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David W. Boykin - One of the best experts on this subject based on the ideXlab platform.
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196 Prevention and treatment of bovine viral diarrhea virus infections in Fetal Fibroblast cells
Reproduction Fertility and Development, 2004Co-Authors: Givens, David A. Stringfellow, Kay P. Riddell, Patricia K. Galik, Eleanor J. Sullivan, James M. Robl, P. Kasinathan, Christine C. Dykstra, David W. BoykinAbstract:Unnoticed infections with bovine viral diarrhea virus (BVDV) can occur in cultured cells used for somatic cell nuclear transfer. Aromatic cationic molecules have exhibited inhibitory activity against in vitro replication of BVDV. The purpose of this research was to evaluate the ability of aromatic cationic compounds to prevent or treat noncytopathic BVDV infections of Fetal Fibroblast cells. Aromatic compounds tested were 2-(4-[2-imidazolinyl]phenyl)-5-(4-methoxyphenyl)furan (DB606); 2-(2-benzimidazolyl)-5-[4-(2-imidazolino) phenyl]furan dihydrochloride (DB772); and 2-(1-methyl-2-benzimidazolyl)-5-[4’-(2-imidazolino)-2’-methylphenyl]furan dihydrochloride (DB824). To evaluate prevention of BVDV infections, 10 cell lines in the absence or presence of 7 dilutions of each of the 3 compounds were inoculated with BVDV. The concentrations of BVDV in medium and cell lysates were determined by serial dilution and virus isolation. Samples were obtained 72 hours post-inoculation. Bovine viral diarrhea virus in cell culture medium and cell lysate samples was evaluated by comparison to equivalent samples from control cultures in which no compound was added (percent of control = cell culture infective doses (50%; CCID50) of BVDV in compound sample/CCID50 of BVDV in control sample lacking compound). The viral inhibitory concentrations (99%) of compounds were calculated with JMP software by least-squares regression techniques. Cumulatively, the 99% endpoints for inhibition of viral replication in Fetal Fibroblast cell lines for the 3 compounds were 0.1 μM, 0.007 μM and 0.028 μM, respectively. To evaluate therapeutic treatment of established BVDV infections, the concentration of BVDV in medium and cell lysates of 2 Fetal Fibroblast cell lines were evaluated. The cell lines were previously determined to be infected with a genotype 1a strain of BVDV. Samples were obtained during 4 sequential passages in the absence or presence of 0.04 μM and 4 μM concentrations of DB772 or DB824. Presence of BVDV was determined by reverse transcription nested polymerase chain reaction and virus isolation. While BVDV persisted in cultures supplemented with no aromatic compound or 0.04 μM, both DB772 and DB824 effectively cured BVDV infections after 1 passage in 4 μM, and cells remained viable. Results indicate that BVDV infections can be effectively prevented or treated in Fetal Fibroblast cultures. Further research is needed to determine if exposed cells are competent for production of normal embryos via nuclear transfer.
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Prevention and elimination of bovine viral diarrhea virus infections in Fetal Fibroblast cells
Antiviral research, 2004Co-Authors: M. Daniel Givens, David A. Stringfellow, Kay P. Riddell, Patricia K. Galik, Eleanor J. Sullivan, James M. Robl, P. Kasinathan, Christine C. Dykstra, Arvind Kumar, David W. BoykinAbstract:Abstract Noncytopathic infections with bovine viral diarrhea virus (BVDV) can compromise research and commercial use of cultured cells. The purpose of this research was to evaluate the ability of aromatic cationic compounds to prevent or treat BVDV infections in Fetal Fibroblast cell lines that are used in somatic cell nuclear transfer. To evaluate preventative use of compounds, 10 cell lines were inoculated with BVDV in the absence or presence of 2-(4-[2-imidazolinyl]phenyl)-5-(4-methoxyphenyl)furan (DB606), 2-(2-benzimidazolyl)-5-[4-(2-imidazolino)phenyl]furan dihydrochloride (DB772), or 2-(1-methyl-2-benzimidazolyl)-5-[4′-(2-imidazolino)-2′-methylphenyl]furan dihydrochloride (DB824). The 99% endpoints for prevention of viral replication by these treatments were 81, 6, and 14 nM. To evaluate therapeutic use of compounds, two Fetal Fibroblast cell lines infected with a genotype 1a strain of BVDV were cultured through four passages in the absence or presence of either 0.04 or 4 μM concentrations of DB772 or DB824. The presence and concentration of BVDV in media and cell lysates were evaluated using reverse transcription nested polymerase chain reaction and virus isolation from titrated sample. A single passage in 4 μM of either compound was sufficient to eliminate BVDV from cells without causing cytotoxicity. Our results demonstrate that in vitro infections with BVDV can be effectively prevented or eliminated by addition of aromatic cations.
Richard Y. Lin - One of the best experts on this subject based on the ideXlab platform.
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the role of the Fetal Fibroblast and transforming growth factor beta in a model of human Fetal wound repair
Seminars in Pediatric Surgery, 1996Co-Authors: Richard Y. Lin, N S AdzickAbstract:The fetus heals skin wounds without scar formation. Human Fetal skin that is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded heals without scar formation, whereas the same Fetal skin heals with scar formation when transplanted to a cutaneous location. To characterize the healing process of human Fetal skin in these two locations, species-specific human and mouse DNA probes were constructed and used to probe graft wounds under high stringency in situ hybridization conditions. Immunostaining for species-specific Fibroblasts, macrophages, and neutrophils was also performed. The cutaneous human Fetal graft healed with scar and showed an influx of adult mouse Fibroblasts and macrophages. In contrast, subcutaneous human Fetal grafts showed exclusively human Fetal Fibroblasts in the wound environment, an absence of inflammatory cells, and scar-free repair. Thus, the highly organized collagen deposition in scarless human Fetal wound repair appears to be intrinsic to the human Fetal Fibroblast and occurs in the absence of an adult-like inflammatory response. One important cytokine involved in wound repair is transforming growth factor-beta (TGF beta). TGF beta promotes inflammatory cell recruitment and collagen deposition in healing wounds. To determine the role of TGF beta 1 in Fetal wound healing, in situ hybridization for TGF beta 1 mRNA expression, immunostaining for TGF beta protein, and species-specific immunohistochemistry for Fibroblasts, macrophages, and neutrophils were performed in human adult wounds, Fetal wounds, and Fetal wounds treated with a TGF beta 1 slow release disk. TGF beta 1 RNA expression and TGF beta protein were found in wounded adult skin. Neither TGF beta 1 mRNA upregulation nor TGF beta protein was detected in human Fetal skin after wounding. However, when exogenous TGF beta 1 was added to human Fetal skin, induction of TGF beta 1 mRNA expression in human Fetal Fibroblasts occurred, an adult-like inflammatory response was detected, and the skin healed with scar formation. Thus, non-scarring Fetal skin is relatively TGF beta 1 deficient when compared to scarring adult skin. Furthermore, when exogenous TGF beta is added to Fetal skin, scarring occurs. TGF beta appears to be an important modulator in scar formation. With a more thorough understanding of scarless repair mechanisms, clinical induction of scarless healing in adults and children soon may be possible.
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The Fetal Fibroblast: The Effector Cell of Scarless Fetal Skin Repair
Plastic and reconstructive surgery, 1995Co-Authors: H P Lorenz, Michael T. Longaker, Richard Y. Lin, David J. WhitbyAbstract:Human Fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human Fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (Fetal) Fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations: cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human Fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal Fibroblasts can heal Fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the Fetal Fibroblast is the major effector cell for scarless Fetal skin repair.
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The Fetal Fibroblast : the effector cell of scarless Fetal skin repair. Discussion
Plastic and Reconstructive Surgery, 1995Co-Authors: H P Lorenz, Michael T. Longaker, Richard Y. Lin, David J. Whitby, P. MartinAbstract:Human Fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human Fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (Fetal) Fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations : cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human Fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal Fibroblasts can heal Fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the Fetal Fibroblast is the major effector cell for scarless Fetal skin repair.
David J. Whitby - One of the best experts on this subject based on the ideXlab platform.
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The Fetal Fibroblast: The Effector Cell of Scarless Fetal Skin Repair
Plastic and reconstructive surgery, 1995Co-Authors: H P Lorenz, Michael T. Longaker, Richard Y. Lin, David J. WhitbyAbstract:Human Fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human Fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (Fetal) Fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations: cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human Fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal Fibroblasts can heal Fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the Fetal Fibroblast is the major effector cell for scarless Fetal skin repair.
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The Fetal Fibroblast : the effector cell of scarless Fetal skin repair. Discussion
Plastic and Reconstructive Surgery, 1995Co-Authors: H P Lorenz, Michael T. Longaker, Richard Y. Lin, David J. Whitby, P. MartinAbstract:Human Fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human Fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (Fetal) Fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations : cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human Fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal Fibroblasts can heal Fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the Fetal Fibroblast is the major effector cell for scarless Fetal skin repair.
H P Lorenz - One of the best experts on this subject based on the ideXlab platform.
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The Fetal Fibroblast: The Effector Cell of Scarless Fetal Skin Repair
Plastic and reconstructive surgery, 1995Co-Authors: H P Lorenz, Michael T. Longaker, Richard Y. Lin, David J. WhitbyAbstract:Human Fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human Fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (Fetal) Fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations: cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human Fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal Fibroblasts can heal Fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the Fetal Fibroblast is the major effector cell for scarless Fetal skin repair.
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The Fetal Fibroblast : the effector cell of scarless Fetal skin repair. Discussion
Plastic and Reconstructive Surgery, 1995Co-Authors: H P Lorenz, Michael T. Longaker, Richard Y. Lin, David J. Whitby, P. MartinAbstract:Human Fetal skin heals without scar formation when it is transplanted to a subcutaneous location on an adult athymic mouse and subsequently wounded. In contrast, human Fetal skin of identical gestational age heals with scar formation when transplanted to a cutaneous location on the athymic mouse recipient. To determine if mouse (adult) or human (Fetal) Fibroblasts are healing the graft wounds, we performed indirect immunohistochemistry for mouse and human collagen types I and III. Full-thickness skin grafts (n = 51) from human fetuses at 18 weeks' (n = 4) or 24 weeks' (n = 2) gestational age were placed onto athymic mice in two locations : cutaneously onto a fascial bed and subcutaneously in a pocket under the murine panniculus carnosus. Linear incisions were made in each graft 7 days after transplantation. Grafts were harvested at 7, 14, and 21 days after wounding and stained with hematoxylin and eosin or Mallory's trichrome. Immunohistochemistry for either human collagen type I or type III or for mouse collagen type I was performed. The subcutaneous grafts healed with human collagen types I and III in a scarless pattern. The wound collagen pattern was reticular and unrecognizable from the surrounding dermis. Hair follicles and sebaceous gland patterns were unchanged in the wounded dermis. Conversely, the cutaneous grafts healed with mouse collagen in a scar pattern with disorganized collagen fibers and no appendages. Mouse collagen scar was present along the base of the cutaneous grafts and as a thin capsule around the subcutaneous grafts. We conclude that (1) subcutaneous grafts heal with human Fetal collagen and no scar formation, and (2) cutaneous grafts heal with mouse collagen in a scar pattern. Fetal Fibroblasts can heal Fetal skin wounds without scar despite being perfused by adult serum and inflammatory cells in an adult environment. These data suggest that the Fetal Fibroblast is the major effector cell for scarless Fetal skin repair.
David A. Stringfellow - One of the best experts on this subject based on the ideXlab platform.
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196 Prevention and treatment of bovine viral diarrhea virus infections in Fetal Fibroblast cells
Reproduction Fertility and Development, 2004Co-Authors: Givens, David A. Stringfellow, Kay P. Riddell, Patricia K. Galik, Eleanor J. Sullivan, James M. Robl, P. Kasinathan, Christine C. Dykstra, David W. BoykinAbstract:Unnoticed infections with bovine viral diarrhea virus (BVDV) can occur in cultured cells used for somatic cell nuclear transfer. Aromatic cationic molecules have exhibited inhibitory activity against in vitro replication of BVDV. The purpose of this research was to evaluate the ability of aromatic cationic compounds to prevent or treat noncytopathic BVDV infections of Fetal Fibroblast cells. Aromatic compounds tested were 2-(4-[2-imidazolinyl]phenyl)-5-(4-methoxyphenyl)furan (DB606); 2-(2-benzimidazolyl)-5-[4-(2-imidazolino) phenyl]furan dihydrochloride (DB772); and 2-(1-methyl-2-benzimidazolyl)-5-[4’-(2-imidazolino)-2’-methylphenyl]furan dihydrochloride (DB824). To evaluate prevention of BVDV infections, 10 cell lines in the absence or presence of 7 dilutions of each of the 3 compounds were inoculated with BVDV. The concentrations of BVDV in medium and cell lysates were determined by serial dilution and virus isolation. Samples were obtained 72 hours post-inoculation. Bovine viral diarrhea virus in cell culture medium and cell lysate samples was evaluated by comparison to equivalent samples from control cultures in which no compound was added (percent of control = cell culture infective doses (50%; CCID50) of BVDV in compound sample/CCID50 of BVDV in control sample lacking compound). The viral inhibitory concentrations (99%) of compounds were calculated with JMP software by least-squares regression techniques. Cumulatively, the 99% endpoints for inhibition of viral replication in Fetal Fibroblast cell lines for the 3 compounds were 0.1 μM, 0.007 μM and 0.028 μM, respectively. To evaluate therapeutic treatment of established BVDV infections, the concentration of BVDV in medium and cell lysates of 2 Fetal Fibroblast cell lines were evaluated. The cell lines were previously determined to be infected with a genotype 1a strain of BVDV. Samples were obtained during 4 sequential passages in the absence or presence of 0.04 μM and 4 μM concentrations of DB772 or DB824. Presence of BVDV was determined by reverse transcription nested polymerase chain reaction and virus isolation. While BVDV persisted in cultures supplemented with no aromatic compound or 0.04 μM, both DB772 and DB824 effectively cured BVDV infections after 1 passage in 4 μM, and cells remained viable. Results indicate that BVDV infections can be effectively prevented or treated in Fetal Fibroblast cultures. Further research is needed to determine if exposed cells are competent for production of normal embryos via nuclear transfer.
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Prevention and elimination of bovine viral diarrhea virus infections in Fetal Fibroblast cells
Antiviral research, 2004Co-Authors: M. Daniel Givens, David A. Stringfellow, Kay P. Riddell, Patricia K. Galik, Eleanor J. Sullivan, James M. Robl, P. Kasinathan, Christine C. Dykstra, Arvind Kumar, David W. BoykinAbstract:Abstract Noncytopathic infections with bovine viral diarrhea virus (BVDV) can compromise research and commercial use of cultured cells. The purpose of this research was to evaluate the ability of aromatic cationic compounds to prevent or treat BVDV infections in Fetal Fibroblast cell lines that are used in somatic cell nuclear transfer. To evaluate preventative use of compounds, 10 cell lines were inoculated with BVDV in the absence or presence of 2-(4-[2-imidazolinyl]phenyl)-5-(4-methoxyphenyl)furan (DB606), 2-(2-benzimidazolyl)-5-[4-(2-imidazolino)phenyl]furan dihydrochloride (DB772), or 2-(1-methyl-2-benzimidazolyl)-5-[4′-(2-imidazolino)-2′-methylphenyl]furan dihydrochloride (DB824). The 99% endpoints for prevention of viral replication by these treatments were 81, 6, and 14 nM. To evaluate therapeutic use of compounds, two Fetal Fibroblast cell lines infected with a genotype 1a strain of BVDV were cultured through four passages in the absence or presence of either 0.04 or 4 μM concentrations of DB772 or DB824. The presence and concentration of BVDV in media and cell lysates were evaluated using reverse transcription nested polymerase chain reaction and virus isolation from titrated sample. A single passage in 4 μM of either compound was sufficient to eliminate BVDV from cells without causing cytotoxicity. Our results demonstrate that in vitro infections with BVDV can be effectively prevented or eliminated by addition of aromatic cations.