Hyun Ju Choi

Multinucleation of koilocytes is in fact multilobation and is related to aberration of the G2 checkpoint.

Aims: To clarify the fine structure of koilocytes and correlate this with genetic aberration of the G2 checkpoint Methods: Three dimensional reconstruction from confocal fluorescent images, together with functional assays for key molecules of the G2 checkpoint—cdc2 and cyclin B1—was performed in human uterine cervical samples. After confirming 22 human papillomavirus (HPV) types using a DNA chip from 30 cervical swabs, previously confirmed as 15 cervical low grade and 15 high grade intraepithelial lesions, the activity of molecules involved in the G2 checkpoint was evaluated using western blotting for cyclin B1, cdc2, and phospho-cdc2 (Y15 and T161), a nuclear extraction fractional assay, and a reverse transcription polymerase chain reaction assay. In addition, three dimensional confocal image restoration was performed on confirmed cervical intraepithelial neoplasia tissue samples. Results: T161 phospho-cdc2 and cyclin B1 expression was higher in HPV infected cervical lesions than in normal samples. Immunofluorescence, revealed that cyclin B1 was present predominantly in the nuclei of HPV infected cells, confirming the results of the nuclear fractional assay. On restoration of three dimensional confocal images, the multinucleation of koilocytes was revealed to be multilobation of a single nucleus, rather than true multinucleation. This multilobation appeared to be associated with chromosomal instability and aberration of the G2 checkpoint. Conclusions: The multiple nuclei of koilocytes are in fact multilobation of a single nucleus, and this phenomenon is associated with upregulation of gene products related to the G2 checkpoint.

The Polysaccharide Fraction AIP1 from Artemisia iwayomogi Suppresses Apoptotic Death of the Mouse Spleen Cells in Culture

A polysaccharide fraction, AIP1, purified from Artemisia iwayomogi was shown to have immu-nomodulating and anti-tumor activities in mice. In order to determine how the AIP1 fraction exhibits the immunomodulating activity, the effect of the fraction on the apoptosis of mouse spleen cells was investigated. Treatment of the mouse spleen cells with the AIP1 fraction resulted in the suppression of apoptotic death and an extension of cell survival in culture, indicating that the fraction might modulate the death of spleen cells. Treatment of the mice with the AIP1 fractionin vivo also resulted in less apoptosis of the spleen cells, which indicates the physiological relevance of the anti-apoptosis effect of the fractionin vitro. A mouse gene array was used to determine the profile of the gene expression change showing a pattern of up-and down-regulated genes by the AIP1 treatment. This study provides preliminary information regarding the immunomodulatory mechanism of the AIP1 fraction.

Quantitative analysis of chemotherapeutic effects in tumors using in vivo staining and correlative histology.

Aims: To microscopically analyze the chemotherapeutic response of tumors using in vivo staining based on an annexinV-Cy5.5 probe and independently asses their apoptotic count using quantitative histological analysis. Methods: Lewis Lung Carcinomas cells, that are sensitive (CS-LLC) and resistant (CR-LLC) to chemotherapy were implanted in nude mice and grown to tumours. Mice were treated with cyclophosphamide and injected with a Cy5.5-annexinV fluorescent probe. In vivo imaging was performed using Fluorescence Molecular Tomography. Subsequently tumours were excised and prepared for histology. The histological tumour sections were stained for apoptosis using a terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assay. A minimum of ten tissue sections were analyzed per tumour for apoptosis quantification by TUNEL staining and corresponding Cy5.5 distribution. Results: We detected higher levels of apoptosis and corresponding higher levels of Cy5.5 fluorescence in the CS-LLC vs. the CR-LLC tumours. The cell count rate on CS-LLC sections over CR-LLC was found to be ∼2u2009:1 where the corresponding area observed on Cy5.5 distribution measurements revealed a ∼1.7u2009:1 ratio of CS-LLC over CR-LLC. These observations are consistent with the higher apoptotic index expected from the CS-LLC cell line. Conclusions: Quantitative analysis of histological slices revealed higher fluorescence and higher apoptotic count in the CS-LLC tumour images compared to the CR-LLC tumour images. These observations demonstrate that the annexinV-Cy5.5 probe sensed the chemotherapeutic effect of cyclophospamide and further confirmed in vivo FMT measurements.

An oligosaccharide fraction from Korean mugwort herb suppresses death of the mouse thymocytes in culture by down-regulating the Fas death receptor gene

Korean mugwort herb is a preparation of dried leaves from Artemisia species and has been used as a traditional medicine in Asia. An oligosaccharide fraction, AVF3, purified from the preparation promoted survival of the mouse thymocytes in culture. A mouse gene array study suggests that the AVF3 may modulate Fas/FasL dependent apoptotic cell death and thus has influence on the survival of the thymocytes in culture. RT-PCR analysis confirmed the down-regulation of the Fas gene by the AVF3 treatment, supporting that the AVF3 modulated thymocyte death by suppressing the Fas gene expression.Korean mugwort herb is a preparation of dried leaves from Artemisia species and has been used as a traditional medicine in Asia. An oligosaccharide fraction, AVF3, purified from the preparation promoted survival of the mouse thymocytes in culture. A mouse gene array study suggests that the AVF3 may modulate Fas/FasL dependent apoptotic cell death and thus has influence on the survival of the thymocytes in culture. RT-PCR analysis confirmed the down-regulation of the Fas gene by the AVF3 treatment, supporting that the AVF3 modulated thymocyte death by suppressing the Fas gene expression.

Development of quantification software using model-based segmentation of left ventricular myocardium in gated myocardial SPECT

Gated myocardial single photon emission computed tomography (SPECT) is being used for the diagnosis of coronary artery diseases. In this study, we developed new software for the quantification of volumes and ejection fraction (EF) on the gated myocardial SPECT data using a cylindrical model. Volumes and EF by developed software were validated by comparing with those quantified by quantitative gated SPECT (QGS) software. Cylinder model for left ventricular myocardium was used to eliminate background activity and count profiles across the myocardium were fitted to the Gaussian curve to determine the endocardial and epicardial boundary. End-diastolic volume (EDV), end-systolic volume (ESV) and EF were calculated using this boundary information. Gated myocardial SPECT was performed in 83 patients. EDV, ESV and EF values estimated using present method were compared to those obtained using the commercialized software QGS, and reproducibility in the parameter estimation was assessed. EF, EDV and ESV obtained using two methods were correlated well (correlation coefficients = 0.96, 0.96 and 0.98). The correlation between the parameters repetitively estimated from the same data set by an operator was very high (correlation coefficients = 0.96, 0.99 and 0.99 for EF, EDV and ESV). On the repeated acquisition, reproducibility was also high with correlation coefficients of 0.89, 0.97 and 0.98. The present software will be useful for the development of new parameters for describing the perfusion and function of the LV.

A case of pemphigus vulgaris associated with ulcerative colitis

Pemphigus vulgaris is an autoimmune bullous disorder characterized by the production of autoantibodies against the intercellular space of the epithelium. It has rarely been reported in association with inflammatory bowel disease. Ulcerative colitis is one of the forms of inflammatory bowel disease. A 62-year-old woman who had been treated for ulcerative colitis for 16 years developed pruritic bullae on the skin of her face and body. Histological findings and direct immunofluorescence examination of the skin showed pemphigus vulgaris. She was treated with systemic steroids, mesalazine, and azathioprine. Her cutaneous lesions have remained in remission and her ulcerative colitis has remained well-controlled. The relationship between pemphigus vulgaris and ulcerative colitis is unclear. An autoimmune response has been suspected in the pathogenesis of ulcerative colitis. Pemphigus vulgaris is also associated with an autoimmune mechanism. To our knowledge, this is the first case of ulcerative colitis associated with pemphigus vulgaris reported in Korea. The association may be causal.

Change in the electroencephalogram delta wave in the frontal cranial region of rats with the hyperventilation.

Hyperventilation is one way to cause activation on the electroencephalogram (EEG) to diagnose brain disorders. The hyperventilation is also known to affect on the delta power in EEG. This study divided the total delta wave into low, middle, and high bands corresponding to the wave frequency. The power in these three delta wave bands was examined in the frontal cranial region of adult male Sprague-Dawley rats hyperventilated with ventilation (VE) of 360, 540, and 720 ml/min for 5 min. The control group was ventilated normally with a volume of 160 ml/min. The results show that the relative power of the low delta band in the rats hyperventilated at 360 ml/min VE was significantly increased compared with powers of pre-hyperventilation (p<0.05). The relative power of the middle delta band was not significantly affected by hyperventilation at any VE, and in the high delta band, all of the relative powers were decreased significantly in all hyperventilated rats compared with powers of pre-hyperventilation (p<0.05). We concluded that hyperventilation affects the frontal cranial region, by increasing the low delta band and decreasing the high delta band.

A computerized program for three-dimensional visualization and quantitative analysis of cell nuclei

In this paper, we have proposed an image processing system for the acquisition and processing of three-dimensional images based on confocal scanning laser microscopy for the purpose of three-dimensional visualization and quantitative analysis of cell nuclei. The three-dimensional visualization methods can be divided into surface rendering and volume rendering. The way that surface rendering is used within this system is based on contour modeling. This method consists of several steps as follows. The first step is to preprocess the volume data obtained. Secondly, the extraction of the contours of each slice is carried out. Thirdly, smoothing algorithms are used to refine the contour data and remove wiggles. Since the surface rendering accounts only for the surface, the inside is not visible. Therefore, based on the basic volume rendering pipeline, we implemented the volume rendering. In the quantification step, in order to extract quantitative features, we made a three-dimensional labeling method based on slice information. Compared to the conventional algorithms, this method has advantages due to the use of memory is highly efficient and it is possible to combine a variety of two-dimensional labeling algorithms to find an appropriate labeling to its application. After applying the labeling algorithm, we extracted the measurements for the three-dimensional quantitative analysis of cell nuclei: nuclear volume, surface area and spherical shape factor. This could become a way to improve the accuracy and reproducibility of quantifying cell nuclei. We believe that our method will become a useful diagnostic tool for the medical image analysis.