Effect of 2 corpora lutea on blood perfusion, peripheral progesterone, and hepatic steroid-inactivating enzymes in dairy cattle.

The luteal structure that develops postovulation is critical to the facilitation and maintenance of pregnancy in dairy cattle. The objectives of this experiment were to determine if the induction of an accessory corpus luteum (CL), via human chorionic gonadotropin, altered blood perfusion of CL, peripheral concentrations of progesterone, or hepatic steroid-inactivating enzymes. Twenty-eight late-lactation Holstein cows were synchronized using the Ovsynch protocol and randomly assigned to 1 of 2 treatment groups. Cows received either an injection of human chorionic gonadotropin (1,000IU, i.m.) to induce an accessory CL (cows had exactly 2CL in 1 ovary) or no treatment (cows had exactly 1CL). Corpora lutea were examined daily from d 10 to 18 (d 0 was induced ovulation) via Doppler ultrasonography and a blood sample was collected. Volume of the CL was recorded, as well as images and videos of each CL, which were analyzed for blood perfusion. On d 13, a liver biopsy was performed to analyze hepatic steroid-inactivating enzymes. Cows with 1 or 2CL had similar peripheral concentrations of progesterone. Cows with 2CL had similar luteal volumes to cows with 1CL but cows with 2CL had greater total luteal blood perfusion. Hepatic enzyme [cytochrome P450 (CYP) 1A, 3A, and 2C, aldo-keto reductase 1C, and uridine diphosphate glucuronosyltransferase] activities did not differ between cows with 1 and 2CL. Overall, the observed increase in total luteal blood perfusion in cows with 2CL did not correspond to differences in peripheral concentrations of progesterone or clearance of progesterone measured by the hepatic enzyme activity. This could indicate that induction of an accessory CL would not affect concentrations of progesterone necessary to maintain pregnancy.

Dependence of ß3-adrenergic signaling on the adipokine leptin in cardiac myocytes.

ß3-Adrenergic receptors (ß3ARs) negatively regulate ß-adrenergic signaling via nitric oxide and are dependent on the adipokine leptin for normal expression in adipocytes, thus making ß3AR an attractive candidate for cross-talk with leptin in the heart. Accordingly, we tested the hypothesis that cardiac ß3AR expression and function are dependent on leptin and are severely diminished in leptin-deficient ob/ob mice. Using isolated cardiac myocyte physiology studies, we found that ß3AR function was significantly diminished in ob/ob myocytes and in wild-type myocytes treated with leptin antagonist. This finding was supported by quantitative PCR demonstrating markedly decreased ß3AR mRNA levels in ob/ob mice. Both ß3AR mRNA and function were restored in ob/ob mice after in vivo leptin repletion. We propose that diminished ß3AR signaling may be the critical element to explain the direct effects of leptin on the myocardium and suggest that this work reveals a key feature in the role of leptin in obesity-related cardiac hypertrophy and heart failure.

In utero gene transfer into the pulmonary epithelium.

In early gestation the internal surface of the lung is structurally simple and an ideal target for somatic gene transfer. The transfer of genes into the growing lung would be particularly useful in the prenatal correction of cystic fibrosis, which has devastating pulmonary complications. In addition, in utero gene therapy has the potential to immunotolerize the individual, and thereby to avoid the immune reactions now seen with the current generation of adenoviral vectors. We injected a replication-defective adenoviral vector containing the lacZ reporter gene (Ad5.CMVlacZ) into the amniotic fluid of rat pups on the 16th day of gestation. At 16 days of gestation, rat lungs are equivalent in maturity to those of a 22-week human fetus as their airways are lined with undifferentiated multipotential stem cells. The pups showed high-level reporter gene expression in their airways a week following birth (13 days following infection). The expression was maintained during a time when the lung volume increased approximately 20-fold, alveolarization occurred, and the epithelial cells differentiated. These data establish gene targeting of undifferentiated fetal cells as an effective means of gene therapy.

Left-handed DNA in vivo.

Left-handed DNA is shown to exist and elicit a biological response in Escherichia coli. A plasmid encoding the gene for a temperature-sensitive Eco RI methylase (MEco RI) was cotransformed with different plasmids containing inserts that had varying capacities to form left-handed helices or cruciforms with a target Eco RI site in the center or at the ends of the inserts. Inhibition of methylation in vivo was found for the stable inserts with the longest left-handed (presumably Z) helices. In vitro methylation with the purified MEco RI agreed with the results in vivo. Supercoil-induced changes in the structure of the primary helix in vitro provided confirmation that left-handed helices were responsible for this behavior. The presence in vivo of left-handed inserts elicits specific deletions and plasmid incompatibilities in certain instances.

Detection of anovulation by heatmount detectors and transrectal ultrasonography before treatment with progesterone in a timed insemination protocol.

Our objective was to determine the accuracy of identifying noncycling lactating dairy cows before the application of a timed artificial insemination (AI) protocol [with or without progesterone supplementation via a controlled internal drug-release (CIDR) insert and 2 different timings of AI] by using heatmount detectors and a single ovarian ultrasound examination. At 6 locations in the Midwest, 1,072 cows were enrolled in a Presynch protocol (2 injections of PGF(2alpha) 14 d apart), with the second injection administered 14 d before initiating the Ovsynch protocol (injection of GnRH 7 d before and 48 h after PGF(2alpha) injection, with timed AI at 0 or 24 h after the second GnRH injection). Heatmount detectors were applied to cows just before the first Presynch injection, assessed 14 d later at the second Presynch injection (replaced when activated or missing), and reassessed at initiation of the Ovsynch protocol. Ovaries were examined for the presence of a corpus luteum (CL) by ultrasound before the initiation of treatment. Treatments were assigned to cows based on the presence or absence of a CL detected by ultrasound: 1) no CL + no CIDR; 2) no CL + CIDR insert for 7 d; and 3) CL present. Further, alternate cows within the 3 treatments were assigned to be inseminated concurrent with the second GnRH injection of Ovsynch (0 h) or 24 h later. Pregnancy was diagnosed at 33 and 61 d after the second GnRH injection. By using low (<1 ng/mL) concentrations of progesterone in serum as the standard for noncycling status, heatmount detectors were activated on a large percentage of noncycling cows (>60%), whereas the single ultrasound examination incorrectly classified noncycling cows only 21% of the time. Conversely, cycling cows (progesterone > or =1 ng/mL) were correctly identified 70 to 78% of the time by heatmount detectors, but 85 to 92% were correctly identified by ultrasound. Overall accuracy of heatmount detectors and ultrasound was 71 and 84%, respectively. Application of progesterone to cows without a CL at the time of the first injection of GnRH reduced the incidence of ovulation but increased the proportions of pregnancies per AI at d 33 or 61 compared with nontreated cows without a CL at the onset of the Ovsynch protocol. Percentages of cows pregnant and pregnancy survival did not differ for cows having a CL before treatment compared with those not having a CL and treated with progesterone. Compared with no response, when a follicle ovulated in response to the first GnRH injection, percentage of cows becoming pregnant after the timed AI increased from 33.3 to 41.6%. Timing of AI at 0 or 24 h after the second GnRH injection did not alter pregnancies per AI, but cows having luteal activity before treatment had improved pregnancies per AI compared with noncycling cows. We conclude that identifying noncycling cows by ultrasound was more accurate than by heatmount detectors. Subsequent progesterone treatment of previously cycling cows not having a CL at the onset of Ovsynch increased the proportion of pregnant cows, equal to that of cows having a CL but not treated with progesterone.

The effect of follicular wave on fertility characteristics in beef cattle.

The objectives of this experiment were to determine the effects of follicular wave (first or second) on diameter of the dominant follicle, concentrations of progesterone and estradiol and the hepatic enzymes that inactivate them, thickness of the endometrium, and pregnancy rates to AI. Beef heifers ( = 101) and cows ( = 106) were randomly assigned to 1 of 2 treatments: insemination to the first follicular wave (FFW) or insemination to the second follicular wave (SFW). Estrous cycles of females were synchronized to ensure appropriate timing for the treatments. The MIXED procedure of SAS was used for analysis. A similar proportion of females in each treatment responded to presynchronization; however, females in the FFW group ovulated in response to the first injection of GnRH of the CO-Synch protocol more frequently. Only females ( = 94) that properly responded to ovulation synchronization were included in further analyses. Cows in the FFW group tended ( 0.06) to have larger ovulatory follicles 36 h post-PGF of the CO-Synch protocol compared to cows in the SFW group (14.22 ± 0.42 and 11.83 ± 0.49, respectively), whereas heifers were similar between treatment groups. Three d prior to AI, circulating concentrations of progesterone were lesser ( 0.01) in females in the FFW (3.63 ± 0.80 ng/mL) than in the SFW (7.12 ± 0.83 ng/mL), whereas concentrations of estradiol tended ( 0.08) to be greater in those in the FFW (82.72 ± 6.48 pg/mL) than in the SFW (65.55 ± 6.74 pg/mL). Concentrations of cytochrome P450 1A in the liver were lesser ( 0.01) in females in the FFW than those in the SFW (0.68 ± 0.08 vs. 0.96 ± 0.06, respectively). Endometrial thicknesses were similar between treatments but were thicker ( < 0.0001) in cows (9.73 ± 0.24 mm) than heifers (7.22 ± 0.26 mm). When considering all females or only those that were properly presynchronized, pregnancy rates were similar between treatments. However, when evaluating females that ovulated to the assigned follicular wave, there was a treatment by parity interaction ( = 0.04) with heifers in the FFW having a lesser pregnancy rate (25.9%) than heifers in the SFW (72.0%) while cows in both treatment groups were intermediate (45.4% in FFW and 50.0% in SFW). The differences in concentrations of steroids between treatment groups may affect fertility of heifers; however, additional research is necessary.

Evaluating blood perfusion of the corpus luteum in beef cows during fescue toxicosis.

The aim of this study was to determine if fescue toxicosis altered blood perfusion in the corpus luteum (CL) and peripheral concentrations of progesterone in cattle. The estrous cycles of 36 nonpregnant Angus or Charolais cows were synchronized in 2 replicates using the CO-Synch+CIDR protocol. Seven days after initiation of the protocol, cows were assigned (d 0) to 1 of 2 dietary treatments: 2.5 kg of 1) Kentucky-31 endophyte-infected (KY31; = 14) or 2) MaxQ novel endophyte (MaxQ; = 12) tall fescue seed. On d 7, ovaries were examined using ultrasonography, and only cows that had 1 CL present remained on the study ( = 26). Images of blood perfusion of CL, blood samples, rectal temperatures, and blood pressure of tails were collected on d 10, 13, 15, and 18. Images of CL blood perfusion were analyzed using ImageJ software for pixel density, and scored visually (0 to 9 with 0 = no perfusion, 9 = complete perfusion) by 2 independent technicians. The MIXED procedure of SAS was used with day as a repeated measure. Least squares means and SEM are reported. Cows receiving KY31 had greater rectal temperatures ( 0.003; 38.76 ± 0.08°C) than those receiving MaxQ (38.44 ± 0.08°C), providing evidence that the cows treated with KY31 were influenced by fescue toxicosis. Pulse pressure and mean arterial pressure were decreased ( < 0.01) in cows receiving KY31 (55.26 ± 2.81 and 80.06 ± 2.72 mmHg, respectively) than MaxQ (66.58 ± 3.03 and 91.38 ± 2.93 mmHg, respectively). Concentrations of progesterone were similar ( = 0.54) between cows receiving KY31 (6.04 ± 0.53 ng/mL) or MaxQ (6.36 ± 0.63 ng/mL). Pixel densities ( = 0.14) and visual perfusion scores were similar ( = 0.11) between cows receiving KY31 (1477.20 ± 655.62 pixels and 2.23 ± 0.34, respectively) or MaxQ (2934.70 ± 718.20 pixels and 3.00 ± 0.36, respectively). Mean CL volume was similar ( 0.95) between treatments. In conclusion, blood perfusion of CL or peripheral concentrations of progesterone were not altered at the onset of fescue toxicosis in this short-term study, indicating that a decrease in blood perfusion of the CL may not be a primary mechanism involved in decreased reproductive efficiency of cattle during fescue toxicosis.

Using estrus detection patches to optimally time insemination improved pregnancy risk in suckled beef cows enrolled in a fixed-time artificial insemination program.

A multilocation study examined pregnancy risk (PR) after delaying AI in suckled beef cows from 60 to 75 h when estrus had not been detected by 60 h in response to a 7-d CO-Synch + progesterone insert (CIDR) timed AI (TAI) program (d -7: CIDR insert concurrent with an injection of GnRH; d 0: PGF injection and removal of CIDR insert; and GnRH injection at TAI [60 or 75 h after CIDR removal]). A total of 1,611 suckled beef cows at 15 locations in 9 states (CO, IL, KS, MN, MS, MT, ND, SD, and VA) were enrolled. Before applying the fixed-time AI program, BCS was assessed, and blood samples were collected. Estrus was defined to have occurred when an estrus detection patch was >50% colored (activated). Pregnancy was determined 35 d after AI via transrectal ultrasound. Cows ( = 746) detected in estrus by 60 h (46.3%) after CIDR removal were inseminated and treated with GnRH at AI (Control). Remaining nonestrous cows were allocated within location to 3 treatments on the basis of parity and days postpartum: 1) GnRH injection and AI at 60 h (early-early = EE; = 292), 2) GnRH injection at 60 h and AI at 75 h (early-delayed = ED; = 282), or 3) GnRH injection and AI at 75 h (delayed-delayed = DD; = 291). Control cows had a greater ( < 0.01) PR (64.2%) than other treatments (EE = 41.7%, ED = 52.8%, DD = 50.0%). Use of estrus detection patches to delay AI in cows not in estrus by 60 h after CIDR insert removal (ED and DD treatments) increased ( < 0.05) PR to TAI when compared with cows in the EE treatment. More ( < 0.001) cows that showed estrus by 60 h conceived to AI at 60 h than those not showing estrus (64.2% vs. 48.1%). Approximately half (49.2%) of the cows not in estrus by 60 h had activated patches by 75 h, resulting in a greater ( < 0.05) PR than their nonestrous herd mates in the EE (46.1% vs. 34.5%), ED (64.2% vs. 39.2%), and DD (64.8% vs. 31.5%) treatments, respectively. Overall, cows showing estrus by 75 h (72.7%) had greater ( < 0.001) PR to AI (61.3% vs. 37.9%) than cows not showing estrus. Use of estrus detection patches to allow for a delayed AI in cows not in estrus by 60 h after removal of the CIDR insert improved PR to TAI by optimizing the timing of the AI in those cows.

Tetracycline promoter mutations decrease non-B DNA structural transitions, negative linking differences and deletions in recombinant plasmids in Escherichia coli.

The ability to clone a variety of sequences with varying capabilities of adopting non-B structures (left-handed Z-DNA, cruciforms or triplexes) into three loci of pBR322 was investigated. In general, the inserts were stable (non-deleted) in the EcoRI site (an untranslated region) of pBR322. However, sequences most likely to adopt left-handed Z-DNA or triplexes in vivo suffered deletions when cloned into the BamHI site, which is located in the tetracycline resistance structural gene (tet). Conversely, when the promoter for the tet gene was altered by filling-in the unique HindIII or ClaI sites, the inserts in the BamHI site were not deleted. Concomitantly, the negative linking differences of the plasmids were reduced. Also, inserts with a high potential to adopt Z-DNA conformations were substantially deleted in the PvuII site of pBR322 (near the replication origin and the copy number control region), but were less deleted if the tet promoter was insertion-mutated. The deletion phenomena are due to the capacity of these sequences to adopt left-handed Z-DNA or triplexes in vivo since shorter inserts, less prone to form non-B DNA structures, or random sequences, did not exhibit this behavior. Sequences with the potential to adopt cruciforms were stable in all sites under all conditions. These results reveal a complex interrelationship between insert deletions (apparently the result of genetic recombination), negative supercoiling, and the formation of non-B DNA structures in living Escherichia coli cells.

Progesterone status, parity, body condition, and days postpartum before estrus or ovulation synchronization in suckled beef cattle influence artificial insemination pregnancy outcomes.

Our objective was to assess the effects of progesterone before initiating an estrus- or ovulation-synchronization program in addition to the influence of parity, BCS, and days postpartum on resulting pregnancy rates per AI. Experimental data were combined from 73 herd-year studies consisting of more than 8,500 suckled beef cows exposed to variants of the CO-Synch program. Blood was harvested from samples collected at 10 and 0 d before the onset of CO-Synch, and progesterone concentrations of the samples were determined. The progesterone environment preceding synchronization was assessed in 3 ways on the basis of progesterone concentrations measured in the 2 defined blood samples. All binomial logistic regression models used procedure GLIMMIX in SAS and included the fixed effects of program duration, inclusion of progesterone via an intravaginal insert, parity, days postpartum at AI, BCS, and appropriate interactions. In addition, model 1 included 3 categories of progesterone concentrations (low [<1 ng/mL], medium [1.00 to 3.99 ng/mL], and high [≥4.00 ng/mL] concentrations) at 10 and 0 d before synchronization and their interaction. Model 2 included 4 categories defining the stage of the estrous cycle (late diestrus, early diestrus, and proestrus-estrus-metestrus) or anestrus, at which cows started the synchronization program. Model 3 defined cows as cycling or noncycling at the onset of the program. Significant effects of progesterone supplementation, which hormone was used to initiate the timed AI program, parity, BCS, days postpartum, and progesterone status assessed in 3 ways were consistent in nearly all models. Progesterone status at the onset of synchronization was not important to pregnancy outcomes in multiparous cows, whereas pregnancy rate per AI was suppressed in primiparous cows that began in a low-progesterone environment (proestrus, estrus, metestrus, or anestrus). A significant 3-way interaction of parity, BCS, and days postpartum in 2 models reinforced the importance of these factors to AI pregnancy outcomes. Ancillary analyses identified the significant effects of cycling status and BCS as well as days postpartum on luteolytic response to PGF(2α). Pregnancy loss of 2.7% to 4.2% was detected to occur between a positive pregnancy diagnosis at 35 d post-AI and later stages of pregnancy. We concluded that progesterone status at the onset of the synchronization program is critical to pregnancy outcomes in primiparous but not multiparous cows.

Effects of administration of prostaglandin F at initiation of the seven-day CO-Synch+controlled internal drug release ovulation synchronization protocol for suckled beef cows and replacement beef heifers.

Two experiments were conducted to determine the effect of administering PGF at the initiation of the 7-d CO-Synch+controlled internal drug release (CIDR) fixed-timed AI (TAI) protocol on pregnancy rates of suckled beef cows and replacement heifers. Within location, cows were stratified by days postpartum (DPP), BCS, and parity (Exp. 1; = 1,551) and heifers were stratified by BCS (Exp. 2; = 999) and randomly assigned to 1 of 2 treatments: 1) CO-Synch+CIDR (100-µg injection of GnRH at CIDR insertion [d -10] with a 25-mg injection of PGF at CIDR removal [d -3] followed by injection of GnRH and TAI on d 0) or 2) PG-CO-Synch+CIDR (a 25-mg injection of PGF on d -10 of the CO-Synch+CIDR protocol). Follicle diameter and corpus luteum (CL) development were assessed on d -10 and -3, and pregnancy status was determined on d 30 to 35. Blood was collected on d -20, -10, -3, and 0 relative to TAI to determine concentrations of progesterone (P4). In Exp. 1, TAI pregnancy rates did not differ ( = 0.667) between treatments and were affected by BCS ( = 0.003) and DPP ( = 0.006). Concentrations of P4 were greater ( < 0.0001) on d -3 for CO-Synch+CIDR than for PG-CO-Synch+CIDR (4.1 ± 0.2 and 3.4 ± 0.2 ng/mL, respectively). Follicle diameter on d -3 differed ( = 0.05) between PG-CO-Synch+CIDR (13.4 ± 0.3 mm) and CO-Synch+CIDR (12.5 ± 0.3 mm) treatments. Cows with P4 > 2.5 ng/mL on d -10 had greater ( = 0.024) pregnancy rate to TAI (56.5%) compared with cows with 2.5 ng/mL < P4 > 1 (43.0%), whereas cows with P4 < 1 ng/mL were intermediate (51.6%). Cows with a CL on d -10 had greater ( = 0.012) pregnancy rates to TAI than cows without a CL (66.3 vs. 39.4%, respectively). In Exp. 2, TAI pregnancy rates did not differ ( = 0.316) between treatments. Concentrations of P4 differed ( < 0.0001) on d -3 with greater concentrations of P4 for CO-Synch+CIDR than for PG-CO-Synch+CIDR (3.75 ± 0.20 ng/mL and 3.60 ± 0.21 ng/mL, respectively). Follicle diameter was similar ( = 0.749) between treatments on d -10 and -3. Regardless of treatment, cyclic status tended ( = 0.062) to improve pregnancy rates to TAI (55 vs. 45%, for cycling and noncycling heifers, respectively). We concluded that addition of PGF to the 7-d CO-Synch+CIDR protocol decreased concentrations of P4 in cows and heifers and increased follicle diameter at CIDR removal in cows but failed to increase TAI pregnancy rates.

Analysis of prognostic factors and survival in patients with ovarian cancer treated with second-line hexamethylmelamine (altretamine).

Hexamethylmelamine (altretamine, HMM) 260 mg/m2/day p.o. for 14 days followed by a 14-day drug-free interval was administered to 52 outpatients with advanced ovarian cancer who had previously been treated with chemotherapy. Prior to HMM, 92% (48/52) of these patients had received cisplatin and cyclophosphamide with or without doxorubicin. Two more patients received other cisplatin-based regimens. At the completion of HMM therapy, 15% (8/52) displayed no evidence of disease (NED). Of these eight patients, five are still alive 32 to 82 months after altretamine therapy (median follow-up of 46 months). At 41 months, one patient died of intercurrent illness with no clinical evidence of recurrence; the other two patients died of their disease at 21 and 31 months following HMM therapy. The median survival of the total group was 11 months: nine months for patients who did not respond to altretamine and 46+ months for patients with NED after altretamine (p less than 0.05). Intermittent oral administration of single-agent altretamine was well tolerated: eight patients reported moderate gastrointestinal symptoms, and only one patient reported severe gastrointestinal symptoms. Moderate neurologic toxicity was reported by five patients. No WBC fell below 2000 mm3 and platelet counts fell below 100,000 mm3 in only three patients; no patient experienced severe hematologic toxicity. In this series of patients, the overall response (15%) was comparable to or better than those reported for more toxic chemotherapeutic regimens. On the basis of these data and those reported by other investigators, HMM warrants consideration as a reasonable option in the management of recurrent or persistent ovarian cancer.

Cytosine methylation as an effector of right-handed to left-handed DNA structural transitions.

Cytosine methylation has energetic and structural influences on left-handed Z-DNA formation in supercoiled plasmids. The restriction and modification enzymes from Haemophilus haemolyticus (HhaI and M.HhaI) provide a system to locate and analyze small segments of Z-DNA in large supercoiled plasmids. An approach is outlined that uses M.HhaI as an in vivo conformational probe for the detection of unusual DNA structures in a living cell. Also, characteristic features of the M.HhaI gene and protein are discussed.

In vivo existence of left-handed DNA.

A genetic-biochemical assay has been developed to investigate the in vivo existence and consequences of unusual DNA structures. Left-handed DNA was shown to exist in living Escherichia coli. The EcoRI methyltransferase gene (temperature-sensitive) was cloned to serve as a probe for perturbed GAATTC sites in vivo. This plasmid was cotransformed with different plasmids containing inserts that had varying capacities to form left-handed helices or cruciforms with a target EcoRI site in the center or at the ends of the inserts. Inhibition of methylation in vivo was found for the stable inserts with the longest left-handed helices. In vitro methylation with the purified M.EcoRI enzyme agreed with the in vivo results.

Combined prostaglandin therapy and ductal formalin infiltration in neonatal pulmonary oligemia.

Prostaglandins and ductal formalin infiltration, singly and together, have been used in efforts to improve pulmonary flow in very ill newborn infants with right ventricular outflow tract obstruction. To evaluate the efficacy of concurrent use of prostaglandins and ductal formalin infiltration, we have reviewed our experience with 25 infants with right ventricular outflow tract obstruction and prostaglandin-ductal formalin infiltration therapy. Prostaglandin therapy was begun 22 +/- 21 hours (range 20 to 93 hours) before and was continued 20 +/- 18 hours (range 0 to 62 hours) following ductal formalin infiltration; prostaglandin administration was initiated at a dose of 0.05 to 0.1 microgram/kg/min and tapered postoperatively. Clinical cyanosis was diminished in 20 of 25 infants (80%) postoperatively. Systemic arterial pH and oxygen saturation both improved following prostaglandin-ductal formalin infiltration therapy from 7.35 to 7.41 (p less than 0.001) and from 35.7 to 50.3 (p less than 0.001), respectively. Persistent ductal patency (mean 219 +/- 191 days) was observed in 17 survivors of the early postoperative period (more than 14 days). Two of five infants who died within 14 days of operation had a widely patent ductus with resultant progressive congestive heart failure. The other three infants died as a result of operative technical problems, dysrhythmias, and thrombotic ductal closure, respectively. No correlation was observed between duration of ductus patency and operatively determined size of ductus, total prostaglandin dose, or duration of prostaglandin infusion. Secondary operative intervention was delayed by 92 +/- 74 days with prostaglandin-ductal formalin infiltration therapy; thus prostaglandin-ductal formalin infiltration therapy may have a role in selected neonates with right ventricular outflow tract obstruction.

Recurrent Paget disease of the vulva in a split-thickness graft.

Recurrent Paget disease developed in a vulvar split-thickness skin graft, the second reported case of recurrence of this type and the first without an underlying dermal adnexal adenocarcinoma.

Primary invasive carcinoma of the vagina.

A retrospective study of 29 patients with invasive carcinoma of the vagina was completed at The Milton S. Hershey Medical Center, Pennsylvania State University, for a ten-year period from 1976-1986. The overall incidence was 1.3% of all gynecologic malignancies. Twenty-four patients (83%) had squamous cell carcinoma and five (17%) had adenocarcinoma. Squamous cell carcinoma was most commonly located in the upper anterior and lateral vaginal vaults, whereas adenocarcinoma was found more often in the lower anterior and lateral vaginal vaults. The majority of the patients (96%) were managed by a combination of whole-pelvis irradiation and brachytherapy. Twenty-four percent of the patients had a recurrence in the vagina only, indicating the need for better local control. The overall survival rate was 48%. Patients with previous hysterectomy were more likely to develop serious treatment-related complications.

Bypassing femoral cortical defects with cemented intramedullary stems.

The objective of this investigation was to examine the effect of cemented intramedullary stem bypass on bone torsional property in the presence of femoral cortical defect. We intended to test two hypotheses; first, intramedullary fixation without bypass will accentuate the stress concentration effect and second, there will be an optimal length of stem bypass beyond the defect. Cemented intramedullary stems were used to bypass 50% diaphyseal diameter unicortical defects in paired, fresh-frozen canine femora. One member of each pair served as an unaltered control and all specimens were tested to failure in torsion. Both single-tailed paired t-test and analysis of variance were used for data analysis. Bones subjected to the cortical defect and no bypass were substantially weakened, exhibiting only 44 +/- 8% of control side maximum torque (p less than 0.001). Positioning of the intramedullary stem with its tip at the center of the defect provided a small degree of strength improvement, achieving 60 +/- 7% of control side maximum torque. One and two diaphyseal diameter bypass, however, significantly (p less than 0.01) improved bone torsional strength, resulting in 80 +/- 6% and 84 +/- 13% of control side maximum torque, respectively. Three diameter bypass achieved only 68 +/- 8% of control side maximum torque. This significant (p less than 0.05) decline in strength when compared to two diameter bypass appears to indicate that the length of stem bypass beyond the cortical defect does have an optimum, probably due to the geometric characteristics of the canine femora.

Left-handed Z-DNA helices in polymers, restriction fragments, and recombinant plasmids.

Studies on DNA polymers, restriction fragments, and recombinant plasmids have revealed the following: A) A family of left-handed DNA conformations exists for (dC-dG)n.(dC-dG)n. The observation of a particular conformation is dependent on the salt, the salt concentration and dehydrating agent. B) In sodium acetate solutions, (dC-dG)n.(dC-dG)n forms left-handed, psi(+)-condensed structures as detected by Raman spectroscopy and circular dichroism. C) (dT-dG)n.(dC-dA)n undergoes a right-to-left-handed transition only when reacted with AAF and at high salt concentrations. D) Transitions observed for polymer DNAs also are observed for restriction fragments containing both (dC-dG).(dC-dG) and (dT-dG).(dC-dA) sequences, but the transitions in the fragments generally require higher salt concentrations than observed for the polymers. E) Studies with recombinant plasmids containing (dC-dG) sequences from 10 to 58 bp in length demonstrate that left-handed Z-DNA segments can exist contiguous to B-DNA segments. F) Negative supercoil density (sigma less than or equal to -0.072) is sufficient to convert the (dC-dG) regions in those plasmids into left-handed structures under physiological ionic conditions (200 mM NaCl). G) The favorable free energy contribution of methylation in stabilizing the Z form in fragments and plasmids is approximately offset by the unfavorable free energy contributions of the B/Z junctions. H) Sl and BAL 31 nucleases recognize aberrant structural features at the confluence of the B and Z regions. I) Detailed mapping of Sl nuclease cleavage on supercoiled plasmids shows that the nuclease sensitive regions extend over at least five to ten bp. J) Even though the (dT-dG)n.(dC-dA)n polymer requires base modification and high salt conditions to undergo the R----L transition, supercoiling (sigma less than or equal to -0.07) can supply enough energy to allow a plasmid containing the intervening sequence of a human fetal globin gene with (dT-dG).(dC-dA) sequences to undergo a R----L transition.