RESUMEN
INTRODUCTION: Transumbilical laparoendoscopic single-site (U-LESS) pyeloplasty may provide improved cosmesis compared with conventional laparoscopic pyeloplasty. However, U-LESS pyeloplasty can be challenging because of the need for extensive suturing. The wristed instrumentation of robot-assisted laparoendoscopic single-site (R-LESS) pyeloplasty provides improved dexterity to facilitate intracorporeal suturing. We therefore present our technique and experience with R-LESS pyeloplasty for ureteropelvic junction obstruction (UPJO). SUBJECTS AND METHODS: The da Vinci(®) S or Si Surgical System (Intuitive Surgical, Sunnyvale, CA) was used in all cases. Ureteral stents were placed cystoscopically at the start of each case. A 3-cm skin incision was made adjacent to the umbilicus. Three ports (12 mm, 8 mm, and 5 mm) were placed either through separate fascial incisions or into a GelPort(®) (Applied Medical, Rancho Santa Margarita, CA). Key techniques included port staggering, a "chopstick" arrangement of the instruments, and use of a 30° lens in an upward configuration. Traditional dismembered pyeloplasty procedures were performed in all cases. Patients less than 45 years of age with no prior abdominal surgery were offered this approach. RESULTS: Eight patients (4 female; 5 right-sided; median age, 22 years) underwent R-LESS pyeloplasty without the need for additional ports. All patients were discharged by the third postoperative day, and 5 were discharged on Day 1. One patient experienced urine leakage, which was managed with a temporary nephrostomy tube. Length of follow-up ranged from 29 to 46 months. No patients have developed symptoms or radiographic evidence of recurrent UPJO. CONCLUSIONS: R-LESS pyeloplasty can be safely performed for selected patients with currently available robotic equipment. Careful patient selection and case setup are key to successfully performing these procedures.
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Pelvis Renal/cirugía , Laparoscopía/métodos , Robótica , Obstrucción Ureteral/cirugía , Adolescente , Adulto , Niño , Femenino , Humanos , Masculino , Estudios Retrospectivos , Resultado del Tratamiento , Ombligo , Adulto JovenRESUMEN
The regulation of a pre-replicative complex (pre-RC) at origins ensures that the genome is replicated only once per cell cycle. Cdt1 is an essential component of the pre-RC that is rapidly degraded at G1-S and also inhibited by Geminin (Gem) protein to prevent re-replication. We have previously shown that destruction of the Drosophila homolog of Cdt1, Double-parked (Dup), at G1-S is dependent upon cyclin-E/CDK2 and important to prevent re-replication and cell death. Dup is phosphorylated by cyclin-E/Cdk2, but this direct phosphorylation was not sufficient to explain the rapid destruction of Dup at G1-S. Here, we present evidence that it is DNA replication itself that triggers rapid Dup destruction. We find that a range of defects in DNA replication stabilize Dup protein and that this stabilization is not dependent on ATM/ATR checkpoint kinases. This response to replication stress was cell-type specific, with neuroblast stem cells of the larval brain having the largest increase in Dup protein. Defects at different steps in replication also increased Dup protein during an S-phase-like amplification cell cycle in the ovary, suggesting that Dup stabilization is sensitive to DNA replication and not an indirect consequence of a cell-cycle arrest. Finally, we find that cells with high levels of Dup also have elevated levels of Gem protein. We propose that, in cycling cells, Dup destruction is coupled to DNA replication and that increased levels of Gem balance elevated Dup levels to prevent pre-RC reformation when Dup degradation fails.
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Proteínas de Ciclo Celular/metabolismo , Replicación del ADN/fisiología , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Animales , Ciclo Celular , Proteínas de Ciclo Celular/genética , División Celular/fisiología , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Femenino , Fase G2/fisiología , Componente 6 del Complejo de Mantenimiento de Minicromosoma , Mutación , Folículo Ovárico/citología , Folículo Ovárico/metabolismoRESUMEN
It is important that chromosomes are duplicated only once per cell cycle. Over-replication is prevented by multiple mechanisms that block the reformation of a pre-replicative complex (pre-RC) onto origins in S and G2 phase. We have investigated the developmental regulation of Double-parked (Dup) protein, the Drosophila ortholog of Cdt1, a conserved and essential pre-RC component found in human and other organisms. We find that phosphorylation and degradation of Dup protein at G1/S requires cyclin E/CDK2. The N terminus of Dup, which contains ten potential CDK phosphorylation sites, is necessary and sufficient for Dup degradation during S phase of mitotic cycles and endocycles. Mutation of these ten phosphorylation sites, however, only partially stabilizes the protein, suggesting that multiple mechanisms ensure Dup degradation. This regulation is important because increased Dup protein is sufficient to induce profound rereplication and death of developing cells. Mis-expression has different effects on genomic replication than on developmental amplification from chorion origins. The C terminus alone has no effect on genomic replication, but it is better than full-length protein at stimulating amplification. Mutation of the Dup CDK sites increases genomic re-replication, but is dominant negative for amplification. These two results suggest that phosphorylation regulates Dup activity differently during these developmentally specific types of DNA replication. Moreover, the ability of the CDK site mutant to rapidly inhibit BrdU incorporation suggests that Dup is required for fork elongation during amplification. In the context of findings from human and other cells, our results indicate that stringent regulation of Dup protein is critical to protect genome integrity.