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Intraarterial injection of muscle-derived CD34(+)Sca-1(+) stem cells restores dystrophin in mdx mice.

Torrente Y, Tremblay JP, Pisati F, Belicchi M, Rossi B, Sironi M, Fortunato F, El Fahime M, D'Angelo MG, Caron NJ, Constantin G, Paulin D, Scarlato G, Bresolin N - J. Cell Biol. (2001)

Bottom Line: One way to circumvent this obstacle would be to use circulating cells capable of homing to the sites of lesions.Normal dystrophin transcripts detected enzymes in the muscles of the hind limb injected intraarterially by the mdx reverse transcription polymerase chain reaction method, which differentiates between normal and mdx message.Our results showed that the muscle-derived stem cells first attach to the capillaries of the muscles and then participate in regeneration after muscle damage.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Ricovero e Cura a Carattere Scientifico (IRCCS) Ospedale Maggiore Policlinico, 20122 Milan, Italy.

ABSTRACT
Duchenne muscular dystrophy is a lethal recessive disease characterized by widespread muscle damage throughout the body. This increases the difficulty of cell or gene therapy based on direct injections into muscles. One way to circumvent this obstacle would be to use circulating cells capable of homing to the sites of lesions. Here, we showed that stem cell antigen 1 (Sca-1), CD34 double-positive cells purified from the muscle tissues of newborn mice are multipotent in vitro and can undergo both myogenic and multimyeloid differentiation. These muscle-derived stem cells were isolated from newborn mice expressing the LacZ gene under the control of the muscle-specific desmin or troponin I promoter and injected into arterial circulation of the hindlimb of mdx mice. The ability of these cells to interact and firmly adhere to endothelium in mdx muscles microcirculation was demonstrated by intravital microscopy after an intraarterial injection. Donor Sca-1, CD34 muscle-derived stem cells were able to migrate from the circulation into host muscle tissues. Histochemical analysis showed colocalization of LacZ and dystrophin expression in all muscles of the injected hindlimb in all of five out of five 8-wk-old treated mdx mice. Their participation in the formation of muscle fibers was significantly increased by muscle damage done 48 h after their intraarterial injection, as indicated by the presence of 12% beta-galactosidase-positive fibers in muscle cross sections. Normal dystrophin transcripts detected enzymes in the muscles of the hind limb injected intraarterially by the mdx reverse transcription polymerase chain reaction method, which differentiates between normal and mdx message. Our results showed that the muscle-derived stem cells first attach to the capillaries of the muscles and then participate in regeneration after muscle damage.

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Intravital microscopy: muscle-derived cells (pp6) firmly adhere in muscle capillaires. To improve contrast between the intra- and extravascular compartment, the animals were injected intravenously with a low dose of FITC-dextran. Venules (A) and arterioles (E) are indicated by asterisks and arrowheads, respectively. The BCECF-labeled cells (bright dots indicated by arrows) can be seen in the muscle capillaries (A, B, and C) 1 min after intraarterial injection. Arrows in D indicate a migrated BCECF-labeled cell in the perivascular space of a muscle venule 1 h after injection. Images before (E) and during the injection (F) are also presented.
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Figure 6: Intravital microscopy: muscle-derived cells (pp6) firmly adhere in muscle capillaires. To improve contrast between the intra- and extravascular compartment, the animals were injected intravenously with a low dose of FITC-dextran. Venules (A) and arterioles (E) are indicated by asterisks and arrowheads, respectively. The BCECF-labeled cells (bright dots indicated by arrows) can be seen in the muscle capillaries (A, B, and C) 1 min after intraarterial injection. Arrows in D indicate a migrated BCECF-labeled cell in the perivascular space of a muscle venule 1 h after injection. Images before (E) and during the injection (F) are also presented.

Mentions: Since intravenous injection of precursor cells offers the advantages of a total body distribution, we believe that this approach is also limited by physiological dispersion of injected cells in the blood-stream of several tissues (especially in liver and lungs). In our experiments, we verified the possibility in an elective biodistribution in muscle limbs after intraarterial injection of muscle-derived stem cells. We tested the adhesive behavior of Sca-1, CD34 positive cells to muscle vessels. Boluses of 7 × 105 fluorescently labeled cells were injected through a carotid artery catheter and their adhesion to the pectoral muscle vessels was recorded (Fig. 6). Sca-1, CD34 cells had a small diameter and interacted especially with muscle capillaries, but rarely with venules. The Sca-1, CD34 positive cells (in pp6) were distinguishable from the other preplates by their significant adhesive properties to the capillaries (data not shown). Adhesion fractions of injected pp6 were determined by counting the number of interacting cells in each muscle vessel per number of cells that passed through the same vessel during an injection. Injected cells adhered prevalently within muscle capillaries and the adhesion fraction was ∼89%.


Intraarterial injection of muscle-derived CD34(+)Sca-1(+) stem cells restores dystrophin in mdx mice.

Torrente Y, Tremblay JP, Pisati F, Belicchi M, Rossi B, Sironi M, Fortunato F, El Fahime M, D'Angelo MG, Caron NJ, Constantin G, Paulin D, Scarlato G, Bresolin N - J. Cell Biol. (2001)

Intravital microscopy: muscle-derived cells (pp6) firmly adhere in muscle capillaires. To improve contrast between the intra- and extravascular compartment, the animals were injected intravenously with a low dose of FITC-dextran. Venules (A) and arterioles (E) are indicated by asterisks and arrowheads, respectively. The BCECF-labeled cells (bright dots indicated by arrows) can be seen in the muscle capillaries (A, B, and C) 1 min after intraarterial injection. Arrows in D indicate a migrated BCECF-labeled cell in the perivascular space of a muscle venule 1 h after injection. Images before (E) and during the injection (F) are also presented.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199616&req=5

Figure 6: Intravital microscopy: muscle-derived cells (pp6) firmly adhere in muscle capillaires. To improve contrast between the intra- and extravascular compartment, the animals were injected intravenously with a low dose of FITC-dextran. Venules (A) and arterioles (E) are indicated by asterisks and arrowheads, respectively. The BCECF-labeled cells (bright dots indicated by arrows) can be seen in the muscle capillaries (A, B, and C) 1 min after intraarterial injection. Arrows in D indicate a migrated BCECF-labeled cell in the perivascular space of a muscle venule 1 h after injection. Images before (E) and during the injection (F) are also presented.
Mentions: Since intravenous injection of precursor cells offers the advantages of a total body distribution, we believe that this approach is also limited by physiological dispersion of injected cells in the blood-stream of several tissues (especially in liver and lungs). In our experiments, we verified the possibility in an elective biodistribution in muscle limbs after intraarterial injection of muscle-derived stem cells. We tested the adhesive behavior of Sca-1, CD34 positive cells to muscle vessels. Boluses of 7 × 105 fluorescently labeled cells were injected through a carotid artery catheter and their adhesion to the pectoral muscle vessels was recorded (Fig. 6). Sca-1, CD34 cells had a small diameter and interacted especially with muscle capillaries, but rarely with venules. The Sca-1, CD34 positive cells (in pp6) were distinguishable from the other preplates by their significant adhesive properties to the capillaries (data not shown). Adhesion fractions of injected pp6 were determined by counting the number of interacting cells in each muscle vessel per number of cells that passed through the same vessel during an injection. Injected cells adhered prevalently within muscle capillaries and the adhesion fraction was ∼89%.

Bottom Line: One way to circumvent this obstacle would be to use circulating cells capable of homing to the sites of lesions.Normal dystrophin transcripts detected enzymes in the muscles of the hind limb injected intraarterially by the mdx reverse transcription polymerase chain reaction method, which differentiates between normal and mdx message.Our results showed that the muscle-derived stem cells first attach to the capillaries of the muscles and then participate in regeneration after muscle damage.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Ricovero e Cura a Carattere Scientifico (IRCCS) Ospedale Maggiore Policlinico, 20122 Milan, Italy.

ABSTRACT
Duchenne muscular dystrophy is a lethal recessive disease characterized by widespread muscle damage throughout the body. This increases the difficulty of cell or gene therapy based on direct injections into muscles. One way to circumvent this obstacle would be to use circulating cells capable of homing to the sites of lesions. Here, we showed that stem cell antigen 1 (Sca-1), CD34 double-positive cells purified from the muscle tissues of newborn mice are multipotent in vitro and can undergo both myogenic and multimyeloid differentiation. These muscle-derived stem cells were isolated from newborn mice expressing the LacZ gene under the control of the muscle-specific desmin or troponin I promoter and injected into arterial circulation of the hindlimb of mdx mice. The ability of these cells to interact and firmly adhere to endothelium in mdx muscles microcirculation was demonstrated by intravital microscopy after an intraarterial injection. Donor Sca-1, CD34 muscle-derived stem cells were able to migrate from the circulation into host muscle tissues. Histochemical analysis showed colocalization of LacZ and dystrophin expression in all muscles of the injected hindlimb in all of five out of five 8-wk-old treated mdx mice. Their participation in the formation of muscle fibers was significantly increased by muscle damage done 48 h after their intraarterial injection, as indicated by the presence of 12% beta-galactosidase-positive fibers in muscle cross sections. Normal dystrophin transcripts detected enzymes in the muscles of the hind limb injected intraarterially by the mdx reverse transcription polymerase chain reaction method, which differentiates between normal and mdx message. Our results showed that the muscle-derived stem cells first attach to the capillaries of the muscles and then participate in regeneration after muscle damage.

Show MeSH
Related in: MedlinePlus