New insight into the morphological characteristics of patella cubiti.

Accessory bones in the elbow region are rare anatomical variations with important clinical significance as they can be misdiagnosed as pathological lesions. Usually, they are asymptomatic and found incidentally during X-ray examination in the context of trauma. Although these bones have been previously described, their development is not fully understood. The present case provides complex macroscopic, X-ray and histological descriptions of a sesamoid bone in the posterior region of the elbow-patella cubiti and the related surrounding structures. Moreover, this report indicates the presence of a well-defined syndesmosis between patella cubiti and the proximal ulna.

Depletion of vascular adaptive mechanisms in hypertension-induced injury of the heart and kidney.

BACKGROUND: Vascular endothelial growth factor (VEGF) is a signalling protein of critical importance for angiogenesis. In an effort to better understand its significance in hypertension-induced injury of the heart and kidney we aimed at studying the changes in its expression in an experimental model and correlated it with capillary density in the myocardium and the renal parenchyma. METHODS: We used two age groups of spontaneously hypertensive rats (6- and 12-month-old), indicative of early and advanced hypertension. VEGF expression was assessed and a semi-quantitative analysis of its immunoreactivity was conducted. Changes in capillary density in the myocardium and kidney were assessed for statistical significance and correlations with VEGF expression were established. RESULTS: We reported stronger VEGF expression in animals with early compared to advanced hypertension in all examined structures. Capillary density decreased significantly at age 12 months compared to 6 months and was significant in all examined structures. A positive correlation was established between capillary density and the expression of VEGF. CONCLUSION: These findings underscore the key significance of VEGF for compensatory angiogenesis and suggest that a statistically significant depletion of these vascular adaptive mechanisms is a major aspect in the cascade of hypertension-induced injury of the heart and kidney (Tab. 3, Fig. 26, Ref. 47).

Sensitive Versatile Fluorogenic Transmembrane Peptide Substrates for Rhomboid Intramembrane Proteases.

Rhomboid proteases are increasingly being explored as potential drug targets, but their potent and specific inhibitors are not available, and strategies for inhibitor development are hampered by the lack of widely usable and easily modifiable in vitro activity assays. Here we address this bottleneck and report on the development of new fluorogenic transmembrane peptide substrates, which are cleaved by several unrelated rhomboid proteases, can be used both in detergent micelles and in liposomes, and contain red-shifted fluorophores that are suitable for high-throughput screening of compound libraries. We show that nearly the entire transmembrane domain of the substrate is important for efficient cleavage, implying that it extensively interacts with the enzyme. Importantly, we demonstrate that in the detergent micelle system, commonly used for the enzymatic analyses of intramembrane proteolysis, the cleavage rate strongly depends on detergent concentration, because the reaction proceeds only in the micelles. Furthermore, we show that the catalytic efficiency and selectivity toward a rhomboid substrate can be dramatically improved by targeted modification of the sequence of its P5 to P1 region. The fluorogenic substrates that we describe and their sequence variants should find wide use in the detection of activity and development of inhibitors of rhomboid proteases.

Substrate binding and specificity of rhomboid intramembrane protease revealed by substrate-peptide complex structures.

The mechanisms of intramembrane proteases are incompletely understood due to the lack of structural data on substrate complexes. To gain insight into substrate binding by rhomboid proteases, we have synthesised a series of novel peptidyl-chloromethylketone (CMK) inhibitors and analysed their interactions with Escherichia coli rhomboid GlpG enzymologically and structurally. We show that peptidyl-CMKs derived from the natural rhomboid substrate TatA from bacterium Providencia stuartii bind GlpG in a substrate-like manner, and their co-crystal structures with GlpG reveal the S1 to S4 subsites of the protease. The S1 subsite is prominent and merges into the 'water retention site', suggesting intimate interplay between substrate binding, specificity and catalysis. Unexpectedly, the S4 subsite is plastically formed by residues of the L1 loop, an important but hitherto enigmatic feature of the rhomboid fold. We propose that the homologous region of members of the wider rhomboid-like protein superfamily may have similar substrate or client-protein binding function. Finally, using molecular dynamics, we generate a model of the Michaelis complex of the substrate bound in the active site of GlpG.

Synthesis of lucifensin by native chemical ligation and characteristics of its isomer having different disulfide bridge pattern.

The antimicrobial 40-amino-acid-peptide lucifensin was synthesized by native chemical ligation (NCL) using N-acylbenzimidazolinone (Nbz) as a linker group. NCL is a method in which a peptide bond between two discreet peptide chains is created. This method has been applied to the synthesis of long peptides and proteins when solid-phase synthesis is imcompatible. Two models of ligation were developed: [15+25] Ala-Cys and [19+21] His-Cys. The [19+21] His-Cys method gives lower yield because of the lower stability of 18-peptide-His-Nbz-CONH2 peptide, as suggested by density functional theory calculation. Acetamidomethyl-deprotection and subsequent oxidation of the ligated linear lucifensin gave a mixture of lucifensin isomers, which differed in the location of their disulfide bridges only. The dominant isomer showed unnatural pairing of cysteines [C1-6], [C3-5], and [C2-4], which limits its ability to form α-helical structure. The activity of isomeric lucifensin toward Bacillus subtilis, Staphylococcus aureus, and Micrococcus luteus was lower than that of the natural lucifensin. The desired product native lucifensin was prepared from this isomer using a one-pot reduction with dithiotreitol and subsequent air oxidation in slightly alkaline medium.

Multipass quantum process tomography.

We introduce a method to enhance the precision and accuracy of Quantum Process Tomography (QPT) by mitigating the errors caused by state preparation and measurement (SPAM), readout and shot noise. Instead of performing QPT solely on a single gate, we propose performing QPT on a sequence of multiple applications of the same gate. The method involves the measurement of the Pauli transfer matrix (PTM) by standard QPT of the multipass process, and then deduce the single-process PTM by two alternative approaches: an iterative approach which in theory delivers the exact result for small errors, and a linearized approach based on solving the Sylvester equation. We examine the efficiency of these two approaches through simulations on IBM Quantum using IBMQ_QASM_SIMULATOR. Compared to the Randomized Benchmarking type of methods, the proposed method delivers the entire PTM rather than a single number (fidelity). Compared to standard QPT, our method delivers PTM with much higher accuracy and precision because it greatly reduces the SPAM, readout and shot noise errors. We use the proposed method to experimentally determine the PTM and the fidelity of the CNOT gate on the quantum processor IBMQ_MANILA (Falcon r5.11L).

The vascular footprint in cardiac homeostasis and hypertensive heart disease-A link between apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase.

Recent studies have suggested a connection between disturbances of the apelin system and various cardiac pathologies, including hypertension, heart failure, and atherosclerosis. Vascular endothelial growth factor is crucial for cardiac homeostasis as a critical molecule in cardiac angiogenesis. Neuronal nitric oxide synthase is an essential enzyme producing nitric oxide, a key regulator of vascular tone. The present study aims to shed light upon the complex interactions between these three vital signaling molecules and examine their changes with the progression of hypertensive heart disease. We used two groups of spontaneously hypertensive rats and age-matched Wistar rats as controls. The expression of the apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase were assessed immunohistochemically. We used capillary density and cross-sectional area of the cardiomyocytes as quantitative parameters of cardiac hypertrophy. Immunoreactivity of the molecules was more potent in both ventricles of spontaneously hypertensive rats compared with age-matched controls. However, capillary density was lower in both ventricles of the two age groups of spontaneously hypertensive rats compared with controls, and the difference was statistically significant. In addition, the cross-sectional area of the cardiomyocytes was higher in both ventricles of the two age groups of spontaneously hypertensive rats compared with controls, and the difference was statistically significant. Our study suggests a potential link between the apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase in cardiac homeostasis and the hypertensive myocardium. Nevertheless, further research is required to better comprehend these interactions and their potential therapeutic implications.

Extensive targeting of chemical space at the prime side of ketoamide inhibitors of rhomboid proteases by branched substituents empowers their selectivity and potency.

Rhomboid intramembrane serine proteases have been implicated in several pathologies, and emerge as attractive pharmacological target candidates. The most potent and selective rhomboid inhibitors available to date are peptidyl α-ketoamides, but their selectivity for diverse rhomboid proteases and strategies to modulate it in relevant contexts are poorly understood. This gap, together with the lack of suitable in vitro models, hinders ketoamide development for relevant eukaryotic rhomboid enzymes. Here we explore the structure-activity relationship principles of rhomboid inhibiting ketoamides by medicinal chemistry and enzymatic in vitro and in-cell assays with recombinant rhomboid proteases GlpG, human mitochondrial rhomboid PARL and human RHBDL2. We use X-ray crystallography in lipidic cubic phase to understand the binding mode of one of the best ketoamide inhibitors synthesized here containing a branched terminal substituent bound to GlpG. In addition, to extend the interpretation of the co-crystal structure, we use quantum mechanical calculations and quantify the relative importance of interactions along the inhibitor molecule. These combined experimental analyses implicates that more extensive exploration of chemical space at the prime side is unexpectedly powerful for the selectivity of rhomboid inhibiting ketoamides. Together with variations in the peptide sequence at the non-prime side, or its non-peptidic alternatives, this strategy enables targeted tailoring of potent and selective ketoamides towards diverse rhomboid proteases including disease-relevant ones such as PARL and RHBDL2.

Chemical Blockage of the Mitochondrial Rhomboid Protease PARL by Novel Ketoamide Inhibitors Reveals Its Role in PINK1/Parkin-Dependent Mitophagy.

The mitochondrial rhomboid protease PARL regulates mitophagy by balancing intramembrane proteolysis of PINK1 and PGAM5. It has been implicated in the pathogenesis of Parkinson's disease, but its investigation as a possible therapeutic target is challenging in this context because genetic deficiency of PARL may result in compensatory mechanisms. To address this problem, we undertook a hitherto unavailable chemical biology strategy. We developed potent PARL-targeting ketoamide inhibitors and investigated the effects of acute PARL suppression on the processing status of PINK1 intermediates and on Parkin activation. This approach revealed that PARL inhibition leads to a robust activation of the PINK1/Parkin pathway without major secondary effects on mitochondrial properties, which demonstrates that the pharmacological blockage of PARL to boost PINK1/Parkin-dependent mitophagy is a feasible approach to examine novel therapeutic strategies for Parkinson's disease. More generally, this study showcases the power of ketoamide inhibitors for cell biological studies of rhomboid proteases.

ß-Cyclodextrin duplexes that are connected through two disulfide bonds: potent hosts for the complexation of organic molecules.

New tubular host molecules, which are composed of two ß-cyclodextrin macrocycles that are connected through two disulfide bonds, have been prepared by the air-promoted oxidation of 6(I),6(IV)-dideoxy-6(I),6(IV)-disulfanyl-ß-cyclodextrin in aqueous solution. This reaction leads to three products: monomeric intramolecular disulfide and two dimeric species, which are termed as "non-eclipsed" and "eclipsed" cyclodextrin duplexes. Oxidation at a concentration of the starting thiol of 0.1 mM gave the intramolecular disulfide as the major product whereas a concentration in the millimolar range afforded the dimeric species as the dominant products. The tubular structure of the "non-eclipsed" isomer was unequivocally determined by X-ray analysis. The binding affinities of the duplexes to a wide range of compounds, including fluorescent dyes and clinically used drugs Imatinib and Esomeprazol, were studied in water by ITC. For most guest compounds, the experimentally determined K(a) values were in the range 10(7)-10(8) M(-1). These binding affinities are significantly higher than those found in the literature for analogous complexes with native cyclodextrins. In cases of binding of neutral or anionic guest molecules cyclodextrin duplexes outperformed cucurbiturils. A complex between a duplex and Nile blue was used to investigate its ability to penetrate the cytoplasmic membrane of HeLa cells. We found that the complex accumulated in the cell membrane but did not pass into cytosol. Importantly, the complex did not decompose to a significant extent under high dilution in the cellular environment.

Mast cells and basic fibroblast growth factor in physiological aging of rat heart and kidney.

Age-related morphological and physiological changes occur in cells, tissues and organs with high metabolic or mitotic activity; these changes decrease their regenerative capacity. One such change is interstitial fibrosis. Mast cells contain basic fibroblast growth factor and have been related to pro-fibrotic activity. We investigated the role of mast cells in physiological aging of the heart and kidney. We analyzed changes in mast cell number and compared the left and right heart ventricles and kidneys of 6- and 12-month-old Wistar rats. We also evaluated the immunohistochemical expression of basic fibroblast growth factor. Finally, we analyzed changes in the extent of interstitial fibrosis and in the glomerular sclerosis index as nonspecific markers of aging and correlated these parameters with of mast cells. Mast cells were visualized by toluidine blue staining and specific immunohistochemical expression of tryptase. The expression of basic fibroblast growth factor was assessed semiquantitatively. The extent of interstitial fibrosis was investigated using Mallory's trichrome staining. Glomerular sclerosis was evaluated using periodic acid-Schiff staining. We found that the number of mast cells increased significantly in the older rats. We also found that the number of mast cells was greatest in the left ventricle followed by the right ventricle, then the kidney. The immunoreactivity of basic fibroblast growth factor also increased in older animals. Correlations between the number of mast cells and immunoreactivity of basic fibroblast growth factor, extent of interstitial fibrosis and glomerular sclerosis index demonstrated the association between mast cells and age-related tissue remodeling of the heart and kidney.

Changes in the number of mast cells, expression of fibroblast growth factor-2 and extent of interstitial fibrosis in established and advanced hypertensive heart disease.

INTRODUCTION: An increasing number of studies have shed light on the role of cardiac mast cells in the pathogenesis of hypertension-induced myocardial remodeling. Mast cells promote fibroblast activation, myofibroblast differentiation and subsequent collagen accumulation through the action of tryptase, chymase, histamine and fibroblast growth factor-2. The aim of the present study was to report on the changes in the number of mast cells as evaluated through toluidine blue, tryptase and c-kit staining, to assess the extent of interstitial fibrosis and correlate it with the changes in the number of mast cells and to analyze the immunohistochemical expression of fibroblast growth factor-2 in two groups of spontaneously hypertensive rats indicative of established and advanced hypertensive heart disease. A novel aspect of our work was the analysis of all parameters in the right ventricle. MATERIAL AND METHODS: For the present study, we used 6- and 12-month-old spontaneously hypertensive rats. A light microscopic study was conducted on sections stained with hematoxylin and eosin and toluidine blue. For the immunohistochemical study we used monoclonal antibodies against mast cell tryptase and fibroblast growth factor-2 and a polyclonal antibody against c-kit. The expression of fibroblast growth factor-2 was assessed semi-quantitatively through ImageJ. The number of mast cells was evaluated on toluidine blue-, tryptase- and c-kit-stained sections and a comparative statistical analysis with the Mann-Whitney test was conducted between the two age groups. A separate statistical analysis between results obtained through immunostaining for tryptase and for c-kit was conducted in each age group with the Wilcoxon signed-rank test. The extent of fibrosis was assessed quantitatively on slides stained with Mallory's trichrome stain as a percentage of the whole tissue and compared between the two age groups. Spearman's correlation was used to test whether a correlation exists between the number of mast cells and the percentage of interstitial fibrosis. RESULTS: Mast cells with typical cytoplasmic granules were visualized in the interstitial tissue and in the perivascular zone in both age groups. In both ventricles, their number increased significantly in 12-month-old animals as evaluated through all three staining methods. Moreover, immunostaining for tryptase and for c-kit yielded comparable results. The immunoreactivity of fibroblast growth factor-2 increased in both ventricles in older animals. Expression of this protein was particularly intensive in the cytoplasm of connective tissue cells with the characteristic features of mast cells mainly found in the areas of fibrotic alterations in 12-month-old spontaneously hypertensive rats. In both ventricles, interstitial fibrosis was more extensive throughout the myocardium of older animals and was positively correlated with the changes in the number of mast cells in both age groups. CONCLUSION: The present study reported for the first time that the increase in the number of mast cells, observed as hypertension-induced myocardial changes progress, is statistically significant and confirmed that this process takes place in both ventricles. This increase is accompanied by a higher expression of fibroblast growth factor-2 and is more strongly correlated with the more pronounced interstitial fibrosis in older animals, further supporting the role of mast cells in the structural changes taking place in the myocardium in response to systemic hypertension.

The potential role of mast cells and fibroblast growth factor-2 in the development of hypertension-induced renal damage.

Hypertension-induced renal injury is a multifactorial process which plays a crucial role in the development of chronic kidney disease. Multiple studies have demonstrated that interstitial rather than glomerular changes correlate better with renal functional capacity. Recent evidence indicates that mast cells and cell signaling proteins such as fibroblast growth factor-2 may contribute to the progression of interstitial changes under hypertensive conditions. The aim of our study was to determine the localization of mast cells in the renal cortex and report on the changes in their number, to analyze the distribution of fibroblast growth factor-2, to assess the extent of renal fibrosis and to evaluate renal damage and correlate it with the changes in the number of mast cells in a model of hypertension-induced renal injury by comparing two age groups of spontaneously hypertensive rats. We used 6- and 12-month-old animals. A light microscopic study was conducted on sections stained with hematoxylin and eosin, periodic acid-Schiff stain, Mallory's trichrome method and toluidine blue. For the immunohistochemical study we used monoclonal antibodies against mast cell tryptase and fibroblast growth factor-2 and a polyclonal antibody against c-kit. The expression of fibroblast growth factor-2 was assessed semi-quantitatively. The number of mast cells was evaluated on toluidine blue-, tryptase- and c-kit-stained sections, as well as double-stained sections and a comparative statistical analysis with the Mann-Whitney test was conducted between the two age groups. Our results showed that mast cells were located mainly in the peritubular and perivascular areas and were absent in the region of the renal corpuscles. Their number increased significantly in 12-month-old animals. Immunostaining for tryptase, c-kit and double staining for both molecules yielded identical results. The immunohistochemical expression of fibroblast growth factor-2 increased in the kidneys of older animals, as did the percentage of collagen fibers. In addition, we described more severe renal damage in 12-month-old spontaneously hypertensive rats and noted a positive correlation in both age groups between the number of mast cells on the one hand and glomerular sclerosis index and tubulointerstitial damage index, on the other. The results obtained in the present study support the pivotal role of mast cells in the development of hypertension-induced kidney damage.

Designed Parasite-Selective Rhomboid Inhibitors Block Invasion and Clear Blood-Stage Malaria.

Rhomboid intramembrane proteases regulate pathophysiological processes, but their targeting in a disease context has never been achieved. We decoded the atypical substrate specificity of malaria rhomboid PfROM4, but found, unexpectedly, that it results from "steric exclusion": PfROM4 and canonical rhomboid proteases cannot cleave each other's substrates due to reciprocal juxtamembrane steric clashes. Instead, we engineered an optimal sequence that enhanced proteolysis >10-fold, and solved high-resolution structures to discover that boronates enhance inhibition >100-fold. A peptide boronate modeled on our "super-substrate" carrying one "steric-excluding" residue inhibited PfROM4 but not human rhomboid proteolysis. We further screened a library to discover an orthogonal alpha-ketoamide that potently inhibited PfROM4 but not human rhomboid proteolysis. Despite the membrane-immersed target and rapid invasion, ultrastructural analysis revealed that single-dosing blood-stage malaria cultures blocked host-cell invasion and cleared parasitemia. These observations establish a strategy for designing parasite-selective rhomboid inhibitors and expose a druggable dependence on rhomboid proteolysis in non-motile parasites.

Ultrastructure of the dorsal claustrum in cat. II. Synaptic organization.

The claustrum is a bilateral subcortical nucleus situated between the insular cortex and the striatum in the brain of all mammals. It consists of two embryologically distinct subdivisions - dorsal and ventral claustrum. The claustrum has high connectivity with various areas of the cortex, subcortical and allocortical structures. It has long been suggested that the various claustral connections have different types of synaptic contacts at the claustral neurons. However, to the best of our knowledge, the literature data on the ultrastructural organization of the different types of synaptic contacts in the dorsal claustrum are very few. Therefore, the aim of our study was to observe and describe the synaptic organization of the dorsal claustrum in the cat. We used a total of 10 adult male cats and conducted an ultrastructural study under a transmission electron microscope as per established protocol. We described a multitude of dendritic spines, which were subdivided into two types - with and without foot processes. Based on the size and shape of the terminal boutons, the quantity and distribution of vesicles and the characteristic features of the active synaptic zone, we described six types of synaptic boutons, most of which formed asymmetrical synaptic contacts. Furthermore, we reported the presence of axo-dendritic, axo-somatic, dendro-dendritic and axo-axonal synapses. The former two likely represent the morphological substrate of the corticoclaustral pathway, while the remaining two types have the ultrastructural features of inhibitory synapses, likely forming a local inhibitory circuit in the claustrum. In conclusion, the present study shares new information about the neuropil of the claustrum and proposes a systematic classification of the types of synaptic boutons and contacts observed in the dorsal claustrum of the cat, thus supporting its key and complex role as a structure integrating various information within the brain.

Cytoarchitecture of the dorsal claustrum of the cat: a quantitative Golgi study.

The claustrum is a subcortical nucleus, found in the telencephalon of all placental mammals. Earlier Golgi studies have mostly focused on a qualitative description of the types of neurons. The aim of the present study was to describe the types of neurons found in the dorsal claustrum of the cat using the Golgi impregnation method and to perform a quantitative analysis of the following morphometric parameters: number of terminals (ends), total dendritic length, dendritic complexity, spine density (in spiny projection neurons), varicosity density (in aspiny interneurons). We used specimens from 5 healthy male cats stained according to the Golgi-Cox method. The dendritic trees of the studied neurons were then reconstructed through the Neurolucida software. Values of the studied quantitative parameters were obtained automatically and tested for statistically significant differences. Five types of spiny neurons were observed-large, medium-sized and small multipolar, bipolar and pyramidal-like. In addition, we described three types of aspiny neurons. The quantitative values and the statistical analysis were presented with tables and diagrams. In conclusion, we have presented a detailed analysis of the cytoarchitecture of the DC of the cat and have reported the first quantitative data on a number of morphometric parameters.

Synthesis, computational study and cytotoxic activity of new 4-hydroxycoumarin derivatives.

Six new 4-hydroxycoumarin derivatives have been synthesized. They were characterized by UV-vis, IR, (1)H NMR, (13)C NMR, mass spectral data, elemental analysis, TLC and melting point determination. The new 4-hydroxycoumarin derivatives are studied by computational methods--DFT (B3LYP) and force field methods (MM2 and OPLS), in order to optimize their geometry and calculate quantum-chemical properties and conformational analysis. Five new 4-hydroxycoumarin derivatives were tested for cytotoxic activity in two tumor cell lines--HL-60 and EJ. The obtained results are compared with the utilized anticancer drug melphalan. Two of these compounds--ethyl 2-[(3,4-dihydroxyphenyl)(4-hydroxy-2-oxo-2H-chromen-3-yl)methyl]-3-oxobutanoate (SS-16) and ethyl 2-[(4-hydroxy-2-oxo-2H-chromen-3-yl)(3-nitrophenyl)methyl]-3-oxobutanoate (SS-21) show comparatively good cytotoxic properties. Their activity is weaker than melphalan. SS-16 seems to be more active than SS-21.

Microcirculatory changes as a hallmark of aging in the heart and kidney / Cambios microcirculatorios como sello distintivo del envejecimiento en el corazón y riñón

SUMMARY: Changes in the microcirculation of multiple tissues and organs have been implicated as a possible mechanism in physiological aging. In particular, vascular endothelial growth factor is a secretory protein responsible for regulating angiogenesis via altering endothelial proliferation, survival, migration, extracellular matrix degradation and cell permeability. The aim of the present study was to evaluate the role of vascular endothelial growth factor in the progression of morphological alterations caused by physiological aging in the heart and kidney and to examine its relation to changes in capillary density. We used two age groups of healthy Wistar rats - 6- and 12-month- old. The expression of vascular endothelial growth factor was examined through immunohistochemistry and immunofluorescence and assessed semi-quantitatively. Changes in capillary density were evaluated statistically and correlated with the expression of vascular endothelial growth factor. We reported stronger immunoreactivity for vascular endothelial growth factor in the left compared to the right ventricle and also observed an increase in its expression in both ventricles in older animals. Contrasting results were reported for the renal cortex and medulla. Capillary density decreased statistically in all examined structures as aging progressed. The studied correlations were statistically significant in the two ventricles in 12-month-old animals and in the renal cortex of both age groups. Our results shed light on some changes in the microcirculation that take place as aging advances and likely contribute to impairment in the function of the examined organs.

Los cambios en la microcirculación de múltiples tejidos y órganos se han implicado como un posible mecanismo en el envejecimiento fisiológico. En particular, el factor de crecimiento endotelial vascular es una proteína secretora responsable de regular la angiogénesis mediante la alteración de la proliferación endotelial, la supervivencia, la migración, la degradación de la matriz extracelular y la permeabilidad celular. El objetivo del presente estudio fue evaluar el papel del factor de crecimiento del endotelio vascular en la progresión de las alteraciones morfológicas causadas por el envejecimiento fisiológico en el corazón y riñón y examinar su relación con los cambios en la densidad capilar. Utilizamos dos grupos de ratas Wistar sanas: 6 y 12 meses de edad. La expresión del factor de crecimiento del endotelio vascular se examinó mediante inmunohistoquímica e inmunofluorescencia y se evaluó semicuantitativamente. Los cambios en la densidad capilar se evaluaron estadísticamente y se correlacionaron con la expresión del factor de crecimiento del endotelio vascular. Informamos una inmunorreactividad más fuerte para el factor de crecimiento endotelial vascular en el ventrículo izquierdo en comparación con el derecho y también observamos un aumento en su expresión en ambos ventrículos en animales mayores. Se informaron resultados contrastantes para la corteza renal y la médula. La densidad capilar disminuyó estadísticamente en todas las estructuras examinadas a medida que avanzaba el envejecimiento. Las correlaciones estudiadas fueron estadísticamente significativas en los dos ventrículos en animales de 12 meses y en la corteza renal de ambos grupos de edad. Nuestros resultados arrojan luz sobre algunos cambios en la microcirculación que tienen lugar a medida que avanza el envejecimiento y probablemente contribuyan a un deterioro en la función de los órganos examinados.

General and Modular Strategy for Designing Potent, Selective, and Pharmacologically Compliant Inhibitors of Rhomboid Proteases.

Rhomboid-family intramembrane proteases regulate important biological processes and have been associated with malaria, cancer, and Parkinson's disease. However, due to the lack of potent, selective, and pharmacologically compliant inhibitors, the wide therapeutic potential of rhomboids is currently untapped. Here, we bridge this gap by discovering that peptidyl α-ketoamides substituted at the ketoamide nitrogen by hydrophobic groups are potent rhomboid inhibitors active in the nanomolar range, surpassing the currently used rhomboid inhibitors by up to three orders of magnitude. Such peptidyl ketoamides show selectivity for rhomboids, leaving most human serine hydrolases unaffected. Crystal structures show that these compounds bind the active site of rhomboid covalently and in a substrate-like manner, and kinetic analysis reveals their reversible, slow-binding, non-competitive mechanism. Since ketoamides are clinically used pharmacophores, our findings uncover a straightforward modular way for the design of specific inhibitors of rhomboid proteases, which can be widely applicable in cell biology and drug discovery.

Synthesis and Inhibiting Activity of Some 4-Hydroxycoumarin Derivatives on HIV-1 Protease.

Six novel 4-hydroxycoumarin derivatives were rationally synthesized, verified, and characterized by molecular docking using crystal HIV-1 protease. Molecular docking studies predicted antiprotease activity of (7) and (10). The most significant functional groups, responsible for the interaction with HIV-1 protease by hydrogen bonds formation are pyran oxygen, atom, lactone carbonyl oxygen and one of the hydroxyl groups. The newly synthesized compounds were biologically tested in MT-4 cells for inhibiting HIV-1 replication, exploring the protection of cells from the cytopathic effect of HIV measured by cell survival in MTT test. One derivative -7 showed 76-78% inhibition of virus infectivity with IC(50) = 0.01 nM, much less than the maximal nontoxic concentration (1 mM). Antiprotease activity of 7 in two different concentrations was detected to be 25%. Nevertheless, the results of study of (7) encourage using it as a pharmacophore for further synthesis and evaluation of anti-HIV activity.