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Table 1 Local and systemic IL-1Ra after administration of IL-1Ra-producing B. subtilis

From: Mucosal delivery of anti-inflammatory IL-1Ra by sporulating recombinant bacteria

 

Delivery of B. subtilis pSM539

Detection of IL-1Ra

Animal

Route

Time

Local

Serum

Mouse

peritoneal cavity

0 h

-

-

  

3 h

++

+

  

6 h

+

-

  

24 h

-

-

Rat

small intestine

0 h

-

-

  

3 h

++

-

  

4 h

+ (B: 149.4 μg)

-

  

6 h

+/-

-

  

8 h

+/- (B: 16.5 μg)

-

  

24 h

-

n.t.

 

large intestine

0 h

-

-

  

2 h

n.t.

++ (B: 0.6–1.3 μg/ml)

  

3 h

+++

n.t.

  

4 h

+++

++ (B: 0.5–1.9 μg/ml)

  

6 h

++

+

  

8 h

++

+ (B: 0.1–0.4 μg/ml)

  

24 h

+

n.t.

Rabbit

large intestine

0 h

-

- (B/E: 0 μg/ml)

  

0.5 h

n.t.

+ (B: 0.2–0.4 μg/ml)

  

1 h

n.t.

++ (B/E: 0.2–1.2 μg/ml)

  

2 h

n.t.

++ (B/E: 0.6–1.6 μg/ml)

  

3 h

n.t.

++ (B: 0.6–2.0 μg/ml)

  

4 h

++

++ (B/E: 0.4–2.1 μg/ml)

  

6 h

n.t.

++ (B/E: 0.3–0.9 μg/ml)

  

8 h

++

++ (E: 0.6–2.4 μg/ml)

  

24 h

n.t.

+ (B: 0.3 μg/ml)

  1. Mice, rats and rabbits were administered live cells of B. subtilis strain pSM539 (engineered to produce human mature IL-1Ra; 1 × 108-2 × 109 cells/kg) into the peritoneal cavity, the small intestine, or the large intestine. Animals were sacrificed at different time points after bacterial inoculum, and samples of serum and intestinal washings were taken. The presence of human IL-1Ra was assessed in all samples by Western blotting. Semi-quantitative analysis was performed after laser scanning densitometry in comparison to different amounts of standard human recombinant IL-1Ra, and scored as follows: -, no detection; +/- very faint and/or inconsistent detection; + consistently positive; ++ abundant detection; +++ very high levels. Western blotting analysis revealed that IL-1Ra recovered from serum and intestinal washings had the same molecular mass as the standard IL-1Ra. No proteolytic fragments or larger aggregates could be detected. For some samples, quantitative assessment of IL-1Ra was performed by ELISA (E) or by BIAcore (B) analysis. Assessed animals were 2–16/group/time. n.t., not tested.