EMMPRIN expression is involved in the development of interstitial fibrosis and tubular atrophy in human kidney allografts.

BACKGROUND: Matrix metalloproteinases (MMP) are involved in the development of interstitial fibrosis and tubular atrophy (IF/TA) in renal disease. The synthesis of MMP is activated by the extracellular matrix metalloproteinases inducer protein (EMMPRIN). To analyze the role of EMMPRIN in IF/TA, we retrospectively detected EMMPRIN expression in specimens of human renal allografts with various levels of IF/TA. METHODS: Immunohistochemistry was performed to detect EMMPRIN expression. In a retrospective analysis, a total cohort of 50 specimens were divided according to BANFF-classification into four subgroups (0-3): no, mild (≤ 25%), moderate (26-50%), or severe (>50%) IF/TA. Among other parameters, renal function was analyzed and compared to EMMPRIN expression. RESULTS: In 24 of 38 biopsies, we detected positive EMMPRIN staining. All nephrectomy (n = 12) samples were negative for EMMPRIN. Positive staining in the biopsy samples was detectable on the basolateral side of tubular epithelial cells. EMMPRIN staining was negatively correlated with IF/TA (p < 0.001). We found significant differences between the mean EMMPRIN expression in IF/TA groups 0 and 3 (p = 0.021) and groups 1 and 3 (p = 0.004). Furthermore, we found significant correlations between EMMPRIN staining and renal function. CONCLUSION: Our data suggest that EMMPRIN is involved in the pathophysiology of IF/TA.

Head-to-Head Comparison of the RNA-Based Aptima Human Papillomavirus (HPV) Assay and the DNA-Based Hybrid Capture 2 HPV Test in a Routine Screening Population of Women Aged 30 to 60 Years in Germany.

Testing for E6/E7 mRNA in cells infected with high-risk (HR) human papillomavirus (HPV) might improve the specificity of HPV testing for the identification of cervical precancerous lesions. Here we compared the RNA-based Aptima HPV (AHPV) assay (Hologic) and the DNA-based Hybrid Capture 2 (HC2) HPV test (Qiagen) to liquid-based cytology (LBC) for women undergoing routine cervical screening. A total of 10,040 women, 30 to 60 years of age, were invited to participate in the study, 9,451 of whom were included in the analysis. Specimens were tested centrally by LBC, the AHPV test, and the HC2 test, and women who tested positive on any test were referred for colposcopy. Genotyping was performed on all HR-HPV-positive samples. Test characteristics were calculated based on histological review. As a result, we identified 90 women with cervical intraepithelial neoplasia grade 2+ (CIN2+), including 43 women with CIN3+. Sensitivity differences between the AHPV test and the HC2 test in detecting CIN2+ (P = 0.180) or CIN3+ (P = 0.0625) lesions were statistically nonsignificant. Of three CIN3 cases that were missed with the AHPV test, two cases presented lesion-free cones and one had a non-HR HPV67 infection. The specificity (

Assessment of tumor vascularity in lung cancer using volume perfusion CT (VPCT) with histopathologic comparison: a further step toward an individualized tumor characterization.

OBJECTIVE: To measure perfusion in different lung cancer subtypes and compare results with histopathological/immunohistochemical results. METHODS: Seventy-two consecutive untreated patients with lung cancer (40 adenocarcinomas, 20 squamous cell, and 12 small cell lung cancers) were enrolled. A 40-second volume perfusion computed tomography of the tumor bulk was obtained. Blood flow (BF), blood volume (BV), and transit constant were determined. Tumor volume and tumor necrosis were determined on contrast-enhanced computed tomography. Pathologic specimens were assessed for microvessel density (MVD), hypoxia-induced transcription (hif-1/-2), and proliferation (Ki-67). RESULTS: Higher MVD is associated with higher BF and BV. Higher tumor grade leads to lower BF but increased necrosis and tumor volume. Markers of hypoxia were independent from perfusion parameters, extent of necrosis or MVD. Blood flow, BV, and MVD were not significantly different among lung cancer subtypes. Transit constant was significantly reduced in small cell lung cancer versus adenocarcinoma. CONCLUSIONS: Perfusion values are related to MVD and tumor grade but vary considerably among lung cancer subtypes.

The cryo-needle: a new tool for histological biopsies. A feasibility study.

PURPOSE: Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is a standard procedure for intrathoracic lymph node biopsies. The newly developed cryo-needle operates in a similar way to the EBUS-TBNA but is able to obtain specimens for histological evaluation. The purpose of this animal study was to evaluate the feasibility, effect, and safety of the cryo-needle biopsies. METHODS: Four EBUS-guided cryo-needle biopsies were obtained from a mediastinal lymph node of a healthy pig. In an open surgery approach, cryo-needle biopsies using activation times of 1, 2, and 3 s (A1/A2/A3) and needle biopsies using a 21-gauge EBUS-TBNA needle were obtained from mesenteric lymph nodes. Cryo-needle biopsies A2 were performed with (A2+) and without (A2-) an oversheath. The size, weight, percentage of lymphatic tissue and artefact-free area of each cryobiopsy were evaluated. Smears were made with the TBNA-needle aspirates to determine the number of lymphocytes per high-power field (HPF). The bleeding duration was measured. RESULTS: We successfully obtained EBUS-guided cryo-needle biopsies. The area and weight of the biopsies A3 and A2+ were significantly larger compared with A1 (1.7 ± 0.8 and 1.4 ± 0.3 vs. 0.9 ± 0.4 mm(2); 5.2 ± 2.4 and 3.4 ± 1.8 vs. 1.5 ± 0.7 mg). The percentage of lymphatic tissue of the cryobiopsies was 90 ± 25 and 98 % of samples were artefact-free. The number of lymphocytes/HPF of TBNA-needle smears was 128 ± 54.3. There was no difference in bleeding duration between the techniques. CONCLUSIONS: The cryo-needle yields large histological specimens of high quality.

Pathological complete response and sphincter-sparing surgery after neoadjuvant radiochemotherapy with regional hyperthermia for locally advanced rectal cancer compared with radiochemotherapy alone.

PURPOSE: To evaluate the influence of regional hyperthermia on rates of complete pathological response (pCR) and sphincter-sparing surgery in the context of an up-to-date radiochemotherapy protocol for locally advanced rectal cancer. METHODS: Between 2007 and 2010, 106 patients with locally advanced cancer of the middle and lower rectum were admitted to neoadjuvant radiochemotherapy either with (n = 61) or without (n = 45) regional hyperthermia. A retrospective comparison was performed between two groups: 45 patients received standard treatment consisting of 5040 cGy in 28 fractions to the pelvis and 5-fluorouracil (RCT group) and 61 patients received the same treatment in combination with regional hyperthermia (HRCT group). Target temperature was 40.5°C for at least 60 min. Total mesorectal excision was performed routinely. RESULTS: pCR was seen in 6.7% of patients in the RCT group and 16.4% in the HRCT group. Patients who received at least four hyperthermia treatments (n = 40) achieved a significantly higher pCR rate (22.5%) than the remaining 66 patients (p = 0.043). Rates of sphincter-sparing surgery were similar in both groups with 64% in the RCT group and 66% in HRCT. When considering only low-lying tumours located within 8 cm of the anal verge prior to treatment, the rate of sphincter-sparing surgery was 57% in the HRCT group compared with 35% in the RCT group (p = 0.077). CONCLUSION: The combination of regional hyperthermia and neoadjuvant radiochemotherapy may lead to an increased pCR rate in locally advanced rectal cancer. Patients with low-lying tumours especially may benefit when additional downsizing allows sphincter-preserving surgery.

Severe paraneoplastic hypereosinophilia in metastatic renal cell carcinoma.

BACKGROUND: Renal cell carcinoma can cause various paraneoplastic syndromes including metabolic and hematologic disturbances. Paraneoplastic hypereosinophilia has been reported in a variety of hematologic and solid tumors. We present the first case in the literature of severe paraneoplastic hypereosinophilia in a patient with renal cell carcinoma. CASE PRESENTATION: A 46 year-old patient patient with a history of significant weight loss, reduced general state of health and coughing underwent radical nephrectomy for metastasized renal cell carcinoma. Three weeks after surgery, the patient presented with excessive peripheral hypereosinophilia leading to profound neurological symptoms due to cerebral microinfarction. Systemic treatment with prednisolone, hydroxyurea, vincristine, cytarabine, temsirolimus and sunitinib led to reduction of peripheral eosinophils but could not prevent rapid disease progression of the patient. At time of severe leukocytosis, a considerable increase of cytokines associated with hypereosinophilia was measurable. CONCLUSIONS: Paraneoplastic hypereosinophilia in patients with renal cell carcinoma might indicate poor prognosis and rapid disease progression. Myelosuppressive therapy is required in symptomatic patients.

Detection of cholangiocarcinoma in vivo using miniprobe-based confocal fluorescence microscopy.

BACKGROUND & AIMS: The preoperative diagnosis of cholangiocarcinoma is associated with a low sensitivity. To overcome this limitation, a new imaging modality was evaluated to detect neoplasia in vivo in the biliary tract. METHODS: Fourteen patients with biliary strictures were examined. Mucosal imaging was performed with a miniaturized confocal laser scanning miniprobe introduced via the accessory channel of a cholangioscope. Thereafter, targeted biopsy specimens were taken from the same regions. RESULTS: All strictures could be reached. Presence of irregular vessels use confocal laser microscopy enabled prediction of neoplasia with an accuracy rate of 86%, sensitivity of 83%, and specificity of 88%. The respective numbers for standard histopathology were 79%, 50%, and 100%. The mean signal-to-noise-ratio of laser microscopic images acquired from malignant strictures differed significantly from those of benign origin (1.8 +/- 0.8 vs 2.6 +/- 1.0; P = .005). CONCLUSIONS: Miniprobe-based confocal laser scanning microscopy considerably increases sensitivity for the detection of biliary neoplasia and therefore represents a promising diagnostic approach.

Percutaneously assisted endoscopic surgery using a new PEG-minitrocar for advanced endoscopic submucosal dissection (with videos).

BACKGROUND: For endoscopic submucosal dissection (ESD), adequate exposure and visualization of the submucosa for controlled dissection is of eminent importance. OBJECTIVE: To determine the feasibility and safety of percutaneously assisted endoscopic surgery (PA-ES) with a new prototype PEG-minitrocar (PMT) for advanced ESD in a porcine model. INTERVENTIONS: Placement of the PMT was done in all pigs by the use of a modified pull-through technique. After endoscopic incision of the mucosa, traction was provided for ESD by grasping the incisional margins of the mucosa with a rigid forceps introduced through the PMT, enabling stepwise dissection of the exposed submucosa under direct vision. MAIN OUTCOME MEASUREMENTS: Feasibility and safety of the new PMT for PA-ES and en bloc resection of prespecified mucosal areas. RESULTS: The study started with acute experiments in 8 animals, followed by a 10-day survival study in another 8 pigs. A total of 20 mucosal pieces were resected. The sizes of the resected pieces varied up to 7.5 x 4.0 cm ex vivo. All but one could be resected en bloc. Percutaneous assistance resulted in an excellent exposure of the submucosal space and enabled stepwise dissection of the submucosal connective tissue. Neither the PMT nor advanced ESD led to relevant complications. CONCLUSIONS: We demonstrated the feasibility and safety of a new PMT for advanced ESD. With the use of PA-ES, mucosal pieces of various sizes can be resected en bloc in gastric locations that are difficult to access by flexible endoscopy alone.

In vivo histopathology for detection of gastrointestinal neoplasia with a portable, confocal miniprobe: an examiner blinded analysis.

BACKGROUND & AIMS: Confocal fluorescence microscopy (CFM) has been mentioned to be a promising tool for in vivo histology. Recently, a portable confocal miniprobe has been developed. Our aim was to evaluate the potential benefit of CFM for detection of gastrointestinal neoplasia. METHODS: A total of 47 patients with known or suspected neoplasia in the upper (n = 34) or lower gastrointestinal tract (n = 13) were examined with standard endoscopes. After mucolyis with 5-10 mL of acetic acid 1.5%, chromoendoscopy with 2-5 mL cresyl violet 0.25% was performed, with the substance also being used as a fluorophore for CFM. Real-time video sequences were recorded. Thereafter, biopsies were taken or mucosectomy/polypectomy was performed from the same examined area. All stored sequences were put into a random order and assessed by a pathologist and a gastroenterologist both blinded to any data. RESULTS: A total of 119 CFM video sequences were recorded of 85 benign or 34 neoplastic areas. Quality of CFM images was regarded too low in 24 (pathologist) and 14 sequences (gastroenterologist). For the pathologist, accuracy of CFM detecting neoplasia was 92.6% (suitable images) and 73.9% (intention to diagnose). The respective accuracy values for the gastroenterologist were 92.4% (suitable images) and 81.5% (intention to diagnose). Agreement between CFM and histopathology was excellent (kappa values, 0.821 and 0.817). CONCLUSIONS: We have demonstrated that CFM with a miniprobe has the potential to diagnose neoplasia during ongoing endoscopy. This system has the advantage that it can be used with standard endoscopes. Further studies are warranted for validation.

High HSP27 and HSP70 expression levels are independent adverse prognostic factors in primary resected colon cancer.

BACKGROUND: The expression of Heat Shock Proteins (HSPs) is increased in various cancers and has been shown to correlate with biological tumor behaviour. This study aimed to investigate the impact of HSP70, HSP60 and HSP27 expression in colon cancer. MATERIAL AND METHODS: HSP expression was determined by immunohistochemistry on a tissue microarray with 355 primary resected colon carcinomas of all stages. Expression patterns were correlated with pathologic features (UICC pTNM category, tumor grading) and survival. RESULTS: Expression of HSP27, HSP60 and HSP70 ranged from negative to high. There was no correlation between HSP27, HSP60 and HSP70 expression among each other and with UICC pT category, presence of lymph node or distant metastases or tumor grading. High HSP70 expression was associated with worse overall survival (p < 0.001) and was an independent prognostic factor (p = 0.004) in multivariate analysis including the pathological parameters mentioned above. For patients without lymph node or distant metastases (UICC stages I/II) and with complete tumor excision, HSP70 expression was the only independent prognostic factor for survival (p = 0.001) and superior to UICC pT category. In left sided UICC stage I/II carcinomas, high HSP27 expression also had adverse prognostic impact and was an independent prognostic factor (p = 0.016) besides HSP70 (p = 0.002). CONCLUSION: High HSP70 and HSP27 expression is associated with worse clinical outcome in colon cancer. Determination of tumoral HSP70 and HSP27 may be used as additional biomarker for risk stratification especially for UICC stage I/II patients.

Application of laser microdissection and quantitative PCR to assess the response of esophageal cancer to neoadjuvant chemo-radiotherapy.

Tissues are complicated three-dimensional structures, composed of different types of interacting cells. Since the cell population of interest might constitute only a minor fraction of the total tissue volume, the problem of tissue heterogeneity has been a major barrier to the molecular analysis of normal versus diseased tissue. Thus, tissue microdissection represents one of the most promising techniques in molecular pathology offering the link between morphology and genetic analysis since it was established in the early 1970s. These first applications and further developments in the techniques enable preparation of morphologically well described and circumscribed cell populations of either tumor cells or surrounding tissue or even cytology specimens without contamination of unwanted cells. Laser capture microdissection is suitable for the dissection of both paraffin embedded and fresh frozen material. Further applications of the dissected genomic material are isolation of DNA and RNA as described later on followed by PCR or RT-PCR and sequencing.

Paratesticular fibrous pseudotumor--an IgG4-related disorder?

Paratesticular fibrous pseudotumor (nodular periorchitis, inflammatory pseudotumor of the spermatic cord) is a rare, benign condition of unknown etiology characterized by solitary or multiple intrascrotal nodules composed of dense fibrous tissue with a variable, sometimes sparse inflammatory infiltrate. Based on certain similarities to other fibroinflammatory disorders characterized by infiltrates of IgG4-expressing plasma cells and recently subsumed under the heading of IgG4-mediated diseases, we investigated the plasma cell distribution and immunoglobulin isotypes in three cases of paratesticular fibrous and inflammatory pseudotumor. All three cases showed a high number of IgG4-positive plasma cells with an IgG4 to IgG ratio of 44-48%. This finding indicates that paratesticular fibrous pseudotumor might belong to the growing list of IgG4-related diseases, which by now includes such diverse entities as retroperitoneal fibrosis, sclerosing pancreatitis and cholangitis, Riedel's thyroiditis, or sclerosing sialadenitis.

Enhanced activation of epidermal growth factor receptor caused by tumor-derived E-cadherin mutations.

Mutations of the tumor suppressor E-cadherin and overexpression of the receptor tyrosine kinase epidermal growth factor receptor (EGFR) are among the most frequent genetic alterations associated with diffuse-type gastric carcinoma. Accumulating evidence suggests a functional relationship between E-cadherin and EGFR that regulates both proteins. We report that somatic mutation of E-cadherin is associated with increased activation of EGFR followed by enhanced recruitment of the downstream acting signaling components growth factor receptor binding protein 2 and Shc, and activation of Ras. Reduced complex formation of mutant E-cadherin - with an in frame deletion of exon 8 in the extracellular domain resulting in reduced adhesion and increased motility - with EGFR was observed compared with wild-type E-cadherin. We conclude that reduced binding of mutant E-cadherin to EGFR in a multicomponent complex or reduced stability of the complex may enhance EGFR surface motility, thereby facilitating EGFR dimerization and activation. Furthermore, reduced surface localization due to enhanced internalization of mutant E-cadherin compared with the wild-type protein was observed. The internalization of EGFR was decreased in response to epidermal growth factor stimulation in cells expressing mutant E-cadherin, suggesting that mutation of E-cadherin also influences the endocytosis of EGFR. Moreover, we show increased activation of EGFR in gastric carcinoma samples with mutant E-cadherin lacking exons 8 or 9. In summary, we describe activation of EGFR by mutant E-cadherin as a novel mechanism in tumor cells that explains the enhanced motility of tumor cells in the presence of an extracellular mutation of E-cadherin.

Combined analysis of Rac1, IQGAP1, Tiam1 and E-cadherin expression in gastric cancer.

Rho GTPases are a family of major regulators of E-cadherin-mediated cell adhesion that are implicated in the carcinogenic process by deregulated expression of the family members itself or of upstream modulators or downstream effectors. Combined investigation of the Rho GTPase Rac1, the effector protein IQGAP1 and the activator Tiam1 in relation to expression or mutation of E-cadherin in gastric adenocarcinomas has not been reported. The aim of the study was to determine the expression and prognostic significance of Rac1, IQGAP1, Tiam1 and E-cadherin in gastric adenocarcinomas. Gastric carcinomas of 76 patients were investigated immunohistochemically in a tissue microarray study for expression of Rac1, IQGAP1, Tiam1 and E-cadherin. Correlations with clinical and follow-up data were examined. Moderate or strong reactivity for Rac1 was observed in 46% and for Tiam1 in 56% of tumors. Expression of IQGAP1 was present in 59% and of E-cadherin in 87% of tumors. While Rac1 and E-cadherin expression were not related to prognosis, a trend was observed between a lack of IQGAP1 expression (log-rank 0.088) as well as presence of Tiam1 (log-rank 0.097) and favorable prognosis in Kaplan-Meier survival analysis. Expression of Rac1 was positively linked to IQGAP1 expression (P=0.007, r=0.343) and tended to be inversely associated with expression of E-cadherin (P=0.055, r=-0.245). In conclusion, we observed deregulated expression of Rac1, IQGAP1, Tiam1 and E-cadherin in gastric cancer. We present evidence that either upregulation (for Rac1 and IQGAP1) or downregulation (for Tiam1 and E-cadherin) occurs. Rac1 and E-cadherin expression were not related to prognosis, while trends pointing to favorable prognosis of patients with Tiam1 expression and a lack of IQGAP1 expression were observed. These results indicate that the investigated regulators of E-cadherin-mediated cell adhesion play a role in gastric carcinogenesis.

High-resolution miniprobe-based confocal microscopy in combination with video mosaicing (with video).

BACKGROUND: A new portable confocal laser microscopy system has recently been developed. OBJECTIVE: Our purpose was to evaluate the feasibility of performing real-time microscopic imaging with a prototype of a new high-resolution miniprobe in conjunction with a video mosaicing algorithm. DESIGN: Feasibility study. SETTING: Tertiary referral center at a large university hospital. PATIENTS: Seven patients referred for endoscopy for various indications. MAIN OUTCOME MEASUREMENTS: High-resolution laser microscopy of the upper and lower GI tract was performed with standard endoscopes. Seven to 10 mL of 1% fluorescein was injected intravenously a few seconds before the procedure. No additional preparation was required. The prototype used has a lateral resolution of 1.2 microm and an axial resolution of 3 mum with a total field of 240 x 200 microm. From all stored video sequences a video mosaicing algorithm was used to combine the successive individual images, cancel motion artifacts, and reconstitute panoramas of the tissues. RESULTS: Cell-to-cell borders, single cell structures, and mucosal inflammation was readily detectable. By the use of the mosaicing algorithm, the image area could be increased 2- to 4-fold, and image definition could be further enhanced to allow finer detail visualization. LIMITATIONS: Low number of patients, early feasibility study. CONCLUSIONS: Our preliminary data show that high-resolution miniprobe-based confocal fluorescence microscopy in conjunction with video mosaicing has the potential to provide images similar to standard histopathologic studies. Dynamic images with a smaller field of view can be combined to reconstruct still images of high resolution covering a fairly large area.