Comparison of Minimally Invasive Surgery with Open Surgery for Remnant Gastric Cancer: A Multi-institutional Cohort Study.

BACKGROUND: Despite growing evidence of the effectiveness of minimally invasive surgery (MIS) for primary gastric cancer, MIS for remnant gastric cancer (RGC) remains controversial due to the rarity of the disease. This study aimed to evaluate the surgical and oncological outcomes of MIS for radical resection of RGC. PATIENTS AND METHODS: Patients with RGC who underwent surgery between 2005 and 2020 at 17 institutions were included, and a propensity score matching analysis was performed to compare the short- and long-term outcomes of MIS with open surgery. RESULTS: A total of 327 patients were included in this study and 186 patients were analyzed after matching. The risk ratios for overall and severe complications were 0.76 [95% confidence interval (CI): 0.45, 1.27] and 0.65 (95% CI: 0.32, 1.29), respectively. The MIS group had significantly less blood loss [mean difference (MD), -409 mL; 95% CI: -538, -281] and a shorter hospital stay (MD, -6.5 days; 95% CI: -13.1, 0.1) than the open surgery group. The median follow-up duration of this cohort was 4.6 years, and the 3-year overall survival were 77.9% and 76.2% in the MIS and open surgery groups, respectively [hazard ratio (HR), 0.78; 95% CI: 0.45, 1.36]. The 3-year relapse-free survival were 71.9% and 62.2% in the MIS and open surgery groups, respectively (HR, 0.71; 95% CI: 0.44, 1.16). CONCLUSIONS: MIS for RGC showed favorable short- and long-term outcomes compared to open surgery. MIS is a promising option for radical surgery for RGC.

Oestrogen receptor α in T cells controls the T cell immune profile and glucose metabolism in mouse models of gestational diabetes mellitus.

AIMS/HYPOTHESIS: The imbalance between maternal insulin resistance and a relative lack of insulin secretion underlies the pathogenesis of gestational diabetes mellitus (GDM). Alterations in T cell subtypes and increased levels of circulating proinflammatory cytokines have been proposed as potential mechanisms underlying the pathophysiology of insulin resistance in GDM. Since oestrogen modulates T cell immunity, we hypothesised that oestrogen plays a homeostatic role in visceral adipose tissue by coordinating T cell immunity through oestrogen receptor α (ERα) in T cells to prevent GDM. METHODS: Female CD4-cre ERαfl/fl (KO) mice on a C57BL/6 background with ERα ablation specifically in T cells, and ERαfl/fl (ERα-floxed [FL]) mice were fed 60 kJ% high-fat diet (HFD) for 4 weeks. Female mice mated with male BALB/c mice to achieve allogenic pregnancy and were maintained on an HFD to generate the GDM model. Mice were divided into four experimental groups: non-pregnant FL, non-pregnant KO, pregnant FL (FL-GDM) and pregnant KO (KO-GDM). GTTs and ITTs were performed on day 12.5 or 13.5 and 16.5 after breeding, respectively. On day 18.5 after breeding, mice were killed and T cell subsets in the gonadal white adipose tissue (gWAT) and spleen were analysed using flow cytometry. Histological examination was also conducted and proinflammatory gene expression in gWAT and the liver was evaluated. RESULTS: KO mice that mated with BALB/c mice showed normal fertility rates and fetal weights as compared with FL mice. Body and tissue weights were similar between FL and KO mice. When compared with FL-GDM mice, KO-GDM mice showed decreased insulin secretion (serum insulin concentration 15 min after glucose loading: 137.3 ± 18.3 pmol/l and 40.1 ± 36.5 pmol/l, respectively; p < 0.05), impaired glucose tolerance (glucose AUC in GTT: 2308.3 ± 54.0 mmol/l × min and 2620.9 ± 122.1 mmol/l × min, respectively; p < 0.05) and increased numbers of T helper (Th)17 cells in gWAT (0.4 ± 0.0% vs 0.8 ± 0.1%; p < 0.05). However, the contents of Th1 and regulatory T cells (Tregs) in gWAT remained similar between FL-GDM and KO-GDM. Glucose-stimulated insulin secretion was similar between isolated islets derived from FL and KO mice, but was reduced by IL-17A treatment. Moreover, the levels of proinflammatory gene expression, including expression of Emr1 and Tnfa in gWAT, were significantly higher in KO-GDM mice than in FL-GDM mice (5.1-fold and 2.7-fold, respectively; p < 0.01 for both). Furthermore, KO-GDM mice showed increased expression of genes encoding hepatokines, Ahsg and Fgf21 (both were 2.4-fold higher vs FL-GDM mice; p < 0.05 and p = 0.09, respectively), with no changes in inflammatory gene expression (e.g., Tnfa and Ifng) in the liver compared with FL-GDM mice. CONCLUSIONS/INTERPRETATION: Deletion of ERα in T cells caused impaired maternal adaptation of insulin secretion, changes in hepatokine profiles, and enhanced chronic inflammation in gWAT alongside an abnormal increase in Th17 cells. These results suggest that the ERα-mediated oestrogen signalling effects in T cells regulate T cell immunity and contribute to glucose homeostasis in pregnancy.

Suspected panosteitis in a crossbred calf.

A male crossbred calf developed a limp and pain upon deep pressure on the right hind limb and the right forelimb. The radiographic findings of affected limbs and pathological findings of bone biopsy were similar to those observed in canine panosteitis. This is the first case of suspected panosteitis reported in cattle.

Panostéite suspectée chez un veau de race croisée. Un veau mâle de race croisée a développé une boiterie et de la douleur à l'application d'une pression profonde sur la jambe arrière droite et la jambe avant droite. Les résultats de la radiographie des membres touchés et les résultats pathologiques d'une biopsie osseuse étaient semblables à ceux observés dans la panostéite canine. Il s'agit du premier cas de panostéite suspectée chez le bétail.(Traduit par Isabelle Vallières).

Osteoporotic fracture risk in women with breast cancer treated with aromatase inhibitors: a health insurance claims database study in Japan.

BACKGROUND: Hormone therapy with aromatase inhibitors (AIs) for estrogen receptor-dependent breast cancer may expose patients to an increased osteoporosis risk. This study was performed to estimate fracture risk in women with breast cancer to whom AIs were prescribed in Japan. METHODS: This retrospective study used data from the Japanese Medical Data Vision database. Women with breast cancer prescribed AIs over a 12-month period were identified and matched to women not prescribed AIs using a propensity score. Fracture rates were estimated by a cumulative incidence function and compared using a cause-specific Cox hazard model. The proportion of women undergoing bone density tests was retrieved. RESULTS: For all fractures sites combined, cumulative fracture incidence at 10 years was 0.19 [95%CI: 0.16-0.22] in women prescribed AIs and 0.18 [95%CI: 0.15-0.21] without AIs. AI prescription was not associated with any changes in risk (adjusted hazard ratio: 1.08 [95%CI: 0.99-1.17] p = 0.08). Women prescribed AI more frequently underwent bone density testing (31.9% [95% CI: 31.2%; 32.6%] versus 2.2% [95% CI: 2.0%; 2.4%]). CONCLUSIONS: The anticipated association between AI exposure and osteoporotic fracture risk in Japanese women with breast cancer was not seen clearly.

Long-Term Outcomes of Self-Expandable Metallic Stents as a Bridge to Surgery for Obstructive and Symptomatic Primary Tumors of Stage IV Colorectal Cancer: A Propensity-Score Analysis.

Background: Self-expandable metallic stent (SEMS) was introduced for the treatment of obstructive colorectal cancer (CRC) a few decades ago. However, its long-term outcomes remain controversial, especially for stage IV CRC. The aim of this study was to clarify the outcomes of SEMS as a "bridge to surgery" (BTS) for obstructive and symptomatic primary tumors in stage IV CRC by one-to-one propensity-score matching. Materials and Methods: This retrospective cohort study was conducted at a single center from January 2007 to December 2017. Patients with obstructive and symptomatic primary tumors of stage IV CRC underwent primary resection (PR) or placement of a SEMS as a BTS. They were divided into SEMS and PR groups, and their short- and long-term outcomes were compared. Results: In total, 52 patients were reviewed (SEMS group, 21; PR group, 31). Sixteen patients in both groups were matched using propensity scores. Patients in the SEMS group more frequently underwent laparoscopic surgery than those in the PR group (75% versus 19%, P = .004). The two groups showed no significant differences in perioperative and pathological outcomes. The 5-year overall survival was not significantly different between groups (29% versus 20%, P = .53). Conclusions: As a BTS, the use of SEMS for obstructive and symptomatic primary tumors in CRC stage IV can be a comparable option to PR in terms of short- and long-term outcomes, and would be less invasive with respect to surgical procedures.

A Rare Case of Severe Post-operative Pyoderma Gangrenosum After Surgery for Perforated Diverticulitis at the Sigmoid Colon.

Pyoderma gangrenosum (PG) is a nonbacterial ulcerating skin condition. It is typically associated with other systemic disorders. However, approximately 20%-30% of cases are idiopathic. Post-operative PG (PPG) is a rare type of PG with a rapidly expanding cutaneous ulcer at a surgical site and is often misdiagnosed as a wound infection. The difficulty in diagnosis can lead to unnecessary surgical interventions and delay in the treatment of PG. Herein, we present the case of a 68-year-old patient with severe PPG with no underlying diseases. He underwent an emergency laparotomy (Hartmann's procedure) for perforated diverticulitis. After the operation, systemic inflammatory response syndrome (SIRS) developed and the skin around the incisional wound, stoma, injection venous lines, and electrocardiogram monitoring pads gradually became erythematous. Skin biopsy and the absence of a source of infection confirmed the diagnosis of PG. Drug therapy for PG with steroids, and tumor necrosis factor-α inhibitors improved SIRS and the patient recovered.

How do magnetic resonance cholangiopancreatography findings predict conversion from laparoscopic cholecystectomy for acute cholecystitis to bailout procedures?

BACKGROUND: Acute cholecystitis is one of the most prevalent surgical abdominal conditions. The Tokyo Guidelines describe the management of acute cholecystitis and recommend bailout procedures for "difficult" cholecystitis cases. This study aimed to identify risk factors for conversion from laparoscopic cholecystectomy to bailout procedures in patients with acute cholecystitis. METHODS: This retrospective cohort study was conducted at a single center between January 2017 and December 2021. Patients who underwent laparoscopic cholecystectomy for acute cholecystitis were enrolled and classified into bailout and non-bailout groups. The patients' characteristics and perioperative data were compared between the 2 groups. RESULTS: In total, 161 patients who underwent laparoscopic cholecystectomy for acute cholecystitis were reviewed. Fourteen were excluded because of a lack of preoperative magnetic resonance cholangiopancreatography; thus, 147 patients were enrolled (bailout group, 21; non-bailout group, 126). Age (74 vs 67 years old; P = .048), days from onset to surgery (3 vs 2 days; P = .02), or defect of cystic duct in magnetic resonance cholangiopancreatography (57% vs 29%; P = .02) were significantly associated with conversion to bailout procedures. In the logistic regression analysis, a defect of the cystic duct in magnetic resonance cholangiopancreatography was an independent predictor for bailout procedures (odds ratio, 2.793; P = .04). CONCLUSION: In this study, defect of the cystic duct in the magnetic resonance cholangiopancreatography can predict conversion to bailout procedures. To the best of our knowledge, this is the first report to describe magnetic resonance cholangiopancreatography finding of the cystic duct as a predictor of surgical difficulty in patients with acute cholecystitis.

Japanese Papilio butterflies puddle using Na+ detected by contact chemosensilla in the proboscis.

Many butterflies acquire nutrients from non-nectar sources such as puddles. To better understand how male Papilio butterflies identify suitable sites for puddling, we used behavioral and electrophysiological methods to examine the responses of Japanese Papilio butterflies to Na(+), K(+), Ca(2+), and Mg(2+). Based on behavioral analyses, these butterflies preferred a 10-mM Na(+) solution to K(+), Ca(2+), and Mg(2+) solutions of the same concentration and among a tested range of 1 mM to 1 M NaCl. We also measured the ion concentrations of solutions sampled from puddling sites in the field. Na(+) concentrations of the samples were up to 6 mM, slightly lower than that preferred by butterflies in the behavioral experiments. Butterflies that sipped the 10 mM Na(+) solution from the experimental trays did not continue to puddle on the ground. Additionally, butterflies puddled at sites where the concentrations of K(+), Ca(2+), and/or Mg(2+) were higher than that of Na(+). This suggests that K(+), Ca(2+), and Mg(2+) do not interfere with the detection of Na(+) by the Papilio butterfly. Using an electrophysiological method, tip recordings, receptor neurons in contact chemosensilla inside the proboscis evoked regularly firing impulses to 1, 10, and 100 mM NaCl solutions but not to CaCl(2) or MgCl(2). The dose-response patterns to the NaCl solutions were different among the neurons, which were classified into three types. These results showed that Japanese Papilio butterflies puddle using Na(+) detected by the contact chemosensilla in the proboscis, which measure its concentration.

A Rare Case of the Coexistence of Pancreaticobiliary Maljunction and Gastrointestinal Tumor in Neurofibromatosis Type 1.

Neurofibromatosis type 1 (NF1) is a congenital condition characterized by "café au lait" spots and subcutaneous fibromas. There are various combined diseases, such as malignant tumors in the abdominal organs or brain tumors. Here, we present a case of a 35-year-old patient with a rare combination of NF1 with a gastrointestinal stromal tumor (GIST) and pancreaticobiliary maljunction (PBM). At the first visit, her main symptom was right upper abdominal pain. Radiological investigations revealed a common bile duct stone, submucosal tumor in the duodenum, PBM, and abnormal findings in the intrahepatic bile ducts. After the common bile duct stone was removed by endoscopic intervention, the patient underwent laparoscopic cholecystectomy, resection of the duodenal submucosal tumor, and liver biopsy. Pathological examination revealed chronic cholecystitis, GIST of the duodenum, and chronic inflammation of the intrahepatic bile ducts. This is the first case report of the rare coexistence of GIST and PBM in a patient with NF1.

Polo-like kinase 1 regulates cell proliferation and is targeted by miR-593* in esophageal cancer.

Polo-like kinase 1 (PLK1) is overexpressed in various human cancers. However, the biological functions and the post-transcriptional regulations of PLK1 in esophageal cancer (EC) are still unknown. The purposes of our study are to determine whether PLK1 can be a molecular target of EC therapy and to identify a microRNA (miRNA) targeting PLK1. We performed loss-of-function and gain-of-function experiments regarding cell proliferation, cell cycle, apoptosis, in vivo tumor formation and luciferase reporter assays, using siRNAs against PLK1 and miRNA. PLK1 protein was expressed in all 11 EC cell lines, but not in normal esophageal epithelial cells (HEEpiC). Knockdown of PLK1 in EC cells induced G2/M arrest (p < 0.001) in cell cycle assay and reduced cell proliferation (p = 0.019) and tumor formation ability in vivo (p < 0.0001). MiR-593*, identified as a miRNA targeting PLK1 by a database search, was less expressed especially in six EC cell lines than HEEpiC cells. Moreover, miR-593* expression level was inversely correlated with PLK1 mRNA level in 48 clinical tissue specimens of EC (p = 0.006). Introduction of synthetic miR-593* suppressed PLK1 expression by 69-73%, reduced cell proliferation (p = 0.008) and increased cell proportion of G2/M phase (p = 0.01) in HSA/c (an EC cells), whereas a miR-593* inhibitor upregulated PLK1 expression by 11-55%. Additionally, luciferase assay demonstrated that miR-593* interacted two binding sites in the PLK1 3'-UTR and reduced 56.8-71.5% of luciferase activity by degrading luciferase mRNA in HSA/c cells. In conclusion, PLK1 is post-transcriptionally regulated by miR-593* and could be a promising molecular target for EC treatment.

Activities to support individual dosimetry of children in Kawamata Town.

This paper does not necessarily reflect the views of the International Commission on Radiological Protection.Kawamata Town in Date District, Fukushima Prefecture is located more than 30 km north-west of Fukushima Daiichi nuclear power plant, but on 22 April 2011, part of the Yamakiya District of Kawamata Town was designated as a planned evacuation area. The exposure of children was a concern in Kawamata Town. Based on the proposal of Kindai University, Kawamata Town Board of Education took the initiative to measure individual radiation doses with an integrated dosimeter (glass badge) for all kindergarten children, nursery school children, elementary school students, and junior high school students in the town. These measurements were continued for nearly 3 years from June 2011 until the end of March 2014. The total number of measurements was approximately 16,800 across 11-cycle measurement, with 3 months' accumulation taken as one-cycle measurement. Kindai University provided financial support for the glass badge measurement service, and cooperated in the analysis of measured values and the development of advice based on the results. The main body implementing the measurements was Kawamata Town Board of Education, and the data obtained belong to Kawamata Town. When measurements were starting to be taken, schools got involved in the collection and distribution of dosimeters after explanations were provided to principals and school nurses who were in charge of risk communication. Thanks to the efforts of the schools, the recovery rate exceeded 90%, increasing the reliability of the measurements. It was clear who needed the information - the children and their parents. Kawamata Town Board of Education summarised the cumulative dose results for each measurement and notified parents via personal reports. These were sent to parents with advice on measurement results prepared by Kindai University, and care was taken to ensure that people could understand the measured results. Further briefing sessions were held as appropriate. At the briefing sessions, at the request of Kawamata Town Board of Education, the faculty members of Kindai University explained the measurement results from a professional point of view, and a professor from the Faculty of Medicine provided individual health consultations. Kawamata Town took the lead in using specialists to gain peace of mind, and this was key to the project's success. The situation was managed by taking measurements by dosimetry, and asking experts to interpret the data and provide advice to help reassure the residents.

Re-pancreaticojejunostomy for Necrosis of the Roux-en-Y Limb Tip 14 Years After Partington-Rochelle Procedure.

Longitudinal pancreaticojejunostomy for chronic pancreatitis, the Partington-Rochelle (PR) procedure, is a good option to control pain caused by dilation of the main pancreatic duct. However, long-term complications related to anastomosis are still unclear. Here, we present a case of a 78-year-old patient with sudden necrosis of the Roux-en-Y limb tip in a PR procedure performed 14 years ago. During emergent laparotomy, we resected the necrotic limb and re-anastomosed the remaining Roux-en-Y limb to the main pancreatic duct. Postoperatively, we managed the inflammation caused by the pancreatic fistula and successfully saved the patient by long-term drainage. Although the cause of necrosis is still unclear, mild kinking and stenosis of the Roux-en-Y limb might be associated with this situation.

The miR-106b-25 polycistron, activated by genomic amplification, functions as an oncogene by suppressing p21 and Bim.

BACKGROUND & AIMS: Barrett's esophagus (BE) is a highly premalignant disease that predisposes to the development of esophageal adenocarcinoma (EAC); however, the involvement of microRNAs (miRs) in BE-EAC carcinogenic progression is not known. METHODS: Esophageal cultured cells (HEEpiC, QhTRT, ChTRT, GihTRT, and OE-33) and esophageal tissues (22 normal epithelia, 24 BE, and 22 EAC) were studied. MiR microarrays and quantitative reverse-transcription polymerase chain reaction (RT-PCR) were employed to explore and verify differentially expressed miRs. Quantitative genomic PCR was performed to study copy number variation at the miR-106b-25 polycistron and MCM7 gene locus on chromosome 7q22.1. In vitro cell proliferation, cell cycle, and apoptosis assays and in vivo tumorigenesis experiments were performed to elucidate biologic effects of the miR-106b-25 polycistron. Western blotting and luciferase assays were performed to confirm direct messenger RNA (mRNA) targeting by the miR-106b-25 polycistron. RESULTS: The miR-106b-25 polycistron exerted potential proliferative, antiapoptotic, cell cycle-promoting effects in vitro and tumorigenic activity in vivo. MiRs-93 and -106b targeted and inhibited p21, whereas miR-25 targeted and inhibited Bim. This polycistron was upregulated progressively at successive stages of neoplasia, in association with genomic amplification and overexpression of MCM7. In addition, miRs-93 and -106b decreased p21 mRNA, whereas miR-25 did not alter Bim mRNA, suggesting the following discrete miR effector mechanisms: (1) for p21, mRNA degradation; (2) for Bim, translational inhibition. CONCLUSIONS: The miR-106b-25 polycistron is activated by genomic amplification and is potentially involved in esophageal neoplastic progression and proliferation via suppression of 2 target genes: p21 and Bim.

Estrogen regulates sex-specific localization of regulatory T cells in adipose tissue of obese female mice.

Regulatory T cells (Treg) play essential roles in maintaining immune homeostasis. Resident Treg in visceral adipose tissue (VAT-Treg) decrease in male obese mice, which leads to the development of obesity-associated chronic inflammations and insulin resistance. Although gender differences in immune responses have been reported, the effects of the difference in metabolic environment on VAT-Treg are unclear. We investigated the localization of VAT-Treg in female mice in comparison with that in male mice. On a high-fat diet (HFD), VAT-Treg decreased in male mice but increased in female mice. The increase was abolished in ovariectomized and HFD-fed mice, but was restored by estrogen supplementation. The IL33 receptor ST2, which is important for the localization and maturation of VAT-Treg in males, was reduced in CD4+CD25+ T cells isolated from gonadal fat of obese mice of both genders, suggesting that a different system exists for VAT-Treg localization in females. Extensive analysis of chemokine expression in gonadal fat and adipose CD4+CD25+T cells revealed several chemokine signals related to female-specific VAT-Treg accumulation such as CCL24, CCR6, and CXCR3. Taken together, the current study demonstrated sexual dimorphism in VAT-Treg localization in obese mice. Estrogen may attenuate obesity-associated chronic inflammation partly through altering chemokine-related VAT-Treg localization in females.

Chromosomal abnormalities and novel disease-related regions in progression from Barrett's esophagus to esophageal adenocarcinoma.

Barrett's esophagus (BE) is a metaplastic condition caused by chronic gastroesophageal reflux which represents an early step in the development of esophageal adenocarcinoma (EAC). Single-nucleotide polymorphism microarray (SNP-chip) analysis is a novel, precise, high-throughput approach to examine genomic alterations in neoplasia. Using 250K SNP-chips, we examined the neoplastic progression of BE to EAC, studying 11 matched sample sets: 6 sets of normal esophagus (NE), BE and EAC, 4 of NE and BE and 1 of NE and EAC. Six (60%) of 10 total BE samples and 4 (57%) of 7 total EAC samples exhibited 1 or more genomic abnormalities comprising deletions, duplications, amplifications and copy-number-neutral loss of heterozygosity (CNN-LOH). Several shared abnormalities were identified, including chromosome 9p CNN-LOH [2 BE samples (20%)], deletion of CDKN2A [4 BE samples (40%)] and amplification of 17q12-21.2 involving the ERBB2, RARA and TOP2A genes [3.1 Mb, 2 EAC (29%)]. Interestingly, 1 BE sample contained a homozygous deletion spanning 9p22.3-p22.2 (1.2 Mb): this region harbors only 1 known gene, basonuclin 2 (BNC2). Real-time PCR analysis confirmed the deletion of this gene and decreased the expression of BNC2 mRNA in the BE sample. Furthermore, transfection and stable expression of BNC2 caused growth arrest of OE33 EAC cells, suggesting that BNC2 functions as a tumor suppressor gene in the esophagus and that deletion of this gene occurs during the development of EAC. Thus, this SNP-chip analysis has identified several early cytogenetic events and novel candidate cancer-related genes that are potentially involved in the evolution of BE to EAC.

Gankyrin oncoprotein overexpression as a critical factor for tumor growth in human esophageal squamous cell carcinoma and its clinical significance.

To elucidate the possible involvement of gankyrin in ESCC progression and the effect of its down-regulation in ESCC, we investigated the expression of gankyrin in ESCC tissues comparing it with the corresponding normal esophageal epithelia and tested a short-hairpin RNA (shRNA) expression vector for gankyrin in ESCC cell lines. Gankyrin protein expression in 11 ESCC cell lines (KYSE series) was examined by RT-PCR and western blot. The expression of gankyrin mRNA in 30 ESCC tissues was compared with the corresponding normal epithelia by Real-time PCR. Expression of gankyrin protein was immunohistochemically analyzed in the ESCC of 103 patients. A gankyrin-shRNA vector was stably transfected into KYSE 170 cells to assess the role of gankyrin in cell motility, invasion and proliferation in vitro and tumor formation in vivo. Gankyrin expression increased in all 11 ESCC cell lines. Real-time PCR revealed that gankyrin expression was higher in the cancerous tissue for all 30 patients. In immunohistochemistry, gankyrin overexpression was correlated with lower survival rate (p = 0.0001), extent of the primary tumor, lymph node metastasis, distant lymph node metastasis and stage (p = 0.0072, p = 0.0004, p = 0.0172 and p = 0.0002, respectively). A shRNA vector against gankyrin repressed growth, cell motility, invasiveness in vitro and tumor formation in vivo. Gankyrin overexpression is associated with poor prognosis. It may play an important role in ESCC tumor progression and could be a potentially important therapeutic gene target in ESCC.

Promoter hypermethylation of CDH13 is a common, early event in human esophageal adenocarcinogenesis and correlates with clinical risk factors.

Although the CDH13 gene has been shown to undergo epigenetic silencing by promoter methylation in many types of tumors, hypermethylation of this gene in Barrett's-associated esophageal adenocarcinogenesis has not been studied. Two hundred fifty-nine human esophageal tissues were therefore examined for CDH13 promoter hypermethylation by real-time methylation-specific PCR. CDH13 hypermethylation showed discriminative receiver-operator characteristic curve profiles, sharply demarcating esophageal adenocarcinoma (EAC) from esophageal squamous cell carcinoma (ESCC) and normal esophagus (NE) (p < 0.0001). CDH13 normalized methylation values (NMV) were significantly higher in Barrett's esophagus (BE), dysplastic BE (D) and EAC than in NE (p < 0.0000001). CDH13 hypermethylation frequency was 0% in NE but increased early during neoplastic progression, rising to 70% in BE, 77.5% in D and 76.1% in EAC. Both CDH13 hypermethylation frequency and its mean NMV were significantly higher in BE with than without accompanying EAC. In contrast, only 5 (19.2%) of 26 ESCCs exhibited CDH13 hypermethylation. Furthermore, both CDH13 hypermethylation frequency and its mean NMV were significantly higher in EAC than in ESCC, as well as in BE or D vs. ESCC. Interestingly, mean CDH13 NMV was significantly lower in short-segment than in long-segment BE, a known clinical risk factor for neoplastic progression. Similarly, BE segment length was significantly lower in specimens with unmethylated than with methylated CDH13 promoters. 5-aza-2'-deoxycytidine treatment of OE33 EAC and KYSE220 ESCC cells reduced CDH13 methylation and increased CDH13 mRNA expression. These findings suggest that hypermethylation of CDH13 is a common, tissue-specific event in human EAC, occurs early during BE-associated neoplastic progression, and correlates with known clinical neoplastic progression risk factors.

Hypermethylation of the AKAP12 promoter is a biomarker of Barrett's-associated esophageal neoplastic progression.

The A-kinase anchoring protein 12 (AKAP12) is a kinase scaffold protein with known tumor suppressor activity. Recently, AKAP12 promoter hypermethylation was reported in gastric and colorectal cancers. We examined AKAP12 promoter hypermethylation using real-time methylation-specific PCR in 259 human esophageal tissues. AKAP12 hypermethylation showed highly discriminative receiver-operator characteristic (ROC) curve profiles, clearly distinguishing esophageal adenocarcinoma (EAC) from esophageal squamous cell carcinoma and normal esophagus (P < 0.0001). AKAP12-normalized methylation values were significantly higher in Barrett's metaplasia (BE), dysplastic Barrett's, and EAC than in normal esophagus (P < 0.0000001). AKAP12 hypermethylation frequency was zero in normal esophagus but increased early during neoplastic progression, to 38.9% in BE from patients with Barrett's alone, 52.5% in dysplastic Barrett's metaplasia, and 52.2% in EAC. AKAP12 hypermethylation levels were significantly higher in normal esophageal epithelia from patients with EAC (mean = 0.00082) than in normal esophagi from patients without Barrett's or esophageal cancer (mean = 0.00007; P = 0.006). There was a significant correlation between AKAP12 hypermethylation and BE segment length, a known clinical neoplastic progression risk factor. In contrast, only 2 (7.7%) of 26 esophageal squamous cell carcinomas exhibited AKAP12 hypermethylation. Treatment of BIC and OE33 EAC cells with 5-aza-2'-deoxycytidine reduced AKAP12 methylation and increased AKAP12 mRNA expression. AKAP12 mRNA levels in EACs with unmethylated AKAP12 (mean = 0.1663) were higher than in EACs with methylated AKAP12 (mean = 0.0668). We conclude that promoter hypermethylation of AKAP12 is a common, tissue-specific event in human EAC, occurs early during Barrett's-associated esophageal neoplastic progression, and is a potential biomarker for the early detection of EAC.

The suppression of aurora-A/STK15/BTAK expression enhances chemosensitivity to docetaxel in human esophageal squamous cell carcinoma.

PURPOSE: We previously reported that the expression of Aurora-A was frequently up-regulated in human esophageal squamous cell carcinoma (ESCC) tissues as well as cell lines and the up-regulation contributed to a poor prognosis. In this study, we assessed the possibility of Aurora-A suppression as a therapeutic target for ESCC using ESCC cell lines. EXPERIMENTAL DESIGN: We established subclones using vector-based short hairpin RNA (shRNA). Then, we investigated the effect of Aurora-A suppression on proliferation and cell cycle changes in vitro. Next, chemosensitivity against docetaxel was investigated by tetrazolium salt-based proliferation assay (WST assay) and cell number determinations, and furthermore, the type of cell death induced by docetaxel was analyzed by flow cytometry. Finally, to examine the effect of Aurora-A shRNA on proliferation and chemosensitivity against docetaxel in vivo, a s.c. tumor formation assay in nude mice was done. RESULTS: We established two genetically different stable cell lines (510 A and 1440 A) in which levels of Aurora-A were reduced. Cell growth was inhibited by 38.7% in 510 A and by 24.3% in 1440 A in vitro compared with empty vector-transfected controls (510 m and 1440 m), and this growth inhibition was mediated through G(2)-M arrest as confirmed by flow cytometry. Next, in a WST assay, the IC(50) for Aurora-A shRNA-transfected cells was lower than that of empty vector-transfected cells (510 A, 2.7 x 10(-7) mol/L; 510 m, 4.8 x 10(-7) mol/L; 1440 A, 2.6 x 10(-7) mol/L; 1440 m, 4.9 x 10(-7) mol/L). In addition, 0.3 nmol/L docetaxel induced a notable level of apoptosis in Aurora-A shRNA-transfected cells compared with empty vector-transfected cells. In the assay of s.c. tumors in nude mice, tumor growth in 510 A was inhibited by 36.1% compared with that in 510 m, and in tumors treated with docetaxel, the suppression of Aurora-A resulted in 44.0% tumor growth suppression in vivo. CONCLUSIONS: These results indicated that Aurora-A might play an important role in chemosensitivity to docetaxel, and the suppression of its expression might be a potential therapeutic target for ESCC.