Laboratory diagnosis of Mycoplasma pneumoniae infection. 2. Comparison of methods for the direct detection of specific antigen or nucleic acid sequences in respiratory exudates

The efficiency of the direct detection of Mycoplasma pneumoniae in respiratory exudates by an antigen capture, indirect enzyme immunoassay (Ag-EIA), has been compared with its detection with a cDNA probe ('Gen-Probe assay') directed against the specific ribosomal RNA sequences of the organism ('Mycoplasma pneumoniae Rapid Diagnostic System', Gen-Probe, San Diego, California). Both assays showed excellent specificity against a range of mycoplasma species suspended in negative nasopharyngeal aspirates; only M. pneumoniae and M. genitalium reacted. In experiments with graded doses of viable M. pneumoniae cells suspended in negative nasopharyngeal aspirate, the Gen-Probe assay was more sensitive than Ag-EIA; detection limits were respectively 2 X 10(3) c.f.u./ml (3.2 X 10(5) genomes) and 2.5 X 10(4) c.f.u./ml (4 X 10(6) genomes); detection levels 10-100 times less sensitive than culture. The two assays were also tested on nasopharyngeal aspirates or sputum specimens from 90 patients with respiratory infection; 67 of these were culture- or seronegative for M. pneumoniae and 23 were culture- or seropositive. Ag-EIA detected 21 (91%) of the latter but the Gen-Probe assay detected only 5 (22%). Both assays were negative with the 67 culture-/sero-negatives; there were no Gen-Probe assay positive/Ag-EIA negatives. Overall, it is concluded that although Ag-EIA and the Gen-Probe assay are effective substitutes for culture as a diagnostic procedure, there is a significant problem with samples which are culture-negative and from patients who have good serological evidence of current infection. Possible reasons for the disparity between the two assays are advanced.     

Regional expression of c-fos and heat shock protein-70 mRNA following hypoxia-ischemia in immature rat brain.

Cerebral ischemia induces the expression of a number of proteins that may have an important influence on cellular injury. The purpose of this study was to compare the regional effects of hypoxia-ischemia on the expression of the proto-oncogene, c-fos, and the heat shock protein-70 (HSP-70) gene in developing brain. Unilateral hypoxia-ischemia was produced in the brain of immature rats (7, 15, and 23 days after birth) using a combination of carotid artery ligation and systemic hypoxia (8% O2). After recovery for 2 and 24 h, the regional expression of c-fos and HSP-70 mRNA was determined using in situ hybridization. Littermates were permitted to recover for 1 week for assessment of histologic injury. Hypoxia-ischemia increased the expression of both c-fos and HSP-70 mRNA, but the topography of expression varied with the age of the animal as well as the mRNA species. In the 7-day-old group, expression of c-fos at 2 h increased in multiple regions of the ipsilateral hemisphere in nearly one-half of the animals, while HSP-70 mRNA was not expressed until 24 h and, then, predominantly in the hippocampus. In 15- and 23-day-old rats, expression of c-fos was increased at 2 h in the entorhinal cortex and in the dendritic field of the upper blade of the hippocampal dentate gyrus, while HSP-70 mRNA was prominently expressed in neocortex and the cell layers of the hippocampus. Interestingly, the strong expression of HSP-70 mRNA in dentate granule cells did not occur in the innermost layer of cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

American Indian--Alaska Native youth health.

OBJECTIVE--To assess risk behaviors, health problems, worries and concerns, and resiliency-promoting factors among American Indian-Alaska Native adolescents. DESIGN--Survey. SETTING--Nonurban schools from eight Indian Health Service areas. PARTICIPANTS--A total of 13,454 seventh- through 12th-grade American Indian-Alaska Native youths. MAIN OUTCOME MEASURES--revised version of the Adolescent Health Survey, a comprehensive, anonymous, self-report questionnaire with 162 items addressing 10 dimensions of health. RESULTS--Poor physical health was reported by 2% of the study sample and was significantly correlated with social risk factors of physical and/or sexual abuse, suicide attempts, substance abuse, poor school performance, and nutritional inadequacies. Injury risk behaviors included never wearing seatbelts (44%), drinking and driving (37.9% of driving 10th through 12th graders), and riding with a driver who had been drinking (21.8%). Physical and sexual abuse prevalence was 10% and 13%, respectively, with 23.9% of females reporting physical abuse and 21.6% of females reporting sexual abuse by the 12th grade. Almost 6% of the entire sample endorsed signs of severe emotional distress. Eleven percent of the teens surveyed knew someone who had killed himself or herself, and 17% had attempted suicide themselves. Sixty-five percent of males and 56.8% of females reported having had intercourse by the 12th grade. Weekly or more frequent alcohol use rose from 8.2% of seventh graders to 14.1% by the 12th grade; for males, the survey noted an increase in regular alcohol use of 3% to 5% a year to 27.3% by the 12th grade. For each variable measured, rates are much higher for American Indian adolescents than those for rural white Minnesota youth, except for age at first intercourse and alcohol use. CONCLUSIONS--American Indian-Alaska Native adolescents reported high rates of health-compromising behaviors and risk factors related to unintentional injury, substance use, poor self-assessed health status, emotional distress, and suicide. Interventions must be culturally sensitive, acknowledge the heterogeneity of Indian populations, be grounded in cultural traditions that promote health, and be developed with full participation of the involved communities.     

Recombinant bone morphogenetic protein (BMP)-2 regulates costochondral growth plate chondrocytes and induces expression of BMP-2 and BMP-4 in a cell maturation-dependent manner

This study examined the effect of recombinant human bone morphogenetic protein-2 on several parameters of growth, differentiation, and matrix synthesis and on the endogenous production of mRNA of bone morphogenetic proteins 2 and 4 by growth plate chondrocytes in culture. Chondrocytes from resting and growth zones were obtained from rat costochondral cartilage and cultured for 24 or 48 hours in medium containing 0.05-100 ng/ml recombinant human bone morphogenetic protein-2 and 10% fetal bovine serum. Incorporation of [³H]thymidine, cell number, alkaline phosphatase specific activity, incorporation of [³H]proline into collagenase-digestible protein and noncollagenase-digestible protein, and incorporation of [³⁵S]sulfate were assayed as indicators of cell proliferation, differentiation, and extracellular matrix synthesis. mRNA levels T for bone morphogenetic proteins 2 andv4 were determined by Northern blot analysis. Recombinant human bone morphogenetic protein-2 increased the incorporation of [³H]thymidine by quiescent resting-zone and growth-zone cells in a similar manner, whereas it had a differential effect on nonquiescent cultures. At 24 and 48 hours, 12.5-100 ng/ml recombinant human bone morphogenetic protein-2 caused a dose-dependent increase in cell number and DNA synthesis in resting-zone chondrocytes. No effect was seen in growth-zone cell Recombinant human bone morphogenetic protein-2 stimulated alkaline phosphatase specific activity in resting-zone chondrocytes in a bimodal manner, causing significant increases between 0.2 and 0.8 ng/ml and again between 25 and 100 ng/ml. In contrast, alkaline phosphatase specific activity in growth-zone chondrocytes was significantly increased only between 12.5 and 100 ng/ml. Recombinant human bone morphogenetic protein-2 increased the production of both collagenase-digestible protein and noncollagenase-digestible protein by resting-zone and growth-zone cells, but incorporation of [³⁵S]sulfate was unaffected. Administration of recombinant human bone morphogenetic protein-2 also increased incorporation of [³H]uridine in both resting-zone and growth-zone chondrocytes; these cells produced mRNA for bone morphogenetic proteins 2 and 4. Bone morphogenetic protein-2 mRNA levels in both resting-zone and growth-zone chondrocytes increased in the presence of recombinant human bone morphogenetic protein-2; however, bone morphogenetic protein-4 mRNA levels in growth-zone cells decreased under its influence, and those in resting-zone cells were upregulated only with a dose of 10 ng/ml. This indicates that recombinant human bone morphogenetic protein-2 regulates chondrocyte proliferation, differentiation, and matrix production, and the effects are dependent on the stage of cell maturation. Resting-zone chondrocytes were more sensitive, suggesting that they are targeted by bone morphogenetic protein-2 and that this growth factor may have autocrine effects on these cells.