Cryopreservation of Galanthus elwesii Hook. apical meristems by droplet vitrification.

The aim of this study was to develop an efficient cryopreservation protocol for the geophyte giant snowdrop (Galanthus elwesii Hook.) that guarantees a high rate of survival and plant regeneration after cryopreservation. The excised apical meristems were obtained from cultures of in vitro grown bulb scales. Using a vitrification procedure and optimizing the duration of the exposure to the loading solution (LS), meristem post-rewarm survival rates higher than 90 percent were achieved. Also regrowth percentages were very high, ranging from 87 to 91 percent. After optimizing the time of exposure to the plant vitrification solution (PVS2), the survival rate was between 83 and 97 percent. During post-rewarm regeneration, good growth recovery was as high as 76 percent; however, hyperhydration and callusing were also observed. The results demonstrate that cryopreservation of Galanthus elwesii germplasm seems to be feasible.

Hepatic iron accumulation over time in European starlings (Sturnus vulgaris) fed two levels of iron.

European starlings (Sturnus vulgaris) were used as a passerine bird model to examine the effect of dietary iron on the level of hepatic iron in birds. Nestling and fledgling starlings (n = 56) were raised on a controlled-iron diet. When birds maintained constant body weight, they were assigned in pairs to cages, and baseline sampling was performed. Pairs were then assigned to one of two diets: the controlled-iron diet (168 ppm, dry basis) or a high-iron diet (3,035 ppm, dry basis). Dry-matter intake and iron consumption were recorded. Dry-matter intake did not differ between the dietary treatment groups and was stable during treatment periods. Iron intake was higher in the high-iron group (P < 0.05). Birds were euthanized at baseline, 8 wk, and 16 wk. Body, liver, and spleen weights were measured. Hepatic iron and copper concentrations were determined. Body weight did not differ between the two treatment groups or among individuals for the study duration. Liver iron concentration differed over time and between treatment groups. Birds receiving both treatments had similar liver iron content at week 8 (3,107 +/- 228.6 ppm and 3,122 +/- 306.2 ppm high and controlled iron, respectively; P > 0.05), but by week 16, birds consuming the high-iron diet had greater hepatic iron levels than those consuming the controlled-iron diet (5,929 +/- 937.2 ppm and 3,683 +/- 229.5 ppm high and controlled iron, respectively; P < 0.05). Birds on the controlled-iron diet also had higher hepatic iron at 16 wk than at 8 wk. Liver copper decreased over time in all birds regardless of treatment. Results show that both dietary iron level and duration of time influenced hepatic iron storage. The controlled-iron diets still allowed accumulation of hepatic iron in an 8-wk period.

Probing interactions between viral DNA and human immunodeficiency virus type 1 integrase using dinucleotides.

Retroviral integrases are essential for viral replication and represent an attractive chemotherapeutic target. In the current study, we demonstrated the activity of micromolar concentrations of dinucleotides against human immunodeficiency virus type 1 (HIV-1), HIV type 2 (HIV-2), simian immunodeficiency virus, and feline immunodeficiency virus integrases. The structure-activity relationship indicates that 5'-phosphorylation enhances potency and that phosphodiester and sugar modifications affect the inhibition of HIV-1 integrase. Base sequence selectivity was observed: pAC, pAT, and pCT were the most potent inhibitors, whereas pAA, pGA, and pGC showed low activity at 100 microM. The inhibition by pAC is consistent with the interaction of the enzyme with the 5' end of the noncleaved strand (5'-AC-3'). The linear and cyclic dinucleotides released by the 3'-processing reaction did not affect enzymatic activity at physiological concentrations. An increase in the length to trinucleotides or tetranucleotides enhanced potency by only 2-3-fold, suggesting that two neighboring bases may be sufficient for significant interactions. Inhibition of a truncated (50-212) integrase mutant and global inhibition of all nucleophiles in the 3'-processing reaction suggest that dinucleotides bind in the catalytic core. All of the active dinucleotides inhibited enzyme/DNA binding in their respective IC50 range. Although the dinucleotides tested showed no antiviral activity, these observations demonstrate the usefulness of dinucleotides in elucidating enzyme mechanisms and as potential ligands for cocrystallization and as lead structures for development of antivirals.

Studies on stereospecific formation of P-chiral internucleotide linkage: synthesis of all-Rp and all-Sp methylphosphonate pentanucleotide d(MMTrAPMeTPMeTPMeCPMeTAc) via Grignard activated coupling.

Synthesis of stereoregular 3',5'-protected all-Rp and all-Sp methylphosphonate heterooligonucleotides d(MMTrAPMeTPMeTPMeCPMeTAc) (12) complementary to the fragment of HIV-1 splicing acceptor site was achieved via stereo-controlled formation of internucleotide linkage. The coupling was based on the transesterification of P-stereodefined monomer type of 5'-O-monomethoxytrityl-2'-O-deoxynucleoside 3'-O-[O-(4-nitrophenyl)methylphosphonate]. The nucleophile was a t-butylmagnesium chloride activated 5'-terminal hydroxyl function of the growing oligonucleotide chain. This and other P-homochiral oligomers will be used as building blocks for the synthesis of biologically significant, longer stereoregular oligonucleotides.

Ketorolac versus meperidine for pain relief after orthopaedic surgery.

In this double-blind, randomized, multicenter study, 244 patients with at least moderate pain after major orthopaedic surgery received intramuscular Ketorolac (60 mg followed by 30 mg) or intramuscular meperidine (100 mg or placebo) every 2 to 6 hours as needed for as many as 5 days. Analgesic response was evaluated for 6 hours after initial study drug administration and thereafter each night at bedtime. Both active treatment groups had similar 3-hour summed pain intensity difference and 3-hour total pain relief scores after the first dose that were superior to placebo. The 6-hour summed pain intensity difference and total pain relief scores were significantly higher with Ketorolac than with meperidine or placebo. The mean daily categorical pain intensity scores were comparable with Ketorolac and meperidine, and both were significantly superior to placebo. Patient ratings of overall medication efficacy were significantly better with Ketorolac than with meperidine. In both patient and observer evaluations, Ketorolac was significantly better tolerated than meperidine, and the number of patients reporting adverse events was lower with Ketorolac than with meperidine. Following major orthopaedic surgery, Ketorolac provided effective analgesia that was superior to placebo and at least comparable with meperidine. Ketorolac was better tolerated than meperidine.

Additives containing bacteria and enzymes for alfalfa silage.

First-cutting alfalfa was wilted, harvested from alternate rows, left untreated or treated with additives containing lactic acid bacteria and enzymes (cellulase, amylase, and pectinase), and ensiled in bag silos. Inoculation increased lactic acid bacteria from 5 x 10(4) to 1 x 10(6) cfu/g of forage. Because treatments were bagged consecutively, the DM of treated silages was higher than that of untreated silage. However, after 4 d of ensiling, the pH of treated silage, about 4.3, was lower than that of untreated silage, 4.7, and remained lower throughout the ensiling period. After 177 d of ensiling, total lactate was about 25% higher, and ammonia N was about 40% lower, in treated silage. In addition, NDF and ADF contents were lower in treated than in untreated silage. Between 51 and 177 d of storage, glucose content increased in treated silage, but not in untreated silage, suggesting that some plant cell-wall hydrolysis occurred during prolonged storage. In vitro digestion of NDF did not differ among treatments during early incubation, but the extent of digestion after 36 and 48 h was lower in treated than in untreated silage. The microbial and enzyme silage additives used in this study improved fermentation characteristics and reduced fiber content of silage but decreased the in vitro digestibility of fiber.

Comparison of oral ketorolac, intramuscular morphine, and placebo for treatment of pain after orthopedic surgery.

Using a randomized, double-blind, placebo-controlled, parallel, single-dose, single-center, 6-hour study, we compared the analgesic response and tolerability of oral ketorolac tromethamine and intramuscular morphine sulfate and placebo. The study group comprised 176 patients with moderate, severe, or very severe pain after hip or knee surgery at a teaching hospital. Patients received either 10 mg of ketorolac orally, 10 mg of morphine intramuscularly, 5 mg of morphine IM, or placebo. Patients rated pain intensity at baseline and pain intensity and pain relief at 30 minutes, 1 hour, and hourly thereafter for 6 hours. At study completion, we evaluated overall patient ratings of pain relief and occurrence of adverse events. Summed pain intensity difference scores and total pain relief scores showed the active medications to be significantly superior to placebo and not significantly different from each other. The 10-mg dose of morphine showed a small advantage over ketorolac in peak analgesic effect, but the onset of pain relief was comparable among the active agents. The incidence of adverse events among the active-treatment groups was similar, though there was a numerical trend favoring ketorolac over 10 mg of morphine. We found oral ketorolac to be an effective alternative to parenteral opioids for the treatment of pain after hip or knee surgery in patients who can tolerate oral medication.

The use of ketorolac in the management of postoperative pain.

Ketorolac tromethamine (Toradol) is a nonsteroidal antiinflammatory drug (NSAID) available in intramuscular (IM) and oral formulations for the management of acute pain. Intramuscular ketorolac is the only parenteral NSAID available for analgesic use in the US. The clinical profile is reviewed, and clinical studies most applicable to a postoperative patient are discussed in detail. The results of a clinical study performed at Emory University School of Medicine are presented. In this single-dose study, 176 patients received either 10 mg of oral ketorolac, 5 mg or 10 mg of IM morphine, or placebo after orthopedic surgery. The analgesic efficacy of ketorolac was comparable to both doses of morphine and significantly superior to placebo. Ketorolac, when administered intramuscularly or orally, is a safe and effective analgesic agent for the short-term management of acute postoperative pain and can be used as an alternative to opioid therapy.