Acid/base responsive pseudo[3]rotaxanes from amine naphthotubes and bis-pyridinium/isoquinolinium guests.

A novel cooperative pseudo[3]rotaxane system was successfully constructed by the inclusion complexation of two identical amine naphthotubes with a bis-pyridinium/isoquinolinium guest. Single crystal structure analysis revealed that weak Csp3-H⋯O hydrogen bonds between the two hosts are responsible for the positive cooperativity during the formation of pseudo[3]rotaxanes. Moreover, intermolecular charge-transfer interactions between the electron-rich host and the electron-poor guests were observed. The pseudo[3]rotaxanes showed pH-controllable association/dissociation processes with naked-eye color changes in solution.

Amide naphthotube as a novel supramolecular sequestration agent for tetracaine and decamethonium.

RATIONALE: Anesthetics are widely used for optimizing surgical conditions, postoperative pain management, and treating various chronic pain conditions. Tetracaine and decamethonium are representative drugs of local anesthetics and neuromuscular blocking agents, respectively. However, overdose and toxicity of the drugs always lead to serious adverse events. Thus, there is a strong demand for effective antidotes. METHODS: The binding interactions of amide naphthotubes with tetracaine and decamethonium were systematically studied using 1H NMR, ITC, and DFT calculations. The antidotal effects of amide naphthotube to tetracaine toxicity were assessed in vitro and in vivo, and the mechanism of detoxification was explored at a cellular level. Additionally, mouse models were established to evaluate the reversal activities of amide naphthotube on decamethonium-induced mortality and muscle relaxation, and the reversal mechanism was investigated through pharmacokinetic experiments. RESULTS: We have demonstrated that the anti-isomer of amide naphthotube exhibits significant binding affinities towards tetracaine (K a = 1.89×107 M-1) and decamethonium (K a = 1.01×107 M-1) in water. The host displayed good biocompatibility both in vitro and in vivo. The administration of amide naphthotube following tetracaine overdose in mouse models notably increased the overall survival rate, indicating its effective antidotal properties. The host could reverse the tetracaine-induced Na+ channels blockage at the cellular level. Moreover, the injection of amide naphthotube also reversed the mortality and paralysis induced by decamethonium in mouse models following a pharmacokinetic mechanism. CONCLUSION: An emerging artificial receptor, amide naphthotube, has strong binding affinities towards tetracaine and decamethonium. It functions as a supramolecular antidote for tetracaine poisoning and a reversal agent for decamethonium by selectively sequestering these compounds in vivo.

Neuregulin-1 immunoreactivity in peripheral plasma is associated with rs6982890 polymorphism-mediated psychotic symptoms in schizophrenia.

OBJECTIVES: Neuregulin 1 (NRG1) is a risk gene for schizophrenia and involved in neurodevelopment and synaptic plasticity. Polymorphisms in NRG1 may affect psychotic symptoms in schizophrenia. This study investigated the effects of the single nucleotide polymorphism (SNP) rs6982890 on peripheral plasma NRG1 immunoreactivity, clinical symptoms and cognitive functions in schizophrenia patients. MATERIAL AND METHODS: We recruited subjects from the Han population of northern China from 2010 to 2022. We first genotyped and analyzed 6 NRG1 SNPS in 1304 patients with schizophrenia and 871 healthy controls. Then, 91 patients with schizophrenia and 40 healthy controls were selected to detect the peripheral plasma NRG1 immunoreactivity by ELISA. Among them, 84 patients were divided into rs6982890 genotypes to analyze the correlation between NRG1 immunoreactivity and clinical symptoms. RESULTS: Rs6982890 allelic frequencies were statistically significant between patients and controls. Baseline peripheral plasma NRG1 immunoreactivity in patients were significantly lower than controls. NRG1 immunoreactivity in patients were significantly increased after 8 weeks of antipsychotic treatment and significantly correlated with clinical symptoms and cognitive function. Genotyping of patients with SNP rs6982890 indicated NRG1 immunoreactivity in CC genotype increased significantly after treatment, while CT genotype had no significant change. Baseline NRG1 immunoreactivity with the CT genotype were significantly higher than CC genotype. CONCLUSIONS: NRG1 SNP rs6982890 is significantly associated with schizophrenia in the Han population of northern China, and it may affect the effect of antipsychotic drug treatment by regulating the peripheral plasma NRG1 immunoreactivity.

An endo-functionalized molecular cage for selective potentiometric determination of creatinine.

Potentiometric ion-selective electrodes (ISEs), which rely on selective and lipophilic ionophores, are commonly employed in clinical diagnostics. However, there are very limited specific ionophores for the detection of creatinine, a critical biomarker for renal function assessment. In the present research, we designed and synthesized an endo-functionalized cage, which is able to selectively bind the creatininium cation (K a = 8.6 × 105 M-1) through the formation of multiple C-H⋯O and N-H⋯N hydrogen bonds and cation⋯π interactions. ISEs prepared with this host show a Nernstian response to creatinine and exhibit excellent selectivity and a low detection limit of 0.95 µM. In addition, the creatinine levels in urine or plasma samples determined by our sensor are consistent with those analyzed using enzymatic assay on a Cobas c702. The method is simple, fast and accurate, and amenable to clinical detection of creatinine levels.

Elevated plasma matrix metalloproteinase 9 in schizophrenia patients associated with poor antipsychotic treatment response and white matter density deficits.

Oxidative stress and neuroinflammation contribute to schizophrenia (SCZ) pathology and may influence treatment efficacy. Matrix metalloproteinase 9 (MMP9) is a critical molecular node mediating the interaction between oxidative stress and inflammation, and so may influence treatment efficacy. Here we examined the associations of plasma MMP9 concentration with antipsychotic drug responses, clinical symptoms, and brain structure. A total of 129 healthy controls and 124 patients with SCZ were included in this study. Patients were monitored clinically during 8 weeks of antipsychotic treatment and classified as poor responders (n = 49) or good responders (n = 75). We then compared plasma MMP9 concentrations in healthy controls at baseline and both SCZ responder groups at baseline and after the 8-week antipsychotic treatment regimen. Cognitive function was also examined using the MATRICS Consensus Cognitive Battery. In addition, we extracted regional white matter density from magnetic resonance images of patients. Compared to healthy controls, plasma MMP9 levels were significantly elevated in poor responders at baseline and negatively correlated with both white matter density in the right superior temporal gyrus and the change in cognitive symptoms after treatment. Conversely, there was no significant difference in plasma MMP9 between good responders and healthy controls, and no associations of plasma MMP9 with cognitive symptoms or regional white matter density among good responders. Elevated plasma MMP9 is associated with poor antipsychotic drug efficacy and white matter deficits in SCZ patients, and so may be a useful biomarker to guide personalized treatment.

Flexible Hydrazone-Linked Metal-Covalent Organic Frameworks with Copper Clusters for Efficient Electrocatalytic Oxygen Evolution Reaction.

Despite the challenges associated with the synthesis of flexible metal-covalent organic frameworks (MCOFs), these offer the unique advantage of maximizing the atomic utilization efficiency. However, the construction of flexible MCOFs with flexible building units or linkages has rarely been reported. In this study, novel flexible MCOFs are constructed using flexible building blocks and copper clusters with hydrazone linkages. The heterometallic frameworks (Cu, Co) are prepared through the hydrazone linkage coordination method and evaluated as catalysts for the oxygen evolution reaction (OER). Owing to the spatial separation and functional cooperation of the heterometallic MCOF catalysts, the as-synthesized MCOFs exhibited outstanding catalytic activities with an overpotential of 268.8 mV at 10 mA cm-2 for the OER in 1 M KOH, which is superior to those of the reported covalent organic frameworks (COFs)-based OER catalysts. Theoretical calculations further elucidated the synergistic effect of heterometallic active sites within the linkages and frameworks, contributing to the enhanced OER activity. This study thus introduces a novel approach to the fundamental design of flexible MCOF catalysts for the OER, emphasizing their enhanced atomic utilization efficiency.

Evaluating extractable work of quantum batteries via entropic uncertainty relations.

In this study, we investigate the effectiveness of entropic uncertainty relations (EURs) in discerning the energy variation in quantum batteries (QBs) modelled by battery-charger field in the presence of bosonic and fermionic reservoirs. Our results suggest that the extractable works (exergy and ergotropy) have versatile characteristics in different scenarios, resulting in a complex relationship between tightness and extractable work. It is worth noting that the tightness of the lower bound of entropic uncertainty can be a good indicator for energy conversion efficiency in charging QBs. Furthermore, we disclose how the EUR including uncertainty and lower bound contributes to energy conversion efficiency in the QB system. It is believed that these findings will be beneficial for better understanding the role of quantum uncertainty in evaluating quantum battery performance.

Selective recognition and enrichment of C70 over C60 using an anthracene-based nanotube.

Selective recognition and enrichment of fullerenes (e.g., C60 and C70) remains challenging due to the same diameter and geometrical similarity. Herein, we report a hexagonal anthracene-based nanotube (1) through a one-pot Suzuki-Miyaura cross-coupling reaction. With anthracene-based side walls and pyridine linkers, 1 features a nano-scale tubular cavity measuring 1.2 nm in diameter and 0.9 nm in depth, along with pH-responsive properties. Interestingly, the electron-rich 1 shows high binding affinity (K a ≈ 106 M-1) and selectivity (K s ≈ 140) to C70 over C60 in toluene, resulting from their different contribution of π-π interactions with the host. The protonation of 1 simultaneously alters the electronic properties within the nanotube, resulting in the release of the fullerene guests. Lastly, the selective recognition and pH stimuli-responsive properties of the nanotube have been utilized to enrich C70 from its low-content mixtures of fullerenes in chloroform.

Drug-eluting bead transarterial chemoembolization as neoadjuvant therapy pre-liver transplantation for advanced-stage hepatocellular carcinoma.

BACKGROUND: The objectives of this study were to assess the safety and efficacy of drug-eluting bead transarterial chemoembolization (DEB-TACE) as neoadjuvant therapy before liver transplantation (LT) for advanced-stage hepatocellular carcinoma (HCC) and to analyze the prognostic factors. AIM: To determine whether DEB-TACE before LT is superior to LT for advanced-stage HCC. METHODS: A total of 99 individuals diagnosed with advanced HCC were studied retrospectively. The participants were categorized into the following two groups based on whether they had received DEB-TACE before LT: DEB-TACE group (n = 45) and control group (n = 54). The participants were further divided into two subgroups based on the presence or absence of segmental portal vein tumor thrombus (PVTT). The DEB-TACE group consisted of two subgroups: Group A (n = 31) without PVTT and group B (n = 14) with PVTT. The control group also had two subgroups: Group C (n = 37) without PVTT and group D (n = 17) with PVTT. Data on patient demographics, disease characteristics, therapy response, and adverse events (AEs) were collected. The overall survival (OS) and recurrence-free survival (RFS) rates were assessed using Kaplan-Meier curves. Univariate and multivariate Cox regression analyses were conducted to determine the parameters that were independently related to OS and RFS. RESULTS: The DEB-TACE group exhibited an overall response rate of 86.6%. Following therapy, there was a significant decrease in the median alpha-fetoprotein (AFP) level (275.1 ng/mL vs 41.7 ng/mL, P < 0.001). The main AE was post-embolization syndrome. The 2-year rates of RFS and OS were significantly higher in the DEB-TACE group than in the control group (68.9% vs 38.9%, P = 0.003; 86.7% vs 63.0%, P = 0.008). Within the subgroups, group A had higher 2-year rates of RFS and OS compared to group C (71.0% vs 45.9%, P = 0.038; 83.8% vs 62.2%, P = 0.047). The 2-year RFS rate of group B was markedly superior to that of group D (64.3% vs 23.5%, P = 0.002). Results from multivariate analyses showed that pre-LT DEB-TACE [hazard ratio (HR) = 2.73, 95% confidence interval (CI): 1.44-5.14, P = 0.04], overall target tumor diameter ≤ 7 cm (HR = 1.98, 95%CI: 1.05-3.75, P = 0.035), and AFP level ≤ 400 ng/mL (HR = 2.34; 95%CI: 1.30-4.19, P = 0.009) were significant risk factors for RFS. Additionally, pre-LT DEB-TACE (HR = 3.15, 95%CI: 1.43-6.96, P = 0.004) was identified as a significant risk factor for OS. CONCLUSION: DEB-TACE is a safe and efficient therapy for advanced-stage HCC and also enhances patient survival after LT.

Development of α-Tocopherol Loaded PLGA Nanoparticles and Its Evaluation as a Novel Immune Adjuvant.

With the continuous development of preventive and therapeutic vaccines, traditional adjuvants cannot provide sufficient immune efficacy and it is of high necessity to develop safe and effective novel nanoparticle-based vaccine adjuvants. α-Tocopherol (TOC) is commonly used in oil-emulsion adjuvant systems as an immune enhancer, yet its bioavailability is limited by poor water solubility. This study aims to develop TOC-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (TOC-PLGA NPs) to explore the potential of TOC-PLGA NPs as a novel nanoparticle-immune adjuvant. TOC-PLGA NPs are prepared by a nanoprecipitation method and their physicochemical properties are characterized. It is shown that TOC-PLGA NPs are 110.8 nm, polydispersity index value of 0.042, and Zeta potential of -13.26 mV. The encapsulation efficiency and drug loading of NPs are 82.57% and 11.80%, respectively, and the cumulative release after 35 days of in vitro testing reaches 47%. Furthermore, TOC-PLGA NPs demonstrate a superior promotion effect on RAW 264.7 cell proliferation compared to PLGA NPs, being well phagocytosed and also promoting antigen uptake by macrophages. TOC-PLGA NPs can strongly upregulate the expression of co-stimulatory surface molecules and the secretion of cytokines. In conclusion, TOC-PLGA NPs can be a novel vaccine adjuvant with excellent biocompatibility and significant immune-enhancing activity.

Alkali-Induced Protein Structural, Foaming, and Air-Water Interfacial Property Changes and Quantitative Proteomic Analysis of Buckwheat Sourdough Liquor.

In this study, the protein structural, foaming, and air-water interfacial properties in dough liquor (DL) ultracentrifugated from buckwheat sourdough with different concentrations of an alkali (1.0-2.5% of sodium bicarbonate) were investigated. Results showed that the alkali led to the cross-linking of protein disulfide bonds through the oxidation of free sulfhydryl groups in DL. The alterations in protein secondary and tertiary structures revealed that the alkali caused the proteins in DL to fold, decreased the hydrophobicity, and led to a less flexible but compact structure. The alkali accelerated the diffusion of proteins and decreased the surface tension of DL. In addition, the alkali notably improved the foam stability by up to 34.08% at 2.5% concentration, mainly by increasing the net charge, reducing the bubble size, and strengthening the viscoelasticity of interfacial protein films. Quantitative proteomic analysis showed that histones and puroindolines of wheat and 13S globulin of buckwheat were closely related to the changes in the alkali-induced foaming properties. This study sheds light on the mechanism of alkali-induced improvement in gas cell stabilization and the buckwheat sourdough steamed bread quality from the aspect of the liquid lamella.

Exploiting the advantages of cationic copolymers and AgBr nanoparticles to optimize the antibacterial activity of chitosan.

Recently, the chitosan (CS)-based composites have attracted increasing attention for controlling and preventing the spread of pathogenic microorganisms. Herein, an amphiphilic copolymer containing epoxy and quaternary ammonium groups (PBGDBr) was synthesized via three common acrylate monomers. The epoxy groups of this copolymer were then crosslinked with the amino groups of CS to synthesize a natural/synthetic (PBGDBr-C) composite to increase the water solubility of CS under alkaline conditions and enhance its antibacterial activity based on chemical contact-type modes. Moreover, silver bromide nanoparticles (AgBr NPs)-decorated PBGDBr-C (AgBr@PBGDBr-C) composite was prepared, which aimed to endow the final AgBr@PBGDBr-C composite with a photodynamic antibacterial mode relying on the formation of Ag/AgBr nanostructures catalyzed by visible light on AgBr NPs. The results showed that the final composite possessed satisfactory bactericidal effects at concentrations higher than 64 and 128 µg/mL against Escherichia coli and Staphylococcus aureus, respectively. Additionally, The L929 cells treated with the final composite retained high cell viability (>80 %) at a concentration of 128 µg/mL, indicating its low toxicity to L929 cells. Overall, our synthetic strategy exploits a multi-modal system that enables chemical-photodynamic synergies to treat infections caused by pathogenic bacteria while delaying the development of bacterial resistance.

Deciphering the folate puzzle: Unraveling the impact of genetic variations and metabolites on cancer risk.

The C677T polymorphism in the MTHFR gene and its role in folate metabolism, impacting serum folate metabolites like THF and 5-MTHF, is a critical but underexplored area in cancer research. This nested case-control study utilized data from CHHRS, involving 87,492 hypertensive adults without prior cancer. During a median of 2.02 years, we identified 1332 cancer cases and matched controls based on age, sex, and residency. Serum levels of folate, THF, and 5-MTHF were measured, and the MTHFR C677T gene polymorphism was considered. Statistical analyses included restricted cubic spline regression and conditional logistic regression models. Serum THF levels were inversely associated with overall cancer risk (ORper SD = 0.90, 95% CI = 0.82-0.99), while 5-MTHF levels showed a negative association in the general cohort (ORQ3 vs. Q1 = 0.76, 95% CI = 0.60-0.96; ORQ4 vs. Q1 = 0.75, 95% CI = 0.58-0.98) and in individuals with MTHFR C677T (CC + CT) polymorphism (ORper SD = 0.87, 95% CI = 0.77-0.99; ORQ4 VS. Q1 = 0.79, 95% CI = 0.61-0.98), but a positive association in the MTHFR C677T (TT) subgroup (ORper SD = 1.89, 95% CI = 1.02-3.72; ORQ4 VS. Q1 = 2.17, 95% CI = 1.06-8.21). The impact of folate, THF, and 5-MTHF on cancer risk varied significantly across different cancer types and MTHFR C677T genotypes. This study provides novel insights into the variable effects of folate and its metabolites on cancer risk, influenced by genetic factors like the MTHFR C677T polymorphism and cancer type.

Early-onset and later-onset cancer: trends, risk factors, and prevention in Northern China.

The characteristics of early-onset (onset age <50 years) and later-onset (onset age ≽ 50 years) cancers differ significantly. Identifying novel risk factors for both types of cancer is crucial for increasing awareness of cancer prevention and for reducing its burden. This study aimed to analyze the trends in incidence and risk factors for early-onset and late-onset cancers. We conducted a prospective study by drawing data from the Kailuan Study. This study included 6,741 participants with cancer (624 with early-onset cancer and 6,117 with later-onset cancer) and 6,780 matched controls among the 186,249 participants who underwent Kailuan health examinations from 2006 to 2019. The primary outcomes were cancer incidence rates, and associated risk factors for early- and later-onset cancer. Weighted Cox regression was used to calculate hazard ratios and 95% confidence intervals of each exposure factor for early- and later-onset cancer by cancer type. Population-attributable risk proportions were used to estimate the number of cases that could be prevented by eliminating a risk factor from the population. Except for liver cancer, incidence rates for nearly all types of cancer increased during the study period. Smoking, alcohol consumption, lipid metabolism disorders, hypertension, diabetes mellitus, fatty liver, and inflammation were associated with a significantly increased risk of cancer at multiple sites, but risk factors for cancer incidence differed by site. Smoking, alcohol consumption, inflammation, and hypertension were the major contributors to preventable cancer. The incidence of several different types of cancer, including early-onset cancer, is increasing in northeastern China. Differences in risk factors between early-onset and later-onset malignancies may contribute to the divergence in the observed changes in incidence trends between these two specific types of cancer.

The causal role of multiple psycho-emotional disorders in gastroesophageal reflux disease: A two-sample Mendelian randomized study.

BACKGROUND: Observational studies have previously shown a potential link between psycho-emotional disorders, such as mood swings, highly strung, anxious feelings, and gastroesophageal reflux disease (GERD). However, the credibility of these associations could be influenced by various confounding factors. Consequently, our study sought to employ a Mendelian randomization (MR) approach to elucidate a potential causal relationship between psycho-emotional disorders and GERD. METHOD: Information on independent genetic variants linked to mood swings, highly strung, and anxious feelings was gathered from European populations participating in the IEU Open GWAS research. The FinnGen Consortium provided the genome-wide association study (GWAS) summary statistics for GERD. Our analysis employed the inverse variance weighted (IVW) method under the random effects model as the main analytical method. To further bolster our findings, we employed the weighted median and MR Egger methods. In addition, we conducted a series of sensitivity analyses. RESULTS: Our study supports the existence of a causal relationship between psycho-emotional disorders and GERD. Mood swings, highly strung, and anxious feelings adversely affected GERD risk (mood swings: OR 2.21, 95% CI 1.19-5.59, p = 3.09 × 10-2; highly strung: OR 5.63, 95% CI 1.77-17.94, p = 3.42 × 10-3; anxious feelings: OR 2.48, 95% CI 1.08-4.33, p = 2.89 × 10-2). CONCLUSION: This Mendelian randomization study provides robust support for the notion that mood swings, highly strung and anxious feelings, are associated with an increased risk of developing GERD.

The relationship between different fatty acids intake and the depressive symptoms: A population-based study.

BACKGROUND: Depression is a common psychological disorder worldwide, affecting mental and physical health. Previous studies have explored the benefits of polyunsaturated fatty acids (PUFAs) intake in depressive symptoms; however, few studies have focused on the association between all types of fatty acids intake and depressive symptoms. Therefore, we explored the relationship between the intake of different fatty acids intake and the risk of depressive symptoms. METHODS: The study was based on the data from the 2005-2018 National Health and Nutrition Examination Survey (NHANES), a large US-based database. We used a nutrient residual model and multi-nutrient density model for the analysis. We calculated the nutrient density and residual in men and women separately, and the fatty acids intake was divided into quartiles based on the sex distribution. The relationship between the depressive symptoms and the intake of different fatty acids was examined using logistic regression; furthermore, we explored the relationships separately in men and women. RESULTS: The intake of monounsaturated fatty acids (MUFAs) and PUFAs, particularly n-3 and n-6 PUFAs, were associated with reduced odds ratios for depressive symptoms. The inverse relationship between the intake of MUFAs, PUFAs, n-3, and n-6 PUFAs and depressive symptoms was stronger in women. The inverse relationship between total fatty acid (TFAs) intake and depressive symptoms existed only in a single model. In contrast, saturated fatty acid (SFAs) intake was not related to depressive symptoms. CONCLUSION: Consuming MUFAs and PUFAs can counteract the depressive symptoms, especially in women.

Ether Naphthotube Host-Guest Complexes and [2]Rotaxanes with Dications.

The interactions between ether naphthotube and a series of dication guests in organic solution were investigated. It was found that ether naphthotube formed stable host-guest complexes selectively with these guests in a 1 : 1 stoichiometric ratio with association constants ranging from 102 to 106 M-1, which were confirmed by 1H-NMR spectra and ITC experiments. The host-guest interactions are driven by enthalpy change as the entropic factors are unfavorable. Positive correlations between ΔH and ΔS have been observed in the host-guest complexes. Furthermore, the para-substitution of the guests can significantly affect the binding affinities through a combination of field/inductive and resonance effects by following a linear free energy relationship. Based on the host-guest complexes composed of ether naphthotube and organic cations, two interlocked [2]rotaxanes were prepared by cationization reaction and Huisgen cycloaddition between the cations and the stopper components. The ether naphthotube-based host-guest complexes are useful for creating sophisticated interlocked molecules.

Human umbilical cord blood mesenchymal stem cells mediate microglia activation and improve anxiety-like behavior in MIA-induced offspring of schizophrenic rats.

Current treatments for schizophrenia (SCZ) remain largely ineffective in one-third of patients. Recent studies using stem cell therapy show a close relationship between stem cell immunomodulatory function and neuroinflammation in SCZ. To better investigate the efficacy of stem cell therapy for SCZ, human umbilical cord blood mesenchymal stem cells (hUC-MSC) with powerful immunomodulatory effects were administered to rats via the tail vein (once a week for 5 consecutive weeks starting from the weaning period) using a maternal immune activation (MIA) rodent model. Open field, PPI, Western blotting, Q-PCR, and immunofluorescence were used to assess the biological effects of repeated tail vein injections of hUC-MSC in offspring rats following the MIA model of SCZ. The results indicated that offspring of MIA rats exhibited schizophrenia-like (SCZ-like) anxiety behavior, with observed microglial activation triggering neuroinflammation. Furthermore, levels of IBA1, HMGB1, and PSD95 were significantly up-regulated, while SYP was significantly down-regulated. It is suggested that hUCB-MSCs may act through HMGB1, Iba1, PSD95, and related pathway molecules to alleviate neuroinflammation and repair synaptic damage by regulating the activity state of microglia. Consequently, this could improve the abnormal behavior observed in MIA offspring rats.

Optimization of seeding density of OP9 cells to improve hematopoietic differentiation efficiency.

BACKGROUND: OP9 mouse stromal cell line has been widely used to induce differentiation of human embryonic stem cells (hESCs) into hematopoietic stem/progenitor cells (HSPCs). However, the whole co-culture procedure usually needs 14-18 days, including preparing OP9 cells at least 4 days. Therefore, the inefficient differentiation system is not appreciated. We aimed to optimize the culture conditions to improve differentiation efficiency. METHODS: In the experimental group, we set six different densities of OP9 cells and just cultured them for 24 h before co-culture, and in the control group, OP9 cells were cultured for 4 days to reach an overgrown state before co-culture. Then we compared the hematopoietic differentiation efficiency among them. RESULTS: OP9 cells were randomly assigned into two groups. In the experimental group, six different plated numbers of OP9 cells were cultured for 1 day before co-culture with hESCs. In contrast, in the control group, OP9 cells were cultured for 4 days at a total number of 3.1 × 104 cells/cm2 in a 6-well plate to reach an overgrown state before co-culture. Hematopoietic differentiation was evaluated with CD34 immunostaining, and compared between these two groups. We could not influence the differentiation efficiency of OP9 cells with a total number of 10.4 × 104 cells/cm2 in a 6-well plate which was cultured just for 1 day, followed by co-culture with hESCs. It reached the same differentiation efficiency 5 days earlier than the control group. CONCLUSION: The peak of CD34 + cells appeared 2 days earlier compared to the control group. A total number of 1.0 × 106 cells in a 6-well plate for OP9 cells was appropriate to have high differentiation efficiency.