Tenascin-C-enriched regeneration-specific extracellular matrix guarantees superior muscle regeneration in Ambystoma mexicanum.

Severe muscle injury causes distress and difficulty in humans. Studying the high regenerative ability of the axolotls may provide hints for the development of an effective treatment for severe injuries to muscle tissue. Here, we examined the regenerative process in response to a muscle injury in axolotls. We found that axolotls are capable of complete regeneration in response to a partial muscle resection called volumetric muscle loss (VML), which mammals cannot perfectly regenerate. We investigated the mechanisms underlying this high regenerative capacity in response to VML, focusing on the migration of muscle satellite cells and the extracellular matrix (ECM) formed during VML injury. Axolotls form tenascin-C (TN-C)-enriched ECM after VML injury. This TN-C-enriched ECM promotes the satellite cell migration. We confirmed the importance of TN-C in successful axolotl muscle regeneration by creating TN-C mutant animals. Our results suggest that the maintenance of a TN-C-enriched ECM environment after muscle injury promotes the release of muscle satellite cells and supports eventually high muscle regenerative capacity. In the future, better muscle regeneration may be achieved in mammals through the maintenance of TN-C expression.

FGF signaling induces the regeneration of collagen fiber structure during skin wound healing in axolotls.

Axolotls have been considered to be able to regenerate their skin completely. Our recent study updated this theory with the finding that the lattice structure of dermal collagen fibers was not fully regenerated after skin injury. We also discovered that nerves induce the regeneration of collagen fibers. The mechanism of collagen fiber regeneration remains unknown, however. In this study, we focused on the structure of collagen fibers with collagen braiding cells, and cell origin in axolotl skin regeneration. In the wounded dermis, cells involved in skin repair/regeneration were derived from both the surrounding dermis and the subcutaneous tissue. Regardless of cell origin, cells acquired the proper cell morphology to braid collagen fiber with nerve presence. We also found that FGF signaling could substitute for the nerve roles in the conversion of subcutaneous fibroblasts to lattice-shaped dermal fibroblasts. Our findings contribute to the elucidation of the fundamental mechanisms of true skin regeneration and provide useful insights for pioneering new skin treatments.

Refinements in the reconstruction of bisphosphonate-related osteonecrosis of the jaw.

The recommended treatment strategy for stage 3 bisphosphonate-related osteonecrosis of the jaw (BRONJ) is currently rigid plate fixation without bone reconstruction. However, a recent systematic review indicated the utility of microsurgical reconstruction after resection of BRONJ. Several types of flaps have been described but their applications are controversial. Here we present a detailed reconstruction plan for obtaining better outcomes in patients with maxillary and mandibular BRONJ. Given that progressive maxillary BRONJ is often invasive to the skin, including the eyelid, leading to functional loss such as leakage of discharge and ectropion, several revision surgeries are needed to increase the volume in the defect after the free flap transfer. For progressive mandibular BRONJ, hemi-mandibulectomy to subtotal mandibulectomy with an adequate margin from the necrotic bone is necessary, followed by a well-designed fibular free flap.

Bead Implantation and Delivery of Exogenous Growth Factors.

Genetic methods in axolotls (Ambystoma mexicanum) remain in their infancy which has hampered the study of limb regeneration. There is much room for advancement, especially with respect to spatiotemporal regulation of gene expression. Secreted growth factors play a major role in each stage of regeneration. The use of slow-release beads is one of the most effective methods to control the spatiotemporal expression of secretory gene products. The topical administration of secreted factors by slow-release beads may also prove effective for future applications in non-regenerative animals and for medical applications in humans, in which genetic methods are not available. In this chapter, we describe a methodology for using and implanting slow-release beads to deliver exogenous growth factors to salamanders.

Spinocerebellar ataxia type 17-digenic TBP/STUB1 disease: neuropathologic features of an autopsied patient.

Spinocerebellar ataxia (SCA) type 17-digenic TBP/STUB1 disease (SCA17-DI) has been recently segregated from SCA17, caused by digenic inheritance of two gene mutations - intermediate polyglutamine-encoding CAG/CAA repeat expansions (polyQ) in TBP (TBP41 - 49) and STUB1 heterozygosity - the former being associated with SCA17, and the latter with SCA48 and SCAR16 (autosomal recessive). In SCA17, most patients carry intermediate TBP41 - 49 alleles but show incomplete penetrance, and the missing heritability can be explained by a new entity whereby TBP41 - 49 requires the STUB1 variant to be symptomatic. The STUB1 gene encodes the chaperone-associated E3 ubiquitin ligase (CHIP) involved in ubiquitin-mediated proteasomal control of protein homeostasis. However, reports of the neuropathology are limited and role of STUB1 mutations in SCA17-DI remain unknown. Here we report the clinicopathologic features of identical twin siblings, one of whom was autopsied and was found to carry an intermediate allele (41 and 38 CAG/CAA repeats) in TBP and a heterozygous missense mutation in STUB1 (p.P243L). These patients developed autosomal recessive Huntington's disease-like symptoms. Brain MRI showed diffuse atrophy of the cerebellum and T2WI revealed hyperintense lesions in the basal ganglia and periventricular deep white matter. The brain histopathology of the patient shared features characteristic of SCA17, such as degeneration of the cerebellar cortex and caudate nucleus, and presence of 1C2-positive neurons. Here we show that mutant CHIP fails to generate the polyubiquitin chain due to disrupted folding of the entire U box domain, thereby affecting the E3 activity of CHIP. When encountering patients with cerebellar ataxia, especially those with Huntington's disease-like symptoms, genetic testing for STUB1 as well as TBP should be conducted for diagnosis of SCA17-DI, even in cases of sporadic or autosomal recessive inheritance.

Impact of different dose reduction criteria for anti-Xa direct oral anticoagulants on bleeding complications: A single center observational study.

Background: Each direct oral anticoagulant (DOAC) has different dose reduction criteria. Here, we evaluated the differences in the doses of three anti-Xa DOACs and clinical events based on the dose reduction criteria in patients with atrial fibrillation (AF). Methods: Consecutive AF patients prescribed with anti-Xa DOACs [rivaroxaban (Riva), apixaban (Apix), and edoxaban (Edox)] between April 2011 and May 2016 were retrospectively evaluated. The incidences of thromboembolic and bleeding events were evaluated by the end of December 2020, focusing on the dose proportion. Results: A total of 786 patients (72 ± 10 years old, 66.9% male) were enrolled in this study [Riva (n = 337), Apix (n = 239), and Edox (n = 210)]. The proportion of reduced dose prescriptions was significantly greater for Edox (79.2%) than Riva (38.7%) or Apix (31.9%). A Kaplan-Meier analysis showed that the incidence of minor bleeding was significantly higher in the Apix than other groups (p < .001), even after propensity score matching. The standard dose of Apix had significantly higher bleeding events than the other DOACs (p < .001). Moreover, 23.2% and 51.6% of the patients with a standard dose of Apix were fulfilled with the dose reduction criteria for Riva and Edox and had more minor bleeding events than the unfulfilled ones (p = .046). Conclusions: The patients with a standard dose of Apix had a higher incidence of minor bleeding events than the other dosages. A reduced dose of apixaban was not prone to being chosen because of the dose reduction criteria, which may have been associated with a higher minor bleeding rate in patients with Apix.

Lattice-patterned collagen fibers and their dynamics in axolotl skin regeneration.

The morphology of collagen-producing cells and the structure of produced collagen in the dermis have not been well-described. This lack of insights has been a serious obstacle in the evaluation of skin regeneration. We succeeded in visualizing collagen-producing cells and produced collagen using the axolotl skin, which is highly transparent. The visualized dermal collagen had a lattice-like structure. The collagen-producing fibroblasts consistently possessed the lattice-patterned filopodia along with the lattice-patterned collagen network. The dynamics of this lattice-like structure were also verified in the skin regeneration process of axolotls, and it was found that the correct lattice-like structure was not reorganized after simple skin wounding but was reorganized in the presence of nerves. These findings are not only fundamental insights in dermatology but also valuable insights into the mechanism of skin regeneration.

An approach for elucidating dermal fibroblast dedifferentiation in amphibian limb regeneration.

Urodele amphibians, Pleurodeles waltl and Ambystoma mexicanum, have organ-level regeneration capability, such as limb regeneration. Multipotent cells are induced by an endogenous mechanism in amphibian limb regeneration. It is well known that dermal fibroblasts receive regenerative signals and turn into multipotent cells, called blastema cells. However, the induction mechanism of the blastema cells from matured dermal cells was unknown. We previously found that BMP2, FGF2, and FGF8 (B2FF) could play sufficient roles in blastema induction in urodele amphibians. Here, we show that B2FF treatment can induce dermis-derived cells that can participate in multiple cell lineage in limb regeneration. We first established a newt dermis-derived cell line and confirmed that B2FF treatment on the newt cells provided plasticity in cellular differentiation in limb regeneration. To clarify the factors that can provide the plasticity in differentiation, we performed the interspecies comparative analysis between newt cells and mouse cells and found the Pde4b gene was upregulated by B2FF treatment only in the newt cells. Blocking PDE4B signaling by a chemical PDE4 inhibitor suppressed dermis-to-cartilage transformation and the mosaic knockout animals showed consistent results. Our results are a valuable insight into how dermal fibroblasts acquire multipotency during the early phase of limb regeneration via an endogenous program in amphibian limb regeneration.

Lmx1b activation in axolotl limb regeneration.

BACKGROUND: Axolotls can regenerate their limbs. In their limb regeneration process, developmental genes are re-expressed and reorganize the developmental axes, in which the position-specific genes are properly re-expressed. However, how such position specificity is reorganized in the regeneration processes has not been clarified. To address this issue, we focused on the reactivation process of Lmx1b, which determines the limb dorsal identity in many animals. RESULTS: Here, we show that Lmx1b expression is maintained in the dorsal skin before amputation and is activated after amputation. Furthermore, we demonstrate that only cells located in the dorsal side prior to limb amputation could reactivate Lmx1b after limb amputation. We also found that Lmx1b activation was achieved by nerve presence. The nerve factors, BMP2+FGF2+FGF8 (B2FF), consistently reactivate Lmx1b when applied to the dorsal skin. CONCLUSIONS: These results imply that the retained Lmx1b expression in the intact skin plays a role in positional memory, which instruct cells about the spatial positioning before amputation. This memory is reactivated by nerves or nerve factors that can trigger the entire limb regeneration process. Our findings highlight the role of nerves in amphibian limb regeneration, including both the initiation of limb regeneration and the reactivation of position-specific gene expression.

Fibrillation cycle length predicts cardiovascular events in patients with long-standing persistent atrial fibrillation.

BACKGROUND: Atrial fibrillation (AF) is associated with an increased risk of heart failure (HF), stroke, and death. Although fibrillation cycle length (FCL) is used as a surrogate for atrial refractoriness, its impact on outcomes remains unclear. This study aimed to identify predictors of cardiovascular events, including FCL, in patients with long-standing persistent AF. METHODS: The study included 190 consecutive patients with long-standing persistent AF (mean age 74 years, 74% male). Patients with valvular AF or hemodialysis-dependent end-stage renal disease and those on anti-arrhythmic drugs were excluded. The primary composite outcome was occurrence of cardiovascular events (myocardial infarction, HF), cerebrovascular events (stroke, transient ischemic attack), and all-cause death. FCL was calculated by fast Fourier transformation analysis of fibrillation waves in the surface electrocardiogram. RESULTS: Over a median follow-up of 2.6 years, the primary outcome occurred in 31 patients (cardiovascular events, n = 18; cerebrovascular events, n = 8; all-cause death, n = 5). In multivariate analysis, longer FCL and history of HF were independent predictors of these outcomes. In a Cox proportional hazards model adjusted for age, sex, and history of HF, patients with an FCL > 160 ms (cut-off determined by receiver-operating characteristic curve analysis) were at increased risk of the outcome (hazard ratio 12.9; 95% confidence interval 4.99-44.10; p < 0.001). CONCLUSIONS: FCL was independently associated with cardiovascular outcomes in patients with long-standing persistent AF.

Comparing nerve-mediated FGF signalling in the early initiation phase of organ regeneration across mutliple amphibian species.

Amphibians have a very high capacity for regeneration among tetrapods. This superior regeneration capability in amphibians can be observed in limbs, the tail, teeth, external gills, the heart, and some internal organs. The mechanisms underlying the superior organ regeneration capability have been studied for a long time. Limb regeneration has been investigated as the representative phenomenon for organ-level regeneration. In limb regeneration, a prominent difference between regenerative and nonregenerative animals after limb amputation is blastema formation. A regeneration blastema requires the presence of nerves in the stump region. Thus, nerve regulation is responsible for blastema induction, and it has received much attention. Nerve regulation in regeneration has been investigated using the limb regeneration model and newly established alternative experimental model called the accessory limb model. Previous studies have identified some candidate genes that act as neural factors in limb regeneration, and these studies also clarified related events in early limb regeneration. Consistent with the nervous regulation and related events in limb regeneration, similar regeneration mechanisms in other organs have been discovered. This review especially focuses on the role of nerve-mediated fibroblast growth factor in the initiation phase of organ regeneration. Comparison of the initiation mechanisms for regeneration in various amphibian organs allows speculation about a fundamental regenerative process.

Clinical study of treatment methods and associated factors in mandibular osteoradionecrosis.

This study aimed to determine the appropriate treatment methods and evaluate associated factors by comparing nonoperative treatment alone with a combination of both nonoperative and surgical treatment in 34 patients with mandibular osteoradionecrosis (mORN). The associated cure factors were analyzed by Cox regression. Propensity scores were calculated from factors that were not significant in the univariate analysis and used as covariates in the multivariate analysis. The cure rate among patients who received nonoperative and surgical treatment was higher than that observed with nonoperative treatment alone. Only the treatment method was associated with cure in both univariate and multivariate analyses.

Intentional Supernumerary Motor Phantom Limb after Right Cerebral Stroke: A Case Report.

A 47-year-old right-handed man was admitted to our hospital for rehabilitation after right basal ganglion hematoma. On day 57, he noticed a supernumerary motor phantom limb (SPL) involving his right arm, originating at the level of the elbow. The most notable finding of his SPL was the motor characteristic. When the subject had the intention to move the upper paralyzed limb simultaneously with the trainer's facilitating action, he said "there is another arm." The intention to move the paralyzed arm alone or passive movement of the paralyzed arm did not induce the SPL. He showed a severe left sensorimotor impairment and mild hemineglect, but no neglect syndromes of the body (e.g., asomatognosia, somatoparaphrenia, personification and misoplegia, or anosognosia) were observed. Brain MRI demonstrated a hematoma in the right temporal lobe subcortex, subfrontal cortex, putamen, internal capsule, and thalamus. Single-photon emission computed tomography images showed more widespread hypoperfusion in the right hemisphere in comparison to the lesions on MRI. However, the premotor cortex was preserved. Our case is different from Staub's case in that SPL was not induced by the intention to move the paralyzed limb alone; rather, it was induced when the patient intended to move the paralyzed limb with a trainer's simultaneous facilitating action. The SPL may reflect that an abnormal closed-loop function of the thalamocortical system underlies the phantom phenomenon. However, despite the severe motor and sensory impairment, the afferent pathway from the periphery to the premotor cortex may have been partially preserved, and this may have been related to the induction of SPL.

Canagliflozin Suppresses Atrial Remodeling in a Canine Atrial Fibrillation Model.

Background Recent clinical trials have demonstrated the possible pleiotropic effects of SGLT2 (sodium-glucose cotransporter 2) inhibitors in clinical cardiovascular diseases. Atrial electrical and structural remodeling is important as an atrial fibrillation (AF) substrate. Methods and Results The present study assessed the effect of canagliflozin (CAN), an SGLT2 inhibitor, on atrial remodeling in a canine AF model. The study included 12 beagle dogs, with 10 receiving continuous rapid atrial pacing and 2 acting as the nonpacing group. The 10 dogs that received continuous rapid atrial pacing for 3 weeks were subdivided as follows: pacing control group (n=5) and pacing+CAN (3 mg/kg per day) group (n=5). The atrial effective refractory period, conduction velocity, and AF inducibility were evaluated weekly through atrial epicardial wires. After the protocol, atrial tissues were sampled for histological examination. The degree of reactive oxygen species expression was evaluated by dihydroethidium staining. The atrial effective refractory period reduction was smaller (P=0.06) and the degree of conduction velocity decrease was smaller in the pacing+CAN group compared with the pacing control group (P=0.009). The AF inducibility gradually increased in the pacing control group, but such an increase was suppressed in the pacing+CAN group (P=0.011). The pacing control group exhibited interstitial fibrosis and enhanced oxidative stress, which were suppressed in the pacing+CAN group. Conclusions CAN and possibly other SGLT2 inhibitors might be useful for preventing AF and suppressing the promotion of atrial remodeling as an AF substrate.

Axolotl liver regeneration is accomplished via compensatory congestion mechanisms regulated by ERK signaling after partial hepatectomy.

BACKGROUND: Axolotls have remarkable organ-level regeneration capability. They can regenerate their limbs, tail, brain, gills, and heart. The liver had been considered to be a regenerative organ in these highly regeneration-competent animals. Therefore, no research had been performed on liver regeneration in urodele amphibians. In the present study, we focused on axolotl liver regeneration and found a unique regeneration mechanism compared with other vertebrates. RESULTS: Partial hepatectomy (PH) was performed to assess axolotl liver regeneration. Regeneration was assessed using block-face imaging (CoMBi), histology, cell proliferation, weight gain, and Albumin (Alb) + area. Axolotl liver histology was compared with other vertebrates. Axolotl liver consists of Glisson's capsule, sinusoids, and hepatic cord with no apparent lobule structures. Hepatocytes were mononucleated or multinucleated. PH increased the multinucleated hepatocytes and the Alb + area, but there was no apparent liver shape recovery even 40 days after PH. Gene expression pattern suggests that no epimorphic regeneration takes place. We also found that the increase in the number of proliferating hepatocytes was regulated by ERK-signaling. CONCLUSION: Our findings suggest that axolotls, which have epimorphic regeneration ability, regenerate their liver via unique mechanisms, compensatory congestion.

Additional posterior wall isolation is associated with gastric hypomotility in catheter ablation of atrial fibrillation.

BACKGROUND: Gastric hypomotility (GH) is a possible complication of catheter ablation (CA) for atrial fibrillation (AF). However, it is unclear which factors are associated with GH. We sought to elucidate the relationship between the CA procedure and GH. METHODS: The study population consisted of 254 patients who underwent CA for AF from November 2017 to October 2018. Finally, 119 patients were enrolled and divided into two groups: with or without GH (GH or non-GH groups). To evaluate the association with GH, the clinical backgrounds and procedure characteristics of the radiofrequency CA (RFCA) were compared between the two groups. RESULTS: The median age was 69 years old with 34% of female. GH were observed in 27.7% of patients who underwent RFCA, which was significantly higher than that in the cohort of patients who underwent esophago-gastro-duodenoscopy during the same time period (1.9%: 151 in 8063 patients, p < 0.0001). According to the detailed RFCA procedure, additional posterior wall isolation with pulmonary vein isolation (PVI) had a higher prevalence of GH than that with only PVI (54.8% vs. 18.2%; odds ratio 5.46, 95%CI 2.24-13.32, p = 0.0002). After an adjustment using a multivariate logistic analysis, a posterior wall isolation with the PVI was identified as the only independent predictor for GH (odds ratio 5.01, 95%CI 1.94-13.43, p = 0.0009). CONCLUSIONS: Additional posterior wall isolation with PVI was associated with gastric hypomotility.

Acute heart failure due to large intimal flap associated with chronic aortic dissection.

Acute heart failure from aortic stenosis secondary to chronic aortic dissection is very rare. We describe a case of acute heart failure secondary to aortic arch stenosis resulting from subacute type A aortic dissection. Resection of large thickened immobile intimal flap with total aortic arch replacement was successful, and cardiac function improved.

Neural regulation in tooth regeneration of Ambystoma mexicanum.

The presence of nerves is an important factor in successful organ regeneration in amphibians. The Mexican salamander, Ambystoma mexicanum, is able to regenerate limbs, tail, and gills when nerves are present. However, the nerve-dependency of tooth regeneration has not been evaluated. Here, we reevaluated tooth regeneration processes in axolotls using a three-dimensional reconstitution method called CoMBI and found that tooth regeneration is nerve-dependent although the dentary bone is independent of nerve presence. The induction and invagination of the dental lamina were delayed by denervation. Exogenous Fgf2, Fgf8, and Bmp7 expression could induce tooth placodes even in the denervated mandible. Our results suggest that the role of nerves is conserved and that Fgf+Bmp signals play key roles in axolotl organ-level regeneration. The presence of nerves is an important factor in successful organ regeneration in amphibians. The Mexican salamander, Ambystoma mexicanum, is able to regenerate limbs, tail, and gills when nerves are present. However, the nervedependency of tooth regeneration has not been evaluated. Here, we reevaluated tooth regeneration processes in axolotls using a three-dimensional reconstitution method called CoMBI and found that tooth regeneration is nerve-dependent although the dentary bone is independent of nerve presence. The induction and invagination of the dental lamina were delayed by denervation. Exogenous Fgf2, Fgf8, and Bmp7 expression could induce tooth placodes even in the denervated mandible. Our results suggest that the role of nerves is conserved and that Fgf+Bmp signals play key roles in axolotl organ-level regeneration.

Unveiling the Dynamical Assembly of Magnetic Nanocrystal Zig-Zag Chains via In Situ TEM Imaging in Liquid.

The controlled assembly of colloidal magnetic nanocrystals is key to many applications such as nanoelectronics, storage memory devices, and nanomedicine. Here, the motion and ordering of ferrimagnetic nanocubes in water via liquid-cell transmission electron microscopy is directly imaged in situ. Through the experimental analysis, combined with molecular dynamics simulations and theoretical considerations, it is shown that the presence of highly competitive interactions leads to the formation of stable monomers and dimers, acting as nuclei, followed by a dynamic growth of zig-zag chain-like assemblies. It is demonstrated that such arrays can be explained by first, a maximization of short-range electrostatic interactions, which at a later stage become surpassed by magnetic forces acting through the easy magnetic axes of the nanocubes, causing their tilted orientation within the arrays. Moreover, in the confined volume of liquid in the experiments, interactions of the nanocube surfaces with the cell membranes, when irradiated at relatively low electron dose, slow down the kinetics of their self-assembly, facilitating the identification of different stages in the process. The study provides crucial insights for the formation of unconventional linear arrays made of ferrimagnetic nanocubes that are essential for their further exploitation in, for example, magnetic hyperthermia, magneto-transport devices, and nanotheranostic tools.

The low carriage prevalence of pneumococcus among community-dwelling older people: A cross-sectional study in Japan.

BACKGROUND: The carriage prevalence of pneumococcus among community-dwelling older adults is not fully understood, especially in superaged societies. Our purpose was to elucidate the carriage prevalence of pneumococcus in the upper respiratory tract among Japanese community-dwelling adults aged ≥65 years. METHODS: We conducted a cross-sectional study of generally healthy community-dwelling adults aged ≥65 years in Nagasaki city, Japan. Demographic and clinical data and nasopharyngeal, oropharyngeal and saliva samples were collected from February 21st, 2018, to December 17th, 2018. The specimens were tested by culture and molecular methods. RESULTS: Among a total of 504 enrolled participants, none were positive for pneumococcus by culture, and 22 were positive by PCR. The overall carriage prevalence was 4.4% (95% CI: 2.8-6.5%). The prevalence was highest in saliva samples, followed by oropharyngeal and nasopharyngeal samples. No demographic characteristics were associated with carriage prevalence, including age (4.7% among participants aged 65-74 years and 4.1% among those 75 years and older). Among the pneumococcal-positive participants, 18.2% were PCV13-covered serotypes. CONCLUSIONS: Our data suggest a low carriage prevalence of S. pneumoniae among community-dwelling older people in Japan.