Characterization of disease-specific alterations in metabolites and effects of mesenchymal stromal cells on dystrophic muscles.

Introduction: Duchenne muscular dystrophy (DMD) is a genetic disorder caused by mutations in the dystrophin-encoding gene that leads to muscle necrosis and degeneration with chronic inflammation during growth, resulting in progressive generalized weakness of the skeletal and cardiac muscles. We previously demonstrated the therapeutic effects of systemic administration of dental pulp mesenchymal stromal cells (DPSCs) in a DMD animal model. We showed preservation of long-term muscle function and slowing of disease progression. However, little is known regarding the effects of cell therapy on the metabolic abnormalities in DMD. Therefore, here, we aimed to investigate the mechanisms underlying the immunosuppressive effects of DPSCs and their influence on DMD metabolism. Methods: A comprehensive metabolomics-based approach was employed, and an ingenuity pathway analysis was performed to identify dystrophy-specific metabolomic impairments in the mdx mice to assess the therapeutic response to our established systemic DPSC-mediated cell therapy approach. Results and Discussion: We identified DMD-specific impairments in metabolites and their responses to systemic DPSC treatment. Our results demonstrate the feasibility of the metabolomics-based approach and provide insights into the therapeutic effects of DPSCs in DMD. Our findings could help to identify molecular marker targets for therapeutic intervention and predict long-term therapeutic efficacy.

Innsbruck-style Retromandibular Anterior Trans-parotid Approach for Condylar Fractures: A Retrospective Review of 39 Fractures.

The retromandibular anterior trans-parotid (RAT) approach and a triangular-positioned double mini-plate osteosynthesis (TDO) technique have been reported from Innsbruck Medical University. This minimally invasive technique involves direct visualization of the condyle and is associated with lower incidence of facial palsy. Methods: A retrospective review was performed on the RAT approach and TDO technique conducted by a surgeon and team at two hospitals in Tokyo during a period of 3 years and 10 months. Results: This technique was performed on 35 patients with 39 condylar fractures. Sixty-nine percent of cases were due to accidental fall, 17% to traffic accidents, and 9% to sports. Furthermore, 92% cases were condylar base fractures. Nighty-seven percent of cases achieved good occlusion. The mean maximum mouth opening was 49 ± 1.3 mm. Postoperatively, facial palsy developed in three patients (7.7%), and two of them developed Frey syndrome at approximately 2.5 years postoperatively (5.1%). All patients completely recovered within 3 months postoperatively. One case each of salivary fistula, visible scar, and condylar resorption was found (2.6%). No case of massive bleeding during surgery, hematoma, or TMJ pain after surgery was found. Conclusion: This technique could achieve good occlusion with low incidence of complications and could contribute to early social reintegration among patients.

Glutamate in the medium of Lactiplantibacillus plantarum FL-664 affects the production of IL-12(p40) on murine spleen cells.

Lactic acid bacteria (LAB) have been attracting attention for their effects on innate immunity, and therefore, it is required to develop an efficient culturing method while maintaining their functionality. In this study, first, we compared the growth and functionality of LAB cultured on food grade (FG) medium with those on standard LAB medium and found that LAB cultured in the FG medium were smaller in cell size with high yield and had a higher ability to induce IL-12(p40) production by murine spleen cells in vitro. Moreover, the higher the glutamate concentration in the medium, the smaller the cell size, and the higher the yield and the higher the ability to induce IL-12 production. Addition of glutamate to the culture medium changes the size of LAB and affects their ability to induce IL-12(p40) production. In conclusion, regulating the concentration of glutamate would be important in the efficient culturing of functional LAB.

Airway Compromise in Mandibular Fracture.

ABSTRACT: Airway compromise is life threatening complication in the maxillofacial fracture. However, the incident of the airway compromise following mandibular fracture was little known. Maxillofacial fractures during the past 3 years from 2017 to 2020 were investigated retrospectively. There were 154 patients with maxillofacial fractures were noted in the past 3 years. Mandibular fractures accounted for 93% of the maxillofacial fracture. The most common cause of injury was falls (75%), followed by sports (12%). Of these, 3 cases (1.9%) needed an airway management at the time of initial treatment, and all were mandibular fractures. The average age was 55.7 ±â€Š10 years and all were male. The cause of the injury was high-energy trauma including traffic accident and work-related injury. Endotracheal intubation was performed at the time of initial treatment. All cases were communicated mandibular fracture included symphysis and para-symphysis region, and the bone fragments including the genial tubercle were largely displaced inward.

Oligonucleotide delivery to antigen presenting cells by using schizophyllan.

Schizophyllan (SPG), a member of the ß-glucan family, can form novel complexes with homo-polynucleotides such as poly(dA) through hydrogen bonding between two main chain glucoses and the one nucleotide base. Dectin-1, one of the major receptors for ß-glucans, is known to be expressed on antigen presenting cells (APCs) such as macrophages and dendritic cells. This suggests that the above-mentioned complexes could deliver bound functional oligonucleotides (ODNs) including antisense (AS)-ODNs, small interfering RNA, and CpG-ODNs to the APCs. Analysis using a quartz crystal microbalance revealed that a complex consisting of SPG and dA60 with a phosphorothioate backbone was recognized by recombinant Dectin-1 protein. Treatment with this complex containing an AS-ODN for tumor necrosis factor alpha protected mice against lipopolysaccharide-induced hepatitis at a very low AS-ODN dose. Moreover, immunization with CpG-ODN/SPG complex and antigenic proteins induced potent antigen specific immune responses. The present review also represents peptide delivery by conjugation with dA60 and the preparation of a nanogel using DNA-DNA hybridization. These findings indicate that the delivery of a specific ODN using ß-glucans could be used for treating various diseases caused by APCs and for activating antigen specific immune responses.

Melanin concentration and depolarization metrics measurement by polarization-sensitive optical coherence tomography.

Imaging of melanin in the eye is important as the melanin is structurally associated with some ocular diseases, such as age-related macular degeneration. Although optical coherence tomography (OCT) cannot distinguish tissues containing the melanin from other tissues intrinsically, polarization-sensitive OCT (PS-OCT) can detect the melanin through spatial depolarization of the backscattered light from the melanin granules. Entropy is one of the depolarization metrics that can be used to detect malanin granules in PS-OCT and valuable quantitative information on ocular tissue abnormalities can be retrived by correlating entropy with the melanin concentration. In this study, we investigate a relationship between the melanin concentration and some depolarization metrics including the entropy, and show that the entropy is linearly proportional to the melanin concentration in double logarithmic scale when noise bias is corrected for the entropy. In addition, we also confirm that the entropy does not depend on the incident state of polarization using the experimental data, which is one of important attributes that depolarization metrics should have. The dependence on the incident state of polarization is also analyzed for other depolarization metrics.

Sulfonamide antibiotics inhibit RNAi by binding to human Argonaute protein 2 PAZ.

We found that sulfisomidine, a sulfonamide antibiotic, potently binds to the Piwi/Argonaute/Zwille (PAZ) domain of human Argonaute protein 2 and inhibits RNA interference (RNAi). To elucidate the effect on RNAi of strong affinity of the 3'-ends in small interfering RNA (siRNA) to the PAZ domain, chemically modified siRNAs bearing sulfisomidine at the 3'-end were synthesized.

Polarization-sensitive optical coherence tomography for estimating relative melanin content of autologous induced stem-cell derived retinal pigment epithelium.

Transplantation of autologous human induced pluripotent stem cell-derived retinal pigment epithelial (hiPSC-RPE) sheets is a promising therapy for age-related macular degeneration (AMD). As melanin content is a representative feature of healthy RPE, we used polarization-sensitive optical coherence tomography (PS-OCT) to estimate the relative melanin content of RPE in diseased and non-diseased area, and in human iPSC-RPE sheets in vitro and in vivo by evaluating the randomness of polarization (entropy). Two aged Japanese women, one with neovascular AMD that underwent transplantation of an autologous hiPSC-RPE cell sheet and another with binocular dry AMD, were selected for this study. Entropy value was minimal in cells containing no melanin, whereas that of human RPE and hiPSC-RPE sheets was high. En face entropy of the cultured hiPSC-RPE sheet was compared with its grey-scale photo and its values were found to be inversely correlated with the extent of absence of pigmentation in vitro. En face entropy maps were compared to colour fundus photographs, fundus autofluorescence images, and fluorescein angiography images from patients. Entropy values of intact and defective RPEs and of iPSC-RPE transplant areas were determined in vivo using PS-OCT B-scan images. PS-OCT was found to be applicable in the estimation of relative melanin content of cultured and transplanted RPEs in regenerative medicine.

Practical and reliable synthesis of 2',3',5',5″-tetradeuterated uridine derivatives.

Deuterated drugs are valuable in the fields of drug discovery and medicinal chemistry. 2',3',5',5″-tetradeuterated uridine derivatives were synthesised from 2,3,5,5'-selectively tetradeuterated ribose using Sajiki's H-D exchanged Ru/C-H2-D2O-NaOH system and silyl-Hilbert-Johnson methods. The total deuterium content of the tetradeuterated uridines was over 92% using either basic or acidic reaction conditions. These derivatives would be expected as building blocks for the synthesis of deuterium-substituted nucleic acid probes for tracking the pharmacokinetics of nucleic acid drugs.

Long-term results of photodynamic therapy or ranibizumab for polypoidal choroidal vasculopathy in LAPTOP study.

BACKGROUND/AIM: We previously reported that ranibizumab performed better on visual prognosis than photodynamic therapy (PDT) in a Ranibizumab (Lucentis) And Photodynamic Therapy On Polypoidal choroidal vasculopathy (LAPTOP) study. To determine if the first-choice treatment, either PDT or intravitreal ranibizumab, has a long-term effect in patients with polypoidal choroidal vasculopathy (PCV). METHODS: We reviewed medical records of patientsrandomised to either PDT (29 eyes) or ranibizumab (27 eyes) from July 2009 to June 2011 in LAPTOP study. Retreatment or switching to other treatments were at the investigator's discretion after release from the 2-year LAPTOP study up to 5years. We evaluated visual acuity (VA), continuity of initial treatment, percentage of dry macula achievement and macular atrophy at 5 years. RESULTS: The logarithm of minimal angle of resolution VA was 0.56 in the PDT and 0.44 in the ranibizumab groups at baseline (p=0.101) and was 0.55 and 0.28 at 5years, respectively (p<0.05). More than 70% of the patients converted to aflibercept in following years. Achievement percentages of dry macula were 74% (PDT) and 63% (ranibizumab) at 5years, and macular atrophy was detected in 78% (PDT) and 60% (ranibizumab) with a mean area of 7.7 and 3.5 mm2, respectively (p=0.155). CONCLUSIONS: The better VA in the initial ranibizumab group compared with the PDT group at 2 years was retained at the 5-year follow-up.

Designing an immunocyte-targeting delivery system by use of beta-glucan.

A ß-1,3-d-glucan called Schizophyllan (SPG) can form a novel complex with homo oligodeoxynucleotides (ODNs) via the combination of hydrogen bonding and hydrophobic interactions. Dectin-1 is a major receptor involved in the recognition of ß-1,3-d-glucans and expressed on antigen presenting cells (APCs) including macrophages, dendritic cells, monocytes, neutrophils, and a subset of T cells. Therefore, the SPG/ODN complex can be used as APCs cell-specific delivery of functional ODNs including unmethylated CpG sequences (CpG-ODNs). In fact, CpG-ODN/SPG complex induced high antibody titers when it was administered to cynomolgus monkeys as adjutant of influenza vaccine. These results indicate that SPG can be an excellent immunocyte-targeting drug delivery system.

Incidence of outer retinal tubulation in eyes with choroidal neovascularization under intravitreal anti-vascular endothelial growth factor therapy in a Japanese population.

PURPOSE: The purpose of this study was to investigate the occurrence of outer retinal tubulation (ORT) among patients with different types of choroidal neovascularization (CNV) over time. MATERIALS AND METHODS: In this retrospective chart review, disease type was classified as typical age-related macular degeneration (t-AMD), polypoidal choroidal vasculopathy (PCV), retinal angiomatous proliferation (RAP), or myopic CNV (mCNV). Spectral domain-optical coherence tomography (SD-OCT) images were evaluated for the appearance of ORT and subretinal fibrosis and fluid. Furthermore, the association of the presence of ORT with clinical data and OCT findings was investigated. RESULTS: Among the 136 eyes studied, the overall rates of occurrence of ORT were 7.8%, 18.8%, and 31.6% after 12, 24, and 36 months from baseline, respectively. Among patients with t-AMD, RAP, and mCNV, the occurrence of ORT increased soon after the initial visit. In contrast, among patients with PCV, the occurrence of ORT increased slowly over time. Patients with and without ORT - ORT(+) and ORT(-) groups, respectively - differed significantly in terms of sex ratio and presence of intraretinal fluid at the initial visit and presence of subretinal fibrosis at 3 years from baseline. The ORT(+) group exhibited lower visual acuity (VA; 0.67±0.43) than that of the ORT(-) group (0.41±0.36; P<0.001). CONCLUSION: The occurrence of ORT tended to increase more slowly among eyes diagnosed with PCV than among eyes with other types of CNV.

Response of eyes with age-related macular degeneration to anti-VEGF drugs and implications for therapy planning.

PURPOSE: To evaluate the response to and dependence on aflibercept or ranibizumab in patients with age-related macular degeneration (AMD). METHODS: We retrospectively reviewed AMD patients who received induction therapy with aflibercept or ranibizumab for the following parameters: whether complete resolution of the retinal fluid ("good response") was achieved and whether recurrence was observed within 3 months ("dependent") after the induction treatment. With aflibercept treatment, treatment-naïve eyes with a good response/non-dependence were recommended a pro re nata regimen, and other eyes were recommended a proactive bimonthly regimen, followed by monitoring of visual acuity (VA) for 12 months. The measured values of the groups were compared using one-way analysis of variance with Tukey's test to evaluate the difference between baseline and postinjection VA. RESULTS: Among the treatment-naïve eyes, 76% had a good response to aflibercept and 37% of these were aflibercept-dependent, while 58% had a good response to ranibizumab but 51% of these were ranibizumab-dependent. Among the eyes that converted from ranibizumab treatment, 92% of the good responders to ranibizumab with dependence and 76% of the poor responders on ranibizumab had a good response to aflibercept. With aflibercept treatment, the mean VA of treatment-naïve patients was significantly better than the baseline VA over 12 months (P<0.001), and the VA of the converted group improved significantly with proactive treatment and the improvement was continuously maintained from 6 to 12 months. CONCLUSION: The evaluation of response to and dependence on anti-vascular endothelial growth factor therapies in AMD was useful and practical in managing therapeutic protocols to obtain a good VA.

A two-component micelle with emergent pH responsiveness by mixing dilauroyl phosphocholine and deoxycholic acid and its delivery of proteins into the cytosol.

Providing appropriate pH responsiveness for drug delivery nanoparticles is one of the major issues in developing a new generation of delivery systems. This paper reports that, when phosphocholine and a bile acid were mixed, the resultant two-component micelle gained pH responsiveness, while the individual components did not show any such responsiveness. The pH responsiveness was shown to be determined by the chemical structure, especially the positions and chirality of the OH groups, of the bile acid, and the sensitivity was determined by the alkyl chain length of the phosphocholine. The best combination for evading endocytosis was dilauroyl phosphocholine (DLPC) and deoxycholic acid (DA). Small-angle X-ray scattering revealed that the pH responsiveness was related to the change of surface hydrophobicity, namely, decreasing pH led to protonation of the carboxylic acid, resulting in aggregation of the preceding micelles. We assume that particles that become hydrophobic in this way can start interacting with the endocytotic bilayer, which eventually leads to rupture of the endocytotic vesicle. This mechanism is well supported by the finding that fluorescein-conjugated ovalbumin proteins were transported into the cytosol when they were co-administered with DLPC/DA.

Restoration of Photoreceptor Outer Segments up to 24 Months after Pars Plana Vitrectomy in Patients with Diabetic Macular Edema.

PURPOSE: To evaluate the ellipsoid zone (EZ) and external limiting membrane (ELM) status on optical coherence tomography images in patients with diabetic macular edema (DME) before and after pars plana vitrectomy (PPV). DESIGN: Retrospective observational analysis. PARTICIPANTS: Sixty-one eyes of 49 patients with DME. METHODS: We retrospectively reviewed the spectral-domain optical coherence tomography images of 61 eyes of 49 patients with DME who were treated with PPV and evaluated the percentage disruption of the EZ and ELM before and up to 24 months after PPV. MAIN OUTCOME MEASURES: Longitudinal changes in visual acuity (VA), central retinal thickness (CRT), and EZ and ELM spectral-domain optical coherence tomography findings in patients with DME. RESULTS: The logarithm of the minimum angle of resolution VA was 0.477 before PPV and then improved to 0.372 and 0.344 at 12 and 24 months after PPV, respectively (P < 0.005 and P < 0.001, respectively). The CRT was 505.5 µm before PPV and then decreased gradually to 345.0 and 301.4 µm at 12 and 24 months, respectively (P < 0.001 for both). The percentage disruption of the EZ was 29.3% and ELM was 9.6% before PPV and then deteriorated to 41.6% (P < 0.001) and 15.1% (P < 0.05), respectively, 1 month after PPV. The disruption of the EZ and ELM decreased gradually. The percentage disruption of the ELM recovered but did not improve by 24 months, compared with baseline (11.6%; P = 0.875), whereas the percentage disruption of the EZ recovered to baseline by 6 months (29.4%; P = 1.000) and then decreased significantly at 24 months (21.3%; P < 0.05). CONCLUSIONS: Both the EZ and ELM deteriorated after PPV. Whereas the ELM recovered but did not improve by 24 months when compared with baseline, the EZ and VA improved up to 24 months.

Adjuvant Activity Enhanced by Cross-Linked CpG-Oligonucleotides in ß-Glucan Nanogel and Its Antitumor Effect.

Cancer vaccine has the ability to directly eradicate tumor cells by creating and activating cytotoxic T lymphocytes (CTLs). To achieve efficient CTL activity and to induce Th1 responses, it is essential to administer an appropriate adjuvant as well as an antigen. CpG-ODN is known as a ligand of Toll-like receptor 9 (TLR9) and strongly induces Th1 responses. In our previous study, we developed a CpG-ODN delivery system by use of the formation of complexes between ODN and a ß-glucan SPG, denoted as CpG/SPG, and demonstrated that CpG/SPG induces high Th1 responses. In this study, we created a nanogel made from CpG/SPG complexes through DNA-DNA hybridization (cross-linked (CL)-CpG). Immunization with CL-CpG induced much stronger antigen-specific Th1 responses in combination with the antigenic protein ovalbumin (OVA) than that with CpG/SPG. Mice preimmunized with CL-CpG and OVA exhibited a long delay in tumor growth and an improved survival rate after tumor inoculation. These immune inductions can be attributed to the improvement of cellular uptake by the combination of increased size and the cluster effect of the ß-glucan recognition site in the nanogel structure. In other words, the particle nature of CL-CpG, instead of the semiflexible rod conformation of CpG/SPG, enhanced the efficacy of a cancer vaccine. The present results indicate that CL-CpG can be used as a potent vaccine adjuvant for the treatment of cancers and infectious diseases.

Changes in Legal Aspects of Public Education about Drugs and Their Importance.

Requirements for education on proper use of drugs were included in the junior high school educational guideline in 2012 incorporating pharmaceutical education in the obligatory school curriculum. This move is closely related to the country's new OTC drug marketing system. The amendment of the Pharmaceutical Affairs Law (PAL) in 2013 highlighted the public's own role in "promoting proper use of drugs and other related products and making greater efforts to acquire knowledge and improve understanding of their effective and safe use". Furthermore, the Law to Amend the PAL and Pharmacists Law enforced in 2014 allowed all OTC drugs to be sold online under appropriate rules. Deregulation of online sale of OTC drugs is expected further to promote self-medication for minor illnesses and require stricter measures to ensure people's safety through their proper use. These legal amendments in recent years have made people's education about proper use of drugs one of the top priorities Japan should pursue at state level. Since 2000, the author has been offering education on drugs to early primary school children as part of their healthcare education program. In the future, dedicated education on drugs will be necessary for people of all ages including not only school children but also their guardians and elderly citizens as well.

Gene silencing using a conjugate comprising Tat peptide and antisense oligonucleotide with phosphorothioate backbones.

Antisense oligonucleotides (ASOs) have a great therapeutic potential for the modulation of gene expression because of the high specificity. The major obstacles for clinical application are enzymatic degradation and low uptake into cells in vivo. In this study, we prepared the conjugate comprising Tat peptide and ASO with phosphorothioate linkages in a simple manner; azide alkyne Huisgen cycloaddition using a copper catalyst. The obtained conjugate showed a high stability in serum, compared with the conjugate with phosphodiester linkages. The conjugates with antisense for c-myb that is transcriptional factor concerning cell growth inhibited the cell proliferation in a dose dependent manner sequence-specifically. These findings suggest Tat-mediated ASOs delivery is useful for the treatment of various diseases.

Predictive factors for recurrence of macular edema after successful intravitreal bevacizumab therapy in branch retinal vein occlusion.

PURPOSE: Our aim was to investigate predictive factors associated with efficacy and recurrence after intravitreal bevacizumab (IVB) therapy for macular edema (ME) in patients with branch retinal vein occlusion (BRVO). METHODS: Fifty-two eyes of 52 patients who underwent IVB as a primary treatment against ME associated with BRVO were included retrospectively. Based on the postoperative central retinal thickness (CRT), the patients were classified into two groups: an effective group in which the CRT decreased to ≤250 µm within postoperative 3 months and an ineffective group in which the CRT remained >250 µm throughout the first 3 months. The effective group was then divided into two subgroups: a recurrent group in which ME had once resolved but recurred afterward, and a nonrecurrent group in which the resolution of ME was maintained throughout the follow-up period without additional injections. Preoperative factors such as age, gender, estimated elapsed time from disease onset to IVB, visual acuity, and CRT were compared between groups. RESULTS: There was no significant difference between effective (n = 37) and ineffective (n = 15) groups in all preoperative factors. Between recurrent (n = 26) and nonrecurrent (n = 11) groups, elapsed time was significantly different (29.7 ± 29.5 vs. 15.7 ± 8.9 weeks, respectively; P = 0.036), and there were no significant differences in the remaining factors. CONCLUSIONS: Early IVB treatment against BRVO may suppress ME recurrence.