Editing of the ethylene biosynthesis gene in carnation using CRISPR-Cas9 ribonucleoprotein complex.

The study aimed to edit ethylene (ET) biosynthesis genes [1-aminocyclopropane-1-carboxylic acid (ACC) synthetase 1 (ACS1) and ACC oxidase 1 (ACO1)] in carnation using the CRISPR/Cas9 ribonucleoprotein (RNP) complex system. Initially, the conserved regions of the target genes (ACS1 and ACO1) were validated for the generation of different single guide RNAs (sgRNAs), followed by the use of an in vitro cleavage assay to confirm the ability of the sgRNAs to cleave the target genes specifically. The in vitro cleavage assay revealed that the sgRNAs were highly effective in cleaving their respective target regions. The complex of sgRNA: Cas9 was directly delivered into the carnation protoplast, and the target genes in the protoplast were deep-sequenced. The results revealed that the sgRNAs were applicable for editing the ET biosynthesis genes, as the mutation frequency ranged from 8.8 to 10.8% for ACO1 and 0.2-58.5% for ACS1. When sequencing the target genes in the callus derived from the protoplasts transformed with sgRNA: Cas9, different indel patterns (+ 1, - 1, and - 8 bp) in ACO1 and (- 1, + 1, and + 11) in ACS1 were identified. This study highlighted the potential application of CRISPR/Cas9 RNP complex system in facilitating precise gene editing for ET biosynthesis in carnation.

Macrophages-derived exo-miR-4449 induced by Cryptococcus affects HUVEC permeability and promotes pyroptosis in BEAS-2B via the HIC1 pathway.

Macrophages have recently been discovered to assume a significant role in the progression of cryptococcosis. However, the potential involvement of macrophage-derived exosomes in the pathogenesis of cryptococcosis remains uncertain. In this study, we investigated the changes of microRNAs in macrophage exosomes (exo-miRNAs) in cryptococcal infections and the role of markedly altered exo-miRNAs in the modulation of Human Umbilical Vein Endothelial Cells (HUVEC) permeability and ROS accumulation and pyroptosis in Human Bronchial Epithelioid Cells (BEAS-2B). Techniques such as microarray analysis and real-time quantitative PCR were used to detect different exo-miRNAs and to screen for the most highly expressed exo-miRNAs. Then its mimics were transfected into HUVEC to study its effect on the monolayer permeability of HUVEC. Finally, the relationship between this exo-miRNAs and the ROS accumulation and pyroptosis was verified by bioinformatics analysis. The results showed that five exo-miRNAs were overexpressed and two exo-miRNAs were reduced, among which, exo-miR-4449 was expressed at the highest level. Exo-miR-4449 could be internalized by HUVEC and enhanced its monolayer permeability. Moreover, exo-miR-4449 was found to promote ROS accumulation and pyroptosis in BEAS-2B through HIC1 pathway. Thus, exo-miR-4449 plays an important role in the pathogenesis of cryptococcosis and holds promise as a significant biomarker for treatment.

A Study and Analysis of the Relationship between Visual-Auditory Logos and Consumer Behavior.

Given enterprises' participation in market competition and the development of sensory marketing, in addition to the traditional visual identity, some enterprises gradually begin to pay attention to auditory and then introduce sound design when designing logos. Audio-visual stimulation and media innovation are committed to creating positive attitudes among consumers. This study constructs a model of visual and auditory interactive relationships with consumer behavior using the SOR model. It tests the conceptual model and checks the hypotheses proposed in the study. This study summarizes and contributes to the visual and auditory interactive relationship between information integration, information synergy, mutual competition, and matching degree. It further proposes the influence of purchase intention and consumer support on consumer behavior of perceived brand perception, credibility, and quality perception. The results and highlights ensure brand identities reflect a significant positive result through consumer behavior. In this paper, we collected questionnaires from a random sample of 1407 respondents. We used regression analysis to test the association between visual and auditory interactive relationships as well as consumer behavior. We further verified the mediating role of consumer perception variables. Adding audiovisual logo design to the marketing process can be an effective way for companies and brands to attract customers and increase their support and purchase intentions.

Risk factors for the recurrence in pulmonary tuberculosis patients with massive hemoptysis.

OBJECTIVES: To evaluate the outcomes of bronchial artery embolization (BAE) for the treatment of massive hemoptysis in patients with pulmonary tuberculosis and identify risk factors that influence recurrence. METHODS: A total of 81 patients with massive hemoptysis who underwent BAE between January 2014 and December 2017 were retrospectively reviewed. All of the patients had either a history of pulmonary tuberculosis or a current diagnosis of pulmonary tuberculosis. Follow-up ranged from 18 to 66 months. RESULTS: Hemoptysis was stopped or markedly decreased, with subsequent clinical improvement in 73 patients, while 11 patients experienced recurrence during the follow-up period. Systemic-pulmonary shunts and clinical failure showed a statistically significant correlation with the recurrence rate. The cumulative non-recurrence rate was 95.3% for 3 months and 81.9% for more than 24 months. Complications were common (12.5%), but self-limiting. CONCLUSIONS: BAE is a safe and effective treatment option for the control of massive hemoptysis in pulmonary tuberculosis patients. Systemic-pulmonary shunts and clinical failure are the risk factors for recurrence.

Promotion of the resistance of human oral epithelial cells to herpes simplex virus type I infection via N6-methyladenosine modification.

OBJECTIVE: This study aimed to explore the mechanism behind N6-methyladenosine (m6A) modification of the total ribonucleic acid (RNA) involved in the resistance to herpes simplex virus type I (HSV-1) infection in oral epithelial cells. METHOD: The variation in m6A modification level on messenger RNA following HSV-1 infection was determined using the RNA dot blot method. The expression levels of alpha-ketoglutarate-dependent dioxygenase lab homolog 5 (ALKBH5) protein and fatty mass and obesity-associated genes (FTO) were determined using real-time fluorescence quantification polymerase chain reaction and the western blot technique, respectively. Next, after suppressing the expression of ALKBH5 or FTO via small interfering RNA, human immortalised oral epithelial cells (HIOECs) were infected with HSV-1, followed by measurement of the viral load or expression level of type I interferon (I-IFN) and interferon-stimulated genes (ISGs). RESULTS: The m6A modification level was significantly increased following HSV-1 infection of the HIOECs (P < 0.05), while the expression of ALKBH5 and FTO genes was reduced (P < 0.01). Moreover, the suppression of ALKBH5 or FTO increased the production of I-IFN and ISGs during the HSV-1 infection of the HIOECs (P < 0.01), and the viral load was significantly reduced (P < 0.01). CONCLUSION: During oral HSV-1 infection, the m6A level was increased through the down-regulation of ALBHK5 and FTO expression, increasing I-IFN production and the promotion of HSV-1 clearing in HIOECs.

A-Kinase Anchor Protein 95 Is Involved in ERK1/2-Elk-1 Signal Transduction in Colon Cancer.

Objectives: To assess A-kinase anchor protein 95 (AKAP95), B-Raf, extracellular regulated protein kinases 1/2 (ERK1/2), and Elk-1 expression in colon cancer tissue, and characterize AKAP95 associations with B-Raf, ERK1/2, Elk-1, and colon cancer clinicopathological indices. Methods: The immunohistochemistry streptavidin-perosidase (SP) method was used to determine protein expression levels in 64 colon cancer and 32 para-carcinoma tissue specimens. Results: (1) Positive AKAP95 expression rates in colon cancer tissue were higher when compared with para-carcinoma tissue (92.19% vs. 59.38%, P < 0.05). Similar findings were determined for B-Raf (76.56% vs. 25%, P < 0.05), ERK1/2 (90.63% vs. 31.25%, P < 0.05), and Elk-1 levels (92.19% vs. 40.63%, P < 0.05). (2) No significant associations were identified between AKAP95, B-Raf, ERK1/2, and Elk-1 protein expression and degree of differentiation, histological type, and lymph node metastasis in colon cancer samples (P > 0.05); however, in The Cancer Genome Atlas and Gene Expression Omnibus datasets, AKAP95 was closely related to immune infiltration, and highly expressed AKAP95 was negatively associated with overall survival and relapse free survival rates in colon cancer patients. (3) Correlations were observed between AKAP95 and ERK1/2, AKAP95 and Elk-1, B-Raf and ERK1/2, B-Raf and Elk-1, and ERK1/2 and Elk-1 (all P < 0.05), but no correlation was observed between AKAP95 and B-Raf (P > 0.05). Conclusions: AKAP95 may affect immune infiltration levels in colon cancer by participating in ERK1/2-Elk-1 signal transduction.

BMSCs mediates endothelial cell autophagy by upregulating miR-155-5p to alleviate ventilator-induced lung injury.

In this study, we explored to detect the effects and mechanism of bone-marrow-derived mesenchymal stem cells (BMSCs) on ventilator-induced lung injury (VILI). We transplanted BMSCs in mice and then induced VILI using mechanical ventilation (MV) treatment. The pathological changes, the content of PaO2 and PaCO2 , wet/dry weight ratio (W/D) of the lung, levels of tumor necrosis factor-α and interleukin-6 in bronchoalveolar lavage fluid, and apoptosis were detected. The autophagy-associated factor p62, LC3, and Beclin-1 expression were analyzed by western blot. The quantitative polymerase chain reaction was applied to detect abnormally expressed microRNAs, including miR-155-5p. Subsequently, we overexpressed miR-155-5p in VILI mice to detect the effects of miR-155-5p on MV-induced lung injury. Then, we carried out bioinformatics analysis to verify the BMSCs-regulated miR-155-5p that target messenger RNA. It was observed that BMSCs transplantation mitigated the severity of VILI in mice. BMSCs transplantation reduced lung inflammation, strengthened the arterial oxygen partial pressure, and reduced apoptosis and the W/D of the lung. BMSCs promoted autophagy of pulmonary endothelial cells accompanied by decreased p62 and increased LC3 II/I and Beclin-1. BMSCs increased the levels of miR-155-5p in VILI mice. Overexpression of miR-155-5p alleviated lung injury in VILI mice following reduced apoptosis and increased autophagy. Finally, TAB2 was identified as a downstream target of miR-155-5p and regulated by miR-155-5p. BMSCs may protect lung tissues from MV-induced injury, inhibit lung inflammation, promote autophagy through upregulating of miR-155-5p.

Role of Ethylene Biosynthesis Genes in the Regulation of Salt Stress and Drought Stress Tolerance in Petunia.

Ethylene plays a critical signaling role in the abiotic stress tolerance mechanism. However, the role of ethylene in regulating abiotic stress tolerance in petunia has not been well-investigated, and the underlying molecular mechanism by which ethylene regulates abiotic stress tolerance is still unknown. Therefore, we examined the involvement of ethylene in salt and drought stress tolerance of petunia using the petunia wild type cv. "Merage Rose" and the ethylene biosynthesis genes (PhACO1 and PhACO3)-edited mutants (phaco1 and phaco3). Here, we discovered that editing PhACO1 and PhACO3 reduced ethylene production in the mutants, and mutants were more sensitive to salt and drought stress than the wild type (WT). This was proven by the better outcomes of plant growth and physiological parameters and ion homeostasis in WT over the mutants. Molecular analysis revealed that the expression levels of the genes associated with antioxidant, proline synthesis, ABA synthesis and signaling, and ethylene signaling differed significantly between the WT and mutants, indicating the role of ethylene in the transcriptional regulation of the genes associated with abiotic stress tolerance. This study highlights the involvement of ethylene in abiotic stress adaptation and provides a physiological and molecular understanding of the role of ethylene in abiotic stress response in petunia. Furthermore, the finding alerts researchers to consider the negative effects of ethylene reduction on abiotic stress tolerance when editing the ethylene biosynthesis genes to improve the postharvest quality of horticultural crops.

Down-regulation of microRNA-155 suppressed Candida albicans induced acute lung injury by activating SOCS1 and inhibiting inflammation response.

Acute lung injury caused by Candida albicans could result in high mortality and morbidity. MicroRNA-155 (miR-155) and suppressor of cytokine signaling 1 (SOCS1) have been believed to play a key in the regulation of inflammatory response. Whether miR-155/SOCS1 axis could regulate the acute lung injury caused by C. albicans has not been reported. The acute lung injury animal model was established with acute infection of C. albicans. miR-155 inhibitor, miR-155 mimic, and sh-SOCS1 were constructed. The binding site between miR-155 and SOCS1 was identified with dual luciferase reporter assay. Knockdown of miR-155 markedly inhibited the germ tube formation of C. albicans. Knockdown of miR-155 significantly up-regulated the expression of SOCS1, and the binding site between miR-155 and SOCS1 was identified. Knockdown of miR-155 improved the acute lung injury, suppressed inflammatory factors and fungus loading through SOCS1. Knockdown of SOCS1 greatly reversed the influence of miR-155 inhibitor on the cell apoptosis in vitro. The improvement of acute lung injury caused by C. albicans, suppression of inflammatory response and C. albicans infection, and inhibitor of cell apoptosis were achieved by knocking down miR-155 through SOCS1. This research might provide a new thought for the prevention and treatment of acute lung injury caused by C. albicans through targeting miR-155/SOCS1 axis.

Inhibition of Candida albicans in vivo and in vitro by antimicrobial peptides chromogranin A-N12 through microRNA-155/suppressor of cytokine signaling 1 axis.

Antimicrobial peptides (AMPs) have proven to inhibit a variety of pathogens. Chromogranin A-N12 (CGA-N12) is a kind of AMP, and it is characterized by stable structure, high anti-Candida activity, and good safety. However, it remains unclear whether CGA-N12 could effectively inhibit the growth of Candida albicans (C. albicans). Colony forming assays were used to measure minimal inhibitory concentration (MIC), minimal fungicidal concentration (MFC), and time-kill curve. Disseminated C. albicans rabbit model was established to investigate the influence of CGA-N12 on histological damage. The protein and mRNA levels of suppressor of cytokine signaling 1 (SOCS1) after treatment were investigated. The MIC and MFC of CGA-N12 against C. albicans was 6 mg/mL. CGA-N12 considerably inhibited germ tube formation of C. albicans. The fungal load in the tissues and inflammatory factors in the serum were suppressed by CGA-N12. CGA-N12 significantly reduced the histological changes caused by C. albicans, and the protein and mRNA levels of SOCS1 were markedly inhibited. The inhibition effect of CGA-N12 on C. albicans and significant improvement of histological damage by CGA-N12 through microRNA-155/SOCS1 axis were proved in this study. This study proposes a novel therapeutic strategy for the treatment and prevention of C. albicans.Abbreviations: AMPs: Antimicrobial peptides; MIC: Minimal inhibitory concentration; MFC: Minimal fungicidal concentration; AIDS: Acquired immune deficiency syndrome; PBS: Phosphate buffer saline; FBS: Fetal bovine serum; ROS: Reactive oxygen species; CFU: Colony formation unit; CGA: Chromogranin A; SOCS1: Suppressor of cytokine signaling 1; SDA: Sabouraud Dextrose Agar; GRAVY: Grand average of hydropathicity; C. parapsilosis: Candida parapsilosis; C. albicans: Candida albicans.

Responses of leaf hydraulic traits of Schoenoplectus tabernaemontani to increasing temperature and CO2 concentrations.

BACKGROUND: Against the background of a changing climate, the responses of functional traits of plateau wetland plants to increasing temperatures and CO2 concentrations need to be understood. Hydraulic traits are the key for plants to maintain their ecological functions and affect their growth and survival. However, few studies have comprehensively considered the response strategies of wetland plants' hydraulic traits to environmental changes in the context of water and matter transport, loss, and retention. According to the latest IPCC prediction results, we performed experiments under increased temperature (2 °C) and CO2 levels (850 µmol/mol) in an artificial Sealed-top Chamber (STC) to investigate the responses of the hydraulic characteristics of Schoenoplectus tabernaemontani, the dominant species in plateau wetlands in China. RESULTS: Compared with the CK group, net photosynthetic rate, transpiration rate, stomatal length, cuticle thickness, vascular bundle length, vascular bundle width, and vascular bundle area of S. tabernaemontani in the ET group were significantly reduced, whereas stomatal density and vein density increased significantly. Compared with the CK group, the hydraulic traits of S. tabernaemontani in the EC group were reduced considerably in stomatal length and cuticle thickness but increased dramatically in stomatal density, and there were no significant differences between other parameter values and the control group. Net photosynthetic rate was significantly positively correlated with stomatal length, cuticle thickness, and vascular bundle length, and stomatal conductance was significantly positively correlated with cuticle thickness. The transpiration rate was significantly positively correlated with cuticle thickness, epidermal cell area, vascular bundle length, vascular bundle width, and vascular bundle area. Regarding the hydraulic traits, there was a significant negative correlation between stomatal density and stomatal length, or cuticle thickness, and a significant positive correlation between the latter two. The epidermal cell area was significantly positively correlated with epidermal thickness, vascular bundle length, vascular bundle width, and vascular bundle area. CONCLUSIONS: Increased temperature and CO2 levels are not conducive to the photosynthetic activity of S. tabernaemontani. Photosynthetic rate, stomatal density and size, vein density, epidermal structure size, and vascular bundle size play an essential role in the adaptation of this species to changes in temperature and CO2 concentration. In the process of adaptation, hydraulic traits are not isolated from each other, and there is a functional association among traits. This study provide a scientific basis for the management and protection of plateau wetlands.

Editing of 1-aminocyclopropane-1-carboxylate oxidase genes negatively affects petunia seed germination.

KEY MESSAGE: Editing of ACO genes involved in ethylene biosynthesis pathway reduces ethylene production in petunia seeds and inhibits seed germination. Ethylene production in the seeds of Petunia hybrida cv. 'Mirage Rose' was associated with expression of 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase (ACO) genes (PhACO1, PhACO3, and PhACO4). Suppression of their expression by ethylene inhibitor silver thiosulphate (STS) significantly reduced ethylene production and inhibited seed germination. When it was combined with ethylene precursor ACC, ethylene production was re-promoted via activation of the genes and higher seed germination was restored. This was confirmed using the mutants editing the genes and WT. In the present study, compared with wild type plants, three different mutants (phaco1, phaco3, and phaco4) showed significantly decreased germination percentages as well as delayed germination time and seedling growth. These reductions were associated with lighter seed weight, lower ACO transcript levels, and lower ethylene production in mutants. Inhibited seed germination owing to reduced ethylene production was further verified by the supplementation of exogenous ACC and gibberellic acid (GA3) to growth medium, which restored high seed germination activity in all mutants via enhanced ethylene production. In this study, we reported a key regulatory role of ethylene in seed germination mechanisms in petunia. Further, we highlighted on need to consider the negative effects of ethylene reduction in seed germination and plant growth when editing genes in the ethylene biosynthesis pathway for the maintenance of postharvest fruit, vegetable, and flower quality.

Transcriptomic Analysis for the Identification of Metabolic Pathway Genes Related to Toluene Response in Ardisia pusilla.

Toluene is an industrial raw material and solvent that can be found abundantly in our daily life products. The amount of toluene vapor is one of the most important measurements for evaluating air quality. The evaluation of toluene scavenging ability of different plants has been reported, but the mechanism of plant response to toluene is only partially understood. In this study, we performed RNA sequencing (RNA-seq) analysis to detect differential gene expression in toluene-treated and untreated leaves of Ardisiapusilla. A total of 88,444 unigenes were identified by RNA-seq analysis, of which 49,623 were successfully annotated and 4101 were differentially expressed. Gene ontology analysis revealed several subcategories of genes related to toluene response, including cell part, cellular process, organelle, and metabolic processes. We mapped the main metabolic pathways of genes related to toluene response and found that the differentially expressed genes were mainly involved in glycolysis/gluconeogenesis, starch and sucrose metabolism, glycerophospholipid metabolism, carotenoid biosynthesis, phenylpropanoid biosynthesis, and flavonoid biosynthesis. In addition, 53 transcription factors belonging to 13 transcription factor families were identified. We verified 10 differentially expressed genes related to metabolic pathways using quantitative real-time PCR and found that the results of RNA-seq were positively correlated with them, indicating that the transcriptome data were reliable. This study provides insights into the metabolic pathways involved in toluene response in plants.

CRISPR/Cas9-mediated editing of 1-aminocyclopropane-1-carboxylate oxidase1 enhances Petunia flower longevity.

The genes that encode the ethylene biosynthesis enzyme 1-aminocyclopropane-1-carboxylate oxidase (ACO) are thought to be involved in flower senescence. Hence, we investigated whether the transcript levels of PhACO genes (PhACO1, PhACO3 and PhACO4) in Petunia cv. Mirage Rose are associated with ethylene production at different flowering stages. High transcript levels were detected in the late flowering stage and linked to high ethylene levels. PhACO1 was subsequently edited using the CRISPR/Cas9 system, and its role in ethylene production was investigated. PhACO1-edited T0 mutant lines, regardless of mutant type (homozygous or monoallelic), exhibited significantly reduced ethylene production and enhanced flower longevity compared with wild-type. Flower longevity and the reduction in ethylene production were observed to be stronger in homozygous plants than in their monoallelic counterparts. Additionally, the transmission of the edited gene to the T1 (lines 6 and 36) generation was also confirmed, with the results for flower longevity and ethylene production proving to be identical to those of the T0 mutant lines. Overall, this study increases the understanding of the role of PhACO1 in petunia flower longevity and also points to the CRISPR/Cas9 system being a powerful tool in the improvement of floricultural quality.

Structural characteristics of humic-like acid from microbial utilization of lignin involving different mineral types.

This paper determines the impact of two clay minerals (kaolinite and montmorillonite) and three oxides (goethite, δ-MnO2, and bayerite) on the elemental composition and FTIR spectra of humic-like acid (HLA) extracted from microbial-mineral residue formed from the microbial utilization of lignin in liquid shake flask cultivation. Goethite, bayerite, and δ-MnO2 showed higher enrichment capabilities of C and O + S in the HLA than kaolinite and montmorillonite. Goethite showed the highest retention of organic C, followed by bayerite, but kaolinite exhibited the least exchangeability. Kaolinite and montmorillonite enhanced microbial consumption of N, resulting in the absence of N in HLA. A few aliphatic fractions were preferentially gathered on the surfaces of kaolinite and montmorillonite, making the H/C ratios of HLA from the clay mineral treatments higher than those of HLA from the oxide treatments. δ-MnO2 was considered the most effective catalyst for abiotic humification, and goethite and bayerite ranked second and third in this regard. This trend was proportional to their specific surface areas (SSAs). However, comparing the effects of different treatments on the promotion of HLA condensation by relying solely on the SSA of minerals was not sufficient, and other influencing mechanisms had to be considered as well. Additionally, Si-O-Al and Si-O of kaolinite participated in HLA formation, and Si-OH, Si-O, and Si-O-Al of montmorillonite also contributed to this biological process. Fe-O and phenolic -OH of goethite, Mn-O of δ-MnO2, and Al-O of bayerite were all involved in HLA formation through ligand exchange and cation bridges. Lignin was better protected from microbial decomposition by the kaolinite, bayerite, and δ-MnO2 treatments, which caused lignin-like humus (HS) formation. Under the treatments of δ-MnO2, goethite, and bayerite, HLA showed a greater degree of condensation compared to HLA precipitated by kaolinite and montmorillonite. Contributions from Si-O, and Si-O-Al of clay minerals, and Fe-O, Mn-O, and Al-O of oxides were the mechanisms by which minerals catalyzed the formation of HS from lignin.

Trimebutine, a small molecule mimetic agonist of adhesion molecule L1, contributes to functional recovery after spinal cord injury in mice.

Curing spinal cord injury (SCI) in mammals is a daunting task because of the lack of permissive mechanisms and strong inhibitory responses at and around the lesion. The neural cell adhesion molecule L1CAM (L1) has been shown to favor axonal regrowth and enhance neuronal survival and synaptic plasticity but delivery of full-length L1 or its extracellular domain could encounter difficulties in translation to therapy in humans. We have, therefore, identified several small organic compounds that bind to L1 and stimulate neuronal survival, neuronal migration and neurite outgrowth in an L1-dependent manner. Here, we assessed the functions of two L1 mimetics, trimebutine and honokiol, in regeneration following SCI in young adult mice. Using the Basso Mouse Scale (BMS) score, we found that ground locomotion in trimebutine-treated mice recovered better than honokiol-treated or vehicle-receiving mice. Enhanced hindlimb locomotor functions in the trimebutine group were observed at 6 weeks after SCI. Immunohistology of the spinal cords rostral and caudal to the lesion site showed reduced areas and intensities of glial fibrillary acidic protein immunoreactivity in both trimebutine and honokiol groups, whereas increased regrowth of axons was observed only in the trimebutine-treated group. Both L1- and L1 mimetic-mediated intracellular signaling cascades in the spinal cord lesion sites were activated by trimebutine and honokiol, with trimebutine being more effective than honokiol. These observations suggest that trimebutine and, to a lesser extent under the present experimental conditions, honokiol have a potential for therapy in regeneration of mammalian spinal cord injuries.

Neuregulin-1 protects mouse cerebellum against oxidative stress and neuroinflammation.

Cerebellum undergoes degenerative changes in neurodegenerative diseases. Two main factors including oxidative stress and neuroinflammation mediate neurodegeneration. Neuregulin-1 (Nrg1) has been implicated in many neurodegenerative diseases, while the underlying mechanisms are unknown. We hypothesized that Nrg1 prevents oxidative stress and neuroinflammation in neurodegeneration. We found a positive correlation between Nrg1 protein levels and ErbB4 and ErbB2 receptor phosphorylation in microarrays of normal human cerebellar tissue. In addition, Nrg1 was also co-localized with pErbB4 and pErbB2. Primary mouse cerebellar granule neurons (CGNs) were treated with H2O2 or LPS combined with recombinant Nrg1ß (rNrg1ß). Western blot analysis and immunofluorescence revealed that H2O2 and LPS-induced neuronal toxicity down-regulated the activation of ErbB receptors and Akt1, and the ratio of Bcl2/Bax, which was reversed by rNrg1ß. In vivo studies showed that LPS-induced neuroinflammation in mouse cerebellum down-regulated pErbB4, pErbB2, pAkt1/Akt1 and Bcl2/Bax levels, whereas rNrg1ß treatment reversed the changes. Immunohistochemistry and Western blot analysis showed that rNrg1ß alleviates neuroinflammation by reducing the number of microglial cells and astrocytes and the expression of IL1ß. Our results indicate that Nrg1 protects against oxidative stress and neuroinflammation in mouse cerebellum, suggesting potential therapeutic application in neuroinflammation associated with neurodegenerative diseases.

Over-expression of heat shock protein 70 protects mice against lung ischemia/reperfusion injury through SIRT1/AMPK/eNOS pathway.

Lung ischemia/reperfusion injury (LIRI) usually occurs during in lung transplantation and extracorporeal circulation operation and may develop into pulmonary infections, acute rejection and bronchiolitis obliterans syndrome. Recent studies have discovered the protective effect of heat shock protein 70 (HSP70) on various types of injuries. In the present study, we firstly explore the role of over-expressed HSP70 on the protection against LIRI. Lung Wet/Dry (W/D) ratio, biomarkers in the bronchoalveolar lavage fluid (BALF), lung histological changes and apoptosis markers, oxidative products and proinflammatory cytokines in the lung tissues were analyzed. Next, the expression of eNOS, SIRT1 and AMPK were measured. Finally, the changes of the lung W/D ratio and biomarkers in the BALF using the inhibitors of SIRT1/AMPK/eNOS pathway were evaluated. Mice exposed to LIRI procedure had significant increases in lung W/D ratio and biomarkers (protein level, LDH level, leukocytes and total cells) in BALF. LIRI also caused histological injury, demonstrated by hemorrhage, alveolar septal thickening and fibrin deposition. Apoptosis, oxidative products and proinflammatory cytokines in lung tissue were also induced by LIRI. The over-expression of HSP70 antagonized the impacts of LIRI by attenuating these parameters. It significantly increased the expression of eNOS, SIRT1 and AMPK, while the inhibition of SIRT1 and AMPK deactivated the eNOS expression. The lung W/D ratio and biomarkers in BALF were increased while mice were given inhibitors of eNOS, SIRT1 and AMPK. We concluded that over-expression of HSP70 had protective effect on LIRI and HSP70 might be involved in the protection through a SIRT1/AMPK/eNOS pathway.

Construction of protein interaction network involved in lung adenocarcinomas using a novel algorithm.

Studies that only assess differentially-expressed (DE) genes do not contain the information required to investigate the mechanisms of diseases. A complete knowledge of all the direct and indirect interactions between proteins may act as a significant benchmark in the process of forming a comprehensive description of cellular mechanisms and functions. The results of protein interaction network studies are often inconsistent and are based on various methods. In the present study, a combined network was constructed using selected gene pairs, following the conversion and combination of the scores of gene pairs that were obtained across multiple approaches by a novel algorithm. Samples from patients with and without lung adenocarcinoma were compared, and the RankProd package was used to identify DE genes. The empirical Bayesian (EB) meta-analysis approach, the search tool for the retrieval of interacting genes/proteins database (STRING), the weighted gene coexpression network analysis (WGCNA) package and the differentially-coexpressed genes and links package (DCGL) were used for network construction. A combined network was also constructed with a novel rank-based algorithm using a combined score. The topological features of the 5 networks were analyzed and compared. A total of 941 DE genes were screened. The topological analysis indicated that the gene interaction network constructed using the WGCNA method was more likely to produce a small-world property, which has a small average shortest path length and a large clustering coefficient, whereas the combined network was confirmed to be a scale-free network. Gene pairs that were identified using the novel combined method were mostly enriched in the cell cycle and p53 signaling pathway. The present study provided a novel perspective to the network-based analysis. Each method has advantages and disadvantages. Compared with single methods, the combined algorithm used in the present study may provide a novel method to analyze gene interactions, with increased credibility.

Differential changes in Neuregulin-1 signaling in major brain regions in a lipopolysaccharide-induced neuroinflammation mouse model.

Neuregulin 1 (Nrg1) is involved in multiple biological processes in the nervous system. The present study investigated changes in Nrg1 signaling in the major brain regions of mice subjected to lipopolysaccharide (LPS)-induced neuroinflammation. At 24 h post­intraperitoneal injection of LPS, mouse brain tissues, including tissues from the cortex, striatum, hippocampus and hypothalamus, were collected. Reverse transcription­polymerase chain reaction was used to determine the expression of Nrg1 and its receptors, Neu and ErbB4, at the mRNA level. Western blotting was performed to determine the levels of these proteins and the protein levels of phosphorylated extracellular signal-regulated kinases (Erk)1/2 and Akt1. Immunohistochemical staining was utilized to detect the levels of pNeu and pErbB4 in these regions. LPS successfully induced sites of neuroinflammation in these regions, in which changes in Nrg1, Neu and ErbB4 at the mRNA and protein levels were identified compared with controls. LPS induced a reduction in pNeu and pErbB4 in the striatum and hypothalamus, although marginally increased pErbB4 levels were found in the hippocampus. LPS increased the overall phosphorylation of Src but this effect was reduced in the hypothalamus. Moreover, increased phosphorylation of Akt1 was found in the striatum and hippocampus. These data suggest diverse roles for Nrg1 signaling in these regions during the process of neuroinflammation.