Enzyme and immunohistochemistry on undecalcified bone and bone marrow biopsies after embedding in plastic: A new embedding method for routine application

Wolf, Eduard; Röser, Kerstin; Hahn, Michael; Welkerling, Heike; Delling, G.

doi:10.1007/bf01605979

Cited by 88 publications

(56 citation statements)

References 19 publications

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“…Infiltrated specimens were placed into embedding molds, and polymerization was performed at Ϫ15°C. 12 Polymerized blocks were initially ground to bring the tissue components closer to the cutting surface. After the opposite side of the block was glued to a Plexiglas slide with Technovit 4000 (Kulzer & Co) in vacuum, a 1200-grit grinding in a micro parallel grinding system (EXAKT, Apparatebau) was used, followed by 4000-grit sandpaper grinding, which smoothed the final lower surface of the samples.…”

Section: Methodsmentioning

confidence: 99%

Angiogenesis, Vascular Endothelial Growth Factor and Platelet-Derived Growth Factor-BB Expression, Iron Deposition, and Oxidation-Specific Epitopes in Stented Human Coronary Arteries

Bräsen

Kivelä

Röser

et al. 2001

ATVB

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Abstract-Pathogenesis of in-stent restenosis remains poorly understood because information from human histopathologic studies is scarce. We used an improved saw-grinding and cutting method on methacrylate-embedded samples containing metal stents, which allows in situ hybridization and immunohistochemical analysis of in-stent restenosis. Twenty-one samples were collected 3 hours to 3 years after stenting from 6 patients aged 36 to 81 years. Except in very early samples collected within hours after the stent deployment, neovascularization was present in all segments studied. At advanced stages, extensive neovascularization was located mainly at the luminal side of the stent struts and was only rarely accompanied by inflammatory cells. The neovessels colocalized with vascular endothelial growth factor (VEGF)-A mRNA and protein expression as well as with iron deposits and oxidation-specific epitopes, which imply the presence of chronic oxidative stress. VEGF-A expression was detected in the same areas containing macrophages, endothelial cells, and, to a lesser extent, smooth muscle cells, which also showed platelet-derived growth factor-BB expression. We conclude that in-stent restenosis features neovascularization, VEGF-A and platelet-derived growth factor-BB expression, and iron deposition, which is most probably derived from microhemorrhages. These mechanisms may play an important role in the development of neointimal thickening and could provide useful targets for the prevention and treatment of in-stent restenosis. Key Words: in-stent-restenosis Ⅲ immunohistochemistry Ⅲ in situ hybridization Ⅲ pathology Ⅲ methylmethacrylate embedding P ercutaneous transluminal coronary angioplasty (PTCA) is the treatment of choice in short single coronary lesions. Acute complications, dissections, and early recoil can be treated or avoided with intracoronary stents, which also reduce the incidence of late restenosis. Nevertheless, angiographic restenosis is seen in 20% to 30% of ideal lesions treated with stents. 1,2 The angiographic restenosis rate with more complex lesions is even higher. 3 Treatment of in-stent restenosis by balloon dilation, rotablation, and coronary atherectomy leads to unsatisfactory results in at least 50% of the cases. 4 Thus, prevention of in-stent restenosis is an important goal. See coverIn-stent restenosis is thought to be a reparative process resembling wound healing, involving a cascade of traumatic, thrombotic, granulating, and proliferative phases as well as late remodeling, with final accumulation of smooth muscle cells and matrix components. 5,6 Glycoprotein IIb/IIIa antagonists aiming at blocking early platelet deposition/thrombus formation and the proliferation and migration of smooth muscle cells reduced but failed to fully inhibit in-stent restenosis. 7,8 The amount of medial damage and stent oversizing have been shown to be correlated with the degree of in-stent restenosis, suggesting that the extent of vessel trauma plays a crucial role in in-stent restenosis. 9,10 One of the major probl...

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Section: Methodsmentioning

confidence: 99%

Angiogenesis, Vascular Endothelial Growth Factor and Platelet-Derived Growth Factor-BB Expression, Iron Deposition, and Oxidation-Specific Epitopes in Stented Human Coronary Arteries

Bräsen

Kivelä

Röser

et al. 2001

ATVB

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“…Animals received daily aspirin (Ϸ40 mg, orally), beginning on Day Ϫ3 and continuing until euthanasia. On Day 1, animals received a single prophylactic injection of benzathine/procaine penicillin-G (42 000 IU/kg, IM) and were preanesthetized (ketamine HCl, 10 mg/kg, IM; atropine SO 4 , 0.04 mg/kg, IM), then intubated and maintained in anesthesia with isoflurane. Animals were euthanized at day 4 (nϭ2), day 7 (nϭ2), and day 14 (nϭ2) after injury.…”

Section: Surgical and Tissue Retrieval Techniquesmentioning

confidence: 99%

“…Stented segments were embedded in methyl methacrylate mixed with n-butyl methacrylate (Sigma Chemical Co) 4,5 to allow for sectioning through metal stent struts. Balloon-injured segments were paraffin embedded.…”

Section: Tissue Processingmentioning

confidence: 99%

Targeting CCR2 or CD18 Inhibits Experimental In-Stent Restenosis in Primates

Horvath

Welt

Nedelman

et al. 2002

Circulation Research

108

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Abstract-A central role for leukocytes in neointimal hyperplasia after arterial injury is suspected. However, the relative importance of neutrophils and monocytes in balloon or stent-induced injury are not well understood, and mechanistic targeting of leukocyte recruitment or function is crude. We determined the temporal and spatial distribution of different leukocytes after balloon and stent-induced injury in primate iliac arteries. Based on these data, we targeted neutrophil and monocyte recruitment selectively after angioplasty or stent implantation and demonstrated that monocyte-specific blockade achieved via blockade of the MCP-1 receptor CCR2, was effective at reducing neointimal hyperplasia after stenting. In contrast, combined neutrophil and monocyte blockade achieved by targeting the leukocyte ␤ 2 -integrin ␤-subunit CD18 was required to reduce neointimal hyperplasia after balloon injury.

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“…Erben nants are lost because of temperature peaks and modification of chemical structures in the tissue by reactive radicals formed during polymerization of MMA (Baskin et al 1992;Wolf et al 1992). It is also possible to initiate polymerization of methacrylates by blue or uv light photons.…”

mentioning

confidence: 99%

“…Recently, an MMA embedding technique has been reported that can be used for embedding of larger bone samples, and which preserves enzyme activities and antigenic determinants in bone tissue (Wolf et al 1992). This embedding mixture polymerizes at Ϫ 15 to Ϫ 20C.…”

mentioning

confidence: 99%

Embedding of Bone Samples in Methylmethacrylate: An Improved Method Suitable for Bone Histomorphometry, Histochemistry, and Immunohistochemistry

Erben

1997

J Histochem Cytochem.

292

242

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SUMMARY Methylmethacrylate (MMA) embedding of undecalcified bone biopsies is a technique widely used for bone histomorphometry. However, conventional MMA embedding causes almost complete loss of enzyme activity and protein antigenicity in the tissues. Recently, an MMA embedding technique has been reported that preserves enzyme activity and antigenic determinants in bone tissue. We describe here a modification of this embedding method. For our modified MMA embedding process, commercially available methacrylates can be used without purification, and the histologic quality of bone sections is comparable to that of conventionally MMA-embedded bone specimens. The technique reported here can be employed for embedding of larger bone samples and is suitable for histochemical and immunohistological applications as well as for routine bone histomorphometry. By addition of methylbenzoate during infiltration and polymerization of the plastic, the antigenicity of the tissue was improved. As applications of this novel technique, demonstration of alkaline phosphatase and tartrate-resistant acid phosphatase as well as positive labeling of Kupffer cells and osteoclasts with the monoclonal antibody ED1 in sections of liver, tibiae, and vertebrae of 3-month-old rats was demonstrated. The method described here might be useful for the inclusion of histochemical and immunohistological methods into bone histomorphometry. (J Histochem Cytochem 45:307-313, 1997)

show abstract

Enzyme and immunohistochemistry on undecalcified bone and bone marrow biopsies after embedding in plastic: A new embedding method for routine application

Cited by 88 publications

References 19 publications

Angiogenesis, Vascular Endothelial Growth Factor and Platelet-Derived Growth Factor-BB Expression, Iron Deposition, and Oxidation-Specific Epitopes in Stented Human Coronary Arteries

Angiogenesis, Vascular Endothelial Growth Factor and Platelet-Derived Growth Factor-BB Expression, Iron Deposition, and Oxidation-Specific Epitopes in Stented Human Coronary Arteries

Targeting CCR2 or CD18 Inhibits Experimental In-Stent Restenosis in Primates

Embedding of Bone Samples in Methylmethacrylate: An Improved Method Suitable for Bone Histomorphometry, Histochemistry, and Immunohistochemistry

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