Robotic surgeries in pediatric patients: an early experience in a single center.

Purpose: Robotic surgery (RS) has the advantages of 3-dimensional view, optical magnification, motional scaling, and improved ergonomics and degree of freedom. Although RS has widely been performed on pediatric patients lately, there are still numerous restrictions and ambiguous indications. The purpose of this study was to report our early experience with RS on pediatric patients at a single center. Methods: Electronic medical records of patients who underwent RS with the da Vinci Xi surgical platform (Intuitive Surgical, Inc.) in Seoul National University Children Hospital from November 2019 to August 2021 were reviewed retrospectively. The median follow-up was 21.0 months (range, 12.3-31.8 months). An online survey was conducted to investigate satisfaction with robotic surgical scars. Results: Fifty-four patients underwent robotic surgeries (median age at operation, 11.1 years [range, 0.1-17.8 years]). In our hospital, patients had 20 different kinds of robotic surgeries, including choledochal cyst excision with hepaticojejunostomy, ovarian mass excision, and others. Median operation time and console time were 157.5 minutes (range, 45-505 minutes) and 40 minutes (range, 11-360 minutes), respectively. All cases were done without conversion into open or laparoscopic methods. Postoperative complications were found in 5 patients. According to an online survey, over half of patients (60.9%) answered that they felt satisfied with scars. Conclusion: Our early experience demonstrated the safety and feasibility of RS in children with a range of diagnoses and complicated procedures. With more experience, RS could be an alternative to traditional open or laparoscopic operations in pediatric patients. Further studies are needed to clarify indications of pediatric RS.

In vitro synthesis and biochemical characterization of acyl-homoserine lactone synthase and its deletion mutant.

Quorum sensing can induce density-dependent gene expressions that cause various problems. For quorum-sensing inhibition, fundamental solutions such as gene manipulation are required, and acyl-homoserine lactone synthase (AHL synthase), which synthesizes the universal quorum-sensing signal of gram-negative bacteria, can be used as a target. In this study, researchers synthesized His-tagged AHL synthase and its deletion mutant that lacks the active site and compared their biochemical characteristics. His-YpeI, the 6x His-tagged AHL synthase of Serratia fonticola, and His-ΔYpeI, its deletion mutant, were designed, and their property conservation were examined using in silico projection tools. For in vitro synthesis of enzymes, the His-YpeI CFPS template was synthesized by in vitro gene synthesis, and the His-ΔYpeI CFPS template was obtained by deletion PCR. CFPS was performed and the products were purified with the 6x His-tag. The enzymes' properties were compared using an enzymatic assay. The bioinformatic analysis confirmed the conservation of biochemical properties between 6x His-tagged and untagged enzymes, including helix-turn-helix interactions, hydropathy profiles, and tertiary structure between His-YpeI and YpeI and between His-ΔYpeI and ΔYpeI. His-YpeI and His-ΔYpeI synthesized by CFPS were found to have the expected molecular weights and demonstrated distinct differences in enzyme activity. The analyzed enzymatic constants supported a significant decrease in substrate affinity and reaction rate as a result of YpeI's enzyme active site deletion. This result showed that CFPS could be used for in vitro protein synthesis, and quorum sensing could be inhibited at the enzymatic level due to the enzyme active site's deletion mutation.

Quorum sensing inhibition through site-directed mutation by deletion PCR.

Quorum sensing induces biofilms and virulence factors that are adverse industrially and medically. Nowadays, quorum sensing inhibitions focus on signal analogs or signal degradation, but these methods have several downsides, which are temporal and affected by several environmental factors. In this research, we used deletion PCR to perform site-directed mutagenesis on the quorum sensing pathway gene and then analyzed its effects on quorum sensing. Serratia fonticola DSM 4576 strain was utilized as the research strain, and the gram-negative bacteria's universal quorum sensing pathway, which is conducted by acyl-homoserine lactone (AHL), was analyzed. The structure and active site of the AHL synthase enzyme encoded by S. fonticola DSM 4576's luxI-type gene were predicted. The gene's partial section solely encodes the enzyme's active site. By using sequence and ligation-independent cloning, the obtained mutagenic gene was cloned into the suicide vector pEX18Ap. The recombinant vector was used to transform wild-type S. fonticola DSM 4576 strains, and the mutants were determined through two-step selections and PCR genotyping. The gene expression level and biofilm formation were quantitatively analyzed through RT-PCR and biofilm assay, and no significant difference was noted in the gene expression between wild types and mutants. However, when mutants were compared to wildtypes, there was a significant decrease in biofilm formation as a result of quorum sensing induced bioreaction. Thus, we propose a quorum sensing inhibitory technique based on enzyme mutation on the quorum sensing pathway, and we proved the feasibility of enzyme active site's site-directed mutation through deletion PCR.

Daclatasvir Plus Asunaprevir for the Treatment of Patients with Hepatitis C Virus Genotype 1b Infection: Real-World Efficacy, Changes in Liver Stiffness and Fibrosis Markers, and Safety.

BACKGROUND/AIMS: The treatment with daclatasvir plus asunaprevir (DCV+ASV) is associated with potent antiviral effects in patients with genotype 1b hepatitis C virus (HCV) infection. We investigated the real-world efficacy, changes in liver stiffness and noninvasive fibrosis markers, and the safety of DCV+ASV treatment in Korean patients. METHODS: In total, 363 patients with chronic hepatitis C were treated with DCV+ASV between August 2015 and January 2017. Finally, we analyzed the data of 270 patients who were monitored for at least 12 weeks after the end of treatment. RESULTS: The mean age was 60.7 years, and females predominated (60.4%). Most patients (64.8%) were treatment-naïve, and 56 patients (20.7%) had cirrhosis. Two hundred fifty-seven (95.2%) and 251 (93.0%) patients achieved end-of-treatment responses and sustained virological responses at 12 weeks posttreatment (SVR12), respectively. The SVR12 rates were higher in patients who were ＜65 years of age, males, without cirrhosis and had lower HCV RNA levels. All LS values and fibrosis-4 and aspartate aminotransferase-to-platelet ratio index values declined from baseline to the time of assessment of SVR12. CONCLUSIONS: The DCV+ASV therapy resulted in a high SVR12 and improved liver fibrosis; the treatment was well tolerated in patients with genotype 1b HCV infections.