Passage
The lumen of the human gut is lined by a monolayer of epithelial cells that acts as a selectively permeable barrier, preventing the passage of harmful intraluminal foreign antigens, flora, and toxins into the circulation while allowing digestion and absorption of essential dietary nutrients along with the transfer of electrolytes and water.Proteins in the tight junctions of intestinal epithelial cells maintain barrier integrity, but barrier dysfunction occurs when these cells are damaged in the setting of infection, burns, shock, or hypoxia (low oxygen levels) . The transcription factor HIF-1, a heterodimer composed of the macromolecules HIF-1α and HIF-1β, regulates the adaptive cellular response to hypoxia and the consequent expression of tight junction proteins.Researchers assessed the concentration of HIF-1 heterodimer components in human intestinal Caco-2 cells subjected to hypoxia/reoxygenation (H/R) in vitro. Caco-2 cells, a colon-derived cell line, were cultured under specific conditions to mimic the functional and morphological phenotype of wild-type enterocytes lining the small intestine. These cells were prepared and grown as a monolayer on a collagen-coated membrane.Next, the monolayer was cultured in hypoxic conditions and then exposed to atmospheric oxygen levels (normoxia) for 30, 60, and 120 minutes. Protein levels were quantified using direct enzyme-linked immunosorbent assay (ELISA) , in which an antibody linked to a reporter enzyme was utilized to bind and detect expression of the target molecule (analyte) in a sample. When the colorless substrate of the reporter enzyme was added, the enzyme generated a visible colored product that could be quantified based on color intensity.
Figure 1 Concentration of (A) HIF-1α and (B) HIF-1β in Caco-2 cells subjected to H/R (Note: 0 minutes = cells cultured in hypoxic conditions) Emodin, an anthraquinone compound that prevents hypoxia-induced epithelial cell disruption, was used in conjunction with a chemical HIF-1α inhibitor (HIF-1α-I) to treat Caco-2 cells in a separate experiment. Transepithelial electrical resistance (TEER) was assessed as a measure of barrier function, with higher TEER values indicating a more intact epithelial cell barrier. HIF-1α-I was found to block emodin's protective effect on epithelial barrier integrity.
Adapted from Lei Q, Qiang F, Chao D, et al. Int J Mol Med. 2014;34(6) :1629-39.
-Which graphic depicts the most likely effect of the HIF-1α inhibitor on emodin? (Note: control = cells cultured under normoxic conditions.)

Question

Passage
The lumen of the human gut is lined by a monolayer of epithelial cells that acts as a selectively permeable barrier, preventing the passage of harmful intraluminal foreign antigens, flora, and toxins into the circulation while allowing digestion and absorption of essential dietary nutrients along with the transfer of electrolytes and water.Proteins in the tight junctions of intestinal epithelial cells maintain barrier integrity, but barrier dysfunction occurs when these cells are damaged in the setting of infection, burns, shock, or hypoxia (low oxygen levels) . The transcription factor HIF-1, a heterodimer composed of the macromolecules HIF-1α and HIF-1β, regulates the adaptive cellular response to hypoxia and the consequent expression of tight junction proteins.Researchers assessed the concentration of HIF-1 heterodimer components in human intestinal Caco-2 cells subjected to hypoxia/reoxygenation (H/R) in vitro. Caco-2 cells, a colon-derived cell line, were cultured under specific conditions to mimic the functional and morphological phenotype of wild-type enterocytes lining the small intestine. These cells were prepared and grown as a monolayer on a collagen-coated membrane.Next, the monolayer was cultured in hypoxic conditions and then exposed to atmospheric oxygen levels (normoxia) for 30, 60, and 120 minutes. Protein levels were quantified using direct enzyme-linked immunosorbent assay (ELISA) , in which an antibody linked to a reporter enzyme was utilized to bind and detect expression of the target molecule (analyte) in a sample. When the colorless substrate of the reporter enzyme was added, the enzyme generated a visible colored product that could be quantified based on color intensity.

Figure 1 Concentration of (A) HIF-1α and (B) HIF-1β in Caco-2 cells subjected to H/R (Note: 0 minutes = cells cultured in hypoxic conditions) Emodin, an anthraquinone compound that prevents hypoxia-induced epithelial cell disruption, was used in conjunction with a chemical HIF-1α inhibitor (HIF-1α-I) to treat Caco-2 cells in a separate experiment. Transepithelial electrical resistance (TEER) was assessed as a measure of barrier function, with higher TEER values indicating a more intact epithelial cell barrier. HIF-1α-I was found to block emodin's protective effect on epithelial barrier integrity.
Adapted from Lei Q, Qiang F, Chao D, et al. Int J Mol Med. 2014;34(6) :1629-39.
-Which graphic depicts the most likely effect of the HIF-1α inhibitor on emodin? (Note: control = cells cultured under normoxic conditions.)

Quizplus · Accepted Answer

11ecba2d_0f31_cff8_a3bf_ff0c78cb343d_MD0008_00 The control cells in this experiment have been cultured under normal oxygen conditions, and therefore have an intact epithelial barrier and high TEER, which is a measure of proper barrier function. Because hypoxia damages the epithelial barrier, cells cultured under low oxygen conditions have decreased TEER compared to controls, signifying compromised barrier integrity. Emodin prevents the epithelial damage induced by hypoxia. As a result, cells grown in hypoxic conditions but treated with emodin have a TEER value comparable to the control. However, cells also grown in hypoxic conditions but treated with both emodin and the HIF-1α inhibitor have a low TEER compared to the control. This result is due to the HIF-1α inhibitor blocking emodin's ability to maintain an intact epithelial barrier under hypoxic conditions, as stated in the passage. (Choice B) This graph shows a reverse finding of the true experimental result, and therefore is invalid. (Choice C) Cells cultured in hypoxic conditions have a low TEER, but this graph shows that these cells have a high TEER. It also shows a reverse finding of the true result for cells treated with emodin and for those treated with both emodin and the HIF-1α inhibitor. (Choice D) Cells cultured with the inhibitor and emodin in hypoxic conditions ("Hypoxia + emodin + inhibitor") should have low TEER. However, this graph shows that TEER is high under these conditions. Educational objective: Inhibitors prevent chemical reactions, block gene/protein expression, or reduce the activity of a particular substance (eg, enzyme, reactant). Transcription factors can be inhibited by certain drugs or substances.