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Elastogenic protein expression of a highly elastic murine spinal ligament: the ligamentum flavum.

Brown JP, Lind RM, Burzesi AF, Kuo CK - PLoS ONE (2012)

Bottom Line: We found that elastogenesis begins in utero with the microfibril constituent fibrillin-1 staining intensely just before birth.These expression patterns correlated with reported skeletal and behavioral changes during murine development.This immunohistochemical characterization of elastogenesis of the LF will be useful for future studies investigating mechanisms for elastogenesis and developing new strategies for treatment or regeneration of spinal ligaments and other highly elastic tissues.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America.

ABSTRACT
Spinal ligaments, such as the ligamentum flavum (LF), are prone to degeneration and iatrogenic injury that can lead to back pain and nerve dysfunction. Repair and regeneration strategies for these tissues are lacking, perhaps due to limited understanding of spinal ligament formation, the elaboration of its elastic fibers, maturation and homeostasis. Using immunohistochemistry and histology, we investigated murine LF elastogenesis and tissue formation from embryonic to mature postnatal stages. We characterized the spatiotemporal distribution of the key elastogenic proteins tropoelastin, fibrillin-1, fibulin-4 and lysyl oxidase. We found that elastogenesis begins in utero with the microfibril constituent fibrillin-1 staining intensely just before birth. Elastic fibers were first detected histologically at postnatal day (P) 7, the earliest stage at which tropoelastin and fibulin-4 stained intensely. From P7 to P28, elastic fibers grew in diameter and became straighter along the axis. The growth of elastic fibers coincided with intense staining of tropoelastin and fibulin-4 staining, possibly supporting a chaperone role for fibulin-4. These expression patterns correlated with reported skeletal and behavioral changes during murine development. This immunohistochemical characterization of elastogenesis of the LF will be useful for future studies investigating mechanisms for elastogenesis and developing new strategies for treatment or regeneration of spinal ligaments and other highly elastic tissues.

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Related in: MedlinePlus

Determining cellularity of LF.Three representative 1000 µm2 areas of the LF were selected for cell counting at approximately equidistant regions along the length of the ligament. Counts were averaged for mice at a given stage for comparison (P56 shown). Scale bar: 200 µm.
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pone-0038475-g001: Determining cellularity of LF.Three representative 1000 µm2 areas of the LF were selected for cell counting at approximately equidistant regions along the length of the ligament. Counts were averaged for mice at a given stage for comparison (P56 shown). Scale bar: 200 µm.

Mentions: H&E-stained sections of the LF from E16 through P2-yrs were imaged using an inverted optical microscope (Leitz Diavert, Wetzlar, Germany) and a DXC-390P color video camera (Sony, Tokyo, Japan). Three representative areas of the LF were selected for analysis at approximately three equidistant regions along the length of the ligament (Fig. 1). Stained nuclei in selected areas of a given LF were counted manually using ImageJ software (NIH, Bethesda, MD) and averaged for the animal. LF from ≥3 different animals were characterized for each stage. Means and standard deviations were calculated for all animals of a given stage or age.


Elastogenic protein expression of a highly elastic murine spinal ligament: the ligamentum flavum.

Brown JP, Lind RM, Burzesi AF, Kuo CK - PLoS ONE (2012)

Determining cellularity of LF.Three representative 1000 µm2 areas of the LF were selected for cell counting at approximately equidistant regions along the length of the ligament. Counts were averaged for mice at a given stage for comparison (P56 shown). Scale bar: 200 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3369910&req=5

pone-0038475-g001: Determining cellularity of LF.Three representative 1000 µm2 areas of the LF were selected for cell counting at approximately equidistant regions along the length of the ligament. Counts were averaged for mice at a given stage for comparison (P56 shown). Scale bar: 200 µm.
Mentions: H&E-stained sections of the LF from E16 through P2-yrs were imaged using an inverted optical microscope (Leitz Diavert, Wetzlar, Germany) and a DXC-390P color video camera (Sony, Tokyo, Japan). Three representative areas of the LF were selected for analysis at approximately three equidistant regions along the length of the ligament (Fig. 1). Stained nuclei in selected areas of a given LF were counted manually using ImageJ software (NIH, Bethesda, MD) and averaged for the animal. LF from ≥3 different animals were characterized for each stage. Means and standard deviations were calculated for all animals of a given stage or age.

Bottom Line: We found that elastogenesis begins in utero with the microfibril constituent fibrillin-1 staining intensely just before birth.These expression patterns correlated with reported skeletal and behavioral changes during murine development.This immunohistochemical characterization of elastogenesis of the LF will be useful for future studies investigating mechanisms for elastogenesis and developing new strategies for treatment or regeneration of spinal ligaments and other highly elastic tissues.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America.

ABSTRACT
Spinal ligaments, such as the ligamentum flavum (LF), are prone to degeneration and iatrogenic injury that can lead to back pain and nerve dysfunction. Repair and regeneration strategies for these tissues are lacking, perhaps due to limited understanding of spinal ligament formation, the elaboration of its elastic fibers, maturation and homeostasis. Using immunohistochemistry and histology, we investigated murine LF elastogenesis and tissue formation from embryonic to mature postnatal stages. We characterized the spatiotemporal distribution of the key elastogenic proteins tropoelastin, fibrillin-1, fibulin-4 and lysyl oxidase. We found that elastogenesis begins in utero with the microfibril constituent fibrillin-1 staining intensely just before birth. Elastic fibers were first detected histologically at postnatal day (P) 7, the earliest stage at which tropoelastin and fibulin-4 stained intensely. From P7 to P28, elastic fibers grew in diameter and became straighter along the axis. The growth of elastic fibers coincided with intense staining of tropoelastin and fibulin-4 staining, possibly supporting a chaperone role for fibulin-4. These expression patterns correlated with reported skeletal and behavioral changes during murine development. This immunohistochemical characterization of elastogenesis of the LF will be useful for future studies investigating mechanisms for elastogenesis and developing new strategies for treatment or regeneration of spinal ligaments and other highly elastic tissues.

Show MeSH
Related in: MedlinePlus