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Apolipoprotein C-II and lipoprotein lipase show a temporal and geographic correlation with surfactant lipid synthesis in preparation for birth.

Côté M, Provost PR, Gérard-Hudon MC, Tremblay Y - BMC Dev. Biol. (2010)

Bottom Line: The major sites of apoC-II and LPL gene expression changed over time and were found mainly in the distal epithelium at the end of gestation but not after birth.Accumulation of apoC-II in secretory granule-like structures was not systematically observed, but was found in the distal epithelium only at the end of gestation and soon after birth, mainly in epithelia with no or small lumina.We propose a model where apoC-II is retained in secretory granules in distal epithelial cells until the lumina reaches a minimum size, and is then secreted when the rate of surfactant production becomes optimal.

View Article: PubMed Central - HTML - PubMed

Affiliation: Reproduction, Perinatal and Child Health Axis, Rm T-1-49, CHUQ Research Center, Centre de Recherche en Biologie de Reproduction, Laval University, Québec City, Québec, Canada.

ABSTRACT

Background: Fatty acids are precursors in the synthesis of surfactant phospholipids. Recently, we showed expression of apolipoprotein C-II (apoC-II), the essential cofactor of lipoprotein lipase (LPL), in the fetal mouse lung and found the protein on the day of the surge of surfactant synthesis (gestation day 17.5) in secretory granule-like structures in the distal epithelium. In the present study, we will answer the following questions: Does apoC-II protein localization change according to the stage of lung development, thus according to the need in surfactant? Are LPL molecules translocated to the luminal surface of capillaries? Do the sites of apoC-II and LPL gene expression change according to the stage of lung development and to protein localization?

Results: The present study investigated whether the sites of apoC-II and LPL mRNA and protein accumulation are regulated in the mouse lung between gestation day 15 and postnatal day 10. The major sites of apoC-II and LPL gene expression changed over time and were found mainly in the distal epithelium at the end of gestation but not after birth. Accumulation of apoC-II in secretory granule-like structures was not systematically observed, but was found in the distal epithelium only at the end of gestation and soon after birth, mainly in epithelia with no or small lumina. A noticeable increase in surfactant lipid content was measured before the end of gestation day 18, which correlates temporally with the presence of apoC-II in secretory granules in distal epithelium with no or small lumina but not with large lumina. LPL was detected in capillaries at all the developmental times studied.

Conclusions: This study demonstrates that apoC-II and LPL mRNAs correlate temporally and geographically with surfactant lipid synthesis in preparation for birth and suggests that fatty acid recruitment from the circulation by apoC-II-activated LPL is regionally modulated by apoC-II secretion. We propose a model where apoC-II is retained in secretory granules in distal epithelial cells until the lumina reaches a minimum size, and is then secreted when the rate of surfactant production becomes optimal.

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Distribution of apolipoprotein C-II mRNA and protein in the perinatal mouse lung. Mouse tissue sections are from saccular stage (A, B, H, GD 19.5; C, G, PN 0; I, J, PN 1; K, PN 2; L, PN 3; D to F, M, N, PN 5) or alveolar stage (O, PN 10). In situ hybridization (above the yellow line) (A to F) was performed with apoC-II anti-sense (A, C to E) and sense (B, F) probes. The major site of apoC-II mRNA synthesis (positive signal, blue) changed after birth (compare A to C). Positive signals were found in newly-formed septa (D, E) and macrophages (E) on PN 5. Immunohistochemistry (below the yellow line) (G-O) was performed using an anti-apoC-II polyclonal antibody (G to I, K to M, O) or goat IgG as negative control (J, N). Positive signals (red) were found in secretory granule-like structures in distal epithelial cells on GD 19.5 (H) and PN0 (G) but not in later timepoints. On PN 3, positive secretory granules were also found in epithelial cells of the respiratory bronchioles near the basal membrane, close to the mesenchyme (L). Macrophages were positive on PN 5 and 10 while capillaries were positive on PN 10 in one third of the analyzed subjects (O and data not shown). Scale bars, 50 μm (A to D, F, H, K, L, M, N) or 20 μm (E, G, I, J, L, O). BV, blood vessel; C, capillary; D, distal epithelium; MA, macrophage; P, proximal epithelium; RB, respiratory bronchiole; S, septa; V, vein.
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Figure 5: Distribution of apolipoprotein C-II mRNA and protein in the perinatal mouse lung. Mouse tissue sections are from saccular stage (A, B, H, GD 19.5; C, G, PN 0; I, J, PN 1; K, PN 2; L, PN 3; D to F, M, N, PN 5) or alveolar stage (O, PN 10). In situ hybridization (above the yellow line) (A to F) was performed with apoC-II anti-sense (A, C to E) and sense (B, F) probes. The major site of apoC-II mRNA synthesis (positive signal, blue) changed after birth (compare A to C). Positive signals were found in newly-formed septa (D, E) and macrophages (E) on PN 5. Immunohistochemistry (below the yellow line) (G-O) was performed using an anti-apoC-II polyclonal antibody (G to I, K to M, O) or goat IgG as negative control (J, N). Positive signals (red) were found in secretory granule-like structures in distal epithelial cells on GD 19.5 (H) and PN0 (G) but not in later timepoints. On PN 3, positive secretory granules were also found in epithelial cells of the respiratory bronchioles near the basal membrane, close to the mesenchyme (L). Macrophages were positive on PN 5 and 10 while capillaries were positive on PN 10 in one third of the analyzed subjects (O and data not shown). Scale bars, 50 μm (A to D, F, H, K, L, M, N) or 20 μm (E, G, I, J, L, O). BV, blood vessel; C, capillary; D, distal epithelium; MA, macrophage; P, proximal epithelium; RB, respiratory bronchiole; S, septa; V, vein.

Mentions: Similarities were found between apoC-II and LPL expression patterns during the perinatal period. Both genes were expressed in the distal epithelium short time before birth (Figures 5A and 6A), but not after birth (Figures 5C and 6C) (delivery occurred during GD 19). In contrast, the proximal epithelium was negative on GD 17.5, showed a weak positive signal on GD 18.5 and 19.5 (data not shown), presented a marked increase in intensity soon after birth (Figures 5C and 6C), and was negative on PN 5 for both genes. Using 6 fetuses on GD 19.5 (3 males and 3 females) and 6 neonates on PN 0, we confirmed that the switch from the distal to the proximal epithelium correlated with birth, not with sex (data not shown). ApoC-II and LPL mRNAs were also found in a few scattered distal epithelial cells after birth. In addition, positive signals by ISH were observed in association with blood vessels before birth (Figures 5A and 6A) but not after birth (data not shown) for both genes. On PN 5, thus at junction between saccularization and alveolarization, apoC-II and LPL mRNA were found in alveolar walls (Figures 5D-E and 6E-F), including newly-formed septa (Figure 5E and data not shown).


Apolipoprotein C-II and lipoprotein lipase show a temporal and geographic correlation with surfactant lipid synthesis in preparation for birth.

Côté M, Provost PR, Gérard-Hudon MC, Tremblay Y - BMC Dev. Biol. (2010)

Distribution of apolipoprotein C-II mRNA and protein in the perinatal mouse lung. Mouse tissue sections are from saccular stage (A, B, H, GD 19.5; C, G, PN 0; I, J, PN 1; K, PN 2; L, PN 3; D to F, M, N, PN 5) or alveolar stage (O, PN 10). In situ hybridization (above the yellow line) (A to F) was performed with apoC-II anti-sense (A, C to E) and sense (B, F) probes. The major site of apoC-II mRNA synthesis (positive signal, blue) changed after birth (compare A to C). Positive signals were found in newly-formed septa (D, E) and macrophages (E) on PN 5. Immunohistochemistry (below the yellow line) (G-O) was performed using an anti-apoC-II polyclonal antibody (G to I, K to M, O) or goat IgG as negative control (J, N). Positive signals (red) were found in secretory granule-like structures in distal epithelial cells on GD 19.5 (H) and PN0 (G) but not in later timepoints. On PN 3, positive secretory granules were also found in epithelial cells of the respiratory bronchioles near the basal membrane, close to the mesenchyme (L). Macrophages were positive on PN 5 and 10 while capillaries were positive on PN 10 in one third of the analyzed subjects (O and data not shown). Scale bars, 50 μm (A to D, F, H, K, L, M, N) or 20 μm (E, G, I, J, L, O). BV, blood vessel; C, capillary; D, distal epithelium; MA, macrophage; P, proximal epithelium; RB, respiratory bronchiole; S, septa; V, vein.
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Figure 5: Distribution of apolipoprotein C-II mRNA and protein in the perinatal mouse lung. Mouse tissue sections are from saccular stage (A, B, H, GD 19.5; C, G, PN 0; I, J, PN 1; K, PN 2; L, PN 3; D to F, M, N, PN 5) or alveolar stage (O, PN 10). In situ hybridization (above the yellow line) (A to F) was performed with apoC-II anti-sense (A, C to E) and sense (B, F) probes. The major site of apoC-II mRNA synthesis (positive signal, blue) changed after birth (compare A to C). Positive signals were found in newly-formed septa (D, E) and macrophages (E) on PN 5. Immunohistochemistry (below the yellow line) (G-O) was performed using an anti-apoC-II polyclonal antibody (G to I, K to M, O) or goat IgG as negative control (J, N). Positive signals (red) were found in secretory granule-like structures in distal epithelial cells on GD 19.5 (H) and PN0 (G) but not in later timepoints. On PN 3, positive secretory granules were also found in epithelial cells of the respiratory bronchioles near the basal membrane, close to the mesenchyme (L). Macrophages were positive on PN 5 and 10 while capillaries were positive on PN 10 in one third of the analyzed subjects (O and data not shown). Scale bars, 50 μm (A to D, F, H, K, L, M, N) or 20 μm (E, G, I, J, L, O). BV, blood vessel; C, capillary; D, distal epithelium; MA, macrophage; P, proximal epithelium; RB, respiratory bronchiole; S, septa; V, vein.
Mentions: Similarities were found between apoC-II and LPL expression patterns during the perinatal period. Both genes were expressed in the distal epithelium short time before birth (Figures 5A and 6A), but not after birth (Figures 5C and 6C) (delivery occurred during GD 19). In contrast, the proximal epithelium was negative on GD 17.5, showed a weak positive signal on GD 18.5 and 19.5 (data not shown), presented a marked increase in intensity soon after birth (Figures 5C and 6C), and was negative on PN 5 for both genes. Using 6 fetuses on GD 19.5 (3 males and 3 females) and 6 neonates on PN 0, we confirmed that the switch from the distal to the proximal epithelium correlated with birth, not with sex (data not shown). ApoC-II and LPL mRNAs were also found in a few scattered distal epithelial cells after birth. In addition, positive signals by ISH were observed in association with blood vessels before birth (Figures 5A and 6A) but not after birth (data not shown) for both genes. On PN 5, thus at junction between saccularization and alveolarization, apoC-II and LPL mRNA were found in alveolar walls (Figures 5D-E and 6E-F), including newly-formed septa (Figure 5E and data not shown).

Bottom Line: The major sites of apoC-II and LPL gene expression changed over time and were found mainly in the distal epithelium at the end of gestation but not after birth.Accumulation of apoC-II in secretory granule-like structures was not systematically observed, but was found in the distal epithelium only at the end of gestation and soon after birth, mainly in epithelia with no or small lumina.We propose a model where apoC-II is retained in secretory granules in distal epithelial cells until the lumina reaches a minimum size, and is then secreted when the rate of surfactant production becomes optimal.

View Article: PubMed Central - HTML - PubMed

Affiliation: Reproduction, Perinatal and Child Health Axis, Rm T-1-49, CHUQ Research Center, Centre de Recherche en Biologie de Reproduction, Laval University, Québec City, Québec, Canada.

ABSTRACT

Background: Fatty acids are precursors in the synthesis of surfactant phospholipids. Recently, we showed expression of apolipoprotein C-II (apoC-II), the essential cofactor of lipoprotein lipase (LPL), in the fetal mouse lung and found the protein on the day of the surge of surfactant synthesis (gestation day 17.5) in secretory granule-like structures in the distal epithelium. In the present study, we will answer the following questions: Does apoC-II protein localization change according to the stage of lung development, thus according to the need in surfactant? Are LPL molecules translocated to the luminal surface of capillaries? Do the sites of apoC-II and LPL gene expression change according to the stage of lung development and to protein localization?

Results: The present study investigated whether the sites of apoC-II and LPL mRNA and protein accumulation are regulated in the mouse lung between gestation day 15 and postnatal day 10. The major sites of apoC-II and LPL gene expression changed over time and were found mainly in the distal epithelium at the end of gestation but not after birth. Accumulation of apoC-II in secretory granule-like structures was not systematically observed, but was found in the distal epithelium only at the end of gestation and soon after birth, mainly in epithelia with no or small lumina. A noticeable increase in surfactant lipid content was measured before the end of gestation day 18, which correlates temporally with the presence of apoC-II in secretory granules in distal epithelium with no or small lumina but not with large lumina. LPL was detected in capillaries at all the developmental times studied.

Conclusions: This study demonstrates that apoC-II and LPL mRNAs correlate temporally and geographically with surfactant lipid synthesis in preparation for birth and suggests that fatty acid recruitment from the circulation by apoC-II-activated LPL is regionally modulated by apoC-II secretion. We propose a model where apoC-II is retained in secretory granules in distal epithelial cells until the lumina reaches a minimum size, and is then secreted when the rate of surfactant production becomes optimal.

Show MeSH