Male C57BL/6J mice have higher presence and abundance of Borrelia burgdorferi in their ventral skin compared to female mice.

Borrelia burgdorferi is a tick-borne spirochete that causes Lyme disease in humans. The host immune system controls the abundance of the spirochete in the host tissues. Recent work with immunocompetent Mus musculus mice strain C3H/HeJ found that males had a higher tissue infection prevalence and spirochete load compared to females. The purpose of this study was to determine whether host sex and acquired immunity interact to influence the prevalence and abundance of spirochetes in the tissues of the commonly used mouse strain C57BL/6. Wildtype (WT) mice and their SCID counterparts (C57BL/6) were experimentally infected with B. burgdorferi via tick bite. Ear biopsies were sampled at weeks 4, 8, and 12 post-infection (PI) and five tissues (left ear, ventral skin, heart, tibiotarsal joint of left hind leg, and liver) were collected at necropsy (16 weeks PI). The mean spirochete load in the tissues of the SCID mice was 260.4x higher compared to the WT mice. In WT mice, the infection prevalence in the ventral skin was significantly higher in males (40.0 %) compared to females (0.0 %), and the spirochete load in the rear tibiotarsal joint was significantly higher (4.3x) in males compared to females. In SCID mice, the spirochete load in the ventral skin was 200.0x higher in males compared to females, but there were no significant sex-specific difference in spirochete load in the other tissues (left ear, heart, tibiotarsal joint, or liver). Thus, the absence of acquired immunity greatly amplified the spirochete load in the ventral skin of male mice. It is important to note that the observed sex-specific differences in laboratory mice cannot be extrapolated to humans. Future studies should investigate the mechanisms underlying the male bias in the abundance of B. burgdorferi in the mouse skin.

Using a testis regeneration model, FGF9, LIF, and SCF improve testis cord formation while RA enhances gonocyte survival.

Implantation of testis cell aggregates from various donors under the back skin of recipient mice results in de novo formation of testis tissue. We used this implantation model to study the putative in vivo effects of six different growth factors on testis cord development. Recipient mice (n = 7/group) were implanted with eight neonatal porcine testis cell aggregates that were first exposed to a designated growth factor: FGF2 at 1 µg/mL, FGF9 at 5 µg/mL, VEGF at 3.5 µg/mL, LIF at 5 µg/mL, SCF at 3.5 µg/mL, retinoic acid (RA) at 3.5 × 10-5 M, or no growth factors (control). The newly developed seminiferous cords (SC) were classified based on their morphology into regular, irregular, enlarged, or aberrant. Certain treatments enhanced implant weight (LIF), implant cross-sectional area (SCF) or the relative cross-sectional area covered by SC within implants (FGF2). RA promoted the formation of enlarged SC and FGF2 led to the highest ratio of regular SC and the lowest ratio of aberrant SC. Rete testis-like structures appeared earlier in implants treated with FGF2, FGF9, or LIF. These results show that even brief pre-implantation exposure of testis cells to these growth factors can have profound effects on morphogenesis of testis cords using this implantation model.

Brief exposure of neonatal testis cells to EGF or GDNF alters the regenerated tissue.

We have previously shown that implantation of testis cell aggregates under the back skin of immunodeficient mice results in de novo regeneration of testis tissue. We used this unique model to investigate the effects of epidermal growth factor (EGF) and glial cell-derived neurotrophic factor (GDNF) on testis cord development. Neonatal piglet testis cells were briefly (<1 h) exposed to either low (L: 0.02 µg/mL) or high (H: 2 µg/mL) doses of EGF, GDNF, or vehicle (control), before implantation in recipient mice. Randomly selected implants were removed from each mouse at 1, 2, 4, and 8 weeks post-implantation. GDNF-L implants showed increased testis cord development over time, and EGF-L implants had increased cross-sectional area. The ratio of regular cords decreased over time in EGF-H and GDNF-H implants and was replaced by a higher ratio of irregular cords in GDNF-H. EGF-L and GDNF-H implants were quickest to display rete testis-like structures. Overall, the lower dose of each growth factor was more effective than its higher dose in improving the implantation outcomes. This is the first comprehensive assessment of these key growth factors on de novo formation (regeneration) of testis tissue. Lay summary: In recent decades, testicular cancer rates have quadrupled in young men while sperm counts have dropped by half. Both conditions may be related to exposure of fetuses or infants to noxious substances causing disruption of normal testis development. To study the effects of any putative factor on testis development, we established an animal model of testis tissue regeneration. We collected newborn piglet testes after routine castration, used enzymes to completely dissociate testis cells, exposed the cells to two key growth factors (EGF or GDNF), and implanted the cells under the back skin of recipient mice, acting as live incubators. We then examined implant samples after 1, 2, 4, or 8 weeks and assessed testis regeneration. Overall, the high dose of each growth factor had adverse effects on the formation of normal testis. Therefore, this novel implantation model may also be used to study the effects of potentially harmful substances on testis development.

Stationary brush use in naive dairy heifers.

Weaned dairy heifers are often housed in environments with few appropriate outlets for grooming or oral manipulation. Our objective was to characterize brush use by naive heifers, including patterns over time. In phase 1, groups of 4 heifers (n = 13 groups, 146.4 ± 9.1 d old, mean ± standard deviation; SD) were introduced to a bedded pack pen with 4 wall-mounted brushes (25.4 × 6.0 cm with 3.8-cm-long bristles). On d 1, 2, and 6 of exposure, continuous video recordings were used to observe 2 focal heifers per group for brush use (oral manipulation, grooming, and the sum of total brush use; all averaged at the group level). Latency to use a brush upon entering the pen was 3.4 ± 4.9 min (mean ± SD; range: 0.1 to 17.8 min among individuals). Heifers used the brushes for oral manipulation (39.7 ± 17.5% of brush use, mean ± SD) and grooming (60.3 ± 17.5%), primarily of their heads (89.9 ± 5.4% of grooming). In phase 2, heifers were moved in pairs (n = 13 pairs/treatment) to freestall pens either with (brush treatment) or without (control) brushes mounted inside the stalls for the first 5 d of phase 2 (d 8-12 of the study); on the last day (d 13 of the study), brushes were provided in both treatments. On d 8 (brush treatment) and 13 (both treatments), one focal heifer/pen was recorded for the same behaviors as in phase 1. Linear mixed models were used to evaluate brush use patterns across days (phase 1: d 1, 2, and 6; phase 2 brush treatment: d 8 vs. 13) and between treatments on d 13. In phase 1, brush use was greatest on d 1 [45.9 min; 95% confidence interval (CI): 33.2-63.3 min, back-transformed from natural-log values], decreased on d 2 (25.0 min, 95% CI: 18.4-34.0 min), but then remained steady until d 6 (21.0 min, 95% CI: 15.4-28.5 min); the initial reduction in total brush use was due to changes in grooming, but oral manipulation remained relatively static. In phase 2, heifers in the brush treatment showed similar usage on d 8 versus d 13 (3.8 vs. 3.7 min, 95% CI: 1.9-6.8 vs. 1.9-6.5 min). Compared with heifers with continuous brush access on d 8-12, those in the control treatment showed more brush use on d 13, both for oral manipulation (6.6 vs. 2.5 min, 95% CI: 3.8-11.1 vs. 1.3-4.5 min) and grooming (3.5 vs. 1.2 min, 95% CI: 1.9-5.7 vs. 0.5-2.3 min). Our study is the first to characterize stationary brush use in weaned dairy heifers. We conclude that, despite lacking previous experience, heifers use brushes for both grooming and oral manipulation.