With clinical and in vitro data supporting the therapeutic potential of probiotics and their secreted products, the direction of my current and future research activities center around demonstrating the anti-inflammatory properties of the secreted products using in vitro models as well as characterizing these products with various laboratory techniques. Equally as important is the investigation of whether single or combination probiotics offer optimal anti-inflammatory effects. Since B.infantis and L.acidophilus were included in three clinical trials showing a statistically significant reduction in the incidence of NEC, these two probiotics had been chosen for the initial stages of my research. To date, we have studied the inflammatory response of mature and immature human intestinal epithelium, quantified by cellular IL-6 secretion with ELISA assays, and the attenuation of this response with concomitant exposure to probiotic-conditioned cell culture medium.
The initial data show that cellular IL-6 secretion by immature, human enterocyte cell lines after TNF-α or IL1-β stimulation is decreased after concomitant exposure to probiotic-conditioned media compared to the positive control. The decrease in cellular IL-6 secretion is greatest with B.infantis-conditioned media in fetal IEC and H4 cells and L.acidophilus-conditioned media in NEC-IEC cells; the data does not support the idea that a combined probiotic preparation of B.infantis and L.acidophilus provides an optimal anti-inflammatory effect.
The observed changes in IL-6 secretion were similar when boiled fractions of the secreted products were used, suggesting that the bioactive effecters are heat-stable. For further characterization, the bioactive products responsible for IL-6 secretion attenuation were size-fractionated using a centrifugal filtering system. Data from our size-fractionated studies suggest that there may be more than one bioactive secreted effector responsible for the anti-inflammatory effect of these particular probiotic strains.
They also suggest that the effector(s) secreted by B.infantis may be very different than that secreted by L.acidophilus. The nature of these differences is unclear, and therefore the mechanism by which inflammatory attenuation occurs is being investigated for both bacteria. As preliminary data show an exaggerated inflammatory response in immature enterocytes, qRT-PCR analysis is being used to determine whether developmentally regulated gene expression contributes to this finding and will characterize the influence probiotic-secreted products have on enterocyte gene expression. The intestinal inflammatory attenuation as well as the mechanism of action determined by our in vitro models will be confirmed using various in vivo NEC models.