COMMENTARY
THE ANATOMICAL RECORD 296:724–725 (2013)
New Approach of Corrosion Casting Using Direct Injection of Mercox Into the Parenchyma of Different Organs MIROSLAV STEFANOV,1,2 JUNG-DAE KIM,2,3* MIN-HO NAM,4 AND KWANG-SUP SOH2 1 Department of Animal Morphology, Physiology and Nutrition, Agricultural Faculty, Trakia University, 6000 Stara Zagora, Bulgaria 2 Nano Primo Research Center, Advanced Institute of Convergence Technology, Seoul National University, Suwon, Republic of Korea 3 Pharmacopuncture Medical Research Center, Korean Pharmacopuncture Institute, Seoul, Republic of Korea 4 Department of Pathology, College of Korean Medicine, Kyung-Hee University, Seoul, Republic of Korea
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Methacrylate resins, such as commercial MercoxV, have been used to visualize vascular and lymphatic vessels and structures of different body cavities and lumens. The routine protocol requires full maceration with subsequent evaluation of the casts by stereomicroscopy and scanning electron microscopy (SEM) (Kobayashi et al., 1976; Lametschwandtner et al., 1984, 1990; Konerding, 1991; Aharinejad and Lametschwandtner, 1992; MartinOrti et al., 1999; Giuvarasteanu, 2007; Verli et al., 2007). Because of the full maceration, the observations are only on the vascular system and cavity surfaces without their relation with the surrounding tissues. The purpose of this report is to present a completely new approach using corrosion casting materials with partial preservation of the surrounding tissues in order to show the relation between all structures of the organs or tissues of interest. Five female ICR mice aged 8–10 weeks old were purchased from the Dooyeol-Biotech, Inc. (Seoul, Korea). The mice were euthanized by an overdose of urethane. Procedures involving animals and their care were compliant with institutional guidelines and current policies. The experiments were conducted within one or two days after obtaining the mice. Mercox was injected into the parenchyma of organs that included liver, lung, brain, heart, spleen, or directly into the skin. The injecting points are: left lobule of the liver, left lung, left wall of the heart, right hemisphere of the brain after making a hole into the skull lateral to the bregma, and hypodermal layer of the dorsal skin, 1–2 mm lateral to the vertebral column. Red and blue Mercox (Acrylic polymer in primary form) and its catalyst were obtained from Ladd Research Industries, USA. One-half milliliters of Mercox of each color and 0.02 mg catalyst were mixed immediately prior to the injections.
Disposable syringes of 1 mL volume with the disposable needle (30G31/200 , Sungshin Medical Co., Korea) were used for injection of 0.1 mL Mercox to each organ. The Mercox was slowly injected over a period of about 2 min. Thirty minutes after injection, the organs were placed in a solution for partial maceration. For this, the organs were partially macerated for 24 hr in separated petri dishes with 1% potassium hydroxide (KOH, Duksan Pure Chemicals, Korea) solution in order to see the distribution of the Mercox inside the organs while partially preserving the surrounding tissues. Microscopic observations were conducted using a stereomicroscope (SZX12, Olympus, Japan), optical microscope (BX51, Olympus, Japan), and polarizing microscope (KSM-BA3, Samwon, Korea). The new approach could be continued with routine histology protocol or with full maceration with 3% KOH and routine SEM procedures. After direct injection of the polymer Mercox into the parenchyma of the different organs, the structures in these organs were observed. In the spleen and lung, the vasculature could be easily observed. In the liver, the vascular bed and gallbladder channels could be observed. Likewise, the heart wall and pericardial vessels would be observed without filling the atrium and ventricular spaces. The brain cavities and arachnoidal space of the spinal cord could be visualized. In the skin, fat cells made a longitudinal trace along the spinal vertebra and the fat cells were apparent under high magnification (Fig. 1). The different distributions of the Mercox into the tissue structures of the organs need a special analysis and interpretation. The most surprising observation was the filling of the adipocytes in the skin (Fig. 1K,L). We can suppose that it is possible because of Mercox dilution in lipids. The
Grant sponsor: Traditional Korean Medicine R&D Project, Ministry of Health and Welfare, Korea; Grant number: B110076. *Correspondence to: J. Kim, PhD, Nano Primo Research Center, Advanced Institute of Convergence Technology, Seoul National University, Building-B, 7th floor, 864-1, Iui-dong,
Yeongtong-gu, Suwon-si, Gyeonggi-do, Republic of Korea. Fax: 182-31-888–9382. E-mail:
[email protected] Received 23 September 2012; Accepted 22 January 2013. DOI 10.1002/ar.22679 Published online 27 February 2013 in Wiley Online Library (wileyonlinelibrary.com).
C 2013 WILEY PERIODICALS, INC. V
NEW APPROACH OF CORROSION CASTING METHOD
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Fig. 1. Visualization of the distribution of the Mercox into different organ structures after partial maceration. (A) Heart, HW: heart wall, (B) heart, HC: heart cavity, (C) heart, PV: pericardial vessels, (D1) liver, GB: gall bladder, LP: liver parenchyma, C: channel, (D2) liver vascular bed, (D3) magnified view of the liver vessels, (E) lung, LV: lung vessel, (F) brain, AC: cerebral aqueduct, BC: brain cavities (arrows), S: skull,
(G) cross-section of spinal cord, SP: spinal process, VB: vertebral body, (H) spleen, (I) skin, IP: injection point, HL: hypodermal layer, (J) fat cell lines formed after full maceration of the skin, HL: hypodermal layer, SP: spinal process, TP: transversal process, (K) protrusion of fat cell line in the skin of hypodermal layer, (L) higher magnification of the fat cells of the skin, F: fat cells.
penetration of Mercox into fat cells may help explain some prior observations of the penetration of this plastic into some types of cells, including endothelial cells, blood cells, and some neighboring structures (Ajarinejad et al., 1993) or perivascular cells (Rodriguez-Baeza et al., 1998). The method described here could facilitate investigations of various tissues in relation to associated vessels, channels, and ducts. This approach also has some advantages over full corrosion casting because of the partial preservation of the surrounding organ and tissue structures, permitting an improved approach to study relationships between these various tissue structures.
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LITERATURE CITED Aharinejad SH, Bock P, Lametschwandtner A, Kikuta A, Castenholz A. 1993. Mercox-methylmethacrylic acid mixture penetrates into cells. A scanning electron microscopic study. Scanning Microsc 7:295–304. Aharinejad SH, Lametschwandtner A. 1992. Microvascular corrosion casting in scanning electron microscopy: techniques and applications. New York: Springer-Verlag. p 375.
