Detection of underlying characteristics of nuclear chromatin patterns of thyroid tumor cells using texture and factor analyses

BACKGROUND: Aspiration biopsy cytology of thyroid tumors has been used more frequently in recent times to differentiate between malignant and benign lesions. Chromatin patterns of the tumor cell nuclei are one of most important factors for cytologic diagnosis. The interpretation of nuclear chromatin patterns is subjective and more difficult than that of nuclear size or shape. In the present report, we investigated how to detect underlying chromatin characteristics of benign and malignant thyroid tumor cells by means of texture and factor analyses. METHODS: We employed a computer-aided system in which light microscopy was combined with an image processor and monochrome camera. Using this system, 100 randomly selected cells in a Papanicolaou stained, aspiration biopsy cytologic smear in each case of 39 benign and malignant thyroid tumor cases were digitized. We applied two-dimensional and higher texture analyses with the use of co-occurrence and run-length matrices to analyze the chromatin patterns. Factor analysis was used to determine whether a large number of independent variables actually measured one or more underlying common variables. RESULTS: According to parameters with high factor-loading values, the morphologic chromatin characters were classified into three categories according to heterogeneity, contrast, and homogeneity of chromatin patterns. On the basis of analyses with these morphologic categories, nuclei of papillary carcinoma showed higher contrast of chromatin patterns than did those of the benign group. Moreover, there was a variety of contrasting chromatin patterns among cells in each papillary carcinoma case in comparison with the benign group. In contrast, follicular carcinomas showed a significant difference in the standard deviation of factor 3, which indicated more monotonous chromatin patterns among cells in each follicular carcinoma case than in each benign case. CONCLUSION: We believe that this technique, using texture and factor analyses, is useful in the detection of underlying characteristics of nuclear chromatin patterns in aspiration biopsy cytology.     

Trypanosome alternative oxidase, a potential therapeutic target for sleeping sickness, is conserved among Trypanosoma brucei subspecies

Trypanosoma brucei rhodesiense and T. b. gambiense are known causes of human African trypanosomiasis (HAT), or “sleeping sickness,” which is deadly if untreated. We previously reported that a specific inhibitor of trypanosome alternative oxidase (TAO), ascofuranone, quickly kills African trypanosomes in vitro and cures mice infected with another subspecies, non-human infective T. b. brucei, in in vivo trials. As an essential factor for trypanosome survival, TAO is a promising drug target due to the absence of alternative oxidases in the mammalian host. This study found TAO expression in HAT-causing trypanosomes; its amino acid sequence was identical to that in non-human infective T. b. brucei. The biochemical understanding of the TAO including its 3 dimensional structure and inhibitory compounds against TAO could therefore be applied to all three T. brucei subspecies in search of a cure for HAT. Our in vitro study using T. b. rhodesiense confirmed the effectiveness of ascofuranone (IC₅₀ value: 1 nM) to eliminate trypanosomes in human infective strain cultures.     

Protective role of hydroxysteroid sulfotransferase in lithocholic acid-induced liver toxicity

Supplement of 1% lithocholic acid (LCA) in the diet for 5-9 days resulted in elevated levels of the marker for liver damage aspartate aminotransferase and alkaline phosphatase activities in both farnesoid X receptor (FXR)-null and wild-type female mice. The levels were clearly higher in wild-type mice than in FXR-null mice, despite the diminished expression of a bile salt export pump in the latter. Consistent with liver toxicity marker activities, serum and liver levels of bile acids, particularly LCA and taurolithocholic acid, were clearly higher in wild-type mice than in FXR-null mice after 1% LCA supplement. Marked increases in hepatic sulfating activity for LCA (5.5-fold) and hydroxysteroid sulfotransferase (St) 2a (5.8-fold) were detected in liver of FXR-null mice. A 7.4-fold higher 3alpha-sulfated bile acid concentration was observed in bile of FXR-null mice fed an LCA diet compared with that of wild-type mice. Liver St2a content was inversely correlated with levels of alkaline phosphatase. In contrast, microsomal LCA 6beta-hydroxylation was not increased and was in fact lower in FXR-null mice compared in wild-type mice. Clear decreases in mRNA encoding sodium taurocholate cotransporting polypeptide, organic anion transporting polypeptide 1, and liver-specific organic anion transporter-1 function in bile acid import were detected in LCA-fed mice. These transporter levels are higher in FXR-null mice than wild-type mice after 1% LCA supplement. No obvious changes were detected in the Mrp2, Mrp3, and Mrp4 mRNAs. These results indicate hydroxysteroid sulfotransferase-mediated LCA sulfation as a major pathway for protection against LCA-induced liver damage. Furthermore, Northern blot analysis using FXR-null, pregnane X receptor-null, and FXR-pregnane X receptor double-null mice suggests a repressive role of these nuclear receptors on basal St2a expression.     

Fatty Acids as Inhibitors of Microbial Cell Wall Synthesis

The Maillard reaction of DNA with ketoses was investigated. Several days of incubation of d-fructose 6-phosphate with deoxyribonucleotides or with polymer DNA in an aqueous buffer resulted in the formation of chromophores and fluorophores. Aminoguanidine and sodium cvanoborohydride inhibited the formation of fluorophores. Transition metal ions such as Cu²⁺, Fe³⁺, Fe²⁺, or Mn^{2 +} promoted the formation of chromophores and fluorophores. Metal-chelating agents such as DETAPAC, citrate, and Desferal inhibited the formation of fluorophores. Superoxide dismutase and catalase also inhibited the formation of fluorophores. The transition metal ion-catalyzed autoxidation of d-fructose 6-phosphate or of the Heyns rearrangement products were to be partially involved in the glycation of DNA and subsequent formation of chromophores and of fluorophores.     

Modification of regression of virally xenogenized tumor cells by cyclophosphamide and busulfan

Summary Rat fibrosarcoma cells infected with Friend leukemia virus (FV-KMT-17) grow for a short time and then regress spontaneously in syngeneic hosts. This regression mechanism was examined by analyzing the immunomodulating action of the antitumor drugs busulfan (BU) and cyclophosphamide (CY). In preliminary experiments, the optimum dosages of BU and CY for the enhancement of DTH responses to SRBC were 10 mg/kg and 40 mg/kg respectively. Treatment of rats with BU (10 mg/kg) on day 5 induced the regression of KMT-17 cells, while in contrast, the same drug delayed the spontaneous regression of FV-KMT-17 cells. Pretreatment with CY (40 mg/kg) on day 5 did not affect the growth of KMT-17 or FV-KMT-17 cells. After the same treatment schedule, BU inhibited humoral antibody formation against SRBC and against virus-associated antigen (VAA), NK cell activity, and ADCC effector cell activity. On the other hand, CY did not affect the activities of NK cells or ADCC effector cells, although it significantly augmented the DTH responses to SRBC and the production of antibody to VAA but had no effect on production of antibodies to SRBC. These results suggest that NK cells and ADCC may play an important role in the initial stage of the spontaneous regression of FV-KMT-17 cells.Supported in part by a Grant-in-Aid for Cancer Research from the Japanese Ministry of Education Abbreviations used: BU, busulfan; CY, cyclophosphamide; PFC assay, plaque forming cell assay; VAA, virus-associated antigen; NK cell, natural killer cell; ADCC, antibody dependent cellular cytotoxicity; MuLV, murine leukemia virus; DTH, delayed type hypersensitivity; SRBC, sheep red blood cells; C.I., cytotoxic index; CRBC, chicken red blood cells; IL-1, interleukin 1; IL-2, interleukin 2; IFN, interferon     

Hic-5 interacts with GIT1 with a different binding mode from paxillin

Hic-5, a member of the paxillin family of adaptor molecules, is localized at focal adhesion and implicated in integrin-mediated signaling. Hic-5 and paxillin exhibit structural homology and share interacting factors, however, diverse functions are suggested for them. In this study, we carried out yeast two-hybrid screening to identify Hic-5 interacting factors using its LD3-4 region, which includes the Hic-5-specific amino acid sequence, as a bait. Through the screening, we identified GIT1, an Arf GTPase-activating protein, as a Hic-5 binding protein. The interaction of these two proteins was mediated by the LD3 motif of Hic-5 and the C-terminal region, which includes a paxillin-binding subdomain, of GIT1. Although GIT1 is known as a paxillin-binding protein, we only observed weak association of paxillin with GIT1 in the overexpression system. In contrast, Hic-5 firmly bound to GIT1 under the same conditions. In addition, the paxillin/GIT1 complex contained PIX, a guanine nucleotide exchange factor, whereas the Hic-5/GIT1 complex contained a smaller amount of PIX. These results suggested that paxillin and Hic-5 associate with GIT1 with different binding modes, and that the Hic-5 complex possesses static features compared with the paxillin complex, which contains both positive and negative regulators of GTPases involved in actin dynamics. Moreover, Hic-5-mediated inhibition of cell spreading was restored by co-expression of the C-terminal fragment of GIT1, which perturbs the interaction of Hic-5 with endogenous GIT1. Thus, it was demonstrated that Hic-5 and GIT1 interact functionally in addition to showing a physical association.     

Association between light exposure at night and manic symptoms in bipolar disorder: cross-sectional analysis of the APPLE cohort

ABSTRACT

Previous studies have found that keeping the room dark at night was associated with a decrease in manic symptoms for patients with bipolar disorder (BD). However, the association between light at night of real-life conditions and manic symptoms is unclear. We investigated the association between bedroom light exposure at night and manic symptoms in BD patients. One-hundred and eighty-four outpatients with BD participated in this cross-sectional study. The average light intensity at night during sleep was evaluated using a portable photometer for seven consecutive nights. Manic symptoms were assessed using the Young Mania Rating Scale (YMRS), and scores ≥5 were treated as a “hypomanic state.” The median (interquartile range) YMRS score was 2.0 (0–5.0), and 52 (28.2%) participants were in a hypomanic state. The prevalence of a hypomanic state was significantly higher in the participants with an average light intensity at night exposure of ≥3 lux than in those with <3 lux (36.7% versus 21.9%; P = .02). In multivariable logistic regression analysis adjusted for BD type, depressive symptoms, sleep duration, and daytime physical activity, the odds ratio (OR) for a hypomanic state was significantly higher for the participants with an average light intensity at night exposure of ≥3 lux than for those with <3 lux (OR: 2.15, 95% confidence interval: 1.09–4.22, P = .02). This association remained significant at the cutoff value of YMRS score ≥6 (OR: 2.51, 95% confidence interval: 1.15–5.46; P = .02). The findings of this study indicate bedroom light exposure at night is significantly associated with manic symptoms in BD patients. Although the results of this cross-sectional investigation do not necessarily imply causality, they may serve to inform beneficial nonpharmacological intervention and personalized treatment of BD patients.     

Acid-induced changes in the in vivo wall-yielding properties of hypocotyl sections of Vigna unguiculata

Elongation growth of hypocotyl sections of Vigna unguiculata under xylem perfusion was significantly enhanced when acid was applied by acid-aerosol to an abraded hypocotyl surface in the air. The in vivo wall extensibility (φ) and the effective turgor (Pⁱ– Y), both of which were determined by the pressure-jump method, increased during acid-induced growth as observed in IAA-induced growth. The intracellular pressure (Pⁱ), however, decreased significantly at the beginning of acid-induced growth whereas Pⁱ scarcely changed in IAA-induced growth. This result indicates that protons increase the effective turgor by decreasing the yield threshold as IAA does. There seems to be no essential difference between proton and auxin in the effects on the in vivo mechanical properties of the surface cell wall.     

Antimicrobial activity of lipoprotein particles containing apolipoprotein Al

Human plasmain vitro inhibits the growth of coagulase negative staphylococci,S. epidermidis, which may be pathogenic in the immunocompromised host. To determine the antimicrobial components, serum was fractionated by column chromatography, which revealed that elution areas where lipoproteins can be yielded had high antimicrobial activity againstS. epidermidis. Therefore, lipoprotein fractions, including very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL), were separated by ultracentrifugation and incubated withS. epidermidis. All 3 lipoprotein fractions suppressed bacterial growth within the first 3 h but VLDL enhanced bacterial growth after 9 h of incubation compared with the control. HDL, however, inhibited bacterial growth throughout 21 h of incubation.To confirm these results, serum from healthy volunteers was separated by ion exchange column chromatography and again by HPLC to purify the antimicrobial fraction. In the protein analysis with gradient polyacrylamide-SDS gel, apolipoprotein Al (apo Al), which is a major apolipoprotein of HDL, was detected in the antimicrobial fraction. Therefore, this fraction was loaded onto an immunoaffinity column coupled with the anti-apo Al monoclonal antibody (Mab). Unbound fraction had no antimicrobial activity, but anti-S. epidermidis activity was recovered from the bound fraction which consisted mainly of apo Al, All and apo C in protein composition.These results indicated that the antimicrobial activity was associated with the apo Al-containing lipoprotein particles (HDL). This property of HDL may directly affect bacterial growth and promote the self-defense mechanisms of normal and immunocompromised individuals.     

A Ca2+- and Voltage-Dependent Cl- -Sensitive Anion Channel in the Chara Plasmalemma: A Patch-Clamp Study

The inside-out patch-clamp technique was applied to the plasmolyzedplasmalemma of inter-nodes of Chara corallina without enzymatictreatment. We found two different types of channel activitythat were CP^–-sensitive. Both types of channel were Ca²⁺-dependent.However, the one that exhibited greater dependence on Ca²⁺ ionswas the focus of our studies, and we named it the Ca²⁺-dependentCP^–-sensitive anion channel. When the concentration ofCa²⁺ ions on the cyto-plasmic side was 1.0 µM, the Ca²⁺-dependentCP^–-sensitive channel opened most frequently between approximately–80 and –100 mV. At 10 µM Ca²⁺, it openedless frequently, and at 0.1 µM Ca²⁺ it scarcely openedat all. These observations indicate that the anion channel ofinterest is voltage-dependent over a restricted range of concentrationsof Ca²⁺ ions. The dependence on Ca²⁺ and voltage of the channelcan explain the behavior of the excitable Ca²⁺-activated Cl^–channel in the Chara plasmalemma. The channel activity was blockedby several antagonists of calmodulin. ⁴ Present Address: Department of Biology, College of GeneralEducation, Osaka University, Toyonaka, 560 Osaka, Japan (Received October 8, 1990; Accepted April 4, 1991)