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Retinal pigment epithelium (RPE) is an important component of the outer blood-retinal barrier and plays a critical role in maintaining retinal homeostasis. Alterations in RPE can be detected during the early stages of diabetic retinopathy (DR). However, the molecular mechanisms underlying these early changes remain unclear. We investigated the molecular changes induced in the RPE by high glucose concentrations by constructing a high glucose-induced ARPE-19 cell injury model simulating the DR environment in vitro. Proteomic analysis was conducted to measure differences in protein expression between cells treated with normal (5 mM) and high (25 mM) glucose concentrations, and bioinformatics techniques were used to analyze the mechanism of action. The results of the proteomic analyses were validated using western blotting. High glucose levels inhibited the proliferation of ARPE-19 cells. We identified 88 upregulated proteins and 114 downregulated proteins. Six of these proteins were selected for further validation. Changes in the proteome mainly affected the lysosome and cell cycle pathways. Proteomic differences between ARPE-19 cells treated with normal and high glucose concentrations indicate that damage to the RPE in DR may be caused by specific mechanisms. Our study verified protein changes in ARPE-19 cells in a high-glucose environment and may provide new strategies for understanding the molecular mechanisms underlying DR.

Introduction: It is known that the metabolic disorder caused by high glucose is one of pathogenesis in diabetic retinopathy (DR), the leading cause of blindness, due to the main pathological change of apoptosis of endothelial cells (ECs). In previous studies, the potential impact of sodium glucose cotransporter-2 (SGLT-2), whose inhibitors slow the progression of DR, has not been elucidated. The purpose of the presented study was to explore the effect of SGLT-2 inhibitors dapagliflozin (DAPA) on apoptosis of diabetic mice retina and human retinal microvascular endothelial cells (HRMECs), examine the effects of dapagliflozin on HRMECs metabolism, and explore the molecular processes that affect DR.Methods and Results: The eyeballs of male streptozotocin (STZ)-induced diabetic C57BL/6N mice were evaluated. C57BL/6N mice were divided into control group (CON), diabetic untreated group (DM), diabetic dapagliflozin treatment group (DM + DAPA) and diabetic insulin treatment group (DM + INS). Hematoxylin-Eosin (HE) staining was performed to observe the pathological structure of the mice retina, and TUNEL staining to detect apoptosis of mice retinal cells. In vitro, DCFH-DA and western blot (WB) were used to evaluate ROS, Bcl-2, BAX, cleaved-caspase 3 in HRMECs and metabolomics detected the effect of dapagliflozin on the metabolism of HRMECs. And then, we performed correlation analysis and verification functions for significantly different metabolites. In vivo, dapagliflozin reduced the apoptosis of diabetic mice retina independently of hypoglycemic. In vitro, SGLT-2 protein was expressed on HRMECs. Dapagliflozin reduced the level of ROS caused by high glucose, decreased the expression of cleaved-caspase3 and the ratio of BAX/Bcl-2. Metabolomics results showed that dapagliflozin did not affect the intracellular glucose level. Compared with the high glucose group, dapagliflozin reduced the production of arachidonic acid (AA) and inhibited the phosphorylation of ERK1/2, therefore, reducing the phosphorylation of cPLA2, which is a key enzyme for arachidonic acid release.Conclusion: Collectively, results unearthed for the first time that dapagliflozin reduced apoptosis of retina induced by DM whether in vivo or in vitro. Dapagliflozin did not affect the glucose uptake while mitigated intracellular arachidonic acid in HRMECs. Dapagliflozin alleviated HRMECs apoptosis induced by high glucose through ERK/1/2/cPLA2/AA/ROS pathway.

In various reactive oxygen species (ROS)‐based antitumor approaches (e.g., photodynamic therapy), increasing attentions are made to improve ROS level, but the short lifetime that is another decisive hurdle of ROS‐based antitumor outcomes is not even explored yet. To address it, a photocleaved O2‐released nanoplatform is constructed to release and switch ROS into reactive nitrogen species (RNS) for repressing hypoxic breast tumor. Systematic explorations validate that the nanoplatforms can attain continuous photocontrolled O2 release, alleviate hypoxia, and elevate ROS level. More significantly, the entrapped PDE5 inhibitor (PDE5‐i) in this nanoplatform can be enzymatically decomposed into nitric oxide that further combines with ROS to generate RNS, enabling the persistent antitumor effect since RNS features longer lifetime than ROS. Intriguingly, ROS conversion into RNS can help ROS to evade the hypoxia‐induced resistance to ROS‐based antitumor. Eventually, RNS production unlocks robust antitumor performances along with ROS elevation and hypoxia mitigation. Moreover, this extraordinary conversion from ROS into RNS also can act as a general method to solve the short lifetime of ROS.

Owing to a nanochannel-based enrichment effect and anti-fouling ability, highly ordered and vertically oriented mesoporous silica thin film (VMSF) modified electrodes have demonstrated their great potential in direct and highly sensitive analysis of complex samples. In this work, a VMSF modified fluorine-doped tin oxide (FTO) electrode (VMSF/FTO) is fabricated for enhanced electrochemiluminescence (ECL) analysis of lidocaine in serum. VMSF with good integrity and mechanical stability can be rapidly and conveniently grown on FTO in a few seconds at room temperature using an electrochemically assisted self-assembly (EASA) method. Due to the strong electrostatic attraction between the cationic ECL probe and negatively charged nanochannel, the VMSF/FTO electrode shows significant enrichment of tris(2,2-bipyridine) ruthenium(II) (Ru(bpy)32+), leading to ∼10 times enhancement of its ECL signal in comparison to the bare FTO electrode. Lidocaine, an anesthetic and antiarrhythmic drug, can act as the co-reactant of Ru(bpy)32+ and promote its ECL signal. Sensitive ECL detection of lidocaine is achieved by the sensor in a wide linear range from 10 nM to 50 μM with a low limit-of-detection (LOD) of 8 nM. Combined with the antifouling ability of VMSF, the VMSF/FTO electrode also realizes the accurate and rapid analysis of lidocaine in real serum samples.

SQUAMOSA Promoter Binding Protein (SBP) family genes act as central players to regulate plant growth and development with functional redundancy and specificity. Addressing the diversity of the SBP family in crops is of great significance to precisely utilize them to improve agronomic traits. Blueberry is an important economic berry crop. However, the SBP family has not been described in blueberry. In the present study, twenty VcSBP genes were identified through data mining against blueberry transcriptome databases. These VcSBPs could be clustered into eight groups, and the gene structures and motif compositions are divergent among the groups and similar within each group. The VcSBPs were differentially expressed in various tissues. Intriguingly, 10 VcSBPs were highly expressed at green fruit stages and dramatically decreased at the onset of fruit ripening, implying that they are important regulators during early fruit development. Computational analysis showed that 10 VcSBPs were targeted by miR156, and four of them were further verified by degradome sequencing. Moreover, their functional diversity was studied in Arabidopsis. Noticeably, three VcSBPs significantly increased chlorophyll accumulation, and qRT-PCR analysis indicated that VcSBP13a in Arabidopsis enhanced the expression of chlorophyll biosynthetic genes such as AtDVR, AtPORA, AtPORB, AtPORC, and AtCAO. Finally, the targets of VcSBPs were computationally identified in blueberry, and the Y1H assay showed that VcSBP13a could physically bind to the promoter region of the chlorophyll-associated gene VcLHCB1. Our findings provided an overall framework for individually understanding the characteristics and functions of the SBP family in blueberry.

Color change is an important event during fruit maturation in blueberry, usually depending on chlorophyll degradation and anthocyanin accumulation. MicroRNA156 (miR156)–SPL modules are an important group of regulatory hubs involved in the regulation of anthocyanin biosynthesis. However, little is known regarding their roles in blueberry or in chlorophyll metabolism during color change. In this study, a MIR156 gene (VcMIR156a) was experimentally identified in blueberry (Vaccinium corymbosum). Overexpression of VcMIR156a in tomato (Solanum lycopersicum) enhanced anthocyanin biosynthesis and chlorophyll degradation in the stem by altering pigment-associated gene expression. Further investigation indicated that the VcSPL12 transcript could be targeted by miR156, and showed the reverse accumulation patterns during blueberry fruit development and maturation. Noticeably, VcSPL12 was highly expressed at green fruit stages, while VcMIR156a transcripts mainly accumulated at the white fruit stage when expression of VcSPL12 was dramatically decreased, implying that VcMIR156a–VcSPL12 is a key regulatory hub during fruit coloration. Moreover, VcSPL12 decreased the expression of several anthocyanin biosynthetic and regulatory genes, and a yeast two-hybrid assay indicated that VcSPL12 interacted with VcMYBPA1. Intriguingly, expression of VcSPL12 significantly enhanced chlorophyll accumulation and altered the expression of several chlorophyll-associated genes. Additionally, the chloroplast ultrastructure was altered by the expression of VcMIR156a and VcSPL12. These findings provide a novel insight into the functional roles of miR156–SPLs in plants, especially in blueberry fruit coloration.

Objective.To evaluate contrast-enhanced ultrasound (CEUS) for monitoring early intrahepatic recurrence of primary hepatocellular carcinoma (HCC) after curative treatment.Methods.We prospectively analyzed 97 patients (124 nodules) with primary HCC who underwent hepatic resection or radiofrequency ablation and subsequently experienced intrahepatic recurrence. Patients were assessed with conventional ultrasound and CEUS. They were also assessed with contrast-enhanced computed tomography (CECT) and/or magnetic resonance imaging (MRI). The image characteristics of CEUS of recurrent hepatocellular carcinoma and high-grade dysplastic nodules (HGDNs) were analyzed. In addition, the ability of CEUS and CECT/MRI to assess internal artery vascularization in recurrent disease was compared.Results.CEUS of recurrent hepatocellular carcinoma showed hyperenhancement in the arterial phase in 96 of 99 nodules, and it showed hypo- or isoenhancement for portal venous and delayed phases. The most common enhancement patterns were “fast-in and slow-out” and “fast-in and fast-out”. Based on the arterial hyperenhancement of lesions and with clinical data such as patient history of HCC and increased level of serum alpha-fetoprotein, the diagnostic accuracy of CEUS for recurrent HCC was significantly higher than that based on the enhancement pattern of “fast-in and fast-out”. CEUS of HGDNs showed local or global hyperenhancement during the arterial phase, isoenhancement during the portal venous phase, and isoenhancement or slight hypoenhancement during the delayed phase. The enhancement pattern was “fast-in and slow-out”. In some cases, it was difficult to differentiate HGDNs from recurrent disease using CEUS. Vascularization in recurrent disease was significantly higher when assessed by CEUS than when assessed with CECT/MRI (P< 0.05). For detecting recurrent disease, CEUS showed sensitivity of 97.0%, specificity of 68.0%, positive predictive value of 92.3%, and negative predictive value of 85.0%. The corresponding parameters for CECT/MRI were 71.7%, 72.0%, 88.8%, and 39.1%.Conclusion.Intrahepatic recurrent HCC and HGDNs with diameter ≤ 3.0 cm have a characteristic appearance on CEUS. This imaging modality may be effective for monitoring early intrahepatic recurrence after curative treatment of primary HCC.

Aux/IAA genes are an important class of players in diverse developmental processes in plants, which generally exert their functions through the auxin signaling pathway. Blueberry is an economically and nutritionally important berry-bearing crop. However, Aux/IAA genes remain unknown in blueberry. In the present study, an Aux/IAA gene (VcIAA27) was identified and characterized in blueberry, and it is most closely related to IAA27 in other plant species. Expression analysis indicated that VcIAA27 transcripts accumulate highly in shoot, flower and fruit. Interestingly, VcIAA27 was highly expressed at early fruit developmental stages, and dramatically decreased from the onset of fruit ripening, implying that VcIAA27 possibly plays important roles during fruit enlargement. Meanwhile, the analysis of promoter activity in Arabidopsis showed that strong GUS signal was detected in the trichome and hydathodes of leaves, receptacle of silique, and lateral roots of seedling. Overexpression of VcIAA27 in Arabidopsis leads to auxin-related defects such as downward-curled leaves, short or sterile siliques, shorter stature, and more shoot branches. Moreover, qPCR analysis indicated that VcIAA27 is able to alter the expression patterns of the auxin-related genes BRU6, SAG13, SAUR26 in Arabidopsis, suggesting that VcIAA27 might be negatively involved in the auxin signaling pathway. The findings will greatly contribute to future investigation of Aux/IAA-mediated mechanisms that control blueberry development, especially fruit development and ripening.

4-Aminophenol (4-AP) served as the drug intermediates has been widely used or found in pharmaceutical preparations. Therefore, direct and rapid determination of 4-AP content in pharmaceuticals is highly important for human health. In this work, we demonstrate a simple and fast electrochemical method for direct analysis of 4-AP in pharmaceuticals by using the indium tin oxide (ITO) electrode modified with vertically-ordered mesoporous silica-nanochannel films (VMSF). Arising from the vertically-aligned nanochannels and high porosity of VMSF, 4-AP could permeate the VMSF and transfer to the underlying electrode surface, ultimately giving rise to the electrochemical signal. By using the differential pulse voltammetry (DPV) method, the VMSF/ITO sensor was able to detect 4-AP in a wide linear range from 0.5 μM to 400 μM. The obtained limit of detection was calculated as 0.32 μM. Owing to the good anti-fouling and anti-interference ability of VMSF, direct analysis of 4-AP in pharmaceuticals was achieved, without any complex sample pretreatments.

BACKGROUND Therapeutic evaluation of 3-dimensional conformal radiotherapy (3DCRT) is rarely reported for non-resectable hepatocellular carcinoma (HCC) with portal vein tumor thrombus (PVTT). The aim of this study was to determine the value of contrast-enhanced ultrasound (CEUS) in evaluating the therapeutic response of HCC with PVTT treated with 3DCRT. MATERIAL AND METHODS PVTT reduction rate in the study was determined after 3DCRT using time intensity curve (TIC) analysis software before and after radiotherapy. Seventy-nine HCC patients with PVTT treated with 3DCRT were studied. HCC and PVTT were performed by CEUS, before and after 3DCRT, over time. The parameters of blood flow, including arrival time (AT), time to peak (TTP), peak intensity (PI), washout time (WT), and area under the curve (AUC), were quantified and evaluated on still images by CEUS. RESULTS After 3DCRT, typing and staging of PVTT in 38 patients was decreased, the reduction rate was 48.1%. HCC was effective in 45 patients, the effective rate was 57%; No differences were found between the PVTT reduction rate and the HCC effective rate (χ2=2.96, P>0.05). In the effective group, the PI and AUC of HCCs and PVTTs after 3DCRT were significantly lower than before 3DCRT, while the other parameters of TIC were not significantly different before and after 3DCRT. CONCLUSIONS CEUS might be a useful monitoring option for the evaluation of HCC with PVTT treated with 3DCRT. CEUS might be useful as an important choice for monitoring and evaluation HCC with PVTT after 3DCRT. TIC parameters might provide quantitative data for efficacy evaluation, which helps to modify treatment strategies timely and accurately.

Background The purpose of this study was to investigate the values of contrast-enhanced ultrasound (CEUS) in the diagnosis and differential diagnosis of hyperechoic liver lesions. Material/Methods The CEUS findings of 102 patients with hyperechoic liver lesions identified by 2-dimensional ultrasound in the Affiliated Tumor Hospital of Guangxi Medical University were reviewed and analyzed. Results A total of 135 lesions were analyzed, of which malignant lesions were found in 72 patients and benign lesions in 63, with a CEUS accuracy rate of 91.11%, which was significantly higher than that of conventional ultrasound (74.81%; P

Hypoxia as one characteristic hallmark of solid tumors has been demonstrated to be involved in cancer metastasis and progression, induce severe resistance to oxygen-dependent therapies, and hamper the transportation of theranostic agents. To address these issues, an oxygen-self-produced sonodynamic therapy (SDT) nanoplatform involving a modified fluorocarbon (FC)-chain-mediated oxygen delivery protocol has been established to realize highly efficient SDT against hypoxic pancreatic cancer. In this nanoplatform, mesopores and FC chains of FC-chain-functionalized hollow mesoporous organosilica nanoparticle carriers can provide sufficient storage capacity and binding sites for sonosensitizers (IR780) and oxygen, respectively. In vitro and in vivo experiments demonstrate the nanoplatform involving this distinctive oxygen delivery protocol indeed breaks the hypoxia-specific transportation barriers, supplies sufficient oxygen to hypoxic PANC-1 cells especially upon exposure to ultrasound irradiation, and relieves hypoxia. Consequently, hypoxia-induced resistance to SDT is inhibited and sufficient highly reactive oxygen species (ROS) are produced to kill PANC-1 cells and shrink hypoxic PANC-1 pancreatic cancer. This distinctive FC-chain-mediated oxygen delivery method provides an avenue to hypoxia oxygenation and holds great potential in mitigating hypoxia-induced resistance to those oxygen-depleted therapies, e.g., photodynamic therapy, radiotherapy, and chemotherapy.

Background Human bocavirus (HBoV) is a novel parvovirus identified in 2005. It has mostly been detected in respiratory and enteric infections and has not been studied large scale in blood products in relation to transfusion. Study Design and Methods An in-house quantitative polymerase chain reaction (Q-PCR) was developed to test HBoV DNA in plasma and plasma derivatives. Plasma samples (n = 6096) collected from healthy donors, 241 plasma pools, and 326 plasma derivatives were screened for HBoV DNA by Q-PCR. Positive samples were confirmed by nested PCR and further amplified for sequence analysis and phylogenetic studies. The prevalence of immunoglobulin (Ig)G and IgM specific to HBoV structural proteins was measured by enzyme-linked immunosorbent assay in 209 samples grouped according to virus load (Group 1, HBoV DNA >104 copies/mL; Group 2, HBoV DNA >5 × 102 copies/mL but below 104 copies/mL; Group 3,HBoV DNA negative). Results The genomic prevalence of HBoV in the plasma donors was 9.06%, ranging from 5.01 × 102 to 3.02 × 106 copies/mL. HBoV-specific IgG and IgM were detected at 20.00 and 7.50% in Group 1, at 20.29 and 2.90% in Group 2, and at 13.00 and 4.0% in Group 3, respectively. Phylogenetic analyses proved that HBoV Genotype 1 was the prevalent genotype in Chinese plasma donors. Conclusion Low levels of HBoV DNA were detectable at high prevalence in Chinese plasma donors and plasma derivatives. Further study is needed to determine whether HBoV screening is necessary.

MicroRNAs (miRNAs) play vital roles in the regulation of fruit development and ripening. Blueberry is an important small berry fruit crop with economical and nutritional value. However, nothing is known about the miRNAs and their targets involved in blueberry fruit ripening. In this study, using high-throughput sequencing of small RNAs, 84 known miRNAs belonging to 28 families and 16 novel miRNAs were identified in white fruit (WF) and blue fruit (BF) libraries, which represent fruit ripening onset and in progress, respectively. Among them, 41 miRNAs were shown to be differentially expressed during fruit maturation, and 16 miRNAs representing 16 families were further chosen to validate the sRNA sequencing data by stem-loop qRT-PCR. Meanwhile, 178 targets were identified for 41 known and 7 novel miRNAs in WF and BF libraries using degradome sequencing, and targets of miR160 were validated using RLM-RACE (RNA Ligase-Mediated (RLM)-Rapid Amplification of cDNA Ends) approach. Moreover, the expression patterns of 6 miRNAs and their targets were examined during fruit development and ripening. Finally, integrative analysis of miRNAs and their targets revealed a complex miRNA-mRNA regulatory network involving a wide variety of biological processes. The findings will facilitate future investigations of the miRNA-mediated mechanisms that regulate fruit development and ripening in blueberry.

Numerous postoperative therapies for preventing recurrence of hepatocellular carcinoma (HCC) have been reported, but their efficacy remains controversial and knowledge about adverse effects is limited. A systematic review of randomized controlled trials (RCTs) was performed to gain a comprehensive picture of the efficacy and risks of these therapies.MEDLINE, EMBASE and the Cochrane Library were systematically searched through July 2011. Risk ratios (RRs) and 95% confidence intervals (CIs) were calculated.A total of 2989 patients from 28 RCTs involving 10 postoperative therapies were included. For interferon therapy, the estimated RR for the 2-year recurrence rate was 0.84 (95% CI 0.73-0.97, P = 0.02) and the overall survival (OS) was 1.15 (95% CI 1.07-1.22, P0.001). Postoperative therapy with the vitamin K2 analog did not lead to a significant reduction in the 1-year recurrence rate, with a pooled RR of 0.60 (95% CI 0.28-1.27, P = 0.18). However, it did slightly improve the 1-year OS, with a pooled RR of 1.03 (95% CI 1.00-1.05, P = 0.03). Transarterial chemotherapy with or without embolization, adoptive immunotherapy and heparanase inhibitor PI-88 therapy may delay tumor recurrence. The effects of acyclic retinoid, lipiodol-iodine-131 and tumor vaccine treatment were promising but require further study. All postoperative therapies except interferon administered intramuscularly were well tolerated by the majority of patients.Use of adjuvant interferon is definitely associated with an increase in OS. Postoperative therapies involving acyclic retinoid, lipidol-iodine-131, or tumor vaccine may improve the OS of patients with HCC after curative treatment.