23 Radial Growth of Muscle Fibres

23.1 Introduction

• The radial growth of myofibres is an example of cellular hypertrophy - the myofibre (a giant cell) is growing in size.
• But when the myofibre undergoes hypertrophy - it also increases its number of myofibrils.
• Thus, myofibre hypertrophy is accompanied by myofibrillar hyperplasia (increase in number - not size).
• Longitudinal growth of myofibres differs from radial growth (although both are examples of cellular hypertrophy), and is examined in the next lecture.

23.2 Myofibre diameters and myofibrillar hyperplasia

• As a myofibre grows in diameter, there is an increase in the number of myofibrils.
• At one time, it was  suggested myofibre diameters might be used to measure muscularity in meat animals, but hopes of this faded rapidly once the complexity of the interactions between myofibre number, length and diameter became known.
  
• Myofibre  diameters are negatively correlated  with the tenderness of cooked meat, but this may not be a direct relationship.  For example, as animals grow older, their myofibres increase in diameter, but they also develop stronger and more abundant connective tissue. Thus, the real relationship may be between connective tissue and tenderness.
  
• In young animals, fast-contracting myofibres usually have more rapid radial growth than slow-contracting myofibres. 
  
• In older animals, radial growth of fast-contracting myofibres slows down, but radial growth of slow-contracting myofibres continues.  This is because the slow myofibres are primarily involved in the maintenance of body posture against gravity - and the animals are now getting very heavy.
• All other things being comparable, animals on a low plane of nutrition will have smaller myofibre diameters than animals on a high plane of nutrition.
• All other things being comparable, entire males will have larger diameter myofibres than females.  This may be detectable to the consumer as coarse meat texture.
• Measurement of radial growth is difficult because myofibres are not really cylindrical - they tend to be prismatic - with flat sides where they are pressed together.  Where would you measure the diameter of the myofibre shown above?
• Another problem is myofibres are often tapered - their diameters may decrease towards one or both ends of the myofibre.
• And, of course, myofibre diameters increase when a myofibres contracts!  Seldom do researchers correct for differences in sarcomere length.

1. New myofilaments are added around the outside of the myofibril.
2. This increases the length of the diffusion pathway for calcium ions as they turn contraction on and off. (Remember - calcium ions are released from the sarcoplasmic reticulum to initiate muscle contraction, then re-sequestered for relaxation).
   
3. This causes mechanical stress in the myofibril - the outside contracts and relaxes before the central axis.
4. Calcium ions building up in the interior of the myofibril activate enzymes (calpains) which release thin myofilaments from their Z-lines.
5. Contraction causes the weakened myofibril to split.
6. But, the myofibril is very long, and a split at one point may not match up to a split at another point.
7. We end up with a complex structure.  Where once there was a single myofibril seen in transverse section, we now see many myofibrils in transverse section - but following them along the myofibril we find they are all linked.
   
8. Thus, we have not really formed any new myofibrils, just increased their size and complexity.
9. But, it appears we have formed new myofibrils, so we will keep on talking about their numbers!

To keep things simple in the diagram above - we are looking at transverse sections at the midlength of relaxed sarcomeres where we will only see thick myofilaments. The new new myofilaments are shown in green. You can see how the progressive addition of new myofilaments will increase the length of the pathway by which calcium ions move in and out of the myofibril.

23.3 New myofibre nuclei

As the myofibre grows radially, it adds new nuclei.

• It is important to remember when we look at transverse sections of myofibres - the number of nuclei seen under the sarcolemma (the cell membrane) depends on the thickness of the transverse section relative to the length of the nuclei.  If the nuclei are long and thin, then they are more likely to be seen in transverse section than short, thick nuclei.  The thicker a transverse section, the more nuclei will be seen.
• Even taking this into account, however, there is no doubt the number of nuclei increases as myofibres undergo radial hypertrophy.
• Where do the extra nuclei come from? This was a puzzle for a hundred years.  New nuclei are normally obtained from mitosis (cell division), and mitosis is normally detectable when chromosomes separate at metaphase - looking rather like bunches of bananas. But, in a hundred years, nobody ever saw any  metaphase chromosomes in a myofibre.

• (1) No clear zone separates them from myofibrils.
• (2) They are more basophilic (stained by basic dyes for light microscopy) than fibroblast nuclei.
• (3) The nucleolus (containing RNA) is well defined and large.
• (4) The outer chromatin (the granular pattern of stained DNA) is tightly distributed along the nuclear membrane.
• (5) The internal cromatin is evenly dispersed.

Myofibre nuclei contain DNA combined with histones and other structural proteins to form chromatin. When DNA is used for protein synthesis, the chromatin is dispersed, only binds weakly to histological stains, and is called euchromatin. In non‑dividing cells, chromatin may form darkly stained irregular clumps called chromatin particles. Nuclei also contain RNA and darkly stained clumps of RNA form nucleoli. The number of nucleoli may vary between animal species. Condensed regions of darkly stained chromosomes sometimes persist between cell divisions and are called heterochromatin. In the mononucleated cells of the body, such as those of the skin or liver, darkly stained chromosomes composed of inactive DNA are seen when cells divide. But, as explained below, the situation in multinucleated myofibres is more complex, and distinct chromosomes are not seen by light microscopy within myofibres.

On or near the myofibre membrane are several types of nuclei.

• True myofibre nuclei are located within the sarcoplasm.
  
• Although the nuclei of satellite cells are seen by light microscopy within the sarcoplasm, electron microscopy shows satellite cells are located in depressions in the myofibre surface.
  
• Thus, the nucleus of a satellite cell is separated from the sarcoplasm of its myofibre by a satellite cell membrane and a myofibre membrane.

The red line above is where the TWO membranes are located - one around the satellite cell and one lining the depression on the myofibre surface.

The existence of satellite cells became accepted in 1961, although they had been seen a hundred years earlier - but no one believed it. Pericytes are mesodermal cells found around very small blood vessels. They contain actomyosin and are probably capable of contraction and phagocytosis.

• (1) They  are indented into a myofibre surface, although sometimes they bulge outwards.
• (2) They have variable amounts of basophilic cytoplasm - usually very little.
• (3) Their outer chromatin is heavily and unevenly deposited along the nuclear membrane.
• (4) Their internal chromatin is scattered in clumps.
• (5) Their nucleolus is small and usually masked by internal chromatin - because mitosis is still possible.
• (6) Sometimes the nuclei are uniformly stained dark.
• (7) Most of them have a clear space of 0.5 to 0.2 micrometres  separating them from the myofibrils - this is where the two membranes are located.

• Muscle nuclei increase in number during postnatal development but the relative magnitude of the increase varies from muscle to muscle.
  
• Before the existence of satellite cells was proved by electron microscopy, it was rather difficult to explain how muscle nuclei were able to increase in number without any evidence, by light microscopy, of mitosis in myofibre nuclei.
  
• Myofibre nuclei are able to withstand longitudinal compression during muscle contraction by means of concertina‑like wrinkles in their nuclear membrane and, for many years, histologists regarded wrinkled muscle nuclei as evidence of an unusual type of nuclear division. They were wrong!
• New nuclei added during the postnatal growth of myofibres come from the daughter cells of satellite cell mitosis.
• Like myoblasts, satellite cells are part of the myogenic cell lineage from somitic mesoderm - essentially - they are premyoblasts which have been trapped on the myofibre surface by the development of the endomysium (the connective tissues around each myofibre).
•  Mitosis in satellite cells has been seen by electron microscopy and the synthesis of DNA by satellite cell nuclei has been proved by radioautography (tritiated thymidine used in the synthesis of new DNA is detected by its radioactivity).
• If a myofibre is damaged by disease, its myofibrils are removed by phagocytic cells - then a new myofibre may be formed from satellite cells within the old endomysial tube.
• There is some evidence satellite cells can move around on the myofibre surface.  Can you think how this might be exploited agriculturally?
  
• On a proportional basis, there are more nuclei in red muscles than in white muscles, and nuclei are more frequent at the ends of myofibres than at their midlength.
• Younger animals have proportionally more satellite cells than older animals.

Further information