Formulation and Characterization of Triamcinolone Acetonide Acetate-Loaded Microspheres Prepared by a Static Mixing Technique.

PURPOSE: Reproducibility and scale-up production of microspheres through spray drying present significant challenges. In this study, biodegradable microspheres of Triamcinolone Acetonide Acetate (TAA) were prepared using a novel static mixing method by employing poly( lactic-co-glycolic acid) (PLGA) as the sustained-release carrier. METHODS: TAA-loaded microspheres (TAA-MSs) were prepared using a static mixing technique. The PLGA concentration, polyvinyl alcohol concentration (PVA), phase ratio of oil/water, and phase ratio of water/solidification were optimized in terms of the particle size, drug loading (DL), and encapsulation efficiency (EE) of TAA-MSs. The morphology of TAA-MSs was examined using Scanning Electron Microscopy (SEM), while the physicochemical properties were evaluated through X-ray diffraction (XRD), Differential Scanning Calorimetry (DSC), and Fourier Transform Infrared Spectroscopy (FT-IR). The in vitro release of TAA-MSs was compared to that of the pure drug (TAA) using a water-bath vibration method in the medium of pH 7.4 at 37°C. RESULTS: The formulation composition and preparation condition for the preparation of TAA-MSs were optimized as follows: the PLGA concentration was 1%, the phase ratio of oil(dichloromethane) /water (PVA solution) was 1:3, the phase ratio of water (PVA solution)/solidification was 1:2. The optimized TAA-MSs displayed spherical particles with a size range of 30-70 µm, and DL and EE values of 27.09% and 98.67%, respectively. Moreover, the drug-loaded microspheres exhibited a significant, sustained release, with 20% of the drug released over a period of 28 days. The XRD result indicated that the crystalline form of TAA in microspheres had been partly converted into the amorphous form. DSC and FT-IR results revealed that some interactions between TAA and PLGA occurred, indicating that the drug was effectively encapsulated into PLGA microspheres. CONCLUSION: TAA-loaded PLGA microspheres have been successfully prepared via the static mixing technique with enhanced EE and sustained-release manner.

Crafting Docetaxel-Loaded Albumin Nanoparticles Through a Novel Thermal-Driven Self-Assembly/Microfluidic Combination Technology: Formulation, Process Optimization, Stability, and Bioavailability.

Background: The commercial docetaxel (DTX) formulation causes severe side effects due to polysorbate 80 and ethanol. Novel surfactant-free nanoparticle (NP) systems are needed to improve bioavailability and reduce side effects. However, controlling the particle size and stability of NPs and improving the batch-to-batch variation are the major challenges. Methods: DTX-loaded bovine serum albumin nanoparticles (DTX-BSA-NPs) were prepared by a novel thermal-driven self-assembly/microfluidic technology. Single-factor analysis and orthogonal test were conducted to obtain the optimal formulation of DTX-BSA-NPs in terms of particle size, encapsulation efficiency (EE), and drug loading (DL). The effects of oil/water flow rate and pump pressure on the particle size, EE, and DL were investigated to optimize the preparation process of DTX-BSA-NPs. The drug release, physicochemical properties, stability, and pharmacokinetics of NPs were evaluated. Results: The optimized DTX-BSA-NPs were uniform, with a particle size of 118.30 nm, EE of 89.04%, and DL of 8.27%. They showed a sustained release of 70% over 96 hours and an increased stability. There were some interactions between the drug and excipients in DTX-BSA-NPs. The half-life, mean residence time, and area under the curve (AUC) of DTX-BSA-NPs increased, but plasma clearance decreased when compared with DTX. Conclusion: The thermal-driven self-assembly/microfluidic combination method effectively produces BSA-based NPs that improve the bioavailability and stability of DTX, offering a promising alternative to traditional formulations.

Insight into the chemical compositions of Anhua dark teas derived from identical tea materials: A multi-omics, electronic sensory, and microbial sequencing analysis.

Anhua dark teas (DTs), including Tianjian tea, Qianliang tea, Hei brick tea, and Fu brick tea, are unique fermented teas from China's Anhua County; yet their chemical composition differences remain unclear. Herein, metabolomics, volatolomics, and electronic sensory assessments were employed to analyze and compare chemical compositions and sensory characteristics of five types of Anhua DTs. All of these teas were derived from identical tea materials. Chemical compositions differed significantly among Anhua DTs, with Tianjian tea remarkable. Long-lasting fermentation and complex processing methods led to transformation of multiple compounds, particularly catechins. Eighteen volatile compounds with OVA > 1 were key aroma contributors in Anhua DTs. Internal transcribed spacer and 16S ribosomal DNA sequencing showed that Eurotium, Pseudomonas, and Bacillus are dominant microorganisms in Anhua DTs. Furthermore, this study unveiled notable differences in chemical compositions between Anhua DTs and five other traditional types of tea. This research enhances our understanding of Anhua DTs processing.

Development of Polyvinyl Alcohol/Polyethylene Glycol Copolymer-based Orodispersible Films Loaded with Entecavir: Formulation and In vitro Characterization.

PURPOSE: The aim of the study is to prepare entecavir (ETV)-loaded orodispersible films (ODFs) using polyvinyl alcohol (PVA)/polyethylene glycol (PEG) graft copolymer (Kollicoat® IR) as a film-forming agent, and further to evaluate the dissolution rate, mechanical and physicochemical properties of films. METHODS: ETV-ODFs were prepared by a solvent casting method. The amount of film-forming agent, plasticizer, and disintegrating agent was optimized in terms of the appearance, thickness, disintegration time and mechanical properties of ODFs. The compatibility between the drug and each excipient was conducted under high temperature (60 °C), high humidity (RH 92.5%), and strong light (4500 Lx) for 10 days. The dissolution study of optimal ODFs compared with the original commercial tablet (Baraclude®) was performed using a paddle method in pH 1.0, pH 4.5, pH 6.8, and pH 7.4 media at 37 °C. The morphology of ODFs was observed via scanning electron microscopy (SEM). The mechanical properties such as tensile strength (TS), elastic modulus (EM), and percentage elongation (E%) of ODFs were evaluated using the universal testing machine. The physicochemical properties of ODFs investigated using X-ray diffraction (XRD), differential scanning calorimetry (DSC), and Fourier transform infrared spectroscopy (FT-IR). RESULTS: The related substances were less than 0.5% under high temperature, high humidity, and strong light for 10 days when ETV was mixed with excipients. The optimal formulation of ODFs was set as the quality ratio of Kollicoat® IR, glycerol, sodium alginate (ALG-Na): TiO2: MCC+CMC-Na: ETV was 60:9:12:1:1:1. The drug-loaded ODFs were white and translucent with excellent stripping property. The thickness, disintegration time, EM, TS, and E% were 103.33±7.02 µm, 25.31±1.95 s, 25.34±8.69 Mpa, 2.14±0.26 Mpa, and 65.45±19.41 %, respectively. The cumulative drug release from ODFs was more than 90% in four different media at 10 min. The SEM showed that the drug was highly dispersible in ODFs, and the XRD, DSC, and FT-IR results showed that there occurred some interactions between the drug and excipients. CONCLUSION: In conclusion, the developed ETV-loaded ODFs showed relatively short disintegration time, rapid drug dissolution, and excellent mechanical properties. This might be an alternative to conventional ETV Tablets for the treatment of chronic hepatitis B.

The R2R3-MYB transcription factor CsMYB42 regulates theanine biosynthesis in albino tea leaves.

Theanine is a unique secondary metabolite in tea plants and contributes to the umami taste and health benefits of tea. However, theanine biosynthesis in tea plants is not fully understood, and its mechanism of transcriptional regulation remains poorly reported. Theanine content was significantly correlated with the expression of theanine biosynthesis-related gene CsGS1c and transcription factor CsMYB42 in different leaf positions and picking times, but there was no significant correlation in different tissues of albino tea plant 'Anjibaicha'. This suggests that CsMYB42 may regulate CsGS1c to synthesize theanine in albino tea leaves, and the regulation is tissue specific. CsMYB42 is a nuclear-localized R2R3-MYB transcription factor gene with transcriptional activation activity. Yeast one-hybrid assay and electrophoretic mobility shift assay confirmed the direct binding of CsMYB42 to the promoter of CsGS1c. Luciferase assay showed that CsMYB42 activates the CsGS1c expression. Furthermore, the inhibition of CsMYB42 using an antisense oligonucleotide in tea leaves decreased CsGS1c expression and theanine content. These results indicate that CsMYB42 plays a crucial role in activating the expression of CsGS1c and may be involved in the biosynthesis of theanine in albino tea leaves. This study provides fresh insights into the tissue-specific regulation of theanine biosynthesis, which laid a foundation for breeding high-theanine tea plants.

Hypomethylation of Thyroid Peroxidase as a Biomarker for Hepatocellular Carcinoma with Tumor Thrombosis.

OBJECTIVE: Thyroid hormones (THs) regulate multiple physiological activities in the liver, including cellular metabolism, differentiation, and cell growth, and play important roles in the pathogenesis of hepatocellular carcinoma (HCC). Thyroid peroxidase (TPO) is a key molecule involved in the THs synthesis and signaling pathway. As an epigenetic modification, DNA methylation has a critical role in tumorigenesis with diagnostic potential. However, the connection between THs and DNA methylation has been rarely investigated. METHODS: The methylation of key TH-related genes was analyzed by in-house epigenome-wide scanning, and we further analyzed the methylation levels of the TPO promotor in 164 sample pairs of HCC and adjacent non-cancerous tissues by Sequenom EpiTYPER assays, and evaluated their clinical implications. RESULTS: We identified that the methylation of the TPO promoter was downregulated in the HCC tissues (P<0.0001) with a mean difference ranging from 18.5% to 22.3%. This methylation pattern correlated with several clinical factors, including a multi-satellite tumor, fibrous capsule, and the presence of tumor thrombus. The receiver operator characteristic (ROC) curve analysis further confirmed that the percent methylated reference (PMR) values for TPO were predictive of the tumor [the area under the curve (AUC) ranged from 0.755 to 0.818] and the thrombosis in the HCC patients (the AUC ranged from 0.706 to 0.777). CONCLUSION: These findings demonstrated that epigenetic alterations of TPO, as indicated by the PMR values, were a potential biomarker for HCC patients with tumor thrombosis.

Protective Effect of Eurotium cristatum Fermented Loose Dark Tea and Eurotium cristatum Particle on MAPK and PXR/AhR Signaling Pathways Induced by Electronic Cigarette Exposure in Mice.

Electronic-cigarette smoke (eCS) has been shown to cause a degree of oxidative stress and inflammatory damage in lung tissue. The aim of this study was to evaluate the repair mechanism of Eurotium cristatum fermented loose dark tea (ECT) and Eurotium cristatum particle metabolites (ECP) sifted from ECT after eCS-induced injury in mice. Sixty C57BL/6 mice were randomly divided into a blank control group, an eCS model group, an eCS + 600 mg/kg ECP treatment group, an eCS + 600 mg/kg ECT treatment group, an eCS + 600 mg/kg ECP prevention group, and an eCS + 600 mg/kg ECT prevention group. The results show that ECP and ECT significantly reduced the eCS-induced oxidative stress and inflammation and improved histopathological changes in the lungs in mice with eCS-induced liver injury. Western blot analysis further revealed that ECP and ECT significantly inhibited the eCS-induced upregulation of the phosphorylation levels of the extracellular Regulated protein Kinases (ERK), c-Jun N-terminal kinase (JNK) and p38mitogen-activated protein kinases (p38MAPK) proteins, and significantly increased the eCS-induced downregulation of the expression levels of the pregnane X receptor (PXR) and aryl hydrocarbon receptor (AhR) proteins. Conclusively, these findings show that ECP and ECT have a significant repairing effect on the damage caused by eCS exposure through the MAPK and PXR/AhR signaling pathways; ECT has a better effect on preventing eCS-induced injury and is suitable as a daily healthcare drink; ECP has a better therapeutic effect after eCS-induced injury, and might be a potential therapeutic candidate for the treatment of eCS-induced injury.

Integrative analysis of transcriptome and metabolome reveals the mechanism of foliar application of Bacillus amyloliquefaciens to improve summer tea quality (Camellia sinensis).

Bacillus amyloliquefaciens is a promising microbial agent for quality improvement in crops; however, the effects of B. amyloliquefaciens biofertilizers on tea leaf metabolites are relatively unknown. Herein, a combination of metabolome profiling and transcriptome analysis was employed to investigate the effects of foliar spraying with B. amyloliquefaciens biofertilizers on tea leaf quality. The tea polyphenol to amino acid ratio (TP/AA), catechin, and caffeine levels decreased, but theanine level increased in tea leaves after foliar spraying with B. amyloliquefaciens. The differentially accumulated metabolites included flavonoids, phenolic acids, organic acids, amino acids, and carbohydrates. The decrease in catechin was correlated with the catechin/flavonoid biosynthesis pathway. The AMPD gene was highly associated with caffeine content, while the GOGAT gene was associated with theanine accumulation. Foliar spraying with B. amyloliquefaciens biofertilizers may improve summer tea quality. Our findings provide a basis for the application of B. amyloliquefaciens biofertilizers in tea plants and new insights on summer tea leaf resource utilization.

Impact of information and Lévy noise on stochastic COVID-19 epidemic model under real statistical data.

In this paper, we consider the dynamical behaviour of a stochastic coronavirus (COVID-19) susceptible-infected-removed epidemic model with the inclusion of the influence of information intervention and Lévy noise. The existence and uniqueness of the model positive solution are proved. Then, we establish a stochastic threshold as a sufficient condition for the extinction and persistence in mean of the disease. Based on the available COVID-19 data, the parameters of the model were estimated and we fit the model with real statistics. Finally, numerical simulations are presented to support our theoretical results.

Identification of a WRKY transcriptional activator from Camellia sinensis that regulates methylated EGCG biosynthesis.

Naturally occurring methylated catechins, especially methylated EGCG in tea leaves are known to have many health benefits. Although the genes involved in methylated EGCG biosynthesis have been studied extensively, the transcriptional factors controlling methylated EGCG biosynthesis are still poorly understood. In the present study, a WRKY domain-containing protein termed CsWRKY57like was identified, which belongs to group IIc of the WRKY family, and contains one conserved WRKY motif. CsWRKY57like was found to localize in the nucleus, function as a transcriptional activator, and its expression positively correlated with methylated EGCG level. In addition, CsWRKY57like activated the transcription of three genes related to methylated EGCG biosynthesis, including CCoAOMT, CsLAR, and CsDFR by specifically interacting with their promoters via binding to the cis-acting element (C/T)TGAC(T/C). Further assays revealed that CsWRKY57like physically interacts with CsVQ4, and participates in the metabolic regulation of O-methylated catechin biosynthesis. Collectively, we conclude that CsWRKY57like may positively impact the biosynthesis of methylated EGCG in the tea plant, which comprehensively enriches the regulatory network of WRKY TFs associated with methylated EGCG and provide a potential strategy for the breeding of specific tea plant cultivars with high methylated EGCG .

The R2R3 Transcription Factor CsMYB59 Regulates Polyphenol Oxidase Gene CsPPO1 in Tea Plants (Camellia sinensis).

Polyphenol oxidase (PPO) plays a role in stress response, secondary metabolism, and other physiological processes during plant growth and development, and is also a critical enzyme in black tea production. However, the regulatory mechanisms of PPO genes and their activity in tea plants are still unclear. In this study, we measured PPO activity in two different tea cultivars, Taoyuandaye (TYDY) and Bixiangzao (BXZ), which are commonly used to produce black tea and green tea, respectively. The expression pattern of CsPPO1 was assessed and validated via transcriptomics and quantitative polymerase chain reaction in both tea varieties. In addition, we isolated and identified an R2R3-MYB transcription factor CsMYB59 that may regulate CsPPO1 expression. CsMYB59 was found to be a nuclear protein, and its expression in tea leaves was positively correlated with CsPPO1 expression and PPO activity. Transcriptional activity analysis showed that CsMYB59 was a transcriptional activator, and the dual-luciferase assay indicated that CsMYB59 could activate the expression of CsPPO1 in tobacco leaves. In summary, our study demonstrates that CsMYB59 represents a transcriptional activator in tea plants and may mediate the regulation of PPO activity by activating CsPPO1 expression. These findings provide novel insights into the regulatory mechanism of PPO gene in Camellia sinensis, which might help to breed tea cultivars with high PPO activity.

Eurotium cristatum Fermented Loose Dark Tea Ameliorates Cigarette Smoke-Induced Lung Injury by MAPK Pathway and Enhances Hepatic Metabolic Detoxification by PXR/AhR Pathway in Mice.

Cigarette smoke- (CS-) induced oxidative stress and inflammation in the lung are serious health problems. Primary and reprocessed tea products contain multiple antioxidants that have been reported to protect the lung against CS-induced injury. However, the beneficial effects of Eurotium cristatum fermented loose dark tea (ECT) and Eurotium cristatum particle metabolites (ECP) on CS-induced lung injury and its potential hepatic metabolic detoxification are still unclear. Therefore, sixty mice were randomly divided into six equal groups. CS-exposed mice were prevented or treated with ECP or ECT infusions for 12 or 8 weeks to determine the antioxidative stress, anti-inflammatory and potential metabolic detoxification of ECT and ECP. Thirty-six mice were randomly divided into six equal groups to observe the effects on hepatic metabolic detoxification by replacing daily drinking water with ECT. Results showed that CS significantly decreased the activities of glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) and upregulated the expressions of malondialdehyde (MDA), tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), IL-8, and IL-1ß in serum. These adverse effects were modulated by ECP and ECT. In addition, ECT upregulated the mRNA expression of pregnane X receptor (PXR) and cytochrome P450 (CYP450) in the liver on daily free drinking ECT mice group. Western blot analysis further revealed that in CS-exposed mice, ECP and ECT significantly decreased the phosphorylation of mitogen-activated protein kinase (MAPK) in the lung but upregulated the protein expressions of PXR and aryl hydrocarbon receptor (AhR) in the liver. Overall, our findings demonstrated that ECT and ECP protected against lung injury induced by CS via MAPK pathway and enhanced hepatic metabolic detoxification via PXR and AhR pathways. Therefore, daily intake of ECT and ECP can potentially protect against CS-induced oxidative and inflammatory injuries.

A novel microsensor for measuring thermal conductivity of fluid based on three omega method.

A novel microsensor has been designed for the measurement of thermal conductivity of fluids based on the three omega (3ω) method. First, we theoretically analyzed the heat conduction using the 3ω method to demonstrate the mechanism of the microsensor to measure the thermal conductivity of a fluid. For the main structure of the microsensor, a heater was supported by the thin dielectric layers. In order to obtain the optimal parameters, we used the finite element method to simulate the working condition of the microsensor. In the simulation model, the effects of the thicknesses of the heater and dielectric layers on the thermal conductivity λ of the fluid were analyzed. The simulation results confirmed the validity and accuracy of conventional analytical calculations. Based on the simulation and theoretical calculation, a microsensor was optimally designed and fabricated to measure the thermal conductivity of fluids. Experimental data are consistent with those reported in the literature and demonstrate that the proposed sensor is effective for measuring thermal conductivity of fluids, including conductive ones.

[Biomechanical stability study on different internal fixation for acetabular fracture].

OBJECTIVE: To investigate the biomechanical stability of the acetabular fracture with three different internal fixation methods. METHODS: Sixteen both-column acetabular fracture models were randomly divided into three groups:The specimens of 16 hip joints were randomly divided into 4 groups. Among them, 1 group of complete acetabulum were used as normal control group, and the other 3 groups simulated two column fracture models and were fixed in the following methods, respectively: anterior wall with screw and posterior with plate(SP), anterior wall with plate and posterior wall with screw(PS) and both wall with plate (PP). The degree of fracture displacement and the contact characteristics of the acetabulum were recorded by continuous vertical loading. RESULTS: The mean longitudinal displacement under the load 800 N of SP, PS and PP three groups were (1.92±0.81), (2.09±1.13) and (3.44±0.75) mm, there was significant difference between SP and PP group (P=0.033). And the mean horizontal displacement of SP, PS and PP three groups were(0.63±0.33), (0.77±0.45) and (1.44±0.56) mm, there was significant difference between SP and PP group(P=0.047).Compared with normal control group in the acetabular area under the loading 800 N, the contact area of SP, PS and PP groups were increased by 6%, 9% and 27%, there was significant difference between PP and normal control group (P=0.027). Meanwhile, the mean stress of SP, PS and PP groups were increased by 4%, 29% and 39%, there was significant difference between PP and intact acetabulum group (P=0.003). CONCLUSIONS: Anterior column screw combined with posterior column plate has better biomechanical stability and contact characteristics than other two methods.

[Delayed union or nonunion of the ulna after intramedullary nailing for pediatric forearm fractures].

OBJECTIVE: To analyze the causes of delayed union or nonunion of the ulna after intramedullary nailing in pediatric forearm fractures. METHODS: From February 2005 to February 2010,5 patients with forearm fractures who were treated with titanium elastic nailing (TEN) were identified to fulfill the criteria of having developed a delayed union or nonunion of the ulna. The causes of delayed union or nonunion were investigated according to mechanism of injury, fracture location, treatments methods and postoperative management. All patients were male and the age was 3 to 14 years old with an average of 9.4 years. All fractures were located on the mid-third part of forearm. Two cases had a re-fracture. Among them, 3 cases caused by high-energy injury and 2 cases by falling down. Open reduction were performed in 4 cases while the other one was treated with closed reduction. Four patients were immobilized in an above-elbow cast, postoperatively. RESULTS: All patients were followed up from 7 to 19 months with an average of 11.4 months. There were 4 delayed union and 1 nonunion. Three patients healed after the removal of the nail and avoidance of weight-bearing. Two patients healed by replacing another fixation. No patients had soft-tissue irritation or nail-entry-site infections.. The clinical effect was evaluated according to Daruwalla and Price scores with 3 excellent and 2 good of the results. CONCLUSIONS: Using titanium elastic nailing for the treatment of pediatric both-bone forearm fractures is a good method. However,strict indication selection should be followed to avoid delayed union or nonunion.