Sensitive and selective detection of uracil-DNA glycosylase activity with a new pyridinium luminescent switch-on molecular probe.

Uracil-deoxyribonucleic acid glycosylase (UDG) is known to function as an important base-excision repair enzyme and eliminate uracil from DNA molecules to maintain genomic integrity. A new small organic molecule (DID-VP) with interesting structural properties was synthesized as a G-quadruplex selective ligand and was demonstrated to be a sensitive luminescent switch-on probe in a convenient luminescent assay specifically for UDG detection in fetal bovine serum samples under rapid and simple conditions. This newly developed analytical method is based on the UDG enzymatic activity to unwind a duplex DNA substrate, and comprises a G-quadruplex-forming sequence (ON1) and uracil-containing DNA strand (ON2) to generate a remarkable fluorescence signal through the specific interaction of DID-VP with ON1. This luminescent switch-on assay is able to achieve high sensitivity and specificity for UDG over other enzymes. The application range of the present analytical system is found to be 0.05 to 1.00 U mL(-1) UDG with a very low detection limit of 0.005 U mL(-1). The recovery study of UDG in real samples gave a very good performance with 75.05%-102.7% recovery. In addition, an extended application of the assay in screening of UDG inhibitors is demonstrated. A good dose-dependence of the luminescence response with respect to the concentration of UDG inhibitors in samples was observed.

Robotic-assisted versus conventional laparoscopic surgery for colorectal cancer: Short-term outcomes at a single center.

BACKGROUND: The robotic technique has been established as an alternative approach to laparoscopy for colorectal surgery. The aim of this study was to compare the short-term outcomes of robot-assisted and laparoscopic surgery in colorectal cancer. METHODS: The cases of robot-assisted or laparoscopic colorectal resection were collected retrospectively between July 2015 and September 2018. We evaluated patient demographics, perioperative characteristics, and pathologic examinations. Short-term outcomes included time to passage of flatus and length of postoperative hospital stay. RESULTS: A total of 580 patients were included in the study. There were 271 patients in the robotic colorectal surgery (RCS) group and 309 in the laparoscopic colorectal surgery (LCS) group. The time to passage of flatus in the RCS group was 3.62 days shorter than the LCS group. The total costs were increased by 2,258.8 USD in the RCS group compared to the LCS group (P < 0.001). CONCLUSION: The present study suggests that colorectal cancer robotic surgery was more beneficial to patients because of a shorter postoperative recovery time of bowel function and shorter hospital stays.

Short-Term Outcomes of Robotic versus Laparoscopic Total Mesorectal Excision for Rectal Cancer: A Cohort Study.

The aim of this study was to evaluate and compare the intestinal function recovery time and other short-term outcomes between robotic-assisted total mesorectal excision (R-TME) and laparoscopic total mesorectal excision (L-TME) for rectal cancer. This is a retrospective study using a prospectively collected database. Patients' records were obtained from Gansu Provincial Hospital between July 2015 and October 2017. Eighty patients underwent R-TME, and 116 with the same histopathological stage of the tumor underwent an L-TME. Both operations were performed by the same surgeon, comparing intra- and postoperative outcomes intergroups. The time to the first passage of flatus (P < 0.001), the time to the first postoperative oral fluid intake (P < 0.001), and the length of hospital stay (P < 0.01) of the R-TME group were about three days faster than those in the L-TME group. The rate of conversion to open laparotomy (P = 0.038) and postoperative urinary retention (P = 0.016) were significantly lower in the R-TME group than in the L-TME group. Intraoperative blood loss of the R-TME group was more than that of the L-TME group (P < 0.01).The operation time, number of lymph nodes harvested, and rate of positive circumferential resection margin were similar intergroup. The total cost of the R-TME group was higher than that of the L-TME group, but with a lack of statistical significance (85,623.91 ± 13,310.50 vs 67,356.79 ± 17,107.68 CNY, P = 0.084). The R-TME is safe and effective and has better postoperative short-term outcomes and faster intestinal function recovery time, contrasting with the L-TME. The large, multicenter, prospective studies were needed to validate the advantages of robotic surgery system used in rectal cancer.

New pyridinium-based fluorescent dyes: A comparison of symmetry and side-group effects on G-Quadruplex DNA binding selectivity and application in live cell imaging.

A series of C1-, C2-and C3-symmetric pyridinium conjugates with different styrene-like side groups were synthesized and were utilized as G-quadruplex selective fluorescent probes. The new compounds were well-characterized. Their selectivity, sensitivity, and stability towards G-quadruplex were studied by fluorescence titration, native PAGE experiments, FRET and circular dichroism (CD) analyses. These new compounds investigated in the fluorescence assays were preferentially bound with G-quadruplex DNA compared with other type of nucleic acids and it is fascinating to realize the effects of molecular symmetry and associated side groups showing unexpectedly great influence on the fluorescent signal enhancement for the discrimination of G-quadruplexes DNA from other nucleic acids. This may correlate with the pocket symmetry and shape of the G-quadruplex DNA inherently. Among the compounds, a C2-symmetric dye (2,6-bis-((E)-2-(1H-indol-3-yl)-vinyl)-1-methylpyridin-1-ium iodide) with indolyl-groups substituted was screened out from the series giving the best selectivity and sensitivity towards G-quadruplexes DNA, particularly telo21, due to its high equilibrium binding constant (K=2.17×10(5)M(-1)). In addition, the limit of detection (LOD) of the dye to determine telo21 DNA in bioassays was found as low as 33nM. The results of the study give insight and certain crucial factors, such as molecular symmetry and the associated side groups, on developing of effective fluorescent dyes for G-quadruplex DNA applications including G-quadruplex structure stabilization, biosensing and clinical applications. The compound was also demonstrated as a very selective G-quadruplex fluorescent agent for living cell staining and imaging.

Molecular Engineering of Thiazole Orange Dye: Change of Fluorescent Signaling from Universal to Specific upon Binding with Nucleic Acids in Bioassay.

The universal fluorescent staining property of thiazole orange (TO) dye was adapted in order to be specific for G-quadruplex DNA structures, through the introduction of a styrene-like substituent at the ortho-position of the TO scaffold. This extraordinary outcome was determined from experimental studies and further explored through molecular docking studies. The molecular docking studies help understand how such a small substituent leads to remarkable fluorescent signal discrimination between G-quadruplex DNA and other types of nucleic acids. The results reveal that the modified dyes bind to the G-quadruplex or duplex DNA in a similar fashion as TO, but exhibit either enhanced or quenched fluorescent signal, which is determined by the spatial length and orientation of the substituent and has never been known. The new fluorescent dye modified with a p-(dimethylamino)styryl substituent offers 10-fold more selectivity toward telomeric G-quadruplexes than double-stranded DNA substrates. In addition, native PAGE experiments, FRET, CD analysis, and live cell imaging were also studied and demonstrated the potential applications of this class of thiazole-orange-based fluorescent probes in bioassays and cell imaging.

A molecular fluorescent dye for specific staining and imaging of RNA in live cells: a novel ligand integration from classical thiazole orange and styryl compounds.

A new RNA-selective fluorescent dye integrated with a thiazole orange and a p-(methylthio)styryl moiety shows better nucleolus RNA staining and imaging performance in live cells than the commercial stains. It also exhibits excellent photostability, cell tolerance, and counterstain compatibility with 4',6-diamidino-2-phenylindole for specific RNA-DNA colocalization in bioassays.