Meptyldinocap induces implantation failure by forcing cell cycle arrest, mitochondrial dysfunction, and endoplasmic reticulum stress in porcine trophectoderm and endometrial luminal epithelial cells.

Meptyldinocap is a dinitrophenol fungicide used to control powdery mildew. Although other dinitrophenol pesticides have been found to exhibit reproductive toxicity, studies of meptyldinocaps are scarce. This study investigated the adverse effects of meptyldinocap on porcine trophectoderm (pTr) and porcine endometrial luminal epithelial (pLE) cells, which play crucial roles in implantation. We confirmed that meptyldinocap decreased cell viability, induced apoptosis, and inhibited proliferation by decreasing proliferation-related gene expression and inducing changes in the cell cycle. Furthermore, meptyldinocap treatment caused mitochondrial dysfunction, endoplasmic reticulum stress, and disruption of calcium homeostasis. Moreover, it induces alterations in mitogen-activated protein kinase signaling cascades and reduces the migration ability, leading to implantation failure. Our findings suggest that meptyldinocap reduces the cellular functions of pTr and pLE cells, which are important for the implantation process, and interferes with interactions between the two cell lines, potentially leading to implantation failure. We also propose a mechanism by which the understudied fungicide meptyldinocap exerts its cytotoxicity.

Divergent Roles of Escherichia Coli Encoded Lon Protease in Imparting Resistance to Uncouplers of Oxidative Phosphorylation: Roles of marA, rob, soxS and acrB.

Uncouplers of oxidative phosphorylation dissipate the proton gradient, causing lower ATP production. Bacteria encounter several non-classical uncouplers in the environment, leading to stress-induced adaptations. Here, we addressed the molecular mechanisms responsible for the effects of uncouplers in Escherichia coli. The expression and functions of genes involved in phenotypic antibiotic resistance were studied using three compounds: two strong uncouplers, i.e., Carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and 2,4-Dinitrophenol (DNP), and one moderate uncoupler, i.e., Sodium salicylate (NaSal). Quantitative expression studies demonstrated induction of transcripts encoding marA, soxS and acrB with NaSal and DNP, but not CCCP. Since MarA and SoxS are degraded by the Lon protease, we investigated the roles of Lon using a lon-deficient strain (Δlon). Compared to the wild-type strain, Δlon shows compromised growth upon exposure to NaSal or 2, 4-DNP. This sensitivity is dependent on marA but not rob and soxS. On the other hand, the Δlon strain shows enhanced growth in the presence of CCCP, which is dependent on acrB. Interestingly, NaSal and 2,4-DNP, but not CCCP, induce resistance to antibiotics, such as ciprofloxacin and tetracycline. This study addresses the effects of uncouplers and the roles of genes involved during bacterial growth and phenotypic antibiotic resistance. Strong uncouplers are often used to treat wastewater, and these results shed light on the possible mechanisms by which bacteria respond to uncouplers. Also, the rampant usage of some uncouplers to treat wastewater may lead to the development of antibiotic resistance.

[Clinical case report of acute dinitrophenol poisoning with a fatal outcome of Udmurt Republic]. / Klinicheskii sluchai ostrogo otravleniya dinitrofenolom s letal'nym iskhodom v Udmurtskoi Respublike.

THE AIM OF THE STUDY: Was to conduct the analysis of patient's clinical observation with acute dinitrophenol poisoning, admitted to a toxicological department of CCH №6 of Izhevsk, Udmurt Republic in 2021 yr. In this clinical case report, a 19 years old girl, who took 20 tablets of dinitrophenol, illegally obtained in online-shop, died. The fatal outcome was realized by the uncoupling of oxidative phosphorylation mechanism and cellular respiration, which in its turn led to serious dystrophic changes in all organs and tissues. Disorders of hemodynamics and blood rheological properties dominated in poisoning pathogenesis, led to congestion, stasis in microcirculatory vessels, hyperpermeability with multiple perivascular hemorrhages in organs, occurrence of piecemeal necrosis in kidneys and liver, nephrosis and nonspecific reactive hepatitis. Production ATP from ADP becomes impossible in these conditions, and respiratory energy chain completely disappears as heat, that explains the heat-increasing and fat-burning effects of dinitrophenol.

Sublethal toxicities of 2,4-dinitrophenol as inferred from online self-reports.

INTRODUCTION: 2,4-dinitrophenol (DNP) is a mitochondrial toxin sometimes used as a weight loss agent. Reports of fatalities from DNP have been increasing since 2000, suggesting an increase in use. Our understanding of DNP toxicity in humans comes from reports to Poison Control and postmortem analyses, sources that are biased to more extreme presentations. This leads to a gap in our knowledge about the adverse effects of DNP at nonlethal doses. Here we investigate the doses and effects of DNP as reported online. METHODS: We analyzed publicly available Internet posts that we collected from 2017-2019. The posts came from anonymous users or users who voluntarily self-identified. We collected data from websites whose terms of use allow for the secondary analysis of data that their users agree to make public. We used natural language processing techniques that we had previously developed to extract doses, effects, and substances mentioned in each post. RESULTS: We collected 1,630 posts across 5 online forums and the Reddit forum r/DNP. The posts were from 1,234 unique usernames. The most commonly reported doses were between 150 to 300 mg each day followed by 300 to 450 mg each day. At those doses, the most reported adverse effects were profuse sweating and fatigue. Reports of thermoregulatory (sweating, feeling hot flashes or flushed), fatigue-related, and neurologically related symptoms were statistically significantly more frequent at reported daily doses greater than 150 mg than doses below 150 mg (post-hoc χ2-test with Bonferroni correction). The effects were judged as plausible by two board-certified medical toxicologists. Triiodothyronine, clenbuterol, testosterone, and trenbolone, an androgenic anabolic steroid were the most significantly co-mentioned substances. CONCLUSIONS: Fatigue, increased body temperature, and paresthesias from DNP are reported more frequently at doses greater than 150 mg each day than at doses less than 150 mg each day. Online discussions of DNP frequently mention androgenic anabolic steroids and other weight loss agents.

2,4-Dinitrophenol does not exert neuro-regenerative potential in experimental autoimmune neuritis.

OBJECTIVE: We evaluated the potential neuro-regenerative effects of the mitochondrial uncoupler 2,4-Dinitrophenol in experimental autoimmune neuritis, an animal model for an acute autoimmune neuropathy. METHODS: Experimental autoimmune neuritis was induced in Lewis rats. Different concentrations of 2,4-Dinitrophenol (1 mg/kg, 0.1 mg/kg and 0.01 mg/kg) were applied during the recovery phase of the neuritis (at days 18, 22 and 26) and compared to the vehicle. Any effects were assessed through functional, electrophysiological, and morphological analysis via electron microscopy of all groups at day 30. Additional immune-histochemical analysis of inflammation markers and remyelination of the sciatic nerves were performed for the dosage of 1 mg/kg and control. RESULTS: No enhancement of functional or electrophysiological recovery was observed in all 2,4-Dinitrophenol-treated groups. Cellular inflammation markers of T cells (CD3+) were comparable to control, and an increase of macrophages (IbA1+) invasion in the sciatic nerves was observed. Treatment with 2,4-Dinitrophenol reduced axonal swelling in myelinated and unmyelinated fibers with an increased production of brain-derived neurotrophic factor. CONCLUSION: Our findings do not support the hypothesis that repurposing of the mitochondrial uncoupler 2,4-Dinitrophenol exerts functionally relevant neuro-regenerative effects in autoimmune neuritis.

CKBA suppresses mast cell activation via ERK signaling pathway in murine atopic dermatitis.

Atopic dermatitis (AD) is a common inflammatory skin disorder. Mast cells play an important role in AD because they regulate allergic reactions and inflammatory responses. However, whether and how the modulation of mast cell activity affects AD has not been determined. In this study, we aimed to determine the effects and mechanisms of 3-O-cyclohexanecarbonyl-11-keto-ß-boswellic acid (CKBA). This natural compound derivative alleviates skin inflammation by inhibiting mast cell activation and maintaining skin barrier homeostasis in AD. CKBA markedly reduced serum IgE levels and alleviated skin inflammation in calcipotriol (MC903)-induced AD mouse model. CKBA also restrained mast cell degranulation both in vitro and in vivo. RNA-seq analysis revealed that CKBA downregulated the extracellular signal-regulated kinase (ERK) signaling in BM-derived mast cells activated by anti-2,4-dinitrophenol/2,4-dinitrophenol-human serum albumin. We proved that CKBA suppressed mast cell activation via ERK signaling using the ERK activator (t-butyl hydroquinone) and inhibitor (selumetinib; AZD6244) in AD. Thus, CKBA suppressed mast cell activation in AD via the ERK signaling pathway and could be a therapeutic candidate drug for AD.

Non-Cardiotoxic Tetradecanoic Acid-2,4-Dinitrophenol Ester Nanomicelles in Microneedles Exert Potent Anti-Obesity Effect by Regulating Adipocyte Browning and Lipogenesis.

Sustained oral uncoupler 2,4-dinitrophenol (DNP) administration exerts prominent anti-obesity effects, but the adipose tissue off-target disadvantage leads to systemic adverse effects. A novel non-cardiotoxicity DNP delivery method using a biocompatible microneedles patch containing the amphiphilic tetradecanoic acid-DNP ester (TADNP) is described, which is synthesized via esterification on the phenolic hydroxyl of DNP. The TADNP is self-assembled as nanomicelles, which enhance the endocytosis rate of DNP by adipocytes and its permeation in isolated adipose tissues. The microenvironment of adipose tissues promotes the massive release of DNP and plasma and simulated gastrointestinal fluids. The microneedles-delivered TADNP nanomicelles (MN-TADNP) effectively deliver DNP in treated adipose tissues and reduce DNP content in off-target organs. Both oral and MN patch-delivered TADNP micelles effectively exert anti-obesity effects in a mouse model of high-fat diet-induced obesity; and noteworthily, MN-TADNP exhibit more satisfactory biosafety than oral administration. Here, a smart MN patch loaded with tetradecanoic acid-modified DNP is reported, which enhances its accumulation in adipose tissues and exerts an anti-obesity effect without causing any systemic toxicity.

Phytoremediation of dinitrophenol from wastewater by atriplex lentiformis: effect of salicylic acid.

Quail bush [Atriplex lentiformis (Torr.) S. Wats] plants were used in removing 2, 4-dinitrophenol (DNP) from wastewater in a hydroponic experiment. The hydroponic system contained three doses of DNP, i.e., 0, 10, and 20 mg L-1. Quail bush plants were sprayed with 0.1 mM salicylic acid (SA) to study its role in resisting DNP toxicity. DNP significantly (p < 0.05) reduced plant growth. Exposure of A. lentiformis plants to 20 mg L-1 of DNP reduced the total chlorophyl and relative water content by 39 and 24%, respectively. SA improved the antioxidant defense in terms of ascorbate peroxidase (APX) and polyphenol oxidase (PPO) activities. SA alleviated DNP toxicity by enhancing the production of osmoprotectants, e.g.,proline, phenols, and carbohydrates. SA enhanced the removal efficiency of DNP and the highest removal efficiency (96%) was recorded in the plants sprayed with SA and grown on 10 mg L-1 of DNP. A. lentiformis is a halophytic plant that has good physiological characteristics to resist 2, 4-dinitrophenol toxicity in wastewaters and is qualified to purify water from these harmful compounds. Exogenous application of 0.1 mM SA increased the defense system in A. lentiformis against 2, 4-dinitrophenol toxicity and enhanced the removal efficiency.

2, 4-dinitrophenol inhibited the synthesis of photosynthetic pigments.Salicylic acid protects the vital bio-compounds in plant cells.Atriplex plants are able to remove (96%) of 2, 4-dinitrophenol from the wastewater.Atriplex plants have a strong antioxidant defense enable them to survive in wastewater.

In-situ synthesis of novel dual S-scheme AgI/Ag6Mo7O24/g-C3N4 heterojunctions with tandem structure for photocatalytic degradation of organic pollutants.

The controllable design of multivariate heterojunction with sequential structures is of significant relevance for breaking the performance limit of binary composite photocatalysts. In this work, the novel dual S-scheme ternary-component AgI/Ag6Mo7O24/exfoliated g-C3N4 (ECN) composite was prepared by a two-step in-situ synthetic strategy. The energy band bending at the heterointerface and the formation of dual built-in electric field could be observed due to distinct work functions of different components in the ternary composite. Benefiting from the sequential heterojunction structure, the AgI/Ag6Mo7O24/ECN composite achieved 98.7% removal efficiency of 2-nitrophenol (2-NP) within 70 min under visible light irradiation, and AgI/Ag6Mo7O24/ECN also showed higher degradation efficiency for a variety of organic pollutants such as methylene blue (MB), rhodamine B (RhB), methyl orange (MO), 4-nitrophenol (4-NP), 2-sec-butyl-4,6-dinitrophenol (DNBP) and tetracycline (TC). Notably, •OH and •O2- played dominant roles in the AgI/Ag6Mo7O24/ECN set up, which was consistent with the dual S-scheme charge transfer mechanism. In-depth insights for the photodegradation of 2-NP were presented based on a combined DFT study and GC-MS analysis. Additionally, the photoreduction of Ag+ in AgI/Ag6Mo7O24/ECN was also evaded by the fast transfer of photogenerated electrons through the dual S-scheme pathway, achieving the effect of killing two birds with one stone.

Efficient Detection of 2,6-Dinitrophenol with Silver Nanoparticle-Decorated Chitosan/SrSnO3 Nanocomposites by Differential Pulse Voltammetry.

Herein, an ultra-sonication technique followed by a photoreduction technique was implemented to prepare silver nanoparticle-decorated Chitosan/SrSnO3 nanocomposites (Ag-decorated Chitosan/SrSnO3 NCs), and they were successively used as electron-sensing substrates coated on a glassy carbon electrode (GCE) for the development of a 2,6-dinitrophenol (2,6-DNP) efficient electrochemical sensor. The synthesized NCs were characterized in terms of morphology, surface composition, and optical properties using FESEM, TEM, HRTEM, BET, XRD, XPS, FTIR, and UV-vis analysis. Ag-decorated Chitosan/SrSnO3 NC/GCE fabricated with the conducting binder (PEDOT:PSS) was found to analyze 2,6-DNP in a wide detection range (LDR) of 1.5~13.5 µM by applying the differential pulse voltammetry (DPV) approach. The 2,6-DNP sensor parameters, such as sensitivity (54.032 µA µM-1 cm-2), limit of detection (LOD; 0.18 ± 0.01 µM), limit of quantification (LOQ; 0.545 µM) reproducibility, and response time, were found excellent and good results. Additionally, various environmental samples were analyzed and obtained reliable analytical results. Thus, it is the simplest way to develop a sensor probe with newly developed nanocomposite materials for analyzing the carcinogenic contaminants from the environmental effluents by electrochemical approach for the safety of environmental and healthcare fields in a broad scale.

2,4-Dinitrophenol as an Uncoupler Augments the Anthracyclines Toxicity against Prostate Cancer Cells.

One of the strategies for the treatment of advanced cancer diseases is targeting the energy metabolism of the cancer cells. The compound 2,4-DNP (2,4-dinitrophenol) disrupts the cell energy metabolism through the ability to decouple oxidative phosphorylation. The aim of the study was to determine the ability of 2,4-DNP to sensitize prostate cancer cells with different metabolic phenotypes to the action of known anthracyclines (doxorubicin and epirubicin). The synergistic effect of the anthracyclines and 2,4-DNP was determined using an MTT assay, apoptosis detection and a cell cycle analysis. The present of oxidative stress in cancer cells was assessed by CellROX, the level of cellular thiols and DNA oxidative damage. The study revealed that the incubation of LNCaP prostate cancer cells (oxidative phenotype) with epirubicin and doxorubicin simultaneously with 2,4-DNP showed the presence of a synergistic effect for both the cytostatics. Moreover, it contributes to the increased induction of oxidative stress, which results in a reduced level of cellular thiols and an increased number of AP sites in the DNA. The synergistic activity may consist of an inhibition of ATP synthesis and the simultaneous production of toxic amounts of ROS, destroying the mitochondria. Additionally, the sensitivity of the LNCaP cell line to the anthracyclines is relatively higher compared to the other two (PC-3, DU-145).

Halide salts induced the photodegradation of a fat-burning compound 2, 4-dinitrophenol by iron-montmorillonite.

Natural montmorillonite clay and anthropogenic organic pollutants frequently coexist in the estuarine environment where freshwater from rivers mixes with saltwater from the ocean. In this environment, the sharply changed aqueous chemistry especially salt content could significantly alter the photochemical behaviors of pollutants. However, this process was rarely investigated. In this study, the photodegradation of a representative anthropogenic weight-loss compound 2,4-dinitrophenol in the presence of Fe3+-montmorillonite and different halide salts was systematically investigated. Results show that 2,4-dinitrophenol was resistant to photodegradation by Fe3+-montmorillonite alone, but the presence of NaCl, NaBr, and sea salts in the system can evoke significant 2,4-dinitrophenol degradation. The enhancement effect was further elucidated as the replacement reaction between the clay associated Fe3+ and Na + which leads to the release of more interlayer Fe3+ from montmorillonite, resulting in increased production of high active hydroxyl radicals (˙OH) that can substantially damage 2,4-dinitrophenol molecule. In addition, halogen radicals from the reaction of halide ions with ˙OH were also confirmed to participate in 2,4-dinitrophenol degradation. Overall, this study implied that the changed salty condition in the estuarine water could induce the rapid transformation of organic pollutants that move from freshwater and have relatively stable photochemical properties.

The photosynthetic physiological response and purification effect of Salix babylonica to 2, 4-dinitrophenol wastewater.

Phytoremediation technology based on living green plants would clean up water pollution. Through hydroponic experiment, the effects of different concentration of 2, 4-dinitrophenol (2, 4-DNP) on the photosynthetic and chlorophyll fluorescence parameters of Salix babylonica, and the absorption and purification effect of S. babylonica on 2, 4-DNP were measured to explore the tolerance of S. babylonica to 2, 4-DNP and the feasibility to purify dinitrophenol waste water by it. The biomass, actual photochemical efficiency (PSII), net photosynthetic rate (Pn), photochemical quenching coefficient (qP), stomatal conductance (Gs), transpiration rate (Tr), maximum photochemical efficiency (Fv/Fm) and chlorophyll content of the S. babylonica showed downward trend with the increasing exposure concentrations of 2,4-DNP, but the intercellular CO2 concentration (Ci) appeared upward trend. Non-photochemical quenching coefficient (NPQ) increased at 5 mg L-1, then declined with the increase concentrations of 2, 4-DNP. In addition, the percent removal of 2, 4-DNP in 20 mg L-1 waste water was 91.4%. In conclusion, 2, 4-DNP significantly inhibits Pn of S. babylonica and the reduction of Pn was caused by decreasing Gs, carboxylation efficiency and chlorophyll content. When the concentration of 2, 4-DNP is not more than 20 mg L-1, S. babylonica can remove 2, 4-DNP efficiently.

Interaction with 2,4-dinitrophenol correlates with polyreactivity, self-binding, and stability of clinical-stage therapeutic antibodies.

Therapeutic antibodies should cover particular physicochemical and functional requirements for successful entry into clinical practice. Numerous experimental and computational approaches have been developed for early identification of different unfavourable features of antibodies. Immune repertoires of healthy humans contain a fraction of antibodies that recognize nitroarenes. These antibodies have been demonstrated to manifest antigen-binding polyreactivity. Here we observed that >20 % of 112 clinical stage therapeutic antibodies show pronounced binding to 2,4-dinitrophenol conjugated to albumin. This interaction predicts a number of unfavourable functional and physicochemical features of antibodies such as polyreactivity, tendency for self-association, stability and expression yields. Based on these findings we proposed a simple approach that may add to the armamentarium of assays for early identification of developability liabilities of antibodies intended for therapeutic use.

Bifunctional small molecules that mediate the degradation of extracellular proteins.

Targeted protein degradation (TPD) has emerged as a promising therapeutic strategy. Most TPD technologies use the ubiquitin-proteasome system, and are therefore limited to targeting intracellular proteins. To address this limitation, we developed a class of modular, bifunctional synthetic molecules called MoDE-As (molecular degraders of extracellular proteins through the asialoglycoprotein receptor (ASGPR)), which mediate the degradation of extracellular proteins. MoDE-A molecules mediate the formation of a ternary complex between a target protein and ASGPR on hepatocytes. The target protein is then endocytosed and degraded by lysosomal proteases. We demonstrated the modularity of the MoDE-A technology by synthesizing molecules that induce depletion of both antibody and proinflammatory cytokine proteins. These data show experimental evidence that nonproteinogenic, synthetic molecules can enable TPD of extracellular proteins in vitro and in vivo. We believe that TPD mediated by the MoDE-A technology will have widespread applications for disease treatment.

Dinitrophenol-Hyaluronan Conjugates as Multivalent Antibody-Recruiting Glycopolymers for Targeted Cancer Immunotherapy.

Multivalent antibody-recruiting glycopolymers (MARGs) composed of hyaluronic acid (HA) grafted with multiple copies of dinitrophenol (DNP) were developed for targeted cancer immunotherapy. Structure-activity studies demonstrated that the MARGs were able to specifically recognize CD44-positive cancer cells and displayed remarkable antibody-recruiting capacities and tumor cell killing activities dependent on the introduced multivalent effect and the length of PEG linker. One of the MARGs, HA-[PEG3 -DNP]8 , showed the best capacity for clustering anti-DNP antibodies onto CD44-positive cancer cells and displayed potent in vitro anti-cancer activity by triggering complement dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC). Moreover, we found that HA-[PEG3 -DNP]8 significantly inhibited the xenograft tumor growth of Babl/c nude mice bearing triple negative breast cancer cells, while it did not cause detectable histological cytotoxicity. Given the easy access of this type of natural glycopolymer and the practical synthesis approach, these MARGs provide promising immunotherapeutics for cancer immunotherapy.

Dinitrophenol-mediated modulation of an anti-PD-L1 VHH for Fc-dependent effector functions and prolonged serum half-life.

Single-domain antibodies, VHHs or nanobodies, represent a promising set of alternatives to conventional therapeutic antibodies, gaining substantial attention in the field of cancer immunotherapy. However, inherent drawbacks of nanobodies such as fast clearance from blood circulation and lack of immune effector functions often led to unsatisfactory therapeutic efficacy. We previously reported that dinitrophenyl modification of an anti-EGFR VHH conferred Fc-dependent immune effector functions and elongated serum half-life on it through recruiting of hapten antibodies, resulting in improved immunotherapy efficacy in vivo. In the present work, we further tested the versatility of this approach in the case of an anti-PD-L1 blockade VHH (KN035). Site-specific dinitrophenyl conjugation did not impair the binding capacity of KN035 portion to PD-L1, but indirectly restored its immune effector functions, manifested by the observed antibody dependent cell-mediated cytotoxicity, antibody-dependent cellular phagocytosis and complement-dependent cytotoxicity against PD-L1 positive tumor cells. Significant delay of blood clearance of dinitrophenylated KN035 was evidenced by the prolonged half-life of ca. 22 h. This approach, using small hapten molecule conjugation, loaded additional antibody-mediated tumor killing mechanisms to PD-L1 blockade VHH and therefore improved efficacy is anticipated in the future in vivo therapeutic studies. Thus, our results underscore the power of this versatile approach for achieving desirable properties of VHH-based or similar therapeutics.

Sargassum horneri as a Functional Food Ameliorated IgE/BSA-Induced Mast Cell Activation and Passive Cutaneous Anaphylaxis in Mice.

Sargassum horneri (S. horneri), an edible brown alga, has been proposed as a functional food with an improvement effect on abnormal skin immune responses. The present study investigates the anti-allergic effect of an ethanol extract from S. horneri (SHE) on immunoglobulin E (IgE)/bovine serum albumin (BSA)-mediated activation in bone marrow-derived cultured-mast cells (BMCMCs) and passive cutaneous anaphylaxis (PCA) reaction in mice. SHE markedly and dose-dependently suppressed the degranulation of BMCMCs by reducing the ß-hexosaminidase and histamine release without cytotoxicity. In addition, SHE significantly decreased the FcεRI expression on the surface of BMCMCs and its IgE binding. Moreover, SHE reduced the mRNA expression and the production of allergic cytokines; interleukin (IL)-1ß, IL-4, IL-5, IL-6, IL-10, IL-13; interferon (IFN)-γ and/or tumor necrosis factor (TNF)-α; and a chemokine, thymus and activation-regulated chemokine (TARC), by suppressing the activation of Src-family kinases and nuclear factor (NF)-κB signaling. In further study, the application of SHE reduced the PCA reaction in an IgE/BSA-induced type I allergic mice model. Taken together, we suggest that SHE has an anti-allergic effect in type I allergic responses.