Increased tube formation and up-regulation of FGFR3 mRNA expression in microvascular endothelial cell by exosomes derived from SW480-7.

INTRODUCTION: Extracellular vesicles (exosome-like vesicles) are small membrane vesicles ranging from 20-200nm in size that are released by various cells into the extracellular space. These extracellular vesicles play a major role in cell-to-cell communication and contain materials, such as proteins, mRNAs, microRNAs (miRNAs) and long non-coding RNAs (lncRNAs). The effect of exosomes derived from an invasive colon cancer cell line on angiogenesis is unclear. Hence, the aim of this study is to investigate the effect of exosomes derived from an invasive colon cancer cell line on angiogenesis of endothelial cells. MATERIALS AND METHODS: In the present study, the exosomes from the cell culture supernatants of an invasive colon cancer cell line SW480-7 were characterised. The effect on tube formation and expression of angiogenic genes in a microvascular endothelial cell, telomerase-immortalised microvascular endothelial cell (TIME) was examined after co-cultured with exosomes secreted from SW480-7. RESULTS: Zetasizer result showed average diameter of exosomes derived from SW480-7 was 246.2 nm and morphological analysis showed the size of majority of exosomes were less than 200 nm. Results showed that exosomes derived from SW480-7 increased tube formation and up-regulated FGFR3 mRNA expression in TIME. CONCLUSION: Our findings suggest that exosomes derived from SW480-7 increased tube formation and up-regulated expression of FGFR3 mRNA in TIME.

Update on the Hong Kong Reference Framework for Hypertension Care for Adults in Primary Care Settings-review of evidence on the definition of high blood pressure and goal of therapy.

The Hong Kong Reference Framework for Hypertension Care for Adults in Primary Care Settings is updated regularly to ensure it reflects the latest medical development and best practice. In 2017, guidelines from the United States included a major change, adopting the lower blood pressure values of 130/80 mm Hg in defining hypertension, in contrast to the prevailing international consensus of 140/90 mm Hg. After thorough review of the literature and international guidelines, the Advisory Group on Hong Kong Reference Framework for Care of Diabetes and Hypertension in Primary Care Settings (Advisory Group) recommends that the definition of hypertension adopted in the Reference Framework should remain unchanged as a blood pressure of ≥140/90 mm Hg, as there is currently inadequate evidence and lack of general consensus to support such change in Hong Kong. The Advisory Group agrees on individualised treatment goals, and recommends that the initial blood pressure goal for individuals with uncomplicated hypertension should be <140/90 mm Hg; for those who can tolerate it, the goal should be ≤130/80 mm Hg. A lower blood pressure is advisable for young or overweight/obese patients, smokers, and patients with other cardiovascular risk factors.

Gene expression profile alone is inadequate in predicting complete response in multiple myeloma.

With advent of several treatment options in multiple myeloma (MM), a selection of effective regimen has become an important issue. Use of gene expression profile (GEP) is considered an important tool in predicting outcome; however, it is unclear whether such genomic analysis alone can adequately predict therapeutic response. We evaluated the ability of GEP to predict complete response (CR) in MM. GEP from pretreatment MM cells from 136 uniformly treated MM patients with response data on an IFM, France led study were analyzed. To evaluate variability in predictive power due to microarray platform or treatment types, additional data sets from three different studies (n=511) were analyzed using same methods. We used several machine learning methods to derive a prediction model using training and test subsets of the original four data sets. Among all methods employed for GEP-based CR predictive capability, we got accuracy range of 56-78% in test data sets and no significant difference with regard to GEP platforms, treatment regimens or in newly diagnosed or relapsed patients. Importantly, permuted P-value showed no statistically significant CR predictive information in GEP data. This analysis suggests that GEP-based signature has limited power to predict CR in MM, highlighting the need to develop comprehensive predictive model using integrated genomics approach.

Vitamin D-binding protein haplotype is associated with hospitalization for RSV bronchiolitis.

BACKGROUND: Between 75 000 and 125 000 U.S. infants are hospitalized for respiratory syncytial virus (RSV) bronchiolitis every year. Up to half will be diagnosed with asthma in later childhood. Vitamin D deficiency has been associated with susceptibility to asthma and respiratory infections. Measured vitamin D is largely bound to vitamin D-binding protein (VDBP); VDBP levels are influenced by its gene (GC) haplotype. OBJECTIVE: We assessed the relationship between polymorphisms rs7041 and rs4588, which define haplotypes GC1s, GC1f, and GC2, and RSV bronchiolitis susceptibility and subsequent asthma. METHODS: We retrospectively recruited 198 otherwise healthy children (93% White) hospitalized for severe RSV bronchiolitis in Boston and 333 parents into a follow-up study to assess asthma diagnosis. Data were analysed using family-based genetic association tests. We independently validated our results in 465 White children hospitalized with RSV bronchiolitis and 930 White population controls from the Netherlands. RESULTS: The rs7041_C allele (denoting haplotype GC1s) was overtransmitted (P = 0.02, additive model) in the entire Boston cohort, in Whites (P = 0.03), and especially in children subsequently diagnosed with asthma (P = 0.006). The GC1f haplotype was undertransmitted in the asthma subgroups (all races and White, both P < 0.05). The rs7041_C allele was also more frequent in the RSV bronchiolitis group compared with controls (OR 1.12, 95% CI 1.02, 1.4, P = 0.03) in the Netherlands, especially in mechanically ventilated patients (P = 0.009). CONCLUSION AND CLINICAL RELEVANCE: GC1s haplotype carriage may increase the risk of RSV bronchiolitis in infancy and subsequent asthma development. The GC1s haplotype is associated with higher VDBP levels, resulting in less freely available vitamin D. KEY MESSAGES: Vitamin D-binding protein (VDBP) haplotypes influence free vitamin D levels. We report an association between a VDBP haplotype and hospitalization for RSV bronchiolitis in infancy in two independent cohorts.

Basic fibroblast growth factor modulates cell cycle of human umbilical cord-derived mesenchymal stem cells.

BACKGROUND: Mesenchymal stem cells (MSC) have great potential in regenerative medicine, immunotherapy and gene therapy due to their unique properties of self-renewal, high plasticity, immune modulation and ease for genetic modification. However, production of MSC at sufficient clinical scale remains an issue as in vitro generation of MSC inadequately fulfils the demand with respect to patients. OBJECTIVES: This study has aimed to establish optimum conditions to generate and characterize MSC from human umbilical cord (UC-MSC). MATERIALS AND METHODS: To optimize MSC population growth, basic fibroblast growth factor (bFGF) was utilized in culture media. Effects of bFGF on expansion kinetics, cell cycle, survival of UC-MSC, cytokine secretion, expression of early stem-cell markers and immunomodulation were investigated. RESULTS: bFGF supplementation profoundly enhanced UC-MSC proliferation by reducing population doubling time without altering immunophenotype and immunomodulatory function of UC-MSC. However, cell cycle studies revealed that bFGF drove the cells into the cell cycle, as a higher proportion of cells resided in S phase and progressed into M phase. Consistent with this, bFGF was shown to promote expression of cyclin D proteins and their relevant kinases to drive UC-MSC to transverse cell cycle check points, thus, committing the cells to DNA synthesis. Furthermore, supplementation with bFGF changed the cytokine profiles of the cells and reduced their apoptotic level. CONCLUSION: Our study showed that bFGF supplementation of UC-MSC culture enhanced the cells' growth kinetics without compromising their nature.

Increase in tumour-infiltrating lymphocytes with regulatory T cell immunophenotypes and reduced zeta-chain expression in nasopharyngeal carcinoma patients.

The pathological significance of the mechanisms of tumour immune-evasion and/or immunosuppression, such as loss of T cell signalling and increase in regulatory T cells (T(regs)), has not been well established in the nasopharyngeal carcinoma (NPC) microenvironment. To evaluate the T(reg) immunophenotypes in tumour-infiltrating lymphocytes (TILs), we performed a double-enzymatic immunostaining for detection of forkhead box P3 (FoxP3) and other markers including CD4, CD8, and CD25 on 64 NPC and 36 non-malignant nasopharyngeal (NP) paraffin-embedded tissues. Expression of CD3 zeta and CD3 epsilon was also determined. The prevalence of CD4(+)FoxP3(+) cells in CD4(+) T cells and the ratio of FoxP3(+)/CD8(+) were increased significantly in NPC compared with those in NP tissues (P < 0.001 and P = 0.025 respectively). Moreover, the ratio of FoxP3(+)/CD25(+)FoxP3(-) in NPC was significantly lower than that in NP tissues (P = 0.005), suggesting an imbalance favouring activated phenotype of T cells in NPC. A significant negative correlation between the abundance of FoxP3(+) and CD25(+)FoxP3(-) cells (P < 0.001) was also identified. When histological types of NPC were considered, a lower ratio of FoxP3(+)/CD25(+)FoxP3(-) was found in non-keratinizing and undifferentiated carcinomas. Increased CD4(+)FoxP3(+)/CD4(+) proportion and FoxP3(+)/CD8(+) ratio were associated with keratinizing squamous cell carcinoma. A reduced expression of CD3 zeta in TILs was found in 20.6% of the NPC tissues but none of the NP tissues. These data provide evidence for the imbalances of T(reg) and effector T cell phenotypes and down-regulation of signal-transducing molecules in TILs, supporting their role in suppression of immune response and immune evasion of NPC.

Effect of sludge fasting/feasting on growth of activated sludge cultures.

Reduction of excess sludge in an oxic-settling-anoxic (OSA) activated sludge process might be attributed to a "sludge fasting (insufficient food under an anoxic condition)/feasting (sufficient food under an oxic condition)" treatment. This paper was to examine this explanation by investigating both the sludge fasting/feasting phenomenon and the effect of a fasting/feasting treatment on sludge growth. In this study, five different activated sludge cultures cultivated using synthetic wastewater composed of mainly glucose and other necessary nutrients: (1) an aerobic batch culture, (2) an intermittently aerated batch culture, (3) an anoxic batch culture, (4) a continuous aerobic culture, and (5) an OSA culture, were employed. It was found that only the aerobic batch culture and the aerobic continuous culture are fastable when the oxidation reduction potential (ORP) level is below 100 mV under no-food condition during a 2-h fasting treatment, showing that both the biomass and carbohydrate storage of these two cultures were reduced after the treatment. When the fasted cultures were treated in a feasting environment, an accumulation of carbohydrate storage did not occur, while specific oxygen uptake rates (SOUR) showed a sharp increase. Both the substrate utilization and biomass growth rates were also accelerated. It was therefore confirmed that a sludge feasting did occur after a fasting treatment for the fastable cultures. However, an increase in sludge ATP content was not brought about by the feasting treatment. The sludge fasting/feasting treatment in this paper could not induce a reduction of the observed growth yield (Y(obs)) in all the cultures cultivated with glucose-based synthetic wastewater.

The promoter of LE-ACS7, an early flooding-induced 1-aminocyclopropane-1-carboxylate synthase gene of the tomato, is tagged by a Sol3 transposon.

Many terrestrial plants respond to flooding with enhanced ethylene production. The roots of flooded plants produce 1-aminocyclopropane-1-carboxylic acid (ACC), which is transported from the root to the shoot, where it is converted to ethylene. In the roots, ACC is synthesized by ACC synthase, which is encoded by a multigene family. Previously, we identified two ACC synthase genes of tomato that are involved in flooding-induced ethylene production. Here, we report the cloning of LE-ACS7, a new tomato ACC synthase with a role early during flooding but also in the early wound response of leaves. The promoter of LE-ACS7 is tagged by a Sol3 transposon. A Sol3 transposon is also present in the tomato polygalacturonase promoter to which it conferred regulatory elements. Thus, Sol3 transposons may affect the regulation of LE-ACS7 and may be involved in the communication between the root and the shoot of waterlogged tomato plants.

Characterization of adrenoceptors involved in the electrogenic chloride secretion by cultured rat epididymal epithelium.

1. Short-circuit current (SCC) technique was used to study the adrenoceptors involved in the electrogenic chloride secretion by cultured cauda epididymal epithelium of rats. Stimulation of the epithelium with noradrenaline (primarily beta 1-adrenoceptor selective agonist), salbutamol (beta 2-adrenoceptor selective agonist) and adrenaline (non-selective beta-adrenoceptor agonist) led to a rise in SCC. At a low chart-speed (2 mm min-1), the response profile to these agonists consisted of a peak followed by a sustained response considerably higher than the basal SCC. 2. The EC50s (doses of agonist producing 50% maximum response) of noradrenaline, salbutamol and adrenaline were 300, 115 and 10 nM respectively. Pretreating the tissues with 1 microM atenolol (beta 1-selective antagonist) and 10 microM butoxamine (beta 2-selective antagonist) shifted the dose-response curves of noradrenaline (shifted EC50 = 4000 nM) and salbutamol (shifted EC50 = 1050 nM) to the right. Atenolol (1 microM) and butoxamine (10 microM) shifted the dose-response curve of adrenaline to the right with new EC50s of 30 nM and 115 nM, respectively. 3. The rapidly rising phase of the SCC response to noradrenaline and adrenaline observed at low chart-speed consisted of a brief and transient retraction followed by a rebound increase in SCC. At a high chart-speed (1 mm s-1), the retraction and rebound phenomenon manifested as a fast initial spike which could be blocked by phentolamine (non-specific alpha-adrenoceptor antagonist) in a dose-dependent fashion. Similar initial spikes were observed when the tissues were stimulated with phenylephrine (alpha-selective agonist) but not with isoprenaline (non-selective beta-agonist) or forskolin (activator of adenylate cyclase). The response of the initial spike triggered by noradrenaline was dose-dependent and the EC50 was 2000 nM.4. The present study showed that the electrogenic chloride secretion by rat epididymis could be stimulated by (alphaxi-, beta131- and beta2-adrenoceptor agonists. The al-mediated response had a faster onset and more transient action than the 3-counterpart. It is postulated that epididymal chloride secretion might be regulated by neural (noradrenaline-mediated) and humoral (adrenaline-mediated) controls and that the stimulus-secretion coupling mechanisms might involve both Ca2+(alpha-mediated response) and adenosine 3':5'-cyclic monophosphate (beta-mediated response) as intracellular second messengers.

Differential accumulation of transcripts for four tomato 1-aminocyclopropane-1-carboxylate synthase homologs under various conditions.

Degenerate oligonucleotide primers corresponding to conserved regions flanking the active-site domain of 1-aminocyclopropane-1-carboxylate (ACC) synthase (EC 4.4.1.14) were used for the polymerase chain reaction (PCR) to amplify DNA fragments from mRNA isolated from tomato fruit and tomato suspension cell culture. Antibodies raised against two conserved peptide sequences (TNPSNPLGTT and SLSKDLGLPGFRVG) were used to screen for positive colonies, after the PCR products were cloned into a Bluescript plasmid and expressed in Escherichia coli. Four distinct cDNA fragments encoding ACC synthase homologs were isolated. While pBTAS1 and pBTAS4 were obtained from fruit mRNA, cell culture mRNA yielded three sequences, pBTAS1, pBTAS2, and pBTAS3. Sequencing of these gene fragments revealed that pBTAS1 and pBTAS4 were identical to those full-length sequences previously reported by Van Der Straeten et al. [Van Der Straeten, D., Van Wiemeersch, L., Goodman, H. & Van Montague, M. (1990) Proc. Natl. Acad. Sci. USA 87, 4859-4863] and Olson et al. [Olson, D. C., White, J. A., Edelman, J., Harkin, R. N. & Kende, H. (1991) Proc. Natl. Acad. Sci. USA 88, 5340-5344] from tomato fruit, whereas pBTAS2 and pBTAS3 represent new sequences. Ribonuclease protection assays were used to examine the expression of these transcripts under three different conditions of enhanced ethylene production--namely, during fruit ripening, in response to mechanical wounding in fruit tissue, and auxin stimulation in vegetative tissue. Transcripts of pBTAS1 accumulated massively during ripening and wounding but only slightly in response to auxin treatment. Although pBTAS4 was associated with fruit ripening, it was unresponsive to auxin treatment in vegetative tissue. In contrast, the expression of pBTAS2 and pBTAS3 was greatly promoted in auxin-treated vegetative tissue but was absent from fruit tissue. While the expression of pBTAS2 was moderately dependent on wounding, pBTAS3 was unresponsive to wounding. These data support the view that ACC synthase is encoded by a multigene family and that the members are differentially expressed in response to developmental, environmental, and hormonal factors.

Induction of 1-aminocyclopropane-1-carboxylate synthase mRNA by auxin in mung bean hypocotyls and cultured apple shoots.

Auxin is known to promote ethylene production in vegetative tissues by increasing the activity of 1-aminocyclopropane-1-carboxylate (ACC) synthase; therefore, we have studied the effect of auxins on ACC synthase mRNA expression. Total RNA was isolated from auxin-incubated cultured apple (Malus sylvestris Mill.) shoots or mung bean (Vigna radiata L.) hypocotyls. These RNAs and a set of oligonucleotide primers corresponding to two conserved amino acid sequences (SNPLGTT and MSSFGLV) found in ACC synthases isolated from other species were used for polymerase chain reaction-based amplification of DNA fragments encoding the ACC synthase-active site domain. We obtained and sequenced a 290-base pair cDNA fragment (pAA1) from cultured apple shoots and a 328-base pair cDNA clone (pMBA1) from mung bean hypocotyls. Comparisons of their deduced amino acid sequences with those of previously characterized ACC synthase cDNAs indicate that both fragments are, indeed, closely related to ACC synthase cDNA. Northern blot analyses further showed that the expression of these transcripts is regulated by auxin treatment. These data indicate that auxin induces ethylene production transcriptionally by increasing the ACC synthase transcripts. The pAA1 shares 46% amino acid sequence homology with ripening-regulated apple fruit ACC synthase, indicating that ripening-regulated and auxin-regulated ACC synthases are encoded by different genes. In mung bean hypocotyls, aminooxyacetic acid, a potent inhibitor of ACC synthase activity, promoted the expression of auxin-induced ACC synthase mRNA, but cycloheximide inhibited this induction.

Purification and characterization of 1-aminocyclopropane-1-carboxylate synthase from apple fruits.

1-Aminocyclopropane-1-carboxylate (ACC) synthase, a key enzyme in ethylene biosynthesis, was isolated and partially purified from apple (Malus sylvestris Mill.) fruits. Unlike ACC synthase isolated from other sources, apple ACC synthase is associated with the pellet fraction and can be solubilized in active form with Triton X-100. Following five purification steps, the solubilized enzyme was purified over 5000-fold to a specific activity of 100 micromoles per milligram protein per hour, and its purity was estimated to be 20 to 30%. Using this preparation, specific monoclonal antibodies were raised. Monoclonal antibodies against ACC synthase immunoglobulin were coupled to protein-A agarose to make an immunoaffinity column, which effectively purified the enzyme from a relatively crude enzyme preparation (100 units per milligram protein). As with the tomato enzyme, apple ACC synthase was inactivated and radiolabeled by its substrate S-adenosyl-l-methionine. Apple ACC synthase was identified to be a 48-kilodalton protein based on the observation that it was specifically bound to immunoaffinity column and it was specifically radiolabeled by its substrate S-adenosyl-l-methionine.

Cloning of a cDNA encoding 1-aminocyclopropane-1-carboxylate synthase and expression of its mRNA in ripening apple fruit.

1-Aminocyclopropane-1-carboxylate (ACC) synthase (EC 4.4.1.14) purified from apple (Malus sylvestris Mill.) fruit was subjected to trypsin digestion. Following separation by reversed-phase high-pressure liquid chromatography, ten tryptic peptides were sequenced. Based on the sequences of three tryptic peptides, three sets of mixed oligonucleotide probes were synthesized and used to screen a plasmid cDNA library prepared from poly(A)(+) RNA of ripe apple fruit. A 1.5-kb (kilobase) cDNA clone which hybridized to all three probes were isolated. The clone contained an open reading frame of 1214 base pairs (bp) encoding a sequence of 404 amino acids. While the polyadenine tail at the 3'-end was intact, it lacked a portion of sequence at the 5'-end. Using the RNA-based polymerase chain reaction, an additional sequence of 148 bp was obtained at the 5'-end. Thus, 1362 bp were sequenced and they encode 454 amino acids. The deduced amino-acid sequence contained peptide sequences corresponding to all ten tryptic fragments, confirming the identity of the cDNA clone. Comparison of the deduced amino-acid sequence between ACC synthase from apple fruit and those from tomato (Lycopersicon esculentum Mill.) and winter squash (Cucurbita maxima Duch.) fruits demonstrated the presence of seven highly conserved regions, including the previously identified region for the active site. The size of the translation product of ACC-synthase mRNA was similar to that of the mature protein on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), indicating that apple ACC-synthase undergoes only minor, if any, post-translational proteolytic processing. Analysis of ACC-synthase mRNA by in-vitro translation-immunoprecipitation, and by Northern blotting indicates that the ACC-synthase mRNA was undetectable in unripe fruit, but was accumulated massively during the ripening proccess. These data demonstrate that the expression of the ACC-synthase gene is developmentally regulated.

Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase.

The pyridoxal phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14), the key enzyme in ethylene biosynthesis, is inactivated by its substrate S-adenosylmethionine (AdoMet). Apple ACC synthase was purified with an immunoaffinity gel, and its active site was probed with NaB3H4 or Ado[14C]Met. HPLC separation of the trypsin digest yielded a single radioactive peptide. Peptide sequencing of both 3H- and 14C-labeled peptides revealed a common dodecapeptide of Ser-Leu-Ser-Xaa-Asp-Leu-Gly-Leu-Pro-Gly-Phe-Arg, where Xaa was the modified, radioactive residue in each case. Acid hydrolysis of the 3H-labeled enzyme released radioactive N-pyridoxyllysine, indicating that the active-site peptide contained lysine at position 4. Mass spectrometry of the 14C-labeled peptide indicated a protonated molecular ion at m/z 1390.6, from which the mass of Xaa was calculated to be 229, a number that is equivalent to the mass of a lysine residue alkylated by the 2-aminobutyrate portion of AdoMet, as we previously proposed. These results indicate that the same active-site lysine binds the PLP and convalently links to the 2-aminobutyrate portion of AdoMet during inactivation. The active site of tomato ACC synthase was probed in the same manner with Ado[14C]Met. Sequencing of the tomato active-site peptide revealed two highly conserved dodecapeptides; the minor peptide possessed a sequence identical to that of the apple enzyme, whereas the major peptide differed from the minor peptide in that methionine replaced leucine at position 6.

Dependence of in vivo ethylene production rate on 1-aminocyclopropane-1-carboxylic Acid content and oxygen concentrations.

1-Aminocyclopropane-1-carboxylic acid (ACC) is aerobically oxidized in plant tissues to form ethylene by ethylene-forming enzyme (EFE). The effect of substrate (ACC and oxygen) concentrations on ethylene production rate by plant tissues was investigated. The K(m) value for O(2) in ethylene production varied greatly depending on the internal ACC content. When ACC levels in the tissue were low (below its K(m) value), the concentration of O(2) giving half-maximal ethylene production rate ([S](0.5)) ranged between 5 and 7%, and was similar among different tissues. As the concentration of ACC was increased (greater than its K(m) value), [S](0.5) for O(2) decreased markedly. In contrast, the K(m) value for ACC was not much dependent on O(2) concentration, but varied greatly among different plant tissues, ranging from 8 micromolar in apple (Malus sylvestris Mill.) tissue to 120 micromolar in etiolated wheat (Triticum aestivum) leaf. Such a great variation was thought to be due to the different compartmentation of ACC within the cells in different tissues. These kinetic data are consistent with the view that EFE follows an ordered binding mechanism in which EFE binds first to O(2) and then to ACC.

Cyanide metabolism in relation to ethylene production in plant tissues.

HCN is the putative product of C-1 and amino moieties of 1-aminocyclopropane-1-carboxylic acid (ACC) during its conversion to ethylene. In apple (Malus sylvestrus Mill.) slices or auxin-treated mungbean (Vigna radiata L.) hypocotyls, which produced ethylene at high rates, the steady state concentration of HCN was found to be no higher than 0.2 micromolar, which was too low to inhibit respiration (reported Ki for HCN to inhibit respiration was 10-20 micromolar). However, these tissues became cyanogenic when treated with ACC, the precursor of ethylene, and with 2-aminoxyacetic acid, which inhibits beta-cyanoalanine synthase, the main enzyme to detoxify HCN; the HCN levels in these tissues went up to 1.7 and 8.1 micromolar, respectively. Although ethylene production by avocado (Persea gratissima) and apple fruits increased several hundred-fold during ripening, beta-cyanoalanine synthase activity increased only one- to two-fold. These findings support the notion that HCN is a co-product of ethylene biosynthesis and that the plant tissues possess ample capacity to detoxify HCN formed during ethylene biosynthesis so that the concentration of HCN in plant tissues is kept at a low level.

The effect of light and phytochrome on 1-aminocyclopropane-1-carboxylic Acid metabolism in etiolated wheat seedling leaves.

While light-grown wheat leaves produced ethylene at a low rate of <0.1 nanomoles per gram per hour and contained 1-aminocyclopropane-1-carboxylic acid (ACC) at low levels of <2.5 nanomoles per gram, etiolated wheat leaves produced ethylene at a rate of 2 nanomoles per gram per hour and accumulated concentrations of ACC at levels of 40 nanomoles per gram. Upon illumination of 8-day-old etiolated wheat seedlings with white light, the ethylene production rate increased initially, due to the activation of ethylene-forming activity, but subsequently declined to a low level (0.1 nanomoles per gram per hour) at the end of the 6-hour illumination. This light-induced decline in ethylene production rate resulted from a decline (more than 35 nanomoles per gram) in ACC level, which was accompanied by a corresponding increase in 1-(malonylamino)cyclopropane-1-carboxylic acid content. These data indicate that illumination promoted ACC malonylation, resulting in reduced ACC level and consequently reduced ethylene production. However, light did not cause any significant increase in the extractable ACC-malonyltransferase activity. The effect of continuous white light on promotion of ACC malonylation was also observed in intermittent white light or red light. A far-red light treatment following red light partially reversed the red light effect, indicating that phytochrome participates in the promotion of ACC malonylation.

Interferences and specificity of the 1-aminocyclopropane-1-carboxylic Acid assay with the hypochlorite reagent.

1-Aminocyclopropane-1-carboxylic Acid (ACC), the immediate precursor of ethylene is routinely assayed by converting it into ethylene with NaOCl, and the ethylene liberated is then determined by gas chromatography (MCC Lizada, SF Yang 1979 Anal Biochem 100: 140-145). However, certain materials which may be present in crude plant extracts or in enzyme reaction mixtures interfere with this assay procedure. Mono, and di-alkyl amines cause poor yields of ethylene from ACC. Ethanol in the presence of NH(3) or amines but in the absence of ACC can produce ethylene under the assay procedure. The characteristics of these interfering reactions were studied and precautions to avoid these problems are suggested. Recovery of ACC during its extraction and purification from plant extracts were tested and are discussed.

Effect of thidiazuron, a cytokinin-active urea derivative, in cytokinin-dependent ethylene production systems.

Cytokinins are known to stimulate ethylene production in mungbean hypocotyls synergistically with indoleacetic acid (IAA), in mungbean hypocotyls synergistically with Ca(2+), and in wilted wheat leaves. Thidiazuron, a substituted urea compound, mimicked the effect of benzyladenine (BA) in all three systems. In the Ca(2+) + cytokinin system and the IAA + cytokinin systems of mungbean hypocotyls, thiadiazuron was slightly more active than BA at equimolar concentration. In mungbean hypocotyls exogenously applied IAA was rapidly conjugated into IAA asparate, and this conjugation process was effectively inhibited by thidiazuron, as by cytokinins. In the wilted wheat leaves system, 10 micromolar thidiazuron exerted stress ethylene production equal to that exerted by 1 millimolar BA, indicating that thidiazuron is more active than BA by two orders. The structure-activity relationship of thidiazuron and its thiadiazolylurea analogs in stimulating Ca(2+)-dependent ethylene production in mungbean hypocotyls was found to agree well with the structure-activity relationship of these derivatives in promoting the growth of callus tissues. These results indicate that thidiazuron and its derivatives are highly active to mimic the adenine-type cytokinin responses in promoting ethylene production and that the structure-activity relationship in promoting the growth of callus and in promoting ethylene production is similar.

Central haemangioma of the mandible.

The literature on central haemangioma of the jaws has been extensively reviewed. The clinical and radiographic features of central haemangiomas of the jaws and the problems in diagnostic and various modalities of treatment of this lesion are discussed. Central haemangioma of the jaws is rare. Up till 1975, only 53 cases have been reported in the literature. A classical case of central haemangioma of the mandible is presented to illustrate the features of this uncommon disease.