Publications by authors named "Ahram Kim"

26 Publications

  • Page 1 of 1

First report of equine parvovirus-hepatitis and equine hepacivirus coinfection in horses in Korea.

Transbound Emerg Dis 2021 Dec 17. Epub 2021 Dec 17.

Department of Infectious Diseases, College of Veterinary Medicine, Konkuk University, Gwangjin-gu, Seoul, Korea.

Equine parvovirus-hepatitis (EqPV-H) and equine hepacivirus (EqHV) are etiologically associated with Theiler's disease (TD), causing fulminant equine hepatitis, but the transmission route and co-infection effect remain unclear. We determined EqPV-H and EqHV prevalence and coinfection rate in 160 serum and 114 faecal samples using nested polymerase chain reaction. Amino acid and nucleotide analyses were performed and phylogenetic trees were constructed. By measuring liver-specific parameters (AST, GGT, TBIL and A/G ratio), hepatopathological changes in viremia status were compared. We found a high prevalence (EqPV-H: 10.6% in serum, 5.3% in faeces; EqHV: 8.1% in serum) and coinfection rate (35.3% in EqPV-H) of TD-causing agents. The newly identified EqPV-H genomes showed high nucleotide and amino acid similarities with previously reported strains in the USA, China and Austria. In phylogenetic tree and recombination analysis, a natural recombination event was confirmed between Chinese and Korean strains. In the EqPV-H or EqHV viremic horses, AST was significantly elevated and at least two liver-specific parameters were outside the reference intervals in 43.5% (10/23) of horses. To our knowledge, this is the first prevalence field study of EqPV-H and EqHV using both serum and faeces, providing further evidence of faecal-oral transmission of TD. These epidemiologic and clinicopathologic analyses specify the risk factors of TD infection and promote disease prevention strategy.
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http://dx.doi.org/10.1111/tbed.14425DOI Listing
December 2021

Synthetic molecule libraries for nucleic acid delivery: Design parameters in cationic/ionizable lipids and polymers.

Drug Metab Pharmacokinet 2021 Oct 30;42:100428. Epub 2021 Oct 30.

Department of Materials Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8656, Japan.

Recent progress in the design of cationic lipids and polymers has successfully translated nucleic acid drugs into clinical applications, such as the treatment of liver diseases and the prevention of virus infection. Small or large libraries of delivery molecules have been used to find the key chemical structures to protect nucleic acids from nucleases in the extracellular milieu and to facilitate the endosomal escape after endocytosis. This review introduces three essential design parameters (i.e., acid dissociation constant, hydrophobicity, and biodegradability) to develop synthetic molecules for nucleic acid delivery. The significance and mechanism of each parameter are described based on the results obtained from in vitro and in vivo evaluations. Other design parameters were then discussed to create the next generation of delivery molecules for future nucleic acid therapeutics.
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http://dx.doi.org/10.1016/j.dmpk.2021.100428DOI Listing
October 2021

First Detection and Genetic Characterization of New Equine Parvovirus Species Circulating among Horses in Korea.

Vet Sci 2021 Nov 7;8(11). Epub 2021 Nov 7.

Department of Infectious Diseases, College of Veterinary Medicine, Konkuk University, Seoul 05029, Korea.

Equine parvovirus-cerebrospinal fluid (EqPV-CSF) and eqcopivirus (EqCoPV) are new parvovirus species (EqPVs) identified from various tissues (CSF, blood, and respiratory swabs) in horses with neurologic and respiratory diseases. In this study, we described the prevalence rate of EqPV-CSF and EqCoPV in 133 and 77 serum and fecal samples, respectively, using polymerase chain reaction. Further, we analyzed the potential risk factors for infection. We calculated the nucleotide and amino acid similarity and constructed phylogenetic trees. There was a moderate-to-high prevalence rate (EqPV-CSF: 3.8%; EqCoPV 9.8%) of each virus in serum; moreover, age, country of foaling, and clinical colic signs were significantly associated with the EqPVs infection. The newly identified EqPV-CSF/EqCoPV genomes had high nucleotide and amino acid identities with previously isolated strains in the USA. In phylogenetic analysis, they clustered and formed a new subgroup in the genus . To our knowledge, this is the first field epidemiologic study on EqPV-CSF and EqCoPV using both serum and fecal samples. Our findings demonstrate the risk factors for infection and could facilitate the development of disease prevention strategies.
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http://dx.doi.org/10.3390/vetsci8110268DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8621016PMC
November 2021

Analgesic and cardiopulmonary effects of premedication with tramadol in calves anesthetized with the infusion of guaifenesin and thiamylal.

J Vet Med Sci 2021 Dec 1;83(12):1988-1993. Epub 2021 Nov 1.

College of Veterinary Medicine and Veterinary Medical Research Institute, Jeju National University, Jeju 63243, Republic of Korea.

This study examined the analgesic and cardiopulmonary effects of intravenous (IV) tramadol during general intravenous anesthesia in calves. Calves were premedicated with diazepam (0.2 mg/kg, IV) with tramadol (2 mg/kg, IV) (group T) or saline (group S). Anesthesia was induced by thiamylal sodium (4 mg/kg, IV) and maintained with an infusion (2 ml/kg/hr) of 5% guaifenesin containing thiamylal sodium (2 mg/ml). Additional thiamylal sodium (1-2 mg/kg, IV) was administered when interference from the calves was observed during surgery. The total counts of additional thiamylal sodium administration, analgesia score using a visual analog scale, recovery time, and cardiopulmonary function in the different groups were assessed and compared. Group T showed significantly fewer counts of additional drug administration and a significantly higher analgesia score. Tramadol may provide adequate analgesia with minimal cardiopulmonary changes in calves during general anesthesia.
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http://dx.doi.org/10.1292/jvms.21-0196DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8762425PMC
December 2021

First Clinical Case of Equine Parvovirus-Hepatitis-Related Theiler's Disease in Asia.

Viruses 2021 09 24;13(10). Epub 2021 Sep 24.

Department of Infectious Diseases, College of Veterinary Medicine, Konkuk University, Seoul 05029, Korea.

Equine parvovirus-hepatitis (EqPV-H) is a newly identified etiologic agent of Theiler's disease (TD). We present a case of EqPV-H-related fulminant hepatitis in a 14-year-old thoroughbred mare in Korea. The mare had acute hepatopathy and gastrointestinal symptoms, with abnormal liver-related blood parameters. The horse was born in the USA and imported to Korea in 2017, with no history of administration of equine biological products after entry into Korea. The horse was diagnosed with EqPV-H-associated hepatitis after abdominal ultrasonography, laparotomy, and nested polymerase chain reaction (PCR) and in situ hybridization (ISH) assays. The serum, nasal swab, oral swab, and liver biopsy were positive for EqPV-H according to the PCR assay. Genetic analysis of the partial NS1 gene of EqPV-H showed a unique nucleotide substitution, distinct from that in previously deposited strains. EqPV-H DNA was found not only in hepatocytes but also in bile duct epithelium and Kupffer cells, particularly via ISH. To the best of our knowledge, this is the first case of EqPV-H-associated TD in Asia, providing the first clinical evidence for viral shedding from the mouth and nose, and identification of EqPV-H in the liver. This study contributes to a better understanding of the pathological features of EqPV-H-associated TD.
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http://dx.doi.org/10.3390/v13101917DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8541225PMC
September 2021

Comparison of the Fecal Microbiota of Horses with Intestinal Disease and Their Healthy Counterparts.

Vet Sci 2021 Jun 17;8(6). Epub 2021 Jun 17.

Faculty of Biotechnology, School of Life Sciences, SARI, Jeju 63243, Korea.

(1) Background: The intestinal microbiota plays an essential role in maintaining the host's health. Dysbiosis of the equine hindgut microbiota can alter the fermentation patterns and cause metabolic disorders. (2) Methods: This study compared the fecal microbiota composition of horses with intestinal disease and their healthy counterparts living in Korea using 16S rRNA sequencing from fecal samples. A total of 52 fecal samples were collected and divided into three groups: horses with large intestinal disease ( = 20), horses with small intestinal disease ( = 8), and healthy horses ( = 24). (3) Results: Horses with intestinal diseases had fewer species and a less diverse bacterial population than healthy horses. Lactic acid bacteria, Lachnospiraceae, and Lactobacillaceae were overgrown in horses with large intestinal colic. The Firmicutes to Bacteroidetes ratio (F/B), which is a relevant marker of gut dysbiosis, was 1.94, 2.37, and 1.74 for horses with large intestinal colic, small intestinal colic, and healthy horses, respectively. (4) Conclusions: The overgrowth of two lactic acid bacteria families, Lachnospiraceae and Lactobacillaceae, led to a decrease in hindgut pH that interfered with normal fermentation, which might cause large intestinal colic. The overgrowth of also led to a decrease in pH in the hindgut, which suppressed the proliferation of the methanogen and reduced methanogenesis in horses with small intestinal colic.
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http://dx.doi.org/10.3390/vetsci8060113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8234941PMC
June 2021

Comparison of the Gut Microbiota of Jeju and Thoroughbred Horses in Korea.

Vet Sci 2021 May 11;8(5). Epub 2021 May 11.

College of Veterinary Medicine and Veterinary Medical Research Institute, Jeju National University, Jeju 63243, Korea.

(1) Background: The large intestine of horses is an anaerobic fermentative chamber filled with fibrolytic bacteria that play essential roles in digesting and absorbing nutrients for energy production. Although Jeju horses are a prominent local breed in Korea, few studies have investigated the gut microbiota of Jeju horses; (2) Methods: This study performed sequencing of V3 and V4 hypervariable regions of the partial 16S rRNA genes obtained from horse fecal samples and compared the gut microbiota between Jeju and Thoroughbred horses. Thirty and 24 fecal samples were obtained from Jeju and Thoroughbred horses, respectively; (3) Results: The gut microbiota belonged to 23 phyla and 159 families. Firmicutes and Bacteroidetes were the most abundant and predominant phyla, followed by Verrucomicrobia, Euryachaeota, and Spirochaete. The ratio of Firmicutes to Bacteroidetes (F/B), which is known as a relevant marker of gut dysbiosis, was 1.84 for Jeju horses, whereas it was 1.76 for Thoroughbred horses. Moreover, at the genus level, 21 genera were significantly different between the Jeju and Thoroughbred horses ( < 0.05); (4) Conclusions: The Thoroughbred horse's gut microbiotas had significantly higher diversity than the Jeju horses ( < 0.05). In addition, beneficial commensal bacteria that produce short-chain fatty acids thus providing a significant source of energy are also more abundant in Thoroughbred horses. These results provide novel information on the horse gut microbiota and insights for further studies related to the horse gut microbiota.
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http://dx.doi.org/10.3390/vetsci8050081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8151153PMC
May 2021

Antioxidant Nanomedicine Significantly Enhances the Survival Benefit of Radiation Cancer Therapy by Mitigating Oxidative Stress-Induced Side Effects.

Small 2021 05 15;17(21):e2008210. Epub 2021 Apr 15.

Department of Materials Science, Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8573, Japan.

Oxidative stress-induced off-target effects limit the therapeutic window of radiation therapy. Although many antioxidants have been evaluated as radioprotective agents, none of them are in widespread clinical use, owing to the side effects of the antioxidants themselves and the lack of apparent benefit. Aiming for a truly effective radioprotective agent in radiation cancer therapy, the performance of a self-assembling antioxidant nanoparticle (herein denoted as redox nanoparticle; RNP) is evaluated in the local irradiation of a subcutaneous tumor-bearing mouse model. Since RNP is covered with a biocompatible shell layer and possesses a core-shell type structure of several tens of nanometers in size, its lifetime in the systemic circulation is prolonged. Moreover, since 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO), one of the most potent antioxidants, is covalently encapsulated in the core of RNP, it exerts intense antioxidant activity and induces fewer adverse effects by avoiding leakage of the TEMPO molecules. Preadministration of RNP to the mouse model effectively mitigates side effects in normal tissues and significantly extends the survival benefit of radiation cancer therapy. Moreover, RNP pretreatment noticeably increases the apoptosis/necrosis ratio of radiation-induced cell death, a highly desirable property to reduce the chronic side effects of ionizing irradiation.
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http://dx.doi.org/10.1002/smll.202008210DOI Listing
May 2021

An Antioxidant Nanoparticle Enhances Exercise Performance in Rat High-intensity Running Models.

Adv Healthc Mater 2021 05 4;10(10):e2100067. Epub 2021 Mar 4.

Department of Materials Science, Graduate School of Pure and Applied Science, University of Tsukuba, Tennodai 1-1-1, Tsukuba, Ibaraki, 305-8573, Japan.

Although the adverse effects of excessively generated reactive oxygen species (ROS) on the body during aerobic exercise have been debated, there are few reports on the remarkable effects of the application of conventional antioxidants on exercise performance. The conventional antioxidants could not enhance exercise performance due to their rapid excretion from the body and serious adverse effects on the cellular respiratory system. In this study, impact of the original antioxidant self-assembling nanoparticle, redox-active nanoparticle (RNP), is investigated on the exercise performance of rats during running experiments. With an increase in the dose of the administered RNP, the all-out time of the rat running extends in a dose-dependent manner. In contrast, with an increase in the dose of the low-molecular-weight (LMW) antioxidant, the all-out running time of the rats decreases. The control group and LMW antioxidant treated group decrease in the number of red blood cells (RBCs) and increase oxidative stress after running. However, the RNP group maintains a similar RBC level and oxidative stress as that of the sedentary group. The results suggest that RNP, which shows long-blood circulation without disturbance of mitohormesis, effectively removes ROS from the bloodstream to suppresses RBC oxidative stress and damage, thus improving exercise performance.
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http://dx.doi.org/10.1002/adhm.202100067DOI Listing
May 2021

Comparison of intraoperative cardiorespiratory and behavioral responses to medetomidine combined with tramadol or butorphanol during standing laparoscopic ovariectomy in horses.

J Vet Med Sci 2021 Apr 11;83(4):643-647. Epub 2021 Feb 11.

Department of Veterinary Clinical Sciences, Jeju National University, Jeju 63243, Republic of Korea.

The purpose of this study was to assess the cardiorespiratory and behavioral responses to the combination of medetomidine and tramadol (M-T) or butorphanol (M-B) in standing laparoscopic ovariectomy in horses. One ovary was removed under M-T and the contralateral ovary was removed under M-B with at least 4 weeks between operations at random. Horses were sedated using intravenous medetomidine (5 µg/kg) followed by tramadol (1 mg/kg) or butorphanol (10 µg/kg) after 5 min. Sedation was maintained through the repeated injection of medetomidine (1 µg/kg) and tramadol (0.4 mg/kg) or medetomidine (1 µg/kg) and butorphanol (4 µg/kg) every 15 min. Cardiorespiratory function and behavioral responses, including, sedation, ataxia, and analgesia, were assessed during the surgery. There were no significant differences in cardiorespiratory values and sedation and analgesia scores between M-T and M-B. Ataxia scores were significantly lower in M-T than in M-B. This result suggests that M-T could maintain smooth and stable standing surgery with minimal cardiorespiratory changes in horses.
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http://dx.doi.org/10.1292/jvms.20-0397DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8111346PMC
April 2021

Molecular design of a high-performance polymeric carrier for delivery of a variety of boronic acid-containing drugs.

Acta Biomater 2021 02 13;121:554-565. Epub 2020 Dec 13.

Department of Materials Science, Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8573, Japan; Master's School of Medical Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8573, Japan; Center for Research in Isotopes and Environmental Dynamics (CRiED), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8573, Japan. Electronic address:

Because of their many useful and unique properties, boronic acids are well suited for biomedical applications such as antitumor chemotherapy and boron neutron capture therapy (BNCT). Bortezomib, a boronic acid derivative, has drawn a lot of attention as a potent proteasome inhibitor. Nevertheless, because of rapid excretion and off-target effects, the clinical translation of boronic acid-containing drugs is limited. To this end, we employed a polymeric carrier to stably encapsulate boronic acid-containing drugs and achieve superior pharmacokinetics with an on-target drug release capability. Accordingly, to construct a supramolecular polymeric nanoparticle, we took advantage of the facile, stable, and pH-sensitive conjugation between boronic acids and diethanolamine-installed polymeric carriers. We demonstrated the feasibility of our molecular design by generating and applying bortezomib-loaded nanoparticles to a subcutaneous tumor-bearing mouse model. Stable encapsulation and pH-sensitive release of bortezomib facilitated antitumor efficacy and alleviated hepatotoxicity. We also verified the versatility of our approach through biological evaluations of the nanoparticles encapsulating benzo(b)thiophene-2-boronic acid, phenylboronic acid, and p-phenylene-diboronic acid.
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http://dx.doi.org/10.1016/j.actbio.2020.12.015DOI Listing
February 2021

Non-isotope enriched phenylboronic acid-decorated dual-functional nano-assembles for an actively targeting BNCT drug.

Biomaterials 2021 01 24;268:120551. Epub 2020 Nov 24.

Department of Materials Science, Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8573, Japan; Master's Program in Medical Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8573, Japan; Center for Research in Isotopes and Environmental Dynamics (CRiED), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8577, Japan. Electronic address:

The feasibility of boron neutron capture therapy (BNCT) greatly depends on the selective accumulation of B in tumors. The p-boronophenylalanine-fructose (BPA-f) complex has been established as a conventional BNCT agent due to its preferential uptake into tumors, which is driven by amino acid transporters. However, the retention of BPA-f in tumors is highly limited because of an antiport mechanism, which is regulated by a gradient of amino acid concentration across the cancer cell membrane. Thus, to preserve a high B concentration in tumors, patients are inevitably subjected to a constant intravenous infusion. To this end, we employed a phenylboronic acid (PBA)-decorated polymeric nanoparticle (Nano) as a sialic acid-targeting BNCT agent. In this manner, the PBA can exhibit dual functionalities, i.e., exhibiting a neutron capture capacity and hypersialyated cancer cell targeting effect. Our developed Nano possesses a supramolecular structure composed of a core and shell comprised of poly(lactic acid) (PLA) and poly(ethylene glycol) (PEG) segments, respectively. The PBA moiety is installed at the PEG end, providing an unusually strong targeting effect, supposedly via multivalent binding onto the cancer cell membrane. As in BNCT, we verified the feasibility of Nano against a B16 melanoma-bearing mouse model. By virtue of efficient tumor targeting, even at a 100-fold lower dose than BPA-f, the Nano achieved a potent antitumor effect.
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http://dx.doi.org/10.1016/j.biomaterials.2020.120551DOI Listing
January 2021

Integrating nanofibers with biochemical gradients to investigate physiologically-relevant fibroblast chemotaxis.

Lab Chip 2019 11 27;19(21):3641-3651. Epub 2019 Sep 27.

Department of Mechanical Engineering, Virginia Tech, Blacksburg, VA 24061, USA. and School of Biomedical Engineering & Sciences, Virginia Tech, Blacksburg, VA 24061, USA.

Persistent cell migration can occur due to anisotropy in the extracellular matrix (ECM), the gradient of a chemo-effector, or a combination of both. Through a variety of in vitro platforms, the contributions of either stimulus have been extensively studied, while the combined effect of both cues remains poorly described. Here, we report an integrative microfluidic chemotaxis assay device that enables the study of single cell chemotaxis on ECM-mimicking, aligned, and suspended nanofibers. Using this assay, we evaluated the effect of fiber spacing on the morphology and chemotaxis response of embryonic murine NIH/3T3 fibroblasts in the presence of temporally invariant, linear gradients of platelet-derived growth factor-BB (PDGF-BB). We found that the strength of PDGF-mediated chemotaxis response depends on not only the gradient slope but also the cell morphology. Low aspect ratio (3.4 ± 0.2) cells on flat substrata exhibited a chemotaxis response only at a PDGF-BB gradient of 0-10 ng mL. However, high aspect ratio (19.1 ± 0.7) spindle-shaped cells attached to individual fibers exhibited maximal chemotaxis response at a ten-fold shallower gradient of 0-1 ng mL, which was robustly maintained up to 0-10 ng mL. Quadrilateral-shaped cells of intermediate aspect ratio (13.6 ± 0.8) attached to two fibers exhibited a weaker response compared to the spindle-shaped cells, but still stronger compared to cells attached to 2D featureless substrata. Through pharmacological inhibition, we show that the mesenchymal chemotaxis pathway is conserved in cells on fibers. Altogether, our findings show that chemotaxis on ECM-mimicking fibers is modulated by fiber spacing-driven cell shape and can be significantly different from the behavior observed on flat 2D substrata. We envisage that this microfluidic platform will have wide applicability in understanding the combined role of ECM architecture and chemotaxis in physiological and pathological processes.
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http://dx.doi.org/10.1039/c9lc00602hDOI Listing
November 2019

Improved Size Determination by Nanoparticle Tracking Analysis: Influence of Recognition Radius.

Anal Chem 2019 08 14;91(15):9508-9515. Epub 2019 Jun 14.

School of Materials Science and Engineering , Nanyang Technological University , 50 Nanyang Avenue 639798 , Singapore.

Nanoparticle tracking analysis (NTA) is a light scattering technique that measures the size distribution of suspended nanoparticles. A key parameter in NTA measurements is the recognition radius, which distinguishes individual, tracked particles from one another. However, by defining a finite radius, the displacement of tracked particles is effectively restricted, translating into an overestimation of particle size. Herein, we introduce a modified probability model that describes the restricted displacement of a tracked particle and enables more accurate nanoparticle size determination. The analytical performance of the modified displacement probability was validated through computational simulations and experimental measurements. While conventional models typically overestimated nanoparticle size by over 20%, the modified probability model estimated nanoparticle size with an error of less than 6%. The modified probability model is compatible with conventional NTA measurement readouts and thus should find wide application for more accurately determining the size distribution of suspended nanoparticles.
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http://dx.doi.org/10.1021/acs.analchem.9b00454DOI Listing
August 2019

Encapsulation of tissue plasminogen activator in pH-sensitive self-assembled antioxidant nanoparticles for ischemic stroke treatment - Synergistic effect of thrombolysis and antioxidant.

Biomaterials 2019 09 13;215:119209. Epub 2019 May 13.

Department of Materials Science, Graduate School of Pure and Applied Science, University of Tsukuba, Tennoudai 1-1-1, Tsukuba, Ibaraki, 305-8573, Japan; Master's School in Medical Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tennoudai 1-1-1, Tsukuba, Ibaraki, 305-8573, Japan; Center for Research in Isotopes and Environmental Dynamics (CRiED), University of Tsukuba, Tennoudai 1-1-1, Tsukuba, Ibaraki, 305-8573, Japan. Electronic address:

The medical treatment for stroke has advanced greatly in recent years. Thrombolytic therapy with tissue plasminogen activator (t-PA) is one of the mainstream treatments, but it still has many problems, including short half-life, and t-PA-induced reperfusion and oxidative injuries. To broaden the therapeutic window of t-PA and reduce its associated oxidative stress after reperfusion, t-PA-installed, nitroxide radical-containing, self-assembled polyion complex nanoparticles ([email protected]) were designed. Encapsulation of t-PA in the self-assembled antioxidant nanoparticles improved its bioavailability and extended its therapeutic window. To suppress reactive oxygen species (ROS) in the ischemic penumbra area, the low-molecular-weight nitroxide antioxidant 4-amino-2,2,6,6-tetramethylpiperidine-1-oxyl was covalently conjugated with the nanoparticle matrix, thus suppressing oxidative damage in the brain after reperfusion. t-PA and nitroxide radicals were confined and protected in the core of [email protected], thereby preventing their rapid metabolism and excretion out of the body after systemic circulation for prolonged period. The nano-sized formulation prevented non-specific internalization of [email protected] in healthy cells, thereby preserving the normal function of redox reactions in the cells, especially important redox reactions such as electron transport chains. This improved pharmacological performance of [email protected] remarkably extended the in vivo half-life of t-PA in systemic circulation. Using a mouse model of photo-thrombotic middle cerebral artery occlusion, we found that [email protected] treatment, compared with naked t-PA, void iRNP, or [email protected] (non-ROS scavenging nanoparticle as a control), significantly suppressed increases in cerebral infarct volume and improved neurological deficit after brain ischemia. t-PA-induced subarachnoid hemorrhage was also suppressed by [email protected] treatment through elimination of overproduced ROS. Based on these data, [email protected] presents therapeutic potential through synergistic effect of thrombolysis and antioxidant effects for preventing and treating ischemia-reperfusion injury.
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http://dx.doi.org/10.1016/j.biomaterials.2019.05.020DOI Listing
September 2019

Enhancing Boron Uptake in Brain Glioma by a Boron-Polymer/Microbubble Complex with Focused Ultrasound.

ACS Appl Mater Interfaces 2019 Mar 18;11(12):11144-11156. Epub 2019 Mar 18.

Department of Biomedical Engineering and Environmental Sciences , National Tsing Hua University , No. 101, Section 2, Kuang-Fu Road , Hsinchu 30013 , Taiwan.

Boron neutron capture therapy (BNCT) is a promising radiotherapy for treating glioblastoma multiforme (GBM). However, the penetration of drugs (e.g., sodium borocaptate and BSH) for BNCT into brain tumors is limited by cerebral vesicular protective structures, the blood-brain barrier, and the blood-brain tumor barrier (BTB). Although BSH has been reported to be selectively taken up by tumors, it is rapidly excreted from the body and cannot achieve a high tumor-to-normal brain ratio (T/N ratio) and tumor-to-blood ratio (T/B ratio). Despite the development of large-molecular weight boron compounds, such as polymers and nanoparticles, to enhance the permeation and retention effect, their effects remain insufficient for clinical use. To improve the efficiency of boron delivery to the tumor site, we propose combinations of self-assembled boron-containing polyanion [polyethylene glycol- b-poly(( closo-dodecaboranyl)thiomethylstyrene) (PEG- b-PMBSH)] nanoparticles (295 ± 2.3 nm in aqueous media) coupled with cationic microbubble (B-MB)-assisted focused ultrasound (FUS) treatment. Upon FUS sonication (frequency = 1 MHz, pressure = 0.3-0.7 MPa, duty cycle = 0.5%, sonication = 1 min), B-MBs can simultaneously achieve safe BTB opening and boron drug delivery into tumor tissue. Compared with the MBs of the PEG- b-PMBSH mixture group (B + MBs), B-MBs showed 3- and 2.3-fold improvements in the T/N (4.4 ± 1.4 vs 1.3 ± 0.1) and T/B ratios (1.4 ± 0.6 vs 0.1 ± 0.1), respectively, after 4 min of FUS sonication. The spatial distribution of PEG- b-PMBSH was also improved by the complex of PEG- b-PMBSH with MBs. The findings presented herein, in combination with the expanding clinical application of FUS, may improve BNCT and treatment of GBM.
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http://dx.doi.org/10.1021/acsami.8b22468DOI Listing
March 2019

Validation of Size Estimation of Nanoparticle Tracking Analysis on Polydisperse Macromolecule Assembly.

Sci Rep 2019 02 25;9(1):2639. Epub 2019 Feb 25.

School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, 639798, Singapore, Singapore.

As the physicochemical properties of drug delivery systems are governed not only by the material properties which they are compose of but by their size that they conform, it is crucial to determine the size and distribution of such systems with nanometer-scale precision. The standard technique used to measure the size distribution of nanometer-sized particles in suspension is dynamic light scattering (DLS). Recently, nanoparticle tracking analysis (NTA) has been introduced to measure the diffusion coefficient of particles in a sample to determine their size distribution in relation to DLS results. Because DLS and NTA use identical physical characteristics to determine particle size but differ in the weighting of the distribution, NTA can be a good verification tool for DLS and vice versa. In this study, we evaluated two NTA data analysis methods based on maximum-likelihood estimation, namely finite track length adjustment (FTLA) and an iterative method, on monodisperse polystyrene beads and polydisperse vesicles by comparing the results with DLS. The NTA results from both methods agreed well with the mean size and relative variance values from DLS for monodisperse polystyrene standards. However, for the lipid vesicles prepared in various polydispersity conditions, the iterative method resulted in a better match with DLS than the FTLA method. Further, it was found that it is better to compare the native number-weighted NTA distribution with DLS, rather than its converted distribution weighted by intensity, as the variance of the converted NTA distribution deviates significantly from the DLS results.
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http://dx.doi.org/10.1038/s41598-019-38915-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6389903PMC
February 2019

Small Delivery Vehicles of siRNA for Enhanced Cancer Targeting.

Biomacromolecules 2018 07 18;19(7):2377-2390. Epub 2018 Jun 18.

Department of Materials Engineering, Graduate School of Engineering , The University of Tokyo , 7-3-1 Hongo , Bunkyo-ku, Tokyo 113-8656 , Japan.

Small interfering RNA (siRNA) drugs have been considered to treat various diseases in major organs. However, siRNA drugs developed for cancer therapy are hindered from proceeding to the clinic. To date, various delivery formulations have been developed from cationic lipids, polymers, and/or inorganic nanoparticles for systemic siRNA delivery to solid tumors. Most of these delivery vehicles do not generate small particle sizes and pharmacokinetics required for accumulation in target cancer cells compared with clinically tested anticancer drug-loaded polymeric micelles. This review describes the significance of small, long-circulating vehicles for efficient delivery of siRNA to cancer tissues via the enhanced permeability and retention (EPR) effect. We summarize recent biological evidence that supports the size effect of delivery vehicles in tumor microenvironments and introduce promising strategies for the construction of small vehicles with sizes of 10-50 nm. We then discuss the feasibility of these delivery vehicles with respect to translation to clinical trials.
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http://dx.doi.org/10.1021/acs.biomac.8b00546DOI Listing
July 2018

Design of Nanofiber Coatings for Mitigation of Microbial Adhesion: Modeling and Application to Medical Catheters.

ACS Appl Mater Interfaces 2018 May 26;10(18):15477-15486. Epub 2018 Apr 26.

Surface-associated microbial communities, known as biofilms, pose significant challenges in clinical and industrial settings. Micro-/nanoscale substratum surface features have been shown to disrupt firm adhesion of planktonic microbes to surfaces, thereby interfering with the earliest stage of biofilm formation. However, the role of geometry and size of surface features in microbial retention is not completely understood. In this study, we developed a biophysical model that describes the changes in the total free energy (adhesion energy and stretching energy) of an adherent Candida albicans cell on nanofiber-coated surfaces as a function of the geometry (i.e., diameter) and configuration (i.e., interfiber spacing) of the surface features (i.e., nanofibers). We then introduced a new nondimensional parameter, Π, to represent the ratio of cell rigidity to cell-substratum interfacial energy. We show that the total free energy is a strong function of topographical feature size at higher Π and lower spacing values. To confirm our biophysical model predictions, we performed 24 h dynamic retention assays and quantified cell attachment number density on surfaces coated with highly ordered polystyrene nanofibers. We show that the total free energy of a single adherent cell on a patterned surface is a key determinant of microbial retention on that surface. The cell attachment density trend closely correlates with the predictions based on the adherent single-cell total energy. The nanofiber coating design (1.2 μm diameter, 2 μm spacing) that maximized the total energy of the adherent cell resulted in the lowest microbial retention. We further demonstrate the utility of our biophysical model by showing close correlation between the computed single-cell total free energy and biofilm nucleation on fiber-coated urinary and central venous catheters of different materials. This biophysical model could offer a powerful new paradigm in ab initio design of patterned surfaces for controlled biofilm growth for medical applications and beyond.
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http://dx.doi.org/10.1021/acsami.8b02907DOI Listing
May 2018

Recent progress in development of siRNA delivery vehicles for cancer therapy.

Adv Drug Deliv Rev 2016 09 25;104:61-77. Epub 2016 Jun 25.

Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan; Department of Materials Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan; Innovation Center of NanoMedicine, Institute of Industry Promotion-Kawasaki, 3-25-14 Tonomachi, Kawasaki-ku, Kawasaki 210-0821, Japan. Electronic address:

Recent progress in RNA biology has broadened the scope of therapeutic targets of RNA drugs for cancer therapy. However, RNA drugs, typically small interfering RNAs (siRNAs), are rapidly degraded by RNases and filtrated in the kidney, thereby requiring a delivery vehicle for efficient transport to the target cells. To date, various delivery formulations have been developed from cationic lipids, polymers, and/or inorganic nanoparticles for systemic delivery of siRNA to solid tumors. This review describes the current status of clinical trials related to siRNA-based cancer therapy, as well as the remaining issues that need to be overcome to establish a successful therapy. It, then introduces various promising design strategies of delivery vehicles for stable and targeted siRNA delivery, including the prospects for future design.
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http://dx.doi.org/10.1016/j.addr.2016.06.011DOI Listing
September 2016

Intracellular Delivery of Charge-Converted Monoclonal Antibodies by Combinatorial Design of Block/Homo Polyion Complex Micelles.

Biomacromolecules 2016 Feb 4;17(2):446-53. Epub 2016 Jan 4.

Polymer Chemistry Division, Chemical Resources Laboratory, Tokyo Institute of Technology , R1-11, 4529 Nagatsuta, Midori-ku, Yokohama 226-8503, Japan.

Direct intracellular delivery of antibodies has gained much attention, although only a few agents have been developed, and none of them has reached clinical stages. The main obstacles here are the insufficient characteristics of delivery systems including stability and appropriate ability for intracellular antibody release. We tailored the structure of polyion complex (PIC) micelles by loading transiently charge-converted antibody derivatives for achieving enhanced stability, delivery to cytosol, and precise antigen recognition inside cells. Citraconic anhydride was used for the charge conversion of the antibody; the optimized degree of modification was identified to balance the stability of PIC micelles in the extracellular compartment and prompt pH-triggered disintegration after their translocation into the acidic endosomal compartment of target cells. The use of a mixture of homo- and block-catiomers in an appropriate ratio to construct PIC micelles substantially enhanced the endosomal escaping efficacy of the loaded antibody, leading to improved recognition of intracellular antigens.
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http://dx.doi.org/10.1021/acs.biomac.5b01335DOI Listing
February 2016

Induction of Secondary Structure through Micellization of an Oppositely Charged Pair of Homochiral Block- and Homopolypeptides in an Aqueous Medium.

Macromol Rapid Commun 2015 Nov 21;36(22):1958-64. Epub 2015 Aug 21.

Graduate School of Engineering, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-8656, Japan.

Polyion complex (PIC) formation is an attractive method for obtaining molecular assemblies owing to their facile fabrication process in aqueous media, but more insights are required in order to control the higher-dimensional structures of polypeptide-based PICs. Herein, the PIC formation behavior of oppositely charged homochiral polypeptides, poly-L-lysine and poly(ethylene glycol)-b-poly(L-glutamate) (PEG-PLG), and their secondary structures are carefully studied in water. PIC formation takes place in a polymer concentration-dependent manner, and clear β-sheet formation is observed at polymer concentrations ≥0.3 mg mL(-1). The results also confirm that multimolecular aggregation is a prerequisite for β-sheet formation, which indicates that the inner hydrophobic environment of PICs is favorable for β-sheet formation. Furthermore, the PEG weight fraction, stereoregularity of the polypeptide, and ionic strength of the solutions are found to be key factors for generating a secondary structure, presumably because these factors can contribute to the tuning of the inner environment of PICs. This method of producing water-soluble nanoassemblies from oppositely charged polypeptides may expedite self-assembly studies in biological systems and be incorporated into various molecular systems to exploit protein-mimicking features.
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http://dx.doi.org/10.1002/marc.201500368DOI Listing
November 2015

Antibody fragment-conjugated polymeric micelles incorporating platinum drugs for targeted therapy of pancreatic cancer.

Biomaterials 2015 Jan 15;39:23-30. Epub 2014 Nov 15.

Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan; Division of Clinical Biotechnology, Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; Department of Materials Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan. Electronic address:

Antibody-mediated therapies including antibody-drug conjugates (ADCs) have shown much potential in cancer treatment by tumor-targeted delivery of cytotoxic drugs. However, there is a limitation of payloads that can be delivered by ADCs. Integration of antibodies to drug-loaded nanocarriers broadens the applicability of antibodies to a wide range of therapeutics. Herein, we developed antibody fragment-installed polymeric micelles via maleimide-thiol conjugation for selectively delivering platinum drugs to pancreatic tumors. By tailoring the surface density of maleimide on the micelles, one tissue factor (TF)-targeting Fab' was conjugated to each carrier. Fab'-installed platinum-loaded micelles exhibited more than 15-fold increased cellular binding within 1 h and rapid cellular internalization compared to non-targeted micelles, leading to superior in vitro cytotoxicity. In vivo, Fab'-installed micelles significantly suppressed the growth of pancreatic tumor xenografts for more than 40 days, outperforming non-targeted micelles and free drugs. These results indicate the potential of Fab'-installed polymeric micelles for efficient drug delivery to solid tumors.
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http://dx.doi.org/10.1016/j.biomaterials.2014.10.069DOI Listing
January 2015

Light-induced cytosolic activation of reduction-sensitive camptothecin-loaded polymeric micelles for spatiotemporally controlled in vivo chemotherapy.

ACS Nano 2014 Nov 14;8(11):11591-602. Epub 2014 Nov 14.

Department of Bioengineering, Graduate School of Engineering, The University of Tokyo , 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.

Nanomedicines capable of smart operation at the targeted site have the potential to achieve the utmost therapeutic benefits. Providing nanomedicines that respond to endogenous stimuli with an additional external trigger may improve the spatiotemporal control of their functions, while avoiding drawbacks from their inherent tissue distribution. Herein, by exploiting the permeabilization of endosomes induced by photosensitizer agents upon light irradiation, we complemented the intracellular action of polymeric micelles incorporating camptothecin (CPT), which can sharply release the loaded drug in response to the reductive conditions of the cytosol, as an effective strategy for precisely controlling the function of these nanomedicines in vivo, while advancing toward a light-activated chemotherapy. These camptothecin-loaded micelles (CPT/m) were stable in the bloodstream, with minimal drug release in extracellular conditions, leading to prolonged blood circulation and high accumulation in xenografts of rat urothelial carcinoma. With the induction of endosomal permeabilization with the clinically approved photosensitizer, Photofrin, the CPT/m escaped from the endocytic vesicles of cancer cells into the cytosol, as confirmed both in vitro and in vivo by real-time confocal laser microscopies, accelerating the drug release from the micelles only in the irradiated tissues. This spatiotemporal switch significantly enhanced the in vivo antitumor efficacy of CPT/m without eliciting any toxicity, even at a dose 10-fold higher than the maximum tolerated dose of free CPT. Our results indicate the potential of reduction-sensitive drug-loaded polymeric micelles for developing safe chemotherapies after activation by remote triggers, such as light, which are capable of permeabilizing endosomal compartments.
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http://dx.doi.org/10.1021/nn504836sDOI Listing
November 2014

Aligned and suspended fiber force probes for drug testing at single cell resolution.

Biofabrication 2014 Sep 29;6(4):045006. Epub 2014 Sep 29.

School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, USA.

The role of physical forces in disease onset and progression is widely accepted and this knowledge presents an alternative route to investigating disease models. Recently, numerous force measurement techniques have been developed to probe single and multi-cell behavior. While these methods have yielded fundamental insights, they are yet unable to capture the fibrous extra-cellular matrix biophysical interactions, involving parameters of curvature, structural stiffness (N m(-1)), alignment and hierarchy, which have been shown to play key roles in disease and developmental biology. Using a highly aggressive glioma model (DBTRG-05MG), we present a platform technology to quantify single cell force modulation (both inside-out and outside-in) with and without the presence of a cytoskeleton altering drug (cytochalasin D) using suspended and aligned fiber networks (nanonets) beginning to represent the aligned glioma environment. The nanonets fused in crisscross patterns were manufactured using the non-electrospinning spinneret based tunable engineering parameters technique. We demonstrate the ability to measure contractile single cell forces exerted by glioma cells attached to and migrating along the fiber axis (inside-out). This is followed by a study of force response of glioma cells attached to two parallel fibers using a probe deflecting the leading fiber (outside-in). The forces are calculated using beam deflection within the elastic limit. Our data shows that cytochalasin D compromises the spreading area of single glioma cells, eventually decreasing their 'inside-out' contractile forces, and 'outside-in' force response to external strain. Most notably, for the first time, we demonstrate the feasibility of using physiologically relevant aligned fiber networks as ultra-sensitive force (∼nanoNewtons) probes for investigating drug response and efficacy in disease models at the single cell resolution.
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http://dx.doi.org/10.1088/1758-5082/6/4/045006DOI Listing
September 2014

siRNA delivery from triblock copolymer micelles with spatially-ordered compartments of PEG shell, siRNA-loaded intermediate layer, and hydrophobic core.

Biomaterials 2014 May 6;35(15):4548-56. Epub 2014 Mar 6.

Department of Materials Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan; Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan; Center for NanoBio Integration, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan. Electronic address:

Hydrophobized block copolymers have widely been developed for construction of polymeric micelles for stable delivery of nucleic acids as well as anticancer drugs. Herein, we elaborated an A-B-C type of triblock copolymer featuring shell-forming A-segment, nucleic acid-loading B-segment, and stable core-forming C-segment, directed toward construction of a three-layered polymeric micelle as a small interfering RNA (siRNA) vehicle. The triblock copolymer was prepared with nonionic and hydrophilic poly(ethylene glycol) (PEG), cationic poly(l-lysine) (PLys), and poly{N-[N-(2-aminoethyl)-2-aminoethyl]aspartamide} [PAsp(DET)] bearing a hydrophobic dimethoxy nitrobenzyl ester (DN) moiety in the side chain [PEG-PLys-PAsp(DET-DN)]. The resulting triblock copolymers spontaneously formed sub-100 nm-sized polymeric micelles with a hydrophobic PAsp(DET-DN) core as well as PEG shell in an aqueous solution. This micelle was able to incorporate siRNA into the intermediate PLys layer, associated with slightly reduced size and a narrow size distribution. The triblock copolymer micelles (TCMs) stably encapsulated siRNA in serum-containing medium, whereas randomly hydrophobized triblock copolymer [PEG-PLys(DN)-PAsp(DET-DN)] control micelles (RCMs) gradually released siRNA with time and non-PEGylated diblock copolymer [PLys-PAsp(DET-DN)] control micelles (DCMs) immediately formed large aggregates. The TCMs thus induced appreciably stronger sequence-specific gene silencing in cultured cancer cells, compared to those control micelles. The siRNA delivery with TCMs was further examined in terms of cellular uptake and intracellular trafficking. The flow cytometric analysis revealed that the cellular uptake of TCMs was more efficient than that of RCMs, but less efficient than that of DCMs. The intracellular trafficking study using confocal laser scanning microscopy combined with fluorescence resonance energy transfer (FRET) revealed that the TCMs could readily release the siRNA payload within cells, which was in contrast to the DCMs exhibiting much slower release profile. This result indicates that PEG shell contributed to the smooth release of siRNA from TCMs within the cells, presumably due to avoiding irreversible aggregate formation. The obtained results demonstrated that the design of separately functionalized polymer segments expanded the performance of polymeric micelles for successful siRNA delivery.
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http://dx.doi.org/10.1016/j.biomaterials.2014.02.016DOI Listing
May 2014
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