The Role of Matrix Metalloproteinases in Thoracic Aortic Disease: Are They Indicators for the Pathogenesis of Dissections?

The pathogenesis of aortic aneurysm and dissection continues to be under discussion. Extracellular matrix (ECM) remodeling processes in the aortic wall are hypothesized to be involved in the development of the disorders. Therefore, in a histological study, we investigated the expression of metalloproteases 1 and 9 (MMP1 and MMP9) and their inhibitors (TIMP 1 and TIMP 2) in cardiac surgery patients. In parallel, we studied the aortic roots by echocardiography. Clinical reports of 111 patients (30 women and 81 men) who suffered from aortic aneurysms and aortic dissection were evaluated and studied by transesophageal echocardiography. Seven patients who had coronary heart disease served as "healthy controls". All patients underwent the necessary surgical procedure according to the diagnosed aortic disease in the period from 2007 to 2015. A tissue sample of the aortic biopsies was collected from each patient during surgery. Immunohistochemical staining was performed for MMP1 and MMP9 and TIMP1 and TIMP2 as well. Vascularization was monitored by a CD 31 antibody. In direct comparison, the expressions are not homogeneous. We found the smallest changes in the intima area at all. TIMP 1 and TIMP 2 distribution increases from the lumen of the vessel outward in the wall layers of the aorta. In the case of arteriosclerotic changes, intima had a capillarization, but not in the media. An opposite pattern was found in the dissected aortas. There are differences in the vascularization between the aneurysm and dissection and the different layers, respectively. A different remodeling process of the ECM in comparison to the vascular layers must be hypothesized. Reading the patterns of staining and with regard to the known inhibitory effect of MMP9 on ECM remodeling, but especially TIMP 2 on neoangiogenesis, disturbed nutrition, and dysfunctional vasa vasorum remodeling must be assumed as causes of dissection.

The ratio of cytochrome c oxidase subunit 4 isoform 4I1 and 4I2 mRNA is changed in permanent atrial fibrillation.

AIMS: The conditions of hypoxia are suggested to induce permanent atrial fibrillation (AF). The regulation of COX4I2 and COX4I1 depends on oxygen availability in tissues. A role of COX4I2 in the myocardium of AF patients is supposed for pathogenesis of AF and subsequent alterations in the electron transfer chain (ETC) under hypoxia. METHODS AND RESULTS: In vitro, influence of hypoxia on HeLa 53 cells was studied and elevated parts of COX 4I2 were confirmed. Myocardial biopsies were taken ex vivo from the patients' Right Atria with SR (n = 31) and AF (n = 11), respectively. RT- PCR for mRNA expresson, mitochondrial respiration by polarography and the protein content of cytochrome c oxidase (CytOx) subunit 4I1 and CytOx subunit 4I2 by ELISA were studied. Clinical data were correlated to the findings of gene expressions in parallel. Patients with permanent AF had a change in isoform 4I2/4I1 expression along with a decrease of isoform COX 4I1 expression. The 4I2/4I1 ratio of mRNA expression was increased from 0.630 to 1.058 in comparison. However, the protein content of CytOx subunit 4 was much lower in the AF group, whereas the respiration/units enzyme activity in both groups remained the same. CONCLUSIONS: This study describes a possible molecular correlate for the development of AF. Due to the known functional significance of COX 4I2, mitochondrial dysfunction can be assumed as a part of the pathogenesis of AF.

Influence of cardioplegic solution on incidence of delirium after CABG surgery: Use of Calafiore blood cardioplegia versus HTK - Bretschneider - solution in a single-center retrospective analysis from 2017 to 2021.

BACKGROUND: Purpose of the present study is an evaluation of postoperative incidence for delirium after coronary artery bypass surgery (CABG). Study addressed whether application of Histidine-Tryptophan-Ketoglutarate (HTK) solution (Bretschneider) or blood cardioplegia (Calafiore) is associated with increased of postoperative delirium cases. MATERIALS AND METHODS: In a retrospective, single center evaluation a total number of 273 patients were enrolled in the study from January 2017 to October 2021. There were 124 patients assigned to the Calafiore group blood cardioplegic solution (BCC) and 149 patients were included in the Bretschneider group (HTK). The primary endpoint was the postoperative delirium rate in its frequency of occurrence. Definition of the dilirium status was performed using the Confusion Assessment Method in the Intensive Care Unit (CAM-ICU) score during the first three postoperative days. Secondary endpoints were the time intervals of intensive care duration of stay, mechanical ventilation, total extracorporeal circulation, ischemia and reperfusion. Serum levels of the electrolytes Sodium, Potassium, ionized Calcium, and Chloride were monitored. RESULTS: Although no significant difference in delirium status between the groups were noticed, on third postoperative day, delirium rate dependent on cardioplegia solution used (HTK 12.0%; BCC 3.0%; p = .024) and duration of intensive care stay differed (HTK 4.5 vs. BCC 3.0 days; p = .001). Although Ischemic time (HTK 73.0 vs. BCC 83.0 min; p < .001) and reperfusion time (HTK 35.0 vs. Calafiore 24.0 min; p < .001) were extended in the BCC group less cases of delirium were diagnosed. Serum sodium levels after HTK cardioplegic infusion were decreased (HTK 129.68 vs. BCC 138.96 mmol/l; p < .001). The significant difference persists up to the hundredth extracorporeal circulation circuit min (p = .005). CONCLUSION: The present data suggest an impact of the cardioplegic solution used upon postoperative delirium rates. Optimization of cardiac arrest protocols is needed. Present data encourage further prospective studies regarding the impact of cardioplegic solutions on electrolyte imbalance for postoperative delirium rates in CABG surgery.

Enhanced Bioaccessibility of Microencapsulated Puerarin Delivered by Pickering Emulsions Stabilized with OSA-Modified Hydrolyzed Pueraria montana Starch: In Vitro Release, Storage Stability, and Physicochemical Properties.

Puerarin is a bioactive flavonoid isolated from Kudzu roots that possesses numerous health benefits. However, its poor bioavailability and existing complex delivery systems with safety issues are challenging tasks for its incorporation into functional foods. Preparing modified-starch-stabilized Pickering emulsions containing microencapsulated puerarin with improved bioaccessibility was the key objective of the present research work. Acid-hydrolyzed high-amylose Pueraria montana starch (PMS) was modified with octenyl succinic anhydride (OSA) and evaluated as an emulsifier to prepare emulsions. The FTIR, SEM, and XRD results showed that PMS was successfully modified. Furthermore, the emulsification index (EI), mean droplet size, and ζ-potential values showed that modified starch with a higher degree of substitution (DS) enhanced the storage stability of emulsions. Similarly, the retention degree and encapsulation efficiency results of puerarin proved the assumption after storage of 16 d. The Pickering emulsions also helped in the controlled release of microencapsulated puerarin in vitro. The study outcomes proved that Pickering emulsions stabilized with OSA-modified PMS have promising applicability in functional foods as efficient food-grade delivery systems, enhancing oral supplementation and accessibility of puerarin.

The ABCT31 Transporter Regulates the Export System of Phenylacetic Acid as a Side-Chain Precursor of Penicillin G in Monascus ruber M7.

The biosynthesis of penicillin G (PG) is compartmentalized, and the transportation of the end and intermediate products, and substrates (precursors) such as L-cysteine (L-Cys), L-valine (L-Val) and phenylacetic acid (PAA) requires traversing membrane barriers. However, the transportation system of PAA as a side chain of PG are unclear yet. To discover ABC transporters (ABCTs) involved in the transportation of PAA, the expression levels of 38 ABCT genes in the genome of Monascus ruber M7, culturing with and without PAA, were examined, and found that one abct gene, namely abct31, was considerably up-regulated with PAA, indicating that abct31 may be relative with PAA transportation. Furthermore the disruption of abct31 was carried out, and the effects of two PG substrate's amino acids (L-Cys and L-Val), PAA and some other weak acids on the morphologies and production of secondary metabolites (SMs) of Δabct31 and M. ruber M7, were performed through feeding experiments. The results revealed that L-Cys, L-Val and PAA substantially impacted the morphologies and SMs production of Δabct31 and M. ruber M7. The UPLC-MS/MS analysis findings demonstrated that Δabct31 did not interrupt the synthesis of PG in M. ruber M7. According to the results, it suggests that abct31 is involved in the resistance and detoxification of the weak acids, including the PAA in M. ruber M7.

Prediction of stroke reconvalescence after coronary bypass surgery indicated by CT scan parameters.

BACKGROUND: Stroke in the postoperative time course after heart surgery remains a serious risk. Cranial computer tomography (CCT) is the first line option to detect severe intracranial damage. However, only few data are available to predict neurological outcome. Using visual rating scales (VRSs), this study addresses reliability and effectivity to indicate neurological status and likelyhood of improvement. METHODS: In a single-center retrospective evaluation, 3719 patients underwent coronary bypass surgery. Because of a delayed recovery phase and neurologic deficits after cardiac surgery 109 patients had a cranial CT scan in the early postoperative period. The incidence of clinically relevant findings within the imaging was rated by an experienced neuroradiologist using two VRS, that is, the age-related white matter changes (ARWMCs) and the Mendes-Ribeiro visual rating scale (MRVRS). Both are computer-assisted measurement schemes to detect stroke-related intracranial damage. Follow-up was investigated with regard to clinical outcome and patient-related risk profiles. RESULTS: Of 109 patients with postoperative cranial CT scans due to prolonged recovery phases or proven neurological damage 44.5% had one cerebral defect in CCT imaging scans only. The others showed multiple defects. During hospital stay, 92.3% experienced neurological improvement exposing reduced ARWMC, while 7.1% had no improvement and correlating high scores. Of both scales, the ARWMC-VRS demonstrated superior accuracy and discrimination. The preoperative ejection fraction (EF), arteriosclerotic degeneration of carotid arteries, and reduced glomerular filtration rate were found to have a high correlation (r = 0.0005) with the latter group. In-hospital mortality of this cohort was 8.18%. CONCLUSION: Both the ARWMC and MRVRS were found to be appropriate. They reliably discriminate the groups of stroke patients after coronary artery bypass grafting (CABG)  in the analysis of CCT images. When applied at the onset of neurological symptoms both scales are able to predict neurological reconvalescence upon hospital dismission. The ARWMC scale appeared superior as it demonstrated better accuracy and discrimination. The use of both VRS in patients with suspected stroke after CABG surgery can give insightful information toward a progression of neurological dysfunction or postoperative improvement.

Cytochrome c Oxidase Inhibition by ATP Decreases Mitochondrial ROS Production.

This study addresses the eventual consequence of cytochrome c oxidase (CytOx) inhibition by ATP at high ATP/ADP ratio in isolated rat heart mitochondria. Earlier, it has been demonstrated that the mechanism of allosteric ATP inhibition of CytOx is one of the key regulations of mitochondrial functions. It is relevant that aiming to maintain a high ATP/ADP ratio for the measurement of CytOx activity effectuating the enzymatic inhibition as well as mitochondrial respiration, optimal concentration of mitochondria is critically important. Likewise, only at this concentration, were the differences in ΔΨm and ROS concentrations measured under various conditions significant. Moreover, when CytOx activity was inhibited in the presence of ATP, mitochondrial respiration and ΔΨm both remained static, while the ROS production was markedly decreased. Consubstantial results were found when the electron transport chain was inhibited by antimycin A, letting only CytOx remain functional to support the energy production. This seems to corroborate that the decrease in mitochondrial ROS production is solely the effect of ATP binding to CytOx which results in static respiration as well as membrane potential.

Role of matrix metalloproteinases in mitral valve regurgitation: Association between the of MMP-1, MMP-9, TIMP-1, and TIMP-2 expression, degree of mitral valve insufficiency, and pathologic etiology.

BACKGROUND: The pathogenesis of mitral valve insufficiency is not yet fully understood. Several studies stressed the role of matrix metalloproteinases (MMPs) in the emergence of valvular pathologies. The primary objective of the present study is to analyze the role of selected MMPs and their inhibitors in mitral valve insufficiency. PATIENTS AND METHODS: Eighty patients (33 female/47 male, mean age 67 years) underwent cardiopulmonary bypass surgery for mitral valve reconstruction between 2007 and 2015. All patients suffered from mitral insufficiency (MI) Stages iii and iv. When tissue resection was acquired specimens were taken immediately frozen and used for histological examination. Expression of MMP-1, MMP-9, tissue inhibitor of metalloproteinase (TIMP)-1, and TIMP-2 was examined immunohistochemically and distribution was analyzed in regard to preoperative clinical, echocardiographic, and histopathological findings. RESULTS: A clear correlation between the MMP expression and the MI degree of severity could be shown. The expression of MMPs proved to be high in relation to mild insufficiencies and relatively weak in the case of severe ones. Additionally, the etiology of the MI was considered in the analysis and a significant difference in the expression of MMPs between the mitral valves with endocarditis and the ones featuring a degenerative disease could be shown. Within the group of valves with degenerative diseases, no significant difference could be established between the subgroups (myxoid and sclerosed valves). CONCLUSION: The increased expression of MMPs and their inhibitors in mild insufficiencies could prove that the molecular changes in the valve precede the macroscopical and thus the echocardiographically diagnosable changes. Hence, new options for early diagnosis and therapy of MIs should be examined in further studies, respectively. Herein, the correlation of the MMP blood levels with MMP tissue expression should be addressed for surgical therapeutical decisions.

Urinary TIMP-2 and IGFBP-7 protein levels as early predictors of acute kidney injury after cardiac surgery.

BACKGROUND: Acute kidney injury (AKI) is a frequent complication associated with on-pump cardiac surgery. Early recognition may alter their prognosis. Therefore, the urinary concentrations of TIMP-2 (tissue inhibitor of metalloproteinases-2) and IGFBP7 (insulin-like growth factor-binding protein) as predictors for AKI were studied. METHODS: Repetitive blood and urine samples were collected consecutively from 50 patients. Demographic, intra-, and postoperative data were recorded prospectively. To calculate the production of the TIMP-2 and IGFBP-7 protein concentrations, urinary samples were taken preoperatively, intraoperatively at 30 and 60 min after aortic clamping and at 0, 6, 12, and 24 h after admission to the intensive care unit (ICU). RESULTS: AKI occurred in 14 patients (28%), all of them at Kidney Disease: Improving Global Outcomes stage 1. Predictive value for [TIMP-2] × [IGFBP7] was shown at 0 and 24 h after admission to ICU. At 0 h, the sensitivity was 84.6% and the specificity 55.6% for an ideal calculated cutoff at 0.07. After 24 h, the ideal cutoff amounted to 0.35 with a sensitivity of 53.8% and a specificity of 88.2%. The receiver operating characteristic curves demonstrated areas under the curve of 0.725 and 0.718. The suggested cutoffs of 0.3 and 2.0 could not be confirmed. The serum creatinine was reached to the peak median within 48 h after admission to ICU. CONCLUSION: Postoperative risk assessment for the development of AKI can be established by [ TIMP - 2 ] × [ IGFBP 7 ] . Previously suggested cutoff values could not be confirmed. A correlation with urinary dilution parameters may enable the identification of more universal cutoffs.

Assessment of In Vitro and In Vivo Bioremediation Potentials of Orally Supplemented Free and Microencapsulated Lactobacillus acidophilus KLDS Strains to Mitigate the Chronic Lead Toxicity.

Lead (Pb) is a pestilent and relatively nonbiodegradable heavy metal, which causes severe health effects by inducing inflammation and oxidative stress in animal and human tissues. This is because of its significant tolerance and capability to bind Pb (430 mg/L) and thermodynamic fitness to sequester Pb in the Freundlich model (R 2 = 0.98421) in vitro. Lactobacillus acidophilus KLDS1.1003 was selected for further in vivo study both in free and maize resistant starch (MRS)-based microencapsulated forms to assess its bioremediation aptitude against chronic Pb lethality using adult female BALB/c mice as a model animal. Orally administered free and microencapsulated KLDS 1.1003 provided significant protection by reducing Pb levels in the blood (127.92 ± 5.220 and 101.47 ± 4.142 µg/L), kidneys (19.86 ± 0.810 and 18.02 ± 0.735 µg/g), and liver (7.27 ± 0.296 and 6.42 ± 0.262 µg/g). MRS-microencapsulated KLDS 1.0344 improved the antioxidant index and inhibited changes in blood and serum enzyme concentrations and relieved the Pb-induced renal and hepatic pathological damages. SEM and EDS microscopy showed that the Pb covered the surfaces of cells and was chiefly bound due to the involvement of the carbon and oxygen elements. Similarly, FTIR showed that the amino, amide, phosphoryl, carboxyl, and hydroxyl functional groups of bacteria and MRS were mainly involved in Pb biosorption. Based on these findings, free and microencapsulated L. acidophilus KLDS 1.0344 could be considered a potential dietetic stratagem in alleviating chronic Pb toxicity.

Mitochondrial respiration is controlled by Allostery, Subunit Composition and Phosphorylation Sites of Cytochrome c Oxidase: A trailblazer's tale - Bernhard Kadenbach.

In memoriam of Bernhard Kadenbach: Although the main focus of his research was the structure, function, and regulation of mitochondrial cytochrome c oxidase (CytOx), he earlier studied the mitochondrial phosphate carrier and found an essential role of cardiolipin. Later, he discovered tissue-specific and developmental-specific protein isoforms of CytOx. Defective activity of CytOx is found with increasing age in human muscle and neuronal cells resulting in mitochondrial diseases. Kadenbach proposed a theory on the cause of oxidative stress, aging, and associated diseases stating that allosteric feedback inhibition of CytOx at high mitochondrial ATP/ADP ratios is essential for healthy living while stress-induced reversible dephosphorylation of CytOx results in the formation of excessive reactive oxygen species that trigger degenerative diseases. This article summarizes the main discoveries of Kadenbach related to mammalian CytOx and discusses their implications for human disease.

Cholate Disrupts Regulatory Functions of Cytochrome c Oxidase.

Cytochrome c oxidase (CytOx), the oxygen-accepting and rate-limiting enzyme of mitochondrial respiration, binds with 10 molecules of ADP, 7 of which are exchanged by ATP at high ATP/ADP-ratios. These bound ATP and ADP can be exchanged by cholate, which is generally used for the purification of CytOx. Many crystal structures of isolated CytOx were performed with the enzyme isolated from mitochondria using sodium cholate as a detergent. Cholate, however, dimerizes the enzyme isolated in non-ionic detergents and induces a structural change as evident from a spectral change. Consequently, it turns off the "allosteric ATP-inhibition of CytOx", which is reversibly switched on under relaxed conditions via cAMP-dependent phosphorylation and keeps the membrane potential and ROS formation in mitochondria at low levels. This cholate effect gives an insight into the structural-functional relationship of the enzyme with respect to ATP inhibition and its role in mitochondrial respiration and energy production.

Multiple Mechanisms Regulate Eukaryotic Cytochrome C Oxidase.

Cytochrome c oxidase (COX), the rate-limiting enzyme of mitochondrial respiration, is regulated by various mechanisms. Its regulation by ATP (adenosine triphosphate) appears of particular importance, since it evolved early during evolution and is still found in cyanobacteria, but not in other bacteria. Therefore the "allosteric ATP inhibition of COX" is described here in more detail. Most regulatory properties of COX are related to "supernumerary" subunits, which are largely absent in bacterial COX. The "allosteric ATP inhibition of COX" was also recently described in intact isolated rat heart mitochondria.

Stress-mediated generation of deleterious ROS in healthy individuals - role of cytochrome c oxidase.

Psychosocial stress is known to cause an increased incidence of coronary heart disease. In addition, multiple other diseases like cancer and diabetes mellitus have been related to stress and are mainly based on excessive formation of reactive oxygen species (ROS) in mitochondria. The molecular interactions between stress and ROS, however, are still unknown. Here we describe the missing molecular link between stress and an increased cellular ROS, based on the regulation of cytochrome c oxidase (COX). In normal healthy cells, the "allosteric ATP inhibition of COX" decreases the oxygen uptake of mitochondria at high ATP/ADP ratios and keeps the mitochondrial membrane potential (ΔΨm) low. Above ΔΨm values of 140 mV, the production of ROS in mitochondria increases exponentially. Stress signals like hypoxia, stress hormones, and high glutamate or glucose in neurons increase the cytosolic Ca2+ concentration which activates a mitochondrial phosphatase that dephosphorylates COX. This dephosphorylated COX exhibits no allosteric ATP inhibition; consequently, an increase of ΔΨm and ROS formation takes place. The excess production of mitochondrial ROS causes apoptosis or multiple diseases.

Transfigured Morphology and Ameliorated Production of Six Monascus Pigments by Acetate Species Supplementation in Monascus ruber M7.

Monascus species have been used for the production of many industrially and medically important metabolites, most of which are polyketides produced by the action of polyketide synthases that use acetyl-CoA and malonyl-CoA as precursors, and some of them are derived from acetate. In this study the effects of acetic acid, and two kinds of acetates, sodium acetate and ammonium acetate at different concentrations (0.1%, 0.25% and 0.5%) on the morphologies, biomasses, and six major Monascus pigments (MPs) of M. ruber M7 were investigated when M7 strain was cultured on potato dextrose agar (PDA) at 28 °C for 4, 8, 12 days. The results showed that all of the added acetate species significantly affected eight above-mentioned parameters. In regard to morphologies, generally the colonies transformed from a big orange fleecy ones to a small compact reddish ones, or a tightly-packed orange ones without dispersed mycelia with the increase of additives concentration. About the biomass, addition of ammonium acetate at 0.1% increased the biomass of M. ruber M7. With respect to six MPs, all acetate species can enhance pigment production, and ammonium acetate has the most significant impacts. Production of monascin and ankaflavin had the highest increase of 11.7-fold and 14.2-fold in extracellular contents at the 8th day when 0.1% ammonium acetate was supplemented into PDA. Intracellular rubropunctatin and monascorubrin contents gained 9.6 and 6.46-fold at the 8th day, when 0.1% ammonium acetate was added into PDA. And the extracellular contents of rubropunctamine and monascorubramine were raised by 1865 and 4100-fold at the 4th day when M7 grew on PDA with 0.5% ammonium acetate.

Genetic Modification of mfsT Gene Stimulating the Putative Penicillin Production in Monascus ruber M7 and Exhibiting the Sensitivity towards Precursor Amino Acids of Penicillin Pathway.

: The biosynthesis of penicillin G (PG) is compartmentalized, which forces penicillin and its intermediates to cross the membrane barriers. Although many aspects around the penicillin intermediates traffic system remain unclosed, the transmembrane transporter protein involvement has been only predicted. In the present work, detection of PG and isopenicillin N (IPN) in Monascus ruber M7 was performed and functions of mfst gene as a transporter were investigated by the combination of gene deletion (Δmfst) complementation (ΔmfsT::mfsT) and overexpression (M7::PtrpC-mfsT). While, the feeding of PG pathway precursor side chain and amino acids, i.e., phenylacetic acid, D-valine, and L-cysteine was performed for the interpretation of mfsT gene role as an intermediate transporter. The results showed that, the feeding of phenylacetic acid, D-valine, and L-cysteine possessed a significant effect on morphologies, secondary metabolites (SMs) production of all above-mentioned strains including M. ruber M7. The results of UPLC-MS/MS revealed that, ΔmfsT interrupt the penicillin G (PG) production in M. ruber M7 by blocking the IPN transportation, while PG and IPN produced by the ΔmfsT::mfsT have been recovered the similar levels to those of M. ruber M7. Conclusively, these findings suggest that the M. ruber M7 is, not only a PG producer, but also, indicate that the mfsT gene is supposed to play a key role in IPN intermediate compound transportation during the PG production in M. ruber M7.

Reversible dimerization of cytochrome c oxidase regulates mitochondrial respiration.

Almost all energy consumed by higher organisms, either in the form of ATP or heat, is produced in mitochondria by respiration and oxidative phosphorylation through five protein complexes in the inner membrane. High-resolution x-ray analysis of crystallized cytochrome c oxidase (CytOx), the final oxygen-accepting complex of the respiratory chain, isolated by using cholate as detergent, revealed a dimeric structure with 13 subunits in each monomer. In contrast, CytOx isolated with non-ionic detergents appeared to be monomeric. Our data indicate in vivo a continuous transition between CytOx monomers and dimers via reversible phosphorylation. Increased intracellular calcium, as a consequence of stress, dephosphorylates and monomerises CytOx, whereas cAMP rephosphorylates and dimerises it. Only dimeric CytOx exhibits an "allosteric ATP-inhibition" which inhibits respiration at high cellular ATP/ADP-ratios and could prevent oxygen radical formation and the generation of diseases.

Protamine Sulfate Induces Mitochondrial Hyperpolarization and a Subsequent Increase in Reactive Oxygen Species Production.

Protamine sulfate (PS) is widely used in heart surgery as an antidote for heparin, albeit its pharmacological effects are not fully understood and applications are often accompanied by unwanted side effects. Here we show the effect of PS on mitochondrial bioenergetics profile resulting in mitochondrial reactive oxygen species (ROS) production. Polarographic measurements were performed in parallel to membrane potential and ROS measurements by FACS analyzer using tetramethylrhodamine ethyl ester and MitoSOX fluorescent dyes, respectively. PS inhibited intact rat heart mitochondrial respiration (stimulated by ADP) to 76% (P < 0.001) from the baseline of 51.6 ± 6.9 to 12.4 ± 2.3 nmol O2⋅min-1⋅ml-1 The same effect was found when respiration was inhibited by antimycin A (101.0 ± 8.9 vs. 38.0 ± 9.9 nmol O2 ⋅min-1⋅ml-1, P < 0.001) and later stimulated by substrates of cytochrome oxidase (CytOx) i.e., ascorbate and tetramethyl phenylene diamine, suggesting that PS exerted its effect through inhibition of CytOx activity. Furthermore, the inhibition of mitochondrial respiration by PS was concentration dependent and accompanied by hyperpolarization of the mitochondrial membrane potential (Δψ m), i.e., 18% increase at 50 µg/ml and an additional 3.3% increase at 250 µg/ml PS compared with control. This effect was associated with a strong consequent increase in the production of ROS, i.e., 85% and 88.6% compared with control respectively. We propose that this excessive increase in ROS concentrations results in mitochondrial dysfunction and thus might relate to the "protamine reaction," contributing to the development of various cardiovascular adverse effects.

Assessment of the Antimicrobial Potentiality and Functionality of Lactobacillus plantarum Strains Isolated from the Conventional Inner Mongolian Fermented Cheese Against Foodborne Pathogens.

Lactobacillus plantarum are amongst the diversified lactic acid bacteria (LAB) species which are being utilized abundantly in the food industry. Numerous L. plantarum strains have been reported to produce several antimicrobial compounds. Diacetyl, hydrogen peroxide, organic acids, as well as bacteriocins can also be exemplified by a variable spectrum of actions. The current study was intended to conduct the screening and characterization of antimicrobial prospective of L.plantarum from traditional Inner Mongolian fermented hard cheese. Foodborne pathogens, Salmonellatyphimurium, Escherichia coli O157:H7, Listeria monocytogenes, and Staphylococcusaureus, were examined by using the Oxford cup technique and the mixed culture inhibition assays. The resulting analyses disclosed that L. plantarum KLDS1.0344 indicated broad antimicrobial spectrum against all selected pathogens as compared to other LAB used in this study. Additionally, the decrement of the pathogen population was observed up to 3.47 logs in mixed culture inhibition assays. L. plantarum KLDS 1.0344 acid production was recorded up to 71.8 ± 3.59 °D in mixed culture while antimicrobial particles released in cell free supernatants demonstrated bacteriocin-like characteristics showing substantial pH stability (2.0-6.0), proteolytic enzyme reduced the antibacterial activity (15.2 ± 0.6 mm-20.4 ± 0.8 mm), heat stability (20 min at 120 °C) against selected pathogens. Moreover, the spectrum range of antimicrobial peptides after the partial purification was decreased as compared to the crude bacteriocin-like compound. The SDS-PAGE analysis showed the molecular weight range of partially purified bacteriocin from 12 to 45 kDa. After analyzing the obtained data from the current experimentation showed that the capability of L. plantarum KLDS 1.0344 to oppose the pathogen growth in vitro relies on the occurrence of organic acids along with bacteriocin-like compounds proving L. plantarum KLDS 1.0344 as a potentially appropriate candidate as an alternative bio-control agent against foodborne pathogens.

Mitochondrial active and relaxed state respiration after heat shock mRNA response in the heart.

Induction of Heat Shock Proteins results in cytoprotection. Beneficial effect results from transcription and translational cellular components' involvement that defends metabolism and thus induce ischemic protection of the tissue. Mitochondrial respiration is also involved in stress- induced conditions. It is not a uniform process. Cytochrome c Oxidase (CytOx) representing complex IV of the Electron Transfer Chain (ETC) has a regulatory role for mitochondrial respiratory activity, which is tested in our study after hsp induction. Moreover, protein translation for mitochondrial components was probed by the detection of MT-CO1 for Subunit 1 of CytOx neosynthesis. Wistar rats were subjected to whole-body hyperthermia at 42.0-42.5 °C for 15 min followed by a normothermic recovery period. Heat shock response was monitored time dependent from LV biopsies of all control and heat treated animals with PCR-analysis for hsp 32, 60, 70.1, 70.2, 90 and MT-CO1 expression at 15, 30, 45, 60, 120 and 360 min recovery (n = 5 in each group), respectively. Enzymatic activity of CytOx were evaluated polarographically. High energy phosphates were detected by chromatographic analysis. The mRNA expression of MT-CO1 peaked at 60 min and was accompanied by hsp 32 (r = 0.457; p = 0.037) and hsp 70.2 (r = 0.615; p = 0.003) upregulation. With hsp induction, mitochondrial respiration was increased initially. Enzymatic activity reconciled from active into relaxed status wherein CytOx activity was completely inhibited by ATP. Myocardial ATP content increased from stress induced point i.e. < 1 µmol g-1 protein w/w to finally 1.5 ±â€¯0.53 µmol g-1 protein w/w at 120 min recovery interval. Hyperthermic, myocardial hsp- induction goes along with increased CytOx activity representing an increased "active" mitochondrial respiration. In parallel, de -novo holoenzyme assembly of CytOx begins as shown by MT-CO1 upregulation at 60 min recovery time crossing with a final return to the physiological "relaxed" state and ATP -inhibited respiration.