Biomedical subjects

K Merker

Publications and source records attributed to K Merker.

9 recordsLinked to original sources

PubMed2001

Proteolysis, caloric restriction and aging.

The nature of the aging process has been the subject of considerable speculation. It is believed that free radical damage to cellular components is one of the main contributors to the aging process. Studies on proteins have shown age-related decline in enzyme activities, age-related accumulation of oxidized proteins and a decline of the proteolytic machinery of the cell. The proteasome, a highly regulated intracellular proteolytic system, is the major enzymatic system responsible for the degradation of damaged proteins. The current knowledge on regulation and of the properties of this unique proteolytic system with special emphasis to the aging process are discussed in this review. Since it is known that caloric restriction (CR) is the only method to delay the aging process and extend the maximal lifespan the effects of CR on the age-related decline in protein degradation is highlighted.

Aging↗

PubMed2001

Lipofuscin accumulation in proliferating fibroblasts in vitro: an indicator of oxidative stress.

The amount of the ageing pigment, lipofuscin, found in replicating cells depends both on its rate of formation as well as its rate of dissolution by cell division. We present a model which allows the calculation of the lipofuscin accumulation rate from measurements of its amount and of the cell cycle duration. In two human fibroblast strains, the accumulation rate correlates well with differences in oxidative stress/antioxidative defence as measured by intracellular peroxide generation, protein carbonyl content, telomere shortening rate and replicative life span. The lipofuscin content increases with replicative age in both cultures. The rather steep increase in presenescent fibroblasts is not solely due to a slowing down of the cell turnover, but is partially caused by an increased rate of lipofuscin formation/ accumulation. This might indicate an increased level of oxidative stress in presenescent fibroblasts, or a decreased efficiency of proteolytic systems, or both. The results are in accordance with data demonstrating an adverse effect of lipofuscin accumulation on cellular protein turnover and suggest an active role for lipofuscin accumulation in cellular senescence.

Biomarkers↗

PubMed2000

Hydrogen peroxide-mediated protein oxidation in young and old human MRC-5 fibroblasts.

It is suggested that the aging process is dependent on the action of free radicals. One of the highlights of age-related changes of cellular metabolism is the accumulation of oxidized proteins. The present investigation was undertaken to reveal the proliferation-related changes in the protein oxidation and proteasome activity during and after an acute oxidative stress. It could be demonstrated that the activity of the cytosolic proteasomal system declines during proliferative senescence of human MRC-5 fibroblasts and is not able to remove oxidized proteins in old cells efficiently. Whereas in young cells removal of oxidized proteins was accompanied by an increase in the overall protein turnover, this increase in protein turnover could not be seen in old MRC-5 fibroblasts. Therefore, our studies demonstrate that old fibroblasts are much more vulnerable to the accumulation of oxidized proteins after oxidative stress and are not able to remove these oxidized proteins as efficiently as young fibroblasts.

Aldehydes↗

PubMed2000

Protein oxidation and degradation during proliferative senescence of human MRC-5 fibroblasts.

One of the highlights of age-related changes of cellular metabolism is the accumulation of oxidized proteins. The aging process on a cellular level can be treated either as the ongoing proliferation until a certain number of cell divisions is reached (the Hayflick limit) or as the aging of nondividing cells, that is, the age-related changes in cells without proliferation. The present investigation was undertaken to reveal the changes in protein turnover, proteasome activity, and protein oxidation status during proliferative senescence. We were able to demonstrate that the activity of the cytosolic proteasomal system declines dramatically during the proliferative senescence of human MRC-5 fibroblasts. Regardless of the loss in activity, it could be demonstrated that there are no changes in the transcription and translation of proteasomal subunits. This decline in proteasome activity was accompanied by an increased concentration of oxidized proteins. Cells at higher proliferation stages were no longer able to respond with increased degradation of endogenous [(35)S]-Met-radiolabeled proteins after hydrogen peroxide- or quinone-induced oxidative stress. It could be demonstrated that oxidized proteins in senescent human MRC-5 fibroblasts are not as quickly removed as they are in young cells. Therefore, our study demonstrates that the accumulation of oxidized proteins and decline in protein turnover and activity of the proteasomal system are not only a process of postmitotic aging but also occur during proliferative senescence and result in an increased half-life of oxidized proteins.

Cell Division↗

PubMed2000

Proteolysis of oxidised proteins and cellular senescence.

Aerobe living is consequently connected with a permanent oxidation of cellular proteins. Free radicals and other oxidants damage the normal intracellular protein pool. Therefore, the prevention of accumulation of oxidised cellular proteins is one of the major functions of the proteolytic machinery of mammalian cells. It is known that the multicatalytic proteinase complex, the proteasome, is the major protease that is able to recognise and degrade oxidised proteins. Cellular models are most useful to investigate biochemical changes of protein catabolism during senescence. Unfortunately, little is known about the protein turnover and the regulation of the proteasomal system as well as under oxidative stress conditions as during senescence. The proteasomal regulation during oxidative stress, protein oxidation and the changes of these processes during the ageing process are highlighted in this review.

Animals↗

PubMed2000

Protein oxidation and degradation during cellular senescence of human BJ fibroblasts: part I--effects of proliferative senescence.

Oxidized and cross-linked proteins tend to accumulate in aging cells. Declining activity of proteolytic enzymes, particularly the proteasome, has been proposed as a possible explanation for this phenomenon, and direct inhibition of the proteasome by oxidized and cross-linked proteins has been demonstrated in vitro. We have further examined this hypothesis during both proliferative senescence (this paper) and postmitotic senescence (see the accompanying paper, ref 1 ) of human BJ fibroblasts. During proliferative senescence, we found a marked decline in all proteasome activities (trypsin-like activity, chymotrypsin-like activity, and peptidyl-glutamyl-hydrolyzing activity) and in lysosomal cathepsin activity. Despite the loss of proteasome activity, there was no concomitant change in cellular levels of actual proteasome protein (immunoassays) or in the steady-state levels of mRNAs for essential proteasome subunits. The decline in proteasome activities and lysosomal cathepsin activities was accompanied by dramatic increases in the accumulation of oxidized and cross-linked proteins. Furthermore, as proliferation stage increased, cells exhibited a decreasing ability to degrade the oxidatively damaged proteins generated by an acute, experimentally applied oxidative stress. Thus, oxidized and cross-linked proteins accumulated rapidly in cells of higher proliferation stages. Our data are consistent with the hypothesis that proteasome is progressively inhibited by small accumulations of oxidized and cross-linked proteins during proliferative senescence until late proliferation stages, when so much proteasome activity has been lost that oxidized proteins accumulate at ever-increasing rates. Lysosomes attempt to deal with the accumulating oxidized and cross-linked proteins, but declining lysosomal cathepsin activity apparently limits their effectiveness. This hypothesis, which may explain the progressive intracellular accumulation of oxidized and cross-linked proteins in aging, is further explored during postmitotic senescence in the accompanying paper (1).

Cell Division↗

PubMed2000

Protein oxidation and degradation during cellular senescence of human BJ fibroblasts: part II--aging of nondividing cells.

Oxidized/cross-linked intracellular protein materials, known as ceroid pigment, age pigment, or lipofuscin, accumulate in postmitotic tissues. It is unclear, however, whether diminishing proteolytic capacities play a role in the accumulation of such oxidized intracellular proteins. Previous studies revealed that the proteasome is responsible for the degradation of most oxidized soluble cytoplasmic and nuclear proteins and, we propose, for the prevention of such damage accumulations. The present investigation was undertaken to test the changes in protein turnover, proteasome activity, lysosome activity, and protein oxidation status during the aging of nondividing cells. Since the companion paper shows that both proteasome activity and the overall protein turnover decline during proliferative senescence whereas the accumulation of oxidized proteins increases significantly, we decided to use the same human BJ fibroblasts, this time at confluency, at different PD levels (including those that are essentially postmitotic) to investigate the same parameters under conditions where cells do not divide. We find that the activity of the cytosolic proteasome declines dramatically during senescence of nondividing BJ fibroblasts. The peptidyl-glutamyl-hydrolyzing activity was particularly affected. This decline in proteasome activity was accompanied by a decrease in the overall turnover of short-lived (radiolabeled) proteins in the nondividing BJ fibroblasts. On the other hand, no decrease in the actual cellular proteasome content, as judged by immunoblots, was found. The decline in the activity of the proteasome was also accompanied by an increased accumulation of oxidized proteins, especially of oxidized and cross-linked material. Unlike the loss of lysosomal function seen in our accompanying studies of proliferative senescence (1), however, the present study of hyperoxic senescence in nondividing cells actually revealed marked increases in lysosomal cathepsin activity in all but the very 'oldest' postmitotic cells. Our comparative studies of proliferating (1) and nonproliferating (this paper) human BJ fibroblasts reveal a good correlation between the accumulation of oxidized/cross-linked proteins and the decline in proteasome activity and overall cellular protein turnover during in vitro senescence, which may predict a causal relationship during actual cellular aging.

Cellular Senescence↗

PubMed1998

Proteasome-dependent degradation of oxidized proteins in MRC-5 fibroblasts.

Fibroblasts were exposed to various concentrations of hydrogen peroxide and the removal of oxidized proteins was followed by determining protein-bound carbonyls. Fibroblasts are able to increase the turnover of metabolically radiolabeled proteins after treatment with hydrogen peroxide. It was demonstrated for the first time, that the increased protein turnover was accompanied by a removal of protein-bound carbonyl groups. The proteasome-specific inhibitor lactacystin was able to inhibit the elimination of protein-bound carbonyl groups. Therefore, the key role of the proteasome in the degradation of oxidized proteins in fibroblasts could be demonstrated.

Acetylcysteine↗

PubMed1990

[Knowledge and ability of patients concerning the administration of nasal drops].

The study tried to find out what patients know on the application of nose drops when leaving the pharmacy. As could be shown knowledge and ability were not satisfying. Two third of the patients applied their nose drops in a correct single dose but in too long intervals. Almost the half did not know the correct technique of application. Only 2% of the patients mentioned the pharmacist as source of their information. The results require a more effective information about the application of drugs by the pharmaceutical staff.

Administration, Intranasal↗