Prostaglandin E2: from clinical applications to its potential role in bone- muscle crosstalk and myogenic differentiation

Abstract

Prostaglandin E(2) (PGE(2)), a prostanoid synthesized from arachidonic acid via the cyclooxygenase pathway, is a modulator of physiological responses including inflammation, fever, and muscle regeneration. Several patents have been filed that are related to PGE(2), one of them being directly related to skeletal muscles. In this report, we first summarize the key patents describing inventions for the utilization of PGE(2) for either diagnostic or therapeutic purposes, including skeletal muscle. In the second part of our work we present new and exciting data that demonstrates that PGE(2) accelerates skeletal muscle myogenic differentiation. Our discovery resulted from our recent and novel concept of bone-muscle crosstalk. Bone and muscle are anatomically intimate endocrine organs and we aimed to determine whether this anatomical intimacy also translates into a biochemical communication from bone cells to muscle cells at the in vitro level. The effects of MLOY4 osteocyte-like cell conditioned medium (CM) and three osteocyte-secreted factors, PGE(2), sclerostin and monocyte chemotactic protein (MCP-3), on C2C12 myogenic differentiation were evaluated using morphological analyses, a customized 96-gene PCR array, and measurements of intracellular calcium levels. MLO-Y4 CM and PGE(2), but not sclerostin and MCP-3, induced acceleration of myogenesis of C2C12 myoblasts that was linked with significant modifications in intracellular calcium homeostasis. This finding should further stimulate the pursuit of new patents to explore the use of PGE(2) and the new concept of bone-muscle crosstalk for the development and application of inventions designed to treat muscle diseases characterized by enhanced muscle wasting, such as sarcopenia.

Figure 1. Histological evidence for the intimate association of bone-circulation-muscle

Mineralized bone is shown in light green. The osteocytes stained in dark blue are the major cell component of mature bone. Vessels can be seen in between the bone and muscle interface. Given the overwhelming proof of tissues' crosstalk it seems feasible to infer that bone and muscle communicate biochemically either through paracrine or endocrine loops.

Figure 2. MLO-Y4 CM and PGE₂ significantly enhance myogenic differentiation of C2C12 myoblasts

(A) Representative pictures of treatments taken 3 days after onset of differentiation. a) control; b) 25% MLO-Y4 CM; c) 50nM PGE₂; d) 2μg/ml sclerostin; e) 150μg/ml MCP-3. (B) myotube area measurement shows that CM and PGE₂, but not sclerostin and MCP-3 treatments significantly increase myotube area. *, significant different from control (P<0.05).

Figure 3. Real time-PCR results of 91-gene array

CM and PGE₂ uprgulate the expression of myogenic regulatory factors, and genes related with Ca²⁺ homeostasis and mitochondria biogenesis in C2C12 myoblasts. Two arrays yielded similar results.

Figure 4. Modulation of Ca²⁺ homeostasis in myotubes by osteocyte signaling

18-30 myotubes were tested for each condition. Arrows indicate addition of 20mM caffeine. While a single peak calcium transient is observed in response to caffeine under control conditions, multiple peaks are observed in cells treated with CM and PGE₂.