Our objectives were to evaluate the efficacy of ampicillin trihydrate for the treatment of metritis in dairy cows compared with ceftiofur hydrochloride and the subsequent effects on pregnancy at first insemination (P/AI). Cows in the first 12 d in milk (DIM) with a uterine discharge score of 5 (watery, reddish or brownish discharge of foul smell) and rectal temperature <39.5°C were diagnosed with metritis based on the fetid discharge, and cows with metritis and rectal temperature ≥39.5°C were diagnosed as having puerperal metritis. Cows with metritis (n=528) were blocked by parity and type of metritis as fetid discharge or puerperal metritis and, within each block, assigned randomly to receive 11mg/kg of ampicillin (n=259) or 2.2mg/kg of ceftiofur (n=269) once daily for 5 d. Day of diagnosis of metritis was considered study d 1. A cohort of 268 cows without metritis was selected randomly at 12 DIM. Rectal temperature was measured in cows with metritis on study d 1 to 7, and 12, and vaginal discharge was scored on study d 5, 7, and 12. Metritis cure was characterized by vaginal discharge score of <5 or by vaginal discharge score of <5 and no fever. At 32±3 DIM, vaginal discharge was scored for diagnosis of purulent vaginal discharge. At 39±3 DIM, endometrial cytology was performed. At 53±3 and 67±3 DIM, ovaries were scanned to determine estrous cyclicity. Pregnancy was evaluated after the first AI. Cure of metritis based on vaginal discharge <5 was greater for ampicillin than ceftiofur on d 5 (37.1 vs. 25.2%) and 7 (57.2 vs. 46.3%), but not on d 12 (82.0 vs. 85.0%). Cure of metritis based on vaginal discharge <5 and no fever was greater for ampicillin than for ceftiofur only on d 7 (50.4 vs. 37.9%), but not on d 5 (23.1 vs. 17.6%) and 12 (66.1 vs. 67.4%). Cows with puerperal metritis had reduced cure compared with cows with fetid discharge on d 5 (30.5 vs. 12.8%), 7 (55.2 vs. 33.6%), and 12 (72.0 vs. 61.1%). The proportion of cows with fever on any day after therapy started did not differ between treatments. Fifty-three percent of cows with metritis based on fetid discharge developed fever after initiating antimicrobial therapy. Cows receiving ampicillin had less prevalence of purulent vaginal discharge than those treated with ceftiofur (57.7 vs. 67.8%), but they were both greater than cows without metritis (21.9%). Prevalence of cytological endometritis did not differ between ampicillin and ceftiofur (30.0 vs. 25.4%), but they were both greater than cows without metritis (14.5%). The proportion of estrous cyclic cows (75.0%) and P/AI did not differ among treatments (ampicillin=28.0% vs. ceftiofur=28.3% vs. without metritis=30.5%). Clinical cure was faster for ampicillin than for ceftiofur, but on study d 12 both treatments resulted in similar cure. Clinical cure was less for cows with puerperal metritis than for cows with fetid uterine discharge. Despite differences in uterine health, P/AI at the first insemination did not differ among treatments.
The kidney androgen-regulated protein (KAP) is specifically expressed and differentially regulated by androgens and tri-iodothyronine (T3) in intact mouse early (PCT) and late (PR) proximal-tubule cells. Until now, detailed characterization of the molecular elements mediating androgen-responsive gene expression in the kidney has been hampered by the lack of appropriate cultured cell systems suitable for DNA transfection studies. In the present study we have analysed the hormone-dependent transactivation of the KAP gene promoter in immortalized differentiated PCT and PR proximal-tubule cells derived from L-PK/Tag1 transgenic mice. Transient transfection studies with different KAP promoter constructs indicated that a 224bp-truncated fragment was sufficient to mediate cell-specific expression of the KAP promoter. Dihydrotestosterone (DHT) stimulated in an androgen-dependent manner the transactivation of KAP in PCT and PR cells, while mutation of a putative androgen-response element (ARE) sequence located at −39bp from the transcription initiation site abolished the transactivation induced by DHT. Furthermore, insulin-like growth factor 1 (IGF-1), but not T3, enhanced the androgen-dependent transactivation of KAP in cultured PCT cells. These results demonstrate that the short 224bp fragment of the KAP promoter is sufficient to drive the proximal-tubule androgen-specific regulated expression of KAP and reveal synergistic interactions between IGF-1 and androgens for KAP regulation in PCT cells.
The objectives of the current study were to evaluate the effects of supplemental progesterone after artificial insemination (AI) on expression of IFN-stimulated genes (ISG) in blood leukocytes and fertility in lactating dairy cows. Weekly cohorts of Holstein cows were blocked by parity (575 primiparous and 923 multiparous) and method of insemination (timed AI or AI on estrus) and allocated randomly within each block to untreated controls, a controlled internal drug release (CIDR) containing 1.38g of progesterone from d 4 to 18 after AI (CIDR4), or a CIDR on d 4 and another on d 7 after AI and both removed on d 18 (CIDR4+7). Blood was sampled to quantify progesterone concentrations in plasma and mRNA expression in leukocytes for the ubiquitin-like IFN-stimulated gene 15-kDa protein (ISG15) and receptor transporter protein-4 (RTP4) genes. Pregnancy was diagnosed on d 34±3 and 62±3 after AI. Treatment increased progesterone concentrations between d 5 and 18 after AI in a dose-dependent manner (control=3.42, CIDR4=4.97, and CIDR4+7=5.46ng/mL). Cows supplemented with progesterone tended to have increased luteolysis by d 19 after AI (control=17.2; CIDR4=29.1; CIDR4+7=30.2%), which resulted in a shorter AI interval for those reinseminated after study d 18. Pregnancy upregulated expression of ISG in leukocytes on d 19 of gestation, but supplementing progesterone did not increase mRNA abundance for ISG15 and RTP4 on d 16 after insemination and tended to reduce mRNA expression on d 19 after AI. For RTP4 on d 19, the negative effect of supplemental progesterone was observed only in the nonpregnant cows. No overall effect of treatment was observed on pregnancy per AI on d 62 after insemination and averaged 28.6, 32.7, and 29.5% for control, CIDR4, and CIDR4+7, respectively. Interestingly, an interaction between level of supplemental progesterone and method of AI was observed for pregnancy per AI. For cows receiving exogenous progesterone, the lower supplementation with CIDR4 increased pregnancy per AI on d 62 in cows inseminated following timed AI (CIDR4=39.2; CIDR4+7=27.5%); in those inseminated following detection of estrus, however, the use of a second insert on d 7 resulted in greater pregnancy per AI (CIDR4=26.9; CIDR4+7=31.5%). Pregnancy loss did not differ among treatments. Supplemental progesterone post-AI using a single intravaginal insert on d 4 was beneficial to pregnancy in cows inseminated following timed AI, but incremental progesterone with a second insert on d 7 did not improve fertility of dairy cows.
Dihydrotestosterone (DHT) binding studies and the effects of DHT on the expression of beta-glucuronidase (Gus) and kidney androgen-regulated protein (KAP) genes and cell growth were investigated in immortalized early PKSV-PCT and late PKSV-PR proximal tubule cells, derived from transgenic mice carrying the L-pyruvate kinase/SV40 hybrid gene. [3H]DHT binding studies indicated that both cell lines have conserved substantial amounts of androgen receptors. The levels of KAP and Gus transcripts in PKSV-PCT cells, and those of KAP transcripts in PKSV-PR cells, decreased when cells were shifted from a serum-supplemented to a steroid-free medium. The addition of 30 nM DHT to the steroid-free medium resulted in a slight increase in Gus and in a more marked increase in KAP transcripts in both cell lines. Dihydrotestosterone also affected the growth of PKSV-PCT and PKSV-PR cells, since this hormone added to the steroid-free medium stimulated the incorporation of [3H]thymidine in a dose-dependent manner and induced the formation of domes, which represent indicators of ionic transport processes. Thus, because these early and late mouse proximal tubule cells have conserved androgen receptors, they represent attractive cell systems to analyze the action of androgens on specific functions of the mouse proximal tubule.
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