La caída casual en Asturias, una causa de lesión medular traumática en ascenso / Accidental falls in Asturias, an increasing cause of traumatic spinal cord injury

Introducción. La caída casual es una causa cada vez más frecuente de lesión medular traumática en nuestro medio. Objetivos. Conocer la incidencia y las características sociodemográficas de los pacientes con lesión medular traumática de etiología casual en Asturias. Pacientes y métodos. Censo de pacientes codificados como «lesión medular traumática con o sin fractura vertebral» de etiología casual en Asturias entre 1932 y 2013. Se recogen variables sociodemográficas, hospitalarias y clínicas. Resultados. Se registraron un total de 317 pacientes, la mayoría varones (68%), con una edad media de 56 años ± 18,8 DT. La incidencia en 2012 fue de 8,4 por millón de habitantes, con tendencia al aumento, siendo actualmente la primera causa de lesión medular traumática. La muestra estudiada presentó una distribución homogénea en cuanto al tipo de lesión (completas el 51,6% e incompletas el 48,4%), con un predominio de localización cervical (32,2%). Las caídas de propia altura representaron el 32%, y se relacionaron con mayor edad media y presencia de lesión incompleta. En la población analizada el 76,8% presentaban lesión ósea, existiendo en más de la mitad de ellos una afectación multinivel. Conclusiones. Se observa una tendencia hacia el aumento en la proporción de lesiones medulares traumáticas provocadas por caída casual, relacionadas con el envejecimiento poblacional, que pone en evidencia la necesidad de crear nuevas estrategias tanto preventivas como terapéuticas (AU)

Introduction. Accidental falls are an increasing cause of traumatic spinal cord injury in Spain. Aims. To assess the incidence and socio-demographic characteristics of traumatic spinal cord injury due to accidental falls in Asturias. Patients and methods. A census was made of patients codified as «traumatic spinal cord injury with or without vertebral fracture» due to falls in Asturias between 1932 and 2013. Socio-demographic, hospital and clinical variables were gathered. Results. In total 317 patients were registered, most of them male (68%), with an average age of 56 years ± 18.8 DT. The estimated incidence in 2012 was 8.4 cases/million/year, with an increasing trend. Accidental falls are currently the leading cause of traumatic spinal cord injury. The distribution in the type of injury was homogeneous (complete in 51.6% and incomplete in 48.4%) and the most common location was cervical (32.2%). Own height falls represented 32% and were associated with a higher mean age and the presence of incomplete injury. Bone injury was present in 76.8%, with multilevel injury in more than half of the patients. Conclusions. An increasing trend was observed in the proportion of traumatic spinal cord injuries due to accidental falls, related to the aging population. There is a need to create specific preventive and therapeutic strategies (AU)

Identification and functional characterization of AclB, a novel cell-separating enzyme from Lactobacillus casei.

Autolysis of nonstarter lactic acid bacteria (NSLAB) was favorable for the development of flavor compounds during cheese manufacture. Among these bacteria, Lb. casei was regarded as the most important microbiota involved in cheese processes. In this study, a novel autolysin named AclB was identified in the genome of Lb. casei BL23 and its modular structure was predicted through bioinformatic approaches. Subsequently, its transcription profile in the exponential phase, hydrolytic activities against cell walls, enzymatic properties under different conditions, physiological function via gene inactivation and upregulation assays, as well as potential applications to NSLAB's autolysis were fully investigated. According to the results, AclB was recognized as a species-specific cell-separating enzyme, responsible for cell separation after cell division in Lb. casei BL23. The purified AclB showed considerable hydrolyzing activities towards cell walls, indicating its enzymatic nature as peptidoglycan hydrolase, or autolysin. The highest activity of AclB was determined at pH5.0 and 37°C, and the expression vector constructed based on AclB was shown to facilitate the controlled lysis of Lb. casei BL23 hosts. In summary, this study provided insight into the enzymatic properties of a novel autolysin involved in cell separation of Lb. casei BL23, which is promising to accelerate cheese ripening and improve cheese quality.

Postmortem radiography of gastromalacia: case reports.

Gastromalacia is a postmortem artifact resulting from autolysis of the gastric walls. Gastromalacia is autolytic rupture of the stomach caused by endogenous enzymes, and it is devoid of any vital reactions. The left leaf of the diaphragm is occasionally perforated by a ragged fenestration, with escape of gastric contents into the pleural cavity. This rupture may lead to pneumoperitoneum or pneumothorax. For diagnostic radiologists, gastromalacia is rarely encountered. Therefore, they should be aware of this entity to avoid misdiagnosis when performing postmortem radiography.

Time course in calpain activity and autolysis in slow and fast skeletal muscle during clenbuterol treatment.

Calpains are Ca2+ cysteine proteases that have been proposed to be involved in the cytoskeletal remodeling and wasting of skeletal muscle. Cumulative evidence also suggests that ß2-agonists can lead to skeletal muscle hypertrophy through a mechanism probably related to calcium-dependent proteolytic enzyme. The aim of our study was to monitor calpain activity as a function of clenbuterol treatment in both slow and fast phenotype rat muscles. For this purpose, for 21 days we followed the time course of the calpain activity and of the ubiquitous calpain 1 and 2 autolysis, as well as muscle remodeling in the extensor digitorum longus (EDL) and soleus muscles of male Wistar rats treated daily with clenbuterol (4 mg·kg-1). A slow to fast fiber shift was observed in both the EDL and soleus muscles after 9 days of treatment, while hypertrophy was observed only in EDL after 9 days of treatment. Soleus muscle but not EDL muscle underwent an early apoptonecrosis phase characterized by hematoxylin and eosin staining. Total calpain activity was increased in both the EDL and soleus muscles of rats treated with clenbuterol. Moreover, calpain 1 autolysis increased significantly after 14 days in the EDL, but not in the soleus. Calpain 2 autolysis increased significantly in both muscles 6 hours after the first clenbuterol injection, indicating that clenbuterol-induced calpain 2 autolysis occurred earlier than calpain 1 autolysis. Together, these data suggest a preferential involvement of calpain 2 autolysis compared with calpain 1 autolysis in the mechanisms underlying the clenbuterol-induced skeletal muscle remodeling.

Acute pancreatitis.

Acute pancreatitis (AP) is an important cause of morbidity and mortality worldwide and the annual incidence appears to be increasing. It presents as a mild self-limiting illness in 80% of patients. However, one-fifth of these develop a severe complicated life-threatening disease requiring intensive and prolonged therapeutic intervention. Alcohol and gallstone disease remain the commonest causes of AP but metabolic abnormalities, obesity and genetic susceptibility are thought be increasingly important aetiological factors. The prompt diagnosis of AP and stratification of disease severity is essential in directing rapid delivery of appropriate therapeutic measures. In this review, the range of diagnostic and prognostic assays, severity scoring systems and radiological investigations used in current clinical practice are described, highlighting their strengths and weaknesses. Increased understanding of the complex pathophysiology of AP has generated an array of new potential diagnostic assays and these are discussed. The multidisciplinary approach to management of severe pancreatitis is outlined, including areas of controversy and novel treatments.

The extracellular ß-1,3-endoglucanase EngA is involved in autolysis of Aspergillus nidulans.

AIMS: To elucidate the roles of the ß-1,3-endoglucanase EngA in autolysis of the filamentous fungus Aspergillus nidulans and to identify the common regulatory elements of autolytic hydrolases. METHODS AND RESULTS: A ß-1,3-endoglucanase was purified from carbon-starving cultures of A. nidulans. This enzyme is found to be encoded by the engA gene (locus ID: AN0472.3). Functional and gene-expression studies demonstrated that EngA is involved in the autolytic cell wall degradation resulting from carbon starvation of the fungus. Moreover, regulation of engA is found to be dependent on the FluG/BrlA asexual sporulation signalling pathway in submerged culture. The deletion of either engA or chiB (encoding an endochitinase) caused highly reduced production of hydrolases in general. CONCLUSIONS: The ß-1,3-endoglucanase EngA plays a pivotal role in fungal autolysis, and activities of both EngA and ChiB are necessary to orchestrate the expression of autolytic hydrolases. The production of cell wall-degrading enzymes was coordinately controlled in a highly sophisticated and complex manner. SIGNIFICANCE AND IMPACT OF THE STUDY: No information was available on the autolytic glucanase(s) of the euascomycete A. nidulans. This study demonstrates that EngA is a key element in fungal autolysis, and normal activities of both EngA and ChiB are crucial for balanced production of hydrolases.

The role of autolysis loop in determining the specificity of coagulation proteases.

We recently demonstrated that the substitution of the autolysis loop (residues 143 to 154 in the chymotrypsin numbering system) of activated protein C (APC) with the corresponding loop of factor Xa (fXa) renders the APC mutant (APC/fX143-154) susceptible to inhibition by antithrombin (AT) in the presence of pentasaccharide. Our recent results further indicated, that in addition to an improvement in the reactivity of APC/fX143-154 with AT, both the amidolytic and anti-factor Va activities of the mutant APC have also been significantly increased. Since the autolysis loop of APC is five residues longer than the autolysis loop of fXa, it could not be ascertained whether this loop in the mutant APC specifically interacts with the activated conformation of AT or if a shorter autolysis loop is responsible for a global improvement in the catalytic activity of the mutant protease. To answer this question, we prepared another APC mutant in which the autolysis loop of the protease was replaced with the corresponding loop of trypsin (APC/Tryp143-154). Unlike an approximately 500-fold improvement in the reactivity of APC/fX143-154 with AT in the presence of pentasaccharide, the reactivity of APC/Tryp143-154 with the serpin was improved approximately 10-fold. These results suggest that both the length and structure of residues of the autolysis loop are critical for the specificity of the coagulation protease interaction with AT. Further factor Va inactivation studies with the APC mutants revealed a similar role for the autolysis loop of APC in the interaction with its natural substrate.

The role of autolysis loop in determining the specificity of coagulation proteases

We recently demonstrated that the substitution of the autolysis loop (residues 143 to 154 in the chymotrypsin numbering system) of activated protein C (APC) with the corresponding loop of factor Xa (fXa) renders the APC mutant (APC/fX143-154) susceptible to inhibition by antithrombin (AT) in the presence of pentasaccharide. Our recent results further indicated, that in addition to an improvement in the reactivity of APC/fX143-154 with AT, both the amidolytic and anti-factor Va activities of the mutant APC have also been significantly increased. Since the autolysis loop of APC is five residues longer than the autolysis loop of fXa, it could not be ascertained whether this loop in the mutant APC specifically interacts with the activated conformation of AT or if a shorter autolysis loop is responsible for a global improvement in the catalytic activity of the mutant protease. To answer this question, we prepared another APC mutant in which the autolysis loop of the protease was replaced with the corresponding loop of trypsin (APC/Tryp143-154). Unlike an ~500-fold improvement in the reactivity of APC/fX143-154 with AT in the presence of pentasaccharide, the reactivity of APC/Tryp143-154 with the serpin was improved ~10-fold. These results suggest that both the length and structure of residues of the autolysis loop are critical for the specificity of the coagulation protease interaction with AT. Further factor Va inactivation studies with the APC mutants revealed a similar role for the autolysis loop of APC in the interaction with its natural substrate.

Autolysosomal membrane-associated betaine homocysteine methyltransferase. Limited degradation fragment of a sequestered cytosolic enzyme monitoring autophagy.

We compared the membrane proteins of autolysosomes isolated from leupeptin-administered rat liver with those of lysosomes. In addition to many polypeptides common to the two membranes, the autolysosomal membranes were found to be more enriched in endoplasmic reticulum lumenal proteins (protein-disulfide isomerase, calreticulin, ER60, BiP) and endosome/Golgi markers (cation-independent mannose 6-phosphate receptor, transferrin receptor, Golgi 58-kDa protein) than lysosomal membranes. The autolysosomal membrane proteins include three polypeptides (44, 35, and 32 kDa) whose amino-terminal sequences have not yet been reported. Combining immunoblotting and reverse transcriptase-polymerase chain reaction analyses, we identified the 44-kDa peptide as the intact subunit of betaine homocysteine methyltransferase and the 35- and 32-kDa peptides as two proteolytic fragments. Pronase digestion of autolysosomes revealed that the 44-kDa and 32-kDa peptides are present in the lumen, whereas the 35-kDa peptide is not. In primary hepatocyte cultures, the starvation-induced accumulation of the 32-kDa peptide occurs in the presence of E64d, showing that the 32-kDa peptide is formed from the sequestered 44-kDa peptide during autophagy. The accumulation is induced by rapamycin but completely inhibited by wortmannin, 3-methyladenine, and bafilomycin. Thus, detection of the 32-kDa peptide by immunoblotting can be used as a streamlined assay for monitoring autophagy.

Purification and properties of a chitinase from Penicillium oxalicum autolysates.

A chitinase (EC. 3.2.1.14) from autolysed culture filtrate of Penicillium oxalicum was purified by precipitation with ammonium sulphate, gel filtration and ion exchange chromatographies. The purified enzyme showed a single protein band in SDS gel electrophoresis. The enzyme is an acidic protein with a pI of 4.5 and has a molecular weight of 54,900 as estimated from SDS gel electrophoresis and 21,500 from gel filtration. The optimum pH and temperature were 5.0 and 35 degrees C, respectively. The enzyme was stable at temperatures up to 45 degrees C and in a pH range between 4.0 and 6.0. The Km was 2.5 mg ml-1 for colloidal chitin, Hg2+ and Ag+ were effective inhibitors. The viscosimetric study carried out using carboxymethyl chitin as substrate revealed the endotype action of this enzyme.

Autolysis and extension of isolated walls from growing cucumber hypocotyls.

Walls isolated from cucumber hypocotyls retain autolytic activities and the ability to extend when placed under the appropriate conditions. To test whether autolysis and extension are related, we treated the walls in various ways to enhance or inhibit long-term wall extension ('creep') and measured autolysis as release of various saccharides from the wall. Except for some non-specific inhibitors of enzymatic activity, we found no correlation between wall extension and wall autolysis. Most notably, autolysis and extension differed strongly in their pH dependence. We also found that exogenous cellulases and pectinases enhanced extension in native walls, but when applied to walls previously inactivated with heat or protease these enzymes caused breakage without sustained extension. In contrast, pretreatment of walls with pectinase or cellulase, followed by boiling in methanol to inactivate the enzymes, resulted in walls with much stronger expansin-mediated extension responses. Crude protein preparations from the digestive tracts of snails enhanced extension of both native and inactivated walls, and these preparations contained expansin-like proteins (assessed by Western blotting). Our results indicate that the extension of isolated cucumber walls does not depend directly on the activity of endogenous wall-bound autolytic enzymes. The results with exogenous enzymes suggest that the hydrolysis of matrix polysaccharides may not induce wall creep by itself, but may act synergistically with expansins to enhance wall extension.

The mutL repair gene of Escherichia coli K-12 forms a superoperon with a gene encoding a new cell-wall amidase.

We report a molecular genetic analysis of the region immediately upstream from the Escherichia coli mutL DNA repair gene at 94.8 min. An open reading frame ending 9 bp upstream from the start of mutL corresponds to a 48 kDa polypeptide detected previously in minicells. The predicted amino acid sequence of this 48 kDa polypeptide shows homology to the major N-acetylmuramoyl-L-alanine amidase autolysin of Bacillus subtilis, a known amidase of Bacillus licheniformis, and the product of a Salmonella typhimurium gene that maps near 50 min. Insertions in this upstream gene, which we named amiB, or in mutL did not affect cell shape or viability; however, overexpression of the AmiB polypeptide caused cell lysis, hypersensitivity to osmotic shock and treatment with water, and temporary autolysis by low levels of antibiotics, which are all consistent with AmiB acting as a cell-wall hydrolase. Analysis of chromosomal transcription demonstrated that amiB forms a complex operon with mutL and two additional upstream genes. mutL transcripts also originated from an internal promoter, designated PmutL, located in amiB 312 bp upstream from the translational start of mutL. Together, these results suggest that E. coli contains a second amidase possibly involved in cell-wall hydrolysis, septation, or recycling, and that transcription of this amidase is directly linked to a gene central for DNA repair.

[The role of induced autolysis during digestion in fishes]. / Rol' indutsirovannogo autoliza v protsesse pishchevareniia u ryb.

A considerable activity of digestive hydrolases (carbohydrases in particular) was found in fish during feeding. The enzyme activity increases 2-6-fold at low values of the pH due, probably, to autolysis. This mechanism is important for the digestion in fish and other wild animals.

Biochemical, morphological and cytochemical studies of enhanced autolysis of Saccharomyces cerevisiae. 2. Morphological and cytochemical studies.

Morphological and cytochemical observation of Saccharomyces cerevisiae undergoing of induced autolysis were done in response to various chemical inducers of autolysis (NaCl, ethanol, fresh autolyzate). Changes in the inner structure of yeast cells were monitored by transmission electron microscopy and the surface of the cell wall was observed by scanning electron microscopy during autolysis. Cytochemical characterization of autolyzed cells was performed using four synthetic substrates for determination of proteinase activities but only carboxypeptidase Y could be detected in the vacuolar membranes. The morphological studies supported the data obtained from biochemical studies and confirmed that optimized conditions of autolysis have a significant effect on the structural changes of autolyzed yeast.

Coagulation autolysis in microorganisms and its relation to coagulase production.

The phenomenon of coagulation autolysis was observed in two model microorganisms, i.e., a bacterial culture and an imperfect fungus. It was characterized by impairment of the cell membranes, followed by condensation and dehydration of the cytoplasm and long-term preservation of the cells in the form of coagulated cytoplasm. In this respect, it was similar to coagulation necrosis of human tissues. The autolysis in the microorganisms was accompanied by increase of their coagulase activity, the substrate specificity of the enzyme rather broad. The coagulase activity of the microorganisms was detected during the culture period between the lag-phase and the exponential growth phase, i.e., the phase of their active growth. It served as a signal to induce biosynthesis of peptidohydrolase and cleavage of proteins. We believe that the phenomenon of coagulation autolysis in these microorganisms is rather typical and can be considered as an adaptative reaction, inducing a cascade of events from synthesis of coagulase to overproduction of peptidohydrolases with proteolytic activity.

[The relation between corneal autolysis and the lysosomal enzyme level in aqueous humor and corneal storage medium].

The activity of acid phosphatase, a marker enzyme of lysosomal enzymes, in aqueous humor of wet-chamber stored eyeball and corneal storage medium was determined. The results showed a significant rise in the enzyme activity as the storage time increased, i.e. the degree of autolysis of the cornea progressed.

[The morphogenesis of pancreonecrosis]. / Morfogenez pankreonekroza.

Pancreas was studied histologically, histochemically and electron microscopically in 117 autopsy cases and in the experiment (25 dogs and 60 white rats). Morphogenesis of pancreonecrosis proceeds through certain stages. The initial stages of an acute pancreatic edema is characterized by degenerative and necrotic changes of pancreocytes with parapedesis of pancreatic enzymes into the interstitium and a reactive stromal edema. The following stages are distinguished in the progressing phase of pancreonecrosis: hemorrhagic pancreonecrosis when the proteolytic enzymes provoke a colliquation necrosis of the acinar tissue, fibrinoid necrosis of vascular walls and disturbances of the intravascular hemorheology resulting in the enhancement of destructive processes and hemorrhagic inhibition of tissues; fat pancreonecrosis in which lipolytic enzymes lead to the coagulation necrosis of the acinar and fat tissue while a non-completed proteolysis of necrotic tissues stipulates the intensity of the reactive inflammation.

[The effect of autolysis on the conduction system in young rats. An enzyme histochemistry study]. / Vliv autolýzy na prevodní systém mladých krys. Enzym-histochemická studie.

The conducting system of young rats was studied by histochemistry during autolysis at room temperature lasting 1, 3, 6, 24 and 48 hours. The results were compared with working muscle tissue. The most sensitive was the "glycogen-dependent" phosphorylase and the activity decreased after 1 hour. It was slightly detectable as late as after 24 hours when the working muscle was already negative. Other enzyme activities (oxidoreductases and hydrolases) were practically uninfluenced by 6 hour autolysis. A substantial decrease of activities was found in conducting system after 48 hours in comparison to the working muscle. A solitary relatively strong residual activity belonged to acetylcholine-esterase in the conducting system. Detection of acetylcholine-esterase can be useful in identification of the conducting system even later after death.

Penicillin amidohydrolases in fungal autolysis.

The production of penicillin G and penicillin V amidohydrolases or acylases (E.C.3.5.1.11) was studied during the autolysis of filamentous fungi in a mineral medium, and in the same medium with phenoxyacetic acid as inducer. In all the studied fungi, enzymes showing penicillin G and penicillin V amidohydrolase activities were found. Generally, an increase of these activities during fungal autolysis was observed. The presence of phenoxyacetic acid in the medium did not increase these activities. The activities found in the culture fluids were generally higher than that found in the mycelial extracts. Under these conditions, beta-lactamases (penicillinases) were not found. The fungi Alternaria alternata, Fusarium culmorum, Penicillium oxalicum, and the species Penicillium 222 were chosen to study penicillin G and penicillin V acylases. The enzymes were precipitated with tannic acid from the culture fluid of their autolyzed cultures. Some kinetic constants of these activities were determined.