생쥐 턱밑샘의 발생, 분화 및 노화에 따른 C-type natriuretic peptide와 natriuretic peptide 수용기의 발현양상

복세미(Se-Mi Bok),김택헌(Tak-Heun Kim),박기랑(Kee-Rang Park),조의식(Eui-Sic Cho) 대한해부학회 2005 Anatomy & Cell Biology Vol.38 No.4

C-type natriuretic peptide (CNP)는 natriuretic peptide family의 하나로서 주로 중추신경계통과 혈관내피세포에서 국소적으로 합성되는 것으로 알려져 있으나 최근 다른 말초기관에서도 합성되어 세포의 성장과 분화 조절에 관여하는 것으로 알려지고 있다. 본 연구자들은 CNP가 정상 성체 턱밑샘의 사이관과 곱슬과립세관세포에서 국소적으로 합성되어 분포하며 생물학적 활성을 보임을 보고한 바 있으나, 발생 및 분화, 그리고 노화에 따르는 발현 및 활성에 대해서는 알려진 바 없다. 따라서 본 연구는 턱밑샘의 발생 및 분화과정에서 CNP와 그 수용기의 발현양상과 노화과정에서 그 수용기의 생물학적 활성변화를 알아보고자, 임신14, 16, 18일된 C57BL/6N 생쥐 배자와 출생 후 1일, 2주, 그리고 1, 2, 12, 24개월된 성체의 턱밑샘을 대상으로 RT-PCR, in situ hybridization, 면역조직화학법 그리고 cGMP assay를 시행하였다. CNP는 임신14일에 턱밑샘의 원기인 외배엽성상피싹 종말부위에서 강하게 발현되었으며 출생 전까지 높게 유지되었으나 출생 후 현저히 감소되었다. CNP는 성체 턱밑샘의 사이관과 곱슬과립세관에 분포하고 있었으며 이들 관계통에서 NPRC는 특이적으로 발현되었으나 NPRB는 발현되지 않았다. RT-PCR결과 턱밑샘에서 CNP전사체는 임신16일부터 출생 후 2개월까지 점차 감소하였으나 ANP전사체는 증가하였다. CNP에 특이적인 수용기인 NPRB와 NPRC는 CNP와 동일한 발현양상이었으나 NPRA는 약하게 발현되었다. 또한 CNP는 치아싹, 혀, 앞위턱뼈, 뼈형성 부위와 같은 두개안면 조직에서도 발현되었으며, 이들 부위에서도 NPRC는 발현되었으나 NPRB는 발현되지 않았다. 출생 후 2개월된 턱밑샘의 관세포는 CNP에 의해 현저히 많은 cGMP를 생성하였으나, 샘꽈리는 CNP보다는 ANP와 BNP에 의해 많은 cGMP를 생성하였다. 노화에 따라 턱밑샘의 cGMP 활성능력은 전반적으로 감소되었으며 출생 후 1개월된 턱밑샘에서는 CNP, 출생 후 2개월 이후로는 ANP에 의한 cGMP 활성이 우세하였다. 이상의 결과로 보아 CNP는 턱밑샘에서 NPRC를 매개로 하는 발생 및 분화유도 기능을 주로 수행하며, 성체이후에는 NPRB를 매개로 하는 관계통 유지에 관여하고 노화에 따라서 활성이 점차 감소하는 것으로 추정된다. C-type natriuretic peptide (CNP), a member of natriuretic peptide family, is mainly synthesized in the endothelium and central nervous system. But CNP is also involved in the growth and differentiation of other peripheral organs. Although we have reported the local synthesis and localization of CNP in the adult submandibular glands (SMG), it is not known that the expression and biological activity of CNP following the morphogenesis, differentiation and aging. This study aimed to examine the expression of CNP and its receptors in the developing and differentiating stages of mouse SMG, and the changes of biological activity of its receptors with aging. The SMG, obtained from 14, 16, 18 days-old embryos (E) and 1 day, 2 weeks, 1, 2, 12, and 24 month-old C57BL/6N mouse, were processed for RT-PCR, in situ hybridization, immunohistochemistry and cGMP assay. CNP was strongly expressed in the epithelial clusters of primitive SMG, which was maintained before birth but was markedly decreased after birth. CNP was localized in the intercalated duct and granular convoluted tubules of adult SMG, where NPRC was specifically expressed but NPRB was not. CNP mRNA was gradually decreased from E16 to 2 M but ANP mRNA was opposed. NPRB and NPRC were the same pattern of the expression of CNP but NPRA was weakly expressed. In addition, CNP mRNA was also expressed in the craniofacial tissues such as tooth germs, tongue, premaxilla and bone forming area in which NPRC was specifically expressed but NPRB was not. In the SMG of 2 M, the membrane of duct cells markedly produced cGMP by CNP whereas acini produced cGMP by ANP and BNP rather than CNP. The biological activity of cGMP production of SMG gradually decreased with age. cGMP production was dominant by CNP in SMG of 1M but was by ANP after 2M. These results shows that CNP may play roles both in the morphogenesis and differentiation via NPRC and in the maintenance of duct system via NPRB in the mouse SMG and that the biological activity of its receptors may decreased with aging.

Expression Patterns of Tbx1 during Development of the Mouse Mandibular First Molar

복세미(Se-Mi Bok),정경철(Kyung-Chul Chung),조의식(Eui-Sic Cho) 대한해부학회 2006 Anatomy & Cell Biology Vol.39 No.3

Tbx1은 T-box transcription factor gene의 하나로서, 사람 염색체 22q11의 결실증후군으로 cardiac outflow tract의 기형과 가슴샘 및 부갑상샘의 저형성, 입천장갈림, facial dysmorphogenesis, 인두굽이와 인두주머니의 비정상적 발생을 포함하는 DiGeorge/velocardiofacial syndrome의 주요한 후보유전자로 알려져 있다. 본 연구에서는 치아의 발생과정에서 Tbx1의 발현양상을 알아보고자, E13.5~E18.5 시기의 생쥐 배아를 대상으로 in situ hybridization을 시행하였다. 아래턱 어금니 치아싹의 분화에 따른 Tbx1의 발현양상을 알아본 결과, 싹시기 치아싹에서는 치아판과 주변 중간엽조직에서 발현되었으며, 모자시기 치아싹에서는 사기질결절을 제외한 치아기에서 강하게 발현되었다. 초기 종시기 치아싹에서는 치아싹목고리와 중간층에서 강하게 발현되었으나 치아유두와 치아주머니에서는 약하게 발현되었고, 후기 종시기 치아싹에서는 별그물을 제외한 치아기 조직에서 매우 강하게 발현되었으며 속사기질상피에 인접한 치아유두 세포와 치아주머니 세포에서도 발현되었다. 어금니의 발생과정에서 이와같은 Tbx1의 발현은 Sox9의 발현양상과 유사하였으나 Pitx2와 ectodin의 발현과는 다른 양상이었다. 이상의 결과로 볼 때, Tbx1은 치아싹의 발생과정동안에 치아판 형성 초기부터 치관형성기까지 주로 상피 특이적으로 발현되나 치아 중간엽조직에서도 약하게 발현됨을 확인하였다. 따라서 Tbx1은 치아발생 초기부터 치관형성기까지 상피세포 뿐만 아니라 중간엽 세포에서 지속적으로 발현하여 이들의 증식과 분화과정을 조절함으로써 치아형성에 기여할 것으로 추정된다. A T-box transcription factor gene, Tbx1 is a principal candidate of the most frequent chromosomal deletion syndrome found in human, DiGeorge/velocardiofacial syndrome which is a complex developmental disorder associated with cardiac outflow tract abnormalities, mid facial dysmorphology, velopharyngeal insufficiency and submucosal cleft palate. We performed in situ hybridization against mouse embryo from E13.5 (bud stage) to E18.5 (late bell stage) in order to analyze the expression patterns of Tbx1 in the developing mouse first molar, a derivative of the first pharyngeal arch. Tbx1 transcripts were found in the dental lamina and its surrounding mesenchyme at E13.5 and in the dental organ except enamel knot at E14.5 (cap stage). Tbx1 was strongly expressed in the cervical loop and stratum intermedium but was weak in the dental papilla and dental follicle at E15.5 (early bell stage). At E18.5, Tbx1 was strongly expressed not only in the dental organ (bell stage) except stellate reticulum but also dental papilla and dental follicle adjacent to the inner dental epithelium. In conclusion, Tbx1 transcripts were specifically expressed both in the dental epithelium and surrounding mesenchyme of developing tooth from initiation to bell stage, which were the most similar with those of Sox9 but little different from those of Pitx2 and ectodin. These results strongly suggested that Tbx1 may play a role as a transcription factor regulating proliferation and differentiation of both dental epithelium and mesenchyme through the tooth development.

Bleomycin 유도 폐 섬유화 쥐 모델에서 미세 전산화단층촬영의 유용성

이재아 ( Jae A Lee ),진공용 ( Gong Yong Jin ),복세미 ( Se Mi Bok ),한영민 ( Young Min Han ),박성주 ( Seoung Ju Park ),이용철 ( Yong Chul Lee ),정명자 ( Myung Ja Chung ),윤건하 ( Gun Ha Youn ) 대한결핵 및 호흡기학회 2009 Tuberculosis and Respiratory Diseases Vol.67 No.5

Background: Micro computed tomography (CT) is rapidly developing as an imaging tool, especially for mice, which have become the experimental animal of choice for many pulmonary disease studies. We evaluated the usefulness of micro CT for evaluating lung fibrosis in the murine model of bleomycin-induced lung inflammation and fibrosis. Methods: The control mice (n=10) were treated with saline. The murine model of lung fibrosis (n=60) was established by administering bleomycin intra-tracheally. Among the 70 mice, only 20 mice had successful imaging analyses. We analyzed the micro CT and pathological findings and examined the correlation between imaging scoring in micro CT and histological scoring of pulmonary inflammation or fibrosis. Results: The control group showed normal findings on micro CT. the abnormal findings on micro CT performed at 3 weeks after the administration of bleomycin were ground-glass opacity (GGO) and consolidation. At 6 weeks after bleomycin administration, micro CT showed various patterns such as GGO, consolidation, bronchiectasis, small nodules, and reticular opacity. GGO (r=0.84) and consolidation (r=0.69) on micro CT were significantly correlated with histological scoring that reflected pulmonary inflammation (p<0.05). In addition, bronchiectasis (r=0.63) and reticular opacity (r=0.83) on micro CT shown at 6 weeks after bleomycin administration correlated with histological scoring that reflected lung fibrosis (p<0.05). Conclusion: these results suggest that micro CT findings from a murine model of bleomycin-induced lung fibrosis reflect pathologic findings, and micro CT may be useful for predicting bleomycin-induced lung inflammation and fibrosis in mice.