Functional aspects of the superoxide dismutative action of Cu-penicillamine.

The superoxide dismutative action of Cu-penicillamine was examined by pulse radiolysis. The second order rate constand of the reaction wpith superoxide was 0.4 +/- o.2.10(9) M-1.s-1, comparable to the action of Fe and Mn-superoxide dismutases. No marked pH-dependence was seen. Neither ethylene diamine tetraacetic acid nor cyanide affected the catalytic action of Cu-penicillamine. The cyanide resistant reactivity as well as further X-ray photoelectron spectrometric measurements supported the suggestion of a Cu(I) stabilized sulphur radical being the active species involved in the catalysis of superoxide dismutation.     

Investigation of the cell structure and organelles during autolytic PCD of Antirrhinum majus “Legend White” petals

Protoplasma - One of the classes of the plant developmental programmed cell death (PCD) is vacuolar cell death or autolysis. The results of the transmission electron microscope (TEM) studies...     

High-performance liquid chromatographic determination of cotinine in urine in isocratic mode

A simple procedure for the determination of cotinine, major metabolite of nicotine in urine, is described. The assay involved a liquid–liquid extraction with dichloromethane in alkaline environment. The extract was dried at ambient temperature under a gentle stream of nitrogen. The residue was dissolved in 300 μl of mobile phase and 30 μl aliquot was injected via an automatic sampler into the liquid chromatograph and eluted with the mobile phase (10–9%, v/v methanol and acetonitrile, respectively in potassium dihydrogenphosphate buffer adjusted to pH 3.4) at a flow rate of 1 ml/min on a C₈ Symmetry cartridge column (5 μm, 150 mm×3.9 mm, Waters) at 25°C. The eluate was detected at 260 nm. Internal standard was 2-phenylimidazole. Sensitive and specific, this technique was performed to test urine of diabetic patients (smokers and non-smokers) admitted in an endocrinology service. Urinary cotinine seems to be a better marker of smoking status than thiocyanates.     

Broad-range PCR,cloning and sequencing of the full 16S rRNA gene for detection of bacterial DNA in synovial fluid samples of Tunisian patients with reactive and undifferentiated arthritis

Introduction

Broad-range rDNA PCR provides an alternative, cultivation-independent approach for identifying bacterial DNA in reactive and other form of arthritis. The aim of this study was to use broad-range rDNA PCR targeting the 16S rRNA gene in patients with reactive and other forms of arthritis and to screen for the presence of DNA from any given bacterial species in synovial fluid (SF) samples.

Methods

We examined the SF samples from a total of 27 patients consisting of patients with reactive arthritis (ReA) (n = 5), undifferentiated arthritis (UA) (n = 9), rheumatoid arthritis (n = 7), and osteoarthritis (n = 6) of which the latter two were used as controls. Using broad-range bacterial PCR amplifying a 1400 bp fragment from the 16S rRNA gene, we identified and sequenced at least 24 clones from each SF sample. To identify the corresponding bacteria, DNA sequences were compared to the EMBL (European Molecular Biology Laboratory) database.

Results

Bacterial DNA was identified in 20 of the 27 SF samples (74, 10%). Analysis of a large number of sequences revealed the presence of DNA from more than one single bacterial species in the SF of all patients studied. The nearly complete sequences of the 1400 bp were obtained for most of the detected species. DNA of bacterial species including Shigella species, Escherichia species, and other coli-form bacteria as well as opportunistic pathogens such as Stenotrophomonas maltophilia and Achromobacter xylosoxidans were shared in all arthritis patients. Among pathogens described to trigger ReA, DNA from Shigella sonnei was found in ReA and UA patients. We also detected DNA from rarely occurring human pathogens such as Aranicola species and Pantoea ananatis. We also found DNA from bacteria so far not described in human infections such as Bacillus niacini, Paenibacillus humicus, Diaphorobacter species and uncultured bacterium genera incertae sedis OP10.

Conclusions

Broad-range PCR followed by cloning and sequencing the entire 16S rDNA, allowed the identification of the bacterial DNA environment in the SF samples of arthritic patients. We found a wide spectrum of bacteria including those known to be involved in ReA and others not previously associated with arthritis.     

The emu oil emulsified in egg lecithin and butylated hydroxytoluene enhanced the proliferation,stemness gene expression,and in vitro wound healing of adipose-derived stem cells

In recent decades, mesenchymal stem cells originated from adipose tissue (adipose-derived stem cells, ASCs) have gained increased attention for production of cell-based therapeutics. Emu oil as a natural compound showed antioxidant effects in previous studies. The goal of this study was to investigate the effect of crude emu oil on the proliferation, cell cycle progression, stemness genes expression, and in vitro wound healing potential of ASCs. An emulsion of emu oil was prepared using egg lecithin and butylated hydroxytoluene to improve bioavailability and solubility of emu oil in the expansion medium. The ASCs were treated using a series of emu oil concentrations in emulsion form, diluted in expansion medium (0.03–3 mg/ml). The emu oil-free emulsion was used as control treatment. The results revealed that emu oil (1.25 mg/ml) in emulsion form significantly (p?<?0.001) increased ASCs proliferation and colony formation. Additionally, emu oil caused upregulation of stemness marker genes (Sox2, Oct4, Nanog, and Nestin) (p?<?0.05). The cell cycle analysis after emu oil treatments showed an increase in the population of ASCs in S-phase of the cell cycle. Besides, an accelerated in vitro scratch wound healing was observed in emu oil-treated ASCs. Emu oil enhanced proliferation, colony formation, stemness genes expression, and in vitro wound healing of ASCs. These findings suggest that emu oil treatment could maintain the stemness of ex vivo cultivated ASCs and enhance their regenerative potential.     

Cholecystokinin-induced residual stimulation of enzyme secretion from mouse pancreatic acini

When dispersed acini from mouse pancreas are first incubated with cholecystokinin octapeptide, washed and then reincubated with no additions there is significant stimulation of amylase secretion during the second incubation (residual stimulation of enzyme secretion). Cholecystokinin-induced residual stimulation of enzyme secretion is modified, but not abolished, by reducing the temperature of the first incubation from 37 degrees C to 4 degrees C. Measurement of binding of 125I-labeled cholecystokinin octapeptide indicated that maximal cholecystokinin induced residual stimulation of enzyme secretion occurs when 12-20% of cholecystokinin receptors are occupied by cholecystokinin octapeptide. Moreover, maximal cholecystokinin-induced residual stimulation of amylase secretion is 25% greater than maximal cholecystokinin-induced direct stimulation of amylase secretion. Cholecystokinin tetrapeptide, which causes the same maximal direct stimulation of amylase secretion as does cholecystokinin octapeptide, causes a maximal residual stimulation of enzyme secretion that is only 30% of that caused by a maximally effective concentration of cholecystokinin octapeptide. Adding dibutyryl cyclic GMP to the second incubation can reverse the residual stimulation caused by adding cholecystokinin to the first incubation. The pattern and extent of the dibutyryl cyclic GMP-induced reversal of residual stimulation varies, depending on the temperature and concentration of cholecystokinin octapeptide in the first incubation. The present results are compatible with the hypothesis that mouse pancreatic acini possess two classes of cholecystokinin receptors. One class has a relatively high affinity for cholecystokinin and produces stimulation of enzyme secretion; the other class has a relatively low affinity for cholecystokinin and produces inhibition of enzyme secretion.