Air contrast of the intervillous space enables non-disruptive Micro-CT analysis of paraffin-embedded archival placental tissue.

The morphometric parameters of the villous tree are a strong indicator of deviant placentas. Methods have been established to digitally reconstruct small peripheral branches by tracing with 3D Microscopy at subcellular resolution. Micro-CT can help scale up the scanning of villous trees with resolution in the range of a few micrometers. As placental tissue samples are routinely conserved and archived by fixation and paraffin embedding, the villous structures are inaccessible to Micro-CT imaging due to poor contrast between paraffin and paraffinized tissue. We present a novel procedure for contrast enhancement by selectively replacing wax by air in the intervillous space.

NKG2-C is a receptor on human natural killer cells that recognizes structures on K562 target cells.

NKG2-C is a member of the recently discovered NKG2 family of genes and proteins, which are preferentially expressed on human natural killer (NK) cells. These potential NK cell receptors belong to a larger class of type II transmembrane proteins with a C-type lectin domain. We show here that NKG2-C is expressed as a 36-kDa glycoprotein by translation in vitro, recombinant expression and immunoprecipitation from a human NK cell clone. Further, a recombinant soluble NKG2-C-receptor binds specifically to K562 cells, which are target cells for NK cell killing, and to RPMI 8866 cells, which are feeder cells for NK cells; several other hematopoietic cell lines tested do not show any binding. The binding structures on the surface of K562 cells disappear, concomitant with a loss in susceptibility to killing when the cells are induced to differentiate with phorbol ester and Ca2+ ionophore. Our data suggest the presence of specific target molecules for NKG2-C on K562 cells, since overall glycosylation, Lewis X and Lewis Y structures, as well as the mucin-like CD43 molecule, do not change following induction of the cells. We propose that NKG2-C mediates a specific interaction of NK cells and their target cells with functional importance for NK cell killing.

Biologically active protease 3C of human rhinovirus 1A is expressed from a cloned cDNA segment in Escherichia coli.

We produced the putative protease 3C of human rhinovirus 1A (HRV-1A), a minor group rhinovirus, by Escherichia coli expression of a segment of HRV-1A cDNA coding for 3A, 3B, 3C, and parts of 2C and 3D (delta 2C3ABC delta 3D). The protease 3C was expected to be processed by intramolecular Q-G cleavages from the virus-specific precursor polypeptide. While the N-terminal 3B-3C site was correctly cleaved, the C-terminal Q-G site of 3C was not processed. Western blotting with a site-specific polyclonal antipeptide antibody showed that not the mature 3C polypeptide, but the 3C-containing precursor 3C delta 3D was the only rhinovirus-specific protein. Mature 3C was obtained by introducing two stop codons at positions glycine-1 and glutamine-2 of 3D by site-specific mutagenesis. This mutant produced the mature 3C protease of HRV-1A. In contrast to poliovirus, the mature 3C protease of HRV-1A is a minor peptide in virus-infected HeLa cells. The 3C protein can be detected only by Western blotting with a polyclonal antipeptide 3C antibody but not by radiolabeling the viral polypeptide.

Pinealocyte subsurface cisterns. I: Cytological aspects including three-dimensional structure.

Subsurface cisterns (ssc) in the pineal gland of Meriones unguiculatus have been characterized with transmission electron microscopy, freeze-fracture, morphometry, and three-dimensional reconstruction. Subsurface cisterns (which are observed only in pinealocytes and never in gliocytes) are cisterns of the endoplasmic reticulum (ER) approaching the plasmalemma at a distance of 15-20 nm. Freeze-fracture preparations show that except for some ribosomes at the cytoplasmic face, the cistern membranes as well as the related portions of the plasmalemma are free of special or specifically arranged particles or pores. All ssc have a lumen of 15-20 nm width and underlie 5.6% of the plasmalemma in a single layer; neither collapsed types nor stacks of ssc could be observed. As seen from reconstructions, large ssc are fenestrated and are situated preferentially in regions where the neighboring pinealocyte also bears large ssc. As a consequence, double-sided ssc, which can be observed in sections of (mostly large) ssc, are not a random phenomenon. In regions of the large ssc, adhering junctions are also concentrated. Tubules of 20-nm diameter link ssc with deeper parts of the ER, particularly with the perinuclear cistern and with the tubulo-cisternal network at the trans-side of the Golgi apparatus. Besides ssc proper, a lot of small endplates of ER-tubules are seen close to the plasmalemma. It is suggested that ssc form by the widening and fusing of such ER-tubules and decrease by retraction of ER-tubules from the cell surface.