MicroRNA-143-3p ameliorates collagen-induced arthritis by polarizing naive CD4+ T cells into Treg cells.

BACKGROUND: Rheumatoid arthritis (RA) is a persistent and systemic autoimmunity disease. The abnormal differentiation of Treg cells is important in pathogenesis. Despite previous studies showed that microRNAs (miRNAs, miR) are pivotal modulators of Treg cells, the effect of miRNAs on Treg cell differentiation and function is not clear. Our study wants to reveal the relationship of miR-143-3p with the differentiative ability and biofunction of Treg cells during the development of RA. METHODS: The Expressing level of miR-143-3p and cell factor generation in peripheral blood (PB) of RA sufferers were identified by ELISA or RT-qPCR. The roles of miR-143-3p in Treg cell differentiation were studied via ShRNA/lentivirus transfection. Male DBA/1 J mice were separated into control, model, control mimics, and miR-143-3p mimics groups to analyze the anti-arthritis efficacy, the differentiative ability of Treg cells, and the expression level of miR-143-3p. RESULTS: Our team discovered that the Expressing level of miR-143-3p was related to RA disease activities in a negative manner, and remarkably related to antiinflammation cell factor IL-10. In vitro, the expression of miR-143-3p in the CD4+ T cells upregulated the percentage of CD4+ CD25+ Fxop3+ cells (Tregs) and forkhead box protein 3 (Foxp3) mRNA expression. Evidently, miR-143-3p mimic intervention considerably upregulated the content of Treg cells in vivo, validly avoided CIA progression, and remarkably suppressed the inflammatory events of joints in mice. CONCLUSION: Our findings indicated that miR-143-3p could ameliorate CIA through polarizing naive CD4+ T cells into Treg cells, which may be a novel strategy to treat autoimmune diseases such as RA.

Natural Variation Confers 'Aiyuan 38' Citrus Mutant a New Color and Unique Flavor.

Citrus exhibits unique nutritional values. Most citrus cultivars are derived from mutations. However, the effect of these mutations on fruit quality is unclear. We have previously found a yellowish bud mutant in the citrus cultivar 'Aiyuan 38'. Therefore, this study aimed to determine the effect of the mutation on fruit quality. 'Aiyuan 38' (WT) and a bud mutant variant (MT) were used to analyze variations in fruit color variation and flavor substances using colorimetric instruments, high-performance liquid chromatography (HPLC), headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS), and odor activity values (OAVs). The mutation in MT conferred yellowish characteristics to its peel. Although the differences in total sugar and acid content of the pulp were not statistically significant between WT and MT, the MT glucose content was significantly lower and the malic acid level was significantly higher. HS-SPME-GC-MS analysis revealed that the MT pulp released more types and contents of volatile organic compounds (VOCs) than the WT, whereas the opposite trend was observed for the peel. Analysis of the OAV revealed that the MT pulp contains 6 unique VOCs, whereas the peel contains only 1. This study provides a useful reference for the study of flavor substances associated with citrus bud mutations.

Effect of Natural Variation and Rootstock on Fruit Quality and Volatile Organic Compounds of 'Kiyomi tangor' (Citrus reticulata Blanco) Citrus.

In this study, we compared the fruit quality and color of 'Kiyomi' (WT) and its mutant (MT) grafted on Ziyang xiangcheng (Cj) (WT/Cj, MT/Cj), and the MT grafted on Trifoliate orange (Pt) (MT/Pt). The differences in sugar, organic acid, flavonoids, phenols, and volatile substances of the three materials were also analyzed by high performance liquid chromatography (HPLC) and headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS). The results showed significant differences in the appearance of WT/Cj, MT/Cj, and MT/Pt. MT/Pt, compared to WT/Cj, MT/Cj, had lower sugar, acid, phenol and flavonoid contents in the pulp. However, MT/Pt pulp was higher in vitamin C (VC), and the peel had significantly higher total phenol and flavonoid contents. In terms of pulp, WT/Cj had the greatest diversity of volatile organic compounds (VOCs). 4-methyl-1-pentanol was significantly higher in MT/Cj pulp, while MT/Pt pulp had a unique octanoic acid, methyl ester. VOCs were more diverse in the peels of the three materials. ß-Myrcene and valencen were significantly higher in MT/Cj peels. In contrast, 16 unique VOCs were detected in MT/Pt, and D-limonene content was significantly higher than in WT/Cj and MT/Cj. The results suggest Trifoliate orange is a suitable rootstock for MT.

Analysis of Codon Usage Bias in Xyloglucan Endotransglycosylase (XET) Genes.

Xyloglucan endotransglycosylase (XET) genes are widely distributed in most plants, but the codon usage bias of XET genes has remained uncharacterized. Thus, we analyzed the codon usage bias using 4500 codons of 20 XET genes to elucidate the genetic and evolutionary patterns. Phylogenetic and hierarchical cluster analyses revealed that the 20 XET genes belonged to two groups. The closer the genetic distance, the more similar the codon usage preference. The codon usage bias of most XET genes was weak, but there was also some codon usage bias. AGA, AGG, AUC, and GUG were the top four codons (RSCU > 1.5) in the 20 XET genes. CitXET had a stronger codon usage bias, and there were eight optimal codons of CitXET (i.e., AGA, AUU, UCU, CUU, CCA, GCU, GUU, and AAA). The RSCU values underwent a correspondence analysis. The two main factors affecting codon usage bias (i.e., Axes 1 and 2) accounted for 54.8% and 17.6% of the total variation, respectively. Multiple correspondence analysis revealed that XET genes were widely distributed, with Group 1 genes being closer to Axis 1 than Group 2 genes, which were closer to Axis 2. Codons with A/U at the third codon position were distributed closer to Axis 1 than codons with G/C at the third codon position. PgXET, ZmXET, VlXET, VrXET, and PcXET were biased toward codons ending with G/C. In contrast, CitXET, DpXET, and BrpXET were strongly biased toward codons ending with A/U, indicating that these XET genes have a strong codon usage bias. Translational selection and base composition (especially A and U at the third codon position), followed by mutation pressure and natural selection, may be the most important factors affecting codon usage of 20 XET genes. These results may be useful in clarifying the codon usage bias of XET genes and the relevant evolutionary characteristics.

Identification of Photosynthesis Characteristics and Chlorophyll Metabolism in Leaves of Citrus Cultivar (Harumi) with Varying Degrees of Chlorosis.

In autumn and spring, citrus leaves with a Ponkan (Citrus reticulata Blanco cv. Ponkan) genetic background (Harumi, Daya, etc.) are prone to abnormal physiological chlorosis. The effects of different degrees of chlorosis (normal, mild, moderate and severe) on photosynthesis and the chlorophyll metabolism of leaves of Citrus cultivar (Harumi) were studied via field experiment. Compared with severe chlorotic leaves, the results showed that chlorosis could break leaf metabolism balance, including reduced chlorophyll content, photosynthetic parameters, antioxidant enzyme activity and enzyme activity related to chlorophyll synthesis, increased catalase and decreased enzyme activity. In addition, the content of chlorophyll synthesis precursors showed an overall downward trend expected for uroporphyrinogen III. Furthermore, the relative expression of genes for chlorophyll synthesis (HEMA1, HEME2, HEMG1 and CHLH) was down-regulated to some extent and chlorophyll degradation (CAO, CLH, PPH, PAO and SGR) showed the opposite trend with increased chlorosis. Changes in degradation were more significant. In general, the chlorosis of Harumi leaves might be related to the blocked transformation of uroporphyrinogen III (Urogen III) to coproporphyrinogen III (Coprogen III), the weakening of antioxidant enzyme system activity, the weakening of chlorophyll synthesis and the enhancement in degradation.

Effects of Exogenous Application of Glycine Betaine Treatment on 'Huangguoggan' Fruit during Postharvest Storage.

Loss of quality in citrus fruit is a common occurrence during postharvest storage due to oxidative stress and energy consumption. In recent years, glycine betaine (GB) has been widely applied to postharvest horticulture fruit. This study aimed to investigate the effect of GB treatment (10 mM and 20 mM) on the quality and antioxidant activity of 'Huangguogan' fruit during postharvest storage at room temperature. Our results indicated that both 10 mM and 20 mM treatments effectively reduced weight and firmness losses and maintained total soluble solid (TSS), titratable acidity (TA), and ascorbic acid contents. Additionally, GB treatment significantly increased the activity of antioxidant enzymes, maintained higher levels of total phenols and total flavonoids, and led to slower accumulation of H2O2. A transcriptome analysis conducted at 28 days after treatment (DAT)identified 391 differentially expressed genes (DEGs) between 20 mM GB (GB-2) and the control (CK) group. These DEGs were enriched in various pathways, particularly related to oxygen oxidoreductase, peroxidase activity, and flavonoid biosynthesis. Overall, the application of GB proved beneficial in enhancing the storability and extending the shelf life of 'Huangguogan' fruit.

Panax notoginseng saponins (PNS) attenuate Th17 cell differentiation in CIA mice via inhibition of nuclear PKM2-mediated STAT3 phosphorylation.

CONTEXT: Rheumatoid arthritis (RA) is an autoimmune disease with aberrant Th17 cell differentiation. Panax notoginseng (Burk.) F. H. Chen (Araliaceae) saponins (PNS) have an anti-inflammatory effect and can suppress Th17 cell differentiation. OBJECTIVE: To investigate mechanisms of PNS on Th17 cell differentiation in RA, and the role of pyruvate kinase M2 (PKM2). MATERIALS AND METHODS: Naive CD4+T cells were treated with IL-6, IL-23 and TGF-ß to induce Th17 cell differentiation. Apart from the Control group, other cells were treated with PNS (5, 10, 20 µg/mL). After the treatment, Th17 cell differentiation, PKM2 expression, and STAT3 phosphorylation were measured via flow cytometry, western blots, or immunofluorescence. PKM2-specific allosteric activator (Tepp-46, 50, 100, 150 µM) and inhibitor (SAICAR, 2, 4, 8 µM) were used to verify the mechanisms. A CIA mouse model was established and divided into control, model, and PNS (100 mg/kg) groups to assess an anti-arthritis effect, Th17 cell differentiation, and PKM2/STAT3 expression. RESULTS: PKM2 expression, dimerization, and nuclear accumulation were upregulated upon Th17 cell differentiation. PNS inhibited the Th17 cells, RORγt expression, IL-17A levels, PKM2 dimerization, and nuclear accumulation and Y705-STAT3 phosphorylation in Th17 cells. Using Tepp-46 (100 µM) and SAICAR (4 µM), we demonstrated that PNS (10 µg/mL) inhibited STAT3 phosphorylation and Th17 cell differentiation by suppressing nuclear PKM2 accumulation. In CIA mice, PNS attenuated CIA symptoms, reduced the number of splenic Th17 cells and nuclear PKM2/STAT3 signaling. DISCUSSION AND CONCLUSIONS: PNS inhibited Th17 cell differentiation through the inhibition of nuclear PKM2-mediated STAT3 phosphorylation. PNS may be useful for treating RA.

OsUGE3-mediated cell wall polysaccharides accumulation improves biomass production, mechanical strength, and salt tolerance.

Cell walls constitute the majority of plant biomass and are essential for plant resistance to environmental stresses. It is promising to improve both plant biomass production and stress resistance simultaneously by genetic modification of cell walls. Here, we report the functions of a UDP-galactose/glucose epimerase 3 (OsUGE3) in rice growth and salt tolerance by characterizing its overexpressing plants (OsUGE3-OX) and loss-of-function mutants (uge3). The OsUGE3-OX plants showed improvements in biomass production and mechanical strength, whereas uge3 mutants displayed growth defects. The OsUGE3 exhibits UDP-galactose/glucose epimerase activity that provides substrates for polysaccharides polymerization, consistent with the increased biosynthesis of cellulose and hemicelluloses and strengthened walls in OsUGE3-OX plants. Notably, the OsUGE3 is ubiquitously expressed and induced by salt treatment. The uge3 mutants were hypersensitive to salt and osmotic stresses, whereas the OsUGE3-OX plants showed improved tolerance to salt and osmotic stresses. Moreover, OsUGE3 overexpression improves the homeostasis of Na+ and K+ and induces a higher accumulation of hemicelluloses and soluble sugars during salt stress. Our results suggest that OsUGE3 improves biomass production, mechanical strength, and salt stress tolerance by reinforcement of cell walls with polysaccharides and it could be targeted for genetic modification to improve rice growth under salt stress.

CsMYB96 confers resistance to water loss in citrus fruit by simultaneous regulation of water transport and wax biosynthesis.

A Citrus sinensis R2R3 MYB transcription factor (CsMYB96) has previously been shown to be strongly associated with the expression of many genes related to wax biosynthesis in the fruit. In this study, CsMYB96 was found to alleviate water loss by simultaneously regulating the expression of genes encoding plasma membrane intrinsic proteins (CsPIPs) and wax-related genes. Expression profiling indicated that CsPIP1;1 and CsPIP2;4 had high expression that was representative of other aquaporins, and they were down-regulated in the peel of post-harvest citrus fruit. CsPIP2;4 was further characterized as the predominant CsPIP, with high expression and high-water channel activity. Transient overexpression of CsPIP2;4 accelerated water loss in citrus fruit. In silico analysis further indicated that the expression of CsMYB96 had a significant negative correlation with that of CsPIPs. In vivo and in vitro experiments confirmed that CsMYB96 was able to directly repress the expression of CsPIPs. In addition, CsMYB96 was able to activate wax-related genes and promote wax biosynthesis for defense against water loss. Transient and stable overexpression of CsMYB96 reduced water loss from both citrus fruit and Arabidopsis.

CitWRKY28 and CitNAC029 promote the synthesis of cuticular wax by activating CitKCS gene expression in citrus fruit.

KEY MESSAGE: CitWRKY28 and CitNAC029 are involved in cuticular wax synthesis as indicated by the comparative analysis of fruit aliphatic wax content between Citrus reticulata and Citrus trifoliata and gene co-expression analysis. Cuticular wax covers the fruit surface, playing important roles in reduction of fruit water loss and resistance to pathogen invasion. However, there is limited research on the synthesis and transcriptional regulation of cuticular wax in citrus fruit. In this study, we characterized the variations of aliphatic wax in HJ (Citrus reticulata) and ZK (Citrus trifoliata) from young fruit to mature fruit, as well as performed transcriptome sequencing on 27 samples at different fruit developmental stages. The results revealed that the ZK fruit always had a higher aliphatic wax content than the HJ fruit during development. qRT-PCR analysis demonstrated that two KCS genes, CitKCS1 and CitKCS12, had the most significant difference in expression between HJ and ZK. Furthermore, a heterologous expression assay in Arabidopsis indicated that CitKCS1 and CitKCS12 are involved in cuticular wax synthesis. Subsequently, gene co-expression network analysis screened CitWRKY28 and CitNAC029. Dual luciferase and EMSA assays indicated that CitWRKY28 might bind to the promoter of CitKCS1 and CitKCS12 and CitNAC029 might bind to that of CitKCS1 to activate their expression. Moreover, CitWRKY28 and CitNAC029 could promote the accumulation of cuticular wax in Arabidopsis leaves. Our findings provide new insights into the synthesis and regulation of cuticular wax and valuable information for further mining of wax-related genes in citrus fruit.

Triptolide inhibits Th17 differentiation via controlling PKM2-mediated glycolysis in rheumatoid arthritis.

CONTEXT: Rheumatoid arthritis (RA) is an autoimmune disease with the aberrant differentiation of T helper 17 (Th17) cells. Pyruvate kinase M2 (PKM2), a key enzyme of glycolysis, was associated with Th17 cell differentiation. AIM: To investigate the potential therapeutic effects of triptolide (TP) in collagen-induced arthritis (CIA) and Th17 cell differentiation, and elucidated the underlying mechanisms. METHODS: PKM2 expression and IL-17A production in peripheral blood of RA patients were detected by RT-qPCR or ELISA. Flow cytometry and ELISA were employed to assess the effect of Th17 cell differentiation by TP. PKM2 expression and other glycolysis-related factors were detected using RT-qPCR and Western Blot. PKM2 specific inhibitor Compound 3 K was used to verify the mechanisms. Male DBA/1J mice were divided into control, model, and TP (60 µg/kg) groups to assess the anti-arthritis effect, Th17 cell differentiation and PKM2 expression. RESULTS: PKM2 expression positively correlated with IL-17A production in RA patients. PKM2 expression was increased upon Th17 cell differentiation. Down-regulating PKM2 expression could strongly reduce Th17 cell differentiation. Molecular docking analysis predicted that TP targeted PKM2. TP treatment significantly reduced Th17 cell differentiation, PKM2 expression, pyruvate, and lactate production. In addition, compared with down-regulating PKM2 alone (Compound 3 K treatment), co-treatment with TP and Compound 3 K further significantly decreased PKM2-mediated glycolysis and Th17 cell differentiation. In CIA mice, TP repressed the PKM2-mediated glycolysis and attenuated joint inflammation. CONCLUSION: TP inhibited Th17 cell differentiation through the inhibition of PKM2-mediated glycolysis. We highlight a novel strategy for the use of TP in RA treatment.

Polycrystalline Few-Layer Graphene as a Durable Anticorrosion Film for Copper.

Corrosion of metals in atmospheric environments is a worldwide problem in industry and daily life. Traditional anticorrosion methods including sacrificial anodes or protective coatings have performance limitations. Here, we report atomically thin, polycrystalline few-layer graphene (FLG) grown by chemical vapor deposition as a long-term protective coating film for copper (Cu). A six-year old, FLG-protected Cu is visually shiny and detailed material characterizations capture no sign of oxidation. The success of the durable anticorrosion film depends on the misalignment of grain boundaries between adjacent graphene layers. Theoretical calculations further found that corrosive molecules always encounter extremely high energy barrier when diffusing through the FLG layers. Therefore, the FLG is able to prevent the corrosive molecules from reaching the underlying Cu surface. This work highlights the interesting structures of polycrystalline FLG and sheds insight into the atomically thin coatings for various applications.

Transcriptomic Identification of Wheat AP2/ERF Transcription Factors and Functional Characterization of TaERF-6-3A in Response to Drought and Salinity Stresses.

AP2/ERF (APETALA2/ethylene responsive factor) is a family of plant-specific transcription factors whose members are widely involved in many biological processes, such as growth, development, and biotic and abiotic stress responses. Here, 20 AP2/ERF genes were identified based on wheat RNA-seq data before and after drought stress, and classified as AP2, ERF, DREB, and RAV. The analysis of gene structure revealed that about 85% of AP2/ERF family members had lost introns, which are presumed to have been lost during the formation and evolution of the wheat genome. The expression of 20 AP2/ERF family genes could be verified by qRT-PCR, which further supported the validity of the RNA-seq data. Subsequently, subcellular localization and transcriptional activity experiments showed that the ERF proteins were mainly located in the nucleus and were self-activating, which further supports their functions as transcription factors. Furthermore, we isolated a novel ERF gene induced by drought, salt, and cold stresses and named it TaERF-6-3A. TaERF-6-3A overexpression increased sensitivity to drought and salt stresses in Arabidopsis, which was supported by physiological and biochemical indices. Moreover, the expression of stress- and antioxidant-related genes was downregulated in TaERF-6-3A-overexpressing plants. Overall, these results contribute to the further understanding of the TaERF-6-3A gene function in wheat.

Citrus NIP5;1 aquaporin regulates cell membrane water permeability and alters PIPs plasma membrane localization.

KEY MESSAGE: The ER or donut-like structures localized aquaporin NIP5;1, which interacts with PIPs and alters their localization from plasma membrane to donut-like structures, regulates water permeability. NOD26-like intrinsic proteins (NIPs) play important roles in nutrient uptake and response to various stresses. However, there have been few studies of their functions in water transportation in citrus. Here, we demonstrate the functions of a novel citrus NIP aquaporin (CsNIP5;1) via multiple physiological and biochemical experiments. CsNIP5;1 showed high water permeability when expressed in Xenopus laevis oocytes and yeast. However, subcellular localization assays showed that this protein was localized in the endoplasmic reticulum (ER) or donut-like structures in citrus callus and tobacco leaf. Meanwhile, overexpression of CsNIP5;1 led to a reduction in the water permeability of citrus callus. Protein-protein interaction experiments and subcellular localization assays further revealed that CsNIP5;1 physically interacted with PIPs (CsPIP1;1 and AtPIP2;1), which altered their subcellular localization from the plasma membrane to donut-like structures. Together, CsNIP5;1 was identified as a good water channel when expressed in oocytes and yeast. Meanwhile, CsNIP5;1 participated in the regulation of water permeability of citrus callus, which may be associated with CsNIP5;1-induced re-localization of water channels PIPs. In summary, these results provide new insights into the regulatory mechanism of AQPs-mediated water diffusion.

MicroRNA-26b-5p alleviates murine collagen-induced arthritis by modulating Th17 cell plasticity.

Rheumatoid arthritis (RA) is a chronic autoimmune disease, and the abnormal differentiation of IL-17-producing T helper (Th17) cells is an important factor in the pathogenesis. Previous studies have shown that microRNAs (miRNAs, miR) act as key regulators of Th17 cells. However, the effects of miRNAs on Th17 cell differentiation and plasticity in RA are not clear. In this study, not only low miR-26b-5p expression and high IL-17A level were observed in the peripheral blood of RA patients, but also the negative correlation between miR-26b-5p and IL-17A was explored. The changes in collagen-induced arthritis (CIA) mice were consistent with those in RA patients. The results of in vitro experiments showed that miR-26b-5p mainly inhibited the initial differentiation of Th17 cells but did not impact the differentiation of induced-Treg into Th17-like cells. Meanwhile, miR-26b-5p mimics treatment alleviated inflammatory responses and reduced Th17 proportion in CIA mice. These results indicated that miR-26b-5p could alleviate the development of mice CIA by inhibiting the excessive Th17 cells, and that miR-26b-5p could modulate the plasticity of Th17 cell differentiation in RA, mainly block the initial differentiation. This may provide a novel strategy for the clinical treatment of RA.

A Wheat TaTOE1-B1 Transcript TaTOE1-B1-3 Can Delay the Flowering Time of Transgenic Arabidopsis.

Flowering time is one of the most important agronomic traits in wheat production. A proper flowering time might contribute to the reduction or avoidance of biotic and abiotic stresses, adjust plant architecture, and affect the yield and quality of grain. In this study, TaTOE1-B1 in wheat produced three transcripts (TaTOE1-B1-1, TaTOE1-B1-2, and TaTOE1-B1-3) by alternative splicing. Compared to the longest transcript, TaTOE1-B1-1, TaTOE1-B1-3 has a deletion in the sixth exon (1219-1264 bp). Under long-day conditions, the heterologous overexpression of the TaTOE1-B1-3 gene delayed flowering, prolonged the vegetative growth time, and enlarged the vegetative body of Arabidopsis, but that of TaTOE1-B1-1 did not. As typical AP2 family members, TaTOE1-B1-1 and TaTOE1-B1-3 are mainly located in the nucleus and have transcriptional activation activities; the transcriptional activation region of TaTOE1-B1-3 is located in the C-terminal. In TaTOE1-B1-3 overexpression lines, the expression of flowering-related AtFT and AtSOC1 genes is significantly downregulated. In addition, this study confirms the protein-protein interaction between TaTOE1-B1-3 and TaPIFI, which may play an important role in flowering inhibition. These results provide a theoretical basis for the precise regulation of wheat flowering time.

A NAC transcription factor and its interaction protein hinder abscisic acid biosynthesis by synergistically repressing NCED5 in Citrus reticulata.

Although abscisic acid (ABA) is a vital regulator of fruit ripening and several transcription factors have been reported to regulate ABA biosynthesis, reports of the effect of ABA on citrus ripening and the regulation of its biosynthesis by a multiple-transcription-factor complex are scarce. In the present study, a systematic metabolic, cytological, and transcriptome analysis of an ABA-deficient mutant (MT) of Citrus reticulata cv. Suavissima confirmed the positive effect of ABA on the citrus ripening process. The analysis of transcriptome profiles indicated that CrNAC036 played an important role in the ABA deficiency of the mutant, most likely due to an effect on the expression of 9-cis-epoxycarotenoid dioxygenase 5 (CrNCED5). Electrophoretic mobility shift assays and dual luciferase assays demonstrated that CrNAC036 can directly bind and negatively regulate CrNCED5 expression. Furthermore, yeast two-hybrid, bimolecular fluorescence complementation, and dual luciferase assays demonstrated that CrNAC036 interacted with CrMYB68, also down-regulating the expression of CrNCED5. Taken together, our results suggest that CrNAC036 and CrMYB68 synergistically inhibit ABA biosynthesis in citrus fruit by regulating the expression of CrNCED5.

Baicalein enhances the antitumor efficacy of docetaxel on nonsmall cell lung cancer in a ß-catenin-dependent manner.

The side effects of docetaxel have limited its antitumor performances in the treatment of nonsmall cell lung cancer (NSCLC). To address the problem, baicalein, a bioactive flavone that exhibits antitumor activity, was combined with docetaxel so as to achieve better efficacy and lower toxicity. The combination treatment enhanced the stabilization of microtubules and halted the cell-cycle progression, thus synergistically inhibiting the proliferation and inducing the apoptosis of A549 cells and Lewis lung carcinoma cells. The decreased expression of Cyclin-dependent kinase 6 and Cyclin B1 confirmed its regulation in cell cycle, with ß-catenin being an important upstream effector, as evidenced by the decreased expression in the cytoplasm and nucleus as well as the attenuated aggregation in the nucleus. Furthermore, baicalein plus docetaxel evinced better antitumor efficacy by the suppressed tumor growth, increased apoptosis, and decreased tumor angiogenesis in vivo, with no increased toxicity discovered in both tumor-bearing and non-tumor-bearing mice, and an improvement in therapeutic index. This study has demonstrated that baicalein plus docetaxel is an appropriate combination simultaneously with augmented antitumor efficacy and acceptable safety, which might be a promising strategy for patients with advanced NSCLC.

A d-Peptide Ligand of Integrins for Simultaneously Targeting Angiogenic Blood Vasculature and Glioma Cells.

The current prognosis of glioma patients remains poor after intensive multimodal treatments, which is partially due to the existence of the blood-brain tumor barrier (BBTB). In the present study, a novel "bifunctional ligand" (termed DVS) was developed by retro-inverso isomerization. DVS is a ligand of integrins highly expressed on glioma cells and tumor neovasculature. DVS exhibited exceptional stability in serum and demonstrated significantly higher targeting efficiency for glioma and HUVEC cells compared with the parent L-peptide. As a result, DVS modified micelles (DVS-MS) exhibited high encapsulation efficiency of doxorubicin, ideal size distribution, and sustained release behavior of the payload. In vivo studies showed that DVS-MS could target and efficiently deliver fluorescence to tumor cells and tumor vasculature not only in the mice bearing subcutaneous tumors but also in those bearing intracranial tumors. Moreover, doxorubicin loaded DVS modified micelles exerted potent tumor growth inhibitory activity against subcutaneous and intracranial human glioma in comparison to drug loaded plain micelles and LVS modified micelles. Therefore, DVS appears to be a suitable targeting ligand with potential applications for glioma targeted drug delivery.

Enhanced Glioblastoma Targeting Ability of Carfilzomib Enabled by a DA7R-Modified Lipid Nanodisk.

The robust proliferation of tumors relies on a rich neovasculature for nutrient supplies. Therefore, a basic strategy of tumor targeting therapy should include not only killing regular cancer cells but also blocking tumor neovasculature. D-peptide DA7R, which was previously reported to specifically bind vascular endothelial growth factor receptor 2 (VEGFR2) and neuropilin-1 (NRP-1), could achieve the goal of multitarget recognition. Accordingly, the main purposes of this work were to establish a carfilzomib-loaded lipid nanodisk modified with multifunctional peptide DA7R (DA7R-ND/CFZ) and to evaluate its anti-glioblastoma efficacy in vitro and in vivo. It is testified that the DA7R peptide-conjugated lipid nanodisk can be specifically taken up by U87MG cells and HUVECs. Furthermore, DA7R-ND demonstrated a more enhanced penetration than that of the nonmodified formulation on the tumor spheroid model in vitro and more tumor region accumulation in vivo on the subcutaneous and intracranial tumor-bearing nude mice model. DA7R-ND was shown to co-localize with tumor neovasculature in vivo. When loaded with proteasome inhibitor carfilzomib, the DA7R-decorated nanodisk could remarkably suppress tumor proliferation, extend survival time of nude mice bearing an intracranial tumor, and inhibit neovasculature formation with an efficacy higher than that of the nonmodified nanodisk in vitro and in vivo. The present study verified that the heptapeptide DA7R-conjugated nanodisk is a promising nanocarrier for glioblastoma targeting therapy.