[Clinical characteristics of neonatal fungal sepsis in neonatal intensive care unit].

OBJECTIVE: To study the characteristics of neonatal fungal sepsis and the difference between bacterial sepsis and fungal sepsis. To improve the understanding of neonatal fungal sepsis. METHODS: Clinical data of neonatal fungal sepsis in neonatal intensive care unit (NICU) were collected from 2011 to 2016 in Peking University first Hospital. The clinical characteristics were analyzed retrospectively. The difference between neonatal fungal sepsis and bacterial sepsis was also analyzed. RESULTS: Fifteen cases of neonatal fungal sepsis were recruited. Over the study period, the incidence of neonatal fungal sepsis was 0.52%, while it was 2.5% in very low birth weight infants. Clinical characteristics were nonspeci-fic. All the infants were treated with parenteral nutrition and broad spectrum antibiotics. Peripheral inserted central catheter (PICC) was placed in thirteen patients. Pathogenic analyses indicated Candida glabrata was the main pathogen in our study. All the pathogens were sensitive to amphotericin B. Only one Candida glabrata was resistant to fluconazole. Thirty-four cases of bacterial sepsis were included. The clinical characteristics and laboratory examination results were compared. The platelet count was 61×109/L in fungal group, while the platelet count was 178×109/L in bacterial group. There was statistical difference between the fungal group and bacterial group (P=0.004). The rate of thrombocytopenia was 80.0% in fungal group, while it was 29.4% in bacterial group. It was much higher in fungal group than in bacterial group (P=0.001). The rate of PICC placement was 86.7% in fungal group, while it was 55.7% in bacterial group. It was much higher in fungal group than in bacterial group (P=0.037). Receiver operating characteristic (ROC) curve analysis showed that the cut-off value of the platelet count for the diagnosis of neonatal fungal sepsis was 145×109/L (sensitivity 61.8%, specificity 92.9%). All the patients were cured after standardized antifungal therapy. The indicators of liver and renal function were also measured before and after antifungal therapy. No significant difference was observed before and after treatment. CONCLUSION: The clinical characteristics of neonatal fungal sepsis was nonspecific. Candida glabrata was the main pathogen in our NICU. It can be cured as the result of standardized treatment. Decreased platelet count and PICC placement may indicate the possibility of fungal sepsis in neonates.

Leucine alleviates dexamethasone-induced suppression of muscle protein synthesis via synergy involvement of mTOR and AMPK pathways.

Glucocorticoids (GCs) are negative muscle protein regulators that contribute to the whole-body catabolic state during stress. Mammalian target of rapamycin (mTOR)-signalling pathway, which acts as a central regulator of protein metabolism, can be activated by branched-chain amino acids (BCAA). In the present study, the effect of leucine on the suppression of protein synthesis induced by GCs and the pathway involved were investigated. In vitro experiments were conducted using cultured C2C12 myoblasts to study the effect of GCs on protein synthesis, and the involvement of mTOR pathway was investigated as well. After exposure to dexamethasone (DEX, 100 µmol/l) for 24 h, protein synthesis in muscle cells was significantly suppressed (P<0.05), the phosphorylations of mTOR, ribosomal protein S6 protein kinase 1 (p70s6k1) and eukaryotic initiation factor 4E binding protein 1 (4EBP1) were significantly reduced (P<0.05). Leucine supplementation (5 mmol/l, 10 mmol/l and 15 mmol/l) for 1 h alleviated the suppression of protein synthesis induced by DEX (P<0.05) and was accompanied with the increased phosphorylation of mTOR and decreased phosphorylation of AMPK (P<0.05). Branched-chain amino transferase 2 (BCAT2) mRNA level was not influenced by DEX (P>0.05) but was increased by leucine supplementation at a dose of 5 mmol/l (P<0.05).

Clival invasion on multi-detector CT in 390 pituitary macroadenomas: correlation with sex, subtype and rates of operative complication and recurrence.

BACKGROUND AND PURPOSE: Clival invasion, a rare but potentially significant complication of pituitary adenoma, is difficult to detect on MR imaging. Because CT is widely used in adjunct guidance of pituitary surgery and it has recently been suggested that preoperative CT may add useful diagnostic information in addition to pituitary MR imaging, we performed the first large cross-sectional imaging study to define the image attributes, clinical correlates, and prognostic implications of clival invasion on CT for pituitary adenoma surgical guidance. MATERIALS AND METHODS: Preoperative CT images from 390 patients with histopathologically diagnosed pituitary macroadenoma were reviewed retrospectively and classified by the presence and degree of clival invasion. Tumor volume, tumor subtype, patient sex, operative complication, and recurrence rates were compared between groups. RESULTS: After we corrected for multiple correlations, the most significant independent risk factor for clival invasion was female sex (OR=3.62, P=.014, multinomial logistic regression), followed by large tumor volume (OR=1.08, P<.001), and null-cell subtype (OR=5.47, P<.001). Larger tumor volume correlated with null-cell subtype (Mann-Whitney U test, P=.006), incidence of clival invasion (P<.001), and extent of clival invasion (P=.038). Clival invasion was associated with a significantly higher ratio of operative complications (15.63%, χ(2)=7.067, P=.008) and recurrence (57.14%, χ(2)=10.739, P=.001). CONCLUSIONS: CT detection of clival invasion by pituitary macroadenoma is significantly more common in women, in patients with large tumors, and in patients with null-cell tumors, and it is associated with a higher rate of operative complications and recurrences. Attention to the presence of clival invasion on preoperative CT and prospective investigation of its prognostic significance are indicated. Attention to this finding on pituitary guidance CT is warranted.

[Effect of liuwei dihuang decoction, on the cytokine expression in splenocytes in AA rats].

OBJECTIVE: The effect of Liuwei Dihuang decoction (LW) on the function of splenic T helper cells (Th) was studied and compared with those of the immune inhibitors such as cyclophosphamide (Cy) and cyclosporin A (CsA) in AA rats. METHODS: The expression of mRNA for IFN-gamma, IL-2, IL-4 and IL-10 in splenic T-Lymphocytes of adjuvant arthritis (AA) rats was evaluated using reverse transcription polymerase chain reaction (RT-PCR) technique. RESULTS: mRNA expression level of IL-2 had a tendency to elevate, but the mRNA expression levels of IFN-gamma, IL-4 and IL-10 were significantly decreased in AA rats in comparison with control groups. LW treatment significantly inhibited the mRNA expression of IL-2 and promoted the expression of IFN-gamma, IL-4 and IL-10 in splenocytes of AA rats, with an obvious characteristic as compared with that of Cy or CsA. CONCLUSION: LW can correct the indifferent balance of the functions of splenocyte Th1/Th2 in AA rats.

The synthesis and vasopressin (AVP) antagonist activity of a novel series of N-aroyl-2,4,5,6-tetrahydropyrazolo[3,4-d]thieno[3,2-b]azepines.

Synthesis and SAR of N-[4-[(4,5-dihydropyrazolo[3,4-d]thieno[3,2-b]azepin-6(2H)-y l)carbonyl]phenyl]benzamides as arginine vasopressin (AVP) receptor antagonists are discussed. Potent orally active AVP receptor antagonists are produced when the benzamide moiety contains a phenyl group at the 2-position. Similar analogues of 4,6,7,8-tetrahydro-5H-thieno[3,2-b]azepine and VPA-985 are reported.

4,10-dihydro-5H-thieno[3,2-c][1]benzazepine derivatives and 9,10-dihydro-4H-thieno[2,3-c][1]benzazepine derivatives as orally active arginine vasopressin receptor antagonists.

Synthesis and structure-activity relationships (SAR) of arginine vasopressin receptor (AVP) antagonists are described. Potent and orally active compounds are prepared when tricyclic 10,11-dihydro-5H-pyrrolo[2,1-c][1,4]benzodiazepine moiety in VPA-985 1 is replaced with a compound 7 or 12.

5-fluoro-2-methyl-N-[5-(5H-pyrrolo[2,1-c][1,4]benzodiazepine-10(11H)-yl carbonyl)-2-pyridinyl]benzamide (CL-385004) and analogs as orally active arginine vasopressin receptor antagonists.

Synthesis and structure-activity relationships (SAR) of orally active arginine vasopressin (AVP) receptor antagonists are discussed. Potent and orally active AVP receptor antagonists are produced when ring A of VPA-985 (1) is replaced with a 3-pyridinyl unit (2b).

[Guiding-evaluation of immunomodulating activity of the Liuwei Dihuang Decoction during the stepwise fractionation].

OBJECTIVE: To conduct the activity evaluation for immunomodulating components from the Liuwei Dihuang Decoction (LWDHD). METHODS: Cyclophosphamide (Cy)-treated mice were used as the immunodeficient model and the antibody production response in plaque-forming cell (PFC) assay was employed as the activity-evaluating parameter. Stepwise fractionation was guided by activity-evaluation. RESULTS: Fr1, the ethanol-soluble fraction, and Fr2, the ethanol insoluble fraction of LWDHD, both significantly improved the antibody production response in Cy-treated mice, in which Fr2 showed stronger activity than that of Fr1. Fr2 was further fractionated. Fr2-CA4, one of the fractions obtained by active carbon chromatography from Fr2, exhibited the strongest activity in comparison with other fractions. CONCLUSION: CA4, which is mainly composed of acidic polysaccharides, is the main immunomodulating active fraction contained in the ethanol-insoluble fraction of LWDHD.

[Effects of low-molecular-weight Rehmannia glutinosa polysaccharides on p53 gene expression].

AIM: To study the effect of low-molecular-weight Rehmannia glutinosa polysaccharides (LRPS) on p53 gene expression. METHODS: The level of p53 gene expression was determined by quantitative polymerase chain reaction and autoradiography. RESULTS: The levels of p53 mRNA in Lewis lung cancer tissue were 0.46, 1.52, 1.48, and 1.60 respectively after saline, LRPS 20 and 40 mg.kg-1, and cyclophosphamide 10 mg.kg-1, which were the effective doses inhibiting the growth of tumor tissue cells in vivo. LRPS markedly increased p53 gene expression. CONCLUSION: LRPS effect on p53 gene expression is one of the mechanisms of antitumor activity.

Mutational analysis of vaccinia DNA ligase defines residues essential for covalent catalysis.

DNA ligation entails AMP transfer from ATP to the 5' end of DNA to form a DNA-adenylate structure, A(5')pp(5')N. A similar reaction involving GMP transfer occurs during 5' capping of eukaryotic mRNA. In both cases, nucleotidyl transfer occurs through a covalent lysyl-NMP intermediate. There is local sequence conservation among ligases and capping enzymes in the vicinity of the active site lysine (KxDG) and at three other collinear motifs. The role of these motifs in DNA ligation was tested by mutating individual conserved residues in the vaccinia virus DNA ligase. Wild-type and mutated versions of vaccinia ligase were expressed in bacteria as His-tagged fusion proteins and purified by Ni-affinity and phosphocellulose chromatography steps. We found that Ala substitution for Lys-231 (the presumptive active site) abrogated enzyme-adenylate formation and DNA ligation activities. Ala mutations at conserved residues Glu-283, Glu-377, and Lys-397 also resulted in loss of ligation activity, which correlated with a defect in ligase-AMP formation. These results are concordant with mutational studies of yeast RNA capping enzyme and suggest a common structural basis for covalent nucleotidyl transfer.

Regulation of mast cell differentiation studied using the diffusion chamber technique.

A homogeneous population of mast cells was obtained by culturing bone marrow cells of WBB6F1(-)+/+ mice. The proliferation of the cultured mast cells in diffusion chambers was investigated to examine whether the diffusion chamber technique was applicable for study of the regulation of mast cell proliferation. WBB6F1-W/Wv mice are genetically deficient in mast cells. When cultured mast cells of WBB6F1(-)+/+ mouse origin were directly injected into the peritoneal cavity of WBB6F1-W/Wv mice, the mast cells survived. In contrast, WBB6F1(-)+/+ mouse-derived cultured mast cells did not survive in diffusion chambers implanted in the peritoneal cavity of either WBB6F1-W/Wv or WBB6F1(-)+/+ mice. Because the coinoculation of NIH/3T3 cells supported the proliferation of mast cells in diffusion chambers, a certain type of cells in the peritoneal cavity appeared to have the same mast cell-supporting activity as NIH/3T3 cells. The magnitude of either interleukin 3-dependent or NIH/3T3 cell-dependent proliferation of mast cells in diffusion chambers was not significantly influenced by the genotype of chambers recipients (i.e., WBB6F1(-)+/+ or WBB6F1-W/Wv mice), suggesting that the previously reported inhibitory effect of mast cells on differentiation of mast cells may be mediated by direct contact between mast cells.

Suppressive effect of Sl/Sld mouse embryo-derived fibroblast cell lines on diffusible factor-dependent proliferation of mast cells.

Two modes of mast cell growth are present, one dependent on diffusible growth factors (interleukins [IL] 3 and 4) and another dependent on contact with fibroblasts. The 3T3 fibroblast cell lines derived from WCB6F1-+/+ mouse embryos supported the proliferation of cultured mast cells (CMC), whereas the 3T3 fibroblast cell lines from WCB6F1-Sl/Sld mouse embryos did not. To investigate the relationship between growth factor-dependent and fibroblast-dependent growths of mast cells, we cocultured CMC and 3T3 fibroblasts in the presence of diffusible growth factors. WCB6F1-+/+ mouse embryo-derived 3T3 cells did not affect the growth factor-dependent proliferation of CMC, but WCB6F1-Sl/Sld mouse embryo-derived 3T3 cells significantly suppressed the proliferation. Close cell-to-cell contact was necessary for the suppression. The NWS1 fibroblast cell line was established from the spleen cells of an adult WBB6F1-+/+ mouse. Although the NWS1 cell line had no supporting effect on the proliferation of CMC in the absence of diffusible growth factors, it did not suppress the proliferation of CMC induced by the growth factors. The present result suggests that a product of mutant Sl genes may be involved in the suppressive activity of WCB6F1-Sl/Sld mouse embryo-derived 3T3 cells.

Fast and easy detection of mouse sex chimeras using electrophoretic polymorphism of phosphoglycerate kinase-1, an X chromosome-linked enzyme.

About half of the chimeras produced by aggregation of two mouse embryos are sex chimeras composed of both XX and XY cells. We developed a fast and easy method to identify sex chimeras by using electrophoretic bimorphism of an X-linked enzyme, phosphoglycerate kinase-1 (PGK-1), as a marker. When embryos resulting from the crossing of a Pgk-1b/Pgk-1b female and a Pgk-1a/Y male are aggregated, the genotype of sex chimeras is Pgk-1b/Pgk-1a----Pgk-1b/Y. Most of these were identifiable from the PGK-1 electrophoretic pattern of blood cells (i.e., AB type) and the appearance of genitalia (male type or apparently abnormal). Genotypes of functional sperm in the testes of the male-type sex chimeras were also identifiable from the PGK-1 electrophoretic pattern of progenies. Examination of gonads of the sex chimeras revealed that a considerable proportion was hermaphorditic. With this method, reasonable numbers of male-type sex chimeras and hermaphrodites may be selected and used as material for investigating sexual differentiation.

Characteristics of hemopoietic stem cells determined by single spleen colony transplantation.

By using a spleen colony transfer technique and sex chromosome determinations as a cytogenetic marker, we compared the spleen colony-forming cells of bone marrow origin from three strains of mice, i.e., LACA, C57, and (LACA female X C57 male) F1. The proliferation potential of spleen colony-forming cells was found to be heterogeneous in nature. Some of the cells have the essential characteristics of hemopoietic stem cells (Ps-CFUS), i.e., the capacity for sustained proliferation and differentiation into myeloid and lymphoid lineages of cells, whereas others exhibit the capacity to form gross colonies on irradiated recipient spleen which are capable of differentiating different lineages of blood cells, but have lost the ability to reconstitute hemopoiesis in lethally irradiated recipients (Pg-CFUS). The results of studies on spleen colony-forming cells of F1 mice indicate the existence of a subpopulation of primitive precursors of spleen colony-forming cells (Pre-CFUs) which are not able to form spleen colonies on recipient spleens, but, under appropriate stimuli, may assume the properties of Ps-CFUS. Therefore the sequential process of blood cell differentiation may be described as follows: Pre-CFUS----Ps-CFUS----Pg-CFUS----different lineages of hemopoietic progenitors----different lineages of blood cells.