Triclosan (TCS) is an antimicrobial used widely in hospitals and personal care products, at ~10 mM. Human skin efficiently absorbs TCS. Mast cells are ubiquitous key players both in physiological processes and in disease, including asthma, cancer, and autism. We previously showed that non-cytotoxic levels of TCS inhibit degranulation, the release of histamine and other mediators, from rat mast cells (RBL-2H3), and in this study we replicate this finding in human (HMC-1.2) mast cells. Our investigation into the molecular mechanisms underlying this effect led to the discovery that TCS disrupts ATP production in RBL-2H3 cells in glucose-free, galactose-containing media (95% CI EC50 = 7.5-9.7 μM), without causing cytotoxicity. Using these same glucose-free conditions, 15 μM TCS dampens RBL-2H3 degranulation by 40%. The same ATP disruption was found with human HMC-1.2 cells (EC50 4.2-13.7 μM), NIH-3T3 mouse fibroblasts (EC50 4.8-7.4 μM), and primary human keratinocytes (EC50 3.0-4.1 μM) all with no cytotoxicity. TCS increases oxygen consumption rate in RBL-2H3 cells. Known mitochondrial uncouplers (e.g., CCCP) previously were found to inhibit mast cell function. TCS-methyl, which has a methyl group in place of TCS’s ionizable proton, affects neither degranulation nor ATP production at non-cytotoxic doses. Thus, triclosan’s effects on mast cell function are due to its proton ionophore structure. Also, 5 μM TCS inhibits thapsigargin-stimulated degranulation of RBL-2H3 cells: further evidence that TCS disrupts mast cell signaling. Our data indicate that TCS is a mitochondrial uncoupler, and TCS may affect numerous cell types and functions via this mechanism.

Exposure to arsenic (As) is a global health concern. We previously documented an inhibitory effect of inorganic arsenite on IgE- mediated degranulation of RBL-2H3 mast cells (Hutchinson et al., 2011). Mast cells are tissue-resident cells that are positioned at the host-environment interface, thereby serving vital roles in many physiological processes and disease states, in addition to their well-known roles in allergy and asthma. Upon activation, mast cells secrete several mediators from cytoplasmic granules, in degranulation. The present study is an investigation of arsenite's molecular target(s) in the degranulation pathway. Here, we report that As does not affect degranulation stimulated by either the Ca2+ionophore A23187 or thapsigargin, which both bypass early signaling events. As also does not alter degranulation initiated by another non-IgE-mediated mast cell stimulant, the G-protein activator compound 48/80. However, As inhibits Ca2+ influx into antigen-activated mast cells. These results indicate that arsenic's target in the degranulation pathway is upstream of Ca2+ influx. Phospho-Syk ELISA and phospho-p85 phosphoinositide 3-kinase ELISA data show that arsenic inhibits early phosphorylation events. Taken together, this evidence indicates that the mechanism underlying arsenic's inhibition of mast cell degranulation occurs at the early tyrosine phosphorylation steps in the degranulation pathway.

Mast cells comprise a physiologically and toxicologically important cell type that is ubiquitous among species and tissues. Mast cells undergo degranulation, in which characteristic intracellular granules fuse with the plasma membrane and release many bioactive substances, such as enzymes β‐hexosaminidase and tryptase. Activity of mast cells in the toxicology model organism, zebrafish, has been monitored via tryptase release and cleavage of substrate N‐α‐benzoyl‐dl‐Arg‐p‐nitroanilide (BAPNA). An extensively used in vitro mast cell model for studying toxicant mechanisms is the RBL‐2H3 cell line. However, instead of tryptase, granule contents such as β‐hexosaminidase have usually been employed as RBL‐2H3 degranulation markers. To align RBL‐2H3 cell toxicological studies to in vivo mast cell studies using zebrafish, we aimed to develop an RBL‐2H3 tryptase assay. Unexpectedly, we discovered that tryptase release from RBL‐2H3 cells is not detectable, using BAPNA substrate, despite optimized assay that can detect as little as 1 ng tryptase. Additional studies performed with another substrate, tosyl‐Gly‐Pro‐Lys‐pNA, and with an enzyme‐linked immunosorbent assay, revealed a lack of tryptase protein released from stimulated RBL‐2H3 cells. Furthermore, none of the eight rat tryptase genes (Tpsb2, Tpsab1, Tpsg1, Prss34, Gzmk, Gzma, Prss29, Prss41) is expressed in RBL‐2H3 cells, even though all are found in RBL‐2H3 genomic DNA and even though β‐hexosaminidase mRNA is constitutively expressed. Therefore, mast cell researchers should utilize β‐hexosaminidase or another reliable marker for RBL‐2H3 degranulation studies, not tryptase. Comparative toxicity testing in RBL‐2H3 cells in vitro and in zebrafish mast cells in vivo will require use of a degranulation reporter different from tryptase.

The present study was conducted to quantify the effects of soil pollutants on soil inhabiting root-associated fungi. Roots of Parthenium hysterophorus (L.) subjected to fungal isolation revealed more diversity in fungal association compared to Desmostachya bipinnata (L.). Aspergillus was found most commonly associated fungi with the roots of both the plants. A total of seven different fungal species were isolated from the two plants. The root extracts of both the plants were found to have antibacterial agents. However, gram negative strains of bacteria were observed least sensitive. It seems that either plants or fungi, on polluted sites have co-adapted themselves in order to survive. Their interdependence is a manifestation of an association that cannot be strictly regarded as symbiotic but leads to an endorsing reciprocity.

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