Expression of SARS-CoV-2 entry factors, electrolyte, and mineral transporters in different mouse intestinal epithelial cell types.

Angiotensin-converting enzyme 2 (ACE2) and transmembrane proteases (TMPRSS) are multifunctional proteins required for SARS-CoV-2 infection or for amino acid (AA) transport, and are abundantly expressed in mammalian small intestine, but the identity of the intestinal cell type(s) and sites of expression are unclear. Here we determined expression of SARS-CoV-2 entry factors in different cell types and then compared it to that of representative AA, electrolyte, and mineral transporters. We tested the hypothesis that SARS-CoV-2, AA, electrolyte, and mineral transporters are expressed heterogeneously in different intestinal cell types by making mouse enteroids enriched in enterocytes (ENT), goblet (GOB), Paneth (PAN), or stem (ISC) cells. Interestingly, the expression of ACE2 was apical and modestly greater in ENT, the same pattern observed for its associated AA transporters B0 AT1 and SIT1. TMPRSS2 and TMPRSS4 were more highly expressed in crypt-residing ISC. Expression of electrolyte transporters was dramatically heterogeneous. DRA, NBCe1, and NHE3 were greatest in ENT, while those of CFTR and NKCC1 that play important roles in secretory diarrhea, were mainly expressed in ISC and PAN that also displayed immunohistochemically abundant basolateral NKCC1. Intestinal iron transporters were generally expressed higher in ENT and GOB, while calcium transporters were expressed mainly in PAN. Heterogeneous expression of its entry factors suggests that the ability of SARS-CoV-2 to infect the intestine may vary with cell type. Parallel cell-type expression patterns of ACE2 with B0 AT1 and SIT1 provides further evidence of ACE2's multifunctional properties and importance in AA absorption.

Cell-Type-Specific, Ketohexokinase-Dependent Induction by Fructose of Lipogenic Gene Expression in Mouse Small Intestine.

BACKGROUND: High intakes of fructose are associated with metabolic diseases, including hypertriglyceridemia and intestinal tumor growth. Although small intestinal epithelia consist of many different cell types, express lipogenic genes, and convert dietary fructose to fatty acids, there is no information on the identity of the cell type(s) mediating this conversion and on the effects of fructose on lipogenic gene expression. OBJECTIVES: We hypothesized that fructose regulates the intestinal expression of genes involved in lipid and apolipoprotein synthesis, that regulation depends on the fructose transporter solute carrier family 2 member a5 [Slc2a5 (glucose transporter 5)] and on ketohexokinase (Khk), and that regulation occurs only in enterocytes. METHODS: We compared lipogenic gene expression among different organs from wild-type adult male C57BL mice consuming a standard vivarium nonpurified diet. We then gavaged twice daily for 2.5 d fructose or glucose solutions (15%, 0.3 mL per mouse) into wild-type, Slc2a5-knockout (KO), and Khk-KO mice with free access to the nonpurified diet and determined expression of representative lipogenic genes. Finally, from mice fed the nonpurified diet, we made organoids highly enriched in enterocyte, goblet, Paneth, or stem cells and then incubated them overnight in 10 mM fructose or glucose. RESULTS: Most lipogenic genes were significantly expressed in the intestine relative to the kidney, liver, lung, and skeletal muscle. In vivo expression of Srebf1, Acaca, Fasn, Scd1, Dgat1, Gk, Apoa4, and Apob mRNA and of Scd1 protein increased (P < 0.05) by 3- to 20-fold in wild-type, but not in Slc2a5-KO and Khk-KO, mice gavaged with fructose. In vitro, Slc2a5- and Khk-dependent, fructose-induced increases, which ranged from 1.5- to 4-fold (P < 0.05), in mRNA concentrations of all these genes were observed only in organoids enriched in enterocytes. CONCLUSIONS: Fructose specifically stimulates expression of mouse small intestinal genes for lipid and apolipoprotein synthesis. Secretory and stem cells seem incapable of transport- and metabolism-dependent lipogenesis, occurring only in absorptive enterocytes.

Protective effects of eicosapentaenoic acid in adipocyte-breast cancer cell cross talk.

Breast cancer is the leading cause of death in women among all cancer types. Obesity is one of the factors that promote progression of breast cancer, especially in post-menopausal women. Increasingly, adipose tissue is recognized for its active role in the tumor microenvironment. We hypothesized that adipocytes conditioned medium can impact breast cancer progression by increasing inflammatory cytokines production by cancer cells, and subsequently increasing their motility. By contrast, eicosapentaenoic acid (EPA), an anti-inflammatory n-3 polyunsaturated fatty acid, reduces adipocyte-secreted inflammatory factors, leading to reduced cancer cell motility. To test these hypotheses, we investigated the direct effects of EPA on MCF-7 and MDA-MB-231 breast cancer cells and the effects of conditioned medium from 3 T3-L1 or human mesenchymal stem cells (HMSC)-derived adipocytes treated with or without EPA supplementation on breast cancer cells. We observed that conditioned medium from HMSC-derived adipocytes significantly increased mRNA transcription levels of cancer-associated genes such as FASN, STAT3 and cIAP2, while EPA-treated HMSC-derived adipocytes significantly reduced mRNA levels of these genes. However, direct EPA treatment significantly reduced mRNA content of these tumor-associated markers (FASN, STAT3, cIAP-2) only in MDA-MB-231 cells not in MCF-7 cells. Conditioned medium from EPA-treated 3 T3-L1 adipocytes further decreased inflammation, cell motility and glycolysis in cancer cells. Our data confirms that adipocytes play a significant role in promoting breast cancer progression and demonstrates that EPA-treated adipocytes reduced the negative impact of adipocyte-secreted factors on breast cancer cell inflammation and migration.

Low dose radiation, inflammation, cancer and chemoprevention.

PURPOSE: Recently, new studies have brought to light the potential risks of low dose radiation (LDR) in cancer. In this review, we discuss in detail the detrimental effects of LDR in some model organisms and animal models, as well as potential risks to human beings from some routine medical screening procedures. Furthermore, cellular mechanisms by which LDR exerts its negative effects like endoplasmic reticulum stress, epigenetic changes and microRNAs are also reviewed. A few studies are discussed that have reported some benefits of LDR through changes in energy metabolism. Lastly, we focus on breast cancer, one of the predominant forms of cancer potentially affected by LDR and some of the benefits of n-3 polyunsaturated fatty acids (PUFA) as dietary compounds that offer protection against radiation effects on cancer cells and cancer progression. CONCLUSIONS: Overall, LDR exerts mainly damaging effects through diverse cell and molecular mechanisms, with a few beneficial effects reported. In some cancers, surrounding adipose tissue of the breast may contribute to obesity-related cancer. Further, preclinical data suggest that anti-inflammatory dietary compounds such as PUFA and other dietary interventions may protect against radiation effects on cancer cells and cancer progression.

Marked differences in tight junction composition and macromolecular permeability among different intestinal cell types.

BACKGROUND: Mammalian small intestinal tight junctions (TJ) link epithelial cells to one another and function as a permselective barrier, strictly modulating the passage of ions and macromolecules through the pore and leak pathways, respectively, thereby preventing the absorption of harmful compounds and microbes while allowing regulated transport of nutrients and electrolytes. Small intestinal epithelial permeability is ascribed primarily to the properties of TJs between adjoining enterocytes (ENTs), because there is almost no information on TJ composition and the paracellular permeability of nonenterocyte cell types that constitute a small but significant fraction of the intestinal epithelia. RESULTS: Here we directed murine intestinal crypts to form specialized organoids highly enriched in intestinal stem cells (ISCs), absorptive ENTs, secretory goblet cells, or Paneth cells. The morphological and morphometric characteristics of these cells in organoids were similar to those in vivo. The expression of certain TJ proteins varied with cell type: occludin and tricellulin levels were high in both ISCs and Paneth cells, while claudin-1, -2, and -7 expression was greatest in Paneth cells, ISCs, and ENTs, respectively. In contrast, the distribution of claudin-15, zonula occludens 1 (ZO-1), and E-cadherin was relatively homogeneous. E-cadherin and claudin-7 marked mainly the basolateral membrane, while claudin-2, ZO-1, and occludin resided in the apical membrane. Remarkably, organoids enriched in ENTs or goblet cells were over threefold more permeable to 4 and 10 kDa dextran compared to those containing stem and Paneth cells. The TJ-regulator larazotide prevented the approximately tenfold increases in dextran flux induced by the TJ-disrupter AT1002 into organoids of different cell types, indicating that this ZO toxin nonselectively increases permeability. Forced dedifferentiation of mature ENTs results in the reacquisition of ISC-like characteristics in TJ composition and dextran permeability, suggesting that the post-differentiation properties of TJs are not hardwired. CONCLUSIONS: Differentiation of adult intestinal stem cells into mature secretory and absorptive cell types causes marked, but potentially reversible, changes in TJ composition, resulting in enhanced macromolecular permeability of the TJ leak pathway between ENTs and between goblet cells. This work advances our understanding of how cell differentiation affects the paracellular pathway of epithelia.

Protective properties of n-3 fatty acids and implications in obesity-associated breast cancer.

Obesity is well documented as a risk factor for developing breast cancer, especially in postmenopausal women. Adipose tissue in the breast under obese conditions induces inflammation by increasing macrophage infiltration and pro-inflammatory cytokines that in turn up-regulates genes and signaling pathways, resulting in increased inflammation, cell proliferation and tumor growth in the breast. Due to their potent anti-inflammatory effects, n-3 polyunsaturated fatty acids (n-3 PUFA) are a promising and safe dietary intervention in reducing breast cancer risk. Here, we briefly review current status of breast cancer and its relationship with obesity. We then review in depth, current research and knowledge on the role of n-3 PUFA in reducing/preventing breast cancer cell growth in vitro, in vivo and in human studies, and how n-3 PUFA may modulate signaling pathways mitigating their effects on breast cancer development.

Metabolic profiling in nutrition and metabolic disorders.

Nutrients exert potent effects on metabolism through a variety of regulatory mechanisms, resulting in local and systemic changes in metabolite levels. Numerous studies have focused on mechanisms by which nutrients and disease states regulate metabolism at the gene or protein levels using genomic and proteomic approaches, respectively. However, few studies have investigated nutritional regulation of the whole metabolome. Thus, metabolomic approaches have recently emerged to complement the genomics and proteomics research and to help identify biologically meaningful metabolites and metabolic networks that control cellular responses to genetic and environmental factors, including diet, and to identify metabolic diseases that are influenced by genetic and dietary factors. These large-scale studies expedite our ability to develop targeted treatments. The goal of this symposium was to provide a forum to introduce the metabolomics field to nutrition researchers. An overview of the state-of-the-art metabolomic technologies used was provided. The impact of some specific nutrients, disease states, or genetic variations and their interaction with the metabolome was discussed by the speakers. Our objectives were as follows: 1) to educate the audience about the use of metabolomics as an innovative tool for linking changes in cell metabolites and genetic variations to nutrient metabolism, energy balance, and the overlying effects on health and disease; 2) to understand the concept of metabolomics and describe the analytical tools and resources available in this area; 3) to introduce the potential application of metabolomics in the field of nutrition research; and 4) to provide specific nutrition-relevant metabolomics study examples in investigating regulation of the metabolic network or metabolic changes resulting from disease states by dietary factors.