Monitoring of Bone Loss Biomarkers in Human Sweat: A Non-Invasive, Time Efficient Means of Monitoring Bone Resorption Markers under Micro and Partial Gravity Loading Conditions

The overall goal of this project was to validate the concept that the rate and extent of unloading-induced bone loss in humans can be assessed by monitoring the levels of two bone resorption markers in sweat, namely ionized calcium (Ca2+) and total collagen cross-links (T-CCL) (i.e., the pyridinium cross-links PYD & DPD) The original funded project plan called for a phased approach consisting of three phases; the first phase focused on selection of the most appropriate and efficient means of collecting a sweat sample from an individual compatible with the microgravity environment of space flight coupled with biochemical validation that these sweat samples contained bone resorption markers at levels capable of being detected using standard laboratory analysis techniques; the second phase focused on validation of the concept that bone resorption marker levels detected in sweat samples accurately and consistently reflected circulating levels and/or urine levels of these biomarkers; and the third phase focused on longitudinal assessment of bone resorption marker level in sweat, blood, and urine in young and old populations undergoing active bone formation or bone loss, respectively.

After successful completion of Phase I, preliminary data generated during Phase II indicated that the NASA criterion measure for bone loss during space flight (i.e., 24 hr urinary ionized calcium excretion), while related to calcium and T-CCL levels in sweat samples actively produced during defined exercise, were not predictive of 24 hr urine calcium excretion rates. After consultation and review of the preliminary Phase II results by representatives of the NASA-Johsnon Space Center-Human Research Program (JSC-HRP), the focus of Phase II was redirected to explore collection of a 24 hr sweat sample, rather than collection of a discrete, exercise-induced “active” sweat sample, to determine if a 24 hr integrated sweat sample was predictive of biomarker concentrations found in 24 hr urine samples. This redirection of effort required the identification and validation of additional commercially available absorbant materials which did not contain endogenous biomarker signal as well as a means of extracting the biomarkers from the absorbant material compatible with fluid handling limitations in the space flight environment. After identifying and developing such a collection method, this approach was then utilized to answer the question of whether or not biomarker levels in an integrated 24 hr sweat sample was predictive of those found in a concurrent 24 hr urine sample in a convenience of healthy individuals.

After successful completion of Phase II, the NASA-JSC-HRP program indicated that they wished to deploy the sweat monitoring technology in a NASA bed-rest campaigns being performed at the GCRC at University of Texas Medical Branch (UTMB) instead of in young and old subject populations as originally planned. Unfortunately however, during the last 9 months of the project NASA had to postpone bed-rest operations resulting in a joint decision by NASA-HRP and the Principal Investigator to utilize spinal-injured patients recruited from the Texas Medical Center (rather than NASA bed rest subjects) in which to test the 24 hr sweat monitoring technology as means of assessing bone loss. The resulting time delay surrounding availability of bed rest subjects and the subsequent decision to utilize spinal chord injury (SCI) patients, coupled with the additional requirement to seek Committee for the Protection of Human Subjects (CPHS) approval for testing in a new subject population resulted in NASA granting a one year no-cost extension to the project. Limited data gathered in the final year of the project provides evidence that biomarker levels in a 24 hr integrated sweat sample are predictive of those levels found in 24 hr urine samples in SCI patients. These data indicate that 24 hr sweat sample collection (using a collection and analysis scheme compatible with space flight operations) in a terrestrial human population undergoing active bone loss in a similar fashion to crew members during space flight is a non-invasive, time-efficient alternative to on-orbit 24 hr urine void collection as means of assessing biomarkers of bone loss. In addition, the ability to perform this type of sample collection using a microgravity compatible approach to liquid sample handling and the use of simple colorimetric based analysis techniques is of notable operational relevancy. Our data and validated methodologies suggest that sweat biomarker analysis (as an operationally compatible means of assessing and/or monitoring bone loss in crew members during space flight) should be considered for further development as a “real-time” analytical method for assessment of space flight-induced bone loss and a valid means of monitoring the efficacy of “in-flight” bone loss countermeasures.

[Editor's note 3/27/2013: No Task Book report received. Progress section and Bibliography compiled from PI's Final Technical Report submitted January 2013]