First Report of Leaf Spot Caused by Alternaria alternata on Cyclocodon lancifolius in China.

Cyclocodon lancifolius Bunge in the family Campanulaceae, and commonly known as Hong Guo Ginseng, is found in the Indonesia, Philippines, Vietnam, Japan, and China. The leaves and roots of C. lancifolius are widely used as tonics by ethnic minorities in Guizhou and Hunan Provinces in China. In addition, the fruit is edible, and it is a new resource for both medicine and food. In June 2022, symptoms of leaf spot (Fig 1 A and B.) were observed on C. lancifolius plants in the medicinal plant greenhouse of Guizhou University of Traditional Chinese Medicine (106°61'E, 26°39'N), Guizhou Province. The incidence of leaf spot on C. lancifolius was approximately 40 to 70% of all leaves in canopy. Early symptoms on leaves were small circular or irregular brown spots. As the disease progressed the lesions gradually expanded, and multiple lesions coalesced to form large irregular brown spots. Eventually the seedlings died and leaves of mature plants wilted. In order to isolate the pathogen, ten leaf pieces (5×5 mm) were cut from the junction of the diseased and the healthy tissues, surface sterilized with 75% ethanol for 30 s, 0.1% mercuric chloride (HgCl2) solution for 60 s, rinsed in sterile water three times, finally dried and placed on potato dextrose agar (PDA) and cultured in the dark at 27°C for 4 days. Five purified fungal isolates were obtained by single spore isolation. The colonies were olivaceous to dark olive with white margins and abundant aerial mycelia. On potato carrot agar (PCA) medium, these fungi produced septate conidiophores. Conidia were obclavate or ellipsoid, brown, with one to four transverse septa and one to two longitudinal septa. Spores measured 7.64 to 14.20 × 3.32 to 6.38 µm (n=50). These morphological characteristics are consistent with Alternaria alternata (S. P. Wiltshire. 1933). To further confirm the identification, four genomic DNA regions including the ribosomal DNA internal transcribed spacer (ITS), translation elongation factor 1-a gene (TEF), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), RNA polymerase II second largest subunit (RPB2), and Alternaria major allergen gene (Alt a1) were amplified and sequenced with primers ITS1/ITS4 (White et al. 1990), TEF1-728F/TEF1-986R (Carbone and Kohn 1999), gpd1/gpd2 (Berbee et al. 1999), RPB2-5F/RPB2-7cR (Liu et al. 1999), and Alt-for/Alt-rev (Hong et al. 2005), respectively. Sequences were deposited in GenBank with accession Nos. ITS: OQ128111, OQ690707, and OQ690708; TEF: OQ200380, OQ700996, and OQ700998; GAPDH: OQ200378, OQ700993, and OQ700995; RPB2:OQ200379, OQ701002, and OQ701004; Alt: OQ675614, OQ700999, and OQ701001. In a BLAST search, the sequences were 99-100% identical with corresponding sequences of A. alternata. A maximum likelihood phylogenetic tree was constructed with the combined sequence data sets of ITS, TEF, GAPDH, RPB2, and Alt a1 using MEGA 11. The isolate DHY0, DHY1, and DHY3 clustered with A. alternata (J. H. C. Woudenberg et al. 2015) (Fig. 2). To fulfill Koch's postulates, leaves on three healthy 3-month-old potted C. lancifolius seedlings were wounded with sterile needles and inoculated with 5 mm diameter mycelium, which was covered moist by sterile cotton for 24 h. Sterile water was used as the control. After inoculation, the plants were incubated at 27°C, 85% relative humidity, and a 12 h photoperiod. The experiment was repeated three times. Fifteen days after inoculation, all the leaves showed leaf spot symptoms that were similar to those observed in the greenhouse, while control leaves were asymptomatic (Fig. 1). A. alternata was successfully re-isolated from the symptomatic leaves and identified by morphology and the molecular methods described above. This pathogen has been reported to cause a leaf disease in a wide range of vegetables (Zhang et al. 2021), flowers (Zhang et al. 2022), and medicinal plants (Xing et al. 2020). To the best of our knowledge, this is the first report of A. alternata causing leaf spot on C. lancifolius in China. The accurate identification of this pathogen will provide a basis for the prevention and control of C. lancifolius leaf spot disease in the future.

Effects of rhizosphere fungi on the chemical composition of fruits of the medicinal plant Cinnamomum migao endemic to southwestern China.

BACKGROUND: This study examined how rhizosphere fungi influence the accumulation of chemical components in fruits of a small population species of Cinnamomum migao. RESULTS: Ascomycota and Basidiomycota were dominant in the rhizosphere fungal community of C. migao. Pestalotiopsis and Gibellulopsis were associated with α-Terpineol and sabinene content, and Gibellulopsis was associated with crude fat and carbohydrate content. There were significant differences in rhizosphere fungal populations between watersheds, and there was no obvious change between fruiting periods. Gibberella, Ilyonectria, Micropsalliota, and Geminibasidium promoted sabinene accumulation, and Clitocybula promoted α-Terpineol accumulation. CONCLUSION: The climate-related differentiation of rhizosphere fungal communities in watershed areas is the main driver of the chemical composition of C. migao fruit. The control of the production of biologically active compounds by the rhizosphere fungal community provides new opportunities to increase the industrial and medicinal value of the fruit of C. migao.

Chloroplast genome characteristics and phylogenetic analysis of the medicinal plant Blumea balsamifera (L.) DC.

Blumea balsamifera (L.) DC., a medicinal plant with high economic value in the Asteraceae family, is widely distributed in China and Southeast Asia. However, studies on the population structure or phylogenetic relationships with other related species are rare owing to the lack of genome information. In this study, through high-throughput sequencing, we found that the chloroplast genome of B. balsamifera was 151,170 bp in length, with a pair of inverted repeat regions (IRa and IRb) comprising 24,982 bp, a large single-copy (LSC) region comprising 82,740 bp, and a small single-copy (SSC) region comprising 18,466 bp. A total of 130 genes were identified in the chloroplast genome of B. balsamifera, including 85 protein-coding, 37 transfer RNA, and 8 ribosomal RNA genes; furthermore, sequence analysis identified 53 simple sequence repeats. Whole chloroplast genome comparison indicated that the inverted regions (IR) were more conserved than large single-copy and SSC regions. Phylogenetic analysis showed that B. balsamifera is closely related to Pluchea indica. Conclusively, the chloroplast genome of B. balsamifera was helpful for species identification and analysis of the genetic diversity and evolution in the genus Blumea and family Asteraceae.

Spectrum-effect relationship of antioxidant and anti-inflammatory activities of Laportea bulbifera based on multivariate statistical analysis.

Laportea bulbifera, named Hong He Ma in Chinese, is a Chinese herbal medicine commonly used by the Miao nationality of China. In this study, 43 batches of L. bulbifera were collected from different origins in China. Ethanol, ethyl acetate and petroleum ether were used to prepare different extracts of the plant. UHPLC technique was used to establish the fingerprints, whereas DPPH assay and RAW264.7 inflammatory cell models were used to evaluate the antioxidant and anti-inflammatory activities, respectively. Moreover, the spectrum-effect relationship between relative peak area of common peaks and efficacy value was set up by multivariate statistical analysis. Furthermore, 10 batches were selected randomly for validation of those models. The results showed that ethyl acetate and petroleum ether extracts possess excellent antioxidant and anti-inflammatory activities, respectively. Peaks A6 and A7 demonstrated the greatest antioxidant activity, while peak A17 showed the strongest anti-inflammatory activity. After a verified experiment, the result was obtained and illustrated that the spectrum-effect relationship which we established could reliably infer antioxidant and anti-inflammatory compounds of the Chinese herbal medicine.

[Study on HPLC fingerprint of Sabia parviflora].

This research was performed to establish the HPLC fingerprint of Sabia parviflora. HPLC method was carried out on a Thermo Accucore-C18(4. 6 mm×150 mm,2. 6 µm) column by 30% tetrahydrofuran in methyl alcohol-acetonitrile-0. 1% phosphate solution as mobile phase at a flow rate of 1. 0 m L·min-1,the column temperature was 30 ℃ and the detection wavelength was 360 nm. The fingerprints were further evaluated by chemometrics methods including similarity analysis,hierarchical clustering analysis,and principal component analysis. In HPLC fingerprint,15 common peaks were selected as the common peaks,and 6 contents of them were identified. The similarity degrees of 38 batches of the samples was more than 0. 710,and the samples were divided into 6 clusters by their quality difference. The method was precision,repeatable,stable,simple and reliable,which could be used for quality control and evaluation of S. parviflora.

[Analysis of medicinal resources diversity of 33 pilots (districts) in Guizhou province].

This study is based on the data analysis of medicinal plant resources and diversity collected from the fourth Chinese traditional medicine resource survey( pilot). Through the analysis of relevant data from 33 census pioneer plots in Guizhou province( area),a total of 265 families,1 432 genera and 5 296 species of medicinal resources were reported,including algae,fungi,lichens,mosses,a total of 43 genera and 35 families,57,48 families,120 genera and 453 species of ferns,gymnosperms 11 families,22 genera and 61 species,167 families,1 243 genera and 4 721 species of angiosperms,4 genera and 4 families four medicinal animals.Compared with the data related to the third survey of traditional Chinese medicine resources,the number of ferns,gymnosperms and angiosperms in the fourth survey has increased far more than that of the third survey. From the regional distribution of medicinal resources,the composition of the genus,the type of life,and the location of the medicine,the richness of the medicinal plant resources in Guizhou province is not only reflected in many types,but also in the variety of medicinal resources. These studies provide a scientific basis for vigorously developing the Chinese herbal medicine industry and the sustainably using medicinal plant resources in Guizhou province.

[The role of human lysozyme-like protein 4 in fertilization and its enzymatic properties].

OBJECTIVE: To elucidate the possible role of human lysozyme-like protein 4 (LYZL4) in fertilization and characterize its enzymatic properties. METHODS: The localization of LYZL4 in human spermatozoa was investigated by immunofluorescence staining, the sources of LYZL4 on the sperm surface examined by RT-PCR, and the role of LYZL4 in fertilization assessed by the zona-free hamster egg penetration test. The recombinant plasmid pPIC9K-LYZL4 was constructed and its expression induced with methanol after transformed into competent Pichia pastoris GS115. The recombinant LYZL4 protein (rLYZL4) was purified from the fermentation supernatant and subsequently identified by Western blot. The hyaluronan binding ability of rLYZL4 was determined by ELISA and the muramidase activity, hyaluronidase activity, and free radical scavenging ability examined by spectrophotometric methods. RESULTS: Immunodetection with a specific antiserum localized LYZL4 on the acrosomal membrane of mature spermatozoa, which was exclusively secreted from the testis and epididymis as shown by RT-PCR. Immunoneutralization of LYZL4 significantly decreased the number of human spermatozoa bound to zona-free hamster eggs in a dose-dependent manner in vitro. The recombinant protein was expressed successfully by the P. pastoris strain GS115. Purified rLYZL4 exhibited a potent hyaluronan binding ability and a strong free radical scavenging ability but no muramidase or hyaluronidase activity. CONCLUSIONS: LYZL4 secreted from the testis and epididymis is localized on the acrosomal membrane of mature spermatozoa and plays a role in sperm-egg binding as well as in binding hyaluronan and scavenging free radicals, which suggests that it might be a multi-functional molecule contributive to sperm protection and sperm-egg binding.

Preventing surgical disputes through early detection and intervention: a case control study in China.

BACKGROUND: Medical disputes have become a serious issue in China. A crisis cannot usually be predicted and managed through a cost-benefit strategy; therefore, researchers believe that prevention is better than containment and post-crisis resolution. This study aimed to identify solutions to prevent medical disputes in surgical cases through early warning and intervention of potential cases. METHODS: A case-control study was conducted to identify early detection indicators of medical disputes in the surgical treatment of liver cancer through Delphi consultation and logistic regression on the basis of which interventions were undertaken to prevent potential cases. RESULTS: The dispute detection model was composed of patient age (P = 0.08), frequency of hospitalization (P = 0.003), length of hospital stay (P < 0.001), terminal condition (P = 0.004), unplanned reoperation (P = 0.048), blood transfusion volume (P = 0.006), and arrearage (P < 0.001). Risk management interventions through quality improvement and enhanced communication in cases with an abnormal performance indicator proved effective in practice. CONCLUSIONS: This study explored the use of an evidence-based medical risk management strategy for medical disputes that involved early detection and intervention and could potentially be adopted by hospitals to prevent medical disputes.

A nursing care classification system for assessing workload and determining optimal nurse staffing in a teaching hospital in China: A pre-post intervention study.

We aimed to evaluate the effectiveness of a nursing care classification system for re-assessing nurse workload and determining staffing needs. Adequate bed-nurse ratios help manage hospital cost-efficiency, quality of care and patient safety. A prospective pre-post intervention study was conducted from January 2010 to December 2012 in 16 medical-surgical units of a tertiary teaching hospital. Nursing tasks were classified into four grades of care reflecting actual workload. Units were re-staffed accordingly and bed-nurse ratios compared with government-authorized bed-nurse ratios. Patient satisfaction, hospital stays and mortality were evaluated pre- and poststaffing changes. Average bed-nurse ratio (1:0.41) exceeded the national standard (1:0.40) in 16 units, but was inadequate in five units. Re-staffing increased average bed-nurse ratio from 1:0.41 to 1:0.48. Patients' satisfaction increased from 96.9% to 97.6%, and hospital stays decreased significantly. Nursing care classification effectively distributes nurse staffing to match patients' care levels, improving patient outcomes.

HDGF-related protein-3 is required for anchorage-independent survival and chemoresistance in hepatocellular carcinomas.

OBJECTIVE: Hepatoma-derived growth factor (HDGF)-related proteins (HRPs) comprise a family of six members and are characterised by a conserved HATH domain. Among the family members, HDGF was the first to be identified as a mitogenic factor and shown to play an important role in hepatocellular carcinoma pathogenesis. The aim of the present study is to examine the relevance of HDGF-related protein-3 (HRP-3), another member of the HRP family in hepatocellular carcinoma (HCC). DESIGN: HRP-3 expression in HCC tissues was measured by quantitative reverse transcriptase PCR, western blot and immunohistochemistry analysis. The biological consequences of overexpression and knockdown of HRP-3 in HCC cell lines were studied in vitro and in vivo. RESULTS: Expression of HRP-3 mRNA and protein was shown to be highly upregulated in HCC tissues. While knockdown of HRP-3 by small interference RNAs failed to affect anchorage-dependent growth of HCC cells, it inhibited anchorage-independent growth of HCC cells in vitro and xenograft tumour growth in vivo. Further, knockdown of HRP-3 was shown to sensitise HCC cells to anoikis. Moreover, HRP-3 specifically activated the extracellular-signal-regulated kinase (ERK) pathway without affecting c-Jun N-terminal kinase (JNK), p38, AKT and signal transducer and activator of transcription 3 (STAT3). Importantly, inhibition of the ERK pathway diminished HRP-3-mediated protection of HCC cells from anoikis. Finally, knockdown of HRP-3 was shown to enhance apoptosis of HCC cells induced by multiple chemotherapeutic drugs. CONCLUSION: These findings indicate that HRP-3 plays an essential role in HCC pathogenesis and suggest that it may serve as a novel prognostic marker and molecular target for development of drugs for treatment of HCC.

Development and optimization of a frequency mixing sensor for adjacent samples quantitative detection on a lateral flow assay.

Frequency-mixing technology has been widely used to precisely identify magnetic nanoparticles in applications of quantitative biomedical detection in recent years. Examples include immune adsorption, lateral flow assays (LFAs), and biomagnetic imaging. However, the signals of magnetic response generated by adjacent magnetic samples interfere with each other owing to the small spacing between them in applications involving multi-sample detection (such as the LFA and multiplexing detection). Such signal interference prevents the biosensor from obtaining characteristic peaks related to the concentration of adjacent biomarkers from the magnetic response signals. Mathematical and physical models of the structure of sensors based on frequency-mixing techniques were developed. The theoretical model was verified and its key parameters were optimized by using simulations. A new frequency-mixing magnetic sensor structure was then designed and developed based on the model, and the key technical problem of signal crosstalk between adjacent samples was structurally solved. Finally, standard cards with stable magnetic properties were used to evaluate the performance of the sensor, and strips of the gastrin-17 (G-17) LFA were used to evaluate its potential for use in clinical applications. The results show that the minimum spacing between samples required by the optimized sensor to accurately identify them was only about 4-5 mm, and the minimum detectable concentration of G-17 was 11 pg mL-1 . This is a significant reduction in the required spacing between samples for multiplexing detection. The optimized sensor also has the potential for use in multi-channel synchronous signal acquisition, and can be used to detect synchronous magnetic signals in vivo.

Identification of keystone taxa in rhizosphere microbial communities using different methods and their effects on compounds of the host Cinnamomum migao.

Exploring keystone taxa affecting microbial community stability and host function is crucial for understanding ecosystem functions. However, identifying keystone taxa from humongous microbial communities remains challenging. We collected 344 rhizosphere and bulk soil samples from the endangered plant C. migao for 2 years consecutively. Used high-throughput sequencing 16S rDNA and ITS to obtain the composition of bacterial and fungal communities. We explored keystone taxa and the applicability and limitations of five methods (SPEC-OCCU, Zi-Pi, Subnetwork, Betweenness, and Module), as well as the impact of microbial community domain, time series, and rhizosphere boundary on the identification of keystone taxa in the communities. Our results showed that the five methods, identified abundant keystone taxa in rhizosphere and bulk soil microbial communities. However, the keystone taxa shared by the rhizosphere and bulk soil microbial communities over time decreased rapidly decrease in the five methods. Among five methods on the identification of keystone taxa in the rhizosphere community, Module identified 113 taxa, SPEC-OCCU identified 17 taxa, Betweenness identified 3 taxa, Subnetwork identified 3 taxa, and Zi-Pi identified 4 taxa. The keystone taxa are mainly conditionally rare taxa, and their ecological functions include chemoheterotrophy, aerobic chemoheterotrophy, nitrate reduction, and anaerobic photoautotrophy. The results of the random forest model and structural equation model predict that keystone taxa Mortierella and Ellin6513 may have an effects on the accumulation of 1, 4, 7, - Cycloundecatriene, 1, 5, 9, 9-tetramethyl-, Z, Z, Z-, beta-copaene, bicyclogermacrene, 1,8-Cineole in C. migao fruits, but their effects still need further evidence. Our study evidence an unstable microbial community in the bulk soil, and the definition of microbial boundary and ecologically functional affected the identification of keystone taxa in the community. Subnetwork and Module are more in line with the definition of keystone taxa in microbial ecosystems in terms of maintaining community stability and hosting function.

Ultrasound-Driven Non-Metallic Fenton-Active Center Construction for Extensive Chemodynamic Therapy.

Chemodynamic therapy (CDT) is an emerging tumor microenvironment-responsive cancer therapeutic strategy based on Fenton/Fenton-like reactions. However, the effectiveness of CDT is subject to the slow kinetic rate and non-homogeneous distribution of H2 O2 . In this study, a conceptual non-metallic "Fenton-active" center construction strategy is proposed to enhance CDT efficiency using Bi0.44 Ba0.06 Na0.5 TiO2.97 (BNBT-6) nanocrystals. The separated charge carriers under a piezoelectric-induced electric field synchronize the oxidation of H2 O and reduction of H2 O2 , which consequently increases hydroxyl radical (·OH) yield even under low H2 O2 levels. Moreover, acceptor doping induces electron-rich oxygen vacancies to facilitate the dissociation of H2 O2 and H2 O and further promote ·OH generation. In vitro and in vivo experiments demonstrate that BNBT-6 induces extensive intracellular oxidative stress and enhances cell-killing efficiency by activating necroptosis in addition to the conventional apoptotic pathway. This study proposes a novel design approach for nanomaterials used in CDT and presents a new treatment strategy for apoptosis-resistant tumors.

[Microstructure characteristics and identification significance of leaf epidermis of medicinal plants from Sabia genus].

OBJECTIVE: To study the feasibility and significance of the method to identify medicinal plants through the observation and statistics of 9 species of Sabia medical plants from Guizhou province. METHODS: Leaf epidermis characteristics were observed, measured by optical microscope and analyzed by SPSS 17.0 software. RESULTS: All of these plants had some differences in indumentum, cell morphology and size, and had significant difference in the length and circumference of lower epidermis cells. CONCLUSION: The method combining microscopic observation with statistics can be used as the classification and identification basis of medicinal plants and materials of Sabia genus.

Immunochromatographic enhancement strategy for SARS-CoV-2 detection based on nanotechnology.

The global outbreak of coronavirus disease 2019 (COVID-19) has been catastrophic to both human health and social development. Therefore, developing highly reliable and sensitive point-of-care testing (POCT) for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a priority. Among all available POCTs, the lateral flow immunoassay (LFIA, also known as immunochromatography) has proved to be effective due to its accuracy, portability, convenience, and speed. In areas with a scarcity of laboratory resources and medical personnel, the LFIA provides an affordable option for the diagnosis of COVID-19. This review offers a comprehensive overview of methods for improving the sensitivity of SARS-CoV-2 detection using immunochromatography based on nanotechnology, sorted according to the different detection targets (antigens, antibodies, and nucleic acids). It also looks into the performance and properties of the various sensitivity enhancement strategies, before delving into the remaining challenges in COVID-19 diagnosis through LFIA. Ultimately, it seeks to provide helpful guidance in selecting an appropriate strategy for SARS-CoV-2 immunochromatographic detection based on nanotechnology.

Dual-signal readout paper-based wearable biosensor with a 3D origami structure for multiplexed analyte detection in sweat.

In this research, we design and implement a small, convenient, and noninvasive paper-based microfluidic sweat sensor that can simultaneously detect multiple key biomarkers in human sweat. The origami structure of the chip includes colorimetric and electrochemical sensing regions. Different colorimetric sensing regions are modified with specific chromogenic reagents to selectively identify glucose, lactate, uric acid, and magnesium ions in sweat, as well as the pH value. The regions of electrochemical sensing detect cortisol in sweat by molecular imprinting. The entire chip is composed of hydrophilically and hydrophobically treated filter paper, and 3D microfluidic channels are constructed by using folding paper. The thread-based channels formed after the hydrophilic and hydrophobic modifications are used to control the rate of sweat flow, which in turn can be used to control the sequence of reactions in the differently developing colored regions to ensure that signals of the best color can be captured simultaneously by the colorimetric sensing regions. Finally, the results of on-body experiments verify the reliability of the proposed sweat sensor and its potential for the noninvasive identification of a variety of sweat biomarkers.

An investigation on the respiratory mechanics of mechanically ventilated patients during spontaneous breathing trials with enhanced low-level pressure support ventilation.

INTRODUCTION: Low-level pressure support ventilation (PSV) is most commonly adopted in spontaneous breathing trials (SBTs), and some have proposed setting the positive end-expiratory pressure (PEEP) to 0 cmH2 O in order to shorten the observation time of SBTs. This study aims to investigate the effects of two PSV protocols on the patients' respiratory mechanics. MATERIAL AND METHOD: A prospective randomized self-controlled crossover design was adopted in this study, which involved enrolling 30 difficult-to-wean patients who were admitted to the intensive care unit of the First Affiliated Hospital of Guangzhou Medical University between July 2019 and September 2021. Patients were subjected to the S group (pressure support: 8 cmH2 O, PEEP: 5 cmH2 O) and S1 group (PS: 8 cmH2 O, PEEP: 0 cmH2 O) for 30 min in a random order, and respiratory mechanics indices were dynamically monitored via a four-lumen multi-functional catheter with an integrated gastric tube. Among the 30 enrolled patients, 27 were successfully weaned. RESULT: The S group showed higher airway pressure (Paw), intragastric pressure (Pga) and airway pressure-time product (PTP) than the S1 group. The S group also showed a shorter inspiratory trigger delay, (93.80 ± 47.85) versus (137.33 ± 85.66) ms (P = 0.004); and fewer abnormal triggers, (0.97 ± 2.65) versus (2.67 ± 4.48) (P = 0.042) compared with the S1 group. Stratification based on the causes of mechanical ventilation revealed that under the S1 protocol, patients with chronic obstructive pulmonary disease (COPD) had a longer inspiratory trigger delay compared to both post-thoracic surgery (PTS) patients and patients with acute respiratory distress syndrome. Despite providing greater respiratory support, S group led to significant reductions in inspiratory trigger delay and less abnormal triggers compared to S1 group, especially among patients with chronic obstructive pulmonary disease. CONCLUSION: These findings suggest that the zero PEEP group was more likely to induce a higher number of patient-ventilator asynchronies in difficult-to-wean patients.

Cytotoxic T lymphocyte-dependent tumor growth inhibition by a vascular endothelial growth factor-superantigen conjugate.

T cells are major lymphocytes in the blood and passengers across the tumor vasculature. If these T cells are retained in the tumor site, a therapeutic potential will be gained by turning them into tumor-reactive cytotoxic T lymphocytes (CTLs). A fusion protein composed of human vascular endothelial growth factor (VEGF) and staphylococcal enterotoxin A (SEA) with a D227A mutation strongly repressed the growth of murine solid sarcoma 180 (S180) tumors (control versus VEGF-SEA treated with 15µg, mean tumor weight: 1.128g versus 0.252g, difference=0.876g). CD4(+) and CD8(+) T cells driven by VEGF-SEA were accumulated around VEGFR expressing tumor cells and the induced CTLs could release the tumoricidal cytokines, such as interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha). Meanwhile, intratumoral CTLs secreted cytolytic pore-forming perforin and granzyme B proteins around tumor cells, leading to the death of tumor cells. The labeled fusion proteins were gradually targeted to the tumor site in an imaging mice model. These results show that VEGF-SEA can serve as a tumor targeting agent and sequester active infiltrating CTLs into the tumor site to kill tumor cells, and could therefore be a potential therapeutical drug for a variety of cancers.

Apigenin enhances the cytotoxic effects of tumor necrosis factor-related apoptosis-inducing ligand in human rheumatoid arthritis fibroblast-like synoviocytes.

Activated rheumatoid arthritis (RA) fibroblast-like synoviocytes (RAFLSs) play a central role in both initiating and driving RA. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been documented to induce apoptosis only in a small proportion of RAFLSs, which is followed by an induction of proliferation in surviving cells. Apigenin, a chemopreventive bioflavonoid, exhibits proapoptotic activity in many types of cells. In the present study, we sought to determine whether apigenin could enhance the cytotoxic effect of TRAIL on activated RAFLSs. Human RAFLSs isolated from patients with RA were treated with TRAIL (1 nM), apigenin (20 µM), or their combination, and subjected to apoptosis analysis after a 24-h incubation and proliferation analysis after a 72-h incubation. Apoptosis assay revealed that TRAIL or apigenin alone induced a marked apoptosis in RAFLS and their combination yielded a synergistic increase in RAFLS apoptosis. Immunoblotting analysis of apoptosis regulators demonstrated that combined treatment with apigenin increased caspase-3 expression and activity and decreased the Bcl-2/Bax ratio relative to treatment with TRAIL alone. The presence of apigenin significantly restrained TRAIL-induced RAFLS proliferation, coupled with restoration of the expression of two cell-cycle inhibitors p21 and p27. Moreover, the combination with apigenin blunted TRAIL-induced activation of the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway. Our data collectively demonstrate that apigenin sensitizes RAFLS to TRAIL-induced apoptosis and counteracts TRAIL-dependent RAFLS proliferation, which is likely mediated through inactivation of PI3-K/Akt signaling pathway.

Investigation of the Potential Key Genes and the Multitarget Mechanisms of Polygonum cuspidatum against Heart Failure Based on Network Pharmacology and Experimental Validation.

In this study, systematic pharmacology and bioinformatic approaches were employed to identify the potential targets of Polygonum cuspidatum (PC) for treating heart failure (HF). The active ingredients of PC were screened by using the TCMSP database, and HF-related genes were identified in the GEO database. Then, the herb-HF targeted-gene networks were constructed using Cytoscape software. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional analyses were performed to obtain the enriched molecular pathways associated with the pathogenesis of HF. Finally, in vitro experiment was performed to evidence network pharmacology analysis. 170 intersection genes were obtained, and key genes (FOXO3, NFKB1, and TNF) were identified. Besides, GO and KEGG findings indicated that PC treatment of HF was achieved via regulating apoptosis, IL-17 signaling pathway, TNF signaling pathway, response to oxidative stress, and response to reactive oxygen species. And cell experiment revealed that PC could decrease the expression of NFKB1 and TNF and increase the expression of FOXO3, SOD1, and GPX1 in H9C2 cells. These findings showed that the therapeutic mechanism of PC in the treatment of HF may be associated with the regulation of inflammation-related and oxidative stress-related genes.