Search

Recombinant protein drugs, which are typically produced by mammalian host cells, have been approved for the treatment of a range of diseases. Accordingly, systems for selecting recombinant cell lines with efficient protein expression and for testing the content of recombinant proteins in vivo are crucial to the large-scale production and application of protein-based therapeutic drugs. In this study, we designed three aptamer beacons to detect His-tag, a common label of recombinant proteins. We found that all three beacons could specifically and quantitatively measure the His-tagged recombinant proteins with a short reaction time. Among these three beacons, the 6H5-MU beacon had the highest sensitivity for His polypeptides with a detection limit of 250 ng/mL and the shortest detection time within 1 min. Furthermore, we established a rapid and highly effective recombinant cell line construction system, which could obtain monoclonal cell lines with high yields of target proteins within 21 days, by combining 6H5-MU with pSB, a novel plasmid composed of a Sleeping Beauty transposase and a transposon. Finally, 6H5-MU also discriminately tested the serum concentration of His-tagged recombinant proteins in vivo, with consistent results compared to enzyme-linked immunosorbent assay (ELISA). We thus established a rapid and high-throughput method for generating recombinant cell lines and in vivo monitoring of recombinant protein levels, thereby providing a new platform for the development and preparation of recombinant protein drugs. KEY POINTS: • The 6H5-MU aptamer beacon rapidly and accurately binds to His-tagged recombinant proteins. • A system for rapid and high-throughput generation of recombinant cell lines is established using 6H5-MU and pSB. • 6H5-MU allows in vivo monitoring of recombinant protein levels.

Phytohormone signalling pathways have an important role in defence against pathogens mediated by cell-surface pattern recognition receptors and intracellular nucleotide-binding leucine-rich repeat class immune receptors

Purpose Reduced physical activity is reported in the elderly, especially in institutional residents. Institutionalized older adults exhibit a high prevalence of frailty. In this work, we developed an artificial intelligence of things (AIoT)-based feedback assistive strengthening ergometer (AIFASE), for the physical strengthening of the elderly with intelligent assistance. Methods We conducted a 12-week intervention in a long-term care facility. In total, 16 participants (84.38 ± 6.0 years; 4 males and 12 females) were recruited with 1:1 randomization of exercise to control groups. The muscle strength of the lower extremities, timed up and go test (TUG), and Short-form Physical Performance Battery (SPPB) of the participants were measured. The AIFASE system allows the clinical staff to record the personal physical performance of the elderly and generates personalized exercise prescriptions accordingly. AIFASE also displays the current usage status of all ergometers and the users’ physiological conditions. The algorithms were developed to generate warning alerts when the training workload was too large by personal physiological detection. AIFASE automatically customized the exercise prescription according to the user’s exercise performance. Results After a 12-week AIFASE intervention, the intervention group exhibited significant improvements in the strength of the hip flexor, Semi-Tandem Stand, and Tandem Stand. Conclusion In this study, we developed an AIoT ergometer that delivered customized physical training prescriptions to improve the physical performance of long-term care facility residents. We believe that the application of AIFASE will help improve the quality of institutional care.

In physiological concentrations, heme is nontoxic to the cell and is essential for cell survival and proliferation. Increasing intracellular heme concentrations beyond normal levels, however, will lead to carcinogenesis and facilitate the survival of tumor cells. Simultaneously, heme in an abnormally high quantity is also a potent inducer of tumor cell death, contributing to its ability to generate oxidative stress on the cells by boosting oxidative phosphorylation and suppressing tumors through ferroptosis. During tumorigenesis and progression, therefore, heme works as a double-edged sword. Heme oxygenase 1 (HO-1) is the rate-limiting enzyme in heme catabolism, which converts heme into physiologically active catabolites of carbon monoxide (CO), biliverdin, and ferrous iron (FeHerein, the authors review recent studies on heme, HO-1, and Bach1 in cancer. Specifically, they cover the following areas: (1) the carcinogenic and anticarcinogenic aspects of heme; (2) the carcinogenic and anticarcinogenic aspects of HO-1; (3) the carcinogenic and anticarcinogenic aspects of Bach1; (4) the interactions of the heme/HO-1/Bach1 axis involved in tumor progression.This review summarized the literature about the dual role of the heme/HO-1/Bach1 axis and their mutual dependence in the carcinogenesis and anti-carcinogenesis intersection.

To describe medication adherence, to analyze the relationships among medication adherence, illness perception, and beliefs about medicines, and to determine the mediating effects of beliefs about medicines on the relationship in breast cancer patients with adjuvant endocrine therapy (AET) in China.A cross-sectional study was conducted on 202 breast cancer patients with AET from September 2017 to February 2019 in China. The Medication Adherence Report Scale (MARS-5), the Chinese version of the revised illness perception questionnaire for Breast Cancer (CIPQ-R-BC) and the Beliefs about Medicines Questionnaire (BMQ) were used.The mean MARS-5 score of our participants was 23.72 (SD = 1.62), and 175 (86.6%) patients were adherent to medications. Moreover, medication adherence was negatively correlated with identity, environmental or immune factors, emotional representations, BMQ-specific concerns, BMQ-general overuse, and BMQ-general harm, as well as being positively correlated with coherence and the total BMQ scores. Furthermore, beliefs in the overuse about medicines functioned as mediators for the influencing effects of coherence and emotional representations on medication adherence.Illness perception not only directly affected medication adherence, but also indirectly affected medication adherence through the beliefs about medicines. Necessary interventions that target beliefs in the overuse about medicines in breast cancer patients with AET with low levels of coherence or high levels of emotional representations could be provided to improve the level of their medication adherence.

A decreased estimated glomerular filtration rate (eGFR) is associated with frailty, but the association between kidney function and frailty using multidimensional assessments has not been entirely examined. We aimed to investigate whether albuminuria and the eGFR using different biomarkers were associated with frailty.A total of 1830 older adults were included. Kidney function was assessed by the eGFR (based on combined creatinine-cystatin C [eGFRAt baseline, kidney function was associated with prevalent frailty. During the 2-year follow-up, a decreased eGFR (per 10 units) was associated with an increased risk of incident frailty using the FP (eGFRKidney function decline and CKD were associated with an increased risk of prevalent and incident frailty in older adults. Physicians should pay more attention to monitoring frailty status in older adults with CKD, even in those with kidney function decline.

Understanding how groundwater is formed and evolves is critical for water resource exploitation and utilization. In this study, hydrochemistry and stable isotope tracing techniques were adopted to determine the key factors influencing groundwater chemical evolution in Feng County. A total of fourteen wells and five surface water samples were investigated in November 2021. The δD and δ

Glioblastoma (GBM) is the most malignant and challenging type of astrocytoma and also notoriously acknowledged as the most common primary brain tumor globally. Currently, chemotherapy is the most master therapy for tumor and is essential in clinical treatment for GBM. Nevertheless, the characterization of chemotherapy resistance seriously hinders clinical chemotherapy treatment. Accordingly, there are imperious demands for the exploitation of novel chemosensitizer to promote the efficacy of chemotherapy. Our current study was conducted to probe into the potential impacts of microRNA (miR)-640 on the chemosensitivity in GBM and the associated underlying mechanism. Initially, TargetScan software was utilized to predict the targeted genes of miR-640, and the target relationship between miR-640 and Bcl-2-modifying factor (BMF) was validated by double luciferase report assay. Additionally, to explore the role of miR-640/BMF in U251 cells, miR-640 inhibitor/BMF-siRNA was used. U251 cells were processed with 100 μM temozolomide (TMZ) and detected with CCK-8 kit. Eventually, RT-qPCR and Western blotting were used for evaluating Bcl-2, Bax mRNA, and protein expression level. Flow cytometry analysis was performed to measure cellular apoptosis. Initially, the results indicated that BMF was the target gene of miR-640. MiR-640 negatively regulated BMF expression in GBM cells. Besides, the findings revealed that miR-640 inhibition significantly inhibited U251 cell proliferation, promoted cell apoptosis, and increased the sensitivity of GBM cells to TMZ by targeting BMF. Moreover, BMF overexpression significantly suppressed U251 cell proliferation, induced cell apoptosis, and increased the sensitivity of GBM cells to TMZ. Inhibition of miR-640 expression enhances chemosensitivity of human GBM cells to TMZ by targeting BMF.

Sub-anesthetic ketamine has rapid-onset effects for the treatment of major depressive disorder (MDD). However, the mechanism underlying ketamine's antidepressant properties remains unclear. Recent studies have reported an interrelationship between autophagy and the inflammasome, both of which are involved in the pathophysiology of MDD. In this study, we assess whether ketamine exerts its antidepressant effects via an association with the autophagy-NLRP3 inflammasome pathway.We established a depressive-like rat model by treating Wistar Kyoto rats with chronic restraint stress (CRS) for 28 days. Microglial cells from newborn Sprague-Dawley rats were used for in vitro experiments.We found sub-anesthetic ketamine treatment reversed depressive-like behavior in CRS rats. Ketamine triggered autophagy in the microglia of prefrontal cortex (PFC) and (hippocampus) HPC, with increased levels of LC3B, decreased levels of p62 protein, and elevated autophagosomes both in vivo and in vitro. Moreover, NLRP3 inflammasome activation was also inhibited by ketamine, with reduced expression of NLRP3-ASC-CASP1 assembly and decreased IL-1β levels in cerebrospinal fluid (CSF) as well as in the serum. Increased BDNF levels and synaptophysin levels were detected in the ketamine-treated group. The rapid anti-depressive effects, elevation of autophagy, reduction in NLRP3, and neuroplasticity-related factors induced by ketamine could be significantly blocked by the autophagy inhibitor Baf A1 (0.1 mg/kg).Our findings demonstrate that sub-anesthetic doses of ketamine exert their antidepressant-like effects by inhibiting inflammation and initiating neuroprotection via autophagy activation. These data might help expand future investigations on the antidepressant properties of ketamine.

Cleft lip and palate is one of the most frequent congenital developmental defects. Autophagy is a highly conserved process of cell self-degradation in eukaryotes, involving multiple biological processes in which chloroquine (CQ) is the most common inhibitor. However, whether CQ affects and how it affects palate development is unknown. Mouse embryonic palatal cells (MEPCs) were treated with CQ to observe cell viability, apoptosis, migration, osteogenic differentiation by cell proliferation assay, flow cytometric analysis, scratch assay, and alizarin red staining. PI staining was used to measure cell cycle distribution. Immunofluorescence (IF) assay and transmission electron microscopy were used to detect autophagosomes. The autophagy-related factors (LC3 and P62), apoptosis-related markers (P53, caspase-3 cleaved caspase-3, BAX, and BCL-2), and cell cycle-related proteins (P21, CDK2, CDK4, cyclin D1, and cyclin E) were all measured by western blot. CQ inhibited the proliferation of MEPCs by arresting the G0/G1 phase of the cell cycle in a concentration- and time-dependent manner with cell cycle-related proteins P21 upregulated and CDK2, CDK4, cyclin D1, and cyclin E downregulated. Then we detected CQ also induced cell apoptosis in a dose-dependent manner by decreasing the BCL-2/BAX ratio and increasing cleaved caspase-3. Next, it was investigated that migration and osteogenesis of MEPCs decreased with CQ treatment in a dose-dependent manner. Meanwhile, CQ blocked the autophagy pathway by upregulating LC3II and P62 expressions which activated the P53 pathway. CQ activates P53 which affects MEPC biological characteristics by changing the proliferation and apoptosis of MEPCs through inhibiting autophagy.

Abstract Ischemia-reperfusion injury (IRI) is the main cause of acute kidney injury (AKI), and there is no effective therapy. Microenvironmental acidification is generally observed in ischemic tissues. Acid-sensing ion channel 1a (ASIC1a) can be activated by a decrease in extracellular pH which mediates neuronal IRI. Our previous study demonstrated that, ASIC1a inhibition alleviates renal IRI. However, the underlying mechanisms have not been fully elucidated. In this study, we determined that renal tubule-specific deletion of ASIC1a in mice (ASIC1afl/fl/CDH16cre) attenuated renal IRI, and reduced the expression of NLRP3, ASC, cleaved-caspase-1, GSDMD-N, and IL-1β. Consistent with these in vivo results, inhibition of ASIC1a by the specific inhibitor PcTx-1 protected HK-2 cells from hypoxia/reoxygenation (H/R) injury, and suppressed H/R-induced NLRP3 inflammasome activation. Mechanistically, the activation of ASIC1a by either IRI or H/R induced the phosphorylation of NF-κB p65, which translocates to the nucleus and promotes the transcription of NLRP3 and pro-IL-1β. Blocking NF-κB by treatment with BAY 11-7082 validated the roles of H/R and acidosis in NLRP3 inflammasome activation. This further confirmed that ASIC1a promotes NLRP3 inflammasome activation, which requires the NF-κB pathway. In conclusion, our study suggests that ASIC1a contributes to renal IRI by affecting the NF-κB/NLRP3 inflammasome pathway. Therefore, ASIC1a may be a potential therapeutic target for AKI. Key messages Knockout of ASIC1a attenuated renal ischemia-reperfusion injury. ASIC1a promoted the NF-κB pathway and NLRP3 inflammasome activation. Inhibition of the NF-κB mitigated the NLRP3 inflammasome activation induced by ASIC1a.

Algorithms and frameworks for distributed machine learning have been widely used in numerous artificial intelligence engineering applications. A cloud platform provides a large number of resources at a lower cost and is a more convenient method for such applications. With the rapid development of containerization, native cloud combinations based on Docker and Kubernetes have provided effective resource support for distributed machine learning. However, native Kubernetes does not provide efficient priority or fair resource scheduling strategies for distributed machine learning in computationally intensive and time-consuming jobs, which easily leads to resource deadlock, resource waste, and low job execution efficiency. Therefore, to utilize the execution order between multiple jobs in distributed machine learning as well as the dependencies between multiple tasks for the same job, considering intra- and inter-group scheduling priorities, a combined priority scheduling method is proposed for distributed machine learning based on Kubernetes and Volcano. Considering the user priority, task priority, longest wait time, task parallelism, and affinity and non-affinity between the parameter server and worker nodes, a combined priority scheduling model of inter- and intra-job priority is proposed, which is mapped into a scheduling strategy of inter- and intra-group priorities of pods, enabling the efficient scheduling and training of distributed machine learning. The experiment results show that the proposed method achieves preferential resource allocation for urgent, high parallelism, and high-priority jobs with high-priority users and improves the job execution efficiency. The affinity and anti-affinity settings among pods reduce the time of information interaction between the parameter server and worker nodes to a certain extent, thereby improving the job completion efficiency. This group scheduling strategy alleviates the problems of resource deadlock and waste caused by insufficient resources in cloud computing.

This research aims to fabricate plane-type substrates for evaluating the axon behaviors of neuronal cells in vitro toward the development of brain-on-chip models by applying the functions of diamond-like carbon (DLC) thin film deposition, which helped to eliminate the costly and time-consuming lithography process by utilizing a shadow mask. The DLC thin films were partially deposited on stretched polydimethylsiloxane (PDMS) substrates covered with a metal mask by the plasma chemical vaper deposition method, and using the substrates culture teats with human neuroblastoma cells (SH-SY5Y) were performed. Three patterns of interconnection structures of axons were created on the substrates with disordered and regular linear wrinkle structures with several μm size formed by the depositions. The patterns were characterized by the structure that the aggregations of axons formed on the linear DLC thin film deposited areas were separately placed in regular intervals and connected each other by plenty of axons, which were individually taut in a straight line at about 100 to over 200 μm in length. The substrates expected of uses for evaluation of axon behaviors are available without fabricating guiding grooves by conventional soft lithographic methods requiring multiple stages and their treating times.