Effects of the antifouling compound, Irgarol 1051, on a simulated estuarine salt marsh ecosystem.

Toxicity effects of the antifouling compound, Irgarol 1051, were examined using a simulated estuarine salt marsh ecosystem. The 35 day mesocosm exposure incorporated tidal flux and contained seawater, sediments, marsh grass, and estuarine biota. Irgarol (10.0 microg/l) caused a significant reduction in phytoplankton biomass and primary productivity. HPLC pigment analysis indicated significant effects of irgarol on both phytoplankton and periphyton community composition, with decreased concentrations of pigments representative of diatom species. There was also a significant decrease in dissolved oxygen levels in the 10.0 microg/l irgarol treatment. Growth of the hard shell clam was significantly reduced in the 1.0 and 10.0 microg/l irgarol treatments. The effects observed occurred at irgarol concentrations greater than those typically measured in the environment. Prolonged exposure, the accumulation of irgarol in sediments, plant, or animal tissues, and the interaction of irgarol with other chemicals in the environment; however, could increase risk.

Toxicity of the antimicrobial compound triclosan and formation of the metabolite methyl-triclosan in estuarine systems.

Triclosan, a commonly used antimicrobial compound, has been measured in aquatic systems worldwide. This study exposed marine species to triclosan to examine effects primarily on survival and to investigate the formation of the degradation product, methyl-triclosan, in the estuarine environment. Acute toxicity was assessed using the bacterium Vibrio fischeri, the phytoplankton species Dunaliella tertiolecta, and three life stages of the grass shrimp Palaemonetes pugio. P. pugio larvae were more sensitive to triclosan than adult shrimp or embryos. Acute aqueous toxicity values (96 h LC50) were 305 microg/L for adult shrimp, 154 microg/L for larvae, and 651 microg/L for embryos. The presence of sediment decreased triclosan toxicity in adult shrimp (24 h LC50s were 620 microg/L with sediment, and 482 microg/L without sediment). The bacterium was more sensitive to triclosan than the grass shrimp, with a 15 min aqueous IC50 value of 53 microg/L and a 15 min spiked sediment IC50 value of 616 microg/kg. The phytoplankton species was the most sensitive species tested, with a 96 h EC50 value of 3.55 microg/L. Adult grass shrimp were found to accumulate methyl-triclosan after a 14-day exposure to 100 microg/L triclosan, indicating formation of this metabolite in a seawater environment and its potential to bioaccumulate in higher organisms. Triclosan was detected in limited surface water sampling of Charleston Harbor, SC at a maximum concentration of 0.001 microg/L, substantially lower than the determined toxicity values. These findings suggest triclosan poses low acute toxicity risk to estuarine organisms; however, the potential for chronic, sublethal, and metabolite effects should be investigated.

Activation of MYCN in a case of non-Hodgkin's lymphoma.

We have examined a series of non-Hodgkin's lymphomas (NHL) for evidence of expression of the MYC gene family. Northern blot analysis of RNA samples derived from 11 non-malignant reactive lymphoid tissues and 33 NHL was used to investigate expression of MYC, MYCL and MYCN. As expected MYC expression was detected in all samples. The levels of MYC expression were quantified by densitometry and appeared to be 3-8 fold higher in high grade NHL than in the low grade NHL or non-malignant lymphoid tissue. No expression of MYCL was detected in any sample. Expression of MYCN was however observed in one sample, which had been diagnosed as a T-cell high grade NHL. A detailed cytogenetic analysis of this sample proved difficult to obtain but, by using fluorescence in-situ hybridization (FISH), we were able to demonstrate that on one of the chromosomes 2 the MYCN gene was localised to a translocation breakpoint region. It therefore appears that in NHL it is possible for MYCN, like MYC in Burkitt lymphoma, to be activated as a result of a chromosome translocation event.

MDR-1 expression in non-Hodgkin's lymphomas is unrelated to treatment intensity or response to therapy.

Over-expression of the MDR-1 gene, which codes for P-glycoprotein, is thought to be an important mechanism in the drug resistance exhibited by many tumours. A number of chemotherapeutic agents which induce MDR-1 expression are also components of combination chemotherapies that are used in the treatment of high grade non-Hodgkin's lymphomas (NHL). We have therefore examined expression of MDR-1 in a series of NHL by Northern blot analysis as well as investigated the localization of P-glycoprotein by immunohistochemistry. The series included 11 hyperplastic reactive nodes and tonsils, 17 low grade NHL and 15 high grade NHL. The levels of MDR-1 mRNA were quantified by scanning densitometry and comparison with levels of glucose-6-phosphate dehydrogenase (G6PD). The MDR-1 mRNA was observed in both non-malignant and NHL tissues. Immunohistochemical staining revealed that expression of MDR-1 mRNA in reactive nodes was related to the presence of P-glycoprotein in lymphocytes, however, P-glycoprotein was apparent in both the reactive lymphocytes and tumour cells in the NHL samples. Elevated mRNA levels (2-3 fold increase) were observed in some low grade and high grade NHL relative to those observed in reactive lymphoid tissue. There appeared to be little correlation, however, between expression of the MDR-1 gene and either treatment intensity or response to therapy. The drug resistance that is often encountered in NHL patients is therefore likely to involve mechanisms other than over-expression of P-glycoprotein.

Elevated levels of MDM-2 and p53 expression are associated with high grade non-Hodgkin's lymphomas.

The role of p53 in the evolution of non-Hodgkin's lymphomas (NHL) is unclear. Mutations of the p53 gene appear to be relatively uncommon but stabilized p53 protein, as detected by immunohistochemistry, has indicated a more frequent involvement of p53. As dysfunction of p53 protein has also been suggested to occur after overexpression of the mdm-2 protein, we have therefore investigated a series of non-malignant hyperplastic reactive lymphoid tissues and NHL to examine whether the levels of expression of MDM-2 correlated to positivity of p53 protein staining. Northern blot analysis of MDM-2 expression was compared to glucose-6-phosphate dehydrogenase (G6PD) expression by densitometry to quantify the relative levels of MDM-2 expression. Consistent low levels of MDM-2 expression were observed in non-malignant lymphoid tissue and in low grade NHL, however, 13/15 high grade NHL exhibited a 2-15-fold increase in MDM-2 expression. Interestingly similar elevations in p53 mRNA expression were also observed in 6/15 high grade NHL. Positive staining of the p53 protein did not, however, correlate with elevated mRNA levels of either MDM-2 or p53. The significance of these observations is discussed.

Analysis of the p53 gene, its expression and protein stabilization in non-Hodgkin's lymphomas.

We have investigated the p53 gene, expression of its mRNA, and stabilization of its protein in a series of non-Hodgkin's lymphomas (NHLs). Immunohistochemistry revealed positive staining of the p53 protein in node biopsies from 6/36 NHL patients, all of whom had high-grade disease. The remaining NHL samples, together with 3 reactive nodes, showed either negative staining or the staining of only occasional cells. In one case that exhibited intense nuclear staining in 90% of the cells, a mutation in the p53 gene was also observed. There was no evidence of rearrangements of the p53 gene in any of the NHL samples. Although p53 mRNA could not be detected in nonmalignant tissue, it was apparently overexpressed in 16/38 NHL, but this did not correlate with positive staining of the p53 protein. These data suggest that p53 dysfunction might play an important role in the evolution of some cases of NHL, and that mechanisms other than mutation of the p53 gene may be involved in stabilizing the p53 protein in these neoplasms.

Investigation of the expression of housekeeping genes in non-Hodgkin's lymphoma.

Studies of quantitative changes in gene expression in malignant cells have often used housekeeping genes as controls against which the level of expression of a gene under study could be compared. We have now examined whether the expression of the most commonly used of these housekeeping genes can be regarded as reliable controls for gene expression studies in non-Hodgkin's lymphoma (NHL). We have used Northern blot analysis to compare the levels of expression of beta-actin, alpha-tubulin, beta 2-microglobulin and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to that of ribosomal RNA. These studies demonstrated that whereas there was a reasonable correlation between the relative levels of rRNA and housekeeping gene expression in reactive hyperplastic nodes, there were major differences in the relative levels of expression of the housekeeping genes in both low and high grade lymphomas; only GAPDH showed any degree of consistency. These observations indicated that housekeeping gene expression was not a reliable control for estimating changes in the level of expression of other genes in NHL, and instead suggested that 18S or 28S rRNA expression offered a more accurate method of RNA quantitation.