Biomedical subjects

A Bertolotto

Publications and source records attributed to A Bertolotto.

At least 37 records · Page 2Linked to original sources

PubMed1999

Quantitative PCR reveals increased levels of tumor necrosis factor-alpha mRNA in peripheral blood mononuclear cells of multiple sclerosis patients during relapses.

Quantification of tumor necrosis factor-alpha (TNF-alpha) mRNA in peripheral blood mononuclear cells (PBMC) could provide information about disease activity in multiple sclerosis (MS); however, specific competitive methods must be utilized. A competitor cDNA, having the same sequence of the target TNF-alpha cDNA, a part from an internal 49-bp deletion, was generated and used to set-up a quantitative polymerase chain reaction (PCR) to quantify mRNA of TNF-alpha. Competitor and target were co-amplified using the same primers. The rates of generation of competitor and target TNF-alpha conformed closely to the prediction of the mathematical model, and a high level of accuracy and reproducibility was achieved. The method was applied to quantify TNF-alpha mRNA in PBMC of normal subjects and multiple sclerosis (MS) patients both during clinical relapses and remissions. A statistically significant higher level of TNF-alpha mRNA was detected during relapses than during remissions. High levels of TNF-alpha mRNA were found in 44% of relapses and 12% of samples during remissions, suggesting that TNF-alpha mRNA synthesis is abnormal in MS.

Adult↗

PubMed1998

5D4 keratan sulfate epitope identifies a subset of ramified microglia in normal central nervous system parenchyma.

Microglia expressing keratan sulfate (KS) was studied in normal central nervous system (CNS) and in rat neonatal brain cultures. The majority of KS+ cells are ramified microglia located in the brain parenchyma; positive cells were only exceptionally found in extraparenchymal structures. KS+ cells are ubiquitous, but their density is heterogeneous throughout the CNS. Serial sections incubated with anti-KS MAb and MAb against the complement receptor type 3 (CR3) revealed a higher number of CR3+ cells and double immunofluorescence showed the presence of two microglial populations: the first expressing both KS and CR3, the second expressing only CR3. Two sets of microglial cells were found also in neonatal rat microglial cultures where only a low percentage of microglial cells expressing CR3 was also KS+. KS was not induced by microglia activation.

Animals↗

PubMed1996

Compound heterozygosity for a structural apolipoprotein A-I variant, apo A-I(L141R)Pisa, and an apolipoprotein A-I null allele in patients with absence of HDL cholesterol, corneal opacifications, and coronary heart disease.

BACKGROUND: The concentration of HDL cholesterol is inversely correlated with the risk of coronary heart disease (CHD). Some rare mutations in the apolipoprotein (apo) A-I gene are associated with low levels of HDL cholesterol. Their association with cardiovascular risk is controversial. METHODS AND RESULTS: We studied the molecular defects underlying corneal opacities and absence of HDL cholesterol in three brothers and a sister. In a family study, the importance of these defects for lipid metabolism and manifestation of coronary heart disease was investigated. The frequency of these apo A-I defects was assessed by genotype and phenotype analysis of 477 DNA- and plasma samples, respectively, from the population. The four patients were compound heterozygotes for a null allele and a missense mutation in the apo A-I gene that leads to a leucine-->arginine substitution at residue 141 [apo A-I(L141R)Pisa]. Heterozygotes for either the null allele or the structural variant had half-normal concentrations of HDL cholesterol and apo A-I compared with unaffected family members. Apo A-I(L141R)Pisa was detected in one more unrelated subject. Coronary angiography of the four compound heterozygotes revealed the presence of CHD in all male patients, whose ages ranged between 45 and 52 years. They presented with additional risk factors, including elevated LDL cholesterol levels, obesity, and arterial hypertension. Despite complete HDL deficiency and hypercholesterolemia, CHD was absent in the 51-year-old premenopausal sister. CONCLUSIONS: Apo A-I deficiency may lead to premature atherosclerosis if present in conjunction with additional cardiovascular risk factors.

Adolescent↗

PubMed1996

Immunohistochemical mapping of perineuronal nets containing chondroitin unsulfated proteoglycan in the rat central nervous system.

Subsets of neurons ensheathed by perineuronal nets containing chondroitin unsulfated proteoglycan have been immunohistochemically mapped throughout the rat central nervous system from the olfactory bulb to the spinal cord. A variable proportion of neurons were outlined by immunoreactivity for the monoclonal antibody (Mab 1B5), but only after chondroitinase ABC digestion. In forebrain cortical structures the only immunoreactive nets were around interneurons; in contrast, throughout the brainstem and spinal cord a large proportion of projection neurons were surrounded by intense immunoreactivity. Immunoreactivity was ordinarily found in the neuropil between neurons surrounded by an immunopositive net. By contrast, within the pyriform cortex the neuropil of the plexiform layer was intensely immunoreactive even though no perineuronal net could be found. The presence of perineuronal nets could not be correlated with any single class of neurons; however a few functionally related groups (e.g., motor and motor-related structures: motor neurons both in the spinal cord and in the efferent somatic nuclei of the brainstem, deep cerebellar nuclei, vestibular nuclei; red nucleus, reticular formation; central auditory pathway: ventral cochlear nucleus, trapezoid body, superior olive, nucleus of the lateral lemniscus, inferior colliculus, medial geniculate body) were the main components of the neuronal subpopulation displaying chondroitin unsulfated proteoglycans in the surrounding extracellular matrix. The immunodecorated neurons found in the present study and those shown by different monoclonal antibodies or by lectin cytochemistry, revealed consistent overlapping of their distribution patterns.

Animals↗

PubMed1995

Keratan sulphate is a marker of differentiation of ramified microglia.

Recently we reported that the keratan sulphate epitope recognised by the monoclonal antibody 5D4 is expressed by a population of ramified microglia in adult rats. As ramified microglia is believed to differentiate from ameboid microglia during postnatal development, we studied the rat brain from birth to 90 postnatal days of life with the monoclonal antibody 5D4. Contrary to all the other microglia markers until now described, keratan sulphate is not expressed by ameboid microglia and by macrophages but appears on the surface of microglia only when the cells are differentiated and show ramified processes. The keratan sulphate positive cells become evident at different times in different central nervous system areas; the first were localised in the pyriform cortex and brainstem from the end of the second postnatal week. These observations suggest that keratan sulphate expression on microglia cells is induced by differentiation and by a resting functional state. Moreover the 5D4 monoclonal antibody showed a strong diffuse positive staining of some cortical, thalamic and white matter areas during the first two postnatal weeks. This staining was transient and it does not seem biologically correlated with the expression of the keratan sulphate on differentiated microglia.

Animals↗

PubMed1995

Disappearance of the Vicia villosa-positivity from the perineuronal net containing chondroitin proteoglycan after chondroitinase digestion.

Anti-chondroitin unsulfated proteoglycan (COS-PG) 1B5 monoclonal antibody (MAb) and Vicia villosa agglutinin (VVA) recognize the same neuronal subset. Moreover, when in double immunofluorescence study the sections were digested with chondroitinase ABC (ChABC), a procedure necessary to create the epitope of 1B5 MAb, and incubated with VVA, no VVA positive neurons were detected. This finding suggests that VVA binds terminal N-acetyl-galactosamine present in the glycidic chains of COS-PG or of glycoproteins that form perineuronal molecular aggregates with COS-PG.

Animals↗

PubMed1993

Congenital muscular dystrophy associated with familial junctional epidermolysis bullosa letalis.

A 20-year-old patient was born with epidermolysis bullosa and a severe, slowly progressive muscle disease. Skin biopsy demonstrated junctional epidermolysis bullosa. Muscle biopsy demonstrated degenerative changes with increase in connective tissue, fibre size variability, rods and cytoplasmic bodies, central nuclei. In muscle biopsy dystrophin, chondroitin unsulphate, chondroitin 4-sulphate, chondroitin 6-sulphate, heparan sulphate, collagen III, collagen IV and VI, laminin, and fibronectin were normally distributed. This is the first report of the association of a form of congenital muscular dystrophy with junctional epidermolysis bullosa and, together with the previous reports of muscle involvement in epidermolysis bullosa simplex and dystrophica, it suggests the existence of a syndrome characterized by the contemporaneous presence of skin and muscle involvement.

Adult↗

PubMed1993

Monoclonal antibodies to keratan sulfate immunolocalize ramified microglia in paraffin and cryostat sections of rat brain.

We used six monoclonal antibodies (MAb) recognizing epitopes within keratan sulfate (KS) chains for an immunocytochemical study of adult rat brain. One of the MAb selectively stained microglia and their ramified processes. KS-positive cells were found throughout the CNS in both paraffin-embedded and cryostat sections; the greatest number were present in hippocampus and brainstem. In the cortex the positive processes of some cells surrounded neuronal somata. In the white matter the processes were both parallel and perpendicular to the axon bundles. Double staining showed that KS-positive cells did not express astrocytic or oligodendroglial markers. By immunoelectron microscopy, the positivity was localized around the perikarya and cell processes of small cells with peripheral chromatin clumps and dark cytoplasm, which often contained secondary lysosomes. The KS-positive cells did not contribute to myelin sheaths and were not surrounded by a basal membrane. In addition to the cellular staining, three other MAb stained the white matter diffusely. Anti-KS MAb are therefore proposed as immunohistochemical markers for ramified microglia in both paraffin and cryostat sections of adult rat brain.

Animals↗

PubMed1993

Selective expression of the Met/HGF receptor in human central nervous system microglia.

The c-MET proto-oncogene encodes the tyrosine kinase receptor for hepatocyte growth factor (HGF), also known as scatter factor, a powerful mitogen and motility factor for epithelial cells. We now show that the two previously described forms of the Met/HGF receptor, the intact p190MET and the truncated p140MET, are expressed in physiological conditions in the human central nervous system (CNS). The receptors were identified by Western blot analysis with monoclonal antibodies directed against different epitopes. By immunohistochemical staining the Met/HGF receptor was found to be expressed in a homogeneous cell population, equally distributed between the grey and the white matter, showing morphological features and immunochemical markers specific for the resident microglial cells. These data suggest a possible role for the c-MET proto-oncogene and HGF in microglial reactions to CNS injuries.

Antibodies, Monoclonal↗

PubMed1991

Chondroitin sulfate proteoglycan surrounds a subset of human and rat CNS neurons.

Chondroitin sulfate proteoglycan (CS-PG) bearing glycosaminoglycan (GAG) chains containing unsulfate (COS) and 6-sulfate (C6S) disaccharides was immunolocalized in rat and human CNS by using monoclonal antibodies (MAb) specific for the two disaccharides. The immunostaining with both MAb was restricted to the periphery of a neuronal subset in rat and human CNS. Double immunofluorescence showed codistribution of the antigens around the same neuronal population. The staining with anti-COS MAb was stronger than with anti-C6S MAb, suggesting that the proteoglycan (PG) contains mainly COS disaccharides. In different rat cortical areas, 40-60/mm2 positive interneurons were found, the visual cortex showing the highest value. In human cortex, positivity was also observed around the soma of some pyramidal cells. In the rat, positive neurons were also localized in deep cerebellar nuclei, reticular nucleus of the thalamus, and other structures of the midbrain and hindbrain. CA3 region of hippocampus and the external layer of pyriform cortex were characterized by positivity of the neuropil. Immunoelectronmicroscopy showed the antigens in the extracellular space around the neuronal soma, the synaptic elements and the cell processes of the neuropil. The neuronal surface of the soma and of the proximal dendrites were positive, but the pre- and postsynaptic membranes and clefts were negative.

Aged↗

PubMed1991

Differential distribution and modulation of expression of alpha 1/beta 1 integrin on human endothelial cells.

In this paper we report that the integrin complex alpha 1/beta 1, a laminin/collagen receptor, is expressed on cultured foreskin microvascular endothelium, but is absent on endothelial cells from large vessels such as the aorta and umbilical and femoral veins. The restricted expression of integrin alpha 1/beta 1 to microvascular endothelium was also demonstrated in vivo, by immunohistochemical staining of human tissue sections. Alpha 1 specific antibodies reacted strongly with endothelial cells of small blood vessels and capillaries in several tissues, but not with endothelium of vein and arteries of umbilical cord. Expression of integrin alpha 1 can be induced in cultured umbilical vein endothelial cells by treatment with 5 ng/ml tumor necrosis factor alpha (TNF alpha). Induction of alpha 1 subunit expression also occurred after treatment of umbilical vein endothelium with 10(-5) M retinoic acid or with 10 nM PMA; Maximal induction of alpha 1 integrin was reached after 48 h of treatment and costimulation with TNF alpha and PMA resulted in a synergistic effect. The induction of alpha 1 integrin changed the adhesive properties of umbilical vein endothelial cells, by increasing the adhesiveness to collagen, laminin, and laminin fragment P1, while adhesion to fibronectin and laminin fragment E8 remained constant. The alpha 1 integrin is thus a marker of a specific population of endothelial cells and its expression confers distinctive properties of interaction with the underlying basal membrane.

Amino Acid Sequence↗

PubMed1990

Extracellular matrix of cultured glial cells: selective expression of chondroitin 4-sulfate by type-2 astrocytes and their progenitors.

We have studied the extracellular matrix composition of cultured glial cells by immunocytochemistry with different monoclonal and polyclonal antibodies. Double immunofluorescence experiments and metabolic labeling with [3H]glucosamine performed in different types of cerebellar and cortical cultures showed that bipotential progenitors for type-2 astrocytes and for oligodendrocytes (recognized by the monoclonal antibody LB1 at early stages of their development) synthesize chondroitin sulfate (CS) and deposit this proteoglycan in their extracellular matrix. The distribution of the various [3H]glucosamine-labeled glycosaminoglycans between the intracellular and the extracellular space was different. CS was present both within the cells and in the culture medium, although in different amounts. Bi-potential progenitors became also O4-positive during their development in vitro. At the stage of O4-positivity they were still stained with antibodies against CS. However, when the progenitor cells were maintained in serum-free medium and differentiated into Gal-C-positive oligodendrocytes, they became CS-negative. In the presence of fetal calf serum in the culture medium, the bipotential progenitors differentiated into GFAP-positive type-2 astrocytes. These cells still expressed CS: their Golgi area and their surface were stained with anti-CS antibodies. Staining with monoclonal antibodies specific for different types of CS (4-sulfate, 6-sulfate, and unsulfated) revealed that both bipotential progenitors and type-2 astrocytes synthesized only chondroitin 4-sulfate. Type-1 astrocytes were negative for both the polyclonal and the monoclonal anti-CS antibodies. Finally, type-2 astrocytes and their progenitors were weakly stained with anti-laminin antibodies and unstained with anti-fibronectin. Type-1 astrocytes were positive for both anti-laminin and anti-fibronectin antibodies and appeared to secrete fibronectin in the extracellular space.

Animals↗

PubMed1990

Chondroitin 4-sulfate proteoglycan forms an extracellular network in human and rat central nervous system.

Chondroitin 4-sulfate proteoglycan (C4S-PG) was localized both in rat and human central nervous system (CNS) by monoclonal and polyclonal antisera recognizing the 4-sulfate disaccharide (C4S). In the rat the whole CNS was studied in serial coronal sections. A positive extracellular meshwork was observed both in white and grey matters. In the white matter (WM) C4S-PG formed a network around myelinated axons, sparing myelin sheaths and axoplasms. The neuropil of the grey matter (GM) showed a positive meshwork constituted by delicate intermingling filaments. The cytoplasms of neuronal, glial and endothelial cells were negative. Stronger straining than in the neuropil was observed around the soma and the proximal part of the cell processes of some neurons located in the cortex, in the deep cerebellar nuclei and in some other CNS nuclei. A similar pattern was also observed in human CNS, the only difference being a smaller amount of cortical neurons surrounded by a rim of C4S-PG. This study shows that a PG bearing C4S disaccharide is located extracellularly in the rodent and human CNS and that C4S disaccharides can be present in different types of CNS proteoglycans (PGs).

Aged↗

PubMed1989

Metabolic effects of three new low-dose pills: a six-month experience.

We evaluated the effects on glucose and lipid metabolism in 57 healthy volunteers randomly assigned to one of three low-dose oral contraceptives: two monophasic (desogestrel + ethinylestradiol, EE, and cyproterone acetate + EE) and one triphasic (gestodene + EE) contraceptives. Glucose and insulin responses during OGTT were slightly affected by the cyproterone pill. The insulin area/glucose area ratio and HbA1c level were unchanged in all women. No preparation affected total and LDL-cholesterol levels. Triglycerides rose in all groups, while HDL-CH did only in women taking the two monophasic pills. The three low-dose pills assessed in this study have negligible effects on glucose and lipid metabolism.

Adult↗

PubMed1989

[Effects of low-dose oral contraceptives on sugar metabolism].

The effects on carbohydrate metabolism by four low-dose oral contraceptives were evaluated in four low-dose oral contraceptives were evaluated-66 young women randomly divided in four groups. In the various preparations there were a different dosage of estrogen (ethinylestradiol) together different doses and types of progestogen (desogestrel, gestodene, cyproterone acetate). After six months of treatment, in all groups a slight increase of glycemic and insulinemic responses during OGTT was observed; the significance was achieved with the preparation containing cyproterone acetate alone. Glycated hemoglobin did not change. Our results suggest that these new low-dose oral contraceptives induced negligible metabolic side effects.

Adolescent↗

PubMed1989

Biochemical, histochemical and immunohistochemical study of glycosaminoglycans in human meningiomas.

The localization and quantitation of glycosaminoglycans classes (GAGs) were studied in human meningiomas. Meningiomas presented high amounts of these compounds and electrophoretic separation revealed that they were 90% sulphated. The Alcian method and a polyclonal antiserum against chondroitin sulphate were used to localize the different GAGs in tissue sections. Quantitative and qualitative differences and different tissue distributions of GAGs were observed among transitional, syncytial and fibroblastic meningiomas. Syncytial meningiomas presented the lowest amount of GAGs and the immuno- and histochemical studies showed that they were located only in vessels and connectival trabeculae. Transitional meningiomas contained the highest concentration of GAGs; the percentage of the different GAG classes was similar to that observed in the syncytial oncotype indicating a quantitative but not qualitative difference between the two oncotypes. The high amount of GAGs in transitional meningiomas was attribute to the whorls, the structures stained by the histochemical and immunohistochemical techniques. The tumoral parenchyma of these two oncotypes was negative. On the contrary, fibroblastic meningiomas showed a fine meshwork among tumoral cells containing chondroitin sulphate and heparan sulphate. Biochemical data were consistent with the histochemical and immunohistochemical findings revealing a high percentage of chondroitin sulphate and heparan sulphate in fibroblastic meningiomas. This study suggests that the three meningioma types have different abilities to produce extracellular matrix components.

Glycosaminoglycans↗

PubMed1988

Urinary albumin excretion in normal subjects and in diabetic patients measured by a radioimmunoassay: methodological and clinical aspects.

We have developed a radioimmunoassay method (RIA) to measure urinary albumin excretion. We determined the albumin excretion rate (AER) (micrograms/min) of 122 healthy subjects and 145 diabetic patients (115 type I, 30 type II). The results indicate that the RIA is sensitive (0.39 +/- 0.08 mg/L), precise (CV 5-8%), and gives reliable results on previously frozen urine samples. The distribution of the AER values in healthy subjects and diabetic patients was not normal. It was normalized by log or square-root transformation of the data. Seventy-three percent of diabetic patients lay within the normal range (0.6-10.6 micrograms/min). Twenty percent could be considered "at risk" to develop overt diabetic nephropathy because their albuminuria exceeded a threshold level of 15 micrograms/min chosen previously as the cutoff value for microalbuminuria. We found no correlation between AER and glycated hemoglobin, and only a weak correlation between AER and diabetes duration in type I diabetic patients.

Adolescent↗

PubMed1988

Glutamate dehydrogenase (GDH) deficiency in different types of progressive hereditary cerebellar ataxia.

Leukocyte glutamate dehydrogenase (GDH) was studied in 29 patients affected by progressive cerebellar ataxia (PCA) and in 20 healthy controls. Eight GDH-deficient patients, with GDH activity 2 SD below mean value of controls, were identified. GDH deficiency did not identify a subgroup of PCA by characteristic pattern of inheritance and/or age of onset of disease. However, the GDH-deficient patients presented more neurological signs than non-GDH-deficient patients. A significant correlation was observed between GDH deficiency and the presence of extrapyramidal signs, supranuclear palsy, absence of osteotendineal reflexes and neurogenic electromyographical findings.

Adolescent↗