Immunohistochemistry section staining is not always easy to interpret. Manual quantification of immunohistochemical staining is limited by the observer visual ability to detect changes in level staining. Hence, the quantification of immunostaining by means of digital image analysis allows us to measure accurately protein expression percentages in immunobiological stained tissues and ensures to overcome the visual limitations. We perform an experimental study to analyse the impact of folic acid (FA) deficiency into collagen IV and laminin-1 expression in the embryonic mouse lens. The study starts with microscope images of embryos mouse lens whose mothers fed a diet deficient in FA during 2 and 8 weeks. A principal component analysis (PCA) image processing is used to analyse these images coming from control and FA deficit groups. The method permits to define an index of over- or infraexpression of collagen IV and laminin-1 associated to different spatial organisation structures (PC processes). Additionally, it permits to determine in precise percentage the exact quantity of the overexpression or infraexpression and finally to comprehend molecular regionalisation and expression in both control and deficient groups. The results suggest that even with 2 weeks of deficit of FA the expression and distribution of both molecules is affected.
Purpose: Folic Acid (FA) is an essential vitamin for embryonic development. It plays particularly a critical role in RNA, DNA, and protein synthesis. On the other hand, the collagen IV and laminin-1 are important proteins during embryonic development. This study was done to find if FA-deficiency at a short and a long-term in mothers could alter the tissue texture of retina and lens of the progeny. Methods: Collagen IV and laminin-1 were localized by immunohistochemistry in the lens and retina of the FA-deficient embryos. To carry out the image processing, texture segmentation was performed through canny edge detection and Fourier transform (FT). We defined a parameter, the grain size, to describe the texture of the lens and retina. A bootstrap method to estimate the distribution and confidence intervals of the mean, standard deviation, skewness and kurtosis of the grain size has been developed. Results: Analysis through image processing using Matlab showed changes in the grain size between control and FA deficient groups in both studied molecules. Measures of texture based on FT exhibited changes in the directionality and arrangements of type IV collagen and laminin-1. Conclusions: Changes introduced by FA deficiency were visible in the short-term (two weeks) and evident in the long term (eight weeks) in both grain size and orientation of fibre structures in the tissues analysed (lens and retina). This is the first work devoted to study the effect of FA deficit in the texture of eye tissues using image processing techniques.
Purpose: Folic acid (FA) is an essential nutrient for normal embryonic development. FA deficiency (FAD) in maternal diet increases the risk of several defects among the progeny, especially, neural tube defects. The eye begins its development from the neural tube; however, the relationship between FAD and ocular development in the offspring has been little explored and it isn't known how the FAD affects the formation of the eye. Our objective was to analyze the effect of maternal FAD on mouse embryos ocular biometry. Methods: Female mice C57/BL/6J were distributed into three different groups, according to the assigned diet: control group fed a standard FA diet (2 mg FA/kg), FAD group for short term fed (0 mg FA/kg + 1% succinylsulfathiazole) from the day after mating until day 14.5 of gestation, and FAD group for long term fed the same FA-deficient diet for 6 weeks prior mating and continued with this diet during gestation. A total of 57 embryos (19 embryos of each dietary group) at 14.5 gestational days were evaluated. As indicators of changes in ocular biometry, we analyze two parameters: area and circularity of the lens and whole eye, and the area of the retina. The program used in the treatment and selection of the areas of interest was ImageJ. The statistical analysis was performed by IBM SPSS Statistics 19. Results: Regarding the measures of the area, FA-deficient lenses and eyes were smaller than that of controls. We have also observed increase in the size of the neural retina, spatially, in embryos from females fed FAD diet during long term. On the other hand, as regard to circularity measures, we have seen that eyes and lenses were more circular than control. Conclusion: Maternal FAD diet for a very short term generates morphological changes in ocular structures to the offspring.
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