Search Results (53)

Hypertension and metabolic dysfunction-associated fatty liver disease (MAFLD) are both common chronic diseases globally. Nearly half of patients with hypertension are complicated by MAFLD. The mechanisms of the bidirectional promotion between the two remain unclear. The (pro) renin receptor ((P)RR) is one of the classic members of the renin–angiotensin system (RAS) and serves as the receptor for prorenin. Although the role of (P)RR in the induction and progression of hypertension has been extensively studied, its role and underlying mechanisms in MAFLD remain underreported. In this study, we aim to investigate the role of (P)RR in the pathogenesis of hypertension combined with MAFLD. In this study, SHRs were used for the model for hypertension combined with MAFLD. Liver lipid content analysis, liver H&E staining, the detection of (P)RR, ERK and downstream proteins related to fatty acid synthesis and transport, and RNA sequencing and data analysis were performed. In the in vitro experiments, we activated (P)RR using renin and established the lipid deposition model of HepG2 cells induced by renin for the first time. (P)RR was specifically blocked using handle region peptide (HRP), and Nile red fluorescence staining, (P)RR/ERK/PPARγ protein expression analysis, and immunofluorescence were performed to further verify the role of (P)RR in the pathogenesis of hypertension combined with MAFLD. Our results demonstrate that (P)RR plays a role in the development and progression of hypertension combined with MAFLD. The hepatic TG and FFA levels in the SHRs were increased, and the protein expression of the (P)RR/ERK/PPARγ pathway and downstream proteins related to fatty acid synthesis and transport were upregulated. HRP reversed the activation of these proteins and reduced intracellular lipid accumulation. In conclusion, our study first reveals that (P)RR is a potential therapeutic target for hypertension combined with MAFLD. And we found the (P)RR/ERK/PPARγ axis for the first time, which plays an important role in the progression of spontaneous hypertension combined with MAFLD. Full article

Aberrant protein glycosylation is closely associated with a number of biological processes and diseases. However, characterizing the types of post-translational modifications (PTMs) from the complex biological samples is challenging for comprehensive glycoproteomic analysis. The development of high-performance enrichment materials and strategies during the sample pretreatment process is a prerequisite to glycoproteome research. Here in this work, a sulfonate-rich covalent organic framework (COF) called TpPa-(SO₃H)₂ (referred to as SCOF-2) was synthesized using the Schiff base reaction for the identification of glycopeptides. Benefiting from high hydrophilicity and abundant sulfonate affinity, a total of 28 and 16 glycopeptides could be efficiently detected from the standard glycoproteins of horseradish peroxidase (HRP) and immunoglobulin G (IgG) tryptic digest, respectively. Moreover, the as-prepared sulfonate-rich SCOF-2 has an ultralow detection limit (0.01 fmol μL⁻¹), excellent enrichment selectivity (molar ratio HRP:BSA = 1:5000), satisfactory recovery rate (89.1%), high adsorption capacity (150 mg g⁻¹) and good reusability in the individual enrichment. Meanwhile, by using the SCOF-2 adsorbent, 196 and 194 endogenous glycopeptides in the serum of ovarian cancer patients and healthy people among triplicates were successfully enriched and identified, respectively, using combined nanoLC–MS/MS technology. It demonstrated its great application potential in glycoproteomics research and provided a novel insight for the design of affinity materials. Full article

In recent years, hydrogel beads and in situ hydrogels have gained wide attention in various fields such as biomedicine. In this study, 3-(4-hydroxyphenyl) propionic acid (HP) was introduced into the side chain of poly(α,β-[N-(2-hydroxyethyl)-D,L-aspartamide]) (PHEA) to synthesize phenolic hydroxyl-functionalized poly(aspartamide) derivative PHEA-HP with enzyme-catalyzed cross-linking potential. First, the chemical structure of PHEA-HP was characterized by FT-IR, UV and ¹H NMR, and the results of in vitro cytotoxicity against L929 cell line and hemolysis experiment showed that PHEA-HP did not have toxicity to cells (viability > 90%) and had good blood compatibility. Then, rheological measurement confirmed the formation of PHEA-HP-based in situ hydrogel with a high storage modulus (G′) around 10⁴ Pa, and the vial-tilting method revealed that the gelation time of PHEA-HP aqueous solution could be tuned in the wide range of 5–260 s by varying the concentrations of hydrogen peroxide (H₂O₂) and horseradish peroxidase (HRP). Finally, hydrogel beads of different diameters containing methylene blue (for easy observation) were prepared using a coaxial needle and syringe pumps, and the effect of the flow rate of the outer phase on the diameters of the hydrogel beads was also investigated. Therefore, PHEA-HP may be a promising and safe poly(aspartamide) derivative that can be used to prepare in situ hydrogels and hydrogel beads for applications closely related to the human body. Full article

In this study, Fe₃(PO₄)₂·8H₂O magnetic nanoparticles (MNPs) were successfully extracted from the strain Burkholderia cepacia CG-1. We subsequently characterized their composition, structure, and morphology, revealing that these nanoparticles consisted of Fe₃(PO₄)₂·8H₂O with an average diameter of 66.87 ± 0.56 nm. Our measurements indicated magnetic parameters of 151 Oe for coercivity, 2 emu/g for saturation remanence, and 16 emu/g for saturation magnetization. Our findings confirmed that these magnetic nanoparticles exhibited intrinsic peroxidase-like activity, catalyzing the oxidation of 3,3,5,5-tetramethylbenzidine (TMB) in the presence of H₂O₂. Compared to horseradish peroxidase (HRP), the peroxidase mimic demonstrated greater stability under various physicochemical conditions. To investigate the mechanisms underlying the peroxidase-like catalysis of Fe₃(PO₄)₂·8H₂O, we employed fluorescence spectroscopy and electron spin resonance (ESR) analysis. The results revealed that the peroxidase-like activity of Fe₃(PO₄)₂·8H₂O stemmed from the generation of hydroxyl radicals (·OH). Furthermore, we established a platform for the colorimetric detection of H₂O₂ and glucose. Our method was capable of detecting H₂O₂ concentrations as low as 1.0 × 10⁻³ mmol/L. Impressively, this sensitive method was successfully applied to determine glucose levels in human serum. Full article

Oriented antibody immobilization has been widely employed in immunoassays and immunodiagnoses due to its efficacy in identifying target antigens. Herein, a heptapeptide ligand, HWRGWVC (HC7), was coupled to poly(glycidyl methacrylate) (PGMA) nanospheres (PGMA-HC7). The antibody immobilization behavior and antigen recognition performance were investigated and compared with those on PGMA nanospheres by nonspecific adsorption and covalent coupling via carbodiimide chemistry. The antibodies tested included bovine, rabbit, and human immunoglobulin G (IgG), while the antigens included horseradish peroxidase (HRP) and β-2-Microglobulin (β2-MG). The nanospheres were characterized using zeta potential and particle size analyzers, scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, and reversed-phase chromatography, proving each synthesis step was succeeded. Isothermal titration calorimetry assay demonstrated the strong affinity interaction between IgG and PGMA-HC7. Notably, PGMA-HC7 achieved rapid and extremely high IgG adsorption capacity (~3 mg/mg) within 5 min via a specific recognition via HC7 without nonspecific interactions. Moreover, the activities of immobilized anti-HRP and anti-β2-MG antibodies obtained via affinity binding were 1.5-fold and 2-fold higher than those of their covalent coupling counterparts. Further, the oriented-immobilized anti-β2-MG antibody on PGMA-HC7 exhibited excellent performance in antigen recognition with a linear detection range of 0–5.3 μg/mL, proving its great potential in immunoassay applications. Full article

Grapes and grape products are rich in secondary metabolites such as phenolic compounds and anthocyanins, which have antioxidant properties. These compounds possess health-promoting attributes, including cardioprotective, antimicrobial, and anticancer effects. In recent years, biotechnological methods have been employed to produce high quantities and purity of secondary metabolites under in vitro conditions, aiming to elucidate their complex functions and optimize production methods. However, the potential effects of harpin proteins on the accumulation of secondary compounds in callus cultures have not been investigated thus far. Harpin proteins, encoded by the hrp gene clusters in Gram-negative phytopathogens, are known to trigger defense responses in various plant species by promoting the accumulation of secondary compounds. These findings suggest that harpin proteins may have the potential to enhance secondary metabolite accumulation in callus cultures. This study therefore investigated the potential of applying different concentrations of harpin protein (0, 0.1, 1, 10, and 100 ppm) to increase secondary metabolite production in calluses derived from petioles of the “Horoz Karası” grape cultivar. Our findings revealed that 1 and 10 ppm harpin treatments resulted in the highest anthocyanin accumulations, with 17.21 and 16.57 CV/g, respectively, representing 1.95- and 1.87-fold increases compared to control treatments, respectively. Total phenolic content peaked at 0.39 mg GAE g⁻¹ FW with the 1 ppm harpin treatment, representing a 4.33-fold increase over the control. Total flavanol levels reached their highest levels at 0.027 mg CE g⁻¹ FW with 1 and 10 ppm harpin concentrations, resulting in a 2.25-fold increase compared to the control. The highest averages for total flavonol content were recorded at 0.024 and 0.021 mg RE g⁻¹ FW with 1 and 10 ppm harpin concentrations, respectively, representing 1.5- and 1.3-fold increases over the control. Principal component analysis (PCA) corroborated the results obtained from the heatmap analysis, indicating that harpin applications at 1 and 10 ppm were the most effective concentration range for maximizing secondary metabolite synthesis, while very low or high concentrations diminished these effects. These findings offered valuable insights for optimizing the production of high-value bioactive compounds, which can be utilized in various fields such as medicine, pharmaceuticals, food, and cosmetics. These results are expected to serve as a valuable reference for elucidating the mechanisms by which harpin proteins, rarely used in vitro, exert their effects on grapevine calluses, contributing to the literature in this domain. Full article

In this study, an innovative conductive hybrid biomaterial was synthetized using collagen (COL) and reduced graphene oxide (rGO) in order for it to be used as a wound dressing. The hydrogels were plasticized with glycerol and enzymatically cross-linked with horseradish peroxidase (HRP). A successful interaction among the components was demonstrated by FTIR, XRD, and XPS. It was demonstrated that increasing the rGO concentration led to higher conductivity and negative charge density values. Moreover, rGO also improved the stability of hydrogels, which was expressed by a reduction in the biodegradation rate. Furthermore, the hydrogel’s stability against the enzymatic action of collagenase type I was also strengthened by both the enzymatic cross-linking and the polymerization of dopamine. However, their absorption capacity, reaching values of 215 g/g, indicates the high potential of the hydrogels to absorb fluids. The rise of these properties positively influenced the wound closure process, achieving an 84.5% in vitro closure rate after 48 h. These findings clearly demonstrate that these original composite biomaterials can be a viable choice for wound healing purposes. Full article

Xanthomonas oryzae pv. oryzicola (Xoc) is a notorious plant pathogen. Like most bacterial pathogens, Xoc has evolved a complex regulatory network to modulate the expression of various genes related to pathogenicity. Here, we have identified TfmR, a transcriptional regulator belonging to the TetR family, as a key player in the virulence mechanisms of this phytopathogenic bacterium. We have demonstrated genetically that tfmR is involved in the hypersensitive response (HR), pathogenicity, motility and extracellular polysaccharide production of this phytopathogenic bacterium. Our investigations extended to exploring TfmR’s interaction with RpfG and HrpX, two prominent virulence regulators in Xanthomonas species. We found that TfmR directly binds to the promoter region of RpfG, thereby positively regulating its expression. Notably, constitutive expression of RpfG partly reinstates the pathogenicity compromised by TfmR-deletion mutants. Furthermore, our studies revealed that TfmR also exerts direct positive regulation on the expression of the T3SS regulator HrpX. Similar to RpfG, sustained expression of HrpX partially restores the pathogenicity of TfmR-deletion mutants. These findings underscore TfmR’s multifaceted role as a central regulator governing key virulence pathways in Xoc. Importantly, our research sheds light on the intricate molecular mechanisms underlying the regulation of pathogenicity in this plant pathogen. Full article

Aim. To identify subgroups of patients with primary osteoarthritis of the hip joint (pHOA) with similar imaging and laboratory findings, disease evolution, and response to conventional therapies. Methods. We performed further statistical analyses on patient data from two published, double-blind, randomized, and placebo-controlled studies (DB-RCTs), which examined the effects of intra-articular corticosteroids (ia-CSs), hyaluronic acid (ia-HA)—KИ-109-3-0008/14.01.2014, and intravenous bisphosphonates (iv-BPs) -KИ- 109-3-0009/14.01.2014 compared to the country’s standard pHOA therapy. The data span an 8-year follow-up of 700 patients with pHOA, including: 1. Clinical parameters (WOMAC-A, B, C, and T; PtGA). 2. Laboratory markers (serum calcium and phosphate levels; 25-OH-D and PTH, markers for bone sCTX-I and cartilage uCTX-II turnover). 3. Radiological indicators: X-ray stage (Kellgren-Lawrence (K/L) and model (Bombelli/OOARSI), width (mJSW), speed (JSN mm/year), and zone of maximum narrowing of the joint space (max-JSN)—determining the type of femoral head migration (FHM). 4. DXA indicators: bone geometry (HAL; NSA; and MNW); changes in regional and total bone mineral density (TH-BMD, LS-BMD, and TB-BMD). 5. Therapeutic responses (OARSI/MCII; mJSW; JSNmm/yearly) to different drug regimens (iv-BP -zoledronic acid (ZA/-5 mg/yearly for 3 years)); ia-CS 40 mg methylprednisolone acetate, twice every 6 months; and ia-HA with intermediate molecular weight (20 mg/2 mL × 3 weekly applications, two courses every 6 months) were compared to standard of care therapy (Standard of Care/SC/), namely D3-supplementation according to serum levels (20–120 ng/mL; target level of 60 ng/mL), simple analgesics (paracetamol, up to 2.0 g/24 h), and physical exercises. The abovementioned data were integrated into a non-supervised hierarchical agglomerative clustering analysis (NHACA) using Ward’s linkage method and the squared Euclidean distance to identify different endophenotypes (EFs). Univariate and multivariate multinomial logistic regression analyses were performed to determine the impact of sex and FHM on clinical and radiographic regression of pHOA. Results. A baseline cluster analysis using incoming (M0) patient data identified three EFs: hypertrophic H-HOA, atrophic A-HOA, and intermediate I-HOA. These EFs had characteristics that were similar to those of patients grouped by radiographic stage and pattern (‘H’-RPs, ‘I’-RPs, and ‘A’-RPs), p < 0.05). The repeated cluster analysis of M36 data identified four EF pHOAs: 1. Hypertrophic (slow progressors, the influence of the type of femoral head migration (FHM) outweighing the influence of sex on progression), progressing to planned total hip replacement (THR) within 5 (K/LIII) to 10 (K/LII) years. 2. Intermediate (sex is more important than the FHM type for progression) with two subgroups: 2#: male-associated (slow progressors), THR within 4 (K/LIII) to 8 years. (K/LII). 2* Female-associated (rapid progressors), THR within 3 (K/LIII) to 5 (K/LII) years. 3. Atrophic (rapid progressors; the influence of FHM type outweighs that of sex), THR within 2 (K/LIII) to 4 (K/LII) years. Each EF, in addition to the patient’s individual progression rate, was also associated with a different response to the aforementioned therapies. Conclusions. Clinical endophenotyping provides guidance for a personalized approach in patients with pHOA, simultaneously assisting the creation of homogeneous patient groups necessary for conducting modern genetic and therapeutic scientific studies. Full article

Chlorophenol compounds pose a health risk to many organisms due to their toxicity. The present paper presents the estimation of the activation and deactivation energies and the optimum temperatures of 2,4-dichlorophenol degradation by horseradish peroxidase (HRP). The activities of horseradish peroxidase depending on temperature were analyzed. In a mathematical model, describing 2,4-dichlorophenol degradation by HRP was assumed that both the 2,4-dichlorophenol degradation and the deactivation of HRP were first-order reactions by the enzyme concentration. The parameters of the optimum temperatures $T_{\mathrm{o}\mathrm{p}\mathrm{t}}$, the activation energies $E_{\mathrm{r}}$, and the deactivation energies $E_{\mathrm{d}}$ in the process of 2,4-dichlorophenol degradation by HRP immobilized on a modified nanofibrous membrane were determined $k_{\mathrm{d}} \mathrm{and} t_{1/2}$ were determined for HRP immobilized at temperatures in the range of 25 °C to 75 °C. Likewise, thermodynamic parameters such as the change in the enthalpy ${∆H}^{\#}$, change in entropy ${∆S}^{\#}$, the change in Gibbs free energy ${∆G}^{\#}$ for native HPR and the change in the enthalpy ${∆H}_d^{\#}$, change in entropy ${∆S}_d^{\#}$, and the change in Gibbs free energy ${∆G}_d^{\#}$ for deactivated HRP were determined at 25 °C. Full article

Biomarkers, ranging from molecules to behavior, can be used to identify thresholds beyond which performance of mission tasks may be compromised and could potentially trigger the activation of countermeasures. Identification of homologous brain regions and/or neural circuits related to operational performance may allow for translational studies between species. Three discussion groups were directed to use operationally relevant performance tasks as a driver when identifying biomarkers and brain regions or circuits for selected constructs. Here we summarize small-group discussions in tables of circuits and biomarkers categorized by (a) sensorimotor, (b) behavioral medicine and (c) integrated approaches (e.g., physiological responses). In total, hundreds of biomarkers have been identified and are summarized herein by the respective group leads. We hope the meeting proceedings become a rich resource for NASA’s Human Research Program (HRP) and the community of researchers. Full article

Acacia melanoxylon (blackwood) is a valuable wood with excellent-quality heartwood extensively utilized worldwide. The main aim of this study was to confirm the horizontal and vertical variation and provide estimated values of genetic gains and clonal repeatabilities for improving the breeding program of A. melanoxylon. Six blackwood clones at 10 years old were analyzed in Heyuan and Baise cities in China. Stem trunk analysis was conducted for sample trees to explore the differences between heartwood and sapwood. The heartwood radius (HR), heartwood area (HA), and heartwood volume (HV) in heartwood properties decreased as the tree height (H) in growth traits increased, and the HV = 1.2502 DBH (diameter at breast height)^1.7009 model can accurately estimate the heartwood volume. Furthermore, G × E analysis showed that the heritabilities of the eleven indices, including DBH, DGH (diameter at ground height), H, HR, SW (sapwood width), BT (bark thickness), HA, SA (sapwood area), HV, HRP (heartwood radius percentage), HAP (heartwood area percentage), and HVP (heartwood volume percentage) were between 0.94 and 0.99, and repeatabilities of the eleven indices were between 0.74 and 0.90. Clonal repeatability of DBH (0.88), DGH (0.88), and H (0.90) in growth traits and HR (0.90), HVP (0.90), and HV (0.88) in heartwood properties were slightly higher than for SA (0.74), SW (0.75), HAP (0.75), HRP (0.75), and HVP (0.75). These data also implied that the growth characteristics of heartwood and sapwood of blackwood clones were less affected by the environment and had substantial heritability. Full article

The multi-detection size exclusion chromatography (SEC) has been recognized as an advanced analytical technique for the characterization of macromolecules and process control, as well as the manufacturing and formulation of biotechnology products. It reveals reproducible molecular characterization data, such as molecular weight and its distribution, and the size, shape, and composition of the sample peaks. The aim of this work was to investigate the potential and suitability of the multi-detection SEC as a tool for surveillance over the molecular processes during the conjugation reaction between the antibody (IgG) and horseradish peroxidase (HRP), and demonstrate the plausibility of its application in the quality control of the final product, the IgG-HRP conjugate. Guinea pig anti-Vero IgG-HRP conjugate was prepared using a modified periodate oxidation method, based on periodate oxidation of the carbohydrate side chains of HRP, followed by the formation of Schiff bases between the activated HRP and amino groups of the IgG. The quantitative molecular characterization data of the starting samples, intermediates, and final product were obtained by multi-detection SEC. Titration of the prepared conjugate was performed by the ELISA and its optimal working dilution was determined. This methodology proved to be a promising and powerful technology for the IgG-HRP conjugate process control and development, as well as for the quality control of the final product, as verified by the analysis of several commercially available reagents. Full article

Contamination of deoxynivalenol (DON) in grains has attracted widespread concern. It is urgently needed to develop a highly sensitive and robust assay for DON high-throughput screening. Antibody against DON was assembled on the surface of immunomagnetic beads orientationally by the aid of Protein G. AuNPs were obtained under the scaffolding of poly(amidoamine) dendrimer (PAMAM). DON-horseradish peroxidase (HRP) was combined on the periphery of AuNPs/PAMAM by a covalent link to develop DON-HRP/AuNPs/PAMAM. Magnetic immunoassay based on DON-HRP/AuNPs/PAMAM was optimized and that based on DON-HRP/AuNPs and DON-HRP was adopted as comparison. The limits of detection (LODs) were 0.447 ng/mL, 0.127 ng/mL and 0.035 ng/mL for magnetic immunoassays based on DON-HRP, DON-HRP/Au and DON-HRP/Au/PAMAM, respectively. Magnetic immunoassay based on DON-HRP/AuNPs/PAMAM displayed higher specificity towards DON and was utilized to analyze grain samples. The recovery for the spiked DON in grain samples was 90.8–116.2% and the method presented a good correlation with UPLC/MS. It was found that the concentration of DON was in the range of ND-3.76 ng/mL. This method allows the integration of dendrimer–inorganic NPs with signal amplification properties for applications in food safety analysis. Full article

Instead of Western blot being considered as a gold standard for intracellular protein expression assays, we developed a novel multiplexed high throughput (180 tests/day) in situ manual protein expression method directly in 96-well plates using 25,000–100,000 cells/well after formaldehyde fixation and Triton X 100 permeabilization. HepG2 cells were treated with ochratoxin A (OTA) and staurosporine (STP) to induce apoptosis. Antioxidant and apoptotic cell signaling protein expression were studied by various rabbit primary antibodies and HRP labeled secondary antibodies. The HRP labeled immune complexes were developed by H₂O₂/Ampliflu Red fluorogenic reagent and measured in a plate reader. Our assay can simultaneously quantify 22 protein antigens in one plate with 4 technical replicates with an interassay imprecision of <10% CV. The fluorescence signals are referred to total intracellular protein contents in the wells and given as fluorescence/protein ratio FPR, expressed as % of the controls (FPR %). OTA caused a dose–response increase (p < 0.05–p < 0.001) in SOD2, CAT, ALB, CASP3,7,9, BCL2, BAX, Nf-kB, phospho-Erk1/2/Erk1/2, phospho-Akt/Akt, phospho-p38/p38, and phospho-PPARg/PPARg levels while phospho-AMPK/AMPK ratios decreased (p < 0.05–p < 0.001). On the contrary, STP induced a dose–response decrease (p < 0.05–p < 0.001) in CASP3,7,9, BAX, BCL2, Nf-kB and phospho-Erk1/2/Erk1/2 expression while B-ACT, phospho-Akt/Akt, phospho-p38/p38 and phospho-PPARg/PPARg ratios increased. Full article