Natural cytotoxicity receptor 1 in mouse uNK cell maturation and function.

Early and midgestational decidua of mice genetically ablated for expression of the natural killer (NK) cell natural cytotoxicity receptor (NCR; Ncr1Gfp/Gfp mice) shows restricted angiogenesis and atypically small uterine (u)NK cells. We hypothesized that NCR1 inactivation disturbs maturation and angiokine production by uterine natural killer (uNK) cells. Using histological and morphometric approaches, we observed that Ncr1Gfp/Gfp but not control C57BL/6 (B6) implantation sites sustain immature, non-granulated uNK cells into midpregnancy. Mouse uNK cells can be subclassified by their reactivity with Dolichos biflorus agglutinin (DBA) lectin; DBA+ uNK cells with greater Ncr1 expression were investigated. DBA+ uNK cells from Ncr1Gfp/Gfp mice show delayed maturation as indicated by shorter diameters and fewer cytoplasmic granules. Granules in mature Ncr1Gfp/Gfp uNK cells are ultrastructurally abnormal and abundance of granule-associated proteins (perforin, granzyme) and of cytoplasmic proteins (vascular endothelial growth factor; placental growth factor) differs from controls. Leukocyte-leukocyte conjugate formation in gestation day 6.5 and 8.5 intact Ncr1Gfp/Gfp decidua was less frequent than in B6; however, this difference involved leukocytes other than DBA+ uNK cells. These studies strongly support roles for NCR1 and its ligands in normal pregnancy promotion.

Contemporary zebrafish transgenesis with Tol2 and application for Cre/lox recombination experiments.

Spatiotemporal transgene regulation by transgenic DNA recombinases is a central tool for reverse genetics in multicellular organisms, with excellent applications for misexpression and lineage tracing experiments. One of the most widespread technologies for this purpose is Cre recombinase-controlled lox site recombination that is attracting increasing interest in the zebrafish field. Tol2-mediated zebrafish transgenesis provides a stable platform to integrate lox cassette transgenes, while the amenability of the zebrafish embryo to drug treatments makes the model an ideal candidate for tamoxifen-inducible CreERT2 experiments. In addition, advanced transgenesis technologies such as phiC31 or CRISPR-Cas9-based knock-ins are even further promoting zebrafish transgenesis for Cre/lox applications. In this chapter, we will first introduce the basics of Cre/lox methodology, CreERT2 regulation by tamoxifen, as well as the utility of Tol2 and other contemporary transgenesis techniques for Cre/lox experiments. We will then outline in detail practical experimental steps for efficient transgenesis toward the creation of single-insertion transgenes and will introduce protocols for 4-hydroxytamoxifen-mediated CreERT2 induction to perform spatiotemporal lox transgene regulation experiments in zebrafish embryos. Last, we will discuss advanced experimental applications of Cre/lox beyond traditional lineage tracing approaches.

Receptors for non-MHC ligands contribute to uterine natural killer cell activation during pregnancy in mice.

INTRODUCTION: Activated uterine natural killer (uNK) cells are abundant in early human and mouse decidual basalis. In mice, distinct uNK cell subsets support early endothelial tip cell induction, the pruning of new vessels and initiation of spiral arterial modification. While genetic studies indicate that NK/uNK cell activation via receptors recognizing Class I MHC-derived peptides promotes human pregnancy, roles for other activation receptors expressed by NK cells, such as the aryl hydrocarbon receptor (AHR) and natural cytotoxicity receptors (NCR) are undefined in human or mouse pregnancies. METHODS: Expression of AHR and NCR1 (ortholog of human NKp46) by gestation day (gd)10.5 mouse uNK cell subsets was measured by quantitative real-time RT-PCR. Early implantation sites from mice lacking expression of either receptor were examined histologically. RESULTS: Gd10.5 uNK cell subsets, separated by reactivity to Dolichos biflorus agglutinin lectin, differed in relative transcript abundance for Ahr and Ncr1. Quantitative histology revealed that, in comparison to C57BL/6 controls, implant sites from gd10.5 Ahr(-/-) and gd6.5-12.5 UkCa:B6.Ncr1(Gfp/Gfp) mice had normal uNK cell abundance but the uNK cells were smaller than normal and unable to trigger spiral arterial remodeling. Whole mount immunohistochemistry comparisons of viable, gd6.5-8.5 Ncr1(Gfp/Gfp) and C57BL/6 implant sites revealed deficits in implant site angiogenesis and conceptus growth in Ncr1(Gfp/Gfp). DISCUSSION: In mice, activation of AHR and of NCR1 by endogenous, as yet undefined ligands, contributes to uNK cell activation/maturation and angiogenic functions during early to mid-gestation pregnancy. MHC-independent activation of uNK cells also likely makes critical contributions to human pregnancy success.

[Information obtainable from hysterosalpingography and laparoscopy and importance of tubal factor in sterile patients (author's transl)]. / Der Aussagewert von Hysterosalpingographie und Laparoskopie und die Bedeutung des Tubenfaktors bei Sterilitätspatientinnen.

Hysterosalpingography was used in 437 instances and laparoscopy in 99 to check 455 women for tubal patency who come to seek special advice for sterility. Hysterosalpingography was used together with laparoscopy, the latter second in time, on 81 women. Findings agreed in 64.2 per cent and differed in 35.8 per cent. Tubal occlusion was recorded from 106 patients (23.3 per cent). They were unilateral in 70 cases (66.04 per cent) and collateral in 36 cases (33.96 per cent). Thirty-one of the latter 106 women (29.24 per cent) with unilateral or collateral tubal occlusion turned pregnant. Yet, pregnancy occurred only to five of the above 36 women with collateral occlusion. The causes of those apparent tubal occlusions are discussed.

[Does the HLA-system enrich our possibilities within the scope of sterility diagnosis?]. / Bereichert das HLA-System unsere Möglichkeiten im Rahmen der Sterilitäts-Diagnostik?

Since the HLA antigens were discovered there were great advances especially in organ transplantation, paternity test and human genetics. --It follows the question after the relevance of HLA antibodies as a reason for infertility. --Bachner, Stolp and coworkers estimated a high frequency of HLA antibodies in infertile women. --In contrast we found in our own investigations: 367 unselected women with undesired infertility showed HLA antibodies in none of the cases.