Identification of an interaction between the m-band protein skelemin and beta-integrin subunits. Colocalization of a skelemin-like protein with beta1- and beta3-integrins in non-muscle cells

Signaling across integrins is regulated by interaction of these receptors with cytoskeletal proteins and signaling molecules. To identify molecules interacting with the cytoplasmic domain of the beta3-integrin subunit (glycoprotein IIIa), a placental cDNA library was screened in the yeast two-hybrid system. Two identical clones coding for a 96-amino acid sequence were identified. This sequence was 100% identical to a sequence in skelemin, a protein identified previously in skeletal muscle. Skelemin is a member of a superfamily of cytoskeletal proteins that contain fibronectin-type III-like motifs and immunoglobulin C2-like motifs and that regulate the organization of myosin filaments in muscle. The amino acid residues in the isolated clones encompassed C2 motifs 4 and 5 of skelemin. A recombinant skelemin protein consisting of C2 motifs 3-7 interacted with beta1- and beta3-integrin cytoplasmic domains expressed as glutathione S-transferase (GST) fusion proteins, but not with GST-beta2-integrin cytoplasmic tail or GST alone. The skelemin-binding region was in the membrane proximal cytoplasmic domains of the integrins. Full-length skelemin interacted with integrin in intact cells as demonstrated by the colocalization of hemagglutinin-tagged skelemin in Chinese hamster ovary (CHO) cells containing alphaIIbbeta3-integrin and by the finding that microinjection of C2 motif 4 of skelemin into C2C12 mouse myoblast cells caused spread cells to round up. A skelemin-like protein was detected in CHO cells, endothelial cells, and platelets, and this protein colocalized with beta1- and beta3-integrins in CHO cells. This study suggests the presence of a skelemin-like protein in non-muscle cells and provides evidence that it may be involved in linking integrins to the cytoskeleton.     

The influence of advanced age on the outcome of assisted reproduction

This study investigates within-subject variations and associations of salivary viscosities and flow rates in a test panel of healthy adults. After several practice sessions, unstimulated and stimulated whole saliva samples were collected 5 times daily (at 0800, 1100, 1400, 1700, and 2000 h) from 30 university students. There was a significant within-subject variation in viscosity and flow rate of unstimulated saliva (P<0.001). Intra-item correlations were significantly different for salivary flow rates (r= 0.82 for unstimulated, r= 0.88 for stimulated, P< 0.001) and viscosity of unstimulated saliva (r= 0.54, P< 0.05), but viscosity of stimulated saliva was different in this respect. Our results indicate that there is a significant within-subject variation in viscosity of unstimulated saliva.     

An experimental evaluation of traumatic globe rupture

PURPOSE: The purpose of this investigation was to evaluate the surgeon's ability to assess various types of globe injury, to determine the force necessary to rupture the globe with these types of injuries, and to determine typical orbital retraction forces used in the clinical setting. MATERIALS AND METHODS: Forty-four enucleated globes from recently killed cows were divided into four equal groups-one uninjured control group, one group with a through-and-through scleral laceration, another group with a subtotal scleral laceration, and the last group with an 18-gauge needle perforation. Twenty-seven boarded or board eligible oral and maxillofacial surgeons were asked to assess one sample from each of the four groups. They were then asked to retract a simulated globe on a custom-fabricated jig to determine clinical retraction forces. Ten globes from each of the four groups were then subjected to forces until rupture on an Instron 8501M mechanical testing unit. Accuracy of the clinical assessment was determined, and means and standard deviations of the retraction forces and globe rupture forces were derived. RESULTS: Through-and-through lacerations were assessed by surgeons with 100% accuracy, subtotal lacerations with 96% accuracy, uninjured globes with 74% accuracy, and perforated globes with 15% accuracy. Globe rupture occurred at 16.72+/-7.87 kg in the control group, 20.36+/-7.87 kg in the perforated group, 15.38+/-6.06 kg in the subtotal laceration group, and 4.94+/-2.56 kg in the through-and-through laceration group. Statistically significant differences (P < .001) were noted between the total laceration group and all other groups. The mean retraction force was 0.35+/-0.47 kg, which was statistically less than the force used in all of the rupture groups (P < .001). CONCLUSIONS: Severe injuries (through-and-through lacerations) were assessed with 100% accuracy by the clinicians, and less severe injuries with less accuracy. Rupture forces for globes with perforations and subtotal lacerations were no different than for the control group, but substantially less than for the total laceration group. The simulated clinical retraction forces were substantially more than the rupture forces in all of the groups, including the through-and-through laceration group.     

Determination of somatostatin receptor subtype 2 in carcinoid tumors by immunohistochemical investigation with somatostatin receptor subtype 2 antibodies

We have shown previously that expression of mRNA for somatostatin receptor subtype 2 (sst2) detected by in situ hybridization correlates to therapeutic outcome in patients with carcinoid tumors treated with somatostatin analogues. However, in situ hybridization is laborious and not practical in clinical routine work. We have, therefore, developed polyclonal antibodies directed against sst2 that may be used for immunohistochemistry on tissue specimens. The staining is specific and is highly correlated to expression of mRNA for sst2 (P < 0.01) as well as to tracer uptake at somatostatin receptor scintigraphy (P < 0.01). There is also a good correlation to the therapeutic response in carcinoid patients treated with somatostatin analogues (P < 0.05). Of 35 patients with carcinoid tumors included in this investigation, 25 stained positive with the antibodies. Twenty-two of these were investigated by somatostatin receptor scintigraphy and showed tracer uptake in metastases. An additional two patients that did not stain with the antibodies showed pathological uptake of the tracer in metastases, which might indicate binding to somatostatin receptor subtype 5. None of the 10 patients without positive immunostaining responded to somatostatin analogue treatment, whereas patients with a positive stain had a biochemical response or remained stable during treatment. Thus, these antibodies may be used to determine the presence of sst2 in carcinoid tumors and to select patients suitable for somatostatin analogue treatment. The method is easily applicable in clinical practice.     

Angiotensin II stimulates cardiac myocyte hypertrophy via paracrine release of TGF-beta 1 and endothelin-1 from fibroblasts

We examined the effects of different cytokine combinations and culture conditions on the expansion and modulation of cell surface antigens of CD34+ derived dendritic cells (DCs), the most efficient antigen-presenting cells capable of stimulating resting T cells in the primary immune response. Cells with a dendritic morphology and expressing HLA-DR, CD1a, S100 and CD83 were maximally expanded under serum-free conditions with the addition of SCF, GM-CSF, TNF-alpha, TGF-beta and Flt-3 ligand (fold increase of CD1a+ cells = 102 +/- 32 after 2 weeks of culture). CD34+ cells were also grown under continuous flow conditions in an artificial capillary system: after 14d of culture, the expansion in the total cell number was lower than that of the static cultures (3.3 +/- 2 v 18.9 +/- 4) but the percentage of CD1a+/CD83+/ CD80+ cells was considerably higher, whereas the CD14+ cells were significantly reduced (8.9 +/- 2 v 26 +/- 13). In continuous perfusion cultures, low levels of DC precursors and of LTC-IC were still present up to day 14. The DCs generated under flow conditions stimulated the mixed leucocyte reaction (MLR) more than the cells grown in static cultures. By electron microscopy, cells grown in the continuous flow system showed an increased number of large cells with numerous dendritic processes and abundant multilamellar complexes. The cells expanded under these conditions were sorted on the basis of their light-scatter properties into two fractions: one containing a predominance of CD1a+/S100+/ CD8 3+/CD80+/CD14- 'large cells' with great internal complexity (mature DCs); the second including 'small cells' either CD33+/CD14+, CD33+/CD15+ or CD33+/CD13-/CD14. The DCs generated and selected with this method are therefore particularly well suited for immunotherapeutic protocols.     

Outplacement: Client characteristics and outcomes.

Outplacement firms developed in response to corporate downsizings to assist former upper-level employees in seeking reemployment. An important question for practitioners who assist clients in outplacement is how the personal characteristics that clients bring to the situation influence their success in finding a new job. The present archival study examined demographic variables, previous job history, and personality characteristics as predictors of outplacement outcomes. Outplacement duration was predicted by previous income, and new earnings were predicted by previous earnings, sex, and years with previous employer. The need for prospective, longitudinal research that examines the coping process in outplacement is discussed. (PsycINFO Database Record (c) 2010 APA, all rights reserved)