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Rectal transmission is one of the main routes of infection by human immunodeficiency virus type 1 (HIV-1).To efficiently study transmission mechanisms and prevention strategies, a small animal model permissive for rectal transmission of HIV is mandatory. We tested the susceptibility of RAG2 ؊/؊ ␥ c ؊/؊ mice transplanted with human cord blood hematopoietic stem cells to rectal infection with HIV. We rectally exposed these humanized mice to cell-free and cell-associated HIV. All mice remained HIV negative as assessed by plasma viral load. The same mice infected intraperitoneally showed high levels of HIV replication. In the gut-associated lymphatic tissue, we found disproportionately smaller numbers of human cells than in other lymphoid organs. This finding may explain the observed resistance to rectal transmission of HIV. To increase the numbers of local HIV target cells and the likelihood of HIV transmission, we treated mice with different proinflammatory stimuli: local application of interleukin-1␤, addition of seminal plasma to the inoculum, or induction of colitis with dextran sodium sulfate. These procedures attracted some human leukocytes, but the transmission rate was still very low. The humanized mice showed low levels of human engraftment in the intestinal tract and seem to be resistant to rectal transmission of HIV, and thus they are an unsuitable model for this application.

The requirement for Notch signaling during T-cell development has been extensively studied. Nevertheless, the developmental stage at which it is required and whether additional signaling pathways are needed are still poorly understood. By using a stromalcell-free culture system, we show that sorted double-negative 3 (DN3) thymocytes only require a Delta-like-4-induced Notch signal to differentiate into double-positive (DP) cells. This differentiation process is preTCR-a dependent. DN3 cells undergo 4-5 proliferation cycles, and the addition of the chemokine CXCL12 improves proliferation. IL-7 blocks the differentiation of DN3 cells to DP cells but not the Notch-induced proliferation of cultured DN3 cells. The impaired differentiation correlates with an inhibition of Rag-2 upregulation. Overall, the in vitro stromal-cell-free culture system presented here also provides a powerful and unique tool for studying the mechanisms involved in the positive and negative selection of T cells.Key words: Delta-like 4 and preTCR . DN3 cells . Double-positive cells . Notch IntroductionTo maintain T lymphopoiesis, progenitor cells from the BM constantly seed the thymus. In transplantation settings, it has been shown that different progenitors can home to the thymus and generate T cells [1][2][3][4][5][6][7][8][9]. However, under physiological conditions the role of each of these progenitors is still unclear. It is generally believed that the so-called thymic seeding precursor (TSP) still has the capability of generating B cells, NK cells, DCs and other myeloid cells [2,5,6,10,11]. Upon receiving the Notch signal, the differentiation potential towards the B-cell lineage of the TSP is rapidly lost [2,10,11], whereas other lineage options are still maintained [2,5,6,10,11]. However, a recent study using an IL-7Ra reporter mouse indicated that the non-T-cell lineage differentiation of these progenitors is not, or only very rarely, occurring under physiologic conditions [12,13].Within the thymus, the earliest thymocytes are characterized by the absence of CD4 and CD8 expression and are therefore called double-negative (DN) cells. Based on the expression of CD25, CD44 and CD117, DN cells can be subdivided into four subpopulations. DN1 cells express high cell surface levels of CD44 and CD117, and are negative for CD25, whereas their DN2 progeny express all three markers [14][15][16]. In vitro, DN1 and DN2 cells still possess the capacity to differentiate into NK cells, DCs and other myeloid cells [5,6,11,17] suggesting that they are not yet restricted to the T-cell lineage. T-cell commitment is achieved at the DN3 stage. However, the mechanisms operating . It is at this stage of development that the rearrangement of the TCR-b chain locus is completed, and the cells are selected for a productive TCR-b chain rearrangement, also known as the b-selection checkpoint [18][19][20]. DN3 cells that have rearranged their TCR-b chain locus unproductively can either differentiate into g/d T cells or will otherwise undergo apoptosis. DN3 cells car...

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