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Stem Cell Therapy for Muscular Dystrophy

Rather than referring to one specific disease, the term “muscular dystrophy” (MD) describes a group of more than 30 diseases of similar manifestation and genetic origin. Commonly characterized by progressive degeneration, weakness and loss of control of musculoskeletal motion, these dystrophies represent a rare group of genetic disorders. The various types of MD differ according to the age of onset, the extent and distribution of muscle weakness, the rate of progression of the disease, and the pattern of inheritance.

In MD, muscle fibers exhibit an abnormal susceptibility to damage, which results in a progressive weakening and atrophy over time of the voluntary muscles throughout the body, which are ultimately replaced by fat and connective tissue. In some forms of MD, involuntary muscles and organs may also be affected, including possibly the cardiovascular system, the gastrointestinal system, the neurological system, the endocrine glands, and the eyes. Respiratory diseases are especially common in MD, and some patients may develop difficulty swallowing. The most common forms of MD are found only in males, especially in the early years of life, although females may be genetic carriers of the disease without exhibiting symptoms. MD is often fatal.

According the National Institute of Neurological Disorders and Stroke (NINDS), a division of the National Institutes of Health (NIH),

“Muscular dystrophy (MD) refers to a group of more than 30 inherited diseases that cause muscle weakness and muscle loss. Some forms of MD appear in infancy or childhood, while others may not appear until middle age or later. The different muscular dystrophies vary in who they affect and the symptoms. All forms of MD grow worse as the person’s muscles get weaker. Most people with MD eventually lose the ability to walk.”

“There is no cure for muscular dystrophy. Treatments include physical and speech therapy, orthopedic devices, surgery and medications. Some people with muscular dystrophy have mild cases that worsen slowly. Others cases are disabling and severe.”

(From the website of the National Library of Medicine, at www.nlm.nih.gov).

The most common forms of MD have been found to be caused by a genetically induced deficiency of dystrophin, which is a protein involved in maintaining the integrity of muscle fiber.

Muscular dystrophy falls under the category of “single gene disorders and disabilities”. Single gene disorders (SGDs) are a group of conditions caused by a mutation in one particular gene. Over 6,000 SGDs have been identified, and although the frequency of occurrence of any one particular SGD is rare, when grouped together all the SGDs combined account for approximately one in 300 births.

In its various forms muscular dystrophy is also known as pseudohypertrophic myopathy and dystrophinopathy.

Background:

The word “dystrophy” is derived from two Greek words: the prefix “dys”, meaning “abnormal”, “impaired” or “difficult, and “trophe”, meaning “nutrition” or “growth”. In general, therefore, “dystrophy” implies inadequate growth or nutrition, with “muscular dystrophy” referring specifically to the group of diseases that are characterized by muscular degeneration and which are now known to be of genetic origin.

The first case of muscular dystrophy was documented in 1830 when Sir Charles Bell published an essay in which he described a progressive muscular weakness that he had observed in some young boys. Shortly thereafter, further historical accounts of MD were documented as other scientists noted the same symptoms among brothers. Although such symptoms were initially assumed to be indications of tuberculosis, successive reports were published in the following years which described not only progressive muscular weakness but also premature death in those who exhibited the symptoms. Documentation of such maladies became increasingly prominent in the medical journals, and by the 1850s the French neurologist Guillaume Duchenne had recorded a comprehensive account of 13 boys who were observed to have the most common and severe form of the disease, which now bears the name of its discoverer. Increasing evidence quickly led physicians to the conclusion that this mysterious illness had more than one form and could strike people of either gender and all ages, although it was also determined that the disease is not contagious and is not caused by injury or activity.

Several genes have now been identified for some forms of MD, although there are other forms for which the underlying genetic cause has yet to be isolated. While MD is most prominent in boys, it is found throughout the world, occurring in all races. Since some forms of the disease are more common than others, precise epidemiological records that distinguish between the various types of MD do not exist in all countries. The most extensive data correspond to the most common forms of MD, namely, the Duchenne and Becker forms, which are estimated to affect approximately 1 in every 3,500 to 5,000 boys, and which in the U.S. would constitute between 400 and 600 live male births per year.

Types and Symptoms:

Before a conclusive diagnosis of MD may be made, other diseases with similar symptoms must be ruled out. Since there are many diseases that affect skeletomusclature and the neurological system, with symptoms that may be very similar to those of MD, a thorough and precise evaluation of the patient must be made. For example, inflammatory myopathy, muscle atrophy and even cardiomyopathy, among other symptoms, may result from a variety of conditions, both genetic and nongenetic, and such disorders do not necessarily imply MD nor any other single disease. Indeed, because of the necessity for thorough and specific medical testing, MD is difficult to diagnose quickly. Accurate means of testing do exist, however, and only by employing such tests can a reliable diagnosis be made.

Of the more than 30 types of MD, nine categories have been designated for the most distinct forms. These nine variations of the disease are classified according to several variables, such as the extent and distribution of muscle weakness, the age of onset, the rate of progression of the disease, the severity of symptoms, and the particular pattern of inheritance that is recognizable within the family. Although differences exist between each of these nine dystrophies, they are all characterized by progressive muscular atrophy. They are named either according to their respective discoverers, or descriptively by their most prominent features. Some of the categories are further subdivided into additional classifications. Symptoms and subdivisions of these nine primary types of MD are listed herein.

Of these nine basic categories of MD, four are most characteristically associated with childhood, while the other forms may strike later in life. Those variations of this disease that result from genetic mutations on the X chromosome occur almost exclusively in males, although females may be carriers of the defective genes without expressing symptoms.

1. Duchenne muscular dystrophy (DMD):

Duchenne MD is the most common form and the most severe form of MD, accounting for approximately 50% of all cases.

DMD is known to occur in approximately one in every 3,500 male births. Currently DMD affects approximately 8,000 boys and young men in the United States. A milder form that occurs in female carriers is much less frequent in occurrence. In males, the disease is typically fatal at an early age, while in females it is much less severe and rarely fatal.

This particular form of MD is caused by a mutation on the X chromosome (Xp21) and is therefore inherited in an X-linked recessive manner. The X chromosome contains 2.5 million base pairs, and the gene implicated in DMD consists of a sequence of base pairs that are known to produce dystrophin, a mutation in which results in an absence of dystrophin. The protein dystrophin is known to be an integral component of the membrane cytoskeleton and is also suspected of playing a role in maintaining plasma membrane stability during muscle contraction or stretching. It is believed that the loss of dystrophin may lead to plasma membrane injury and a subsequent cascading of events that ultimately result in muscle cell necrosis. In healthy individuals, muscles are normally able to rebuild themselves by anabolic metabolism whenever subjected to catabolic breakdown and daily wear and tear, but muscles are unable to rebuild themselves in people with MD.

The specific gene on which the DMD mutation occurs was discovered in 1986, and further research has since shown that dystrophin also binds to other proteins at the edge of muscle fibers and probably plays a role in “anchoring” muscles to the adjoining connective tissue.

Duchenne is one of several forms of MD which are known to result from an absence of dystrophin, although all forms of MD are believed to result from a genetic defect in protein metabolism. Approximately one-third of all cases of DMD represent new mutations, not inherited from either parent, while the remaining two-thirds follow a familial pattern of inheritance.

Blood tests of children with DMD show an abnormally high level of creatine kinase (CK), an enzyme that leaks out of damaged muscle and which is measurable at birth. In the absence of newborn screening, however, DMD is usually first noticeable when the affected child begins to walk, and the disease may not be officially diagnosed until the child is between 3 to 6 years of age.

The degenerative muscle weakness characterized by DMD is first apparent in the upper legs and pelvis, spreading later to the upper arms and the rest of the body as the disease progresses. Pseudohypertrophy is common in the initial stages, where certain muscles, especially the calf muscles, become enlarged due to an accumulation of fat and connective tissue, which may give a deceptively healthy appearance to degenerating muscle. Contractures are another hallmark of DMD, in which muscles become permanently tightened, most commonly in the calf muscles. Such a tightening only exacerbates the progressive muscle atrophy, compounding problems with movement and balance. Premature death is usually inevitable.

Early symptoms are characterized by problems with walking and balance, which may also involve a loss of some reflexes. In particular, physicians may observe the absence of deep tendon reflexes in the upper extremities and knees even in the early stages of DMD, whereas ankle reflexes may remain detectable until the terminal stages. Associated symptoms may include delayed walking, frequent falling, a “waddling” gait, clumsiness, difficulty running, difficulty when rising from a sitting or lying position, difficulty climbing stairs, and changes to overall posture. General diaphragm weakness in DMD is a serious problem which often makes breathing, coughing and swallowing difficult while simultaneously increasing susceptibility to lung infections. Scoliosis (a side-to-side curvature of the spine) may also appear at intermediate stages in DMD patients, along with osteoporosis (bone thinning) and cardiomyopathy, which is a pathological, and usually irreversible, condition of the heart muscle. Many children with DMD are unable to run or jump, and most are unable to walk by the age of 12, at which point a wheelchair is commonly needed, although orthopedic devices and physical therapy may prolong the ability to walk. Approximately one-third of affected boys with DMD exhibit a “nonprogressive cognitive deficit”, i.e., mental retardation and learning disabilities. DMD is often fatal by the late teens or early 20s, due usually to severe respiratory problems, congestive heart failure, infections, or some combination thereof. Some patients with DMD, however, are able to live beyond their 20s with the aid of a ventilator. While antibiotics are usually successful in treating respiratory infections in the early stages of the disease, the increasing severity and continued recurrence of such infections often make treatment more and more difficult as the disease progresses.

In families in which this particular mutated gene is present, there is a 50% probability that the gene may be inherited by females as well as by males, in which case the females will then become carriers of the gene, and may or may not exhibit any symptoms themselves. If they do exhibit symptoms, such symptoms are usually mild by comparison to those expressed in males, and the disease is rarely fatal in females.

In the Middle East and North Africa, a rare autosomal recessive form of MD has been found which is clinically similar to Duchenne but is less severe and progresses more slowly. In this particular form of the disease, the onset of muscle weakness typically occurs between the ages of 5 and 10, with most individuals losing the ability to walk in their early twenties, and death from cardiac or respiratory complications may be delayed until the age of 40.

A very similar but milder form of DMD is known as Becker muscular dystrophy (BMD).

2. Becker muscular dystrophy (BMD):

BMD occurs in approximately one in every 30,000 live male births, and mainly afflicts older boys and young men. It is similar to Duchenne MD but is slightly less severe. Whereas DMD is characterized by a virtual absence of dystrophin, BMD is characterized by the presence of some dystrophin, though it is not enough to prevent the atrophy of muscles. The age of onset is slightly later, the rate of progression is slightly slower and life expectancy is slightly longer in Becker than in Duchenne. The typically age of onset for Becker MD is between the ages of 11 and 25, and it is not uncommon for BMD patients to live into middle age. The exact rate of progression of the disease varies widely among individuals, with some patients maintaining their ability to walk into their 30s or later, while others are unable to walk past their teens. Early symptoms of BMD are similar to those of Duchenne, although mental impairment and cardiac complications are not as severe in Becker as in Duchenne MD. The overall severity of Becker MD varies, depending on the amount of dystrophin that is present in the body.

Becker MD was originally believed to be an entirely separate disease, unrelated to MD, until it was discovered that defects in the same dystrophin gene were responsible for both Becker and Duchenne MD.

As with Duchenne, respiratory problems often become severe in Becker MD, with associated cardiovascular complications. Arrhythmias (irregular heartbeats), fatigue, shortness of breath, chest pain, dizziness and ultimately congestive heart failure are common, although patients are able to achieve some degree of improvement in their breathing with the help of a ventilator.

In clinical evaluations, the symptoms of Becker MD are often combined with those of Duchenne MD to form the “spectrum” known as Duchenne/Becker muscular dystrophy (DBMD).

3. Congenital MD (CMD):

This is a rare form of MD, present from birth. Infants who are born with CMD have severe muscle weakness, with very little muscle tone and a sharply reduced ability for voluntary movement. With careful attention to physical therapy, however, some children with CMD are able to learn to walk, and some live into young adulthood or beyond.

A further subclassification within CMD is Fukuyama CMD, which is also congenital, involves a severe weakness of the facial and limb muscles, and is seen almost exclusively in Japan. Children with Fukuyama CMD are rarely able to walk and have severe mental retardation. Seizures are often part of the disease and must be controlled with medication. Most children with this type of CMD die in childhood.

Symptoms of CMD usually progress slowly and may include general weakness, a flaccid muscular tone, abnormally bent joints, and slow motor development. Such muscular weakness may only be noticed by parents when the child fails to achieve certain milestones in motor function and control.

CMDs constitute a group of autosomal recessive muscular dystrophies that affect both boys and girls. The symptoms may range from mild to severe and muscle degeneration is restricted primarily to skeletal muscle. Without physical therapy and other forms of treatment, most children with CMDs are unable to sit or stand without support, and some affected children may never learn to walk.

Congenital MDs are further subdivided into 3 groups, according to etiology:

1/ merosin-negative disorders, which are due to an absence of the protein merosin, which is found in the connective tissue that surrounds muscle fibers,

2/ merosin-positive disorders, in which merosin is present but there is an absence of other proteins that are necessary for proper muscle metabolism, and

3/ neuronal migration disorders, in which there has been a disruption in the migration of nerve cells during the formation of neurons that occurs in the very early developmental stages of the fetal nervous system.

The protein merosin normally lies outside muscle cells and links the cells to surrounding tissue. Nearly half of all cases of congenital MD are caused by merosin disorders, which are responsible for the most common symptoms such as muscle and tendon contracture, scoliosis, and respiratory difficulties that ultimately lead to respiratory failure. CMDs involving disorders of neuronal migration may also affect the central nervous system, causing vision and speech problems, seizures, and structural changes in the brain. It is not uncommon for children with these symptoms to die in infancy.

CMDs have been linked to genetic defects on chromosome 9, although the precise genes that are involved on this chromosome are yet to be discovered.

Fukuyama CMD has been linked to a mutation on chromosome 6, but its precise gene and corresponding protein have yet to be identified. Fukuyama CMD is also known as “congenital muscular dystrophy with merosin deficiency”.

4. Emery-Dreifuss MD (EDMD):

EDMD is the rarest of all forms of MDs. Fewer than 300 cases have been identified.

The most striking feature of EDMD which distinguishes it from other dystrophies is progressive cardiac disease, the severity of which is much greater than the skeletomuscular atrophy found throughout the body. Primarily affecting young boys, EDMD involves problems in the electrical conductivity of the heart, with secondary symptoms including a distinct pattern of muscle atrophy.

In people with EDMD, ECGs (electrocardiograms) typically become abnormal by the age of 20 to 30 years, at which time first-degree atrioventricular blockage is detectable, with the atria becoming involved earlier than the ventricles, such that atrial fibrillation and flutter are observable, including permanent atrial standstill and junctional bradycardia. By the age of 35 to 40 years, virtually all individuals with EDMD exhibit abnormalities in impulse generation or conduction such that permanent ventricular pacing is often required.

The age of onset of EDMD is usually prior to the age of 10, although symptoms may appear as late as the mid-twenties. EDMD causes slow but progressive wasting of the upper arm and lower leg muscles, which is less severe than in Duchenne but which is often preceded by contractures in the spine, ankles, knees, elbows, and back of the neck. Mild facial weakness may also occur, and serum creatine kinase levels may be moderately elevated. Life-threatening heart problems are the main hallmark of EDMD, however, and a high percentage of EDMD patients have serious cardiac complications by the age of 30, which are specifically due to defects in the electrical conduction of the heart and often require a pacemaker or other type of cardiac assistive device. Since female carriers of the disorder often exhibit cardiac problems without muscle weakness, the sisters and mothers of boys with EDMD should be tested. Patients can live until middle age but ultimately die from progressive pulmonary or cardiac failure, or both.

The skeletal muscle weakness in EDMD is less severe than that of other dystrophies, such as Duchenne, and the pattern of progression is the opposite of that usually seen, affecting the shoulder and upper arm muscles in the initial stages and progressing to the lower extremities in later stages. Also unlike other forms of MD, contractures may appear in EDMD prior to any symptoms of muscle weakness.

EDMD has two forms: one that is X-linked recessive and one that is autosomal dominant. The X-linked recessive form is caused by a defect in the gene on the X chromosome that codes for the protein emerin, the precise function of which has not yet been identified. The precise genetic locus of the autosomal dominant form also has not yet been identified.

5. Facioscapulohumeral MD (FSH):

FSH is the third most common form of MD, after Duchenne and Becker MD.

Symptoms may begin during infancy, late childhood or early adulthood. The first sign is usually facial weakness, with difficulty smiling, whistling and closing the eyes. Later, there is difficulty raising the arms or flexing the wrists and ankles. The disease occurs in both genders and in all racial groups.

Also known as Landouzy-Dejerine disease, FSH occurs in approximately 1 in every 20,000 people. In the United States, it has been estimated that approximately 15,000 people are currently afflicted with FSH.

This particular form of MD takes its name from the muscles of the face (facio), the shoulders (scapulo), and the upper arms (humera), which are the areas affected by FSH. Muscles of the legs and pelvis are often, though not always, spared, as FSH is characterized primarily by wasting of the upper body, including the chest muscles, with rare but occasional secondary wasting of the hip and leg muscles. A loss of facial expression is one of the most noticeable symptoms of this disorder.

Muscles around the eyes and mouth are often affected first, which are followed by weakness around the lower shoulders and chest. Reflexes in the biceps and triceps are also impaired. Early symptoms may also include lordosis, which is an abnormal backward curvature of the spine, and hearing loss, particularly at high frequencies. Contractures are rare, although some individuals experience severe pain in the affected limbs.

The usual age of onset is not until late childhood or early adulthood, occurring most commonly in the teenage years. In some cases the disease may not appear until the age of 40, however, and an infantile form of FSH also exists, in which symptoms become evident shortly after birth and most commonly include retinal disease and hearing loss.

The rate of progression of FSH is slow in comparison to that of other forms of MD, and the severity of symptoms can range from mild to disabling. Life expectancy is normal, even though some individuals may become severely handicapped. The course of the disease may be punctuated by long periods of stability that are interspersed with shorter periods of rapid muscle deterioration and increased weakness.

Approximately half of all people with the FSH form of MD retain the ability to walk throughout their lives. Orthopedic supports are helpful in the early stages although surgery is often prescribed in the latter stages to improve certain functions, especially in the shoulder area.

FSH is known to be an autosomal dominant disorder, although the precise genetic defects that are responsible for this particular form of MD have not yet been discovered. Nevertheless, scientists are narrowing in on the specific genetic locus of this disease, which is believed to be a particular allele on the chromosomal region that is linked to FSH. Researchers at the Leiden University Medical Center and the University Medical Center of Nijmegen in The Netherlands have recently compared the alleles 4qA and 4qB in a group of 80 individuals known to have FSH with those of a control group consisting of 80 healthy individuals. Led by Silvèere van der Maarel, Ph.D., the scientists discovered that only the 4qA allele was affected in the FSH group, thus shedding some light on the exact location of the genetic defect that is responsible for FSH. Their research was supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMSD).

6. Limb-Girdle MD (LGMD):

LGMD refers to more than a dozen disorders characterized by a degeneration of muscles that are closest to the trunk.

Clinically, LGMD is classified into 2 primary categories, namely, the severe childhood form that is similar in appearance to DMD, and a slightly milder form that appears in a person’s teens or twenties. Both males and females may be affected with LGMD, which is most noticeable in the upper legs and shoulders. Life expectancy may be reduced only slightly, and intelligence remains normal. Although the rate of progression is slow when compared to other forms of MD, most people with LGMD become severely disabled within 20 years after the onset of the disease.

In LGMD, the proximal reflexes, which are those closest to the center of the body, are frequently impaired, corresponding to the “limb-girdle” region implied by the name. Consequently, patients may also experience cardiomyopathy and respiratory complications. The course of the disease consists of progressive atrophy of the muscles that are closest to the trunk.

LGMD has been linked to several genetic defects that were found on chromosomes 2, 13, 15 and 17. At least three variations of an autosomal dominant form of LGMD have been identified, which are classified as “type 1” and are known to result from genetic defects on chromosome 5. Eight variations of an autosomal recessive form of LGMD are also known to exist, which are classified as “type 2”, although the associated genes have not yet been identified. Chromosome 15 is known to contain a gene that codes for the caplain 3 enzyme, and chromosome 17 is known to contain a gene which codes for the protein adhalin, mutations in which have been implicated in certain cases of LGMD. Some types of the autosomal recessive forms of this disease are now understood to be the result of a deficiency of any of the four sarcoglycans that constitute the dystrophin-glycoproteins, although their precise genetic components have not yet been located.

The autosomal recessive forms of LGMD occur more frequently than the autosomal dominant forms, usually beginning in childhood or in the teenage years, and are marked by dramatic increases in serum creatine kinase levels. The autosomal dominant forms of LGMD usually begin in adulthood.

LGMD is first noticeable in muscular weakness that begins around the hips and later spreads to the shoulders, legs, and neck. A “waddling” gait may develop, along with difficulty in climbing stairs, carrying heavy objects or rising from a sitting or lying position. Falls become more frequent and patients ultimately lose the ability to run. Unlike in other forms of MD, contractures in the back muscles are common but not at the elbows or knees.

The number of people with LGMD is not known with certainty, but in the United States it is estimated to be in the low thousands.

7. Distal MD (DD):

Distal muscular dystrophy occurs almost exclusively in Sweden, with rare exceptions.

Also known as “distal myopathy”, DD describes a group of at least six specific muscle diseases which are typically less severe, progress more slowly, and involve fewer muscles than other forms of MD. Life span is not usually affected, although distal MD can spread to additional muscle groups, including those of the cardiac and respiratory systems, and patients may eventually require the use of a ventilator.

In its most common forms, symptoms of distal MD do not begin until middle age or later, and are confined primarily to muscular weakness in the feet and hands. As the name implies, the muscles affected by this disease are those farthest away (distal) from the trunk.

Early signs may include difficulty with activities that require fine motor skills, such as tying shoes or fastening buttons. Diagnosis includes testing of fine hand and finger control, as well as control over the muscles of the foot, such as the ability to hop or to stand on one’s heels. Onset of distal MD is typically between 40 and 60 years of age.

Distal MD is primarily an autosomal dominant disorder in which there is an absence or lack of dysferlin, which is a key component in a muscle membrane protein complex. Autosomal recessive forms have also been found in young adults, in which the symptoms are similar to those of Duchenne MD but with a different pattern of muscle atrophy. An infantile-onset form of autosomal recessive distal MD has also been reported, in which slow but progressive weakness is first noticed around the age of one year, when the child begins to walk, and continues to progress very slowly throughout adult life.

One of the autosomal dominant forms of distal MD is known as “Welander”, in which the muscles of the hands are affected first. Conversely, “Markesbery-Griggs” and “Nonaka” are two other autosomal dominant forms in which the front of the lower legs are affected first. “Miyoshi” is an autosomal recessive form of distal MD that is caused by a genetic defect on chromosome 2, in which the backs of the lower legs are affected first.

8. Myotonic MD:

Also known as “dystrophia myotonica” and Steinert’s disease, myotonic MD is the most common form of MD in adults, afflicting approximately 30,000 adults in the United States alone.

Symptoms may begin at any time throughout the course of one’s life span, from birth through adulthood, although the typical age of onset is around the third decade of life. The disease affects both males and females, and is characterized by general weakness in the muscles of the face, hands and feet, with systemic complications that often result in the latter stages of the disease. As the name implies, people with this form of MD also experience myotonia, which is the failure of muscles to relax after use, and this is the only form of MD in which myotonia is found.

Life span is not usually affected, and the rate of progression is slow. Most people with myotonic MD are severely disabled within 20 years of the onset of symptoms, but do not usually require a wheelchair. The muscles of the face and the front of the neck are the first to weaken, although myotonic MD also affects the central nervous system, the cardiovascular system, the endocrine system, the gastrointestinal tract and the eyes. Symptoms may include prolonged muscle spasms, retinal degeneration, the formation of cataracts, cardiac abnormalities, endocrine disturbances, insulin resistance, ptosis (drooping eyelids), mild mental impairment, low motivation, abnormal emotional indifference, and an excessive need for sleep. Muscular atrophy is usually most severe in the forearms and calves. Gastrointestinal problems often require continuous medical attention in myotonic MD, as do cardiovascular complications which often require a cardiac pacemaker in combination with drug therapy, although patients with myotonic MD also tend to exhibit abnormally adverse reactions to general anesthesia. A difficulty relaxing the grasp, especially after having held cold objects, is one of the first signs of myotonia in this disease.

Myotonic MD is an autosomal dominant disease that results from a mutation of a gene on chromosome 19 which codes for the myotonin protein kinase enzyme. This particular mutation results in an abnormally long repetition of a three-letter sequence in the genetic code, which is already repeated several times in healthy individuals, but in people with myotonic MD the sequence is repeated too many times. The result is an abnormal lengthening of the triplet with each successive generation. This particular triplet-repeat mechanism has now been linked to more than 15 other disorders, including Huntington's disease and the spinocerebellar ataxias.

Unlike some other forms of MD, myotonic MD may be expressed in both men and women, and some symptoms are specific to each gender. Males may experience early frontal balding, impotence and testicular atrophy, for example, while females may exhibit irregular menstruation and infertility. Other symptoms vary by age, and a childhood form of myotonic MD may become apparent between the ages of 5 and 10 years, with symptoms that include general muscle weakness, especially in the face and distal muscles, an overall lack of muscle tone, and mental impairment. In such cases, the gene is usually inherited from the father.

A rare and severe congenital form occurs almost exclusively in the children of women who are carriers of myotonic MD but who do not exhibit symptoms of the disease themselves, and who therefore often do not know that they are carriers of the disease. In this case, symptoms in the child are present from birth and may include difficulty swallowing or sucking, impaired breathing, absence of reflexes, skeletal deformities such as club feet, and noticeable muscle weakness, especially in the face. It is not uncommon for children with congenital myotonic MD to experience mental impairment and delayed motor development.

9. Oculopharyngeal MD (OPMD):

OPMD differs from other forms of MD in a number of ways, most notably in the age of onset, which is not usually before the age of 40 and it may even occur after the age of 60. As the name implies, OPMD affects muscles in the eyes and throat, which exhibit a progressive weakening over time. The disease is seen both in men and in women, and in the United States OPMD is most commonly found among families of French-Canadian and Hispanic descent.

Early signs often include drooping eyelids, facial muscular weakness and pharyngeal changes that may alter the sound of the voice. Vision problems may be severe and may include retinitis pigmentosa, which is a progressive degeneration of the retina that affects night vision and peripheral vision. Muscle atrophy in the neck, shoulders and limbs may occur in advanced stages, and cardiac irregularities are not uncommon. Although the progression of the disease is slow, in its most severe cases people who are afflicted with OPMD may eventually lose the ability to walk. Dysphagia (difficulty swallowing) is common and may lead to choking and recurrent pneumonia.

OPMD has been linked to a genetic defect on chromosome 14.

Causes:

To a certain extent, epidemiology reflects etiology, as the statistics associated with any disease vary, to some degree, from nation to nation. The underlying reasons for such associations, however, linking genetics and pathology with culture, environment and lifestyle, have yet to be fully elucidated.

Although certain types of MD are more common to some nations than to others, all of the muscular dystrophies appear to result from mutations in any of the thousands of genes that are involved in protein metabolism. Some of the mutations that occur in MD are the result of spontaneous mutations, not found in the genes of either parent, while other mutations are known to follow a pattern of inheritance within the family.

Everyone has 46 chromosomes, half of which are inherited from each parent. Of the 23 chromosomes that each parent contributes to each child, one chromosome determines gender (X or Y) while the other 22 chromosomes are known as autosomes. Muscular dystrophies may therefore be inherited in one of three possible ways:

1/ through autosomal dominant inheritance, which occurs when only one copy of the defective gene, inherited from one parent, is sufficient to cause the disease,

2/ through autosomal recessive inheritance, which occurs when two copies of the gene must be inherited to cause the disease, one gene from each parent, with each parent being a carrier of the defective gene but without expressing symptoms themselves, and

3/ through X-linked recessive inheritance, which occurs when the gene is inherited by a son from his mother who is a “carrier” of the gene on one of her two X chromosomes.

In autosomal dominant inheritance, each child has a 50% chance of inheriting the defective gene and being afflicted with the disease. In autosomal recessive inheritance, each child has a 25% chance of inheriting both copies of the defective gene from both parents, in which case the child will be afflicted with the disease, and for each child there also exists a 50% chance of inheriting only one copy of the defective gene from one parent, in which case the child will not be afflicted with the disease but will become a carrier of the gene, possibly transmitting the disease to his or her children. In autosomal recessive inheritance each child also has a 25% chance of not inheriting the defective gene from either parent, in which case the child will neither be afflicted with, nor be a carrier of, the disease.

Both sons and daughters of carrier mothers have the same 50% chance of inheriting the disorder, although daughters are not usually affected by the disease when they inherit the gene, since the healthy X chromosome that is inherited from the father is usually capable of offsetting the faulty gene received from the mother. Fathers afflicted with the disease cannot pass an X-linked disorder to their sons but they can transmit the disease to their daughters, who will not be afflicted with symptoms but will become carriers of the defective gene. In rare instances, carrier females do exhibit mild symptoms of MD, but such cases are exceptional.

Muscle Physiology:

As with any disease, an understanding of the cellular mechanisms that are at work in normal, healthy individuals will facilitate an understanding of the processes that go awry and result in the specific manifestations of the disease.

Muscles are composed of thousands of muscle fibers, each one of which in turn is composed of numerous individual cells that have joined together during development, are encased by an outer membrane, and are bound together by connective tissue.

Muscles are activated to contract when an electrical impulse originating in the brain travels along the peripheral nerves of the central nervous system to the various sensory organs and to the muscles. An important component of such neurophysiological pathways is the “neuromuscular junction”, which is a gap that exists between the nerve fiber and the particular muscle that is activated by the particular nerve fiber. Assuming no interruptions in the transmission of the electrical signal along this route, acetylcholine will be released at the neuromuscular junction, which in turn will trigger a series of events that will cause the muscle to contract.

In healthy individuals, the ordinary wear and tear of daily life results in a continuous catabolism (breaking down) of muscle fiber, which in turn stimulates a continuous anabolism (building up) of muscle fiber, during which the body’s natural ability to heal and repair itself is active. In degenerative muscular diseases such as MD, however, these automatic repair mechanisms are rendered ineffective.

As previously mentioned, muscle fiber consists of a protective membrane which contains a group of proteins known as the dystrophin-glycoprotein complex, which prevents damage to muscle fibers during repeated contraction and relaxation. In the event that the membrane is damaged nonetheless, creatine kinase enzymes leak out of the membrane while calcium ions leak into the membrane. This exchange of enzymes and ions causes further damage which ultimately results in death of the muscle fibers. Creatine kinase is a key component in the chemical reactions that produce energy for muscle contractions, and any malfunction in the activity of this enzyme will result in a series of abnormal cellular events.

In the various muscular dystrophies, this ongoing cellular damage is compounded by the complete or partial absence of normal proteins, thereby interrupting the natural repair mechanisms of the muscle cells.

The end result is a series of abnormal events not only at the cellular level but also at the gross systemic level as well. Progressive damage to muscle tissue without repair results also in the branching and splitting of muscle fiber, and in phagocytosis, which is a process in which muscle tissue is specifically targeted and destroyed by scavenger cells. The absence of normal proteins, and the continuous damage of muscle fiber without repair, results ultimately in the chronic or permanent shortening of tendons with an eventual loss of tendon reflexes and muscular tone. This muscular damage in turn triggers a progressive degeneration of related organs and systems throughout the body. In its most severe cases, death is inevitable.

Diagnosis:

A wide variety of tests are available not only to detect the presence of MD, but also to identify with molecular specificity the precise type of MD that an individual may have. In fact, many of the new treatment strategies that are currently being developed for MD require a precise molecular diagnosis and an exact identification of the specific genetic mutations that are involved.

When MD is suspected, a physician will typically begin with the most generalized tests and advance through increasingly more sophisticated and specialized tests as necessary, to verify positive results. It is not uncommon, however, for repeated negative results of such testing to confirm the absence of MD, in which case other types of neuromuscular pathologies must be investigated.

Laboratory evaluations for MD may include tests to measure serum levels of the muscle enzyme creatine kinase (CK), which leaks into the blood from muscle fibers whenever the muscle membrane is damaged. Elevated levels of CK may be detectable prior to the manifestation of physical symptoms, and may indicate either Duchenne or Becker MD. Measurement of CK levels is also useful in identifying women who are carriers of the gene.

Blood and urine tests are useful in identifying specific neuromuscular and skeletomuscular disorders, which may be detectable by serum aldolase levels. This enzyme is involved in the breakdown of glucose, and elevated levels have been found in patients with MD.

High levels of myglobin have also been found in patients with MD. A protein that binds with oxygen, myoglobin is present in cardiac and skeletal muscle cells and is known to be elevated whenever injury or disease are present.

Muscle biopsies may be conducted for histological analysis of muscle fiber. Because of the increased specificity of molecular techniques, however, muscle biopsies are no longer as important for the diagnosis of most types of MD as they once were, although such tests are still useful in detecting neuropathy, inflammation and various types of myopathy. Muscle biopsies are also useful in carrier testing, and tissue samples derived from muscle biopsies may be stained with fluorescent markers for the immunofluorescent detection of certain proteins, such as dystrophin, or the absence thereof, within the fibers.

Neurophysiological testing may be prescribed to identify physical and chemical changes in the nervous system. Similarly, electromyograms (EMGs) measure nerve conduction velocity and the response of stimulated muscle, and may be prescribed for assessing nerve damage. Electrocardiograms (ECG) may be ordered if cardiac irregularities are suspected.

If any of these tests indicate abnormalities, genetic evaluation may be prescribed for confirmation of the presence of any of the various mutated genes that are known to be linked to MD.

Known also as “molecular diagnosis”, genetic testing with polymerase chain reaction (PCR) is a method of generating and analyzing multiple copies of DNA fragments. DNA analysis and associated enzyme assays that test for genetic linkage and chromosomal markers are especially useful techniques when studying family members across generations.

Serum electrophoresis may also be prescribed, in which a blood sample is exposed to an electric current which causes the different proteins to separate into bands according to their conductivity, thereby indicating the relative proportion of each protein fragment.

Electron microscopy is capable of detecting the specific muscle fiber changes that are evident in one particular variation of Distal MD. Electron microscopy may also identify changes in mitochondria and increased amounts of connective tissue.

Exercise tests may be prescribed to assess muscle strength, especially respiratory weakness, as well as to detect elevated levels of metabolic chemicals following exercise.

Prior to the birth of a child, amniocentesis and chorionic villus sampling (CVS) are often employed as part of routine genetic counseling, whenever MD may be present in a family.

Magnetic resonance imaging (MRI) and ultrasound may both be useful in assessing the extent of the disease by showing the amount of fat and connective tissue that have replaced muscle.

A person’s particular symptomology, and the specific type of MD that is suspected, will dictate the particular combination of tests that should be prescribed. For example, in the case of Becker MD (BMD), diagnosis may be established on the basis of clinical and neurological assessments if myopathic EMG (electromyography) patterns are revealed along with elevated serum levels of the creatine phosphokinase enzyme. Additionally, muscle biopsies should include histochemical and enzyme histochemistry techniques as well as an immunohistochemical study of the dystrophin protein, in order to detect abnormal changes that are specific to BMD. If the results of such tests are positive, diagnosis may be confirmed by DNA analysis of the dystrophin gene, if mutations are revealed which consist either of single or multiple deletions within the gene itself.

In Duchenne MD (DMD), however, multiplex polymerase chain reaction analysis of DNA for deletion mutations may detect the presence of DMD only in approximately 65% of patients, due to the frequency of false-negatives. An additional cause of inaccuracy results from the fact that approximately 20% of patients with DMD do not possess the mutated dystrophin gene, so in patients who test negative for a gene deletion mutation, therefore, it is necessary to analyze the dystrophin protein further by muscle biopsy and more refined immunohistochemical techniques such as by cryostat sections or immunoblotting. Dystrophin deficiency alone is often a sufficient indication of the presence of DMD, even in the absence of DNA confirmation.

As with any disease, a complete family medical history is always worthy of important consideration.

Treatment:

It is commonly recognized that a single comprehensive treatment for MD does not exist, although a number of therapies are being developed. Even with the specificity of genetic testing, however, conventional medical treatments are not able to stop or reverse the progression of MD. The goal of most therapies is therefore to alleviate symptoms as much as possible while preventing complications for as long as possible.

Conventional medical treatment of MD may include respiratory therapy, physical therapy, speech therapy, drug therapy, orthopedic supports, corrective orthopedic surgery, and any combination thereof. Assisted ventilation is often a part of respiratory therapy, and pacemakers are frequently necessary to correct cardiac abnormalities.

Whenever surgery is involved, extra care should be taken among physicians in the selection of anesthesia, since muscular dystrophy patients regularly exhibit malignant hyperthermia, which is a severe reaction to the halothane anesthesia.

Drug therapy may include corticosteroids to slow muscle degeneration, anticonvulsants to control seizures and irregular muscle activity, immunosuppressants to delay some types of damage to dying muscle cells, and antibiotics to fight infections.

In an attempt to delay muscle deterioration, especially in Duchenne MD, corticosteroids such as prednisone are often prescribed, and these drugs have shown some success in prolonging independent ambulation in children, although prednisone has proven to be most effective when administered relatively early in the disease. Side effects of corticosteroids are numerous and well documented, however, and frequently include, among other symptoms, weight gain and bone fragility which are especially dangerous in children with MD.

As stated by the National Institute of Neurological Disorders and Stroke (NINDS), a division of the National Institutes of Health (NIH),

“Corticosteroids are known to extend the ability of Duchenne MD patients to walk by up to 2 years, but steroids have substantial side effects and their mechanism of action is unknown.” (From www.ninds.nih.gov)

In fact, the side effects of corticosteroids have reached such a level of concern that the National Institutes of Health (NIH) have funded a group of scientists to establish a new set of clinical standards for the treatment of Duchenne MD with steroids, with the specific objective of modifying the steroids in such a way as to reduce or eliminate their side effects.

Deflazacort has been found to be as effective as prednisone in slowing the progression of MD, and with fewer side effects, but this medication has not been approved for use in the United States.

The anabolic steroids oxandrolone and creatine are also frequently prescribed to increase muscle strength. Creatine is produced naturally by the liver in small quantities and is also obtained from some foods such as animal proteins, and it is known to play an important role in the production and circulation of ATP (adenosine triphosphate) in the muscles. Creatine levels are abnormally low in people with MD, and pharmaceutical supplementation is often prescribed. Such “performance enhancing drugs”, however, have been a recurrent topic of public controversy among athletes, with well documented side effects that most commonly include, in their mildest forms, hypertension resulting from elevated blood pressure and elevated cholesterol levels, and in high doses creatine has been known to damage the kidneys, the liver and the heart. Oxandrolone is a synthetically modified form of testosterone, with numerous side effects that may include serious and even fatal liver complications. Oxandrolone is also a known tetragen, meaning that it causes birth defects and therefore should never be taken by anyone who is pregnant or who may become pregnant while on this medication. Tetragenic substances, in fact, should not even be handled by anyone who is pregnant or who may become pregnant, since such chemicals may be absorbed into the bloodstream through the skin.

According to experimental results from mouse models, the amino acid glutamine in combination with creatine monohydrate may be therapeutic for Duchenne MD, and this combination is being tested for safety and efficacy in humans.

Clinical trials are also underway to test the safety and efficacy of oxatomide, a steroid that interferes with the release of histamine from mast cells and which is involved in inflammatory diseases. If determined to be safe and effective, oxatomide may be prescribed for boys with Duchenne MD.

Immunosuppressive drugs such as cyclosporin and azathioprine have been found to delay some damage to dying muscle cells in patients with MD. Mexiletine, phenytoin and baclofen, which block signals from the spinal cord to the muscles, are often prescribed to provide temporary relief from myotonia, as are quinine and dantrolene, which interfere with the process of muscle contraction.

Paradoxically, drugs that have been specifically formulated for the treatment of myotonia are not effective in treating myotonic MD.

Anticonvulsants, also known as antiepileptics, may be prescribed to control seizures and some types of erratic muscle activity. Some oral anticonvulsants that are commonly prescribed to MD patients include carbamazepine, phenytoin, clonazepam, gabapentin, topiramate, and felbamate.

Respiratory infections are typically treated with antibiotics.

Using techniques of high-throughput screening (HTS), which allows for the testing of hundreds of chemicals with rapid results, various pharmaceutical compounds are being experimentally tested in the upregulation of compensatory proteins, such as utrophin, in the replacement of dystrophin. Utrophin has shown some therapeutic success in animal models as a substitute for dystrophin.

Scientists at the National Institute of Neurological Disorders and Stroke (NINDS), in collaboration with researchers at several universities, are investigating the potential enzymatic inhibitory capabilities of various agents that may prevent specific enzymes from degrading muscle, thereby delaying muscle degeneration. Although oxandrolone has already been mentioned, it is being further tested in combination with albuterol as a potential medication specifically for patients with FSH. It is believed that such combinations of drugs may also be more effective when given in a “pulsed” manner rather than when given continuously.

As with all drugs, side effects and contraindications should be discussed thoroughly with one’s physician.

As a result of growing concern over the numerous side effects of pharmaceutically formulated medications, patients and researchers alike are increasingly turning their attention to nutritional supplements, and to the potentially therapeutic benefits of such supplements. One such example is coenzyme Q10 (CoQ10), which is already known to protect cell membranes from oxidative stress and is also believed to play a key role in mitochondrial function and in the production of cellular energy. This and other nutritional supplements are of increasing scientific interest for the roles that they play in cellular health, and for their possible use in the treatment of diseases such as MD.

Research is also underway to enhance the body’s natural muscle repair mechanisms, which are of increasing interest to scientists, as it is hoped that drug or gene therapy may be able to target the specific points along these repair “pathways” that are defective in MD. A new class of potentially therapeutic modalities is therefore being studied with the objective of enhancing the body’s natural repair processes.

Myostatin, for example, is an inhibitor of muscle growth, and methods of blocking myostatin have resulted in enhanced muscle repair in animal models of MD. The protein dysferlin, by contrast, is involved in muscle repair, and researchers are investigating various methods of increasing the cellular expression and efficacy of dysferlin. Insulin-like growth factor 2 (IGF-2) has also been found to enhance natural muscle repair mechanisms, and is being used in clinical trials for the treatment of Myotonic Dystrophy.

Future Research Directions:

In the United States, most MD research is funded by The National Institute of Neurological Disorders and Stroke (NINDS), which supports a broad program of research not only on MD but also on numerous other diseases, as its name implies.

NINDS and its sister institutes, the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) and the National Institute of Child Health and Human Development (NICHD), together lead the MD research efforts that are conducted at the National Institutes of Health (NIH) and at grantee institutions throughout the country.

With support from such organizations as these, protein replacement therapy and gene replacement therapy constitute the two main areas of greatest research focus in the development of new treatments for MD.

In one example of successful protein replacement therapy, researchers at the University of Pennsylvania conducted a study led by Tejvir S. Khurana, M.D., Ph.D. in which a fragment of the protein heregulin was injected into mice that were bred to develop a disease similar to DMD. After 3 months, the heregulin was found to improve both the structure and function of muscle tissue in the mice, with a partial restoration of the mechanical properties and a reduction in the inflammation of muscles in the mice. Heregulin is known to increase the body’s natural production of utrophin, which is a muscle protein similar in structure and function to dystrophin. Utrophin is naturally present in mammals prior to birth, after which time levels gradually diminish throughout life. By increasing utrophin levels, muscle degeneration is slowed. This method uses the body’s own cells to produce and regulate utrophin as a natural substitute for dystrophin in maintaining muscle function. The procedure may be therapeutic for certain forms of MD such as Duchenne and Becker, which are characterized by a complete absence of dystrophin in the former and a partial absence of dystrophin in the latter, to such a severe extent that few people with Duchenne MD live past their 20s.

In gene replacement therapy, researchers are looking for specific ways in which to replace defective genes with functional genes. A number of obstacles have presented challenges to this objective in the past, however, such as the large size of the dystrophin gene, for example, which has prevented transmission of this gene by the viral vectors that are typically employed as gene-delivery systems. Now researchers have developed a “mini-dystrophin” gene, which resembles the larger gene in its properties and function yet which is small enough to be carried and delivered by a viral vector, and which therefore allows the targeting of specific muscle tissue. Gene replacement therapy which uses this “mini-dystrophin” gene has proven to be a successful form of treatment in mouse models of Duchenne MD.

Researchers in the related field of “genetic modification therapy” are developing ways to circumvent inherited mutations via manipulation of the protein synthesis process. Such methods have been shown to produce a gene that is capable of ignoring the genetic mutations that stop the production of dystrophin.

The antibiotic gentamicin has also been shown to ignore such genetic mutations, and it has been found to be effective in treating approximately 15% of individuals with Duchenne MD, in a NINDS-funded clinical trial.

An alternative approach to gene therapy uses gene splicing technology to jump past the mutations in the dystrophin gene to a point where the genetic information is complete, thereby producing a functional protein. This strategy has also been found to be effective in a mouse model of Duchenne MD.

Many of the new therapies that are being developed are the result of research that is specifically focused on Duchenne MD, since this is the most common form of the disease, although discoveries made with Duchenne MD may ultimately have applications to other types of MD.

“Translational research” is a major focus of NINDS and other funding organizations, which involves bringing (“translating”) discoveries that are made in the laboratory to a clinical setting.

One such example involves insulin-like growth factor 1 (IGF-1). Dr. H. L. Sweeney and colleagues at the University of Pennsylvania have created a new mouse model which is a cross between a strain with muscular dystrophy symptoms and another strain with high levels of IGF-1. Since IGF-1 is known to be involved in the manufacture of proteins and therefore in the regeneration of muscle, this particular hybrid mouse not only exhibits an increase in muscle mass but also a reduction in muscle damage. The director of the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), Stephen Katz, M.D., Ph.D., announced

“This is indeed good news for the muscular dystrophy community. The combination of better muscle regeneration and less muscle wasting could lead to a better muscle capacity over time. Less muscle effort would be needed to produce a required force, so muscle would be less likely to be damaged by normal activity.”

Scientists and physicians collaborating together in translational research are now exploring methods of application by which IGF-1 may be administered, safely and effectively, in the treatment of humans with MD. This particular research was funded jointly by the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) and by the Muscular Dystrophy Association.

Together NINDS and NIAMS are funding a research registry specifically for FSH and Myotonic MD. According to their websites, this registry “serves as a resource for scientists seeking a cure for these diseases, in addition to enhancing research on what changes occur in MD.” (From www.nih.gov).

The registry is based at the University of Rochester in New York, and scientists have access to statistical analyses of the registry data and to registry members who have agreed to participate in particular research studies.

Similar registries for Duchenne MD are supported by the U.S. Centers for Disease Control and Prevention.

In December of 2001, President George W. Bush signed into U.S. law the Muscular Dystrophy Community Assistance, Research, and Education Amendments Act of 2001 (the MD CARE Act, Public Law 107-84). As part of the Act, certain branches of NIH, namely, NIAMS, NINDS (the National Institute of Neurological Disorders and Stroke), and NICHD (the National Institute of Child Health and Human Development), in collaboration with the Muscular Dystrophy Association, have funded a consortium of Centers of Excellence for research on muscular dystrophy known as the Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Centers. Six Centers exist thus far, three of which were established in 2003 and three in 2005. As part of their charter they are required to make core resources and services available to the national muscular dystrophy research community. They also fund translational research on gene therapy, stem cell therapy, molecular and pharmacological treatments, and clinical studies of such genetic and pharmacological therapies. These Centers are intended to serve as focal points of research, collaboration, and training for the MD scientific and patient care communities.

The Act also authorized the Centers for Disease Control and Prevention to award grants for epidemiological studies and data collection, and other federal agencies also contribute to this research initiative. In response to the MD CARE Act, NIH also formed the Muscular Dystrophy Coordinating Committee to help guide research on MD. As described on their website,

“The MD Coordinating Committee (MDCC) is made up of physicians, scientists, NIH professional staff, and representatives of other federal agencies and voluntary health organizations with a focus on MD. The purpose of the group is to help NIH add new capabilities to the national effort to understand and treat MD, without duplicating existing programs. The MD Coordinating Committee has developed a broad research and education plan and continues to refine the plan to improve basic, translational, and clinical research in MD, with the goal of improving the quality of life for patients with MD.” (From www.ninds.nih.gov).

NINDS itself is a member of the Muscular Dystrophy Coordinating Committee.

Stem Cells:

The stem cell-based therapies of regenerative medicine continue to offer increasing evidence for the successful treatment of a variety of diseases such as MD. In fact, stem cell therapy has already shown very positive results in reversing the effects of MD in animal models.

Stem cells replace the missing proteins, such as dystrophin in Duchenne and Becker MD, merosin in Congenital MD, emerin in Emery-Dreifuss MD, adhalin and the four sarcoglycans that constitute the dystrophin-glycoprotein complex in Limb-Girdle MD, dysferlin in Distal MD, and stem cells may also correct the abnormally long repetition of the DNA sequence from the myotonin protein kinase enzyme gene that is found in Myotonic MD.

Replacing such proteins that are either absent or defective results not only in the regeneration of damaged tissue but also in the protection of muscle fiber from further degeneration.

As already described, muscle tissue exhibits a natural regenerative capacity in healthy individuals. This innate, characteristic feature of muscle fiber makes muscle-related diseases such as the muscular dystrophies prime candidates for stem cell therapy. In fact, since stem cells are naturally present within muscles, one approach is to stimulate those stem cells that already exist within the muscle fibers. Even when stem cells from an external source are used, the muscle cells of the body are highly responsive to such stimuli, with a natural tendency to regenerate themselves, and in fact such properties are prime hallmarks that distinguish muscle tissue from most other tissue in the body. While basic catabolic (breaking down) and anabolic (building up) processes occur in all tissue types throughout the body, at all times, such fundamental properties of metabolism are most actively “programmed” into muscle tissue.

Muscle fibers contain a highly specialized type of cellular environment that naturally promotes the continuous regenerative capacity of this tissue. Such a cellular milieu consists of a number of built-in mechanisms and “cues” that automatically trigger regeneration whenever fibers are worn down or injured. More so than any other types of tissue throughout the body, muscle tissue is constantly being worn down and rebuilt, even under normal circumstances in healthy individuals, and it is part of the very nature of muscle that it cannot be built up unless it is first torn down. Such natural processes are of increasing interest to researchers, and a number of national and government organizations, such as NIAMS, have funded various studies that are directed toward understanding these molecular and cellular processes. Stem cells are therefore particularly effective in such a natural cellular environment that is already highly conducive to regeneration.

Stem cells may also be used as a delivery system for functional genes, such as the dystrophin and other genes, to replace the defective genes in MD patients. In fact, such corrective gene replacement therapy with stem cells has already been successfully performed with dystrophic mice.

A family of natural growth-stimulating proteins known as “Wnt” has been found to prompt stem cells to rebuild damaged muscles in mouse models. The Wnt family of proteins constitute a group of signaling molecules that are known to regulate cell-to-cell interactions during embryogenesis and cancer, but given the right cellular signals and “cues”, the Wnt proteins have also been shown to stimulate stem cells that naturally exist within the muscle tissue, thereby regenerating muscle fiber after an injury. Wnt itself is naturally present in cells throughout life and becomes activated whenever needed.

Stem cells known as mesoangioblasts have shown the ability to cross the inner lining of blood vessels and migrate extensively throughout tissue in the regeneration of muscle. Mesoangioblasts may be collected from a person’s blood vessels, genetically “fixed” in the laboratory where they are allowed to multiply, and then injected back into the bloodstream of the patient where they migrate to the muscles and begin the process of regenerating cells. Such a procedure represents autologous transplantation (in which the donor of the stem cells is the same person as the recipient), in which immune rejection does not occur. This process has been successfully demonstrated in various animal models.

One such example involved Dr. Giulio Cossu of the Stem Cell Research Institute in Milan, Italy, who directed a study in which mesoangioblasts were used to treat MD in a mouse model. The mice showed measurable improvement in their musculature as a result of receiving the stem cells. The findings were published in the journal Science.

Another example involved scientists in Italy and France who collaborated on a study in which dogs with Duchenne MD were treated with autologous mesoangioblasts and consequently experienced a regeneration of dystrophin as well as a regeneration of some of their muscle strength. Results of the study, which was conducted in 2006 and led by Dr. Maurilio Sampaolesi, were published in the journal Nature.

Similarly, researchers at the Children's Hospital in Boston conducted a study with autologous stem cells that were derived from bone marrow and transplanted into mice which were bred to be deficient in dystrophin. The stem cells were found to differentiate into a variety of cell types including muscle cells, with 10% of the muscle fibers expressing dystrophin within 12 weeks after the mice were injected with the bone marrow stem cells.

John Huard, Ph.D., and colleagues at the Children’s Hospital of Pittsburgh, in collaboration with the University of Bonn in Germany, conducted a study in which they isolated a subpopulation of muscle stem cells which were then administered to mice that had been bred to express symptoms of MD. The stem cells were shown specifically to replace dystrophin in the mice, resulting in measurable improvement in the musculature of the mice. The study was funded by the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), the Muscular Dystrophy Association, the Parent Project Muscular Dystrophy Research and the Children’s Hospital of Pittsburgh.

Additionally, muscle-derived stem cells (MDSCs) have been isolated and cloned from postnatal muscle and have been shown to differentiate into muscle tissue and other cell lineages. A NIAMS-supported laboratory has developed a method for isolating MDSCs from young mice and expanding these colonies of cells for more than 200 doublings, during which time the cells retained their ability to synthesize muscle-specific proteins and to develop muscle cell morphology, both in culture and when transplanted back into the mouse muscle tissue. Such a high level of cultural expansion had previously been attributed exclusively to embryonic stem cells, but such properties have now been widely replicated with multiple types of adult stem cells including bone marrow-derived and muscle-derived stem cells.

Adult stem cells offer the same pluripotency of embryonic stem cells, but without the danger of forming teratomas (tumors), which remains a serious risk from embryonic stem cells. It is neither necessary nor desirable to use embryonic stem cells in the treatment of MD or other diseases, since a growing number of studies are showing increasing success with adult stem cells. In fact, the only stem cell studies that have ever shown success in the treatment of any human disease have involved adult stem cells, since no study has ever been conducted in which a disease was successfully treated with human embryonic stem cells, although this fact is not generally reported by the media. Ethics and politics aside, adult stem cells are highly preferable to embryonic stem cells purely for scientific reasons. (Please see the section entitled “Stem Cell Primer” for an explanation of the different properties of the different types of stem cells).

The various muscular dystrophies constitute a tremendous burden, not only to the individuals afflicted with these diseases, but also to their families and indeed to national economies.

Adult stem cell therapy offers a safe and potentially effective treatment of a disease which previously has been considered irreversible.


 

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