Spin-induced band modifications of graphene through intercalation of magnetic iron atoms.

Intercalation of magnetic iron atoms through graphene formed on the SiC(0001) surface is found to induce significant changes in the electronic properties of graphene due mainly to the Fe-induced asymmetries in charge as well as spin distribution. From our synchrotron-based photoelectron spectroscopy data together with ab initio calculations, we observe that the Fe-induced charge asymmetry results in the formation of a quasi-free-standing bilayer graphene while the spin asymmetry drives multiple spin-split bands. We find that Fe adatoms are best intercalated upon annealing at 600 °C, exhibiting split linear π-bands, characteristic of a bilayer graphene, but much diffused. Subsequent changes in the C 1s, Si 2p, and Fe 3p core levels are consistently described in terms of Fe-intercalation. Our calculations together with a spin-dependent tight binding model ascribe the diffuse nature of the π-bands to the multiple spin-split bands originated from the spin-injected carbon atoms residing only in the lower graphene layer.

Humoral and cellular immune responses to Blomia tropicalis and concanavalin A-binding fractions in atopic patients.

Blomia tropicalis, Dermatophagoides pteronyssinus and D. farinae are prevalent house dust mites. Concanavalin A-binding components derived from B. tropicalis (Bt-ConA extract) are highly immunogenic in allergic diseases. The aim of the present study was to evaluate the humoral and cellular immune responses to B. tropicalis in mite-sensitized patients. A total of 137 patients with allergic rhinitis with/without asthma and 109 non-atopic subjects were selected and analyzed by the skin prick test, and for total serum IgE and specific IgE levels to both Bt-total and Bt-ConA extracts, their proliferative response and cytokine (IFN-gamma and IL-5) production by peripheral blood mononuclear cells (PBMC) stimulated with both extracts. Skin prick test showed that 70% of the patients were sensitized to Bt (Bt+) and similar levels of specific IgE to Bt-total and Bt-ConA extracts were demonstrable in Bt+ patients. Significant PBMC proliferation was observed in response to Bt-total extract in Bt+, but not in Bt- patients and non-atopic subjects (P < 0.001). Bt-ConA extract induced increased proliferative responses in all patient groups compared to medium alone (P < 0.05), but these responses were significantly decreased in the presence of the mannopyranoside ConA inhibitor (P < 0.05). Significant IFN-gamma production was observed after Bt-ConA stimulation of Bt+ patients (P < 0.05), while Bt-total extract had no effect. IL-5 production was consistently detected in Bt+ patients after allergen-specific stimulation or with no stimulus, indicating that PBMC from allergic patients are prone to produce Th2 profile cytokines, spontaneously or inductively by allergen restimulation. These data showed that ConA-binding components isolated from B. tropicalis may contain relevant antigens that are involved in both humoral and cellular immune responses. However, without an additional purification procedure to eliminate the residual contamination with ConA, its use in immunotherapeutic procedures cannot be recommended.

Humoral and cellular immune responses to Blomia tropicalis and concanavalin A-binding fractions in atopic patients

Blomia tropicalis, Dermatophagoides pteronyssinus and D. farinae are prevalent house dust mites. Concanavalin A-binding components derived from B. tropicalis (Bt-ConA extract) are highly immunogenic in allergic diseases. The aim of the present study was to evaluate the humoral and cellular immune responses to B. tropicalis in mite-sensitized patients. A total of 137 patients with allergic rhinitis with/without asthma and 109 non-atopic subjects were selected and analyzed by the skin prick test, and for total serum IgE and specific IgE levels to both Bt-total and Bt-ConA extracts, their proliferative response and cytokine (IFN-ã and IL-5) production by peripheral blood mononuclear cells (PBMC) stimulated with both extracts. Skin prick test showed that 70 percent of the patients were sensitized to Bt (Bt+) and similar levels of specific IgE to Bt-total and Bt-ConA extracts were demonstrable in Bt+ patients. Significant PBMC proliferation was observed in response to Bt-total extract in Bt+, but not in Bt- patients and non-atopic subjects (P < 0.001). Bt-ConA extract induced increased proliferative responses in all patient groups compared to medium alone (P < 0.05), but these responses were significantly decreased in the presence of the mannopyranoside ConA inhibitor (P < 0.05). Significant IFN-ã production was observed after Bt-ConA stimulation of Bt+ patients (P < 0.05), while Bt-total extract had no effect. IL-5 production was consistently detected in Bt+ patients after allergen-specific stimulation or with no stimulus, indicating that PBMC from allergic patients are prone to produce Th2 profile cytokines, spontaneously or inductively by allergen restimulation. These data showed that ConA-binding components isolated from B. tropicalis may contain relevant antigens that are involved in both humoral and cellular immune responses. However, without an additional purification procedure to eliminate the residual contamination with...

Mite-specific immunotherapy using allergen and/or bacterial extracts in atopic patients in Brazil.

OBJECTIVE: This study aimed to evaluate the clinical efficacy and antibody response changes after specific immunotherapy (SIT) using Dermatophagoides pteronyssinus (Dpt) allergens with or without bacterial extracts in Brazilian mite-atopic patients. METHODS: One-hundred patients with allergic rhinitis were selected for a randomized double-blind, placebo-controlled trial and distributed into 4 groups: Dpt (Dpt allergen extract), Dpt+MRB (Dpt allergen plus mixed respiratory bacterial extracts), MRB (MRB extract only) and placebo. Rhinitis symptom and medication scores; skin prick test (SPT) to Dpt extract; and serum immunoglobulin (Ig) E, IgG4, and IgG1 levels to Dpt, Der p 1, and Der p 2 allergens were evaluated before and after a year of treatment. RESULTS: After 1 year, the SPT response was reduced in the Dpt group (P=.03), whereas IgE levels to Der p 2 decreased only in the Dpt (P = .048) and Dpt+MRB (P = .005) groups. IgG4 and IgG1 levels to Dpt and Der p 1 increased in the Dpt group (P < .05), whereas in the Dpt + MRB group the IgG1 level only increased to Dpt (P=.001) and the IgG4 only increased to Der p 1 (P=.049). IgE levels to Dpt decreased only in the MRB (P= .005) and Dpt + MRB (P= .001) groups. Rhinitis symptom and medication scores fell in all groups, including the placebo group (P<.001). CONCLUSIONS: SIT using Dpt extract alone was effective in reducing SPT response and IgE levels to Der p 2 allergen, while bacterial extracts induced decreases in IgE levels to whole Dpt extract. However, only groups receiving Dpt allergen had higher levels of IgG1 and IgG4 to Dpt and Der p 1 after a year of treatment.

Mite and pet allergen exposure in hotels in Uberlândia, Midwestern Brazil.

UNLABELLED: Mite allergens are involved in airway sensitization and allergic diseases. We evaluated the exposure to house dust-mite (Der p 1 and Der f 1) and pet (Fel d 1 and Can f 1) allergens in hotels in Uberlândia, Midwestern Brazil. A total of 140 dust samples were collected from bedding (n = 98) and carpet (n = 42) of bedrooms in 20 hotels enrolled for this study. Geometric mean (GM) levels of Der f 1 (11.30 microg/g of dust; 95% CI: 8.34-15.30 microg/g) were significantly higher than Der p 1 (0.15 microg/g of dust; 95% CI: 0.13-0.18 microg/g) in bedding dust samples (P < 0.001), regardless of the hotel classes. Der f 1 levels were significantly higher in bedding (11.30 microg/g of dust; 95% CI: 8.34-15.30 microg/g) than carpet (6.32 microg/g of dust; 95% CI: 4.31-9.26 microg/g) dust samples (P < 0.05). High levels of Der f 1 (>10 microg/g of dust) were found in 58%, 76%, and 69% of dust samples from Simple, Economical, and Tourist/Superior hotels, respectively, while GM levels of Fel d 1 (0.11 microg/g of dust; 95% CI: 0.09-0.14 microg/g) and Can f 1 (0.30 microg/g of dust; 95% CI: 0.20-0.44 microg/g) were relatively low. These results indicate that Der f 1 is the predominant allergen in hotels in Uberlândia, especially in bedding dust samples, regardless of the hotel classes and could represent an important risk factor for exacerbation of allergic symptoms in previously mite-sensitized guests. PRACTICAL IMPLICATIONS: Mites and pets are important sources of indoor allergens. Most people spend the greatest part of their time indoors. Hotels can constitute an important allergen reservoir of the indoor environment and could represent an important risk for exacerbation of allergic symptoms in previously sensitized guests. Thus, hotels should also be included for planning indoor allergen avoidance as part of a global management strategy, especially in patients with respiratory allergy.

Responses of IgE, IgG1, and IgG4 to concanavalin A-binding Blomia tropicalis antigens in allergic patients.

Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the prevalent house dust mites in tropical countries and are associated with allergic diseases. Glycosylated antigens are highly immunogenic and involved in different pathologies. We evaluated the presence of IgE, IgG1, and IgG4 to concanavalin A-binding antigens (Bt-Con-A) isolated from Bt-total extract in sera of allergic and non-allergic subjects. Bt-total and Bt-Con-A extracts were evaluated by SDS-PAGE and ELISA for reacting with IgE, IgG1, and IgG4 in sera of 121 patients with allergic rhinitis and 36 non-allergic individuals. All subjects were skin prick tested with Bt-total extract and inhibition tests were performed for IgE, IgG1, and IgG4 using both extracts (Bt-total and Bt-Con-A). Skin prick test showed that 58% of the patients were sensitized to Bt (Bt+), with 52% reactive to both mites (Bt and Dp) and 6% to Bt only. A broad spectrum of proteins (14-152 kDa) was visualized in Bt-total and components >27 kDa for the Bt-Con-A extract. ELISA showed a similar profile of IgE, IgG1 and IgG4 levels in response to Bt-total and Bt-Con-A extracts in different groups, although Bt+ patients showed a lower IgG4 reactivity to Bt-Con-A extract. Specific IgG1 levels were higher in Bt+ patients than in control subjects, and IgG4 levels showed no significant difference among groups. ELISA inhibition showed a partial IgE and total IgG1 and IgG4 cross-reactivity with Dp extract for Bt-total and Bt-Con-A extracts. We conclude that Con-A-binding components isolated from Bt constitute major allergens and are involved in both allergen sensitization (IgE response) and homeostasis maintenance (IgG1 and IgG4 responses).

Responses of IgE, IgG1, and IgG4 to concanavalin A-binding Blomia tropicalis antigens in allergic patients

Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the prevalent house dust mites in tropical countries and are associated with allergic diseases. Glycosylated antigens are highly immunogenic and involved in different pathologies. We evaluated the presence of IgE, IgG1, and IgG4 to concanavalin A-binding antigens (Bt-Con-A) isolated from Bt-total extract in sera of allergic and non-allergic subjects. Bt-total and Bt-Con-A extracts were evaluated by SDS-PAGE and ELISA for reacting with IgE, IgG1, and IgG4 in sera of 121 patients with allergic rhinitis and 36 non-allergic individuals. All subjects were skin prick tested with Bt-total extract and inhibition tests were performed for IgE, IgG1, and IgG4 using both extracts (Bt-total and Bt-Con-A). Skin prick test showed that 58 percent of the patients were sensitized to Bt (Bt+), with 52 percent reactive to both mites (Bt and Dp) and 6 percent to Bt only. A broad spectrum of proteins (14-152 kDa) was visualized in Bt-total and components >27 kDa for the Bt-Con-A extract. ELISA showed a similar profile of IgE, IgG1 and IgG4 levels in response to Bt-total and Bt-Con-A extracts in different groups, although Bt+ patients showed a lower IgG4 reactivity to Bt-Con-A extract. Specific IgG1 levels were higher in Bt+ patients than in control subjects, and IgG4 levels showed no significant difference among groups. ELISA inhibition showed a partial IgE and total IgG1 and IgG4 cross-reactivity with Dp extract for Bt-total and Bt-Con-A extracts. We conclude that Con-A-binding components isolated from Bt constitute major allergens and are involved in both allergen sensitization (IgE response) and homeostasis maintenance (IgG1 and IgG4 responses).

Differential IgE reactivity to Der p 1 and Der p 2 allergens of Dermatophagoides pteronyssinus in mite-sensitized patients.

Several studies have shown that the presence of IgE antibodies to house dust mites (HDM), particularly Dermatophagoides pteronyssinus (Dpt), is an important risk factor for asthma. Allergen immunotherapy is indicated for patients with IgE antibodies to clinically relevant allergens. The aims of this study were to analyze the levels of specific serum IgE to Der p 1 and Der p 2 allergens in mite-sensitized atopic patients and to compare them with both in vivo (skin prick test) and in vitro (IgE-ELISA) sensitizations to Dpt crude extract. Forty-seven atopic patients with allergic rhinitis with or without intermittent or persistent mild asthma and positive skin prick test (SPT) to Dpt total extract were studied. Thirty age-matched healthy subjects with negative SPT to HDM were included as controls. Levels of total IgE and Dpt-, Der p 1- and Der p 2-specific IgE were measured by ELISAs in SPT-positive atopic patients and SPT-negative control subjects. Among 47 symptomatic atopic patients, 27 (57.4%) were double positive IgE to Der p 1 and Der p 2 allergens, 3 (6.4%) were single positive IgE to Der p 1, 4 (8.5%) were single positive IgE to Der p 2, and 13 (27.6%) were double negative IgE to both allergens. There was a significant correlation between Der p 1- and Der p 2-specific IgE levels, but not between Der p 1- or Der p 2-IgE levels and SPT results. The double negative IgE patients had the smallest skin test reactions although they showed high mean levels of total serum IgE. Therefore, the knowledge of specific IgE levels to Der p 1 and Der p 2 major allergens might support physicians for indication or follow-up in mite-sensitized patients under allergen-specific immunotherapy. These approaches might be important for obtaining improved safety and efficacy of the current clinical practice of allergen immunotherapy.

Taxis but not private cars are mite allergen reservoirs in Brazil.

Indoor allergens are major causative agents in allergic disease development. Besides homes, public transport vehicles have been considered important mite and pet allergen reservoirs. Our recent studies on allergen exposure in automobiles showed that different allergen levels are found in private cars versus taxis. We quantified group 1 Dermatophagoides spp. (Der 1), Felis domesticus (Fel d 1), and Canis familiaris (Can f 1) allergen levels by ELISA in dust samples from 60 taxi and 60 private car upholstered seats. Mean levels of Der 1 and Fel d 1 were significantly higher in taxis than private cars. A significantly higher percentage of taxis (42%) harboring sensitizing levels of Der 1 compared to private cars (5%) was also found. In spite of the low mean Fel d 1 levels, comparison of the percentage of vehicles with moderate Fel d 1 levels showed a significant difference between taxis and private cars (43% vs. 20%). On the other hand, mean Can f 1 levels were significantly higher in private cars compared to taxis concomitant with a significantly higher percentage of private cars containing moderate Can f 1 levels than taxis (53% vs. 28%). We conclude that upholstered seats from Brazilian taxis but not private cars constitute an important mite allergen reservoir. Thus, additional effective measures for the reduction of allergen exposure in vehicles within the global allergen avoidance strategy should also be routinely accomplished to minimize the induction of sensitization and symptoms in allergic patients.

IgE, IgG1, and IgG4 antibody responses to Blomia tropicalis in atopic patients.

BACKGROUND: Allergens from house dust mites (HDMs), Dermatophagoides pteronyssinus and Blomia tropicalis are clinically relevant in atopic respiratory diseases in tropical countries. AIMS OF THE STUDY: To evaluate immunoglobulin (Ig)E, IgG1, and IgG4 antibody responses to B. tropicalis in Brazilian atopic patients. METHODS: About 110 patients with allergic rhinitis with/without asthma and 33 control subjects underwent skin prick testing (SPT) with HDM extracts, and their sera were tested for IgE and IgG subclass antibodies to D. pteronyssinus and B. tropicalis by enzyme-linked immunosorbent assay (ELISA) and immunoblotting. RESULTS: Most patients (56%) had positive SPT to B. tropicalis extract (B. tropicalis+ group), although 51% were reactive to both B. tropicalis and D. pteronyssinus and 6% were sensitized to B. tropicalis only. IgE-ELISA detected 43%B. tropicalis positivity with high-specific IgE levels in B. tropicalis+ patients. Specific IgG4 levels were higher in B. tropicalis+ than B. tropicalis- groups and correlated with specific IgE levels. The IgG1 levels to B. tropicalis were higher in patients than controls. The major allergenic B. tropicalis components recognized by B. tropicalis+ patient sera were the 54, 66, and 68 kDa proteins. The IgG4-binding protein profiles closely resembled that of IgE. The IgG1 antibodies recognizing multiple B. tropicalis protein species were detected in sera of all three patient groups. CONCLUSIONS: A large percentage of our allergic patients are B. tropicalis+. They are more frequently sensitized to high-molecular weight (HMW) B. tropicalis components than the major low-molecular weight (11-15 kDa) allergens detected in other studies. The results suggest that HMW B. tropicalis antigenic components are potential candidates for evaluating allergen exposure and sensitization, and for immunotherapy treatment.

Mite allergen levels and acarologic analysis in house dust samples in Uberaba, Brazil.

Mite allergen exposure has been widely related to sensitization and development of allergic diseases. This study intended to evaluate the degree of allergen exposure in Uberaba, Brazil, through the measurements of Der f 1 and Der p 1 allergen levels associated with the acarologic analysis in house dust samples. A total of 240 dust samples were collected from 60 houses through vacuuming sofas and bedding, during the months of March and July 2000. Indoor temperature and relative humidity were also measured. Mites were counted and identified under light microscopy and allergen levels were measured by two-site monoclonal antibody ELISAs. The major mite family was Pyroglyphidae (39.4%), having D. pteronyssinus as the most frequent species (15.6%), followed by D. farinae (12.3%) and E. maynei (7.9%). The family Glycyphagidae was less commonly found (4.8%), with Blomia tropicalis (4.4%) as its majoritary member. The highest levels of Der f 1 and Der p 1 allergens were found in bedding samples in March (31.7 and 0.9 microg/g of dust, respectively), with Der f 1 levels significantly higher than Der p 1 (p < 0.0001). There was a significant positive correlation between the mite number and allergen levels. These results indicate that Dermatophagoides sp are the most frequent mites in our region followed by E. maynei. Therefore, the knowledge of the local mite fauna would improve the means of investigating the association between allergen exposure and sensitization, allowing the addition of new mite extracts in diagnostic tests.

Efficient presentation of house dust mite allergen Der p 2 by monocyte-derived dendritic cells and the role of beta 2 integrins.

Monocyte-derived dendritic cells (MDDC) are potent antigen-presenting cells (APC). The APC functions of MDDC in allergy were examined. MDDC presentation of the major house dust mite allergen Der p 2 resulted in 4-12-fold higher T-cell proliferation and markedly higher IFN-gamma and IL-5 production than PBMC cultures. Comparable T-cell proliferation was obtained with 10-fold fewer MDDC than purified monocytes. MDDC cultured from adherent cells, or CD14+, CD11b+ or Percoll-purified monocytes were comparable in presenting soluble Ag, and in stimulating allogeneic MLR. Importantly, MDDC presentation of Der p 2 resulted in both Th1 and Th2 stimulation, although MDDC are known to produce high levels of IL-12 and stimulate biased Th1 responses. The basis for the potent APC function of MDDC was further examined. MDDC were found to be highly phagocytic. Immunoprecipitation studies showed markedly elevated ICAM-1 expression but > 10-fold reduction in LFA-1 expression on MDDC compared with monocytes. Monoclonal antibody (MoAb)-blocking experiments showed that ICAM-LFA-1 interaction was essential for MDDC stimulation of Der p 2-specific T-cell proliferative responses. These results show that the use of MDDC as APC provides a simple, sensitive and versatile method for detecting T-cell responses to allergens and that the strong phagocytic capability and the increased ICAM-1 expression of MDDC contribute significantly to their Ag presentation potency.

Assessment of skeletal and dental changes by maxillary protraction.

To further evaluate the effect of maxillary protraction on facial growth, cephalometric changes in 129 subjects with conditions diagnosed as skeletal Class III malocclusion and who had been treated with maxillary protraction were compared to 9 male and 12 female subjects with annual cephalometric records from the Yonsei growth study sample. The control subjects had Angle Class I malocclusions with normal overjet and overbite. More maxillary forward displacement and mandibular growth inhibition were observed in the protraction group during treatment, and the difference from the untreated controls was statistically significant. When changes due to treatment according to ages were compared, there was no statistical difference. The direction of maxillary growth was similar in the untreated and protraction groups. Maxillary protraction had a growth-stimulating effect on the maxilla during the treatment period.

Pyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon, Pyrococcus furiosus, functions as a CoA-dependent pyruvate decarboxylase.

Pyruvate ferredoxin oxidoreductase (POR) has been previously purified from the hyperthermophilic archaeon, Pyrococcus furiosus, an organism that grows optimally at 100 degrees C by fermenting carbohydrates and peptides. The enzyme contains thiamine pyrophosphate and catalyzes the oxidative decarboxylation of pyruvate to acetyl-CoA and CO2 and reduces P. furiosus ferredoxin. Here we show that this enzyme also catalyzes the formation of acetaldehyde from pyruvate in a CoA-dependent reaction. Desulfocoenzyme A substituted for CoA showing that the cofactor plays a structural rather than a catalytic role. Ferredoxin was not necessary for the pyruvate decarboxylase activity of POR, nor did it inhibit acetaldehyde production. The apparent Km values for CoA and pyruvate were 0.11 mM and 1.1 mM, respectively, and the optimal temperature for acetaldehyde formation was above 90 degrees C. These data are comparable to those previously determined for the pyruvate oxidation reaction of POR. At 80 degrees C (pH 8.0), the apparent Vm value for pyruvate decarboxylation was about 40% of the apparent Vm value for pyruvate oxidation rate (using P. furiosus ferredoxin as the electron acceptor). Tentative catalytic mechanisms for these two reactions are presented. In addition to POR, three other 2-keto acid ferredoxin oxidoreductases are involved in peptide fermentation by hyperthermophilic archaea. It is proposed that the various aldehydes produced by these oxidoreductases in vivo are used by two aldehyde-utilizing enzymes, alcohol dehydrogenase and aldehyde ferredoxin oxidoreductase, the physiological roles of which were previously unknown.

Long- and short-term flooding effects on survival and sink-source relationships of swamp-adapted tree species.

About 95% of swamp tupelo (Nyssa sylvatica var. biflora (Walt.) Sarg.) and sweetgum (Liquidambar styraciflua L.) seedlings survived continuous root flooding for more than two years, whereas none of the swamp chestnut oak (Quercus michauxii Nutt.) and cherrybark oak (Q. falcata var. pagodifolia Ell.) seedlings survived one year of flooding. Death of oak seedlings occurred in phases associated with periods of major vegetative growth, e.g., after bud burst in spring, after summer stem elongation, and during the winter deciduous stage, suggesting that stored reserves and sources were inadequate to maintain the seedlings when vegetative sinks were forming. Additional evidence that flooding induced a source deficiency in oak was that leaves of flooded oak were 65 to 75% smaller than leaves of nonflooded oak. Flooded swamp tupelo seedlings had a normal leaf size and patchy stomatal opening compared with nonflooded seedlings. Flooding caused increases in alcohol dehydrogenase (ADH) specific activity in taproot cambial tissues and increases in starch concentrations of swamp tupelo seedlings that were reversed when seedlings were removed from flooding. Flooding had little effect on soluble sugar concentrations in swamp tupelo or sweetgum. In the long-term flood-dry-flood treatment, in which all species had survivors, upper canopy leaf photosynthetic rates were higher in all species during the dry period than in nonflooded controls, whereas their starch and soluble sugars concentrations were similar to those of nonflooded controls. Based on seedling survival and the sink-source relationships, the order of flood tolerance was: swamp tupelo > sweetgum > swamp chestnut oak > cherrybark oak.

Effects of the protein kinase C stimulant bryostatin 1 on the proliferation and colony formation of irradiated human T-lymphocytes.

The protein kinase C stimulant bryostatin 1 (Bryo) was used in examining human peripheral blood T-lymphocyte radiosensitivities in proliferation assays. Bryo was similar to PMA in inducing T-cell proliferation by the CD3, CD28 and CD69 pathways. No difference in radiosensitivities was observed in T-cells stimulated by the three independent surface antigen-mediated activation pathways. CD3 was chosen as the second signal for comparing the potencies of the three different first signals Bryo, phorbol 12-myristate, 13-acetate (PMA), and interleukin 2 (IL-2) in stimulating T-cell proliferation and in maintaining this response after radiation. Though there were radioresponse differences among various individuals, the irradiated lymphocytes consistently showed significantly greater proliferation when treated with Bryo or PMA than with IL-2 (p < 0.05- < 0.005). No difference in proliferative responses was observed in T-cells irradiated between 4 h before and 15 h after the addition of stimulants. Colony forming assays showed higher colony survival for irradiated T-cells stimulated with Bryo than with PMA. These results support the important role of protein kinase C in T-cell radiation responses, and suggest a potential role for Bryo in enhancing T-lymphocyte survival during radiation therapy.

Cytotoxicity and manganese superoxide dismutase induction by tumor necrosis factor-alpha and ionizing radiation in MCF-7 human breast carcinoma cells.

Both tumor necrosis factor-alpha (TNF) and ionizing radiation cause active-oxygen radical-mediated cell injuries and cell death. Thus cells treated by both TNF and radiation may suffer greater injuries than cells treated by either agent alone. On the other hand, TNF or radiation treatment can stimulate the expression of a mitochondrial superoxide scavenging enzyme, manganese superoxide dismutase (MnSOD), which can lower the cytotoxic effects of both agents. Thus, the induction of MnSOD by radiation may interfere with the cytotoxic action of TNF and vice versa. We used a human breast tumor cell line, MCF-7, to determine the interaction of TNF and radiation on cytotoxicity and MnSOD expression. TNF was found to be more effective as a cytotoxic agent when used before than after radiation treatment. These observations suggest that radiation induced-MnSOD was more effective in reducing the cytotoxic effect of TNF whereas TNF induction of MnSOD was less effective in counteracting the radiation action. Our results not only underscore the different effects of the treatment order of TNF and radiation, but also point to potential implication in the radiotherapy of breast tumors.

Combined treatments of heat, radiation, or cytokines with flavone acetic acid on the growth of cultured endothelial cells.

The antitumour effects of flavone acetic acid (FAA) against a broad spectrum of established experimental tumours has been demonstrated. Damage to the vasculature, which rapidly disrupts blood flow and induces haemorrhagic necrosis, is believed to be a major mechanism contributing to the observed antitumour effects. Despite these established observations, FAA has shown little effect against human tumours. However, other applications of FAA, for examples, for an extended period of treatments or in combination with other antitumour modalities, have not been sufficiently explored. In order to test the direct effects of FAA on vasculature, endothelial cells isolated from human umbilical vein (HUVEC) and bovine pulmonary artery (CPAEC) were used in this study. FAA at the concentrations of 50 to 200 micrograms/ml causes reduction in cell number (from 20 to > 30% of the cells) of HUVEC as measured by MTT assay after 1, 3, and 5 h of treatment at 37 degrees C. FAA did not produce significant effects on similarly treated human squamous cell carcinoma, cell line UM-SCC-2. After 1 h treatment of FAA at 300 micrograms/ml, a large number of HUVECs failed to react with an actin stain, NBD-phallacidin. The growth of HUVECs and CPAEC in the presence of FAA for 1-3 days was progressively reduced. The number of HUVEC treated for 3 days at the concentrations of 100, 200, and 300 micrograms/ml were reduced by 75-86% in comparison with the control culture. The experiments with CPAEC showed similar results. The inhibition of the growth of endothelial cells by FAA was enhanced when it combines with tumour necrosis factor-alpha but not with interleukin-1, interferon-gamma, heat, or radiation. We observed that FAA can initiate both immediate effects and growth inhibition on cultured endothelial cells. These results support the notion that FAA rapidly induces vasculature damage. Furthermore, cytokines such as tumour necrosis factor-alpha can enhance the toxicity of FAA on endothelial cells.

Vascular cambial sucrose metabolism and growth in loblolly pine (Pinus taeda L.) in relation to transplanting stress.

Sucrose synthase (SS) was the dominant enzyme of sucrose metabolism in both stem and root vascular cambial zone tissues of nursery-grown loblolly pine (Pinus taeda L.) seedlings. Acid invertase (AI) and neutral invertase (NT) activities were generally less than 10% of the SS activity in both tissues. In both cambial tissues, seasonal patterns of enzyme activity were observed for SS but not for AI or NI. The seasonal patterns of SS activity in stem and root cambia paralleled the periodic growth of stems and roots. Stems had high SS activity and growth during summer and early fall. Roots had substantial SS activity and growth during summer and fall, but SS activity and growth were even higher in winter. When seedlings were transplanted, about eight months elapsed before stem and root cambia resumed rates of growth and sucrose metabolism similar to those in control nontransplanted seedlings. Two months after transplanting, root SS was at its lowest, whereas AI activity in transplants was 50% higher than in control nontransplanted seedlings. In stems, SS activity decreased in response to transplanting, whereas AI and NI activities did not change appreciably. In loblolly pine tissues, SS was specific for uridylates, whereas the nucleotide triphosphate-dependent phosphofructokinase (NTP-PFK) had similar activity with either UTP or ATP. Except in winter, the NTP-PFK was less active than the pyrophosphate-dependent phosphofructokinase (PPi-PFK) during all seasons. The PPi-dependent PFK activity in nontransplanted seedlings followed similar seasonal and spatial patterns to those of SS activity. In actively growing tissues, such as stem cambial tissues in summer and root cambial tissues in winter, the measured total PFK to SS ratio ranged between 1.5/1 and 3/1. In contrast, in less actively growing tissues or transplanted seedlings, a greater decrease occurred in SS than in PFK activity, hence the ratio rose to as high as 12/1. It was concluded that: (1) SS was the dominant enzyme for sucrose metabolism in root and stem cambial tissues of loblolly pine seedlings; (2) both SS and PPi-PFK in the cambial tissues can be used as biochemical indicators of growth sink strength in stems and roots; and (3) both enzymes can be used as indicators of seedling stress caused by events such as transplanting and winter freezing.

Stimulation of interleukin-1 alpha and interleukin-1 beta production in human monocytes by protein phosphatase 1 and 2A inhibitors.

Protein phosphatases 1 and 2A are important in regulating cellular functions by controlling the phosphorylation state of their substrates. In human monocytes, the inhibitors of these phosphatases, okadaic acid and calyculin A, were found to increase the mRNA accumulation and cytokine production of interleukin-1 beta and interleukin-1 alpha. The increased mRNA accumulation was found to be primarily because of the increase in the transcription rate of the interleukin-1 genes. Stimulation of interleukin-1 gene transcription may be caused by the stimulation of transcription factor activities, including those of AP-1, by these protein phosphatase inhibitors. Okadaic acid increased the synthesis of the interleukin-1 beta precursor and mature forms and their secretion. This increased processing and secretion correlated with the stimulation of IL-1 beta convertase mRNA accumulation. The stimulation of interleukin-1 alpha production by okadaic acid was more modest than that of interleukin-1 beta. However, the phosphorylation of the precursor interleukin-1 alpha cytokine was increased. These results show that protein phosphatase 1 and 2A inhibitors exert multiple effects on cytokine production in human monocytes and suggest that these two phosphatases play important roles in regulating interleukin-1 production.