The inhibitory effect of naringenin on atopic dermatitis induced by DNFB in NC/Nga mice

Aims

Atopic dermatitis (AD) is a chronic and relapsing inflammatory dermatitis characterized by pruritic and eczematous skin lesions. Here, we investigated the therapeutic effect of the fruit flavonoid naringenin on DNFB induced atopic dermatitis mice model.

Main methods

AD-like skin lesion was induced by repetitive skin contact with DNFB in NC/Nga mice and the effects of the fruit flavonoid naringenin were evaluated on the basis of histopathological findings of skin, ear swelling and cytokine production of CD4⁺T cells.

Key findings

Intraperitoneal injection of naringenin for one week after DNFB challenge significantly lowered ear swelling and improved back skin lesions. In addition, naringenin significantly suppressed production of interferon-gamma (IFN-γ) by activated CD4⁺ T cells and serum IgE level. Furthermore, naringenin reduced DNFB-induced infiltration of eosinophils, mast cells, CD4⁺ T cells, and CD8⁺ T cells in skin lesions.

Significance

Naringenin may suppress the development of AD-like skin lesions in DNFB-treated NC/Nga mice by reducing IFN-γ production of activated CD4⁺ T cells, serum IgE levels and infiltration of immune cells to skin lesion.     

Clonal deletion of T cell repertoires with specific T cell receptor Vβ chains by two endogenous superantigens in NC/Nga mice

Superantigens (SAgs) are powerful T-cell stimulatory proteins. Because an atopic dermatitis (AD) model NC/Nga mice had two endogenous SAgs, namely minor lymphocyte-stimulating locus-1^a (Mls-1^a) and mouse mammary tumor virus (MMTV)(SHN), SAg-responsive T-cells bearing Vβ5.1, Vβ6, Vβ8.1, Vβ8.2, Vβ8.3, Vβ9, and Vβ11 should be endogenously deleted. Here, we discuss that the endogenous SAgs-expression may be involved in AD-sensitivity in NC/Nga mice.     

Supplemental feeding of phospholipid-enriched alkyl phospholipid from krill relieves spontaneous atopic dermatitis and strengthens skin intercellular lipid barriers in NC/Nga mice

Plasmalogen (Pls) is a glycerophospholipid derived from alkyl phospholipid (Alk) with antioxidant functions in vivo. The present study investigated the effects of ether phospholipids, such as Pls and Alk, on intercellular lipid barriers in the skin of NC/Nga mice, a model of atopic dermatitis (AD). NC/Nga mice fed Alk showed increased plasma levels of Alk and Pls. The AD-related changes in ceramide composition in the skin were abrogated by oral administration of Alk. Moreover, Alk suppressed skin inflammation in AD mice. These results indicate that Alk partially fortifies the stratum corneum lipid barrier and may be an effective treatment for AD.

Abbreviations: Pls: plasmalogen; PlsCho: choline plasmalogen; PlsEtn: ethanolamine plasmalogen; Alk: alkyl phospholipid; TJ: tight junction; FA: fatty acid; AD: atopic dermatitis; SO: soybean oil; FO: fish oil; DHA: docosahexaenoic acid; EPA: eicosapentaenoic acid; TG: triglyceride; PL: phospholipid; RF: retention factor; AlkCho: choline-type alkyl phospholipid; AlkEtn: ethanolamine-type alkyl phospholipid; LC-MS/MS: liquid chromatography-tandem mass spectrometry; FAR1: fatty acyl-coenzyme (Co)A reductase 1     

Lack of effect of the abnormal fatty acid metabolism in NC/Nga mice on their atopic dermatitis

Although clinical evidence has suggested that dysregulated fatty acid metabolism is associated with atopic disorders, the molecular basis for such a correlation remains to be demonstrated. In the present study, we analyzed the fatty acid composition in peripheral blood cells of NC/Nga mice, a model for atopic dermatitis (AD). We found that arachidonic acid significantly accumulated in mice with the AD manifestation. In addition, the leucotriene B4-releasing ability upon calcium ionophore A23187 stimulation was potentiated in blood cells. An arachidonic acid accumulation was not apparent in the non-atopic BALB/c strain, but was still observed in healthy NC/Nga mice fed under specific pathogen-free conditions. These results indicate that a disturbed fatty acid metabolism in NC/Nga mice was not a trigger factor for their dermatitis development.     

Orally supplemented Lactobacillus acidophilus strain L‐92 inhibits passive and active cutaneous anaphylaxis as well as 2,4‐dinitroflurobenzene and mite fecal antigen induced atopic dermatitis‐like skin lesions in mice

Oral supplementation of lactic acid bacteria is a potential approach to the prevention and manipulation of allergic diseases such as atopic dermatitis. Our previous report showed that heat‐killed Lactobacillus acidophilus strain L‐92 (L‐92) possessed anti‐allergic properties, although its physiological function in atopic dermatitis has largely remained undefined. To evaluate the anti‐allergic efficacy of L‐92, we used four experimental animal models with the major features of atopic dermatitis and compared the results to those of clinically active drugs. ICR mice were passively sensitized by anti‐dinitrophenyl mouse monoclonal IgE for passive cutaneous anaphylaxis (PCA), and BALB/c mice were actively sensitized by ovalbumin for active cutaneous anaphylaxis (ACA). Allergic reaction was induced by repeated exposure to 2,4‐dinitroflurobenzene (DNFB) and mite (Dermatophagoides farinae) fecal allergen, in BALB/c and NC/Nga mice, respectively. Orally administrated L‐92 significantly inhibited the vascular permeability increase in both PCA and ACA, and the elevation of ovalbumin‐specific IgE titer in ACA. Moreover, repeated applications of DNFB and mite fecal antigen onto the BALB/c and NC/Nga mouse ear, respectively, caused clinical symptoms similar to atopic dermatitis such as ear swelling, scratching behavior and elevation of total serum IgE levels that were also moderately suppressed by L‐92. In addition, L‐92 treated mice exhibited lower levels of mast cells, eosinophil infiltration and Th1/Th2 cytokine expression. Our results, therefore, suggest that oral administration of L‐92 might be useful for alleviating allergic symptoms.     

Inhibitory Effects of Heartwood Extracts of Broussonetia kazinoki Sieb on the Development of Atopic Dermatitis in NC/Nga Mice

We investigated the effects of a topically applied extract of the heartwood of Broussonetia kazinoki Sieb (B. kazinoki) on atopic dermatitis (AD)-like skin lesions induced by an extract of the house-dust mite Dermatophagoides farina in NC/Nga mice. We found that topically applied B. kazinoki extract suppressed the histological manifestations of AD-like skin lesions, and decreased the levels of plasma immunoglobulin E (IgE) and interleukin-4 (IL-4) in the mice. Moreover, B. kazinoki inhibited the induction of thymus-and-activation-regulated chemokine (TARC/CCL17), macrophage-derived chemokine (MDC/CCL22), and regulated-on-activation-normal T cell-expressed-and-secreted chemokine (RANTES/CCL5) in HaCaT cells activated by tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). In conclusion, our results suggest that B. kazinoki extract has therapeutic advantages in the treatment of AD.     

A novel hairless mouse model on an atopic dermatitis-prone genetic background generated by receptor-mediated transgenesis

Current mouse models for atopic dermatitis (AD) have a serious drawback, being the existence of dense hair on the body. Thus, a hairless animal model on an AD-prone genetic background will be a powerful tool to investigate the basis of and therapy for this complex disease. We applied the Toxin Receptor-mediated Cell Knockout (TRECK) method to generate a hairless transgenic (Tg) mice on the NC/Nga background, an AD-prone inbred strain. A minigene with the mouse Keratin71 (Krt71) promoter and human diphtheria toxin receptor, which intrinsically functions as the heparin-binding EGF-like growth factor, was introduced into the pronucleus of NC/Nga oocytes. Unexpectedly NCN24, one NC/Nga Tg line, showed a dominant hairless phenotype without diphtheria toxin administration. Furthermore, the atopic dermatitis-like predisposition and IgE elevation was observed in both NCN24 and the NC/Nga wildtype strain. NCN24 mice, which we have newly developed, will be useful to assess drugs for AD therapy, being able to monitor skin inflammation without shaving. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Oral Intake of <Emphasis Type="Italic">Lactobacillus helveticus</Emphasis> NS8 Alleviates Ovalbumin-Induced Atopic Dermatitis in SKH-1 Hairless Mice

Atopic dermatitis (AD) is the most common chronic inflammatory skin disease and is driven by strong type 2 immune responses. Lactobacillus helveticus NS8 (NS8), a probiotic strain isolated from Mongolian koumiss, has anti-inflammatory activities. Here, we evaluated the therapeutic potential of NS8 on AD-like skin lesions by using SKH-1 hairless mice that underwent three cycles of epicutaneous sensitization (EC) with ovalbumin (OVA). NS8 (5?×?10⁸ CFU/day) was orally administered to mice from 2 weeks before the first sensitization until the end of the study. NS8 attenuated the symptoms and pathological changes in the skin of AD mice. For example, NS8 reduced epidermal and dermal thickening and significantly restrained the infiltration of mast cells, eosinophils, and CD4⁺ T cells into the dermis. By analysing the Th1/Th2 cytokines produced in skin lesions, we found that NS8 significantly suppressed the expression of IL-4, IL-5, and IL-13 (P?<?0.05), while it had no discernible effect on the expression of IFN-γ. Systemically, NS-8 reduced the total IgE and OVA-specific IgE levels in serum (P?<?0.05). Our study demonstrates that oral administration of L. helveticus NS8 effectively alleviates AD severity in mice by suppressing the Th2 immune response. NS8 may be a promising candidate for prophylactic and therapeutic treatments of allergic diseases, such as AD.     

Oral administration of Lactobacillus strains from Kimchi inhibits atopic dermatitis in NC / Nga mice

Aims: Atopic dermatitis (AD) is marked by elevated levels of immunoglobulin E and skin lesions such as oedema and haemorrhage. Kimchi is a Korean fermented food that contains beneficial bacteria for human health. In this study, Lactobacillus plantarum CJLP55, CJLP56, CJLP133 and CJLP136 isolated from Kimchi were investigated for their capacity to inhibit AD. Methods and Results: The three strains, CJLP55, CJLP133 and CJLP136, suppressed AD‐like skin lesions, high serum IgE levels and epidermal thickening. The three strains diminished the accumulation of eosinophils and mast cells into topical inflammatory sites and the enlargement of axillary lymph nodes, which are responsible for the dorsal dermatitis. CJLP55, CJLP133 and CJLP136 decreased production of type 2 cytokines such as IL‐4 and IL‐5 in lymph node cell culture. CJLP133 and CJLP136 increased IFN‐γ secretion, while CJLP55 enhanced IL‐10 production. Conclusions: The three strains isolated from Kimchi suppress house‐dust mite‐induced dermatitis in NC/Nga mouse, a representative animal model of human AD. Significance and Impact of the Study: These findings suggest that lactobacilli isolated from Kimchi inhibit AD, probably by altering the balance of Th1/Th2 ratio or inducing IL‐10 production.     

Effect of dendritic cells treated with CpG ODN on atopic dermatitis of Nc/Nga mice

Atopic dermatitis (AD) is a chronic inflammatory skin disease and the pathogenesis of AD is associated with the release of various cytokines/chemokines due to activated Th(2) immune responses. Synthetic oligodeoxynucleotides (ODNs) containing unmethylated CpG dinucleotide in the context of particular base sequence (CpG motifs) are known to have the immunostimulatory activities in mice and to convert from Th(2) to Th(1) immune responses in AD. We aimed to investigate that CpG ODN, especially phosphodiester form, can stimulate the protective immunity in NC/Nga mice with AD. We isolated BMDCs from NC/Nga mice and then, cultured with GM-CSF and IL-4 for 6 days, and treated for 2 days by either phosphorothioate ODN or phosphodiester ODN. CpG ODN-treated DCs resulted in more production of IL-12. When CpG ODN-treated DCs were intravenously injected into the NC/Nga mice, the NC/Nga mice with CpG ODN-treated DCs showed significant improvement of AD symptoms and decrease of IgE level. Histopathologically, the NC/Nga mice skin with CpG ODN-treated DCs showed the decreased IL-4 and TARC expression comparing with non-injected mice. These results may suggest that phosphodiester CpG ODN-treated DCs might function as a potent adjuvant for AD in a mouse model.     

Atopic dermatitis‐mitigating effects of new Lactobacillus strain,Lactobacillus sakei probio 65 isolated from Kimchi

Aims

Atopic dermatitis (AD) is an inflammatory skin disease. Probiotics have been reported to modulate immune responses and thus are now being suggested as potential treatments for allergies. In this study, we investigated the inhibitory effects of Lactobacillus sakei probio 65 isolated from Kimchi on artificially inducing AD in NC/Nga mice.

Methods and Results

Oral administration of viable or heat‐inactivated Lact. sakei probio 65 improved the condition of skin and reduced scratching frequency. Serum levels of IgE and cutaneous T‐cell‐attracting chemokine (CTACK) were significantly decreased by this therapy. Dead Lact. sakei probio 65 also decreased IL‐4 and IL‐6 serum concentrations. Moreover, both live and dead Lact. sakei probio 65 inhibited the expression of Thymus and activation‐regulated chemokine and CTACK in AD‐like skin lesions. The increased levels of Foxp3 expression in the lesional skin and ears were also suppressed by Lact. sakei probio 65. In addition, Lact. sakei probio 65 inhibited β‐hexosaminidase release and the secretion of IL‐4, TNF‐α and IL‐6 from RBL‐2H3 cells.

Conclusions

Oral treatment with both viable and heat‐inactivated Lact. sakei probio 65 inhibits skin inflammation and AD‐like skin lesions, as well as mast cell activation.

Significance and Impact of the Study

Lactobacillus sakei probio 65 has an inhibitory effect on atopic dermatitis‐like skin lesions and may represent an effective new anti‐inflammatory agent.     

Enhancement of the Non‐Specific Defence Activities in Carp (Cyprinus carpio) and Flounder (Paralichthys olivcaces) by Oral Administration of Schizophyllan

The effect of an immuno‐stimulative β‐glucan from Schizophyllum commune, schizophyllan, on the enhancement of fish immunity was evaluated with carp and flounder. The oral administration of schizophyllan induced a reduction in cumulative mortality after a bacterial challenge. Whereas mortality in the control groups was 100% after seven days, it was only 60% after challenging carp (Cyprinus carpio) with 1 × 10⁶ cells of Aeromonas hydrophila and 70% for flounder (Paralichthys olivaceus) using 1 × 10⁶ cells of Edwardsiella tarda. The numbers of peripheral macrophages and neutrophiles, the phagocytic activities of leukocytes and the activity of serum lysozyme were increased in the fish being fed a schizophyllan‐supplemented diet. These results support the findings that non‐specific defence activities in fish such as the number of leukocytes, phagocytic activities and serum lysozyme activity could be enhanced by oral administration of schizophyllan. Moreover, the oral administration of schizophyllan can reduce mortality after bacterial infections depending on the size of fish and the concentrations of bacteria.     

A combination of Olea europaea leaf extract and Spirodela polyrhiza extract alleviates atopic dermatitis by modulating immune balance and skin barrier function in a 1-chloro-2,4-dinitrobenzene-induced murine model

BackgroundAtopic dermatitis is a chronic inflammatory skin disease in humans. Although Olea europaea leaf extract (OLE) and Spirodela polyrhiza extract (SPE) have been used to protect against skin damage, the effects of their combined administration on atopic dermatitis have yet to studied.PurposeIn this study, we evaluated the potential therapeutic effects of an OLE and SPE combination on the progression of atopic dermatitis and the possible mechanisms underlying these effects in 1-chloro-2,4-dinitrobenzene (DNCB)-treated NC/Nga mice.MethodsAtopic dermatitis was induced by topical application of 0.2% w/v DNCB prepared in an olive oil:acetone solution (1:3), and thereafter OLE, SPE and OLE + SPE were administered orally for 5 weeks. We determined atopic dermatitis symptoms, serum IgE levels, and levels of cytokine- and gene expression in the dorsal skin and splenocytes, and performed histological and immune cell subtype analyses. The expression of skin barrier-related proteins (filaggrin, sirtuin 1, and claudin 1) was also evaluated.ResultsThe OLE + SPE combination significantly ameliorated atopic dermatitis symptoms, including dermatitis scores, and reduced epidermal thickness and infiltration of different inflammatory cells in mice with DNCB-induced atopic dermatitis. It also significantly reduced the number of CD4⁺, CD8⁺, and CD4⁺/CD69⁺ T cells; immunoglobulin E-producing B cells (CD23⁺/B220⁺) in the axillary lymph nodes; CD3⁺ T-cell eosinophils (chemokine–chemokine receptor 3⁺/CD11b⁺) in the skin; and CD3⁺ T cells, immunoglobulin E-producing B cells (CD23⁺/B220⁺), and eosinophils in peripheral blood mononuclear cells. Additionally, the experimental combination lowered levels of serum immunoglobulin E and histamine, as well as Th2-mediated cytokines, and interleukin-4, -5, and -13, whereas it increased the levels of Th1-mediated cytokine interferon-γ in splenocytes. Furthermore, the preparation significantly restored expression of the skin barrier-related proteins filaggrin, sirtuin 1, and claudin 1, and also reduced the expression of the inflammatory cytokine interleukin-6 and chemokine–chemokine receptor 3, as well as the pruritus-related cytokine interleukin-31 and interleukin-31 receptor, in atopic dermatitis skin lesions.ConclusionTaken together, our findings indicate that administration of a combination of OLE and SPE can alleviate atopic dermatitis symptoms by regulating immune balance and skin barrier function and may be an effective therapeutic option for the treatment of atopic dermatitis.     

Transplantation of expanded bone marrow‐derived very small embryonic‐like stem cells (VSEL‐SCs) improves left ventricular function and remodelling after myocardial infarction

Adult bone marrow‐derived very small embryonic‐like stem cells (VSEL‐SCs) exhibit a Sca‐1⁺/Lin^–/CD45^– phenotype and can differentiate into various cell types, including cardiomyocytes and endothelial cells. We have previously reported that transplantation of a small number (1 × 10⁶) of freshly isolated, non‐expanded VSEL‐SCs into infarcted mouse hearts resulted in improved left ventricular (LV) function and anatomy. Clinical translation, however, will require large numbers of cells. Because the frequency of VSEL‐SCs in the marrow is very low, we examined whether VSEL‐SCs can be expanded in culture without loss of therapeutic efficacy. Mice underwent a 30 min. coronary occlusion followed by reperfusion and, 48 hrs later, received an intramyocardial injection of vehicle (group I, n= 11), 1 × 10⁵ enhanced green fluorescent protein (EGFP)‐labelled expanded untreated VSEL‐SCs (group II, n= 7), or 1 × 10⁵ EGFP‐labelled expanded VSEL‐SCs pre‐incubated in a cardiogenic medium (group III, n= 8). At 35 days after myocardial infarction (MI), mice treated with pre‐incubated VSEL‐SCs exhibited better global and regional LV systolic function and less LV hypertrophy compared with vehicle‐treated controls. In contrast, transplantation of expanded but untreated VSEL‐SCs did not produce appreciable reparative benefits. Scattered EGFP⁺ cells expressing α‐sarcomeric actin, platelet endothelial cell adhesion molecule (PECAM)‐1, or von Willebrand factor were present in VSEL‐SC‐treated mice, but their numbers were very small. No tumour formation was observed. We conclude that VSEL‐SCs expanded in culture retain the ability to alleviate LV dysfunction and remodelling after a reperfused MI provided that they are exposed to a combination of cardiomyogenic growth factors and cytokines prior to transplantation. Counter intuitively, the mechanism whereby such pre‐incubation confers therapeutic efficacy does not involve differentiation into new cardiac cells. These results support the potential therapeutic utility of VSEL‐SCs for cardiac repair.     

Metabolie Aspects in Mice Administered Live Salmonella typhimurium

The influence of intraperitoneal inoculation of live Salmonella typhimurium on carbohydrate and lipid metabolisms in mice was investigated at doses of 9.2 × 10⁷ cells, 1.9 × 10⁸ cells, and 3.8 × 10⁸ cells. The hepatic glycogen content in mice at 18 hr after the inoculation decreased in inverse proportion to an increase in the injection dose. The activities of hepatic phosphorylase and G-6-P_ase increased significantly after 2 hr, but after 18 hr the levels of both enzyme activities, especially G-6-P_ase, declined in inverse porportion to an increase in dose of viable cells administered to the mice. The levels of serum glucose and free fatty acids (FFA) in mice markedly decreased at doses of 1.9 × 10⁸ and 3.8 × 10⁸ cells after a transient rise at early stage (1 hr) after the injection. Marked hypertriglyceridemia was seen in infected mice. However, the activity in serum lipoprotein lipase (LPL) was reduced by an increase in the injection dose. The effect of intraperitoneal administration of viable cells on the serum triglyceride level was prevented in mice immunized with S. typhimurium endotoxin or administered with the anti-endotoxin serum. These results indicate that hypertriglyceridemia mainly results from the action of endotoxin in the pathogen. Serum lactate dehydrogenase (LDH) activity markedly increased at the dose of 3.8 × 10⁸ cells within 8–16 hr, and the infected mice exhibited a leakage of isozymes LDH-3 and 5 in the serum 16 hr post-inoculation.     

Very high density of Chinese hamster ovary cells in perfusion by alternating tangential flow or tangential flow filtration in WAVE bioreactor™—part II: Applications for antibody production and cryopreservation

A high cell density perfusion process of monoclonal antibody (MAb) producing Chinese hamster ovary (CHO) cells was developed in disposable WAVE Bioreactor? using external hollow fiber (HF) filter as cell separation device. Tangential flow filtration (TFF) and alternating tangential flow (ATF) systems were compared and process applications of high cell density perfusion were studied here: MAb production and cryopreservation. Operations by perfusion using microfiltration (MF) or ultrafiltration (UF) with ATF or TFF and by fed‐batch were compared. Cell densities higher than 10⁸ cells/mL were obtained using UF TFF or UF ATF. The cells produced comparable amounts of MAb in perfusion by ATF or TFF, MF or UF. MAbs were partially retained by the MF using ATF or TFF but more severely using TFF. Consequently, MAbs were lost when cell broth was discarded from the bioreactor in the daily bleeds. The MAb cell‐specific productivity was comparable at cell densities up to 1.3 × 10⁸ cells/mL in perfusion and was comparable or lower in fed‐batch. After 12 days, six times more MAbs were harvested using perfusion by ATF or TFF with MF or UF, compared to fed‐batch and 28× more in a 1‐month perfusion at 10⁸ cells/mL density. Pumping at a recirculation rate up to 2.75 L/min did not damage the cells with the present TFF settings with HF short circuited. Cell cryopreservation at 0.5 × 10⁸ and 10⁸ cells/mL was performed using cells from a perfusion run at 10⁸ cells/mL density. Cell resuscitation was very successful, showing that this system was a reliable process for cell bank manufacturing. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:768–777, 2013     

Large Voltage Generation of Flexible Thermoelectric Nanocrystal Thin Films by Finger Contact

This paper demonstrates that thermal energy radiated from a human finger can be converted efficiently into electricity by a nanocrystal (NC) thin film that substantially suppresses thermal conduction, but still allows electric conduction. The converting efficiencies of the chalcogenide NC thin films with dimensions 40 µm × 20 µm × 20 nm, prepared on flexible substrates by a solution process, are maximized by adjusting the NC size. A Seebeck coefficient of S = 1829 µV K^?1, and a dimensionless thermoelectric figure‐of‐merit, ZT = 0.68 are achieved at ambient temperature for p‐ and n‐type NC thin films, respectively. A thermoelectric array consisting of p‐ and n‐type NC thin films generates a voltage of 645 mV for a temperature gradient of 10 K. Furthermore, the donut‐shaped pn array can generate a voltage of 170 mV from the heat supplied by an individual's finger.     

Real‐time PCR quantification of Bemisia tabaci (Homoptera: Aleyrodidae) B‐biotype remains in predator guts

The cotton whitefly, Bemisia tabaci (Gennadius) B‐biotype, is fed on by a wide variety of generalist predators, but there is little information on these predator–prey interactions, especially under field conditions. In this study, a real‐time polymerase chain reaction (PCR) assay was developed to quantify B. tabaci B‐biotype remains in predator gut. The B. tabaci B‐biotype genomic DNA copy number was referred to the actual amount of BT1 isolate, the B. tabaci B‐biotype specific DNA fragment. The numbers of BT1 isolate in one B. tabaci B‐biotype egg, individual adult and a single red‐eyed nymph were 2.56 × 10³, 2.56 × 10⁴, and 1.29 × 10⁴ copies, respectively. When Propylaea japonica adults fed on one, two, four, eight or 16 red‐eyed nymphs, the detected numbers of BT1 isolate ranged from 2.77 × 10⁴ to 4.05 × 10⁵ copies, forming a strong linear relationship (R² = 0.9899). Following the consumption of two red‐eyed nymphs, prey DNA was detectable in 100% of P. japonica at t = 0, decreasing to 80.0% and 60.0% after 1–4 h and 8 h of digestion, respectively, with 3.36 × 10⁴–1.25 × 10³ BT1 isolate copies. The predation by field‐collected predators, 26 larvae of P. japonica, and of Harmonia axyridis each, Chrysopa spp. larvae (Chrysopa pallens and C. formosa, 18 individuals in total), and a single adult of Scymnus hoffmanni, 19 adults of Orius sauteri and nine adult spiders (Erigonnidium graminicolum and Neoscona doenitzi), on B. tabaci B‐biotype were quantified. Of the 99 analysed predator individuals, 3.65 × 10²–4.60 × 10⁵ copies of BT1 isolate, equivalent to 0.8–18.8 red‐eyed nymphs were detected. These results suggest that TaqMan real‐time PCR technology may provide a rapid and sensitive method for quantifying B. tabaci B‐biotype remains in predator guts and will be invaluable in assessing the food web relationship between prey and arthropod predators.     

Inability of IL-12 to down-regulate IgE synthesis due to defective production of IFN-gamma in atopic NC/Nga mice.

NC/Nga mice raised in nonsterile circumstances spontaneously suffer from atopic dermatitis-like skin lesions with IgE hyperproduction. We investigated effects of rIL-12 on the IgE production in NC/Nga mice. rIL-12 administration was successful to suppress the increase of IgE levels in BALB/c mice immunized with OVA and aluminum hydroxide, but failed to abrogate that in NC/Nga mice. Both in vivo and in vitro IFN-gamma production induced by rIL-12 was less in NC/Nga mice than in BALB/c mice. Addition of rIFN-gamma to rIL-4 and LPS completely abrogated IgE production by B cells of BALB/c mice, but was insufficient to suppress it by B cells of NC/Nga mice. In splenic cells pretreated with Con A, STAT4 was phosphorylated at the tyrosine residue by addition of rIL-12, which was more weakly inducible in NC/Nga mice than in BALB/c mice. Finally, we examined the preventive ability of rIL-12 on the clinical aspects of atopic dermatitis in NC/Nga mice. rIL-12 administration resulted in exacerbation of development of the skin lesions and IgE production in NC/Nga mice raised in nonsterile circumstances. These results suggest that defective production of IFN-gamma by T cells less sensitive to IL-12 and low responsiveness of B cells to IFN-gamma may contribute to IgE hyperproduction in NC/Nga mice, and that IL-12 may have no ability to improve the clinical aspects of NC/Nga mice.