Exposure to fine particulate matter in the air alters placental structure and the renin-angiotensin system.

METHODS: Female Wistar rats were exposed to filtered air (F) or to concentrated fine particulate matter (P) for 15 days. After mating, the rats were divided into four groups and again exposed to F or P (FF, FP, PF, PP) beginning on day 6 of pregnancy. At embryonic day 19, the placenta was collected. The placental structure, the protein and gene expression of TGFß1, VEGF-A, and its receptor Flk-1 and RAS were evaluated by indirect ELISA and quantitative real-time PCR. RESULTS: Exposure to P decreased the placental mass, size, and surface area as well as the TGFß1, VEGF-A and Flk-1 content. In the maternal portion of the placenta, angiotensin II (AngII) and its receptors AT1 (AT1R) and AT2 (AT2R) were decreased in the PF and PP groups. In the fetal portion of the placenta, AngII in the FP, PF and PP groups and AT2R in the PF and PP groups were decreased, but AT1R was increased in the FP group. VEGF-A gene expression was lower in the PP group than in the FF group. CONCLUSIONS: Exposure to pollutants before and/or during pregnancy alters some characteristics of the placenta, indicating a possible impairment of trophoblast invasion and placental angiogenesis with possible consequences for the maternal-fetal interaction, such as a limitation of fetal nutrition and growth.

Isolated total RNA and protein are preserved after thawing for more than twenty-four hours.

OBJECTIVE: The preservation of biological samples at a low temperature is important for later biochemical and/or histological analyses. However, the molecular viability of thawed samples has not been studied sufficiently in depth. The present study was undertaken to evaluate the viability of intact tissues, tissue homogenates, and isolated total RNA after defrosting for more than twenty-four hours. METHODS: The molecular viability of the thawed samples (n = 82) was assessed using the A260/A280 ratio, the RNA concentration, the RNA integrity, the level of intact mRNA determined by reverse transcriptase polymerase chain reaction, the protein level determined by Western blotting, and an examination of the histological structure. RESULTS: The integrity of the total RNA was not preserved in the thawed intact tissue, but the RNA integrity and level of mRNA were perfectly preserved in isolated defrosted samples of total RNA. Additionally, the level of ß-actin protein was preserved in both thawed intact tissue and homogenates. CONCLUSION: Isolated total RNA does not undergo degradation due to thawing for at least 24 hours, and it is recommended to isolate the total RNA as soon as possible after tissue collection. Moreover, the protein level is preserved in defrosted tissues.

Isolated total RNA and protein are preserved after thawing for more than twenty-four hours

OBJECTIVE: The preservation of biological samples at a low temperature is important for later biochemical and/or histological analyses. However, the molecular viability of thawed samples has not been studied sufficiently in depth. The present study was undertaken to evaluate the viability of intact tissues, tissue homogenates, and isolated total RNA after defrosting for more than twenty-four hours. METHODS: The molecular viability of the thawed samples (n = 82) was assessed using the A260/A280 ratio, the RNA concentration, the RNA integrity, the level of intact mRNA determined by reverse transcriptase polymerase chain reaction, the protein level determined by Western blotting, and an examination of the histological structure. RESULTS: The integrity of the total RNA was not preserved in the thawed intact tissue, but the RNA integrity and level of mRNA were perfectly preserved in isolated defrosted samples of total RNA. Additionally, the level of β-actin protein was preserved in both thawed intact tissue and homogenates. CONCLUSION: Isolated total RNA does not undergo degradation due to thawing for at least 24 hours, and it is recommended to isolate the total RNA as soon as possible after tissue collection. Moreover, the protein level is preserved in defrosted tissues.

Expressão de mediadores citotóxicos (perforina, granzima B, FAS e FAS-l) em biópsias de aloenxerto renal / Expression of cytotoxic mediators (perforin, granzyme B, FAS,and FAS-l) in renal allograft biopsies

O objetivo deste trabalho foi analisar por imunoistoquímica a expressão in situ de perforina, granzima B, FAS-L e FAS em biópsias de enxerto renal,correlacionando esses achados com o grau histológico de rejeição eprognóstico do enxerto. Biópsias renais (n = 96) foram divididas em três grupos: rejeição aguda (n = 56), rejeição crônica (n = 31), função renal estável (sem rejeição; n = 9). A expressão de perforina, granzima B, FAS-L e FAS foi avaliada por imunoistoquímica. A expressão de perforina e granzima B foi significativamente maior na rejeição aguda (4,84 ± 0,65 e 30,05 ±7,93) quando comparada à rejeição crônica (0,71 ± 0,13 céls./mm2e 11,40 ± 3,84; p < 0,001 e p< 0,05 respectivamente), mas não deFAS-L (24,44 ± 5,56 na rejeição aguda vs. 18,87 ± 6,83 na rejeição crônica). As marcações de perforina, granzima B e FAS-L na rejeição aguda foram significativamente maiores na rejeição aguda que nos casos sem rejeição e controle. A expressão de FAS foi semelhante entre os grupos. Houve modesta correlação entre a expressão de perforina e a gravidade da rejeição aguda (r = 0,28, p = 0,05). A perforina foi o marcador mais confiável para o diagnóstico de rejeição aguda, com 80% de sensibilidade e 84,4% de especificidade. A presença in situ de perforina, granzima B e FAS-L embiópsias de aloenxerto renal na rejeição aguda indica a ocorrênciade intensa atividade citotóxica. Estes dados apontam para a possibilidade de utilização destas moléculas como marcadores de rejeição, podendo vir a se tornar um instrumento na monitorização pós-transplante.