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AIM: The objective of this study was to investigate the effects of a combination of emulsified isoflurane, lidocaine, and fentanyl (EI-L-F) compared with the efficacy of emulsified isoflurane alone (EI), a combination of emulsified isoflurane and lidocaine (EI-L) or emulsified isoflurane and fentanyl (EI-F) for anaesthetising dogs. METHODS: Eight mongrel dogs were anesthetised with EI (8 mL/kg/hour), EI-L (3 mg/kg/hour lidocaine and 6 mL/kg/hour of emulsified isoflurane), EI-F (1.5 μg/kg/hour fentanyl and 6 mL/kg/hour of emulsified isoflurane), and EI-L-F (5 mL/kg/hour of emulsified isoflurane, 1 μg/kg/hour of fentanyl and 2.4 mg/kg/hour of lidocaine). Each dog received all four treatments and there was a 15-day washout period between the treatments. The dogs' anaesthesia and analgesia scores and physiological parameters were determined before and 5, 10, 20, 30, 40, 50, 60, 70 and 80 minutes after the administration of anaesthetic agents. RESULTS: The dogs in each of the four groups became laterally recumbent within 1 minute. Respiration rate and heart rate increased (p<0.05) during the first 5 minutes of anaesthesia in all groups. Respiration rate in the EI-F-L group was higher (p=0.037) than other groups from 30 to 50 minutes. Heart rate was higher in the EI than EI-F-L group (p=0.018) from 10 to 20 minutes, then returned to near baseline. Arterial oxygen saturation decreased during the period of anaesthesia but was higher (p=0.032) from 10 to 50 minutes in EI-F-L group than in other groups. The total anaesthesia scores in the EI-L-F group were higher than the EI and EI-L groups (p<0.05). The mean time to body movement was 5 (SD 2), 5 (SD 2), 7 (SD 2) and 8 (SD 2) minutes for the EI, EI-L, EI-F and EI-F-L groups, respectively. The mean time to standing was 8 (SD 2), 9 (SD 2), 10 (SD 2) and 13 (SD 3) minutes for the EI, EI-L, EI-F and EI-F-L groups, respectively. No excitement was observed during recovery after anaesthesia. CONCLUSIONS: The EI-F-L combination that was used in this study provided an adequate anaesthesia effect in dogs, which was characterised by adequate analgesia and muscle relaxation without any complications. [ABSTRACT FROM PUBLISHER]

The single-component Mollerup model, with over 40 direct applications and 442 citations, is the most widely used activity model for chromatographic mechanistic modeling. Many researchers have extended this formula to multi-component systems by directly adding subscripts, a modification deemed thermodynamically inconsistent (referred to as the reference model). In this work, we rederived the asymmetric activity model for multi-component systems, using the van der Waals equation of state, and termed it the multi-component Mollerup model. In contrast to the reference model, our proposed model accounts for the contributions of all components to the activity. Three numerical experiments were performed to investigate the impact of the three different activity models on the chromatographic modeling. The results indicate that our proposed model represents a thermodynamically consistent generalization of the single-component Mollerup model to multi-component systems. This communication advocates adopting of the multi-component Mollerup model for activity modeling in multi-component chromatographic separation to enhance thermodynamic consistency., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Mechanistic models are powerful tools for chromatographic process development and optimization. However, hydrophobic interaction chromatography (HIC) mechanistic models lack an effective and logical parameter estimation method, especially for multi-component system. In this study, a parameter-by-parameter method for multi-component system (called as mPbP-HIC) was derived based on the retention mechanism to estimate the six parameters of the Mollerup isotherm for HIC. The linear parameters (k s,i and k eq,i ) and nonlinear parameters (n i and q max,i ) of the isotherm can be estimated by the linear regression (LR) and the linear approximation (LA) steps, respectively. The remaining two parameters (k p,i and k kin,i ) are obtained by the inverse method (IM). The proposed method was verified with a two-component model system. The results showed that the model could accurately predict the protein elution at a loading of 10 g/L. However, the elution curve fitting was unsatisfactory for high loadings (12 g/L and 14 g/L), which is mainly attributed to the demanding experimental conditions of the LA step and the potential large estimation error of the parameter q max . Therefore, the inverse method was introduced to further calibrate the parameter q max , thereby reducing the estimation error and improving the curve fitting. Moreover, the simplified linear approximation (SLA) was proposed by reasonable assumption, which provides the initial guess of q max without solving any complex matrix and avoids the problem of matrix unsolvable. In the improved mPbP-HIC method, q max would be initialized by the SLA and finally determined by the inverse method, and this strategy was named as SLA+IM. The experimental validation showed that the improved mPbP-HIC method has a better curve fitting, and the use of SLA+IM reduces the error accumulation effect. In process optimization, the parameters estimated by the improved mPbP-HIC method provided the model with excellent predictive ability and reasonable extrapolation. In conclusion, the SLA+IM strategy makes the improved mPbP-HIC method more rational and can be easily applied to the practical separation of protein mixture, which would accelerate the process development for HIC in downstream of biopharmaceuticals., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

In the present study, a new degraded konjac glucomannan (DKGM) was prepared using a crude enzyme from abalone ( Haliotis discus hannai ) viscera, and its physicochemical properties were investigated. After enzymatic hydrolysis, the viscosity of KGM obviously decreased from 15,500 mPa·s to 398 mPa·s. The rheological properties analysis of KGM and DKGMs revealed that they were pseudoplastic fluids, and pseudoplasticity, viscoelasticity, melting temperature, and gelling temperature significantly decreased after enzymatic hydrolysis, especially for KGM-180 and KGM-240. In addition, the molecular weight of KGM decreased from 1.80 × 10 6 Da, to 0.45 × 10 6 Da and the polydispersity index increased from 1.17 to 1.83 after 240 min of degradation time. Compared with natural KGM, the smaller particle size distribution of DKGM further suggests enzyme hydrolysis reduces the aggregation of molecular chains with low molecular weight. FT-IR and FESEM analyses showed that the fragmented KMG chain did not affect the structural characteristics of molecular monomers; however, the dense three-dimensional network microstructure formed by intermolecular interaction changed to fragment microstructure after enzyme hydrolysis. These results revealed that the viscosity and rheological properties of KGM could be controlled and effectively changed using crude enzymes from abalone viscera. This work provides theoretical guidance for the promising application of DKGM in the food industry.

Regulatory authorities recommend using residence time distribution (RTD) to address material traceability in continuous manufacturing. Continuous virus filtration is an essential but poorly understood step in biologics manufacturing in respect to fluid dynamics and scale-up. Here we describe a model that considers nonideal mixing and film resistance for RTD prediction in continuous virus filtration, and its experimental validation using the inert tracer NaNO 3 . The model was successfully calibrated through pulse injection experiments, yielding good agreement between model prediction and experiment ( R 2 > ${R}^{2}\gt $  0.90). The model enabled the prediction of RTD with variations-for example, in injection volumes, flow rates, tracer concentrations, and filter surface areas-and was validated using stepwise experiments and combined stepwise and pulse injection experiments. All validation experiments achieved R 2 > ${R}^{2}\gt $  0.97. Notably, if the process includes a porous material-such as a porous chromatography material, ultrafilter, or virus filter-it must be considered whether the molecule size affects the RTD, as tracers with different sizes may penetrate the pore space differently. Calibration of the model with NaNO 3 enabled extrapolation to RTD of recombinant antibodies, which will promote significant savings in antibody consumption. This RTD model is ready for further application in end-to-end integrated continuous downstream processes, such as addressing material traceability during continuous virus filtration processes., (© 2024 The Authors. Biotechnology and Bioengineering published by Wiley Periodicals LLC.)

Digital twin (DT) is a virtual and digital representation of physical objects or processes. In this paper, this concept is applied to dynamic control of the collection window in the ion exchange chromatography (IEC) toward sample variations. A possible structure of a feedforward model-based control DT system was proposed. Initially, a precise IEC mechanistic model was established through experiments, model fitting, and validation. The average root mean square error (RMSE) of fitting and validation was 8.1% and 7.4%, respectively. Then a model-based gradient optimization was performed, resulting in a 70.0% yield with a remarkable 11.2% increase. Subsequently, the DT was established by systematically integrating the model, chromatography system, online high-performance liquid chromatography, and a server computer. The DT was validated under varying load conditions. The results demonstrated that the DT could offer an accurate control with acidic variants proportion and yield difference of less than 2% compared to the offline analysis. The embedding mechanistic model also showed a positive predictive performance with an average RMSE of 11.7% during the DT test under >10% sample variation. Practical scenario tests indicated that tightening the control target could further enhance the DT robustness, achieving over 98% success rate with an average yield of 72.7%. The results demonstrated that the constructed DT could accurately mimic real-world situations and perform an automated and flexible pooling in IEC. Additionally, a detailed methodology for applying DT was summarized., (© 2024 Wiley Periodicals LLC.)

Developing an accurate and reliable model for chromatographic separation that meets regulatory requirements and ensures consistency in model development remains challenging. In order to address this challenge, a standardized approach was proposed in this study with ion-exchange chromatography (IEC). The approach includes the following steps: liquid flow identification, system and column-specific parameters determination and validation, multi-component system identification, protein amount validation, steric mass action parameters determination and evaluation, and validation of the calibrated model's generalization ability. The parameter-by-parameter (PbP) calibration method and the consideration of extra-column effects were integrated to enhance the accuracy of the developed models. The experiments designed for implementing the PbP method (five gradient experiments for model calibration and one stepwise experiment for model validation) not only streamline the experimental workload but also ensure the extrapolation abilities of the model. The effectiveness of the standardized approach is successfully validated through an application about the IEC separation of industrial antibody variants, and satisfactory results were observed with R 2 ≈ 0.9 for the majority of calibration and validation experiments. The standardized approach proposed in this work contributes significantly to improve the accuracy and reliability of the developed IEC models. Models developed using this standardized approach are ready to be applied to a broader range of industrial separation systems, and are likely find further applications in model-assisted decision-making of process development., (© 2024 Wiley-VCH GmbH.)

Hydrophobic interaction chromatography (HIC) is used as a critical polishing step in the downstream processing of biopharmaceuticals. Normally the process development of HIC is a cumbersome and time-consuming task, and the mechanical models can provide a powerful tool to characterize the process, assist process design and accelerate process development. However, the current estimation of model parameters relies on the inverse method, which lacks an efficient and logical parameter estimation strategy. In this study, a parameter-by-parameter (PbP) method based on the theoretical derivation and simplifying assumptions was proposed to estimate the Mollerup isotherm parameters for HIC. The method involves three key steps: (1) linear regression (LR) to estimate the salt-protein interaction parameter and the equilibrium constant; (2) linear approximation (LA) to estimate the stoichiometric parameter and the maximum binding capacity; and (3) inverse method to estimate the protein-protein interaction parameter and the kinetic coefficient. The results indicated that the LR step should be used for dilution condition (loading factor below 5%), while the LA step should be conducted when the isotherm is in the transition or nonlinear regions. Six numerical experiments were conducted to implement the PbP method. The results demonstrated that the PbP method developed allows for the systematic estimation of HIC parameters one-by-one, effectively reducing the number of parameters required for inverse method estimation from six to two. This helps prevent non-identifiability of structural parameters. The feasibility of the PbP-HIC method was further validated by real-world experiments. Moreover, the PbP method enhances the mechanistic understanding of adsorption behavior of HIC and shows a promising application to other stoichiometric displacement model-derived isotherms., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objective: The clinical impact of baseline mitral regurgitation (MR) on the outcomes after transcatheter aortic valve replacement (TAVR) is not clear. This study sought to assess the clinical impact of baseline MR on outcomes after TAVR., Methods: The study was a retrospective analysis. Data was from 120 consecutive patients with severe aortic stenosis (AS) undergoing TAVR at our center from June 2018 and July 2020. Clinical outcomes were assessed at 30-day, 1- and 2-year follow-up., Results: The median follow-up was 736.0 (interquartile range, 666.0-965.0) days. Overall survival in patients with nonsignificant and significant baseline MR was not significantly different, while patients from the improved MR group after TAVR demonstrated a significantly higher survival than unchanged or worsened MR group during 2-year follow-up. NYHA functional class had generally improved at 1 year, with only 8.3 % of patients with nonsignificant MR and 17.5 % of patients with significant MR in class III or IV. Patients with improved MR at 1 year after TAVR had a significantly higher LVEF, smaller LVEDD and LVESD than those with unchanged or worsened MR. Among the significant baseline MR group, 70.4 % and 80.0 % of patients had improved to nonsignificant MR at 30-day and 1-year follow-up after TAVR, respectively., Conclusions: Significant baseline MR was not associated with the increased risk of all-cause mortality 2 years after TAVR. Significant baseline MR was improved in most patients at 1 year after TAVR. Patients with unchanged or worsened MR had an increased all-cause mortality., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Author(s).)

Multi-column periodic counter-current chromatography is a promising technology for continuous antibody capture. However, dynamic changes due to disturbances and drifts pose some potential risks for continuous processes during long-term operation. In this study, a model-based approach was used to describe the changes in breakthrough curves with feedstock variations in target proteins and impurities. The performances of continuous capture of three-column periodic counter-current chromatography under ΔUV dynamic control were systematically evaluated with modeling to assess the risks under different feedstock variations. As the concentration of target protein decreased rapidly, the protein might not breakthrough from the first column, resulting in the failure of ΔUV control. Small reductions in the concentrations of target proteins or impurities would cause protein losses, which could be predicted by the modeling. The combination of target protein and impurity variations showed complicated effects on the process performance of continuous capture. A contour map was proposed to describe the comprehensive impacts under different situations, and nonoperation areas could be identified due to control failure or protein loss. With the model-based approach, after the model parameters are estimated from the breakthrough curves, it can rapidly predict the process stability under dynamic control and assess the risks under feedstock variations or UV signal drifts. In conclusion, the model-based approach is a powerful tool for continuous process evaluation under dynamic changes and would be useful for establishing a new real-time dynamic control strategy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Objectives: The aim of the present study was to assess the differences between BAV and TAV patients with chronic moderate to severe or severe AS regarding presentation, incidence of TAVR, survival, ascending aorta diameter and dilatation rate before and after TAVR., Methods: The study included 667 consecutive patients with chronic moderate to severe or severe AS from January 2012 and December 2022. Outcomes included all-cause mortality, incidence of TAVR, and ascending aorta diameter and dilatation rate., Results: There were 185 BAV-AS and 482 TAV-AS patients, and BAV-AS patients were younger (67 vs 78 years, P = 0.027). Total follow-up was 4.5 years (IQR: 2.7-8.9 years), 290 patients underwent TAVR, and 165 patients died. The 8-year TAVR incidence was higher in BAV-AS (55% ± 4%) vs TAV-AS (41% ± 5%; P = 0.02). The 8-year survival was higher in BAV-AS (85% ± 6%) vs TAV-AS (71% ± 6%; P < 0.0001) and became insignificant after age adjustment ( P = 0.33). The dilatation rate of ascending aorta was significantly faster in BAV-AS patients compared with TAV-AS patients before TAVR. However, the ascending aorta dilatation rate for BAV-AS and TAV-AS patients was not significantly different after TAVR., Conclusions: Compared with TAV-AS, BAV-AS patients were younger and underwent TAVR more frequently, resulting in a considerable survival advantage. After TAVR, ascending aorta dilatation rates were similar in BAV-AS and TAV-AS patients, suggesting an important role of hemodynamics on ascending aorta dilatation in BAV-AS., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Author(s).)

Objective: We designed a simplified total arch reconstruction (s-TAR) technique which could be performed under mild hypothermia (30-32 °C) with distal aortic perfusion. This study aimed to compare its efficacy of organ protection with the conventional total arch reconstruction (c-TAR)., Methods: We reviewed the clinical data of 195 patients who had ascending aortic aneurysm with extended aortic arch dilation and underwent simultaneous ascending aorta replacement and TAR procedure between January 2018 and December 2022 in our center. 105 received c-TAR under moderate hypothermia (25-28 °C) with circulatory arrest (c-TAR group); rest 90 received s-TAR under mild hypothermia (30-32 °C) with distal aortic perfusion (s-TAR group)., Results: The s-TAR group demonstrated shorter CPB time, cross-clamp time and lower body circulatory arrest time compared with the c-TAR group. The 30-day mortality was 2.9% for the c-TAR group and 1.1% for the s-TAR group (P = 0.043). The mean duration of mechanical ventilation was shorter in the s-TAR group. Paraplegia was observed in 4 of 105 patients (3.8%) in the c-TAR group, while no such events were observed in the s-TAR group. The incidence of temporary neurologic dysfunction was significantly higher in the c-TAR group. The incidence of permanent neurologic dysfunction also showed a tendency to be higher in the c-TAR group, without statistical significance. Furthermore, the incidence of reoperation for bleeding were significantly lower in the s-TAR group. The rate of postoperative hepatic dysfunction and all grades of AKI was remarkably lower in the s-TAR group. The 3-year survival rate was 95.6% in the s-TAR group and 91.4% in the c-TAR group., Conclusions: s-TAR under mild hypothermia (30-32℃) with distal aortic perfusion is associated with lower mortality and morbidity, offering better neurological and visceral organ protection compared with c-TAR., (© 2023. The Author(s).)

Background and Aims: Understanding adaptive genetic variation and whether it can keep pace with predicted future climate change is critical in assessing the genetic vulnerability of species and developing conservation management strategies. The lack of information on adaptive genetic variation in relict species carrying abundant genetic resources hinders the assessment of genetic vulnerability. Using a landscape genomics approach, this study aimed to determine how adaptive genetic variation shapes population divergence and to predict the adaptive potential of Pterocarya macroptera (a vulnerable relict species in China) under future climate scenarios., Methods: We applied restriction site-associated DNA sequencing (RAD-seq) to obtain 8244 single-nucleotide polymorphisms (SNPs) from 160 individuals across 28 populations. We examined the pattern of genetic diversity and divergence, and then identified outliers by genetic differentiation (FST) and genotype-environment association (GEA) methods. We further dissected the effect of geographical/environmental gradients on genetic variation. Finally, we predicted genetic vulnerability and adaptive risk under future climate scenarios., Key Results: We identified three genetic lineages within P. macroptera: the Qinling-Daba-Tianmu Mountains (QDT), Western Sichuan (WS) and Northwest Yunnan (NWY) lineages, which showed significant signals of isolation by distance (IBD) and isolation by environment (IBE). IBD and IBE explained 3.7-5.7 and 8.6-12.8 % of the genetic structure, respectively. The identified GEA SNP-related genes were involved in chemical defence and gene regulation and may exhibit higher genetic variation to adapt to the environment. Gradient forest analysis revealed that the genetic variation was mainly shaped by temperature-related variables, indicating its adaptation to local thermal environments. A limited adaptive potential was suggested by the high levels of genetic vulnerability in marginal populations., Conclusions: Environmental gradient mainly shaped the population differentiation of P. macroptera. Marginal populations may be at high risk of extinction, and thus proactive management measures, such as assisted gene flow, are required to ensure the survival of these populations., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Annals of Botany Company.)

Numerical method is widely used for solving the mechanistic models of chromatography process, but it is time-consuming and hard to response in real-time. Physics-informed neural network (PINN) as an emerging technology combines the structure of neural network with physics laws, and is getting noticed for solving physics problems with a balanced accuracy and calculation speed. In this research, a proof-of-concept study was carried out to apply PINN to chromatography process simulation. The PINN model structure was designed for the lumped kinetic model (LKM) with all LKM parameters. The PINN structure, training data and model complexity were optimized, and an optimal mode was obtained by adopting an in-series structure with a nonuniform training data set focusing on the breakthrough transition region. A PINN for LKM (LKM-PINN) consisting of four neural networks, 12 layers and 606 neurons was then used for the simulation of breakthrough curves of chromatography processes. The LKM parameters were estimated with two breakthrough curves and used to infer the breakthrough curves at different residence times, loading concentrations and column sizes. The results were comparable to that obtained with numerical methods. With the same raw data and constraints, the average fitting error for LKM-PINN model was 0.075, which was 0.081 for numerical method. With the same initial guess, the LKM-PINN model took 160 s to complete the fitting, while the numerical method took 7 to 72 min, depending on the fitting settings. The fitting speed of LKM-PINN model was further improved to 30 s with random initial guess. Thus, the LKM-PINN model developed in this study is capable to be applied to real-time simulation for digital twin., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Conduction disturbances requiring permanent pacemaker implantation (PPI) are common following transcatheter aortic valve replacement (TAVR). There were conflicting data regarding the impact of new PPI on clinical outcomes after TAVR., Objectives: The study sought to evaluate the impact of new PPI on clinical outcomes in patients undergoing TAVR., Methods: This study was a retrospective analysis of prospectively collected data. Data were from 210 consecutive patients without prior PPI who underwent TAVR due to severe symptomatic aortic stenosis at our center between June 2018 and July 2020. Clinical, echocardiographic, and pacing data were assessed at 30-day, 1- and 2-year follow-up., Results: New PPI was required in 35 (16.7%) patients within 30 days after TAVR. The median time from TAVR to PPI was 3 days. The most common indication for PPI was high-degree or complete atrioventricular block. The median follow-up was 798.0 (interquartile range, 669.0-1115.0) days. There were no differences in all-cause mortality (adjusted hazard ratio [HR]: 1.18; 95% confidence interval [CI]: 0.85-2.36; p = 0.415) and cardiovascular mortality (adjusted HR: 0.92; 95% CI: 0.57-1.89; p = 0.609) between groups. However, PPI group had a higher risk of heart failure (HF) rehospitalization (adjusted HR: 1.53; 95% CI: 1.26-2.28; p = 0.027). Echocardiography showed no significant improvement of LVEF over time in patients with PPI. At the latest follow-up, 31.3% of patients exhibited low (≤10%) pacing burdens, whereas 28.1% of patients had near constant (>90%) right ventricular pacing., Conclusions: New PPI within 30 days following TAVR was not associated with an increased risk of all-cause or cardiovascular mortality. However, patients with PPI had a higher risk of HF rehospitalization and lack of LVEF improvement., (© 2023 Wiley Periodicals LLC.)

Background: One of the crucial aspects of ascending aorta replacement is to achieve hemostasis of the proximal anastomosis. This study aimed to describe a modified prosthesis eversion technique for proximal anastomosis in ascending aorta replacement and compare its operative outcomes with the conventional prosthesis eversion technique., Methods: We conducted a retrospective analysis of all consecutive patients who had ascending aortic aneurysm and underwent ascending aorta replacement with the modified or conventional prosthesis eversion technique between January 2019 and December 2022 in our center., Results: A total of 108 patients were included: 55 in the modified group and 53 in the conventional group. The durations of cardiopulmonary bypass, aortic cross-clamping and total operation in the conventional group were longer than those in the modified group. Furthermore, perioperative blood loss and the incidence of re-exploration for bleeding were significantly lower in the modified group. Accordingly, patients in the conventional group accepted more blood transfusion. The modified group had a shorter duration in intensive care unit (ICU) and hospital, and lower total hospitalization costs than those in the conventional group., Conclusions: The modified prosthesis eversion technique is an effective alternative for proximal anastomosis in ascending aorta replacement, with less blood loss, shorter operation time, and lower rate of postoperative complications compared with the conventional technique., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://jtd.amegroups.com/article/view/10.21037/jtd-23-550/coif). The authors have no conflicts of interest to declare., (2023 Journal of Thoracic Disease. All rights reserved.)

Continuous manufacturing in monoclonal antibody production has generated increased interest due to its consistent quality, high productivity, high equipment utilization, and low cost. One of the major challenges in realizing continuous biological manufacturing lies in implementing continuous chromatography. Given the complex operation mode and various operation parameters, it is challenging to develop a continuous process. Due to the process parameters being mainly determined by the breakthrough curves and elution behaviors, chromatographic modeling has gradually been used to assist in process development and characterization. Model-assisted approaches could realize multi-parameter interaction investigation and multi-objective optimization by integrating continuous process models. These approaches could reduce time and resource consumption while achieving a comprehensive and systematic understanding of the process. This paper reviews the application of modeling tools in continuous chromatography process development, characterization and design. Model-assisted process development approaches for continuous capture and polishing steps are introduced and summarized. The challenges and potential of model-assisted process characterization are discussed, emphasizing the need for further research on the design space determination strategy and parameter robustness analysis method. Additionally, some model applications for process design were highlighted to promote the establishment of the process optimization and process simulation platform., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The multicolumn counter-current solvent gradient purification (MCSGP) method has proven effective in addressing the issue of elution profile overlap for difficult-to-separate proteins, leading to improved purity and recovery. However, during the MCSGP process, the flow rate and proportion of loaded proteins undergo changes, causing a significant discrepancy between the elution profiles of batch process design and the actual MCSGP process. This mismatch negatively impacts the purity and recovery of the target protein. To address this challenge, an improved process design (reDesign) was proposed with the first-run MCSGP to mimic the actual continuous process and obtain elution profiles that closely resemble the real ones. The reDesign was demonstrated with both a model protein mixture and a sample of monoclonal antibody (mAb) with charge variants. For model protein mixture, the reDesign-based MCSGP process (reMCSGP) showed a remarkable improvement in recovery, increasing from 83.6% to 97.8% while maintaining a purity of more than 95%. For mAb sample, the recovery of reMCSGP improved significantly to 93.9%, surpassing the performance of normal MCSGP processes at a given purity level of more than 84%. In general, the new process design strategy developed in this work could generate a more representative elution profile that closely mirrors actual conditions in continuous processes, which enhances the separation performance of MCSGP., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Crystallization of organic steric molecules often leads to multiple polyhedral crystal morphologies. However, the relationships among the molecular structure, supramolecular interaction, aggregation mode and crystal morphology are still unclear. In this work, we elaborate two model crystals formed by spiro[fluorene-9,9'-xanthene] (SFX) and spiro[cyclopenta[1,2-b : 5,4-b']dipyridine-5,9'-xanthene] (SDAFX) to demonstrate the feasibility of morphology prediction by periodic bond chain (PBC) theory based on interaction energy (IE) values in terms of single point energy. With non-directional van der Waals forces, only one PBC direction is found in SFX crystal, leading to the irregular 1D rod-like structure. Compared with SFX, the extra N heteroatoms in SDAFX can bring additional hydrogen bonds and some other interactions into the bulky molecular skeletons, inducing 3-dimensionally oriented PBCs to form the explicit F-face network in SDAFX which leads to the final octahedral structure. A simple and accurate method has been provided to quantify PBC vector on the supramolecular level in the organic molecular system, and the PBC theory has also been further demonstrated and developed in the morphology prediction of organic spiro-molecules., (© 2023 Wiley-VCH GmbH.)

Ultrathin 2D organic nanosheets (2DONs) with high mobility have received tremendous attention due to thickness of few molecular layers. However, ultrathin 2DONs with high luminescence efficiency and flexibility simultaneously are rarely reported. Here, the ultrathin 2DONs (thickness: 19 nm) through the modulation of tighter molecular packing (distance: ≈3.31 Å) achievable from the incorporation of methoxyl and dipenylamine (DPA) groups into 3D spirofluorenexanthene (SFX) building blocks is successfully prepared. Even with closer molecular stacking, ultrathin 2DONs still enable the suppression of aggregation quenching to exhibit higher quantum yields of blue emission (Φ F  = 48%) than that on amorphous film (Φ F  = 20%), and show amplified spontaneous emission (ASE) with a mediate threshold (332 mW cm -2 ). Further, through drop-casting method, the ultrathin 2DONs are self-organized into large-scale flexible 2DONs films (1.5 × 1.5 cm) with the low hardness (H: 0.008 Gpa) and low Young's modulus (E r : 0.63 Gpa). Impressively, the large-scale 2DONs film can realize electroluminescence performances with a maximum luminance (445 cd m -2 ) and low turn on voltage (3.7 V). These ultrathin 2DONs provide a new avenue for the realization of flexible electrically pumping lasers and intelligent quantum tunneling systems., (© 2023 Wiley-VCH GmbH.)

Background: Evidence about safety and efficacy of transcatheter aortic valve replacement (TAVR) with the Venus A-Valve system (Venus Medtech, Hangzhou, China) remains limited for patients with pure native aortic regurgitation (PNAR)., Objectives: The single-center study sought to report the one-year clinical outcomes of the Venus A-Valve in the treatment of PNAR., Methods: This study was a retrospective analysis of prospectively collected data. Data was from all consecutive patients who had PNAR and underwent TAVR with the Venus A-Valve system at our center from July 2020 and June 2021. Procedural and clinical outcomes up to one year were analyzed using Valve Academic Research Consortium-2 criteria., Results: A total of 45 consecutive patients with PNAR underwent transfemoral TAVR with the Venus A-Valve system. The Mean age was 73.5 ± 5.5 years and 26.7% were female. All the TAVR procedures were performed via transfemoral access. Implantations were successful in 44 cases (97.8%). Only one patient was converted to surgical aortic valve replacement. No patient died intraoperatively. No second valve was implanted. In-hospital mortality rate was 2.3%. The one-year all-cause mortality rate was 4.7% without cardiovascular related death. No patient had moderate or severe paravalvular leakage during follow-up. At one year, the mean pressure gradient was 8.8 ± 0.9 mmHg, and left ventricular ejection fraction increased to 61.5 ± 3.6%., Conclusions: This single-center study demonstrated the safety and efficacy of transfemoral TAVR with the Venus A-Valve in the treatment of patients with PNAR., (© 2023. The Author(s).)

Background: Reconstruction of the aortic arch and its three supra-aortic vessels remains a great surgical challenge with postoperative complications. We present a simplified total arch reconstruction with a modified stent graft (s-TAR) and compared its operative outcomes with conventional total arch replacement (c-TAR)., Methods: This retrospective analysis of prospectively collected data from all consecutive patients who had ascending aortic aneurysm with extended aortic arch dilation and underwent simultaneous ascending aorta replacement and aortic arch reconstruction with the s-TAR or c-TAR between 2018 and 2021. The indication for intervention was maximum diameter of ascending aorta >55 mm and aortic arch in zone II >35 mm., Results: A total of 84 patients were analyzed: 43 in the s-TAR group and 41 in the c-TAR group. No inter-group differences were found for sex, age, comorbidities, or EuroSCORE II results. All patients were successfully treated with s-TAR or c-TAR, and none died intraoperatively. Cardiopulmonary bypass, selective cerebral perfusion, and lower-body circulatory arrest time were significantly shorter in the s-TAR group, which also had a lower incidence of prolonged ventilation and transient neurologic dysfunction. No patient in either group experienced permanent neurologic dysfunction. The incidence of recurrent laryngeal nerve injury and paraplegia was markedly increased in the c-TAR group; however, no such events were observed in the s-TAR group. Both perioperative blood loss and the incidence of reoperation for bleeding were significantly lower in the s-TAR group. The in-hospital mortality rate was 0% in the s-TAR group and 4.9% in the c-TAR group. The s-TAR group had significantly shorter intensive care unit (ICU) stay and lower total hospitalization costs., Conclusions: The s-TAR technique is a safe and effective alternative for total arch reconstruction with shorter operation time, lower rate of postoperative complications and lower total hospitalization costs compared with c-TAR., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://jtd.amegroups.com/article/view/10.21037/jtd-22-1283/coif). The authors have no conflicts of interest to declare., (2023 Journal of Thoracic Disease. All rights reserved.)

Mechanistic models play a crucial role in the process development and optimization of ion-exchange chromatography (IEC). Recent researches in steric mass action (SMA) model have heightened the need for better estimation of nonlinear parameter, steric shielding factor σ. In this work, a straightforward approach combination of simplified linear approximation (SLA) and inverse method (IM) was proposed to initialize and further determine σ, respectively. An existed, unique, and positive σ can be derived from SLA. Compared with linear approximation (LA) developed in our previous study, σ of the multi-component system can be calculated easily without solving the complex system of linear equations, leading to a time complexity reduction from O(n 3 ) to O(n). The proposed method was verified first in numerical experiments about the separation of three charge variants. The calculated σ was more reasonable than that of LA, and the error of elution profiles with the parameters estimated by SLA+IM was only one-sixth of that by LA in numerical experiments. Moreover, the error accumulation effect could also be reduced. The proposed method was further confirmed in real-world experiments about the separation of monomer-dimer mixtures of monoclonal antibody. The results gave a lower error and better physical understanding compared to LA. In conclusion, SLA+IM developed in the present work provides a novel and straightforward way to determine σ. This simplification would help to save the effort of calibration experiments and accelerate the process development for the multi-component IEC separation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Continuous process is a promising alternative for tradition batch process in biomanufacturing, which has higher productivity and lower material consumption. However, despite the maturation of necessary technologies for continuous process, there are few discussion about optimization of full continuous process. One possible reason is that the continuous process is such a complex and interacted process that the traditional experiment-based optimization approach is not sufficient anymore. To address that problem, the process simulation tool SuperPro Designer and continuous capture chromatography model were integrated into a model-assisted design platform for better development of continuous process. The influences of different continuous capture operation modes and sub-lot number under various upstream conditions were analyzed for pilot-scale production. The best combination of operation mode and sub-lot number were determined for the highest equipment utilization without any conflict. After the process optimization, the equipment utilization of continuous process was significantly improved to 27.3%. Then, a pilot-scale case study was carried out to validate the proposed approach. The results showed that the scaling up and process design of continuous process were successful. No time conflict and process failure happened and the final product met the release specification. Finally, the cost of goods was evaluated with SuperPro Designer, and the results showed a 17.4% cost reduction for optimized continuous downstream process compared to batch process, which is promoting for the future industrial applications., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The biopharmaceutical market is driving the revolution from traditional batch processes to continuous manufacturing for higher productivity and lower costs. In this work, a batch mAb downstream process has been converted into an integrated continuous process with the combination of multiple techniques. For process intensification, two batch mode unit operations (protein A capture chromatography, ultrafiltration/diafiltration) were converted into continuous ones; for continuity, surge tanks were used between adjacent steps, and level signals were used to trigger process start or stop, forming a holistic continuous process. For process automation, manual operations (e.g., pH and conductivity adjustment) were changed into automatic operation and load mass was controlled with process analytical technology (PAT). A model-based simulation was applied to estimate the loading conditions for the continuous capture process, resulting in 21% resin capacity utilization and 28% productivity improvement as compared to the batch process. Automatic load mass control of cation exchange chromatography (CEX) was achieved through a customized in-line protein quantity monitoring system, with a difference of less than 1.3% as compared to off-line analysis. Total process time was shortened from 4 days (batch process) to less than 24 hours using the continuous downstream process with the overall productivity of 23.8 g mAb per day for the bench-scale system. Comparable yield and quality data were obtained in three test runs, indicating a successful conversion from a batch process to a continuous process. The insight of this work could be a reference to other similar situations., (© 2022 Wiley-VCH GmbH.)

Short peptide biomimetic chromatography technology as a developing protein separation technology has huge potential for antibody purification. In this study, four tetrapeptide ligands (Ac-FYKH, Ac-YEHF, Ac-YFLH and Ac-FYHI) with high potential binding ability to antibody were selected for the optimal ligand to antibody purification. The results showed that Ac-YEHF-4FF resin had higher binding capacity and selectivity for hIgG among the four resins. And at pH 7.0 and 0.3 ml/min, the highest Q 10%-hIgG of Ac-YEHF-4FF resin was 26.2 mg/ml resin while its Q 10%-BSA was just 2.2 mg/ml resin. Further, Ac-YEHF-4FF resin was used to purify protein mixtures. By binding at pH 7.0 and being eluted at pH 5.0 and pH 4.0, Ac-YEHF-4FF resin was well used to separate hIgG from BSA containing feedstock, HSA containing feedstock and human serum with the purity and yield both more than 95 %. And the screened resin could also separate mAb from CHO cell culture supernatant with purity 94.3 % and yield 97.5 %. The adsorption and separation results of Ac-YEHF-4FF resin indicated that the goal of getting the efficacy of critical residues from protein A to biomimetic its structure and function could be achieved, which had great significance to the establishment and improvement of tetrapeptide biomimetic chromatography, and also provided a new method for the field of antibody separation and purification., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

Ion exchange chromatography (IEC) is one of the most widely-used techniques for protein separation and has been characterized by mechanistic models. However, the time-consuming and cumbersome model calibration hinders the application of mechanistic models for process development. A new methodology called "parameter-by-parameter method (PbP)" was proposed with mechanistic derivations of the steric mass action (SMA) model of IEC. The protocol includes four steps: (1) first linear regression (LR1) for characteristic charge; (2) second linear regression (LR2) for equilibrium coefficient; (3) linear approximation (LA) for shielding factor; (4) inverse method (IM) for kinetic coefficient. Four SMA parameters could be one-by-one determined in sequence, reducing the number of unknown parameters per species from four to one, and predicting almost consistent retention. Numerical single-component experiments were investigated firstly, and the PbP method showed excellent agreement between experiments and simulations. The effects of loadings on the PbP and Yamamoto methods were compared. It was found that the PbP method had higher accuracy and robustness than the Yamamoto method. Moreover, a five-experiment strategy was suggested to implement the PbP method, which is straightforward to reduce the cost of calibration experiments. Finally, a real-world multi-component separation was challenged and further confirmed the feasibility of the PbP method. In general, the proposed method can not only reliably estimate the SMA parameters with comprehensive physical understanding but also accurately predict retention over a wide range of loading conditions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Multi-column counter-current chromatography is an advanced technology used for continuous capture processes to improve process productivity, resin capacity utilization and product consistency. However, process development is difficult due to process complexity. In this work, some general and convenient guidances for three-column periodic counter-current chromatography (3C-PCC) were developed. Boundaries and distributions of operating windows of 3C-PCC processes were clarified by model-based predictions. Interactive effects of feed concentration (c 0 ), resin properties (q max and D e ), recovery and regeneration times (t RR ) were evaluated over a wide range for maximum productivity (P max ). Furthermore, variation of P max was analyzed considering the constraint factors (capacity utilization target and flow rate limitation). The plateau value of P max was determined by q max and t RR . The operating conditions for P max were controlled by q max , t RR and c 0 interactively, and a critical concentration existed to judge whether the operating conditions of P max under constraints. Based on the comprehensive understanding on 3C-PCC processes, a model-free strategy was proposed for process development. The optimal operating conditions could be determined based on a set of breakthrough curves, which was used to optimize process performance and screen resins. The approach proposed was validated using monoclonal antibody (mAb) capture with a 3C-PCC system under various mAb and feed concentrations. The results demonstrated that a thorough model-based process understanding on multi-column counter-current chromatography is important and could improve process development and establish a model-free strategy for more convenient applications., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The advancement of recombinant virus-like particle-based vaccines has attracted global attention owing to substantially safety and high efficacy in provoking a protective immunity against various chronic and infectious diseases in humans and animals. A robust, low-cost, and scalability separation and purification technology is of utmost importance in the downstream processing of recombinant virus-like particles to produce affordable and safe vaccines. Being a relatively simple, environmentally friendly, and efficient biomolecules recovery approach, aqueous two-phase systems have received great attention from researchers worldwide. This review aims to highlight the challenges and outlook in addition to the current applications of aqueous two-phase systems in downstream processing of virus-like particles. The efforts will confidently reinforce scholars' knowledge and fill in the valuable research gap in the aspect of concerning recombinant virus-like particle-based vaccines development, particularly related to the virus-like particles downstream production processes., (© 2022 Wiley-VCH GmbH.)

The cellulase cocktail of marine Aspergillus niger exhibited salt-tolerant and thermostable properties, which is of great potential in industrial application. In order to excavate the single tolerant cellulase components from complex cellulase cocktail, constitutive homologous expression was employed for direct obtainment of the endoglucanase (AnEGL). Enzymatic property study revealed that AnEGL exhibited a property of salt tolerance and a strong thermostability in high salinity environment. Significantly, its activity increased to 129% and the half-life at 65 °C increased to 27.7-fold with the presence of 4.5 M NaCl. Molecular dynamics simulation revealed that Na + and Cl - could form salt bridges with charged residues, and then influenced the activity of loops and the stability of substrate binding pocket, which accounted for the salt tolerance and thermostability. Further, site-specific mutagenesis study proved that the residues Asp95 and Asp99 in the pocket were of great concern for the tolerant properties. The salt-tolerant and thermostable AnEGL was of great value in lignocellulosic utilization and the conjectural mechanisms were of referential significance for other tolerant enzymes., (© 2022. The Author(s).)

Continuous chromatography is a promising technology for downstream processing of biopharmaceuticals. The operation of continuous processes is significantly different to batch-mode chromatography and needs comprehensive evaluation. In this work, the performances of four Protein A affinity resins were studied systematically for twin-column continuous capture processes. A model-based approach was used to evaluate the process performance (productivity and capacity utilization) under varying operation conditions, and the objective was to reveal the crucial resin properties for continuous capture. The trade-off between productivity and capacity utilization was found, and it is necessary to select appropriate resins for different feedstock and operation conditions. The capacity utilization heavily depends on mass transfer, and steep breakthrough curves are favorable for high capacity utilization. The productivity is determined by both equilibrium binding capacity and mass transfer, and the balance of feed amount and feed time is critical. Moreover, the influence of binding capacity and mass transfer on process productivity and parameter sensitivity with two important resin properties (equilibrium binding capacity q max and effective pore diffusion coefficient D e ) were assessed by the model, and suitable resin parameter ranges for twin-column continuous capture were determined. The model-based approach is an effective and useful tool to evaluate the complex performance of different resins and guide the design of next-generation resins for continuous processes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier B.V.)

Purpose: The association between the process of postoperative pneumonia and lung cancer recurrence remains elusive in lung cancer surgery. Herein, the association between postoperative pneumonia and lung cancer recurrence was investigated, emphasizing the warning role of postoperative specific pneumonia in primary lung cancer resection patients., Methods: The occurrence of postoperative pneumonia was assessed in 4-6 months (PPFS), 7-12 months (PPST), and lung cancer recurrence within 1 year (LRO) in 332 patients. The primary outcome was the development of PPST and LRO according to PPFS occurrence. The relevant risk factors of PPFS, PPST, and LRO were identified through multivariable regression analysis., Results: During follow-up, 151 (45.48%) participants experienced PPFS. Irrespective of the existing postoperative pneumonia in 1-3 months (PPOT), PPFS significantly increased the risk of PPST (P < 0.01) and LRO (P < 0.01), and persistent PPST further increased the risk of LRO (P < 0.001). The generalized estimating equation identified chemotherapy as an independent risk factor for PPFS and PPST., Conclusion: PPFS was associated with the increased risk of PPST and LRO. Postoperative pulmonary inflammation assessed 4 months post-surgery also significantly influenced LRO development, indicating a need for close follow-up of lung inflammatory conditions to improve patient outcomes., Competing Interests: The authors report no conflicts of interest in this study., (© 2021 Lin et al.)

The increasing demand for efficient and robust processes in the purification of monoclonal antibodies (mAbs) has recently brought frontal chromatography to the forefront. Applied during the polishing step, it enables the removal of high molecular weight aggregates from the target product, achieving high purities. Typically, this process is operated in batch using a single column, which makes it intrinsically subjected to a purity-yield tradeoff. This means that high purities can only be achieved at the cost of lowering the product yield and vice versa. Recently, a two-column continuous implementation of frontal chromatography, referred to as Flow2, was developed. Despite being able of alleviating the purity-yield tradeoff typical of batch operations, the increase in the number of process parameters complicates its optimal design, with the risk of not exploiting its full potential. In this study, we developed an ad hoc design procedure (DP) suitable for the optimization of both batch frontal chromatography and Flow2 in terms of purity, yield, and productivity. This procedure provided similar results as a multiobjective optimization based on genetic algorithm but with lower computational effort. Then, batch and Flow2 operated at their optimal conditions were compared. Besides showing a more favorable Pareto front of yield and productivity at a specified purity, the Flow2 process demonstrated improved robustness compared to the batch process with respect to modifications in the loading linear velocity, washing buffer ionic strength and loading time, thus providing an appealing operation for integrated processes., (© 2021 Wiley Periodicals LLC.)

Continuous capture with affinity chromatography is one of the most important units for continuous manufacturing of monoclonal antibody (mAb). Due to the complexity of three-column periodic counter-current chromatography (3C-PCC), three approaches (experimental, model-based, and simplified approaches) were studied for process development and optimization. The effects of residence time for interconnected load (RT C ), breakthrough percentage of the first column for interconnected load (s) and feed protein concentration (c 0 ) on productivity and capacity utilization were focused. The model-based approach was found superior to the experimental approach in process optimization and evaluation. Two phases of productivity were observed and the optimal RT C for the maximum productivity was located at the boundary of the two phases. The comprehensive effects of the operating parameters (RT C , s, and c 0 ) were evaluated by the model-based approach, and the operation space was predicted. The best performance of 34.5 g/L/h productivity and 97.6% capacity utilization were attained for MabSelect SuRe LX resin under 5 g/L concentration at RT C  = 2.8 min and s = 87.5%. Moreover, a simplified approach was suggested to obtain the optimal RT C for the maximum productivity. The results demonstrated that model-assisted tools are useful to determine the optimum conditions for 3C-PCC continuous capture with high productivity and capacity utilization., (© 2021 Wiley Periodicals LLC.)

Background: Doxorubicin is a first-line chemotherapy agent on human myelogenous leukemia clinical treatment, but the development of chemoresistance has largely limited curative effect. In this study, we aimed to evaluate the biological function and molecular mechanisms of CrkL to Doxorubicin resistance., Methods: Quantitative reverse transcription-PCR (qRT-PCR) assay was performed to examine the expression of CrkL in K562 and K562/ADR cells. The expression of CrkL was silenced through RNA interference technology. MTT assay and flow cytometry were performed to detect the proliferation inhibition and apoptosis rate after CrkL siRNA transfection. The protein expression changes of PI3K/AKT/MRP1 pathway induced by CrkL siRNA were observed by Western Blot assay. Xenograft tumor model was carried out to observe tumor growth in vivo., Results: We observed that silencing of CrkL could effectively increase apoptosis rate induced by doxorubicin and dramatically reversed doxorubicin resistance in K562/ADR cells. Further studies revealed knockdown CrkL expression suppressed PI3K/Akt/MRP1 signaling, which indicated CrkL siRNA reversed doxorubicin effect through regulating PI3K/Akt/MRP1 pathway. In addition, overexpression of MRP1 could evidently reduce apoptosis rate and reversed the inhibitory effects of doxorubicin resistance caused by CrkL siRNA on K562/ADR cells. Finally, in vivo experiments revealed that CrkL silencing acted a tumor-suppressing role in myelogenous leukemia via regulating PI3K/Akt/MRP1 signaling., Conclusion: Together, we indicated that CrkL is up-regulated in myelogenous leukemia cells and silencing of CrkL could reverse Doxorubicin resistance effectively. These results show a potential novel strategy for intervention chemoresistance in myelogenous leukemia during chemotherapy., (© 2021 The Authors. Journal of Clinical Laboratory Analysis published by Wiley Periodicals LLC.)

Downstream processing of mAb charge variants is difficult owing to their similar molecular structures and surface charge properties. This study aimed to apply a novel twin-column continuous chromatography (called N-rich mode) to separate and enrich acidic variants of an IgG1 mAb. Besides, a comparison study with traditional scaled-up batch-mode cation exchange (CEX) chromatography was conducted. For the N-rich process, two 3.93 mL columns were used, and the buffer system, flow rate and elution gradient slope were optimized. The results showed that 1.33 mg acidic variants with nearly 100% purity could be attained after a 22-cycle accumulation. The yield was 86.21% with the productivity of 7.82 mg/L/h. On the other hand, for the batch CEX process, 4.15 mL column was first used to optimize the separation conditions, and then a scaled-up column of 88.20 mL was used to separate 1.19 mg acidic variants with the purity of nearly 100%. The yield was 59.18% with the productivity of 7.78 mg/L/h. By comparing between the N-rich and scaled-up CEX processes, the results indicated that the N-rich method displays a remarkable advantage on the product yield, i.e. 1.46-fold increment without the loss of productivity and purity. Generally, twin-column N-rich continuous chromatography displays a high potential to enrich minor compounds with a higher yield, more flexibility and lower resin cost., Competing Interests: The authors have declared no conflict of interest., (© 2021 The Authors. Engineering in Life Sciences published by Wiley‐VCH GmbH.)

Meat, except marine sources, is a highly nutritious food but generally lacks some healthy ingredients, such as omega-3 fatty acids (ω-3 FA) and dietary fiber. However, ω-3 FA and dietary fiber could be incorporated during the manufacture of surimi-like products. In our previous study, chicken surimi was successfully developed from spent-hen breast. Although there was no (P > 0.05) difference in water-holding capacity between wheat fiber and carrageenan, an increased (P < 0.05) flaxseed oil-holding capacity was observed in wheat fiber samples. Furthermore, an addition of 5% wheat fiber resulted in optimal emulsification capacity and less cooking loss at 4°C for 14 d and at -20°C for 60 d (P < 0.05). Because of the lower (P < 0.05) purge and centrifugation losses, thiol group content, and thiobarbituric acid reactive substance value than those formulated with more flaxseed oil, 12% flaxseed oil was an optimal level in chicken surimi with 5% wheat fiber. Scanning electron microscopy results also showed better emulsification of surimi batters with wheat fiber compared with those without wheat fiber, and meanwhile, the formulation with 5% wheat fiber could hold up to 12% flaxseed oil as well. To enhance flaxseed-oil addition, semi-manufactured chicken surimi batter was successfully fortified with a combination of 12% flaxseed oil and 5% wheat fiber. This surimi-like product with dietary fiber and ω-3 FA would fit the need in the current market regarding consumers' demands for high nutritional value and improved processing characteristics., (Copyright © 2020. Published by Elsevier Inc.)

Continuous bioprocessing is a promising trend in biopharmaceutical production, and multi-column continuous chromatography shows advantages of high productivity, high resin capacity utilization, small footprint, low buffer consumption and less waste. Due to the complexity and dynamic nature of continuous processing, traditional experiment-based approaches are often time-consuming and inefficient. In this review, model-assisted approaches were focused and their applications in continuous chromatography process development, validation and control were discussed. Chromatographic models are useful in describing particular process performances of continuous capture and polishing with multi-column chromatography. Model-assisted tools showed powerful ability in evaluating multiple operating parameters and identifying optimal points over the entire design space. The residence time distribution models, model-assisted process analytical technologies and model-predictive control strategies were also developed to reveal the propagation of disturbances, enhance real time monitor and achieve adaptive control to match the transient disturbances and deviations of continuous processes. Moreover, artificial neural networks and machine learning concepts were integrated into modeling approaches to improve data treatment. In general, further development in research and applications of model-assisted approaches for continuous chromatography are needed urgently to support the continuous manufacturing., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Postoperative functional training for fracture or osteoarthritis is mainly focused on functional self-exercise, which aims to recover the function of the lower limbs., Purpose: To compare the function and life quality recovery in patients with fracture or arthritis treated with novel muscle training device (NMT) or conventional rehabilitation training (CRT) following surgery., Patients and Methods: A total of 32 fracture patients were randomly divided into the NMT or the CRT groups. The evaluation was performed on the first and 7th day after surgery. The outcome measurements included the incidence of foot drop, Deep Vein Thrombosis and pressure ulcers, Hospital for Special Surgery knee score (HSS scores), pain scores for the Visual Analogue Scale (Pain scores for VAS), Zung self-rating anxiety scale (SAS), Pittsburgh sleep quality index (PSQI) and the Barthel Index score., Results: The comparison of the change scores between the two groups indicated significant differences on day 7 following surgery in the Barthel Index score (P<0.01). The Pain scores for VAS between the two groups indicated a significant difference (P<0.05, U=20.0). The HSS scores between the two groups indicated a significant difference (P<0.05, U=19.0). The HSS scores exhibited a highly significant difference in the NMT group (P<0.01). The Mann-Whitney test was used to analyze the various components of the HSS scores. The comparison of the change scores on the function between the two groups indicated a significant difference (P<0.05). The Range of Motion difference between groups exhibited highly significant differences (P<0.01)., Conclusion: The novel muscle training device positively influenced the decrease in pain score, which resulted in a range increase of knee joint movement and a significant overall improvement in motion., Competing Interests: The authors report no conflicts of interest in this work., (© 2020 Cai et al.)

Human serum albumin is widely used in clinical practice, and the development of new ligands with high affinity is beneficial to improve its separation efficiency. The Site II of human serum albumin is an active binding site of various molecules such as l-tryptophan, which was studied with molecular simulation to obtain insights for the design of new ligands. The results showed that the carboxyl and indolyl groups of l-tryptophan were critical for the binding on Site II. Seven ligands containing carboxyl groups and indolyl groups were designed, and molecular simulation showed that indole-3-pentanoic acid was the best ligand. A new ligand combined indole-3-acetic acid and cysteine was designed for easier resin preparation, and molecular simulation also indicated that the new ligand bound strongly to Site II. Resins with the new ligand designed was prepared and static adsorption experiments indicated that the new resin had high adsorption capacity of human serum albumin and strong salt tolerance. Finally, recombinant human serum albumin was separated from yeast broth with high purity of 90.4% and recovery of 94.2%, which indicated that the new resin had good adsorption selectivity and strong potential for applications., (© 2020 Wiley-VCH GmbH.)

Short peptide biomimetic affinity chromatography as a novel antibody separation chromatography is a potential alternative to protein A chromatography. However, if directly attaching ligand to matrix, the adsorption capacity and mass transfer rate would be affected by pore blockage and steric effect. Grafting resin is an effective method to solve this problem by using polymer as a bridge between matrix and ligand. In this work, a novel resin was prepared by grafting a tetrapeptide to the dextran-grafted matrix. Then, the adsorption properties for human immunoglobin G and BSA were determined. The results showed the saturation adsorption capacity could reach to 133 mg/g resin at pH 8.9 with a significantly low dissociation constant (0.03 mg/mL). The influence of flow rates to dynamic binding capacity of this resin was less than that of the non-grafted resin. The separation performance of the resin showed monoclonal antibody could be well isolated from the Chinese hamster ovary culture supernatant at pH 9.0 with the purity of 93.0% and yield of 84.7% by one step. Overall, this resin could achieve higher binding capacity by the possible of gaining higher ligand density, indicating its potential significance for separation in larger scale systems., (© 2020 Wiley-VCH GmbH.)

Multi-column continuous chromatography has advantages of high resin capacity utilization and productivity, low buffer consumption and small footprint. Experimental optimization is often time-consuming and inefficient due to the complexity of continuous processes. In this study, a model-based approach was investigated to improve process development of twin-column continuous capture with Protein A affinity resin MabSelect PrismA. Breakthrough curves under various conditions, productivity and capacity utilization (CU) of the continuous processes under varying operating conditions were predicted. Effects of three key operating parameters (feed concentration (c 0 ), interconnected feed residence time (RT) and breakthrough percentage control of the first column during interconnected feeding (s)) on the productivity and CU were evaluated. A recommended working window can be determined directly from contour maps to balance the trade-off between productivity and CU. The model-optimized operating conditions at varying feed concentrations were verified by experiments, which indicated that the model-based approach was feasible and reliable. The results showed that the suitable RT was 1~2 min and suitable s was 0.6~0.75 for the continuous IgG capture with MabSelect PrismA. The maximum productivity varied from 14 to 47 g/L/h with the feed IgG concentrations at the range of 1 to 10 mg/mL. The results indicated that model-based approach could assist process development efficiently and promote target-orientated process design for continuous processes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Multi-column periodic counter-current chromatography (PCC) has been developed for continuous antibody capture, but the complexity of continuous processes makes experimental optimization time consuming and costly. In this work, with twin-column continuous system as a typical case, mathematical models were established and used to evaluate the impacts of operating parameters for process development. The model fitted well with the experimental breakthrough curves and process performance under varying protein concentrations and residence times. Three important operating parameters, residence time for interconnected feeding (RT C ), breakthrough percentage control for interconnected feeding (s) and disconnected feeding time (t DC ), were evaluated systematically. The profiles of productivity and resin capacity utilization showed three phases as a function of RT C , which resulted in different optimization strategies towards s and t DC . Based on the model prediction, a working window of RT C and s can be determined for process development. Finally, a model-based design approach was proposed to determine the optimum operating conditions and to design a suitable continuous process for high productivity and capacity utilization. With the model-based design approach developed, the best performance of 12.8 g/L/h productivity and 91.9% capacity utilization was found for MabSelect SuRe resin under 1 mg/mL feeding IgG concentration at RT C  = 2 min, s = 65% and t DC  = 26 min., Competing Interests: Declaration of Competing interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier B.V.)

Hippocampal neurons undergo various forms of cell death after status epilepticus. Necrostatin-1 specifically inhibits necroptosis mediated by receptor interacting protein kinase 1 (RIP1) and RIP3 receptors. However, there are no reports of necroptosis in mouse models of status epilepticus. Therefore, in this study, we investigated the effects of necrostatin-1 on hippocampal neurons in mice with status epilepticus, and, furthermore, we tested different amounts of the compound to identify the optimal concentration for inhibiting necroptosis and apoptosis. A mouse model of status epilepticus was produced by intraperitoneal injection of kainic acid, 12 mg/kg. Different concentrations of necrostatin-1 (10, 20, 40, and 80 μM) were administered into the lateral ventricle 15 minutes before kainic acid injection. Hippocampal damage was assessed by hematoxylin-eosin staining 24 hours after the model was successfully produced. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining, western blot assay and immunohistochemistry were used to evaluate the expression of apoptosis-related and necroptosis-related proteins. Necrostatin-1 alleviated damage to hippocampal tissue in the mouse model of epilepsy. The 40 μM concentration of necrostatin-1 significantly decreased the number of apoptotic cells in the hippocampal CA1 region. Furthermore, necrostatin-1 significantly downregulated necroptosis-related proteins (MLKL, RIP1, and RIP3) and apoptosis-related proteins (cleaved-Caspase-3, Bax), and it upregulated the expression of anti-apoptotic protein Bcl-2. Taken together, our findings show that necrostatin-1 effectively inhibits necroptosis and apoptosis in mice with status epilepticus, with the 40 μM concentration of the compound having an optimal effect. The experiments were approved by the Animal Ethics Committee of Fujian Medical University, China (approval No. 2016-032) on November 9, 2016., Competing Interests: None

Biomimetic affinity chromatography with short peptide ligands is a developing technology, which has great potential for antibody separation and purification. In this study, a tetrapeptide library with critical residues of natural ligands to hIgG was constructed and a novel tetrapeptide ligand (Ac-FYHE) with high LibDock scores was selected by molecular docking. Then, Ac-FYHE ligand was linked to agarose bead to prepare a new chromatography resin. The properties of antibody adsorption were measured and evaluated by static/dynamic adsorption. It was found that the resin with ligand Ac-FYHE has high binding capacity and selectivity for hIgG. The results showed the Q m-hIgG of Ac-FYHE-4FF resin was 87.9 mg/g resin while the Q m-BSA of this resin was only 16.5 mg/g resin at pH 7.0. Moreover, at pH 7.0, Q 10% of Ac-FYHE-4FF resin was 24.1 mg/mL for hIgG but just 2.1 mg/mL for BSA, which presented high selectivity of the screened resin at pH 7.0. Subsequently, the adsorption and separation properties of the Ac-FYHE-4FF resin were further investigated. As a result, with the addition of 0.5 M NaCl, Q m decreased by less than 20% but Q m decreased by 70% with the addition of 50% (v/v) ethylene glycol, which indicated that hydrophobic interaction would be the driving force for the binding between resin and hIgG. Besides, pH 7.5 and pH 4.5 could be the optimal loading and elution condition for hIgG, respectively. Finally, the Ac-FYHE-4FF resin was applied to separate mAb or/and hIgG from BSA containing feedstock, CHO cell culture supernatant and human serum, and the purity and recovery were both more than 90% with only one-step separation. The results indicate that the Ac-FYHE-4FF resin developed in this work would be promising for antibody separation and purification., (Copyright © 2019. Published by Elsevier B.V.)

Using density functional theory calculations and photoemission measurements, we have studied the interaction between the non-fullerene small-molecule acceptor ITIC and K atoms (a representative of reactive metals). It is found that the acceptor-donor-acceptor-type geometric structure and the electronic structure of ITIC largely decide the interaction process. One ITIC molecule can combine with more than 20 K atoms. For stoichiometries K x ≤6 ITIC, the K atoms are attracted to the acceptor units of the molecule and donate their 4s electrons to the unoccupied molecular orbitals. K-ITIC organometallic complexes, characterized by the breaking of some S-C bonds in the donor unit of ITIC and the formation of K-S bonds, are formed for stoichiometries K x ≥7 ITIC. The complexes are still conjugated despite the breaking of some S-C bonds., Competing Interests: The authors declare no competing financial interest.

Three pyrene-based spirocyclic aromatic hydrocarbons (Py-SAHs) were prepared to clarify the roles of molecular segments in regulating the morphologies and photophysical properties of organic microcrystals. Due to the different supramolecular steric hindrance (SSH) effect between bulky groups and pyrene rings, distinct nanocrystal morphologies with unique photoluminescence properties were realized.

The hydrolysis step by β-glucosidase (BGL) is generally recognized as the major limiting step in cellulose degradation and the BGLs with prominent enzymatic properties are of great importance for efficient utilization of lignocellulosic biomass. In order to identify some salt-tolerant BGLs, two BGL genes were cloned from marine Aspergillus niger ZJUBE-1 genome. Then two bgl expression cassettes driven by gpdA promoter were respectively transformed into marine A. niger for homologous constitutive expression. Directed expression was achieved for the domination of target BGLs in fermentation broth. Conveniently, two BGLs were purified to homogeneity by two separation steps, ultrafiltration and anion exchange chromatography. The purified BGL1 and BGL2 showed maximum activity at pH 3.0-4.0 and 3.5-4.5, respectively, suggesting these two BGLs were relatively acidophilic, especially for BGL1. Besides, BGL1 was stable to most of metal ions, while BGL2 was sensitive to Cu 2+ , Fe 3+ and Ag + . Most specially, BGL2 activity increased by 44% in the presence of 4 M NaCl, suggesting BGL2 was halophilic. Homology modeling revealed that longer loops and linkers as well as polymerous Glu 492 may contribute to the halophilism of BGL2. At last, the medium for directed expression was optimized and the content as well as the purity of target protein was improved., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objective: Atherosclerosis is a major risk factor for cardiovascular disease, but its mechanism of progression remained unclear. However, many long non-coding RNAs (lncRNAs) have recently been implicated in different processes for cardiovascular disease. In this study, we mainly focused on the role of lncRNA TUG1 in atherosclerosis., Patients and Methods: qRT-PCR was used to detect the expression of lncRNA TUG1 in atherosclerosis patients and animal model. Moreover, the expression of TUG1 in vascular smooth muscle cell dysfunction model was also measured. Proliferation ability was tested by CCK-8 and cyclin D1 assay, through loss- and gain-of function approaches. Western-blot was used to measure the expression of PTEN, when TUG1 was in different levels., Results: We found that the lncRNA TUG1 was highly expressed in serum samples from 38 patients with atherosclerosis, compared with 24 healthy volunteers.  LncRNA TUG1 was dramatically upregulated in atherosclerotic plaques of ApoE-/- mice. We also found that the expression of TUG1 was upregulated in vascular smooth muscle cell injury model. Through loss- and gain-of function approaches, we showed that TUG1 promotes cell proliferation and induces apoptosis in vitro. What's more, TUG1 expression level was reversely correlated with PTEN expression in patients with atherosclerosis. LncRNA TUG1 could compete with PTEN for miR-21 binding., Conclusions: We found that lncRNA TUG1 was closely related to the progression of atherosclerosis, which could be a potential target for treating atherosclerosis.