Volume 15 Number 2 February i992
patients with severe hypotension (blood pressure less than 60 m m Hg). 2 They cited no less than 12 references to corroborate the following statement, "A correlation between preoperative blood pressure and death in this condition has been noted by most other investigators, and this relationship is further reinforced by the data in the present study." Wakefield et al. 3 observed a 78% mortality rate when the blood pressure was less than 80 m m H g in patients with ruptured abdominal aortic aneurysms. They concluded, "Patients whose systolic blood pressures were lower than 80 mm H g had the highest overall and intraoperative mortality..." This leads to a disturbing aspect of the editorial. Dr. Crawford suggests that, " . . . no significant attempt should be made for blood volume resuscitation until at the time of operation. Systolic blood pressure should be maintained at 50-70 mm H g . . . . " H e claims that hypotension slows bleeding and allows local clot formation, and he labels the Harborview policy o f fluid resuscitation naive and unsophisticated. The essential problem underlying all shock states is maldistribution of blood flow at the microcirculatory level. This compromises tissue oxygenation and results in less oxygen consumption than needed with increased metabolic activity. As blood flow increases to the liver, heart, and brain, it is shunted away from the skin, gut, and kidneys. Oxygenation of these tissues is further reduced, and eventually metabolic acidosis and resistance to catecholamines results. We were unable to find any exception to the principle that hypotension from hemorrhagic shock should be treated by rapid administration of volume to achieve systolic blood pressures above 100 m m Hg. 4,s The "plugging" and "tamponade" of bleeding sites with hypotension that Dr. Crawford hails as therapeutic actually play a major role in the progression of hemorrhagic shock. With low flow, sludging of red blood cells, white blood cells, and platelets increase a microcirculatory viscosity and decrease a tissue perfusion. 5 Again, the result is hypoxia, acidosis, and, if prolonged, irreversible shock. Based on the available physiologic data pertaining to hypovolemic shock, its treatment by maintenance of a blood pressure between 50 and 70 mm H g is dangerous at best and clearly unfounded. Dr. Crawford's patients comprise a highly selected group. Ninety percent of them are able to endure an ambulance ride to a helicopter followed by an air flight of variable duration. Many of the patients we, and doubtlessly the Harborview group, see are too sick and unstable to survive a first-class helicopter ride. They do, however, make it to our operating rooms. Dr. Crawford criticizes the knowledge, experience and technical ability of the Harborview group because his resuks contrast so strikingly with theirs. We reiterate that Dr. Crawford's results contrast strikingly,~fith those of the rest o f the nation. A comparison of his helicopter triage system to what is available to other skilled and competent vascular surgeons around the nation is unfair.
Letters to the Editors
457
While we praise the Harborview Group for their management of a difficult problem, we may also continue to attempt to emulate the standards set by Dr. Crawford. Steven G. Friedman, M D IC K I&ishnasastry, M2D
REFERENCES 1. Ouriel K, Geary K, Green RM, et al. Factors determining survival after ruptured aortic aneurysm: the hospital, the surgeon, and the patient, l Vasc SuR6 1990;11:493-6. 2. Shackleton CR, Schecter MT, Bianco R, et al. Preoperative predictors of mortality risk in ruptured abdominal aortic aneurysm. J Vasc SURG 1987;6:583-9. 3. Wakefield TW, Whitehouse WM, Wu SC, et al. Abdominal aortic aneurysm rupture: statistical analysis of factors affecting outcome of surgical treatment. Surgery 1982;91:586-96. 4. Shoemaker WC. Pathophysiology and therapy of shock syndromes. In: Shoemaker WC, Thompson VffL, Holbrook PR, eds. Textbook of critical care. 1~ted. Philadelphia: WB Saunders Co, i984;52-72. 5. Peityman A. Principles of circulatory support and the treatment of hemorrhagic shock. In: Snyder IV, Pursky MR, eds. Oxygen transport in the critically ill. I st ed. Chicago: Year Book Medical Publishers, Inc., 1987;407-18. 24/41/31837
Cells derived from omental fat tissue and used for seeding vascular prostheses are not endothelial in origin To the Editors:
A recent report by Visser et al. 1 presents evidence that cells isolated and cultured from human omentum represent a mixture of mesothelial and endothelial cells. Their subsequent conclusion that a major cell type in human omentum is rnesothelium is well substantiated by their own data as well as numerous previous reports by other authors on this subject. 2~ However, Visser et al. extend the interpretation of their data to claim a large number of investigators, including ourselves, have misinterpreted our own findings. We strongly disagree with many of t h e subsequent conclusions made in this manuscript and wish to draw attention to several misinterpretations and omissions. First, Dr. Visser uses the terms omentum, omental tissue, and omental-associated fat almost interchangeably. This is incorrect since these terms describe at least two morphologically distinct tissues. O m e n m m is an anatomic structure consisting of a serosal-covered surface enclosing larger vessels, many lymphatic structures, and adipose tissue. Mesothelial cells forming the serosal surface represent a large proportion of the cell number because of the high surface area-to-contained-volume ratio, and thus one would expect a predominant number of mesothelial cells from collagenase digestion. On the other hand, fat tissue is morphologically quite different from omentum and can be
458
Journal of VASCULAR SURGERY
Letters to the Editors
described as adipocytes perfused by a rich network of microvascular endothelial cells. This critical distinction has been previously discussed in a publication by us, 6 but a discussion of this distinction is not found in Dr. Visser's paper. Thus an important point not discussed by Dr. Visser is the importance of isolating fat free from surrounding mesothelial cell serosal capsule. When this is performed the sheets of cells described by Dr. Visser are not observed in the isolate, thus significantly reducing the number o f mesothelial cells present. Dr. Visser also omitted descriptions o f alternate sources of fat for microvessel endothelial cell isolation, which can be used for vascular graft endothelialization studies. We have previously reported the use o f subcutaneous fat for endothelial cell isolation, a tissue that is not surrounded by a mesothelial cell capsule. Subcutaneous fat has numerous advantages over the use of fat associated with the omentum since it can be obtained by liposuction. Fat occurs in numerous other sites in the body in morphologically distinct form. In conclusion Dr. Visser and colleagues, through significant omissions of published papers and misapplication of their own findings on omentum, have suggested that cells isolated and cultured from fat are mesothelium and not endothelium. Although the merits o f using omentum to derive cells for vascular graft treatments deserve continued study as proposed by Dr. Visser, most o f the conclusions concerning the identity of cells derived from fat drawn by Dr. Visser and colleagues are inappropriate. Stuart IC Williams, PhD Bruce E. Jarrell, ME) Department of Surgery University of Arizona Tucson, AZ 85724 REFERENCES
1. Visser MJT, van Bockel JH, van Muijen GNP, van Hindbergh VWM. Cells derived from omental fat tissue and used for seeding vascular prostheses are not endothelial in origin. J VAsc SURG 1991;13:378-81. 2. Potzsch B, Gmlich-Henn J, Rossing R, Wille D, MullerBerghans G. Identification of endothelial and mesothelial cells in human omental tissue and in omentum-derived culture cells by specific cell markers. Lab Invest 1990;63:841-52. 3. Stylianou E, Jenner LA, Davies M, Coles GA, Williams JD. Isolation culture and characterization of human peritoneal mesothelial cells. Kid Int 1990;37:1563-70. 4. van Hinsberg VWM, Kooistra T, Scheffer MA, van Bockel JH, van Muijen GNP. Characterization and fibrinolytic of human omental tissue mesothelial cells. Comparison with endothelial cells. Blood 1990;75:1490-7. 5. Hata J, Goto T, Mukai K, Sawasaki Y, Takahashi K. Cobblestone monolayer cells from human omental adipose tissue are possibly mesothelial not endothelial. In Vitro Cell Dev Biol 1989;25:109-11. 6. Williams SK, Jarrell BE, Rose DR, et al. Human microvessel endothelial cell isolation and vascular graft sodding in the operating room. Ann Vasc Surg 1989;3:146-52. 24/41/32075
Reply To the Editors." We appreciate the opportunity to respond to Drs. Williams' and JarreU's criticisms, which we strongly dispute. They raise an important issue that gives us the chance to stress that the nature o f cells derived from omental fat tissue needs to be properly evaluated. The point made by Drs. Williams and Jarrell that the terms omentum, omental tissue, and omental-associated fat were used almost interchangeably in our report is well taken. The greater omentum can be defined as an anatomic structure consisting o f a fourfold layer of serosal tissue. This anatomic structure contains adipose tissue in the adult human. In the reports we cited in our article either the term omentum 1 or omentum and omental tissue 2 were used to refer to the source of the presumed microvascular endothelial cells. Of course, these investi gators did not use the 'omentum' as 'serosal covere~ surfaces' as defined by Drs. Williams and Jarrell, because it is not logical to isolate endothelial cells from this material, which contains no microvessels. In their previous work, which was cited by us, 3-5 Drs. Williams and Jarrell introduce confusion by not precisely mentioning how the omental tissue was prepared. Particularly, they never mentioned that the serosal layers were separated from the actual fat tissue. There can, however, be no misunderstanding as to whether we used true omentum or omental fat as the source of the epitheloid cells we described. In the "Materials and Methods" section of our report the precise type of tissue used is defined in the second sentence: "The cells derived from omentum, which were presumed to be microvascular endothelial cells, were isolated from human omental f a t tissue." Drs. Williams and Jarrell state that no sheet structures are observed after collagenase digestion of isolated omental fat. They then mistakenly draw the conclusion that the number of mesothelial cells is significantly reduced. The absence of sheet structures is not a proof that the number o f mesothelial cells is reduced, because the presence of sheet structures depends on the applied isolation techniques like collagenase digestion o f omental tissue: type of collagenase, batch differences, concentration, and incubation time. Although the number of mesothelial cells contaminating the yield of microvascular endothelial cells might be reduced by isolation of omental fat tissue, free from serosal capsule, the adequate identification of the nature of the cells remains an important issue. The nature of the epitheloid cells that are isolated after collagenase digestion must certainly not be evaluated by the presence or absence of sheet structures neither must it be exclusively evaluated by immunofluorescence staining with polyclonal anti-yon Willebrand factor antibodies, as many investigators do. A more appropriate method to establish the nature o f these cells is to evaluate the presence of yon Willebrand factor antigens by mouoclonal antibodies and the presence or absence of cytokeratins 8 and 18. The second point Drs. Williams and Jarrell make is that we omitted descriptions o f alternate sources of fat for
patients with severe hypotension (blood pressure less than 60 m m Hg). 2 They cited no less than 12 references to corroborate the following statement, "A correlation between preoperative blood pressure and death in this condition has been noted by most other investigators, and this relationship is further reinforced by the data in the present study." Wakefield et al. 3 observed a 78% mortality rate when the blood pressure was less than 80 m m H g in patients with ruptured abdominal aortic aneurysms. They concluded, "Patients whose systolic blood pressures were lower than 80 mm H g had the highest overall and intraoperative mortality..." This leads to a disturbing aspect of the editorial. Dr. Crawford suggests that, " . . . no significant attempt should be made for blood volume resuscitation until at the time of operation. Systolic blood pressure should be maintained at 50-70 mm H g . . . . " H e claims that hypotension slows bleeding and allows local clot formation, and he labels the Harborview policy o f fluid resuscitation naive and unsophisticated. The essential problem underlying all shock states is maldistribution of blood flow at the microcirculatory level. This compromises tissue oxygenation and results in less oxygen consumption than needed with increased metabolic activity. As blood flow increases to the liver, heart, and brain, it is shunted away from the skin, gut, and kidneys. Oxygenation of these tissues is further reduced, and eventually metabolic acidosis and resistance to catecholamines results. We were unable to find any exception to the principle that hypotension from hemorrhagic shock should be treated by rapid administration of volume to achieve systolic blood pressures above 100 m m Hg. 4,s The "plugging" and "tamponade" of bleeding sites with hypotension that Dr. Crawford hails as therapeutic actually play a major role in the progression of hemorrhagic shock. With low flow, sludging of red blood cells, white blood cells, and platelets increase a microcirculatory viscosity and decrease a tissue perfusion. 5 Again, the result is hypoxia, acidosis, and, if prolonged, irreversible shock. Based on the available physiologic data pertaining to hypovolemic shock, its treatment by maintenance of a blood pressure between 50 and 70 mm H g is dangerous at best and clearly unfounded. Dr. Crawford's patients comprise a highly selected group. Ninety percent of them are able to endure an ambulance ride to a helicopter followed by an air flight of variable duration. Many of the patients we, and doubtlessly the Harborview group, see are too sick and unstable to survive a first-class helicopter ride. They do, however, make it to our operating rooms. Dr. Crawford criticizes the knowledge, experience and technical ability of the Harborview group because his resuks contrast so strikingly with theirs. We reiterate that Dr. Crawford's results contrast strikingly,~fith those of the rest o f the nation. A comparison of his helicopter triage system to what is available to other skilled and competent vascular surgeons around the nation is unfair.
Letters to the Editors
457
While we praise the Harborview Group for their management of a difficult problem, we may also continue to attempt to emulate the standards set by Dr. Crawford. Steven G. Friedman, M D IC K I&ishnasastry, M2D
REFERENCES 1. Ouriel K, Geary K, Green RM, et al. Factors determining survival after ruptured aortic aneurysm: the hospital, the surgeon, and the patient, l Vasc SuR6 1990;11:493-6. 2. Shackleton CR, Schecter MT, Bianco R, et al. Preoperative predictors of mortality risk in ruptured abdominal aortic aneurysm. J Vasc SURG 1987;6:583-9. 3. Wakefield TW, Whitehouse WM, Wu SC, et al. Abdominal aortic aneurysm rupture: statistical analysis of factors affecting outcome of surgical treatment. Surgery 1982;91:586-96. 4. Shoemaker WC. Pathophysiology and therapy of shock syndromes. In: Shoemaker WC, Thompson VffL, Holbrook PR, eds. Textbook of critical care. 1~ted. Philadelphia: WB Saunders Co, i984;52-72. 5. Peityman A. Principles of circulatory support and the treatment of hemorrhagic shock. In: Snyder IV, Pursky MR, eds. Oxygen transport in the critically ill. I st ed. Chicago: Year Book Medical Publishers, Inc., 1987;407-18. 24/41/31837
Cells derived from omental fat tissue and used for seeding vascular prostheses are not endothelial in origin To the Editors:
A recent report by Visser et al. 1 presents evidence that cells isolated and cultured from human omentum represent a mixture of mesothelial and endothelial cells. Their subsequent conclusion that a major cell type in human omentum is rnesothelium is well substantiated by their own data as well as numerous previous reports by other authors on this subject. 2~ However, Visser et al. extend the interpretation of their data to claim a large number of investigators, including ourselves, have misinterpreted our own findings. We strongly disagree with many of t h e subsequent conclusions made in this manuscript and wish to draw attention to several misinterpretations and omissions. First, Dr. Visser uses the terms omentum, omental tissue, and omental-associated fat almost interchangeably. This is incorrect since these terms describe at least two morphologically distinct tissues. O m e n m m is an anatomic structure consisting of a serosal-covered surface enclosing larger vessels, many lymphatic structures, and adipose tissue. Mesothelial cells forming the serosal surface represent a large proportion of the cell number because of the high surface area-to-contained-volume ratio, and thus one would expect a predominant number of mesothelial cells from collagenase digestion. On the other hand, fat tissue is morphologically quite different from omentum and can be
458
Journal of VASCULAR SURGERY
Letters to the Editors
described as adipocytes perfused by a rich network of microvascular endothelial cells. This critical distinction has been previously discussed in a publication by us, 6 but a discussion of this distinction is not found in Dr. Visser's paper. Thus an important point not discussed by Dr. Visser is the importance of isolating fat free from surrounding mesothelial cell serosal capsule. When this is performed the sheets of cells described by Dr. Visser are not observed in the isolate, thus significantly reducing the number o f mesothelial cells present. Dr. Visser also omitted descriptions o f alternate sources of fat for microvessel endothelial cell isolation, which can be used for vascular graft endothelialization studies. We have previously reported the use o f subcutaneous fat for endothelial cell isolation, a tissue that is not surrounded by a mesothelial cell capsule. Subcutaneous fat has numerous advantages over the use of fat associated with the omentum since it can be obtained by liposuction. Fat occurs in numerous other sites in the body in morphologically distinct form. In conclusion Dr. Visser and colleagues, through significant omissions of published papers and misapplication of their own findings on omentum, have suggested that cells isolated and cultured from fat are mesothelium and not endothelium. Although the merits o f using omentum to derive cells for vascular graft treatments deserve continued study as proposed by Dr. Visser, most o f the conclusions concerning the identity of cells derived from fat drawn by Dr. Visser and colleagues are inappropriate. Stuart IC Williams, PhD Bruce E. Jarrell, ME) Department of Surgery University of Arizona Tucson, AZ 85724 REFERENCES
1. Visser MJT, van Bockel JH, van Muijen GNP, van Hindbergh VWM. Cells derived from omental fat tissue and used for seeding vascular prostheses are not endothelial in origin. J VAsc SURG 1991;13:378-81. 2. Potzsch B, Gmlich-Henn J, Rossing R, Wille D, MullerBerghans G. Identification of endothelial and mesothelial cells in human omental tissue and in omentum-derived culture cells by specific cell markers. Lab Invest 1990;63:841-52. 3. Stylianou E, Jenner LA, Davies M, Coles GA, Williams JD. Isolation culture and characterization of human peritoneal mesothelial cells. Kid Int 1990;37:1563-70. 4. van Hinsberg VWM, Kooistra T, Scheffer MA, van Bockel JH, van Muijen GNP. Characterization and fibrinolytic of human omental tissue mesothelial cells. Comparison with endothelial cells. Blood 1990;75:1490-7. 5. Hata J, Goto T, Mukai K, Sawasaki Y, Takahashi K. Cobblestone monolayer cells from human omental adipose tissue are possibly mesothelial not endothelial. In Vitro Cell Dev Biol 1989;25:109-11. 6. Williams SK, Jarrell BE, Rose DR, et al. Human microvessel endothelial cell isolation and vascular graft sodding in the operating room. Ann Vasc Surg 1989;3:146-52. 24/41/32075
Reply To the Editors." We appreciate the opportunity to respond to Drs. Williams' and JarreU's criticisms, which we strongly dispute. They raise an important issue that gives us the chance to stress that the nature o f cells derived from omental fat tissue needs to be properly evaluated. The point made by Drs. Williams and Jarrell that the terms omentum, omental tissue, and omental-associated fat were used almost interchangeably in our report is well taken. The greater omentum can be defined as an anatomic structure consisting o f a fourfold layer of serosal tissue. This anatomic structure contains adipose tissue in the adult human. In the reports we cited in our article either the term omentum 1 or omentum and omental tissue 2 were used to refer to the source of the presumed microvascular endothelial cells. Of course, these investi gators did not use the 'omentum' as 'serosal covere~ surfaces' as defined by Drs. Williams and Jarrell, because it is not logical to isolate endothelial cells from this material, which contains no microvessels. In their previous work, which was cited by us, 3-5 Drs. Williams and Jarrell introduce confusion by not precisely mentioning how the omental tissue was prepared. Particularly, they never mentioned that the serosal layers were separated from the actual fat tissue. There can, however, be no misunderstanding as to whether we used true omentum or omental fat as the source of the epitheloid cells we described. In the "Materials and Methods" section of our report the precise type of tissue used is defined in the second sentence: "The cells derived from omentum, which were presumed to be microvascular endothelial cells, were isolated from human omental f a t tissue." Drs. Williams and Jarrell state that no sheet structures are observed after collagenase digestion of isolated omental fat. They then mistakenly draw the conclusion that the number of mesothelial cells is significantly reduced. The absence of sheet structures is not a proof that the number o f mesothelial cells is reduced, because the presence of sheet structures depends on the applied isolation techniques like collagenase digestion o f omental tissue: type of collagenase, batch differences, concentration, and incubation time. Although the number of mesothelial cells contaminating the yield of microvascular endothelial cells might be reduced by isolation of omental fat tissue, free from serosal capsule, the adequate identification of the nature of the cells remains an important issue. The nature of the epitheloid cells that are isolated after collagenase digestion must certainly not be evaluated by the presence or absence of sheet structures neither must it be exclusively evaluated by immunofluorescence staining with polyclonal anti-yon Willebrand factor antibodies, as many investigators do. A more appropriate method to establish the nature o f these cells is to evaluate the presence of yon Willebrand factor antigens by mouoclonal antibodies and the presence or absence of cytokeratins 8 and 18. The second point Drs. Williams and Jarrell make is that we omitted descriptions o f alternate sources of fat for
