Development of an mRNA vaccine against a panel of heterologous H1N1 seasonal influenza viruses using a consensus hemagglutinin sequence.

Seasonal influenza, causes hundreds of thousands of deaths annually, posing a severe threat to human health. Currently available influenza vaccines are targeted only at specific strains or conserved epitopes; however, these vaccines are not completely efficacious because influenza viruses can undergo mutation during circulation, leading to antigenic mismatch between recommended strains and circulating strains and elusion from the immune system. Therefore, developing an influenza vaccine that is quick, effective, and broadly protective has become crucial, and the integral part of hemagglutinin (HA) remains an ideal target for vaccine development. This study developed a lipid nanoparticle-encapsulated nucleoside-modified mRNA vaccine (mRNA-LNPs) encoding a consensus full-length HA sequence (H1c) and evaluated its protective efficacy and immunogenicity through in vitro and in vivo assays. Following two intramuscular immunizations (2, 10 µg, or 20 µg) at a 3-week interval in BALB/c mice, H1c-mRNA-LNP vaccine induced strong antibodies as shown in the hemagglutination-inhibition test and protective neutralizing antibodies against numerous heterologous H1N1 influenza viruses as shown in the microneutralization assay. Additionally, both Th1- and Th2-biased cellular immune responses were elicited, with the Th1-biased response being stronger. Two doses of the H1c-mRNA-LNP vaccine could neutralize a panel of heterologous H1N1 influenza viruses and could confer protection in mice. Taken together, these findings suggest that the H1c-mRNA-LNP vaccine encoding a consensus full-length HA is a feasible strategy for developing a cross-protective vaccine against a panel of heterologous H1N1 influenza viruses.

Automated machine learning-based model for the prediction of delirium in patients after surgery for degenerative spinal disease.

OBJECTIVE: This study used machine learning algorithms to identify critical variables and predict postoperative delirium (POD) in patients with degenerative spinal disease. METHODS: We included 663 patients who underwent surgery for degenerative spinal disease and received general anesthesia. The LASSO method was used to screen essential features associated with POD. Clinical characteristics, preoperative laboratory parameters, and intraoperative variables were reviewed and were used to construct nine machine learning models including a training set and validation set (80% of participants), and were then evaluated in the rest of the study sample (20% of participants). The area under the receiver-operating characteristic curve (AUROC) and Brier scores were used to compare the prediction performances of different models. The eXtreme Gradient Boosting algorithms (XGBOOST) model was used to predict POD. The SHapley Additive exPlanations (SHAP) package was used to interpret the XGBOOST model. Data of 49 patients were prospectively collected for model validation. RESULTS: The XGBOOST model outperformed the other classifier models in the training set (area under the curve [AUC]: 92.8%, 95% confidence interval [CI]: 90.7%-95.0%), validation set (AUC: 87.0%, 95% CI: 80.7%-93.3%). This model also achieved the lowest Brier Score. Twelve vital variables, including age, serum albumin, the admission-to-surgery time interval, C-reactive protein level, hypertension, intraoperative blood loss, intraoperative minimum blood pressure, cardiovascular-cerebrovascular disease, smoking, alcohol consumption, pulmonary disease, and admission-intraoperative maximum blood pressure difference, were selected. The XGBOOST model performed well in the prospective cohort (accuracy: 85.71%). CONCLUSION: A machine learning model and a web predictor for delirium after surgery for the degenerative spinal disease were successfully developed to demonstrate the extent of POD risk during the perioperative period, which could guide appropriate preventive measures for high-risk patients.

A Label-free aptasensor based on Aptamer/NH2 Janus particles for ultrasensitive electrochemical detection of Ochratoxin A.

In this study, a novel aptasensor based on Aptamer/NH2 Janus particles is developed for the detection of Ochratoxin A(OTA). By coating gold on the hemispherical surface of the aminated polystyrene particles, Ochratoxin A aptamer is immobilized on the surface of the gold layer for selective identification and the other hemispherical able to bind to Glassy carbon electrode via peptide bond. Under optimum conditions, the sensor exhibited a wide dynamic range of OTA concentration from 1 × 10-5 nM to 10 nM, and the detection limit is 3.3 × 10-3 pM on condition of acceptable stability and reproducibility. The sensors were showed excellent performance in the detection of OTA in red wine sample with recoveries between 95.7% and 100.18%, which studied by the standard addition spiking technique. This work provides a new idea and method for preparing immune electrochemical sensors and is expected to be used for the OTA detection in red wine sample analysis.

Let-7i inhibits the malignant phenotype of osteosarcoma cells by targeting Aurora-B.

Our previous study indicated that Aurora-B is involved in osteosarcoma (OS) cell invasion and metastasis; however, the mechanism underlying Aurora-B overexpression in OS remains unknown. In the present study, significantly downregulated let-7i expression in OS tissues and OS cells was observed compared with that in normal adjacent tumorous tissues and human osteoblast cell lines. Bioinformatic predictions have revealed a conserved binding site in a microRNA locus on Aurora­B, suggesting the potential of let­7i targeting the Aurora­B gene. To validate this, a luciferase reporter assay was performed on OS cells. The results indicated that Aurora­B is a likely to be a direct target negatively regulated by let­7i. The expression of let­7i in OS cells was restored by infection with let­7i mimics. Results revealed that Aurora­B mRNA and protein expression levels were significantly decreased. Furthermore, the proliferation, migration and invasion abilities of OS cells were significantly suppressed by infection with let­7i mimics. Notably, the inhibitory effect of silencing Aurora­B by LV­shAurora­B on cell proliferation, migratory and invasive ability was significantly lower than that by let­7i mimics, which indicated that let­7i inhibits cell malignant phenotypes partially by targeting Aurora­B in OS cells. All data suggested that let­7i may be a novel potential target for OS treatment.

LY294002 suppresses the malignant phenotype and sensitizes osteosarcoma cells to pirarubicin chemotherapy.

Pirarubicin is frequently used in chemotherapy against tumors. However, clinical use is severely limited by the development of progressive dose-dependent cardiomyopathy and acquired drug resistance. LY294002 is a commonly used pharmacologic inhibitor, which selectively inhibits the phosphoinositide 3-kinase-AKT nexus. The aim of this study was to investigate the combined inhibitory effect of LY294002 and pirarubicin on human osteosarcoma (OS) cells in vitro. The inhibitory effect of LY294002 plus pirarubicin on U2-OS and MG-63 OS cell proliferation, apoptosis, migration and invasion was investigated by cell proliferation, wound healing and Transwell invasion assays. The results revealed that LY294002 and pirarubicin synergistically induced apoptosis, and inhibited the growth, migration and invasion of OS cells. This indicates that LY294002 enhanced the effects of pirarubicin on OS in vitro. LY294002 combined with pirarubicin may thus be a future therapeutic strategy in OS.

Let-7g reverses malignant phenotype of osteosarcoma cells by targeting Aurora-B.

Accumulating studies revealed that the expression levels of several miRNAs are up or down-regulated in osteosarcoma (OS). The aim of this study was to investigate the functional significance and molecular of the let-7g in OS cells. The expression levels of let-7g was significantly down-regulated in OS cell lines U2-OS and HOS cell compared to osteoblast cell lines HOB cell. Moreover, bioinformatic prediction suggested that Aurora-B, which is overexpressed and functions as an oncogene in OS cells, is a putative target gene of let-7g. Using mRNA and protein expression analysis and luciferase assays, we further identified let-7g directly regulated Aurora-B expression in OS cells. Functional investigation revealed both restoration of let-7g and silencing Aurora-B induce cell apoptosis and suppressed cell viability, migratory and invasive ability in OS cells. Finally, we found that silencing Aurora-B in OS cells could partly dampen anti-let-7g mediated tumor promotion. Thus, our findings suggested that let-7g inhibits OS cell malignant phenotype at least partly through targeting Aurora-B. Targeting of let-7g and Aurora-B may be a novel therapeutic strategy for treating OS.

Lapatinib alters the malignant phenotype of osteosarcoma cells via downregulation of the activity of the HER2-PI3K/AKT-FASN axis in vitro.

Lapatinib, an inhibitor of human epidermal growth factor receptor 2 (HER2) phosphorylation, has been reported to inhibit several types of tumors such as HER2-overexpressing breast cancer. However, the effect of lapatinib on the malignant phenotype of human osteosarcoma (OS) cells and the potential molecular mechanisms remain unclear. To elucidate the effect of lapatinib on OS, two OS cell lines, U2-OS and MG-63, were utilized in the present study. Various concentrations of lapatinib were used to treat OS cells for different time durations. Cell proliferation was evaluated by MTT and colony formation assays. Flow cytometry (FCM) was used to evaluate cell apoptosis. Wound healing and Transwell invasion assays were performed to examine the migratory and invasive abilities of the cells. To investigate the possible molecular mechanisms involved, the expression of p-HER2, phosphatidylinositol 3-kinase (PI3K), p-AKT, AKT and fatty acid synthase (FASN) protein was detected by western blotting. MTT assays showed that lapatinib inhibited the proliferation of U2-OS and MG-63 cells in a dose- and time-dependent manner, and the rate of colony formation of the lapatinib-treated cells was significantly lower when compared to those cells not treated with lapatinib in both cell lines. FCM assay revealed a significantly higher apoptotic rate in the lapatinib-treated OS cells. Wound healing and Transwell invasion assays revealed that the migratory and invasive abilities of OS cells were significantly inhibited by lapatinib (P<0.05). Western blotting showed that lapatinib suppressed the activity of HER2-PI3K/AKT-FASN in U2-OS and MG-63 cells in vitro. These results suggest that lapatinib may alter the malignant phenotype of OS cells via downregulation of the activity of the HER2-PI3K/AKT-FASN signaling pathway in vitro. Thus, lapatinib may be an effective chemotherapeutic agent for the treatment of osteosarcoma.

Interaction between fatty acid synthase and human epidermal growth receptor 2 (HER2) in osteosarcoma cells.

Elevated expression of fatty acid synthase (FASN) and human epidermal growth factor receptor-2 (HER2) are observed in human osteosarcoma (OS). The aim in this study is to investigate a possible connection between FASN expression and the activity of HER2. The immunohistochemistry staining was conducted on 24 OS specimens from patients, which revealed a significant positive correlation between FASN and HER2 as well as p-HER2 protein expression. Furthermore, the human OS cell lines MG-63 and U2-OS were treated with FASN-specific RNAi Plasmid or Lapatinib (an inhibitor of HER2). The mRNA of HER2 and FASN was measured using RT-PCR. Western blot was performed to detect the protein expression of HER2, p-HER2 and FASN. The results demonstrated that HER2 modulates FASN expression, inhibition of FASN resulted in down-regulation of HER2 and p-HER2 protein in OS cells. Our findings suggested that there was positive feedback regulation between FASN and HER2 expression and phosphorylation in OS cells.

Positive feedback regulation between Akt phosphorylation and fatty acid synthase expression in osteosarcoma.

The activation of PI3K/Akt and the overexpression of fatty acid synthase (FASN) are frequently observed in human osteosarcoma (OS). In the present study, in order to investigate the possible association between the phosphorylation of Akt and FASN expression, immunohistochemical staining was conducted on 24 OS specimens from patients with pulmonary metastasis, which revealed a significant positive correlation between phosphorylated Akt (p-Akt) and the expression of FASN (R=0.469, P=0.04). To investigate the association between p-Akt and FASN in vitro, human U2-OS OS cells were treated with FASN-specific RNAi plasmid or LY294002 (an inhibitor of PI3k/Akt). The mRNA levels of Akt and FASN were measured by real-time PCR. Western blot analysis was also performed to detect the protein experession of PI3K, Akt, p-Akt and FASN. The results demonstrated that the PI3K/Akt signaling pathway modulates FASN expression; the inhibition of FASN resulted in the downregulation of p-Akt in the U2-OS cells. Furthermore, the effects induced by the inhibition of the activity of p-Akt or FASN on the malignant phenotype of U2-OS cells were investigated, demonstrating that the malignant phenotype was inhibited by suppressing the activity of PI3K/Akt or FASN in the U2-OS cells. The findings from our study suggest the existence of a positive feedback regulation between Akt phosphorylation and FASN expression and that this loop may play an important role in the malignant phenotype of OS cells.

[Study on the detoxification of alcohol extracts from orientvine and its effective component on withdrawal syndromes of morphine].

OBJECTIVE: To observe the effect of alcohol extracts from orientivne and its effective component sinomenine on withdrawal syndromes and neurotransmitter of morphine-abstinent mice and on intracellular calcium level in morphine-dependent neuronal-cells line. To study the detoxification of alcohol extracts from orientvine and sinomenine on morphine-dependent animal and explore the mechanism of its effect. METHODS: The effect of alcohol extracts from orientivne and sinomenineon on abstinent syndromes was observed by experiment study on morphine-dependent ex vivo ileum from guinea pigs and morphine-dependent mice. The morphine-dependent model mice were established by injection on dosage increasing by degrees. The hypothalamic monomine neurotransmitters such as NA, DA, 5-HT were tested by fluorospectrophotometry. Morphine-dependent cell line was established by administering morphine at different doses into the culture medium. The cells were stained with fluo-3 and the intracellular calcium level was measured by flow cytometry. RESULTS: Alcohol extracts from orientvine and sinomenine could alleviate withdrawal contractile response of morphine-dependent ex vivo ileum from guinea pigs and withdrawal syndromes of morphine-dependent mice, decrease the concentration of the neurotransmitters, and elevate the intracellular calcium level and inhibit the decreasing of Ca2+ induced by naloxone. CONCLUSIONS: Alcohol extracts from orientvine and sinomenine have some effects on withdrawal syndromes which may be related to inhibiting neurotransmitters and the regulation of intracellular calcium concentration.

[Study of the effect of aliphatic alcohol on the inclusion interaction of qucertin-cyclodextrin complexes].

The inclusion interaction of qucertin with beta-cyclodextrin and its derivatives was studied by fluorimetry. The formation constants of their inclusion complexes were determined, and the stoichiometry ratio on the formation of the inclusion complexes was determined by equimolar continueous variation method. The effect of aliphatic alcohols with short chains on inclusion process was discussed. The results showed that the inclusion ability of beta-CD and its derivatives was in the order: HP-beta-CD>M-beta-CD>beta-CD. The effect of aliphatic alcohol on complexes was related to the carbon number and volume of alcohol molecules.

[The application of the integrative management with no blind spot in the cooperative projects for prevention of blindness].

OBJECTIVE: To evaluate the management for donated funds in cooperative projects for prevention of blindness. METHODS: Retrospectively analyze the integrative management with no blind spot in the cooperative projects conducted by the Center of Preventing and Treating Blindness in Anhui Province and the International Organization for Prevention of Blindness from 1992 to 2001. RESULTS: The integrative management such as standardization of fund budget, establishment of inner-controlling system, improvement of inner-management and enhancement of project audit, returning information to donors and obtaining consent were applied in the cooperative projects for prevention of blindness. Therefore, the funds were used reasonably and projects were smoothly accomplished. CONCLUSION: The integrative management with no blind spot is effective for the management of donated funds in the cooperative projects for blindness.