SPA, a Stigma-style-transmitting tract Physical microenvironment Assay for investigating mechano-signaling in pollen tubes.

Accurate sensing and responding to physical microenvironment are crucial for cell function and survival, but the underlying molecular mechanisms remain elusive. Pollen tube (PT) provides a perfect single-cell model for studying mechanobiology since it's naturally subjected to complex mechanical instructions from the pistil during invasive growth. Recent reports have revealed discrepant PT behaviors between in vivo and flat, two-dimensional in vitro cultures. Here, we established the Stigma-style-transmitting tract (TT) Physical microenvironment Assay (SPA) to recapitulate pressure changes in the pistil. This biomimetic assay has enabled us to swiftly identify highly redundant genes, GEF8/9/11/12/13, as new regulators for maintaining PTs integrity during style-to-TT emergence. In contrast to normal growth on solid medium, SPA successfully phenocopied gef8/9/11/12/13 PT in vivo growth-arrest deficiency. Our results suggest the existence of distinct signaling pathways regulating in vivo and in vitro PT integrity maintenance, underscoring the necessity of faithfully mimicking the physical microenvironment for studying plant cell biology.

Polyketides with α-glucosidase inhibitory and neuroprotective activities from Aspergillus versicolor associated with Pedicularis sylvatica.

Aspergillus versicolor, an endophytic fungus associated with the herbal medicine Pedicularis sylvatica, produced four new polyketides, aspeversins A-D (1-2 and 5-6) and four known compounds, O-methylaverufin (2), aversin (3), varilactone A (7) and spirosorbicillinol A (8). Their structures were elucidated by extensive spectroscopic data analysis, and their absolute configurations were determined by calculated electronic circular dichroism (ECD) and Mo2(AcO)4-induced CD data. Compound 5 was found to exhibit α-glucosidase inhibitory activity with an IC50 value of 25.57 µM. An enzyme kinetic study indicated that 5 was a typical uncompetitive inhibitor toward α-glucosidase, which was supported by a molecular docking study. Moreover, compounds 1-3 and 5 also improved the cell viability of PC12 cells on a 1-methyl-4-phenylpyridinium (MPP+)-induced Parkinson's disease model, indicating their neuroprotective potential as antiparkinsonian agents.

Evaluation of Phenazine Derivatives from the Lichen-Associated Streptomyces flavidovirens as Potent Antineuroinflammatory Agents In Vitro and In Vivo.

Eighteen nitrogen-containing compounds (1-18) were isolated from cultures of the lichen-associated Streptomyces flavidovirens collected from the Qinghai-Tibet Plateau, including seven phenazine derivatives with three new ones, named subphenazines A-C (2-4), two new furan pyrrolidones (8-9), and nine known alkaloids. The structures were elucidated by spectroscopic data analysis, and absolute configurations were determined by single-crystal X-ray diffraction and ECD calculations. The phenazine-type derivatives, in particular compound 3, exhibited significantly better antineuroinflammatory activity than other isolated compounds (8-18). Compound 3 inhibited the release of proinflammatory cytokines including IL-6, TNF-α, and PGE2, and the nuclear translocation of NF-κB; it also reduced the oxidative stress and activated the Nrf2 signaling pathway in LPS-induced BV2 microglia cells. In vivo anti-inflammatory activity in zebrafish indicated that 3 inhibited LPS-stimulated ROS generation. These findings suggested that compound 3 might be a potent antineuroinflammatory agent through the regulation of the NF-κB/Nrf2 signaling pathways.

Fungal Coculture of Herpotrichia sp. and Trametes versicolor Induces Production of Diverse Metabolites with Anti-Parkinson's Neuroprotective Activity.

Co-cultivation of isopod-associated fungi Herpotrichia sp. SF09 and Trametes versicolor SF09A led to the reciprocal induction of thirteen new compounds (1-7 and 9-13) with diverse architectures. Importantly, compounds 1 and 2 are rare fungal sesquiterpene-saccharide hybrids incorporating a xylopyranose moiety, compound (±)-3 represents the first example of a natural linear sesquiterpene racemate, and compound 7 is a rare α-pyrone derivative with a xylopyranose motif. Their structures were elucidated by analysis of NMR and mass spectrometry data, and their absolute configurations were determined by Mosher's method, microscale derivatization, and single-crystal X-ray diffraction, as well as ECD calculations. All the isolated compounds ameliorated MPP+-induced oxidative damage in PC12 cells in a dose-dependent fashion. Among them, compounds 5 and 15 showed significant protective action against neuronal injury by MPP+ at 5 µM. Meanwhile, transcriptome sequencing was performed to evaluate the molecular mechanism of the neuroprotective activity for compound 5. Results indicated that compound 5 might mitigate MPP+-induced neuronal injury through the regulation of multiple signaling pathways, including the PI3K-Akt and MAPK pathways. Our findings suggested that compound 5 could be a promising neuroprotective agent for the treatment of Parkinson's disease.

Abietane-Type Diterpenoids from the Arils of Torreya grandis.

A chemical investigation of the arils of Torreya grandis led to the isolation of seven abietane-type diterpenoids (compounds 1-7) including three previously undescribed compounds, one unreported natural product, and three known analogs. The structures of these compounds were determined by means of spectroscopy, single-crystal X-ray diffraction, and ECD spectra. An antibacterial activity assay showed that compounds 5 and 6 had significant inhibitory effects on methicillin-resistant Staphylococcus aureus, with MIC values of 100 µM. Moreover, compounds 1, 3, 4, and 7 exhibited anti-neuroinflammatory activity in LPS-stimulated BV-2 microglia cells, with the IC50 values ranging from 38.4 to 67.9 µM.

Matrix stiffness, endothelial dysfunction and atherosclerosis.

Atherosclerosis (AS) is the leading cause of the human cardiovascular diseases (CVDs). Endothelial dysfunction promotes the monocytes infiltration and inflammation that participate fundamentally in atherogenesis. Endothelial cells (EC) have been recognized as mechanosensitive cells and have different responses to distinct mechanical stimuli. Emerging evidence shows matrix stiffness-mediated EC dysfunction plays a vital role in vascular disease, but the underlying mechanisms are not yet completely understood. This article aims to summarize the effect of matrix stiffness on the pro-atherosclerotic characteristics of EC including morphology, rigidity, biological behavior and function as well as the related mechanical signal. The review also discusses and compares the contribution of matrix stiffness-mediated phagocytosis of macrophages and EC to AS progression. These advances in our understanding of the relationship between matrix stiffness and EC dysfunction open the avenues to improve the prevention and treatment of now-ubiquitous atherosclerotic diseases.

Mechanistic exploration of Yiqi Liangxue Shengji prescription on restenosis after balloon injury by integrating metabolomics with network pharmacology.

CONTEXT: Yiqi Liangxue Shengji prescription (YQLXSJ) is a traditional Chinese medicine (TCM) formula that has long been used for treatment after percutaneous coronary intervention (PCI). OBJECTIVE: To investigate the putative pharmacological mechanism of YQLXSJ on restenosis through an integrated approach utilizing metabolomics and network pharmacology. MATERIALS AND METHODS: Forty male Sprague-Dawley rats were divided into sham, model, YQLXSJ, and positive groups. YQLXSJ group received the treatment of YQLXSJ (6 g/kg/d, i.g.) and the positive group was treated with atorvastatin (2 mg/kg/d, i.g.). After 4 weeks, the improvement in intimal hyperplasia was evaluated by ultrasound, H&E staining, and immunofluorescence. UPLC-MS/MS technology was utilized to screen the differential metabolites. Network pharmacology was conducted using TCMSP, GeneCards, and Metascape, etc., in combination with metabolomics. Eventually, the core targets were acquired and validated. RESULTS: Compared to models, YQLXSJ exhibited decreased intima-media thickness on ultrasound (0.23 ± 0.02 mm vs. 0.20 ± 0.01 mm, p < 0.01) and reduced intima thickness by H&E (30.12 ± 6.05 µm vs. 14.32 ± 1.37 µm, p < 0.01). We identified 18 differential metabolites and 5 core targets such as inducible nitric oxide synthase (NOS2), endothelial nitric oxide synthase (NOS3), vascular endothelial growth factor-A (VEGFA), ornithine decarboxylase-1 (ODC1) and group IIA secretory phospholipase A2 (PLA2G2A). These targets were further confirmed by molecular docking and ELISA. DISCUSSION AND CONCLUSIONS: This study confirms the effects of YQLXSJ on restenosis and reveals some biomarkers. TCM has great potential in the prevention and treatment of restenosis by improving metabolic disorders.

Tricrilactones A-H, Potent Antiosteoporosis Macrolides with Distinctive Ring Skeletons from Trichocladium crispatum, an Alpine Moss-Associated Fungus.

Tricrilactones A-H (1-8), a new family of oligomeric 10-membered macrolides featuring collectively five unique ring skeletons, were isolated from a hitherto unexplored fungus, Trichocladium crispatum. Compounds 1 and 7 contain two unconventional bridged (aza)tricyclic core skeletons, 2, 3, 5, and 6 share an undescribed tetracyclic 9/5/6/6 ring system, 4 bears an uncommon 9/5/6/10/3-fused pentacyclic architecture, and 8 is a dimer bridged by an unexpected C-C linkage. Their structures, including absolute configurations, were elucidated by spectroscopic analysis, quantum chemical calculations, and X-ray diffraction analysis. Importantly, the absolute configuration of the highly flexible side chain of 1 was resolved by the asymmetric synthesis of its four stereoisomers. The intermediate-trapping and isotope labeling experiments facilitated the proposal of the biosynthetic pathway for these macrolides. In addition, their antiosteoporosis effects were evaluated in vivo (zebrafish).

Tumor mutation burden in blood predicts benefit from neoadjuvant chemo/radiotherapy in locally advanced rectal cancer.

Distant metastasis has been the major concern of prognosis in patients with locally advanced rectal cancer (LARC). The purpose of this study was to investigate the prognostic value of TMB in blood (bTMB) in LARC patients after receiving neoadjuvant chemoradiotherapy (nCRT) and surgery. Using targeted ctDNA sequencing, we revealed that bTMB level at baseline was positively correlated with recurrence-free survival (RFS). Following nCRT, the patients with decreasing TMB tends to have a longer median RFS. bTMB level after surgery was negatively correlated with RFS. The serum cytokines including IFNγ, IFNα2, IL-1ß, IL-2 and MIP-1ß were significantly higher in pre-nCRT serum with higher bTMB group than that of lower bTMB group. Clonal evolution analysis showed that the pre- and post-nCRT ctDNAs of most cases had shared mutations. In conclusion, we presume that bTMB could potentially improve pre- and post-treatment risk assessment and facilitate individualized therapy for patients with LARC.

Nanoscale Metal-Organic Layers Detect Mitochondrial Dysregulation and Chemoresistance via Ratiometric Sensing of Glutathione and pH.

Mitochondrial dysregulation controls cell death and survival by changing endogenous molecule concentrations and ion flows across the membrane. Here, we report the design of a triply emissive nanoscale metal-organic layer (nMOL), NA@Zr-BTB/F/R, for sensing mitochondrial dysregulation. Zr-BTB nMOL containing Zr6 secondary building units (SBUs) and 2,4,6-tris(4-carboxyphenyl)aniline (BTB-NH2) ligands was postsynthetically functionalized to afford NA@Zr-BTB/F/R by exchanging formate capping groups on the SBUs with glutathione(GSH)-selective (2E)-1-(2'-naphthyl)-3-(4-carboxyphenyl)-2-propen-1-one (NA) and covalent conjugation of pH-sensitive fluorescein (F) and GSH/pH-independent rhodamine-B (R) to the BTB-NH2 ligands. Cell imaging demonstrated NA@Zr-BTB/F/R as a ratiometric sensor for mitochondrial dysregulation and chemotherapy resistance via GSH and pH sensing.

Nanoscale Coordination Polymers for Combined Chemotherapy and Photodynamic Therapy of Metastatic Cancer.

Combination therapy enhances anticancer efficacy through synergistic effects of different drugs/modalities and can potentially address the challenges in the treatment of metastatic diseases. Here we report the design of carb/pyro nanoscale coordination polymer nanoparticles that carry carboplatin (carb) in the core and the photosensitizer pyrolipid (pyro) on the shell for the treatment of metastatic triple negative breast cancer. Upon light irradiation, carb/pyro generated reactive oxygen species to cause severe cell apoptosis and early calreticulin exposure. Upon intravenous injection and local light irradiation, carb/pyro significantly regressed tumor growth in the 4T1 murine metastatic breast cancer model. When combined with an anti-CD47 antibody, carb/pyro with light irradiation completely eradicated primary and metastatic 4T1 tumors in 50% mice. The anticancer efficacy of carb/pyro was also demonstrated in the CT26 murine colorectal cancer model.

The CD68+ macrophages to CD8+ T-cell ratio is associated with clinical outcomes in hepatitis B virus (HBV)-related hepatocellular carcinoma.

BACKGROUND: Tumor microenvironment plays an essential role during the progression of hepatocellular carcinoma (HCC). Tumor infiltrating immune cells (TILs) was an important component of tumor microenvironment. However, whether TIL features are correlated with the prognosis of HCC patients remains unclear. METHODS: Cancer tissue and paired paracancerous tissues from 220 stage II∼III HBV-related HCC patients were collected. TILs were analyzed using a tyramide signal amplification system combined with immunohistochemistry. Kaplan-Meier survival analysis was conducted to investigate the associations between the prognosis and the infiltrating pattern of TILs. RESULTS: The patients were classified into three distinct subgroups (Clusters (C)1-3) with different overall survival (OS) and disease-free survival (DFS) according to the distribution pattern of TILs. The CD68/CD8 ratio in the cancer SA was correlated with the prognosis. Patients with a higher CD68/CD8 ratio exhibited poorer OS and DFS than those with a lower ratio. The CD68/CD8 ratio in the cancer SA was an independent factor for OS prediction but not DFS. CONCLUSION: CD68+ macrophages and CD8+ T-cells are essential immunological determinants for HBV-related HCC prognosis, and the CD68/CD8 ratio in cancer SA is a novel, prognostic factor for OS prediction in HBV-related HCC patients.

Kanamycin-induced production of 2',3'-cyclic AMP in Escherichia coli.

In contrast to the well-characterized second messenger adenosine 3',5'-cyclic monophosphate (3',5'-cAMP), the biological roles of its isomer 2',3'-cAMP remain largely unknown, especially in bacteria. Recent work reported that RNase I-dependent elevation of 2',3'-cNMP levels in Escherichia coli correlated with reduced biofilm production, and separate studies demonstrated E. coli ribonuclease activation in response to aminoglycoside antibiotics. Here we report that E. coli produced 2',3'-cAMP in response to kanamycin at sub-inhibitory levels. Surprisingly, other aminoglycosides like streptomycin or gentamicin did not generate levels of 2',3'-cAMP detectable by 31P NMR. Interestingly, because 2',3'-cAMP is also produced in E. coli strains expressing a plasmid-encoded kanamycin resistance gene but not by other ribosome-targeting antibiotics, this kanamycin-specific production may not reflect disrupted protein synthesis. Overall, this finding provides a link between aminoglycoside-induced ribonuclease activity and 2',3'-cAMP production in E. coli.

Phaeosphaones: Tyrosinase Inhibitory Thiodiketopiperazines from an Endophytic Phaeosphaeria fuckelii.

Phaeosphaeria fuckelii, an endophytic fungus associated with the herbal medicine Phlomis umbrosa, produced four new thiodiketopiperazine alkaloids, phaeosphaones A-D (1-4), featuring an unusual ß-(oxy)thiotryptophan motif, along with four known analogues, phaeosphaone E (5), chetoseminudin B (6), polanrazine B (7), and leptosin D (8). Their structures were elucidated by extensive spectroscopic data analysis, and their absolute configurations were determined by single-crystal X-ray diffraction and ECD calculations. Compounds 4, 6, and 8 were found to display mushroom tyrosinase inhibitory activity with IC50 values of 33.2 ± 0.2, 31.7 ± 0.2, and 28.4 ± 0.2 µM, respectively, more potent than that of the positive control, kojic acid (IC50 = 40.4 ± 0.1 µM). A molecular-docking study disclosed the π-π stacking interaction between the indole moiety of 8 and the His243 residue of tyrosinase.

Bioremediation of malachite green by cyanobacterium Synechococcus elongatus PCC 7942 engineered with a triphenylmethane reductase gene.

Malachite green is a carcinogenic dye that has been detected in fish tissues and freshwater. Here we evaluated the malachite green decoloring ability of a photoautotrophic cyanobacterium, Synechococcus elongatus PCC 7942 (Synechococcus), that lives in freshwater. Results show that 99.5% of the dye was removed by Synechococcus through bioabsorption and bioaccumulation; however, the dye was not degraded or chemically modified. Next, we established an engineered Synechococcus strain to degrade the dye after uptake. The triphenylmethane reductase gene katmr was heterologously expressed, resulting in high production of a soluble recombinant protein. The engineered strain showed advanced decoloring abilities at a low cell density and in stressful environments. It degraded malachite green into the smaller molecules 4-methylaminobenzoic acid and 4-hydroxyl-aniline. After treatment with the engineered cyanobacterium, the growth of wheat seeds was fully recovered in the presence of malachite green. These results demonstrate the potential application of the engineered Synechococcus as a photosynthetic cell factory for the removal of malachite green from wastewater.

Development of an inoculation technique for rapidly evaluating maize inbred lines for resistance to stalk rot caused by Fusarium spp. in the field.

Maize (Zea mays L.) stalk rot, caused primarily by the soil-borne fungal pathogen Fusarium spp., reduces maize quality and yield worldwide. This study was undertaken to develop and utilize a rapid continuous injection inoculation technique to evaluate maize inbred lines for resistance to Fusarium spp. under field conditions, which could facilitate the identification and development of new sources of host resistance to manage the disease. Continuous injection inoculation is a rapid, stable, and simple method that can evaluate the resistance of maize inbred lines to Fusarium stalk rot (FSR) within 20 days. To verify the feasibility and reliability of inoculation method, Fusarium graminearum, F. proliferatum, and F. subglutinans were isolated, identified, and inoculated into maize at the six-leaf stage (V6) by a veterinary adjustable bottle continuous vaccination syringe. Our results showed that out of a total of 97 inbred maize lines, six (6.2%) showed high resistance to maize stalk rot, 20 showed resistance (20.6%), 32 were susceptible (33.0%), and 39 were very susceptible (40.2%). Based on simple sequence repeat (SSR) markers, an analysis of molecular variance indicated a significant correlation between population of the inbred maize line and resistance to FSR (P = 0.001). Overall, this study provided a systematic, rapid, stable, and simple identification method for maize inbred lines resistant to FSR in the field. At the same time, this method was also suitable for genetic diversity analysis of maize inbred lines resistant to FSR.

Structure-activity relationship of daptomycin analogues with substitution at (2S, 3R) 3-methyl glutamic acid position.

Daptomycin is a highly effective lipopeptide antibiotic against Gram-positive pathogens. The presence of (2S, 3R) 3-methyl glutamic acid (mGlu) in daptomycin has been found to be important to the antibacterial activity. However the role of (2S, 3R) mGlu is yet to be revealed. Herein, we reported the syntheses of three daptomycin analogues with (2S, 3R) mGlu substituted by (2S, 3R) methyl glutamine (mGln), dimethyl glutamic acid and (2S, 3R) ethyl glutamic acid (eGlu), respectively, and their antibacterial activities. The detailed synthesis of dimethyl glutamic acid was also reported.

Photodynamic Therapy Mediated by Nontoxic Core-Shell Nanoparticles Synergizes with Immune Checkpoint Blockade To Elicit Antitumor Immunity and Antimetastatic Effect on Breast Cancer.

An effective, nontoxic, tumor-specific immunotherapy is the ultimate goal in the battle against cancer, especially the metastatic disease. Checkpoint blockade-based immunotherapies have been shown to be extraordinarily effective but benefit only the minority of patients whose tumors have been pre-infiltrated by T cells. Here, we show that Zn-pyrophosphate (ZnP) nanoparticles loaded with the photosensitizer pyrolipid (ZnP@pyro) can kill tumor cells upon irradiation with light directly by inducing apoptosis and/or necrosis and indirectly by disrupting tumor vasculature and increasing tumor immunogenicity. Furthermore, immunogenic ZnP@pyro photodynamic therapy (PDT) treatment sensitizes tumors to checkpoint inhibition mediated by a PD-L1 antibody, not only eradicating the primary 4T1 breast tumor but also significantly preventing metastasis to the lung. The abscopal effects on both 4T1 and TUBO bilateral syngeneic mouse models further demonstrate that ZnP@pyro PDT treatment combined with anti-PD-L1 results in the eradication of light-irradiated primary tumors and the complete inhibition of untreated distant tumors by generating a systemic tumor-specific cytotoxic T cell response. These findings indicate that nanoparticle-mediated PDT can potentiate the systemic efficacy of checkpoint blockade immunotherapies by activating the innate and adaptive immune systems in tumor microenvironment.

Nanoscale Coordination Polymers Codeliver Carboplatin and Gemcitabine for Highly Effective Treatment of Platinum-Resistant Ovarian Cancer.

Due to the ability of ovarian cancer (OCa) to acquire drug resistance, it has been difficult to develop efficient and safe chemotherapy for OCa. Here, we examined the therapeutic use of a new self-assembled core-shell nanoscale coordination polymer nanoparticle (NCP-Carbo/GMP) that delivers high loadings of carboplatin (28.0 ± 2.6 wt %) and gemcitabine monophosphate (8.6 ± 1.5 wt %). A strong synergistic effect was observed between carboplatin and gemcitabine against platinum-resistant OCa cells, SKOV-3 and A2780/CDPP, in vitro. The coadministration of carboplatin and gemcitabine in the NCP led to prolonged blood circulation half-life (11.8 ± 4.8 h) and improved tumor uptake of the drugs (10.2 ± 4.4% ID/g at 24 h), resulting in 71% regression and 80% growth inhibition of SKOV-3 and A2780/CDDP tumors, respectively. Our findings demonstrate that NCP particles provide great potential for the codelivery of multiple chemotherapeutics for treating drug-resistant cancer.

Characterization of CsUGT73AC15 as a Multifunctional Glycosyltransferase Impacting Flavonol Triglycoside Biosynthesis in Tea Plants.

Flavonol glycosides, contributing to the health benefits and distinctive flavors of tea (Camellia sinensis), accumulate predominantly as diglycosides and triglycosides in tea leaves. However, the UDP-glycosyltransferases (UGTs) mediating flavonol multiglycosylation remain largely uncharacterized. In this study, we employed an integrated proteomic and metabolomic strategy to identify and characterize key UGTs involved in flavonol triglycoside biosynthesis. The recombinant rCsUGT75AJ1 exhibited flavonoid 4'-O-glucosyltransferase activity, while rCsUGT75L72 preferentially catalyzed 3-OH glucosylation. Notably, rCsUGT73AC15 displayed substrate promiscuity and regioselectivity, enabling glucosylation of rutin at multiple sites and kaempferol 3-O-rutinoside (K3R) at the 7-OH position. Kinetic analysis revealed rCsUGT73AC15's high affinity for rutin (Km = 9.64 µM). Across cultivars, CsUGT73AC15 expression inversely correlated with rutin levels. Moreover, transient CsUGT73AC15 silencing increased rutin and K3R accumulation while decreasing their respective triglycosides in tea plants. This study offers new mechanistic insights into the key roles of UGTs in regulating flavonol triglycosylation in tea plants.