Thoracoscopic Esophagectomy for a Patient With Perforated Esophageal Epiphrenic Diverticulum After Kidney Transplantation: A Case Report.

A 58-year-old man who underwent cadaveric kidney transplantation twice presented to hospital with a perforated epiphrenic diverticulum. Computed tomography revealed epiphrenic diverticulitis and right pleural effusion. Upper gastrointestinal fibroscopy showed an epiphrenic diverticulum full of food residue. He was transferred to our hospital, where we performed percutaneous endoscopic gastrostomy under general anesthesia in the supine position before thoracoscopy. Thoracoscopic esophagectomy was performed in the semi-prone position under 6-10 mmHg artificial pneumothorax via the right thoracic cavity. We performed subtotal esophagectomy to remove sources of infection because the esophageal wall surrounding the diverticulum was too thick to close or to perform diverticulectomy. A cervical esophagostomy was constructed after the thoracic procedure. The patient was managed with continuous hemodiafiltration and administered immunosuppressants and steroids to preserve the transplanted kidney. Continuous hemodiafiltration was stopped on postoperative day (POD) 4. The patient was discharged from the intensive care unit on POD 10 and transferred to the original hospital on POD 24 for rehabilitation. The second operative stage was performed on POD 157 at our hospital. We performed gastric tube reconstruction via the ante-sternal route and anastomosed the tube to the cervical esophagus. The postoperative course was uneventful; the patient was transferred to the original hospital on POD 15 after the second operation. Minimally invasive surgery was sufficient to treat perforated epiphrenic diverticulum while preserving the transplanted kidney. We recommend completely removing the source of infection and reducing surgical invasiveness to preserve the transplanted kidney in cases of esophageal perforation following kidney transplantation.

IL-11 contribution to tumorigenesis in an NRF2 addiction cancer model.

The interaction between cancer cells and their microenvironment is an important determinant of the pathological nature of cancers, particularly their tumorigenic abilities. The KEAP1-NRF2 system, originally identified as a critical defense mechanism against oxidative stress, is often dysregulated in various human cancers forming solid tumors, resulting in the aberrant activation of NRF2. Increased accumulation of NRF2 in cancers is strongly associated with the poor prognoses of cancer patients, including those with lung and breast cancers. Multiple lines of evidence suggest that aberrantly activated NRF2 in cancer cells drives their malignant progression and that the cancer cells consequently develop 'NRF2 addiction.' Although the downstream effectors of NRF2 that are responsible for cancer malignancy have been extensively studied, mechanisms of how NRF2 activation contributes to the aggressive tumorigenesis remains to be elucidated. In this study, we found a significant correlation between NRF2 and IL-11 status in breast cancer patients. Based on a recent report demonstrating that IL-11 is induced downstream of NRF2, we examined the significance of IL-11 in NRF2-driven tumorigenesis with a newly established NRF2 addiction cancer model. Expression of Il11 was elevated during the tumorigenesis of the NRF2 addiction cancer model, but intriguingly, it was hardly detected when the cancer model cells were cultured in vitro. These results imply that a signal originating from the microenvironment cooperates with NRF2 to activate Il11. To the best of our knowledge, this is the first report showing the influence of the microenvironment on the NRF2 pathway in cancer cells and the contribution of NRF2 to the secretory phenotypes of cancers. Disruption of Il11 in the NRF2 addiction cancer model remarkably inhibited the tumorigenesis, suggesting an essential role of IL-11 in NRF2-driven tumorigenesis. Thus, this study suggests that IL-11 is a potential therapeutic target for NRF2-addicted breast cancers.

The Rho guanine nucleotide exchange factor ARHGEF5 promotes tumor malignancy via epithelial-mesenchymal transition.

Epithelial tumor cells often acquire malignant properties, such as invasion/metastasis and uncontrolled cell growth, by undergoing epithelial-mesenchymal transition (EMT). However, the mechanisms by which EMT contributes to malignant progression remain elusive. Here we show that the Rho guanine nucleotide exchange factor (GEF) ARHGEF5 promotes tumor malignancy in a manner dependent on EMT status. We previously identified ARHGEF5, a member of the Dbl family of GEFs, as a multifunctional mediator of Src-induced cell invasion and tumor growth. In the present study, ARHGEF5 was upregulated during tumor growth factor-ß-induced EMT in human epithelial MCF10A cells, and promoted cell migration by activating the Rho-ROCK pathway. ARHGEF5 was necessary for the invasive and in vivo metastatic activity of human colorectal cancer HCT116 cells. These findings underscore the crucial role of ARHGEF5 in cell migration and invasion/metastasis. An in vivo tumorigenesis assay revealed that ARHGEF5 had the potential to promote tumor growth via the phosphatidylinositol 3-kinase (PI3K) pathway. However, ARHGEF5 was not required for tumor growth in epithelial-like human colorectal cancer HCT116 and HT29 cells, whereas the growth of mesenchymal-like SW480 and SW620 cells depended on ARHGEF5. Induction of EMT by tumor necrosis factor-α or Slug in HCT116 cells resulted in the dependence of tumor growth on ARHGEF5. In these mesenchymal-like cells, Akt was activated via ARHGEF5 and its activity was required for tumor growth. Analysis of a transcriptome data set revealed that the combination of ARHGEF5 upregulation and E-cadherin downregulation or Snail upregulation was significantly correlated with poor prognosis in patients with colorectal cancers. Taken together, our findings suggest that EMT-induced ARHGEF5 activation contributes to the progression of tumor malignancy. ARHGEF5 may serve as a potential therapeutic target in a subset of malignant tumors that have undergone EMT.

ZEB1 induces EPB41L5 in the cancer mesenchymal program that drives ARF6-based invasion, metastasis and drug resistance.

Onset of the cancer mesenchymal program is closely associated with cancer malignancy and drug resistance. Among the different epithelial-mesenchymal transition (EMT)-associated transcriptional factors, ZEB1 has a key role in inducing the mesenchymal phenotypes and stem cell-like properties of different breast cancer cells. ARF6 and its effector AMAP1 are frequently overexpressed in breast cancer cells, and promote invasion, metastasis and drug resistance. EPB41L5 is induced during EMT, and mediates the disruption of E-cadherin-based cell-cell adhesion and the promotion of focal adhesion dynamics. Here we show that EPB41L5 is an integral component of the ARF6-based pathway, which is induced by ZEB1. We found that EPB41L5 is expressed at high levels in malignant breast cancer cells and binds to AMAP1. ZEB1 induced EPB41L5 both in cancer cells and normal cells. This relationship was recaptured with The Cancer Genome Atlas RNASeq data set, and correlated with the poor outcome of the patients. In contrast, diversified events, such as tumor growth factor ß1 stimulation, expression of SNAI1 and TP53 mutation, can each cause the induction of ZEB1 and EPB41L5, depending on the cellular context. Our results demonstrated that the ZEB1-EPB41L5 axis is at the core of the cancer mesenchymal program that drives ARF6-based invasion, metastasis and drug resistance of significant populations of primary breast cancers, and is tightly correlated with the poor outcomes of patients.

Molecular evidence that the genes for dioecism and monoecism in Spinacia oleracea L. are located at different loci in a chromosomal region.

Spinach (Spinacia oleracea L.) is widely known to be dioecious. However, monoecious plants can also occur in this species. Sex expression in dioecious spinach plants is controlled by a single gene pair termed X and Y. Our previous study showed that a single, incompletely dominant gene, which controls the monoecious condition in spinach line 03-336, should be allelic or linked to X/Y. Here, we developed 19 AFLP markers closely linked to the monoecious gene. The AFLP markers were mapped to a 38.2-cM chromosomal region that included the monoecious gene, which is bracketed between flanking markers with a distance of 7.1 cM. The four AFLP markers developed in our studies were converted into sequence-characterized amplified region (SCAR) markers, which are linked to both the monoecious gene and Y and are common to both populations segregating for the genes. Linkage analysis using the SCAR markers suggested that the monoecious gene (M) and Y are located in different intervals, between different marker pairs. Analysis of populations segregating for both M and Y also directly demonstrates linkage of the genes at a distance of ~12 cM. The data presented in this study may be useful for breeding dioecious and highly male monoecious lines utilized as the pollen parents for hybrid seed production, as well as for studies of the evolutionary history of sexual systems in this species, and can provide a molecular basis for positional cloning of the sex-determining genes.

Lysyl oxidase secreted by tumour endothelial cells promotes angiogenesis and metastasis.

BACKGROUND: Molecules that are highly expressed in tumour endothelial cells (TECs) may be candidates for specifically targeting TECs. Using DNA microarray analysis, we found that the lysyl oxidase (LOX) gene was upregulated in TECs compared with its expression in normal endothelial cells (NECs). LOX is an enzyme that enhances invasion and metastasis of tumour cells. However, there are no reports on the function of LOX in isolated TECs. METHODS: TECs and NECs were isolated to investigate LOX function in TECs. LOX inhibition of in vivo tumour growth was also assessed using ß-aminopropionitrile (BAPN). RESULTS: LOX expression was higher in TECs than in NECs. LOX knockdown inhibited cell migration and tube formation by TECs, which was associated with decreased phosphorylation of focal adhesion kinase (Tyr 397). Immunostaining showed high LOX expression in human tumour vessels in vivo. Tumour angiogenesis and micrometastasis were inhibited by BAPN in an in vivo tumour model. CONCLUSION: LOX may be a TEC marker and a possible therapeutic target for novel antiangiogenic therapy.

Biglycan is a specific marker and an autocrine angiogenic factor of tumour endothelial cells.

BACKGROUND: We isolated tumour endothelial cells (TECs), demonstrated their abnormalities, compared gene expression profiles of TECs and normal endothelial cells (NECs) by microarray analysis and identified several genes upregulated in TECs. We focused on the gene encoding biglycan, a small leucine-rich repeat proteoglycan. No report is available on biglycan expression or function in TECs. METHODS: The NEC and TEC were isolated. We investigated the biglycan expression and function in TECs. Western blotting analysis of biglycan was performed on sera from cancer patients. RESULTS: Biglycan expression levels were higher in TECs than in NECs. Biglycan knockdown inhibited cell migration and caused morphological changes in TECs. Furthermore, immunostaining revealed strong biglycan expression in vivo in human tumour vessels, as in mouse TECs. Biglycan was detected in the sera of cancer patients but was hardly detected in those of healthy volunteers. CONCLUSION: These findings suggested that biglycan is a novel TEC marker and a target for anti-angiogenic therapy.

Induction of hypertrophic chondrocyte-like phenotypes by oxidized LDL in cultured bovine articular chondrocytes through increase in oxidative stress.

OBJECTIVE: It has been reported that the lectin-like oxidized low-density lipoprotein (Ox-LDL) receptor 1 (LOX-1) is expressed by chondrocytes in osteoarthritis (OA) cartilage and that Ox-LDL binding to LOX-1 increases intracellular oxidative stress in cultured bovine articular chondrocytes (BACs). It was recently demonstrated that reactive oxygen species (ROS) induce hypertrophic differentiation of chondrocytes in the growth plate. It has also been shown that activated chondrocytes in OA have hypertrophic chondrocyte-like phenotypes. The purpose of this study was to determine whether Ox-LDL induces hypertrophic chondrocyte-like phenotypes in BACs. DESIGN: Changes in type X collagen (COL10) and runt-related transcription factor 2 (Runx2) mRNA expression in BACs after Ox-LDL stimulation were investigated using real-time polymerase chain reaction (PCR). Western blotting and immunofluorescent cell staining were used to investigate changes in protein level. The antioxidant N-acetyl cysteine (NAC) was used to ascertain whether oxidative stress is involved in COL10 and Runx2 expression. We induced LOX-1 knockdown cells using small interfering RNA (siRNA) to examine the receptor specificity of Ox-LDL. RESULTS: COL10 expression was upregulated by Ox-LDL in a time- and dose-dependent manner. Immunofluorescent staining showed that Ox-LDL increased COL10 production in the extracellular matrix. Ox-LDL-induced upregulation of COL10 was suppressed by pretreatment with NAC and siRNA. Expression of Runx2 was upregulated by Ox-LDL and H(2)O(2), and these effects were suppressed by NAC pretreatment. CONCLUSION: Ox-LDL binding to LOX-1 induces a hypertrophic chondrocyte-like phenotype through oxidative stress, indicating that Ox-LDL plays a role in the degeneration of cartilage.

Characteristics of prolactin-releasing response to salsolinol (SAL) and thyrotropin-releasing hormone (TRH) in ruminants.

The secretion of prolactin (PRL) is stimulated by thyrotropin-releasing hormone (TRH), and inhibited by dopamine (DA). However, we have recently demonstrated that salsolinol (SAL), a DA-derived endogenous compound, is able to stimulate the release of PRL in ruminants. The aims of the present study were to compare the characteristics of the PRL-releasing response to SAL and TRH, and examine the relation between the effects that SAL and DA exert on the secretion of PRL in ruminants in vivo and in vitro. Three consecutive intravenous (i.v.) injections of SAL (5mg/kg body weight (b.w.): 19.2micromol/kgb.w.) or TRH (1microg/kgb.w.: 2.8nmol/kgb.w.) at 2-h intervals increased plasma PRL levels after each injection in goats (P<0.05); however, the responses to SAL were different from those to TRH. There were no significant differences in each peak value between the groups. The rate of decrease in PRL levels following the peak was attenuated in SAL-treated compare to TRH-treated animals (P<0.05). PRL-releasing responses to SAL were similar to those to sulpiride (a DA receptor antagonist, 0.1mg/kgb.w.: 293.3nmol/kgb.w.). In cultured bovine anterior pituitary (AP) cells, TRH (10(-8)M) significantly increased the release of PRL following both 15- and 30-min incubation periods (P<0.05), but SAL (10(-6)M) did not increase the release during the same periods. DA (10(-6)M) completely blocked the TRH-induced release of PRL for a 2-h incubation period in the AP cells (P<0.05). Sulpiride (10(-6)M) reversed this inhibitory effect but SAL (10(-6)M) did not have any influence on the action of DA. These results show that the mechanism(s) by which SAL releases PRL is different from the mechanism of action of TRH. Furthermore, they also show that the secretion of PRL is under the inhibitory control of DA, and SAL does not antagonize the DA receptor's action.

Clinical features of chromosome 16q22.1 linked autosomal dominant cerebellar ataxia in Japanese.

Chromosome 16q22.1-linked autosomal dominant cerebellar ataxia (16q-ADCA) is strongly associated with a substitution in the puratrophin-1 gene. This locus overlaps with spinocerebellar ataxia type 4 (SCA4) which shows ataxia with prominent sensory axonal neuropathy. We found that 16q-ADCA is a common ADCA subtype in the Tohoku District of Japan. The clinical feature of Japanese 16q-ADCA is characterized as late-onset pure cerebellar ataxia.

Management and algorithm for focal nodular hyperplasia of the liver in children.

PURPOSE: The aim of this study was to determine an appropriate management plan for childhood and adolescent FNH, in particular to establish an algorithm for preoperative diagnosis and treatment. PATIENTS AND METHODS: Between 1985 and 2003, 4 children with FNH were diagnosed. Of these 4 patients, 3 (Group A) underwent tumor resection, and 1 (Group B) was treated by conservative management. Clinical data, pathological findings and follow-up were evaluated retrospectively. RESULTS: The 3 patients in Group A were symptomatic, while the 1 patient in Group B was asymptomatic. In 3 of 4 patients, a homogeneous tumor with a central stellate area was noted on abdominal ultrasonography, CT scan and MR imaging. In case 2, SPIO-enhanced MR imaging was useful for differentiating FNH from hepatocellular carcinoma. Though percutaneous needle biopsy was performed in case 3, a pathologically definitive diagnosis was impossible. An open biopsy was performed in case 4 and FNH was diagnosed. In case 4 treated by conservative management, the tumor size did not change during the 7 years after the diagnosis of FNH. CONCLUSION: FNH is usually treated conservatively because of the good evolutionary outcome of the lesion. Surgery is indicated in cases of complications, compressed adjacent organs, lesion progression, or for symptomatic patients. We advocate the use of less invasive SPIO-enhanced MR imaging instead of open biopsy when the diagnosis of focal liver lesions is not clear after contrast-enhanced CT scan and non-enhanced MR imaging.

Endobronchial ultrasonography with guide-sheath for peripheral pulmonary lesions.

The usefulness of endobronchial ultrasonography (EBUS) with guide-sheath (GS) as a guide for transbronchial biopsy (TBB) for diagnosing peripheral pulmonary lesions (PPL)s and for improving diagnostic accuracy was evaluated in this study. EBUS-GS-guided TBB was performed in 24 patients with 24 PPLs of < or =30 mm in diameter (average diameter=18.4 mm). A 20-MHz radial-type ultrasound probe, covered with GS was inserted via a working bronchoscope channel and advanced to the PPL in order to produce an EBUS image. The probe with the GS was confirmed to reach the lesion by EBUS imaging and X-ray fluoroscopy. When the lesion was not identified on the EBUS image, the probe was removed and a curette was used to lead the GS to the lesion. After localising the lesion, the probe was removed, and TBB and bronchial brushing were performed via the GS. Nineteen peripheral lesions (79.2%) were visualised by EBUS. All patients whose PPLs were visible on EBUS images subsequently underwent an EBUS-GS-guided diagnostic procedure. A total of 14 lesions (58.3%) were diagnosed. Even when restricted to PPLs <20 mm in diameter, the diagnostic sensitivity was 53%. In conclusion, endobronchial ultrasonography with guide sheath-guided transbronchial biopsy was feasible and effective for diagnosing peripheral pulmonary lesions.

Dysferlin mutations in Japanese Miyoshi myopathy: relationship to phenotype.

OBJECTIVE: To study dysferlin gene mutations and genotype-phenotype correlations in Japanese patients with Miyoshi myopathy (MM). BACKGROUND: MM is an autosomal recessive distal muscular dystrophy that arises from mutations in the dysferlin gene. This gene is also mutated in families with limb girdle muscular dystrophy 2B. METHODS: The authors examined 25 Japanese patients with MM. Genomic DNA was extracted from the peripheral lymphocytes of the patients. The PCR products of each of 55 exons were screened by single strand conformation polymorphism or direct sequencing from the PCR fragments. RESULTS: The authors identified 16 different mutations in 20 patients with MM; 10 were novel. Mutations in Japanese patients are distributed along the entire length of the gene. CONCLUSIONS: Four mutations (C1939G, G3370T, 3746delG, and 4870delT) are relatively more prevalent in this population, accounting for 60% of the mutations in this study. This study revealed that the G3370T mutation was associated with milder forms of MM and the G3510A mutation was associated with a more severe form.

Oxidative stress and predominant Abeta42(43) deposition in myopathies with rimmed vacuoles.

This study was undertaken to determine the C terminus of amyloid beta protein (Abeta), accumulated in vacuolated muscle fibers, and compare these findings to the level of oxidative stress. Eight patients with myopathies characterized by rimmed vacuoles (RVs) were analyzed. Monoclonal antibodies specific to Abeta40 or Abeta42(43) revealed that the Abeta42(43) immunoreactivity was solely distributed in the vacuolated muscle fibers, and that only a part was also immunopositive for anti-Abeta40. Quantitative analyses in four specimens, in which eight or more vacuolated muscle fibers were observed, revealed that the mean incidence of Abeta42(43)-positive muscle fibers was 79.5+/-6.2% in total vacuolated muscle fibers, whereas that of the Abeta40-positive fibers was 42.9+/-12.6%. The predominance of Abeta42(43) deposition was statistically significant ( P<0.05). Abeta deposition was then compared with the distribution of oxidative nucleic acid damage in muscle fibers using a monoclonal antibody against 8-hydroxy-2'-deoxyguanosine and 8-hydroxyguanosine (8OHdG&G). The cytoplasmic staining for anti-8OHdG&G was found not only in vacuolated muscle fibers, but also in other muscle fibers including morphologically normal ones. Positive staining was completely abolished by RNase pretreatment and, thus, was suggested to reflect an increase of cellular RNA oxidation. The distribution of 8OHdG&G was much broader than the Abeta deposition. These data suggest that Abeta42(43) is predominantly involved in the pathogenesis of muscle fiber degeneration with RVs, and that oxidative damage may precede Abeta deposition in muscle fibers and play a key role in the pathomechanism of myopathies with RVs.

A novel A97P amino acid substitution in alpha-galactosidase A leads to a classical Fabry disease with cardiac manifestations.

BACKGROUND: Fabry disease results from a genetic deficiency of alpha-galactosidase A (GLA) activity. Phenotype-genotype correlations in this condition have not as yet been fully elucidated. OBJECTIVE: To report a case of a male patient with classical Fabry disease and his mother, a heterozygous female with Fabry disease, showing cardiac involvement, and to identify the underlying GLA gene mutation in this particular phenotype. PATIENTS/METHODS: Genomic DNA was extracted from the patient, his mother and the unaffected family members. Biopsy specimens of skin, heart and kidney were examined using light and electron microscopy. The mutation was identified by polymerase chain reaction and direct sequencing and was confirmed by restriction enzyme fragment length polymorphism. RESULTS: The G-->C transversion was identified in codon 97 of the GLA gene and resulted in an A97P amino acid substitution that was a novel pathogenic GLA gene mutation. The male patient who had classical Fabry disease was hemizygous and his mother was heterozygous for this mutation. CONCLUSIONS: These results indicate that the A97P amino acid substitution in GLA might tend to induce classical Fabry disease.

Mutation in the caveolin-3 gene causes a peculiar form of distal myopathy.

The authors describe a patient with sporadic distal myopathy associated with reduced caveolin-3 in muscle fibers in which the muscle atrophy was restricted to the small muscles of the hands and feet. Gene analysis disclosed a heterozygous 80 G-->A substitution in the caveolin-3 gene that was identical to that of reported cases of elevated serum creatine kinase. This patient further demonstrated possible clinical heterogeneity of myopathies with mutations in the caveolin-3 gene.

Inhibitory activity of quinolones against DNA gyrase of Mycobacterium tuberculosis.

The in vitro inhibitory activities of quinolones against Mycobacterium tuberculosis DNA gyrase were measured. The 50% inhibitory concentrations (IC(50)s) of sitafloxacin (DU-6859a), sparfloxacin, ciprofloxacin and levofloxacin against supercoiling activity of DNA gyrase were 1.67, 4.80, 12.2 and 13.9 mg/L, respectively, and correlated well with their MICs. Two altered proteins of GyrA containing Ala-90Val, or Ala-90Val and Asp-94Gly were also purified and the inhibitory activities of the quinolones ranged from 12 to >83 times weaker than those against the wild-type enzyme. These results suggest that mutations in the corresponding genes confer quinolone resistance.