The Pichia pastoris HIS4 gene: nucleotide sequence,creation of a non-reverting his4 deletion mutant,and development of HIS4-based replicating and integrating plasmids

We have obtained a clone of the Pichia pastoris HIS4 gene and have determined its nucleotide sequence. Based upon its deduced amino-acid sequence, the product of the P. pastoris HIS4 gene has the same structural organization as the Saccharomyces cerevisiae His4 protein and appears to encode a trifunctional enzyme catalyzing the second (phosphoribosyl-ATP pyrophosphohydrolase), third (phosphoribosyl-AMP cyclohydrolase), and tenth (histidinol dehydrogenase) steps in histidine biosynthesis. The chromosomal copy of the HIS4 gene was disrupted by homologous recombination, creating the strain SGY58. The his4 deletion mutation in this strain lacks the entire coding region of this gene and has a reversion rate that is undetectable. A set of complementary plasmids that carry the HIS4 gene was also developed. Among these are nine E. coli-P. pastoris shuttle vectors that transform the His4 deletion mutant at high efficiency and an integration vector for creating site-specific alterations of the P. pastoris genome.     

Ovulation triggering in clomiphene citrate-stimulated cycles: Human chorionic gonadotropin versus a gonadotropin releasing hormone agonist

Purpose To compare the use of human chorionic gonadotropin (hCG) to a gonadotropin releasing hormone (GnRH) agonist, nafarelin, in initiating ovulation and supporting the luteal phase after priming with clomiphene.Methods In 26 infertile women 50 mg clomiphene citrate produced a preovulatory-size follicle. Then, 11 women were randomized to receive two 400-g doses of nafarelin intranasally 16 h apart, and 15 women were injected intramuscularly with 5000 IU of hCG (luteal day 0 = LD0). Starting on LD6, 7 more 400-g doses of nafarelin were repeated on an every 16-h schedule or a single 2500 IU dose of hCG was given, respectively. Serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E₂), progesterone (P), and hCG were measured. On LD13, endometrium was evaluated with ultrasonography and biopsy in 19 nonpregnant women.Results As judged by a threefold rise in serum LH, an LH surge was detected on LD1 in all 11 nafarelin patients, but in only 8 hCG patients (P = 0.01). LH and FSH levels were significantly higher on LD1, 7, and 8 and were significantly suppressed on LD13 in the nafarelin group. All patients had mid-luteal P levels greater than 10 ng/ml and luteal phases longer than 13 days. Significantly different luteal E₂ or P levels were noted only on LD13, with lower values in the nafarelin group. Pregnancies were achieved in 3 of 11 nafarelin cycles and 2 of 15 hCG cycles. Luteal phase defects were also similar: 4 of 8 nafarelin patients and 7 of 11 hCG patients.Conclusion Nafarelin or hCG in conjunction with clomiphene can result in viable pregnancies, but is associated with low pregnancy rates and a high incidence of luteal phase defects.     

Self-Directed Treatment of Intermittent Heartburn: A Randomized, Multicenter, Double-Blind, Placebo-Controlled Evaluation of Antacid and Low Doses of an H(2)-Receptor Antagonist (Famotidine)

BACKGROUND: Heartburn, a common symptom, is self-treated with oral antacids. Efficacy of antacids has not been demonstrated for individual, spontaneous heartburn episodes. METHODS: We conducted a double-blind, randomized, placebo-controlled, parallel-group study of self-directed treatment for episodic heartburn comparing famotidine (FAM) 5, 10, or 20 mg and antacid (11 mEq ANC) to placebo (PBO) during a 4-week period. Twenty-nine US investigators enrolled a total of 565 outpatients, ages 18--81 years (mean 44.1 years) with heartburn but not seeking care for heartburn. Treatment of spontaneous heartburn episodes was permitted as needed, up to twice daily, with self-administered test drug. An open-label, backup antacid was provided to use if test drug did not provide adequate relief. Patients assessed heartburn relief hourly and recorded use of backup antacid. Relief was defined as complete relief of symptoms without the use of backup antacid. RESULTS: The media proportion of episodes relieved was: PBO, 41%; FAM 5 mg, 59%, 0.05 less-than-or-equal p < 0.10; FAM 10 mg, 70%, p < 0.001; FAM 20 mg, 69%, p < 0.001; antacid, 62%, p < 0.05 (p-values versus PBO). Supplemental analyses incorporating time to relief confirmed that famotidine and antacid provided more rapid and more frequent relief than placebo (odds ratio for relief relative to PBO: FAM 5 mg, 1.55, p = 0.003; FAM 10 mg, 1.94, p < 0.001; FAM 20 mg, 2.13, p < 0.001; antacid 1.57, p = 0.003). The tolerability profile was similar with famotidine, antacid, and placebo. CONCLUSIONS: The positive results with antacid demonstrated for the first time the efficacy of antacid in self-treatment of individual heartburn episodes and provided internal validation of this study paradigm. Patients in this study self-medicated effectively using low doses of famotidine on an as needed basis for spontaneous episodes of heartburn.     

Comparison of the polyvinyl alcohol sponge and expanded polytetrafluoroethylene subcutaneous implants as models to evaluate wound healing potential in human beings

Our current understanding of the complex processes involved in wound healing is based mainly on studies of animal models. Although this information has been useful, it may not totally reflect the response found in human beings. For example, human beings have a tendency to either "overheal," as seen in keloids and hypertrophic scar formation, or have deficient healing, as seen in chronic ulcer formation. No animal models are available to analyze these human clinical pathologic conditions. Therefore the objective of this study was to analyze the wound healing response in a large population (n = 40) of normal healthy human beings as a first step to begin studies of abnormal human wound healing. Simultaneously, a comparison was made between the polyvinyl alcohol implant and the expanded polytetrafluoroethylene implant model. Under sterile conditions with the use of local anesthesia, two preweighed polyvinyl alcohol implants and two standard 6 cm expanded polytetrafluoroethylene implants were placed subcutaneously in the upper arm of each subject. High-performance liquid chromatography was used to quantitate isoleucine and hydroxy-l-proline in acid hydrolysates of each implant. Isoleucine was used as an indicator of protein content in the tissue sample, whereas hydroxyproline reflected collagen content. No infectious or hemorrhagic complications were found in the 40 volunteers included in the study. No significant difference was found in isoleucine or hydroxy-l-proline content between postoperative day 7 polyvinyl alcohol implants and day 14 polyvinyl alcohol implants. In contrast, both isoleucine and hydroxy-l-proline content were significantly increased in day 14 expanded polytetrafluoroethylene implants compared with day 7 implants (p < 0.005 and p < 0.001, respectively). In addition, the ratio of hydroxy-l-proline to isoleucine was significantly increased in day 14 expanded polytetrafluoroethylene implants compared with day 7 expanded polytetrafluoroethylene and both day 7 and day 14 polyvinyl alcohol implants (p < 0.001). This observation suggests that by 14 days implantation of expanded polytetrafluoroethylene stimulated an increased deposition of collagen. No significant differences were found in the hydroxy-l-proline to isoleucine ratios among day 7 expanded polytetrafluoroethylene, day 7 polyvinyl alcohol, and day 14 polyvinyl alcohol implants. Histologic analyses correlated with the biochemical findings. These results suggest that expanded polytetrafluoroethylene may be the preferred implant for studies designed to examine pathologic processes associated with retarded wound healing. In contrast, the polyvinyl alcohol implant may be better suited for studies where a low background response is required. Moreover, the extreme variability in normal healthy volunteers seen in this study correlates clinically with the finding that, among the normal adult human population, there is a heterogeneous wound healing response.     

Antibiotic resistance in nosocomial isolates in Scotland

In this first multi-centre study in Scotland, 1028 consecutive Gram-negative and staphylococci strains were obtained from four major teaching hospitals. E. coli was the most common organism among both intensive care units (ICUs) (39%) and non-ICU strains (46.6%). The prevalence of antibiotic resistance among E. coli was always higher in isolates from ICUs than non-ICUs: ceftazidime (14.1%, 7.2%), ceftriaxone (12.7%, 6.1%), cefotaxime (15.5%, 8.7%), cefuroxime (28.8%, 20.8%), amoxicillin plus clavulanic acid (52.2%, 38.8%) and gentamicin (7.0%, 2.8%). The highest incidences of resistance were identified among Enterobacter/Citrobacter spp. from ICUs; 43.8%, 41.7%, 45.8%, 54.2%, 87.5% and 10.4% of these organisms were resistant to ceftazidime, ceftriaxone, cefotaxime, cefuroxime, amoxicillin plus clavulanic acid and gentamicin, respectively.     

Trunk Testing Using a Prototype Cybex II lsokinetic Dynamometer Stabilization System

This pilot study represents Cybex II isokinetic dynamometer trunk testing on 160 subjects (98 males, 62 females). A prototype apparatus which provides stabilization in the standing (functional) position was used in the testing. The testing protocols were performed first for trunk flexion and then trunk extension. The protocols consisted of two isometric tests for 5 seconds at 0 degrees trunk angle and 45 degrees trunk angle followed by three repetitions each at the isokinetic speeds of 30, 60, 90, and 120 degrees per second. Peak torque values were measured for the isometric tests. lsokinetic data interpretation consisted of: peak torque, time rate of tension development, range of motion where peak torque occurred, and total work performed. The ratios of trunk flexors to extensors at each test were also calculated. The conclusion from this pilot study is that the use of an isokinetic testing of trunk musculature and establishing normative data provide clinically useful guidelines for sports screening, industrial medicine screening, and objective parameters for discharging patients with trunk dysfunction. J Orthop Sports Phys Ther 1982;3(4):164-170.     

Strontium-90 in newborns and childhood disease

Radioactive strontium-90 concentrations in baby teeth obtained from Suffolk County, New York, rose steadily during the 1980s. Recent levels of strontium-90 are similar to those reported for babies born in the late 1950s-at the height of atmospheric nuclear weapons testing in Nevada. Strontium-90 concentrations increased concomitantly with increases in cancer incidence among Suffolk children under the age of 5 y, a result that mimicked parallel trends observed in the 1950s and early 1960s. Given that effects of strontium-90 on developing cells are most pronounced during the fetal and infant periods, escalating levels should be viewed as a factor in the recent decline in various child health status measures.     

Antioxidant-rich diets improve cerebellar physiology and motor learning in aged rats

The free radical theory of aging predicts that reactive oxygen species are involved in the decline in function associated with aging. The present paper reports that diets supplemented with either spinach, strawberries or blueberries, nutritional sources of antioxidants, reverse age-induced declines in beta-adrenergic receptor function in cerebellar Purkinje neurons measured using electrophysiological techniques. In addition the spinach diet improved learning on a runway motor task, previously shown to be modulated by cerebellar norepinephrine. Motor learning is important for adaptation to changes in the environment and is thus critical for rehabilitation following stroke, spinal cord injury, and the onset of some neurodegenerative diseases. These data are the first to indicate that age-related deficits in motor learning and memory can be reversed with nutritional interventions.