Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques

Noemy Sandoval-Skeet,1 Jason A Kaufman,2 Monica J Castro,2 Layla Al-Nakkash3 1Department of Biomedical Sciences, 2Department of Anatomy, 3Department of Physiology, Midwestern University, Glendale, AZ 85308, USA Introduction: Diabetes is commonly associated with gastrointestinal dysfunction. We have...

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Autores principales:	Sandoval-Skeet N, Kaufman JA, Castro MJ, Al-Nakkash L
Formato:	article
Lenguaje:	EN
Publicado:	Dove Medical Press 2018
Materias:	Genistein intestine epithelia jejunum optical clearing diabetes Specialties of internal medicine RC581-951
Acceso en línea:	https://doaj.org/article/24c7a25aab6840e7979d651b0760aacc
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spelling	oai:doaj.org-article:24c7a25aab6840e7979d651b0760aacc2021-12-02T07:10:46ZGenistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques1178-7007https://doaj.org/article/24c7a25aab6840e7979d651b0760aacc2018-11-01T00:00:00Zhttps://www.dovepress.com/genistein-diet-does-not-modify-crypt-morphology-in-the-obob-mouse-jeju-peer-reviewed-article-DMSOhttps://doaj.org/toc/1178-7007Noemy Sandoval-Skeet,1 Jason A Kaufman,2 Monica J Castro,2 Layla Al-Nakkash3 1Department of Biomedical Sciences, 2Department of Anatomy, 3Department of Physiology, Midwestern University, Glendale, AZ 85308, USA Introduction: Diabetes is commonly associated with gastrointestinal dysfunction. We have previously shown that transepithelial short circuit current, Isc (chloride secretion), is significantly reduced in the jejunum from ob/ob mice vs lean controls, and consumption of 600 mg genistein/kg of diet (600 G) for 4 weeks significantly rescues Isc. We aimed to evaluate whether morphological changes in the jejunal crypts contribute to the rescue of Isc. Methods: Male mice (ob/ob and lean controls) were fed either a genistein-free diet or genistein-containing diet (600 G). Comparisons of crypt morphology were made for crypt depth, length, and numbers of proliferative cells. Assessments of crypt measures using DAPI and 5-ethynyl-2′-deoxyuridine (EdU) were performed using traditional cryostat sectioning and an innovative 3D optical clearing method. Results: We found that crypt length in the ob/ob genistein-fed group was significantly greater when measured with cleared tissue (85.19±4.73 µm, P<0.05, n=8) compared to lengths measured with cryostat (65.42±3.48 µm, n=8). In addition, proliferative EdU+ counts were approximately fivefold greater with clearing, compared to counts obtained via single plane images from cryostat sections for all groups measured. The average length to EdU+ ratio was unchanged between groups. Conclusion: Thus, we conclude that genistein diet does not affect overall cellular proliferation or crypt morphology, other than for the modest increased crypt length measured via clearing in the ob/ob genistein group. The increase in crypt length is likely indicative of the greater accuracy of the 3D measures compared to single plane. Genistein diet-induced increases in the intestinal Isc are therefore likely not attributed to changes in intestinal crypt morphology. Keywords: genistein, intestine epithelia, jejunum, optical clearing, diabetesSandoval-Skeet NKaufman JACastro MJAl-Nakkash LDove Medical PressarticleGenisteinintestine epitheliajejunumoptical clearingdiabetesSpecialties of internal medicineRC581-951ENDiabetes, Metabolic Syndrome and Obesity: Targets and Therapy, Vol Volume 11, Pp 863-873 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Genistein intestine epithelia jejunum optical clearing diabetes Specialties of internal medicine RC581-951
spellingShingle	Genistein intestine epithelia jejunum optical clearing diabetes Specialties of internal medicine RC581-951 Sandoval-Skeet N Kaufman JA Castro MJ Al-Nakkash L Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
description	Noemy Sandoval-Skeet,1 Jason A Kaufman,2 Monica J Castro,2 Layla Al-Nakkash3 1Department of Biomedical Sciences, 2Department of Anatomy, 3Department of Physiology, Midwestern University, Glendale, AZ 85308, USA Introduction: Diabetes is commonly associated with gastrointestinal dysfunction. We have previously shown that transepithelial short circuit current, Isc (chloride secretion), is significantly reduced in the jejunum from ob/ob mice vs lean controls, and consumption of 600 mg genistein/kg of diet (600 G) for 4 weeks significantly rescues Isc. We aimed to evaluate whether morphological changes in the jejunal crypts contribute to the rescue of Isc. Methods: Male mice (ob/ob and lean controls) were fed either a genistein-free diet or genistein-containing diet (600 G). Comparisons of crypt morphology were made for crypt depth, length, and numbers of proliferative cells. Assessments of crypt measures using DAPI and 5-ethynyl-2′-deoxyuridine (EdU) were performed using traditional cryostat sectioning and an innovative 3D optical clearing method. Results: We found that crypt length in the ob/ob genistein-fed group was significantly greater when measured with cleared tissue (85.19±4.73 µm, P<0.05, n=8) compared to lengths measured with cryostat (65.42±3.48 µm, n=8). In addition, proliferative EdU+ counts were approximately fivefold greater with clearing, compared to counts obtained via single plane images from cryostat sections for all groups measured. The average length to EdU+ ratio was unchanged between groups. Conclusion: Thus, we conclude that genistein diet does not affect overall cellular proliferation or crypt morphology, other than for the modest increased crypt length measured via clearing in the ob/ob genistein group. The increase in crypt length is likely indicative of the greater accuracy of the 3D measures compared to single plane. Genistein diet-induced increases in the intestinal Isc are therefore likely not attributed to changes in intestinal crypt morphology. Keywords: genistein, intestine epithelia, jejunum, optical clearing, diabetes
format	article
author	Sandoval-Skeet N Kaufman JA Castro MJ Al-Nakkash L
author_facet	Sandoval-Skeet N Kaufman JA Castro MJ Al-Nakkash L
author_sort	Sandoval-Skeet N
title	Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
title_short	Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
title_full	Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
title_fullStr	Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
title_full_unstemmed	Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
title_sort	genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques
publisher	Dove Medical Press
publishDate	2018
url	https://doaj.org/article/24c7a25aab6840e7979d651b0760aacc
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Genistein diet does not modify crypt morphology in the ob/ob mouse jejunum: a comparison of cryostat and clearing techniques

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