Discovery of long-chain polyamines embedded in the biosilica on the Bacillus cereus spore coat.

Biosilicification is the process by which organisms incorporate soluble, monomeric silicic acid, Si(OH)4, in the form of polymerized insoluble silica, SiO2. Although the mechanisms underlying eukaryotic biosilicification have been intensively investigated, prokaryotic biosilicification has only recently begun to be studied. We previously reported that biosilicification occurs in the gram-positive, spore-forming bacterium Bacillus cereus, and that silica is intracellularly deposited on the spore coat as a protective coating against acids, although the underlying mechanism is not yet fully understood. In eukaryotic biosilicifying organisms, such as diatoms and siliceous sponges, several relevant biomolecules are embedded in biogenic silica (biosilica). These biomolecules include peptides, proteins, and long-chain polyamines. In this study, we isolated organic compounds embedded in B. cereus biosilica to investigate the biomolecules involved in the prokaryotic biosilicification process and identified long-chain polyamines with a chemical structure of H2N-(CH2)4-[NH-(CH2)3]n-NH2 (n: up to 55). Our results demonstrate the common presence of long-chain polyamines in different evolutionary lineages of biosilicifying organisms, i.e., diatoms, siliceous sponges, and B. cereus, suggesting a common mechanism underlying eukaryotic and prokaryotic biosilicification.

Usefulness of Five-Parameter System Reconfirmed for Cytopathology of Oral Squamous Cell Carcinoma regardless of Differentiation Degree.

BACKGROUND: We previously introduced the Five-Parameter System (FPS), which exclusively evaluates keratinized cellular findings, for use in cytology examinations of oral well-differentiated squamous cell carcinoma (SCC) and carcinoma in situ (CIS) specimens, as they occasionally lack nuclear atypia and can be challenging for categorization by The Bethesda System (TBS). This study was conducted to determine whether FPS parameters are detectable even in oral SCC/CIS specimens with apparent nuclear atypia. SUMMARY: Oral cytology specimens were obtained together with biopsy tissue samples. They were obtained from 59 malignant (HSIL and SCC) and 29 not-definitely malignant (NILM to ASC-H) specimens diagnosed using TBS. Following re-confirmation of the original TBS categorization, the specimens were re-evaluated using FPS. One or more of the FPS parameters were noted in 69 of 70 malignant specimens examined, of which 11 had been diagnosed by TBS as not-definitely malignant. The remaining one malignant specimen was diagnosed as SCC with only TBS. FPS parameters #1 (concentric arrangement), #2 (large cell number), #3 (bizarre-shaped cells), #4 (keratoglobules), and #5 (uneven filamentous cytoplasm) were observed only in malignant cases, while none were revealed in not-definitely malignant specimens. Finally, TBS supplemented with FPS achieved sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of 100%. KEY MESSAGES: FPS parameters are included in most examinations of oral cytology specimens. Thus, FPS is highly recommended for use in cytology examinations of oral SCC regardless of differentiation degree to confirm judgment based on TBS, a mandatory standard, as well as to cover its limitation of mainly evaluating nuclear atypia. FPS is considered to be an important diagnostic tool for oral cytology, especially in triage cases, which are challenging for TBS. Cytopathology should not be limited to only nuclear findings but be based on whole-cell morphology.

Differential diagnosis of well-differentiated squamous cell carcinoma from non-neoplastic oral mucosal lesions: New cytopathologic evaluation method dependent on keratinization-related parameters but not nuclear atypism.

BACKGROUND: The cytology of oral squamous cell carcinoma (SCC) is challenging because oral SCC cells tend to be well differentiated and lack nuclear atypia, often resulting in a false negative diagnosis. The purpose of this study was to establish practical cytological parameters specific to oral SCCs. METHODS: We reviewed 123 cases of malignancy and 53 of non-neoplastic lesions of the oral mucosa, which had been diagnosed using both cytology and histopathology specimens. From those, we selected 12 SCC and 4 CIS cases that had initially been categorized as NILM to ASC-H with the Bethesda system, as well as 4 non-neoplastic samples categorized as LSIL or ASC-H as controls, and compared their characteristic findings. After careful examinations, we highlighted five cytological parameters, as described in Results. Those 20 cytology samples were then reevaluated by 4 independent examiners using the Bethesda system as well as the 5 parameters. RESULTS: Five cytological features, (i) concentric arrangement of orangeophilic cells (indicating keratin pearls), (ii) large number of orangeophilic cells, (iii) bizarre-shaped orangeophilic cells without nuclear atypia, (iv) keratoglobules, and (v) uneven filamentous cytoplasm, were found to be significant parameters. All malignant cases contained at least one of those parameters, while none were observed in the four non-neoplastic cases with nuclear atypia. In reevaluations, the Bethesda system did not help the screeners distinguish oral SCCs from non-neoplastic lesions, while use of the five parameters enabled them to make a diagnosis of SCC. CONCLUSION: Recognition of the present five parameters is useful for oral SCC cytology. Diagn. Cytopathol. 2017;45:406-417. © 2017 Wiley Periodicals, Inc.

Properties of Zip4 accumulation during zinc deficiency and its usefulness to evaluate zinc status: a study of the effects of zinc deficiency during lactation.

Systemic and cellular zinc homeostasis is elaborately controlled by ZIP and ZnT zinc transporters. Therefore, detailed characterization of their expression properties is of importance. Of these transporter proteins, Zip4 functions as the primarily important transporter to control systemic zinc homeostasis because of its indispensable function of zinc absorption in the small intestine. In this study, we closely investigated Zip4 protein accumulation in the rat small intestine in response to zinc status using an anti-Zip4 monoclonal antibody that we generated and contrasted this with the zinc-responsive activity of the membrane-bound alkaline phosphatase (ALP). We found that Zip4 accumulation is more rapid in response to zinc deficiency than previously thought. Accumulation increased in the jejunum as early as 1 day following a zinc-deficient diet. In the small intestine, Zip4 protein expression was higher in the jejunum than in the duodenum and was accompanied by reduction of ALP activity, suggesting that the jejunum can become zinc deficient more easily. Furthermore, by monitoring Zip4 accumulation levels and ALP activity in the duodenum and jejunum, we reasserted that zinc deficiency during lactation may transiently alter plasma glucose levels in the offspring in a sex-specific manner, without affecting homeostatic control of zinc metabolism. This confirms that zinc nutrition during lactation is extremely important for the health of the offspring. These results reveal that rapid Zip4 accumulation provides a significant conceptual advance in understanding the molecular basis of systemic zinc homeostatic control, and that properties of Zip4 protein accumulation are useful to evaluate zinc status closely.

Cytological features of atypical carcinoid combined with adenocarcinoma of the uterine cervix.

Neuroendocrine tumors of the uterine cervix (UC) are rare, and atypical carcinoid (AC) combined with adenocarcinoma of the uterine cervix (ACAUC) is particularly rare. Only the histopathology has been investigated in the English literature. A 49-year-old female with a polypoid lesion of the UC visited Yamanashi Prefectural Central Hospital. Scraping cytology, biopsy, and hysterectomy was performed. EC smears showed solid, rosette, honeycomb,true tubular, and trabecular clusters. Solid clusters were oval, thin-edge, delicate, small-large nuclei, pale, granular, scant, nothing,and well-preserved (though ill-defined border) cytoplasm. Rosette clusters were eccentric, oval nuclei, mixture of coarsely granular chromatin and euchromatin, and cyanophilic luminal content. Solid and rosette clusters impress AC. Honeycomb clusters involved a clearly defined border and translucent mucin. True tubular clusters were oval nuclei of fine chromatin or euchromatin, thick cytoplasm,and orange luminal content. Honeycomb and true tubular clusters suspected adenocarcinoma. Trabecular clusters were fusiform, columnar, cuboidal, and polygonal cell shapes of small,monotonous nuclei, and contained coarsely granular chromatin with occasionally intracytoplasmic translucent mucin and were difficult to differentiate typical carcinoid and adenocarcinoma.Histology was AC combined with adenocarcinoma. The aim of this study was to investigate the cytological characteristics of ACAUC.

Adequate design of customized cDNA macroarray for convenient multiple gene expression analysis.

To establish a convenient, cost-effective, and reasonably reliable method for monitoring multiple gene expression using customized membrane-based macroarray, we constructed a cDNA macroarray with multiple probes for 13 human vascular endothelial genes and assessed the accuracy of the macroarray measurements. For each gene, two cDNA probes (450-550 bp) were designed from different regions (coding region and 3'-untranslated region [3'-UTR], respectively) on the basis of simple criteria concerning length and sequence specificity and spotted on the macroarray. In addition, unmodified oligonucleotide probes (80 mer) targeted to a unique sequence from the coding region of each gene were spotted on the same macroarray. Using this macroarray, shear stress-induced mRNA expression changes were analyzed in human coronary artery endothelial cells. Comparison of the expression ratios obtained with those measured using quantitative real-time polymerase chain reaction (PCR) as a reference method revealed that cDNA probes designed from a sequence within the coding region provided a highly accurate expression profile, whereas results obtained from oligonucleotide probes showed no correlation with real-time PCR data, which might be caused by inadequate immobilization of oligonucletotide probes on the nylon membrane. In addition, we observed that cDNA probes targeting different regions of a gene yielded different signal intensities. Most cDNA probes designed from a sequence within the coding region showed detectable signals, whereas few cDNA probes designed from 3'-UTR did.

Prenatal diagnosis of trisomy 9.

Fetal trisomy 9, especially its nonmosaic form, is a rare chromosomal abnormality and there are only 8 cases reported to have been sonographically detected in the prenatal period. We report a case of nonmosaic fetal trisomy 9, mimicking trisomy 13 on sonographic findings at 32 weeks' gestation. Although the incidence of trisomy 9 is rare, diagnosing trisomy 9 is important because the sonographic features are similar to those of trisomies 13 and 18, and cannot to be identified by routine fluorescencein situ hybridization. Because nonmosaic trisomy 9 is universally lethal, correct diagnosis and appropriate counseling is essential in patient care and clinical management.

Amplification of the synovial inflammatory response through activation of mitogen-activated protein kinases and nuclear factor kappaB using ligation of CD40 on CD14+ synovial cells from patients with rheumatoid arthritis.

OBJECTIVE: To determine the signal transduction pathways in CD14+ synovial cells from patients with rheumatoid arthritis (RA) after CD40 ligation, and to examine their role in amplifying synovial inflammation in affected joints. METHODS: Expression of messenger RNA was analyzed using quantitative reverse transcription-polymerase chain reaction. Cytokines and chemokines were measured using enzyme-linked immunosorbent assay. Activation of kinases was detected using Western blotting. Nuclear translocation of NF-kappaB was examined using immunohistochemistry. CD14+ synovial cells were enriched using magnetic cell sorting. Fibroblast-like synoviocytes (FLS) were obtained by passaging primary synovial cell culture. RESULTS: Stimulation of CD14+ synovial cells from RA patients by recombinant soluble CD154 (rsCD154) significantly induced expression of tumor necrosis factor alpha (TNFalpha), interleukin-1alpha (IL-1alpha), and IL-1beta. CD14+ RA synovial cells stimulated with rsCD154 plus interferon-gamma (IFNgamma) induced significantly higher production of IL-6, IL-8, and monocyte chemoattractant protein 1 by FLS compared with unstimulated CD14+ synovial cells, through TNFalpha-, IL-1alpha-, and IL-1beta-mediated pathways. Stimulation with rsCD154 plus IFNgamma induced the activation of ERK-1/2, p38 MAPK, and NF-kappaB. Specific inhibitors of MAPK/ERK-1/2 kinases and p38 MAPK significantly reduced the production of TNFalpha and IL-1beta by rsCD154 plus IFNgamma-stimulated CD14+ synovial cells, and also inhibited production of these cytokines by freshly isolated synovial cells from RA patients. CONCLUSION: These data indicate that the CD40-CD154 interaction activates the ERK, p38, and NF-kappaB pathways in CD14+ synovial cells from RA patients to produce TNFalpha, IL-1alpha, and IL-1beta, which in turn amplifies inflammatory responses by stimulating FLS. Inhibition of the CD40-CD154 interaction or its signal transduction pathways would be a strong and efficient strategy for the management of synovial inflammation in RA.