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Biomedical subjects

D Bosco

Publications and source records attributed to D Bosco.

At least 19 recordsLinked to original sources
PubMed1999

Loss of long-duration response to levodopa over time in PD: implications for wearing-off.

OBJECTIVE: To determine the modifications of the long-duration response to levodopa in PD over a 1-year period. BACKGROUND: The development of predictable motor fluctuations in PD has been attributed mainly to modifications over time of the short-duration response to levodopa, whereas the role of the long-duration response has not been widely investigated. METHODS: In 17 patients with PD the authors examined prospectively both the short-duration response and the long-duration response to levodopa under standardized conditions on two different occasions separated by a period of approximately 1 year (11.7 +/- 3.6 months). RESULTS: At the end of the follow-up period, the short-duration response increased in magnitude but did not change significantly in duration. A total of 24% of patients lost the long-duration response 1 year after their first examination, but a sustained long-duration response could be reestablished by shortening the interdose interval for levodopa intake. Moreover, the duration of the long-duration response after discontinuation of treatment became significantly shorter during 1 year. CONCLUSION: Modifications of the long-duration response may have a pivotal role in generating a fluctuating response, and suggest that therapeutic strategies based on maintenance of the long-duration response should be sought to avoid the appearance of motor fluctuations.

Aged↗
PubMed1999

Loss of alpha 1 connexin does not alter the prenatal differentiation of pancreatic beta cells and leads to the identification of another islet cell connexin.

Connexin alpha 1, also referred to as Cx43, has thus far been the only gap junction protein identified between the hormone-producing cells of pancreatic islets. To investigate whether loss of this connexin affects the development of endocrine pancreas and the differentiation of insulin-producing beta cells, we have taken advantage of a transgenic line in which the gene coding for connexin alpha 1 had been functionally deleted by homologous recombination. Analysis of pancreas at embryonal day 19.5 (E 19.5) after immunostaining for the four main types of islet hormones, showed that islet cell development was similar in homozygous transgenic mice that completely lacked alpha 1 connexin, in mice that were heterozygous for the transgene, and in age-matched controls with a genetic background similar to that of the transgenic animals. In particular, the three animal groups featured beta cells that had a similar insulin content and ultrastructural organization, including the presence of typical gap junction plaques on the membrane. However, quantitative analysis of freeze-fractured membranes showed that these plaques were less frequent in the transgenic mice lacking alpha 1 connexin. This finding prompted us to revisit the connexin pattern of normal pancreatic beta cells. Using RT-PCR amplification and primers specific for nine of the mammalian connexins, we have found that normal rat and mouse pancreas contain six connexin transcripts, including one that codes for alpha 6 connexin, a protein also referred to as Cx45. This transcript was also identified in isolated pancreatic islets, in FACS-purified suspensions of primary beta cells and in the insulin-producing cells of an experimental tumor. Using antibodies, we found that connexin alpha 6 is expressed by the latter cells, as well as by pancreatic fibroblasts and epithelial duct cells. The data show that pancreatic islets have a normal prenatal development in mice that no longer express alpha 1 connexin. They further provide evidence that normal and tumoral insulin-producing cells natively coexpress connexins alpha 1 and alpha 6.

Animals↗
PubMed1999

Effects of TNF-alpha/colchicine combined treatment on Burkitt lymphoma cells: molecular and ultrastructural changes.

Tumour necrosis factor alpha (TNF-alpha) kills Daudi cells (Human Burkitt Lymphoma), inducing either necrosis or apoptosis without DNA fragmentation. Therefore, we were interested in studying the molecular and ultrastructural events occurring when the nucleus is more accessible and cells are blocked in mitosis, following colchicine treatment. In fact, as early as after 1 h treatment a typical ladder pattern was shown by means of DNA gel electrophoresis. In parallel the quantitative analysis of the different morphological patterns observed gave evidence of an increased percentage of primary necrosis after 6 h treatment, and a higher incidence of cells in late apoptosis as well as in secondary necrosis after 24 h treatment. Our findings show that Daudi cells respond to the combined treatment with an increased formation of micronuclei and nuclear alterations which follow a number of early mitochondrial changes and result in enhanced cell death. These data imply that TNF-alpha-induced apoptosis of Daudi cells can be triggered by mitochondrial changes and is somehow related to microtubule organization.

Apoptosis↗
PubMed1998

Enhanced secretion of amylase from exocrine pancreas of connexin32-deficient mice.

To determine whether junctional communication between pancreatic acinar cells contributes to their secretory function in vivo, we have compared wild-type mice, which express the gap junctional proteins connexin32 (Cx32) and connexin26, to mice deficient for the Cx32 gene. Pancreatic acinar cells from Cx32 (-/-) mice failed to express Cx32 as evidenced by reverse transcription-PCR and immunolabeling and showed a marked reduction (4.8- and 25-fold, respectively) in the number and size of gap junctions. Dye transfer studies showed that the extent of intercellular communication was inhibited in Cx32 (-/-) acini. However, electrical coupling was detected by dual patch clamp recording in Cx32 (-/-) acinar cell pairs. Although wild-type and Cx32 (-/-) acini were similarly stimulated to release amylase by carbamylcholine, Cx32 (-/-) acini showed a twofold increase of their basal secretion. This effect was caused by an increase in the proportion of secreting acini, as detected with a reverse hemolytic plaque assay. Blood measurements further revealed that Cx32 (-/-) mice had elevated basal levels of circulating amylase. The results, which demonstrate an inverse relationship between the extent of acinar cell coupling and basal amylase secretion in vivo, support the view that the physiological recruitment of secretory acinar cells is regulated by gap junction mediated intercellular communication.

Amylases↗
PubMed1998

Oral focal mucinosis of the tongue.

Oral focal mucinosis (OFM) is an uncommon clinicopathological entity which is considered to be the oral counterpart of cutaneous focal mucinosis and cutaneous myxoid cyst. It is comprised of a clinically elevated mass with a histological feature of localised areas of myxomatous connective tissue. The present study adds a rare case of OFM of the tongue to the literature, and we present a review of the most characteristic oral myxomatous lesions.

Aged↗
PubMed1998

Upregulation of connexin 26 between keratinocytes of psoriatic lesions.

To assess whether the expression of connexins (Cx) by keratinocytes is altered under conditions of abnormal epidermal differentiation, we have compared Cx26, Cx32, Cx37, Cx40, and Cx43 in the epidermis of 11 psoriatic patients who had not been treated for at least 1 mo and of seven healthy individuals. In all samples of fully mature psoriatic plaques, we have observed a massive expression of Cx26, as judged at both the transcript level (northern blot) and the protein level (immunofluorescence). This protein became consistently detected between keratinocytes of the basal and granular layers at the periphery of psoriatic plaques and in all layers of fully developed psoriatic epidermis, except in regions of parakeratosis. None or a minimal amount of Cx26 was observed in both control and nonlesional regions of psoriatic epidermis. Psoriatic plaques also contained Cx43, the prominent gap junction protein in the interfollicular epidermis of normal human skin. The levels of this protein appeared to be slightly higher in psoriatic than in control skin, as judged at both the transcript level (northern blot) and the protein level (immunofluorescence). Three other connexins (Cx32, Cx37, and Cx40), which are not observed in control interfollicular epidermis, were not induced in either nonlesional or lesional regions of psoriatic skin. The data indicate that selective changes in the normal expression of connexins by keratinocytes are associated with the changes in the proliferation and differentiation program that these cells undergo in psoriasis.

Adult↗
PubMed1998

Sorting human beta-cells consequent to targeted expression of green fluorescent protein.

Pancreatic islets of Langerhans are composed of four major endocrine cell types with a smaller number of nonendocrine cells. To study the molecular constituents and function of just one subpopulation of islet cells, it is necessary to sort them from the other cell types. While rat beta-cells can be sorted by autofluorescence-activated flow cytometry, this has not proved possible on a routine and reproducible basis for human beta-cells. In the present study, we have selectively labeled human beta-cells with green fluorescent protein (GFP), allowing for their sorting by flow cytometry. Human islet cells were infected with replication-defective (attenuated) recombinant adenovirus expressing GFP driven by the rat insulin I promoter (Ad-RIP-GFP) for targeted expression in beta-cells, or beta-galactosidase driven by the promiscuous cytomegalovirus (CMV) promoter (Ad-CMV-beta-gal) as control. Whereas the majority of islet cells can be infected by adenovirus, as shown by control infection with Ad-CMV-beta-gal, increased fluorescence after infection with Ad-RIP-GFP was limited to insulin-containing beta-cells. Infection of islet cells with Ad-RIP-GFP resulted reproducibly in the appearance of a population of intensely fluorescent cells, when analyzed by flow cytometry. These cells were sorted using a fluorescence-activated cell sorter (FACS) and shown by immunofluorescence to consist of >95% beta-cells. The targeted expression of GFP thus allows for preparation of human beta-cells purified close to homogeneity. This method should be readily applicable in any laboratory with FACS capability.

Adenoviridae↗
PubMed1998

Ultrastructural changes in rat cornea after prolonged hypobaric hypoxia.

A study was performed to evaluate the effect of prolonged environmental hypobaric hypoxia on the ultrastructure of rat cornea. 60-day-old female Wistar rats were exposed to a simulated altitude of 5,500 m (350 mmHg) and pO2 of 76 mmHg for 30 days. Control rats were exposed to atmospheric sea level pressure (760 mmHg) and pO2 of 150 mmHg, for the same period. Ultrastructural analysis of the corneal epithelium did not reveal any relevant difference between control and treated rats. In contrast, the corneal stroma of rats subjected to hypoxia showed vascularization with advanced vessel differentiation and signs of active proliferation. The endothelium of hypoxic cornea showed swollen mitochondria and large empty cytoplasmic areas. The endothelial intercellular junctions could hardly be identified in the hypoxic condition. Nevertheless, the most evident change in hypoxic cornea was in Descemet's membrane, which was considerably thickened, to approximately twice that of the control specimen. These results suggest that environmental hypobaric hypoxia induces marked alterations in the corneal stroma and endothelium morphology, probably related to reduced oxygen tension in the aqueous humor, consequent to hypoxemia.

Animals↗
PubMed1997

Diabetic microangiopathy: lupus anticoagulant dependent thrombotic tendency in type 1 (insulin-dependent) diabetes mellitus.

Type 1 (insulin-dependent) diabetes mellitus is associated with long-term vascular complications. In addition to metabolic factors, immunological and haemostatic mechanisms may be involved. Lupus anticoagulant (LA), an immunoglobulin which interferes with endothelial cell function, is frequently associated with a high risk of thromboembolic events. LA has been described in several diseases but never in diabetes mellitus. The aim of this study was to evaluate if endothelial dysfunction and unmodulated haemostasis are amplified by the presence of LA in Type 1 diabetic patients. Plasma samples collected from clinically and biochemically well-characterized Type 1 diabetic patients were examined for LA, fibrinogen, prothrombin (PT), PTT, prothrombin degradation products (F1 + 2) and activated protein C (APC). The results revealed significantly decreased APC and increased F1 + 2 plasma concentrations in LA-positive but not in LA-negative patients; 60% of LA-positive and only 18% of LA-negative patients had microangiopathy (not significant). No thrombotic episodes in large vessels were found in LA-positive patients. These findings suggest that LA could be considered an additional factor in the onset and/or progression of diabetic complications, acting as a link between the immunological and haemostatic systems in the pathogenesis of diabetic microangiopathy.

Adult↗
PubMed1997

Tenascin expression in elastotic cuffs of invasive ductal carcinoma of the breast.

We studied immunohistochemically one thousand one hundred and thirty-seven cases of primary invasive breast cancers (NST) and adjacent normal mammary glands for tenascin expression, and compared their elastic content to verify if a relationship exists between tenascin expression and elastosis. Periductal, perivascular and stromal elastosis were graded on a scale from 0 to 3 (absent to massive). All carcinomas showed tenascin expression and elastosis with various histological appearances. In the adjacent breast, teanscon was distributed around the normal ducts or with extasia and uctal hyperplasia without atypia. Digestion of the sections with elastase prior to staining resulted in a loss of the specific staining reactions in all areas where elastosis was present. Tenascin staining was observed in the mesenchyme closely surrounding the neoplastic ducts and the cancer cell nests. Stromal tenascin staining appeared stronger in those carcinomas that exhibited marked desmoplastic reactions. The highly differentiated tumours contained more elastosis in their tumour tissue than the poorly differentiated ones, whereas tenascin expression was stronger in poorly differentiated tumours than well differentiated tumours. A strong staining for tenascin was observed in the elastotic cuff. Tenascin staining did not disappear afterwards with elastase. We did not find a statistically significant correlation between tenascin expression, elastosis and prognostic factors such as size of the tumour, lymph node metastasis, tumour necrosis and age. In our study tenascin proved to be an additional element in elastotic areas even though the significance of an association between elastosis and tenascin is still unknown, as is that of elastosis itself.

Adult↗
PubMed1997

Identification of phytoplasmas in eggs, nymphs and adults of Scaphoideus titanus Ball reared on healthy plants.

A survey for the presence of aster yellows-related phytoplasmas in the different life stages of Scaphoideus titanus was carried out by means of PCR and nested-PCR assays. Using a phytoplasma 16Srl group-specific primer pair followed by RFLP analysis of amplified products, we were able to detect and identify phytoplasmas from eggs, newly hatched nymphs, fourth and fifth instar nymphs and adults reared on ten phytoplasma-free Vicia faba seedlings. Two of these broad beans became infected, whereas PCR failed to detect phytoplasmas in the same ten plants before leafhopper rearing. These results suggest the possibility of transovarial transmission of aster yellows-related phytoplasmas in S. titanus.

Animals↗
PubMed1997

Hyperinsulinemia-induced hypoglycemia is enhanced by overexpression of connexin 43.

To assess whether cell to cell communications via connexins (Cx) participate to insulin secretion in vivo, we studied insulinoma cells (INS1) implanted in rats after stable transfection with connexin 43 (Cx43). We found that compared to wild-type and transfected cells, which in vivo express modest levels of Cx43 and junctional communication, cells overexpressing Cx43 communicated extensively, featured decreased growth, and induced a much higher hyperinsulinemia. As a result, rats with insulinomas made of these cells became more severely hypoglycemic than rats implanted with either wild-type, neomycin-transfected cells or cells transfected with a Cx43 antisense complementary DNA. Rats implanted with transfected cells that expressed modest level of Cx43 showed levels of circulating insulin similar to those in rats implanted with wild-type INS1 cells. The data show that overexpression of Cx43 influences the growth and secretion of the implanted insulinoma cells, providing evidence for a contribution of Cx-mediated cell to cell communication in the functioning of insulin-producing cells in vivo.

Animals↗
PubMed1996

Protein kinase A-dependent phosphorylation of GLUT2 in pancreatic beta cells.

In pancreatic beta cells, cyclic AMP-dependent protein kinase regulates many cellular processes including the potentiation of insulin secretion. The substrates for this kinase, however, have not been biochemically characterized. Here we demonstrate that the glucose transporter GLUT2 is rapidly phosphorylated by protein kinase A following activation of adenylyl cyclase by forskolin or the incretin hormone glucagon-like peptide-1. We show that serines 489 and 501/503 and threonine 510 in the carboxyl-terminal tail of the transporter are the in vitro and in vivo sites of phosphorylation. Stimulation of GLUT2 phosphorylation in beta cells reduces the initial rate of 3-O-methyl glucose uptake by approximately 48% but does not change the Michaelis constant. Similar differences in transport kinetics are observed when comparing the transport activity of GLUT2 mutants stably expressed in insulinoma cell lines and containing glutamates or alanines at the phosphorylation sites. These data indicate that phosphorylation of GLUT2 carboxyl-terminal tail modifies the rate of transport. This lends further support for an important role of the transporter cytoplasmic tail in the modulation of catalytic activity. Finally, because activation of protein kinase A stimulates glucose-induced insulin secretion, we discuss the possible involvement of GLUT2 phosphorylation in the amplification of the glucose signaling process.

3-O-Methylglucose↗
PubMed1996

Quantitative determination of tomato yellow leaf curl geminivirus DNA by chemiluminescent assay using digoxigenin-labeled probes.

A quantitative dot-blot hybridisation assay was developed for tomato yellow leaf curl geminivirus (TYLCV). The assay is based on chemiluminescent detection of viral and plasmid DNA using digoxigenin-labeled probes on nylon membranes. The response-error relationship was studied and a square root transformation was found to stabilise the variance of the response. An asymmetric sigmoid (gompertz) curve was used to describe the dose-response relationship. The detection limits and the precision profiles of the curves were studied. A method is suggested for setting an upper limit on the maximum DNA amount that can be discriminated from the upper asymptote. With respect to different times of exposure of an X-ray film to a membrane, the shortest times gave better upper limits and the longest times provided better detection limits. Purified virus and plasmid preparations were studied in various dilution media, such as TYLCV-free Bemisia tabaci (the whitefly vector) and tomato extracts, with particular attention to parallelism with standard calibration curves. Plasmid diluted in buffer was found useful for calibration of purified virus and virus in the vector, while extraction of known amounts of virus, in parallel with samples to be examined, was needed to quantify viral DNA in plant hosts.

DNA Probes↗
PubMed1996

Induced hypotensive anesthesia for adolescent orthognathic surgery patients.

PURPOSE: This study compared blood loss, quality of surgical field, and duration of procedure with and without induced hypotensive anesthesia in adolescent orthognathic surgery patients. PATIENTS AND METHODS: Fifty orthognathic surgery patients were studied in a prospective, randomized, blocked, stratified, and single-blind fashion. All patients underwent either sagittal ramus split osteotomy or Le Fort I osteotomy or genioplasty. One group of patients (n = 25) had induced hypotension; the other group (n = 25) received anesthesia with no attempt to deliberately reduce blood pressure during the operation. The surgeon, unaware of to which group the patient had been assigned, rated the surgical field every 15 minutes. At completion of surgery, three different methods were used to estimate or calculate blood loss. Duration of the procedure was recorded from time of first incision to time of last suture placement. The data were analyzed using ANOVA, chi-squared, and linear regression, where appropriate. RESULTS: Estimated blood loss was significantly less when induced hypotensive anesthesia was used. The surgical field was better, but there was no significant difference in duration of the procedure with induced hypotensive anesthesia. CONCLUSION: Induced hypotensive anesthesia results in both reduced blood loss and improvement in surgical field.

Adolescent↗
PubMed1996

Histiocytic necrotizing lymphadenitis (Kikuchi's disease).

We present a case report of a young asiatic women presenting Kikuchi's lymphadenitis, a suprahyoidea mass, pharyngitis, fever, weakness and lack of appetite. The clinical picture quickly improved after biopsy until complete remission within two months. Sierologic studies showed significant antibody titres revealing a previous EBV infection; they were also negative for Toxotest, HbsAg, HIV and serogroup 3 and 9 Yersinia Enterocolitica. Histological findings showed necrotic changes and large foci of infiltration in the cortex and/or paracortex. They consisted of variable numbers of small lymphocytes, immunoblasts, macrophages and so-called plasmacytoid monocytes, neutrophils were absent. Immunohistochemistry revealed within the lesion T-Lymphocytes displaying a phenotype CD4+ while T-Lymphocytes CD8+ were observed in the paracortex and in the peripheral region of the lesion.

Adult↗