Molecular cloning and characterization of two novel NAC genes from Mikania micrantha (Asteraceae).

NAC proteins, which are plant-specific transcription factors, have been identified to play important roles in plant response to stresses and in plant development. The full-length cDNAs that encode 2 putative NAC proteins, designated as MmATAF1 and MmNAP, respectively, were cloned from Mikania micrantha by rapid amplification of cDNA ends. The full-length cDNAs of MmATAF1 and MmNAP were 1329 and 1072 bp, respectively, and they encoded deduced proteins of 260- and 278-amino acid residues, respectively. The proteins MmATAF1 and MmNAP had a calculated molecular mass of 29.81 and 32.55 kDa and a theoretical isoelectric point of 7.08 and 9.00, respectively. Nucleotide sequence data indicated that both MmATAF1 and MmNAP contained 2 introns and 3 exons and that they shared a conserved genomic organization. Multiple sequence alignments showed that MmATAF1 showed high sequence identity with ATAF1 of Arabidopsis thaliana (61%) and that MmNAP showed high sequence identity with NAP of A. thaliana (67%) and CitNAC of Citrus sinensis Osbeck (62%). Phylogenetic analysis showed that the predicted MmATAF1 and MmNAP proteins were classified into the ATAF and NAP subgroups, respectively. Transient expression analysis of onion epidermal cells indicated nuclear localization of both MmATAF1-GFP and MmNAP-GFP fusion proteins. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) analysis indicated that MmATAF1 was expressed in all the tissues tested, but in varying abundance, while MmNAP was specifically expressed in stems, petioles, shoots, and leaves, but not in roots. The transcript levels of MmATAF1 and MmNAP in shoots and in infected stems were induced and strengthened by wounding, exogenous ZnSO(4), abscisic acid, salicylic acid, and Cuscuta campestris infection on the basis of semi-quantitative RT-PCR and real-time PCR analyses, respectively. Collectively, these results indicated that MmATAF1 and MmNAP, besides having roles in M. micrantha adaptation to C. campestris infection and abiotic stresses, also integrated signals derived from both C. campestris infection and abiotic stresses.

The use of dual-phase 18F-FDG PET in characterizing thyroid incidentalomas.

AIM: To examine the usefulness of dual-phase 2-[18F]-fluoro-2-deoxy-d-glucose (FDG) positron-emission tomography (PET) for the evaluation of thyroid incidentalomas. MATERIALS AND METHODS: In this retrospective study, cases with focal thyroid lesions seen incidentally at FDG PET in which the histopathological diagnosis was available and in which dual-phase FDG PET imaging was performed at 1 and 2 h after FDG injection were reviewed. In the included cases, the 1 and 2 h maximal standard uptake value (1-hour maximal SUV and 2-hour maximal SUV, respectively) and retention index (RI) were calculated, and the differences between benign and malignant thyroid incidentalomas were analysed. Receiver operating characteristic (ROC) analysis was performed to evaluate the ability of 1-hour maximal SUV, 2-hour maximal SUV, and RI to discriminate benign from malignant lesions. RESULTS: A total of 39 patients (25 females, 14 males) with 45 lesions (17 malignant, 28 benign) were included. In malignant thyroid incidentalomas, the average 1-hour maximal SUV, 2-hour maximal SUV, and RI were 5.20, 5.72, and 7.67%, respectively, and in benign thyroid incidentalomas the values were 4.67, 4.97, and 7.38%, respectively. There were no significant differences in 1-hour maximal SUV, 2-hour maximal SUV, and RI between benign and malignant lesions. The area under the ROC curve did not differ from 0.5. CONCLUSION: Dual-phase FDG PET is not useful for differentiating benign from malignant thyroid incidentalomas.

A tumor-suppressor function for Fas (CD95) revealed in T cell-deficient mice.

Fas (CD95) and its ligand are central regulatory molecules in hematopoietic cells. Previous studies have suggested a role for Fas in the regulation of tumor progression, but Fas has not yet been conclusively identified as a tumor suppressor. Fas-deficient individuals lack malignant tumors, perhaps because of regulation by T cells. To investigate such a possibility, mice deficient in both T cells and Fas were generated, and they were found to develop severe B cell dysregulation characterized by malignant, lethal B cell lymphoma. Lymphoma arose from a monoclonal B220+CD19-CD5-CD23- B cell secreting immunoglobulin M, kappa rheumatoid factor. In contrast, animals containing alpha beta T cells, gamma delta T cells, and/or functional Fas suppressed the development of lymphoma. These data indicate that Fas functions as a tumor suppressor, and identifies roles for both alpha beta T cells and gamma delta T cells in Fas-independent tumor regulation.

Extracellular matrix stability of primary mammalian chondrocytes and intervertebral disc cells cultured in alginate-based microbead hydrogels.

Three-dimensional alginate constructs are widely used as carrier systems for transplantable cells. In the present study, we evaluated the chondrogenic matrix stability of primary rat chondrocytes and intervertebral disc (IVD) cells cultured in three different alginate-based microbead matrices to determine the influence of microenvironment on the cellular and metabolic behaviors of chondrogenic cells confined in alginate microbeads. Cells entrapped in calcium, strontium, or barium ion gelled microbeads were monitored with the live/dead dual fluorescent cell viability assay kit and the 1,9-dimethylmethylene blue (DMB) assay designed to evaluate sulfated glycosaminoglycan (s-GAG) production. Expression of chondrogenic extracellular matrix (ECM) synthesis was further evaluated by semiquantitative RT-PCR of sox9, type II collagen, and aggrecan mRNAs. Results indicate that Ca and Sr alginate maintained significantly higher population of living cells compared to Ba alginate (p < 0.05). Production of s-GAG was similarly higher in Ca and Sr alginate microbead cultures compared to Ba alginate microbeads. Although there was no significant difference between strontium and calcium up to day 14 of culture, Sr alginate showed remarkably improved cellular and metabolic activities on long-term cultures, with chondrocytes expressing as much as 31% and 44% greater s-GAG compared to calcium and barium constructs, respectively, while IVD cells expressed 63% and 74% greater s-GAG compared to calcium and barium constructs, respectively, on day 28. These findings indicate that Sr alginate represent a significant improvement over Ca- and Ba alginate microbeads for the maintenance of chondrogenic phenotype of primary chondrocytes and IVD cells.

Roles of interferon-gamma and interleukin-4 in murine lupus.

The systemic autoimmune syndrome of MRL/Mp-lpr/lpr (MRL/lpr) mice consists of severe pan-isotype hypergammaglobulinemia, autoantibody production, lymphadenopathy, and immune complex-associated end-organ disease. Its pathogenesis has been largely attributed to helper alphabeta T cells that may require critical cytokines to propagate pathogenic autoantibody production. To investigate the roles of prototypical Th1 and Th2 cytokines in the pathogenesis of murine lupus, IFN-gamma -/- and IL-4 -/- lupus-prone mice were generated by backcrossing cytokine knockout animals against MRL/lpr breeders. IFN-gamma -/- animals produced significantly reduced titers of IgG2a and IgG2b serum immunoglobulins as well as autoantibodies, but maintained comparable levels of IgG1 and IgE in comparison to cytokine-intact controls; in contrast, IL-4 -/- animals produced significantly less IgG1 and IgE serum immunoglobulins, but maintained comparable levels of IgG2a and IgG2b as well as autoantibodies in comparison to controls. Both IFN-gamma -/- and IL-4 -/- mice, however, developed significantly reduced lymphadenopathy and end-organ disease. These results suggest that IFN-gamma and IL-4 play opposing but dispensable roles in the development of lupus-associated hypergammaglobulinemia and autoantibody production; however, they both play prominent roles in the pathogenesis of murine lupus-associated tissue injury, as well as in lpr-induced lymphadenopathy.

[Gene mutations from 511 myelodysplastic syndromes patients performed by targeted gene sequencing].

Objective: To study the characteristics of gene mutations in Chinese myelodysplastic syndromes (MDS) patients. Methods: A total of 511 Chinese patients with MDS performed 112-gene targeted sequencing were retrospectively analyzed. Results: Eighty-three distinct mutant genes were found in 511 patients with MDS. Amongst these, the most frequent mutations was associated with epigenetics (50%) , followed by spliceosome (37%) , signal transduction (34%) , transcription factors (24%) and cell cycle/apoptosis (17%) . 439 subjects (86%) had at least one gene mutation. The mean number of mutations in refractory anemia with unilineage dysplasia (RCUD) was 1.25, refractory anemia with multilineage dysplasia (RCMD) was 1.73, refractory anemia with ring sideroblasts (RARS) was 2.79, refractory anemia with excess blasts-1 (RAEB-1) was 2.22, RAEB-2 was 2.34, MDS with isolated 5q- was 2.67, MDS, unclassified (MDS-U) was 2.00. U2AF1 mutant subjects were more likely to have isolated+8[Q<0.001, OR=4.42 (95% CI 2.23-8.68) ]and less likely to have complex karyotypes[Q=0.005, OR=0.22 (95% CI 0.04-0.72) ]. According to the number of gene mutations, all subjects were categorized into three groups, namely group with 0-1 mutation, with 2 mutations and with three or more mutations. There was a significant difference in overall survival (OS) among three groups (P=0.041) . Conclusion: About 90% patients with MDS have at least one gene mutation. Genes associated with epigenetics and spliceosome are most common mutated genes in MDS. The increased numbers of gene mutations accompany with disease evolution and associate with poor prognosis.

Prognostic value of PUMA expression in patients with HBV-related hepatocellular carcinoma.

OBJECTIVE: The p53 upregulated modulator of apoptosis (PUMA) is a potent apoptosis inducer that is downexpressed in various tumor types. The aim of this study was to explore the prognostic significance of PUMA expression in patients with hepatitis B virus-related hepatocellular carcinoma (HCC). MATERIALS AND METHODS: PUMA expressions were examined in 80 pairs of tissues to compare its expression between cancer tissues and paired noncancerous liver tissues using immunohistochemistry (IHC). Relationship between the PUMA expression level and clinicopathological characteristics and clinic outcomes was analyzed. RESULTS: PUMA protein was all positive in paired non-tumor tissue samples. PUMA were downregulated in 61.25% (49/80) of tumor tissues compared with non-tumor tissues. PUMA levels in cancer tissues were significantly lower than non-tumor in patients with recurrence-related factors and patients at higher stage (stage II, III) (p < 0.05). In addition, the expressions level in tumor tissues showed a significant correlation with advanced TNM stage (p = 0.013) and present of recurrence-related factors (p = 0.002). Furthermore, Kaplan-Meier survival analysis revealed that weak PUMA expression was associated with poor 3-year disease-free survival (DFS) and overall survival (OS) in HCC patients. Finally, multivariate analyses identified that PUMA was an independent poor-prognostic predictor for DFS and OS in patients with HBV-related HCC. CONCLUSIONS: Our results suggest that PUMA expression is a novel prognostic indicator in HBV-related HCC and may be a potential target for diagnosis and gene therapy.

Systemic autoimmunity in LG/J mice.

Humoral and end-organ parameters of autoimmunity were investigated in LG/J mice, which have traditionally been considered normal, non-diseased animals. Surprisingly, LG/J mice were found to possess autoantibodies, including antinuclear antibodies and rheumatoid factor, and to develop renal disease, including glomerulonephritis, interstitial nephritis, and perivasculitis, but not hepatic or cutaneous disease. In contrast, age-matched, identically-housed control animals failed to develop autoantibodies or end-organ disease. These findings have complications for the genetic study of lupus erythematosus in the MRL murine model, which derives heavily from the LG/J background. Thus, the LG/J strain may provide a useful model in the analysis of autoimmunity.

A clinical study of schizophrenic suicides. 42 cases in Taiwan.

A clinical study of 25 male and 17 female schizophrenic suicides is presented. Jumping from a height was the most common method used. The 42 suicidal schizophrenics in Taiwan were compared with both 84 sex- and age-matched and 60 5-year illness course non-suicidal schizophrenic control groups. The suicidal schizophrenics were not significantly different from the non-suicidal counterparts of both control groups with regard to age, sex, ethnicity, religion, educational background, the presence of suicide cases in the family history, and the presence of insight, but were significantly different in characteristics of a history of previous suicide attempts, presence of psychotic symptoms during their final month, depression during their final month, a history of depression, a history of previous psychiatric hospitalizations, and the number of hospitalizations. We discuss the findings from this study and others in the literature in the context of the different clinical and socio-cultural backgrounds of these Taiwanese schizophrenic suicides.

Triterpenoid saponins from Berneuxia thebetica.

Four triterpenoid saponins were isolated from Berneuxia thibetica. On the basis of chemical and spectroscopic evidence, three new saponins, berneuxia saponins A, B and C, were elucidated as 21-tigloylbarringtogenol C-3 beta-O-alpha-L-rhamnopyranosyl(1-->2)-beta-D-galactopyranosyl(1-->3)[bet a- D-glucopyranosyl(1-->2)-beta-D-glucuronopyranoside], 28-tigloylbarringtogenol C-3 beta-O-alpha-L-rhamnopyranosyl(1-->2)-beta-D-galactopyranosyl(1-->3)[bet a- D-glucopyranosyl(1-->2)-beta-D-glucuronopyranoside] and 16 alpha-28-dihydroxyolean-12-en-21-one-3-O-alpha-L-rhamnopyranosyl(1 -->2) -beta-D-galactopyranosyl(1-->3)[beta-D-glucopyranosyl(1-->2)-beta-D- glucuronopyranoside]. The known saponin was desacyl jegosaponin.

[The alkaloids of Dactylicapnos torulosa (Hook f. et Thomas) Hutchins].

Six ingredients were isolated from the whole plant of Dactylicapnos torulosa (Hook. f. et Thomas.) Hutchins. Four of them were identified by UV, IR, 1HNMR and 13CNMR. One of them is a new compound named zijinlongine(I). The three others are 1-methoxyberberium chloride(II), triethylphenylammonium chloride (III) and isocorydine(IV). II and III were isolated from plants for the first time.

[Chemical constituents of Diuranthera inarticulata].

AIM: To investigate the chemical constituents from the roots of Diuranthera inarticulata Wang et K. Y. Lang. METHODS: The compounds were isolated with silica gel column chromatography and their structures were identified on the basis of spectral data and TLC with authentic samples. RESULTS: From the methanolic extracts of the roots of Diuranthera inarticulata, ten compounds were obtained and nine of them were identified as chloromaloside A (III), diuranthoside A (IV), diuranthoside B (V), diuranthoside F (VI), diuranthoside G (VII), 2 alpha, 3 alpha, 19 alpha-trihydroxy-urs-12-en-28-oic acid (VIII), 3,6,7,3',4'-pentamethoxy flavone (IX), gitogenin (X) and daucosterol (XI). III-VI showed inhibition of growth with the yeast, Candida albicans. III, IV and VI also showed inhibition of growth with fungus, Aspergillus niger, in the test of antibacterial activity. CONCLUSION: All of these compounds were isolated from this plant for the first time. Among them, VI and VII are new steroidal saponins.

[Studies on chemical constituents of Curcuma aromatica salisb].

OBJECTIVE: To study the effective ingredients of Curcuma aromatica. METHOD: Solvent extraction was used. The constituents were isolated with resin D-101 silica gel column and thin-layer chromatography, and the structures were elucidated by physico-chemical properties and spectral analysis. RESULT: Curdione, neocurdione, curcumol, tetramethylpyrazine and (R)-(+)-1,2-hexadecanediol were isolated from C. aromatica. CONCLUSION: Neocurdione and (R)-(+)-1,2-hexadecanediol were isolated from C. aromatica for the first time, and was isolated from Curcuma and reported for the first time.

[Chemical constituents from the fruit of Rosa bella Rehd. et Wils].

OBJECTIVE: To investigate the chemical constituents from the fruit of Rosa bella and its bioactivity. METHOD: The compounds were isolated from the methanol extract by column chromatography, then identified by 1H, 13C NMR and by comparison with authentic samples. The unorganized hoptoad heart was used for testing the contracting force of cardiac muscle and enzymology for measuring the bloody lipid of serum in rats. RESULT: Seven compounds were isolated and identified as pomolic acid, tiliroside, euscaphic acid, daucosterol, quercetin, oleanolic acid and beta-sitosterol, respectively. In the concentration of 0.4-1.0 mg.ml-1, the methanol extract had obvious cardiotonic effect. While at 400 mg.kg-1, there are no obvious effect to the TC, HDL-ac and TG of serum in rats. CONCLUSION: The isolated compounds may be the active components of this plant.

Reductionism and encephalitis lethargica, 1916-1939.

This paper is the 1992 winner of the Annual Stephen Wickes Prize in Medicine, presented by the Medical History Society of New Jersey. The history of encephalitis lethargica reflects a complex interaction between research-oriented science and clinical medicine.