Horizontal deformation of skeletal muscle thick sections visualised by confocal microscopy
Certain morphological parameters of the skeletal muscle tissue can be better understood via 3D considerations. Fluorescent confocal microscopy of thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, thick...
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description | Certain morphological parameters of the skeletal muscle tissue can be better understood via 3D considerations. Fluorescent confocal microscopy of thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, thick tissue sections are prone to deformations which may significantly influence some stereological and morphometric results like muscle fibre diameter and capillary length, but not dimensionless parameters like object number and Euler‐Poincaré characteristics. To better understand this phenomenon, we studied the horizontal deformation of thick (100 µm) transverse skeletal muscle sections, by comparing the muscle fibre diameters measured on thick sections to muscle fibre diameters measured on thin (10 µm) sections of the same sample. Diameter changes were further correlated with shrinkage in the Z direction (axial shrinkage) and deviation of the muscle fibre preferential axis from the Z‐axis. We showed that the thick sections dilated in horizontal and shrunk in Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in the Z direction. The latter was more pronounced in transversely than obliquely cut tissue sections. The results emphasise that even when shrinkage in the Z direction can be corrected using calibration, it is important to optimise histological protocols to minimise the Z‐axis collapse that could cause horizontal dilation.
Lay description
In skeletal muscle research, 3D analysis is especially important for studying the microvasculature. Laser scanning confocal microscopy of skeletal muscle thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, such sections are prone to deformations which may significantly influence the study results. To better understand this phenomenon, we studied the horizontal deformation of thick transverse skeletal muscle sections. We compared the average muscle fibre diameters measured on thick skeletal muscle sections, thin fixed skeletal muscle sections and immunohistochemically stained thin skeletal muscle sections with the muscle fibre diameters measured on thin native skeletal muscle sections of the same sample, with the latter condition serving as the standard diameters (ie the control condition). We further studied the association among muscle fibre diameter changes, shrinkage of the |
doi_str_mv | 10.1111/jmi.12985 |
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Lay description
In skeletal muscle research, 3D analysis is especially important for studying the microvasculature. Laser scanning confocal microscopy of skeletal muscle thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, such sections are prone to deformations which may significantly influence the study results. To better understand this phenomenon, we studied the horizontal deformation of thick transverse skeletal muscle sections. We compared the average muscle fibre diameters measured on thick skeletal muscle sections, thin fixed skeletal muscle sections and immunohistochemically stained thin skeletal muscle sections with the muscle fibre diameters measured on thin native skeletal muscle sections of the same sample, with the latter condition serving as the standard diameters (ie the control condition). We further studied the association among muscle fibre diameter changes, shrinkage of the thick skeletal muscle sections in the Z direction and their sectioning angle. We showed that the thick skeletal muscle sections dilated in the horizontal direction and shrunk in the Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in Z direction. The shrinkage in the Z direction was more pronounced in transversely than obliquely cut tissue sections. These results emphasise that even when shrinkage in the Z direction can be corrected using Z‐axis calibration, it is very important to optimise histological protocols to minimise the Z‐axis collapse that could cause horizontal dilation in order to enhance the integrity of study results.</description><identifier>ISSN: 0022-2720</identifier><identifier>EISSN: 1365-2818</identifier><identifier>DOI: 10.1111/jmi.12985</identifier><identifier>PMID: 33202057</identifier><language>eng</language><publisher>England: Wiley Subscription Services, Inc</publisher><subject>Calibration ; Capillaries ; Collapse ; Confocal microscopy ; Deformation ; Diameters ; Dilation ; Dimensionless numbers ; fibre diameter ; Fluorescence ; horizontal dilation ; Horizontal orientation ; Microscopy ; Microscopy, Confocal ; Microscopy, Fluorescence ; Microvasculature ; Muscle Fibers, Skeletal ; Muscle, Skeletal ; Muscles ; Musculoskeletal system ; Parameters ; sample deformation ; Sectioning ; Shrinkage ; shrinkage in Z direction ; Skeletal muscle</subject><ispartof>Journal of microscopy (Oxford), 2021-05, Vol.282 (2), p.113-122</ispartof><rights>2020 Royal Microscopical Society</rights><rights>2020 Royal Microscopical Society.</rights><rights>2021 Royal Microscopical Society</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><cites>FETCH-LOGICAL-c3135-6a4a2160507aac6c07abd804e4a2a642567d0a7f44b1e885a20be4b4439e1ac93</cites><orcidid>0000-0002-4414-5928 ; 0000-0003-0286-4999 ; 0000-0001-5831-2216</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1111%2Fjmi.12985$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1111%2Fjmi.12985$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,780,784,1417,1433,27924,27925,45574,45575,46409,46833</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/33202057$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Umek, Nejc</creatorcontrib><creatorcontrib>Janáček, Jiří</creatorcontrib><creatorcontrib>Cvetko, Erika</creatorcontrib><creatorcontrib>Eržen, Ida</creatorcontrib><title>Horizontal deformation of skeletal muscle thick sections visualised by confocal microscopy</title><title>Journal of microscopy (Oxford)</title><addtitle>J Microsc</addtitle><description>Certain morphological parameters of the skeletal muscle tissue can be better understood via 3D considerations. Fluorescent confocal microscopy of thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, thick tissue sections are prone to deformations which may significantly influence some stereological and morphometric results like muscle fibre diameter and capillary length, but not dimensionless parameters like object number and Euler‐Poincaré characteristics. To better understand this phenomenon, we studied the horizontal deformation of thick (100 µm) transverse skeletal muscle sections, by comparing the muscle fibre diameters measured on thick sections to muscle fibre diameters measured on thin (10 µm) sections of the same sample. Diameter changes were further correlated with shrinkage in the Z direction (axial shrinkage) and deviation of the muscle fibre preferential axis from the Z‐axis. We showed that the thick sections dilated in horizontal and shrunk in Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in the Z direction. The latter was more pronounced in transversely than obliquely cut tissue sections. The results emphasise that even when shrinkage in the Z direction can be corrected using calibration, it is important to optimise histological protocols to minimise the Z‐axis collapse that could cause horizontal dilation.
Lay description
In skeletal muscle research, 3D analysis is especially important for studying the microvasculature. Laser scanning confocal microscopy of skeletal muscle thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, such sections are prone to deformations which may significantly influence the study results. To better understand this phenomenon, we studied the horizontal deformation of thick transverse skeletal muscle sections. We compared the average muscle fibre diameters measured on thick skeletal muscle sections, thin fixed skeletal muscle sections and immunohistochemically stained thin skeletal muscle sections with the muscle fibre diameters measured on thin native skeletal muscle sections of the same sample, with the latter condition serving as the standard diameters (ie the control condition). We further studied the association among muscle fibre diameter changes, shrinkage of the thick skeletal muscle sections in the Z direction and their sectioning angle. We showed that the thick skeletal muscle sections dilated in the horizontal direction and shrunk in the Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in Z direction. The shrinkage in the Z direction was more pronounced in transversely than obliquely cut tissue sections. These results emphasise that even when shrinkage in the Z direction can be corrected using Z‐axis calibration, it is very important to optimise histological protocols to minimise the Z‐axis collapse that could cause horizontal dilation in order to enhance the integrity of study results.</description><subject>Calibration</subject><subject>Capillaries</subject><subject>Collapse</subject><subject>Confocal microscopy</subject><subject>Deformation</subject><subject>Diameters</subject><subject>Dilation</subject><subject>Dimensionless numbers</subject><subject>fibre diameter</subject><subject>Fluorescence</subject><subject>horizontal dilation</subject><subject>Horizontal orientation</subject><subject>Microscopy</subject><subject>Microscopy, Confocal</subject><subject>Microscopy, Fluorescence</subject><subject>Microvasculature</subject><subject>Muscle Fibers, Skeletal</subject><subject>Muscle, Skeletal</subject><subject>Muscles</subject><subject>Musculoskeletal system</subject><subject>Parameters</subject><subject>sample deformation</subject><subject>Sectioning</subject><subject>Shrinkage</subject><subject>shrinkage in Z direction</subject><subject>Skeletal muscle</subject><issn>0022-2720</issn><issn>1365-2818</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2021</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp10EFLwzAUB_AgipvTg19ACl700C1JkzY7ylA3UbzoxUtI01fM1jazaZX66U3t9CCYy4PHjz95f4ROCZ4S_2br0kwJnQu-h8YkinlIBRH7aIwxpSFNKB6hI-fWGGPBBT5EoyiimGKejNHL0tbm01aNKoIMcluXqjG2CmweuA0U0O_L1ukCgubV6E3gQPfABe_GtaowDrIg7QJtq9zqHhtdW6fttjtGB7kqHJzs5gQ931w_LZbh_ePtanF1H-qIRDyMFVOUxJjjRCkdaz_STGAGfq1iRnmcZFglOWMpASG4ojgFljIWzYEoPY8m6GLI3db2rQXXyNI4DUWhKrCtk5TFxN_NBfH0_A9d27au_O8k5YRG8yQhsVeXg-ovcTXkclubUtWdJFj2hUtfuPwu3NuzXWKblpD9yp-GPZgN4MMU0P2fJO8eVkPkFx8Pikg</recordid><startdate>202105</startdate><enddate>202105</enddate><creator>Umek, Nejc</creator><creator>Janáček, Jiří</creator><creator>Cvetko, Erika</creator><creator>Eržen, Ida</creator><general>Wiley Subscription Services, Inc</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7U5</scope><scope>8FD</scope><scope>L7M</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0002-4414-5928</orcidid><orcidid>https://orcid.org/0000-0003-0286-4999</orcidid><orcidid>https://orcid.org/0000-0001-5831-2216</orcidid></search><sort><creationdate>202105</creationdate><title>Horizontal deformation of skeletal muscle thick sections visualised by confocal microscopy</title><author>Umek, Nejc ; Janáček, Jiří ; Cvetko, Erika ; Eržen, Ida</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3135-6a4a2160507aac6c07abd804e4a2a642567d0a7f44b1e885a20be4b4439e1ac93</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2021</creationdate><topic>Calibration</topic><topic>Capillaries</topic><topic>Collapse</topic><topic>Confocal microscopy</topic><topic>Deformation</topic><topic>Diameters</topic><topic>Dilation</topic><topic>Dimensionless numbers</topic><topic>fibre diameter</topic><topic>Fluorescence</topic><topic>horizontal dilation</topic><topic>Horizontal orientation</topic><topic>Microscopy</topic><topic>Microscopy, Confocal</topic><topic>Microscopy, Fluorescence</topic><topic>Microvasculature</topic><topic>Muscle Fibers, Skeletal</topic><topic>Muscle, Skeletal</topic><topic>Muscles</topic><topic>Musculoskeletal system</topic><topic>Parameters</topic><topic>sample deformation</topic><topic>Sectioning</topic><topic>Shrinkage</topic><topic>shrinkage in Z direction</topic><topic>Skeletal muscle</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Umek, Nejc</creatorcontrib><creatorcontrib>Janáček, Jiří</creatorcontrib><creatorcontrib>Cvetko, Erika</creatorcontrib><creatorcontrib>Eržen, Ida</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>Technology Research Database</collection><collection>Advanced Technologies Database with Aerospace</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of microscopy (Oxford)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Umek, Nejc</au><au>Janáček, Jiří</au><au>Cvetko, Erika</au><au>Eržen, Ida</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Horizontal deformation of skeletal muscle thick sections visualised by confocal microscopy</atitle><jtitle>Journal of microscopy (Oxford)</jtitle><addtitle>J Microsc</addtitle><date>2021-05</date><risdate>2021</risdate><volume>282</volume><issue>2</issue><spage>113</spage><epage>122</epage><pages>113-122</pages><issn>0022-2720</issn><eissn>1365-2818</eissn><abstract>Certain morphological parameters of the skeletal muscle tissue can be better understood via 3D considerations. Fluorescent confocal microscopy of thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, thick tissue sections are prone to deformations which may significantly influence some stereological and morphometric results like muscle fibre diameter and capillary length, but not dimensionless parameters like object number and Euler‐Poincaré characteristics. To better understand this phenomenon, we studied the horizontal deformation of thick (100 µm) transverse skeletal muscle sections, by comparing the muscle fibre diameters measured on thick sections to muscle fibre diameters measured on thin (10 µm) sections of the same sample. Diameter changes were further correlated with shrinkage in the Z direction (axial shrinkage) and deviation of the muscle fibre preferential axis from the Z‐axis. We showed that the thick sections dilated in horizontal and shrunk in Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in the Z direction. The latter was more pronounced in transversely than obliquely cut tissue sections. The results emphasise that even when shrinkage in the Z direction can be corrected using calibration, it is important to optimise histological protocols to minimise the Z‐axis collapse that could cause horizontal dilation.
Lay description
In skeletal muscle research, 3D analysis is especially important for studying the microvasculature. Laser scanning confocal microscopy of skeletal muscle thick tissue sections is a well‐established method for visualising and measuring skeletal muscle fibres and surrounding capillaries in 3D. However, such sections are prone to deformations which may significantly influence the study results. To better understand this phenomenon, we studied the horizontal deformation of thick transverse skeletal muscle sections. We compared the average muscle fibre diameters measured on thick skeletal muscle sections, thin fixed skeletal muscle sections and immunohistochemically stained thin skeletal muscle sections with the muscle fibre diameters measured on thin native skeletal muscle sections of the same sample, with the latter condition serving as the standard diameters (ie the control condition). We further studied the association among muscle fibre diameter changes, shrinkage of the thick skeletal muscle sections in the Z direction and their sectioning angle. We showed that the thick skeletal muscle sections dilated in the horizontal direction and shrunk in the Z direction, and that the magnitude of horizontal dilation was associated with the magnitude of shrinkage in Z direction. The shrinkage in the Z direction was more pronounced in transversely than obliquely cut tissue sections. These results emphasise that even when shrinkage in the Z direction can be corrected using Z‐axis calibration, it is very important to optimise histological protocols to minimise the Z‐axis collapse that could cause horizontal dilation in order to enhance the integrity of study results.</abstract><cop>England</cop><pub>Wiley Subscription Services, Inc</pub><pmid>33202057</pmid><doi>10.1111/jmi.12985</doi><tpages>10</tpages><orcidid>https://orcid.org/0000-0002-4414-5928</orcidid><orcidid>https://orcid.org/0000-0003-0286-4999</orcidid><orcidid>https://orcid.org/0000-0001-5831-2216</orcidid></addata></record> |
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subjects | Calibration Capillaries Collapse Confocal microscopy Deformation Diameters Dilation Dimensionless numbers fibre diameter Fluorescence horizontal dilation Horizontal orientation Microscopy Microscopy, Confocal Microscopy, Fluorescence Microvasculature Muscle Fibers, Skeletal Muscle, Skeletal Muscles Musculoskeletal system Parameters sample deformation Sectioning Shrinkage shrinkage in Z direction Skeletal muscle |
title | Horizontal deformation of skeletal muscle thick sections visualised by confocal microscopy |
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