Correction to: Comprehensive genetic analyses using targeted next-generation sequencing and genotype-phenotype correlations in 53 Japanese patients with osteogenesis imperfecta.

The original article has been corrected.

The risk of interstitial lung disease during biological treatment in Japanese patients with psoriasis.

BACKGROUND: With the increasing use of biological agents for the treatment of psoriasis, the numbers of patients with interstitial lung disease (ILD) associated with biologics have also increased. Many of these cases were associated with tumour necrosis factor (TNF)-α inhibitors, but cases associated with other families of biologics have also been reported in Japan. AIM: To analyse the background factors of patients who developed ILD, and to discuss better management of biological treatment. METHOD: We reviewed 246 patients with psoriasis who were treated with biological agents in our department to identify any pulmonary adverse events (AEs). Data on patients who developed ILD were extracted to analyse background factors, clinical type of psoriasis, time to onset of ILD, pre-existing ILD, smoking habit and prescribed drugs. RESULTS: Pulmonary AEs were seen in 22 cases, of which 11 were diagnosed as drug-induced ILD. The causative drugs were mainly TNF-α inhibitors, accounting for eight cases (six treated with infliximab, two with adalimumab). The remaining three cases were associated with secukinumab, ustekinumab and ixekizumab (n = 1 each). Notably, these three cases also had a history of drug-induced ILD. CONCLUSION: Patients with a history of drug-induced ILD seem to be more susceptible to developing another ILD induced by biologics, even if treated with interleukin-17 inhibitors. Thorough screening of risk factors and evaluation for eligibility, and careful monitoring during treatment are the best solutions to avoid serious pulmonary AE. Early detection and precise diagnosis of pulmonary AEs, especially differentiation from infectious diseases, is essential for managing biological treatment.

Randomized clinical trial of extensive intraoperative peritoneal lavage versus standard treatment for resectable advanced gastric cancer (CCOG 1102 trial).

BACKGROUND: A survival benefit of extensive intraoperative peritoneal lavage (EIPL) has been reported in patients with gastric cancer with positive peritoneal cytology. The hypothesis of this study was that EIPL may reduce peritoneal recurrence in patients with advanced gastric cancer who undergo surgery with curative intent. METHODS: This was an open-label, multi-institutional, randomized, phase 3 trial to assess the effects of EIPL versus standard treatment after curative gastrectomy for resectable gastric cancer of T3 status or above. The primary endpoint was disease-free survival (DFS); secondary endpoints were overall survival, peritoneal recurrence-free survival and incidence of adverse events. RESULTS: Between July 2011 and January 2014, 314 patients were enrolled from 15 institutions and 295 patients were analysed (145 and 150 in the EIPL and no-EIPL groups respectively). The 3-year DFS rate was 63·9 (95 per cent c.i. 55·5 to 71·2) per cent in the EIPL group and 59·7 (51·3 to 67·1) per cent in the control group (hazard ratio (HR) 0·81, 95 per cent c.i. 0·57 to 1·16; P = 0·249). The 3-year overall survival rate was 75·0 (67·1 to 81·3) per cent in the EIPL group and 73·7 (65·9 to 80·1) per cent in the control group (HR 0·91, 0·60 to 1·37; P = 0·634). Peritoneal recurrence-free survival was not significantly different between the two groups (HR 0·92, 0·62 to 1·36; P = 0·676). No intraoperative complications related to EIPL were observed. CONCLUSION: EIPL did not improve survival or peritoneal recurrence in patients who underwent gastrectomy for advanced gastric cancer. Registration number: 000005907 (http://www.umin.ac.jp/ctr/index.htm).

ANTECEDENTES: Se ha descrito que un lavado peritoneal extenso intraoperatorio (extensive intraoperative peritoneal lavage, EIPL) proporciona un beneficio en la supervivencia en pacientes con cáncer gástrico con citología peritoneal positiva. La hipótesis de este estudio era que el EIPL podría disminuir la recidiva peritoneal en pacientes con cáncer gástrico avanzado sometidos a cirugía con intención curativa. MÉTODOS: Ensayo clínico fase 3, abierto, multicéntrico y aleatorizado para evaluar los efectos de un lavado peritoneal extenso intraoperatorio (EIPL) frente a tratamiento estándar tras gastrectomía curativa por cáncer gástrico ≥T3 resecable. La variable de resultado primaria fue la supervivencia libre de enfermedad (disease-free survival, DFS), y las variables de resultado secundarias fueron la supervivencia global (overall survival, OS), la supervivencia libre de recidiva peritoneal y la incidencia de efectos adversos. RESULTADOS: Entre julio de 2011 y enero de 2014, se reclutaron 314 pacientes de 15 instituciones y se analizaron los datos de 295 pacientes (145 en el grupo con EIPL y 150 en el grupo sin EIPL). La DFS a los 3 años fue 63,9% (i.c. del 95% 55,5-71,2) en el grupo con EIPL y 59,7% (i.c. del 95% 51,3-67,1) en el grupo control (cociente de riesgos instantáneos, hazard ratio, HR 0,81 (i.c. del 95% 0,57-1,16), P = 0,249). La OS a los 3 años fue 75,0% (i.c. del 95% 67,1-81,3) en el grupo con EIPL y 73,7% (i.c. del 95% 65,9-80,1) en el grupo control (HR 0,91 i.c. del 95% 0,60-1,37), P = 0,634). No se observaron diferencias estadísticamente significativas entre los dos grupos en la supervivencia libre de recidiva peritoneal (P = 0,676, HR 0,92 (i.c. del 95% 0,62-1,36). No se observaron complicaciones intraoperatorias relacionadas con EIPL. CONCLUSIÓN: El EIPL no mejoró la supervivencia o la recidiva peritoneal en pacientes sometidos a gastrectomía por cáncer gástrico avanzado.

Comprehensive genetic analyses using targeted next-generation sequencing and genotype-phenotype correlations in 53 Japanese patients with osteogenesis imperfecta.

To elucidate mutation spectrum and genotype-phenotype correlations in Japanese patients with OI, we conducted comprehensive genetic analyses using NGS, as this had not been analyzed comprehensively in this patient population. Most mutations were located on COL1A1 and COL1A2. Glycine substitutions in COL1A1 resulted in the severe phenotype. INTRODUCTION: Most cases of osteogenesis imperfecta (OI) are caused by mutations in COL1A1 or COL1A2, which encode α chains of type I collagen. However, mutations in at least 16 other genes also cause OI. The mutation spectrum in Japanese patients with OI has not been comprehensively analyzed, as it is difficult to identify using classical Sanger sequencing. In this study, we aimed to reveal the mutation spectrum and genotype-phenotype correlations in Japanese patients with OI using next-generation sequencing (NGS). METHODS: We designed a capture panel for sequencing 15 candidate OI genes and 19 candidate genes that are associated with bone fragility or Wnt signaling. Using NGS, we examined 53 Japanese patients with OI from unrelated families. RESULTS: Pathogenic mutations were detected in 43 out of 53 individuals. All mutations were heterozygous. Among the 43 individuals, 40 variants were identified including 15 novel mutations. We found these mutations in COL1A1 (n = 30, 69.8%), COL1A2 (n = 12, 27.9%), and IFITM5 (n = 1, 2.3%). Patients with glycine substitution on COL1A1 had a higher frequency of fractures and were more severely short-statured. Although no significant genotype-phenotype correlation was observed for bone mineral density, the trabecular bone score was significantly lower in patients with glycine substitutions. CONCLUSION: We identified pathogenic mutations in 81% of our Japanese patients with OI. Most mutations were located on COL1A1 and COL1A2. This study revealed that glycine substitutions on COL1A1 resulted in the severe phenotype among Japanese patients with OI.

Development of the double level osteotomy in severe varus osteoarthritis showed good outcome by preventing oblique joint line.

INTRODUCTION: The purpose of the study was to describe the development of the surgical technique of double level osteotomy in patients with severe varus malalignment and to investigate the clinical and radiological outcome. It was hypothesized that good clinical results without a higher complication rate can be achieved by double level osteotomy to normalize joint angles and avoid joint line obliquity even in cases of progressed osteoarthritis. MATERIALS AND METHODS: Between 2011 and 2014, 33 patients (37 knees) undergoing double level osteotomies (open wedge HTO and closed wedge DFO) were included; of these, 24 patients (28 knees) were available in mean of 18 ± 10 months for the follow-up examination. Indication was symptomatic varus malalignment and medial compartment osteoarthritis. Postoperatively, these patients were assigned to 20 kg partial weight-bearing using two crutches for 6 weeks followed by full weight-bearing. No braces or casts were used. Full weight-bearing long leg anteroposterior radiographs were obtained preoperatively, after 6 weeks and at the time of final follow-up. Mechanical tibiofemoral angle (mTFA), mechanical lateral distal femoral angle (mLDFA) and medial proximal tibia angle (MPTA) were measured. Clinical outcome was evaluated using Lequesne-, Lysholm-, Oxford-, and IKDC-score at the time of follow-up. RESULTS: The preoperative mTFA of - 11 ± 3° increased to 0 ± 2° at final follow-up. The difference between mTFA-planning and final follow-up was - 2 ± 3° (p < 0.0006). At final follow-up, MPTA and mLDFA were 89.2 ± 2° and 87 ± 2°, respectively. The Lysholm, Oxford, Lequesne, and IKDC scores were 88 ± 13, 44 ± 3, 2 ± 2, and 77 ± 12, respectively. CONCLUSIONS: This study showed that double level osteotomy for the patients with severe varus malalignment and medial compartment osteoarthritis normalises the alignment, joint-angles, avoids joint line obliquity, and leads to good clinical results, despite progressive osteoarthritis. LEVEL OF EVIDENCE: Case series, Level IV.

IgA Antibodies Against Gliadin and Tissue Transglutaminase in Dogs With Chronic Enteritis and Intestinal T-Cell Lymphoma.

Molecular clonality analysis of T-cell receptor (TCR) genes for diagnosing T-cell lymphoma is widely used in veterinary medicine. However, differentiating chronic enteritis (CE) from intestinal lymphoma is challenging because of the incompatibility between histopathologic and clonality analysis results. On the basis of findings that canine intestinal T-cell lymphoma and celiac disease share some common features, we conducted serologic examinations in combination with histopathologic and T-cell receptor clonality analyses in 48 dogs diagnosed with either CE or intestinal lymphoma. Immunoglobulin A (IgA) and immunoglobulin G (IgG) antibodies against gliadin and tissue transglutaminase (tTG) were quantitatively measured using ELISA. The conditions were classified according to the histopathologic diagnosis, clonality analysis, and combined histopathologic/clonality analysis. Histopathologic analysis showed that dogs with intestinal lymphoma were likely to have high levels of serum IgA antibodies against gliadin and tTG, and serum IgG antibodies against tTG. No correlation between the diagnosed groups and control group was observed in the results of the clonality analysis and histopathologic/clonality analysis. It is interesting that dogs with intestinal lymphoma had a higher serum IgA titer against gliadin and tTG than did dogs with CE. These results suggest an association between repetitive inflammatory stimulation by gliadin peptides and subsequent intestinal lymphoma in dogs.

Enhanced regeneration potential of mobilized dental pulp stem cells from immature teeth.

OBJECTIVES: We have previously demonstrated that dental pulp stem cells (DPSCs) isolated from mature teeth by granulocyte colony-stimulating factor (G-CSF)-induced mobilization method can enhance angiogenesis/vasculogenesis and improve pulp regeneration when compared with colony-derived DPSCs. However, the efficacy of this method in immature teeth with root-formative stage has never been investigated. Therefore, the aim of this study was to examine the stemness, biological characteristics, and regeneration potential in mobilized DPSCs compared with colony-derived DPSCs from immature teeth. MATERIALS AND METHODS: Mobilized DPSCs isolated from immature teeth were compared to colony-derived DPSCs using methods including flow cytometry, migration assays, mRNA expression of angiogenic/neurotrophic factor, and induced differentiation assays. They were also compared in trophic effects of the secretome. Regeneration potential was further compared in an ectopic tooth transplantation model. RESULTS: Mobilized DPSCs had higher migration ability and expressed more angiogenic/neurotrophic factors than DPSCs. The mobilized DPSC secretome produced a higher stimulatory effect on migration, immunomodulation, anti-apoptosis, endothelial differentiation, and neurite extension. In addition, vascularization and pulp regeneration potential were higher in mobilized DPSCs than in DPSCs. CONCLUSIONS: G-CSF-induced mobilization method enhances regeneration potential of colony-derived DPSCs from immature teeth.

Variations in Histiocytic Differentiation of Cell Lines From Canine Cerebral and Articular Histiocytic Sarcomas.

Two newly established canine histiocytic sarcoma (HS) cell lines, designated as PWC-HS01 and FCR-HS02, were obtained from brain and articular tumors, respectively. These 2 HS cell lines had phagocytic ability and modal chromosome aberrations. Although morphologic features of both HS cells were similar, immunocytochemical examinations revealed that the PWC-HS01 cell line expressed both dendritic cell (ie, S100, CD208, CD1, and CD4) and macrophage (ie, CD68, CD163, and CD204) markers. In contrast, the FCR-HS02 cell line was immunonegative for CD204 and CD68 but consistently positive for the dendritic cell markers. Moreover, reverse transcription polymerase chain reaction analyses confirmed histiocytic differentiation of both HS cell lines. These results suggest that HS from the central nervous system may have a tendency to be more undifferentiated compared with cases from other organs. In addition, the 2 newly established HS cell lines were also tumorigenic and metastatic in immunodeficient mice, supporting that these cell lines can be used as new tumor models for investigating canine histiocytic diseases.

Deposition, Clearance, and Reinduction of Amyloid A Amyloid in Interleukin 1 Receptor Antagonist Knockout Mice.

Amyloid A (AA) amyloidosis is characterized by the extracellular deposition of AA amyloid and results in the irreversible dysfunction of parenchymal organs. In experimental models, AA amyloid deposits are cleared following a decrease in circulating serum amyloid A (SAA) concentrations. Additional inflammatory stimuli during this recovery process may induce more severe amyloid redeposition. In the present study, we confirmed the deposition, clearance, and reinduction of AA amyloid deposits in interleukin 1 receptor antagonist knockout mice (IL-1raKO) and studied the SAA levels and amyloid-enhancing factor activity based on the time-dependent changes of amyloid deposition. Histopathologically, following initial (day 0) injection of amyloid-enhancing factor in combination with an inflammatory stimulus (silver nitrate [AgNO3]), amyloid deposition peaked by day 20, and its deposition gradually decreased after day 35. SAA concentrations in serum were precipitously elevated on day 1 but returned to normal levels by day 10, whereas the SAA dimer was detected in serum after day 45. An additional AgNO3 injection was administered to mice with amyloidosis on day 5, 10, 35, or 50, and all mice developed large amyloid deposits. Amyloid deposition was most severe in mice treated with AgNO3 on day 35. The inoculation of sera from mice with AA amyloidosis, combined with AgNO3, induced AA amyloidosis. Serum samples collected on days 35 and 50, which contained high concentrations of the SAA dimer, induced amyloidosis in a high proportion (83%) of mice. Therefore, increased SAA and/or its dimer in serum during the recovery process may markedly exacerbate the development of AA amyloidosis.

Early full weight-bearing versus 6-week partial weight-bearing after open wedge high tibial osteotomy leads to earlier improvement of the clinical results: a prospective, randomised evaluation.

PURPOSE: Open wedge high tibial osteotomy is a widespread treatment option in patients with varus malalignment and medial compartment osteoarthritis. There is no standardised protocol for post-operative rehabilitation available. The purpose of this study was to compare two post-operative rehabilitation protocols and to evaluate the clinical outcome of early full weight-bearing after open wedge HTO. METHODS: One hundred and twenty consecutive patients with varus malalignment and medial compartment osteoarthritis received an open wedge HTO using an angular locking plate fixation between December 2008 and December 2011. All patients were assigned randomly into one of two groups with different post-operative rehabilitation protocols (11-day vs. 6-week 20-kg partial weight-bearing). Clinical outcome was evaluated using established instruments (Lequesne, Lysholm, HSS and IKDC scores) preoperatively, 6, 12 and 18 months post-operatively. Deformity analysis was performed preoperatively and during follow-up. RESULTS: All clinical scores showed a significant pre- to post-operative improvement. After 6 months, there was a higher improvement in the group of early full weight-bearing. The difference between preoperative and 6-month follow-up for the group with early full weight-bearing and for the group with 20-kg PWB for 6 weeks was 28 ± 26 and 18 ± 22, respectively, for the Lysholm score and -5.0 ± 5.1 and -3.0 ± 3.6, respectively, for the Lequesne score. CONCLUSIONS: Early full weight-bearing (11-day 20-kg partial weight-bearing) after open wedge HTO without bone graft leads to earlier improvement of the clinical results and can be recommended for post-operative rehabilitation after open wedge HTO and fixation with an angular locking plate. LEVEL OF EVIDENCE: Therapeutic study, Level I.

Pathology of the Aging Brain in Domestic and Laboratory Animals, and Animal Models of Human Neurodegenerative Diseases.

According to the WHO, the proportion of people over 60 years is increasing and expected to reach 22% of total world's population in 2050. In parallel, recent animal demographic studies have shown that the life expectancy of pet dogs and cats is increasing. Brain aging is associated not only with molecular and morphological changes but also leads to different degrees of behavioral and cognitive dysfunction. Common age-related brain lesions in humans include brain atrophy, neuronal loss, amyloid plaques, cerebrovascular amyloid angiopathy, vascular mineralization, neurofibrillary tangles, meningeal osseous metaplasia, and accumulation of lipofuscin. In aging humans, the most common neurodegenerative disorder is Alzheimer's disease (AD), which progressively impairs cognition, behavior, and quality of life. Pathologic changes comparable to the lesions of AD are described in several other animal species, although their clinical significance and effect on cognitive function are poorly documented. This review describes the commonly reported age-associated neurologic lesions in domestic and laboratory animals and the relationship of these lesions to cognitive dysfunction. Also described are the comparative interspecies similarities and differences to AD and other human neurodegenerative diseases including Parkinson's disease and progressive supranuclear palsy, and the spontaneous and transgenic animal models of these diseases.

Pathologic Features of Colorectal Inflammatory Polyps in Miniature Dachshunds.

The histopathologic characteristics of colorectal inflammatory polyps that formed in Miniature Dachshunds were compared with those of other colorectal proliferative lesions, including adenomas and adenocarcinomas. Fifty-three colorectal polypoid lesions were histopathologically classified into inflammatory polyps (26 cases), adenoma (18 cases), and adenocarcinoma (9 cases). All 26 dogs that were diagnosed with inflammatory polyps were Miniature Dachshunds, indicating that colorectal inflammatory polyps exhibit a marked predilection for this breed. The inflammatory polyps had complex histopathologic features and were classified into 3 stages based on their epithelial composition. In early stage (stage 1), the polyps tended to exhibit a thickened mucosa containing hyperplastic goblet cells, dilated crypts filled with a large amount of mucus, and mild lymphocyte and macrophage infiltration. In later stages (stages 2 and 3), more severe neutrophil infiltration, interstitial mucus accumulation, granulation tissue, and occasional osteoid tissue were seen. Also, a few small foci of dysplastic epithelial cells were detected. The hyperplastic goblet cells, which were a major component of the epithelium of the inflammatory polyps, were positive for cytokeratin 20 (CK20), while the dysplastic epithelial cells found in inflammatory polyps (stage 3) and the tumor cells of the adenomas and adenocarcinomas were negative for CK20. These CK20-negative epithelial cells exhibited cytoplasmic and nuclear immunoreactivity for beta-catenin. In addition, the epithelial cells in the inflammatory polyps demonstrated significantly higher cyclooxygenase 2 and fibroblast growth factor 2 expression than did those of the adenomas and adenocarcinomas, suggesting that the arachidonate cascade is involved in the development of colorectal inflammatory polyps in miniature dachshunds.

Expression of Autophagy-Related Proteins in the Spinal Cord of Pembroke Welsh Corgi Dogs With Canine Degenerative Myelopathy.

Canine degenerative myelopathy (DM) is a progressive neurodegenerative disease frequently found in Pembroke Welsh Corgi (PWC) dogs, and it has clinical and pathologic similarities to human amyotrophic lateral sclerosis. Autophagy is a major intracellular protein degradation system. Abnormalities of autophagy--resulting in cell death through mechanisms called type II programmed cell death--have recently been reported to occur in various neurodegenerative diseases, including amyotrophic lateral sclerosis. Thus, the distribution and expression levels of proteins involved in autophagy were examined in the spinal cords of 8 PWC dogs suffering from DM with superoxide dismutase mutation, 5 non-DM PWC dogs, and 6 Beagle dogs without neurologic signs. There was no significant difference in the ratio of neurons with microtubule-associated protein light chain 3 (LC3)-positive somata relative to those that were LC3 negative among the 3 groups, whereas the number of LC3-positive neurites was significantly increased in DM dogs. Punctate LC3 immunoreactivity did not colocalize with a lysosome marker, LAMP2 (lysosome-associated membrane protein 2). NBR1 (neighbor of BRCA gene 1) was localized mostly in reactive astrocytes, whereas there were p62 (p62/A170/SQSTM1)-positive foci in the neuropil of the spinal cord of DM dogs. Western blotting revealed in DM dogs the decreased expression of Beclin1 and Atg16 L, which are molecules involved in formation of the isolation membrane. These findings suggest that altered autophagosome degradation may result in LC3 and p62 accumulation in the DM spinal cord, whereas the early stage of membrane formation is likely to be downregulated.

Experimental transmission of AA amyloidosis by injecting the AA amyloid protein into interleukin-1 receptor antagonist knockout (IL-1raKO) mice.

The incidence of AA amyloidosis is high in humans with rheumatoid arthritis and several animal species, including cats and cattle with prolonged inflammation. AA amyloidosis can be experimentally induced in mice using severe inflammatory stimuli and a coinjection of AA amyloid; however, difficulties have been associated with transmitting AA amyloidosis to a different animal species, and this has been attributed to the "species barrier." The interleukin-1 receptor antagonist knockout (IL-1raKO) mouse, a rodent model of human rheumatoid arthritis, has been used in the transmission of AA amyloid. When IL-1raKO and BALB/c mice were intraperitoneally injected with mouse AA amyloid together with a subcutaneous pretreatment of 2% AgNO3, all mice from both strains that were injected with crude or purified murine AA amyloid developed AA amyloidosis. However, the amyloid index, which was determined by the intensity of AA amyloid deposition, was significantly higher in IL-1raKO mice than in BALB/c mice. When IL-1raKO and BALB/c mice were injected with crude or purified bovine AA amyloid together with the pretreatment, 83% (5/6 cases) and 38% (3/8 cases) of IL-1raKO mice and 17% (1/6 cases) and 0% (0/6 cases) of BALB/c mice, respectively, developed AA amyloidosis. Similarly, when IL-1raKO and BALB/c mice were injected with crude or purified feline AA amyloid, 33% (2/6 cases) and 88% (7/8 cases) of IL-1raKO mice and 0% (0/6 cases) and 29% (2/6 cases) of BALB/c mice, respectively, developed AA amyloidosis. These results indicated that IL-1raKO mice are a useful animal model for investigating AA amyloidogenesis.

Clinical and Pathologic Study of Feline Merkel Cell Carcinoma With Immunohistochemical Characterization of Normal and Neoplastic Merkel Cells.

The authors herein describe the morphologic and immunohistochemical features of normal Merkel cells as well as the clinicopathologic findings of Merkel cell carcinoma in cats. Merkel cells were characterized as vacuolated clear cells and were individually located in the epidermal basal layer of all regions examined. Clusters of Merkel cells were often observed adjacent to the sinus hair of the face and carpus. Immunohistochemically, Merkel cells were positive for cytokeratin (CK) 20, CK18, p63, neuron-specific enolase, synaptophysin, and protein gene product 9.5. Merkel cell carcinoma was detected as a solitary cutaneous mass in 3 aged cats (13 to 16 years old). On cytology, large lymphocyte-like cells were observed in all cases. Histologic examinations of surgically resected tumors revealed nests of round cells separated by various amounts of a fibrous stroma. Tumor cells were commonly immunopositive for CK20, CK18, p63, neuron-specific enolase, and synaptophysin, representing the characteristics of normal Merkel cells.

The origin and role of autophagy in the formation of cytoplasmic granules in canine lingual granular cell tumors.

Granular cell tumors (GCTs) are histologically characterized by polygonal neoplastic cells with abundant eosinophilic cytoplasmic granules. In humans, these cells are considered to be derived from Schwann cells, and the cytoplasmic granules are assumed to be autophagosomes or autophagolysosomes. However, the origin and nature of the cytoplasmic granules in canine GCTs have not been well characterized. The present study examined 9 canine lingual GCTs using immunohistochemistry, transmission electron microscopy (TEM), and cell culture and xenotransplantation experiments. In some cases, the tumor cells expressed S100, CD133, and desmin. The cytoplasmic granules were positive for LC3, p62, NBR1, and ubiquitin. TEM revealed autophagosome-like structures in the cytoplasm of the granule-containing cells. The cultured GCT cells were round to spindle shaped and expressed S100, nestin, Melan-A, CD133, LC3, p62, NBR1, and ubiquitin, suggesting that they were of neural crest origin, redifferentiated into melanocytes, and exhibited upregulated autophagy. The xenotransplanted tumors consisted of spindle to polygonal cells. Only a few cells contained cytoplasmic granules, and some had melanin pigments in their cytoplasm. The xenotransplanted cells expressed S100, nestin, Melan-A, and CD133. P62 and ubiquitin were detected, regardless of the presence or absence of cytoplasmic granules, while LC3 and NBR1 were detected only in the neoplastic cells containing cytoplasmic granules. These findings suggest that some xenotransplanted cells redifferentiated into melanocytes and that autophagy was upregulated in the cytoplasmic granule-containing cells. In conclusion, canine lingual GCTs originate from the neural crest and develop cytoplasmic granules via autophagy. In addition, the microenvironment of GCT cells affects their morphology.

Prophylactic intra-arterial administration of fasudil hydrochloride for vasospasm following subarachnoid haemorrhage.

PURPOSE: We evaluated patients treated with prophylactic intra-arterial administration of fasudil hydrochloride (IAF) after subarachnoid haemorrhage (SAH). MATERIALS AND METHODS: Between August 1998 and December 2012, 92 patients with aneurysmal SAH were treated with IAF for angiographic vasospasm without ischemic symptoms after their follow-up angiography. Patients comprised 50 women and 42 men, aged 24-83 (mean 56.6) years. IAF consisted of 15 mg of fasudil hydrochloride dissolved in 20 ml physiological saline and injected through a catheter during approximately 15 min, after diagnostic angiography. The clinical outcome was evaluated using the Glasgow Outcome Scale (GOS) at discharge and ischemic lesions resulting from vasospasm were assessed on computed tomography (CT) scan at discharge. RESULTS: Forty-eight patients underwent surgical clipping and 44 patients underwent endovascular coiling. Angiographic improvement was observed in all patients (100 %). At discharge, 76 (83.0 %) of 92 patients showed good recovery on GOS. Nine patients developed progression of delayed ischemic neurological deficits (DIND) and three of these patients had ischemic lesions on CT scans. No patient had any significant changes in vital signs or any other adverse effects resulting from IAF. CONCLUSION: IAF therapy was safe and effective for patients with vasospasm following SAH. Prophylactic IAF therapy may prevent symptomatic vasospasm.

Overexpression of nucleostemin contributes to an advanced malignant phenotype and a poor prognosis in oral squamous cell carcinoma.

BACKGROUND: Nucleostemin (NS) is essential for the maintenance of stem cell properties, the functions of which remain poorly understood in cancer cells. The purpose of this study was to explore the impact of NS on malignancy and its clinical significance in oral squamous cell carcinoma (OSCC) patients. METHODS: We investigated the effects of NS on the proliferation and invasion of OSCC using NS-overexpressing or -knockdown OSCC cells. We assessed the activation of the STAT3 (signal transducer and activator of transcription 3) signalling pathway and the downstream targets in the cells with different expression levels of NS. An immunohistochemical analysis of NS was also performed in 54 OSCC patients who were treated with preoperative chemoradiotherapy and surgery. RESULTS: The overexpression of NS significantly enhanced the proliferation and invasive potential of OSCC cells. On the other hand, downregulation of NS suppressed the invasiveness of the cells. The alterations of these malignant phenotypes were associated with the activation of STAT3 signalling and its downstream targets. An immunohistochemical analysis demonstrated that a high NS tumour expression level significantly correlated with an advanced T-stage and N-stage. Furthermore, a Cox regression analysis revealed that the NS status (hazard ratio, 9.09; P=0.002) was a significant progression factor for OSCC patients. CONCLUSIONS: Our results suggest that targeting NS may provide a promising treatment for highly malignant OSCC.

The effects of tumor location on diagnostic criteria for canine malignant peripheral nerve sheath tumors (MPNSTs) and the markers for distinction between canine MPNSTs and canine perivascular wall tumors.

Canine malignant peripheral nerve sheath tumors (MPNSTs) occur not only in the peripheral nervous system (PNS) but also in soft tissue and various organs (non-PNS). The most important diagnostic criterion is proof of peripheral nerve sheath origin. This is difficult in non-PNS MPNSTs, and its differential diagnosis is challenging. Canine perivascular wall tumors (PWTs) also commonly arise in soft tissue. Their histopathological features are quite similar to those of canine MPNSTs, making their differential diagnosis challenging. To elucidate whether the morphological features are applicable to diagnose non-PNS MPNSTs and to demonstrate useful markers for distinction between canine MPNSTs and PWTs, the authors examined 30 canine MPNSTs and 31 PWTs immunohistochemically for S100, nestin, NGFR, Olig2, claudin-1, CD57, PRX, α-SMA, desmin, and calponin. Among canine MPNSTs, the PNS tumors displayed significantly higher S100 and Olig2 expression than the non-PNS tumors. The expression levels of the other markers did not differ significantly, suggesting that the same morphological diagnostic criteria are applicable regardless of their location. The PWT cells displayed significantly weaker immunoreactivity than MPNSTs to markers used except α-SMA and desmin. Cluster analysis sorted most canine MPNSTs and PWTs into 2 distinctly different clusters, whereas 3 MPNSTs and 6 PWTs were assigned to the opposing cluster. These 3 MPNSTs were negative for almost all markers, while these 6 PWTs were positive for only neuronal markers. In particular, NGFR and Olig2 were almost negative in the rest of PWT cases. These findings suggest that NGFR and Olig2 are useful to distinguish these 2 tumors.

Establishment of a rat model for canine necrotizing meningoencephalitis (NME).

The pathogenesis of necrotizing meningoencephalitis (NME), necrotizing leukoencephalitis (NLE), and granulomatous meningoencephalomyelitis (GME) is still uncertain, although they are considered immune-mediated diseases. The purpose of the present study is to generate a rodent model(s) of these diseases. Rats were injected with rat cerebral or cerebellar homogenate. Rats injected with cerebral homogenate (Cbr) exhibited vacuolar or malacic changes mainly in the cerebral cortex. CD3-positive T cells and Iba-1-positive and CD163-negative microglia infiltrated and activated around the lesions. IgG deposited in the glial fibrillary acid protein (GFAP)-positive glia limitans from the early phase, and CD3-positive T cells attached to GFAP-positive astrocytes. Autoantibodies against GFAP were detected in the sera. These pathological features of Cbr rats were consistent with those of canine NME. In contrast, rats injected with cerebral homogenate (Cbe) exhibited demyelinating lesions with inflammatory reactions in the cerebellum, brainstem, and spinal cord. The presence of demyelination and autoantibodies against myelin proteins in Cbe rats was similar to murine experimental autoimmune encephalitis and differed from NME, NLE, and GME. All the present findings indicate that autoantibodies together with microglia and T cells may play a major role in the pathogenesis of idiopathic canine meningoencephalomyelitis.