Biomedical subjects

W L Lin

Publications and source records attributed to W L Lin.

At least 55 records · Page 3Linked to original sources

PubMed1999

Optimal configuration of multiple-focused ultrasound transducers for external hyperthermia.

External ultrasound hyperthermia is considered to be a very flexible modality for heating deep-seated tumors owing to its penetration and focusing ability. However, using this flexibility requires that many complicated, interacting decisions be made to obtain optimal treatment. This paper presents the feasibility of arranging multiple-focused ultrasound transducers to produce an appropriate heating pattern for a specific treatment, based on the optimal scan parameters obtained from an optimization algorithm. The variable scan parameters of the heating system optimized are the transducer tilt and rotation angles, focal depth, scan radius, and output acoustical power. After obtaining the optimal scan parameters, multiple transducers are systematically arranged according to these scan parameters. Three-dimensional ultrasound power deposition and temperature distribution for a specific treatment are calculated for this multiple ultrasound transducer system. A more uniform temperature distribution in the treatment region for a large, highly perfused tumor can be achieved by scanning the system with respect to the central scan axis and/or swinging the transducers inwards and outwards. The maximum heating depth of focused ultrasound transducers used in this heating system is also studied. Simulation results demonstrate that the optimal arrangement of this multiple-focused ultrasound transducer system is highly promising for heating deep, large, and highly perfused tumors.

Algorithms↗

PubMed1999

Alkaline serine proteinase is a major allergen of Aspergillus flavus, a prevalent airborne Aspergillus species in the Taipei area.

BACKGROUND: Aspergillus species are prevalent indoor airborne fungi and have been identified to be a causative agent of human allergic disorders. In the present study, we identified, purified and characterized the allergen(s) from Aspergillus flavus, a predominant airborne Aspergillus species in the Taipei area. METHODS: The IgE-binding components of A. flavus were identified by SDS-PAGE immunoblotting with sera from asthmatic patients. The N-terminal amino acid sequences of the major allergens were determined by Edman degradation. The allergenic cross-reactivity among allergens from different fungi was analyzed by immunoblot inhibition using sera from asthmatic patients. The detected major allergen was purified from the culture medium. It was further characterized in terms of its N-terminal amino acid sequence, its IgE-binding activity and its enzymatic activity. RESULTS: The results of the immunoblot analysis indicate that a 34-kD component that has high IgE-binding (63%) frequency is a major allergen of A. flavus. The N-terminus of this 34-kD major allergen (GLTTQKSAP) has high sequence identity with that of the 34-kD alkaline serine proteinase major allergen of A. oryzae. Results from immunoblot inhibition studies indicate that IgE cross-reactivity occurs among the 34-kD major allergens of A. flavus, A. fumigatus and Penicillium citrinum. The 34-kD major allergen of A. flavus was purified from the culture medium by ammonium sulfate precipitation and DEAE ion exchange chromatography. The N-terminal amino acid sequence of the purified allergen (Asp fl 13) is identical to that determined previously for the 34-kD major allergen in the crude extract of A. flavus. The IgE immunoblot reactivity to the 34-kD major allergen in the crude extract can be dose-dependently inhibited by the purified Asp fl 13. The degree of IgE binding to the 34-kD major allergen in the crude extract correlates with that of the purified Asp fl 13 in sera of 8 asthmatic patients. The purified Asp fl 13 has proteolytic activity with casein as substrate at pH 8.0. This enzymatic activity can be inhibited by either phenylmethylsulfonyl fluoride or diethylpyrocarbonate. CONCLUSION: Our results suggest that the 34-kD alkaline serine proteinase is a major allergen of A. flavus. There was IgE cross-reactivity among allergens of A. flavus, A. fumigatus and P. citrinum.

Adolescent↗

PubMed1999

Inhibitory effect of 12-O-tetradecanoylphorbol-13-acetate-caused tumor promotion in benzo[a]pyrene-initiated CD-1 mouse skin by baicalein.

The effects of topical application of baicalein on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced promotion of skin tumors, hyperplasia, ornithine decarboxylase activity, and inflammation were evaluated in female CD-1 mice. Topical application of baicalein (0.08, 0.16, or 0.2 mumol) with TPA (5 nmol) twice weekly for 24 weeks to mice previously initiated with benzo[a]pyrene inhibited the number of TPA-induced tumors per mouse significantly. Preapplication of the same amount of baicalein also afforded significant protection against TPA-induced hyperplasia in the ear skin. Topical application of baicalein inhibited tumor promoter-caused induction of epidermal ornithine decarboxylase activity by TPA (5 nmol). The topical application of baicalein (0.008, 0.016, or 0.02 mumol) inhibited TPA-induced edema of mouse ears by 88%, 96%, or 97%, respectively. Pretreatment of mouse skin with various amounts of baicalein caused inhibition of H2O2 and myeloperoxidase formation by TPA. These results indicate that baicalein can be a potential cancer-chemopreventive agent against tumor promotion.

Animals↗

PubMed1999

N-cadherin expression during periodontal ligament cell differentiation in vitro.

BACKGROUND: When confluent periodontal ligament (PDL) cells were cultured in the presence of dexamethasone (Dex), ascorbic acid (AA), and beta-glycerophosphate (GP), they underwent sequential differentiation, demonstrating distinct morphological characteristics. At 1 week, localized cell proliferation led to the formation of multilayers of cells. As cell differentiation progressed, they formed nodules by deposition of matrix in the clusters of cells at 2 weeks, and mineralized the nodules at 3 weeks. These changes implicate extensive cell-to-cell interactions. Cadherins are known to play an important role in establishing cell contacts during tissue formation. METHODS: To determine whether cadherins are involved in PDL cell differentiation, and the formation and mineralization of nodules by the cells in vitro, we investigated the expression of N-cadherin using immunofluorescence labeling and Northern blot analysis. RESULTS: Immunolabeling showed that N-cadherin was expressed in PDL cells in the stages of nodule formation and mineralization. Northern blot analysis demonstrated a 3-fold increase in the expression of N-cadherin mRNA in the stages. However, neither E-cadherin nor P-cadherin was expressed. CONCLUSIONS: Our data suggest that N-cadherin may play an important role in PDL cell differentiation and the formation of mineralized nodules by PDL cells.

Alkaline Phosphatase↗

PubMed1998

Promotional effect of N-nitroso-N-(3-keto-1,2-butanediol)-3'-nitrotyramine (a nitrosated Maillard reaction product) in mouse fibroblast cells.

N-Nitroso-N-(3-keto-1,2-butanediol)-3'-nitrotyramine (NO-NTA) is a product of model browning system generated in the presence of sodium nitrite. The chemical structure of this compound has been confirmed by UV, mass and nuclear magnetic resonance, and infrared spectroscopy in our previous study. A two-stage transformation protocol was used to chemically transform the mouse embryo fibroblasts C3H10T1/2 cells. To initiate transformation, the cells were treated with benzo[a]pyrene (BaP) (0.1 mg/ml), and NO-NTA (0.01, 0.1 and 1 mg/ml) was employed subsequently to complete the transformation process. Malignant transformed foci were formed in BaP-initiated and NO-NTA promoted C3H10T1/2 cells after 8 wk. Cells treated with NO-NTA alone failed to induce transformation. However, cells initiated with BaP and promoted by cells initiated with BaP and promoted by NO-NTA demonstrated oncogenic properties. Cell lines transformed with NO-NTA-transformed colonies exhibited enhanced growth rate, anchorage independence and tumorigenicity in animals relative to parent cells. These results indicate that NO-NTA is a new tumour promoter and may induce tumour promotion by two-stage oncogenesis. Further studies on the mechanism of action of NO-NTA are now in progress.

Animals↗

PubMed1998

Specific absorption rate ratio patterns of cylindrical ultrasound transducers for breast tumors.

The purpose of this paper is to examine the optimal driving frequency and to configure the ultrasound energy deposition schema for a various size and location of breast tissues when a portion or the entire cylindrical ultrasound transducer is employed for breast hyperthermia treatments. This work employs a computer simulation program based on an ideal ultrasound power deposition from a cylindrical transducer. The ultrasound power within the breast is assumed to be exponentially attenuated according to the penetration depth of the ultrasound beam and a uniform absorption for the entire breast is also assumed. The distribution of the specific absorption rate (SAR) ratio is employed to determine the heating pattern of a set of given parameters. The control parameters considered are the ultrasound frequency in the breast tissue, the active portion of cylindrical transducer, and the shifting distance between the central axes of the breast and the transducer. The effect of the breast size on the SAR ratio is also considered. Simulation results demonstrate that the breast size, the ultrasound frequency in breast tissue, the shifting distance, and the active portion of the cylindrical transducer are the potential parameters for influencing the distribution of the SAR ratio. High frequencies should be used for the superficial heating treatments and the active portion of the transducer can be changed to obtain a region with an appropriate SAR ratio to cover the treatment region. Low frequencies are used for deep heating treatments and the region of the high SAR ratio can be moved by shifting the transducer and its pattern is varied with the transducer's active portion. The distribution of the SAR ratio indicates the domain of treatable tumor size and tumor depth for a given set of parameters (driving frequency, shifting distance and active portion of the transducer, as well as breast diameter). Findings of this study can be used to know whether or not the tumor is treatable as well as to select the optimal driving frequency and the appropriate active portion of the cylindrical transducer for a treatment, and hopefully to design an appropriate cylindrical ultrasound heating system for breast tumors.

Biophysical Phenomena↗

PubMed1998

Alkaline serine proteinase: a major allergen of Aspergillus oryzae and its cross-reactivity with Penicillium citrinum.

BACKGROUND: Aspergillus species are common indoor airborne fungi and have been considered as causative agents of human allergic disorders. However, allergens of different Aspergillus species have not been effectively characterized. The object of this study was to identify and characterize IgE-binding components of Aspergillus oryzae. METHODS: Allergens of A. oryzae were identified by immunoblot analysis using sera from asthmatic patients. The N-terminal amino acid sequences of allergens thus identified were determined by Edman degradation. The antigenic and the allergenic cross-reactivities between allergens of different fungi were analyzed by immunoblotting and immunoblot inhibition analysis, respectively, using a monoclonal antibody (MoAb) 55A against the 33-kD major allergen of Penicillium citrinum and a mixture of IgE-containing asthmatic serum samples. RESULTS: Thirteen components of A. oryzae ranging in apparent molecular weight from 16 to 42 kD react with IgE antibodies. A 34-kD component that showed intense IgE-binding reactivity and was detectable in the highest frequency in our asthmatic serum samples tested was considered a major allergen of A. oryzae. The 34-kD component also reacted with MoAb 55A. Results from immunoblot inhibition studies also demonstrated the IgE cross-reactivity between the 34-kD major allergens of A. oryzae and P. citrinum. In addition, the sequence of the N-terminal 18 amino acid residues of the 34-kD major allergen of A. oryzae was found to be identical to that of the alkaline serine proteinase from the same Aspergillus species. CONCLUSION: The 34-kD major allergen of A. oryzae is an alkaline serine proteinase. There is IgE cross-reactivity between the major serine proteinase allergens of A. oryzae and P. citrinum.

Allergens↗

PubMed1998

Amplification of T-cell receptor alpha- and beta-chain transcripts from mouse spleen lymphocytes by the nonpalindromic adaptor-polymerase chain reaction.

We employed the nonpalindromic adaptor-PCR (NPA-PCR) method to amplify T-cell receptor (TCR) alpha- and beta-chain transcripts from the spleen of normal SJL mice. The NPA-PCR method has been specifically designed for the amplification of transcripts with variable or unknown 5' ends, such as TCRs and immunoglobulins (Ig). This method has certain distinct advantages over existing two-sided PCR methods for the amplification of TCR transcripts. Two NPA-PCR amplifications are sufficient to amplify all the TCR transcripts (one for the alpha-chain and another one for the beta-chain). Amplification of TCR transcripts by classical two-sided PCR requires a minimum of 45 amplification reactions for the murine TCR (20 for the V alpha families and 25 for the V beta families), using 45 different V-family-specific amplification primers. cDNA was synthesized from spleen RNA, using oligonucleotides complementary to sequences of either the murine TCR C alpha or C beta regions. The NotI restriction site was conjugated to these primers and therefore, a NotI restriction site was incorporated at the 3' end of the cDNA. A double-stranded nonpalindromic adaptor (EcoRI-XmnI strand and XmnI G strand, which are complementary to each other) was ligated onto both ends of the double-stranded cDNA. The adaptor was removed from the 3' end by NotI nuclease digestion whereas the adaptor was retained at the 5' end. Two rounds of PCR amplification were carried out. In the first, the EcoRI-XmnI adaptor was used as 5' end amplification primer; an antisense C region primer, designated mC alpha 2 or mC beta 2 (for the alpha- and beta-chain, respectively), was used as 3' amplification primer. In the second round of PCR amplification the same 5' end primer and a 3' end antisense primer, designated mC alpha 1 or mC beta 1, were used. These mC alpha 1 and mC beta 1 primers are located 5' to the mC alpha 2/mC beta 2 primers that were used for the first amplification. The amplified transcripts were cloned. Colonies were screened using a 32P-labeled probe, either C alpha or C beta, located 5' to those used for the last amplification and many positive clones were isolated and sequenced. All clones were unique when compared to each other, as anticipated for polyclonal T-cell populations. Comparison of the sequences obtained to those in the GENBANK/EMBL database revealed that they were typical of mouse alpha- or beta-chain TCR. With the exception of two beta-chain TCR transcripts, all the sequences shown here (36 alpha-chain and 20 beta-beta chain) have not been previously reported to the GENBANK/EMBL database.

Amino Acid Sequence↗

PubMed1997

Changes in serum tumour necrosis factor-alpha in burned patients.

Dynamic tumour necrosis factor-alpha (TNF-alpha) changes in serial serum samples of 10 burned patients were analysed in this study. The total body surface areas (TBSA) of the burn injury were from 30 to 85 per cent. Among these 10 patients, five recovered and another five died with proved sepsis. On admission which was about 5-13 h postburn, eight of the 10 patients showed their serum TNF-alpha levels to be higher than the mean serum TNF-alpha value of five healthy laboratory personnel. Furthermore, an initial peak serum TNF-alpha response which could be detected within 2.5 days after burn injury has also been observed. However, significant differences in both the serum TNF-alpha values on admission, as well as the first peak serum TNF-alpha levels, were not found (P > 0.05) between patients with TBSA of greater or less than 50 per cent and patients who survived or died from burn injury. In the survivors, serum TNF-alpha stayed at low levels, while it increased markedly in four of the five non-survivors with proven sepsis starting at about 1 week postburn. A significant difference in the maximum serum TNF-alpha levels (P < 0.05) was detected between patients who recovered and died from the thermal injury. In conclusion, great increases in serum TNF-alpha levels have been detected in burned patients with the occurrence of bacterial infection postburn. It is suggested that strategies for the inhibition of TNF-alpha production or in the neutralization of TNF-alpha activity should also be considered in the better treatment of burned patients.

Adolescent↗

PubMed1997

Adaptive control of arterial blood pressure with a learning controller based on multilayer neural networks.

We discuss a two-model multilayer neural network controller for adaptive control of mean arterial blood pressure (MABP) using sodium nitroprusside. A model with an autoregressive moving average (ARMA), representing the dynamics of the system, and a modified back-propagation training algorithm are used to design the control system to meet specified objectives of design (settling time and undershoot/overshoot) and clinical constraints. The controller is associated with a weighting-determinant unit (WDU) to determine and update the output weighting factor of the parallel two-model neural network for adequate control action and a control-signal modification unit (CMU) to comply with clinical constraints and to suppress the effect of adverse noise and to improve the WDU performance. Extensive computer simulations indicate satisfactory performance and robustness of the proposed controller in the presence of much noise, over the full range of plant parameters, uncertainties, and large variations of parameters.

Adaptation, Physiological↗

PubMed1997

Characterization of the 33-kilodalton major allergen of Penicillium citrinum by using MoAbs and N-terminal amino acid sequencing.

BACKGROUND: The 33 kD component has been identified as a major allergen of Penicillium citrinum, the most prevalent Penicillium species in the Taipei area of Taiwan. OBJECTIVE: This study analyses the isoforms, antigenic cross-reactivity and the N-terminal amino acid sequence of the 33 kD allergen of P. citrinum. METHODS: The composition of isoforms and antigenic cross-reactivity was analysed by SDS-PAGE and 2D-immunoblotting using MoAbs generated. The N-terminal sequence was analysed by using an automatic gas/liquid phase sequencer. RESULTS: Two MoAbs (55A and 34H) against the 33 kD allergen were generated in the present study. In addition to the 33 kD component, MoAb 34H also showed immunoblot reactivity to other components in the crude extract of P. citrinum. Analysed by 2D-immunoblotting, at least six different isoforms of the 33 kD component with pI values ranging from 6.75 to greater than 7.0 were shown to be reactive to both MoAbs and IgE antibodies in serum of an asthmatic patient. Different immunoblot patterns were observed when both MoAbs were reacted with four different strains of P. citrinum used in the present study. Among another six different Penicillium and four different Aspergillus species tested, only an immunoblot reactivity of MoAb 55A to the 33 kD component of P. brevicompactum was observed. In 2D-immunoblotting, components of P. brevicompactum with an MW of about 33 kD and pI values similar to those of the 33 kD component of P. citrinum reacted with MoAb 55A and IgE antibodies in serum of the asthmatic patient. The N-terminal amino acid sequence of the 33 kD component of P. citrinum was determined to be ANVVQSNVP which was identical to the first 9 N-terminal amino acids of a heat-labile alkaline serine proteinase from P. citrinum. CONCLUSION: Results obtained in the present study suggest that the 33 kD major allergen of P. citrinum may be an alkaline serine proteinase.

Allergens↗

PubMed1997

Molecular cloning and expression of a Penicillium citrinum allergen with sequence homology and antigenic crossreactivity to a hsp 70 human heat shock protein.

BACKGROUND: Penicillium citrinum has been identified as the most prevalent airborne Penicillium species in the Taipei area. However, detailed studies on allergens of this ubiquitous Penicillium species are still lacking. OBJECTIVE: For the characterization of allergens of this prevalent Penicillium species, molecular cloning and expression of the allergen genes of P. citrinum were performed in the present study. METHODS: Molecular cloning of the allergen genes was performed by using a lambda Uni-Zap XR cDNA library of P. citrinum and serum from an asthmatic patient. The cloned cDNA was excised from the phage vector as a recombinant pBluescript phagemid and sequenced. The cDNA of the IgE-binding clone was expressed as fusion protein with the glutathione-S-transferase. The corresponding natural allergen was analysed by absorption immunoblotting using monoclonal antibody and serum from asthmatic patient. The frequency of IgE-binding to the allergen cloned was analysed by dot immunoassay using recombinant allergen and by immunoblotting using the whole extract of P. citrinum. RESULTS: In the screening of cDNA library of P. citrinum using serum from an asthmatic patient, IgE-binding cDNA clones designated SC4 and XL were obtained. The 5'-truncated, 0.7-kb and 1.7-kb cDNA inserts of clones SC4 and XL contained open reading frames of 163 and 503 amino acids, respectively. On alignment, the deduced amino acid sequences showed that 97 (60%) of the 163 amino acids and 376 (75%) of the 503 amino acids were identical to the corresponding amino acid sequence of the human heat shock protein in the hsp70 family. Both recombinant SC4 and XL showed positive SDS-PAGE-immunoblot reactivity to a monoclonal antibody MA3-006 against the human hsp 70 protein. For characterization of the corresponding natural allergen, immunoblotting reactivities of MA3-006 and IgE antibodies to the 70 kDa component of P. citrinum have been shown to be disappeared after absorption of these antibodies with the recombinant SC4 protein. Sera from 14 (41%) of 34 Penicillium-allergic patients showed IgE-binding to the recombinant XL protein and the 70 kDa component in the extract of P. citrinum. CONCLUSION: Results obtained suggest that hsp 70 is an allergen of P. citrinum and that clones SC4 and XL contain partial cDNAs of this allergen gene.

Allergens↗

PubMed1997

Characterization of the allergen Der f 7 from house dust mite extracts by species-specific and crossreactive monoclonal antibodies.

BACKGROUND: The group 7 mite allergens react with IgE in 50% of sera from allergic patients. OBJECTIVE: To determine the molecular and antigenic characteristics and heterogeneity of Der f 7 in mite extracts. METHODS: Monoclonal antibodies (MoAbs) produced from mice immunized with recombinant Der f 7 were examined for crossreactivity to Der p 7 and then used for immunoblotting of 1 and 2-D gel electrophoresis. Deglycosylation was studied with N-glycosidase-F and N-terminal sequencing by Edman degradation. The epitopes of the monoclonal antibodies were compared by cross-inhibitory immunoassays. RESULTS: Immunoblotting of D. farinae extracts with all the anti Der f 7 MoAbs showed major reactivities at 31, 30 and 25 kDa. The strongest immunostaining was at 25 kDa which contrasted with Der p 7 where the 31 and 30 kDa bands were strongest. The relative strength of staining however varied between extracts. The 31 and 30 kDa components were glycosylation products of the 25 kDa form which had the N-terminal sequence predicted from cDNA analysis. Two MoAbs stained an 18 kDa band consistent with a degradation product. The 2-D gels showed that different components with pIs from 5.6-6.4. Both species-specific and Der p 7 crossreactive MoAbs were produced and a two-site ELISA assay for detecting group 7 allergen was developed with MoAbs recognizing different epitopes. CONCLUSIONS: Der f 7 has been defined by its natural N-terminal sequence and MoAbs. It apparently exists as different glycosylation and degradation products in mite extracts, the relative abundance of which differs with different preparations. A two-site ELISA to measure the allergen was developed.

Allergens↗

PubMed1996

L ferritin accumulation in macrophages infiltrating the lung during rat Angiostrongylus cantonensis infection.

A 22-kDa protein was increased quantitatively, as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis techniques, in lung microsomes prepared from Angiostrongylus cantonensis-infected rats. However, it was almost absent in normal rats. The protein was purified by sequential chromatography on Superdex 200 columns and identified chemically and immunologically as ferritin. Using isoelectric focusing and anion exchange chromatography, it was identified as L ferritin. Distribution of this 22-kDa protein in the lung tissue of A. cantonensis-infected rate was studied by immunocytochemistry. Positively stained cells were mainly infiltrated macrophages. Our results suggest that L ferritin accumulation in the macrophages may be related to the proliferation of connective tissue elements and the inflammatory response to A. cantonensis dwelling in the pulmonary arteries of the rat.

Angiostrongylus cantonensis↗

PubMed1996

IgE and monoclonal antibody binding by the mite allergen Der p 7.

BACKGROUND: The recently characterized group 7 house dust mite allergens give positive skin-test reactions in 53% of allergic patients. OBJECTIVE: This study was performed to compare the IgE binding activity of natural and recombinant Der p 7, to measure the binding in allergic sera in comparison to major allergen Der p 2 and characterize the response by competitive inhibition with monoclonal antibodies. METHODS: IgE anti Der p 2 and Der p 7 antibodies against the recombinant allergens and monoclonal binding activities were measured by a solid phase radioimmune assay. RESULTS: A competitive binding assay showed that rDer p 7 inhibited 91% of IgE-binding to natural Der p 7 in 2 sera and 73% in a further two. The IgE binding of rDer p 2 and Der p 7 from 41 sera was then compared. Of the sera 88% and 46% respectively showed positive binding. All of the 19 sera which bound Der p 7 also bound Der p 2 but 11 (58%) had bound IgE to Der p 7 as high or higher than the binding to Der p 2. These sera were mostly high responders to both allergens. A panel of six monoclonal antibodies produced against either rDer p 7 or rDer f7 was used for epitope analysis. All of these reacted with each allergen by enzyme linked immunosorbent assay (ELISA) and immunoblotting. Two patterns of cross inhibition of monoclonal antibody binding were observed and of five monoclonal antibodies tested, four could inhibit the binding of IgE (WH9, WH22, WP8 and HD19) while one (WH14) could not. CONCLUSIONS: Although Der p 7 only reacts with 50% of allergic sera it often has a high IgE binding activity and may be more important than the major Der p 2 allergen in a high percentage of subjects. The combined competitive inhibition experiments show the IgE response is directed at several specificities.

Antibodies, Monoclonal↗

PubMed1996

Allergenic components in three different species of Penicillium: crossreactivity among major allergens.

BACKGROUND: Penicillium species have been considered as important causative agents of extrinsic bronchial asthma. However, little is known about the allergens of these ubiquitous airborne fungal species. OBJECTIVE: This study compares the allergenic profiles and allergenic crossreactivity among allergens of three prevalent airborne Penicillium species. METHODS: IgE-binding Penicillium components were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE)-immunoblotting using sera from 67 asthmatic patients. The presence of allergenic crossreactivity was analysed by immunoblot inhibition. RESULTS: Among the 67 serum samples tested, 15, 14 and 11 samples showed IgE reactivity to components of P. citrinum, P. notatum and P. brevicompactum, respectively. All 15 P. citrinum-positive serum samples showed IgE-binding to a 33 kDa extract component of this species. Thirteen (93%) of the 14 P. notatum-positive serum samples and 10 (91%) of the 11 P. brevicompactum-positive sera also showed IgE reactivity to components with a molecular weight of about 33 kDa in individual Penicillium species. All of the 10 P. brevicompactum 33 kDa component-positive serum samples showed IgE reactivity to the 33 kDa components of the other two Penicillium species tested. Dose-dependent inhibition of IgE-binding to these major allergens was observed when the positive serum sample was absorbed with different amounts of individual allergenic extract as well as with different amounts of extracts prepared from the other two Penicillium species. CONCLUSION: Although different allergenic profiles were observed in the three different Penicillium species tested, results showed that there was an IgE crossreactivity among the 33 kDa group major allergens of P. citrinum, P. notatum and P. brevicompactum.

Allergens↗

PubMed1995

Synthesis of noncollagenous extracellular matrix proteins during development of mineralized nodules by rat periodontal ligament cells in vitro.

To characterize the mineralized nodules produced by rat periodontal ligament (PDL) cells in vitro, we have studied the synthesis and distribution of mineralized tissue proteins at various stages of nodule formation. PDL cells were obtained from coagulum in the socket at 2 days after tooth extraction and cultured in Dulbecco's Modified Eagles Medium (DMEM) containing 10% fetal bovine serum and antibiotics. Confluent cells were grown in the presence of ascorbic acid (50 micrograms/ml), dexamethasone (5 microM), and beta-glycerophosphate (10 mM) for 3 weeks. Four stages showing distinct morphological characteristics during development of mineralized nodules were identified. Protein synthesis and deposition of proteins into the matrix were studied during these stages by metabolic labeling with [35S]methionine for 24 hours. Large quantities of SPARC (secreted protein, acidic and rich in cysteine) were synthesized by confluent cells but decreased during the progress of mineralized nodule formation. Two forms of osteopontin (OPN) (67 kDa and 61 kDa) were synthesized in comparable quantities by confluent cells; OPN and bone sialoprotein (BSP) were induced by dexamethasone and represented the major proteins in the mineralized matrix. The 67 kDa form of OPN was the predominant species in the mineralized matrix. Both OPN and BSP were localized by immunogold electron microscopy on globular as well as fused electron-dense structures at sites of tissue mineralization.

Animals↗

PubMed1995

Molecular cloning and immunological characterization of the house dust mite allergen Der f 7.

BACKGROUND: The allergen Der p 7 from Dermatophagoides pteronyssinus has been defined by molecular cloning and shown to be an important specificity in 50% of mite-allergic patients. OBJECTIVE: This study compares the cDNA sequence and serological reactivity of Der f 7 from D. farinae with Der p 7. METHOD: cDNA encoding Der f 7 was amplified by polymerase chain reaction, sequenced and expressed as a fusion with glutathione-S-transferase for IgE and monoclonal antibody binding studies. RESULTS: Der f 7 cDNA encoded a 213 polypeptide containing a predicted 17 amino acid leader sequence, no cysteines and a single N-glycosylation site similar to Der p 7. The predicted 196 residue mature polypeptide had 86% identity to Der p 7 and a calculated molecular weight of 22,348Da. No homologues were found in searches of the data banks. The Der f 7 fusion protein showed a single band of 46 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and reacted with IgE antibodies in 19/41 (46%) of sera from asthmatic children. The degree of binding was usually 30% of that to Der p 7 consistent with the exposure of the patients to D. pteronyssinus. Monoclonal antibodies (WH9 and WH22) against Der p 7 reacted with Der f 7 but inhibition studies showed a 10-fold difference in reactivity. CONCLUSION: Der f 7 has a predicted 213 residue polypeptide with 86% homology and serological crossreactivity to Der p 7.

Allergens↗