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Lipoprotein(a) deposition is associated with the development of transplant renal arteriosclerosis in humans
METABOLIC CONSIDERATIONS
Lipoprotein(a) Deposition Is Associated With the Development of Transplant Renal Arteriosclerosis in Humans Y.-S. Han, M. Ueda, S. Tanabe, T. Nakatani, T. Kishimoto, S. Suzuki, H. Amemiya, and J. Kimura
T
RANSPLANT RENAL arteriosclerosis (TRA) is a major cause of chronic rejection in human kidney transplantation. We have previously investigated the cellular composition of TRA using immunohistochemical techniques, and demonstrated that smooth muscle cell migration and proliferation are a crucial event in the development of TRA in human renal allografts.1–3 Recently, we have also shown that extracellular matrix glycoproteins, such as fibronectin and tenascin, and endothelinconverting enzyme which is a key enzyme in the biosynthesis pathway of endothelin, may play important roles during the development of TRA in humans.2– 4 Lipoprotein(a) (Lp(a)) is a macromolecular complex in which increased concentration in plasma is correlated with the development of atherosclerosis.5,6 The characteristic protein component of Lp(a) is apolipoprotein(a) which is disulphide-linked to apolipoprotein B-100.7 Recent in vitro
study has demonstrated that Lp(a) promotes vascular smooth muscle cell proliferation.8 To our knowledge, however, the potential significance of Lp(a) during the development of TRA has not been reported. We therefore investigated, with the use of immunohistochemical techniques, the deposition of Lp(a) at sites of TRA lesions in human renal allografts.
From the Department of Urology (Y.-S.H., S.T., T.N., T.K.); Department of Pathology (M.U.), Osaka City University Medical School, Osaka, Japan; National Children’s Medical Research Center (S.S., H.A.), Tokyo, Japan; and Vessel Research Laboratory Co. Ltd. (J.K.), Kanagawa, Japan. Address reprint requests to Makiko Ueda, MD, Department of Pathology, Osaka City University Medical School, 1-4-54, Asahimachi, Abeno-ku, Osaka, Japan.
Table 1. Relevant Clinical Data Case No.
Age (Y)
Sex
Interval Transplant./Hemodialysis
Interval Transplant./Nephrectomy
1 2 3 4 5 6 7 8 9 10
37 43 40 37 27 17 49 41 43 42
M F M F F F F M M F
— — — 3m 3m 2m 1y 6m 2y 3m 2y 4m 3y
1m 2m 2m 4m 4m 5m 2y 7m 3y 3m 3y 4m 3y 6m
© 1998 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
0041-1345/98/$19.00 PII S0041-1345(98)01289-5
Transplantation Proceedings, 30, 3017–3020 (1998)
3017
3018
HAN, UEDA, TANABE ET AL
Fig 1. Micrographs of an intrarenal artery with TRA, 1 month after transplantation. (A) Hematoxylin-eosin stain. (B) Staining with anti-macrophage antibody, HAM56. (C) Staining with anti-muscle actin antibody, HHF35. (D) Staining with anti-human Lp(a) antibody, 1D1.
MATERIALS AND METHODS In this study, 10 renal allografts that had been removed due to rejection were examined. Intervals between transplantation and removal of the allografts in these patients ranged from 1 month to 3 years. Table 1 gives the relevant clinical data of the patients. Intrarenal arteries in ten normal kidneys were also examined as controls. All specimens were fixed in methanol-Carnoy’s fixative, and tissue blocks were obtained and embedded in paraffin. Thirty serial sections, 5 mm thick, were cut from each block. Every first, second, and third section was stained with hematoxylin and eosin, Weigert’s elastic van Gieson’s, and periodic-acid Schiff stain, respectively. The other sections were used for immunohistochemical staining. The monoclonal antibodies used were as follows: anti-human Lp(a) antibody, 1D1; anti-macrophage antibody, HAM56; anti-T cell antibody, UCHL1; anti-B cell antibody, L26; anti-vimentin antibody; anti-muscle actin antibody, HHF35; and anti-smooth muscle actin antibody, CGA7. The specificity of the anti-human Lp(a) antibody, 1D1 has been reported previously.9 The labeled streptavidin-biotin complex system with nickle chloride color modification was performed in all instances. Sections were counterstained with methyl green.
RESULTS
In normal kidneys, no Lp(a) deposition was found in intrarenal arteries.
In all renal allografts, TRA characterized by concentric intimal thickening was observed, frequently in the interlobular and arcuate arteries. In the earliest stage of TRA, 1 month after transplantation, the neointima was composed of macrophages, T cells, and smooth muscle cells (Fig 1A–C). At this stage, marked deposition of Lp(a) was observed in the media and neointima (Fig 1D). In two cases, 2 months after transplantation, the neointima consisted of macrophages and smooth muscle cells (Fig 2A–C). At this stage, the neointima showed distinct Lp(a) deposition, while the media revealed no or only weak Lp(a) deposition (Fig 2D). In older stages of TRA, from 4 months onward, the neointima was composed almost entirely of smooth muscle cells (Fig 3A–C). At these stages, no or little Lp(a) deposition was observed in the media and neointima (Fig 3D). DISCUSSION
TRA is known to be a major limitation to the long-term survival of human renal allografts. Our previous studies have shown that smooth muscle cell proliferation and migration, associated with phenotypic modulation of smooth muscle cells to the de-differentiation state, are an essential event at early stages after transplantation in the lesions with TRA.2– 4
Fig 2. Micrographs of an intrarenal artery with TRA, 2 months after transplantation. (A) Hematoxylin-eosin stain. (B) Staining with anti-macrophage antibody, HAM56. (C) Staining with anti-muscle actin antibody, HHF35. (D) Staining with anti-human Lp(a) antibody, 1D1.
Fig 3. Micrographs of an intrarenal artery with TRA, 3 years and 4 months after transplantation. (A) Hematoxylin-eosin stain. (B) Staining with anti-macrophage antibody, HAM56. (C) Staining with anti-muscle actin antibody, HHF35. (D) Staining with anti-human Lp(a) antibody, 1D1.
3020
The present study documents that, at the early stages, within 2 months after transplantation, distinct Lp(a) deposition occurred in the preexistent media and young neointima. However, at later stages from 4 months onward, Lp(a) deposition was no longer found in the media and young neointima. These findings strongly suggest that Lp(a) deposition in the media and young neointima at early stages after transplantation is closely related with the development of neointima in human renal allografts. Recent experimental studies have demonstrated that Lp(a) stimulates the proliferation of human smooth muscle cells in culture,8 and increases the amounts of smooth muscle cell migration by inhibition of TGF-b activation.10 These in vitro studies strongly suggest that Lp(a) may contribute to the development of neointimal lesions by promoting the proliferation and migration of vascular smooth muscle cells. Our observations in humans, therefore, are keeping with these experimental data and seem to indicate that Lp(a) could play an important role as one of the mediators in the growth of neointima in human renal allografts. The present study thus provides data that Lp(a) deposition contributes to the formation of neointimal lesions at
HAN, UEDA, TANABE ET AL
early stages after transplantation in humans. These findings add new information to the nature of the cellular mechanisms that lead to the development of TRA in humans.
REFERENCES 1. Han Y-S, Ueda M, Tanabe S, et al: Transplant Proc 26:931, 1994 2. Tanabe S, Ueda M, Han Y-S, et al: Transplant Proc 27:1078, 1995 3. Tanabe S, Ueda M, Han Y-S, et al: Transpl Int 9:S45, 1996 4. Tanabe S, Ueda M, Han Y-S, et al: Transplant Proc 29:1517, 1997 5. Genest JJ, Martin-Munley SS, McNamara JR, et al: Circulation 85:2025, 1992 6. Schreiner PJ, Morrisett JD, Sharrett AR, et al: Arterioscler Thromb 13:826, 1993 7. Utermann G: Science 246:904, 1989 8. Grainger DJ, Kirschenlohr HL, Metcalfe JC, et al: Science 260:1655, 1993 9. Lafferty MA, Salamon AM, Usher DC: J Lipid Res 32:277, 1991 10. Kojima S, Harpel PC, Rifkin DB: J Cell Biol 113:1439, 1991
Lipoprotein(a) Deposition Is Associated With the Development of Transplant Renal Arteriosclerosis in Humans Y.-S. Han, M. Ueda, S. Tanabe, T. Nakatani, T. Kishimoto, S. Suzuki, H. Amemiya, and J. Kimura
T
RANSPLANT RENAL arteriosclerosis (TRA) is a major cause of chronic rejection in human kidney transplantation. We have previously investigated the cellular composition of TRA using immunohistochemical techniques, and demonstrated that smooth muscle cell migration and proliferation are a crucial event in the development of TRA in human renal allografts.1–3 Recently, we have also shown that extracellular matrix glycoproteins, such as fibronectin and tenascin, and endothelinconverting enzyme which is a key enzyme in the biosynthesis pathway of endothelin, may play important roles during the development of TRA in humans.2– 4 Lipoprotein(a) (Lp(a)) is a macromolecular complex in which increased concentration in plasma is correlated with the development of atherosclerosis.5,6 The characteristic protein component of Lp(a) is apolipoprotein(a) which is disulphide-linked to apolipoprotein B-100.7 Recent in vitro
study has demonstrated that Lp(a) promotes vascular smooth muscle cell proliferation.8 To our knowledge, however, the potential significance of Lp(a) during the development of TRA has not been reported. We therefore investigated, with the use of immunohistochemical techniques, the deposition of Lp(a) at sites of TRA lesions in human renal allografts.
From the Department of Urology (Y.-S.H., S.T., T.N., T.K.); Department of Pathology (M.U.), Osaka City University Medical School, Osaka, Japan; National Children’s Medical Research Center (S.S., H.A.), Tokyo, Japan; and Vessel Research Laboratory Co. Ltd. (J.K.), Kanagawa, Japan. Address reprint requests to Makiko Ueda, MD, Department of Pathology, Osaka City University Medical School, 1-4-54, Asahimachi, Abeno-ku, Osaka, Japan.
Table 1. Relevant Clinical Data Case No.
Age (Y)
Sex
Interval Transplant./Hemodialysis
Interval Transplant./Nephrectomy
1 2 3 4 5 6 7 8 9 10
37 43 40 37 27 17 49 41 43 42
M F M F F F F M M F
— — — 3m 3m 2m 1y 6m 2y 3m 2y 4m 3y
1m 2m 2m 4m 4m 5m 2y 7m 3y 3m 3y 4m 3y 6m
© 1998 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
0041-1345/98/$19.00 PII S0041-1345(98)01289-5
Transplantation Proceedings, 30, 3017–3020 (1998)
3017
3018
HAN, UEDA, TANABE ET AL
Fig 1. Micrographs of an intrarenal artery with TRA, 1 month after transplantation. (A) Hematoxylin-eosin stain. (B) Staining with anti-macrophage antibody, HAM56. (C) Staining with anti-muscle actin antibody, HHF35. (D) Staining with anti-human Lp(a) antibody, 1D1.
MATERIALS AND METHODS In this study, 10 renal allografts that had been removed due to rejection were examined. Intervals between transplantation and removal of the allografts in these patients ranged from 1 month to 3 years. Table 1 gives the relevant clinical data of the patients. Intrarenal arteries in ten normal kidneys were also examined as controls. All specimens were fixed in methanol-Carnoy’s fixative, and tissue blocks were obtained and embedded in paraffin. Thirty serial sections, 5 mm thick, were cut from each block. Every first, second, and third section was stained with hematoxylin and eosin, Weigert’s elastic van Gieson’s, and periodic-acid Schiff stain, respectively. The other sections were used for immunohistochemical staining. The monoclonal antibodies used were as follows: anti-human Lp(a) antibody, 1D1; anti-macrophage antibody, HAM56; anti-T cell antibody, UCHL1; anti-B cell antibody, L26; anti-vimentin antibody; anti-muscle actin antibody, HHF35; and anti-smooth muscle actin antibody, CGA7. The specificity of the anti-human Lp(a) antibody, 1D1 has been reported previously.9 The labeled streptavidin-biotin complex system with nickle chloride color modification was performed in all instances. Sections were counterstained with methyl green.
RESULTS
In normal kidneys, no Lp(a) deposition was found in intrarenal arteries.
In all renal allografts, TRA characterized by concentric intimal thickening was observed, frequently in the interlobular and arcuate arteries. In the earliest stage of TRA, 1 month after transplantation, the neointima was composed of macrophages, T cells, and smooth muscle cells (Fig 1A–C). At this stage, marked deposition of Lp(a) was observed in the media and neointima (Fig 1D). In two cases, 2 months after transplantation, the neointima consisted of macrophages and smooth muscle cells (Fig 2A–C). At this stage, the neointima showed distinct Lp(a) deposition, while the media revealed no or only weak Lp(a) deposition (Fig 2D). In older stages of TRA, from 4 months onward, the neointima was composed almost entirely of smooth muscle cells (Fig 3A–C). At these stages, no or little Lp(a) deposition was observed in the media and neointima (Fig 3D). DISCUSSION
TRA is known to be a major limitation to the long-term survival of human renal allografts. Our previous studies have shown that smooth muscle cell proliferation and migration, associated with phenotypic modulation of smooth muscle cells to the de-differentiation state, are an essential event at early stages after transplantation in the lesions with TRA.2– 4
Fig 2. Micrographs of an intrarenal artery with TRA, 2 months after transplantation. (A) Hematoxylin-eosin stain. (B) Staining with anti-macrophage antibody, HAM56. (C) Staining with anti-muscle actin antibody, HHF35. (D) Staining with anti-human Lp(a) antibody, 1D1.
Fig 3. Micrographs of an intrarenal artery with TRA, 3 years and 4 months after transplantation. (A) Hematoxylin-eosin stain. (B) Staining with anti-macrophage antibody, HAM56. (C) Staining with anti-muscle actin antibody, HHF35. (D) Staining with anti-human Lp(a) antibody, 1D1.
3020
The present study documents that, at the early stages, within 2 months after transplantation, distinct Lp(a) deposition occurred in the preexistent media and young neointima. However, at later stages from 4 months onward, Lp(a) deposition was no longer found in the media and young neointima. These findings strongly suggest that Lp(a) deposition in the media and young neointima at early stages after transplantation is closely related with the development of neointima in human renal allografts. Recent experimental studies have demonstrated that Lp(a) stimulates the proliferation of human smooth muscle cells in culture,8 and increases the amounts of smooth muscle cell migration by inhibition of TGF-b activation.10 These in vitro studies strongly suggest that Lp(a) may contribute to the development of neointimal lesions by promoting the proliferation and migration of vascular smooth muscle cells. Our observations in humans, therefore, are keeping with these experimental data and seem to indicate that Lp(a) could play an important role as one of the mediators in the growth of neointima in human renal allografts. The present study thus provides data that Lp(a) deposition contributes to the formation of neointimal lesions at
HAN, UEDA, TANABE ET AL
early stages after transplantation in humans. These findings add new information to the nature of the cellular mechanisms that lead to the development of TRA in humans.
REFERENCES 1. Han Y-S, Ueda M, Tanabe S, et al: Transplant Proc 26:931, 1994 2. Tanabe S, Ueda M, Han Y-S, et al: Transplant Proc 27:1078, 1995 3. Tanabe S, Ueda M, Han Y-S, et al: Transpl Int 9:S45, 1996 4. Tanabe S, Ueda M, Han Y-S, et al: Transplant Proc 29:1517, 1997 5. Genest JJ, Martin-Munley SS, McNamara JR, et al: Circulation 85:2025, 1992 6. Schreiner PJ, Morrisett JD, Sharrett AR, et al: Arterioscler Thromb 13:826, 1993 7. Utermann G: Science 246:904, 1989 8. Grainger DJ, Kirschenlohr HL, Metcalfe JC, et al: Science 260:1655, 1993 9. Lafferty MA, Salamon AM, Usher DC: J Lipid Res 32:277, 1991 10. Kojima S, Harpel PC, Rifkin DB: J Cell Biol 113:1439, 1991