Challenges in the use of nanostructures as carriers of nucleic acids in clinical practice

ABSTRACT The delivery of nucleic acids to cells is considered a crucial step for the success of genetic modifications aimed at therapeutic purposes or production of genetically modified animals. In this context, nanotechnology is one of the most promising fields of science, with the potential to solve several existing problems. Nanostructures have desirable characteristics to be used as carriers, such as nanometric size, large surface area, cell internalization capacity, prolonged and controlled release, among others. Genetically modified animals can contribute to the production of biopharmaceuticals, through the expression of high-associated-value molecules. The production of these animals, also known as biofactories, further enhances Brazilian agribusiness, since it allows adding value to the final product, and favors the integration between the agricultural market and the pharmaceutical sector. However, there is a growing concern about the safety and possible harmful effects of nanostructures, since data on the safe use of these materials are still insufficient. The objective of this review was to address aspects of the use of nanostructures, mainly carbon nanotubes as nucleic acid carriers, aiming at the production of genetically modified animals, with the certainty that progress in this field of knowledge depends on more information on the mechanisms of interaction between nanostructures, cells and embryos, as well as on its toxicity.

Desenlaces en ciclos de transferencia con embriones vitrificados: análisis de dos esquemas de preparación endometrial / Results in cycles of frozen-thawed embryo transfer: Analysis of two protocols of endometrial preparation

Resumen: OBJETIVO: Comparar la tasa de recién nacido vivo con la de embarazo clínico-transferencia embrionaria, las características clínicas y concentraciones hormonales entre dos esquemas de preparación endometrial para transferencia de embriones desvitrificados: con un agonista de GnRH versus su sustitución con dosis altas de estrógenos. MATERIALES Y MÉTODOS: Estudio observacional, de cohorte histórica, efectuado en pacientes atendidas en el Instituto Nacional de Perinatología en protocolo FIV-ICSI entre enero 2017-marzo 2019. Se compararon dos esquemas de preparación endometrial: grupo A con agonista de GnRH y grupo B con estradiol a dosis de 8 mg al día sin agonista de GnRH. A todas las pacientes se les tomaron muestras de sangre para determinación de FSH, LH, estradiol y progesterona y se dio seguimiento ultrasonográfico durante la preparación endometrial. RESULTADOS: En 99 pacientes entre 23 y 42 años, con embriones congelados, la tasa de recién nacido vivo-transferencia embrionaria fue, respectivamente, de 17.2 y 8.6% (OR 1.98; IC95%: 077-1.53) y la de embarazo clínico de 26.5 y 22.8% (OR1.09; IC95%: 0.77-1.53) en los grupos A y B, respectivamente. La duración total del ciclo, los días con estrógenos y la concentración sérica de estradiol al inicio de la progesterona alcanzaron diferencia significativa entre los grupos. No se detectaron datos clínicos ni de laboratorio sugerentes de ovulación. CONCLUSIONES: La tasa de recién nacido vivo fue mayor con agonistas de GnRH sin alcanzar significación; sin embargo, la preparación endometrial solo con estrógenos es un esquema seguro, de menor costo y más amigable para la paciente y el médico.

Abstract: OBJECTIVE: To compare the rate of live newborn with clinical pregnancy-embryo transfer, clinical characteristics and hormonal concentrations between two endometrial preparation schemes for devitrified embryo transfer: with a GnRH agonist (aGnRH) versus its replacement with high doses of estrogens. MATERIALS AND METHODS: Historical cohort observational study conducted in patients treated at the National Institute of Perinatology under IVF-ICSI protocol between January 2017-March 2019. Two endometrial preparation schemes were compared: group A with GnRH agonist and group B with estradiol at 8 mg per day without aGnRH. All patients had blood samples taken for FSH, LH, estradiol and progesterone determination and ultrasonographic follow-up during endometrial preparation. RESULTS: in 99 patients between 23 and 42 years of age, with frozen embryos, the live birth-embryo transfer rate was 17.2 and 8.6%, (OR 1.98; CI95%: 077-1.53) and the clinical pregnancy rate was 26.5 and 22.8% (OR1.09; CI95%: 0.77-1.53) in groups a and b, respectively. total cycle length, estrogen days and serum estradiol concentration at the beginning of progesterone reached significant difference between groups. no clinical or laboratory data suggesting ovulation were detected. CONCLUSIONS: The rate of live newborn was higher with GnRH agonists without reaching significance; however, endometrial preparation with estrogens alone is a safe, lower cost and more patient- and physician-friendLy scheme.

Developmental stages and viability of Toxocara canis eggs outside the host / Desarrollo y viabilidad de huevos de Toxocara canis

ABSTRACT Introduction: Toxocariasis is a soil-transmitted zoonotic disease caused mainly by ingestion of larvated eggs of Toxocara canis. Objectives: To study the morphology of the intraovular developmental stages of Toxocara canis in culture, characterize non-viable eggs and the sequences of larval molting and compare the viability of eggs at the early stages of division and at reaching full maturation. Material and methods: Observation of developing embryos and characterization of non-viable eggs were done using light microscope. The proportions of viable eggs during embryonation were compared to the proportions of viable mature eggs. Results: Cell division commenced after 24 hours of cultivation. Early stages were found to be present over a period of 3-5 days. The developmental stages identified were eggs with: One cell, two cells, three cells, four cells, early morula, late morula, blastula, gastrula, tadpole, pre-larva, first, second and third stage larva. Two larval molts occurred. Non-viable eggs had degenerated cytoplasm, thin or collapsed shell and the larvae did not move after exposure to light. No significant differences were found between the proportions of viable eggs from day five to day 21 as compared to viability of fully mature eggs (30 days). Conclusion: Developing embryos in the environment may be considered as a potential threat to the public health. The precise identification of developmental stages and the clear differentiation of viable and non-viable eggs can help in determining an accurate baseline rate of development that could be used in studies of ovicidal compounds.

RESUMEN Introducción. La toxocariasis es una enfermedad zoonótica transmitida por contacto con el suelo contaminado y causada principalmente por la ingestión de huevos larvados de Toxocara canis. Objetivos. Estudiar la morfología de los estadios intraovulares en desarrollo de T. canis en cultivo, caracterizar los huevos no viables y las secuencias de las mudas larvarias, y comparar la viabilidad de los huevos en las etapas tempranas de división y al alcanzar la maduración completa. Materiales y métodos. Se observó el desarrollo de los embriones y se caracterizaron los huevos no viables, mediante microscopía de luz. Se comparó la proporción de huevos viables con embrión con la de huevos maduros viables. Resultados. La división celular comenzó 24 horas después de iniciado el cultivo. Los estadios tempranos estuvieron presentes por un periodo de tres a cinco días. Los estadios de desarrollo identificados fueron: huevos con una célula, con dos células, con tres células y con cuatro células;mórula temprana, mórula tardía, blástula, gástrula, renacuajo, prelarva, primer, segundo y tercer estado larvario. Se presentaron dos mudas larvarias. Los huevos no viables tenían el citoplasma degradado, cubierta exterior delgada o colapsada, y su larva no se movía al exponerla a la luz. No se encontraron diferencias significativas entre la proporción de huevos viables del día 5 al día 21, al compararla con la viabilidad de los huevos completamente maduros (30 días). Conclusión. Los embriones en desarrollo en el medio ambiente pueden considerarse como un riesgo potencial para la salud pública. La identificación precisa de los estadios de desarrollo y la clara diferenciación de huevos viables y no viables, pueden ayudar a determinar con exactitud una tasa basal de desarrollo, la cual sería útil en el estudio de compuestos ovicidas.

Embrioteca: espacio para la investigación científica y la docencia en la universidad médica villaclareña / Embrioteca: space for scientific research and teaching at the medical university of Villa Clara

Las embriotecas humanas son valiosas colecciones de material embrionario humano que honran las instituciones que las poseen. Entre las más citadas actualmente están prestigiosas embriotecas en España, Estados Unidos y Argentina. Resultan de un laborioso trabajo de recolección y procesamiento de delicadas muestras embrionarias con potencialidades de uso docente e investigativo. Han sido múltiples las aportaciones al conocimiento científico a partir del estudio de sus series, además del inestimable valor didáctico de las imágenes que atesoran. La Universidad de Ciencias Médicas de Villa Clara presenta la modesta colección que posee, abierta a nuevas contribuciones y a los interesados en su consulta. El objetivo de esta revisión es exponer su relevancia científica y docente en un contexto de revisión histórica y actualidad.

Human embryos are valuable collections of human embryonic material that honor the institutions that own them. Among the most cited are currently prestigious embryos in Spain, the United States and Argentina. They result from a laborious work of collection and processing of delicate embryonic samples with potential for teaching and research use. There have been many contributions to scientific knowledge from the study of their series, in addition to the invaluable didactic value of the images they treasure. The University of Medical Sciences of Villa Clara presents the modest collection that it has, open to new contributions and to those interested in its consultation. The objective of this review is to expose its scientific and teaching relevance in a context of historical review and current affairs.

Relationship between mitochondrial DNA copy number, membrane potential of human embryo and embryo morphology / 中华妇产科杂志

Objective To explore the relationship between the embryo with the different morphological types in the third day and its mitochondrial copy number,the membrane potential.Methods Totally 117 embryos with poor development after normal fertilization and were not suitable transferred in the fresh cycle and 106 frozen embryos that were discarded voluntarily by infertility patients with in vitro fertilization-embryo transfer after successful pregnancy were selected.According to evaluation of international standard in embryos,all cleavage stage embryos were divided into class Ⅰ frozen embryo group (n=64),class Ⅱ frozen embryo group (n=42) and class Ⅲ fresh embryonic group (not transplanted embryos;n=117).Real-time PCR and confocal microscopy methods were used to detect mitochondrial DNA (mtDNA) copy number and the mitochondrial membrane potential of a single embryo.The differences between embryo quality and mtDNA copy number and membrane potential of each group were compared.Results The copy number of mtDNA and the mitochondrial membrane potential in class Ⅲ fresh embryonic group [(1.7± 1.0)× 105 copy/μl,1.56±0.32] were significantly lower than those in class Ⅰ frozen embryo group [(3.4±1.7)×105 copy/μl,2.66±0.21]and class Ⅱ frozen embryo group [(2.6± 1.2)× 105 copy/μl,1.80±0.32;all P＜0.05].The copy number of mtDNA and the mitochondrial membrane potential in class Ⅰ frozen embryo group were significantly higher than those in class Ⅱ frozen embryo group (both P＜0.05).Conclusion The mtDNA copy number and the mitochondrial membrane potential of embryos of the better quality embryo are higher.

Preservação da fertilidade em mulheres com câncer: atualização e perspectivas / Fertility preservation in women with cancer: update and perspectives

Com o aumento do diagnóstico de câncer em mulheres jovens e os avanços no seu tratamento, muitas pacientes que poderão ter sua fertilidade comprometida com a quimioterapia têm manifestado desejo de engravidar futuramente. O congelamento de embriões, após fertilização in vitro, para preservar a fertilidade está bem estabelecido. A criopreservação de oócitos por vitrificação evoluiu bastante nos últimos anos, deixando de ser experimental. Até 2009 nasceram mais de 900 crianças a partir de oócitos criopreservados, sem aumento do risco de anomalias congênitas. O uso de análogos do GnRH para a supressão ovariana durante a quimioterapia na tentativa de prevenir a falência ovariana prematura apresenta resultados incertos. Outras técnicas ainda são consideradas experimentais, como a criopreservação e posterior autotransplante de tecido ovariano. Já foram relatados 24 nascimentos com o seu uso, persistindo, entretanto, dúvidas que motivam o seu estudo. A maturação de folículos ovarianos in vitro é alternativa promissora para preservação da fertilidade nessas pacientes e tem apresentado resultados positivos em roedores, macacos e humanos. Muita cautela deve ser tomada com o uso de técnicas experimentais, especialmente quando oferecidas para pacientes diante de fragilidade emocional. Por isso, é importante transmitir corretamente informações sobre chances de gravidez com tratamentos existentes e as limitações das técnicas experimentais...

With the increased number of cancer diagnoses among young women and the advances in treatment, many patients who may have had their fertility compromised by chemotherapy express desire to become pregnant in the future. Freezing embryos for later IVF so as to preserve fertility is a well established process. Oocyte cryopreservation by vitrification has evolved greatly in recent years and is no longer considered experimental. By 2009 more than 900 children were born from cryopreserved oocytes, without increased risk of congenital anomalies. The preventive use of GnRH analogues for ovarian suppression during chemotherapy to avoid premature ovarian failure has uncertain outcomes. Other techniques such as cryopreservation of ovarian tissue for later autograft are still considered experimental. Although use has already been reported in 24 births, doubts still persist and motivate further study. In vitro maturation of ovarian follicles is a promising alternative for preserving patient fertility and has shown positive results in rodents, monkeys, andhumans. Caution should be used with experimental techniques, especially when offered to emotionally fragile patients. Therefore it is important to thoroughly convey information on the chances of pregnancy with existing treatments and the limitations of experimental techniques...

Prenatal ultrasound of fetus with echogenic bowel and complicated structural abnormalities/chromosome abnormality / 中华医学超声杂志（电子版）

Objective To explore the prenatal ultrasonographic features of fetuses diagnosed as echogenic bowel and chromosomal abnormalities. Methods From September 2009 to June 2013, eighty cases diagnosed as echogenic bowel by prenatal ultrasound screening in our hospitals underwent chromosomal karyotype analysis and were followed up till fetal birth. The prenatal ultrasonographic features were carefully correlated with the postnatal findings. Results Prenatal ultrasoundand chromosomal findings of the 80 cases were:(1) Fifty-eight cases of fetuses with echogenic bowel alone (72.5%, 58/80, without chromosomal karyotype abnormalities). (2) Eleven cases of fetuses with additional ultrasound soft markers (13.8%, 11/80, without chromosomal karyotype abnormalities). The ultrasound examination revealed 8 cases of choroid plexus cysts, 1 case of single umbilical artery, 1 case of shorter nasal bone, and 1 case of echogenic intracardiac focus. (3) Five cases of fetuses with severe functional abnormalities or complex structural malformations and normal chromosomal karyotype. The ultrasound examination revealed 3 cases of structural cardiac malformations, 1 case of heart failure and 2 cases of central nervous system malformations. Of them,one case showed complex malformations (right ventricular dysplasia syndrome, cerebellar dysplasia and single umbilical artery), whereas the remaining 4 cases only involve single malformation. (4)Six cases had chromosomal abnormalities (7.5%, 6/80), including 3 cases of abnormal chromosome microdeletions, and 3 cases of abnormal chromosome numbers (1 21-trisomy syndrome, 1 18-trisomy syndrome, and 1 triploid syndrome). All of them had severe structural malformations and abnormal soft markers. The ultrasound examination revealed 2 cases of severe cardiac malformations, 2 cases of central nervous system malformations, 1 case of pericardial/peritoneal effusion, and 1 case of placental chorionic multiple hemangioma. One case of holoprosencephaly (donor of the twins) ended with intrauterine death. The associated abnormal soft markers include nuchal fold (NF) thickening, choroid plexus cysts, umbilical cord cysts, single umbilical artery, and placental chorionic hemangioma. Conclusions The fetuses with echogenic bowelaloneusualyhaveagoodprognosis.Insomecases,echogenicbowelmaydisappearduringvfolow-up.The fetuses complicated with severe structural malformations and chromosomal abnormalities have poor prognosis. For the fetuses diagnosed of echogenic bowel by prenatal ultrasound screening, systematic ultrasound examination should be carried out and clinical chromosomal karyotype analysis was suggested.

E16.5 embryonic mouse pancreatic tissue transplantation for the treatment of experimental diabetes / 中华普通外科杂志

Objective To investigate the effect of early embryonic mouse pancreatic tissue transplantation in the treatment of mouse experimental diabetes.Methods C57BL/6 mouse models of streptozocin-induced diabetes mellitus were established and then randomly divided into two groups:transplantation group,in which,five to seven pieces of pancreatic tissue of mice at embryonic 16.5 days were transplanted into mouse renal capsule,and sham-operated (SO) control group,in which,0.05 ml RPMI1640 culture medium was injected into mouse renal capsule.When blood glucose level of the transplantation group mouse was ≤ 11.2 mmol/L,the endocrine function of embryonic pancreatic tissue transplanted was detected by IPGTT and IPITT methods and then the transplanted graft was removed for observing the blood glucose surge.In vitro,levels of insulin secretion were measured in serum,E16.5 pancreatic tissue and graft (3.3 mmol/L and 16.7 mmol/L) glucose stimulation by ELISA.Histology and immunohistochemistry were observed before or after pancreatic tissue transplantation to detect insulin and glycagon production.Results (1) The glucose level decreased significantly in the TX group after 4-6 weeks of transplantation (13.4 ± 6.5 vs 28.9 ± 2.5,P ＜ 0.05),and the body weight gaining (P ＜ 0.05).Compared to SO group,glucose level decreased significantly (P ＜ 0.001).The graft can modulate insulin secretion; (2) Insulin can be detected in E16.5 embryonic mouse.Insulin and glycagon were produced more in TX group than pretransplantated pancreatic tissue.Conclusions E16.5 embryonic mouse pancreatic tissue transplantation effectively controls serum glucose level and restore the nondiabetic pattern of weight gaining in diabetic mouse.

Odontogenic cell culture in PEGDA hydrogel scaffolds for use in tooth regeneration protocols

In order to obtain a tooth-like structure, embryonic oral ectoderm cells (EOE) and bone marrow-derived stem cells (BMSC) were stratified within a synthetic hydrogel matrix (PEGDA) and implanted in the ileal mesentery of adult male Lewis rats. Wholemount in situ hybridization was used to evaluate the expression of Pitx2, Shh and Wnt10a signals indicative of tooth initiation. In rats, expression of the three markers was present in the oral ectoderm starting at embryonic stage E12.5, which was therefore selected for cell harvesting. Embryos were obtained by controlled service of young female Lewis rats in which estrus was detected by impedance reading. At E12.5, pregnant rats were humanely euthanized and embryos were collected. The mandibular segment of the first branchial arch was dissected and the mesenchyme separated from the ectoderm by enzymatic digestion with pancreatin trypsin solution. BMSCs were collected by flushing the marrow of tibiae and femurs of adult Lewis rats with a-MEM and cultured in a-MEM in 25 cm2 flasks. Second passage BMSC's were recombined with competent oral ectoderm (E12.5-E13) stratifying them within a 3D PEGDA scaffold polymerized by exposure to UV (365nm) inside a pyramidal polypropylene mold. Constructs were incubated from 24 to 48 hrs in a-MEM and then implanted for four to six weeks in the mesentery of adult male (3- 6 month old) Lewis rats. 76 constructs were implanted (37 experimental, 27 negative controls and 12 positive controls). Upon maturation, constructs were harvested, fixed in buffered formalin, processed and stained with hematoxylin eosin (HE). Histological evaluation of the experimental and negative constructs showed that BMSCs underwent an apoptotic process due to lack of matrix interactions, known as anoikis, and were thus incapable of interacting with the competent ectoderm. In contrast, embryonic oral ectoderm was able to proliferate during the mesenteric implantation. In conclusion, PEGDA scaffolds are incompatible with BMSCs, therefore it is essential to continue the search for an ideal scaffold that allows proper tissue interactions.

Para lograr la formacion de una estructura similar a un diente se estratificaron celulas de ectodermo oral embrionario (EOE) con celulas troncales de medula osea (BMSC) dentro de una matriz de diacrilato de polietilenglicol (PEGDA) que se implanto en el mesenterio ileal de machos adultos de ratas Lewis. Mediante hibridizacion in situ de bloque completo se evaluo la expresion de tres genes iniciadores putativos de la formacion dental (Pitx2, Shh y Wnt10a), estableciendo que en ratas las senales iniciadoras de dentogenesis aparecen entre E12.5 y E13. El tejido ectodermico embrionario se obtuvo haciendo cruces controlados de hembras a las que se les detecto el estro mediante impedanciometria. En E12.5 las hembras se sacrificaron y se extrajeron los embriones. Se diseco la porcion mandibular del primer arco branquial y el ectodermo se separo del mesenquima mediante disociacion enzimatica con una solucion de pancreatina tripsina. Las BMSC se extrajeron de los huesos largos de las extremidades inferiores de ratas mediante lavado con a-MEM y se cultivaron en cajas de 25cm2 hasta un segundo pasaje. Las BMSC fueron recombinadas con ectodermo embrionario competente (E12.5 - E13) estratificandolas en un soporte tridimensional de PEGDA, polimerizado con luz ultravioleta (365nm) dentro de un molde piramidal de polipropileno (PP). Los constructos se cultivaron entre 48 y 72 horas en a-MEM y posteriormente fueron implantados en el mesenterio ileal de machos adultos (3 a 6 meses de edad) por un periodo de cuatro a seis semanas. Se implantaron 76 constructos (37 experimentales, 27 controles negativos y 12 controles positivos). En la fecha determinada los animales se sacrificaron mediante asfixia con una mezcla de CO2 y aire recuperando los constructos que se fijaron en formalina tamponada para luego procesarlos y tenirlos con hematoxilina eosina (HE). La evaluacion histologica de los constructos experimentales, positivos y negativos mostro que las BMSC incluidas en el hidrogel sufrieron un proceso de apoptosis conocido como anoikis que impidio su interaccion con las celulas ectodermicas. En contraste el EOE prolifero durante el periodo de implantacion. A futuro se debe buscar la matriz portadora ideal que permita el confinamiento de los dos grupos celulares y que brinde el soporte estructural necesario para la proliferacion de las BMSC facilitando su interaccion con las celulas inductoras de origen ectodermico.

Miocardiopatia não compactada: uma visão atual / Noncompaction cardiomyopathy: a current view / Miocardiopatía no compactada: una visión actual

A miocardiopatia não compactada isolada é uma doença rara, que provavelmente surge no período embrionário, com a parada intrauterina da compactação miocárdica no início do desenvolvimento fetal, e que determina trabeculações miocárdicas proeminentes com recessos intertrabeculares profundos e espessamento do miocárdio em duas camadas distintas (compactado e não compactado). Embora descrita inicialmente na população pediátrica ou em conjunto com cardiopatia congênita, pode-se compreender que essa doença ocorre de forma isolada, porque o diagnóstico é cada vez mais comum em pacientes adultos que não apresentam outra doença cardíaca. As manifestações clínicas são altamente variáveis, porque partem da ausência de sintomas à insuficiência cardíaca congestiva, arritmias e tromboembolismo sistêmico. A ecocardiografia é o procedimento diagnóstico mais utilizado, porém o pouco conhecimento sobre essa doença, sua semelhança com outras afecções miocárdicas e a limitação da técnica ecocardiográfica empregada fazem com que o diagnóstico seja postergado. Esta revisão objetiva mostrar que outras modalidades de imagem como ressonância magnética, tomografia computadorizada e ventriculografia esquerda são alternativas diagnósticas.

Isolated non-compaction cardiomyopathy is a rare disease that is likely to develop in the embryonic period. It is caused by the intrauterine arrest of the myocardial compaction process in the beginning of the fetal development. It is characterized by prominent myocardial trabeculations and deep intertrabecular recesses, as well as the thickening of the myocardium into two distinct layers (compacted and not compacted). Even though this disease is said to be prevalent in the pediatric population or together with congenital heart disease, one can understand that this disease occurs in isolation, because the diagnosis is becoming more common in adult patients that have no other heart disease. The clinical manifestations vary greatly, because they range from absence of symptoms to congestive heart failure, arrhythmias and systemic thromboembolism. Echocardiography is the most widely used diagnostic procedure, but the little knowledge about this disease, its similarity to other myocardial diseases and the limitation of the echocardiographic technique used delay the diagnosis. The purpose of this review is to show that that other imaging techniques, such as MRI, CT and left ventriculography have emerged as diagnostic alternatives.

La miocardiopatía no compactada aislada es una enfermedad rara, que probablemente surge en el período embrionario, con la parada intrauterina de la compactación miocárdica en el comienzo del desarrollo fetal, y que determina trabeculaciones miocárdicas prominentes con recesos intertrabeculares profundos y espesamiento del miocardio en dos capas distintas (compactado y no compactado). Aunque descripta inicialmente en la población pediátrica o en conjunto con cardiopatía congénita, se puede comprender que esa enfermedad ocurre de forma aislada, porque el diagnóstico es cada vez más común en pacientes adultos que no presentan otra enfermedad cardíaca. Las manifestaciones clínicas son altamente variables, porque parten de la ausencia de síntomas a la insuficiencia cardíaca congestiva, arritmias y tromboembolismo sistémico. La ecocardiografía es el procedimiento diagnóstico más utilizado, sin embargo el poco conocimiento sobre esa enfermedad, su semejanza con otras afecciones miocárdicas y la limitación de la técnica ecocardiográfica empleada hacen que el diagnóstico sea postergado. Esta revisión pretende mostrar que otras modalidades de imagen como resonancia magnética, tomografía computarizada y ventriculografía izquierda son alternativas diagnósticas.

Desenvolvimento embrionário em ratas tratadas com tacrolimus durante a fase de pré-implantação / Embryo development in rats treated with tacrolimus during the preimplantation phase

OBJETIVO: avaliar a toxicidade do tacrolimus sobre o desenvolvimento embrionário em ratas tratadas durante o período de trânsito tubário. MÉTODOS: sessenta ratas Wistar foram distribuídas em quatro grupos (15 animais cada), que receberam diferentes doses de tacrolimus por via intragástrica: (T1) 1,0 mg/kg/dia, (T2) 2,0 mg/kg/dia e (T3) 4,0 mg/kg/dia. O grupo controle (C) recebeu água destilada. As ratas foram observadas diariamente para detectar sinais clínicos de toxicidade. O tratamento foi realizado do primeiro ao quinto dia de gestação. As seguintes variáveis maternas foram analisadas: peso corporal, de ovários, fígados e rins, consumo de alimento, número de corpos lúteos, implantes, fetos vivos e mortos e índice de implantação. Os fetos e placentas foram pesados e os primeiros foram observados para detectar malformações externas. Estatística: análise de variância (ANOVA), uma via, seguida de teste de Dunnett (alfa=0,05). RESULTADOS: não ocorreram indícios clínicos de toxicidade materna, tais como perda de peso, redução do consumo de alimento ou do peso de órgãos (p>0,05). Também não houve diferença significativa no peso corporal dos fetos (C: 1,8±0,6; T1: 2,2±0,5; T2: 1,9±0,5 e T3: 2,0±0,5 g) e de placentas (C: 1,6±0,4; T1: 1,5±0,4; T2: 1,8±0,4 e T3: 1,6±0,4 g), com p>0,05. Nenhuma malformação externa foi detectada. CONCLUSÕES: a administração de tacrolimus a ratas prenhes durante o período de trânsito tubário não parece ter qualquer efeito tóxico e materno ou embrionário.

PURPOSE: to evaluate the toxicity of tacrolimus on embryonic development in rats treated during the tubal transit period. METHODS: sixty Wistar rats were distributed into four groups (15 animals each), which received different doses of tacrolimus through intragastric administration: (T1) 1.0 mg/kg/day, (T2) 2.0 mg/kg/day and (T3) 4.0 mg/kg/day. The control group (C) received distilled water. The rats were observed daily to detect clinical signs of toxicity. The treatments were performed from the first to the fifth day of pregnancy. The following maternal variables were analyzed: body, ovary, liver, and kidney weights, food intake, number of corpora lutea, implants, alive and dead fetuses, and implantation rates. The fetuses and placentae were weighed and the former were observed in order to detect external malformation. Statistical analysis was performed by one way: analysis of variance (ANOVA), folowed by the Dunnet test (alpha=0.05). RESULTS: there were no signs of maternal toxicity, such as body weight loss, decrease in food intake or in organ weights (p>0.05). There was also no significant difference among weights of fetuses (C: 1.8±0.6; T1: 2.2±0.5; T2: 1.9±0.5 and T3: 2.0±0.5 g) and placentae (C: 1,6±0.4; T1: 1.5±0.4; T2: 1.8±0.4 e T3: 1.6±0.4 g), with p>0.05; no external malformation was detected in the fetuses. CONCLUSIONS: the administration of tacrolimus to pregnant rats during the tubal transit period does not seem to generate any toxic effect to mother or embryo.

Volume do embrião estimado pela ultra-sonografia tridimensional entre a sétima e a décima semana de gestação / Embryo volume estimated by three-dimensional ultrasonography at seven to ten weeks of pregnancy

OBJETIVO: avaliar a evolução do volume do embrião (VE) entre a sétima e a décima semana de gestação por meio da ultra-sonografia tridimensional. MÉTODOS: realizou-se um estudo de corte transversal com 63 gestantes normais entre a sétima e a décima semana. Os exames ultra-sonográficos foram realizados por meio de um transdutor endocavitário volumétrico. Para o cálculo do VE, utilizou-se o método VOCAL (Virtual Organ Computer-aided Analysis) com ângulo de rotação de 12º, com delimitação de 15 planos seqüenciais. Para o VE foram calculadas médias, medianas, desvios padrão e valores máximo e mínimo em todas as idades gestacionais. Para se avaliar a correlação entre o VE e o comprimento cabeça-nádega (CCN) foi criado gráfico de dispersão, sendo o ajuste realizado pelo coeficiente de determinação (R²). Para se determinarem intervalos de referência do VE em função do CCN, utilizou-se a seguinte fórmula: percentil =VE+K versus dp, com K=1,96. RESULTADOS: o CCN variou de 9,0 a 39,7 mm, com média de 23,9 mm (±7,9 mm), enquanto o VE variou de 0,1 a 7,6 cm³, com média de 2,7 cm³ (±3,2 cm³). O VE foi altamente correlacionado com o CCN, sendo que o melhor ajuste foi obtido com regressão quadrática (VE=0,165 - 0,055 x CCN + 0,005 x CCN²; R²=0,853). O VE médio variou de 0,1 (-0,3 a 0,5 cm³) a 6,7 cm³ (3,8 a 9,7 cm³) no intervalo de 9 a 40 mm do CCN. Neste intervalo o VE aumentou 67 vezes, enquanto o CCN aumentou apenas 4,4 vezes. CONCLUSÕES: o VE é um parâmetro mais sensível que o CCN para avaliar o crescimento embrionário entre a sétima e a décima semana de gestação.

PURPOSE: to evaluate the embryo's volume (EV) between the seventh and the tenth gestational week, through tridimensional ultrasonography. METHODS: a transversal study with 63 normal pregnant women between the seventh and the tenth gestational week. The ultrasonographical exams have been performed with a volumetric abdominal transducer. Virtual Organ Computer-aided Analysis (VOCAL) has been used to calculate EV, with a rotation angle of 12º and a delimitation of 15 sequential slides. The average, median, standard deviation and maximum and minimum values have been calculated for the EV in all the gestational ages. A dispersion graphic has been drawn to assess the correlation between EV and the craniogluteal length (CGL), the adjustment being done by the determination coefficient (R²). To determine EV's reference intervals as a function of the CGL, the following formula was used: percentile=EV+K versus SD, with K=1.96. RESULTS: CGL has varied from 9.0 to 39.7 mm, with an average of 23.9 mm (±7.9 mm), while EV has varied from 0.1 to 7.6 cm³, with an average of 2.7 cm³ (±3.2 cm³). EV was highly correlated to CGL, the best adjustment being obtained with quadratic regression (EV=0.2-0.055 versus CGL+0.005 versus CGL²; R²=0.8). The average EV has varied from 0.1 (-0.3 to 0.5 cm³) to 6.7 cm³ (3.8 to 9.7 cm³) within the interval of 9 to 40 mm of CGL. EV has increased 67 times in this interval, while CGL, only 4.4 times. CONCLUSIONS: EV is a more sensitive parameter than CGL to evaluate embryo growth between the seventh and the tenth week of gestation.