Clinical Outcomes of Surgical Treatments for Traumatic Spinal Injuries due to Snowboarding

Takahiro Masuda,Kei Miyamoto,Kazuhiko Wakahara,Kazu Matsumoto,Akira Hioki,Tetsuya Shimokawa,Katsuji Shimizu,Shinji Ogura,Haruhiko Akiyama 대한척추외과학회 2015 Asian Spine Journal Vol.9 No.1

Study Design: Retrospective study. Purpose: To assess treatment outcomes of snowboarding-related spinal and spinal cord injuries. Overview of Literature: Snowboarding-related spinal or spinal cord injury have a great impact on social and sporting activities. Methods: A retrospective review of 19 cases of surgically treated snowboard-related injury was done. Analyzed parameters included site of injury, type of fracture, peri- and postoperative complications, pre- and postoperative neurological status, activities of daily living, and participation in sports activities at the final follow-up. Results: The major site of injury was the thoracolumbar junction caused by fracture-dislocation (13/19 cases). The remaining 6 cases had cervical spine injuries. Over 60% of the patients had Frankel A and B paralysis. All patients were surgically treated by posterior fusion with instrumentation. Five underwent additional anterior fusion. Surgical outcome was restoration of ambulatory capacity in 12 patients (63.2%). Ultimately, 15 patients (78.9%) could return to work. Patients with complete paralysis upon admission showed reduced ambulatory capacity compared to those with incomplete paralysis. None of the patients again participated in any sports activities, including snowboarding. Conclusions: Snowboarding-related spinal or spinal cord injury has a great impact on social as well as sports activities. It is necessary to enhance promotion of injury prevention emphasizing the snowboarders’ responsibility code.

Production of a biologically active human basic fibroblast growth factor using silkworm-baculovirus expression vector system

Akitsu Masuda,Jian Xu,Kosuke Minamihata,Genki Kagawa,Yusei Hamada,Yoshiki Morifuji,Takumi Yano,Masato Hino,Daisuke Morokuma,Noriko Karasaki,Hiroaki Mon,Noriho Kamiya,Takahiro Kusakabe,이재만 한국응용곤충학회 2018 Journal of Asia-Pacific Entomology Vol.21 No.2

As a therapeutic treatment, recombinant human basic fibroblast growth factor (rhbFGF) is usually employed in tissue regeneration, and as an essential component in culture medium for maintaining the induced pluripotent stem (iPS) cell and embryonic stem (ES) cell in an undifferentiated state. Therefore, a large amount of biologically active rhbFGF is required. In this study, silkworm-baculovirus expression vector system (silkworm-BEVS) is employed to achieve a high productivity of recombinant rhbFGF with two small affinity tags (His-tag and STREP-tag) at the N or C-terminus. It is observed that rhbFGF with 30 K signal peptide of silkworm were successfully expressed but are not sufficiently secreted into the culture medium of cultured insect cells. Then we purified the N- or C-tagged intracellular rhbFGF protein and obtained a yield of about 0.7 mg/larva and 1.2 mg/ larva, respectively. Although the final yield of the C-tagged rhbFGF is higher than that of the N-tagged, rhbFGF with N-tag demonstrated promising and comparable biological activity, which is evaluated through a mammalian cell proliferation assay. Taken together, these results indicate that silkworm-BEVS could contribute to the mass-production of the biologically active rhbFGF for medical uses.

Improvement of Endo-β-N-acetylglucosaminidase H production using silkworm–baculovirus protein expression system

Atsushi Masuda,Jian Xu,TakumiMitsudome,Daisuke Morokuma,Hiroaki Mon,Yutaka Banno,Takahiro Kusakabe,이재만 한국응용곤충학회 2015 Journal of Asia-Pacific Entomology Vol.18 No.2

Endo-β-N-acetylglucosaminidase H (Endo H) catalyzes cleavage between the GlcNAc residues of the chitobiose core of N-linked glycans, leaving one GlcNAc residues attached to asparagine. Endo H cleaves high mannose and hybrid, but not complex, N-linked oligosaccharides on glycoproteins. Because of its unique specificity, Endo H is widely used for the structural and functional analyses of glycoproteins. In our previous study, the recombinant Endo H was produced as a secreted protein using silkworm–baculovirus expression system, but the yield was low (30 μg Endo H/10 ml larval hemolymph) compared to that of Escherichia coli. In this study, we purified active recombinant Endo H as an intracellular protein from fat body of silkworm infected with the recombinant baculovirus expressing Endo H without the exogenous signal peptide. Remarkably, the yield (9.3mgfrom20 silkwormlarvae)was about 310-fold higher than that secreted into larval hemolymph as reported previously. In addition,we screened the silkwormstrains maintained in Kyushu University and identified n17 as a high-level expression strain for Endo H.

Minimally Invasive Transtubular Endoscopic Decompression for L5 Radiculopathy Induced by Lumbosacral Extraforaminal Lesions

Ko Ikuta,Takahiro Kitamura,Keigo Masuda,Kensuke Hotta,Hideyuki Senba,Satoshi Shidahara 대한척추외과학회 2018 Asian Spine Journal Vol.12 No.2

Study Design: Retrospective study. Purpose: This study aimed to evaluate the efficacy of minimally invasive transtubular endoscopic decompression for the treatment of lumbosacral extraforaminal lesion (LSEFL). Overview of Literature: Conventional procedures for surgical decompression for the treatment of LSEFL involve certain technical challenges because the lumbosacral extraforaminal region has unique anatomical features. Moreover, the efficacy of minimally invasive procedures performed via the posterolateral approach for LSEFL has been reported. Methods: Twenty-five patients who had undergone minimally invasive transtubular endoscopic decompression for the treatment of LSEFL and could be followed up for at least 1 year postoperatively were enrolled. Five of these patients had a history of lumbar surgery, and seven had concomitant adjacent-level spinal stenosis. The clinical outcomes were evaluated using the Japanese Orthopaedic Association (JOA) lumbar score, numeric rating scale (NRS), and the JOA Back Pain Evaluation Questionnaire (JOABPEQ). The mean postoperative follow-up (FU) duration was 3.8 years. Results: All procedures could be completed without any severe surgical complications, and all patients could resume their previous activity level within 1 month postoperatively. The JOA score significantly increased from 14.1±4.0 at baseline to 23.1±3.7 at the 1-year FU and 22.1±3.8 at the last FU. Similarly, there were significant improvements in the postoperative NRS and JOABPEQ scores. An additional surgery was performed in two patients (8%) during the FU period. Patients with degenerative scoliosis exhibited significantly poorer outcomes compared with those without this condition. Conclusions: Transtubular endoscopic decompression can overcome certain technical challenges involved in the conventional procedures for LSEFL treatment; therefore, it can be recommended as a useful procedure for treating LSEFL. This procedure can provide some benefits to LSEFL patients and offer a well-illuminated surgical field and high surgical safety for the surgeon. However, the procedure should be carefully adapted for LSEFL patients with concomitant degenerative scoliosis.

Efficient production of recombinant T7 endonuclease I using silkwormbaculovirus expression vector system

Kakino Kohei,Masuda Akitsu,Hino Masato,Ebihara Takeru,Xu Jian,Mon Hiroaki,Fujita Ryosuke,Fujii Tsuguru,Kusakabe Takahiro,Lee Jae Man 한국응용곤충학회 2020 Journal of Asia-Pacific Entomology Vol.23 No.3

Recently, T7 Endonuclease I (T7E1) cleavage assay has been widely employed as an efficient approach for detecting mutations from CRISPR/Cas9 targeted samples. This enzyme is sufficient to detect single- and multiple-base mismatches from various heteroduplex DNA samples. However, T7E1 is quite expensive for researchers to use it only for screening mutations, especially in the condition of a large number of test samples. Regarding the production of this enzyme, to data, only the E. coli system has been reported and the highly overexpressed T7E1 seems toxic to the E. coli host cells. Thus, in this study, we tested whether the silkwormbaculovirus expression vector system (BEVS) is suitable to produce recombinant T7 Endonuclease I (rT7E1). The rT7E1 with N- or C-tags in cultured silkworm cells and silkworm pupae were successfully expressed. Our results demonstrated that the rT7E1-Ntag was highly expressed in silkworm pupae and we obtained rT7E1 proteins in high purity. Moreover, rT7E1 from silkworm-BEVS sufficiently recognized and cleaved the mismatches of designed and CRISPR/Cas9-mediated DNA substrates, which was equivalent to the commercial rT7E1 of the E. coli system. Taken together, our study would greatly support the genome-editing research by providing a cost-effective and active rT7E1 enzyme.

Acoustic Excitation of Jet Diffusion Flame ( Stability Limits and Flame Shapes )

Kazuhiro Yamane,Takahiro Hori,Yoshinori Nakajima,Manabu Hishida,Wataru Masuda 대한기계학회 1998 KSME/JSME THERMAL and FLUID Engineering Conference Vol.1 No.1

Percutaneous Transpedicular Interbody Fusion Technique in Percutaneous Pedicle Screw Stabilization for Pseudoarthrosis Following Pyogenic Spondylitis

Ko Ikuta,Keigo Masuda,Yutaka Yonekura,Takahiro Kitamura,Hideyuki Senba,Satoshi Shidahara 대한척추외과학회 2016 Asian Spine Journal Vol.10 No.2

This report introduces a percutaneous transpedicular interbody fusion (PTPIF) technique in posterior stabilization using percutaneous pedicle screws (PPSs). An 81-year-old man presented with pseudoarthrosis following pyogenic spondylitis 15 months before. Although no relapse of infection was found, he complained of obstinate low back pain and mild neurological symptoms. Radiological evaluations showed a pseudoarthrosis following pyogenic spondylitis at T11–12. Posterior stabilization using PPSs from Th9 to L2 and concomitant PTPIF using autologous iliac bone graft at T11–12 were performed. Low back pain and neurological symptoms were immediately improved after surgery. A solid interbody fusion at T11–12 was completed 9 months after surgery. The patient had no restriction of daily activity and could play golf at one year after surgery. PTPIF might be a useful option for perform segmental fusion in posterior stabilization using PPSs.

Optimal silkworm larva host for high-level production of Mus musculus IL-4 using a baculovirus expression vector system

Masahiko Kobayashi,Jian Xu,Kohei Kakino,Akitsu Masuda,Masato Hino,Naoki Fujimoto,Kosuke Minamihata,Noriho Kamiya,Hiroaki Mon,Hiroshi Iida,Masateru Takahashi,Takahiro Kusakabe,이재만 한국응용곤충학회 2020 Journal of Asia-Pacific Entomology Vol.23 No.1

Interleukine-4 (IL-4) is a cytokine that plays an important role in the immune system and recognized as a biological medicine. Therefore, there is a demand for the production of IL-4 with high performance. The expression of a recombinant IL-4 protein in the prokaryotic system usually results in the formation of an inclusion body. To date, the solution to obtain those active products without the refolding process remains to be established. In this study, we tried to acquire a biologically active recombinant Mus musculus IL-4 (rMmIL-4) using a silkworm-baculovirus expression vector system (silkworm-BEVS). We constructed two recombinant baculoviruses coding rMmIL-4 with the distinct location of affinity purification tags and succeeded in the expression and purification of rMmIL-4 proteins directly without the refolding process. Both purified proteins displayed comparable biological activity to the commercial proteins produced by the E. coli expression system. Besides, we performed screening of silkworm strains to seek optimal hosts for the mass-production of rMmIL-4. Intriguingly, we found that some silkworm strains showed significantly higher secretion levels of rMmIL-4 in silkworm sera. Our study provides meaningful insights into the industrial-scale production of rMmIL-4 with high productivity for pharmaceutical applications in the future.

Surgical Outcomes of Minimally Invasive Stabilization for Spinal Fractures in Patients with Ankylosing Spinal Disorders

Kazuhiro Kai,Ko Ikuta,Keigo Masuda,Takahiro Kitamura,Hideyuki Senba,Satoshi Shidahara 대한척추외과학회 2018 Asian Spine Journal Vol.12 No.3

Study Design: A retrospective study. Purpose: To evaluate the clinical and radiological outcomes of ankylosing spinal disorder (ASD) patients with spinal fractures treated by minimally invasive stabilization (MISt) using percutaneous pedicle screws (PPSs). Overview of Literature: ASDs, such as ankylosing spondylitis (AS) and diffuse idiopathic skeletal hyperostosis (DISH), increase susceptibility to spinal fractures because of extremely decreased spinal flexibility. Such fractures tend to be unstable and, consequently, should be treated with multiple-segmental internal fixation. However, conventional internal fixation procedures can severely damage the soft tissue, resulting in severe hemorrhage. Therefore, MISt is the preferred approach to treat spinal fractures in ASD patients. Methods: Nine ASD patients (four males and five females; three AS and six DISH patients) with spinal fractures who were treated by MISt using PPSs, were reviewed from April 2009 to August 2016. One patient died of aspiration pneumonia during follow-up (FU), and the remaining eight patients underwent clinical and radiological evaluation. Results: The mean age at surgery was 79.6 years (range, 68–95 years). The mean duration of postoperative FU was 14.2 months (range, 3–30 months). All treated fractures were anterior and posterior element injuries with distraction. Three patients presented delayed onset preoperative neurological deficit following trauma. The mean operation time was 179.6 minutes (range, 92–340 minutes). The mean hemorrhage was 103.6 mL (range, unquantifiable to 480 mL). Radiological evaluations at FU showed preservation of the acceptable postoperative correction of the fractured vertebra, as there were no re-collapses of the fractured vertebrae during FU. Conclusions: ASD patients must be acknowledged as highly susceptible to unstable spinal fractures, even after relatively mild trauma. MISt using PPSs may be an effective treatment for spinal fractures in such patients.

Production of an active Mus musculus IL-3 using updated silkworm-based baculovirus expression vector system

Nagai Ryo,Ebihara Takeru,Kakino Kohei,Masuda Akitsu,Xu Jian,Minamihata Kosuke,Kamiya Noriho,Kongkrongtong Tatphon,Kawahara Masahiro,Mon Hiroaki,Fujii Tsuguru,Kusakabe Takahiro,Lee Jae Man 한국응용곤충학회 2021 Journal of Asia-Pacific Entomology Vol.24 No.3

Due to the biological significance and therapeutic potential of Interleukin-3 (IL-3) secreted mainly by activated T cells, various protein expression systems have been challenged to produce recombinant IL3 to meet the increasing demands worldwide. Recently, we established an updated silkworm-based baculovirus expression vector system (silkworm-BEVS), which in most cases, produces eukaryotic proteins in biological or enzymatical active forms with considerable amounts. We attempted to reconstruct and express a recombinant mouse IL-3 (rMmIL-3) with C-terminal His8-Strep tags in silkworm-BEVS in the current study. From our results, we gained an active glycosylated rMmIL-3 protein in a substantial amount and quality. As compared with the E. coli expression system, silkworm-BEVS is a better choice regarding the glycosylations attached in rMmIL-3 and upscalable system in case that a commercial amount is required in the future. Collectively, our method shares an excellent model to produce interleukin molecular for approaching pharmaceutical applications.