Effect of Observer's Sex and Chin Prominences on the Perception of the Lower Lip-Chin Prominence Angle.

ABSTRACT: This study aimed to identify the preferred range of lower lip-chin prominence angles in the Korean population and evaluate the effect of the individual lower lip-chin prominence angle on perceptions of esthetic chin profile.Chin prominence silhouettes were used to assess the lower lip-chin prominence preference. The observers randomly categorized each image as (1) normal, (2) slightly abnormal but not requiring surgical correction, and (3) abnormal and requiring surgery. Individual lower-chin prominence angles of all observers were analyzed using standardized clinical photographs.The normal range of lower lip-chin prominence angle is 0° to 25°; socially acceptable range is 0° to -10°, 25° to 40°; range needing surgery is -10° to -30° and 40° to 45°. Women are more tolerant to chin protrusion. A protrusive chin is more acceptable in observers with retrusive chin profile.Skeletal Class II profile is more acceptable than skeletal Class III in the Korean population. The individual lower-chin prominence angle could affect perception of desired surgery. These findings indicate that patient-specific treatment planning is important in achieving satisfaction in chin surgery.

Phytosphingosine promotes megakaryocytic differentiation of myeloid leukemia cells.

We report that phytosphingosine, a sphingolipid found in many organisms and implicated in cellular signaling, promotes megakaryocytic differentiation of myeloid leukemia cells. Specifically, phytosphingosine induced several hallmark changes associated with megakaryopoiesis from K562 and HEL cells including cell cycle arrest, cell size increase and polyploidization. We also confirmed that cell type specific markers of megakaryocytes, CD41a and CD42b are induced by phytosphingosine. Phospholipids with highly similar structures were unable to induce similar changes, indicating that the activity of phytosphingosine is highly specific. Although phytosphingosine is known to activate p38 MAPK-mediated apoptosis, the signaling mechanisms involved in megakaryopoiesis appear to be distinct. In sum, we present another model for dissecting molecular details of megakaryocytic differentiation which in large part remains obscure.

Flow cytometric detection of neutrophil-associated immunoglobulin in patients with or without neutropenia and establishment of the reference interval.

We measured neutrophil-associated immunoglobulin (NAIg) levels using flow cytometry to establish the reference interval for NAIg and to estimate NAIg in patients with or without neutropenia. Peripheral blood from 152 individuals was analyzed for NAIg detection by flow cytometry. Using fluorescescent-conjugated anti-CD10 monoclonal antibody and anti-human immunoglobulins, proportions of NAIgG, NAIgA, and NAIgM bound to neutrophils were measured. Reference intervals for NAIg were set as NAIgG <2.6%, NAIgA <3.2%, and NAIgM <3.4%, representing the 95th percentile of data from 40 healthy individuals. 63 patients with neutropenia showed positivities of 49.2% (31/63) for NAIgG, 50.0% (19/38) for NAIgA, and 42.9% for (27/63) NAIgM. The proportion of NAIgA-bound neutrophils was higher in females (median 10.7% vs 3.0%, P=0.024), and NAIgA positivity rates were increased in patients aged less than 10 years (83.3%, P=0.043). NAIg was associated with the severity of neutropenia. In particular, NAIgM levels were significantly increased in patients with severe neutropenia (P=0.019). In addition, NAIg was commonly detected in patients with autoimmune diseases, solid organ tumors, hematologic disorders, and lymphoma. Flow cytometry permitted rapid detection of NAIg in small samples. Using this method, and using the reference intervals defined herein, patients with neutropenia or adverse transfusion reactions may be evaluated in a clinically relevant manner.

Arrest defective 1 autoacetylation is a critical step in its ability to stimulate cancer cell proliferation.

The N-acetyltransferase arrest defective 1 (ARD1) is an important regulator of cell growth and differentiation that has emerged recently as a critical molecule in cancer progression. However, the regulation of the enzymatic and biological activities of human ARD1 (hARD1) in cancer is presently poorly understood. Here, we report that hARD1 undergoes autoacetylation and that this modification is essential for its functional activation. Using liquid chromatography-tandem mass spectrometry and site-directed mutational analyses, we identified K136 residue as an autoacetylation target site. K136R mutation abolished the ability of hARD1 to promote cancer cell growth in vitro and tumor xenograft growth in vivo. Mechanistic investigations revealed that hARD1 autoacetylation stimulated cyclin D1 expression through activation of the transcription factors beta-catenin and activator protein-1. Our results show that hARD1 autoacetylation is critical for its activation and its ability to stimulate cancer cell proliferation and tumorigenesis.

Detection of mutant p53 using field-effect transistor biosensor.

We assessed the abilities of wild p53 and mutant p53 proteins to interact with the consensus DNA-binding sequence using a MOSFET biosensor. This is the first report in which mutant p53 has been detected on the basis of DNA-protein interaction using a FET-type biosensor. In an effort to evaluate the performance of this protocol, we constructed the core domain of wild p53 and mutant p53 (R248W), which is DNA-binding-defective. After the immobilization of the cognate DNA to the sensing layer, wild p53 and mutant p53 were applied to the DNA-coated gate surface, and subsequently analyzed using a semiconductor analyzer. As a consequence, a significant up-shift in drain current was noted in response to wild p53, but not mutant p53, thereby indicating that sequence-specific DNA-protein interactions could be successfully monitored using a field-effect-based biosensor. These data also corresponded to the results obtained using surface plasmon resonance (SPR) measurements. Taken together, our results show that a FET-type biosensor might be promising for the monitoring of mutant p53 on the basis of its DNA-binding activity, providing us with very valuable insights into the monitoring for diseases, particularly those associated with DNA-protein binding events.

Flow cytometric detection of platelet-associated immunoglobulin in patients with immune thrombocytopenic purpura and nonimmune thrombocytopenia.

We increased the specificity of flow cytometric detection of platelet-associated immunoglobulin (PAIg) by a combination of platelet gating and cutoff for positivity determined by the use of receiver operating characteristic (ROC) curve analysis, and we evaluated the significance of elevated PAIg in non-immune thrombocytopenic purpura (ITP) patients. Blood samples from 118 patients with a platelet count <100 x 10(9)/L were used in this study. Flow cytometric detection of PAIg was performed. To obtain the cutoff of the surface-bound immunoglobulin for the discrimination of ITP and non-ITP, ROC curve analysis was used. The sensitivity of a positive PAIgG and PAIgM test for ITP in thrombocytopenic patients was 74.6%; the specificity was 79.7%; the positive predictive value 78.6%; and the negative predictive value 75.8%. Among 3 patients with myelodysplastic syndrome, 2 showed increased PAIg. Six of 20 patients with benign disease showed positivity for PAIg. Among these patients, 4 with elevated PAIg were diagnosed with liver disease. This study demonstrates that flow cytometric detection of PAIg combined with ROC curve analysis is a convenient, sensitive, and specific test, compared to previous methods, and it is useful for the differential diagnosis of thrombocytopenic patients.

Blockade of processing/activation of caspase-3 by hypoxia.

Tumor hypoxia, which is caused by the rapid proliferation of tumor cells and aberrant vasculature in tumors, results in inadequate supplies of oxygen and nutrients to tumor cells. Paradoxically, these unfavorable growth conditions benefit tumor cell survival, although the mechanism is poorly understood. We have demonstrated for the first time that hypoxia inhibits TRAIL-induced apoptosis by blocking translocation of Bax from cytosol to the mitochondria in tumor cells. However, it is largely unknown how hypoxia-inhibited Bax translocation attenuates TRAIL-induced apoptosis. Here, we demonstrate that despite its inhibitory activity in TRAIL-induced apoptosis, hypoxia does not affect TRAIL-triggered proximal apoptotic signaling events, including caspase-8 activation and Bid cleavage. Instead, hypoxia inhibited processing of caspase-3, leading to incomplete activation of the caspase. Importantly, hypoxia-blocked translocation of Bax to the mitochondria significantly inhibited releasing the mitochondrial factors, such as cytochrome c and Smac/DIABLO, to the cytosol in response to TRAIL. It is well-known that complete processing/activation of caspase-3 requires Smac/DIABLO released from mitochondria. Therefore, our data indicate that an engagement of the apoptotic mitochondrial events leading to caspase-3 activation is blocked by hypoxia. Our data shed new light on understanding of the apoptotic signal transduction and targets regulated by tumor hypoxia.

A potent reporter applicable to the monitoring of caspase-3-dependent proteolytic cleavage.

In this study, we developed a chimeric caspase-3 substrate (GST:DEVD:EGFP) comprised of glutathione-S transferase (GST) and enhanced green fluorescent protein (EGFP) with a specialized linker peptide harboring the caspase-3 cleavage sequence, DEVD. Using this reporter, we assessed the proteolytic cleavage of the artificial caspase-3 substrate for caspase-3. The common feature of this approach is that the presence of the DEVD sequence between GST and EGFP allows for caspase-3-dependent cleavage after the Asp (D) residue, resulting in the elimination of EGFP from the GST:DEVD:EGFP reporter. To the best of our knowledge, this study reports the first application employing a chimeric protein substrate, with the similar accuracy level compared to the conventional methods such as fluorometric assays. As a result, using this GST:DEVD:EGFP reporter, caspase-3 activation based on proteolytic properties could be monitored via a variety of bioanalytical techniques such as immunoblot analysis, glutathione-agarose bead assay, and on-chip visualization, providing both technical and economical advantages over the extensively utilized fluorogenic peptide assay. Our results convincingly showed that this versatile reporter (GST:DEVD:EGFP) constitutes a useful system for the monitoring of caspase-3 activation, potentially enabling the monitoring of the proteolytic activities of different intra-cellular proteases via the substitution of the cleavage sequence within the same schematic construct.

Expression, crystallization and preliminary X-ray crystallographic analyses of two N-terminal acetyltransferase-related proteins from Thermoplasma acidophilum.

N-terminal acetylation is one of the most common protein modifications in eukaryotes, occurring in approximately 80-90% of cytosolic mammalian proteins and about 50% of yeast proteins. ARD1 (arrest-defective protein 1), together with NAT1 (N-acetyltransferase protein 1) and possibly NAT5, is responsible for the NatA activity in Saccharomyces cerevisiae. In mammals, ARD1 is involved in cell proliferation, neuronal development and cancer. Interestingly, it has been reported that mouse ARD1 (mARD1(225)) mediates epsilon-acetylation of hypoxia-inducible factor 1alpha (HIF-1alpha) and thereby enhances HIF-1alpha ubiquitination and degradation. Here, the preliminary X-ray crystallographic analyses of two N-terminal acetyltransferase-related proteins encoded by the Ta0058 and Ta1140 genes of Thermoplasma acidophilum are reported. The Ta0058 protein is related to an N-terminal acetyltransferase complex ARD1 subunit, while Ta1140 is a putative N-terminal acetyltransferase-related protein. Ta0058 shows 26% amino-acid sequence identity to both mARD1(225) and human ARD1(235). The sequence identity between Ta0058 and Ta1140 is 28%. Ta0058 and Ta1140 were overexpressed in Escherichia coli fused with an N-terminal purification tag. Ta0058 was crystallized at 297 K using a reservoir solution consisting of 0.1 M sodium acetate pH 4.6, 8%(w/v) polyethylene glycol 4000 and 35%(v/v) glycerol. X-ray diffraction data were collected to 2.17 A. The Ta0058 crystals belong to space group P4(1) (or P4(3)), with unit-cell parameters a = b = 49.334, c = 70.384 A, alpha = beta = gamma = 90 degrees. The asymmetric unit contains a monomer, giving a calculated crystal volume per protein weight (V(M)) of 2.13 A(3) Da(-1) and a solvent content of 42.1%. Ta1140 was also crystallized at 297 K using a reservoir solution consisting of 0.1 M trisodium citrate pH 5.6, 20%(v/v) 2-propanol, 20%(w/v) polyethylene glycol 4000 and 0.2 M sodium chloride. X-ray diffraction data were collected to 2.40 A. The Ta1140 crystals belong to space group R3, with hexagonal unit-cell parameters a = b = 75.174, c = 179.607 A, alpha = beta = 90, gamma = 120 degrees. Two monomers are likely to be present in the asymmetric unit, with a V(M) of 2.51 A(3) Da(-1) and a solvent content of 51.0%.

Overexpression, crystallization and preliminary X-ray crystallographic analysis of a putative transposase from Thermoplasma acidophilum encoded by the Ta0474 gene.

IS200 transposases, originally identified in Salmonella typhimurium LT2, are present in many bacteria and archaea and are distinct from other groups of transposases. To facilitate further structural comparisons among IS200-like transposases, structural analysis has been initiated of a putative transposase from Thermoplasma acidophilum encoded by the Ta0474 gene. Its 137-residue polypeptide shows high levels of sequence similarity to other members of the IS200 transposase family. The protein was overexpressed in intact form in Escherichia coli and crystallized at 297 K using a reservoir solution consisting of 100 mM Na HEPES pH 7.5 and 20%(v/v) ethanol. X-ray diffraction data were collected to 1.78 A. The crystals belong to the monoclinic space group P2(1), with unit-cell parameters a = 65.00, b = 34.07, c = 121.58 A, alpha = 90, beta = 100.20, gamma = 90 degrees. Four monomers, representing two copies of a dimeric molecule, are present in the asymmetric unit, giving a crystal volume per protein weight (V(M)) of 2.02 A(3) Da(-1) and a solvent content of 39.2%.

[Fine structure of Toxoplasma gondii]

The importance of Toxoplasma gondii in human disease stimulated a number of electron microscope studies on the structure of this protozoan parasite. Gustafson et al. first studied the fine structure by means of thin sections in 1954. Many other papers havs subsequantly appeared. It is well known that Toxoplasma gondii has two stages in its life cycle-the proliferative forms and the cyst. The purpose of the electron microscopical work reported here was to study the fine structure of Toxoplasma gondii with recent techniques clarifying the correlation between the proliferative forms and cyst. RH strain and KM strain as proliferative forms on the one hand and Beverley strain as a cyst form of Toxoplasma gondii on the other hand were used throughout this study. 1) The conoid, toxoneme, nucleus, nucleolus, osmiophilic granules, mitochondria and vacuoles were found in RH strain as wsll as in KM strain and Beverley strain. 2) The endoplasmic reticulum was found in the cytoplasm of RH strain and KM strain. It was better developed in KM strain than in RH strain. 3) The outside contour of the organism of Beverley strain was somewhat irregular and toxoneme of this organism was better developed than in the other two strains. 4) Vacuoles were found in RH strain, KM strain and Beverley strain. Furthermore, tube-like bodies were observed in the vacuoles of the organism of RH strain. 5) In KM strain, two organisms of the same size were demonstrated in the leucocytes. It was presumed that they were products of longitudinal division.

[Immuno-Diffusion And Complement-Fixation Test In Paragonimiasis Of Cat]

In order to observe the complement fixation test and immuno-diffusion test of paragonimiasis, the sera taken at 10 days intervals up to 150 days from cats infected with Paragonimus westermani were examined by the above two immunological methods. The resultant findings were as follows: 1)The complement fixation test showed positive reaction 20 days after the infection with 20 metacercariae, and 40~50 days after the infection with 10 metacercariae. The highest titer was observed 110 days later following the acceleration at 80 days later. 2)In immuno-diffusion test, one are appeared 30 days after the infection with 20 metacercariae, but 60 days after the infection with 10 metacercariae. However, more than two arcs were observed since 70 days after infection. 3)A relatively wide band appeared by the antigens of Fresh worm material and Somatic material. But relatively clear precipitin lines were observed in the diffusion test with V.B.S. antigen, increasing to 3~4 arcs after 110days. 4)In general, complement fixation test showed earlier and higher sensitive reaction than immuno-diffusion test, and was considered to be more valuable method forr immunological diagnosis.