Association of procalcitonin with voriconazole concentrations: a retrospective cohort study.

Inflammation is a potential risk factor of voriconazole (VCZ) overdose, procalcitonin (PCT) is reported to act as a diagnostic marker for bacterial infections. However, the association of PCT with VCZ trough serum concentrations (VCZ-Cmin) is not fully clear. Our study aims to investigate the associations between PCT and VCZ-Cmin. In this retrospective cohort study, we collected the clinical data of 147 patients who received VCZ and monitored the VCZ concentration of them in our hospital from August 2017 to August 2021. All patients underwent routine clinical examinations on the day or the day before VCZ administration. General information and clinical symptoms of these patients were recorded. Multivariate liner analysis showed that PCT was significantly associated with VCZ-Cmin (p < 0.001). Overall, it was shown that VCZ-Cmin was significantly increased by 0.32 µg/mL for each fold increment in PCT in crude model. In the minor adjusted model (Model 1, adjustment for sex, age, albumin, direct bi1irubin, WBC) and fully adjusted model (Model 2, adjustment for sex, age, albumin, direct bilirubin, WBC, AST and ALT), VCZ-Cmin was significantly increased by 0.23 µg/mL and 0.21 µg/mL, respectively, for each fold increment in PCT. In conclusion, this research reveals the correlation between PCT and VCZ-Cmin, indicating that PCT has the potential to serve as a valuable biomarker for drug monitoring in the treatment of VCZ.

Engineered mesenchymal stem cell-derived extracellular vesicles: A state-of-the-art multifunctional weapon against Alzheimer's disease.

With the increase of population aging, the number of Alzheimer's disease (AD) patients is also increasing. According to current estimates, approximately 11% of people over 65 suffer from AD, and that percentage rises to 42% among people over 85. However, no effective treatment capable of decelerating or stopping AD progression is available. Furthermore, AD-targeted drugs composed of synthetic molecules pose concerns regarding biodegradation, clearance, immune response, and neurotoxicity. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) are essential intercellular communication mediators holding great promise as AD therapeutics owing to their biocompatibility, versatility, effortless storage, superior safety, and the ability to transport messenger and noncoding RNAs, proteins, lipids, DNAs, and other bioactive compounds derived from cells. The functionalisation and engineering strategies of MSC-EVs are highlighted (e.g. preconditioning, drug loading, surface modification, and artificial EV fabrication), which could improve AD treatment by multiple therapeutic effects, including clearing abnormal protein accumulation and achieving neuroprotection and immunomodulatory effects. Herein, this review summarises state-of-the-art strategies to engineer MSC-EVs, discusses progress in their use as AD therapeutics, presents the perspectives and challenges associated with the related clinical applications, and concludes that engineered MSC-EVs show immense potential in AD therapy.

Involvement of microRNA/cystine/glutamate transporter in cold-stressed gastric mucosa injury.

Stress ulcers are complicated by severe trauma and other critical diseases, the mechanism of which remains unclear. An increasing number of studies have shown that microRNAs (miRNAs) are important regulators of stress responses such as hypoxia, abnormal temperature, and inflammation. The evidence indicates that miRNAs are also involved in regulating stress-induced ulcers. Recently, we demonstrated that gastric mucosal injury induced by aspirin is related to the reduction of glutamate levels by inhibition of cystine/glutamate transporter (xCT) activity. In the present study, the effect of a miRNA/xCT on gastric mucosal injury induced by cold stimulation was investigated. We found that cold stimulation induced gastric mucosa injury with a reduction in glutamate levels and xCT activity and upregulation of miR-143, miR-152, and miR-181 expression. Exogenous glutamate significantly alleviated gastric mucosa injury by cold stimulation. In vitro experiments demonstrated that treatment with miR-143, miR-152, or miR-181 mimics directly induced cell damage. The effects of these mimics were alleviated by exogenous glutamate. The present study suggests that miR-143, miR-152, and miR-181 are involved in cold stimulation-induced acute gastric mucosal injury. Furthermore, the regulatory effect of miRNAs on gastric mucosa injury induced by cold stimulation is related to a decrease in glutamate release by reduction of cystine/glutamate transporter activity.

Gold nanoparticle-directed autophagy intervention for antitumor immunotherapy via inhibiting tumor-associated macrophage M2 polarization.

Tumor-associated macrophages (TAMs), one of the dominating constituents of tumor microenvironment, are important contributors to cancer progression and treatment resistance. Therefore, regulation of TAMs polarization from M2 phenotype towards M1 phenotype has emerged as a new strategy for tumor immunotherapy. Herein, we successfully initiated antitumor immunotherapy by inhibiting TAMs M2 polarization via autophagy intervention with polyethylene glycol-conjugated gold nanoparticles (PEG-AuNPs). PEG-AuNPs suppressed TAMs M2 polarization in both in vitro and in vivo models, elicited antitumor immunotherapy and inhibited subcutaneous tumor growth in mice. As demonstrated by the mRFP-GFP-LC3 assay and analyzing the autophagy-related proteins (LC3, beclin1 and P62), PEG-AuNPs induced autophagic flux inhibition in TAMs, which is attributed to the PEG-AuNPs induced lysosome alkalization and membrane permeabilization. Besides, TAMs were prone to polarize towards M2 phenotype following autophagy activation, whereas inhibition of autophagic flux could reduce the M2 polarization of TAMs. Our results revealed a mechanism underlying PEG-AuNPs induced antitumor immunotherapy, where PEG-AuNPs reduce TAMs M2 polarization via induction of lysosome dysfunction and autophagic flux inhibition. This study elucidated the biological effects of nanomaterials on TAMs polarization and provided insight into harnessing the intrinsic immunomodulation capacity of nanomaterials for effective cancer treatment.

LC-MS/MS method for quantifying aescinate A and B and assessing their relationship with phlebitis.

The purpose of this study is to establish and validate a sensitive, robust and rapid liquid chromatography-tandem mass spectrometry method for quantifying the aescinate A and aescinate B in human plasma and assessing the association of phlebitis and aescinate A and aescinate B in vivo exposure. The chromatographic separation was completed on Agilent ZORBAX SB-C18 (2.1 mm × 100 mm, 3.5 µm, Agilent, USA) column with isocratic elution. The flow rate was 0.3 mL/min and the total run time was optimized within 5 min. The protein precipitation was applied to pretreat plasma sample using methanol as precipitant. The data acquisition was achieved with positive electrospray ionization in multi-reaction monitoring mode for both aescinate A and aescinate B. The calibration range of aescinate A and aescinate B are constructed in 100-2000 ng/mL, and their correlation coefficients are both >0.990. The intra-day and inter-day precision and accuracy of this method are less than 9.04% and within -13.75% and -0.93%. This analytical method has been successfully applied for the determination of plasma aescinate A and aescinate B concentrations in patients with cerebral infarction, and the results showed that the incidence and grade of phlebitis were not associated with the in vivo exposure of aescinate A and aescinate B.

Regional cerebral perfusion in patients with amnestic mild cognitive impairment: effect of cerebral small vessel disease.

OBJECTIVE: To explore the effort of cerebral small vessel disease (CSVD) on regional cerebral perfusion in patients with mild cognitive impairment (MCI) using NeuroGam™ software and evaluate the capability of brain perfusion single photon emission computed tomography (SPECT) in distinguishing MCI with and without CSVD. METHODS: 34 amnestic MCI subjects entered this study, conducting neuropsychological tests, MRI and 99mTechnetium ethyl cystine dimer brain perfusion SPECT imaging. All subjects were divided into those with CSVD and those without CSVD. Perfusion value was measured with Brodmann area (BA) mapping in these two groups. Automated software (NeuroGam™) was used for semi-quantitative analyses of perfusion value and comparison with normal database. RESULTS: Compared with normal database, perfusion levels in BAs 23-left, 28 and 36-left of MCI without CSVD group had great deviations, while perfusion levels in BAs 21, 23, 24, 25, 28, 36, 38 and 47-left of MCI with CSVD group had great deviations. Furthermore, compared with CSVD group, there was significantly lower perfusion value in BA 7-left (P < 0.001) in MCI without CSVD group. CONCLUSIONS: CSVD could interact with pathological changes related to AD, exacerbating hypoperfusion in BAs 21, 23, 28, 36, 38 while compensating for cerebral blood perfusion disorder in BA 7-left in MCI patients. Meanwhile, MCI patients with CSVD shared similar hypoperfusion with vascular cognitive impairment (VCI) in BAs 24, 25 and 47L. Brain perfusion SPECT may help improve our ability to differentiate MCI with and without CSVD.

[The efficacy of topiramate and flunarizine hydrochloride for prophylactic treatment of vestibular migraine].

Objective:To evaluate the efficacy and safety of topiramate and flunarizine hydrochloride in the prophylactic treatment of vestibular migraine prophylaxis. Methods:47 patients with confirmed or probable vestibular migraine（VM） treated at the vertigo clinic of our neurology department from August 2020 to April 2021 were reviewed, and 42 patients were finally included. They were divided into topiramate group （n=22） and flunarizine hydrochloride group （n=20）. The two groups were treated with topiramate 50 mg daily and flunarizine hydrochloride 10 mg daily, respectively. The visual analogue scale, vertigo duration, vertigo frequency, and Dizziness Handicap Inventory （DHI） scores of patients with VM before and 3 months after treatment were compared. The anxiety screening scale （GAD-7） and depression screening scale （PHQ-9） were recorded to assess the improvement of patients' anxiety and depression, and the occurrence of adverse events. Results:Topiramate and flunarizine hydrochloride effectively reduced vertigo intensity, vertigo duration, and vertigo frequency in VM patients （P<0.05）. Meanwhile, total DHI score, DHI physical （DHI-P）, DHI emotional （DHI-E）, DHI functional （DHI-F）, PHQ-9 and GAD-7 were significantly decreased（P<0.05）. Furthermore, topiramate was superior to flunarizine hydrochloride in reducing vertigo intensity, vertigo duration, vertigo frequency, DHI-P, and DHI-F, while there was no significant difference between two drugs in improving patients' mood（P>0.05）. No serious adverse events were reported in either group. Conclusion:This study suggests that topiramate and flunarizine hydrochloride are safe and effective in the prevention of VM, and the daily dose of topiramate 50 mg is superior to the daily dose of flunarizine hydrochloride 10 mg. However, there was no significant difference between the two drugs in terms of mood improvement.

Tumor RNA-loaded nanoliposomes increases the anti-tumor immune response in colorectal cancer.

PURPOSE: Tumor RNA vaccines can activate dendritic cells to generate systemic anti-tumor immune response. However, due to easily degraded of RNA, direct RNA vaccine is less effective. In this study, we optimized the method for preparing PEGylated liposom-polycationic DNA complex (LPD) nanoliposomes, increased encapsulate amount of total RNA derived from CT-26 colorectal cancer cells. Tumor RNA LPD nanoliposomes vaccines improved anti-tumor immune response ability of tumor RNA and can effectively promote anti-tumor therapeutic effect of oxaliplatin. METHODS: Total tumor-derived RNA was extracted from colorectal cancer cells (CT-26 cells), and loaded to our optimized the LPD complex, resulting in the LPD nanoliposomes. We evaluated the characteristics (size, zeta potential, and stability), cytotoxicity, transfection ability, and tumor-growth inhibitory efficacy of LPD nanoliposomes. RESULTS: The improved LPD nanoliposomes exhibited a spherical shape, RNA loading efficiency of 9.07%, the average size of 120.37 ± 2.949 nm and zeta potential was 3.34 ± 0.056 mV. Also, the improved LPD nanoliposomes showed high stability at 4 °C, with a low toxicity and high cell transfection efficacy toward CT-26 colorectal cancer cells. Notably, the improved LPD nanoliposomes showed tumor growth inhibition by activating anti-tumor immune response in CT-26 colorectal cancer bearing mice, with mini side effects toward the normal organs of mice. Furthermore, the effect of the improved LPD nanoliposomes in combination with oxaliplatin can be better than that of oxaliplatin alone. CONCLUSION: The improved LPD nanoliposomes may serve as an effective vaccine to induce antitumor immunity, presenting a new treatment option for colorectal cancer.

Immunotherapy of Tumor RNA-Loaded Lipid Nanoparticles Against Hepatocellular Carcinoma.

PURPOSE: Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide. Most current therapeutic strategies primarily include localized treatment, lacking effective systemic strategies. Meanwhile, recent studies have suggested that RNA vaccines can effectively activate antigen-presenting cells (APCs) and lymphocytes to produce a strong systemic immune response and inhibit tumor growth. However, tumor vaccines loaded with a single tumor antigen may induce immunosuppression and immune evasion, while identifying tumor-specific antigens can require expensive and laborious procedures. Therefore, the use of whole tumor cell antigens are currently considered to be promising, potentially effective, methods. Previously, we developed a targeted liposome-polycation-DNA (LPD) complex nanoparticle that possess a small size, high RNA encapsulation efficiency, and superior serum stability. These particles were found to successfully deliver RNA to tumor sites. In the current study, we encapsulated total tumor-derived RNA in lipid nanoparticles (LNPs) to target dendritic cells (DCs) to incite expeditious and robust anti-tumor immunity. METHODS: Total tumor-derived RNA was extracted from liver cancer cells (Hepa1-6 cells). LNPs loaded with tumor RNA were then prepared thin-film hydration method. The ability of RNA LNPs to induce DC maturation, cytotoxicity, and anti-tumor activity, was investigated in vitro and in vivo. RESULTS: The average particle size of LNPs and RNA LNPs was 102.22 ± 4.05 nm and 209.68 ± 6.14 nm, respectively, while the zeta potential was 29.97 ± 0.61 mV and 42.03 ± 0.42 mV, respectively. Both LNPs and RNA LNP vaccines exhibited good distribution and stability. In vitro, RNA LNP vaccines were capable of promoting DC maturation and inducing T lymphocytes to kill Hepa1-6 cells. In vivo, RNA LNP vaccines effectively prevent and inhibit HCC growth. CONCLUSION: RNA LNPs may serve as an effective antigen specific vaccine to induce anti-tumor immunity for HCC.

PEG-Ceramide Nanomicelles Induce Autophagy and Degrade Tau Proteins in N2a Cells.

PURPOSE: Alzheimer's disease (AD) is a neurodegenerative disorder that manifests as abnormal behavior and a progressive decline in memory. Although the pathogenesis of AD is due to the excessive deposition of amyloid ß protein (Aß) outside the neurons in the brain, evidence suggests that tau proteins may be a better target for AD therapy. In neurodegenerative diseases, a decrease in autophagy results in the failure to eliminate abnormally deposited or misfolded proteins. Therefore, induction of autophagy may be an effective way to eliminate tau proteins in the treatment of AD. We investigated the effects of polyethylene glycol (PEG)-ceramide nanomicelles on autophagy and on tau proteins in N2a, a murine neuroblastoma metrocyte cell line. METHODS: Ceramide is a sphingolipid bioactive molecule that induces autophagy. PEG-ceramide is a polymer that is composed of the hydrophobic chain of ceramide and the hydrophilic chain of PEG-2000. In this study, we prepared PEG-ceramide nanomicelles that were 10-20 nm in size and had nearly neutral zeta potential. RESULTS: The results show that PEG-ceramide nanomicelles caused an increase in the LC3-II/LC3-I ratio, while p62 protein levels decreased. Confocal microscopy revealed a significant increase in the number of dots corresponding to autophagosomes and autolysosomes, which indicated autophagic activation. Moreover, PEG-ceramide nanomicelles induced tau degradation in N2a cells through autophagy. CONCLUSION: In summary, we have confirmed that PEG-ceramide nanomicelles enhanced autophagic flux and degraded overexpressed human tau proteins in N2a cells by regulating the autophagy pathway. Thus, PEG-ceramide nanomicelles show great promise as agents to induce autophagy and degrade tau proteins in the treatment of AD.

Variations of soil microbial communities accompanied by different vegetation restoration in an open-cut iron mining area.

Overexploitation of iron mining in China has caused serious environmental pollution. Therefore, establishing a stable ecological restoration with vegetation in mining areas has gradually aroused people's awareness and obtained extensive concerns. This study aimed to evaluate vegetation restoration with Robinia pseudoacacia (RP), Acer mono (AM) and Pinus koraiensis (PK) in iron mining compared with unrestored area, to investigate the soil environment factors and microbial communities, and to better understand the correlations between soil environment factors and soil microbial communities. Vegetation restoration could reduce soil pH and alleviate soil alkaline, and remarkably increase soil nutrients, especially in RP site. Analysis of 16S rRNA and ITS rRNA gene sequences provided a total of 645,004 and 906, 276 valid sequences clustered into 7091 OTUs and 1689 OTUs at a 0.03 genetic distance for bacteria and fungi, respectively. The predominant bacterial and fungal phyla were Actinobacteria and Ascomycota in studied sites, respectively. Additionally, revegetation significantly increased the relative abundances of Proteobacteria, Gemmatimonadetes, Bacteroidetes and Patescibacteria, and decreased the relative abundance of Actinobacteria. Robinia pseudoacacia harbored the highest soil fungal community diversity, and bacterial Simpson index and Shannon index. Vegetation restoration with RP could clearly shifted soil communities compared to AM and PK. Along with the restoration of vegetation, the remarkable abiotic changes were the accumulation of total C, total N, total P, available P, available N and available K and the decreasing of soil pH, which were the most important factors affecting soil microbial communities. Our results addressed that Robinia pseudoacacia was the best preferable species than AM and PK in improving soil nutrients, soil community diversity and structure in Fe mining, providing a helpful guideline for selection of tree species.

Development of liposomal pemetrexed for enhanced therapy against multidrug resistance mediated by ABCC5 in breast cancer.

PURPOSE: Breast cancer is the most common cancer among women. Pemetrexed, a new generation antifolate drug, is one of the primary treatments for breast cancer. However, multidrug resistance (MDR) in breast cancer greatly hampers the therapeutic efficacy of chemotherapies such as pemetrexed. Nanomedicine is emerging as a promising alternative technique to overcome cancer MDR. Thus, pemetrexed-loaded d-alpha tocopheryl polyethylene glycol 1000 succinate (vitamin E TPGS) liposomes (liposomal pemetrexed) were developed as a strategy to overcome MDR to pemetrexed in breast cancer. MATERIALS AND METHODS: Liposomal pemetrexed was developed using the calcium acetate gradient method. The cytotoxic effects, apoptosis-inducing activity, in vivo distribution, and antitumor activity of liposomal pemetrexed were investigated. RESULTS: Liposomal pemetrexed was small in size (160.77 nm), with a small polydispersity of <0.1. The encapsulation efficacy of liposomal pemetrexed was 63.5%, which is rather high for water-soluble drugs in liposomes. The IC50 of liposomal pemetrexed following treatment with MDR breast cancer cells (MCF-7 cells overexpressing ABCC5) was 2.6-fold more effective than pemetrexed. The in vivo biodistribution study showed that the liposomes significantly accumulated in tumors 24 h after injection. The antitumor assay in mice bearing MDR breast cancer xenograft tumors confirmed the superior antitumor activity of liposomal pemetrexed over pemetrexed. It was also found that the improved therapeutic effect of liposomal pemetrexed may be attributed to apoptosis through both extrinsic and intrinsic pathways. CONCLUSION: Liposomal pemetrexed represents a potential therapeutic approach for overcoming breast cancer MDR.

Cationic liposomes induce cell necrosis through lysosomal dysfunction and late-stage autophagic flux inhibition.

AIM: The application of cationic liposomes (CLs) as nonviral vectors is hampered by their cellular toxicity. Thus we aim to investigate the mechanisms underlying the cellular toxicity of CLs. MATERIALS & METHODS: The effect of CLs on the autophagic flux, autophagosome-lysosome fusion, lysosome membrane permeabilization and cell necrosis of liver cells was investigated. RESULTS & CONCLUSION: Our results reveal a novel mechanism of CL-induced cell necrosis involving the induction of lysosome membrane permeabilization and late-stage autophagic flux inhibition that resulted in cytoplasmic release of cathepsin B, mitochondrial dysfunction and reactive oxygen species production, which are the key mediators of cell necrosis. Our study is important for revealing the cellular toxicity of CLs and designing safer gene delivery systems.

Interleukin-1ß Promoter Polymorphism Enhances the Risk of Sleep Disturbance in Alzheimer's Disease.

Sleep alleviates Alzheimer's disease (AD)-related neuropathological processes, whereas sleep disturbance in AD patients is associated with elevated peripheral inflammatory cytokine levels. In the present study, we assessed interleukin (IL)-1ß and APOEε4 polymorphisms for association with susceptibility of sleep disturbances in AD patients. A total of 123 pretreated AD patients and 120 age-, gender- and education level-matched healthy controls were recruited for two consecutive full-night polysomnography and measurement of Epworth Sleepiness Scale (ESS) scores for sleep-wake disturbance. Their genomic DNA was analyzed for IL-1ß and APOEε4 SNPs using ligase detection reaction (LDR) technology. Blood levels of IL-1ß, IL-6, and tumor necrosis factor alpha (TNF-α) were measured using ELISA after lipopolysaccharide (LPS) stimulation. The odds ratio and 95% confidence interval for genotype-specific risk were calculated using an unconditional logistic regression model and adjusted by age, gender, educational levels, body mass index (BMI), and activities of daily living (ADL). Compared to the non-APOEε4/ε4 genotype, APOEε4/ε4 significantly increased the risk of AD (APOEε4/ε4 vs. non-APOEε4/ε4, adjusted OR = 4.33, 95% CI = 1.33-14.10, p = 0.015). Compared to the IL-1ß CC genotype (-31), the TT genotype significantly increased the risk of AD (TT vs. CC, adjusted OR = 1.72, 95% CI = 1.13-2.61, p = 0.010). AD patients carrying the APOEε4 allele and the IL-1ß TT genotype showed less time in bed, longer sleep latency and REM latency, more awakenings, and a lower SWS percentage than those carrying CC/CT combined genotypes. In addition, blood IL-1ß levels were significantly greater in AD patients carrying both the APOEε4 allele and the IL-1ß-31TT genotype than in those carrying the APOEε4 allele and the -31 TC or CC genotype. In conclusion, this study provides the first evidence indicating that the IL-1ß-31TT genotype and homozygous APOEε4 combined are associated with increased risk of developing AD with sleep disturbance.

[Correlation of high-risk HPV 16/18 infections with prostate cancer].

OBJECTIVE: To study the correlation of high-risk human papillomavirus 16 and 18 (HPV16/18) infections with the risk of prostate cancer (PCa) and their association with the clinicopathologic indexes of PCa. METHODS: We collected tissue samples from 75 cases of PCa and 73 cases of benign prostatic hyperplasia (BPH). We detected HPV16/18 infections in the samples by immunohistochemistry and PCR combined with reverse dot blot (RDB) assay. RESULTS: Immunohistochemistry revealed 16 cases of HPV16/18 positive in the PCa (21.3%) and 7 cases in the BPH samples (9.5%), with statistically significant difference between the two groups (P=0.049). PCR combined with RDB assay showed 17 cases of HPV16 infection (22.6%) and 13 cases of HPV18 infection (17.8%), including 4 cases of HPV16/18 positive, in the PCa group, remarkably higher than 6 cases of HPV16 infection (8.2%), 3 cases of HPV18 infection (4.1%) and no HPV16/18 positive in the BPH controls (P=0.001). No significant differences were observed between the result of immunohistochemistry and that of PCR combined with RDB assay (P=0.069). The risk of HPV16/18 infections was found to be correlated with the clinical T-stage and Gleason score of PCa (P<0.05 ) but not with the patient's age, PSA level or lymph node metastasis (P>0.05 ). CONCLUSIONS: High-risk HPV16/18 infections are correlated with the risk of prostate cancer.

Heat shock protein 90 inhibition by 17-Dimethylaminoethylamino-17-demethoxygeldanamycin protects blood-brain barrier integrity in cerebral ischemic stroke.

Metalloproteinase (MMP)9 plays a pivotal role in ischemic stroke induced blood brain barrier (BBB) disruption. Correlation between HSP90 and MMP9 in several diseases prompted us to evaluate the efficacy of HSP90 inhibition as a novel approach to protect BBB integrity in ischemic stroke. ELISA was used to detect HSP90α and MMP9 in serum samples of stroke patients, which showed that HSP90α significantly correlated with MMP9 among 63 serum samples of stroke patients. Male C57/BL6 mice were pretreated with 17-Dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) or vehicle before being subjected to transient occlusion of middle cerebral artery and reperfusion (MCAO). Infarction, neurological scores, Evans blue (EB) extravasation, inflammatory responses and tight junction protein expression were examined 24 h after MCAO. We also investigated if 17-DMAG protected BBB integrity by suppressing inflammation and MMP9 activation. Oxygen glucose deprivation (OGD) was performed on bEnd.3 cells to explore the mechanisms of HSP90 inhibition in inhibiting MMP9. The results demonstrated that infarct volume was reduced in 17-DMAG-treated mice compared to control group following MCAO. Neurological outcomes were greatly improved in 17-DMAG-treated mice. Inflammatory responses, MMP9 activity and EB extravasation were decreased by 17-DMAG. In addition, 17-DMAG inhibited nuclear factor kappa B (NF-κB) activation following MCAO. Furthermore, HSP90 inhibition decreased NF-κB dependent MMP9 expression in bEnd.3 after OGD /reoxygenation. These findings suggested that HSP90 could be a novel therapeutic target in BBB breakdown during ischemic stroke. As several HSP90 inhibitors are in clinical trials for cancer, these findings have translational implications.

Identification and expression analysis of QM-like gene from Spodoptera litura after challenge by the entomopathogenic fungus Nomuraea rileyi.

A partial sequence of QM homologue was isolated from a Spodoptera litura fatbody suppression subtractive hybridization library. The full-length Spodoptera litura QM (SpLQM) cDNA of 838 bp contains a 5' untranslated region (UTR) of 112 bp, a 3' UTR of 66 bp, and an open reading frame of 660 nucleotides coding for a 219 amino acid peptide with a molecular weight of 25.5 kDa. Analysis of SpLQM sequence revealed the presence of characteristic motifs, including the ribosomal protein L10 signature and SH3-binding motif. Multiple alignment analysis revealed that SpLQM shares an overall identity of 57.1-99.1% with other members of QM family. Phylogenetic analysis confirmed that SpLQM is closely related to other insect QMs. Analysis of the tissue expression pattern showed that the SpLQM mRNA was expressed in all tissues tested, with highest levels measured in hemocytes, followed by fat bodies. Upon Nomuraea rileyi challenge, SpLQM showed significant upregulation in fat bodies and hemocytes, while slightly upregulation in midguts. The results suggest that SpLQM might play an important role in the innate immunity of S. litura in response to N. rileyi infection. SpLQM was also successfully overexpressed in Escherichia coli, and the recombinant fusion protein SpLQM-His has a molecular weight of 32 kDa.

Identification of suitable reference genes for gene expression studies by qRT-PCR in the blister beetle Mylabris cichorii.

The blister beetle Mylabris cichorii L. (Coleoptera: Meloidae) is a traditional medicinal insect recorded in the Chinese Pharmacopoeia. It synthesizes cantharidin, which kills cancer cells efficiently. Only males produce large amounts of cantharidin. Reference genes are required as endogenous controls for the analysis of differential gene expression in M. cichorii. Our study chose 10 genes as candidate reference genes. The stability of expression of these genes was analyzed by quantitative PCR and determined with two algorithms, geNorm and Normfinder. We recommend UBE3A and RPL22e as suitable reference genes in females and UBE3A, TAF5, and RPL22e in males.

Association of ABCC2 -24C>T polymorphism with high-dose methotrexate plasma concentrations and toxicities in childhood acute lymphoblastic leukemia.

Methotrexate (MTX) is a key agent for the treatment of childhood acute lymphoblastic leukemia (ALL). Increased MTX plasma concentrations are associated with a higher risk of adverse drug effects. ATP-binding cassette subfamily C member 2 (ABCC2) is important for excretion of MTX and its toxic metabolite. The ABCC2 -24C>T polymorphism (rs717620) reportedly contributes to variability of MTX kinetics. In the present study, we assessed the association between the ABCC2 -24C>T polymorphism and methotrexate (MTX) toxicities in childhood ALL patients treated with high-dose MTX. A total of 112 Han Chinese ALL patients were treated with high-dose MTX according to the ALL-Berlin-Frankfurt-Muenster 2000 protocol. Our results showed that presence of the -24T allele in ABCC2 gene led to significantly higher MTX plasma concentrations at 48 hours after the start of infusion, which would strengthen over repeated MTX infusion. The -24T allele in ABCC2 gene was significantly associated with higher risks of high-grade hematologic (leucopenia, anemia, and thrombocytopenia) and non-hematologic (gastrointestinal and mucosal damage/oral mucositis) MTX toxicities. This study provides the first evidence that the -24T allele in ABCC2 gene is associated with the severity of MTX toxicities, which add fresh insights into clinical application of high-dose MTX and individualization of MTX treatment.

Nur77 suppresses pulmonary artery smooth muscle cell proliferation through inhibition of the STAT3/Pim-1/NFAT pathway.

The orphan nuclear receptor 4A (NR4A) family plays critical roles in the regulation of cell proliferation, differentiation, and survival in the cardiovascular system. However, the molecular mechanisms underlying the regulation of NR4A receptor expression and its role in pulmonary artery smooth muscle cell (PASMC) function remain unclear. Here, we investigated whether the NR4A family regulates PASMC proliferation, and if so, which mechanisms are involved. By using quantitative real-time RT-PCR, we showed that the orphan nuclear receptor Nur77 was the most abundant member of NR4A family expressed in rat PASMCs, as compared with the two other members, NOR-1 and Nurr1. In rat PASMCs, expression of Nur77 was robustly induced in response to several pathologic stimuli of pulmonary arterial hypertension (PAH), such as hypoxia, 5-hydroxytryptamine (5-HT), platelet-derived growth factor, and endothelin-1. Importantly, Nur77 was also significantly increased in lungs of rats with monocrotaline-induced PAH. Furthermore, we demonstrated that 5-HT markedly up-regulated Nur77 expression through the mitogen-activated protein kinases/extracellular signal-regulated kinase 1/2 pathway. Overexpression of Nur77 inhibited 5-HT-induced PASMC proliferation, as well as the expression of cyclin D1 and proliferating cell nuclear antigen. Mechanistically, we demonstrated that Nur77 specifically interacts with signal transducer and activator of transcription 3, thus inhibiting its phosphorylation and expression of its target genes, such as Pim-1, nuclear factor of activated T cells c2, and survivin in PASMCs. These results indicate that Nur77 is a novel negative-feedback regulator of PASMC proliferation through inhibition of the signal transducer and activator of transcription 3/Pim-1/nuclear factor of activated T cells axis. Modulation of Nur77 activity may potentially represent a novel therapeutic strategy for the treatment of PAH.