Adoption of an Enhanced Recovery after Surgery Protocol for Neuromuscular Scoliosis Shortens Length of Hospital Stay.

Introduction: We aimed to implement the enhanced recovery after surgery (ERAS) protocol for pediatric neuromuscular scoliosis (NMS) surgery and to examine the effectiveness of this program in this study. Methods: Subjects were children with NMS who underwent scoliosis surgery at our department by a surgeon using a single posterior approach. A series of 27 cases before the introduction of ERAS and 27 cases during program stabilization were included in the study. Patient backgrounds did not show significant differences before and after introducing ERAS. Perioperative data, complications, length of hospital stay (LOS), and readmission within 90 days were investigated and statistically analyzed. Results: When the pre- and post-ERAS induction groups were compared, no significant differences in anesthesia induction time (p=0.979), pelvic fixation (p=0.586), fusion levels (p=0.479), intraoperative hypothermia duration (p=0.154), end-of-surgery body temperature (p=0.197), operative time (p=0.18), postoperative main Cobb angle (p=0.959), main Cobb angle correction rate (p=0.91), postoperative spino-pelvic obliquity (SPO) (p=0.849), and SPO correction rate (p=0.267) were observed. However, significant differences in using V-flap technique (p=0.041), intraoperative blood loss (p=0.001), and LOS (p=0.001) were observed. Intraoperative blood loss was weakly correlated with LOS (p=0.432 and 0.001). No statistically significant difference existed between the V-flap method and LOS (p=0.265). Multiple regression analysis using LOS as the objective variable and ERAS protocols and intraoperative blood loss as explanatory variables revealed that the effect of ERAS on LOS was greater than that of intraoperative blood loss. No statistically significant differences in the readmission rates within 90 days were found. Conclusions: After the introduction of ERAS, LOS decreased without an increase in complications or readmissions within 90 days.

Characterization of the membrane penetration-enhancing peptide S19 derived from human syncytin-1 for the intracellular delivery of TAT-fused proteins.

Although cell-penetrating peptides such as the HIV-derived TAT peptide have been used as tools for the intracellular delivery of therapeutic peptides and proteins, a problem persists: the endosomal escape efficiency is low. Previously, we found that the fusogenic peptide S19, derived from the human protein syncytin-1, enhance the endosomal escape efficiency of proteins that incorporated by endocytosis via TAT. In this study, we first performed Ala-scanning mutagenesis of S19, and found that all Ile, Val, Leu and Phe with high ß-sheet forming propensities in S19 are important for the intracellular uptake of S19-TAT-fused proteins. In a secondary structure analysis of the mutated S19-TAT peptides in the presence of liposomes mimicking late endosomes (LEs), the CD spectra of V3A and I4A mutants with low uptake activity showed the appearance of an α-helix structure, whereas the mutant G5A retained both the uptake activity and the ß-structure. In addition, we investigated the appropriate linking position and order of the S19 and TAT peptides to a cargo protein including an apoptosis-induced peptide and found that both the previous C-terminal S19-TAT tag and the N-terminal TAT-S19 tag promote the cytoplasmic delivery of the fusion protein. These results and previous results suggest that the interaction of TAT with the LE membrane causes a structural change in S19 from a random coil to a ß-strand and that the subsequent parallel ß-sheet formation between two S19 peptides may promote adjacent TAT dimerization, resulting in endosomal escape from the LE membrane.

Isolation of a methylated mannose-binding protein from terrestrial worm Enchytraeus japonensis.

To elucidate a biological role of the methylated mannose residues found in N-glycans of terrestrial worm Enchytraeus japonensis, we first synthesized 3-O-methyl mannose- and 4-O-methyl mannose-derivatives and immobilized them to Sepharose 4B beads in order to isolate the sugar-binding protein. When whole protein extracts from the worms was applied to a series of the columns immobilized with the modified and unmodified mannose-derivatives, respectively, a protein with a molecular weight of 25,000 was isolated by 4-O-methyl mannose-immobilized column chromatography, and termed as a methylated mannose-binding protein (mMBP). mMBP bound weakly to a mannose-immobilized column and moderately to a 3-O-methyl mannose-immobilized column. The N-terminal amino acid sequences of mMBP and its endoprotease-digested peptides were determined. Using the degenerate first primers synthesized based on the primary sequence, a genomic DNA fragment was isolated. Then, the second primers were synthesized based on the genomic DNA fragment, and with use of them two cDNA fragments were obtained by the 3'- and 5'-RACE methods. Finally, the third primers were synthesized based on the sequences of the two cDNA fragments and one genomic DNA fragment, and with use of them a full-length cDNA of mMBP was isolated and shown to comprise a putative 633 bp open reading frame encoding 210 amino acid residues. BLAST analysis revealed that mMBP has identities by 26 ~ 55% to several proteins including the regeneration-upregulated protein 3 from the same species. Whether mMBP is involved in the regeneration of the worm is under investigation.