Continuous Centrifugation Research Articles

Analysis of changes in serum calcium and magnesium levels during plateletpheresis on different apheresis machines in healthy donors: A cross-sectional study

Abstract: BACKGROUND: The most common observed complication of plateletpheresis procedure is hypocalcemia due to citrate’s chelation of free calcium. Magnesium is also affected by citrate infusion. We have evaluated the impact of plateletpheresis on the total serum calcium (Ca++) and magnesium (Mg++) levels of the donor. MATERIALS AND METHODS: One hundred and thirty-seven healthy plateletpheresis donors were included in the study. Samples were collected from the donor at 0 min (baseline), 30 min, and 60 min, at the end of the procedure and 30 min after completion of the procedure. The pattern of changes in serum Ca++ and serum Mg++ levels at different time points for different apheresis machines (Fenwal Amicus, Fresenius COM.TEC, Spectra Optia, and Hemonetics MCS+) was studied. RESULTS: A significant decline in serum Ca++ and serum Mg++ levels from baseline to the end of the procedure was observed. This decline was observed maximum for Hemonetics MCS+. The baseline values were regained 30 min after completion of the procedure. Difference from baseline values 30 min after the procedure was lowest in amicus in the case of serum Ca++ and COM.TEC in case of serum Mg++. Twenty donors (14.60%) experienced adverse donor reactions. CONCLUSION: Serum calcium and serum magnesium levels decreased transiently during the procedure and regained normal values after 30 min of the completion of the plateletpheresis procedure on all four apheresis machines (whether based on continuous or intermittent centrifugation). Hence, it is concluded from our study that plateletpheresis is a well-tolerated and safe procedure.

Effects of aeration and centrifugation conditions on omega-3 fatty acid production by the mixotrophic dinoflagellate Gymnodinium smaydae in a semi-continuous cultivation system on a pilot scale

High production and efficient harvesting of microalgae containing high omega-3 levels are critical concerns for industrial use. Aeration can elevate production of some microalgae by providing CO2 and O2. However, it may lower the production of others by generating shear stress, causing severe cell damage. The mixotrophic dinoflagellate Gymnodinium smaydae is a new, promising microalga for omega-3 fatty acid production owing to its high docosahexaenoic acid content, and determining optimal conditions and methods for high omega-3 fatty acid production and efficient harvest using G. smaydae is crucial for its commercial utilization. Therefore, to determine whether continuous aeration is required, we measured densities of G. smaydae and the dinoflagellate prey Heterocapsa rotundata in a 100-L semi-continuous cultivation system under no aeration and continuous aeration conditions daily for 9 days. Furthermore, to determine the optimal conditions for harvesting through centrifugation, different rotational speeds of the continuous centrifuge and different flow rates of the pump injecting G. smaydae + H. rotundata cells into the centrifuge were tested. Under continuous aeration, G. smaydaeproduction gradually decreased; however, without aeration, the production remained stable. Harvesting efficiency and the dry weights of omega-3 fatty acids of G. smaydae + H. rotundata cells at a rotational speed of 16,000 rpm were significantly higher than those at 2,000–8,000 rpm. However, these parameters did not significantly differ at injection pump flow rates of 1.0–4.0 L min-1. The results of the present study provide a basis for optimized production and harvest conditions for G. smaydae and other microalgae.

Use of plasma exchange as therapeutic tool in clinical practice

ObjectiveTo descrive the use of therapeutic plasma exchange in several pathologies and its adjustment to international reference guides. MethodObservational, descriptive, retrospective study, of all the patients that received plasmapheresis between January 2014-December 2015. We analized the appropriate indication according to the bibliography consulted, and the therapeutic outcome. Indication, replaced volume of plasma, number of sessions and periodicity were established by the Hematology Service depending on the disease and its clinical course. Results10 patients (8 women), between 28-72 years old, received therapeutic plasma exchange. The pathologies treated were neurological (9 patients), Waldenstrom disease (1 patient). The technique used was continuous centrifugation with albumin 5% as replacement fluid. ConclusionsThe therapeutic plasma exchange in reviewed patients agreed to reference guides. There was not a direct relation between the recommendation grade and the response obtained. The reduced number of patients is a limitation to obtain conclusive results.

Continuous cold ethanol precipitation of immunoglobulin G from human plasma

Currently intravenous immunoglobulin G (IVIG) is manufactured by cold ethanol precipitation in batch, which results in a slow process with a high footprint due to the use of large tanks. A batch precipitation method using ethanol for the production of IVIG from “cryo-poor plasma”, a starting material obtained after removal of cryoprecipitate from thawed fresh frozen plasma was rendered into a continuous method using a tubular reactor. This reactor consisted of double jacketed stainless-steel tubes filled with static mixers for efficient cooling, because the heat transfer in stainless steel is very high. The pH adjusted, precooled plasma and ethanol were fed into a tubular reactor and cooled down to − 7 °C and precipitation was induced and completed after having left the reactor. The precipitate at the reactor effluent was harvested by tandem precoat filtration or pseudo continuous batch centrifugation. Yield and purity were in the same range as the batch precipitation method, with 95% yield. Even with a residence time of one minute, complete precipitation with same yield and purity could be achieved. A tubular reactor with a length of 3.5 m and 3.7 cm internal diameter would be suited to process 4000 L of plasma per day. This concept will be the basis to substantially shrink the plant footprint of new plants or be a way to intensify existing manufacturing plants.

Effects of daily artificial gravity training on orthostatic tolerance following 60-day strict head-down tilt bedrest

PurposeOrthostatic intolerance commonly occurs following immobilization or space flight. We hypothesized that daily artificial gravity training through short-arm centrifugation could help to maintain orthostatic tolerance following head-down tilt bedrest, which is an established terrestrial model for weightlessness.MethodsWe studied 24 healthy persons (eight women; age 33.3 ± 9.0 years; BMI 24.3 ± 2.1 kg/m2) who participated in the 60-days head-down tilt bedrest (AGBRESA) study. They were assigned to 30 min/day continuous or 6 × 5 min intermittent short-arm centrifugation with 1Gz at the center of mass or a control group. We performed head-up tilt testing with incremental lower-body negative pressure until presyncope before and after bedrest. We recorded an electrocardiogram, beat-to-beat finger blood pressure, and brachial blood pressure and obtained blood samples from an antecubital venous catheter. Orthostatic tolerance was defined as time to presyncope. We related changes in orthostatic tolerance to changes in plasma volume determined by carbon dioxide rebreathing.ResultsCompared with baseline measurements, supine and upright heart rate increased in all three groups following head-down tilt bedrest. Compared with baseline measurements, time to presyncope decreased by 323 ± 235 s with continuous centrifugation, by 296 ± 508 s with intermittent centrifugation, and by 801 ± 354 s in the control group (p = 0.0249 between interventions). The change in orthostatic tolerance was not correlated with changes in plasma volume.ConclusionsDaily artificial gravity training on a short-arm centrifuge attenuated the reduction in orthostatic tolerance after 60 days of head-down tilt bedrest.

The Impact of Nutrient Limitation and Harvest Method on the Wet Preservation of Chlorella vulgaris Biomass.

The temporary storage of wet algae concentrates enables the decoupling in time of algae harvests and their biorefinery. However, the impact of cultivation and of the harvest conditions on algae quality during preservation is largely unknown. This study aimed to determine the impact of nutrient limitation and of harvest methods on the preservation of Chlorella vulgaris biomass. Algae were either well-fed until harvest or received no nutrients for one week, and were harvested by either batch or continuous centrifugation. The organic acid formation, lipid levels, and lipolysis were monitored. Nutrient limitation had a large impact and resulted in lower pH values (4.9 ± 0.4), high levels of lactic acid and acetic acid, and a slightly higher degree of lipid hydrolysis. Concentrates of well-fed algae had a higher pH (7.4 ± 0.2) and another pattern of fermentation products with mainly acetic acid, succinic acid, and, to a smaller extent, lactic acid and propionic acid. The effect of the harvest method was smaller, with, most often, higher lactic acid and acetic acid levels for algae harvested by continuous centrifugation than for those obtained by batch centrifugation. In conclusion, nutrient limitation, a well-known method to enhance algae lipid levels, can impact several quality attributes of algae during their wet storage.

Single‐use centrifugal separator enables intensification of the clarification process in biomanufacturing of recombinant proteins

AbstractBACKGROUNDThe need to produce more biologics at lower costs and hence intensify bioprocessing has significantly improved upstream productivity (cell culture titers ≥3 g L−1 and cell densities ≥20 × 106 cells mL−1), shifting the economic and production bottleneck to downstream manufacturing. The first downstream unit operation is the clarification of the cell culture broth to remove various insoluble (such as cells and cell debris) and soluble (such as host cell DNA and proteins) contaminants, producing a feed stream suitable for further downstream purification. The high cell‐density cultures obtained in modern intensified bioprocessing have made legacy single‐use clarification trains (based primarily on depth filtration of the cell culture broth) ineffective and impractical, significantly increasing the economic and material burden of the clarification process.RESULTSWe have developed and characterized approaches that enable intensification of the clarification operation, by increasing process capacities while reducing material requirements and footprint, for single‐use manufacturing facilities. These approaches are based on primary clarification using the Alfa Laval CultureOne™ centrifuge (a continuous solid discharge disc‐stack centrifuge with a single‐use flow path) followed by secondary clarification using single‐use disposable depth filters. We show that these intensified clarification trains are suitable for processing high cell‐density cultures (6–12% packed cell volume) with significantly high yields (≥90%) and impurity clearance, along with reduced operating costs (30–60% savings).CONCLUSIONIncorporating a single‐use continuous discharge centrifuge into the clarification train enables clarification of high cell‐density cultures in single‐use manufacturing facilities and enhances process economic and environmental sustainability. © 2023 Society of Chemical Industry (SCI).

Development of Hetero-Cell Type Spheroids Via Core-Shell Strategy for Enhanced Wound Healing Effect of Human Adipose-Derived Stem Cells.

Stem cell-based therapies have been developed to treat various types of wounds. Human adipose-derived stem cells (hADSCs) are used to treat skin wounds owing to their outstanding angiogenic potential. Although recent studies have suggested that stem cell spheroids may help wound healing, their cell viability and retention rate in the wound area require improvement to enhance their therapeutic efficacy. We developed a core-shell structured spheroid with hADSCs in the core and human dermal fibroblasts (hDFs) in the outer part of the spheroid. The core-shell structure was formed by continuous centrifugation and spheroid incubation. After optimizing the method for inducing uniform-sized core-shell spheroids, cell viability, cell proliferation, migration, and therapeutic efficacy were evaluated and compared to those of conventional spheroids. Cell proliferation, migration, and involucrin expression were evaluated in keratinocytes. Tubular assays in human umbilical vein endothelial cells were used to confirm the improved skin regeneration and angiogenic efficacy of core-shell spheroids. Core-shell spheroids exhibited exceptional cell viability under hypoxic cell culture conditions that mimicked the microenvironment of the wound area. The improvement in retention rate, survival rate, and angiogenic growth factors secretion from core-shell spheroids may contribute to the increased therapeutic efficacy of stem cell treatment for skin wounds.

Acoustofluidics for simultaneous droplet transport and centrifugation facilitating ultrasensitive biomarker detection.

Trace biological sample detection is critical for the analysis of pathologies in biomedicine. Integration of microfluidic manipulation techniques typically strengthens biosensing performance. For instance, using isothermal amplification reactions to sense trace miRNA in peripheral circulation lacks a sufficiently complex pretreatment process that limits the sensitivity of on-chip detection. Herein we propose an orthogonal tunable acoustic tweezer (OTAT) to simultaneously actuate the transportation and centrifugation of μ-droplets on a single device. The OTAT enables diversified modes of droplet transportation such as unidirectional transport, multi-direction transport, round-trip transport, tilt angle movement, multi-droplet fusion, and continuous centrifugation of the dynamic droplets simultaneously. The multiplicity of modalities enables the focusing of a loaded analyte at the center of the droplet or constant rotation about the center axis of the droplet. We herein demonstrate the OTAT's ability to actuate transportation, fusion, and centrifugation-based pretreatment of two biological sample droplets loaded with miRNA biomarkers and multiple mixtures, as well as facilitating the increase of fluorescence detection sensitivity by an order of magnitude compared to traditional tube reaction methods. The results herein demonstrate the OTAT-based droplet acoustofluidic platform's ability to combine a wide range of biosensing mechanisms and provide a higher accuracy of detection for one-stop point-of-care disease diagnosis.

Cross‐Batch Contamination in a Continuous Horizontal Decanter Centrifuge during Virgin Olive Oil Production

AbstractCross‐batch contamination in a decanter centrifuge during virgin olive oil production cannot be avoided using current technology. The extent of this contamination is investigated using industrial‐scale tests, by measuring the volatile profile and color on three consecutive oil batches, collected at the decanter outlet at different extraction times. The extent of contamination varied, pointing out qualitative consequences, as defective molecules are found. The latter are often active at low concentrations, and the measured cross‐batch contamination can lead both to the downgrading of large batches of virgin olive oils and to the adulteration of monovarietal and certified productions. An innovative method, based on the direct determination of the color (L and a* coordinates) of oil at the outlet of the decanter is able to identify the same compositional change point indicated by gas chromatography, and could be successfully used to mitigate the effects of cross‐batch contamination.Practical applications: An in‐line colorimetric system can be implemented at the decanter outlet to detect the point of change between different olive batches. Otherwise, the virgin olive oil exiting from the decanter at the beginning of one batch can be collected separately in order to avoid the contamination due to the previous batch.

White-flesh guava juice clarification by a fixed-angle conical rotor centrifuge laboratory and characterization of continuous disk stack centrifuges

The two objectives of this paper were to determine the effect of centrifugation parameters on guava juice physicochemical characteristics and to identify operational characteristics of continuous disc-stack centrifuges based on the performance of a laboratory centrifuge. Effects of g-force (149 g–3731 g) and centrifugation time (10–40 min) on juice physicochemical characteristics (protein, pectin, galacturonic acid, dry matter, total soluble sugars contents; pH; electrical conductivity, clarity and particle size distribution) were assessed. Laboratory centrifuge performance was evaluated for 1343 g, 2388 g and 3731 g. At 1343 g, separation limits (xmax), feed flow-rates of disc-stack centrifuges and cut-off sizes (x50) were determined for corresponding laboratory centrifuge operation times. Significant decrease in average particle size, protein and pectin contents was observed, contrary to juice clarity. Similar clarification efficiency was obtained for g-forces ≥ 1343 g. xmax were 2669 nm, 2200 nm and 1783 nm, with corresponding x50 for continuous centrifuges, 1887 nm, 1556 nm, 1261 nm, for 20 min, 30 min and 40 min, respectively.

Use of ultrafiltration ceramic membranes as a first step treatment for olive oil washing wastewater

Olive oil is a food product in great demand throughout the world, its production generating large volumes of wastewater with a high organic load, where phenolic compounds are present. These compounds have outstanding antioxidant characteristics, so their recovery is of great interest. In this way, the treatment of these wastewaters should be based on reusing water and, at the same time, on recovering valuable compounds. Ultrafiltration with ceramic membranes (5, 15 and 50 kDa) is proposed as a first treatment step for olive oil washing wastewater (OOWW) from the continuous two-phase centrifugation process. The effect of cross flow velocity and transmembrane pressure was evaluated against permeate flux values and the removal of turbidity, colour, chemical oxygen demand (COD), sugars and the phenolic compounds recovery. The CFV had a great influence on the removal of colour and turbidity. COD rejection increased with increasing TMP, being the highest rejection obtained under the most extreme conditions. The rejection of phenolic compounds and sugars did not show great variation between conditions. The 50 kDa membrane was the one that presented the largest permeate flux decline and the lowest permeability recovery after the cleaning, confirming the great fouling that this membrane suffered. The 15 kDa membrane at the operating conditions of 3 m.s-1 and 3 bar was observed as a good option to eliminate much of the colour (72%) and turbidity (99%) and to reduce considerably the organic load (54%) without greatly affecting the concentration of phenolic compounds (rejection of 21%) for future recovery.

Continuous Percoll Gradient Centrifugation of Erythrocytes—Explanation of Cellular Bands and Compromised Age Separation

(1) Background: When red blood cells are centrifuged in a continuous Percoll-based density gradient, they form discrete bands. While this is a popular approach for red blood cell age separation, the mechanisms involved in banding were unknown. (2) Methods: Percoll centrifugations of red blood cells were performed under various experimental conditions and the resulting distributions analyzed. The age of the red blood cells was measured by determining the protein band 4.1a to 4.1b ratio based on western blots. Red blood cell aggregates, so-called rouleaux, were monitored microscopically. A mathematical model for the centrifugation process was developed. (3) Results: The red blood cell band pattern is reproducible but re-centrifugation of sub-bands reveals a new set of bands. This is caused by red blood cell aggregation. Based on the aggregation, our mathematical model predicts the band formation. Suppression of red blood cell aggregation reduces the band formation. (4) Conclusions: The red blood cell band formation in continuous Percoll density gradients could be explained physically by red blood cell aggregate formation. This aggregate formation distorts the density-based red blood cell age separation. Suppressing aggregation by osmotic swelling has a more severe effect on compromising the RBC age separation to a higher degree.

Greener production of cellulose nanocrystals: An optimised design and life cycle assessment

Cellulose nanocrystals (CNCs) are usually prepared by sulphuric acid hydrolysis in industrial production, resulting in a large amount of alkaline and water consumption. In this paper, three acid-CNCs separation methods, including gravity settling, centrifugation and microfiltration, are used to directly separate sulphuric acid from hydrolysate mixture to reduce alkaline and water consumption. Microfiltration was selected as the best method for acid-CNCs separation. Using microfiltration, the highest sulphuric acid recovery ratio was about 65.0%, with an acid concentration range of 8–10 wt% in the pilot-scale CNCs production. Continuous centrifugation was introduced to replace gravity settling to separate CNCs from the neutralised mixture for faster production and reducing water consumption. The consumption of alkaline and water decreased by 63.0% and 68.0%, and the cost was reduced by 26%. The result of life cycle assessment (LCA) showed that the process with acid-CNCs separation using microfiltration achieved a reduction of 33.4% in aquatic ecotoxicity, 38.1% in terrestrial ecotoxicity, 47.9% in aquatic acidification, 55.8% in aquatic eutrophication, 40.0% in global warming potential and 47.8% in non-renewable energy consumptions. This study provided a promising approach for separating sulphuric acid from hydrolysate mixture and reducing consumption of alkaline and water. Such clean and economic process has great potential in the industrial production of CNCs.

Rapidly Deployable Algae Cleaning System for Applications in Freshwater Reservoirs and Water Bodies

Occurrence of large-scale harmful algal blooms (HABs) in our reservoirs and water bodies threaten both quality of our drinking water and economy of aquaculture immensely. Hence, rapid removal of HAB biomass during and after a bloom is crucial in protecting the quality of our drinking water and preserve our water resources. We reported here a rapidly deployable algae cleaning system based on a high-capacity high-throughput (HCHT) spiral blade continuous centrifuge connected with inlet and effluent water tanks and a series of feed-in and feed-out pumps as well as piping, all fitted into a standard 20 feet metal shipping container. The system separates algal biomass from algae-laden water with a maximum flow rate of 4000 L/h and a centrifugal force of 4500× g. Cells collected by the system are still intact due to the low centrifugal force used. We showed that after HCHT centrifugation, cellular contents of HAB biomass were not found in the effluent water, and hence, could be discharged directly back to the water body. Furthermore, the addition of flocculants and chemicals prior to the separation process is not required. The system could operate continuously with proper programmed procedures. Taken overall, this system offered a much better alternative than the traditional flocculation- and sonication-based methods of HAB removal in a freshwater environment. This deployable system is the first of its kind being built and had been field-tested successfully.

P68.10 PFN1 Induces Tumor Metastasis Through Promoting Secretion of Microvesicles By ROCK I/p-MLC Pathway in Non-Small Cell Lung Cancer

Profilin1(PFN1) plays confusing roles in metastasis of different cancers, here we explored the roles of PFN1 in metastasis of non-small cell lung cancer (NSCLC). Considering that PFN1 is an important mediator of membrane trafficking and microvesicles(MVs) have already been proved closely associated with tumor metastasis, we further explored relationship of PFN1 and MVs in NSCLC metastasis and the underlying mechanisms. We first detect PFN1 and p-MLC expression in tissue microarray. Then we extracted MVs of NSCLC patients’ serums by continuous differential centrifugation, quantified MVs by WB, electron microscopy and spectral flow cytometry. And PFN1 was overexpression/ knockdown to study the effect of MVs secretion and cell migration. Protein interaction was tested by CO-IP. ROCK kinase assay was used to detect ROCK kinase activity. For in vivo experiment, intracardiac injection of H1299 cell lines were used to establish metastatic tumor model for in vivo assay. PFN1 were highly expressed in late stage NSCLC tissues, and the expression of PFN1 is positively correlated with p-MLC in IHC. Overexpression of PFN1 promotes MVs secretion and metastasis in vitro. Further investigation found that PFN1 could interact with ROCK I and affect the activity of ROCK I to phosphorylate MLC, which could regulate the secretion of MVs. Inhibition of ROCK I activity by Y27632 partially reverse PFN1’ s promotion of MVs secretion and NSCLC metastasis in vivo and in vitro. PFN1 affects NSCLC metastasis via promoting MVs’ secretion, which provided us a new insight into PFN1’s roles in cancer metastasis and a new target of therapy in NSCLC metastasis.

Abstract P192: Orthostatic Tolerance Before And After 60 Days Of Strict Head Down Tilt Bedrest With And Without Daily Artificial Gravity Training

Background: Orthostatic intolerance occurs after space flight, immobilization and in patients with autonomic diseases, so there is a need for more effective countermeasures. We hypothesized that daily artificial gravity elicited through short-arm centrifugation attenuates plasma volume loss and orthostatic intolerance following 60 days of HDTBR, which models cardiovascular responses to weightlessness. Methods: We studied 24 healthy persons (8 women, 33.4±9.3 yr, 24.3±2.1 kg/m2) exposed to 60d HDTBR. Subjects were assigned to 30 min/d continuous short arm centrifugation (cAG), 6x5 min short arm centrifugation (iAG), or a control group (ctr, no countermeasures). Head-up tilt testing (15 min of 80°) followed by incremental lower body negative pressure (-10 mmHg every 3 min) until presyncope was performed before and at the end of HDTBR. Plasma volume was measured (CO rebreathing) 12-2 days before and after 56d of HDTBR. Stroke volume was measured by cMRI. Norepinephrine, epinephrine, aldosterone, and renine plasma levels were measured before and after HDTBR. Results: Time to presyncope decreased in all groups following bedrest (ctr: 22:56 min pre and 9:35 min post, cAG 15:34 min pre and 10:11 min post; iAG 14:56 min pre and 10:00 min post, p<0.001). The significant interaction (p=0.025) between bedrest and intervention was explained by greater baseline orthostatic tolerance time in the ctr. AG Data was pooled analysis. The reduction in stroke volume (ctr, pre: 93±19 ml, HDTBR: 69±13 ml, AG, pre: 88±20 ml, HDTBR: 67±17 ml) and plasma volume was similar (ctr, pre: 4155±1085 ml, HDTBR: 3855±1087 ml, AG, pre: 4114±1250 ml, HDTBR: 3674± 1313 ml). Catechols and aldosterone did not change significantly during bedrest. The increase in renine was similar between groups (ctr pre: 18±12 mE/L, HDTBR: 21±8 mE/L, AG pre: 21±10 mE/L, HDTBR: 31±12 mE/L). Conclusions: 30 min daily AG didn’t prevent a reduction in orthostatic tolerance following 60d HDTBR. Whether numerically smaller reductions in orthostatic tolerance in the AG groups indicate efficacy or result from baseline differences can’t be ascertained. A stronger AG stimulus or combination with other countermeasures might be required to maintain orthostatic tolerance and to attenuate the volume reduction.

Optic disc edema and chorioretinal folds develop during strict 6° head-down tilt bed rest with or without artificial gravity.

Spaceflight associated neuro‐ocular syndrome (SANS) is hypothesized to develop as a consequence of the chronic headward fluid shift that occurs in sustained weightlessness. We exposed healthy subjects (n = 24) to strict 6° head‐down tilt bed rest (HDTBR), an analog of weightlessness that generates a sustained headward fluid shift, and we monitored for ocular changes similar to findings that develop in SANS. Two‐thirds of the subjects received a daily 30‐min exposure to artificial gravity (AG, 1 g at center of mass, ~0.3 g at eye level) during HDTBR by either continuous (cAG, n = 8) or intermittent (iAG, n = 8) short‐arm centrifugation to investigate whether this intervention would attenuate headward fluid shift‐induced ocular changes. Optical coherence tomography images were acquired to quantify changes in peripapillary total retinal thickness (TRT), retinal nerve fiber layer thickness, and choroidal thickness, and to detect chorioretinal folds. Intraocular pressure (IOP), optical biometry, and standard automated perimetry data were collected. TRT increased by 35.9 µm (95% CI, 19.9–51.9 µm, p < 0.0001), 36.5 µm (95% CI, 4.7–68.2 µm, p = 0.01), and 27.6 µm (95% CI, 8.8–46.3 µm, p = 0.0005) at HDTBR day 58 in the control, cAG, and iAG groups, respectively. Chorioretinal folds developed in six subjects across the groups, despite small increases in IOP. Visual function outcomes did not change. These findings validate strict HDTBR without elevated ambient CO2 as a model for investigating SANS and suggest that a fluid shift reversal of longer duration and/or greater magnitude at the eye may be required to prevent or mitigate SANS.

Extraction, Characterization and Immune Effect Analysis of Listeria Monocytogenes Membrane Vesicles

To establish a method for extracting Listeria monocytogenesmembrane vesicles (LM-MVs) and to analyze the characteristics of LM-MVs and their ability to induce innate immune effect in vitro so as to lay the foundation for research into using LM-MVs as vaccine carrier and drug delivery platform. The membrane vesicles secreted by Listeria monocytogenes were extracted through a continuous process, including culturing, centrifugation, filtration, ultrafiltration concentration and ultracentrifugation. The morphological characteristics of LM-MVs were observed with transmission electron microscope, and particle size distribution were measured by dynamic light scattering analysis. SDS-PAGE and Western blot were used to analyze the protein composition of LM-MVs. CCK-8 cell proliferation and toxicity determination experiments were done to analyze their effect on the proliferation of innate immune cells, and qPCR was used to analyze their ability to induce innate immune responses. A method for extracting LM-MVs was successfully established. Under the transmission electron microscope, LM-MVs presented a nearly circular film-like structure, and dynamic light scattering analysis showed that their sizes were between 65 and 190 nm. SDS-PAGE and Western blot showed that LM-MVs contained proteins, including listeriolysin O (LLO). CCK-8 cell proliferation and toxicity experiment showed that after intervention with 10, 20 and 50 μg/mL of LM-MVs for 24 hours, the proliferation rate of DC 2.4 mouse dendritic cell line was higher than that of non-interventional DC 2.4 cells ( P<0.05); after intervention with 0.1, 1, 10, 20 and 50 μg/mL of LM-MVs for 24 hours, the proliferation rate of RAW 264.7 cells was higher than that of non-interventional RAW 264.7 cells ( P<0.01). The results of qPCR showed that, after intervention with 50 μg/mL of LM-MVs, the expression levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6 and IL-10 in RAW 264.7 cells were higher than those of non-intervention control cells ( P<0.05). The method established in the study can be used to extract LM-MVs. The extracted LM-MVs have a diameter of 65-190 nm and a nearly circular membrane-like structure. They can secrete a variety of protein components and stimulate innate immune responses.

Numerical Analysis of Emulsion Separation Processes in a Continuous Centrifuge

Numerical Analysis of Emulsion Separation Processes in a Continuous Centrifuge