|LE Magazine January 2001 |
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The cell cycle
The concept of protein removal brings to mind structural proteins such as collagen that the body breaks down after a relatively long life. In order to understand the implications of proteolytic decline and buildup of aberrant proteins, it is necessary to revise this picture.
Proteasomal decline in Alzheimer's disease
The brain depends upon the proteasome for timely disposal of proteins. When the proteasome is inhibited, oxidized and aggregated proteins accumulate, and neurons degenerate and die. A bottleneck in protein removal could shift the balance toward the accumulation of deposits characteristic of Alzheimer's disease.
A new study at the University of Kentucky provides the first direct evidence of reduced proteasome activity in a neurodegenerative disorder. The scientists compared proteasome activity in five brain regions of normal and Alzheimer's disease brains, using specimens removed during autopsies. They found the activity of the proteasome significantly reduced in three brain regions showing severe degeneration in Alzheimer's disease. By contrast, proteasome activity was not reduced in two brain regions showing less or no degenerative change in Alzheimer's disease (Keller JN et al., 2000).
New research from France suggests a mechanism by which proteasomal impairment could in turn increase production of the amyloid-beta material that makes up senile plaques. Proteins called presenilins influence the production of amyloid-beta from its “parent” protein called amyloid precursor protein, or APP. Mutations in the presenilin genes that lead to early onset Alzheimer's disease upset the balance between production of amyloid-beta and of a neuroprotective derivative of APP called secreted APP. These mutations especially favor production of a long form of amyloid-beta that more readily collects into aggregates and eventually plaques.
There is considerable evidence that presenilin protein concentrations are regulated by the proteasome. The French research shows that inhibition of the proteasome increases production of the long aggregable form of amyloid-beta. Amyloid-beta in turn inhibits the proteasome. The researchers propose that proteasome activators could reverse this imbalance, potentially in sporadic as well as genetic forms of Alzheimer's disease (Checler F et al., 2000; Marambaud P et al., 1998).
Think instead of a highly dynamic population of diverse proteins playing critical roles in the body's regulatory and signaling pathways. These proteins must be selectively synthesized at just the right moment so that they can do their precisely timed jobs, then they must be swiftly degraded at the correct point in a tightly regulated sequence of events. Normally such processes run like clockwork, but when damaged proteins accumulate the system can bog down.
It is in this way that physiological processes—both fast and slow—could become deranged by excessive buildup of proteins marked for removal by the proteasome-ubiquitin system. A case in point is the cell cycle. It consists of four phases culminating in mitosis (cell division). The key steps in this cycle are controlled by proteasomal degradation of proteins. For example, entry into the DNA synthesis phase, separation of sister chromatids, and the exit from mitosis are all dependent upon the timely removal of proteins such as cyclins by the proteasome (Hershko A, 1997; Ford HL et al., 1999).
By inhibiting the proteasome, carbonylated proteins could interfere with cell cycle progression and control. To understand how this can happen, consider an engine whose oil isn't changed regularly. When the detergent in the oil is used up, contaminants precipitate and sludge forms on vital engine parts. The sludge accumulates, impairing engine performance, until finally the engine dies.
The body too needs an efficient sludge removal system. When protein “sludge” accumulates, the gears of the cell cycle can get clogged up. This could impair the efficiency of cell division, and perhaps more importantly, enable damaged cells to reproduce. The result is increasing chromosomal instability, leading to degeneration and cancer (Schmutte C et al., 1999). Another possible outcome is cellular senescence, when the cell cycle grinds to a halt. Protein carbonylation thus becomes a potentially terminal condition.
Cell cycle control represents one more pathway along which damage to, and inefficient removal of protein could contribute to both cellular mortality and the cellular immortalization seen in cancer. In this scenario, the buildup of carbonylated protein feeds a vicious circle of proteasomal impairment, chromosomal instability, and increasing numbers of defective and senescent cells, which the body cannot remove. Insofar as the cellular life cycle is bound up with the life cycles of proteins, it behooves us to maintain healthy intact proteins and to ensure their timely turnover.
While the epidermis (outer skin layer) changes only subtly with age, profound changes take place in the dermis (inner skin layer).
The aging processes discussed above—cellular senescence, protein carbonylation and proteasomal decline—play leading roles in the changes that aging brings to the skin. While the epidermis (outer skin layer) changes only subtly with age, profound changes take place in the dermis (inner skin layer). There, the population of fibroblasts (connective tissue cells) is cut in half by age 80. Collagen becomes disorganized with broken fibers, while the extracellular matrix shows widespread destruction (West MD, 1994).
Protein carbonylation damages all components of the epidermis and dermis, leading to loss of elasticity, wrinkles, macromolecular disorganization, loss of extracellular matrix and reduced capacity for wound repair—all of which are primary characteristics of aged skin. Protein carbonylation rises with age in the epidermis and in cultured keratinocytes (dividing cells that migrate into the epidermis). As elsewhere in the body, it results from protein oxidation, glycation (protein-sugar reactions) and reactions with lipid peroxidation products (Petropoulos I et al., 2000).
Collagen, the protein substance of connective tissue, tends to cross-link with age. It is well known that collagen is cross-linked in the course of glycation and the consequent formation of advanced glycation end products (AGEs). This robs the skin of elasticity and youthful tone. Recent laboratory research demonstrates that this problem can be self perpetuating. Once AGEs form, they can directly induce the cross-linking of collagen—even in the absence of glucose and oxidation reactions (Sajithlal GB et al., 1998). The researchers found that neither antioxidants nor metal chelators could inhibit direct cross-linking of collagen by AGEs. Only an anti-glycating agent, in this case the drug aminoguanidine, could inhibit this process. The natural dipeptide carnosine offers a superior efficacy and toxicity profile compared to aminoguanidine (Munch G et al., 1997; Preston JE et al., 1998; Burcham PC, 2000).
The researchers propose that reactive carbonyl compounds have the ability to induce collagen cross-linking regardless of oxidative conditions. Their findings underline the importance of preventing protein carbonylation and in particular glycation before the cycle of collagen cross-linking gets started.
The dynamic fibroblast
Connective tissue cells, called fibroblasts, play the leading role in the ongoing regeneration of the dermis. In order to function properly, fibroblasts must strike a delicate balance between destruction of extracellular protein and synthesis of new protein.
Normally fibroblasts are quiescent, dividing at a low rate. They produce only small amounts of the matrix metalloproteinase enzymes (collagenase and stromelysin) that break down the surrounding extracellular matrix, and large amounts of matrix metalloproteinase inhibitors (TIMP-1 and TIMP-2). But in response to various stimuli including wounding and inflammation, they undergo a drastic transformation into activated fibroblasts. They then secrete large amounts of enzymes that break down collagen and destroy the extracellular matrix.
New research confirms that proteasome activity declines in keratinocytes and epidermal cells with age.
Cellular senescence locks fibroblasts and keratinocytes into an approximation of this activated state (West MD, 1994). They switch from a matrix-producing to a matrix-degrading mode, secreting more matrix metalloproteinases and less matrix metalloproteinase inhibitors. Senescent fibroblasts and keratinocytes are known to accumulate in aging skin, as demonstrated by a biomarker of skin cell senescence (Dimri GP et al., 1995). In addition to breaking down the extracellular matrix, they secrete proinflammatory mediators such as interleukin-1 alpha and growth factors such as heregulin (regulator of breast and epithelial cell growth) whose influence extends far beyond the cell secreting them (Campisi J, 1998; Campisi J, 1997).
Proteolysis of connective tissue is a normal part of skin cell development and wound healing. Proteolytic enzymes and their inhibitors sculpt structural proteins and break them down at the appropriate times. Unfortunately, as aging skin cells senesce and increase their proteolytic activity, the proteasome (the main enzyme complex for protein degradation) enters an age-related decline. The balance between protein creation and destruction is again upset, compromising the integrity and regeneration of skin tissue.
New research confirms that proteasome activity declines in keratinocytes and epidermal cells with age. At the same time, protein carbonyl levels are rising and the increasing numbers of senescent cells are secreting more proteolytic enzymes. This has been demonstrated clearly in keratinocytes, both in culture and in specimens from humans, where there is an inverse relationship between proteasome content and biomarkers of cellular senescence (Petropoulos I et al., 2000).
The skin makes visible the changes that occur throughout the body as damaged proteins and senescent cells accumulate. As we have seen, the life cycles of cells and proteins may regulate both our appearance as we age and how long we live. Preserving the integrity and regular turnover of protein is thus a key defense against the downward spirals of degeneration in the later years. Carnosine is the only agent that has shown multi-modal protective effects against protein degradation and cellular senescence.
References on Page 3