Cytokine engineered NK-92 therapy to improve persistence and anti-tumor activity.

Natural killer (NK) cells are an attractive cell source in cancer immunotherapy due to their potent antitumor ability and promising safety for allogenic applications. However, the clinical outcome of NK cell therapy has been limited due to poor persistence and loss of activity in the cytokine-deficient tumor microenvironment. Benefits from exogenous administration of soluble interleukin-2 (IL-2) to stimulate the activity of NK cells have not been significant due to cytokine consumption and activation of other immune cells, compromising both efficacy and safety. Methods: To overcome these drawbacks, we developed a novel membrane-bound protein (MBP) technology to express IL-2 on the surface of NK-92 cells (MBP NK) inducing autocrine signal for proliferation without IL-2 supplementation. Results: The MBP NK cells exhibited not only improved proliferation in IL-2 deficient conditions but also stronger secretion of cytolytic granules leading to enhanced anti-tumor activity both in vitro and in vivo. Furthermore, the experiment with a spheroid solid tumor model exhibited enhanced infiltration by MBP NK cells creating higher local effector-to-target ratio for efficient tumor killing. These results suggest MBP technology can be an effective utility for NK-92 cell engineering to increase anti-tumor activity and reduce potential adverse effects, providing a higher therapeutic index in clinical applications.

Comparison of the UEscope Video Laryngoscope with the Traditional Direct Laryngoscope in Neonates and Infants: A Randomized Clinical Trial.

The role of video laryngoscopy in adults is well established, but its role in children is still inconclusive. Previous studies on the UEscope in pediatric patients with difficult airways showed that it could reduce the time to intubation (TTI) compared to a conventional direct laryngoscope. The main objective of the current study was to investigate if the use of the UEscope could reduce the TTI in neonates and infants. Forty patients under 12 months old were recruited from a single tertiary hospital from March 2020 to September 2021 and were randomly assigned to the direct laryngoscope group (n = 19, neonates = 4, infants = 15) or UEscope group (n = 21, neonates = 6, infants = 15). Although the quality of glottic view was comparable in both groups, the TTI was significantly lower in the UEscope group in both the "intention-to-treat" (-19.34 s, 95% confidence interval = -28.82 to -1.75, p = 0.0144) and "as treated" (-11.24 s, 95% confidence interval: -21.73 to 0, p = 0.0488) analyses. The UEscope may be a better choice for tracheal intubation than conventional direct laryngoscope in neonates and infants.

Non-invasive optical control of endogenous Ca2+ channels in awake mice.

Optogenetic approaches for controlling Ca2+ channels provide powerful means for modulating diverse Ca2+-specific biological events in space and time. However, blue light-responsive photoreceptors are, in principle, considered inadequate for deep tissue stimulation unless accompanied by optic fiber insertion. Here, we present an ultra-light-sensitive optogenetic Ca2+ modulator, named monSTIM1 encompassing engineered cryptochrome2 for manipulating Ca2+ signaling in the brain of awake mice through non-invasive light delivery. Activation of monSTIM1 in either excitatory neurons or astrocytes of mice brain is able to induce Ca2+-dependent gene expression without any mechanical damage in the brain. Furthermore, we demonstrate that non-invasive Ca2+ modulation in neurons can be sufficiently and effectively translated into changes in behavioral phenotypes of awake mice.

Structure-based glycoengineering of interferon lambda 4 enhances its productivity and anti-viral potency.

Interferon lambda 4 (IFNλ4) has been recently known and studied for its role in hepatitis C virus (HCV) infection, but its clinical potential is significantly hampered due to its poor expression in vitro. Our study reports the successful production of IFNλ4 from a mammalian cell line through a glycoengineering and structure-based approach. We introduced de novo N-glycosylation of IFNλ4, guided by structural analysis, and produced IFNλ4 variants in Expi293F that displayed improved expression and potency. To preserve the structure and functionality of IFNλ4, the model structure of the IFNλ4 signaling complex was analyzed and the N-glycosylation candidate sites were selected. The receptor binding activity of engineered IFNλ4 variants and their receptor-mediated signaling pathway were similar to the E. coli version of IFNλ4 (eIFNλ4), while the antiviral activity and induction levels of interferon-stimulated gene (ISG) were all more robust in our variants. Our engineered IFNλ4 variants may be further developed for clinical applications and utilized in basic research to decipher the immunological roles of IFNλ4.

Comparison between Post-Operative Analgesic Efficacy of Low-Concentration High-Volume and High-Concentration Low-Volume Combinations of Ropivacaine for Transverse Abdominis Plane Block in Pediatric Open Inguinal Hernia Repair.

Transverse abdominis plane (TAP) block can provide post-operative analgesia in children undergoing open inguinal hernia repair. However, the optimal anesthetic dose, and concentration for TAP block in the pediatric population, is not well defined. This study compared the post-operative analgesic effect of TAP block between low-concentration, with high-volume (LCHV) and high-concentration, with low-volume (HCLV) combinations of local anesthetic. Forty-four patients who underwent open inguinal hernia repair were randomly assigned to two groups. The patients in the LCHV group received 0.67 mL/kg of 0.15% ropivacaine. Whereas, those in the HCLV group received 0.4 mL/kg of 0.25% ropivacaine. Both groups received the same amount of ropivacaine (1 mg/kg). The primary outcome measure was the face, leg, activity, cry, consolability (FLACC) scale score at post-anesthetic care unit (PACU; T1). FLACC scale score at T1 was significantly lower in the HCLV group (2.91 versus 1.43; mean difference, -1.49; 95% confidence interval, -0.0245 to -2.936; p = 0.0464). FLACC scale scores one hour and six hours after the surgery were not different between the two groups. This study reports better post-operative analgesic effects after unilateral open inguinal hernia repair with 1 mg/kg of 0.25% ropivacaine than 1 mg/kg of 0.15% ropivacaine at PACU.

Psychometric properties of transitional care instruments and their relationships with health literacy: Brief PREPARED and Care Transitions Measure.

OBJECTIVE: To assess the psychometric properties of the Brief PREPARED (B-PREPARED) and the Care Transitions Measure (CTM) in patients discharged from hospitals in Korea, and examine their relationships with health literacy. DESIGN: A cross-sectional study with a follow-up telephone survey 4 weeks post-discharge. SETTING: Six medical and surgical wards in a tertiary hospital in Seoul. PARTICIPANTS: 293 patients discharged from general wards. MAIN OUTCOME MEASURES: Psychometric properties of the Korean versions of the B-PREPARED and the 15- and 3-item CTM (CTM-15 and CTM-3), and the 3-item Brief Screening Questionnaire for health literacy. RESULTS: All instruments discriminated well between patients who were satisfied with hospital care and the discharge process, and those who were not. One month post-discharge, the CTM-15 score was significantly higher in patients with good health status than the others, and the CTM-3 score was significantly higher in patients who used outpatient care than the others. However, no significant difference was found in the scores for all instruments by emergency department visits, rehospitalization, and medication adherence. Cronbach's alpha values were 0.69 for B-PREPARED, 0.91 for CTM-15 and 0.67 for CTM-3. Intraclass correlation coefficients were 0.64, 0.75 and 0.66, respectively. Approximately 33% of the participants had limited health literacy. After adjusting for other patient variables, those with inadequate health literacy had lower scores on all instruments. CONCLUSIONS: Although the three instruments had acceptable validity and reliability, they showed limited criterion validity. Patients with limited health literacy should be supported to ensure the quality of transitional care.

IFN-λ4 potently blocks IFN-α signalling by ISG15 and USP18 in hepatitis C virus infection.

Genetic polymorphisms in IFNL4 have been shown to predict responses to IFN-α-based therapy in hepatitis C virus (HCV)-infected patients. The IFNL4-ΔG genotype, which encodes functional IFN-λ4 protein, is associated with a poor treatment response. In the present study, we investigated the induction and biological effects of IFN-λ4 in HCV-infected hepatocytes and their association with responsiveness to IFN-α. We also studied the effects of direct-acting antiviral (DAA) treatment on IFN-λ4 expression and IFN-α responsiveness. HCV infection induced IFN-λ4 expression at mRNA and protein levels in primary human hepatocytes (PHHs). In hepatoma cells, IFNL4 gene transfection or recombinant IFN-λ4 protein treatment robustly increased the protein levels of ISG15 and USP18 in an IFNLR1-dependent manner and potently blocked IFN-α signalling. The ISG15/USP18-mediated IFN-α unresponsiveness was demonstrated by transfection of siRNAs targeting ISG15 and/or USP18. This potent IFN-λ4 effect was related to prolonged ISG expression after IFNL4 gene transfection. DAA treatment of HCV-infected PHHs reduced the expression of IFN-λs, including IFN-λ4, and restored IFN-α responsiveness. These results demonstrate that virus-induced IFN-λ4 potently blocks IFN-α signalling by inducing high protein levels of ISG15 and USP18. Moreover, the data clearly demonstrate that DAA therapy restores IFN-α responsiveness in HCV-infected cells.

Molecular Architecture of Yeast Chromatin Assembly Factor 1.

Chromatin Assembly Complex 1 (CAF-1) is a major histone chaperone involved in deposition of histone H3 and H4 into nucleosome. CAF-1 is composed of three subunits; p150, p60 and p48 for human and Cac1, Cac2 and Cac3 for yeast. Despite of its central role in chromatin formation, structural features of the full CAF-1 in complex with histones and other chaperones have not been well characterized. Here, we dissect molecular architecture of yeast CAF-1 (yCAF-1) by cross-linking mass spectrometry (XL-MS) and negative stain single-particle electron microscopy (EM). Our work revealed that Cac1, the largest subunit of yCAF-1, might serve as a major histone binding platform linking Cac2 and Cac3. In addition, EM analysis showed that yCAF-1 adopts a bilobal shape and Cac1 connecting Cac2 and Cac3 to generate a platform for binding histones. This study provides the first structural glimpse of the full CAF-1 complex and a structural framework to understand histone chaperoning processes.

Designed nucleases for targeted genome editing.

Targeted genome-editing technology using designed nucleases has been evolving rapidly, and its applications are widely expanding in research, medicine and biotechnology. Using this genome-modifying technology, researchers can precisely and efficiently insert, remove or change specific sequences in various cultured cells, micro-organisms, animals and plants. This genome editing is based on the generation of double-strand breaks (DSBs), repair of which modifies the genome through nonhomologous end-joining (NHEJ) or homology-directed repair (HDR). In addition, designed nickase-induced generation of single-strand breaks can also lead to precise genome editing through HDR, albeit at relatively lower efficiencies than that induced by nucleases. Three kinds of designed nucleases have been used for targeted DSB formation: zinc-finger nucleases, transcription activator-like effector nucleases, and RNA-guided engineered nucleases derived from the bacterial clustered regularly interspaced short palindromic repeat (CRISPR)-Cas (CRISPR-associated) system. A growing number of researchers are using genome-editing technologies, which have become more accessible and affordable since the discovery and adaptation of CRISPR-Cas9. Here, the repair mechanism and outcomes of DSBs are reviewed and the three types of designed nucleases are discussed with the hope that such understanding will facilitate applications to genome editing.

Gliomatosis peritonei of the scrotal sac associated with an immature gastric teratoma.

A male infant presented with bilateral scrotal masses 7 months after removal of an immature gastric teratoma. After surgical excision, histology showed gliomatosis peritonei. We report an unusual presentation of gliomatosis peritonei arising from a rare lesion of immature gastric teratoma.