Domain > xenomuscle.com

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DNS Resolutions

Date	IP Address
2021-06-15	160.153.60.163 (ClassC)
2026-02-08	107.180.57.185 (ClassC)

HTTP/1.1 200 OKDate: Sun, 08 Feb 2026 18:29:31 GMTServer: ApacheX-Powered-By: PHP/8.1.34Permissions-Policy: private-state-token-redemption(self https://www.google.com https://www.gstatic.com https://recaptcha.net https://challenges.cloudflare.com https://hcaptcha.com), private-state-token-issuance(self https://www.google.com https://www.gstatic.com https://recaptcha.net https://challenges.cloudflare.com https://hcaptcha.com)Link: https://xenomuscle.com/wp-json/>; relhttps://api.w.org/, https://xenomuscle.com/wp-json/wp/v2/pages/28>; relalternate; titleJSON; typeapplication/json, https://xenomuscle.com/>; relshortlinkUpgrade: h2,h2cConnection: UpgradeVary: Accept-EncodingTransfer-Encoding: chunkedContent-Type: text/html; charsetUTF-8

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!important;}.has-nv-light-bg-border-color{border-color: var(--wp--preset--color--nv-light-bg) !important;}.has-nv-dark-bg-border-color{border-color: var(--wp--preset--color--nv-dark-bg) !important;}.has-neve-text-color-border-color{border-color: var(--wp--preset--color--neve-text-color) !important;}.has-nv-text-dark-bg-border-color{border-color: var(--wp--preset--color--nv-text-dark-bg) !important;}.has-nv-c-1-border-color{border-color: var(--wp--preset--color--nv-c-1) !important;}.has-nv-c-2-border-color{border-color: var(--wp--preset--color--nv-c-2) !important;}.has-vivid-cyan-blue-to-vivid-purple-gradient-background{background: var(--wp--preset--gradient--vivid-cyan-blue-to-vivid-purple) !important;}.has-light-green-cyan-to-vivid-green-cyan-gradient-background{background: var(--wp--preset--gradient--light-green-cyan-to-vivid-green-cyan) !important;}.has-luminous-vivid-amber-to-luminous-vivid-orange-gradient-background{background: var(--wp--preset--gradient--luminous-vivid-amber-to-luminous-vivid-orange) !important;}.has-luminous-vivid-orange-to-vivid-red-gradient-background{background: var(--wp--preset--gradient--luminous-vivid-orange-to-vivid-red) !important;}.has-very-light-gray-to-cyan-bluish-gray-gradient-background{background: var(--wp--preset--gradient--very-light-gray-to-cyan-bluish-gray) !important;}.has-cool-to-warm-spectrum-gradient-background{background: var(--wp--preset--gradient--cool-to-warm-spectrum) !important;}.has-blush-light-purple-gradient-background{background: var(--wp--preset--gradient--blush-light-purple) !important;}.has-blush-bordeaux-gradient-background{background: var(--wp--preset--gradient--blush-bordeaux) !important;}.has-luminous-dusk-gradient-background{background: var(--wp--preset--gradient--luminous-dusk) !important;}.has-pale-ocean-gradient-background{background: var(--wp--preset--gradient--pale-ocean) !important;}.has-electric-grass-gradient-background{background: var(--wp--preset--gradient--electric-grass) !important;}.has-midnight-gradient-background{background: var(--wp--preset--gradient--midnight) !important;}.has-small-font-size{font-size: var(--wp--preset--font-size--small) !important;}.has-medium-font-size{font-size: var(--wp--preset--font-size--medium) !important;}.has-large-font-size{font-size: var(--wp--preset--font-size--large) !important;}.has-x-large-font-size{font-size: var(--wp--preset--font-size--x-large) !important;}:where(.wp-block-columns.is-layout-flex){gap: 2em;}:where(.wp-block-columns.is-layout-grid){gap: 2em;}/*# sourceURLglobal-styles-inline-css *//style>link relstylesheet idneve-style-css hrefhttps://xenomuscle.com/wp-content/themes/neve/style-main-new.min.css mediaall />style idneve-style-inline-css>.is-menu-sidebar .header-menu-sidebar { visibility: visible; }.is-menu-sidebar.menu_sidebar_slide_left .header-menu-sidebar { transform: translate3d(0, 0, 0); left: 0; }.is-menu-sidebar.menu_sidebar_slide_right .header-menu-sidebar { transform: translate3d(0, 0, 0); right: 0; }.is-menu-sidebar.menu_sidebar_pull_right .header-menu-sidebar, .is-menu-sidebar.menu_sidebar_pull_left .header-menu-sidebar { transform: translateX(0); }.is-menu-sidebar.menu_sidebar_dropdown .header-menu-sidebar { height: auto; }.is-menu-sidebar.menu_sidebar_dropdown .header-menu-sidebar-inner { max-height: 400px; padding: 20px 0; }.is-menu-sidebar.menu_sidebar_full_canvas .header-menu-sidebar { opacity: 1; }.header-menu-sidebar .menu-item-nav-search:not(.floating) { pointer-events: none; }.header-menu-sidebar .menu-item-nav-search .is-menu-sidebar { pointer-events: unset; }@media screen and (max-width: 960px) { .builder-item.cr .item--inner { --textalign: center; --justify: center; } }.nv-meta-list li.meta:not(:last-child):after { content:/ }.nv-meta-list .no-mobile{			display:none;		}.nv-meta-list li.last::after{			content: !important;		}@media (min-width: 769px) {			.nv-meta-list .no-mobile {				display: inline-block;			}			.nv-meta-list li.last:not(:last-child)::after {		 		content: / !important;			}		} :root{ --container: 748px;--postwidth:100%; --primarybtnbg: var(--nv-primary-accent); --primarybtnhoverbg: var(--nv-primary-accent); --primarybtncolor: #fff; --secondarybtncolor: var(--nv-primary-accent); --primarybtnhovercolor: #fff; --secondarybtnhovercolor: var(--nv-primary-accent);--primarybtnborderradius:3px;--secondarybtnborderradius:3px;--secondarybtnborderwidth:3px;--btnpadding:13px 15px;--primarybtnpadding:13px 15px;--secondarybtnpadding:calc(13px - 3px) calc(15px - 3px); --bodyfontfamily: Roboto; --bodyfontsize: 15px; --bodylineheight: 1.6; --bodyletterspacing: 0px; --bodyfontweight: 400; --headingsfontfamily: IBM Plex Mono; --h1fontsize: 1.5em; --h1fontweight: 800; --h1lineheight: 1.6em; --h1letterspacing: 0px; --h1texttransform: none; --h2fontsize: 1.3em; --h2fontweight: 600; --h2lineheight: 1.6em; --h2letterspacing: 0px; --h2texttransform: none; --h3fontsize: 24px; --h3fontweight: 700; --h3lineheight: 1.4; --h3letterspacing: 0px; --h3texttransform: none; --h4fontsize: 20px; --h4fontweight: 700; --h4lineheight: 1.6; --h4letterspacing: 0px; --h4texttransform: none; --h5fontsize: 16px; --h5fontweight: 700; --h5lineheight: 1.6; --h5letterspacing: 0px; --h5texttransform: none; --h6fontsize: 14px; --h6fontweight: 700; --h6lineheight: 1.6; --h6letterspacing: 0px; --h6texttransform: none;--formfieldborderwidth:2px;--formfieldborderradius:3px; --formfieldbgcolor: var(--nv-site-bg); --formfieldbordercolor: #dddddd; --formfieldcolor: var(--nv-text-color);--formfieldpadding:10px 12px; } .nv-index-posts{ --borderradius:0px; } .has-neve-button-color-color{ color: var(--nv-primary-accent)!important; } .has-neve-button-color-background-color{ background-color: var(--nv-primary-accent)!important; } .alignfull > class*__inner-container, .alignwide > class*__inner-container{ max-width:718px;margin:auto } .nv-meta-list{ --avatarsize: 20px; } .single .nv-meta-list{ --avatarsize: 20px; } .nv-post-cover{ --height: 250px;--padding:40px 15px;--justify: flex-start; --textalign: left; --valign: center; } .nv-post-cover .nv-title-meta-wrap, .nv-page-title-wrap, .entry-header{ --textalign: left; } .nv-is-boxed.nv-title-meta-wrap{ --padding:40px 15px; --bgcolor: var(--nv-dark-bg); } .nv-overlay{ --opacity: 50; --blendmode: normal; } .nv-is-boxed.nv-comments-wrap{ --padding:20px; } .nv-is-boxed.comment-respond{ --padding:20px; } .single:not(.single-product), .page{ --c-vspace:0 0 0 0;; } .scroll-to-top{ --color: var(--nv-text-dark-bg);--padding:8px 10px; --borderradius: 3px; --bgcolor: var(--nv-primary-accent); --hovercolor: var(--nv-text-dark-bg); --hoverbgcolor: var(--nv-primary-accent);--size:16px; } .global-styled{ --bgcolor: var(--nv-site-bg); } .header-top{ --rowbcolor: var(--nv-light-bg); --color: var(--nv-text-color); --bgcolor: var(--nv-site-bg); } .header-main{ --rowbcolor: var(--nv-light-bg); --color: var(--nv-text-color); --bgcolor: var(--nv-site-bg); } .header-bottom{ --rowbcolor: var(--nv-light-bg); --color: var(--nv-text-color); --bgcolor: var(--nv-site-bg); } .header-menu-sidebar-bg{ --justify: flex-start; --textalign: left;--flexg: 1;--wrapdropdownwidth: auto; --color: var(--nv-text-color); --bgcolor: var(--nv-site-bg); } .header-menu-sidebar{ width: 360px; } .builder-item--logo{ --maxwidth: 120px; --color: #193840; --fs: 24px;--padding:10px 0;--margin:0; --textalign: left;--justify: flex-start; } .builder-item--nav-icon,.header-menu-sidebar .close-sidebar-panel .navbar-toggle{ --borderradius:0; } .builder-item--nav-icon{ --label-margin:0 5px 0 0;;--padding:10px 15px;--margin:0; } .builder-item--primary-menu{ --hovercolor: var(--nv-secondary-accent); --hovertextcolor: var(--nv-text-color); --activecolor: var(--nv-primary-accent); --spacing: 20px; --height: 25px;--padding:0;--margin:0; --fontsize: 1em; --lineheight: 1.6; --letterspacing: 0px; --fontweight: 500; --texttransform: none; --iconsize: 1em; } .hfg-is-group.has-primary-menu .inherit-ff{ --inheritedfw: 500; } .footer-top-inner .row{ grid-template-columns:1fr 1fr 1fr; --valign: flex-start; } .footer-top{ --rowbcolor: var(--nv-light-bg); --color: var(--nv-text-color); --bgcolor: var(--nv-site-bg); } .footer-main-inner .row{ grid-template-columns:1fr 1fr 1fr; --valign: flex-start; } .footer-main{ --rowbcolor: var(--nv-light-bg); --color: var(--nv-text-color); --bgcolor: var(--nv-site-bg); } .footer-bottom-inner .row{ grid-template-columns:1fr 1fr 1fr; --valign: flex-start; } .footer-bottom{ --rowbcolor: var(--nv-light-bg); --color: var(--nv-text-dark-bg); --bgcolor: var(--nv-dark-bg); } @media(min-width: 576px){ :root{ --container: 992px;--postwidth:50%;--btnpadding:13px 15px;--primarybtnpadding:13px 15px;--secondarybtnpadding:calc(13px - 3px) calc(15px - 3px); --bodyfontsize: 16px; --bodylineheight: 1.6; --bodyletterspacing: 0px; --h1fontsize: 1.5em; --h1lineheight: 1.6em; --h1letterspacing: 0px; --h2fontsize: 1.3em; --h2lineheight: 1.6em; --h2letterspacing: 0px; --h3fontsize: 26px; --h3lineheight: 1.4; --h3letterspacing: 0px; --h4fontsize: 22px; --h4lineheight: 1.5; --h4letterspacing: 0px; --h5fontsize: 18px; --h5lineheight: 1.6; --h5letterspacing: 0px; --h6fontsize: 14px; --h6lineheight: 1.6; --h6letterspacing: 0px; } .nv-meta-list{ --avatarsize: 20px; } .single .nv-meta-list{ --avatarsize: 20px; } .nv-post-cover{ --height: 320px;--padding:60px 30px;--justify: flex-start; --textalign: left; --valign: center; } .nv-post-cover .nv-title-meta-wrap, .nv-page-title-wrap, .entry-header{ --textalign: left; } .nv-is-boxed.nv-title-meta-wrap{ --padding:60px 30px; } .nv-is-boxed.nv-comments-wrap{ --padding:30px; } .nv-is-boxed.comment-respond{ --padding:30px; } .single:not(.single-product), .page{ --c-vspace:0 0 0 0;; } .scroll-to-top{ --padding:8px 10px;--size:16px; } .header-menu-sidebar-bg{ --justify: flex-start; --textalign: left;--flexg: 1;--wrapdropdownwidth: auto; } .header-menu-sidebar{ width: 360px; } .builder-item--logo{ --maxwidth: 120px; --fs: 24px;--padding:10px 0;--margin:0; --textalign: left;--justify: flex-start; } .builder-item--nav-icon{ --label-margin:0 5px 0 0;;--padding:10px 15px;--margin:0; } .builder-item--primary-menu{ --spacing: 20px; --height: 25px;--padding:0;--margin:0; --fontsize: 1em; --lineheight: 1.6; --letterspacing: 0px; --iconsize: 1em; } }@media(min-width: 960px){ :root{ --container: 1170px;--postwidth:33.333333333333%;--btnpadding:13px 15px;--primarybtnpadding:13px 15px;--secondarybtnpadding:calc(13px - 3px) calc(15px - 3px); --bodyfontsize: 16px; --bodylineheight: 1.7; --bodyletterspacing: 0px; --h1fontsize: 2em; --h1lineheight: 1.6em; --h1letterspacing: 0px; --h2fontsize: 1.6em; --h2lineheight: 1.6em; --h2letterspacing: 0px; --h3fontsize: 28px; --h3lineheight: 1.4; --h3letterspacing: 0px; --h4fontsize: 24px; --h4lineheight: 1.5; --h4letterspacing: 0px; --h5fontsize: 20px; --h5lineheight: 1.6; --h5letterspacing: 0px; --h6fontsize: 16px; --h6lineheight: 1.6; --h6letterspacing: 0px; } #content .container .col, #content .container-fluid .col{ max-width: 70%; } .alignfull > class*__inner-container, .alignwide > class*__inner-container{ max-width:789px } .container-fluid .alignfull > class*__inner-container, .container-fluid .alignwide > class*__inner-container{ max-width:calc(70% + 15px) } .nv-sidebar-wrap, .nv-sidebar-wrap.shop-sidebar{ max-width: 30%; } .nv-meta-list{ --avatarsize: 20px; } .single .nv-meta-list{ --avatarsize: 20px; } .nv-post-cover{ --height: 400px;--padding:60px 40px;--justify: flex-start; --textalign: left; --valign: center; } .nv-post-cover .nv-title-meta-wrap, .nv-page-title-wrap, .entry-header{ --textalign: left; } .nv-is-boxed.nv-title-meta-wrap{ --padding:60px 40px; } .nv-is-boxed.nv-comments-wrap{ --padding:40px; } .nv-is-boxed.comment-respond{ --padding:40px; } .single:not(.single-product), .page{ --c-vspace:0 0 0 0;; } .scroll-to-top{ --padding:8px 10px;--size:16px; } .header-menu-sidebar-bg{ --justify: flex-start; --textalign: left;--flexg: 1;--wrapdropdownwidth: auto; } .header-menu-sidebar{ width: 360px; } .builder-item--logo{ --maxwidth: 120px; --fs: 24px;--padding:10px 0;--margin:0; --textalign: left;--justify: flex-start; } .builder-item--nav-icon{ --label-margin:0 5px 0 0;;--padding:10px 15px;--margin:0; } .builder-item--primary-menu{ --spacing: 20px; --height: 25px;--padding:0;--margin:0; --fontsize: 1em; --lineheight: 1.6; --letterspacing: 0px; --iconsize: 1em; } }.scroll-to-top {right: 20px; border: none; position: fixed; bottom: 30px; display: none; opacity: 0; visibility: hidden; transition: opacity 0.3s ease-in-out, visibility 0.3s ease-in-out; align-items: center; justify-content: center; z-index: 999; } @supports (-webkit-overflow-scrolling: touch) { .scroll-to-top { bottom: 74px; } } .scroll-to-top.image { background-position: center; } .scroll-to-top .scroll-to-top-image { width: 100%; height: 100%; } .scroll-to-top .scroll-to-top-label { margin: 0; padding: 5px; } .scroll-to-top:hover { text-decoration: none; } .scroll-to-top.scroll-to-top-left {left: 20px; right: unset;} .scroll-to-top.scroll-show-mobile { display: flex; } @media (min-width: 960px) { .scroll-to-top { display: flex; } }.scroll-to-top { color: var(--color); padding: var(--padding); border-radius: var(--borderradius); background: var(--bgcolor); } .scroll-to-top:hover, .scroll-to-top:focus { color: var(--hovercolor); background: var(--hoverbgcolor); } .scroll-to-top-icon, .scroll-to-top.image .scroll-to-top-image { width: var(--size); height: var(--size); } .scroll-to-top-image { background-image: var(--bgimage); background-size: cover; }:root{--nv-primary-accent:#0366d6;--nv-secondary-accent:#0e509a;--nv-site-bg:#ffffff;--nv-light-bg:#ededed;--nv-dark-bg:#14171c;--nv-text-color:#393939;--nv-text-dark-bg:#ffffff;--nv-c-1:#77b978;--nv-c-2:#f37262;--nv-fallback-ff:Arial, Helvetica, sans-serif;} #content.neve-main .container .alignfull > class*__inner-container,#content.neve-main .alignwide > class*__inner-container{ max-width: 718px; } @media(min-width: 576px){ #content.neve-main .container .alignfull > class*__inner-container,#content.neve-main .alignwide > class*__inner-container{ max-width: 962px; } } @media(min-width: 960px) { #content.neve-main .container .alignfull > class*__inner-container,#content.neve-main .container .alignwide > class*__inner-container{ max-width: 1140px; } #content.neve-main > .container > .row > .col{ max-width: 100%; } body:not(.neve-off-canvas) #content.neve-main > .container > .row > .nv-sidebar-wrap, body:not(.neve-off-canvas) #content.neve-main > .container > .row > .nv-sidebar-wrap.shop-sidebar { max-width: 0%; } } /*# sourceURLneve-style-inline-css *//style>link relstylesheet idneve-google-font-roboto-css href//fonts.googleapis.com/css?familyRoboto%3A400%2C500&displayswap&ver4.2.2 mediaall />link relstylesheet idneve-google-font-ibm-plex-mono-css href//fonts.googleapis.com/css?familyIBM+Plex+Mono%3A400%2C800%2C600%2C700&displayswap&ver4.2.2 mediaall />link relhttps://api.w.org/ hrefhttps://xenomuscle.com/wp-json/ />link relalternate titleJSON typeapplication/json hrefhttps://xenomuscle.com/wp-json/wp/v2/pages/28 />link relEditURI typeapplication/rsd+xml titleRSD hrefhttps://xenomuscle.com/xmlrpc.php?rsd />meta namegenerator contentWordPress 6.9.1 />link relcanonical hrefhttps://xenomuscle.com/ />link relshortlink hrefhttps://xenomuscle.com/ />	style>img#wpstats{display:none}/style>		link relicon hrefhttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/cropped-mouse_favicon_512x512.png?fit32%2C32&ssl1 sizes32x32 />link relicon hrefhttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/cropped-mouse_favicon_512x512.png?fit192%2C192&ssl1 sizes192x192 />link relapple-touch-icon hrefhttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/cropped-mouse_favicon_512x512.png?fit180%2C180&ssl1 />meta namemsapplication-TileImage contenthttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/cropped-mouse_favicon_512x512.png?fit270%2C270&ssl1 />		style idwp-custom-css>			/*Limit max site width*/body {	max-width: 1300px;	margin-left: auto !important;	margin-right: auto !important;}/*Top Menu Font Changes*/.builder-item--primary-menu {	font-family: IBM Plex Mono, Arial, Helvetica, sans-serif;	font-weight: 500;	}/*Site Title/Logo Changes*/.builder-item--logo .site-logo {	font-family: IBM Plex Mono, Arial, Helvetica, sans-serif;	font-weight: 600;}/*Restrict full width image size*/@media (min-width: 1920px) {	body {    max-width: 1920px;    padding-right: 15px;    padding-left: 15px;		margin-left: auto;		margin-right: auto;}}/*styling links*/a {  color: #327080/*#1a4c58*/ !important;  text-decoration: none !important;}a:hover {	color: #193840/*#387a8a*/ !important;  text-decoration: underline!important;}a:active {  color: #fc348d !important;}a:visited {  color: #541750;}/*style site 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idneve_body  >div classwrapper>		header classheader  >		a classneve-skip-link show-on-focus href#content >			Skip to content		/a>		div idheader-grid  classhfg_header site-header>	nav classheader--row header-main hide-on-mobile hide-on-tablet layout-full-contained nv-navbar header--row	data-row-idmain data-show-ondesktop>	div		classheader--row-inner header-main-inner>		div classcontainer>			div				classrow row--wrapper				data-sectionhfg_header_layout_main >				div classhfg-slot left>div classbuilder-item desktop-left>div classitem--inner builder-item--logo		data-sectiontitle_tagline		data-item-idlogo>	div classsite-logo>	a classbrand hrefhttps://xenomuscle.com/ aria-labelxenomuscle relhome>div classnv-title-tagline-wrap>p classsite-title>xenomuscle/p>small>/small>/div>/a>/div>	/div>/div>/div>div classhfg-slot right>div classbuilder-item desktop-left>div classitem--inner builder-item--nav-icon		data-sectionheader_menu_icon		data-item-idnav-icon>	div classmenu-mobile-toggle item-button 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classwrap>a hrefhttps://xenomuscle.com/#engrafted aria-currentpage>Engrafted Muscle/a>/div>/li>li idmenu-item-244 classmenu-item menu-item-type-custom menu-item-object-custom current-menu-item current_page_item menu-item-home menu-item-244 nv-active>div classwrap>a hrefhttps://xenomuscle.com/#markers aria-currentpage>Pathological Markers/a>/div>/li>li idmenu-item-245 classmenu-item menu-item-type-custom menu-item-object-custom current-menu-item current_page_item menu-item-home menu-item-245 nv-active>div classwrap>a hrefhttps://xenomuscle.com/#uses aria-currentpage>Xenografting Uses/a>/div>/li>li idmenu-item-354 classmenu-item menu-item-type-post_type menu-item-object-page menu-item-354>div classwrap>a hrefhttps://xenomuscle.com/licensing-inquiries/>Licensing Inquiries/a>/div>/li>li idmenu-item-43 classmenu-item menu-item-type-post_type menu-item-object-page menu-item-43>div classwrap>a hrefhttps://xenomuscle.com/contact/>Contact/a>/div>/li>/ul>	/div>/div>	/div>/div>					/div>	/div>/div>div classheader-menu-sidebar-overlay hfg-ov hfg-pe onclickif(undefined ! typeof toggleAriaClick ) { toggleAriaClick() }>/div>/div>	/header>	style>.nav-ul li:focus-within .wrap.active + .sub-menu { opacity: 1; visibility: visible; }.nav-ul li.neve-mega-menu:focus-within .wrap.active + .sub-menu { display: grid; }.nav-ul li > .wrap { display: flex; align-items: center; position: relative; padding: 0 4px; }.nav-ul:not(.menu-mobile):not(.neve-mega-menu) > li > .wrap > a { padding-top: 1px }/style>		main idcontent classneve-main>div classcontainer single-page-container>	div classrow>				div classnv-single-page-wrap col>			div classnv-content-wrap entry-content>figure classwp-block-image alignfull size-full>img data-recalc-dims1 fetchpriorityhigh decodingasync width1022 height621 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/whole-xenograft_horiz.jpg?resize1022%2C621 alt classwp-image-23 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/whole-xenograft_horiz.jpg?w1022&ssl1 1022w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/whole-xenograft_horiz.jpg?resize300%2C182&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/whole-xenograft_horiz.jpg?resize768%2C467&ssl1 768w sizes(max-width: 1022px) 100vw, 1022px />/figure>h2 classwp-block-heading idinmice>Making Human Skeletal Muscle Tissue in Mice/h2>p>We have developed a method to xenograft immortalized human muscle precursor cells from patients with FSHD and first-degree relative controls into the em>tibialis anterior/em> muscle compartment of immuno-deficient mice, generating human muscle xenografts. We report that FSHD cells mature into organized and innervated human muscle fibers with minimal contamination of murine myonuclei. They also reconstitute the satellite cell niche within the xenografts. FSHD xenografts express em>DUX4/em> and DUX4 downstream targets, retain the 4q35 epigenetic signature of their original donors, and express two novel protein biomarkers of FSHD, TPM3 and SLC34A2. Ours is the first scalable, mature em>in vivo/em>human model of FSHD. It should be useful for studies of the pathogenic mechanism of the disease as well as for testing therapeutic strategies targeting em>DUX4/em> expression. /p>h2 classwp-block-heading idapproach>Our Approacha href#approachasterisk>sup>*/sup>/a>/h2>p>We use immunodeficient Nod-RAG (NRG; NOD.Cg-Rag1sup>tm1Mom/sup>l2rgsup>tm-Wjl/sup>/SzJ) mice.  We first X-irradiate their hindlimbs, to prevent the regeneration of the hindlimb muscles in later steps. We then inject a myotoxin along the length of the em>tibialis anterior /em>(TA) muscles of each limb. (We have tested a number of different toxins, the best being cardiotoxin and BaClsub>2/sub>.  As cardiotoxin is no longer commercially available, we typically use BaClsub>2/sub>.)  The following day, we inject a concentrated suspension of human myogenic precursor cells (hMPCs) along the muscle.  One week later, we electrically stimulate the lower hindlimb muscles, including the TA, via the peroneal nerve for 30 min a day, 3 times a week.  This intermittent neuromuscular electrical stimulataion (iNMES) is continued for 4 weeks.  At that point, the xenograft has developed hundreds of human muscle fibers that are mature and that contain very few, if any, murine myonuclei (<2%). Mice may then be euthanized or used for further studies. /p>div classwp-block-image>figure classaligncenter size-large>img data-recalc-dims1 decodingasync width938 height464 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/engraftment_timeline.png?resize938%2C464&ssl1 altEngraftment Timeline classwp-image-376 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/engraftment_timeline.png?w938&ssl1 938w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/engraftment_timeline.png?resize300%2C148&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/engraftment_timeline.png?resize768%2C380&ssl1 768w sizes(max-width: 938px) 100vw, 938px />/figure>/div>p>An overview of the timeline is shown above. Details of the procedures are reported in a href#references>references 1 and 2/a>./p>p idapproachasterisk>em>*These methods are patented in the US (a href#references>reference 3/a>)/em>/p>hr classwp-block-separator alignfull is-style-wide/>div classwp-block-columns is-layout-flex wp-container-core-columns-is-layout-9d6595d7 wp-block-columns-is-layout-flex>div classwp-block-column is-layout-flow wp-block-column-is-layout-flow>h2 classhas-text-align-center wp-block-heading>a href#engrafted data-typeinternal data-id#engrafted>Engrafted Muscle/a>/h2>figure classwp-block-image size-large>img data-recalc-dims1 decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES-1024x1024.jpg?resize1024%2C1024 alt classwp-image-15 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?resize768%2C768&ssl1 768w sizes(max-width: 1024px) 100vw, 1024px />/figure>/div>div classwp-block-column is-layout-flow wp-block-column-is-layout-flow>h2 classhas-text-align-center wp-block-heading>a href#markers>Pathological Markers/a>/h2>figure classwp-block-image size-large>img data-recalc-dims1 loadinglazy decodingasync width438 height438 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/DXUr_FSHD_square.png?resize438%2C438 altDXUr FSHD classwp-image-234 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/DXUr_FSHD_square.png?w438&ssl1 438w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/DXUr_FSHD_square.png?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/DXUr_FSHD_square.png?resize150%2C150&ssl1 150w sizesauto, (max-width: 438px) 100vw, 438px />/figure>/div>div classwp-block-column is-layout-flow wp-block-column-is-layout-flow>h2 classhas-text-align-center wp-block-heading>a href#uses data-typeinternal data-id#uses>Xenografting Uses/a>/h2>figure classwp-block-image size-large>img data-recalc-dims1 loadinglazy decodingasync width646 height646 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_char_squaret.jpg?resize646%2C646 altFulcrum Chart classwp-image-233 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_char_squaret.jpg?w646&ssl1 646w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_char_squaret.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_char_squaret.jpg?resize150%2C150&ssl1 150w sizesauto, (max-width: 646px) 100vw, 646px />/figure>/div>/div>hr classwp-block-separator is-style-wide/>h2 classwp-block-heading idengrafted>Characteristics of the Engrafted Human Muscle/h2>p>The fibers in graft are human, with minimal contamination by murine myonuclei.  They are also mature, by several criteria./p>h3 classwp-block-heading>Identifying the Human Myofibers/h3>p>Human myofibers in the grafts are readily identified by immunofluorescence methods, following incubation with antibodies that recognize human but not murine muscle proteins.  We typically use monoclonal antibodies to human β-spectrin (h-β-spectrin)  and to human lamin A/C (h-lamin A/C; see figure).  Human fibers in the xenograft are labeled green with antibodies to h-β-spectrin; fibers around the periphery of the graft are not, and thus are murine in origin.  Similarly, myonuclei in fibers labeled for h-β-spectrin are also positive for h-lamin A/C (purple). The occasional murine myonucleus (blue) is not./p>div classwp-block-columns is-layout-flex wp-container-core-columns-is-layout-9d6595d7 wp-block-columns-is-layout-flex>div classwp-block-column is-layout-flow wp-block-column-is-layout-flow>figure classwp-block-image size-large is-resized>img data-recalc-dims1 loadinglazy decodingasync srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/h-bspectrin_desmin.jpg?resize431%2C543 alth-β-spectrin  classwp-image-191 width431 height543 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/h-bspectrin_desmin.jpg?w574&ssl1 574w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/h-bspectrin_desmin.jpg?resize238%2C300&ssl1 238w sizesauto, (max-width: 431px) 100vw, 431px />/figure>/div>div classwp-block-column is-layout-flow wp-block-column-is-layout-flow>figure classwp-block-gallery columns-2 is-cropped wp-block-gallery-1 is-layout-flex wp-block-gallery-is-layout-flex>ul classblocks-gallery-grid>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width210 height270 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/mouse_myonucleus.png?resize210%2C270 altMouse Myonucleus data-id51 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/mouse_myonucleus.png data-linkhttps://xenomuscle.com/characteristics-of-the-engrafted-human-muscle/attachment/mouse_myonucleus/ classwp-image-51/>figcaption classblocks-gallery-item__caption>Mouse myonucleus br>– for h-lamin A/C /figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width206 height262 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/human_myonucleus.png?resize206%2C262 altHuman Myonucleus data-id50 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/human_myonucleus.png data-linkhttps://xenomuscle.com/characteristics-of-the-engrafted-human-muscle/attachment/human_myonucleus/ classwp-image-50/>figcaption classblocks-gallery-item__caption>Human myonucleus  br>+  for h-lamin A/C            /figcaption>/figure>/li>/ul>/figure>/div>/div>h3 classwp-block-heading>State of Differentiation/h3>p>We examined myosin in the contractile apparatus in the human myofibers generated following our standard protocol, with iNMES, and without electrical stimulation.  Fibers stimulated electrically eliminate nearly all their embryonic myosin, whereas unstimulated human fibers do not (see figure).  This suggests that our protocol generates mature human skeletal muscle tissue./p>figure classwp-block-gallery columns-2 is-cropped wp-block-gallery-2 is-layout-flex wp-block-gallery-is-layout-flex>ul classblocks-gallery-grid>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES-1024x1024.jpg?resize1024%2C1024 alt data-id15 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg data-linkhttps://xenomuscle.com/myh-emb_noinmes/ classwp-image-15 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_noiNMES.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>-iNMES/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_iNMES-1024x1024.jpg?resize1024%2C1024 alt data-id14 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_iNMES.jpg data-linkhttps://xenomuscle.com/myh-emb_inmes/ classwp-image-14 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_iNMES.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_iNMES.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_iNMES.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/MYH-emb_iNMES.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>+iNMES/figcaption>/figure>/li>/ul>/figure>p>The presence of fully differentiated fibers in the xenografts is also indicated by the fact that desmin is organized in striations, when fibers are observed in longitudinal section, and in a reticulum surrounding the myofibrils, when fibers are observed in cross section (desmin in red; see Figure)./p>figure classwp-block-gallery columns-2 is-cropped wp-block-gallery-3 is-layout-flex wp-block-gallery-is-layout-flex>ul classblocks-gallery-grid>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width322 height322 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/human_spectrin_desmin.png?resize322%2C322 althuman spectrin desmin data-id195 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2021/03/human_spectrin_desmin.png data-linkhttps://xenomuscle.com/introduction/attachment/human_spectrin_desmin/ classwp-image-195 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/human_spectrin_desmin.png?w322&ssl1 322w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/human_spectrin_desmin.png?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/03/human_spectrin_desmin.png?resize150%2C150&ssl1 150w sizesauto, (max-width: 322px) 100vw, 322px />/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/desmin-composite-RGB-1024x1024.jpg?resize1024%2C1024 alt data-id7 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/desmin-composite-RGB.jpg data-linkhttps://xenomuscle.com/desmin-composite-rgb/ classwp-image-7 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/desmin-composite-RGB.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/desmin-composite-RGB.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/desmin-composite-RGB.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/desmin-composite-RGB.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />/figure>/li>/ul>/figure>h3 classwp-block-heading>Innervation/h3>p>The human fibers are innervated, as indicated by the presence of presynaptic termini labeled for synaptophysin (Syp, green) and postsynaptic membranes enriched in acetylcholine receptors (AchR, labeled with fluorescent α-bungarotoxin, blue) on the fiber surfaces (labeled for h-β-spectrin, red; see Figure)./p>figure classwp-block-gallery aligncenter columns-3 wp-block-gallery-4 is-layout-flex wp-block-gallery-is-layout-flex>ul classblocks-gallery-grid>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-synaptophysin-1024x1024.jpg?resize1024%2C1024 alt data-id13 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/innervation-synaptophysin.jpg data-linkhttps://xenomuscle.com/innervation-synaptophysin/ classwp-image-13 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-synaptophysin.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-synaptophysin.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-synaptophysin.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-synaptophysin.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>Syp/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-ACHR-1024x1024.jpg?resize1024%2C1024 alt data-id9 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/innervation-ACHR.jpg data-linkhttps://xenomuscle.com/innervation-achr/ classwp-image-9 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-ACHR.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-ACHR.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-ACHR.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-ACHR.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>AchR/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-merged-1024x1024.jpg?resize1024%2C1024 alt data-id11 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/innervation-merged.jpg data-linkhttps://xenomuscle.com/innervation-merged/ classwp-image-11 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-merged.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-merged.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-merged.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/innervation-merged.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>Merge/figcaption>/figure>/li>/ul>/figure>h3 classwp-block-heading>Satellite Cells/h3>p>The human fibers in the grafts are associated with human satellite cells, that have repopulated their niche between the plasma membrane (labeled for h-β-spectrin and h-lamin A/C; red) and the basal lamina (labeled for laminin; purple).   The satellite cell nucleus is labeled with DAPI (blue) and antibodies to Pax7 (green).  Thus, the grafts have the capacity to grow and regenerate./p>figure classwp-block-gallery columns-5 wp-block-gallery-5 is-layout-flex wp-block-gallery-is-layout-flex>ul classblocks-gallery-grid>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Dapi-1024x1024.jpg?resize1024%2C1024 alt data-id16 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Dapi.jpg data-linkhttps://xenomuscle.com/satellite-cell-dapi/ classwp-image-16 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Dapi.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Dapi.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Dapi.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Dapi.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>DAPI/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Pax7-1024x1024.jpg?resize1024%2C1024 alt data-id20 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Pax7.jpg data-linkhttps://xenomuscle.com/satellite-cell-pax7/ classwp-image-20 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Pax7.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Pax7.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Pax7.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-Pax7.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>Pax7/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-speclamAC-1024x1024.jpg?resize1024%2C1024 alt data-id21 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-speclamAC.jpg data-linkhttps://xenomuscle.com/satellite-cell-speclamac/ classwp-image-21 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-speclamAC.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-speclamAC.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-speclamAC.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-speclamAC.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>h-β-spec, h-lam/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-laminin-1024x1024.jpg?resize1024%2C1024 alt data-id17 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-laminin.jpg data-linkhttps://xenomuscle.com/satellite-cell-laminin/ classwp-image-17 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-laminin.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-laminin.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-laminin.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-laminin.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>Laminin/figcaption>/figure>/li>li classblocks-gallery-item>figure>img data-recalc-dims1 loadinglazy decodingasync width1024 height1024 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-merged-1024x1024.jpg?resize1024%2C1024 alt data-id18 data-full-urlhttps://xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-merged.jpg data-linkhttps://xenomuscle.com/satellite-cell-merged/ classwp-image-18 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-merged.jpg?w1024&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-merged.jpg?resize300%2C300&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-merged.jpg?resize150%2C150&ssl1 150w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/Satellite-cell-merged.jpg?resize768%2C768&ssl1 768w sizesauto, (max-width: 1024px) 100vw, 1024px />figcaption classblocks-gallery-item__caption>Merge/figcaption>/figure>/li>/ul>/figure>h2 classwp-block-heading idmarkers>Pathological Markers/h2>p>The disease we have been focusing on is Facioscapulohumeral Muscular Dystrophy (FSHD) which occurs in individuals with chromosomal contractions at position 4q135 that result in a reduction in the number of D4Z4 satellite repeats at that locus. This contraction leads to a decrease in methylation of the terminal D4Z4 repeat and an increase in its transcription. When a polyadenylation sequence is included in the transcript, an mRNA encoding the transcription factor, DUX4, is produced. DUX4’s activity as a transcription factor activates dozens of downstream genes.  Our results confirm that the xenografts prepared with FSHD cells replicate these biochemical features of FSHD.  They have additional, unexpected features, that we are documenting:/p>h3 classwp-block-heading>Genetic/h3>p>DUX4 is expressed in grafts developed from FSHD but not control cells./p>div classwp-block-image>figure classaligncenter size-large>img data-recalc-dims1 loadinglazy decodingasync width432 height390 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/dxu4.png?resize432%2C390 alt classwp-image-61 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/dxu4.png?w432&ssl1 432w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/dxu4.png?resize300%2C271&ssl1 300w sizesauto, (max-width: 432px) 100vw, 432px />/figure>/div>p>All the gene products downstream of DUX4 that we have tested in FSHD vs control grafts are also upregulated at the mRNA level, including TRIM43, MBD3L2, ZSCAN4, LEUTX, and SLC34A2.  Data for the first three are shown here.  Control gene products (not shown) do not differ between FSHD and control grafts./p>figure classwp-block-image size-large>img data-recalc-dims1 loadinglazy decodingasync width1024 height378 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/04/dux4_chart-1024x378.jpg?resize1024%2C378 alt classwp-image-210 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/04/dux4_chart.jpg?resize1024%2C378&ssl1 1024w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/04/dux4_chart.jpg?resize300%2C111&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/04/dux4_chart.jpg?resize768%2C284&ssl1 768w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/04/dux4_chart.jpg?w1430&ssl1 1430w sizesauto, (max-width: 1024px) 100vw, 1024px />/figure>h3 classwp-block-heading>Epigenetic/h3>p>Methylation of the terminal D4Z4 region of chromosome 4q35 is reduced in grafts prepared from FSHD but not control cells. (Methylated sites are indicated by red boxes, unmethylated, by blue boxes)/p>div classwp-block-image>figure classaligncenter size-large>img data-recalc-dims1 loadinglazy decodingasync width936 height206 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/FSHD-xenograft.png?resize936%2C206 alt classwp-image-62 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/FSHD-xenograft.png?w936&ssl1 936w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/FSHD-xenograft.png?resize300%2C66&ssl1 300w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/FSHD-xenograft.png?resize768%2C169&ssl1 768w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/FSHD-xenograft.png?resize930%2C206&ssl1 930w sizesauto, (max-width: 936px) 100vw, 936px />/figure>/div>h3 classwp-block-heading>Morphology/h3>p>Overall fiber diameter and spacing is not significantly different between FSHD and control grafts, but necrotic fibers, identified with an antibody to a potential biomarker of FSHD, SLC34A2, are present in significantly higher numbers in the FSHD grafts (arrow, Figure)/p>div classwp-block-image>figure classaligncenter size-large>img data-recalc-dims1 loadinglazy decodingasync width402 height445 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/arrow_biomarker.jpg?resize402%2C445 alt classwp-image-63 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/arrow_biomarker.jpg?w402&ssl1 402w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2020/08/arrow_biomarker.jpg?resize271%2C300&ssl1 271w sizesauto, (max-width: 402px) 100vw, 402px />/figure>/div>hr classwp-block-separator is-style-wide/>h2 classwp-block-heading>Work in Progress with the Xenografts/h2>p>Collaborations with a number of laboratories and biopharma companies have been established to test the abilities of different approaches to treating FSHD with small molecules (inhibitors of p38 kinase, gapmers), oligonucleotide-antibody conjugates, and adeno-associated viral vectors carrying CRISPR/Cas9 or DUX4 pseudosubstrates./p>p>We are also using the grafts in experiments to identify biomarkers of FSHD that have the potential to be used to monitor the efficacy of different therapies currently being developed for the clinic.  Our primary focus is currently on SLC43A2, a protein that is upregulated by DUX4 expression./p>p>Experiments to explore variations of our xenografting protocols are also in progress, with3  goals: producing grafts with still higher numbers of human skeletal myofibers, reducing variability among grafts, and minimizing the effort required to generate the grafts./p>h2 classwp-block-heading iduses>strong>Uses of the Xenografting Methods/strong>/h2>h3 classwp-block-heading>Disease Models/h3>p>Many studies of neuromuscular diseases, such as muscular dystrophies and myopathies, have been performed in animal models, especially in mice, in which the mutations that cause disease in man can be generated.  Although the murine models can sometimes only approximate the human disease, they have been very useful in identifying some of the underlying pathogenic mechanisms and in initial testing of therapies.  But murine models of human diseases that are not the result of simple genetic mutations are much harder to generate.  Our ability to prepare mice carrying mature human skeletal muscle tissue for individuals with such diseases offers the best murine model available for mechanistic and therapeutic investigations.  /p>h3 classwp-block-heading>Testing the Efficacy and Specificity of Therapies/h3>p>Therapies designed to treat muscular dystrophies typically use either small molecules as drugs or genetic approaches to replace the protein missing in the disease or to suppress the expression of a protein that causes the disease. Testing the efficacy and specificity of these treatments on patients is a lengthy and expensive process.  Having mature human muscle tissue to test in mice provides a quicker and cheaper alternative.  Off-target and other non-specific effects as well as the efficacy of different treatments can be readily assessed in our xenografts. An example of our use of the xenografts to test a small molecule therapy for FSHD is the work we did with Fulcrum Therapeutics to examine the effects of p38 kinase inhibitors on the expression of em>DUX4/em> and its downstream products. Data we obtained with Fulcrum testing the Lilly drug, LY 2228820, is shown here.  The results show that the drug reduced expression of mRNA encoding 3 of DUX4’s downstream products (black, controls; blue, + LY compound)/p>div classwp-block-image>figure classaligncenter size-large>img data-recalc-dims1 loadinglazy decodingasync width720 height405 srchttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_chart.jpg?resize720%2C405 altFulcrum Chart classwp-image-226 srcsethttps://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_chart.jpg?w720&ssl1 720w, https://i0.wp.com/xenomuscle.com/wp-content/uploads/2021/06/fulcrum_chart.jpg?resize300%2C169&ssl1 300w sizesauto, (max-width: 720px) 100vw, 720px />/figure>/div>h2 classwp-block-heading idlicensing>Licensing/h2>p>Any parties interested in licensing this technology for research and/or commercial purposes should contact the Office of Technology Transfer at the University of Maryland, Baltimore a hrefhttps://xenomuscle.com/licensing-inquiries/ data-typeURL data-idhttps://xenomuscle.com/licensing-inquiries/>here/a>./p>h5 classwp-block-heading idreferences>strong>References/strong>/h5>ol classwp-block-list>li>Sakellariou, P., A. O’Neill, A.L. Mueller, G. Stadler, W.E. Wright, J.A. Roche and R.J. Bloch (2016) Neuromuscular Electrical Stimulation Promotes Development in Mice  of Mature Human Muscle from Immortalized Human Myoblasts. Skeletal Muscle 6, 4   a href26925213 target_blank relnoreferrer noopener>PMID:26925213/a>; a hrefhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4769538/ target_blank relnoreferrer noopener>PMC4769538/a>./li>li>Mueller, A.L., A. O’Neill, T.I. Jones, A. Llach-Martinez, L.A. Rojas, P. Sakellariou, G. Stadler,  W.E. Wright, D. Eyerman, P.L. Jones, R.J. Bloch (2019) Muscle Xenografts Reproduce Key Molecular Features of Facioscapulohumeral Muscular Dystrophy. Exp. Neurol 320:113011 a hrefhttps://pubmed.ncbi.nlm.nih.gov/31306642/ data-typeURL data-idhttps://pubmed.ncbi.nlm.nih.gov/31306642/ target_blank relnoreferrer noopener>PMID:31306642/a>; a hrefhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC6730665/>PMC6730665/a>/li>li>a>Bloch, R.J., /a>P. Sakellariou, A. O’Neill, and J.A. Roche (2020) “Methods of Generating Mature Human Muscle Fibers” a hrefhttps://patents.google.com/patent/US20160256681A1/en data-typeURL data-idhttps://patents.google.com/patent/US20160256681A1/en target_blank relnoreferrer noopener>US Patent Number 10,632,305/a> issued 4/28/2020. Valid thru 12/8/2036. /li>/ol>p>/p>/div>		/div>			/div>/div>/main>!--/.neve-main-->button tabindex0 idscroll-to-top classscroll-to-top scroll-to-top-right  scroll-show-mobile icon aria-labelScroll To Top>svg classscroll-to-top-icon aria-hiddentrue roleimg xmlnshttp://www.w3.org/2000/svg width15 height15 viewBox0 0 15 15>rect width15 height15 fillnone/>path fillcurrentColor dM2,8.48l-.65-.65a.71.71,0,0,1,0-1L7,1.14a.72.72,0,0,1,1,0l5.69,5.7a.71.71,0,0,1,0,1L13,8.48a.71.71,0,0,1-1,0L8.67,4.94v8.42a.7.7,0,0,1-.7.7H7a.7.7,0,0,1-.7-.7V4.94L3,8.47a.7.7,0,0,1-1,0Z/>/svg>/button>footer classsite-footer idsite-footer  >	div classhfg_footer>		div classfooter--row footer-bottom layout-full-contained	idcb-row--footer-desktop-bottom	data-row-idbottom data-show-ondesktop>	div		classfooter--row-inner footer-bottom-inner footer-content-wrap>		div classcontainer>			div				classhfg-grid nv-footer-content hfg-grid-bottom row--wrapper row 				data-sectionhfg_footer_layout_bottom >				div classhfg-slot left>div classbuilder-item cr>div classitem--inner>div classcomponent-wrap>div>p>© 2022 xenomuscle/p>/div>/div>/div>/div>/div>div classhfg-slot c-left>/div>div classhfg-slot center>/div>							/div>		/div>	/div>/div>	/div>/footer>/div>!--/.wrapper-->script typespeculationrules>{prefetch:{source:document,where:{and:{href_matches:/*},{not:{href_matches:/wp-*.php,/wp-admin/*,/wp-content/uploads/*,/wp-content/*,/wp-content/plugins/*,/wp-content/themes/neve/*,/*\\?(.+)}},{not:{selector_matches:arel~\nofollow\}},{not:{selector_matches:.no-prefetch, .no-prefetch a}}},eagerness:conservative}}/script>script idneve-script-js-extra>var NeveProperties  {ajaxurl:https://xenomuscle.com/wp-admin/admin-ajax.php,nonce:fffecc5148,isRTL:,isCustomize:};//# sourceURLneve-script-js-extra/script>script srchttps://xenomuscle.com/wp-content/themes/neve/assets/js/build/modern/frontend.js idneve-script-js async>/script>script idneve-script-js-after>	var html  document.documentElement;	var theme  html.getAttribute(data-neve-theme) || light;	var variants  {logo:{light:{src:false,srcset:false,sizes:false},dark:{src:false,srcset:false,sizes:false},same:true}};	function setCurrentTheme( theme ) {		var pictures  document.getElementsByClassName( neve-site-logo );		for(var i  0; ipictures.length; i++) {			var picture  pictures.item(i);			if( ! picture ) {				continue;			};			var fileExt  picture.src.slice((Math.max(0, picture.src.lastIndexOf(.)) || Infinity) + 1);			if ( fileExt  svg ) {				picture.removeAttribute(width);				picture.removeAttribute(height);				picture.style  width: var(--maxwidth);			}			var compId  picture.getAttribute(data-variant);			if ( compId && variantscompId ) {				var isConditional  variantscompIdsame;				if ( theme  light || isConditional || variantscompIddarksrc  false ) {					picture.src  variantscompIdlightsrc;					picture.srcset  variantscompIdlightsrcset || ;					picture.sizes  variantscompIdlightsizes;					continue;				};				picture.src  variantscompIddarksrc;				picture.srcset  variantscompIddarksrcset || ;				picture.sizes  variantscompIddarksizes;			};		};	};	var observer  new MutationObserver(function(mutations) {		mutations.forEach(function(mutation) {			if (mutation.type  attributes) {				theme  html.getAttribute(data-neve-theme);				setCurrentTheme(theme);			};		});	});	observer.observe(html, {		attributes: true	});function toggleAriaClick() { function toggleAriaExpanded(toggle  true) { document.querySelectorAll(button.navbar-toggle).forEach(function(el) { if ( el.classList.contains(caret-wrap) ) { return; } el.setAttribute(aria-expanded, true  el.getAttribute(aria-expanded) ? false : toggle); }); } toggleAriaExpanded(); if ( document.body.hasAttribute(data-ftrap-listener) ) { return; } document.body.setAttribute(data-ftrap-listener, true); document.addEventListener(ftrap-end, function() { toggleAriaExpanded(false); }); }//# sourceURLneve-script-js-after/script>script idneve-scroll-to-top-js-extra>var neveScrollOffset  {offset:0};//# sourceURLneve-scroll-to-top-js-extra/script>script srchttps://xenomuscle.com/wp-content/themes/neve/assets/js/build/modern/scroll-to-top.js idneve-scroll-to-top-js async>/script>script idjetpack-stats-js-before>_stq  window._stq || ;_stq.push( view, {v:ext,blog:181703194,post:28,tz:0,srv:xenomuscle.com,j:1:15.4} );_stq.push( clickTrackerInit, 181703194, 28 );//# sourceURLjetpack-stats-js-before/script>script srchttps://stats.wp.com/e-202606.js idjetpack-stats-js defer data-wp-strategydefer>/script>script idwp-emoji-settings typeapplication/json>{baseUrl:https://s.w.org/images/core/emoji/17.0.2/72x72/,ext:.png,svgUrl:https://s.w.org/images/core/emoji/17.0.2/svg/,svgExt:.svg,source:{concatemoji:https://xenomuscle.com/wp-includes/js/wp-emoji-release.min.js}}/script>script typemodule>/*! 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Domain > xenomuscle.com

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