Anti-inflammatory effects of bifidobacteria by inhibition of LPS-induced NF-κB activation

Table 1 Bacterial strains used in this study

Strain oroligonucleotides	Relevant characteristics	Source orPCR product
E. coli
TG1	Cloning host	DSMZ1
D2241	Non-pathogenic intestinal isolate, control	NCC2
Bifidobacterium
NCC362	B. lactis, type strain	NCC2
NCC2705	B. longum, type strain	NCC2
NCC251	B. adolescentis, type strain	NCC2
NCC189	B. bifidum	NCC2
S16	B. bifidum, intestinal isolate from a breast-fed infant	[57]
S17	B. bifidum, intestinal isolate from a breast-fed infant	[57]
E18	B. infantis/longum, intestinal isolate from an adult	[57]
MB226	B. breve, type strain	D. Matteuzzi

¹ DSMZ: Deutsche Sammlung von Mikroorganismen und Zellkulturen; ² NCC: Nestlé Culture Collection.

Table 2 Quantification of NF-κB-driven SEAP activity in the supernatants of HT-29 clone 34 cells

Treatment	RLU ± SD 1
None (untreated control)	2 ± 1
HM	2 ± 1
LPS	5 ± 1
LPS + HM	100 ± 6

¹ Values are mean ± SD of at least 3 independent experiments for each condition performed in triplicate. For each individual experiment luminescence measurements were normalized to give 100 RLU for stimulation with LPS + HM.

Table 3 IL-8 protein in the supernatants of HT-29 cells (mean ± SD, n = 3)

Treatment	IL-8 [μg/L] 1
	moi = 10	moi = 100
None (untreated control)	2 ± 1
LPS+HM	36 ± 3
NCC2705/LPS+HM 1	30 ± 2	7 ± 3
S17/LPS+HM 1	7 ± 2	2 ± 0

¹ Pre-incubation with B. longum NCC2705 or B. bifidum S17 at moi = 10 or 100 was followed by stimulation with LPS + HM for 16 h.

Table 4 Real-time quantification of mRNA levels of several genes in HT-29 cells

Treatment	Relative mRNA level 1± SD of gene for
	IL-8	TNF-α	COX-2	ICAM-1
LPS+HM (positive control)	40.6 ± 17.4	37.4 ± 12.6	5.9 ± 1.7	28.0 ± 10.7
B. longum NCC2705/ LPS+HM 2	11.4 ± 1.1 a	12.7 ± 1.8 a	2.5 ± 1.6	12.2 ± 9.2
B. bifidum S17/ LPS+HM 2	3.5 ± 2.6 a	2.9 ± 2.4 a	1.4 ± 0.6 a	2.0 ± 0.8  a

¹ Results are mRNA levels relative to the negative control (no bacteria, no LPS) and are mean ± standard deviation (SD) of three independent pools of RNA for each condition.

² Pre-incubation with B. longum NCC2705 or B. bifidum S17 at moi = 100 was followed by stimulation with LPS + HM for 4 h.

^aP < 0.05 vs positive control; Student’s t-test.