Human Umbilical Cord Wharton's Jelly Derived Mesenchymal Stromal Cells May Attenuate Sarcopenia in Aged Mice Induced by Hindlimb Suspension

Abstract

BACKGROUND Since the use of human umbilical cord Wharton's Jelly derived mesenchymal stromal cells (hWJ-MSCs) to treat sarcopenia has not been explored, we studied the effects of hWJ-MSCs in aged male C57BL/6J mice with sarcopenia induced by hindlimb suspension, and explored the potential mechanism. MATERIAL AND METHODS Hindlimb suspension was used to induce sarcopenia in 24-month-old C57BL/6J mice and green fluorescent protein-tagged hWJ-MSCs and controls were transplanted into mice via tail vein or local intramuscular injection. After hWJ-MSC transplantation, changes in whole body muscle strength and endurance, gastrocnemius muscle weight and myofiber cross-sectional area (CSA) were studied. Proliferation of skeletal muscle stem cell, apoptosis, and chronic inflammation were also investigated. RESULTS We demonstrated that whole body muscle strength and endurance, gastrocnemius muscle mass, and CSA were significantly increased in hWJ-MSC-transplanted mice than in controls (P<0.05). In hWJ-MSC-transplanted mice, apoptotic myonuclei was reduced, and BrdU and Pax-7 expression indices of gastrocnemius muscles were increased (P<0.05). Tumor necrosis factor (TNF)-α and interleukin (IL)-6 were downregulated, and IL-4 and IL-10 were upregulated (P<0.05). CONCLUSIONS hWJ-MSCs may ameliorate sarcopenia in aged male C57BL/6J mice induced by hindlimb suspension, and this may be via activation of resident skeletal muscle satellite cells, reduction of apoptosis, and less chronic inflammation.

Figure 1

Effect of human umbilical cord Wharton’s Jelly derived mesenchymal stromal cell (hWJ-MSC) transplantation on whole body muscle strength and endurance in sarcopenic mouse model. Quantification of whole body muscle strength and endurance using latency to fall during a wire hang test. Pre-hindlimb suspension (HS), before hindlimb suspension; post-HS, after 14 days of hindlimb suspension; after-T, after 8 weeks of hWJ-MSC transplantation. MT – muscular transplant group; MT-Con – MT-control group; TVT – tail vein transplant group; TVT-Con – TVT-control group. * P<0.05, compared to relative control in after-T group; ^# P<0.05, compared to relative post-HS group; ^&&& P<0.001, compared to relative pre-HS group.

Figure 2

Effect of hWJ-MSC transplantation on muscle mass in sarcopenic mouse model. Absolute gastrocnemius mass (A), gastrocnemius mass (body weight standardized) (B). * P<0.05, ** P<0.01, *** P<0.001, compared to relative control; ^# P<0.05, compared to MT group. hWJ-MSC – human umbilical cord Wharton’s Jelly derived mesenchymal stromal cell; MT – muscular transplant.

Figure 3

Histology of gastrocnemius muscle and measurements of the cross-sectional area (CSA). Hematoxylin and eosin (H&E) stain (A), absolute muscle fiber CSA (B), muscle fiber CSA (body weight standardized) (C). * P<0.05, *** P<0.001, compared to relative control.

Figure 4

Changes in BrdU and Pax-7 expression in gastrocnemius muscle after hWJ-MSC transplantation. BrdU expression (A, B), Pax-7 expression (C, D). A and C show BrdU and Pax-7 immunofluorescent staining of gastrocnemius muscle cross section: Pax7 (red) and BrdU (red) expression, basal lamina (green) and nuclear (DAPI blue) identification. B and D show BrdU and Pax-7 positive cell index for each group. *** P<0.001, compared to relative control; ^## P<0.01, compared to MT group. Pax-7 – paired box 7; hWJ-MSC – human umbilical cord Wharton’s Jelly derived mesenchymal stromal cell; MT – muscular transplant.

Figure 5

Changes of nuclear apoptotic indices after hWJ-MSC transplantation into gastrocnemius muscles of aged C57BL/6J mice. (A) TUNEL immunofluorescent staining of gastrocnemius muscle cross section: representative TUNEL labeled (green), basal lamina (red), and nuclear (DAPI blue) identification. (B) Nuclear apoptotic indices as indicated by the number of TUNEL-positive myonuclei to total myonuclei. *** P<0.001, compared with relative control. hWJ-MSC – human umbilical cord Wharton’s Jelly derived mesenchymal stromal cell; TUNEL – terminal deoxynucleotidyl transferase (TdT)-mediated deoxy-UTP nick end labeling.

Figure 6

Cytokine expression after hWJ-MSC transplantation into gastrocnemius muscles of aged C57BL/6J mice. The expression levels of IL-4, IL-10, TNF-α, and IL-6 were detected by ELISA. * P<0.05, ** P<0.01, *** P<0.001, versus relative control. ^# P<0.05, compared to MT group. Data are means ± standard deviation and analyzed by a 2-way ANOVA. hWJ-MSC – human umbilical cord Wharton’s Jelly derived mesenchymal stromal cell; IL – interleukin; TNF – tumor necrosis factor; ELISA – enzyme-linked immunosorbent assay; MT – muscular transplant.