Herrera-Ruiz D, Wang Q, Gudmundsson OS, Cook TJ, Smith RL, Faria TN and Knipp GT Spatial Expression Patterns of Peptide Transporters in the Human and Rat Gastrointestinal Tracts, Caco-2 In Vitro Cell Culture Model, and Multiple Human Tissues AAPS PharmSci 2001; 3 (1) article 9 (https://www.pharmsci.org/scientificjournals/pharmsci/journal/01_09.html).

Figures and Tables

Table 1. Sequence-specific Primers Used for the Amplification of the Human and Rat Peptide Transporter Isoforms and β-actin, a Positive Control, Based on Their Respective, Reported Encoding cDNA Regions

*Based on preliminary human sequence; see text.

† Based on the human PTR3 sequence.

Figure 1.RT-PCR analysis of human PepT1, PTR3, PHT1, and HPT-1 mRNAs in the human esophagus (A), stomach (B), duodenum (C), jejunum (D), ileum (E), ileocecum (F), cecum (G), ascending colon (H), transverse colon (I), descending colon (J), rectum (K), and in Caco-2 cells (L). RT-PCR was performed with specific primers for each mRNA and amplified products of PepT1, PTR3, PHT1, and HPT-1 were 588, 470, 443, and 1004 bp, respectively. Reaction products were electrophoretically separated in 1.4% agarose gels, stained with ethidium bromide (top panels), and identity confirmed by Southern Blot analysis (lower panels). Commercially available human β-actin primers were used to generate a mRNA expression positive control, amplifying a product of 303 bp.

Figure 2.RT-PCR analysis of rat PepT1, PTR3, PHT1, and RPT1 mRNAs in the rat stomach (A), duodenum (B), jejunum (C), ileum (D), ileocecal junction (E), cecum (F), and colon (G). RT-PCR was performed using specific primers for rat PepT1, PHT1, and RPT-1 mRNAs amplifying products of 523, 437, and 860 bp, respectively. Analysis of PTR3 mRNA expression in the rat tissues was performed using primers designed from the human PTR3 mRNA sequence (Table 1). Reaction products were electrophoretically separated in 1.4% agarose gels, stained with ethidium bromide (top panels), and identity confirmed by Southern Blot analysis (lower panels). Rat-specific β-actin primers were designed to generate a mRNA expression-positive control, amplifying a product of 375 bp.

Figure 3.RT-PCR analysis of human PepT1, PTR3, PHT1, and HPT-1 mRNAs in Caco-2 cells at day 4 (A), day 10 (B), day 15 (C), day 20 (D), day 25 (E), and day 30 (F) after being passaged. RT-PCR was performed with the same set of primers used for the analysis of mRNA expression in the human gastrointestinal tract. The Caco-2 cell line was obtained from the American Type Culture Collection (Rockville, MD) and cultured as described previously. Reaction products were electrophoretically separated in 1.4% agarose gels, stained with ethidium bromide (top panels), and identity confirmed by Southern Blot analysis (lower panels). Commercially available human β-actin primers were used to generate a mRNA expression-positive control, amplifying a product of 303 bp.

Figure 4.RT-PCR analysis of human PepT1, PTR3, PHT1, and HPT-1 mRNAs in the brain (A), colon (B), heart (C), kidney (D), leukocytes (E), liver (F), lung (G), ovary (H), pancreas (I), placenta (J), prostate (K), skeletal muscle (L), small intestine (M), spleen (N), testis (O), and thymus (P). RT-PCR was performed with specific primers for each mRNA, and amplified products of human PepT1, PTR3, PHT1, and HPT-1 were 588, 470, 443, and 1004 bp, respectively. Reaction products were electrophoretically separated in 1.4% agarose gels, stained with ethidium bromide (top panels), and identity confirmed by Southern Blot analysis (lower panels). Commercial available human β-actin primers were used to generate a mRNA expression-positive control, amplifying a product of 303 bp.