Keratin 6A Is Expressed at the Invasive Front and Enhances the Progression of Colorectal Cancer.

Keratins (KRTs) are intermediate filament proteins in epithelial cells, and they are important for cytoskeletal organization. KRT6A, classified as a type II KRT, is normally expressed in stratified squamous epithelium and squamous cell carcinomas. Little is known about the expression and role of KRT6A in adenocarcinomas. We investigated the clinicopathologic and molecular biological significance of KRT6A in colorectal adenocarcinoma. Immunostaining of colorectal adenocarcinoma cases treated at our institution demonstrated that KRT6A showed significantly stronger expression at the invasive front than that at the tumor center (P < .0001). The high KRT6A-expression cases (n = 47) tended to have a high budding grade associated with significantly worse prognoses. A multivariate analysis revealed that the KRT6A expression status was an independent prognostic factor for overall survival (P = .0004), disease-specific survival (P = .0097), and progression-free survival (P = .0033). The correlation between KRT6A and patient prognoses was also validated in an external cohort from a published data set. To determine the function of KRT6A in vitro, KRT6A was overexpressed in 3 colon cancer cell lines: DLD-1, SW620, and HCT 116. KRT6A overexpression increased migration and invasion in DLD-1 but did not in SW620 and HCT116. In 3-dimensional sphere-forming culture, KRT6A expression enhanced the irregular protrusion around the spheroid in DLD-1. Our findings in this study indicated that KRT6A expression is a valuable prognostic marker of colorectal cancer and KRT6A may be involved the molecular mechanism in the progression of invasive areas of colorectal cancer.

Ki-67 distribution, α-methylacyl-CoA racemase (AMACR) expression and mucin phenotypes are associated with non-polypoid growth in ulcerative colitis-associated neoplasia.

AIMS: Ulcerative colitis-associated neoplasia (UCAN) is characterised by multifocal tumourigenesis. A wide range of metachronous lesions have been reported to occur after endoscopic treatment of UCAN, which suggests the development of sporadic tumours in lesions treated as UCAN. Therefore, we aimed to evaluate differences of immunohistochemistry (IHC) in features and clinicopathological characteristics of intramucosal lesions in patients with ulcerative colitis (UC). METHODS AND RESULTS: We examined 35 intramucosal lesions resected for carcinoma or dysplasia by total colectomy from patients with UC and 71 sporadic adenomas (SAs) endoscopically resected from patients without UC. UC lesions were divided into the conventional UCAN group, defined as p53 mutant pattern and normal expression of ß-catenin, and the non-conventional UCAN group, defined as the rest. Ki-67 distribution, α-methylacyl-CoA racemase (AMACR) expression and mucin phenotypes were compared using IHC, and clinicopathological characteristics were investigated. Conventional and non-conventional UCAN lesions were located in the left colon and rectum. Relative to the SA lesions, UCAN lesions occurred in much younger patients and exhibited more frequent basal distribution of Ki-67 in tumour crypts. Conventional UCAN lesions tended to be non-polyploid and exhibited a higher frequency of normal AMACR expression than SA lesions. UC lesions were heterogeneous-only two of the eight patients with multiple lesions had lesions (both non-conventional UCAN lesions) exhibiting concordant IHC staining features. CONCLUSIONS: The basal pattern of Ki-67 distribution, normal expression of AMACR and a non-intestinal mucin phenotype were determined as characteristic features suggestive of UCAN. Non-polypoid growth was another a key feature of UCAN.

Vitamin D Receptor Mediates Attenuating Effect of Lithocholic Acid on Dextran Sulfate Sodium Induced Colitis in Mice.

Bile acids are major components of bile; they emulsify dietary lipids for efficient digestion and absorption and act as signaling molecules that activate nuclear and membrane receptors. The vitamin D receptor (VDR) is a receptor for the active form of vitamin D and lithocholic acid (LCA), a secondary bile acid produced by the intestinal microflora. Unlike other bile acids that enter the enterohepatic circulation, LCA is poorly absorbed in the intestine. Although vitamin D signaling regulates various physiological functions, including calcium metabolism and inflammation/immunity, LCA signaling remains largely unknown. In this study, we investigated the effect of the oral administration of LCA on colitis in a mouse model using dextran sulfate sodium (DSS). Oral LCA decreased the disease activity of colitis in the early phase, which is a phenotype associated with the suppression of histological injury, such as inflammatory cell infiltration and goblet cell loss. These protective effects of LCA were abolished in VDR-deleted mice. LCA decreased the expression of inflammatory cytokine genes, but this effect was at least partly observed in VDR-deleted mice. The pharmacological effect of LCA on colitis was not associated with hypercalcemia, an adverse effect induced by vitamin D compounds. Therefore, LCA suppresses DSS-induced intestinal injury in its action as a VDR ligand.

Enhanced oxidative phosphorylation of IgG plasma cells can contribute to hypoxia in the mucosa of active ulcerative colitis.

Mucosal hypoxia is detected in the mucosa of ulcerative colitis (UC), however the mechanism and the cause of hypoxia is not fully understood, while a dense infiltration of plasma cells is observed in the inflamed mucosa of UC. When differentiating from a B cell to a plasma cell, the energy metabolism dramatically shifts from glycolysis to oxidative phosphorylation, which results in a large amount of oxygen consumption of the plasma cell. We hypothesized that the plasma cell infiltration into the inflamed mucosa contributes to the mucosal hypoxia in UC in part. We examined the association between mucosal hypoxia and plasma cell infiltration in UC. More IgG plasma cells (but not IgA plasma cells) were distributed, and the nuclear and cell sizes were enlarged in hypoxic mucosa compared to normoxic mucosa in UC. Oxidative phosphorylation signature genes of these IgG plasma cells were markedly upregulated compared to those of other lymphoid cells infiltrating the lamina propria of inflamed mucosa of UC. Enlarged IgG plasma cells, which increase in number in the inflamed mucosa of UC, can be related to the hypoxic state of the inflamed mucosa of UC.

[Multiple Minute Pulmonary Meningothelial-like Nodules;Report of a Case].

A 57-year-old woman was referred to our hospital for investigation of multiple tiny nodules in the lung fields bilaterally on computed tomography (CT). Video-assisted thoracoscopic lung biopsy was performed to diagnose the pulmonary lesions. Histological analysis showed nodular lesions with interstitial proliferation of uniform, round to oval cells with variable widening of the alveolar septa. Immunohistochemically, the cells were positive for EMA, CD56 and the progesterone receptor, but negative for chromogranin and synaptophysin. The diagnosis was "diffuse pulmonary meningotheliomatosis", with multiple diffuse "minute pulmonary meningothelial-like nodules". Diffuse pulmonary meningotheliomatosis should be kept in mind when we encounter small nodular shadows on a CT scan.

[Pancreatic Pleural Effusion;Report of a Case].

A 50-year-old man was admitted with respiratory failure. Chest X-ray and computed tomography revealed massive left pleural effusion and mediastinal shift. Pleural effusion showed abnormally high amylase levels of 42,600 IU/l and a high protein level of 3.2 g/dl. The serum amylase level was also 42,100 IU/l, and the proportion of pancreatic-type amylase was 88%. We diagnosed the patient with pancreatic effusion. Chest and abdominal enhanced computed tomography and magnetic resonance cholangiopancreatography revealed no pancreaticopleural fistula. He underwent a thoracoscopic examination that revealed brown pleural effusion as well as fibrin clots and thickness of the pleura. Histologically, there was no malignancy and the cause of pleural effusion was considered to be chronic pancreatitis.

Adenocarcinoma within anorectal fistulae: different clinicopathological characteristics between Crohn's disease-associated type and the usual type.

Adenocarcinoma within anorectal fistulae is rare and is sometimes associated with Crohn's disease. Crohn's disease-associated adenocarcinoma within anorectal fistulae has a poor prognosis; however, little is known about the clinicopathological differences between Crohn's disease-associated adenocarcinoma within anorectal fistulae and usual adenocarcinoma within anorectal fistulae. We retrospectively searched patients' charts and pathology archives at Tokyo Yamate Medical Center and Tokyo Medical and Dental University Hospital for adenocarcinoma within anorectal fistulae. Clinical and pathological data were collected and immunohistochemical examinations were conducted. Overall survival rate was estimated using the Kaplan-Meier method. Prognostic factors of overall survival were assessed using univariate and multivariate Cox regression analyses. We examined 82 cases of adenocarcinoma within anorectal fistulae. Fifty-nine of 82 cases (72%) had usual adenocarcinoma within anorectal fistulae, while the remaining 23 cases (28%) had Crohn's disease-associated adenocarcinoma within anorectal fistulae. Patients with Crohn's disease-associated adenocarcinoma within anorectal fistulae were diagnosed at a younger age and at a more advanced stage than those with usual adenocarcinoma within anorectal fistulae. Macroscopic and histological types were also different between usual adenocarcinoma within anorectal fistulae and Crohn's disease-associated adenocarcinoma within anorectal fistulae. Crohn's disease-associated adenocarcinoma within anorectal fistulae included more ulcerative types and high-grade adenocarcinomas. The rate of lymphovascular invasion was higher in Crohn's disease-associated adenocarcinoma within anorectal fistulae. Immunohistochemically, the expression of E-cadherin, p53, and MUC5AC differed between usual adenocarcinoma within anorectal fistulae and Crohn's disease-associated adenocarcinoma within anorectal fistulae. Patients with Crohn's disease-associated adenocarcinoma within anorectal fistulae exhibited worse overall survival than those with usual adenocarcinoma within anorectal fistulae, and vascular invasion was the strongest significant independent predictor of overall survival in patients with adenocarcinoma within anorectal fistulae. In conclusion, usual adenocarcinoma within anorectal fistulae and Crohn's disease-associated adenocarcinoma within anorectal fistulae have different clinicopathological characteristics and should be considered separate clinical entities.

Macroscopic Localized Subicular Thinning as a Potential Indicator of Amyotrophic Lateral Sclerosis.

Subicular degeneration occurs in amyotrophic lateral sclerosis (ALS) patients. However, it was unknown whether microscopic subicular degeneration could be observed as macroscopic changes and whether these changes were associated with the transactive-response DNA binding protein 43 kDa (TDP-43) pathology. Topographic differences between subicular degeneration caused by ALS and Alzheimer disease (AD) had also not been characterized. Here we investigated the subiculum and related areas in autopsied brains from 3 ALS and 3 AD patients. Macroscopic subicular thinning and corresponding astrocytosis were pronounced in ALS compared to AD. This thinning was frequently accompanied by TDP-43 pathology in the transentorhinal cortex and nucleus accumbens. The preferential susceptibility of the perforant pathway to TDP-43 deposition may be an underlying cause of subicular thinning in ALS.

Failing Left Ventricles Have an Enhanced Post-Stimulation Potentiation Despite Their Impaired Force Frequency Relationship.

The left ventricular contractile force (LV dP/dtmax) of patients with left ventricular systolic dysfunction does not increase effectively with an increase in heart rate. In other words, their force-frequency relationship (FFR) is impaired. However, it is unknown whether a longer coupling interval subsequent to tachycardia causes a stronger contraction (poststimulation potentiation, PSP) in a rate-dependent manner.In 16 patients with idiopathic dilated cardiomyopathy (DCM) (48 ± 2 years old, LVEF 30 ± 10%) and 6 control patients (58 ± 4 years old, LVEF 70 ± 7%), FFR was assessed by right atrial pacing using a micro-manometer-tipped catheter. At each pacing rate, the increase of LV dP/dtmax over basal LV dP/dt (ΔFFR) and the increase of LV dP/dtmax of the first beat after pacing cessation over LV dP/dtmax during pacing (ΔPSP) were evaluated.Patients with DCM had smaller LV dP/dtmax at baseline (872 ± 251 versus 1370 ± 123 mmHg/second, P = 0.0002) and developed smaller ΔFFR (eg, at 120/minute, 77 ± 143 versus 331 ± 131 mmHg/second, P = 0.0011). In contrast, they showed a rate-dependent increase of LV dP/dtmax of PSP and had greater ΔPSP (eg, at 120/minute, 294 ± 173 versus -152 ± 131 mmHg/second, P < 0.0001).Failing left ventricles develop little contractile force during tachycardia despite their rate-dependent enhancement in post-stimulation potentiation, suggesting that refractoriness of contractile force underlies impaired FFR.

Relative refractoriness of left ventricular contraction underlies human tachycardia-induced mechanical and electrical alternans.

BACKGROUND: Mechanical alternans (MA) and electrical alternans (EA) are predictors of cardiac events. Experimental studies have suggested that refractoriness of calcium cycling underlies these cardiac alternans. However, refractoriness of left ventricular contraction has not been examined in patients with cardiac alternans. METHODS: In 51 patients with miscellaneous heart diseases, incremental right atrial pacing was performed to induce MA and EA. MA was quantified by alternans amplitude (AA: the difference between left ventricular dP/dt of a strong beat and that of a weak beat), and AA at 100/min (AA100) and maximal AA (AAmax) were measured. EA was defined as alternation of T wave morphology in 12-lead electrocardiogram. Relative refractoriness of left ventricular contraction was examined by drawing the mechanical restitution curve under a basal coupling interval (BCL) of 600 ms (100/min) and was assessed by the slope at BCL (Δmechanical restitution). Postextrasystolic potentiation (PESP) was also examined and the slope of PESP curve (ΔPESP) was assessed as a property to alternate strong and weak beats. RESULTS: MA and EA were induced in 19 patients and in none at 100/min or less, and at any heart rate in 32 and in 10, respectively. AA100 and AAmax correlated positively with Δmechanical restitution and negatively with ΔPESP. Patients with EA had a significantly larger Δmechanical restitution and a significantly larger absolute value of ΔPESP than those without. CONCLUSIONS: In patients with MA and EA, the left ventricular contractile force during tachycardia is under relative refractoriness and prone to cause large fluctuation of contractile force.

Longitudinal and regional association between dietary factors and prevalence of Crohn's disease in Japan.

Although a Western diet has been identified as a risk factor for Crohn's disease (CD), there is still controversy surrounding the specific foods that may contribute to the development of the disease. In this study, we examined the association between food intake and the prevalence of CD in Japan, as Japanese patients with CD are known to have limited genetic involvement. We identified changes in food intake associated with an increase in the number of patients with CD by analyzing the per capita consumption of food types from 1981 to 2014. Additionally, we examined the association between CD prevalence and food intake in each prefecture. Finally, the relationship between food intake and estimated age at CD onset was also investigated. Between 1981 and 2014, we observed Increased consumption of meat, eggs, milk and dairy products, oil, and potatoes and decreased consumption of grains, beans, vegetables, fruit, fish, sugar, and seaweed. The annual incidence of CD increased by 1388% over the same period. We found that meat consumption was significantly associated with CD prevalence (ß = 0.503, p = 0.0003), while a significant negative correlation was observed between CD prevalence and fruit and vegetable consumption (fruit, ß = 0.464, p = 0.0012; vegetables, ß = 0.404, p = 0.0023). Furthermore, we estimated that the peak consumption of more meat and less fruit and vegetables and the peak age of CD onset occurred within the age range of 20-24 years. Our study identified a clear correlation between the consumption of meat, fruits, and vegetables and the prevalence of CD in Japan. Additionally, we found an association between meat, fruit, and vegetable consumption and the age at CD onset.

Variability in morphology and immunohistochemistry of Crohn's disease-associated small bowel neoplasms: implications of Claudin 18 and Cadherin 17 expression for tumor-targeted immunotherapies.

AIMS: Inflammatory bowel disease-associated colorectal carcinomas are known to have different morphology, immunoprofile, and genetic findings from sporadic colorectal carcinomas; however, little is known for Crohn's disease-associated small bowel neoplasms (CD-SBNs). Cadherin 17 is a useful biomarker of adenocarcinomas with intestinal phenotype and recently reported as an ideal target for chimeric antigen receptor T-cells (CAR-T) therapy for gastrointestinal carcinoma. Claudin 18 is a cell adhesion protein, and Claudin18 isoform 2 (CLDN18.2) is frequently expressed at high levels in gastric-type adenocarcinoma. Zolbetuximab, a targeted monoclonal antibody, has been developed for CLDN18.2-positive gastroesophageal adenocarcinoma. We examined a series of CD-SBNs for both Cadherin 17 and Claudin 18, and also hypothesized that expression of Claudin 18 was associated with gastric phenotype. METHODS AND RESULTS: We performed histological and immunohistochemical examinations on 25 CD-SBNs. Most of adenocarcinomas showed tubular morphology as seen in gastric carcinomas, whereas a subset of dysplasia was morphologically similar to that of the large bowel. Cadherin17 and Claudin 18 expression was identified in 93% and 57% CD-associated adenocarcinomas respectively. In Cadherin 17-positive CD-SBNs, frequent MUC5AC, MUC6, and Claudin18 expression was identified (61%, 57%, and 57%, respectively). Claudin 18-positive CD-SBNs showed significantly more MUC5AC and MUC6 expression than Claudin 18-negative CD-SBNs (P = 0.005, < 0.001 respectively). CONCLUSION: In CD-associated small bowel adenocarcinomas, Cadherin 17 expression was frequently retained and Claudin 18 was frequently co-expressed. Claudin 18 had a positive correlation with the expression of gastric mucins. These results suggest that CD-associated small bowel adenocarcinomas may be candidates for Cadherin 17- and Claudin 18-targeted immunotherapies.

Efficacy of repeated intravenous injection of peramivir against influenza A (H1N1) 2009 virus infection in immunosuppressed mice.

The efficacy of intravenous peramivir against influenza A (H1N1) 2009 virus infection was evaluated in mice in which the immune system was suppressed by cyclophosphamide (CP) treatment. The mortality rate of the vehicle control group was 100%, and the mice lost 20% of their body weight on average by day 13 postinfection (p.i.). Repeated administration of peramivir (40 mg/kg of body weight once a day, given intravenously for 20 days), starting at 1 h p.i., significantly reduced mortality, body weight loss, viral titers, and cytokine production in infected mice compared with results for administration of vehicle (P < 0.01). In addition, repeated administration of peramivir, starting at 24 h, 48 h, or 72 h p.i., also resulted in increases in survival rates and reduction of viral titers in the lungs (P < 0.01). The mean days to death (MDD) of the vehicle group was 14.5 days, while in the groups treated with peramivir starting at 24 h, 48 h, and 72 h p.i., the MDDs were >23.0, 20.9, and 21.8 days, respectively. In comparison, repeated administration of oseltamivir phosphate (5 mg/kg twice a day, given orally for 20 days), starting at 24 h, 48 h, and 72 h p.i., also significantly prevented body weight loss, whereas no significant differences in mortality rates and viral titers in the lungs were observed compared with results for the vehicle group. These data indicated that repeated administration of peramivir was effective in promoting the survival and reducing virus replication in immunosuppressed mice infected with influenza A (H1N1) 2009 virus.

Estimation of paracrine signaling using quantitative RT-PCR from multiple patchy lesion samples.

We investigated whether correlations between mRNA levels of cytokines versus other proteins from patchy lesion could estimate cytokine paracrine signaling in vivo. Experiments with rat experimental autoimmune myocarditis (EAM), a patchy myocarditis model, indicated IL-1 and other protein levels were correlated, indicating paracrine signaling pathways in vivo.

Mechanical alternans in human idiopathic dilated cardiomyopathy is caused with impaired force-frequency relationship and enhanced poststimulation potentiation.

Mechanical alternans (MA) is frequently observed in patients with heart failure, and is a predictor of cardiac events. However, there have been controversies regarding the conditions and mechanisms of MA. To clarify heart rate-dependent contractile properties related to MA, we performed incremental right atrial pacing in 17 idiopathic dilated cardiomyopathy (DCM) patients and in six control patients. The maximal increase in left ventricular dP/dt during pacing-induced tachycardia was assessed as the force gain in the force-frequency relationship (FG-FFR), and the maximal increase in left ventricular dP/dt of the first post-pacing beats was examined as the force gain in poststimulation potentiation (FG-PSP). As a result, MA was induced in 9 DCM patients (DCM MA(+)) but not in the other 8 DCM patients (DCM MA(-)), and not in any of the control patients. DCM MA(+) had significantly lower FG-FFR (34.7 ± 40.9 vs 159.4 ± 103.9 mmHg/s, P = 0.0091) and higher FG-PSP (500.0 ± 96.8 vs 321.9 ± 94.9 mmHg/s, P = 0.0017), and accordingly a wider gap between FG-PSP and FG-FFR (465.3 ± 119.4 vs 162.5 ± 123.6 mmHg/s, P = 0.0001) than DCM MA(-) patients. These characteristics of DCM MA(+) showed clear contrasts to those of the control patients. In conclusion, MA is caused with an impaired force-frequency relationship despite significant poststimulation potentiation, suggesting that MA reflects ineffective utilization of the potentiated intrinsic force during tachycardia.

Unencapsulated thymoma: a case report.

Thymomas, the most common mediastinal tumors, form capsules. Only a few reports have presented small thymomas without capsule formation, so-called microthymomas. Here, we report a case of an unencapsulated thymoma measuring 18 mm. A 42-year-old female presented with an anterior mediastinal tumor. Computed tomography revealed an 18-mm nodule in the anterior mediastinum. Magnetic resonance imaging revealed a solid tumor that was iso-intense on T1-weighted images and hypo-intense on T2-weighted images. Thoracoscopic partial thymectomy was performed. The histopathological diagnosis was a type B1 thymoma. The tumor was localized within the thymic tissue lacked a fibrous capsule and partially invaded the surrounding fat tissue. To our knowledge, this is the first report of an unencapsulated thymoma, except for microthymomas.

Value of apparent diffusion coefficient on MRI for prediction of histopathological type in anal fistula cancer.

The main histopathological types of anal fistula cancers are mucinous adenocarcinoma and tubular adenocarcinoma. The purpose of this study was to investigate the utility of the apparent diffusion coefficient (ADC) value in magnetic resonance imaging (MRI) to determine the histopathological type of an anal fistula cancer, and to investigate the relationship between ADC values and histopathological type (mucinous type or tubular carcinoma), clinical information, and surgical findings. We retrospectively identified 69 patients diagnosed with anal fistula cancer at our hospital from January 2013 to December 2021. Among them, we selected the patients diagnosed using the same 1.5-T MRI machine, underwent surgery, and a pathological sample was obtained during the operation. Finally, these 25 patients were selected for the analysis since they underwent the imaging scan using the same MRI machine. The ADC value was compared between mucinous and tubular adenocarcinomas, and between tumors at the Tis-T1-T2 and T3-T4 stages. Finally, 25 patients were selected. The mean age of the 25 patients included in the analysis was 60.8 ± 13.3 years and all were males. The median ADC of anal fistula cancers was 1.97 × 10-3 mm2/s for mucinous adenocarcinomas and 1.36 × 10-3 mm2/s for tubular adenocarcinomas; this difference was statistically significant (P < .01). Furthermore, the median ADC was 1.62 × 10-3 mm2/s for tumors in Tis-T1-T2 stages and 2.01 × 10-3 mm2/s for T3-T4 tumors (P = .02). The ADC value in MR images may predict the histopathological type and depth of anal fistula cancers. Also, the different ADC values between Tis-T1-T2 and T3-T4 tumors could help predict the classification of progression.

Systemic transplantation of allogenic fetal membrane-derived mesenchymal stem cells suppresses Th1 and Th17 T cell responses in experimental autoimmune myocarditis.

We have reported that systemic administration of autologous bone marrow or allogenic fetal membrane (FM)-derived mesenchymal stem cells (MSCs) similarly attenuated myocardial injury in rats with experimental autoimmune myocarditis (EAM). Since rat EAM is a T-helper (Th) cell-mediated autoimmune disease, and recent evidence has indicated that both autologous and allogenic MSCs exert an immunosuppressive effect on Th cell activity, we focused on Th cell differentiation in allogenic FM-MSC administered EAM rats. EAM was induced in Lewis rats by injecting porcine cardiac myosin (day 0). Allogenic FM-MSCs, obtained from major histocompatibility complex mismatched ACI rats, were intravenously injected (5 × 10(5)cells/rat) on days 7, 10, or 14 (MSCd7, MSCd10, or MSCd14 groups, respectively). At day 21, echocardiography confirmed that reduced ejection fraction in the untreated EAM group (63 ± 2%) was significantly improved in the MSCd10 and MSCd14 groups (74 ± 1 and 75 ± 2%, respectively, P<0.01). CD68 immunostaining revealed that prominent macrophage infiltration in the myocardium of the EAM group (1466 ± 93 cells/mm(2)) was significantly decreased in the MSCd10 group (958 ± 139 cells/mm(2), P<0.05). To evaluate Th cell differentiation, we used flow cytometry to determine the percentage of interferon (IFN)-γ positive Th1 and interleukin (IL)-17 positive Th17 cells in peripheral CD4-positive Th cells. The percentage of Th1 cells at day 16 was significantly lower in the MSCd10 (1.3 ± 0.2%) and MSCd14 (1.6 ± 0.3%) groups compared to the EAM group (2.4 ± 0.3%, P<0.05), as was the percentage of Th17 cells in the MSCd10 group (1.9 ± 0.5%) compared to the EAM group (2.2 ± 0.9%, P<0.05). At day 21, infiltrating Th17 cells in myocardium were significantly decreased in the MSCd10 group (501 ± 132 cells/mm(2), P<0.05) compared to EAM (921 ± 109 cells/mm(2)). In addition, human CD4+ Th cells co-cultured with human FM-MSCs exhibited reduced Th1 and Th17 cell-differentiation and proliferation, with increased expression of immunosuppressive molecules including indoleamine 2,3-dioxygenase 2 and IL-6 in co-cultured FM-MSCs. These results suggest that intravenous administration of allogenic FM-MSCs ameliorates EAM via the suppression of Th1/Th17 immunity.

Beneficial effects of edaravone, a novel antioxidant, in rats with dilated cardiomyopathy.

Edaravone, a novel antioxidant, acts by trapping hydroxyl radicals, quenching active oxygen and so on. Its cardioprotective activity against experimental autoimmune myocarditis (EAM) was reported. Nevertheless, it remains to be determined whether edaravone protects against cardiac remodelling in dilated cardiomyopathy (DCM). The present study was undertaken to assess whether edaravone attenuates myocardial fibrosis, and examine the effect of edaravone on cardiac function in rats with DCM after EAM. Rat model of EAM was prepared by injection with porcine cardiac myosin 28 days after immunization, we administered edaravone intraperitoneally at 3 and 10 mg/kg/day to rats for 28 days. The results were compared with vehicle-treated rats with DCM. Cardiac function, by haemodynamic and echocardiographic study and histopathology were performed. Left ventricular (LV) expression of NADPH oxidase subunits (p47(phox), p67(phox), gp91(phox) and Nox4), fibrosis markers (TGF-ß(1) and OPN), endoplasmic reticulum (ER) stress markers (GRP78 and GADD 153) and apoptosis markers (cytochrome C and caspase-3) were measured by Western blotting. Edaravone-treated DCM rats showed better cardiac function compared with those of the vehicle-treated rats. In addition, LV expressions of NADPH oxidase subunits levels were significantly down-regulated in edaravone-treated rats. Furthermore, the number of collagen-III positive cells in the myocardium of edaravone-treated rats was lower compared with those of the vehicle-treated rats. Our results suggest that edaravone ameliorated the progression of DCM by modulating oxidative and ER stress-mediated myocardial apoptosis and fibrosis.

Involvement of AMPK and MAPK signaling during the progression of experimental autoimmune myocarditis in rats and its blockade using a novel antioxidant.

There are various reports suggesting the role of angiotensin (Ang) receptor blockers, Ang converting enzyme inhibitors, calcium channel blockers, diuretics and antioxidants against the progression of experimental autoimmune myocarditis (EAM) to dilated cardiomyopathy (DCM). Most of them were reported to be effective during this adverse cardiac remodeling. Recently much attention has been paid to studying the involvement of AMP-activated protein kinase (AMPK) and mitogen activated protein kinase (MAPK) in various cardiovascular ailments. AMPK acts as a master sensor of cellular energy balance via maintenance of lipid and glucose metabolism. Evidences also suggest the relation between AMPK and oxidative stress during physiological and pathological myocardial cellular function. Since, it is of interest to identify the roles of AMPK and MAPK during the progression of EAM to DCM and also the effect of edaravone, a novel free radical scavenger, against its progression. For this, we have carried out western blotting, histopathological staining and immunohistochemical analyses to measure the myocardial expressions of AMPK signaling and oxidative stress related parameters in normal and vehicle or edaravone-treated EAM rats, respectively. We identified the myocardial levels of phospho Akt and phosphoinositide 3-kinase, which are the upstream proteins of AMPK and MAPK activation and both were up-regulated in the vehicle-treated rats, whereas candesartan treatment significantly reversed these changes. We have also measured the myocardial levels of p-AMPKα, different isoforms of protein kinase C and MAPK signaling proteins. All of these protein levels were significantly elevated in the hearts of DCM rats whereas edaravone treatment significantly reversed these changes. In viewing these results, we can suggest that along with MAPK, AMPK signaling also plays a crucial role in the progression of EAM and it can be effectively blocked by the treatment with a novel antioxidant, edaravone.