ABSTRACT
Overexpression of Short and Raji variants of Cellular FLICE-like inhibitory protein (c-FLIP) is capable of inhibiting apoptosis, while the function of the Long isoform depends of c-FLIPL concentration in cells. The aim of this study was to determine the effects of c-FLIPL knockdown in cervical cell lines. SiHa, C-4I and C-33A cervical cancer cell lines were analyzed. c-FLIPL level expression was determined by quantitative real-time PCR and western blotting. c-FLIPL was transiently downregulated by siRNA. The effects of knockdown of c-FLIPL on cell viability, proliferation and apoptosis were assessed by comparing with scrambled siRNA-transfected cells. SiHa and C-4I c-FLIPL knockdown cells showed increased viability compared with scrambled siRNA-transfected cells (P<0.05), while C-33A cells did not show significant differences. Ki-67 and PCNA immunocytochemistry was performed to evaluate proliferation on these cervical cancer cell lines. SiHa cells with c-FLIPL knockdown showed elevated expression of Ki-67 protein compared with their scrambled counterparts (P<0.0001), while C-33A c-FLIPL knockdown cells showed a significantly lower in PCNA expression (P<0.01) compared with control. All three c-FLIP-transfected cell lines showed a higher level of apoptosis compared with their scrambled controls. Our results suggest that c-FLIPL could have effects in proliferation and apoptosis in cervical cancer cell lines.
Cuando las variantes Short y Raji de la proteína Cellular FLICE-like inhibitory protein (c-FLIP) se encuentran sobrexpresadas son capaces de inhibir la apoptosis, mientras la función de la isoforma Long (c-FLIPL), depende de la concentración de esta molécula en las células. El objetivo de este estudio fue determinar los efectos de la inhibición de c-FLIPL en líneas celulares de cáncer de cuello uterino. Para realizar el estudio fueron utilizadas SiHa, C-4I y C-33A, líneas celulares de cáncer cervical. La expresión de c-FLIPL en estas líneas fue establecida mediante PCR en tiempo real y western blot. Posteriormente la expresión de c-FLIPL fue inhibida, mediante transfeción transiente con siRNA complementario al mRNA mensajero de c.-FLIPL. Los efectos de esta inhibición en la viabilidad celular, proliferación y apoptosis fue comparada con células transfectadas con un siRNA control (scrambled). Una vez reprimido c-FLIPL, las líneas celulares SiHa y C-4I presentaron un aumento de la viabilidad celular (P<0,05). Para evaluar la proliferación celular se utilizó inmunocitoquímica de los marcadores Ki-67 y PCNA. Las células SiHa transfectadas con siRNA c-FLIPL, mostraron una elevada expresión de Ki-67 (P<0,0001), mientras que las células C-33A con c-FLIPL inhibido mostraron una menor expresión de PCNA (P<0,01). Las tres líneas celulares con c-FLIPL reprimido mostraron un mayor nivel de apoptosis que las células control. Estos resultados sugieren que c-FLIPL puede tener efectos en la proliferación y apoptosis de líneas celulares de cáncer de cuello uterino.
Subject(s)
Humans , Female , Uterine Cervical Neoplasms/metabolism , Uterine Cervical Neoplasms/pathology , Cell Line, Tumor , CASP8 and FADD-Like Apoptosis Regulating Protein/metabolism , Immunohistochemistry , Cell Survival , Apoptosis , In Situ Nick-End Labeling , RNA, Small Interfering , Cell Proliferation , Real-Time Polymerase Chain ReactionABSTRACT
The aim of this work was the description of fetal mummification in silky shark. During July of 2001 in Santa Maria fishing camp, Gulf of California Mexico, fourteen pregnant female of silky sharks were captured. In all of them the embryos were found to be dehydrated. However, total observations were possible in only two litters. Six embryos in each analyzed litters were found. All the embryos were mummified, complete and without maceration. Because the litters were totally mummificated in several females, it was suggested that fetal mummification was produced by the environmental condition.
A mumificação fetal é caracterizada pela desidatração de embriões mortos dentro do útero da mãe. A perda de embriões pode afetar a sustentabilidade da população explorada, tornando o estudo deste fenômeno de suma importância. Em Julho de 2001 quatorze femeas grávidas do tubarão lombo preto foram capituradas durante uma amostragem no campo pesqueiro de Santa Maria, Golfo de California, México. Todas as fêmeas capturadas apresentaram embriões mumificados. Infelizmente, observações detalhadas dos embriões foram possíveis em apenas duas fêmeas. Cada fêmea analizada apresentava seis embriões, todos completamente mumificados e sem maceração. Embriões completamente mumificados foram encontrados em muitas fêmeas, o que sugere que a mumificação fetal desses embriões pode ter sido causada por condições ambientais.