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Protein-protein interactions are essential to cellular mechanisms at all levels in biologically responsive systems. These interactions occur extracellularly and include ligand-receptor interactions, cell adhesion, antigen recognition, and virus-host recognition. Intracellular protein-protein interactions occur in the formation of multi-protein complexes, during the assembly of cytoskeletal elements, and between receptor-effector, as well as effector-effector, molecules of signal transduction pathways. Finally, assembly of transcriptional machinery involves protein interactions. The yeast two-hybrid method is a powerful technique for analyzing these protein-protein interactions. Since the publication of this technique in the late 1980s, the robust nature and far-reaching utility of yeast two-hybrid systems for functional expression library cloning has led to the identification of many novel proteins in all areas of biological life science research. Additionally, two-hybrid techniques provide a rapid and versatile system for the further characterization of discrete protein-protein interactions. Recent variations on the basic system have enabled application well beyond protein pairs, to investigate multi-protein complexes and protein-nucleotide interactions. Yeast two-hybrid methods necessitate expression and subsequent interaction between a "protein of interest" functional pair within the yeast cell, ultimately driving reporter gene expression and thus effectively linking protein-protein interaction(s) to a change in yeast cell phenotype. Functional protein-protein interactions using the two-hybrid techniques have been demonstrated for all levels of cellular biology; however, until recently, extracellular protein-protein interactions were excluded from investigations using this technique. Investigations from several labs have now demonstrated that extracellular proteins can be studied using two-hybrid methods, thereby enabling intense study of extracellular protein partners using the robust nature and the genetic power of yeast.

Gene expression profiling (GEP) has stratified diffuse large B-cell lymphoma (DLBCL) into molecular subgroups that correspond to different stages of lymphocyte development-namely germinal center B-cell-like and activated B-cell-like. This classification has prognostic significance, but GEP is expensive and not readily applicable into daily practice, which has lead to immunohistochemical algorithms proposed as a surrogate for GEP analysis. We assembled tissue microarrays from 475 de novo DLBCL patients who were treated with rituximab-CHOP chemotherapy. All cases were successfully profiled by GEP on formalin-fixed, paraffin-embedded tissue samples. Sections were stained with antibodies reactive with CD10, GCET1, FOXP1, MUM1, and BCL6 and cases were classified following a rationale of sequential steps of differentiation of B-cells. Cutoffs for each marker were obtained using receiver operating characteristic curves, obviating the need for any arbitrary method. An algorithm based on the expression of CD10, FOXP1, and BCL6 was developed that had a simpler structure than other recently proposed algorithms and 92.6% concordance with GEP. In multivariate analysis, both the International Prognostic Index and our proposed algorithm were significant independent predictors of progression-free and overall survival. In conclusion, this algorithm effectively predicts prognosis of DLBCL patients matching GEP subgroups in the era of rituximab therapy.Leukemia accepted article preview online, 22 March 2012; doi:10.1038/leu.2012.83.

Epitaxial YBa2Cu3O7−δ thin films grown by laser ablation on MgO (100) substrates were investigated for microwave applications. By systematically varying the growth conditions, we obtained films with various microstructures, low‐frequency superconducting properties, and microwave performance. The surface resistances were determined from a measured unloaded quality factor in a 8.6‐GHz microstrip resonator. Surface resistance was found to correlate most directly with the degree of grain alignment as revealed by electron channeling and x‐ray diffraction studies. Films grown at optimal conditions gave a scaled surface resistance of 0.6 mΩ at 77 K and 10 GHz.

The Tl oxidation state in the precursor of Tl2Ba2CaCu2O8 superconducting films plays an important role in the post-deposition process and has been carefully examined by X-ray photoelectron spectroscopy (XPS). The Tl4f 5 2 and Tl4f 5 2 peaks in the XPS spectra have been deconvoluted into several peaks corresponding to different oxidation states. Under normal deposition conditions for laser ablation, it is found that mixtures of Tl-I(+ 1 oxidation state) and Tl-III (+ 3) are present with various percentages. The study of time evolution and depth profile data suggests that the surface oxidation state is a function of time and 200 A of film should be removed before collecting data. The Tl-oxidation state depends strongly on the oxygen background pressure during deposition and the optimal pressure is found to be about 180 mTorr. Negatively charged Tl (presumably due to alloying with other metal constituents) is found when deposited with no background oxygen pressure.

During gestation, pigs have constant circulating levels of prolactin (PRL), and lack decidual PRL and placental lactogens. Effects of PRL on uterine physiology In pigs may be due to changes in endometrial PRL receptors. In this study, effects of the conceptus and cyclic hormonal environment on endometrial PRL receptors were investigated. Endometrial PHI receptor numbers were similar between Days 8 and 15 for cyclic gilts. In contrast, endometrial PHI receptor numbers for pregnant gilts were similar between Days 8 and 11, then increased (p < 0.05) on Day 12 and remained elevated between Days 14 and 30. This day-bystatus Interaction approached significance (p < 0.06) and overall receptor numbers were higher (p

We have fabricated and measured 5 GHz microstrip resonators from a series of YBa2Cu3O7−δ thin films grown on LaAlO3(001) substrates by in situ laser ablation. We have studied the correlations between unloaded quality factor and various film properties, such as transition temperature, width of transition, critical current density, narrowness of x‐ray rocking curve, sharpness of electron channeling pattern, and most important substrate temperature during growth. We found that in general, higher transition temperature, higher critical current density, sharper transition, sharper channeling pattern, and narrower x‐ray rocking curve correlate positively with good microwave performance. The best quality factor exists for a narrow growth temperature window (around 800 °C). We also report the dependence of quality factor on device power for each film.

A systematic study of the surface morphology and chemical composition of several ceramic targets after pulsed laser ablation has been conducted using a scanning electron microscope with x-ray energy dispersive spectroscopy. We have examined single crystal LaAIO3 and NdGaO3 disks, as well as powder sintered LaAIO3 and SrTiO3 discs. These targets were rotated during laser ablation and concentric trenches were formed with distinctive surface morphologies due to different doses of laser energy. We observed varied light-solid surface interactions on the targets prepared by different methods. The sintered targets exhibited surface explosions after long period laser ablation, presumably due to sub-surface overheating [1]. The single crystal targets have smoother surface morphology and the resulting films have much fewer particles on the surface. As such, they are preferred for laser ablation applications.

During gestation, pigs have constant circulating levels of prolactin (PRL), and lack decidual PRL and placental lactogens. Effects of PRL on uterine physiology in pigs may be due to changes in endometrial PRL receptors. In this study, effects of the conceptus and cyclic hormonal environment on endometrial PRL receptors were investigated. Endometrial PRL receptor numbers were similar between Days 8 and 15 for cyclic gilts. In contrast, endometrial PRL receptor numbers for pregnant gilts were similar between Days 8 and 11, then increased (p less than 0.05) on Day 12 and remained elevated between Days 14 and 30. This day-by-status interaction approached significance (p less than 0.06) and overall receptor numbers were higher (p less than 0.01) for pregnant than for cyclic gilts. Pig conceptuses secrete estrogen between Days 11 and 12; therefore, regulation of endometrial PRL receptors by acute administration of estradiol was investigated. Uterine flushings and endometrium were collected from one uterine horn of cyclic gilts on Day 11; then, at 1, 6, 12, and 24 h following a single injection of estradiol valerate (EV; 5 mg, into adipose tissue), uterine flushings and endometrium were collected from the second uterine horn. Endometrial PRL receptor numbers were higher (p less than 0.05) at both 1 h and 6 h after treatment with EV and then decreased (p less than 0.02) by 12 h to below pretreatment values. In uterine flushings, total recoverable protein (p less than 0.05), uteroferrin (p less than 0.01), leucine aminopeptidase (p less than 0.05), calcium (p less than 0.03), sodium (p less than 0.01), and potassium (p less than 0.05) increased between 12 and 24 h following EV treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Establishment of pregnancy in pigs requires a shift in endometrial prostaglandin (PG) F secretion from an endocrine (toward the myometrium and uterine vasculature) to an exocrine (toward the uterine lumen) orientation. Three experiments utilized bilateral endometrial perfusion devices for separate perifusion of myometrial and luminal surfaces to determine whether promoting calcium cycling across the luminal epithelial surface, which may be induced by interactive effects of estradiol and PRL, is involved in the reorientation of PGF secretion during establishment of pregnancy in pigs. In Exp 1, endometrium from cyclic gilts (n = 7) on day 14 after estrus was perifused with either saline (control) or calcium ionophore A23187 added to luminal surface perifusion buffer. In Exp 2, cyclic gilts (n = 7) at day 11 after estrus received an im injection of estradiol valerate (EV) after unilateral hysterectomy and the remaining EV-stimulated uterine horn removed at 6 h (n = 3) or 12 h (n = 4) after EV. Both surfaces of these endometrial samples were perifused with buffer containing either 250 ng/ml BSA or porcine PRL. In Exp 3, endometrium from cyclic (n = 3), pregnant (n = 4), and EV-induced pseudopregnant (n = 4) gilts was collected on day 14 after estrus and perifused with BSA or PRL to both surfaces. Orientation of PGF secretion was initially endocrine in Exps 1 and 2 (P less than 0.01). In Exp 1, calcium ionophore shifted the orientation of PG secretion from endocrine to exocrine (P less than 0.01). In Exp 2, neither EV in vivo nor PRL in vitro alone altered the orientation of PGF secretion. However, EV and PRL interacted to reorient secretion of PGF from endocrine to exocrine at 6 h (P less than 0.01) and 12 h (P less than 0.01) after EV. In Exp 3, the orientation of PGF secretion was endocrine in cyclic gilts and exocrine in pregnant and pseudopregnant gilts (status x side; P less than 0.05). PRL did not alter the orientation of PGF secretion regardless of reproductive status. These results suggest that the reorientation of endometrial PG secretion in pigs during the establishment of pregnancy involves interactive effects of estrogens and PRL, possibly through increased calcium cycling across the uterine epithelium.

High Tc superconducting (HTSC) films synthesis for demanding applications requires epitaxial c-axis growth without high angle grain boundaries. Electron channeling pattern (ECP) analysis using a scanning electron microscope is a non-destructive, fast, direct,and economical tool to determine the orientation and crystallinity of epitaxial films. We have successfully employed this technique to examine our in-situ laser ablated YBa2Cu307−δ superconducting thin films grown on LaAI03 (100) substrates. By changing the energy of the incident electron beam, we can also monitor the film quality vs. depth. The surface bending of YBa2Cu3O7−δ thin films due to the cubic-rhombohedral transition of LaAI03 substrates is also visiblewhen sharp ECPs are present. For films with rough surfaces, the ECP contrast is obscured by unwanted contrast arising from surface features. This unwanted contrast can be minimized by defocusing the electron beam.

A twin-free substrate material, CaYAlO4, was synthesized and used for the epitaxial growth of single crystal YBa2Cu3O7-δ and Tl2Ba2CaCu2O8 thin films for the first time. The lattice parameters of ttus new substrate are a=b=3.648 Å and c=11.89 Å. The dielectric properties, similar to those of LaAlO3, are 20(dielectric constant) and 4×10-5 (loss tangent) at 5 GHz and 77 K. Initial attempts at film growth on this substrate indicate degradation of both superconducting and microwave properties possibly due to interdiffusion.

High-quality epitaxial Tl2Ba2CaCu2O8 superconducting thin films have been grown on NdGaO3(001) by a two step process. One film, annealed at 850°C, shows interdiffusion of Ga by secondary ion mass spectroscopy, resulting in an incomplete AC susceptibility transition at 80 K. For films annealed at lower temperature, excellent crystallinity and transport properties were obtained. Ground plane substitution of the film in a 5 GHz microstrip resonator indicated worse microwave performance than similar films on LaAlO3(001) substrates. The 2.3 GHz resonator made from the film on NdGaO3 gave significantly worse quality factor (Q) values (1850) than those made on LaAlO3.

High-quality YBa2Cu3O7-δ thin films have been grown on NdGaO3(001) substrates by in situ laser ablation. The best transition temperature determined by AC susceptibility is 88.9 K and transition width is as narrow as 0.4 K. Critical current density of 1.2×106 A/cm2 at 77 K was obtained. However, the loss tangent of this substrate is measured to be 3×10-4 at 77 K and 5 GHz which is ten times poorer than that of LaAlO3. A 2.3 GHz resonator made from films on NdGaO3 exhibited a low power Q of 4500 which is very close to the calculated limit based on the loss tangent data. The microwave performance is thus severely limited by the substrate loss in our test geometries.

High-quality epitaxial Tl2Ba2CaCu2O8 superconducting thin films have been grown on CaNdAlO4 (001) substrates by a two step process. Analysis results from secondary ion mass spectroscopy and X-ray energy dispersive spectroscopy of a film annealed at 850°C indicated non-uniform Nd and Ca interdiffusion from the substrate into the film. A second film, annealed at 800°C, exhibited better superconducting properties with a T c of 100.7 K, transition width of 2.5 K, and J c of 3.5×105 A/cm2 at 77 K. Microwave measurement at 77 K of a 2.3 GHz microstrip resonator made from the 800°C annealed film on a CaNdAlO4 substrate exhibited a low power Q of 2,100.

High-quality YBa2Cu3O7-δ thin films have been grown on CaNdAlO4 (001) substrates by in situ laser ablation. The best transition temperature determined by AC susceptibility midpoint was 88.3 K with a critical current density of 1.0×106 A/cm2 at 77 K. The full width at half maximum of the X-ray rocking curve of the YBCO (004) peak was 0.33°. The film crystallinity was slightly worse than those grown on LaAlO3 and NdGaO3 judging from electron channeling pattern. A 2.3 GHz resonator made from a YBCO film on CaNdAlO4 exhibited a low power Q of 3,945 which is worse than films on LaAlO3 (Q∼18,000) and also lower than the expected value from loss tangent data.

This study investigated acute effects of 17 beta-estradiol (E) on ions, cyclic nucleotides, and catecholamines and their association with temporal changes in uterine secretory products in pregnant, cyclic, and nonpregnant gilts. Uterine flushings (UTF) and endometrium (ENDOM) from one uterine horn of nonpregnant and pregnant gilts (n = 9) were collected on days 10, 12, and 14 (n = 3). Protein, plasma inhibitor (P less than 0.05), Na+, and K+ (P less than 0.01) increased linearly in UTF of pregnant gilts. Ca2+ changed biphasically, with higher concentrations (P less than 0.01) in pregnant gilts on day 12. Endometrial cAMP and cGMP (P less than 0.05) increased between days 12 and 14 of pregnancy. The UTF norepinephrine (NE) concentrations increased (P less than 0.01) in cyclic gilts between days 12 and 14, while endometrial NE increased between days 10 and 12 and then decreased on day 14. The UTF of pregnant gilts had higher (P less than 0.05) concentrations of dopamine (DA), which peaked on day 12 and then decreased (P less than 0.01) by day 14. DA in UTF of nonpregnant gilts decreased between days 10 and 12 and remained low on day 14. A catecholamine metabolite, 3,4-dihydroxyphenylglycol, in ENDOM (P less than 0.05) and UTF (P less than 0.01) decreased linearly between days 10 and 14. The turnover rate of endometrial catecholamines was 2.4-fold higher (P less than 0.05) during early pregnancy compared with that in cyclic gilts. Effects of acute administration of exogenous E (0.5 mg) were also studied. The UTF and ENDOM were from day 11 nonpregnant gilts 0, 30, 60, and 360 min (n = 3) post-E or post-saline-ethanol (C) injection (n = 9). Potassium, plasma inhibitor (P less than 0.01), and cGMP (P less than 0.05) increased rapidly (30 min) after E injection. The K+ level changed biphasically, with increased concentrations again at 360 min. Plasmin inhibitor returned to levels similar to controls by 60 min, whereas cGMP remained elevated until 360 min postinjection when E, Ca2+, K+ (P less than 0.01), and Na+ (P less than 0.03) were increased. ENDOM NE (P less than 0.05) and UTF DA (P less than 0.05) were lower in E-treated gilts. Additionally, the turnover rate was significantly lower (P less than 0.05) for NE and DA in the UTF of E-treated gilts. An increase in Ca2+ in UTF preceded increases in protein secretions (except plasmin inhibitor) and may play a role in E-stimulated endometrial epithelial secretory vesicle exocytosis.(ABSTRACT TRUNCATED AT 400 WORDS)

Hypoprolactinaemia was induced by bromocriptine (CB154; 100 mg/day) which decreased circulating prolactin by 40% (P less than 0.06), but did not affect conceptus survival at Day 25 when administered on Days 10-16 when compared to saline:ethanol-treated control gilts. Bromocriptine or vehicle was administered to cyclic gilts on Days 10-11, oestradiol valerate was injected on Day 11 and uterine flushings were collected on Day 12. Total recoverable protein and uteroferrin in uterine flushings were not affected by treatment. However, leucine aminopeptidase activity (P less than 0.02) and total recoverable Ca2+, Na+, K+ and Cl- (P less than 0.05) were decreased in uterine flushings of gilts that received bromocriptine, suggesting that hypoprolactinaemia decreased general secretory activity of the endometrial epithelium and modulated ionic changes, respectively, in the uterine environment of pigs. Subcutaneous administration of pig prolactin (1 mg/12 h) increased (P less than 0.001) serum prolactin 4.5-fold. The interaction between hyperprolactinaemia and progesterone, without oestrogen, on components of uterine flushings were determined using gilts that received progesterone (200 mg/day) and prolactin or saline on Days 4-14 after ovariectomy on Day 4. On Day 15, there were no differences (P greater than 0.05) in any of the uterine secretory components measured. Hyperprolactinaemia (1 mg pig prolactin on Days 6-11) enhanced overall uterine secretory response on Day 12 to oestradiol (5 mg) administered on Day 11 compared to gilts that received 1 ml saline on Days 6-11 of the oestrous cycle. Total recoverable protein and leucine aminopeptidase activity were greater (P less than 0.05) for oestradiol-treated gilts, but effects of prolactin were not significant. Total recoverable glucose (P less than 0.01), PGF-2 alpha (P less than 0.02), uteroferrin (P less than 0.01) and specific activity of uteroferrin (P less than 0.001) were increased by prolactin and oestradiol, but not oestradiol alone. Calcium (P less than 0.05), chloride (P less than 0.05) and potassium (P less than 0.01) were increased in response to oestradiol. These results indicate an interaction between oestradiol and prolactin, but not progesterone and prolactin, which enhances secretion of some products of the pig uterine endometrium.

Ethyl acetate appears to be a satisfactory subsitute solvent for diethyl ether in the Formalin-ether sedimentation technique. In comparative studies, concentration of organisms with ethyl acetate was equal to or greater than that with diethyl ether. No distortion or alteration of morphology was observed with eigher solvent, and preparations were comparable in appearance and ease of examination. In addition, ethyl acetate is less flammable and less hazardous to use than diethyl ether.