Biomedical subjects

Carlo Bertoni-Freddari

Publications and source records attributed to Carlo Bertoni-Freddari.

At least 19 recordsLinked to original sources

PubMed2006

Synaptic and mitochondrial physiopathologic changes in the aging nervous system and the role of zinc ion homeostasis.

Brain performances, e.g. learning and memory, decay during aging. Deterioration of synaptic junctions, as structural correlates of these key functions of the central nervous system, may play a central role in this impairment. Current research on the age-related changes of synapses is documenting that the numeric loss of contacts appears to trigger a compensatory reaction by the old CNS, i.e. the surviving junctional areas in old individuals are larger than in adult subjects. The final outcome of the balanced changes in synaptic number and size is that the overall synaptic junctional area per cubic micron of neuropil is also reduced in aging and this may account for the age-associated functional decay of CNS performances. Among the suggested determinants of synaptic deterioration in aging, a considerable number of recent studies support an early and pivotal role of the progressive decline of the mitochondrial metabolic competence, i.e. the capacity of select pools of organelles to provide adequate amounts of adenosine triphosphate. Quantitative ultrastructural studies together with cytochemistry of key enzymes of the respiratory chain (cytochrome oxidase and succinic dehydrogenase) have shown that mitochondrial dysfunctions play an early and central role in synaptic deterioration events associated with aging and neurodegenerative diseases. Among the various causes, the multiple mechanisms and molecules involved in zinc ion homeostasis have been supposed to be less efficient in the aging brain. Thus, a transient imbalance of free zinc ion concentration in the cytosol ([Zn2+]i) can be considered an unfavourable trigger of subtle mitochondrial damage and synaptic pathology.

Aged↗

PubMed2006

Cytochemical estimation of cytochrome oxidase activity as a morphofunctional mitochondrial check-up.

Cytochemical estimation of COX activity and morphometric measurement of mitochondrial ultrastructure were carried out in organelles from adult and old rats. Although no age-related difference was found in cytochemical precipitate (CPA) and mitochondrial area (MA), the ratio CPA:MA (R) decreased by 25.7% in aging. R was the same in oversized mitochondria, but in smaller organelles it was significantly decreased during aging. R reports on the functional mitochondrial surface involved in energy providing mechanisms; thus, by revealing age-related differences, these data suggest that the COX preferential cytochemistry associated with morphometry may serve as a reliable and sensitive mitochondrial morphofunctional checkup procedure.

Aging↗

PubMed2006

Testing mitochondrial metabolic competence by cytochrome oxidase preferential cytochemistry versus immunoreactivity of subunits I and IV.

Cytochemically evidenced COX activity was compared with levels of immunohistochemically stained mitochondrial- and nuclear-encoded subunits (CO I and CO IV) in the dentate gyrus outer molecular layer (OML) and cerebellar granular layer (GL) of adult and old rats. COX activity decreased significantly in aging, whereas CO I and CO IV levels were significantly increased both in GL and OML of old animals. These findings suggest that the age-related decay of the mitochondrial metabolic competence is not caused by a reduction of COX subunits levels, but causal events affecting mitochondria as discrete morphofunctional units of the cellular bioenergetic machinery.

Aging↗

PubMed2006

Synaptic pathology in the brain cortex of old monkeys as an early alteration in senile plaque formation.

Synaptic numeric density (Nv), average size (area: S), surface density (Sv) and number of synapses/neurone (Syn/Neur) were morphometrically measured in frontal (FC) and temporal (TC) cortex of adult and old monkeys. Sv was constant, a clear age-related trend to decrease by Nv and increase by S were observed in both areas investigated. Syn/Neur significantly decreased in TC of aged animals (-21.1%), whereas FC showed a not significant reduction (-2.6%). The present data support the hypothesis of an increased sensitivity to deterioration of TC synapses in aged monkeys, which might constitute a predisposing condition to the development of senile plaques.

Aging↗

PubMed2006

Level and distribution of microtubule-associated protein-2 (MAP2) as an index of dendritic structural dynamics.

Optical density of MAP2 immunoreactivity (OD), the ratio between the MAP2 stained area/total test area (area fraction: AF), the total length of MAP2 labeled profiles (TL) and the ratio perimeter/area of the immunostained profiles (pleomorphism index [PI]) were measured by quantitative immunohistochemistry in the brain of rats of different ages. In old rats versus young and adult animals, OD and AF were significantly lower, whereas PI was significantly higher, in dentate gyrus molecular layer, CA1 stratum radiatum and olfactory bulb. These findings lend support to the many converging results on the higher vulnerability to aging of the CNS areas featuring higher plasticity.

Aging↗

PubMed2005

Aging affects the distribution of the circadian CLOCK protein in rat hepatocytes.

Several biochemical, physiological, and behavioral processes exhibit cyclic oscillations of about 24 h, which have been defined as circadian rhythms. In mammals, the primary circadian pacemaker resides in the suprachiasmatic nuclei; however, cell-autonomous circadian oscillators occur also in extraneural tissues, including the liver. CLOCK protein is a transcription factor essential for normal circadian rhythms and recent studies have demonstrated that it undergoes intranuclear redistribution in hepatocytes, along the daily cycle. It is known that aging leads to a progressive deterioration of the circadian rhythm at the behavioral, physiological, and cellular levels; in addition, aging affects the organization of nuclear structural components involved in transcription and splicing. In this view, we carried out ultrastructural immunocytochemical analyses on hepatocytes of adult and old rats, so as to investigate possible qualitative and quantitative modifications of CLOCK protein, in relation to the aging process. Our observations demonstrated that most CLOCK protein was always located in the cell nucleus, where it accumulated on perichromatin fibrils (the sites of premRNA transcription and early splicing); in addition, CLOCK showed daily oscillations in the different nuclear compartments, but these oscillations differed significantly between adult and old animals. This unusual distribution of CLOCK protein during aging could be related to the prolonged diurnal activity of old animals and/or to altered nuclear pathways.

Aging↗

PubMed2005

The effect of chronic physical exercise on succinic dehydrogenase activity in the heart muscle of old rats.

Succinic dehydrogenase (SDH) activity, preferentially evidenced by cytochemical methods, has been measured by computer-assisted morphometry in the heart muscle of old sedentary and age-matched animals chronically undergone physical exercise (20 min, twice a day, 5 days a week). The area of the SDH-positive mitochondria (MA) and the overall area of the cytochemical precipitates due to SDH activity (PA) were semiautomatically measured and the ratios PA/MA as well as MA/overall myocardial tissue area analysed (MA/TA) were the parameters taken into account. No significant difference was found between the two groups investigated as regards PA/MA, whereas the MA/TA value is significantly increased in the animals undergone physical training. The present findings document that chronic physical exercise significantly increases the overall mitochondrial area involved in energy provision in the old myocardial tissue. Considering that myocardial function highly relies on mitochondrial metabolism, our results support a beneficial effect of chronic physical exercise on the old heart muscle.

Aging↗

PubMed2005

Brain, aging and neurodegeneration: role of zinc ion availability.

Actual fields of research in neurobiology are not only aimed at understanding the different aspects of brain aging but also at developing strategies useful to preserve brain compensatory capacity and to prevent the onset of neurodegenerative diseases. Consistent with this trend much attention has been addressed to zinc metabolism. In fact, zinc acts as a neuromodulator at excitatory synapses and has a considerable role in the stress response and in the functionality of zinc-dependent enzymes contributing to maintaining brain compensatory capacity. In particular, the mechanisms that modulate the free zinc pool are pivotal for safeguarding brain health and performance. Alterations in zinc homeostasis have been reported in Parkinson's and Alzheimer's disease as well as in transient forebrain ischemia, seizures and traumatic brain injury, but little is known regarding aged brain. There is much evidence that that age-related changes, frequently associated to a decline in brain functions and impaired cognitive performances, could be related to dysfunctions affecting the intracellular zinc ion availability. A general agreement emerges from studies of humans' and rodents' old brains about an increased expression of metallothionein (MT) isoforms I and II, but dyshomogenous results are reported for MT-III, and it is still uncertain whether these proteins maintain in aging the protective role, as it occurs in adult/young age. At the same time, there is considerable evidence that amyloid-beta deposition in Alzheimer's disease is induced by zinc, but the pathological significance and the causes of this phenomenon are still an open question. The scientific debate on the role of zinc and of some zinc-binding proteins in aging and neurodegenerative disorders, as well as on the beneficial effect of zinc supplementation in aged brain and neurodegeneration, is extensively discussed in this review.

Aging↗

PubMed2004

Age-related decline in metabolic competence of small and medium-sized synaptic mitochondria.

A computer-assisted morphometric investigation of cytochrome oxidase (COX) activity, selectively evidenced by preferential diaminobenzidine cytochemistry, has been carried out on synaptic mitochondria in the cerebellar cortex of adult and old rats. The ratio (R) of the area of the cytochemical precipitate (CPA) to the overall area of each mitochondrion (MA) was calculated. R refers to the fraction of the inner mitochondrial membrane actively involved in cellular respiration, thus its quantitative estimation constitutes a reliable index of the mitochondrial metabolic competence (MMC). In adult rats a significant negative correlation between MA and R values was found, while in old animals there was just a positive trend. Paired-quartile comparisons of R values showed a significant age-related decrease in small and medium-sized mitochondria, whereas the lowest and not significant age-related reduction was found in oversized organelles. A paired decrease in number and increase in size is reported to be a general trend for mitochondria during aging, but oversized organelles, according to their low R value, constitute a scanty, though functional, compensating reaction. Thus, the present findings support the argument that the currently reported age-related cellular metabolic decay appears to rely both on the decline in MMC of the small and medium-sized mitochondria, and on their specific reduction in number. This novel result is of biological relevance since it is largely the small and medium-sized mitochondria that are required for the provision of adequate amounts of ATP for actual cellular performance, while the significantly enlarged organelles are thought to represent an intermediate ultrastructural feature in mitochondrial genesis and/or remodelling.

Animals↗

PubMed2004

Metallothionein isoforms (I+II and III) and interleukin-6 in the hippocampus of old rats: may their concomitant increments lead to neurodegeneration?

Metallothionein (MT)-III isoform is a brain metal-binding protein that, like the MT-I + II isoform, binds zinc with high affinity. In the young-adult age, MT-III isoform increases during transient stress while MT-I + II isoform decreases, suggesting compensatory phenomena between the two isoforms and a protective role of MT-III against oxidative damage. This role may be questioned during ageing, because the stress-like condition is chronic in ageing due to high persistent levels of interleukin-6. In the present study, high expression of MT-III and MT-I + II genes (examined by RT-PCR and in situ hybridisation) was found in the hippocampus of old rats. These results indicate that a large amount of free zinc ions can be sequestered by MT isoforms, leading to impaired zinc-dependent functions in the ageing brain. In addition, zinc (tested with the Timm's method) was found to be low in mossy fibres from the old hippocampus. As this method tests bound and unbound zinc, we also investigated free zinc ion bioavailability based on the ratio active thymulin/total thymulin. We found that zinc ion bioavailability was low in old rats, together with increased interleukin-6 mRNA, high expression of both MT isoforms and reduced number of synapses whose function is zinc-dependent, in the old hippocampus. The results indicate that concomitant increments of both MT isoforms may provoke detrimental synergistic effects leading to reduced free zinc ion bioavailability for synapses. As a consequence, compensatory phenomena between MT isoforms may not occur in the old hippocampus due to chronic stress-like condition elicited by high persistent levels of interleukin-6.

Aging↗

PubMed2004

Pineal graft in old rats improves erythrocyte resistance to peroxyl radical-induced hemolysis.

Pineal graft from young to old rats was performed and red blood cell hemolysis, induced by the water-soluble radical initiator 2,2'-azobis (2-amidino-propane) dihydrochloride (AAPH), was evaluated 6 months after graft. Pineal graft modified the hemolysis curve kinetic profile in grafted rats versus age-matched controls, the 50% hemolysis time as well as the lag time were longer, whereas the maximal amount of hemolysis was lower, and it occurred over a longer period of time. Thiobarbituric acid reactive substances production was lower in pineal-grafted rats than in controls and the age-related decrease of erythrocyte membrane fluidity was prevented by pineal graft. The present findings support an important physiological role of pineal gland in preventing age-related alterations of erythrocyte membranes and suggest a possible antioxidant action of melatonin.

Animals↗

PubMed2004

Decay of mitochondrial metabolic competence in the aging cerebellum.

Cytochemically evidenced cytochrome oxidase activity was morphometrically measured in the cerebellar cortex of adult and old rats. The ratio (R) between the area of the precipitate due to the cytochemical reaction and the overall area of each mitochondrion was calculated. While in adult rats an inverse correlation between mitochondrial size and R values (r = -.905) was envisaged, in old animals increasing values of R were paired by increases in mitochondrial area (r =.561). Paired-quartile comparisons of the R values from adult and old animals documented a marked age-related impairment of the mitochondrial metabolic competence in small (I quartile: -31.6%) and medium-sized (II quartile: -26.4; III quartile: -16.4) mitochondria, while large organelles showed the lowest age-related decrease (IV quartile: -3.0%). The present findings support that a marked dysfunction of small and medium-sized mitochondria contributes to the significant decay of energy metabolism currently reported in physiological aging.

Aging↗

PubMed2004

Cytochrome oxidase activity in hippocampal synaptic mitochondria during aging: a quantitative cytochemical investigation.

Synaptic mitochondria, cytochemically positive to cytochrome oxidase (COX) activity, were investigated by morphometric methods in the hippocampal dentate gyrus of adult and old rats. The number of mitochondria/microm(3) of tissue (Nv), the volume fraction occupied by mitochondria/microm(3) of tissue (Vv), the average mitochondrial volume (V), the longer mitochondrial diameter (F(max)), and the ratio R:mitochondrial area/overall area of the cytochemical precipitate due to COX activity were measured on COX-positive organelles. In old animals, Nv, Vv, V, and F(max) increased at a not significant extent; R was not significantly decreased. The complement (%) of longer organelles was higher in old animals. COX activity is currently considered an endogenous marker of neuronal oxidative metabolism; thus, although our findings refer to the discrete subpopulation of COX-positive organelles located at synaptic terminals, they support that changes of mitochondrial ultrastructure and metabolic competence may contribute to the age-related alterations of neuronal performances.

Aging↗

PubMed2004

Vitamin E deficiency and aging effect on expression levels of GAP-43 and MAP-2 in selected areas of the brain.

The expression levels of GAP-43 and MAP-2, two proteins involved, respectively, in axonal and dendritic remodeling, in control adult (11 months), old (24 months), and vitamin E-deficient (11 months) rats were evaluated. mRNA levels were determined by means of a quantitative in situ hybridization procedure in subregions of hippocampus and cerebellum. Though a general trend can be observed indicating a reduction in GAP-43 expression in aging as compared to adult animals and an increase in vitamin E-deprived rats in comparison with adult animals, no statistically significant change was found in any region analyzed. In the same way, MAP-2 mRNA levels show an increase in vitamin E-deprived rats in comparison with other groups tested; only one variation was statistically significant, namely the increase in cerebellar cortex MAP-2 nRNA levels in vitamin E-deficient versus adult rats. These results suggest that oxidative stress and aging negatively affect neuroplasticity, showing different characteristics at the dendritic and axonal levels.

Aging↗

PubMed2004

Chronic aluminum administration to old rats results in increased levels of brain metal ions and enlarged hippocampal mossy fibers.

The effect of chronic aluminium administration (2 g/L/6 months) was investigated in the central nervous system (CNS) of old rats. The content of Al(3+), Cu(2+), Zn(2+), and Mn(2+) was measured in prosencephalon + mesencephalon, pons-medulla, and cerebellum. The area occupied by the mossy fibers in the hippocampal CA3 zone was also measured. In Al-treated rats the contents of Al(3+), Cu(2+), Zn(2+), and Mn(2+) were significantly increased in prosencephalon + mesencephalon and pons-medulla, while no change was observed in the cerebellum except a Cu(2+) decrease. The area occupied by the mossy fibers in the CA3 field was significantly increased (+32%) in Al-treated rats. Taken together, the present findings document that the aging CNS is particularly susceptible to aluminum toxic effects that may be responsible for a consistent rise in the cell load of oxidative stress. This may contribute, as an aggravating factor, to the development of neurodegenerative events, as observed in Alzheimer disease.

Aluminum↗

PubMed2004

Aging and vitamin E deficiency are responsible for altered RNA pathways.

Fibrillar centers (FCs), dense fibrillar (DFC) and granular (GC) components in nucleoli, and perichromatin granules (PGs) in nucleoplasm were measured by morphometry. FC size and their nucleolar surface fraction significantly decreased in aging and vitamin E deficiency. The GC and DFC nucleolar fraction was unchanged in adult and old rats, but in vitamin E-deficient animals GC increased and DFC decreased significantly. PG density significantly increased in aging and decreased in vitamin E deficiency. The quantitative evaluation of immunolabeled transcription and splicing factors revealed that polymerase II and SC-35 significantly decreased in old and vitamin E-deficient versus adult animals. Fibrillarin and snRNPs did not change between adult and old rats, but were significantly lower in vitamin E-deficient rats. These data document altered RNA pathways in aging and vitamin E deficiency. Considering the antioxidant role of vitamin E, they lend further support to the importance of free radical production and control in the aging process.

Aging↗

PubMed2004

Increased intracellular ionic content is correlated with a decreased perichromatin granule density in old neurons.

The intracellular content of monovalent ions (Na(+), Cl(-), and K(+)) in brain cortical cells from young, adult, and old rats was measured by X-ray microanalysis. By referring the peak of each ion to the respective value of the dry mass, we obtained the intracellular content as a percentage of dry mass. A significant increase of the intracellular content of Na(+), Cl(-), and K(+) was found in old animals. In the same type of cells from rats of different ages, we estimated the density of perichromatin granules (PGs), the RNA structural correlates containing ribonucleoproteins (RNPs). PGs were discovered by Bernhard's method, which allows the preferential staining of RNPs. The PG density (i.e., the number of PGs/microm2 of nuclear area) was significantly lower (-27.8%) in the neurons of old animals. Taken together, the present findings document that in the nuclei of brain cortical cells of old animals the ionic strength is markedly increased and that this alteration may affect chromatin functions (e.g., DNA template activity). The ionic content of the nucleoplasm is reported to modulate the structure of chromatin molecules and regulate gene expression; thus, the increased content of the three ions found by us in old animals, by increasing the viscosity of the nucleoplasm, may affect gene expression by disturbing the inhibition (negative regulation) or potentiation (positive regulation) of RNA polymerase binding. In physiological aging, increased ionic strength may lead to age-related DNA dysfunction as well as to alterations in the processing of RNA structural constituents, such as PGs.

Animals↗

PubMed2004

Increased mitochondrial and nuclear gene expression of cytochrome oxidase subunits I and IV in neuronal aging.

To assess the role of mitochondrial metabolic competence (MMC) in neuronal aging, quantitative immunohistochemistry of cytochrome oxidase (COX) subunits I (mitochondrial-encoded) and IV (nuclear-encoded) was carried out in the cerebellar cortex of adult and old rats. The optical density (OD) values of the immunostained COX subunits I and IV were measured on an overall area of 75,000 microm(2) in the granular and molecular layers of the cerebellar cortex of each animal. In old animals, OD values of subunit I were increased by 35.5 and 34.2% in the molecular and granular layers, respectively, but only the difference found in the latter cerebellar zone was statistically significant (p < 0.05%). As regards subunit IV, old animals showed higher, not significant, densitometric values in the molecular (120.6%) and granular (126.8%) layers. The present findings sustain that gene expression of COX subunits I and IV appears not to be involved in the well-documented time-related mitochondrial decay. The proper functioning of COX depends on several factors that can affect MMC in the aging cell. In the fully assembled holoenzyme, both the subunits I and IV span the inner mitochondrial membrane. On the basis of these molecular biology data, it is reasonable to suppose that any alteration of the physicochemical features and chemical composition of the mitochondrial membranes reported to occur in aging (e.g., decreased membrane fluidity and cardiolipin content, increased cholesterol/phospholipid molar ratio and free-radical damage, etc.) may significantly affect the proper assembling of the enzyme and, in turn, its activity. Considering the reported significant decline of COX activity with advancing age, our findings further support that an adequate mitochondrial metabolic competence, while including proper nuclear and mitochondrial gene expression of subunits of the respiratory chain, relies on the overall balance among various determinants that can be differently damaged by aging and represent critical causative events responsible for the age-related functional decline of selected mitochondrial populations.

Animals↗