Capacitively coupled electrical stimulation of rat chondroepiphysis explants: A histomorphometric analysis.

The growth plate is a cartilaginous layer present from the gestation period until the end of puberty where it ossifies joining diaphysis and epiphysis. During this period several endocrine, autocrine, and paracrine processes within the growth plate are carried out by chondrocytes; therefore, a disruption in cellular functions may lead to pathologies affecting bone development. It is known that electric fields impact the growth plate; however, parameters such as stimulation time and electric field intensity are not well documented. Accordingly, this study presents a histomorphometrical framework to assess the effect of electric fields on chondroepiphysis explants. Bones were stimulated with 3.5 and 7 mV/cm, and for each electric field two exposure times were tested for 30 days (30 min and 1 h). Results evidenced that electric fields increased the hypertrophic zones compared with controls. In addition, a stimulation of 3.5 mV/cm applied for 1 h preserved the columnar cell density and its orientation. Moreover, a pre-hypertrophy differentiation in the center of the chondroepiphysis was observed when explants were stimulated during 1 h with both electric fields. These findings allow the understanding of the effect of electrical stimulation over growth plate organization and how the stimulation modifies chondrocytes morphophysiology.

Gene's hubs in retinal diseases: A retinal disease network.

BACKGROUND: Retinal diseases associated with the dysfunction or death of photoreceptors are a major cause of blindness around the world, improvements in genetics tools, like next generation sequencing (NGS) allows the discovery of genes and genetic changes that lead to many of those retinal diseases. Though, there very few databases that explores a wide spectrum of retinal diseases, phenotypes, genes, and proteins, thus creating the need for a more comprehensive database, that groups all these parameters. METHODS: Multiple open access databases were compiled into a new comprehensive database. A biological network was then crated, and organized using Cytoscape. The network was scrutinized for presence of hubs, measuring the concentration of grouped nodes. Finally, a trace back analysis was performed in areas were the power law reports a high r-squared value near one, that indicates high nodes density. RESULTS: This work leads to creation of a retinal database that includes 324 diseases, 803 genes, 463 phenotypes, and 2461 proteins. Four biological networks (1) a disease and gene network connected by common phenotypes, (2) a disease and phenotype network connected by common genes, (3) a disease and gene network with shared disease or gene as the cause of an edge, and (4) a protein and disease network. The resulting networks will allow users to have easier searching for retinal diseases, phenotypes, genes, and proteins and their interrelationships. CONCLUSIONS: These networks have a broader range of information than previously available ones, helping clinicians in the comprehension of this complex group of diseases.

Mechanobiological modeling of endochondral ossification: an experimental and computational analysis.

Long bone formation starts early during embryonic development through a process known as endochondral ossification. This is a highly regulated mechanism that involves several mechanical and biochemical factors. Because long bone development is an extremely complex process, it is unclear how biochemical regulation is affected when dynamic loads are applied, and also how the combination of mechanical and biochemical factors affect the shape acquired by the bone during early development. In this study, we develop a mechanobiological model combining: (1) a reaction-diffusion system to describe the biochemical process and (2) a poroelastic model to determine the stresses and fluid flow due to loading. We simulate endochondral ossification and the change in long bone shapes during embryonic stages. The mathematical model is based on a multiscale framework, which consisted in computing the evolution of the negative feedback loop between Ihh/PTHrP and the diffusion of VEGF molecule (on the order of days) and dynamic loading (on the order of seconds). We compare our morphological predictions with the femurs of embryonic mice. The results obtained from the model demonstrate that pattern formation of Ihh, PTHrP and VEGF predict the development of the main structures within long bones such as the primary ossification center, the bone collar, the growth fronts and the cartilaginous epiphysis. Additionally, our results suggest high load pressures and frequencies alter biochemical diffusion and cartilage formation. Our model incorporates the biochemical and mechanical stimuli and their interaction that influence endochondral ossification during embryonic growth. The mechanobiochemical framework allows us to probe the effects of molecular events and mechanical loading on development of bone.

Cyclic Concatenated Genetic Encoder: A mathematical proposal for biological inferences.

The organization of the genetic information and its ability to be conserved and translated to proteins with low error rates have been the subject of study by scientists from different disciplines. Recently, it has been proposed that living organisms display an intra-cellular transmission system of genetic information, similar to a model of digital communication system, in which there is the ability to detect and correct errors. In this work, the concept of Concatenated Genetic Encoder is introduced and applied to the analysis of protein sequences as a tool for exploring evolutionary relationships. For such purposes Error Correcting Codes (ECCs) are used to represent proteins. A methodology for representing or identifying proteins by use of BCH codes over ℤ20 and F4×ℤ5 is proposed and cytochrome b6-f complex subunit 6-OS sequences, corresponding to different plants species, are analyzed according to the proposed methodology and results are contrasted to phylogenetic and taxonomic analyses. Through the analyses, it was observed that using BCH codes only some sequences are identified, all of which differ in one amino acid from the original sequence. In addition, mathematical relationships among identified sequences are established by considering minimal polynomials, where such sequences showed a close relationship as revealed in the phylogenetic reconstruction. Results, here shown, point out that communication theory may provide biology of interesting and useful tools to identify biological relationships among proteins, however the proposed methodology needs to be improved and rigorously tested in order to become into an applicable tool for biological analysis.

Cellular scale model of growth plate: An in silico model of chondrocyte hypertrophy.

The growth plate is the responsible for longitudinal bone growth. It is a cartilaginous structure formed by chondrocytes that are continuously undergoing a differentiation process that starts with a highly proliferative state, followed by cellular hypertrophy, and finally tissue ossification. Within the growth plate chondrocytes display a characteristic columnar organization that potentiates longitudinal growth. Both chondrocyte organization and hypertrophy are highly regulated processes influenced by biochemical and mechanical stimuli. These processes have been studied mainly using in vivo models, although there are few computational approaches focused on the rate of ossification rather than events at cellular level. Here, we developed a model of cellular behavior integrating biochemical and structural factors in a single column of cells in the growth plate. In our model proliferation and hypertrophy were controlled by biochemical regulatory loop formed between Ihh and PTHrP (modeled as a set of reaction-diffusion equations), while cell growth was controlled by mechanical loading. We also examined the effects of static loading. The model reproduced the proliferation and hypertrophy of chondrocytes in organized columns. This model constitutes a first step towards the development of mechanobiological models that can be used to study biochemical interactions during endochondral ossification.

Cellular automata model for human articular chondrocytes migration, proliferation and cell death: An in vitro validation.

Articular cartilage is characterized by low cell density of only one cell type, chondrocytes, and has limited self-healing properties. When articular cartilage is affected by traumatic injuries, a therapeutic strategy such as autologous chondrocyte implantation is usually proposed for its treatment. This approach requires in vitro chondrocyte expansion to yield high cell number for cell transplantation. To improve the efficiency of this procedure, it is necessary to assess cell dynamics such as migration, proliferation and cell death during culture. Computational models such as cellular automata can be used to simulate cell dynamics in order to enhance the result of cell culture procedures. This methodology has been implemented for several cell types; however, an experimental validation is required for each one. For this reason, in this research a cellular automata model, based on random-walk theory, was devised in order to predict articular chondrocyte behavior in monolayer culture during cell expansion. Results demonstrated that the cellular automata model corresponded to cell dynamics and computed-accurate quantitative results. Moreover, it was possible to observe that cell dynamics depend on weighted probabilities derived from experimental data and cell behavior varies according to the cell culture period. Thus, depending on whether cells were just seeded or proliferated exponentially, culture time probabilities differed in percentages in the CA model. Furthermore, in the experimental assessment a decreased chondrocyte proliferation was observed along with increased passage number. This approach is expected to having other uses as in enhancing articular cartilage therapies based on tissue engineering and regenerative medicine.

Acid ceramidase treatment enhances the outcome of autologous chondrocyte implantation in a rat osteochondral defect model.

OBJECTIVE: The overall aim of this study was to evaluate how supplementation of chondrocyte media with recombinant acid ceramidase (rhAC) influenced cartilage repair in a rat osteochondral defect model. METHODS: Primary chondrocytes were grown as monolayers in polystyrene culture dishes with and without rhAC (added once at the time of cell plating) for 7 days, and then seeded onto Bio-Gide® collagen scaffolds and grown for an additional 3 days. The scaffolds were then introduced into osteochondral defects created in Sprague-Dawley rat trochlea by a microdrilling procedure. Analysis was performed 6 weeks post-surgery macroscopically, by micro-CT, histologically, and by immunohistochemistry. RESULTS: Treatment with rhAC led to increased cell numbers and glycosaminoglycan (GAG) production (∼2 and 3-fold, respectively) following 7 days of expansion in vitro. Gene expression of collagen 2, aggrecan and Sox-9 also was significantly elevated. After seeding onto Bio-Gide®, more rhAC treated cells were evident within 4 h. At 6 weeks post-surgery, defects containing rhAC-treated cells exhibited more soft tissue formation at the articular surface, as evidenced by microCT, as well as histological evidence of enhanced cartilage repair. Notably, collagen 2 immunostaining revealed greater surface expression in animals receiving rhAC treated cells as well. Collagen 10 staining was not enhanced. CONCLUSION: The results further demonstrate the positive effects of rhAC treatment on chondrocyte growth and phenotype in vitro, and reveal for the first time the in vivo effects of the treated cells on cartilage repair.

Growth plate stress distribution implications during bone development: a simple framework computational approach.

Mechanical stimuli play a significant role in the process of long bone development as evidenced by clinical observations and in vivo studies. Up to now approaches to understand stimuli characteristics have been limited to the first stages of epiphyseal development. Furthermore, growth plate mechanical behavior has not been widely studied. In order to better understand mechanical influences on bone growth, we used Carter and Wong biomechanical approximation to analyze growth plate mechanical behavior, and explore stress patterns for different morphological stages of the growth plate. To the best of our knowledge this work is the first attempt to study stress distribution on growth plate during different possible stages of bone development, from gestation to adolescence. Stress distribution analysis on the epiphysis and growth plate was performed using axisymmetric (3D) finite element analysis in a simplified generic epiphyseal geometry using a linear elastic model as the first approximation. We took into account different growth plate locations, morphologies and widths, as well as different epiphyseal developmental stages. We found stress distribution during bone development established osteogenic index patterns that seem to influence locally epiphyseal structures growth and coincide with growth plate histological arrangement.

Penetrating stab wounds to the abdomen: use of serial US and contrast-enhanced CT in stable patients.

PURPOSE: To evaluate the usefulness of computed tomography (CT) and ultrasonography (US) for the initial assessment of penetrating abdominal stab wounds in patients who presented to the emergency department without indication for immediate laparotomy. MATERIALS AND METHODS: During 36 months, 32 patients with a penetrating stab wound to the abdomen were examined with serial US (at admission and 12 hours later) and helical CT, with contrast material administered orally, intravenously, and rectally. Presence of hemoperitoneum and integrity of solid and hollow viscera were evaluated with both methods. Sonograms were interpreted by the radiologist who performed the examination, and CT images were independently evaluated by two radiologists. Findings of both techniques were compared with clinical outcome and/or surgical findings. RESULTS: One (3.1%) of 32 patients required surgery: Surgical findings were massive hemoperitoneum and an extensive hepatic laceration. Both US and CT depicted these abnormalities. Thirty-one (96.9%) patients were treated conservatively, without surgery, and remained asymptomatic during 28 days of clinical follow-up after discharge from the hospital. US and/or CT showed intraperitoneal abnormalities in 21 of these patients. In 11 patients, both methods showed no evidence of visceral injury or hemoperitoneum, and none of these patients required surgery. CONCLUSION: Serial US and CT help guide treatment for stable patients with penetrating stab injuries to the abdomen.

Determination of beta-globin gene cluster haplotypes and prevalence of alpha-thalassemia in sickle cell anemia patients in Venezuela.

Sickle cell anemia and alpha-thalassemia have a heterogeneous distribution in Venezuela with a high frequency in the coastal area (sea level) and few cases in the mountains. Most of our population is an ethnic admixture of Europeans (Spaniards colonists), Africans (slaves), and Amerindians. The purpose of our study was to determine the origin of the beta(s) globin haplotype, age and survival dependency, and the admixture among the different African groups in our population. The alpha(3.7) globin gene deletion status was also studied and found in a very high frequency. DNA from peripheral blood of 191 non-related patients (81 with HbS homozygous and 15 patients compound heterozygous for HbS, HbC, HbD with beta-thalassemia, and 95 with sickle cell trait) were studied. The beta(s) chromosome was linked 51% to the Benin Haplotype, 29.5% with the CAR, 12.5% to the Senegal, and 2.5% to the Cameroon. We did not find any significant difference between the haplotype distribution among adults and children and among sickle cell patients and traits. Only 8.6% of the patients have homozygosity for the Benin haplotype. These results show a very high frequency of admixture in our African origin population.

[Detection of beta thalassemia by the technique of refractory amplification of mutation systems (ARMS-PCR)]. / Detección de beta Talasemia mediante la técnica de amplificación refractaria de sistemas de mutaciones (ARMS-PCR).

beta Thalassemia (Thal) mutations were studied in DNA from 80/159 patients with hemolytic anemia and high levels of Hb A2 by the amplification refractory mutation system technique (ARMS-PCR). This method detects point mutations and insertions or deletions of just a few nucleotides in the beta globin gene by the polymerase chain reaction of allele-specific priming. In 43/80 patients with different clinical presentations of beta Thalassemia and 37/80 compound heterozygous for hemoglobinopathies and beta Thalassemia the most frequent mutation found was the -29 (of African origin), followed by the CD39 (of Mediterranean origin) and in a lower frequency also was found the -88, the IVSI-6 and the IVSI-110. We conclude that this technique is an useful approach in determining the beta thalassemia mutations in population surveys, because it allows to make a differential diagnosis between beta Thalassemia minor and individuals with high levels of Hb A2. It helps to clarify the diagnosis of patients with structural hemoglobinopathies that also presents high levels of Hb A2.

Antibiotic-resistant Salmonella typhi from two outbreaks: few ribotypes and IS200 types harbor Inc HI1 plasmids.

To investigate factors that could be involved in the emergence of antibiotic resistant S. typhi, we characterized R plasmids and antibiotic resistant S. typhi strains from two outbreaks of typhoid in Peru and Chile. Differences in the Inc HI1 plasmids of Peruvian and Chilean strains were identified by conjugation and incompatibility studies and plasmid DNA characterization. Antibiotic-resistant S. typhi harboring Inc HI1 plasmids belonged to a reduced number of Pst1 and Cla1 ribotypes and IS200 types, in contrast to the high genetic diversity found among epidemic antibiotic-susceptible S. typhi. The low diversity of antibiotic-resistant S. typhi suggests that they may express properties that are related to both their ability to harbor Inc HI1 R plasmids and to disseminate.

Resistencia a oxacilina en staphylococcus aislados de bacteremias intrahospitalarias / Resistance a oxaciline in Staphylococcus isolate of bacteremias intrahospital

Se identificó la bacteria asociada a 264 bacteremias ocurridas en pacientes de 4 grandes hospitales de Lima y se evaluó la sensibilidad a oxacilina y a otros antibióticos (amikacina, cefotixina, ceftazimida, ceftriaxona, ciprofloxacina, gentamicina y vancomicina) de las cepas de estafilococo coagulasa negativo (209) y de estafilococo dorado (63) aisladas. La frecuencia relativa de cepas resistentes a oxacilina para cada hospital varió entre 52 por ciento y 0 por ciento para los estafilococos coagulasa negativos, y entre 63 por ciento y 7 por ciento para los estafilococos dorados. Igualmente variable fue la sensibilidad de los estafilococos resistentes a oxacilina a otros antibióticos, excepto para la vancomicina a la que fueron sensibles el 100 por ciento de estas cepas. Se concluye que es necesario considerar la posibilidad de resistencia a oxacilina en los aislamientos intrahospitalarios de estafilococos, casos en los que que la vancomicina debe considerarse como una alternativa, y que es importante mantener una vigilancia de la sensibilidad antibiótica de los gérmenes en general en cada hospital.

[The in vitro action of plants on Vibrio cholerae]. / Acción in vitro de plantas sobre Vibrio cholerae.

Natural products of several plants, according to the geographic location, are used by Peruvian people in the popular treatment of diarrhea, with good success. When cholerae cases appeared in Peru, we were interested to know the "in vitro" effect against Vibrio cholerae 01, of these useful plants to treat diarrhea. The following plants were tested: Cichorium intybus, Althaea officinalis, Psorela glandulosa, Geranium maculatum, Punica granatum, Malus sativa, Cydonia oblonga, Chenopodium ambrosoides, Krameria triandria, Tea chinensis, Daucus carota, Persea gratissima, Psidium guayaba and Lippia dulcis. Decoction or infusion of the plants were used in the "in vitro" experiments. The following plants showed no "in vitro" effect against V. cholerae: Cichorium intybus, Althaea officinalis, Psorela glandulosa, Geranium maculatum, Chenopodium ambrosoides, Krameria triandria, Psidium guayaba, Lippia dulcis and Daucus carota. Decoction of Malus sativa and Cydenia oblonga showed bactericidal effect for their acidity and stone avocado (Persea gratissima) a late bactericidal effect. Tea infusión and the decoction of Punica granatum peel, showed the best bactericidal effect and we suggest to use them as to stop cholera spreading.

Plasmid-mediated resistance to multiple antibiotics in Salmonella typhi.

Two hundred forty-one strains of Salmonella typhii isolated in Lima, Peru, from October 1981 through February 1983 were tested for susceptibility to antimicrobial agents. Seventy-two strains (29.9%) were resistant to chloramphenicol and other antibiotics, including ampicillin, sulfonamides, and trimethoprim. The minimal inhibitory concentrations of 17 antimicrobial agents were determined for all of these chloramphenicol-resistant strains. Sch 25393, new beta-lactams, new quinolones, and the formulation clavulanic acid-amoxicillin were effective against all the strains. Four different resistance patterns distributed among eight phage types were observed. The 72 resistant S. typhi could transfer the resistance marker into Escherichia coli C1, and all the plasmids belonged to the incompatibility group H1.

Hemoglobin Hofu associated with beta 0-thalassemia.

A mild anemia (hemoglobin 9 g/dl) was found in a patient from Seville (Spain) with marked morphological abnormalities in the peripheral blood smear. The red cell osmotic fragility showed a mild resistance curve with a mean cell fragility (MCF) of 0.375% NaCl (normal = 0.450). Chemical Chemical and thermal instability test and search for inclusion bodies gave positive results. Hemoglobin electrophoresis at pH 8.9 revealed absence of Hb A, a major component of fast mobility (94%), and increased Hb F and Hb A2 levels (1.5% and 4.6%, respectively). The fast fraction, isolated and purified by means of cellulose acetate electrophoresis, precipitated in acid acetone and treated with urea 8 M and mercaptoethanol, revealed an anomalous beta chain. Trypsin-digested globin peptides were separated by high-voltage electrophoresis at pH 6.4 and ascendant chromatography. With differential staining, an extra peptide was detected in an unusual site, more anodic than alpha Tp4 but in lower position. Peptide map of the fast beta chain, stained with ninhydrin, and also for Tyr, confirmed the position of the new peptide and the absence of the usual beta Tp13. The new peptide, separated by high-voltage electrophoresis at pH 3.5, revealed absence of Val and the presence of an additional Glu residue, which should appear only in position beta 126. The diagnosis of Hb Hofu (alpha 2 beta 2 126 Val----Glu; H4) was reached, thus interpreting its increase and the absence of Hb A, as an association with beta o-thalassemia, producing a mild hemolytic anemia. Evidence was obtained that Hb Hofu is a mild unstable hemoglobin variant.

A case of hematometra infected by Listeria monocytogenes.

A case of Listeria infection is reported of a 10 year old girl with symptoms of the first menstruation but without the discharge of blood because of the absence of the vaginal duct. The blood retained in the uterus obtained by puncture through the rectum contained Listeria monocytogenes 4b. The infection was possibly acquired from a household pet.

Human listeriosis in Peru.

In a survey on listeriosis in Peru a study was made of 653 stool samples; material from 8 cases of perinatal death, in which a macroscopic diagnosis of listeriosis had been made at autopsy; 1 sample of vaginal secretion from a woman with abortion due to listeriosis; and 1 case of abscess which turned out to be an hematometra. The following results have been obtained: 1. From the faeces 7 isolates of Listeria (1.07%) were obtained: 3 belonged to L. monocytogenes serovar 4ab, 3 to L. innocua (the former "4g"--nownow serovar 6b) and 1 to L. monocytogenes serovar 4a. 2. The 8 strains of autopsy materials belonged to L. monocytogenes: 6 serovar 4b and 2 serovar 4d. 3. From the vaginal secretion L. monocytogenes serovar 4b was cultured. 4. The strain from the hematometra was identified as L. monocytogenes serovar 4b. The serovars 4a, 4ab and 4d are at this time the only ones identified in Peru as cause of human listeriosis. The serovars of serogroup 1/2 frequent in Western and primarily in Central Europe, were so far never cultivated.