Hanseul Oh

Spinal cord fusion with PEG-GNRs (TexasPEG): Neurophysiological recovery in 24 hours in rats

Background: The GEMINI spinal cord fusion protocol has been developed to achieve a successful cephalosomatic anastomosis. Here, for the first time, we report the effects of locally applied water-soluble, conductive PEG(polyethylene glycol)ylated graphene nanoribbons (PEG-GNRs) on neurophysiologic conduction after sharp cervical cord transection in rats. PEG-GNRs were produced by the polymerization of ethylene oxide from anion-edged graphene nanoribbons. These combine the fusogenic potential of PEG with the electrical conducting properties of the graphene nanoribbons. Methods: Laminectomy and transection of cervical spinal cord (C5) was performed on Female Sprague-Dawley (SD) rats. After applying PEG-GNR on the severed part, electrophysiological recovery of the reconstructed cervical spinal cord was confirmed by somatosensory evoked potentials (SSEPs) at 24 h after surgery. Results: While no SSEPs were detected in the control group, PEG-GNR treated group showed fast recovery of SSEPs at 24 h after the surgery. Conclusion: In this preliminary dataset, for the first time, we report the effect of a novel form of PEG with the goal of rapid reconstruction of a sharply severed spinal cord.

Immunohistochemical evidence of axonal regrowth across polyethylene glycol-fused cervical cords in mice

It is generally accepted that a severed spinal cord is associated with permanent paralysis. Recently, a spinal cord fusion protocol (GEMINI) has been proposed, whereby an acutely controlled, sharp, operative transection of the spinal cord is carried out. This scenario is not comparable (even in principle) to the clinical situation of a traumatic spinal cord injury, in which major tissue disruption (mechanical, hemorrhagic, scarand cyst-associated) occurs (Canavero, 2015). During 1950s–1960s, neurosurgeon Dr. Freeman made extensive observations of what happens when a spinal cord is sharply transected. He reported slow recovery of behavioral motor function in several animals over months (reviewed in Canavero et al., 2016), with clear signs of electrophysiological conduction. Most importantly, silver stained histologic sections showed numerous growing axons connecting the divided ends (Freeman, 1963). This recovery can be accelerated by treating the severed cord with a fusogen (e.g., polyethylene glycol) (see for full discussion and rationale from Canavero, 2013; Kim, 2016). Here, we clearly prove that axonal regrowth is possible across the severance interface using immunohistochemistry and electron microscopy. The experiment was approved by the Institutional Animal Care and Use Committee of the Konkuk University (Seoul, Korea). Our previous experiment reported a mild recovery of the limb to the level of a sweeping behavior without weight support at the 4 week after C5 laminectomy, equal to a Basso, Beattie and Bresnahan (BBB) locomotor scale score of 6–8 (Kim et al., 2016), and aiming at confirming the possibility of direct reconnection of a fully severed spinal cord. In this model, reproducing traumatic severance with a blade as happens after clinical stab wounds, no gap exists between the severed stumps. In this study, to confirm neuroregeneration across the contact interface, C5 laminectomy was performed in female ICR mice as described previously (Kim et al., 2016). After gently raising the cervical cord with a hook, complete severance was performed with surgical sharp blades #11. Polyethylene glycol (PEG MW 400, Sigma-Aldrich, St. Louis, MO, USA), a cell membrane fusogen (Ye et al., 2016)) was dripped onto the cut area. After the muscle and fascia were sutured and the skin closed, normal saline solution was provided with total parenteral nutrition (TPN, Chong Kun Dang, Korea) through the tail vein four times a day. All mice sacrificed at 4 weeks after operation showed partial behavior recovery as reported in a previous study (Kim et al., 2016). The spinal cervical cords were removed, frozen on dry ice and sectioned into longitudinal and transverse slices. Sections were fixed with 4% paraformaldehyde solution. Fixed sections were immunohistochemically stained for anti-neurofilament 200 (NF200; Sigma, 1:50), and mounted with 4′,6-diamidino-2-phenyindole (DAPI). Fluorescence was visualized using fluorescence or confocal microscopy. The transverse slices of fixed tissue were examined by transmission electron microscopy

GEMINI: Initial behavioral results after full severance of the cervical spinal cord in mice.

Background: The GEMINI spinal cord fusion protocol has been developed to achieve a successful cephalosomatic anastomosis. Here, we report the preliminary data on the use of a fusogen [polyethylene glycol (PEG)] after full cervical cord transection in mice to facilitate the fusion of both ends of a sharply transected spinal cord. Methods: Cervical laminectomy and a complete, visually confirmed cervical cord (C 5) transection was performed on female albino mice (n = 16). In Group 1 (n = 8), a fusogen, (PEG) was used to bridge the gap between the cut ends of the spinal cord. Group 2 received the same spinal cord transection but was treated with saline. Outcome was assessed daily using a standard scale (modified 22-point Basso-Beattie-Bresnahan scale) and filmed on camera. Results: The PEG group (group 1) showed partial restoration of motor function after 4 weeks of observation; group 2 (placebo) did not recover any useful motor activity. Conclusion: In this preliminary experiment, PEG, but not saline, promoted partial motor recovery in mice submitted to full cervical transection.

First detection of West Nile virus in domestic pigeon in Korea

West Nile virus (WNV) is a mosquito-borne zoonotic pathogen that has spread throughout Europe and the United States. Recently, WNV spread to East and Southeast Asia, and great efforts have been made in South Korea to prevent the spread of WNV from neighboring countries. In this study, we diagnosed the first case of WNV in pigeons (Columba livia domestica) residing in cities using a competitive enzyme-linked immunosorbent assay and confirmed it with nested reverse transcription polymerase chain reaction analysis and sequencing. This is the first report to provide convincing evidence that WNV is present within South Korea.

Detection and Molecular Characterization of Cryptosporidium spp. from Wild Rodents and Insectivores in South Korea

In order to examine the prevalence of Cryptosporidium infection in wild rodents and insectivores of South Korea and to assess their potential role as a source of human cryptosporidiosis, a total of 199 wild rodents and insectivore specimens were collected from 10 regions of South Korea and screened for Cryptosporidium infection over a period of 2 years (2012-2013). A nested-PCR amplification of Cryptosporidium oocyst wall protein (COWP) gene fragment revealed an overall prevalence of 34.2% (68/199). The sequence analysis of 18S rRNA gene locus of Cryptosporidium was performed from the fecal and cecum samples that tested positive by COWP amplification PCR. As a result, we identified 4 species/genotypes; chipmunk genotype I, cervine genotype I, C. muris, and a new genotype which is closely related to the bear genotype. The new genotype isolated from 12 Apodemus agrarius and 2 Apodemus chejuensis was not previously identified as known species or genotype, and therefore, it is supposed to be a novel genotype. In addition, the host spectrum of Cryptosporidium was extended to A. agrarius and Crosidura lasiura, which had not been reported before. In this study, we found that the Korean wild rodents and insectivores were infected with various Cryptosporidium spp. with large intra-genotypic variationa, indicating that they may function as potential reservoirs transmitting zoonotic Cryptosporidium to livestock and humans.

Glutathione Peroxidase 3 Inhibits Prostate Tumorigenesis in TRAMP Mice.

Glutathione peroxidase 3 (GPx3) is involved in protecting cells from oxidative damage, and down‐regulated levels of expression have been found in prostate cancer samples. We hypothesize that loss of the GPx3 increases the rate of prostate carcinogenesis and generated GPx3‐deficient transgenic adenocarcinoma of the mouse prostate (TRAMP) mice.

A Synthetic Tul4 and FopA Peptide Cocktail of Francisella tularensis Induces Humoral and Cell-Mediated Immune Responses in Mice.

Francisella tularensis is a highly virulent pathogen of humans and other mammals. Moreover, F. tularensis has been designated a category A biothreat agent, and there is growing interest in the development of a protective vaccine. In the present study, we determine the in vitro and in vivo immune responses of a subunit vaccine composed of recombinant peptides Tul4 and FopA from epitopes of the F. tularensis outer membrane proteins. The recombinant peptides with adjuvant CpG induced robust immunophenotypic change of dendritic cell (DC) maturation and secretion of inflammatory cytokines (IL-6, IL-12). In addition, the matured DCs enabled ex vivo proliferation of naive splenocytes in a mixed lymphocyte reaction. Lastly, we determined the in vivo immune response by assessment of antibody production in C57BL/ 6 mice. Total IgG levels were produced after immunization and peaked in 6 weeks, and moreover, Tul4-specific IgG was confirmed in the mice receiving peptides with or without CpG. Based on these results, we concluded that the recombinant peptides Tul4 and FopA have immunogenicity and could be a safe subunit vaccine candidate approach against F. tularensis.

Respiratory Toxicity of Polyhexamethylene Guanidine Phosphate Exposure in Zebrafish

Humidifier disinfectants containing polyhexamethylene guanidine phosphate (PHMG-P) can induce pulmonary toxicity and has caused human casualties in South Korea since 2006. Thereby, the safety evaluation of household chemicals such as PHMG-P has garnered increased importance. However, many limitations, such as the lack of specialized facilities and animal welfare concerns associated with the use of murine models, persist. Zebrafish gills have high functional and structural similarity to mammalian lungs. Moreover, zebrafish are sensitive to toxic substances, resulting in changes in behavioral or ventilatory patterns. Based on these facts, in this study, we aimed to evaluate the pulmonary toxicity of PHMG-P in zebrafish. Zebrafish exposed to PHMG-P showed an increase in mRNA levels of inflammatory factors persisting for 28 days along with histopathologic changes in the gills. An exposure time-dependent alteration in infiltration of inflammatory cells and destruction of gill lamellae was observed. In addition, an increase in mRNA levels of fibrosis factors was observed in gills exposed to PHMG-P for 28 days, as assessed by collagen staining with Massons trichrome. These results supported the cellular level results. Taken together, our results reveal pulmonary toxic effects of PHMG-P and suggest useful markers for evaluating pulmonary toxicity.

Troglitazone inhibits the migration and invasion of PC‑3 human prostate cancer cells by upregulating E‑cadherin and glutathione peroxidase 3

Troglitazone (TGZ) is a synthetic peroxisome proliferator-activated receptor γ (PPARγ) ligand that exhibits potential antitumor effects on a number of cancer subtypes, including prostate cancer. However, little is known about the effect of TGZ on metastasis in prostate cancer. The aim of the present study was to determine the inhibitory effect and mechanism underlying TGZ on cell growth, migration and invasion using the prostate cancer PC-3 cell line. Cellular migration and invasion were evaluated by performing a wound healing assay and Matrigel assay, respectively. The expression levels of mRNA and protein were determined by reverse transcription-quantitative polymerase chain reaction and western blotting. The results demonstrated that TGZ dose-dependently inhibited cell migration and invasion of PC-3 cells. The present study also revealed that TGZ increased the mRNA and protein levels of E-cadherin and glutathione peroxidase 3 (GPx3) in human prostate cancer PC-3 cells. In addition, GW9662, a PPARγ antagonist, attenuated the increased mRNA and protein levels of E-cadherin and GPx3, suggesting that the PPARγ-dependent signaling pathway was involved. Taken together, these results suggested that the anti-migration and anti-invasion effect of TGZ on PC-3 prostate cancer cells is, at least in part, mediated via upregulation of E-cadherin and GPx3. The present study also concluded that PPARγ may be used as a potential remedial target for the prevention and treatment of prostate cancer cell invasion and metastasis.

A case of active incomplete biliary cirrhosis in an aged female Japanese macaque (Macaca fuscata)

We describe the first case of biliary cirrhosis in Japanese macaque. Clinical signs had not been detected. The liver was nodular. Histopathologically, portal‐to‐portal pattern of fibrosis might have indicated chronic cholestasis. Fibrotic septa were infiltrated with inflammatory cells. Therefore, this case could be diagnosed as active incomplete biliary cirrhosis.