Effects of Neuroinflammation and Autophagy on the Structure of the Blood-Brain Barrier in ADHD Model.

Spontaneously hypertensive rats (SHR) are the most common animal model used to study attention deficit hyperactivity disorder (ADHD). The purpose of this study was to look at the impact of neuroinflammation and autophagy on blood-brain barrier function in the prefrontal cortex and hippocampus of ADHD rats. The rats were separated into three groups: juvenile SHR (6 weeks), mature SHR (12 weeks), and comparable age WKY groups. An open-field test was used to assess rats' ability to move on their own. Immunofluorescence was used to detect the Iba1-immunopositive microglia, ZO-1 and TNF-α. Meanwhile, the expression of p62, Beclin-1, LC3B, and MMP9, MMP2, TNF-α, ZO-1, and occludin were detected by Western blot. The results have shown that Iba1-immunopositive microglia and TNF-α protein in the brain of SHR rats were significantly increased. Moreover, autophagy of cells and the level of MMP2 and MPP9 in the prefrontal cortex and hippocampus increased in SHR rats. In addition, the expression of ZO-1 and occludin was decreased in SHR rats. To sum up, the increase of neuroinflammation and excessive autophagy were essential factors for the damage of blood-brain barrier structure and function.

FBXW7 loss of function promotes esophageal squamous cell carcinoma progression via elevating MAP4 and ERK phosphorylation.

BACKGROUND: Increasing evidence suggests that FBXW7 has a high frequency of mutations in esophageal squamous cell carcinoma (ESCC). However, the function of FBXW7, especially the mutations, is not clear. This study was designed to investigate the functional significance of FBXW7 loss of function and underlying mechanism in ESCC. METHODS: Immunofluorescence was applied to clarify the localization and main isoform of FBXW7 in ESCC cells. Sanger sequencing were performed to explore mutations of FBXW7 in ESCC tissues. Proliferation, colony, invasion and migration assays were performed to examine the functional roles of FBXW7 in ESCC cells in vitro and in vivo. Real-time RT-PCR, immunoblotting, GST-pulldown, LC-MS/MS and co-immunoprecipitation assay were used to explore the molecular mechanism underlying the actions of FBXW7 functional inactivation in ESCC cells. Immunohistochemical staining were used to explore the expression of FBXW7 and MAP4 in ESCC tissues. RESULTS: The main FBXW7 isoform in ESCC cells was the ß transcript in the cytoplasm. Functional inactivation of FBXW7 led to activation of the MAPK signaling pathway and upregulation of the downstream MMP3 and VEGFA, which enhanced tumor proliferation cell invasion and migration. Among the five mutation forms screened, S327X (X means truncated mutation) had an effect similar to the FBXW7 deficiency and led to the inactivation of FBXW7 in ESCC cells. Three other point mutations, S382F, D400N and R425C, attenuated but did not eliminate FBXW7 function. The other truncating mutation, S598X, which was located outside of the WD40 domain, revealed a tiny attenuation of FBXW7 in ESCC cells. Notably, MAP4 was identified as a potential target of FBXW7. The threonine T521 of MAP4, which was phosphorylated by CHEK1, played a key role in the FBXW7-related degradation system. Immunohistochemical staining indicated that FBXW7 loss of function was associated with tumor stage and shorter survival of patients with ESCC. Univariate and multivariate Cox proportional hazards regression analyses showed that high FBXW7 and low MAP4 was an independent prognostic indicator and prospective longer survival. Moreover, a combination regimen that included MK-8353 to inhibit the phosphorylation of ERK and bevacizumab to inhibit VEGFA produced potent inhibitory effects on the growth of FBXW7 inactivation xenograft tumors in vivo. CONCLUSIONS: This study provided evidence that FBXW7 loss of function promoted ESCC via MAP4 overexpression and ERK phosphorylation, and this novel FBXW7/MAP4/ERK axis may be an efficient target for ESCC treatment.

Influence of preoperative simulation on the reduction quality and clinical outcomes of open reduction and internal fixation for complex proximal humerus fractures.

PURPOSE: Proximal humerus fractures (PHFs) are common. With the development of locking plates, open reduction and internal fixation (ORIF) of the proximal humerus can provide excellent clinical outcomes. The quality of fracture reduction is crucial in the locking plate fixation of proximal humeral fractures. The purpose of this study was to determine the impact of 3-dimensional (3D) printing technology and computer virtual technology assisted preoperative simulation on the reduction quality and clinical outcomes of 3-part and 4-part proximal humeral fractures. METHOD: A retrospective comparative analysis of 3-part and 4-part PHFs undergoing open reduction internal fixation was performed. Patients were divided into 2 groups according to whether computer virtual technology and 3D printed technology were used for preoperative simulation: the simulation group and the conventional group. Operative time, intraoperative bleeding, hospital stay, quality of fracture reduction, Constant scores, American Society for Shoulder and Elbow Surgery (ASES) scores, shoulder range of motion, complications, and revision surgeries were assessed. RESULTS: This study included 67 patients (58.3%) in the conventional group and 48 patients (41.7%) in the simulation group. The patient demographics and fracture characteristics were comparable in these groups. Compared with the conventional group, the simulation group had shorter operation time and less intraoperative bleeding (P < 0.001, both). Immediate postoperative assessment of fracture reduction showed a higher incidence of greater tuberosity cranialization of < 5 mm, neck-shaft angle of 120° to 150°, and head shaft displacement of < 5 mm in the simulation group. The incidence of good reduction was 2.6 times higher in the simulation group than in the conventional group (95% CI, 1.2-5.8). At the final follow-up, the chance of forward flexion > 120° (OR 5.8, 95% CI 1.8-18.0) and mean constant score of > 65 (OR 3.4, 95% CI 1.5-7.4) was higher in the simulation group than the conventional group, as well as a lower incidence of complications in the simulation group was obtained (OR 0.2, 95% CI 0.1-0.6). CONCLUSIONS: This study identified that preoperative simulation assisted by computer virtual technology and 3D printed technology can improve reduction quality and clinical outcomes in treatment of 3-part and 4-part PHFs.

Construction of graphene quantum dots ratiometric fluorescent probe by intermolecular electron transfer effect for intelligent and real-time visual detection of ofloxacin and its L-isomer in daily drink.

The design of intelligent and real-time sensing devices is significant in the medical drug monitoring field, but it is still highly challenging. Here, ratiometric fluorescent detections of ofloxacin (OFL) and its L-isomer levofloxacin (LEV) constructed from tri-doped graphene quantum dots (T-GQDs) are reported, and the detection limits reach as low as 46/67 nM toward OFL/LEV due to the intermolecular electron transfer (intermolecular ET) effect. After adding OFL/LEV, the generation of electrostatic bond provides a channel for the intermolecular ET from the edge of T-GQDs to OFL/LEV, resulting in the fluorescence quenching at 414 nm and the fluorescence promoting at 498 nm. Furthermore, a smartphone can be used for the visual and quantitative detection of OFL and LEV by identifying the RGB values of test paper and drink samples. This work not only reveals the physics mechanism of ratiometric detection, but also develops a convenient smartphone diagnostic for OFL and LEV.

Promising energy transfer system between fuorine and nitrogen Co-doped graphene quantum dots and Rhodamine B for ratiometric and visual detection of doxycycline in food.

A novel sensor based on dual emissive fluorescent graphene quantum dots is developed for a rapid, selective, sensitive and visual detection of doxycycline (DOX). The ratiometric fluorescent probe is designed by grafting fluorescent group (Rhodamine B, RhB) on F, N-doped graphene quantum dots (FNGQDs). In the presence of DOX, the fluorescence at 466 nm is remarkably quenched due to inner filter effect and fluorescence resonance energy transfer, whereas the peak at 592 nm is attenuated slightly due to the energy transfer in the emission peaks of FNGQDs and RhB functional group. The sensor shows good linear relationship from 0.04 to 100 µM with a low detection limit of 40 nM. Furthermore, the flexible solid-state fluorescent sensing platform is used for detecting DOX in milk, pork and water samples. Therefore, this dual-emission FGQD-RhB can be used as a high-performance fluorescent and visual sensor for food safety and environmental monitoring.

Mechanisms behind multicolor tunable Near-Infrared triple emission in graphene quantum dots and ratio fluorescent probe for water detection.

Developing environmental-friendly and low-cost strategies of synthesizing red-emission graphene quantum dots (R-GQDs) is a considerable challenge. Herein, we present the green and facile preparation of R-GQDs by using o-phenylenediamine (o-PD) and catechol as precursors via oxidation/polymerization and Schiff base reaction at low temperature (80 °C) for 3 hrs. Results show that the prepared R-GQDs exhibit triple fluorescence emissions, which is enabled by the introduce of different nitrogen (pyrrolic N, pyridinic N, and amino N) species on the surface of R-GQDs. Moreover, the R-GQDs are implemented to detect the moisture in different organic solvent. A highly sensitive ratiometric sensing of water in organic solvents is achieved, and the relationship between the change of fluorescence signal caused by moisture and the corresponding internal emission site is also determined. In the end, the multicolor light emissions of R-GQDs are realized by simply adjusting the polarity of surrounding solvents. And based on the solvatochromism of R-GQDs, the multicolor solid fluorescent powder and ink are prepared for illumination application. All in all, the above research work presents a novel R-GQDs for wide application in detecting and illumination.

A smartphone-integrated optical sensing platform based on Lycium ruthenicum derived carbon dots for real-time detection of Ag.

Growing global environmental pollution problems challenge the need for converting biomass into an advantageous product. In this paper, Lycium ruthenicum is successfully turned into beneficial green emissive (527 nm) fluorescent nitrogen doping carbon dots (N-CDs) via the hydrothermal treatment for the first time. The horizontal and vertical dimensions of N-CDs are demonstrated to be about 4.5 and 0.73 nm, respectively. The N-CDs possess an extremely stable green fluorescence and quantum yield up to 21.8%. Meaningfully, N-CDs exhibit a good linear relationship with Ag+ in the range of 0.7-36 µM, and its detection limit is determined to be 59 nM. The practicability of the fluorescent probe is further validated in lake water and the satisfactory spiked recoveries of Ag+ ranges from 98.99% to 104.19%. Besides, based on the sensitive and selective photoluminescence quenching properties, a smartphone-based laboratory device and RGB analysis software are used to directly capture and analyze fluorescence images with a sensitive detection limit of 83 nM for Ag+. This novel sensor based on N-CDs and smartphone provides a reliable way for on-site monitoring of Ag+ and expands application prospect in the field of environmental pollution detection.

PVDF-triggered multicolor fluorine-doped graphene quantum dots for water detection and anti-counterfeiting.

Fluorescent fluorine-doped graphene quantum dots (F-GQDs) have been synthesized via the hydrothermal method using long-chain polymer polyvinylidene fluoride (PVDF) as the precursor. Due to the unique molecular structure of PVDF, a possible synthesis process of F-GQDs has been put forward. F-GQDs have adjustable emission wavelength by simply adjusting the concentration of the solution. As the concentration increases, the emission wavelength of F-GQDs gradually red shifts from 455 nm (blue) to 551 nm (yellow-green). In addition, F-GQDs also exhibit a sensitive fluorescence response to water content in organic solvents, and the ultralow detections limit are 0.056% in ethanol and 0.124% in DMF. Besides, due to strong UV absorption capacity, a photothermal film is fabricated by embedding F-GQDs in PDMS. The temperature of F-GQDs/PDMS polymer film can reach 33.4 oC under simulated sunlight, while the maximum temperature of blank PDMS film only reach 29.4 oC. Based on this phenomenon, a new type of anti-counterfeiting device is designed by combining F-GQDs/PDMS film with temperature change ink.

Synthesis of multi-color fluorine and nitrogen co-doped graphene quantum dots for use in tetracycline detection, colorful solid fluorescent ink, and film.

Fluorine-doped graphene quantum dots have unique chemical bonds and charge distribution, which can bring unexpected properties compared to other common atom-doped graphene quantum dots. In the present work, fluorine and nitrogen co-doped graphene quantum dots (F, N-GQDs) are synthesized from levofloxacin via a simple hydrothermal method. Systematic studies demonstrate that F, N-GQDs can emit various fluorescence with the wavelength ranging from blue to green by dispersing F, N-GQDs into different solvents. Moreover, multi-color fluorescence is available by simply changing the concentration of F, N-GQDs. In addition to these unique characteristics, F, N-GQDs also exhibit a sensitive fluorescence response to tetracycline with an ultralow detection limit of 77 nM in water. Because of high photostability and high quantum yield, the F, N-GQDs are exploited as a unique invisible ink, which is printable and writable on paper. Meanwhile, based on the solvatochromism of F, N-GQDs, we realized the color adjustable fluorescent ink. Finally, large-area flexible multi-color fluorescent films are realized. Our synthesized F, N-GQDs, with tunable fluorescence in wavelength and intensity, have numerous opportunities for optical molecular sensors, information security, flexible optics, and others.

Circulating microRNA let­7e is decreased in knee osteoarthritis, accompanied by elevated apoptosis and reduced autophagy.

Knee osteoarthritis (KOA) is a major cause of leg disability in the elderly population. Recently, the expression levels of circulating microRNA (miRNA) let­7e have been reported to be significantly reduced in KOA. The aims of the present study were to assess the feasibility of let­7e as a serum marker for detecting KOA and to explore the underlying mechanisms of its involvement. Based on previous studies and bioinformatics analysis, let­7e may regulate apoptosis and autophagy of articular chondrocytes. A total of 10 patients with KOA and 10 patients with trauma without KOA were recruited to examine the levels of let­7e in peripheral blood. Subsequently, KOA rat models were established, and the levels of let­7e in the cartilage and serum were examined, the expression of apoptotic proteins and autophagy­related proteins in the cartilage were investigated, and apoptotic and autophagic activities of primary cultured chondrocytes were also detected. In patients with KOA, let­7e levels in the peripheral serum were significantly decreased compared with the control group, and this result was confirmed in the peripheral serum and cartilage of KOA rats. In addition, the expression levels of proteins involved in the apoptotic pathway were increased in the cartilage of KOA rats, and apoptotic activity was increased. The expression of autophagy­related proteins beclin 1 and microtubule associated protein 1 light chain 3 ß (LC3B) II/LC3BI in the articular cartilage of KOA rats was lower compared with the controls, and autophagy was decreased. Si­Miao­San (SMS) treatment restored the expression of let­7e and reversed the changes in apoptosis and autophagy. Therefore, the present study provided additional evidence that circulating let­7e may be a potential serum biomarker for the diagnosis and treatment of KOA. Elevated apoptosis levels and decreased autophagy levels of cartilage tissue are involved in KOA, and treatment with SMS may reverse these effects.

Intra-articular nodular fasciitis: a rare lesion case report and an updated review of the literature.

BACKGROUND: Nodular fasciitis is a benign proliferation of myofibroblasts that usually arises in subcutaneous tissues of the trunk, neck, head, and upper extremities of young adults. It is not reported to arise in the joints. CASE PRESENTATION: In this report, we describe a rare case where nodular fasciitis occurred in an intra-articular location in the right knee of a 20-year-old man. The patient presented with 3-months' duration of knee pain without history of trauma to the extremity. Physical examination revealed pain, joint effusion, and limited range of motion (ROM) of the affected knee. Magnetic resonance imaging (MRI) showed a 2.5 × 2 × 1 cm lesion in front of the posterior cruciate ligament. Arthroscopically, the soft tissue mass was removed and pathologically diagnosed as a rare, benign, intra-articular nodular fasciitis. Symptoms resolved 1 month after the operation and no recurrence was found at the 6 months follow-up. CONCLUSION: The present paper describes detailed characteristics of intra-articular nodular fasciitis and provides an updated comprehensive summary of 21 prior case reports.

Retrovirus-Mediated Transfection of the Tissue-type Plasminogen Activator Gene Results in Increased Thrombolysis of Blood Clots.

Tissue-type plasminogen activator (tPA) is involved in the lysis of blood clots. In this study, we attempted to target thrombolysis and enhance blood clot lysis by generating a construct (pLEGFP-N1-tPA) to integrate tPA gene into the genome of different cell lines. pLEGFP-N1-tPA construct was generated and used to target the tPA gene in different cell lines. The thrombolytic effects mediated by the supernatant from transfected HeLa cells and Linx cells were assessed using plasma thrombus plates. Furthermore, enhanced green fluorescent protein (EGFP), which was fused to the tPA gene in the pLEGFP-N1-tPA construct, was analyzed under the fluorescent microscope to assess tPA localization. We also monitored tPA activity and expression in the transfected cell lines. As part of the study, we successfully generated the pLEGFP-N1-tPA construct. The sequence of this construct was verified and the construct was subsequently used to generate the PT67/pLEGFP-N1-tPA cell line. The pLEGFP-N1-tPA construct was also used to transfect HeLa cells and Linx cells. We observed that supernatants from transfected cells were capable of lysing thrombi. In addition, tPA activity and tPA concentration were elevated in the latter supernatants and tPA was rapidly and stably expressed in the transfected cell lines. These results reveal a potentially important thrombolytic role for tPA-targeted gene therapy following cardiac valve replacement.

The post-therapeutic effect of rapamycin in mild traumatic brain-injured rats ensuing in the upregulation of autophagy and mitophagy.

Mild traumatic brain injury (mTBI), common in juveniles, has been reported to be caused by sports-related concussion. Many young children may suffer from post-concussion syndrome. mTBI, in early stages of life, could play a part in neuron apoptosis and degeneration, cognitive and motor coordination impairment, as well as dementia. Our study was aimed at further investigating the post-therapeutic efficacy of rapamycin in the recuperation of mTBI while at the same time investigating the metamorphosis in both autophagy and mitophagy in mTBI. We created a weight-drop rat mTBI model with the administration of rapamycin at 4 h after every mTBI. Behavioral tests of beam walking and open field task indicated the expected improvement of cognitive and motor coordination functions. Both Western blot and immunofluorescence examinations revealed increased Beclin-1 and PINK1 in the treated rats as well as reduction of caspase-3 and cytochrome C (Cyt C). More so, the TUNEL staining evidenced curtailment of apoptotic cells following treatment with rapamycin. The upregulation of Beclin-1 and PINK1 and the downregulation of caspase-3 and Cyt C extrapolate that rapamycin plays neuroprotective as well as anti-apoptotic role via interposition of both autophagy and mitophagy.

Improved Neural Regeneration with Olfactory Ensheathing Cell Inoculated PLGA Scaffolds in Spinal Cord Injury Adult Rats.

BACKGROUND/AIMS: Every year, around the world, between 250000 and 500000 people suffer from spinal cord injury (SCI). This study investigated the potential for poly (lactic-co-glycolic acid) (PLGA) complex inoculated with olfactory ensheathing cells (OECs) to treat spinal cord injury in a rat model. METHODS: OECs were identified by immunofluorescence based on the nerve growth factor receptor (NGFR) p75. The Basso, Beattie, and Bresnahan (BBB) score, together with an inclined plane (IP) test were used to detect functional recovery. Nissl staining along with the luxol fast blue (LFB) staining were independently employed to illustrate morphological alterations. More so, immunofluorescence labeling of the glial fibrillary acidic protein (GFAP) and the microtubule-associated protein-2 (MAP-2), representing astrocytes and neurons respectively, were investigated at time points of weeks 2 and 8 post-operation. RESULTS: The findings showed enhanced locomotor recovery, axon myelination and better protected neurons post SCI when compared with either PLGA or untreated groups (P < 0.05). CONCLUSION: PLGA complexes inoculated with OECs improve locomotor functional recovery in transected spinal cord injured rat models, which is most likely due to the fact it is conducive to a relatively benevolent microenvironment, has nerve protective effects, as well as the ability to enhance remyelination, via a promotion of cell differentiation and inhibition of astrocyte formation.

Puerarin attenuates inflammation and oxidation in mice with collagen antibody-induced arthritis via TLR4/NF-κB signaling.

Puerarin is an important active ingredient in the root of kudzu vine due to its pharmacological properties. The aim of the present study is to contribute to the existing knowledge of the effect of puerarin in the attenuation of inflammation and oxidation in mice with collagen antibody-induced arthritis via toll­like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling. Arthritis was induced using injection of anti­type II collagen antibodies. Treatment with puerarin was observed to significantly decrease clinical scoring of the collagen antibody­induced arthritis and suppress oxidative stress and the inflammatory response in mice. Furthermore, puerarin was demonstrated to inhibit mRNA expression of matrix metalloproteinase­9 and protein expression of TLR4 following collagen antibody-induced arthritis in mice. The effect of puerarin may be associated with the suppression of NF­κB activity in collagen antibody­induced arthritis mice. Furthermore, upregulation of phosphorylated (p)­Janus kinase 2 and p­signal transducer and activator of transcription 3 protein expression was suppressed by puerarin. The results of the present study indicate, for the first time, the effect of puerarin to attenuate inflammation and oxidation in mice with collagen antibody­induced arthritis via TLR4/NF-κB signaling.

Estrogen receptor alpha inhibits RLR-mediated immune response via ubiquitinating TRAF3.

RIG-I-like receptors (RLRs) function as key sentinel receptor for invading viruses. Moderate activation of RLR signaling is critical for efficient viral clearance without harmful immunopathology. Estrogen receptor alpha (ERα) is a member of the nuclear receptor superfamily of ligand-activated transcription factors and is involved in the regulation of innate immune responses. However, the effects of ERα on RLR signaling and the molecular mechanisms are poorly understood. In this study, we identify ERα as a negative regulator of RLR-triggered antiviral immune responses. The expression level of ERα is upregulated following RLR activation in macrophages. In the absence of ligand, VSV infection phosphorylates ERα at serine 167. ERα inhibits VSV-induced IRF3 activation. We further demonstrate that ERα directly interacts with TRAF3 and promotes K48-linked proteasomal degradation of TRAF3. Consistently, ERα inhibits VSV-triggered IFN-ß production in macrophages in a ligand independent mechanism. Thus, ERα functions as a negative feedback regulator of RLR-triggered antiviral immune responses. These findings also provide the insights that separate the immune effects of ERα from its ligand-induced hormonal effects.

Gastric foregut cystic developmental malformation: case series and literature review.

Foregut cystic developmental malformation (FCDM) is a very rare lesion of the alimentary tract, especially in the stomach. We discuss the concepts of gastric duplication cyst, bronchogenic cysts, and FCDM. Nomenclature has been inconsistent and confusing, but, by some definitions, gastric duplication cysts involve gastric mucosa and submucosal glands, bronchogenic cysts involve respiratory mucosa with underlying cartilage and glands, and FCDM lacks gastric mucosa or underlying glands or cartilage but has pseudostratified ciliated columnar epithelium (PCCE). We searched our departmental case files from the past 15 years and identified 12 cases of FCDM in the alimentary tract. We summarize the features of these 12 cases including a report in detail on a 52-year-old man with a submucosal cyst lined with simple PCCE and irregular and stratified circular muscle layers that merged with gastric smooth muscle bundles near the lesser curvature of the gastric cardia. A literature review of cases with this histology yielded 25 cases. We propose the term gastric-FCDM for such cases. Our own series of 12 cases confirms that preoperative recognition of the entity is infrequent and problematic. The rarity of this developmental disorder, as well as a lack of understanding of its embryologic origins, may contribute to missing the diagnosis. Not appreciating the diagnosis preoperatively can lead to an inappropriate surgical approach. In contrast, presurgical recognition of the entity will contribute to a good outcome and reduced risk of complications.

Effect of the timing of surgery on the fracture healing process and the expression levels of vascular endothelial growth factor and bone morphogenetic protein-2.

The aim of the present study was to observe the effect of varying the timing of surgery on the fracture healing process and the expression levels of vascular endothelial growth factor (VEGF) and bone morphogenetic protein (BMP)-2 in rats. A total of 192 rats underwent closed femur fracture modelling. The rats underwent open reduction and internal fixation surgery 1, 3, 5, 7, 11 and 14 days subsequent to the fracture occurring. Immunohistochemical staining and analysis of the VEGF and BMP-2 expression levels were simultaneously conducted on bone from the fracture site of the rats on various days. The VEGF and BMP-2 expression levels at the fracture sites were higher and were maintained for a longer period of time in the 7- and 11-day surgery groups than in the other surgery groups and the rats that did not undergo surgery. The 5-day surgery group demonstrated a greater intensity in BMP-2 expression compared with the remaining surgery groups; however, no significant differences were identified between 1-day surgery and non surgery groups. In the 3-day surgery group, the expression levels VEGF and BMP-2 were low at each stage of the fracture-healing process and were lower compared with those observed in the non-surgery group. The timing of the surgical procedures affected the VEGF and BMP-2 expression levels at the fracture sites of the experimental rats and, the optimal time for performing surgery was identified to be within the first two weeks. However, surgery may not be conducive to fracture healing if it is performed within the first few days following fracture.