Dipole-moment-induced supramolecular assembly of a donor-acceptor-type molecule on a metal surface and in a crystal.

The conformation and alignment of molecules in organic materials are important because they affect the materials' bulk physical properties. Because two-dimensional (2D) materials offer a simpler model of three-dimensional (3D) materials, the conformation and alignment of molecules in 2D assemblies have been investigated at the atomic scale by scanning tunnelling microscopy (STM). However, differences in the conformation and alignment of molecules between 2D and 3D assemblies have not been clarified. In this work, the conformation and alignment of a donor-acceptor-type molecule, 4-(3,3-dimethyl-2,3-dihydro-1H-indol-1-yl)benzonitrile (IBN), are studied in 2D and 3D assemblies. Thus, the 2D assembly of IBN on the Au(111) surface was investigated by STM and the 3D assembly of IBN in a single crystal was investigated by X-ray crystallography. Our survey revealed that the conformation of IBN is planar in both 2D and 3D assemblies because of the electron-delocalised structure resulting from the electron-donating and electron-accepting groups of IBN; thus, the values of the dipole moment of IBN in 2D and 3D assemblies are essentially the same. In both the 2D and 3D assemblies, IBN molecules align to cancel out the dipole moment even though the self-assembled structures differ. In the 2D assemblies, the orientation and self-assembled structure of IBN are changed by the surface density of IBN, and they are affected by the crystal orientation and superstructure of Au(111) because of the strong interaction between IBN and Au(111). In addition, scanning tunnelling spectroscopy revealed that the coordination structure is not included in the self-assembled structure of IBN on Au(111).

An atomic resolution scanning tunneling microscope that applies external tensile stress and strain in an ultrahigh vacuum.

We have developed an ultrahigh vacuum scanning tunneling microscope with an in situ external stress application capability in order to determine the effects of stress and strain on surface atomistic structures. It is necessary to understand these effects because controlling them will be a key technology that will very likely be used in future nanometer-scale fabrication processes. We used our microscope to demonstrate atomic resolution imaging under external tensile stress and strain on the surfaces of wafers of Si(111) and Si(001). We also successfully observed domain redistribution induced by applying uniaxial stress at an elevated temperature on the surface of a wafer of vicinal Si(100). We confirmed that domains for which an applied tensile stress is directed along the dimer bond become less stable and shrink. This suggests that it may be feasible to fabricate single domain surfaces in a process that controls surface stress and strain.

[A survey on the dioxin level in breast milk in coastal and inland region].

OBJECTIVE: To study the dioxin level of breast milk among Chinese mothers, and to assess the dioxin intake of new-born babies from mother's milk and compare with the Tolerable Daily Intake (TDI) of dioxin. METHODS: The CALUX bioassay was used to detect the dioxin concentration of the first time mother's milk among the inland samples (Shenyang region; 32 cases) and the coastal city samples (Dalian region; 47 cases). RESULTS: The median value of the dioxin Toxic Equivalence (TEQ) in breast milk in the Dalian region was 15.84 pg TEQs.g(-1) fat, which was significantly higher than that in the Shenyang region 7.21 pg TEQs.g(-1) fat (P < 0.01). CONCLUSION: The dioxin level in breast milk in Chinese is at the world's average level. The dioxin intake of the new-born babies during the period of lactation was higher than the lowest limit of the Tolerable Daily Intake (TDI) proposed by WHO. This situation should be noticed by the related authorities.

Geographical north-south decline in DNASE1*2 in Japanese populations.

Allele frequencies for human deoxyribonuclease I (DNase I) phenotypes were determined using blood samples from about 2000 Japanese subjects living in nine prefectures, and compared with one another. DNase I phenotyping was performed principally using isoelectric focusing electrophoresis and activity staining. The DNase I system was shown to have enhanced potential for anthropologic, genetic, and clinical studies of Japanese populations. DNase I phenotypes were analyzed to evaluate the degree of genetic variation at the DNASE1 locus. Our examination of DNase I types revealed a decreasing north-to-south gradient in the DNASE1 allele.

Allele frequencies of eight STRs in Japanese and Chinese.

Allele frequencies for the eight STR loci Hum-CSF1P0, F13A01, F13B, FES/FPS, LPL, TH01, TPOX and VWA were investigated in Japanese and Chinese populations. No significant deviations from Hardy-Weinberg equilibrium could be found for all loci. In the Japanese population VWA, CSF1PO, TH01, FES/FPS and TPOX were found to be useful for forensic applications and in the Chinese population, VWA, CSF1PO, TH01 and TPOX were found to be useful. Allele distributions were similar between both populations except for FES/FPS.

[Effect of postmortem change on detection of apoptosis in rats].

Terminal deoxynucleotidyl transferase(TdT)-mediated dUTP-digoxigenin nick end labeling (TUNEL) assay is useful to detect apoptotic cells in situ. We examined by hematoxylin-eosin (H-E) and TUNEL assay whether or not postmortem delay affects the development of apoptotic signals of cells in various organs. Wistar Imamichi rats were radiated by X-ray and sacrificed six hours after radiation. The spleen, thymus, adrenal and testis were excised and kept in a moist chamber at room temperature. Each tissue was fixed after different time intervals 0, 6, 12, 24 hours and paraffin-embedded sections were made. In the no-radiation group, a few of TUNEL positive cells were observed in the spleen, thymus and testis sections, but not in the adrenal. No increase in the number of apoptotic cells was observed with postmortem delay. In the radiation group, we observed in the spleen and thymus, much increase in the number of TUNEL positive cells, of which nuclei were clearly and deeply stained, corresponding to the area where shrinking nuclei were observed in H-E section. In testis sections, there was a little increase in the number of positively stained cells, and no change was observed in H-E section. With postmortem delay, the margin of the TUNEL positive cells changed from clear to indistinct, and the positive area was spread around. Our results show that it is difficult to distinguish apoptotic cells from postmortem change. It is possible, however, to detect TUNEL positive cell together with postmortem changes as the spread of the TUNEL positive area after 24 hours postmortem delay. It is important to consider the effect of the postmortem change when we adapt TUNEL assay to autopsy cases.

[Simple and rapid detection of HLA-DRB polymorphism from forensic samples].

Simple and rapid detection of HLA-DRB polymorphism has been performed using AMPLICOR HLA-DRB Typing Kit. We tried to apply this kit to various forensic samples. When DNA was extracted from the forensic samples using conventional phenol-chloroform method, addition of 7.5 mM MgCl2 was required to PCR amplification. HLA-DRB types were detected from DNA more than 0.1 ng by PCR amplification. Typing of unrelated 50 Japanese showed 38 different patterns, of which 30 patterns occurred once in the group. A total of 16 serotypes were deduced from the HLA-DRB DNA types. Out of them, high frequency serotypes were DR4 (24%), DR9 (18%) and DR15/16 (14%). This kit was very useful in forensic cases such as rape and in paternity cases. When we tried to detect HLA-DRB types from a single hair shaft of 3 cm in length, we were successful in detection from only one of five persons.

[Spreadsheets of a conventional application software for calculation of plausibility of paternity: application to parentage testing with highly polymorphic markers in deceased party].

We designed a spreadsheet package for the computation of plausibility of paternity, that can cope with highly polymorphic genetic markers and cases of deceased parties. The application program is Microsoft EXCEL, which is one of the best-selling spreadsheet software running on both Microsoft Windows and Macintosh OS. Komatsu's formula for paternity testing was mainly employed in the spreadsheet package. Probability of the mother-child-alleged father combination was calculated using "IF" function to compare the members' genotypes, whereas "VLOOKUP (or HLOOKUP)" function was employed to refer to a list of genes and their frequencies. In case of a phenotype consisting of several genotypes, the list of phenotypes versus genotypes was also given, to which the function referred. To extend these spreadsheets available for the test of deceased party, additional sheets were also created to estimate frequencies of alleged father's possible genotypes. These probabilities were calculated on the basis of types of his parents and siblings, those of his wife and their biological children, and those of both. This package would be cut out to compute the probability of paternity with extremely polymorphic loci with gentle user interface. Calculation time is satisfactorily short, although it requires considerably large disk space in some extremely complicated cases. Japanese version of this package is freely available at anonymous FTP site of the Department of Forensic Medicine, Tohoku University School of Medicine.

Polygalacturonase isolated from the culture of the psychrophilic fungus Sclerotinia borealis.

A polygalacturonase was isolated from the culture medium of Sclerotinia borealis, a psychrophilic fungus that grows on lawn and wheat seedling under the snow in winter and induces the snow mold disease. Pectic acid was a better substrate of this enzyme than pectin when the activity was determined by measuring the reducing sugar produced. However, when the activity was measured by viscosity change, the viscosity of pectin decreased more rapidly than that of pectic acid. The results of viscosity change apparently indicate that the polygalacturonase catalyzes pectin hydrolysis as an endo-type enzyme. Highly methyl-esterified pectin was a poor substrate, as determined by measurements of reducing sugar production and viscosity change. It is suggested from the results that the methoxy group of pectin affects the polygalacturonase reaction. A reaction mechanism was proposed for the polygalacturonase reaction. Molecular mass of this enzyme was 40 kDa and its isoelectric point was pH 7.5. Optimum pH of the enzyme reaction was 4.5 and its optimum temperature was 40-50 degrees C. Thirty percent of the maximum activity was observed at 5 degrees C, but it was only slightly active above 60 degrees C. The activity was preserved for more than 2 years at 5 degrees C and pH 4.5, but it was lost when kept at room temperature overnight or heated at 50 degrees C for 30 min. The amino acid sequence of the N-terminal region of the psychrophilic polygalacturonase of Sclerotinia borealis is compared with those of polygalacturonases of mesophilic fungi. The function of this enzyme against the target plants is discussed with reference to the reaction of polygalacturonases of mesophilic fungi.

ABO genotyping by PCR-direct sequencing.

The PCR-direct sequence method was applied to ABO genotyping. At the 261st nucleotide of the genes of A and B glycosyltrasferase, it was easily detected that the nucleotide was guanine in AA, AB and BB genotypes and that the nucleotide was ademine in only OO. In AO and BO, substitution of A to G was confirmed by the dye primer method, but it was difficult to detect correctly by the dye terminator method. At the 297th, nucleotide substitution between A and B alleles was confirmed by the both methods. As this position, O allele was subdivided into three types, OAOA, OGOG and OAOG. At the 703rd, nucleotide substitution between A and B alleles was easily detected by the both methods. The PCR-direct sequence method was suitable to confirm the nucleotide substitution or deletion directly and to prevent the mistyping by other methods.

[Application of stochastical analysis to maternity testing without putative mother].

In a maternity test in which the putative mother was deceased, the cumulative probability of maternity (PM) was calculated at 0.822 from 24 genetic markers by the stochastical method. This PM may not be evaluated in the same way as that of usual paternity cases. We applied the same method to two families whose blood relationships were undoubted. We compared the PMs in the cases in which maternal genotypes were estimated and were defined. Also, we calculated the PMs in the case of real maternal relationship and false maternal one. The estimated PM from real maternity relationship was significantly higher than that from false maternal one.

Laceration of the liver with delayed massive intra-abdominal hemorrhage: a case report of child abuse.

A 3-year-old girl was found unresponsive in the bedroom and expired at a hospital. Autopsy revealed massive intra-abdominal bleeding due to laceration of the liver and mesenterium with multiple rib fractures and multiple fresh and old bruises. The time of the assault causing the liver trauma was questioned because the perpetrator, her mother's boyfriend, denied any outrages on that particular day although he confessed that he had physically abused her for several months. Microscopically, numerous polymorph leucocytes infiltrated exclusively surrounding the lacerated area of the liver. Many hepatocytes were necrotic and cord arrangement of the parenchymal cells was destroyed. There should be a certain time lag between the major assault and massive intra-abdominal hemorrhage, which was not inconsistent with the statement of the perpetrator.

Polymorphism of short tandem repeat (STR) loci PLA2, D3S2459, D8S315 (kw38), CYP19, D3S1359, FGA, and D8S1132 in the Japanese and Chinese populations.

We report a study of polymorphism for seven short tandem repeat (STR) loci in Japanese and Chinese populations. Among 104 to 134 individuals in the both population samples, eight alleles were revealed for locus PLA2, thirteen for D3S1359, eleven for FGA, eight for D8S315 (kw38), ten for D8S1132, five for CYP19, and seven for D3S2459. They correspondingly constituted 10 to 39 genotypes therein. For most of the STRs, there was only a single allele active as the most frequent one among the others, except locus D3S1359 in Chinese samples (two alleles, 206 bp and 210 bp, frequency = 0.273 each). Also, the population genotype configurations were locus specific, varying in the patterns of commonest genotypes on each locus, e.g., one pattern for loci CYP19, D3S1359, and D8S315, one and two for loci PLA2 and D3S2459, two for locus D8S1132, and one and four for locus FGA. The distributions of observed genotypes were in Hardy-Weinberg Equilibrium. Furthermore, the seven STRs were exhibited highly polymorphic and informative for the both populations, and the alleles could be easily separated in electrophoresis and correctly interpreted with side-to-side allelic ladders. Together, the results suggest that the tri- and tetra-meric STRs are useful genetic markers for forensic practice.

IGF2 but not H19 shows loss of imprinting in human glioma.

Genomic imprinting is a gamete-specific modification resulting in the allele-specific expression of genes in somatic cells. A loss of imprinting (LOI) has been found in many embryonal and adult tumors, suggesting that it plays a role in tumor development. The incidence of LOI, however, does not seem to be ubiquitous among tumors because neuroblastoma and colorectal cancer revealed no LOI. We examined the involvement of LOI of IGF2 and H19 genes in human gliomas. The two genes were imprinted in normal brain subcortex tissues. In glioma, 8 of 14 informative cases (57%) revealed LOI in IGF2. The frequency did not depend on the tumor grade. For H19, in contrast, all 13 informative cases maintained imprinting. These results suggest that LOI of IGF2 but not H19 plays a role in the development of human glioma.

Alterations of c-fos gene methylation in the processes of aging and tumorigenesis in human liver.

The state of DNA methylation in the c-fos gene was examined in human livers of different ages, cirrhosis and hepatocellular carcinoma. The degree of methylation in the intron 1 to exon 4 region increased with age, whereas all of the 10 cirrhosis samples revealed a decrease in methylation when compared to normal livers of similar ages. The 11 hepatocellular carcinomas showed varied alterations suggesting that the alteration of the c-fos gene methylation is related to aging as well as to early-step of hepatocarcinogenesis.

Alteration of c-fos gene methylation in human gliomas.

In an attempt to find a common DNA alteration occurring in human glioma, we examined DNA methylation in 34 gliomas of various pathological grades and compared them with those in normal cerebral subcortex DNA. The total methylated cytosine levels in the genome did not differ appreciably between the tumors and the normal tissues; however, the degree of DNA methylation in several proto-oncogenes and suppressor oncogenes showed some alterations. Among them, the c-fos gene demonstrated deviation from that of normal tissues in all cases examined, suggesting that the alteration of c-fos gene methylation plays a role in the early steps of human glioma development.

Sudden unexpected death in childhood due to eosinophilic myocarditis.

A 12-year-old boy with no previous serious medical history experienced abdominal discomfort and chest pains for 5 days and suddenly died. The autopsy revealed diffuse and extensive infiltration of eosinophils into the myocardium, with poorly formed granulomas and few fibrotic changes. The necrotic changes was so extensive that Charcot-Leyden crystals formed. The other visceral organs had no specific pathologic changes except for mild lymphocytic infiltration with an increase in goblet cells in the bronchial areas and eosinocytosis in the blood vessels. An initial viral infection seemed to have caused subsequent eosinophil activation due to an allergic condition. Eosinophilic myocarditis is a rare cause of sudden death in apparently healthy children. Cardiac toxicity of eosinophils is, however, well established and dominates the ultimate prognosis of patients with complicated eosinophilia.

Monoclonal antibody against an amniotic protein carrying ABH blood group epitopes and its forensic application.

A mouse monoclonal antibody against an amniotic protein carrying ABH antigenic epitopes was established. BALB/c mice were immunized by an amniotic protein of molecular weight over 200 kDa, which had proved to be the carrier protein of ABH blood group epitopes by analysis with SDS-PAGE and immunoblotting. The antibody, ASP-1, was directed to the amniotic carrier protein without affecting the ABH blood group antigenicity, and did not cross-react with other body fluids which included blood, saliva, semen, urine or vaginal secretion. The immunoglobulin class of ASP-1 was IgG1 with a titer of 1 : 1,600. ASP-1 was used to detect the ABH blood group of amniotic fluid by the sandwich ELISA in which wells of plates were coated with ASP-1, and the ABH blood group of the captured protein was detected with mouse IgM anti-A and -B antibodies and enzyme conjugated anti-mouse IgM. The sandwich ELISA could successfully detect the blood group of amniotic fluid in mixed body fluids.

Molecular biologic analyses of tetragametic chimerism in a true hermaphrodite with 46,XX/46,XY.

OBJECTIVE: To investigate the mechanism of the formation of a tetragametic chimera with true hermaphroditism (46,XX/46,XY). DESIGN: Molecular biologic analyses. SETTING: Outpatient clinic and laboratories of a university hospital. SUBJECTS: A true hermaphrodite with 46,XX/46,XY and the parents. MAIN OUTCOME ANALYSIS: Restriction fragment length polymorphism (RFLP) of the pseudoautosomal region on sex chromosomes. RESULTS: Whereas a normal diploid individual showed two bands, the true hermaphrodite showed four bands in the RFLP analyses. Evaluation of the molecular weights of the bands revealed two of them to be of maternal origin and the other two to be of paternal origin. CONCLUSION: The two cell lineages composing the true hermaphrodite are heterogeneous because those originated from the fertilization of two genetically different maternal haploid cells by two different spermatozoa.