A. Leövey

Effect of elastin peptides on human monocytes: Ca2+ mobilization, stimulation of respiratory burst and enzyme secretion.

The effect of elastin peptides (Kappa-elastin) was investigated on human monocytes. The data presented here indicate that elastin peptides increase the intracellular Ca2+ level measured by Quin 2 fluorescence and mediate the release of beta glucuronidase and elastase. The O2 consumption and H2O2 release were stimulated in a dose-dependent manner. The early rise of cAMP was followed by a return to the original level at 30 min and by a concomitant increase of cGMP level. The action of elastin peptides on intracellular calcium level and cGMP levels may well be related to its previously demonstrated chemotactic activity. These activities may well play a role in the modifications of the extracellular matrix following elastin degradation as observed in atherosclerosis, emphysema and aging.

Age related variations of some polymorphonuclear leukocyte functions

The age-related alterations of some biochemical processes in polymorphonuclear leukocytes (PMNL) obtained from 20 healthy aged male and 20 healthy aged female (age: 60-94 years) subjects were investigated. The basic level of luminol dependent chemiluminescence (LDCL) was increased, whereas the basal value of reduced/oxidized glutathione (GSH per GS-SG) was decreased in the aged groups. The enhancement of LDCL was more significant by PMNLs of aged males than of females. The change of both GSH and GS-SG levels during phagocytosis were diminished in the PMNLs of aged subjects. The L-alanine beta-naphthylamide specific elastase like protease (ELP) activity measured in living cell suspension was markedly increased with aging in male subjects. Therefore, it was concluded that the aging of PMNLs is partly a sex related process. The intensive release of both beta-glucuronidase (beta-G) and ELP by the PMNLs of aged subjects after in vitro treatment with calcium ionophore A23187, Cytochalasin B or low density lipoprotein (LDL) suggests that they could play an important role in some age-related disorders.

The role of technetium-99m methoxyisobutylisonitrile scintigraphy in the differential diagnosis of cold thyroid nodules

Abstract. Various diagnostic techniques have been successfully used in the clinical management of cold nodules; however, the decision on whether to employ surgery or a conservative treatment is not always easy. This study was designed to appraise the diagnostic value of technetium-99m methoxyisobutylisonitrile (MIBI) scintigraphy in the assessment of cold nodules detected using 99mTc-pertechnetate. Fifty-two patients were included in the study. All had already been selected for surgery, based on their clinical and laboratory findings, including fine-needle aspiration biopsy. The total number of cold nodules on 99mTc-pertechnetate scans was 59. The thyroid scan was performed 20-40 min after i.v. injection of 400 MBq of 99mTc-MIBI. Uptake of MIBI in thyroid nodules was compared with that in the surrounding normal thyroid tissue, and a score of between 0 and 3 was assigned to each nodule as follows: 0, cold; 1, decreased; 2, equal; 3, hot. Definitive histology revealed nodular goitre in 24 cases, adenoma in 19, thyroiditis in 1, differentiated cancer in 12, medullary cancer in 2, and anaplastic cancer in 1. None of the degenerative nodules were hot on MIBI scan, while the adenomas showed a variety of MIBI imaging patterns, most frequently the score 3 pattern. In the diagnosis of differentiated thyroid cancer the sensitivities of score 3 and score 2+3 MIBI uptake patterns were 83% (10/12) and 100%, respectively. The score 3 MIBI uptake pattern had a specificity of 100% and a positive predictive value of 100% with respect to thyroid (benign and malignant) neoplastic diseases, whereas a specificity of 72% and a positive predictive value of 43% were observed in the detection of differentiated cancer. After a cold nodule had been detected using 99mTc-pertechnetate, a second scan with high MIBI uptake increased by 7.8 times the probability that this nodule would be a differentiated cancer. In conclusion, 99mTc-MIBI scintigraphy is a useful method in the differential diagnosis of cold thyroid nodules if the primary aim is to differentiate degenerative from neoplastic diseases rather than to differentiate benign from malignant nodules. High MIBI uptake considerably increases the probability of a differentiated thyroid cancer and facilitates immediate surgical removal, while decreased uptake actually excludes it. We suggest a combination of fine-needle aspiration biopsy and MIBI scan as a routine diagnostic approach to cold thyroid nodules.

Age-dependent alterations of human recombinant GM-CSF effects on human granulocytes

The granulocyte-macrophage colony stimulating factor (GM-CSF) is an important in vivo regulator of granulopoiesis and neutrophil functions. It is well-known that the immune response and the transmembrane signalling in immune cells change with aging. We wished to elucidate the effects of GM-CSF in itself and in priming the activities of other inflammatory agents on neutrophils of elderly persons. Neutrophils of 20 healthy elderly (aged 60-90 years) and 20 healthy young (aged 20-25 years) subjects were studied for superoxide anion production, intracellular free calcium mobilization, antibody dependent cellular cytotoxicity (ADCC) and intracellular killing activities. It was found that GM-CSF is unable to prime neutrophils of elderly subjects to the action of FMLP, metenkephalin or opsonized zymosan. By the use of Pertussis toxin and H7 it was demonstrated that a different signal transduction pathway in neutrophils of elderly subjects is activated by GM-CSF or FMLP if compared to that of young subjects. These results suggest that the lack of priming could contribute to the greater susceptibility of the elderly to infections and that the change of the signal transduction mechanism in neutrophils of elderly subjects might partly explain this phenomenon.

Altered phosphatidylinositol breakdown after K-elastin stimulation in PMNLs of elderly.

The degradation of elastic fibres during atherosclerotic plaque formation in arterial wall is a well known process. The liberated elastin peptides such as K-elastin possess various biological activities: They are chemotactic for monocytes and fibroblasts, stimulate the oxidative burst and the intracellular free Ca2+ mobilisation through the phosphatidylinositol (PIP2) breakdown in PMNLs. It was found that the PIP2 breakdown induced by K-elastin is a pertussis toxin sensitive process in PMNLs of young subjects. In the case of the elderly, the K-elastin-induced oxidative burst, intracellular free Ca2+ elevation was less than in young, and could not be inhibited by pertussis toxin. Studying the K-elastin-induced inositol phosphate (IP) formation in PMNLs of elderly a disturbed PIP2 breakdown was found. K-elastin stimulated the IP formation at a very low level in PMNLs of elderly. This alteration of the second messenger formation (e.g. IP3 and Ca2+) after KE stimulation, might be the consequence of their originally elevated levels in resting PMNLs of elderly.

Age-related changes in cAMP and cGMP levels during phagocytosis in human polymorphonuclear leukocytes.

Alterations of cyclic nucleotides have been studied during the incorporation of opsonized yeast cells into human polymorphonuclear leukocytes (PMNL) from both young and aged subjects. In PMNLs obtained from healthy young males the cAMP level rose to a maximal value during the first 15 min and returned to the starting level at the 60th min of phagocytosis. The cGMP level started to rise only at the 30th min of incorporation and enhanced progressively up to the 120th min. In contrast, the elevated cAMP level remained increased during the 120 min of phagocytosis in PMNLs obtained from aged subjects, whereas the cGMP level was not altered during the same period. The basic level of cAMP diminished, while the cGMP level was found to be elevated with ageing in PMNLs. It is concluded that phagocytosis was not impaired with age yet cytotoxic functions were diminished and that the data presented support the idea that cyclic nucleotides may be directly involved only in the latter process.

Cell surface markers, inositol phosphate levels and membrane potential of lymphocytes from young and old human patients

It is well known that most physiological functions change with aging, including the immune response. Data concerning the aging of lymphocyte subpopulations are conflicting. The antigen density of peripheral blood lymphocytes has been determined by fluorescently tagged OKT-3, OKT-4, OKT-8, OKT-11 and OKM1 monoclonal antibodies in a carefully selected aged (over 87 years) population, and compared to that of young subjects. A substantial difference was found in the percentage distribution of OKT8 and OKM1 subsets. The volume of lymphocytes of the elderly population was significantly less than that of the young. The effect of various monoclonal antibodies on phosphatidylinositol breakdown has also been studied. It was found that only OKT3, acting through the CD3 antigen receptor, was able to induce inositol phosphate formation in both young and elderly, although in the latter population this occurred at a lower level. Because the plasma membrane plays a regulatory role in this process, an important and sensitive functional parameter, the membrane potential, was also monitored and influenced by changing the extracellular K+ concentration. The lymphocytes of the elderly population responded less sensitively to changes in extracellular potassium concentration.

Age-Dependent Variations of Intralysosomal Enzyme Release from Human PMN Leukocytes under Various Stimuli

The intralysosomal beta glucuronidase and elastase release from polymorphonuclear leukocytes (PMNL) of young and aged male subjects were determined after 60-min incubation with 10 micrograms/ml Cytochalasin B (CB), 10(-6) M of Ca ionophore A 23187 and various concentrations of human low density lipoprotein (LDL). The beta glucoronidase secretion was triggered by both A 23187 and LDL; however, no significant differences were found between the enzyme release from PMNLs of young and aged subjects. In contrast, a marked elastase release was triggered in the young group only by LDL, whereas in the aged group, all of the applied drugs induced a significant elastase release. LDL caused the most dramatic enzyme release from PMNLs of aged males. It was concluded that the release of PMNL-elastase after LDL incorporation as well as by CB and Ca ionophore stimulation may be an age-related process.

Blood laboratory parameters of carefully selected healthy elderly people

Hematological and biochemical parameters were measured in carefully selected healthy elderly (50 males and 50 females, aged 60-97 years) and young (10 males and 10 females, aged 18-23 years) populations. Most of the parameters measured did not show any age-related variations. It means that any deviation from the present normal ranges established for healthy young subjects should be considered as pathologic in the case of elderly, too. Some parameters showed age-specific changes and hence there is a possibility that their actual normal ranges might be extended towards the upper limit (globulin, IgG, IgA, CIC, C3, BUN, AP for females), or towards the lower limit (creatinine clearance, albumin). Verification of these modifications may indicate that a lot of unnecessary investigations in the case of elderly could be avoided.

Immunology of elastin: study of anti-elastin peptide antibodies by DOT immunobinding assay

In order to further investigate the role of the immune system in the arteriosclerotic process, we investigated the anti-elastin peptide antibodies (AEAb) of the IgG and IgM types by DOT immunobinding assay in the sera of patients suffering from various arteriosclerotic diseases. In total 232 control and pathological sera were studied. In obliterative arteriosclerosis of the legs 90%, ischemic heart disease 67% and hypertension 60% of sera were positive for AEAb of the IgG type independent of age. In the case of diabetes mellitus, however, the duration of the disease was determinant. In rheumatoid arthritis, the results were negative. No clear-cut positivity could be demonstrated in stroke patients either. These results indicate that AEAb can be detected in some diseases and DOT appears to be an appropriate method for the AEAb screening in various diseases.