Hepatocytes polarize by forming functionally distinct sinusoidal (basolateral) and canalicular (apical) plasma membrane domains. Two distinct routes are used for delivery of membrane proteins to the canaliculus. Proteins having glycosylphosphatidylinositol anchors or single transmembrane domains are targeted to the sinusoidal plasma membrane from where they transcytose to the canalicular domain. In contrast, apical ATP-binding-cassette (ABC) transporters, which are required for energy-dependent biliary secretion of bile acids (ABCB11), phospholipids (ABCB4), and nonbile acid organic anions (ABCC2), lack initial residence in the basolateral plasma membrane and traffic directly from Golgi membranes to the canalicular membrane. While investigating mechanisms of apical targeting in WIF-B9 cells, a polarized hepatic epithelial cell line, we observed that rab11a is required for canalicular formation. Knockdown of rab11a or overexpression of the rab11a-GDP locked form prevented canalicular formation as did overexpression of the myosin Vb motorless tail domain. In WIF-B9 cells, which lack bile canaliculi, apical ABC transporters colocalized with transcytotic membrane proteins in rab11a-containing endosomes and, unlike the transcytotic markers, did not distribute to the plasma membrane. We propose that polarization of hepatocytes (i.e., canalicular biogenesis) requires recruitment of rab11a and myosin Vb to intracellular membranes that contain apical ABC transporters and transcytotic markers, permitting their targeting to the plasma membrane. In this model, polarization is initiated upon delivery of rab11a-myosin Vb-containing membranes to the surface, which causes plasma membrane at the site of delivery to differentiate into apical domain (bile canaliculus).
An antibody produced against rat Y protein, the major cytoplasmic organic anion-binding protein in liver, was characterized. The antibody precipitated Y protein from liver supernatant fractions and specifically removed the organic anion-binding capacity from this fraction.Y protein was detected by immunodiffusion with this antibody in the supernates of rat liver, kidney, and small intestinal mucosa and was not detected in supernates of 16 other tissues including bile and serum. Precipitation with anti-Y was not detected with supernates of liver from 20 other species, including man. Quantitative radial immunodiffusion revealed Y protein to constitute 4.5% of supernatant protein in rat liver and approximately 2% of supernatant protein in rat kidney and small intestinal mucosa. Phenobarbital administration increased the concentration of Y protein in rat liver by 280%, but not in kidney or small intestinal mucosa, and was associated with increased plasma disappearance of sulfobromphthalein sodium, indocyanine green, and bilirubin, and increased hepatic, but not kidney or small intestinal mucosal, content of these organic anions. These observations provide further evidence indicating that the concentration of Y protein is a major determinant of organic anion flux across the plasma membrane of the liver cell.Immunodiffusion and immunoelectrophoresis revealed serological identity between Y protein, cortisol metabolite-binding protein I. and the major azocarcinogen-binding protein.
Microcirculation (2010) 17, 32–38. doi: 10.1111/j.1549-8719.2009.00004.x Objective: Fenestrations are pores in the liver sinusoidal endothelium that facilitate the transfer of particulate substrates between the sinusoidal lumen and hepatocytes. Fenestrations express caveolin-1 and have structural similarities to caveolae, therefore might be a form of caveolae and caveolin-1 may be integral to fenestration structure and function. Therefore, fenestrations were studied in the livers of caveolin-1 knockout mice. Methods: Scanning, transmission and immunogold electron microscopic techniques were used to study the liver sinusoidal endothelium and other tissues in caveolin-1 knockout and wild-type mice. Results: Comparison of fenestrations in wild-type and knockout mice did not reveal any differences on either scanning or transmission electron microscopy. The diameter of the fenestrations was not significantly different (74 ± 13 nm knockout mice vs 78 ± 12 nm wild-type mice) nor was the fenestration porosity (6.5 ± 2.1 knockout vs 7.3 ± 2.4% wild-type mice). In contrast, adipocytes and blood vessels in other tissues lacked caveolae in the knockout mice. Caveolin-1 immunogold of livers of wild-type mice indicated sparse expression in sinusoidal endothelial cells. Conclusions: The normal structure of fenestrations in the liver sinusoidal endothelium is not dependent upon caveolin-1 and fenestrations are not a form of caveolae.
On April 2, 2012, Nelson Fausto, Professor and former Chairman of the Department of Pathology at Washington University School of Medicine, died at the age of 75. During his long struggle with multiple myeloma, he sustained professional and personal activities with characteristics that, throughout his professional life, spawned appreciation, admiration, and friendship from hundreds of colleagues, students and trainees worldwide. A masterful teacher, talented researcher, and caring mentor, Nelson was a role model for young investigators and a leader in advancing knowledge regarding hepatic biology and pathobiology. Nelson was a kindly, intellectual, and emotional giant—a renaissance man who combined the joys of life and work and who profoundly influenced the lives of those with whom he interacted. Like his parents, Nelson was an immigrant, which profoundly influenced his experience, achievements, and humanity. He was born in São Paulo, Brazil, where his parents married after his mother had left Turkey and his father had left the Austro-Hungarian empire to escape pogroms and seek a better life. After the death of his mother when Nelson was an infant, his family struggled to sustain themselves economically and intellectually. Nelson's brothers became distinguished: Boris as a historian and Ruy as a philosopher. Nelson's interest in medicine resulted in an M.D. from the University of São Paulo where, following residency training, he became a faculty member. In 1962, he went to the McCardle Laboratory at the University of Wisconsin for 2 years of postdoctoral training in liver regeneration; however, political upheaval in Brazil changed his life. With family members threatened or fleeing for their lives and with his history of political activism, Nelson remained in the United States and acquired citizenship. Like his parents, he escaped repression and persecution in search of self, freedom, adventure, and growth. In 1967, the leaders of a new medical school at Brown University in Providence, Rhode Island, were prescient in recruiting Nelson as an Assistant Professor of Medical Science, a position he held until 1994. During his tenure at Brown University, Nelson manifested great skills in leadership in teaching, research, administration, and human relationships. His research program in liver regeneration thrived and, with the advent of molecular biology, Nelson was one of the first to use molecular techniques to study regeneration, cancer, and hepatocellular biology. He became Professor of Pathology and founding Chair of the Department of Pathology and Laboratory Medicine, in which capacity he organized and directed the General Pathology course. During the following 11 years, hundreds of students selected him for teaching awards annually. In 1994, he was recruited to the University of Washington Medical School as Chairman of the Department of Pathology. Under his leadership, the Department was spectacularly successful in research and teaching, and became a leading center in modern pathology. Research remained his intellectual pursuit, and Nelson and his colleagues made major contributions in understanding liver regeneration, lineage development, and transformation. Nelson was an academic trailblazer. With colleagues and students, he expanded the field of liver research at a critical time when, in contrast to other areas of medical research, molecular biology still had only a limited impact on liver biology and disease. Nelson encouraged his colleagues to entire this brave new world and tackle long-standing difficult problems in liver biology and disease. Nelson was a leader in academic pathology, a field in which he served as author, spokesman, and innovator. He was President of the American Society of Investigative Pathology (ASIP), and from 1992 to 2001 he served as Editor-in-Chief of The American Journal of Pathology. In 2010, in recognition of his seminal contributions and as "an individual who represents the highest ideals in pathology and medicine," he received the Gold-Headed Cane award from the ASIP, the highest honor offered by that organization. Many international awards followed. Among his most cherished was the Spinoza Chair (University of Amsterdam, The Netherlands, 2000), the Distinguished Scientist Award from the American Liver Foundation (2004), the Distinguished Achievement Award from the American Association for the Study of Liver Disease (2009), the Arnaldo Vieira de Carvalho Medal from the University of São Paulo (2009), and the Distinguished Service Award from the Association of Pathology Chairs (2012). Nelson's influence in pathology and, in particular, liver pathobiology was profound and global. He co-edited the books Robbins and Kumar: The Pathologic Basis of Disease and The Liver: Biology and Pathobiology. Through editorials, reviews, books, lectures, and over 250 peer-reviewed articles, he disseminated his knowledge to an unlimited number of researchers. He influenced several generations of physicians and scientists, including 31 postdoctoral fellows who began their academic careers in his laboratory, 22 graduate students who received their Ph.D. under his mentorship, and innumerable colleagues worldwide who benefitted from his knowledge and willingness to share. Extensive travel to lecture at scientific meetings and educational events provided a format for exchange of ideas with all who were interested. Because of his patient style, frequent traveling, and willingness to learn about all aspects of life, conversations with Nelson were memorable and ranged widely to include science, politics, art, and culture. An avid reader in a wide range of topics, Nelson enjoyed discussing and sharing books that excited his intellect. In addition to mentorship, writing, and teaching, Nelson shared his knowledge and experience in other venues, which also reflected his sense of responsibility to the scientific community. For many years, he served on Study Sections and Advisory Councils for the National Institutes of Health, national and international committees that embraced a range of activities, and as an effective reviewer and editor of professional journals, including HEPATOLOGY. A scholar of many languages and cultures, Nelson bridged these disciplines and lived by the principles he learned from life. Although interested in all cultures, he and his wife, Ann DeLancey, were particularly involved in his home region of the Pacific Northwest. Appreciative of the cultural contributions of the Native American population and aware of the painful shortcomings of reservation life, Nelson and Ann brought Native American middle school children to visit the University of Washington and contributed to medical scholarships for students from Native American tribes. In a symposium held in his honor several months before his death, Native American representatives poignantly praised Nelson's philanthropy, leadership, and concern. Physician, scientist, humanist, and friend, Nelson Fausto's contributions will long influence our lives, even those who did not have the privilege of knowing him personally. He was a giant who was modest about his accomplishments. Nelson enjoyed life to the fullest. A caring, sensitive man, he described his capacity to love by saying that "all the other stuff does not matter." Because I believe he would approve, I have included a favorite photograph of Nelson and Ann in happier days.
