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Orlando Paciello » 5.Specific alterations to muscle fibres


Specific alterations to muscle fibres

  • Muscle lesions can often be seen and assessed using only a microscope.
  • A proper microscope examination involves looking at transverse and longitudinal sections. The diameter of the fibres, changes to the cell architecture and the percentage of abnormal fibres are all easier to see in transverse sections.
  • Routine staining tests like hematoxylin-eosin (H&E) are not adequate for a complete assessment of myopathic lesions, intramuscular nerve lesions or for the presence of abnormal accumulations of substances.

Pathology of muscle tissue

  • Neuropathic myopathies (neuromyopathies) resulting from effects or absence of innervation.
  • Myopathic myopathies (real myopathies) caused by alterations at the primary site of muscle fibres (e.g. necrosis from nutritional deficit).

Specific pathological alterations – Neuropathies

  • Angular atrophy
  • Pyknotic nuclear clumps
  • Compensatory hypertrophy
  • Central nuclei
  • Fibre target
  • Muscle atrophy in small or large groups
  • Fibre type grouping
  • Predominance of fibre type
  • Pathologies of intramuscular nerve (axon/myelin degeneration, loss of myelinated axons)

Specific pathological alterations – Myopathies

  • Non-angular atrophy
  • Myonecrosis and cellular infiltrate
  • Compensatory hypertrophy
  • Nuclei in central position
  • Regen/degeneration of fibres
  • Endomysial fibrosis
  • Vacuolization of muscle fibres
  • Degenerative
  • Accumulations (glycogen, lipids, etc.)

Alterations in muscle fibre


Denervation

  • Loss of any nerve impulse quickly results in atrophy of the muscle and, without innervation, practically half the muscle mass can be lost in the space of a few weeks.
  • Generalized neuropathies or neuronopathies like equine motor neurone disease lead to widespread, symmetrical atrophy. Where there is damage to a particular nerve then asymmetrical atrophy is observed.
  • One example is equine laryngeal hemiplegia secondary to a recurrent left laryngeal nerve lesion.

Img.2 Diagram illustrating denervation in muscle fibres. The fibres appear uniformly atrophied. Source: modified from Anderson J.R. Atlas pf Skeletal Muscle pathology MTP Press Limited

Img.1 Diagram illustrating denervation and re-innervation of muscle fibres. Source: modified from Anderson J.R. Img.1 Atlas pf Skeletal Muscle pathology MTP Press Limited

Img.1 Diagram illustrating denervation and re-innervation of muscle fibres. Source: modified from Anderson J.R. Img.1 Atlas pf Skeletal Muscle pathology MTP Press Limited

Img.2

Img.2


Neuropathic myopathies – morphology

Atrophic myofibres

Enervation leads to a particular type of atrophy, called pointed or angular (angular atrophy). In transverse section, the atrophic myofibres present in various bundles appear small and spiky with rounded corners (angular myofibres), which is because fibres near the atrophic ones become hypertrophic and so compress them.

Angular atrophic fibres are easily visible with NADH histoenzymatic staining (arrow)

Angular atrophic fibres are easily visible with NADH histoenzymatic staining (arrow)


Neuropathic myopathies – morphology (cont.)

Small group atrophy

Progression of preceding lesion whereby the number of atrophic angular myofibres increases and they tend to form small clusters.

Example of small-group atrophy (hematoxylin-eosin 40x)

Example of small-group atrophy (hematoxylin-eosin 40x)


Fiber-type Grouping

Denervated myofibres (A-C) can be reinnervated (D) by collateral nerves thus the fibres acquire the characteristics of the new motor unit (D).

Many angular atrophic fibres can be seen as well as different ATPase at PH 9.4 fibre type grouping

Many angular atrophic fibres can be seen as well as different ATPase at PH 9.4 fibre type grouping

Diagram illustrating denervation of muscle fibres and reinnervation with formation of fibre-type groupings. Source: modified from Anderson J.R. Atlas pf Skeletal Muscle pathology MTP Press

Diagram illustrating denervation of muscle fibres and reinnervation with formation of fibre-type groupings. Source: modified from Anderson J.R. Atlas pf Skeletal Muscle pathology MTP Press


Pyknotic Nuclear Clumps

If a denervated fibre is not reinnervated, atrophy continues with loss of myofibrils so that only the myonuclei remain in the sarcolemma.

This is the last stage in chronic denervation.

Numerous atrophic angular fibres and pyknotic-nuclear clumps (arrows). Hematoxylin eosin stain

Numerous atrophic angular fibres and pyknotic-nuclear clumps (arrows). Hematoxylin eosin stain


Big-group atrophy

When all the myofibres, or at least the majority, are enervated at the same time it gives rise to widespread, simultaneous atrophy, so the fibres do not assume their characteristic angular form but appear uniform and rounded in transverse section. In chronic states it is possible to find target fibres.

Large groups of atrophic fibres and an increase in endomysial connective tissue (asterix). Hematoxylin esoin stain

Large groups of atrophic fibres and an increase in endomysial connective tissue (asterix). Hematoxylin esoin stain


Target fibres

  • Often observed in cases of acute denervation and reinnervation.
  • Clearly seen with NADH stain. Characterized by a light or dark centre with a light halo round it and hyperactivity apparent in the area adjacent to the rest of the normal fibre.
A target fibre can be seen in the centre of the picture. NADH histoenzymatic stain

A target fibre can be seen in the centre of the picture. NADH histoenzymatic stain


Unusual alterations in myofibres

  • Ragged Red Fibres
  • Nemalinic myopathy
  • Myopathy with Central Core Disease
  • Target fibres)
  • Myotubular myopathy
  • Splitting
  • Predominance or lack of one type of myofibre

Ragged Red Fibres

Vacuoles and aggregates of substances can be seen in many muscle pathologies. Vacuoles may contain lipids, glycogen or the products of degradation. Accumulation of abnormal mitochondria can be seen with Gomori’s trichrome.

Examples of RRFs. These fibres look ragged and are characterised by deposits of red-coloured material. The stain of choice for observation of these fibres is Engel’s trichrome

Examples of RRFs. These fibres look ragged and are characterised by deposits of red-coloured material. The stain of choice for observation of these fibres is Engel's trichrome


Ragged-Red Fibres


- Mitochondrial myopathies

RRFs are typical of mitochondrial myopathies and are often associated with “Ragged Blue Fibers” which can be seen with SDH and with COX negative fibres
SDH and COX are both purely mitochondrial histoenzymatic stains.

RRFs and RBFs will be negative to the cytochrome oxidase stain (COX) when the abnormal proliferation of the mitochondria is associated with malfunctioning of these organelles.


Myopathy with enzymatic hole in the middle

This type presents with areas where enzyme activity is absent and are typical of Central Core Disease.

Central core disease

Central core disease


Splitting

This is a normal way for muscle fibres to divide but it can also occur in muscle pathologies like muscular dystrophy where the division is incomplete, along only part of the length of the fibre. Splitting is a process of regeneration and differentiation starting from a single fibre but it can also be a process of subdivision of hypertrophic myofibres.

Splitting phenomena

Splitting phenomena


Internal nuclei

The nuclei in a myofibre are normally located sub-sarcolemma. Migration towards the centre is a non-specific response.
Internal nuclei can be observed in myotonia and muscular dystrophy.

Numerous nuclei moved into central position

Numerous nuclei moved into central position


Increase in connective

Connective and adipose tissue substitute the muscle after necrosis.
In figure 1: Normal muscle. H.E.stain.

In figure 2: Serious increase in endomysial connective tissue in a case of muscular dystrophy (asterisk). H.E.stain


Necrosis

Necrosis of a muscle usually occurs as partial or segmental necrosis and only involves the sarcoplasm. Necrosis involving the sarcolemma as well is less common but it spreads to many fibres (total necrosis).

When the muscle fibre is destroyed, certain enzymes are released which are traceable in the blood:

  • Creatinphosphokinase (CPK);
  • Glutamic oxaloacetic transaminase (SGOT);
  • Myoglobin (excreted in urine–myoglobinuria).

Segmental (hyaline) necrosis (Zenker degeneration)

This is a process of coagulation necrosis involving the sarcoplasm but not the sarcolemma so regeneration is possible.
There are different degrees of this type of necrosis:

  1. Necrosis of the myofibrils (rhabdomyolysis)
  2. Necrosis of the myofibrils and sarcoplasm
  3. Necrosis which only leaves nuclei of satellite cells and basal lamina intact.
  4. Necrosis involving the sarcolemma (e.g. ischemia) – Regeneration is not possible.

Segmental necrosis (Zenker degeneration)

  • Histologically, myofibres in necrosis can have very different appearances. The earliest lesions show segmental hypercontraction, where segments are slightly larger in diameter and slightly darker in colour (A).
  • The cytoplasm in really necrotic fibres is often homogeneously oesinophilic and pale-coloured (hyaline degeneration) with loss of cytoplasmic striations and adjacent nuclei.
  • The increase in intra-cellular calcium is the common trigger for necrosis in all cells, and myofibres contain large amounts of calcium ions in the sarcoplasmic reticulum. This is why muscle fibres can be very sensitive to calcium-induced necrosis either when the sarcolemma is damaged leading to an influx of extra-cellular calcium, or when the sarcoplasm is damaged leading to the release of intra-cellular calcium deposits (B).
  • Macrophages resulting from the transformation of monocytes in the blood quickly infiltrate the site of necrosis (sarcoclastosis [C]).

Regeneration

Skeletal muscle generally has good regenerative capacity though there are a few exceptions. These regenerative capacities, however, can only work and the muscle be properly restored if the endomysial stroma and the basal lamina are intact.
Regeneration occurs in one of two possible ways depending on whether the sarcolemma is affected or not.

Regeneration by gemmation

After a few hours the sarcolemma nuclei start to multiply where the stumps are and appear surrounded by newly-formed sarcoplasma which emerges from each stump like a club-shaped bud. These grow into the young connective tissue which has continually filled the solution.

Regeneration of muscle fibre with centralisation of nucleus. (H.E. stain 40X)

Regeneration of muscle fibre with centralisation of nucleus. (H.E. stain 40X)


Regeneration through myoblast profileration

This type of regeneration occurs when the satellite cells are activated as myoblasts capable of synthesising new sarcoplasma within the framework provided by the sarcolemma sheath. This process involves intense protein synthesis and the regenerating myofibres are thus recognizable because of their slender, fusiform, nastriform shape and because of the basophilia (regenerating basophile myofibres). The nuclei in the regenerating myofibres are more numerous, larger and have prominent nucleoli.

Immunohistochemistry

  • Histochemical tests performed on non-fixed frozen sections of muscle represent the “gold standard” for muscle pathology.
  • But we often turn to immunohistochemical tests for the study of cytoskeleton proteins, like dystrophyn and dystrophin-associated proteins.
Sarcolemma is positive to anti-dystrophin antibodies (COOH) (immunohistochemical stain, avidyne biodyne oxidase method. 40X)

Sarcolemma is positive to anti-dystrophin antibodies (COOH) (immunohistochemical stain, avidyne biodyne oxidase method. 40X)


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