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. 2004 May;53(5):694-700.
doi: 10.1136/gut.2003.027789.

Is the mucosal route of administration essential for probiotic function? Subcutaneous administration is associated with attenuation of murine colitis and arthritis

B Sheil  1 , J McCarthy, L O'Mahony, M W Bennett, P Ryan, J J Fitzgibbon, B Kiely, J K Collins, F Shanahan
Affiliations

Is the mucosal route of administration essential for probiotic function? Subcutaneous administration is associated with attenuation of murine colitis and arthritis

B Sheil et al. Gut. 2004 May.

Erratum in

  • Gut. 2004 Aug;53(8):1216

Abstract

Background: We and others have reported the prophylactic efficacy of oral consumption of probiotic lactobacilli in the interleukin 10 knockout (IL-10 KO) model of colitis. It has not been demonstrated that the oral route is essential for probiotic efficacy.

Aims: (i) To determine the effect of parenteral administration (subcutaneous) of Lactobacillus salivarius 118 on colitis of IL-10 KO mice; and (ii) to determine if observed responses are disease specific.

Methods: (i) IL-10 KO mice were injected subcutaneously with L salivarius 118 or saline over 19 weeks. At sacrifice, the bowels were histologically scored. Isolated splenocytes were cultured in vitro and cytokine levels measured. (ii) In the collagen induced arthritis model, DBA/1 mice were injected subcutaneously with the probiotic or saline. At sacrifice, paw thickness was measured and joints were histologically scored.

Results: (i) Colonic inflammatory scores were significantly decreased in IL-10 KO mice injected with L salivarius 118 compared with controls (p<0.05). Proinflammatory cytokine production from stimulated splenocytes was significantly lower for the probiotic group whereas stimulated transforming growth factor beta (TGF-beta) levels were significantly increased (p<0.05). (ii) Scoring of arthritis and paw thickness were significantly improved in the group of mice injected with L salivarius 118 compared with controls.

Conclusions: (1) Subcutaneous administration of L salivarius 118 significantly attenuated colitis in the IL-10 KO model and suppressed collagen induced arthritis, suggesting that the oral route may not be essential for probiotic anti-inflammatory effects and that responses are not disease specific. (2) The probiotic effect was associated with reduced production of proinflammatory (T helper 1) cytokines and maintained production of anti-TGF-beta.

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Figures

Figure 1
Figure 1
Post sacrifice, a histological score of 0–4 was awarded to each examined area of bowel; caecum, proximal colon (ascending and transverse colon), and distal colon (descending colon, rectum and anal canal) with a total possible score of 12 to be awarded to each mouse gut. Gastrointestinal inflammatory score in the probiotic and control fed groups is shown. *p<0.05, n = 10 per group. Results are expressed as mean (SEM) inflammatory score.
Figure 2
Figure 2
Cytokine production by stimulated splenocytes in control and probiotic interleukin 10 knockout mice. Results are expressed as mean (SEM) cytokine level per group. There was a reduction in the proinflammatory cytokines tumour necrosis factor α (TNF) (A) and interleukin 12 p40 (IL-12) (B), with an increase in the anti-inflammatory cytokine transforming growth factor β (TGF) (C). *p<0.05; n = 10 per group.
Figure 3
Figure 3
Comparison of daily clinical scores of mice subcutaneously inoculated with L salivarius 118 and phosphate buffered saline (PBS) following induction of arthritis. Mice were examined daily from week 10 (day 1) to week 12 (day 16) for visual signs of arthritis using an established scoring system on a scale of 0–4. The macroscopic score (mean (SEM)) was expressed as a cumulative value for all paws, with a maximum possible score of 16 (n = 10 per group, except unaffected negative control group, n = 6). The L salivarius 118 group had a significantly reduced daily clinical score compared with the PBS control on days 3–day 9 (p = 0.01), on days 10–12, and on day 15 (p = 0.05).
Figure 4
Figure 4
Effect of subcutaneous inoculation of L salivarius 118 and phosphate buffered saline (PBS) on paw thickness (A), total inflammation in all limbs (B), inflammation in the fore limbs (C), and inflammation in the hind limbs (D), in collagen induced arthritis. The thickness of each paw was measured with spring loaded callipers. Total paw swelling for each mouse was calculated by adding the thickness of the individual paws. Paw thickness of the unaffected/negative control group was taken as 0. Results (A–D) are expressed as mean (SEM). The L salivarius 118 group showed a significant reduction in paw thickness compared with controls (n = 10 per group).
Figure 5
Figure 5
Joint histology in collagen induced arthritis. Following sacrifice, histopathological changes were scored using a scale of 0–3, based on cartilage and bone destruction by pannus formation. Infiltration of inflammatory cells was also scored on a scale of 0–3, ranging from no cell infiltration to severe infiltration. The range of the histological grading system used in the control and L salivarius groups is illustrated. (A) Example of a normal joint with no signs of arthritis. (B) A joint with mild inflammation and synovial hyperplasia. (C) A joint with moderate inflammation and pannus invasion of cartilage. (D) A joint with severe inflammation and invasion of pannus to underlying bone.
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