Chronic delivery of a thrombospondin-1 mimetic decreases skeletal muscle capillarity in mice

Abstract

Angiogenesis is an essential process for normal skeletal muscle function. There is a growing body of evidence suggesting that thrombospondin-1 (TSP-1), a potent antiangiogenic protein in tumorigenesis, is an important regulator of both physiological and pathological skeletal muscle angiogenesis. We tested the hypothesis that chronic exposure to a TSP-1 mimetic (ABT-510), which targets the CD36 TSP-1 receptor, would decrease skeletal muscle capillarity as well as alter the balance between positive and negative angiogenic proteins under basal conditions. Osmotic minipumps with either ABT-510 or vehicle (5% dextrose) were implanted subcutaneously in the subscapular region of C57/BL6 mice for 14 days. When compared to the vehicle treated mice, the ABT-510 group had a 20% decrease in capillarity in the superficial region of the gastrocnemius (GA), 11% decrease in the plantaris (PLT), and a 35% decrease in the soleus (SOL). ABT-510 also decreased muscle protein expression of vascular endothelial growth factor (VEGF) in both the GA (-140%) and SOL (-62%); however there was no change in VEGF in the PLT. Serum VEGF was not altered in ABT-510 treated animals. Endogenous TSP-1 protein expression in all muscles remained unaltered. Tunnel staining revealed no difference in muscle apoptosis between ABT-510 and vehicle treated groups. These data provide evidence that the anti-angiogenic effects of TSP-1 are mediated, at least in part, via the CD36 receptor. It also suggests that under physiologic conditions the TSP-1/CD36 axis plays a role in regulating basal skeletal muscle microvessel density.

Figure 1. Representative figures of the histology sections.

A) Superficial Gastrocnemius Muscle B) Deep Gastrocnemius Muscle C) Plantaris Muscle, D) Soleus Muscle.

Figure 2. Superficial gastrocnemius capillarity, but not deep, is decreased in the ABT-510 group.

Chronic administration of ABT-510 30 mg/kg/day decreased capillary to fiber ratio (C∶F) and capillary density (CD) in the superficial gastrocnemius (GA). Despite a similar trend, no significant changes in capillarity were seen in the deep GA. There was no change in fiber cross sectional area in either the deep or superficial portions of the GA. * = P≤0.05.

Figure 3. Plantaris and soleus capillarity is decreased in the ABT-510 group.

Chronic administration of ABT-510 30 mg/kg/day decreased capillary to fiber ratio (C∶F) in the plantaris muscle (PLT) but did not decrease capillary density (CD). ABT-510 decreased capillary to fiber ratio (C∶F) and capillary density (CD) in the soleus (SOL). There was no change in fiber cross sectional area in either the PLT or SOL muscle. * = P≤0.05.

Figure 4. Skeletal muscle VEGF protein content is decreased in the ABT-510 group, but not serum levels.

Chronic administration of the mimetic (ABT-510 30 mg/kg/day) decreased VEGF protein in the GA, and SOL muscles of C57/BLK6 mice. There was no change in the PLT muscle or in Serum levels. * = P≤0.05.

Figure 5. There is no difference in skeletal muscle apoptosis between the mimetic and control groups.

TUNEL staining shows that chronic administration of the mimetic (ABT-510 30 mg/kg/day) did not increase or decrease skeletal muscle apoptosis in any of the muscles tested. Representative GA samples, PLT and SOL data not shown. A) Representative mimetic GA B) Representative positive control mimetic GA C) Representative control GA D) Representative positive control vehicle GA.