Development and optimization of the Glabridin-loaded dissolving microneedle for enhanced treatment of keloid.

Glabridin (Gla) has been reported to have significant effects in scar treatment, and however, the water insolubility of Gla leads to its poor transdermal absorption ability, which affects its bioactivities. Therefore, we attempted to prepare the Gla dissolving microneedles (Gla-MN) to improve the absorbtion of Gla. After investigation of the 3 factors including the needle tip matrix concentration, the prescription concentration of backing material, and the dissolution method of Gla, we finally determined the process parameters of 10% hyaluronic acid (HA) as the needle tip and 5% polyvinyl alcohol (PVA) as the backing, according to which the Gla-MN was prepared with the good characteristics of high hardness, complete appearance and good in vitro dissolution ability. We then loaded Gla onto the microneedles and measured that the average drug loading of Gla-MN was 2.26 ± 0.11 µg/mg and the cumulative transdermal release of Gla-MN was up to 76.9% after 24 h. In addition, Gla-MN had good skin penetration properties, with Gla-MN penetrating at least 4 to 5 layers of parafilm. And the skin basically could return to normal after 4 h of piercing. Importantly, our results showed that Gla-MN had higher transdermal delivery and therapeutic effects against keloid than that of Gla at the same dosage.

In vitro and in vivo inhibition of the host TRPC4 channel attenuates Zika virus infection.

Zika virus (ZIKV) infection may lead to severe neurological consequences, including seizures, and early infancy death. However, the involved mechanisms are still largely unknown. TRPC channels play an important role in regulating nervous system excitability and are implicated in seizure development. We investigated whether TRPCs might be involved in the pathogenesis of ZIKV infection. We found that ZIKV infection increases TRPC4 expression in host cells via the interaction between the ZIKV-NS3 protein and CaMKII, enhancing TRPC4-mediated calcium influx. Pharmacological inhibition of CaMKII decreased both pCREB and TRPC4 protein levels, whereas the suppression of either TRPC4 or CaMKII improved the survival rate of ZIKV-infected cells and reduced viral protein production, likely by impeding the replication phase of the viral life cycle. TRPC4 or CaMKII inhibitors also reduced seizures and increased the survival of ZIKV-infected neonatal mice and blocked the spread of ZIKV in brain organoids derived from human-induced pluripotent stem cells. These findings suggest that targeting CaMKII or TRPC4 may offer a promising approach for developing novel anti-ZIKV therapies, capable of preventing ZIKV-associated seizures and death.

Chemogenetic Evolution of Diversified Photoenzymes for Enantioselective [2 + 2] Cycloadditions in Whole Cells.

Artificial photoenzymes with novel catalytic modes not found in nature are in high demand; yet, they also present significant challenges in the field of biocatalysis. In this study, a chemogenetic modification strategy is developed to facilitate the rapid diversification of photoenzymes. This strategy integrates site-specific chemical conjugation of various artificial photosensitizers into natural protein cavities and the iterative mutagenesis in cell lysates. Through rounds of directed evolution, prominent visible-light-activatable photoenzyme variants were developed, featuring a thioxanthone chromophore. They successfully enabled the enantioselective [2 + 2] photocycloaddition of 2-carboxamide indoles, a class of UV-sensitive substrates that are traditionally challenging for known photoenzymes. Furthermore, the versatility of this photoenzyme is demonstrated in enantioselective whole-cell photobiocatalysis, enabling the efficient synthesis of enantioenriched cyclobutane-fused indoline tetracycles. These findings significantly expand the photophysical properties of artificial photoenzymes, a critical factor in enhancing their potential for harnessing excited-state reactivity in stereoselective transformations.

[Mechanism of Modified Huoluo Xiaoling Pills against colorectal cancer based on network pharmacology, molecular docking, and experimental validation].

This study aims to predict the possible targets and related signaling pathways of Modified Huoluo Xiaoling Pills against colorectal cancer(CRC) by both network pharmacology and molecular docking and verify the mechanism of action by experiments. TCMSP was used to obtain the active ingredients and targets of Modified Huoluo Xiaoling Pills, and GeneCards, DrugBank, OMIM, and TTD were employed to acquire CRC-related targets. Cytoscape software was utilized to construct the drug-active ingredient-target network, and the STRING database was applied to establish the protein-protein interaction(PPI) network. DAVID platform was adopted to investigate the targets in terms of GO function and KEGG pathway enrichment analysis. Molecular docking was performed in AutoDock Vina. HCT 116 cells were intervened by different concentrations of Modified Huoluo Xiaoling Pills-containing serum, and CCK-8 was used to detect the proliferation inhibition of HCT 116 cells in each group. Transwell was employed to show the invasive abi-lity of HCT 116 cells, and Western blot was taken to reveal the expression levels of ß-catenin, cyclinD1, c-Myc, as well as epithelial-mesenchymal transition(EMT) marker proteins E-cadherin, N-cadherin, vimentin, MMP2, MMP7, MMP9, and TWIST in HCT 116 cells. The network pharmacological analysis yielded 242 active ingredients of Modified Huoluo Xiaoling Pills, 1 844 CRC targets, and 127 overlapping targets of CRC and Modified Huoluo Xiaoling Pills, and the signaling pathways related to CRC involved PI3K-Akt, TNF, HIF-1, IL-17, Wnt, etc. Molecular docking showed that the key active ingredients had a stable binding conformation with the core proteins. CCK-8 indicated that Modified Huoluo Xiaoling Pills significantly inhibited the proliferation of HCT 116 cells. Transwell assay showed that with increasing concentration of Modified Huoluo Xiaoling Pills containing serum, the invasive ability of HCT 116 cells was more obviously inhibited. The expression of ß-catenin, cyclinD1, c-Myc, N-cadherin, vimentin, MMP2, MMP7, MMP9, and TWIST proteins were suppressed, and the expression of E-cadherin was improved by the intervention of drug-containing serum. Thus, it can be seen that Modified Huoluo Xiaoling Pills restrains the proliferation, invasion, and metastasis of CRC cells through multiple components, multiple targets, and multiple pathways, and the mechanism of action may be related to the inhibition of the activation of the Wnt/ß-catenin signaling pathway, thereby affecting the occurrence of EMT.

Identification of the cytochrome P450s responsible for the biosynthesis of two types of aporphine alkaloids and their de novo biosynthesis in yeast.

Aporphine alkaloids have diverse pharmacological activities; however, our understanding of their biosynthesis is relatively limited. Previous studies have classified aporphine alkaloids into two categories based on the configuration and number of substituents of the D-ring and have proposed preliminary biosynthetic pathways for each category. In this study, we identified two specific cytochrome P450 enzymes (CYP80G6 and CYP80Q5) with distinct activities toward (S)-configured and (R)-configured substrates from the herbaceous perennial vine Stephania tetrandra, shedding light on the biosynthetic mechanisms and stereochemical features of these two aporphine alkaloid categories. Additionally, we characterized two CYP719C enzymes (CYP719C3 and CYP719C4) that catalyzed the formation of the methylenedioxy bridge, an essential pharmacophoric group, on the A- and D-rings, respectively, of aporphine alkaloids. Leveraging the functional characterization of these crucial cytochrome P450 enzymes, we reconstructed the biosynthetic pathways for the two types of aporphine alkaloids in budding yeast (Saccharomyces cerevisiae) for the de novo production of compounds such as (R)-glaziovine, (S)-glaziovine, and magnoflorine. This study provides key insight into the biosynthesis of aporphine alkaloids and lays a foundation for producing these valuable compounds through synthetic biology.

Exogenous NADH promotes the bactericidal effect of aminoglycoside antibiotics against Edwardsiella tarda.

The global surge in multidrug-resistant bacteria owing to antibiotic misuse and overuse poses considerable risks to human and animal health. With existing antibiotics losing their effectiveness and the protracted process of developing new antibiotics, urgent alternatives are imperative to curb disease spread. Notably, improving the bactericidal effect of antibiotics by using non-antibiotic substances has emerged as a viable strategy. Although reduced nicotinamide adenine dinucleotide (NADH) may play a crucial role in regulating bacterial resistance, studies examining how the change of metabolic profile and bacterial resistance following by exogenous administration are scarce. Therefore, this study aimed to elucidate the metabolic changes that occur in Edwardsiella tarda (E. tarda), which exhibits resistance to various antibiotics, following the exogenous addition of NADH using metabolomics. The effects of these alterations on the bactericidal activity of neomycin were investigated. NADH enhanced the effectiveness of aminoglycoside antibiotics against E. tarda ATCC15947, achieving bacterial eradication at low doses. Metabolomic analysis revealed that NADH reprogrammed the ATCC15947 metabolic profile by promoting purine metabolism and energy metabolism, yielding increased adenosine triphosphate (ATP) levels. Increased ATP levels played a crucial role in enhancing the bactericidal effects of neomycin. Moreover, exogenous NADH promoted the bactericidal efficacy of tetracyclines and chloramphenicols. NADH in combination with neomycin was effective against other clinically resistant bacteria, including Aeromonas hydrophila, Vibrio parahaemolyticus, methicillin-resistant Staphylococcus aureus, and Listeria monocytogenes. These results may facilitate the development of effective approaches for preventing and managing E. tarda-induced infections and multidrug resistance in aquaculture and clinical settings.

MOF-Derived Nanoparticles with Enhanced Acoustical Performance for Efficient Mechano-Sonodynamic Therapy.

Ultrasound (US) generates toxic reactive oxygen species (ROS) by acting on sonosensitizers for cancer treatment, and the mechanical damage induced by cavitation effects under US is equally significant. Therefore, designing a novel sonosensitizer that simultaneously possesses efficient ROS generation and enhanced mechanical effects is promising. In this study, carbon-doped zinc oxide nanoparticles (C-ZnO) are constructed for mechano-sonodynamic cancer therapy. The presence of carbon (C) doping optimizes the electronic structure, thereby enhancing the ROS generation triggered by US, efficiently inducing tumor cell death. On the other hand, the high specific surface area and porous structure brought about by C doping enable C-ZnO to enhance the mechanical stress induced by cavitation bubbles under US irradiation, causing severe mechanical damage to tumor cells. Under the dual effects of sonodynamic therapy (SDT) and mechanical therapy mediated by C-ZnO, excellent anti-tumor efficacy is demonstrated both in vitro and in vivo, along with a high level of biological safety. This is the first instance of utilizing an inorganic nanomaterial to achieve simultaneous enhancement of ROS production and US-induced mechanical effects for cancer therapy. This holds significant importance for the future development of novel sonosensitizers and advancing the applications of US in cancer treatment.

Tip60-FOXO regulates JNK signaling mediated apoptosis in Drosophila.

The JNK signaling pathway plays crucial roles in various physiological processes, including cell proliferation, differentiation, migration, apoptosis, and stress response. Dysregulation of this pathway is closely linked to the onset and progression of numerous major diseases, such as developmental defects and tumors. Identifying and characterizing novel components of the JNK signaling pathway to enhance and refine its network hold significant scientific and clinical importance for the prevention and treatment of associated cancers. This study utilized the model organism Drosophila and employed multidisciplinary approaches encompassing genetics, developmental biology, biochemistry, and molecular biology to investigate the interplay between Tip60 and the JNK signaling pathway, and elucidated its regulatory mechanisms. Our findings suggest that loss of Tip60 acetyltransferase activity results in JNK signaling pathway activation and subsequent induction of JNK-dependent apoptosis. Genetic epistasis analysis reveals that Tip60 acts downstream of JNK, paralleling with the transcription factor FOXO. The biochemical results confirm that Tip60 can bind to FOXO and acetylate it. Introduction of human Tip60 into Drosophila effectively mitigates apoptosis induced by JNK signaling activation, underscoring conserved regulatory role of Tip60 in the JNK signaling pathway from Drosophila to humans. This study further enhances our understanding of the regulatory network of the JNK signaling pathway. By revealing the role and mechanism of Tip60 in JNK-dependent apoptosis, it unveils new insights and potential therapeutic avenues for preventing and treating associated cancers.

Negative Thermal Expansion in ABC(MoO4)3 Compounds.

Negative thermal expansion (NTE) compounds provide a solution for the mismatch of coefficients of thermal expansion in highly integrated device design. However, the current NTE compounds are rare, and how to effectively design new NTE compounds is still challenging. Here, a new concept is proposed to design NTE compounds, that is, to increase the flexibility of framework structure by expanding the space in framework structure compounds. Taking the parent compound NaZr2(PO4)3 as a case, a new NTE system AIBIICIII(MoO4)3 (A = Li, Na, K, and Rb; B = Mg and Mn; C = Sc, In, and Lu) is designed. In these compounds, the large volume of MoO4 tetrahedron is used to replace the small volume of PO4 tetrahedron in NaZr2(PO4)3 to enhance structural space and NTE performance. Simultaneously, a joint study of temperature-dependent X-ray diffraction, Raman spectroscopy, and the first principles calculation reveals that the NTE in AIBIICIII(MoO4)3 series compounds arise from the coupled oscillation of polyhedral. Large-radius ions are conducive to enhancing the space and softening the framework structure to achieve the enhancement of NTE. The current strategy for designing NTE compounds is expected to be adopted in other compounds to obtain more NTE compounds.

Metabolomics analysis of the lactobacillus plantarum ATCC 14917 response to antibiotic stress.

BACKGROUND: Lactobacillus plantarum has been found to play a significant role in maintaining the balance of intestinal flora in the human gut. However, it is sensitive to commonly used antibiotics and is often incidentally killed during treatment. We attempted to identify a means to protect L. plantarum ATCC14917 from the metabolic changes caused by two commonly used antibiotics, ampicillin, and doxycycline. We examined the metabolic changes under ampicillin and doxycycline treatment and assessed the protective effects of adding key exogenous metabolites. RESULTS: Using metabolomics, we found that under the stress of ampicillin or doxycycline, L. plantarum ATCC14917 exhibited reduced metabolic activity, with purine metabolism a key metabolic pathway involved in this change. We then screened the key biomarkers in this metabolic pathway, guanine and adenosine diphosphate (ADP). The exogenous addition of each of these two metabolites significantly reduced the lethality of ampicillin and doxycycline on L. plantarum ATCC14917. Because purine metabolism is closely related to the production of reactive oxygen species (ROS), the results showed that the addition of guanine or ADP reduced intracellular ROS levels in L. plantarum ATCC14917. Moreover, the killing effects of ampicillin and doxycycline on L. plantarum ATCC14917 were restored by the addition of a ROS accelerator in the presence of guanine or ADP. CONCLUSIONS: The metabolic changes of L. plantarum ATCC14917 under antibiotic treatments were determined. Moreover, the metabolome information that was elucidated can be used to help L. plantarum cope with adverse stress, which will help probiotics become less vulnerable to antibiotics during clinical treatment.

Inclusion Complexes of Ethanamizuril with ß- and Hydroxypropyl-ß-Cyclodextrin in Aqueous Solution and in Solid State: A Comparison Study.

Ethanamizuril (EZL) is a new anticoccidial drug developed by our Shanghai Veterinary Research Institute. Since EZL is almost insoluble in water, we conducted a study to improve the solubility of EZL by forming inclusion complexes with ß-cyclodextrin (ß-CD) and hydroxypropyl-ß-cyclodextrin (HP-ß-CD). In this study, we performed molecular docking and then systematically compared the interactions of EZL with ß-CD and HP-ß-CD in both aqueous solution and the solid state, aiming to elucidate the solubilization effect and mechanism of cyclodextrins (CDs). The interactions were also examined in the solid state using DSC, PXRD, and FT-IR. The interactions of EZL with CDs in an aqueous solution were investigated using PSA, UV-vis spectroscopy, MS, 1H NMR, and 2D ROESY. The results of phase solubility experiments revealed that both ß-CD and HP-ß-CD formed inclusion complexes with EZL in a 1:1 molar ratio. Among them, HP-ß-CD exhibited higher Kf (stability constant) and CE (complexation efficiency) values as well as a stronger solubilization effect. Furthermore, the two cyclodextrins were found to interact with EZL in a similar manner. The results of our FT-IR and 2D ROESY experiments are in agreement with the theoretical results derived from molecular simulations. These results indicated that intermolecular hydrogen bonds existing between the C=O group on the triazine ring of EZL and the O-H group of CDs, as well as the hydrophobic interactions between the hydrogen on the benzene ring of EZL and the hydrogen of CDs, played crucial roles in the formation of EZL/CD inclusion complexes. The results of this study can lay the foundation for the future development of high-concentration drinking water delivery formulations for EZL.

Visual analysis of the research frontiers, hotspots and trends of exercise therapy intervention in tumor-related sleep-wake disorders.

Objective: To systematically understand the research frontiers, hotspots and development trends of exercise therapy in the intervention of tumor-related sleep-wake disorders, and to provide scientific basis for follow-up research. Methods: Downloaded the original research papers on February 26, 2024, from the Web of Science core collection database, on tumor-associated sleep-wake disorders. The data that met the inclusion criteria were imported into the Bibliometric Analysis Platform (http://biblimetric.com), CiteSpace 6.3.R1 and VOSviwer1.6.20 software for visual analysis, and imported into Excel2021. Scientometric analysis was performed with Oringin2021 and PyCharm Community Edition 2022.1.3. Results: A total of 512 original research papers on tumor-related sleep-wake disorders were obtained. The most influential countries in the subject area are the United States, Spain and German, the institutions are the University of California System, Sun Yat Sen University and Northwestern University, et al., the authors are Berger AM, Aaronson NK, Bower JE, et al., and the journals are Cancer, Brit J Cancer and Cancer Nurs. The co-cited references suggest that the current research frontier in the field mainly involves the level, place and program of exercise therapy, including the relationship between physical activity, sedentary behavior and cancer prevention and control. The results of co-occurrence keyword network analysis showed that quality of life, physical activity, breast cancer, exercise, fatigue, and survivors may be the research hotspots in this field, with breast cancer, health, aerobic exercise, adults, and chemotherapy being the most popular. Conclusions: The number of papers published and the research enthusiasm in this field show a steady upward trend. However, there is a lack of influential institutions and scholars, and there is relatively little research collaboration across countries/regions/institutions. The scientific research influence of institutions and scholars in most European and American countries/regions is significantly ahead of that of institutions and scholars in Asian and African countries/regions. But Sun Yat Sen University in China is a relatively active and influential scientific research institution in recent years, which is worthy of attention. In addition, the research frontier of this discipline is the level, place and program of exercise therapy auxiliary intervention, and the research hotspots involve breast cancer, health, aerobic exercise, adults, chemotherapy, et al. Their clinical efficacy needs to be further demonstrated in multi-center, large-sample and high-quality prospective studies.

Adipose-derived stem cells ameliorate radiation-induced lung injury by activating the DDAH1/ADMA/eNOS signaling pathway.

Background: Ionizing radiation-induced lung injury is caused by the initial inflammatory reaction and leads to advanced fibrosis of lung tissue. Adipose-derived stem cells (ASCs) are a type of mesenchymal stem cell that can differentiate into various functional cell types with broad application prospects in the treatment of tissue damage. The purpose of this study was to explore the protective effect of ASCs against radiation-induced lung injury and to provide a novel basis for prevention and treatment of radiation-induced lung injury. Materials and methods: Fifty mice were randomly divided into a control group (Ctrl), radiation exposure group (IR), radiation exposure plus ASC treatment group (IR + ASC), radiation exposure plus L-257 group (IR + L-257), and radiation exposure plus ASC treatment and L-257 group (IR + ASC + L-257). Mice in IR, IR + ASC, and IR + ASC + L-257 groups were exposed to a single whole-body dose of 5 Gy X-rays (160 kV/25 mA, 1.25 Gy/min). Within 2 h after irradiation, mice in IR + ASC and IR + ASC + L-257 groups were injected with 5 × 106 ASCs via the tail vein. Mice in IR + L-257 and IR + ASC + L-257 groups were intraperitoneally injected with 30 mg/kg L-257 in 0.5 mL saline. Results: The mice in the IR group exhibited lung hemorrhage, edema, pulmonary fibrosis, and inflammatory cell infiltration, increased release of proinflammatory cytokines, elevation of oxidative stress and apoptosis, and inhibition of the dimethylarginine dimethylamino hydratase 1 (DDAH1)/ADMA/eNOS signaling pathway. ASC treatment alleviated radiation-induced oxidative stress, apoptosis, and inflammation, and restored the DDAH1/ADMA/eNOS signaling pathway. However, L-257 pretreatment offset the protective effect of ASCs against lung inflammation, oxidative stress, and apoptosis. Conclusions: These data suggest that ASCs ameliorate radiation-induced lung injury, and the mechanism may be mediated through the DDAH1/ADMA/eNOS signaling pathway.

rESWT promoted angiogenesis via Bach1/Wnt/ß-catenin signaling pathway.

Previous reports have established that rESWT fosters angiogenesis, yet the mechanism by which rESWT promotes cerebral angiogenesis remains elusive. rESWT stimulated HUVECs proliferation as evidenced by the CCK-8 test, with an optimal dosage of 2.0 Bar, 200 impulses, and 2 Hz. The tube formation assay of HUVECs revealed that tube formation peaked at 36 h post-rESWT treatment, concurrent with the lowest expression level of Bach1, as detected by both Western blot and immunofluorescence. The expression level of Wnt3a, ß-catenin, and VEGF also peaked at 36 h. A Bach1 overexpression plasmid was transfected into HUVECs, resulting in a decreased expression level of Wnt3a, ß-catenin, and VEGF. Upon treatment with rESWT, the down-regulation of Wnt3a, ß-catenin, and VEGF expression in the transfected cells was reversed. The Wnt/ß-catenin inhibitor DKK-1 was utilized to suppress Wnt3a and ß-catenin expression, which led to a concurrent decrease in VEGF expression. However, rESWT treatment could restore the expression of these three proteins, even in the presence of DKK-1. Moreover, in the established OGD model, it was observed that rESWT could inhibit the overexpression of Bach1 and enhance VEGF and VEGFR-2 expression under the OGD environment.

Assessment of cognitive dysfunction and its influencing factors after acute ischemic stroke.

OBJECTIVE: In order to provide a more accurate and effective basis for clinical diagnosis and treatment, patients with cognitive dysfunction after acute ischemic stroke (AIS) were evaluated and their influencing factors were analyzed. METHODS: A rigorous and systematic logistic regression analysis was conducted to comprehensively investigate the various influencing factors that contribute to cognitive dysfunction. RESULTS: Among them, the sex granulocyte/lymphocyte ratio (NLR), low-density lipoprotein cholesterol (LDL-C) level, and C-reactive protein (CRP) were also higher than those in the control group (p < 0.05). The scores of memory, orientation, visual and spatial function, abstract thinking and language in the control group were higher than those in the experimental group (p < 0.05). The results of multivariate logistic regression analysis showed that history of diabetes mellitus, high NLR, high LDL-C, high CRP, smoking and temporal lobe infarction were risk factors for cognitive dysfunction after AIS, while elevated BMI and love of exercise were protective factors for cognitive dysfunction after AIS. CONCLUSION: Patients with cognitive dysfunction had the highest incidence of temporal lobe infarction, and they scored lower than the control group on memory, orientation, visual and spatial function, abstract thinking, and language function. Multivariate logistic regression analysis showed that a history of diabetes mellitus, high NLR, high LDL-C, high CRP, smoking, and temporal lobe infarction were independent risk factors for cognitive dysfunction after acute ischemic stroke, while elevated BMI and a love of exercise were protective factors for cognitive dysfunction after acute ischemic stroke.

Hai-Honghua medicinal liquor is a reliable remedy for fracture by promotion of osteogenic differentiation via activation of PI3K/Akt pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Hai-Honghua medicinal liquor (HHML), an external Chinese herbal formula preparation, is often applied to treat freshly closed tibia/fibular fractures, ankle fractures, and other bone-related disorders, but the related molecular mechanism is unclear. AIM OF THE STUDY: To evaluate the therapeutic effect of HHML in patients with tibial/fibular and ankle fractures, and to explore its related possible mechanism. METHODS AND MATERIALS: A total of 182 patients with tibia/fibular fractures and 183 patients with ankle fractures were enrolled in this study. A randomized, controlled, unblinded clinical trial was designed to evaluate the therapeutic effect of HHML on tibial/fibular and ankle fractures. The chemical compositions of HHML were analyzed by the HPLC-Q-Extractive MS/MS. Furthermore, a rat tibial fracture model was established to evaluate the therapeutic effects of HHML in promoting fracture healing, and the mouse embryonic osteoblasts cell line of MC3T3-E1 was further carried out to explore the mechanisms of HHML on osteoblast differentiation. RESULTS: In the clinical evaluation, HHML treatment significantly shortened the time for pain and swelling in patients with tibial/fibular fractures (P < 0.01) and ankle fractures (P < 0.01), and the incidence of complications was significantly reduced as well. Subsequently, 116 constituents were identified from HHML via HPLC-Q-TOF-MS/MS analysis. In vivo, no obvious changes in weight were observed in HHML-treated rats. Moreover, the levels of bone formation markers (including osteocalcin (OCN), N-terminal propeptide of type I procollagen (PINP), alkaline phosphatase (ALP), calcium (Ca) and substance P) in rat serum were significantly increased in HHML-treated rats compared with model rats (P < 0.05). Micro-CT analysis showed bone mineral density (BMD), bone volume fraction (BV/TV), trabecular thickness (Tb.Th) of the HHML-treated rats were significantly increased (P < 0.05, vs. Model) while trabecular separation (Tb.Sp) and structure model index (SMI) values were significantly reduced (P < 0.05, vs. Model). Histological analysis showed that HHML treatment promoted the healing of fractures and cartilage repair, and increased the osteoblasts and collagen fibers. Furthermore, our results also revealed HHML could promote MC3T3-E1 cells proliferation and osteoblast differentiation via regulation of the runt-related transcription factor 2 (RUNX2), bone alkaline phosphatase (BALP), and OCN by activating phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) pathway, which confirmed by adding PI3K chemical inhibitor of LY294002. CONCLUSION: HHML treatment is a reliable remedy for fractures in tibial and ankle by promotion of osteogenic differentiation via activation of PI3K/Akt pathway.

Extraction and analysis of high-quality chloroplast DNA with reduced nuclear DNA for medicinal plants.

BACKGROUND: Obtaining high-quality chloroplast genome sequences requires chloroplast DNA (cpDNA) samples that meet the sequencing requirements. The quality of extracted cpDNA directly impacts the efficiency and accuracy of sequencing analysis. Currently, there are no reported methods for extracting cpDNA from Erigeron breviscapus. Therefore, we developed a suitable method for extracting cpDNA from E. breviscapus and further verified its applicability to other medicinal plants. RESULTS: We conducted a comparative analysis of chloroplast isolation and cpDNA extraction using modified high-salt low-pH method, the high-salt method, and the NaOH low-salt method, respectively. Subsequently, the number of cpDNA copies relative to the nuclear DNA (nDNA ) was quantified via qPCR. As anticipated, chloroplasts isolated from E. breviscapus using the modified high-salt low-pH method exhibited intact structures with minimal cell debris. Moreover, the concentration, purity, and quality of E. breviscapus cpDNA extracted through this method surpassed those obtained from the other two methods. Furthermore, qPCR analysis confirmed that the modified high-salt low-pH method effectively minimized nDNA contamination in the extracted cpDNA. We then applied the developed modified high-salt low-pH method to other medicinal plant species, including Mentha haplocalyx, Taraxacum mongolicum, and Portulaca oleracea. The resultant effect on chloroplast isolation and cpDNA extraction further validated the generalizability and efficacy of this method across different plant species. CONCLUSIONS: The modified high-salt low-pH method represents a reliable approach for obtaining high-quality cpDNA from E. breviscapus. Its universal applicability establishes a solid foundation for chloroplast genome sequencing and analysis of this species. Moreover, it serves as a benchmark for developing similar methods to extract chloroplast genomes from other medicinal plants.

[Chidamide Promotes Osteogenic Differentiation of Bone Marrow Mesenchymal Stromal Cells from Patients with Myelodysplastic Syndromes].

OBJECTIVE: To explore the effects and mechanisms of chidamide on the osteogenic differentiation of bone marrow mesenchymal stromal cells (MSC) from myelodysplastic syndromes (MDS). METHODS: MSC were isolated and cultured from bone marrow of MDS patients and healthy donors. CCK-8 assay was used to detect the effects of chidamide on the proliferation of MSC. The effects of chidamide on the activity of histone deacetylase (HDAC) in MSC was measured by a fluorescence assay kit and Western blot. Alkaline phosphatase (ALP) activity was detected on day 3 and calcium nodule formation was observed by Alizarin Red staining on day 21 after osteogenic differentiation. The expression of early and late osteogenic genes was detected on day 7 and day 21, respectively. RT-PCR and Western blot were used to detect the effects of chidamide on mRNA and protein expression of RUNX2 which is the key transcription factor during osteogenesis. RESULTS: As the concentration of chidamide increased, the proliferation of MSC was inhibited. However, at a low concentration (1 µmol/L), chidamide had no significant inhibitory effect on MSC proliferation but significantly inhibited HDAC activity. In MSC from both MDS patients and healthy donors, chidamide (1 µmol/L) significantly increased ALP activity, calcium nodule formation, thereby mRNA expression of osteogenic genes, and restored the reduced osteogenic differentiation ability of MDS-MSC compared to normal MSC. Mechanistic studies showed that the osteogenic-promoting effect of chidamide may be related to the upregulation of RUNX2 . CONCLUSION: Chidamide can inhibit HDAC activity in MSC, upregulate the expression of the osteogenic transcription factor RUNX2, and promote the osteogenic differentiation of MDS-MSC.

Opening the black box: explainable deep-learning classification of wood microscopic image of endangered tree species.

BACKGROUND: Traditional method of wood species identification involves the use of hand lens by wood anatomists, which is a time-consuming method that usually identifies only at the genetic level. Computer vision method can achieve "species" level identification but cannot provide an explanation on what features are used for the identification. Thus, in this study, we used computer vision methods coupled with deep learning to reveal interspecific differences between closely related tree species. RESULT: A total of 850 images were collected from the cross and tangential sections of 15 wood species. These images were used to construct a deep-learning model to discriminate wood species, and a classification accuracy of 99.3% was obtained. The key features between species in machine identification were targeted by feature visualization methods, mainly the axial parenchyma arrangements and vessel in cross section and the wood ray in tangential section. Moreover, the degree of importance of the vessels of different tree species in the cross-section images was determined by the manual feature labeling method. The results showed that vessels play an important role in the identification of Dalbergia, Pterocarpus, Swartzia, Carapa, and Cedrela, but exhibited limited resolutions on discriminating Swietenia species. CONCLUSION: The research results provide a computer-assisted tool for identifying endangered tree species in laboratory scenarios, which can be used to combat illegal logging and related trade and contribute to the implementation of CITES convention and the conservation of global biodiversity.

Polyvinyl alcohol/kappa-carrageenan-based package film with simultaneous incorporation of ferric ion and polyphenols from Capsicum annuum leaves for fruit shelf-life extension.

Bio-based food packaging materials have elicited growing interests due to their great degradability, high safety and active biofunctions. In this work, by simultaneously introducing the polyphenolic extracts from Capsicum annuum leaves and ferric ion (Fe3+) into the Polyvinyl alcohol/kappa-carrageenan (PVA/κ-carrageenan)-based film-forming matrix, an active package film was developed, with the purpose to improve the food shelf life. The experimental results indicated that the existence of Fe3+ can not only improve the mechanical properties owing to the multiple dynamic coordinated interactions, but also endow the composite films with excellent fire-retardancy. Moreover, the composite films could display excellent UV resistant performance, water vapor/oxygen gas barrier properties and antioxidant activities with the corporation of polyphenols. In particular, the highest DPPH and ABTS radical scavenging capacities for composite film (PC-PLP7 sample) were evaluated to be 82.5 % and 91.1 %, respectively. Higher polyphenol concentration is favorable to the bio-functions of the materials. Benefitting from these features, this novel kind of films with a dense and steady micro-structure could be further applicated in fruit preservations, where the ripening bananas were ensured with the high storage quality. This integration as a prospective food packaging material provides an economic and eco-friendly approach to excavate the high added-values of biomass.