Lymphonodular cryptococcosis diagnosed by fine needle aspiration cytology in hyper-IgM syndrome. A case report

BACKGROUND: Most cases of cryptococcosis are diagnosed when signs of meningitis have appeared. We report a case of lymphonodular cryptococcosis that was diagnosed by fine needle aspiration cytology (FNAC), excisional biopsy of a cervical lymph node and culture of aspirated material. CASE: An 11-year-old boy presented with a history of fever and enlarged bilateral cervical lymph nodes of two weeks' duration. Past medical history included immunoglobulin replacement for hyper-IgM syndrome for the previous eight years. FNAC smears from a cervical lymph node showed numerous yeasts of various sizes, ranging from 5 to 15 microns in diameter, located in the cytoplasm of multinucleated giant cells and in the background. In air-dried, Diff-Quik-stained slides, the yeasts stained blue and were surrounded by clear halos. Aspirated material collected in the syringe was cultured, and Cryptococcus neoformans was isolated. CONCLUSION: This case report suggests that a combination of FNAC and culture is a simple and useful method of diagnosing fungal infections. Early diagnosis by FNAC makes possible the early initiation of treatment.     

Dramatically accelerated growth and extraordinary gigantism of transgenic mud loach Misgurnus mizolepis

Transgenic mud loaches (Misgurnus mizolepis), in which the entire transgene originated from the same species, have been generated by microinjecting the mud loach growth hormone (mlGH) gene fused to the mud loach -actin promoter. Out of 4,100 eggs injected, 7.5% fish derived from the injected eggs showed dramatically accelerated growth, with a maximum of 35-fold faster growth than their non-transgenic siblings. Many fast-growing transgenic individuals showed extraordinary gigantism: their body weight and total length (largest fish attained to 413g and 41.5cm) were larger and longer than even those of 12-year-old normal broodstock (maximum size reached to 89g and 28cm). Of 46 transgenic founders tested, 30 individuals transmitted the transgene to next generation with a wide range of germ-line transmission frequencies ranging from 2% to 33%. The growth performance of the subsequent generation (F₁) was also dramatically accelerated up to 35-fold, although the levels of enhanced growth were variable among transgenic lines. Three transgenic germ-lines up to F₄ were established, showing the expected Mendelian inheritance of the transgene. Expression of GH mRNA in many tissues was detected by RT-PCR analyses. The time required to attain marketable size (10g) in these transgenic lines was only 30–50 days after fertilization, while at least 6 months in non-transgenic fish. Besides growth enhancement, significantly improved feed-conversion efficiency up to 1.9-fold was also observed.     

The erythropoietin receptor cytosolic juxtamembrane domain contains an essential, precisely oriented, hydrophobic motif

We report that the erythropoietin receptor cytosolic juxtamembrane region is conformationally rigid and contains a hydrophobic motif, composed of residues L253, I257, and W258, that is crucial for Janus kinase 2 (JAK2) activation and receptor signaling. Alanine insertion mutagenesis shows that the orientation of this motif and not its distance from the membrane bilayer is critical. Intragenic complementation studies suggest that L253 is contained within an alpha helix functionally continuous to the transmembrane alpha helix. The alpha-helical orientation of L53 is required not for JAK2 activation but for activated JAK2 to induce phosphorylation of the erythropoietin receptor. This motif is highly conserved among cytokine receptors and couples ligand-induced conformational changes in the receptor to intracellular activation of JAK2.     

Nanoscale fabrication of biomolecular layer and its application to biodevices

Biodevices composed of biomolecular layer have been developed in various fields such as medical diagnosis, pharmaceutical screening, electronic device, photonic device, environmental pollution detection device, and etc. The biomolecules such as protein, DNA and pigment, and cells have been used to construct the biodevices such as biomolecular diode, biostorage device, bioelectroluminescence device, protein chip, DNA chip, and cell chip. Substantial interest has focused upon thin film fabrication or the formation of biomaterials mono- or multi-layers on the solid surfaces to construct the biodevices. Based on the development of nanotechnology, nanoscale fabrication technology for biofilm has been emerged and applied to biodevices due to the various advantages such as high density immobilization and orientation control of immobilized biomolecules. This review described the nanoscale fabrication of biomolecular film and its application to bioelectronic devices and biochips.     

Regulation of lymphocyte clustering by CD30-mediated ICAM-1 up-regulation

CD30 is expressed transiently on activated B and T lymphocytes and constitutively on several B- and T cell lymphomas. CD30 functions include participation in negative selection of thymocytes, costimulation of activated T cells, isotype switching of B cells, and regulation of the effector activity of cytotoxic lymphocytes. Although CD30 is not a marker for T helper 2 (TH2) cells, it may participate in the polarization of TH1 and TH2 cells. The pleiotropic functions of CD30 are initiated by interaction of CD30-expressing cells with other immune competent cells expressing CD30-L and providing the signals for modulation of effector cell activity. Here, we report that CD30 signals generated by anti-CD30 on activated, normal murine T cells strongly up-regulate the expression of intercellular adhesion molecule 1 (ICAM-1, CD54), and to a lesser extent, ICAM-2 (CD102). CD30 signals moreover delay the subsequent decline of ICAM expression. CD30 cross-linking did not alter the expression of CD11a/CD18 (LFA-1), the counter receptor for ICAM abundant on T cells. CD30-mediated ICAM-1 up-regulation is independent of cytokine secretion and appears to be transmitted directly through NF-kappaB activation. CD30-mediated up-regulation of ICAM-1 expression led to a significant increase in cluster formation of lymph node cells. Increased lymphocyte self-aggregation mediated by CD30 may set the stage for fraternal signaling to modulate lymphocyte function.     

Depletion of mitochondrial DNA alters glucose metabolism in SK-Hep1 cells

Maternally inherited mitochondrial DNA (mtDNA) has been suggested to be a genetic factor for diabetes. Reports have shown a decrease of mtDNA content in tissues of diabetic patients. We investigated the effects of mtDNA depletion on glucose metabolism by use of rho(0) SK-Hep1 human hepatoma cells, whose mtDNA was depleted by long-term exposure to ethidium bromide. The rho(0) cells failed to hyperpolarize mitochondrial membrane potential in response to glucose stimulation. Intracellular ATP content, glucose-stimulated ATP production, glucose uptake, steady-state mRNA and protein levels of glucose transporters, and cellular activities of glucose-metabolizing enzymes were decreased in rho(0) cells compared with parental rho(+) cells. Our results suggest that the quantitative reduction of mtDNA may suppress the expression of nuclear DNA-encoded glucose transporters and enzymes of glucose metabolism. Thus this may lead to diabetic status, such as decreased ATP production and glucose utilization.     

Cloning of Japanese flounder Paralichthys olivaceus CD3 cDNA and gene, and analysis of its expression

Two distinct CD3 homologue cDNAs, CD3-1 and CD3-2, were isolated from a Japanese flounder leukocyte cDNA library. CD3-1 consisted of 961 bp encoding 178 amino acid residues, and CD3-2 consisted of 927 bp encoding 182 amino acid residues. The two deduced amino acid sequences had an identity of 95.1%, and neither had N-linked glycosylation sites. The identities between the Japanese flounder CD3s and previously reported CD3s (CD3 epsilon, CD3 gamma, or CD3 delta) of Xenopus laevis, chicken, and various mammals were approximately 25%. The Japanese flounder CD3s had an extracellular domain, a CXXCXE motif, and an immunoreceptor tyrosine-based activation motif (ITAM), each of which are important characteristics of CD3 chains. Furthermore, the positions of four cysteine residues in the extracellular domain were preserved in both of the Japanese flounder CD3s. A phylogenetic tree based on the amino acid sequences confirmed that the Japanese flounder CD3s are closer to CD3 epsilon than to CD3 gamma and CD3 delta. However, the gene structure of Japanese flounder CD3 is identical to the chicken and Xenopus CD3 gamma/delta genes and the mammalian CD3 delta gene. Southern blot hybridization and the DNA sequence of the CD3 gene of homocloned Japanese flounder indicated that the CD3 gene exists as a single copy. Southern blot hybridization also showed the presence of a polymorphic variant of Japanese flounder CD3. An RT-PCR analysis detected Japanese flounder CD3 mRNA in several organs that contained lymphocytes. The proportion of CD3-positive cells in the peripheral blood leukocytes was 34.9%.     

Contribution of the hydrogen-bond network involving a tyrosine triad in the active site to the structure and function of a highly proficient ketosteroid isomerase from Pseudomonas putida biotype B

Delta(5)-3-Ketosteroid isomerase from Pseudomonas putida biotype B is one of the most proficient enzymes catalyzing an allylic isomerization reaction at rates comparable to the diffusion limit. The hydrogen-bond network (Asp99... Wat504...Tyr14...Tyr55...Tyr30) which links the two catalytic residues, Tyr14 and Asp99, to Tyr30, Tyr55, and a water molecule in the highly apolar active site has been characterized in an effort to identify its roles in function and stability. The DeltaG(U)(H2O) determined from equilibrium unfolding experiments reveals that the elimination of the hydroxyl group of Tyr14 or Tyr55 or the replacement of Asp99 with leucine results in a loss of conformational stability of 3.5-4.4 kcal/mol, suggesting that the hydrogen bonds of Tyr14, Tyr55, and Asp99 contribute significantly to stability. While decreasing the stability by about 6.5-7.9 kcal/mol, the Y55F/D99L or Y30F/D99L double mutation also reduced activity significantly, exhibiting a synergistic effect on k(cat) relative to the respective single mutations. These results indicate that the hydrogen-bond network is important for both stability and function. Additionally, they suggest that Tyr14 cannot function efficiently alone without additional support from the hydrogen bonds of Tyr55 and Asp99. The crystal structure of Y55F as determined at 1.9 A resolution shows that Tyr14 OH undergoes an alteration in orientation to form a new hydrogen bond with Tyr30. This observation supports the role of Tyr55 OH in positioning Tyr14 properly to optimize the hydrogen bond between Tyr14 and C3-O of the steroid substrate. No significant structural changes were observed in the crystal structures of Y30F and Y30F/Y55F, which allowed us to estimate approximately the interaction energies mediated by the hydrogen bonds Tyr30...Tyr55 and Tyr14...Tyr55. Taken together, our results demonstrate that the hydrogen-bond network provides the structural support that is needed for the enzyme to maintain the active-site geometry optimized for both function and stability.