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Abstract Akkermansia muciniphila has received great attention because of its beneficial roles in gut health by regulating gut immunity, promoting intestinal epithelial development, and improving barrier integrity. However, A. muciniphila-derived functional molecules regulating gut health are not well understood. Microbiome-secreted proteins act as key arbitrators of host-microbiome crosstalk through interactions with host cells in the gut and are important for understanding host-microbiome relationships. Herein, we report the biological function of Amuc_1409, a previously uncharacterised A. muciniphila-secreted protein. Amuc_1409 increased intestinal stem cell (ISC) proliferation and regeneration in ex vivo intestinal organoids and in vivo models of radiation- or chemotherapeutic drug-induced intestinal injury and natural aging with male mice. Mechanistically, Amuc_1409 promoted E-cadherin/β-catenin complex dissociation via interaction with E-cadherin, resulting in the activation of Wnt/β-catenin signaling. Our results demonstrate that Amuc_1409 plays a crucial role in intestinal homeostasis by regulating ISC activity in an E-cadherin-dependent manner and is a promising biomolecule for improving and maintaining gut health.

Abstract Background In cases of wrist arthritis, proximal row carpectomy (PRC) has been widely utilized and shown favorable long-term outcomes. However, its applicability is limited in cases where arthritis extends to the lunate fossa or capitate. Recently, surgical approaches combining various methods of interposition arthroplasty have been introduced to overcome these drawbacks. The purpose of this study was to perform PRC and interposition arthroplasty with dorsal capsule and acellular dermal matrix(ADM),and analyze the clinical outcomes of these procedures. Methods Fourteen cases who underwent PRC and interposition arthroplasty using both dorsal capsular flap and ADM were retrospectively recruited. The researchers assessed the patients’ Visual Analog Scale (VAS) pain score, Disabilities of the Arm, Shoulder and Hand (DASH) scores, range of motion (ROM), retear, and radiocarpal distance (RCD). Results One year post-surgery, both the VAS pain scores, DASH scores, and ROM showed statistically significant improvement compared to before the surgery. Upon reviewing the radiological results, the postoperative mean RCD was 4.8 ± 0.8 mm and one year follow up mean RCD was 3.6 ± 0.5 mm at one year post-surgery. Moreover, in the one year follow-up, there was no observed failure of the allodermis graft in any of the cases. Conclusion The PRC and interposition arthroplasty with ADM demonstrated significantly improved clinical outcomes after surgery, showing a maintain of RCD without graft failure effectively.

In this paper, a heating device is implemented by considering two large factors in a 100 cmm RTO design. First, when the combustion chamber is used for a long time with a high temperature of 750–1100 °C depending on the high concentration VOC gas capacity, there is a problem that the combustion chamber explodes or the function of the rotary is stopped due to the fatigue and load of the device. To prevent this, the 100 cmm RTO design with a changed rotary position is improved. Second, an RTO design with a high-heating element is implemented to combust VOC gas discharged from the duct at a stable temperature. Through this, low-emission combustion emissions and energy consumption are reduced. By implementing a high heat generation device, the heat storage combustion oxidation function is improved through the preservation of renewable heat. Over 177 h of demonstration time, we improved the function of 100 cm by discharging 99% of VOC’s removal efficiency, 95.78% of waste heat recovery rate, 21.95% of fuel consumption, and 3.9 ppm of nitrogen oxide concentration.

Corn peptide (CP) is a short, naturally occurring, and physiologically active peptide generated from corn-protease-catalyzed hydrolysis. CP plays a role in preventing obesity-related disorders, but its impact on reducing inflammation is unknown. Hence, this study examined the possible protective effects of corn peptide powder (CPP) against the harmful effects of lipopolysaccharide (LPS), with a particular emphasis on reducing oxidative damage and inflammation in adipocytes. Hence, mature 3T3-L1 adipocytes underwent exposure to 10 ng/mL LPS, with or without CPP (10 and 20 μg/mL). LPS stimulation increased reactive oxygen species and superoxide anion generation. However, this effect was reduced in a dose-dependent manner by pretreatment with CPP. CPP treatment elevated the mRNA expressions of the antioxidant enzymes manganese superoxide dismutase (mnSOD) and glutathione peroxidase 1 (Gpx1) while reducing the mRNA expressions of the cytosolic reactive oxygen species indicators p40 and p67 (NADPH oxidase 2). In addition, CPP inhibited the monocyte chemoattractant protein-1, tumor necrosis factor-alpha, Toll-like receptor 4, and nuclear factor kappa B mRNA expressions induced by LPS. These findings demonstrate that CPP may ameliorate adipocyte dysfunction by suppressing oxidative damage and inflammatory responses through a new mechanism known as Toll-like receptor 4/nuclear factor kappa B-mediated signaling.

Introduction Low-dose radiation therapy (LDRT) for osteoarthritis (OA) has been performed for several decades. However, supporting evidence from randomised studies using modern methodologies is lacking, and a recently published randomised study failed to show the significant benefit of LDRT. The presented trial aims to evaluate the efficacy and safety of LDRT for patients with knee OA.Methods and analysis This prospective, multicentre, randomised trial will be conducted in the Republic of Korea. A total of 114 participants will be randomly assigned (1:1:1) to receive sham irradiation, 0.3 Gy/6 fractions of LDRT or 3 Gy/6 fractions of LDRT. Key inclusion criteria are primary knee OA with Kellgren-Lawrence grade 2–3 and visual analogue scale 50–90 when walking at the baseline. The primary endpoint is the rate of responders at 4 months after LDRT according to the OARSI-OMERACT criteria. Concomitant use of analgesics is prohibited until the primary efficacy evaluation is scheduled.Ethics and dissemination Currently, approval from the Ministry of Food and Drug Safety of the Republic of Korea and the institutional review board of each participating hospital has been obtained. Patient enrolment began in October 2022 and is ongoing at three participating sites. The results will be disseminated to academic audiences and the public via publication in an international peer-reviewed journal and presentation at conferences. This trial will provide valuable information on the safety and efficacy of LDRT for patients with knee OA.Trial registration number NCT05562271.

Abstract Background The gut microbiota is associated with diverse age-related disorders. Several rejuvenation methods, such as probiotic administration and faecal microbiota transplantation, have been applied to alter the gut microbiome and promote healthy ageing. Nevertheless, prolongation of the health span of aged mice by remodelling the gut microbiome remains challenging. Results Here, we report the changes in gut microbial communities and their functions in mouse models during ageing and three rejuvenation procedures including co-housing, serum-injection and parabiosis. Our results showed that the compositional structure and gene abundance of the intestinal microbiota changed dynamically during the ageing process. Through the three rejuvenation procedures, we observed that the microbial community and intestinal immunity of aged mice were comparable to those of young mice. The results of metagenomic data analysis underscore the importance of the high abundance of Akkermansia and the butyrate biosynthesis pathway in the rejuvenated mouse group. Furthermore, oral administration of Akkermansia sufficiently ameliorated the senescence-related phenotype in the intestinal systems in aged mice and extended the health span, as evidenced by the frailty index and restoration of muscle atrophy. Conclusions In conclusion, the changes in key microbial communities and their functions during ageing and three rejuvenation procedures, and the increase in the healthy lifespan of aged mice by oral administration of Akkermansia. Our results provide a rationale for developing therapeutic strategies to achieve healthy active ageing. Video abstract

Background Surgical techniques related to soft tissue management play critical roles in optimizing surgical outcomes and patient satisfaction in total knee arthroplasty (TKA). Despite the importance of wound closure and bleeding management approaches, no published guidelines/consensus are available. Methods Twelve orthopedic surgeons participated in a modified Delphi panel consisting of 2 parts (each part comprising two rounds) from September–October 2018. Questionnaires were developed based on published evidence and guidelines on surgical techniques/materials. Questionnaires were administered via email (Round 1) or at a face-to-face meeting (subsequent rounds). Panelists ranked their agreement with each statement on a five-point Likert scale. Consensus was achieved if ≥70% of panelists selected 4/5, or 1/2. Statements not reaching consensus in Round 1 were discussed and repeated or modified in Round 2. Statements not reaching consensus in Round 2 were excluded from the final consensus framework. Results Consensus was reached on 13 goals of wound management. Panelists agreed on 38 challenges and 71 strategies addressing surgical techniques or wound closure materials for each tissue layer, and management strategies for blood loss reduction or deep vein thrombosis prophylaxis in TKA. Statements on closure of capsular and skin layers, wound irrigation, dressings and drains required repeat voting or modification to reach consensus. Conclusion Consensus from Asia-Pacific TKA experts highlights the importance of wound management in optimizing TKA outcomes. The consensus framework provides a basis for future research, guidance to reduce variability in patient outcomes, and can help inform recommendations for wound management in TKA.

Background: Few studies have evaluated the impact of obesity on operation time in patients with ACL reconstruction. The purpose of this study was to understand the effect of obesity on operation time in patients with arthroscopic anterior cruciate ligament (ACL) reconstruction. Methods: A total of 103 patients were included. The mean pure operation time was 45.9 ± 13.4 min. Considering that 15 min incremental increases in operation time are an independent risk factor for complications, all patients were classified into two groups according to operation time: more or less than 61 min. Demographic data were compared between both groups. Pure operation time was defined as operative time without suture time (pure operation time = suture start time − operation start time). Correlation analysis between demographic data and pure operation time was performed, and multiple linear regression analysis was used to identify the predictors of pure operation time. Results: The pure operation time ≥61 min group (n = 34) had a 14.7 kg higher weight and 4.5 kg/m2 higher body mass index (BMI) than those with pure operation time < 61 min (n = 69). Weight (r = 0.635, p < 0.001) and BMI (r = 0.584, p < 0.001) were positively correlated with operation time. Multiple linear regression analysis showed that weight (β = 0.635, p < 0.001) was the only predictor of operation time. A weight of 74.25 kg was a cut-off value for a pure operation time of >61 min. Conclusions: The weight and BMI of the group with pure operation time of ≥61 min were 14.7 kg and 4.5 kg/m2 higher, respectively. The weight of patients with ACL tears was a factor affecting delay in the operation time. Patients weighing over 74.25 kg were more likely to delay ACL reconstruction.

This study investigated the rate of cartilage regeneration after an open-wedge high tibial osteotomy (HTO) without cartilage regeneration by second-look arthroscopy. This study included patients who underwent an open-wedge HTO between July 2014 and March 2019. A total of 65 patients were enrolled. Pre- and postoperative (second-look arthroscopy) hip–knee–ankle (HKA) angle and tibial slope were measured. All patients underwent arthroscopic examination prior to osteotomy. Medial femoral condyle (MFC) and medial tibial plateau (MTP) articular cartilage were evaluated according to the International Cartilage Repair Society (ICRS) grading system. After 26.5 months, second-look arthroscopy was performed with plate removal to identify the cartilage status of the MFC and MTP. The preoperative HKA angle (6.4° ± 2.7°) was well corrected postoperatively (−2.7° ± 2.7°, p < 0.001). In terms of MFC on second-look arthroscopy, 29 patients (44.6%) showed an improved ICRS grade, 31 patients (47.7%) were maintained, and 5 patients (7.7%) showed a worse ICRS grade since the prior operation. In the MTP group, 19 patients (29.2%) improved, 44 patients (67.7%) were maintained, and 2 patients (3.1%) worsened. Approximately 44.6% and 29.2% of patients showed improved cartilage statuses on the MFC and MTP after open-wedge HTO without any cartilage regeneration procedures. Cartilage regenerations in both the MFC and MTP did not influence clinical outcomes.

Abstract Background Only limited data are available regarding postural stability between anterior cruciate ligament (ACL)-injured patients with medial meniscus (MM) tear and those with lateral meniscus (LM) tear. The purpose of this study was to compare preoperative postural stability for both involved and uninvolved knees in ACL rupture combined with MM and LM tears. It was hypothesized that there would be a significant difference in postural stability between these two groups. Methods Ninety-three ACL-injured patients (53 combined with MM tears vs. 40 combined with LM tears) were included. Static and dynamic postural stability were evaluated with the overall stability index (OSI), anterior–posterior stability index (APSI), and medial–lateral stability index (MLSI) using stabilometry. Knee muscle strength was evaluated using an isokinetic testing device. Results In the static postural stability test, none of the stability indices showed significant differences between the two groups for both knees (p > 0.05). In the dynamic postural stability test for involved side knees, the OSI and APSI were significantly higher in the LM tear group compared to the MM tear group (OSI: 2.0 ± 0.8 vs. 1.6 ± 0.5, p = 0.001; APSI: 1.5 ± 0.6 vs. 1.3 ± 0.5, p = 0.023), but not the MLSI (p > 0.05). In the static and dynamic postural stability tests in each group, there were no significant differences between the involved and uninvolved side knees (p > 0.05). There was no significant difference in the knee muscle strength between the two groups (p > 0.05). All postural stability showed no significant correlation with knee muscle strength (p > 0.05). Conclusion Dynamic postural stability was poorer in patients with ACL rupture combined with LM tear than in those with MM tear. Therefore, close monitoring for postural stability would be necessary during preoperative and postoperative rehabilitation, especially for patients with ACL rupture combined with LM tear. Level of evidence: Level III

Abstract Background The aim was to compare tunnel widening of autogenous hamstring anterior cruciate ligament reconstruction (ACLR) using cortical button versus cross-pin femoral fixation. Methods The PubMed, Embase, and Cochrane Central Register of Controlled Trials databases were searched from inception to 11 April 2019. The study included all levels of evidence in studies that reported femoral tunnel widening and compared cortical button and cross-pin femoral fixation for ACLR. Results Six studies were included, covering a total of 344 knees. Using transtibial techniques for ACLR, the mean absolute amount of femoral tunnel widening was significantly greater with cortical button fixation than with transfemoral cross-pin fixation (−0.30 mm; 95% confidence interval (CI) −0.56,−0.05 mm; p= 0.02). Using the transtibial technique, the mean relative percentage of femoral tunnel widening was significantly greater with cortical button fixation than with transfemoral cross pin fixation (−5.73%; 95% CI −10.32, −1.14% ; p= 0.01). Conclusion The present meta-analysis revealed greater widening of the femoral tunnel when using cortical button fixation for hamstring ACLR via the transtibial technique than when using transfemoral cross-pin fixation.

Summary Targeted gene regulation is indispensable for reprogramming a cellular network to modulate a microbial phenotype. Here, we adopted the type II CRISPR interference (CRISPRi) system for simple and efficient regulation of target genes in Pseudomonas putida KT2440. A single CRISPRi plasmid was generated to express a nuclease‐deficient Cas9 gene and a designed single guide RNA, under control of l‐rhamnose‐inducible PrhaBAD and the constitutive Biobrick J23119 promoter respectively. Two target genes were selected to probe the CRISPRi‐mediated gene regulation: exogenous green fluorescent protein on the multicopy plasmid and endogenous glpR on the P. putida KT2440 chromosome, encoding GlpR, a transcriptional regulator that represses expression of the glpFKRD gene cluster for glycerol utilization. The CRISPRi system successfully repressed the two target genes, as evidenced by a reduction in the fluorescence intensity and the lag phase of P. putida KT2440 cell growth on glycerol. Furthermore, CRISPRi‐mediated repression of glpR improved both the cell growth and glycerol utilization, resulting in the enhanced production of mevalonate in an engineered P. putida KT2440 harbouring heterologous genes for the mevalonate pathway. CRISPRi is expected to become a robust tool to reprogram P. putida KT2440 for the development of microbial cell factories producing industrially valuable products.

Abstract Background Genipin is a compound derived from gardenia fruit extract. Although Genipin has anti-tumor effects in various cancers, its effect and mechanism in gastric cancer remain unclear. Here, we investigated the relationship between the anticancer effect of Genipin and signal transducer and activator of transcription (Stat3)/myeloid cell leukemia-1 (Mcl-1) in human gastric cancers. Methods MTT assays were performed to determine the cell viability of gastric cancer and gastric epithelial cell lines (AGS, MKN45, SNU638, MKN74, HFE-145). A TUNEL assay and Western blotting were carried out to investigate apoptosis. Stat3 activity was measured by proteome profiler phospho kinase array, immunofluorescence and immunoblotting. Mitochondria function was monitored with an XF24 analyzer and by flow cytometry, confocal microscopy using fluorescent probes for general mitochondrial membrane potential (MMP). Results Genipin induced apoptosis in gastric cancer cells, including AGS and MKN45 cells. Genipin also reduced Mcl-1 mRNA and protein levels. Furthermore, we found that phosphorylation of Stat3 is regulated by Genipin. Additionally, the protein level of phospho Janus kinase 2 (JAK2) was decreased by Genipin treatment, indicating that the Stat3/JAK2/Mcl-1 pathway is suppressed by Genipin treatment in gastric cancer cells. Mcl-1 is closely related to mitochondrial function. These findings suggest that Genipin contributes to the collapse of mitochondrial functions like MMP. Conclusions Genipin induced apoptosis by suppressing the Stat3/Mcl-1 pathway and led to mitochondrial dysfunction. Our results reveal a novel mechanism for the anti-cancer effect of Genipin in gastric cancer.

Abstract Background This meta-analysis was designed to quantify adduction moment loss, to evaluate the relationship between changes in mechanical axis alignment and adduction moment, and to assess whether sagittal plane moment is altered after medial open wedge high tibial osteotomy (HTO). Methods Following preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines, all studies reporting preoperative and postoperative peak knee adduction moment or change in peak knee adduction moment from before to after surgery in patients who underwent medial open wedge HTO were included. Results Nine studies were included in the meta-analysis. The pooled mean difference in adduction moment from before to after medial open wedge HTO was 1.44% Nm/body weight (BW)xheight (HT) (95% confidence interval [CI]: 1.33 to 1.55% Nm/BWxHT; P

Background and Objectives: The TomoFix anatomical plate was developed to improve plate position, proximal screw direction, and post-correction tibial contouring. The purpose of this study was to compare postoperative configurations between the TomoFix anatomical plate and the TomoFix conventional plate. It was hypothesized that the new modified plate provides a better fixative coaptation than the conventional plate. Materials and Methods: A total of 116 cases (112 patients) were enrolled in this study from March 2015 to February 2021. Among them, 63 patients underwent surgery using the TomoFix conventional plate, and 53 underwent surgery using the TomoFix anatomical plate. The radiographic outcomes, including the hip–knee–ankle (HKA) angle, medial proximal tibial angle (MPTA), tibial slope, plate angle, proximal screw angles, and plate-to-cortex distance at #1 hole (just below the osteotomy site) were compared between the two groups. Results: Patients with the TomoFix anatomical plate showed similar results in terms of the pre- and postoperative HKA angle, MPTA, and tibial slope. The TomoFix anatomical group showed a significantly greater plate angle (39.2° ± 8.1° vs. 31.7° ± 7.0°, p < 0.001) and less screw angles, indicating that the TomoFix anatomical plates allowed a more posterior plate position than the conventional plate. The plate-to-cortex distance was significantly less in the TomoFix anatomical group than in the TomoFix conventional group (p < 0.001). Conclusion: The TomoFix anatomical plate showed a more posteromedial plating position, better proximal screw direction to the lateral hinge, and improved post-correction tibial contour compared to the TomoFix conventional plate.

Background/Aims Nilotinib is used for treating patients with imatinib-sensitive or -resistant chronic myeloid leukemia (CML); however, nilotinib-resistant cases have been observed in recent years. In addition, a considerable number of patients receiving nilotinib developed diabetes. Metformin is a front-line drug for the treatment of type 2 diabetes, and several studies have shown that diabetes patients treated with metformin have reduced incidence of cancer. This study aimed to define the effect of metformin on CML cells to determine whether metformin overcomes nilotinib resistance, and to identify novel targets for the treatment of nilotinib resistance. Methods We observed the effects of metformin and nilotinib on K562 and KU812 human CML cell lines. Nilotinib-resistant CML cell lines were generated by exposing cells to gradually increasing doses of nilotinib. Then, we investigated the driving force that makes resistance to nilotinib and the effect of metformin on the driving force. Results Sub-toxic doses of metformin enhanced nilotinib efficacy by reducing Bcl-xL expression, which induces apoptosis in CML cells. Next, we generated nilotinib-resistant K562 and KU812 cell lines that overexpressed the c-Jun N-terminal kinase (JNK) gene. JNK silencing by a JNK inhibitor restored sensitivity to nilotinib. Furthermore, metformin was effective in decreasing phosphorylated JNK levels, restoring nilotinib sensitivity. Combined treatment with nilotinib and metformin was more effective than combined treatment with nilotinib and a JNK inhibitor in terms of cell proliferation inhibition. Conclusions This study suggested that combination therapy with metformin and nilotinib may have clinical benefits of enhancing antileukemia efficacy and overcoming resistance to nilotinib.

Genetic circuits have been developed for quantitative measurement of enzyme activity, metabolic engineering of strain development, and dynamic regulation of microbial cells. A genetic circuit consists of several bio-elements, including enzymes and regulatory cassettes, that can generate the desired output signal, which is then used as a precise criterion for enzyme screening and engineering. Antagonists and inhibitors are small molecules with inhibitory effects on regulators and enzymes, respectively. In this study, an antagonist and an inhibitor were applied to a genetic circuit for a dynamic detection range. We developed a genetic circuit relying on regulators and enzymes, allowing for straightforward control of its output signal without additional genetic modification. We used para-nitrophenol and alanine as an antagonist of DmpR and inhibitor of tyrosine phenol-lyase, respectively. We show that the antagonist resets the detection range of the genetic circuit similarly to a resistor in an electrical logic circuit. These biological resistors in genetic circuits can be used as a rapid and precise controller of variable outputs with minimal circuit configuration.

Methanotrophs with soluble methane monooxygenase (sMMO) show high potential for various ecological and biotechnological applications. Here, we developed a high throughput method to identify sMMO-producing microbes by integrating droplet microfluidics and a genetic circuit-based biosensor system. sMMO-producers and sensor cells were encapsulated in monodispersed droplets with benzene as the substrate and incubated for 5 h. The sensor cells were analyzed as the reporter for phenol-sensitive transcription activation of fluorescence. Various combinations of methanotrophs and biosensor cells were investigated to optimize the performance of our droplet-integrated transcriptional factor biosensor system. As a result, the conditions to ensure sMMO activity to convert the starting material, benzene, into phenol, were determined. The biosensor signals were sensitive and quantitative under optimal conditions, showing that phenol is metabolically stable within both cell species and accumulates in picoliter-sized droplets, and the biosensor cells are healthy enough to respond quantitatively to the phenol produced. These results show that our system would be useful for rapid evaluation of phenotypes of methanotrophs showing sMMO activity, while minimizing the necessity of time-consuming cultivation and enzyme preparation, which are required for conventional analysis of sMMO activity.

Bioconversion of C1 chemicals such as methane and methanol into higher carbon-chain chemicals has been widely studied. Methanol oxidation catalyzed by methanol dehydrogenase (Mdh) is one of the key steps in methanol utilization in bacterial methylotrophy. In bacteria, few NAD+-dependent Mdhs have been reported that convert methanol to formaldehyde. In this study, an uncharacterized Mdh gene from Lysinibacillus xylanilyticus (Lxmdh) was cloned and expressed in Escherichia coli. The maximum alcohol oxidation activity of the recombinant enzyme was observed at pH 9.5 and 55°C in the presence of 10 mM Mg2+. To improve oxidation activity, rational approach-based, site-directed mutagenesis of 16 residues in the putative active site and NAD+-binding region was performed. The mutations S101V, T141S, and A164F improved the enzyme’s specific activity toward methanol compared to that of the wild-type enzyme. These mutants show a slightly higher turnover rate than that of wild-type, although their KM values were increased compared to that of wild-type. Consequently, according the kinetic results, S101, T141, and A164 positions may related to the catalytic activity in the active site for methanol dehydrogenation. It should be further studied other mutant variants with high activity for methanol. In conclusion, we characterized a new Lxmdh and its variants that may be potentially useful for the development of synthetic methylotrophy in the future.

PURPOSE:Although rotation of the distal portion of the tibia below the osteotomy site is considered an inevitable change in the axial plane in open wedge high tibial osteotomy (HTO), several studies on this issue have shown contradictory results. The purpose of this study was, therefore, to determine the direction and amount of distal tibial rotation following open wedge HTO using a three-dimensional reconstructed model. METHODS:This study involved 41 patients (42 knees) undergoing open wedge HTO for primary medial osteoarthritis. Distal tibial rotation was measured on the overlaid tibial plateau of a preoperative and postoperative 3-dimensional reconstructed model based on computed tomography. RESULTS:The mean distal tibial external rotation was 2.7° ± 2.3° (range, -0.9° to 9.9°), and the opening gap was larger in the group with > 3° distal tibial rotation than in the group with ≤ 3° distal tibial rotation (11.4 mm vs. 9.6 mm, P = 0.027). Multiple regression analysis showed that the opening gap was the only predictor of distal tibial rotation. On receiver operating characteristics analysis, an opening gap of 10 mm was found to be the optimal cutoff value for achieving greater than 3° of distal tibial rotation. CONCLUSIONS:Following medial opening wedge HTO, the distal tibial portion below the osteotomy site rotated approximately 3° externally. The magnitude of the external rotation of the distal tibia was affected by the opening gap width.

Efficient biosynthesis of lactams is still undesirable due to lacking of suitable enzyme. Here, the authors develop a sensitive transcription factor-based biosensor for high-throughput screening of marine metagenome and find a cyclase that can cyclize ω-amino fatty acids to lactam.

Purpose: (1) To compare postoperative range of motion (ROM), stability, and clinical outcomes between fixed-bearing posterior-stabilized (PS) and ultracongruent (UC). (2) The effect of postoperative stability on ROM and clinical outcomes was also evaluated in both designs. Materials and methods: Propensity score matching was conducted for age, gender, body mass index, preoperative ROM, Western Ontario and McMaster Universities Osteoarthritis (WOMAC) index, Knee Society (KS) scores, hip–knee–ankle (HKA) alignment, and follow-up period. Two hundred patients (100 PS and 100 UC) were enrolled. Preoperative and final follow-up outcomes including postoperative ROM, anteroposterior (AP) stability (good, fair, and poor), WOMAC index, and KS scores were compared. Then, postoperative outcomes compared between the PS and UC. We also analyzed if AP stability was associated with the postoperative outcomes in both implant designs. Results: In both groups, ROM and clinical outcomes of final follow-up showed improvement than preoperation. Statistical significance was not determined between the PS and UC groups in terms of postoperative ROM (PS vs. UC, 134.6° vs. 133.4°, p = 0.13), stability (good/fair/poor, 91/9/0 vs. 84/14/0, p = 0.376), WOMAC index, KS scores, and outliers of HKA alignment (15% vs. 10%, p = 0.393). “Fair” stability showed inferior KS scores but greater ROM than “good” stability in both designs. Conclusion: TKA with UC insert provided similar ROM, AP stability, and clinical outcomes when compared to PS insert. In both designs, greater postoperative ROM was found but inferior clinical outcomes were found when TKA resulted in fair stability instead of good stability. Level of evidence III: Retrospective comparative study.

Abstract Background Isoprene is a five-carbon chemical that is an important starting material for the synthesis of rubber, elastomers, and medicines. Although many plants produce huge amounts of isoprene, it is very difficult to obtain isoprene directly from plants because of its high volatility and increasing environmental regulations. Over the last decade, microorganisms have emerged as a promising alternative host for efficient and sustainable bioisoprene production. Isoprene synthase (IspS) has received much attention for the conversion of isoprene from dimethylallyl diphosphate (DMAPP). Herein, we isolated a highly expressible novel IspS gene from Metrosideros polymorpha (MpIspS), which was cloned and expressed in Escherichia coli, using a plant cDNA library and characterized its molecular and biochemical properties. Results The signal sequence deleted MpIspS was cloned and expressed in E. coli as a 65-kDa monomer. The maximal activity of the purified MpIspS was observed at pH 6.0 and 55 °C in the presence of 5 mM Mn2+. The K m, k cat, and k cat/K m for DMAPP as a substrate were 8.11 mM, 21 min− 1, and 2.59 mM− 1 min− 1, respectively. MpIspS was expressed along with the exogenous mevalonate pathway to produce isoprene in E. coli. The engineered cells produced isoprene concentrations of up to 23.3 mg/L using glycerol as the main carbon source. Conclusion MpIspS was expressed in large amounts in E. coli, which led to increased enzymatic activity and resulted in isoprene production in vivo. These results demonstrate a new IspS enzyme that is useful as a key biocatalyst for bioisoprene production in engineered microbes.

Swift and continuous phagocytosis of apoptotic cells can be achieved by modulation of calcium flux in phagocytes. However, the molecular mechanism by which apoptotic cells modulate calcium flux in phagocytes is incompletely understood. Here, using biophysical, biochemical, pharmaceutical, and genetic approaches, we show that apoptotic cells induced the Orai1-STIM1 interaction, leading to store-operated calcium entry (SOCE) in phagocytes through the Mertk-phospholipase C (PLC) γ1-inositol 1,4,5-triphosphate receptor (IP3R) axis. Apoptotic cells induced calcium release from the endoplasmic reticulum, which led to the Orai1-STIM1 association and, consequently, SOCE in phagocytes. This association was attenuated by masking phosphatidylserine. In addition, the depletion of Mertk, which indirectly senses phosphatidylserine on apoptotic cells, reduced the phosphorylation levels of PLCγ1 and IP3R, resulting in attenuation of the Orai1-STIM1 interaction and inefficient SOCE upon apoptotic cell stimulation. Taken together, our observations uncover the mechanism of how phagocytes engulfing apoptotic cells elevate the calcium level.

The clearance of apoptotic cells is known to be a critical step in maintaining tissue and organism homeostasis. This process is rapidly/promptly mediated by recruited or resident phagocytes. Phagocytes that engulf apoptotic cells have been closely linked to the release of anti-inflammatory cytokines to eliminate inflammatory responses. Defective clearance of apoptotic cells can cause severe inflammation and autoimmune responses due to secondary necrosis of apoptotic cells. Recently accumulated evidence indicates that apoptotic cells and their clearance have important physiological roles in addition to immune-related functions. Herein, we review the current understanding of the mechanisms and fundamental roles of apoptotic cell clearance and the beneficial roles of apoptotic cells in physiological processes such as differentiation and development.

ε-Polylysine (ε-PL) is a safe food additive that is used in the food industry globally. This study evaluated the antimicrobial and antibiofilm activity of antibacterial peptides (ε-PL) against food poisoning pathogens detected in chicken (Salmonella Enteritidis, Listeria monocytogenes, and Escherichia coli). The results showed that minimum inhibitory concentrations (MICs) ranged between 0.031–1.0 mg/mL, although most bacterial groups (75%) showed MICs of 1.0 mg/mL. The reduction in the cell viability of pathogens due to ε-PL depended on the time and concentration, and 1/2 × MIC of ε-PL killed 99.99% of pathogens after 10 h of incubation. To confirm biofilm inhibition and degradation effects, crystal violet assay and confocal laser scanning microscopy (CLSM) were used. The biofilm formation rates of four bacterial groups (Salmonella, Listeria, E. coli, and multi-species bacteria) were 10.36%, 9.10%, 17.44%, and 21.37% at 1/2 × MIC of ε-PL, respectively. Additionally, when observed under a CLSM, ε-PL was found to induce biofilm destruction and bacterial cytotoxicity. These results demonstrated that ε-PL has the potential to be used as an antibiotic and antibiofilm material for chicken meat processing.

The distribution of differential extracellular matrix (ECM) in the lateral and medial menisci can contribute to knee instability, and changes in the meniscus tissue can lead to joint disease. Thus, deep proteomic identification of the lateral and medial meniscus cartilage is expected to provide important information for treatment and diagnosis of various knee joint diseases. We investigated the proteomic profiles of 12 lateral/medial meniscus pairs obtained from excess tissue of osteoarthritis patients who underwent knee arthroscopy surgery using mass spectrometry-based techniques and measured 75 ECM protein levels in the lesions using a multiple reaction monitoring (MRM) assay we developed. A total of 906 meniscus proteins with a 1% false discovery rate (FDR) was identified through a tandem mass tag (TMT) analysis showing that the lateral and medial menisci had similar protein expression profiles. A total of 131 ECM-related proteins was included in meniscus tissues such as collagen, fibronectin, and laminin. Our data showed that 14 ECM protein levels were differentially expressed in lateral and medial lesions (p < 0.05). We present the proteomic characterization of meniscal tissue with mass spectrometry-based comparative proteomic analysis and developed an MRM-based assay of ECM proteins correlated with tissue regeneration. The mass spectrometry dataset has been deposited to the MassIVE repository with the dataset identifier MSV000087753.

Abstract Background Multiplex control of metabolic pathway genes is essential for maximizing product titers and conversion yields of fuels, chemicals, and pharmaceuticals in metabolic engineering. To achieve this goal, artificial transcriptional regulators, such as clustered regularly interspaced short palindromic repeats (CRISPR) interference (CRISPRi), have been developed to specifically repress genes of interest. Results In this study, we deployed a tunable CRISPRi system for multiplex repression of competing pathway genes and, thus, directed carbon flux toward production of molecules of interest in Escherichia coli. The tunable CRISPRi system with an array of sgRNAs successfully repressed four endogenous genes (pta, frdA, ldhA, and adhE) individually and in double, triple, or quadruple combination that are involved in the formation of byproducts (acetate, succinate, lactate, and ethanol) and the consumption of NADH in E. coli. Single-target CRISPRi effectively reduced the amount of each byproduct and, interestingly, pta repression also decreased ethanol production (41%), whereas ldhA repression increased ethanol production (197%). CRISPRi-mediated multiplex repression of competing pathway genes also resulted in simultaneous reductions of acetate, succinate, lactate, and ethanol production in E. coli. Among 15 conditions repressing byproduct-formation genes, we chose the quadruple-target CRISPRi condition to produce n-butanol in E. coli as a case study. When heterologous n-butanol-pathway enzymes were introduced into E. coli simultaneously repressing the expression of the pta, frdA, ldhA, and adhE genes via CRISPRi, n-butanol yield and productivity increased up to 5.4- and 3.2-fold, respectively. Conclusions We demonstrated the tunable CRISPRi system to be a robust platform for multiplex modulation of endogenous gene expression that can be used to enhance biosynthetic pathway productivity, with n-butanol as the test case. CRISPRi applications potentially enable the development of microbial “smart cell” factories capable of producing other industrially valuable products.

Abstract This meta-analysis was designed to compare the longevity of the survivorship of opening versus closing wedge high tibial osteotomy (HTO). All studies reporting survival rates in patients who underwent open or closed wedge HTO with more than 5-year follow-up duration were included in the meta-analysis. Survival time was considered as time to conversion to TKA. Twenty three studies were included in meta-analysis, 20 of which were of level IV evidence. The pooled 5-year survival rates were 95.1% (95% CI: 93.1 to 97.1%) in open wedge HTO and 93.9% (95% CI: 93.1 to 94.6%) in closed wedge HTO. Although there was 1.2% greater survival rate in open wedge HTO than in closed wedge HTO, this difference did not reach statistical significance (P = 0.419). Pooled 10-year survival rates were 91.6% (95% CI: 88.5 to 94.8%) in open wedge HTO and 85.4% (95% CI: 84.0 to 86.7%) in closed wedge HTO, indicating that open wedge HTO had 6.2% greater survival rate 10 years after surgery than did closed wedge HTO (P = 0.002). No difference in 5-year survivorship was found between open- and closed-wedge HTO. However, the survival rate was higher in open-wedge HTOs than in closed wedge HTO at 10 years.

Lysozyme is widely used as a model protein in studies of structure–function relationships. Recently, lysozyme has gained attention for use in accelerating the degradation of secondary sludge, which mainly consists of bacteria. However, a high-throughput screening system for lysozyme engineering has not been reported. Here, we present a lysozyme screening system using a genetically encoded biosensor. We first cloned bacteriophage T4 lysozyme (T4L) into a plasmid under control of the araBAD promoter. The plasmid was expressed in Escherichia coli with no toxic effects on growth. Next, we observed that increased soluble T4L expression decreased the fluorescence produced by the genetic enzyme screening system. To investigate T4L evolution based on this finding, we generated a T4L random mutation library, which was screened using the genetic enzyme screening system. Finally, we identified two T4L variants showing 1.4-fold enhanced lytic activity compared to native T4L. To our knowledge, this is the first report describing the use of a genetically encoded biosensor to investigate bacteriophage T4L evolution. Our approach can be used to investigate the evolution of other lysozymes, which will expand the applications of lysozyme.

Previously, we demonstrated that γ-ionizing radiation (IR) triggers the invasion/migration of A549 cells via activation of an EGFR–p38/ERK–STAT3/CREB-1–EMT pathway. Here, we have demonstrated the involvement of a novel intracellular signaling mechanism in γ-ionizing radiation (IR)-induced migration/invasion. Expression of receptor-interacting protein (RIP) 1 was initially increased upon exposure of A549, a non-small cell lung cancer (NSCLC) cell line, to IR. IR-induced RIP1 is located downstream of EGFR and involved in the expression/activity of matrix metalloproteases (MMP-2 and MMP-9) and vimentin, suggesting a role in epithelial-mesenchymal transition (EMT). Our experiments showed that IR-induced RIP1 sequentially induces Src-STAT3-EMT to promote invasion/migration. Inhibition of RIP1 kinase activity and expression blocked induction of EMT by IR and suppressed the levels and activities of MMP-2, MMP-9 and vimentin. IR-induced RIP1 activation was additionally associated with stimulation of the transcriptional factor NF-κB. Specifically, exposure to IR triggered NF-κB activation and inhibition of NF-κB suppressed IR-induced RIP1 expression, followed by a decrease in invasion/migration as well as EMT. Based on the collective results, we propose that IR concomitantly activates EGFR and NF-κB and subsequently triggers the RIP1–Src/STAT3–EMT pathway, ultimately promoting metastasis.

A large proportion of the recombinant proteins manufactured today rely on microbe-based expression systems owing to their relatively simple and cost-effective production schemes. However, several issues in microbial protein expression, including formation of insoluble aggregates, low protein yield, and cell death are still highly recursive and tricky to optimize. These obstacles are usually rooted in the metabolic capacity of the expression host, limitation of cellular translational machineries, or genetic instability. To this end, several microbial strains having precisely designed genomes have been suggested as a way around the recurrent problems in recombinant protein expression. Already, a growing number of prokaryotic chassis strains have been genome-streamlined to attain superior cellular fitness, recombinant protein yield, and stability of the exogenous expression pathways. In this review, we outline challenges associated with heterologous protein expression, some examples of microbial chassis engineered for the production of recombinant proteins, and emerging tools to optimize the expression of heterologous proteins. In particular, we discuss the synthetic biology approaches to design and build and test genome-reduced microbial chassis that carry desirable characteristics for heterologous protein expression.

BACKGROUND:Achieving proper rotational alignment of the femoral component in total knee arthroplasty (TKA) for valgus knee is challenging because of lateral condylar hypoplasia and lateral cartilage erosion. Gap-based navigation-assisted TKA enables surgeons to determine the angle of femoral component rotation (FCR) based on the posterior condylar axis. This study evaluated the possible factors that affect the rotational alignment of the femoral component based on the posterior condylar axis. MATERIALS AND METHODS:Between 2008 and 2016, 28 knees were enrolled. The dependent variable for this study was FCR based on the posterior condylar axis, which was obtained from the navigation system archives. Multiple regression analysis was conducted to identify factors that might predict FCR, including body mass index (BMI), Kellgren-Lawrence grade (K-L grade), lateral distal femoral angles obtained from the navigation system and radiographs (NaviLDFA, XrayLDFA), hip-knee-ankle (HKA) axis, lateral gap under varus stress (LGVS), medial gap under valgus stress (MGVS), and side-to-side difference (STSD, MGVS - LGVS). RESULTS:The mean FCR was 6.1° ± 2.0°. Of all the potentially predictive factors evaluated in this study, only NaviLDFA (β = -0.668) and XrayLDFA (β = -0.714) predicted significantly FCR. CONCLUSIONS:The LDFAs, as determined using radiographs and the navigation system, were both predictive of the rotational alignment of the femoral component based on the posterior condylar axis in gap-based TKA for valgus knee. A 1° increment with NaviLDFA led to a 0.668° decrement in FCR, and a 1° increment with XrayLDFA led to a 0.714° decrement. This suggests that symmetrical lateral condylar hypoplasia of the posterior and distal side occurs in lateral compartment end-stage osteoarthritis with valgus deformity.

Metformin, a first-line drug used to treat type 2 diabetes, has also been shown to have anticancer effects against a variety of malignancies, including colorectal cancer. Although inhibition of the mTOR pathway is known to be the most important mechanism for the antitumor effects of metformin, other mechanisms remain unclear. The purpose of this study was to identify the antitumor mechanism of metformin in colorectal cancer using high-throughput data, and then test the mechanism experimentally. We identified the gene signature of metformin-treated colon cancer cells. This signature was processed for prediction using colon adenocarcinoma patient data from the Cancer Genome Atlas to classify the patients showing a gene expression pattern similar to that in metformin-treated cells. This patient group showed better overall and disease-free survival. Furthermore, pathway analysis revealed that the metformin-predicted group was characterized by decreased interleukin (IL)-6 pathway signaling, epithelial-mesenchymal transition, and colon cancer metastatic signaling. We induced epithelial-mesenchymal transition in colon cancer cell lines via IL-6 treatment, which increased cell motility and promoted invasion. However, these effects were blocked by metformin. These findings suggest that blockade of IL-6-induced epithelial-mesenchymal transition is an antitumor mechanism of metformin.

A newly designed Tissue Equivalent Proportional Counter (TEPC) has been developed for the CubeSat mission, SIGMA (Scientific cubesat with Instruments for Global Magnetic field and rAdiation) to investigate space radiation. In order to test the performance of the TEPC, we have performed heavy ion beam experiments with the Heavy Ion Medical Accelerator in Chiba (HIMAC), Japan. In space, human cells can be exposed to complex radiation sources, such as X-ray, Gamma ray, energetic electrons, protons, neutrons and heavy charged particles in a huge range of energies. These generate much a larger range of Linear Energy Transfer (LET) than on the ground and cause unexpected effects on human cells. In order to measure a large range of LET, from 0.3 to 1,000 keV/μm, we developed a compact TEPC which measures ionized particles produced by collisions between radiation sources and tissue equivalent materials in the detector. By measuring LET spectra, we can easily derive the equivalent dose from the complicated space radiation field. In this HIMAC experiment, we successfully obtained the linearity response for the TEPC with Fe 500 MeV/u and C 290 MeV/u beams and demonstrated the performance of the active radiation detector.

The sonic hedgehog signaling pathway is critical for cell proliferation, cell differentiation, and organ development during embryogenesis. Aberrant activation of the sonic hedgehog pathway has been associated with a variety of human cancers. Currently, there is no target molecular drug that clinically affects sonic hedgehog activation in patients with gastric cancer. In this review, we will focus on the current clinical treatment options for advanced gastric cancer and discuss the sonic hedgehog signaling pathway. We also review our understanding of the role of sonic hedgehog signaling in advanced gastric cancer progression. Finally, we will describe current known molecular pathways that crosstalk with the sonic hedgehog pathway in cancer stem cells.

Posterior cruciate ligament (PCL) reconstruction for patients with PCL insufficiency has been associated with postoperative improvements in proprioceptive function due to mechanoreceptor regeneration. However, it is unclear whether reconstructed PCL or contralateral normal knees have better proprioceptive function outcomes. This meta-analysis was designed to compare the proprioceptive function of reconstructed PCL or contralateral normal knees in patients with PCL insufficiency. All studies that compared proprioceptive function, as assessed with threshold to detect passive movement (TTDPM) or joint position sense (JPS) in PCL reconstructed or contralateral normal knees were included. JPS was calculated by reproducing passive positioning (RPP). Five studies met the inclusion/exclusion criteria for the meta-analysis. The proprioceptive function, defined as TTDPM (95% CI: 0.25 to 0.51°; P1°) of the loss of proprioception are clinically relevant.

Reactive oxygen species modulator-1 (Romo1) is a novel protein that has been reported to be crucial for cancer cell proliferation and invasion. However, its clinical implications in colorectal cancer patients are not well-known. For the first time, we investigated the association between Romo1 and the clinical outcomes of colorectal cancer patients.We examined Romo1 expression in resected tumor tissues immunohistochemically and assessed it with histological scores. We conducted survival analyses for patients who had curative resection (n = 190) in accordance with clinical parameters including level of Romo1 expression, and we examined the association between Romo1 expression and cell invasion using Matrigel invasion assay in colorectal cancer cells.We observed significantly longer mean disease-free survival in the low Romo1 group compared with the high Romo1 group (161 vs 127.6 months, p = 0.035), and the median overall survival of the low Romo1 group was significantly longer than that of the high Romo1 group (196.9 vs 171.3 months, p = 0.036). Cell invasiveness decreased in the Romo1 knockdown colorectal cancer cells in contrast to the controlled cells. Romo1 overexpression in tumor tissue was associated with a high lymph node ratio between the metastatic and examined lymph nodes (p = 0.025).Romo1 overexpression in tumor tissue was significantly associated with survival in curatively resected colorectal cancer patients, suggesting Romo1 expression as a potential adverse prognostic marker. Increased Romo1 expression was found to be associated with high lymph node ratio. Cancer invasiveness appeared to be a key reason for the poor survival related to highly expressed Romo1.

Cellulose-binding domains (CBDs) are protein domains with cellulose-binding activity, and some act as leaders in the localization of cellulosomal scaffoldin proteins to the hydrophobic surface of crystalline cellulose. In this study, we found that a CBD fusion enhanced and improved soluble β-glucuronidase (GusA) enzyme properties through the formation of an artificially oligomeric state. First, a soluble CBD fused to the C-terminus of GusA (GusA-CBD) was obtained and characterized. Interestingly, the soluble GusA-CBD showed maximum activity at higher temperatures (65°C) and more acidic pH values (pH 6.0) than free GusA did (60°C and pH 7.5). Moreover, the GusA-CBD enzyme showed higher thermal and pH stabilities than the free GusA enzyme did. Additionally, GusA-CBD showed higher enzymatic activity in the presence of methanol than free GusA did. Evaluation of the protease accessibility of both enzymes revealed that GusA-CBD retained 100% of its activity after 1 h incubation in 0.5 mg/ml protease K, while free GusA completely lost its activity. Simple fusion of CBD as a single domain may be useful for tunable enzyme states to improve enzyme stability in industrial applications.

Theoretical considerations suggest that leg length increases after opening wedge high tibial osteotomy (HTO) and decreases after closing wedge HTO; however, in vivo studies have yielded conflicting results. This meta-analysis therefore assessed changes in leg length after opening wedge and closing wedge HTO.All studies comparing pre- and postoperative leg length in patients who underwent opening and/or closing wedge HTO were included. Two reviewers independently recorded data from each study in terms of sample size as well as preoperative and postoperative leg length of open wedge and/or closed wedge HTO groups.Four studies were included in the meta-analysis. Although pooled results showed leg length changes from before to after surgery were -6.93 mm (95% confidence interval [CI]: -17.53 to 3.67 mm; P = 0.20) in opening wedge HTO and 1.97 mm (95% CI: -7.13 to 11.07 mm; P = 0.67) in closing wedge HTO, respectively, these values were statistically not significant. However, the difference in the pooled mean leg length change from before to after surgery between opening wedge and closing wedge HTO was 8 mm, a difference that was significant (95% CI: 6.53 to 9.46 mm; P

The communications link in a space program is a crucial point for upgrading its performance by handling data between spacecraft bus and payloads, because spacecraft’s missions are related to the data handling mechanism using communications ports such as a controlled area network bus (CAN Bus) and a universal asynchronous receiver and transmitter (UART). The NEXTSat-1 has a lot of communications ports for performing science and technology missions. However, the top level system requirements for the NEXTSat-1 are mass and volume limitations. Normally, the communications for units shall be conducted by using point to point link which require more mass and volume to interconnect. Thus, our approach for the novel communications link in the NEXTSat-1 program is to use CAN and serializer and deserializer low voltage differential signal (SerDesLVDS) to meet the system requirements of mass and volume. The CAN Bus and SerDesLVDS were confirmed by using already defined communications link for our missions in the NEXTSat-1 program and the analysis results were reported in this study in view of data flow and size analysis.

The NISS onboard NEXTSat-1 is being developed by Korea astronomy and space science institute (KASI). For the study of the cosmic star formation history, the NISS performs the imaging spectroscopic observation in the near-infrared range for nearby galaxies, low background regions, star-forming regions and so on. It is designed to cover a wide field of view (2 × 2 deg) and a wide wavelength range from 0.95 to 3.8 μm by using linear variable filters. In order to reduce the thermal noise, the telescope and the infrared sensor are cooled down to 200 K and 80 K, respectively. Evading a stray light outside the field of view and making the most use of limited space, the NISS adopts the off-axis reflective optical system. The primary and the secondary mirrors, the opto-mechanical part and the mechanical structure are designed to be made of aluminum material. It reduces the degradation of optical performance due to a thermal variation. This paper presents the study on the conceptual design of the NISS.

Anaerobic digestion (AD) of secondary sludge is a rate-limiting step due to the bacterial cell wall. In this study, experiments were performed to characterize secondary sludges from three wastewater treatment plants (WWTPs), and to investigate the feasibility of using bacteriophage lysozymes to speed up AD by accelerating the degradation of bacterial cell walls. Protein was the main organic material (67.7% of volatile solids in the sludge). The bacteriophage T4 lysozyme (T4L) was tested for hydrolysis and biochemical methane potential. Variations in the volatile suspended solid (VSS) concentration and biogas production were monitored. The VSS reduction efficiencies by hydrolysis using T4L for 72 h increased and ranged from 17.8% to 26.4%. Biogas production using T4L treated sludges increased and biogas production was increased by as much as 82.4%. Biogas production rate also increased, and the average reaction rate coefficient of first-order kinetics was 0.56 ± 0.02/d, which was up to 47.5% higher compared to the untreated samples at the maximum. Alphaproteobacteria, Betaproteobacteria, Flavobacteriia, Gammaproteobacteria, and Sphingobacteriia were major microbial classes in all sludges. The interpretation of the microbial community structure indicated that T4L treatment is likely to increase the rate of cell wall digestion.

Background: Although the anticancer activity of Korean Red Ginseng (KRG) has been known in various cancers, the mechanism of KRG-induced apoptosis is unknown in colorectal cancer (CRC). In our study, we examined whether KRG induces apoptosis in CRC cells. Methods: In the cell viability assay, the concentration of the appropriate KRG extracts was fixed at 2.5 mg/mL in numerous CRC cells. This fixed concentration was in other experiments, and it was confirmed that the KRG extracts induce apoptosis in CRC cells. Results: We found that KRG induced Noxa activation and apoptosis and increased endoplasmic reticulum stress via reactive oxygen species production. This indicated that KRG efficiently enhanced cell death in CRC cells. Conclusion: Our results show that KRG can be used as a possible anticancer drug for patients with CRC

Circulating tumor cells (CTCs) have attracted increasing attention as important biomarkers for clinical and biological applications. Several microfluidic approaches have been demonstrated to separate CTCs using immunoaffinity or size difference from other blood cells. This study demonstrates a sheathless, high-throughput separation of CTCs from white blood cells (WBCs) using a viscoelastic fluid. To determine the fluid viscoelasticity and the flow rate for CTC separation, and to validate the device performance, flow characteristics of 6, 13, and 27 μm particles in viscoelastic fluids with various concentrations were estimated at different flow rates. Using 0.2% hyaluronic acid (HA) solution, MCF-7 (Michigan Cancer Foundation-7) cells mimicking CTCs in this study were successfully separated from WBCs at 500 μL/min with a separation efficiency of 94.8%. Small amounts of MCF-7 cells (~5.2%) were found at the center outlet due to the size overlap with WBCs.

The microbial assimilation of one-carbon (C1) gases is a topic of interest, given that products developed using this pathway have the potential to act as promising substrates for the synthesis of valuable chemicals via enzymatic oxidation or C−C bonding. Despite extensive studies on C1 gas assimilation pathways, their key enzymes have yet to be subjected to high-throughput evolution studies on account of the lack of an efficient analytical tool for C1 metabolites. To address this challenging issue, we attempted to establish a fine-tuned single-cell−level biosensor system constituting a combination of transcription factors (TFs) and several C1-converting enzymes that convert target compounds to the ligand of a TF. This enzymatic conversion broadens the detection range of ligands by the genetic biosensor systems. In this study, we presented new genetic enzyme screening systems (GESSs) to detect formate, formaldehyde, and methanol from specific enzyme activities and pathways, named FA-GESS, Frm-GESS, and MeOH-GESS, respectively. All the biosensors displayed linear responses to their respective C1 molecules, namely, formate (1.0−250 mM), formaldehyde (1.0−50 μM), and methanol (5−400 mM), and they did so with high specificity. Consequently, the helper enzymes, including formaldehyde dehydrogenase and methanol dehydrogenase, were successfully combined to constitute new versatile combinations of the C1-biosensors.

(−)-α-Bisabolol, a naturally occurring sesquiterpene alcohol, has been used in pharmaceuticals and cosmetics owing to its beneficial effects on inflammation and skin healing. Previously, we reported the high production of (−)-α-bisabolol by fed-batch fermentation using engineered Escherichia coli (E. coli) expressing the exogenous mevalonate (MVA) pathway genes. The productivity of (−)-α-bisabolol must be improved before industrial application. Here, we report enhancement of initial (−)-α-bisabolol productivity to 3-fold higher than that observed in our previous study. We first harnessed a farnesyl pyrophosphate (FPP)-resistant mevalonate kinase 1 (MvaK1) from an archaeon Methanosarcina mazei (M. mazei) to create a more efficient heterologous MVA pathway that produces (−)-α-bisabolol in the engineered E. coli. The resulting strain produced 1.7-fold higher (−)-α-bisabolol relative to the strain expressing a feedback-inhibitory MvaK1 from Staphylococcus aureus (S. aureus). Next, to efficiently convert accumulated MVA to (−)-α-bisabolol, we additionally overexpressed genes involved in the lower MVA mevalonate pathway in E. coli containing the entire MVA pathway genes. (−)-α-Bisabolol production increased by 1.8-fold with reduction of MVA accumulation, relative to the control strain. Finally, we optimized the fermentation conditions including inducer concentration, aeration and enzymatic cofactor. The strain was able to produce 8.5 g/L of (−)-α-bisabolol with an initial productivity of 0.12 g/L h in the optimal fed-batch fermentation. Thus, the microbial production of (−)-α-bisabolol would be an economically viable bioprocess for its industrial application.

Natural products have shown great promise in sensitizing cells to TNF-related apoptosis-inducing ligand (TRAIL) therapy. Sea cucumber (SC) extracts possess antitumor activity, and hence their potential to sensitize colorectal cancer (CRC) cells to TRAIL therapy was evaluated. This study used Western blotting to evaluate the combination effects of SC and TRAIL in CRC, and determined the molecular mechanism underlying these effects. SC fractions and TRAIL alone did not affect apoptosis; however, combined treatment dramatically induced the apoptosis of CRC cells, but not of normal colon cells. Combined treatment induced the expression of apoptotic proteins (poly (ADP-ribose) polymerase (PARP), caspase 3, and 8), and this effect was markedly inhibited by the ubiquitination of X-linked inhibitor of apoptosis protein (XIAP). SC did not affect the mRNA levels, but it increased proteasomal degradation and ubiquitination of the XIAP protein. Furthermore, SC induced reactive oxygen species (ROS) production, thereby activating c-Jun N-terminal kinase (JNK) and endoplasmic reticulum (ER) stress-related apoptotic pathways in CRC. Altogether, our results demonstrate that the SC F2 fraction may sensitize CRC cells to TRAIL-induced apoptosis through XIAP ubiquitination and ER stress.

Tissue equivalent proportional counter (TEPC) can measure the Linear Energy Transfer (LET) spectrum and calculate the equivalent dose for the complicated radiation field in space. In this paper, we developed and characterized a TEPC for radiation monitoring in International Space Station (ISS). The prototype TEPC which can simulate a 2 μm of the site diameter for micro-dosimetry has been tested with a standard alpha source (241Am, 5.5 MeV). Also, the calibration of the TEPC was performed by the 252Cf neutron standard source in Korea Research Institute of Standards and Science (KRISS). The determined calibration factor was kf = 3.59×10-7 mSv/R.

Plasmid-based gene expression is a fundamental tool in the field of biotechnology. However, overexpression of genes of interest with multi-copy plasmids often causes detrimental effects on host cells. To overcome this problem, chromosomal integration of target genes has been used for decades; however, insufficient protein expression occurred with this method. In this study, we developed a novel cloning and expression system named the chromosomal vector (ChroV) system, that has features of stable and high expression of target genes on the F' plasmid in the Escherichia coli JM109(DE3) strain. We used an RMT cluster (KCTC 11994BP) containing a silent cat gene from a previous study to clone a gene into the F' plasmid. The ChroV system was applied to clone two model targets, GFPuv and carotenoids gene clusters (4 kb), and successfully used to prove the inducible tightly regulated protein expression in the F' plasmid. In addition, we verified that the expression of heterologous genes in ChroV system maintained stably in the absence of antibiotics for 1 week, indicating ChroV system is applicable to antibiotics-free production of valuable proteins. This protocol can be widely applied to recombinant protein expression for antibiotics-free, stable, and genome-based expression, providing a new platform for recombinant protein synthesis in E. coli. Overall, our approach can be widely used for the economical and industrial production of proteins in E. coli.