Genome editing: A perspective on the application of CRISPR/Cas9 to study human diseases (Review).

Genome editing reemerged in 2012 with the development of CRISPR/Cas9 technology, which is a genetic manipulation tool derived from the defense system of certain bacteria against viruses and plasmids. This method is easy to apply and has been used in a wide variety of experimental models, including cell lines, laboratory animals, plants, and even in human clinical trials. The CRISPR/Cas9 system consists of directing the Cas9 nuclease to create a site­directed double­strand DNA break using a small RNA molecule as a guide. A process that allows a permanent modification of the genomic target sequence can repair the damage caused to DNA. In the present study, the basic principles of the CRISPR/Cas9 system are reviewed, as well as the strategies and modifications of the enzyme Cas9 to eliminate the off­target cuts, and the different applications of CRISPR/Cas9 as a system for visualization and gene expression activation or suppression. In addition, the review emphasizes on the potential application of this system in the treatment of different diseases, such as pulmonary, gastrointestinal, hematologic, immune system, viral, autoimmune and inflammatory diseases, and cancer.

Olfactomedin-like 2 A and B (OLFML2A and OLFML2B) expression profile in primates (human and baboon)

BACKGROUND: The olfactomedin-like domain (OLFML) is present in at least four families of proteins, including OLFML2A and OLFML2B, which are expressed in adult rat retina cells. However, no expression of their orthologous has ever been reported in human and baboon. OBJECTIVE: The aim of this study was to investigate the expression of OLFML2A and OLFML2B in ocular tissues of baboons (Papio hamadryas) and humans, as a key to elucidate OLFML function in eye physiology. METHODS: OLFML2A and OLFML2B cDNA detection in ocular tissues of these species was performed by RT-PCR. The amplicons were cloned and sequenced, phylogenetically analyzed and their proteins products were confirmed by immunofluorescence assays. RESULTS: OLFML2A and OLFML2B transcripts were found in human cornea, lens and retina and in baboon cornea, lens, iris and retina. The baboon OLFML2A and OLFML2B ORF sequences have 96% similarity with their human's orthologous. OLFML2A and OLFML2B evolution fits the hypothesis of purifying selection. Phylogenetic analysis shows clear orthology in OLFML2A genes, while OLFML2B orthology is not clear. CONCLUSIONS: Expression of OLFML2A and OLFML2B in human and baboon ocular tissues, including their high similarity, make the baboon a powerful model to deduce the physiological and/or metabolic function of these proteins in the eye.

Molecular evolution and expression profile of the chemerine encoding gene RARRES2 in baboon and chimpanzee.

BACKGROUND: Chemerin, encoded by the retinoic acid receptor responder 2 (RARRES2) gene is an adipocytesecreted protein with autocrine/paracrine functions in adipose tissue, metabolism and inflammation with a recently described function in vascular tone regulation, liver, steatosis, etc. This molecule is believed to represent a critical endocrine signal linking obesity to diabetes. There are no data available regarding evolution of RARRES2 in non-human primates and great apes. Expression profile and orthology in RARRES2 genes are unknown aspects in the biology of this multigene family in primates. Thus; we attempt to describe expression profile and phylogenetic relationship as complementary knowledge in the function of this gene in primates. To do that, we performed A RT-PCR from different tissues obtained during necropsies. Also we tested the hypotheses of positive evolution, purifying selection, and neutrality. And finally a phylogenetic analysis was made between primates RARRES2 protein. RESULTS: RARRES2 transcripts were present in liver, lung, adipose tissue, ovary, pancreas, heart, hypothalamus and pituitary tissues. Expression in kidney and leukocytes were not detectable in either species. It was determined that the studied genes are orthologous. CONCLUSIONS: RARRES2 evolution fits the hypothesis of purifying selection. Expression profiles of the RARRES2 gene are similar in baboons and chimpanzees and are also phylogenetically related.

Molecular evolution and expression profile of the chemerine encoding gene RARRES2 in baboon and chimpanzee

BACKGROUND: Chemerin, encoded by the retinoic acid receptor responder 2 (RARRES2) gene is an adipocytesecreted protein with autocrine/paracrine functions in adipose tissue, metabolism and inflammation with a recently described function in vascular tone regulation, liver, steatosis, etc. This molecule is believed to represent a critical endocrine signal linking obesity to diabetes. There are no data available regarding evolution of RARRES2 in non-human primates and great apes. Expression profile and orthology in RARRES2 genes are unknown aspects in the biology of this multigene family in primates. Thus; we attempt to describe expression profile and phylogenetic relationship as complementary knowledge in the function of this gene in primates. To do that, we performed A RT-PCR from different tissues obtained during necropsies. Also we tested the hypotheses of positive evolution, purifying selection, and neutrality. And finally a phylogenetic analysis was made between primates RARRES2 protein. RESULTS: RARRES2 transcripts were present in liver, lung, adipose tissue, ovary, pancreas, heart, hypothalamus and pituitary tissues. Expression in kidney and leukocytes were not detectable in either species. It was determined that the studied genes are orthologous. CONCLUSIONS: RARRES2 evolution fits the hypothesis of purifying selection. Expression profiles of the RARRES2 gene are similar in baboons and chimpanzees and are also phylogenetically related.

Extrapituitary growth hormone synthesis in humans.

The gene for pituitary growth hormone (GH-N) in man belongs to a multigene locus located at chromosome 17q24.2, which also harbors four additional genes: one for a placental variant of GH-N (named GH-V) and three of chorionic somatommamotropin (CSH) type. Their tandem arrangement from 5' to 3' is: GH-N, CSH-L, CSH-1, GH-V and CSH-2. GH-N is mainly expressed in the pituitary from birth throughout life, while the remaining genes are expressed in the placenta of pregnant women. Pituitary somatotrophs secrete GH into the bloodstream to act at receptor sites in most tissues. GH participates in the regulation of several complex physiological processes, including growth and metabolism. Recently, the presence of GH has been described in several extrapituitary sites, such as neural, ocular, reproductive, immune, cardiovascular, muscular, dermal and skeletal tissues. It has been proposed that GH has an autocrine action in these tissues. While the body of evidence for its presence is constantly growing, research of its possible function and implications lag behind. In this review we highlight the evidence of extrapituitary synthesis of GH in humans.

Histoplasmosis oral atípica en paciente VIH reactivo: reporte de un caso / Atypical oral histoplasmosis in a HIV-reactive patient: report of a case

La histoplasmosis es una infección granulomatosa causada por un hongo dimórfico, el Histoplasma capsulatum. Clínicamente, existen tres formas: aguda, crónica y diseminada. Nosotros presentamos un caso de un paciente masculino de 19 años, reactivo para el VIH, en el estado C, quien fue derivado al Servicio de Odontología Hospital Juan A. Fernández, Buenos Aires, Argentina, en donde se realizó una evaluación estomatológica. El paciente presentaba erosiones, pápulas y nódulos indoloros de aproximadamente 2,4 mm en la mucosa bucal. Se realizaron tres biopsias correspondientes a cada tipo de lesión presente. Los estudios histológicos revelaron invasión celular por Histoplasma capsulatum. Este caso de reporta como una forma atípica de histoplasmosis en base a lesiones diferentes entre sí, lo cual hizo el diagnóstico difícil. Es común la observación de presentaciones inusuales de lesiones bucales en pacientes VIH positivos.

Histoplasmosis oral atípica en paciente VIH reactivo: reporte de un caso / Atypical oral histoplasmosis in a HIV-reactive patient: report of a case

La histoplasmosis es una infección granulomatosa causada por un hongo dimórfico, el Histoplasma capsulatum. Clínicamente, existen tres formas: aguda, crónica y diseminada. Nosotros presentamos un caso de un paciente masculino de 19 años, reactivo para el VIH, en el estado C, quien fue derivado al Servicio de Odontología Hospital Juan A. Fernández, Buenos Aires, Argentina, en donde se realizó una evaluación estomatológica. El paciente presentaba erosiones, pápulas y nódulos indoloros de aproximadamente 2,4 mm en la mucosa bucal. Se realizaron tres biopsias correspondientes a cada tipo de lesión presente. Los estudios histológicos revelaron invasión celular por Histoplasma capsulatum. Este caso de reporta como una forma atípica de histoplasmosis en base a lesiones diferentes entre sí, lo cual hizo el diagnóstico difícil. Es común la observación de presentaciones inusuales de lesiones bucales en pacientes VIH positivos.(AU)

[Dinoflagellates (Dinophyta) of orders Dinophysiales and Prorocentrales of the Veracruz Reef System, Mexico]. / Dinoflagelados (Dinophyta) de los ordenes Prorocentrales y Dinophysiales del Sistema Arrecifal Veracruzano, México.

Dinoflagellates are a major taxonomic group in marine phytoplankton communities in terms of diversity and biomass. Some species are also important because they form blooms and/or produce toxins that may cause diverse problems. The composition of planktonic dinoflagellates of the orders Prorocentrales and Dinophysiales, in the Veracruz Reef System, were obtained during the period of October 2006 to January 2007. For this, samples were taken from the surface at 10 stations with net of 30 microm mesh, and were analyzed by light and scanning electron microscopy. Each species was described and illustrated, measured and their distribution and ecological data is also given. A total of nine species were found and identified, belonging to four genera: Dinophysis was represented by three species; Prorocentrum by three, Phalacroma by two, and only one species of Ornithocercus was detected. From the samples, four potentially toxin-producer species were found: Dinophysis caudata, D. rapa, Phalacroma rotundata and Prorocentrum micans. The number of species found in this study is low, especially considering the higher numbers observed in other areas of the Gulf of Mexico, where some reports have recorded up to 53 species of the order Dinophysiales and 14 for Prorocentrales. Identification keys for orders, genera and species for the study area are provided with this study.

Dinoflagelados (Dinophyta) de los órdenes Prorocentrales y Dinophysiales del Sistema Arrecifal Veracruzano, México

Dinoflagellates (Dinophyta) of orders Dinophysiales and Prorocentrales of the Veracruz Reef System, Mexico. Dinoflagellates are a major taxonomic group in marine phytoplankton communities in terms of diversity and biomass. Some species are also important because they form blooms and/or produce toxins that may cause diverse problems. The composition of planktonic dinoflagellates of the orders Prorocentrales and Dinophysiales, in the Veracruz Reef System, were obtained during the period of October 2006 to January 2007. For this, samples were taken from the surface at 10 stations with net of 30μm mesh, and were analyzed by light and scanning electron microscopy. Each species was described and illustrated, measured and their distribution and ecological data is also given. A total of nine species were found and identified, belonging to four genera: Dinophysis was represented by three species; Prorocentrum by three, Phalacroma by two, and only one species of Ornithocercus was detected. From the samples, four potentially toxin-producer species were found: Dinophysis caudata, D. rapa, Phalacroma rotundata and Prorocentrum micans. The number of species found in this study is low, especially considering the higher numbers observed in other areas of the Gulf of Mexico, where some reports have recorded up to 53 species of the order Dinophysiales and 14 for Prorocentrales. Identification keys for orders, genera and species for the study area are provided with this study. Rev. Biol. Trop. 59 (1): 501-514. Epub 2011 March 01.

Los dinoflagelados son considerados como el grupo más abundante y diverso de microalgas del fitoplancton después de las diatomeas. Esta investigación da a conocer la composición específica de dinoflagelados planctónicos de los ordenes Prorocentrales y Dinophysiales, recolectados mediante arrastres superficiales con red (30μm de luz de malla) en el Sistema Arrecifal Veracruzano (SAV), de las costas de Veracruz, México, en el período octubre 2006 a enero2007. Para el análisis del material se utilizaron microscopios de luz y electrónico de barrido. Se identificaron 9 especies de cuatro géneros: Dinophysis estuvo representado por 3 especies, al igual que Prorocentrum, dos de Phalacroma y una de Ornithocercus. Se presentan claves para identificación de los órdenes, géneros y especies de dinoflagelados, asimismo se describe e ilustra cada especie, incluyendo datos morfométricos y se menciona la distribución de las mismas. Dinophysis caudata, D. rapa, D. ovum y Phalacroma rotundata se han documentado como especies potencialmente productoras de toxinas. La riqueza de especies en el SAV se considera muy baja, especialmente considerando mayores riquezas en otras zonas del Golfo de México, como el sur del Golfo de México, donde se han listado 53 especies del orden Dinophysiales y 14 especies de Prorocentrales.

Serologic surveillance for West Nile virus and other flaviviruses in febrile patients, encephalitic patients, and asymptomatic blood donors in northern Mexico.

A clinical and serological investigation was performed to determine the presence of West Nile virus (WNV) among febrile and encephalitic patients in northern Mexico. In addition, asymptomatic blood donors were serologically assayed for WNV to determine the seroprevalence of WNV in the general population. The study cohort consisted of 1432 individuals (588 febrile patients, 44 encephalitic patients, and 800 asymptomatic blood donors). All subjects were negative for WNV IgM. Sixty subjects were reactive for dengue virus (DENV) IgM (16 blood donors and 44 febrile patients). A subset (n = 425) of individuals was also screened by ELISA for flavivirus IgG. The prevalence of flavivirus IgG in febrile patients, encephalitic patients, and blood donors ranged from 40% to 59%. A subset (n = 147) of sera reactive for flavivirus IgG was further tested by plaque reduction neutralization test. Six individuals with no history of travel during the preceding 12 months were seropositive for WNV. Another 65 individuals were seropositive for DENV1 and 24 were seropositive for DENV2. The high prevalence of dengue antibodies in northern Mexico appears to limit the incidence of WNV infection in this region. Article Summary Line: Antibodies to WNV, DENV-1, and DENV-2 were identified in humans in northern Mexico.

Hallazgos patológicos del tracto reproductivo en cerdas de descarte en Venezuela

En el avance hacia la obtención de la productividad del rebaño, es fundamental el adecuado conocimiento tanto de la anatomía como de la fisiología de la reproducción porcina. Dentro de la producción, uno de los parámetros que incide con mayor relevancia en la productividad es el número de lechones destetados por cerda por año, ya que de él depende el número final de cerdos que va a matadero. Este parámetro va a estar influenciado directamente por el desempeño reproductivo del rebaño de cría. Las fallas reproductivas son una de las principales causas de descarte, de ahí que el examen de los órganos reproductivos es una herramienta de gran valor diagnóstico en el manejo reproductivo porcino. La evaluación de estos órganos provee de una serie de datos sobre las posibles causas de falla reproductiva y a su vez, va a permitir tomar medidas correctivas a nivel de rebaño. En Venezuela, no existen reportes que describan los hallazgos patológicos observados en cerdas de descarte. De aquí que se plantea como objetivo general de esta investigación describir cuáles eran los hallazgos patológicos presentes en el tracto reproductivo de cerdas a través del análisis de muestras recolectadas en un matadero industrial del estado Aragua. Se evaluó un total de 125 tractos reproductivos encontrándose diferentes hallazgos macroscópicos en 23 animales (18,4%), desglosándose los mismos de la manera siguiente: 8,94% (11/123) de animales con ovarios inactivos o anestro (sin cuerpos lúteos ni folículos o con folículos con un diámetro < 5 mm), 13,01% (16/123) presentaron quistes en los ovarios, 5,6% (7/125) mostraron úteros con material purulento y de éstos en uno se encontró un feto retenido en proceso de descomposición y otro con ausencia de oviducto (cuerno ciego); en 17,6% (22/125) de los animales se evidenció procesos inflamatorios compatibles con cuadros de endometritis.

Pathological Findings of the Reproductive Tract from Culled Sows in Venezuela Abstract In order to advance towards the attainment of productivity in swine production, the adequate knowledge of the anatomy, as well as of the reproductive physiology of the sow is essential. In terms of production, one of the parameters that greatly influence productivity is the number of pigs weaned by sow per year, as this parameter determines the number of pigs delivered to the market. This parameter is directly influenced by the reproductive performance of the herd. Reproductive failure is one of the major causes of culling sows; therefore, a thorough examination of the reproductive tract is a valuable diagnostic tool in swine reproductive management. The evaluation of these organs provides us with a series of data on the possible causes of reproductive failure. At the same time, such an evaluation, will allow us to take corrective measures in the herd. In Venezuela, there is no available data describing the pathological findings observed in the reproductive tract of culled sows. The purpose of this investigation was to describe the pathological findings that affect the reprodcutive tract of culled sows. Samples were collected from a slaughterhouse, located in the State of Aragua, Venezuela. A total of 125 reproductive tracts was collected. Results of this investigation show that only 23 animals (18.4%)exhibited abnormal findings which were classified as follows: 8.94% (11/123) sows with inactive ovaries or anestro (neither corpora lutea nor follicles, or follicles with a diameter < 5 mm); 13.01% (16/123) had evidence of ovarian cysts, and 5.6% (7/125) showed uteri with purulent material: one had a retained fetus in decomposition process, and another lacked an oviduct (blind horn); 17.6% of the animals showed inflammatory processess compatible with endometritis.

Presencia del virus del oeste del Nilo en el noreste de México / Presence of west Nile virus in northeast Mexico

OBJETIVO: Detectar la presencia del virus del oeste del Nilo (VON) en aves, equinos y seres humanos en el noreste de México. MATERIAL Y MÉTODOS: Se buscó en diferentes localidades del noreste de México la presencia de anticuerpos antivirus del oeste del Nilo (anti-VON) en suero de 33 aves, 24 caballos y 237 personas mediante pruebas de ELISA durante el periodo de julio de 2003 a julio de 2006. En los sueros humanos se buscó también el RNA-VON mediante RT-PCR. RESULTADOS: Se encontraron tres aves seropositivas y 15 equinos. En el hombre, 40 por ciento de los sueros fue positivo para anticuerpos IgG y ninguno para anticuerpos IgM. CONCLUSIONES: El VON se encuentra activo en México y se suma a otras enfermedades emergentes transmitidas por vectores que representan un reto a la investigación y a los programas de prevención.

OBJECTIVE: To investigate the presence of WNV in birds, horses and humans in northeast Mexico. MATERIAL AND METHODS: Serum samples from 33 birds, 24 horses and 237 humans were screened by ELISA for Anti-WNV antibodies. Human serum samples were also screened for WNV RNA using an RT-PCR assay. RESULTS: Positive sera were found in three birds and 15 horses. Forty percent of the human serum samples were positive for IgG antibodies and 0 percent for IgM antibodies and viral RNA. CONCLUSIONS: The results of this study show that WNV is present in northeast Mexico and it is a new emergent infectious agent that represents a challenge for research and prevention programs in Mexico.

Cloning of the growth hormone cDNA of alligator gar Atractosteus spatula and its expression through larval development.

The alligator gar (Atractosteus spatula) is the largest freshwater fish inhabiting rivers draining into the Gulf of Mexico. This primitive fish shows a fast growth rate since its early larval stages. This is attributed to the action of growth hormone (GH), an anterior pituitary gland hormone responsible for linear growth in vertebrates that can also be expressed in extrapituitary adult tissues and in fish embryos. The present research was aimed at obtaining the GH coding sequence of the alligator gar and studying its expression through larval development. A cDNA was obtained by RT-PCR, cloned and sequenced. The alligator gar GH cDNA sequence shares 98% nucleotide similarity with that reported for Lepisosteus osseus, indicating a very slow evolution of the GH within the primitive fish, in contrast with the burst of changes observed in euteleosts. Using RT-PCR and RNA nuclease protection assays, GH transcripts were detected at very high levels in eggs, embryos and in several larval stages. These data suggest that the GH may play an important role during embryogenesis in fish. The better understanding of alligator gar larval physiology will facilitate the culture of larvae and juvenile gar and consequently may allow the restoration of their natural populations.

Correlación de los hallazgos histopatológicos con las variables ultrasonográficas del endometrio conforme al peso corporal y distribución del tejido adiposo en la posmenopausia / Correlation of histopathologic findings with ultrasonographic variables of endometrium in relation with body weight and adipose tissue distribution in postmenopause

El objetivo de este trabajo fue correlacionar los hallazgos histopatologícos y las variables ultrasonográficas del endometrio en relación al peso corporal y distribución del tejido adiposo, en dos grupos de mujeres: uno con sangrado uterino posmenopáusico y otro bajo terapia hormonal de reemplazo (THR). fueron estudiadas 31 mujeres posmenopáusicas divididas en dos grupos: I. mujeres con sangrado uterino posmenopáusico (n= 19) y II. mujeres con THR (estrógenos conjugados y clormadinona) (n= 12). En ambos grupos se realizó el mismo día ultrasonido transvaginal previo a la biopsia de endometrio (BE). La BE se tomó en el grupo I el día que acudieron a consulta refiriendo sangrado y en el grupo II en el día 21 del ciclo de THR. Se observó que en el grupo I existió correlación significativa entre el peso corporal y los hallazgos histológicos y entre la refringencia del endometrio y los hallazgos histológicos. En el grupo II hubo correlación significativa entre la distribución de tejido adiposo y la refringencia del endometrio y entre el espesor endometrial y los hallazgos histológicos. Por lo que se puede concluir que en la mujer con sangrado posmenopáusico el peso corporal influye sobre los hallazgos histológicos y la refringencia está en relación del hallazgo histológico, mientras que en aquellas con THR es más importante la distribución del tejido adiposo sobre la refringencia del endometrio y el espesor endometrial tiene relación con los hallazgos histológicos.