The induction of baboon glycodelin expression by progesterone is not through Sp1

Glycodelin is a major secretory product of the uterine glandular epithelial cells of the human and non-human primate during the late luteal phase of the menstrual cycle and early pregnancy. Since progesterone levels are elevated during these periods we sought to determine how progesterone modulates glycodelin gene expression. Co-transfection of various deletions of the baboon glycodelin promoter with the progesterone receptor (PR) into Ishikawa cells, a human endometrial cell line, revealed that full progesterone responsiveness is retained within the region -119/+48. In COS-1 cells, a kidney cell line, progesterone failed to elevate luciferase levels when various deletion constructs and the PR were co-transfected. Mutation of the Sp1 site in the -67/+48 region lowered basal expression but did not affect the ability of progesterone to increase expression of the luciferase reporter in Ishikawa cells. These findings suggest that Sp1 sites are not involved in the progesterone regulation of the baboon glycodelin gene. We propose that progesterone induces a factor that regulates glycodelin gene expression in the uterus since we failed to obtain a similar response in a non-uterine cell line.     

Immunolocalization of Fas and Fas ligand in the ovaries of women with polycystic ovary syndrome: relationship to apoptosis

Both Fas (APO-1, CD95), an apoptosis-inducing receptor, and its ligand, Fas ligand (FasL, CD95L), have been localized to the ovary. Granulosa cell apoptosis occurs in antral follicular atresia. In polycystic ovary syndrome (PCOS), antral follicles accumulate with some atretic features. The ovarian expression of Fas and FasL was examined in PCOS by immunohistochemistry and correlated with immunodetection of apoptotic cells. Fas immunostaining was present in pre-antral follicle oocytes, some primary and secondary pre-antral follicle granulosa cells, and both granulosa and theca of antral follicles. Thecal staining persisted with advancing atresia, while granulosa staining declined. In antral follicles, abundant Fas-positive cells co-localized with scattered nuclei immunopositive for apoptosis. Ovarian vascular myocytes were strongly Fas-immunopositive. FasL immunostaining was present in pre-antral follicles in oocytes and variably in granulosa. In antral follicles, granulosa and thecal FasL staining increased with advancing atresia. Normal control ovaries showed follicular Fas and FasL staining patterns similar to those in PCOS, but vascular staining was less prominent. In one healthy follicle, Fas immunostaining was seen in the oocyte and weakly in mural granulosa and theca interna. The results suggest that in PCOS, an alteration in Fas-mediated apoptosis, does not cause abnormal folliculogenesis, but may promote ovarian vascular remodelling.     

Multiple effects of dopamine on layer V pyramidal cell excitability in rat prefrontal cortex

The mechanisms underlying the inhibitory effects of dopamine (DA) on layer V pyramidal neuron excitability in the prelimbic region of the rat medial prefrontal cortex were investigated. Under control conditions, DA depressed both action potential generation (driven by somatic current injection) and input resistance (R(N)). The presence of GABA(A) receptor antagonists blocked DA-induced depression of action potential generation and revealed a delayed increase in excitability that persisted for the duration of experimental recording, up to 20 min following the washout of DA. In contrast to spike generation, disinhibition did not affect the transient depression of R(N) produced by DA, suggesting independent actions of DA on spike generation and R(N). Consistent with the hypothesis that DA acts to decrease pyramidal cell output via a GABAergic mechanism, DA increased the frequency of spontaneous inhibitory postsynaptic currents in both the absence and presence of TTX. Furthermore focal application of GABA to a perisomatic region mimicked the inhibitory effect of DA on spike production without affecting R(N). Focal application of bicuculline to the same location reversed the inhibitory effect of bath-applied DA on spike generation, while again having no effect on R(N). The depression of R(N) by DA was both occluded and mimicked by the Na(+) channel blocker TTX, suggesting the involvement of a Na(+) conductance in reducing pyramidal cell R(N) during the acute presence of DA. Together these data demonstrate that the acute presence of DA decreases pyramidal neuron excitability by two independent mechanisms. At the same time DA triggers a delayed and longer-lasting increase in excitability that is partially masked by synaptic inhibition.     

Mantle Cell Lymphomas Lack Expression of p27kip1, a Cyclin-Dependent Kinase Inhibitor

p27^Kip1 is a cyclin-dependent kinase inhibitor that regulates the decision to enter S phase or withdraw from the cell cycle. In resting cells, the level of p27^Kip1 provides an inhibitory threshold above which G1 cyclin D/E/cyclin-dependent kinases accumulate before activation; however, in cycling cells, p27^Kip1 protein is sequestered by high levels of active cyclin D/cyclin-dependent kinase 4 complexes. As a group, the cyclin-dependent kinase inhibitors have been proposed to act as tumor suppressor genes, and several members have been implicated in the pathogenesis of a variety of human cancers. We examined p27^Kip1 expression in 116 non-Hodgkin’s lymphomas including 50 cases of MCL (40 typical and 10 blastic variants), 21 follicular lymphomas, 20 diffuse large B-cell lymphomas, 16 chronic lymphocytic leukemias, 8 marginal zone B-cell lymphomas, and 1 splenic marginal zone lymphoma, and correlated its expression with that of the proliferation marker Ki67 (MiB1) and with p53. p27^Kip1gene structure was analyzed by Southern blot in the group of MCLs. In all cases of non-Hodgkin’s lymphoma other than MCL, p27^Kip1 expression was inversely related to the proliferation index as measured by Ki67. In contrast, in typical MCL, p27^Kip1 expression was negative in 35 of 40 (88%) cases, irrespective of the proliferative rate (median 15%; range 2 to 90%). Paradoxically, in the blastic variant of MCL, 8 of 10 (80%) cases showed expression of p27^Kip1, despite a high proliferation rate (median 60%; range 32 to 100%). However, the staining in most of the cases was less intense than in the reactive T lymphocytes. Deletions of p27^Kip1gene were not found in any of the 25 cases examined. p53 expression was found in 15 of 50 cases of MCL: 7 of 10 (70%) in the blastic variant and 8 of 40 (20%) in the typical MCL (70% vs. 20%, P < 0.0045). These results demonstrate that MCLs, in contrast to other non-Hodgkin’s lymphomas and normal lymphoid tissue, fail to correlate p27^Kip1 expression with the proliferation rate. This peculiar uncoupling of p27^Kip1 protein expression from the proliferation rate may be related to the high levels of cyclin D1 expressed in MCL and is likely to have profound effects on cell cycle regulation and contribute to the pathogenesis of MCL.     

The pathology of posttransplant lymphoproliferative disorders occurring in the setting of cyclosporine A-prednisone immunosuppression.

Posttransplant lymphoproliferative disorders (PTLDs) were diagnosed in 43 patients from the Pittsburgh-Denver series between June 1980 and March 1987. This constitutes a detection rate of 1.7%. Major categories of clinical presentation included a mononucleosislike syndrome, gastrointestinal/abdominal disease, and solid organ disease. The median time of onset in patients initially immunosuppressed with cyclosporine-A (CsA)-containing regimens was 4.4 months after transplant, regardless of tumor clonality. A strong association of PTLD with Epstein-Barr virus (EBV) was observed. A histologic spectrum of lesions from polymorphic to monomorphic was observed. Whereas polymorphic lesions could be either clonal or nonclonal, monomorphic lesions appeared to be clonal in composition. The presence of large atypical cells (atypical immunoblasts) or necrosis did not appreciably worsen the prognosis. Twelve patients had clonal, 13 had nonclonal, and five had both clonal and nonclonal tumors. Clonality was indeterminate in 13 cases. Most patients were treated with a regimen based on reduced immunosuppression and supportive surgery. Almost all nonclonal and about half of the clonal lesions respond to this conservative therapy, indicating that it is an appropriate first line of treatment. This behavior suggests that a spectrum of lesions ranging from infectious mononucleosis to malignant lymphoma constitutes the entity known as PTLD. Some monoclonal tumors can undergo regression, however, apparently in response to host immune control mechanisms. Because of its short latency and strong association with EBV, PTLD is an important model for the study of virus-associated tumor progression in humans.     

Infant gaze, head, face and self-touch at 4 months differentiate secure vs. avoidant attachment at 1 year: a microanalytic approach

The study attempted to distinguish avoidant vs. secure infants at 1 year from 4-month infant behavior only, during a face-to-face play interaction with the mother. Thirty-five 4-month-old infants were coded second by second for infant gaze, head orientation, facial expression and self-touch/mouthing behavior. Mother behavior was not coded. At 1 year, 27 of these infants were classified as secure (B), and 8 as avoidant (A) attachment in the Ainsworth Strange Situation. Compared with the B infant, the future A infant spent less time paying 'focused' visual attention (a look of a minimum 2 seconds duration) to the mother's face. Only if the A infant engaged in self-touch/mouthing behavior did its focused visual attention match that of the B. Markovian t to t+1 transition matrices then showed that both for future A and for future B infants, focused visual attention on the mother constrained the movements of the head to within 60 degrees from center vis-à-vis, defining head/gaze co-ordination within an attentional-interpersonal space. However, infant maintenance of head/gaze co-ordination was associated with self-touch/mouthing behavior for the A infant but not the B. Positive affect was associated with a disruption of head/gaze co-ordination for the A but not the B. Whereas the B had more variable facial behavior, potentially providing more facial signaling for the mother, the A had more variable tactile/mouthing behavior, changing patterns of self-soothing more often. Thus, infants classified as A vs. B at 12 months showed different behavioral patterns in face-to-face play with their mothers as early as 4 months.     

Tumours of histiocytes and accessory dendritic cells: an immunohistochemical approach to classification from the International Lymphoma Study Group based on 61 cases

Neoplasms of histiocytes and dendritic cells are rare, and their phenotypic and biological definition is incomplete. Seeking to identify antigens detectable in paraffin-embedded sections that might allow a more complete, rational immunophenotypic classification of histiocytic/dendritic cell neoplasms, the International Lymphoma Study Group (ILSG) stained 61 tumours of suspected histiocytic/dendritic cell type with a panel of 15 antibodies including those reactive with histiocytes (CD68, lysozyme (LYS)), Langerhans cells (CD1a), follicular dendritic cells (FDC: CD21, CD35) and S100 protein. This analysis revealed that 57 cases (93%) fit into four major immunophenotypic groups (one histiocytic and three dendritic cell types) utilizing six markers: CD68, LYS, CD1a, S100, CD21, and CD35. The four (7%) unclassified cases were further classifiable into the above four groups using additional morphological and ultrastructural features. The four groups then included: (i) histiocytic sarcoma (n=18) with the following phenotype: CD68 (100%), LYS (94%), CD1a (0%), S100 (33%), CD21/35 (0%). The median age was 46 years. Presentation was predominantly extranodal (72%) with high mortality (58% dead of disease (DOD)). Three had systemic involvement consistent with 'malignant histiocytosis'; (ii) Langerhans cell tumour (LCT) (n=26) which expressed: CD68 (96%), LYS (42%), CD1a (100%), S100 (100%), CD21/35 (0%). There were two morphological variants: cytologically typical (n=17) designated LCT; and cytologically malignant (n=9) designated Langerhans cell sarcoma (LCS). The LCS were often not easily recognized morphologically as LC-derived, but were diagnosed based on CD1a staining. LCT and LCS differed in median age (33 versus 41 years), male:female ratio (3.7:1 versus 1:2), and death rate (31% versus 50% DOD). Four LCT patients had systemic involvement typical of Letterer-Siwe disease; (iii) follicular dendritic cell tumour/sarcoma (FDCT) (n=13) which expressed: CD68 (54%), LYS (8%), CD1a (0%), S100 (16%), FDC markers CD21/35 (100%), EMA (40%). These patients were adults (median age 65 years) with predominantly localized nodal disease (75%) and low mortality (9% DOD); (iv) interdigitating dendritic cell tumour/sarcoma (IDCT) (n=4) which expressed: CD68 (50%), LYS (25%), CD1a (0%), S100 (100%), CD21/35 (0%). The patients were adults (median 71 years) with localized nodal disease (75%) without mortality (0% DOD). In conclusion, definitive immunophenotypic classification of histiocytic and accessory cell neoplasms into four categories was possible in 93% of the cases using six antigens detected in paraffin-embedded sections. Exceptional cases (7%) were resolvable when added morphological and ultrastructural features were considered. We propose a classification combining immunophenotype and morphology with five categories, including Langerhans cell sarcoma. This simplified scheme is practical for everyday diagnostic use and should provide a framework for additional investigation of these unusual neoplasms.     

Whole-genome sequence assembly for mammalian genomes: Arachne 2

We previously described the whole-genome assembly program Arachne, presenting assemblies of simulated data for small to mid-sized genomes. Here we describe algorithmic adaptations to the program, allowing for assembly of mammalian-size genomes, and also improving the assembly of smaller genomes. Three principal changes were simultaneously made and applied to the assembly of the mouse genome, during a six-month period of development: (1) Supercontigs (scaffolds) were iteratively broken and rejoined using several criteria, yielding a 64-fold increase in length (N50), and apparent elimination of all global misjoins; (2) gaps between contigs in supercontigs were filled (partially or completely) by insertion of reads, as suggested by pairing within the supercontig, increasing the N50 contig length by 50%; (3) memory usage was reduced fourfold. The outcome of this mouse assembly and its analysis are described in (Mouse Genome Sequencing Consortium 2002).