High-throughput ultra-high-performance liquid chromatography/tandem mass spectrometry quantitation of insulin-like growth factor-I and leucine-rich alpha-2-glycoprotein in serum as biomarkers of recombinant human growth hormone administration.

Insulin-like growth factor-I (IGF-I) is a known biomarker of recombinant human growth hormone (rhGH) abuse, and is also used clinically to confirm acromegaly. The protein leucine-rich alpha-2-glycoprotein (LRG) was recently identified as a putative biomarker of rhGH administration. The combination of an ACN depletion method and a 5-min ultra-high-performance liquid chromatography/tandem mass spectrometry (uHPLC/MS/MS)-based selected reaction monitoring (SRM) assay detected both IGF-I and LRG at endogenous concentrations. Four eight-point standard addition curves of IGF-I (16-2000 ng/mL) demonstrated good linearity (r(2) = 0.9991 and coefficients of variance (CVs) <13%). Serum samples from two rhGH administrations were extracted and their uHPLC/MS/MS-derived IGF-I concentrations correlated well against immunochemistry-derived values. Combining IGF-I and LRG data improved the separation of treated and placebo states compared with IGF-I alone, further strengthening the hypothesis that LRG is a biomarker of rhGH administration. Artificial neural networks (ANNs) analysis of the LRG and IGF-I data demonstrated an improved model over that developed using IGF-I alone, with a predictive accuracy of 97%, specificity of 96% and sensitivity of 100%. Receiver operator characteristic (ROC) analysis gave an AUC value of 0.98. This study demonstrates the first large scale and high throughput uHPLC/MS/MS-based quantitation of a medium abundance protein (IGF-I) in human serum. Furthermore, the data we have presented for the quantitative analysis of IGF-I suggest that, in this case, monitoring a single SRM transition to a trypsin peptide surrogate is a valid approach to protein quantitation by LC/MS/MS.

The role of lactate in the exercise-induced human growth hormone response: evidence from McArdle disease.

PURPOSE: Increased blood lactate concentration has been suggested as a primary stimulus for the exercise-induced growth hormone response (EIGR). Patients with McArdle disease are unable to produce lactate in response to exercise and thus offer a unique model to assess the role of lactate in the EIGR. Accordingly, McArdle's patients were exercised to test the hypothesis that lactate is a major stimulus of the EIGR. METHODS: 11 patients with McArdle disease (3 male, 8 female; age: 35.5 (SD 13.9) years, height: 166 (8) cm, body mass: 75.2 (13.1) kg) were recruited for the study. The patients walked initially at 0.42 m/s, increasing by 0.14 m/s per 3 min stage. Exercise was terminated when participants completed 3 minutes at 1.80 m/s or when a Borg CR10 pain scale rating of "4" was reached. Stages were separated by 60 s for capillary blood sampling for analysis of hGH and blood lactate concentration. RESULTS: McArdle's patients' blood lactate levels remained at resting levels (0.3-1.2 mmol/l) as exercise intensity increased. Nine out of 11 participants failed to demonstrate an EIGR obtaining hGH values below the clinical definition of a response (>3 microg/l). CONCLUSION: The absence of an EIGR in nine out of 11 participants suggests that lactate could play a major role in the EIGR in humans.

The detraining and retraining of an elite rower: a case study.

A heavyweight male rower, and current Olympic champion, undertook a laboratory-based incremental rowing test on four separate occasions; eight weeks prior to the Sydney Olympics (Pre OG), after eight weeks of inactivity (Post-IA), after 8 weeks of retraining (Post 8) and after a further 12 weeks of training (Post 20). Following the period of inactivity, peak oxygen uptake (VO2peak) declined by 8%, power at reference blood lactate concentrations declined by approximately 100 W (25%), and power at VO2peak was 20% lower. With eight weeks of retraining, rapid improvements were seen. For most parameters, however, the rate of improvement slowed and after 20 weeks of retraining the individual was approaching pre-Olympic levels. VO2 at lactate threshold as a percentage of VO2peak remained unchanged. These results show that detraining in the elite athlete can be pronounced, with rapid improvements upon retraining which slow, so that retraining takes considerably longer to achieve than detraining did. Complete cessation of training should be limited to short periods only in the preparation of the elite heavyweight rower. Any break should, if possible, include 'maintenance training'. In this way any decrements in those physiological parameters associated with 2000 m rowing performance will be minimised.

The validity of capillary blood sampling in the determination of human growth hormone concentration during exercise in men.

BACKGROUND: Studies measuring human growth hormone (hGH) in blood during exercise have mainly used venous sampling. The invasive nature of this procedure makes evaluation of hGH impossible in various exercise environments. OBJECTIVE: To determine whether capillary sampling could offer an alternative sampling method. METHODS: Capillary and venous blood samples were collected for determination of hGH at the end of each exercise stage during an incremental exercise test in 16 male club level competitive cyclists (mean (SD) age 30.8 (8.0) years, body mass 72.2 (7.1) kg, body fat 12.9 (3.5)%, peak oxygen consumption 4.18 (0.46) l x min(-1)). Linear regression, from a plot of venous v capillary blood hGH concentration, showed a correlation coefficient of r = 0.986 (p<0.001). When geometric means and log transformations were used, a coefficient of variation of 14.2% was demonstrated between venous and capillary flow for hGH concentration. The mean ratio limits of agreement were 0.62 (1.72)-that is, 95% of the ratios were contained between 0.36 and 1.07, with a mean of 0.62. CONCLUSIONS: Capillary blood sampling is an acceptable alternative to venous sampling for determining hGH concentration during rest and exercise. Sample sites should not be used interchangeably: one site should be chosen and its use standardised.