Steroid esters are a frequently used ergogenic aid among athletes, and their direct detection represents an important approach to anti-doping efforts. However, the presence of esterase enzymes in blood complicates these efforts. Although dried blood spots (DBS) have emerged as a promising solution to the esterase problem, the limited sample volume creates both analytical and practical challenges. The larger collection volume for liquid blood may allow for higher analytical sensitivity, thus offsetting the effects of esterase activity in practical scenarios. First, to evaluate the effects of blood esterases ex vivo, 10 individuals were administered a single dose of either testosterone propionate or testosterone cypionate. Venous blood was collected into serum separator tubes (SST) serum and NaF-containing tubes and stored at varying conditions. Both esters significantly decreased in serum while no ester loss was observed in the NaF-containing plasma. Next, to compare the detectability across matrices stored at conditions meant to mimic real-world scenarios, the individuals administered testosterone propionate collected capillary serum, capillary NaF-containing plasma, and capillary DBS at various times postadministration. Each sample type was stored at different conditions designed to mimic what may be encountered during real-world shipping scenarios. Similar detectability of testosterone propionate was observed between all NaF, DBS, and SST samples stored under cool conditions, whereas detectability was better in NaF and DBS samples under ambient and warm conditions. These results indicate that for samples not shipped in temperature regulated conditions, DBS and NaF-containing plasma are the preferred matrices.

BACKGROUND: A lack of safety and dosing information on restoring hormone levels in neutered dogs has hindered treatment of spay-neuter syndrome, the diverse set of health and behavior conditions resulting from the loss of gonadal hormones. The purpose of this study was to provide basic information on the use of testosterone treatment in male neutered dogs using a target animal safety study design. Twelve previously neutered dogs were divided into four equal groups, receiving 0 (controls), 1x, 3x, or 5x the standard weekly dose of injectable testosterone cypionate (0.5 mg/kg) for 90 days. Bloodwork, including endocrine assays, body condition, prostate health, and qualitative behavioral observations were used to evaluate any changes in health during treatment. RESULTS: Testosterone levels increased in a dose-dependent fashion, and luteinizing hormone decreased after a month of treatment in dogs receiving 5x the standard dose. Behavioral measures, Zambelli prostate health score, body condition scores, clinical evaluations, and routine blood hematology and chemistries showed minor variation over time or across groups. Two seizures were documented in a dog from group 3x with previous suspected idiopathic epilepsy. CONCLUSIONS: The results of the initial safety study indicated that weekly injectable testosterone therapy over a 3-month period was safe and capable of increasing testosterone levels in neutered dogs to within the normal range for intact dogs. The limitations of the study included small group sizes and no long-term follow up. A summary of the risks and recommendations for the use of testosterone restoration in dogs was provided.

The abusive use of anabolic androgenic steroids has become a serious health problem worldwide, but its effects on oral health are still poorly understood. Therefore, the objective of this study was to evaluate the effects of a supraphysiological dose of testosterone cypionate (TC) on salivary biochemical, histomorphology, immunohistochemistry, and redox state parameters of parotid and submandibular glands. Twenty male Wistar rats, 12 weeks old, were divided into two groups (n=10/group): a control group and TC group, which received a dose of 20 mg/kg, once a week, for 6 weeks. Post treatment, the saliva and glands were collected. A supraphysiological dose of TC increased plasma and salivary testosterone concentrations. Although TC did not alter salivary flow, pH, and buffering capacity, the treatment increased the salivary secretion of total protein and reduced amylase, calcium, phosphate, and potassium. TC reduced the connective tissue area in the parotid gland and acinar area of the submandibular gland, while increasing the granular convoluted tubule area in the submandibular gland. Proliferating cell nuclear antigen was higher in the acinar cells of the submandibular glands from the TC group. Moreover, TC increased concentrations of total oxidant capacity and damaged lipids in both salivary glands, while total antioxidant activity and uric acid were lower in the submandibular gland, and reduced glutathione was higher in both glands. Superoxide dismutase, catalase, and glutathione peroxidase activities were higher in the parotid gland, while only glutathione peroxidase activity was lower in the submandibular gland of the TC group. In conclusion, TC abuse may be a potential factor for dysfunction of the parotid and submandibular glands, becoming a risk factor for the oral and systemic health of users.

Transgender men undergoing hormone therapy are at risk for insulin resistance. However, how virilizing testosterone therapy affects serum insulin and peripheral insulin sensitivity in transgender men is unknown. This study assessed the effect of acute, virilizing testosterone on serum insulin concentrations and insulin signaling in liver, skeletal muscle, and white adipose tissue (WAT) of female pigs as a translational model for transgender men. Females received three doses of intramuscular testosterone cypionate (TEST females; 50 mg/day/pig) or corn oil (control) spaced 6 days apart starting on the day of estrus (D0). Fasting blood was collected on D0, D3, D5, D11, and D13, and females were euthanized on D13. On D13, TEST females had virilizing concentrations of serum testosterone with normal concentrations of serum estradiol. Virilizing serum testosterone concentrations (D13) were associated with decreased serum insulin and C-peptide concentrations. Blood glucose and serum glycerol concentrations were not altered by testosterone. Virilizing concentrations of testosterone downregulated AR and ESR1 in subcutaneous (sc) WAT and upregulated transcript levels of insulin-signaling pathway components in WAT and liver. At the protein level, virilizing testosterone concentrations were associated with increased PI3K 110α in liver and increased insulin receptor (INSR) and phospho(Ser256)-FOXO1 in visceral (v) WAT but decreased phospho(Ser473)-AKT in vWAT and scWAT. These results suggest that acute exposure to virilizing concentrations of testosterone suppresses circulating insulin levels and results in increased abundance of proteins in the insulin-signaling pathway in liver and altered phosphorylation of key proteins in control of insulin sensitivity in WAT.NEW & NOTEWORTHY Acute virilizing doses of testosterone administered to females suppress circulating insulin levels, upregulate components of the insulin-signaling pathway in liver, and suppress insulin signaling in white adipose tissue. These results suggest that insulin resistance in transgender men may be due to suppression of the insulin-signaling pathway and decreased insulin sensitivity in white adipose tissue.