Int J Stem Cells.  2016 Nov;9(2):239-249. 10.15283/ijsc16011.

Intramuscular Transplantation of Allogeneic Mesenchymal Stromal Cells Derived from Equine Umbilical Cord

• Dias MC
• da Cruz Landim-Alvarenga F
• de Moraes
• da Costa L
• Geraldini
• de Vasconcelos Machado VM
• Maia L

• KMID: 2362518
• DOI: http://doi.org/10.15283/ijsc16011

Abstract

BACKGROUND AND OBJECTIVES
Mesenchymal stromal cells (MSCs) have great therapeutic potential, particularly in the process of tissue repair and immunomodulation through the secretion of biomolecules. Thus, the aim of this study was to evaluate the hypothesis that intramuscular transplantation of allogeneic MSCs obtained from equine umbilical cord (UC-MSCs) is safe, demonstrating that this is a suitable source of stem cells for therapeutic use.
METHODS AND RESULTS
For this, UC-MSCs were cultured, characterized and cryopreserved for future transplantation in six healthy mares. On day 0, transplantation of three million UC-MSCs diluted in Hank's Balanced Solution (HBSS) was performed on right and left sides of the rump muscle. As a control, HBSS injections were performed caudally in the same muscle. Muscle biopsies were obtained as a control 30 days before transplantation (D-30). The biopsies were collected again on day 2 (left side) and day 7 (right side) post transplantation and examined histologically. All procedures were preceded by ultrasound examination and blood sampling. Hematologic evaluation remained within normal limits and no differences were observed between time points (p>0.05). Ultrasound examination was suggestive of inflammation 48 hours after transplantation in both groups (control and treated). At histological evaluation it was found only discrete inflammation signals between D-30×D2 (p<0.05) in the treated group, without differences (p> 0.05) between the groups at different time points.
CONCLUSIONS
Equine UC-MSCs under the experimental conditions did not promote severe inflammation that causes tissue damage or lead to its rejection by the host organism and therefore has a good potential for clinical use.

Keyword

Mesenchymal stromal cells; Allogeneic; Transplantation; Muscle; Umbilical cord

MeSH Terms

Biopsy
Immunomodulation
Inflammation
Mesenchymal Stromal Cells*
Stem Cells
Transplantation
Ultrasonography
Umbilical Cord*

Figure

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  Fig. 1 Scheme of UC-MSCs transplantation and muscle biopsies. (A) Location of transplanting with UC-MSCs and HBBS. (B) Location of biopsies performed after transplantation (D2 on the left side and D7 on the right side).

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  Fig. 2 Immunophenotypic analysis by flow cytometry of the markers CD34, CD44, CD90 and MHC-II of fresh UC- MSCs (A, C, E, G) and after cryopreservation (B, D, F, H). (A, B) Representative histograms of marker CD34 fresh (A) and after cryopreservation (B). (C, D) Representative histograms of marker CD44 (C) fresh and after cryopreservation (D). (E, F) Representative histograms of marker CD90 (E) fresh and after cryopreservation (F). (G, H) Representative histograms of marker MHCII (G) fresh and after cryopreservation (H).

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  Fig. 3 Sonograms of equine superficial gluteal muscle at D0, D2 and D7 moments. (A) D0. Region that did not received UC-MSCs/HBSS injection. Note the arrangement of the muscle fibers of a healthy tissue. (B) Needle image (yellow arrow) being guided by ultrasound at a depth of about 2 cm. (C) D2. Muscles with heterogeneous aspect (red ellipse) and presence of hypo echogenic regions (yellow arrow), after UC-MSCs injection. (D) D7. Improved aspect of the muscles with the presence of discrete hypoechoic regions permeated at the muscle fibers (red ellipse).

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  Fig. 4 Photomicrographs of biopsied superficial gluteal muscle without histological changes. (A, B) Moment D-30. (C) Moment D2, treated. (D) Moment D2, control. (E) Moment D7, treated. (F) Moment D7, control.

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  Fig. 5 Photomicrographs of superficial gluteal muscle biopsies with signals of mild inflammation. (A) Treated group at moment D2: Inflammatory infiltrate composed mostly of macrophages, and rare neutrophils. Degeneration (arrowhead) and necrosis (arrow) of muscle fibers indicated by the increase of sarcoplasmic eosinophilia and nuclear pyknosis were noticed. (B) Control group at moment D2: Interfascicular inflammatory infiltrate composed by macrophages and rare lymphocytes. Degenerated muscle fibers (arrowhead). (C) Treated group at moment D7: Inflammatory infiltrate involving the collagenous stroma (perimysium). (D) Control group at moment D7: Photomicrograph showing discrete perivascular inflammatory infiltrate composed by macrophages and discrete lymphocytes, associated with interfascicular edema.

Reference

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