Low lymphatic vessel density associates with chronic rhinosinusitis with nasal polyps.

OBJECTIVES: Chronic rhinosinusitis with and without nasal polyps (CRSwNP and CRSsNP) and antrochoanal polyps (ACP) are different upper airway inflammation phenotypes with different pathomechanisms. In order to understand the development of tissue edema, the present study aimed to evaluate lymphatic vessel density in CRSsNP, CRSwNP and ACP. MATERIALS AND METHODS: 120 retrospective nasal and maxillary sinus specimens were stained immunohistochemically with a von Willebrand factor polyclonal antibody recognizing vascular and lymphatic endothelium, and with a podoplanin monoclonal antibody recognizing lymphatic endothelium. Vessels were studied by microscopy in a blinded fashion, and the vessel density and the relative density of lymphatic vessels were calculated. Patient characteristic factors and follow-up data of in average 9 years were collected from patient records. RESULTS AND CONCLUSION: In the nasal cavity, the low absolute and relative density of vessels and of lymphatic vessels was associated with CRSwNP and ACP tissues compared to control inferior turbinate. This was observed also in the inflammatory hotspot area. In the maxillary sinus, lower absolute and relative density of lymphatic vessels associated with the CRSwNP phenotype. High lymphatic vessel density in polyp tissue associated with the need for revision CRS-surgery. As a conclusion, low density of lymphatic vessels distinguished patients with CRSwNP not only in the hotspot area of polyp tissue, but also in maxillary sinus mucosa. Yet, higher lymphatic vessel density seems to associate with polyp recurrence. Further studies are still needed to explore if formation of nasal polyps could be diminished by intranasal therapeutics affecting lymphangiogenesis.

Indoleamine 2,3-dioxygenase expression is associated with chronic rhinosinusitis with nasal polyps and antrochoanal polyps.

Chronic rhinosinusitis without and with nasal polyps (CRSwNP and CRSsNP), and antrochoanal polyps are different phenotypes with different pathomechanisms. Indoleamine 2,3-dioxygenase (IDO) is an enzyme expressed in many cells involved in the catabolism of the essential amino acid tryptophan to kynurenine. IDO might have a role in allergic airway inflammation. The aim was to evaluate if IDO expression is associated with CRSsNP, CRSwNP, or ACP. One hundred fifty specimens from the nasal cavity and sinus mucosa were immunohistochemically stained with mAb anti-IDO. The expression of epithelial and leukocyte IDO was associated with CRSwNP and ACP. The presence of ASA intolerance, asthma, atopy, smoking and use of medication did not significantly change the results. The different expression of IDO could putatively indicate the differences in the pathomechanisms of CRSsNP, CRSwNP and ACP. Further studies on the role of IDO in upper airways pathologies are required.

Proliferative and anti-apoptotic activity of esophageal mucosa in gastroesophageal reflux disease is not affected by fundoplication: a 4-year follow-up study.

BACKGROUND: The role of fundoplication in the prevention of esophageal adenocarcinoma is controversial. Development of cancer is associated with proliferation and anti-apoptosis, for which little data exist regarding their response to fundoplication. METHODS: Ki-67 and Bcl-2 expression was assessed in the esophagogastric junction (EGJ) and the distal and proximal esophagus of 20 patients with gastroesophageal reflux disease (GERD) treated by fundoplication and in 7 controls. Endoscopy was performed preoperatively and 6 (20 patients) and 48 months (16 patients) postoperatively. RESULTS: There were positive correlations between Ki-67 and Bcl-2 levels in the EGJ (p > 0.001) and in the distal (p = 0.001) and proximal esophagus (p = 0.013). Compared to the preoperative level, Ki-67 expression was elevated in the distal (p = 0.012) and proximal (p = 0.007) esophagus at 48 months. In addition, compared to control values, Ki-67 expression was lower at the 6-month follow-up in the EGJ (p = 0.037) and the proximal esophagus (p = 0.003), and higher at the 48-month follow-up in the distal esophagus (p = 0.002). Compared to control values, Bcl-2 was lower at 6 months in the EGJ (p = 0.038). CONCLUSIONS: Proliferative activity after fundoplication increased in the long term in the distal esophagus despite a normal fundic wrap and healing of GERD.

Altered expression of HSP27 and HSP70 in distal oesophageal mucosa in patients with gastro-oesophageal reflux disease subjected to fundoplication.

BACKGROUND: Gastro-oesophageal reflux disease (GERD) is a risk factor for oesophageal adenocarcinoma. Although fundoplication cures reflux symptoms and oesophagitis, it remains controversial whether it is capable of preventing the development of oesophageal adenocarcinoma. Hsp27 and Hsp70 are associated with the development of cancer, whereas the effect of fundoplication on them is not known. METHODS: The expression of Hsp27 and Hsp70 was assessed semiquantitatively from biopsies of oesophageal mucosa for a prospective cohort of 19 patients with GERD treated with fundoplication and 7 controls without GERD. Upper gastrointestinal endoscopy with biopsies from the oesophagogastric junction (EGJ) and the distal and proximal oesophagus were performed preoperatively (19 patients) and after recovery from GERD at 6 (19 patients) and 48 months (16 patients) postoperatively. RESULTS: The expressions of both Hsp27 (p = 0.001) and Hsp70 (p = 0.002) in the distal oesophagus were lower in patients preoperatively and at 48 months postoperatively (p < 0.001 for both) than in controls. The patients' Hsp27 and Hsp70 levels were lower preoperatively in the proximal oesophagus (p = 0.048 for both) than in controls. Both Hsp27 (p = 0.002) and Hsp70 (p = 0.003) were lower in the distal oesophagus preoperatively and at 48 months postoperatively (p = 0.003 for Hsp27, p = 0.004 for Hsp70) than in the proximal oesophagus. CONCLUSIONS: Our results indicate that there may be some factor interfering with the mucosal defence system of the distal oesophagus in GERD that is uninfluenced by fundoplication and not associated with the acid-reflux-normalizing effect.

Differences in retinal neovascular tissue and vitreous humour in patients with type 1 and type 2 diabetes.

AIMS: The aim of the study was to evaluate the histopathology of neovascular tufts and vitreous samples collected from patients with diabetes. METHODS: Vitreous samples and neovascular tufts were collected from patients with type 1 (n = 13) and (n = 17) type 2 diabetes with proliferative retinopathy, and from controls with a macular hole (n = 5). Neovessels were analysed using immunohistochemistry and vitreous samples with an enzyme-linked immunosorbent assay (ELISA). The main outcome measure was to examine differences in the levels of growth factors in patients with type 1 and type 2 diabetes with proliferative retinopathy. RESULTS: Vascular endothelial growth factor (VEGF)-A was most strongly present in the samples from patients with type 1 diabetes. In type 2 diabetes, VEGF-D was more abundantly present than in type 1 diabetes. Angiopoietin (ANG)-2 was also abundantly present. Macrophages and nuclear factor kappa B (NFkappaB) were found, indicating the presence of an inflammatory process in the neovascular tissues. CONCLUSIONS: VEGF-A and ANG-2 are equally important in the neovascular process in both type 1 and type 2 diabetes. VEGF-D is abundantly present in type 2 diabetes. In order to achieve better control of diabetic retinopathy, it might be beneficial to develop treatments that prevent the actions of ANG-2 and VEGF-D.

Birch pollen allergen Bet v 1 binds to and is transported through conjunctival epithelium in allergic patients.

BACKGROUND: Previous work in type-I pollen allergies has mainly focused on lymphocytes and immune responses. Here, we begin to analyse with a systems biology view the differences in conjunctival epithelium obtained from healthy and allergic subjects. METHODS: Transcriptomics analysis combined with light and electron microscopic analysis of birch pollen allergen Bet v 1 located within conjunctival epithelial cells and tissues from birch allergic subjects and healthy controls was carried out. RESULTS: Bet v 1 pollen allergen bound to conjunctival epithelial cells within minutes after the exposure even during the nonsymptomatic winter season only in allergic, but not in healthy individuals. Light- and electron microscopy showed that Bet v 1 was transported through the epithelium within lipid rafts/caveolae and reached mast cells only in allergic patients, but not in healthy individuals. Transcriptomics yielded 22 putative receptors expressed at higher levels in allergic epithelium compared with healthy specimens. A literature search indicated that out of these receptors, eight (i.e. 37%) were associated with lipid rafts/caveolae, which suggested again that Bet v 1 transport is lipid raft/caveola-dependent. CONCLUSIONS: We show a clear difference in the binding and uptake of Bet v 1 allergen by conjunctival epithelial cells in allergic vs healthy subjects and several putative lipid raft/caveolar receptors were identified, which could mediate or be co-transported with this entry. The application of discovery driven methodologies on human conjunctival epithelial cells and tissues can provide new hypotheses worth a further analysis to the molecular mechanisms of a complex multifactorial disease such as type-I birch pollen allergy.

Ischemic intestinal injury during cardiopulmonary bypass does not show an association with neutrophil activation: a porcine study.

BACKGROUND: Neutrophil activation and tissue sequestration are crucial events in intestinal ischemia-reperfusion injury, but their role in the gut wall after clinical cardiopulmonary bypass (CPB) remains unclear. We tested whether local post-CPB inflammatory response in the gut wall would be associated with intestinal mucosal perfusion. METHODS: Twenty pigs underwent 60 min of aortic clamping and 75 min of normothermic perfusion. Intestinal biopsies were taken after 120 min of reperfusion. Based on ileal myeloperoxidase activity (MPO), the animals were divided into 2 groups, CPB-induced increase in MPO (MPO+) versus no such increase (MPO-), for comparison of the parameters that measure gut mucosal perfusion. Ileal p(CO)((2)) and intramucosal pH were determined, and arterial gases were analyzed. Additionally, several hemodynamic parameters and blood thrombin-antithrombin complexes (TAT) were measured. RESULTS: Myocyte degeneration, endothelial activation and vasculitis were more pronounced in the MPO+ group (p < 0.05), while the MPO- group showed significantly increased pi(CO)((2)) and lower mucosal pH values during reperfusion. Hemodynamics and TAT levels did not differ between the groups. CONCLUSION: Tissue sequestration of neutrophils was poorly associated with perturbed mucosal perfusion after CPB. Mechanisms of gut wall injury after a low-flow/reperfusion setting can differ from those in reperfusion injury after total ischemia.

Is Cystatin C more sensitive than creatinine in detecting early chronic allograft nephropathy?

BACKGROUND: Cystatin C (CyC) has been suggested as a more accurate indicator of renal function than creatinine (Crea). CyC performance against graft histopathology has not been investigated. AIM: To compare CyC and Crea-based methods as predictors of chronic allograft damage index (CADI). MATERIAL AND METHODS: 105 protocol biopsies obtained at 6 months post-transplantation were classified with Banff'97 and CADI. CyC and Crea were measured concomitantly. Histology was correlated to CyC, Crea, their reciprocals, CyC-estimated GFR (Larsson), Cockroft and Gault (C&G) and abbreviated MDRD using Kendall's Tau. The area under ROC curve (ROC-auc),sensitivity/specificity, positive and negative predictive values were calculated at CADI cut-off of 2. RESULTS: Mild histological changes were best revealed by Crea, although with modest sensitivity/ specificity. A Crea threshold of 111 micromol/l distinguished 74% of the patients with CADI > 2 and excluded this condition in 66%. For Crea, ROC-auc was 0.72 (p < 0.001). Crea and 1/Crea correlated best to CADI, chronic allograft nephropathy, chronic inflammation, tubular atrophy, vascular changes and glomerulopathy. Neither C&G nor MDRD improved Crea performance alone. CyC and Larsson formula performed the same (ROC-auc 0.67). A CyC threshold of 1.12 mg/l distinguished 69% of the patients with CADI > 2 and excluded it in 60%. Significant Tau correlation was found between CyC, 1/CyC and Larsson with CADI, chronic inflammation, tubular atrophy and chronic vascular changes. CONCLUSIONS: CyC, 1/CyC and Larsson-estimated GFR did not offer significant advantages over Crea in predicting mild histological allograft changes. Protocol biopsy provides information that cannot be sensitively predicted by biochemical measurements used in clinical practice.

Antithrombin reduces pulmonary hypertension during reperfusion after cardiopulmonary bypass in a pig.

BACKGROUND: Antithrombin (AT) may alleviate many cardiopulmonary bypass (CPB) and ischemia-reperfusion (I/R)-related adverse effects. Using a porcine model of clinical cardiac surgery on CPB, we tested the effects of supplementary AT on myocardial and lung I/R injury. METHODS: Twenty pigs undergoing 60-min aortic clamping and 75-min normothermic perfusion were randomized in a blinded setting to receive an intravenous (i.v.) bolus of AT (250 IU/kg) (AT group, n = 10) or placebo (n = 10) 15 min before aortic declamping. An additional group of five animals received 500 IU/kg AT in an open-label setting (AT+). Thrombin-antithrombin complexes (TAT), activated clotting times (ACT), AT and myeloperoxidase (MPO) activities, troponin T, and several hemodynamic parameters were measured before CPB and after weaning from CPB up to 120 min after aortic declamping. After 120 min of reperfusion, myocardial and lung biopsies were taken for histological examination. RESULTS: AT effectively inhibited coagulation as assessed by ACT. In the AT and AT+ groups only, cardiac output (CO) and stroke volume (SV) showed a trend of post-ischemic recovery during the first 15 min after CPB. AT-attenuated reperfusion induced an increase in pulmonary arterial diastolic pressure (PAPD) but did not have significant effects on systemic or pulmonary vascular resistance. The effects of AT on SV, CO, and PAPD were fortified in the AT+ group. AT did not show effects on inflammatory changes in either myocardial or pulmonary tissue specimens. AT did not reduce post-ischemic troponin T release. CONCLUSION: Supplementary AT, in doses with significant anticoagulant effect, did not alleviate myocardial I/R injury in terms of histological inflammatory changes or post-ischemic troponin T release. Instead, however, AT-attenuated reperfusion induced an increase in pulmonary pressure after CPB. Mechanisms and clinical implications of these effects remain to be explored.

Ezrin expression in tissue microarray of primary and recurrent gliomas.

Malignant progression, infiltrative growth pattern and recurrencies after surgery are characteristic features of diffuse gliomas. Ezrin is a membrane-cytoskeleton linker protein expressed in several types of neoplasms. In experimental models, increased ezrin expression correlates with invasion of malignant glioma cells. We studied ezrin expression and its correlation with patient survival in 229 primary and recurrent astrocytomas (WHO grades II-IV), oligodendrogliomas (II-III) and oligoastrocytomas (II-III) of 113 patients. Ezrin expression as evaluated by immunohistochemistry and immunoblotting was detected in all studied glioma types. Staining intensity and number of immunoreactive cells correlated with increasing malignancy of astrocytomas and oligoastrocytomas (P = 0.001). Ezrin expression was strongest in astrocytomas (P = 0.006). Also oligodendrogliomas were positive for ezrin. High ezrin expression in primary gliomas correlated with shorter time to recurrence (P < 0.05) and poor overall survival (P < 0.05) of the patients. Ezrin expression increased during progression of the tumours (P < 0.05). However, ezrin was not an independent prognostic factor. The results of this study show that ezrin expression is associated with progression of gliomas and correlates with histological cell type and WHO tumour grade.

Protocol core needle biopsy and histological chronic allograft damage index as surrogate endpoint for Long-Term graft survival.

Following encouraging results from several single-center studies showing that early histological manifestations of chronic rejection are seen in the graft before a decline in transplant function, we tested this concept in a multicenter study and investigated whether protocol needle biopsy may be used as a surrogate to late graft survival in multicenter renal transplantation trials. During two mycophenolate mofetil trials, 621 representative protocol biopsies were obtained at baseline, 1 year, and 3 years. The samples were coded and evaluated blindly by two pathologists and a Chronic Allograft Damage Index (CADI) score was constructed. At 1 year only 20% of patients had elevated (>1.5 mg/100 mL) serum creatinine, whereas 60% of the biopsies demonstrated an elevated (>2.0) CADI score. The mean CADI score at baseline, 1.3 +/- 1.1, increased to 3.3 +/- 1.8 at 1 year and to 4.1 +/- 2.2 at 3 years. The patients at 1 year were divided into 3 groups, those with CADI <2, between 2 and 3.9, and >4.0, the first two groups having normal (1.4 +/- 0.3 and 1.5 +/- 0.6 mg/dL) and the third group pathological (1.9 +/- 0.8 mg/dL) levels of serum creatinine. At 3 years there were no lost grafts in the "low" CADI group, six lost grafts (4.6%) in the "elevated" CADI group, and 17 lost grafts (16.7%) in the "high" CADI group (P <.001). One-year histological CADI score predicts graft survival even when the graft function is still normal. This observation makes it possible to use CADI as a surrogate endpoint in prevention trials and to identify the patients at risk for intervention trials.

Tumour microvessel density as predictor of chemotherapy response in breast cancer patients.

The aim of this study was to evaluate the predictive value of intratumoural microvessel density in breast cancer. We studied immunohistochemically primary tumours of 104 patients with metastasised breast cancer who took part in a randomised multicentre trial comparing docetaxel to sequential methotrexate and 5-fluorouracil. Vessels were highlighted with factor VIII staining and counted microscopically. Microvessel density was compared with clinical response to chemotherapy and patient survival. The microvessel density of the primary tumour was not significantly associated with patient's response to chemotherapy, time to progression or overall survival in the whole patient population or in the docetaxel or methotrexate and 5-fluorouracil groups. However, disease-free survival was longer in patients with low microvessel density (P=0.01). These findings suggest that microvessel density of the primary tumour cannot be used as a predictive marker for chemotherapy response in advanced breast cancer.

Effects of extrinsic denervation with or without ischemia-reperfusion injury on constitutional mucosal characteristics in porcine jejunoileum.

We investigated the effects of jejunoileal denervation with or without ischemia-reperfusion on mucosal characteristics and small intestinal structure. Growing pigs underwent sham laparotomy, jejunal transection, or extrinsic jejunoileal denervation with or without in situ ischemia-reperfusion. Small intestinal morphology, crypt cell proliferation, enterocyte ultrastructure, and disaccharidase activities were analyzed from jejunum and ileum after eight weeks. Immunohistological analysis of the ileum showed no staining of catecholaminergic neurons after extrinsic denervation. Neural isolation of the jejunoileum with or without ischemia-reperfusion injury reduced weight gain and villous enterocyte density in the ileum, abolished the proximodistal gradient of sucrase activity, and increased mucosal thickness, villus height, and villus surface area in the ileum. However, gross jejunoileal morphology, crypt cell proliferation, and enterocyte ultrastructure remained unchanged. In conclusion, jejunoileal denervation in growing pigs selectively modulates constitutional mucosal characteristics in the ileum, presumably due to altered enterocyte turnover, without a decrease in small intestinal absorptive surface area. These changes are independent of short ischemia and subsequent reperfusion.

Autotransplantation modulates ileal enteroendocrine cell expression in the pig.

BACKGROUND: Enteroendocrine cell-derived peptides modulate postresectional small bowel adaptation, which may be attenuated by transplantation. We investigated whether autotransplantation modulates the number and distribution of ileal enteroendocrine cells in pigs with proximal small bowel resection. MATERIALS AND METHODS: Fifteen pigs were assigned into either small intestinal transection or 75% proximal small intestinal resection with or without autotransplantation of the remaining ileum. After 14 weeks the number and subtype distribution of enteroendocrine cells, crypt cell proliferation, and mucosal histology were analyzed from the proximal and distal ends of the remaining ileum. RESULTS: When compared to resected controls, autotransplantation of the ileum decreased the absolute (P < 0.05 in proximal ileum) and proportional (P < 0.05 in distal ileum) crypt enteroendocrine cell number. In addition, autotransplantation reduced somatostatin and glicentin expressing cell counts and abolished the proximodistal gradient of the enteroendocrine cell number. When compared to transected controls, villus height, crypt depth, number of proliferating crypt cells, and crypt cell proliferation index increased after the proximal resection (P < 0.05 in all except in crypt depth and proliferation index of the distal ileum) but remained virtually unchanged after autotransplantation of the ileal remnant. CONCLUSIONS: Autotransplantation decreases the crypt enteroendocrine cell number and alters their proximodistal and subtype distribution in the remaining ileum in pigs with proximal small bowel resection. These alterations are associated with attenuated adaptive response of the autotransplanted ileum.

Endothelial sulfated sialyl Lewis x glycans, putative L-selectin ligands, are preferentially expressed in bronchial asthma but not in other chronic inflammatory lung diseases.

Lymphocyte infiltrate is a hallmark of inflammatory responses. We have previously shown that de novo-induced endothelial sialyl Lewis x (sLex) expression guides lymphocytes in an L-selectin-dependent manner to sites of acute organ transplant rejections. In this research, we have analyzed five groups of chronic lung inflammations to determine the presence of properly glycosylated, i.e., sulfated, sLex-decorated, L-selectin ligands. Two anti-sLex (2F3 and HECA-452) and one anti-6- and/or 6'-sulfated and/or 6,6'-bisulfated (MECA-79) monoclonal antibodies (mAbs) were used. The control lung specimens did not express L-selectin ligands on endothelium. In contrast, the endothelial staining intensity and the number of positive peribronchial venules and capillaries with mAbs 2F3, HECA-452, and MECA-79 were significantly greater in bronchial biopsies from patients with asthma compared with normal specimens (P<0.003). However, no significant increase of peribronchial endothelial reactivity with these antibodies was observed in adult respiratory distress syndrome, chronic bronchitis, fibrosing alveolitis, and granulomatous inflammation compared with controls. These data suggest that sulfated sLex glycans, acting putatively as ligands for L-selectin, could be instrumental in lymphocyte extravasation into human peribronchial lung tissue during asthma, but not so important in several other inflammatory lung diseases.

MR imaging of overuse injuries of the Achilles tendon.

OBJECTIVE: This study was conducted to illustrate and classify the abnormalities found on high-resolution MR imaging of symptomatic Achilles tendons in athletic adult patients. SUBJECTS AND METHODS: One hundred patients with 118 painful Achilles tendons were imaged with a 1.5-T magnet. The tendon, peritendinous tissues, tendon insertion, and musculotendinous junction were examined on MR imaging. Twenty-eight patients underwent surgery, and histopathologic samples were taken in 13. Long-term follow-up was performed, on average, 3.4 years after MR imaging. RESULTS: Of 118 painful Achilles tendons, abnormalities were detected in 111. These were in the tendon (n = 90), surrounding structures, or both. Fifty-four tendons had a focal area of increased intratendinous signal, best detected on axial high-resolution T1-weighted gradient-echo MR imaging. Histopathology confirmed abnormal tendon structure. Of the 21 surgically proven foci of tendinosis, 20 were revealed on MR imaging. At the level of the insertion, changes were found in the tendon in 15%, in the retrocalcaneal bursa in 19%, and in the calcaneal bone marrow in 8% of the studies. Abnormalities in peritendinous soft tissues were detected in 67%. More than one type of abnormality was found in 64% of the studies. CONCLUSION: Lesions in the Achilles tendon and in the peritendinous structures can have similar clinical presentation. MR imaging detects and characterizes these changes. A more specific diagnosis and prognosis can be made with the use of MR imaging than with clinical examination alone.

Endothelial and epithelial expression of sialyl Lewis(x) in squamous carcinoma of the tongue.

In this retrospective study we describe the immunohistochemical expression pattern of sLe(x) epitopes in endothelial and epithelial cells of 59 squamous carcinomas of the tongue, and relate this to the relative survival rates of the patients. Endothelial sLe(x) expression was significantly elevated in malignant lesions compared to normal tissues, but did not have any prognostic value for the relative survival rate. In contrast, epithelial sLe(x) expression was decreased in carcinomas compared to normal tongue. The patients whose carcinoma showed only moderate epithelial HECA-452 reactivity had a significantly better relative survival rate than the patients with tumor specimens with neglible or very high HECA-452 reactivity. The epithelial staining with the two other anti-sLe(x) antibodies (CSLEX-1 and 2F3) did not correlate with the survival rates of tongue carcinoma patients.