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Urethral closure mechanisms during passive increments in intravesicular pressure (P(ves)) were investigated using microtip transducer catheters in urethane-anesthetized female rats. After a block of reflex bladder contractions by spinal cord transection at T8-T9, abruptly raising P(ves) to 20, 40, or 60 cmH(2)O for 2 min induced a bladder pressure-dependent contractile response in a restricted portion of the middle urethra (12.5-15 mm from the urethral orifice) that was abolished by cutting the pelvic nerves bilaterally. In pelvic nerve-intact rats, the bilateral transection of either the pudendal nerves, the nerves to the iliococcygeous/pubococcygeous muscles, or the hypogastric nerves significantly reduced (49-74%) the urethral reflex response induced by passive P(ves) increases, and combined transection of these three sets of nerves totally abolished the urethra-closing responses. In spinal cord-intact rats, similar urethral contractile responses were elicited during P(ves) elevation (20 or 40 cmH(2)O) and were also eliminated by bilateral pelvic nerve transection. After spinal cord and pelvic nerve transection, leak point pressures, defined as the pressure inducing fluid leakage from the urethral orifice during passive P(ves) elevation by either bladder pressure clamping in 2.5-cmH(2)O steps or direct compression of the bladder, were significantly lowered by 30-35% compared with sham-operated (spinal cord-transected and pelvic nerve-intact) rats. These results indicate that 1) passive elevation of P(ves) can elicit pelvic afferent nerve-mediated contractile reflexes in the restricted portion of the urethra mediated by activation of sympathetic and somatic nerves and 2) bladder-to-urethral reflexes induced by passive P(ves) elevation significantly contribute to the prevention of stress urinary incontinence.

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We compared three different methods of testing leak point pressure (LPP) in rats with or without the pudendal nerves and nerves to the iliococcygeus/pubococcygeus muscles transected: (1) sneeze induced with a whisker in the nostril (sneeze LPP), (2) manually increased abdominal pressure (Crede LPP), and (3) increased intravesical pressure using the vertical tilt table method (vertical tilt table LPP). In sham rats, passive intravesical pressure rises in Crede and vertical tilt table methods induced active urethral closure mechanisms that contributed to high LPPs (41.4 and 35.5 cm H2O, respectively), which were significantly reduced by nerve transection. During sneezing, leakage was observed in nerve-transected rats, but not in sham rats, indicating that sneezing can activate an additional urethral closure mechanism. Measuring LPP during sneezing or passive intravesical pressure rises in the vertical tilt table and Crede method seems to be useful for assessing the continence mechanisms under different stress conditions in rats.

Human pluripotent stem cells (hPSCs) hold significant promise for use in regenerative medicine, or as a model to understand human embryo development. However, the basic mechanisms required for proliferation and self-renewal of hPSCs have not been fully uncovered. Proliferation in all eukaryotes is dependent upon highly regulated expression of the histone H3 variant Centromere protein A (CENP-A). In the current study, we demonstrate that hPSCs have a unique messenger ribonucleic acid (mRNA) reserve of CENP-A not found in somatic fibroblasts. Using short hairpin RNA technology to reduce but not ablate CENP-A, we show that CENP-A-depleted hPSCs are still capable of maintaining a functional centromeric mark, whereas fibroblasts are not. However, upon induction of differentiation or DNA damage, hPSCs with depleted CENP-A arrest in G2/M and undergo apoptosis. Analysis of CENP-A dynamics following DNA damage in hPSCs reveals that 60 min after irradiation, CENP-A is found in multiple small nuclear foci that are mutually exclusive to γH2AX as well as CENP-C. Furthermore, following irradiation, hPSCs with depleted CENP-A mount a normal apoptotic response at 6 h; however at 24 h, apoptosis is significantly increased in CENP-A-depleted hPSCs relative to control. Taken together, our results indicate that hPSCs exhibit a unique mechanism for maintaining genomic integrity by possessing the flexibility to reduce the amount of CENP-A required to maintain a functional centromere under self-renewing conditions, and maintaining a reserve of CENP-A mRNA to rebuild the centromere following differentiation or DNA damage.

Objective To determine if hyperglycosylated hCG (hhCG), produced by invasive trophoblasts, measured as early as 9 days after egg retrieval can predict ongoing pregnancies (OP) after in vitro fertilization and fresh embryo transfer (IVF-ET). Design Cohort Setting Academic ART center Patients Consecutive patients undergoing IVF-ET Interventions Serum hhCG and hCG levels measured 9 (D9) and 16 (D16) days after egg retrieval Outcome Ongoing pregnancy (OP) beyond 9 weeks of gestation Results OP (62 of 112 participants) was associated with higher D9 levels of hhCG and hCG However, hhCG was detectable in all D9 OP samples, while hCG was detectable in only 22%. D9 hhCG levels >110 pg/mL was 96% specific for OP, yielding a positive predictive value of 95%. Compared to D9 hCG levels, hhCG was more sensitive and had a larger area under the curve (0.87 vs. 0.67). Diagnostic test characteristics were similar between D16 hhCG and hCG levels. Conclusions In patients undergoing assisted reproduction, a test to detect pregnancy early and predict outcomes is highly desirable. HhCG is detectable in serum 9 days after egg retrieval IVF-ET cycles. At this early assessment, hhCG is superior to traditional hCG and highly predictive of ongoing pregnancies.

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