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M L Watson

Publications and source records attributed to M L Watson.

At least 19 recordsLinked to original sources
PubMed2001

Characterization of initiated cells in N-methylnitrosourea-induced carcinogenesis of the CNS in the adult rat.

Glial tumors may originate from the malignant transformation of multipotent glial progenitor cells, but tools to study malignant transformation leading to gliomas are limited by the lack of biological systems that represent early stages of this disease in adult animals. In order to characterize the initiated cells that give rise to gliomas, we have employed the N-methylnitrosourea (MNU) model for induction of brain tumors in adult rats (Rushing et al., 1998). Specifically, we have isolated and cultured transformed (premalignant) cells from normal-appearing brains of rats exposed to MNU for 10 weeks and from histologically abnormal brains of rats exposed to MNU for 15 weeks. We compared them with cells cultured from control animals under identical conditions. Cultured cells were classified according to their morphology, immunophenotype, karyotype, proliferation capacity, and tumorigenicity in athymic mice. Cultures from untreated normal rat brains grew as monolayers and had normal karyotypes (42 X,Y), epithelioid morphology, and slow proliferative capacity (doubling time > 120 h). In contrast, cultured cells from brains of MNU-exposed animals had karyotypes that ranged from normal to highly aneuploid. Aneuploid lines grew rapidly in multilayers (doubling time < 24 h), had differentiated astrocytic or oligodendroglial morphology and immunohistochemical staining profile, and yielded tumors in athymic mice. Initiated cells with minor chromosomal aberrations assumed mixed bipolar or tripolar morphologies in high density cultures, proliferated rapidly, but showed contact inhibition and failed to induce tumors when injected s.c. in athymic mice. In general, lines showing no evidence of chromosomal aberrations had the most epithelioid morphology, proliferated slowly (doubling time > 72 h), and retained strict contact growth inhibition. The presumed undifferentiated glial progenitor cells in culture from either control or MNU-treated rats variably expressed markers such as vimentin, nestin, and NG2 proteoglycan, and they weakly expressed the mature astrocytic or oligodendroglial markers glial fibrillary acidic protein or galactocerbroside, respectively. These cultures differentiated to bipolar-tripolar morphology with concomitant maturation to a GFAP+ or GalC+ phenotype upon exposure to secondary messengers such as dibutyryl-cyclic-AMP and/or growth factors such as basic fibrillary growth factor. Continuous stimulation with these messengers resulted in terminal differentiation and consequent death upon withdrawal of the stimulus. These results provide information that could lead to detailed characterization of initiated, premalignant cells in the adult brain and to a better understanding of glial carcinogenesis.

Animals↗
PubMed2000

Investigation and treatment of patients with teeth affected by tooth substance loss: a review.

Tooth substance loss, an increasing problem, may result from erosion, abrasion and attrition, often with more than one of these acting together. Investigation requires a detailed history and examination. The aim of treatment may be prevention of further damage in less affected cases. The treatment of severe tooth substance loss may be complex, especially in view of the reduced amounts of tooth substance which may be available and the need to find space because of the compensatory over-eruption of worn teeth.

Bruxism↗
PubMed2000

Recurrent (nonfamilial) hemangioblastomas involving spinal nerve roots: case report.

OBJECTIVE AND IMPORTANCE: Spinal nerve root hemangioblastomas are rare and are reported mainly in patients with von Hippel-Lindau (VHL) syndrome. The pathogenesis of so-called nonfamilial lesions is virtually unknown. We discuss, mainly from a molecular perspective, a unique patient with sporadic, recurrent hemangioblastomas restricted to spinal nerve roots. CLINICAL PRESENTATION: A 53-year-old man who had had a surgically corrected lumbosacral meningomyelocele presented on at least three occasions during a 17-year period with multifocal capillary hemangioblastomas involving spinal nerve roots. On each occasion, tumors appeared on a different nerve root, with the majority located in the midcervical segments. The patient had no clinical features or family history of VHL syndrome. TECHNIQUE: To obtain a clearer understanding of the pathogenesis of this unusual case and its relationship to VHL syndrome, molecular analysis of the VHL gene was performed by use of complete sequence analysis and loss of heterozygosity studies on deoxyribonucleic acid derived from the patient's blood leukocytes and three separately resected hemangioblastomas. CONCLUSION: Germ-line molecular analysis performed on all three exons in the VHL gene coding region did not indicate that any mutations were present. Loss of heterozygosity analysis of deoxyribonucleic acid from the three hemangioblastoma resections showed normal heterozygosity in the 3p25-26 region. Complete VHL gene sequence analysis did not demonstrate a somatic mutation in the coding region of the VHL gene in any of the three tumors, thereby supporting the loss of heterozygosity data that a molecular event directly involving the VHL gene may not be the causative factor in their tumorigenesis.

Base Sequence↗
PubMed1999

Burkholderia cepacia is resistant to the antimicrobial activity of airway epithelial cells.

There has been much interest recently in the antimicrobial properties of cationic peptides called beta-defensins from epithelial cells. Human beta-defensin (hBD)-1 and -2 have been particularly implicated in cystic fibrosis (CF) patients, where their inhibition by high salt concentrations may explain in part the susceptibility of the CF lung to bacterial infection. In this work, we have employed a simple co-culture system using the 16-HBE human bronchial epithelial cell line to assess growth inhibitory activity against Pseudomonas aeruginosa and Burkholderia cepacia. In medium alone, P. aeruginosa proliferated more than 100,000-fold, whereas in the presence of 16-HBE cells or 16-HBE-conditioned medium, bacterial proliferation was less than 100-fold. Raising the salt concentration of cell-free 16-HBE conditioned medium to approximately 200 mM significantly reduced this growth inhibitory activity. In contrast, there was no evidence of epithelial-derived growth inhibitory activity against two strains of B. cepacia. RT-PCR analysis indicated expression of the hBD-2 mRNA in 16-HBE cells, but not hBD-1. These data demonstrate for the first time that B. cepacia is resistant to epithelial-derived antimicrobial substances and argue against them being important in the defense against this organism in the lung.

Burkholderia cepacia↗
PubMed1999

New gene family defined by MORC, a nuclear protein required for mouse spermatogenesis.

Mammalian spermatogenesis is a complex developmental process. The analysis of mouse mutations has provided insight into biochemical pathways required for completion of this process. We previously described the autosomal recessive mouse morc TgN(Tyr)1Az(microrchidia) mutation, a serendipitous transgenic insertional mutation which causes arrest of spermatogenesis prior to the pachytene stage of meiosis prophase I. We now report the molecular characterization of the morc locus and positional cloning of a gene disrupted by the morc TgN(Tyr)1Az mutation. This gene, which we term Morc, encodes a 108 kDa protein expressed specifically in male germ cells. The transgene integrated within the first intron of Morc and was accompanied by an intragenic deletion of approximately 13 kb of genomic sequences, removing exons 2-4 and abrogating expression of the wild-type transcript. Analysis of the MORC protein sequence revealed putative nuclear localization signals, two predicted coiled-coil structural motifs and limited homology to GHL (GyraseB, Hsp90, MutL) ATPase. Epitope-tagged MORC protein expressed in COS7 cells localized to the nucleus. We also cloned the human MORC homolog and show that it too is testis-specific, but closely related human genes are transcribed in multiple somatic tissues. Homologous proteins are also present in zebrafish, nematodes, slime mold and plants. Thus, cloning of Morc defines a novel gene family whose members are likely to serve important biological functions in both meiotic and mitotic cells of multicellular organisms.

Amino Acid Sequence↗
PubMed1999

Dioleylphosphatidylglycerol inhibits the expression of type II phospholipase A2 in macrophages.

We have recently shown that modified natural pulmonary surfactant Curosurf inhibits the synthesis of type II phospholipase A2 (sPLA2-II) by cultured guinea-pig alveolar macrophages (AM). The goal of the present study was to identify the surfactant components and the mechanisms involved in this process. We show that protein-free artificial surfactant (AS) mimicked the inhibitory effect of Curosurf, suggesting that phospholipid components of surfactant play a role in the inhibition of sPLA2-II expression. Among surfactant phospholipids, dioleylphosphatidylglycerol (DOPG) was the most effective in inhibiting the synthesis of sPLA2-II. By contrast, the concentrations of platelet-activating factor (PAF)-acetylhydrolase and lysophospholipase activities remained unchanged, indicating that inhibition of sPLA2-II synthesis was caused by a specific effect of surfactant. The effect of DOPG on sPLA2-II synthesis was concentration-dependent and was accompanied by a rapid and time-dependent uptake of DOPG by AM whereas dipalmitoylphosphatidylcholine (DPPC) was only marginally taken up. Curosurf, AS, and DOPG inhibited tumor necrosis factor-alpha (TNF-alpha) secretion, a key step in the induction of sPLA2-II synthesis by AM, in contrast to DPPC which had only a marginal effect. We conclude that phospholipid components, especially DOPG, play a major role in the inhibition of sPLA2-II synthesis by surfactant and that this effect can be explained, at least in part, by an impairment of TNF-alpha secretion.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1999

Anti-inflammatory actions of interleukin-13: suppression of tumor necrosis factor-alpha and antigen-induced leukocyte accumulation in the guinea pig lung.

The Th2 cytokine interleukin (IL)-13 is believed to play an important role in the development of allergy, although it has also been ascribed anti-inflammatory roles in several experimental models. In this study, we have examined the effects of human recombinant IL-13 on eosinophilic lung inflammation in the guinea pig. IL-13 (1 to 100 ng, given by intratracheal instillation) did not elicit airway eosinophil recruitment. A pronounced accumulation of eosinophils, as well as monocyte/macrophages, was elicited by intratracheal instillation of guinea pig tumor necrosis factor alpha (gpTNF-alpha). Intratracheal administration of IL-13 (1 to 100 ng) given immediately prior to exposure to gpTNF-alpha resulted in a dose-related suppression of eosinophil and monocyte/macrophage accumulation in the airways, as assessed by bronchoalveolar lavage (BAL) and eosinophil peroxidase activity in whole-lung homogenates. IL-13 treatment also reduced BAL fluid (BALF) leukocyte accumulation induced by subsequent aerosol antigen challenge of sensitized guinea pigs. Antigen challenge also resulted in elevated levels of immunoreactive eotaxin and eosinophil-stimulating activity in BALF, although only the latter was reduced significantly by IL-13 instillation prior to challenge. In contrast to the suppressive effects of IL-13, instillation of human recombinant IL-4 (100 ng) alone elicited an increase in BALF monocyte/macrophage numbers, and IL-4 was unable to inhibit gpTNF-alpha-induced leukocyte accumulation. Hence, IL-13 (but not human IL-4) exhibits an anti-inflammatory action in the airways of gpTNF-alpha- or antigen-challenged guinea pigs, by mechanisms that may involve the decreased generation of eosinophil-stimulating activity in the airways.

Animals↗
PubMed1998

Identification of morc (microrchidia), a mutation that results in arrest of spermatogenesis at an early meiotic stage in the mouse.

The microrchidia, or morc, autosomal recessive mutation results in the arrest of spermatogenesis early in prophase I of meiosis. The morc mutation arose spontaneously during the development of a mouse strain transgenic for a tyrosinase cDNA construct. Morc -/- males are infertile and have grossly reduced testicular mass, whereas -/- females are normal, indicating that the Morc gene acts specifically during male gametogenesis. Immunofluorescence to synaptonemal complex antigens demonstrated that -/- male germ cells enter meiosis but fail to progress beyond zygotene or leptotene stage. An apoptosis assay revealed massive numbers of cells undergoing apoptosis in testes of -/- mice. No other abnormal phenotype was observed in mutant animals, with the exception of eye pigmentation caused by transgene expression in the retina. Spermatogenesis is normal in +/- males, despite significant transgene expression in germ cells. Genomic analysis of -/- animals indicates the presence of a deletion adjacent to the transgene. Identification of the gene inactivated by the transgene insertion may define a novel biochemical pathway involved in mammalian germ cell development and meiosis.

Animals↗
PubMed1998

Interleukin 8 and monocyte chemoattractant protein 1 production by cultured human airway smooth muscle cells.

Leukocyte accumulation and activation are key events in the pathogenesis of inflammatory lung disease. The ability of human airway smooth muscle cells (HASM) to contribute to the inflammatory process by its ability to produce the chemokines interleukin (IL) 8, monocyte chemotactic protein (MCP-1) and regulated on activation, normal T cell expressed and secreted (RANTES) was investigated. Cultured HASM, when stimulated with the pro-inflammatory cytokines IL-1 alpha (0.01-1 ng/ml) or tumour necrosis factor alpha (TNF-alpha, 0.3-30 ng/ml), synthesize and release substantial amounts of IL-8, as assessed by specific immunoassay, bioasssay (elevation of intracellular free calcium in human neutrophils), and upregulation of mRNA. These stimuli also increased MCP-1 production and mRNA expression, but RANTES mRNA expression was not detected at 24 h. The smooth muscle spasmogen endothelin 1 (1 microM) was unable to stimulate IL-8 or MCP-1 release or mRNA expression. These data indicate that HASM may constitute an important source of leukocyte attractants in the inflamed lung, where the inducing stimuli, IL-1 alpha and TNF-alpha, are also likely to be present.

Bronchi↗
PubMed1998

Glial tumors in the MNU rat model: induction of pure and mixed gliomas that do not require typical missense mutations of p53.

Gliomas were induced in adult male Sprague-Dawley rats by continuous exposure to 100 ppm of N-nitrosmethylurea (MNU) in drinking water. Latency periods for such tumors were 20 and 50 weeks following completion of exposure intervals of 20, 15, and 10 weeks, respectively. Based on histomorphology and the pattern of GFAP immunoreactivity, a large percentage of MNU-induced tumors (>40%) were anaplastic mixed gliomas, having both neoplastic astrocytic and oligodendroglial components. Typical oligodendrogliomas and astrocytomas also occurred less frequently. Unlike the majority of tumors induced by ethylnitrosourea (ENU), MNU yielded glial tumors that did not express synaptophysin. Anaplastic mixed gliomas and glioblastoma multiforme (GBMs) had no missense p53 mutations in the commonly mutated exons 4 through 8 and did not overexpress wild-type p53, suggesting that MNU-induced oncogenesis in rat brain tumors may not require inactivation/alteration of the p53 tumor suppressor gene. The K-ras gene was also analyzed and found to have no activating mutations in brain tumors. This model is suitable for studying genetic events leading to the majority of gliomas that apparently express functional p53.

Animals↗
PubMed1997

Recombinant guinea pig and human RANTES activate macrophages but not eosinophils in the guinea pig.

To characterize the biologic activities of potential mediators of allergic inflammation, we have cloned, expressed, and purified guinea pig RANTES (gpRANTES). cDNA for gpRANTES was cloned from Con A-stimulated guinea pig spleen cells. A high level of gpRANTES expression in Escherichia coli was achieved by mutation of a human RANTES (hRANTES) expression construct to obtain a 68-amino acid protein identical with the predicted guinea pig amino acid sequence, assuming an equivalent amino terminus as the human protein. Purified gpRANTES was an effective stimulus of human eosinophils as assessed by increases in intracellular free calcium in fura-2-loaded cells and chemotactic responses in vitro. gpRANTES exhibits similar potency and efficacy to hRANTES. In marked contrast, neither gpRANTES nor hRANTES was able to activate guinea pig peritoneal eosinophils in these assays, even in the presence of IL-5. However, gpRANTES was found to be a potent stimulator of guinea pig peritoneal macrophages. Following tracheal instillation of gpRANTES, a dose-dependent increase in macrophages, but not eosinophils, was observed in gpBAL. Macrophage accumulation was detectable by 6 h and sustained for at least 48 h. These results indicate that RANTES in the guinea pig may have a different cellular selectivity than that described in the human, which may be important in the use of animal models in the analysis of allergic disorders. These selectivities do not appear to be accounted for by differences in guinea pig and human RANTES sequences.

Amino Acid Sequence↗
PubMed1997

Chemokine production by human vascular smooth muscle cells: modulation by IL-13.

1. The production of chemokines by vascular smooth muscle cells (SMC) is implicated in the pathogenesis of atherosclerosis, although the factors regulating chemokine production by these cells are incompletely characterized. 2. We describe the differential stimulation of interleukin-(IL)-8, monocyte chemoattractant protein (MCP)-1 and regulated on activation normal T-cell expressed and secreted (RANTES) synthesis following treatment of human vascular SMC with IL-1alpha or tumour necrosis factor alpha (TNFalpha). Under basal conditions, cultured SMC release very low amounts of IL-8, MCP-1 and RANTES as assessed by specific ELISA. Concentration-response studies with IL-1alpha or TNFalpha revealed that each stimulus induced a similar amount of MCP-1. In contrast approximately three fold more IL-8 was induced by IL-1alpha than by TNFalpha whereas significant RANTES production was induced only by TNFalpha. These findings point to a divergence in the regulation of synthesis of the different chemokines in response to IL-1alpha or TNFalpha stimulation. 3. The T-cell derived cytokines IL-10 and IL-13 were also found to have differential effects on chemokine production by SMC. IL-13, but not IL-10, significantly enhanced IL-8 and MCP-1 release in response to IL-1alpha or TNFalpha. This increase in chemokine release appeared to be accounted for by increased mRNA expression. 4. These findings provide support for the concept that smooth muscle cells can have an active role in a local immune response via the production of chemokines which can be selectively modulated by T-cell derived cytokines.

Cells, Cultured↗
PubMed1997

Ambulatory blood pressure in hypertensive patients with autosomal dominant polycystic kidney disease.

BACKGROUND: Ambulatory blood pressure is more closely correlated with various indices of hypertensive target-organ damage, and is a better prognostic predictor of cardiovascular morbidity and mortality than conventional methods of blood pressure measurement. Autosomal dominant polycystic kidney disease (ADPKD) is complicated by hypertension, progressive renal failure, and an increased risk of cardiovascular mortality. This study investigated the 24-h ambulatory blood pressure profile in patients with ADPKD in view of the sparsity of such data in these patients and the possibility that abnormal diurnal blood pressure variations may have prognostic consequences. METHODS: Ambulatory blood pressure was measured over a 24-h period by the oscillometric method with an automatic non-invasive recorder (SpaceLabs 90207 system) in matched groups of 25 hypertensive patients with ADPKD and 25 patients with essential hypertension. RESULTS: Both groups showed a nocturnal decrease in blood pressure, but this was significantly smaller in patients with ADPKD. There was no evidence of enhanced lability of blood pressure in ADPKD. CONCLUSIONS: The nocturnal fall in blood pressure was attenuated in patients with ADPKD. Further studies are required to assess the importance of this finding and its possible contribution to the progression of renal failure or increased cardiovascular mortality in these patients.

Adult↗