Biology Reference
In-Depth Information
6.5.1.1
In vitro studies with hPBMC's
LPS (5 μg/mL) was added to hPBMC's followed by treatment with
different concentrations of β-PGG (
66
) to explore this hypothesis (two
independent trials, Fig. 6.21). After 4 hours of incubation, the TNFα
level in the supernatant, as measured by an ELISA kit, was significantly
depleted compared to the control where hPBMC's were treated only with
LPS. A concentration of 5 μM for β-PGG appeared to be the point of
maximum inhibition (as much as 90% decrease in TNFα levels
compared to the control), since higher concentrations did not result in
significant further lowering of the TNFα output. Incubation beyond 4
hours (to 24 hours) eliminated any inhibition caused by β-PGG. Thus,
there appeared to be a time dependence as well. After these promising
in vitro
studies, β-PGG was tested next with live rats.
Fig. 6.22
In vivo
suppression by β-PGG of LPS-stimulated TNFα secretion in rats.
6.5.1.2
In vivo testing of live rats experiencing septic shock
Initial
in vivo
experiments determined that a 50-60 mg/rat administration
of β-PGG (
66
) led to a lethal level of hypotension. Use of a 30 mg/rat
dose led to no changes in blood glucose or blood pressure levels,
whereas use of 4 mg/rat caused no inhibition of TNFα production with
LPS treated rats. Therefore, a “safe” value of 17 mg/rat was utilized in
all subsequent experiments. Injection of the rats with either saline or β-
PGG was followed by LPS administration 10 minutes later. The TNFα
