Comparison of Apoptosis and Bcl-2 Expression in Normal Pregnancy, Pregnancy Induced Hypertension and Intrauterine Growth Restriction / 대한산부인과학회잡지

OBJECTIVE: The purpose of this study was to investigate the incidence of apoptosis and expression of bcl-2 in the placenta of normal pregnancy, Pregnancy Induced Hypertension, and Intrauterine Growth Restriction. METHODS: Placenta samples were collected from 15 cases of normal full-term pregnancies, 15 cases of second trimester pregnancies, 17 cases of Pregnancy Induced Hypertension, and 13 cases of Intrauterine Growth Restriction. Hematoxylin and eosin staining and TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate marker nick end-labeling) staining were used to quantify the incidence of apoptosis and the electron microscopy was used to confirm it. Expression of bcl-2 was confirmed by using immunohistochemical stain in relation to apoptosis. RESULTS: 1. In TUNEL staining, quantification of apoptosis was 1.05 in 2nd trimester (n=15), 3.65 in pregnancy induced hypertensive pregnancy (n=17), 2.92 in intrauterine growth restrictive pregnancy (n=13) and 1.93 in normal full-term pregnancy (n=15). The incidence of apoptosis was significantly higher in placental tissues from full-term pregnancies than second trimester pregnancies (p0.05, t test), and between intrauterine growth restriction and normal full-term pregnancy (p>0.05, Mann-Whitney U test). CONCLUSION: These data suggest that apoptosis increases with gestational age, and in pathophysiologic states such as pregnancy induced hypertension and intrauterine growth restriction, and that bcl-2 expression is lower with gestational age.

Comparison of Apoptosis and Bcl-2 Expression in Normal Pregnancy, Pregnancy Induced Hypertension and Intrauterine Growth Restriction / 대한산부인과학회잡지

OBJECTIVE: The purpose of this study was to investigate the incidence of apoptosis and expression of bcl-2 in the placenta of normal pregnancy, Pregnancy Induced Hypertension, and Intrauterine Growth Restriction. METHODS: Placenta samples were collected from 15 cases of normal full-term pregnancies, 15 cases of second trimester pregnancies, 17 cases of Pregnancy Induced Hypertension, and 13 cases of Intrauterine Growth Restriction. Hematoxylin and eosin staining and TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate marker nick end-labeling) staining were used to quantify the incidence of apoptosis and the electron microscopy was used to confirm it. Expression of bcl-2 was confirmed by using immunohistochemical stain in relation to apoptosis. RESULTS: 1. In TUNEL staining, quantification of apoptosis was 1.05 in 2nd trimester (n=15), 3.65 in pregnancy induced hypertensive pregnancy (n=17), 2.92 in intrauterine growth restrictive pregnancy (n=13) and 1.93 in normal full-term pregnancy (n=15). The incidence of apoptosis was significantly higher in placental tissues from full-term pregnancies than second trimester pregnancies (p0.05, t test), and between intrauterine growth restriction and normal full-term pregnancy (p>0.05, Mann-Whitney U test). CONCLUSION: These data suggest that apoptosis increases with gestational age, and in pathophysiologic states such as pregnancy induced hypertension and intrauterine growth restriction, and that bcl-2 expression is lower with gestational age.

The Effect of RGP Lens and Reverse Geometry Lens on Apoptosis in Rabbit Cornea

PURPOSE: To investigate the cytotoxic effect of rigid gas permeable(RGP) and reverse geometry lens(RGL) on rabbit corneal cells. METHODS: In 36 white rabbits, right eyes were worn with either RGP or RGL. Left eyes were served as control. The rabbits were sacrificed at 1, 3, 7, 14 days after fitting and 7, 14 days after removal. Morphological changes of the basal epithelial cells and stromal keratocytes were observed using the light and transmission electron microscopy. The samples were examined after TUNEL staining in 3 high power fields (x400). RESULTS: In control eyes, 2.26% of central corneal epithelial cells showed apoptosis. After RGP wearing, the epithelial apoptosis decreased by 0.32% compared to control eyes(2.17%) at 14 days(p<0.001), and returned to 1.55% at 14 days after lens removal(2.04% in control, p=0.161). After RGL wearing, the degree of apoptosis was also lower than control(3.22%) by 0.30% at 14 days(p=0.003). However, it increased to 0.75% at 14 days after lens removal(1.67% in control, p=0.077). A few TUNEL positive fibroblasts were found in superficial stroma in both RGP and RGL group, but not in control eyes. CONCLUSIONS: Both RGP and RGL wearing may decrease the degree of apoptosis in corneal epithelium and lead the apoptosis in corneal stroma. The degree of apoptosis was recovered after lens removal, suggesting the process might to be reversible. The fitting with RGL induced apoptosis of keratocyte more than the fitting with RGP did. It suggests that the RGL wearing may be less physiologic than RGP wearing.

Effects of MgSO4 on Cerebral Infarction after Permanent Middle Cerebral Artery Occlusion in Rat

OBJECTIVE: This study is aimed to prove the neuroprotective effect of magnesium ion, which is known to have competitive action against calcium influx in the process of neuronal damage. METHODS: One hundred twenty five male Sprague-Dawley rats weighing 300-350gm were divided into control and MgSO4 injected groups. Middle cerebral artery occlusion was induced by nylon insertion near internal carotid artery bifurcation via external carotid artery. Forty rats which were alive for 48 hours after the induction of cerebral infarction were drawn from each group. Thirty minutes after the induction of cerebral infarction, 750mumol/Kg of MgSO4 was intramuscullary injected for the experimental group and same amount of normal saline was injected for the control group. Each group was subdivided into groups of 20 rats 48 hours after the cerebral infarction. Brain-sections of a subgroup were obtained after transcardially perfusion of 2% 2', 3', 5'-triphenyl-tetrazolium chloride and infarction volume was measured using imaging analysis program. Brain-sections of the other subgroup were stained with terminal deoxynucleotidyl-transferase-mediated biotin dUTP nick end labeling(TUNEL) and apoptotic index(AI) was estimated. RESULTS: The volume of cerebral infarction of the control and MgSO4 injected groups were 52.44+/-5.68% and 38.38+/-9.02% respectively, and the infarction volume was significantly reduced in MgSO4 injected group(p<0.001). Brain edema volume were 209.01+/-75.41mm3 in control group and 140.7+/-81.89mm3 in MgSO4 injected group that was a significant reduction in MgSO4 injected group(p<0.001). AI of 53.68+/-6.38 for control group and 29.80+/-6.77 for MgSO4 injected group were estimated and MgSO4 injected groups showed a significant reduction(p<0.001). CONCLUSION: Magnesium ion has neuroprotective effect in reduction of cerebral infarction volume, brain edema volume and AI. In the future, various attempts to prove protecting mechanism of magnesium ion should be made and TUNEL stain is thought to be an effective comparison method in such studies.

Correlation between Activity of Nitric Oxide Synthases and Apoptotic Change in the Closed Head Injury Animal Model

OBJECTIVE: The authors present an investigation of alterations in nitric oxide synthase(NOS) activity and histopathological response after moderate diffuse axonal injury(mDAI). METHODS: A total of 40 anesthetized Sprague-Dawley adult rats were injured utilizing the Marmarou's weight-drop device through a Plexiglas guide tube. These rats were divided into 8 groups(control, 1 hr, 2 hr, 3 hr, 6 hr, 12 hr, 24 hr, 48 hr after trauma). The temporal pattern of apoptosis after mDAI in the adult rat brain was characterized using TUNEL stain and cDNA for NOS activity was amplified using RT-PCR. The PCR products were electrophoresed on a 2% agarose gel and then observed under a UV-transilluminator. The enhanced chemiluminescence method(Pierce, Rockford, IL) was used to visualize the protein bands, and the quantification was performed by using the image analysis soft-ware Bio-1D. RESULTS: eNOS activity was not detected at all groups, but nNOS activity was expressed at 3 hr and continuously by 48 hr after impact , which was especially showed about 2 times larger than control group at 12 and 24 hr(mean value=28314+/-35.07 in injury group, 13386+/-26.14 in control group, p<0.05), followed by being on the decrease at 48 hr. iNOS activity was showed fivetmes larger than control group at 12 hr and 24 hr(mean value=15264+/-38.37 in injury group, 3002+/-31.62 in control group, p<0.05) followed by dramatically decrease below the level of control group(2299+/-14.06). Histologically, significant apoptotic changes after brain injury were occurred from 12 to 24 hr post-injury(AI=24.8%, 27.5%, p<0.001). CONCLUSION: These data indicate that nNOS and iNOS activity are affected after mDAI in a time-dependent manner and there is a close relation between apoptotic changes and NOS activity. nNOS activity is not stronger than iNOS, which lately expressed. However, iNOS activity is markedly reduced by 48 hr compared to nNOS.

Placental apoptosis in preeclampsia / 대한산부인과학회잡지

OBJECTIVE: Our purpose was to investigate the incidence of placental apoptosis in pregnancies complicated by preeclampsia. METHODS: Placental samples were obtained from 15 uncomplicated third-trimester pregnancies and from 17 pregnancies complicated by preeclampsia. TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling) staining and electron microscopy were performed on all placental samples and identified apoptosis within the cells of the placenta. TUNEL positive stained cells were counted at each photograph(five sections were photoragphed at 1,500 magnifications for each sample). We focused on only the syncytiotrophoblast nuclei of the placenta. The number of apoptotic syncytiotrophoblast nuclei identified was expressed as a percentage of total number of syncytiotrophoblast nuclei counted. RESULTS: Quantification of apoptosis (mean+/-SD) was as follows: normal third trimester (n=15) 0.57+/- 0.47% of cells and preeclampsia third trimester (n=17) 1.41+/-0.67% of cells. The incidence of apoptosis was significantly increased in placentas from pregnancies with preeclampsia compared with normal placentas (p< 0.01). CONCLUSION: These results suggest that placental apoptosis may play a role in the pathophysiologic mechanisms of preeclampsia.

Light and Electron Microscopic Study on Epithelial Cell Death During Implantation in Rat Uterus / 대한해부학회지

The loss of the uterine epithelium surrounding blastocyst is important in bringing the trophoblast into close association with the endometrial stroma during embryonal implantation in several laboratory rodents. It is usually assumed that the epithelial cells die during this process, but the cause of epithelial cell death remains speculative. This study was performed to investigate the mechanism of epithelial cell death during implantation in the endometrium of rat uterus throughout days 5-9 of pregnancy by light and electron microscopy. Cathepsin-D reactivity, detected by imunohistochemistry, was localized to all viable epithelia, some degenerating epithelial and decidual cells between days 5-9 of pregnancy. DNA fragmentation, detected by TUNEL staining, was observed in degenerating epithelial and decidual cells throughout days 6-9 of pregnancy. Cellular fragments within lumen and phagosome within trophoblasts were reacted positively to TUNEL staining. Electron microscopy revealed that the epithelium lining the antimesometrial chamber was sloughed off into the lumen on day 6 of pregnancy. Its cell organelles appeared healthy but its nucleus was condensed and irregular shaped. The detached epithelial cells lining the antimesometrial chamber on day 7 of pregnancy contained condensed and fragmented nuclei, but exhibited different morphological pattern according to the intactness or loss of cytoplamic organelles and membrane. And these cells were surrounded and phagocytosed by trophoblasts. On day 7 of pregnancy, syncytial knots were formed, and later these knots and most of detached epithelial cells were severely degenerated. Some decidual cells nearby the lumen were also degenerated at this period. These results indicate that the epithelial cell death of rat uterine endometrium during implantation is mainly due to the process of apoptosis and these dying cells are removed by trophoblasts.

An Electron Microscopic Study on the Effect of Hypokalemia in Endometrial Epithelium of Rat Uterus / 대한해부학회지

To investigate morphologic alterations in the rat uterine epithelium after chronic potassium depletion, light and electron microscopic studies were made in rats fed a potassium-free diet for 2 weeks. The uterine epithelium was simple columnar and the uterine gland was simple cuboid cells in toluidine blue staining. In experimental uterus, the epithelial cells contain many brownish granules in the supranuclear and basal cytoplasm. The cells containing dense body with pale halo appeared occasionally among the epithelial cells. By TUNEL staining, more positive cells were observed in experimental uterus than those in control group. By electron microscopy, uterine epithelial cells of control rat had short, irregular microvilli and a few apical vesicles and cytoplasm. Increased microvilli, apical vesicles and fat droplets were observed in uterine epithelial cells of the experimental rat. Apoptotic cells or cells containing apoptotic bodies were sometimes present among the epithelial cells. These results indicate that morphological alterations in the rat uterine epithelium are occurred at chronic potassium depletion, and suggest that hypokalemia may induce apoptosis of uterine epithelium.

The Mechanism of Tissue Injury in the Animal Model of Experimental Retinal Vein Occlusion

We investigated the development of programmed cell death of apoptosis as a mechanism of cell death in experimental retinal vein occlusion of rats. We induced hemicentral retinal veinocclusion by photocoagulation 3 or 4 out of 6 retinal veins after infecting rose bengal intravenously.The eyes were enucleated sequentially between day one to day fourteen, and the retinas were examined after H&E and TUNEL staining. Exudative retinal detachment developed one day after vein occlusion and was reabsorbed completely by day 4. The number of ganglion cells in vein-occluded retina began to decrease one day after vein occlusion and continue to decrease until day fourteen. Many cells in ganglion cell layer and inner nuclear layer were stained positively by TUNEL staining. These results suggest that apoptosis is an important mechanism of cell death in early stage of experimental retinal vein occlusion.