Effects of pre-surgical nasoalveolar moulding on maxillary arch and nasal form in unilateral cleft lip and palate before lip surgery.

OBJECTIVES: To investigate the effects of pre-surgical nasoalveolar moulding (PNAM) on the maxillary arch and nasal form in patients with unilateral cleft lip and palate (UCLP). SETTING AND SAMPLE POPULATION: This is a retrospective case series study. The subjects were infants with complete UCLP who were treated with PNAM (n = 18) at Kagoshima University Medical and Dental Hospital (Japan) between 2006 and 2013. MATERIAL AND METHODS: Maxillary dental casts and facial photographs were taken at the time of the first visit and immediately prior to lip surgery to evaluate the maxillary arch and nasal form changes. The dental casts were scanned with a laser scanner, and changes in the 3-Dimensional coordinates of anatomical landmarks and alveolar cleft width were analysed. Moreover, we investigated the correlation between the changes in the maxillary alveolar arch and nasal form. RESULTS: Regarding the maxillary alveolar arch form, the anterior points of the major segment had moved significantly to the cleft side just prior to the time of lip repair, and the alveolar cleft width was significantly decreased. For nasal form, the inclination and displacement of the columella were significantly improved. The improvement of columella inclination was moderately correlated with the posterior movement of the anterior points of the major segment. CONCLUSIONS: These findings indicate that PNAM for infants with UCLP enhanced symmetry in the maxillary alveolar arch and nasolabial form. In addition, the posterior movement of the anterior points of the maxillary alveolar arch was correlated with the improvement of columella deformation.

Structure-activity relationships of triterpenoid derivatives extracted from Gymnema inodorum leaves on glucose absorption.

The leaves of Gymnema inodorum (GI) have been known to be effective for some diseases including diabetes mellitus, rheumatic arthritis and gout. The crude saponin mixtures extracted from GI leaves inhibited glucose absorption in the isolated intestinal tract and suppressed the increased blood glucose in rats. In this study, we examined the relationship between chemical structure and pharmacological activity of the four components from GI leave extracts (GiA-1, GiA-2, GiA-5 and GiA-7). These components were the derivatives of (3beta,4alpha,16beta)-16,23,28-trihydroxyolean-12-en-3-yl-beta-D-glucopyranosiduroic acid. GiA-2, GiA-5 and GiA-7 that have suppressive effects on the high K+-induced contraction, an increase in deltaPD and the increased blood glucose level in the glucose tolerance test have -H at the 21st position and -CH2OH at 4beta of aglycon. On the other hand, GiA-1 that does not have any effects on the three parameters mentioned above has -H at the 21st position and -CH3 at 4beta of aglycon. In conclusion, it is suggested that the inhibitory effect of triterpenoids in Gymnema leaves on glucose absorption from the intestinal tract relies on -CH2OH at 4beta.

Mice containing a human chromosome 21 model behavioral impairment and cardiac anomalies of Down's syndrome.

Trisomy 21 (Ts21) is the most common live-born human aneuploidy; it results in a constellation of features known as Down's syndrome (DS). Ts21 is the most frequent cause of congenital heart defects and the leading genetic cause of mental retardation. To investigate the gene dosage effects of an extra copy of human chromosome 21 (Chr 21) on various phenotypes, we used microcell-mediated chromosome transfer to create embryonic stem (ES) cells containing Chr 21. ES cell lines retaining Chr 21 as an independent chromosome were used to produce chimeric mice with a substantial contribution from Chr 21-containing cells. Fluorescence in situ hybridization and PCR-based DNA analysis revealed that Chr 21 was substationally intact but had sustained a small deletion. The freely segregating Chr 21 was lost during development in some tissues, resulting in a panel of chimeric mice with various mosaicism as regards retention of the Chr 21. These chimeric mice showed a high correlation between retention of Chr 21 in the brain and impairment in learning or emotional behavior by open-field, contextual fear conditioning and forced swim tests. Hypoplastic thymus and cardiac defects, i.e. double outlet right ventricle and riding aorta, were observed in a considerable number of chimeric mouse fetuses with a high contribution of Chr 21. These chimeric mice mimic a wide variety of phenotypic traits of DS, revealing the utility of mice containing Chr 21 as unique models for DS and for the identification of genes responsible for DS.

Development of a self-oscillating ultrasonic micro-motor and its application to a watch

We have developed an ultrasonic micro-motor for use as a micro-actuator in place of an electromagnetic motor. This ultrasonic micro-motor, which can be driven by a single signal and in which the change of the direction of the rotor movement can be made easily by selecting the electrode to apply the driving signal, can easily construct a self-oscillating circuit and simplify the driving circuit. We have also simplified the motor structure, which is easy to miniaturize and mass-produce. We applied a version of this motor with a diameter of 8 mm to a vibration alarm, and one with a diameter of 4.5 mm to a driving source of a calendar mechanism in a watch. This ultrasonic micro-motor is expected to be of use as a new driving source in a broad range of fields.

A new concept of the pattern of structural changes with bone loss by histomorphometric analysis using bone slabs.

We examined naked bone slabs (1.2 mm thick) from iliac bone biopsied cores obtained from 33 women aged 33-89 years. The number, size, and shape of the pores in the bone slabs were analyzed. The results revealed that the % bone area (the percentage area occupied by bone in the slab) was linearly correlated with age and other parameters, such as the size of pores, irregularity of pores, and pore distance, but was not correlated to the number of pores. We found a second-degree polynomial relationship between the % bone area and the number of pores. Based on three parameters--% bone area, number of pores, and size of pores--cluster analysis was performed and the specimens divided into three groups. The group with sufficient bone mass showed few small round pores, and the group with severe bone loss revealed a few large pores that were caved in. The characteristics of these groups represented the relationship between bone mass and structural change. The remaining group with moderate bone loss was divided into two subgroups, one with an increased number of pores without expansion and one with expanded pores without an increase in number. We presumed that the variations between the groups were caused by differences between fine and rough structures in the trabeculae caused during the process of bone loss. We concluded that this analysis of bone slabs allowed the pattern of trabecular structural change that occurred with bone loss to be determined easily and visually.

Fine structure of hydrous chromosomes observed by low vacuum scanning electron microscopy.

Chinese hamster metaphase chromosomes were stained with platinum (Pt) blue and observed in the hydrous state with low vacuum scanning electron microscopes (LVSEM). The coiled structure of the chromatin fibers in the chromosomes was well recognized through the surrounding perichromosomal substances in the backscattered electron (BSE) mode at accelerating voltages of over 20 kV. Findings indicated that chromatin fibers in native chromosomes have a structure similar to the hierarchic coiled model. The present study also demonstrated that not only surface structures but also subsurface structures can be studied in the BSE mode of LVSEM, when the subsurface structures have been stained with heavy metal salts such as Pt blue.

Scanning electron microscopic and cytochemical analyses of alveolar and peritoneal macrophages of mice, dogs and cats.

Alveolar and peritoneal macrophages (MPs) of mouse, dog and cat were compared in relation to their scanning electron microscopic features and the lysosomal activities of nonspecific esterase, acid phosphatase, and beta-glucuronidase. The long spindle shape of peritoneal MPs differed from the spherical form of alveolar MPs in all species. There was no difference in the morphological findings among the three animals. Murine alveolar and peritoneal MPs were strongly positive for all three enzymes. Canine and feline alveolar and peritoneal MPs were strongly positive for acid phosphatase and beta-glucuronidase, but weakly positive for nonspecific esterase. These results strongly suggest that acid phosphatase and beta-glucuronidase can be used as markers of the MPs in healthy dogs and cats.

[Study on the organization of chromatin and chromosome in mouse spermatogenic cells by scanning electron microscopy].

It remains unclear about the intermediate construction of chromosome due to its highly compact nature and the limitation in methods. The present study was designed to investigate the construction of chromatin and mitotic chromosome in situ with scanning electron microscopy. Mouse testes were selected as the material, because of in which the spermatogenic cells divide actively and successively to form the sperm. Such a feature would be able to study the structure of mammalian chromatin and chromosomes along with the change of nuclear cycle. The animal were perfused with 200 ml of 0.075 mol/L KCl hypotonic solution to remove blood and placed for 15-20 min on ice followed by 0.5% glutaraldehyde and 0.5% formaldehyde for fixing. Through treated by the routine process of fractured and freeze dried with t-butyl alcohol, the specimens were then coated with a 3 nm thick platinum and observed with Hitachi S-430 scanning electron microscopy. It was found that the hypotonic treatment with 0.075 mol/L KCl solution was suit for demonstrating the nuclear structure, when the organelles were well preserved. The chromatin fibers of 10-30 nm and 80-125 nm in diameter could be recognized in the interphase nuclei, which were arranged losely at the region of euchromatin, and folded with each other into chromatin masses at the region of heterochromatin, while the chromatin fibers with the diameter of 80-125 nm often could be viewed on the mitotic chromosomes. Since its presence in interphase nuclei and mitotic chromosomes, it was considered that the chromatin fibers with 80-125 nm in diameter might play a role in the condensation of chromosome, serve as a type of the intermediate structure.

Scanning and transmission electron-microscopic study of the development of the podocyte in the human fetus.

The metanephric kidneys of seven human fetuses at 11-17 weeks' gestation were examined by scanning and transmission electron microscopy in order to evaluate differentiation of glomerular podocytes. When glomeruli were in the stage of S-shaped bodies, the surface of the visceral epithelium of renal corpuscles was smooth, with indistinct cell borders. As the glomeruli developed, visceral epithelial cells of renal corpuscles became spherical and resembled clusters of grapes in dense aggregation. In this stage, processes and foot processes were simultaneously formed at the base of epithelial cells. As glomeruli further differentiated, visceral epithelial cells of renal corpuscles began to separate from one another and became flat with the development of vascular loops. Processes and foot processes were exposed for the first time in Bowman's space. In this stage, the degree of differentiation of epithelial cells varied widely among various sites of the glomerulus. As glomeruli developed further, projections became more complex and epithelial cells began to show structures similar to those of adult epithelial cells. The adjacent foot processes arose from different cells throughout the period of morphological differentiation.

Suppression of glucose absorption by some fractions extracted from Gymnema sylvestre leaves.

Extracts containing gymnemic acids, which were extracted from the leaves of Gymnema sylvestre (GS) as nine fractions, were evaluated for their effects on a high K(+)-induced contraction of guinea-pig ileal longitudinal muscles, on glucose transport mediated by the difference of glucose-evoked transmural potential difference (delta PD) in the inverted intestine of guinea-pig and rat, and on blood glucose in rat. Among nine fractions obtained by high performance liquid chromatography from the extract, f-2 and f-4 strongly suppressed the high K(+)-induced contraction of the ileal muscle, f-3 and f-5 did so moderately, and f-8 and f-9 did so weakly, whereas the other fractions did not affect it. The degree of suppression of high K(+)-induced contraction by f-2 at 74% was almost the same as that of f-4 at 67%, at concentrations of 0.1 mg/ml. The suppressed contraction by f-2 or f-4 was recovered by adding 5.5 mM pyruvate. The delta PD increased by 5.5 mM glucose in the inverted intestines of guinea-pig and rat were equally suppressed by 0.1 mg/ml of f-2 or f-4 to 40%. In a rat sucrose tolerance test, f-2 and f-4 suppressed the elevation of blood glucose level. Both f-2 and f-4 suppressed the contraction of guinea-pig ileal longitudinal muscle, interfered with the increase in delta PD induced by glucose in the inverted intestines of guinea-pig and rat, and inhibited the elevation of blood glucose level. In conclusion, it is suggested that some of the extracts containing gymnemic acids from GS leaves suppress the elevation of blood glucose level by inhibiting glucose uptake in the intestine.

Diagnostic revolution of microhematuria by real time confocal scanning laser microscope: Hyodo-Iino-Miyagawa method, third report.

The real time confocal scanning laser microscope provides excellent three-dimensional images free of out-of-focus information. The objective of this study was to evaluate the usefulness of the laser microscope for the diagnosis of microhematuria. Characteristics of the test were evaluated in 81 patients with definite causes of hematuria. 30 erythrocytes in urinary sediments were examined for each patient, and those in whom less than 20% of the erythrocytes were poikilocytes were considered to have urological diseases and those in whom 80% or more of the erythrocytes were poikilocytes to have nephritis. According to these criteria, the sensitivity and the specificity of the examination to nephritis were 100 and 98.1%. 91.4% of the patients with urological disease had the nonglomerular type. The time required for the examination was less than 2-5 min in samples containing 1-3 erythrocytes in one field under an ordinary light microscope (x400).

Immunocytochemical localization of actin in the nucleolus of rat oocytes.

In order to determine the localization of actin, growing and fully grown rat oocytes were immunocytochemically examined using a post-embedding ultrastructural protein-A gold technique. In quiescent oocytes, the nucleoplasm showed slightly lower levels of actin signal when compared to the surrounding cytoplasm. The highest levels of labeling were found on nucleoli showing a reticular type morphology. In oocytes at the diakinesis stage in which nucleolar compaction had occurred, the levels of labeling increased by 5-6 times those found in quiescent oocytes. Except for conspicuous accumulation of actin under the plasma membrane, compact nucleoli had significantly higher levels of labeling when compared with those found on the general cytoplasm, while the nucleoplasm with homogeneously dispersed chromatin showed significantly lower levels of associated actin signal than the general cytoplasm. In oocytes at metaphase I, the cytoplasmic region had comparable or lower levels of labeling than the cytoplasm of oocytes at diakinesis. The meiotic spindle embedded in material with medium electron density showed a similar level of labeling as the surrounding cytoplasm. On the other hand, significantly higher levels of associated actin were observed on the chromosomes of metaphase I. The actin signals were dispersed over the chromosomes and not concentrated on a specific region. These results suggest that nuclear actin may be involved in the process of chromosome construction and also the formation of the compacted structure of the nucleolus.

Confocal scanning laser microscopy of morphometric human sperm parameters: correlation with acrosin profiles and fertilizing capacity.

OBJECTIVES: To develop quantitative criteria for assessing sperm morphology and to determine the correlation between the percentage of morphologically normal spermatozoa and the outcome of the sperm hypo-osmotic swelling test, sperm acrosin profile, and sperm capacity for fertilization. DESIGN: The maximal length and width of the sperm head, the length of the midpiece and principal piece of the sperm tail, and the ratio of the surface of the acrosomal region to the total surface of the head were determined in specimens obtained from a group of infertile men and a group of fertile men using a confocal scanning laser microscope. Group A consisted of 53 infertile men who were participating in an IVF program, and group B consisted of 98 fertile men. The mean +/- 2 SD of the morphometric parameters in group B was established as representing the lowest and highest normal values in both groups. A normal spermatozoon was defined as one with morphometric parameters within normal levels. The lowest percentage of morphologically normal spermatozoa, hypo-osmotic swelling test result, and acrosin activity in group B were also taken as the lowest normal values in group A. SETTING: In vitro fertilization program at the Tottori University School of Medicine, Yonago, Japan. MAIN OUTCOME MEASURES: Sperm morphometric parameters, percentage of morphologically normal spermatozoa, hypo-osmotic swelling test, and acrosin activity. RESULTS: The length of the midpiece, ratio (x 100) of the surface of the acrosomal region to the total surface of the sperm head, percentage of morphologically normal spermatozoa, outcome of hypo-osmotic swelling test, and acrosin activity were significantly higher in group B than in group A. The maximal width of the head was significantly lower in group B than in group A. Strongly positive correlations were observed between percentage of morphologically normal spermatozoa or length of midpiece and the proportion of fertilized oocytes in group A and between ratio (x 100) of the surface of the acrosomal region to the total surface of the head and acrosin activity in groups A and B. Sperm morphology showed high positive and negative predictive values for acrosin activity (normal/abnormal) and fertility potential (present/absent). CONCLUSIONS: Using quantitative strict criteria, we found that sperm morphology was an important predictor of sperm fertilizing capacity. The confocal scanning laser microscope provided useful information about the sperm cytoskeleton and its importance in fertilization.

Structural changes of iliac cancellous bone in postmenopausal women: a scanning electron microscopic study.

The functional state of iliac cancellous bones in healthy postmenopausal women was investigated by scanning electron microscopy (SEM) using a modified organic preparation method. SEM analysis of the location, number, size and phase of the remodeling sites showed that normal iliac cancellous bone of postmenopausal women has high resorbing activity in the early stage of postmenopause in both the region near the cortico-endosteum and the midway region, while in the late stage of postmenopause there is a great difference in resorbing activity between the two regions; namely, the resorbing activity progressively decreases with age, but high resorbing activity is continuously present in the region near the cortico-endosteum. Our results suggest the possibility that at the region near the cortico-endosteum the bone volume loss occurs continuously throughout all the stages of post-menopause, while at the midway region it occurs only in the early stage and not in the late stage of postmenopause.

The spatial distribution of spindle microtubules in anaphase 3Y1 cells.

The spatial distribution of the microtubules (MT) in the rat 3Y1 cells in mitosis was investigated by immunoelectron microscopy and computer-graphic reconstruction of serial thin sections. In anaphase the interzone-MT increased in number gradually with advancing phase, while the kinetochore-MT in half-spindles decreased. The interzone-MT overlapped with each other at the equatorial region of the cell, and they formed a specific structure called the 'stem bodies'. The ends of the interzone-MT opposite to the stem bodies often attached to chromosomes but not to the poles. The stem bodies were not labeled with immunogold particles of anti-alpha tubulin. Some of the stem bodies or MT which originate from stem bodies were found just beneath the plasma membrane in the equatorial region where abundant actin filaments appear showing the formation of the contractile ring and subsequently the cleavage furrow begins. On the basis of these observations it is assumed that the interzone-MT is involved both in the separation of chromosomes in anaphase and in the formation of the cleavage furrow in telophase.

Acrosin profiles of human spermatozoa recovered from the new Sperm Prep II filtration column.

Twenty semen samples were collected and two aliquots of 1.5 ml were prepared from each sample. One sample of each pair (fraction-one) was used for evaluation of semen parameters and total acrosin activity of spermatozoa. The other sample of each pair (fraction-two) was mixed with an equal volume of Ham's F-10 medium and filtered through the Sperm Prep II. At the end of the filtration, the filtrate was pooled, sperm parameters were evaluated and total acrosin activity was assessed. Percentage of normal spermatozoa, sperm motility, and total acrosin activity of spermatozoa were significantly higher in the postfiltered fraction-two than in the fresh fraction-one. Considering the great importance of acrosin for fertilization, it is suggested that Sperm Prep II filtration method may be beneficial in preparing spermatozoa from infertile men with low acrosin profiles for assisted reproduction programs. Further studies will be necessary, however, in order to confirm this, since the present study concerned men with known fertility.

The tannin-osmium conductive staining after dehydration: an attempt to observe the chromosome structure by SEM without metal coating.

The tannin-osmium conductive staining method was modified in order to observe chromosomal structures without metal coating. The treatment with tannic acid and OsO4, which has been used in aqueous solution in the original method, was performed in acetone solution after dehydration in the present study. The usefulness of this method is discussed showing ultrastructural images of chromosomes of Drosophila and Tradescantia, which can be achieved by the method.

Application of colour laser microscope for observing living biological specimens.

Pollen-mother cells from Lilium longiflorum, human metaphase chromosomes and human spermatozoa, were observed under a colour laser microscope. Helical mitochondrial strands were clearly evident in the middle piece of human spermatozoa. The technique enables living specimens to be conveniently and effectively examined.