall length amounted 0.1 m(3) year(-1) m(-1) (n = 9). A strong correlation was found between the headcut retreat rate (upsilon (m(ortho)) m(3) year(-1)) and the drainage-basin area (A, m(2)) of the gullies, expressed by upsilon (m(ortho)) = 0.02 A(0.57) (R-2 = 0.93, n = 9). Comparing the findings from this study with those obtained by short-term headcut retreat monitoring suggests a high reliability of the estimated retreat rates, supporting the applicability of the developed dendrochronological method. (C) 2001 Elsevier Science B.V. All rights reserved.status: publishe

Human IG production and isotype switching in severe combined immunodeficient: human mice

Severe combined immunodeficient (SCID) mice were transplanted with different human fetal organs (SCID-hu mice), including thymus, liver, spleen, and omentum, and the serum levels of human IgM, IgG, IgE, and IgA were measured. In all SCID-hu mice significant levels (up to 590 ng/ml) of IgM were detected, irrespective of the organs transplanted. In contrast, IgG was present (up to 530 ng/ml) only when the fetal thymus was transplanted together with the fetal liver, indicating that the presence of human T cells is a prerequisite for in vivo isotype switching by human B cells in SCID-hu mice. Additional transplantation of fetal spleen did not significantly increase IgG levels. Human IgA and IgE were never detected in the serum of these SCID-hu mice. The peak of IgM and IgG production was observed 4 months after transplantation. At that time, analysis by IEF showed that human IgG present in SCID-hu serum was at least oligoclonal. Furthermore, all IgG subclasses were represented in the human IgG pool. Human B cells were undetectable in the peripheral blood, spleen, and bone marrow of-these SCID-hu mice; in contrast, B cells expressing CD19 could be isolated from the SCID-hu thymus. Considerable proportions of the CD19+ B cells coexpressed CD5, CD7, CD10, CD40, and CD2. These B cells spontaneously produced IgM and IgG in vitro and could be induced to switch to IgE-producing cells when cocultured with cloned activated CD4+ T cells in the presence of IL-4. Collectively, these data demonstrate that functionally mature B cells able to produce IgM and IgG in vivo, and IgE in vitro, are present in the SCID-hu human thymus