Spatiotemporal expression of RCAN1 and its isoform RCAN1-4 in the mouse hippocampus after pilocarpine-induced status epilepticus

Regulator of calcineurin 1 (RCAN1) can be induced by an intracellular calcium increase and oxidative stress, which are characteristic features of temporal lobe epilepsy. Thus, we investigated the spatiotemporal expression and cellular localization of RCAN1 protein and mRNA in the mouse hippocampus after pilocarpine-induced status epilepticus (SE). Male C57BL/6 mice were given pilocarpine hydrochloride (280 mg/kg, i.p.) and allowed to develop 2 h of SE. Then the animals were given diazepam (10 mg/kg, i.p.) to stop the seizures and sacrificed at 1, 3, 7, 14, or 28 day after SE. Cresyl violet staining showed that pilocarpine-induced SE resulted in cell death in the CA1 and CA3 subfields of the hippocampus from 3 day after SE. RCAN1 immunoreactivity showed that RCAN1 was mainly expressed in neurons in the shammanipulated hippocampi. At 1 day after SE, RCAN1 expression became detected in hippocampal neuropils. However, RCAN1 signals were markedly enhanced in cells with stellate morphology at 3 and 7 day after SE, which were confirmed to be reactive astrocytes, but not microglia by double immunofluorescence. In addition, real-time reverse transcriptase–polymerase chain reaction showed a significant upregulation of RCAN1 isoform 4 (RCAN1-4) mRNA in the SE-induced hippocampi. Finally, in situ hybridization with immunohistochemistry revealed astrocytic expression of RCAN1-4 after SE. These results demonstrate astrocytic upregulation of RCAN1 and RCAN1-4 in the mouse hippocampus in the acute and subacute phases of epileptogenesis, providing foundational information for the potential role of RCAN1 in reactive astrocytes during epileptogenesis.

Connected in cinema: educational effects of filmmaking classes on medical students / 한국의학교육

PURPOSE@#The purpose of this study is to explore what the students experienced in short filmmaking class performed to 1st grade premedical students of a medical school, and to trace educational effects of the class.@*METHODS@#Taking a qualitative approach, the authors used semi-structured interviews to collect the data. This study employed the quota sampling method to purposefully select students to interview. Data coding and analysis were performed based on the grounded theory. The filmmaking experiences consistently described by the interviewees were labeled and reorganized into categories through the open, axial, and selective coding.@*RESULTS@#The students experience the group filmmaking class as a participatory class. Learners also experienced the procedure of performing complicated group tasks according to detailed and scheduled processes. Participation leads to collaboration. Collaboration here is through communication and participation, not through mechanical cooperation. Students also experience various dimensions of communication. The students learned that successful performance of the group filmmaking process is enabled through consideration towards others, and experience a sense of connectedness resulting in a type of community spirit. Having fun and interest, finally, the students experience the sense of accomplishment and sharing through joint screening.@*CONCLUSION@#Students' shared experiences and their education effects of the filmmaking class can be explained in terms of the above mentioned seven closely intertwined categories. In this class, the students were able to express emotions they would not normally express. Through this, the students were able to find the true character and new aspects of their fellow students, forming intimacy, which led to a sense of belonging and connectedness.

Connected in cinema: educational effects of filmmaking classes on medical students / 한국의학교육

PURPOSE: The purpose of this study is to explore what the students experienced in short filmmaking class performed to 1st grade premedical students of a medical school, and to trace educational effects of the class. METHODS: Taking a qualitative approach, the authors used semi-structured interviews to collect the data. This study employed the quota sampling method to purposefully select students to interview. Data coding and analysis were performed based on the grounded theory. The filmmaking experiences consistently described by the interviewees were labeled and reorganized into categories through the open, axial, and selective coding. RESULTS: The students experience the group filmmaking class as a participatory class. Learners also experienced the procedure of performing complicated group tasks according to detailed and scheduled processes. Participation leads to collaboration. Collaboration here is through communication and participation, not through mechanical cooperation. Students also experience various dimensions of communication. The students learned that successful performance of the group filmmaking process is enabled through consideration towards others, and experience a sense of connectedness resulting in a type of community spirit. Having fun and interest, finally, the students experience the sense of accomplishment and sharing through joint screening. CONCLUSION: Students' shared experiences and their education effects of the filmmaking class can be explained in terms of the above mentioned seven closely intertwined categories. In this class, the students were able to express emotions they would not normally express. Through this, the students were able to find the true character and new aspects of their fellow students, forming intimacy, which led to a sense of belonging and connectedness.

Expression of ciliary neurotrophic factor and its receptor in experimental obstructive nephropathy / 대한해부학회지

Ciliary neurotrophic factor (CNTF) is well known as a growth/survival factor of neuronal tissue. We investigated the expression of CNTF and its specific receptor alpha (CNTFRalpha) in a unilateral ureteral obstruction (UUO) model. Complete UUO was produced by left ureteral ligation in Sprague-Dawley rats. The animals were sacrificed on days 1, 3, 5, 7, 14, 21, and 28 after UUO. The kidneys were fixed, and processed for both immunohistochemistry and in situ hybridization. CNTF immunoreactivity in sham-operated kidneys was observed only in the descending thin limb (DTL) of the loop of Henle. In UUO kidneys, CNTF expression was induced in the S3 segment (S3s) of the proximal tubule from day 1, and progressively expanded into the entire S3s and a part of the convoluted proximal tubules, distal tubules (DT), and glomerular parietal epithelium up to day 7. Upregulated CNTF expression was maintained to day 28. From day 14, the inner medullary collecting duct showed weak CNTF immunoreactivity. The CNTFRalpha mRNA hybridization signal in sham-operated kidneys was weakly detected in the DTL, DT, medullary thick ascending limb, and in a few S3s cells. After UUO, CNTFRalpha mRNA expression increased progressively in both the renal cortex and the medulla up to day 7 and increased expression was maintained until day 28. The results suggest that the S3s may be the principal induction site for CNTF in response to renal injury, and that CNTF may play a role in chronic renal injury.

Expression of calponin in periglomerular myofibroblasts of rat kidney with experimental chronic injuries / 대한해부학회지

Our previous research demonstrated that calponin-immunoreactivity was localized in myofibroblasts of the periglomerular region of human kidney specimens obtained at the time of transplantation from organ recipients. In the present study we examined calponin expression in two chronic nephropathy models, puromycin aminonucleoside (PAN) nephropathy and subtotal nephrectomy (SNx), to investigate the role of calponin in chronic renal injury. Male Sprague-Dawley rats were used, and both nephropathy models were established at 1, 2, 4, and 8 weeks after surgery. There were no periglomerular calponin-positive cells in sham, PAN 1 and 2 week, and SNx 1, 2, and 4 week groups. In SNx 8 week and PAN 4 and 8 week groups, only a few glomeruli with periglomerular calponin-reactivity, which covered half or a very small part of the periglomerular space, were observed. All glomeruli with periglomerular calponin-reactivity showed sclerotic changes, especially thickening of parietal epithelial cells (PECs). In conjunction with our previous report, this data represents the first documentation of the expression of calponin in renal myofibroblasts. We suggest that interactions between PECs and calponin-positive myofibroblasts may play a key role in the late stage of glomerulosclerosis.

Expression of iNOS and NADPH-diaphroase Reactivity in the Lipopolysaccharide Treated Rat Kidney / 대한해부학회지

Inducible nitric oxide synthase (iNOS) has been known to be involved in the various physiological metabolim and has been attracting topic. However, there are extensive differences in the reports about the localization of iNOS expression. To resolve this discrepancy, we compared immunohistochemical data from four iNOS antibody produced by different company (Chemicon, CH; Sigma, SI; Transduction Laboratories, TL; Upstate, UP), and NADPHdiaphorase (NADPH-d) enzyme-histochemical results using light- and transmission electorn-microscope in the lipopolysaccharide (LPS)-treated rat kidney. Electron microscopical examination revealed two different distribution of the NADPH-d reaction product. In the majority of NADPH-d reaction-positive cells, reaction depositions were restricted to the mitochondia, and in the cells of macula densa, descending thin limb (DTL), capsular epithelium (CE) and interstitial wandering cells (WC), NADPH-d positivities were found in the cytoplasm. In immunohistochemical results from LPStreated animal, DTL, CE and WC were positively stained with TL and UP antibodies but with CH and SI antibodies. We conclude that NADPH-d histochemistry may be usefull for identifing the iNOS-positive cells morphologically.

Expression of Nestin in the Rat Kidney with Ischemia-Reperfusion Injury / 대한해부학회지

Nestin, a type VI intermediate filament,is a marker for stem cells.Although it is expressed abundantly in various organs during development,its expression in adults is restricted to certain types of cells.However,nestin is reinduced in activated cells involved in the regeneration and survival of injured tissues.We investigated the expression of nestin in the ischemia-reperfusion injured rat kidney by immunohistochemistry using anti-nestin antiserum. Kidneys were preserved from normal adults and at 1,3,5,7 and 14 d after ischemia-reperfusion injury,and processed using pre-embedding.In the normal adult kidney,nestin was expressed strongly in the glomerular podocyte.Capillary endothelial cells,except for those of the glomeruli,showed nestin positivity.Fibroblast-like interstitial cells were also nestin positive except for lipid-laden interstitial cells of the inner medulla.After ischemia-reperfusion injury,the renal expression of nestin increased progressively to 7 d and then returned almost to a normal level by 14 d.These changes of nestin expression were attributed mainly to changes in the number and staining intensity of immunostained interstitial cells.The podocytes and endothelial cells showed no change in immunoreactivity throughout any stage in the experimental animals.Interestingly,nestin-positive tubular cells,which were nestin negative in normal kidney,were observed from 3 to 14 d.Nestin immunostained cells were increased in the interstitium around these nestin-positive tubules.These results suggest that nestin is induced in both interstitial cells and regenerating tubular cells and that it can be used as a histological marker related to epithelial-mesenchymal transformation in the injured kidney.

The Expression of Interleukin-6 and Its Receptor in the Developing Rat Kidney / 체질인류학회지

Interleukin-6 (IL-6) and its receptor are presumed to play important roles in the developing nervous system. However, little is known about their potential role(s) in the developing kidney. To investigate this, we have studied the expression of IL-6 and its receptor (IL-6R) in the developing rat kidney. Kidneys from 16- (F16), 18- and 20-day-old (F20) fetuses, 1- (P1), 3- (P3), 7- (P7) and 14-day-old (P14) pups, and adult rats were extracted. Renal expressions of IL-6 and its receptors were examined by immunohistochemistry and in situ hybridization respectively. Il-6 protein already appeared in F16. The early stage of renal development before birth, IL-6 showed strong immunoreactivity in the ureteric bud, metanephric mesenchymal cells (MMC) and developing glomerulus. The expression pattern of IL-6 in nephrogenic zone are very similar even after birth. In matured nephron after birth, IL-6 immunoreactivities were detected in distal tubules strongly, and collecting ducts moderately and thick ascending limb weekly. IL-6R hybridization signals have also already appeared in 16-day old fetal kidney. Before birth, IL-6R mRNAs were expressed in ureteric bud, MMC and developing glomerulus. In the matured nephron after birth, IL-6R mRNA was expressed in the thick ascending limb, distal tubules, collecting ducts and S3 segment of proximal tubule. These results suggest that IL-6 and its receptor may be involved in regulation of nephron formation in nephrogenic zone of rat, and play a role in distal nephron including collecting duct after birth.

Expression of mullerian inhibiting substance and its receptor in ovarian neoplsia / 대한산부인과학회지

Mullerian inhibiting substance (MIS) is a glycoprotein hormone produced by fetal Sertoli cells that causes regression of the Mullerian ducts in males during sexual differentiation. Cell lines derived from human ovarian epithelium and rodent Leydig cell tumors, which respond to MIS in growth inhibition assays and express the MIS type II receptors (MISR II). But the pathophysiological role of MIS in human ovarian neoplasia development has not yet been fully established. In order to understand its role in pathogenesis of ovarian neoplasia, the expression and localization of the MIS and MISR II were studied in 5 normal ovaries, 11 benign tumors, 9 borderline ovarian malignancies, 40 ovarian malignancies in paraffin embedded tissue and tissue microarrays by using immunohistochemical stain. The results were as follows; 1. The first staining for MIS and MISR II were detected in granulosa cells in primary follicles of normal ovary. Among the growing follicles, larger developing follicles stained more intensely than smaller follicles. 2. In benign ovarian tumors, 8 (72.73%) in MIS and 5 (45.45%) in MISR II out of 11 cases were stained. The intensity scores of staining were 1.18 in MIS and 0.64 in MISR II. 3. In borderline malignancies, 6 (66.67%) in MIS and 7 (77.78%) in MISR II out of 9 cases were stained. The intensity scores of staining were 0.89 in MIS and 1.22 in MISR II. 4. In ovarian malignancies, the expression of MIS and MISR II were 50% (9/18) and 50% (9/18) in epithelial, 92.30% (12/13) and 76.72% (10/13) in germ cell, and 88.9% (8/9) and 100% (9/9) in sex-cord stromal tumors. The intensity scores of MIS and MISR II expression were 0.72 and 0.72 in epithelial, 1.45 and 1.62 in germ cell, and 1.78 and 1.67 in sex-cord stromal tumors. 5. There was significant high expression of MIS and MISR II in non-epithelial (90.91%, 86.36%) than epithelial ovarian cancers (50%, 50%). The scores of expression intensity was also higher in non-elithelial cancers (MIS: 1.67 +/- 0.16 vs 0.72 +/- 0.20, p=0.003, MISR II: 1.64 +/- 0.20 vs 0.72 +/- 0.21, p=0.022). In conlusion, the expression of MIS and MISR II were not different according to the differentiation, but tissue type specific. The frequency of MIS and MISR II expression was higher in non-epithelial cancers, especially in sex-cord stromal tumors. The results of this experiment could be utilized as scientific basis of researches, furthermore clinical applications in diagnosis and treatment of non-epithelial ovarian malignancies.

Expression of Nestin in the Developing Rat Kidney / 대한해부학회지

Nestin is abundantly expressed in stem cells of the developmental stage of both central nervous system and some non-neuronal organs. The aim of this study was to examine the expression of nestin in the developing rat kidney. Kidneys from 16-(F16), 18- and 20-day-old fetuses, 1-, 3-, 7-, 14-, and 21-day-old pups (P21) and adult were preserved and processed for immunohistochemistry. The nestin was already expressed at all areas of kidney from F16, especially strong in nephrogenic zone. Throughout the development, nestin immunoreactivity was exclusively localized in both glomerulus and interstitium, not in renal tubules. In the vesicle and the S-shaped body stages of the glomerulogenesis, nestin was negative. In the capillary loop stage, the immature podocytes became positive for nestin. Aggregated mesenchymal cells at the root of immature glomeruli were nestin-positive, and then lost the immunoreactivity progressively. In the maturing stage, nestin was expressed only in podocytes. In the renal interstitium except renal papilla, nestin was positive and colocalized with vimentin in almost all the interstitial cells at the prenatal age except both ED-1-positive macrophages and in MHC class II-positive dendritic cells. After birth, the number of nestin-positive cells in the interstitium was decreased, and at P21 pattern of nestin expression in the interstitium was similar with that of adult kidney. In the renal papilla, lipid-laden cells show nestin-negative but vimentin-positive.

Evaluation of Nutrient Intake Quality Over 40 Year-Old People Living in Rural and Suburban Areas / 대한지역사회영양학회지

ABSTRACT To assess the quality of nutrient intake by area of Korean adults, a dietary survey with the 3-day record method was obtained from 324 subjects aged 40 years and older but younger than 70 (52.4 +/- 8.7) living in a rural area (Ansung) and suburban area of a middle-sized city (Ansan). The quality of nutrient intake was assessed by analyzing Nutrient Adequacy Ratio (NAR), Mean Adequacy Ratio (MAR) and Index of Nutritional Quality (INQ). The average daily mean energy intakes were 1,832 kcal for Ansung and 1,842 kcal for Ansan, respectively. Daily intakes of fat for Ansung and Ansan subjects were 40.9 and 40.3 g, and those for protein were 75.1 and 73.1 g, respectively. The overall calorie: protein: fat ratio (CPF) of energy intake was 63 : 17 : 20. Daily mean intakes of protein, fat, calcium, phosphorus, iron, potassium, carotene, sodium, thiamin, and niacin were significantly higher in Ansung residents than in Ansan subjects (p< .05). The average intakes of energy, calcium, vitamin A were lower than Recommend Dietary Allowance (RDA) in both areas. Note, over 30% of the study subjects had less than 75% of RDA of calcium, vitamin A and riboflavin. The MAR was higher in Ansung than Ansan residents (0.86 and 0.85, respectively; p< .05). INQs were over 1 for most nutrients except calcium (0.87), and that of calcium and phosphorus was each significantly higher in Ansung than Ansan subjects. Based on these results, nutrient intake quality of subjects aged 40 to 69 years living in the surveyed rural area is comparable to that of semi-industrialized suburban area in Korea. Dietary deficiency in all of calcium, vitamin A, and riboflavin, however, was a common problem for both rural and suburban residents.

Localization and Morphology of Serotonin Cells in Intestinal Gland of Rodents / 대한해부학회지

This study was an attempt to investigate the relative distribution and morphology of serotonin cells (SC) in intestinal glands of adult rodents, rats, guinea pigs and mice. The intact isolated epithelial sheets of intestinal glands from duodenum, jejunum, ileum, cecum, and proximal and distal colon were prepared for immunohistochemistry using antiserotonin antisera. Examination of isolated epithelia reveals an actual number of SC in one intestinal gland and whole image of individual serotonin cell. In small intestine of all species in this study, the average number of SC per one intestinal gland was the highest in duodenum, and decreased in jejunum and ileum. The distributional patterns of SC in large intestine of three species, however, were different. The number of SC decreased towards distal colon in both rat and guinea pig, and vice versa in mouse. And in the rat, the number of SC in colon was even higher than in duodenum, while in the guinea pig the number of SC in colon was lower than any other part of small intestine. In all the intestinal region of three species, SC were more numerous towards the bases of glands. The open type of SC whose apical cytoplasmic process reach glandular lumen were predominant (over 97% in average) in small intestine of all species in this study. The frequency of closed type was increased in large intestine (up to 44.9% in proximal colon of guinea pig). And closed type was more frequently detected towards the upper part of gland. In small intestine of all species in this study, SC were predominantly flask-like in shape without basal processes. In large intestine, SC with basal processes were often detected, and their frequencies increased towards the upper part of gland. In mice, basal processes were usually long in length (over the long axis of cell), while all the basal precesses of SC of guinea pig were short. We found that the isolated epithelium were very useful to figure out the actual number and whole images of enteroendocrine cells in intestinal mucosal epithelium. The present results demonstrated that relative distribution and morphology of SC were very different among the species especially in large intestine.

Developmental Expression of eNOS in the Rat Renal Vasculature / 대한해부학회지

Nitric oxide (NO) has an important role in maintaining basal renal blood flow (RBF) and glomerular filtration rate (GFR) in the developing kidney. However, renal endothelial NO synthase (eNOS) has not been localized in the developing kidney. The purpose of this study was to examine the expression and localization of eNOS in the developing rat kidney using immunohistochemistry and western blotting. Kidneys from 14 (E14)-, 16 (E16)-, 18 (E18)- and 20-day-old (E20) fetuses, 1 (P1)-, 4 (P4)-, 7 (P7)-, 14 (P14)- and 21-day-old (P21) pups, and adult rats were extracted for immunohistochemistry, and western blot analysis. In the adult rat kidney, eNOS was expressed strongly in the endothelial cells of the arcuate artery and the vascular bundle in the medulla. Endothelial cells of the glomerulus and peritubular capillary network were weakly labeled for eNOS. There was no eNOS immunoreactivity in the uriniferous tubules, including the proximal tubules. In the developing rat kidney, eNOS appeared in the endothelial cells of the capillary network from E14. In the developing glomerular capillary, immunoreactivity for eNOS was observed in the S-shaped bodies (stage II glomeruli) and stage III glomeruli, whereas mature glomeruli (stage IV glomeruli) were faintly immunolabeled for eNOS. These eNOS-positive early-stage developing glomeruli were observed in the nephrogenic zone until seven days after birth. In the endothelial cells of the peritubular capillary network, eNOS was strongly expressed in the fetus and gradually decreased in intensity after birth. The endothelial cells of the arcuate artery were strongly immunoreactive for eNOS from E16 to the adult stages. In the renal medulla, eNOS was expressed in the endothelial cells of the capillary network surrounding the developing medullary collecting ducts of the fetal kidney. After birth, eNOS immunoreactivity gradually disappeared from the vasculature of the renal medulla and only remained in the vasa recta. In conclusion, the strong expression of eNOS in the early stages of the developing vasculature suggests that eNOS may contribute to angiogenesis and/or critically participate in the hemodynamics of the immature kidney.

Expression of FSHR mRNA in female genital organs / 대한산부인과학회잡지

FSH is the pivotal hormone in the regulation of ovarian function and acts by binding to specific receptor, FSH receptor (FSHR), which is belong to the family of G-protein coupled receptor. It have been considered that ovary is the only target organ of FSH because FSHR mRNA was first detected in ovarian follicles. However expression of FSHR mRNA was also detected on fallopian tube in experimental animal study and it is related wih tumorigenesis in postmenopausal women.In this study, in order to understand the FSH function in female genital organs, the ontogeny of the production profile of FSHR and the pattern of its localization in female genital organs were studied. We obtained the fresh tissues of ovary, fallopian tube, uterine body and uterine cervix with blood samples during proliferative phase in women with regular menstrual cycle. To establish FSHR mRNA expression of human internal genital organ, we studied by using in situ hybridization and quantitative competitive reverse transcription polymerase chain reaction (QC RT-PCR). To localize FSHR transcripts by in situ hybridization, we synthesized digoxigenin-labelled ssRNA probe (about 800 bp) from the cloned FSHR cDNA. For QC RT-PCR, we designed oligonucleotide primers (antisense: 5'-GGCCCTGCTCCTGGTCTCTTTG-3', sense: 3'-AACAGCGGGAGTACCTTCGG-5') which produced 799 bp sized PCR products. Simultaneously we synthesized 149 bp deleted DNA competitor by site-directed mutagenesis to quantify target FSHR mRNA expression comparing as internal control.In situ hybridization with digoxigenin-labelled ssRNA probe showed no signal above the background in primordial follicles. FSHR mRNA was first detected in the single layer of cuboidal granulosa cells surrounding primary follicles. As follicular growth progressed, FSHR mRNA expression increased gradually in antral and graafian follicles. Similary, in fallopian tube, the epithelium stained intensly. But FSHR mRNA expression was absent in uterine body including endometrium and myometrium and uterine cervix. Total RNA was extracted and quantitated by QC RT-PCR. The amounts of FSHR transcript measured were 840.00+/-516.29 in the ovarian tissue, 240.00+/-154.91 in the fallopian tube, 6.06+/-4.13 in the uterine body, 5.48+/-5.00 fg in the uterine cervix. These experiments demonstrated that FSHR mRNA is expressed in the ovary and fallopian tube, albeit only small amount was expressed in uterine body and cervix.In conclusion, the presence of FSHR mRNA in female internal genital organ with site specific pattern suggested that FSH may have some role in female genital organs during the adult reproductive cycle and may act as an factor in the tumorigenesis. Further study about the functional role and tumorigenesis of FSH should be performed in human internal genital organ.

Influence of aging on expression of FSHR mRNA in human ovary / 대한산부인과학회잡지

FSH is the central hormone for the regulation of ovarian function and acts by binding with specific receptor, FSHR, which is one of the G-protein coupled receptor family. The aging of ovary decreases the number and the activity of follicle, which results in the increase of FSH by reduction of inhibin and estrogen. The study on FSH level and FSHR mRNA expression in the ovary of perimenopausal women is the crucial step for the understanding of the menopausal mechanism.We studied FSHR mRNA expression of ovarian follicle by using in situ hybridization and QC RT-PCR. The fresh ovarian tissues and blood samples were obtained from premenopausal women in mid-follicular stage and postmenopausal women. The experimental samples were grouped as below 40 years old women, 40-44 years old ones, 45-49 years old ones, 50-54 years old ones, and postmenopausal women as negative control. To localize FSHR transcripts by in situ hybridization, we synthesized digoxigenin-labelled ssRNA probe (about 800 bp) and measured the degree of staining as 0, 1+, 2+ in the primary follicles which were independent to FSH effect. To do QC RT-PCR, we synthesized oligonucleotide primers (antisense: 5-GGCCCTGCTCCTGG- TCTCTTTG-3, sense: 3-AACAGCGGGAGTACCTTCGG-5) to form the 799 bp sized PCR products. We also synthesized 149 bp deleted DNA competitor by site-directed mutagenesis and then calculated the relative amount of target FSHR mRNA by comparing with competitor after PCR.There were significant reverse relationships between follicular number and aging (r=-0.934, P=0.01), and FSH level (r=-0.713, P<0.001). The similar amount of FSHR mRNA was expressed in the group of below 40 years by in situ hybridization. In the groups of above 40 years, the FSHR mRNA expression decreased progressively according to aging (r=-0.744, P<0.001) and FSH level (r=-0.771, P<0.001). But we could not find FSHR mRNA expression in menopausal ovaries. The amount of follicular FSHR mRNA was measured as 840.00+/-516.29 for the below 40 years group, 240.00+/-154.91 for the 40-44 years group, 40.00+/-21.90 for the 45-49 years group, 6.06+/-4.13 for the 50-54 years group, and 0.48+/-0.00 fg in the postmenopausal ovary. The amount of FSHR mRNA decreased with ovarian aging (r=-0.857, P<0.001) and FSH level (r=-0.771, P<0.001).These results demonstrate that the gradual increase of FSH and the decrease of FSHR mRNA expression in older than 40 years women are related to the changes of sex hormones. However the gradual decrease of the FSHR mRNA expression in the primary follicle may be due to the follicular aging itself. Therefore the menopausal transition already starts at the beginning of 40 years and one of the major cause of the menopause may be the reduction of FSHR mRNA expression followed the decrease of ovarian response to gonadotropins. The further studies should be required to elucidate the underlying mechanism and the associated factors of menopause.

Expression of alphasmooth Muscle Actin, Aquaporin 1 and Renin after Blocking AT1 Receptor in Developing Renal Arterial System of Rat / 대한신장학회잡지

BACKGROUND: Recent studies have demonstrated that renin, alphasmooth muscle actin(ASMA) and aquaporin-1(AQP1) participate in the development of renal arterial system. The components of the renin- angiotensin system have been shown to function as growth factors, apart from their classical roles in controlling blood volume and homeostasis. Interestingly, the vasoconstrictor angiotensin II(ANG II) appears to participate in the regulation of angiogenesis in various tissues. The present study examined the effect of ANG II type-1(AT1) receptor blocker losartan given during pregnancy or newborn rats on the expression of renin, ASMA and AQP1 in the developing renal arterial system. METHODS: Pregnant and newborn rats received losartan(10 mg/kg/day) or saline for 4 and 8 days from E14 to parturition, and for 4 and 9 days starting at day 1 after birth, respectively. Kidneys of 17-day-old fetuses and 1-, 4-, and 9-day- old pups were processed for immunohistochemistry using antibodies to renin(1 : 10,000), ASMA(1 : 1,000), and AQP1(1 : 1,000). RESULTS: In all pregnant groups, there were no differences in immunostaining for renin, ASMA, and AQP1 between losartan treated groups and saline treated groups. In all newborn groups, however, blockade of AT1 receptor with losartan found to increase expression of renin and ASMA but to have no effect on expression of AQP1 in the developing renal arterial system. CONCLUSION: These results suggest that AQP1 expression is not associated with renin or ASMA expression during development of renal arterial system.

Developmental Expression of Epidermal Growth Factor in Rat Kidney / 대한해부학회지

Epidermal growth factor (EGF) is well known to be one of regulating factor in proliferation. The aims of the present study were to elucidate the expression time and localization of expression of EGF in developing rat (Sprague-Dawley) kidney. Kidneys of 16-, 18- and 20-day-old fetuses, 1-, 3-. 7-, 14-, and 21-day-old pups and adult (3months) were preserved for light and electron microscopic immunocytochemistry. In adult rat kidney, EGF immunoreactivity was well localized in thick ascending limb and distal convoluted tubule. In developing rat kidney, EGF-positive cells appeared first in cortical thick ascending limbs and distal convoluted tubules of juxtamedullary nephrons at 3 days and 7 days after birth respectively, in cortical thick ascending limbs and distal convoluted tubules of cortical nephrons at 14 days after birth, and in medullary thick ascending at 21 days after birth. These findings suggest that renal EGF synthesized and secreted from distal tubule may possibly accelerate not proliferation but differentiation of the renal tubular epithelium at least in neonatal rat kidney.

Effect of Furosemide on the Morphogenesis of Neonatal Rat Renal Papilla / 대한해부학회지

In developing rat kidney, apoptosis plays an important role in the morphogenesis of renal papilla, and hyperosmolality is well known to be one of regulating factor in apoptosis. The aim of this study was to determine the effect of hypoosmolality induced furosemide in neonate on cell proliferation and apoptosis in renal papilla. One-dayold pups were given two times a day, at 12 hour intervals, subcutaneous injection of furosemide (10 mg/kg BW) or saline for 4 days or 7days. We identified thick ascending limb by labeling with antibody to BSC1 (bumetanide-sensitive Na/K/2Cl cotransporter) and type A intercalated cell in collecting duct by labeling with antibody to band 3 protein. We also inspected apoptosis with TdT-mediated dUTP nick end labeling (TUNEL) method and cell proliferation with immunostaining for proliferating cell nuclear antigen (PCNA). The effect of hypoosmolality induced furosemide on neonatal rat kidney. 1) In furosemide-treated animals, body and kidney weights were reduced compare to control groups. the length of renal papillae of furosemide-treated groups were shortened compare to control groups. 2) Transformation from a cuboidal epithelium of thick ascending limb to a squamous epithelium of ascending thin limb in renal papilla was delayed in furosemide-treated groups. 3) In the inner medullary collecting duct of furosemide-treated groups, elimination of type A intercalated cells was delayed compared to control groups. 4) Furosemide treatment reduced the apoptotic index in transforming thick ascending limb of Henle's loop and collecting duct in renal medulla. 5) PCNA-positive cells in the transforming thick ascending limb of Henle's loop and the collecting duct in renal medulla were decreased in number in furosemide-treated groups compared to control groups. These finding suggest that renal hypoosmolality induced furosemide treatment decrease not only apoptosis but also cell proliferation and may retard renal papilla growth at least in neonatal rat kidney.

Developmental Expression of Renin and Aquaporin-1 in the Rat Renal Vasculature / 대한신장학회잡지

Recent studies in the developing rat kidney have demonstrated that renin in juxtaglomerular cells and aquaporin-1(AQP1), a kind of water channel, participate in the development of renal arterial system. The purpose of this study was to identify the association among renin and AQP1 in the developing renal arterial system. Sprague-Dawley rat kidneys from 14-, 16-, 18- and 20-day-old fetuses, and 1-, 4-, 7-, 14-, 21- and 28-day-old pups were preserved with periodate-lysine-2% paraformaldehyde solution for single or multiple immunohistochemistry. Renin- positive, smooth muscle, and endothelial cells were detected using renin polyclonal(1 : 5,000), alpha-smooth muscle actin(ASMA) monoclonal(1 : 1,000), and AQP1 polyclonal(1 : 500) antibodies, respectively. Immunoreactivity for renin and AQP1 was not detected in the developing vessels in fetal kidneys on the 14 th day of gestation. At the 16th day of gestation, AQP1 appeared in the developing kidney. Immunoreactivity for AQP1 was observed in endothelial cells of the arterial capillary plexus and of the arcuate artery, but not in the venous capillary plexus or the arcuate vein. At the 18th day of gestation, renin-positive cells appeared throughout the arterial system including the arcuate artery. After birth, immunoreactivity for renin and AQP1 was gradually decreased in the developing artery. No renin immunoreactivity was detected in the arcuate artery from 7 days after birth, and in the interlobular artery from 14 days after birth. Renin immunoreactivity was confined to juxtaglomerular cells in the afferent arteriole from 21 days after birth. AQP1 immunoreactivity in endothelial cells was not observed in the arcuate artery from right after birth, or in the interlobular artery, or afferent arteriole from 14 days after birth. This study indicates that the expression and loss of AQP1 in endothelial cells spatiotemporally coexists with renin in smooth muscle cells during the development of arterial system in the rat kidney. It is suggested that AQP1 and renin might play an important role in the development and growth of the rat kidney arterial system by functioning through a paracrine or autocrine mechanism.

Expression of Osteopontin in the Adult and Developing Rat Kidney / 대한해부학회지

Osteopontin (OPN), originally considered to be a bone protein, is now reported to be expressed in other tissues, notably in kidney. OPN has been demonstrated in the kidney by Northern and Western analyses, immunohistochemistry and in situ hybridization. However, studies of the cellular distribution of OPN in the kidney, especially in normal condition, have given highly conflicting results. This study is designed to establish the expression of OPN in kidney from the fetus to adult. Kidneys from 16-(F16), 18-(F18), and 20 day-old (F20) fetuses and 1-(P1), 3-(P3), 7-(P7), 14-(P14), 21 day-old pups, and adult were studied by in situ hybridization and immunohistochemistry. OPN mRNA and protein were expressed in the thick ascending limb (TAL) at F18 and F20, but disappeared gradually after birth. In the collection duct (CD), weak labeling appeared in a few cells at F20. After birth cells with strong labeling were scattered throughout the CD in the medulla and inner cortex at P1-P7 and in the outer cortex at P14. There was little OPN expression in the CD at P21. OPN mRNA and immunoreactivity appeared in the papillary surface epithelium (PSE) at F20 and in the descending thin limb (DTL) at P1. After birth OPN expression gradually increased to adult levels in the PSE. In the DTL, adult levels of expression were reached at P21. Proximal tubules exhibited a punctate subapical OPN immunos-taining from F18, but no hybridization signal. During renal development, the transient expression of OPN in the TAL and CD suggests that OPN may have a role in the developing kidney, and from P21 OPN expression was localized at DTL and PSE exclusively, which was similar to adult OPN expression pattern.