A Histological Analysis of Regeneration in Watermelon

The optimization of regeneration protocol for different genotypes increases the yield in transformation studies. Cotyledon explants of watermelon [Citrullus lanatus (Thunb) Matsum & Nakai] cv Crimson Sweet were cultured on MS medium containing combinations of benzyl adenine (BA) (0, 5, 10, 20 µM) and indole-3-acetic acid (IAA) (0, 0.5, 5 µM). Maximum shoot growth and subsequent rooting from explants on regeneration medium were obtained from the media containing 10 µM BA + 0.5 µM IAA and 20 µM BA (75 and 78%) by direct organogenesis, respectively. Histological analysis showed that cell division was observed in the epidermal and subepidermal layers. Protuberant structures were observed in tissues between 7 and 12 days in culture. Meristematic structures were observed after 12days in culture which later developed into buds.     

Cytotoxicity of a novel lipid-like bacterial product

A highly cytotoxic, lipid-like compound was isolated from a Serratia marcescens strain currently under identification. We have named the compound DCX for its direct cytotoxic activity on various cell types in culture. DCX was purified by preparative thin layer chromatography from chloroform: methanol = 4:1 extracts of whole bacteria, and is chromatographically homogeneous. The effect of DCX on cells is dose, time, and temperature dependent. DCX is particularly toxic to the mastocytoma cell line P815 (TD50 = 75 pg/ml). Three other malignant or transformed murine cell lines were sensitive to the cytotoxic action of DCX. The effect of DCX was also tested on normal cells (human gingival fibroblasts), which showed greater resistance to DCX than the other cells tested.     

Aluminium causes nutrient imbalance and structural changes in the needles of Scots pine without inducing clear root injuries

The effects of aluminium chloride (AICI3) treatments (50 and 150 mg/l) on 3-year-old Scots pine (Pinus sylvestris L.) seedlings were studied in a sand culture during 2 growing periods in an open field experiment. Even by the end of the first growing period, a decline was observed in the concentrations of Ca, Mg and P within the needles, and of Ca and Mg in the roots. After the second growing period, increased N and K concentrations were observed in the needles of Al-treated seedlings. Both the needles and roots of Al-treated seedlings showed, after the second growing period, a decline in growth and increased concentrations of AI as the amount of AICI3 in the nutrient solution increased. Al-induced changes in needle structure were found to be symptomatic of a nutrient imbalance, particularly of Mg and P. Al-stress did not result in any observable changes in root anatomy or in the number of mycorrhizas. Scots pine proved to be rather resistant to Al-stress, indicating that direct Al-injuries are not likely in the field, though Al-stress may be a contributing factor in the formation of nutrient imbalances.     

Different states of sarcoplasmic reticulum membrane in the presence of acetyl phosphate and adenosine triphosphate.

In the presence of acetyl phosphate, approximately 0.8 extra sulphydryl groups/10⁵ g protein of sarcoplasmic reticulum membrane vesicles are exposed to reaction with $\text{N}$-ethylmaleimide, whereas in the presence of ATP approximately 0.6 groups/10⁵ g protein are protected. Dithiobis (nitrobenzoic acid) reacts with the membrane sulphydryl groups more slowly in the presence of ATP than in the presence of acetyl phosphate or in the absence of substrate. Sarcoplasmic reticulum membrane is degraded by trypsin at a faster rate than normal when acetyl phosphate is present as seen from changes in electrophoretic patterns, ATPase activity and Ca²⁺ uptake capacity, and at a slower rate when ATP is present as seen from the last two properties. These differences in reactivity are interpreted as being due to differences in membrane conformations induced by the two substrates.     

Ammonia induces amyloidogenesis in astrocytes by promoting amyloid precursor protein translocation into the endoplasmic reticulum

Hyperammonemia is known to cause various neurological dysfunctions such as seizures and cognitive impairment. Several studies have suggested that hyperammonemia may also be linked to the development of Alzheimer’s disease (AD). However, the direct evidence for a role of ammonia in the pathophysiology of AD remains to be discovered. Herein, we report that hyperammonemia increases the amount of mature amyloid precursor protein (mAPP) in astrocytes, the largest and most prevalent type of glial cells in the central nervous system that are capable of metabolizing glutamate and ammonia, and promotes amyloid beta (Aβ) production. We demonstrate the accumulation of mAPP in astrocytes was primarily due to enhanced endocytosis of mAPP from the plasma membrane. A large proportion of internalized mAPP was targeted not to the lysosome, but to the endoplasmic reticulum, where processing enzymes β-secretase BACE1 (beta-site APP cleaving enzyme 1) and γ-secretase presenilin-1 are expressed, and mAPP is cleaved to produce Aβ. Finally, we show the ammonia-induced production of Aβ in astrocytic endoplasmic reticulum was specific to Aβ42, a principal component of senile plaques in AD patients. Our studies uncover a novel mechanism of Aβ42 production in astrocytes and also provide the first evidence that ammonia induces the pathogenesis of AD by regulating astrocyte function.     

Role of MGMT in protecting against cyclophosphamide-induced toxicity in cells and animals

O(6)-Methylguanine-DNA methyltransferase (MGMT) is a DNA repair protein that protects cells from the biological consequences of alkylating agents by removing alkyl groups from the O(6)-position of guanine. Cyclophosphamide and ifosfamide are oxazaphosphorines used clinically to treat a wide variety of cancers; however, the role of MGMT in recognizing DNA damage induced by these agents is unclear. In vitro evidence suggests that MGMT may protect against the urotoxic oxazaphosphorine metabolite, acrolein. Here, we demonstrate that Chinese hamster ovary cells transfected with MGMT are protected against cytotoxicity following treatment with chloroacetaldehyde (CAA), a neuro- and nephrotoxic metabolite of cyclophosphamide and ifosfamide. The mechanism by which MGMT recognizes damage induced by acrolein and CAA is unknown. CHO cells expressing a mutant form of MGMT (MGMT(R128A)), known to have >1000-fold less repair activity towards alkylated DNA while maintaining full active site transferase activity towards low molecular weight substrates, exhibited equivalent CAA- and acrolein-induced cytotoxicity to that of CHO cells transfected with plasmid control. These results imply that direct reaction of acrolein or CAA with the active site cysteine residue of MGMT, i.e. scavenging, is unlikely a mechanism to explain MGMT protection from CAA and acrolein-induced toxicity. In vivo, no difference was detected between Mgmt-/- and Mgmt+/+ mice in the lethal effects of cyclophosphamide. While MGMT may be important at the cellular level, mice deficient in MGMT are not significantly more susceptible to cyclophosphamide, acrolein or CAA. Thus, our data does not support targeting MGMT to improve oxazaphosphorine therapy.     

Vegetation feedbacks of nutrient addition lead to a weaker carbon sink in an ombrotrophic bog

To study vegetation feedbacks of nutrient addition on carbon sequestration capacity, we investigated vegetation and ecosystem CO₂ exchange at Mer Bleue Bog, Canada in plots that had been fertilized with nitrogen (N) or with N plus phosphorus (P) and potassium (K) for 7–12 years. Gross photosynthesis, ecosystem respiration, and net CO₂ exchange were measured weekly during May–September 2011 using climate‐controlled chambers. A substrate‐induced respiration technique was used to determine the functional ability of the microbial community. The highest N and NPK additions were associated with 40% less net CO₂ uptake than the control. In the NPK additions, a diminished C sink potential was due to a 20–30% increase in ecosystem respiration, while gross photosynthesis rates did not change as greater vascular plant biomass compensated for the decrease in Sphagnum mosses. In the highest N‐only treatment, small reductions in gross photosynthesis and no change in ecosystem respiration led to the reduced C sink. Substrate‐induced microbial respiration was significantly higher in all levels of NPK additions compared with control. The temperature sensitivity of respiration in the plots was lower with increasing cumulative N load, suggesting more labile sources of respired CO₂. The weaker C sink potential could be explained by changes in nutrient availability, higher woody : foliar ratio, moss loss, and enhanced decomposition. Stronger responses to NPK fertilization than to N‐only fertilization for both shrub biomass production and decomposition suggest that the bog ecosystem is N‐P/K colimited rather than N‐limited. Negative effects of further N‐only deposition were indicated by delayed spring CO₂ uptake. In contrast to forests, increased wood formation and surface litter accumulation in bogs seem to reduce the C sink potential owing to the loss of peat‐forming Sphagnum.