Lack of Association Between ACE2 Expression and Serum Testosterone Concentrations in Peripheral Mononuclear Cells in Males

Background: Male sex is a risk factor for developing severe COVID-19 illness, hospitalization, and mortality. It is possible that the male sex hormone, testosterone, contributes to the morbidity from COVID-19. SARS-CoV2 viruses use cell membrane protein Angiotensin-Converting Enzyme 2 (ACE2) receptor and undergo S protein priming by the Type II Transmembrane Serine Protease (TMPRSS2) to enter the cells. Hence, the expression level of ACE2 and TMPRSS2 may affect disease susceptibility and possible severity. TMPRSS2 is regulated by the androgen receptor. We, therefore, examined if an association exists between serum testosterone concentrations and ACE2 or TMPRSS2 expression level in men. Methods: We analyzed fasting serum samples and peripheral blood mononuclear cells (MNC) from 42 men. Total and free testosterone and estradiol were measured by liquid chromatography/equilibrium dialysis. Sex hormone binding globulin (SHBG) was measured by chemiluminescence. mRNA was prepared from MNC. Quantitative RT-PCR was conducted using commercially available, pre-designed TaqMan primers and probes targeting ACE2. The ACE2 relative level was calculated after normalization to beta Actin and GAPDH, with lowest ACE2 level being set to 1. Results: Subjects’ age ranged from 20 to 65 years. Type 2 diabetes was present in 74% of the men and the mean HbA1 was 7.2±1.6% (mean ± S.D.). Fifteen subjects had subnormal free testosterone (<50 pg/ml). Compared to the 27 subjects with normal free testosterone, they were older (49±12 vs 40±13 years, p=0.03) but had similar BMI (36±10, 35±10 kg/m(2), p=0.71). As expected, they had lower total testosterone (Median [25(th), 75(th) percentile]; 222 [171-266] vs 431 [335, 618] ng/dl, p<0.001) and free testosterone concentrations (39 [21, 44] vs 72 [59, 92], p<0.001). Total estradiol was also lower in this group (19±1 vs 29±13 pg/ml, p=0.03) but free estradiol (0.44±0.33 vs 0.56±0.34 pg/ml, p=0.44) and SHBG (27 [19, 33] vs 30 [21, 39] nmol/L, p=0.52) were similar. Quantitative PCR data showed there was a large inter-individual variation of ACE2 expression level, up to 15-fold difference. Average ACE2 level did not differ between subnormal and normal testosterone groups (3.4 [2.7, 5.0] vs 3.9 [2.5, 5.7] arbitrary units). ACE2 expression was not related to free testosterone (r=0.11), free estradiol (r=0.18) or sex hormone binding globulin (r=0.15) on linear regression analyses (p>0.30 for all). ACE2 expression was also not related to age (r= -0.15, p=0.34), BMI (r= -0.2, p=0.23) or presence of diabetes. We were not able to detect a significant expression of TMPRSS2 in MNC. Conclusion: Our results do not support a role for testosterone or estradiol as regulators of ACE2 expression in peripheral blood MNC in males.