• Asian Pacific Journal of Cancer Prevention
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• 제16권17호
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• pp.7575-7582
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• 2015

Cyclin E, a key coordinator of the G1 to S transition in the cell cycle, may be deregulated in several malignancies, including breast cancer. The most significant aberration in cyclin E is its elastase mediated proteolytic cleavage into tumor specific low molecular weight isoforms (LMW-Es). LMW-Es are biochemically hyperactive and biologically drive tumorigenesis in transgenic mouse models. Additionally, expression of LMW-Es has been correlated with poor survival in breast cancer cases. Here we determine whether expression of LMW-Es in a breast cancer cell line that is naturally devoid of these deregulated forms would alter their progression through each phase of the cell cycle. The results revealed that LMW-Es expression resulted in an increased doubling time, concomitant with a predominant increase in the population in the S phase of the cell cycle. Moreover, downregulation of p53 in LMW-Es cells resulted in additional shortening of the doubling time and enrichment of cells in the S and G2/M phases of the cell cycle. Furthermore, expression of LMW-Es sensitized cells to ${\beta}$-estradiol (E2) mediated growth and changed expression patterns of estrogen receptor and Bcl-2. Intriguingly, expression of LMW-Es could surpass anti-apoptotic effects raised by p53 upregulation. Taken together these studies suggest that overexpression of LMW-Es in collaboration with p53 loss results in altered g rowth properties of MCF-7 cells, enhancing the oncogenic activity of these ER positive breast cancer cells.

• 한국발생생물학회지:발생과생식
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• 제19권2호
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• pp.69-77
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• 2015

Cell-cell direct communication through channel-forming molecules, connexin (Cx), is essential for a tissue to exchange signaling molecules between neighboring cells and establish unique functional characteristics during postnatal development. The corpus epididymis is a well-known androgen-responsive tissue and involves in proper sperm maturation. In the present research, it was attempted to determine if expression of Cx isoforms in the corpus epididymis in the adult is modulated by exposure to estrogenic or anti-androgenic compound during the early postnatal period. The neonatal male rats at 7 days of age were subcutaneously injected by estradiol benzoate (EB) at low-dose ($0.015{\mu}g/kg$ body weight) or high-dose ($1.5{\mu}g/kg$ body weight) or flutamide (Flu) at low-dose ($500{\mu}g/kg$ body weight) or high-dose (50 mg/kg body weight). The corpus epididymis collected at 4 months of age was subjected to evaluate expressional changes of Cx isoforms by quantitative real-time PCR. Treatment of low-dose EB resulted in increases of Cx32, Cx37, and Cx45 transcript levels, while exposure to high-dose EB decreased expression of Cx26, Cx30.3, Cx31, Cx31.1, Cx32, Cx40, Cx43, and Cx45. Treatments of Flu caused significant decreases of expression of all examined Cx isoforms, except Cx37 and Cx43 shown no expressional change with high-dose Flu treatment. These findings imply that expression of most Cx isoforms present in the corpus epididymis would be transcriptionally regulated by actions of androgen and/or estrogen during postnatal period.

• 한국발생생물학회지:발생과생식
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• 제19권1호
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• pp.43-51
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• 2015

Connexin (Cx) is a complex which allows direct communication between neighboring cells via exchange of signaling molecules and eventually leads to functional harmony of cells in a tissue. The initial segment (IS) is an excurrent duct of male reproductive tract and expression of numerous genes in the IS are controlled by androgens and estrogens. The effects of these steroid hormones on gene expression in the IS during postnatal development have not extensively examined. The present research investigated expressional modulation of Cx isoforms in the IS by exogenous exposure to estrogen agonist, estradiol benzoate (EB), or androgen antagonist, flutamide (Flu), at weaning age. Two different doses of EB or Flu were subcutaneously administrated in 21-day old of male rats, and expressional changes of Cx isoforms in the adult IS were analyzed by quantitative real-time PCR. Treatment of a low-dose EB ($0.015{\mu}g/kg$ body weight) resulted in an increased expression of Cx31 gene and a decreased expression of Cx37 gene. A high-dose EB ($1.5{\mu}g/kg$ body weight) treatment caused an increase of Cx31 gene expression. Increased levels of Cx30.3 and Cx40 transcripts were observed with a low-dose Flu ($500{\mu}g/kg$ body weight) treatment. Treatment of high-dose Flu (50 mg/kg body weight) led to expressional increases of Cx30.3, 40, and 43 genes. Our previous and present findings suggest differential responsiveness on gene expression of Cx isoforms in the IS by androgens and estrogens at different postnatal ages.

• 한국발생생물학회지:발생과생식
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• 제20권4호
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• pp.349-357
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• 2016

The present research was chiefly designed to determine the effect of the treatment of estrogenic agonist, estradiol benzoate (EB), or antiandrogenic compound, flutamide (Flu), at the weaning age on the expression of connexin (Cx) isoforms in the caudal epididymis of adult male rat. Animals were subcutaneously administrated with a single shot of either EB at a low-dose ($0.015{\mu}g$ of EB/kg body weight (BW)) or a high-dose ($1.5{\mu}g$ of EB/kg BW) or Flu at a low-dose ($500{\mu}g$ of EB/kg BW) or a high-dose (5 mg of EB/kg BW). Expressional changes of Cx isoforms in the adult caudal epididymis were examined by quantitative real-time PCR analysis. The treatment of a low-dose EB caused significant increases of Cx30.3, Cx31, Cx32, and Cx43 transcript levels but reduction of Cx31.1, Cx37, and Cx45 expression. Exposure to a high-dose EB resulted in very close responses observed in a low-dose EB treatment, except no significant expressional change of Cx37 and a significant induction of Cx40. Expression of all Cx isoforms, except Cx45, was significantly increased by a low-dose Flu treatment. Expressional increases of all Cx isoforms were detected by a high-dose Flu treatment. The current study demonstrates that a single exposure to estrogenic or antiandrogenic compound during the early postnatal developmental period is sufficient to disrupt normal expression of Cx isoforms in the adult caudal epididymis.

• 한국발생생물학회지:발생과생식
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• 제21권3호
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• pp.317-325
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• 2017

Direct communication between neighboring cells through connexin (Cx)-based gap junction is a crucial biological manner to regulate functions of a tissue consisting of multi-cell types. The present research evaluated expressional changes of Cx isoforms in the caput epididymis of adult rat exposed to estradiol benzoate (EB) or flutamide (Flu) at the early postnatal age. A single subcutaneous administration of EB at a low-dose [$0.015{\mu}g/kg$ body weight (BW)] or a high-dose ($1.5{\mu}g/kg\;BW$) or Flu at a low-dose ($500{\mu}g/kg\;BW$) or a high-dose (5 mg/kg BW) was performed to an animal at 1 week of age. Quantitative real-time PCR analysis was employed to determine expressional changes of Cx isoforms. The transcript levels of Cxs30.3 and 37 were decreased by a low-dose EB treatment, while decreases of Cxs31, 31.1, 32, 40, and 45 transcript levels were observed with a low-dose EB treatment. The treatment of a high-dose EB resulted in expressional reduction of Cxs30.3, 31, 31.1, 37, 40, 43, and 45. The Flu treatment at a low dose caused increases of Cxs26, 37, and 40 transcript levels but decreases of Cxs31.1, 43, and 45 transcript levels. Increases of Cxs30.3, 31, 37, and 40 mRNA amounts were induced by a high-dose Flu treatment. However, exposure to a high-dose Flu produced expressional decreases of Cxs31.1, 32, and 43 in the adult caput epididymis. These observations suggest that exposure to EB or Flu at the neonatal period could lead to aberrant expression of Cx isoforms in the adult caput epididymis.

• 한국발생생물학회지:발생과생식
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• 제21권4호
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• pp.379-389
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• 2017

Connexin (Cx) involves in the regulation of various physiological functions of tissue by forming a channel, a gap junction which allows direct cell-cell communication, between adjacent cells. The effect of a single subcutaneous treatment of estradiol benzoate (EB) or flutamide (Flu) at the weaning age on the expression of Cx isoforms in the adult caput epididymis was evaluated in this research. Using quantitative real-time PCR analysis, a low-dose of EB [$0.015{\mu}g/kg$ body weight (BW)] caused significant decreases of Cx30.3, Cx32, Cx40, Cx43, and Cx45 mRNA levels and no change of Cx26, Cx31, Cx31.1, Cx37 transcript levels. The treatment of a high-dose EB ($1.5{\mu}g/kg\;BW$) resulted in reduced expression of Cx30.3, Cx31, Cx43, and Cx45 but increased expression of Cx37 and Cx40. Expression of all Cx isoforms examined, except Cx31, was significantly increased by the treatment of a low-dose Flu ($500{\mu}g/kg\;BW$). However, the treatment of a high-dose Flu (5 mg/kg BW) led significant expressional suppression of Cx30.3, Cx31, Cx31.1, Cx32, Cx40, Cx43, and Cx45 but an increase of Cx37 transcript level. With the comparison of previous findings, the expression of Cx isoforms in the adult epididymis after the exposure to EB or Flu is likely differentially regulated in regional-specific and/or exposed postnatal age-specific manner.

• 생명과학회지
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• 제13권5호
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• pp.658-667
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• 2003

환경 스트레스에 대해 내성을 가지는 것으로 알려진 명아주과에 속하는 근대(지상부길이 15 cm)를 이용하여, 다양한 염 농도에서의 건물함량 측정을 통한 생장반응과 항산화 효소(SOD, APX, GR)의 효과를 밝히기 위하여 다양한 농도(0, 50, 200, 1000 mM NaCl)의 염을 처리한 후 24시간 동안의 효소의 활성변화를 측정하였다. 근대는 처리 2시간째 200 mM NaCl처리구 에서 SOD, APX, GR의 최대활성을 보였으며, 50 mM NaCl처리구에서 가장 낮은 활성을 나타내었다. PAGE에 의한 isoforms의 확인결과, 근대는 3개의 SOD isoforms(Fe-SOD, CuZn-SOD, Mn-SOD)를 함유하고 있었으며, major isoform은 CuZn-SOD로 밝혀졌다. APX의 경우, 9개의 bands 중 특별히 저분자 isoforms (No. 7,8)의 강한 발현양상을 보였다. SOD의 경우 50 mM NaCl처리에서 Mn-SOD isoform의 불활성을 보여 활성의 증감에 있어 Mn-SOD가 직접적인 연관성을 가질 것으로 생각된다. 근대의 항산화 효소는 염 처리후 단시간내 효소 활성의 증가양상(특별히 처리후 2시간째 200 mM NaCl처리구)을 보여, 고농도 염 환경하에서 항산화시스템의 빠른 작동을 통해 염스트레스에 의해 생성된 활성산소를 제거함으로써 염에 의한 산화적 스트레스에 대해 효과적으로 대처해 나가는 것으로 생각된다. 검색어-근대, 염, 활성산소, SOD, APX, GR.

• Applied Biological Chemistry
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• 제41권7호
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• pp.505-509
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• 1998

Pathogenesis-related proteins중 chitinase를 지황에서 추출하고 분류하였다. 지황 chitinase는 chitin affinity column chromatography로 염기성 및 산성 두개 군으로 분류되었으며 native PAGE gel상 효소활성을 조사한 결과 이동도가 작은 염기성 chitinase는 pH 2.9의 산성추출 조건에서, 이동도가 큰 산성은 pH 8.8의 염기성추출 조건에서 상대적으로 많이 추출되었다. 지황에는 염기성 3개, 산성 4개등 총 7개의 chitinase isoform이 존재하며 이들의 분자량은 $28{\sim}32\;kD$내외이고 지상부와 달리 뿌리에는 주로 염기성 chitinase만 검출되어 일부 산성효소는 조직 특이적으로 발현될 가능성을 보여주고 있다.

• Asian-Australasian Journal of Animal Sciences
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• 제16권7호
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• pp.962-967
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• 2003

The purpose of the study were to characterize the proteins secreted by elongating caprine conceptus, to identify a group of low molecular weight proteins as retinol-binding protein (RBP), to identify RBP cell-specific localization in conceptus tissue, and to demonstrate that the conceptuses secreted continuously RBP during the time period maternal recognition of pregnancy. Caprine conceptuses were removed from the uterus between days 16 and 22 of pregnancy, the time period maternal recognition of pregnancy. Isolated conceptuses were cultured in a modified minimum essential medium in the presence of radiolabeled amino acids. Proteins synthesized and secreted into medium were analyzed by fluorography of two-dimensional polyacrylamide gel electrophoresis and fluorography. At least five proteins showed consistently a grouping of spots with characteristic location on two-dimensional gels. A major low molecular weight protein consisted of two major isoforms (pI 5.3-6.0) of similar molecular mass (21 kDa) was identified as RBP by using antiserum against RBP. Presence of RBP in conceptus culture medium and uterine flushings between days 16 and 22 of pregnancy were determined by immunoprecipitation and Western blotting using anti-RBP serum. In immunocytochemical study, strong immunostaining for RBP was localized in trophectoderm and endoderm of conceptus. These results clearly demonstrated that the caprine conceptus was active in protein synthesis as early as day 16 of pregnancy. Secretion of RBP by caprine conceptuses (days 16-22) coincident with the rapid transformation of the conceptus from a spherical blastocyst to a filamentous structure. Production of RBP by the elongating conceptuses may be indicative of an important role for conceptus RBP in the transport, availability and metabolism of retinol during maternal recognition of pregnancy.

• Asian-Australasian Journal of Animal Sciences
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• 제15권12호
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• pp.1708-1713
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• 2002

Endometrial explants obtained from does between days 13 and 21 of pregnancy were cultured in a modified minimum essential medium in the presence of [$^35S$]methionine and [$^3H$]-leucine. Proteins synthesized and secreted into medium were analyzed by fluorography of two-dimensional polyacrylamide gel electrophoresis and fluorography. No marked qualitative changes in patterns of protein production by caprine endometrium between days 13-21 of pregnancy. At least 11 proteins showed consistently a clear spot or a grouping of spots with characteristic location on two-dimensional gels. A major low molecular weight protein consisted of two major isoforms (pI 5.3-6.0) of similar molecular mass (21 kDa). Limited N-terminal sequence analysis of these two isoforms showed that the protein had complete homology with bovine placental and plasma retinol-binding protein (RBP) over the first 20 amino acids. Through use of the antiserum raised against bovine placental RBP, immunoreactive RBP was detected in cultures conditioned by uterine explants prepared at days 13, 15 and 21 of pregnancy. In the present study, proteins synthesized and secreted by caprine endometrium during periattachment period of early pregnancy were characterized. The pregnant endometrium secreted a number of neutral-to-acidic proteins which constituted, in part, the histotroph. A vitamin A-transport protein, RBP, was identified in cultures conditioned by endometrium of days 13-21 of pregnancy. The uterine endometrium is the only source of retinol for embryonic tissues. The uterine RBP appears to transport retinol locally toward embryonic tissues. Secretion of RBP by caprine endometrium of days 13, 15 and 21 of pregnancy suggested that retinol played an important role in conceptus development during periattachment period of early pregnancy.