[Blaps rynchopetera combined with cyclophosphamide affects proliferation and apoptosis of lung cancer cells via Wnt/ß-catenin signaling pathway].

This study aims to investigate the effects of Blaps rynchopetera Fairmaire and/or cyclophosphamide on the proliferation and apoptosis of lung cancer cells and decipher the underlying mechanism. B. rynchopetera and cyclophosphamide-containing serum and blank serum were prepared from SD rats. Cell counting kit-8(CCK-8) assay was employed to examine the proliferation of lung cancer cell lines A549 and Lewis treated with corresponding agents. The Jin's formula method was used to evaluate the combined effect of the two drugs. According to the evaluation results, appropriate drug concentrations and lung cancer cell line were selected for subsequent experiments, which included control, B. rynchopetera, cyclophosphamide, B. rynchopetera + cyclophosphamide, and B. rynchopetera + Wnt/ß-catenin pathway agonist lithium chloride(LiCl) groups. Immunocytochemistry was employed to measure the expression of proliferation-related proteins in Lewis cells after drug interventions. Flow cytometry was employed to determine the cell cycle and apoptosis. The expression levels of proliferating cell nuclear antigen(PCNA), cyclinD1, B-cell lymphoma 2(Bcl-2), Bcl-2-assiocated X protein(Bax), Wnt1, and ß-catenin were determined by Western blot. The results showed that B. rynchopetera and/or cyclophosphamide significantly inhibited the proliferation of A549 and Lewis cells. Compared with B. rynchopetera alone, the combination increased the inhibition rate on cell proliferation. The combination of B. rynchopetera and cyclophosphamide demonstrated a synergistic effect according to Jin's formula-based evaluation. Compared with the control group, the B. rynchopetera, cyclophosphamide, and B. rynchopetera + cyclophosphamide groups showed increased proportion of Lewis cells in G_0/G_1 phase, increased apoptosis rate, up-regulated expression of Bax, and down-regulated expression of PCNA, cyclinD1, Bcl-2, Wnt1, and ß-catenin. Compared with the cyclophosphamide group, the combination group showed increased proportion of cells in G_0/G_1 phase, increased apoptosis rate, up-regulated expression of Bax, and down-regulated expression of PCNA, cyclinD1, Bcl-2, Wnt1, and ß-catenin. Compared with the B. rynchopetera group, the B. rynchopetera + LiCl group had deceased proportion of cells in G_0/G_1 phase, decreased apoptosis rate, down-regulated expression of Bax, and up-regulated expression of PCNA, cyclinD1, Bcl-2, Wnt1, and ß-catenin. The results indicated that B. rynchopetera could inhibit the proliferation, arrest the cell cycle, and induce the apoptosis of lung cancer cells by inhibiting the Wnt/ß-catenin signaling pathway. Moreover, B. rynchopetera had a synergistic effect with cyclophosphamide.

Size Restriction Is Required for Proper Functioning of a Bipartite Begomovirus AC4 Protein.

Many geminiviruses, including members of the genus Begomovirus, produce a protein known as C4 or AC4. Whereas C4/AC4 typically consists of more than 80 amino acid residues, a few are much shorter. The significance of these shorter C4/AC4 proteins in viral infection and why the virus maintains their abbreviated length is not yet understood. The AC4 of the begomovirus Tomato leaf curl Hsinchu virus contains only 65 amino acids, but it extends to 96 amino acids when the natural termination codon is replaced with a normal codon. We discovered that both interrupting and extending AC4 were harmful to tomato leaf curl Hsinchu virus (ToLCHsV). The extended AC4 (EAC4) also showed a reduced ability to promote the infection of the heterologous virus Potato virus X than the wild-type AC4. When the wild-type AC4 was fused with yellow fluorescent protein (AC4-YFP), it was predominantly found in chloroplasts, whereas EAC4-YFP was mainly localized to the cell periphery. These results suggest that ToLCHsV's AC4 protein is important for viral infection, and the virus may benefit from the abbreviated length, because it may lead to chloroplast localization. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

[Blaps rynchopetera affects proliferation, migration, and invasion of non-small cell lung cancer: a study based on network pharmacology and in vivo and in vitro experiments].

Network pharmacology, molecular docking, and in vivo and in vitro experiments were employed to study the molecular mechanism of Blaps rynchopetera Fairmaire in the treatment of non-small cell lung cancer(NSCLC). The components of B. rynchopetera were collected by literature review, and the active components were screened out through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP). PharmMapper was used to obtain the targets of the active components. The targets of NSCLC were obtained from DrugBank, GeneCards, OMIM, TTD, and PharmGKB. The Venn diagram was drawn to identify the common targets shared by the active components of B. rynchopetera and NSCLC. The "drug component-target" network and protein-protein interaction(PPI) network were constructed by Cytoscape, and the key targets were screened by Centiscape. Gene Ontology(GO) annotation and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment of the above key targets were performed by DAVID. AutoDock and PyMOL were used for the molecular docking between the key targets and corresponding active components. A total of 31 active components, 72 potential targets, and 11 key targets of B. rynchopetera against NSCLC were obtained. The active components of B. rynchopetera had good binding activity with key targets. Further, the serum containing B. rynchopetera was prepared and used to culture human lung adenocarcinoma A549 cells. The CCK-8 assay was employed to determine the inhibition rates on the growth of A549 cells in blank control group and those exposed to different concentrations of B. rynchopetera-containing serum, cisplatin, and drug combination(B. rynchopetera-containing serum+cisplatin) for different time periods. The cell migration and invasion of A549 cells were detected by cell scratch assay and Transwell assay, respectively. Western blot was employed to determine the expression levels of B-cell lymphoma-2(Bcl-2), Bcl-2-associated X(Bax), caspase-3, cell division cycle 42(CDC42), proto-oncogene tyrosine-protein kinase SRC, and vascular endothelial growth factor(VEGF) in A549 cells. C57BL/6 mice were inoculated with Lewis cells and randomly assigned into a model control group, a B. rynchopetera group, a cisplatin group, and a drug combination(B. rynchopetera+cisplatin) group, with 12 mice per group. The body weight and the long diameter(a) and short diameter(b) of the tumor were monitored every other day during treatment, and the tumor volume(mm~3) was calculated as 0.52ab~2. After 14 days of continuous medication, the mice were sacrificed for the collection of tumor, spleen, and thymus, and the tumor inhibition rate and immune organ indexes were calculated. The tissue morphology of tumors was observed by hematoxylin-eosin(HE) staining, and the positive expression of Bax, Bcl-2, caspase-3, CDC42, SRC, and VEGF in the tumor tissue was detected by immunohistochemistry. The results indicated that B. rynchopetera and the drug combination regulated the expression levels of Bax, Bcl-2, caspase-3, CDC42, SRC, and VEGF to inhibit the proliferation, migration, and invasion of A549 cells and Lewis cells, thus playing a role in the treatment of NSCLC via multiple ways.

Advances in the Therapeutic Applications of Plant-Derived Exosomes in the Treatment of Inflammatory Diseases.

Plant-derived exosomes (PLDEs) are small extracellular vesicles that encapsulate proteins, nucleic acids and lipids, and they are usually involved in intercellular communication and molecular transport in plants. PLDEs are widely used in the therapy of diseases due to their abundance and easy availability. The diverse roles of PLDEs, which include transportation of drugs, acting as biomarkers for diagnosis of diseases and their roles in different therapies, suggest that there is a need to fully understand all the mechanisms involved in order to provide the optimum conditions for their therapeutic use. This review summarizes the biogenesis, components and functions of PLDEs and focuses on their use as therapeutic agents in the treatment of inflammatory diseases. It also explores new ideas for novel approaches in which PLDEs could potentially help patients with inflammatory diseases in the future.

Astilbin Attenuates Cadmium-Induced Adipose Tissue Damage by Inhibiting NF-κB Pathways and Regulating the Expression of HSPs in Chicken.

Cadmium (Cd) can damage tissues by inducing oxidative stress, lymphocyte infiltration, and inflammation in these sites. Meanwhile, astilbin (Ast) is an antioxidant agent. At present, only a few mechanisms of Cd-induced adipose tissue damage have been described. Herein, we assessed the potential protective effects and the molecular mechanism underlying the antioxidant properly of Ast after Cd intake in chicken adipose tissue. In this study, a total of 160 7-day-old roosters were randomly divided into four groups. Roosters were fed with a basic diet (C group), Ast 40 mg/kg (Ast group), CdCl2 150 mg/kg + Ast 40 mg/kg (Cd/Ast group), and CdCl2 150 mg/kg (Cd group) for 60 days. We found that Cd intake changed the morphology and structure of adipose tissues and decreased the expression of several antioxidants, including total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), catalase (CAT), and total antioxidant capacity (T-AOC), but increased those of oxidative stress markers including malondialdehyde (MDA), inducible nitric oxide synthase (iNOS), NO, and H2O2. Cd further activated the nuclear factor kappa B (NF-κB) signaling pathway and increased the expression of the inflammation-related mediators, interleukin 1beta (IL-1ß), interleukin 6 (IL-6), interleukin 8 (IL-8), interleukin 10 (IL-10), cyclooxygenase-2 (COX-2), iNOS, prostaglandin E synthase (PTGES), tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). Cd-induced oxidative stress upregulated the expression of three heat shock proteins (HSPs), including HSP27, HSP70, and HSP90. Summarily, Cd causes oxidative stress-mediated tissue damage by activating the NF-κB pathway, promoting inflammation and upregulating the expression of HSPs. However, Ast supplementation modulates oxidative stress in adipose tissue by inhibiting inflammation mediated by the NF-κB pathway and regulating the expression of HSPs.

Modified Xiaochaihu Decoction Combined with Mirtazapine in the Treatment of Persistent Depression: A Pilot Randomized Controlled Trial.

Background: Western drugs effectively manage persistent depressive disorder (PDD) but are associated with side effects. Objective: To observe the efficacy and safety of modified Xiaochaihu Decoction combined with mirtazapine in treating PDD. Methods: Patients with PDD were enrolled at the Naval General Hospital (06/2018-02/2019) and randomized to modified Xiaochaihu Decoction and modified Xiaochaihu Decoction with mirtazapine. The self-rating depression scale (SDS) and traditional Chinese medicine (TCM) scale were assessed at baseline and after 12 weeks. The overall clinical efficacy (primary outcome) and adverse reactions were observed. Results: Sixty-four participants completed the trial in the combined and control groups (30 and 28), respectively. In controls, the total effective rate was 78.6%, compared with 96.7% in the combined group (P=0.035). The scores of the SDS and TCM syndrome scale in the two groups were lower after treatment (P < 0.001) but without difference between groups (P=0.077). The combined group showed higher improvement rates regarding insomnia (96.4% vs. 44.0%, P < 0.001), bitter taste (90.5% vs. 52.6%, P=0.007), languid (72.0% vs. 31.8%, P=0.006), and belching/anorexia (100% vs. 52.6%, P < 0.001). The combined group showed a higher frequency of adverse events (73.3% vs. 3.6%) (P < 0.001). Conclusion: Modified Xiaochaihu Decoction combined with mirtazapine effectively treats PDD, and its curative effect is better than that of TCM alone. Trial Registration. This trial was registered with https://www.chictr.org.cn/index.aspx/ChiCTR2100048188.

[Effect of acupuncture for dysphagia after stroke based on fiberoptic endoscopic swallowing function evaluation].

OBJECTIVE: To observe the effect of acupuncture combined with regular treatment and swallowing function training on pharyngeal motor, sensory function and penetration-aspiration function in patients with dysphagia after stroke. METHODS: A total of 60 patients with dysphagia after stroke were randomly divided into a control group and an observation group, 30 patients in each group. Both groups were treated with conventional treatment and swallowing function training; in addition, the observation group was treated with acupuncture at Lianquan (CV 23), Fengfu (GV 16), Yifeng (TE 17). All the treatments were given once a day, 5 days a week, for totally 4 weeks. In the two groups, the pharyngeal motor and sensory function, penetration-aspiration scores were evaluated by fiberoptic endoscopic evaluation of swallowing (FEES), and the Kubota water swallowing test scores were assessed before and after treatment, and the clinical effects were compared. RESULTS: After treatment, the pharyngeal motor and sensory function in the two groups were all higher than those before treatment (P<0.05), and those in the observation group were better than the control group (P<0.05). After treatment, the penetration-aspiration scores and Kubota water swallowing test scores in the two groups were all lower than those before treatment (P<0.05), and those in the observation group were lower than the control group (P<0.05). The total effective rate was 93.3% (28/30) in the observation group, which was better than 73.3% (22/30) in the control group (P<0.05). CONCLUSION: Acupuncture combined with regular treatment and swallowing training could improve the pharyngeal motor and sensory function, and penetration-aspiration scores in patients with dysphagia after stroke.

Protective Effects of Astilbin Against Cadmium-Induced Apoptosis in Chicken Kidneys via Endoplasmic Reticulum Stress Signaling Pathway.

Cadmium (Cd) can cause endoplasmic reticulum stress (ERS) and apoptosis in animals. The kidney is an organ seriously affected by Cd because it can accumulate metal ions. Astilbin (ASB) is a dihydroflavonol rhamnoside, which has an anti-renal injury effect. This study aimed to evaluate the protective effect of ASB on Cd-induced ERS and apoptosis in the chicken kidney. In this study, a total of 120 1-day-old chickens were randomly divided into 4 groups. Chickens were fed with a basic diet (Con group), ASB 40 mg/kg (ASB group), CdCl2 150 mg/kg + ASB 40 mg/kg (ASB/Cd group), and CdCl2 150 mg/kg (Cd group) for 90 days. The results showed that Cd exposure induced pathological and ultrastructural damages and apoptosis in chicken kidneys. Compared with the Con group, metallothionein (MT1/MT2) level, nitric oxide (NO) content, inducible nitric oxide synthase (iNOS) activity, ERS-related genes 78-kDa glucose-regulated protein (Grp78), protein kinase PKR-like endoplasmic reticulum kinase (Perk), activating transcription factor 4 (Atf4) and CAAT/enhancer-binding protein (C/EBP) homologous protein (Chop), and pro-apoptotic gene B-cell lymphoma 2 (Bcl-2)-associated X (Bax), caspase-12, caspase-9, caspase-3 expression levels, and apoptotic rate were significantly increased in the Cd group. The expression level of Bcl-2 was significantly decreased in the Cd group. ASB/Cd combined treatment significantly improves the damage of chicken kidneys by ameliorating Cd-induced kidney ERS and apoptosis. Cd can cause the disorder of the GRP78 signal axis, activate the PERK-ATF4-CHOP pathway, aggravate the structural damage and dysfunction of ER, and promote the apoptosis of chicken kidneys, while the above changes were significantly alleviated in the ASB/Cd group. The results showed that ASB antagonizes the negative effects of Cd and against Cd-induced apoptosis in chicken kidneys via ERS signaling pathway.

Hyperoside Attenuates Zearalenone-induced spleen injury by suppressing oxidative stress and inhibiting apoptosis in mice.

Zearalenone (ZEA) is a ubiquitous mycotoxin contaminant that causes immune toxicity, apoptosis, and oxidative stress in animals. Hyperoside (Hyp) is a flavonol glycoside compound with antioxidant and anti-apoptotic properties. However, the potential of Hyp to prevent ZEA-induced spleen injury remains unknown. To evaluate the chemoprotective effect of Hyp against ZEA-induced spleen injury, 60 male Kunming mice were randomly assigned into five groups. The first two groups were orally treated with ZEA (40 mg/kg) for 30 days, and combined with Hyp (0, 100 mg/kg) treatment. The other three groups are orally treated with normal saline, olive oil, or Hyp (100 mg/kg) for 30 days. Hyperoside had an inhibitory effect against ZEA-induced spleen lesions. In addition, Hyp significantly increased the activity of antioxidant enzymes [superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT)], the total antioxidant capacity (T-AOC), and significantly reduced the malondialdehyde (MDA) content reducing ZEA-induced oxidative stress in the spleen. Moreover, the translation of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream target genes (CAT, NQO1, SOD1, GSS, GCLM, and GCLC) were ameliorated using co-therapy with Hyp before treatment with ZEA. Hyperoside also significantly inhibited the translation and expression of apoptotic genes (caspase3, casepase9, Bax, Bcl-2) and the production of apoptotic bodies induced by ZEA in the spleen. In conclusion, the findings revealed that Hyp inhibited ZEA-induced spleen injury through its antioxidant and anti-apoptotic effects. Thus, it provides a new treatment option for immune system diseases caused by ZEA.

EDLm6APred: ensemble deep learning approach for mRNA m6A site prediction.

BACKGROUND: As a common and abundant RNA methylation modification, N6-methyladenosine (m6A) is widely spread in various species' transcriptomes, and it is closely related to the occurrence and development of various life processes and diseases. Thus, accurate identification of m6A methylation sites has become a hot topic. Most biological methods rely on high-throughput sequencing technology, which places great demands on the sequencing library preparation and data analysis. Thus, various machine learning methods have been proposed to extract various types of features based on sequences, then occupied conventional classifiers, such as SVM, RF, etc., for m6A methylation site identification. However, the identification performance relies heavily on the extracted features, which still need to be improved. RESULTS: This paper mainly studies feature extraction and classification of m6A methylation sites in a natural language processing way, which manages to organically integrate the feature extraction and classification simultaneously, with consideration of upstream and downstream information of m6A sites. One-hot, RNA word embedding, and Word2vec are adopted to depict sites from the perspectives of the base as well as its upstream and downstream sequence. The BiLSTM model, a well-known sequence model, was then constructed to discriminate the sequences with potential m6A sites. Since the above-mentioned three feature extraction methods focus on different perspectives of m6A sites, an ensemble deep learning predictor (EDLm6APred) was finally constructed for m6A site prediction. Experimental results on human and mouse data sets show that EDLm6APred outperforms the other single ones, indicating that base, upstream, and downstream information are all essential for m6A site detection. Compared with the existing m6A methylation site prediction models without genomic features, EDLm6APred obtains 86.6% of the area under receiver operating curve on the human data sets, indicating the effectiveness of sequential modeling on RNA. To maximize user convenience, a webserver was developed as an implementation of EDLm6APred and made publicly available at www.xjtlu.edu.cn/biologicalsciences/EDLm6APred . CONCLUSIONS: Our proposed EDLm6APred method is a reliable predictor for m6A methylation sites.

A layered aluminum-based metal-organic framework as a superior trap for nitrobenzene capture via an intercalation role.

Layered materials with porous layers are of great interest due to their intriguing structural topologies and potential applications as new adsorbents. In this study, a layered aluminum-based metal-organic framework, i.e. Al-TCPP, was successfully synthesized via a facile method for the adsorptive removal of nitrobenzene (NB). The as-synthesized Al-TCPP exhibited a typical layered structure and can be stable in water at pH = 5-7. Batch experimental results showed a superior adsorption performance towards NB with a maximum adsorption capability of 1.85 mg mg-1, and an exceptionally rapid equilibrium within 1 min, yielding an overall adsorptive performance superior to the state-of-the-art NB adsorbents reported so far. The morphology and crystallinity of the Al-TCPP adsorbent basically retain the original status after the capture of NB. Importantly, X-ray diffraction patterns of the samples after the NB adsorption revealed that the possible NB intercalation took place in layered Al-TCPP and expanded the interlayer space during the adsorption, which greatly enriched the adsorption sites and thus achieved the outstanding performance. This work highlights new prospects in designing layered materials for use in environmental remediation.

Clinical characteristics and treatments for bronchial Dieulafoy's disease.

BACKGROUND: Dieulafoy's disease of the bronchus is an arterial abnormality characterized by enlarged mucosal arterial branches that are susceptible to lethal bleeding. To date, this disease is rarely reported in the literature. We recently encountered three patients from February 2010 to March 2017, each with such a vascular anomaly in a bronchus with massive hemoptysis. AIM: This paper describes the clinical characteristics and treatments for Dieulafoy's disease. METHODS: We report three cases with recurrent massive hemoptysis. Bronchoscopic examination was performed on two patients, one with a non-pulsating polypoid nodule and the other without. One patient had fatal bleeding after biopsy and could not withstand bronchial artery embolization or thoracotomy. Angiography and bronchial artery embolization on another two patients successfully stopped the bleeding. In addition, we retrospectively reviewed the literature on all reported cases with cryptogenic hemoptysis, obtained through PubMed and Chinese journal searches. RESULTS: The intervention with embolization was successful, and no new episodes of acute hemoptysis were observed. CONCLUSION: Angiography can be used for diagnosis of Dieulafoy's disease of the bronchus, whereas bronchoscopy biopsy should be avoided. Interventions such as embolization or bronchial coagulation play an important role in patients with coughing with massive hemoptysis.

Evaluation of the Genetic Diversity and Differentiation of Black Locust (Robinia pseudoacacia L.) Based on Genomic and Expressed Sequence Tag-Simple Sequence Repeats.

Understanding the genetic diversity and differentiation of the genetic resources of a species is important for the effective use and protection of forest tree resources. Ex situ development is a common method for the protection of genetic diversity and an essential resource for users who require ready access to a species' germplasm. In this study, we collected seeds of black locust (Robinia pseudoacacia L.) from 19 provenances, covering most of its natural distribution; we randomly selected 367 tender leaves with well-grown and different maternal strains from this group for further analysis. Forty-eight simple sequence repeat (SSR) primers were successfully selected from 91 pairs of SSR primers using native-deformation polyacrylamide gel electrophoresis. In addition, we identified identical genotypes among all individuals and evaluated the quality of the markers. From this, 35 loci were confirmed for analyses of genetic diversity and differentiation of the black locust provenances, which contained 28 expressed sequence tag-derived simple sequence repeats (EST-SSRs) and 7 genomic DNA-derived simple sequence repeats (G-SSRs). We observed high genetic diversity among the native black locust provenances, from which Wright's fixation index and molecular variance suggested that a majority of the genetic differentiation variation could be attributed to within-provenance differences. The genetic distance and identity results indicated that geographic distance was not a dominating factor influencing the distribution of black locust. This is the first study to evaluate provenance genetic variation in native black locust samples using two types of SSR markers, which provides a comprehensive theoretical basis for ex situ conservation and utilization of genetic resources, with an emphasis on breeding applications.

Genetic diversity of begomoviruses in Pakistan captured through a vector based survey.

Begomoviruses (Geminiviridea), transmitted by whiteflies, constitute one of the most dangerous groups of plant viruses posing a severe threat to economically important crops in tropical and sub-tropical areas. In this study, whiteflies were collected from various locations all over Pakistan. The begomoviruses carried by these whiteflies were detected by PCR with the degenerative primers pair AV94/Dep3. Analysis of the 177 sequences obtained in our study, revealed 14 distinct begomovirus species, including five which were not previously reported in this country. Putative novel strains of Corchorus yellow vein virus (CoYVV) and Chilli leaf curl virus (ChiLCV) showing less than 90% identity with the previously available taxa were also identified. The greatest number of begomoviruses per single site was detected in Sindh province, where up to five different begomovirus species were identified from the same cropping field. Moreover, Cotton leaf curl Multan virus - Rajasthan (CLCuMuV-Ra) was found prevalent in all the cotton growing areas. The data reported here may be useful in the development of control measures against begomoviruses.

Visible light crosslinkable human hair keratin hydrogels.

Keratins extracted from human hair have emerged as a promising biomaterial for various biomedical applications, partly due to their wide availability, low cost, minimal immune response, and the potential to engineer autologous tissue constructs. However, the fabrication of keratin-based scaffolds typically relies on limited crosslinking mechanisms, such as via physical interactions or disulfide bond formation, which are time-consuming and result in relatively poor mechanical strength and stability. Here, we report the preparation of photocrosslinkable keratin-polyethylene glycol (PEG) hydrogels via the thiol-norbornene "click" reaction, which can be formed within one minute upon irradiation of visible light. The resulting keratin-PEG hydrogels showed highly tunable mechanical properties of up to 45 kPa in compressive modulus, and long-term stability in buffer solutions and cell culture media. These keratin-based hydrogels were tested as cell culture substrates in both two-dimensional surface seeding and three-dimensional cell encapsulation, demonstrating excellent cytocompatibility to support the attachment, spreading, and proliferation of fibroblast cells. Moreover, the photocrosslinking mechanism makes keratin-based hydrogel suitable for various microfabrication techniques, such as micropatterning and wet spinning, to fabricate cell-laden tissue constructs with different architectures. We believe that the unique features of this photocrosslinkable human hair keratin hydrogel promise new opportunities for their future biomedical applications.

Immune response of interferon-γ-inducible lysosomal thiol reductase (GILT) from Chinese sturgeon (Acipenser sinensis) to microbial invasion and its antioxdative activity in lipopolysaccharides-treated mammalian dentritic cells.

Interferon-γ-inducible lysosomal thiol reductase (GILT) plays an important role in the major histocompatibility complex-restricted antigen processing of endocytosed proteins via catalyzing the disulfide bond reduction in the endocytic pathway. Here, the cDNA of Chinese sturgeon (Acipenser sinensis) GILT (CsGILT) was cloned. It contained an open reading frame of 762 nucleotides encoding a protein of 254 amino acids with an estimated molecular weight of 28.1 kDa. The characteristic structural features, including a signature sequence CQHGX2ECX2NX4C, a CXXC motif, two potential N-glycosylation sites, and eight conserved cysteines were detected in the deduced amino acid sequence of CsGILT. CsGILT was widely expressed in Chinese sturgeon with the highest expression in the spleen, and CsGILT mRNA expression was significantly up-regulated when Chinese sturgeons were challenged with polyinosinic polycytidylic acid or Vibrio anguillarum. The recombinant CsGILT displayed obvious thiol reductase activity demonstrated by catalyzing the reduction of mouse IgG(H+L) by dithiothreitol into heavy chain and light chain. CsGILT also displayed significant antioxidant activity in mouse dentritic cells as indicated by its increasing GSH level and GSH/GSSG ratio, decreasing intracellular reactive oxygen species and nitric oxide levels and lipid peroxidation, as well as enhancing the activities of the antioxidative redox enzymes including catalase and superoxide dismutase. Our results suggested an important role for CsGILT in the immune response in Chinese sturgeon to pathogen invasion possibly via a conserved functional mechanism throughout vertebrate evolution, contributing to our understanding the immune biology and protection of Chinese sturgeon.

MHC class II alpha, beta and MHC class II-associated invariant chains from Chinese sturgeon (Acipenser sinensis) and their response to immune stimulation.

The major histocompatibility complex class II (MHC II) molecules play a vital role in adaptive immune response through presenting antigenic peptides to CD4+ T lymphocytes. To accomplish this physiologic function, the MHC class II-associated invariant chain interacts with the MHC II α/ß subunits and promotes their correct assembly and efficient traffic. Here, we isolated the cDNAs of MHC II α, ß and MHC II-associated invariant chains (designated as CsMHC II α, CsMHC II ß, and CsMHC II γ) from Chinese sturgeon (Acipenser sinensis). The CsMHC II α, ß, and γ mRNAs were widely expressed in Chinese sturgeon, and the highest expression was found in spleen for CsMHC II α and ß chains, while in head kidney for CsMHC II γ chain. Stimulation to Chinese sturgeon with inactivated trivalent bacterial vaccine or polyinosinic polycytidylic acid (poly(I:C)) up-regulated the expressions of CsMHC II α, and ß mRNAs, and their transcripts were overall more quickly up-regulated by poly(I:C) than by bacterial vaccine. Poly(I:C) induced higher CsMHC II γ expression than bacterial vaccine in intestine and spleen, while lower than bacterial vaccine in head kidney and liver. When co-expressed in mouse dendritic cells, the CsMHC II γ chain bound to both the MHC II α and ß chains. Furthermore, the over-expressed CsMHC II γ chain, not CsMHC II α or CsMHC II ß chain, activated NF-κB and STAT3 in mouse dendritic cells, and induced TNF-α and IL-6 expressions as well. This activity was nearly abolished by mutation of the Ser29/Ser34 to Ala29/Ala34 in CsMHC II γ. These results suggested that CsMHC II α, ß, and γ chains might play important role in immune response to pathogen microbial infection of Chinese sturgeon possibly via a conserved functional mechanism throughout vertebrate evolution, which might contribute to our understanding the immune biology of sturgeons.

Structural analysis of photocrosslinkable methacryloyl-modified protein derivatives.

Biochemically modified proteins have attracted significant attention due to their widespread applications as biomaterials. For instance, chemically modified gelatin derivatives have been widely explored to develop hydrogels for tissue engineering and regenerative medicine applications. Among the reported methods, modification of gelatin with methacrylic anhydride (MA) stands out as a convenient and efficient strategy to introduce functional groups and form hydrogels via photopolymerization. Combining light-activation of modified gelatin with soft lithography has enabled the materialization of microfabricated hydrogels. So far, this gelatin derivative has been referred to in the literature as gelatin methacrylate, gelatin methacrylamide, or gelatin methacryloyl, with the same abbreviation of GelMA. Considering the complex composition of gelatin and the presence of different functional groups on the amino acid residues, both hydroxyl groups and amine groups can possibly react with methacrylic anhydride during functionalization of the protein. This can also apply to the modification of other proteins, such as recombinant human tropoelastin to form MA-modified tropoelastin (MeTro). Here, we employed analytical methods to quantitatively determine the amounts of methacrylate and methacrylamide groups in MA-modified gelatin and tropoelastin to better understand the reaction mechanism. By combining two chemical assays with instrumental techniques, such as proton nuclear magnetic resonance (1H NMR) and liquid chromatography tandem-mass spectrometry (LC-MS/MS), our results indicated that while amine groups had higher reactivity than hydroxyl groups and resulted in a majority of methacrylamide groups, modification of proteins by MA could lead to the formation of both methacrylamide and methacrylate groups. It is therefore suggested that the standard terms for GelMA and MeTro should be defined as gelatin methacryloyl and methacryloyl-substituted tropoelastin, respectively, to remain consistent with the widespread abbreviations used in literature.

Vaccination efficacy with marrow mesenchymal stem cell against cancer was enhanced under simulated microgravity.

Stem cell vaccination can induce consistent and strong anti-tumor immunity against cancer in mice model. The antigenic similarity between tumors and embryos has been appreciated for many years and reflects the expression of embryonic gene products by cancer cells and/or cancer-initiating stem cells. Taking advantage of this similarity, we have tested a prophylactic lung cancer vaccine composed of allogeneic murine MSCs. Based on this conception, we first compared their tumor vaccines intervention effects of adult MSCs and MSCs under simulated microgravity (MSC/SMG). In this study, BALB/c mice were vaccinated with MSCs or MSC/SMG, compared with mice vaccinated with phosphate buffered saline (PBS) as negative controls. We then subcutaneously implanted the A549 human lung cancer cell line into vaccinated mice and monitored tumor growth potential in vivo. The smaller tumor size and less tumor weight were observed in mice vaccinated with MSCs or MSC/SMG, compared with that of the Control group. Particularly, it was much more significant in the group of MSC/SMG than that group of the MSCs. Vaccination with SMG treated MSCs inhibited proliferation and promoted apoptosis of tumor tissue. SMG/MSC vaccination induced bothTh1-mediated cytokine response; CD8-dependent cytotoxic response which reduced the proportion of Treg cells. Furthermore, SMG/MSC vaccination significantly increased MHC1 and HSPs proteins expression. In conclusion, we demonstrated the SMG could improve tumor-suppressive activity of MSC. The enhanced anti-tumor immune response of MSCs/SMG was strongly associated with the higher expression of MHC class I molecule on DCs, and the abundance of HSPs in the SMG treated MSCs may make antigens in the MSC more cross-presentable to the host DCs for generating protective antitumor activity. This study gains an insight into the mechanism of MSCs anti-tumor efficacy and gives a new strategy for cancer therapies in the future.

[Non-real-time endobronchial bronchoscopy ultrasound assisted transbronchial lung biopsy in diagnosing peripheral pulmonary lesions].

OBJECTIVE: To evaluate the role of non-real-time endobronchial bronchoscopy ultrasound(EBUS) assisted transbronchial lung biopsy (TBLB) in diagnosing peripheral pulmonary lesions (PPL). METHODS: One hundred and five patients [68 males and 37 females, mean age (59 ± 12) years, ranged from 39 - 81 years] with PPL confirmed by computered tomography (CT) were recruited in this study between June 1st 2011 and March 1st 2012. All cases received bronchoscopy examinations and presented with roughly normal results. Fifty-four cases received EBUS examinations. For peripheral lesions with accessible EBUS images, blind biopsy was performed with biopsy forceps through pathways of the ultrasonic probe after the retreat of the probe. In those cases without accessible EBUS images, blind biopsy was performed based on the localization by image data. The other 51 cases without EBUS testing underwent blind biopsy on the localization by image data. Positive rates of pathological diagnosis of the 2 groups were compared. Analysis was by χ(2)-test. RESULTS: In 54 patients who received EBUS examinations, 76% (41/54) of PPLs were detected performed by EBUS. The positive rate of the EBUS assisted TBLB group was 67% (36/54), compared with 45% (23/51) in the general TBLB group. There was a better diagnostic rate (P < 0.05) in the EBUS assisted TBLB group than the general TBLB group. Thirteen patients without accessible EBUS images obtained negative pathological results. The diagnosis rate of EBUS assisted TBLB on lesions with ≤ 30 mm minimum diameter was 44% (8/18), lower than 78% (28/36) on lesions with > 30 mm minimum diameter (P < 0.05). In terms of diagnosis rate on lesions with ≤ 30 mm minimum diameter, EBUS assisted TBLB was 44% (8/18), higher than 12% (2/17) of TBLB alone (P < 0.05). As for lesions with > 30 mm minimum diameter, diagnosis rate of EBUS assisted TBLB was 52% (28/54) and TBLB alone was 41% (21/51), representing insignificant difference (P > 0.05). In the EBUS assisted TBLB group, we performed 269 blind biopsies, with an average of 4.8 times per case, whereas the general TBLB group required 398 times, with an average of 7.8 times per case. EBUS assisted TBLB decreased the operation times of blind biopsy (P < 0.05) to acquire adequate and appropriate specimen. Complications of biopsy occurred in this study included slight haemoptysis (61/105, 58.1%), chest pain (25/105, 23.8%) and pneumothorax (2/105, 1.9%). Patients with these complications recovered spontaneously without special managements. CONCLUSIONS: Non-real-time EBUS assisted TBLB could improve diagnostic positive rate without increasing operational risk. In most cases, the blind biopsy did not succeed if EBUS failed to detect the lesions. The success rate of non-real-time EBUS assisted TBLB was related to the minimum diameter of PPL. In terms of diagnosis rate on lesions with ≤ 30 mm minimum diameter, EBUS assisted TBLB was higher than TBLB alone. As for lesions with >30mm minimum diameter, there was no significant difference in the diagnosis rate between these 2 groups. EBUS assisted TBLB decreased the times of blind biopsy process (P < 0.05) to obtain adequate and appropriate specimen.