Enhancement of lipoprotein lipase activity by tissue factor pathway inhibitor

The effect of a basic synthetic peptide, representing the C-terminal region of tissue factor pathway inhibitor (TFPI - Lys254 - Met276), as well as that of the whole protein, on the activity of lipoprotein lipase (LPL) is described. The activity of bovine LPL was measured by chromogenic assay using a water-soluble chromogenic substrate, p-nitrophenyl butyrate. Five and 10 microM concentrations of the peptide increased Vmax of bovine LPL by 48.9% and 85.6% respectively as compared with the buffer control without affecting Km. Poly l-lysine, though positively charged did not have any effect, suggesting the importance of the amino acid sequence of the test peptide. On the other hand, 0.25, 0.5 and 1.0 mM n-butyric acid - a product of LPL catalysis in the chromogenic assay, when added to the incubation mixture decreased Vmax non competitively by 22.8%, 40.4% and 63% respectively as compared with buffer control, confirming the known product inhibition of LPL. A 100-fold molar excess of n-butyric acid produced inhibition of the LPL reaction as compared with the synthetic peptide which produced potentiation, suggesting a 1:100 stoichiometric interaction of the peptide with n-butyric acid. At a fixed concentration of 0.25 mM substrate, 10 nM full length recombinant TFPI, containing the basic C-terminal domain, increased velocity of LPL reaction by 39.4% as compared with buffer control. The same concentration of two-domain recombinant TFPI (TFPI1-160) had no effect. It is possible that negatively charged n-butyric acid is sequestered by the positively charged peptide or the basic region of recombinant full length TFPI. Relieving of product inhibition could then be a possible mechanism of the observed potentiation of bovine LPL activity by the basic peptide or full length recombinant TFPI. The 39.4% increase in reaction velocity of LPL catalysis produced by 10 nM full length recombinant TFPI was comparable to 38.9% increase produced by 5 microM of the basic peptide under the same conditions. A further increase of 78.7% was brought about by 10 microM concentration of the same peptide. The reason for about 500-fold increase in the potency of the whole protein as compared with that of the peptide is not clear. It is possible that in its tertiary conformational state, the whole protein is able to sequester product and relieve product inhibition more effectively than the short linear peptide. Rabbit polyclonal antiserum against the basic peptide partially inhibited LPL activity of human post heparin plasma, measured by radioenzymatic assay using triolein substrate. Since post heparin plasma contains full length TFPI, binding of the added antibody to its basic C-terminus and hence the relative unavailability of latter for product sequestration (oleic acid in this case) could explain the observed inhibition of human LPL activity by antibody against the peptide. Thus by enhancing lipase activity, full length TFPI may facilitate hydrolysis of triglyceride and concomitantly lower factor VII coagulant activity as demonstrated earlier, particularly after heparin injection when both TFPI and LPL are released in circulation.     

Monoamine oxidase A inhibition by fluoxetine: an in vitro and in vivo study

Monoamine oxidase A (MAO-A) inhibition was investigated both in vitro and in vivo in rat brains by using the radioligand, 18F-fluoroclorgyline (N-[3-(2',4'-dichlorophenoxy)-2-18F-fluoropropyl]-N-methylpropa rgylamine). In vitro binding affinities of six compounds, clorgyline, Ro 41-1049, deprenyl, fluoxetine, norfluoxetine and citalopram, were studied. Fluoxetine and norfluoxetine showed in vitro affinities of 36.5 and 68 microM for MAO-A, respectively. Fluoxetine and norfluoxetine also significantly inhibited (more than 20%) the binding of the radioligand in vivo while citalopram and deprenyl showed very poor affinities in vitro for MAO-A and had no effect in vivo. The in vivo effects of the various drugs were directly comparable to their in vitro affinities for binding to MAO-A as seen in the correlation plot of percent control in vivo binding of 18F-fluoroclorgyline and binding affinity, -log IC50 (R2 = 0.979). An acute dose of 20 mg/kg of fluoxetine inhibited binding of 18F-fluoroclorgyline by more than 20%, while lower doses had some significant effects. These results provide evidence on the in vitro and in vivo inhibition of monoamine oxidase A by fluoxetine.     

Tumor proliferative activity and response to first-line chemotherapy in advanced breast carcinoma

Summary The relationship between tumor proliferative activity and response to first-line chemotherapy and survival was investigated in 76 advanced breast cancer patients. Proliferative activity was determined by means of Ki-67 immunohistologic staining on primary tumors (55 patients) or at the relapse site (21 patients), and was classified as low ( 25% of stained cells) or high (> 25% of stained cells). The usual WHO response criteria were used. The median duration of follow-up was 18 months (range 3–58).Forty-seven patients (62%) had tumors with low, and 29 (38%) had tumors with a high rate of proliferative activity. The two groups were well balanced in terms of important variables such as disease-free survival, performance status, age, menopausal status, and the type of first-line chemotherapy (anthracycline-based regimens versus cyclophosphamide-methotrexate-5-fluorouracil). The estrogen receptor (ER) content, measured by means of immunohistochemical assay, was markedly different in the two groups, with 27/47 tumors with low proliferative activity (57%) and 6/29 with high-proliferative activity (21%) being ER positive ( 45% of stained cells) (p = 0.003). Moreover, a significant difference in the metastatic pattern was also evident, with a higher incidence of bone and a lower incidence of soft tissue metastases in the group of patients with tumors with low proliferative activity (p = 0.004). Overall, 10/47 responses (21%: PR = 7, and CR = 3) were observed in the group with a low rate of proliferative activity, versus 14/29 (48%: PR = 9, and CR = 5) in the group with highly proliferative tumors, the difference being statistically significant (p = 0.03). When a multivariate analy-sis was performed, the only factor that retained independent prognostic significance was the predominant site of disease, particularly soft tissues (p = 0.003). Despite the difference in response rate, when survival analysis was performed according to the Kaplan-Meier method, no significant difference was observed in the two groups, but when the analysis was limited to responsive patients, the median survival observed in those with a low and those with a high rate of proliferation was 35 and 19 months respectively (p = 0.02). The same results were obtained when multivariate survival analysis was carried out using Cox's regression model. These data suggest that there is a link between tumor proliferative activity and response to chemotherapy in advanced breast cancer, and may indicate the need to use more intensive treatments in selected patients with highly proliferative tumors.Presented in part at the Annual Meeting of the American Society of Clinical Oncology, May 14–17, 1994, Dallas, TX, USA     

Complete inversion of the urinary bladder through a vesicovaginal fistula

This is the case of an Indian woman who was hospitalized after a diagnosis of chronic inversion of the uterus with a vesicovaginal fistula. She suffered from urinary incontinence for 40 years, the condition having developed following a difficult labor. Ultimately, she proved to have a complete inversion of the bladder through a vesicovaginal fistula.     

Epirubicin, methotrexate and bleomycin (EMB) in the treatment of recurrent epidermoid cancer of the head and neck.

During the period May 1989 to November 1990, at the "O. Alberti" Radium Institute of Brescia's General Hospital, 35 patients affected by epidermoid head and neck carcinoma were treated every 28 days with the salvage chemotherapy regimen EMB (epirubicin, 50 mg/m2 i.v. day 1; methotrexate, 40 mg/m2. i.v. days 1, 18; bleomycin, 10 mg/m2 i.v. days 4, 11, 18). Sixteen patients had been previously treated with surgery, 15 with radiotherapy and 4 with chemotherapy. Six patients (Group A) received only 1 cycle of chemotherapy because of disease progression and subsequent death. In another 15 patients (Group B) it was possible to administer 2 cycles of EMB, and 9 of them showed local disease progression and died. Among the remaining 6 patients, evaluated as PR, 1 refused further therapy and 5 were amenable to a previously impossible radiotherapy (4 of them are still alive). Fourteen patients received 3 or more cycles of EMB (Group C): 8 subjects showed progression and died; 1 reached CR and is alive without any evidence of tumor; 5 are in PR (3 of them underwent subsequent radiotherapy and 1 chemotherapy with CDDP). Out of 35 patients, 12 (34%) reached a favorable response (CR or PR) and 8 (22%) are still alive. As regards toxicity, the following adverse events were recorded (< or = 2 Miller's scale): leukopenia (8.5%), thrombocytopenia (5.7%), anemia (14.2%), stomatitis (5.7%), vomiting (5.7%), alopecia (8.5%), and fever (11.4%). It can be concluded that the EMB regimen is very well tolerated and shows good effects in the treatment of patients with relapsed head and neck carcinoma.     

Incidence and outcome of chromosomal mosaicism found at the time of chorionic villus sampling

OBJECTIVE: Chromosomal mosaicism has been reported in about 1% to 3% of chorionic villus sampling specimens. This report provides incidence and outcome information that should be useful in counseling patients found to have mosaicism on chorionic villus sampling.STUDY DESIGN: A retrospective analysis of 11,200 consecutive patients undergoing chorionic villus sampling at the University of California, San Francisco, during the period from Jan. 1, 1984, to June 1, 1996, was undertaken.RESULTS: A total of 140 cases of mosaicism were identified for an incidence of 1.3%. Follow-up information was available for 130 cases, 26 of which (20%) were confirmed in fetal tissue. Confirmation rates for specific types of mosaicism were as follows: autosomal trisomy 7.6%, sex chromosome 25%, structural abnormality 27.3%, and marker chromosome 77.8%. Neonatal outcome was normal in all cases for which pregnancy continued.CONCLUSION: The data indicate that in most cases of chromosomal mosaicism found by chorionic villus sampling the mosaicism is unlikely to be clinically significant in the fetus. (Am J Obstet Gynecol 1997;176:1349-53.)     

Cerebral autoregulation in pediatric traumatic brain injury.

OBJECTIVE: The aims of this study were to document the incidence of impaired cerebral autoregulation in children with traumatic brain injury using transcranial Doppler ultrasonography and to examine the relationship between autoregulatory capacity and outcome in children following traumatic brain injury. DESIGN: Prospective cohort study. SETTING: Harborview Medical Center (level I pediatric trauma center) in Washington state. PATIENTS: Thirty-six children <15 yrs old with traumatic brain injury: Glasgow Coma Scale score <9 (n = 12, group 1), Glasgow Coma Scale score 9-12 (n = 12, group 2), and Glasgow Coma Scale score 13-15 (n = 12, group 3). INTERVENTIONS: Cerebral autoregulation testing was conducted during extracranial surgery. Mean middle cerebral artery flow velocities were measured using transcranial Doppler as mean arterial pressure was increased to whichever variable was greater: 20% above baseline or a set value (80 mm Hg for <9 yrs and 90 mm Hg for 9-14 yrs). Autoregulatory capacity was quantified by the Autoregulatory Index. Autoregulatory Index <0.4 was considered impaired cerebral autoregulation. Discharge outcome using the Glasgow Outcome Scale score was considered good if the Glasgow Outcome Scale score was > or =4. MEASUREMENTS AND MAIN RESULTS: Twenty-four (67%) of 36 children had an Autoregulatory Index > or =0.4. The incidence of impaired cerebral autoregulation was 42% (five of 12) in group 1, 42% (five of 12) in group 2, and 17% (two of 12) in group 3. Ten (42%) of the 24 children with intact cerebral autoregulation had a good outcome compared with only one of 12 (8%) children with impaired cerebral autoregulation (p =.04). Six of 12 (50%) children with impaired cerebral autoregulation had hyperemia compared with one of 24 (4%) children with intact cerebral autoregulation (p <.01). Hyperemia was associated with poor outcome (p =.01). CONCLUSIONS: The incidence of impaired cerebral autoregulation was greatest following moderate to severe traumatic brain injury. Impaired cerebral autoregulation was associated with poor outcome. Hyperemia was associated with impaired cerebral autoregulation and poor outcome.     

Trends in compliance with the national colposcopy guidelines.

This study aims to assess trends in compliance with current colposcopy guidelines in 10 gynaecological units in four English counties since 1996; to identify constraints on compliance and suggest change in practice. All 10 gynaecology units in Oxfordshire, Buckinghamshire, Northamptonshire and Berkshire participated. Data were collected prospectively by colposcopists from 23,500 new referrals across a 55-month period from September 1996 to March 2001. The Oxford Cancer Intelligence Unit performed collation, quality assurance and retrieval of data for incomplete records. Audit results were disseminated annually to colposcopists via the Regional Colposcopy Group. Colposcopy waiting times exceeded the standards, but waiting times for high-grade referrals showed statistically significant improvement. Six standards were achieved; relating to accuracy, appropriateness of management and outcomes. The seven unmet standards relate to waiting times, colposcopist's caseload, follow-up policy and the proportion of cervical epithelial neoplasia (CIN) on histology. Changes in practice are suggested, constraints on compliance are identified and the appropriateness of some guidelines is questioned.