- Email: [email protected]
Effects of external magnetic field on biodistribution of nanoparticles: A histological study
ARTICLE IN PRESS
Journal of Magnetism and Magnetic Materials 311 (2007) 372–375 www.elsevier.com/locate/jmmm
Effects of external magnetic field on biodistribution of nanoparticles: A histological study Tony Wua, Mu-Yi Huab, Jyh-ping Chenb, Kuo-Chen Weic, Shih-Ming Jungd, Yeu-Jhy Changa, Mei-Jie Joue, Yunn-Hwa Mae, a
Department of Neurology, Chang Gung University College of Medicine and Memorial Hospital, 199 Tung-Hwa N Rd, Taipei, Taiwan Department of Chemical and Material Engineering, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Tao-Yuan, Taiwan c Department of Neurosurgery, Chang Gung University College of Medicine and Memorial Hospital, 199 Tung-Hwa N Rd, Taipei, Taiwan d Department of Pathology, Chang Gung University College of Medicine and Memorial Hospital, 199 Tung-Hwa N Rd, Taipei, Taiwan e Department of Physiology and Pharmacology, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Tao-Yuan 333, Taiwan b
Available online 9 January 2007
Abstract This study investigates the effect of external magnetic fields on the biodistribution of nanoparticles (NP). A NdFeB magnet of 2.4 kG was externally applied over the left femoral artery or right kidney. The 250 nm dextran-coated Fe3O4 NP was injected via tail vein in healthy rats, and organs were taken 1 or 24 h later. Prussian blue stain revealed that NP were more rapidly retained in the liver and spleen than in the lungs. NP aggregation observed in the kidney and femoral artery after application of external magnets was time dependent. Hollow organs such as the intestine, colon, and urinary bladder retained little NP. r 2007 Elsevier B.V. All rights reserved. Keywords: Nanoparticle; Biodistribution; External magnetic field; Prussian blue stain; Histology
1. Introduction Development of efficient drug delivery systems has attracted great attention during the last two decades. Drug delivery from carrier systems can avoid unwanted effects of the free drugs because of controlled biodistribution [1]. In medicine, magnetic fluids have been used since 1960 for magnetically controlled metallic thrombosis of intracranial aneurysms [2]. Considerable attention has been drawn to using nanoparticles (NP) as drug carriers due to their stability, biodegradability, and ease of preparation [3]. Magnetic NP are also used in highresolution magnetic resonance imaging (MRI) to allow the detection of small lymph node metastases in patients with prostate cancer [4]. Magnetic-targeted drugs can be injected into the bloodstream and guided to specific body sites with external Corresponding author. Tel.: +886 3 3283016x5266; fax: +886 3 3283031. E-mail address: [email protected] (Y.-H. Ma).
0304-8853/$ - see front matter r 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.jmmm.2006.10.1202
magnetic fields. This reduces systemic toxicity of chemotherapeutic agents, requiring a much smaller dose of anticancer drugs. In vivo studies revealed that a magnetic field applied at the tumor site accumulates the NP in tumor tissue and cells [5]. Specifically, the concentration of 123 I-labeled NP at the tumor site was dependent on the strength of the magnetic field [6]. The biodistribution of magnetic NP with external magnetic fields has been well studied in animal tumor models. The NP commonly used as drug carriers have a diameter of around 250 nm and are modified with dextran, polyethylene glycol (PEG), or polybutyl cyanoacrylate (PBC). However, the vasculature in tumor tissue has more enhanced permeability than normal blood vessels and cannot serve as a healthy rat model. For further clinical application of magnetictargeted drug delivery in cancer treatment, the distribution, clearance, and excretion of NP must be well understood. There is also an increasing need of targeted drug delivery of antibiotics, thrombolytic agents, and biologically active substances (cytokines, growth factors, and DNA sequences) [7,8]. Therefore, the behavior of the magnetic
ARTICLE IN PRESS T. Wu et al. / Journal of Magnetism and Magnetic Materials 311 (2007) 372–375
NP after injection into blood vessels, and the changes of biodistribution caused by the external magnetic field needed to be studied in models without tumors. This study investigates, in healthy rats, the effect of an external magnetic field on the biodistribution of dextran-modified NP after intravenous injection. 2. Material and methods 2.1. Magnetic nanoparticles The NP were obtained from Micromod (Rostock, Germany) and consisted of 75–80% (w/w) magnetite in a matrix of dextran (40.000 D) and can easily be separated with a conventional permanent magnet. The dextrancoated Fe3O4 NP (nanomag-D-COOH, 250 nm) had surface functionalities COOH for the covalent binding of proteins, antibodies, or other molecules. 2.2. Animals
2.5. Histology The excised tissue samples were immediately fixated at room temperature for 24 h in 10% neutral buffered formalin. These samples were dehydrated in ethanol, embedded in paraffin, sectioned, and stained with H&E and Prussian blue. Histological distributions of NP were examined after staining with Prussian blue, a specific stain to identify iron [9]. A dark blue crystalline salt Fe4[Fe(CN)6]3 was obtained by precipitation from ferric salt treated with ferrocyanic acid, whereas the nuclei were stained red with nuclear-fast red. The distribution of NP in the tissue is seen in sharp contrast under microscopic examination. 3. Results and discussions In this study, the biodistribution of NP in rats after single intravenous injection of NP suspension was
a
Pathogen-free (healthy) male Sprague–Dawley rats weighing about 350 g (14–18 weeks old) were purchased from Taipei, Taiwan, housed and used according to the protocol approved by the Chang Gung University’s Institutional Animal Care and Use Committee. During the study, animals were housed under normal conditions with 12 h light/dark cycles and were given access to food and water ad libitum. 2.3. Magnetic field
373
Number of rat: control group (no magnet), 1 hour / 24 hour group Brain: 0, 0/1 Carotid artery-R: 0, 0/0
Carotid artery-L: 0, 0/0
Lung-R: 0, 2/2
Lung-L: 0, 1/2
Stomach: 0, 0/1
Heart: 0, 0/1
Liver: 3, 3/3
Spleen: 3, 3/3
Vena cava: 0, 1/1
Aorta: 0, 0/1
Kidney-R: 0, 3/2
Kidney-L: 0, 2/1
Small intestine: 0, 0/0
Bone marrow: 0, 0/1
Femoral artery-R: 0, 0/0
Femoral artery-L: 0, 0/0 Colon: 0, 0/1
Urinary bladder: 0, 0/0
A NdFeB permanent magnet with a maximum magnetic flux density of 2.4 kG was used to produce an inhomogeneous magnetic field. The magnet was fixed securely onto the back region close to the right kidney or the inguinal region close to the proximal segment of left femoral artery of all experimental rats. The magnetic field was present for 1 h or 24 h after the NP injection.
Tail vessels: 2, 2/3 Nanoparticle injection site (tail vein) External magnet
b
Number of rat: control group (no magnet)1hr/24hr, magnet group1 hr/24 hr Brain: 0/1, 2/0
Carotid artery-R: 0/0, 0/0
Carotid artery-L: 0/0, 0/0
Lung-R: 0/3, 2/2
Lung-L: 0/3, 2/2
2.4. Surgical intervention
Stomach: 0/0, 0/1 Heart: 0/0, 0/0 Liver: 3/3, 3/3
After induction of light anesthesia by inhalation of isoflurane at a dose of 0.3 ml for approximately 15 s, the animals received an intravenous dose, via the tail vein, of 0.1 mg of the NP suspended in 100 ml normal saline using a 25 gauge, 1 in butterfly needle fitted to a syringe. The rats were sacrificed (three rats per time point for both control and experiment groups) at 1 and 24 h after the injection of NP. The heart, lungs, liver, kidneys, spleen, brain, stomach, small intestine, colon, urinary bladder, aorta, periaortic lymph nodes, inferior vena cava, carotid arteries, femoral arteries, bone marrow of lumbar vertebraes, and tail were surgically excised from the animals and prepared for histomorphologic investigation.
Spleen: 3/3, 3/3
Vena cava: 0/0, 0/2
Aorta: 0/1, 0/1
Kidney-R: 0/1, 0/2
Kidney-L: 0/1, 1/2
Small intestine: 0/0, 0/1
Bone marrow: 0/1, 0/2
Femoral artery-R: 0/0, 1/0
Femoral artery-L: 0/0, 2/1
Urinary bladder: 0/0, 0/1
Colon: 0/1, 0/2 1
Nanoparticle injection site (tail vein) External magnet
Tail vessels: 3/2, 2/3
Fig. 1. The biodistribution of nanoparticles 1 and 24 h after single intravenous injection of nanoparticles: (a) magnetic field applied to the right kidney retained the NP in both kidneys with more in the right kidney and (b) magnetic field applied at the proximal part of the left femoral artery retained NP in two of three rats after 1 h and one of three rats after 24 h.
ARTICLE IN PRESS 374
T. Wu et al. / Journal of Magnetism and Magnetic Materials 311 (2007) 372–375
investigated. Fig. 1(a) shows the results of tissue distribution when magnet was placed over surface of back close to right kidney, and Fig. 1(b) when magnet was placed over body surface close to the proximal part of left femoral artery. Fig. 2 shows the NP seen as a blue pigment in crosssection of various organ tissues.
cells and NP passed through the sinusoid of the liver and spleen red pulp, but through the capillaries of the lung. The NP were filtered out in the liver and spleen easily, but needed to pass through the fenestra of the capillary wall in order to reach the interstitial or alveolar space of the lung. 3.2. Biodistribution to magnetic field over right kidney
3.1. Biodistribution of control (without magnet) In accordance with the results reported by other authors, the removal of the NP is performed by the liver, spleen, and lungs [10,11], which belong to the reticuloendothelial system (RES). The RES is responsible for removing foreign particles or aged blood cells from the circulatory system. Results of the current study are consistent with the known theory. In the control groups, the clearance of NP in liver and spleen was evident at 1 h after injection of NP. On the other hand, the appearance of NP in lung tissue is delayed. NP were noted in none of 12 lungs after 1 h, and six of six lungs at 24 h. This may be related to the specific tissue structure of these RES organs. NP were noted in Kuppfer cells and hepatocytes of the liver and red pulp of the spleen. Blood
Magnetic field applied to the right kidney retained the NP over either kidney with more on the right side. The branch point of each renal artery from abdominal aorta is rather close. The magnetic field not only trapped the NP passing through the right kidney, but might also have attracted NP at the origin of each renal artery. Thus, NP might have flown to the left kidney with the blood stream, and have been retained in the kidney tissue. Other organ tissues close to the right kidney (i.e., stomach, small intestine, and colon) did not have significant retention of NP. 3.3. Biodistribution to magnetic field over left femoral artery External magnetic field applied at the proximal part of the left femoral artery retained NP in two of three
Fig. 2. Sections of various tissue organs removed after intravenous injection of nanoparticles: (a) NP in Kupffer cells and hepatocytes of liver tissue, (b) NP in the cytoplasm of the histocytes in the lymph node tissue, (c) NP in the tubules of the kidney, (d) NP in the alveolar space and interstitium of lung tissue, (e) NP in the red pulp of splenic tissue, and (f) NP in the red pulp of splenic tissue.
ARTICLE IN PRESS T. Wu et al. / Journal of Magnetism and Magnetic Materials 311 (2007) 372–375
experimental rats at 1 h and one of three rats at 24 h, as opposed to zero of three control rats at both times. The NP retained at femoral artery may flow away with blood circulation. The positive finding of NP in a total of five of 12 kidneys (control, two of 12) and one of six right femoral artery (control, zero of six) may be related to the weak magnetic field from nearby magnet. Other organ tissue close to the right kidney (i.e.: small intestine, colon, urinary bladder) did not have significant increased retention of NP. 3.4. Comparing the two biodistribution patterns The NP tend to aggregate more in kidney tissue than in the femoral artery. The small vessels in the kidneys have slower blood flow velocity than the femoral artery. All rats in each study group had blue pigment particles in liver and spleen tissue. The long-term outcome of these NP in the RES needs to be further clarified. On the other hand, the NP in the kidney and femoral artery by external magnetic field seems to be time dependent. 4. Conclusions In the present study, we have examined the biodistribution profiles of magnetic NP following intravenous administration in normal rats. The NP were more rapidly retained in the liver and spleen than in the lungs. The externally applied magnetic field modified the biodistribution pattern of magnetic NP in normal rat. The NP attracted in the kidney and femoral artery by external magnets seems to be time dependent. Hollow organs such as the intestine, colon, and urinary bladder retained little NP. The results of our findings will be further confirmed by an ongoing MRI study, which will also obtain the important pharmacokinetics parameters (effective clear-
375
ance and peak concentration) for the liver and spleen. Ongoing electron microscopy evaluation of the tissues will elucidate the cellular localization of the NP. Further histological examination done 28 days after intravenous NP injection will provide the biocompatibility data. Before the magnetic NP can be used as therapeutic drug carriers, these experiments are crucial for delineating the physiological (or pathophysiological) response of various organ tissues to the injected NP. Acknowledgments This study was supported by grants (CMRPD33151, CMRPD, CMRPD250011, and CMRPD140041) from the Chang Gung Memorial Hospital and Chang Gung University. References [1] J. Kreuter, in: J. Kreuter (Ed.), Colloidal Drug Delivery Systems, Marcel Dekker, New York, 1994. [2] J.F. Alksne, A. Fingerhut, R. Rand, Surgery 60 (1966) 212. [3] S.D. Troster, U. Muller, J. Kreuter, Int. J. Pharm. 61 (1990) 85. [4] M.G. Harisinghani, J. Barentsz, P.F. Hahn, et al., N. Engl. J. Med. 348 (2003) 2491. [5] C. Alexiou, R.J. Schmid, R. Jurgons, et al., Eur. Biophys. J. 35 (2006) 446. [6] C. Alexiou, A. Schmidt, R. Klein, et al., J. Magn. Magn. Mater. 252 (2002) 363. [7] A.S. Lu¨bbe, C. Alexiou, C. Bergemann, J. Surg. Res. 95 (2001) 200. [8] Y. Xie, M.D. Kaminski, S.G. Guy, et al., J. Biomed. Nanotech. 1 (2005) 410. [9] T. Grogan, K. Reinhardt, M. Jaramillo, et al., Adv. Anat. Pathol. 7 (2000) 110. [10] W.L. Monsky, D. Fukumura, T. Gohongi, et al., Cancer Res. 59 (1999) 4129. [11] S. Vishal, S. Mostafa, S. Jun, Int. J. Pharm. 308 (2006) 200.
Journal of Magnetism and Magnetic Materials 311 (2007) 372–375 www.elsevier.com/locate/jmmm
Effects of external magnetic field on biodistribution of nanoparticles: A histological study Tony Wua, Mu-Yi Huab, Jyh-ping Chenb, Kuo-Chen Weic, Shih-Ming Jungd, Yeu-Jhy Changa, Mei-Jie Joue, Yunn-Hwa Mae, a
Department of Neurology, Chang Gung University College of Medicine and Memorial Hospital, 199 Tung-Hwa N Rd, Taipei, Taiwan Department of Chemical and Material Engineering, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Tao-Yuan, Taiwan c Department of Neurosurgery, Chang Gung University College of Medicine and Memorial Hospital, 199 Tung-Hwa N Rd, Taipei, Taiwan d Department of Pathology, Chang Gung University College of Medicine and Memorial Hospital, 199 Tung-Hwa N Rd, Taipei, Taiwan e Department of Physiology and Pharmacology, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Tao-Yuan 333, Taiwan b
Available online 9 January 2007
Abstract This study investigates the effect of external magnetic fields on the biodistribution of nanoparticles (NP). A NdFeB magnet of 2.4 kG was externally applied over the left femoral artery or right kidney. The 250 nm dextran-coated Fe3O4 NP was injected via tail vein in healthy rats, and organs were taken 1 or 24 h later. Prussian blue stain revealed that NP were more rapidly retained in the liver and spleen than in the lungs. NP aggregation observed in the kidney and femoral artery after application of external magnets was time dependent. Hollow organs such as the intestine, colon, and urinary bladder retained little NP. r 2007 Elsevier B.V. All rights reserved. Keywords: Nanoparticle; Biodistribution; External magnetic field; Prussian blue stain; Histology
1. Introduction Development of efficient drug delivery systems has attracted great attention during the last two decades. Drug delivery from carrier systems can avoid unwanted effects of the free drugs because of controlled biodistribution [1]. In medicine, magnetic fluids have been used since 1960 for magnetically controlled metallic thrombosis of intracranial aneurysms [2]. Considerable attention has been drawn to using nanoparticles (NP) as drug carriers due to their stability, biodegradability, and ease of preparation [3]. Magnetic NP are also used in highresolution magnetic resonance imaging (MRI) to allow the detection of small lymph node metastases in patients with prostate cancer [4]. Magnetic-targeted drugs can be injected into the bloodstream and guided to specific body sites with external Corresponding author. Tel.: +886 3 3283016x5266; fax: +886 3 3283031. E-mail address: [email protected] (Y.-H. Ma).
0304-8853/$ - see front matter r 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.jmmm.2006.10.1202
magnetic fields. This reduces systemic toxicity of chemotherapeutic agents, requiring a much smaller dose of anticancer drugs. In vivo studies revealed that a magnetic field applied at the tumor site accumulates the NP in tumor tissue and cells [5]. Specifically, the concentration of 123 I-labeled NP at the tumor site was dependent on the strength of the magnetic field [6]. The biodistribution of magnetic NP with external magnetic fields has been well studied in animal tumor models. The NP commonly used as drug carriers have a diameter of around 250 nm and are modified with dextran, polyethylene glycol (PEG), or polybutyl cyanoacrylate (PBC). However, the vasculature in tumor tissue has more enhanced permeability than normal blood vessels and cannot serve as a healthy rat model. For further clinical application of magnetictargeted drug delivery in cancer treatment, the distribution, clearance, and excretion of NP must be well understood. There is also an increasing need of targeted drug delivery of antibiotics, thrombolytic agents, and biologically active substances (cytokines, growth factors, and DNA sequences) [7,8]. Therefore, the behavior of the magnetic
ARTICLE IN PRESS T. Wu et al. / Journal of Magnetism and Magnetic Materials 311 (2007) 372–375
NP after injection into blood vessels, and the changes of biodistribution caused by the external magnetic field needed to be studied in models without tumors. This study investigates, in healthy rats, the effect of an external magnetic field on the biodistribution of dextran-modified NP after intravenous injection. 2. Material and methods 2.1. Magnetic nanoparticles The NP were obtained from Micromod (Rostock, Germany) and consisted of 75–80% (w/w) magnetite in a matrix of dextran (40.000 D) and can easily be separated with a conventional permanent magnet. The dextrancoated Fe3O4 NP (nanomag-D-COOH, 250 nm) had surface functionalities COOH for the covalent binding of proteins, antibodies, or other molecules. 2.2. Animals
2.5. Histology The excised tissue samples were immediately fixated at room temperature for 24 h in 10% neutral buffered formalin. These samples were dehydrated in ethanol, embedded in paraffin, sectioned, and stained with H&E and Prussian blue. Histological distributions of NP were examined after staining with Prussian blue, a specific stain to identify iron [9]. A dark blue crystalline salt Fe4[Fe(CN)6]3 was obtained by precipitation from ferric salt treated with ferrocyanic acid, whereas the nuclei were stained red with nuclear-fast red. The distribution of NP in the tissue is seen in sharp contrast under microscopic examination. 3. Results and discussions In this study, the biodistribution of NP in rats after single intravenous injection of NP suspension was
a
Pathogen-free (healthy) male Sprague–Dawley rats weighing about 350 g (14–18 weeks old) were purchased from Taipei, Taiwan, housed and used according to the protocol approved by the Chang Gung University’s Institutional Animal Care and Use Committee. During the study, animals were housed under normal conditions with 12 h light/dark cycles and were given access to food and water ad libitum. 2.3. Magnetic field
373
Number of rat: control group (no magnet), 1 hour / 24 hour group Brain: 0, 0/1 Carotid artery-R: 0, 0/0
Carotid artery-L: 0, 0/0
Lung-R: 0, 2/2
Lung-L: 0, 1/2
Stomach: 0, 0/1
Heart: 0, 0/1
Liver: 3, 3/3
Spleen: 3, 3/3
Vena cava: 0, 1/1
Aorta: 0, 0/1
Kidney-R: 0, 3/2
Kidney-L: 0, 2/1
Small intestine: 0, 0/0
Bone marrow: 0, 0/1
Femoral artery-R: 0, 0/0
Femoral artery-L: 0, 0/0 Colon: 0, 0/1
Urinary bladder: 0, 0/0
A NdFeB permanent magnet with a maximum magnetic flux density of 2.4 kG was used to produce an inhomogeneous magnetic field. The magnet was fixed securely onto the back region close to the right kidney or the inguinal region close to the proximal segment of left femoral artery of all experimental rats. The magnetic field was present for 1 h or 24 h after the NP injection.
Tail vessels: 2, 2/3 Nanoparticle injection site (tail vein) External magnet
b
Number of rat: control group (no magnet)1hr/24hr, magnet group1 hr/24 hr Brain: 0/1, 2/0
Carotid artery-R: 0/0, 0/0
Carotid artery-L: 0/0, 0/0
Lung-R: 0/3, 2/2
Lung-L: 0/3, 2/2
2.4. Surgical intervention
Stomach: 0/0, 0/1 Heart: 0/0, 0/0 Liver: 3/3, 3/3
After induction of light anesthesia by inhalation of isoflurane at a dose of 0.3 ml for approximately 15 s, the animals received an intravenous dose, via the tail vein, of 0.1 mg of the NP suspended in 100 ml normal saline using a 25 gauge, 1 in butterfly needle fitted to a syringe. The rats were sacrificed (three rats per time point for both control and experiment groups) at 1 and 24 h after the injection of NP. The heart, lungs, liver, kidneys, spleen, brain, stomach, small intestine, colon, urinary bladder, aorta, periaortic lymph nodes, inferior vena cava, carotid arteries, femoral arteries, bone marrow of lumbar vertebraes, and tail were surgically excised from the animals and prepared for histomorphologic investigation.
Spleen: 3/3, 3/3
Vena cava: 0/0, 0/2
Aorta: 0/1, 0/1
Kidney-R: 0/1, 0/2
Kidney-L: 0/1, 1/2
Small intestine: 0/0, 0/1
Bone marrow: 0/1, 0/2
Femoral artery-R: 0/0, 1/0
Femoral artery-L: 0/0, 2/1
Urinary bladder: 0/0, 0/1
Colon: 0/1, 0/2 1
Nanoparticle injection site (tail vein) External magnet
Tail vessels: 3/2, 2/3
Fig. 1. The biodistribution of nanoparticles 1 and 24 h after single intravenous injection of nanoparticles: (a) magnetic field applied to the right kidney retained the NP in both kidneys with more in the right kidney and (b) magnetic field applied at the proximal part of the left femoral artery retained NP in two of three rats after 1 h and one of three rats after 24 h.
ARTICLE IN PRESS 374
T. Wu et al. / Journal of Magnetism and Magnetic Materials 311 (2007) 372–375
investigated. Fig. 1(a) shows the results of tissue distribution when magnet was placed over surface of back close to right kidney, and Fig. 1(b) when magnet was placed over body surface close to the proximal part of left femoral artery. Fig. 2 shows the NP seen as a blue pigment in crosssection of various organ tissues.
cells and NP passed through the sinusoid of the liver and spleen red pulp, but through the capillaries of the lung. The NP were filtered out in the liver and spleen easily, but needed to pass through the fenestra of the capillary wall in order to reach the interstitial or alveolar space of the lung. 3.2. Biodistribution to magnetic field over right kidney
3.1. Biodistribution of control (without magnet) In accordance with the results reported by other authors, the removal of the NP is performed by the liver, spleen, and lungs [10,11], which belong to the reticuloendothelial system (RES). The RES is responsible for removing foreign particles or aged blood cells from the circulatory system. Results of the current study are consistent with the known theory. In the control groups, the clearance of NP in liver and spleen was evident at 1 h after injection of NP. On the other hand, the appearance of NP in lung tissue is delayed. NP were noted in none of 12 lungs after 1 h, and six of six lungs at 24 h. This may be related to the specific tissue structure of these RES organs. NP were noted in Kuppfer cells and hepatocytes of the liver and red pulp of the spleen. Blood
Magnetic field applied to the right kidney retained the NP over either kidney with more on the right side. The branch point of each renal artery from abdominal aorta is rather close. The magnetic field not only trapped the NP passing through the right kidney, but might also have attracted NP at the origin of each renal artery. Thus, NP might have flown to the left kidney with the blood stream, and have been retained in the kidney tissue. Other organ tissues close to the right kidney (i.e., stomach, small intestine, and colon) did not have significant retention of NP. 3.3. Biodistribution to magnetic field over left femoral artery External magnetic field applied at the proximal part of the left femoral artery retained NP in two of three
Fig. 2. Sections of various tissue organs removed after intravenous injection of nanoparticles: (a) NP in Kupffer cells and hepatocytes of liver tissue, (b) NP in the cytoplasm of the histocytes in the lymph node tissue, (c) NP in the tubules of the kidney, (d) NP in the alveolar space and interstitium of lung tissue, (e) NP in the red pulp of splenic tissue, and (f) NP in the red pulp of splenic tissue.
ARTICLE IN PRESS T. Wu et al. / Journal of Magnetism and Magnetic Materials 311 (2007) 372–375
experimental rats at 1 h and one of three rats at 24 h, as opposed to zero of three control rats at both times. The NP retained at femoral artery may flow away with blood circulation. The positive finding of NP in a total of five of 12 kidneys (control, two of 12) and one of six right femoral artery (control, zero of six) may be related to the weak magnetic field from nearby magnet. Other organ tissue close to the right kidney (i.e.: small intestine, colon, urinary bladder) did not have significant increased retention of NP. 3.4. Comparing the two biodistribution patterns The NP tend to aggregate more in kidney tissue than in the femoral artery. The small vessels in the kidneys have slower blood flow velocity than the femoral artery. All rats in each study group had blue pigment particles in liver and spleen tissue. The long-term outcome of these NP in the RES needs to be further clarified. On the other hand, the NP in the kidney and femoral artery by external magnetic field seems to be time dependent. 4. Conclusions In the present study, we have examined the biodistribution profiles of magnetic NP following intravenous administration in normal rats. The NP were more rapidly retained in the liver and spleen than in the lungs. The externally applied magnetic field modified the biodistribution pattern of magnetic NP in normal rat. The NP attracted in the kidney and femoral artery by external magnets seems to be time dependent. Hollow organs such as the intestine, colon, and urinary bladder retained little NP. The results of our findings will be further confirmed by an ongoing MRI study, which will also obtain the important pharmacokinetics parameters (effective clear-
375
ance and peak concentration) for the liver and spleen. Ongoing electron microscopy evaluation of the tissues will elucidate the cellular localization of the NP. Further histological examination done 28 days after intravenous NP injection will provide the biocompatibility data. Before the magnetic NP can be used as therapeutic drug carriers, these experiments are crucial for delineating the physiological (or pathophysiological) response of various organ tissues to the injected NP. Acknowledgments This study was supported by grants (CMRPD33151, CMRPD, CMRPD250011, and CMRPD140041) from the Chang Gung Memorial Hospital and Chang Gung University. References [1] J. Kreuter, in: J. Kreuter (Ed.), Colloidal Drug Delivery Systems, Marcel Dekker, New York, 1994. [2] J.F. Alksne, A. Fingerhut, R. Rand, Surgery 60 (1966) 212. [3] S.D. Troster, U. Muller, J. Kreuter, Int. J. Pharm. 61 (1990) 85. [4] M.G. Harisinghani, J. Barentsz, P.F. Hahn, et al., N. Engl. J. Med. 348 (2003) 2491. [5] C. Alexiou, R.J. Schmid, R. Jurgons, et al., Eur. Biophys. J. 35 (2006) 446. [6] C. Alexiou, A. Schmidt, R. Klein, et al., J. Magn. Magn. Mater. 252 (2002) 363. [7] A.S. Lu¨bbe, C. Alexiou, C. Bergemann, J. Surg. Res. 95 (2001) 200. [8] Y. Xie, M.D. Kaminski, S.G. Guy, et al., J. Biomed. Nanotech. 1 (2005) 410. [9] T. Grogan, K. Reinhardt, M. Jaramillo, et al., Adv. Anat. Pathol. 7 (2000) 110. [10] W.L. Monsky, D. Fukumura, T. Gohongi, et al., Cancer Res. 59 (1999) 4129. [11] S. Vishal, S. Mostafa, S. Jun, Int. J. Pharm. 308 (2006) 200.