Development of a device for cultivation and isolation of microbes using a specialized cellulose film.

A device was developed for the cultivation and isolation of microbes using a specialized cellulose film (SCF). This method can solve problems associated with the agar plate method. It can also be used for other culture-related operations, which is not possible with the traditional agar plate culture method.

A variant of death-receptor 3 associated with rheumatoid arthritis interferes with apoptosis-induction of T cell.

Rheumatoid arthritis (RA) is a chronic polyarthritis of unknown etiology. To unravel the molecular mechanisms in RA, we performed targeted DNA sequencing analysis of patients with RA. This analysis identified a variant of the death receptor 3 (DR3) gene, a member of the family of apoptosis-inducing Fas genes, which contains four single-nucleotide polymorphisms (SNPs) and a 14-nucleotide deletion within exon 5 and intron 5. We found that the deletion causes the binding of splicing regulatory proteins to DR3 pre-mRNA intron 5, resulting in a portion of intron 5 becoming part of the coding sequence, thereby generating a premature stop codon. We also found that this truncated DR3 protein product lacks the death domain and forms a heterotrimer complex with wildtype DR3 that dominant-negatively inhibits ligand-induced apoptosis in lymphocytes. Myelocytes from transgenic mice expressing the human DR3 variant produced soluble truncated DR3, forming a complex with TNF-like ligand 1A (TL1A), which inhibited apoptosis induction. In summary, our results reveal that a DR3 splice variant that interferes with ligand-induced T cell responses and apoptosis may contribute to RA pathogenesis.

Synthesis, Structure, and Photophysical/Chiroptical Properties of Benzopicene-Based π-Conjugated Molecules.

The convenient synthesis of substituted benzopicenes and azabenzopicenes has been achieved by the cationic rhodium(I)/H8-BINAP or BINAP complex-catalyzed [2+2+2] cycloaddition under mild conditions. This method was applied to the synthesis of benzopicene-based long ladder and helical molecules. The X-ray crystal structure analysis revealed that the benzopicene-based helical molecule is highly distorted and the average distance of overlapped rings is markedly shorter than that in the triphenylene-based helical molecule. Photophysical and chiroptical properties of these benzopicene and azabenzopicene derivatives have also been examined. With respect to photophysical properties, substituted benzopicenes and azabenzopicenes showed red shifts of absorption and emission maxima compared with the corresponding triphenylenes and azatriphenylenes. With respect to chiroptical properties, the CPL spectra of the benzopicene-based helical molecule showed two opposite peaks, and thus the value of the CPL was smaller than that of the triphenylene-based helical molecule presumably due to the presence of two chiral fluorophores.

Rh-Mediated Enantioselective Synthesis, Crystal Structures, and Photophysical/Chiroptical Properties of Phenanthrenol-Based [9]Helicene-like Molecules.

The enantioselective synthesis of phenanthrenol-based [9]helicene-like molecules has been achieved via the rhodium-mediated intramolecular [2 + 2 + 2] cycloadditions of 3-phenanthrenol-linked triynes. Crystal structures and photophysical/chiroptical properties of these [9]helicene-like molecules were compared with the corresponding [7]helicene-like molecules.

Rapid tumor necrosis and massive hemorrhage induced by bevacizumab and paclitaxel combination therapy in a case of advanced breast cancer.

Bevacizumab when combined with chemotherapy exerts significant activity against many solid tumors through tumor angiogenesis inhibition; however, it can induce severe side effects. We report the rare case of a 27-year-old premenopausal woman with locally advanced breast cancer that was marked by rapid tumor necrosis followed by massive hemorrhage shortly after bevacizumab and paclitaxel administration. On the basis of histopathological examination of a biopsy specimen and computed tomography findings, she was diagnosed with stage IV estrogen and progesterone receptor-negative and human epidermal growth factor receptor type 2-positive breast cancer with multiple organ metastases when she had entered gestational week 24. Cyclophosphamide, Adriamycin®, fluorouracil therapy was initiated, but multiple liver metastases continued to progress. A healthy fetus was delivered by induced delivery and trastuzumab-based treatment was initiated. Although the multiple liver metastases were controlled successfully by trastuzumab combined with paclitaxel, the primary tumor continued to expand even after subsequent administration of three other treatment regimens including anti-human epidermal growth factor receptor type 2 agents and cytotoxic drugs. To inhibit primary tumor growth, a combination therapy with paclitaxel and bevacizumab was subsequently initiated. Following therapy initiation, however, the large tumor occupying the patient's entire left breast became necrotic and ulcerated rapidly. Furthermore, massive hemorrhage from the tumor occurred 5 weeks after bevacizumab-based therapy initiation. Although hemostasis was achieved by manual compression, the patient required blood transfusion for the massive blood loss. She eventually succumbed to respiratory failure. This case report demonstrates that primary breast cancer lesions with skin involvement have the potential to cause massive hemorrhage after bevacizumab-based treatment.

Synthesis of triphenylene derivatives by rhodium-catalyzed [2 + 2 + 2] cycloaddition: application to the synthesis of highly fluorescent triphenylene-based long ladder molecules.

The convenient synthesis of substituted triphenylenes and azatriphenylenes has been achieved by the cationic rhodium(I)/H8-BINAP or BINAP complex-catalyzed [2 + 2 + 2] cycloaddition under mild conditions. Photophysical properties of representative triphenylenes and azatriphenylenes were examined, which revealed that azatriphenylenes showed higher fluorescence quantum yields than triphenylenes. This method was successfully applied to the synthesis of highly fluorescent triphenylene-based long ladder molecules.

[A quality of life questionnaire for Japanese allergic conjunctival disease].

PURPOSE: To establish a specific quality of life (QOL) questionnaire for Japanese allergic conjunctival disease (ACD) (Japanese allergic conjunctival disease QOL questionnaire: JACQLQ). SUBJECTS AND METHODS: A multicenter study was conducted in 521 patients with ACD and 127 healthy volunteers (total 648 cases). The JACQLQ ver. 0 was developed by modifying the Japanese rhino-conjunctivitis QOL questionnaire (JRQLQ). The participants were asked to complete the questionnaire, and objective scores were determined by an ophthalmologist using a slit lamp. To confirm validity, item and factor analyses were conducted and correlation coefficients were calculated. RESULTS: The items were grouped into four subscales (Daily activity, Psychological well-being, Eye symptoms, Nasal symptoms) after factor analysis. The JACQLQ had good item-internal consistency (Cronbach's alpha: 0.846-0.934). QOL scores were correlated with eye itching, eye irritation and tearing. Objective scores were correlated with eye redness, eye itching and eye irritation. Face scores were correlated with eye itching, eye irritation and eye redness. CONCLUSION: The JACQLQ is a useful tool for assessing disease specific QOL in ACD.

Multimodality therapeutic outcomes in anaplastic thyroid carcinoma: improved survival in subgroups of patients with localized primary tumors.

BACKGROUND: The aim of the present study was to investigate the role of a multimodality treatment for anaplastic thyroid carcinoma (ATC). METHODS: Clinical data of 40 consecutive patients treated between 1985 and 2009 were retrospectively analyzed. RESULTS: The median survival time (MST) stratified by clinical stage was 6.0 months for stage IVB and 4.2 months for stage IVC. When the stage IVB patients were classified into 2 groups (IVB-a and IVB-b) in accord with the extent of involvement of the tumor, the MST of the patients with IVB-a (9.6 months) was significantly longer than that of patients with IVB-b (4.0 months) (p < .05). The MST of the patients treated with surgery followed by radiation and chemotherapy (13.7 months) tended to be longer compared with the patients treated with 2 or fewer modalities. CONCLUSION: The patients with a less-invasive primary tumor may gain a survival benefit from aggressive multimodality therapeutic approaches.

FcεRI-induced mast cell cytokine production critically involves an aspartic acid residue (D234) in the C-terminal intracellular domain of the FcεRIß chain.

The high affinity IgE Fc receptor (FcεRI) ß chain is well implicated as a signal amplifier through the immunoreceptor tyrosine-based activation motif (ITAM) in its C-terminal intracellular region. Our previous study, however, demonstrated that mutation in all of the three tyrosine residues within the FcεRIß ITAM did not impair FcεRI-induced cytokine production, suggesting a possible functional region other than the ITAM. To investigate the ITAM-independent mechanism by which FcεRIß regulates FcεRI-induced cytokine production, mouse mast cells expressing various FcεRIß mutants were generated. We observed that truncation of the FcεRIß C-terminus downstream of the ITAM resulted in a considerable decrease in FcεRI-induced IL-6 production but not degranulation. Furthermore, mutagenesis of a single C-terminal aspartic acid (D234) to alanine (ß-D234A) also significantly impaired IL-6 production. In addition, the similarity between the circular dichroism (CD) spectra of the wild type and ß-D234A suggests that the secondary structure of the FcεRIß C-terminus was not affected by the D234A mutation. Consistently, we did not observe any effect of this mutation on FcεRI-induced tyrosine phosphorylation of FcεRIß. These observations strongly suggest a novel signaling pathway mediated by the cytoplasmic tail downstream of the FcεRIß ITAM.

CSLEX (Sialyl Lewis X) is a useful tumor marker for monitoring of breast cancer patients.

BACKGROUND: CSLEX is a type II carbohydrate antigen that interacts with the CSLEX-1 monoclonal antibody. CSLEX in combination with carbohydrate antigen 15-3 may be more useful than Carcinoembryonic Antigen with carbohydrate antigen 15-3 as tumor markers for monitoring of breast cancer. METHODS: The serum levels of tumor markers, including CSLEX, were measured in 480 consecutive breast cancer patients with or without metastasis who visited the outpatient clinic of the Division of Breast and Endocrine Surgery, Shinshu University Hospital, between April 2007 and September 2007. RESULTS: Serum levels of each of the tumor markers correlated significantly with the status of metastasis (P < 0.01). Combinations of Carcinoembryonic Antigen and carbohydrate antigen 15-3, Carcinoembryonic Antigen and Nation Cancer Center-Stomach-439, Carcinoembryonic Antigen and CSLEX, carbohydrate antigen 15-3 and Nation Cancer Center-Stomach-439, and carbohydrate antigen 15-3 and CSLEX levels also correlated significantly with the status of metastasis (P < 0.01). The sensitivity, specificity, positive predictive value, negative predictive value and accuracy were almost the same for CSLEX and Nation Cancer Center-Stomach-439, which are both type II carbohydrate antigens. The cutoff indexes of serum CSLEX and Nation Cancer Center-Stomach-439 for detection of breast cancer metastasis were 38.8 ± 52.7-fold and 22.1 ± 27.8-fold, respectively (P = 0.16). CONCLUSIONS: These data suggest that the diagnostic values of CSLEX and Nation Cancer Center-Stomach-439 are similar in single or combined use. However, the cutoff index of serum CSLEX tended to be higher than that of Nation Cancer Center-Stomach-439, which may make CSLEX more useful for detection of breast cancer metastasis.

Full-thickness chest-wall resection followed by thorax reconstruction for recurrent malignant phyllodes tumor.

We present a case of a 39-year-old woman with a giant recurrent malignant phyllodes tumor accompanied with bleeding and infection. She underwent full-thickness chest-wall resection. Bony thorax reconstruction and stabilization was accomplished using a Composix mesh™, and soft tissue reconstruction was performed with a musculocutaneous flap of latissimus dorsi muscle. The patient had a good postoperative outcome, and the surgical treatment remarkably improved her quality of life. Because chemotherapy and radiation are not established for treating malignant phyllodes tumors, an aggressive surgical approach should be considered for patients with a locally advanced malignant phyllodes tumor.

Study of thermal dynamics of defatted bovine serum albumin in D2O solution by Fourier transform infrared spectra and evolving factor analysis.

Fourier transform infrared (FT-IR) spectra have been measured for defatted bovine serum albumin (BSA) in D(2)O with a concentration of 2.0 wt % over a temperature range of 26-90 degrees C and the corresponding difference spectra have been calculated by subtracting the contribution of D(2)O at the same temperature. Evolving factor analysis (EFA) by selecting two factors and three factors has been employed to analyze the temperature-dependent difference IR spectra in the 1700-1600 cm(-1) spectral region of the defatted BSA in D(2)O solution. Three-factor EFA has been employed to determine the distinction of the three protein species involved in the process of temperature elevation: native, transitional, and denatured protein. The temperature profiles obtained from three-factor EFA indicate that heat-induced conformational change in the secondary structures of defatted BSA in D(2)O undergoes two two-state transitions, a drastic transition and a slight transition, which occur in the temperature ranges of 68-82 degrees C and 56-76 degrees C, respectively.

An infrared spectroscopy study of acid stability and thermal unfolding process of granulocyte-colony stimulating factor.

Temperature-dependent (25-80 degrees C) infrared (IR) spectra were obtained for recombinant methionyl human granulocyte-colony stimulating factor (rmethuG-CSF) in aqueous solutions over the pD range of 5.5-2.1 to investigate its thermal stability at various pDs. Second derivative, Fourier self-deconvolution, and curve-fitting analyses were performed to analyze the obtained spectra. These spectral analyses demonstrated that in the thermal unfolding process the alpha-helix structure of rmethuG-CSF partially changes to an unordered structure and then the unordered structure forms aggregates. The temperature-dependent IR spectra revealed that the structure of rmethuG-CSF is the most stable at pD 2.5 in the pD range of 5.5-2.1. It has been suggested that the unordered structure formed before the marked structural change in the whole molecule is a perturbed form of the native structure of rmethuG-CSF and plays a role as a precursor for the aggregation. This alteration to the perturbed form is likely to be the first secondary structure change that occurs along the aggregation pathway. Of particular note is that the stability at pD 2.1 is slightly lower than that at pD 2.5, but that aggregates are formed at higher temperature at pD 2.1 than at pD 2.5, probably because the repulsive interaction between the unordered structure is stronger at pD 2.1.

Heat-induced secondary structure and conformation change of bovine serum albumin investigated by Fourier transform infrared spectroscopy.

Fourier transform infrared (FT-IR) spectra were measured for an aqueous solution (pD = 5.40) of defatted monomer bovine serum albumin (BSA) over a temperature range of 25-90 degrees C to investigate temperature-induced secondary structure and conformation changes. The curve fitting method combined with the Fourier self-deconvolution technique allowed us to explore details of the secondary structure and conformation changes in defatted BSA. Particularly striking in the FT-IR spectra was an observation of the formation of an irreversible intermolecular beta-sheet of BSA on heating above 70 degrees C. A band at 1630 cm(-1) in the spectra was assigned to short-segment chains connecting alpha-helical segments. The transition temperature for the short-segment chains connecting alpha-helical segments is lower by 17-18 degrees C, when compared to those of the alpha-helix, turn, and intermolecular beta-sheet structures of BSA, suggesting that the alpha-helix and turn structures of BSA are cooperatively denatured on heating. Moreover, the results give an important feature in heat-induced denaturation of BSA that the conformation changes occur twice around both 57 and 75 degrees C. The appearance of two peaks is interpreted by the collapse of the N-terminal BSA domain due to the crevice in the vicinity between domains I and II at low-temperature transition and by the change in cooperative unit composed of the other two BSA domains at high-temperature transition.

Temperature-dependent near-infrared spectra of bovine serum albumin in aqueous solutions: spectral analysis by principal component analysis and evolving factor analysis.

Fourier transform near-infrared (FT-NIR) spectra have been measured for bovine serum albumin (BSA) in an aqueous solution (pH 6.8) with a concentration of 5.0 wt% over a temperature range of 45-85 degrees C. Not only conventional spectral analysis methods, such as second-derivative spectra and difference spectra, but also chemometrics, such as principal component analysis (PCA) and evolving factor analysis (EFA), have been employed to analyze the temperature-dependent NIR spectra in the 7500-5500 and 4900-4200 cm-1 regions of the BSA aqueous solution. Intensity changes of bands in the 7200-6600 cm-1 and 4650-4500 cm-1 regions in the difference spectra indicate variations of the hydration and secondary structure of BSA in the aqueous solution, respectively. The plot of a band intensity at 7080 cm-1 in the different spectra shows a clear turning point at 63 degrees C, revealing that a significant change in the hydration occurs at about 63 degrees C. The forward and backward eigenvalues (EVs) from EFA suggest that marked changes in the hydration and secondary structure of BSA take place in the temperature ranges of 61-65 degrees C and 59-63 degrees C, respectively. In addition, the temperature of 71 degrees C marked in the EFA plots may correspond to the onset temperature of increase in the intermolecular beta-sheet structure.

Near-infrared spectroscopy for liquids of microliter volume using capillaries with wall transmission.

In the present study we propose a capillary tube method for measuring near-infrared (NIR) spectra of microliter liquid and solution samples. This method enables one to measure NIR spectra of liquids and solutions of only 2.5 microl. As an example of the capillary tube method, Fourier-transform NIR spectra of benzene were measured using a capillary tube with a diameter of 1 mm. Positions and intensities of the NIR bands in the spectra obtained by the capillary tube were almost identical to those measured using a quartz cuvette cell. Moreover, capillary NIR spectra of human blood serum obtained from a patient with backbone neoplasm were also measured. Subtraction of the NIR spectrum of water from those of human blood serum develops unambiguously some NIR bands due to blood components such as proteins and glucose. Our results suggest that capillary NIR spectroscopy will open new areas of NIR applications for small amounts of liquid and solution samples.

The redox states of serum and synovial fluid of patients with temporomandibular joint disorders.

The redox states of albumin in serum and synovial fluid (SF) were analyzed in 35 female patients with temporomandibular joint disorders and 9 asymptomatic female healthy volunteers by high-performance liquid chromatographic (HPLC) analysis. Human serum albumin (HSA) is divided by HPLC analysis into three fractions: mercaptalbumin (HMA, reduced form) and nonmercaptalbumin (HNA-1 and HNA-2, oxidized forms). It was found that the characterized values of albumin in SF of patients with the disorders were the fraction of HMA (f (HMA), 65.8 +/- 9.1%), the fraction of HNA-1 (f (HNA-1), 31.3 +/- 8.8%), and the fraction of HNA-2 (f (HNA-2), 2.9 +/- 0.9%), respectively, and similarly the values of albumin in serum of same subjects were f (HMA), 79.4 +/- 4.0%; f (HNA-1), 18.7 +/- 3.8%; and f (HNA-2), 1.9 +/- 0.5%. The HNA fractions in SF were significantly higher than those in serum (p < 0.0001). The SF of patients and even control showed more oxidative condition compared with sera of the same subjects. Each fraction of SF showed significant difference in control (p < 0.05), and greater significance in patients (p < 0.0001). Furthermore, the HNA-2 fraction (f (HNA-2), 2.9 +/- 0.9%) in SF of the patients with temporomandibular joint disorders revealed a significantly higher value (p < 0.01) than that (f (HNA-2), 1.7 +/- 0.3%) of controls. These findings suggested that SF of the temporomandibular joint is more locally oxidized than serum; thus oxidative factors are concerned in the pathogenesis of the temporomandibular joint disorders. It was also suggested that synovial albumin may play a scavenging role against the intraarticular oxidative stress.

Simultaneous determination of human serum albumin, gamma-globulin, and glucose in a phosphate buffer solution by near-infrared spectroscopy with moving window partial least-squares regression.

Near-infrared (NIR) spectra in the 12,000-4,000 cm(-1) region were measured for phosphate buffer solutions containing human serum albumin (HSA), gamma-globulin, and glucose with various concentrations at 37 degrees C. Five levels of full factorial design were used to prepare a sample set consisting of 125 samples of three component mixtures. The concentration ranges of HSA, gamma-globulin and glucose were 0.00-6.00 g dl(-1), 0.00-4.00 g dl(-1) and 0.00-2.00 g dl(-1), respectively. The 125 sample data were split into two sets, the calibration set with 95 data and the prediction set with 30 data. The most informative spectral ranges of 4648-4323, 4647-4255 and 4912-4304 cm(-1) were selected by moving window partial least-squares regression (MWPLSR) for HSA, [gamma]-globulin, and glucose in the mixtures, respectively. For HSA, the correlation coefficient (R) of 0.9998 and the root mean square error of prediction (RMSEP) of 0.0289 g dl(-1) were obtained. For [gamma]-globulin, R of 0.9997 and RMSEP of 0.0252 g dl(-1) were obtained. The corresponding statistic values of glucose were 0.9997 and 0.0156 g dl(-1), respectively. These statistical values obtained by MWPLSR are highly significant and better than those calculated by using the regions reported in the literature. The results presented here show that MWPLSR can select the informative regions with a simple procedure and increase the power of NIR spectroscopy for simultaneous determination of the concentrations of HSA, [gamma]-globulin and glucose in the mixture systems.

Two-dimensional near-IR correlation spectroscopy study of molten globule-like state of ovalbumin in acidic pH region: simultaneous changes in hydration and secondary structure.

The molten globule-like states of ovalbumin (OVA) in acid aqueous solutions are investigated by generalized two-dimensional (2D) Fourier transform near-IR (FT-NIR) correlation spectroscopy. This new method allows us to explore the changes in hydration and the secondary structure simultaneously. FT-NIR spectra are measured for OVA aqueous solutions with concentrations of 1, 2, 3, 4, and 5 wt % over a pH range of 2.4-5.4. Concentration-perturbed 2D correlation spectra are calculated for the spectra in the 4850-4200 and 7500-5350 cm(-1) regions at different pH values. The 2D NIR synchronous spectrum in the 4850-4200 cm(-1) region shows a significant change upon going from pH 5.4 to 3.6. An autopeak at 4265 cm(-1) that is due to a combination of a symmetric CH(2) stretching mode and a CH(2) bending mode of side chains seen at pH 5.0 disappears completely in the synchronous spectrum at pH 3.6. This suggests that some amino acid residues of OVA are subjected to microenvironmental changes with decreasing pH. More remarkable changes are observed in the synchronous spectra at pHs below 2.8. A band near 4600 cm(-1) arising from a combination of amide B and amide II modes (amide B/II) shifts downward with considerable broadening between pH 3.0 and 2.4, suggesting that the strength of the hydrogen bonds of amide groups of OVA changes significantly. The synchronous and asynchronous spectra in the 4850-4200 cm(-1) region show that the intensities of the bands attributable to amide groups and side chains of OVA and that of the band near 4800 cm(-1) arising from water change in phase with the increase in the concentration above pH 2.8, but they vary out of phase below pH 2.8. The 2D synchronous map in the 7500-5350 cm(-1) region also shows marked changes upon going from pH 2.8 to 2.6. A broad autopeak at around 6950 cm(-1) assigned to free water and bound water with weak hydrogen bonds becomes very weak in the synchronous spectrum at pH 2.6, while broad autopeaks around 6450 cm(-1) suddenly appear that are due to bound water with several hydrogen bonds and the first overtone of an NH stretching mode of the amide groups of OVA. Therefore, it is very likely that protein hydration and the hydrogen bonds of amide groups change simultaneously in a narrow pH region of 2.8-2.6. It is probably that below pH 2.6 the protein assumes a molten globule-like state in which the whole molecule is very flexible, and side chains (but not the backbone chain) fluctuate significantly.