Table 2. Effects of TLR4 and TLR4 inhibitors on palmitic acid-induced cytokine release from cultured astrocytes.

Primary astrocyte cultures were established as described in Methods, and exposed to BSA, or BSA-conjugated palmitic acid (100 μM final concentration, see Methods). Selected cells were also exposed to PA in the presence of increasing concentrations of a pharmacologic inhibitor of TLR2/4 (OxPAPC), a neutralizing antibody to TLR4 (FA6.152), or a neutralizing antibody to TLR2 (FA6.152). All inhibitors were applied to cells 45 min before administration of palmitic acid. Release of TNFα, IL-6, and IL-1β; and MTT conversion was measured after 24 h. Data were compiled from 2 separate experiments, and are means and SEM of 10–30 dishes per group.

		TNFα (pg/ml)	IL-6 (pg/ml)	IL-1β (pg/ml)	MTT (% BSA)
BSA		125.7 ± 22.1	758.9 ± 75.6	128.5 ± 18.2	100.0 ± 5.3
100 μM PA		995.1 ± 46.5***	4632.2 ± 177.6***	1191.6 ± 61.5***	101.0 ± 6.9
PA with OxPAPC	10 μM	108.3 ± 13.6###	2428.5 ± 159.3###	129.3 ± 23.8###	124.1 ± 14.3
	30 μM	98.1 ± 16.9###	2382.2 ± 290.6###	112.3 ± 40.1###	125.0 ± 11.4
	50 μM	110.0 ± 20.2###	2165.5 ± 350.8###	163.4 ± 47.8###	120.8 ± 16.1
PA with TLR4 Ab	0.5 μg	677.2 ± 54.2##	2774.2 ± 479.1###	881.1 ± 116.6	116.6 ± 13.0
	1 μg	648.2 ± 67.5###	2530.3 ± 502.0###	752.5 ± 65.0##	120.8 ± 14.3
	5 μg	528.6 ± 38.9###	1924.4 ± 341.0###	619.3 ± 100.8###	109.4 ± 15.7
PA with TLR2 Ab	0.5 μg	1004.9 ± 77.2	4083.2 ± 231.4	1180.0 ± 83.5	101.0 ± 5.2
	1 μg	953.5 ± 69.4	4379.2 ± 163.2	1249.7 ± 115.8	111.9 ± 4.4
	5 μg	1155.9 ± 179.3	4705.8 ± 342.1	1727.7 ± 279.6	90.4 ± 6.6

^***indicates significant (p<0.001) increases in cytokine release from astrocytes treated with palmitic acid as compared to control cells treated with just BSA, and

^{### and ##}indicates significant (p < 0.01 and p<0.001, respectively) decreases in cytokine release from astrocytes treated with palmitic acid in the presence of inhibitors as compared to cells treated with palmitic acid alone.