Biomedical subjects

B Delpech

Publications and source records attributed to B Delpech.

At least 37 records · Page 2Linked to original sources

PubMed1999

Ligation of the CD44 adhesion molecule reverses blockage of differentiation in human acute myeloid leukemia.

Blockage in myeloid differentiation characterizes acute myeloid leukemia (AML); the stage of the blockage defines distinct AML subtypes (AML1/2 to AML5). Differentiation therapy in AML has recently raised interest because the survival of AML3 patients has been greatly improved using the differentiating agent retinoic acid. However, this molecule is ineffective in other AML subtypes. The CD44 surface antigen, on leukemic blasts from most AML patients, is involved in myeloid differentiation. Here, we report that ligation of CD44 with specific anti-CD44 monoclonal antibodies or with hyaluronan, its natural ligand, can reverse myeloid differentiation blockage in AML1/2 to AML5 subtypes. The differentiation of AML blasts was evidenced by the ability to produce oxidative bursts, the expression of lineage antigens and cytological modifications, all specific to normal differentiated myeloid cells. These results indicate new possibilities for the development of CD44-targeted differentiation therapy in the AML1/2 to AML5 subtypes.

Acute Disease↗

PubMed1998

Nucleotide sequence and analysis of the new chromosomal abortive infection gene abiN of Lactococcus lactis subsp. cremoris S114.

A 7.275-kb DNA fragment which encodes resistance by abortive infection (Abi+) to bacteriophage was cloned from Lactococcus lactis subsp. cremoris S114. The genetic determinant for abortive infection was subcloned from this fragment. This gene was found to confer a reduction in efficiency of plating and plaque size for prolate-headed bacteriophage phi 53 (group I homology) and for small isometric-headed bacteriophage phi 59 (group III homology). This new gene, termed abiN, is predicted to encode a polypeptide of 178 amino acid residues with a deduced molecular mass of 20,461 Da and an isoelectric point of 4.63. No homology with any previously described genes was found. A probe was used to determine the presence of this gene only in S114 from 31 strains tested.

Bacteriophages↗

PubMed1998

Acute and long-lasting changes in extracellular-matrix chondroitin-sulphate proteoglycans induced by injection of chondroitinase ABC in the adult rat brain.

Lattice-like perineuronal accumulations of extracellular-matrix proteoglycans have been shown to develop during postnatal maturation and to persist throughout life as perineuronal nets (PNs) in many brain regions. However, the dynamics of their reorganization in adults are as yet unknown. The aim of the present study was to examine the capability of PNs for reconstitution after experimental destruction and to search for possible consequences of extracellular-matrix degradation for neurons and glial cells. The changes were induced by single intracortical injections of Proteus vulgaris chondroitinase ABC and studied after postinjection periods of 1 day to 5 months. The N-acetylgalactosamine-binding Wisteria floribunda agglutinin (WFA), an antibody against chondroitin-sulphate proteoglycans, three antibodies recognizing initial chondroitin or chondroitin-sulphate moieties ('stubs') of proteoglycan core proteins, an antibody against the hyaluronan-binding protein component of versican, and biotinylated hyaluronectin, which binds to hyaluronan, were used as cytochemical markers. One day postinjection, the WFA-binding sites and hyaluronan were shown to be almost completely removed within a circumscribed digestion zone. The staining of different core-protein components revealed only fragments of PNs. These changes were found to be partly compensated 4 weeks after injection of chondroitinase ABC. After 8 and 12 weeks postinjection, the cytochemical and structural characteristics as well as the area-specific distribution patterns of PNs were progressively reconstituted. At 5 months postinjection, they could not be distinguished from those in untreated tissue. In contrast to such transient changes, a diffuse chondroitin-sulphate proteoglycan immunoreactivity persisted in the neuropil. Loss of neurons or alterations of their structure as well as reactions of glial cells were not observed. We conclude from this study that PNs, enzymatically destroyed in the adult rat brain, can be completely reconstituted, but the restoration of their extracellular-matrix components needs several months.

Age Factors↗

PubMed1998

Effect of aging on neointima formation and hyaluronan, hyaluronidase and hyaluronectin production in injured rat aorta.

Few studies have examined the effect of aging on arterial wall response to injury, and the results are discordant. Moreover, the effect of aging on hyaluronan synthesis in injured vessels is unknown. The aim of this present study was to determine the effect of aging on neointima formation and hyaluronan (HA), hyaluronidase and hyaluronectin production in injured rat aorta. Aorta was analysed in sham-operated rats (group D0) and 14 (D14) and 28 (D28) days after injury using biochemical and immunohistochemical techniques. Uninjured aorta of old rats was more thickened than that of young rats; it showed a decreased number of arterial smooth muscle cells (ASMC) and was characterized by HA accumulation in the intima and increased hyaluronidase activity. Intima-media wet weight was significantly increased in young rats at D14 and D28 but remained unchanged in old rats. DNA content was significantly enhanced at D14 in both young and old rats. DNA decreased slightly in young rats at D28 but significantly in old rats to return to control level. HA content and hyaluronidase activity in the intima-media were markedly increased in young rats at D14 (+148% and +116% respectively) but slightly in old rats (+23% and +15% respectively). Both HA and hyaluronidase activity continued to increase at D28, but remained more produced in young rats. The immunohistochemical analysis showed the formation of a thickened neointima in young rats, which was associated with strong expression of HA and HN. Neointima of old rats was reduced; it also showed strong expression of HA and HN but their distributions were different from those observed in neointima of young rats. In conclusion, aorta of old rats showed an increased amount of HA in the intima and elevated activity of hyaluronidase. Injury induced formation of a significant neointima in young rats but not in old rats. This was correlated with more HA and hyaluronidase production in injured aorta of young rats. As HA is considered to increase extracellular matrix space and to promote ASMC proliferation and migration, our findings suggest that HA may be implicated in intima thickening with age and after injury.

Aging↗

PubMed1998

Hyaluronectin is produced by oligodendrocytes and Schwann cells in vitro.

A hyaluronectin (HN)-like antigen was found in rat O-2A progenitors and oligodendrocytes, as well as in Schwann cells and in their culture medium. The HN-like antigen secreted in culture supernatants had a higher molecular mass than HN extracted from rat brain at acidic pH. In vitro the secreted HN-like antigen was spontaneously and slowly degraded into species whose Mr was close to that of HN found in acidic brain extract. In brain or nerve neutral pH extracts, both HN-like antigen and HN were present. The high Mr of the secreted antigen, the homology in amino acid sequences between HN and N-terminal domain of PG-M/versican, in addition to a positive hybridization between Schwann cell RNAs and a probe obtained with primers derived from HN sequences also found in versican suggested that HN is closely related to the large proteoglycan PG-M/versican. The presence in Schwann cell extract of a HN mRNA whose Mr was compatible with the size expected for HN showed that HN may be directly secreted by cells and not only the consequence of a proteolytic cleavage. The similarity of HN with PG-M (V3) suggested that HN found in vivo could be the result of an alternative splicing of a single gene. We conclude that HN as other members of the PG-M/versican family is a marker of oligodendrocytes and Schwann cells in culture.

Animals↗

PubMed1998

Differentially expressed genes in C6.9 glioma cells during vitamin D-induced cell death program.

C6.9 rat glioma cells undergo a cell death program when exposed to 1, 25-dihydroxyvitamin D3 (1,25-D3). As a global analytical approach, we have investigated gene expression in C6.9 engaged in this cell death program using differential screening of a rat brain cDNA library with probes derived from control and 1,25-D3-treated cells. Using this methodology we report the isolation of 61 differentially expressed cDNAs. Forty-seven cDNAs correspond to genes already characterized in rat cells or tissues. Seven cDNAs are homologous to yeast, mouse or human genes and seven are not related to known genes. Some of the characterized genes have been reported to be differentially expressed following induction of programmed cell death. These include PMP22/gas3, MGP and beta-tubulin. For the first time, we also show a cell death program induced up-regulation of the c-myc associated primary response gene CRP, and of the proteasome RN3 subunit and TCTP/mortalin genes. Another interesting feature of this 1,25-D3 induced-cell death program is the down-regulated expression of transcripts for the microtubule motor dynein heavy chain/MAP 1C and of the calcium-binding S100beta protein. Finally 15 upregulated cDNAs encode ribosomal proteins suggesting a possible involvement of the translational apparatus in this cell program. Alternatively, these ribosomal protein genes could be up-regulated in response to altered rates of cellular metabolism, as has been demonstrated for most of the other isolated genes which encode proteins involved in metabolic pathways. Thus, this study presents to our knowledge the first characterization of genes which are differentially expressed during a cell death program induced by 1, 25-D3. Therefore, this data provides new information on the fundamental mechanisms which participate in the antineoplastic effects of 1,25-D3 and on the machinery of a cell death program in a glioma cell line.

Animals↗

PubMed1997

Hyaluronectin blocks the stimulatory effect of hyaluronan-derived fragments on endothelial cells during angiogenesis in vitro.

Hyaluronic acid (HA) is a glycosaminoglycan of the extracellular matrix. Its fragmentation by the hyaluronidase, secreted by tumor cells, facilitates tumor invasion and the HA degradation products generated stimulate angiogenesis. We report here that the HA-binding protein hyaluronectin (HN) inhibits the stimulatory effect of HA-derived fragments on the proliferation and migration of endothelial cells in vitro, and hampers the organization of endothelial cells into capillary-like structures. Since HN strongly inhibits endothelial cell adhesion to immobilized HA, it is postulated that HN acts by impairing the binding to endothelial cells of HA fragments generated by hyaluronidase, thereby neutralizing the effect of HA degradation products on angiogenesis. Our results reveal a new mechanism by which the angiogenesis induced by HA fragments is modulated by HN.

Animals↗

PubMed1997

Increased hyaluronidase levels in breast tumor metastases.

Hyaluronidase, a matrix-degrading enzyme, was assayed in extracts from breast primary tumors and regional metastases using a pool of human sera as a standard. Optimal activities of tumor extracts and serum were found for concentrations of 0.15-0.20 M NaCl in pH 3.8-4.0 buffer. In evaluating contamination by serum due to vascular proliferation, we expressed our results as the ratio of the entire activity (mU/l extract) on serum albumin content of tumors (g/l). Median (interquartile range) activities were 9.02 (6.04-14.34) for primary tumors and 37.36 (24.06-99.63) mU/g albumin for metastases. The difference was significant. Zymographic analysis showed that 3 bands of activity were detected which corresponded to 68, 53 and 49 kDa for tumoral hyaluronidase. The same pattern was observed for cellular extracts of breast cancer cell line CAL51, demonstrating that hyaluronidase detected in tumor extracts had mainly a cellular origin. Our results suggest that hyaluronidase may be involved in the metastatic process.

Breast Neoplasms↗

PubMed1997

Maternal protein restriction early in rat pregnancy alters brain development in the progeny.

We assessed the effects of a dietary protein restriction (5% vs. 20% casein in diet) initiated at conception and imposed during the first 2 weeks of rat gestation on postnatal brain development. At the end of the malnutrition period, protein-restricted animals exhibited significantly smaller fetal body weight and brain cortical thickness than controls. At birth and thereafter, body weight was normalized in the progeny. Similarly, brain weight and cytoarchitecture were normal in postnatal animals. In contrast, we observed, during the first 2 postnatal weeks, several abnormalities of brain development which affected all the studied areas for most of the studied parameters: (i) delayed astrocytogenesis as shown by a reduced GFAP staining; (ii) delayed production of hyaluronan in the extracellular matrix studied with binding of biotinylated hyaluronectin; (iii) abnormal neuronal differentiation as shown by reduced expression of MAP-5 and increased expression of MAP-1; (iv) abnormal synaptogenesis as shown by the increased expression of synaptophysin in the basal ganglia; (v) decreased programmed cell death. In adult prenatally protein-restricted animals, all the above parameters were normalized excepted MAP-1 labeling which remained high. In addition, we observed slight alterations of the ventilatory response to hypoxia in adult animals. The present study demonstrates that early protein malnutrition during embryonic development induces multiple, transient alterations of brain development. However, the almost complete normalization in adults of brain architecture and differentiation as well as our physiological data strongly suggest a remarkable plasticity of the developing brain following an early aggression.

Aging↗

PubMed1997

The origin of hyaluronectin in human tumors.

The origin of tumor stroma hyaluronectin (HN), a glycoprotein that binds to hyaluronan (HA), has long remained unknown. Histological observations of human tumors suggest that tumor HN could originate from stroma fibroblasts, and in some cases from inflammatory cells. The fibroblast origin was confirmed by the discovery of HN-like antigen along with hyaluronan in culture medium of tumor-derived fibroblasts. An HA-binding protein was characterized in the culture medium of peripheral blood mononuclear cells (PBMC) in both normal subjects and tumor-bearing patients and was found to be human HN. Cultivated monocytes did not produce HA. HN was not related to the HA-binding site CD44. Sequencing of brain HN-derived peptides demonstrated that each determined peptide sequence was similar to a sequence of the proteoglycan PG-M/versican, suggesting that HN is the HA-binding moiety of the proteoglycan. One probe was synthesized from human PBMC by polymerase chain reaction with primers derived from HN sequences also found in versican. Northern blots were positive only with HN-producing cells. The main RNAs were in the 6-8 kb range, and there was a limited proportion of smaller RNA, which was compatible with the size expected from the HN molecular mass. Southern blotting of monocytes and tumor cells demonstrated that the gene was limited to a unique band. We conclude that HN, an extracellular component of brain, connective embryonic, inflammatory and tumoral tissues, is a PG-M/versican-derived molecule. Our results suggest that tumor HN, which originates from fibroblasts and monocytes of tumor stroma, is a molecular component of the host-tumor relationship and could play a role in the regulation of HA activity in oncogenesis.

Adenocarcinoma↗

PubMed1997

[Production of hyaluronidase by cultured human tumor cells].

The presence of hyaluronidase was detected at pH 3.8 in eight out of twelve human cancer cell line culture media. Eight cell lines derived from primary tumours and four from metastases. In three culture media the enzymatic activity was lower than 0.035 pU/cell/h. In five others (in a hepatoma cell line and in four metastasis-derived cell lines) the activity was higher than 0.057 pU/cell/h. A tumour-derived fibroblast culture was negative. The optimal activity was observed at a pH comprised between 3.6 and 4. Salt inhibition of hyaluronidase was reversible. The enzyme was denaturated by a 10-min heating at 70 degrees C. The enzyme was not strictly specific for hyaluronan hydrolysis but also digested chondroitin sulfates. PH20, a spermatozoid protein that has homologies with the bee venom hyaluronidase, was not expressed by cell lines tested.

Chromatography, High Pressure Liquid↗

PubMed1997

Characterization of proteoglycan-containing perineuronal nets by enzymatic treatments of rat brain sections.

Proteoglycans are among the major extracellular matrix components of the central nervous system. In the cerebral cortex and many subcortical regions, chondroitin sulphate proteoglycans, which are related to the aggrecan-versican-neurocan family, have been detected immunocytochemically in perineuronal nets that surround various types of neurons. This indicates that, in the brain, there is a nonhomogeneous but defined distribution of extracellular matrix components. The present study is a further attempt to characterize the perineuronal nets in the cerebral cortex. Sections obtained from fixed and unfixed rat brains were subjected to different enzymatic treatments prior to the visualization of perineuronal nets using N-acetylgalactosamine-binding Wisteria floribunda agglutinin, antibodies against chondroitin sulphate proteoglycans or hyaluronectin, and biotinylated hyaluronectin which detects hyaluronan. In all perineuronal nets the binding of the Wisteria floribunda agglutinin was abolished after the incubation of sections with chondroitinase ABC. The protein components of the proteoglycan complexes became easier to digest after removal of chondroitin sulphate chains or hyaluronan. Since only quantitative, and not qualitative, differences in the labelling properties and the structural appearance of cortical perineuronal nets were observed after the various treatments, it is concluded that, with regard to their proteoglycan composition, these structures have common basic properties.

Animals↗

PubMed1997

Hyaluronan: fundamental principles and applications in cancer.

Hyaluronan (HYA) plays a particular role in cancer cell microenvironment. A component of the desmoplasia. HYA is associated to other macromolecules and contributes to the net structure of the matrix. Cancer cells exhibit binding sites (CD44, RHAMM) for HYA. The cell adhesion to HYA can influence the cell motility and different factors could interplay to facilitate cell detachment from HYA. HYA protects cancer cells against immune cell attack. Serum HYA is often increased in metastatic patients.

Animals↗

PubMed1996

Hyaluronan and hyaluronectin production in injured rat thoracic aorta.

The aim of our study was to investigate the production of hyaluronan (HA) by the intima-media during the sclerotic response to aortic injury with a catheter balloon in the rat. In addition we analyzed, for the first time in this model, the production of a glycoprotein (hyaluronectin, HN) which binds specifically to HA. HA and HN were analyzed in control (D0), 14 (D14) and 28 (D28) days after injury using biochemical and immunohistochemical techniques. Intima-media DNA content and wet weight increased significantly on D14 and declined on D28 (but remained significantly increased in comparison to controls). HA content (median in D0 = 448 ng) increased significantly on D14 (2P < 0.04) and on D28 (2P < 0.02). HN content (median in D0 = 920 ng) increased significantly on D14 (2P < 0.05) but decreased on D28 to return to the control level. On D0 the amount of HN was about 3 times higher than that of HA (median ratio HA/HN = 0.34). The ratio remained unchanged on D14 but significantly increased on D28 (2P < 0.02). HPLC and Western blotting showed no difference between HN extracted from normal aorta and HN extracted from injured aorta at D14. Different isoforms of HN were present in both cases, ranging from 400 to 45 kDa. The HA increase on D14 and D28 was not related to a change in hyaluronidase activity of aortic tissue. Immunohistochemical analysis showed at D0 a small amount of HA around arterial smooth muscle cells (ASMC) in media, at D14 more HA was localized around and between ASMC in media and neointima but at D28 it was localized mainly near the vessel lumen. HN formed all the time (D0, D14 and D28) a continuous layer localized near the vessel lumen. In vitro studies showed that production of HA and HN was stimulated when ASMC proliferate and HA at high concentrations (1-100 micrograms/ml) reduced, in a dose dependent manner, ASMC growth. In conclusion our results show that both neointima formation in vivo and ASMC proliferation in vitro correlated with increased HA and HN production. This suggests that HA and HN are probably involved in the formation of neointima. On the other hand, the finding that HA continued to increase in the aorta when neointima decreased and that high concentrations of HA reduce ASMC proliferation in culture suggest that HA might be involved in the regression of neointima.

Animals↗

PubMed1996

Evidence of involvement of CD44 in endothelial cell proliferation, migration and angiogenesis in vitro.

Angiogenesis is essential for tumor growth and metastasis. In the process of angiogenesis, the interaction between adhesive proteins of endothelial cells and extracellular matrix components plays an important role by mediating cell attachment, which is indispensable for their motility, and by transmitting the regulatory signals for cell locomotion and proliferation. In this study, we examined the hypothesis that CD44 expressed on the endothelial cell surface is involved in the angiogenesis process. The experiments using calf pulmonary artery endothelial cells (CPAE) and a human microvascular endothelial cell line (HMEC-1) show that a monoclonal antibody against CD44 (clone J 173) inhibits endothelial cell proliferation by about 30% and migration by 25-50%, and abolishes the stimulating effect of hyaluronan polysaccharides on endothelial cell migration and proliferation. This antibody also suppresses the capillary formation of CPAE in an in vitro model of angiogenesis using fibrin matrix. These results provide evidence of the involvement of endothelial-cell-associated CD44 in angiogenesis.

Animals↗

PubMed1996

Molecular cloning of a levocabastine-sensitive neurotensin binding site.

A search for sequences homologous to the neurotensin receptor cDNA in a rat hypothalamic library has identified a novel neurotensin receptor (NTR-2). The 1539 bp cDNA encodes a 416 amino acid protein and shows highest homology to the previously cloned neurotensin receptor (NTR-1) (64% homology and 43% identity). Binding and pharmacological studies demonstrate that NTR-2 expressed in COS cells recognizes neurotensin (NT) with high affinity as well as several other agonists and antagonists. However, a fundamental difference was found; unlike NTR-1, NTR-2 recognizes, with high affinity, levocabastine, a histamine H1 receptor antagonist previously shown to compete with NT for low-affinity binding sites in brain.

Amino Acid Sequence↗

PubMed1996

CD44 exon variant 6 epitope and hyaluronate synthase are expressed on HT29 human colorectal carcinoma cells in a SCID mouse model of metastasis formation.

The factors which lead to the formation of metastases are generally poorly understood; however the expression of a particular variant of the cell adhesion molecule CD44 may be important in facilitating metastasis formation in colon cancer. The aim of the present study was to investigate the expression of CD44 exon v 6 (CD44v6), hyaluronate (one of its ligands), and hyaluronate synthase, in a clinically relevant animal model of metastatic colon carcinoma. HT29 human colon carcinoma cells were injected subcutaneously between the scapulae of severe combined immunodeficient (SCID) mice and left for 3 weeks (by which time the tumours had produced metastases in the lungs). Morphological observations at the tumour-host interface were consistent with the dissociation of neoplastic cells from the primary tumours, and the ability of these cells to migrate through the extracellular matrix facilitating metastasis formation. Immunohistochemically detectable hyaluronate synthase expression was increased in vivo compared with the parent cell line in vitro. CD44v6 expression and hyaluronate were increased around single cells at the periphery of tumours compared with the central regions. CD44v6 and hyaluronate snythase expression were co-expressed in the same cells. Indeed, the present study is the first to demonstrate hyaluronate synthase expression by an epithelial cell type.

Animals↗

PubMed1996

Does the lectin Helix pomatia agglutinin bind to hyaluronic acid in breast and colon cancer?

The lectin Helix pomatia agglutinin (HPA) has been successfully used as an indicator of metastatic spread in a number of epithelial neoplasms including breast, colorectal, gastric, prostate and oesophageal cancers. Despite its use, the binding partners of HPA in tissue sections have not been defined at the molecular level. HPA has two main binding specificities: 1) N-acetylgalactosamine (GalNac) and 2) N-acetylglucosamine (GlucNac). In order to determine whether HPA binds to hyaluronic acid (a GlucNac-containing glycosaminoglycan) a dot blot assay was performed which confirmed hyaluronic acid as a binding partner for HPA. In the next step, experiments were performed using hyaluronate lyase predigestion before HPA application on clinical and experimental human tumours grown in severe combined immunodeficient (SCID) mice to assess whether the HPA binding in the dot blot system also occurred in tissue sections. The results from all samples indicate that hyaluronate lyase pretreatment does not alter HPA binding to tumour cells both in the patients' samples and in the human cancer cell lines grown in SCID mice. This also indicates that HPA binds to similar carbohydrate residues in patients' samples and in SCID mouse-grown human tumour cells. It seems, therefore, unlikely that HPA recognises hyaluronic acid as a binding partner in tissue sections of human tumours, and hence GalNac-containing glycoproteins seem to be the more likely ligands for HPA in cancer cells.

Animals↗