RBC AChE activity

Table 1. The effect of 1 mM NaF + 20 pM A1C13 on the acetylcholinesterase activity (AChE) in freshly prepared intact RBC and in hemolysate of patients with AD (mean age 72.5 5.1 years), age-matched healthy controls (AM-HS) (72.1 1.6 years), and the group of young healthy subjects (YS) (35.9 8.5 years). Whole venous blood samples were drawn from each subject after overnight fasting., always at 07 30 AM. Red blood cells (RBC) were isolated from the blood of patients with AD, AM-HS, and YS by centrifugation [68], RBC AChE activity was evaluated in intact freshly prepared RBC or hemolyzate following the spectrophotometric method [45] with modifications. Buffer was Tris-HCl, pH 7.5 in the solution of 154 mmol L 1 NaCl, acetylthiocholine iodide was a substrate. Measurement of enzymatic activity was performed in fluorimeter polystyrene cuvettes for 3 min (UV/VIS spectrophotometer Shimadzu, Japan). The effects of 1 mmol L-1 NaF in the presence of 20 pmol L 1 A1C13 were measured. Data are expressed in percentage of the AChE activity in the absence of aluminum and fluoride ions. No differences between the AChE activity were found between the investigated groups...

The LOAELadj (0.054 mg/kg per day) used by ORNL for derivation of the RfD for GB was based on the lowest dose that caused a significant depression in RBC AChE activity in rats (Bucci and Parker 1992). The subcommittee notes that ChE inhibition is typically considered a biomarker of exposure to organophosphate agents rather than an adverse effect. However, it is generally agreed that inhibition of ChE contributes... 

ORNL assigned a factor of 10 for the nncertainty factor for the extrapolation of data from animals to humans (UFa) becanse no evidence suggests that humans are less susceptible than rats to GD. ORNL cited the evidence that rodents have much lower RBC-AChE activity than humans (EUin 1981), suggesting that rats might be more susceptible than hnmans to anti-ChE compounds, but also noted that the lower RBC-AChE... 

The subcommittee considered other possible critical studies for the derivation of the RfD for VX. In a study by Goldman et al. (1988), VX was administered to Sprague-Dawley rats (25 males and 25 females per group) by subcutaneous (s.c.) injection at doses of 0, 0.25, 1.0, and 4.0 pg/kg for 5 days per week for up to 90 days. A dose-dependent decrease in RBC-AChE concentrations was observed in male and female rats compared with controls. Plasma ChE was significantly depressed at day 30 in both sexes administered VX at a dose of 1.0 pg/kg per day, and at days 30, 60, and 90 in both sexes at a dose of 4.0 pg/kg per day. The data from rats exposed for 30 days was reanalyzed by ORNL using analysis of variance and Dunnett s and Scheffe s comparisons. ORNL reported that RBC-AChE activity was significantly lower in both sexes in all dose groups. 

In addition to being found in the nervous system, acetylcholinesterase also occurs in the blood where it is bound to the surface of red blood cells (termed RBC-ChE or RBC-AChE). RBC-AChE activity, as well as the activity of a second type of cholinesterase found in blood plasma (butyrylchoUnesterase, or plasma cholinesterase) have been used to monitor exposure to organophosphate compounds (pesticides and nerve agents). Both RBC-AChE and plasma-ChE activity have been used as bioindicators of potential toxic effects. There is some evidence that RBC-AChE is as sensitive as brain ChE to the effects of nerve agents. Grob and Harvey (1958) reported that the in vitro concentrations producing 50% depression of brain-ChE and RBC-AChE activity were the same in the case of GA (1.5 x 10 mol/L),... 

Acetylthiocholine iodide concentration for maximum RBC-AChE activity. 

Changes in plasma-ChE activity in dosed and control animals are shown in Table 4. Over the course of the study, plasma-ChE activity levels in dosed and control animals appear to be more stable than RBC-AChE activity. It was reported that plasma-ChE activity was decreased by about 55% in dosed females at week 7, and by 37.5% in dosed males at week 3. Mean plasma-ChE activity in the female controls exhibited a slow increase over the 13-week test period (from 1743 lU/L at week -1 to 2891 lU/L at week 13). A similar response was seen in the two lowest dose groups of females. In males, mean plasma-ChE activity in controls was lower than preexposure levels (401 lU/L at week -1) at all weeks except week 3 (413 lU/L). In the dosed groups of males, mean plasma-ChE levels were lower than pre-exposure values at all sampling times. Statistical analysis of the plasma-ChE activity indicated that mean values were significantly lower than controls in the mid- and high-dose females at weeks -1, 1, 3, and 7 but not at week 13, and in the high-dose males at weeks 3 and 7. 

The use of a rat study for developing an RfD for GA is complicated by the fact that rodents have a much lower RBC-AChE activity level compared to humans (Ellin, 1981). By itself, this could cause rats to be relatively more sensitive than humans to anticholinesterase compounds however, the lower RBC-AChE activity may be offset by the presence of ahesterases in the blood of rats. Aliesterases, which are not found in human blood plasma, are known to bind to and, therefore, reduce the toxicity of GB, and a similar mechanism may operate in the case of GA. Other species differences, such as in the rates of aging of the GA-ChE complex, in the rates of synthesis of plasma-ChE in the liver, and in the levels of AChE in the nervous system (see Ivanov et al., 1993) may also result in difference between species in sensitivity to GA. Data are insufficient to more fuUy evaluate these possibihties. There is httle human acute toxicity data that can be compared with the available rat data however, acute toxicity data for primates in general (see Table 2) suggests that humans are likely to be more sensitive than rats. Therefore, for the purpose of this assessment, the standard EPA method will be followed which assumes that humans can be as much as ten times more sensitive to a chemical than laboratory animals. 

Sheep have a much lower RBC-AChE activity level compared to humans, 2.9 /rmoEmL/min versus... 

Comparative analysis of biochemical and physiological parameters studied is indicative of the complete absence of significant changes of RBC-AChE activity in rats of all three groups relative to the control, after exposiue to the RVX doses given above (not shown here). On the contrary, platelets of test animals exposed to RVX differed from the control by their pronoimced instability, an indication of which was development of their spontaneous activation and aggregation (Figure 7.5). 

To study the embryotoxicity of RVX, it was administered to pregnant female rats perorally at a dose I/IOOLD50 (Kiryukhin et at, 2007). RVX had a toxic effect in the females judged by decrease of their RBC-AChE activity at... 

GA, a unitary chemical munition, inhibits AChE, the enzyme responsible for the breakdown of the neurotransmitter ACh. When inhaled, its toxicity is half that of sarin. It depresses plasma and RBC-AChE activities significantly in the blood. At 20-25% of red blood cell AChE baseline, the effect of the nerve agent becomes noticeable. There is no evidence of systemic toxicity other than the cholinesterase activity (Parker et al, 1990 Munro et al, 1994). GA has not been shown to produce OPIDN except at extremely high doses. The cardiac effect of GA conforms to OP-caused arrhythmias and AV block. 

RBC AChE activity also varies among species. In humans, low RBC AChE activity has been associated with increased susceptibility to nerve agents (Leng and Lewalter, 1999). Pigs, sheep, dogs, rabbits, and cats have less RBC cholinesterase activity than humans (Anonymous, 2000b Ellin, 1982). This would make these species more sensitive to nerve agents unless they have an alternative system to combat cholinesterase inhibition. 

Figure 58.2. Selected results of RBC AChE activity determination are shown in Table 58.1.

In general, there were certain tendencies to decrease cholinesterase activity (especially RBC AChE activity) during their work at the department. Some rare examples of suspect intoxications were observed in the cholinesterase monitoring as well as an RBC AChE decrease in workers during their work with high concentrations of nerve agents (e.g. inhalation exposure experiments) (Figure 58.4). 

GB, GD, and VX to determine the in vitro mass balance of these OPs. They developed a mathematically based toxicokinetic model for the estimation of the upper limit of Hu BChE dose required for protection against OP toxicity. The model addressed the relationship between the Hu BChE dose needed to maintain 30% of residual red blood cell (RBC) AChE activity and other parameters (level and duration of OP exposure, bimolecular rate constants of inhibition of Hu AChE and Hu BChE by OPs, and time elapsed from enzyme administration). They validated the Hu BChE dose by in vitro experiments and data from published human studies (Ashani et al., 1998 Ashani and Pistinner, 2004). The proposed model suggested that the upper limit doses of 134, 115, and 249 mg/kg of Hu BChE were sufficient to protect RBC AChE above 30% following a challenge with 1 X LD50 of VX, GD, and GB, respectively. 

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