Phosphorylation of Smad2/3 at the specific linker threonine residue indicates slow-cycling esophageal stem-like cells before re-entry to the cell cycle.

UNLABELLED: The stem cell compartment in the esophageal epithelium is possibly located in the basal layer. We have identified significant expression of Smad2/3, phosphorylated at specific linker threonine residues (pSmad2/3L-Thr), in the epithelial cells of murine stomach and intestine, and have suggested that these cells are epithelial stem cells. In this study, we explore whether pSmad2/3L-Thr could serve as a biomarker for esophageal stem cells. We examined esophageal tissues from normal C57BL/6 mice and those with esophagitis. Double immunofluorescent staining of pSmad2/3L-Thr with Ki67, CDK4, p63, or CK14 was performed. After immunofluorescent staining, we stained the same sections with hematoxylin-eosin and observed these cells under a light microscope. We used the 5-bromo-2-deoxyuridine (BrdU) labeling assay to examine label retention of pSmad2/3L-Thr immunostaining-positive cells. We collected specimens 5, 10, 15 and 20 days after repeated BrdU administrations and observed double immunofluorescent staining of pSmad2/3L-Thr with BrdU. In the esophagus, pSmad2/3L-Thr immunostaining-positive cells were detected in the basal layer. These cells were detected between Ki67 immunostaining-positive cells, but they were not co-localized with Ki67. pSmad2/3L-Thr immunostaining-positive cells showed co-localization with CDK4, p63, and CK14. Under a light microscope, pSmad2/3L-Thr immunostaining-positive cells indicated undifferentiated morphological features. Until 20 days follow-up period, pSmad2/3L-Thr immunostaining-positive cells were co-localized with BrdU. pSmad2/3L-Thr immunostaining-positive cells significantly increased in the regeneration phase of esophagitis mucosae, as compared with control mice (esophagitis vs. CONTROL: 6.889 ± 0.676/cm vs. 4.293 ± 0.659/cm; P < 0.001). We have identified significant expression of pSmad2/3L-Thr in the specific epithelial cells of murine esophagi. We suggest that these cells are slow-cycling epithelial stem-like cells before re-entry to the cell cycle.

Progressive irreversible hearing loss is caused by stria vascularis degeneration in an Slc26a4-insufficient mouse model of large vestibular aqueduct syndrome.

Hearing loss of patients with enlargement of the vestibular aqueduct (EVA) can fluctuate or progress, with overall downward progression. The most common detectable cause of EVA is mutations of SLC26A4. We previously described a transgenic Slc26a4-insufficient mouse model of EVA in which Slc26a4 expression is controlled by doxycycline administration. Mice that received doxycycline from conception until embryonic day 17.5 (DE17.5; doxycycline discontinued at embryonic day 17.5) had fluctuating hearing loss between 1 and 6 months of age with an overall downward progression after 6 months of age. In this study, we characterized the cochlear functional and structural changes underlying irreversible hearing loss in DE17.5 mice at 12 months of age. The endocochlear potential was decreased and inversely correlated with auditory brainstem response thresholds. The stria vascularis was thickened and edematous in ears with less severe hearing loss, and thinned and atrophic in ears with more severe hearing loss. There were pathologic changes in marginal cell morphology and gene expression that were not observed at 3 months. We conclude that strial dysfunction and degeneration are the primary causes of irreversible progressive hearing loss in our Slc26a4-insufficient mouse model of EVA. This model of primary strial atrophy may be used to explore the mechanisms of progressive hearing loss due to strial dysfunction.

Splenectomy and antiviral treatment for thrombocytopenic patients with chronic hepatitis C virus infection.

Thrombocytopenic patients with chronic hepatitis C virus (HCV) infection are poor candidates for antiviral treatment with interferon (IFN), but no standard treatment for thrombocytopenia has yet been established. We evaluated the safety of splenectomy and its efficacy for the initiation and continuation of antiviral therapy. From March 2003 to April 2006, 10 patients (mean age 62.5 years) with HCV-related cirrhosis, low platelet count (<==106 000/mm(3)) and splenomegaly (spleen size >==10 cm) underwent splenectomy. Platelet counts significantly increased at 4-8 weeks after splenectomy [pre: 64 200 +/- 6900/mm(3)vs post 209 000 +/- 40 600/mm(3) (P = 0.004)]. No severe operative complications were observed. All patients subsequently received antiviral therapy. Of the eight patients who were infected with HCV genotype 1 and had a high viral load (>==100 KIU/mL), four received combination therapy with pegylated IFNalpha-2b plus ribavirin, and the other four received standard IFNalpha-2b plus ribavirin. One patient infected with HCV genotype 2 and another with HCV genotype 1 and a low viral load (<100 KIU/mL) were treated with pegylated IFNalpha-2a. Six patients achieved sustained virologic response (SVR). Among four patients who failed to achieve SVR, one was given retreatment with pegylated IFN plus ribavirin, and the other three received low-dose long-term IFN therapy. Although this study was small, the treatment results were similar to those for patients without thrombocytopenia and suggested that splenectomy would not reduce the antiviral efficacy of IFNalpha-based treatment.

Techniques to facilitate ERCP with a conventional endoscope in patients with previous pancreatoduodenectomy.

There is little guidance on the performance of endoscopic retrograde cholangiopancreatography (ERCP) in patients with previous pancreatoduodenectomy. We reviewed techniques for ERCP with a conventional endoscope and assessed its value in 10 patients with previous pancreatoduodenectomy (15 ERCPs). After exploration of the surgical reconstruction, we used a front-viewing endoscope, and we used a small firm pillow under the abdomen and hand compression for preventing loop formation. Successful insertion to the ductal anastomoses and biliary cannulation were achieved in 13 / 15 procedures (87 %). In 6 procedures where we attempted pancreatic cannulation, we could not identify the pancreatojejunostomy, but after spraying contrast around the suspected location of the ductal anastomosis we obtained a pancreatogram in 4 / 6 procedures (67 %). Endoscopic biliary interventions were successful in 6 / 7 procedures (86 %). No complications were encountered. Use of appropriate techniques makes ERCP with a conventional endoscope feasible, effective, and safe in patients with previous pancreatoduodenectomy. Endoscopic therapy can be performed successfully in the bile duct, but has limited value regarding the pancreatic duct.

Characterization of bradykinin-induced endothelium-independent contraction in equine basilar artery.

We investigated the effect of bradykinin (BK) on isolated equine basilar arterial rings with and without endothelium. BK induced concentration-dependent contraction of resting arterial rings and no relaxation when the rings were precontracted by prostaglandin F(2alpha). The maximal response and pD(2) value were 161.2 +/- 28.1% (to 60 mm KCl-induced contraction) and 8.24 +/- 0.25 respectively. The cumulative concentration-response curve for BK was not shifted to the right by des-Arg(9)-[Leu(8)]-BK (a B(1)-receptor antagonist), HOE140 (a B(2)-receptor antagonist) or NPC567 (another B(2)-receptor antagonist). In four of six basilar arteries, NPC567 induced concentration-dependent contraction. Indomethacin (a cyclooxygenase inhibitor), nordihydroguaiaretic acid (a lipoxygenase inhibitor), quinacrine (a phospholipase A(2) inhibitor), tetrodotoxin (a selective blocker of Na(+) channels), guanethidine (a nor-adrenergic neuron blocking drug), phentolamine (an alpha-adrenoceptor antagonist), Nomega-nitro-L-arginine (L-NNA, a nitric oxide (NO) synthase inhibitor) and endothelial denudation did not affect the BK-induced contraction. L-NNA and indomethacin induced contraction and relaxation under resting vascular tone respectively. These results suggest that endothelial cells are not involved in BK-induced contraction and that the contraction is not mediated via activation of known B(1) and B(2) receptors. Arachidonic acid metabolites and neurotransmitters like norepinephrine and NO might not play a role in BK-induced contraction in equine basilar artery.

Human TSLP and TLR3 ligands promote differentiation of Th17 cells with a central memory phenotype under Th2-polarizing conditions.

UNLABELLED: BACKGROUND" Human thymic stromal lymphopoietin (TSLP) is expressed in the human asthmatic lung and activates dendritic cells (DCs) to strongly induce proallergic T-helper type 2 (Th2) cell responses, suggesting that TSLP plays a critical role in the pathophysiology of human asthma. Th2 cells are predominantly involved in mild asthma, whereas a mixture of Th1 and Th2 cells with neutrophilic inflammation, probably induced by Th17, affects more severe asthmatic disease. Exacerbation of asthmatic inflammation is often triggered by airway-targeting RNA viral infection; virus-derived double-stranded RNA, Toll-like receptor (TLR)3 ligand, activates bronchial epithelial cells to produce pro-inflammatory mediators, including TSLP. OBJECTIVE: Because TSLPR-expressing DCs express TLR3, we examined how the relationship between TSLP and TLR3 ligand stimulation influences DC activation. METHODS: CD11c(+)DCs purified from adult peripheral blood were cultured in TLR ligands containing media with or without TSLP and then co-cultured with allogeneic naïve CD4(+)T cells. RESULTS: CD11c(+) DCs responded to a combination of TSLP and TLR3 ligand, poly(I : C), to up-regulate expression of the functional TSLP receptor and TLR3. Although TSLP alone did not induce IL-23 production by DCs, poly(I : C) alone primed DCs for the production of IL-23, and a combination of TSLP and poly(I : C) primed DCs for further production of IL-23. The addition of poly(I : C) did not inhibit TSLP-activated DCs to prime naïve CD4(+) T cells to differentiate into inflammatory Th2 cells. Furthermore, DCs activated by a combination of TSLP and poly(I : C) primed more naïve CD4(+) T cells to differentiate into Th17-cytokine-producing cells with a central memory T cell phenotype compared with DCs activated by poly(I : C) alone. CONCLUSIONS: These results suggest that through DC activation, human TSLP and TLR3 ligands promote differentiation of Th17 cells with the central memory T cell phenotype under Th2-polarizing conditions.

Anomalous origin of the anterior choroidal artery.

SUMMARY: We describe a very rare case of anomalous origin of the anterior choroidal artery. In our case the anterior choroidal artery arises from the internal carotid artery proximal to the posterior communicating artery.

Human TSLP directly enhances expansion of CD8+ T cells.

Human thymic stromal lymphopoietin (TSLP) promotes CD4(+) T-cell proliferation both directly and indirectly through dendritic cell (DC) activation. Although human TSLP-activated DCs induce CD8(+) T-cell proliferation, it is not clear whether TSLP acts directly on CD8(+) T cells. In this study, we show that human CD8(+) T cells activated by T-cell receptor stimulation expressed TSLP receptor (TSLPR), and that TSLP directly enhanced proliferation of activated CD8(+) T cells. Although non-stimulated human CD8(+) T cells from peripheral blood did not express TSLPR, CD8(+) T cells activated by anti-CD3 plus anti-CD28 did express TSLPR. After T-cell receptor stimulation, TSLP directly enhanced the expansion of activated CD8(+) T cells. Interestingly, using monocyte-derived DCs pulsed with a cytomegalovirus (CMV)-specific pp65 peptide, we found that although interleukin-2 allowed expansion of both CMV-specific and non-specific CD8(+) T cells, TSLP induced expansion of only CMV-specific CD8(+) T cells. These results suggest that human TSLP directly enhances expansion of CD8(+) T cells and that the direct and indirect action of TSLP on expansion of target antigen-specific CD8(+) T cells may be beneficial to adoptive cell transfer immunotherapy.

Morphologic evaluation of the caudal end of the inferior petrosal sinus using 3D rotational venography.

BACKGROUND AND PURPOSE: The inferior petrosal sinus (IPS) is the main transvenous access route used to examine or treat lesions involving the cavernous sinus. To carry out these procedures successfully, one must have a detailed knowledge of the anatomy of the venous system around the junction of the IPS and the internal jugular vein (IJV). MATERIALS AND METHODS: Eighty-three sides in 63 patients (26 men, 37 women; mean, 56.5 years of age) were examined by using 3D rotational venography (3DRV). RESULT: The drainage patterns of the IPS could be classified into the following 6 types, with emphasis on the level of IPS-IJV junction: type A, the IPS drains into the jugular bulb in 1/83 sides (1.2%); type B, the IPS drains into the IJV at the level of the extracranial opening of the hypoglossal canal in 29/83 sides (34.9%); type C, the IPS drains into the lower extracranial IJV in 31/83 sides (37.3%); type D, the IPS forms a plexus and has multiple junctions to the IJV near the jugular foramen in 5/83 sides (6.0%); type E, the IPS drains directly into the vertebral venous plexus (VVP) with no connection to the IJV in 3/83 sides (3.6%); and type F, the IPS is absent in 14/83 sides (16.9%). Each type is also characterized by the way of anastomosis with the VVP. CONCLUSION: This classification seemed to be rational from the embryologic viewpoint, and it may be useful in establishing treatment strategies that involve endovascular manipulation via the IPS.

Oral treatment with probiotic Lactobacillus johnsonii NCC533 (La1) for a specific part of the weaning period prevents the development of atopic dermatitis induced after maturation in model mice, NC/Nga.

BACKGROUND: The inhibitory effect of probiotic bacteria on atopic dermatitis has been shown in human infants, but the mechanism is still unclear. OBJECTIVE: This study aimed to show the effects of the administration of a probiotic during the weaning period in mouse models on production of the intestinal secretory IgA (sIgA) and on the development of atopic dermatitis (AD) in later life. METHODS: The effects of the administration of Lactobacillus johnsonii NCC533 (La1) during weaning were evaluated using a mouse model (Balb/c). The weaning period of mice was divided into three phases according to the evolution of faecal IgA. La1 was administered in each phase and the evolution of the faecal IgA was estimated. In the next experiment, the effect of the administration of La1 in phase 2 on host immunity after maturation was further assessed by using the model NC/Nga mouse for human AD. RESULTS: Administration of La1 in each phase showed a distinct effect on the production of sIgA. But sIgA production was only positively stimulated when La1 was administrated in phase 2. The development of AD induced by mite antigen from 6 weeks old was significantly prevented by the primary administration of La1 in phase 2. AD-like lesions were significantly milder than those of the control mice, and histological observations showed an almost normal appearance of the epidermis and upper dermis of the mice treated with La1. CONCLUSION: This study suggested that the primary administration of La1 in a specific part of the weaning period is effective in preventing or inhibiting the development of AD after maturation by modulating or accelerating the gut immune response.

Atypical spinal dural arteriovenous fistula with supply from the lateral sacral artery.

We report a dural arteriovenous fistula (AVF) that developed at a site on the midline dorsal surface of the dura mater that had been damaged by repeated lumbar punctures. A 61-year-old male patient had undergone repeated lumbar punctures and discectomy for severe lumbago 40 years before the present admission. After surgery, the lumbago symptoms resolved. However, 30 years after the operation, he started to experience dysaesthesia, motor weakness in both legs, and urinary disturbance. Physical examination revealed bilateral leg weakness, diminished deep tendon reflexes in the patellar and Achilles tendons bilaterally, and decreased superficial sensation below L1. Magnetic resonance imaging revealed swelling with intramedullary high intensity and multiple flow voids around the conus and spinal cord on T(2)-weighted images, and adhesive arachnoiditis. Spinal angiography revealed an AVF between the left lateral sacral artery and the S1 radicular vein at the site of the previous operation. Surgery was conducted to carry out excision of the dural AVF at the shunting point, the arterialized intradural vein, and lysis of the arachnoiditis. This case of dural AVF may have been caused by repeated lumbar punctures.

Gastric mucosal hyperplasia via upregulation of gastrin induced by persistent activation of gastric innate immunity in major histocompatibility complex class II deficient mice.

BACKGROUND AND AIMS: Major histocompatibility complex class II deficient (Aalpha0/0) mice have decreased CD4+ T cells, making them immunologically similar to patients with acquired immunodeficiency syndrome (AIDS). Both patients with AIDS and Aalpha0/0 mice have hypertrophic gastric folds. To clarify the mechanism of gastric mucosal hyperplasia, we investigated the pathophysiology and the role of the innate immunity in the stomach of Aalpha0/0 mice. METHODS: Stomachs from 1-6 month old Aalpha0/0 mice, kept under specific pathogen free conditions, were examined at 1 month intervals histologically and immunohistochemically. Gene expression of proinflammatory cytokines, Toll-like receptors (TLRs), cyclooxygenase (COX)-2, and myeloperoxidase (MPO) activity in the gastric mucosa was investigated. Serum gastrin levels and gastric acidity were measured. Bacterial culture of the stomach was performed. To clarify the roles of hypergastrinaemia in the gastric mucosa, a gastrin receptor antagonist (AG041R) was administered. RESULTS: Aalpha0/0 mice had a diffusely thick corpus mucosa with infiltration of CD11b+ granulocytes and macrophages. Anti-Ki67 staining demonstrated expansion of the proliferating neck zone. Gene expression of interleukin 1beta, interferon gamma, TLR-2, TLR-4, and COX-2 were upregulated, and MPO activity was increased. Only a small amount of non-pathogenic bacteria was detected in the stomach. Serum gastrin levels and Reg-Ialpha positive cells in the gastric mucosa increased, despite normal gastric acidity. After treatment with AG041R, gastric mucosal thickness was significantly reduced. CONCLUSION: Persistent activation of innate immunity in the stomach induced gastric mucosal hyperplasia through upregulation of gastrin synthesis in Aalpha0/0 mice, suggesting a pathophysiology similar to the gastric changes in patients with AIDS.

Pitfalls during the Embolization and Evaluation after the Embolization for the Skull Base Meningiomas.

SUMMARY: Pitfall during the embolization and evaluation after the embolization for skull base meningiomas supplied by meningeal arteries of internal carotid artery (ICA) are reported. This study includes 15 cases of skull base meningiomas (two males and 13 females) that supplied by meningeal branches of internal carotid artery. The preoperative embolization was performed by these feeders. MRI findings and serum levels of C-reactive protein (CRP) after the embolization were examined. In ten patients among 15 patients the meningeal branches of ICA were dominant feeders. In ten patients out of 15 patients, the embolization from the meningeal branches of ICA was possible. Eight patients out of these ten patients were suffered from high fever and increase of serum level of CRP after the embolization. During the embolization for skull base meningiomas, the existence of collateral pathways between the ICA system and external carotid artery system were identified. The increase of serum levels of CRP might be recognized in the patients that effective embolization were performed.

In vitro application of endotoxin enhances nitric oxide production in thoracic aortas from Mg-deficient rats.

Since endotoxin-induced vascular hyporeactivity to phenylephrine is enhanced in Mg-deficient rats, this study was designed to determine whether endotoxin directly enhances nitric oxide (NO) production in thoracic aortas isolated from Mg-deficient rats in vitro. Thoracic aortas isolated from Mg-deficient and control rats were cultured for 6 h with or without endotoxin (LPS). LPS (0.01-1.0 microg) increased NO production in a concentration-dependent manner. NO production in the presence of 0.1 and 1.0 microg/mL LPS was significantly higher in Mg-deficient rat aortas compared to aortas from control rats. The enhanced NO production was not significantly affected by endothelium-denudation. LPS-stimulated NO production was fully inhibited by a selective iNOS inhibitor, 1400W (0.1, 1.0 microM), in control rat aortas, but in Mg-deficient rat aortas inhibition by 1400W was only partial. A similar inhibitory effect was observed with anti-CD14 and anti-TLR4 antibodies. These results suggest that endotoxin enhances NO production in Mg-deficient rat aortas directly, and that endotoxin receptors might, at least in part, contribute to this enhancement.