Limits...
A central function for perlecan in skeletal muscle and cardiovascular development.

Zoeller JJ, McQuillan A, Whitelock J, Ho SY, Iozzo RV - J. Cell Biol. (2008)

Bottom Line: In the perlecan morphants, primary intersegmental vessel sprouts, which develop through angiogenesis, fail to extend and show reduced protrusive activity.The phenotype is partially rescued by microinjection of human perlecan or endorepellin.These findings indicate that perlecan is essential for the integrity of somitic muscle and developmental angiogenesis and that endorepellin mediates most of these biological activities.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Anatomy, and Cell, Thomas Jefferson University, Philadelphia, PA 19107, USA.

ABSTRACT
Perlecan's developmental functions are difficult to dissect in placental animals because perlecan disruption is embryonic lethal. In contrast to mammals, cardiovascular function is not essential for early zebrafish development because the embryos obtain adequate oxygen by diffusion. In this study, we use targeted protein depletion coupled with protein-based rescue experiments to investigate the involvement of perlecan and its C-terminal domain V/endorepellin in zebrafish development. The perlecan morphants show a severe myopathy characterized by abnormal actin filament orientation and disorganized sarcomeres, suggesting an involvement of perlecan in myopathies. In the perlecan morphants, primary intersegmental vessel sprouts, which develop through angiogenesis, fail to extend and show reduced protrusive activity. Live videomicroscopy confirms the abnormal swimming pattern caused by the myopathy and anomalous head and trunk vessel circulation. The phenotype is partially rescued by microinjection of human perlecan or endorepellin. These findings indicate that perlecan is essential for the integrity of somitic muscle and developmental angiogenesis and that endorepellin mediates most of these biological activities.

Show MeSH

Related in: MedlinePlus

Spatiotemporal expression patterns of zebrafish perlecan mRNA and protein. (A–G) Whole mount ISH with a digoxigenin-labeled perlecan domain V antisense probe for the localization of perlecan mRNA (blue/purple staining). Perlecan mRNA can be detected at the 20-somite stage in regions of the developing brain (A) and along both sides of the notochord (B, arrows). At 1 dpf, perlecan expression is seen in the head region (C), in the developing somites (C, black arrows), and within the developing axial vasculature (D, arrows). Also note perlecan expression localized within the duct of Cuvier (C, red arrows). By 5 dpf (F and G [magnified image of the region boxed in F]), perlecan expression significantly increases throughout the trunk musculature (G, arrows), the major trunk/tail vessels including the DLAV, DA, and PCV, and in the developing gastrointestinal tract and future fin regions. (E) ISH with a digoxigenin-labeled perlecan domain V sense probe. ISH was performed in groups of two to five samples, and representative images are shown. (H–N) Whole mount immunohistochemistry with an affinity-purified anti–mouse perlecan antibody. Perlecan can be detected as early as the 64- and 1,000-cell stage of embryonic development (2 and 3 hpf, respectively) throughout the cell mound (I and J). At 1 dpf, perlecan protein (L and M) is detected throughout the head (L, arrow), trunk, and tail and is specifically localized to the developing muscle myoseptae (M, arrows) and the developing vasculature. By 2 dpf (N), perlecan protein is specifically detected in the trunk vasculature, including the DLAV (N, black arrows) and axial vessels (N, white arrows). (H and K) Immunohistochemistry images in which the primary antibody was omitted. Bars, 500 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2315682&req=5

fig2: Spatiotemporal expression patterns of zebrafish perlecan mRNA and protein. (A–G) Whole mount ISH with a digoxigenin-labeled perlecan domain V antisense probe for the localization of perlecan mRNA (blue/purple staining). Perlecan mRNA can be detected at the 20-somite stage in regions of the developing brain (A) and along both sides of the notochord (B, arrows). At 1 dpf, perlecan expression is seen in the head region (C), in the developing somites (C, black arrows), and within the developing axial vasculature (D, arrows). Also note perlecan expression localized within the duct of Cuvier (C, red arrows). By 5 dpf (F and G [magnified image of the region boxed in F]), perlecan expression significantly increases throughout the trunk musculature (G, arrows), the major trunk/tail vessels including the DLAV, DA, and PCV, and in the developing gastrointestinal tract and future fin regions. (E) ISH with a digoxigenin-labeled perlecan domain V sense probe. ISH was performed in groups of two to five samples, and representative images are shown. (H–N) Whole mount immunohistochemistry with an affinity-purified anti–mouse perlecan antibody. Perlecan can be detected as early as the 64- and 1,000-cell stage of embryonic development (2 and 3 hpf, respectively) throughout the cell mound (I and J). At 1 dpf, perlecan protein (L and M) is detected throughout the head (L, arrow), trunk, and tail and is specifically localized to the developing muscle myoseptae (M, arrows) and the developing vasculature. By 2 dpf (N), perlecan protein is specifically detected in the trunk vasculature, including the DLAV (N, black arrows) and axial vessels (N, white arrows). (H and K) Immunohistochemistry images in which the primary antibody was omitted. Bars, 500 μm.

Mentions: To determine the spatiotemporal expression of zebrafish perlecan, we used whole mount in situ hybridization (ISH) and immunohistochemistry. ISH using a domain V antisense RNA probe showed the prominent expression of perlecan at the 20-somite stage in the head and somite region (Fig. 2, A and B).Perlecan was more clearly expressed at 1 dpf in these regions and was localized along the lateral myotomes (Fig. 2 C) and in the major axial vessels (Fig. 2 D). At 5 dpf, perlecan was expressed in the head and intestinal organs, with strong positivity in the axial vessels, including the dorsal aorta (DA) and the dorsal longitudinal anastomotic vessel (DLAV) as well as in the myoseptae, the ISVs, and posterior cardinal vein (PCV; Fig. 2, F and G).


A central function for perlecan in skeletal muscle and cardiovascular development.

Zoeller JJ, McQuillan A, Whitelock J, Ho SY, Iozzo RV - J. Cell Biol. (2008)

Spatiotemporal expression patterns of zebrafish perlecan mRNA and protein. (A–G) Whole mount ISH with a digoxigenin-labeled perlecan domain V antisense probe for the localization of perlecan mRNA (blue/purple staining). Perlecan mRNA can be detected at the 20-somite stage in regions of the developing brain (A) and along both sides of the notochord (B, arrows). At 1 dpf, perlecan expression is seen in the head region (C), in the developing somites (C, black arrows), and within the developing axial vasculature (D, arrows). Also note perlecan expression localized within the duct of Cuvier (C, red arrows). By 5 dpf (F and G [magnified image of the region boxed in F]), perlecan expression significantly increases throughout the trunk musculature (G, arrows), the major trunk/tail vessels including the DLAV, DA, and PCV, and in the developing gastrointestinal tract and future fin regions. (E) ISH with a digoxigenin-labeled perlecan domain V sense probe. ISH was performed in groups of two to five samples, and representative images are shown. (H–N) Whole mount immunohistochemistry with an affinity-purified anti–mouse perlecan antibody. Perlecan can be detected as early as the 64- and 1,000-cell stage of embryonic development (2 and 3 hpf, respectively) throughout the cell mound (I and J). At 1 dpf, perlecan protein (L and M) is detected throughout the head (L, arrow), trunk, and tail and is specifically localized to the developing muscle myoseptae (M, arrows) and the developing vasculature. By 2 dpf (N), perlecan protein is specifically detected in the trunk vasculature, including the DLAV (N, black arrows) and axial vessels (N, white arrows). (H and K) Immunohistochemistry images in which the primary antibody was omitted. Bars, 500 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2315682&req=5

fig2: Spatiotemporal expression patterns of zebrafish perlecan mRNA and protein. (A–G) Whole mount ISH with a digoxigenin-labeled perlecan domain V antisense probe for the localization of perlecan mRNA (blue/purple staining). Perlecan mRNA can be detected at the 20-somite stage in regions of the developing brain (A) and along both sides of the notochord (B, arrows). At 1 dpf, perlecan expression is seen in the head region (C), in the developing somites (C, black arrows), and within the developing axial vasculature (D, arrows). Also note perlecan expression localized within the duct of Cuvier (C, red arrows). By 5 dpf (F and G [magnified image of the region boxed in F]), perlecan expression significantly increases throughout the trunk musculature (G, arrows), the major trunk/tail vessels including the DLAV, DA, and PCV, and in the developing gastrointestinal tract and future fin regions. (E) ISH with a digoxigenin-labeled perlecan domain V sense probe. ISH was performed in groups of two to five samples, and representative images are shown. (H–N) Whole mount immunohistochemistry with an affinity-purified anti–mouse perlecan antibody. Perlecan can be detected as early as the 64- and 1,000-cell stage of embryonic development (2 and 3 hpf, respectively) throughout the cell mound (I and J). At 1 dpf, perlecan protein (L and M) is detected throughout the head (L, arrow), trunk, and tail and is specifically localized to the developing muscle myoseptae (M, arrows) and the developing vasculature. By 2 dpf (N), perlecan protein is specifically detected in the trunk vasculature, including the DLAV (N, black arrows) and axial vessels (N, white arrows). (H and K) Immunohistochemistry images in which the primary antibody was omitted. Bars, 500 μm.
Mentions: To determine the spatiotemporal expression of zebrafish perlecan, we used whole mount in situ hybridization (ISH) and immunohistochemistry. ISH using a domain V antisense RNA probe showed the prominent expression of perlecan at the 20-somite stage in the head and somite region (Fig. 2, A and B).Perlecan was more clearly expressed at 1 dpf in these regions and was localized along the lateral myotomes (Fig. 2 C) and in the major axial vessels (Fig. 2 D). At 5 dpf, perlecan was expressed in the head and intestinal organs, with strong positivity in the axial vessels, including the dorsal aorta (DA) and the dorsal longitudinal anastomotic vessel (DLAV) as well as in the myoseptae, the ISVs, and posterior cardinal vein (PCV; Fig. 2, F and G).

Bottom Line: In the perlecan morphants, primary intersegmental vessel sprouts, which develop through angiogenesis, fail to extend and show reduced protrusive activity.The phenotype is partially rescued by microinjection of human perlecan or endorepellin.These findings indicate that perlecan is essential for the integrity of somitic muscle and developmental angiogenesis and that endorepellin mediates most of these biological activities.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Anatomy, and Cell, Thomas Jefferson University, Philadelphia, PA 19107, USA.

ABSTRACT
Perlecan's developmental functions are difficult to dissect in placental animals because perlecan disruption is embryonic lethal. In contrast to mammals, cardiovascular function is not essential for early zebrafish development because the embryos obtain adequate oxygen by diffusion. In this study, we use targeted protein depletion coupled with protein-based rescue experiments to investigate the involvement of perlecan and its C-terminal domain V/endorepellin in zebrafish development. The perlecan morphants show a severe myopathy characterized by abnormal actin filament orientation and disorganized sarcomeres, suggesting an involvement of perlecan in myopathies. In the perlecan morphants, primary intersegmental vessel sprouts, which develop through angiogenesis, fail to extend and show reduced protrusive activity. Live videomicroscopy confirms the abnormal swimming pattern caused by the myopathy and anomalous head and trunk vessel circulation. The phenotype is partially rescued by microinjection of human perlecan or endorepellin. These findings indicate that perlecan is essential for the integrity of somitic muscle and developmental angiogenesis and that endorepellin mediates most of these biological activities.

Show MeSH
Related in: MedlinePlus