Insolubility of Cr2O3 in Bioaccessibility Tests Points to Requirement for a New Human Oral Reference Dose for Trivalent Chromium

Bioaccessibility measurements have the potential to improve the accuracy of risk assessments and reduce the potential costs of remediation when they reveal that the solubility of chemicals in a matrix (e.g., soil) differs markedly from that in the critical toxicity study (i.e., the key study from which a toxicological or toxicity reference value is derived). We aimed to apply this approach to a brownfield site contaminated with chromium, and found that the speciation was Cr^III, using a combination of alkaline digestion/diphenylcarbazide complexation and X-ray absorption near edge structure analysis. The bioaccessibility of Cr₂O₃, the compound on which a reference dose for Cr^III is based, was substantially lower (<0.1%) than that of the Cr^III in the soils, which was a maximum of 9%, giving relative bioaccessibility values of 13,000% in soil. This shows that the reference dose is based on essentially an insoluble compound, and thus we suggest that other compounds be considered for toxicity testing and derivation of reference dose. Two possibilities are CrCl₃·6H₂O and KCr(SO₄)₂·12H₂O, which have been used for derivation of ecological toxicity reference values and are soluble at a range of dosing levels in our bioaccessibility tests.     

THE ABSORPTION OF P,P′-DDE BY LARVAE OF AGLAIS URTICAE (LEPIDOPTERA: NYMPHALIDAE)

Fifth instar larvae of Aglais urticae L. absorb single and double doses of 27.5 g of p,p-DDE at a constant rate 8–72 hours after dosing. There is little difference between the rates of absorption by the blood of larvae given doses on two sites, and the sum of the rates of absorption by the blood of larvae given single doses, so that the blood does not limit the rate at which apolar substances pass through the cuticle.

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Zusammenfassung Die Rate, mit welcher p,p-DDE durch die Kutikula des 5.Larvenstadiums von A. urticae eindringt, wurde durch direkte Messung der absorbierten Menge bestimmt: Verluste durch Stoffwechsel und Exkretion waren zu vernachlässigen. Die Raupen erhielten entweder einzelne Dosen von 27,5 g DDE in 1 l 1,4-Dioxan auf den Thorax oder das Abdomen, oder zwei Dosen, eine auf den Thorax und eine auf das Abdomen. Die Larven wurden 8, 24 und 72 Stunden nach der Applikation auf DDE untersucht. Die Aufnahmerate durch die Larven, die zwei Dosen erhalten hatten, war mit 0,6–1,1 ppm/Std ähnlich der Summe der Aufnahmeraten durch die Larven, denen einzelne Dosen auf Thorax oder Abdomen verabreicht worden waren, so daß die Aufnahmerate wahrscheinlich völlig durch Faktoren innerhalb der Kutikula begrenzt wird und die Absorptionskapazität des Blutes für diese apolare Substanz keinen entscheidenden Faktor darstellt.

     

Validity of the tritiated thymidine method for estimating bacterial growth rates: measurement of isotope dilution during DNA synthesis.

The rate of tritiated thymidine incorporation into DNA was used to estimate bacterial growth rates in aquatic environments. To be accurate, the calculation of growth rates has to include a factor for the dilution of isotope before incorporation. The validity of an isotope dilution analysis to determine this factor was verified in experiments reported here with cultures of a marine bacterium growing in a chemostat. Growth rates calculated from data on chemostat dilution rates and cell density agreed well with rates calculated by tritiated thymidine incorporation into DNA and isotope dilution analysis. With sufficiently high concentrations of exogenous thymidine, de novo synthesis of deoxythymidine monophosphate was inhibited, thereby preventing the endogenous dilution of isotope. The thymidine technique was also shown to be useful for measuring growth rates of mixed suspensions of bacteria growing anaerobically. Thymidine was incorporated into the DNA of a range of marine pseudomonads that were investigated. Three species did not take up thymidine. The common marine cyanobacterium Synechococcus species did not incorporate thymidine into DNA.     

Volume regulation by flounder red blood cells in anisotonic media

The nucleated high K, low Na red blood cells of the winter flounder demonstrated a volume regulatory response subsequent to osmotic swelling or shrinkage. During volume regulation the net water flow was secondary to net inorganic cation flux. Volume regulation the net water flow was secondary to net inorganic cation flux. Volume regulation after osmotic swelling is referred to as regulatory volume decrease (RVD) and was characterized by net K and water loss. Since the electrochemical gradient for K is directed out of the cell there is no need to invoke active processes to explain RVD. When osmotically shrunken, the flounder erythrocyte demonstrated a regulatory volume increase (RVI) back toward control cell volume. The water movements characteristic of RVI were a consequence of net cellular NaCl and KCl uptake with Na accounting for 75 percent of the increase in intracellular cation content. Since the Na electrochemical gradient is directed into the cell, net Na uptake was the result of Na flux via dissipative pathways. The addition of 10(-4)M ouabain to suspensions of flounder erythrocytes was without effect upon net water movements during volume regulation. The presence of ouabain did however lead to a decreased ration of intracellular K:Na. Analysis of net Na and K fluxes in the presence and absence of ouabain led to the conclusion that Na and K fluxes via both conservative and dissipative pathways are increased in response to osmotic swelling or shrinkage. In addition, the Na and K flux rate through both pump and leak pathways decreased in a parallel fashion as cell volume was regulated. Taken as a whole, the Na and K movements through the flounder erythrocyte membrane demonstrated a functional dependence during volume regulation.     

In vivo and in vitro inhibition of rat liver vitamin D3-25-hydroxylase activity by 19-hydroxy-10(S),19-dihydrovitamin D3

Rats treated with varying amounts of 19-hydroxy-10(S),19-dihydrovitamin D3 prior to administration of physiologic doses of vitamin D3 exhibit normal intestinal calcium transport but are unable to mobilize bone calcium. In contrast, 19-hydroxy-10(R),19-dihydrovitamin D3 had no inhibitory activity. Circulating serum levels of 25-hydroxy[3H]vitamin D3 and 1 alpha, 25-dihydroxy[3H]vitamin D3 are markedly suppressed but not totally eliminated in animals predosed with 19-hydroxy-10(S),19-dihydrovitamin D3 before [3H]vitamin D3. Hepatic 25-hydroxy[3H]vitamin D3 levels were approximately equal in both 19-hydroxy-10(S),19-dihydroviotamin D3 treated and untreated rats. However, the rate of conversion of [3H]vitamin D3 to 25-hydroxyvitamin D3 in vivo is greatly reduced in the treated rats. The inhibitory vitamin analogue was also show to block hepatic microsomal 25-hydroxylation in vitro. These results indicate that 19-hydroxy-10(S),19-dihydrovitamin D3 is a specific inhibitor for a hepatic microsomal vitamin D3-25-hydroxylase system.     

Cytochemical analysis of intracellular calcium distribution in the anterior pituitary of the rat

Summary In an attempt to assign morphologic identities to previously distinguished functional calcium compartments in the anterior pituitary of the rat, we employed the potassium pyroantimonate technique for cation localization. Tissues were incubated for In at 37°C in control medium; with 10mM theophylline; or with depolarizing amounts of potassium. Precipitate was quantified on photomicrographs of tissue prepared for electron microscopy with a Talos Systems Digitizer. The nature of the electron dense precipitate was dependent on the experimental state of the tissue. Treatment with 5 mM EGTA abolished the dense precipitate. Electron microprobe analysis also confirmed that calcium was the predominant cation in the observed precipitate. The most significant changes in precipitate deposition occurred along the plasma membrane, the limiting membrane of secretory granules and within mitochondria. Dense precipitate was present along the plasma membrane only in cells treated with potassium. Control tissue exhibited higher levels of precipitate associated with the limiting membrane of secretory granules than either theophylline-treated or potassium-treated tissue. Mitochondria contained more precipitate in potassium-treated tissue than in controls; the mitochondria of theophylline-treated tissue contained intermediate levels of precipitate. Addition of either theophylline or depolarizing amounts of potassium has been associated with hormone secretion in anterior pituitary tissue of normal rats. Kinetic studies in our laboratory indicate that intracellular calcium shifts occur. The pyroantimonate technique is useful in verifying morphologically the calcium compartments involved in shifts in intracellular calcium.     

Production of infectious hepatitis delta virus in vitro and neutralization with antibodies directed against hepatitis B virus pre-S antigens.

Hepatitis delta virus (HDV) particles were produced in Huh7 human hepatoma cells by transfection with cloned hepatitis B virus (HBV) DNA and HDV cDNA. The particles were characterized by their buoyant density, the presence of encapsidated viral RNA, and their ability to infect primary cultures of chimpanzee hepatocytes. Successful infection was evidenced by the appearance of increasing amounts of intracellular HDV RNA after exposure to particles. Infection was prevented when particles were incubated with antibodies directed against synthetic peptides specific for epitopes of the pre-S1 or pre-S2 domains of the HBV envelope proteins before exposure to hepatocytes. These data demonstrate that HDV particles produced in vitro are infectious and indicate (i) that infectious particles are coated with HBV envelope proteins that contain the pre-S1 and pre-S2 regions, (ii) that epitopes of the pre-S1 and pre-S2 domains of HBV envelope proteins are exposed at the surface of HDV particles, and (iii) that antibodies directed against those epitopes have neutralizing activity against HDV.     

Ultrastructure of bacteria and the proportion of Gram-negative bacteria in marine sediments

Bacteria in sediments from the surface aerobic layer (0–1 cm) and a deeper anaerobic layer (20–21 cm) of a seagrass bed were examined in section by transmission electron microscopy. Bacteria with a Gram-negative ultrastructure made up 90% of bacteria in the surface layer, and Gram-positive bacteria comprised 10%. In the anaerobic zone, Gram-negative bacteria comprised 70% and Gram-positive bacteria 30% of the bacterial population. These differences were highly significant and support predictions of these proportions made from muramic acid measurements and direct counting with fluorescence microscopy. Most cells were enveloped in extracellular slime layers or envelopes, some with considerable structural complexity. The trophic value to animals of these envelopes is discussed. A unique organism with spines was observed.     

SIVcpz cross-species transmission and viral evolution toward HIV-1 in a humanized mouse model

HIV-1 evolved from its progenitor SIV strains, but details are lacking on its adaptation to the human host. We followed the evolution of SIVcpz in humanized mice to mimic cross-species transmission. Increasing viral loads, CD4⁺ T-cell decline, and non-synonymous mutations were seen in the entire genome reflecting viral adaptation.