QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online December 15, 2003
doi: 10.1242/10.1242/jcs.00856

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Levy, E. Articles by Bendayan, M. Search for Related Content PubMed PubMed Citation Articles by Levy, E. Articles by Bendayan, M. Social Bookmarking                         What's this?

Ontogeny, immunolocalisation, distribution and function of SR-BI in the human intestine

¹ Department of Nutrition, Hôpital Sainte-Justine and University of Montreal, Montreal QC H3T 1C5, Canada
² Group on the Functional Development and Physiopathology of the Digestive Tract, Canadian Institute of Health Research and Department of Cellular Biology, Faculty of Medicine, Université de Sherbrooke, Sherbrooke QC J1H 5N4, Canada
³ Department of Biochemistry, Hôpital Sainte-Justine and University of Montreal, Montreal QC H3T 1C5, Canada
⁴ Department of Biological Sciences, Université du Québec, Montréal QC H3C 3P8, Canada
⁵ Departments of Pathology and Cell Biology, Hôpital Sainte-Justine and University of Montreal, Montreal QC H3T 1C5, Canada

View larger version (100K): [in a new window]   Fig. 1. Expression and distribution of SR-BI protein along the duodenal crypt-villus axis. Representative indirect immunofluorescence micrographs of cryosections are shown for 14 weeks (A,B) and 20 weeks (C) of gestation, as well as for the adult period (D). Immunofluorescence was also visualized in the microvascular endothelial cells of the lamina propria (E) and muscle cells (F). In all cases, no fluorescent staining was observed when the primary SR-BI antibody was replaced with the appropriate nonimmune serial (data not shown).

View larger version (130K): [in a new window]   Fig. 2. Expression and distribution of SR-BI protein along the jejunal, ileal and colonic crypt-villus axis for 14 weeks of gestation. Representative indirect immunofluorescence micrographs of cryosections of human fetal jejunum (A), ileum (B), proximal colon (C) and distal colon (D). In all cases, no fluorescent staining was observed when the primary SR-BI antibody was replaced with the appropriate nonimmune serial (data not shown). Bars, 50 µm.

View larger version (143K): [in a new window]   Fig. 3. Expression and distribution of SR-BI protein along the jejunal, ileal and colonic crypt-villus axis for 20 weeks of gestation. Representative indirect immunofluorescence micrographs of cryosections of human fetal jejunum (A), ileum (B), proximal colon (C) and distal colon (D). In all cases, no fluorescent staining was observed when the primary SR-BI antibody was replaced with the appropriate nonimmune serial (data not shown). Bars, 50 µm.

View larger version (161K): [in a new window]   Fig. 4. Expression and distribution of SR-BI in adult human gut tissues. The staining is present in all epithelial cells of the crypt-villus axis in the jejunum (A) and ileum (B) as well as in the crypt-surface epithelium of the proximal (C) and distal (D) colon. Bars, 30 µm.

View larger version (104K): [in a new window]   Fig. 5. Immunocytochemical detection of SR-BI in fetal duodenal mucosa. Protein A-gold immunocytochemical technique was applied to localize SR-BI in absorptive cells of duodenal tissue at 17 weeks (A) and 20 weeks (B) of gestation as well as at the adult period (C,D). The gold particles revealing SR-BI antigenic sites are mainly associated with the luminal plasma membrane lining the microvilli (mv). They are also present in endosomal invaginations and vesicles (e). The labelling within the cell, although of lower intensity, is observed in the rough endoplasmic reticulum (RER), the Golgi apparatus (G) and the basolateral membrane (blm). Mitochondria (m) are devoid of labelling.

View larger version (18K): [in a new window]   Fig. 6. Pattern of SR-BI distribution in different regions of human fetal intestine. Immunoblotting analysis was performed at week 20 of the gestational period. The intestinal regions are: duodenum (D), jejunum (J), ileum (I), proximal colon (PC) and distal colon (DC). Values are means ± s.e. of 4-6 experiments/group.

View larger version (31K): [in a new window]   Fig. 7. SR-BI protein expression in human small intestine and colon from 15-20 weeks of gestation. Each intestinal segment was analyzed separately with similar protein amounts at the various periods of gestation. However, protein amounts were different among intestinal regions in order to increase the electrophoretic resolution.

View larger version (104K): [in a new window]   Fig. 8. Expression and distribution of caveolin-1 along the duodenal and colonic crypt-villus axis. Representative indirect immunofluorescence micrographs of cryosections are shown for the 20 weeks of gestation (A,C) as well as for the adult period (B,D).

View larger version (14K): [in a new window]   Fig. 9. Association of SR-BI with caveolin-1 in intestinal epithelial cells. Homogenates of epithelial cells (detached from the duodenum) and Caco-2 cells were incubated with anti-SR-BI (a-SR-BI) antibody. The immunoprecipitates were run on SDS-PAGE and transferred to a nitrocellulose membrane. The membrane was blotted with anti-SR-BI (a-SR-BI) or anti-caveolin-1 (a-cav-1) antibodies. Immunoblotting with a-SR-BI revealed the SR-BI protein, whereas immunoblotting with a-cav-1 did not display any signal. The lack of co-precipitation and the absence of caveolin-1 signal refuted the presence of caveolin-1 and potential physical interaction between SR-BI and caveolin-1. STD, standard.

View larger version (32K): [in a new window]   Fig. 10. Disruption of SR-BI expression using SR-BI antisense (AS) oligodeoxynucleotides in Caco-2 cells. Following antisense treatment, cells were incubated with fresh MEM medium allowed to grow and tested for SR-BI expression at the differentiated state. Three clones (AS1, AS2, AS3) presenting decreased SR-BI protein levels (compared with cells transfected with vector without insert) were selected. Values are means ± s.e. of three experiments.

View larger version (22K): [in a new window]   Fig. 11. Effect of SR-BI antisense treatment on cell proliferation and differentiation. As examined by [3H]-thymidine incorporation (A), sucrase activity (B) and transepithelial resistance (C), antisense treatment did not affect cell growth differentiation and cell integrity.Values are means ± s.e. of three experiments.

View larger version (91K): [in a new window]   Fig. 12. Immunocytochemical labelling of SR-BI in Caco-2 cells following antisense treatment. To confirm the disruption of SR-BI expression using SR-BI antisense oligodeoxynucleotides, we applied the immunogold protein A technique on Caco-2 cells transfected with the vector pZeoSV without insert (A) or with the human SR-BI cDNA inserted in an antisense orientation (B). Upper panels show the apical region of the cell, and lower panels illustrate the basolateral region of the cell. Reduced labelling is noticed in microvilli (mv) and the basolateral membrane as well as within endocytotic vesicles (arrows). Magnification: A, upper panel x38,000; lower panel x36,000; B, upper panel x40,000; lower panel x56,000.

View larger version (25K): [in a new window]   Fig. 13. Impact of SR-BI disruption on free cholesterol, phosphatidylcholine and cholesteryl ester uptake in Caco-2 cells. Following the inhibition of SR-BI expression using SR-BI antisense oligodeoxynucleotides, Caco-2 cells were incubated with free [14C]-cholesterol (A), [14C]-phosphatidylcholine (B) or [14C]-cholesteryl ester (C).

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter