Impact of prior systemic therapy on lymphocytic infiltration in surgically resected breast cancer brain metastases.

PURPOSE: Tumor-infiltrating lymphocytes (TILs) have been positively correlated with response to systemic therapy for triple-negative and HER2 + subtypes and improved clinical outcomes in early breast cancer (BC). Less is known about TILs in metastatic sites, particularly brain metastases (BM), where unique immune regulation governs stromal composition. Reactive glial cells actively participate in cytokine-mediated T cell stimulation. The impact of prior medical therapy (chemotherapy, endocrine, and HER2-targeted therapy) on the presence of TILs and gliosis in human breast cancer brain metastases (BCBM) has not been previously reported. METHODS: We examined prior treatment data for 133 patients who underwent craniotomy for resection of BMs from the electronic medical record. The primary endpoint was overall survival (OS) from the time of BM diagnosis. We examined the relationship between prior systemic therapy exposure and the histologic features of gliosis, necrosis, hemorrhage, and lymphocyte infiltration (LI) in BCBMs resected at subsequent craniotomy in univariate analyses. RESULTS: Complete treatment data were available for 123 patients. BCBM LI was identified in 35 of 116 (30%) patients who had received prior systemic treatment versus 5 of 7 (71.4%) who had not {significant by Fisher's exact test p = 0.045}. There were no statistically significant relationships between prior systemic therapy and the three other histologic variables examined. CONCLUSIONS: This observation suggests that systemic therapy may interfere with the immune response to BCBMs and cause exhaustion of anti-tumor immunity. This motivates clinical investigation of strategies to enhance LI for therapeutic benefit to improve outcomes for patients with BCBMs.

Implementation of a non-lethal biopsy punch monitoring program for mercury in smallmouth bass, Micropterus dolomieu Lacepède, from the Eleven Point River, Missouri.

A non-lethal biopsy method for monitoring mercury (Hg) concentrations in smallmouth bass (Micropterus dolomieu; smallmouth) from the Eleven Point River in southern Missouri USA was evaluated. A biopsy punch was used to remove a muscle tissue plug from the area immediately below the anterior dorsal fin of 31 smallmouth. An additional 35 smallmouth (controls) were held identically except that no tissue plug was removed. After sampling, all fish were held in a concrete hatchery raceway for 6 weeks. Mean survival at the end of the holding period was 97 % for both groups. Smallmouth length, weight and Fulton's condition factor at the end of the holding period were also similar between plugged and non-plugged controls, indicating that the biopsy procedure had minimal impact on growth under these conditions. Tissue plug Hg concentrations were similar to smallmouth Hg data obtained in previous years by removing the entire fillet for analysis.

Effects of historical lead-zinc mining on riffle-dwelling benthic fish and crayfish in the Big River of southeastern Missouri, USA.

The Big River (BGR) drains much of the Old Lead Belt mining district (OLB) in southeastern Missouri, USA, which was historically among the largest producers of lead-zinc (Pb-Zn) ore in the world. We sampled benthic fish and crayfish in riffle habitats at eight sites in the BGR and conducted 56-day in situ exposures to the woodland crayfish (Orconectes hylas) and golden crayfish (Orconectes luteus) in cages at four sites affected to differing degrees by mining. Densities of fish and crayfish, physical habitat and water quality, and the survival and growth of caged crayfish were examined at sites with no known upstream mining activities (i.e., reference sites) and at sites downstream of mining areas (i.e., mining and downstream sites). Lead, zinc, and cadmium were analyzed in surface and pore water, sediment, detritus, fish, crayfish, and other benthic macro-invertebrates. Metals concentrations in all materials analyzed were greater at mining and downstream sites than at reference sites. Ten species of fish and four species of crayfish were collected. Fish and crayfish densities were significantly greater at reference than mining or downstream sites, and densities were greater at downstream than mining sites. Survival of caged crayfish was significantly lower at mining sites than reference sites; downstream sites were not tested. Chronic toxic-unit scores and sediment probable effects quotients indicated significant risk of toxicity to fish and crayfish, and metals concentrations in crayfish were sufficiently high to represent a risk to wildlife at mining and downstream sites. Collectively, the results provided direct evidence that metals associated with historical mining activities in the OLB continue to affect aquatic life in the BGR.

Toxicokinetics of 2,2',4,4'- and 3,3',4,4'-tetrachlorobiphenyl congeners in the house cricket.

Recently, investigators have reported bioaccumulation of polychlorinated biphenyls (PCBs) in insects collected at hazardous waste sites. Little is known about differences in bioaccumulation of particular PCB congeners in insects. The authors report on the toxicokinetics of a non-coplanar ortho substituted congener, 2,2',4,4'-tetrachlorobiphenyl (PCB-47) and a coplanar non-ortho substituted PCB, 3,3',4,4'-tetrachlorobiphenyl (PCB-77) in topically treated house crickets. Penetration and excretion were more rapid for PCB-47 than for PCB-77. PCB-47 demonstrated a high affinity for abdomen, gut, and gonads, whereas, PCB-77 had a greater affinity for the head and thorax. Sex differences were observed in excretion and distribution. Females had an increased excretion of PCBs and increased accumulation in gut and gonads compared to males. Differences in lipophilicity can explain differences in distribution between sexes and of compounds within sexes. However, to explain the observed difference in penetration, it is necessary to invoke explanations based on stearic factors associated with coplanarity of the molecules. Differences in excretion are likely related to sex differences in mixed-function oxidase activities. These toxicokinetic data suggest that the more toxic coplanar congener, PCB-77, is likely to be more persistent in crickets compared to PCB-47, despite its lower lipophilicity.

Alkoxyresorufin metabolism in white-footed mice at relevant environmental concentrations of Aroclor 1254.

Recent investigations have detected polychlorinated biphenyl (PCB) body burdens in wild white-footed mice (Peromyscus leucopus) captured at hazardous waste sites. Insufficient information is currently available to interpret the toxicological significance of these body burdens. In an effort to provide this information, we investigated hepatic changes and PCB body burdens in white-footed mice following a 21-day dietary exposure to a PCB mixture, Aroclor 1254. Dietary concentrations tested were 0, 2.5, 25, 50, and 100 mg Aroclor 1254/kg diet (reported as ppm). Liver weights were significantly increased at all concentrations except 2.5 ppm. Ethoxyresorufin O-dealkylase (EROD) activity, an aryl hydrocarbon hydroxylase-type substrate, was significantly increased at all PCB concentrations, but the dose-response tended to plateau above 25 ppm. Pentoxyresorufin O-dealkylase (PROD) activity, a putative phenobarbital-type substrate, was significantly increased in a dose-dependent manner at 25 ppm PCB and above, with no plateau response. Pentobarbital sleep time was significantly decreased at 25 ppm, but not at 2.5 ppm. Results indicate white-footed mice undergo a mixed-type induction pattern following exposure to Aroclor 1254, with EROD the most sensitive indicator of PCB exposure. This investigation identified a no observed effect concentration for liver weights and PROD activity at 2.5 ppm in the diet which is equivalent to a body burden of 2.0 mg Aroclor 1254/kg wet wt of mice; the no observed effect concentration for EROD is below these levels. These results support the use of EROD, PROD, and liver weight as biomarkers of PCB exposure in field-captured rodents.

Dopamine and octopamine in whole body extracts of the bulb mite.

Using high performance liquid chromatography with electrochemical detection, whole body extracts of the bulb mite, Rhizoglyphus echinopus (Fumouze and Robin), were found to contain the biogenic amines dopamine and octopamine at concentrations of 4.3 +/- 0.6 and 2.3 +/- 1.4 ng g-1 wet weight, respectively. Adrenaline, noradrenaline, tyramine, N-methyldopamine, N-acetyldopamine, and 5-hydroxytryptamine, if present, were below the limits of detectability. This is the initial demonstration of the presence of octopamine in a mite species.

Effects of aluminum on the biochemical composition of Atlantic salmon.

Biomolecule content, dry weight, and wet weight of Atlantic salmon, Salmo salar L., were monitored during 60-day exposure to aluminum, which began before hatch and extended to about the end of the alevin stage. The Atlantic salmon exposed to aluminum concentrations of 0, 33, 71, 124, or 264 micrograms/L at pH 5.5 for 60 days were evaluated for growth, survival, and biochemical effects on days 15, 30, and 60. Growth and survival were significantly reduced at the two highest aluminum concentrations by day 60. RNA and DNA were the most sensitive biomolecules monitored and were significantly reduced at the three highest aluminum concentrations by day 60. RNA:protein and RNA:DNA ratios were no more responsive than growth and survival. The lipid content of salmon was not significantly affected by exposure to aluminum. It was concluded that biomolecule content or ratios were not particularly sensitive indicators of growth effects in Atlantic salmon exposed to aluminum.

Structural characterization of phospholamban in cardiac sarcoplasmic reticulum membranes by cross-linking.

The native form of phospholamban in cardiac sarcoplasmic reticulum membranes was investigated using photosensitive heterobifunctional cross-linkers, both cleavable and noncleavable, and common protein modifiers. The photosensitive heterobifunctional cleavable cross-linker ethyl 4-azidophenyl-1, 4-dithiobutyrimidate was used in native SR vesicles and it cross-linked phospholamban into an apparent phospholamban-phospholamban dimer and into an approximately 110,000-Da species. The phospholamban dimer migrated at approximately 12,000 Da on sodium dodecyl sulfate-polyacrylamide gels, and upon cleavage of the cross-linker before electrophoresis the dimer disappeared. The approximately 110,000-Da cross-linked species was not affected by boiling in sodium dodecyl sulfate prior to electrophoresis. This cross-linked form of phospholamban migrated approximately 5500 Da above the Ca2(+)-ATPase, which was visualized using fluorescein 5'-isothiocynate, a fluorescent marker that binds specifically to the Ca2(+)-ATPase. p-Azidophenacyl bromide, iodoacetic acid, and N-ethylmaleimide, all of which react with sulfhydryl groups, were also employed to further characterize phospholamban in native sarcoplasmic reticulum membranes. Cross-linking with p-azidophenacyl bromide resulted in only monomeric and dimeric forms of phospholamban as observed on sodium dodecyl sulfate-polyacrylamide gels. Iodoacetic acid and N-ethylmalemide were found to be effective in disrupting the pentameric form of phospholamban only when reacted with sodium dodecyl sulfate solubilized sarcoplasmic reticulum. In view of these findings, the amino acid sequence of phospholamban was examined for possible protein-protein interaction sites. Analysis by hydropathic profiling and secondary structure prediction suggests that the region of amino acids 1-14 may form an amphipathic alpha helix and the hydrophobic surface on one of its sites could interact with the reciprocal hydrophobic surface of another protein, such as the Ca2(+)-ATPase.

Temporal and spatial variation of RNA content in nymphs of the mayfly, Stenonema femoratum (Say).

Temporal variation of RNA: weight, RNA: protein, RNA: DNA, and protein: DNA ratios in Stenonema femoratum (Say) was assessed by monitoring at the same site in a small midwestern stream over a two-year period. Variation components were estimated using a two-way mixed model analysis of variance with weight class and collection date as main effects. About 30% of the variation in RNA concentration was within-cell variation associated with differences among individuals and analytical error. Temporal variation accounted for about 50% of the total variability in RNA concentration data. RNA: weight had 11% and RNA: protein had 17% variation attributed to weight class effects indicating that these ratios were dependent on the size of the mayfly. RNA: DNA ratios were not influenced by mayfly weight, suggesting that this ratio would be the most appropriate for comparing populations based on means of mixed individual sizes. Comparison of mean RNA: DNA ratios among collection dates yielded significant increases in the ratio during late fall in two consecutive years. Spatial variations in RNA: DNA ratios of S. femoratum collected from sites of similar habitats were not significantly different. These data encourage the future development of RNA concentration as an in situ biochemical indicator of growth rate and possibly of stress related effects on growth rate.

Protein and nucleic acid content in Daphnia magna during chronic exposure to cadmium.

Daphnia magna Straus were exposed to cadmium concentrations of 0, 0.4, 0.8, 2.1, 4.3, and 7.2 micrograms/liter for 21 days. Protein, RNA, DNA, glycogen and lipid content were measured after 4, 7, and 21 days of exposure, and the results were related to survival and reproduction. The no observed effect concentration (NOEC) for survival, protein growth, and reproduction at Day 21 was 2.1, 0.8, and 0.8 microgram/liter cadmium, respectively. Protein growth was most sensitive to cadmium exposure following the rapid growth phase which occurred between Days 4 and 6 of growth and development. All concentrations of cadmium produced significant reductions in protein growth at Day 7 indicating that the NOEC was less than 0.4 micrograms/liter. RNA:protein and protein:RNA:DNA ratios, which are related to the growth rate of daphnids, were most appropriately monitored at Day 4 which was prior to the rapid growth phase. Protein:RNA:DNA ratio at Day 4 identified the same NOEC as identified by protein growth and reproduction after 21 days of exposure.

Effects of fenvalerate on biochemical parameters, survival, and reproduction of Daphnia magna.

Daphnia magna were exposed to fenvalerate at nominal concentrations of 0.5, 0.25, 0.13, 0.06, and 0.03 micrograms/liter for 21 days. On Days 7 and 21 of exposure, levels of ribonucleic acid (RNA), deoxyribonucleic acid (DNA), adenosine diphosphate (ADP), adenosine triphosphate (ATP), glycogen, and lipid were measured, and the results were related to survival and reproduction during the 21-day test period. Survival was not significantly (alpha = 0.05) affected by the 21-day exposure; however, reproduction was reduced at fenvalerate concentrations of 0.25 and 0.5 micrograms/liter. On Day 7, protein, RNA, ADP, caloric equivalents, and glycogen were also significantly reduced at fenvalerate concentrations of 0.25 and 0.5 micrograms/liter. Thus, these biochemical parameters identified the same no observable effect concentration (NOEC) as did reproduction. Variables derived from biochemical parameters were related to reduced protein growth and reproduction. Decreases in protein/RNA and glycogen/lipid ratios and increases in ADP/DNA, ADP/ATP, protein/RNA/DNA, and lipid/DNA ratios were observed at Day 7 for those daphnia exposed to 0.5 micrograms/liter of fenvalerate. The only derived variable that reflected the reduced protein growth at Day 7 in daphnia exposed to 0.25 micrograms/liter of fenvalerate was the glycogen/lipid ratio. Biochemical determinations at Day 21 indicated that the organisms exposed to 0.25 micrograms/liter of fenvalerate were not different from controls, whereas those exposed to 0.5 micrograms/liter were still affected.