Supermicrovascular anastomosis training using chicken wings and colored water.

Plastic surgeons require experience in supermicroscopic vascular anastomosis. Herein, we report a simple, rapid, and cost-effective training method using chicken wings and colored water. The avian ventral metacarpal artery was selected for dissection and anastomosis to mimic supermicrosurgery. Over 14 weeks (one anastomosis per day), the ulnar artery in 100 chicken wings was exposed by dissection, cut proximally, and injected with blue food dye-colored water by an inexperienced surgeon. After ligating the artery branches, it was cut and subjected to end-to-end anastomosis. Next, colored water was injected into the ulnar artery to check for suture sufficiency. The vessel was re-dissected to inspect the lumen and sutures qualitatively. Of the 100 wings, the first and last 20 wings' ventral metacarpal artery dissection, anastomosis times, and leakage frequency were compared. Avian ventral metacarpal artery diameter was recorded, and the cumulative anastomosis time where individual anastomosis times started decreasing was determined. Leakage rates before and after this point were compared. The avian ventral metacarpal artery diameter was 0.7-0.8 mm. The last 20 wings had significantly shorter median dissection times (12:27 vs. 17:45 min), anastomosis times (9:02 vs. 12:29 min), and leakage rates (15% vs. 70%); more even stitching and parallel ligature points; and less vessel layer inversion than the first 20 wings. After a cumulative anastomosis time of 10 h 26 min, individual times sharply decreased, and the leakage rate decreased significantly (58.3% vs. 23.8%). The proposed method significantly improved supermicrosurgical anastomosis. Thus, we believe that this method will help surgeons improve their supermicrosurgical skills.

Activating GNAS and KRAS mutations in gastric foveolar metaplasia, gastric heterotopia, and adenocarcinoma of the duodenum.

BACKGROUND: Heterotopic gastric-type epithelium, including gastric foveolar metaplasia (GFM) and gastric heterotopia (GH), is a common finding in duodenal biopsy specimens; however, there is still controversy regarding their histogenetic backgrounds. METHODS: We analysed a total of 177 duodenal lesions, including 66 GFM lesions, 81 GH lesions, and 30 adenocarcinomas, for the presence of GNAS, KRAS, and BRAF mutations. RESULTS: Activating GNAS mutations were identified in 27 GFM lesions (41%) and 23 GH lesions (28%). The KRAS mutations were found in 17 GFM lesions (26%) and 2 GH lesions (2%). A BRAF mutation was found in only one GFM lesion (2%). These mutations were absent in all 32 normal duodenal mucosa specimens that were examined, suggesting a somatic nature. Among the GFM lesions, GNAS mutations were more common in lesions without active inflammation. Analyses of adenocarcinomas identified GNAS and KRAS mutations in 5 (17%) and 11 lesions (37%), respectively. Immunohistochemically, all the GNAS-mutated adenocarcinomas diffusely expressed MUC5AC, indicating gastric epithelial differentiation. CONCLUSIONS: A significant proportion of GFM and GH harbours GNAS and/or KRAS mutations. The common presence of these mutations in duodenal adenoma and adenocarcinoma with a gastric epithelial phenotype implies that GFM and GH might be precursors of these tumours.

Frequent MED12 mutations in phyllodes tumours of the breast.

BACKGROUND: Phyllodes tumours are rare fibroepithelial tumours of the breast, that include benign, borderline, and malignant lesions. Although the molecular basis of phyllodes tumours largely remains unknown, a recent exome study identified MED12 mutations as a sole recurrent genetic alteration in fibroadenoma, a common benign fibroepithelial tumour that shares some histological features with the phyllodes tumour. METHODS: Forty-six phyllodes tumours and 58 fibroadenomas of the breast were analysed for MED12 mutations by using Sanger sequencing. RESULTS: MED12 mutations were identified in 37 out of the 46 phyllodes tumours (80%). The prevalence of MED12 mutations was similar among benign (15/18, 83%), borderline (12/15, 80%), and malignant tumours (10/13, 77%). MED12 mutations were also identified in 36 of the 58 fibroadenomas (62%). The mutations were frequent among intracanalicular-type (24/32, 75%) and complex-type lesions (4/6, 67%), but were significantly less common among the pericanalicular-type lesions (8/20, 40%). A microdissection-based analysis showed that MED12 mutations were confined to the stromal components in both phyllodes tumours and fibroadenomas. CONCLUSIONS: MED12 mutations were frequent among the phyllodes tumours of the breast, regardless of the tumour grade. Phyllodes tumours and fibroadenomas share, at least in part, a common genetic background.

NOTCH1 expression predicts patient prognosis in esophageal squamous cell cancer.

PURPOSE: The prognosis of patients with esophageal cancer remains poor, and the classification of tumor node metastasis has proven insufficient to predict patient prognosis. Therefore, novel predictive markers of esophageal cancer prognosis are needed. Notch receptors and their ligands have been reported to be upregulated in cervical, lung, colon, renal, and pancreatic cancers, but NOTCH1 expression has not been studied in esophageal cancer. METHODS: Expression of NOTCH1 was quantified by real-time reverse transcription-polymerase chain reaction in 55 primary esophageal squamous cell carcinomas (ESCCs) and their paired normal esophageal mucosa. We then examined the correlations between NOTCH1 expression, clinicopathological factors, and prognosis in patients with ESCC. RESULTS: The probability of overall survival was significantly lower for patients with high NOTCH1 expression (p = 0.0028; log-rank test). Overexpression of NOTCH1 was identified as a significant and independent prognostic factor (p = 0.0061) in patients who had undergone surgical treatment for ESCCs. The hazard ratio for predicting early death was 4.298 (95% confidence interval 1.515-12.195) for high versus low NOTCH1 expression. CONCLUSIONS: Our data indicate that NOTCH1 may be a candidate molecular prognostic marker and a molecular target for the development of an effective therapeutic intervention for patients with ESCC.

Frequent GNAS mutations in low-grade appendiceal mucinous neoplasms.

BACKGROUND: The molecular basis for the development of appendiceal mucinous tumours, which can be a cause of pseudomyxoma peritonei, remains largely unknown. METHODS: Thirty-five appendiceal mucinous neoplasms were analysed for GNAS and KRAS mutations. A functional analysis of mutant GNAS was performed using a colorectal cancer cell line. RESULTS: A mutational analysis identified activating GNAS mutations in 16 of 32 low-grade appendiceal mucinous neoplasms (LAMNs) but in none of three mucinous adenocarcinomas (MACs). KRAS mutations were found in 30 LAMNs and in all MACs. We additionally analysed a total of 186 extra-appendiceal mucinous tumours and found that GNAS mutations were highly prevalent in intraductal papillary mucinous tumours of the pancreas (88%) but were rare or absent in mucinous tumours of the colorectum, ovary, lung and breast (0-9%). The prevalence of KRAS mutations was quite variable among the tumours. The introduction of the mutant GNAS into a colorectal cancer cell line markedly induced MUC2 and MUC5AC expression, but did not promote cell growth either in vitro or in vivo. CONCLUSION: Activating GNAS mutations are a frequent and characteristic genetic abnormality of LAMN. Mutant GNAS might play a direct role in the prominent mucin production that is a hallmark of LAMN.

[Treatment of surgical site infection and hypertrophic scars].

For the treatment of surgical site infection (SSI), removal of foreign body materials and débridement of infected/necrotized tissues should be performed after careful imaging diagnosis. Then, wound irrigation should be performed everyday. For critically colonized wounds, iodine preparations are useful. If healthy granulation tissues start to grow, wound dressing with the basic fibroblast growth factor (bFGF) spray and/or prostaglandin E1 (PGE1) ointment should be recommended. Poor increase in granulation tissues, is a good indication for vacuum assisted closure (VAC) therapy. In Japan, the V.A.C. system is the only device for negative pressure wound therapy (NPWT) that is covered by the health insurance scheme. If entire wounds are covered by healthy granulation, chest wall reconstruction using skin graft or flap can be considered. For the treatment of hypertrophic scars (HSs), multimodal therapies should be performed. Tranilast and/or Saireito are widely used as internal agents in Japan. Corticosteroid ointments/creams/tapes/ injections are useful to decrease chronic inflammation in HSs. Moreover, wound stabilization, compression and keeping the affected area moist are effective to decrease subjective symptoms including itch and pain. Surgery, radiation therapy, pulsed dye laser, neodymium yttrium aluminum garnet(Nd:YAG) laser, make-up therapy such as rehabilitation make-up should be selected on a case-by-case basis.

An artificially constructed radiation-responsive promoter is activated by doxorubicin.

We previously developed an artificially constructed promoter that was activated in response to X-ray irradiation in LNCap, a prostate cancer cell line. Anticancer drugs were examined to see whether some of them could stimulate the activity of the promoter. It was found that doxorubicin (Dox) treatment to LNCap transfected with a gene cassette of the luciferase gene under control of the promoter-enhanced luciferase activity in a dose-dependent manner, indicating that the promoter could be controlled by Dox. When the luciferase gene was replaced with the fcy::fur gene whose product facilitates conversion of 5-fluorocytosine into 5-fluorouracil that is highly toxic, Dox stimulated the expression of the gene product, resulting in facilitation of cell killing effect in the presence of 5-fluorocytosine. These results suggest that therapeutic gene expression controlled with an anticancer drug may lead to a more effective cancer therapy with less hazardous side effects.

Regulation of gene expression in prostate cancer cells with an artificially constructed promoter responsive to radiation.

A promoter library was developed that is composed of DNA fragments constructed by randomly elongating the cis-acting elements of transcription factors presumably activated in prostate cancer by radiation, and linking to the TATA-box sequence. One promoter with the strongest reactivity to X-ray in the LNCap cells of the library was chosen and improved by the introduction of random mutations. The resultant promoter was designated clone 880-8, showing the highest dose-dependent activity enhancement with X-ray irradiation (X-irradiation). A recombinant retrovirus expressing the luciferase gene under the control of clone 880-8 was infected into LNCap cells that showed 9.12±0.36-fold enhancement of luciferase activity 12 h after X-irradiation at 10 Gy. When the infected cells were inoculated onto nude mice, enhancement of luciferase expression was 4.27±1.36-fold 12 h after X-irradiation at 10 Gy. When LNCap was infected with another recombinant carrying the fcy::fur gene downstream from clone 880-8, fcy::fur expression was enhanced by X-irradiation. It was also shown to increase the dose-dependent cell killing ratio with 5-FC as compared with a counterpart without X-irradiation. These results suggest that the method used in this study is effective to construct a promoter responsive to stimulation. Such promoters can be used for stimulation-controlled gene therapies.

Propeller flaps: classification and clinical applications.

Propeller flaps feature a highly reliable reconstructive method, based on a perforator vessel. Since their introduction in 1991, a great variety of propeller flaps have been described, according to their shape and their potential of coverage. Indeed, these flaps have progressively been refined and modified, concerning their vascularity and space design. The authors present a classification of propeller flaps. This anatomical classification is necessary to understand the dissection procedure and the differences between the numerous types of propeller flaps nowadays described. It is the international classification, which should be used for the description and conception of these flaps.

Serum interleukin-5 levels correlate with disease activity of Churg-Strauss syndrome in a patient treated with a leucotriene receptor antagonist, pranlukast, and inhaled corticosteroid.

This case report demonstrates that interleukin (IL)-5 levels and eosinophil cationic protein (ECP) correlated well with disease activity of Churg-Strauss syndrome (CSS) in a patient receiving treatment with leucotriene receptor antagonist and inhaled corticosteroid. In addition, ECP was localized in the inflamed tissue. IL-5 levels may thus provide a clue to therapeutic efficacy in patients with CSS using leucotriene receptor antagonists and inhaled corticosteroid.

Identification of candidate genes involved in the radiosensitivity of esophageal cancer cells by microarray analysis.

Radiotherapy plays a key role in the control of tumor growth in esophageal cancer patients. To identify the patients who will benefit most from radiation therapy, it is important to know the genes that are involved in the radiosensitivity of esophageal cancer cells. Hence, we examined the global gene expression in radiosensitive and radioresistant esophageal squamous cell carcinoma cell lines. Radiosensitivities of 13 esophageal cancer cell lines were measured. RNA was extracted from each esophageal cancer cell line and a normal esophageal epithelial cell line, and the global gene expression profiles were analyzed using a 34 594-spot oligonucleotide microarray. In the clonogenic assay, one cell line (TE-11) was identified to be highly sensitive to radiation, while the other cell lines were found to be relatively radioresistant. We identified 71 candidate genes that were differentially expressed in TE-11 by microarray analysis. The up-regulated genes included CABPR, FABP5, DSC2, GPX2, NME, CBR3, DOCK8, and ABCC5, while the down-regulated genes included RPA1, LDOC1, NDN, and SKP1A. Our investigation provided comprehensive information on genes related to radiosensitivity of esophageal cancer cells; this information can serve as a basis for further functional studies.

Gene expression profiling of the response of esophageal carcinoma cells to cisplatin.

Cisplatin is the most common chemotherapeutic agent used in esophageal cancer. However, sensitivity to cisplatin varies greatly between patients. It is important to identify the gene(s) that are related to the sensitivity to cisplatin in esophageal cancer patients. The IC50 for cisplatin was measured for 15 esophageal cancer cell lines (TE1-5, TE8-15, KYSE140, and KYSE150). RNA was extracted from each of these cell lines and a normal esophageal epithelial cell line, namely, Het1A, and gene expression profiles were analyzed using an oligonucleotide microarray consisting of 34 594 genes. TE4 was highly resistant and TE12, 14, and 15 were sensitive to cisplatin. Thirty-seven genes were differentially expressed in the cisplatin-resistant esophageal cancer cell line. Our investigation provides a list of candidate genes that may be associated with resistance to cisplatin in esophageal cancer cells, which may serve as a basis for additional functional studies.

Relationship between expression of 5-fluorouracil metabolic enzymes and 5-fluorouracil sensitivity in esophageal carcinoma cell lines.

5-Fluorouracil (5-FU) is a key drug in the treatment of esophageal squamous cell carcinoma (ESCC). Gene expression of 5-FU metabolic enzymes such as thymidylate synthase (TS), thymidine phosphorylase (TP), dihydropyrimidine dehydrogenase (DPD) and orotate phosphoribosyl transferase (OPRT), has recently been investigated in order to predict the 5-FU sensitivity of several cancers. We examined the relationship between such gene expression and 5-FU sensitivity in 25 ESCC cell lines. TS, DPD, TP and OPRT mRNA levels were assessed by real-time polymerase chain reaction. The 50% inhibitory concentrations (IC50) of 5-FU in 25 ESCC cell lines were determined by cell proliferation assay. IC50 values for 5-FU ranged from 1.00 to 39.81 micromol/L. There were significant positive correlations between IC50 and TS mRNA expression (R(2) = 0.5781, P < 0.0001) and DPD mRNA expression (R(2) = 0.3573, P = 0.0016). There were no correlations between IC50 and TP or OPRT mRNA expression. TS and DPD mRNA expression levels may be useful indicators in predicting the anti-tumor activity of 5-FU in ESCC.

Decreased expression of NDRG1 is correlated with tumor progression and poor prognosis in patients with esophageal squamous cell carcinoma.

NDRG1 (N-myc downstream regulated gene-1) was reported to be necessary for p53-mediated apoptosis and to be regulated by PTEN (phosphatase and tensin homolog). In several cancers, it was suggested to be a tumor suppressor gene. Its significance in esophageal squamous cell carcinoma (ESCC) has not been studied. The objective of this study was to clarify the relation between clinicopathological and biologic factors in esophageal carcinoma and to determine the prognostic significance of the expression of NDRG1. Expression of NDRG1 mRNA was quantified by real-time reverse transcription polymerase chain reaction using a Lightcycler in 47 esophageal ESCC specimens. The data were analyzed with reference to clinicopathological factors. Among the esophageal cancer tissues, NDRG1 mRNA expression was significantly lower in tumors of more advanced pathological stage (0-I vs. II-IV; P = 0.0027) and local tumor invasion (T1-2 vs. T3-4; P = 0.0136). Patients who had low NDRG1 mRNA expression had a significantly shorter survival after surgery compared with patients who had high NDRG1 mRNA expression (log-rank test, P = 0.0478). Impaired NDRG1 expression may lead to more aggressive invasion of ESCC.

Preexpanded distant "super-thin" intercostal perforator flaps for facial reconstruction without the need for microsurgery.

BACKGROUND: Concept of the 'super-thin perforator flap' was introduced in 1994 by authors. Since then, various types of 'super-thin perforator flaps' were applied successfully especially for contour sensitive reconstruction such as face and neck. METHODS: Eleven patients requiring large flaps who presented with extensive disfiguring facial scar (male: seven cases, female: four cases). On the consideration of flaps' colour, texture and thickness requirements, the authors selected 'super-thin' anterior intercostal perforator flaps (AICP, range from 4 x 14 cm to 25 cm x 9 cm) for reconstruction purpose. First, tissue expanders (volume range from 800 cc to 1200 cc) were carefully inserted under the AICP. After the flaps were expanded for 2 months, distant scars were removed and the covering super-thinned flaps were transferred into recipient site. Two weeks later, pedicles in the anterior chest were cut down and flaps were transferred to replace all the left scars. RESULTS: Flap were survived without any complications. The colour, texture and thickness of the transferred flap were satisfactory, shrink of flaps were not observed after long term follow-up. The authors present a method of facial reconstruction that has the advantages of creating a large amount of thin tissue of both good colour and texture, without the need of microsurgery and few disadvantages of donor-site morbidity. The disadvantages are three-staged procedures, complications of tissue expansion and uncomfortable compulsory posture for patients. In our opinion, this is an alternative method of choice for reconstructing all large defects in the lower two-thirds of the face.