Upon contact with biofluids, proteins are quickly adsorbed onto the nanoparticle (NP) surface to form a protein corona, which initiates the opsonization and facilitates the rapid clearance of the NP ...by macrophage uptake. Although polyethylene glycol (PEG) functionalization has been the standard approach to evade macrophage uptake by reducing protein adsorption, it cannot fully eliminate nonspecific uptake. Herein, polyglycerol (PG) grafting is demonstrated as a better alternative to PEG. NPs of various size and material were grafted with PG and PEG at 30, 20, and 10 wt % contents by controlling the reaction conditions, and the resulting NP-PG and NP-PEG were characterized qualitatively by IR spectroscopy and quantitatively by thermogravimetric analysis. Their resistivity to adsorption of the proteins in fetal bovine serum and human plasma were compared by polyacrylamide gel electrophoresis, bicinchoninic acid assay, and liquid chromatography-tandem mass spectrometry, giving a consistent conclusion that PG shields protein adsorption more efficiently than does PEG. The macrophage uptake was assayed by transmission electron microscopy and by extinction spectroscopy or inductively coupled plasma mass spectrometry, revealing that PG avoids macrophage uptake more efficiently than does PEG. In particular, a NP coated with PG at 30 wt % (NP-PG-h) prevents corona formation almost completely, regardless of NP size and core material, leading to the complete evasion of macrophage uptake. Our findings demonstrate that PG grafting is a promising strategy in nanomedicine to improve anti-biofouling property and stealth efficiency in nanoformulations.
Lewy body disease (LBD) is a spectrum of progressive neurodegenerative disorders characterized by the wide distribution of Lewy bodies and neurites in the central and peripheral nervous system (CNS, ...PNS). Clinical diagnoses include Parkinson’s disease (PD), dementia with Lewy bodies, or pure autonomic failure. All types of LBD are accompanied by non-motor symptoms (NMSs) including gastrointestinal dysfunctions such as constipation. Its relationship to Lewy body-related α-synucleinopathy (Lewy pathology) of the enteric nervous system (ENS) is attracting attention because it can precede the motor symptoms. To clarify the role of ENS Lewy pathology in disease progression, we performed a clinicopathological study using the Brain Bank for Aging Research in Japan. Five-hundred and eighteen cases were enrolled in the study. Lewy pathology of the CNS and PNS, including the lower esophagus as a representative of the ENS, was examined via autopsy findings. Results showed that one-third of older people (178 cases, 34%) exhibited Lewy pathology, of which 78 cases (43.8%) exhibited the pathology in the esophagus. In the esophageal wall, Auerbach’s plexus (41.6%) was most susceptible to the pathology, followed by the adventitia (33.1%) and Meissner’s plexus (14.6%). Lewy pathology of the esophagus was significantly associated with autonomic failures such as constipation (
p
< 0.0001) and among PNS regions, correlated the most with LBD progression (
r
= 0.95,
p
< 0.05). These findings suggest that the propagation of esophageal Lewy pathology is a predictive factor of LBD.
Thioredoxin interacting protein (Txnip) is an α-arrestin protein that regulates pleiotropic biological responses. Txnip acts as a cancer suppressor and is a critical regulator of energy metabolism. ...To investigate molecular mechanisms involving Txnip, we searched for its protein binding partners using tandem affinity purification and proteomics analyses and identified several viable candidates, including HSP90, HSP70, and Prp31. We showed, by native PAGE, that Txnip is involved in the formation of high molecular weight complexes (1000–1300 kDa) in the nuclear fraction of cells treated with glucose and bortezomib. DTT treatment partly dissolved these high molecular weight complexes, suggesting that Txnip forms redox sensitive high-order nucleoprotein complexes. RNAse treatment slightly decreased the complex and RNA-seq showed differential expression of RNAs in the complex between Txnip protein overexpressing and control cells, indicating the involvement of RNAs in the complex. These results collectively provide a model whereby Txnip exerts its functions through multiple binding partners, forming transient higher-order complexes to regulate other signaling molecules.
•Proteomic analyses showed HSP90, HSP70, and Prp31 as Txnip binding partners.•Txnip forms a high molecular weight complex (1000–1300 kDa).•The high molecular complex contains disulfide bonds and RNAs.•Differential expression of RNAs in the complex between Txnip and control cells.
Epstein-Barr virus-associated gastric cancer (EBVaGC) has been reported to be associated with chronic inflammation of the gastric epithelium caused by Helicobacter pylori infection. Autoimmune ...gastritis (AIG) is also believed to increase the risk of carcinogenesis. We herein report a case of multiple EBVaGCs that arose in a patient with AIG, highlighting the potential for multiplicity of this entity. In this case, a total of four metachronous EBVaGCs were found after initial Endoscopic submucosal dissection for EBVaGC, all of which were treated endoscopically. This case demonstrates that patients with AIG should be monitored closely for development of EBVaGC.
Hemocyanin (Hc) and phenoloxidase (PO) are members of the type 3 copper protein family. Although arthropod Hc and PO exhibit similar three-dimensional structures of the copper-containing active site, ...Hc functions as an oxygen transport protein, showing minimal or no phenoloxidase activity. Here, we present the crystal structure of the oxy form of Hc from Panulirus japonicus (PjHc) at 1.58 Å resolution. The structure of the di-copper active site of PjHc was found to be almost identical to that of PO. Although conserved amino acids and the water molecule crucial for the enzymatic activity were observed in PjHc at almost the same positions as those in PO, PjHc showed no enzymatic activity under our experimental conditions. One striking difference between PjHc and arthropod PO was the presence of a “blocker residue” near the binuclear copper site of PjHc. This blocker residue comprised a phenylalanine residue tightly stacked with an imidazole ring of a CuA coordinated histidine and hindered substrates from accessing the active site. Our results suggest that the blocker residue is also a determining factor of the catalytic activity of type 3 copper proteins.
•Hemocyanin and phenoloxidase are two major members of the type 3 copper proteins.•We determined the high-resolution crystal structure of hemocyanin from a crustacean.•The structure of its active site is almost identical to that of phenoloxidases.•However, hemocyanin has an active site blocker residue that is absent in phenoloxidase.•The blocker residue would cause the functional difference between these type 3 copper proteins.
Cytoplasmic DNA activates cyclic GMP-AMP synthase (cGAS) to produce cyclic 2'-5'3'-5'GMP-AMP dinucleotide (2'5 'cGAMP). The binding of 2'5'cGAMP to an adaptor protein, stimulator of IFN genes ...(STING), activates a transcription factor, IFN regulatory factor 3, leading to the induction of IFN and chemokine gene expression. In this study, we found that the 2'5'cGAMP-dependent STING activation induced highly upregulated CXCL10 gene expression. Formation of a distinct STING dimer, which was detected by native PAGE, was induced by 2'5'cGAMP, but not 3'-5'3'-5'cGAMP. Analysis of DNase II(-/-) mice, which constitutively produce IFN-β and CXCL10, showed the accumulation of 2'5'cGAMP in their fetal livers and spleens, suggesting that the undigested DNA accumulating in DNase II(-/-) cells may have leaked from the lysosomes into the cytoplasm. The DNase II(-/-) mouse embryonic fibroblasts produced 2'5'cGAMP in a cGAS-dependent manner during apoptotic cell engulfment. However, cGAS deficiency did not impair the STING-dependent upregulation of CXCL10 in DNase II(-/-) mouse embryonic fibroblasts that was induced by apoptotic cell engulfment or DNA lipofection. These results suggest the involvement of a cGAS-independent additional DNA sensor(s) that induces the STING-dependent activation of innate immunity.
Idiopathic normal pressure hydrocephalus usually exhibits triad of symptoms including gait disturbance, urinary incontinence, and dementia with ventriculomegaly. Currently, its pathogenesis remains ...to be fully elucidated.
To provide a better understanding of this order, we examined whether dysmetabolism of sphingolipids as major lipid components in the brain present in cerebrospinal fluid (CSF) of the patients. Here, we measured various sphingolipidsincluding ceramide and sphingomyelin and glycolipids by electrospray ionization-tandem mass spectrometry in the cerebrospinal fluid of 19 consecutive idiopathic normal pressure hydrocephalus patients, 49 Parkinson's disease patients, and 17 neurologically normal controls.
The data showed that there was a significant and specific reduction of all galactosylceramide subspecies levels in idiopathic normal pressure hydrocephalus patients compared with other groups, whereas ceramide and sphingomyelin levels as well as other neutral glycolipids such as glucosylceramide and lactosylceramide were similar in both disease states. Multiple regression analysis of sex and age did not show any correlation with galactosylceramide levels. We also examined whether MMSE scores are correlated with sphingolipid levels in iNPH patients. A specific subspecies of sphingomyelin (d18:1/18:0) only exhibited a statistically significant negative correlation (p = 0.0473, R = −0.4604) with MMSE scores but no other sphingolipids in iNPH patients. These data strongly suggest that myelin-rich galactosylceramide metabolism is severely impaired in idiopathic normal pressure hydrocephalus patients and might serve as the basis of biomarker for this disorder.
•Pathogenesis of idiopathic normal pressure hydrocephalus (iNPH) is currently unknown.•Although clinical diagnostic criteria have been available, its excellent surrogate marker is still unavailable.•Brain white matter is usually involved in iNPH patients.•All galactosylceramide subspecies in CSF are severely reduced in iNPH compared with Parkinson's disease and normal subjects.•Whereas ceramides and sphingomyelin as well as other neutral glycolipids were unaltered.
Iron deficiency and iron overload are among the most prevalent nutritional disorders worldwide. Duodenal cytochrome b (DcytB) and divalent metal transporter 1 (DMT1) are regulators of iron ...absorption. Their expression is increased during high systemic requirements for iron, but the molecular mechanisms that regulate DcytB and DMT1 expression are undefined. Hypoxia-inducible factor (HIF) signaling was induced in the intestine following acute iron deficiency in the duodenum, resulting in activation of DcytB and DMT1 expression and an increase in iron uptake. DcytB and DMT1 were demonstrated as direct HIF-2α target genes. Genetic disruption of HIF signaling in the intestine abolished the adaptive induction of iron absorption following iron deficiency, resulting in low systemic iron and hematological defects. These results demonstrate that HIF signaling in the intestine is a critical regulator of systemic iron homeostasis.
Many types of molecular targeted drugs that inhibit cancer growth by acting on specific molecules have been developed. The runt‐related transcription factor (RUNX) family, which induces cancer ...development by binding to a specific DNA sequence, has attracted attention as a new target for cancer treatment. We have developed Chb‐M′, which targets the RUNX‐binding sequence. Chb‐M′ was developed by conjugating pyrrole‐imidazole (PI) polyamides and chlorambucil as an anticancer agent. It was recently reported that Chb‐M′ had a remarkable anticancer effect in vivo. In this study, to explore the possibility of an alternative structure, we designed a new series of CBI–PI polyamides, in which seco‐CBI was applied as a DNA‐alkylating agent. We examined the characteristics of the CBI–PI polyamides targeting the RUNX‐binding sequence and found that these conjugates have great potential for cancer treatment.
A better warhead: The DNA‐alkylating mechanism of seco‐CBI–pyrrole‐imidazole (CBI–PI) polyamide conjugates, which inhibit cancer proliferation by targeting the RUNX‐binding sequence 5′‐TGTGGT‐3′, were chemically examined.
In patients with neurofibromatosis type 1 (NF1), it is important to accurately determine when plexiform neurofibroma (pNF) transforms to a malignant peripheral nerve sheath tumor (MPNST). The purpose ...of this study is to investigate the usefulness of diffusion-weighted imaging (DWI) in differentiating pNF and MPNST in NF1 patients.
Among the NF1 patients who were referred to our hospital between 1985 and 2015, 10 cases of MPNST and 19 cases of pNF were included. We evaluated features of standard magnetic resonance imaging according to the differentiation criteria of malignancy from benignancy as previously reported, apparent diffusion coefficient (ADC) value based on the DWI and the correlation between ADC value and benignancy/malignancy. ROC analysis was performed to determine the appropriate cutoff value of ADC.
There were significant differences between MPNST and pNF in the size of the tumor (P = 0.009), peripheral enhancement pattern (P = 0.002), perilesional edema-like zone (P = 0.0008), and intratumoral cystic change (P = 0.02). The mean and minimum values of ADC were significantly lower in MPNST than those in pNF (P = 0.03 and P = 0.003, respectively). When we set a cutoff value of mean ADC as 1.85 × 10–3 mm2/s, the sensitivity and specificity were 80% and 74%, respectively. The area under the curve value improved by adding the Wasa score to the mean ADC evaluation.
ADC values determined by DWI are useful in differentiating MPNST from pNF and adding ADC evaluation to standard MRI evaluation improved the diagnostic accuracy.
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