Industrial control of recombinant E. coli fed-batch culture: new perspectives on traditional controlled variables.

Three industrially relevant fed-batch algorithms were applied to the production of recombinant proteins in Escherichia coli BL21 DE3. Starvation dissolved-oxygen (DO)-transient control sustained growth rates greater than those in pH-stat (0.16 h(-1) versus 0.11-0.13 h(-1)) while feed-up DO-transient control better tracked the measured threshold for acetate production (mu approximately 0.2 h(-1)). All controllers supported growth without acetate production, resulting in end concentrations of recombinant protein up to 20 times greater than in batch culture. Both DO-transient control systems were judged superior to pH-stat for their ability to detect and track the acetate threshold. Results also showed that although high cell density at the stationary phase is desirable, this parameter may be dictated by the choice of media and reactor design as opposed to controller type. Controller selection, however, has a great impact on the capacity to track the acetate threshold and therefore to enhance productivity without concomitant acetate production.

Normalization of retinal vascular permeability in experimental diabetes with genistein.

PURPOSE: To study the effects of genistein, a tyrosine kinase inhibitor, on retinal vascular permeability in an experimental diabetic rat model. METHODS: Seventy-two rats were equally divided into four groups: (1) nondiabetic control group, (2) diabetic control group, (3) diabetic rats receiving 150 mg genistein/kg food, and (4) diabetic rats receiving 300 mg genistein/kg food. Diabetes was induced by streptozotocin injection in the three diabetic groups. Rats were fed diets with or without genistein and followed for 6 months. Retinal vascular permeability was assessed by measuring radiolabeled sucrose leakage into the retina and by Western blot analysis for total retinal albumin. Retinal phosphotyrosine levels and proliferating cell nuclear antigen (PCNA) were also evaluated by Western blot analysis. RESULTS: Diabetic control rats had markedly increased retinal vascular leakage of radiolabeled sucrose compared with nondiabetic control rats. Diabetic rats receiving oral genistein had significantly less retinal vascular leakage of radiolabeled sucrose than diabetic control rats in a dose-response fashion. Diabetic control rats had increased levels of phosphotyrosine, retinal albumin, and PCNA by Western blot analysis compared with nondiabetic control rats. Rats receiving 300 mg of genistein had decreased retinal albumin by Western blot analysis. Western blot analysis demonstrated a dose-response decrease in retinal phosphotyrosine levels and PCNA in genistein-treated diabetic rats compared with diabetic control rats. CONCLUSIONS: Long-term oral administration of genistein significantly inhibits retinal vascular leakage in experimentally induced diabetic rats. Tyrosine kinase inhibition may be a useful pharmacological approach for the treatment of diabetic-induced retinal vascular leakage.

Flash vacuum pyrolysis of methoxy-substituted lignin model compounds.

The flash vacuum pyrolysis (FVP) of methoxy-substituted beta-O-4 lignin model compounds has been studied at 500 degrees C to provide mechanistic insight into the primary reaction pathways that occur under conditions of fast pyrolysis. FVP of PhCH(2)CH(2)OPh (PPE), a model of the dominant beta-O-4 linkage in lignin, proceeds by C-O and C-C cleavage, in a 37:1 ratio, to produce styrene plus phenol as the dominant products and minor amounts of toluene, bibenzyl, and benzaldehyde. From the deuterium isotope effect in the FVP of PhCD(2)CH(2)OPh, it was shown that C-O cleavage occurs by homolysis and by 1,2-elimination in a ratio of 1.4:1, respectively. Methoxy substituents enhance the homolysis of the beta-O-4 linkage, relative to PPE, in o-CH(3)O-C(6)H(4)OCH(2)CH(2)Ph (o-CH(3)O-PPE) and (o-CH(3)O)(2)-C(6)H(3)OCH(2)CH(2)Ph ((o-CH(3)O)(2)-PPE) by a factor of 7.4 and 21, respectively. The methoxy-substituted phenoxy radicals undergo a complex series of reactions, which are dominated by 1,5-, 1,6-, and 1,4-intramolecular hydrogen abstraction, rearrangement, and beta-scission reactions. In the FVP of o-CH(3)O-PPE, the dominant product, salicylaldehyde, forms from the methoxyphenoxy radical by a 1,5-hydrogen shift to form 2-hydroxyphenoxymethyl radical, 1,2-phenyl shift, and beta-scission of a hydrogen atom. The 2-hydroxyphenoxymethyl radical can also cleave to form formaldehyde and phenol in which the ratio of 1, 2-phenyl shift to beta-scission is ca. 4:1. In the FVP of o-CH(3)O-PPE and (o-CH(3)O)(2)-PPE, products (ca. 20 mol %) are also formed by C-O homolysis of the methoxy group. The resulting phenoxy radicals undergo 1,5- and 1,6-hydrogen shifts in a ratio of ca. 2:1 to the aliphatic or benzylic carbon, respectively, of the phenethyl chain. In the FVP of (o-CH(3)O)(2)-PPE, o-cresol was the dominant product. It was formed by decomposition of 2-hydroxy-3-hydroxymethylbenzaldehyde and 2-hydroxybenzyl alcohol, which are formed from a complex series of reactions from the 2, 6-dimethoxyphenoxy radical. The key step in this reaction sequence was the rapid 1,5-hydrogen shift from 2-hydroxy-3-methoxybenzyloxy radical to 2-hydroxymethyl-6-methoxyphenoxy radical before beta-scission of a hydrogen atom to give the substituted benzaldehyde. The 2-hydroxybenzyl alcohols rapidly decompose under the reaction conditions to o-benzoquinone methide and pick up hydrogen from the reactor walls to form o-cresol.

Hypertensive encephalopathy: complication in children treated for myeloproliferative disorders--report of three cases.

We routinely perform echo-planar diffusion-weighted sequences in all brain magnetic resonance (MR) imaging studies. When three children undergoing chemotherapy for acute leukemia presented with seizures, conventional MR images demonstrated what appeared to be acute, posterior, parasagittal infarcts. However, diffusion-weighted images were normal. These MR imaging findings were consistent with those of hypertensive encephalopathy. Early recognition and treatment of minimal hypertension in these patients allows reversal of encephalopathy.

Comparison of endoscopic retrograde cholangiopancreatography with MR cholangiopancreatography in patients with pancreatitis.

PURPOSE: To assess the usefulness of magnetic resonance (MR) cholangiopancreatography (MRCP) in the evaluation of disease in patients with acute or chronic pancreatitis. MATERIALS AND METHODS: MR imaging was performed at 1.5 T in 39 patients with chronic (n = 30) or acute (n = 9) pancreatitis. The patients underwent a pancreas MR imaging protocol that included an MRCP sequence. Comparison was made with findings at endoscopic retrograde cholangiopancreatography (ERCP), performed within 30 days. Three blinded readers used a scoring system to evaluate nine segments of the pancreatic and biliary ducts as depicted on the ERCP and MRCP images. MRCP image quality was also evaluated. RESULTS: Of 196 segments analyzed, 17 were not seen at MRCP (sensitivity, 91%). Of the segments visualized at MRCP, 14 were incorrectly characterized (accuracy, 92%). At MRCP, segments not detected or mischaracterized were either normal, slightly dilated, or narrowed. At ERCP, 42 segments in 19 patients were not visualized. MRCP findings were considered useful in all those cases. MRCP image quality was not interpretable in two cases due to artifacts. CONCLUSION: Very good correlation between ERCP and MRCP findings was demonstrated. Both modalities failed to depict pathologic conditions depicted by the alternative method. MRCP may obviate ERCP, particularly in patients who cannot undergo ERCP or in whom ERCP has been unsuccessful.

Current concepts in the management of central retinal vein occlusion.

The results of the Central Vein Occlusion Study guide the management of complications of macular edema and neovascularization. Panretinal photocoagulation was shown to be beneficial for eyes with at least 2 hours of iris neovascularization or any angle neovascularization. Grid-pattern laser photocoagulation for perfused macular edema did not show a significant beneficial effect. Recently, several reports have suggested restoring venous outflow by 1) creating a chorioretinal anastomosis, 2) administering recombinant tissue plasminogen activator, 3) cannulating the retinal vein transvitreally, or 4) transecting the posterior scleral ring.

Outcome of pregnancies in women with uterine duplication anomalies.

PURPOSE: This study was conducted to compare pregnancy outcomes in women with uterine duplication anomalies with outcomes in women with normal uteri. METHODS: We studied 22 women with uterine duplication anomalies in whom first-trimester sonography demonstrated a live fetus and compared pregnancy outcomes in these patients with outcomes in 66 women with normal-appearing uteri (controls). The 2 groups were matched for gestational age at the time of first-trimester sonography. RESULTS: Uterine duplication anomalies were associated with a higher miscarriage rate than were normal-appearing uteri (23% versus 6%, p < 0.05, Fisher's exact test). Deliveries resulting in liveborn infants occurred earlier in women with anomalous uteri. The mean gestational age at delivery was 37.0 weeks for women with duplication anomalies versus 38.8 weeks for controls (p < 0.05, Student's t-test), and a higher percentage of duplication anomaly patients delivered before 34 weeks' gestation (29% versus 3%, p < 0.01, Fisher's exact test). Neonates born from anomalous uteri had lower mean birth-weight percentiles than did neonates in the control group (51st versus 68th percentile, p < 0.05, t-test). There was no difference in Apgar scores between the 2 groups at 1 and 5 minutes. CONCLUSIONS: Women with duplication anomalies of the uterus have a higher miscarriage rate, deliver earlier, and have infants with lower birth-weight percentiles than do women with normal-appearing uteri.

Eye injuries caused by bungee cords.

OBJECTIVE: Bungee cords are common workplace and household items that have many uses. Despite their utility, they represent a potential source of severe ocular injury. DESIGN: A retrospective review was conducted to identify patients presenting to an ocular emergency department with bungee cord-related eye injuries over a 42-month period. PARTICIPANTS: The authors identified 17 patients with bungee cord-related ocular injuries. INTERVENTION: In identified patients, the hospital inpatient and outpatient charts were reviewed in detail. MAIN OUTCOME MEASURES: Patient demographics, mechanism of injury, use of ocular protection, presenting and final best-corrected visual acuity, associated ocular injuries, therapeutic interventions, and anatomic results were noted. RESULTS: Fourteen (82%) of the patients had closed globe injuries and 3 (18%) had open globe injuries. A wide array of periocular, anterior segment, and posterior segment injuries were identified with hyphema being the most common associated ocular injury. Ten patients (59%) had a final visual acuity of 20/25 or better. Five patients (29%) had a final visual acuity that was less than or equal to 20/60, with three of these patients having a final visual acuity less than or equal to 4/200. Two of the three patients with open globe injuries had a final visual acuity of 20/60 or better, whereas one had no light perception. Poor visual outcome was associated with posterior segment involvement. CONCLUSIONS: The authors advocate the use of printed warnings on the packaging of bungee cords and extreme caution by those who use them. A modification in design and the use of certified safety glasses may help to decrease the incidence of bungee cord-related ocular trauma.

Early singleton pregnancy outcome: effects of maternal age and mode of conception.

PURPOSE: To assess the effect of prognostic factors on the outcome of singleton pregnancies. MATERIALS AND METHODS: First-trimester ultrasonographic (US) scans that demonstrated a living fetus in 4,156 consecutive singleton pregnancies were studied. The relationship between outcome and maternal age, mode of conception, maternal symptoms, and US findings was evaluated. RESULTS: Spontaneous abortion occurred in 371 of 4,156 (8.9%) cases. Higher pregnancy-loss rates were associated with older maternal age (P < 10(-5)), assisted mode of conception (P < 10(-8)), maternal symptoms of pain and/or bleeding (P < .001), and abnormal US findings (P < 10(-8)). US abnormalities were more frequent in older women than in younger women (P < 10(-5)) and in assisted conceptions than in natural conceptions (P < 10(-8)). At stepwise logistic regression, with gestational age as a covariate, US abnormalities and maternal symptoms independently affected pregnancy outcome. Maternal age and mode of conception had no further statistically significant effect on pregnancy outcome. CONCLUSION: The prognosis for older mothers and for those with assisted conception is not statistically significantly different from that for younger mothers and for those with natural conception if maternal symptoms, US findings, and gestational age are the same.

Physiologic studies with the sulfate-reducing bacterium Desulfovibrio desulfuricans: evaluation for use in a biofuel cell.

The growth kinetics of the sulfate-reducing bacteria Desulfovibrio desulfuricans Essex 6 was investigated under various conditions for potential use in a microbial fuel cell that recovers electrons generated from the reduction of sulfate to hydrogen sulfide. Hydrogen sulfide was found to inhibit growth and decrease both the growth yields and the sulfate-specific reduction rate. Hydrogen sulfide inhibition was direct, reversible, and not due to limitation by iron deficiency. A high initial lactate concentration also retarded bacterial growth, reduced the specific sulfate reduction rates, and gave variable biomass growth yields. This effect resulted from a bottleneck in the lactate oxidation pathway which induced the production of the secondary product butanol. The use of pyruvate as a carbon source was more advantageous than lactate in terms of growth rate and biomass growth yields, with only a slight decrease in the rate of specific sulfate reduction. For equal biomass, a slightly higher current density was generated from lactate than pyruvate, but pyruvate required nearly 40% less sulfate.

Optimal dynamic experiments for bioreactor model discrimination.

This paper describes a general approach for dynamic model discrimination for continuous cultures and presents dynamic models for pure cultures of E. coli and C. utilis obtained using the method. For each pure culture system, four candidate models representing various levels of structure were considered. All models reduce to Monod growth kinetics at steady state. An optimized set of multivariable step inputs in selected manipulative variables was used to discriminate between candidate models. The models that best predicted the dynamic behavior were selected by comparison of model predictions with experimental data. Two discrimination functions were compared in terms of their ability to determine the optimal set of multivariable step inputs to discriminate between candidate models. Results indicate that model discrimination based on maximizing the minimum absolute difference between any two models for a given set of inputs possessed good potential for discrimination between candidate models. Models selected for E. coli and C. utilis from the model discrimination work are presented and compared with experimental data.

Orbital polymyositis and giant cell myocarditis.

BACKGROUND: Orbital polymyositis associated with giant cell myocarditis rarely has been reported in the literature. The authors report the clinical, neuroradiographic, and histopathologic features of the only patient to survive this usually fatal syndrome after cardiac transplantation. FINDINGS: This 22-year-old white woman presented in 1991 with periorbital redness, swelling, and pain in both eyes that was unresponsive to antibiotic therapy. Results of her examination were significant for limited extraocular movements, ptosis, erythema, edema, chemosis, and exophthalmos. Electrocardiogram and chest x-ray were normal. Orbital computed tomographic scan showed swelling of the extraocular muscles up to and including their insertions. The patient was given the diagnosis of orbital polymyositis and her condition improved clinically and radiographically while taking parenteral steroids. One month after discharge, the patient was in cardiogenic shock. Endomyocardial biopsy showed giant cell myocarditis, and the patient underwent emergent cardiac transplantation. Despite a complicated postoperative course, the patient has done remarkably well. CONCLUSION: Although this disorder is rare, this case suggests the need for a high index of suspicion for giant cell myocarditis in patients with inflammatory orbital polymyositis. In non-Graves orbital polymyositis the patient should be questioned and instructed concerning the signs and symptoms of congestive heart failure. Chest x-ray, Holter monitoring, and electrocardiogram also should be performed and be repeated with an echocardiogram if there are any cardiac symptoms. In addition, early endomyocardial biopsy should be considered in the proper clinical setting, allowing timely diagnosis and expeditious cardiac transplantation.