Metabolic assessment of wallaby blastocysts during embryonic diapause and subsequent reactivation

The metabolism of tammar wallaby (Macropus eugenii) blastocysts was analysed by means of quantitative fluorescence microscopy during embryonic diapause and 2, 3, 4, 5, 8 and 10 days after reactivation to determine nutrient preferences during metabolic reactivation of the blastocyst. The surface area of quiescent blastocysts was 0.16 +/- 0.02 mm2 (mean +/- s.e.m.), and increased to 0.44 +/- 0.04 mm2 (P < 0.05) by Day 8 after removal of the sucking stimulus of the pouch young (RPY). Day-10 blastocysts, analysed over two successive breeding seasons, were significantly different in size from each other (Group A, 1992: 4.44 +/- 1.47 mm2; Group B, 1993: 18.87 +/- 4.62 mm2; P < 0.01), and both groups were significantly different in size from diapausing blastocysts (P < 0.01). There was no significant difference in carbohydrate uptake or production by blastocysts during the first five days after RPY. Glucose uptake by blastocysts recovered 8 days after RPY (61.9 +/- 30.0 pmol embryo-1 h-1) was significantly greater than that by Day-0 blastocysts (17.9 +/- 5.5 pmol embryo-1 h-1) and glucose uptake by both groups of Day-10 blastocysts (Group A, 174.0 +/- 28.4 pmol embryo-1 h-1; Group B, 616.0 +/- 239.0 pmol embryo-1 h-1) was significantly different from that by Day-0 blastocysts (P < 0.01). Pyruvate uptake by Day-10 blastocysts (Group A, 46.0 +/- 32.2 pmol embryo-1 h-1; Group B, 250.0 +/- 136.0 pmol embryo-1 h-1; P < 0.01) increased significantly compared with that by Day-0 blastocysts (6.4 +/- 1.6 pmol embryo-1 h-1; P < 0.01). Lactate production by Day-10 blastocysts (Group A, 186.7 +/- 30.3 pmol embryo-1 h-1; Group B, 285 +/- 129 pmol embryo-1 h-1; P > 0.01) was also significantly different from that by quiescent blastocysts (41.20 +/- 9.6 pmol embryo-1 h-1). There was a linear relationship between surface area and glucose uptake and surface area and pyruvate uptake (r2 = 0.965 and r2 = 0.971 respectively). Despite increases in carbohydrate uptake, there was a proportional decrease in lactate production indicating an increase in oxidative metabolism during reactivation. This suggests that there may be a metabolic switch at, or around, Day 5 after RPY.     

Factors associated with sexual behavior problems in young sexually abused children

OBJECTIVE: To identify variables associated with the presence of sexual behavior problems in young sexually abused children. METHOD: Data were gathered from the clinical records of 100 sexually abused boys and girls ages 3-7 years enrolled in two treatment programs. Information was coded systematically on approximately 350 areas related to the child and family's history and functioning, the sexual abuse experience, and treatment outcome. The children were grouped and compared according to their presenting sexual behavior into three categories: (1) developmentally "expected"; (2) "sexualized/self-focused"; and (3) problematic "interpersonal" sexual behavior. RESULTS: Bivariate and multivariate analyses highlighted five variables which were predictive of sexual behavior problems among sexually abused children. Sexual arousal of the child during his/her sexual abuse, the perpetrator's use of sadism, and a history of physical and emotional abuse differentiated between those children with and without "interpersonal" sexual behavior problems. Who the child blamed for his/her sexual abuse further contributed to the distinction between children whose sexual behavior was exclusively "self-focused" (sexualized) versus "interpersonal." CONCLUSIONS: The five major predictor variables, as well as other variables identified in this study, have potential utility in assessing child risk for negative outcomes and determining referral priorities for sexual abuse treatment. Given that sexual arousal and who the child blames for the abuse are prominent variables associated with sexual problems and self-blame, clinicians will need to ensure that sexually abused children and their caregivers are given specific opportunities to deal with these areas in the supportive context of treatment. Children with sexual behavior problems differ not only in the type and level of sexual behavior they exhibit but in most other areas as well, suggesting a need for differential assessment and individualized treatment approaches.     

Instrumented measurement of patellar mobility

To provide an objective analysis of medial and lateral patellofemoral laxity, we examined 94 uninjured athletic subjects and 22 patients with unilateral lateral patellar dislocation. We developed an instrument to measure the compliance of the medial and lateral patellar restraints. The instrument recorded the force-displacement relationship as the patella was pushed medially and laterally. Subtracting the medial displacement from the lateral displacement at a given force level allowed the tester to assess the peripatellar soft tissue "balance." The results for both the 2.5- and the 5-pound tests were significant. Paired comparisons differentiated the three groups, with significant differences between control and affected (P = 0.0001), control and contralateral (P = 0.0036), and affected and contralateral (P = 0.0157) knees. The mean result of the lateral minus medial displacement test for our sample population of control subjects was -2.1 mm for the 5-pound test. A negative value in this test indicates that medial displacement exceeds lateral displacement. This finding was present in 81% of control subjects. In contrast, the mean result for the patients' affected knees was +3.2 mm for the 5-pound test. Using the value of 0.0 mm as the diagnostic determinant for peripatellar imbalance, we found a test sensitivity of 91% and a specificity of 81%.     

Neonatal hypoglycemia, Part I: Background and definition

Hypoglycemia in the neonate remains a common problem. The association of low blood glucose concentrations and abnormal development has prompted extensive research into the anticipation, evaluation, and treatment of neonatal hypoglycemia. Glucose homeostasis in the fetus and neonate is a developmentally regulated dynamic process involving a number of intricate physiologic mechanisms. In addition, the determination of glucose concentrations is dependent upon both the type of tissue analyzed and the limitations of the specific method employed. The complexity of glucose metabolism makes it difficult to precisely define "normal" and "abnormal" glucose levels in preterm and term neonates.     

Major increase in endopeptidase activity of human cathepsin B upon removal of occluding loop contacts

The main feature distinguishing cathepsin B from other cysteine proteases of the papain family is the presence of a large insertion loop, termed the occluding loop, which occupies the S' subsites of the enzyme. The loop is held in place mainly by two contacts with the rest of the enzyme, involving residues His110 and Arg116 on the loop that form salt bridges with Asp22 and Asp224, respectively. The influence of this loop on the endopeptidase activity of cathepsin B has been investigated using site-directed mutagenesis and internally quenched fluorogenic (IQF) substrates. Wild-type cathepsin B displays poor activity against the substrates Abz-AFRSAAQ-EDDnp and Abz-QVVAGA-EDDnp as compared to cathepsin L and papain. Appreciable increases in kcat/KM were observed for cathepsin B containing the single mutations D22A, H110A, R116A, and D224A. The highest activity however is observed for mutants where both loop to enzyme contacts are disrupted. For the triple-mutant D22A/H110A/R116A, an optimum kcat/KM value of 12 x 10(5) M-1 s-1 was obtained for hydrolysis of Abz-AFRSAAQ-EDDnp, which corresponds to a 600-fold increase relative to wild-type cathepsin B and approaches the level of activity observed with cathepsin L or papain. By comparison, the mutations have little effect on the hydrolysis of Cbz-FR-MCA. The influence of the mutations on the pH dependency of activity also indicates that the complexity of pH activity profiles normally observed for cathepsin B is related to the presence of the occluding loop. The major increase in endopeptidase activity is attributed to an increase in loop "flexibility" and suggests that the occluding loop might move when an endopeptidase substrate binds to the enzyme. The possible contribution of these interactions in regulating endopeptidase activity and the implications for cathepsin B activity in physiological or pathological conditions are discussed.