Muscle cells have learned to "fall into childhood"
Bioengineers reprogrammed mature muscle tissue
into progenitor stem cells without interfering with the genomeNanonewsnet based on materials from UC Berkeley:
Bioengineers reprogram muscles to combat degeneration
Scientists from the University of California at Berkeley (University of California Berkeley) turned back the clock of mature muscle tissue, achieving its return to an earlier stage of differentiation. In experiments on mice, they showed that de-differentiated muscle progenitor stem cells can be used to repair damaged tissue.
According to Irina Conboy, Associate Professor of Bioengineering at UC Berkeley, this achievement, described in an article in the journal Chemistry & Biology (Inhibitors of Tyrosine Phosphatases and Apoptosis Reprogram Lineage-Marked Differentiated Muscle to Myogenic Progenitor Cells), opens the way to the development of new methods to combat muscle degeneration associated with muscular dystrophy or aging.
Musculoskeletal tissue consists of bundles of elongated muscle fibers, or myofibrils, which are fused together muscle cells – myoblasts. The fusion of individual myoblasts into a muscle fiber is considered the final stage of differentiation of skeletal muscle cells.
"Until now, muscle formation seemed like a one–way trip – from stem cells to myoblasts and muscle fibers, but we managed to reverse the development of multinucleated myofibrils and divide them into separate myoblasts," says Conboy.
The practical application of treatment methods based on pluripotent cells capable of becoming a differentiated cell of any type is associated with many problems. Pluripotent cells can be obtained either from embryonic tissue, a source of ongoing ethical disputes, or from adult differentiated cells returned by de-differentiation to a state similar to that of embryonic stem cells. The latter method produces induced pluripotent stem (iPS) cells, and this technology is based on the delivery of specific genes into the cell.
Pluripotent stem cells can divide almost indefinitely, and without targeted differentiation into a certain type, they quickly form teratomas (tumors from disorganized immature tissues) – a serious problem for using iPS cell transplantation as a treatment method.
The biggest problem, relevant for both embryonic and induced pluripotent stem cells, is that even one such undifferentiated cell, once in the body and starting to multiply, leads to a tumor. Muscle progenitor cells obtained by de-differentiation do not form tumors when transplanted into a living muscle.
Unlike pluripotent stem cells, which can differentiate into any type of adult cells, the fate of tissue-specific stem cells of an adult organism is predetermined. Muscle progenitor cells (progenitors) are destined to become muscle tissue, hepatocyte progenitor cells can only become liver tissue, etc..
In addition, cells with embryonic properties are difficult to differentiate into functional adult tissues, for example, blood, brain or muscles. Therefore, instead of returning cells to a pluripotent state, American scientists focused their attention on the progenitor stage, at which cells are already doomed to become skeletal muscles and can grow and divide in culture. Progenitor cells effectively differentiate into muscle fibers - both in vitro and when injected into damaged muscle.
Stem muscle cells, as a rule, are located next to mature fibers, hence their second name – satellite cells. These cells are at rest and begin to repair or form new tissue only after it is damaged or worn out. This happens more than once during our life, when, for example, when lifting weights, old muscle fibers tear, and satellite cells form a new tissue.
When muscle fiber (A) is damaged, chemical signals activate satellite cells.
They multiply (In), forming one new resting cell and one proliferating one.
Proliferating satellite cells can either form a new muscle fiber (C),
or restore the damaged (D).
However, in patients with Duchenne muscular dystrophy – a genetic disease expressed in muscle degeneration due to a defect in structural proteins and subsequent depletion of muscle stem cells - the tissue repair process is grossly disrupted.
In order to force the multinucleated muscle fibers to reverse their course and divide into separate myoblasts, the scientists acted on differentiated muscle tissue with tyrosine phosphatase inhibitors, giving mature cells a signal to start dividing.
The effect on muscle fibers with tyrosine phosphatase inhibitors gives a signal for the beginning of division, but it can cause too strong changes in them. These cells have already merged into one large structure – they have a common cytoplasm and a common cytoskeleton. If you just "order" them to share, many of them will start dying.
To solve this problem, scientists used inhibitors of apoptosis, or programmed cell death.
As Conboy notes, the use of molecular inhibitors for the de-differentiation of mature tissue is a popular direction in the field of stem cells. "These tiny molecules get into the cell and change the line of its behavior without interfering with the genome," explains Conboy. "The inhibitors are used for just 48 hours – enough time for the muscle fibers to separate into individual myoblasts – and then washed out. Cells continue to live and die like normal ones, so there is no risk of their uncontrolled division leading to tumors."
To prove that the myoblasts they obtained were de-differentiated from mature muscle tissue, and not formed from resting satellite cells accompanying muscle fibers, the scientists genetically labeled the fused muscle fibers with a green fluorescent protein (left).
Now they could make sure that the glowing green myoblasts (on the right) could only come from de-differentiated muscle fibers.
To test the viability of the newly obtained myoblasts, that is, to make sure that they can grow, multiply and merge normally into new muscle fibers, the scientists first cultured them in the laboratory, and then transplanted the de-differentiated myoblasts into the damaged muscles of live mice.
After two to three weeks, they saw new glowing green muscle fibers, which proved that progenitor cells derived from mature muscle tissue contributed to muscle recovery in the animals.
The muscle cells obtained by de-differentiation, fluorescing green, were injected into the muscles of mice and successfully merged to form new muscle fibers (left).
On the right, for comparison, is normal muscle tissue formed by non-experimental muscle cells.
Photo: Preeti Paliwal, UC Berkeley.
According to the scientists, the next steps in their work will be to test this process on human muscle tissue and search for other chemical compounds that can help de-differentiate mature tissue.
"This approach is not for all degenerative diseases," explains Conboy. "It will only work in those diseases where we can start with differentiated tissue, such as neurons or liver cells. But in patients, for example, with type 1 diabetes, there are no islet beta cells of the pancreas that produce insulin, so there is no differentiated functional tissue with which we could start. Our approach is not a replacement for the pluripotent cell method, but an additional tool in the arsenal of cell therapy."
Portal "Eternal youth" http://vechnayamolodost.ru 30.09.2011