Evaluation of the in vivo and in vitro interleukin-12 p40 and p35 subunit response in yellowtail (Seriola quinqueradiata) to heat-killed Lactobacillus plantarum strain L-137 (HK L-137) supplementation, and immersion challenge with Lactococcus garvieae

Dietary supplementation of immunostimulants might be effective to reduce the economic losses due to infectious diseases and the use of antibiotics in aquaculture. To investigate the immune response of interleukin-12 (IL-12) in yellowtail Seriola quinqueradiata to heat-killed Lactobacillus plantarum strain L-137 (HK L-137), we performed a leukocyte culture, feeding trial with diets containing L-137 and an immersion challenge with Lactococcus garvieae. IL-12 (IL-12p70) is a heterodimeric cytokine composed of IL-12p35 and IL-12p40 subunits. In the yellowtail-leukocyte culture, HK L-137 treatment stimulated the mRNA expression of one IL12p35 subunit (p35a) and all IL12p40 subunits (p40a, p40b, and p40c) in a dose-dependent manner. Furthermore, mRNA expression of type-I helper (Th-1) cytokine (tumor necrosis factor α, TNF-α, and interferon γ, IFN-γ) was also stimulated by HK L-137. After 6 weeks of feeding yellowtails with diets containing 0, 20, and 100 ppm of HK L-137, the mRNA expression of p35a and p40b in the spleen leukocytes increased with the dietary concentration of HK L-137, and that of p40b, p40c, and ifng in the head kidney leukocytes were the highest in the 20 ppm HK L-137 group. Survival rates in the 20 ppm HK L-137 group after immersion challenge with L. garvieae were significantly higher than the control (0 ppm of HK L-137). The 100 ppm HK L-137 group did not significantly suppress mortality. HK L-137 showed immunostimulant activity by increasing the expression of il-12, tnfa, and ifng mRNA in both in vitro and in vivo tests in yellowtail. Our results suggest that dietary supplementation with 20 ppm HK L-137 is the most efficient dose for improving immunity in yellowtail. Furthermore, a high dose of HK L-137 and/or long-term feeding of a diet containing HK L-137 might suppress the immune response, which probably decreases the survival rate of fish. To maintain a high immune response in yellowtail, the optimal dietary concentration of HK L-137 and/or feeding regime should be investigated further

Surface distribution and depths profiling of particulate organic UV absorbers by Raman imaging and tape stripping

Tape stripping in conjunction with scanning Raman microscopy was used for assessing the lateral and vertical distribution of an organic particulate UV filter, methylene bis-benzotriazolyl tetramethylbutylphenol (MBBT), in a sunscreen formulation. On the volar forearms of three volunteers, 1 mg cm(-2) formulation containing 10% MBBT was applied, and the average amount of MBBT was measured by Raman scanning microscopy in 15 consecutive tape strippings. The recovery of MBBT was 91.1% with 30.2% localizing on the skin surface (first strip), 42.5% in the upper stratum corneum (strips 2-5) and from 3.6 down to 0.8% in each of the 10 consecutive layers. The concentration of surface deposits of MBBT differed by a factor of 300 between folds, furrows and pores on the one hand and the interjacent ridges on the other hand. Seventy-five per cent of the applied particles occupied a fifth of the evaluated area - where concentrating in folds and furrows - as was confirmed by 3-D reconstruction. On interjacent ridges, 8.6% of MBBT distributed as very thin films preferentially. MBBT localized at sites not connected with the surface, such as in truncated pores or as potentially penetrated material amounted to 0.06% or to a twentieth of the 1.4% found in the lowest skin strippings. Scanning Raman microscopy in combination with tape stripping documented the lateral and vertical distribution quantitatively and at cellular (12.5 ?m) lateral resolution. Our results confirmed an earlier report on the vertical distribution of organic particles applied to skin and was in line with similar reports on TiO(2) distribution