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Effect of E-cadherin expression on hormone production in rat anterior pituitary lactotrophs in vitro.

Kusumoto K, Kikuchi M, Fujiwara K, Horiguchi K, Kouki T, Kawanishi K, Yashiro T - Acta Histochem Cytochem (2010)

Bottom Line: Recent studies of the developing rat adenohypophysis found that primordial cells co-expressed E- and N-cadherins, but that hormone-producing cells lost E-cadherin and ultimately possessed only N-cadherin.The mean signal intensity of prolactin protein in rE-cad-IZ-transfected cells was approximately one fourth that in intact cells and in -IZ-transfected cells (P<0.01).These results suggest that the expression of E-cadherin does not affect prolactin mRNA transcription; rather, it reduces prolactin protein content, presumably by affecting trafficking of secretory granules.

View Article: PubMed Central - PubMed

Affiliation: Division of Histology and Cell Biology, Department of Anatomy, Jichi Medical University School of Medicine, Yakushiji 3311-1, Shimotsuke-shi, Tochigi 329-0498.

ABSTRACT
Cadherins are a family of transmembrane glycoproteins that mediate cell-to-cell adhesion. A change in cadherin type in cells, i.e., cadherin switching, induces changes in the character of the cell. Recent studies of the developing rat adenohypophysis found that primordial cells co-expressed E- and N-cadherins, but that hormone-producing cells lost E-cadherin and ultimately possessed only N-cadherin. In the present study, we examined the roles of cadherin switching in cytogenesis of anterior pituitary cells by observing prolactin mRNA and protein expression in lactotrophs that were transformed with an E-cadherin expression vector. In hormone-producing cells that were transfected with a pIRES2-ZsGreen1 plasmid with a full-length E-cadherin cDNA (rE-cad-IZ) insert in primary culture, we detected E- and N-cadherins on plasma membrane and E-cadherin in cytoplasm. In these rE-cad-IZ-transfected cells, in situ hybridization revealed prolactin mRNA signals that were at a level identical to that in control cells, while prolactin protein was barely detectable using immunocytochemistry. The mean signal intensity of prolactin protein in rE-cad-IZ-transfected cells was approximately one fourth that in intact cells and in -IZ-transfected cells (P<0.01). These results suggest that the expression of E-cadherin does not affect prolactin mRNA transcription; rather, it reduces prolactin protein content, presumably by affecting trafficking of secretory granules.

No MeSH data available.


Related in: MedlinePlus

Immunohistochemistry of E-cadherin and N-cadherin in transformed anterior pituitary cells. Anterior pituitary cells were transfected with pIRES2-ZsGreen1 plasmid with a -insert, -IZ (A, C, E and G, in the identical field), and the same plasmid with cDNA for the entire region of rat E-cadherin, rE-cad-IZ (B, D, F and H, in the identical field). A and B: Differential interference contrast images of transformed cells. C and D: Merged images of immunocytochemistry for E-cadherin (light blue), N-cadherin (red), and fluorescence from ZsGreen1 (green). E and F: Immunoreaction of N-cadherin. G and H: Immunoreaction of E-cadherin. Immunocytochemistry for E-cadherin and a phase-contrast image of transfected hormone-producing cells sorted by fluorescence-activated cell sorting are also shown in I and J. I: Purified hormone-producing cells were transfected with -IZ. J: Purified hormone-producing cells were transfected with rE-cad-IZ. The differential interference contrast image is merged. The differential interference contrast image is merged. Arrows and arrowheads indicate cells transfected with rE-cad-IZ and -IZ, respectively. A small arrow shows the boundary of cells transfected with rE-cad-IZ. Bar=5 µm.
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Figure 1: Immunohistochemistry of E-cadherin and N-cadherin in transformed anterior pituitary cells. Anterior pituitary cells were transfected with pIRES2-ZsGreen1 plasmid with a -insert, -IZ (A, C, E and G, in the identical field), and the same plasmid with cDNA for the entire region of rat E-cadherin, rE-cad-IZ (B, D, F and H, in the identical field). A and B: Differential interference contrast images of transformed cells. C and D: Merged images of immunocytochemistry for E-cadherin (light blue), N-cadherin (red), and fluorescence from ZsGreen1 (green). E and F: Immunoreaction of N-cadherin. G and H: Immunoreaction of E-cadherin. Immunocytochemistry for E-cadherin and a phase-contrast image of transfected hormone-producing cells sorted by fluorescence-activated cell sorting are also shown in I and J. I: Purified hormone-producing cells were transfected with -IZ. J: Purified hormone-producing cells were transfected with rE-cad-IZ. The differential interference contrast image is merged. The differential interference contrast image is merged. Arrows and arrowheads indicate cells transfected with rE-cad-IZ and -IZ, respectively. A small arrow shows the boundary of cells transfected with rE-cad-IZ. Bar=5 µm.

Mentions: Transfected cells were identified with fluorescence derived from the ZsGreen1 plasmid, as shown in Figure 1 (indicated by arrows and arrowheads). The transfection rate was usually 2 to 3% and no difference in the rate was observed among cell types. The transfected cells did not differ in shape on differential interference contrast images (Fig. 1A and 1B). We detected N-cadherin on the plasma membrane (Fig. 1D and 1F) and E-cadherin both on the plasma membrane and in the cytoplasm of all cells transfected with the rE-cad-IZ plasmid (indicated by arrows in Fig. 1D, 1F, and 1H). In contrast, we detected only N-cadherin on the plasma membrane of cells transfected with the -IZ plasmid (indicated by arrowheads in Fig. 1C, 1E, and 1G). These results are not sufficient to confirm that E-cadherin was successfully induced in hormone-producing cells, because rE-cad-IZ transfection of non-hormone-producing folliculo-stellate cells, which express both E- and N-cadherin [8], may yield similar results. Thus, to confirm forced E-cadherin expression in hormone-producing cells, we transfected purified hormone-producing cells with rE-cad-IZ. Hormone-producing cells were purified from transgenic S100b-GFP rat anterior pituitary cells [6] by means of fluorescence-activated cell sorting. Transfected hormone-producing cells did not differ in cell shape on differential interference contrast images (Fig. 1I and 1J). E-cadherin was detected on the plasma membrane of rE-cad-IZ-transfected cells (indicated by the small arrow in Fig. 1J) and in the cytoplasm of hormone-producing cells. However, we did not detect E-cadherin in -IZ-transfected hormone-producing cells (Fig. 1I).


Effect of E-cadherin expression on hormone production in rat anterior pituitary lactotrophs in vitro.

Kusumoto K, Kikuchi M, Fujiwara K, Horiguchi K, Kouki T, Kawanishi K, Yashiro T - Acta Histochem Cytochem (2010)

Immunohistochemistry of E-cadherin and N-cadherin in transformed anterior pituitary cells. Anterior pituitary cells were transfected with pIRES2-ZsGreen1 plasmid with a -insert, -IZ (A, C, E and G, in the identical field), and the same plasmid with cDNA for the entire region of rat E-cadherin, rE-cad-IZ (B, D, F and H, in the identical field). A and B: Differential interference contrast images of transformed cells. C and D: Merged images of immunocytochemistry for E-cadherin (light blue), N-cadherin (red), and fluorescence from ZsGreen1 (green). E and F: Immunoreaction of N-cadherin. G and H: Immunoreaction of E-cadherin. Immunocytochemistry for E-cadherin and a phase-contrast image of transfected hormone-producing cells sorted by fluorescence-activated cell sorting are also shown in I and J. I: Purified hormone-producing cells were transfected with -IZ. J: Purified hormone-producing cells were transfected with rE-cad-IZ. The differential interference contrast image is merged. The differential interference contrast image is merged. Arrows and arrowheads indicate cells transfected with rE-cad-IZ and -IZ, respectively. A small arrow shows the boundary of cells transfected with rE-cad-IZ. Bar=5 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 1: Immunohistochemistry of E-cadherin and N-cadherin in transformed anterior pituitary cells. Anterior pituitary cells were transfected with pIRES2-ZsGreen1 plasmid with a -insert, -IZ (A, C, E and G, in the identical field), and the same plasmid with cDNA for the entire region of rat E-cadherin, rE-cad-IZ (B, D, F and H, in the identical field). A and B: Differential interference contrast images of transformed cells. C and D: Merged images of immunocytochemistry for E-cadherin (light blue), N-cadherin (red), and fluorescence from ZsGreen1 (green). E and F: Immunoreaction of N-cadherin. G and H: Immunoreaction of E-cadherin. Immunocytochemistry for E-cadherin and a phase-contrast image of transfected hormone-producing cells sorted by fluorescence-activated cell sorting are also shown in I and J. I: Purified hormone-producing cells were transfected with -IZ. J: Purified hormone-producing cells were transfected with rE-cad-IZ. The differential interference contrast image is merged. The differential interference contrast image is merged. Arrows and arrowheads indicate cells transfected with rE-cad-IZ and -IZ, respectively. A small arrow shows the boundary of cells transfected with rE-cad-IZ. Bar=5 µm.
Mentions: Transfected cells were identified with fluorescence derived from the ZsGreen1 plasmid, as shown in Figure 1 (indicated by arrows and arrowheads). The transfection rate was usually 2 to 3% and no difference in the rate was observed among cell types. The transfected cells did not differ in shape on differential interference contrast images (Fig. 1A and 1B). We detected N-cadherin on the plasma membrane (Fig. 1D and 1F) and E-cadherin both on the plasma membrane and in the cytoplasm of all cells transfected with the rE-cad-IZ plasmid (indicated by arrows in Fig. 1D, 1F, and 1H). In contrast, we detected only N-cadherin on the plasma membrane of cells transfected with the -IZ plasmid (indicated by arrowheads in Fig. 1C, 1E, and 1G). These results are not sufficient to confirm that E-cadherin was successfully induced in hormone-producing cells, because rE-cad-IZ transfection of non-hormone-producing folliculo-stellate cells, which express both E- and N-cadherin [8], may yield similar results. Thus, to confirm forced E-cadherin expression in hormone-producing cells, we transfected purified hormone-producing cells with rE-cad-IZ. Hormone-producing cells were purified from transgenic S100b-GFP rat anterior pituitary cells [6] by means of fluorescence-activated cell sorting. Transfected hormone-producing cells did not differ in cell shape on differential interference contrast images (Fig. 1I and 1J). E-cadherin was detected on the plasma membrane of rE-cad-IZ-transfected cells (indicated by the small arrow in Fig. 1J) and in the cytoplasm of hormone-producing cells. However, we did not detect E-cadherin in -IZ-transfected hormone-producing cells (Fig. 1I).

Bottom Line: Recent studies of the developing rat adenohypophysis found that primordial cells co-expressed E- and N-cadherins, but that hormone-producing cells lost E-cadherin and ultimately possessed only N-cadherin.The mean signal intensity of prolactin protein in rE-cad-IZ-transfected cells was approximately one fourth that in intact cells and in -IZ-transfected cells (P<0.01).These results suggest that the expression of E-cadherin does not affect prolactin mRNA transcription; rather, it reduces prolactin protein content, presumably by affecting trafficking of secretory granules.

View Article: PubMed Central - PubMed

Affiliation: Division of Histology and Cell Biology, Department of Anatomy, Jichi Medical University School of Medicine, Yakushiji 3311-1, Shimotsuke-shi, Tochigi 329-0498.

ABSTRACT
Cadherins are a family of transmembrane glycoproteins that mediate cell-to-cell adhesion. A change in cadherin type in cells, i.e., cadherin switching, induces changes in the character of the cell. Recent studies of the developing rat adenohypophysis found that primordial cells co-expressed E- and N-cadherins, but that hormone-producing cells lost E-cadherin and ultimately possessed only N-cadherin. In the present study, we examined the roles of cadherin switching in cytogenesis of anterior pituitary cells by observing prolactin mRNA and protein expression in lactotrophs that were transformed with an E-cadherin expression vector. In hormone-producing cells that were transfected with a pIRES2-ZsGreen1 plasmid with a full-length E-cadherin cDNA (rE-cad-IZ) insert in primary culture, we detected E- and N-cadherins on plasma membrane and E-cadherin in cytoplasm. In these rE-cad-IZ-transfected cells, in situ hybridization revealed prolactin mRNA signals that were at a level identical to that in control cells, while prolactin protein was barely detectable using immunocytochemistry. The mean signal intensity of prolactin protein in rE-cad-IZ-transfected cells was approximately one fourth that in intact cells and in -IZ-transfected cells (P<0.01). These results suggest that the expression of E-cadherin does not affect prolactin mRNA transcription; rather, it reduces prolactin protein content, presumably by affecting trafficking of secretory granules.

No MeSH data available.


Related in: MedlinePlus