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ABSTRACT: Nonsyndromic cleft lip with or without cleft palate (CL/P) is the most common orofacial malformation, having a non-Mendelian and multifactorial aetiology. It has been shown that polymorphic variants of genes encoding key proteins of folate and methionine metabolism might be important maternal risk factors for having a child with these craniofacial anomalies. The aim of this study was to evaluate the role of two polymorphisms of the methylenetetrahydrofolate dehydrogenase 1 (MTHFD1) gene, the A1958G and the G401A variants, on the risk of CL/P in the Italian population. A1958G and G401A polymorphism genotyping of MTHFD1 was performed on 216 CL/P triads, (patient and parents), for this study by restriction endonuclease digestion of PCR products. Linkage disequilibrium between markers and disease was tested using both pairwise and haplotype analyses. In our case-parents triad design no significant association between MTHFD1 and the disease is evident. Our data do not support MTHFD1 involvement in CL/P onset among the Italian population.
Annals of Human Genetics 06/2008; 72(Pt 3):297-9. · 2.57 Impact Factor
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K M Lammers,
S Ouburg,
S A Morré,
J B A Crusius,
P Gionchett,
F Rizzello,
C Morselli, E Caramelli,
R Conte,
G Poggioli,
M Campieri,
A S Peña
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ABSTRACT: To investigate the single nucleotide polymorphisms (SNPs) in genes involved in bacterial recognition and the susceptibility to pouchitis or pouchitis severity.
Analyses of CD14 -260C>T, CARD15/NOD2 3020insC, Toll-like receptor (TLR)4 +896A>G, TLR9 -1237T>C, TLR9+2848G>A, and IRAKM + 22148G>A SNPs were performed in 157 ileal-pouch anal anastomosis (IPAA) patients (79 patients who did not develop pouchitis, 43 infrequent pouchitis patients, 35 chronic relapsing pouchitis patients) and 224 Italian Caucasian healthy controls.
No significant differences were found in SNP frequencies between controls and IPAA patients. However, a significant difference in carriership frequency of the TLR9-1237C allele was found between the infrequent pouchitis and chronic relapsing pouchitis groups [P = 0.028, oddos ratio (OR) = 3.2, 95%CI = 1.2-8.6]. This allele uniquely represented a 4-locus TLR9 haplotype comprising both studied TLR9 SNPs in Caucasians. Carrier trait analysis revealed an enhanced combined carriership of the alleles TLR9 -1237C and CD14 -260T in the chronic relapsing pouchitis and infrequent pouchitis group (P = 0.018, OR = 4.1, 95%CI = 1.4 -12.3).
There is no evidence that the SNPs predispose to the need for IPAA surgery. The significant increase of the combined carriership of the CD14 -260T and TLR9 -1237C alleles in the chronic relapsing pouchitis group suggests that these markers identify a subgroup of IPAA patients with a risk of developing chronic or refractory pouchitis.
World Journal of Gastroenterology 12/2005; 11(46):7323-9. · 2.47 Impact Factor
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ABSTRACT: It has been demonstrated that cyclosporin A (CyA) blocks the immune system, acts on cytoskeleton and stimulates the production of extracellular matrix (ECM) and transforming growth factor-beta1 (TGF-beta1). This cytokine, such as transforming growth factor-alpha (TGF-alpha), induces deposition of glycosaminoglycans (GAG), proteoglycans and collagen fibres in the ECM.
In this work, we examined the effect induced by CyA, TGF-beta1 and TGF-alpha on cultures of healthy and overgrown human gingival fibroblasts in order to evaluate the glycosaminoglycan, cytoskeletal changes and the behaviour of fibroblasts after concanavalin A (Con A) treatment. Moreover, we examined gingival biopsies by Alcian blue histochemical staining and electron transmission microscopy.
Total and extracellular sulphated GAG in overgrown gingiva specimens and in derived fibroblast cultures treated with CyA and cytokines were significantly higher than controls. The action of cytokines was increased (P < or = 0.01) compared with CyA with a greater effect of TGF-alpha in comparison with TGF-beta1; the electron microscopy showed ECM accumulation. The agglutinations showed the heterogeneity of fibroblast populations.
Stimulation with Con A showed that the fibroblast population had cell surface heterogeneity, and could respond in a different way to both CyA and cytokine stimulus. Moreover, increased synthesis of GAG in overgrown gingiva compared with synthesis in normal fibroblasts before CyA treatment suggests a possible genetic origin of damage. As not all CyA-treated patients develop gingival overgrowth, a genetic predisposition may explain the different responses of gingival fibroblast populations.
Journal of Oral Pathology and Medicine 07/2004; 33(6):346-53. · 1.63 Impact Factor
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F Pezzetti,
M Martinelli,
L Scapoli,
F Carinci,
A Palmieri,
J Marchesini,
P Carinci, E Caramelli,
R Rullo,
F Gombos,
M Tognon
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ABSTRACT: The pathogenesis of cleft lip with or without cleft palate (CL/P) is complex; its onset could be due to the interaction of various genetic and environmental factors. Recently MTHFR functional polymorphisms were found to increase the risk of this common malformation; however, this finding is still debated. We investigated 110 sporadic CL/P patients, their parents and 289 unrelated controls for c.665C>T (commonly known as 677C>T; p.Ala222Val) and c.1286A>C (known as 1298A>C; p.Glu429Ala) polymorphism in the MTHFR gene. Transmission disequilibrium test (TDT) showed no distortion in allele transmission. Nevertheless, association studies revealed significant differences in allele frequencies between mothers of CL/P patients and controls. This work supports the hypothesis that a lower MTHFR enzyme activity in pregnant women, mostly related to the c.665C>T variant form, is responsible for a higher risk of having CL/P affected offspring.
Human Mutation 07/2004; 24(1):104-5. · 5.69 Impact Factor
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ABSTRACT: During organ differentiation, cell-extracellular matrix (ECM) interactions are required. The components of the ECM, such as glycosaminoglycans, fibronectin, laminin, and collagens, change in relation to cytokine and enzyme activity. Moreover, glycosaminoglycans (GAGs) are components of the ECM that play an important role in both cytokine regulation and cell activities. In this work we studied the accumulation of hyaluronic acid and chondroitin sulfate and heparan sulfate proteoglycans (PGs), beta-N-acetyl-D-glucosaminidase activity, the presence of transforming growth factor beta(2) (TGF beta(2)), and interleukin-1 (IL-1), and the localization of fibronectin, laminin, and collagen I and IV during the early stages of chick embryo lung development. We also determined the levels of hyaluronic acid, chondroitin sulfate, dermatan sulfate, and heparan sulfate GAGs and the activity of beta-N-acetyl-D-glucosaminidase with biochemical methods. Our data show that beta-N-acetyl-D-glucosaminidase activity increases in each cell, especially in the epithelial growth front at the emergence of each bronchial bud, where hyaluronic acid and IL-1 are located in the surrounding mesenchymal areas. Chondroitin sulfate and heparan sulfate PGs, fibronectin, laminin, and collagen I and IV are evident in the area near the basal membrane along the sides where the forming structures are stabilized. Biochemical data show that beta-N-acetyl-D-glucosaminidase activity increases in cells during lung development and is related to GAG decrease and to modifications of the nonsulfated/sulfated GAG ratio. These modifications could change cytokine activity and play an important role in bronchial branching development.
Journal of Histochemistry and Cytochemistry 04/2004; 52(3):325-34. · 2.72 Impact Factor
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ABSTRACT: Promising results from clinical studies on the effect of probiotics as maintenance therapy in inflammatory bowel disease and in the prevention of onset of pouchitis ask for studies to unravel the still poorly understood mechanism of action of probiotics.
To evaluate whether the probiotic bacteria that were used in the clinical studies (VSL#3, Escherichia coli Nissle 1917, and Lactobacillus GG) are able to induce chemokine production in epithelial cells, HT29/19A monolayers were incubated with cell debris and cell extract fractions of single strains of the probiotic bacteria in doses ranging from 10(3) to 10(9) colony-forming units/ml for 32 h. Supernatants were measured for interleukin 8 by ELISA.
Lactobacilli and bifidobacteria strains from VSL#3 and Lactobacillus GG did not induce interleukin 8, whereas both cell debris and cell extracts from E. coli Nissle 1917 induced interleukin 8 production in a dose-dependent way. Cell extracts from streptococcal strains induced interleukin 8 when applied at high concentrations.
Probiotic Gram-positive bacteria did not induce interleukin 8, whereas the nonpathogenic, Gram-negative E. coli Nissle 1917 strain induced interleukin 8 in a dose-dependent way in this culture model. These results suggest that probiotic Gram-positive bacteria and E. coli Nissle 1917 may exert their beneficial effects on the host by a different mechanism of action.
The American Journal of Gastroenterology 06/2002; 97(5):1182-6. · 7.28 Impact Factor
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M Marchisio,
A Di Baldassarre,
D Angelucci, E Caramelli,
A Cataldi,
S Castorina,
A Antonucci,
L Di Giovannantonio,
C Schiavone,
R Di Biagio,
M Falconi,
G Zauli,
S Miscia
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ABSTRACT: The expression, cellular distribution, and activity of PIP(2)-specific phospholipase C (PLC) in healthy human gastric-mucosa cells have been recently studied in our laboratories and a direct evidence for an almost exclusive expression of PLC beta isoforms, with the exception of PLC beta4, has been provided. These results addressed our attention to possible modification of PLC expression and activity during neoplastic transformation of the human gastric mucosa. In the present article we present results indicating that PLC delta2 is markedly expressed in type II intestinal metaplasia and in the adenocarcinoma whereas traces of other PLC isoforms were sometime detected. Interestingly, we found that type I intestinal metaplasia was in the majority of the cases PLC delta2-negative, but when expressed, this type of metaplasia generally considered as benignant, always evolved toward neoplastic transformation. These results therefore readdress the question of surveillance of the patients with type I intestinal metaplasia and suggest that PLC delta2 expression might be a possible marker of gastric malignant transformation.
American Journal Of Pathology 10/2001; 159(3):803-8. · 4.89 Impact Factor
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ABSTRACT: This paper evaluated how the pregnancy after 41 completed weeks' gestation with amniotic fluid index (AFI) > 6 has a slower response to the prostaglandin E2 (PGE2) induction. Eighty-one post-term pregnancies (41 completed gestations' weeks) with unfavorable cervix were considered in this follow-up. Induction was performed by means of intracervical PGE2 gel (Dinoprostone 0.5 mg). After 12 hours, if the cervix was still unfavorable, then another gel administration followed. Cases that had oxytocin administration were excluded from the study. The median time of spontaneous delivery in the overall series was 25 hours, 14 minutes. We had 18 cases of cesarean section (22.2%). In the group of pregnancies with AFI > 6 (60 cases) and in the group with AFI < or =6 (21 cases), the median time of spontaneous delivery was 29 hours, 25 minutes and 23 hours, 39 minutes, respectively (p-value = 0.02). The rate of cesarean sections was 26.67 and 9.52, respectively in the two groups (p-value >0.05). Two out of four cases of cesarean sections for fetal distress belonged to the group of AFI > 6. All the 14 cases of cesarean section for dystocia belonged to the group with AFI > 6. Considering just patients who did not deliver within 12 hours (57 cases), median time of spontaneous delivery was 33 hours and 24 hours 40 minutes for group AFI > 6 (42 cases) and AFI < or =6 (15 cases), respectively (p-value = 0.0009). Thirty-one cases out of 57 had another PGE2 gel administration. Adjusted odds ratio was 0.33 (0.16-0.65, 95% C.I.) for AFI < or =6 versus AFI > 6.
American Journal of Perinatology 02/2000; 17(6):319-24. · 1.32 Impact Factor
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ABSTRACT: Little is known about the natural history of polycystic ovary syndrome (PCOS), although preliminary data indicate that affected women are more susceptible than the general population to diabetes and cardiovascular diseases at post-menopausal ages. The aim of this study was to follow-up all main features of the metabolic syndrome in a group of young women with PCOS and to investigate the long-term effects on metabolism and body composition of oestrogen-progestagen (OP) compounds, which are frequently used in these women to treat hyperandrogenism and related clinical features.
Long-term follow-up study.
Thirty-seven women with PCOS were re-evaluated 10.3 +/- 0.8 years (range 6-18 years) after their first assessments (age: before 19.8 +/- 4.9 years; after 29.9 +/- 4.4 years). When first examined, women were instructed to follow a hypocaloric diet if they were obese plus OP, if they agreed to such treatment. Main anthropometric parameters, basal sex hormones and lipids, fasting and glucose-stimulated glucose and insulin levels and several clinical data were recorded before and after follow-up.
In the whole group of women with PCOS we found no changes in body weight and fat mass, whereas both the waist-to-hip ratio and the waist-to-thigh ratio were significantly reduced. No significant changes occurred in mean fasting and glucose-stimulated glucose and insulin concentrations, whereas a significant increase in high-density lipoprotein-cholesterol was found. No significant changes occurred in testosterone levels. During the follow-up period 16 women took OP for an average of 97 +/- 18 months (range 12-180 months) (OP-users) whereas 21 women never took OP (non-OP-users). All OP-users were still taking OP when re-evaluated at the follow-up examination. With respect to baseline values, body mass index was higher in non-OP-users than in their counterparts. Waist circumference (P < 0.025), the waist-to-hip (P < 0.05) and the waist-to-thigh (P < 0.01) ratios decreased significantly only in the OP-users. In addition, percentage changes in waist circumference (P < 0.05) and waist-to-hip ratio (P < 0.05) during the follow-up period were significantly different between the groups. Glucose tolerance (as area under the curve (AUC)) improved (P < 0.05) in OP-users but not in non-OP-users. Moreover, compared to baseline values, basal insulin levels were significantly (P < 0.01) reduced in OP-users but not in non-OP-users. On the contrary, no significant change was found in insulinAUC in the former, whereas it significantly increased (P < 0.05) in the latter. Accordingly, fasting C-peptide decreased (P < 0.05) in OP-users, whereas both fasting (P < 0.01) and stimulated (P < 0.01) C-peptide significantly increased in non-OP-users. Changes in fasting or stimulated insulin and C-peptide in non-OP-users were not associated with parallel changes in testosterone levels. Total cholesterol and triglycerides did not change in either group, but HDL-cholesterol increased (P < 0.05) only in OP-users. Sex hormone-binding globulin concentrations increased significantly (P < 0.01) in OP-users, without any significant change in non-OP-users. Testosterone concentrations did not change significantly in either group, but the testosterone: SHBG ratio significantly decreased in OP-users (P < 0.05) but not in the non-OP-users. Among the clinical features, acanthosis nigricans significantly (P < 0.01) worsened in non-OP-users but not in the OP-users, without any significant change in the hirsutism and acne scores. Pregnancy rates during the follow-up were similar in both groups.
These data indicate that hyperinsulinaemia and insulin resistance tended to worsen spontaneously in women with PCOS, without any worsening of the hyperandrogenism. Long-term oestrogen-progestagen treatment countered this tendency, probably because it improved the pattern of body fat distribution, by reducing abdominal fat depots.
Clinical Endocrinology 04/1999; 50(4):517-27. · 3.17 Impact Factor
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ABSTRACT: The granulocytic differentiation of HL-60 cells induced by all-trans retinoic acid was accompanied by a progressive tyrosine phosphorylation of specific proteins in either cells or isolated nuclei. Among these phosphoproteins, we identified the Vav adaptor in whole cells as well as in the inner nuclear compartment, where the increase in its tyrosine phosphorylation level was more conspicuous. We also demonstrated the differentiation-dependent association of nuclear phosphorylated Vav to phospholipase C-gamma1 and to the p85 regulatory subunit of phosphoinositide 3-kinase. The role of the Vav/phospholipase C-gamma1/phosphoinositide 3-kinase phosphoprotein complexes in the nuclei of HL-60 induced to differentiate along the granulocytic lineage is discussed.
FEBS Letters 01/1999; 441(3):480-4. · 3.54 Impact Factor
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ABSTRACT: Signal transduction elements, including protein kinase C, have been identified in mammalian spermatozoa. In order to evaluate the pattern of expression and the subcellular localization of nine different protein kinase C isoforms in the course of spermatogenesis, we utilized quantitative electron microscopy immunocytochemistry on thin sections of rat seminiferous tubules. The results indicate a progressive reduction of the protein kinase C isoforms present in the early stages of spermatogenesis, so that in late spermatids none of them is present in the nucleus, while the isoforms alpha, gamma and beta II are specifically retained in the acrosome, the isoforms beta I and zeta in the neck, and the isoform epsilon in the tail. These isoforms, except for beta II, are maintained at the same sites in spermatozoa. Western blotting analysis indicates the presence of alpha and gamma isoforms in the head subfraction, and of beta I, zeta and epsilon isoforms in the tail subfraction of spermatozoa. These findings suggest that specific protein kinase C isoforms may be functionally involved in some events of spermatozoa differentiation and, eventually, in the fertilization process.
European Journal of Cell Biology 03/1997; 72(2):142-50. · 2.81 Impact Factor
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ABSTRACT: The effect of thrombopoietin (TPO) on the functional activity of surface alpha IIb beta 3 (GPIIbIIIa) was investigated in both primary human megakaryocytic cells, derived from peripheral blood CD34+ cells, and HEL hematopoietic cell line. TPO (100 ng/mL) induced a sixfold to ninefold enhancement of adhesion of both primary megakaryocytic and HEL cells to plates coated with either fibrinogen or fibronectin and a parallel increase of immunoreactivity to the PAC1 monoclonal antibody (MoAb) and fluorescein isothiocyanate-fibrinogen, both of which recognize an activated state of alpha IIb beta 3. The enhanced adhesion to fibrinogen or fibronectin was mediated by the Arg-Gly-Asp (RGD) recognition sequence of alpha IIb beta 3, as it was abolished by pretreatment of cells with saturating concentrations of RGDS peptide. A MoAb specific for the alpha IIb beta subunit of alpha IIb beta 3 also inhibited cell attachment to fibrinogen or fibronectin, while MoAb to anti-alpha v beta 3 or anti-alpha 5 integrins were completely ineffective, clearly indicating that alpha IIb beta 3 participates in this association. A role for PI 3 kinase (PI 3-K) in the TPO-mediated increase in alpha IIb beta 3 function in megakaryocytic cells was suggested by the ability of the PI 3-K inhibitor wortmannin (100 nmol/L) and antisense oligonucleotides directed against the p85 regulatory subunit of PI 3-K to completely block the TPO-induced increase in alpha IIb beta 3 integrin activity upon TPO stimulation. The modulation of adhesiveness to extracellular matrix proteins containing the RGD motif mediated by TPO likely plays a physiologic role in megakaryocytopoiesis, as pretreatment of CD34+ cells with RGDS or anti-alpha IIb MoAb significantly reduced the number of megakaryocytic colonies obtained in a fibrinclot semisolid assay.
Blood 03/1997; 89(3):883-95. · 9.90 Impact Factor
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ABSTRACT: Presence and intracellular distribution of phosphoinositide-specific phospholipase C, phosphatidylinositol 4,5-bisphosphate and protein kinase C have been investigated in rat maturing germ cells and spermatozoa. The isoforms beta 1 and gamma 1 of phosphoinositide-specific phospholipase C were immunologically identified and found to be predominantly nuclear or cytoplasmic and nuclear, respectively. The two enzymes were present in the maturing cell lineage of the seminiferous tubule, except for the nucleus of late spermatids, and absent in spermatozoa, in which, however, a phosphoinositide-specific phospholipase C activity persisted, due to yet uncharacterized enzyme(s). Protein kinase C paralleled these developmental changes, and was completely down-regulated in both total cell homogenates and isolated nuclei obtained from spermatozoa. On the contrary, phosphatidylinositol 4,5-bisphosphate, present at the nuclear level in all cell types, accumulated in the nuclei of late spermatids and spermatozoa. These data support the contention that the spermatozoon nucleus stores a lipid-dependent signaling apparatus which could be reactivated either during sperm maturation or at fertilization.
European Journal of Cell Biology 11/1996; 71(2):154-64. · 2.81 Impact Factor
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ABSTRACT: We investigated whether members of the protein kinase C (PKC) family of enzymes were involved in the nuclear events underlying granulocytic differentiation induced by 10(-6) M all-trans retinoic acid (ATRA) in HL-60 cells. PKC activity was analysed by using a serine substituted specific peptide which enabled the evaluation of the whole catalytic activity of both Ca2+ -dependent and Ca2+ -independent PKC isoforms. In parallel, the subcellular distribution of various PKC isoforms was evaluated by Western blot, immunoprecipitation and in situ immunocytochemistry analyses. The level of PKC catalytic activity in the nuclei of HL-60 cells significantly (P < 0.01) and progressively increased from 1 h of ATRA treatment onwards. Consistently, PKC-alpha and -zeta showed a striking and selective accumulation inside the nucleus upon treatment with ATRA. On the other hand, PKC-beta I and -beta II, the only two other isoforms present at nuclear level, did not show any significant modification upon ATRA treatment. The remaining PKC isoforms were not detectable inside the nucleus and showed only modest and non-significant variations, also in whole cell homogenates, upon ATRA treatment, except PKC-delta which showed a progressive down-regulation. Our data suggest that a selective nuclear translocation of PKC-alpha and -zeta might be involved in the process of granulocytic differentiation induced by ATRA in HL-60 cells.
British Journal of Haematology 06/1996; 93(3):542-50. · 4.94 Impact Factor
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ABSTRACT: We investigated whether members of the protein kinase C (PKC) family of enzymes could play a role in the nuclear events involved in megakaryocytic differentiation. PKC activity was analysed using a serine substituted specific peptide, which enabled us to evaluate the whole catalytic activity in the pluripotent haemopoietic HEL cell line treated with 10(-7)M phorbol myristate acetate (PMA) or haemin. In parallel, the subcellular distribution of different PKC isoforms (alpha, beta I, beta II, gamma, delta, epsilon, theta, eta, zeta) was evaluated by Western blot. PKC catalytic activity in the nuclei of HEL cells showed a peak after acute (30 min) treatment with PMA, followed by a significant (P < 0.05) decline after prolonged exposure (72 h) to the same agonist, when most HEL cells had acquired a differentiated megakaryocytic phenotype. Western blot analysis of nuclear lysates consistently showed a significant increase of PKC-alpha, -beta I, -epsilon, theta and -zeta isoforms after 30 min of PMA treatment, followed by a drastic decline of all but PKC-zeta isoforms. Moreover, PKC-6 delta appeared in HEL nuclei only after 72 h of exposure to PMA. On the other hand, neither the catalytic activity nor the immunoreactivity of the different PKC isoforms showed remarkable variations in nuclei of HEL cells induced to differentiate along the erythroid lineage with 10(-7)M haemin. The possible implications of these findings for a better understanding of the molecular events underlying the process of megakaryocytic differentiation are discussed.
British Journal of Haematology 03/1996; 92(3):530-6. · 4.94 Impact Factor
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ABSTRACT: We investigated the effect of extracellular Tat protein of human immunodeficiency virus-type 1 (HIV-1) on the phosphatidylinositol (PI) cycle, which represents a major signal transduction pathway in lymphoid cells. Recombinant Tat, recombinant HIV-1 p24 and cross-linked anti-CD3 monoclonal antibody (mAb) were added in culture for 1-60 min to Jurkat lymphoblastoid CD4+ T cells. The stimulation of T cell receptor by cross-linked anti-CD3 mAb resulted in a rapid increase of the phosphatidylinositol-specific phospholipase C (PI-PLC) activity in whole cell lysates. On the other hand, Tat protein, either alone or in combination with anti-CD3 mAb, showed little effect on the PI turnover of whole cell extracts. Tat, however, selectively stimulated a nuclear-specific PI-PLC with a peak of activity after 30 min from the addition in culture to Jurkat cells. Interestingly, this time corresponded to that required for the uptake and nuclear localization of recombinant Tat protein, as demonstrated by electron microscope immunocytochemistry experiments with anti-Tat mAb. Moreover, exogenous Tat reached the nucleus of Jurkat cells in a bioactive form, as shown in a HIV-1 long terminal repeat-chloramphenicol acetyl transferase transactivation assay. The specific increase of a nuclear PI-PLC activity was further demonstrated by the ability of Tat to stimulate PI turnover also when added directly to isolated nuclei. As a whole, these data demonstrate that Tat selectively stimulates a nuclear polyphosphoinositide hydrolysis, which appears to be independent of the cellular PI turnover. The relevance of these findings for a better understanding of the biological functions of extracellular Tat is discussed.
European Journal of Immunology 10/1995; 25(9):2695-700. · 5.10 Impact Factor
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ABSTRACT: The organization of DNA-protamine complexes and their association with the nuclear matrix have been analyzed in sperm nuclei by in situ Nick Translation at the electron microscope. The data obtained indicate that the chromatin organization in sperm nuclei is maintained during the sperm condensation by means of interaction with the nuclear matrix at fixed sites. The fine structure of the sperm nucleus and sperm nuclear matrix, investigated by sectioning and replica of freeze-fractured specimens, suggests that the lamellar array observed by freeze-fracturing in the sperm nuclei, could depend on the inner matrix which presents a regular organization of globular structures involved in the maintenance of chromatin domains.
Italian journal of anatomy and embryology = Archivio italiano di anatomia ed embriologia 02/1995; 100 Suppl 1:39-46.
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ABSTRACT: In this study we evaluated the effect of human immunodeficiency virus type 1 (HIV-1) recombinant Tat protein on mRNA expression and protein synthesis of two inflammatory cytokines-interleukin-6 (IL-6) and transforming growth factor-beta 1 (TGF-beta 1)-by peripheral blood (PB) monocytes. Whereas maximal levels of IL-6 protein were recovered in PB monocyte culture supernatants after 24-48 h from the addition of 1 micrograms/ml of recombinant Tat, TGF-beta 1 showed a slower and progressive increase, reaching maximal levels only after 72-96 h of culture. Consistently, the analysis of the steady-state levels of mRNA showed a sharp increase of IL-6 mRNA expression after 24h of culture, with a slow decline thereafter. On the other hand, TGF-beta 1 mRNA expression showed a slow increase only after 72-96 h of culture. Moreover, IL-6 appeared involved in the up-regulation of TGF-beta 1, because the addition of a neutralizing anti-IL-6 antibody to Tat-treated PB monocyte cultures significantly reduced the amounts of TGF-beta 1 recovered in the culture supernatants after 96 h. The present demonstration that HIV-1 Tat protein directly up-regulates IL-6 expression and stimulates TGF-beta 1 production both directly and indirectly, through early IL-6 production, could have important implications in the pathogenesis of HIV-1 disease.
British Journal of Haematology 11/1994; 88(2):261-7. · 4.94 Impact Factor
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ABSTRACT: In the sperm nuclei the DNA is packaged into a highly condensed form and is not organized into nucleosome and solenoid but is bound and stabilized mainly by the protamines that arrange the DNA in an almost crystalline state. As demonstrated for somatic cells, the sperm DNA has been reported to be organized in loop domains attached to the nuclear matrix structures. However, the possible role of the sperm head matrix in maintaining the loop organization in absence of a typical nucleosomal structures has not been fully elucidated. By using in situ nick translation at confocal and electron microscope level, we analyzed the organization of the DNA-protamine complex and its association with the sperm nuclear matrix. The data obtained indicate that the chromatin organization in sperm nuclei is maintained during the sperm condensation by means of interactions with the nuclear matrix at fixed sites. The fine structure of sperm nucleus and of sperm nuclear matrix, investigated on sections and replicas of freeze-fractured specimens, suggests that the lamellar array, observed by freeze-fracturing in the sperm nuclei, could depend on the inner matrix which presents a regular organization of globular structures possibly involved in the maintenance of chromatin domains in highly condensed sperm nuclei also.
Biology of the Cell 02/1994; 81(1):47-57. · 3.60 Impact Factor
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ABSTRACT: The water-soluble probe carboxyfluorescein (CF), contained in the internal aqueous phase of liposomes, was used to investigate the interaction of phospholipid vesicles with isolated nuclei. Ultrastructural analysis indicated that adherent liposomes coated the nuclear surface, and fluorescence microscopy showed that they contained quenching concentrations of the dye. Flow cytometry revealed that the transfer of the entrapped dye from the adhering liposomes to nuclei was blocked by chilling at 0 degrees C. Chase experiments demonstrated that the most reliable mechanism of dye transfer involved fusion phenomena between the liposomal and the nuclear membranes. After the release of the fluorophore into the nucleus, empty liposomes could withdraw the intranuclear soluble fraction of the dye.
Cell Biochemistry and Function 02/1989; 7(1):71-4. · 1.77 Impact Factor
