Expression of Selected miRNAs in Normal and Cancer-Associated Fibroblasts and in BxPc3 and MIA PaCa-2 Cell Lines of Pancreatic Ductal Adenocarcinoma.

Therapy for pancreatic ductal adenocarcinoma remains challenging, and the chances of a complete cure are very limited. As in other types of cancer, the expression and role of miRNAs in controlling the biological properties of this type of tumor have been extensively studied. A better insight into miRNA biology seems critical to refining diagnostics and improving their therapeutic potential. In this study, we focused on the expression of miR-21, -96, -196a, -210, and -217 in normal fibroblasts, cancer-associated fibroblasts prepared from a ductal adenocarcinoma of the pancreas, and pancreatic carcinoma cell lines. We compared these data with miRNAs in homogenates of paraffin-embedded sections from normal pancreatic tissues. In cancer-associated fibroblasts and cancer cell lines, miRNAs differed significantly from the normal tissue. In detail, miR-21 and -210 were significantly upregulated, while miR-217 was downregulated. Similar transcription profiles were earlier reported in cancer-associated fibroblasts exposed to hypoxia. However, the cells in our study were cultured under normoxic conditions. We also noted a relation to IL-6 production. In conclusion, cultured cancer-associated fibroblasts and carcinoma cells reflect miR-21 and -210 expression similarly to the cancer tissue samples harvested from the patients.

Diagnostic accuracy of DSA in carotid artery stenosis: a comparison between stenosis measured on carotid endarterectomy specimens and DSA in 644 cases.

BACKGROUND AND PURPOSE: DSA (digital subtraction angiography) is the gold standard for measuring carotid artery stenosis (CS). Yet, the correlation between DSA and stenosis is not well documented. MATERIAL AND METHODS: We compared CS as measured by DSA to carotid artery specimens obtained from carotid endarterectomy surgery. Patients were divided into three groups according to NASCET criteria (North American Symptomatic Carotid Endarterectomy Trial): stenosis of 30-49% (mild), stenosis of 50-69% (moderate), and stenosis of 70-99% (severe). RESULTS: This prospective cohort study involved 644 patients. The mean stenosis in the mild stenosis group (n = 128 patients) was 54% ECST (European Carotid Surgery Trial), 40% NASCET, and 72% ESs (endarterectomy specimens). The mean absolute difference between ECST and NASCET was 14%. The mean stenosis in the moderate stenosis group (n = 347 patients) was 66% ECST, 60% NASCET, and 77% ES. The mean absolute difference between ECST and NASCET was 6%. The mean stenosis in the severe group (n = 169 patients) was 80% ECST, 76% NASCET, and 79% ES. No significant correlation coefficients were found between DSA and ES methods. In the mild group, the CC was 0.16 (ESCT) and 0.13 (NASCET); in the moderate group, the CC was 0.05 (ESCT) and 0.01 (NASCET); and in the severe group, the CC was 0.23 (ESCT) and 0.10 (NASCET). For all groups combined, CC was 0.22 for the ECST and 0.20 for the NASCET method. CONCLUSION: The relationship between DSA and ES methods to measure CS is almost random. This lack of a relationship between the DSA and ES techniques questions the validity of current DSA-based guidelines.

Histological Analysis of Carotid Plaques: The Predictors of Stroke Risk.

OBJECTIVES: The morphology and histological structure of the atherosclerotic plaque seem critical to its stability. Our study aimed to identify the epidemiological, morphological and histological parameters associated with stable and unstable plaques. MATERIALS AND METHODS: The study included 280 plaques harvested from 269 consecutive patients. Epidemiological and demographic data were recorded as well as the histological features of plaque, i.e. calcifications, myxoid changes, bleeding into plaque, presence of thrombus, inflammation, macrophages, giant cell reaction, siderophages, neovascularization and plaque ossification. All specimens were classified according to the American Heart Association (AHA). RESULTS: Monofactorial analysis identified three significant histological predictors for the symptomatic plaque: a plaque with a large necrotic core (odds ratio, OR=2.0, p = 0.03), thrombosis (OR=3.7, p = 0.01) and the formation of foamy macrophages (OR=2.0, p = 0.01). Multifactorial logistic regression revealed that the presence of foamy macrophages (OR=1.9, p = 0.03) and thrombosis (OR=3.5, p = 0.02) were significant predictors of symptomatic stenosis. Symptomatic plaques were significantly more frequently classified as AHA type VI than AHA type IV-V compared to asymptomatic ones (OR=1.8, p = 0.03). CONCLUSIONS: Our study shows that no single histological feature, except for the presence of foamy macrophages and thrombosis on the plaque, is predictive of plaque instability. Rather, a complex plaque structure (AHA type VI) is predictive of plaque instability. Our findings should be kept in mind during the assessment of non-invasive imaging and stroke risk estimation.

Changes in histopathological classification of neuroendocrine tumors in 5th edition of WHO classification of gastrointestinal tract tumors (2019).

In a brief review is presented a summary of news in the classification of neuroendocrine neoplasms of the digestive system, as they were introduced in the 5th edition of the WHO Classification of Digestive System Tumors published in summer 2019.

[Isolated infectious endocarditis of the pulmonary valve: a case report]. / Izolovaná infekcní endokarditida pulmonální chlopne: kazuistika.

Isolated infectious endocarditis of the pulmonary valve is a rare condition and represents 1,5-2% of all cases of infectious endocarditis. We present a case of a 37year-old woman without any relevant medical history. The woman was hospitalized with hallmarks of severe sepsis and bilateral pneumonia; she died several hours after admission with progressive multiorgan failure and disseminated intravascular coagulopathy. Microbiologic examination approved Staphylococcus aureus as the etiological agent. The autopsy showed isolated endocarditis of the pulmonary valve, without any known predisposing factor. Literary data refer single cases or small groups of patients with isolated pulmonary infectious endocarditis. The clinical suspicion of this rare disease in differential diagnosis of febrile conditions is an essential factor in prognosis of afflicted persons. The crucial diagnostic methods for infectious endocarditis are echocardiography and CT examination.

Detection of carotid artery stenosis using histological specimens: a comparison of CT angiography, magnetic resonance angiography, digital subtraction angiography and Doppler ultrasonography.

BACKGROUND: Carotid endarterectomy (CEA) is accepted as a primary modality to treat carotid stenosis. The accuracy of measuring carotid stenosis is important for indication of the CEA procedure. Different diagnostic tools have been developed and used in the past 2 decades for the diagnosis of carotid stenosis. Only a few studies, however, have focused on the comparison of different diagnostic tools to histological findings of carotid plaque. METHOD: Patients with internal carotid artery (ICA) stenosis were investigated primarily by computed tomography angiography (CTA). Digital subtraction angiography (DSA), Doppler ultrasonography (DUS) and magnetic resonance angiography (MRA) were performed as well. Atherosclerotic plaque specimens were transversally cut into smaller segments and histologically processed. The slides were scanned and specimens showing maximal stenosis were determined; the minimal diameter and the diameter of the whole plaque were measured. High quality histological specimen and histological measurement was considered to be the prerequisite for inclusion into the analysis. The preoperative findings were compared with histological measurement. CTA and histological measurements were obtained from 152 patients. DSA measurements were available in 138 of these cases, MRA in 107 and DUS in 88. A comparison between preoperative and histological findings was performed. In addition, correlation coefficients were computed and tested. RESULTS: A significant correlation was found for each of the diagnostic procedures. The strongest correlation coefficient and the best allocation of stenosis into clinical significant groups (<50 %, 50-69 %, ≥70 %) was observed for CTA. Mean differences in the whole cohort between preoperative and histological measurements were as follows: CTA underestimated histological measurement by 2.4 % (based on European Carotid Surgery Trial [ECST] methodology) and 11.9 % (based on North American Symptomatic Carotid Endarterectomy Trial [NASCET] methodology). DSA underestimated the histological measurement by 7 % (ECST) and 12.2 % (NASCET). MRA overestimated the histological measurement by 2.6 % (ECST) and underestimated by 0.6 % (NASCET). DUS overestimated the stenosis by 1.8 %. CONCLUSIONS: CTA yields the best accuracy in detection of carotid stenosis, provided that all axial slices of the stenosis are checked and carefully analysed. DSA underestimates moderate and mild ICA stenosis, whereas DUS overestimates high-grade ICA stenosis. For MRA, a relatively low correlation coefficient was observed with histological findings. We conclude that CTA-ecst technique is the most reliable technique for carotid stenosis measurement.

Human papillomavirus DNA prevalence and type distribution in anal carcinomas worldwide.

Knowledge about human papillomaviruses (HPV) types involved in anal cancers in some world regions is scanty. Here, we describe the HPV DNA prevalence and type distribution in a series of invasive anal cancers and anal intraepithelial neoplasias (AIN) grades 2/3 from 24 countries. We analyzed 43 AIN 2/3 cases and 496 anal cancers diagnosed from 1986 to 2011. After histopathological evaluation of formalin-fixed paraffin-embedded samples, HPV DNA detection and genotyping was performed using SPF-10/DEIA/LiPA25 system (version 1). A subset of 116 cancers was further tested for p16(INK4a) expression, a cellular surrogate marker for HPV-associated transformation. Prevalence ratios were estimated using multivariate Poisson regression with robust variance in the anal cancer data set. HPV DNA was detected in 88.3% of anal cancers (95% confidence interval [CI]: 85.1-91.0%) and in 95.3% of AIN 2/3 (95% CI: 84.2-99.4%). Among cancers, the highest prevalence was observed in warty-basaloid subtype of squamous cell carcinomas, in younger patients and in North American geographical region. There were no statistically significant differences in prevalence by gender. HPV16 was the most frequent HPV type detected in both cancers (80.7%) and AIN 2/3 lesions (75.4%). HPV18 was the second most common type in invasive cancers (3.6%). p16(INK4a) overexpression was found in 95% of HPV DNA-positive anal cancers. In view of the results of HPV DNA and high proportion of p16(INK4a) overexpression, infection by HPV is most likely to be a necessary cause for anal cancers in both men and women. The large contribution of HPV16 reinforces the potential impact of HPV vaccines in the prevention of these lesions.

Small nucleolar RNA U91 is a new internal control for accurate microRNAs quantification in pancreatic cancer.

BACKGROUND: RT-qPCR quantification of miRNAs expression may play an essential role in pancreatic ductal adenocarcinoma (PDAC) diagnostics. RT-qPCR-based experiments require endogenous controls for the result normalization and reliability. However, expression instability of reference genes in tumors may introduce bias when determining miRNA levels. METHODS: We investigated expression of 6 miRNAs, isolated from FFPE samples of pancreatic adenocarcinomas. Four internal controls were utilized for RT-qPCR result normalization: artificial miR-39 from C. elegans, U6 snRNA, miR-16 and snoRNA U91. RESULTS: We found miR-21, miR-155 or miR-217 expression values in tumors may differ up to several times, depending on selected internal controls. Moreover, different internal controls can produce controversial results for miR-96, miR-148a or miR-196a quantification. Also, expression of our endogenous controls varied significantly in tumors. U6 demonstrated variation from -1.03 to 8.12-fold, miR-16 from -2.94 up to 7.38-fold and the U91 from -3.05 to 4.36-fold respectively. On the other hand, the most stable gene, determined by NormFinder algorithm, was U91. Each miRNA normalized relatively to the spike or U91, demonstrated similar expression values. Thus, statistically significant and insignificant differences between tumors and normal tissues for miRNAs were equal for the spike and the U91. Also, the differences between the spike and U91 were statistically insignificant for all of miRs except miR-217. Among three endogenous controls, U91 had the lowest average expression values and standard deviation in cancer tissues. CONCLUSIONS: We recommend U91 as a new normalizer for miRNA quantification in PDACs.

[Factors causing damage and destruction of beta-cells of the islets of Langerhans in the pancreas]. / Faktory vedoucí k poskození a destrukci B-bunek Langerhansových ostruvku pankreatu.

Insulin secretion in patients with manifested diabetes mellitus tends to disappear months to decades after the diagnosis, which is a clear sign of a gradual loss of pancreatic islet beta-cells. In our sample of 30 type 2 diabetic patients, whose disease manifested between 30 and 45 years of age, about a half have retained or even increased insulin secretion 30 years later, while the other half exhibit a much diminished or lost insulin secretion. Factors that can damage or destroy beta-cells can be divided into the following groups: Metabolic factors: hyperglycemia and glucotoxicity, lipotoxicity, hypoxia, reactive oxygen species; Pharmacological factors: antimicrobial medication pentamidine, SSRI antidepressants; Factors related to impaired insulin secretion: MODY type diabetes; Environmental toxic factors: rat poison Vacor, streptozotocin, polychlorinated and polybrominated hydrocarbons; Disorders of the exocrine pancreas: tumor infiltration, fibrous infiltration, chronic pancreatitis, cystic fibrosis; Infections, inflammation, autoimmunity, viral factors: Coxsackie viruses, H1N1 influenza, enteroviruses. We are currently working on finding other factors leading to beta-cell damage, studying their effect on apoptosis and necrosis and looking for possible protective factors to prevent this damage. We our increasing knowledge about the mechanisms of beta-cell damage and destruction we come ever closer to suggest measures for their prevention. In this review we offer a brief and simplified summary of some of the findings related to this area.Key words: pancreatic islet beta-cells of Langerhans - factors damaging or destroying beta-cells - insulin secretion.

Metastatic Adenoid Cystic Carcinoma at the Base of the Tongue and Duplicate Breast Cancer Diagnosed During Restaging.

BACKGROUND/AIM: Adenoid cystic carcinoma (AdCC) is a rare malignant tumor that primarily affects the salivary glands but can also occur in other organs. Low incidence and unpredictable clinical behavior make AdCC one of the most difficult head and neck tumors to treat. CASE REPORT: We present the case of a 54-year-old woman with AdCC localized at the base of the tongue, following radical surgical and oncological therapy. Due to advances in palliative oncological treatment, there is a more than five-year survival period before the progression of metastatic disease. Considering the rare occurrence of this disease, a literature search was also conducted, and therapy options are discussed. Ensuring a sufficient extent of the surgical procedure is still a challenge, and most specialists agree that subsequent postoperative radiotherapy reduces the risk of local recurrence. The effective dose of radiotherapy to the area of the primary tumor and lymph nodes is not clearly defined. CONCLUSION: The distinct biological behavior of AdCC results in varying sensitivity to chemotherapy or radiotherapy compared to treatments commonly used for head and neck squamous cell carcinomas. Treatment recommendations for these rarer tumors are based mainly on case reports and small clinical trials. The acquired therapeutic experience can contribute to prolonging the survival period of patients and improving their prognosis and quality of life.

Plasma lipidome differences in patients with and without significant carotid plaque.

BACKGROUND: Atherosclerosis is a major cause of ischemic stroke, and early detection of advanced atherosclerosis in the carotid artery is important for reducing morbidity and mortality. What is even more important is not only detection of atherosclerosis but early determination whether the patients are at high risk of an event with adverse effects as the size of the plaque does not necessarily reflect its potential to trigger such events. AIM: We studied whether plasma lipidomics profile can be used as a diagnostic tool for stratification of stable or unstable plaques without the need of removing the carotid plaque. METHODS: This study used liquid chromatography high-resolution tandem mass spectrometry lipidomics to characterize lipid profiles in patients' plasma and found that patients with significant and complicated (vulnerable) atherosclerotic plaque had distinct lipid profiles compared to those with insignificant plaques. RESULTS: The lipid classes that were most predictive of vulnerable plaque were lysophosphoethanolamines, fatty acyl esters of hydroxy fatty acids, free fatty acids, plasmalogens, and triacylglycerols. Most of these compounds were found decreased in plasma of patients with unstable plaques which enabled sufficient performance of a statistical model used for patient stratification. CONCLUSIONS: Plasma lipidomes measured by liquid chromatography-mass spectrometry show differences in patients with stable and unstable carotid plaques, therefore these compounds could potentially be used as biomarkers for unstable plaque in future clinical diagnosis.

Expression of Selected miRNAs in Undifferentiated Carcinoma with Osteoclast-like Giant Cells (UCOGC) of the Pancreas: Comparison with Poorly Differentiated Pancreatic Ductal Adenocarcinoma.

Undifferentiated carcinoma with osteoclast-like giant cells (UCOGC) of the pancreas represents a rare subtype of pancreatic ductal adenocarcinoma (PDAC). Despite a distinct morphology and specific clinical behavior, UCOGCs exhibit unexpected similarities in regard to DNA mutational profiles with conventional PDAC. Treating pancreatic ductal adenocarcinoma is particularly challenging, with limited prospects for cure. As with many other malignant neoplasms, the exploration of microRNAs (miRNAs, miRs) in regulating the biological characteristics of pancreatic cancer is undergoing extensive investigation to enhance tumor diagnostics and unveil the therapeutic possibilities. Herein, we evaluated the expression of miR-21, -96, -148a, -155, -196a, -210, and -217 in UCOGCs and poorly differentiated (grade 3, G3) PDACs. The expression of miR-21, miR-155, and miR-210 in both UCOGCs and G3 PDACs was significantly upregulated compared to the levels in normal tissue, while the levels of miR-148a and miR-217 were downregulated. We did not find any significant differences between cancerous and normal tissues for the expression of miR-96 and miR-196a in G3 PDACs, whereas miR-196a was slightly, but significantly, downregulated in UCOGCs. On the other hand, we have not observed significant differences in the expression of the majority of miRNAs between UCOGC and G3 PDAC, with the exception of miR-155. UCOGC samples demonstrated lower mean levels of miR-155 in comparison with those in G3 PDACs.

[Microscopic colitis]. / Mikroskopické zánety tlustého streva.

Microscopic colitis is characterized by chronic or intermittent watery diarrhoea. Microscopic colitis is a common cause of chronic diarrhoea in predominantly older adults. The underlying mechanism in the pathogenesis of microscopic colitis remains unspecified. Microscopic colitis including colitis collagenous, lymphocytic, microscopic colitis with incomplete findings, minimal change colitis, eosinophilic colitis, Brainerd´s diarrhoea, graft-versus-host disease, mastocytic enterocolitis and postinfectious irritable bowel syndrome. Careful consideration of the clinical features and colonic mucosal biopsies usually lead to correct diagnosis. Treatments of microscopic colitis were based primarily on case reports and personal experience. Many medications have been proposed that either offer symptomatic relief (loperamide, cholestyramine) or had anti-inflammatory or immunosuppressive properties (aminosalicylates, steroids, adalimumab, azathioprine).

Nanofiber Wound Dressing Materials-A Comparative Study of Wound Healing on a Porcine Model.

BACKGROUND: Nanofiber wound dressings remain the domain of in vitro studies. The purpose of our study was to verify the benefits of chitosan (CTS) and polylactide (PLA)-based nanofiber wound dressings on a porcine model of a naturally contaminated standardized wound and compare them with the conventional dressings, i.e., gauze and Inadine. MATERIAL AND METHODS: The study group included 32 pigs randomized into four homogeneous groups according to the wound dressing type. Standardized wounds were created on their backs, and wound dressings were regularly changed. We evaluated difficulty of handling individual dressing materials and macroscopic appearance of the wounds. Wound swabs were taken for bacteriological examination. Blood samples were obtained to determine blood count values and serum levels of acute phase proteins (serum amyloid A, C-reactive protein, and haptoglobin). The crucial point of the study was histological analysis. Microscopic evaluation was focused on the defect depth and tissue reactions, including formation of the fibrin exudate with neutrophil granulocytes, the layer of granulation and cellular connective tissue, and the reepithelialization. Statistical analysis was performed by using SPSS software. The analysis was based on the Kruskal-Wallis H test and Mann-Whitney U test followed by Bonferroni correction. Significance was set at P < .05. RESULTS: Macroscopic examination did not show any difference in wound healing among the groups. However, evaluation of histological findings demonstrated that PLA-based nanofiber dressing accelerated the proliferative (P = .025) and reepithelialization (P < .001) healing phases, while chitosan-based nanofiber dressing potentiated and accelerated the inflammatory phase (P = .006). No statistically significant changes were observed in the blood count or acute inflammatory phase proteins during the trial. Different dynamics were noted in serum amyloid A values in the group treated with PLA-based nanofiber dressing (P = .006). CONCLUSION: Based on the microscopic examination, we have documented a positive effect of nanofiber wound dressings on acceleration of individual phases of the healing process. Nanofiber wound dressings have a potential to become in future part of the common wound care practice.

Oxidative Stress as a Reliable Biomarker of Carotid Plaque Instability: A Pilot Study.

Background: Predicting stroke risk in patients with carotid artery stenosis (CS) remains challenging. Circulating biomarkers seem to provide improvements with respect to risk stratification. Methods: Study patients who underwent carotid endarterectomy were categorized into four groups according to symptomatology and compared as follows: symptomatic with asymptomatic patients; and asymptomatic patients including amaurosis fugax (AF) (asymptomatic + AF group) with patients with a transient ischemic attack (TIA) or brain stroke (BS) (hemispheric brain stroke group). Carotid specimens were histologically analyzed and classified based on the American Heart Classification (AHA) standard. As a marker of OS, the plasma levels of malondialdehyde (MDA) were measured. Comparisons of MDA plasma levels between groups were analyzed. Results: In total, 35 patients were included in the study. There were 22 (63%) patients in the asymptomatic group and 13 (37%) in the symptomatic group. Atheromatous plaque (p = 0.03) and old hemorrhage (p = 0.05), fibrous plaque (p = 0.04), myxoid changes (p = 0.02), plaques without hemorrhage (p = 0.04), significant neovascularization (p = 0.04) and AHA classification (p = 0.006) had significant correlations with clinical presentation. There were 26 (74%) patients in the asymptomatic group and 9 (26%) in the hemispheric brain stroke group. Atheromatous plaque (p = 0.02), old hemorrhage (p = 0.05) and plaques without neovascularization (p = 0.02), fibrous plaque (p = 0.03), plaques without hemorrhage (p = 0.02) and AHA classification (p = 0.01) had significant correlations with clinical presentation. There was no significant difference between symptomatic and asymptomatic groups with respect to MDA plasma levels (p = 0.232). A significant difference was observed when MDA plasma levels were compared to asymptomatic + AF and the hemispheric stroke group (p = 0.002). Conclusions: MDA plasma level correlates with the risk of hemispheric stroke (TIA or BS) and is a reliable marker of plaque vulnerability in carotid artery stenosis.

Human keratinocyte growth and differentiation on acellular porcine dermal matrix in relation to wound healing potential.

A number of implantable biomaterials derived from animal tissues are now used in modern surgery. Xe-Derma is a dry, sterile, acellular porcine dermis. It has a remarkable healing effect on burns and other wounds. Our hypothesis was that the natural biological structure of Xe-Derma plays an important role in keratinocyte proliferation and formation of epidermal architecture in vitro as well as in vivo. The bioactivity of Xe-Derma was studied by a cell culture assay. We analyzed growth and differentiation of human keratinocytes cultured in vitro on Xe-Derma, and we compared the results with formation of neoepidermis in the deep dermal wounds treated with Xe-Derma. Keratinocytes cultured on Xe-Derma submerged in the culture medium achieved confluence in 7-10 days. After lifting the cultures to the air-liquid interface, the keratinocytes were stratified and differentiated within one week, forming an epidermis with basal, spinous, granular, and stratum corneum layers. Immunohistochemical detection of high-molecular weight cytokeratins (HMW CKs), CD29, p63, and involucrin confirmed the similarity of organization and differentiation of the cultured epidermal cells to the normal epidermis. The results suggest that the firm natural structure of Xe-Derma stimulates proliferation and differentiation of human primary keratinocytes and by this way improves wound healing.

Senescence-Associated miRNAs and Their Role in Pancreatic Cancer.

Replicative senescence is irreversible cell proliferation arrest for somatic cells which can be circumvented in cancers. Cellular senescence is a process, which may play two opposite roles. On the one hand, this is a natural protection of somatic cells against unlimited proliferation and malignant transformation. On the other hand, cellular secretion caused by senescence can stimulate inflammation and proliferation of adjacent cells that may promote malignancy. The main genes controlling the senescence pathways are also well known as tumor suppressors. Almost 140 genes regulate both cellular senescence and cancer pathways. About two thirds of these genes (64%) are regulated by microRNAs. Senescence-associated miRNAs can stimulate cancer progression or act as tumor suppressors. Here we review the role playing by senescence-associated miRNAs in development, diagnostics and treatment of pancreatic cancer.

Coating Ti6Al4V implants with nanocrystalline diamond functionalized with BMP-7 promotes extracellular matrix mineralization in vitro and faster osseointegration in vivo.

The present study investigates the effect of an oxidized nanocrystalline diamond (O-NCD) coating functionalized with bone morphogenetic protein 7 (BMP-7) on human osteoblast maturation and extracellular matrix mineralization in vitro and on new bone formation in vivo. The chemical structure and the morphology of the NCD coating and the adhesion, thickness and morphology of the superimposed BMP-7 layer have also been assessed. The material analysis proved synthesis of a conformal diamond coating with a fine nanostructured morphology on the Ti6Al4V samples. The homogeneous nanostructured layer of BMP-7 on the NCD coating created by a physisorption method was confirmed by AFM. The osteogenic maturation of hFOB 1.19 cells in vitro was only slightly enhanced by the O-NCD coating alone without any increase in the mineralization of the matrix. Functionalization of the coating with BMP-7 resulted in more pronounced cell osteogenic maturation and increased extracellular matrix mineralization. Similar results were obtained in vivo from micro-CT and histological analyses of rabbit distal femurs with screws implanted for 4 or 12 weeks. While the O-NCD-coated implants alone promoted greater thickness of newly-formed bone in direct contact with the implant surface than the bare material, a further increase was induced by BMP-7. It can be therefore concluded that O-NCD coating functionalized with BMP-7 is a promising surface modification of metallic bone implants in order to improve their osseointegration.

Heat Shock Proteins 27, 70, and 110: Expression and Prognostic Significance in Colorectal Cancer.

Heat shock proteins (HSPs) are evolutionarily conserved chaperones occurring in virtually all living organisms playing a key role in the maintenance of cellular homeostasis. They are constitutively expressed to prevent and repair protein damage following various physiological and environmental stressors. HSPs are overexpressed in various types of cancers to provide cytoprotective function, and they have been described to influence prognosis and response to therapy. Moreover, they have been used as a tumor marker in blood serum biochemistry and they represent a potentially promising therapeutic target. To clarify prognostic significance of two canonical HSPs (27 and 70) and less known HSP110 (previously known as HSP105) in colorectal carcinoma (CRC), we retrospectively performed HSP immunohistochemistry on tissue microarrays from formalin-fixed paraffin-embedded tumor tissue from 297 patients with known follow-up. Survival analysis (univariate Kaplan-Meier analysis with the log-rank test and multivariate Cox regression) revealed significantly shorter overall survival (OS, mean 5.54 vs. 7.07, p = 0.033) and borderline insignificantly shorter cancer specific survival (CSS, mean 6.3 vs. 7.87 years, p = 0.066) in patients with HSP70+ tumors. In the case of HSP27+ tumors, there was an insignificantly shorter OS (mean 6.36 vs. 7.13 years, p = 0.2) and CSS (mean 7.17 vs. 7.95 years, p = 0.2). HSP110 showed no significant impact on survival. Using Pearson's chi-squared test, there was a significant association of HSP27 and HSP70 expression with advanced cancer stage. HSP27+ tumors were more frequently mismatch-repair proficient and vice versa (p = 0.014), and they occurred more often in female patients and vice versa (p = 0.015). There was an enrichment of left sided tumors with HSP110+ compared to the right sided (p = 0.022). In multivariate Cox regression adjusted on the UICC stage, grade and right/left side; both HSPs 27 and 70 were not independent survival predictors (p = 0.616 & p = 0.586). In multivariate analysis, only advanced UICC stage (p = 0) and right sided localization (p = 0.04) were independent predictors of worse CSS. In conclusion, from all three HSPs examined in our study, only HSP70 expression worsened CRC prognosis, although stage-dependent. The contribution of this article may be seen as a large survival analysis of HSPs 27 and 70 and the largest analysis of HSP110 described in CRC.

Expression profiles of proliferative and antiapoptotic genes in sporadic and colitis-related mouse colon cancer models.

Elevated levels of survivin, telomerase catalytic subunit (TERT), integrin-linked kinase (ILK), cyclooxygenase 2 (COX-2), inducible nitric oxide synthase (iNOS) and the regulatory factors c-MYB and Tcf-4 are often found in human cancers including colorectal cancer (CRC) and have been implicated in the development and progression of tumorigenesis. The aim of this study was to determine the expression of these genes in mouse models of sporadic and colitis-associated CRC. To address these issues, we used qRT-PCR approach to determine changes in gene expression patterns of neoplastic cells (high-grade dysplasia/intramucosal carcinoma) and surrounding normal epithelial cells in A/J and ICR mouse strains using laser microdissection. Both strains were injected with azoxymethane and ICR mice were also given drinking water that contained 2% dextran sodium sulphate. In both sporadic (A/J mice) and colitis-associated (ICR mice) models of CRC, the levels of TERT mRNA, COX-2 mRNA and Tcf-4 mRNA were higher in neoplastic cells than in surrounding normal epithelial cells. In contrast, survivin mRNA was upregulated only in neoplastic cells from A/J mice and ILK mRNA was upregulated only in neoplastic cells from ICR mice. However, the expression of iNOS mRNA was similar in normal and neoplastic cells in both models and c-MYB mRNA was actually downregulated in neoplastic cells compared with normal cells in both models. These findings suggest that the genetic background and/or the molecular mechanisms of tumorigenesis associated with genotoxic insults and colonic inflammation influence the gene expression of mTERT, COX-2, Tcf-4, c-MYB, ILK and survivin in colon epithelial neoplasia.