Characterization of Caco-2 and HT29-MTX cocultures in an in vitro digestion/cell culture model used to predict iron bioavailability☆

Mahler, Gretchen J.; Shuler, Michael L.; Glahn, Raymond P.

doi:10.1016/j.jnutbio.2008.05.006

Cited by 253 publications

(194 citation statements)

References 35 publications

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“…The 3D architecture of the silk scaffold system with a geometrically engineered hollow lumen allows seeding of Caco-2 and HT29-MTX cells circumferentially on the surface of the lumen and recapitulates the structure of the human small intestine. Two types of intestinal epithelial cells which are routinely used in coculture for intestinal model systems (30), cells of the Caco-2 and the HT29-MTX cell lines, which are differentiated for 2 weeks in situ, are included, resulting in enterocyte-like and goblet cell-like cells, respectively. These cells grow as highly polarized monolayers characterized by microvilli with continuous brush borders and the formation of tight junctions, identified by zonula occludens protein 1 (ZO-1) immunostaining (26), in the scaffold lumen.…”

Section: Discussionmentioning

confidence: 99%

Novel Bioengineered Three-Dimensional Human Intestinal Model for Long-Term Infection of Cryptosporidium parvum

et al. 2017

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Cryptosporidium spp. are apicomplexan parasites of global importance that cause human diarrheal disease. In vitro culture models that may be used to study this parasite and that have physiological relevance to in vivo infection remain suboptimal. Thus, the pathogenesis of cryptosporidiosis remains poorly characterized, and interventions for the disease are limited. In this study, we evaluated the potential of a novel bioengineered three-dimensional (3D) human intestinal tissue model (which we developed previously) to support long-term infection by Cryptosporidium parvum. Infection was assessed by immunofluorescence assays and confocal and scanning electron microscopy and quantified by quantitative reverse transcription-PCR. We found that C. parvum infected and developed in this tissue model for at least 17 days, the extent of the study time used in the present study. Contents from infected scaffolds could be transferred to fresh scaffolds to establish new infections for at least three rounds. Asexual and sexual stages and the formation of new oocysts were observed during the course of infection. Additionally, we observed ablation, blunting, or distortion of microvilli in infected epithelial cells. Ultimately, a 3D model system capable of supporting continuous Cryptosporidium infection will be a useful tool for the study of hostparasite interactions, identification of putative drug targets, screening of potential interventions, and propagation of genetically modified parasites. KEYWORDS 3D model, Cryptosporidium, in vitro culture, intestinal epithelial cells C ryptosporidium spp. are apicomplexan parasites of global importance that cause diarrheal disease in humans and animals worldwide (1, 2). Though immunocompetent individuals experience self-limiting or asymptomatic infection, immunocompromised hosts, such as untreated patients with HIV infection or AIDS (3) and malnourished children (1, 2) in resource-limited settings, may experience severe diarrhea, wasting, and death. In a recent case-control study, Cryptosporidium was one of four pathogens responsible for moderate to severe diarrhea in children under the age of 5 years and was associated with an increased risk of death in children from 1 to 2 years of age in seven countries in sub-Saharan Africa and South Asia (4). The only FDA-approved drug for the treatment of cryptosporidiosis is nitazoxanide, a drug with broad antiparasitic properties (5). However, this drug is not effective in immunocompromised patients (6) and has not been widely tested in malnourished children in resource-limited countries. Though significant improvements in water purification methods have occurred (7) since the outbreak of waterborne cryptosporidiosis in Milwaukee, WI, in 1993 (8), industrialized countries are seeing increased numbers of cases, largely due to recreational water outbreaks (9).Unlike many other apicomplexan parasites which often require more than one host to complete their life cycles (10), Cryptosporidium completes its entire life cycle in a

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Section: Discussionmentioning

confidence: 99%

Novel Bioengineered Three-Dimensional Human Intestinal Model for Long-Term Infection of Cryptosporidium parvum

et al. 2017

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“…A few months later, coculture of cells that model enterocytes with the ones that model goblet (mucinsecreting) cells was described [27]. Such cocultures appear to predict iron bioavailability for foods [34] and look like a system where new insights can develop. Perhaps they can also encourage a new level of insight into how phytates, polyphenols, and other inhibitors render iron-rich plant foods like spinach into poor sources of iron.…”

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confidence: 99%

Human iron transporters

Garrick

2010

Genes Nutr

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Human iron transporters manage iron carefully because tissues need iron for critical functions, but too much iron increases the risk of reactive oxygen species. Iron acquisition occurs in the duodenum via divalent metal transporter (DMT1) and ferroportin. Iron trafficking depends largely on the transferrin cycle. Nevertheless, nondigestive tissues have a variety of other iron transporters that may render DMT1 modestly redundant, and DMT1 levels exceed those needed for the just-mentioned tasks. This review begins to consider why and also describes advances after 2008 that begin to address this challenge.

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“…Caco-2 cells are composed solely of absorptive cells whereas the intestinal epithelium is a conglomerate of absorptive enterocytes and other cell types such as goblet cells, endocrine cells, and microfold (M) cells. Several modifications have been made to more closely mimic the heterogeneity of the intestinal epithelium; e.g., co-culturing Caco-2 and HT29 cells introduces the presence of mucin-secreting goblet cells and provides a physiological transport barrier [42,43,44]. Co-culturing Caco-2, HT29, and Raji B cells may more closely resemble intestinal mucosa [48].…”

Section: Discussionmentioning

confidence: 99%

Cell Culture Model for Examining Fat-Soluble Nutrient Absorption <i>In Vitro</i>

Kamil¹,

Blumberg²,

Chen³

2017

JFNR

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Permeable support systems (PS) are employed in in vitro nutrient absorption studies but data are absent on their efficacy compared to conventional cell culture models (CONV). The in vivo absorption of fat soluble nutrients is influenced by its delivery vehicle, yet a fundamental understanding of the influence of the vehicle on cells in culture is lacking. We compared the efficacy of lutein absorption in Caco-2 cells cultured with CONV and PS, and examined the role of micelles, the physiological vehicle within the small intestine. After plating for 2 and 21 d to attain confluence and differentiation in CONV and PS, respectively, cells were treated with lutein in micelles or ethanol. After incubation, lutein in cell lysate, as well as apical and basolateral mediums, were quantified by HPLC-UV. After 24 h, cellular lutein in CONV was ≥460 and 8% greater in ethanol and micelle, respectively, than in PS. However, the intracellular AUC over time was only different for ethanol (P ≤ 0.05). In PS, 0.15% of micellized lutein was secreted into the basolateral medium in contrast to 0.016% of lutein in ethanol. The absorption of lutein (uptake + secretion), independent of the vehicle, in CONV increased in a linear manner with dose (0.35 to 4 or to 14.6 µg/mL for ethanol or micelle, respectively), while that in PS peaked at 1.18 μg/mL. Caco-2 cells cultured in PS grow to display the phenotype and function of small intestine enterocytes and suggest this in vitro platform generates information closest to the natural physiology of the absorptive process. However, although the CONV has the physiology of colonic tissue, it appears to display a greater efficacy for lutein uptake by Caco-2 cells and so can provide a more rapid, preliminary method for nutrient absorption studies.

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Characterization of Caco-2 and HT29-MTX cocultures in an in vitro digestion/cell culture model used to predict iron bioavailability☆

Cited by 253 publications

References 35 publications

Novel Bioengineered Three-Dimensional Human Intestinal Model for Long-Term Infection of Cryptosporidium parvum

Novel Bioengineered Three-Dimensional Human Intestinal Model for Long-Term Infection of Cryptosporidium parvum

Human iron transporters

Cell Culture Model for Examining Fat-Soluble Nutrient Absorption <i>In Vitro</i>

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