β-Conglycinin Reduces the Tight Junction Occludin and ZO-1 Expression in IPEC-J2

Soybean allergy presents a health threat to humans and animals. The mechanism by which food/feed allergen β-conglycinin injures the intestinal barrier has not been well understood. In this study, the changes of epithelial permeability, integrity, metabolic activity, the tight junction (TJ) distribution and expression induced by β-conglycinin were evaluated using IPEC-J2 model. The results showed a significant decrease of trans-epithelial electrical resistance (TEER) (p < 0.001) and metabolic activity (p < 0.001) and a remarkable increase of alkaline phosphatase (AP) activity (p < 0.001) in a dose-dependent manner. The expression levels of tight junction occludin and ZO-1 were decreased (p < 0.05). The reduced fluorescence of targets and change of cellular morphology were recorded. The tight junction occludin and ZO-1 mRNA expression linearly declined with increasing β-conglycinin (p < 0.001).     

Wnt3a Promotes the Vasculogenic Mimicry Formation of Colon Cancer via Wnt/β-Catenin Signaling

Our previous study provided evidence that non-canonical Wnt signaling is involved in regulating vasculogenic mimicry (VM) formation. However, the functions of canonical Wnt signaling in VM formation have not yet been explored. In this study, we found the presence of VM was related to colon cancer histological differentiation (p < 0.001), the clinical stage (p < 0.001), and presence of metastasis and recurrence (p < 0.001). VM-positive colon cancer samples showed increased Wnt3a expression (p < 0.001) and β-catenin nuclear expression (p < 0.001) compared with the VM-negative samples. In vitro, over-regulated Wnt3a expression in HT29 colon cancer cells promoted the capacity to form tube-like structures in the three-dimensional (3-D) culture together with increased expression of endothelial phenotype-associated proteins such as VEGFR2 and VE-cadherin. The mouse xenograft model showed that Wnt3a-overexpressing cells grew into larger tumor masses and formed more VM than the control cells. In addition, the Wnt/β-catenin signaling antagonist Dickkopf-1(Dkk1) can reverse the capacity to form tube-like structures and can decrease the expressions of VEGFR2 and VE-cadherin in Wnt3a-overexpressing cells. Taken together, our results suggest that Wnt/β-catenin signaling is involved in VM formation in colon cancer and might contribute to the development of more accurate treatment modalities aimed at VM.     

Fenofibrate Regulates Visceral Obesity and Nonalcoholic Steatohepatitis in Obese Female Ovariectomized C57BL/6J Mice

Fibrates, including fenofibrate, are a class of hypolipidemic drugs that activate peroxisome proliferator-activated receptor α (PPARα), which in-turn regulates the expression of lipid and lipoprotein metabolism genes. We investigated whether fenofibrate can reduce visceral obesity and nonalcoholic fatty liver disease via adipose tissue PPARα activation in female ovariectomized (OVX) C57BL/6J mice fed a high-fat diet (HFD), a mouse model of obese postmenopausal women. Fenofibrate reduced body weight gain (−38%, p < 0.05), visceral adipose tissue mass (−46%, p < 0.05), and visceral adipocyte size (−20%, p < 0.05) in HFD-fed obese OVX mice. In addition, plasma levels of alanine aminotransferase and aspartate aminotransferase, as well as free fatty acids, triglycerides, and total cholesterol, were decreased. Fenofibrate also inhibited hepatic lipid accumulation (−69%, p < 0.05) and infiltration of macrophages (−72%, p < 0.05), while concomitantly upregulating the expression of fatty acid β-oxidation genes targeted by PPARα and decreasing macrophage infiltration and mRNA expression of inflammatory factors in visceral adipose tissue. These results suggest that fenofibrate inhibits visceral obesity, as well as hepatic steatosis and inflammation, in part through visceral adipose tissue PPARα activation in obese female OVX mice.     

Different Role of Tumor Necrosis Factor-α Polymorphism in Non-Hodgkin Lymphomas among Caucasian and Asian Populations: A Meta-Analysis

Tumor necrosis factor-α (TNF-α) is an immunoregulatory cytokine involved in B- and T-cell function, and also plays an important role in inflammation and cancer. TNF-α-308G>A has been associated with constitutively elevated TNF-α expression. Several studies have reported the association between the TNF-α-308G>A polymorphism and non-Hodgkin lymphomas (NHL) risk, however, results are still inconsistent. To solve these conflicts, we conducted the first meta-analysis to assess the effect of TNF-α-308G>A polymorphism on the risk of NHL and various subtypes (additive model) including 10,619 cases and 12,977 controls in Caucasian and Asian populations. Our meta-analysis indicated that TNF-α-308G>A polymorphism is not associated with NHL risk when pooling all studies together (OR = 1.06, 95% CI: 0.92–1.23, p = 0.413). In stratified analyses, we found TNF-α-308A allele was significantly associated with higher risk of NHL, B-cell lymphomas (BCL), T-cell lymphomas (TCL) and diffuse large B-cell lymphomas (DLBCL) in Caucasians (OR = 1.22, 95% CI: 1.06–1.40, p = 0.007; OR = 1.18, 95% CI: 1.03–1.34, p = 0.014; OR = 1.20, 95% CI: 1.01–1.42, p = 0.040; OR = 1.21, 95% CI: 1.11–1.32, p < 0.001, respectively). Interestingly, it was associated with decreased risk of NHL, BCL and DLBCL in Asians (OR = 0.75, 95% CI: 0.66–0.86, p < 0.001; OR = 0.70, 95% CI: 0.52–0.94, p = 0.018; OR = 0.70, 95% CI: 0.57–0.86, p = 0.001). These findings also suggest TNF-α might play a distinct role in pathogenesis of NHL in different populations.     

Telmisartan Attenuates Kanamycin-Induced Ototoxicity in Rats

Telmisartan (TM) has been proposed to relieve inflammatory responses by modulating peroxisome proliferator activator receptor-γ (PPARγ) signaling. This study aimed to investigate the protective effects of TM on kanamycin(KM)-induced ototoxicity in rats. Forty-eight, 8-week-old female Sprague Dawley rats were divided into four groups: (1) control group, (2) TM group, (3) KM group, and (4) TM + KM group. Auditory brainstem response was measured. The histology of the cochlea was examined. The protein expression levels of angiotensin-converting enzyme 2 (ACE2), HO1, and PPARγ were measured by Western blotting. The auditory threshold shifts at 4, 8, 16, and 32 kHz were lower in the TM + KM group than in the KM group (all p < 0.05). The loss of cochlear outer hair cells and spiral ganglial cells was lower in the TM + KM group than in the KM group. The protein expression levels of ACE2, PPARγ, and HO1 were higher in the KM group than in the control group (all p < 0.05). The TM + KM group showed lower expression levels of PPARγ and HO1 than the KM group.TM protected the cochlea from KM-induced injuries in rats. TM preserved hearing levels and attenuated the increase in PPARγ and HO1 expression levels in KM-exposed rat cochleae.     

(De)stabilization of Alpha-Synuclein Fibrillary Aggregation by Charged and Uncharged Surfactants

Parkinson’s disease (PD) is the second most common neurodegenerative disorder. An important hallmark of PD involves the pathological aggregation of proteins in structures known as Lewy bodies. The major component of these proteinaceous inclusions is alpha (α)-synuclein. In different conditions, α-synuclein can assume conformations rich in either α-helix or β-sheets. The mechanisms of α-synuclein misfolding, aggregation, and fibrillation remain unknown, but it is thought that β-sheet conformation of α-synuclein is responsible for its associated toxic mechanisms. To gain fundamental insights into the process of α-synuclein misfolding and aggregation, the secondary structure of this protein in the presence of charged and non-charged surfactant solutions was characterized. The selected surfactants were (anionic) sodium dodecyl sulphate (SDS), (cationic) cetyltrimethylammonium chloride (CTAC), and (uncharged) octyl β-D-glucopyranoside (OG). The effect of surfactants in α-synuclein misfolding was assessed by ultra-structural analyses, in vitro aggregation assays, and secondary structure analyses. The α-synuclein aggregation in the presence of negatively charged SDS suggests that SDS-monomer complexes stimulate the aggregation process. A reduction in the electrostatic repulsion between N- and C-terminal and in the hydrophobic interactions between the NAC (non-amyloid beta component) region and the C-terminal seems to be important to undergo aggregation. Fourier transform infrared spectroscopy (FTIR) measurements show that β-sheet structures comprise the assembly of the fibrils.     

Activation of Apoptosis in a βB1-CTGF Transgenic Mouse Model

To reveal the pathomechanisms of glaucoma, a common cause of blindness, suitable animal models are needed. As previously shown, retinal ganglion cell and optic nerve degeneration occur in βB1-CTGF mice. Here, we aimed to determine possible apoptotic mechanisms and degeneration of different retinal cells. Hence, retinae were processed for immunohistology (n = 5–9/group) and quantitative real-time PCR analysis (n = 5–7/group) in 5- and 10-week-old βB1-CTGF and wildtype controls. We noted significantly more cleaved caspase 3⁺ cells in βB1-CTGF retinae at 5 (p = 0.005) and 10 weeks (p = 0.02), and a significant upregulation of Casp3 and Bax/Bcl2 mRNA levels (p < 0.05). Furthermore, more terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL⁺) cells were detected in transgenic mice at 5 (p = 0.03) and 10 weeks (p = 0.02). Neurofilament H staining (p = 0.01) as well as Nefh (p = 0.02) and Tubb3 (p = 0.009) mRNA levels were significantly decreased at 10 weeks. GABAergic synapse intensity was lower at 5 weeks, while no alterations were noted at 10 weeks. The glutamatergic synapse intensity was decreased at 5 (p = 0.007) and 10 weeks (p = 0.01). No changes were observed for bipolar cells, photoreceptors, and macroglia. We conclude that apoptotic processes and synapse loss precede neuronal death in this model. This slow progression rate makes the βB1-CTGF mice a suitable model to study primary open-angle glaucoma.     

Decreased Levels of Microfibril-Associated Glycoprotein (MAGP)-1 in Patients with Colon Cancer and Obesity Are Associated with Changes in Extracellular Matrix Remodelling

Objective: The protein microfibril-associated glycoprotein (MAGP)-1 constitutes a crucial extracellular matrix protein. We aimed to determine its impact on visceral adipose tissue (VAT) remodelling during obesity-associated colon cancer (CC). Methods: Samples obtained from 79 subjects (29 normoponderal (NP) (17 with CC) and 50 patients with obesity (OB) (19 with CC)) were used in the study. Circulating concentrations of MAGP-1 and its gene expression levels (MFAP2) in VAT were analysed. The impact of inflammation-related factors and adipocyte-conditioned media (ACM) on MFAP2 mRNA levels in colon adenocarcinoma HT-29 cells were further analysed. The effects of MAGP-1 in the expression of genes involved in the extracellular matrix (ECM) remodelling and tumorigenesis in HT-29 cells was also explored. Results: Obesity (p < 0.01) and CC (p < 0.001) significantly decreased MFAP2 gene expression levels in VAT whereas an opposite trend in TGFB1 mRNA levels was observed. Increased mRNA levels of MFAP2 after the stimulation of HT-29 cells with lipopolysaccharide (LPS) (p < 0.01) and interleukin (IL)-4 (p < 0.01) together with a downregulation (p < 0.05) after hypoxia mimicked by CoCl₂ treatment was observed. MAGP-1 treatment significantly enhanced the mRNA levels of the ECM-remodelling genes collagen type 6 α3 chain (COL6A3) (p < 0.05), decorin (DCN) (p < 0.01), osteopontin (SPP1) (p < 0.05) and TGFB1 (p < 0.05). Furthermore, MAGP-1 significantly reduced (p < 0.05) the gene expression levels of prostaglandin-endoperoxide synthase 2 (COX2/PTGS2), a key gene controlling cell proliferation, growth and adhesion in CC. Interestingly, a significant decrease (p < 0.01) in the mRNA levels of MFAP2 in HT-29 cells preincubated with ACM from volunteers with obesity compared with control media was observed. Conclusion: The decreased levels of MAGP-1 in patients with obesity and CC together with its capacity to modulate key genes involved in ECM remodelling and tumorigenesis suggest MAGP-1 as a link between AT excess and obesity-associated CC development.     

Supplementing with L-Tryptophan Increases Medium Protein and Alters Expression of Genes and Proteins Involved in Milk Protein Synthesis and Energy Metabolism in Bovine Mammary Cells

The objective of this study was to investigate the effects of supplementing with L-tryptophan (L-Trp) on milk protein synthesis using an immortalized bovine mammary epithelial (MAC-T) cell line. Cells were treated with 0, 0.3, 0.6, 0.9, 1.2, and 1.5 mM of supplemental L-Trp, and the most efficient time for protein synthesis was determined by measuring cell, medium, and total protein at 0, 24, 48, 72, and 96 h. Time and dose tests showed that the 48 h incubation time and a 0.9 mM dose of L-Trp were the optimal values. The mechanism of milk protein synthesis was elucidated through proteomic analysis to identify the metabolic pathway involved. When L-Trp was supplemented, extracellular protein (medium protein) reached its peak at 48 h, whereas intracellular cell protein reached its peak at 96 h with all L-Trp doses. β-casein mRNA gene expression and genes related to milk protein synthesis, such as mammalian target of rapamycin (mTOR) and ribosomal protein 6 (RPS6) genes, were also stimulated (p < 0.05). Overall, there were 51 upregulated and 59 downregulated proteins, many of which are involved in protein synthesis. The results of protein pathway analysis showed that L-Trp stimulated glycolysis, the pentose phosphate pathway, and ATP synthesis, which are pathways involved in energy metabolism. Together, these results demonstrate that L-Trp supplementation, particularly at 0.9 mM, is an effective stimulus in β-casein synthesis by stimulating genes, proteins, and pathways related to protein and energy metabolism.     

Cloning,Expression and Characterization of a Thermostable Esterase HydS14 from Actinomadura sp. Strain S14 in Pichia pastoris

A thermostable esterase gene (hydS14) was cloned from an Actinomadura sp. S14 gene library. The gene is 777 bp in length and encodes a polypeptide of 258 amino acid residues with no signal peptide, no N-glycosylation site and a predicted molecular mass of 26,604 Da. The encoded protein contains the pentapeptide motif (GYSLG) and catalytic triad (Ser88-Asp208-His235) of the esterase/lipase superfamily. The HydS14 sequence shows 46%–64% identity to 23 sequences from actinomycetes (23 α/β-hydrolases), has three conserved regions, and contains the novel motif (GY(F)SLG), which distinguishes it from other clusters in the α/β-hydrolase structural superfamily. A plasmid containing the coding region (pPICZαA-hydS14) was used to express HydS14 in Pichia pastoris under the control of the AOXI promoter. The recombinant HydS14 collected from the supernatant had a molecular mass of ~30 kDa, which agrees with its predicted molecular mass without N-glycosylation. HydS14 had an optimum temperature of approximately 70 °C and an optimum pH of 8.0. HydS14 was stable at 50 and 60 °C for 120 min, with residual activities of above 80% and above 90%, respectively, as well as 50% activity at pH 6.0–8.0 and pH 9.0, respectively. The enzyme showed higher activity with p-nitrophenyl-C2 and C4. The K_m and V_max values for p-nitrophenyl-C4 were 0.21 ± 0.02 mM and 37.07 ± 1.04 μmol/min/mg, respectively. The enzyme was active toward short-chain p-nitrophenyl ester (C2–C6), displaying optimal activity with p-nitrophenyl-C4 (K_cat/K_m = 11.74 mM⁻¹·S⁻¹). In summary, HydS14 is a thermostable esterase from Actinomadura sp. S14 that has been cloned and expressed for the first time in Pichia pastoris.     

Assessment of Amyloid Forming Tendency of Peptide Sequences from Amyloid Beta and Tau Proteins Using Force-Field,Semi-Empirical,and Density Functional Theory Calculations

A wide variety of neurodegenerative diseases are characterized by the accumulation of protein aggregates in intraneuronal or extraneuronal brain regions. In Alzheimer’s disease (AD), the extracellular aggregates originate from amyloid-β proteins, while the intracellular aggregates are formed from microtubule-binding tau proteins. The amyloid forming peptide sequences in the amyloid-β peptides and tau proteins are responsible for aggregate formation. Experimental studies have until the date reported many of such amyloid forming peptide sequences in different proteins, however, there is still limited molecular level understanding about their tendency to form aggregates. In this study, we employed umbrella sampling simulations and subsequent electronic structure theory calculations in order to estimate the energy profiles for interconversion of the helix to β-sheet like secondary structures of sequences from amyloid-β protein (KLVFFA) and tau protein (QVEVKSEKLD and VQIVYKPVD). The study also included a poly-alanine sequence as a reference system. The calculated force-field based free energy profiles predicted a flat minimum for monomers of sequences from amyloid and tau proteins corresponding to an α-helix like secondary structure. For the parallel and anti-parallel dimer of KLVFFA, double well potentials were obtained with the minima corresponding to α-helix and β-sheet like secondary structures. A similar double well-like potential has been found for dimeric forms for the sequences from tau fibril. Complementary semi-empirical and density functional theory calculations displayed similar trends, validating the force-field based free energy profiles obtained for these systems.     

Suppression of Hypoxia-Inducible Factor 1α by Low-Molecular-Weight Heparin Mitigates Ventilation-Induced Diaphragm Dysfunction in a Murine Endotoxemia Model

Mechanical ventilation (MV) is required to maintain life for patients with sepsis-related acute lung injury but can cause diaphragmatic myotrauma with muscle damage and weakness, known as ventilator-induced diaphragm dysfunction (VIDD). Hypoxia-inducible factor 1α (HIF-1α) plays a crucial role in inducing inflammation and apoptosis. Low-molecular-weight heparin (LMWH) was proven to have anti-inflammatory properties. However, HIF-1α and LMWH affect sepsis-related diaphragm injury has not been investigated. We hypothesized that LMWH would reduce endotoxin-augmented VIDD through HIF-1α. C57BL/6 mice, either wild-type or HIF-1α–deficient, were exposed to MV with or without endotoxemia for 8 h. Enoxaparin (4 mg/kg) was administered subcutaneously 30 min before MV. MV with endotoxemia aggravated VIDD, as demonstrated by increased interleukin-6 and macrophage inflammatory protein-2 levels, oxidative loads, and the expression of HIF-1α, calpain, caspase-3, atrogin-1, muscle ring finger-1, and microtubule-associated protein light chain 3-II. Disorganized myofibrils, disrupted mitochondria, increased numbers of autophagic and apoptotic mediators, substantial apoptosis of diaphragm muscle fibers, and decreased diaphragm function were also observed (p < 0.05). Endotoxin-exacerbated VIDD and myonuclear apoptosis were attenuated by pharmacologic inhibition by LMWH and in HIF-1α–deficient mice (p < 0.05). Our data indicate that enoxaparin reduces endotoxin-augmented MV-induced diaphragmatic injury, partially through HIF-1α pathway inhibition.     

Isolation and Self-Association Studies of Beta-Lactoglobulin

The aim of this study was to investigate isolated β-lactoglobulin (β-LG) from the whey protein isolate (WPI) solution using the column chromatography with SP Sephadex. The physicochemical characterization (self-association, the pH stability in various salt solutions, the identification of oligomeric forms) of the protein obtained have been carried out. The electrophoretically pure β-LG fraction was obtained at pH 4.8. The fraction was characterized by the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/TOF MS) technique. The use of the HCCA matrix indicated the presence of oligomeric β-LG forms, while the SA and DHB matrices enabled the differentiation of A and B isoforms in the sample. The impact of sodium chloride, potassium chloride, ammonium sulfate, and sodium citrate in dispersion medium on β-LG electrophoretic stability in solution was also studied. Type of the dispersion medium led to the changes in the isoelectric point of protein. Sodium citrate stabilizes protein in comparison to ammonium sulfate. Additionally, the potential of capillary electrophoresis (CE) with UV detection using bare fused capillary to monitor β-LG oligomerization was discussed. Obtained CE data were further compared by the asymmetric flow field flow fractionation coupled with the multi-angle light scattering detector (AF4-MALS). It was shown that the β-LG is a monomer at pH 3.0, dimer at pH 7.0. At pH 5.0 (near the isoelectric point), oligomers with structures from dimeric to octameric are formed. However, the appearance of the oligomers equilibrium is dependent on the concentration of protein. The higher quantity of protein leads to the formation of the octamer. The far UV circular dichroism (CD) spectra carried out at pH 3.0, 5.0, and 7.0 confirmed that β-sheet conformation is dominant at pH 3.0, 5.0, while at pH 7.0, this conformation is approximately in the same quantity as α-helix and random structures.     

A20 Overexpression Inhibits Lipopolysaccharide-Induced NF-κB Activation,TRAF6 and CD40 Expression in Rat Peritoneal Mesothelial Cells

Zinc finger protein A20 is a key negative regulator of inflammation. However, whether A20 may affect inflammation during peritoneal dialysis (PD)-associated peritonitis is still unclear. This study was aimed to investigate the effect of A20 overexpression on lipopolysaccharide (LPS)-induced inflammatory response in rat peritoneal mesothelial cells (RPMCs). Isolated and cultured RPMCs in vitro. Plasmid pGEM-T easy-A20 was transfected into RPMCs by Lipofectamine™2000. The protein expression of A20, phospho-IκBα, IκBα, TNF receptor-associated factor (TRAF) 6 and CD40 were analyzed by Western blot. The mRNA expression of TRAF6, CD40, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were determined by real time-PCR. NF-κB p65 DNA binding activity, IL-6 and TNF-α levels in cells culture supernatant were determined by ELISA. Our results revealed that RPMCs overexpression of A20 lead to significant decrease of LPS-induced IκBα phosphorylation and NF-κB DNA binding activity (all p < 0.01). In addition, A20 also attenuated the expression of TRAF6, CD40, IL-6 and TNF-α as well as levels of IL-6 and TNF-α in cells culture supernatant (all p < 0.05). However, A20 only partly inhibited CD40 expression. Our study indicated that A20 overexpression may depress the inflammatory response induced by LPS in cultured RPMCs through negatively regulated the relevant function of adaptors in LPS signaling pathway.     

Construction of IL-13 Receptor α2-Targeting Resveratrol Nanoparticles against Glioblastoma Cells: Therapeutic Efficacy and Molecular Effects

Glioblastoma multiforme (GBM) is the most common lethal primary brain malignancy without reliable therapeutic drugs. IL-13Rα2 is frequently expressed in GBMs as a molecular marker. Resveratrol (Res) effectively inhibits GBM cell growth but has not been applied in vivo because of its low brain bioavailability when administered systemically. A sustained-release and GBM-targeting resveratrol form may overcome this therapeutic dilemma. To achieve this goal, encapsulated Res 30 ± 4.8 nm IL-13Rα2-targeting nanoparticles (Pep-PP@Res) were constructed. Ultraviolet spectrophotometry revealed prolonged Res release (about 25%) from Pep-PP@Res in 48 h and fluorescent confocal microscopy showed the prolonged intracellular Res retention time of Pep-PP@Res (>24 h) in comparison with that of free Res (<4 h) and PP@Res (<4 h). MTT and EdU cell proliferation assays showed stronger suppressive effects of Pep-PP@Res on rat C6 GBM cells than that of PP@Res (p = 0.024) and Res (p = 0.009) when used twice for 4 h/day. Pep-PP@Res had little toxic effect on normal rat brain cells. The in vivo anti-glioblastoma effects of Res can be distinctly improved in the form of Pep-PP@Res nanoparticles via activating JNK signaling, upregulating proapoptosis gene expression and, finally, resulting in extensive apoptosis. Pep-PP@Res with sustained release and GBM-targeting properties would be suitable for in vivo management of GBMs.     

A Protease Inhibitor with Induction Therapy with Natural Interferon-β in Patients with HCV Genotype 1b Infection

The restoration of innate immune responses has potential as a novel therapeutic strategy for chronic hepatitis C (CHC). We compared the efficacy and safety of induction therapy (IT) with natural interferon-β (n-IFN-β) followed by pegylated-IFN-α/ribavirin (PR) alone (group A, n = 30) and IT with a protease inhibitor (PI) (simeprevir or vaniprevir)/PR (group B, n = 13) in CHC patients with genotype 1b and high viral loads. During IT with nIFN-β, virologic response rates in group A and group B were 10% and 8% (p = 0.6792) at week 4, 30% and 16% (p = 0.6989) at week 12 and 47% and 20% (p = 0.0887) at week 24 respectively. During and after the treatment with PR alone or PI/PR, virologic response rates in groups A and B were 50% and 82% (p = 0.01535) at week 4, 53% and 91% (p = 0.006745) at week 8, 57% and 91% (p = 0.001126) at week 12, 57% and 100% (p < 0.001845) at the end of the treatment and 57% and 80% (p < 0.005166) after treatment cessation. IT with PI/PR linked to the restoration of innate immune response was tolerated well, overcame virological breakthrough, enhanced early virologic responses, and resulted in a sustained virologic response in difficult-to-treat CHC patients. IT with PI/PR is beneficial for treating difficult-to-treat CHC patients.     

Amyloid Structural Changes Studied by Infrared Microspectroscopy in Bigenic Cellular Models of Alzheimer’s Disease

Alzheimer’s disease affects millions of lives worldwide. This terminal disease is characterized by the formation of amyloid aggregates, so-called amyloid oligomers. These oligomers are composed of β-sheet structures, which are believed to be neurotoxic. However, the actual secondary structure that contributes most to neurotoxicity remains unknown. This lack of knowledge is due to the challenging nature of characterizing the secondary structure of amyloids in cells. To overcome this and investigate the molecular changes in proteins directly in cells, we used synchrotron-based infrared microspectroscopy, a label-free and non-destructive technique available for in situ molecular imaging, to detect structural changes in proteins and lipids. Specifically, we evaluated the formation of β-sheet structures in different monogenic and bigenic cellular models of Alzheimer’s disease that we generated for this study. We report on the possibility to discern different amyloid signatures directly in cells using infrared microspectroscopy and demonstrate that bigenic (amyloid-β, α-synuclein) and (amyloid-β, Tau) neuron-like cells display changes in β-sheet load. Altogether, our findings support the notion that different molecular mechanisms of amyloid aggregation, as opposed to a common mechanism, are triggered by the specific cellular environment and, therefore, that various mechanisms lead to the development of Alzheimer’s disease.     

Using Synchrotron Radiation-Based Infrared Microspectroscopy to Reveal Microchemical Structure Characterization: Frost Damaged Wheat vs. Normal Wheat

This study was conducted to compare: (1) protein chemical characteristics, including the amide I and II region, as well as protein secondary structure; and (2) carbohydrate internal structure and functional groups spectral intensities between the frost damaged wheat and normal wheat using synchrotron radiation-based Fourier transform infrared microspectroscopy (SR-FTIRM). Fingerprint regions of specific interest in our study involved protein and carbohydrate functional group band assignments, including protein amide I and II (ca. 1774–1475 cm⁻¹), structural carbohydrates (SCHO, ca. 1498–1176 cm⁻¹), cellulosic compounds (CELC, ca. 1295–1176 cm⁻¹), total carbohydrates (CHO, ca. 1191–906 cm⁻¹) and non-structural carbohydrates (NSCHO, ca. 954–809 cm⁻¹). The results showed that frost did cause variations in spectral profiles in wheat grains. Compared with healthy wheat grains, frost damaged wheat had significantly lower (p < 0.05) spectral intensities in height and area ratios of amide I to II and almost all the spectral parameters of carbohydrate-related functional groups, including SCHO, CHO and NSCHO. Furthermore, the height ratio of protein amide I to the third peak of CHO and the area ratios of protein amide (amide I + II) to carbohydrate compounds (CHO and SCHO) were also changed (p < 0.05) in damaged wheat grains. It was concluded that the SR-FTIR microspectroscopic technique was able to examine inherent molecular structure features at an ultra-spatial resolution (10 × 10 μm) between different wheat grains samples. The structural characterization of wheat was influenced by climate conditions, such as frost damage, and these structural variations might be a major reason for the decreases in nutritive values, nutrients availability and milling and baking quality in wheat grains.