Fluorine-19 nuclear magnetic resonance studies of binary and ternary complexes of thymidylate synthase utilizing a fluorine-labeled folate analogue

Traditional fluorine-19 nuclear magnetic resonance (19F NMR) studies of thymidylate synthase (TS) have utilized the fluorine substituent of 5-fluorodeoxyuridine 5'-monophosphate (FdUMP), a mechanism-based inhibitor of the enzyme, in complexes with various folate and folate analogues in order to establish a paradigm for the formation of binary and ternary complexes. In order to extent and confirm this paradigm, complexes of thymidylate synthase (TS) and the N-10-(fluoroethyl)quinazolinylfolate analogue CB3731 with either deoxyuridine 5'-monophosphate (dUMP), deoxythymidine 5'-monophosphate (dTMP), or FdUMP were examined from the perspective of the folate analogue using 19F NMR. The spectrum of the free folate analogue gave rise to a multiplet centered at -57.0 ppm, which was broadened by approximately 50% upon incubation with the enzyme. The use of FdUMP with CB3731 afforded us the opportunity to compare the spectrum obtained for the folate with that of the nucleotide. This comparison led to the assignment of the resonance at -53.5 ppm as representing the noncovalent TS:FdUMP:CB3731 ternary complex, while a new resonance at -52.0 ppm corresponded to the species in which the nucleotide is covalently attached to the enzyme and the folate is noncovalently associated. Ternary complexes consisting of TS, CB3731, and either dUMP or dTMP displayed a resonance at -53.5 ppm which represented the noncovalent TS-nucleotide adduct. None of the TS:nucleotide:CB3731 ternary complexes, however, was stable to native polyacrylamide gel electrophoresis.     

Do substance abuse patients with more psychopathology receive more treatment?

The association between psychopathology at treatment entry and the amount of treatment services received was evaluated in 104 alcohol-dependent and 100 cocaine-dependent male veterans treated for 1 month in either a day hospital or inpatient program. Measures of psychopathology included the Addiction Severity Index psychiatric composite score, the presence or absence of an antisocial personality disorder diagnosis, and the total number of additional lifetime or current psychiatric diagnoses. Patients with higher admission Addiction Severity Index psychiatric composite scores received more medical, alcohol, family/social, and psychiatric services. There was also preliminary evidence that patients who received more treatment showed greater improvement 7 months after admission. The relationships between the other measures of psychopathology and treatment services failed to achieve overall statistical significance, although significant relationships were found in several individual areas.     

Thyroid cancer

Management of thyroid cancer varies somewhat between communities and institutions depending on tumor type and individual treatment philosophy. The differentiated thyroid cancers have a significantly better outlook than the medullary and anaplastic. This article provides an overview of the literature that describes pathogenesis, diagnosis, and treatment currently recommended for these thyroid cancers.     

In vivo modulation of vaccine-induced immune responses toward a Th1 phenotype increases potency and vaccine effectiveness in a herpes simplex virus type 2 mouse model

Several vaccines have been investigated experimentally in the herpes simplex virus type 2 (HSV-2) model system. While it is believed that CD4(+)-T-cell responses are important for protection in general, the correlates of protection from HSV-2 infection are still under investigation. Recently, the use of molecular adjuvants to drive vaccine responses induced by DNA vaccines has been reported in a number of experimental systems. We sought to take advantage of this immunization model to gain insight into the correlates of immune protection in the HSV-2 mouse model system and to further explore DNA vaccine technology. To investigate whether the Th1- or Th2-type immune responses are more important for protection from HSV-2 infection, we codelivered the DNA expression construct encoding the HSV-2 gD protein with the gene plasmids encoding the Th1-type (interleukin-2 [IL-2], IL-12, IL-15, and IL-18) and Th2-type (IL-4 and IL-10) cytokines in an effort to drive immunity induced by vaccination. We then analyzed the modulatory effects of the vaccine on the resulting immune phenotype and on the mortality and the morbidity of the immunized animals following a lethal challenge with HSV-2. We observed that Th1 cytokine gene coadministration not only enhanced the survival rate but also reduced the frequency and severity of herpetic lesions following intravaginal HSV challenge. On the other hand, coinjection with Th2 cytokine genes increased the rate of mortality and morbidity of the challenged mice. Moreover, of the Th1-type cytokine genes tested, IL-12 was a particularly potent adjuvant for the gD DNA vaccination.     

Cocaine and tobacco use and the risk of spontaneous abortion

BACKGROUND: Cigarette smoking and cocaine use may be risk factors for spontaneous abortion, but data supporting such a link are limited. METHODS: We studied the associations between cocaine and tobacco use and spontaneous abortion among pregnant adolescents and women (age range, 14 to 40 years) who sought care at an inner-city emergency department. A total of 400 adolescents and women had spontaneous abortions either at study entry or during follow-up (which lasted until 22 weeks' gestation), and 570 adolescents and women remained pregnant past 22 weeks' gestation. Cocaine use was measured at base line by self-reports and analysis of urine and hair samples. Cigarette smoking was measured by self-reports and urine analysis. RESULTS: The adolescents and women in both groups were predominantly black and of lower socioeconomic status. Among those who had spontaneous abortions, 28.9 percent used cocaine on the basis of hair analysis and 34.6 percent smoked on the basis of a urine cotinine assay, as compared with 20.5 percent and 21.8 percent, respectively, of the adolescents and women who did not have spontaneous abortions. The presence of cocaine in hair samples was independently associated with an increase in the occurrence of spontaneous abortion (odds ratio, 1.4; 95 percent confidence interval, 1.0 to 2.1) after adjustment for demographic and drug-use variables. However, the use of cocaine as measured by self-reports and by urine analysis was not. The presence of cotinine in urine was also independently associated with an increased risk of spontaneous abortion (odds ratio, 1.8; 95 percent confidence interval, 1.3 to 2.6). Twenty-four percent of the risk of spontaneous abortion could be related to cocaine or tobacco use. CONCLUSIONS: Cocaine and tobacco use were common in our study population and were associated with a significant risk of spontaneous abortion.     

Specific delivery of captopril to the kidney with the prodrug captopril-lysozyme

Low-molecular-weight proteins (LMWPs) accumulate in the proximal tubular cells of the kidney, which makes these proteins interesting tools for renal drug targeting. We studied this approach using the LMWP lysozyme as a carrier for the angiotensin-converting enzyme inhibitor captopril. Captopril was conjugated to lysozyme via a disulfide bond. The pharmacokinetics of the captopril-lysozyme conjugate was studied in the rat. Only intact conjugate could be detected in the circulation. The total amount of captopril disulfides in the kidney was six times higher after administration of the conjugate than after the administration of an equivalent amount of free captopril. The conjugate was recovered in the urine partially as intact conjugate and partially as low-molecular-weight disulfides. The excretion of conjugate in the urine was not a consequence of the coupling of captopril to lysozyme because an equivalent bolus dose of native lysozyme was similarly excreted into the urine. By determination of the renal angiotensin-converting enzyme activity, we showed that the conjugate was degraded to the pharmacologically active captopril in vivo. We conclude that the coupling of captopril to the LMWP lysozyme results in increased captopril concentrations in the kidney and reduced captopril concentrations in the circulation.     

Antimicrobial resistance and serotype distribution of Streptococcus pneumoniae strains causing childhood infections in Bangladesh, 1993 to 1997

Three hundred sixty-two Streptococcus pneumoniae strains were isolated from children under 5 years of age at Dhaka Shishu (Children) Hospital from 1993 to 1997. The strains were isolated from blood (n = 105), CSF (n = 164), ear swab (n = 61), eye swab (n = 20), and pus (n = 12). Of the 362 isolates, 42 (11.6%) showed intermediate resistance (MIC, <0.1 microgram/ml) and only 4 (1.1%) showed complete resistance (MIC, >2.0 microgram/ml) to penicillin. Penicillin resistance exhibited a strong relationship with serotype 14; 47.8% of the penicillin-resistant strains belonged to this type. A remarkably high (64.1%) resistance to co-trimoxazole was observed, along with a significant increase during the time period studied; there was no relationship to capsular type. By way of contrast, penicillin resistance did not show any significant change during the study period. Resistance to chloramphenicol (2.2%) and erythromycin (1.1%) was rare. The high resistance to co-trimoxazole and its increasing trend demand elucidation of the clinical impact of pneumonia treatment by this antimicrobial and reconsideration of the World Health Organization recommendation for co-trimoxazole administration to children with community-acquired pneumonia at the health care worker level in Bangladesh.     

Rat hindlimb muscle blood flow during level and downhill locomotion

During eccentrically biased exercise (e.g., downhill locomotion), whole body oxygen consumption and blood lactate concentrations are lower than during level locomotion. These general systemic measurements indicate that muscle metabolism is lower during downhill exercise. This study was designed to test the hypothesis that hindlimb muscle blood flow is correspondingly lower during downhill vs. level exercise. Muscle blood flow (determined by using radioactive microspheres) was measured in rats after 15 min of treadmill exercise at 15 m/min on the level (L, 0 degrees) or downhill (D, -17 degrees). Blood flow to ankle extensor muscles was either lower (e.g., white gastrocnemius muscle: D, 9 +/- 2; L, 15 +/- 1 ml. min-1. 100 g-1) or not different (e.g., soleus muscle: D, 250 +/- 35; L, 230 +/- 21 ml. min-1. 100 g-1) in downhill vs. level exercise. In contrast, blood flow to ankle flexor muscles was higher (e.g., extensor digitorum longus muscle: D, 53 +/- 5; L, 31 +/- 6 ml. min-1. 100 g-1) during downhill vs. level exercise. When individual extensor and flexor muscle flows were summed, total flow to the leg was lower during downhill exercise (D, 3.24 +/- 0.08; L, 3.47 +/- 0. 05 ml/min). These data indicate that muscle blood flow and metabolism are lower during eccentrically biased exercise but are not uniformly reduced in all active muscles; i.e., flows are equivalent in several ankle extensor muscles and higher in ankle flexor muscles.