# Major Royal Jelly Proteins promote C2C12 myotubes differentiation by improving mitochondrial function

## TL;DR

This study shows that Major Royal Jelly Proteins (MRJPs) help muscle cells develop by improving mitochondrial function, but they cannot reverse muscle atrophy caused by dexamethasone.

## Contribution

The study reveals a novel role of MRJPs in promoting muscle cell differentiation through mitochondrial dynamics and biogenesis.

## Key findings

- MRJPs increased myotube density and length in C2C12 cells.
- MRJPs stimulated mitochondrial biogenesis and mitophagy while reducing apoptosis.
- MRJPs could not reverse dexamethasone-induced muscle atrophy.

## Abstract

Royal Jelly (RJ) has been widely used as a health-promoting supplement and its bioactive component is the Major Royal Jelly Proteins (MRJPs). Whether MRJPs promote skeletal-muscle-cell development remains unresolved. Muscle dysfunction is linked to mitochondrial depletion and protein breakdown. Thus, we evaluated how MRJPs affect myotube differentiation. Myotubes morphology and number were measured using fluorescence microscopy and Coomassie brilliant blue staining, and C2C12 myotube differentiation was assessed using Western blotting or qRT-PCR analysis of the expression of MyoD, MyoG, Myosin Heavy Chain (MyHC) and muscle ring finger 1 (MuRF-1). Mitochondrial function was assessed with fluorescent probes, whereas the content of mitochondria was determined by analyzing the expression of related proteins. Western blotting was used to examine the expression of myosin-associated proteins, autophagy-associated proteins, apoptosis-associated proteins, and mitochondria-associated proteins. This was demonstrated by increased myotubes density and length, and increased mRNA and proteins expression of MyoD, MyoG and MyHC. In this study, we found that MRJPs promoted the differentiation of C2C12 myoblasts to form myotubes, but could not reverse Dex-induced muscle atrophy. The possible mechanism is that MRJPs reduced apoptosis increased cellular autophagy, stimulated mitochondrial biogenesis, promoted mitochondrial dynamics homeostasis and mitophagy, and prevented the loss of mitochondrial membrane potential.

Schematic representation of the molecular mechanism of MRJPs in the induced differentiation of C2C12 myotubes. Some of the pieces in the figure were created with Servier Medical Art, and some were created with PowerPoint.Diagram illustrating the effects of major royal jelly proteins on myotubes in C2C12 cells. It shows royal jelly extraction, intervention, and the resulting increase in myotube density and length. Three main processes are highlighted: mitochondrial dynamics involving DRP1 and MFN2, mitochondrial biogenesis and uncoupling with PGC-1α and UCP-1, and mitophagy involving proteins like PINK1 and Parkin. MyHC, MyoD, and MyoG are indicated in the myotubes. Connected steps are marked with dotted arrows and positive effects are denoted by red arrows. Various cellular components are symbolized with corresponding icons.

Schematic representation of the molecular mechanism of MRJPs in the induced differentiation of C2C12 myotubes. Some of the pieces in the figure were created with Servier Medical Art, and some were created with PowerPoint.