Peroxidase Complex Research Articles

Sandwich-Type Electrochemical Aptasensor with Supramolecular Architecture for Prostate-Specific Antigen.

A novel sandwich-type electrochemical aptasensor based on supramolecularly immobilized affinity bioreceptor was prepared via host-guest interactions. This method utilizes an adamantane-modified, target-responsive hairpin DNA aptamer as a capture molecular receptor, along with a perthiolated β-cyclodextrin (CD) covalently attached to a gold-modified electrode surface as the transduction element. The proposed sensing strategy employed an enzyme-modified aptamer as the signalling element to develop a sandwich-type aptasensor for detecting prostate-specific antigen (PSA). To achieve this, screen-printed carbon electrodes (SPCEs) with electrodeposited reduced graphene oxide (RGO) and gold nanoferns (AuNFs) were modified with the CD derivative to subsequently anchor the adamantane-modified anti-PSA aptamer via supramolecular associations. The sensing mechanism involves the affinity recognition of PSA molecules on the aptamer-enriched electrode surface, followed by the binding of an anti-PSA aptamer-horseradish peroxidase complex as a labelling element. This sandwich-type arrangement produces an analytical signal upon the addition of H2O2 and hydroquinone as enzyme substrates. The aptasensor successfully detected the biomarker within a concentration range of 0.5 ng/mL to 50 ng/mL, exhibiting high selectivity and a detection limit of 0.11 ng/mL in PBS.

Biodegradation and Detoxification of Some Dyes by Crude Lignin Peroxidase Complex Produced by Escherichia coli Accession No: LR0250096.1 and Pseudomonas aeruginosa Accession No: CP031449.2

Synthetic and untreated dyes discharged in wastewater effluents are a threat to an ecosystem. This study investigated dye degradation and detoxification efficiency of crude lignin peroxidase separately obtained from the cultures of Escherichia coli (LR0250096.1) and Pseudomonas aeruginosa (CP031449.2). The ability of the crude lignin peroxidase to degrade Malachite Green (MG), Remazol Brilliant Blue R (RBBR), Congo Red (CR), and Azure B (AZ) was evaluated at different operating conditions (enzyme, dye, and hydrogen peroxide concentrations; pH; temperature; and contact time). The ability of the degraded dyes to support the growth of bacteria was also investigated. The observed optimum operating conditions for lignin peroxidase extracts of the Escherichia coli on AZ were 20 mg/mL enzyme concentration, 50 mg/L dye, pH 7.0, temperature 50 °C, and 1.5 mM hydrogen peroxide within 20–50 min of incubation time and on MG were 20 mg/mL, 50 mg/L, 9.0, 30 °C, 0.1 mM, and 20 min, respectively. The enzyme extract from Pseudomonas aeruginosa on AZ demonstrated optimum operation conditions of 20 mg/mL, 50 mg/L, pH 9.0, 40 °C, 1.5 mM, and 50 min, respectively and on MG, they were 20 mg/mL, 50 mg/L, 6.0, 30 °C, 1.0 mM, and 20 min, respectively). The prepared enzyme showed an appreciable degradative effect on CR and RBBR compared with commercial lignin peroxidase. The degraded dyes were able to support the growth of two Gram-positive (Bacillus cereus and Staphylococcus aureus), and two Gram-negative (Proteus mirabilis and Escherichia coli) bacteria, indicating the efficiency and the potential use of the enzyme complexes in the clean-up of industrial dyes’ waste.

The recent research progress in neurobiological characteristics and pain regulation of the cerebrospinal fluid-contacting nucleus

AbstractThe ependymal epithelium forms the cerebrospinal fluid barrier, separating the brain and spinal cord from the cerebrospinal fluid. However, in specific regions of the central nervous system, there are neurons that directly interface with the cerebrospinal fluid, including neuronal bodies, dendrites, or axons, This constitutes what is referred to as the "cerebrospinal fluid contacting neurons system (CSF-CNS)". The research team led by Professor Zhang has successfully utilized cholera toxin subunit B coupled horseradish peroxidase complex (CB-HRP) to selectively label the specialized neuron system that interfaces with cerebrospinal fluid, pioneeringly designating it as the "cerebrospinal fluid-contacting nucleus", commonly referred to as the "CSF-contacting nucleus". For the first time, the discovery of the CSF-contacting nucleus provides compelling morphological evidence for the existence of a distinct neural structure within the brain parenchyma that establishes a connection with the cerebrospinal fluid, thereby suggesting its potential significance in facilitating material and information exchange between the brain parenchyma and cerebrospinal fluid. After conducting a comprehensive series of studies on the morphological structure, material expression, gene analysis and functional aspects of the CSF-contacting nucleus in rodents and non-human primates, it has been revealed that there are fibrous connections between the CSF-contacting nucleus and the cerebral cortex and subcortical nuclei being involved in the regulatory mechanisms of pain, cognition, learning and memory, emotion, addiction, stress and anxiety responses, visceral activity, olfaction, vision processing and perception, auditory processing, perception, motor control and coordination, homeostasis regulation including maintenance of body energy and fluid balance, as well as the control of sleep–wake cycles and synchronization of biological rhythms. Current experiments have confirmed that the CSF-contacting nucleus is related to pain, morphine dependence and withdrawal, learning and memory, as well as stress. This present article offers a comprehensive review of the neurobiological characteristics and recent advancements in pain regulation of the CSF-contacting nucleus. The aim is to provide novel insights into the investigation of pain regulation within bidirectional regulatory pathway between the brain and cerebrospinal fluid, with a specific focus on elucidating the role of the CSF-contacting nucleus as a bridge structure. Additionally, the objective of this research is to propose innovative strategies for pain management and associated disorders in the future.

Actinobacterial peroxidase-mediated biodeterioration of hazardous explosive, 2, 4, 6, trinitrophenol by in silico and in vitro approaches.

Explosives are perilous and noxious to aquatic biota disrupting their endocrinal systems. Supplementarily, they exhibit carcinogenic, teratogenic and mutagenic effects on humans and animals. Henceforth, the current study has been targeted to biotransform the explosive, 2, 4, 6 trinitrophenol (TNP) by wetland peroxidase from Streptomyces coelicolor. A total peroxidase yield of 20,779mg/l with 51.6 folds of purification was observed. In silico molecular docking cum in vitro appraisals were accomplished to assess binding energy and interacting binding site residues of peroxidase and TNP complex. TNP required a minimal binding energy of-6.91kJ/mol and was subjected to biodeterioration (89.73%) by peroxidase in purified form, with 45kDa and a similarity score of 34 by MASCOT protein analysis. Moreover, the peroxidase activity was confirmed with Zymogram analysis. Characterization of peroxidase revealed that optimum values of pH and temperature as 6 and 40 °C, respectively, with their corresponding stability varying from 3.5 to 7. Interestingly, the kinetic parameters such as Km and Vmax on 2,2'-azino-bis 3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and H2O2 were 19.27µm and 0.41µm/min; 21.4µm and 0.1µm/min, respectively. Among the diverse substrates, chemicals and trace elements, ABTS (40mM), citric acid (5mM) and Fe2+ (5mM) displayed the highest peroxidase activity. Computational docking and in vitro results were corroborative and UV-Vis spectroscopy, HPLC, FTIR and GC-MS indicated the presence of simple metabolites of TNP such as nitrophenols and benzoquinone, showcasing the efficacy of S. coelicolor peroxidase to biotransform TNP. Henceforth, the current study offers a promising channel for biological treatment of explosive munitions, establishing a sustainable green earth.

The oral administration of Lotus corniculatus L. attenuates acute and chronic pain models in male rats

Ethnopharmacological relevanceLotus corniculatus L. (Fabaceae) traditionally used in Persian folk medicine to heal peritoneal inflammation and back pain. Aim of the studyTo explore the antinociceptive (acute pain) and anti-neuropathic (chronic pain) activities of Lotus corniculatus leaves essential oil (LCEO) in addition to uncovering the possible mechanisms of antinociception. Materials and methodsLCEO as well as the pure oleanolic acid (OA) compound, were assayed for their effects on acute (formalin induced paw licking test or FIPT) and chronic (cervical contusion injury models on the fifth cervical vertebra or CCS; 14-day intervals) pain. The possible involvements of NO-cGMP-K+ channel, TRPV, dopamine, cannabinoid, PPAR, adrenergic, and opioid mechanisms in the antinociceptive activity of LCEO have studied by formalin test. The levels of p53 and inflammatory markers were measured using a streptavidin biotin immune peroxidase complex and ELISA methods, respectively. ResultsThe LCEO and OA exerted antinociceptive activity in the first-phase of FIPT. Pretreatment with antagonists of TRPV1, dopamine D2, cannabinoid type1 and 2, and NO-cGMP-K+ channel blockers (glibenclamide, L-NAME and methylene blue) attenuated the antinociceptive effect of LCEO in FIPT. In addition, LCEO and OA meaningfully reduced hyperalgesia (days 6–14) and mechanical allodynia (days 2–14) in the CCS model. LCEO suppressed the apoptotic marker (p53) in CCS model and also ameliorated IL-2, TNF-α, and IL-1 in the spinal cord. ConclusionFinally, LCEO inhibited acute (possibly via the modulation of opioid, TRPV, dopamine, cannabinoid mechanisms as well as NO-cGMP-K+ channel) and chronic pain (via suppressing apoptotic and inflammatory markers) in male rats. The results also suggest that OA has analgesic activity against acute and chronic pain conditions.

Unveiling the Potential of Antioxidant Proteins with the Integration of Little Millet Phytochemicals from GC-MS Studies through In silico Approach

Aim: Millet extracts contain bioactive compounds that have antioxidant properties, anti-diabetic, anti-inflammatory, and other health-promoting properties. Little millet contains more protein, minerals, vitamins, and carbohydrates than rice and wheat. The dynamics of soil organic matter may be significantly impacted by the presence of antioxidants compounds in the soil. Antioxidants and protein-modifying substances are dietary components that change numerous characteristics and, in some cases, reverse ageing. Thus, exploring the phytochemicals in little millet is very much essential in understanding its biological functional implications.
 Methodology: We have carried out the GCMS analysis for the little millet (seed). Further, we have performed the molecular docking and molecular dynamics simulation for the shortlisted phytochemicals. 
 Results: We screened the metabolites using GCMS analysis due to the unexplored phytochemicals of little millet. Docking against the little millet phytochemicals was done with a focus on key antioxidants such as superoxide dismutase, catalase, and glutathione peroxidase. Acetin compound displayed strong binding with superoxide dismutase and glutathione peroxidase, while hexadecenoic acid exhibited best affinity with catalase. Through molecular dynamics simulations, we found the glutathione peroxidase complex to be the most stable. This stability implies enhanced antioxidant activity, crucial in counteracting oxidative stress.
 Conclusion: This study uncovers the untapped potential of little millet’s phytochemicals. By elucidating their interaction with vital antioxidant proteins, it opens avenues for innovative anti-aging strategies, health interventions and helps in enhancing the plant defence mechanism.

Co-catalysis of melanin degradation by laccase-manganese peroxidase complex from Trametes hirsuta OK271075 for application in whitening cosmetics

Melanins are a complex natural pigment that darken the skin, a phenomenon regarded as an esthetic problem in tropical countries such as Indonesia. Melanin can be degraded by various oxidative enzymes such as laccase (Lac) and manganese peroxidase (MnP) which are produced by white rot fungi (WRF). In the current study, we successfully obtained a partial purified Lac-MnP from a tropical WRF called Trametes hirsuta OK271075. Co-catalysis of Lac and MnP in the melanin degradation existed as indicated by the enhancement of degradation upon the addition of mediators for the respective enzymes. The highest degradation was up to 42% in the presence of TEMPO (0.2 mM) and MnSO4-H2O2 (0.75 mM, 0.2 mM) as mediators. The increment was up to 20% compared to treatment without mediators. A higher degradation rate achieved when specific mediators of the enzymes were utilized indicated a synergistic effect of both enzymes that could be used simultaneously for melanin degradation. FTIR and FESEM analysis revealed significant changes in melanin structure between enzyme-treated and untreated melanin. Melanin granules were cracked and exfoliated due to an enzyme attack. Based on Pyrolysis-GCMS analysis, several compounds such as phenols, epoxides, ketones, and carboxylic acids were found in higher amount in the enzyme-treated than untreated melanin. This finding revealed the potency of a new isolated T. hirsuta for new enzyme applications in biotechnology fields, especially to degrade melanin for application in natural whitening cosmetics.

Hydroxylated Graphene Porous Membrane-Based Biosensor for Exosome Isolation and Detection

Exosomes are nanoscale (about 100–200 nm) extracellular vesicles (EVs) that are secreted by cells into the extracellular space and in body liquid circulation. Recent studies on exosomes have shown that they play an important role in various biological processes and can be detected for an early diagnosis of cancer. In this work, we developed a graphene-based biosensor that could isolate exosomes of breast cancer cells from EVs and simultaneously achieve quantitative detection of exosomes. G-OH (hydroxylated graphene), a novel nanomaterial with high conductivity, good biocompatibility, and huge surface area, is quite suitable for the preparation of biosensors. First, the G-OH nanosheets were deposited to form the membrane with the micropores to screen exosomes and microvesicles in EVs. Then, rabbit α-human CD44 antibody was incubated in the membrane to capture MDA-MB-231 (MM-231) exosomes. Finally, modified mouse α-human CD9 antibodies and the third anti-horseradish peroxidase complex were incubated in the membrane, respectively. Under the optimal experimental conditions, this biosensor showed a wide linear range of 25 to 1 × 106 particles·μL–1 and a low detection limit of 9 particles·μL–1 (S/N = 3), spanning 5 orders of magnitude. The quantitative detection of the cancer cell exosomes could be achieved in a mixture of cancer cells and healthy cell exosomes.

Immunohistochemical Detection of Pro-Inflammatory and Anti-Inflammatory Interleukins in the Lungs of Sheep with Jaagsiekte

Objective: In this study, it was aimed to evaluate the levels of interleukins such as IL-1β, IL-6, IL-10 and IL-12β in Jaagsiekte sheep by immunohistochemical methods. In this way, it will be revealed whether interleukins are effective in the progression of Jaagsiekte and how useful they are in the diagnosis of the disease. 
 Material-Method: The material of current study consisted of lung tissues of 26 sheep (Control, n=6 and Jaagsiekte, n=20) brought to Department of Pathology for routine histopathological diagnosis. Tissue samples taken were fixed in 10% buffered formaldehyde solution. 5 µm-thick sections were taken from the paraffin blocks prepared after routine tissue follow-up procedures. Hematoxylin & Eosin staining was applied to the sections in order to detect histopathological changes. Sections were examined and photographed under a light microscope. The routine streptavidin–biotin peroxidase complex method was used. 
 Results: In sheep with Jaagsiekte, tumoral foci with large and small acinar or papillary growths were observed in the alveolar and bronchiole lumens. The control group was negative for IL-1β, IL-6, IL-10 and IL-12β immunoreactivity. IL 1β-6-10 and 12β levels were dramatically increased in the Jaagsiekte group compared to the control group. 
 Conclusion: It was determined that interleukins were produced from tumoral cells and tumor microenvironment elements, and these interleukins showed pro-inflammatory effects, except for IL-10. Data from the current study show that interleukins are very useful markers for the diagnosis of Jaagsiekte.

Detection of Dengue Virus Transovarial Transmission in Dengue Hemorrhagic Fever Endemic Areas

Dengue virus is a group of RNA viruses that are highly pathogenic in humans and spread quickly through the bites of Aedes aegypti and Aedes albopictus mosquitoes, especially in tropical countries. More than half a billion out of 100 countries worldwide are at serious risk of dengue virus infection. Vector surveillance activities with Ovitrap and detection of dengue virus types in Aedes aegypti and Aedes albopictus have never been carried out in Pontianak City. It is important in early alert systems at transmission foci. The purpose of this study was to prove the transovarial transmission of dengue virus in Aedes aegypti and Aedes albopictus mosquitoes with a transovarial transmission index (TTI) in endemic areas in Pontianak City, West Kalimantan. The method used in this research is descriptive observational, viral examination method with immunocytochemistry streptavidin-biotin peroxidase complex (ISBPC) and Polymerase Chain Reaction Transcription Reaction (PCR) aimed at proving the presence of transovarial transmission of dengue virus in the same period. The conclusion in this study is that there is evidence of transovarial transmission of dengue virus in Aedes mosquitoes in endemic areas by 29.3% in Sungai Jawi Dalam sub-district, West Pontianak sub-district, and 39.6% in Batu Layang sub-district, North Pontianak sub-district, mosquito density from the results of the Ovitrap Index measurement (OI) in Batu Layang Village is denser, namely 41.3%, compared to Sungai Jawi Village, which is 38.22% and has succeeded in identifying the type of dengue virus, namely the Dengue virus strain, in the two research locations.

The effects of hypoxia-inducible factors-1α and -2α and erythroferrone on hepcidin in patients with chronic kidney disease stages 3–5 and renal anemia

Objective This study aimed to investigate the effects of hypoxia-inducible factor-1α (HIF-1α), hypoxia-inducible factor-2α (HIF-2α), and erythroferrone (ERFE) on hepcidin in patients with chronic kidney disease (CKD) stages 3–5 and renal anemia. Methods A total of 90 patients with CKD stages 3–5 and renal anemia were selected for the study at the Nephrology Department of Fujian Provincial People’s Hospital and divided into three groups, according to CKD stage, while another 30 healthy subjects who underwent a physical examination at the hospital during the same period were selected as the normal group. The serum levels of hepcidin, HIF-1α, HIF-2α, ERFE, and furin were measured using an avidin biotin peroxidase complex enzyme-linked immunosorbent assay to compare the differences between the groups in the related indicators. Results ① Serum HIF-2α, HIF-1α, ERFE, and furin levels increased gradually in the patients with CKD stages 3–5 ( p < 0.05, p < 0.01). ②Simple correlation analysis:Serum hepcidin was positively correlated with HIF-2α, ERFE, and HIF-1α in the CKD patients ( p < 0.01). ③Serum hepcidin was positively correlated with HIF-2α, HIF-1α, and ERFE in the CKD patients injected with erythropoietin (EPO) ( p < 0.01), while serum hepcidin was positively correlated with HIF-2α and HIF-1α ( p < 0.01) in the patients not injected with EPO. ④ Multivariate linear regression analysis showed that HIF-1α, (β = 4.36, p < 0.01), serum ferritin(SF) (β = 0.13, p < 0.01), and HIF-2α (β = 0.66, p < 0.01) were significantly correlated with hepcidin. Conclusion HIF-1α, HIF-2α, and SF are factors which have an effect on hepcidin in patients with CKD stages 3–5 and renal anemia. The increase of HIF-1α, HIF-2α, and ERFE does not seem to inhibit the increase of hepcidin.

Morphological Description of Den-3 Virus Infection Cells through EDTA Blood Feed In Aedes Aegypti Mosquitoe

Mosquito Ae. aegypti is the main dengue virus vector which causes the virus to grow and develop properly. The Artificial Membrane Feeding (AMF) method is a method of indirectly transmitting the dengue virus in the Ae. aegypti. This method uses blood feed with EDTA anticoagulant and DEN-3 virus in the insectarium. One way to detect the dengue virus found in Ae mosquito cells. aegypti is an immunohistochemical Streptavidin Biotin Peroxidase Complex (SBPC). In this method, the morphological picture of DEN-3 virus infection cells will be seen through the heparin anticoagulant blood feed using the AMF method. Anticoagulants function to slow the blood clotting to feed the Ae mosquitoes. Aegypti. The aim of this study was to determine the Positive Infection Rate of DEN-3 virus and the morphological picture of DEN-3 virus infection cells using the AMF method of EDTA anticoagulant blood feed through SBPC immunocytochemical examination of Ae mosquitoes. aegypti. The design method of this study was experimental, infected Ae.aegypti mosquitoes using anticoagulant blood containing DEN-3 virus orally as an infectious sample. Negative control used Culex Sp mosquitoes and positive control used adult Ae.aegypti mosquitoes which were infected with DEN-3 virus by injection. Detection of DEN-3 virus in Ae.aegypti mosquitoes through SBPC immunositochemical examination. Results of Positive Infection Rate for Ae.aegypti mosquitoes with EDTA anticoagulant as much as 23.20% through SBPC immunocytochemical examination with a picture of cell morphology of DEN-3 virus infection. The conclusion of SBPC immunositochemical examination showed positive cell morphology of DEN-3 virus infection with EDTA anticoagulant in Ae.aegypti mosquitoes fed by human blood orally through the AMF method.

Prognostic significance of Ezrin expression in colorectal carcinoma

Objectives This study aimed to evaluate the immunohistochemical expression of Ezrin in various histopathological types of colorectal carcinoma, and to examine its association with various clinicopathological features and so evaluate its prognostic significance. Background Colorectal cancer is considered one of the most common cancers worldwide, it developed because of the cumulative effect of sequential genetic alterations. Thus, more extensive studies are needed to demonstrate the molecular mechanism of colorectal cancer initiation and progression. Ezrin gene has an important role in cell movement, migration, mitosis, and other physiological functions, and is involved in maintaining cell structure and cell motility. Its expression correlates with many human malignancies. Patients and methods Ezrin antibody immunostaining was performed using the avidin–biotin–peroxidase complex method in 204 cases of primary colorectal carcinomas, including 118 (57.8%) cases of adenocarcinoma, 56 (27.5%) cases of a mucoid carcinoma, and 30 (14.7%) cases of signet ring cell carcinoma. Results Cytoplasmic Ezrin expression was significantly higher in lymph-node-positive cases (P Conclusion High Ezrin expression is considered a poor prognostic factor for colorectal adenocarcinoma.

Screening of Fv Antibodies with Specific Binding Activities to Monosodium Urate and Calcium Pyrophosphate Dihydrate Crystals for the Diagnosis of Gout and Pseudogout.

To date, medical diagnosis of gout and pseudogout has been performed by observing the crystals in the joint fluid of patients under a polarized microscope. Conventional diagnostic methods using a polarized microscope have disadvantages, such as time-consuming analysis, a high false negative rate, and difficulty in distinguishing gout with monosodium urate (MSU) crystals and pseudogout with calcium pyrophosphate dihydrate (CPPD) crystals in synovial fluids. In this study, a chromogenic assay for the diagnosis of gout and pseudogout, without the requirement of a polarized microscope and trained experts, was proposed using Fv antibodies with specific binding activities to MSU and CPPD crystals. The IgG VH chain Fv library with randomized complementarity-determining region 3 (CDR3) region was expressed on the outer membrane of Escherichia coli using autodisplay technology. The target Fv antibodies with binding activity to MSU and CPPD crystals were screened from the autodisplayed Fv library on the E. coli outer membrane, and five clones were selected. On the basis of the binding properties of the screened Fv antibodies, peptides with the selected clone of amino acid sequences of the CDR3 region (15 residues) were chemically synthesized. The binding properties of the synthetic peptides with amino acid sequences of CDR3 regions from the selected clones were analyzed using fluorescence imaging and flow cytometry, and the affinity constants (Kd) of each peptide for binding to MSU and CPPD crystals were calculated by fitting based on the isotherm model. A chromogenic assay configuration for gout and pseudogout was developed using synthetic peptides. In this chromogenic assay, synthetic peptides labeled with biotin and streptavidin-horseradish peroxidase (HRP) complex were used, and crystal detection was possible using a chromogenic reaction between HRP and a chromogenic substrate (TMB). Finally, gout and pseudogout were diagnosed by detecting MSU and CPPD crystals in the synovial fluid in the concentration range of 0-300 μg/mL.

Evaluation of MMP-9 and iNOS expressions in sheep with encephalitic listeriosis

This study aimed to correlate Matrix Metalloproteinase-9 and iNOS expressions with the severity of histopathological findings in tissue samples taken from sheep with encephalitic listeriosis. Thus, the role of these molecules in the pathogenesis of the disease can be elucidated. After systemic necropsy, tissue samples of adult sheeps with meningoencephalitis were investigated by the culture, histopathological and immunohistochemical methods in the presence of Listeria spp. isolation from tissues was performed in accordance with the USDA-FSIS method with some modifications. Tissue samples were fixed in a 10% buffered formaldehyde solution. Following routine procedures, tissue sections at 5 μm were stained with Hematoxylin and Eosin, investigated under light microscope and photographed. Immunohistochemical staining was performed on the tissues using the avidin-biotin immune peroxidase complex method. Listeria spp. were obtained in 20 (83.3%) of 24 tissue samples with the presence of bright grey-black centred smooth colonies on Listeria Selective Agar and identified as Listeria monocytogenes through the phenotypically supportive tests. Liquefaction necrosis, purulent meningoencephalitis, perivascular cuffing, microabscesses and glial nodules were the most important histopathological findings. MMP-9 immunpositive reactions were observed in the cytoplasm of microglial cells and neurons in areas where inflammatory and necrotic areas are concentrated in medulla oblongata and pons. In perivascular cuffing areas, immune reactions in endothelial cells were detected. We detected iNOS positive reactions in the medulla oblangata and pons, especially in inflammatory cells in the microabscesses. Consequently, a positive correlation (p < 0.05) was found between MMP-9 expression and the severity of histopathological findings in sheep with encephalitic listeriosis. expression. In addition, we found that iNOS expression increased in parallel with the increase in MMP-9

Unraveling cardiolipin-induced conformational change of cytochrome c through H/D exchange mass spectrometry and quartz crystal microbalance

Cardiolipin (CL), a crucial component in inner mitochondrial membranes, interacts with cytochrome c (cyt c) to form a peroxidase complex for the catalysis of CL oxidation. Such interaction is pivotal to the mitochondrial regulation of apoptosis and is affected by the redox state of cyt c. In the present study, the redox-dependent interaction of cyt c with CL was investigated through amide hydrogen/deuterium exchange coupled with mass spectrometry (HDXMS) and quartz crystal microbalance with dissipation monitoring (QCM-D). Ferrous cyt c exhibited a more compact conformation compared with its ferric form, which was supported by the lower number of deuterons accumulated and the greater amplitude reduction on dissipation. Upon association with CL, ferrous cyt c resulted in a moderate increase in deuteration, whereas the ferric form caused a drastic increase of deuteration, which indicated that CL-bound ferric cyt c formed an extended conformation. These results were consistent with those of the frequency (f) − dissipation (D) experiments, which revealed that ferric cyt c yielded greater values of |ΔD/Δf| within the first minute. Further fragmentation analysis based on HDXMS indicated that the effect of CL binding was considerably different on ferric and ferrous cyt c in the C-helix and the Loop 9–24. In ferric cyt c, CL binding affected Met80 and destabilized His18 interaction with heme, which was not observed with ferrous cyt c. An interaction model was proposed to explain the aforementioned results.

Recently Approved and Under Investigation Drugs for Treating Patients with Heart Failure.

Heart Failure (HF) represents a leading cause of morbidity and mortality worldwide. Despite the recent advances in the treatment of this condition, patients´ prognosis remains unfavorable in most cases. Sacubitril/valsartan and ivabradine have been recently approved to improve clinical outcomes in patients with HF with reduced ejection fraction. Drugs under investigation for treating patients with HF encompass many novel mechanisms including vasoactive peptides, blocking inflammatory-mediators, natriuretic peptides, selective non-steroidal mineralocorticoid-receptor antagonists, myocardial β3 adrenoreceptor agonists, inhibiting the cytochrome C/cardiolipin peroxidase complex, neuregulin-1/ErbB signaling and inhibiting late inward sodium current. The aim of this manuscript is to review the main drugs under investigation for the treatment of patients with HF and give perspectives for their implementation into clinical practice.

Cerebrospinal fluid-contacting neurons affect the expression of endogenous neural progenitor cells and the recovery of neural function after spinal cord injury

Objective To explore the relationship between cerebrospinal fluid-contacting neurons (CSF-cNs) and endogenous neural progenitor cells (ENPCs) and whether CSF-cNs are involved in nerve repair after spinal cord injury (SCI). Methods Cholera toxin B-horseradish peroxidase complex (CB-HRP) and cholera toxin B conjugated with saporin (CB-SAP) were injected into the lateral ventricles of spinal cord injured rats to mark and destroy the CSF-cNs. Then the rats in the experimental group were injured by SCI. Observe the content and co-expression of CSF-cNs and ENPCs in rats of each group, and observe the recovery of motor function after SCI in each group. Results After the destruction of CSF-cNs, the number of ENPCs decreased significantly in the long term after the surgery, and the recovery of motor function also deteriorated as compared to the group with intact CSF-cNs. Meanwhile some cells in the spinal cord express both the biological marker of CSF-cNs and ENPCs. Conclusion This study shows that the population of ENPCs and motor function recovery in SCI rats declined after the destruction of CSF-cNs, suggesting that CSF-cNs affect the ENPCs population and may be involved in the recovery of neural function after SCI.

Expression of PCNA, MMP-9, p53, Bax and Bcl-2 in canine transmissible venereal tumors

In this study, we aimed to evaluate the proliferative, metastatic and apoptotic capacities of TVT cases, which are in various phases of development, by using immunohistochemical markers. The material of this study consisted of twelve female and six male dogs diagnosed with TVT brought to our department between 2007 and 2020 years. Diff-quick staining was applied to the smear taken from tumoral masses for cytological examinations. Tumoral tissues from dogs were fixed in a 10% neutral buffered formaldehyde solution. After routine tissue procedures Hematoxylin & Eosin stain was applied to the sections. Tissue sections were investigated under a light microscope and photographed. Immunohistochemical staining was performed on the tissues using the avidin-biotin immune peroxidase complex method. As a result of macroscopic, cytological and histopathological examinations, TVT positive cases were divided into three according to their developmental stages. While the expression of PCNA, MMP-9, mutant p53 and Bcl-2 increased significantly in progressive cases compared to regressive and stable cases, Bax expression increased significantly in regressive cases compared to progressive and stable cases. In conclusion, we thought that the mentioned markers are very useful for understanding the prognosis of TVT, the tumor aggressiveness and the survival of the malignant cells.

Pengaruh Lama Penyimpanan Telur terhadap Transovarial Infection Rate Virus DEN-3 pada Nyamuk Aedes aegypti

Transovarial transmission of dengue virus is important phenomenon causes dengue virus survival during inter-epidemic period. The purpose of this study was to analyze effect of egg storage duration on DEN-3 virus Transovarial Infection Rate (TIR) in Aedes aegypti mosquitoes. This was laboratory research with experimental design. Ae. aegypti was infected with the DEN-3 virus per-orally and allowed to undergo its gonotrophic cycle. Eggs stored at room temperature (25±3oC) and relative humidity (70±5%) for 0, 1, 2 and 3 months. Hatched eggs then colonized until become adult mosquito. Sample was taken from colonies at 4, 8, 16 days old. Headsquash of mosquitoes was preparedwith Streptavidin Biotin Peroxidase Complex (SBPC) immunocytochemical method using DSSE 10 monoclonal antibody and detected the presence virus antigen. Transovarial infection rate was determined by counting percentage of positive samples. Data were analyzed by linear regression test. Result of this study showed that in 4, 8, 16 samples showed lowest TIR on 0 month storage (33,33%, 33,33%; 40,00%) and highest TIR observed on 2 month storage (76,67%; 66,67%, 76,67%). Statistical test showed significantly different result (p = 0,013) with α = 0,05 and R2 = 0,476. The duration of egg storage effect on DEN-3 virus TIR in Ae. aegypti mosquito with 47.6% contribution.