Determination of D-sorbitol in human erythrocytes by an improved enzymatic method with fluorometric detection

In this enzymatic method to analyze erythrocytes for D-sorbitol, the erythrocytes were separated from plasma by centrifugation, washed with isotonic saline (9 g/L NaCl), then diluted threefold with more saline. We lysed 1.5 mL of the diluted erythrocytes with chloroform and precipitated the protein with 58 g/L HClO4 solution. The resulting supernates were mixed with buffer, NAD+, and sorbitol dehydrogenase (EC 1.1.1.14). For standards, we added sorbitol to the erythrocytes before lysing. After 25 min of incubation at 37 degrees C, the fluorescence responses were recorded. Results for sorbitol were corrected for sample-blank and enzyme fluorescence, and were then normalized for hemoglobin content. Response was linear for a sorbitol concentration range of 1 to 30 mumol/L. Reproducibility was good, with an average CV of 2.5% at 10 mumol/L. Results for healthy individuals and diabetic patients are presented.     

Gene typing of Chlamydia trachomatis by polymerase chain reaction and restriction endonuclease digestion.

A portion of the major outer membrane protein (MOMP) gene from 15 Chlamydia trachomatis serovars was amplified by polymerase chain reaction (PCR) and the product was analyzed by restriction fragment length polymorphism (RFLP). A set of primers was used to amplify an 871 base pair gene fragment encompassing the 4 hypervariable regions of MOMP. AluI digestion of the product gave distinctive patterns for the 15 serovars as demonstrated on silver-stained polyacrylamide gels. A triple digest with EcoRI, HinfI, and HpaII allowed improved discrimination between closely related serovars (C, H, I, J, L3). PCR and RFLP were used to type 50 wild-type clinical isolates and results were compared to results of the solid-phase enzyme immunoassay typing method. These isolates represented the most prevalent genital serovars (D, E, F, K, I and J) in the local sexually transmitted diseases clinic population. For specimens containing 1 serovar, the results of the two methods were similar for 42 samples and discordant for 1 sample. In addition, two samples showed evidence of mixed infection with two serovars as identified by both methods. Five additional specimens contained two serovars, as shown by one or both methods. In all five such specimens, the two typing methods agreed on at least one of the two serovars. For both single and multiple serovar specimens, there was concordance between the two typing methods for 16/17 E serovars, 8/9 D serovars, 8/8 F serovars, 7/7 I serovars, 7/7 J serovars, 5/8 K serovars, and 0/2 G serovars.(ABSTRACT TRUNCATED AT 250 WORDS)     

Epikeratophakia without annular keratectomy

Nineteen patients (20 eyes) underwent epikeratophakia for correction of high myopia. In contrast to the conventional surgical method utilizing an annular keratectomy, no keratectomy was performed in these patients. The surgical procedure is discussed. Preliminary results (up to six months) indicate that the omission of an annular keratectomy has no effect on graft clarity or viability. The weighted-average percent correction over six months postoperatively was 102%.     

Patterns of cognitive change over time and relationship to age following successful treatment of Cushing's disease.

Chronically elevated levels of cortisol have been associated with changes in cognitive functioning and brain morphology. Using Cushing's disease as a model to assess the effects of high levels of cortisol on cognitive functioning, 27 patients with Cushing's disease were examined at baseline and three successive follow-up periods up to 18 months after successful surgical treatment. At all follow-up periods, patients were administered cognitive tests as well as measures of plasma and urinary free cortisol. Structural MRIs and a depression measure were taken at baseline and one-year follow-up. Results showed that there is a specific pattern of significant cognitive and morphological improvement following successful treatment. Verbal fluency and recall showed recovery, although brief attention did not. Age of participants was a significant factor as to when recovery of function occurred; younger patients regained and sustained their improvement in cognitive functioning more quickly than older participants. Improvement in verbal recall also was associated with a decrease in cortisol levels as well as an increase in hippocampal formation volume one year after treatment. Overall, these findings suggest that at least some of the deleterious effects of prolonged hypercortisolemia on cognitive functioning are potentially reversible, up to at least 18 months post treatment.     

Isolated noncompaction of the ventricular myocardium

Isolated noncompaction of the ventricular myocardium is a recently described anomaly. We report the first case of this anomaly presenting as a restrictive cardiomyopathy, and the first association of this entity with endocardial fibrosis.