Screening and instability of FMR1 alleles in a prospective sample of 24,449 mother-newborn pairs from the general population.

To study the instability of FMR1 triplet repeats in the general population, we screened a prospective sample of 24,449 anonymized mother-offspring pairs and analyzed transmissions of intermediate-size (45-54 triplets) and premutation-size (55-200 triplets) alleles. We screened all mothers for alleles > or = 45 triplets by Southern blot and studied transmission of 545 maternal alleles to their offspring using polymerase chain reaction. Out of 21,411 maternal samples with conclusive results, we identified 250 carriers of at least one intermediate-size allele and 39 carrying a premutation-size allele. Out of a subsample of 430 transmissions of normal-size alleles (< 45 triplets), we observed four (< 1%) unstable transmissions. There were 6/90 intermediate-size unstable alleles (7%) and 11/25 unstable premutation-size alleles (44%). Two mothers transmitted a typical full mutation. The incidence of fragile X syndrome was thus 1/12,225 newborns (upper limit of 95% confidence interval: 1/4638 newborns), but larger in males (1/6209) than females (none detected in over 12,000 newborn females). Intermediate-size alleles were more unstable than normal-size alleles (p = 0.0027), but more stable (about sixfold) than premutation-size alleles (p < 0.0001). Unstable premutation-size alleles harbored the major fragile X haplotype (T50-T42-T62), and this haplotype appeared to be a good predictor of instability in premutations (p = 0.02). Incidence and instability are important to determine the feasibility and cost effectiveness of putative FMR1 screening programs. Carriers of FMR1 alleles of 55+ triplets with no family history of the disease may have a significant risk of expansion to a full mutation in a single generation.

High throughput and economical mutation detection and RFLP analysis using a minimethod for DNA preparation from whole blood and acrylamide gel electrophoresis.

We report a simple, rapid, and high throughput method which allows the simultaneous processing of multiple whole blood samples for routine DNA purification and analysis. The method is based on a microscale DNA preparation and digestion using minimal amounts of reagents and handling. The amount of material necessary for a Southern blot analysis (5-7 micrograms) is obtained from 200 microliters of whole blood. All steps involved in DNA preparation and restriction digestion are processed in a single 1.5-ml Eppendorf tube. DNA preparation is performed using a salting out procedure with a proteinase K digestion step but no phenol/chloroform extraction. Restricted fragments are separated by electrophoresis through polyacrylamide slab gels followed by electrotransfer to nylon membranes. By varying the electrophoresis parameters (V/cm or duration), fragments of interest up to 12 kb length can be separated with high resolution. At least 80 samples can be processed at once per DNA preparation, and multiples of this number depend on the available equipment. This economical and rapid method allows routine DNA analysis for mutation or RFLP detection to be performed on a large scale which is a mandatory feature in any DNA-based population screening program. In addition, the DNA purified by the minimethod can be used as an economical source for PCR analysis.

Cardiovascular responses during recovery from exercise and thermal stress.

Although the central cardiovascular adjustments to exercise in the heat have been identified, little is known about the post-exercise hemodynamics during recovery from exercise and heat stress. This study examined heart rate (HR), stroke index (SI), cardiac index (CI), systemic vascular resistance (SVR), systolic (SBP) and diastolic (DBP) blood pressure in 8 males during 15 min of passive seated recovery preceded by 30 min of cycle ergometry (60% VO2max) on two separate occasions: under control (C) and heat stress (HS) conditions. During both recovery conditions, SI significantly declined (p < 0.05) to below pre-exercise values. No differences were observed between groups with respect to SI. The decrease in recovery HR was slower (p < 0.05) in HS than C. The greater elevation in HR during HS accounted for the relative increase in CI above that observed prior to exercise. The estimated SVR measured immediately following exercise in both groups was lower (p < 0.05) than pre-exercise values. By 5 min of C recovery, SVR returned to baseline values but remained significantly depressed (p < 0.05) for the entire HS condition. These results indicate that the pressor responses were attenuated during HS; however, CI was maintained above pre-exercise levels due to higher HR responses compensating for the reduction in SI. Stimulation of the baroreceptor reflex and increased myocardial contractility could possibly explain the maintenance of output at a time when preload and afterload were reduced.

Facial cooling does not benefit cardiac dynamics during recovery from exercise hyperthermia.

Cardiovascular responses during passive recovery following exercise-induced hyperthermia were investigated in male subjects (n = 8) on two separate occasions; once with facial cooling (FC; 2 degrees C, 2.5 m/s) and once without (NC). Recovery heart rate (HR) responses were significantly (p less than 0.05) lower during the FC than NC condition. No statistical differences were observed between groups with respect to stroke volume (SV), cardiac output (CO), systemic vascular resistance (SVR), and mean arterial blood pressure (MAP). After 5 min of recovery, FC caused a significant (p less than 0.05) reduction in HR however, the NC group response remained unchanged. The decline in HR with FC was moderately associated with the decrease in forehead temperature (r = 0.69; p less than 0.05) but poorly correlated with rectal temperature (r = 0.09; p greater than 0.05). As expected, SV, CO, and MAP significantly declined (p less than 0.05) while the estimated SVR progressively increased (p less than 0.05) over time. The results suggest that FC is not beneficial in terms of improving central and peripheral cardiovascular dynamics during recovery from exercise-induced hyperthermia.