Systemically administered zoledronic acid activates locally implanted synthetic hydroxyapatite particles enhancing peri-implant bone formation: A regenerative medicine approach to improve fracture fixation.

Fracture fixation in an ageing population is challenging and fixation failure increases mortality and societal costs. We report a novel fracture fixation treatment by applying a hydroxyapatite (HA) based biomaterial at the bone-implant interface and biologically activating the biomaterial by systemic administration of a bisphosphonate (zoledronic acid, ZA). We first used an animal model of implant integration and applied a calcium sulphate (CaS)/HA biomaterial around a metallic screw in the tibia of osteoporotic rats. Using systemic ZA administration at 2-weeks post-surgery, we demonstrated that the implant surrounded by HA particles showed significantly higher peri­implant bone formation compared to the unaugmented implants at 6-weeks. We then evaluated the optimal timing (day 1, 3, 7 and 14) of ZA administration to achieve a robust effect on peri­implant bone formation. Using fluorescent ZA, we demonstrated that the uptake of ZA in the CaS/HA material was the highest at 3- and 7-days post-implantation and the uptake kinetics had a profound effect on the eventual peri­implant bone formation. We furthered our concept in a feasibility study on trochanteric fracture patients randomized to either CaS/HA augmentation or no augmentation followed by systemic ZA treatment. Radiographically, the CaS/HA group showed signs of increased peri­implant bone formation compared with the controls. Finally, apart from HA, we demonstrated that the concept of biologically activating a ceramic material by ZA could also be applied to ß-tricalcium phosphate. This novel approach for fracture treatment that enhances immediate and long-term fracture fixation in osteoporotic bone could potentially reduce reoperations, morbidity and mortality. STATEMENT OF SIGNIFICANCE: • Fracture fixation in an ageing population is challenging. Biomaterial-based augmentation of fracture fixation devices has been attempted but lack of satisfactory biological response limits their widespread use. • We report the biological activation of locally implanted microparticulate hydroxyapatite (HA) particles placed around an implant by systemic administration of the bisphosphonate zoledronic acid (ZA). The biological activation of HA by ZA enhances peri­implant bone formation. •Timing of ZA administration after HA implantation is critical for optimal ZA uptake and consequently determines the extent of peri­implant bone formation. • We translate the developed concept from small animal models of implant integration to a proof-of-concept clinical study on osteoporotic trochanteric fracture patients. • ZA based biological activation can also be applied to other calcium phosphate biomaterials.

Advanced image analysis-based evaluation of protein antibody microarray chemiluminescence signal improves glioma type identification by blood serum proteins concentrations.

BACKGROUND AND OBJECTIVE: Gliomas are the most common brain tumors usually classified as benign low-grade or aggressive high-grade glioma. One of the promising possibilities of glioma diagnostics and tumor type identification could be based on concentration measurements of glioma secreted proteins in blood. However, several published approaches of quantitative proteomic analysis emphasize limits of one single protein to be used as biomarker of these types of tumors. Simultaneous multi-protein concentrations analysis giving antibody array-based methods suffer from poor measurement accuracy due to technical limitations of imaging systems. METHODS: We applied Principal Component Analysis (PCA) for series of repeated antibody array chemiluminescence images to extract the component representing relative values of protein concentrations, free from zero-mean noise and uneven background illumination - main factors corrupting evaluation result. RESULTS: The proposed method increased accuracy of protein concentration estimates at least 2-fold. Decision tree classifier applied to the relative concentration values of three proteins TIMP-1, PAI-1 and NCAM-1 estimated by proposed image analysis method effectively distinguished between low-grade glioma, high-grade glioma and healthy control subjects showing validation accuracy of 74.9% with the highest positive predictive value of 81.2% for high grade glioma and 57.1% for low grade glioma cases. CONCLUSIONS: PCA-based image processing could be applied in protein antibody microarray and other multitarget detection/evaluation investigations to increase estimation accuracy.

Serum protein triplet TGF-ß1, TIMP-1, and YKL-40 serve as diagnostic and prognostic profile for astrocytoma.

Astrocytoma is the most common glial tumour of the CNS. The most malignant form is grade IV Astrocytoma, also called Glioblastoma. Due to its heterogeneity, aggressiveness and lethal nature scientists are trying to find less invasive methods for early prediction of tumour onset, recurrence, response to therapy and patients' survival. Here, applying decision tree classification algorithm we performed astrocytoma specific protein profile analysis on serum proteins TIMP-1, active and latent form of TGF-ß1, IP-10, ANGPT-1, OPN, and YKL-40 using enzyme-linked immunosorbent detection assay (ELISA). Results have demonstrated that astrocytoma specific profile consisted of three proteins-active form of TGF-ß1, TIMP-1 and YKL-40 and was able to correctly classify 78.0% (103/132) of sample and 83.3% (60/72) of astrocytoma sample. Calculating decision tree algorithm associated with astrocytoma patient survival, prediction model reached an accuracy of 83.3% (60/72). All together these results indicate that glioma detection and prediction from patient serum using glioma associated proteins and applying mathematical classification tools could be achieved, and applying more comprehensive research further could be implemented in clinic.

Predictors of 30-Day In-Hospital Mortality in Patients Undergoing Urgent Abdominal Surgery Due to Acute Peritonitis Complicated with Sepsis.

BACKGROUND Sepsis is a life-threatening condition with high morbidity and mortality rate. Identifying early prediction factors of critical situations in intra-abdominal sepsis patients can help reduce mortality rates. This prospective study was carried out to evaluate the association of technically available factors with 30-day in-hospital mortality. MATERIAL AND METHODS There were 67 intra-abdominal sepsis patients included in the study; patients were observed for 30 days postoperatively. The data was processed using SPSS24.0 statistical analysis package. All tests that had a significance level of 0.05 were selected. RESULTS Septic shock in association with increase in age per year showed increase the odds of mortality and prognosed 30-days in hospital mortality correctly in 79% of cases. The observed OR was 12.24 (P<0.001). Multiple logistic regression model 2 for the 30-day mortality identified a combination of septic shock, age (≥70 years), time from peritonitis symptoms to surgery prognose mortality with accuracy of 82%. The most accurate model to prognose 30-day in-hospital mortality included the presents of septic shock, age, time from peritonitis symptoms to surgery, drop of MAP <65 mmHg) post-induction, the odds of mortality 8.86 (P=0.001). Severe hypotension post-induction was more frequent in patients who were not diagnosed with sepsis (P=0.035). CONCLUSIONS The present study revealed a simple indicator for the risk for death under diffuse peritonitis patients complicated with sepsis. Septic shock, increase in age per year, peritonitis symptoms lasting more than 30 hours, and severe hypotension post-induction had a negative prognostic value for mortality in patients with intra-abdominal sepsis, and might be a high risk for 30-day mortality.

Mathematical morphology-based imaging of gastrointestinal cancer cell motility and 5-aminolevulinic acid-induced fluorescence.

The aim of this study was the quantitative evaluation of gastrointestinal cancer cell motility and 5-aminolevulinic acid (5-ALA)-induced fluorescence in vitro using mathematical morphology and structural analysis methods. The results of our study showed that MKN28 cells derived from the lymph node have the highest motility compared with AGS or HCT116 cells derived from primary tumors. Regions of single cells were characterized as most moving, and "tightly packed" cell colonies as nearly immobile. We determined the reduction of cell motility in late passage compared to early passage. Application of 5-ALA caused fluorescence in all investigated cells, and the fluorescence was different with regard to the cell type and application time. We observed higher fluorescence in MKN28 cells. Comprehensive image analysis did not reveal any statistically significant difference in fluorescence intensity between "tightly packed" cell regions, where nearly no motility was registered and loosely distributed cells, where the highest cell motility was registered. In conclusions, our study revealed that MKN28 cells derived from the lymph node have higher motility and 5-ALA-induced fluorescence than AGS or HCT116 derived from primary tumors. Moreover, image analysis based on a large amount of processed data is an important tool to study these tumor cell properties.

SCARB1 rs5888 is associated with the risk of age-related macular degeneration susceptibility and an impaired macular area.

BACKGROUND: Age-related macular degeneration (ARMD), a progressive retinal disease, is responsible for an impaired central vision in about 180 million people worldwide. Current options for ARMD prevention and treatment are limited due to an incomplete understanding of disease etiopathogenesis. We aimed to test the hypothesis that the single nucleotide polymorphism rs5888 of SCARB1 gene reflecting lipid and antioxidant micronutrient metabolism pathways is associated with ARMD susceptibility and to evaluate if there is any relation between SCARB1 rs5888 and the macular lesion area. MATERIALS AND METHODS: The prospective case-control study included patients with ARMD (n = 215) and the reference group (n = 238) drawn from a random sample of the Lithuanian population (n = 1436). The genotyping test of SCARB1 rs5888 was carried out using the real-time polymerase chain reaction method. RESULTS: Regression analysis adjusted by gender and age demonstrated that SCARB1 rs5888 TT genotype significantly decreased the odds for ARMD development (OR: 0.61, 95%; CI: 0.380-0.981, p = 0.04). A smoking habit and leading an outdoor life are associated with larger macular lesion areas in ARMD patients (0.54 (0.00-39.06) vs. 3.09 (0.02-19.30) and 0.27 (0.00-34.57) vs. 0.75 (0.00-39.06), respectively). In late stage ARMD subjects with CT genotype, the macular lesion area was larger than in TT carriers (7.64 (0.49-39.06) mm2 vs. 5.02 (0.03-37.06) mm2, p = 0.006). CONCLUSIONS: SCARB1 rs5888 and environmental oxidative stress have a prominent role in ARMD susceptibility, early ARMD progression to advanced stage disease and even in the outcome of the disease-an area of macular lesion.

A method for reconstruction of visually evoked potentials from limited amount of sweeps.

Visually evoked potentials (VEPs) are signals evoked by a visual stimulus. They consist of brief discrete deflections embedded in background electroencephalographic (EEG) activity, which often has larger amplitude. Background EEG cancelation is a major part of VEPs analysis algorithms often realized by coherent averaging or other methods requiring large minimal amount of registered sweeps. In some cases, especially for pediatric patients, or in poor patient compliance cases, long procedure duration and fatigue might cause impaired attention and non-steady target fixation, affecting the quality of recorded VEPs. The possibility to reconstruct VEPs in every single sweep from limited size ensembles opens new diagnostic possibilities and shortens the registration procedure improving its quality. A proposed method is based on truncated expansion (Karhunen-Loève transform) of VEP signals applying generalized universal basis functions (eigenvectors of covariation matrix) calculated from learning set of sweeps, i.e. an ensemble of collected typical recordings. It realizes the possibility to reconstruct a signal from every single sweep even in limited size ensembles of registered sweeps. Application of adaptively time-shifted basis functions enables optimal reconstruction of the signal with latency shift or jitter.

Digital imaging of colon tissue: method for evaluation of inflammation severity by spatial frequency features of the histological images.

BACKGROUND: The efficacy of histological analysis of colon sections used for evaluation of inflammation severity can be improved by means of digital imaging giving quantitative estimates of main diagnostic features. The aim of this study was to reveal most valuable diagnostic features reflecting inflammation severity in colon and elaborate the evaluation method for computer-aided diagnostics. METHODS: Tissue specimens from 24 BALB/c mice and 15 patients were included in the study. Chronic and acute colon inflammation in mice was induced by oral administration of dextran sulphate sodium (DSS) solution, while mice in the control group did not get DSS. Human samples of inflamed colon tissue were obtained from patients with ulcerative colitis (n = 6). Non-inflamed colon tissue of control subjects (n = 9) was obtained from patients with irritable bowel syndrome or functional obstipation. Analysis of morphological changes in mice and human colon mucosa was performed using 4-µm haematoxylin-eosin (HE) sections. The features reflecting morphological changes in the images of colon mucosa were calculated by convolution of Gabor filter bank and array of pixel values. All features were generalized by calculating mean, histogram skewness and entropy of every image response. Principal component analysis was used to construct optimal representation of morphological changes. RESULTS: First principal component (PC1) was representing the major part of features variation (97 % in mice and 71 % in human specimens) and was selected as a measure of inflammation severity. Validation of new measure was performed by means of custom-made software realizing double blind comparison of differences in PC1 with expert's opinion about inflammation severity presented in two compared pictures. Overall accuracy of 80 % for mice and 67 % for human was reached. CONCLUSION: Principal component analysis of spatial frequency features of histological images may provide continuous scale estimation of inflammation severity of colon tissue.

Multistage principal component analysis based method for abdominal ECG decomposition.

Reflection of fetal heart electrical activity is present in registered abdominal ECG signals. However this signal component has noticeably less energy than concurrent signals, especially maternal ECG. Therefore traditionally recommended independent component analysis, fails to separate these two ECG signals. Multistage principal component analysis (PCA) is proposed for step-by-step extraction of abdominal ECG signal components. Truncated representation and subsequent subtraction of cardio cycles of maternal ECG are the first steps. The energy of fetal ECG component then becomes comparable or even exceeds energy of other components in the remaining signal. Second stage PCA concentrates energy of the sought signal in one principal component assuring its maximal amplitude regardless to the orientation of the fetus in multilead recordings. Third stage PCA is performed on signal excerpts representing detected fetal heart beats in aim to perform their truncated representation reconstructing their shape for further analysis. The algorithm was tested with PhysioNet Challenge 2013 signals and signals recorded in the Department of Obstetrics and Gynecology, Lithuanian University of Health Sciences. Results of our method in PhysioNet Challenge 2013 on open data set were: average score: 341.503 bpm(2) and 32.81 ms.

Small Aß1-42 oligomer-induced membrane depolarization of neuronal and microglial cells: role of N-methyl-D-aspartate receptors.

Although it is well documented that soluble beta amyloid (Aß) oligomers are critical factors in the pathogenesis of Alzheimer's disease (AD) by causing synaptic dysfunction and neuronal death, the primary mechanisms by which Aß oligomers trigger neurodegeneration are not entirely understood. We sought to investigate whether toxic small Aß(1-42) oligomers induce changes in plasma membrane potential of cultured neurons and glial cells in rat cerebellar granule cell cultures leading to neuronal death and whether these effects are sensitive to the N-methyl-D-aspartate receptor (NMDA-R) antagonist MK801. We found that small Aß(1-42) oligomers induced rapid, protracted membrane depolarization of both neurons and microglia, whereas there was no change in membrane potential of astrocytes. MK801 did not modulate Aß-induced neuronal depolarization. In contrast, Aß1(-42) oligomer-induced decrease in plasma membrane potential of microglia was prevented by MK801. Small Aß(1-42) oligomers significantly elevated extracellular glutamate and caused neuronal necrosis, and both were prevented by MK801. Also, small Aß(1-42) oligomers decreased resistance of isolated brain mitochondria to calcium-induced opening of mitochondrial permeability transition pore. In conclusion, the results suggest that the primary effect of toxic small Aß oligomers on neurons is rapid, NMDA-R-independent plasma membrane depolarization, which leads to neuronal death. Aß oligomers-induced depolarization of microglial cells is NMDA-R dependent.

Detection and evaluation of ventricular repolarization alternans: an approach to combined ECG, thoracic impedance, and beat-to-beat heart rate variability analysis.

BACKGROUND AND OBJECTIVE: Beat-to-beat alteration in ventricles repolarization reflected by alternans of amplitude and/or shape of ECG S-T,T segment (TWA) is known as phenomena related with risk of severe arrhythmias leading to sudden cardiac death. Technical difficulties have caused limited its usage in clinical diagnostics. Possibilities to register and analyze multimodal signals reflecting heart activity inspired search for new technical solutions. First objective of this study was to test whether thoracic impedance signal and beat-to-beat heart rate reflect repolarization alternans detected as TWA. The second objective was revelation of multimodal signal features more comprehensively representing the phenomena and increasing its prognostic usefulness. MATERIALS AND METHODS: ECG, and thoracic impedance signal recordings made during 24h follow-up of the patients hospitalized in acute phase of myocardial infarction were used for investigation. Signal morphology variations reflecting estimates were obtained by the principal component analysis-based method. Clinical outcomes of patients (survival and/or rehospitalization in 6 and 12 months) were compared to repolarization alternans and heart rate variability estimates. RESULTS: Repolarization alternans detected as TWA was also reflected in estimates of thoracic impedance signal shape and variation in beat-to-beat heart rate. All these parameters showed correlation with clinical outcomes of patients. The strongest significant correlation showed magnitude of alternans in estimates of thoracic impedance signal shape. CONCLUSIONS: The features of ECG, thoracic impedance signal and beat-to-beat variability of heart rate, give comprehensive estimates of repolarization alternans, which correlate, with clinical outcomes of the patients and we recommend using them to improve diagnostic reliability.

Prognosis of in-hospital myocardial infarction course for diabetic and nondiabetic patients using a noninvasive evaluation of hemodynamics and heart rate variability.

BACKGROUND AND OBJECTIVE: The objective of our study was to investigate whether the combination of markers of heart rate variability (HRV) and impedance cardiography (ICG) help evaluate the risk of in-hospital death, ventricular arrhythmia, or complicated course secondary to myocardial infarction (STEMI) and to clarify whether combined analysis of HRV and ICG improve prognosis of STEMI, comparing 3 groups: 1) diabetic, 2) nondiabetic, and 3) diabetes-unselected patients. MATERIAL AND METHODS: The parameters reflecting heart rate variability and central hemodynamics were estimated from a 24-hour synchronic electrocardiogram and thoracic impedance signal recordings in 232 patients (67 diabetic) on the third day after myocardial infarction. Logistic regression analysis was used to determine the predictors of selected outcomes. Different prognostic models were compared with the receiver operating characteristic curve analysis. RESULTS: The model consisting of low- and high-frequency power ratio (LF/HF) and cardiac output (CO) was elaborated for the prognosis of in-hospital death in the group 3 (odds ratios [ORs] were 9.74 and 4.85, respectively). Very low-frequency power (VLF), cardiac index (CIN), and cardiac power output (CPO) were the predictors of ventricular arrhythmia in the group 2 (ORs of 1.005, 5.09, and 66.7, respectively) and the group 3 (ORs of 1.004, 3.84, and 37.04, respectively). The predictors of the complicated in-hospital course in the group 1 were the baseline width of the minimum square difference triangular interpolation of the highest peak of the histogram of all NN intervals (TINN) and stroke volume (SV) (ORs of 1.006, and 1.009, respectively); in the group 2, the mean of the standard deviations of all NN intervals for all 5-minute segments of the recording (SDNN index) and CPO (ORs of 1.06 and 2.44, respectively); and in the group 3, SDNN index, VLF, LF/HF, CIN (ORs of 1.04, 1.004, 2.3, and 3.49, respectively). CONCLUSIONS: The patients with decreased HRV and low estimates of central hemodynamics evaluated by ICG are at an increased risk of the adverse in-hospital course of STEMI. The combined analysis of HRV and ICG hemodynamic estimates contributes to the risk assessment of the complicated in-hospital course of STEMI, in-hospital hemodynamically significant ventricular arrhythmia, and in-hospital death secondary to STEMI. The in-hospital prognostic value of the combined estimates of HRV and ICG is lower in the STEMI patients with diabetes mellitus as compared with the nondiabetic patients.

Impedance cardiography and heart rate variability for long-term cardiovascular outcome prediction after myocardial infarction.

BACKGROUND AND OBJECTIVE: The objective of our study was to evaluate the predictive power of a combined assessment of heart rate variability (HRV) and impedance cardiography (ICG) measures in order to better identify the patients at risk of serious adverse events after ST-segment elevation myocardial infarction (STEMI): all-cause or cardiac mortality (primary outcomes) and in-hospital recurrent ischemia, recurrent nonfatal MI, and need for revascularization (secondary outcomes). MATERIAL AND METHODS: A total of 213 study patients underwent 24-hour electrocardiogram (used for HRV analysis) and thoracic bioimpedance monitoring (used for calculation of hemodynamic measures) immediately after admission. The patients were examined on discharge and contacted after 1 and 5 years. Cox regression analysis was used to determine the predictors of selected outcomes. RESULTS. The standard deviation of all normal-to-normal intervals (SDNN) and cardiac power output (CPO) were found to be the significant determinants of 5-year all-cause mortality (SDNN ≤ 100.42 ms and CPO ≤ 1.43 W vs. others: hazard ratio [HR], 11.1; 95% CI, 4.48-27.51; P<0.001). The standard deviation of the averages of NN intervals (SDANN) and CPO were the significant predictors of 5-year cardiac mortality (SDANN ≤ 85.41 ms and CPO ≤ 1.43 W vs. others: HR, 11.05; 95% CI, 3.75-32.56; P<0.001). None of the ICG measures was significant in predicting any secondary outcome. CONCLUSIONS: The patients with both impaired autonomic heart regulation and systolic function demonstrated by decreased heart rate variability and impedance hemodynamic measures were found to be at greater risk of all-cause and cardiac death within a 5-year period after STEMI. An integrated analysis of electrocardiogram and impedance cardiogram helps estimate patient's risk of adverse outcomes after STEMI.

Monitoring changes in heart tissue temperature and evaluation of graft function after coronary artery bypass grafting surgery.

Thermography is a relatively new contact-free method used in experimental and clinical studies and in cardiovascular surgery to investigate the myocardium and coronary artery function. Objects of complex study included mongrel dogs and patients with coronary artery disease who underwent cardiac surgery. For active dynamic thermography, we used a thermovision camera "A20V" (FLIR Systems, USA). Our data indicate that both experimental and clinical study performed on beating hearts could be an important approach to interoperation inspection of autovenous graft function. An infrared camera also can be successfully used to determine the extent of ischemic damage to the myocardium, heart, and blood vessels during surgery as a significant prognostic tool for evaluating outcome after cardiac operation.

[Hemodynamic studies for prediction of acute myocardial infarction outcomes]. / Hemodinamikos tyrimai uminio miokardo infarkto pasekmems prognozuoti.

Prediction of outcomes after acute myocardial infarction was initiated more than 40 years ago. Improvement of the management options significantly reduced mortality of patients with acute myocardial infarction. In the 1960s, the mortality rate of inpatients was around 25-30%, whereas in 2007, according to the guidelines for the diagnosis and treatment of non-ST-segment elevation acute coronary syndromes, issued by the European Society of Cardiology, hospital mortality in patients with ST-elevation acute myocardial infarction was 7%, while in patients with non-ST-elevation acute coronary syndrome just 5%, but at 6 months, mortality rates were very similar in both conditions (12% vs. 13%, respectively). There are different criteria for prediction of acute myocardial infarction: demographic, clinical, laboratory, instrumental, and epidemiological. Data of hemodynamic studies are ones of the possible criteria for prediction of outcomes after acute myocardial infarction. Methods and findings of hemodynamic studies used for prediction of the outcomes are presented in this article.

Evaluation of ischemic injury of the cardiac tissue by using the principal component analysis of an epicardial electrogram.

Monitoring and control of the heart tissue viability is of crucial importance during heart surgery operations. In most cases the heart tissue suffers from an ischemic injury that causes a decrease in the velocity of electrical excitation propagation in it and influences the shape of the excitation wave front that spreads over the injured area. It is reflected in a more complex shape of the registered epicardial electrogram as compared to normal. A method for quantitative evaluation of the complexity of the shape of the epicardial electrogram based on the principal component analysis is here proposed for evaluation of the ischemic injury of the cardiac tissue. A minimal, yet sufficient, number of the principal components (the optimal basis functions) for truncated expansion of the epicardial electrogram signals could be used as an estimate of signal complexity. The method for determination of such a minimal, yet sufficient, number of principal components were developed by using epicardial electrograms registered during in situ experiments on dogs in which local ischemia was evoked by ligation of a coronary vessel.