Genetic and embryological experiments have established the Caenorhabditis elegans adult hermaphrodite gonad as a paradigm for studying the control of germline development and the role of soma-germline interactions. We describe ultrastructural features relating to essential germline events and the soma-germline interactions upon which they depend, as revealed by electron and fluorescence microscopy. Gap junctions were observed between oocytes and proximal gonadal sheath cells that contract to ovulate the oocyte. These gap junctions must be evanescent since individual oocytes lose contact with sheath cells when they are ovulated. In addition, proximal sheath cells are coupled to each other by gap junctions. Within proximal sheath cells, actin/myosin bundles are anchored to the plasma membrane at plaque-like structures we have termed hemi-adherens junctions, which in turn are closely associated with the gonadal basal lamina. Gap junctions and hemi-adherens junctions are likely to function in the coordinated series of contractions required to ovulate the mature oocyte. Proximal sheath cells are fenestrated with multiple small pores forming conduits from the gonadal basal lamina to the surface of the oocyte, passing through the sheath cell. In most instances where pores occur, extracellular yolk particles penetrate the gonadal basal lamina to directly touch the underlying oocytes. Membrane-bounded yolk granules were generally not found in the sheath cytoplasm by either electron microscopy or fluorescence microscopy. Electron microscopic immunocytochemistry was used to confirm and characterize the appearance of yolk protein in cytoplasmic organelles within the oocyte and in free particles in the pseudocoelom. The primary route of yolk transport apparently proceeds from the intestine into the pseudocoelom, then through sheath pores to the surface of the oocyte, where endocytosis occurs. Scanning electron microscopy was used to directly visualize the distal tip cell which extends tentacle-like processes that directly contact distal germ cells. These distal tip cell processes are likely to play a critical role in promoting germline mitosis. Scanning electron microscopy also revealed thin filopodia extending from the distal sheath cells. Distal sheath filopodia were also visualized using a green fluorescent protein reporter gene fusion and confocal microscopy. Distal sheath filopodia may function to stretch the sheath over the distal arm.

The placental memebranes and uterus of the garter snake, Thamnophis sirtalis, were studied using histological, histochemical, electron microscopic, dye transfer, and radioisotopic techniques. The conceptuses are completely enclosed throughout gestation by a transparent shell membrane which is produced by glandular epithelia in the uterine segment of the oviduct.Both chorio-allantoic and omphalo ( yolk-sac ) placentation are observed in this snake. The growth of the extra-embryonic mesoderm takes place i n a manner peculiar to placental reptiles, and results in the isolation of the omphaloplacenta from the yolk-sac wall. On the basis of morphology, enzyme histochemistry, and phagocytosis of Trypan blue particles, this structure is interpreted as a site of histiotrophic absorption.The chorio-allantoic region of placentation is simple in structure. Fetal and maternal capillaries are closely apposed, but always separated by layers of uterine and chorionic epithelium and the thin shell membrane. The placental membranes of the garter snake are similar in many respects to those of other live-bearing snakes, but less specialized than most lizard placentate.Isotopically labelled sodium and glycine are passed to the fetus following maternal injection, the latter at least apparently via the omphaloplacenta. The permeability to iron and phospate is extremely low. On the basis of these results and the selective transfer df certain dyestuffs, -it appears dialyzing membrane.

The mechanisms by which Neisseria meningitidis establishes a carrier state or invades mucosal surfaces of the host to cause septicemia and meningitis are unknown. An experimental model of human columnar nasopharyngeal tissue in organ culture was developed, and the interaction of encapsulated, piliated N meningitidis with this mucosal surface was studied. Electron microscopic studies showed that meningococci attached selectively to nonciliated columnar cells of the nasopharynx. After attachment, the microvilli of these nonciliated cells elongated and surrounded the organisms. Six to twelve hours after infection, endocytic vacuoles containing meningococci were seen in the apical portion of some nonciliated columnar cells. Later, diplococci were seen in the subepithelial tissues adjacent to lymphoid tissue; this observation suggested that meningococci had penetrated the epithelial layer. The interaction of meningococci with the nasopharyngeal epithelium may be an important means whereby these bacteria establish a carrier state or invade the host.

In Drosophila, the Dorsal protein establishes the embryonic dorso-ventral axis during development. Here we show that the vertebrate homologue of Dorsal, nuclear factor-kappa B (NF-kappaB), is vital for the formation of the proximo-distal organizer of the developing limb bud, the apical ectodermal ridge (AER). Transcription of the NF-kappaB proto-oncogene c-rel is regulated, in part, during morphogenesis of the limb bud by AER-derived signals such as fibroblast growth factors. Interruption of NF-kappaB activity using viral-mediated delivery of an inhibitor results in a highly dysmorphic AER, reduction in overall limb size, loss of distal elements and reversal in the direction of limb outgrowth. Furthermore, inhibition of NF-kappaB activity in limb mesenchyme leads to a reduction in expression of Sonic hedgehog and Twist but derepresses expression of the bone morphogenetic protein-4 gene. These results are the first evidence that vertebrate NF-kappaB proteins act to transmit growth factor signals between the ectoderm and the underlying mesenchyme during embryonic limb formation.

Surfactant-associated protein A (SP-A) is a C-type lectin that is involved in surfactant metabolism as well as host defense functions in the lung. We have recently identified a receptor on macrophages [specific 210-kDa SP-A receptor (SPR210)] that binds SP-A. In the current study we have investigated the role of SP-A in mediating uptake of bacillus Calmette-Guérin (BCG) by rat macrophages and human monocytes and have examined the role of the macrophage SPR210 in this process. 125I-labeled SP-A bound BCG in a Ca(2+)-, carbohydrate-, and dose-dependent manner. To examine association of SP-A-BCG complexes with macrophages, BCG were opsonized with SP-A and were incubated with rat bone marrow-derived macrophages (RBMM), rat alveolar macrophages (RAM), or human monocytes at a 1-to-1 ratio for 4 h. The cells were washed, fixed in formalin, and stained with auramine-rhodamine. Cell-associated organisms were enumerated by fluorescent microscopy. The percentage of cells with one or more associated BCG was increased by SP-A from 27% of RBMM with BCG alone to 54% with SP-A-BCG complexes; 1-16% in RAM; and 39-67% in human monocytes. This enhanced uptake was dependent on the dose of SP-A, with maximal increases seen with 10 micrograms/ml. Electron microscopic analysis supported the conclusion that organisms were ingested by and not simply bound to the macrophages. Inclusion of SPR210 antibodies blocked association of SP-A-BCG complexes, suggesting a role for SPR210 in mediating the interaction of SP-A-BCG with the macrophages. This was further supported by the finding that modulation of SPR210 activity resulted in altered SP-A-BCG uptake. These results demonstrate that SP-A binds to BCG and that uptake of these SP-A-BCG complexes is mediated in part by the SPR210 on rat macrophages and human monocytes.

In Caenorhabditis elegans, specialized contractile myoepithelial cells of the somatic gonad, the gonadal sheath cells, are closely apposed to oocytes and are required for normal meiotic maturation and ovulation. Previously we found that mutations in the ceh-18 gene, which encodes a POU-class homeoprotein expressed in sheath cells, result in oocyte defects. To determine the basis for these oocyte defects, we have used time-lapse video Nomarski microscopy to observe meiotic maturation, ovulation, and early embryogenesis in ceh-18 mutants. In ceh-18 mutants sheath cell contractions are weaker, less frequent, and uncoordinated throughout the sequence of ovulation events, and ovulation is defective. Defective ovulation can result in the formation of endomitotic oocytes in the gonad, the formation of haploid embryos, and reversals in embryonic polarity. ceh-18 mutant oocytes exhibit defects prior to nuclear envelope breakdown, suggesting that they are physiologically different from the wild type. We observed delays in meiotic maturation, as well as maturation out of the normal spatial and temporal sequence, suggesting that proximal sheath cells directly or indirectly promote and spatially restrict meiotic maturation. Analysis of sheath cell differentiation in ceh-18 mutants using antibodies to proteins of the contractile apparatus reveals that although contractile proteins are expressed, the sheath cells appear disorganized. Transmission electron microscopy reveals that ceh-18 mutant sheath cells are morphologically irregular and only loosely cover oocytes. Taken together, these observations indicate that ceh-18 is a crucial determinant of sheath cell differentiation, a function required for normal meiotic maturation and ovulation.

No abstract

Mammalian uteri are unreceptive to blastocyst implantation except during a relatively brief period. The transmembrane, cell surface mucin, Muc1, is present on epithelial cells of nonreceptive uteri in various species and has been demonstrated to have antiadhesive properties. These activities of Muc1 may prevent interaction of the embryonic trophoblast cells with the uterine epithelium. A previous study indicated that Muc1 expression in the rabbit, as in primates, is up-regulated by progesterone. This response would be expected to create a nonadhesive uterine surface during the progesterone-dominated receptive phase. In the current study, Northern blot analysis was used to evaluate Muc1 messenger RNA expression in the endometrium of estrous and progesterone-treated estrous rabbits and in endometrium from different stages of pregnancy or pseudopregnancy. Steady state levels of Muc1 messenger RNA were increased 10-fold when estrous animals were treated with progesterone for 5 days. Muc1 message was elevated 2- to 6-fold over estrous levels in endometrium of pseudopregnant females and 30-fold in preimplantation stage (6.75 days postcoitum) uteri. During implantation (7.25 day postcoitum), the high level of Muc1 expression continued in nonimplantation regions, but was dramatically reduced in endometrium from implantation sites. Using immunofluorescence localization, Muc1 protein was present on the apical surface of epithelial cells of estrous, pseudopregnant (4 and 6.75 days), preimplantation (6.75 days), and implantation (7.25 day) stage uteri. At the latter stage, luminal epithelium apposed to blastocysts had a marked reduction or absence of Muc1 immunostaining. Muc1-immunoreactive cells included luminal and cryptal epithelium in pregnant/pseudopregnant uteri, whereas the glandular cells stained weakly. Short term coculture of uterine epithelial cells with trophoblastic vesicles derived from 6.75-day blastocysts also resulted in a local reduction in apical epithelial Muc1 staining. These findings demonstrate that Muc1 expression is up-regulated by progesterone in the rabbit uterine epithelium and increases incrementally during pre- and periimplantation stages. Removal of Muc1 from the epithelial surface at implantation sites is accomplished locally via signals apparently produced by the blastocyst.