Biomedical subjects

G Arienti

Publications and source records attributed to G Arienti.

At least 19 recordsLinked to original sources

PubMed2001

Lipid fatty acid and protein pattern of equine prostasome-like vesicles.

The semen of several mammals contains vesicles of different composition and origin. We have recently reported on the presence of lipoprotein vesicles in stallion semen. To a certain extent, these resemble human prostasomes, but differ from them in amount and composition. These horse-semen prostasome-like vesicles may be important, not only in horse reproductive physiology, but also in view of stallion semen cryopreservation. In this paper, we have studied horse-semen prostasome-like vesicles and found that they possess less saturated fatty acid than human prostasomes. Moreover, their protein pattern (SDS-PAGE electrophoresis) shows that the 30-50-kDa fraction is less abundant in stallion vesicles. In addition, fluidity (measured as fluorescence anisotropy of diphenylhexatriene) is higher in horse prostasome-like vesicles than in human prostasomes, albeit being much lower than that of most membranes. These findings may be connected to some species-related differences in reproductive physiology: the vaginal milieu of the mare is not acidic and the deposition of semen is intrauterine in the horse but vaginal in humans.

Animals↗

PubMed2001

Progesterone-induced increase of sperm cytosolic calcium is enhanced by previous fusion of spermatozoa to prostasomes.

Ejaculated spermatozoa must undergo a number of modifications before becoming able to fertilize the oocyte. The interaction of sperm with other semen components may influence these phenomena; human semen contains vesicles of prostatic origin, called prostasomes that may fuse to sperm at slightly acidic to neutral pH values. Prostasomes contain calcium and it has been demonstrated that their fusion with spermatozoa produces a transient increase (wave) of [Ca(2+)](i) in these cells. The fusion process also transfers protein and lipid to spermatozoa. These phenomena may induce long-lasting changes of sperm properties. We test the hypothesis that spermatozoa, as modified by fusion, change their ability to undergo the progesterone-induced increase of [Ca(2+)](i) and we find that the increase of [Ca(2+)](i) produced by the fusion with prostasomes and by the stimulation with progesterone are independent and additive phenomena. We also find that spermatozoa present a stronger response to the progesterone-induced increase of [Ca(2+)](i) if they are previously made to fuse with prostasomes. This effect does not depend directly on the [Ca(2+)](i) increase due to fusion, since it is still present after the [Ca(2+)](i) has returned to resting values.

Adult↗

PubMed2000

Determination of S-nitrosohemoglobin using a solid-state amperometric sensor.

Nitric oxide (NO, nitrogen monoxide), generated in biological systems, plays important roles as a regulatory molecule. Its ability to bind to hemoglobin (Hb) iron is well known. Moreover, it may lose an electron, forming the nitrosonium ion, involved in the synthesis of nitrosothiols (RSNO). It has been suggested that S-nitrosohemoglobin (SNO-Hb) may act as a reservoir of NO. The S-nitrosylation of Hb can be detected after the incubation of CysNO and Hb for 60 min with a molecular ratio (CysNO/hem) of 1:1. Upon increasing the ratio to 10:1, about 50% of total Hb (100% of beta-chain -SH 93) was derivatized in 60 min. In this paper, we describe a new method for the quantitative assay of SNO-Hb, after the liberation of NO by Cu(2+)/Cu(+) and the simultaneous assessment of NO by solid-state amperometric sensor. The assay described by us is sensitive, rapid, easy to perform, and inexpensive. For this reason, we believe that it may represent an important analytical improvement for the study of the S-transnitrosylation reactions between RSNO and the Hb Cys-beta 93 and SNO-Hb and glutathione.

Biosensing Techniques↗

PubMed1999

Distribution of lipid and protein in human semen fractions.

Human semen is formed by the secretions of different glands. We fractionated semen by centrifugation and obtained four main fractions: (a) spermatozoa, (b) material precipitating at 10¿ omitted¿000xg, (c) prostasomes (precipitate at 105¿ omitted¿000xg), and (d) a soluble fraction. When required, fractions were purified further. We find that most semen protein (about 85%) is in the soluble fraction, 7% in spermatozoa and the remainder is scattered in the other fractions. We compared the electrophoretic pattern of soluble protein with the protein of prostasomes and found marked differences. On the other hand, prostasomes, that comprises only about 3% of total semen protein, contain about 45% of cholesterol and almost 15% of lipid phosphorus with a cholesterol to phospholipid molar ratio greater than 2. On the contrary, phospholipid is largely bound to the fraction containing spermatozoa (about 46% of total lipid phosphorus). This fraction is poor in cholesterol and has a cholesterol to phospholipid molar ratio of about 0.2. The distribution of lipid phosphorus among lipid classes shows some similarity in the soluble fraction and in prostasomes; in both fractions, sphingomyelin is the most abundant phospholipid (about 50%). On the other hand, phosphatidylcholine is the main phospholipid in spermatozoa-enriched fractions (about 35% of total lipid phosphorus). We conclude that the various fractions of seminal plasma obtained by centrifugation differ markedly from each other as to lipid and protein content.

Adolescent↗

PubMed1999

Increase of human spermatozoa intracellular Ca2+ concentration after fusion with prostasomes.

Prostasomes are membranous vesicles (150-200 nm diameter) present in human semen. They are secreted by the prostate gland and contain large amounts of cholesterol, sphingomyelin and calcium, and some of their proteins are enzymes. Prostasomes are involved in a number of biological functions. In previous work, we discovered that prostasomes may fuse to sperm at neutral or at slightly acidic pH values. This mechanism may deliver calcium to sperm, thereby influencing sperm functions. We measured sperm [Ca2+]i with the fura-2 AM method and found that it increased after mixing prostasomes and sperm at pH values allowing fusion (pH 5-7). The increase of [Ca2+]i was proportional to the extent of fusion as measured through the relief of R18 self-quenching. We also examined the increase of sperm [Ca2+]i and the extent of fusion as a function of sperm to prostasome ratio and, also in this case, there was proportionality between the extent of fusion and the increase of [Ca2+]i that reached its maximal values in about 10-20 min. However, a detectable increase of [Ca2+]i was attained after 2 min of fusion. This would represent a new mechanism to influence sperm [Ca2+]i besides ion-exchange systems and ATP-dependent pumps. The value of [Ca2+]i remained elevated, unless Na+ was also present in the external medium. Therefore, the mechanism of fusion might influence deeply the physiology of sperm by producing a transient increase of [Ca2+]i.

Adolescent↗

PubMed1999

The motility of human spermatozoa as influenced by prostasomes at various pH levels.

Human semen contains several components among which spermatozoa, membranous vesicles called 'prostasomes', secreted by the prostate gland and unorganized material. Prostasomes possess an unusual lipid composition, contain a number of proteins and small molecules and have been claimed to take a part in the immune response, in seminal fluid liquefaction and in sperm motility. Since sperm may come in contact with an acidic environment in the vagina, it may be of some interest to know whether prostasomes may affect spermatozoon motility or may protect spermatozoa upon the exposure to an acidic milieu. Human semen was supplied by donors. From whole semen we collected spermatozoa by centrifugation and used the supernatant to prepare prostasomes (centrifugation at 105,000 g for 120 min, followed by purification step on Sephadex G 200); spermatozoa were then collected by a swim-up procedure and exposed to an acidic pH medium (from 5 to 7) in the presence or absence of prostasomes. Spermatozoa motility was subsequently assessed with a superimposed image analysis system (SIAS). Results indicate that the motility of spermatozoa was affected by the pH value of the medium. Acidic media reduced the percentage of motile cells and decreased the straight line velocity of spermatozoa (VLS). Prostasomes had a protective effect and increased the percentage of motile cells. However, they did not change the characteristics of motility (curvilinear and straight). Prostasomes may be considered as a system for counteracting the negative effects of acidic pH values that may be present in the vagina after coitus.

Adult↗

PubMed1998

Fatty acid pattern of human prostasome lipid.

Prostasomes are organelles of prostatic origin found in human semen. Their average diameter is about 150 nm and they appear as a lipoprotein membrane surrounding less organized material. Their lipid composition is peculiar, having much cholesterol and sphingomyelin. On the other hand, many of their proteins possess catalytic activity and are involved in the immune response. In previous work, we have shown that prostasomes may fuse to sperm at slightly acidic pH values, thereby modifying the composition of the sperm plasma membrane. In this paper, we examine the fatty acid pattern of prostasome lipid and find that it is completely different from that of sperm membrane lipid. Polyunsaturated phosphatidylcholines, common in sperm membrane, are rare in prostasome. Therefore, the fusion between prostasomes and sperm should stabilize sperm plasma membrane by enriching it in cholesterol, sphingomyelin, and saturated glycerophospholipid. This would prevent the untimely occurrence of the acrosome reaction.

Cytoplasmic Granules↗

PubMed1998

Interactions between prostasomes and leukocytes.

Prostasomes are membranous vesicles (150-200 nm diameter) present in human semen. They are secreted by the prostate gland and contain large amounts of cholesterol, sphingomyelin and Ca2+. In addition, some of their proteins are enzymes. Prostasomes enhance the motility of ejaculated sperm and are involved in a number of biological functions. In a previous work, we found that prostasome can fuse to spermatozoa at slightly acidic pH values, as demonstrated by the transfer of the lipophilic octadecylrhodamine probe. In this paper, we study the interactions of two leukocyte populations (polymorphonuclear and mononuclear) with prostasomes and find a pH-dependent adhesion (revealed by microscopic observation), but no fusion. These phenomena may be relevant for the functions of leukocytes in human reproduction.

Adult↗

PubMed1998

A new assay for the determination of low-molecular-weight nitrosothiols (nitrosoglutathione), NO, and nitrites by using a specific and sensitive solid-state amperometric gas sensor.

Nitric oxide (NO) is generated in biological systems and plays an important role as a bioregulatory molecule. Its ability to bind hemoglobin and myoglobin is well known. Moreover, it may lose an electron forming the nitrosyl group involved in the formation of S-nitrosothiols. The main problem in analyzing NO is its extreme reactivity. We have tackled this task by using an amperometric sensor to determine free NO, S-nitrosothiols (such as S-nitrosoglutathione), and nitrite in cell-free systems and murine microglial cell cultures. The determination of nitrosothiols is of biochemical relevance and a difficult task particularly at low concentration values. In this article we describe a new method based on the reductive cleavage of the S-NO bond by cuprous ions followed by a solid-state amperometric determination. The system described by us is sensitive, rapid, does not require previous purification steps, is easy to perform, and is inexpensive. For this reason, we think that it may represent an important analytical improvement. It has been suggested that nitrosothiols may exert biological activity by acting as a reservoir of NO. We tested the production of nitrite and of RSNO in stimulated, cultured murine microglial cells and we have shown that nitrite accumulates in these conditions. GSNO also accumulates, provided that GSH is present in the medium.

Animals↗

PubMed1998

Prostasome-like particles in stallion semen.

Human semen contains membranous vesicles called prostasomes. They are secreted by the prostate gland and contain large amounts of cholesterol, sphingomyelin, and Ca2+. Prostasomes enhance the motility of ejaculated spermatozoa and are involved in a number of additional biological functions. No prostasome-like vesicles have been described in horse semen up to now. We have demonstrated the presence of prostasome-like vesicles in the equine semen and characterized them as to size, morphology, and lipid composition; we have found that they are similar to human prostasomes in many respects. We propose that these vesicles might be important for the fecundity of horse semen. This is of interest since the success of artificial insemination is limited by the fact that stallion sperm barely survive cryopreservation.

Animals↗

PubMed1997

Prostasome to sperm transfer of CD13/aminopeptidase N (EC 3.4.11.2).

Prostasomes are membranous vesicles (150-200 nm in diameter) that are present in human semen. They are secreted by the prostate gland and contain large amounts of cholesterol, sphingomyelin and Ca2+. In addition, some of their proteins are enzymes. Prostasomes enhance the motility of ejaculated spermatozoa and are involved in a number of additional biological functions. In previous papers, we demonstrated that lipid can be transferred from prostasomes to sperm by a fusion process occurring at slightly acidic pH. CD (cluster antigens) are ubiquitous proteins; in this paper, we demonstrate that CD13/aminopeptidase N is present is semen, where it is bound to prostasomes. Upon mixing prostasomes and sperm at slightly acidic pH (7 or less), aminopeptidase is transferred from prostasomes to sperm. This evidence comes from enzymatic activity determinations and from the use of the monoclonal antibody, anti-human CD13. The transfer was about 8% of total prostasomal activity at pH 5 and with a prostasome to sperm ratio of 2 (on a protein basis). The transfer did not occur at pH 8.0, but was measurable at pH 7. Therefore, this mechanism may represent a means of modifying the composition and the biological properties of ejaculated sperm.

Antigens, CD↗

PubMed1997

Transfer of aminopeptidase activity from prostasomes to sperm.

Prostasomes are membranous vesicles (150-200 nm diameter) present in human semen. They are secreted by the prostate and contain large amounts of cholesterol, sphingomyelin and Ca2+. In addition, some of their proteins are enzymes. Prostasomes enhance the motility of ejaculated spermatozoa and are involved in a number of additional biological functions. It has been demonstrated that lipid can be transferred from prostasomes to sperm by a fusion process occurring at slightly acidic pH. In this paper, we show that an aminopeptidase activity is transferred from prostasome to sperm. This may be of particular interest since it indicates the involvement of protein in the process of fusion and because sperm may acquire new membrane-bound proteins by this procedure.

Aminopeptidases↗

PubMed1997

Fusion of sperm with prostasomes: effects on membrane fluidity.

Prostasomes are membranous vesicles (150-200 nm diameter) present in human semen. They are secreted by the prostate gland and contain large amounts of cholesterol, sphingomyelin, and Ca2+. In addition, some of their proteins are enzymes. Prostasomes enhance the motility of ejaculated sperm and are involved in a number of biological functions. In this work, we study the fusion of prostasomes to sperm by determining the relief of octadecylrhodamine self-quenching and the fluidity of membranes by measuring the fluorescence anisotropy of diphenylhexatriene. We present the following findings: (a) the contact of sperm cells with prostasomes at slightly acidic pH causes the fusion of the membranes; (b) the amount of transferred lipid depends on the prostasome/sperm ratio; (c) the fluidity of sperm is much higher than that of prostasomes; (d) the fusion changes some properties of sperm cells, such as fluidity, which decreases greatly; and (e) the extent of fluidity variations depends on the prostasome to sperm ratio. We propose that the H(+)-dependent fusion of prostasomes to sperm may have physiological consequences. In fact, this process can modify the lipid and protein pattern of sperm plasma membranes.

Humans↗

PubMed1997

Transfer of CD26/dipeptidyl peptidase IV (E.C. 3.5.4.4) from prostasomes to sperm.

Prostasomes are vesicles present in human semen. They are secreted by the prostate and contain large amounts of cholesterol and sphingomyelin. Some of their proteins are enzymes. Prostasomes are involved in a number of biological functions. In previous papers we demonstrated that lipid can be transferred from prostasomes to sperm by a fusion process occurring at neutral or slightly acidic pH. In this paper we demonstrate that CD26/dipeptidyl peptidase IV, an enzymatic activity absent in sperm, is transferred to sperm from prostasomes. This may be of particular interest since, by this procedure, sperm may acquire new membrane-bound enzymes and modify the catalytic activity of their surface.

Adult↗

PubMed1997

Fusion of human sperm to prostasomes at acidic pH.

Prostasomes are membranous vesicles (150-200 nm diameter) present in human semen. They are secreted by the prostate and contain large amounts of cholesterol, sphingomyelin and Ca2+. In addition, some of their proteins are enzymes. Prostasomes enhance the motility of ejaculated spermatozoa and are involved in a number of additional biological functions. The possibility that they may fuse to sperm has never been proved. In this work, we studied the fusion of sperm to prostasomes by using various methods (relief of octadecyl Rhodamine B fluorescence self-quenching, fluorescence microscopy and flow cytometry) and we found that it occurs at acidic pH (4-5), but not at pH 7.5 pH-dependent fusion relies on the integrity of one or more proteins and is different from the Ca2+-stimulated fusion between rat liver liposomes and spermatozoa that does not require any protein and occurs at neutral pH. We think that the H+-dependent fusion of prostasomes to sperm may have physiological importance by modifying the lipid and protein pattern of sperm membranes.

Animals↗

PubMed1996

Formylcolchicine bound to lactosaminated serum albumin is a more active antifibrotic agent than free colchicine.

Liver fibrosis was induced by chronically (7 weeks) administering CCl4 to rats. Animals were divided into four groups: (a) controls, (b) treated with CCl4 alone, (c) treated with CCl4 and colchicine and (d) treated with CCl4 and formyl-colchicine bound to lactosaminated serum albumin (FC-LASA). Liver dysfunction was monitored by biochemical tests (alkaline phosphatase [ALP], gamma-glutamyltransferase [gamma GT], aspartate and alanine transaminases [AST and ALT], albumin and total bilirubin). Fibrosis was evaluated by determining hydroxyproline and by microscopic examination. The exposure to CCl4 produced major alterations of liver structure and collagen deposition. These effects were partially counteracted by colchicine and to a greater extent by FC-LASA. Morphological findings paralleled biochemical data. The information reported here indicates that colchicine has an antifibrotic activity on the liver of intoxicated rats and that FC-LASA is more active than colchicine itself as an antifibrotic agent.

Amino Sugars↗

PubMed1996

Red blood cell ghosts are affected by adrenoleucodystrophy.

X-linked adrenoleucodystrophy is a disorder occurring in different clinical forms, characterized by adrenal, gonadal and nervous system dysfunction. The basis of the illness is a derangement of the peroxisomal system necessary to oxidize very long-chain fatty acids that accumulate in various tissues. The diagnosis relies on clinical signs and symptoms and on biochemical findings. The six reported cases presented idiopathic adrenal insufficiency. We measured the lipid composition of red blood cell (RBC) ghosts of patients and control subjects. The distribution of phosphorus among phospholipid classes was unaffected; we could not demonstrate any differences between the fatty acid patterns of RBC membrane, either in total lipid extracts or in separated lipid classes. However, we found an increase in total lipid (both phospholipid and cholesterol), in membrane viscosity and in the Na+/K(+)-dependent ATPase. Therefore, we report four main findings on ghosts in adrenoleucodystrophy patients: (a) very long-chain fatty acids do not accumulate; (b) the lipid-protein ratio increases; (c) fluidity decreases; and (d) the activity of ATPase increases. The last finding is proposed as a possible biochemical marker of the illness. We conclude that adrenoleucodystrophy affects deeply RBC membranes.

Adrenoleukodystrophy↗

PubMed1995

Liver alcoholic cirrhosis and spur-cell (acanthocytic) anaemia. A study of erythrocyte ghost composition and fluidity.

BACKGROUND: The occurrence of spur-cell anaemia in the course of cirrhosis is rare. Alterations of the lipid composition and fluidity of erythrocyte (RBC) ghosts may be present and participate in the phenomenon. METHODS: A 59-year-old male patient with alcoholic cirrhosis developed severe spur-cell haemolytic anaemia before death. We compared his RBC ghosts with those of 10 cirrhotic patients and used a group of 9 healthy subjects as controls. RESULTS: The cholesterol to protein ratio was higher in cirrhotic patients; besides, they had less unsaturated fatty acid. The ratio of phospholipid phosphorus to protein did not change; yet, the distribution of phosphorus among phospholipid classes was altered. No difference in 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy (membrane fluidity) was observed between the ghosts of cirrhotic patients and those of healthy people. However, the ghosts of the patient with spur-cell anaemia were more rigid than those of either group. CONCLUSIONS: The values of most variables of cirrhotic patients' ghosts lay between those of healthy subjects and those of the one who developed spur-cell anaemia. It is concluded that some homeostatic mechanisms must control fluidity during cirrhosis; in some cases alterations are particularly great, and fluidity cannot be maintained.

Acanthocytes↗