The partitioning of phosphoramide mustard and its aziridinium ions among alkylation and P-N bond hydrolysis reactions

NMR (1H and 31P) and HPLC techniques were used to study the partitioning of phosphoramide mustard (PM) and its aziridinium ions among alkylation and P-N bond hydrolysis reactions as a function of the concentration and strength of added nucleophiles at 37 degrees C and pH 7.4. With water as the nucleophile, bisalkylation accounted for only 10-13% of the product distribution given by PM. The remainder of the products resulted from P-N bond hydrolysis reactions. With 50 mM thiosulfate or 55-110 mM glutathione (GSH), bisalkylation by a strong nucleophile increased to 55-76%. The rest of the PM was lost to either HOH alkylation or P-N bond hydrolysis reactions. Strong experimental and theoretical evidence was obtained to support the hypothesis that the P-N bond scission observed at neutral pH does not occur in the parent PM to produce nornitrogen mustard; rather it is an aziridinium ion derived from PM which undergoes P-N bond hydrolysis to give chloroethylaziridine. In every buffer studied (bis-Tris, lutidine, triethanolamine, and Tris), the decomposition of PM (with and without GSH) gave rise to 31P NMR signals which could not be attributed to products of HOH or GSH alkylation or P-N bond hydrolysis. The intensities of these unidentified signals were dependent on the concentration of buffer.     

Tendons attached to prostheses by tendon-bone block fixation: an experimental study in dogs

To develop a method of tendon attachment to a metallic endoprosthesis, we evaluated fixation strength, clinical function of the tendon, and morphological changes in an experimental model. The canine supraspinatus tendon was removed from the greater tubercle of the humerus and attached to a titanium prosthesis. In 12 animals, the bone block underlying the tendon insertion was preserved and attached in one limb; the soft part of the tendon was attached directly to the prosthesis in the contralateral limb. Fixation strength was evaluated after 16 weeks of in vivo implantation (12 specimens) and compared with the in vitro fixation strength (12 specimens) and with intact normal controls (six specimens from cadavera). Function of the tendon in vivo was evaluated by force-plate analysis (at 3-week intervals). All specimens were evaluated histologically. Sixteen weeks after surgery, the tendon-bone block attachment was significantly stronger (mean, 16%) than the direct tendon attachment and not significantly different from the normal control, and the direct tendon attachment was significantly weaker (mean, 68%) than the normal control. There was significantly more weight-bearing on the limbs with a tendon-bone block attachment than on the limbs with a direct tendon attachment at both 3 and 6 weeks postoperatively. Both front legs showed increased weight-bearing with time, but the differences were not statistically significant. Anchorage by tissue ingrowth to the titanium prosthesis was found consistently--there was bone ingrowth in the tendon-bone block attachments and fibrous tissue ingrowth in the direct tendon attachments. When a bone block was preserved, the strength and stiffness were comparable with those of a normal tendon insertion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Development of a patient-reported measure of function of the knee

The purpose of the present study was to demonstrate the reliability, validity, and responsiveness of the Activities of Daily Living Scale of the Knee Outcome Survey, a patient-reported measure of functional limitations imposed by pathological disorders and impairments of the knee during activities of daily living. The study comprised 397 patients; 213 were male, 156 were female, and the gender was not recorded for the remaining twenty-eight. The mean age of the patients was 33.3 years (range, twelve to seventy-six years). The patients were referred to physical therapy because of a wide variety of disorders of the knee, including ligamentous and meniscal injuries, patellofemoral pain, and osteoarthrosis. The Activities of Daily Living Scale was administered four times during an eight-week period: at the time of the initial evaluation and after one, four, and eight weeks of therapy. Concurrent measures of function included the Lysholm Knee Scale and several global measures of function. The subjects also provided an assessment of the change in function, with responses ranging from greatly worse to greatly better, at one, four, and eight weeks. The Activities of Daily Living Scale was administered to an additional sample of fifty-two patients (thirty-two male and twenty female patients with a mean age of 31.6 years [range, fourteen to sixty-six years]) before and after treatment within a single day to establish test-retest reliability. Factor analysis revealed two dominant factors: one that reflected a combination of symptoms and functional limitations and the other, only symptoms. The internal consistency of the Activities of Daily Living Scale was substantially higher than that of the Lysholm Knee Scale (coefficient alpha, 0.92 to 0.93 compared with 0.60 to 0.73), resulting in a smaller standard error of measurement for the former scale. Validity was demonstrated by moderately strong correlations with concurrent measures of function, including the Lysholm Knee Scale (r = 0.78 to 0.86) and the global assessment of function as measured on a scale ranging from 0 to 100 points (r = 0.66 to 0.75). Analysis of variance with repeated measures revealed significant improvements in the score on the Activities of Daily Living Scale during the eight weeks of physical therapy (F2,236 = 108.13; p < 0.0001); post hoc testing indicated that the change in the score at eight weeks was significantly greater than the change at four weeks and that the change at four weeks was significantly greater than that at one week (p < 0.0001 for both). As had been hypothesized, the patients in whom the knee had somewhat improved had a significantly smaller change in the score, both at four weeks (F1,189 = 33.50; p < 0.001) and at eight weeks (F1,156 = 22.48; p < 0.001), compared with those in whom the knee had greatly improved. The test-retest reliability coefficient (intraclass correlation coefficient[2,1]) was 0.97. These results suggest that the Activities of Daily Living Scale is a reliable, valid, and responsive instrument for the assessment of functional limitations that result from a wide variety of pathological disorders and impairments of the knee.     

Triple therapy with ursodeoxycholic acid, prednisone and azathioprine in primary biliary cirrhosis: a 1-year randomized, placebo-controlled study

BACKGROUND/AIMS: Treatment with ursodeoxycholic acid has been shown to decrease the rate of disease progression in patients with primary biliary cirrhosis, although the effect is modest. Since primary biliary cirrhosis has many features of an autoimmune disorder, immunosuppressives added to ursodeoxycholic acid may be of value in the treatment of primary biliary cirrhosis. METHODS: A 1-year randomized, double-blind, placebo-controlled trial was carried out in 50 patients with primary biliary cirrhosis, who had already been treated with ursodeoxycholic acid for at least 1 year, but had not achieved complete disease remission. Patients were randomized to additional prednisone (30 mg per day initially, tapered to 10 mg daily after 8 weeks) and azathioprine (50 mg daily) or placebo. A subgroup of patients received cyclical etidronate and calcium. The principal aim of the study was to assess the short-term benefits and risks of the combined bile acid and low-dose immunosuppressive regimen. Primary endpoints were effects on symptoms, liver biochemistry, liver histology, bone mass and the occurrence of adverse events. RESULTS: Pruritus (p=0.02), alkaline phosphatase, aspartate aminotransferase, IgM and procollagen-III-propeptide improved significantly (all p<0.002) in the combined treatment group as compared to the placebo group. Histological scores for disease activity and disease stage decreased significantly within the combination treatment group (p<0.001). CONCLUSIONS: In patients with primary biliary cirrhosis receiving ursodeoxycholic acid, there is an additional beneficial effect of 1-year treatment with prednisone and azathioprine on symptoms and biochemical, fibrogenetic and histological parameters. These results strongly encourage the evaluation of this triple treatment regimen in long-term controlled trials of adequate size to document its effect on clinical events.     

In vitro and in vivo effect of glucocorticoids on IgE and IgG subclass secretion

Hydrocortisone (HC) as well as its synthetic derivatives have been shown to strongly enhance interleukin-4 (IL-4)-induced in vitro IgE synthesis. To investigate possible effects on IgG subclasses, peripheral blood mononuclear cells (PBMC) were incubated with different glucocorticosteroids in the absence or presence of IL-4. The glucocorticoids alone led to a strongly enhanced secretion of IgG1, IgG2 and IgG3, but not IgG4. The addition of IL-4 induced marked increases in IgG1 and IgG4, no changes in IgG3, but a consistent decrease in IgG2 synthesis. In order to find out whether these profound in vitro effects of corticosteroids are also reflected by changes in antibody serum levels during steroid treatment, 10 healthy volunteers took 25 mg prednisone for 7 consecutive days. We could not observe any significant changes of IgE or IgG subclass serum levels during or after this period. However, cell cultures performed after the glucocorticoid treatment revealed a marked decrease in the ability to produce IgG4 and a significantly lower potential to produce IgE in response to IL-4 alone or IL-4 and HC. We conclude that, although strongly implicated by the in vitro results, glucocorticosteroid treatment does not result in an increased allergy risk.     

Circadian variations of the urinary excretion of catecholamines and electrolytes

Concomitant measurements of circadian variations in the urinary excretion of dopamine (DA), homovanillic acid (HVA), norepinephrine (NE), epinephrine (E) as well as of creatinine, sodium and potassium under controlled dietary conditions during relative physical and emotional rest in 13 volunteers have shown that maximum excretion of all these substances occurred in the afternoon period between 14:30h and 18:00h, and minimum excretion in the morning between 4:00h and 5:00h. The changes were in some cases progressive from one collection period to the other, and synchronized for NE and E. DA and HVA excretions fluctuated from subject to subject. Excretory rhythms of sodium and potassium were found to be similar to those of the catecholamines. This can be explained by diurnal changes in renal blood flow and different renal excretory mechanisms of catecholamines. None of the catecholamines correlated with the urinary volume but urinary NE and E positively correlated with urinary creatinine, urinary NE and E with urinary DA and urinary sodium with urinary E. There are some common patterns in the diurnal rhythms of catecholamines and electrolytes but their interrelationship is different for individual catecholamines.     

Mental symptoms in Parkinson''s disease during chronic treatment with levodopa

The influence of epidural block with bupivacaine (Marcaine-adrenalin) on fetal heart rate, uterine activity, and the frequency and intensity of contractions was studied in twenty-five nulliparae at term. Uterine activity was found to decrease during the first 30 minutes following epidural block. In the time interval 30 to 40 minutes after epidural block uterine activity increased again and attained the same level as during the last 10 minutes before analgesia. The frequency of uterine contractions did not decrease after the block. The lower levels of uterine contractility were due only to a lower amplitude of the contractions. The regularizing effect of epidural analgesia on incoordinate uterine action was noted. The recording of fetal heart rate in the time interval studied revealed no pathological findings. Mode of delivery, mean labour duration and Apgar scores after epidural block were comparable with earlier studies of a larger patient population. This study suggest that epidural analgesia does not induce any important changes in fetal heart rate but temporarily decreases uterine activity.     

Proliferation, differentiation, and long-term culture of primary hippocampal neurons

Primary embryonic hippocampal neurons can develop morphologically and functionally in culture but do not survive more than a few weeks. It has been reported that basic fibroblast growth factor (bFGF) promotes the survival of and neurite elongation from fetal hippocampal neurons. We report that bFGF, in a dose-dependent manner, can induce the survival (50 pg to 1 ng/ml) and proliferation (10-20 ng/ml) of embryonic hippocampal progenitor neurons in vitro. In serum-free medium containing high concentrations of bFGF, neurons not only proliferated (4-day doubling time) and differentiated morphologically but also could be passaged and grown as continuous cell lines. The neuronal nature of the proliferating cells was positively established by immunostaining with several different neuron-specific markers and by detailed ultrastructural analyses. The proliferative effect of bFGF was used to generate nearly pure neuronal cell cultures that can be passaged, frozen, thawed, and cultured again. Neurons have been maintained > 5 months in culture. The ability to establish long-term primary neuronal cultures offers the possibility that clonal lines of distinct neuronal cell types may be isolated from specific areas of the central nervous system. Such long-term neuronal cultures should prove valuable in studying neurons at the individual cell level and also in exploring interactions between neurons in vitro. The observed dose dependence raises the possibility that cell survival and proliferation in vivo may be influenced by different levels of bFGF.